COD

BOD

Color
Boron

Cobalt
Acidity

Barium

Copper
Arsenic

Cyanide
Calcium

Fluoride
Bacteria

Chlorine
Chloride
Bromine

Chelants
Alkalinity

Cadmium

Chromate
Acid/Base

Aluminum

Detergents
Conductivity

Cyanuric Acid
Ascorbic Acid

Formaldehyde
Carbon Dioxide

Erythorbic Acid
Chlorine Dioxide

Chromium (Total)

Dissolved Oxygen
Section 1

Chromium (Hexavalent)
1.1 Applications guide

Agriculture
Aquaculture
Aquarium Testing
Beverages/Bottled Water
Boiler/Cooling Water
Chemical Manufacture
Chlorine Production
Commercial Laundries
Drinking Water
Education
Water Analysis Guide

Environmental Testing
Food/Feed Industry
Metals/Mining, Mfg, Finishing
Petroleum Industry
Pharmaceutical Manufacture
Pools, Spas
Power Plant Utilities
Pulp, Paper Mills
Semiconductor Manufacture
Solid Waste/Sludge
Textile Industry
Ultrapure Water
Wastewater, Industrial
Wastewater, Municipal
Water Conditioning

5
6
pH
PCB
Lead

Nickel
Iodine
Iodide

Ozone
Glycols

Phenols
Mercury
Hardness
Hydrazine

Phosphate
Iron (Total)

Manganese

Molybdenum
Iron (Ferrous)

Permanganate
Oil and Grease
Nitrogen (TKN)
Gluteraldehyde

Nitrogen (Total)

Nitrogen (Nitrite)
Nitrogen (Nitrate)
Hydrogen Sulfide

Nitrogen Ammonia

Oxygen Scavenger
Hydrogen Peroxide

Nitrogen (Inorganic)

Nitrogen (Monochloramine)
Water Analysis Guide

Agriculture
Aquaculture
Aquarium Testing
Beverages/Bottled Water
Boiler/Cooling Water
Chemical Manufacture
Chlorine Production
Commercial Laundries
Drinking Water
Education
Environmental Testing
Food/Feed Industry
Metals/Mining, Mfg, Finishing
Petroleum Industry
Pharmaceutical Manufacture
Pools, Spas
Power Plant Utilities
Pulp, Paper Mills
Semiconductor Manufacture
Solid Waste/Sludge
Textile Industry
Ultrapure Water
Wastewater, Industrial
Wastewater, Municipal
Water Conditioning
 TPH
TDS

Zinc
QAC

Silica
Silver

Sulfite
Tannin
Sulfide
Sulfate
Salinity

Sodium

Toxicity

Triazole
Turbidity
Selenium
Potassium

Water in Oil
Phosphorus

Volatile Acids
Phosphonates

Sodium Chromate
Sodium Hydroxide
Agriculture
Aquaculture
Aquarium Testing
Beverages/Bottled Water
Boiler/Cooling Water
Chemical Manufacture
Chlorine Production
Commercial Laundries
Drinking Water
Education
Environmental Testing
Food/Feed Industry
Metals/Mining, Mfg, Finishing
Petroleum Industry
Pharmaceutical Manufacture
Pools, Spas
Power Plant Utilities
Pulp, Paper Mills
Semiconductor Manufacture
Solid Waste/Sludge
Textile Industry
Ultrapure Water
Wastewater, Industrial
Wastewater, Municipal
Water Conditioning

7
Water Analysis Guide
Water Analysis Guide

1.2 Abbreviations and conversions

1.2.1 Procedure abbreviations
Table 1 shows common abbreviations used in written chemical procedures.
Table 1 Abbreviations
Abbreviation Definition Abbreviation Definition
°C degree(s) Celsius (Centigrade) L liter—volume equal to one cubic
decimeter (dm3)
°F degree(s) Fahrenheit LR low range
ACS American Chemical Society MDL method detection limit
reagent grade purity
APHA Standard Standard Methods for the MDB marked dropper bottle
Methods Examination of Water and
mg/L milligrams per liter (ppm)
Wastewater, published jointly by
the American Public Health µg/L micrograms per liter (ppb)
Association (APHA), the American
Water Works Association mL milliliter—1/1000 of a liter. It is
(AWWA) and the Water approximately the same as a
Environment Federation (WEF), is cubic centimeter (and is
the standard reference work for sometimes called a “cc”).
water analysis. Many procedures MR medium range
contained in this manual are
based on Standard Methods. NIPDWR National Interim Primary Drinking
Water Regulations
®
AV AccuVac NPDES National Pollutant Discharge
Elimination System
Bicn bicinchoninate P phosphorus
conc concentrated PCB poly chlorinated biphenyl
DB dropping bottle ppb parts per billion
DBP disinfection by-products ppm parts per million
™
CFR Code of Federal Regulations RL Rapid Liquid
EDL Estimated detection limit SCDB self-contained dropping bottle
EPA Environmental Protection Agency THM trihalomethane
™
F&T free and total TNT Test ‘N Tube
®
FM FerroMo TOC total organic carbon
®
FV FerroVer TPH total petroleum hydrocarbons
®
FZ FerroZine TPTZ 2,4,6-Tri-(2-Pyridyl)-1,3,5-Triazine
g grams USEPA United States Environmental
Protection Agency
gr/gal grains per gallon (1 gr/gal = ULR ultra low range
17.12 mg/L)
HR high range

8
 Water Analysis Guide

1.2.2 Conversions
1.2.2.1 Chemical species
Table 2 shows species conversion factors for many commonly used chemicals.
Table 2 Conversion factors
To convert from... To... Multiply by...
mg/L Al mg/L Al2O3 1.8895
mg/L B mg/L H3BO3 5.7
mg/L Ca-CaCO3 mg/L Ca2+ 0.4004
mg/L CaCO3 mg/L Ca2+ 0.4004
mg/L CaCO3 mg/L Mg2+ 0.2428
µg/L Carbohydrazide µg/L Hydroquinone 1.92
µg/L Carbohydrazide µg/L ISA 2.69
µg/L Carbohydrazide µg/L MEKO 3.15
mg/L Cr6+ mg/L CrO42– 2.231
mg/L Cr6+ mg/L Na2CrO4 3.115
mg/L Cr6+ mg/L Cr2O7 2– 2.077
mg/L Mg-CaCO3 mg/L Mg2+ 0.2428
mg/L Mn mg/L KMnO4 2.876
mg/L Mn mg/L MnO4 – 2.165
mg/L Mo6+ mg/L MoO42– 1.667
mg/L Mo6+ mg/L Na2MoO4 2.146
mg/L N mg/L NH3 1.216
mg/L N mg/L NO3– 4.427
mg/L Cl2 mg/L NH2Cl 0.726
mg/L Cl2 mg/L N 0.197
mg/L NH3-N mg/L NH3 1.216
mg/L NH3-N mg/L NH4 + 1.288
mg/L NO2- mg/L NaNO2 1.5
mg/L NO2 - mg/L NO2 ––N 0.3045
mg/L NO2--N mg/L NaNO2 4.926
µg/L NO2--N µg/L NaNO2 4.926
mg/L NO2 --N mg/L NO2 – 3.284
µg/L NO2--N µg/L NO2– 3.284
mg/L NO3--N mg/L NO3– 4.427
mg/L PO4 3- mg/L P 0.3261
µg/L PO43- µg/L P 0.3261
mg/L PO4 3- mg/L P2O5 0.7473
µg/L PO43- µg/L P2O5 0.7473

9
Water Analysis Guide

Table 2 Conversion factors (continued)

To convert from... To... Multiply by...
mg/L SiO2 mg/L Si 0.4674
µg/L SiO2 µg/L Si 0.4674

1.2.2.2 Hardness conversion

Table 3 shows the factors to convert hardness from one unit of measure to another. For
example, to convert mg/L CaCO3 to German parts/100,000 CaO, multiply the value in
mg/L x 0.056.
Note: meq/L = N × 1000
Table 3 Hardness conversion factors
Units of measure mg/L British American French parts/ German parts/ meq/L g/L CaO lb/ft3
CaCO3 gr/gal gr/gal 100,000 CaCO3 100,000 CaCO3 1 CaCO3
(Imperial) (US)
CaCO3 CaCO3
mg/L CaCO3 1.0 0.07 0.058 0.1 0.056 0.02 5.6x10–4 6.23x10–5
English gr/gal 14.3 1.0 0.83 1.43 0.83 0.286 8.0x10–3 8.9x10–4
CaCO3
US gr/gal CaCO3 17.1 1.2 1.0 1.72 0.96 0.343 9.66x10–3 1.07x10–3
French 10.0 0.7 0.58 1.0 0.56 0.2 5.6x10–3 6.23x10–4
p/100,000 CaCO3
German 17.9 1.25 1.04 1.79 1.0 0.358 1x10–2 1.12x10–3
p/100,000 CaO
meq/L 50.0 3.5 2.9 5.0 2.8 1.0 2.8x10–2 3.11x10–2
g/L CaO 1790.0 125.0 104.2 179.0 100.0 35.8 1.0 0.112
lb/ft3 CaCO3 16,100.0 1123.0 935.0 1610.0 900.0 321.0 9.0 1.0
1 epm/L or mval/L

1.3 Laboratory practices

1.3.1 Temperature
Most methods are completed accurately when the sample temperature is between 20 and
25 °C (68 to 77 °F). A note in the individual procedure shows any special temperature
requirements.
1.3.2 Mixing
When reagent is added to a graduated cylinder or titration flask, swirl the sample gently.
A gentle swirl motion decreases the risk of atmospheric contamination in carbon dioxide
and other tests for gases.
1. Hold the cylinder (or flask) firmly with the tips of the thumb and first two fingers
(Figure 1).
2. Hold the cylinder at a 45-degree angle and make a circular motion from the wrist.
3. Move the cylinder in approximately 305-mm (12-in.) circles. Make enough rotation to
complete the mixing in a few turns.

10
 Water Analysis Guide

Figure 1 Swirl a cylinder and invert a sample cell

To mix a sample in a closed sample cell or a mixing cylinder:

1. Hold the cell or cylinder, in a vertical position with the cap on top.
2. Invert so that the cap is on the bottom. Return the cell to its original position
(Figure 1). Repeat as needed.
To mix a sample in a square sample cell:
1. Hold the neck of the cell with the thumb and index finger of one hand. Put the
concave bottom of the cell on the tip of the index finger of the other hand.
2. Rotate the cell quickly one way and then in the reverse direction to mix (Figure 2).

Figure 2 Rotate a sample cell

1.3.3 Digestion
Several procedures use sample digestion. Digestion uses chemicals and heat to break
down a substance into components that can be analyzed. This section briefly describes
three different digestion procedures.
The Digesdahl system gives a digested substance applicable for the determination of
metals, total phosphorus and total Kjeldahl nitrogen (TKN). It is fast and is very effective
at destroying interfering organic materials.
For USEPA reporting purposes, USEPA-approved digestions are necessary. USEPA
presents two digestions (mild and vigorous) for metals analysis. Other digestion
procedures are necessary for mercury, arsenic, phosphorus and TKN.
Refer to Sample pretreatment by digestion on page 35 for more information on sample
digestion.

11
Water Analysis Guide

1.3.4 Distillation
Distillation is an effective and safe method used to separate some chemical components
for analysis. The equipment that follows is recommended for distillation:
• General Purpose Distillation Apparatus (22653-00), shown in Figure 3
• Arsenic Distillation Apparatus Set (22654-00)
• Cyanide Distillation Apparatus Set (22658-00)
• General Purpose Heater and Support Apparatus (22744-00, 115 VAC, 60 Hz)
• General Purpose Heater and Support Apparatus (22744-02, 230 VAC, 50 Hz)
The Distillation Apparatus is applicable for water and wastewater that use sample
pretreatment by distillation. Applications for the General Purpose Apparatus include:
fluoride, albuminoid nitrogen, ammonia nitrogen, phenols, selenium and volatile acids.
The General Purpose Heater and Support Apparatus gives efficient heating and
anchoring of the glassware.

Figure 3 General purpose distillation apparatus

1.3.5 Filtration
Filtration separates particulates from an aqueous sample. Filtration uses a porous
medium that keeps particulates but lets liquids pass through. Filtration removes turbidity
from water samples. Turbidity can interfere in colorimetric analyses.
The two filtration methods most frequently used are vacuum and gravity filtration.

1.3.5.1 Vacuum filtration

Vacuum filtration uses both suction and gravity to pull the liquid through the filter. An
aspirator or vacuum pump is used to make suction (Figure 4). Vacuum filtration is faster
than gravity filtration alone.

12
 Water Analysis Guide

To filter with a vacuum:

1. Use tweezers to put a filter paper into the filter holder.

2. Put the filter holder assembly in the filtering flask.
3. Dampen a filter paper with deionized water to make sure that there is adhesion to the
holder.
4. Put the funnel housing on the filter holder assembly.
5. While a vacuum is applied to the filtering flask, transfer the sample to the filtering
apparatus.
6. When the filtration is complete, slowly release the vacuum from the filtering flask and
transfer the solution from the filter flask to another container.

Figure 4 Vacuum filtration

1.3.5.2 Necessary apparatus for vacuum filtration

Description Unit Item no.
Filter discs, glass fiber, 47-mm 100/pkg 253000
Filter holder, membrane, 47-mm each 1352900
Flask, filtering, 500-mL each 54649
Select one of the following:
Pump, vacuum, hand operated each 1428300
Pump, vacuum, portable, 115 VAC each 2824800
Pump, vacuum, portable, 230 VAC each 2824801
Tubing, vacuum — 2074145
Tweezers each 1428200

1.3.5.3 Gravity filtration

Many chemical procedures use gravity filtration with the items in Table 4. Gravity filtration
is better for fine particles (Figure 5). The rate of filtration increases as the volume
increases in the filter cone, but do not fill the cone more than three-quarters full.
Note: Pretreatment with acid and heat is often necessary for metal tests. Because filter paper does
not withstand acid and heat pretreatment, use a glass filter disc in the vacuum filtration. Glass filter
discs do not keep color species like the paper filters.
To filter with gravity:

1. Put a folded filter paper into the funnel.

2. Dampen the filter paper with deionized water so that it bonds to the funnel.

13
Water Analysis Guide

3. Put the funnel into an Erlenmeyer flask or graduated cylinder.

4. Pour the sample into the funnel.
Table 4 Necessary apparatus for gravity filtration
Description Unit Item no.
Cylinder, graduated, 100-mL each 50842
Funnel, poly, 65-mm each 108367
Filter paper, 12.5-cm, pleated 100/pkg 189457
Flask, Erlenmeyer, 125-mL each 50543

Figure 5 Gravity filtration

1.3.6 Reagents
1.3.6.1 Reagent and standard stability
In general, reagents and standards have the maximum shelf life when they are put in a
location that is cool, dark and dry. The product label gives any special storage needs.
It is always good laboratory practice to put the date on chemicals upon receipt and to
move supplies so that the older supplies are used first. When the reagent shelf life is
unknown or in doubt, use a standard to measure reagent effectiveness.
Absorption of moisture, carbon dioxide or other gases from the atmosphere, bacterial
action, high temperatures or light (with photosensitive compounds) may affect the reagent
shelf life. In some cases, reaction with the storage container or interaction of reagent
components may occur.

1.3.6.2 Reagent blank

In several tests, the contribution of the reagent(s) to the final reading is of such a
magnitude that it must be compensated for whenever the test is completed. Reagent
blank refers to that portion of the test result contributed solely by the reagent. This makes
a positive error in the test results.
Reagents are made with the lowest possible blank. For most reagents, it is less than
0.009 absorbance units. However, it is sometimes impossible or impractical to make
reagents with such a low blank. When such reagents are used, it is best to find the
reagent blank with the procedure that uses high-quality water (deionized, distilled, etc.) in
place of sample to “zero” the instrument. The resulting value is then shown in the
concentration units of the test and is subtracted from each sample determination that
uses the same reagent lot. Spectrophotometer and colorimeter software lets the reagent

14
 Water Analysis Guide

blank value be kept and subtracted automatically from each sample value. The reagent
blank needs to be found only at first use, when a new lot of reagent has been opened or if
contamination is suspected.
In most tests, the reagent blank is so small the instrument may be set to zero on either an
untreated portion of the original water sample or on deionized water. This will not result in
a significant loss of accuracy unless the test is for very low levels of the species of
interest. When a test is for very low levels of the species, it is best to use a reagent blank
prepared as above.
1.3.7 Sample dilution
Most colorimetric tests use volumes of 10 and 25 mL. However, in some tests, the color
developed in the sample may be too intense to be measured due to high levels of analyte
or unexpected colors may develop due to an interference. In one or the other case, dilute
the sample to make a measurable endpoint or to find out if interfering substances are
present.
To dilute the sample:

1. Use a pipet to add the selected sample portion to a clean graduated cylinder (or
volumetric flask for more accurate work).
2. Fill the cylinder (or flask) to the necessary volume with deionized water.
3. Mix well. Use the diluted sample to complete the test.
Table 5 shows the relative quantities and multiplication factors to use with a 25-mL
graduated cylinder. The concentration of the sample is equal to the diluted sample
result multiplied by the multiplication factor.
Note: For sample sizes of 10 mL or less, use a pipet to measure and add the sample to the
graduated cylinder or volumetric flask.
Table 5 Sample dilution volumes
Sample volume (mL) mL of deionized water used to bring the volume to 25 mL Multiplication factor
25.0 0.0 1
12.5 12.5 2
10.0 15.0 2.5
5.0 20.0 5
2.5 22.5 10
1.0 24.0 25
0.250 24.75 100

More accurate dilutions can be made with a pipet and a 100-mL volumetric flask
(Table 6).
1. Use a pipet to add the sample. Dilute to volume with deionized water.
2. Put in the stopper and invert to mix.
Table 6 Multiplication factors for dilution to 100 mL
Sample volume (mL) Multiplication factor
1 100
2 50
5 20
10 10
25 4
50 2

15
Water Analysis Guide

1.3.7.1 Sample dilution with interfering substances

Sample dilution may affect the level at which a substance interferes. The effect of the
interferences decreases as the dilution increases. In other words, higher levels of an
interfering substance can be tolerated in the original sample if it is diluted before analysis.
Example:
Copper does not interfere at or below 100 mg/L for a 25-mL sample in a procedure. If the
sample volume is diluted with an equal volume of water, what is the level at which copper
will not interfere?
Total volume ÷ Sample volume = Dilution factor
25 ÷ 12.5 = 2
Interference level × Dilution factor = Interference level in sample
100 × 2 = 200
The level at which copper will not interfere in the diluted sample is at or below 200 mg/L.
®
1.3.8 AccuVac Ampuls

CAUTION
Personal injury hazard. Glass ampules have sharp edges after they are opened. Use personal
protective equipment to work with glass ampules.

AccuVac Ampuls contain pre-measured powder or liquid vacuum-packed in optical-quality

glass ampules.
To use AccuVac Ampuls:

1. Collect the sample in a beaker or other open container.

2. Use one of the methods that follow to break the tip off the ampule:
• Use the optional AccuVac Snapper (2405200). Refer to Use the AccuVac
Snapper on page 16 for instructions.
• Put the ampule tip well below the sample surface and break the tip off against the
beaker wall (Figure 6). The break must be far enough below the surface that air
does not come in as the level of the sample drops.
3. Secure an ampule cap over the tip of the ampule. Invert the ampule several times to
dissolve the reagent. The cap protects from broken glass and supplies a grip to insert
and remove the ampul from the cell holder. Wipe the ampule with a lint-free cloth to
remove fingerprints.
Note: Without the cap, the liquid stays in the ampule when the ampule is inverted.
4. Insert the ampule into the sample cell holder and read the results directly.
Figure 6 Use the AccuVac Ampuls

1.3.8.1 Use the AccuVac Snapper

1. Hold the snapper with the open end up.

2. Gently slip the ampule into the snapper, point first, until the tip touches the ramp at
the bottom of the snapper.
3. Hold the snapper between the index and middle finger (like a syringe). With the
ampule tip down, lower the snapper into the sample until the ampule shoulder is wet.

16
 Water Analysis Guide

4. Push on the flat end of ampule with the thumb (as if depressing the plunger on a
syringe) until the tip snaps (Figure 7). Let the ampule fill before the sample is
removed.
5. Rinse the wet end of the snapper and ampule with clean water, if necessary. Remove
the ampule from the snapper.
6. Discard the ampule tip (kept in the snapper) in an applicable waste receptacle.
Figure 7 Use the AccuVac Snapper

®
1.3.9 PermaChem pillows
PermaChem pillows use powdered reagents to minimize deterioration and the risk of
reagent spills (Figure 8). Hold the pillow away from the face as it is opened.
Figure 8 Open the PermaChem pillows

1.3.10 Sample cells

A set of sample cells are shipped with each photometric instrument. The same solution in
both cells gives the same absorbance (within ±0.002 Abs for properly matched cells). For
more information, refer to Match the sample cells on page 18.
For accurate results, use only the sample cells specified in each procedure. Due to
differences in cell path lengths, sample cell substitution introduces bias in test results. For
example, 25.4-mm (1-inch) square cells have a path length approximately 8% longer than
25.4-mm (1-inch) round cells. Substitution of round cells for square cells introduces a bias
in the reading.

1.3.10.1 Orientation of the sample cells

To minimize measurement variability when a particular cell is used, always orient the cell
in the same manner before it is put into the cell holder. The fill marks on the cells can be
used as orientation guides to position the cells.

1.3.10.2 Maintain the sample cells

Keep the sample cells in the supplied boxes to protect them from scratches and
breakage. After use, empty and clean the sample cells. Do not leave color solutions in the
sample cells for extended periods of time.

17
Water Analysis Guide

1.3.10.3 Clean the sample cells

CAUTION
Chemical exposure hazard. Obey laboratory safety procedures and wear all of the
personal protective equipment appropriate to the chemicals that are handled. Refer to the
current material safety data sheets (MSDS) for safety protocols.

CAUTION
Chemical exposure hazard. Dispose of chemicals and wastes in accordance with local,
regional and national regulations.

Most laboratory detergents are used at recommended concentrations. Neutral detergents,

such as Liquinox, are safer to use when regular cleaning is necessary. To decrease the
cleaning times, increase the temperature or use an ultrasonic bath. To complete the
cleaning, rinse a few times with deionized water and then let the sample cell air dry.
Sample cells may also be cleaned with acid, followed by a thorough rinse with deionized
water.
Note: Always use acid to clean sample cells that were used for low-level metal tests.
Special cleaning methods are necessary for individual procedures. When a brush is used
to clean sample cells, take extra care to avoid scratches on the interior surfaces of the
sample cells.

1.3.10.4 Match the sample cells

The sample cells supplied with the spectrophotometer instrument are distortion-free.
Nicks and scratches from movement may cause an optical mismatch between two
sample cells and introduce error into the test results. To prevent this type of error,
optically match the sample cells.
Note: Refer to the spectrophotometer user manual for the specific steps necessary to select
wavelengths and set the instrument to zero.

1. Set the instrument power switch to on. Make sure that the Display Lock is off or the
Reading mode is set to Continuous.
2. Select a wavelength of 510 nm or the wavelength to be used for the test.
3. Pour at least 10 mL (25 mL for 25-mL cells) of deionized water into each of the two
sample cells.
4. Put one sample cell into the cell holder with the fill mark toward the user.
5. Set the instrument to zero.
6. Put the other sample cell into the cell holder with the fill line toward the user.
7. Let the value stabilize and then read the absorbance. Record the resulting
absorbance.
8. Turn the cell 180° and do step 6 again. Try to get an absorbance value within
±0.002 Abs of the first cell. Record the orientation of the cell.
If the sample cells cannot be matched to within ±0.002 Abs, they can still be used if
an adjustment is made for the difference. For example, if the second cell reads
0.003 absorbance units higher than the first cell, adjust future readings (when these
two cells are used). Subtract 0.003 absorbance units (or the equivalent concentration)
from the reading. Likewise, if the second cell reads –0.003 absorbance units, add
0.003 absorbance units to the reading.

18
 Water Analysis Guide

1.3.11 Other apparatus

1.3.11.1 Boiling aids
Boiling is necessary for some procedures. Under some conditions, bumping may occur
and cause sample loss or injury. Bumping is caused by the sudden, almost explosive,
conversion of water to steam as it is heated. Use of a boiling aid, such as boiling chips
(1483531), decreases bumping.
Make sure that the boiling aids do not contaminate the sample. Do not use boiling aids
(except glass beads, 259600) more than once. Use a large sufficient flask or beaker to
give significant head space above the solution. Loosely cover the sample during boiling to
prevent splash, reduce the chance of contamination and minimize sample loss.
Individual procedures recommend the specific boiling aid to use.
1.3.12 Achieve accuracy in measurement
1.3.12.1 Pipets and graduated cylinders

CAUTION
Chemical exposure hazard. The top of the pipet is open. Always use a pipet filler bulb to pull the
liquid into the pipet.

When smaller sample quantities are used, the accuracy of measurements becomes
increasingly important. Figure 9 shows the correct way to read the sample level with the
meniscus formed when the liquid wets the graduated cylinder or pipet walls.
Before use, rinse the pipet or cylinder two or three times with the sample to be tested.
Use a pipet filler or pipet bulb to pull the sample into the pipet. When a pipet is filled, keep
the tip of the pipet below the surface of the sample as the sample is pulled into the pipet.
Serological pipets have marks that show the volume of liquid delivered by the pipet. The
marks may extend to the tip of the pipet or may be only on the straight portion of the tube.
If the marks are only on the straight part of the tube:
1. Fill the serological pipets to the zero mark.
2. To discharge the sample, drain the sample until the meniscus is level with the
necessary mark.
If the serological pipet has marks that extend to the tip of the pipet:
1. Fill the pipet to the applicable volume.
2. Drain all of the sample from the pipet.
3. For accurate measurements, use a pipet filler to blow the sample out of the pipet tip.
Volumetric (transfer) pipets have a bulb in the middle and a single ring above the bulb to
show the volume of liquid when it is filled to the mark. To discharge a volumetric pipet,
hold the tip of the pipet at a slight angle against the container wall and drain. Do not
discharge the solution still in the tip of the pipet after it is drained. Volumetric pipets are
made to keep a small amount of sample in the pipet tip.

19
Water Analysis Guide

If droplets of the sample hold to the walls of the pipet, the pipet is dirty and will not supply
the correct amount of sample. Fully clean the pipet with a laboratory detergent or cleaning
solution and then rinse several times with deionized water.

Figure 9 Read the meniscus

1.3.12.2 Pour-Thru™ Cell

The Pour-Thru Cell is an optional accessory that increases accuracy and makes
measurements more convenient for the rapid liquid methods. Methods that use 25-mL
samples and sample cells can use the Pour-Thru Cell if specified in the procedure. The
Pour-Thru Cell cannot be used with 10-mL sample sizes and reagents. The Pour-Thru
Cell cannot be used directly with a method unless it is specified in the procedure. For
more information, refer to the photometer user manual.
Refer to the photometer user manual for installation and operation instructions.
• Pour the solution into the funnel of the installed Pour-Thru Cell Module. Do not spill
solution on the instrument.
• The funnel height and orientation may be adjusted. The funnel height increases the
speed of the sample flow through the cell. The higher the funnel, the faster the flow.
• To minimize air bubbles, adjust the funnel so that it drains fully with the final level of
liquid in the tube about 5 cm (2 inches) below the tip of the funnel.
• Take instrument readings after the solution has stopped flowing through the cell.
• Always rinse the cell thoroughly with deionized water after each series of tests or as
often as specified in the procedure.
Occasionally, remove the Pour-Thru Cell to look for any accumulation of film on the
windows. If the windows are not clear (have a film), soak the cell in a detergent bath and
rinse thoroughly with deionized water.

1.4 Chemical analysis

1.4.1 Sample collection preservation and storage
Correct sampling and storage are critical for accurate testing. Sampling devices and
containers must be thoroughly cleaned to prevent carryover from previous samples.
Preserve the sample with the test-specific information about sample preservation.

1.4.1.1 Collect water samples

Use a clean container. Rinse the container several times with the water to be sampled,
and then take the sample. Document the location and procedure used for each sample
taken. For example:
From a tap—Take samples as close as possible to the source of the supply. This
decreases the influence of the distribution system on the sample. Make sure that there is
sufficient water to flush the system. Fill sample containers slowly with a gentle stream to
avoid turbulence and air bubbles.

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 Water Analysis Guide

From a well—Let the pump run long enough to pull fresh groundwater into the system.
Collect a sample from a tap near the well.
From open waters—Take the sample as near the middle of the body of water as is
practical, at least several feet from the shore or edge of the tank. Take the sample under
the surface of the water. When a capped container is used, submerge it before the cap is
removed.
1.4.1.1.1 Types of containers
Different containers are recommended for specific parameters.
• Polypropylene and Polyethylene
®
• Quartz or TFE (tetrafluoroethylene, Teflon )—higher quality and price
• Glass—Glass supplies a good general-purpose container. Do not use soft-glass
containers to collect samples to be tested for metals in the µg/L range.
To find silver, put samples in dark containers such as amber or brown glass.
Acid wash the sample containers to fully clean them before use.
1.4.1.1.2 Acid washing
If a procedure suggests acid washing, do the steps that follow:
1. Clean the glassware or plasticware with laboratory detergent. Phosphate-free
detergent is best. To find phosphates, always use phosphate-free detergent.
2. Rinse well with tap water.
3. Rinse with a 1:1 hydrochloric acid solution or a 1:1 nitric acid solution. To test for lead
or other metals, nitric acid is best.
4. Rinse well with deionized water. For chromium, 12–15 rinses may be necessary. To
test for ammonia and Kjeldahl nitrogen, make sure that the rinse water is ammonia-
free.
5. Air dry the container. Protect the glassware from fumes and other sources of
contamination during storage.
Use chromic acid or chromium-free substitutes to remove organic deposits from glass
containers. Afterward, rinse thoroughly with water to remove all traces of chromium.
Do not use metal contaminants from containers, distilled water or membrane filters.
1.4.1.1.3 Sample splits
Samples must often divided into separate containers for intra- or inter-laboratory use in
studies, confirmation, alternative techniques or to keep additional sample for reference
and stability studies.
It is very important that sample be divided done correctly:
• Collect a large volume of sample in a single container and transfer to smaller
containers. Do not fill the smaller containers individually from the water source.
• Fully mix samples that contain particulates or solids before they are divided so that all
the samples are homogeneous.
• If it is necessary to filter the sample before analysis or storage, filter all the sample
before it is divided.
• Use the same kind of container for all the samples.
• Analyze biologically active splits on the same day or as close to the same day as is
possible.
• Preserve all splits in the same way. If this is not done, fully record the differing
methods.
• When the sample is to be tested for volatile contaminants, fill containers so that they
overflow and then put on a cap carefully. Do not leave any head space or air in the
container.

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Water Analysis Guide

1.4.1.2 Storage and preservation

Because chemical and biological processes continue after collection, analyze the sample
as soon as possible. This also reduces the chance for error and minimizes labor. When
an immediate analysis is not possible, preserve the sample. Preservation methods
include pH control, chemical addition, refrigeration and freezing.
Comparison of international drinking water and FDA bottled water guidelines gives an
overview of preservation methods and holding times for specific procedures.
Preserve aluminum, cadmium, chromium, cobalt, copper, iron, lead, nickel, potassium,
silver and zinc samples for at least 24 hours with the steps that follow.
1. Add approximately 2.5 mL Nitric Acid 1:1 solution (254049) per 1 L of sample until a
pH less than 2 is reached.
2. Use pH indicator paper or a pH meter to make sure that the pH is 2 or less. Add
additional pillows if necessary.
3. Adjust the sample pH before analysis. Increase the pH to 4.5 with Sodium Hydroxide
Standard Solution, 1 N or 5 N.
1.4.1.2.1 Sample preservation
Comparison of international drinking water and FDA bottled water guidelines gives an
overview of preservation methods and holding times* for specific procedures. Refer to
Table 7.
Table 7 Necessary containers, preservation techniques and holding times
Parameter name Container1 Preservation2,3 Maximum holding
time4
Bacterial tests
Coliform, fecal and total P, G Cool, 4 °C to less than 10 °C, 0.0008% 6 hours
Na2S2O3
Fecal streptococci P, G Cool, 4 °C to less than 10 °C, 0.0008% 6 hours
Na2S2O3
Aquatic toxicity tests
Toxicity, acute and chronic P, G Cool, 4 ≤ 6 °C 36 hours
Chemical tests
Acidity P, G Cool, 4 ≤ 6 °C 14 days
Alkalinity P, G Cool, 4 ≤ 6 °C 14 days
Ammonia P, G Cool, 4 ≤ 6 °C H2SO4 to pH less than 2 28 days
Biochemical oxygen demand P, G Cool, 4 ≤ 6 °C 48 hours
(BOD)
Biochemical oxygen P, G Cool, 4 ≤ 6 °C 48 hours
demand, carbonaceous
(CBOD)
Boron P, PFTE or quartz HNO3 to pH less than 2 6 months
Bromide P, G Not necessary 28 days
Chemical oxygen demand P, G Cool, 4 ≤ 6 °C, H2SO4 to pH less than 2 28 days
(COD)
Chloride P, G Not necessary 28 days
Chlorine, total residual P, G Not necessary Analyze immediately

* This table was adapted from Table II in the Code of Federal Regulations, Vol 77, No. 97/Friday, May 18,
2012/Rules and Regulations, pages 29806–29809. Most organic tests are not included.

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 Water Analysis Guide

Table 7 Necessary containers, preservation techniques and holding times (continued)

Parameter name Container1 Preservation2,3 Maximum holding
time4
Color P, G Cool, 4 ≤ 6 °C 48 hours
Cyanide, total and amenable P, G Cool, 4 ≤ 6 °C, NaOH to pH higher than 14 days6
to chlorination 12, 0.6 g ascorbic acid5
Fluoride P Not necessary 28 days
Hardness P, G HNO3 to pH less than 2, H2SO4 to pH less 6 months
than 2
Hydrogen ion (pH) P, G Not necessary Analyze immediately
Kjeldahl and organic nitrogen P, G Cool, 4 ≤ 6 °C, H2SO4 to pH less than 2 28 days
Metals7
Chromium VI P, G Cool, 4 ≤ 6 °C, (NH4)2SO4 buffer to pH 28 days8
9.3 – 9.7
Mercury P, G HNO3 to pH less than 2 28 days
Metals, except boron, P, G HNO3 to pH less than 2 6 months
chromium VI and mercury
Nitrate P, G Cool, 4 ≤ 6 °C 48 hours
Nitrate-nitrite P, G Cool, 4 ≤ 6 °C, H2SO4 to pH less than 2 28 days
Nitrite P, G Cool, 4 ≤ 6 °C 48 hours
Oil and grease G Cool, 4 ≤ 6 °C, HCl or H2SO4 to pH less 28 days
than 2
Organic Carbon P, G Cool, 4 ≤ 6 °C, HCl or H2SO4 or H3PO4 to 28 days
pH less than 2
Orthophosphate P, G Filter immediately; Cool, 4 ≤ 6 °C 48 hours
Oxygen, dissolved probe G Bottle and top Not necessary Analyze immediately
Winkler G Bottle and top Fix on site and store in dark 8 hours
48. Phenols G only Cool, 4 ≤ 6 °C, H2SO4 to pH less than 2 28 days
Phosphorus, elemental G Cool, 4 ≤ 6 °C 48 hours
Phosphorus, total P, G Cool, 4 ≤ 6 °C, H2SO4 to pH less than 2 28 days
Residue, Total P, G Cool, 4 ≤ 6 °C 7 days
Residue, Filterable P, G Cool, 4 ≤ 6 °C 7 days
Residue, Nonfilterable (TSS) P, G Cool, 4 ≤ 6 °C 7 days
Residue, Settleable P, G Cool, 4 ≤ 6 °C 48 hours
Residue, Volatile P, G Cool, 4 ≤ 6 °C 7 days
Silica P, PFTE or quartz Cool, 4 ≤ 6 °C 28 days
Specific Conductance P, G Cool, 4 ≤ 6 °C 28 days
Sulfate P, G Cool, 4 ≤ 6 °C 28 days
Sulfide P, G Cool, 4 ≤ 6 °C, add zinc acetate plus 7 days
sodium hydroxide to pH higher than 9
Sulfite P, G Not necessary Analyze immediately
Surfactants P, G Cool, 4 ≤ 6 °C 48 hours

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Water Analysis Guide

Table 7 Necessary containers, preservation techniques and holding times (continued)

Parameter name Container1 Preservation2,3 Maximum holding
time4
Temperature P, G Not necessary Analyze immediately
Turbidity P, G Cool, 4 ≤ 6 °C 48 hours
1 Polyethylene (P), glass (G) or PTFE Teflon
2 Sample preservation should be completed immediately upon sample collection. For composite chemical samples, each
portion should be preserved at the time of collection. When use of an automated sampler makes it impossible to preserve
each portion, chemical samples may be preserved at 4 ≤ 6 °C until compositing and sample splitting is completed.
3 When any sample is to be shipped by common carrier or sent through United States mail, it must comply with the
Department of Transportation Hazardous Material Regulations (49 CFR Part 172). The person offering such material for
transportation is responsible for making sure of such compliance. For the preservation requirements of Table II, the Office
of Hazardous Materials, Materials Transportation Bureau, Department of Transportation have found that the Hazardous
Materials Regulations do not apply to the following materials: Hydrochloric acid (HCl) in water solutions at concentrations
of 0.04% by weight or less (pH about 1.96 or higher); Nitric acid (HNO3) in water solutions at concentrations of 0.15% by
weight or less (pH about 1.62 or higher); Sulfuric acid (H2SO4) in water solutions at concentrations of 0.35% by weight or
less (pH about 1.15 or higher); and Sodium hydroxide (NaOH) in water solutions at concentrations of 0.080% by weight or
less (pH about 12.30 or less).
4 Samples should be analyzed as soon as possible after collection. The times listed are the maximum times that samples
may be held before analysis and still be considered valid. Samples may be held for longer periods only if the permittee or
monitoring laboratory has data on file to show that the specific types of samples under study are stable for the longer time
and the permitee has received a variance from the Regional Administrator under §136.3(e). Some samples may not be
stable for the maximum time period given in the table. A permittee or monitoring laboratory is obligated to hold the sample
for a shorter time if knowledge exists to show that this is necessary to keep sample stability. Refer to §136.3(e) for details.
The term “analyze immediately” usually means within 15 minutes or less after sample collection.
5 Should only be used in the presence of residual chlorine.
6 Maximum holding time is 24 hours when sulfide is present. Optionally, all samples may be analyzed with lead acetate
paper before pH adjustments to find sulfide. If sulfide is present, it can be removed by the addition of cadmium nitrate
powder until a negative spot test is obtained. The sample is filtered and then NaOH is added to pH 12.
7 Samples should be filtered immediately on-site before preservative is added for dissolved metals.
8 From 40 CFR 136.3 - To get the 28-day holding time, use the ammonium sulfate buffer solution specified in EPA Method
218.6. Dissolve 38 g ammonium sulfate in 75 mL ASTM type I water and add 6.5 mL of ammonium hydroxide. Dilute to
100 mL with ASTM type I water. Adjust to pH 9 – 9.5 with the buffer and correct for volume additions.

1.4.1.2.2 Correct for volume additions

When a large volume of preservative or neutralizer is used, account for dilution by the
acid that was added to preserve the sample and/or the base used to adjust the pH to the
range of the procedure. Do the steps that follow to make this correction:

1. Find the volume of the initial sample, the volume of acid and base added and the total
final volume of the sample.
2. Divide the total volume by the initial volume.
3. Multiply the test result by the result of step 2.
Example:
A 1 L sample was preserved with 2 mL of nitric acid. It was neutralized with 5 mL of
5 N sodium hydroxide. The result of the analysis procedure was 10.00 mg/L. What is
the volume correction factor and correct result?
1. Total volume = 1000 mL + 2 mL + 5 mL = 1007 mL
2. 1007 ÷ 1000 = 1.007 = volume correction factor
3. 10 mg/L × 1.007 = 10.07 mg/L = correct result

1.4.1.3 About accuracy and precision

Accuracy defines how near a test result is to the true value. Precision defines how near
repeated measurements are to each other. Although precise results suggest accuracy,
they can be inaccurate. Both the accuracy and the precision of test results can be
evaluated with standard additions or standard solutions. Refer to Figure 10.

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 Water Analysis Guide

Figure 10 Precision vs accuracy

1 Not accurate, not precise 3 Precise, not accurate

2 Accurate, not precise 4 Accurate and precise

1.4.1.4 Standard solutions

A standard solution may be ordered as a prepared reagent or it may be made in the
laboratory. It is a solution of a known composition and concentration. The accuracy of the
analysis system may be identified with a standard solution in place of the sample water in
a procedure.

1.4.1.5 Standard additions

Standard additions is a common technique to identify the accuracy of the test results.
Other names are “spiking” and “known additions.” The technique can identify
interferences, bad reagents, faulty instruments and incorrect procedures.
To complete the standard additions technique, add a measured small amount of a
standard solution to the sample and do the test again. Use the same reagents, equipment
and technique. The result should be about 100% recovery. If not, there is an identifiable
problem.
If the standard additions technique is satisfactory for the test, a standard additions
method section will be in the procedure under Accuracy Check. Complete the detailed
instructions given.
If the result is approximately 100% recovery for each addition, everything is satisfactory
and the results are correct.
If the result is not approximately 100% recovery for each addition, a problem is present.
To identify if the cause is an interference, do the standard additions technique again with
deionized water as the sample. If the result is approximately 100% recovery for each
addition, an interference exists.
If the results show good recoveries with the deionized water, use this checklist to find the
problem:

1. Make sure that the steps in the procedure are done correctly:
a. Are the correct reagents used in the correct order?
b. Is the correct time used to let the color develop?
c. Is the correct glassware used?
d. Is the glassware clean?
e. Does the test need a specific sample temperature?
f. Is the sample pH in the correct range?
Refer to the written procedure to answer these questions.
2. Examine the performance of the instrument with the instructions in the user manual.

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Water Analysis Guide

3. Examine the reagents. Repeat the standard additions technique with new, fresh
reagents. If the results are good, the original reagents were faulty.
4. If nothing else is wrong, the standard is almost certainly defective. Do the standard
additions technique again with a new standard.

If the problem is still not known, contact technical support. Contact information is provided
on the website for all countries.

1.4.1.6 Troubleshoot a test when the results are in doubt

If the results from any chemistry are in doubt, do the steps that follow to troubleshoot.

1. Do a proof-of-accuracy check. Take a standard solution, which has a known

concentration, through the same steps as the original sample. Include sampling and
storage, digestion and colorimetric determination, if applicable. If the results of the
standard solution check are correct, go to step 4. If there is a variation in the
expected results, go to step 2.
2. If the standard solutions check is not the same as the expected results, examine the
instrument set-up and method procedure with the steps that follow:
a. Make sure that the correct program number for the test is selected.
b. Make sure that the units of concentration of the standard are the same as the
units shown. (One of the alternative forms of the analyte may be in the display.)
For example: Molybdenum may be shown as Mo instead of MoO4.
c. Make sure that the sample cells specified in the procedures are used.
d. Make sure that the reagents are correct for the sample size being analyzed.
e. Make sure that the reagent blank value saved is for the current procedure. It may
be from a previous lot of reagents and therefore not representative of the current
reagent lot.
f. Make sure that the calibration curve adjustment (Standard Adjust) is currently in
use. The factory-stored default calibration should be used initially to examine the
standard.
g. Make sure that the dilution factor option is correct.
If the instrument setup is correct and the method procedure specifics are completed
correctly, go to step 3.
3. If the standard solution check does not match the expected results, examine the
reagents used in the test and the analytical technique with the steps that follow:
a. Find the age of the reagents used in the test. Many factors affect reagent shelf
life (i.e., storage temperature, storage conditions, microbial contamination).
Replace suspect reagents and do the standards check again.
b. Do a deionized or distilled water blank through the full process (include sampling
and storage, digestion and colorimetric determination). Some chemicals will add
a small amount of color to a test. This is typical. However, color development
higher than 10% of the range of the test may show a problem with one of the
reagents or the dilution water.
c. To troubleshoot the procedure, delete the parts one by one. First, do the standard
solution, leave out preservation and storage, and do only digestion and
colorimetry. If this analysis is correct, examine the procedure used to keep the
sample. Make sure that it is the procedure prescribed for the selected parameter.
If the sample is acidified for storage, make sure that the correct acid is used and
the sample is adjusted to the proper pH level before the sample is examined.
If the standards check is still incorrect, do the standard on just the colorimetry. If the
results are correct, examine the digestion procedure. Make sure that the amount of

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 Water Analysis Guide

reagents used and the pH after the digestion are correct for the procedure. Refer to
the procedure for the parameter in question.
4. If the standard solution gives a correct value, but the results of the sample
measurement are questionable, there may be an interference in the sample. To look
for an interference:
a. Spike the sample. Use a standard addition test instead of a standard solution test
to include any possible interferences.
b. To test cells that contain fresh sample water, add an amount of standard equal to
two times the concentration of the sample.
c. Do both samples with the same reagents, instruments and technique. The spiked
sample should show an increase equal to the amount of standard added.
d. Calculate percent recovery as shown below. Ideally, the results should be 100%.
The results from 90 to 110% are acceptable. Refer to the procedure notes for
possible interferences and ways to prevent them.
e. Do a series of dilutions on the sample. Make sure that the sample is within the
range of the test. An out-of-range sample for the method may give erroneous
results because of under- or over-development of the color, too much turbidity or
even sample bleaching. Do a series of dilutions to look for this possibility.
f. If the cause of the interference cannot be found, dilute the sample past the point
of interference. This is often the most economical and efficient way to get the
correct result. If it is not possible to dilute out an interference without diluting out
the parameter to be measured, use a different method, such as a different
chemistry or an ion-selective electrode to measure the parameter.

1.4.1.6.1 Calculate the percent recovery

To calculate the percent recovery:

1. Measure the unknown sample concentration.

2. Calculate the theoretical concentration of the spiked sample:
Theoretical concentration = (Cu × Vu) + (Cs × Vs) ÷ Vu + Vs
Where:
Cu = measured concentration of the unknown sample
Vu = volume of the unknown sample
Cs = concentration of the standard
Vs = volume of the standard
3. Measure the spiked sample concentration.
4. Divide the spiked sample concentration by the theoretical concentration and multiply
by 100.
For example:
A sample was tested for manganese and the result was 4.5 mg/L. A separate 97-mL
portion of the same sample was spiked with 3 mL of a 100 mg/L standard solution of
manganese. This spiked solution was examined again for manganese with the same
method. The result was 7.1 mg/L.
The theoretical concentration of the spiked sample is:
(4.5 mg/L × 97 mL) + (100 mg/L × 3 mL) ÷ 97 mL + 3 mL = 7.4 mg/L
The percent spike recovery is:
(7.1 mg/L ÷ 7.4 mg/L) x 100 = 96%

1.4.1.6.2 USEPA calculation

The USEPA uses a more stringent calculation requirement for percent recovery. This
formula calculates the percent recovery only for the standard added to the spiked sample
and gives a lower value than the above calculation. A complete explanation for the

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Water Analysis Guide

USEPA formula is shown in USEPA Publication SW-846. The USEPA percent recovery
formula is:
%R = 100 (Xs – Xu) ÷ K
Where:
Xs = measured value of the spiked sample
Xu = measured value for the unspiked sample, adjusted for the dilution of the spike
volume
K = known value of the spike in the sample
For example:
A sample measures 10 mg/L. A separate 100-mL portion of the sample was spiked with
5 mL of a 100-mg/L standard solution. The spiked solution was measured by the same
method as the original sample. The result was 13.7 mg/L.
Xs = 13.7 mg/L
Xu = (10 mg/L × 100 mL) ÷ 105 mL = 9.5 mg/L
K = (5 mL × 100 mg/L) ÷ 105 mL = 4.8 mg/L
%R = (100 × (13.7 mg/L – 9.5 mg/L)) ÷ 4.8 mg/L = 88%
Acceptable percent recovery values are 80–120%.

1.4.1.7 Adjust the standard curve

Spectrophotometers typically have many programs permanently installed in memory.
Many programs include a pre-programmed calibration curve. Each curve is the result of
an extensive calibration completed under ideal conditions and is normally sufficient for
most testing. Deviations from the curve can occur from compromised testing reagents,
defective sample cells, incorrect test procedure, incorrect technique or other correctable
causes. Interfering substances or other causes may be beyond the control of the analyst.
The pre-programmed curve may not be convenient in the situations that follow:
• Tests are done where the reagents are highly variable from lot to lot.
• Tests are done where frequent calibration curve checks are necessary.
• Tests are done where samples give a consistent test interference.
• Think about the questions that follow before the calibration curve is adjusted:
• Will future test results be better when the curve is adjusted?
• Are interfering substances consistent in all the samples tested?
• Estimated detection limit, sensitivity, precision and test range information
provided with the procedure may not apply to an adjusted curve calibration.
The calibration curves can be adjusted with the steps found in the test procedure.
Generally, add test reagents to a blank and standard solution. It is important to do the
procedure carefully. After the adjustment, it is wise to do standard solutions of several
concentrations to make sure that the adjusted curve is satisfactory. Do standard additions
on typical samples to find out if the adjusted curve is acceptable.
To adjust a measurement is a two-step process. First, the instrument measures the
sample with the pre-programmed calibration. Second, the instrument multiplies this
measurement by an adjustment factor. The factor is the same for all concentrations. The
instrument remembers the factor until the program is exited and shows the standard
adjustment icon when it is used. To return to the pre-programmed curve at any time,
select the original stored program from the main menu.
1.4.2 Interferences
Interferences are contaminants in a sample that can cause changes in color
development, turbidity or unusual colors and odors, and thereby make errors in the
results. A list of common interferences is included in each procedure. Reagents are
formulated to remove many interferences. To remove other interferences, pretreat the
sample as instructed in the procedure.

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 Water Analysis Guide

Test strips are available for many of the common interferences. These test strips can be
used to screen samples for the presence of interferences.
1. Repeat the test on a sample diluted with deionized water. Refer to Sample dilution
on page 15.
2. Correct the results for the dilution and compare them with those from the original test.
3. If they are significantly different, make a second dilution and compare it against the
first.
4. Repeat the dilutions until the same result (after volume corrections) is achieved twice
in succession.
For more information on interferences, refer to Standard additions on page 25. The APHA
Standard Methods book (an excellent reference for water analysis) also shows
interferences in the general introduction section.

1.4.2.1 pH interference
Chemical reactions are often pH dependent. Reagents contain buffers to adjust the pH of
the sample to the correct range. However, the reagent buffer may not be strong enough
for samples that are highly buffered or have an extreme pH. The sampling and storage
section of each procedure gives the pH range for that test.
Before the test, adjust the sample to the proper pH as instructed in the procedure or do
the steps that follow:

1. Measure the pH of the analyzed sample with a pH meter.

Note: Use pH paper to test for chloride, potassium or silver to avoid contamination.
2. Prepare a reagent blank with deionized water as the sample. Add all reagents
specified in the procedure. Timer sequences, etc., may be ignored.
3. Mix well.
4. Measure the pH of the reagent blank with a pH meter.
5. Compare the pH values of the analyzed sample with the reagent blank.
6. If there is little difference in the values of the analyzed sample and the reagent blank,
the pH interference is not the problem. Do the accuracy check for the specific
procedure to more clearly identify the problem.
7. If there is a large difference between the value of the analyzed sample and the
reagent blank, adjust the sample pH to the value of the reagent blank. Adjust the
sample pH to this same pH for all future samples before analysis.
• Use the applicable acid, usually nitric acid, to lower the pH.
• Use the applicable base, usually sodium hydroxide, to increase the pH.
If acid or base was added, adjust the final result for any dilution that was caused.
Refer to Correct for volume additions on page 24.
8. Analyze the sample as done previously.
9. Some purchased standards may be very acidic and do not work directly with test
procedures. Adjust the pH of these standards as described previously. Adjust the final
concentration of the standard for the dilution. The standard solutions suggested in the
procedures are formulated so that no pH adjustment is necessary.

1.4.3 Method performance

1.4.3.1 Estimated detection limit (EDL)
Ranges for chemical measurements have limits. The lower limit is important because it
identifies whether a measurement is different from zero. Many experts disagree about the
definition of this detection limit and find that it can be difficult. The Code of Federal
Regulations (40 CFR, Part 136, Appendix B) provides a procedure to find the Method
Detection Limit (MDL). The MDL is the lowest concentration that is different from zero
with a 99% level of confidence. A measurement below this MDL is highly suspect.

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Water Analysis Guide

The MDL is not fixed. It is different for each reagent lot, instrument, analyst, sample type,
etc. Therefore, a published MDL may be a useful guide, but is only accurate for a specific
set of circumstances. Each analyst should find a more accurate MDL for each specific
sample matrix with the same equipment, reagents and standards that will routinely be
used for measurements.
A sensitivity value (concentration change equivalent to an absorbance change of
0.010 abs) is provided as an estimate of the lower detection limit of each test. The
sensitivity value may be used as an EDL for the purposes of MDL determination. It is a
good starting concentration when a MDL is to be found.
Note: Do not use the EDL as the MDL.
The conditions for MDL determination must be exactly the same as the conditions used
for analysis. The EDL may be useful to the analyst as a starting point when a MDL is to
be found or as a way to compare methods. Measurements below the EDL may also be
valuable because they can show a trend, show the presence of analyte and/or provide
statistical data. However, these values have a large uncertainty.

1.4.3.2 Method detection limit (MDL)

This method is in accordance with the USEPA definition in 40 CFR, Part 136, Appendix B
in the 7-1-94 edition. The USEPA defines the method detection limit (MDL) as the
minimum concentration that can be found with a 99% level of confidence that the true
concentration is higher than zero. Since the MDL is different from analyst to analyst, it is
important that the MDL be found under actual operating conditions.
The procedure to find the MDL is based on replicate analyses at a concentration 1 to
5 times the estimated detection limit. The MDL value is calculated from the standard
deviation of the replicate study results multiplied by the appropriate t value for a 99%
confidence interval. For this definition, the MDL does not account for variation in sample
composition and can only be achieved under ideal conditions.

1. Make an estimate of the detection limit. Use the sensitivity value stated in the Method
performance section of the analysis procedure.
2. Prepare a laboratory standard of the analyte, 1 to 5 times the estimated detection
limit, in deionized water that is free of the analyte.
3. Make an analysis of at least seven portions of the laboratory standard and record
each result.
4. Calculate the average and the standard deviation(s) of the results.
5. Calculate the MDL with the appropriate t value (Table 8) and the standard deviation
value:
MDL = t × s
Table 8 Test portions and t values
Number of test portions t value
7 3.143
8 2.998
9 2.896
10 2.821

For example:
The EDL to measure iron with an iron test is 0.003 mg/L. An analyst accurately
prepared 1 L of a 0.010 mg/L (about 3x the EDL) laboratory standard with a mixture
of a 10-mg/L iron standard in iron-free deionized water.
Eight portions of the standard were examined with the FerroZine method. The results
are shown in Table 9.

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Table 9 Samples and results

Sample # Result (mg/L)
1 0.009
2 0.010
3 0.009
4 0.010
5 0.008
6 0.011
7 0.010
8 0.009

Use a calculator program. The average concentration = 0.010 mg/L and the standard
deviation (s) = 0.0009 mg/L.
Based on the USEPA definition, calculate the MDL:
MDL for iron test = 2.998 (t) x 0.0009 (s)
MDL = 0.003 mg/L (the same as the initial estimate)
Note: Occasionally, the calculated MDL may be very different from the estimate of the
detection limit. To test how reasonable the calculated MDL is, repeat the procedure with a
standard near the calculated MDL. The average result calculated for the second MDL
derivation should agree with the initial calculated MDL. Refer to 40 CFR, Part 136, Appendix B
(7-1-94), pages 635–637 for detailed procedures to make sure that the MDL determination is
correct.

1. Put a laboratory blank (that contains deionized water without analyte) through the
test procedure to to make sure that the blank measurement is less than the
calculated MDL.
2. If the blank measurement is near the calculated MDL, repeat the MDL procedure
with a separate blank for analysis for each portion of standard solution analyzed.
3. Subtract the average blank measurement from each standard and use the
corrected standard values to calculate the average and standard deviation used
in the MDL.

1.4.3.3 Precision
Every chemical measurement has some degree of uncertainty. The quality of the entire
calibration curve determines the precision.
Uncertainty in chemical measurements may be due to systematic errors and/or random
errors. A systematic error is a mistake that is always the same for every measurement
made. For example, a blank can add to each measurement for a specific compound, and
gives consistently high results (a positive bias). Random errors are different for every test
and can add a positive or a negative variation in response. Random errors are most often
caused by variation in analytical technique. Even with reliable reagents developed to
prevent systematic errors, response variation occurs in all chemical measurements.

1.4.3.4 Estimate the precision

The method performance section in each procedure gives an estimate of test precision.
Most procedures use a replicate analysis estimate, based on real data. For precision
found in this manner, the 95% confidence interval of the distribution is reported.
In replicate analysis, the chemist prepares a specific concentration of the analyte in a
deionized water matrix. The standard is analyzed seven individual times on a single
instrument. The standard deviation is calculated and the 95% confidence interval of the
distribution is reported in the method. The reported value gives an estimate of the
“scatter” of results at a particular point in the calibration curve.

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Water Analysis Guide

Precision estimates are based on a deionized water matrix. Precision on real samples
with varying matrices can be quite different from these estimates.
If the concentration achieved from the use of a standard solution is not as expected or
when the results are in doubt, refer to Troubleshoot a test.

1.4.3.5 Sensitivity
The definition of the sensitivity of a method is a change in concentration (ΔConcentration)
for a 0.010 change in absorbance (ΔAbs).
Use sensitivity when different methods are compared. For example, when three methods
are used to find iron (Table 10).
Table 10 Concentration and absorbance
Iron analysis method Portion of curve ΔAbs ΔConcentration
FerroVer Entire range 0.010 0.022 mg/L
FerroZine Entire range 0.010 0.009 mg/L
TPTZ Entire range 0.010 0.012 mg/L

Notice that the FerroZine method has the larger sensitivity of the three methods because
it measures the smallest change in concentration. The technical definition of sensitivity
comes from a calibration curve with Abs on the x-axis and concentration on the y-axis.
• If the calibration is a line, the sensitivity is the slope of the line multiplied by 0.010.
• If the calibration is a curve, the sensitivity is the slope of the tangent line to the curve
at the concentration of interest multiplied by 0.010.
The sensitivity value is also used as an estimate of the lower limit of the test. The value
may be used as a starting point to find the MDL.
1.4.4 Prepare a calibration curve
Note: Calibration curves are recommended when tests are done on different instruments or where
necessary by a regulator.

1. Prepare five or more standards of known concentration that cover the expected range
of the test.
2. Do tests as described in the procedure on each prepared standard.
3. Pour the customary volume of each known solution into a separate clean sample cell
of the type specified for the instrument.
4. Select the proper wavelength. Standardize (zero) the instrument with an untreated
water sample or a reagent blank as specified in the procedure instructions.
5. Measure and record the absorbance of the known solutions within the time
constraints specified in the procedure. To use absorbance vs. concentration, refer to
Absorbance versus concentration calibration on page 32.

1.4.4.1 Absorbance versus concentration calibration

1. If absorbance values are measured, plot the results on linear graph paper.
a. Plot the absorbance value on the y-axis and the concentration on the x-axis.
b. Plot increasing absorbance values from bottom to top.
c. Plot increasing concentration values from left to right.
Values of 0.000 absorbance units and 0 concentration will start at the bottom left
corner of the graph. A calibration table can be extrapolated from the curve or the

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concentration values can be read directly from the graph. Another alternative is to
find an equation for the line with the slope and y-intercept.
2. As an alternative, use the user program software in the spectrophotometer or a
curve-fitting program (such as a spreadsheet software) to calculate the calibration
curve.

1.4.5 Adapt procedures to other spectrophotometers

Test procedures may be used with more than one spectrophotometer if calibration curves
are made that convert absorbance to concentration. Regardless of the spectrophotometer
used, prepare the sample and calibration standards with the procedure and use the
optimum wavelength used in the procedure.
To calibrate for a given analyte, a series of standards are prepared and measured to
make the calibration curve. The absorbance vs. concentration is plotted as described in
Absorbance versus concentration calibration on page 32. Points on the graph are
connected with a smooth line (curved or straight). If necessary, use the curve to make a
calibration table.

1.4.5.1 Select the best wavelength

When a new procedure is developed or procedures that are sensitive to wavelength are
used, select the wavelength where the instrument gives the highest absorbance. Refer to
Figure 11 on page 35. Because procedures are usually developed to use the best
wavelength for the test, the selection of the wavelength is not necessary for most stored
procedures.
1.4.5.1.1 Select the best wavelength on a spectrophotometer
Note: When available, use of the Scan function is the easiest way to find the optimum wavelength.

1. Refer to the user manual for specific instructions for wavelength adjustments.
2. Select single wavelength adjustment.
3. Enter a wavelength in the range of interest.
Note: Sample color provides a good indication of which wavelength region to use.
• A yellow solution absorbs light in the 400–500 nm region.
• A red solution absorbs light between 500–600 nm.
• A blue solution absorbs light in the 600–700 nm range.
4. Prepare the blank and sample for analysis. Fill the applicable sample cells with the
blank and the reacted sample solution.
5. Put the blank in the cell holder. Set the instrument to zero.
6. Put the prepared sample into the cell holder. Read the absorbance level.
7. Increase the wavelength so that it is at least 100 nm higher than the range of interest.
Set the instrument to zero as in step 5. Measure and record the absorbance of the
sample.
8. Repeat, decrease the wavelength by 50 nm. Set the instrument to zero, then
measure and record the absorbance at each increment. Continue this process
throughout the wavelength range of interest. Record the wavelength of highest
absorbance. Refer to Table 11.
Table 11 Absorbance values at 50-nm increments
Wavelength Absorbance
550 nm 0.477
500 nm 0.762

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Water Analysis Guide

Table 11 Absorbance values at 50-nm increments (continued)

Wavelength Absorbance
450 nm 0.355
400 nm 0.134

9. Adjust the wavelength to 50 nm more than the highest absorbance point on the initial
search (step 8). Set the instrument to zero, then measure and record the absorbance
at each increment.
Repeat, decreasing the absorbance in 5-nm steps. Set the instrument to zero, then
measure and record the absorbance at each increment. Continue until the entire
range of interest is measured. Refer to Table 12.
Table 12 Absorbance values at 5-nm increments
Wavelength Absorbance
520 nm 0.748
515 nm 0.759
510 nm 0.780
505 nm 0.771
500 nm 0.771
495 nm 0.651
490 nm 0.590

Make sure that there is enough difference in absorbance between samples with low
and high analyte concentrations. Measure two sample solutions that contain the
expected low and high concentrations of analyte at the optimum wavelength. The
change in absorbance caused by increases/decreases in concentration depends on
the sensitivity of the procedure and the chemistry. Chemistries with small absorbance
changes are less sensitive, but tend to have larger ranges. Chemistries with large
absorbance changes are more sensitive, but tend to have smaller ranges.

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 Water Analysis Guide

Figure 11 Select the best wavelength

1 Absorbance 2 Wavelength (nm)

1.5 Sample pretreatment by digestion

Several procedures use sample digestion before the total metal content is found.
Digestion uses acid and heat to break organo-metallic bonds and free ions for analysis.
1.5.1 USEPA-approved digestions
For USEPA reporting, USEPA-approved digestions are necessary. There are two
methods for metals analysis: mild and vigorous.

1.5.1.1 USEPA mild digestion

1. Add concentrated nitric acid to the entire sample at the time of collection. Add 5 mL of
acid per liter (or quart) of sample.
2. Move 100 mL of well-mixed sample to a beaker or flask.
3. Add 5 mL of distilled 1:1 hydrochloric acid (HCl).
4. Increase the temperature of the liquid with a steam bath or hot plate until the volume
has been reduced to 15–20 mL. Do not boil.
5. Use a filter to remove any insoluble material from the sample.
6. Adjust the pH of the digested sample to pH 4. Add 5.0 N Sodium Hydroxide Standard
Solution a drop at a time. Mix thoroughly and examine the pH after each addition.
7. Pour the reduced sample into a 100-mL volumetric flask.
8. Use a small amount of demineralized water to rinse the beaker. Pour the rinse water
into the volumetric flask.
9. Repeat the rinse process a few more times to remove all of the reduced sample from
the beaker.
10. Add demineralized water to fill the volumetric flask to the 100-mL mark.
11. Use the diluted sample in the test procedure. Record the results.

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Water Analysis Guide

12. Prepare a blank: Repeat steps 1-11 with demineralized water instead of the sample.
13. Subtract the results of the blank analysis from the results of the sample analysis.

1.5.1.2 USEPA vigorous digestion

For some samples mild digestion will not be sufficient. Use a vigorous digestion to make
sure that all of the organo-metallic bonds are broken.

1. Use redistilled 1:1 Nitric Acid Solution to acidify the entire sample to a pH of less than
pH 2. Do not filter the sample before digestion.
2. Move an appropriate sample volume into a beaker and add 3 mL of concentrated
redistilled nitric acid. Refer to Table 13.
3. Put the beaker on a hot plate and evaporate to near dryness. Make sure that the
sample does not boil.
4. Cool the beaker and add another 3 mL of the concentrated re-distilled nitric acid.
5. Put the cover on the beaker with a watch glass and return it to the hot plate. Increase
the temperature of the hot plate so that a gentle reflux occurs. Add additional acid, if
necessary, until the digestion is complete (generally shown when the digestate is light
in color or does not change color or appearance with continued refluxing).
6. Again, evaporate to near dryness (do not bake) and cool the beaker. If any residue or
precipitate results from the evaporation, add redistilled 1:1 hydrochloric acid (5 mL
per 100 mL of final volume). Refer to Table 13.
7. Warm the beaker. Adjust the sample to pH 4 by drop-wise addition of 5.0 N Sodium
Hydroxide Standard Solution. Mix thoroughly and examine the pH after each addition.
8. Pour the reduced sample into a 100-mL volumetric flask.
9. Use a small amount of demineralized water to rinse the beaker. Pour the rinse water
into the volumetric flask.
10. Repeat the rinse process a few more times to remove all of the reduced sample from
the beaker.
11. Add demineralized water to fill the volumetric flask to the 100-mL mark.
12. Use the diluted sample in the test procedure. Record the results.
13. Multiply the result by the correction factor in Table 13.
14. Prepare a blank: Repeat steps 1-13 with demineralized water instead of the sample.
15. Subtract the results of the blank analysis from the results of the sample analysis.
Table 13 Vigorous digestion volumes
Expected metal Suggested sample Suggested volume of Suggested final Correction factor
concentration volume for digestion 1:1 HCl volume after digestion
1 mg/L 50 mL 10 mL 200 mL 4
10 mg/L 5 mL 10 mL 200 mL 40
100 mg/L 1 mL 25 mL 500 mL 500

1.5.2 General Digesdahl digestion

Many samples may be digested with the Digesdahl Digestion Apparatus (2313020). It is
designed to digest samples such as oils, wastewater, sludges, feeds, grains, plating
baths, food and soils. In this procedure, the sample is oxidized by a mixture of sulfuric
acid and hydrogen peroxide. Less than 10 minutes is necessary for the digestion of a dry
sample. About 1 minute/mL is necessary for the digestion of liquid samples. The digestion
is done in a special flat-bottomed, 100-mL volumetric flask. Aliquots (sample portions) are
used for analysis with the colorimetric methods.
Procedures for digestion with the Digesdahl Digestion Apparatus are based on the type
and form of the sample. Refer to the Digesdahl Digestion Apparatus Instruction manual
supplied with the Digesdahl Digestion Apparatus.

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 Water Analysis Guide

Digesdahl digestion is a process that yields a digest that can be used to find metals, total
phosphorus and total Kjeldahl nitrogen (TKN). It is faster than traditional methods, but has
comparable accuracy and precision. The digest can be used with colorimetric,
turbidimetric or titrimetric tests.
The procedures for the Digesdahl Digestion Apparatus vary with the sample type. Sample
types include food products, feeds, grains, wastewater sludges, plating baths, plant
tissues, fertilizers, beverages and oils. Most procedures use a two-phase digestion
process that uses concentrated sulfuric acid and 50% hydrogen peroxide. Sulfuric acid
dehydrates and chars the sample. Hydrogen peroxide is added through the capillary flow
funnel to complete the decomposition. The analyst varies the volume of hydrogen
peroxide used to control the digestion time (exposure to the hydrogen peroxide).
Some samples are more difficult to digest completely (e.g., resistant or refractory
materials, such as nicotinic acid). Several minutes of continued peroxide digestion are
necessary after clearing to get 100% nitrogen recovery. To make sure that there is
complete sample digestion, think about variables such as sample size, solution
temperature and sample contamination. Refer to the Digesdahl Manual (2313018) for
complete information.

1.5.2.1 Frequently asked questions for digestion procedures

This section provides answers to common questions about digestion.
What should be done if the reading on the instrument is over-range?
The concentration range tables found in digestion procedures are only guidelines. Use a
smaller analysis volume and repeat the procedure. Record the new analysis volume and
use it in the calculation.
Should a reagent blank be prepared each time reagents with the same lot number
are used?
To decide, first find the reading of the reagent blank. Set the instrument to zero with
deionized or distilled water. If the reagent blank has an insignificant concentration reading
and the reagents have the same lot number, a reagent blank does not have to be
prepared every time. If the reagent blank shows a reading, analyze it daily or subtract the
reading from the sample reading. If a reagent blank is not analyzed daily, set the
instrument to zero with deionized water.
Does the exact sample amount and analysis volume given in each procedure need
to be used?
The sample amount and the analysis volume for each procedure are only suggested
guidelines. Digest any aqueous solution or suspension sample amount up to 40 mL. Less
than 0.5 g of anhydrous material is necessary for solid or organic liquid samples—as a
routine practice, 0.25 g of sample is used.
How can the initial amount of sample (necessary for digestion) and the analysis
volume to be used be refined?
The amount of sample to be digested is a critical aspect of the digestion. The aliquot size
of the digest to be used in the analysis is also very important. Tables are provided in each
method to find the amount of initial sample to be digested. In order to optimize the
specific test to be done, the equations that follow have been developed. Before these
equations are used, refer to the manual specifications for the sample type.
To use the equations, find the approximate concentration (in ppm, mg/L or mg/kg). Next,
find the range of the colorimetric test to be used (e.g., 0–50 mg/L) and the midpoint of the
test range. This midpoint range is optimum but can be lowered to accommodate very low
sample concentrations. To find the midpoint of the test range, subtract the lower limit of
the range from the higher limit and then divide by 2.
After these determinations are finished, use the equation that follows:
A = (B × C × D) ÷ (E x F)
Where:
A = approximate concentration of sample

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Water Analysis Guide

B = midpoint of colorimetric test range

C = final volume of digest
D = final volume of analysis
E = sample amount to digest
F = analysis volume of digest
Use algebra to obtain the equations that follow:
Equation 1 is E = (B × C × D) ÷ (A × F)
Equation 2 is F = (B × C × D) ÷ (A × E)
Both equations contain two unknown values, E and F. Some trial and error may be
necessary to get the optimum values.
Use equation 1: If the analysis is for copper, use the CuVer™ method with an initial
sample that contains approximately 150 ppm Cu. The amount of sample necessary for
digestion and the aliquot volume to be used can be found as follows:
Find the test range. In this example, the test range is thought to be 0–5.0 ppm and the
midpoint is 2.5. When the Digesdahl system is used, the final volume of digest is 100 mL
and the procedure calls for a final analysis volume of 25 mL.
Therefore:
A = 150
B = 2.5
C = 100
D = 25
E = unknown
F = unknown
Substitute values into equation (1) gives:
E = (2.5 × 100 × 25) ÷ (150 × F) or E = 41.7 ÷ F
Since CuVer Copper Reagent is pH sensitive, a small analysis volume (0.5 mL) is
necessary so that pH adjustment would not be necessary.
With this in mind, a 0.5-mL analysis volume would give:
E = 41.7 ÷ 0.5 = 83.4 mL digestion sample amount
Because the maximum digestion sample amount is 40 mL for Digesdahl digestions, a 0.5-
mL analysis volume is not acceptable for the range. This is where trial and error is
necessary. Next, try a 5.0-mL analysis volume and the equation gives:
E = 41.7 ÷ 5.0 = 8.0 mL digestion sample amount
(Round to the nearest whole number for ease of measure.)
From the calculation, an 8.0 mL sample is digested and a 5.0-mL analysis volume is
taken. A pH adjustment is necessary before analysis.
Use equation 2: Equation 2 may be used when a minimum sample size is necessary or
when a sample has already been digested for one parameter (such as copper) and
measurement for another parameter (such as zinc) is necessary. Continue the example
for copper, above, a zinc test may also be done. The undigested sample contains
approximately 3 ppm zinc and the Zincon method is used. The analysis volume can be
found as follows.
In this example, the Zincon method test range is thought to be 0–2.5 ppm so that the
midpoint of the range is 1.25. Therefore values are:
A=3
B =1.25
C = 100
D = 50
E = 8 (as found above)

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 Water Analysis Guide

substitute: F = (1.25 × 100 × 50) ÷ (3 × 8) = 260 mL

This is an extreme example, but it shows the need to compare the values of D and F to
make sure that the analysis volume (F) is no more than the final analysis volume (D). If F
exceeds D, the analysis cannot be done. A test with a more applicable range is
necessary or a larger sample may be digested for this test. Care must also be taken to
make sure that the volume of digest taken for analysis (F) is higher than 0.1 mL because
accurately pipetting less than 0.1 mL is difficult.
As a comparison, think of the zinc concentration as 75 ppm (A = 75 instead of 3) and
substitute again to get:
F = (1.25 × 100 × 50) ÷ (75 × 8) = 10.5 mL
In this case, the aliquot volume is less than the final analysis volume so analysis may be
done as specified in the procedure.
Why is the factor in the calculation step 75, 2500 or 5000 (depends on the method
used) and where does the factor come from?
In all cases, the factor is a correction for sample dilution. For example, in some tests the
factor is 2500. The Digesdahl digestion total volume is 100 mL, the analysis total volume
is 25 mL and 100 x 25 = 2500. The mL units are not included with the factor because they
cancel out in the formula.
When a slurry is analyzed, how is the total concentration on a dry basis reported?
The sample must be analyzed for moisture content. For necessary apparatus, refer to
Table 14 and Table 15.
To find the dry basis weight:
1. Weigh an aluminum dish and record the weight as “A”.
2. Weigh out approximately 2 g of solid sample into the dish. Record the exact weight
added as “B.”
3. Put the dish in the oven (103–105 °C, 217–221 °F) for 2 hours.
4. Put in a desiccator and cool to room temperature.
5. Weigh the aluminum dish with the oven-dried sample. Record as “C.”
Note: The oven-dried material generally is not meant for additional testing and should be
discarded.
6. Use this formula to calculate the sample on a “dry basis.” Test result (dry basis) = (C
– A) ÷ (B – A).
Note: Multiply the test result on an “as is” basis, by the factor above, to report as “dry basis".

Table 14 Necessary apparatus for dry basis weight

Description Unit Item no.
Balance, analytical, 120-g 454 g 2936801
Desiccant, Drierite (without indicator) each 2285901
Desiccator, vacuum (uses ceramic plate) 100/pkg 2088800
Dish, moisture determination, aluminum, 63 x 17.5 mm each 2164000
Tongs, crucible each 56900
Oven, laboratory, 120 VAC each 1428900
or
Oven, laboratory, 240 VAC each 1428902

Table 15 Optional apparatus

Description Unit Item no.
Desiccator, without stopcock each 1428500

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Water Analysis Guide

1.5.2.2 Adjust the pH

1.5.2.2.1 For a metals procedure
Note: If aliquots smaller than 0.5 mL are analyzed, pH adjustment is not necessary.

1. Find the necessary volume of sample for analysis from the Sample and Analysis
Volume Tables after each digestion procedure. Use a pipet to add this volume into a
graduated mixing cylinder.
Note: To use a pipet to add a volume into a volumetric flask or a regular graduated cylinder is
necessary for some methods.
2. Dilute to about 20 mL with deionized water.
3. Add one drop of 2,4 Dinitrophenol Indicator Solution.
4. Add one drop of 8 N Potassium Hydroxide (KOH) Standard Solution (28232H). Swirl
after each addition until the first flash of yellow shows (pH 3). If the sample is
analyzed for potassium, use 5 N sodium hydroxide (245026) instead. Do not use a
pH meter if the sample is analyzed for potassium or silver.
5. Add one drop of 1 N KOH (2314426). Put the stopper in the cylinder and invert
several times to mix. If the sample is analyzed for potassium, use 1 N sodium
hydroxide instead.
Note: Use pH paper to make sure that the pH is 3. If it is higher than 4, do not adjust again with
acid. Start over with a fresh aliquot.
6. Continue to add 1 N KOH in this manner until the first permanent yellow color shows
(pH 3.5–4.0).
7. Look at the cylinder from the top against a white background. Compare the cylinder to
a second cylinder filled to the same volume with deionized water.
Note: High iron content will cause precipitation (brown cloud) which will co-precipitate other
metals. Do this procedure again with a smaller aliquot volume.
8. Add deionized water to the volume specified in the colorimetric procedure for the
parameter under analysis.
9. Continue with the colorimetric procedure.

1.5.2.2.2 For the Total Kjeldahl Nitrogen colorimetric method

Consult the spectrophotometer or colorimeter procedure to complete the TKN analysis.
The procedure that follows is only a guide to use if a procedure is not available.

1. Use a pipet to add an appropriate analysis volume to a graduated mixing cylinder.

2. Add one drop of TKN Indicator (2251900).
3. Add one drop of 8 N KOH Standard Solution (28232H), swirl after each addition until
the first flash of pale blue shows (pH 3).
4. Add one drop of 1 N KOH (2314426). Put the stopper in the cylinder and invert
several times to mix.
Note: Look at the cylinder from the top against a white background. Compare the cylinder to a
second cylinder filled to the same volume with deionized water.
5. Continue to add 1 N KOH in this manner until the first permanent blue color shows.
6. Add deionized water to the volume shown in the colorimetric procedure for the
parameter under analysis.
7. Continue with the colorimetric procedure.

1.6 Waste management and safety

This section provides guidelines for laboratory waste management. These guidelines are
only a summary of basic USEPA requirements and do not relieve the user from
compliance with the complete regulations contained in the Code of Federal Regulations
(CFR). The regulations may change or additional state and local laws may apply. Waste
generators are responsible for knowing and obeying all the laws and regulations that
apply to their operations.

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 Water Analysis Guide

1.6.1 Waste minimization

The most effective way to decrease waste management problems and expense is
through waste minimization. To do this:
• Use the smallest sample size that will make accurate results.
• Where possible, do methods that use reagents that pose fewer hazards.
• Purchase in smaller quantities to remove the need to dispose of out-dated materials.
• Use biodegradable detergents to clean glassware and apparatus unless solvents or
acids are a specific requirement.

1.6.2 Regulatory overview

The Resource Conservation and Recovery Act (RCRA) controls all solid waste disposal
with an emphasis on hazardous waste. Title 40 Code of Federal Regulations (CFR) part
260 contains the federal hazardous waste disposal regulations issued in accordance with
the RCRA. The regulations create a system to identify hazardous wastes and track waste
generation, transport and ultimate disposal from beginning to end. Each facility involved
in hazardous waste management must be registered with the USEPA, with the exception
of conditionally exempt small quantity generators.
Federal regulations recognize three categories of generators and those who make larger
amounts of waste are under stricter control. The categories are:
• Conditionally exempt small quantity generator—less than 100 kg (220 lb) per month
• Small quantity generator—between 100 kg (220 lb) and 1000 kg (2200 lb) per month
• Large quantity generator—higher than 1000 kg (2200 lb) per month

1.6.3 Hazardous waste

1.6.3.1 Definition
For regulatory purposes, a hazardous waste is a material that is subject to special
consideration by the USEPA under 40 CFR 261. State or local authorities may also
designate additional materials as hazardous waste in their areas.
Many toxic compounds are not regulated, but improper management or disposal may
lead to legal problems under CERCLA (Superfund) or common law tort.
The definition given by 40 CFR 261 identifies a hazardous waste as a solid waste that is
not excluded from regulation and meets one or more of the criteria that follow:
• It is a discarded commercial chemical product, off-specification species, container
residue or spill residue of materials specifically listed in 40 CFR 261.33 (P- and U-
codes)
• It is a waste from a specific source listed in 40 CFR 261.32 (K-code)
• It is a waste from a non-specific source listed in 40 CFR 261.31 (F-code), and/or
• It shows any of the characteristics of hazardous waste that follow:
• Ignitability
• Corrosivity
• Reactivity
• Toxicity
There are exceptions to these regulations. Look at the regulations to find applicable
exclusions.

1.6.3.2 Sample codes

Hazardous wastes are managed by specific codes assigned in 40 CFR 261.20–261.33.
These codes are provided to help identify hazardous waste. The generator is responsible
to make the actual waste code determination.

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Water Analysis Guide

Selected characteristic waste codes for chemicals which may be generated with methods
for water analysis are given in Table 16. A complete list of waste codes is found in
40 CFR 261.20 through 40 CFR 261.33.
Table 16 Hazardous waste codes
Characteristic USEPA code Chemical abstract services (CAS) number Regulatory level (mg/L)
Corrosivity D002 — —
Ignitability D001 — —
Reactivity D003 — —
Arsenic D004 6440-38-2 5.0
Barium D005 6440-39-3 100.0
Benzene D018 71-43-2 0.5
Cadmium D006 7440-43-9 1.0
Chloroform D022 67-66-3 6.0
Chromium D007 7440-47-3 5.0
Lead D008 7439-92-1 5.0
Mercury D009 7439-97-6 0.2
Selenium D010 7782-49-2 1.0
Silver D011 7440-22-4 5.0

1.6.3.3 How to tell if waste is hazardous

Federal laws do not require material testing to find out if waste is hazardous. If the
product is not specifically shown in the regulations, apply product or generator knowledge
to find out if it is hazardous. Often, there is enough information on a Safety Data Sheet
(SDS) to decide. Look for characteristics of a hazardous waste:
• The flash point is below 60 °C (140 °F) or it is classified by DOT as an oxidizer
(D001).
• The pH of the material is less than or equal to pH 2, or higher than or equal to
12.5 (D002).
• The material is unstable, reacts violently with water, may make toxic gases when
mixed with water (D003).
• It is toxic (D004–D043).
Use the chemical composition data to find out if a material is toxic based on the
concentration of certain contaminants (heavy metals and a number of organic
compounds). If the waste is a liquid, compare the concentration of contaminants to the
concentrations shown in 40 CFR 26. If the waste is a solid, make an analysis of the
sample with the Toxicity Characteristic Leachability Procedure (TCLP) and then compare
the results to the concentrations in 40 CFR 261.24. Levels above the threshold amounts
should be thought of as hazardous.
For more information about SDS usage, refer to Safety data sheets on page 45.
Some tests use or make a number of chemicals that make the end product a hazardous
waste (e.g., the COD tests and Nessler reagent). Hazardous waste status may also result
from substances present in the sample.

1.6.3.4 Disposal
Hazardous waste must be managed and discarded according to federal, state and local
regulations. The waste generator is responsible for hazardous waste determinations.
Analysts should speak with their facility environmental compliance department for specific
instructions.

42
 Water Analysis Guide

Most hazardous wastes should be moved by treatment, storage and disposal facilities
(TSDF) that have USEPA permits. In some cases, the generator may treat the hazardous
waste, but may need a permit from the USEPA and/or state agency. Laboratories are not
exempt from these regulations. If the facility is a “Conditionally Exempt Small Quantity
Generator,” special rules may apply. Look at 40 CFR 261 to find the laws and rules that
apply for a given generator.
The most common acceptable treatment is elementary neutralization. This applies to
wastes that are hazardous only because they are corrosive or are listed only for that
reason. Many generated wastes may be treated with the steps that follow:

1. To neutralize acidic solutions, add a base such as sodium hydroxide. To neutralize

basic solutions, add an acid such as hydrochloric acid.
2. Slowly add the neutralizing agent while the solution is stirred.
3. Monitor the pH.
4. When the pH is at or near pH 7, the material is neutralized and may be flushed down
the drain.

For other chemical or physical treatments, such as cyanide destruction or evaporation, a

permit may be necessary. Speak with the environmental department or local regulators to
find which rules apply to each facility.
Laboratory chemicals may be mixed and disposed of with other hazardous wastes
generated at a facility. They may also be collected in accordance with 40 CFR
262.34 satellite accumulation rules. After collection, they may be disposed of in a
labpack. Many environmental and hazardous waste companies offer labpacking services.
These companies will inventory, sort, pack and arrange for proper disposal of hazardous
waste. Find companies that offer these services in the telephone book under “Waste
Disposal—Hazardous” or contact state and local regulators for assistance.
1.6.4 Management of specific waste
Recycling programs for some forms of laboratory waste are available through Hach
Company. To get information on recycling services, call 1-800-227-4224 or visit
www.hach.com.
Several documents are also available to assist in the management of generated waste.
To get the documents, call 1-800-227-4224 or 970-669-3050 and request the literature
codes in Table 17.
Table 17 Waste management literature
Literature code Title
9323 Mercury Waste Disposal Firms
9324 RCRA Waste Disposal Information
9325 COD Waste Disposal Information
9326 COD Heavy Metal Concentrations

1.6.4.1 Special considerations for Cyanide-containing materials

Several procedures in this manual use reagents that contain cyanide compounds. These
materials are regulated as reactive waste (D003) by the Federal RCRA. Instructions
provided with each procedure tell how to collect these materials for proper disposal. It is
imperative that these materials be moved safely to prevent the release of hydrogen
cyanide gas (an extremely toxic material with the smell of bitter almonds). Most cyanide
compounds are stable and can be safely kept for disposal, in highly alkaline solutions (pH
>11) such as 2 N sodium hydroxide. Never mix these wastes with other laboratory wastes
that may contain lower pH materials such as acids or even water.

43
Water Analysis Guide

If a cyanide-containing compound is spilled, avoid exposure to hydrogen cyanide gas. Do

the steps that follow to destroy the cyanide compounds in an emergency:

1. Use a fume hood, supplied air or self-contained breathing apparatus.

2. Stir as the waste is added to a beaker that contains a strong solution of sodium
hydroxide and calcium hypochlorite or sodium hypochlorite (household bleach).
3. Add an excess of hydroxide and hypochlorite. Let the solution stand for 24 hours.
4. Neutralize the solution and flush it down the drain with a large amount of water. If the
solution contains other regulated materials such as chloroform or heavy metals, it
may still need to be collected for hazardous waste disposal. Do not flush untreated
hazardous wastes down the drain.

1.6.5 Resources
Many sources of information on proper waste management are available. The USEPA
has a hotline number for questions about the Resource Conservation and Recovery Act
(RCRA). The RCRA Hotline number is 1-800-424-9346. Copies of the applicable
regulations are available. Federal hazardous waste regulations are found in 40 CFR
260-99. Get this book from the U.S. Government Printing Office or an alternate vendor.
Other documents that may be helpful to the hazardous waste manager in the laboratory
include:
• Task Force on Laboratory Waste Management. Laboratory Waste Management, A
Guidebook; American Chemical Society, Department of Government Relations and
Science Policy: Washington, DC 1994.
• Task Force on Laboratory Waste Management. Waste Management Manual for
Laboratory Personnel; American Chemical Society, Department of Government
Relations and Science Policy: Washington, DC 1990.
• Task Force on Laboratory Waste Management. Less is Better; 2nd ed.; American
Chemical Society, Department of Government Relations and Science Policy:
Washington, DC 1993.
• Committee on Chemical Safety. Safety in Academic Chemistry Laboratories, 5th ed.;
American Chemical Society: Washington, DC, 1990.
• Armour, Margaret-Ann. Hazardous Laboratory Chemicals Disposal Guide; CRC
Press: Boca Raton, FL, 1991.
• Environmental Health and Safety Manager’s Handbook; Government Institutes, Inc.:
Rockville, MD, 1988.
• Lunn, G.; Sansone, E.B. Destruction of Hazardous Chemicals in the Laboratory; John
Wiley and Sons: New York, 1990.
• National Research Council. Prudent Practices for Disposal of Chemicals from
Laboratories; National Academy Press: Washington, DC, 1983.
• National Research Council. Prudent Practices for Handling Hazardous Chemicals in
Laboratories; National Academy Press: Washington, DC, 1981.
• Environmental Protection Agency, Office of Solid Waste and Emergency Response.
The RCRA Orientation Manual; U.S. Government Printing Office: Washington, DC,
1991.
• Environmental Protection Agency, Office of Solid Waste and Emergency Response.
Understanding the Small Quantity Generator Hazardous Waste Rules: A Handbook
for Small Business; U.S. Government Printing Office: Washington, DC, 1986.

1.6.6 Safety
Safety is the responsibility of every analyst. Many chemical procedures use potentially
hazardous chemicals and equipment. It is important to use good laboratory techniques
and prevent accidents. The guidelines that follow apply to water analysis and are not
intended to cover every aspect of safety.

44
 Water Analysis Guide

1.6.6.1 Read labels carefully

Read each reagent label carefully. Note the precautions given. Do not remove or cover
the label on a container while it contains reagent. If a different reagent is put into a
labeled container, the label must be changed. When a reagent or standard solution is
prepared, label the container clearly. If a label is hard to read, replace the label promptly
according to the hazard communication program.
Warning labels also show on some of the apparatus used with the test procedures. The
protective shields with the Digesdahl Digestion Apparatus point out potential hazards.
Make sure that these shields are installed during use and obey the precautions on the
label.

1.6.6.2 Protective equipment

Use the applicable protective equipment for the chemicals and procedures. The SDS
contains this information. Protective equipment may include:
• Eye protection, such as safety glasses or goggles, to protect from flying objects or
chemical splashes.
• Gloves to protect skin from toxic or corrosive materials, sharp objects, very hot or
very cold materials or broken glass. Use tongs or finger cots when a hot apparatus is
transferred.
• Laboratory coats or splash aprons to protect skin and clothing from splashes.
• Footwear to protect feet from spills. Open toed shoes should not be worn in chemistry
settings.
• Respirators may be necessary if sufficient ventilation, such as fume hoods, are not
available. Use fume hoods when directed to do so by the procedure or as
recommended in the SDS. For many procedures, adequate ventilation is enough if
there is plenty of fresh air and air exhaust to protect against unnecessary exposure to
chemicals.

1.6.6.3 First aid equipment and supplies

Most first aid instructions for chemical splashes in eyes or on skin call for a thorough flush
with water. Laboratories should have eyewash and shower stations. For field work, carry
a portable eyewash unit. Laboratories should also have the necessary fire extinguishers
and fume hoods.

1.6.6.4 General safety rules

Obey these rules when work is done with toxic and hazardous chemicals:
• Never pipet by mouth. Always use a mechanical pipet or pipet bulb to avoid ingestion
of chemicals.
• Follow test procedures carefully and observe all precautionary measures. Read the
entire procedure before the procedure is started.
• Clean all spills promptly. Get proper training and have the right response equipment
to clean up spills. Refer to the safety director for more information.
• Do not smoke, eat or drink in an area where toxic or irritating chemicals are used.
• Use reagents and equipment only as directed in the test procedure.
• Do not use damaged labware and broken equipment.
• Minimize all chemical exposures. Do not breathe vapors or let chemicals touch the
skin. Wash hands after chemicals are used.
• Keep work areas neat and clean.
• Do not block exits or access to emergency equipment.

1.6.7 Safety data sheets

Safety data sheets (SDS) describe the hazards of chemical products. This section
describes the information found on the SDS and tells how to find important information for
safety and waste disposal. The information provided on the SDS applies to the product as

45
Water Analysis Guide

sold by a specific manufacturer. The properties of any mixtures made with this product
will be different.

1.6.7.1 How to get an SDS

The SDS is shipped to a customer with the first order of any chemical product. A new
SDS may be sent when the information on the data sheet is updated. Review all new
SDS documents for new information. To get another copy of an SDS, call
1-800-227-4224 or download it directly from www.hach.com.

1.6.7.2 Sections of an SDS

Each SDS has 10 sections. The sections and the information included are described
below.
Header information
The manufacturer order number, SDS date, change number, company address and
telephone number and emergency telephone numbers are shown at the top of the SDS.
1.6.7.2.1 Product identification
This sections contains:
• Product name
• Chemical Abstract Services (CAS) number
• Chemical name
• Chemical formula, if applicable
• Chemical family to which the material belongs
1.6.7.2.2 Ingredients
This section shows each component in the product. It contains the information that
follows for each component:
• PCT: Percent by weight of this component
• CAS NO.: Chemical Abstract Services (CAS) registry number for this component
• SARA: Superfund Amendments and Reauthorization Act, better known as the
“Community Right to Know Law,” tells if the component is shown in SARA 313. If the
component is shown and the facility uses more than the specified amount, report use
to the USEPA every year.
• TLV: Threshold Limit Value. The maximum airborne concentration for an 8-hour
exposure that is recommended by the American Conference of Governmental
Industrial Hygienists (ACGIH).
• PEL: Permissible Exposure Limit. The maximum airborne concentration for an 8-hour
exposure that is regulated by the Occupational Safety and Health Administration
(OSHA).
• HAZARD: Physical and health hazards of the component are explained.
1.6.7.2.3 Physical data
The physical properties of the product are given in this section. The physical properties
include the physical state, color, odor, solubility, boiling point, melting point, specific
gravity, pH, vapor density, evaporation rate, corrosivity, stability and storage precautions.
1.6.7.2.4 Fire and explosion hazard and reactivity data
This section contains the flash point and flammable limits of the material. It also includes
how to fight fires if the material catches on fire. Key terms in this section include:
• Flash point: The temperature at which a liquid will give off enough flammable vapor to
ignite.
• Flammability and ignitability are usually defined by the flash point.
• Lower Flammable Limit (LFL or LEL): The lowest concentration that will cause a fire
or flash when an ignition source is present.
• Upper Flammable Limit (UFL or UEL): The vapor concentration in air that has a
concentration that is too rich to burn.

46
 Water Analysis Guide

• NFPA Codes: The National Fire Protection Association (NFPA) has a system to rate
the degree of hazards given by a chemical. These codes are usually put in a colored
diamond. The codes range from 0 for minimal hazard to 4 for extreme hazard. The
codes are grouped into the hazards that follow: health (blue), flammability (red),
reactivity (yellow) and special hazards (white).
1.6.7.2.5 Health hazard data
This section describes the pathways for a chemical to enter the body (i.e., ingestion,
inhalation, skin contact). It also gives acute (immediate) and chronic (long-term) health
effects. If the material causes cancer or genetic damage, it is specified in this section.
1.6.7.2.6 Precautionary measures
This section contains special precautions for the material. The precautions may include
special storage instructions, handling instructions, conditions to avoid and protective
equipment necessary to use this material safely.
1.6.7.2.7 First aid
First aid instructions for exposures to the chemical are given in this section. Be sure to
read this section before a victim is induced to vomit. Some chemicals are better treated if
the victim does not vomit. Get immediate medical attention for all chemical exposures.
1.6.7.2.8 Spill and disposal procedures
This section describes safe practices for the clean up and disposal of spilled material. For
more information, refer to Hazardous waste on page 41. The waste generator is
ultimately responsible to meet the federal, state and local laws that apply to each facility.
1.6.7.2.9 Transportation data
Domestic and international shipping information is provided in this section. The shipping
name, hazard class and ID number of the product are given.
1.6.7.2.10 References
This section shows the reference materials used to write the SDS.
Refer to the Reference section that shows that this product has SARA 313 chemicals or
California Proposition 65 List Chemicals, if applicable. Any special information about the
product is found here.

1.6.7.3 OSHA chemical hygiene plan

The Occupational Safety and Health Administration (OSHA) enforces laws controlling
exposure to hazardous chemicals in laboratories. These regulations are found in Title
29 CFR 1910.1450. The regulations apply to all employers who use hazardous chemicals
and make it necessary for employers to develop and use a written Chemical Hygiene
Plan and to appoint a qualified person as the Chemical Hygiene Officer.

47
Water Analysis Guide

48
Aluminum DOC316.53.01002

Aluminon Method1 Method 8012

0.008 to 0.800 mg/L Al3+ (spectrophotometers) Powder Pillows
0.01 to 0.80 mg/L Al3+ (colorimeters)
Scope and application: For water and wastewater.
1 Adapted from Standard Methods for the Examination of Water and Wastewater.

Test preparation

Instrument specific information

The table in this section shows all of the instruments that have the program for this test.
Table 1 shows sample cell and orientation requirements for reagent addition tests, such
as powder pillow or bulk reagent tests.
To use the table, select an instrument, then read across to find the corresponding
information for this test.
Table 1 Instrument-specific information for powder pillows
Instrument Sample cell orientation Sample cell
DR 6000 The fill line is to the right. 2495402
DR 3800
DR 2800
DR 2700
DR 5000 The fill line is toward the user.
DR 3900
DR 900 The orientation mark is toward the user. 2401906

Before starting
Install the instrument cap on the DR 900 cell holder before ZERO or READ is pushed.
The sample must be digested with heat and acid to make sure that all forms of the metal are measured. Use the mild or
vigorous digestion. Refer to the Water Analysis Guide for more information.
Clean all glassware with 6.0 N (50%) hydrochloric acid, then rinse thoroughly with deionized water to remove contaminants.
The sample temperature must be between 20–25 °C (68–77 °F) for accurate results.
The Pour-Thru Cell can be used (for applicable instruments) if rinsed well with deionized water between the blank and the
prepared samples.
For best results, measure the reagent blank value for each new lot of reagent. Replace the sample with deionized water in
the test procedure to get the reagent blank value. Subtract the reagent blank value from the sample results automatically
with the reagent blank adjust option.
Review the Safety Data Sheets (MSDS/SDS) for the chemicals that are used and use any recommended personal protective
equipment.
Dispose of reacted solutions according to local, state and federal regulations. Use the Safety Data Sheets for disposal
information for unused reagents. Consult the environmental, health and safety staff for your facility and/or local regulatory
agencies for further disposal information.

1
Items to collect
Description Quantity
®
AluVer 31 Aluminum Reagent Powder Pillow 1
Ascorbic Acid Powder Pillow 1
Bleaching 3 Reagent Powder Pillow 1
Cylinder, graduated mixing, 50-mL, with glass stopper 1
Sample cells. For information about sample cells, adapters or light shields, refer to Instrument
2
specific information on page 1.
1 AluVer is a registered trademark of Hach Company.

Refer to Consumables and replacement items on page 6 for reorder information.

Sample collection and storage

• Collect samples in clean glass or plastic bottles.
• To preserve samples for later analysis, adjust the sample pH to less than 2 with
concentrated nitric acid (about 2 mL per liter). No acid addition is necessary if the
sample is tested immediately.
• Keep preserved samples at room temperature for a maximum of 6 months.
• Before analysis, adjust the pH to 3.5–4.5 with 5.0 N sodium hydroxide standard
solution.
• Correct the test result for the dilution from the volume additions.

Powder pillow procedure

Start

1. Start program 2. Fill a graduated mixing 3. Add the contents of one 4. Close the cylinder. Invert
10 Aluminum Alumin. For cylinder to the 50-mL mark Ascorbic Acid Powder the cylinder several times to
information about sample with sample. Pillow. mix.
cells, adapters or light
shields, refer to Instrument
specific information
on page 1.
Note: Although the program
name may vary between
instruments, the program
number does not change.

2 Aluminum, Aluminon Method (0.800 mg/L)

5. Add one AluVer 6. Start the instrument 7. Invert the cylinder 8. Prepare the blank: Pour
3 Aluminum reagent powder timer. A 1-minute reaction repeatedly during the timer. 10 mL of the reacted sample
pillow. time starts. Undissolved powder will into a sample cell.
An orange to orange-red cause inconsistent results.
color shows if aluminum is
present.

9. Add one Bleaching 10. Start the instrument 11. Swirl the sample cell 12. Start the instrument
3 Reagent powder pillow to timer. A 30-second reaction vigorously. The solution will timer. A 15-minute reaction
the blank. time starts. show a light to medium time starts.
orange.

Zero

13. Prepare the sample: 14. When the timer expires, 15. Insert the blank into the 16. Push ZERO. The
Pour 10 mL of solution from clean the blank. cell holder. display shows 0.00 or
the cylinder into a second 0.000 mg/L Al3+.
sample cell.

Aluminum, Aluminon Method (0.800 mg/L) 3

 Read

17. Clean the prepared 18. Within five minutes after 19. Push READ. Results 20. Clean the graduated
sample. the timer expires, insert the show in mg/L Al3+. cylinder and sample cells
prepared sample into the with soapy water and a
cell holder. brush immediately after the
test. Rinse with deionized
water.

Interferences
Interfering Interference level
substance
Acidity More than 300 mg/L as CaCO3. Pre-treat samples that have more than 300 mg/L acidity as CaCO3 as
follows:

1. Add one drop of m-Nitrophenol Indicator Solution to 50 mL of fresh sample.

2. Add one drop of 5.0 N Sodium Hydroxide Standard Solution. Put the stopper on the cylinder. Invert
to mix. Repeat as often as necessary until the color changes from colorless to yellow.
3. Add one drop of 5.25 N Sulfuric Acid Standard Solution to change the solution from yellow to
colorless. Use this treated sample in the test procedure.

Alkalinity 1000 mg/L as CaCO3. Pre-treat samples that have higher alkalinity concentrations as follows:

1. Add one drop of m-Nitrophenol Indicator Solution to 50 mL of fresh sample. A yellow color indicates
excessive alkalinity.
2. Add one drop of 5.25 N Sulfuric Acid Standard Solution. Put the stopper on the cylinder. Invert to
mix. If the yellow color continues, repeat until the sample becomes colorless. Use this treated
sample in the test procedure.

Fluoride Interferes at all levels. Refer to Fluoride interference on page 4.

Iron More than 20 mg/L
Phosphate More than 50 mg/L
Polyphosphate Polyphosphate interferes at all levels and causes negative errors and must not be present. The sample
must be pre-treated with acid hydrolysis before the test is started to convert polyphosphate to
orthophosphate. Use the Phosphorus, Acid Hydrolyzable Digestion procedure.

Fluoride interference
Fluoride forms complexes with aluminum and interferes at all levels. If the fluoride
concentration of the sample is known, use Figure 1 to find the actual aluminum
concentration.
To use the fluoride interference graph:

1. On the top of the graph, find the aluminum result from the test procedure.
2. On the left side of the graph, find the fluoride concentration of the sample.
3. Go along the grid lines to find the point on the graph where the values intersect.
4. Go along the curved lines on either side of the intersect point to find the actual
aluminum concentration on the bottom of the graph.
Example: The aluminum test result is 0.7 mg/L Al3+ and the fluoride concentration is
1 mg/L F–. The 0.7 mg/L Al3+ grid line intersects with the 1 mg/L F– grid line between

4 Aluminum, Aluminon Method (0.800 mg/L)

 the 1.2 and 1.3 mg/L Al3+ curves. In this example, the actual aluminum concentration
is 1.27 mg/L.
Figure 1 Fluoride interference graph

1 mg/L F– in the sample 2 mg/L Al3+ from test procedure 3 Actual mg/L Al3+

Accuracy check
Standard additions method (sample spike)
Use the standard additions method (for applicable instruments) to validate the test
procedure, reagents and instrument and to find if there is an interference in the sample.
Items to collect:
• 50-mg/L Aluminum Voluette® Ampule Standard
• Ampule breaker
• Pipet, TenSette®, 0.1–1.0 mL and tips
• Mixing cylinders (3)

1. Use the test procedure to measure the concentration of the sample, then keep the
(unspiked) sample in the instrument.
2. Go to the Standard Additions option in the instrument menu.
3. Select the values for standard concentration, sample volume and spike volumes.
4. Open the standard solution.
5. Prepare three spiked samples: use the TenSette pipet to add 0.1 mL, 0.2 mL and
0.3 mL of the standard solution, respectively, to three 50-mL portions of fresh sample.
Mix well.
6. Use the test procedure to measure the concentration of each of the spiked samples.
Start with the smallest sample spike. Measure each of the spiked samples in the
instrument.
7. Select Graph to compare the expected results to the actual results.
Note: If the actual results are significantly different from the expected results, make sure that
the sample volumes and sample spikes are measured accurately. The sample volumes and
sample spikes that are used should agree with the selections in the standard additions menu. If
the results are not within acceptable limits, the sample may contain an interference.

Aluminum, Aluminon Method (0.800 mg/L) 5

Standard solution method
Use the standard solution method to validate the test procedure, reagents and
instrument.
Items to collect:
• 100-mg/L aluminum standard solution
• 250-mL volumetric flask, Class A
• 1.00-mL volumetric pipet, Class A and pipet filler
• Deionized water

1. Prepare a 0.4 mg/L aluminum standard solution as follows:

a. Use a pipet to add 1.00 mL of 100-mg/L aluminum standard solution into the
volumetric flask. (Alternate preparation: Use a pipet to add 0.8 mL of a 50-mg/L
aluminum standard solution into a 100-mL volumetric flask.)
b. Dilute to the mark with deionized water. Mix well. Prepare this solution daily.
2. Use the test procedure to measure the concentration of the prepared standard
solution.
3. Compare the expected result to the actual result.
Note: The factory calibration can be adjusted slightly with the standard adjust option so that the
instrument shows the expected value of the standard solution. The adjusted calibration is then
used for all test results. This adjustment can increase the test accuracy when there are slight
variations in the reagents or instruments.

Method performance
The method performance data that follows was derived from laboratory tests that were
measured on a spectrophotometer during ideal test conditions. Users may get different
results under different test conditions.
Program Standard Precision (95% Confidence Interval) Sensitivity
Concentration change per 0.010 Abs change
10 0.40 mg/L Al3+ 0.385–0.415 mg/L Al3+ 0.008 mg/L Al3+

Summary of method
Aluminon indicator combines with aluminum in the sample to form a red-orange color.
The intensity of color is proportional to the aluminum concentration. Ascorbic acid is
added before the AluVer 3 reagent to remove iron interference. To establish a reagent
blank, the sample is divided after the addition of the AluVer 3. Bleaching 3 Reagent is
then added to one-half of the divided sample to bleach out the color of the aluminum
aluminon complex. The AluVer 3 Aluminum Reagent, packaged in powder form, shows
exceptional stability and is applicable for fresh water applications. The measurement
wavelength is 522 nm for spectrophotometers or 520 nm for colorimeters.
Consumables and replacement items
Required reagents

Description Quantity/test Unit Item no.

Aluminum Reagent Set, includes: — 100 tests 2242000

®
AluVer 31 Aluminum Reagent Powder Pillow 1 100/pkg 1429099
Ascorbic Acid Powder Pillow 1 100/pkg 1457799
Bleaching 3 Reagent Powder Pillow 1 100/pkg 1429449
Hydrochloric Acid, 6.0 N varies 500 mL 88449
Water, deionized varies 4L 27256
1 AluVer is a registered trademark of Hach Company.

6 Aluminum, Aluminon Method (0.800 mg/L)

Required apparatus

Description Quantity/test Unit Item no.

Cylinder, graduated mixing, 50-mL, with glass stopper 1 each 189641

Recommended standards or Recommended standards and apparatus

Description Unit Item no.

Aluminum Standard Solution, 100-mg/L as Al3+ 100 mL 1417442

Aluminum Standard Solution, 10-mg/L as Al3+ 100 mL 2305842
®
Aluminum Standard Solution, 10-mL Voluette Ampule, 50-mg/L as Al 16/pkg 1479210
®
Ampule Breaker, Voluette ampules each 2196800

Optional reagents and apparatus

Description Unit Item no.

Flask, volumetric, Class A, 100-mL each 1457442

Flask, volumetric, Class A, 250-mL each 1457446
Liqui-Nox Phosphate-free detergent 946 mL 2088153
m-Nitrophenol Indicator Solution 100 mL 247632
Nitric Acid Solution, 1:1 500 mL 254049
Paper, pH, 0–14 pH range 100/pkg 2601300
®
Pipet, TenSette , 0.1–1.0 mL each 1970001
Pipet tips for TenSette Pipet 1970001 50/pkg 2185696
Sodium Hydroxide, 5 N 50 mL 245026
Sulfuric Acid Standard Solution, 5.25 N 100 mL 244932
Test tube brush each 69000

Aluminum, Aluminon Method (0.800 mg/L) 7

 FOR TECHNICAL ASSISTANCE, PRICE INFORMATION AND ORDERING: HACH COMPANY
In the U.S.A. – Call toll-free 800-227-4224 WORLD HEADQUARTERS
Outside the U.S.A. – Contact the HACH office or distributor serving you. Telephone: (970) 669-3050
On the Worldwide Web – www.hach.com; E-mail – [email protected] FAX: (970) 669-2932

© Hach Company/Hach Lange GmbH, 1989–2013. All rights reserved. 04/2013, Edition 8
Barium DOC316.53.01315

Turbidimetric Method1 Method 10251

2 to 100, 20 to 1000, 200 to 10,000 mg/L Ba (spectrophotometers) Powder Pillows
2 to 80, 20 to 800, 200 to 8000 mg/L Ba (colorimeters)
Scope and application: For oil and gas field waters.
1 Adapted from Snell and Snell, Colorimetric Methods of Analysis, Vol. II, 769 (1959).

Test preparation

Instrument specific information

The table in this section shows all of the instruments that have the program for this test.
Table 1 shows sample cell and orientation requirements for reagent addition tests, such
as powder pillow or bulk reagent tests.
To use the table, select an instrument, then read across to find the corresponding
information for this test.
Table 1 Instrument-specific information for reagent addition
Instrument Sample cell orientation Sample cell
DR 6000 The fill line is to the right. 2495402
DR 3800
DR 2800
DR 2700
DR 5000 The fill line is toward the user.
DR 3900
DR 900 The orientation mark is toward the user. 2401906

Before starting
Install the instrument cap on the DR 900 cell holder before ZERO or READ is pushed.
Use the Standard Adjust option with each new lot of reagent for the best results.
For best results, measure the reagent blank value for each new lot of reagent. Replace the sample with deionized water in
the test procedure to get the reagent blank value. Subtract the reagent blank value from the sample results automatically
with the reagent blank adjust option.
Filter samples that are turbid with filter paper and a funnel.
Do not use the Pour-Thru Cell or sipper module (for applicable instruments) with this test.
Review the Safety Data Sheets (MSDS/SDS) for the chemicals that are used and use any recommended personal protective
equipment.
Dispose of reacted solutions according to local, state and federal regulations. Use the Safety Data Sheets for disposal
information for unused reagents. Consult the environmental, health and safety staff for your facility and/or local regulatory
agencies for further disposal information.

1
Items to collect
Description Quantity

BariVer™ 4 Barium Reagent Powder Pillows 1

Sample cells (For information about sample cells, adapters or light shields, refer to Instrument
2
specific information on page 1.)

Refer to Consumables and replacement items on page 5 for reorder information.

Sample collection and storage

• Collect samples in clean glass or plastic bottles.
• To preserve samples for later analysis, adjust the sample pH to less than 2 with
concentrated nitric acid (about 2 mL per liter). No acid addition is necessary if the
sample is tested immediately.
• Keep preserved samples at room temperature for a maximum of 6 months.
• Before analysis, adjust the pH to 5 with 5.0 N sodium hydroxide standard solution.
• Correct the test result for the dilution from the volume additions.

Powder pillow procedure

Start

1. Start program 2. Prepare the blank: Add 3. If the sample volume is 4. Swirl to mix.
20 Barium. For information the sample volume that is less than 10 mL, add Refer to Set the dilution
about sample cells, specified for the test range deionized water to the 10- factor on page 3. A 10-
adapters or light shields, to a clean sample cell: mL line. mL graduated mixing
refer to Instrument specific cylinder can be used in
information on page 1. • 2–80 mg/L: 10 mL steps 2 and 3.
• 20–800 mg/L: 1.0 mL
Note: Although the program
name may vary between • 200–8000 mg/L: 0.1 mL
instruments, the program
Use a pipet to add the
number does not change.
1.0 mL and 0.1 mL volumes.

Zero

5. Clean the blank. 6. Insert the blank into the 7. Push ZERO. The display 8. Remove the sample from
cell holder. shows 0 mg/L Ba2+. the cell holder.

2 Barium, Turbidimetric Method (multi-range: 100, 1000, 10,000 mg/L)

9. Prepare the sample: 10. Swirl to mix. 11. Start the instrument 12. Clean the prepared
Add the contents of one The sample will become timer. A 5-minute reaction sample.
BariVer™ 4 Barium Reagent cloudy if barium is in the time starts.
Powder Pillow to the sample sample. Accuracy is not Do not move the sample cell
cell. affected by undissolved during the reaction period.
The solution will get cloudy powder.
if barium is in the sample.

Read

13. Within 5 minutes after 14. Push READ. Results 15. Clean the sample cell
the timer expires, insert the show in mg/L Ba2+. immediately after each test
prepared sample into the with soap, water and a
cell holder. brush.

Interferences
Interfering substance Interference level
Calcium 10,000 mg/L as CaCO3
Magnesium 100,000 mg/L as CaCO3
Silica 500 mg/L
Sodium Chloride 130,000 mg/L as NaCl
Strontium The interference level is dependent on the sample matrix and the barium concentration.
When the barium concentration is zero, there is no interference from strontium. The best
results occur when the barium concentration is less than 20 mg/L and when the strontium
concentration (as mg/L) is equal to or less than the barium concentration.
Highly buffered samples or Can prevent the pH adjustment by the reagent(s) and cause incorrect results.
extreme sample pH

Set the dilution factor

Instruments that have a dilution factor option can include the dilution factor in the result
and show the concentration of the original, undiluted sample. For example, if the sample
is diluted by a factor of 10, the instrument multiplies the result by 10 and shows the
calculated result in the instrument display.

1. Select Options>More>Dilution factor from the instrument menu.

Barium, Turbidimetric Method (multi-range: 100, 1000, 10,000 mg/L) 3

 Note: Colorimeters include a dilution factor when the chemical form is set. Go to
Options>Advanced Options>Chemical Form and select LR, MR or HR.
2. Enter the dilution factor:
• 1 mL sample diluted to 10 mL: dilution factor is 10.
• 0.1 mL sample diluted to 10 mL: dilution factor is 100.
3. Push OK to confirm. Push OK again.
4. Push RETURN to go back to the measurement screen.

Accuracy check
Standard additions method (sample spike)
Use the standard additions method (for applicable instruments) to validate the test
procedure, reagents and instrument and to find if there is an interference in the sample.
Items to collect:
• Barium Standard Solution, 1000-mg/L Ba
• Pipet, TenSette®, 0.1–1.0 mL
• Pipet tips

1. Use the test procedure to measure the concentration of the sample, then keep the
(unspiked) sample in the instrument.
2. Go to the Standard Additions option in the instrument menu.
3. Select the values for standard concentration, sample volume and spike volumes.
4. Open the standard solution.
5. Prepare three spiked samples: use the TenSette pipet to add of the standard solution,
respectively, to three 10-mL portions of fresh sample. Mix well.
6. Use the test procedure to measure the concentration of each of the spiked samples.
Start with the smallest sample spike. Measure each of the spiked samples in the
instrument.
7. Select Graph to compare the expected results to the actual results.
Note: If the actual results are significantly different from the expected results, make sure that
the sample volumes and sample spikes are measured accurately. The sample volumes and
sample spikes that are used should agree with the selections in the standard additions menu. If
the results are not within acceptable limits, the sample may contain an interference.

Standard solution method

Use the standard solution method to validate the test procedure, reagents and
instrument.
Items to collect:
• Barium Standard Solution, 1000-mg/L Ba
• 100-mL volumetric flask, Class A
• 5-mL volumetric pipet, Class A and pipet filler
• Deionized water

1. Prepare a 50.0-mg/L barium standard solution as follows:

a. Use a pipet to add 5.00 mL of 1000-mg/L barium standard solution into the
volumetric flask.
b. Dilute to the mark with deionized water. Mix well. Prepare this solution daily.
2. Use the test procedure to measure the concentration of the prepared standard
solution.
3. Compare the expected result to the actual result.
Note: The factory calibration can be adjusted slightly with the standard adjust option so that the
instrument shows the expected value of the standard solution. The adjusted calibration is then
used for all test results. This adjustment can increase the test accuracy when there are slight
variations in the reagents or instruments.

4 Barium, Turbidimetric Method (multi-range: 100, 1000, 10,000 mg/L)

Method performance
The method performance data that follows was derived from laboratory tests that were
measured on a spectrophotometer during ideal test conditions. Users may get different
results under different test conditions.
Program Standard Precision (95% Confidence Interval) Sensitivity
Concentration change per 0.010 Abs change
20 30 mg/L Ba 25–35 mg/L Ba 1 mg/L Ba

Summary of method
The BariVer™ 4 Barium Reagent Powder combines with barium to form a barium sulfate
precipitate, which is held in suspension by a protective colloid. The amount of precipitate
is proportional to the barium concentration. The measurement wavelength is 450 nm for
spectrophotometers or 520 for colorimeters.
Consumables and replacement items
Required reagents

Description Quantity/test Unit Item no.

BariVer™ 4 Barium Reagent Powder Pillows 1 100/pkg 1206499

Recommended standards

Description Unit Item no.

Barium Standard Solution, 1000-mg/L Ba 100 mL 1461142

Water, deionized 4L 27256

Optional reagents and apparatus

Description Unit Item no.

Brush, test tube each 69000

Filter paper, 12.5-cm 100/pkg 189457
Flask, volumetric, Class A, 100-mL each 1457442
Funnel, poly, 65-mm each 108367
Liqui-Nox Phosphate-free detergent 946 mL 2088153
Nitric Acid Solution, 1:1 500 mL 254049
Paper, pH, 0–14 pH range 100/pkg 2601300
®
Pipet, TenSette , 0.1–1.0 mL each 1970001
Pipet tips for TenSette Pipet 1970001 50/pkg 2185696
Pipet tips for TenSette Pipet 1970001 1000/pkg 2185628
Pipet, volumetric 5.00-mL each 1451537
Pipet filler, safety bulb each 1465100
100 mL
Sodium Hydroxide Standard Solution, 5.0 N 245032
MDB

Barium, Turbidimetric Method (multi-range: 100, 1000, 10,000 mg/L) 5

 FOR TECHNICAL ASSISTANCE, PRICE INFORMATION AND ORDERING: HACH COMPANY
In the U.S.A. – Call toll-free 800-227-4224 WORLD HEADQUARTERS
Outside the U.S.A. – Contact the HACH office or distributor serving you. Telephone: (970) 669-3050
On the Worldwide Web – www.hach.com; E-mail – [email protected] FAX: (970) 669-2932

© Hach Company/Hach Lange GmbH, 1989–2013. All rights reserved. 04/2013, Edition 8
Benzotriazole/Tolyltriazole DOC316.53.01008

UV Photolysis Method1 Method 8079

1.0 to 16.0 mg/L Benzotriazole Powder Pillows
1.0 to 20.0 mg/L Tolyltriazole
Scope and application: For cooling or boiler water.
1 Adapted from Harp, D., Proceedings 45th International Water Conference, 299 (October 22-24, 1984)

Test preparation

Instrument specific information

The table in this section shows all of the instruments that have the program for this test.
Table 1 shows sample cell and orientation requirements for reagent addition tests, such
as powder pillow or bulk reagent tests.
To use the table, select an instrument, then read across to find the corresponding
information for this test.
Table 1 Instrument-specific information for powder pillows
Instrument Sample cell orientation Sample cell
DR 6000 The fill line is to the right. 2495402
DR 3800
DR 2800
DR 2700
DR 5000 The fill line is toward the user.
DR 3900
DR 900 The orientation mark is toward the user. 2401906

Before starting
Install the instrument cap on the DR 900 cell holder before ZERO or READ is pushed.
Wear UV safety goggles while the UV lamp is on.
Do not touch the UV lamp surface with bare fingers. Fingerprints can damage the glass. Rinse the lamp and wipe with a soft,
clean tissue between tests.
The sample temperature must be between 20–25 °C (68–77 °F) for accurate results.
Review the Safety Data Sheets (MSDS/SDS) for the chemicals that are used and use any recommended personal protective
equipment.
Dispose of reacted solutions according to local, state and federal regulations. Use the Safety Data Sheets for disposal
information for unused reagents. Consult the environmental, health and safety staff for your facility and/or local regulatory
agencies for further disposal information.

1
Items to collect
Description Quantity

Triazole Reagent Powder Pillows 1

Square mixing bottle, 25-mL 1
Ultra-violet lamp with power supply 1
UV safety goggles 1
Sample cells (For information about sample cells, adapters or light shields, refer to Instrument
2
specific information on page 1.)

Refer to Consumables and replacement items on page 5 for reorder information.

Sample collection
• Collect samples in clean glass or plastic bottles.
• Analyze the samples as soon as possible for best results.

Powder pillow procedure with UV photolysis

Start

1. Start program 2. Prepare the sample: Fill 3. Add the contents of one 4. Swirl to mix.
30 Benzotriazole or a marked mixing bottle to Triazole Reagent Powder Make sure all of the powder
program 730 Tolyltriazole. the 25-mL line with sample. Pillow. is dissolved.
For information about
sample cells, adapters or
light shields, refer to
Instrument specific
information on page 1.
Note: Although the program
name may vary between
instruments, the program
number does not change.

2 Benzotriazole/Tolyltriazole, UV Photolysis Method (Benzo 16.0 mg/L, Tolyl 20 mg/L)

5. Put on the UV safety 6. Put the ultraviolet lamp 7. Start the instrument 8. When the timer expires,
goggles. into the mixing bottle. Turn timer. A 5-minute reaction turn off the lamp. Remove
on the UV lamp. time starts. the lamp from the mixing
Low results will occur if UV bottle. Swirl the mixing
photolysis is more than or bottle to mix thoroughly.
less than 5 minutes. A
yellow color will show if
triazole is in the sample.

9. Fill a sample cell with 10. Prepare the blank: Fill 11. Clean the blank. 12. Insert the blank into the
10 mL of the reacted a second sample cell with cell holder.
(prepared) sample. 10 mL of unreacted sample.

Zero Read

13. Push ZERO. The 14. Clean the prepared 15. Insert the prepared 16. Push READ. Results
display shows 0.0 mg/L sample. sample into the cell holder. show in mg/L Benzotriazole
Benzotriazole or or Tolyltriazole.
Tolyltriazole.

Interferences
Interfering substance Interference level
Acrylates (as methyl acrylate) More than 50 mg/L
Alum More than 400 mg/L
Borate (as sodium tetraborate (borax)) More than 4000 mg/L. Adjust the sample pH to 4–6 with 1 N sulfuric acid, then start
the test procedure.
Chlorine (as Cl2) More than 20 mg/L

Benzotriazole/Tolyltriazole, UV Photolysis Method (Benzo 16.0 mg/L, Tolyl 20 mg/L) 3

Interfering substance Interference level
Chromium (as chromate) More than 12 mg/L
Copper More than 10 mg/L
Hardness More than 500 mg/L as CaCO3. Add 10 drops of Rochelle Salt Solution before the
reagent is added.
Iron More than 20 mg/L
Lignosulfonates More than 40 mg/L
Magnesium More than 300 mg/L as CaCO3
Molybdenum (as molybdate) More than 200 mg/L
Nitrite More than 4000 mg/L. Adjust the sample pH to 4–6 with 1 N sulfuric acid, then start
the test procedure.
Phosphonates (AMP or HEDP) More than 100 mg/L
Sulfate More than 200 mg/L
Zinc More than 80 mg/L
Strong oxidizing or reducing agents Interfere at all levels

Accuracy check
Standard additions method (sample spike)
Use the standard additions method (for applicable instruments) to validate the test
procedure, reagents and instrument and to find if there is an interference in the sample.
Items to collect:
• Benzotriazole Standard Solution, 500 mg/L
®
• Pipet, TenSette , 0.1–1.0 mL
®
• Pipet tips for TenSette Pipet, 0.1–1.0 mL

1. Use the test procedure to measure the concentration of the sample, then keep the
(unspiked) sample in the instrument.
2. Go to the Standard Additions option in the instrument menu.
3. Select the values for standard concentration, sample volume and spike volumes.
4. Open the standard solution.
5. Prepare three spiked samples: use the TenSette pipet to add 0.1 mL, 0.2 mL and
0.3 mL of the standard solution, respectively, to three 25-mL portions of fresh sample.
Mix well.
6. Use the test procedure to measure the concentration of each of the spiked samples.
Start with the smallest sample spike. Measure each of the spiked samples in the
instrument.
7. Select Graph to compare the expected results to the actual results.
Note: If the actual results are significantly different from the expected results, make sure that
the sample volumes and sample spikes are measured accurately. The sample volumes and
sample spikes that are used should agree with the selections in the standard additions menu. If
the results are not within acceptable limits, the sample may contain an interference.

Standard solution method

Complete this procedure to make sure that the UV lamp operates correctly. If the result is
significantly low, replace the lamp. The normal UV lamp life is approximately 5000 hours.
Items to collect:
• Benzotriazole Standard Solution, 500 mg/L
• 1-L volumetric flask, Class A
• 5-mL volumetric pipet, Class A and pipet filler

4 Benzotriazole/Tolyltriazole, UV Photolysis Method (Benzo 16.0 mg/L, Tolyl 20 mg/L)

 • Deionized water

1. Prepare a 5.0 mg/L benzotriazole standard solution as follows:

a. Use a pipet to add 10.0 mL of 500-mg/L benzotriazole standard solution into the
volumetric flask.
b. Dilute to the mark with deionized water. Mix well. Prepare this solution daily.
2. Use the test procedure to measure the concentration of the prepared standard
solution.
3. Compare the expected result to the actual result.
Note: If the test result is significantly below 5 mg/L, replace the lamp.

Method performance
The method performance data that follows was derived from laboratory tests that were
measured on a spectrophotometer during ideal test conditions. Users may get different
results under different test conditions.
Program Standard Precision (95% Confidence Interval) Sensitivity
Concentration change per 0.010 Abs change
30 10 mg/L benzotriazole 9.7–10.3 benzotriazole 0.2 mg/L benzotriazole
730 12 mg/L tolyltriazole 11.6-12.4 mg/L tolyltriazole 0.2 mg/L tolyltriazole

Summary of method
Benzotriazole or tolyltriazole, used in many applications as corrosion inhibitors for copper
and copper alloys, are determined by a proprietary catalytic ultraviolet (UV) photolysis
procedure requiring less than 10 minutes to perform. The measurement wavelength is
425 nm for spectrophotometers or 420 nm for colorimeters.
Consumables and replacement items
Required reagents

Description Quantity/test Unit Item no.

Triazole Reagent Powder Pillow 1 100/pkg 2141299

Required apparatus

Description Quantity/test Unit Item no.

UV safety goggles 1 each 2113400

Bottle, square, with 25 mL mark 1 each 1704200
Select one based on available voltage:
Lamp kit, UV, with power supply, 115 VAC, 60 Hz 1 each 2704500
Lamp kit, UV, with power supply, 230 VAC, 50 Hz 1 each 2704502

Recommended standards

Description Unit Item no.

Benzotriazole Standard Solution, 500-mg/L 100 mL 2141342

Benzotriazole/Tolyltriazole, UV Photolysis Method (Benzo 16.0 mg/L, Tolyl 20 mg/L) 5

Optional reagents and apparatus

Description Unit Item no.

Flask, volumetric, Class A, 1000-mL each 1457453

Paper, pH, 0–14 pH range 100/pkg 2601300
Pipet filler, safety bulb each 1465100
®
Pipet, TenSette , 0.1–1.0 mL each 1970001
Pipet tips for TenSette Pipet 1970001 50/pkg 2185696
Pipet tips for TenSette Pipet 1970001 1000/pkg 2185628
100 mL
Sulfuric Acid Standard Solution, 1 N 127032
MDB
Rochelle Salt Solution 29 mL 172533
Pipet, volumetric, Class A, 10-mL each 1451538

FOR TECHNICAL ASSISTANCE, PRICE INFORMATION AND ORDERING: HACH COMPANY

In the U.S.A. – Call toll-free 800-227-4224 WORLD HEADQUARTERS
Outside the U.S.A. – Contact the HACH office or distributor serving you. Telephone: (970) 669-3050
On the Worldwide Web – www.hach.com; E-mail – [email protected] FAX: (970) 669-2932

© Hach Company/Hach Lange GmbH, 1989–2013. All rights reserved. 04/2013, Edition 8
Boron DOC316.53.01313

Carmine Method Method 10252

2 to 50 mg/L B Powder Pillows
Scope and application: For oil and gas field waters

Test preparation

Instrument specific information

The table in this section shows all of the instruments that have the program for this test.
Table 1 shows adapter and light shield requirements for the instruments that use them.
To use the table, select an instrument, then read across to find the corresponding
information for this test.
Table 1 Instrument-specific information for test tubes
Instrument Adapters Light shield
DR 6000 — —
DR 5000 — —
DR 900 4846400 Cover supplied with the instrument
DR 3900 — LZV849
DR 3800 — LZV646
DR 2800 —

Before starting
Install the instrument cap on the DR 900 cell holder before ZERO or READ is pushed.
DR 3900, DR 3800, DR 2800 and DR 2700: Install the light shield in Cell Compartment #2 before this test is started.
The reagent that is used in this test is corrosive. Use protection for eyes and skin and be prepared to flush any spills with
running water.
Review the Safety Data Sheets (MSDS/SDS) for the chemicals that are used and use any recommended personal protective
equipment.
Dispose of reacted solutions according to local, state and federal regulations. Use the Safety Data Sheets for disposal
information for unused reagents. Consult the environmental, health and safety staff for your facility and/or local regulatory
agencies for further disposal information.

Items to collect
Description Quantity

BoroVer 3 Reagent Powder Pillow 1

Sufuric Acid, Concentrated 75 mL
Water, deionized 2 mL
Tubes, glass, 16 mm x 100 mm 2
Caps, white Teflon 2
Flask, poly, screw-on cap, 250-mL 1
Cylinder, graduated, poly, 100-mL 1
Light shield or adapter (For information about sample cells, adapters or light shields, refer to
1
Instrument specific information on page 1.)

1
Items to collect (continued)
Description Quantity

Select:
Pipet, 0.2 - 1.0 mL , BBP078 1
Pipet Tip, for BBP078 2
Pipet, 1.0 - 1.0 mL, BBP065 1
Pipet Tip for BBP065 2
OR
Pipet, TenSette, 0.1- to 1.0-mL 1
Pipet tips for 0.1- to 1.0-mL TenSette 2
Pipet, TenSette, 1.0- to 10.0-mL 1
Pipet tips for 1.0- to 10.0-mL TenSette 2

Refer to Consumables and replacement items on page 6 for reorder information.

Sample collection
Collect samples in clean polyethylene or polypropylene bottles.
Prepare the glass tubes for first use
New glass tubes can contain residual amounts of reactive boron from the glass
manufacturing process. For best results, precondition the tubes before the first use.
Previously used tubes do not need to be preconditioned.

1. Prepare the BoroVer 3/Sulfuric Acid Solution.

2. Add 3 to 4 mL of the prepared BoroVer 3/Sulfuric Acid Solution into the tubes.
3. After 30 minutes, discard the solution.
4. Rinse and dry the tubes before use.

Powder pillow procedure

Start

1. Start Boron HR. Refer to 2. Use a 100-mL graduated 3. In a well-ventilated area 4. Swirl the flask
Instrument setup cylinder to measure 75 mL or fume hood, add the immediately to mix. Swirl for
on page 5 for of concentrated sulfuric contents of one BoroVer up to 5 minutes to dissolve
programming instructions. acid. Pour the acid into a 3 Reagent Powder Pillow to the powder completely.
For information about plastic 250-mL Erlenmeyer the flask.
sample cells, adapters or flask.
light shields, refer to
Instrument specific
information on page 1.
Note: Although the program
name may vary between
instruments, the program
number does not change.

2 Boron, Carmine method (50 mg/L)

5. Prepare the blank: 6. Prepare the sample: 7. Add 3.5 mL of the 8. Put the cap on the
Remove the cap from a Remove the cap from a BoroVer 3 Solution from prepared sample and invert
clean 16-mm tube. Add clean 16-mm tube. Add step 4 to the prepared to mix.
0.2 mL of deionized water. 0.2 mL of sample. sample tube. The solution in the tube will
Refer to Prepare the glass Note: If a 3.5-mL pipet is Note: If a 3.5-mL pipet is get warm.
tubes for first use on page 2 not available, 0.4 mL of not available, 7.0 mL of
and Clean the glass tubes sample can be used. BoroVer 3 Solution can be
after use on page 4. used with 0.4 mL of sample.
Note: If a 3.5-mL pipet is
not available, 0.4 mL of DI
water can be used.

9. Add 3.5 mL of the 10. Put the cap on the 11. Start the instrument 12. When the timer expires,
BoroVer 3 Solution from blank. Iinvert to mix. timer. A 30-minute reaction clean the blank.
step 4 to the blank sample The solution in the tube will time starts.
tube. get warm.
Note: If a 3.5-mL pipet is
not available, 7.0 mL of
BoroVer 3 Solution can be
used with 0.4 mL of
deionized water.

Zero

13. Insert the blank into the 14. Push ZERO. The 15. Clean the prepared 16. Insert the prepared
cell holder. display shows 0.0 mg/L B sample. sample into the cell holder.
HR.

Boron, Carmine method (50 mg/L) 3

 Read

17. Push READ. Results

show in mg/L B.

Reagent preparation
To prepare additional BoroVer 3/Sulfuric Acid Solution, mix one BoroVer 3 Reagent
Powder Pillow per 75 mL of concentrated sulfuric acid.
Preparation notes
• Gaseous hydrochloric acid (HCl) forms when the powder pillow is added to sulfuric
acid. Always mix under a fume hood.
• The solution is stable for up to 48 hours when it is stored in plastic containers.
• To prevent boron contamination from the glassware, do not keep the solution in
® ®
borosilicate glassware (Pyrex or Kimax ) for more than 1 hour.
• The BoroVer 3/Sulfuric Acid Solution is highly acidic. Refer to the current MSDS/SDS
for safe handling and disposal instructions.

1. Under a fume hood, measure the concentrated sulfuric acid with a graduated
cylinder.
2. Pour the acid into a Erlenmeyer flask.
3. Stir the acid and add the contents of one BoroVer 3 Reagent Powder Pillow to the
flask. Swirl to mix. Allow up to 5 minutes for the powder to completely dissolve. Add
one powder pillow at a time and stir to dissolve after each powder pillow is added.
4. Pour this solution into plastic containers.

Clean the glass tubes after use

NOTICE
The BoroVer 3/Sulfuric Acid solution is highly acidic. Neutralize the solution to pH 6–9 before
disposal. Refer to a current SDS (Safety Data Sheet) for safe handling and disposal instructions of
reacted boron.

Glass tubes and caps can be reused.

1. Thoroughly drain the boron solution.

2. Rinse the vials several times with deionized water.
3. Let the vials dry completely before the next use.

Accuracy check
Standard solution method
Use the standard solution method to validate the test procedure, reagents and
instrument.
Items to collect:
• 1000 mg/L Boron Standard Solution
• 100-mL volumetric flask, Class A
• 3-mL volumetric pipet, Class A and pipet filler

4 Boron, Carmine method (50 mg/L)

 • Deionized water

1. Prepare a 30.0 mg/L boron standard solution as follows:

a. Use a pipet to add 3.0 mL of 1000 mg/L boron standard solution into the
volumetric flask.
b. Dilute to the mark with deionized water. Mix well. Prepare this solution daily.
2. Use the test procedure to measure the concentration of the prepared standard
solution.
3. Compare the expected result to the actual result.
Note: The factory calibration can be adjusted slightly with the standard adjust option so that the
instrument shows the expected value of the standard solution. The adjusted calibration is then
used for all test results. This adjustment can increase the test accuracy when there are slight
variations in the reagents or instruments.

Method performance
The method performance data that follows was derived from laboratory tests that were
measured on a spectrophotometer during ideal test conditions. Users may get different
results under different test conditions.
Program Standard Precision (95% Confidence Interval) Sensitivity
Concentration change per 0.010 Abs change
As entered 25 mg/L B 24.2–25.8 mg/L B 2.2 mg/L B

Instrument setup
Some spectrophotometers do not have the Boron HR method in the stored programs.
List. Use the instructions in this section to add the test.

1. Set the instrument power to on. The instrument will complete a self-check.
2. From the Main Menu, go to User Programs>Program Options>New>Program
Number.
3. Enter a program number. Push OK.
4. Enter Boron_HR for the program name. Push Next.
5. Set the program type to Single Wavelength. Push Next.
6. Set the units to mg/L. Push Next.
7. Enter 605 for the wavelength. Push Next.
8. Enter 1 for the concentration resolution. Push Next.
9. Enter B for the chemical form. Push Next.
10. Select Enter Formula for Calibration. Push Next.
11. Enter: C = a + bA.
12. Enter: C = a + bA = cA2. Push OK.
13. Enter the values for the equations.
Option Description
DR 2700 a = 0.0, b= 25.51, c = 4.12
DR 2800 a = 0.0, b= 25.51, c = 4.12
DR 3800 a = 0.0, b= 25.51, c = 4.12
DR 3900 a = 0.0, b= 25.51, c = 4.12
DR 6000 a = 0.0, b= 25.51, c = 4.12
DR 5000 a = 0.0, b = 25.02, c = 4.22
14. Push OK, then Done.
15. In the User Program for Number Assigned screen, enter Upper Limit. Select EDIT.
16. Enter ON>2. Push OK twice.

Boron, Carmine method (50 mg/L) 5

 17. In the User Program for Number Assigned screen, enter Timer1. Select EDIT.
18. Enter 1>30.00. Push OK twice.
19. In the User Program for Number Assigned screen, enter Store.
20. Record the User Program Number Assigned.

Summary of method
Boron is determined by its reaction with carminic acid in the presence of sulfuric acid to
produce a reddish to bluish color. The amount of color is directly proportional to the boron
concentration. The measurement wavelength is 605 nm for spectrophotometers or
610 nm for colorimeters.
Consumables and replacement items
Required reagents

Description Quantity/test Unit Item no.

®
BoronVer 3 Boron Reagent Powder Pillows 1pillow/10 tests 100/pkg 1417099
Sulfuric Acid, concentrated, ACS varies 500 mL 97949
Water, deionized varies 100 mL 27242

Required apparatus

Description Quantity/test Unit Item no.

Tubes, glass, 16 mm x 100 mm 1 6/pkg 2275806

Caps, white, Teflon lining, for 16-mm tubes 2 6/pkg 2241106
Cylinder, graduated, polypropylene, 100 mL 1 each 108142
Flask, Polymethylpentene, screw cap, 250 mL, 125 mL 1 each 2089846
Pipet, 0.2 –1.0 mL 1 each BBP078
Pipet Tip for BBP078 2 100/pkg BBP079
Pipet, 1.0–5.0 mL 1 each BBP065
Pipet Tip for BBP065 1 75/pkg BBP068
OR
®
Pipet, TenSette , 0.1- to 1.0-mL 1 each 1970001
Pipet Tips, for TenSette Pipet 1970001 2 50/pkg 2185696
Pipet, TenSette 1.0–10.0 mL 1 each 1970010
Pipet Tips, for TenSette Pipet 1970010 varies 50/pkg 2199796
Tubes, glass, 16 mm x 100 mm 1 6/pkg 2275806

Optional reagents

Description Unit Item no.

Boron Standard Solution, 1000 mg/L as B 100 mL 191442

Optional apparatus

Description Unit Item no.

Gloves, chemical resistant, size 10 pair 2410105

Goggles, safety, standard each 2927902
Pipet tips for TenSette Pipet 1970001 1000/pkg 2185628

6 Boron, Carmine method (50 mg/L)

Consumables and replacement items (continued)
Description Unit Item no.

Pipet tips for TenSette Pipet 1970010 250/pkg 2199725

Test tube rack each 1864100
Pipets, includes one BBP078 and BBP065 pipet plus tips each LZP320

Boron, Carmine method (50 mg/L) 7

 FOR TECHNICAL ASSISTANCE, PRICE INFORMATION AND ORDERING: HACH COMPANY
In the U.S.A. – Call toll-free 800-227-4224 WORLD HEADQUARTERS
Outside the U.S.A. – Contact the HACH office or distributor serving you. Telephone: (970) 669-3050
On the Worldwide Web – www.hach.com; E-mail – [email protected] FAX: (970) 669-2932

© Hach Company/Hach Lange GmbH, 1989–2013. All rights reserved. 04/2013, Edition 8
Bromine DOC316.53.01011

DPD Method1 Method 8016

®
0.05 to 4.50 mg/L Br2 Powder Pillows or AccuVac Ampuls
Scope and application: For testing bromine residuals (including hypobromite, hypobromous acid and
bromamines) used as disinfectants in process waters, treated water, estuary water and seawater.
1 Adapted from Standard Methods for the Examination of Water and Wastewater.

Test preparation

Instrument-specific information
The tables in this section show all of the instruments that have the program for this test.
Table 1 shows sample cell and orientation requirements for reagent addition tests, such
as powder pillow or bulk reagent tests. Table 2 shows sample cell and adapter
requirements for AccuVac Ampul tests.
To use either table, select an instrument, then read across to find the corresponding
information for this test.
Table 1 Instrument-specific information for powder pillows
Instrument Sample cell orientation Sample cell
DR 6000 The fill line is to the right. 2495402
DR 3800
DR 2800
DR 2700
DR 5000 The fill line is toward the user.
DR 3900
DR 900 The orientation mark is toward the user. 2401906

Table 2 Instrument-specific information for AccuVac Ampuls

Instrument Adapter Sample cell
DR 6000 — 2427606
DR 5000
DR 900
DR 3900 LZV846 (A)
DR 3800 LZV584 (C) 2122800
DR 2800
DR 2700

Before starting
Samples must be analyzed immediately after collection and cannot be preserved for later analysis.
Install the instrument cap on the DR 900 cell holder before ZERO or READ is pushed.

1
For best results, measure the reagent blank value for each new lot of reagent. Replace the sample with deionized water in
the test procedure to get the reagent blank value. Subtract the reagent blank value from the sample results automatically
with the reagent blank adjust option.
Review the Safety Data Sheets (MSDS/SDS) for the chemicals that are used and use any recommended personal protective
equipment.
Dispose of reacted solutions according to local, state and federal regulations. Use the Safety Data Sheets for disposal
information for unused reagents. Consult the environmental, health and safety staff for your facility and/or local regulatory
agencies for further disposal information.

Items to collect
Powder pillows

Description Quantity

DPD Total Chlorine Reagent Powder Pillow, 10-mL 1

Sample cells. (For information about sample cells, adapters or light shields, refer to Instrument-
2
specific information on page 1.)

Refer to
Consumable and replacement items on page 6
for reorder information.

AccuVac Ampuls

Description Quantity
®
DPD Total Chlorine Reagent AccuVac Ampul 1
Beaker, 50-mL 1
Sample cells (For information about sample cells, adapters or light shields, refer to Instrument-
1
specific information on page 1.)
Stopper for 18-mm tubes and AccuVac Ampuls 1

Refer to
Consumable and replacement items on page 6
for reorder information.

Sample collection
• Analyze samples for bromine immediately after collection.
• Bromine is a strong oxidizing agent and it is unstable in natural waters. It reacts
quickly with various inorganic compounds and more slowly with organic compounds.
Many factors, including reactant concentrations, sunlight, pH, temperature and
salinity influence the decomposition of bromine in water.
• Collect samples in clean glass bottles. Avoid plastic containers since these may have
a large bromine demand.
• Pre-treat glass sample containers to remove any bromine demand. Soak the
containers in a dilute bleach solution (1 mL commercial bleach to 1 liter of deionized
water) for at least 1 hour. Rinse thoroughly with deionized or distilled water. If sample
containers are rinsed thoroughly with deionized or distilled water after use, only
occasional pre-treatment is necessary.
• Be sure to get a representative sample. If the sample is taken from a spigot or faucet,
let the water flow for at least 5 minutes. Then let the container overflow with the
sample several times and then put the cap on the sample container so that there is
no headspace (air) above the sample. If a sample cell is used, rinse the cell several
times with the sample, then carefully fill to the 10-mL mark.

2 Bromine, DPD Method (4.50 mg/L)

Powder pillow procedure

Start

1. Start program 2. Fill a sample cell with 3. Prepare the sample: 4. Swirl the sample cell for
50 Bromine. For 10 mL of sample. Add the contents of one 20 seconds to mix.
information about sample powder pillow to the sample A pink color shows if
cells, adapters or light cell. bromine is in the sample.
shields, refer to Instrument-
specific information
on page 1.
Note: Although the program
name may vary between
instruments, the program
number does not change.

5. Start the instrument 6. Prepare the blank: Fill a 7. Clean the blank. 8. Insert the blank into the
timer. A 3-minute reaction second sample cell with cell holder.
time starts. 10 mL of sample.
The instrument can be set to
zero with the blank during
the reaction timer.

Zero Read

9. Push ZERO. The display 10. Clean the prepared 11. Within 3 minutes after 12. Push READ. Results
shows 0.00 mg/L Br2. sample. the timer expires, insert the show in mg/L Br2.
prepared sample into the
cell holder.

Bromine, DPD Method (4.50 mg/L) 3

AccuVac Ampul procedure

Start

1. Start program 2. Prepare the blank: Fill 3. Prepare the sample: 4. Quickly invert the Ampul
55 Bromine AV. For the sample cell with 10 mL Collect at least 40 mL of several times to mix.
information about sample of sample. sample in a 50-mL beaker. A pink color shows if
cells, adapters or light Fill the AccuVac Ampul with bromine is in the sample.
shields, refer to Instrument- sample. Keep the tip
specific information immersed while the Ampul
on page 1. fills completely.
Note: Although the program
name may vary between
instruments, the program
number does not change.

Zero

5. Start the instrument 6. Clean the blank. 7. Insert the blank into the 8. Push ZERO. The display
timer. A 3-minute reaction cell holder. shows 0.00 mg/L Br2.
time starts.
The instrument can be set to
zero with the blank during
the reaction timer.

Read

9. Clean the AccuVac 10. Within 3 minutes after 11. Push READ. Results
Ampul. the timer expires, insert the show in mg/L Br2.
prepared sample AccuVac
Ampul into the cell holder.

4 Bromine, DPD Method (4.50 mg/L)

Interferences
Interfering substance Interference level
Acidity More than 150 mg/L CaCO3. The full color may not develop or the color may fade instantly.
Neutralize to pH 6–7 with 1 N Sodium Hydroxide. Measure the amount to be added on a
separate sample aliquot, then add the same amount to the sample that is tested. Correct the
test result for the dilution from the volume addition.
Alkalinity More than 250 mg/L CaCO3. The full color may not develop or the color may fade instantly.
Neutralize to pH 6–7 with 1 N Sulfuric Acid. Measure the amount to add on a separate sample
aliquot, then add the same amount to the sample that is tested. Correct the test result for the
dilution from the volume addition.
Chlorine Interferes at all levels
Chlorine Dioxide Interferes at all levels
Chloramines, organic May interfere
Hardness No effect at less than 1000 mg/L as CaCO3
Iodine Interferes at all levels
Manganese, Oxidized Pre-treat the sample as follows:
(Mn4+, Mn7+) or Chromium,
Oxidized (Cr6+) 1. Adjust the sample pH to 6–7.
2. Add 3 drops of Potassium Iodide (30-g/L) to 25 mL of sample.
3. Mix and wait 1 minute.
4. Add 3 drops of Sodium Arsenite (5-g/L) and mix.
5. Use the test procedure to measure the concentration of 10 mL of the treated sample.
6. Subtract this result from the result without the treatment to obtain the correct bromine
concentration.

Monochloramine Interferes at all levels

Ozone Interferes at all levels
Peroxides May interfere
Highly buffered samples or Can prevent the correct pH adjustment of the sample by the reagents. Sample pretreatment
extreme sample pH may be necessary. Adjust to pH 6–7 with acid (Sulfuric Acid, 1.000 N) or base (Sodium
Hydroxide, 1.00 N).

Pollution prevention and waste management

If sodium arsenite was added to the sample for manganese or chromium interferences,
the reacted samples will contain arsenic and must be disposed of as a hazardous waste.
Dispose of reacted solutions according to local, state and federal regulations.
Method performance
The method performance data that follows was derived from laboratory tests that were
measured on a spectrophotometer during ideal test conditions. Users may get different
results under different test conditions.
Program Standard Precision (95% Confidence Interval) Sensitivity
Concentration change per 0.010 Abs change
50 2.80 mg/L Br2 2.75–2.85 mg/L Br2 0.05 mg/L Br2
55 2.80 mg/L Br2 2.71–2.89 mg/L Br2 0.05 mg/L Br2

Summary of method
Bromine residuals reacts with DPD (N,N-diethyl-p-phenylenediamine) to form a pink color
which is proportional to the total bromine concentration. The measurement wavelength is
530 nm for spectrophotometers or 520 nm for colorimeters.

Bromine, DPD Method (4.50 mg/L) 5

Consumable and replacement items
Required reagents

Description Quantity/Test Unit Item no.

DPD Total Chlorine Reagent Powder Pillow, 10-mL 1 100/pkg 2105669

OR
®
DPD Total Chlorine Reagent AccuVac Ampul 1 25/pkg 2503025

Required apparatus

Description Quantity/Test Unit Item no.

AccuVac Snapper 1 each 2405200

Beaker, 50-mL 1 each 50041H
Stoppers for 18-mm tubes and AccuVac Ampuls 2 6/pkg 173106

Optional reagents and apparatus

Description Unit Item no.

®
AccuVac vials for sample blanks 25/pkg 2677925
®
Ampule Breaker, PourRite ampules each 2484600
®
Ampule Breaker, Voluette ampules each 2196800
Cylinder, mixing, 25-mL each 2088640
Cylinder, mixing, 50-mL each 189641
®
Chlorine Standard Solution, 2-mL PourRite Ampules, 50–75 mg/L 20/pkg 1426820
®
Chlorine Standard Solution, 10-mL Voluette Ampule, 50–75 mg/L 16/pkg 1426810
®
Chlorine Standard Solution, 2-mL PourRite Ampules, 25–30 mg/L 20/pkg 2630020
DPD Total Chlorine Reagent Powder Pillows, 10-mL 1000/pkg 2105628
DPD Total Chlorine Reagent Powder Pillows, 10-mL 300/pkg 2105603
DPD Total Chlorine Reagent, 10 mL, SwifTest™ Dispenser refill vial 250 tests 2105660
Paper, pH, 0–14 pH range 100/pkg 2601300
®
Pipet, TenSette , 0.1–1.0 mL each 1970001
Pipet tips for TenSette Pipet 1970001 50/pkg 2185696
Pipet tips for TenSette Pipet 1970001 1000/pkg 2185628
Potassium Iodide, 30 g/L 100 mL 34332
Sodium Arsenite, 5 g/L 100 mL 104732
100 mL
Sodium Hydroxide Standard Solution, 1.0 N 104532
MDB
SpecCheck™ Secondary Standard Kit, Chlorine DPD, 0-2.0 mg/L Set each 2635300
100 mL
Sulfuric Acid Standard Solution, 1 N 127032
MDB
Water, Chlorine-demand Free 500 mL 2641549
Water, deionized 4L 27256

FOR TECHNICAL ASSISTANCE, PRICE INFORMATION AND ORDERING: HACH COMPANY

In the U.S.A. – Call toll-free 800-227-4224 WORLD HEADQUARTERS
Outside the U.S.A. – Contact the HACH office or distributor serving you. Telephone: (970) 669-3050
On the Worldwide Web – www.hach.com; E-mail – [email protected] FAX: (970) 669-2932

© Hach Company/Hach Lange GmbH, 1989–2013. All rights reserved. 04/2013, Edition 8
Chloramine (Mono) and Nitrogen, DOC316.53.01016

Free Ammonia
Indophenol Method1 Method 10200
0.04 to 4.50 mg/L Cl2 Powder Pillows
0.01 to 0.50 mg/L NH3–N
Scope and application: For the determination of free ammonia and monochloramine simultaneously in finished
chloraminated water. This product has not been evaluated to test for chlorine and chloramines in medical
applications in the United States.
1 U.S. Patent 6,315,950

Test preparation

Instrument specific information

Table 1 shows all of the instruments that have the program for this test. The table also
shows requirements that may vary between instruments, such as adapter and sample cell
requirements. To use this table, select an instrument, then read across to find the
corresponding information for this test.
Table 1 Instrument-specific information
Instrument Adapter Sample cell orientation Sample cell
DR 6000 A23618 The orientation key is toward the user. 4864302
DR 5000 The orientation key is toward the user.
DR 3900 LZV846 (A) The orientation key is away from the user.
DR 900 — The orientation key is toward the user.
DR 3800 LZV585 (B) The 1-cm path is aligned with the arrow on the adapter. 5940506
DR 2800
DR 2700

Before starting
Samples must be analyzed immediately after collection and cannot be preserved for later analysis.
Install the instrument cap on the DR 900 cell holder before ZERO or READ is pushed.
In bright light conditions (e.g., direct sunlight), close the cell compartment on spectrophotometers, if applicable, with the
protective cover during measurements.
For best results, measure the reagent blank value for each new lot of reagent. Replace the sample with deionized water in
the test procedure to get the reagent blank value. Subtract the reagent blank value from the sample results automatically
with the reagent blank adjust option.
Review the Safety Data Sheets (MSDS/SDS) for the chemicals that are used and use any recommended personal protective
equipment.
Dispose of reacted solutions according to local, state and federal regulations. Use the Safety Data Sheets for disposal
information for unused reagents. Consult the environmental, health and safety staff for your facility and/or local regulatory
agencies for further disposal information.

1
Items to collect
Description Quantity

Free Ammonia Reagent Solution 1 drop

Monochlor F Reagent Pillows 2
Sample cells (For information about sample cells, adapters or light shields, refer to Instrument
2
specific information on page 1.)

Refer to Consumables and replacement items 01015 for reorder information.

Sample collection and storage

Collect samples in clean glass bottles. Open the sample valve or spigot and let the water
flow for a minimum of 5 minutes. Rinse the sample bottle several times with the sample
and let the sample overflow each time, then cap the container so that there is no head
space (air) above the sample. Analyze samples immediately after collection.
Powder pillow procedure

Start

1. Start program 66, 2. Fill two sample cells to 3. Clean the 4. Insert the
Monochloramine LR. For the 10-mL line with sample. monochloramine sample monochloramine sample
information about sample Write Free Ammonia on cell. cell into the cell holder.
cells, adapters or light one sample cell. Write
shields, refer to Instrument Monochloramine on the
specific information second sample cell.
on page 1.
Note: Although the program
name may vary between
instruments, the program
number does not change.

Zero

5. Push ZERO. The display 6. Remove the sample from 7. Add the contents of one 8. Close the sample cell.
shows 0.00 mg/L Cl2. the cell holder. Monochlor F Reagent Shake the sample cell for
Powder Pillow to the sample about 20 seconds to
for monochloramine dissolve the reagent. A
measurement. green color will show if
monochloramine is present.

2 Chloramine (Mono) (4.50 mg/L) and Free Ammonia (0.50 mg/L), Indophenol Method
9. Add one drop of Free 10. Close the free 11. Start the instrument 12. When the timer expires,
Ammonia Reagent Solution ammonia sample cell. timer. A 5-minute reaction clean the monochloramine
to the sample cell for free Invert the sample cell to mix time starts. sample cell.
ammonia measurement. the reagent. For samples colder than
Close the reagent bottle to If the sample gets cloudy by 18 °C, refer to Table 2
keep the reagent stable. the end of the reaction on page 4.
period, pre-treat the sample
and do the test again. Refer
to Interferences
on page 4.

Read Start Zero

13. Insert the 14. Push READ. Results 15. Exit program 66. Start 16. Leave the
monochloramine sample show in mg/L Cl2. program 388 N, Ammonia monochloramine sample
cell into the cell holder. Free. cell in the cell holder. Push
ZERO. The display will
show 0.00 mg/L NH3–N f.

17. Remove the 18. When the reaction time 19. Close the sample cell. 20. Start the instrument
monochloramine sample cell in step 11 is complete, add Shake the sample cell for timer. A 5-minute reaction
from the cell holder. the contents of one about 20 seconds to time starts.
Monochlor F Powder Pillow dissolve the reagent. A For samples colder than
to the Free Ammonia green color will show if 18 °C, refer to Table 2
sample cell. monochloramine is present. on page 4.

Chloramine (Mono) (4.50 mg/L) and Free Ammonia (0.50 mg/L), Indophenol Method 3
 Read

21. When the timer expires, 22. Insert the free 23. Push READ. The
clean the free ammonia ammonia sample cell into results are in mg/L NH3–N f.
sample cell. the cell holder.

Color development time

Test results are strongly influenced by the sample temperature. The reaction period
indicated in the procedure is for a sample temperature of 18–20 ºC (64–68 ºF). Adjust the
reaction period according to Table 2. Samples can be read up to 15 minutes after the
listed development time.
Table 2 Color development time
Sample temperature (°C) Sample temperature (°F) Development time (mintues)
5 40 10
7 42 9
9 48 8
10 50 8
12 54 7
14 58 7
16 61 6
18 68 4
20 73 3
23 75 2.5
25 77 2
> 25 > 77 2

Interferences
This method is intended for finished, chloraminated drinking water samples that have a
measurable combined (total) chlorine disinfectant residual. Samples that do not have a
disinfectant residual and samples that have a chlorine demand may show low ammonia
test results. Blanks and ammonia standards that are analyzed without a disinfectant
residual must be prepared with high quality, reagent grade water.
The substances that are shown in Interferences on page 4 do not interfere in the free
ammonia determination at or below the given concentration.
Table 3 Non-interfering substances
Substance Maximum level tested
Aluminum 0.2 mg/L Al
Chloride 1200 mg/L Cl–
Copper 1 mg/L Cu

4 Chloramine (Mono) (4.50 mg/L) and Free Ammonia (0.50 mg/L), Indophenol Method
 Table 3 Non-interfering substances (continued)
Substance Maximum level tested
Iron 0.3 mg/L Fe
Manganese 0.05 mg/L Mn
Nitrate 10 mg/L NO3––N
Nitrite 1 mg/L NO2––N
Phosphate 2 mg/L o-PO4
Silica 100 mg/L SiO2
Sulfate 1600 ppm as SO42–
Zinc 5 ppm Zn

Samples that contain high levels of both Total Hardness and Alkalinity may become
cloudy after the addition of the Free Ammonia Reagent Solution. If this occurs by the end
of the first reaction period, the sample for Free Ammonia measurement must be
pretreated as follows:
1. Measure 10 mL of sample into the sample cell for Free Ammonia.
2. Add the contents of one Hardness Treatment Reagent Powder Pillow to the sample.
3. Tighten the cap on the sample cell and invert until the reagent is dissolved.
4. Remove the cap.
5. Use the pretreated sample in the test procedure for the Free Ammonia sample.
Note: The sample for Monochloramine measurement does not need pretreatment.

Accuracy check
Standard solution method
Items to collect:
• Buffer Powder Pillow, pH 8.3
• Nitrogen, Ammonia Standard Solution, 100-mg/L as NH3–N
• Chlorine Solution Ampules, 50–70 mg/L
• 100-mL Class A volumetric flask
• 50-mL graduated cylinder
• Pipet, TenSette®, 0.1–1.0 mL and tips
• Pipets, Volumetric, 2-mL Class A and Mohr, 5-mL
• Pipet bulb
• Organic-free water

1. Prepare a 4.5-mg/L (as Cl2) monochloramine standard immediately before use as

follows.
a. Add the contents of one Buffer Powder Pillow, pH 8.3 to approximately 50 mL of
organic-free water in a clean 100-mL Class A volumetric flask. Swirl to dissolve
the powder.
b. Use a Class A volumetric pipet to add 2.00 mL of Nitrogen, Ammonia Standard
Solution, 100-mg/L as NH3–N into the flask.
c. Dilute to the mark with organic-free water. Mix well. This is a 2.00-mg/L buffered
ammonia standard.
d. Use a graduated cylinder to add 50.00 mL of the buffered ammonia standard into
a clean 100-mL beaker. Add a stir bar.
e. Find the exact concentration of the Chlorine Solution Ampules, 50–70 mg/L from
the label on the package.
f. Calculate the volume of the Chlorine Solution to add to the ammonia standard:
mL chlorine solution required = 455/(free chlorine concentration).

Chloramine (Mono) (4.50 mg/L) and Free Ammonia (0.50 mg/L), Indophenol Method 5
 g. Open an ampule and use a glass Mohr pipet to add the calculated amount of
Chlorine Solution slowly to the ammonia standard in the beaker. Keep the beaker
on a stir-plate at medium speed during the chlorine addition.
h. Stir the monochloramine solution for 1 minute after the Chlorine Solution addition
is complete.
i. Quantitatively transfer the monochloramine solution to a clean 100-mL Class A
volumetric flask. Dilute to the mark with organic-free water and mix well. This is a
nominal 4.5-mg/L (as Cl2) monochloramine standard.
2. Use this standard within 1 hour of preparation. Use the test procedure to measure the
concentration of the monochloramine standard solution.
3. Compare the expected result to the actual result.
Note: The factory calibration can be adjusted slightly with the standard adjust option so that the
instrument shows the expected value of the standard solution. The adjusted calibration is then
used for all test results. This adjustment can increase the test accuracy when there are slight
variations in the reagents or instruments.

Standard additions method (sample spike)

Use the standard additions method (for applicable instruments) to validate the test
procedure, reagents and instrument and to find if there is an interference in the sample.
Items to collect:
• Ammonium Nitrogen Standard Solution, 10 mg/L NH3–N
• 50-mL mixing cylinders
• Pipet, TenSette®, 0.1–1.0 mL and tips

1. Use the test procedure to measure the concentration of the sample, then keep the
(unspiked) sample in the instrument.
2. Go to the Standard Additions option in the instrument menu.
3. Select the values for standard concentration, sample volume and spike volumes.
4. Open the standard solution.
5. Prepare three spiked samples: use the TenSette pipet to add 0.3 mL, 0.6 mL and
1.0 mL of the standard solution, respectively, to three 50-mL portions of fresh sample.
Mix well.
6. Use the test procedure to measure the concentration of each of the spiked samples.
Start with the smallest sample spike. Measure each of the spiked samples in the
instrument.
7. Select Graph to compare the expected results to the actual results.
Note: If the actual results are significantly different from the expected results, make sure that
the sample volumes and sample spikes are measured accurately. The sample volumes and
sample spikes that are used should agree with the selections in the standard additions menu. If
the results are not within acceptable limits, the sample may contain an interference.

Standard solution method

Use the standard solution method to validate the test procedure, reagents and
instrument.
Items to collect:
• Ammonium Nitrogen Standard Solution, 10 mg/L NH3–N
• 100-mL plastic volumetric flask with stopper, Class A
• 2-mL volumetric pipet, Class A and pipet filler
• Deionized water—must be free of ammonia, chlorine and chlorine demand, for
example 18 MΩ-cm water from a deionizer system.

1. Prepare a 0.20 mg/L ammonia nitrogen standard solution as follows:

a. Use a pipet to add 2.00 mL of 10 mg/L ammonia nitrogen standard solution into
the volumetric flask. (Alternate preparation: add 0.4 mL of a 50 mg/L ammonia
nitrogen standard solution to the volumetric flask.)

6 Chloramine (Mono) (4.50 mg/L) and Free Ammonia (0.50 mg/L), Indophenol Method
 b. Dilute to the mark with deionized water. Mix well. Prepare this solution daily.
2. Use the test procedure to measure the concentration of the prepared standard
solution.
3. Compare the expected result to the actual result.
Note: The factory calibration can be adjusted slightly with the standard adjust option so that the
instrument shows the expected value of the standard solution. The adjusted calibration is then
used for all test results. This adjustment can increase the test accuracy when there are slight
variations in the reagents or instruments.

Method performance
The method performance data that follows was derived from laboratory tests that were
measured on a spectrophotometer during ideal test conditions. Users may get different
results under different test conditions.
Program Standard Precision (95% Confidence Interval) Sensitivity
Concentration change per 0.010 Abs change
66 2.60 mg/L Cl2 2.58–2.62 mg/L Cl2 0.04 mg/L Cl2
388 0.20 mg/L NH3–N 0.19–0.21 mg/L NH3–N 0.01 mg/L NH3–N

Summary of method
Monochloramine (NH2Cl) and free ammonia (NH3 and NH4+) can exist in the same water
sample. Added hypochlorite combines with free ammonia to form more monochloramine.
In the presence of a cyanoferrate catalyst, monochloramine in the sample reacts with a
substituted phenol to form an intermediate monoimine compound. The intermediate
couples with excess substituted phenol to form a green-colored indophenol, which is
proportional to the amount of monochloramine present in the sample. Free ammonia is
determined by comparing the color intensities, with and without added hypochlorite. The
measurement wavelength is 655 nm for spectrophotometers or 610 nm for colorimeters.
Consumables and replacement items
Required reagents

Description Quantity/test Unit Item no.

Free Ammonia Reagent Set, includes: — 50/pkg 2879700

Free Ammonia Reagent Solution 1 drop 4 mL SCDB 2877336
Monochlor F Reagent Pillows 2 100/pkg 2802299

Recommended standards or Recommended standards and apparatus

Description Unit Item no.

Buffer Powder Pillow, pH 8.3 25/pkg 89868

®
Chlorine Standard Solution, 10-mL Voluette Ampule, 50-75 mg/L 16/pkg 1426810
®
Chlorine Standard Solution, 2-mL PourRite Ampule, 50-75 mg/L 20/pkg 1426820
®
Chlorine Standard Solution, 2-mL PourRite Ampules, 25–30 mg/L 20/pkg 2630020
Hardness Treatment Reagent Pillows 50/pkg 2882346
Nitrogen Ammonia Standard Solution, 10-mg/L NH3–N 500 mL 15349
®
Nitrogen Ammonia Standard Solution, 10-mL Voluette Ampule, 50-mg/L NH3–N 16/pkg 1479110
Nitrogen Ammonia Standard Solution, 100-mg/L as NH3–N 500 mL 2406549
®
PourRite Ampule Breaker, 2-mL each 2484600
®
Ampule Breaker, Voluette ampules each 2196800
Organic-free water 500 mL 2641549

Chloramine (Mono) (4.50 mg/L) and Free Ammonia (0.50 mg/L), Indophenol Method 7
Optional reagents and apparatus

Description Unit Item no.

Beaker, 100 mL, polypropylene each 108042

Beaker, glass, 100-mL each 50042H
Cylinder, mixing, 50-mL each 2088641
Flask, volumetric, Class A, 100-mL each 1457442
Free Ammonia Reagent Set 250/pkg 2879701
Monochloramine/Free Ammonia SpecCheck™ Kit each 2507500
Pipet filler, safety bulb each 1465100
®
Pipet, TenSette , 0.1–1.0 mL each 1970001
Pipet tips for TenSette Pipet 1970001 50/pkg 2185696
Pipet tips for TenSette Pipet 1970001 1000/pkg 2185628
Pipet, Mohr, glass, 10-mL each 2093438
Pipet, volumetric, Class A, 2-mL each 1451536
Pipet, volumetric, Class A, 50-mL each 1451541
Scissors each 2883100
Stir bar, octagonal each 2095352
Stirrer, magnetic each 2881200
Thermometer, -10 to 110 °C each 187701
Wipes, disposable 280/pkg 2097000

FOR TECHNICAL ASSISTANCE, PRICE INFORMATION AND ORDERING: HACH COMPANY

In the U.S.A. – Call toll-free 800-227-4224 WORLD HEADQUARTERS
Outside the U.S.A. – Contact the HACH office or distributor serving you. Telephone: (970) 669-3050
On the Worldwide Web – www.hach.com; E-mail – [email protected] FAX: (970) 669-2932

© Hach Company/Hach Lange GmbH, 1989–2013. All rights reserved. 04/2013, Edition 8
Chloramine (Mono) DOC316.53.01014

Indophenol Method1 Method 10172

0.1 to 10.0 mg/L Cl2 (HR) Test 'N Tube
Scope and application: For chloraminated drinking water and chlorinated wastewater. This product has not been
evaluated to test for chlorine and chloramines in medical applications in the United States.
1 Patent Pending

Test preparation

Instrument specific information

The table in this section shows all of the instruments that have the program for this test.
Table 1 shows adapter and light shield requirements for the instruments that use them.
To use the table, select an instrument, then read across to find the corresponding
information for this test.
Table 1 Instrument-specific information for test tubes
Instrument Adapters Light shield
DR 6000 — —
DR 5000 — —
DR 900 4846400 Cover supplied with the instrument
DR 3900 — LZV849
DR 3800 — LZV646
DR 2800 —

Before starting
Samples must be analyzed immediately after collection and cannot be preserved for later analysis.
Install the instrument cap on the DR 900 cell holder before ZERO or READ is pushed.
DR 3900, DR 3800, DR 2800 and DR 2700: Install the light shield in Cell Compartment #2 before this test is started.
For best results, measure the reagent blank value for each new lot of reagent. Replace the sample with deionized water in
the test procedure to get the reagent blank value. Subtract the reagent blank value from the sample results automatically
with the reagent blank adjust option.
Review the Safety Data Sheets (MSDS/SDS) for the chemicals that are used and use any recommended personal protective
equipment.
Dispose of reacted solutions according to local, state and federal regulations. Use the Safety Data Sheets for disposal
information for unused reagents. Consult the environmental, health and safety staff for your facility and/or local regulatory
agencies for further disposal information.

Items to collect
Description Quantity

Light shield (For information about sample cells, adapters or light shields, refer to Instrument
1
specific information on page 1.)
High range Monochloramine Diluent Vials 1
Monochlor F Reagent Powder Pillow 1
Funnel, micro, poly 1

1
Items to collect (continued)
Description Quantity

Pipet, Mohr glass, 2.00 mL 1

Test tube rack 1

Refer to Consumables and replacement items on page 5 for reorder information.

Sample collection and storage

Collect samples in clean glass bottles. Open the sample valve or spigot and let the water
flow for a minimum of 5 minutes. Rinse the sample bottle several times with the sample
and let the sample overflow each time, then cap the container so that there is no head
space (air) above the sample. Analyze samples immediately after collection.
Test 'N Tube procedure

Start

1. Start program 67, 2. Remove the cap from a 3. Put the cap on the vial. 4. Clean the vial.
Monochloramine HR TNT. HR Monochloramine Diluent Invert to mix.
For information about vial. Use a pipet to add
sample cells, adapters or 2.0 mL of sample to the
light shields, refer to vial..
Instrument specific
information on page 1.
Note: Although the program
name may vary between
instruments, the program
number does not change.

Zero

5. Insert the vial into the 16- 6. Push ZERO. The display 7. Remove the vial from the 8. Add the contents of one
mm cell holder. shows 0.00 mg/L Cl2. cell holder. Monochlor F powder pillow
to the sample.

2 Chloramine (Mono), Indophenol TNT Method (10.0 mg/L)

 Read

9. Put the cap on the vial. 10. Start the instrument 11. When the timer expires, 12. Push READ. Results
Shake the vial for about timer. A 5-minute reaction insert the vial into the cell show in mg/L Cl2.
20 seconds to dissolve the time starts. holder.
reagent.

Interferences
Table 2 shows the substances that have been tested for interference and do not interfere
at or below the indicated levels. Table 3 suggests treatments for interferences.
Table 2 Non-interfering substances
Interfering substance Interference level
Alanine 1 mg/L N
Aluminum 10 mg/L Al
Bromide 100 mg/L Br–
Bromine 15 mg/L Br2
Calcium 1000 mg/L as CaCO3
Chloride 18,000 mg/L Cl–
Chlorine Dioxide 5 mg/L ClO2
Chromium (III) 5 mg/L Cr3+
Copper 10 mg/L Cu
Cyanide 10 mg/L CN–
Dichloramine 10 mg/L as Cl2
Fluoride 5 mg/L F–
Free Chlorine 10 mg/L Cl2
Glycine 1 mg/L N
Iron (II) 10 mg/L Fe2+
Iron (III) 10 mg/L Fe3+
Lead 10 mg/L Pb
Magnesium 1000 mg/L as CaCO3
Manganese (VII) 10 mg/L
Nitrate 100 mg/L N
Nitrite 50 mg/L N
Phosphate 100 mg/L PO4
Silica 100 mg/L SiO2
Sulfate 2600 mg/L SO42+
Sulfite 50 mg/L SO32–

Chloramine (Mono), Indophenol TNT Method (10.0 mg/L) 3

 Table 2 Non-interfering substances (continued)
Interfering substance Interference level
Tyrosine 1 mg/L N
Urea 10 mg/L N
Zinc 5 mg/L Zn

Table 3 Interfering substances

Substance Effect Interference level Recommended treatment
Ozone – Above 1 mg/L Usually does not coexist with monochloramine.
Sulfide + A "rust" color develops if present. Usually does not coexist with monochloramine.
Thiocyanate – Above 0.5 mg/L This method will tolerate up to 2 mg/L.

Accuracy check
Standard solution method
Items to collect:
• Buffer Powder Pillow, pH 8.3
• Nitrogen, Ammonia Standard Solution, 100-mg/L as NH3–N
• Chlorine Solution Ampules, 50–70 mg/L
• 100-mL Class A volumetric flask
• 50-mL graduated cylinder
• Pipet, TenSette®, 0.1–1.0 mL and tips
• Pipets, Volumetric, 2-mL Class A and Mohr, 5-mL
• Pipet bulb
• Organic-free water

1. Prepare a 4.5-mg/L (as Cl2) monochloramine standard immediately before use as

follows.
a. Add the contents of one Buffer Powder Pillow, pH 8.3 to approximately 50 mL of
organic-free water in a clean 100-mL Class A volumetric flask. Swirl to dissolve
the powder.
b. Use a Class A volumetric pipet to add 2.00 mL of Nitrogen, Ammonia Standard
Solution, 100-mg/L as NH3–N into the flask.
c. Dilute to the mark with organic-free water. Mix well. This is a 2.00-mg/L buffered
ammonia standard.
d. Use a graduated cylinder to add 50.00 mL of the buffered ammonia standard into
a clean 100-mL beaker. Add a stir bar.
e. Find the exact concentration of the Chlorine Solution Ampules, 50–70 mg/L from
the label on the package.
f. Calculate the volume of the Chlorine Solution to add to the ammonia standard:
mL chlorine solution required = 455/(free chlorine concentration).
g. Open an ampule and use a glass Mohr pipet to add the calculated amount of
Chlorine Solution slowly to the ammonia standard in the beaker. Keep the beaker
on a stir-plate at medium speed during the chlorine addition.
h. Stir the monochloramine solution for 1 minute after the Chlorine Solution addition
is complete.

4 Chloramine (Mono), Indophenol TNT Method (10.0 mg/L)

 i.Quantitatively transfer the monochloramine solution to a clean 100-mL Class A
volumetric flask. Dilute to the mark with organic-free water and mix well. This is a
nominal 4.5-mg/L (as Cl2) monochloramine standard.
2. Use this standard within 1 hour of preparation. Use the test procedure to measure the
concentration of the monochloramine standard solution.
3. Compare the expected result to the actual result.
Note: The factory calibration can be adjusted slightly with the standard adjust option so that the
instrument shows the expected value of the standard solution. The adjusted calibration is then
used for all test results. This adjustment can increase the test accuracy when there are slight
variations in the reagents or instruments.

Method performance
The method performance data that follows was derived from laboratory tests that were
measured on a spectrophotometer during ideal test conditions. Users may get different
results under different test conditions.
Program Standard Precision (95% Confidence Interval) Sensitivity
Concentration change per 0.010 Abs change
67 5.9 mg/L Cl2 5.6–6.2 mg/L Cl2 0.1 mg/L Cl2

Summary of method
In the presence of a cyanoferrate catalyst, monochloramine (NH2Cl) in the sample reacts
with a substituted phenol to form an intermediate monoimine compound. The
intermediate couples with excess substituted phenol to form a green-colored indophenol,
which is proportional to the amount of monochloramine present in the sample. The
measurement wavelength is 655 nm for spectrophotometers or 610 nm for colorimeters.
Consumables and replacement items
Required reagents

Description Quantity/Test Unit Item no.

Monochloramine, HR, Test 'N Tubes, includes: — — 2805145

HR Monochloramine Diluent Vials1 1 50/pkg —
Funnel, micro, poly 1 each 2584335
Monochlor F Reagent Pillows 1 50/pkg 2802246
1 Not sold separately

Required apparatus

Description Quantity/Test Unit Item no.

Pipet, Mohr, glass 2.00 mL 1 each 2093636

Pipet filler, safety bulb 1 each 1465100
Test tube rack 1 each 1864100

Recommended standards

Description Unit Item no.

Buffer Powder Pillow, pH 8.3 25/pkg 89868

®
Chlorine Standard Solution, 10-mL Voluette Ampule, 50–75 mg/L 16/pkg 1426810
®
Chlorine Standard Solution, 2-mL PourRite Ampules, 50–75 mg/L 20/pkg 1426820
®
Chlorine Standard Solution, 2-mL PourRite Ampules, 25–30 mg/L 20/pkg 2630020
Nitrogen Ammonia Standard Solution, 100-mg/L as NH3–N 500 mL 2406549

Chloramine (Mono), Indophenol TNT Method (10.0 mg/L) 5

Consumables and replacement items (continued)
Description Unit Item no.

Nitrogen, Ammonia Standard Solution, 1000-mg/L NH3-N 1L 2354153

Organic-free water 500 mL 2641549

Optional reagents and apparatus

Description Unit Item no.

®
Ampule Breaker, PourRite ampules each 2484600
®
Ampule Breaker, Voluette ampules each 2196800
Beaker, glass, 100-mL each 50042H
Clippers each 96800
Flask, volumetric, Class A, 100-mL each 1457442
Monochloramine/Free Ammonia SpecCheck™ Kit each 2507500
Paper, pH, 0–14 pH range 100/pkg 2601300
Pipet, Mohr, glass, 5-mL each 2093437
®
Pipet, TenSette , 0.1–1.0 mL each 1970001
Pipet tips for TenSette Pipet 1970001 50/pkg 2185696
Pipet tips for TenSette Pipet 1970001 1000/pkg 2185628
Pipet, volumetric, Class A, 2-mL each 1451536
Pipet, volumetric, Class A, 50-mL each 1451541
Rochelle Salt Solution 29 mL 172533
Shears each 2369400
Stir bar, octagonal each 2095352
Stirrer, magnetic each 2881200

FOR TECHNICAL ASSISTANCE, PRICE INFORMATION AND ORDERING: HACH COMPANY

In the U.S.A. – Call toll-free 800-227-4224 WORLD HEADQUARTERS
Outside the U.S.A. – Contact the HACH office or distributor serving you. Telephone: (970) 669-3050
On the Worldwide Web – www.hach.com; E-mail – [email protected] FAX: (970) 669-2932

© Hach Company/Hach Lange GmbH, 1989–2013. All rights reserved. 04/2013, Edition 8
Chloramine (Mono) DOC316.53.01015

Indophenol Method1 Method 10171

0.04 to 4.50 mg/L Cl2 (LR) Powder Pillows
Scope and application: For chloraminated drinking water and chlorinated wastewater. This product has not been
evaluated to test for chlorine and chloramines in medical applications in the United States.
1 U.S. Patent 6,315,950

Test preparation

Instrument specific information

Table 1 shows all of the instruments that have the program for this test. The table also
shows requirements that may vary between instruments, such as adapter and sample cell
requirements. To use this table, select an instrument, then read across to find the
corresponding information for this test.
Table 1 Instrument-specific information
Instrument Adapter Sample cell orientation Sample cell
DR 6000 A23618 The orientation key is toward the user. 4864302
DR 5000 The orientation key is toward the user.
DR 3900 LZV846 (A) The orientation key is away from the user.
DR 900 — The orientation key is toward the user.
DR 3800 LZV585 (B) The 1-cm path is aligned with the arrow on the adapter. 5940506
DR 2800
DR 2700

Before starting
Samples must be analyzed immediately after collection and cannot be preserved for later analysis.
Install the instrument cap on the DR 900 cell holder before ZERO or READ is pushed.
In bright light conditions (e.g., direct sunlight), close the cell compartment on spectrophotometers, if applicable, with the
protective cover during measurements.
To measure chloramine (mono) and free ammonia on the same sample, use Method 10200 Nitrogen, Free Ammonia and
Chloramine (Mono).
For best results, measure the reagent blank value for each new lot of reagent. Replace the sample with deionized water in
the test procedure to get the reagent blank value. Subtract the reagent blank value from the sample results automatically
with the reagent blank adjust option.
Review the Safety Data Sheets (MSDS/SDS) for the chemicals that are used and use any recommended personal protective
equipment.
Dispose of reacted solutions according to local, state and federal regulations. Use the Safety Data Sheets for disposal
information for unused reagents. Consult the environmental, health and safety staff for your facility and/or local regulatory
agencies for further disposal information.

1
Items to collect
Description Quantity

Monochlor F reagent pillows 1

Sample cells (For information about sample cells, adapters or light shields, refer to Instrument
2
specific information on page 1.)

Refer to Consumables and replacement items on page 6 for reorder information.

Sample collection and storage

Collect samples in clean glass bottles. Open the sample valve or spigot and let the water
flow for a minimum of 5 minutes. Rinse the sample bottle several times with the sample
and let the sample overflow each time, then cap the container so that there is no head
space (air) above the sample. Analyze samples immediately after collection.
Color development time
Test results are strongly influenced by the sample temperature. The reaction period
indicated in the procedure is for a sample temperature of 18–20 ºC (64–68 ºF). Adjust the
reaction period according to Table 2. Samples can be read up to 15 minutes after the
listed development time.
Table 2 Color development time
Sample temperature (°C) Sample temperature (°F) Development time (mintues)
5 40 10
7 42 9
9 48 8
10 50 8
12 54 7
14 58 7
16 61 6
18 68 4
20 73 3
23 75 2.5
25 77 2
> 25 > 77 2

2 Chloramine (Mono), Indophenol Method (4.50 mg/L)

Indophenol method, powder pillows

Start

1. Start program 66, 2. Fill the sample cell with 3. Clean the prepared 4. Insert the blank into the
Monochloramine LR. For 10 mL of sample. sample. cell holder.
information about sample
cells, adapters or light
shields, refer to Instrument
specific information
on page 1.
Note: Although the program
name may vary between
instruments, the program
number does not change.

Zero

5. Push ZERO. The display 6. Remove the sample from 7. Add the contents of one 8. Close the sample cell.
shows 0.00 mg/L Cl2. the cell holder. Monochlor F powder pillow Shake the sample cell for
to the sample cell. about 20 seconds to
dissolve the reagent.

Read

9. Start the instrument 10. Clean the prepared 11. When the timer expires, 12. Push READ. Results
timer. A 5-minute reaction sample. insert the prepared sample show in mg/L Cl2.
time starts. into the cell holder.
For samples colder than
18 °C, refer to Table 2
on page 2.

Chloramine (Mono), Indophenol Method (4.50 mg/L) 3

Interferences
Table 3 shows the substances that have been tested for interference and do not interfere
at or below the indicated levels. Table 4 suggests treatments for interferences.
Table 3 Non-interfering substances
Interfering substance Interference level
Alanine 1 mg/L N
Aluminum 10 mg/L Al
Bromide 100 mg/L Br–
Bromine 15 mg/L Br2
Calcium 1000 mg/L as CaCO3
Chloride 18,000 mg/L Cl–
Chlorine Dioxide 5 mg/L ClO2
Chromium (III) 5 mg/L Cr3+
Copper 10 mg/L Cu
Cyanide 10 mg/L CN–
Dichloramine 10 mg/L as Cl2
Fluoride 5 mg/L F–
Free Chlorine 10 mg/L Cl2
Glycine 1 mg/L N
Iron (II) 10 mg/L Fe2+
Iron (III) 10 mg/L Fe3+
Lead 10 mg/L Pb
Nitrate 100 mg/L N
Nitrite 50 mg/L N
Phosphate 100 mg/L PO4
Silica 100 mg/L SiO2
Sulfate 2600 mg/L SO42+
Sulfite 50 mg/L SO32–
Tyrosine 1 mg/L N
Urea 10 mg/L N
Zinc 5 mg/L Zn

Table 4 Interfering substances

Substance Effect Interference level Recommended treatment
Magnesium + Above 400 mg/L CaCO3 Add 5 drops of Rochelle Salt Solution prior to testing. OR: use
the high range (HR) test.
Manganese (+7) – Above 3 mg/L Use the HR test; it will tolerate up to 10 mg/L.
Ozone – Above 1 mg/L Usually does not coexist with monochloramine.
Sulfide + A "rust" color develops if present. Usually does not coexist with monochloramine.
Thiocyanate – Above 0.5 mg/L This method will tolerate up to 2 mg/L.

4 Chloramine (Mono), Indophenol Method (4.50 mg/L)

Accuracy check
Standard solution method
Items to collect:
• Buffer Powder Pillow, pH 8.3
• Nitrogen, Ammonia Standard Solution, 100-mg/L as NH3–N
• Chlorine Solution Ampules, 50–70 mg/L
• 100-mL Class A volumetric flask
• 50-mL graduated cylinder
• Pipet, TenSette®, 0.1–1.0 mL and tips
• Pipets, Volumetric, 2-mL Class A and Mohr, 5-mL
• Pipet bulb
• Organic-free water

1. Prepare a 4.5-mg/L (as Cl2) monochloramine standard immediately before use as

follows.
a. Add the contents of one Buffer Powder Pillow, pH 8.3 to approximately 50 mL of
organic-free water in a clean 100-mL Class A volumetric flask. Swirl to dissolve
the powder.
b. Use a Class A volumetric pipet to add 2.00 mL of Nitrogen, Ammonia Standard
Solution, 100-mg/L as NH3–N into the flask.
c. Dilute to the mark with organic-free water. Mix well. This is a 2.00-mg/L buffered
ammonia standard.
d. Use a graduated cylinder to add 50.00 mL of the buffered ammonia standard into
a clean 100-mL beaker. Add a stir bar.
e. Find the exact concentration of the Chlorine Solution Ampules, 50–70 mg/L from
the label on the package.
f. Calculate the volume of the Chlorine Solution to add to the ammonia standard:
mL chlorine solution required = 455/(free chlorine concentration).
g. Open an ampule and use a glass Mohr pipet to add the calculated amount of
Chlorine Solution slowly to the ammonia standard in the beaker. Keep the beaker
on a stir-plate at medium speed during the chlorine addition.
h. Stir the monochloramine solution for 1 minute after the Chlorine Solution addition
is complete.
i. Quantitatively transfer the monochloramine solution to a clean 100-mL Class A
volumetric flask. Dilute to the mark with organic-free water and mix well. This is a
nominal 4.5-mg/L (as Cl2) monochloramine standard.
2. Use this standard within 1 hour of preparation. Use the test procedure to measure the
concentration of the monochloramine standard solution.
3. Compare the expected result to the actual result.
Note: The factory calibration can be adjusted slightly with the standard adjust option so that the
instrument shows the expected value of the standard solution. The adjusted calibration is then
used for all test results. This adjustment can increase the test accuracy when there are slight
variations in the reagents or instruments.

Method performance
The method performance data that follows was derived from laboratory tests that were
measured on a spectrophotometer during ideal test conditions. Users may get different
results under different test conditions.
Program Standard Precision (95% Confidence Interval) Sensitivity
Concentration change per 0.010 Abs change
66 2.60 mg/L Cl2 2.58–2.62 mg/L Cl2 0.04 mg/L Cl2

Chloramine (Mono), Indophenol Method (4.50 mg/L) 5

Summary of method
In the presence of a cyanoferrate catalyst, monochloramine (NH2Cl) in the sample reacts
with a substituted phenol to form an intermediate monoimine compound. The
intermediate couples with excess substituted phenol to form a green-colored indophenol,
which is proportional to the amount of monochloramine present in the sample. The
measurement wavelength is 655 nm for spectrophotometers or 610 nm for colorimeters.
Consumables and replacement items
Required reagents

Description Quantity/Test Unit Item no.

Monochlor F Reagent Pillows 1 50/pkg 2802246

Recommended standards

Description Unit Item no.

Buffer Powder Pillow, pH 8.3 25/pkg 89868

®
Chlorine Standard Solution, 10-mL Voluette Ampule, 50–75 mg/L 16/pkg 1426810
®
Chlorine Standard Solution, 2-mL PourRite Ampules, 50–75 mg/L 20/pkg 1426820
®
Chlorine Standard Solution, 2-mL PourRite Ampules, 25–30 mg/L 20/pkg 2630020
Nitrogen Ammonia Standard Solution, 100-mg/L as NH3–N 500 mL 2406549
Nitrogen, Ammonia Standard Solution, 1000-mg/L NH3-N 1L 2354153
Organic-free water 500 mL 2641549

Optional reagents and apparatus

Description Unit Item no.

®
Ampule Breaker, PourRite ampules each 2484600
®
Ampule Breaker, Voluette ampules each 2196800
Beaker, glass, 100-mL each 50042H
Flask, volumetric, Class A, 100-mL each 1457442
Monochlor F Reagent Powder Pillows 100/pkg 2802299
Monochloramine/Free Ammonia SpecCheck™ Kit each 2507500
Paper, pH, 0–14 pH range 100/pkg 2601300
Pipet, Mohr, glass, 5-mL each 2093437
®
Pipet, TenSette , 0.1–1.0 mL each 1970001
Pipet tips for TenSette Pipet 1970001 50/pkg 2185696
Pipet tips for TenSette Pipet 1970001 1000/pkg 2185628
Pipet, volumetric, Class A, 2-mL each 1451536
Pipet, volumetric, Class A, 50-mL each 1451541
Rochelle Salt Solution 29 mL 172533
Shears each 2369400
Stir bar, octagonal each 2095352
Stirrer, magnetic each 2881200
Thermometer, non-mercury, -10 to +225 °C each 2635700

6 Chloramine (Mono), Indophenol Method (4.50 mg/L)

Chloramine (Mono), Indophenol Method (4.50 mg/L) 7
 FOR TECHNICAL ASSISTANCE, PRICE INFORMATION AND ORDERING: HACH COMPANY
In the U.S.A. – Call toll-free 800-227-4224 WORLD HEADQUARTERS
Outside the U.S.A. – Contact the HACH office or distributor serving you. Telephone: (970) 669-3050
On the Worldwide Web – www.hach.com; E-mail – [email protected] FAX: (970) 669-2932

© Hach Company/Hach Lange GmbH, 1989–2013. All rights reserved. 04/2013, Edition 8
Chlorine Dioxide DOC316.53.01020

Direct Reading Method Method 8345

1 to 50 mg/L ClO2 (MR)
Scope and application: For water and wastewater.

Test preparation

Instrument specific information

The table in this section shows all of the instruments that have the program for this test.
Table 1 shows sample cell and orientation requirements for reagent addition tests, such
as powder pillow or bulk reagent tests.
To use the table, select an instrument, then read across to find the corresponding
information for this test.
Table 1 Instrument-specific information for reagent addition
Instrument Sample cell orientation Sample cell
DR 6000 The fill line is to the right. 2495402
DR 3800
DR 2800
DR 2700
DR 5000 The fill line is toward the user.
DR 3900
DR 900 The orientation mark is toward the user. 2401906

Before starting
Samples must be analyzed immediately after collection and cannot be preserved for later analysis.
Install the instrument cap on the DR 900 cell holder before ZERO or READ is pushed.
Gloves and goggles are recommended.

Items to collect
Description Quantity

Deionized water 10 mL
Sample cells (For information about sample cells, adapters or light shields, refer to Instrument
2
specific information on page 1.)

Refer to Consumables and replacement items on page 3 for reorder information.

Sample collection
• Samples must be analyzed immediately and cannot be preserved for later analysis.
• Chlorine dioxide is a strong oxidizing agent and it is unstable in natural waters. It
reacts quickly with various inorganic compounds and more slowly with organic

1
 compounds. Many factors, including reactant concentrations, sunlight, pH,
temperature and salinity influence the decomposition of chlorine dioxide in water.
• Collect samples in clean glass bottles. Avoid plastic containers since these may have
a large chlorine dioxide demand.
• Pre-treat glass sample containers to remove any chlorine dioxide demand. Soak the
containers in a dilute bleach solution (1 mL commercial bleach to 1 liter of deionized
water) for at least 1 hour. Rinse thoroughly with deionized or distilled water. If sample
containers are rinsed thoroughly with deionized or distilled water after use, only
occasional pre-treatment is necessary.
• Be sure to get a representative sample. If the sample is taken from a spigot or faucet,
let the water flow for at least 5 minutes. Then let the container overflow with the
sample several times and then put the cap on the sample container so that there is
no headspace (air) above the sample. If a sample cell is used, rinse the cell several
times with the sample, then carefully fill to the 10-mL mark.

Direct reading method

Start

1. Start program 73 Chlor 2. Prepare the blank: Fill 3. Clean the blank. 4. Insert the blank into the
Diox MR. For information the sample cell with 10 mL cell holder.
about sample cells, of deionized water.
adapters or light shields,
refer to Instrument specific
information on page 1.
Note: Although the program
name may vary between
instruments, the program
number does not change.

Zero

5. Push ZERO. The display 6. Prepare the sample: Fill 7. Clean the prepared 8. Insert the prepared
shows 0 mg/L ClO2. a second sample cell with sample. sample into the cell holder.
10 mL of sample.

2 Chlorine Dioxide, Direct Read Method (50 mg/L)

 Read

9. Push READ. Results

show in mg/L ClO2.

Accuracy check
Standard solution method
The preparation of chlorine dioxide standards is difficult and hazardous. These standards
are explosive and volatile! Only a trained chemist should prepare the standards with
appropriate safety equipment and precautions. The manufacturer does not recommend
preparation of chlorine dioxide standards. If independent standard preparation is required,
refer to the instructions in Standard Methods for the Examination of Water and
Wastewater, Part 4500-ClO2 Chlorine Dioxide, under the headings "Stock chlorine
dioxide solution" and "Standard chlorine dioxide solution".
Method performance
The method performance data that follows was derived from laboratory tests that were
measured on a spectrophotometer during ideal test conditions. Users may get different
results under different test conditions.
Program Standard Precision (95% Confidence Interval) Sensitivity
Concentration change per 0.010 Abs change
73 43 mg/L ClO2 41–45 mg/L ClO2 0.3 mg/L ClO2

Summary of method
Chlorine dioxide, a yellow gas, can be measured directly in a water solution. The
measurement wavelength is 360 nm for spectrophotometers or 420 nm for colorimeters.
Consumables and replacement items
Required reagents and apparatus

Description Quantity/test Unit Item no.

Water, deionized varies 4L 27256

Sample cell, 10 mL square, matched pair 2 2/pkg 2495402

Optional apparatus

Description Unit Item no.

Gloves, chemical resistant, size 9-9.5 pair 24101041

Safety goggles, vented each 2550700
Standard Methods Book, most current edition each 2270800
1 Other sizes available

Chlorine Dioxide, Direct Read Method (50 mg/L) 3

 FOR TECHNICAL ASSISTANCE, PRICE INFORMATION AND ORDERING: HACH COMPANY
In the U.S.A. – Call toll-free 800-227-4224 WORLD HEADQUARTERS
Outside the U.S.A. – Contact the HACH office or distributor serving you. Telephone: (970) 669-3050
On the Worldwide Web – www.hach.com; E-mail – [email protected] FAX: (970) 669-2932

© Hach Company/Hach Lange GmbH, 1989–2013. All rights reserved. 04/2013, Edition 8
Chlorine Dioxide DOC316.53.01021

DPD Method1 Method 10126

®
0.04 to 5.00 mg/L ClO2 Powder Pillows or AccuVac Ampuls
Scope and application: For water and wastewater. USEPA accepted for reporting for drinking water analysis.2
1 Adapted from Standard Methods for the Examination of Water and Wastewater.
2 Procedure is equivalent to Standard Methods, 18 ed., 4500 ClO2 D.

Test preparation

Instrument-specific information
The tables in this section show all of the instruments that have the program for this test.
Table 1 shows sample cell and orientation requirements for reagent addition tests, such
as powder pillow or bulk reagent tests. Table 2 shows sample cell and adapter
requirements for AccuVac Ampul tests.
To use either table, select an instrument, then read across to find the corresponding
information for this test.
Table 1 Instrument-specific information for powder pillows
Instrument Sample cell orientation Sample cell
DR 6000 The fill line is to the right. 2495402
DR 3800
DR 2800
DR 2700
DR 5000 The fill line is toward the user.
DR 3900
DR 900 The orientation mark is toward the user. 2401906

Table 2 Instrument-specific information for AccuVac Ampuls

Instrument Adapter Sample cell
DR 6000 — 2427606
DR 5000
DR 900
DR 3900 LZV846 (A)
DR 3800 LZV584 (C) 2122800
DR 2800
DR 2700

Before starting
Samples must be analyzed immediately after collection and cannot be preserved for later analysis.
Install the instrument cap on the DR 900 cell holder before ZERO or READ is pushed.

1
For best results, measure the reagent blank value for each new lot of reagent. Replace the sample with deionized water in
the test procedure to get the reagent blank value. Subtract the reagent blank value from the sample results automatically
with the reagent blank adjust option.
If the chlorine dioxide concentration in the sample exceeds the upper limit of the test, the color may fade or the sample color
may change to yellow. Dilute the sample with a known volume of high quality, chlorine demand-free water and repeat the
test. Some loss of chlorine dioxide may occur due to the dilution. Multiply the result by the dilution factor.
Review the Safety Data Sheets (MSDS/SDS) for the chemicals that are used and use any recommended personal protective
equipment.
Dispose of reacted solutions according to local, state and federal regulations. Use the Safety Data Sheets for disposal
information for unused reagents. Consult the environmental, health and safety staff for your facility and/or local regulatory
agencies for further disposal information.

Items to collect
Powder pillows

Description Quantity

DPD Free Chlorine Powder Pillow, 10-mL 1

Glycine Reagent 4 drops
Sample cells. (For information about sample cells, adapters or light shields, refer to Instrument-
2
specific information on page 1.)
Stopper for 18-mm tubes and AccuVac Ampuls 2

Refer to Consumables and replacement items on page 7 for reorder information.

AccuVac Ampuls

Description Quantity
®
DPD Free Chlorine Reagent AccuVac Ampul 1
Glycine Reagent 16 drops
Beaker, 50-mL 1
Sample cells (For information about sample cells, adapters or light shields, refer to Instrument-
1
specific information on page 1.)
Stopper for 18-mm tubes and AccuVac Ampuls 1

Refer to Consumables and replacement items on page 7 for reorder information.

Sample collection
• Samples must be analyzed immediately and cannot be preserved for later analysis.
• Chlorine dioxide is a strong oxidizing agent and it is unstable in natural waters. It
reacts quickly with various inorganic compounds and more slowly with organic
compounds. Many factors, including reactant concentrations, sunlight, pH,
temperature and salinity influence the decomposition of chlorine dioxide in water.
• Collect samples in clean glass bottles. Avoid plastic containers since these may have
a large chlorine dioxide demand.
• Pre-treat glass sample containers to remove any chlorine dioxide demand. Soak the
containers in a dilute bleach solution (1 mL commercial bleach to 1 liter of deionized
water) for at least 1 hour. Rinse thoroughly with deionized or distilled water. If sample
containers are rinsed thoroughly with deionized or distilled water after use, only
occasional pre-treatment is necessary.
• Be sure to get a representative sample. If the sample is taken from a spigot or faucet,
let the water flow for at least 5 minutes. Then let the container overflow with the

2 Chlorine Dioxide, DPD Method (5.00 mg/L)

 sample several times and then put the cap on the sample container so that there is
no headspace (air) above the sample. If a sample cell is used, rinse the cell several
times with the sample, then carefully fill to the 10-mL mark.

Powder pillow procedure

Start

1. Start program 76 Chlor 2. Prepare the blank: Fill 3. Prepare the sample: Fill 4. Clean the blank.
Diox DPD. For information the sample cell with 10 mL a second sample cell with
about sample cells, of sample. 10 mL of sample.
adapters or light shields, Put the stopper in the blank. Put the stopper in the
refer to Instrument-specific prepared sample.
information on page 1.
Note: Although the program
name may vary between
instruments, the program
number does not change.

Zero

5. Insert the blank into the 6. Push ZERO. The display 7. Add 4 drops of Glycine 8. Swirl to mix.
cell holder. shows 0.00 mg/L ClO2. Reagent to the sample cell.

9. Add the contents of one 10. Swirl the sample cell for 11. Wait 30 seconds for any 12. Clean the prepared
DPD Free Chlorine Powder 20 seconds to mix. undissolved powder to sample.
Pillow to the sample cell. settle. Undissolved powder
will not affect accuracy.

Chlorine Dioxide, DPD Method (5.00 mg/L) 3

 Read

13. Within one minute of 14. Push READ. Results

adding the reagent, insert show in mg/L ClO2.
the prepared sample into
the cell holder.

AccuVac Ampul procedure

Start

1. Start program 77 Chlor 2. Prepare the blank: Fill 3. Clean the blank. 4. Insert the blank into the
Diox DPD AV. For the sample cell with 10 mL cell holder.
information about sample of sample.
cells, adapters or light Put the stopper in the blank.
shields, refer to Instrument-
specific information
on page 1.
Note: Although the program
name may vary between
instruments, the program
number does not change.

Zero

5. Push ZERO. The display 6. Prepare the sample: 7. Swirl to mix. 8. Fill the AccuVac Ampul
shows 0.00 mg/L ClO2. Collect at least 40 mL of with the prepared sample.
sample in a 50-mL beaker. Keep the tip immersed while
Add 16 drops of Glycine the Ampul fills completely.
Reagent to the sample in Close the Ampul.
the beaker.

4 Chlorine Dioxide, DPD Method (5.00 mg/L)

9. Quickly invert the Ampul 10. Wait 30 seconds for any 11. Clean the AccuVac 12. Within one minute of
several times to mix. undissolved powder to Ampul. adding the reagent, insert
Wait 30 seconds for any settle. Undissolved powder the prepared sample
undissolved powder to will not affect accuracy. AccuVac Ampul into the cell
settle. holder.

Read

13. Push READ. Results

show in mg/L ClO2.

Interferences
Interfering substance Interference level
Acidity More than 150 mg/L CaCO3. The full color may not develop or the color may fade instantly. Adjust
to pH 6–7 with 1 N Sodium Hydroxide. Measure the amount to be added on a separate sample
aliquot, then add the same amount to the sample that is tested. Correct the test result for the
dilution from the volume addition.
Alkalinity More than 250 mg/L CaCO3. The full color may not develop or the color may fade instantly. Adjust
to pH 6–7 with 1 N Sulfuric Acid. Measure the amount to add on a separate sample aliquot, then
add the same amount to the sample that is tested. Correct the test result for the dilution from the
volume addition.
Bromine, Br2 Interferes at all levels
Chlorine, Cl2 May interfere at more than 6 mg/L Cl2. Additional glycine may be able to remove this interference.
Chloramines, organic May interfere
Flocculating agents High levels of most flocculating agents are acceptable. The acceptable level is decreased when
chlorine is present. Refer to the information about metals in this table. In the presence of 0.6 mg/L
Cl2, Al(SO4)3 (< 500 mg/L) and FeCl2 (<200 mg/L) may be tolerated.
Hardness No effect at less than 1000 mg/L as CaCO3
Iodine, I2 Interferes at all levels

Chlorine Dioxide, DPD Method (5.00 mg/L) 5

Interfering substance Interference level
Manganese, Oxidized Pre-treat the sample as follows:
(Mn4+, Mn7+) or
Chromium, Oxidized 1. Adjust the sample pH to 6–7.
(Cr6+) 2. Add 3 drops of Potassium Iodide (30-g/L) to 10 mL of sample.
3. Mix and wait 1 minute.
4. Add 3 drops of Sodium Arsenite (5-g/L) and mix.
5. Use the test procedure to measure the concentration of the treated sample.
6. Subtract this result from the result without the treatment to obtain the correct chlorine
concentration.

Metals Various metals can combine with the glycine that is used to remove chlorine from the sample.
Metal interference is minimal except when chlorine is present. In the presence of 0.6 mg/L Cl2,
both copper (>10 mg/L) and nickel (>50 mg/L) interfere. Other metals that combine with glycine
may also interfere. It may be necessary to add more glycine to overcome this interference.
Monochloramine Causes a gradual drift to higher readings. When read within 1 minute after reagent addition,
3 mg/L monochloramine causes less than a 0.1 mg/L increase in the reading.
Ozone Interferes at all levels
Peroxides May interfere
Highly buffered samples Can prevent the correct pH adjustment of the sample by the reagents. Sample pretreatment may
or extreme sample pH be necessary.

Pollution prevention and waste management

If sodium arsenite was added to the sample for manganese or chromium interferences,
the reacted samples will contain arsenic and must be disposed of as a hazardous waste.
Dispose of reacted solutions according to local, state and federal regulations.
Accuracy check
Standard solution method
The preparation of chlorine dioxide standards is difficult and hazardous. These standards
are explosive and volatile! Only a trained chemist should prepare the standards with
appropriate safety equipment and precautions. The manufacturer does not recommend
preparation of chlorine dioxide standards. If independent standard preparation is required,
refer to the instructions in Standard Methods for the Examination of Water and
Wastewater, Part 4500-ClO2 Chlorine Dioxide, under the headings "Stock chlorine
dioxide solution" and "Standard chlorine dioxide solution".
Method performance
The method performance data that follows was derived from laboratory tests that were
measured on a spectrophotometer during ideal test conditions. Users may get different
results under different test conditions.
Program Standard Precision (95% Confidence Interval) Sensitivity
Concentration change per 0.010 Abs change
76 3.00 mg/L ClO2 2.89–3.11 mg/L ClO2 0.04 mg/L ClO2
77 3.00 mg/L ClO2 2.91–3.09 mg/L ClO2 0.04 mg/L ClO2

Summary of method
Chlorine dioxide reacts with DPD (N, N-diethyl-p-phenylenediamine) to the extent of one-
fifth of its total available chlorine content, which corresponds to the reduction of chlorine
dioxide to chlorite. A pink color forms, the intensity of which is proportional to the chlorine
dioxide concentration in the sample. Chlorine interference is removed with the addition of
glycine, which converts free chlorine to chloroaminoacetic acid, but has no effect on
chlorine dioxide at the test pH. The measurement wavelength is 530 nm for
spectrophotometers or 520 nm for colorimeters.

6 Chlorine Dioxide, DPD Method (5.00 mg/L)

Consumables and replacement items
Required reagents

Description Quantity/test Unit Item no.

Chlorine Dioxide DPD/Glycine Reagent Set 1 100/pkg 2770900

Includes:
DPD Free Chlorine Reagent Powder Pillow, 10-mL 1 100/pkg 2105569
Glycine Reagent 4 drops 29 mL 2762133
OR
®
Chlorine Dioxide DPD/Glycine AccuVac Ampul Reagent Set 1 25/pkg 2771000
Includes:
®
DPD Free Chlorine Reagent AccuVac Ampul 1 25/pkg 2502025
Glycine Reagent 4 drops 29 mL 2762133

Required apparatus

Description Quantity/test Unit Item no.

AccuVac Snapper 1 each 2405200

Beaker, 50-mL 1 each 50041H
Stoppers for 18-mm tubes and AccuVac Ampuls 2 6/pkg 173106
Sample cell, 10 mL round, 25 x 54 mm 1 each 2122800
Sample cell, 10 mL round, 25 x 60 mm 1 6/pkg 2427606
Sample cell, 10 mL square, matched pair 2 2/pkg 2495402

Recommended standards and apparatus

Description Unit Item no.

®
Chlorine Standard Solution, 10-mL Voluette Ampule, 50–75 mg/L 16/pkg 1426810
®
Ampule Breaker, Voluette ampules each 2196800
Organic-free water 500 mL 2641549

Optional reagents and apparatus

Description Unit Item no.

®
AccuVac vials for sample blanks 25/pkg 2677925
DPD Free Chlorine Reagent Powder Pillows, 10-mL 1000/pkg 2105528
DPD Free Chlorine Reagent Powder Pillows, 10-mL 300/pkg 2105503
Potassium Iodide, 30 g/L 100 mL 34332
Sodium Arsenite, 5 g/L 100 mL 104732
100 mL
Sodium Hydroxide Standard Solution, 1.0 N 104532
MDB
Standard Methods Book, most current edition each 2270800
Stoppers for 18-mm tube 25/pkg 173125
100 mL
Sulfuric Acid Standard Solution, 1 N 127032
MDB

Chlorine Dioxide, DPD Method (5.00 mg/L) 7

 FOR TECHNICAL ASSISTANCE, PRICE INFORMATION AND ORDERING: HACH COMPANY
In the U.S.A. – Call toll-free 800-227-4224 WORLD HEADQUARTERS
Outside the U.S.A. – Contact the HACH office or distributor serving you. Telephone: (970) 669-3050
On the Worldwide Web – www.hach.com; E-mail – [email protected] FAX: (970) 669-2932

© Hach Company/Hach Lange GmbH, 1989–2013. All rights reserved. 04/2013, Edition 8
Chlorine, Free DOC316.53.01023

USEPA DPD Method1 Method 8021

®
0.02 to 2.00 mg/L Cl2 Powder Pillows or AccuVac Ampuls
Scope and application: For testing free chlorine (hypochlorous acid and hypochlorite ion) in water, treated
waters, estuary and seawater. USEPA accepted for reporting for drinking water analyses.2 This product has not
been evaluated to test for chlorine and chloramines in medical applications in the United States.
1 Adapted from Standard Methods for the Examination of Water and Wastewater.
2 Procedure is equivalent to USEPA and Standard Method 4500-Cl G for drinking water.

Test preparation

Instrument-specific information
The tables in this section show all of the instruments that have the program for this test.
Table 1 shows sample cell and orientation requirements for reagent addition tests, such
as powder pillow or bulk reagent tests. Table 2 shows sample cell and adapter
requirements for AccuVac Ampul tests.
To use either table, select an instrument, then read across to find the corresponding
information for this test.
Table 1 Instrument-specific information for powder pillows
Instrument Sample cell orientation Sample cell
DR 6000 The fill line is to the right. 2495402
DR 3800
DR 2800
DR 2700
DR 5000 The fill line is toward the user.
DR 3900
DR 900 The orientation mark is toward the user. 2401906

Table 2 Instrument-specific information for AccuVac Ampuls

Instrument Adapter Sample cell
DR 6000 — 2427606
DR 5000
DR 900
DR 3900 LZV846 (A)
DR 3800 LZV584 (C) 2122800
DR 2800
DR 2700

1
Before starting
Samples must be analyzed immediately after collection and cannot be preserved for later analysis.
Install the instrument cap on the DR 900 cell holder before ZERO or READ is pushed.
Do not use the same sample cells for free and total chlorine. If trace iodide from the total chlorine reagent is carried over into
the free chlorine determination, monochloramine will interfere. It is best to use separate, dedicated sample cells for free and
total chlorine measurements.
If the test result is over-range, or if the sample temporarily turns yellow after the reagent addition, dilute the sample with a
known volume of high quality, chlorine demand-free water and repeat the test. Some loss of chlorine may occur due to the
dilution. Multiply the result by the dilution factor. Additional methods are available to measure chlorine without dilution.
For best results, measure the reagent blank value for each new lot of reagent. Replace the sample with deionized water in
the test procedure to get the reagent blank value. Subtract the reagent blank value from the sample results automatically
with the reagent blank adjust option.
An AccuVac Ampule for Blanks can be used to zero the instrument in the AccuVac test procedure.
Review the Safety Data Sheets (MSDS/SDS) for the chemicals that are used and use any recommended personal protective
equipment.
Dispose of reacted solutions according to local, state and federal regulations. Use the Safety Data Sheets for disposal
information for unused reagents. Consult the environmental, health and safety staff for your facility and/or local regulatory
agencies for further disposal information.
The SwifTest Dispenser for Free Chlorine can be used in place of the powder pillow in the test procedure.

Items to collect
Powder pillows

Description Quantity

DPD Free Chlorine Reagent Powder Pillows, 10-mL 1

Sample cells. (For information about sample cells, adapters or light shields, refer to Instrument-
2
specific information on page 1.)

Refer to Consumables and replacement items on page 6 for reorder information.

AccuVac Ampuls

Description Quantity

DPD Free Chlorine Reagent AccuVac Ampuls 1

Beaker, 50-mL 1
Sample cells (For information about sample cells, adapters or light shields, refer to Instrument-
1
specific information on page 1.)
Stopper for 18-mm tubes and AccuVac Ampuls 1

Refer to Consumables and replacement items on page 6 for reorder information.

Sample collection
• Samples must be analyzed immediately and cannot be preserved for later analysis.
• Chlorine is a strong oxidizing agent and it is unstable in natural waters. It reacts
quickly with various inorganic compounds and more slowly with organic compounds.
Many factors, including reactant concentrations, sunlight, pH, temperature and
salinity influence the decomposition of chlorine in water.
• Collect samples in clean glass bottles. Avoid plastic containers since these may have
a large chlorine demand.

2 Chlorine, Free, DPD Method (2.00 mg/L)

 • Pre-treat glass sample containers to remove any chlorine demand. Soak the
containers in a dilute bleach solution (1 mL commercial bleach to 1 liter of deionized
water) for at least 1 hour. Rinse thoroughly with deionized or distilled water. If sample
containers are rinsed thoroughly with deionized or distilled water after use, only
occasional pre-treatment is necessary.
• Be sure to get a representative sample. If the sample is taken from a spigot or faucet,
let the water flow for at least 5 minutes. Then let the container overflow with the
sample several times and then put the cap on the sample container so that there is
no headspace (air) above the sample. If a sample cell is used, rinse the cell several
times with the sample, then carefully fill to the 10-mL mark.

Powder pillow procedure

Start

1. Start program 2. Prepare the blank: Fill 3. Clean the prepared 4. Insert the blank into the
80 Chlorine F&T PP. For the sample cell with 10 mL sample. cell holder.
information about sample of sample.
cells, adapters or light
shields, refer to Instrument-
specific information
on page 1.
Note: Although the program
name may vary between
instruments, the program
number does not change.

Zero

5. Push ZERO. The display 6. Prepare the sample: Fill 7. Add the contents of one 8. Swirl the sample cell for
shows 0.00 mg/L. a second sample cell with powder pillow to the sample 20 seconds to mix.
10 mL of sample. cell. A pink color will develop if
chlorine is present. Proceed
to the next step
immediately.

Chlorine, Free, DPD Method (2.00 mg/L) 3

 Read

9. Clean the prepared 10. Within 60 seconds of 11. Push READ. Results
sample. adding the reagent, insert show in mg/L Cl2.
the prepared sample into
the cell holder.

AccuVac Ampul procedure

Start

1. Start program 2. Prepare the blank: Fill 3. Clean the blank. 4. Insert the blank into the
85 Chlorine F&T AV. For the sample cell with 10 mL cell holder.
information about sample of sample.
cells, adapters or light
shields, refer to Instrument-
specific information
on page 1.
Note: Although the program
name may vary between
instruments, the program
number does not change.

Zero

5. Push ZERO. The display 6. Prepare the sample: 7. Quickly invert the Ampul 8. Clean the AccuVac
shows 0.00 mg/L. Collect at least 40 mL of several times to mix. Ampul.
sample in a 50-mL beaker.
Fill the AccuVac Ampul with
sample. Keep the tip
immersed while the Ampul
fills completely.

4 Chlorine, Free, DPD Method (2.00 mg/L)

 Read

9. Within 60 seconds of 10. Push READ. Results

adding the reagent, insert show in mg/L Cl2.
the prepared sample
AccuVac Ampul into the cell
holder.

Interferences
Interfering substance Interference level
Acidity More than 150 mg/L CaCO3. The full color may not develop or the color may fade instantly.
Adjust to pH 6–7 with 1 N Sodium Hydroxide. Measure the amount to be added on a separate
sample aliquot, then add the same amount to the sample that is tested. Correct the test result
for the dilution from the volume addition.
Alkalinity More than 250 mg/L CaCO3. The full color may not develop or the color may fade instantly.
Adjust to pH 6–7 with 1 N Sulfuric Acid. Measure the amount to add on a separate sample
aliquot, then add the same amount to the sample that is tested. Correct the test result for the
dilution from the volume addition.
Bromine, Br2 Interferes at all levels
Chlorine Dioxide, ClO2 Interferes at all levels
Chloramines, organic May interfere
Hardness No effect at less than 1000 mg/L as CaCO3
Iodine, I2 Interferes at all levels
Manganese, Oxidized Pre-treat the sample as follows:
(Mn4+, Mn7+) or Chromium,
Oxidized (Cr6+) 1. Adjust the sample pH to 6–7.
2. Add 3 drops of Potassium Iodide (30-g/L) to 10 mL of sample.
3. Mix and wait 1 minute.
4. Add 3 drops of Sodium Arsenite (5-g/L) and mix.
5. Use the test procedure to measure the concentration of the treated sample.
6. Subtract this result from the result without the treatment to obtain the correct chlorine
concentration.

Monochloramine Causes a gradual drift to higher readings. When read within 1 minute after reagent addition,
3 mg/L monochloramine causes less than a 0.1 mg/L increase in the reading.
Ozone Interferes at all levels
Peroxides May interfere
Highly buffered samples or Can prevent the correct pH adjustment of the sample by the reagents. Sample pretreatment
extreme sample pH may be necessary. Adjust to pH 6–7 with acid (Sulfuric Acid, 1.000 N) or base (Sodium
Hydroxide, 1.00 N).

Pollution prevention and waste management

If sodium arsenite was added to the sample for manganese or chromium interferences,
the reacted samples will contain arsenic and must be disposed of as a hazardous waste.
Dispose of reacted solutions according to local, state and federal regulations.

Chlorine, Free, DPD Method (2.00 mg/L) 5

Accuracy check
Standard additions method (sample spike)
Use the standard additions method (for applicable instruments) to validate the test
procedure, reagents and instrument and to find if there is an interference in the sample.
Items to collect:
• Chlorine Standard Solution, 2-mL PourRite® Ampule, 25–30 mg/L (use mg/L on label)
• Breaker, PourRite Ampules
• Pipet, TenSette®, 0.1–1.0 mL and tips

1. Use the test procedure to measure the concentration of the sample, then keep the
(unspiked) sample in the instrument.
2. Go to the Standard Additions option in the instrument menu.
3. Select the values for standard concentration, sample volume and spike volumes.
4. Open the standard solution.
5. Prepare three spiked samples: use the TenSette pipet to add 0.1 mL, 0.2 mL and
0.3 mL of the standard solution, respectively, to three 10-mL portions of fresh sample.
Mix well.
®
Note: For AccuVac Ampuls, add 0.4 mL, 0.8 mL and 1.2 mL of the standard solution to three
50-mL portions of fresh sample.
6. Use the test procedure to measure the concentration of each of the spiked samples.
Start with the smallest sample spike. Measure each of the spiked samples in the
instrument.
7. Select Graph to compare the expected results to the actual results.
Note: If the actual results are significantly different from the expected results, make sure that
the sample volumes and sample spikes are measured accurately. The sample volumes and
sample spikes that are used should agree with the selections in the standard additions menu. If
the results are not within acceptable limits, the sample may contain an interference.

Method performance
The method performance data that follows was derived from laboratory tests that were
measured on a spectrophotometer during ideal test conditions. Users may get different
results under different test conditions.
Program Standard Precision (95% Confidence Interval) Sensitivity
Concentration change per 0.010 Abs change
80 1.25 mg/L Cl2 1.23–1.27 mg/L Cl2 0.02 mg/L Cl2
85 1.25 mg/L Cl2 1.21–1.29 mg/L Cl2 0.02 mg/L Cl2

Summary of method
Chlorine in the sample as hypochlorous acid or hypochlorite ion (free chlorine or free
available chlorine) immediately reacts with DPD (N,N-diethyl-p-phenylenediamine)
indicator to form a pink color, the intensity of which is proportional to the chlorine
concentration. The measurement wavelength is 530 nm for spectrophotometers or
520 nm for colorimeters.
Consumables and replacement items
Required reagents

Description Quantity/Test Unit Item no.

DPD Free Chlorine Reagent Powder Pillow, 10-mL 1 100/pkg 2105569

OR
®
DPD Free Chlorine Reagent AccuVac Ampul 1 25/pkg 2502025

6 Chlorine, Free, DPD Method (2.00 mg/L)

Required apparatus

Description Quantity/Test Unit Item no.

AccuVac Snapper 1 each 2405200

Beaker, 50-mL 1 each 50041H
Stoppers for 18-mm tubes and AccuVac Ampuls 2 6/pkg 173106

Recommended standards

Description Unit Item no.

®
Chlorine Standard Solution, 2-mL PourRite Ampules, 25–30 mg/L 20/pkg 2630020

Optional reagents and apparatus

Description Unit Item no.

®
AccuVac vials for sample blanks 25/pkg 2677925
®
Ampule Breaker, PourRite ampules each 2484600
®
Ampule Breaker, Voluette ampules each 2196800
Water, Chlorine-demand Free 500 mL 2641549
Cylinder, mixing, 25-mL each 2088640
Cylinder, mixing, 50-mL each 189641
®
Chlorine Standard Solution, 2-mL PourRite Ampules, 50–75 mg/L 20/pkg 1426820
®
Chlorine Standard Solution, 10-mL Voluette Ampule, 50–75 mg/L 16/pkg 1426810
DPD Free Chlorine Reagent Powder Pillows, 10-mL 1000/pkg 2105528
DPD Free Chlorine Reagent Powder Pillows, 10-mL 300/pkg 2105503
DPD Free Chlorine Reagent, 10 mL, SwifTest™ Dispenser refill vial 250 tests 2105560
Paper, pH, 0–14 pH range 100/pkg 2601300
®
Pipet, TenSette , 0.1–1.0 mL each 1970001
Pipet tips for TenSette Pipet 1970001 50/pkg 2185696
Pipet tips for TenSette Pipet 1970001 1000/pkg 2185628
Potassium Iodide, 30 g/L 100 mL 34332
Sodium Arsenite, 5 g/L 100 mL 104732
100 mL
Sodium Hydroxide Standard Solution, 1.0 N 104532
MDB
SpecCheck™ Secondary Standard Kit, Chlorine DPD, 0-2.0 mg/L Set each 2635300
100 mL
Sulfuric Acid Standard Solution, 1 N 127032
MDB

Chlorine, Free, DPD Method (2.00 mg/L) 7

 FOR TECHNICAL ASSISTANCE, PRICE INFORMATION AND ORDERING: HACH COMPANY
In the U.S.A. – Call toll-free 800-227-4224 WORLD HEADQUARTERS
Outside the U.S.A. – Contact the HACH office or distributor serving you. Telephone: (970) 669-3050
On the Worldwide Web – www.hach.com; E-mail – [email protected] FAX: (970) 669-2932

© Hach Company/Hach Lange GmbH, 1989–2013. All rights reserved. 04/2013, Edition 8
Chlorine, Free DOC316.53.01025

DPD Method1 Method 10069

0.1 to 10.0 mg/L Cl2 (HR) Powder Pillows
Scope and application: For determinations of higher levels of free chlorine (hypochlorous acid and hypochlorite
ion) in drinking water, cooling water and industrial process waters. This product has not been evaluated to test for
chlorine and chloramines in medical applications in the United States.
1 Adapted from Standard Methods for the Examination of Water and Wastewater.

Test preparation

Instrument specific information

Table 1 shows all of the instruments that have the program for this test. The table also
shows requirements that may vary between instruments, such as adapter and sample cell
requirements. To use this table, select an instrument, then read across to find the
corresponding information for this test.
Table 1 Instrument-specific information
Instrument Adapter Sample cell orientation Sample cell
DR 6000 A23618 The orientation key is toward the user. 4864302
DR 5000 The orientation key is toward the user.
DR 3900 LZV846 (A) The orientation key is away from the user.
DR 900 — The orientation key is toward the user.
DR 3800 LZV585 (B) The 1-cm path is aligned with the arrow on the adapter. 5940506
DR 2800
DR 2700

Before starting
Samples must be analyzed immediately after collection and cannot be preserved for later analysis.
Install the instrument cap on the DR 900 cell holder before ZERO or READ is pushed.
In bright light conditions (e.g., direct sunlight), close the cell compartment on spectrophotometers, if applicable, with the
protective cover during measurements.
If the chlorine concentration is less than 2 mg/L, use Method 8021, program number 80.
Review the Safety Data Sheets (MSDS/SDS) for the chemicals that are used and use any recommended personal protective
equipment.
Dispose of reacted solutions according to local, state and federal regulations. Use the Safety Data Sheets for disposal
information for unused reagents. Consult the environmental, health and safety staff for your facility and/or local regulatory
agencies for further disposal information.

Items to collect
Description Quantity

DPD Free Chlorine Reagent Powder Pillows, 25-mL 1

Sample cells (For information about sample cells, adapters or light shields, refer to Instrument
2
specific information on page 1.)

1
Refer to Consumables and replacement items on page 5 for reorder information.

Sample collection
• Samples must be analyzed immediately and cannot be preserved for later analysis.
• Chlorine is a strong oxidizing agent and it is unstable in natural waters. It reacts
quickly with various inorganic compounds and more slowly with organic compounds.
Many factors, including reactant concentrations, sunlight, pH, temperature and
salinity influence the decomposition of chlorine in water.
• Collect samples in clean glass bottles. Avoid plastic containers since these may have
a large chlorine demand.
• Pre-treat glass sample containers to remove any chlorine demand. Soak the
containers in a dilute bleach solution (1 mL commercial bleach to 1 liter of deionized
water) for at least 1 hour. Rinse thoroughly with deionized or distilled water. If sample
containers are rinsed thoroughly with deionized or distilled water after use, only
occasional pre-treatment is necessary.
• Be sure to get a representative sample. If the sample is taken from a spigot or faucet,
let the water flow for at least 5 minutes. Then let the container overflow with the
sample several times and then put the cap on the sample container so that there is
no headspace (air) above the sample. If a sample cell is used, rinse the cell several
times with the sample, then carefully fill to the 10-mL mark.

Powder pillow procedure

Start

1. Start program 2. Prepare the blank: Fill a 3. Clean the blank. 4. Insert the blank into the
88 Chlorine F&T HR. For sample cell to the 5-mL cell holder.
information about sample mark with sample.
cells, adapters or light
shields, refer to Instrument
specific information
on page 1.
Note: Although the program
name may vary between
instruments, the program
number does not change.

2 Chlorine, Free, DPD Method (10 mg/L)

 Zero

5. Push ZERO. The display 6. Prepare the sample: Fill 7. Add the contents of one 8. Close the sample cell.
shows 0.0 mg/L Cl2. a second sample cell to the DPD Free Chlorine Powder Shake the sample cell for
5-mL mark with sample. Pillow for 25-mL samples to about 20 seconds to
the sample. dissolve the reagent. A pink
color shows if chlorine is in
the sample.

Read

9. Clean the sample cell. 10. Insert the prepared 11. Push READ. Results
sample into the cell holder. show in mg/L Cl2.

Interferences
Interfering substance Interference level
Acidity More than 150 mg/L CaCO3. The full color may not develop or the color may fade instantly. Adjust
to pH 6–7 with 1 N Sodium Hydroxide. Measure the amount to be added on a separate sample
aliquot, then add the same amount to the sample that is tested. Correct the test result for the
dilution from the volume addition.
Alkalinity More than 250 mg/L CaCO3. The full color may not develop or the color may fade instantly. Adjust
to pH 6–7 with 1 N Sulfuric Acid. Measure the amount to add on a separate sample aliquot, then
add the same amount to the sample that is tested. Correct the test result for the dilution from the
volume addition.
Bromine, Br2 Interferes at all levels
Chlorine Dioxide, ClO2 Interferes at all levels
Chloramines, organic May interfere
Hardness No effect at less than 1000 mg/L as CaCO3
Iodine, I2 Interferes at all levels
Manganese, Oxidized Pre-treat the sample as follows:
(Mn4+, Mn7+) or
Chromium, Oxidized 1. Adjust the sample pH to 6–7.
(Cr6+) 2. Add 3 drops of Potassium Iodide (30-g/L) to 10 mL of sample.
3. Mix and wait 1 minute.
4. Add 3 drops of Sodium Arsenite (5-g/L) and mix.
5. Use the test procedure to measure the concentration of the treated sample.
6. Subtract this result from the result without the treatment to obtain the correct chlorine
concentration.

Chlorine, Free, DPD Method (10 mg/L) 3

Interfering substance Interference level
Ozone Interferes at all levels
Peroxides May interfere
Highly buffered samples Can prevent the correct pH adjustment of the sample by the reagents. Sample pretreatment may
or extreme sample pH be necessary. Adjust to pH 6–7 with acid (Sulfuric Acid, 1.000 N) or base (Sodium Hydroxide,
1.00 N).

Monochloramine interference
For conventional free chlorine disinfection (beyond the breakpoint), typical
monochloramine concentrations are very low. If monochloramine is present in the
sample, its interference in the free chlorine test depends on the sample temperature,
relative amount of monochloramine to free chlorine and the time required to do the
analysis. Typical interference levels of monochloramine as mg/L Cl2 in the free chlorine
test are shown in Table 2 (1 minute test time). Measure the monochloramine levels with
method 10200 for Chloramine (Mono) and Free Ammonia.
Table 2 Monochloramine interference at different sample temperatures
NH2Cl (as Cl2) 5 °C (41 °F) 10 °C (50 °F) 20 °C (68 °F) 30 °C (83 °F)
1.2 mg/L 0.15 0.19 0.30 0.29
2.2 mg/L 0.35 0.38 0.55 0.61
3.2 mg/L 0.38 0.56 0.69 0.73

Pollution prevention and waste management

If sodium arsenite was added to the sample for manganese or chromium interferences,
the reacted samples will contain arsenic and must be disposed of as a hazardous waste.
Dispose of reacted solutions according to local, state and federal regulations.
Accuracy check
Standard additions method (sample spike)
Use the standard additions method (for applicable instruments) to validate the test
procedure, reagents and instrument and to find if there is an interference in the sample.
Items to collect:
• Chlorine Standard Solution, 2-mL PourRite® Ampule, 50–75 mg/L (use mg/L on label)
• Breaker, PourRite Ampules
• Pipet, TenSette®, 0.1–1.0 mL and tips
• Mixing cylinders, 10-mL (3)

1. Use the test procedure to measure the concentration of the sample, then keep the
(unspiked) sample in the instrument.
2. Go to the Standard Additions option in the instrument menu.
3. Select the values for standard concentration, sample volume and spike volumes.
4. Open the standard solution.
5. Prepare three spiked samples: use the TenSette pipet to add 0.1 mL, 0.2 mL and
0.3 mL of the standard solution, respectively, to three 5-mL portions of fresh sample.
Mix well.
6. Use the test procedure to measure the concentration of each of the spiked samples.
Start with the smallest sample spike. Measure each of the spiked samples in the
instrument.
7. Select Graph to compare the expected results to the actual results.
Note: If the actual results are significantly different from the expected results, make sure that
the sample volumes and sample spikes are measured accurately. The sample volumes and

4 Chlorine, Free, DPD Method (10 mg/L)

 sample spikes that are used should agree with the selections in the standard additions menu. If
the results are not within acceptable limits, the sample may contain an interference.

Method performance
The method performance data that follows was derived from laboratory tests that were
measured on a spectrophotometer during ideal test conditions. Users may get different
results under different test conditions.
Program Standard Precision (95% Confidence Interval) Sensitivity
Concentration change per 0.010 Abs change
88 5.4 mg/L Cl2 5.3–5.5 mg/L Cl2 0.04 mg/L Cl2

Summary of method
The range of analysis using the DPD method for free chlorine can be extended by adding
more indicator in proportion to sample volume. Thus, a larger fill powder pillow of DPD
Free Chlorine Reagent is added to a 5-mL sample portion. Chlorine in the sample as
hypochlorous acid or hypochlorite ion (free chlorine or free available chlorine)
immediately reacts with DPD (N,N-diethyl-p-phenylenediamine) indicator to form a pink
color, the intensity of which is proportional to the chlorine concentration. The
measurement wavelength is 530 nm for spectrophotometers or 520 nm for colorimeters.
Consumables and replacement items
Required reagents

Description Quantity/Test Unit Item no.

DPD Free Chlorine Reagent Powder Pillow, 25-mL 1 100/pkg 1407099

Recommended standards and apparatus

Description Unit Item no.

®
Ampule Breaker, PourRite ampules each 2484600
®
Ampule Breaker, Voluette ampules each 2196800
®
Chlorine Standard Solution, 2-mL PourRite Ampules, 50–75 mg/L 20/pkg 1426820
®
Chlorine Standard Solution, 2-mL PourRite Ampules, 25–30 mg/L 20/pkg 2630020
®
Chlorine Standard Solution, 10-mL Voluette Ampule, 50–75 mg/L 16/pkg 1426810
SpecCheck™ Gel Secondary Standard Kit, Chlorine DPD, 0–10 mg/L 4/pkg 2893300

Optional reagents and apparatus

Description Unit Item no.

Cylinder, mixing, 25-mL each 2088640

DPD Free Chlorine Reagent Powder Pillows, 10-mL 1000/pkg 2105528
DPD Free Chlorine Reagent Powder Pillows, 10-mL 300/pkg 2105503
Paper, pH, 0–14 pH range 100/pkg 2601300
®
Pipet, TenSette , 0.1–1.0 mL each 1970001
Pipet tips for TenSette Pipet 1970001 50/pkg 2185696
Pipet tips for TenSette Pipet 1970001 1000/pkg 2185628
Potassium Iodide, 30 g/L 100 mL 34332
Sodium Arsenite, 5 g/L 100 mL 104732

Chlorine, Free, DPD Method (10 mg/L) 5

Consumables and replacement items (continued)
Description Unit Item no.

100 mL
Sodium Hydroxide Standard Solution, 1.0 N 104532
MDB
100 mL
Sulfuric Acid Standard Solution, 1 N 127032
MDB
Test tube rack each 1864100
Thermometer, non-mercury, -10 to +225 °C each 2635700

FOR TECHNICAL ASSISTANCE, PRICE INFORMATION AND ORDERING: HACH COMPANY

In the U.S.A. – Call toll-free 800-227-4224 WORLD HEADQUARTERS
Outside the U.S.A. – Contact the HACH office or distributor serving you. Telephone: (970) 669-3050
On the Worldwide Web – www.hach.com; E-mail – [email protected] FAX: (970) 669-2932

© Hach Company/Hach Lange GmbH, 1989–2013. All rights reserved. 04/2013, Edition 8
Chlorine, Free DOC316.53.01303

USEPA DPD Method2 Method 102451

0.05 to 4.00 mg/L Cl2 (MR) Powder Pillows
Scope and application: For free chlorine (hypochlorous acid and hypochlorite ion) measurements in water,
treated waters, estuary and seawater. This product has not been evaluated to test for chlorine and chloramines in
medical applications in the United States.
1 USEPA accepted for reporting wastewater and drinking water analyses.
2 Procedure is equivalent to USEPA method 330.5 for wastewater and Standard Method 4500-Cl G for drinking water.

Test preparation

Instrument specific information

The table in this section shows all of the instruments that have the program for this test.
Table 1 shows sample cell and orientation requirements for reagent addition tests, such
as powder pillow or bulk reagent tests.
To use the table, select an instrument, then read across to find the corresponding
information for this test.
Table 1 Instrument-specific information for reagent addition
Instrument Sample cell orientation Sample cell
DR 6000 The fill line is to the right. 2495402
DR 3800
DR 2800
DR 2700
DR 5000 The fill line is toward the user.
DR 3900
DR 900 The orientation mark is toward the user. 2401906

Before starting
Samples must be analyzed immediately after collection and cannot be preserved for later analysis.
Install the instrument cap on the DR 900 cell holder before ZERO or READ is pushed.
Do not use the same sample cells for free and total chlorine. If trace iodide from the total chlorine reagent is carried over into
the free chlorine determination, monochloramine will interfere. It is best to use separate, dedicated sample cells for free and
total chlorine measurements.
If the test result is over-range, or if the sample temporarily turns yellow after the reagent addition, dilute the sample with a
known volume of high quality, chlorine demand-free water and repeat the test. Some loss of chlorine may occur due to the
dilution. Multiply the result by the dilution factor. Additional methods are available to measure chlorine without dilution.
For best results, measure the reagent blank value for each new lot of reagent. Replace the sample with deionized water in
the test procedure to get the reagent blank value. Subtract the reagent blank value from the sample results automatically
with the reagent blank adjust option.
Cold waters can cause condensation on the sample cells during color development. Examine the sample cells for
condensation before measurements.

1
Review the Safety Data Sheets (MSDS/SDS) for the chemicals that are used and use any recommended personal protective
equipment.
Dispose of reacted solutions according to local, state and federal regulations. Use the Safety Data Sheets for disposal
information for unused reagents. Consult the environmental, health and safety staff for your facility and/or local regulatory
agencies for further disposal information.

Items to collect
Description Quantity

DPD Free Chlorine Reagent Powder Pillows, 25-mL 1

Sample cells (For information about sample cells, adapters or light shields, refer to Instrument
2
specific information on page 1.)

Refer to Consumables and replacement items on page 5 for reorder information.

Sample collection
• Samples must be analyzed immediately and cannot be preserved for later analysis.
• Chlorine is a strong oxidizing agent and it is unstable in natural waters. It reacts
quickly with various inorganic compounds and more slowly with organic compounds.
Many factors, including reactant concentrations, sunlight, pH, temperature and
salinity influence the decomposition of chlorine in water.
• Collect samples in clean glass bottles. Avoid plastic containers since these may have
a large chlorine demand.
• Pre-treat glass sample containers to remove any chlorine demand. Soak the
containers in a dilute bleach solution (1 mL commercial bleach to 1 liter of deionized
water) for at least 1 hour. Rinse thoroughly with deionized or distilled water. If sample
containers are rinsed thoroughly with deionized or distilled water after use, only
occasional pre-treatment is necessary.
• Be sure to get a representative sample. If the sample is taken from a spigot or faucet,
let the water flow for at least 5 minutes. Then let the container overflow with the
sample several times and then put the cap on the sample container so that there is
no headspace (air) above the sample. If a sample cell is used, rinse the cell several
times with the sample, then carefully fill to the 10-mL mark.

2 Chlorine, Free, DPD Method (4.00 mg/L)

Powder pillow procedure

Start

1. Start program 2. Prepare the blank: Fill 3. Clean the blank. 4. Insert the blank into the
87 Chlorine, F&T PP MR. the sample cell with 10 mL cell holder.
For information about of sample.
sample cells, adapters or
light shields, refer to
Instrument specific
information on page 1.
Note: Although the program
name may vary between
instruments, the program
number does not change.

Zero

5. Push ZERO. The display 6. Prepare the sample: Fill 7. Add the contents of one 8. Close the prepared
shows 0.00 mg/L Cl2. a second sample cell with DPD Free Chlorine Powder sample cell. Invert the
10 mL of sample. Pillow for 25-mL sample to sample cell several times to
the prepared sample cell. mix. A pink color shows if
chlorine is present. Go to
the next step immediately.

Read

9. Clean the prepared 10. Within one minute of 11. Push READ. Results
sample. adding the reagent, insert show in mg/L Cl2.
the prepared sample into
the cell holder.

Chlorine, Free, DPD Method (4.00 mg/L) 3

Interferences
Interfering substance Interference level
Acidity More than 150 mg/L CaCO3. The full color may not develop or the color may fade instantly. Adjust
to pH 6–7 with 1 N Sodium Hydroxide. Measure the amount to be added on a separate sample
aliquot, then add the same amount to the sample that is tested. Correct the test result for the
dilution from the volume addition.
Alkalinity More than 250 mg/L CaCO3. The full color may not develop or the color may fade instantly. Adjust
to pH 6–7 with 1 N Sulfuric Acid. Measure the amount to add on a separate sample aliquot, then
add the same amount to the sample that is tested. Correct the test result for the dilution from the
volume addition.
Bromine, Br2 Interferes at all levels
Chlorine Dioxide, ClO2 Interferes at all levels
Chloramines, organic May interfere
Hardness No effect at less than 1000 mg/L as CaCO3
Iodine, I2 Interferes at all levels
Manganese, Oxidized Pre-treat the sample as follows:
(Mn4+, Mn7+) or
Chromium, Oxidized 1. Adjust the sample pH to 6–7.
(Cr6+) 2. Add 3 drops of Potassium Iodide (30-g/L) to 10 mL of sample.
3. Mix and wait 1 minute.
4. Add 3 drops of Sodium Arsenite (5-g/L) and mix.
5. Use the test procedure to measure the concentration of the treated sample.
6. Subtract this result from the result without the treatment to obtain the correct chlorine
concentration.

Ozone Interferes at all levels

Peroxides May interfere
Highly buffered samples Can prevent the correct pH adjustment of the sample by the reagents. Sample pretreatment may
or extreme sample pH be necessary. Adjust to pH 6–7 with acid (Sulfuric Acid, 1.000 N) or base (Sodium Hydroxide,
1.00 N).

Monochloramine interference
For conventional free chlorine disinfection (beyond the breakpoint), typical
monochloramine concentrations are very low. If monochloramine is present in the
sample, its interference in the free chlorine test depends on the sample temperature,
relative amount of monochloramine to free chlorine and the time required to do the
analysis. Typical interference levels of monochloramine as mg/L Cl2 in the free chlorine
test are shown in Table 2 (1 minute test time). Measure the monochloramine levels with
method 10200 for Chloramine (Mono) and Free Ammonia.
Table 2 Monochloramine interference at different sample temperatures
NH2Cl (as Cl2) 5 °C (41 °F) 10 °C (50 °F) 20 °C (68 °F) 30 °C (83 °F)
1.2 mg/L 0.15 0.19 0.30 0.29
2.2 mg/L 0.35 0.38 0.55 0.61
3.2 mg/L 0.38 0.56 0.69 0.73

Pollution prevention and waste management

If sodium arsenite was added to the sample for manganese or chromium interferences,
the reacted samples will contain arsenic and must be disposed of as a hazardous waste.
Dispose of reacted solutions according to local, state and federal regulations.

4 Chlorine, Free, DPD Method (4.00 mg/L)

Accuracy check
Standard additions method (sample spike)
Use the standard additions method (for applicable instruments) to validate the test
procedure, reagents and instrument and to find if there is an interference in the sample.
Items to collect:
• Chlorine Standard Solution, 2-mL PourRite® Ampule, 25–30 (or 50–75) mg/L
• Ampule breaker
• Pipet, TenSette®, 0.1–1.0 mL and tips

1. Use the test procedure to measure the concentration of the sample, then keep the
(unspiked) sample in the instrument.
2. Go to the Standard Additions option in the instrument menu.
3. Select the values for standard concentration, sample volume and spike volumes.
4. Open the standard solution.
5. Prepare three spiked samples: use the TenSette pipet to add 0.1 mL, 0.2 mL and
0.3 mL of the standard solution, respectively, to three 10-mL portions of fresh sample.
Mix well.
6. Use the test procedure to measure the concentration of each of the spiked samples.
Start with the smallest sample spike. Measure each of the spiked samples in the
instrument.
7. Select Graph to compare the expected results to the actual results.
Note: If the actual results are significantly different from the expected results, make sure that
the sample volumes and sample spikes are measured accurately. The sample volumes and
sample spikes that are used should agree with the selections in the standard additions menu. If
the results are not within acceptable limits, the sample may contain an interference.

Verification of on-line analyzers

This procedure can be used to meet the requirements of USEPA Method 334.0 -
Determination of Residual Chlorine in Drinking Water Using an On-line Chlorine Analyzer.
The procedure and requirements for compliance with EPA Method 334.0 can be
downloaded directly from http://www.hach.com/method334.
Method performance
The method performance data that follows was derived from laboratory tests that were
measured on a spectrophotometer during ideal test conditions. Users may get different
results under different test conditions.
Program Standard Precision (95% Confidence Interval) Sensitivity
Concentration change per 0.010 Abs change
89 2.68 mg/L Cl2 2.63–2.73 mg/L Cl2 0.03 mg/L Cl2

Summary of method
Chlorine in the sample as hypochlorous acid or hypochlorite ion (free chlorine or free
available chlorine) immediately reacts with DPD (N,N-diethyl-p-phenylenediamine)
indicator to form a pink color, the intensity of which is proportional to the chlorine
concentration. The measurement wavelength is 530 nm for spectrophotometers or
520 nm for colorimeters.
Consumables and replacement items
Required reagents

Description Quantity/test Unit Item no.

DPD Free Chlorine Reagent Powder Pillow, 25-mL 1 100/pkg 1407099

Sample cell, 10 mL round, 25 x 60 mm 1 6/pkg 2427606

Chlorine, Free, DPD Method (4.00 mg/L) 5

Recommended standards

Description Unit Item no.

®
Chlorine Standard Solution, 2-mL PourRite Ampules, 25–30 mg/L 20/pkg 2630020
®
Ampule Breaker, Voluette ampules each 2196800
®
PourRite Ampule Breaker, 2-mL each 2484600
®
Chlorine Standard Solution, 2-mL PourRite Ampule, 50-75 mg/L 20/pkg 1426820
®
Chlorine Standard Solution, 10-mL Voluette Ampule, 50–75 mg/L 16/pkg 1426810

Optional reagents and apparatus

Description Unit Item no.

100 mL
Sodium Hydroxide Standard Solution, 1.0 N 104532
MDB
100 mL
Sulfuric Acid Standard Solution, 1 N 127032
MDB
Potassium Iodide, 30 g/L 100 mL 34332
Sodium Arsenite, 5 g/L 100 mL 104732
®
Pipet, TenSette , 0.1–1.0 mL each 1970001
Pipet tips for TenSette Pipet 1970001 50/pkg 2185696
Pipet tips for TenSette Pipet 1970001 1000/pkg 2185628
Paper, pH, 0–14 pH range 100/pkg 2601300
DPD Free Chlorine Reagent Powder Pillows, 25-mL 1000/pkg 1407028
SpecCheck™ Secondary Standard Kit, Chlorine DPD, MR each 2980500
Organic-free water 500 mL 2641549
50 mL
Freechlor F Reagent Solution 2964926
SCDB
Monochlor F Reagent Powder Pillows 100/pkg 2802299

FOR TECHNICAL ASSISTANCE, PRICE INFORMATION AND ORDERING: HACH COMPANY

In the U.S.A. – Call toll-free 800-227-4224 WORLD HEADQUARTERS
Outside the U.S.A. – Contact the HACH office or distributor serving you. Telephone: (970) 669-3050
On the Worldwide Web – www.hach.com; E-mail – [email protected] FAX: (970) 669-2932

© Hach Company/Hach Lange GmbH, 1989–2013. All rights reserved. 04/2013, Edition 8
Chlorine, Free DOC316.53.01024

DPD Method1 Method 10102

0.09 to 5.00 mg/L Cl2 Test 'N Tube™ Vials
Scope and application: For testing higher levels of free chlorine (hypochlorous acid and hypochlorite ion) in
drinking water, cooling water and industrial process water. This product has not been evaluated to test for chlorine
and chloramines in medical applications in the United States.
1 Adapted from Standard Methods for the Examination of Water and Wastewater.

Test preparation

Instrument specific information

The table in this section shows all of the instruments that have the program for this test.
Table 1 shows adapter and light shield requirements for the instruments that use them.
To use the table, select an instrument, then read across to find the corresponding
information for this test.
Table 1 Instrument-specific information for test tubes
Instrument Adapters Light shield
DR 6000 — —
DR 5000 — —
DR 900 4846400 Cover supplied with the instrument
DR 3900 — LZV849
DR 3800 — LZV646
DR 2800 —

Before starting
Samples must be analyzed immediately after collection and cannot be preserved for later analysis.
Install the instrument cap on the DR 900 cell holder before ZERO or READ is pushed.
DR 3900, DR 3800, DR 2800 and DR 2700: Install the light shield in Cell Compartment #2 before this test is started.
For best results, measure the reagent blank value for each new lot of reagent. Replace the sample with deionized water in
the test procedure to get the reagent blank value. Subtract the reagent blank value from the sample results automatically
with the reagent blank adjust option.
Review the Safety Data Sheets (MSDS/SDS) for the chemicals that are used and use any recommended personal protective
equipment.
Dispose of reacted solutions according to local, state and federal regulations. Use the Safety Data Sheets for disposal
information for unused reagents. Consult the environmental, health and safety staff for your facility and/or local regulatory
agencies for further disposal information.

Items to collect
Description Quantity

Light shield (For information about sample cells, adapters or light shields, refer to Instrument
1
specific information on page 1.)
Test 'N Tube™ DPD Free Chlorine Reagent 1
Wipes, disposable varies

1
Refer to Consumables and replacement items on page 5 for reorder information.

Sample collection
• Samples must be analyzed immediately and cannot be preserved for later analysis.
• Chlorine is a strong oxidizing agent and it is unstable in natural waters. It reacts
quickly with various inorganic compounds and more slowly with organic compounds.
Many factors, including reactant concentrations, sunlight, pH, temperature and
salinity influence the decomposition of chlorine in water.
• Collect samples in clean glass bottles. Avoid plastic containers since these may have
a large chlorine demand.
• Pre-treat glass sample containers to remove any chlorine demand. Soak the
containers in a dilute bleach solution (1 mL commercial bleach to 1 liter of deionized
water) for at least 1 hour. Rinse thoroughly with deionized or distilled water. If sample
containers are rinsed thoroughly with deionized or distilled water after use, only
occasional pre-treatment is necessary.
• Be sure to get a representative sample. If the sample is taken from a spigot or faucet,
let the water flow for at least 5 minutes. Then let the container overflow with the
sample several times and then put the cap on the sample container so that there is
no headspace (air) above the sample. If a sample cell is used, rinse the cell several
times with the sample, then carefully fill to the 10-mL mark.

Test 'N Tube procedure

Start

1. Start program 2. Prepare the blank: Fill 3. Clean the blank vial. 4. Insert the blank vial into
89 Chlorine F&T TNT. For an empty Test 'N Tube vial the 16-mm cell holder.
information about sample to the top of the label with
cells, adapters or light sample.
shields, refer to Instrument
specific information
on page 1.
Note: Although the program
name may vary between
instruments, the program
number does not change.

2 Chlorine, Free, DPD TNT Method (5.00 mg/L)

 Zero

5. Push ZERO. The display 6. Prepare the sample: 7. Clean the sample vial. 8. Insert the sample vial
shows 0.00 mg/L Cl2. Remove the cap from a into the 16-mm cell holder.
Free Chlorine DPD Test 'N
Tube. Fill the vial to the top
of the label with sample.

Read

9. Push READ. Results

show in mg/L Cl2.

Interferences
Interfering substance Interference level
Acidity More than 150 mg/L CaCO3. The full color may not develop or the color may fade instantly. Adjust
to pH 6–7 with 1 N Sodium Hydroxide. Measure the amount to be added on a separate sample
aliquot, then add the same amount to the sample that is tested. Correct the test result for the
dilution from the volume addition.
Alkalinity More than 250 mg/L CaCO3. The full color may not develop or the color may fade instantly. Adjust
to pH 6–7 with 1 N Sulfuric Acid. Measure the amount to add on a separate sample aliquot, then
add the same amount to the sample that is tested. Correct the test result for the dilution from the
volume addition.
Bromine, Br2 Interferes at all levels
Chlorine Dioxide, ClO2 Interferes at all levels
Chloramines, organic May interfere
Hardness No effect at less than 1000 mg/L as CaCO3
Iodine, I2 Interferes at all levels
Manganese, Oxidized Pre-treat the sample as follows:
(Mn4+, Mn7+) or
Chromium, Oxidized 1. Adjust the sample pH to 6–7.
(Cr6+) 2. Add 3 drops of Potassium Iodide (30-g/L) to 10 mL of sample.
3. Mix and wait 1 minute.
4. Add 3 drops of Sodium Arsenite (5-g/L) and mix.
5. Use the test procedure to measure the concentration of the treated sample.
6. Subtract this result from the result without the treatment to obtain the correct chlorine
concentration.

Chlorine, Free, DPD TNT Method (5.00 mg/L) 3

Interfering substance Interference level
Ozone Interferes at all levels
Peroxides May interfere
Highly buffered samples Can prevent the correct pH adjustment of the sample by the reagents. Sample pretreatment may
or extreme sample pH be necessary. Adjust to pH 6–7 with acid (Sulfuric Acid, 1.000 N) or base (Sodium Hydroxide,
1.00 N).

Monochloramine interference
For conventional free chlorine disinfection (beyond the breakpoint), typical
monochloramine concentrations are very low. If monochloramine is present in the
sample, its interference in the free chlorine test depends on the sample temperature,
relative amount of monochloramine to free chlorine and the time required to do the
analysis. Typical interference levels of monochloramine as mg/L Cl2 in the free chlorine
test are shown in Table 2 (1 minute test time). Measure the monochloramine levels with
method 10200 for Chloramine (Mono) and Free Ammonia.
Table 2 Monochloramine interference at different sample temperatures
NH2Cl (as Cl2) 5 °C (41 °F) 10 °C (50 °F) 20 °C (68 °F) 30 °C (83 °F)
1.2 mg/L 0.15 0.19 0.30 0.29
2.2 mg/L 0.35 0.38 0.55 0.61
3.2 mg/L 0.38 0.56 0.69 0.73

Pollution prevention and waste management

If sodium arsenite was added to the sample for manganese or chromium interferences,
the reacted samples will contain arsenic and must be disposed of as a hazardous waste.
Dispose of reacted solutions according to local, state and federal regulations.
Accuracy check
Standard additions method (sample spike)
Use the standard additions method (for applicable instruments) to validate the test
procedure, reagents and instrument and to find if there is an interference in the sample.
Items to collect:
• Chlorine Standard Solution, 2-mL PourRite® Ampule, 50–75 mg/L (use mg/L on label)
• Breaker, PourRite Ampules
• Pipet, TenSette®, 0.1–1.0 mL and tips

1. Use the test procedure to measure the concentration of the sample, then keep the
(unspiked) sample in the instrument.
2. Go to the Standard Additions option in the instrument menu.
3. Select the values for standard concentration, sample volume and spike volumes.
4. Open the standard solution.
5. Prepare three spiked samples: use the TenSette pipet to add 0.1 mL, 0.2 mL and
0.3 mL of the standard solution, respectively, to three 10-mL portions of fresh sample.
Mix well.
6. Use the test procedure to measure the concentration of each of the spiked samples.
Start with the smallest sample spike. Measure each of the spiked samples in the
instrument.
7. Select Graph to compare the expected results to the actual results.
Note: If the actual results are significantly different from the expected results, make sure that
the sample volumes and sample spikes are measured accurately. The sample volumes and
sample spikes that are used should agree with the selections in the standard additions menu. If
the results are not within acceptable limits, the sample may contain an interference.

4 Chlorine, Free, DPD TNT Method (5.00 mg/L)

Method performance
The method performance data that follows was derived from laboratory tests that were
measured on a spectrophotometer during ideal test conditions. Users may get different
results under different test conditions.
Program Standard Precision (95% Confidence Interval) Sensitivity
Concentration change per 0.010 Abs change
89 2.68 mg/L Cl2 2.63–2.73 mg/L Cl2 0.03 mg/L Cl2

Summary of method
Chlorine in the sample as hypochlorous acid or hypochlorite ion (free chlorine or free
available chlorine) immediately reacts with DPD (N,N-diethyl-p-phenylenediamine)
indicator to form a pink color, the intensity of which is proportional to the chlorine
concentration. The measurement wavelength is 530 nm for spectrophotometers or
520 nm for colorimeters.
Consumables and replacement items
Required reagents

Description Quantity/Test Unit Item no.

Test 'N Tube™ DPD Free Chlorine Reagent 1 50/pkg 2105545

Recommended standards and apparatus

Description Unit Item no.

®
Chlorine Standard Solution, 10-mL Voluette Ampule, 50–75 mg/L 16/pkg 1426810
®
Chlorine Standard Solution, 2-mL PourRite Ampules, 50–75 mg/L 20/pkg 1426820
®
Chlorine Standard Solution, 2-mL PourRite Ampules, 25–30 mg/L 20/pkg 2630020
®
Ampule Breaker, PourRite ampules each 2484600
®
Ampule Breaker, Voluette ampules each 2196800

Optional reagents and apparatus

Description Unit Item no.

Paper, pH, 0–14 pH range 100/pkg 2601300

®
Pipet, TenSette , 0.1–1.0 mL each 1970001
Pipet tips for TenSette Pipet 1970001 50/pkg 2185696
Pipet tips for TenSette Pipet 1970001 1000/pkg 2185628
Potassium Iodide, 30 g/L 100 mL 34332
Sodium Arsenite, 5 g/L 100 mL 104732
100 mL
Sodium Hydroxide Standard Solution, 1.0 N 104532
MDB
100 mL
Sulfuric Acid Standard Solution, 1 N 127032
MDB
Test tube rack each 1864100
Thermometer, non-mercury, -10 to +225 °C each 2635700
Wipes, disposable 280/pkg 2097000

Chlorine, Free, DPD TNT Method (5.00 mg/L) 5

 FOR TECHNICAL ASSISTANCE, PRICE INFORMATION AND ORDERING: HACH COMPANY
In the U.S.A. – Call toll-free 800-227-4224 WORLD HEADQUARTERS
Outside the U.S.A. – Contact the HACH office or distributor serving you. Telephone: (970) 669-3050
On the Worldwide Web – www.hach.com; E-mail – [email protected] FAX: (970) 669-2932

© Hach Company/Hach Lange GmbH, 1989–2013. All rights reserved. 04/2013, Edition 8
Chlorine, Total DOC316.53.01027

USEPA DPD Method1 Method 8167

®
0.02 to 2.00 mg/L Cl2 Powder Pillows or AccuVac Ampuls
Scope and application: For testing residual chlorine and chloramines in water, wastewater, estuary water and
seawater; USEPA-accepted for reporting for drinking and wastewater analyses.2 This product has not been
evaluated to test for chlorine and chloramines in medical applications in the United States.
1 Adapted from Standard Methods for the Examination of Water and Wastewater.
2 Procedure is equivalent to USEPA and Standard Method 4500-Cl G for drinking water and wastewater analysis.

Test preparation

Instrument-specific information
The tables in this section show all of the instruments that have the program for this test.
Table 1 shows sample cell and orientation requirements for reagent addition tests, such
as powder pillow or bulk reagent tests. Table 2 shows sample cell and adapter
requirements for AccuVac Ampul tests.
To use either table, select an instrument, then read across to find the corresponding
information for this test.
Table 1 Instrument-specific information for powder pillows
Instrument Sample cell orientation Sample cell
DR 6000 The fill line is to the right. 2495402
DR 3800
DR 2800
DR 2700
DR 5000 The fill line is toward the user.
DR 3900
DR 900 The orientation mark is toward the user. 2401906

Table 2 Instrument-specific information for AccuVac Ampuls

Instrument Adapter Sample cell
DR 6000 — 2427606
DR 5000
DR 900
DR 3900 LZV846 (A)
DR 3800 LZV584 (C) 2122800
DR 2800
DR 2700

1
Before starting
Samples must be analyzed immediately after collection and cannot be preserved for later analysis.
Install the instrument cap on the DR 900 cell holder before ZERO or READ is pushed.
If the test result is over-range, or if the sample temporarily turns yellow after the reagent addition, dilute the sample with a
known volume of high quality, chlorine demand-free water and repeat the test. Some loss of chlorine may occur due to the
dilution. Multiply the result by the dilution factor. Additional methods are available to measure chlorine without dilution.
For chloramination disinfection control, use one of the available Chloramine (Mono) methods.
For best results, measure the reagent blank value for each new lot of reagent. Replace the sample with deionized water in
the test procedure to get the reagent blank value. Subtract the reagent blank value from the sample results automatically
with the reagent blank adjust option.
Review the Safety Data Sheets (MSDS/SDS) for the chemicals that are used and use any recommended personal protective
equipment.
Dispose of reacted solutions according to local, state and federal regulations. Use the Safety Data Sheets for disposal
information for unused reagents. Consult the environmental, health and safety staff for your facility and/or local regulatory
agencies for further disposal information.
The SwifTest Dispenser for Total Chlorine can be used in place of the powder pillow in the test procedure.
An AccuVac Ampule for Blanks can be used to zero the instrument in the AccuVac test procedure.

Items to collect
Powder pillows

Description Quantity

DPD Total Chlorine Reagent Powder Pillow, 10-mL 1

Sample cells. (For information about sample cells, adapters or light shields, refer to Instrument-
2
specific information on page 1.)

Refer to Consumables and replacement items on page 7 for reorder information.

AccuVac Ampuls

Description Quantity
®
DPD Total Chlorine Reagent AccuVac Ampul 1
Beaker, 50-mL 1
Sample cells (For information about sample cells, adapters or light shields, refer to Instrument-
1
specific information on page 1.)
Stopper for 18-mm tubes and AccuVac Ampuls 1

Refer to Consumables and replacement items on page 7 for reorder information.

Sample collection
• Samples must be analyzed immediately and cannot be preserved for later analysis.
• Chlorine is a strong oxidizing agent and it is unstable in natural waters. It reacts
quickly with various inorganic compounds and more slowly with organic compounds.
Many factors, including reactant concentrations, sunlight, pH, temperature and
salinity influence the decomposition of chlorine in water.
• Collect samples in clean glass bottles. Avoid plastic containers since these may have
a large chlorine demand.
• Pre-treat glass sample containers to remove any chlorine demand. Soak the
containers in a dilute bleach solution (1 mL commercial bleach to 1 liter of deionized
water) for at least 1 hour. Rinse thoroughly with deionized or distilled water. If sample

2 Chlorine, Total, DPD Method (2.00 mg/L)

 containers are rinsed thoroughly with deionized or distilled water after use, only
occasional pre-treatment is necessary.
• Be sure to get a representative sample. If the sample is taken from a spigot or faucet,
let the water flow for at least 5 minutes. Then let the container overflow with the
sample several times and then put the cap on the sample container so that there is
no headspace (air) above the sample. If a sample cell is used, rinse the cell several
times with the sample, then carefully fill to the 10-mL mark.

Powder pillow procedure

Start

1. Start program 2. Fill a sample cell with 3. Prepare the sample: 4. Swirl the sample cell for
80 Chlorine F&T PP. For 10 mL of sample. Add the contents of one 20 seconds to mix.
information about sample powder pillow to the sample A pink color shows if
cells, adapters or light cell. chlorine is present in the
shields, refer to Instrument- sample.
specific information
on page 1.
Note: Although the program
name may vary between
instruments, the program
number does not change.

5. Start the instrument 6. Prepare the blank: Fill a 7. Clean the blank. 8. Insert the blank into the
timer. A 3-minute reaction second sample cell with cell holder.
time starts. 10 mL of sample.
Prepare the sample blank
and set the instrument to
zero during the reaction
time.

Chlorine, Total, DPD Method (2.00 mg/L) 3

 Zero Read

9. Push ZERO. The display 10. Clean the prepared 11. Within 3 minutes after 12. Push READ. Results
shows 0.00 mg/L Cl2. sample. the timer expires, insert the show in mg/L Cl2.
prepared sample into the
cell holder.

AccuVac Ampul procedure

Start

1. Start program 2. Prepare the blank: Fill 3. Prepare the sample: 4. Quickly invert the Ampul
85 Chlorine F&T AV. For the sample cell with 10 mL Collect at least 40 mL of several times to mix.
information about sample of sample. sample in a 50-mL beaker.
cells, adapters or light Fill the AccuVac Ampul with
shields, refer to Instrument- sample. Keep the tip
specific information immersed while the Ampul
on page 1. fills completely.
Note: Although the program
name may vary between
instruments, the program
number does not change.

Zero

5. Start the instrument 6. Clean the blank. 7. Insert the blank into the 8. Push ZERO. The display
timer. A 3-minute reaction cell holder. shows 0.00 mg/L Cl2.
time starts.
Prepare the sample blank
and set the instrument to
zero during the reaction
time.

4 Chlorine, Total, DPD Method (2.00 mg/L)

 Read

9. Clean the AccuVac 10. Within 3 minutes after 11. Push READ. Results
Ampul. the timer expires, insert the show in mg/L Cl2.
prepared sample AccuVac
Ampul into the cell holder.

Interferences
Interfering substance Interference level
Acidity More than 150 mg/L CaCO3. The full color may not develop or the color may fade instantly. Adjust
to pH 6–7 with 1 N Sodium Hydroxide. Measure the amount to be added on a separate sample
aliquot, then add the same amount to the sample that is tested. Correct the test result for the
dilution from the volume addition.
Alkalinity More than 250 mg/L CaCO3. The full color may not develop or the color may fade instantly. Adjust
to pH 6–7 with 1 N Sulfuric Acid. Measure the amount to add on a separate sample aliquot, then
add the same amount to the sample that is tested. Correct the test result for the dilution from the
volume addition.
Bromine, Br2 Interferes at all levels
Chlorine Dioxide, ClO2 Interferes at all levels
Chloramines, organic May interfere
Hardness No effect at less than 1000 mg/L as CaCO3
Iodine, I2 Interferes at all levels
Manganese, Oxidized Pre-treat the sample as follows:
(Mn4+, Mn7+) or
Chromium, Oxidized 1. Adjust the sample pH to 6–7.
(Cr6+) 2. Add 3 drops of Potassium Iodide (30-g/L) to 10 mL of sample.
3. Mix and wait 1 minute.
4. Add 3 drops of Sodium Arsenite (5-g/L) and mix.
5. Use the test procedure to measure the concentration of the treated sample.
6. Subtract this result from the result without the treatment to obtain the correct chlorine
concentration.

Ozone Interferes at all levels

Peroxides May interfere
Highly buffered samples Can prevent the correct pH adjustment of the sample by the reagents. Sample pretreatment may
or extreme sample pH be necessary. Adjust to pH 6–7 with acid (Sulfuric Acid, 1.000 N) or base (Sodium Hydroxide,
1.00 N).

Pollution prevention and waste management

If sodium arsenite was added to the sample for manganese or chromium interferences,
the reacted samples will contain arsenic and must be disposed of as a hazardous waste.
Dispose of reacted solutions according to local, state and federal regulations.

Chlorine, Total, DPD Method (2.00 mg/L) 5

Accuracy check
Standard additions method (sample spike)
Use the standard additions method (for applicable instruments) to validate the test
procedure, reagents and instrument and to find if there is an interference in the sample.
Items to collect:
• Chlorine Standard Solution, 2-mL PourRite® Ampule, 25–30 mg/L (use mg/L on label)
• Breaker, PourRite Ampules
• Pipet, TenSette®, 0.1–1.0 mL and tips

1. Use the test procedure to measure the concentration of the sample, then keep the
(unspiked) sample in the instrument.
2. Go to the Standard Additions option in the instrument menu.
3. Select the values for standard concentration, sample volume and spike volumes.
4. Open the standard solution.
5. Prepare three spiked samples: use the TenSette pipet to add 0.1 mL, 0.2 mL and
0.3 mL of the standard solution, respectively, to three 10-mL portions of fresh sample.
Mix well.
®
Note: For AccuVac Ampuls, add 0.4 mL, 0.8 mL and 1.2 mL of the standard solution to three
50-mL portions of fresh sample.
6. Use the test procedure to measure the concentration of each of the spiked samples.
Start with the smallest sample spike. Measure each of the spiked samples in the
instrument.
7. Select Graph to compare the expected results to the actual results.
Note: If the actual results are significantly different from the expected results, make sure that
the sample volumes and sample spikes are measured accurately. The sample volumes and
sample spikes that are used should agree with the selections in the standard additions menu. If
the results are not within acceptable limits, the sample may contain an interference.

Method performance
The method performance data that follows was derived from laboratory tests that were
measured on a spectrophotometer during ideal test conditions. Users may get different
results under different test conditions.
Program Standard Precision (95% Confidence Interval) Sensitivity
Concentration change per 0.010 Abs change
80 1.25 mg/L Cl2 1.23–1.27 mg/L Cl2 0.02 mg/L Cl2
85 1.25 mg/L Cl2 1.21–1.29 mg/L Cl2 0.02 mg/L Cl2

Summary of method
Chlorine can be present in water as free chlorine and as combined chlorine. Both forms
can exist in the same water and be determined together as total chlorine. Free chlorine is
present as hypochlorous acid and/or hypochlorite ion. Combined chlorine exists as
monochloramine, dichloramine, nitrogen trichloride and other chloro derivatives. The
combined chlorine oxidizes iodide in the reagent to iodine. The iodine and free chlorine
react with DPD (N,N-diethyl-p-phenylenediamine) to form a pink color which is
proportional to the total chlorine concentration.
To find the concentration of combined chlorine, run a free chlorine test and a total
chlorine test. Subtract the results of the free chlorine test from the total chlorine test to
obtain the combined chlorine concentration. The measurement wavelength is 530 nm for
spectrophotometers or 520 nm for colorimeters.

6 Chlorine, Total, DPD Method (2.00 mg/L)

Consumables and replacement items
Required reagents

Description Quantity/Test Unit Item no.

DPD Total Chlorine Reagent Powder Pillow, 10-mL 1 100/pkg 2105669

OR
®
DPD Total Chlorine Reagent AccuVac Ampul 1 25/pkg 2503025

Required apparatus

Description Quantity/Test Unit Item no.

AccuVac Snapper 1 each 2405200

Beaker, 50-mL 1 each 50041H
Stoppers for 18-mm tubes and AccuVac Ampuls 2 6/pkg 173106

Recommended standards

Description Unit Item no.

®
Chlorine Standard Solution, 10-mL Voluette Ampule, 50–75 mg/L 16/pkg 1426810
®
Chlorine Standard Solution, 2-mL PourRite Ampules, 50–75 mg/L 20/pkg 1426820
®
Chlorine Standard Solution, 2-mL PourRite Ampules, 25–30 mg/L 20/pkg 2630020

Optional reagents and apparatus

Description Unit Item no.

®
AccuVac vials for sample blanks 25/pkg 2677925
®
Ampule Breaker, PourRite ampules each 2484600
®
Ampule Breaker, Voluette ampules each 2196800
Water, Chlorine-demand Free 500 mL 2641549
Cylinder, mixing, 25-mL each 2088640
Cylinder, mixing, 50-mL each 189641
®
Chlorine Standard Solution, 2-mL PourRite Ampules, 50–75 mg/L 20/pkg 1426820
®
Chlorine Standard Solution, 10-mL Voluette Ampule, 50–75 mg/L 16/pkg 1426810
DPD Total Chlorine Reagent Powder Pillows, 10-mL 1000/pkg 2105628
DPD Total Chlorine Reagent Powder Pillows, 10-mL 300/pkg 2105603
DPD Total Chlorine Reagent, 10 mL, SwifTest™ Dispenser refill vial 250 tests 2105660
Paper, pH, 0–14 pH range 100/pkg 2601300
®
Pipet, TenSette , 0.1–1.0 mL each 1970001
Pipet tips for TenSette Pipet 1970001 50/pkg 2185696
Pipet tips for TenSette Pipet 1970001 1000/pkg 2185628
Potassium Iodide, 30 g/L 100 mL 34332
Sodium Arsenite, 5 g/L 100 mL 104732
100 mL
Sodium Hydroxide Standard Solution, 1.0 N 104532
MDB
SpecCheck™ Secondary Standard Kit, Chlorine DPD, 0-2.0 mg/L Set each 2635300

Chlorine, Total, DPD Method (2.00 mg/L) 7

Consumables and replacement items (continued)
Description Unit Item no.

100 mL
Sulfuric Acid Standard Solution, 1 N 127032
MDB
Water, deionized 4L 27256

FOR TECHNICAL ASSISTANCE, PRICE INFORMATION AND ORDERING: HACH COMPANY

In the U.S.A. – Call toll-free 800-227-4224 WORLD HEADQUARTERS
Outside the U.S.A. – Contact the HACH office or distributor serving you. Telephone: (970) 669-3050
On the Worldwide Web – www.hach.com; E-mail – [email protected] FAX: (970) 669-2932

© Hach Company/Hach Lange GmbH, 1989–2013. All rights reserved. 04/2013, Edition 8
Chlorine, Total DOC316.53.01029

USEPA DPD Method1 Method 10070

0.1 to 10.0 mg/L Cl2 (HR) Powder Pillows
Scope and application: For testing higher levels of total chlorine (free and combined) in drinking water, cooling
water and industrial process waters.2 This product has not been evaluated to test for chlorine and chloramines in
medical applications in the United States.
1 USEPA accepted for reporting drinking water analyses.
2 Procedure is equivalent to USEPA and Standard Method 4500-Cl G for drinking water and wastewater.

Test preparation

Instrument specific information

Table 1 shows all of the instruments that have the program for this test. The table also
shows requirements that may vary between instruments, such as adapter and sample cell
requirements. To use this table, select an instrument, then read across to find the
corresponding information for this test.
Table 1 Instrument-specific information
Instrument Adapter Sample cell orientation Sample cell
DR 6000 A23618 The orientation key is toward the user. 4864302
DR 5000 The orientation key is toward the user.
DR 3900 LZV846 (A) The orientation key is away from the user.
DR 900 — The orientation key is toward the user.
DR 3800 LZV585 (B) The 1-cm path is aligned with the arrow on the adapter. 5940506
DR 2800
DR 2700

Before starting
Samples must be analyzed immediately after collection and cannot be preserved for later analysis.
Install the instrument cap on the DR 900 cell holder before ZERO or READ is pushed.
In bright light conditions (e.g., direct sunlight), close the cell compartment on spectrophotometers, if applicable, with the
protective cover during measurements.
If the chlorine concentration is less than 2 mg/L, use Method 8167, program number 80. If the chlorine concentration is less
that 500 µg/L, use Method 8370 (for applicable instruments).
Review the Safety Data Sheets (MSDS/SDS) for the chemicals that are used and use any recommended personal protective
equipment.
Dispose of reacted solutions according to local, state and federal regulations. Use the Safety Data Sheets for disposal
information for unused reagents. Consult the environmental, health and safety staff for your facility and/or local regulatory
agencies for further disposal information.

1
Items to collect
Description Quantity

DPD Total Chlorine Reagent Powder Pillows, 25-mL 1

Sample cells (For information about sample cells, adapters or light shields, refer to Instrument
1
specific information on page 1.)

Refer to Consumables and replacement items on page 5 for reorder information.

Sample collection
• Samples must be analyzed immediately and cannot be preserved for later analysis.
• Chlorine is a strong oxidizing agent and it is unstable in natural waters. It reacts
quickly with various inorganic compounds and more slowly with organic compounds.
Many factors, including reactant concentrations, sunlight, pH, temperature and
salinity influence the decomposition of chlorine in water.
• Collect samples in clean glass bottles. Avoid plastic containers since these may have
a large chlorine demand.
• Pre-treat glass sample containers to remove any chlorine demand. Soak the
containers in a dilute bleach solution (1 mL commercial bleach to 1 liter of deionized
water) for at least 1 hour. Rinse thoroughly with deionized or distilled water. If sample
containers are rinsed thoroughly with deionized or distilled water after use, only
occasional pre-treatment is necessary.
• Be sure to get a representative sample. If the sample is taken from a spigot or faucet,
let the water flow for at least 5 minutes. Then let the container overflow with the
sample several times and then put the cap on the sample container so that there is
no headspace (air) above the sample. If a sample cell is used, rinse the cell several
times with the sample, then carefully fill to the 10-mL mark.

Powder pillow procedure

Start

1. Start program 2. Prepare the blank: Fill a 3. Clean the blank. 4. Insert the blank into the
88 Chlorine F&T HR. For sample cell to the 5-mL cell holder.
information about sample mark with sample.
cells, adapters or light
shields, refer to Instrument
specific information
on page 1.
Note: Although the program
name may vary between
instruments, the program
number does not change.

2 Chlorine, Total, DPD Method (10.0 mg/L)

 Zero

5. Push ZERO. The display 6. Prepare the sample: Fill 7. Add the contents of one 8. Close the sample cell.
shows 0.0 mg/L Cl2. a second sample cell to the DPD Total Chlorine Powder Shake the sample cell about
5-mL mark with sample. Pillow for 25-mL samples to 20 seconds to dissolve the
the sample. reagent.

Read

9. Start the instrument 10. Clean the sample cell. 11. When the timer expires, 12. Push READ. Results
timer. A 3-minute reaction insert the prepared sample show in mg/L Cl2.
time starts. into the cell holder.

Interferences
Interfering substance Interference level
Acidity More than 150 mg/L CaCO3. The full color may not develop or the color may fade instantly.
Adjust to pH 6–7 with 1 N Sodium Hydroxide. Measure the amount to be added on a separate
sample aliquot, then add the same amount to the sample that is tested. Correct the test result for
the dilution from the volume addition.
Alkalinity More than 250 mg/L CaCO3. The full color may not develop or the color may fade instantly.
Adjust to pH 6–7 with 1 N Sulfuric Acid. Measure the amount to add on a separate sample
aliquot, then add the same amount to the sample that is tested. Correct the test result for the
dilution from the volume addition.
Bromine, Br2 Interferes at all levels
Chlorine Dioxide, ClO2 Interferes at all levels
Chloramines, organic May interfere
Hardness No effect at less than 1000 mg/L as CaCO3
Iodine, I2 Interferes at all levels
Manganese, Oxidized Pre-treat the sample as follows:
(Mn4+, Mn7+) or
Chromium, Oxidized 1. Adjust the sample pH to 6–7.
(Cr6+) 2. Add 3 drops of Potassium Iodide (30-g/L) to 10 mL of sample.
3. Mix and wait 1 minute.
4. Add 3 drops of Sodium Arsenite (5-g/L) and mix.
5. Use the test procedure to measure the concentration of the treated sample.
6. Subtract this result from the result without the treatment to obtain the correct chlorine
concentration.

Chlorine, Total, DPD Method (10.0 mg/L) 3

Interfering substance Interference level
Ozone Interferes at all levels
Peroxides May interfere
Highly buffered samples Can prevent the correct pH adjustment of the sample by the reagents. Sample pretreatment may
or extreme sample pH be necessary.

Pollution prevention and waste management

If sodium arsenite was added to the sample for manganese or chromium interferences,
the reacted samples will contain arsenic and must be disposed of as a hazardous waste.
Dispose of reacted solutions according to local, state and federal regulations.
Accuracy check
Standard additions method (sample spike)
Use the standard additions method (for applicable instruments) to validate the test
procedure, reagents and instrument and to find if there is an interference in the sample.
Items to collect:
• Chlorine Standard Solution, 2-mL PourRite® Ampule, 50–75 mg/L (use mg/L on label)
• Ampule breaker
• Pipet, TenSette®, 0.1–1.0 mL and tips

1. Use the test procedure to measure the concentration of the sample, then keep the
(unspiked) sample in the instrument.
2. Go to the Standard Additions option in the instrument menu.
3. Select the values for standard concentration, sample volume and spike volumes.
4. Open the standard solution.
5. Prepare three spiked samples: use the TenSette pipet to add 0.1 mL, 0.2 mL and
0.3 mL of the standard solution, respectively, to three 5-mL portions of fresh sample.
Mix well.
6. Use the test procedure to measure the concentration of each of the spiked samples.
Start with the smallest sample spike. Measure each of the spiked samples in the
instrument.
7. Select Graph to compare the expected results to the actual results.
Note: If the actual results are significantly different from the expected results, make sure that
the sample volumes and sample spikes are measured accurately. The sample volumes and
sample spikes that are used should agree with the selections in the standard additions menu. If
the results are not within acceptable limits, the sample may contain an interference.

Method performance
The method performance data that follows was derived from laboratory tests that were
measured on a spectrophotometer during ideal test conditions. Users may get different
results under different test conditions.
Program Standard Precision (95% Confidence Interval) Sensitivity
Concentration change per 0.010 Abs change
88 5.4 mg/L Cl2 5.3–5.5 mg/L Cl2 0.1 mg/L Cl2

Summary of method
The range of analysis using the DPD method for total chlorine can be extended by adding
more indicator in proportion to sample volume. Thus, a larger fill powder pillow of DPD
Total Chlorine Reagent is added to a 5-mL sample portion. The combined chlorine
oxidizes iodide in the reagent to iodine. The iodine reacts with DPD (N, N-diethyl-p-
phenylenediamine) along with free chlorine present in the sample to form a pink color
which is proportional to the total chlorine concentration. The measurement wavelength is
530 nm for spectrophotometers or 520 nm for colorimeters.

4 Chlorine, Total, DPD Method (10.0 mg/L)

Consumables and replacement items
Required reagents

Description Quantity/test Unit Item no.

DPD Total Chlorine Reagent Powder Pillow, 25-mL 1 100/pkg 1406499

Recommended standards

Description Unit Item no.

®
Chlorine Standard Solution, 2-mL PourRite Ampule, 50-75 mg/L 20/pkg 1426820
®
Chlorine Standard Solution, 10-mL Voluette Ampule, 50–75 mg/L 16/pkg 1426810

Optional reagents and apparatus

Description Unit Item no.

®
Ampule Breaker, Voluette ampules each 2196800
®
PourRite Ampule Breaker, 2-mL each 2484600
Cylinder, mixing, 25-mL each 189640
®
Pipet, TenSette , 0.1–1.0 mL each 1970001
Pipet tips for TenSette Pipet 1970001 50/pkg 2185696
Pipet tips for TenSette Pipet 1970001 1000/pkg 2185628
Paper, pH, 0–14 pH range 100/pkg 2601300
Potassium Iodide, 30 g/L 100 mL 34332
Sodium Arsenite, 5 g/L 100 mL 104732
100 mL
Sodium Hydroxide Standard Solution, 1.0 N 104532
MDB
100 mL
Sulfuric Acid Standard Solution, 1 N 127032
MDB
®
Chlorine Standard Solution, 2-mL PourRite Ampules, 25–30 mg/L 20/pkg 2630020
DPD Total Chlorine Reagent Powder Pillows, 25-mL 1000/pkg 1406428

Chlorine, Total, DPD Method (10.0 mg/L) 5

 FOR TECHNICAL ASSISTANCE, PRICE INFORMATION AND ORDERING: HACH COMPANY
In the U.S.A. – Call toll-free 800-227-4224 WORLD HEADQUARTERS
Outside the U.S.A. – Contact the HACH office or distributor serving you. Telephone: (970) 669-3050
On the Worldwide Web – www.hach.com; E-mail – [email protected] FAX: (970) 669-2932

© Hach Company/Hach Lange GmbH, 1989–2013. All rights reserved. 04/2013, Edition 8
Chlorine, Total, MR DOC316.53.01304

USEPA1 DPD Method2 Method 10250

0.05 to 4.00 mg/L Cl2 (MR) Powder Pillows
Scope and application: For testing residual chlorine and chloramines in water, wastewater, estuary water and
seawater; USEPA-accepted for reporting drinking and wastewater analyses. This product has not been evaluated
to test for chlorine and chloramines in medical applications in the United States.
1 Procedure is equivalent to USEPA and Standard Method 4500-Cl G for drinking water and wastewater analyses.
2 Adapted from Standard Methods for the Examination of Water and Wastewater.

Test preparation

Instrument specific information

The table in this section shows all of the instruments that have the program for this test.
Table 1 shows sample cell and adapter requirements for this test.
To use the table, select an instrument, then read across to find the corresponding
information.
Table 1 Instrument-specific information
Instrument Adapter Sample cell
DR 6000 — 2427606
DR 5000 A23618
DR 3900 LZV846 (A)
DR 3800 LZV584 (C)
DR 2800
DR 2700
DR 900 — 2401906

Before starting
Samples must be analyzed immediately after collection and cannot be preserved for later analysis.
Install the instrument cap on the DR 900 cell holder before ZERO or READ is pushed.
If the test result is over-range, or if the sample temporarily turns yellow after the reagent addition, dilute the sample with a
known volume of high quality, chlorine demand-free water and repeat the test. Some loss of chlorine may occur due to the
dilution. Multiply the result by the dilution factor. Additional methods are available to measure chlorine without dilution.
For best results, measure the reagent blank value for each new lot of reagent. Replace the sample with deionized water in
the test procedure to get the reagent blank value. Subtract the reagent blank value from the sample results automatically
with the reagent blank adjust option.
A powder pillow for 25-mL samples is intentionally added to 10 mL of sample in this test to get the correct working range.
Review the Safety Data Sheets (MSDS/SDS) for the chemicals that are used and use any recommended personal protective
equipment.
Dispose of reacted solutions according to local, state and federal regulations. Use the Safety Data Sheets for disposal
information for unused reagents. Consult the environmental, health and safety staff for your facility and/or local regulatory
agencies for further disposal information.

1
For chloramination disinfection control, use one of the available Chloramine (Mono) methods.
Cold waters can cause condensation on the sample cells during color development. Examine the sample cells for
condensation before measurements.
Do not use the same sample cells for free and total chlorine. If trace iodide from the total chlorine reagent is carried over into
the free chlorine determination, monochloramine will interfere. It is best to use separate, dedicated sample cells for free and
total chlorine measurements.

Items to collect
Description Quantity

DPD Total Chlorine Reagent powder pillow, 25-mL 1

Sample cells (For information about sample cells, adapters or light shields, refer to Instrument
2
specific information on page 1.)

Refer to Consumables and replacement items on page 5 for reorder information.

Sample collection
• Samples must be analyzed immediately and cannot be preserved for later analysis.
• Chlorine is a strong oxidizing agent and it is unstable in natural waters. It reacts
quickly with various inorganic compounds and more slowly with organic compounds.
Many factors, including reactant concentrations, sunlight, pH, temperature and
salinity influence the decomposition of chlorine in water.
• Collect samples in clean glass bottles. Avoid plastic containers since these may have
a large chlorine demand.
• Pre-treat glass sample containers to remove any chlorine demand. Soak the
containers in a dilute bleach solution (1 mL commercial bleach to 1 liter of deionized
water) for at least 1 hour. Rinse thoroughly with deionized or distilled water. If sample
containers are rinsed thoroughly with deionized or distilled water after use, only
occasional pre-treatment is necessary.
• Be sure to get a representative sample. If the sample is taken from a spigot or faucet,
let the water flow for at least 5 minutes. Then let the container overflow with the
sample several times and then put the cap on the sample container so that there is
no headspace (air) above the sample. If a sample cell is used, rinse the cell several
times with the sample, then carefully fill to the 10-mL mark.

2 Chlorine, Total, DPD Method (4.00 mg/L)

Powder pillow procedure

Start

1. Start program 2. Prepare the sample: Fill 3. Add the contents of one 4. Close the sample cell.
87 Chlorine,F&T PP MR. a sample cell with 10 mL of DPD Total Chlorine Reagent Invert the sample cell for
For information about sample. Powder Pillow (for 25-mL 20 seconds. It is not
sample cells, adapters or samples). necessary for all of the
light shields, refer to reagent to dissolve.
Instrument specific
information on page 1.
Note: Although the program
name may vary between
instruments, the program
number does not change.

5. Start the instrument 6. Prepare the blank: Fill a 7. Clean the blank. 8. Insert the blank into the
timer. A 3-minute reaction second sample cell with cell holder.
time starts. 10 mL of sample.
Do the next four steps
during the timer period.

Zero Read

9. Push ZERO. The display 10. Clean the prepared 11. Within 3 minutes after 12. Push READ. Results
shows 0.00 mg/L Cl2. sample. the timer expires, insert the show in mg/L Cl2.
prepared sample into the
cell holder.

Chlorine, Total, DPD Method (4.00 mg/L) 3

Interferences
Interfering substance Interference level
Acidity More than 150 mg/L CaCO3. The full color may not develop or the color may fade instantly. Adjust
to pH 6–7 with 1 N Sodium Hydroxide. Measure the amount to be added on a separate sample
aliquot, then add the same amount to the sample that is tested. Correct the test result for the
dilution from the volume addition.
Alkalinity More than 250 mg/L CaCO3. The full color may not develop or the color may fade instantly. Adjust
to pH 6–7 with 1 N Sulfuric Acid. Measure the amount to add on a separate sample aliquot, then
add the same amount to the sample that is tested. Correct the test result for the dilution from the
volume addition.
Bromine, Br2 Interferes at all levels
Chlorine Dioxide, ClO2 Interferes at all levels
Chloramines, organic May interfere
Hardness No effect at less than 1000 mg/L as CaCO3
Iodine, I2 Interferes at all levels
Manganese, Oxidized Pre-treat the sample as follows:
(Mn4+, Mn7+) or
Chromium, Oxidized 1. Adjust the sample pH to 6–7.
(Cr6+) 2. Add 3 drops of Potassium Iodide (30-g/L) to 10 mL of sample.
3. Mix and wait 1 minute.
4. Add 3 drops of Sodium Arsenite (5-g/L) and mix.
5. Use the test procedure to measure the concentration of the treated sample.
6. Subtract this result from the result without the treatment to obtain the correct chlorine
concentration.

Ozone Interferes at all levels

Peroxides May interfere
Highly buffered samples Can prevent the correct pH adjustment of the sample by the reagents. Sample pretreatment may
or extreme sample pH be necessary. Adjust to pH 6–7 with acid (Sulfuric Acid, 1.000 N) or base (Sodium Hydroxide,
1.00 N).

Pollution prevention and waste management

If sodium arsenite was added to the sample for manganese or chromium interferences,
the reacted samples will contain arsenic and must be disposed of as a hazardous waste.
Dispose of reacted solutions according to local, state and federal regulations.
Accuracy check
Standard additions method (sample spike)
Use the standard additions method (for applicable instruments) to validate the test
procedure, reagents and instrument and to find if there is an interference in the sample.
Items to collect:
• Chlorine Standard Solution, 2-mL PourRite® Ampule, 25–30 mg/L or 50–75 mg/L
(use mg/L on label)
• Ampule breaker
• Pipet, TenSette®, 0.1–1.0 mL and tips

1. Use the test procedure to measure the concentration of the sample, then keep the
(unspiked) sample in the instrument.
2. Go to the Standard Additions option in the instrument menu.
3. Select the values for standard concentration, sample volume and spike volumes.
4. Open the standard solution.

4 Chlorine, Total, DPD Method (4.00 mg/L)

 5. Prepare three spiked samples: use the TenSette pipet to add 0.1 mL, 0.2 mL and
0.3 mL of the standard solution, respectively, to three 10-mL portions of fresh sample.
Mix well.
6. Use the test procedure to measure the concentration of each of the spiked samples.
Start with the smallest sample spike. Measure each of the spiked samples in the
instrument.
7. Select Graph to compare the expected results to the actual results.
Note: If the actual results are significantly different from the expected results, make sure that
the sample volumes and sample spikes are measured accurately. The sample volumes and
sample spikes that are used should agree with the selections in the standard additions menu. If
the results are not within acceptable limits, the sample may contain an interference.

Verification of on-line analyzers

This procedure can be used to meet the requirements of USEPA Method 334.0 -
Determination of Residual Chlorine in Drinking Water Using an On-line Chlorine Analyzer.
The procedure and requirements for compliance with EPA Method 334.0 can be
downloaded directly from http://www.hach.com/method334.
Method performance
The method performance data that follows was derived from laboratory tests that were
measured on a spectrophotometer during ideal test conditions. Users may get different
results under different test conditions.
Program Standard Precision (95% Confidence Interval) Sensitivity
Concentration change per 0.010 Abs change
87 2.10 mg/L Cl2 2.07–2.13 mg/L Cl2 0.02 mg/L Cl2

Summary of method
Chlorine can be present in water as free chlorine and as combined chlorine. Both forms
can exist in the same water and be determined together as total chlorine. Free chlorine is
present as hypochlorous acid and/or hypochlorite ion. Combined chlorine exists as
monochloramine, dichloramine, nitrogen trichloride and other chloro derivatives. The
combined chlorine oxidizes iodide in the reagent to iodine. The iodine and free chlorine
react with DPD (N,N-diethyl-p-phenylenediamine) to form a pink color which is
proportional to the total chlorine concentration.
To find the concentration of combined chlorine, run a free chlorine test and a total
chlorine test. Subtract the results of the free chlorine test from the total chlorine test to
obtain the combined chlorine concentration. The measurement wavelength is 530 nm for
spectrophotometers or 520 nm for colorimeters.
Consumables and replacement items
Required reagents

Description Quantity/test Unit Item no.

DPD Total Chlorine Reagent Powder Pillow, 25-mL 1 100/pkg 1406499

Recommended standards and apparatus

Description Unit Item no.

®
Chlorine Standard Solution, 2-mL PourRite Ampules, 25–30 mg/L 20/pkg 2630020
®
Chlorine Standard Solution, 2-mL PourRite Ampules, 50–75 mg/L 20/pkg 1426820
®
Chlorine Standard Solution, 10-mL Voluette Ampule, 50–75 mg/L 16/pkg 1426810
®
Ampule Breaker, Voluette ampules each 2196800
®
PourRite Ampule Breaker, 2-mL each 2484600

Chlorine, Total, DPD Method (4.00 mg/L) 5

Optional reagents and apparatus

Description Unit Item no.

Beaker, 50-mL each 50041H

Cylinder, mixing, 25-mL each 2088640
Water, deionized 4L 27256
Potassium Iodide, 30 g/L 100 mL 34332
Sodium Arsenite, 5 g/L 100 mL 104732
100 mL
Sodium Hydroxide Standard Solution, 1.0 N 104532
MDB
100 mL
Sulfuric Acid Standard Solution, 1 N 127032
MDB
®
Pipet, TenSette , 0.1–1.0 mL each 1970001
Pipet tips for TenSette Pipet 1970001 50/pkg 2185696
Pipet tips for TenSette Pipet 1970001 1000/pkg 2185628
Paper, pH, 0–14 pH range 100/pkg 2601300
DPD Total Chlorine Reagent Powder Pillows, 25-mL 1000/pkg 1406428
SpecCheck™ Secondary Standard Kit, Chlorine DPD, MR each 2980500
Organic-free water 500 mL 2641549

FOR TECHNICAL ASSISTANCE, PRICE INFORMATION AND ORDERING: HACH COMPANY

In the U.S.A. – Call toll-free 800-227-4224 WORLD HEADQUARTERS
Outside the U.S.A. – Contact the HACH office or distributor serving you. Telephone: (970) 669-3050
On the Worldwide Web – www.hach.com; E-mail – [email protected] FAX: (970) 669-2932

© Hach Company/Hach Lange GmbH, 1989–2013. All rights reserved. 04/2013, Edition 3
Chlorine, Total DOC316.53.01028

DPD Method1 Method 10101

0.09 to 5.00 mg/L Cl2 Test 'N Tube™ Vials
Scope and application: For testing higher levels of total (free plus combined) chlorine in drinking water, treated
wastewater, cooling water or industrial process water. This product has not been evaluated to test for chlorine and
chloramines in medical applications in the United States.
1 Adapted from Standard Methods for the Examination of Water and Wastewater.

Test preparation

Instrument specific information

The table in this section shows all of the instruments that have the program for this test.
Table 1 shows adapter and light shield requirements for the instruments that use them.
To use the table, select an instrument, then read across to find the corresponding
information for this test.
Table 1 Instrument-specific information for test tubes
Instrument Adapters Light shield
DR 6000 — —
DR 5000 — —
DR 900 4846400 Cover supplied with the instrument
DR 3900 — LZV849
DR 3800 — LZV646
DR 2800 —

Before starting
Samples must be analyzed immediately after collection and cannot be preserved for later analysis.
Install the instrument cap on the DR 900 cell holder before ZERO or READ is pushed.
DR 3900, DR 3800, DR 2800 and DR 2700: Install the light shield in Cell Compartment #2 before this test is started.
For best results, measure the reagent blank value for each new lot of reagent. Replace the sample with deionized water in
the test procedure to get the reagent blank value. Subtract the reagent blank value from the sample results automatically
with the reagent blank adjust option.
For chloramination disinfection control, use one of the available Chloramine (Mono) methods.
Review the Safety Data Sheets (MSDS/SDS) for the chemicals that are used and use any recommended personal protective
equipment.
Dispose of reacted solutions according to local, state and federal regulations. Use the Safety Data Sheets for disposal
information for unused reagents. Consult the environmental, health and safety staff for your facility and/or local regulatory
agencies for further disposal information.

Items to collect
Description Quantity

Light shield (For information about sample cells, adapters or light shields, refer to Instrument
1
specific information on page 1.)
Test 'N Tube™ DPD Total Chlorine Reagent 1
Wipes, disposable varies

1
Refer to Consumables and replacement items on page 5 for reorder information.

Sample collection
• Samples must be analyzed immediately and cannot be preserved for later analysis.
• Chlorine is a strong oxidizing agent and it is unstable in natural waters. It reacts
quickly with various inorganic compounds and more slowly with organic compounds.
Many factors, including reactant concentrations, sunlight, pH, temperature and
salinity influence the decomposition of chlorine in water.
• Collect samples in clean glass bottles. Avoid plastic containers since these may have
a large chlorine demand.
• Pre-treat glass sample containers to remove any chlorine demand. Soak the
containers in a dilute bleach solution (1 mL commercial bleach to 1 liter of deionized
water) for at least 1 hour. Rinse thoroughly with deionized or distilled water. If sample
containers are rinsed thoroughly with deionized or distilled water after use, only
occasional pre-treatment is necessary.
• Be sure to get a representative sample. If the sample is taken from a spigot or faucet,
let the water flow for at least 5 minutes. Then let the container overflow with the
sample several times and then put the cap on the sample container so that there is
no headspace (air) above the sample. If a sample cell is used, rinse the cell several
times with the sample, then carefully fill to the 10-mL mark.

DPD method for Test N' Tubes

Start

1. Start program 2. Prepare the blank: Fill 3. Clean the blank vial. 4. Insert the blank vial into
89 Chlorine F&T TNT. For an empty Test 'N Tube vial the 16-mm cell holder.
information about sample to the top of the label with
cells, adapters or light sample.
shields, refer to Instrument
specific information
on page 1.
Note: Although the program
name may vary between
instruments, the program
number does not change.

2 Chlorine, Total, DPD TNT Method (5.00 mg/L)

 Zero

5. Push ZERO. The display 6. Prepare the sample: 7. Put the cap on the vial. 8. Start the instrument
shows 0.00 mg/L Cl2. Remove the cap from a Invert to mix. timer. A 3-minute reaction
Total Chlorine DPD Test N' time starts.
Tube. Fill the vial to the top
of the label with sample.

Read

9. Clean the sample vial. 10. When the timer expires, 11. Push READ. Results
insert the prepared sample show in mg/L Cl2.
into the cell holder.

Interferences
Interfering substance Interference level
Acidity More than 150 mg/L CaCO3. The full color may not develop or the color may fade instantly.
Adjust to pH 6–7 with 1 N Sodium Hydroxide. Measure the amount to be added on a separate
sample aliquot, then add the same amount to the sample that is tested. Correct the test result for
the dilution from the volume addition.
Alkalinity More than 250 mg/L CaCO3. The full color may not develop or the color may fade instantly.
Adjust to pH 6–7 with 1 N Sulfuric Acid. Measure the amount to add on a separate sample
aliquot, then add the same amount to the sample that is tested. Correct the test result for the
dilution from the volume addition.
Bromine, Br2 Interferes at all levels
Chlorine Dioxide, ClO2 Interferes at all levels
Chloramines, organic May interfere
Hardness No effect at less than 1000 mg/L as CaCO3
Iodine, I2 Interferes at all levels
Manganese, Oxidized Pre-treat the sample as follows:
(Mn4+, Mn7+) or
Chromium, Oxidized 1. Adjust the sample pH to 6–7.
(Cr6+) 2. Add 3 drops of Potassium Iodide (30-g/L) to 10 mL of sample.
3. Mix and wait 1 minute.
4. Add 3 drops of Sodium Arsenite (5-g/L) and mix.
5. Use the test procedure to measure the concentration of the treated sample.
6. Subtract this result from the result without the treatment to obtain the correct chlorine
concentration.

Chlorine, Total, DPD TNT Method (5.00 mg/L) 3

Interfering substance Interference level
Ozone Interferes at all levels
Peroxides May interfere
Highly buffered samples Can prevent the correct pH adjustment of the sample by the reagents. Sample pretreatment may
or extreme sample pH be necessary.

Pollution prevention and waste management

If sodium arsenite was added to the sample for manganese or chromium interferences,
the reacted samples will contain arsenic and must be disposed of as a hazardous waste.
Dispose of reacted solutions according to local, state and federal regulations.
Accuracy check
Standard additions method (sample spike)
Use the standard additions method (for applicable instruments) to validate the test
procedure, reagents and instrument and to find if there is an interference in the sample.
Items to collect:
• Chlorine Standard Solution, 2-mL PourRite® Ampule, 50–75 mg/L (use mg/L on label)
• Ampule breaker
• Pipet, TenSette®, 0.1–1.0 mL and tips

1. Use the test procedure to measure the concentration of the sample, then keep the
(unspiked) sample in the instrument.
2. Go to the Standard Additions option in the instrument menu.
3. Select the values for standard concentration, sample volume and spike volumes.
4. Open the standard solution.
5. Prepare three spiked samples: use the TenSette pipet to add 0.1 mL, 0.2 mL and
0.3 mL of the standard solution, respectively, to three 10-mL portions of fresh sample.
Mix well.
6. Use the test procedure to measure the concentration of each of the spiked samples.
Start with the smallest sample spike. Measure each of the spiked samples in the
instrument.
7. Select Graph to compare the expected results to the actual results.
Note: If the actual results are significantly different from the expected results, make sure that
the sample volumes and sample spikes are measured accurately. The sample volumes and
sample spikes that are used should agree with the selections in the standard additions menu. If
the results are not within acceptable limits, the sample may contain an interference.

Method performance
The method performance data that follows was derived from laboratory tests that were
measured on a spectrophotometer during ideal test conditions. Users may get different
results under different test conditions.
Program Standard Precision (95% Confidence Interval) Sensitivity
Concentration change per 0.010 Abs change
89 2.68 mg/L Cl2 2.63–2.73 mg/L Cl2 0.03 mg/L Cl2

Summary of method
Chlorine can be present in water as free chlorine and as combined chlorine. Both forms
can exist in the same water and be determined together as total chlorine. Free chlorine is
present as hypochlorous acid and/or hypochlorite ion. Combined chlorine exists as
monochloramine, dichloramine, nitrogen trichloride and other chloro derivatives. The
combined chlorine oxidizes iodide in the reagent to iodine. The iodine and free chlorine
react with DPD (N,N-diethyl-p-phenylenediamine) to form a pink color which is
proportional to the total chlorine concentration.

4 Chlorine, Total, DPD TNT Method (5.00 mg/L)

 To find the concentration of combined chlorine, run a free chlorine test and a total
chlorine test. Subtract the results of the free chlorine test from the total chlorine test to
obtain the combined chlorine concentration. The measurement wavelength is 530 nm for
spectrophotometers or 520 nm for colorimeters.
Consumables and replacement items
Required reagents

Description Quantity/test Unit Item no.

Test 'N Tube™ DPD Total Chlorine Reagent 1 50/pkg 2105645

Recommended standards and apparatus

Description Unit Item no.

®
Chlorine Standard Solution, 2-mL PourRite Ampules, 50–75 mg/L 20/pkg 1426820
®
PourRite Ampule Breaker, 2-mL each 2484600

Optional reagents and apparatus

Description Unit Item no.

®
Pipet, TenSette , 0.1–1.0 mL each 1970001
Pipet tips for TenSette Pipet 1970001 50/pkg 2185696
Pipet tips for TenSette Pipet 1970001 1000/pkg 2185628
Paper, pH, 0–14 pH range 100/pkg 2601300
®
Ampule Breaker, Voluette ampules each 2196800
Test tube rack each 1864100
Potassium Iodide, 30 g/L 100 mL 34332
Sodium Arsenite, 5 g/L 100 mL 104732
100 mL
Sodium Hydroxide Standard Solution, 1.0 N 104532
MDB
100 mL
Sulfuric Acid Standard Solution, 1 N 127032
MDB
®
Chlorine Standard Solution, 10-mL Voluette Ampule, 50–75 mg/L 16/pkg 1426810
®
Chlorine Standard Solution, 2-mL PourRite Ampule, 25-30 mg/L 20/pkg 2630020

Chlorine, Total, DPD TNT Method (5.00 mg/L) 5

 FOR TECHNICAL ASSISTANCE, PRICE INFORMATION AND ORDERING: HACH COMPANY
In the U.S.A. – Call toll-free 800-227-4224 WORLD HEADQUARTERS
Outside the U.S.A. – Contact the HACH office or distributor serving you. Telephone: (970) 669-3050
On the Worldwide Web – www.hach.com; E-mail – [email protected] FAX: (970) 669-2932

© Hach Company/Hach Lange GmbH, 1989–2013. All rights reserved. 04/2013, Edition 8
Chromium, Hexavalent DOC316.53.01033

USEPA1 1,5-Diphenylcarbohydrazide Method2 Method 8023

®
0.010 to 0.700 mg/L Cr6+ (spectrophotometers) Powder Pillows or AccuVac Ampuls
0.01 to 0.60 mg/L Cr6+ (colorimeters)
Scope and application: For water and wastewater; USEPA accepted for reporting for wastewater analysis.3
1 Accepted USEPA and Standard Method 3500 Cr B.
2 Adapted from Standard Methods for the Examination of Water and Wastewater.
3 Procedure is equivalent to USGS method 1-1230-85 for wastewater.

Test preparation

Instrument-specific information
The tables in this section show all of the instruments that have the program for this test.
Table 1 shows sample cell and orientation requirements for reagent addition tests, such
as powder pillow or bulk reagent tests. Table 2 shows sample cell and adapter
requirements for AccuVac Ampul tests.
To use either table, select an instrument, then read across to find the corresponding
information for this test.
Table 1 Instrument-specific information for powder pillows
Instrument Sample cell orientation Sample cell
DR 6000 The fill line is to the right. 2495402
DR 3800
DR 2800
DR 2700
DR 5000 The fill line is toward the user.
DR 3900
DR 900 The orientation mark is toward the user. 2401906

Table 2 Instrument-specific information for AccuVac Ampuls

Instrument Adapter Sample cell
DR 6000 — 2427606
DR 5000
DR 900
DR 3900 LZV846 (A)
DR 3800 LZV584 (C) 2122800
DR 2800
DR 2700

1
Before starting
Install the instrument cap on the DR 900 cell holder before ZERO or READ is pushed.
For best results, measure the reagent blank value for each new lot of reagent. Replace the sample with deionized water in
the test procedure to get the reagent blank value. Subtract the reagent blank value from the sample results automatically
with the reagent blank adjust option.
At high chromium levels, a precipitate forms. Sample dilution may be necessary.
Review the Safety Data Sheets (MSDS/SDS) for the chemicals that are used and use any recommended personal protective
equipment.
The final samples are highly acidic.
Dispose of reacted solutions according to local, state and federal regulations. Use the Safety Data Sheets for disposal
information for unused reagents. Consult the environmental, health and safety staff for your facility and/or local regulatory
agencies for further disposal information.

Items to collect
Powder pillows

Description Quantity
®
ChromaVer 3 Chromium Reagent Powder Pillows, 5– or 10–mL 1
Sample cells. (For information about sample cells, adapters or light shields, refer to Instrument-
2
specific information on page 1.)

Refer to Consumables and replacement items on page 6 for reorder information.

AccuVac Ampuls

Description Quantity
® ®
ChromaVer 3 AccuVac Ampuls 1
Beaker, 50-mL 1
Sample cells (For information about sample cells, adapters or light shields, refer to Instrument-
1
specific information on page 1.)
Stopper for 18-mm tubes and AccuVac Ampuls 1

Refer to Consumables and replacement items on page 6 for reorder information.

Sample collection and storage

• Collect samples in clean glass or plastic bottles.
• To preserve samples for later analysis, keep the samples at or below 6 °C (43 °F) for
up to 24 hours.
• Let the sample temperature increase to room temperature before analysis.

2 Chromium, Hexavalent, 1,5-Diphenylcarbohydrazide Method (0.700 mg/L)

Powder pillow procedure

Start

1. Start program 2. Prepare the sample: Fill 3. Add the contents of one 4. Swirl to mix.
®
90 Chromium, Hex. For a sample cell with 10 mL of ChromaVer 3 Reagent A purple color will show if
information about sample sample. Powder Pillow to the sample hexavalent chromium is
cells, adapters or light cell. present.
shields, refer to Instrument-
specific information
on page 1.
Note: Although the program
name may vary between
instruments, the program
number does not change.

5. Start the instrument 6. Prepare the blank: Fill a 7. When the timer expires, 8. Insert the blank into the
timer. A 5-minute reaction second sample cell with clean the blank. cell holder.
time starts. 10 mL of sample.

Zero Read

9. Push ZERO. The display 10. Clean the prepared 11. Insert the prepared 12. Push READ. Results
shows 0.00 or 0.000 mg/L sample. sample into the cell holder. show in mg/L Cr6+.
Cr6+*.

* Colorimeter display shows 0.00 mg/L.

Chromium, Hexavalent, 1,5-Diphenylcarbohydrazide Method (0.700 mg/L) 3

AccuVac Ampul procedure

Start

1. Start program 2. Prepare the blank: Fill 3. Prepare the sample: 4. Quickly invert the Ampul
95 Chromium, Hex. AV. the sample cell with 10 mL Collect at least 40 mL of several times to mix.
For information about of sample. sample in a 50-mL beaker.
sample cells, adapters or Fill the AccuVac Ampul with
light shields, refer to sample. Keep the tip
Instrument-specific immersed while the Ampul
information on page 1. fills completely.
Note: Although the program
name may vary between
instruments, the program
number does not change.

Zero

5. Start the instrument 6. When the timer expires, 7. Insert the blank into the 8. Push ZERO. The display
timer. A 5-minute reaction clean the blank. cell holder. shows 0.00 or 0.000 mg/L
time starts. Cr6+**.

Read

9. Clean the AccuVac 10. Insert the prepared 11. Push READ. Results
Ampul. sample AccuVac Ampul into show in mg/L Cr6+.
the cell holder.

** Colorimeter display shows 0.00 mg/L.

4 Chromium, Hexavalent, 1,5-Diphenylcarbohydrazide Method (0.700 mg/L)

Interferences
Interfering substance Interference level
Iron May interfere above 1 mg/L
Mercurous and Mercuric Ions Interfere slightly
Highly buffered samples or Can prevent the correct pH adjustment of the sample by the reagents. Sample pretreatment
extreme sample pH may be necessary.
Vanadium May interfere above 1 mg/L. Allow 10 minutes for the reaction period before reading.
Turbidity For turbid samples, treat the blank with the contents of one Acid Reagent Powder Pillow.
This will make sure that any turbidity dissolved by the acid in the ChromaVer 3 Chromium
Reagent will also be dissolved in the blank.

Accuracy check
Standard additions method (sample spike)
Use the standard additions method (for applicable instruments) to validate the test
procedure, reagents and instrument and to find if there is an interference in the sample.
Items to collect:
• 12.5-mg/L Hexavalent Chromium Standard Solution, 10-mL Voluette® Ampules
• Ampule breaker
• Pipet, TenSette®, 0.1–1.0 mL and tips
• Mixing cylinders, 25-mL (3)

1. Use the test procedure to measure the concentration of the sample, then keep the
(unspiked) sample in the instrument.
2. Go to the Standard Additions option in the instrument menu.
3. Select the values for standard concentration, sample volume and spike volumes.
4. Open the standard solution.
5. Prepare three spiked samples: use the TenSette pipet to add 0.1 mL, 0.2 mL and
0.3 mL of the standard solution, respectively, to three 25-mL portions of fresh sample.
Mix well.
®
Note: For AccuVac Ampuls, add 0.2 mL, 0.4 mL and 0.6 mL of the standard solution to three
50-mL portions of fresh sample.
6. Use the test procedure to measure the concentration of each of the spiked samples.
Start with the smallest sample spike. Measure each of the spiked samples in the
instrument.
7. Select Graph to compare the expected results to the actual results.
Note: If the actual results are significantly different from the expected results, make sure that
the sample volumes and sample spikes are measured accurately. The sample volumes and
sample spikes that are used should agree with the selections in the standard additions menu. If
the results are not within acceptable limits, the sample may contain an interference.

Standard solution method

Use the standard solution method to validate the test procedure, reagents and
instrument.
Items to collect:
• 50.0-mg/L Hexavalent Chromium Standard Solution
• 500-mL volumetric flask, Class A
• 5-mL volumetric pipet, Class A and pipet filler
• Deionized water

1. Prepare a 0.50 mg/L hexavalent chromium standard solution as follows:

Chromium, Hexavalent, 1,5-Diphenylcarbohydrazide Method (0.700 mg/L) 5

 a. Use a pipet to add 5.00 mL of 50.0 mg/L hexavalent chromium standard solution
into the volumetric flask.
b. Dilute to the mark with deionized water. Mix well. Prepare this solution daily.
2. Use the test procedure to measure the concentration of the prepared standard
solution.
3. Compare the expected result to the actual result.
Note: The factory calibration can be adjusted slightly with the standard adjust option so that the
instrument shows the expected value of the standard solution. The adjusted calibration is then
used for all test results. This adjustment can increase the test accuracy when there are slight
variations in the reagents or instruments.

Method performance
The method performance data that follows was derived from laboratory tests that were
measured on a spectrophotometer during ideal test conditions. Users may get different
results under different test conditions.
Program Standard Precision (95% Confidence Interval) Sensitivity
Concentration change per 0.010 Abs change
90 0.500 mg/L Cr6+ 0.497–0.503 mg/L Cr6+ 0.005 mg/L Cr6+
95 0.500 mg/L Cr6+ 0.496–0.504 mg/L Cr6+ 0.006 mg/L Cr6+

Summary of method
Hexavalent chromium is determined by the 1,5-Diphenylcarbohydrazide method using a
single dry powder formulation called ChromaVer 3 Chromium Reagent. This reagent
contains an acidic buffer combined with 1,5-Diphenylcarbohydrazide, which reacts to give
a purple color when hexavalent chromium is present. The measurement wavelength is
540 nm for spectrophotometers or 560 nm for colorimeters.
Consumables and replacement items
Required reagents

Description Quantity/test Unit Item no.

®
ChromaVer 3 Chromium Reagent Powder Pillow, 5– or 10–mL 1 100/pkg 1271099
OR
® ®
ChromaVer 3 AccuVac Ampul 1 25/pkg 2505025
Water, deionized varies 4L 27256

Required apparatus

Description Quantity/test Unit Item no.

AccuVac Snapper 1 each 2405200

Beaker, 50-mL 1 each 50041H
Stoppers for 18-mm tubes and AccuVac Ampuls 2 6/pkg 173106

Recommended standards

Description Unit Item no.

®
Chromium, Hexavalent Standard Solution, 10-mL Voluette Ampules, 12.5-mg/L Cr6+ 16/pkg 1425610
Chromium Hexavalent Standard Solution, 50.0-mg/L Cr6+ 100 mL 81042H

6 Chromium, Hexavalent, 1,5-Diphenylcarbohydrazide Method (0.700 mg/L)

Optional reagents and apparatus

Description Unit Item no.

Acid Reagent Powder Pillow 100/pkg 212699

®
Ampule Breaker, Voluette ampules each 2196800
Flask, volumetric, Class A, 500-mL each 1457449
Pipet, volumetric 5.00-mL each 1451537
Pipet filler, safety bulb each 1465100
Sodium Hydroxide, 5 N 50 mL 245026

Chromium, Hexavalent, 1,5-Diphenylcarbohydrazide Method (0.700 mg/L) 7

 FOR TECHNICAL ASSISTANCE, PRICE INFORMATION AND ORDERING: HACH COMPANY
In the U.S.A. – Call toll-free 800-227-4224 WORLD HEADQUARTERS
Outside the U.S.A. – Contact the HACH office or distributor serving you. Telephone: (970) 669-3050
On the Worldwide Web – www.hach.com; E-mail – [email protected] FAX: (970) 669-2932

© Hach Company/Hach Lange GmbH, 1989–2013. All rights reserved. 04/2013, Edition 8
Chromium, Total DOC316.53.01034

Alkaline Hypobromite Oxidation Method1 Method 8024

0.01 to 0.70 mg/L Cr (spectrophotometers) Powder Pillows
0.01 to 0.60 mg/L Cr (colorimeters)
Scope and application: For water and wastewater.2
1 Adapted from Standard Methods for the Examination of Water and Wastewater.
2 Procedure is equivalent to USEPA and Standard Method 3500-Cr D for wastewater.

Test preparation

Instrument specific information

The table in this section shows all of the instruments that have the program for this test.
Table 1 shows sample cell and orientation requirements for reagent addition tests, such
as powder pillow or bulk reagent tests.
To use the table, select an instrument, then read across to find the corresponding
information for this test.
Table 1 Instrument-specific information for reagent addition
Instrument Measurement cell Mixing cell Measurement cell
orientation
DR 6000 The fill line is to the right. 2401906 2495402
DR 3800
DR 2800
DR 2700
DR 5000 The fill line is toward the user.
DR 3900
DR 900 The orientation mark is 2401906 2401906
toward the user.

Before starting
Install the instrument cap on the DR 900 cell holder before ZERO or READ is pushed.
For best results, measure the reagent blank value for each new lot of reagent. Replace the sample with deionized water in
the test procedure to get the reagent blank value. Subtract the reagent blank value from the sample results automatically
with the reagent blank adjust option.
Prepare a boiling water bath for the test procedure. Use finger cots to hold hot sample cells.
Review the Safety Data Sheets (MSDS/SDS) for the chemicals that are used and use any recommended personal protective
equipment.
Dispose of reacted solutions according to local, state and federal regulations. Use the Safety Data Sheets for disposal
information for unused reagents. Consult the environmental, health and safety staff for your facility and/or local regulatory
agencies for further disposal information.

1
Items to collect
Description Quantity

Acid Reagent Powder Pillow 1

®
ChromaVer 3 Chromium Reagent Powder Pillow, 25–mL 1
Chromium 1 Reagent Powder Pillow 1
Chromium 2 Reagent Powder Pillow 1
Hot plate 1
Water bath and rack 1
Finger cots 1 pair
Sample cell for mixing (For information about sample cells, adapters or light shields, refer to
1
Instrument specific information on page 1.)
Sample cells for measurement (For information about sample cells, adapters or light shields,
2
refer to Instrument specific information on page 1.)

Refer to Consumables and replacement items on page 6 for reorder information.

Sample collection and storage

• Collect samples in clean glass or plastic bottles.
• To preserve samples for later analysis, adjust the sample pH to less than 2 with
concentrated nitric acid (about 2 mL per liter). No acid addition is necessary if the
sample is tested immediately.
• Keep preserved samples at room temperature for a maximum of 6 months.
• Before analysis, adjust the pH to 4 with 5.0 N sodium hydroxide standard solution.
• Correct the test result for the dilution from the volume additions.

Powder pillow procedure (alkaline hypobromite oxidation method)

Start

1. Start program 2. Fill a 25-mL sample cell 3. Prepare the sample: 4. Swirl to mix.
100 Chromium, Total. For with 25 mL of sample Add the contents of one
information about sample Chromium 1 Reagent
cells, adapters or light Powder Pillow.
shields, refer to Instrument
specific information
on page 1.
Note: Although the program
name may vary between
instruments, the program
number does not change.

2 Chromium, Total, Alkaline Hypobromite Oxidation Method (0.70 mg/L)

5. Keep the sample cell cap 6. Start the instrument 7. When the timer expires, 8. Add the contents of one
off. Put the prepared sample timer. A 5-minute reaction remove the prepared Chromium 2 Reagent
in a boiling water bath. time starts. sample from the water bath. Powder Pillow.
Put the cap on. Use running
water to cool the sample cell
to 25 °C.

9. Close the sample cell. 10. Add the contents of one 11. Swirl to mix. 12. Add the contents of one
Invert the sample cell to mix. Acid Reagent Powder ChromaVer 3 Chromium
Pillow. Reagent Powder Pillow.

13. Swirl to mix. 14. Start the instrument 15. During the reaction, 16. Prepare the blank:
timer. A 5-minute reaction pour 10-mL from the mixing When the timer expires, fill
time starts. bottle into a sample cell for the second measurement
meansurement. This is the cell with 10 mL of the
prepared sample. original sample.

Chromium, Total, Alkaline Hypobromite Oxidation Method (0.70 mg/L) 3

 Zero

17. Clean the blank. 18. Insert the blank into the 19. Push ZERO. The 20. Clean the prepared
cell holder. display shows 0.00 mg/L Cr. sample.

Read

21. Insert the prepared 22. Push READ. Results

sample into the cell holder. show in mg/L Cr.

Interferences
Interfering substance Interference level
Organic material May inhibit complete oxidation of trivalent chromium. If high levels of organic material are present,
digestion may be required. Complete the analysis as described in this procedure on the digested
sample.
Turbidity For turbid samples, use the test procedure to prepare a 25-mL blank in the same way as the
prepared sample, but do not the add the ChromaVer 3 Chromium Reagent Powder Pillow. Use this
prepared blank to zero the instrument.
Highly buffered samples Can prevent the correct pH adjustment of the sample by the reagents. Sample pretreatment may
or extreme sample pH be necessary.

Accuracy check
Standard additions method (sample spike)
Use the standard additions method (for applicable instruments) to validate the test
procedure, reagents and instrument and to find if there is an interference in the sample.
Items to collect:
• Trivalent Chromium Standard Solution, 50 mg/L Cr3+
• 5-mL volumetric pipet, Class A and pipet filler
• Mixing cylinders (4), 25 mL
• Deionized water
• Pipet, TenSette®, 0.1–1.0 mL and tips

1. Prepare a 12.5 mg/L trivalent chromium standard solution as follows:

a. Use a pipet to add 5.00 mL of a 50 mg/L Cr3+ standard solution into a 25-mL
mixing cylinder.

4 Chromium, Total, Alkaline Hypobromite Oxidation Method (0.70 mg/L)

 b. Dilute to the 20-mL mark with deionized water. Mix well. Prepare this solution
daily.
2. Use the test procedure to measure the concentration of the sample, then keep the
(unspiked) sample in the instrument.
3. Go to the Standard Additions option in the instrument menu.
4. Select the values for standard concentration, sample volume and spike volumes.
5. Prepare three spiked samples: use the TenSette pipet to add 0.1 mL, 0.2 mL and
0.3 mL of the prepared standard solution, respectively, to three 25-mL portions of
fresh sample. Mix well.
6. Use the test procedure to measure the concentration of each of the spiked samples.
Start with the smallest sample spike. Measure each of the spiked samples in the
instrument.
7. Select Graph to compare the expected results to the actual results.
Note: If the actual results are significantly different from the expected results, make sure that
the sample volumes and sample spikes are measured accurately. The sample volumes and
sample spikes that are used should agree with the selections in the standard additions menu. If
the results are not within acceptable limits, the sample may contain an interference.

Standard solution method

Use the standard solution method to validate the test procedure, reagents and
instrument.
Items to collect:
• Trivalent Chromium Standard Solution, Voluette® Ampule, 50 mg/L Cr3+
• 500-mL volumetric flask, Class A
• 5-mL volumetric pipet, Class A and pipet filler
• Deionized water

1. Prepare a 0.50 mg/L trivalent chromium standard solution as follows:

a. Use a pipet to add 5.00 mL of 50 mg/L trivalent chromium standard solution into
the volumetric flask.
b. Dilute to the mark with deionized water. Mix well. Prepare this solution daily.
2. Use the test procedure to measure the concentration of the prepared standard
solution.
3. Compare the expected result to the actual result.
Note: The factory calibration can be adjusted slightly with the standard adjust option so that the
instrument shows the expected value of the standard solution. The adjusted calibration is then
used for all test results. This adjustment can increase the test accuracy when there are slight
variations in the reagents or instruments.

Method performance
The method performance data that follows was derived from laboratory tests that were
measured on a spectrophotometer during ideal test conditions. Users may get different
results under different test conditions.
Program Standard Precision (95% Confidence Interval) Sensitivity
Concentration change per 0.010 Abs change
100 0.500 mg/L Cr 0.47–0.53 mg/L Cr 0.005 mg/L Cr

Summary of method
Trivalent chromium in the sample is oxidized to the hexavalent form by hypobromite ion
under alkaline conditions. The sample is acidified. The total chromium content is
determined by the 1,5-Diphenylcarbohydrazide method. Determine trivalent chromium by
subtracting the results of a separate hexavalent chromium test from the results of the total
chromium test. The measurement wavelength is 540 nm for spectrophotometers or
560 nm for colorimeters.

Chromium, Total, Alkaline Hypobromite Oxidation Method (0.70 mg/L) 5

Consumables and replacement items
Required reagents

Description Quantity/test Unit Item no.

Chromium, total, Reagent Set 1 100/pkg 2242500

Includes:
Acid Reagent Powder Pillow 1 100/pkg 212699
®
ChromaVer 3 Chromium Reagent Powder Pillow, 25-mL 1 100/pkg 1206699
Chromium 1 Reagent Powder Pillow 1 100/pkg 204399
Chromium 2 Reagent Powder Pillow 1 100/pkg 204499

Required apparatus

Description Quantity/test Unit Item no.

Hot plate, 3½-inch diameter, 120 VAC, 50/60 Hz 1 each 1206701

OR
Hot plate, stirrer, 220 - 240 VAC 1 each 2881602
Water bath and rack 1 each 195555

Recommended standards

Description Unit Item no.

Chromium Trivalent Standard Solution, 50-mg/L Cr3+ 100 mL 1415142

Optional reagents and apparatus

Description Unit Item no.

Finger cots 2/pkg 1464702

Flask, volumetric, Class A, 500-mL each 1457449
Pipet, volumetric 5.00-mL each 1451537
®
Pipet, TenSette , 0.1–1.0 mL each 1970001
Pipet tips for TenSette Pipet 1970001 50/pkg 2185696
Pipet, volumetric Class A, 15-mL each 1451539
Pipet filler, safety bulb each 1465100
Cylinder, mixing, 25-mL each 189640

FOR TECHNICAL ASSISTANCE, PRICE INFORMATION AND ORDERING: HACH COMPANY

In the U.S.A. – Call toll-free 800-227-4224 WORLD HEADQUARTERS
Outside the U.S.A. – Contact the HACH office or distributor serving you. Telephone: (970) 669-3050
On the Worldwide Web – www.hach.com; E-mail – [email protected] FAX: (970) 669-2932

© Hach Company/Hach Lange GmbH, 1989–2013. All rights reserved. 04/2013, Edition 8
Color, True and Apparent DOC316.53.01037

Platinum-Cobalt Standard Method1, 2, 3 Method 8025

15 to 500 color units
Scope and application: For water, wastewater and seawater; equivalent to NCASI method 253 and NCASI
Method Color 71.01 for pulp and paper effluent using 465 nm (requires pH adjustment).
1 Adapted from Standard Methods for the Examination of Water and Wastewater and National Council for Air and Stream Improvement
(NCASI) Methods Manual.
2 Adapted from Wat. Res. Vol. 30, No. 11, pp. 2771–2775, 1996.
3 NCASI Method 253 approved at 40 CFR part 136.

Test preparation

Instrument specific information

The table in this section shows all of the instruments that have the program for this test.
Table 1 shows sample cell and orientation requirements for reagent addition tests, such
as powder pillow or bulk reagent tests.
To use the table, select an instrument, then read across to find the corresponding
information for this test.
Table 1 Instrument-specific information for filtered sample
Instrument Sample cell orientation Sample cell
DR 6000 The fill line is to the right. 2495402
DR 3800
DR 2800
DR 2700
DR 5000 The fill line is toward the user.
DR 3900
DR 900 The orientation mark is toward the user. 2401906

Before starting
Install the instrument cap on the DR 900 cell holder before ZERO or READ is pushed.
The NCASI procedure is available only for spectrophotometers and requires pH adjustment to pH 7.6 with 1.0 N HCl or 1.0 N
NaOH. If the overall volume change during the adjustment is more than 1%, start over and use a stronger acid or base. Use
program 125 for the NCASI procedure.
One pH 8 buffer powder pillow (sodium phosphate/potassium phosphate) can be added to 50 mL of sample before the final
pH adjustment to minimize the volume change from the dilution. Mix thoroughly to dissolve before the final pH adjustment.
To test for apparent color, do not filter the sample or the deionized water blank.
For very low levels, use the Pour-Thru Cell (for applicable instruments) for the best results.
Review the Safety Data Sheets (MSDS/SDS) for the chemicals that are used and use any recommended personal protective
equipment.
Dispose of reacted solutions according to local, state and federal regulations. Use the Safety Data Sheets for disposal
information for unused reagents. Consult the environmental, health and safety staff for your facility and/or local regulatory
agencies for further disposal information.

1
Items to collect
Description Quantity

Buffer, pH 8.0 (optional) 1

Hydrochloric Acid Solution, 1.0 N (for program 125) varies
Sodium Hydroxide, 1.00 N (for program 125) varies
Water, deionized 100 mL
Filter apparatus: membrane filter, filter holder, filter flask and aspirator 1
Stopper, rubber, one hole number 7 1
Tubing, rubber 1
Sample cells (For information about sample cells, adapters or light shields, refer to Instrument
2
specific information on page 1.)

Refer to Consumables and replacement items on page 4 for reorder information.

Sample collection and storage

• Collect samples in clean glass or plastic bottles.
• Analyze the samples as soon as possible for best results.
• If prompt analysis is impossible, fill the bottle completely full, then tighten the cap on
the bottle. Avoid excessive agitation or prolonged contact with air.
• To preserve samples for later analysis, keep the samples at or below 6 °C (43 °F) for
up to 48 hours.
• Let the sample temperature increase to room temperature before analysis.

Platinum-Cobalt procedure

Start

1. Start program 120 Color, 2. Collect 200 mL of 3. Assemble the 4. Pour approximately
455 nm or program sample in a 400-mL beaker. 0.45 micron filter apparatus. 50 mL of deionized water
125 Color, 465 nm for the NCASI: Adjust the pH as NCASI: The NCASI test through the filter to rinse the
NCASI test. Colorimeters described in Before starting uses a 0.8-micron filter. A filter. Discard the rinse
use program 122 Color, on page 1. 1.0 micron prefilter can be water.
420 nm. For information used first for samples that
about sample cells, are difficult to filter.
adapters or light shields,
refer to Instrument specific
information on page 1.
Note: Although the program
name may vary between
instruments, the program
number does not change.

2 Color, True and Apparent, Platinum-Cobalt Method (500 units)

5. Pour another 50 mL of 6. Prepare the blank: Fill 7. Pour approximately 8. Prepare the sample: Fill
deionized water through the the sample cell with 10 mL 50 mL of sample through a second sample cell with
filter. of filtered deionized water the filter. 10 mL of filtered sample.
from the previous step.
Discard the excess water in
the flask.

Zero

9. Clean the blank. 10. Insert the blank into the 11. Push ZERO. The 12. Clean the prepared
cell holder. display shows 0 units Pt-Co. sample.

Read

13. Insert the prepared 14. Push READ. Results

sample into the cell holder. show in units Pt-Co.

Accuracy check
Standard solution method
Use the standard solution method to validate the test procedure, reagents and
instrument.
Items to collect:
• 500 Platinum-Cobalt Units Standard Solution
• 100-mL volumetric flask, Class A
• 50-mL volumetric pipet, Class A and pipet filler
• Deionized water

1. Prepare a 250 platinum-cobalt units standard solution as follows:

a. Use a pipet to add 50.00 mL of 500 platinum-cobalt units standard solution into
the volumetric flask.

Color, True and Apparent, Platinum-Cobalt Method (500 units) 3

 b. Dilute to the mark with deionized water. Mix well. Prepare this solution daily.
2. Use the test procedure to measure the concentration of the prepared standard
solution.
3. Compare the expected result to the actual result.
Note: The factory calibration can be adjusted slightly with the standard adjust option so that the
instrument shows the expected value of the standard solution. The adjusted calibration is then
used for all test results. This adjustment can increase the test accuracy when there are slight
variations in the reagents or instruments.

Method performance
The method performance data that follows was derived from laboratory tests that were
measured on a spectrophotometer during ideal test conditions. Users may get different
results under different test conditions.
Program Standard Precision (95% Confidence Interval) Sensitivity
Concentration change per 0.010 Abs change
120 250 units Pt-Co 245–255 units Pt-Co 16 units Pt-Co
125 250 units Pt-Co 245–255 units Pt-Co 16 units Pt-Co

Summary of method
Color may be expressed as “apparent” or “true” color. The apparent color includes color
from dissolved materials plus color from suspended matter. The true color is determined
by removal of the suspended materials with a filter or a centrifuge. This procedure uses
filtration for true color analysis. To measure apparent color, do not filter the sample or the
deionized water blank. The same instrument program is used for both true and apparent
color. The stored program is calibrated in color units based on the APHA-recommended
standard of 1 color unit being equal to 1 mg/L platinum as chloroplatinate ion. Test results
for Programs 120 and 125 are measured at 455 and 465 nm, respectively in
spectrophotometers. Test results for Program 122 are measured at 420 nm in
colorimeters.
Consumables and replacement items
Required reagents

Description Quantity/test Unit Item no.

Hydrochloric Acid Solution, 1.0 N varies 1L 2321353

Sodium Hydroxide, 1.00 N varies 1L 104553
Water, deionized varies 4L 27256

Required apparatus

Description Quantity/test Unit Item no.

Aspirator, vacuum pump 1 each 213100

Beaker, 400 mL 1 each 50048
Filter, membrane, 47-mm, 0.8-microns, Program 125 1 100/pkg 2640800
Filter holder, 47 mm, magnetic base 1 each 1352900
Filter, membrane, 47-mm, 0.45-microns, Program 120 1 100/pkg 1353000
Flask, filtering, 500-mL 1 each 54649
Stopper, poly, hollow 1 6/pkg 211907
Tubing, rubber, 7.9 x 2.4 mm varies 12 ft 56019

4 Color, True and Apparent, Platinum-Cobalt Method (500 units)

Recommended standards and apparatus

Description Unit Item no.

Buffer, pH 8.0 15/pkg 1407995

Color Standard Solution, 500 platinum-cobalt units 1L 141453
Color Standard Solution, 15 platinum-cobalt units 1L 2602853
®
Color Standard Solution, 500 platinum-cobalt units, 10-mL Voluette Ampules 16/pkg 141410
Filter, glass microfiber, 47-mm, 1.0 micron 100/pk 2551400
Flask, volumetric, Class A, 100-mL each 1457442
Pipet, volumetric, Class A, 50-mL each 1451541
Pipet filler, safety bulb each 1465100

Color, True and Apparent, Platinum-Cobalt Method (500 units) 5

 FOR TECHNICAL ASSISTANCE, PRICE INFORMATION AND ORDERING: HACH COMPANY
In the U.S.A. – Call toll-free 800-227-4224 WORLD HEADQUARTERS
Outside the U.S.A. – Contact the HACH office or distributor serving you. Telephone: (970) 669-3050
On the Worldwide Web – www.hach.com; E-mail – [email protected] FAX: (970) 669-2932

© Hach Company/Hach Lange GmbH, 1989–2013. All rights reserved. 04/2013, Edition 8
Copper DOC316.53.01039

USEPA1 Bicinchoninate Method2 Method 8506 (CuVer 1) and Method 8026 (CuVer 2)
®
0.04 to 5.00 mg/L Cu Powder Pillows or AccuVac Ampuls
Scope and application: For water, wastewater and seawater3; Method 8506 USEPA approved for reporting
wastewater analysis (digestion required)4
1 Approved, USEPA and Standard Method 3500 Cu C or E.
2 Adapted from Nakano, S., Yakugaku Zasshi, 82 486-491 (1962) [Chemical Abstracts, 58 3390e (1963)].
3 Pretreatment required for the powder pillow method- refer to the Interference section.
4 Federal Register, 45 (105) 36166 (May 29, 1980).

Test preparation

Instrument-specific information
The tables in this section show all of the instruments that have the program for this test.
Table 1 shows sample cell and orientation requirements for reagent addition tests, such
as powder pillow or bulk reagent tests. Table 2 shows sample cell and adapter
requirements for AccuVac Ampul tests.
To use either table, select an instrument, then read across to find the corresponding
information for this test.
Table 1 Instrument-specific information for powder pillows
Instrument Sample cell orientation Sample cell
DR 6000 The fill line is to the right. 2495402
DR 3800
DR 2800
DR 2700
DR 5000 The fill line is toward the user.
DR 3900
DR 900 The orientation mark is toward the user. 2401906

Table 2 Instrument-specific information for AccuVac Ampuls

Instrument Adapter Sample cell
DR 6000 — 2427606
DR 5000
DR 900
DR 3900 LZV846 (A)
DR 3800 LZV584 (C) 2122800
DR 2800
DR 2700

1
Before starting
Install the instrument cap on the DR 900 cell holder before ZERO or READ is pushed.
The sample must be digested with heat and acid to make sure that all forms of the metal are measured. Use the mild or
vigorous digestion. Refer to the Water Analysis Guide for more information.
For best results, measure the reagent blank value for each new lot of reagent. Replace the sample with deionized water in
the test procedure to get the reagent blank value. Subtract the reagent blank value from the sample results automatically
with the reagent blank adjust option.
Review the Safety Data Sheets (MSDS/SDS) for the chemicals that are used and use any recommended personal protective
equipment.
Dispose of reacted solutions according to local, state and federal regulations. Use the Safety Data Sheets for disposal
information for unused reagents. Consult the environmental, health and safety staff for your facility and/or local regulatory
agencies for further disposal information.

Items to collect
Powder pillows

Description Quantity
®
CuVer 1 Copper Reagent Powder Pillow, 10-mL 1
Sample cells. (For information about sample cells, adapters or light shields, refer to Instrument-
2
specific information on page 1.)

Refer to Consumables and replacements items on page 7 for reorder information.

AccuVac Ampuls

Description Quantity
® ®
CuVer 2 Reagent AccuVac Ampul 1
Beaker, 50-mL 1
Sample cells (For information about sample cells, adapters or light shields, refer to Instrument-
1
specific information on page 1.)
Stopper for 18-mm tubes and AccuVac Ampuls 1

Refer to Consumables and replacements items on page 7 for reorder information.

Sample collection and storage

• Collect samples in clean glass or plastic bottles.
• To preserve samples for later analysis, adjust the sample pH to less than 2 with
concentrated nitric acid (about 2 mL per liter). No acid addition is necessary if the
sample is tested immediately.
• If only dissolved copper is to be determined, filter the sample before the acid addition.
• Keep preserved samples at room temperature for a maximum of 6 months.
• Before analysis, adjust the pH to 4–6 with 8.0 N potassium hydroxide standard
solution (do not exceed pH 6, as copper may precipitate).
• Correct the test result for the dilution from the volume additions.

2 Copper, Bicinchoninate Method (5.00 mg/L)

Powder pillow procedure (Method 8506)

Start

1. Start program 2. Prepare the sample: Fill 3. Add the contents of one 4. Swirl to mix.
135 Copper, Bicin. For a sample cell with 10 mL of CuVer 1 Copper reagent
information about sample sample. powder pillow.
cells, adapters or light
shields, refer to Instrument-
specific information
on page 1.
Note: Although the program
name may vary between
instruments, the program
number does not change.

5. Start the instrument 6. Prepare the blank: Fill a 7. Clean the blank. 8. Insert the blank into the
timer. A 2-minute reaction second sample cell with cell holder.
time starts. 10 mL of sample.
The sample shows a purple
color when copper in the
sample mixes with the
reagent powder.
Undissolved powder does
not affect accuracy.

Zero Read

9. Push ZERO. The display 10. Clean the prepared 11. Within 30 minutes after 12. Push READ. Results
shows 0.00 mg/L Cu. sample. the timer expires, insert the show in mg/L Cu.
prepared sample into the
cell holder.

Copper, Bicinchoninate Method (5.00 mg/L) 3

AccuVac Ampul procedure (Method 8026)

Start

1. Start program 2. Prepare the blank: Fill 3. Prepare the sample: 4. Quickly invert the Ampul
140 Copper, Bicin. AV. For the sample cell with 10 mL Collect at least 40 mL of several times to mix.
information about sample of sample. sample in a 50-mL beaker.
cells, adapters or light Fill the AccuVac Ampul with
shields, refer to Instrument- sample. Keep the tip
specific information immersed while the Ampul
on page 1. fills completely.
Note: Although the program
name may vary between
instruments, the program
number does not change.

Zero

5. Start the instrument 6. When the timer expires, 7. Insert the blank into the 8. Push ZERO. The display
timer. A 2-minute reaction clean the blank. cell holder. shows 0.00 mg/L Cu.
time starts.
The sample shows a purple
color when copper in the
sample mixes with the
reagent powder.
Undissolved powder does
not affect accuracy.

Read

9. Clean the AccuVac 10. Within 30 minutes of 11. Push READ. Results
Ampul. adding the reagent, insert show in mg/L Cu.
the prepared sample
AccuVac Ampul into the cell
holder.

4 Copper, Bicinchoninate Method (5.00 mg/L)

Interferences
Table 3 gives treatments for powder pillows. Table 4 gives treatments for AccuVac
Ampuls. To differentiate free copper from that complexed to EDTA or other complexing
agents, use a 25-mL sample cell and Free Copper Reagent Powder Pillow instead of the
CuVer 1 Powder Pillow in the test procedure. Add a Hydrosulfite Reagent Powder Pillow
to the same sample and read the result again. This result is the total dissolved copper
(free and complexed). Unlike CuVer 1 Reagent, CuVer 2 Reagent Powder Pillows and
AccuVac Ampuls react directly with copper that is complexed by chelants such as EDTA.
Table 3 Interfering substances and suggested treatments for powder pillows
Interfering Interference level
substance
Acidity If the sample is extremely acidic (pH 2 or less), a precipitate may form. Add 8 N Potassium Hydroxide
Standard Solution by drops until the sample pH is above 4, then start the test.
Aluminum, Al3+ Use the powder pillow procedure, but use a CuVer 2 Copper Reagent Powder Pillow and not the
CuVer 1 Pillow. Results include total dissolved copper (free and complexed). Use a 25-mL sample
volume.
Cyanide, CN- Prevents full color development. Before the CuVer 1 Powder Pillow Reagent is added, add 0.2 mL of
formaldehyde to the 10-mL sample. Wait 4 minutes, then take the reading. Multiply the test results by
1.02 to correct for sample dilution by the formaldehyde.
Hardness Use the powder pillow procedure, but use a CuVer 2 Copper Reagent Powder Pillow and not the
CuVer 1 Pillow. Results include total dissolved copper (free and complexed). Use a 25-mL sample
volume.
Iron, Fe3+ Use the powder pillow procedure, but use a CuVer 2 Copper Reagent Powder Pillow and not the
CuVer 1 Pillow. Results include total dissolved copper (free and complexed). Use a 25-mL sample
volume.
Silver, Ag+ If a turbidity remains and turns black, silver interference is likely. Add 20 drops of 50% saturated
Potassium Chloride Solution to 75 mL of sample, then filter through a fine or highly retentive filter. Use
the filtered sample in the test procedure.

Table 4 Interfering substances and suggested treatments for AccuVac Ampuls

Interfering substance Interference level
Acidity If the sample is extremely acidic (pH 2 or less), a precipitate may form. Add 8 N Potassium
Hydroxide Standard Solution by drops until the sample pH is above 4, then start the test.
Aluminum, Al3+ Reagents accommodate high levels.
Cyanide, CN- Prevents full color development. Add 0.5 mL of formaldehyde per 25-mL of sample, then use the
CuVer 2 Reagent AccuVac Ampul. Wait 4 minutes, then take the reading. Multiply the test results by
1.02 to correct for sample dilution by the formaldehyde.
Hardness Reagents accommodate high levels.
Iron, Fe3+ Reagents accommodate high levels.
Silver, Ag+ If a turbidity remains and turns black, silver interference is likely. Add 10 drops of saturated
Potassium Chloride Solution to 75 mL of sample, then filter through a fine or highly retentive filter.
Use the filtered sample in the procedure.

Accuracy check
Standard additions method (sample spike)
Use the standard additions method (for applicable instruments) to validate the test
procedure, reagents and instrument and to find if there is an interference in the sample.
Items to collect:
• 100 mg/L Copper Standard Solution
• 5-mL volumetric pipet, Class A and pipet filler

Copper, Bicinchoninate Method (5.00 mg/L) 5

 • Mixing cylinder, 50 mL
• Deionized water
• Pipet, TenSette®, 0.1–1.0 mL and tips

1. Prepare a 12.5 mg/L copper standard solution as follows:

a. Use a pipet to add 5.00 mL of a 100 mg/L copper standard solution into a 50-mL
mixing cylinder.
b. Dilute to the 40-mL mark with deionized water. Mix well. Prepare this solution
daily.
2. Use the test procedure to measure the concentration of the sample, then keep the
(unspiked) sample in the instrument.
3. Go to the Standard Additions option in the instrument menu.
4. Select the values for standard concentration, sample volume and spike volumes.
5. Prepare three spiked samples: use the TenSette pipet to add 0.1 mL, 0.2 mL and
0.3 mL of the prepared standard solution, respectively, to three 10-mL portions of
fresh sample. Mix well.
Note: For AccuVac® Ampuls, add 0.2 mL, 0.4 mL and 0.6 mL of a 75 mg/L Copper Voluette
Ampule Standard to three 50-mL portions of fresh sample.
6. Use the test procedure to measure the concentration of each of the spiked samples.
Start with the smallest sample spike. Measure each of the spiked samples in the
instrument.
7. Select Graph to compare the expected results to the actual results.
Note: If the actual results are significantly different from the expected results, make sure that
the sample volumes and sample spikes are measured accurately. The sample volumes and
sample spikes that are used should agree with the selections in the standard additions menu. If
the results are not within acceptable limits, the sample may contain an interference.

Standard solution method

Use the standard solution method to validate the test procedure, reagents and
instrument.
Items to collect:
• Copper Standard Solution, 100-mg/L
• 100-mL volumetric flask, Class A
• 4-mL volumetric pipet, Class A and pipet filler
• Deionized water

1. Prepare a 4.00 mg/L copper standard solution as follows:

a. Use a pipet to add 4.00 mL of 100 mg/L copper standard solution into the
volumetric flask.
b. Dilute to the mark with deionized water. Mix well. Prepare this solution daily.
2. Use the test procedure to measure the concentration of the prepared standard
solution.
3. Compare the expected result to the actual result.
Note: The factory calibration can be adjusted slightly with the standard adjust option so that the
instrument shows the expected value of the standard solution. The adjusted calibration is then
used for all test results. This adjustment can increase the test accuracy when there are slight
variations in the reagents or instruments.

6 Copper, Bicinchoninate Method (5.00 mg/L)

Method performance
The method performance data that follows was derived from laboratory tests that were
measured on a spectrophotometer during ideal test conditions. Users may get different
results under different test conditions.
Program Standard Precision (95% Confidence Interval) Sensitivity
Concentration change per 0.010 Abs change
135 1.00 mg/L Cu 0.97–1.03 mg/L Cu 0.04 mg/L Cu
140 1.00 mg/L Cu 0.97–1.03 mg/L Cu 0.03 mg/L Cu

Summary of method
Copper in the sample reacts with a salt of bicinchoninic acid contained in CuVer 1 or
CuVer 2 Copper Reagent to form a purple colored complex in proportion to the copper
concentration. The measurement wavelength is 560 nm.
Consumables and replacements items
Required reagents

Description Quantity/test Unit Item no.

®
CuVer 1 Copper Reagent Powder Pillow, 10-mL 1 100/pkg 2105869
OR
® ®
CuVer 2 Copper Reagent AccuVac Ampul 1 25/pkg 2504025

Required apparatus

Description Quantity/test Unit Item no.

AccuVac Snapper 1 each 2405200

Beaker, 50-mL 1 each 50041H
Stoppers for 18-mm tubes and AccuVac Ampuls 2 6/pkg 173106

Recommended standards

Description Unit Item no.

Copper Standard Solution, 100-mg/L as Cu 100 mL 12842

®
Copper Voluette Ampule Standard, 75-mg/L as Cu, 10-mL 16/pkg 1424710
Metals Drinking Water Standard, LR for Cu, Fe, Mn 500 mL 2833749
Metals Drinking Water Standard, HR for Cu, Fe, Mn 500 mL 2833649

Optional reagents and apparatus

Description Unit Item no.

Beaker, 50-mL each 50041H

CuVer 2 Copper Reagent Powder Pillow 100/pkg 2188299
Cylinder, mixing, 50-mL each 189641
100 mL
Formaldehyde, ACS 205932
MDB
Nitric Acid, concentrated 500 mL 15249
Potassium Chloride Solution, 50% saturated 25 mL 1429323

Copper, Bicinchoninate Method (5.00 mg/L) 7

Consumables and replacements items (continued)
Description Unit Item no.

100 mL
Potassium Hydroxide Standard Solution, 8 N 28232H
MDB
Sample cells, 25 mL, matched, 1" square 2/pkg 2612602
®
AccuVac Snapper each 2405200
®
Ampule Breaker, Voluette ampules each 2196800
Sample cells, 25 mm round 6/pkg 2401906
Copper, Free and Total Reagent set, includes: each 2439200
Hydrosulfite Reagent Powder Pillows 100/pkg 2118869
Copper, Free, Reagent Powder Pillows 100/pkg 2182369

FOR TECHNICAL ASSISTANCE, PRICE INFORMATION AND ORDERING: HACH COMPANY

In the U.S.A. – Call toll-free 800-227-4224 WORLD HEADQUARTERS
Outside the U.S.A. – Contact the HACH office or distributor serving you. Telephone: (970) 669-3050
On the Worldwide Web – www.hach.com; E-mail – [email protected] FAX: (970) 669-2932

© Hach Company/Hach Lange GmbH, 1989–2013. All rights reserved. 04/2013, Edition 8
Copper DOC316.53.01038

Porphyrin Method1 Method 8143

1 to 210 µg/L Cu (LR) Powder Pillows
Scope and application: For water, wastewater and seawater.
1 Adapted from Ishii and Koh, Bunseki Kagaku, 28 (473), 1979.

Test preparation

Instrument specific information

The table in this section shows all of the instruments that have the program for this test.
Table 1 shows sample cell and orientation requirements for reagent addition tests, such
as powder pillow or bulk reagent tests.
To use the table, select an instrument, then read across to find the corresponding
information for this test.
Table 1 Instrument-specific information for powder pillows
Instrument Sample cell orientation Sample cell
DR 6000 The fill line is to the right. 2495402
DR 3800
DR 2800
DR 2700
DR 5000 The fill line is toward the user.
DR 3900
DR 900 The orientation mark is toward the user. 2401906

Before starting
Install the instrument cap on the DR 900 cell holder before ZERO or READ is pushed.
The sample must be digested with heat and acid to make sure that all forms of the metal are measured. Use the mild or
vigorous digestion. Refer to the Water Analysis Guide for more information.
Wash all glassware with detergent. Rinse with tap water. Rinse again with 1:1 nitric acid solution. Rinse a third time with
high-quality deionized water. These steps will remove deposits that can cause slightly high results.
If samples contain high levels of metals, a slight metallic deposit or yellow buildup may form in the sample cell. Wash the cell
as described above.
For best results, measure the reagent blank value for each new lot of reagent. Replace the sample with deionized water in
the test procedure to get the reagent blank value. Subtract the reagent blank value from the sample results automatically
with the reagent blank adjust option.
Review the Safety Data Sheets (MSDS/SDS) for the chemicals that are used and use any recommended personal protective
equipment.
Dispose of reacted solutions according to local, state and federal regulations. Use the Safety Data Sheets for disposal
information for unused reagents. Consult the environmental, health and safety staff for your facility and/or local regulatory
agencies for further disposal information.

1
Items to collect
Description Quantity

Copper Masking Reagent Powder Pillows, 10-mL 1

Porphyrin 1 Reagent Powder Pillows, 10-mL 2
Porphyrin 2 Reagent Powder Pillows, 10-mL 2
Nitric Acid Solution, 1:1 varies
Sample cells (For information about sample cells, adapters or light shields, refer to Instrument
2
specific information on page 1.)

Refer to Consumables and replacement items on page 5 for reorder information.

Sample collection and storage

• Collect samples in acid-washed plastic bottles.
• To preserve samples for later analysis, adjust the sample pH to less than 2 with
concentrated nitric acid (about 2 mL per liter). No acid addition is necessary if the
sample is tested immediately.
• Keep preserved samples at room temperature for a maximum of 6 months.
• Before analysis, adjust the pH to 2–6 with 5.0 N sodium hydroxide standard solution.
• Correct the test result for the dilution from the volume additions.

Powder pillow procedure

Start

1. Start program 2. Prepare the blank: Fill 3. Add the contents of one 4. Swirl to dissolve the
145 Copper, Porphyrin. the sample cell with 10 mL Copper Masking Reagent reagent.
For information about of sample. powder pillow to the sample
sample cells, adapters or cell to create the blank.
light shields, refer to
Instrument specific
information on page 1.
Note: Although the program
name may vary between
instruments, the program
number does not change.

2 Copper, Porphyrin Method (210 µg/L)

5. Prepare the sample: Fill 6. Add the contents of one 7. Swirl to mix. 8. Add the contents of one
a second sample cell with Porphyrin 1 Reagent Porphyrin 2 Reagent
10 mL of sample. Powder Pillow to each Powder Pillow to each
sample cell. sample cell.

9. Swirl to mix. 10. Start the instrument 11. When the timer expires, 12. Insert the blank into the
If copper is present in the timer. A 3-minute reaction clean the blank. cell holder.
sample, the sample will time starts.
show blue, then go back to
a yellow color.

Zero Read

13. Push ZERO. The 14. Clean the prepared 15. Insert the prepared 16. Push READ. Results
display shows 0 µg/L Cu. sample. sample into the cell holder. show in µg/L Cu.

Interferences
Interfering substance Interference level
Aluminum, Al3+ 60 mg/L
Cadmium, Cd2+ 10 mg/L
Calcium, Ca2+ 1500 mg/L
Chelating agents Interfere at all levels unless either the Digesdahl or vigorous digestion is
completed.
Chloride, Cl- 90,000 mg/L
Chromium, Cr6+ 110 mg/L

Copper, Porphyrin Method (210 µg/L) 3

Interfering substance Interference level
Cobalt, Co2+ 100 mg/L
Fluoride, F- 30,000 mg/L
Iron, Fe2+ 6 mg/L
Lead, Pb2+ 3 mg/L
Magnesium 10,000 mg/L
Manganese 140 mg/L
Mercury, Hg2+ 3 mg/L
Molybdenum 11 mg/L
Nickel, Ni2+ 60 mg/L
Potassium, K+ 60,000 mg/L
Sodium, Na+ 90,000 mg/L
Zinc, Zn2+ 9 mg/L
Highly buffered samples or extreme sample pH Can prevent the correct pH adjustment of the sample by the reagents.
Sample pretreatment may be necessary.

Accuracy check
Standard additions method (sample spike)
Use the standard additions method (for applicable instruments) to validate the test
procedure, reagents and instrument and to find if there is an interference in the sample.
Items to collect:
• Copper Standard Solution, 4 mg/L (PourRite® Ampule or prepare from a dilution of a
higher concentration copper standard solution)
• Ampule breaker
• Pipet, TenSette®, 0.1–1.0 mL and tips

1. Use the test procedure to measure the concentration of the sample, then keep the
(unspiked) sample in the instrument.
2. Go to the Standard Additions option in the instrument menu.
3. Select the values for standard concentration, sample volume and spike volumes.
4. Open the standard solution.
5. Prepare six spiked 10-mL samples: use the TenSette pipet to add 0.1 mL, 0.2 mL and
0.3 mL of the standard solution, respectively, to two 10-mL portions of fresh sample.
Mix well.
6. Use the test procedure to measure the concentration of each of the spiked samples.
Start with the smallest sample spike. Measure each of the spiked samples in the
instrument.
7. Select Graph to compare the expected results to the actual results.
Note: If the actual results are significantly different from the expected results, make sure that
the sample volumes and sample spikes are measured accurately. The sample volumes and
sample spikes that are used should agree with the selections in the standard additions menu. If
the results are not within acceptable limits, the sample may contain an interference.

Standard solution method

Use the standard solution method to validate the test procedure, reagents and
instrument.
Items to collect:
• Copper Standard Solution, 10 mg/L
• 1000-mL volumetric flask, Class A

4 Copper, Porphyrin Method (210 µg/L)

 • 15-mL volumetric pipet, Class A and pipet filler
• Deionized water

1. Prepare a 150 µg/L copper standard solution as follows:

a. Use a pipet to add 15.00 mL of 10-mg/L copper standard solution into the
volumetric flask.
b. Dilute to the mark with deionized water. Mix well. Prepare this solution daily.
2. Use the test procedure to measure the concentration of the prepared standard
solution.
3. Compare the expected result to the actual result.
Note: The factory calibration can be adjusted slightly with the standard adjust option so that the
instrument shows the expected value of the standard solution. The adjusted calibration is then
used for all test results. This adjustment can increase the test accuracy when there are slight
variations in the reagents or instruments.

Method performance
The method performance data that follows was derived from laboratory tests that were
measured on a spectrophotometer during ideal test conditions. Users may get different
results under different test conditions.
Program Standard Precision (95% Confidence Interval) Sensitivity
Concentration change per 0.010 Abs change
145 50 µg/L Cu 47–53 µg/L Cu 1 µg/L Cu

Summary of method
The porphyrin method is very sensitive to trace amounts of free copper. The method is
free from most interferences and does not require any sample extraction or concentration
before analysis. Interferences from other metals are removed with the copper masking
reagent. The porphyrin indicator forms an intense, yellow-colored complex with any free
copper present in sample. The measurement wavelength is 425 nm for
spectrophotometers or 420 nm for colorimeters.
Consumables and replacement items
Required reagents and apparatus

Description Quantity/test Unit Item no.

Nitric Acid Solution, 1:1 varies 500 mL 254049

Copper Reagent Set, Porphyrin, 10-mL 1 100/pkg 2603300
Includes:
Copper Masking Reagent Powder Pillow, 10-mL 1 100/pkg 2603449
Porphyrin 1 Reagent Powder Pillow, 10-mL 2 100/pkg 2603549
Porphyrin 2 Reagent Powder Pillow, 10-mL 2 100/pkg 2603649

Recommended standards

Description Unit Item no.

Copper Standard Solution, 4 mg/L, 2 mL Pour-Rite Ampules 20/pkg 2605720

Copper Standard Solution, 10-mg/L Cu 100 mL 12932
Water, deionized 4L 27256

Copper, Porphyrin Method (210 µg/L) 5

Optional reagents and apparatus

Description Unit Item no.

100 mL
Sodium Hydroxide Standard Solution, 5.0 N 245032
MDB
®
Pipet, TenSette , 0.1–1.0 mL each 1970001
Pipet tips for TenSette Pipet 1970001 50/pkg 2185696
Pipet, volumetric Class A, 15-mL each 1451539
Flask, volumetric, Class A, 1000-mL each 1457453
Pipet filler, safety bulb each 1465100
Sample cells, 1" square matched set 8/pkg 2495408
Paper, pH, 0–14 pH range 100/pkg 2601300

FOR TECHNICAL ASSISTANCE, PRICE INFORMATION AND ORDERING: HACH COMPANY

In the U.S.A. – Call toll-free 800-227-4224 WORLD HEADQUARTERS
Outside the U.S.A. – Contact the HACH office or distributor serving you. Telephone: (970) 669-3050
On the Worldwide Web – www.hach.com; E-mail – [email protected] FAX: (970) 669-2932

© Hach Company/Hach Lange GmbH, 1989–2013. All rights reserved. 04/2013, Edition 8
Cyanide DOC316.53.01040

Pyridine-Pyrazalone Method1 Method 8027

0.002 to 0.240 mg/L CN– Powder Pillows
Scope and application: For water, wastewater and seawater.
1 Adapted from Epstein, Joseph, Anal. Chem. 19(4), 272 (1947).

Test preparation

Instrument specific information

The table in this section shows all of the instruments that have the program for this test.
Table 1 shows sample cell and orientation requirements for reagent addition tests, such
as powder pillow or bulk reagent tests.
To use the table, select an instrument, then read across to find the corresponding
information for this test.
Table 1 Instrument-specific information for powder pillows
Instrument Sample cell orientation Sample cell
DR 6000 The fill line is to the right. 2495402
DR 3800
DR 2800
DR 2700
DR 5000 The fill line is toward the user.
DR 3900
DR 900 The orientation mark is toward the user. 2401906

Before starting
Install the instrument cap on the DR 900 cell holder before ZERO or READ is pushed.
All samples to be analyzed for cyanide should be treated by acid distillation except when experience has shown that there is
no difference in results obtained with or without distillation.
Use a water bath to keep the temperature for the reaction in this test at the optimum 25 °C for best results. Samples less
than 23 °C need longer reaction times and samples more than 25 °C give low results.
The timing during the test procedure is critical. Open the necessary reagents before this procedure is started for best results.
For best results, measure the reagent blank value for each new lot of reagent. Replace the sample with deionized water in
the test procedure to get the reagent blank value. Subtract the reagent blank value from the sample results automatically
with the reagent blank adjust option.
Review the Safety Data Sheets (MSDS/SDS) for the chemicals that are used and use any recommended personal protective
equipment.
Dispose of reacted solutions according to local, state and federal regulations. Use the Safety Data Sheets for disposal
information for unused reagents. Consult the environmental, health and safety staff for your facility and/or local regulatory
agencies for further disposal information.

1
Items to collect
Description Quantity
®
CyaniVer Cyanide 3 Reagent Powder Pillow, 10-mL 1
®
CyaniVer Cyanide 4 Reagent Powder Pillow, 10-mL 1
®
CyaniVer Cyanide 5 Reagent Powder Pillow, 10-mL 1
Cylinder, graduated, 10-mL 1
Sample cells (For information about sample cells, adapters or light shields, refer to Instrument
2
specific information on page 1.)

Refer to Consumables and replacement items on page 7 for reorder information.

Sample collection and storage

• Collect samples in clean glass or plastic bottles.
• The presence of oxidizing agents, sulfides and fatty acids can cause the loss of
cyanide during sample storage. Samples that contain these substances must be
pretreated as described in the sections that follow before preservation with sodium
hydroxide. If the sample contains sulfide and is not pretreated, it must be analyzed
within 24 hours.
• To preserve samples for later analysis, adjust the sample pH to a minimum pH
12 with 5.0 N sodium hydroxide standard solution (about 4 mL per liter). Use a glass
serological pipet and pipet filler. Measure the pH and add more sodium hydroxide if
necessary.
• Keep the preserved samples at or below 6°C (43 °F) for up to 14 days.
• Before analysis, adjust the pH to 7 with 2.5 N hydrochloric acid standard solution.
• Let the sample temperature increase to room temperature before analysis.
• Correct the test result for the dilution from the volume additions.
Oxidizing agents
Oxidizing agents such as chlorine decompose cyanides during storage. To test for and
remove oxidizing agents, pretreat the sample as follows:
1. Measure 25-mL of the sample and add one drop of 10-g/L m-Nitrophenol Indicator
Solution. Swirl to mix.
2. Add 2.5 N Hydrochloric Acid Standard Solution by drops until the color changes from
yellow to colorless. Swirl the sample thoroughly after the addition of each drop.
3. Add two drops of Potassium Iodide Solution, 30-g/L and two drops of Starch Indicator
Solution to the sample. Swirl to mix. The solution will turn blue if oxidizing agents are
present.
4. If the color is blue, add two level, 1-g measuring spoonfuls of ascorbic acid per liter of
sample.
5. Remove a 25-mL portion of the treated sample and repeat steps 1 to 3. If the sample
turns blue, repeat steps 4 and 5.
6. If the 25-mL sample remains colorless, preserve the remaining sample to pH 12 for
storage with 5 N Sodium Hydroxide Standard Solution.
7. Complete the procedure given under Interfering Substances and Levels, Reducing
Agents, to eliminate the effect of excess ascorbic acid, before the cyanide procedure
is started.
Sulfides
Sulfides will quickly convert cyanide to thiocyanate (SCN–). To test for and remove
sulfide, pretreat the sample as follows:
1. Put a drop of sample on a disc of Hydrogen Sulfide Test Paper that has been wetted
with pH 4 Buffer Solution.

2 Cyanide, Pyridine-Pyrazalone Method (0.240 mg/L)

 2. If the test paper darkens, add a 1-g measuring spoon of Lead Acetate to the sample.
Repeat step 1.
3. If the test paper continues to turn dark, keep adding Lead Acetate until the sample
tests negative for sulfide.
4. Filter the lead sulfide precipitate through Filter Paper and a Funnel. Preserve the
sample for storage with 5 N Sodium Hydroxide Standard Solution or neutralize to a
pH of 7 for analysis.
Fatty acids
CAUTION Perform this operation under a ventilation hood and complete as quickly as
possible.
When distilled, fatty acids will pass over with cyanide and under the alkaline conditions of
the absorber, will form soaps. If the presence of fatty acid is suspected, use the following
pretreatment before preserving samples with sodium hydroxide.
1. Acidify 500 mL of sample to pH 6 or 7 with a 4:1 dilution of glacial Acetic Acid.
2. Pour the sample into a 1000-mL separation funnel and add 50 mL of Hexane.
3. Stopper the funnel and shake for 1 minute. Allow the layers to separate.
4. Drain off the lower sample layer into a 600-mL beaker. If the sample is to be stored,
add enough 5 N Sodium Hydroxide Standard Solution to raise the pH to a minimum
pH 12.

Powder pillow procedure

Start

1. Start program 2. Prepare the sample: Fill 3. Add the contents of one 4. Close the sample cell.
160 Cyanide. For a sample cell with 10 mL of CyaniVer 3 Cyanide Shake the sample cell for
information about sample sample. Reagent Powder Pillow. 30 seconds. Let the sample
cells, adapters or light cell sit for another
shields, refer to Instrument 30 seconds.
specific information
on page 1.
Note: Although the program
name may vary between
instruments, the program
number does not change.

Cyanide, Pyridine-Pyrazalone Method (0.240 mg/L) 3

5. Add the contents of one 6. Close the sample cell. 7. Add the contents of one 8. Close the sample cell.
CyaniVer 4 Cyanide Shake the sample cell for CyaniVer 5 Cyanide Shake the sample cell
Reagent Powder Pillow. 10 seconds. Immediately do Reagent powder Pillow. vigorously.
the next step. A delay of If cyanide is in the sample, a
more than 30 seconds will pink color will show.
produce low test results.

9. Start the instrument 10. Prepare the blank: 11. Clean the blank. 12. Insert the blank into the
timer. A 30-minute reaction When the timer expires, fill a cell holder.
time starts. second sample cell with
The solution will show pink, 10 mL of sample.
then will show blue.
Samples less than 25 °C
require a longer reaction
time. Samples greater than
25 °C give low test results..

Zero Read

13. Push ZERO. The 14. Clean the prepared 15. Insert the prepared 16. Push READ. Results
display shows 0.000 mg/L sample. sample into the cell holder. show in mg/L CN–.
CN–.

4 Cyanide, Pyridine-Pyrazalone Method (0.240 mg/L)

Interferences
Interfering Interference level
substance
Chlorine Large amounts of chlorine in the sample cause a milky white precipitate after the addition of the CyaniVer®
5 Reagent. If chlorine or other oxidizing agents are known to be present, pretreat the sample before the test
with the procedure in this table for oxidizing agents.
Metals Nickel or cobalt in concentrations up to 1 mg/L do not interfere. Eliminate the interference from up to 20 mg/L
copper and 5 mg/L iron: add the contents of one HexaVer Chelating Reagent Powder Pillow to a fresh portion
of sample and mix. Use this treated sample in the test procedure. Prepare a reagent blank of deionized water
and reagents to zero the instrument.
Oxidizing 1. Adjust a 25-mL portion of the alkaline sample to pH 7–9 with 2.5 N Hydrochloric Acid Standard Solution.
agents Count the number of drops of acid added.
2. Add two drops of Potassium Iodide Solution and two drops of Starch Indicator Solution to the sample.
Swirl to mix. The sample will turn blue if oxidizing agents are present.
3. Add Sodium Arsenite Solution drop-wise until the sample turns colorless. Swirl the sample thoroughly
after each drop. Count the number of drops.
4. Take another 25-mL sample and add the total number of drops of Hydrochloric Acid Standard Solution
counted in step 1.
5. Subtract one drop from the amount of Sodium Arsenite Solution added in step 3. Add this amount to the
sample and mix thoroughly. Use this treated sample in the cyanide test procedure.

Reducing 1. Adjust a 25-mL portion of the alkaline sample to pH 7–9 with 2.5 N Hydrochloric Acid Standard Solution.
agents Count the number of drops added.
2. Add four drops of Potassium Iodide Solution and four drops of Starch Indicator Solution to the sample.
Swirl to mix. The sample should be colorless.
3. Add Bromine Water drop-wise until a blue color shows. Swirl the sample thoroughly after each addition.
Count the number of drops.
4. Take another 25-mL sample and add the total number of drops of Hydrochloric Acid Standard Solution
counted in step 1.
5. Add the total number of drops of Bromine Water counted in step 3 to the sample and mix thoroughly.
6. Use this treated sample in the cyanide test procedure.

Turbidity Large amounts of turbidity will cause high readings. Use filter paper and a funnel to filter highly turbid water
samples. Use the filtered sample for the blank and sample preparation in the test procedure. The test results
should then be recorded as soluble cyanide.

Acid distillation
All samples to be analyzed for cyanide should be treated by acid distillation except when
experience has shown that there is no difference in results obtained with or without
distillation. With most compounds, a 1-hour reflux is adequate.
If thiocyanate is present in the original sample, a distillation step is absolutely necessary
as thiocyanate causes a positive interference. High concentrations of thiocyanate can
yield a substantial quantity of sulfide in the distillate. The “rotten egg” smell of hydrogen
sulfide will accompany the distillate when sulfide is present. The sulfide must be removed
from the distillate prior to testing.
If cyanide is not present, the amount of thiocyanate can be determined. The sample is not
distilled and the final reading is multiplied by 2.2. The result is mg/L SCN–.
The distillate can be tested and treated for sulfide after the last step of the distillation
procedure by using the following lead acetate treatment procedure.
1. Put a drop of the distillate (already diluted to 250 mL) on a disc of Hydrogen Sulfide
Test Paper that has been wetted with pH 4 Buffer Solution.
2. If the test paper darkens, add 2.5 N Hydrochloric Acid Standard Solution by drops to
the distillate until a neutral pH is obtained.
3. Add a 1-g measuring spoon of Lead Acetate to the distillate and mix. Repeat step 1.

Cyanide, Pyridine-Pyrazalone Method (0.240 mg/L) 5

 4. If the test paper continues to turn dark, keep adding lead acetate until the distillate
tests negative for sulfide. Filter the black lead sulfide precipitate through filter paper
and a funnel. Neutralize the liquid filtrate to pH 7 and immediately analyze for
cyanide.
Distillation procedure
The following steps describe the distillation process using distillation apparatus and
cyanide glassware offered by the manufacturer:

1. Set up the distillation apparatus for cyanide recovery, leaving off the thistle tube.
Refer to the Distillation Apparatus Manual. Turn on the water and make certain it is
flowing steadily through the condenser.
2. Fill the distillation apparatus cylinder to the 50-mL mark with 0.25 N Sodium
Hydroxide Standard Solution.
3. Fill a clean 250-mL graduated cylinder to the 250-mL mark with sample and pour it
into the distillation flask. Put a stir bar in the flask and attach the thistle tube.
4. Arrange the vacuum system as shown in the Distillation Apparatus Manual, but do not
connect the vacuum tubing to the gas bubbler. Turn on the water to the aspirator to
full flow and adjust the flow meter to 0.5 SCFH.
5. Connect the vacuum tubing to the gas bubbler, making certain that air flow is
maintained (check the flow meter) and that air is bubbling from the thistle tube and
the gas bubbler.
6. Turn the power switch on and set the stir control to 5. Using a 50-mL graduated
cylinder, pour 50 mL of 19.2 N Sulfuric Acid Standard Solution through the thistle tube
and into the distillation flask.
7. Using a water bottle, rinse the thistle tube with a small amount of deionized water.
8. Allow the solution to mix for 3 minutes, then add 20 mL Magnesium Chloride Reagent
through the thistle tube and rinse again. Allow the solution to mix for 3 more minutes.
9. Make sure that there is a constant flow of water through the condenser.
10. Turn the heat control to 10.
11. Carefully monitor the distillation flask at this point in the procedure. Once the sample
begins to boil, slowly lower the air flow to 0.3 SCFH. If the contents of the distillation
flask begin to back up through the thistle tube, increase the air flow by adjusting the
flow meter until the contents do not back up through the thistle tube. Boil the sample
for 1 hour.
12. After 1 hour, turn off the still, but maintain the air flow for 15 minutes more.
13. After 15 minutes, remove the rubber stopper on the 500-mL vacuum flask to break
the vacuum and turn off the water to the aspirator. Turn off the water to the
condenser.
14. Remove the gas bubbler/cylinder assembly from the distillation apparatus. Separate
the gas bubbler from the cylinder and pour the contents of the cylinder into a 250-mL,
Class A volumetric flask. Rinse the gas bubbler, cylinder and J-tube connector with
deionized water and add the washings to the volumetric flask.
15. Fill the flask to the mark with deionized water and mix thoroughly. Neutralize the
contents of the flask. Use the distilled sample in the test procedure for cyanide.

Pollution prevention and waste management

Reacted samples may contain cyanide and must be disposed of as a hazardous waste. It
is imperative that these materials be handled safely to prevent the release of hydrogen
cyanide gas (an extremely toxic material with the smell of almonds). Most cyanide
compounds are stable and can be safely stored for disposal in highly alkaline solutions
(pH >11) such as 2 N sodium hydroxide. Never mix these wastes with other laboratory
wastes which may contain lower pH materials such as acids or even water. Dispose of
reacted solutions according to local, state and federal regulations.

6 Cyanide, Pyridine-Pyrazalone Method (0.240 mg/L)

Accuracy check
Standard solution method
Use the standard solution method to validate the test procedure, reagents and
instrument.
Items to collect:
• 0.2503 g potassium cyanide
• 1-L volumetric flask, Class A (2)
• 2-mL volumetric pipet, Class A and pipet filler
• Deionized water

1. Prepare a 100-mg/L cyanide stock solution as follows:

a. Add 0.2503 g of potassium cyanide into a 1-L volumetric flask.
b. Dilute to the mark with deionized water. Mix well. Prepare the stock solution each
week.
2. Prepare a 0.200 mg/L cyanide standard solution as follows:
a. Use a pipet to add 2.00 mL of the 100-mg/L cyanide stock solution into a 1-L
volumetric flask.
b. Dilute to the mark with deionized water. Mix well. Prepare the standard solution
immediately before use.
3. Use the test procedure to measure the concentration of the prepared standard
solution.
4. Compare the expected result to the actual result.
Note: The factory calibration can be adjusted slightly with the standard adjust option so that the
instrument shows the expected value of the standard solution. The adjusted calibration is then
used for all test results. This adjustment can increase the test accuracy when there are slight
variations in the reagents or instruments.

Method performance
The method performance data that follows was derived from laboratory tests that were
measured on a spectrophotometer during ideal test conditions. Users may get different
results under different test conditions.
Program Standard Precision (95% Confidence Interval) Sensitivity
Concentration change per 0.010 Abs change
160 0.100 mg/L CN– 0.090–0.110 mg/L CN– 0.002 mg/L CN–

Summary of method
The Pyridine-Pyrazalone method used for cyanide measurements gives an intense blue
color with free cyanide. A sample distillation is necessary to determine cyanide that is
complexed with transition and heavy metals. The measurement wavelength is 612 nm for
spectrophotometers or 610 nm for colorimeters.
Consumables and replacement items
Required reagents

Description Quantity/test Unit Item no.

Cyanide Reagent Set, CyaniVer, 10-mL 1 1 2430200

Includes:
®
CyaniVer 3 Cyanide Reagent Powder Pillow, 10-mL 1 100/pkg 2106869
®
CyaniVer 4 Cyanide Reagent Powder Pillow, 10-mL 1 100/pkg 2106969
®
CyaniVer 5 Cyanide Reagent Powder Pillow, 10-mL 1 100/pkg 2107069

Cyanide, Pyridine-Pyrazalone Method (0.240 mg/L) 7

Required apparatus

Description Quantity/test Unit Item no.

Cylinder, graduated, 10-mL 1 each 50838

Sample cell, 10 mL square, matched pair 2 2/pkg 2495402
Stoppers for 18 mm-tubes and AccuVac Ampuls 1 6/pkg 1448000

Recommended standards

Description Unit Item no.

Potassium Cyanide, ACS 125 g 76714

Water, deionized 4L 27256

Optional reagents and apparatus

Description Unit Item no.

Acetic Acid, ACS 500 mL 10049

Ascorbic Acid 100 g 613826
Bromine Water, 30 g/L 29 mL 221120
Buffer Solution, pH 4 500 mL 1222349
Filter paper, 12.5-cm 100/pkg 189457
Funnel, poly, 65-mm each 108367
Hexane Solution, ACS 500 mL 1447849
HexaVer Chelating Reagent Powder Pillow 100/pkg 24399
100 mL
Hydrochloric Acid Standard Solution, 2.5 N 141832
MDB
Hydrogen Sulfide Test Paper 100/pkg 2537733
m-Nitrophenol Indicator Solution 100 mL 247632
Magnesium Chloride Reagent 1L 1476253
Potassium Iodide, 30 g/L 100 mL 34332
Sodium Arsenite, 5 g/L 100 mL 104732
Sodium Hydroxide Standard Solution, 0.25 N 1000 mL 1476353
Sodium Hydroxide Standard Solution, 5.0 N 1L 245053
100 mL
Starch Indicator Solution 34932
MDB
Sulfuric Acid Standard Solution, 19.2 N 500 mL 203849
Cyanide glassware each 2265800
Distillation heater and support for 2265300, 115 VAC, 60 Hz each 2274400
Distillation heater and support for 2265300, 230 VAC, 50 Hz each 2274402
Distillation apparatus set, general purpose each 2265300
Pipet, serological, 5-mL each 53237
Pipet filler, safety bulb each 1465100
Paper, pH, 0–14 pH range 100/pkg 2601300

8 Cyanide, Pyridine-Pyrazalone Method (0.240 mg/L)

Consumables and replacement items (continued)
Description Unit Item no.

Spoon, measuring, 1 g each 51000

Thermometer, non-mercury, -10 to +225 °C each 2635700

Cyanide, Pyridine-Pyrazalone Method (0.240 mg/L) 9

 FOR TECHNICAL ASSISTANCE, PRICE INFORMATION AND ORDERING: HACH COMPANY
In the U.S.A. – Call toll-free 800-227-4224 WORLD HEADQUARTERS
Outside the U.S.A. – Contact the HACH office or distributor serving you. Telephone: (970) 669-3050
On the Worldwide Web – www.hach.com; E-mail – [email protected] FAX: (970) 669-2932

© Hach Company/Hach Lange GmbH, 1989–2013. All rights reserved. 04/2013, Edition 8
Cyanuric Acid DOC316.53.01183

Turbidimetric Method Method 8139

5 to 50 mg/L (spectrophotometers) Powder Pillows
7 to 55 mg/L (colorimeters)
Scope and application: For water, pools and spas.

Test preparation

Instrument specific information

The table in this section shows all of the instruments that have the program for this test.
Table 1 shows sample cell and orientation requirements for reagent addition tests, such
as powder pillow or bulk reagent tests.
To use the table, select an instrument, then read across to find the corresponding
information for this test.
Table 1 Instrument-specific information for powder pillows
Instrument Sample cell orientation Sample cell
DR 6000 The fill line is to the right. 2495402
DR 3800
DR 2800
DR 2700
DR 5000 The fill line is toward the user.
DR 3900
DR 900 The orientation mark is toward the user. 2401906

Before starting
Install the instrument cap on the DR 900 cell holder before ZERO or READ is pushed.
Clean sample cells with soap, water and a brush soon after each test to prevent a build-up of film on the sample cells.
Filter samples that are turbid with filter paper and a funnel.
Do not use the Pour-Thru Cell or sipper module (for applicable instruments) with this test.
Review the Safety Data Sheets (MSDS/SDS) for the chemicals that are used and use any recommended personal protective
equipment.
Dispose of reacted solutions according to local, state and federal regulations. Use the Safety Data Sheets for disposal
information for unused reagents. Consult the environmental, health and safety staff for your facility and/or local regulatory
agencies for further disposal information.

1
Items to collect
Description Quantity

Bottle, mixing, square glass 1

Cyanuric Acid 2 Reagent Powder Pillow 1
Sample cells (For information about sample cells, adapters or light shields, refer to Instrument
2
specific information on page 1.)

Refer to Consumables and replacement items on page 4 for reorder information.

Sample collection
• Collect samples in clean glass or plastic bottles.
• Samples must be analyzed within 24 hours.

Turbidimetric method

Start

1. Start program 2. Prepare the sample: Fill 3. Add the contents of one 4. Swirl to mix.
170 Cyanuric Acid. For a marked bottle to the 25- Cyanuric Acid 2 Reagent After the reagent is added, a
information about sample mL line with sample. Powder Pillow. white turbidity will show if
cells, adapters or light For instruments that cyanuric acid is in the
shields, refer to Instrument measure with a 25-mL sample. Accuracy is not
specific information sample cell, prepare the affected by undissolved
on page 1. sample in the sample cell. powder.
Note: Although the program
name may vary between
instruments, the program
number does not change.

5. Start the instrument 6. Prepare the blank: Fill a 7. Clean the blank. 8. Insert the blank into the
timer. A 3-minute reaction sample cell with 10 mL of cell holder.
time starts. unreacted sample.

2 Cyanuric Acid, Turbidimetric Method (50 mg/L)

 Zero

9. Push ZERO. The display 10. When the timer expires, 11. Clean the prepared 12. Within 7 minutes of
shows 0 mg/L Cyan Acid. fill a second sample cell with sample. adding the reagent, insert
10 mL of prepared sample the prepared sample into
from the mixing bottle. the cell holder.

Read

13. Push READ. Results 14. Clean sample cells with

show in mg/L Cyan Acid. soap, water and a brush
soon after each test. Cells
that are not cleaned may
form a white film inside the
sample cell.

Interferences
Turbidity interferes. Filter turbid samples before the test is started.
Accuracy check
Standard solution method
Use the standard solution method to validate the test procedure, reagents and
instrument.
Items to collect:
• 1.000 g cyanuric acid
• 1-L volumetric flask, Class A
• 100-mL volumetric flask, Class A
• 3-mL volumetric pipet, Class A and pipet filler
• Deionized water

1. Prepare a 1000-mg/L cyanuric acid stock solution as follows:

a. Add 1.000 g of cyanuric acid into a 1-L volumetric flask. Add several hundred mL
of deionized water and mix well. The cyanuric acid can take several hours to
dissolve.
b. Dilute to the mark with deionized water. Mix well. This solution is stable for
several weeks.
2. Prepare a 30 mg/L cyanuric acid standard solution as follows:
a. Use a pipet to add 3.00 mL of the 1000-mg/L cyanuric acid stock solution into a
100-mL volumetric flask.

Cyanuric Acid, Turbidimetric Method (50 mg/L) 3

 b. Dilute to the mark with deionized water. Mix well. Prepare the standard solution
each day.
3. Use the test procedure to measure the concentration of the prepared standard
solution.
4. Compare the expected result to the actual result.
Note: The factory calibration can be adjusted slightly with the standard adjust option so that the
instrument shows the expected value of the standard solution. The adjusted calibration is then
used for all test results. This adjustment can increase the test accuracy when there are slight
variations in the reagents or instruments.

Method performance
The method performance data that follows was derived from laboratory tests that were
measured on a spectrophotometer during ideal test conditions. Users may get different
results under different test conditions.
Program Standard Precision (95% Confidence Sensitivity
Interval) Concentration change per 0.010 Abs change
170 10 mg/L cyanuric acid 7–13 mg/L cyanuric acid at 10 and 30 mg/L: 0.3 mg/L; at 50 mg/L: 0.4 mg/L
cyanuric acid

Summary of method
The test for cyanuric acid uses the turbidimetric method. Cyanuric Acid 2 Reagent
precipitates any cyanuric acid in the sample and holds it in suspension. The amount of
turbidity caused by the suspended particles is directly proportional to the amount of
cyanuric acid in the sample. The measurement wavelength is 480 nm for
spectrophotometers or 520 nm for colorimeters.
Consumables and replacement items
Required reagents

Description Quantity/test Unit Item no.

Cyanuric Acid 2 Reagent Powder Pillow 1 50/pkg 246066

Required apparatus

Description Quantity/test Unit Item no.

Bottle, square, with 25 mL mark 1 each 1704200

Sample cell, 10 mL square, matched pair 2 2/pkg 2495402

Recommended standards

Description Unit Item no.

Cyanuric Acid 25 g 712924

Water, deionized 4L 27256

Optional reagents and apparatus

Description Unit Item no.

Balance, 600 g x 0.01 g, 100-240 VAC each 2937201

Brush, test tube each 69000
Filter paper, 12.5-cm 100/pkg 189457
Flask, volumetric, 100-mL each 2636642

4 Cyanuric Acid, Turbidimetric Method (50 mg/L)

Consumables and replacement items (continued)
Description Unit Item no.

Flask, volumetric, Class A, 1000-mL each 2636653

Funnel, poly, 65-mm each 108367
Liqui-Nox Phosphate-free detergent 946 mL 2088153
®
Pipet, TenSette , 1.0 to 10.0 mL each 1970010
Pipet tips for TenSette Pipet 1970010 50/pkg 2199796
Pipet, volumetric, Class A, 3-mL each 1451503
Pipet filler, safety bulb each 1465100

Cyanuric Acid, Turbidimetric Method (50 mg/L) 5

 FOR TECHNICAL ASSISTANCE, PRICE INFORMATION AND ORDERING: HACH COMPANY
In the U.S.A. – Call toll-free 800-227-4224 WORLD HEADQUARTERS
Outside the U.S.A. – Contact the HACH office or distributor serving you. Telephone: (970) 669-3050
On the Worldwide Web – www.hach.com; E-mail – [email protected] FAX: (970) 669-2932

© Hach Company/Hach Lange GmbH, 1989–2013. All rights reserved. 04/2013, Edition 8
Fluoride DOC316.53.01041

USEPA SPADNS Method1 Method 8029

®
0.02 to 2.00 mg/L F– Reagent Solution or AccuVac Ampuls
Scope and application: For water, wastewater and seawater; USEPA accepted for reporting for drinking and
wastewater analyses (distillation required).2
1 Adapted from Standard Methods for the Examination of Water and Wastewater, 4500-F B & D.
2 Procedure is equivalent to USEPA method 340.1 for drinking water and wastewater.

Test preparation

Instrument-specific table
The tables in this section show all of the instruments that have the program for this test.
Table 1 shows sample cell and orientation requirements for reagent addition tests, such
as powder pillow or bulk reagent tests. Table 2 shows sample cell and adapter
requirements for AccuVac Ampul tests.
To use either table, select an instrument, then read across to find the corresponding
information for this test.
Table 1 Instrument-specific information for reagent solution
Instrument Sample cell orientation Sample cell
DR 6000 The fill line is to the right. 2495402
DR 3800
DR 2800
DR 2700
DR 5000 The fill line is toward the user.
DR 3900
DR 900 The fill line is toward the user. 2401906

Table 2 Instrument-specific information for AccuVac Ampuls

Instrument Adapter
DR 6000 —
DR 5000
DR 900
DR 3900 LZV846 (A)
DR 3800 LZV584 (C)
DR 2800
DR 2700

Before starting
Install the instrument cap on the DR 900 cell holder before ZERO or READ is pushed.
The sample and deionized water must be at the same temperature (±1 °C). Temperature adjustments can be made before
or after the reagent addition.

1
Make sure that the sample cells are clean and dry before the test.
Measure the volume of the reagent accurately. Use a pipet if possible.
The reagent that is used in this test is corrosive and toxic. Use protection for eyes and skin and be prepared to flush any
spills with running water.
The SPADNS Reagent contains sodium arsenite. The reacted solutions must be disposed of according to local, state and
federal regulations.
Minor variations between lots of reagent become measurable above 1.5 mg/L. While results above 1.5 mg/L are usable for
most purposes, for the best accuracy dilute the sample to a lower concentration.
Do not use the Pour-Thru Cell or sipper module (for applicable instruments) with this test.
Review the Safety Data Sheets (MSDS/SDS) for the chemicals that are used and use any recommended personal protective
equipment.
Dispose of reacted solutions according to local, state and federal regulations. Use the Safety Data Sheets for disposal
information for unused reagents. Consult the environmental, health and safety staff for your facility and/or local regulatory
agencies for further disposal information.

Items to collect
Reagent solution

Description Quantity

SPADNS Reagent Solution 4 mL

Deionized water 10 mL
Pipet, volumetric, 2-mL 1
Pipet, volumetric, 10-mL 1
Pipet filler bulb 1
Sample cells (For information about sample cells, adapters or light shields, refer to Instrument-
2
specific table PPAV.)
Thermometer, –10 to 110 °C 1

Refer to Consumables and replacement items on page 6 for reorder information.

AccuVac Ampuls

Description Quantity
®
SPADNS Fluoride Reagent AccuVac Ampuls 2
Deionized water 40 mL
Beaker, 50-mL 1
Stoppers for 18 mm tubes and AccuVac Ampuls 2

Refer to Consumables and replacement items on page 6 for reorder information.

Sample collection and storage

• Collect samples in clean glass or plastic bottles.
• Samples can be kept for up to 28 days.
• Let the sample temperature increase to room temperature before analysis.

2 Fluoride, SPADNS Method (2.00 mg/L)

SPADNS reagent solution method

Start

1. Start program 2. Prepare the blank: Use 3. Prepare the sample: 4. Use a pipet to add
190 Fluoride. For a pipet to add 10.0 mL of Use a pipet to add 10.0 mL 2.0 mL of SPADNS Reagent
information about sample deionized water to a sample of sample to a sample cell. Solution into each sample
cells, adapters or light cell. cell.
shields, refer to Instrument-
specific table PPAV.
Note: Although the program
name may vary between
instruments, the program
number does not change.

5. Swirl to mix. 6. Start the instrument 7. When the timer expires, 8. Insert the blank into the
timer. A 1-minute reaction clean the blank. cell holder.
time starts.

Zero Read

9. Push ZERO. The display 10. Clean the prepared 11. Insert the prepared 12. Push READ. Results
shows 0.00 mg/L F–. sample. sample into the cell holder. show in mg/L F–.

Fluoride, SPADNS Method (2.00 mg/L) 3

AccuVac Ampul procedure

Start

1. Start program 2. Prepare the blank: Pour 3. Prepare the sample: 4. Quickly invert the Ampuls
195 Fluoride AV. For at least 40 mL of deionized Collect at least 40 mL of several times to mix.
information about sample water in a 50-mL beaker. Fill sample in a 50-mL beaker.
cells, adapters or light one SPADNS Fluoride Fill the second SPADNS
shields, refer to Instrument- Reagent AccuVac Ampul Fluoride Reagent AccuVac
specific table PPAV. with deionized water. Keep Ampul with sample. Keep
Note: Although the program the tip immersed while the the tip immersed while the
name may vary between Ampul fills completely. Ampul fills completely.
instruments, the program
number does not change.

Zero

5. Start the instrument 6. When the timer expires, 7. Insert the blank AccuVac 8. Push ZERO. The display
timer. A 1-minute reaction clean the blank AccuVac Ampul into the cell holder. shows 0.00 mg/L F–.
time starts. Ampul.

Read

9. Clean the AccuVac 10. Insert the prepared 11. Push READ. Results
Ampul. sample AccuVac Ampul into show in mg/L F–.
the cell holder.

4 Fluoride, SPADNS Method (2.00 mg/L)

Interferences
This test is sensitive to small amounts of contamination. Glassware must be very clean
(acid rinse before each use). Repeat the test with the same glassware to make sure that
the results are accurate.
Interfering substance Interference level
Alkalinity (as CaCO3) At 5000 mg/L, it causes a –0.1 mg/L F– error.
Aluminum At 0.1 mg/L, it causes a –0.1 mg/L F– error. To find whether there is an aluminum interference,
read the concentration 1 minute after reagent addition, then again after 15 minutes. An
appreciable increase in concentration suggests aluminum interference. To remove the effect of
up to 3.0 mg/L aluminum, wait 2 hours, then take the final reading.
Chloride At 7000 mg/L, it causes a +0.1 mg/L F– error.
Chlorine SPADNS Reagent contains enough arsenite to remove up to 5 mg/L chlorine. For higher
chlorine levels, add one drop of Sodium Arsenite Solution, 5.0 g/L, to 25 mL of sample to
remove each additional 2 mg/L of Chlorine.
Iron, ferric At 10 mg/L, it causes a –0.1 mg/L F– error.
Phosphate, ortho At 16 mg/L, it causes a +0.1 mg/L F– error.
Sodium hexametaphosphate At 1.0 mg/L, it causes a +0.1 mg/L F– error.
Sulfate At 200 mg/L, it causes a +0.1 mg/L F– error.

Distillation
To eliminate most interferences, distill the sample, then use the distilled sample in the test
procedure.
Prerequisite—prepare the distillation solution:
1. Measure 60 mL of deionized water into a 250-mL, glass Erlenmeyer flask.
2. With constant stirring, add 120 mL of concentrated sulfuric acid. Caution: The
mixture will become very hot. Put the flask in an ice bath to decrease the
temperature of the solution.
Distillation procedure:

1. Set up the distillation apparatus for general purpose distillation. Refer to the
Distillation Apparatus manual for proper assembly. Use a 125-mL Erlenmeyer flask to
collect the distillate.
2. Turn on the water and maintain a steady flow through the condenser.
3. Use a 100-mL graduated cylinder to add 100 mL of sample into the distillation flask.
Add a magnetic stir bar and 5 glass beads.
4. Turn the stirrer power switch on. Turn the stir control to 5.
5. Use a 250-mL graduated cylinder to carefully add 150 mL of distillation solution into
the flask.
Note: For samples with large amounts of chloride, add 5 mg of silver sulfate to the sample for
every mg/L of chloride in the sample.
6. With the thermometer inserted, turn the heat control to 10. The yellow pilot lamp is an
indication that the heater is on.
7. When the temperature is 180 °C (356 °F) or when 100 mL of distillate has been
collected, turn the still off (takes about 1 hour).
8. Dilute the distillate to a volume of 100 mL, if necessary. Use the diluted distillate in
the test procedure.

Pollution prevention and waste management

Reacted samples contain sodium arsenite and must be disposed of as a hazardous
waste. Dispose of reacted solutions according to local, state and federal regulations.

Fluoride, SPADNS Method (2.00 mg/L) 5

Accuracy check
Standard solution method
Use the standard solution method to validate the test procedure, reagents and
instrument.
Items to collect:
• Standard solution within the test range

1. Use the test procedure to measure the concentration of the standard solution.
2. Compare the expected result to the actual result.
Note: The factory calibration can be adjusted slightly with the standard adjust option so that the
instrument shows the expected value of the standard solution. The adjusted calibration is then
used for all test results. This adjustment can increase the test accuracy when there are slight
variations in the reagents or instruments.

Method performance
The method performance data that follows was derived from laboratory tests that were
measured on a spectrophotometer during ideal test conditions. Users may get different
results under different test conditions.
Program Standard Precision (95% Confidence Interval) Sensitivity
Concentration change per 0.010 Abs change
190 1.00 mg/L F– 0.97–1.03 mg/L F– 0.024 mg/L F–
195 1.00 mg/L F– 0.92–1.08 mg/L F– 0.03 mg/L F–

Summary of method
The SPADNS Method for fluoride determination involves the reaction of fluoride with a
red zirconium-dye solution. The fluoride combines with part of the zirconium to form a
colorless complex, thus bleaching the red color in an amount proportional to the fluoride
concentration. This method is accepted by the EPA for NPDES and NPDWR reporting
purposes when the samples have been distilled. Seawater and wastewater samples
require distillation. The measurement wavelength is 580 nm for spectrophotometers or
610 nm for colorimeters.
Consumables and replacement items
Required reagents

Description Quantity/test Unit Item no.

SPADNS Reagent Solution 4 mL 500 mL 44449

OR
®
SPADNS Fluoride Reagent AccuVac Ampul 2 25/pkg 2506025
Water, deionized varies 4L 27256

Required apparatus (solution)

Description Quantity/test Unit Item no.

Pipet filler, safety bulb 1 each 1465100

Pipet, volumetric, Class A, 2.00-mL 1 each 1451536
Pipet, volumetric, Class A, 10.00-mL 1 each 1451538
Sample cell, 10 mL square, matched pair 2 2/pkg 2495402
Thermometer, -10 to 110 °C 1 each 187701

6 Fluoride, SPADNS Method (2.00 mg/L)

Required apparatus (AccuVac)

Description Quantity/test Unit Item no.

Beaker, 50-mL 1 each 50041H

Sample cell, 10 mL round, 25 x 54 mm 1 each 2122800
Sample cell, 10 mL round, 25 x 60 mm 1 6/pkg 2427606

Recommended standards

Description Unit Item no.

Fluoride Standard Solution, 0.2-mg/L F- 500 mL 40502

Fluoride Standard Solution, 0.5-mg/L F- 500 mL 40505
Fluoride Standard Solution, 0.8-mg/L F- 500 mL 40508
Fluoride Standard Solution, 1.0-mg/L F- 1000 mL 29153
Fluoride Standard Solution, 1.0-mg/L F- 500 mL 29149
Fluoride Standard Solution, 1.2-mg/L F- 500 mL 40512
Fluoride Standard Solution, 1.5-mg/L F- 500 mL 40515
Fluoride Standard Solution, 2.0-mg/L F- 500 mL 40520
Fluoride Standard Solution, 100-mg/L F- 500 mL 23249
Drinking Water Standard, Mixed Parameter, Inorganic for F-, NO3, PO4, SO4 500 mL 2833049

Distillation reagents and apparatus

Description Unit Item no.

Cylinder, graduated, 100-mL each 50842

Cylinder, graduated, 250-mL each 50846
Distillation heater and support for 2265300, 115 VAC, 60 Hz each 2274400
OR
Distillation heater and support for 2265300, 230 VAC, 50 Hz each 2274402
AND
Distillation apparatus set, general purpose each 2265300
Flask, Erlenmeyer, 125-mL each 2089743
Glass beads 100/pkg 259600
Stir bar, magnetic each 1076416
Sulfuric Acid, ACS 500 mL 97949

Optional reagents and apparatus

Description Unit Item no.

Silver Sulfate 113 g 33414

Sodium Arsenite, 5 g/L 100 mL 104732
®
AccuVac Snapper each 2405200
Wipes, disposable 280/pkg 2097000

Fluoride, SPADNS Method (2.00 mg/L) 7

 FOR TECHNICAL ASSISTANCE, PRICE INFORMATION AND ORDERING: HACH COMPANY
In the U.S.A. – Call toll-free 800-227-4224 WORLD HEADQUARTERS
Outside the U.S.A. – Contact the HACH office or distributor serving you. Telephone: (970) 669-3050
On the Worldwide Web – www.hach.com; E-mail – [email protected] FAX: (970) 669-2932

© Hach Company/Hach Lange GmbH, 1989–2013. All rights reserved. 04/2013, Edition 8
Fluoride DOC316.53.01184

USEPA SPADNS 2 Method1 Method 10225

®
0.02 to 2.00 mg/L F– Reagent Solution or AccuVac Ampuls
Scope and application: For water, wastewater and seawater; USEPA accepted for reporting for drinking and
wastewater analyses (distillation required).2
1 Adapted from Standard Methods for the Examination of Water and Wastewater, 4500-F B & D.
2 Procedure is equivalent to USEPA Method 340.1 for drinking water and wastewater.

Test preparation

Instrument-specific table
The tables in this section show all of the instruments that have the program for this test.
Table 1 shows sample cell and orientation requirements for reagent addition tests, such
as powder pillow or bulk reagent tests. Table 2 shows sample cell and adapter
requirements for AccuVac Ampul tests.
To use either table, select an instrument, then read across to find the corresponding
information for this test.
Table 1 Instrument-specific information for reagent addition
Instrument Sample cell orientation Sample cell
DR 6000 The fill line is to the right. 2495402
DR 3800
DR 2800
DR 2700
DR 5000 The fill line is toward the user.
DR 3900
DR 900 The fill line is toward the user. 2401906

Table 2 Instrument-specific information for AccuVac Ampuls

Instrument Adapter
DR 6000 —
DR 5000
DR 900
DR 3900 LZV846 (A)
DR 3800 LZV584 (C)
DR 2800
DR 2700

Before starting
Install the instrument cap on the DR 900 cell holder before ZERO or READ is pushed.
The sample and deionized water must be at the same temperature (±1 °C). Temperature adjustments can be made before
or after the reagent addition.

1
Measure the volume of the reagent accurately. Use a pipet if possible.
If the test result is over-range, dilute a fresh sample with a known volume of deionized water and repeat the test. Multiply the
result by the dilution factor.
Minor variations between lots of reagent become measurable above 1.5 mg/L. While results above 1.5 mg/L are usable for
most purposes, for the best accuracy dilute the sample to a lower concentration.
The SPADNS 2 Reagent contains a non-toxic reducing agent to prevent chlorine interference. SPADNS 2 Reagent does not
contain sodium arsenite.
The reagent that is used in this test is corrosive. Use protection for eyes and skin and be prepared to flush any spills with
running water.
Review the Safety Data Sheets (MSDS/SDS) for the chemicals that are used and use any recommended personal protective
equipment.
Dispose of reacted solutions according to local, state and federal regulations. Use the Safety Data Sheets for disposal
information for unused reagents. Consult the environmental, health and safety staff for your facility and/or local regulatory
agencies for further disposal information.

Items to collect
Reagent solution test

Description Quantity

Pipet filler, safety bulb 1

Pipet, volumetric, Class A, 2.00-mL 1
Pipet, volumetric, Class A, 10.00-mL 1
SPADNS 2 Reagent Solution 4 mL
Thermometer 1
Sample cells (For information about sample cells, adapters or light shields, refer to Instrument-
2
specific table PPAV.)
Water, deionized 10 mL

Refer to Consumable and replacement items on page 6 for reorder information.

AccuVac Ampuls

Description Quantity

Beaker, 50-mL 1
®
SPADNS 2 Fluoride Reagent AccuVac Ampul 1
Stoppers, for 18-mm tubes and AccuVac Ampuls 2
Water, deionized 40 mL

Refer to Consumable and replacement items on page 6 for reorder information.

Sample collection and storage

• Collect samples in clean glass or plastic bottles.
• Samples can be kept for up to 28 days.
• Let the sample temperature increase to room temperature before analysis.

2 Fluoride, SPADNS 2 Method (2.00 mg/L)

Reagent solution test

Start

1. Start program 2. Prepare the sample: 3. Prepare the blank: Use 4. Use a pipet to add
190 Fluoride. For Use a pipet to add 10.0 mL a pipet to add 10.0 mL of 2.0 mL of SPADNS
information about sample of sample to a dry sample deionized water to a dry 2 reagent to each cell.
cells, adapters or light cell. sample cell.
shields, refer to Instrument-
specific table PPAV.
Note: Although the program
name may vary between
instruments, the program
number does not change.

5. Swirl to mix. 6. Start the instrument 7. When the timer expires, 8. Insert the blank into the
timer. A 1-minute reaction clean the blank. cell holder.
time starts.

Zero Read

9. Push ZERO. The display 10. Clean the prepared 11. Insert the prepared 12. Push READ. Results
shows 0.00 mg/L F–. sample. sample into the cell holder. show in mg/L F–.

Fluoride, SPADNS 2 Method (2.00 mg/L) 3

AccuVac Ampul test

Start

1. Start program 2. Prepare the sample: 3. Prepare the blank: Pour 4. Quickly invert the Ampuls
195 Fluoride AV. For Collect at least 40 mL of at least 40 mL of deionized several times to mix.
information about sample sample in a 50-mL beaker. water into a 50-mL beaker.
cells, adapters or light Fill the AccuVac Ampul with Fill an AccuVac Ampul with
shields, refer to Instrument- sample. Keep the tip deionized water. Keep the
specific table PPAV. immersed while the Ampul tip immersed while the
Note: Although the program fills completely. Ampul fills completely.
name may vary between
instruments, the program
number does not change.

Zero

5. Start the instrument 6. When the timer expires, 7. Insert the blank AccuVac 8. Push ZERO. The display
timer. A 1-minute reaction clean the blank AccuVac Ampul into the cell holder. shows 0.00 mg/L F–.
time starts. Ampul.

Read

9. Clean the AccuVac 10. Insert the prepared 11. Push READ. Results
Ampul. sample AccuVac Ampul into show in mg/L F–.
the cell holder.

4 Fluoride, SPADNS 2 Method (2.00 mg/L)

Interferences
This test is sensitive to small amounts of contamination. Glassware must be very clean
(acid rinse before each use). Repeat the test with the same glassware to make sure that
the results are accurate.
Interfering substance Interference level
Alkalinity (as CaCO3) At 5000 mg/L, it causes a –0.1 mg/L F– error.
Aluminum At 0.1 mg/L, it causes a –0.1 mg/L F– error. To find whether there is an aluminum interference,
read the concentration 1 minute after reagent addition, then again after 15 minutes. An
appreciable increase in concentration suggests aluminum interference. To remove the effect of
up to 3.0 mg/L aluminum, wait 2 hours, then take the final reading.
Chloride At 7000 mg/L, it causes a +0.1 mg/L F– error.
Chlorine SPADNS 2 Reagent contains enough non-toxic reductant to remove interference of up to
5 mg/L chlorine. For higher chlorine levels:

1. Dilute the sample with deionized water by a factor that will lower the chlorine
concentration to below 5 mg/L.
2. Use the test procedure to measure the fluoride concentration.
3. Multiply the result by the dilution factor to get mg/L fluoride.

Iron, ferric At 10 mg/L, it causes a –0.1 mg/L F– error.

Phosphate, ortho At 16 mg/L, it causes a +0.1 mg/L F– error.
Sodium hexametaphosphate At 1.0 mg/L, it causes a +0.1 mg/L F– error.
Sulfate At 200 mg/L, it causes a +0.1 mg/L F– error.

Distillation
To eliminate most interferences, distill the sample, then use the distilled sample in the test
procedure.
Prerequisite—prepare the distillation solution:
1. Measure 60 mL of deionized water into a 250-mL, glass Erlenmeyer flask.
2. With constant stirring, add 120 mL of concentrated sulfuric acid. Caution: The
mixture will become very hot. Put the flask in an ice bath to decrease the
temperature of the solution.
Distillation procedure:

1. Set up the distillation apparatus for general purpose distillation. Refer to the
Distillation Apparatus manual for proper assembly. Use a 125-mL Erlenmeyer flask to
collect the distillate.
2. Turn on the water and maintain a steady flow through the condenser.
3. Use a 100-mL graduated cylinder to add 100 mL of sample into the distillation flask.
Add a magnetic stir bar and 5 glass beads.
4. Turn the stirrer power switch on. Turn the stir control to 5.
5. Use a 250-mL graduated cylinder to carefully add 150 mL of distillation solution into
the flask.
Note: For samples with large amounts of chloride, add 5 mg of silver sulfate to the sample for
every mg/L of chloride in the sample.
6. With the thermometer inserted, turn the heat control to 10. The yellow pilot lamp is an
indication that the heater is on.
7. When the temperature is 180 °C (356 °F) or when 100 mL of distillate has been
collected, turn the still off (takes about 1 hour).
8. Dilute the distillate to a volume of 100 mL, if necessary. Use the diluted distillate in
the test procedure.

Fluoride, SPADNS 2 Method (2.00 mg/L) 5

Accuracy check
Standard solution method
Use the standard solution method to validate the test procedure, reagents and
instrument.
Items to collect:
• Standard solution within the test range.

1. Use the test procedure to measure the concentration of the standard solution.
2. Compare the expected result to the actual result.
Note: The factory calibration can be adjusted slightly with the standard adjust option so that the
instrument shows the expected value of the standard solution. The adjusted calibration is then
used for all test results. This adjustment can increase the test accuracy when there are slight
variations in the reagents or instruments.

Method performance
The method performance data that follows was derived from laboratory tests that were
measured on a spectrophotometer during ideal test conditions. Users may get different
results under different test conditions.
Program Standard Precision (95% Confidence Interval) Sensitivity
Concentration change per 0.010 Abs change
190 1.00 mg/L F– 0.97–1.03 mg/L F– 0.024 mg/L F– at 1 mg/L
195 1.00 mg/L F– 0.92–1.08 mg/L F– 0.03 mg/L F– at 1 mg/L

Summary of method
The SPADNS 2 Method for fluoride determination involves the reaction of fluoride with a
red zirconium-dye solution. The fluoride combines with part of the zirconium to form a
colorless complex that bleaches the red color in an amount proportional to the fluoride
concentration. This method is equivalent to the EPA method for NPDES and NPDWR
reporting purposes when the samples have been distilled. Seawater and wastewater
samples require distillation. The measurement wavelength is 580 nm for
spectrophotometers or 610 nm for colorimeters.
Consumable and replacement items
Required reagents

Description Quantity/Test Unit Item no.

SPADNS 2 Reagent Solution 4 mL 500 mL 2947549

OR
®
SPADNS 2 Fluoride Reagent AccuVac Ampul 2 25/pkg 2527025
Water, deionized varies 4L 27256

Required apparatus

Description Quantity/Test Unit Item no.

Pipet filler, safety bulb 1 each 1465100

Pipet, volumetric, Class A, 2.00-mL 1 each 1451536
Pipet, volumetric, Class A, 10.00-mL 1 each 1451538
Thermometer 1 each 2635700
Beaker, 50-mL 1 each 50041H
Stoppers for 18-mm tubes and AccuVac Ampuls 2 6/pkg 173106

6 Fluoride, SPADNS 2 Method (2.00 mg/L)

Recommended standards

Description Unit Item no.

Fluoride Standard Solution, 0.2-mg/L F- 500 mL 40502

Fluoride Standard Solution, 0.5-mg/L F- 500 mL 40505
Fluoride Standard Solution, 0.8-mg/L F- 500 mL 40508
Fluoride Standard Solution, 1.0-mg/L F- 1000 mL 29153
Fluoride Standard Solution, 1.0-mg/L F- 500 mL 29149
Fluoride Standard Solution, 1.2-mg/L F- 500 mL 40512
Fluoride Standard Solution, 1.5-mg/L F- 500 mL 40515
Fluoride Standard Solution, 2.0-mg/L F- 500 mL 40520
Fluoride Standard Solution, 100-mg/L F- 500 mL 23249
Drinking Water Standard, Mixed Parameter, Inorganic for F -, NO3, PO4, SO4 500 mL 2833049

Distillation reagents and apparatus

Description Unit Item no.

Cylinder, graduated, 100-mL each 50842

Cylinder, graduated, 250-mL each 50846
Distillation apparatus set, general purpose each 2265300
Distillation heater and support for 2265300, 115 VAC, 60 Hz each 2274400
Distillation heater and support for 2265300, 230 VAC, 50 Hz each 2274402
Flask, Erlenmeyer, 125-mL each 2089743
Flask, Erlenmeyer, 250-mL each 50546
Glass beads 100/pkg 259600
Stir bar, magnetic each 1076416
Sulfuric Acid, ACS 500 mL 97949

Optional reagents and apparatus

Description Unit Item no.

Silver Sulfate 113 g 33414

Balance, analytical, 80 g x 0.1 mg 100-240 VAC each 2936701
Paper, for weighing, 100 x 100 mm 500/pkg 1473885

Fluoride, SPADNS 2 Method (2.00 mg/L) 7

 FOR TECHNICAL ASSISTANCE, PRICE INFORMATION AND ORDERING: HACH COMPANY
In the U.S.A. – Call toll-free 800-227-4224 WORLD HEADQUARTERS
Outside the U.S.A. – Contact the HACH office or distributor serving you. Telephone: (970) 669-3050
On the Worldwide Web – www.hach.com; E-mail – [email protected] FAX: (970) 669-2932

© Hach Company/Hach Lange GmbH, 1989–2013. All rights reserved. 04/2013, Edition 8
Hardness, Calcium and Magnesium DOC316.53.01043

Calmagite Colorimetric Method Method 8030

0.05 to 4.00 mg/L Ca and Mg as CaCO3
Scope and application: For water, wastewater and seawater.

Test preparation

Instrument specific information

The table in this section shows all of the instruments that have the program for this test.
Table 1 shows sample cell and orientation requirements for reagent addition tests, such
as powder pillow or bulk reagent tests.
To use the table, select an instrument, then read across to find the corresponding
information for this test.
Table 1 Instrument-specific information for reagent solution
Instrument Sample cell orientation Sample cell
DR 6000 The fill line is to the right. 2495402
DR 3800
DR 2800
DR 2700
DR 5000 The fill line is toward the user.
DR 3900
DR 900 The orientation mark is toward the user. 2401906

Before starting
Install the instrument cap on the DR 900 cell holder before ZERO or READ is pushed.
For the most accurate magnesium test results, keep the sample temperature between 21–29 °C (70–84 °F).
This test detects any calcium or magnesium contamination in the mixing cylinder, measuring droppers or sample cells. To
test cleanliness, repeat the test until the results are consistent.
Total hardness in mg/L equals mg/L Ca as CaCO3 plus mg/L Mg as CaCO3.
Traces of EDTA or EGTA that remain from previous tests will give incorrect results. Rinse sample cells thoroughly before
each use.
Review the Safety Data Sheets (MSDS/SDS) for the chemicals that are used and use any recommended personal protective
equipment.
Dispose of reacted solutions according to local, state and federal regulations. Use the Safety Data Sheets for disposal
information for unused reagents. Consult the environmental, health and safety staff for your facility and/or local regulatory
agencies for further disposal information.

1
Items to collect
Description Quantity

Alkali Solution, for calcium and magnesium tests 1 mL

Calcium and Magnesium Indicator Solution 1 mL
EDTA Solution, 1 M 1 drop
EGTA Solution 1 drop
Cylinder, graduated mixing, 100-mL 1
Dropper, measuring, 0.5-mL and 1.0-mL 2
Sample cells (For information about sample cells, adapters or light shields, refer to Instrument
3
specific information on page 1.)

Refer to Consumables and replacement items on page 5 for reorder information.

Sample collection and storage

• Collect samples in acid-washed plastic bottles.
• To preserve samples for later analysis, adjust the sample pH to less than 2 with
concentrated nitric acid (about 2 mL per liter). No acid addition is necessary if the
sample is tested immediately.
• Keep preserved samples at room temperature for a maximum of 6 months.
• Before analysis, adjust the pH to 3–8 with 5.0 N sodium hydroxide standard solution.
• Correct the test result for the dilution from the volume additions.

Calmagite procedure

Start

1. Start program 2. Pour 100 mL of sample 3. Use a 1.0 mL dropper to 4. Close the cylinder. Invert
225 Hardness, Mg. For into a 100-mL graduated add 1.0 mL of Calcium and the cylinder several times to
information about sample mixing cylinder. Magnesium Indicator mix.
cells, adapters or light Solution.
shields, refer to Instrument
specific information
on page 1.
Note: Although the program
name may vary between
instruments, the program
number does not change.

2 Hardness, Ca and Mg, Calmagite Method (4.00 mg/L)

5. Use a 1.0 mL dropper to 6. Close the cylinder. Invert 7. Pour 10 mL of the 8. Blank preparation: Add
add 1.0 mL of Alkali Solution the cylinder several times to solution into each of three one drop of 1 M EDTA
for Calcium and Magnesium mix. sample cells. solution to the first sample
Test. cell.

9. Swirl to mix. 10. Magnesium sample: 11. Swirl to mix. 12. Clean the blank.
Add one drop of EGTA
Solution to the second
sample cell.

Zero

13. Insert the blank into the 14. Push ZERO. The 15. Clean the prepared 16. Magnesium sample:
cell holder. display shows 0.00 mg/L Mg sample. Insert the prepared
CaCO3. magnesium sample cell into
the cell holder.

Hardness, Ca and Mg, Calmagite Method (4.00 mg/L) 3

 Read Start Zero

17. Push READ. Results 18. Do not remove the 19. Exit the magnesium 20. Push ZERO. The
show in mg/L magnesium as sample cell from the program. Start program display shows 0.00 mg/L Mg
calcium carbonate. This instrument. Record or select 220 Hardness, Ca. CaCO3.
value is the amount of STORE to save the
magnesium in the sample magnesium results before
expressed as CaCO3. the next step.

Read

21. Calcium sample: Insert 22. Push READ. Results

the third sample cell into the show in mg/L calcium as
cell holder. calcium carbonate. This
value is the amount of
calcium in the sample
expressed as CaCO3.

Interferences
Interfering substance Interference level
Ca >1.0 mg/L; Mg >0.25 mg/L For the most accurate calcium test result, run the test again on a diluted sample if the calcium
is over 1.0 and the magnesium is over 0.25 mg/L as CaCO3. No retesting is needed if either
is below those respective concentrations.
Chromium (Cr 3+) Above 0.25 mg/L
Copper (Cu 2+) Above 0.75 mg/L
EDTA Above 0.2 mg/L as CaCO3
EDTA or EGTA Traces remaining in sample cells from previous tests will give erroneous results. Rinse cells
thoroughly before use.
Iron (Fe 2+) Above 1.4 mg/L
Iron (Fe 3+) Above 2.0 mg/L
Manganese (Mn 2+) Above 0.20 mg/L
Zinc (Zn 2+) Above 0.050 mg/L

Accuracy check
Standard solution method
Use the standard solution method to validate the test procedure, reagents and
instrument.

4 Hardness, Ca and Mg, Calmagite Method (4.00 mg/L)

 Items to collect:
• 2.00 mg/L (as CaCO3) Calcium Standard Solution

1. Use the test procedure to measure the concentration of the standard solution.
2. Compare the expected result to the actual result.
Note: The factory calibration can be adjusted slightly with the standard adjust option so that the
instrument shows the expected value of the standard solution. The adjusted calibration is then
used for all test results. This adjustment can increase the test accuracy when there are slight
variations in the reagents or instruments.

Method performance
The method performance data that follows was derived from laboratory tests that were
measured on a spectrophotometer during ideal test conditions. Users may get different
results under different test conditions.
Program Standard Precision (95% Confidence Interval) Sensitivity
Concentration change per 0.010 Abs change
220 2.00 mg/L Ca 1.90–2.10 mg/L Ca 0.05 mg/L Ca
225 2.00 mg/L Mg 1.92–2.08 mg/L Mg 0.05 mg/L Mg

Summary of method
The colorimetric method for measuring hardness supplements the conventional titrimetric
method because the colorimetric method can measure very low levels of calcium and
magnesium. Also, some metals that interfere in the titrimetric method may not interfere
when the sample is diluted to bring it within the range of this test. The indicator dye is
calmagite, which forms a purplish-blue color in a strongly alkaline solution and changes to
red when it reacts with free calcium or magnesium.
Calcium and magnesium determinations are made by chelation of calcium with EGTA to
remove the red color from calcium and then chelation of calcium and magnesium with
EDTA to remove the red color from both calcium and magnesium. The measurement of
the red color in the different states is used to measure the calcium and magnesium
concentrations. The measurement wavelength is 522 nm for spectrophotometers or
520 nm for colorimeters.
Consumables and replacement items
Required reagents

Description Quantity/test Unit Item no.

Hardness Reagent Set, includes: — 100 tests 2319900

100 mL
Alkali Solution, for calcium and magnesium tests 1 mL 2241732
MDB
100 mL
Calcium and Magnesium Indicator Solution 1 mL 2241832
MDB
50 mL
EDTA Solution, 1 M 2 drops 2241926
SCDB
50 mL
EGTA Solution 1 drop 2229726
SCDB

Required apparatus

Description Quantity/test Unit Item no.

Cylinder, graduated mixing, 100-mL , tall form 1 each 2088642

Dropper, measuring, 0.5 and 1.0 mL plastic 2 20/pkg 2124720

Hardness, Ca and Mg, Calmagite Method (4.00 mg/L) 5

Optional reagents and apparatus

Description Unit Item no.

Nitric Acid, concentrated 500 mL 15249

100 mL
Sodium Hydroxide Standard Solution, 5.0 N 245032
MDB
Paper, pH, 0–14 pH range 100/pkg 2601300

FOR TECHNICAL ASSISTANCE, PRICE INFORMATION AND ORDERING: HACH COMPANY

In the U.S.A. – Call toll-free 800-227-4224 WORLD HEADQUARTERS
Outside the U.S.A. – Contact the HACH office or distributor serving you. Telephone: (970) 669-3050
On the Worldwide Web – www.hach.com; E-mail – [email protected] FAX: (970) 669-2932

© Hach Company/Hach Lange GmbH, 1989–2013. All rights reserved. 04/2013, Edition 8
Hydrazine DOC316.53.01046

p-Dimethylaminobenzaldehyde Method1 Method 8141

®
4 to 600 µg/L N2H4 (spectrophotometers) Reagent Solution or AccuVac Ampuls
10 to 500 µg/L N2H4 (colorimeters)
Scope and application: For boiler water/feedwater.
1 Adapted from ASTM Manual of Industrial Water, D1385-78, 376 (1979).

Test preparation

Instrument-specific table
The tables in this section show all of the instruments that have the program for this test.
Table 1 shows sample cell and orientation requirements for reagent addition tests, such
as powder pillow or bulk reagent tests. Table 2 shows sample cell and adapter
requirements for AccuVac Ampul tests.
To use either table, select an instrument, then read across to find the corresponding
information for this test.
Table 1 Instrument-specific information for reagent solution
Instrument Sample cell orientation Sample cell
DR 6000 The fill line is to the right. 2495402
DR 3800
DR 2800
DR 2700
DR 5000 The fill line is toward the user.
DR 3900
DR 900 The fill line is toward the user. 2401906

Table 2 Instrument-specific information for AccuVac Ampuls

Instrument Adapter
DR 6000 —
DR 5000
DR 900
DR 3900 LZV846 (A)
DR 3800 LZV584 (C)
DR 2800
DR 2700

Before starting
Samples must be analyzed immediately after collection and cannot be preserved for later analysis.
Install the instrument cap on the DR 900 cell holder before ZERO or READ is pushed.
The sample temperature must be between 21 ± 4 °C (70 ± 7 °F) for accurate results.

1
The reagent that is used in this test is corrosive. Use protection for eyes and skin and be prepared to flush any spills with
running water.
Review the Safety Data Sheets (MSDS/SDS) for the chemicals that are used and use any recommended personal protective
equipment.
Dispose of reacted solutions according to local, state and federal regulations. Use the Safety Data Sheets for disposal
information for unused reagents. Consult the environmental, health and safety staff for your facility and/or local regulatory
agencies for further disposal information.

Items to collect
Reagent solution

Description Quantity

HydraVer 2 Reagent Solution 1 mL

Deionized water 10 mL
Graduated cylinder, 25-mL 1
Sample cells (For information about sample cells, adapters or light shields, refer to Instrument-
2
specific table PPAV.)

Refer to Consumables and replacement items on page 6 for reorder information.

AccuVac Ampuls

Description Quantity

HydraVer 2 Reagent AccuVac Ampuls 2

Deionized water 40 mL
Beaker, 50-mL 1
Stoppers for 18 mm tubes and AccuVac Ampuls 2

Refer to Consumables and replacement items on page 6 for reorder information.

Sample collection
• Samples must be analyzed immediately after collection and cannot be preserved for
later analysis.
• Collect samples in clean glass or plastic bottles with tight-fitting caps. Fill the bottle
completely and immediately tighten the cap.
• Prevent agitation of the sample or exposure to air.

2 Hydrazine, p-Dimethylaminobenzaldehyde Method (600 µg/L)

Reagent solution procedure

Start

1. Start program 2. Prepare the blank: Use 3. Prepare the sample: 4. Add 0.5 mL of HydraVer
231 Hydrazine. For a graduated cylinder to pour Use a graduated cylinder to 2 Hydrazine Reagent to
information about sample 10 mL of deionized water pour 10 mL of sample into a each sample cell.
cells, adapters or light into a sample cell. second sample cell. A yellow color shows if
shields, refer to Instrument- hydrazine is present in the
specific table PPAV. sample. The blank may also
Note: Although the program show a light yellow color.
name may vary between
instruments, the program
number does not change.

5. Swirl to mix. 6. Start the instrument 7. Clean the blank. 8. Insert the blank into the
timer. A 12-minute reaction cell holder.
time starts.
Complete the blank zero
steps and insert the
prepared sample during the
reaction period.

Zero Read

9. Push ZERO. The display 10. Clean the prepared 11. Insert the prepared 12. Immediately after the
shows 0 µg/L N2H4. sample. sample into the cell holder. timer expires, push READ.
Results show in µg/L N2H4.

Hydrazine, p-Dimethylaminobenzaldehyde Method (600 µg/L) 3

AccuVac Ampul procedure

Start

1. Start program 2. Prepare the sample: 3. Quickly invert the Ampul 4. Start the instrument
232 Hydrazine AV. For Collect at least 40 mL of several times to mix. timer. A 12-minute reaction
information about sample sample in a 50-mL beaker. time starts.
cells, adapters or light Fill one HydraVer Hydrazine Complete the blank zero
shields, refer to Instrument- AccuVac Ampul with the steps and insert the
specific table PPAV. sample. Keep the tip prepared sample during the
Note: Although the program immersed while the Ampul reaction period.
name may vary between fills completely.
instruments, the program
number does not change.

5. Prepare the blank: Pour 6. Quickly invert the Ampul 7. Clean the blank. 8. Insert the blank into the
at least 40 mL of deionized several times to mix. cell holder.
water into a second 50-mL
beaker. Fill the second
HydraVer Hydrazine
AccuVac Ampul with
deionized water. Keep the
tip immersed while the
Ampul fills completely.

Zero

9. Insert the blank AccuVac 10. Push ZERO. The 11. Clean the AccuVac 12. Insert the prepared
Ampul into the cell holder. display shows 0 µg/L N2H4. Ampul. sample AccuVac Ampul into
the cell holder.

4 Hydrazine, p-Dimethylaminobenzaldehyde Method (600 µg/L)

 Read

13. Immediately after the

timer expires, push READ.
Results show in µg/L N2H4.

Interferences
Interfering substance Interference level
Ammonia No interference up to 10 mg/L. May cause a positive interference of up to 20% at 20 mg/L.
Highly colored or turbid Prepare a 1:1 mixture of deionized water and household bleach. Add one drop of this mixture to
samples 25 mL of sample in a graduated mixing cylinder and invert to mix. Use this solution, instead of
deionized water, to prepare the blank in the test procedure.
Morpholine No interference up to 10 mg/L.

Accuracy check
Standard solution method
Use the standard solution method to validate the test procedure, reagents and
instrument.
Items to collect:
• Hydrazine sulfate, reagent grade
• 1-L volumetric flask, Class A (2)
• 10-mL volumetric pipet, Class A and pipet filler
• Deionized water, oxygen-free

1. Prepare a 25-mg/L hydrazine stock solution as follows:

a. Add 0.1016 g of hydrazine sulfate into a 1-L volumetric flask.
b. Dilute to the mark with oxygen-free deionized water. Mix well. Prepare the stock
solution each day.
2. Prepare a 0.25 mg/L hydrazine standard solution as follows:
a. Use a pipet to add 10.00 mL of the 25-mg/L hydrazine stock solution into a 1-L
volumetric flask.
b. Dilute to the mark with oxygen-free deionized water. Mix well. Prepare the
standard solution immediately before use.
3. Use the test procedure to measure the concentration of the prepared standard
solution.
4. Compare the expected result to the actual result.
Note: The factory calibration can be adjusted slightly with the standard adjust option so that the
instrument shows the expected value of the standard solution. The adjusted calibration is then
used for all test results. This adjustment can increase the test accuracy when there are slight
variations in the reagents or instruments.

Hydrazine, p-Dimethylaminobenzaldehyde Method (600 µg/L) 5

Method performance
The method performance data that follows was derived from laboratory tests that were
measured on a spectrophotometer during ideal test conditions. Users may get different
results under different test conditions.
Program Standard Precision (95% Confidence Interval) Sensitivity
Concentration change per 0.010 Abs change
231 250 µg/L N2H4 247–253 µg/L N2H4 4 µg/L N2H4
232 250 µg/L N2H4 246–254 µg/L N2H4 4 µg/L N2H4

Summary of method
Hydrazine in the sample reacts with the p-dimethylaminobenzaldehyde from the
HydraVer 2 Reagent to form a yellow color which is proportional to the hydrazine
concentration. The measurement wavelength is 455 nm for spectrophotometers or
420 nm for colorimeters.
Consumables and replacement items
Required reagents

Description Quantity/test Unit Item no.

Water, deionized varies 4L 27256

® 100 mL
HydraVer 2 Hydrazine Reagent 1 mL 179032
MDB
OR
®
HydraVer 2 Hydrazine Reagent AccuVac Ampul 2 25/pkg 2524025

Required apparatus for reagent solution

Description Quantity/test Unit Item no.

Cylinder, graduated, 25-mL. 1 each 50840

Required apparatus for AccuVac Ampuls

Description Quantity/test Unit Item no.

Beaker, 50-mL 1 each 50041H

Stoppers for 18-mm tubes and AccuVac Ampuls 2 6/pkg 173106

Recommended standards

Description Unit Item no.

Hydrazine Sulfate, ACS 100 g 74226

Optional reagents and apparatus

Description Unit Item no.

Cylinder, mixing, 25-mL each 189640

Flask, volumetric, Class A, 1000-mL each 1457453
Pipet, volumetric, Class A, 10-mL each 1451538
Pipet filler, safety bulb each 1465100
®
AccuVac Snapper each 2405200

6 Hydrazine, p-Dimethylaminobenzaldehyde Method (600 µg/L)

Hydrazine, p-Dimethylaminobenzaldehyde Method (600 µg/L) 7
 FOR TECHNICAL ASSISTANCE, PRICE INFORMATION AND ORDERING: HACH COMPANY
In the U.S.A. – Call toll-free 800-227-4224 WORLD HEADQUARTERS
Outside the U.S.A. – Contact the HACH office or distributor serving you. Telephone: (970) 669-3050
On the Worldwide Web – www.hach.com; E-mail – [email protected] FAX: (970) 669-2932

© Hach Company/Hach Lange GmbH, 1989–2013. All rights reserved. 04/2013, Edition 8
Iron, Ferrous DOC316.53.01049

1,10-Phenanthroline Method1 Method 8146

®
0.02 to 3.00 mg/L Fe2+ Powder Pillows or AccuVac Ampuls
Scope and application: For water, wastewater and seawater.
1 Adapted from Standard Methods for the Examination of Water and Wastewater, 15th ed. 201 (1980).

Test preparation

Instrument-specific information
The tables in this section show all of the instruments that have the program for this test.
Table 1 shows sample cell and orientation requirements for reagent addition tests, such
as powder pillow or bulk reagent tests. Table 2 shows sample cell and adapter
requirements for AccuVac Ampul tests.
To use either table, select an instrument, then read across to find the corresponding
information for this test.
Table 1 Instrument-specific information for powder pillows
Instrument Sample cell orientation Sample cell
DR 6000 The fill line is to the right. 2495402
DR 3800
DR 2800
DR 2700
DR 5000 The fill line is toward the user.
DR 3900
DR 900 The orientation mark is toward the user. 2401906

Table 2 Instrument-specific information for AccuVac Ampuls

Instrument Adapter Sample cell
DR 6000 — 2427606
DR 5000
DR 900
DR 3900 LZV846 (A)
DR 3800 LZV584 (C) 2122800
DR 2800
DR 2700

Before starting
Samples must be analyzed immediately after collection and cannot be preserved for later analysis.
Install the instrument cap on the DR 900 cell holder before ZERO or READ is pushed.

1
For best results, measure the reagent blank value for each new lot of reagent. Replace the sample with deionized water in
the test procedure to get the reagent blank value. Subtract the reagent blank value from the sample results automatically
with the reagent blank adjust option.
Review the Safety Data Sheets (MSDS/SDS) for the chemicals that are used and use any recommended personal protective
equipment.
Dispose of reacted solutions according to local, state and federal regulations. Use the Safety Data Sheets for disposal
information for unused reagents. Consult the environmental, health and safety staff for your facility and/or local regulatory
agencies for further disposal information.

Items to collect
Powder pillows

Description Quantity

Ferrous Iron Reagent Powder Pillows, 25-mL 1

Sample cells. (For information about sample cells, adapters or light shields, refer to Instrument-
2
specific information on page 1.)

Refer to Consumables and replacement items on page 6 for reorder information.

AccuVac Ampuls

Description Quantity

Ferrous Iron Reagent AccuVac Ampuls 1

Beaker, 50-mL 1
Sample cells (For information about sample cells, adapters or light shields, refer to Instrument-
1
specific information on page 1.)
Stopper for 18-mm tubes and AccuVac Ampuls 1

Refer to Consumables and replacement items on page 6 for reorder information.

Sample collection
• Samples must be analyzed immediately after collection and cannot be preserved for
later analysis.
• Collect samples in clean glass or plastic bottles with tight-fitting caps. Fill the bottle
completely and immediately tighten the cap.
• Prevent agitation of the sample or exposure to air.

2 Iron, Ferrous, 1,10-Phenanthroline Method (3.00 mg/L)

Powder pillow procedure

Start

1. Start program 255 Iron, 2. Prepare the blank: Fill 3. Prepare the sample: Fill 4. Add the contents of one
Ferrous. For information the sample cell with 10 mL a mixing cylinder to the 25- Ferrous Iron Reagent
about sample cells, of sample. mL line with sample. Powder Pillow to the mixing
adapters or light shields, cylinder.
refer to Instrument-specific An orange color shows if
information on page 1. ferrous iron is present in the
Note: Although the program sample
name may vary between
instruments, the program
number does not change.

5. Close the cylinder. Invert 6. Start the instrument 7. When the timer expires, 8. Insert the blank into the
the cylinder several times to timer. A 3-minute reaction clean the blank. cell holder.
mix. time starts.
Undissolved powder does
not affect accuracy.

Zero

9. Push ZERO. The display 10. Fill a second sample 11. Clean the prepared 12. Insert the prepared
shows 0.00 mg/L Fe2+. cell with 10 mL of the sample. sample into the cell holder.
reacted prepared sample.

Iron, Ferrous, 1,10-Phenanthroline Method (3.00 mg/L) 3

 Read

13. Push READ. Results

show in mg/L Fe2+.

AccuVac Ampul procedure

Start

1. Start program 257 Iron, 2. Prepare the blank: Fill 3. Prepare the sample: 4. Quickly invert the Ampul
Ferrous AV. For information the sample cell with 10 mL Collect at least 40 mL of several times to mix.
about sample cells, of sample. sample in a 50-mL beaker.
adapters or light shields, Fill the AccuVac Ampul with
refer to Instrument-specific sample. Keep the tip
information on page 1. immersed while the Ampul
Note: Although the program fills completely.
name may vary between
instruments, the program
number does not change.

Zero

5. Start the instrument 6. When the timer expires, 7. Insert the blank into the 8. Push ZERO. The display
timer. A 3-minute reaction clean the blank. cell holder. shows 0.00 mg/L Fe2+.
time starts.

4 Iron, Ferrous, 1,10-Phenanthroline Method (3.00 mg/L)

 Read

9. Clean the AccuVac 10. Insert the prepared 11. Push READ. Results
Ampul. sample AccuVac Ampul into show in mg/L Fe2+.
the cell holder.

Accuracy check
Standard solution method
Use the standard solution method to validate the test procedure, reagents and
instrument.
Items to collect:
• Ferrous Ammonium Sulfate, hexahydrate
• 1-L volumetric flask, Class A
• 100-mL volumetric flask, Class A
• 2-mL volumetric pipet, Class A and pipet filler
• Deionized water

1. Prepare a 100-mg/L Fe2+ ferrous iron stock solution as follows:

a. Add 0.7022 g of ferrous ammonium sulfate, hexahydrate into a 1-L volumetric
flask.
b. Dilute to the mark with deionized water. Mix well.
2. Prepare a 2 mg/L ferrous iron standard solution as follows:
a. Use a pipet to add 2.00 mL of the 100-mg/L Fe2+ ferrous iron stock solution into a
100-mL volumetric flask.
b. Dilute to the mark with deionized water. Mix well. Prepare the standard solution
immediately before use.
3. Use the test procedure to measure the concentration of the prepared standard
solution.
4. Compare the expected result to the actual result.
Note: The factory calibration can be adjusted slightly with the standard adjust option so that the
instrument shows the expected value of the standard solution. The adjusted calibration is then
used for all test results. This adjustment can increase the test accuracy when there are slight
variations in the reagents or instruments.

Method performance
The method performance data that follows was derived from laboratory tests that were
measured on a spectrophotometer during ideal test conditions. Users may get different
results under different test conditions.
Program Standard Precision (95% Confidence Interval) Sensitivity
Concentration change per 0.010 Abs change
255 2.00 mg/L Fe2+ 1.99–2.01 mg/L Fe2+ 0.021 mg/L Fe2+
257 2.00 mg/L Fe2+ 1.98–2.02 mg/L Fe2+ 0.023 mg/L Fe2+

Iron, Ferrous, 1,10-Phenanthroline Method (3.00 mg/L) 5

Summary of method
The 1,10-phenanthroline indicator in the Ferrous Iron Reagent reacts with ferrous iron
(Fe2+) in the sample to form an orange color in proportion to the iron concentration. Ferric
iron (Fe3+) does not react. The ferric iron concentration can be determined by subtracting
the ferrous iron concentration from the results of a total iron test. The measurement
wavelength is 510 nm for spectrophotometers or 520 nm for colorimeters.
Consumables and replacement items
Required reagents

Description Quantity/test Unit Item no.

Ferrous Iron Reagent Powder Pillow, 25-mL 1 100/pkg 103769

OR
®
Ferrous Iron Reagent AccuVac Ampul 1 25/pkg 2514025

Required apparatus

Description Quantity/test Unit Item no.

Beaker, 50-mL 1 each 50041H

Stoppers for 18 mm-tubes and AccuVac Ampuls 1 6/pkg 1448000

Recommended standards and apparatus

Description Unit Item no.

Balance, analytical, 80 g x 0.1 mg 100-240 VAC each 2936701

Ferrous Ammonium Sulfate, hexahydrate, ACS 113 g 1125614
Flask, volumetric, Class A, 1000-mL each 1457453
Pipet filler, safety bulb each 1465100
Pipet, volumetric, Class A, 1.00-mL each 1451535
Water, deionized 4L 27256
Wipes, disposable 280/pkg 2097000

FOR TECHNICAL ASSISTANCE, PRICE INFORMATION AND ORDERING: HACH COMPANY

In the U.S.A. – Call toll-free 800-227-4224 WORLD HEADQUARTERS
Outside the U.S.A. – Contact the HACH office or distributor serving you. Telephone: (970) 669-3050
On the Worldwide Web – www.hach.com; E-mail – [email protected] FAX: (970) 669-2932

© Hach Company/Hach Lange GmbH, 1989–2013. All rights reserved. 04/2013, Edition 8
Iron DOC316.53.01048

FerroZine® Method1 Method 8147

0.009 to 1.400 mg/L Fe Reagent Solution Pillows
Scope and application: For water and seawater.
1 Adapted from Stookey, L.L., Anal. Chem., 42(7), 779 (1970).

Test preparation

Instrument specific information

The table in this section shows all of the instruments that have the program for this test.
Table 1 shows sample cell and orientation requirements for reagent addition tests, such
as powder pillow or bulk reagent tests.
To use the table, select an instrument, then read across to find the corresponding
information for this test.
Table 1 Instrument-specific information for powder pillows
Instrument Sample cell orientation Sample cell
DR 6000 The fill line is to the right. 2495402
DR 3800
DR 2800
DR 2700
DR 5000 The fill line is toward the user.
DR 3900
DR 900 The orientation mark is toward the user. 2401906

Before starting
Install the instrument cap on the DR 900 cell holder before ZERO or READ is pushed.
The sample must be digested with heat and acid to make sure that all forms of the metal are measured. Use the mild or
vigorous digestion. Refer to the Water Analysis Guide for more information.
Clean all glassware with 6.0 N (50%) hydrochloric acid, then rinse thoroughly with deionized water to remove contaminants.
For best results, measure the reagent blank value for each new lot of reagent. Replace the sample with deionized water in
the test procedure to get the reagent blank value. Subtract the reagent blank value from the sample results automatically
with the reagent blank adjust option.
If the sample contains rust, refer to Interferences on page 3.
Use clean clippers to open the solution pillows that are free of rust. Wipe with a dry towel. Do not allow the clippers to
contact the contents of the pillow.
As an alternative to the solution pillows, use 0.5 mL of FerroZine® Iron Reagent Solution in the test procedure.
The FerroZine Iron Reagent can crystallize or precipitate if kept at cold temperatures during shipment. The reagent quality is
not affected. Put the reagent in warm water to dissolve the precipitate.

1
 Review the Safety Data Sheets (MSDS/SDS) for the chemicals that are used and use any recommended personal protective
equipment.
Dispose of reacted solutions according to local, state and federal regulations. Use the Safety Data Sheets for disposal
information for unused reagents. Consult the environmental, health and safety staff for your facility and/or local regulatory
agencies for further disposal information.

Items to collect
Description Quantity

FerroZine Iron Reagent Solution Pillows, OR 1

FerroZine Iron Reagent Solution 0.5 mL
Cylinder, 25-mL graduated mixing, with stopper 1
Clippers for solution pillows 1
Sample cells (For information about sample cells, adapters or light shields, refer to Instrument
2
specific information on page 1.)

Refer to Consumables and replacement items on page 5 for reorder information.

Sample collection and storage

• Collect samples in clean glass or plastic bottles.
• To preserve samples for later analysis, adjust the sample pH to less than 2 with
concentrated nitric acid (about 2 mL per liter). No acid addition is necessary if the
sample is tested immediately.
• To measure only dissolved iron, filter the sample immediately after collection and
before acidification.
• Keep preserved samples at room temperature for a maximum of 6 months.
• Before analysis, adjust the pH to 3–5 with 10% ammonium hydroxide solution. Do not
exceed pH 5 to prevent precipitation of the iron.
• Correct the test result for the dilution from the volume additions.

Solution pillow or bulk solution procedure

Start

1. Start program 260 Iron, 2. Prepare the blank: Fill 3. Prepare the sample: Fill 4. Add the contents of one
FerroZine. For information the sample cell with 10 mL a mixing cylinder to the 25- FerroZine Iron Reagent
about sample cells, of sample. mL line with sample. Solution Pillow to the mixing
adapters or light shields, cylinder.
refer to Instrument specific
information on page 1.
Note: Although the program
name may vary between
instruments, the program
number does not change.

2 Iron, FerroZine Method (1.400 mg/L)

5. Close the cylinder. Invert 6. Start the instrument 7. When the timer expires, 8. Clean the blank.
the cylinder several times to timer. A 5-minute reaction pour 10 mL of the prepared
mix. time starts. sample from the mixing
A purple color shows if iron cylinder into the second
is present in the sample. sample cell.

Zero

9. Insert the blank into the 10. Push ZERO. The 11. Clean the prepared 12. Insert the prepared
cell holder. display shows 0.000 mg/L sample. sample into the cell holder.
Fe.

Read

13. Push READ. Results

show in mg/L Fe.

Interferences
Interfering Interference level
substance
Strong chelants Interfere at all levels. Use the FerroVer® or TPTZ methods for these samples. Use the TPTZ method
(EDTA) for low iron concentrations.
Cobalt May give slightly high results
Copper May give slightly high results
Hydroxides Add the FerroZine® Iron Reagent to 25 mL of sample, then boil the sample for 1 minute in a boiling
water bath. Let the sample cool to 24 °C (75 °F), then start the instrument timer. Return the sample
volume to 25 mL with deionized water.

Iron, FerroZine Method (1.400 mg/L) 3

Interfering Interference level
substance
Magnetite (black Add the FerroZine® Iron Reagent to 25 mL of sample, then gently boil the sample for 20 to 30 minutes
iron oxide) or in a boiling water bath.
Ferrites Note: Do not let the sample boil dry. A purple color will show if iron is present.
Let the sample cool to 24 °C (75 °F). Return the sample volume to 25 mL with deionized water.
Continue with the test procedure after the timer step.
Rust Add the FerroZine® Iron Reagent to 25 mL of sample, then boil the sample for 1 minute in a boiling
water bath. Cool to 24 °C (75 °F), then start the instrument timer. Return the sample volume to 25 mL
with deionized water.

Accuracy check
Standard additions method (sample spike)
Use the standard additions method (for applicable instruments) to validate the test
procedure, reagents and instrument and to find if there is an interference in the sample.
Items to collect:
• Iron Voluette® Ampule Standard, 10 mg/L Fe
• Ampule breaker
• Pipet, TenSette®, 0.1–1.0 mL and tips

1. Use the test procedure to measure the concentration of the sample, then keep the
(unspiked) sample in the instrument.
2. Go to the Standard Additions option in the instrument menu.
3. Select the values for standard concentration, sample volume and spike volumes.
4. Open the standard solution.
5. Prepare three spiked samples: use the TenSette pipet to add 0.1 mL, 0.2 mL and
0.3 mL of the standard solution, respectively, to three 25-mL portions of fresh sample.
Mix well.
6. Use the test procedure to measure the concentration of each of the spiked samples.
Start with the smallest sample spike. Measure each of the spiked samples in the
instrument.
7. Select Graph to compare the expected results to the actual results.
Note: If the actual results are significantly different from the expected results, make sure that
the sample volumes and sample spikes are measured accurately. The sample volumes and
sample spikes that are used should agree with the selections in the standard additions menu. If
the results are not within acceptable limits, the sample may contain an interference.

Standard solution method

Use the standard solution method to validate the test procedure, reagents and
instrument.
Items to collect:
• Iron Standard Solution, 100-mg/L
• 500-mL volumetric flask, Class A
• 5-mL volumetric pipet, Class A and pipet filler
• Deionized water

1. Prepare a 1.0 mg/L iron standard solution as follows:

a. Use a pipet to add 5.00 mL of 100 mg/L iron standard solution into the volumetric
flask.
b. Dilute to the mark with deionized water. Mix well. Prepare this solution daily.
2. Use the test procedure to measure the concentration of the prepared standard
solution.
3. Compare the expected result to the actual result.

4 Iron, FerroZine Method (1.400 mg/L)

 Note: The factory calibration can be adjusted slightly with the standard adjust option so that the
instrument shows the expected value of the standard solution. The adjusted calibration is then
used for all test results. This adjustment can increase the test accuracy when there are slight
variations in the reagents or instruments.

Method performance
The method performance data that follows was derived from laboratory tests that were
measured on a spectrophotometer during ideal test conditions. Users may get different
results under different test conditions.
Program Standard Precision (95% Confidence Interval) Sensitivity
Concentration change per 0.010 Abs change
260 1.000 mg/L Fe 0.985–1.015 mg/L Fe 0.009 mg/L Fe

Summary of method
The FerroZine® Iron Reagent forms a purple-colored complex with trace amounts of iron
in samples that are buffered to a pH of 3.5. This method is applicable for determining
trace levels of iron in chemical reagents and glycols and with digestion can be used to
analyze samples containing magnetite (black iron oxide) or ferrites. The measurement
wavelength is 562 nm for spectrophotometers or 560 nm for colorimeters.
Consumables and replacement items
Required reagents

Description Quantity/test Unit Item no.

®
FerroZine Iron Reagent Solution 0.5 mL 500 mL 230149
OR
®
FerroZine Iron Reagent Solution Pillows 1 50/pkg 230166

Required apparatus

Description Quantity/test Unit Item no.

Clippers for solution pillows 1 each 96800

Cylinder, graduated mixing, 25 mL with stopper 1 each 2088640
Sample cell, 10 mL square, matched pair 2 2/pkg 2495402

Recommended standards and apparatus

Description Unit Item no.

Iron Standard Solution, 100 mg/L Fe 100 mL 1417542

®
Iron Standard Solution, 10 mL Voluette ampule, 25 mg/L Fe 16/pkg 1425310
Metals Drinking Water Standard, LR for Cu, Fe, Mn 500 mL 2833749
Flask, volumetric, Class A, 500-mL each 1457449
®
Pipet, TenSette , 0.1–1.0 mL each 1970001
Pipet tips for TenSette Pipet 1970001 50/pkg 2185696
Pipet tips for TenSette Pipet 1970001 1000/pkg 2185628
Pipet, volumetric 5.00-mL each 1451537
Pipet filler, safety bulb each 1465100

Iron, FerroZine Method (1.400 mg/L) 5

Optional reagents and apparatus

Description Unit Item no.

100 mL
Ammonium Hydroxide, 10% 1473632
MDB
Hydrochloric Acid, 1:1, 6N 500 mL 88449
Nitric Acid, concentrated 500 mL 15249

FOR TECHNICAL ASSISTANCE, PRICE INFORMATION AND ORDERING: HACH COMPANY

In the U.S.A. – Call toll-free 800-227-4224 WORLD HEADQUARTERS
Outside the U.S.A. – Contact the HACH office or distributor serving you. Telephone: (970) 669-3050
On the Worldwide Web – www.hach.com; E-mail – [email protected] FAX: (970) 669-2932

© Hach Company/Hach Lange GmbH, 1989–2013. All rights reserved. 04/2013, Edition 8
Iron, Total DOC316.53.01052

FerroMo Method1 Method 8365

0.01 to 1.80 mg/L Fe Powder Pillows
Scope and application: For cooling water that contains molybdate-based treatment.
1 Adapted from G. Frederick Smith Chemical Co., The Iron Reagents, 3rd ed. (1980).

Test preparation

Instrument specific information

The table in this section shows all of the instruments that have the program for this test.
Table 1 shows sample cell and orientation requirements for reagent addition tests, such
as powder pillow or bulk reagent tests.
To use the table, select an instrument, then read across to find the corresponding
information for this test.
Table 1 Instrument-specific information for powder pillows
Instrument Sample cell orientation Sample cell
DR 6000 The fill line is to the right. 2495402
DR 3800
DR 2800
DR 2700
DR 5000 The fill line is toward the user.
DR 3900
DR 900 The orientation mark is toward the user. 2401906

Before starting
Install the instrument cap on the DR 900 cell holder before ZERO or READ is pushed.
The sample must be digested with heat and acid to make sure that all forms of the metal are measured. Use the mild or
vigorous digestion. Refer to the Water Analysis Guide for more information.
Wash all glassware with detergent. Rinse with tap water. Rinse again with 1:1 hydrochloric acid solution. Rinse a third time
with high-quality deionized water. These steps will remove deposits that can cause slightly high results.
If the sample contains 100 mg/L or more molybdate (MoO42–), read the sample immediately after the instrument zero.
For best results, measure the reagent blank value for each new lot of reagent. Replace the sample with deionized water in
the test procedure to get the reagent blank value. Subtract the reagent blank value from the sample results automatically
with the reagent blank adjust option.
Review the Safety Data Sheets (MSDS/SDS) for the chemicals that are used and use any recommended personal protective
equipment.
Dispose of reacted solutions according to local, state and federal regulations. Use the Safety Data Sheets for disposal
information for unused reagents. Consult the environmental, health and safety staff for your facility and/or local regulatory
agencies for further disposal information.

1
Items to collect
Description Quantity
®
FerroMo Reagent 1 Powder Pillow 1
®
FerroMo Reagent 2 Powder Pillow 1
Cylinder, graduated mixing, 25-mL with stopper 1
Cylinder, graduated mixing, 50-mL with stopper 1
Sample cells (For information about sample cells, adapters or light shields, refer to Instrument
2
specific information on page 1.)

Refer to Consumables and replacement items on page 5 for reorder information.

Sample collection and storage

• Collect samples in clean acid-washed glass or plastic bottles.
• To preserve samples for later analysis, adjust the sample pH to less than 2 with
concentrated hydrochloric acid (about 2 mL per liter). No acid addition is necessary if
the sample is tested immediately.
• To measure only dissolved iron, filter the sample through a 0.45 micron filter or
equivalent medium immediately after collection and before acidification.
• Keep preserved samples at room temperature for a maximum of 6 months.
• Before analysis, adjust the pH to 3–5 with 5.0 N sodium hydroxide standard solution.
Do not exceed pH 5 to prevent precipitation of the iron.
• Correct the test result for the dilution from the volume additions.

Powder pillow procedure

Start

1. Start program 275 Iron, 2. Prepare the sample: Fill 3. Add the contents of one 4. Close the cylinder. Invert
FerroMo. For information a 50-mL mixing cylinder with FerroMo Iron Reagent several times to mix.
about sample cells, 50 mL of sample. 1 Powder Pillow to the
adapters or light shields, mixing cylinder.
refer to Instrument specific
information on page 1.
Note: Although the program
name may vary between
instruments, the program
number does not change.

2 Iron, Total, FerroMo Method (1.80 mg/L)

5. Fill a clean 25-mL mixing 6. Prepare the blank: Fill a 7. Develop the sample: 8. Close the cylinder. Invert
cylinder to the 25-mL mark second sample cell with Add the contents of one the cylinder several times to
with the prepared sample. 10 mL of the prepared FerroMo Iron Reagent mix.
Use the rest of the sample sample. 2 Powder Pillow to the A blue color will show if iron
to prepare the blank. prepared sample in the 25- is present in the sample. A
mL mixing cylinder. small amount of undissolved
reagent will not affect the
results of the test.

9. Start the instrument 10. When the timer expires, 11. Clean the blank. 12. Insert the blank into the
timer. A 3-minute reaction pour 10 mL of the cell holder.
time starts. developed sample into a
sample cell. This is the
prepared sample for the
test.

Zero Read

13. Push ZERO. The 14. Clean the developed 15. Insert the developed 16. Push READ. Results
display shows 0.00 mg/L Fe. sample. sample into the cell holder. show in mg/L Fe.

Iron, Total, FerroMo Method (1.80 mg/L) 3

Interferences
Interfering Interference level
substance
pH After the addition of reagent, a sample pH of less than 3 or more than 4 may inhibit color formation, cause
the developed color to fade quickly or result in turbidity. Adjust the sample pH to between 3 and 8 in the
graduated cylinder before the addition of reagent:

1. Add by drops an applicable amount of iron-free acid or base such as 1.0 N Sulfuric Acid Standard
Solution or 1.0 N Sodium Hydroxide Standard Solution.
2. Make a volume correction if significant volumes of acid or base are used.

Accuracy check
Standard additions method (sample spike)
Use the standard additions method (for applicable instruments) to validate the test
procedure, reagents and instrument and to find if there is an interference in the sample.
Items to collect:
• Iron Voluette® Ampule Standard, 50 mg/L Fe
• Ampule breaker
• Pipet, TenSette®, 0.1–1.0 mL and tips

1. Use the test procedure to measure the concentration of the sample, then keep the
(unspiked) sample in the instrument.
2. Go to the Standard Additions option in the instrument menu.
3. Select the values for standard concentration, sample volume and spike volumes.
4. Open the standard solution.
5. Prepare three spiked samples: use the TenSette pipet to add 0.1 mL, 0.2 mL and
0.3 mL of the standard solution, respectively, to three 50-mL portions of fresh sample.
Mix well.
6. Use the test procedure to measure the concentration of each of the spiked samples.
Start with the smallest sample spike. Measure each of the spiked samples in the
instrument.
7. Select Graph to compare the expected results to the actual results.
Note: If the actual results are significantly different from the expected results, make sure that
the sample volumes and sample spikes are measured accurately. The sample volumes and
sample spikes that are used should agree with the selections in the standard additions menu. If
the results are not within acceptable limits, the sample may contain an interference.

Standard solution method

Use the standard solution method to validate the test procedure, reagents and
instrument.
Items to collect:
• 100 mg/L iron standard solution
• 100-mL volumetric flask, Class A
• 1-mL volumetric pipet, Class A and pipet filler
• Deionized water

1. Prepare a 1.00 mg/L iron standard solution as follows:

a. Use a pipet to add 1.0 mL of 100 mg/L iron standard solution into the volumetric
flask.
b. Dilute to the mark with deionized water. Mix well. Prepare this solution daily.
2. Use the test procedure to measure the concentration of the prepared standard
solution.
3. Compare the expected result to the actual result.

4 Iron, Total, FerroMo Method (1.80 mg/L)

 Note: The factory calibration can be adjusted slightly with the standard adjust option so that the
instrument shows the expected value of the standard solution. The adjusted calibration is then
used for all test results. This adjustment can increase the test accuracy when there are slight
variations in the reagents or instruments.

Method performance
The method performance data that follows was derived from laboratory tests that were
measured on a spectrophotometer during ideal test conditions. Users may get different
results under different test conditions.
Program Standard Precision (95% Confidence Interval) Sensitivity
Concentration change per 0.010 Abs change
275 1.00 mg/L Fe 0.98–1.02 mg/L Fe 0.01 mg/L Fe

Summary of method
FerroMo Iron Reagent 1 contains a reducing agent combined with a masking agent. The
masking agent removes interference from high levels of molybdate. The reducing agent
converts precipitated or suspended iron, such as rust, to the ferrous state. FerroMo Iron
Reagent 2 contains the indicator combined with a buffering agent. The indicator reacts
with ferrous iron in the sample, buffered between pH 3 and 5, which results in a deep
blue-purple color. The measurement wavelength is 590 nm for spectrophotometers or
610 nm for colorimeters.
Consumables and replacement items
Required reagents

Description Quantity/test Unit Item no.

®
FerroMo Iron Reagent Set 1 100/pkg 2544800
Includes:
®
FerroMo Reagent 1 Powder Pillow 1 25/pkg 2543768
®
FerroMo Reagent 2 Powder Pillow 1 50/pkg 2543866

Required apparatus

Description Quantity/test Unit Item no.

Cylinder, graduated mixing, 25 mL with stopper 1 each 2088640

Cylinder, graduated mixing, 50 mL, with stopper 1 each 2088641

Recommended standards

Description Unit Item no.

Iron Standard Solution, 100 mg/L Fe 100 mL 1417542

Iron Standard Solution, 1 mg/L Fe 500 mL 13949
®
Iron Standard Solution, 10 mL Voluette ampule, 50 mg/L Fe. 16/pkg 1425410
Water, deionized 4L 27256

Optional reagents and apparatus

Description Unit Item no.

Flask, volumetric, Class A, 100-mL each 1457442

Pipet, volumetric, Class A, 1.00-mL each 1451535
Pipet filler, safety bulb each 1465100

Iron, Total, FerroMo Method (1.80 mg/L) 5

Consumables and replacement items (continued)
Description Unit Item no.

100 mL
Sodium Hydroxide Standard Solution, 1.0 N 104532
MDB
100 mL
Sodium Hydroxide Standard Solution, 5.0 N 245032
MDB
100 mL
Sulfuric Acid Standard Solution, 1 N 127032
MDB
Hydrochloric Acid, 6.0 N 1:1, 50% 500 mL 88449

FOR TECHNICAL ASSISTANCE, PRICE INFORMATION AND ORDERING: HACH COMPANY

In the U.S.A. – Call toll-free 800-227-4224 WORLD HEADQUARTERS
Outside the U.S.A. – Contact the HACH office or distributor serving you. Telephone: (970) 669-3050
On the Worldwide Web – www.hach.com; E-mail – [email protected] FAX: (970) 669-2932

© Hach Company/Hach Lange GmbH, 1989–2013. All rights reserved. 04/2013, Edition 8
Iron, Total DOC316.53.01053

®
USEPA1 FerroVer Method2 Method 8008
®
0.02 to 3.00 mg/L Fe Powder Pillows or AccuVac Ampuls
Scope and application: For water, wastewater and seawater; digestion is required for determining total iron.
1 USEPA approved for reporting wastewater analysis, Federal Register, June 27, 1980; 45 (126:43459).
2 Adapted from Standard Methods for the Examination of Water and Wastewater.

Test preparation

Instrument-specific information
The tables in this section show all of the instruments that have the program for this test.
Table 1 shows sample cell and orientation requirements for reagent addition tests, such
as powder pillow or bulk reagent tests. Table 2 shows sample cell and adapter
requirements for AccuVac Ampul tests.
To use either table, select an instrument, then read across to find the corresponding
information for this test.
Table 1 Instrument-specific information for powder pillows
Instrument Sample cell orientation Sample cell
DR 6000 The fill line is to the right. 2495402
DR 3800
DR 2800
DR 2700
DR 5000 The fill line is toward the user.
DR 3900
DR 900 The orientation mark is toward the user. 2401906

Table 2 Instrument-specific information for AccuVac Ampuls

Instrument Adapter Sample cell
DR 6000 — 2427606
DR 5000
DR 900
DR 3900 LZV846 (A)
DR 3800 LZV584 (C) 2122800
DR 2800
DR 2700

1
Before starting
Install the instrument cap on the DR 900 cell holder before ZERO or READ is pushed.
The reagent in this test procedure converts all soluble iron and most insoluble forms of iron in the sample to soluble ferrous
iron for measurement. For regulatory reporting, however, the sample must be digested with heat and acid to make sure that
all forms of the metal are measured.
For best results, measure the reagent blank value for each new lot of reagent. Replace the sample with deionized water in
the test procedure to get the reagent blank value. Subtract the reagent blank value from the sample results automatically
with the reagent blank adjust option.
For turbid samples, treat the blank with one 0.1-g scoop of RoVer Rust Remover. Swirl to dissolve.
Review the Safety Data Sheets (MSDS/SDS) for the chemicals that are used and use any recommended personal protective
equipment.
Dispose of reacted solutions according to local, state and federal regulations. Use the Safety Data Sheets for disposal
information for unused reagents. Consult the environmental, health and safety staff for your facility and/or local regulatory
agencies for further disposal information.

Items to collect
Powder pillows

Description Quantity
®
FerroVer Iron Reagent Powder Pillows, 10-mL1 1
Sample cells. (For information about sample cells, adapters or light shields, refer to Instrument-
2
specific information on page 1.)
1 FerroVer is a registered trademark of Hach Company.

Refer to Consumables and replacement items on page 6 for reorder information.

AccuVac Ampuls

Description Quantity
® ®
FerroVer Iron Reagent AccuVac Ampul 1
Beaker, 50-mL 1
Sample cells (For information about sample cells, adapters or light shields, refer to Instrument-
1
specific information on page 1.)
Stopper for 18-mm tubes and AccuVac Ampuls 1

Refer to Consumables and replacement items on page 6 for reorder information.

Sample collection and storage

• Collect samples in clean glass or plastic bottles.
• To measure only dissolved iron, filter the sample immediately after collection and
before acidification.
• To preserve samples for later analysis, adjust the sample pH to less than 2 with
concentrated nitric acid (about 2 mL per liter). No acid addition is necessary if the
sample is tested immediately.
• Keep preserved samples at room temperature for a maximum of 6 months.
• Before analysis, adjust the pH to 3–5 with 5.0 N sodium hydroxide standard solution.
• Correct the test result for the dilution from the volume additions.

2 Iron, Total, FerroVer Method (3.00 mg/L)

Powder pillow procedure

Start

1. Start program 265 Iron, 2. Prepare the sample: Fill 3. Add FerroVer Iron 4. Swirl the sample cell to
FerroVer. For information a sample cell with 10 mL of Reagent Powder Pillow to mix. Undissolved powder
about sample cells, sample. the sample cell. will not affect accuracy.
adapters or light shields,
refer to Instrument-specific
information on page 1.
Note: Although the program
name may vary between
instruments, the program
number does not change.

5. Start the instrument 6. Prepare the blank: Fill a 7. Clean the blank. 8. When the timer expires,
timer. A 3-minute reaction second sample cell with insert the blank into the cell
time starts. 10 mL of sample. holder.
An orange color will show if
iron is present. Let samples
that contain rust react for
5 minutes or more.

Zero Read

9. Push ZERO. The display 10. Clean the prepared 11. Insert the prepared 12. Push READ. Results
shows 0.00 mg/L Fe. sample. sample into the cell holder. show in mg/L Fe.

Iron, Total, FerroVer Method (3.00 mg/L) 3

AccuVac procedure

Start

1. Start program 267 Iron, 2. Prepare the blank: Fill 3. Prepare the sample: 4. Quickly invert the Ampul
FerroVer AV. For the sample cell with 10 mL Collect at least 40 mL of several times to mix.
information about sample of sample. sample in a 50-mL beaker. Undissolved powder will not
cells, adapters or light Fill the AccuVac Ampul with affect accuracy.
shields, refer to Instrument- sample. Keep the tip
specific information immersed while the Ampul
on page 1. fills completely.
Note: Although the program
name may vary between
instruments, the program
number does not change.

Zero

5. Start the instrument 6. Clean the blank. 7. When the timer expires, 8. Push ZERO. The display
timer. A 3-minute reaction insert the blank into the cell shows 0.00 mg/L Fe.
time starts. holder.
An orange color will show if
iron is present. Let samples
that contain rust react for
5 minutes or more.

Read

9. Clean the AccuVac 10. Insert the prepared 11. Push READ. Results
Ampul. sample AccuVac Ampul into show in mg/L Fe.
the cell holder.

4 Iron, Total, FerroVer Method (3.00 mg/L)

Interferences
Interfering substance Interference level
Calcium, Ca2+ No effect at less than 10,000 mg/L as CaCO3.
Chloride, Cl– No effect at less than 185,000 mg/L.
Copper, Cu2+ No effect. Masking agent is contained in FerroVer Reagent.
High iron levels Inhibit color development. Dilute sample and re-test to verify results.
Iron oxide A mild, vigorous or Digesdahl digestion is necessary. After digestion, adjust the sample pH to 3–
5 with sodium hydroxide, then analyze.
Magnesium No effect at 100,000 mg/L as CaCO3.
Molybdate molybdenum No effect at 50 mg/L as Mo.
High sulfide levels, S2– Pre-treat the sample in a fume hood or well-ventilated area before analysis:

1. Add 5 mL of hydrochloric acid, ACS to 100 mL of sample in a 250-mL Erlenmeyer flask.

2. Boil for 20 minutes.
3. Let the solution cool to room temperature.
4. Adjust the pH to 3–5 with sodium hydroxide.
5. Add deionized water until the volume is 100 mL.
6. Use the treated sample in the test procedure.

Turbidity Pre-treat the sample before analysis:

®
1. Add 0.1 g scoop of RoVer Rust Remover to the blank. Swirl to mix.
2. If sample remains turbid, add three 0.2 g scoops of RoVer Rust Remover to a 75 mL sample.
Let stand 5 minutes.
3. Filter through a glass membrane filter and filter holder.
4. Use the treated sample in the test procedure.

Highly buffered samples Can prevent the correct pH adjustment of the sample by the reagents. Sample pretreatment may
or extreme sample pH be necessary. Adjust the pH to 3–5.

Accuracy check
Standard additions method (sample spike)
Use the standard additions method (for applicable instruments) to validate the test
procedure, reagents and instrument and to find if there is an interference in the sample.
Items to collect:
• Iron Voluette® Ampule Standard, 25 mg/L
• Ampule Breaker
®
• Pipet, TenSette , 0.1–1.0 mL and tips

1. Use the test procedure to measure the concentration of the sample, then keep the
(unspiked) sample in the instrument.
2. Go to the Standard Additions option in the instrument menu.
3. Select the values for standard concentration, sample volume and spike volumes.
4. Open the standard solution.
5. Prepare three spiked samples: use the TenSette pipet to add 0.1 mL, 0.2 mL and
0.3 mL of the standard solution, respectively, to three 10-mL portions of fresh sample.
Mix well.
®
Note: For AccuVac Ampuls, add 0.2 mL, 0.4 mL and 0.6 mL of the standard solution to three
50-mL portions of fresh sample.

Iron, Total, FerroVer Method (3.00 mg/L) 5

 6. Use the test procedure to measure the concentration of each of the spiked samples.
Start with the smallest sample spike. Measure each of the spiked samples in the
instrument.
7. Select Graph to compare the expected results to the actual results.
Note: If the actual results are significantly different from the expected results, make sure that
the sample volumes and sample spikes are measured accurately. The sample volumes and
sample spikes that are used should agree with the selections in the standard additions menu. If
the results are not within acceptable limits, the sample may contain an interference.

Standard solution method

Use the standard solution method to validate the test procedure, reagents and
instrument.
Items to collect:
• Iron standard solution, 100 mg/L
• 100-mL volumetric flask, Class A
• 2-mL volumetric pipet, Class A and pipet filler
• Deionized water

1. Prepare a 2.00 mg/L iron standard solution as follows:

a. Use a pipet to add 2 mL of the 100 mg/L iron standard solution into the volumetric
flask.
b. Dilute to the mark with deionized water. Mix well. Prepare this solution daily.
2. Use the test procedure to measure the concentration of the prepared standard
solution.
3. Compare the expected result to the actual result.
Note: The factory calibration can be adjusted slightly with the standard adjust option so that the
instrument shows the expected value of the standard solution. The adjusted calibration is then
used for all test results. This adjustment can increase the test accuracy when there are slight
variations in the reagents or instruments.

Method performance
The method performance data that follows was derived from laboratory tests that were
measured on a spectrophotometer during ideal test conditions. Users may get different
results under different test conditions.
Program Standard Precision (95% Confidence Interval) Sensitivity
Concentration change per 0.010 Abs change
265 2.00 mg/L Fe 1.99–2.01 mg/L Fe 0.021 mg/L Fe
267 2.00 mg/L Fe 1.98–2.02 mg/L Fe 0.023 mg/L Fe

Summary of method
FerroVer Iron Reagent converts all soluble iron and most insoluble forms of iron in the
sample to soluble ferrous iron. The ferrous iron reacts with the 1-10 phenanthroline
indicator in the reagent to form an orange color in proportion to the iron concentration.
The measurement wavelength is 510 nm for spectrophotometers or 520 nm for
colorimeters.
Consumables and replacement items
Required reagents

Description Quantity/Test Unit Item no.

®
FerroVer Iron Reagent Powder Pillow, 10-mL 1 100/pkg 2105769
OR
® ®
FerroVer Iron Reagent AccuVac Ampul 1 25/pkg 2507025

6 Iron, Total, FerroVer Method (3.00 mg/L)

Required apparatus

Description Quantity/Test Unit Item no.

Beaker, 50-mL 1 each 50041H

Stoppers for 18-mm tubes and AccuVac Ampuls 2 6/pkg 173106

Recommended standards and apparatus

Description Unit Item no.

Flask, volumetric, Class A, 100-mL each 1457442

Iron Standard Solution, 100 mg/L Fe 100 mL 1417542
®
Iron Standard Solution, 10 mL Voluette ampule, 25 mg/L Fe 16/pkg 1425310
Metals Drinking Water Standard, LR for Cu, Fe, Mn 500 mL 2833749
Metals Drinking Water Standard, HR for Cu, Fe, Mn 500 mL 2833649
Pipet filler, safety bulb each 1465100
®
Pipet, TenSette , 0.1–1.0 mL each 1970001
Pipet tips for TenSette Pipet 1970001 50/pkg 2185696
Pipet tips for TenSette Pipet 1970001 1000/pkg 2185628
Pipet, volumetric, Class A, 2-mL each 1451536
Water, deionized 4L 27256

Optional reagents and apparatus

Description Unit Item no.

®
AccuVac Snapper each 2405200
Cylinder, mixing, 50-mL each 189641
Filter, glass membrane, 47-mm 100/pkg 253000
Filter holder for glass membrane filter each 234000
Hydrochloric Acid, concentrated 500 mL 13449
Nitric Acid, concentrated 500 mL 15249
RoVer Rust Remover 454 g 30001
100 mL
Sodium Hydroxide Standard Solution, 5.0 N 245032
MDB
Spoon, measuring, 0.1-g each 51100

Iron, Total, FerroVer Method (3.00 mg/L) 7

 FOR TECHNICAL ASSISTANCE, PRICE INFORMATION AND ORDERING: HACH COMPANY
In the U.S.A. – Call toll-free 800-227-4224 WORLD HEADQUARTERS
Outside the U.S.A. – Contact the HACH office or distributor serving you. Telephone: (970) 669-3050
On the Worldwide Web – www.hach.com; E-mail – [email protected] FAX: (970) 669-2932

© Hach Company/Hach Lange GmbH, 1989–2013. All rights reserved. 04/2013, Edition 8
Iron, Total DOC316.53.01314

FerroVer® Method1 Method 10249

0.1 to 3.0, 1.0 to 30.0 and 10.0 to 300.0 mg/L Fe Powder Pillows
Scope and application: For oil and gas field waters; digestion is required for total iron determinations.2
1 USEPA approved for reporting wastewater analysis, Federal Register, June 27, 1980; 45 (126:43459).
2 Adapted from Standard Methods for the Examination of Water and Wastewater.

Test preparation

Instrument specific information

The table in this section shows all of the instruments that have the program for this test.
Table 1 shows sample cell and orientation requirements for reagent addition tests, such
as powder pillow or bulk reagent tests.
To use the table, select an instrument, then read across to find the corresponding
information for this test.
Table 1 Instrument-specific information for powder pillows
Instrument Sample cell orientation Sample cell
DR 6000 The fill line is to the right. 2495402
DR 3800
DR 2800
DR 2700
DR 5000 The fill line is toward the user.
DR 3900
DR 900 The orientation mark is toward the user. 2401906

Before starting
Install the instrument cap on the DR 900 cell holder before ZERO or READ is pushed.
The sample must be digested with heat and acid to make sure that all forms of the metal are measured. Use the mild or
vigorous digestion. Refer to the Water Analysis Guide for more information.
For best results, measure the reagent blank value for each new lot of reagent. Replace the sample with deionized water in
the test procedure to get the reagent blank value. Subtract the reagent blank value from the sample results automatically
with the reagent blank adjust option.
Review the Safety Data Sheets (MSDS/SDS) for the chemicals that are used and use any recommended personal protective
equipment.
Dispose of reacted solutions according to local, state and federal regulations. Use the Safety Data Sheets for disposal
information for unused reagents. Consult the environmental, health and safety staff for your facility and/or local regulatory
agencies for further disposal information.

1
Items to collect
Description Quantity
®
FerroVer Iron Reagent Powder Pillow, 10-mL 1
EDTA solution, 1M 2 drops
Sample cells (For information about sample cells, adapters or light shields, refer to Instrument
1
specific information on page 1.)

Refer to Consumables and replacement items on page 6 for reorder information.

Sample collection and storage

• Collect samples in clean glass or plastic bottles.
• To preserve samples for later analysis, adjust the sample pH to less than 2 with
concentrated nitric acid (about 2 mL per liter). No acid addition is necessary if the
sample is tested immediately.
• To measure only dissolved iron, filter the sample immediately after collection and
before acidification.
• Keep preserved samples at room temperature for a maximum of 6 months.
• Before analysis, adjust the pH to 3–5 with 5.0 N sodium hydroxide standard solution.
• Correct the test result for the dilution from the volume additions.

Powder pillow procedure

Start

1. Start program 265 Iron, 2. Fill a clean sample cell 3. If the sample volume is 4. Swirl to mix.
FerroVer. For information with sample: less than 10 mL, add
about sample cells, deionized water to the 10-
adapters or light shields, • Use 10 mL of sample mL line.
refer to Instrument specific for the 0.02 to 3.0 mg/L
information on page 1. range.
• Use 1.0 mL of sample
Note: Although the program
name may vary between for the 0.2 to 30.0 mg/L
instruments, the program range with a dilution
number does not change. factor of 10.
• Use 0.1 mL of sample
for the 2.0 to
300.0 range with a
dilution factor of 100.
Note: Refer to Set the
dilution factor on page 4.

2 Iron, Total, FerroVer Method (multi-range: 3.0, 30.0, 300.0 mg/L)

5. Add 2 drops of EDTA 6. Swirl to mix. 7. Clean the sample cell. 8. Insert the sample cell
Solution 1 M to the sample. into the cell holder.

Zero

9. Push ZERO. The display 10. Remove the sample 11. Add the contents of one 12. Swirl to mix.
shows 0.0 mg/L Fe. from the cell holder. FerroVer Iron Reagent Accuracy is not affected by
Powder Pillow to the sample undissolved powder.
cell.

Read

13. Start the instrument 14. When the timer expires, 15. Insert the sample cell 16. Push READ. Results
timer. A 3-minute reaction clean the sample cell. into the cell holder. show in mg/L Fe.
time starts.
If iron is present in the
sample, an orange color will
show.

Interferences
Interfering Interference level
substance
Barium, Ba2+ The dilution of samples lowers most barium concentrations below interference levels. No effects are
seen on analyzed samples that contain less than 50 mg/L of Ba. No effects are seen when a 1.0 or
0.1 mL sample volume is used in the test procedure. A turbidity may show at higher levels. Use
5 drops of EDTA Solution in the test procedure and allow the sample to react for 5 minutes.
Calcium, Ca2+ No effect at less than 10,000 mg/L as CaCO3.
Chloride, Cl– No effect at less than 185,000 mg/L.

Iron, Total, FerroVer Method (multi-range: 3.0, 30.0, 300.0 mg/L) 3

Interfering Interference level
substance
Copper, Cu2+ No effect. Masking agent is contained in FerroVer Reagent.
High iron levels Inhibit color development. Dilute sample and re-test to verify results.
Magnesium No effect at 100,000 mg/L as CaCO3.
Molybdate No effect at 50 mg/L as Mo.
molybdenum
High sulfide levels, Pre-treat the sample in a fume hood or well-ventilated area before analysis:
S2–
1. Add 5 mL of hydrochloric acid, ACS to 100 mL of sample in a 250-mL Erlenmeyer flask.
2. Boil for 20 minutes.
3. Let the solution cool to room temperature.
4. Adjust the pH to 3–5 with sodium hydroxide.
5. Add deionized water until the volume is 100 mL.
6. Use the treated sample in the test procedure.

Strontium, Sr2+ Strontium by itself does not interfere. Strontium in combination with Barium will cause a precipitate to
form. The dilution of samples lowers most strontium concentrations below interference levels. No
effects are seen on analyzed samples that contain less than 50 mg/L of combined Ba and Sr. No
effects are seen when a 1.0 or 0.1 mL sample volume is used in the test procedure. A turbidity may
show at higher levels. Use 5 drops of EDTA Solution in the test procedure and allow the sample to
react for 5 minutes.
Highly buffered Can prevent the correct pH adjustment of the sample by the reagents. Sample pretreatment may be
samples or extreme necessary. Adjust the sample pH to 3–5 before the test is started. Correct the test result for the
sample pH dilution from the volume addition.

Set the dilution factor

Instruments that have a dilution factor option can include the dilution factor in the result
and show the concentration of the original, undiluted sample. For example, if the sample
is diluted by a factor of 10, the instrument multiplies the result by 10 and shows the
calculated result in the instrument display.

1. Select Options>More>Dilution factor from the instrument menu.

Note: Colorimeters include a dilution factor when the chemical form is set. Go to
Options>Advanced Options>Chemical Form and select LR, MR or HR.
2. Enter the dilution factor:
• 1 mL sample diluted to 10 mL: dilution factor is 10.
• 0.1 mL sample diluted to 10 mL: dilution factor is 100.
3. Push OK to confirm. Push OK again.
4. Push RETURN to go back to the measurement screen.

Accuracy check
Standard additions method (sample spike)
Use the standard additions method (for applicable instruments) to validate the test
procedure, reagents and instrument and to find if there is an interference in the sample.
Items to collect:
• Iron Voluette® Ampule Standard, 25 mg/L
• Ampule breaker
• Pipet, TenSette®, 0.1–1.0 mL and tips

1. Use the test procedure to measure the concentration of the sample, then keep the
(unspiked) sample in the instrument.
2. Go to the Standard Additions option in the instrument menu.

4 Iron, Total, FerroVer Method (multi-range: 3.0, 30.0, 300.0 mg/L)

 3. Select the values for standard concentration, sample volume and spike volumes.
4. Open the standard solution.
5. Prepare three spiked samples: use the TenSette pipet to add 0.1 mL, 0.2 mL and
0.3 mL of the standard solution, respectively, to three 10-mL portions of fresh sample.
Mix well.
6. Use the test procedure to measure the concentration of each of the spiked samples.
Start with the smallest sample spike. Measure each of the spiked samples in the
instrument.
7. Select Graph to compare the expected results to the actual results.
Note: If the actual results are significantly different from the expected results, make sure that
the sample volumes and sample spikes are measured accurately. The sample volumes and
sample spikes that are used should agree with the selections in the standard additions menu. If
the results are not within acceptable limits, the sample may contain an interference.

Standard solution method

Use the standard solution method to validate the test procedure, reagents and
instrument.
Items to collect:
• Iron Standard Solution, 100 mg/L
• 100-mL volumetric flask, Class A
• 2-mL volumetric pipet, Class A and pipet filler
• Deionized water

1. Prepare a 2.00 mg/L iron standard solution as follows:

a. Use a pipet to add 2.00 mL of 100 mg/L iron standard solution into the volumetric
flask.
b. Dilute to the mark with deionized water. Mix well. Prepare this solution daily.
2. Use the test procedure to measure the concentration of the prepared standard
solution.
3. Compare the expected result to the actual result.
Note: The factory calibration can be adjusted slightly with the standard adjust option so that the
instrument shows the expected value of the standard solution. The adjusted calibration is then
used for all test results. This adjustment can increase the test accuracy when there are slight
variations in the reagents or instruments.

Method performance
The method performance data that follows was derived from laboratory tests that were
measured on a spectrophotometer during ideal test conditions. Users may get different
results under different test conditions.
Program Standard Precision (95% Confidence Interval) Sensitivity
Concentration change per 0.010 Abs change
265 2.00 mg/L Fe 1.99–2.01 mg/L Fe 0.021 mg/L Fe

Summary of method
FerroVer Iron Reagent converts all soluble iron and most insoluble forms of iron in the
sample to soluble ferrous iron. The ferrous iron reacts with the 1-10 phenanthroline
indicator in the reagent to form an orange color in proportion to the iron concentration.
The measurement wavelength is 510 nm for spectrophotometers or 520 nm for
colorimeters.

Iron, Total, FerroVer Method (multi-range: 3.0, 30.0, 300.0 mg/L) 5

Consumables and replacement items
Required reagents

Description Quantity/test Unit Item no.

®
FerroVer Iron Reagent Powder Pillow, 10-mL 1 100/pkg 2105769
50 mL
EDTA Solution, 1 M 2 drops 2241926
SCDB

Recommended standards

Description Unit Item no.

Iron Standard Solution, 100 mg/L Fe 100 mL 1417542

®
Iron Standard Solution, 10 mL Voluette ampule, 25 mg/L Fe 16/pkg 1425310
Water, deionized 4L 27256
®
Pipet, TenSette , 0.1–1.0 mL each 1970001
Pipet tips for TenSette Pipet 1970001 50/pkg 2185696
Pipet tips for TenSette Pipet 1970001 1000/pkg 2185628
Flask, volumetric, Class A, 100-mL each 1457442
Pipet, volumetric, Class A, 2-mL each 1451536
Pipet filler, safety bulb each 1465100

Optional reagents and apparatus

Description Unit Item no.

Hydrochloric Acid, concentrated 500 mL 13449

Nitric Acid, concentrated 500 mL 15249
100 mL
Sodium Hydroxide Standard Solution, 5.0 N 245032
MDB
Filter, glass membrane, 47-mm 100/pkg 253000
Filter holder for glass membrane filter each 234000
RoVer Rust Remover 454 g 30001
Spoon, measuring, 0.1-g each 51100

FOR TECHNICAL ASSISTANCE, PRICE INFORMATION AND ORDERING: HACH COMPANY

In the U.S.A. – Call toll-free 800-227-4224 WORLD HEADQUARTERS
Outside the U.S.A. – Contact the HACH office or distributor serving you. Telephone: (970) 669-3050
On the Worldwide Web – www.hach.com; E-mail – [email protected] FAX: (970) 669-2932

© Hach Company/Hach Lange GmbH, 1989–2013. All rights reserved. 04/2013, Edition 8
Iron, Total DOC316.53.01051

TPTZ Method1 Method 8112

®
0.012 to 1.800 mg/L Fe (spectrophotometers) Powder Pillows or AccuVac Ampuls
0.04 to 1.80 mg/L Fe (colorimeters)
Scope and application: For water, wastewater and seawater.
1 Adapted from G. Frederic Smith Chemical Co., The Iron Reagents, 3rd ed. (1980).

Test preparation

Instrument-specific information
The tables in this section show all of the instruments that have the program for this test.
Table 1 shows sample cell and orientation requirements for reagent addition tests, such
as powder pillow or bulk reagent tests. Table 2 shows sample cell and adapter
requirements for AccuVac Ampul tests.
To use either table, select an instrument, then read across to find the corresponding
information for this test.
Table 1 Instrument-specific information for powder pillows
Instrument Sample cell orientation Sample cell
DR 6000 The fill line is to the right. 2495402
DR 3800
DR 2800
DR 2700
DR 5000 The fill line is toward the user.
DR 3900
DR 900 The orientation mark is toward the user. 2401906

Table 2 Instrument-specific information for AccuVac Ampuls

Instrument Adapter Sample cell
DR 6000 — 2427606
DR 5000
DR 900
DR 3900 LZV846 (A)
DR 3800 LZV584 (C) 2122800
DR 2800
DR 2700

Before starting
Install the instrument cap on the DR 900 cell holder before ZERO or READ is pushed.
The sample must be digested with heat and acid to make sure that all forms of the metal are measured. Use the mild or
vigorous digestion. Refer to the Water Analysis Guide for more information.

1
Wash all glassware with detergent. Rinse with tap water. Rinse again with 1:1 hydrochlorioc acid solution. Rinse a third time
with high-quality deionized water. These steps will remove deposits that can cause slightly high results.
For best results, measure the reagent blank value for each new lot of reagent. Replace the sample with deionized water in
the test procedure to get the reagent blank value. Subtract the reagent blank value from the sample results automatically
with the reagent blank adjust option.
Review the Safety Data Sheets (MSDS/SDS) for the chemicals that are used and use any recommended personal protective
equipment.
Dispose of reacted solutions according to local, state and federal regulations. Use the Safety Data Sheets for disposal
information for unused reagents. Consult the environmental, health and safety staff for your facility and/or local regulatory
agencies for further disposal information.

Items to collect
Powder pillows

Description Quantity

TPTZ Iron Reagent Powder Pillows, 10-mLS 2

Sample cells For information about sample cells, adapters or light shields, refer to Instrument-
2
specific information on page 1.

Refer to Consumables and replacement items on page 7 for reorder information.

AccuVac Ampuls

Description Quantity

TPTZ Low Range Iron Reactent AccuVac Ampul 1

Beaker, 50-mL 1
Sample cells (For information about sample cells, adapters or light shields, refer to Instrument-
1
specific information on page 1.)
Stopper for 18-mm tubes and AccuVac Ampuls 1

Refer to Consumables and replacement items on page 7 for reorder information.

Sample collection and storage

• Collect samples in clean glass or plastic bottles.
• To preserve samples for later analysis, adjust the sample pH to less than 2 with
concentrated nitric acid (about 2 mL per liter). No acid addition is necessary if the
sample is tested immediately.
• To measure only dissolved iron, filter the sample immediately after collection and
before acidification.
• Keep preserved samples at room temperature for a maximum of 6 months.
• Before analysis, adjust the pH to 3–4 with 5.0 N sodium hydroxide standard solution.
Do not exceed pH 5 or iron may precipitate.
• Correct the test result for the dilution from the volume additions.

2 Iron, Total, TPTZ Method (1.800 mg/L)

Powder pillow procedure

Start

1. Start program 270 Iron, 2. Prepare the sample: Fill 3. Add the contents of one 4. Swirl the sample cell for
TPTZ. For information about a sample cell with 10 mL of 10-mL TPTZ Iron Reagent at least 30 seconds to mix.
sample cells, adapters or sample. Powder Pillow to the
light shields, refer to prepared sample .
Instrument-specific
information on page 1.
Note: Although the program
name may vary between
instruments, the program
number does not change.

5. Start the instrument 6. Prepare the blank: Fill a 7. Add the contents of the 8. Swirl the sample cell for
timer. A 3-minute reaction sample cell with 10 mL of contents of one 10-mL at least 30 seconds to mix.
time starts. deionized water. TPTZ Iron Reagent Powder
Prepare the blank during the Pillow to the blank. .
reaction time.

Zero

9. When the timer expires, 10. Insert the blank into the 11. Push ZERO. The 12. Clean the prepared
clean the blank. cell holder. display shows 0.000 mg/L sample.
Fe.

Iron, Total, TPTZ Method (1.800 mg/L) 3

 Read

13. Insert the prepared 14. Push READ. Results

sample into the cell holder. show in mg/L Fe.

AccuVac Ampul procedure

Start

1. Start program 272 Iron, 2. Prepare the blank: Fill 3. Prepare the sample: 4. Quickly invert the Ampul
TPTZ AV. For information the sample cell with 10 mL Collect at least 40 mL of several times to mix.
about sample cells, of sample. sample in a 50-mL beaker.
adapters or light shields, Fill the AccuVac Ampul with
refer to Instrument-specific sample. Keep the tip
information on page 1. immersed while the Ampul
Note: Although the program fills completely.
name may vary between
instruments, the program
number does not change.

Zero

5. Start the instrument 6. When the timer expires, 7. Insert the blank into the 8. Push ZERO. The display
timer. A 3-minute reaction clean the blank. cell holder. shows 0.000 mg/L Fe.
time starts.

4 Iron, Total, TPTZ Method (1.800 mg/L)

 Read

9. Clean the AccuVac 10. Insert the prepared 11. Push READ. Results
Ampul. sample AccuVac Ampul into show in mg/L Fe.
the cell holder.

Interferences
Interferences were tested with an iron concentration of 0.5 mg/L Fe. The following do not
interfere with this method when present up to the levels shown.
Interfering substance Interference level
Cadmium 4.0 mg/L
Chromium3+ 0.25 mg/L
Chromium6+ 1.2 mg/L
Cobalt 0.05 mg/L
Copper 0.6 mg/L
Cyanide 2.8 mg/L
Manganese 50.0 mg/L
Mercury 0.4 mg/L
Molybdenum 4.0 mg/L
Nickel 1.0 mg/L
Nitrite Ion 0.8 mg/L
Color or turbidity If the sample, without a TPTZ Iron Reagent Powder Pillow, has a color or turbidity more than the
blank (deionized water plus TPTZ Iron Reagent), then use the sample as the blank. Refer to the
powder pillow procedure.
pH After the addition of reagent, a sample pH of less than 3 or more than 4 may inhibit color formation.
The developed color fades quickly or causes turbidity. Adjust the sample pH in the sample cell
before the addition of reagent:

1. Use a pH meter or pH paper to measure the current pH.

2. Add an applicable amount of iron-free acid or base such as 1.0 N Sulfuric Acid Standard
Solution or 1.0 N Sodium Hydroxide Standard Solution to adjust the sample pH to between
3 and 4.1
3. Make a volume correction if significant volumes of acid or base are used.

1 Refer to Consumables and replacement items on page 7 for reorder information.

Accuracy check
Standard additions method (sample spike)
Use the standard additions method (for applicable instruments) to validate the test
procedure, reagents and instrument and to find if there is an interference in the sample.
Items to collect:
• Iron Standard Solution, 10 mg/L Fe

Iron, Total, TPTZ Method (1.800 mg/L) 5

 • Pipet, TenSette®, 0.1–1.0 mL and pipet tips
• Mixing cylinders, 50-mL (3) (for AccuVac Ampuls)

1. Use the test procedure to measure the concentration of the sample, then keep the
(unspiked) sample in the instrument.
2. Go to the Standard Additions option in the instrument menu.
3. Select the values for standard concentration, sample volume and spike volumes.
4. Open the standard solution.
5. Prepare three spiked samples: use the TenSette pipet to add 0.1 mL, 0.2 mL and
0.3 mL of the standard solution, respectively, to three 10-mL portions of fresh sample.
Mix well.
®
Note: For AccuVac Ampuls, add 0.5 mL, 1.0 mL and 1.5 mL of the standard solution to three
50-mL portions of fresh sample.
6. Use the test procedure to measure the concentration of each of the spiked samples.
Start with the smallest sample spike. Measure each of the spiked samples in the
instrument.
7. Select Graph to compare the expected results to the actual results.
Note: If the actual results are significantly different from the expected results, make sure that
the sample volumes and sample spikes are measured accurately. The sample volumes and
sample spikes that are used should agree with the selections in the standard additions menu. If
the results are not within acceptable limits, the sample may contain an interference.

Standard solution method

Use the standard solution method to validate the test procedure, reagents and
instrument.
Items to collect:
• 100 mg/L iron standard solution
• 500-mL volumetric flask, Class A
• 5-mL volumetric pipet, Class A and pipet filler
• Deionized water

1. Prepare a 1.00 mg/L iron standard solution as follows:

a. Use a pipet to add 5.00 mL of 100 mg/L iron standard solution into the volumetric
flask.
b. Dilute to the mark with deionized water. Mix well. Prepare this solution daily.
2. Use the test procedure to measure the concentration of the prepared standard
solution.
3. Compare the expected result to the actual result.
Note: The factory calibration can be adjusted slightly with the standard adjust option so that the
instrument shows the expected value of the standard solution. The adjusted calibration is then
used for all test results. This adjustment can increase the test accuracy when there are slight
variations in the reagents or instruments.

Method performance
The method performance data that follows was derived from laboratory tests that were
measured on a spectrophotometer during ideal test conditions. Users may get different
results under different test conditions.
Program Standard Precision (95% Confidence Interval) Sensitivity
Concentration change per 0.010 Abs change
270 1.000 mg/L Fe 0.989–1.011 mg/L Fe 0.011 mg/L Fe
272 1.000 mg/L Fe 0.984–1.016 mg/L Fe 0.012 mg/L Fe

6 Iron, Total, TPTZ Method (1.800 mg/L)

Summary of method
The TPTZ Iron Reagent forms a deep blue-purple color with ferrous iron (Fe2+). The
indicator is combined with a reducing agent that converts precipitated or suspended iron,
such as rust, to the ferrous state. The amount of ferric iron (Fe3+) can be determined as
the difference between the results of a ferrous iron test and the concentration of total iron.
The measurement wavelength is 590 nm for spectrophotometers or 610 nm for
colorimeters.
Consumables and replacement items
Required reagents

Description Quantity/test Unit Item no.

TPTZ Iron Reagent Powder Pillow, 10-mL 1 100/pkg 2608799

OR
®
TPTZ Low Range Iron Reagent AccuVac Ampul 1 25/pkg 2510025

Required apparatus

Description Quantity/test Unit Item no.

Beaker, 50-mL 1 each 50041H

Sample cell, 10 mL round, 25 x 54 mm 1 each 2122800
Sample cell, 10 mL round, 25 x 60 mm 1 6/pkg 2427606
Sample cell, 10 mL square, matched pair 2 2/pkg 2495402

Recommended standards

Description Unit Item no.

Iron Standard Solution, 100 mg/L Fe 100 mL 1417542

Iron Standard Solution, 10 mg/L Fe 500 mL 14049
Iron Standard Solution, 1 mg/L Fe 500 mL 13949
Metals Drinking Water Standard, LR for Cu, Fe, Mn 500 mL 2833749
Metals Drinking Water Standard, HR for Cu, Fe, Mn 500 mL 2833649
Water, deionized 4L 27256

Optional reagents and apparatus

Description Unit Item no.

Cylinder, mixing, 50-mL each 189641

Nitric Acid, concentrated 500 mL 15249
50 mL
Sodium Hydroxide Standard Solution, 5.0 N 245026
SCDB
100 mL
Sulfuric Acid Standard Solution, 1 N 127032
MDB
100 mL
Sodium Hydroxide Standard Solution, 1.0 N 104532
MDB
Stoppers for 18-mm tubes and AccuVac Ampuls 6/pkg 173106
®
Pipet, TenSette , 0.1–1.0 mL each 1970001
Pipet tips for TenSette Pipet 1970001 50/pkg 2185696

Iron, Total, TPTZ Method (1.800 mg/L) 7

Consumables and replacement items (continued)
Description Unit Item no.

Flask, volumetric, Class A, 500-mL each 1457449

Pipet, volumetric 5.00-mL each 1451537
Pipet filler, safety bulb each 1465100

FOR TECHNICAL ASSISTANCE, PRICE INFORMATION AND ORDERING: HACH COMPANY

In the U.S.A. – Call toll-free 800-227-4224 WORLD HEADQUARTERS
Outside the U.S.A. – Contact the HACH office or distributor serving you. Telephone: (970) 669-3050
On the Worldwide Web – www.hach.com; E-mail – [email protected] FAX: (970) 669-2932

© Hach Company/Hach Lange GmbH, 1989–2013. All rights reserved. 04/2013, Edition 8
Manganese, LR DOC316.53.01057

1-(2-Pyridylazo)-2-Naphthol PAN Method1 Method 8149

0.006 to 0.700 mg/L Mn (LR) Powder Pillows
Scope and application: For water and wastewater; digestion is necessary for total manganese determinations.
1 Adapted from Goto, K., et al., Talanta, 24, 652-3 (1977).

Test preparation

Instrument specific information

The table in this section shows all of the instruments that have the program for this test.
Table 1 shows sample cell and orientation requirements for reagent addition tests, such
as powder pillow or bulk reagent tests.
To use the table, select an instrument, then read across to find the corresponding
information for this test.
Table 1 Instrument-specific information for powder pillows
Instrument Sample cell orientation Sample cell
DR 6000 The fill line is to the right. 2495402
DR 3800
DR 2800
DR 2700
DR 5000 The fill line is toward the user.
DR 3900
DR 900 The orientation mark is toward the user. 2401906

Before starting
Install the instrument cap on the DR 900 cell holder before ZERO or READ is pushed.
The sample must be digested with heat and acid to make sure that all forms of the metal are measured. Use the mild or
vigorous digestion. Refer to the Water Analysis Guide for more information.
Rinse all glassware with a 1:1 nitric acid solution. Rinse again with deionized water.
Review the Safety Data Sheets (MSDS/SDS) for the chemicals that are used and use any recommended personal protective
equipment.
The alkaline cyanide solution contains cyanide. Make sure to read the Safety Data Sheets and obey the safety precautions.
Dispose of reacted solutions according to local, state and federal regulations. Use the Safety Data Sheets for disposal
information for unused reagents. Consult the environmental, health and safety staff for your facility and/or local regulatory
agencies for further disposal information.

Items to collect
Description Quantity

Alkaline Cyanide Reagent 12 drops

Water, deionized 10 mL

1
Items to collect (continued)
Description Quantity

Ascorbic Acid Powder Pillow 2

PAN Indicator Solution, 0.1% 12 drops
Stoppers for 18 mm-tubes and AccuVac Ampuls 2
Sample cells (For information about sample cells, adapters or light shields, refer to Instrument
2
specific information on page 1.)

Refer to Consumables and replacement items on page 5 for reorder information.

Sample collection and storage

• Collect samples in clean glass or plastic bottles.
• To preserve samples for later analysis, adjust the sample pH to less than 2 with
concentrated nitric acid (about 2 mL per liter). No acid addition is necessary if the
sample is tested immediately.
• Keep preserved samples at room temperature for a maximum of 6 months.
• Before analysis, adjust the pH to 4–5 with 5.0 N sodium hydroxide standard solution.
• Correct the test result for the dilution from the volume additions.

Powder pillow procedure

Start

1. Start Program 2. Prepare the blank: Fill 3. Prepare the sample: Fill 4. Add the contents of one
290 Manganese, LR PAN. the sample cell with 10 mL a second sample cell with Ascorbic Acid Powder Pillow
For information about of deionized water. 10 mL of sample. to each sample cell.
sample cells, adapters or
light shields, refer to
Instrument specific
information on page 1.
Note: Although the program
name may vary between
instruments, the program
number does not change.

2 Manganese, PAN Method (0.700 mg/L)

5. Close both sample cells. 6. Add 12 drops of Alkaline 7. Swirl to mix. 8. Add 12 drops of PAN
Invert to dissolve the Cyanide Reagent Solution The solution may start to Indicator Solution, 0.1% to
powder. to each cell. show turbidity. This should each cell.
dissipate in the next step.

9. Swirl to mix. 10. Start the instrument 11. When the timer expires, 12. Insert the blank into the
The sample will show an timer. A 2-minute reaction clean the blank. cell holder.
orange color if manganese time starts.
is present.

Zero Read

13. Push ZERO. The 14. Clean the prepared 15. Insert the prepared 16. Push READ. Results
display shows 0.000 mg/L sample. sample into the cell holder. show in mg/L Mn.
Mn.

Interferences
Interfering substance Interference level
Aluminum 20 mg/L
Cadmium 10 mg/L
Calcium 1000 mg/L as CaCO3
Cobalt 20 mg/L
Copper 50 mg/L
Hardness For samples that contain more than 300 mg/L hardness as CaCO3, add 4 drops of Rochelle Salt
Solution to the sample after the Ascorbic Acid Powder Pillow is added.
Iron 25 mg/L (if the sample contains more than 5 mg/L iron, increase the reaction period to 10 minutes.)

Manganese, PAN Method (0.700 mg/L) 3

Interfering substance Interference level
Lead 0.5 mg/L
Magnesium 300 mg/L as CaCO3
Nickel 40 mg/L
Zinc 15 mg/L

Accuracy check
Standard additions method (sample spike)
Use the standard additions method (for applicable instruments) to validate the test
procedure, reagents and instrument and to find if there is an interference in the sample.
Items to collect:
®
• Manganese PourRite Ampule Standard, 10 mg/L Mn
• Ampule breaker
• Pipet, TenSette®, 0.1–1.0 mL and tips

1. Use the test procedure to measure the concentration of the sample, then keep the
(unspiked) sample in the instrument.
2. Go to the Standard Additions option in the instrument menu.
3. Select the values for standard concentration, sample volume and spike volumes.
4. Open the standard solution.
5. Prepare three spiked samples: use the TenSette pipet to add 0.1 mL, 0.2 mL and
0.3 mL of the standard solution, respectively, to three 10-mL portions of fresh sample.
Mix well.
6. Use the test procedure to measure the concentration of each of the spiked samples.
Start with the smallest sample spike. Measure each of the spiked samples in the
instrument.
7. Select Graph to compare the expected results to the actual results.
Note: If the actual results are significantly different from the expected results, make sure that
the sample volumes and sample spikes are measured accurately. The sample volumes and
sample spikes that are used should agree with the selections in the standard additions menu. If
the results are not within acceptable limits, the sample may contain an interference.

Standard solution method

Use the standard solution method to validate the test procedure, reagents and
instrument.
Items to collect:
• Manganese Voluette Standard Solution, 250 mg/L Mn
• 1-L volumetric flask, Class A
• 2.0-mL volumetric pipet, Class A and pipet filler
• Deionized water

1. Prepare a 0.5 mg/L manganese standard solution as follows:

a. Use a pipet to add 2.0 mL of the 250 mg/L standard solution into the volumetric
flask.
b. Dilute to the mark with deionized water. Mix well. Prepare this solution daily.
2. Use the test procedure to measure the concentration of the prepared standard
solution.
3. Compare the expected result to the actual result.
Note: The factory calibration can be adjusted slightly with the standard adjust option so that the
instrument shows the expected value of the standard solution. The adjusted calibration is then
used for all test results. This adjustment can increase the test accuracy when there are slight
variations in the reagents or instruments.

4 Manganese, PAN Method (0.700 mg/L)

Method performance
The method performance data that follows was derived from laboratory tests that were
measured on a spectrophotometer during ideal test conditions. Users may get different
results under different test conditions.
Program Standard Precision (95% Confidence Interval) Sensitivity
Concentration change per 0.010 Abs change
290 0.500 mg/L Mn 0.491–0.509 mg/L Mn 0.006 mg/L Mn

Summary of method
The PAN method is a highly sensitive and rapid procedure to measure low levels of
manganese. An ascorbic acid reagent is used initially to reduce all oxidized forms of
manganese to Mn2+. An alkaline-cyanide reagent is added to mask potential
interferences. PAN Indicator is then added to combine with the Mn2+ to form an orange-
colored complex. The measurement wavelength is 560 nm.
Consumables and replacement items
Required reagents

Description Quantity/Test Unit Item no.

Manganese Reagent Set, 10 mL, includes: — 50 tests 2651700

50 mL ,
Alkaline Cyanide Reagent 12 drops 2122326
SCDB
Ascorbic Acid Powder Pillow 1 100/pkg 1457799
50 mL
PAN Indicator Solution, 0.1% 12 drops 2122426
SCDB
Water, deionized varies 4L 27256

Recommended standards and apparatus

Description Unit Item no.

®
Manganese Standard Solution, 10-mg/L Mn, 2 mL PourRite ampule 20/pkg 2605820
®
Manganese Standard Solution, 250-mg/L Mn, 10-mL Voluette ampule 16/pkg 1425810
®
Ampule Breaker, Voluette ampules each 2196800
®
PourRite Ampule Breaker, 2-mL each 2484600
Metals Drinking Water Standard, HR for Cu, Fe, Mn 500 mL 2833649

Optional reagents and apparatus

Description Unit Item no.

Cylinder, mixing, 25-mL each 2088640

Flask, volumetric, Class A, 1000-mL each 1457453
®
Manganese Standard Solution, 2-mL PourRite Ampule, 25 mg/L 20/pkg 2112820
Nitric Acid, concentrated 500 mL 15249
Paper, pH, 0–14 pH range 100/pkg 2601300
Pipet filler, safety bulb each 1465100
®
Pipet, TenSette , 0.1–1.0 mL each 1970001
®
Pipet, TenSette , 1.0 to 10.0 mL each 1970010
Pipet tips for TenSette Pipet 1970001 50/pkg 2185696

Manganese, PAN Method (0.700 mg/L) 5

Consumables and replacement items (continued)
Description Unit Item no.

Pipet tips for TenSette Pipet 1970001 1000/pkg 2185628

Pipet tips for TenSette Pipet 1970010 50/pkg 2199796
Pipet tips for TenSette Pipet 1970010 250/pkg 2199725
Pipet, volumetric, Class A, 2-mL each 1451536
Rochelle Salt Solution 29 mL 172533
100 mL
Sodium Hydroxide Standard Solution, 5.0 N 245032
MDB

FOR TECHNICAL ASSISTANCE, PRICE INFORMATION AND ORDERING: HACH COMPANY

In the U.S.A. – Call toll-free 800-227-4224 WORLD HEADQUARTERS
Outside the U.S.A. – Contact the HACH office or distributor serving you. Telephone: (970) 669-3050
On the Worldwide Web – www.hach.com; E-mail – [email protected] FAX: (970) 669-2932

© Hach Company/Hach Lange GmbH, 1989–2013. All rights reserved. 04/2013, Edition 8
Manganese DOC316.53.01058

USEPA1 Periodate Oxidation Method2 Method 8034

0.1 to 20.0 mg/L Mn (HR) Powder Pillows
Scope and application: For soluble manganese in water and wastewater.
1 USEPA Approved for reporting wastewater analyses (digestion required). Federal Register, 44(116)34 193 (June 14, 1979).
2 Adapted from Standard Methods for the Examination of Water and Wastewater.

Test preparation

Instrument specific information

The table in this section shows all of the instruments that have the program for this test.
Table 1 shows sample cell and orientation requirements for reagent addition tests, such
as powder pillow or bulk reagent tests.
To use the table, select an instrument, then read across to find the corresponding
information for this test.
Table 1 Instrument-specific information for powder pillows
Instrument Sample cell orientation Sample cell
DR 6000 The fill line is to the right. 2495402
DR 3800
DR 2800
DR 2700
DR 5000 The fill line is toward the user.
DR 3900
DR 900 The orientation mark is toward the user. 2401906

Before starting
Install the instrument cap on the DR 900 cell holder before ZERO or READ is pushed.
The sample must be digested with heat and acid to make sure that all forms of the metal are measured. Use the mild or
vigorous digestion. Refer to the Water Analysis Guide for more information.
For best results, measure the reagent blank value for each new lot of reagent. Replace the sample with deionized water in
the test procedure to get the reagent blank value. Subtract the reagent blank value from the sample results automatically
with the reagent blank adjust option.
Review the Safety Data Sheets (MSDS/SDS) for the chemicals that are used and use any recommended personal protective
equipment.
Dispose of reacted solutions according to local, state and federal regulations. Use the Safety Data Sheets for disposal
information for unused reagents. Consult the environmental, health and safety staff for your facility and/or local regulatory
agencies for further disposal information.

1
Items to collect
Description Quantity

High Range Manganese Reagent Set, 10-mL 1

Sample cells (For information about sample cells, adapters or light shields, refer to Instrument
2
specific information on page 1.)

Refer to Consumables and replacement items on page 5 for reorder information.

Sample collection and storage

• Collect samples in acid-washed plastic bottles. Do not use glass containers to
prevent possible adsorption of manganese to glass.
• If only dissolved manganese is to be determined, filter the sample before acid
addition.
• To preserve samples for later analysis, adjust the sample pH to less than 2 with
concentrated nitric acid (about 2 mL per liter). No acid addition is necessary if the
sample is tested immediately.
• Keep preserved samples at room temperature for a maximum of 6 months.
• Before analysis, adjust the pH to 4–5 with 5.0 N sodium hydroxide standard solution.
Do not exceed pH 5 to prevent precipitation of the manganese.
• Correct the test result for the dilution from the volume additions.

Powder pillow procedure

Start

1. Start program 2. Prepare the sample: Fill 3. Add the contents of one 4. Close the sample cell.
295 Manganese, HR. For a sample cell with 10 mL of Buffer Powder Pilow, Citrate Invert the sample cell to mix.
information about sample sample. Type for Manganese.
cells, adapters or light
shields, refer to Instrument
specific information
on page 1.
Note: Although the program
name may vary between
instruments, the program
number does not change.

2 Manganese, Periodate Oxidation Method (20.0 mg/L)

5. Add the contents of one 6. Close the sample cell. 7. Start the instrument 8. Prepare the blank: Fill a
Sodium Periodate Powder Invert to mix. A violet color timer. A 2-minute reaction second sample cell with
Pillow to the sample cell. will show if manganese is time starts. 10 mL of sample.
present in the sample.

Zero

9. When the timer expires, 10. Insert the blank into the 11. Push ZERO. The 12. Clean the prepared
clean the blank. cell holder. display shows 0.0 mg/L Mn. sample.

Read

13. Within eight minutes 14. Push READ. Results

after the timer expires, insert show in mg/L Mn.
the prepared sample into
the cell holder.

Interferences
Interfering substance Interference level
Calcium 700 mg/L
Chloride 70,000 mg/L
Iron 5 mg/L
Magnesium 100,000 mg/L
Highly buffered samples or extreme sample pH Can prevent the correct pH adjustment of the sample by the reagents.
Sample pretreatment may be necessary.

Manganese, Periodate Oxidation Method (20.0 mg/L) 3

Accuracy check
Standard additions method (sample spike)
Use the standard additions method (for applicable instruments) to validate the test
procedure, reagents and instrument and to find if there is an interference in the sample.
Items to collect:
• Manganese Voluette® Ampule Standard, 250 mg/L Mn
• Ampule breaker
• Pipet, TenSette®, 0.1–1.0 mL and tips

1. Use the test procedure to measure the concentration of the sample, then keep the
(unspiked) sample in the instrument.
2. Go to the Standard Additions option in the instrument menu.
3. Select the values for standard concentration, sample volume and spike volumes.
4. Open the standard solution.
5. Prepare three spiked samples: use the TenSette pipet to add 0.1 mL, 0.2 mL and
0.3 mL of the standard solution, respectively, to three 10-mL portions of fresh sample.
Mix well.
6. Use the test procedure to measure the concentration of each of the spiked samples.
Start with the smallest sample spike. Measure each of the spiked samples in the
instrument.
7. Select Graph to compare the expected results to the actual results.
Note: If the actual results are significantly different from the expected results, make sure that
the sample volumes and sample spikes are measured accurately. The sample volumes and
sample spikes that are used should agree with the selections in the standard additions menu. If
the results are not within acceptable limits, the sample may contain an interference.

Standard solution method

Use the standard solution method to validate the test procedure, reagents and
instrument.
Items to collect:
• Manganese Standard Solution, 1000 mg/L
• 1-L volumetric flask, Class A
• 10-mL volumetric pipet, Class A and pipet filler
• Deionized water

1. Prepare a 10.0 mg/L manganese standard solution as follows:

a. Use a pipet to add 10.00 mL of 1000 mg/L manganese standard solution into the
volumetric flask.
b. Dilute to the mark with deionized water. Mix well. Prepare this solution daily.
2. Use the test procedure to measure the concentration of the prepared standard
solution.
3. Compare the expected result to the actual result.
Note: The factory calibration can be adjusted slightly with the standard adjust option so that the
instrument shows the expected value of the standard solution. The adjusted calibration is then
used for all test results. This adjustment can increase the test accuracy when there are slight
variations in the reagents or instruments.

4 Manganese, Periodate Oxidation Method (20.0 mg/L)

Method performance
The method performance data that follows was derived from laboratory tests that were
measured on a spectrophotometer during ideal test conditions. Users may get different
results under different test conditions.
Program Standard Precision (95% Confidence Interval) Sensitivity
Concentration change per 0.010 Abs change
295 10.0 mg/L Mn 9.6–10.4 mg/L Mn 0.1 mg/L Mn

Summary of method
Manganese in the sample is oxidized to the purple permanganate state by sodium
periodate, after buffering the sample with citrate. The purple color is directly proportional
to the manganese concentration. The measurement wavelength is 525 nm for
spectrophotometers or 520 nm for colorimeters.
Consumables and replacement items
Required reagents

Description Quantity/test Unit Item no.

Manganese Reagent Set, High Range, 10-mL includes: 1 100/pkg 2430000

Buffer Powder Pillows, Citrate for Manganese, 10-mL 1 100/pkg 2107669
Sodium Periodate Powder Pillow for Manganese, 10-mL 1 100/pkg 2107769

Required apparatus

Description Quantity/test Unit Item no.

Sample cell, 10 mL square, matched pair 2 2/pkg 2495402

Stoppers for 18-mm tubes and AccuVac Ampuls 2 6/pkg 173106

Recommended standards and apparatus

Description Unit Item no.

Manganese Standard Solution, 1000 mg/L Mn 100 mL 1279142

®
Manganese Standard Solution, 250-mg/L Mn, 10-mL Voluette ampule 16/pkg 1425810
Water, deionized 4L 27256
®
Ampule Breaker, Voluette ampules each 2196800

Optional reagents and apparatus

Description Unit Item no.

®
Manganese Standard Solution, 2-mL PourRite Ampule, 25 mg/L 20/pkg 2112820
®
Manganese Standard Solution, 10-mg/L Mn, 2 mL PourRite ampule 20/pkg 2605820
Paper, pH, 0–14 pH range 100/pkg 2601300
Pipet filler, safety bulb each 1465100
®
Pipet, TenSette , 0.1–1.0 mL each 1970001
Pipet tips for TenSette Pipet 1970001 50/pkg 2185696
Pipet tips for TenSette Pipet 1970001 1000/pkg 2185628
®
Pipet, TenSette , 1.0 to 10.0 mL each 1970010
Pipet tips for TenSette Pipet 1970010 250/pkg 2199725

Manganese, Periodate Oxidation Method (20.0 mg/L) 5

Consumables and replacement items (continued)
Description Unit Item no.

Pipet tips for TenSette Pipet 1970010 50/pkg 2199796

®
PourRite Ampule Breaker, 2-mL each 2484600
100 mL
Sodium Hydroxide Standard Solution, 5.0 N 245032
MDB
Flask, volumetric, Class A, 1000-mL each 1457453
Pipet, volumetric, Class A, 10-mL each 1451538

FOR TECHNICAL ASSISTANCE, PRICE INFORMATION AND ORDERING: HACH COMPANY

In the U.S.A. – Call toll-free 800-227-4224 WORLD HEADQUARTERS
Outside the U.S.A. – Contact the HACH office or distributor serving you. Telephone: (970) 669-3050
On the Worldwide Web – www.hach.com; E-mail – [email protected] FAX: (970) 669-2932

© Hach Company/Hach Lange GmbH, 1989–2013. All rights reserved. 04/2013, Edition 8
Molybdenum, HR DOC316.53.01061

Mercaptoacetic Acid Method1 Method 8036

®
0.2 to 40.0 mg/L Mo (HR) Powder Pillows or AccuVac Ampuls
Scope and application: For water, wastewater, boiler and cooling waters.
1 Adapted from Analytical Chemistry, 25(9) 1363 (1953).

Test preparation

Instrument-specific information
The tables in this section show all of the instruments that have the program for this test.
Table 1 shows sample cell and orientation requirements for reagent addition tests, such
as powder pillow or bulk reagent tests. Table 2 shows sample cell and adapter
requirements for AccuVac Ampul tests.
To use either table, select an instrument, then read across to find the corresponding
information for this test.
Table 1 Instrument-specific information for powder pillows
Instrument Sample cell orientation Sample cell
DR 6000 The fill line is to the right. 2495402
DR 3800
DR 2800
DR 2700
DR 5000 The fill line is toward the user.
DR 3900
DR 900 The orientation mark is toward the user. 2401906

Table 2 Instrument-specific information for AccuVac Ampuls

Instrument Adapter Sample cell
DR 6000 — 2427606
DR 5000
DR 900
DR 3900 LZV846 (A)
DR 3800 LZV584 (C) 2122800
DR 2800
DR 2700

Before starting
Install the instrument cap on the DR 900 cell holder before ZERO or READ is pushed.
For best results, measure the reagent blank value for each new lot of reagent. Replace the sample with deionized water in
the test procedure to get the reagent blank value. Subtract the reagent blank value from the sample results automatically
with the reagent blank adjust option.

1
Filter samples that are turbid with filter paper and a funnel.
Review the Safety Data Sheets (MSDS/SDS) for the chemicals that are used and use any recommended personal protective
equipment.
Dispose of reacted solutions according to local, state and federal regulations. Use the Safety Data Sheets for disposal
information for unused reagents. Consult the environmental, health and safety staff for your facility and/or local regulatory
agencies for further disposal information.

Items to collect
Powder pillows

Description Quantity
®
MolyVer 1 Molybdenum Reagent Powder Pillow, 10-mL 1
®
MolyVer 2 Molybdenum Reagent Powder Pillow, 10-mL 1
®
MolyVer 3 Molybdenum Reagent Powder Pillow, 10-mL 1
Sample cells (For information about sample cells, adapters or light shields, refer to Instrument-
2
specific information on page 1.)

Refer to Consumables and replacement items on page 7 for reorder information.

AccuVac Ampuls

Description Quantity

CDTA Solution, 0.4 M 4 drops

® ®
MolyVer 6 Reagent AccuVac Ampul 1
Beaker, 50 mL 1
Stopper for 18-mm tubes and AccuVac Ampuls 1
Sample cells (For information about sample cells, adapters or light shields, refer to Instrument-
1
specific information on page 1.)

Refer to Consumables and replacement items on page 7 for reorder information.

Sample collection and storage

• Collect samples in clean glass or plastic bottles.
• To preserve samples for later analysis, adjust the sample pH to less than 2 with
concentrated nitric acid (about 2 mL per liter). No acid addition is necessary if the
sample is tested immediately.
• Keep preserved samples at room temperature for a maximum of 6 months.
• Before analysis, adjust the pH to 7 with 5.0 N sodium hydroxide standard solution.
• Correct the test result for the dilution from the volume additions.

2 Molybdenum, Mercaptoacetic Acid Method (40.0 mg/L)

Powder pillow procedure

Start

1. Start program 2. Prepare the sample: Fill 3. Add the contents of one 4. Swirl to mix.
320 Molybdenum HR. For a sample cell with 10 mL of MolyVer 1 reagent powder
information about sample sample. pillow.
cells, adapters or light
shields, refer to Instrument-
specific information
on page 1.
Note: Although the program
name may vary between
instruments, the program
number does not change.

5. Add the contents of one 6. Swirl to mix. 7. Add the contents of one 8. Swirl to mix.
MolyVer 2 reagent powder MolyVer 3 reagent powder A yellow color will show if
pillow. pillow. molybdate molybdenum is
present.

9. Start the instrument 10. Prepare the blank: Fill 11. When the timer expires, 12. Insert the blank into the
timer. A 5-minute reaction a second sample cell with clean the blank. cell holder.
time starts. 10 mL of sample.

Molybdenum, Mercaptoacetic Acid Method (40.0 mg/L) 3

 Zero Read

13. Push ZERO. The 14. Clean the prepared 15. Insert the prepared 16. Push READ. Results
display shows 0.0 mg/L sample. sample into the cell holder. show in mg/L Mo6+.
Mo6+.

AccuVac Ampul procedure

Start

1. Start program 2. Prepare the blank: Fill 3. Prepare the sample: 4. Add Add four drops of
322 Molybdenum HR. For the sample cell with 10 mL Collect 40 mL of sample in a 0.4 M CDTA Standard
information about sample of sample. 50-mL beaker. Solution to the sample in the
cells, adapters or light beaker .
shields, refer to Instrument-
specific information
on page 1.
Note: Although the program
name may vary between
instruments, the program
number does not change.

5. Swirl to mix. 6. Fill an AccuVac Ampul 7. Quickly invert the Ampul 8. Start the instrument
with the treated sample. several times to mix. timer. A 5-minute reaction
Keep the tip immersed while A yellow color will show if time starts.
the Ampul fills completely. molybdate molybdenum is
present.

4 Molybdenum, Mercaptoacetic Acid Method (40.0 mg/L)

 Zero

9. When the timer expires, 10. Insert the blank into the 11. Push ZERO. The 12. Clean the AccuVac
clean the blank. cell holder. display shows 0.0 mg/L Ampul.
Mo6+.

Read

13. Insert the prepared 14. Push READ. Results

sample AccuVac Ampul into show in mg/L Mo6+.
the cell holder.

Interferences
Interfering substance Interference level
Aluminum More than 50 mg/L
Chromium More than 1000 mg/L
Copper Samples that contain 10 mg/L or more of copper will show a positive interference that
increases over time. Read these samples as soon as possible after the 5-minute reaction
period.
Iron More than 50 mg/L
Nickel More than 50 mg/L
Nitrite Add one Sulfamic Acid Powder Pillow to the sample to remove interference from up to
2000 mg/L as NO2–.
Highly buffered samples or Can prevent the correct pH adjustment of the sample by the reagents. Sample pretreatment
extreme sample pH may be necessary.

Accuracy check
Standard additions method (sample spike)
Use the standard additions method (for applicable instruments) to validate the test
procedure, reagents and instrument and to find if there is an interference in the sample.
Items to collect:
• Molybdenum Standard Solution, 1000-mg/L Mo6+
• Mixing cylinders, 50-mL (100-mL for AccuVac Ampuls) (3)

Molybdenum, Mercaptoacetic Acid Method (40.0 mg/L) 5

 • Pipet, TenSette®, 0.1–1.0 mL and tips

1. Use the test procedure to measure the concentration of the sample, then keep the
(unspiked) sample in the instrument.
2. Go to the Standard Additions option in the instrument menu.
3. Select the values for standard concentration, sample volume and spike volumes.
4. Open the standard solution.
5. Prepare three spiked samples: use the TenSette pipet to add 0.2 mL, 0.4 mL and
0.6 mL of the standard solution, respectively, to three 30-mL portions of fresh sample.
Mix well.
®
Note: For AccuVac Ampuls, add 0.4 mL, 0.8 mL and 1.2 mL of the standard solution to three
60-mL portions of fresh sample.
6. Use the test procedure to measure the concentration of each of the spiked samples.
Start with the smallest sample spike. Measure each of the spiked samples in the
instrument.
7. Select Graph to compare the expected results to the actual results.
Note: If the actual results are significantly different from the expected results, make sure that
the sample volumes and sample spikes are measured accurately. The sample volumes and
sample spikes that are used should agree with the selections in the standard additions menu. If
the results are not within acceptable limits, the sample may contain an interference.

Standard solution method

Use the standard solution method to validate the test procedure, reagents and
instrument.
Items to collect:
• Molybdenum Standard Solution, 10 mg/L Mo6+

1. Use the test procedure to measure the concentration of the standard solution.
2. Compare the expected result to the actual result.
Note: The factory calibration can be adjusted slightly with the standard adjust option so that the
instrument shows the expected value of the standard solution. The adjusted calibration is then
used for all test results. This adjustment can increase the test accuracy when there are slight
variations in the reagents or instruments.

Method performance
The method performance data that follows was derived from laboratory tests that were
measured on a spectrophotometer during ideal test conditions. Users may get different
results under different test conditions.
Program Standard Precision (95% Confidence Interval) Sensitivity
Concentration change per 0.010 Abs change
320 10 mg/L Mo6+ 9.7–10.3 mg/L Mo6+ 0.2 mg/L Mo6+
322 10 mg/L Mo6+ 9.7–10.3 mg/L Mo6+ 0.2 mg/L Mo6+

Summary of method
MolyVer 1 and 2 Reagents are added to buffer and condition the sample. MolyVer
3 provides the mercaptoacetic acid which reacts with molybdate molybdenum to form a
yellow color proportional to the molybdenum concentration. The measurement
wavelength is 420 nm.

6 Molybdenum, Mercaptoacetic Acid Method (40.0 mg/L)

Consumables and replacement items
Required reagents

Description Quantity/test Unit Item no.

Molybdenum Reagent Set, 10-mL, includes: — 100 tests 2604100

®
MolyVer 1 Molybdenum Reagent Powder Pillow, 10-mL 1 100/pkg 2604299
®
MolyVer 2 Molybdenum Reagent Powder Pillow, 10-mL 1 100/pkg 2604399
®
MolyVer 3 Molybdenum Reagent Powder Pillow, 10-mL 1 100/pkg 2604499
OR
® ®
MolyVer 6 Reagent AccuVac Ampul 1 25/pkg 2522025
15 mL
CDTA Solution, 0.4 M 4 drops 2615436
SCDB

Required apparatus

Description Quantity/test Unit Item no.

AccuVac Snapper 1 each 2405200

Beaker, 50-mL 1 each 50041H
Stoppers for 18-mm tubes and AccuVac Ampuls 2 6/pkg 173106

Recommended standards

Description Unit Item no.

Molybdenum Standard Solution, 10-mg/L as Mo 100 mL 1418742

Molybdenum Standard Solution, 1000-mg/L as Mo 100 mL 1418642
Water, deionized 4L 27256

Optional reagents and apparatus

Description Unit Item no.

Beaker, 50-mL each 50041H

Cylinder, mixing, 50-mL each 189641
Filter paper, 12.5-cm 100/pkg 189457
Funnel, poly, 65-mm each 108367
®
Pipet, TenSette , 0.1–1.0 mL each 1970001
Pipet tips for TenSette Pipet 1970001 50/pkg 2185696
Sulfamic Acid Powder Pillows 100/pkg 105599

Molybdenum, Mercaptoacetic Acid Method (40.0 mg/L) 7

 FOR TECHNICAL ASSISTANCE, PRICE INFORMATION AND ORDERING: HACH COMPANY
In the U.S.A. – Call toll-free 800-227-4224 WORLD HEADQUARTERS
Outside the U.S.A. – Contact the HACH office or distributor serving you. Telephone: (970) 669-3050
On the Worldwide Web – www.hach.com; E-mail – [email protected] FAX: (970) 669-2932

© Hach Company/Hach Lange GmbH, 1989–2013. All rights reserved. 04/2013, Edition 8
Molybdenum DOC316.53.01062

Ternary Complex Method Method 8169

0.02 to 3.00 mg/L Mo (LR) Powder Pillows
Scope and application: For boiler and cooling tower waters.

Test preparation

Instrument specific information

The table in this section shows all of the instruments that have the program for this test.
Table 1 shows sample cell and orientation requirements for reagent addition tests, such
as powder pillow or bulk reagent tests.
To use the table, select an instrument, then read across to find the corresponding
information for this test.
Table 1 Instrument-specific information for powder pillows
Instrument Sample cell orientation Sample cell
DR 6000 The fill line is to the right. 2495402
DR 3800
DR 2800
DR 2700
DR 5000 The fill line is toward the user.
DR 3900
DR 900 The orientation mark is toward the user. 2401906

Before starting
Samples must be analyzed immediately after collection and cannot be preserved for later analysis.
Install the instrument cap on the DR 900 cell holder before ZERO or READ is pushed.
After several analyses, the sample cells may have a blue discoloration. Clean the sample cells with 6.0 N (50%) hydrochloric
acid, then rinse thoroughly with deionized water.
Review the Safety Data Sheets (MSDS/SDS) for the chemicals that are used and use any recommended personal protective
equipment.
Dispose of reacted solutions according to local, state and federal regulations. Use the Safety Data Sheets for disposal
information for unused reagents. Consult the environmental, health and safety staff for your facility and/or local regulatory
agencies for further disposal information.

Items to collect
Description Quantity

Molybdenum Reagent Set for 20-mL sample 1

Molybdenum 1 Reagent (LR) Molybdate Powder Pillow, 20-mL 1
Molybdenum 2 Reagent Solution 0.5 mL

1
Items to collect (continued)
Description Quantity

Cylinder, graduated mixing, 25-mL 1

Sample cells (For information about sample cells, adapters or light shields, refer to Instrument
2
specific information on page 1.)

Refer to Consumables and replacement items on page 6 for reorder information.

Sample collection
• Samples must be analyzed immediately after collection and cannot be preserved for
later analysis.
• Collect samples in clean glass or plastic bottles.
• Filter samples that are turbid with filter paper and a funnel.

Powder pillow procedure

Start

1. Start program 2. Prepare the sample: Fill 3. Add the contents of one 4. Close the cylinder.
315 Molybdenum LR. For a 25-mL mixing cylinder with Molybdenum 1 Reagent Shake the cylinder to
information about sample 20 mL of sample. Powder Pillow to the mixing completely dissolve the
cells, adapters or light cylinder. reagent.
shields, refer to Instrument
specific information
on page 1.
Note: Although the program
name may vary between
instruments, the program
number does not change.

5. Fill a sample cell with 6. Develop the sample: 7. Swirl to mix. 8. Start the instrument
10 mL of the prepared Add 0.5 mL of Molybdenum timer. A 2-minute reaction
sample. 2 Reagent Solution to the time starts.
prepared sample cell.

2 Molybdenum, Ternary Complex Method (3.00 mg/L)

 Zero

9. Prepare the blank: 10. Clean the blank. 11. Insert the blank into the 12. Push ZERO. The
When the timer expires, fill a cell holder. display shows 0.00 mg/L
second sample cell with Mo6+.
10 mL of the rest of the
prepared sample.

Read

13. Clean the developed 14. Insert the developed 15. Push READ. Results
sample. sample into the cell holder. show in mg/L Mo6+.

Interferences
Interference studies were completed with a molybdenum standard solution of 2 mg/L
Mo6+ that included the potential interfering ion. When the standard solution concentration
changed by ± 5% with a given ion concentration, the ion was considered to be a
substance that interferes. Interference results are summarized in Table 2, Table 3 and
Table 4.
Table 2 Substances that cause a negative interference
Interfering substance Interference level
Alum More than 7 mg/L
Aluminum More than 2 mg/L
AMP (Phosphonate) More than 15 mg/L
Bicarbonate More than 5650 mg/L
Bisulfate More than 3300 mg/L
Borate More than 5250 mg/L
Chloride More than 1400 mg/L
Chromium More than 4.5 mg/L: read the molybdenum concentration immediately after
the 2-minute reaction period.
Copper More than 98 mg/L
Diethanoldithiocarbamate More than 32 mg/L
EDTA More than 1500 mg/L
Ethylene Glycol More than 2% (by volume)

Molybdenum, Ternary Complex Method (3.00 mg/L) 3

 Table 2 Substances that cause a negative interference (continued)
Interfering substance Interference level
Highly buffered samples or extreme sample pH Can prevent the correct pH adjustment of the sample by the reagents.
Sample pretreatment may be necessary. Adjust to pH 3–5 with acid (Sulfuric
Acid, 1.0 N) or base (Sodium Hydroxide, 1.0 N). Correct the test result for
the dilution from the volume additions.
Iron More than 200 mg/L
Lignin Sulfonate More than 105 mg/L
Nitrite More than 350 mg/L
Orthophosphate More than 4500 mg/L
Phosphonohydroxyacetic Acid More than 32 mg/L
Phosphonate HEDP Positive interference of about 10% up to 30 mg/L. As the concentration
increases above 30 mg/L, a decrease in the molybdenum concentration
reading occurs (negative interference).
Sulfite More than 6500 mg/L

Table 3 Substances that cause a positive interference

Interfering substance Interference level
Benzotriazole More than 210 mg/L
Carbonate More than 1325 mg/L
Morpholine More than 6 mg/L
Phosphonate HEDP The presence of the phosphonate HEDP at concentrations up to 30 mg/L will increase the apparent
molybdenum concentration reading by approximately 10% (positive interference). Multiply the test
result by 0.9 to get the actual Mo6+ concentration.
Silica More than 600 mg/L

Table 4 Non-interfering substances

Interfering substance Interference level
Bisulfite 9600 mg/L
Calcium 720 mg/L
Chlorine 7.5 mg/L
Magnesium 8000 mg/L
Manganese 1600 mg/L
Nickel 250 mg/L
PBTC (phosphonate) 500 mg/L
Sulfate 12,800 mg/L
Zinc 400 mg/L

Accuracy check
Standard additions method (sample spike)
Use the standard additions method (for applicable instruments) to validate the test
procedure, reagents and instrument and to find if there is an interference in the sample.
Items to collect:
• Molybdenum Standard Solution, 1000 mg/L Mo6+
• Graduated cylinder, 250 mL

4 Molybdenum, Ternary Complex Method (3.00 mg/L)

 • Pipet, TenSette®, 0.1–1.0 mL and tips
• Erlenmeyer flasks (3)

1. Use the test procedure to measure the concentration of the sample, then keep the
(unspiked) sample in the instrument.
2. Go to the Standard Additions option in the instrument menu.
3. Select the values for standard concentration, sample volume and spike volumes.
4. Open the standard solution.
5. Prepare three spiked samples: use the TenSette pipet to add 0.1 mL, 0.2 mL and
0.3 mL of the standard solution, respectively, to three 200-mL portions of fresh
sample. Mix well.
6. Use the test procedure to measure the concentration of each of the spiked samples.
Start with the smallest sample spike. Measure each of the spiked samples in the
instrument.
7. Select Graph to compare the expected results to the actual results.
Note: If the actual results are significantly different from the expected results, make sure that
the sample volumes and sample spikes are measured accurately. The sample volumes and
sample spikes that are used should agree with the selections in the standard additions menu. If
the results are not within acceptable limits, the sample may contain an interference.

Standard solution method

Use the standard solution method to validate the test procedure, reagents and
instrument.
Items to collect:
• Molybdenum Standard Solution, 10.00-mg/L
• 50-mL volumetric flask, Class A
• 10-mL volumetric pipet, Class A and pipet filler
• Deionized water

1. Prepare a 2.00 mg/L molybdenum standard solution as follows:

a. Use a pipet to add 10.00 mL of 10.00 mg/L molybdenum standard solution into
the volumetric flask.
b. Dilute to the mark with deionized water. Mix well. Prepare this solution daily.
2. Use the test procedure to measure the concentration of the prepared standard
solution.
3. Compare the expected result to the actual result.
Note: The factory calibration can be adjusted slightly with the standard adjust option so that the
instrument shows the expected value of the standard solution. The adjusted calibration is then
used for all test results. This adjustment can increase the test accuracy when there are slight
variations in the reagents or instruments.

Method performance
The method performance data that follows was derived from laboratory tests that were
measured on a spectrophotometer during ideal test conditions. Users may get different
results under different test conditions.
Program Standard Precision (95% Confidence Interval) Sensitivity
Concentration change per 0.010 Abs change
315 2.00 mg/L Mo6+ 1.94–2.06 mg/L Mo6+ 0.02 mg/L Mo6+

Summary of method
The ternary complex method for molybdenum determination is a method in which
molybdate molybdenum reacts with an indicator and a sensitizing agent to give a stable
blue complex. The measurement wavelength is 610 nm.

Molybdenum, Ternary Complex Method (3.00 mg/L) 5

Consumables and replacement items
Required reagents

Description Quantity/test Unit Item no.

Molybdenum Reagent Set, 20-mL, includes: — 100 tests 2449400

Molybdenum 1 Reagent (LR) Molybdate Powder Pillow, 20-mL 1 100/pkg 2352449
Molybdenum 2 Reagent Solution 0.5 mL 50 mL MDB 2352512

Required apparatus

Description Quantity/test Unit Item no.

Cylinder, graduated mixing, 25-mL 1 each 189640

Sample cell, 10 mL square, matched pair 2 2/pkg 2495402

Recommended standards

Description Unit Item no.

Molybdenum Standard Solution, 10-mg/L as Mo 100 mL 1418742

Molybdenum Standard Solution, 1000-mg/L as Mo 100 mL 1418642
Water, deionized 4L 27256

Optional reagents and apparatus

Description Unit Item no.

Cylinder graduated, 250-mL each 108146

Filter paper, 12.5-cm 100/pkg 189457
Funnel, poly, 65-mm each 108367
®
Pipet, TenSette , 0.1–1.0 mL each 1970001
Pipet tips for TenSette Pipet 1970001 50/pkg 2185696
Flask, Erlenmeyer, 250-mL each 50546
Pipet, volumetric, Class A, 10-mL each 1451538
Pipet filler, safety bulb each 1465100
Hydrochloric Acid, 6.0 N 1:1, 50% 500 mL 88449
100 mL
Sodium Hydroxide Standard Solution, 1.0 N 104532
MDB
100 mL
Sulfuric Acid Standard Solution, 1 N 127032
MDB

FOR TECHNICAL ASSISTANCE, PRICE INFORMATION AND ORDERING: HACH COMPANY

In the U.S.A. – Call toll-free 800-227-4224 WORLD HEADQUARTERS
Outside the U.S.A. – Contact the HACH office or distributor serving you. Telephone: (970) 669-3050
On the Worldwide Web – www.hach.com; E-mail – [email protected] FAX: (970) 669-2932

© Hach Company/Hach Lange GmbH, 1989–2013. All rights reserved. 04/2013, Edition 8
Nickel DOC316.53.01063

1-(2 Pyridylazo)-2-Napthol (PAN) Method1 Method 8150

0.006 to 1.000 mg/L Ni Powder Pillows
Scope and application: For water and wastewater.
1 Adapted from Watanabe, H., Talanta, 21 295 (1974).

Test preparation

Instrument specific information

The table in this section shows all of the instruments that have the program for this test.
Table 1 shows sample cell and orientation requirements for reagent addition tests, such
as powder pillow or bulk reagent tests.
To use the table, select an instrument, then read across to find the corresponding
information for this test.
Table 1 Instrument-specific information for powder pillows
Instrument Sample cell orientation Sample cell
DR 6000 The fill line is to the right. 2495402
DR 3800
DR 2800
DR 2700
DR 5000 The fill line is toward the user.
DR 3900
DR 900 The orientation mark is toward the user. 2401906

Before starting
Install the instrument cap on the DR 900 cell holder before ZERO or READ is pushed.
The sample must be digested with heat and acid to make sure that all forms of the metal are measured. Use the mild or
vigorous digestion. Refer to the Water Analysis Guide for more information.
For spectrophotometers, this method can measure the cobalt concentration on the same sample with Program Number 110.
Review the Safety Data Sheets (MSDS/SDS) for the chemicals that are used and use any recommended personal protective
equipment.
Dispose of reacted solutions according to local, state and federal regulations. Use the Safety Data Sheets for disposal
information for unused reagents. Consult the environmental, health and safety staff for your facility and/or local regulatory
agencies for further disposal information.

Items to collect
Description Quantity

EDTA Powder Pillow 2

Phthalate-Phosphate Reagent Powder Pillow 2
PAN Indicator Solution 0.3% 1 mL

1
Items to collect (continued)
Description Quantity

Deionized water 25 mL
Sample cells (For information about sample cells, adapters or light shields, refer to Instrument
2
specific information on page 1.)

Refer to Consumables and replacement items on page 6 for reorder information.

Sample collection and storage

• Collect samples in clean glass or plastic bottles.
• If the sample temperature is less than 10 °C (50 °F), warm the sample to room
temperature before analysis.
• To preserve samples for later analysis, adjust the sample pH to less than 2 with
concentrated nitric acid (about 2 mL per liter). No acid addition is necessary if the
sample is tested immediately.
• Keep preserved samples at room temperature for a maximum of 6 months.
• Before analysis, adjust the pH to 3–8 with 5.0 N sodium hydroxide standard solution.
Do not exceed pH 8 to prevent precipitation of the metal.
• Correct the test result for the dilution from the volume additions.

Powder pillow procedure

Start

1. Start program 2. Prepare the blank: Fill a 3. Prepare the sample: Fill 4. Add the contents of one
340 Nickel, PAN. For sample cell with 10 mL of a second sample cell with Phthalate-Phosphate
information about sample deionized water. 10 mL of sample. Reagent Powder Pillow to
cells, adapters or light each cell.
shields, refer to Instrument
specific information
on page 1.
Note: Although the program
name may vary between
instruments, the program
number does not change.

2 Nickel, PAN Method (1.000 mg/L)

5. Close the sample cells. 6. Add 0.5 mL of 0.3% PAN 7. Close the sample cells. 8. Start the instrument
Immediately shake to Indicator Solution to each Invert several times to mix. timer. A 15-minute reaction
dissolve the reagent. cell. time starts.
If the sample contains iron, During color development
make sure that all the the sample solution color
powder is dissolved before may vary from yellow-
the PAN Indicator is added. orange to dark red, based
on the chemical makeup of
the sample. The blank will
be yellow.

9. When the timer expires, 10. Close the sample cells. 11. Clean the blank. 12. Insert the blank into the
add the contents of one Shake to dissolve the cell holder.
EDTA Reagent Powder reagent powder.
Pillow to each cell.

Zero Read

13. Push ZERO. The 14. Clean the prepared 15. Insert the prepared 16. Push READ. Results
display shows 0.00 mg/L Ni sample. sample into the cell holder. show in mg/L Ni and Co
and Co (spectrophotometers) or
(spectrophotometers) or mg/L Ni (colorimeters).
mg/L Ni (colorimeters) *.

* Spectrophotometers "zero" at 560 nm and 620 nm. Colorimeters "zero" at 560 nm.

Nickel, PAN Method (1.000 mg/L) 3

Interferences
Interfering substance Interference level
Al3+ 32 mg/L
Ca2+ 1000 mg/L as (CaCO3)
Cd2+ 20 mg/L
Cl– 8000 mg/L
Chelating agents (e.g., EDTA) Interfere at all levels. Use either the Digesdahl or vigorous digestion to
remove this interference.
Cr3+ 20 mg/L
Cr6+ 40 mg/L
Cu2+ 15 mg/L
F– 20 mg/L
Fe3+ 10 mg/L
Fe2+ Interferes directly and must not be present.
K+ 500 mg/L
Mg2+ 400 mg/L
Mn2+ 25 mg/L
Mo6+ 60 mg/L
Na+ 5000 mg/L
Pb2+ 20 mg/L
Zn2+ 30 mg/L
Highly buffered samples or extreme sample pH Can prevent the correct pH adjustment of the sample by the reagents.
Sample pretreatment may be necessary.

Accuracy check
Standard additions method (sample spike)
Use the standard additions method (for applicable instruments) to validate the test
procedure, reagents and instrument and to find if there is an interference in the sample.
Items to collect:
• 1000-mg/L Nickel Standard Solution
• 100-mL volumetric flask, Class A
• 5-mL volumetric pipet, Class A and pipet filler
• Deionized water
• Pipet, TenSette®, 0.1–1.0 mL and tips
• Mixing cylinders, 25-mL (3)

1. Prepare a 50 mg/L nickel standard solution as follows:

a. Use a pipet to add 5.00 mL of a 1000 mg/L nickel standard solution into a 100-mL
volumetric flask.
b. Dilute to the mark with deionized water. Mix well. Prepare this solution daily.
2. Use the test procedure to measure the concentration of the sample, then keep the
(unspiked) sample in the instrument.
3. Go to the Standard Additions option in the instrument menu.
4. Select the values for standard concentration, sample volume and spike volumes.

4 Nickel, PAN Method (1.000 mg/L)

 5. Prepare three spiked samples: use the TenSette pipet to add 0.1 mL, 0.2 mL and
0.3 mL of the prepared standard solution, respectively, to three 25-mL portions of
fresh sample. Mix well.
6. Use the test procedure to measure the concentration of each of the spiked samples.
Start with the smallest sample spike. Measure each of the spiked samples in the
instrument.
7. Select Graph to compare the expected results to the actual results.
Note: If the actual results are significantly different from the expected results, make sure that
the sample volumes and sample spikes are measured accurately. The sample volumes and
sample spikes that are used should agree with the selections in the standard additions menu. If
the results are not within acceptable limits, the sample may contain an interference.

Standard solution method

Use the standard solution method to validate the test procedure, reagents and
instrument.
Items to collect:
• 1000-mg/L Nickel Standard Solution
• 1-L volumetric flask, Class A
• 100-mL volumetric flask, Class A
• 10-mL volumetric pipet, Class A and pipet filler
• 5-mL volumetric pipet, Class A and pipet filler
• Deionized water

1. Prepare a 5.00-mg/L nickel stock solution as follows:

a. Use a pipet to add 5.00 mL of a 1000-mg/L nickel standard solution into a 1-L
volumetric flask.
b. Dilute to the mark with deionized water. Mix well. Prepare the stock solution each
day.
2. Prepare a 0.5 mg/L nickel standard solution as follows:
a. Use a pipet to add 10.00 mL of the 5.00-mg/L nickel stock solution into a 100-mL
volumetric flask.
b. Dilute to the mark with deionized water. Mix well. Prepare the standard solution
each day.
3. Use the test procedure to measure the concentration of the prepared standard
solution.
4. Compare the expected result to the actual result.
Note: The factory calibration can be adjusted slightly with the standard adjust option so that the
instrument shows the expected value of the standard solution. The adjusted calibration is then
used for all test results. This adjustment can increase the test accuracy when there are slight
variations in the reagents or instruments.

Method performance
The method performance data that follows was derived from laboratory tests that were
measured on a spectrophotometer during ideal test conditions. Users may get different
results under different test conditions.
Program Standard Precision (95% Confidence Interval) Sensitivity
Concentration change per 0.010 Abs change
340 0.500 mg/L Ni 0.492–0.508 mg/L Ni 0.006 mg/L Ni

Summary of method
After pyrophosphate is added to buffer the sample and mask any Fe3+, the nickel reacts
with 1-(2-Pyridylazo)-2-Naphthol indicator. The indicator forms complexes with most
metals present. After color development, EDTA is added to destroy all metal-PAN
complexes except nickel and cobalt. Spectrophotometers automatically adjust for cobalt

Nickel, PAN Method (1.000 mg/L) 5

 interference by measuring the absorbance of the sample at both 560 nm and 620 nm.
This method is unique because both nickel and cobalt can be determined on the same
sample with a spectrophotometer. The measurement wavelength is 560 nm for
colorimeters.
Consumables and replacement items
Required reagents

Description Quantity/test Unit Item no.

Nickel Reagent Set, PAN, 10-mL 1 100/pkg 2651600

Includes:
EDTA Reagent Powder Pillow 2 100/pkg 700599
Phthalate-Phosphate Reagent Powder Pillow, 10-mL 2 100/pkg 2615199
100 mL
PAN Indicator Solution, 0.3% 1 mL 2150232
MDB
Water, deionized varies 4L 27256

Required apparatus

Description Quantity/test Unit Item no.

Stoppers for 18-mm tubes and AccuVac Ampuls 2 6/pkg 173106

Recommended standards

Description Unit Item no.

Nickel Standard Solution, 1000-mg/L Ni (NIST) 100 mL 1417642

Optional reagents and apparatus

Description Unit Item no.

Cylinder, mixing, 25-mL each 189640

Flask, volumetric, Class A, 100-mL each 1457442
Pipet, volumetric 5.00-mL each 1451537
Pipet filler, safety bulb each 1465100
Pipet, volumetric, Class A, 10-mL each 1451538
Flask, volumetric, Class A, 1000-mL each 1457453
Water, deionized 4L 27256
Nitric Acid Solution, 1:1 500 mL 254049
100 mL
Sodium Hydroxide Standard Solution, 5.0 N 245032
MDB

FOR TECHNICAL ASSISTANCE, PRICE INFORMATION AND ORDERING: HACH COMPANY

In the U.S.A. – Call toll-free 800-227-4224 WORLD HEADQUARTERS
Outside the U.S.A. – Contact the HACH office or distributor serving you. Telephone: (970) 669-3050
On the Worldwide Web – www.hach.com; E-mail – [email protected] FAX: (970) 669-2932

© Hach Company/Hach Lange GmbH, 1989–2013. All rights reserved. 04/2013, Edition 8
Nitrate DOC316.53.01066

Cadmium Reduction Method Method 8039

®
0.3 to 30.0 mg/L NO3–N (HR) Powder Pillows or AccuVac Ampuls
Scope and application: For water, wastewater and seawater.

Test preparation

Instrument-specific information
The tables in this section show all of the instruments that have the program for this test.
Table 1 shows sample cell and orientation requirements for reagent addition tests, such
as powder pillow or bulk reagent tests. Table 2 shows sample cell and adapter
requirements for AccuVac Ampul tests.
To use either table, select an instrument, then read across to find the corresponding
information for this test.
Table 1 Instrument-specific information for powder pillows
Instrument Sample cell orientation Sample cell
DR 6000 The fill line is to the right. 2495402
DR 3800
DR 2800
DR 2700
DR 5000 The fill line is toward the user.
DR 3900
DR 900 The orientation mark is toward the user. 2401906

Table 2 Instrument-specific information for AccuVac Ampuls

Instrument Adapter Sample cell
DR 6000 — 2427606
DR 5000
DR 900
DR 3900 LZV846 (A)
DR 3800 LZV584 (C) 2122800
DR 2800
DR 2700

Before starting
Install the instrument cap on the DR 900 cell holder before ZERO or READ is pushed.
For best results, measure the reagent blank value for each new lot of reagent. Replace the sample with deionized water in
the test procedure to get the reagent blank value. Subtract the reagent blank value from the sample results automatically
with the reagent blank adjust option.

1
This method is technique-sensitive. Shaking time and technique influence the color development. For most accurate results,
use a standard solution that is within the test range and run the test several times. Increase or decrease the shaking time to
get the expected result. Use the adjusted shaking time for sample measurements.
The reagents that are used in this test contain cadmium. Rinse the sample cell immediately after use to remove all cadmium
particles. Collect the reacted samples for proper disposal.
A deposit of unoxidized metal will remain at the bottom of the sample cell after the reagent dissolves. The deposit will not
affect results.
Review the Safety Data Sheets (MSDS/SDS) for the chemicals that are used and use any recommended personal protective
equipment.
Dispose of reacted solutions according to local, state and federal regulations. Use the Safety Data Sheets for disposal
information for unused reagents. Consult the environmental, health and safety staff for your facility and/or local regulatory
agencies for further disposal information.

Items to collect
Powder pillows

Description Quantity
®
NitraVer 5 Nitrate Reagent Powder Pillow, 10-mL 1
Sample cells. (For information about sample cells, adapters or light shields, refer to Instrument-
2
specific information on page 1.)

Refer to Consumables and replacement items on page 8 for reorder information.

AccuVac Ampuls

Description Quantity
® ®
NitraVer 5 Nitrate Reagent AccuVac Ampul 1
Beaker, 50-mL 1
Sample cells (For information about sample cells, adapters or light shields, refer to Instrument-
1
specific information on page 1.)
Stopper for 18-mm tubes and AccuVac Ampuls 1

Refer to Consumables and replacement items on page 8 for reorder information.

Sample collection and storage

• Collect samples in clean glass or plastic bottles.
• Analyze the samples as soon as possible for best results.
• If prompt analysis is not possible, immediately filter and keep the samples at or below
6 °C (43 °F) for a maximum of 48 hours.
• To preserve samples for up to 28 days, adjust the sample pH to 2 or less with
concentrated sulfuric acid (about 2 mL per liter) and keep at or below 6 °C (43 °F).
The test results then include nitrate and nitrite.
• Let the sample temperature increase to room temperature before analysis.
• Before analysis, adjust the pH to 7 with 5.0 N sodium hydroxide standard solution.
• Correct the test result for the dilution from the volume additions.

2 Nitrate, Cadmium Reduction Method (30.0 mg/L)

Powder pillow procedure

CAUTION
Hazardous waste exposure. Prepared samples contain cadmium. Refer to the SDS for safe handling and disposal
instructions. Obey all local and regional disposal regulations.

Start

1. Start program 355 N, 2. Prepare the sample: Fill 3. Add . 4. Start the instrument
Nitrate HR PP. For a sample cell with 10 mL of timer. A 1-minute reaction
information about sample sample. time starts.
cells, adapters or light
shields, refer to Instrument-
specific information
on page 1.
Note: Although the program
name may vary between
instruments, the program
number does not change.

5. Close the sample cell. 6. Start the instrument 7. Prepare the blank: 8. Clean the blank.
Shake the cell vigorously timer. A 5-minute reaction When the second timer
until the timer expires. Some time starts. expires, fill a second sample
powder may not dissolve. An amber color shows if cell with 10 mL of sample.
Undissolved powder will not nitrate is present.
affect results.

Nitrate, Cadmium Reduction Method (30.0 mg/L) 3

 Zero

9. Insert the blank into the 10. Push ZERO. The 11. Clean the prepared 12. Within one minute after
cell holder. display shows 0.0 mg/L sample. the timer expires, insert the
NO3––N. prepared sample into the
cell holder.

Read

13. Push READ. Results

show in mg/L NO3––N.

AccuVac Ampul procedure

CAUTION
Hazardous waste exposure. Prepared samples contain cadmium. Refer to the SDS for safe handling and disposal
instructions. Obey all local and regional disposal regulations.

Start

1. Start program 361 N, 2. Prepare the sample: 3. Tap the bottom of a 4. Start the instrument
Nitrate HR AV. For Collect at least 40 mL of NitraVer 5 Nitrate AccuVac timer. A 1-minute reaction
information about sample sample in a 50-mL beaker. Ampul to dislodge the time starts.
cells, adapters or light powder. Fill the Ampul with
shields, refer to Instrument- sample. Keep the tip
specific information immersed while the Ampul
on page 1. fills completely.
Note: Although the program
name may vary between
instruments, the program
number does not change.

4 Nitrate, Cadmium Reduction Method (30.0 mg/L)

5. Invert the Ampul 48 to 6. Start the instrument 7. Prepare the blank: 8. Clean the blank.
52 times as the timer counts timer. A 5-minute reaction When the second timer
down. time starts. expires, fill a sample cell
Keep the sample still while with 10 mL of sample.
the timer counts down. An
amber color shows if nitrate
is present.

Zero

9. Insert the blank into the 10. Push ZERO. The 11. Clean the AccuVac 12. Within one minute after
cell holder. display shows 0.0 mg/L Ampul. the timer expires, insert the
NO3––N. prepared sample AccuVac
Ampul into the cell holder.

Read

13. Push READ. Results

show in mg/L NO3––N.

Interferences
Interfering substance Interference level
Chloride Chloride concentrations above 100 mg/L cause low results. The test can be used at high chloride
concentrations (seawater) if a calibration is made with standards that have the same chloride
concentration as the samples (refer to Seawater calibration on page 6).
Ferric iron Interferes at all levels

Nitrate, Cadmium Reduction Method (30.0 mg/L) 5

Interfering substance Interference level
Nitrite Interferes at all levels
Compensate for nitrite interference as follows:

1. Add 30-g/L Bromine Water by drops to the sample until a yellow color remains.
2. Add one drop of 30-g/L Phenol Solution to remove the color.
3. Use the test procedure to measure the concentration of the treated sample. Report the results
as total nitrate and nitrite.

Highly buffered samples Can prevent the correct pH adjustment of the sample by the reagents. Sample pretreatment may
or extreme sample pH be necessary.
Strong oxidizing and Interfere at all levels
reducing substances

Seawater calibration
Chloride concentrations above 100 mg/L cause low results. To use this method for
samples with high chloride concentrations, calibrate the instrument with nitrate standard
solutions that contain the same amount of chloride as the samples.
Prepare calibration standards that contain chloride and 1.0, 3.0, 5.0 and 10.0 mg/L nitrate
(as NO3––N) as follows:

1. Prepare 1 liter of chloride water that has the same chloride concentration as the
samples.
a. Weigh the applicable amount of ACS-grade sodium chloride: (chloride
concentration of samples in g/L) x (1.6485) = g of NaCl per liter.
Note: 18.8 g/L is the typical chloride concentration of seawater.
b. Add the sodium chloride to a 1-liter volumetric flask.
c. Dilute to the mark with deionized water. Mix thoroughly. Use this water as the
dilution water to prepare the nitrate standard solutions.
2. Use a pipet to add 1.0, 3.0, 5.0 and 10.0 mL of a 100 mg/L nitrate-nitrogen (NO3––N)
standard solution into four different 100-mL Class A volumetric flasks.
3. Dilute to the mark with the prepared chloride water. Mix thoroughly.
4. Complete the test procedure for each of the standard solutions and for the prepared
chloride water (for a 0-mg/L standard solution).
5. Measure the absorbance of the standard solutions and enter a user calibration into
the instrument.
6. Use the user program to measure samples that contain high concentrations of
chloride.

Accuracy check
Standard additions method (sample spike)
Use the standard additions method (for applicable instruments) to validate the test
procedure, reagents and instrument and to find if there is an interference in the sample.
Items to collect:
• 1000 mg/L Nitrate Nitrogen (NO3––N) Standard Solution
• 100-mL volumetric flask, Class A
• 25-mL volumetric pipet, Class A and pipet filler
• Deionized water
• Pipet, TenSette®, 0.1–1.0 mL and tips

1. Prepare a 250 mg/L nitrate-nitrogen standard solution as follows:

a. Use a pipet to add 25 mL of a 1000 mg/L nitrate nitrogen standard solution into a
100-mL volumetric flask.

6 Nitrate, Cadmium Reduction Method (30.0 mg/L)

 b. Dilute to the mark with deionized water. Mix well. Prepare this solution daily.
2. Use the test procedure to measure the concentration of the sample, then keep the
(unspiked) sample in the instrument.
3. Go to the Standard Additions option in the instrument menu.
4. Select the values for standard concentration, sample volume and spike volumes.
5. Prepare three spiked samples: use the TenSette pipet to add 0.1 mL, 0.2 mL and
0.3 mL of the prepared standard solution, respectively, to three 10-mL portions of
fresh sample. Mix well.
®
Note: For AccuVac Ampuls, add 0.4 mL, 0.8 mL and 1.2 mL of the prepared standard solution
to three 50-mL portions of fresh sample.
6. Use the test procedure to measure the concentration of each of the spiked samples.
Start with the smallest sample spike. Measure each of the spiked samples in the
instrument.
7. Select Graph to compare the expected results to the actual results.
Note: If the actual results are significantly different from the expected results, make sure that
the sample volumes and sample spikes are measured accurately. The sample volumes and
sample spikes that are used should agree with the selections in the standard additions menu. If
the results are not within acceptable limits, the sample may contain an interference.

Standard solution method

Use the standard solution method to validate the test procedure, reagents and
instrument.
Items to collect:
• Nitrate Nitrogen Standard, Solution, 10.0-mg/L NO3––N

1. Use the test procedure to measure the concentration of the standard solution.
2. Compare the expected result to the actual result.
Note: The factory calibration can be adjusted slightly with the standard adjust option so that the
instrument shows the expected value of the standard solution. The adjusted calibration is then
used for all test results. This adjustment can increase the test accuracy when there are slight
variations in the reagents or instruments.

Method performance
The method performance data that follows was derived from laboratory tests that were
measured on a spectrophotometer during ideal test conditions. Users may get different
results under different test conditions.
Program Standard Precision (95% Confidence Sensitivity
Interval) Concentration change per 0.010 Abs change
355 10 mg/L NO3––N 9.3–10.7 mg/L NO3––N 0.3 mg/L at 0 ppm, 0.5 mg/L at 10 ppm, 0.8 mg/L at 30 ppm
NO3––N
361 10 mg/L NO3––N 9.3–10.7 mg/L NO3––N 0.5 mg/L at 0 ppm, 0.6 mg/L at 10 ppm, 0.8 mg/L at 30 ppm
NO3––N

Summary of method
Cadmium metal reduces nitrate in the sample to nitrite. The nitrite ion reacts in an acidic
medium with sulfanilic acid to form an intermediate diazonium salt. The salt couples with
gentisic acid to form an amber colored solution. The measurement wavelength is 500 nm
for spectrophotometers or 520 nm for colorimeters.
Pollution prevention and waste management
Reacted samples contain cadmium and must be disposed of as a hazardous waste.
Dispose of reacted solutions according to local, state and federal regulations.

Nitrate, Cadmium Reduction Method (30.0 mg/L) 7

Consumables and replacement items
Required reagents

Description Quantity/test Unit Item no.

®
NitraVer 5 Nitrate Reagent Powder Pillows, 10-mL 1 100/pkg 2106169
OR
® ®
NitraVer 5 Nitrate Reagent AccuVac Ampul 1 25/pkg 2511025

Required apparatus (powder pillows)

Description Quantity/test Unit Item no.

Stopper, Neoprene, solid, size #1 2 12/pkg 1480801

OR
Stoppers for 18-mm tubes and AccuVac Ampuls 2 6/pkg 173106

Required apparatus (AccuVac)

Description Quantity/test Unit Item no.

Beaker, 50-mL 1 each 50041H

Recommended standards

Description Unit Item no.

Nitrate Nitrogen Standard Solution, 10.0-mg/L NO3--N 500 mL 30749

Nitrate Nitrogen Standard Solution 1000 mg/L NO3--N 500 mL 1279249
Wastewater Influent Standard, Mixed Parameter, for NH3-N, NO3-N, PO4, COD, SO4,
500 mL 2833149
TOC
Water, deionized 4L 27256

Optional reagents and apparatus

Description Unit Item no.

Bromine Water, 30 g/L 29 mL 221120

Cylinder, mixing, 50-mL each 2088641
®
Pipet, TenSette , 0.1–1.0 mL each 1970001
Pipet, Volumetric, Class A, 0.5 mL each 1451534
Pipet, volumetric, Class A, 1.00-mL each 1451535
Pipet, volumetric, Class A, 2-mL each 1451536
Pipet, volumetric, Class A, 3-mL each 1451503
Pipet tips for TenSette Pipet 1970001 50/pkg 2185696
Pipet tips for TenSette Pipet 1970001 1000/pkg 2185628
Phenol Solution, 30-g/L 29 mL 211220
Pipet, volumetric, Class A, 25-mL each 1451540
Pipet filler, safety bulb each 1465100
®
AccuVac Snapper each 2405200
Sodium Hydroxide, 5 N 50 mL 245026

8 Nitrate, Cadmium Reduction Method (30.0 mg/L)

Consumables and replacement items (continued)
Description Unit Item no.

Sulfuric Acid, ACS 500 mL 97949

Flask, volumetric, Class A, 100-mL each 1457442

Nitrate, Cadmium Reduction Method (30.0 mg/L) 9

 FOR TECHNICAL ASSISTANCE, PRICE INFORMATION AND ORDERING: HACH COMPANY
In the U.S.A. – Call toll-free 800-227-4224 WORLD HEADQUARTERS
Outside the U.S.A. – Contact the HACH office or distributor serving you. Telephone: (970) 669-3050
On the Worldwide Web – www.hach.com; E-mail – [email protected] FAX: (970) 669-2932

© Hach Company/Hach Lange GmbH, 1989–2013. All rights reserved. 04/2013, Edition 8
Nitrate DOC316.53.01067

Cadmium Reduction Method Method 8192

0.01 to 0.50 mg/L NO3––N (LR) Powder Pillows
Scope and application: For water, wastewater and seawater.

Test preparation

Instrument specific information

The table in this section shows all of the instruments that have the program for this test.
Table 1 shows sample cell and orientation requirements for reagent addition tests, such
as powder pillow or bulk reagent tests.
To use the table, select an instrument, then read across to find the corresponding
information for this test.
Table 1 Instrument-specific information for powder pillows
Instrument Sample cell orientation Sample cell
DR 6000 The fill line is to the right. 2495402
DR 3800
DR 2800
DR 2700
DR 5000 The fill line is toward the user.
DR 3900
DR 900 The orientation mark is toward the user. 2401906

Before starting
Install the instrument cap on the DR 900 cell holder before ZERO or READ is pushed.
For best results, measure the reagent blank value for each new lot of reagent. Replace the sample with deionized water in
the test procedure to get the reagent blank value. Subtract the reagent blank value from the sample results automatically
with the reagent blank adjust option.
This method is technique-sensitive. Shaking time and technique influence the color development. For most accurate results,
use a standard solution that is within the test range and run the test several times. Increase or decrease the shaking time to
get the expected result. Use the adjusted shaking time for sample measurements.
The reagents that are used in this test contain cadmium. Rinse the sample cell immediately after use to remove all cadmium
particles. Collect the reacted samples for proper disposal.
A deposit of unoxidized metal will remain at the bottom of the sample cell after the reagent dissolves. The deposit will not
affect results.
Review the Safety Data Sheets (MSDS/SDS) for the chemicals that are used and use any recommended personal protective
equipment.
Dispose of reacted solutions according to local, state and federal regulations. Use the Safety Data Sheets for disposal
information for unused reagents. Consult the environmental, health and safety staff for your facility and/or local regulatory
agencies for further disposal information.

1
Items to collect
Description Quantity
®
NitraVer 6 Nitrate Reagent Powder Pillow, 10-mL 1
®
NitriVer 3 Nitrite Reagent Powder Pillow, 10-mL 1
Cylinder, graduated mixing, 25-mL 1
Sample cells (For information about sample cells, adapters or light shields, refer to Instrument
2
specific information on page 1.)

Refer to Consumables and replacement items on page 6 for reorder information.

Sample collection and storage

• Collect samples in clean glass or plastic bottles.
• Analyze the samples as soon as possible for best results.
• If prompt analysis is not possible, immediately filter and keep the samples at or below
6 °C (43 °F) for a maximum of 48 hours.
• To preserve samples for up to 28 days, adjust the sample pH to 2 or less with
concentrated sulfuric acid (about 2 mL per liter) and keep at or below 6 °C (43 °F).
The test results then include nitrate and nitrite.
• Let the sample temperature increase to room temperature before analysis.
• Before analysis, adjust the pH to 7 with 5.0 N sodium hydroxide standard solution.
• Correct the test result for the dilution from the volume additions.

Powder pillow procedure

CAUTION
Hazardous waste exposure. Prepared samples contain cadmium. Refer to the SDS for safe handling and disposal
instructions. Obey all local and regional disposal regulations.

Start

1. Start program 351 N, 2. Fill the mixing cylinder 3. Add the contents of one 4. Start the instrument
Nitrate LR. For information with 15 mL of sample. NitraVer 6 Reagent Powder timer. A 3-minute reaction
about sample cells, Pillow to the cylinder. Close time starts.
adapters or light shields, the cylinder.
refer to Instrument specific
information on page 1.
Note: Although the program
name may vary between
instruments, the program
number does not change.

2 Nitrate, Cadmium Reduction Method (0.50 mg/L)

5. Shake the cylinder 6. When the timer expires, 7. Prepare the sample: 8. Add the contents of one
vigorously during the start the timer again. A 2- When the timer expires, NitriVer 3 Reagent Powder
reaction period. Some minute reaction time starts. carefully pour 10 mL of Pillow to the prepared
powder may not dissolve. sample into a sample cell. sample cell.
Do not transfer cadmium
particles to the sample cell.

9. Start the instrument 10. Close the sample cell. 11. Start the instrument 12. Prepare the blank:
timer. A 30-second reaction Shake the sample cell timer. A 15-minute reaction When the timer expires, fill a
time starts. gently during the 30-second time starts. second sample cell with
timer. A pink color shows if 10 mL of the original
nitrate is present in the sample.
sample.

Zero

13. Clean the blank. 14. Insert the blank into the 15. Push ZERO. The 16. Clean the prepared
cell holder. display shows 0.00 mg/L sample.
NO3––N.

Nitrate, Cadmium Reduction Method (0.50 mg/L) 3

 Read

17. Insert the prepared 18. Push READ. Results

sample into the cell holder. show in mg/L NO3––N.

Interferences
Interfering substance Interference level
Calcium 100 mg/L
Chloride Chloride concentrations above 100 mg/L cause low results. The test can be used at high chloride
concentrations (seawater) if a calibration is made with standards that have the same chloride
concentration as the samples (refer to Seawater calibration on page 4).
Ferric iron Interferes at all levels
Nitrite Interferes at all levels
Compensate for nitrite interference as follows:

1. Add 30-g/L Bromine Water by drops to the sample until a yellow color remains.
2. Add one drop of 30-g/L Phenol Solution to remove the color.
3. Use the test procedure to measure the concentration of the treated sample. Report the
results as total nitrate and nitrite.

Highly buffered samples Can prevent the correct pH adjustment of the sample by the reagents. Sample pretreatment may
or extreme sample pH be necessary.
Strong oxidizing and Interfere at all levels
reducing substances

Seawater calibration
Chloride concentrations above 100 mg/L cause low results. To use this method for
samples with high chloride concentrations, calibrate the instrument with nitrate standard
solutions that contain the same amount of chloride as the samples.
Prepare calibration standards that contain chloride and 1.0, 3.0, 5.0 and 10.0 mg/L nitrate
(as NO3––N) as follows:

1. Prepare 1 liter of chloride water that has the same chloride concentration as the
samples.
a. Weigh the applicable amount of ACS-grade sodium chloride: (chloride
concentration of samples in g/L) x (1.6485) = g of NaCl per liter.
Note: 18.8 g/L is the typical chloride concentration of seawater.
b. Add the sodium chloride to a 1-liter volumetric flask.
c. Dilute to the mark with deionized water. Mix thoroughly. Use this water as the
dilution water to prepare the nitrate standard solutions.
2. Use a pipet to add 1.0, 3.0, 5.0 and 10.0 mL of a 100 mg/L nitrate-nitrogen (NO3––N)
standard solution into four different 100-mL Class A volumetric flasks.
3. Dilute to the mark with the prepared chloride water. Mix thoroughly.
4. Complete the test procedure for each of the standard solutions and for the prepared
chloride water (for a 0-mg/L standard solution).

4 Nitrate, Cadmium Reduction Method (0.50 mg/L)

 5. Measure the absorbance of the standard solutions and enter a user calibration into
the instrument.
6. Use the user program to measure samples that contain high concentrations of
chloride.

Accuracy check
Standard additions method (sample spike)
Use the standard additions method (for applicable instruments) to validate the test
procedure, reagents and instrument and to find if there is an interference in the sample.
Items to collect:
• Nitrate Nitrogen Standard Solution, 100-mg/L NO3––N
• 50-mL volumetric flask, Class A
• 6-mL volumetric pipet, Class A and pipet filler
• Deionized water
• Pipet, TenSette®, 0.1–1.0 mL and tips
• Mixing cylinders, 25 mL (3)

1. Prepare a 12 mg/L nitrate nitrogen standard solution as follows:

a. Use a pipet to add 6.0 mL of a 100 mg/L NO3––N standard solution into a 50-mL
volumetric flask.
b. Dilute to the mark with deionized water. Mix well. Prepare this solution daily.
2. Use the test procedure to measure the concentration of the sample, then keep the
(unspiked) sample in the instrument.
3. Go to the Standard Additions option in the instrument menu.
4. Select the values for standard concentration, sample volume and spike volumes.
5. Prepare three spiked samples: use the TenSette pipet to add 0.1 mL, 0.2 mL and
0.3 mL of the prepared standard solution, respectively, to three 15-mL portions of
fresh sample. Mix well.
6. Use the test procedure to measure the concentration of each of the spiked samples.
Start with the smallest sample spike. Measure each of the spiked samples in the
instrument.
7. Select Graph to compare the expected results to the actual results.
Note: If the actual results are significantly different from the expected results, make sure that
the sample volumes and sample spikes are measured accurately. The sample volumes and
sample spikes that are used should agree with the selections in the standard additions menu. If
the results are not within acceptable limits, the sample may contain an interference.

Standard solution method

Use the standard solution method to validate the test procedure, reagents and
instrument.
Items to collect:
• Nitrate Nitrogen Standard Solution, 10-mg/L NO3––N
• 100-mL volumetric flask, Class A
• 4-mL volumetric pipet, Class A and pipet filler
• Deionized water

1. Prepare a 0.40 mg/L nitrate nitrogen standard solution as follows:

a. Use a pipet to add 4.00 mL of 10 mg/L nitrate nitrogen standard solution into the
volumetric flask.

Nitrate, Cadmium Reduction Method (0.50 mg/L) 5

 b. Dilute to the mark with deionized water. Mix well. Prepare this solution daily.
2. Use the test procedure to measure the concentration of the prepared standard
solution.
3. Compare the expected result to the actual result.
Note: The factory calibration can be adjusted slightly with the standard adjust option so that the
instrument shows the expected value of the standard solution. The adjusted calibration is then
used for all test results. This adjustment can increase the test accuracy when there are slight
variations in the reagents or instruments.

Method performance
The method performance data that follows was derived from laboratory tests that were
measured on a spectrophotometer during ideal test conditions. Users may get different
results under different test conditions.
Program Standard Precision (95% Confidence Interval) Sensitivity
Concentration change per 0.010 Abs change
351 0.40 mg/L NO3––N 0.35–0.45 mg/L NO3––N 0.003 mg/L NO3––N

Summary of method
Cadmium metal reduces nitrate in the sample to nitrite. The nitrite ion reacts in an acidic
medium with sulfanilic acid to form an intermediate diazonium salt. The salt couples with
chromotropic acid to form a pink-colored product. The measurement wavelength is
507 nm for spectrophotometers or 520 nm for colorimeters.
Pollution prevention and waste management
Reacted samples contain cadmium and must be disposed of as a hazardous waste.
Dispose of reacted solutions according to local, state and federal regulations.
Consumables and replacement items
Required reagents

Description Quantity/test Unit Item no.

Nitrate Reagent Set, low range, 10-mL 1 100/pkg 2429800

Includes:
®
NitraVer 6 Nitrate Reagent Powder Pillow, 10-mL 1 100/pkg 2107249
®
NitriVer 3 Nitrite Reagent Powder Pillow, 10-mL 1 100/pkg 2107169

Required apparatus

Description Quantity/test Unit Item no.

Cylinder, graduated mixing, 25 mL with stopper 1 each 2088640

Stoppers for 18-mm tubes and AccuVac Ampuls 2 6/pkg 173106

Recommended standards and apparatus

Description Unit Item no.

Flask, volumetric, Class A, 100-mL each 1457442

Nitrate Nitrogen Standard Solution, 10.0-mg/L NO3--N 500 mL 30749
Nitrate Nitrogen Standard Solution, 100 mg/L NO3--N 500 mL 194749
Water, deionized 4L 27256

6 Nitrate, Cadmium Reduction Method (0.50 mg/L)

Optional reagents and apparatus

Description Unit Item no.

Bromine Water, 30 g/L 29 mL 221120

®
Pipet, TenSette , 0.1–1.0 mL each 1970001
Pipet tips for TenSette Pipet 1970001 50/pkg 2185696
Pipet tips for TenSette Pipet 1970001 1000/pkg 2185628
Pipet, volumetric, Class A, 4.00-mL each 1451504
Flask, volumetric, 50-mL each 1457441
Pipet filler, safety bulb each 1465100
Phenol Solution, 30-g/L 29 mL 211220
Sodium Hydroxide Standard Solution, 5.0 N 1L 245053
Sulfuric Acid, concentrated, ACS 500 mL 97949
Sodium Chloride, ACS 454 g 18201H
®
Pipet, TenSette , 1.0 to 10.0 mL each 1970010
Pipet tips for TenSette Pipet 1970010 50/pkg 2199796

Nitrate, Cadmium Reduction Method (0.50 mg/L) 7

 FOR TECHNICAL ASSISTANCE, PRICE INFORMATION AND ORDERING: HACH COMPANY
In the U.S.A. – Call toll-free 800-227-4224 WORLD HEADQUARTERS
Outside the U.S.A. – Contact the HACH office or distributor serving you. Telephone: (970) 669-3050
On the Worldwide Web – www.hach.com; E-mail – [email protected] FAX: (970) 669-2932

© Hach Company/Hach Lange GmbH, 1989–2013. All rights reserved. 04/2013, Edition 8
Nitrate, MR DOC316.53.01069

Cadmium Reduction Method Method 8171

®
0.1 to 10.0 mg/L NO3––N (MR, spectrophotometers) Powder Pillows or AccuVac
0.2 to 5.0 mg/L NO3––N (MR, colorimeters) Ampuls
Scope and application: For water, wastewater and seawater.

Test preparation

Instrument-specific information
The tables in this section show all of the instruments that have the program for this test.
Table 1 shows sample cell and orientation requirements for reagent addition tests, such
as powder pillow or bulk reagent tests. Table 2 shows sample cell and adapter
requirements for AccuVac Ampul tests.
To use either table, select an instrument, then read across to find the corresponding
information for this test.
Table 1 Instrument-specific information for powder pillows
Instrument Sample cell orientation Sample cell
DR 6000 The fill line is to the right. 2495402
DR 3800
DR 2800
DR 2700
DR 5000 The fill line is toward the user.
DR 3900
DR 900 The orientation mark is toward the user. 2401906

Table 2 Instrument-specific information for AccuVac Ampuls

Instrument Adapter Sample cell
DR 6000 — 2427606
DR 5000
DR 900
DR 3900 LZV846 (A)
DR 3800 LZV584 (C) 2122800
DR 2800
DR 2700

Before starting
Install the instrument cap on the DR 900 cell holder before ZERO or READ is pushed.
For best results, measure the reagent blank value for each new lot of reagent. Replace the sample with deionized water in
the test procedure to get the reagent blank value. Subtract the reagent blank value from the sample results automatically
with the reagent blank adjust option.

1
This method is technique-sensitive. Shaking time and technique influence the color development. For most accurate results,
use a standard solution that is within the test range and run the test several times. Increase or decrease the shaking time to
get the expected result. Use the adjusted shaking time for sample measurements.
The reagents that are used in this test contain cadmium. Rinse the sample cell immediately after use to remove all cadmium
particles. Collect the reacted samples for proper disposal.
A deposit of unoxidized metal will remain at the bottom of the sample cell after the reagent dissolves. The deposit will not
affect results.
Review the Safety Data Sheets (MSDS/SDS) for the chemicals that are used and use any recommended personal protective
equipment.
Dispose of reacted solutions according to local, state and federal regulations. Use the Safety Data Sheets for disposal
information for unused reagents. Consult the environmental, health and safety staff for your facility and/or local regulatory
agencies for further disposal information.

Items to collect
Powder pillows

Description Quantity
®
NitraVer Nitrate 5 Reagent powder pillow, 10-mL 1
Sample cells (For information about sample cells, adapters or light shields, refer to Instrument-
2
specific information on page 1.)
Stopper, Neoprene #1 2

Refer to Consumables and replacement items on page 7 for reorder information.

AccuVac Ampuls

Description Quantity
® ®
NitraVer Nitrate 5 Reagent AccuVac Ampul 1
Beaker, 50 mL 1
Stoppers for 18-mm tubes and AccuVac Ampuls 1
Sample cells (For information about sample cells, adapters or light shields, refer to Instrument-
1
specific information on page 1.)

Refer to Consumables and replacement items on page 7 for reorder information.

Sample collection and storage

• Collect samples in clean glass or plastic bottles.
• Analyze the samples as soon as possible for best results.
• If prompt analysis is not possible, immediately filter and keep the samples at or below
6 °C (43 °F) for a maximum of 48 hours.
• To preserve samples for up to 28 days, adjust the sample pH to 2 or less with
concentrated sulfuric acid (about 2 mL per liter) and keep at or below 6 °C (43 °F).
The test results then include nitrate and nitrite.
• Let the sample temperature increase to room temperature before analysis.
• Before analysis, adjust the pH to 7 with 5.0 N sodium hydroxide standard solution.
• Correct the test result for the dilution from the volume additions.

2 Nitrate, Cadmium Reduction Method (10.0 mg/L)

Powder pillow procedure

Start

1. Start program 353 N, 2. Prepare the sample: Fill 3. Add the contents of one 4. Start the instrument
Nitrate MR PP. For a sample cell with 10 mL of powder pillow to the sample timer. A 1-minute reaction
information about sample sample. cell. time starts.
cells, adapters or light
shields, refer to Instrument-
specific information
on page 1.
Note: Although the program
name may vary between
instruments, the program
number does not change.

5. Close the sample cell. 6. Start the instrument 7. Prepare the blank: Fill a 8. When the timer expires,
Shake the sample cell timer. A 5-minute reaction second sample cell with clean the blank.
vigorously until the timer time starts. 10 mL of sample.
expires. Some solid material An amber color shows if
will not dissolve. nitrate is present.
Undissolved powder will not
affect results.

Zero

9. Insert the blank into the 10. Push ZERO. The 11. Clean the prepared 12. Within 2 minutes after
cell holder. display shows 0.0 mg/L sample. the timer expires, insert the
NO3––N. prepared sample into the
cell holder.

Nitrate, Cadmium Reduction Method (10.0 mg/L) 3

 Read

13. Push READ. Results

show in mg/L NO3––N.

AccuVac Ampul procedure

Start

1. Start program 359 N, 2. Prepare the blank: Fill 3. Prepare the sample: 4. Start the instrument
Nitrate MR AV. For the sample cell with 10 mL Collect at least 40 mL of timer. A 1-minute reaction
information about sample of sample. sample in a 50-mL beaker. time starts.
cells, adapters or light Fill the AccuVac Ampul with
shields, refer to Instrument- sample. Keep the tip
specific information immersed while the Ampul
on page 1. fills completely.
Note: Although the program
name may vary between
instruments, the program
number does not change.

5. Use a stopper to close 6. Start the instrument 7. When the timer expires, 8. Insert the blank into the
the Ampul tip. Invert the timer. A 5-minute reaction clean the blank. cell holder.
Ampul 48–52 times as the time starts.
timer counts down. An amber color shows if
nitrate is present.

4 Nitrate, Cadmium Reduction Method (10.0 mg/L)

 Zero Read

9. Push ZERO. The display 10. Clean the AccuVac 11. Within 2 minutes after 12. Push READ. Results
shows 0.0 mg/L NO3––N. Ampul. the timer expires, insert the show in mg/L NO3––N.
prepared sample AccuVac
Ampul into the cell holder.

Interferences
Interfering substance Interference level
Chloride Chloride concentrations above 100 mg/L cause low results. The test can be used at high chloride
concentrations (seawater) if a calibration is made with standards that have the same chloride
concentration as the samples (refer to Seawater calibration on page 5).
Ferric iron Interferes at all levels
Nitrite Interferes at all levels
Compensate for nitrite interference as follows:

1. Add 30-g/L Bromine Water by drops to the sample until a yellow color remains.
2. Add one drop of 30-g/L Phenol Solution to remove the color.
3. Use the test procedure to measure the concentration of the treated sample. Report the results
as total nitrate and nitrite.

Highly buffered samples Can prevent the correct pH adjustment of the sample by the reagents. Sample pretreatment may
or extreme sample pH be necessary.
Strong oxidizing and Interfere at all levels
reducing substances

Seawater calibration
Chloride concentrations above 100 mg/L cause low results. To use this method for
samples with high chloride concentrations, calibrate the instrument with nitrate standard
solutions that contain the same amount of chloride as the samples.
Prepare calibration standards that contain chloride and 1.0, 3.0, 5.0 and 10.0 mg/L nitrate
(as NO3––N) as follows:

1. Prepare 1 liter of chloride water that has the same chloride concentration as the
samples.
a. Weigh the applicable amount of ACS-grade sodium chloride: (chloride
concentration of samples in g/L) x (1.6485) = g of NaCl per liter.
Note: 18.8 g/L is the typical chloride concentration of seawater.
b. Add the sodium chloride to a 1-liter volumetric flask.
c. Dilute to the mark with deionized water. Mix thoroughly. Use this water as the
dilution water to prepare the nitrate standard solutions.
2. Use a pipet to add 1.0, 3.0, 5.0 and 10.0 mL of a 100 mg/L nitrate-nitrogen (NO3––N)
standard solution into four different 100-mL Class A volumetric flasks.
3. Dilute to the mark with the prepared chloride water. Mix thoroughly.
4. Complete the test procedure for each of the standard solutions and for the prepared
chloride water (for a 0-mg/L standard solution).

Nitrate, Cadmium Reduction Method (10.0 mg/L) 5

 5. Measure the absorbance of the standard solutions and enter a user calibration into
the instrument.
6. Use the user program to measure samples that contain high concentrations of
chloride.

Accuracy check
Standard additions method (sample spike)
Use the standard additions method (for applicable instruments) to validate the test
procedure, reagents and instrument and to find if there is an interference in the sample.
Items to collect:
• Nitrate Nitrogen Standard Solution, 100 mg/L NO3–– N
• Ampule breaker
• Pipet, TenSette®, 0.1–1.0 mL and tips

1. Use the test procedure to measure the concentration of the sample, then keep the
(unspiked) sample in the instrument.
2. Go to the Standard Additions option in the instrument menu.
3. Select the values for standard concentration, sample volume and spike volumes.
4. Open the standard solution.
5. Prepare three spiked samples: use the TenSette pipet to add 0.1 mL, 0.2 mL and
0.3 mL of the standard solution, respectively, to three 10-mL portions of fresh sample.
Mix well.
Note: For AccuVac® Ampuls, add 0.1 mL, 0.2 mL and 0.3 mL of a 500 mg/L NO3––N standard
solution to three 50-mL portions of fresh sample.
6. Use the test procedure to measure the concentration of each of the spiked samples.
Start with the smallest sample spike. Measure each of the spiked samples in the
instrument.
7. Select Graph to compare the expected results to the actual results.
Note: If the actual results are significantly different from the expected results, make sure that
the sample volumes and sample spikes are measured accurately. The sample volumes and
sample spikes that are used should agree with the selections in the standard additions menu. If
the results are not within acceptable limits, the sample may contain an interference.

Standard solution method

Use the standard solution method to validate the test procedure, reagents and
instrument.
Items to collect:
• Nitrate Nitrogen Standard Solution, 100 mg/L NO3–– N
• 100-mL volumetric flask, Class A
• 5-mL volumetric pipet, Class A and pipet filler
• Deionized water

1. Prepare a 5.0 mg/L nitrate-nitrogen standard solution as follows:

a. Use a pipet to add 5.0 mL of 100 mg/L nitrate-nitrogen standard solution into the
volumetric flask.
b. Dilute to the mark with deionized water. Mix well. Prepare this solution daily.
2. Use the test procedure to measure the concentration of the prepared standard
solution.
3. Compare the expected result to the actual result.
Note: The factory calibration can be adjusted slightly with the standard adjust option so that the
instrument shows the expected value of the standard solution. The adjusted calibration is then
used for all test results. This adjustment can increase the test accuracy when there are slight
variations in the reagents or instruments.

6 Nitrate, Cadmium Reduction Method (10.0 mg/L)

Method performance
The method performance data that follows was derived from laboratory tests that were
measured on a spectrophotometer during ideal test conditions. Users may get different
results under different test conditions.
Program Standard Precision (95% Confidence Interval) Sensitivity
Concentration change per 0.010 Abs change
353 5.0 mg/L NO3––N 4.8–5.2 mg/L NO3––N 0.04 mg/L NO3––N
359 5.0 mg/L NO3––N 4.6–5.4 mg/L NO3––N 0.05 mg/L NO3––N

Summary of method
Cadmium metal reduces nitrate in the sample to nitrite. The nitrite ion reacts in an acidic
medium with sulfanilic acid to form an intermediate diazonium salt. The salt couples with
gentisic acid to form an amber colored solution. The measurement wavelength is 400 nm
for spectrophotometers or 420 nm for colorimeters.
Pollution prevention and waste management
Reacted samples contain cadmium and must be disposed of as a hazardous waste.
Dispose of reacted solutions according to local, state and federal regulations.
Consumables and replacement items
Required reagents

Description Quantity/test Unit Item no.

®
NitraVer 5 Nitrate Reagent Powder Pillows, 10-mL 1 100/pkg 2106169
OR
® ®
NitraVer 5 Nitrate Reagent AccuVac Ampul 1 25/pkg 2511025

Required apparatus for powder pillows

Description Quantity/test Unit Item no.

Stopper, Neoprene, solid, size #2 2 12/pkg 1480802

Required apparatus for AccuVac Ampuls

Description Quantity/test Unit Item no.

Beaker, 50-mL 1 each 50041H

Stoppers for 18-mm tubes and AccuVac Ampuls 2 6/pkg 173106

Recommended standards

Description Unit Item no.

Drinking Water Standard, Mixed Parameter, Inorganic for F-, NO3, PO4, SO4 500 mL 2833049
Nitrate Nitrogen Standard Solution, 100-mg/L NH3-N 500 mL 194749
Nitrate Nitrogen Standard Solution, Voluette® Ampule, 500-mg/L NO3–N 16/pkg 1426010

Optional reagents and apparatus

Description Unit Item no.

Bromine Water, 30 g/L 29 mL 221120

Cylinder, mixing, 50-mL each 2088641

Nitrate, Cadmium Reduction Method (10.0 mg/L) 7

Consumables and replacement items (continued)
Description Unit Item no.

Flask, volumetric, Class A, 100-mL each 1457442

®
Pipet, TenSette , 0.1–1.0 mL each 1970001
Pipet tips for TenSette Pipet 1970001 50/pkg 2185696
Pipet tips for TenSette Pipet 1970001 1000/pkg 2185628
Pipet, volumetric 5.00-mL each 1451537
Pipet filler, safety bulb each 1465100
Phenol Solution, 30-g/L 29 mL 211220
Sodium Hydroxide Standard Solution, 5.0 N 1L 245053
Sulfuric Acid, concentrated, ACS 500 mL 97949
Water, deionized 4L 27256

FOR TECHNICAL ASSISTANCE, PRICE INFORMATION AND ORDERING: HACH COMPANY

In the U.S.A. – Call toll-free 800-227-4224 WORLD HEADQUARTERS
Outside the U.S.A. – Contact the HACH office or distributor serving you. Telephone: (970) 669-3050
On the Worldwide Web – www.hach.com; E-mail – [email protected] FAX: (970) 669-2932

© Hach Company/Hach Lange GmbH, 1989–2013. All rights reserved. 04/2013, Edition 8
Nitrate, HR DOC316.53.01068

Chromotropic Acid Method Method 10020

0.2 to 30.0 mg/L NO3––N (HR) Test ‘N Tube™ Vials
Scope and application: For water and wastewater.

Test preparation

Instrument specific information

The table in this section shows all of the instruments that have the program for this test.
Table 1 shows adapter and light shield requirements for the instruments that use them.
To use the table, select an instrument, then read across to find the corresponding
information for this test.
Table 1 Instrument-specific information for test tubes
Instrument Adapters Light shield
DR 6000 — —
DR 5000 — —
DR 900 4846400 Cover supplied with the instrument
DR 3900 — LZV849
DR 3800 — LZV646
DR 2800 —

Before starting
Install the instrument cap on the DR 900 cell holder before ZERO or READ is pushed.
DR 3900, DR 3800, DR 2800 and DR 2700: Install the light shield in Cell Compartment #2 before this test is started.
The vials must be mixed carefully for accurate results. Start each vial inversion with the vial in the vertical position, with the
cap on the top. Turn the vial upside-down and wait for all of the solution to flow down to the cap. Return the vial to the
vertical position and wait for all of the solution to flow down to the bottom of the vial. This mixing method equals one
inversion.
For best results, measure the reagent blank value for each new lot of reagent. Replace the sample with deionized water in
the test procedure to get the reagent blank value. Subtract the reagent blank value from the sample results automatically
with the reagent blank adjust option.
Review the Safety Data Sheets (MSDS/SDS) for the chemicals that are used and use any recommended personal protective
equipment.
Dispose of reacted solutions according to local, state and federal regulations. Use the Safety Data Sheets for disposal
information for unused reagents. Consult the environmental, health and safety staff for your facility and/or local regulatory
agencies for further disposal information.

Items to collect
Description Quantity

Light shield (For information about sample cells, adapters or light shields, refer to Instrument
1
specific information on page 1.)
®
NitraVer X Nitrate Reagent Set, Test 'N Tube™ 1
Funnel, micro, poly 1

1
Items to collect (continued)
Description Quantity
®
Pipet, TenSette , 0.1- to 1.0-mL 1
Test tube rack 1

Refer to Consumables and replacement items on page 4 for reorder information.

Sample collection and storage

• Collect samples in clean glass or plastic bottles.
• Analyze the samples as soon as possible for best results.
• If prompt analysis is not possible, immediately filter and keep the samples at or below
6 °C (43 °F) for a maximum of 48 hours.
• To preserve samples for up to 28 days, adjust the sample pH to 2 or less with
concentrated sulfuric acid (about 2 mL per liter) and keep at or below 6 °C (43 °F).
The test results then include nitrate and nitrite.
• Let the sample temperature increase to room temperature before analysis.
• Before analysis, adjust the pH to 7 with 5.0 N sodium hydroxide standard solution.
• Correct the test result for the dilution from the volume additions.

TNT procedure

Start

1. Start program 344 N, 2. Prepare the blank: Use 3. Close the vial. Invert the 4. Clean the blank vial.
Nitrate HR, TNT. For a pipet to add 1.00 mL of vial 10 times to mix.
information about sample sample to a NitraVer X
cells, adapters or light Reagent A Test 'N Tube
shields, refer to Instrument vial.
specific information
on page 1.
Note: Although the program
name may vary between
instruments, the program
number does not change.

2 Nitrate, Chromotropic Acid TNT Method (30.0 mg/L)

 Zero

5. Insert the vial into the cell 6. Push ZERO. The display 7. Remove the TNT vial 8. Prepare the sample:
holder. shows 0.0 mg/L NO3––N. from the cell holder. Use a funnel to add the
contents of one NitraVer X
Reagent B Powder Pillow to
the vial.

Read

9. Close the vial. Invert the 10. Start the instrument 11. Within 5 minutes after 12. Push READ. Results
vial 10 times to mix. timer. A 5-minute reaction the timer expires, clean the show in mg/L NO3––N.
Some solid matter will not time starts. vial. Insert the vial into the
dissolve. Do not invert the vial again. cell holder.
A yellow color shows if
nitrate is present.

Interferences
Interfering substance Interference level
Barium Negative interference at more than 1 mg/L.
Chloride Does not interfere at less than 1000 mg/L.
Nitrite A positive interference at concentrations more than 12 mg/L. To remove nitrite interference up to
100 mg/L, add 400 mg (one full 0.5 g measuring spoon) of urea to 10 mL of sample. Swirl to
dissolve. Proceed with the nitrate test as usual.
Copper Positive interference at all levels.

Accuracy check
Standard additions method (sample spike)
Use the standard additions method (for applicable instruments) to validate the test
procedure, reagents and instrument and to find if there is an interference in the sample.
Items to collect:
• 500 mg/L Nitrate Nitrogen Standard Solution, Voluette® Ampule
• Ampule breaker
• Pipet, TenSette®, 0.1–1.0 mL and tips

Nitrate, Chromotropic Acid TNT Method (30.0 mg/L) 3

 • 25-mL mixing cylinders (3)

1. Use the test procedure to measure the concentration of the sample, then keep the
(unspiked) sample in the instrument.
2. Go to the Standard Additions option in the instrument menu.
3. Select the values for standard concentration, sample volume and spike volumes.
4. Open the standard solution.
5. Prepare three spiked samples: use the TenSette pipet to add 0.1 mL, 0.2 mL and
0.3 mL of the standard solution, respectively, to three 25-mL portions of fresh sample.
Mix well.
6. Use the test procedure to measure the concentration of each of the spiked samples.
Start with the smallest sample spike. Measure each of the spiked samples in the
instrument.
7. Select Graph to compare the expected results to the actual results.
Note: If the actual results are significantly different from the expected results, make sure that
the sample volumes and sample spikes are measured accurately. The sample volumes and
sample spikes that are used should agree with the selections in the standard additions menu. If
the results are not within acceptable limits, the sample may contain an interference.

Standard solution method

Use the standard solution method to validate the test procedure, reagents and
instrument.
Items to collect:
• 10.0 mg/L Nitrate Nitrogen Standard Solution

1. Use the test procedure to measure the concentration of the standard solution.
2. Compare the expected result to the actual result.
Note: The factory calibration can be adjusted slightly with the standard adjust option so that the
instrument shows the expected value of the standard solution. The adjusted calibration is then
used for all test results. This adjustment can increase the test accuracy when there are slight
variations in the reagents or instruments.

Method performance
The method performance data that follows was derived from laboratory tests that were
measured on a spectrophotometer during ideal test conditions. Users may get different
results under different test conditions.
Program Standard Precision (95% Confidence Interval) Sensitivity
Concentration change per 0.010 Abs change
344 10.0 mg/L NO3––N 9.5–10.5 mg/L NO3––N 0.2 mg/L NO3––N

Summary of method
Nitrate in the sample reacts with chromotropic acid under strongly acidic conditions to
yield a yellow product. The measurement wavelength is 410 nm for spectrophotometers
or 420 nm for colorimeters.
Consumables and replacement items
Required reagents

Description Quantity/test Unit Item no.

®
NitraVer X Nitrate Reagent Set, Test 'N Tube™ — 50 tests 2605345

4 Nitrate, Chromotropic Acid TNT Method (30.0 mg/L)

Required apparatus

Description Quantity/test Unit Item no.

Funnel, micro, poly 1 each 2584335

®
Pipet, TenSette , 0.1- to 1.0-mL 1 each 1970001
Pipet Tips, for TenSette Pipet 1970001 2 50/pkg 2185696
Test tube rack 1 each 1864100

Recommended standards

Description Unit Item no.

Nitrate Nitrogen Standard Solution, 10.0-mg/L NO3--N 500 mL 30749

Nitrate Nitrogen Standard Solution, Voluette® Ampule, 500-mg/L NO3–N 16/pkg 1426010
Wastewater Influent Standard, Mixed Parameter, for NH3-N, NO3-N, PO4, COD, SO4,
500 mL 2833149
TOC
Water, deionized 4L 27256

Optional reagents and apparatus

Description Unit Item no.

®
Ampule Breaker, Voluette ampules each 2196800
Cylinder, mixing, 25-mL each 2088640
Pipet tips for TenSette Pipet 1970001 1000/pkg 2185628
Sodium Hydroxide, 5 N 50 mL 245026
Spoon, measuring, 0.5-g each 90700
Sulfuric Acid, ACS 500 mL 97949
Urea, ACS grade 100 g 1123726

Nitrate, Chromotropic Acid TNT Method (30.0 mg/L) 5

 FOR TECHNICAL ASSISTANCE, PRICE INFORMATION AND ORDERING: HACH COMPANY
In the U.S.A. – Call toll-free 800-227-4224 WORLD HEADQUARTERS
Outside the U.S.A. – Contact the HACH office or distributor serving you. Telephone: (970) 669-3050
On the Worldwide Web – www.hach.com; E-mail – [email protected] FAX: (970) 669-2932

© Hach Company/Hach Lange GmbH, 1989–2013. All rights reserved. 04/2013, Edition 8
Nitrite DOC316.53.01074

USEPA Diazotization Method1 Method 8507

®
0.002 to 0.300 mg/L NO2––N (LR, spectrophotometers) Powder Pillows or AccuVac
0.005 to 0.350 mg/L NO2––N (LR, colorimeters) Ampuls
Scope and application: For water, wastewater and seawater.
1 USEPA approved for wastewater analysis, Federal Register, 44(85), 25505 (May 1, 1979).

Test preparation

Instrument-specific information
The tables in this section show all of the instruments that have the program for this test.
Table 1 shows sample cell and orientation requirements for reagent addition tests, such
as powder pillow or bulk reagent tests. Table 2 shows sample cell and adapter
requirements for AccuVac Ampul tests.
To use either table, select an instrument, then read across to find the corresponding
information for this test.
Table 1 Instrument-specific information for powder pillows
Instrument Sample cell orientation Sample cell
DR 6000 The fill line is to the right. 2495402
DR 3800
DR 2800
DR 2700
DR 5000 The fill line is toward the user.
DR 3900
DR 900 The orientation mark is toward the user. 2401906

Table 2 Instrument-specific information for AccuVac Ampuls

Instrument Adapter Sample cell
DR 6000 — 2427606
DR 5000
DR 900
DR 3900 LZV846 (A)
DR 3800 LZV584 (C) 2122800
DR 2800
DR 2700

1
Before starting
Install the instrument cap on the DR 900 cell holder before ZERO or READ is pushed.
For best results, measure the reagent blank value for each new lot of reagent. Replace the sample with deionized water in
the test procedure to get the reagent blank value. Subtract the reagent blank value from the sample results automatically
with the reagent blank adjust option.
Review the Safety Data Sheets (MSDS/SDS) for the chemicals that are used and use any recommended personal protective
equipment.
Dispose of reacted solutions according to local, state and federal regulations. Use the Safety Data Sheets for disposal
information for unused reagents. Consult the environmental, health and safety staff for your facility and/or local regulatory
agencies for further disposal information.

Items to collect
Powder pillows

Description Quantity
®
NitriVer 3 Reagent Powder Pillows, 10-mL 1
Sample cells. (For information about sample cells, adapters or light shields, refer to Instrument-
2
specific information on page 1.)

Refer to Consumables and replacement items on page 6 for reorder information.

AccuVac Ampuls

Description Quantity
® ®
NitriVer 3 Reagent AccuVac Ampuls 1
Beaker, 50-mL 1
Sample cells (For information about sample cells, adapters or light shields, refer to Instrument-
1
specific information on page 1.)
Stopper for 18-mm tubes and AccuVac Ampuls 1

Refer to Consumables and replacement items on page 6 for reorder information.

Sample collection and storage

• Collect samples in clean glass or plastic bottles.
• To preserve samples for later analysis, keep the samples at or below 6 °C (43 °F) for
up to 48 hours.
• Let the sample temperature increase to room temperature before analysis.

2 Nitrite, Diazotization Method (0.300 mg/L)

Powder pillow procedure

Start

1. Start program 371 N, 2. Prepare the sample: Fill 3. Add the contents of one 4. Swirl to mix.
Nitrite LR PP. For a sample cell with 10 mL of NitriVer 3 Reagent Powder A pink color shows if nitrite
information about sample sample. Pillow. is present in the sample.
cells, adapters or light
shields, refer to Instrument-
specific information
on page 1.
Note: Although the program
name may vary between
instruments, the program
number does not change.

5. Start the instrument 6. Prepare the blank: 7. Clean the blank. 8. Insert the blank into the
timer. A 20-minute reaction When the timer expires, fill a cell holder.
time starts. second sample cell with
10 mL of sample.

Zero Read

9. Push ZERO. The display 10. Clean the prepared 11. Insert the prepared 12. Push READ. Results
shows 0.000 mg/L NO2––N. sample. sample into the cell holder. show in mg/L NO2––N.

Nitrite, Diazotization Method (0.300 mg/L) 3

AccuVac Ampul procedure

Start

1. Start program 375 N, 2. Prepare the sample: 3. Quickly invert the Ampul 4. Start the instrument
Nitrite LR AV. For Collect at least 40 mL of several times to mix. timer. A 20-minute reaction
information about sample sample in a 50-mL beaker. A pink color shows if nitrite time starts.
cells, adapters or light Fill the AccuVac Ampul with is present in the sample.
shields, refer to Instrument- sample. Keep the tip
specific information immersed while the Ampul
on page 1. fills completely.
Note: Although the program
name may vary between
instruments, the program
number does not change.

Zero

5. Prepare the blank: 6. Clean the blank. 7. Insert the blank into the 8. Push ZERO. The display
When the timer expires, fill a cell holder. shows 0.000 mg/L NO2––N.
sample cell with 10 mL of
sample.

Read

9. Clean the AccuVac 10. Insert the prepared 11. Push READ. Results
Ampul. sample AccuVac Ampul into show in mg/L NO2––N.
the cell holder.

Interferences
Interfering substance Interference level
Antimonous ions Interfere by causing precipitation
Auric ions Interfere by causing precipitation

4 Nitrite, Diazotization Method (0.300 mg/L)

Interfering substance Interference level
Bismuth ions Interfere by causing precipitation
Chloroplatinate ions Interfere by causing precipitation
Cupric ions Cause low results
Ferric ions Interfere by causing precipitation
Ferrous ions Cause low results
Lead ions Interfere by causing precipitation
Mercurous ions Interfere by causing precipitation
Metavanadate ions Interfere by causing precipitation
Nitrate Very high levels of nitrate (>100 mg/L nitrate as N) appear to undergo a slight
amount of reduction to nitrite, either spontaneously or during the course of the
test. A small amount of nitrite will be found at these levels.
Silver ions Interfere by causing precipitation
Strong oxidizing and reducing substances Interfere at all levels

Accuracy check
Standard solution method
Use the standard solution method to validate the test procedure, reagents and
instrument.
Items to collect:
• 0.150 mg/L NO2––N standard solution (Nitrite standard solutions are difficult to
prepare. Use the instructions in Standard Methods for the Examination of Water and
Wastewater, Method 4500—NO2-B)

1. Use the test procedure to measure the concentration of the standard solution.
2. Compare the expected result to the actual result.
Note: The factory calibration can be adjusted slightly with the standard adjust option so that the
instrument shows the expected value of the standard solution. The adjusted calibration is then
used for all test results. This adjustment can increase the test accuracy when there are slight
variations in the reagents or instruments.

Method performance
The method performance data that follows was derived from laboratory tests that were
measured on a spectrophotometer during ideal test conditions. Users may get different
results under different test conditions.
Program Standard Precision (95% Confidence Interval) Sensitivity
Concentration change per 0.010 Abs change
371 0.150 mg/L NO2––N 0.147–0.153 mg/L NO2––N 0.002 mg/L NO2––N
375 0.150 mg/L NO2––N 0.147–0.153 mg/L NO2––N 0.002 mg/L NO2––N

Summary of method
Nitrite in the sample reacts with sulfanilic acid to form an intermediate diazonium salt.
This couples with chromotropic acid to produce a pink colored complex directly
proportional to the amount of nitrite present. The measurement wavelength is 507 nm for
spectrophotometers or 520 nm for colorimeters.

Nitrite, Diazotization Method (0.300 mg/L) 5

Consumables and replacement items
Required reagents

Description Quantity/test Unit Item no.

®
NitriVer 3 Nitrite Reagent Powder Pillow, 10-mL 1 100/pkg 2107169
OR
® ®
NitriVer 3 Nitrite Reagent AccuVac Ampul 1 25/pkg 2512025

Required apparatus

Description Quantity/test Unit Item no.

Beaker, 50-mL 1 each 50041H

Recommended standards, reagents and apparatus

Description Unit Item no.

Balance, analytical, 80 g x 0.1 mg 100-240 VAC each 2936701

®
AccuVac vials for sample blanks 25/pkg 2677925
®
AccuVac Snapper each 2405200
®
AccuVac Drainer each 4103600
Standard Methods Book, most current edition each 2270800
Sodium Nitrite, ACS 454 g 245201
Water, deionized 4L 27256

FOR TECHNICAL ASSISTANCE, PRICE INFORMATION AND ORDERING: HACH COMPANY

In the U.S.A. – Call toll-free 800-227-4224 WORLD HEADQUARTERS
Outside the U.S.A. – Contact the HACH office or distributor serving you. Telephone: (970) 669-3050
On the Worldwide Web – www.hach.com; E-mail – [email protected] FAX: (970) 669-2932

© Hach Company/Hach Lange GmbH, 1989–2013. All rights reserved. 04/2013, Edition 8
Nitrite DOC316.53.01073

Diazotization Method Method 10019

0.003 to 0.500 mg/L NO2––N (LR) Test ‘N Tube™ Vials
Scope and application: For water, wastewater and seawater.

Test preparation

Instrument specific information

The table in this section shows all of the instruments that have the program for this test.
Table 1 shows adapter and light shield requirements for the instruments that use them.
To use the table, select an instrument, then read across to find the corresponding
information for this test.
Table 1 Instrument-specific information for test tubes
Instrument Adapters Light shield
DR 6000 — —
DR 5000 — —
DR 900 4846400 Cover supplied with the instrument
DR 3900 — LZV849
DR 3800 — LZV646
DR 2800 —

Before starting
Install the instrument cap on the DR 900 cell holder before ZERO or READ is pushed.
DR 3900, DR 3800, DR 2800 and DR 2700: Install the light shield in Cell Compartment #2 before this test is started.
For best results, measure the reagent blank value for each new lot of reagent. Replace the sample with deionized water in
the test procedure to get the reagent blank value. Subtract the reagent blank value from the sample results automatically
with the reagent blank adjust option.
Review the Safety Data Sheets (MSDS/SDS) for the chemicals that are used and use any recommended personal protective
equipment.
Dispose of reacted solutions according to local, state and federal regulations. Use the Safety Data Sheets for disposal
information for unused reagents. Consult the environmental, health and safety staff for your facility and/or local regulatory
agencies for further disposal information.

Items to collect
Description Quantity

Light shield (For information about sample cells, adapters or light shields, refer to Instrument
1
specific information on page 1.)
®
Test 'N Tube™ NitriVer 3 Nitrite Reagent Set 1
®
Pipet, TenSette 1.0 to 10.0 mL with tips varies

Refer to Consumables and replacement items on page 4 for reorder information.

Sample collection and storage

• Collect samples in clean glass or plastic bottles.

1
 • To preserve samples for later analysis, keep the samples at or below 6 °C (43 °F) for
up to 48 hours.
• Let the sample temperature increase to room temperature before analysis.

Test 'N Tube procedure

Start

1. Start program 345 N, 2. Prepare the sample: Fill 3. Put the cap on the vial. 4. Start the instrument
Nitrite LR TNT. For a Test 'N Tube NitriVer Shake to dissolve the timer. A 20-minute reaction
information about sample 3 Nitrite vial with 5 mL of powder. A pink color shows time starts.
cells, adapters or light sample. if nitrite-nitrogen is present
shields, refer to Instrument in the sample .
specific information
on page 1.
Note: Although the program
name may vary between
instruments, the program
number does not change.

Zero

5. Prepare the blank: 6. Clean the blank vial. 7. Insert the blank vial into 8. Push ZERO. The display
When the timer expires, fill the 16-mm cell holder. shows 0.000 mg/L NO2––N.
and empty Test 'N Tube vial
with 5 mL of sample.

Read

9. Clean the sample vial. 10. Insert the sample vial 11. Push READ. Results
into the 16-mm cell holder. show in mg/L NO2––N.

2 Nitrite, Diazotization TNT Method (0.500 mg/L)

Interferences
Interfering substance Interference level
Antimonous ions Interfere by causing precipitation
Auric ions Interfere by causing precipitation
Bismuth ions Interfere by causing precipitation
Chloroplatinate ions Interfere by causing precipitation
Cupric ions Cause low results
Ferric ions Interfere by causing precipitation
Ferrous ions Cause low results
Lead ions Interfere by causing precipitation
Mercurous ions Interfere by causing precipitation
Metavanadate ions Interfere by causing precipitation
Nitrate Very high levels of nitrate (>100 mg/L nitrate as N) appear to undergo a slight
amount of reduction to nitrite, either spontaneously or during the course of the
test. A small amount of nitrite will be found at these levels.
Silver ions Interfere by causing precipitation
Strong oxidizing and reducing substances Interfere at all levels

Accuracy check
Standard solution method
Use the standard solution method to validate the test procedure, reagents and
instrument.
Items to collect:
• 0.300 mg/L NO2––N standard solution (Nitrite standard solutions are difficult to
prepare. Use the instructions in Standard Methods for the Examination of Water and
Wastewater, Method 4500—NO2-B)

1. Use the test procedure to measure the concentration of the standard solution.
2. Compare the expected result to the actual result.
Note: The factory calibration can be adjusted slightly with the standard adjust option so that the
instrument shows the expected value of the standard solution. The adjusted calibration is then
used for all test results. This adjustment can increase the test accuracy when there are slight
variations in the reagents or instruments.

Method performance
The method performance data that follows was derived from laboratory tests that were
measured on a spectrophotometer during ideal test conditions. Users may get different
results under different test conditions.
Program Standard Precision (95% Confidence Interval) Sensitivity
Concentration change per 0.010 Abs change
345 0.300 mg/L NO2––N 0.294–0.306 mg/L NO2––N 0.003 mg/L NO2––N

Summary of method
Nitrite in the sample reacts with sulfanilic acid to form an intermediate diazonium salt.
This couples with chromotropic acid to produce a pink colored complex directly
proportional to the amount of nitrite present. The measurement wavelength is 507 nm for
spectrophotometers or 520 nm for colorimeters.

Nitrite, Diazotization TNT Method (0.500 mg/L) 3

Consumables and replacement items
Required reagents

Description Quantity/test Unit Item no.

®
NitriVer 3 Nitrite Reagent Set, Test 'N Tube™ 1 50/pkg 2608345

Required apparatus

Description Quantity/test Unit Item no.

®
Pipet, TenSette , 1.0- to 10.0-mL 1 each 1970010
Pipet Tips, for TenSette Pipet 1970010 varies 50/pkg 2199796

Recommended standards, reagents and apparatus

Description Unit Item no.

Standard Methods Book, most current edition each 2270800

Balance, analytical, 80 g x 0.1 mg 100-240 VAC each 2936701
Pipet tips for TenSette Pipet 1970010 250/pkg 2199725
Sodium Nitrite, ACS 454 g 245201
Water, deionized 4L 27256

FOR TECHNICAL ASSISTANCE, PRICE INFORMATION AND ORDERING: HACH COMPANY

In the U.S.A. – Call toll-free 800-227-4224 WORLD HEADQUARTERS
Outside the U.S.A. – Contact the HACH office or distributor serving you. Telephone: (970) 669-3050
On the Worldwide Web – www.hach.com; E-mail – [email protected] FAX: (970) 669-2932

© Hach Company/Hach Lange GmbH, 1989–2013. All rights reserved. 04/2013, Edition 8
Nitrite DOC316.53.01075

Ferrous Sulfate Method1 Method 8153

2 to 250 mg/L NO2– (HR, spectrophotometers) Powder Pillows
2 to 150 mg/L NO2– (HR, colorimeters)™
Scope and application: For cooling systems.
1 Adapted from McAlpine, R. and Soule, B., Qualitative Chemical Analysis, New York, 476, 575 (1933).

Test preparation

Instrument specific information

The table in this section shows all of the instruments that have the program for this test.
Table 1 shows sample cell and orientation requirements for reagent addition tests, such
as powder pillow or bulk reagent tests.
To use the table, select an instrument, then read across to find the corresponding
information for this test.
Table 1 Instrument-specific information for powder pillow
Instrument Sample cell orientation Sample cell
DR 6000 The fill line is to the right. 2495402
DR 3800
DR 2800
DR 2700
DR 5000 The fill line is toward the user.
DR 3900
DR 900 The orientation mark is toward the user. 2401906

Before starting
Install the instrument cap on the DR 900 cell holder before ZERO or READ is pushed.
For best results, measure the reagent blank value for each new lot of reagent. Replace the sample with deionized water in
the test procedure to get the reagent blank value. Subtract the reagent blank value from the sample results automatically
with the reagent blank adjust option.
Review the Safety Data Sheets (MSDS/SDS) for the chemicals that are used and use any recommended personal protective
equipment.
Dispose of reacted solutions according to local, state and federal regulations. Use the Safety Data Sheets for disposal
information for unused reagents. Consult the environmental, health and safety staff for your facility and/or local regulatory
agencies for further disposal information.

Items to collect
Description Quantity
®
NitriVer 2 Nitrite Reagent Powder Pillows, 10-mL 1
Deionized water varies

1
Items to collect (continued)
Description Quantity

Stopper, Neoprene, solid #1 2

Sample cells (For information about sample cells, adapters or light shields, refer to Instrument
2
specific information on page 1.)

Refer to Consumables and replacement items on page 4 for reorder information.

Sample collection and storage

• Collect samples in clean glass or plastic bottles.
• To preserve samples for later analysis, keep the samples at or below 6 °C (43 °F) for
up to 48 hours.
• Let the sample temperature increase to room temperature before analysis.

Powder pillow procedure

Start

1. Start program 373 N, 2. Prepare the sample: Fill 3. Add the contents of one 4. Close the sample cell.
Nitrite HR PP. For a sample cell with 10 mL of NitriVer 2 Nitrite Reagent Shake to dissolve the
information about sample sample. Powder Pillow. A greenish- reagent.
cells, adapters or light brown color starts to show if
shields, refer to Instrument nitrite is present in the
specific information sample.
on page 1.
Note: Although the program
name may vary between
instruments, the program
number does not change.

5. Start the instrument 6. Prepare the blank: Fill a 7. Clean the blank. 8. Insert the blank into the
timer. A 10-minute reaction second sample cell with cell holder.
time starts. 10 mL of sample.
To prevent low results,
leave the sample cell on a
flat surface. Do not move
or disturb the sample cell
during the reaction period.

2 Nitrite, Ferrous Sulfate Method (250 mg/L)

 Zero

9. Push ZERO. The display 10. After the timer expires, 11. Clean the prepared 12. Insert the prepared
shows 0 mg/L NO2–. gently invert the prepared sample. sample into the cell holder.
sample two times.
Excessive mixing causes
low results..

Read

13. Push READ. Results

show in mg/L NO2–.

Interferences
This test does not measure nitrates nor is it applicable to glycol-based samples. Dilute
glycol-based samples and follow the Low Range Nitrite procedure, Method 8507.
Accuracy check
Standard solution method
Use the standard solution method to validate the test procedure, reagents and
instrument.
Items to collect:
• 200 mg/L NO2– standard solution (Nitrite standard solutions are difficult to prepare.
Use the instructions in Standard Methods for the Examination of Water and
Wastewater, Method 4500—NO2-B)

1. Use the test procedure to measure the concentration of the standard solution.
2. Compare the expected result to the actual result.
Note: The factory calibration can be adjusted slightly with the standard adjust option so that the
instrument shows the expected value of the standard solution. The adjusted calibration is then
used for all test results. This adjustment can increase the test accuracy when there are slight
variations in the reagents or instruments.

Nitrite, Ferrous Sulfate Method (250 mg/L) 3

Method performance
The method performance data that follows was derived from laboratory tests that were
measured on a spectrophotometer during ideal test conditions. Users may get different
results under different test conditions.
Program Standard Precision (95% Confidence Interval) Sensitivity
Concentration change per 0.010 Abs change
373 200 mg/L NO2– 191–209 mg/L NO2– 1.4 mg/L NO2–

Summary of method
The method uses ferrous sulfate in an acidic medium to reduce nitrite to nitrous oxide.
Ferrous ions combine with the nitrous oxide to form a greenish-brown complex in direct
proportion to the nitrite present. The measurement wavelength is 585 nm for
spectrophotometers or 560 nm for colorimeters.
Consumables and replacement items
Required reagents

Description Quantity/test Unit Item no.

®
NitriVer 2 Nitrite Reagent Powder Pillow, 10-mL 1 100/ pkg 2107569

Required apparatus

Description Quantity/test Unit Item no.

Stopper, Neoprene, solid, size #1 2 12/pkg 1480801

Optional reagents and apparatus

Description Unit Item no.

Balance, analytical, 80 g x 0.1 mg 100-240 VAC each 2936701

Standard Methods Book, most current edition each 2270800
Water, deionized 4L 27256
Sodium Nitrite, ACS 454 g 245201

FOR TECHNICAL ASSISTANCE, PRICE INFORMATION AND ORDERING: HACH COMPANY

In the U.S.A. – Call toll-free 800-227-4224 WORLD HEADQUARTERS
Outside the U.S.A. – Contact the HACH office or distributor serving you. Telephone: (970) 669-3050
On the Worldwide Web – www.hach.com; E-mail – [email protected] FAX: (970) 669-2932

© Hach Company/Hach Lange GmbH, 1989–2013. All rights reserved. 04/2013, Edition 8
Nitrogen, Ammonia DOC316.53.01077

Salicylate Method1 Method 8155

0.01 to 0.50 mg/L NH3–N Powder Pillows
Scope and application: For water, wastewater and seawater.
1 Adapted from Clin. Chim. Acta., 14, 403 (1966).

Test preparation

Instrument specific information

The table in this section shows all of the instruments that have the program for this test.
Table 1 shows sample cell and orientation requirements for reagent addition tests, such
as powder pillow or bulk reagent tests.
To use the table, select an instrument, then read across to find the corresponding
information for this test.
Table 1 Instrument-specific information for powder pillows
Instrument Sample cell orientation Sample cell
DR 6000 The fill line is to the right. 2495402
DR 3800
DR 2800
DR 2700
DR 5000 The fill line is toward the user.
DR 3900
DR 900 The orientation mark is toward the user. 2401906

Before starting
Install the instrument cap on the DR 900 cell holder before ZERO or READ is pushed.
The reagents that are used in this test contain sodium nitroferricyanide. Keep cyanide solutions at pH > 11 to prevent
exposure to hydrogen cyanide gas. Collect the reacted samples for proper disposal.
Keep the samples sealed at all times to prevent ammonia contamination from the air.
Review the Safety Data Sheets (MSDS/SDS) for the chemicals that are used and use any recommended personal protective
equipment.
Dispose of reacted solutions according to local, state and federal regulations. Use the Safety Data Sheets for disposal
information for unused reagents. Consult the environmental, health and safety staff for your facility and/or local regulatory
agencies for further disposal information.

Items to collect
Description Quantity

Ammonia Cyanurate Reagent Powder Pillow, 10-mL 2

Ammonia Salicylate Reagent Powder Pillow, 10-mL 2

1
Items to collect (continued)
Description Quantity

Stoppers for 18-mm tubes and AccuVac Ampuls 2

Sample cells (For information about sample cells, adapters or light shields, refer to Instrument
2
specific information on page 1.)

Refer to Consumables and replacement items on page 5 for reorder information.

Sample collection and storage

• Collect samples in clean glass or plastic bottles.
• If the sample contains chlorine, add one drop of 0.1 N sodium thiosulfate to 1 liter of
sample to remove each 0.3 mg/L of chlorine.
• To preserve samples for later analysis, adjust the sample pH to less than 2 with
concentrated sulfuric acid (about 2 mL per liter). No acid addition is necessary if the
sample is tested immediately.
• Keep the preserved samples at or below 6 °C (43 °F) for up to 28 days.
• Let the sample temperature increase to room temperature before analysis.
• Before analysis, adjust the pH to 7 with 5.0 N sodium hydroxide standard solution.
• Correct the test result for the dilution from the volume additions.

Powder pillow procedure

Start

1. Start program 385 N, 2. Prepare the blank: Fill a 3. Prepare the sample: Fill 4. Add the contents of one
Ammonia, Salic. For sample cell with 10 mL of a second sample cell with Ammonia Salicylate powder
information about sample deionized water. 10 mL of sample. pillow to each sample cell.
cells, adapters or light
shields, refer to Instrument
specific information
on page 1.
Note: Although the program
name may vary between
instruments, the program
number does not change.

2 Nitrogen-Ammonia, Salicylate Method (0.50 mg/L)

5. Close the sample cell. 6. Start the instrument 7. After the timer expires, 8. Close the sample cell.
Shake to dissolve the timer. A 3-minute reaction add the contents of one Shake to dissolve the
reagent. time starts. Ammonia Cyanurate powder reagent.
pillow to each sample cell.

Zero

9. Start the instrument 10. When the timer expires, 11. Insert the blank into the 12. Push ZERO. The
timer. A 15-minute reaction clean the blank. cell holder. display shows 0.00 mg/L
time starts. NH3–N.
A green color shows when
ammonia-nitrogen is
present.

Read

13. Clean the prepared 14. Insert the prepared 15. Push READ. Results
sample. sample into the cell holder. show in mg/L NH3–N.

Interferences
Interfering substance Interference level
Calcium 1000 mg/L as CaCO3
Iron All levels. Correct for iron interference as follows:

1. Use one of the Iron, Total procedures to measure the iron concentration of the sample.
2. Use an iron standard solution to add iron to the deionized water blank so that the blank has the
same iron concentration as the sample. The iron interference will be zeroed out from the test
result.

Magnesium 6000 mg/L as CaCO3

Nitrogen-Ammonia, Salicylate Method (0.50 mg/L) 3

Interfering substance Interference level
Monochloramine Monochloramine that is in chloraminated drinking water interferes directly at all levels and gives high
results. Use a Free Ammonia and Monochloramine method to determine free ammonia in these
sample matrices.
Nitrate 100 mg/L as NO3––N
Nitrite 12 mg/L as NO2––N
pH Adjust acidic or basic samples to approximately pH 7. Use 1 N sodium hydroxide standard solution
for acidic samples and 1 N hydrochloric acid standard solution for basic samples.
Phosphate 100 mg/L as PO43––P
Sulfate 300 mg/L as SO42–
Sulfide Sulfide will intensify the color. Remove sulfide interference as follows:

1. Measure approximately 350 mL of sample in a 500-mL Erlenmeyer flask.

2. Add the contents of one Sulfide Inhibitor Reagent Powder Pillow. Swirl to mix.
3. Filter the sample through a folded filter paper and filter funnel.
4. Use the filtered sample in the test procedure.

Other Substances Less common interferences such as hydrazine and glycine cause intensified colors in the prepared
sample. Turbidity and color will give incorrect high values. Samples with severe interferences require
distillation. Use the distillation procedure that is supplied with the distillation set.

Accuracy check
Standard additions method (sample spike)
Use the standard additions method (for applicable instruments) to validate the test
procedure, reagents and instrument and to find if there is an interference in the sample.
Items to collect:
• Ammonia Nitrogen Standard Solution, 10 mg/L as NH3–N
• 25-mL mixing cylinders (3)
• Pipet, TenSette®, 0.1–1.0 mL and tips

1. Use the test procedure to measure the concentration of the sample, then keep the
(unspiked) sample in the instrument.
2. Go to the Standard Additions option in the instrument menu.
3. Select the values for standard concentration, sample volume and spike volumes.
4. Open the standard solution.
5. Prepare three spiked samples: use the TenSette pipet to add 0.2 mL, 0.4 mL and
0.6 mL of the standard solution, respectively, to three 25-mL portions of fresh sample.
Mix well.
6. Use the test procedure to measure the concentration of each of the spiked samples.
Start with the smallest sample spike. Measure each of the spiked samples in the
instrument.
7. Select Graph to compare the expected results to the actual results.
Note: If the actual results are significantly different from the expected results, make sure that
the sample volumes and sample spikes are measured accurately. The sample volumes and
sample spikes that are used should agree with the selections in the standard additions menu. If
the results are not within acceptable limits, the sample may contain an interference.

Standard solution method

Use the standard solution method to validate the test procedure, reagents and
instrument.
Items to collect:
• Ammonia Nitrogen Standard Solution, 10 mg/L as NH3–N

4 Nitrogen-Ammonia, Salicylate Method (0.50 mg/L)

 • 100-mL volumetric flask, Class A
• 4-mL volumetric pipet, Class A and pipet filler
• Deionized water

1. Prepare a 0.40 mg/L ammonia nitrogen standard solution as follows:

a. Use a pipet to add 4 mL of the 10 mg/L ammonia nitrogen standard solution into
the volumetric flask. (Alternate preparation: pipet 0.8 mL of a 50-mg/L ammonia
nitrogen standard solution into a 100-mL volumetric flask.)
b. Dilute to the mark with deionized water. Mix well. Prepare this solution daily.
2. Use the test procedure to measure the concentration of the prepared standard
solution.
3. Compare the expected result to the actual result.
Note: The factory calibration can be adjusted slightly with the standard adjust option so that the
instrument shows the expected value of the standard solution. The adjusted calibration is then
used for all test results. This adjustment can increase the test accuracy when there are slight
variations in the reagents or instruments.

Method performance
The method performance data that follows was derived from laboratory tests that were
measured on a spectrophotometer during ideal test conditions. Users may get different
results under different test conditions.
Program Standard Precision (95% Confidence Interval) Sensitivity
Concentration change per 0.010 Abs change
385 0.40 mg/L NH3–N 0.38–0.42 mg/L NH3–N 0.004 mg/L NH3–N

Summary of method
Ammonia compounds combine with chlorine to form monochloramine. Monochloramine
reacts with salicylate to form 5-aminosalicylate. The 5-aminosalicylate is oxidized in the
presence of a sodium nitroprusside catalyst to form a blue-colored compound. The blue
color is masked by the yellow color from the excess reagent to give a final green-colored
solution. The measurement wavelength is 655 nm for spectrophotometers or 610 nm for
colorimeters.
Pollution prevention and waste management
The ammonia salicylate reagent contains sodium nitroferricyanide and must be disposed
of as a hazardous waste. Dispose of reacted solutions according to local, state and
federal regulations.
Consumables and replacement items
Required reagents

Description Quantity/Test Unit Item no.

Nitrogen Ammonia, reagent set, 10-mL, includes: — 100 tests 2668000

Ammonia Cyanurate Reagent Powder Pillow, 10-mL 2 100/pkg 2653199
Ammonia Salicylate Reagent Powder Pillow, 10-mL 2 100/pkg 2653299

Recommended standards and apparatus

Description Unit Item no.

Flask, volumetric, Class A, 100-mL each 1457442

Nitrogen Ammonia Standard Solution, 10-mg/L NH3–N 500 mL 15349
®
Nitrogen Ammonia Standard Solution, 10-mL Voluette Ampule, 50-mg/L NH3–N 16/pkg 1479110
®
Pipet, TenSette , 0.1–1.0 mL each 1970001

Nitrogen-Ammonia, Salicylate Method (0.50 mg/L) 5

Consumables and replacement items (continued)
Description Unit Item no.

Pipet tips for TenSette Pipet 1970001 50/pkg 2185696

Pipet tips for TenSette Pipet 1970001 1000/pkg 2185628
Pipet, volumetric, Class A, 4.00-mL each 1451504
Stoppers for 18-mm tubes and AccuVac Ampuls 6/pkg 173106
Wastewater, Effluent Inorganics, for NH3-N, NO3-N, PO4, COD, SO4, TOC 500 mL 2833249
Water, deionized 4L 27256

Optional reagents and apparatus

Description Unit Item no.

®
Ampule Breaker, Voluette ampules each 2196800
Cylinder, mixing, 25-mL each 2088640
Distillation heater and support for 2265300, 115 VAC, 60 Hz each 2274400
Distillation apparatus set, general purpose each 2265300
Flask, Erlenmeyer, 500-mL each 50549
Funnel, poly, 65-mm each 108367
Distillation heater and support for 2265300, 230 VAC, 50 Hz each 2274402
Paper, filter, folded, 12.5-cm 100/pkg 189457
Pipet, serological, 2-mL each 53236
Sodium Hydroxide, 5 N 50 mL 245026
Sulfide Inhibitor Reagent Powder Pillow 100/pkg 241899
Sulfuric Acid, concentrated, ACS 500 mL 97949

FOR TECHNICAL ASSISTANCE, PRICE INFORMATION AND ORDERING: HACH COMPANY

In the U.S.A. – Call toll-free 800-227-4224 WORLD HEADQUARTERS
Outside the U.S.A. – Contact the HACH office or distributor serving you. Telephone: (970) 669-3050
On the Worldwide Web – www.hach.com; E-mail – [email protected] FAX: (970) 669-2932

© Hach Company/Hach Lange GmbH, 1989–2013. All rights reserved. 04/2013, Edition 8
Nitrogen, Ammonia DOC316.53.01079

Salicylate Method Method 10031

0.4 to 50.0 mg/L NH3–N (HR) Test ‘N Tube™ Vials
Scope and application: For water, wastewater and seawater.

Test preparation

Instrument specific information

The table in this section shows all of the instruments that have the program for this test.
Table 1 shows adapter and light shield requirements for the instruments that use them.
To use the table, select an instrument, then read across to find the corresponding
information for this test.
Table 1 Instrument-specific information for test tubes
Instrument Adapters Light shield
DR 6000 — —
DR 5000 — —
DR 900 4846400 Cover supplied with the instrument
DR 3900 — LZV849
DR 3800 — LZV646
DR 2800 —

Before starting
Install the instrument cap on the DR 900 cell holder before ZERO or READ is pushed.
DR 3900, DR 3800, DR 2800 and DR 2700: Install the light shield in Cell Compartment #2 before this test is started.
Small sample sizes (such as 0.1 mL) may not be representative of the entire sample. Mix the sample well before the test or
use a different portion of the sample to repeat the test.
The reagents that are used in this test contain sodium nitroferricyanide. Keep cyanide solutions at pH > 11 to prevent
exposure to hydrogen cyanide gas. Collect the reacted samples for proper disposal.
Keep the samples sealed at all times to prevent ammonia contamination from the air.
To prevent airborne cross-contamination of the blank, complete the preparation of the blank before samples and standards
are opened. If the sample or standard containers are open, move to a separate area of the lab to prepare the blank.
Review the Safety Data Sheets (MSDS/SDS) for the chemicals that are used and use any recommended personal protective
equipment.
Dispose of reacted solutions according to local, state and federal regulations. Use the Safety Data Sheets for disposal
information for unused reagents. Consult the environmental, health and safety staff for your facility and/or local regulatory
agencies for further disposal information.

Items to collect
Description Quantity

Light shield (For information about sample cells, adapters or light shields, refer to Instrument
1
specific information on page 1.)
®
High Range Test 'N Tube™ AmVer 3 Nitrogen Ammonia Reagent Set 2

1
Items to collect (continued)
Description Quantity

Funnel, micro (for reagent addition) 1

®
Pipet, TenSette 0.1 to 1.0 mL, with tips varies

Refer to Consumables and replacement items on page 5 for reorder information.

Sample collection and storage

• Collect samples in clean glass or plastic bottles.
• Analyze the samples as soon as possible for best results.
• If chlorine is known to be present, add one drop of 0.1 N Sodium Thiosulfate to 1 L of
sample for each 0.3 mg/L Cl2.
• To preserve samples for later analysis, adjust the sample pH to less than 2 with
concentrated hydrochloric acid (about 2 mL per liter). No acid addition is necessary if
the sample is tested immediately.
• Keep the preserved samples at or below 6 °C (43 °F) for up to 28 days.
• Let the sample temperature increase to room temperature before analysis.
• Before analysis, adjust the pH to 7 with 5.0 N sodium hydroxide standard solution.
• Correct the test result for the dilution from the volume additions.

Test 'N Tube procedure

Start

1. Start program 343 N, 2. Prepare the blank: Add 3. Prepare the sample: 4. Add the contents of one
Ammonia HR TNT. For 0.1 mL of ammonia-free Add 0.1 mL of sample to Ammonia Salicylate
information about sample water to one AmVer™ one AmVer™ Diluent Reagent Powder Pillow for
cells, adapters or light Diluent Reagent Test 'N Reagent Test 'N Tube for 5-mL samples to each vial.
shields, refer to Instrument Tube for High Range High Range Ammonia
specific information Ammonia Nitrogen. Nitrogen.
on page 1.
Note: Although the program
name may vary between
instruments, the program
number does not change.

2 Nitrogen-Ammonia, Salicylate TNT Method (50.0 mg/L)

5. Add the contents of one 6. Put the caps on both 7. Start the instrument 8. Clean the blank vial.
Ammonia Cyanurate vials. Shake thoroughly to timer. A 20-minute reaction
Reagent Powder Pillow to dissolve the powder. time starts.
each vial.

Zero

9. Insert the blank vial into 10. Push ZERO. The 11. Clean the sample vial. 12. Insert the sample vial
the 16-mm cell holder. display shows 0.0 mg/L into the 16-mm cell holder.
NH3–N.

Read

13. Push READ. Results

show in mg/L NH3–N.

Interferences
Interfering substance Interference level
Calcium 50,000 mg/L as CaCO3
Iron All levels. Correct for iron interference as follows:

1. Use one of the Iron, Total procedures to measure the iron concentration of the sample.
2. Use an iron standard solution to add iron to the deionized water blank so that the blank has the
same iron concentration as the sample. The iron interference will be zeroed out from the test
result.

Magnesium 300,000 mg/L as CaCO3

Monochloramine Monochloramine that is in chloraminated drinking water interferes directly at all levels and gives high
results. Use a Free Ammonia and Monochloramine method to determine free ammonia in these
sample matrices.

Nitrogen-Ammonia, Salicylate TNT Method (50.0 mg/L) 3

Interfering substance Interference level
Nitrate 5000 mg/L as NO3––N
Nitrite 600 mg/L as NO2––N
pH Adjust acidic or basic samples to approximately pH 7. Use 1 N sodium hydroxide standard solution
for acidic samples and 1 N hydrochloric acid standard solution for basic samples.
Phosphate 5000 mg/L as PO43––P
Sulfate 6000 mg/L as SO42–
Sulfide Sulfide will intensify the color. Remove sulfide interference as follows:

1. Measure approximately 350 mL of sample in a 500-mL Erlenmeyer flask.

2. Add the contents of one Sulfide Inhibitor Reagent Powder Pillow. Swirl to mix.
3. Filter the sample through a folded filter paper and filter funnel.
4. Use the filtered sample in the test procedure.

Other Substances Less common interferences such as hydrazine and glycine cause intensified colors in the prepared
sample. Turbidity and color will give incorrect high values. Samples with severe interferences require
distillation. Use the distillation procedure that is supplied with the distillation set.

Accuracy check
Standard additions method (sample spike)
Use the standard additions method (for applicable instruments) to validate the test
procedure, reagents and instrument and to find if there is an interference in the sample.
Items to collect:
• Nitrogen, Ammonia Ampule Standard, 150-mg/L NH3–N
• Ampule breaker
• Pipet, TenSette®, 0.1–1.0 mL and tips
• 25-mL mixing cylinders (3)

1. Use the test procedure to measure the concentration of the sample, then keep the
(unspiked) sample in the instrument.
2. Go to the Standard Additions option in the instrument menu.
3. Select the values for standard concentration, sample volume and spike volumes.
4. Open the standard solution.
5. Prepare three spiked samples: use the TenSette pipet to add 0.2 mL, 0.4 mL and
0.6 mL of the standard solution, respectively, to three 25-mL portions of fresh sample.
Mix well.
6. Use the test procedure to measure the concentration of each of the spiked samples.
Start with the smallest sample spike. Measure each of the spiked samples in the
instrument.
7. Select Graph to compare the expected results to the actual results.
Note: If the actual results are significantly different from the expected results, make sure that
the sample volumes and sample spikes are measured accurately. The sample volumes and
sample spikes that are used should agree with the selections in the standard additions menu. If
the results are not within acceptable limits, the sample may contain an interference.

Standard solution method

Use the standard solution method to validate the test procedure, reagents and
instrument.
Items to collect:
• 100-mg/L Ammonia Nitrogen standard
• 50-mL volumetric flask, Class A
• 20-mL volumetric pipet, Class A and pipet filler

4 Nitrogen-Ammonia, Salicylate TNT Method (50.0 mg/L)

 • Deionized water

1. Prepare a 40.0 mg/L ammonia nitrogen standard solution as follows:

a. Use a pipet to add 20.0 mL of 100 mg/L ammonia nitrogen standard solution into
the volumetric flask.
b. Dilute to the mark with deionized water. Mix well. Prepare this solution daily.
2. Use the test procedure to measure the concentration of the prepared standard
solution.
3. Compare the expected result to the actual result.
Note: The factory calibration can be adjusted slightly with the standard adjust option so that the
instrument shows the expected value of the standard solution. The adjusted calibration is then
used for all test results. This adjustment can increase the test accuracy when there are slight
variations in the reagents or instruments.

Method performance
The method performance data that follows was derived from laboratory tests that were
measured on a spectrophotometer during ideal test conditions. Users may get different
results under different test conditions.
Program Standard Precision (95% Confidence Interval) Sensitivity
Concentration change per 0.010 Abs change
343 40.00 mg/L NH3–N 38.1–41.9 mg/L NH3–N 0.312 mg/L NH3–N

Summary of method
Ammonia compounds combine with chlorine to form monochloramine. Monochloramine
reacts with salicylate to form 5-aminosalicylate. The 5-aminosalicylate is oxidized in the
presence of a sodium nitroprusside catalyst to form a blue colored compound. The blue
color is masked by the yellow color from the excess reagent present to give a green-
colored solution. The measurement wavelength is 655 nm for spectrophotometers or
610 nm for colorimeters.
Pollution prevention and waste management
The ammonia salicylate reagent contains sodium nitroferricyanide and must be disposed
of as a hazardous waste. Dispose of reacted solutions according to local, state and
federal regulations.
Consumables and replacement items
Required reagents

Description Quantity/test Unit Item no.

Nitrogen Ammonia, Reagent Set, High Range Test 'N Tube™ AmVer™ 2 25 tests 2606945

Required apparatus

Description Quantity/test Unit Item no.

Funnel, micro, poly 1 each 2584335

®
Pipet, TenSette , 0.1- to 1.0-mL 1 each 1970001
Pipet Tips, for TenSette Pipet 1970001 2 50/pkg 2185696
Test tube rack 1 each 1864100

Nitrogen-Ammonia, Salicylate TNT Method (50.0 mg/L) 5

Recommended standards

Description Unit Item no.

Nitrogen, Ammonia Standard Solution, 10-mg/L NH3-N 500 mL 15349

Nitrogen, Ammonia Standard Solution, 100-mg/L NH3-N 500 mL 2406549
®
Nitrogen, Ammonia Standard Solution, 10-mL Voluette Ampules, 150 mg/L 16/pkg 2128410
®
Nitrogen Ammonia Standard Solution, 10-mL Voluette Ampule, 50-mg/L NH3–N 16/pkg 1479110
Wastewater, Effluent Inorganics, for NH3-N, NO3-N, PO4, COD, SO4, TOC 500 mL 2833249
Water, deionized 4L 27256

Optional reagents and apparatus

Description Unit Item no.

Cylinder, mixing, 25-mL each 2088640

Distillation apparatus set, general purpose each 2265300
Filter paper, folded, 12.5-cm 100/pkg 69257
Pipet tips for TenSette Pipet 1970001 1000/pkg 2185628
®
Ampule Breaker, Voluette ampules each 2196800
Distillation heater and support for 2265300, 115 VAC, 60 Hz each 2274400
Distillation heater and support for 2265300, 230 VAC, 50 Hz each 2274402
Funnel, poly, 65-mm each 108367
Pipet, serological, 2-mL each 50549
Pipet filler, safety bulb each 1465100
Hydrochloric Acid, concentrated 500 mL 13449
Hydrochloric Acid Standard Solution, 1 N 1000 mL 2321353
100 mL
Sodium Hydroxide Standard Solution, 1.0 N 104532
MDB
Sulfide Inhibitor Reagent Powder Pillow 100/pkg 241899
50 mL
Sodium Hydroxide Standard Solution, 5.0 N 245026
SCDB
Sodium Thiosulfate, 0.1 N 100 mL 32332

FOR TECHNICAL ASSISTANCE, PRICE INFORMATION AND ORDERING: HACH COMPANY

In the U.S.A. – Call toll-free 800-227-4224 WORLD HEADQUARTERS
Outside the U.S.A. – Contact the HACH office or distributor serving you. Telephone: (970) 669-3050
On the Worldwide Web – www.hach.com; E-mail – [email protected] FAX: (970) 669-2932

© Hach Company/Hach Lange GmbH, 1989–2013. All rights reserved. 04/2013, Edition 8
Nitrogen, Ammonia DOC316.53.01080

Salicylate Method1 Method 10023

0.02 to 2.50 mg/L NH3–N (LR) Test ‘N Tube™ Vials
Scope and application: For water, wastewater and seawater.
1 Adapted from Clin. Chim. Acta, 14, 403 (1966).

Test preparation

Instrument specific information

The table in this section shows all of the instruments that have the program for this test.
Table 1 shows adapter and light shield requirements for the instruments that use them.
To use the table, select an instrument, then read across to find the corresponding
information for this test.
Table 1 Instrument-specific information for test tubes
Instrument Adapters Light shield
DR 6000 — —
DR 5000 — —
DR 900 4846400 Cover supplied with the instrument
DR 3900 — LZV849
DR 3800 — LZV646
DR 2800 —

Before starting
Install the instrument cap on the DR 900 cell holder before ZERO or READ is pushed.
DR 3900, DR 3800, DR 2800 and DR 2700: Install the light shield in Cell Compartment #2 before this test is started.
The reagents that are used in this test contain sodium nitroferricyanide. Keep cyanide solutions at pH > 11 to prevent
exposure to hydrogen cyanide gas. Collect the reacted samples for proper disposal.
Keep the samples sealed at all times to prevent ammonia contamination from the air.
Review the Safety Data Sheets (MSDS/SDS) for the chemicals that are used and use any recommended personal protective
equipment.
Dispose of reacted solutions according to local, state and federal regulations. Use the Safety Data Sheets for disposal
information for unused reagents. Consult the environmental, health and safety staff for your facility and/or local regulatory
agencies for further disposal information.

Items to collect
Description Quantity

Light shield (For information about sample cells, adapters or light shields, refer to Instrument
1
specific information on page 1.)
Nitrogen Ammonia, Reagent Set, Low Range Test 'N Tube™ AmVer™ 2
Funnel, micro, poly 1
®
Pipet, TenSette , 1.0- to 10.0-mL 1
Pipet Tips, for TenSette Pipet 1970010 2

1
Refer to Consumables and replacement items on page 5 for reorder information.

Sample collection and storage

• Collect samples in clean glass or plastic bottles.
• If the sample contains chlorine, add one drop of 0.1 N sodium thiosulfate to 1 liter of
sample to remove each 0.3 mg/L of chlorine.
• To preserve samples for later analysis, adjust the sample pH to less than 2 with
concentrated hydrochloric acid (about 2 mL per liter). No acid addition is necessary if
the sample is tested immediately.
• Keep the preserved samples at or below 6 °C (43 °F) for up to 28 days.
• Let the sample temperature increase to room temperature before analysis.
• Before analysis, adjust the pH to 7 with 5.0 N sodium hydroxide standard solution.
• Correct the test result for the dilution from the volume additions.

Test 'N Tube procedure

Start

1. Start program 342, 2. Prepare the blank: Add 3. Prepare the sample: 4. Use a funnel to add the
Ammonia LR TNT. For 2.0 mL of ammonia-free Add 2.0 mL of sample to contents of one Ammonia
information about sample water to one AmVer™ one AmVer™ Diluent Salicylate Reagent Powder
cells, adapters or light Diluent Reagent Test ‘N Reagent Test ‘N Tube for Pillow to each vial.
shields, refer to Instrument Tube for Low Range Low Range Ammonia
specific information Ammonia Nitrogen. Nitrogen.
on page 1.
Note: Although the program
name may vary between
instruments, the program
number does not change.

5. Use a funnel to add the 6. Close the vials tightly. 7. Start the instrument 8. After the timer expires,
contents of one Ammonia Shake thoroughly to timer. A 20-minute reaction clean the blank vial.
Cyanurate Reagent Powder dissolve the powder. time starts.
Pillow to each vial.

2 Nitrogen-Ammonia, Salicylate TNT Method (2.50 mg/L)

 Zero

9. Insert the blank vial into 10. Push ZERO. The 11. Clean the sample vial. 12. Insert the sample vial
the cell holder. display shows 0.00 mg/L into the cell holder.
NH3–N.

Read

13. Push READ. Results

show in mg/L NH3–N.

Interferences
Interfering substance Interference level
Calcium 2500 mg/L as CaCO3
Iron All levels. Correct for iron interference as follows:

1. Use one of the Iron, Total procedures to measure the iron concentration of the sample.
2. Use an iron standard solution to add iron to the deionized water blank so that the blank has the
same iron concentration as the sample. The iron interference will be zeroed out from the test
result.

Magnesium 15,000 mg/L as CaCO3

Monochloramine Monochloramine that is in chloraminated drinking water interferes directly at all levels and gives high
results. Use a Free Ammonia and Monochloramine method to determine free ammonia in these
sample matrices.
Nitrate 250 mg/L as NO3––N
Nitrite 30 mg/L as NO2––N
pH Adjust acidic or basic samples to approximately pH 7. Use 1 N sodium hydroxide standard solution
for acidic samples and 1 N hydrochloric acid standard solution for basic samples.
Phosphate 250 mg/L as PO43––P
Sulfate 300 mg/L as SO42–

Nitrogen-Ammonia, Salicylate TNT Method (2.50 mg/L) 3

Interfering substance Interference level
Sulfide Sulfide will intensify the color. Remove sulfide interference as follows:

1. Measure approximately 350 mL of sample in a 500-mL Erlenmeyer flask.

2. Add the contents of one Sulfide Inhibitor Reagent Powder Pillow. Swirl to mix.
3. Filter the sample through a folded filter paper and filter funnel.
4. Use the filtered sample in the test procedure.

Other Substances Less common interferences such as hydrazine and glycine cause intensified colors in the prepared
sample. Turbidity and color will give incorrect high values. Samples with severe interferences require
distillation. Use the distillation procedure that is supplied with the distillation set.

Accuracy check
Standard additions method (sample spike)
Use the standard additions method (for applicable instruments) to validate the test
procedure, reagents and instrument and to find if there is an interference in the sample.
Items to collect:
®
• 50 mg/L Nitrogen-Ammonia Standard Solution, 10-mL Voluette Ampule
• Ampule breaker
• Pipet, TenSette®, 0.1–1.0 mL and tips
• 25-mL mixing cylinders (3)

1. Use the test procedure to measure the concentration of the sample, then keep the
(unspiked) sample in the instrument.
2. Go to the Standard Additions option in the instrument menu.
3. Select the values for standard concentration, sample volume and spike volumes.
4. Open the standard solution.
5. Prepare three spiked samples: use the TenSette pipet to add 0.1 mL, 0.2 mL and
0.3 mL of the standard solution, respectively, to three 25-mL portions of fresh sample.
Mix well.
6. Use the test procedure to measure the concentration of each of the spiked samples.
Start with the smallest sample spike. Measure each of the spiked samples in the
instrument.
7. Select Graph to compare the expected results to the actual results.
Note: If the actual results are significantly different from the expected results, make sure that
the sample volumes and sample spikes are measured accurately. The sample volumes and
sample spikes that are used should agree with the selections in the standard additions menu. If
the results are not within acceptable limits, the sample may contain an interference.

Standard solution method

Use the standard solution method to validate the test procedure, reagents and
instrument.
Items to collect:
• 1.0 mg/L Nitrogen-Ammonia Standard Solution

1. Use the test procedure to measure the concentration of the standard solution.
2. Compare the expected result to the actual result.
Note: The factory calibration can be adjusted slightly with the standard adjust option so that the
instrument shows the expected value of the standard solution. The adjusted calibration is then
used for all test results. This adjustment can increase the test accuracy when there are slight
variations in the reagents or instruments.

4 Nitrogen-Ammonia, Salicylate TNT Method (2.50 mg/L)

Method performance
The method performance data that follows was derived from laboratory tests that were
measured on a spectrophotometer during ideal test conditions. Users may get different
results under different test conditions.
Program Standard Precision (95% Confidence Interval) Sensitivity
Concentration change per 0.010 Abs change
342 1.00 mg/L NH3–N 0.90–1.10 mg/L NH3–N 0.014 mg/L NH3–N

Summary of method
Ammonia compounds combine with chlorine to form monochloramine. Monochloramine
reacts with salicylate to form 5-aminosalicylate. The 5-aminosalicylate is oxidized in the
presence of a sodium nitroprusside catalyst to form a blue colored compound. The blue
color is masked by the yellow color from the excess reagent present to give a green-
colored solution. The measurement wavelength is 655 nm for spectrophotometers or
610 nm for colorimeters.
Pollution prevention and waste management
The ammonia salicylate reagent contains sodium nitroferricyanide and must be disposed
of as a hazardous waste. Dispose of reacted solutions according to local, state and
federal regulations.
Consumables and replacement items
Required reagents

Description Quantity/test Unit Item no.

Nitrogen Ammonia, Reagent Set, Low Range Test 'N Tube™ AmVer™ 2 25 tests 2604545

Required apparatus

Description Quantity/test Unit Item no.

Funnel, micro, poly 1 each 2584335

®
Pipet, TenSette , 1.0- to 10.0-mL 1 each 1970010
Pipet Tips, for TenSette Pipet 1970010 varies 50/pkg 2199796
Test tube rack 1 each 1864100

Recommended standards and apparatus

Description Unit Item no.

Nitrogen Ammonia Standard Solution, 1.0-mg/L NH3–N 500 mL 189149

®
Nitrogen Ammonia Standard Solution, 10-mL Voluette Ampule, 50-mg/L NH3–N 16/pkg 1479110
Wastewater, Effluent Inorganics, for NH3-N, NO3-N, PO4, COD, SO4, TOC 500 mL 2833249
Water, deionized 4L 27256

Optional reagents and apparatus

Description Unit Item no.

Cylinder, mixing, 25-mL each 2088640

Distillation apparatus set, general purpose each 2265300
Funnel, poly, 65-mm each 108367
Paper, filter, folded, 12.5-cm 100/pkg 189457

Nitrogen-Ammonia, Salicylate TNT Method (2.50 mg/L) 5

Consumables and replacement items (continued)
Description Unit Item no.

Pipet tips for TenSette Pipet 1970001 50/pkg 2185696

®
Ampule Breaker, Voluette ampules each 2196800
Distillation heater and support for 2265300, 115 VAC, 60 Hz each 2274400
Distillation heater and support for 2265300, 230 VAC, 50 Hz each 2274402
Pipet, serological, 2-mL each 50549
Pipet filler, safety bulb each 1465100
Hydrochloric Acid Standard Solution, 1 N 1000 mL 2321353
Hydrochloric Acid, concentrated 500 mL 13449
100 mL
Sodium Hydroxide Standard Solution, 1.0 N 104532
MDB
50 mL
Sodium Hydroxide Standard Solution, 5.0 N 245026
SCDB
Sulfide Inhibitor Reagent Powder Pillow 100/pkg 241899

FOR TECHNICAL ASSISTANCE, PRICE INFORMATION AND ORDERING: HACH COMPANY

In the U.S.A. – Call toll-free 800-227-4224 WORLD HEADQUARTERS
Outside the U.S.A. – Contact the HACH office or distributor serving you. Telephone: (970) 669-3050
On the Worldwide Web – www.hach.com; E-mail – [email protected] FAX: (970) 669-2932

© Hach Company/Hach Lange GmbH, 1989–2013. All rights reserved. 04/2013, Edition 8
Nitrogen, Total Inorganic DOC316.53.01090

Titanium Trichloride Reduction Method Method 10021

0.2 to 25.0 mg/L N Test ‘N Tube™ Vials
Scope and application: For water, wastewater and seawater.

Test preparation

Instrument specific information

The table in this section shows all of the instruments that have the program for this test.
Table 1 shows adapter and light shield requirements for the instruments that use them.
To use the table, select an instrument, then read across to find the corresponding
information for this test.
Table 1 Instrument-specific information for test tubes
Instrument Adapters Light shield
DR 6000 — —
DR 5000 — —
DR 900 4846400 Cover supplied with the instrument
DR 3900 — LZV849
DR 3800 — LZV646
DR 2800 —

Before starting
Install the instrument cap on the DR 900 cell holder before ZERO or READ is pushed.
DR 3900, DR 3800, DR 2800 and DR 2700: Install the light shield in Cell Compartment #2 before this test is started.
For safety, wear gloves to break open the reagent ampules.
The reagents that are used in this test contain sodium nitroferricyanide. Keep cyanide solutions at pH > 11 to prevent
exposure to hydrogen cyanide gas. Collect the reacted samples for proper disposal.
Review the Safety Data Sheets (MSDS/SDS) for the chemicals that are used and use any recommended personal protective
equipment.
Dispose of reacted solutions according to local, state and federal regulations. Use the Safety Data Sheets for disposal
information for unused reagents. Consult the environmental, health and safety staff for your facility and/or local regulatory
agencies for further disposal information.

Items to collect
Description Quantity

Total Inorganic Nitrogen Pretreatment Reagent Set (TiCl3 Reduction Method) 1

Test 'N Tube AmVer™ Nitrogen-Ammonia Reagent Set 1
Centrifuge 1
Funnel, micro 1
Light shield or adapter (For information about sample cells, adapters or light shields, refer to
1
Instrument specific information on page 1.)
®
Pipet, TenSette , 0.1- to 1.0-mL, with pipet tips 1
Pipet, volumetric, Class A, 1.00-mL 1

1
Items to collect (continued)
Description Quantity

Test tube rack 1

Water, deionized 1 mL

Refer to Consumables and replacement items on page 6 for reorder information.

Sample collection and storage

• Collect samples in clean glass or plastic bottles.
• Analyze the samples as soon as possible for best results.
• If the sample contains chlorine, add one drop of 0.1 N sodium thiosulfate to 1 liter of
sample to remove each 0.3 mg/L of chlorine.
• To preserve samples for later analysis, adjust the sample pH to less than 2 with
concentrated hydrochloric acid (about 2 mL per liter). No acid addition is necessary if
the sample is tested immediately.
• Keep the preserved samples at or below 6 °C (43 °F) for up to 28 days.
• Let the sample temperature increase to room temperature before analysis.
• Before analysis, adjust the pH to 7 with 5.0 N sodium hydroxide standard solution.
• Correct the test result for the dilution from the volume additions.

Test 'N Tube procedure

Start

1. Start program 346 N 2. Add 1 mL of Total 3. Prepare the sample: 4. Prepare the blank: Add
Inorganic TNT. For Inorganic Nitrogen Add 1 mL of sample to one 1 mL of deionized water to
information about sample Pretreatment Base of the vials. the second vial.
cells, adapters or light concentrate into two Total
shields, refer to Instrument Inorganic Nitrogen
specific information Pretreatment Diluent vials.
on page 1.
Note: Although the program
name may vary between
instruments, the program
number does not change.

2 Nitrogen, Total Inorganic, Titanium Trichloride Reduction TNT Method (25.0 mg/L)
5. Put the caps on both 6. Add the contents of one 7. Put the caps on both 8. Let the vials stand for at
vials. Shake for 30 seconds Total Inorganic Nitrogen vials. Shake gently for at least one minute.
to mix. Reductant ampule to the 30 seconds to mix.
prepared sample vial. Add The precipitate should be
the contents of another black after shaking.
Total Inorganic Nitrogen Excessive shaking will
Reductant ampule to the cause the precipitate to
blank vial. change to a white color and
A black precipitate will form give low test results.
immediately.

9. Put the vials in a 10. Start the instrument 11. After the timer expires, 12. Add 2 mL of blank from
centrifuge. timer. A 3-minute reaction add 2 mL of prepared the centrifuge to a second
time starts. sample from the centrifuge AmVer Diluent Reagent
Put the vials in a centrifuge to an AmVer Diluent Test 'N Tube for Low Range
and run the centrifuge to for Reagent Test 'N Tube for Ammonia Nitrogen.
three minutes. If no Low Range Ammonia
centrifuge is available, let Nitrogen.
the vials sit for 30 minutes Do not disturb the sediment
so the solids settle at the in the bottom of the vials.
bottom of the vials.

13. Add the contents of one 14. Add the contents of one 15. Put the caps on both 16. Start the instrument
Ammonia Salicylate Ammonia Cyanurate vials. Shake to dissolve the timer. A 20-minute reaction
Reagent Powder Pillow (for Reagent Powder Pillow )for powder completely. A green time starts.
5-mL samples) to each vial. 5-mL samples) to each vial. color shows if nitrogen is
present.

Nitrogen, Total Inorganic, Titanium Trichloride Reduction TNT Method (25.0 mg/L) 3
 Zero

17. When the timer expires, 18. Insert the blank vial into 19. Push ZERO. The 20. Clean the sample vial.
clean the blank vial. the 16-mm cell holder. display shows 0.0 mg/L N.

Read

21. Insert the sample vial 22. Push READ. Results

into the 16-mm cell holder. show in mg/L N.

Interferences
The substances in Table 2 may interfere when present. The substances in Table 3 do not
interfere below the levels listed.
Table 2 Interfering substances
Interfering substance Interference level
Calcium Causes a positive interference at 1000 mg/L as CaCO3
Manganese (IV) Causes a negative interference at 3 mg/L
Magnesium Causes a positive interference at 1000 mg/L as CaCO3
Sulfide Causes a negative interference at 3 mg/L
Sulfate Causes a negative interference at 250 mg/L

Table 3 Non-interfering substances

Interfering substance Interference level
Al3+ 8 mg/L
Ba2+ 40 mg/L
Cu2+ 40 mg/L
Fe3+ 8 mg/L
Zn2+ 80 mg/L
F– 40 mg/L
PO43--P 8 mg/L
SiO2 80 mg/L
EDTA 80 mg/L

4 Nitrogen, Total Inorganic, Titanium Trichloride Reduction TNT Method (25.0 mg/L)
Accuracy check
Standard additions method (sample spike)
Use the standard additions method (for applicable instruments) to validate the test
procedure, reagents and instrument and to find if there is an interference in the sample.
Items to collect:
• Nitrate Nitrogen PourRite Ampule Standard, 500-mg/L NO3––N
• Ampule breaker
• Pipet, TenSette®, 0.1–1.0 mL and tips
• 25-mL mixing cylinders (3)

1. Use the test procedure to measure the concentration of the sample, then keep the
(unspiked) sample in the instrument.
2. Go to the Standard Additions option in the instrument menu.
3. Select the values for standard concentration, sample volume and spike volumes.
4. Open the standard solution.
5. Prepare three spiked samples: use the TenSette pipet to add 0.1 mL, 0.2 mL and
0.3 mL of the standard solution, respectively, to three 25-mL portions of fresh sample.
Mix well.
6. Use the test procedure to measure the concentration of each of the spiked samples.
Start with the smallest sample spike. Measure each of the spiked samples in the
instrument.
7. Select Graph to compare the expected results to the actual results.
Note: If the actual results are significantly different from the expected results, make sure that
the sample volumes and sample spikes are measured accurately. The sample volumes and
sample spikes that are used should agree with the selections in the standard additions menu. If
the results are not within acceptable limits, the sample may contain an interference.

Standard solution method

Use the standard solution method to validate the test procedure, reagents and
instrument.
Items to collect:
• 10.0-mg/L Nitrate Nitrogen Standard Solution

1. Use the test procedure to measure the concentration of the standard solution.
2. Compare the expected result to the actual result.
Note: The factory calibration can be adjusted slightly with the standard adjust option so that the
instrument shows the expected value of the standard solution. The adjusted calibration is then
used for all test results. This adjustment can increase the test accuracy when there are slight
variations in the reagents or instruments.

Method performance
The method performance data that follows was derived from laboratory tests that were
measured on a spectrophotometer during ideal test conditions. Users may get different
results under different test conditions.
Program Standard Precision (95% Confidence Interval) Sensitivity
Concentration change per 0.010 Abs change
346 20.0 mg/L NO3––N 19.6–20.4 mg/L NO3––N 0.2 mg/L NO3––N

Species recovery
The total inorganic nitrogen test is designed to provide an estimate of the total nitrite,
nitrate and ammonia nitrogen load in a water or wastewater sample. This test is most
applicable to the monitoring of samples taken from an industrial process stream or a
wastewater treatment stream where it is important to track the inorganic nitrogen load as
it passes through the treatment process. The test does show different recoveries of each

Nitrogen, Total Inorganic, Titanium Trichloride Reduction TNT Method (25.0 mg/L) 5
 of the three nitrogen species, as shown in Table 4. The test is not recommended for use
when quantifying only one of the three species. In that case, specific procedures for each
particular analyte would be more appropriate.
Table 4 Species recovery
Nitrogen form Percent recovery
NH3–N 112%
NO3 ––N 100%
NO2––N 77%

Summary of method
Titanium (III) ions reduce nitrate and nitrite to ammonia in a basic environment. After
centrifugation to remove solids, the ammonia is combined with chlorine to form
monochloramine. Monochloramine reacts with salicylate to form 5-aminosalicylate. The 5-
aminosalicylate is oxidized in the presence of a sodium nitroprusside catalyst to form a
blue colored compound. The blue color is masked by the yellow color from the excess
reagent present to give a final green colored solution. The measurement wavelength is
655 nm for spectrophotometers or 610 nm for colorimeters.
Consumables and replacement items
Required reagents

Description Quantity/test Unit Item no.

Nitrogen, Total Inorganic, Pretreatment Reagent Set (TiCl3 Reduction

— 25 tests 2604945
Method)
Nitrogen Ammonia, Reagent Set, Low Range Test 'N Tube™ AmVer™ 2 25 tests 2604545
Water, deionized varies 100 mL 27242

Required apparatus

Description Quantity/test Unit Item no.

Centrifuge, 115 VAC, 6 x 15 mL 1 each 2676500

OR
Centrifuge, 220 VAC, 6 x 15 mL 1 each 2676502
Funnel, micro, poly 1 each 2584335
®
Pipet, TenSette , 0.1- to 1.0-mL 1 each 1970001
Pipet Tips, for TenSette Pipet 1970001 2 50/pkg 2185696
Test tube rack 1 each 1864100
Gloves, nitrile, large (other sizes available) 1 pair 100/pkg 2550503

Recommended standards and apparatus

Description Unit Item no.

Flask, volumetric, 50-mL each 1457441

Nitrate Nitrogen Standard Solution, 10.0-mg/L NO3--N 500 mL 30749
®
Nitrate Nitrogen Standard Solution, 2-mL PourRite Ampule, 500 mg/L 20/pkg 1426020
Pipet filler, safety bulb each 1465100
®
Pipet, TenSette , 0.1–1.0 mL each 1970001
Pipet tips for TenSette Pipet 1970001 50/pkg 2185696

6 Nitrogen, Total Inorganic, Titanium Trichloride Reduction TNT Method (25.0 mg/L)
Consumables and replacement items (continued)
Description Unit Item no.

Pipet tips for TenSette Pipet 1970001 1000/pkg 2185628

Pipet tips for TenSette Pipet 1970010 250/pkg 2199725
Water, deionized 4L 27256

Optional reagents and apparatus

Description Unit Item no.

Cylinder, mixing, 25-mL each 2088640

Hydrochloric Acid, concentrated 500 mL 13449
50 mL
Sodium Hydroxide Standard Solution, 5.0 N 245026
SCDB
Sodium Thiosulfate, 0.1 N 100 mL 32332
Pipet, volumetric, Class A, 1.00-mL each 1451535
®
PourRite Ampule Breaker, 2-mL each 2484600
®
Ampule Breaker, Voluette ampules each 2196800
Nitrate Nitrogen Standard Solution, 1-mg/L NH3-N 500 mL 204649
Nitrate Nitrogen Standard Solution, 100-mg/L NH3-N 500 mL 194749
Nitrate Nitrogen Standard Solution, 1000-mg/L NH3-N 500 mL 1279249
100 mL
Nitrate Nitrogen Standard Solution, 15-mg/L NH3-N 2415132
MDB

Nitrogen, Total Inorganic, Titanium Trichloride Reduction TNT Method (25.0 mg/L) 7
 FOR TECHNICAL ASSISTANCE, PRICE INFORMATION AND ORDERING: HACH COMPANY
In the U.S.A. – Call toll-free 800-227-4224 WORLD HEADQUARTERS
Outside the U.S.A. – Contact the HACH office or distributor serving you. Telephone: (970) 669-3050
On the Worldwide Web – www.hach.com; E-mail – [email protected] FAX: (970) 669-2932

© Hach Company/Hach Lange GmbH, 1989–2013. All rights reserved. 04/2013, Edition 8
Nitrogen, Total Kjeldahl DOC316.53.01091

Nessler Method1 Method 8075

1 to 150 mg/L TKN Reagent Solution
Scope and application: For water, wastewater and sludge; digestion is required.
1 Adapted from Hach, et. al., Journal of Association of Official Analytical Chemists, 70(5) 783-787 (1987); Hach, et. al., Journal of
Agricultural and Food Chemistry, 33(6) 1117-1123 (1985); Standard Methods for the Examination of Water and Wastewater.

Test preparation

Instrument specific information

The table in this section shows all of the instruments that have the program for this test.
Table 1 shows sample cell and orientation requirements for reagent addition tests, such
as powder pillow or bulk reagent tests.
To use the table, select an instrument, then read across to find the corresponding
information for this test.
Table 1 Instrument-specific information for powder pillows
Instrument Sample cell orientation Sample cell
DR 6000 The fill line is to the right. 2495402
DR 3800
DR 2800
DR 2700
DR 5000 The fill line is toward the user.
DR 3900
DR 900 The orientation mark is toward the user. 2401906

Before starting
Install the instrument cap on the DR 900 cell holder before ZERO or READ is pushed.
For best results, measure the reagent blank value for each new lot of reagent. Replace the sample with deionized water in
the test procedure to get the reagent blank value. Subtract the reagent blank value from the sample results automatically
with the reagent blank adjust option.
If the Pour-Thru Cell is used (for applicable instruments), clean the cell periodically. Pour a few sodium thiosulfate
pentahydrate crystals into the cell funnel or rinse the cell with a solution of sodium thiosulfate. Flush the crystals through the
funnel and cell with enough deionized water to dissolve. Rinse the cell with deionized water.
The Nessler reagent contains mercuric iodide. Both the reacted sample and blank will contain mercury. Do not pour these
solutions down the drain. Collect the reacted samples and the blank for proper disposal.
Hold the reagent droppers and dropper bottles vertically, not at an angle, when the reagent is added.
Use the Standard Adjust option with each new lot of reagent for the best results.
Review the Safety Data Sheets (MSDS/SDS) for the chemicals that are used and use any recommended personal protective
equipment.
Dispose of reacted solutions according to local, state and federal regulations. Use the Safety Data Sheets for disposal
information for unused reagents. Consult the environmental, health and safety staff for your facility and/or local regulatory
agencies for further disposal information.

1
Items to collect
Description Quantity

Boiling chips, silicon carbide 2-3

Cylinder, graduated mixing, 25-mL 2
Finger cots 2
Digesdahl Digestion Apparatus 1
Hydrogen Peroxide, 50% 20 mL
Mineral Stabilizer 6 drops
Nessler Reagent 2 mL
Polyvinyl Alcohol Dispersing Agent 6 drops
Potassium Hydroxide (KOH) Standard Solution, 1.0 N varies
Potassium Hydroxide (KOH) Standard Solution, 8.0 N varies
Sulfuric Acid, ACS, concentrated 6 mL
TKN Indicator Solution 2 drops
Pipet, TenSette, 0.1–1.0 mL, plus tips 1
Safety shield 1
Sample cells (For information about sample cells, adapters or light shields, refer to Instrument
2
specific information on page 1.)

Refer to Consumables and replacement items on page 7 for reorder information.

Sample collection and storage

• Collect samples in clean glass or plastic bottles.
• To preserve samples for later analysis, adjust the sample pH to less than 2 with
concentrated sulfuric acid (about 2 mL per liter). No acid addition is necessary if the
sample is tested immediately.
• Keep the preserved samples at or below 6 °C (43 °F) for up to 28 days.
• Let the sample temperature increase to room temperature before analysis.
• Before analysis, adjust the pH to 3–5 with 5.0 N sodium hydroxide standard solution.
• Correct the test result for the dilution from the volume additions.

2 Nitrogen, Total Kjeldahl, Nessler Method (150 mg/L)

Nessler method

Start

1. Start program 2. Prepare the sample: 3. Prepare the blank: 4. Use a pipet to move an
399 Nitrogen, TKN. For Use the Digesdahl Digestion Digest an equal amount of analysis volume of the
information about sample Apparatus Instruction deionized water for use as digested sample to a
cells, adapters or light Manual to digest the sample the blank . graduated mixing cylinder.
shields, refer to Instrument amount. Refer to Digested Refer to Digested sample
specific information sample volumes volumes on page 5.
on page 1. on page 5.
Note: Although the program
name may vary between
instruments, the program
number does not change.

5. Use a pipet to transfer an 6. Add one drop of TKN 7. If the aliquot is less that 8. Add 1.0 N KOH to each
equal amount of digested Indicator to eacy cylinder. 1 mL, go to step 8. If the cylinder, one drop at a time.
deionized water to a second aliquot is greater than 1 mL, Mix after each addition.
graduated mixing cylinder. add drops of 8.0 KOH to Continue until the first
each cylinder until the first permanent blue color
flash of blue color shows. shows.
Put the stopper in the
cylinder and invert after
each addition.

9. Fill both cylinders to the 10. Add three drops of 11. Put the stoppers in the 12. Add three drops of
20-mL mark with deionized Mineral Stabilizer to each cylinders and invert to mix. Polyvinyl Alcohol Dispersing
water. cylinder. Agent to each cylinder.

Nitrogen, Total Kjeldahl, Nessler Method (150 mg/L) 3

13. Put the stoppers in the 14. Fill both cylinders to the 15. Put the stoppers in the 16. Use a pipet to add
cylinders and invert to mix. 25-mL mark with deionized cylinders and invert several 1.00 mL of Nessler Reagent
water. times to mix. to each cylinder.

17. Put the stoppers in the 18. Start the instrument 19. When the timer expires, 20. Clean the blank.
cylinders and invert to mix. timer. A 2-minute reaction pour the contents of each
The solution should not be time starts. cylinder into separate
hazy. Any turbidity (haze) sample cells.
will cause incorrect results.

Zero

21. Insert the blank into the 22. Push ZERO. The 23. Clean the prepared 24. Insert the prepared
cell holder. display shows 0 mg/L TKN. sample. sample into the cell holder.

4 Nitrogen, Total Kjeldahl, Nessler Method (150 mg/L)

 Read

25. Push READ. Results 26. Calculate the sample

show in mg/L TKN. TKN in ppm:
TKN = (75 x A) ÷ (B x C)
Where:

• A = mg/L read from the

display
• B = g (or mL of water)
sample taken for the
digestion
• C = mL analysis volume
of the digested sample

Digested sample volumes

Table 2 Aqueous samples (solutions or suspensions in water—less than 1% solids)
Expected nitrogen concentration (mg/L) Analysis volume (mL)
0.5–28 10
2–112 5
11–560 2
45–2250 1
425–22500 0.5

Table 3 Dry samples

Expected nitrogen concentration (mg/L) Analysis volume (mL)
42–2200 10
106–5600 5
350–18,000 2
1000–56,000 1
4200–220,000 0.5

Table 4 Oils and fats

Expected nitrogen concentration (mg/L) Analysis volume (mL)
85–4500 10
210–11,000 5
2100–110,000 1

Nitrogen, Total Kjeldahl, Nessler Method (150 mg/L) 5

Accuracy check
Digestion method
To validate the digestion method, use the Primary Standards for Kjeldahl Nitrogen that
are given in the Accuracy Check section of the Digesdahl® Digestion Apparatus
Instruction Manual. Use the accuracy check procedure to find the digestion efficiency and
the amount of bound nitrogen that is released during digestion.
Use the digested Kjeldahl standard in the Nessler test procedure to measure the TKN of
the primary standard. The TKN value should be within ± 3% of the value of the prepared
Kjeldahl standard.

Standard solution method

Items to collect:
• 1.0-mg/L NH3–N standard solution
• TKN indicator
• Dropper
• 25-mL graduated mixing cylinders (2)
• Deionized water
• Mineral Stabilizer
• Polyvinyl Alcohol Dispersing agent

1. Add one drop of TKN Indicator to each 25-mL graduated mixing cylinder.
2. Fill one cylinder to the 20-mL mark with deionized water. Fill the other cylinder to the
20-mL mark with a 1.0-mg/L NH3–N standard solution.
3. Add 3 drops of Mineral Stabilizer to each cylinder. Invert several times to mix.
4. Add 3 drops of Polyvinyl Alcohol Dispersing agent to each cylinder. Invert several
times to mix.
5. Continue with the TKN procedure to measure the concentration of the standard
solution. Accurate calibrations will show 26–27 mg/L TKN.
Note: The factory calibration can be adjusted slightly with the standard adjust option so that the
instrument shows the expected value of the standard solution. The adjusted calibration is then
used for all test results. This adjustment can increase the test accuracy when there are slight
variations in the reagents or instruments.

Method performance
The method performance data that follows was derived from laboratory tests that were
measured on a spectrophotometer during ideal test conditions. Users may get different
results under different test conditions.
Program Standard Precision (95% Confidence Interval) Sensitivity
Concentration change per 0.010 Abs change
399 76 mg/L NH3–N 70–82 mg/L NH3–N 1 mg/L NH3–N

Summary of method
The term Total Kjeldahl Nitrogen refers to the combination of ammonia and organic
nitrogen. However, only the organic nitrogen compounds that are present as organically
bound nitrogen in the trinegative state are determined in this test. Nitrogen in this form is
converted into ammonium salts by the action of sulfuric acid and hydrogen peroxide. The
ammonia is then analyzed by a modified Nessler method test. The measurement
wavelength is 460 nm for spectrophotometers or 420 nm for colorimeters.
Pollution prevention and waste management
The Nessler reagent contains mercuric iodide. The reacted samples and blanks will
contain mercury and must be disposed of as a hazardous waste. Dispose of reacted
solutions according to local, state and federal regulations.

6 Nitrogen, Total Kjeldahl, Nessler Method (150 mg/L)

Consumables and replacement items
Required reagents

Description Quantity/test Unit Item no.

Nitrogen Reagent Set, 0-150 mg/L, Nessler Method — 250 tests 2495300
Includes:
Hydrogen Peroxide, 50% 20 mL 490 mL 2119649
50 mL
Mineral Stabilizer 6 drops 2376626
SCDB
Nessler Reagent 2 mL 500 mL 2119449
50 mL
Polyvinyl Alcohol Dispersing Agent 6 drops 2376526
SCDB
50 mL
Potassium Hydroxide Standard Solution, 1.0 N varies 2314426
SCDB
100 mL
Potassium Hydroxide Standard Solution, 8.0 N varies 28232H
MDB
Sulfuric Acid, concentrated, ACS varies 500 mL 97949
50 mL
TKN Indicator Solution 2 drops 2251926
SCDB

Required apparatus

Description Quantity/test Unit Item no.

Boiling chips, silicon carbide 2-3 500 g 2055734

Cylinder, graduated mixing, 25-mL 2 each 2636240
®
Digesdahl Digestion Apparatus, 115 VAC 1 each 2313020
OR
®
Digesdahl Digestion Apparatus, 220 VAC 1 each 2313021
Finger cots 2 2/pkg 1464702
®
Pipet, TenSette , 0.1- to 1.0-mL 1 each 1970001
Pipet Tips, for TenSette Pipet 1970001 2 50/pkg 2185696
Safety shield 1 each 5003000

Recommended standards

Description Unit Item no.

Kjeldahl Nitrogen Primary Standard Set set of 3 2277800

Nitrogen Ammonia Standard Solution, 1.0-mg/L NH3–N 500 mL 189149
®
Nitrogen, Ammonia Standard Solution, 10-mL Voluette Ampules, 150 mg/L 16/pkg 2128410
Wastewater Influent Standard, Mixed Parameter, for NH3-N, NO3-N, PO4, COD, SO4,
500 mL 2833149
TOC

Optional reagents and apparatus

Description Unit Item no.

Sodium Thiosulfate, Pentahydrate 454 g 46001

Pour-Thru Cell Kit (DR 2700, DR 2800) each 5940400

Nitrogen, Total Kjeldahl, Nessler Method (150 mg/L) 7

Consumables and replacement items (continued)
Description Unit Item no.

Pour-Thru Cell Kit (DR 5000) each LZV479

®
PourRite Ampule Breaker, 2-mL each 2484600
®
Ampule Breaker, Voluette ampules each 2196800
Paper, for weighing, 100 x 100 mm 500/pkg 1473885
®
Pipet, TenSette , 1.0 to 10.0 mL each 1970010
Pipet tips for TenSette Pipet 1970010 50/pkg 2199796
Pipet tips for TenSette Pipet 1970010 250/pkg 2199725
Pipet tips for TenSette Pipet 1970001 1000/pkg 2185628
Nitrogen Ammonia Standard Solution, 10-mg/L NH3–N 500 mL 15349
Nitrogen Ammonia Standard Solution, 100-mg/L as NH3–N 500 mL 2406549
Nitrogen, Ammonia Standard Solution, 1000-mg/L NH3-N 1L 2354153
®
Nitrogen Ammonia Standard Solution, 10-mL Voluette Ampule, 50-mg/L NH3–N 16/pkg 1479110
Balance, analytical, 80 g x 0.1 mg 100-240 VAC each 2936701
®
Nitrogen Ammonia Standard Solution, 2-mL PourRite Ampules, 50 mg/L 20/pkg 1479120

FOR TECHNICAL ASSISTANCE, PRICE INFORMATION AND ORDERING: HACH COMPANY

In the U.S.A. – Call toll-free 800-227-4224 WORLD HEADQUARTERS
Outside the U.S.A. – Contact the HACH office or distributor serving you. Telephone: (970) 669-3050
On the Worldwide Web – www.hach.com; E-mail – [email protected] FAX: (970) 669-2932

© Hach Company/Hach Lange GmbH, 1989–2013. All rights reserved. 04/2013, Edition 8
Nitrogen, Total DOC316.53.01085

Persulfate Digestion Method Method 10072

2 to 150 mg/L N (HR) Test ‘N Tube™ Vials
Scope and application: For water and wastewater.

Test preparation

Instrument specific information

The table in this section shows all of the instruments that have the program for this test.
Table 1 shows adapter and light shield requirements for the instruments that use them.
To use the table, select an instrument, then read across to find the corresponding
information for this test.
Table 1 Instrument-specific information for test tubes
Instrument Adapters Light shield
DR 6000 — —
DR 5000 — —
DR 900 4846400 Cover supplied with the instrument
DR 3900 — LZV849
DR 3800 — LZV646
DR 2800 —

Before starting
Install the instrument cap on the DR 900 cell holder before ZERO or READ is pushed.
DR 3900, DR 3800, DR 2800 and DR 2700: Install the light shield in Cell Compartment #2 before this test is started.
Digestion is required for total nitrogen determinations.
The vials must be mixed carefully for accurate results. Start each vial inversion with the vial in the vertical position, with the
cap on the top. Turn the vial upside-down and wait for all of the solution to flow down to the cap. Return the vial to the
vertical position and wait for all of the solution to flow down to the bottom of the vial. This mixing method equals one
inversion.
If the test result is over-range, dilute a fresh portion of sample and repeat the complete test procedure. The digestion must
be repeated for accurate results.
Use the deionized water that is supplied in the reagent set or organic-free water for the blank vial and for the preparation of
standard solutions.
Review the Safety Data Sheets (MSDS/SDS) for the chemicals that are used and use any recommended personal protective
equipment.
Dispose of reacted solutions according to local, state and federal regulations. Use the Safety Data Sheets for disposal
information for unused reagents. Consult the environmental, health and safety staff for your facility and/or local regulatory
agencies for further disposal information.

Items to collect
Description Quantity

Test 'N Tube HR Total Nitrogen Reagent Set 1

DRB200 Reactor 1
Finger cots 2

1
Items to collect (continued)
Description Quantity

Funnel, micro 1
Light shield or adapter (For information about sample cells, adapters or light shields, refer to
1
Instrument specific information on page 1.)
®
Pipet, TenSette , 0.1- to 1.0-mL, with pipet tips 1
®
Pipet, TenSette , 1.0- to 10.0-mL, with pipet tips 1
Test tube rack 1

Refer to Consumables and replacement items on page 7 for reorder information.

Sample collection and storage

• Collect samples in clean glass or plastic bottles.
• Analyze the samples as soon as possible for best results.
• To preserve samples for later analysis, adjust the sample pH to less than 2 with
concentrated sulfuric acid (about 2 mL per liter). No acid addition is necessary if the
sample is tested immediately.
• Keep the preserved samples at or below 6 °C (43 °F) for up to 28 days.
• Let the sample temperature increase to room temperature before analysis.
• Before analysis, adjust the pH to 7 with 5.0 N sodium hydroxide standard solution.
• Correct the test result for the dilution from the volume additions.

Persulfate digestion for Test 'N Tubes

1. Start the 2. Use a funnel to add the 3. Prepare the sample: 4. Prepare the blank: Add
DRB200 Reactor. Set the contents of one Total Add 0.5 mL of sample to 0.5 mL of deionized water
temperature to 105 °C. Nitrogen Persulfate Reagent one of the vials. (included in the kit) to the
Powder Pillow to each of second vial.
two HR Total Nitrogen Use only water that is free of
Hydroxide Digestion all nitrogen-containing
Reagent vials. species as a substitute for
Make sure to clean any the provided deionized
reagent that gets on the lip water.
of the vials or on the vial
threads.

2 Nitrogen, Total, Persulfate Digestion TNT Method (150 mg/L)

 Start

5. Put the caps on both 6. Put the vials in the 7. At 30 minutes, use finger 8. Start program 394 N,
vials. Shake vigorously for reactor and close the lid. cots to immediately remove Total HR TNT. For
at least 30 seconds to mix. Leave the vials in the the vials from the reactor. information about sample
Undissolved powder will not reactor for exactly Let the vials cool to room cells, adapters or light
affect the accuracy of the 30 minutes. temperature. shields, refer to Instrument
test. specific information
on page 1.
Note: Although the program
name may vary between
instruments, the program
number does not change.

9. Add the contents of one 10. Put the caps on both 11. Start the instrument 12. After the timer expires,
Total Nitrogen (TN) Reagent vials. Shake for 30 seconds. timer. A 3-minute reaction remove the caps from the
A Powder Pillow to each time starts. vials. Add one TN Reagent
vial. B Powder Pillow to each
vial.

13. Put the caps on both 14. Start the instrument 15. Prepared sample: 16. Blank: When the timer
vials. Shake vigorously for timer. A 2-minute reaction When the timer expires, use expires, use a pipet to put
15 seconds to mix. The time starts. a pipet to put 2 mL of the 2 mL of the digested,
reagent will not dissolve digested, treated prepared treated blank into the
completely. Undissolved sample into one TN second TN Reagent C vial.
powder will not affect the Reagent C vial.
accuracy of the test.
The solution will start to turn
yellow.

Nitrogen, Total, Persulfate Digestion TNT Method (150 mg/L) 3

17. Put the caps on both 18. Start the instrument 19. When the timer expires, 20. Insert the blank vial into
vials. Invert 10 times to mix. timer. A 5-minute. reaction clean the blank vial. the 16-mm cell holder.
Use slow, deliberation time starts.
inversions for complete The yellow color will
recovery. The vials will be intensify.
warm to the touch.

Zero Read

21. Push ZERO. The 22. Clean the sample vial. 23. Insert the sample vial 24. Push READ. Results
display shows 0 mg/L N. into the 16-mm cell holder. show in mg/L N.

Blanks for colorimetric measurement

The reagent blank can be used for up to 7 days for measurements that use the same lot
of reagents. Keep the reagent blank in the dark at room temperature (18–25 °C). If a
small amount of white floc appears within a week, discard the reagent blank and prepare
a new one.
Interferences
The substances in the Table 2 have been tested and found not to interfere up to the
indicated levels (in mg/L). Interfering substances that resulted in a concentration change
of ±10% appear in the Table 3.
Table 2 Non-interfering substances
Interfering substance Interference level
Barium 10.4 mg/L
Calcium 1200 mg/L
Chromium (3+) 2 mg/L
Iron 8 mg/L
Lead 26.4 µg/L
Magnesium 2000 mg/L
Organic Carbon 600 mg/L
Phosphorus 400 mg/L
Silica 600 mg/L

4 Nitrogen, Total, Persulfate Digestion TNT Method (150 mg/L)

 Table 2 Non-interfering substances (continued)
Interfering substance Interference level
Silver 3.6 mg/L
Tin 6 mg/L

Table 3 Interfering substances

Interfering substance Interference level
Bromide > 240 mg/L; positive interference
Chloride > 3000 mg/L; positive interference

This test performed with standard nitrogen solutions prepared from the following
compounds obtained 95% recovery:
• Ammonium chloride
• Ammonium sulfate
• Ammonium acetate
• Glycine
• Urea
Ammonium chloride or nicotinic-PTSA spikes in domestic influent, effluent and the ASTM
standard specification for substitute wastewater (D 5905-96) also resulted in ≥ 95%
recovery.
The large amounts of nitrogen-free organic compounds in some samples may decrease
digestion efficiency by consuming some of the persulfate reagent. Samples known to
contain high levels of organics should be diluted and re-run to verify digestion efficiency.
Accuracy check
Digestion method
For proof of accuracy use Primary Standards for Kjeldahl Nitrogen. This method generally
gives 95–100% recovery on organic nitrogen standards. Analysts have found Ammonia-
PTSA (p-Toluenesulfonate) to be the most difficult to digest. Other compounds may yield
different percent recoveries.
Items to collect:
• Primary Standard for Kjeldahl Nitrogen (Ammonia-PTSA, Glycine-PTSA or Nicotinic-
PTSA)
• 1-L volumetric flask, Class A
• Deionized water (use the deionized water supplied in the reagent set or water that is
free of all organic and nitrogen-containing species)

1. Prepare a 120-mg/L N equivalent standard.

a. Weigh the applicable standard:
• Ammonia-PTSA: 1.6208 g
• Glycine-PTSA: 2.1179 g
• Nicotinic-PTSA: 2.5295 g
b. Use a funnel to add the standard to the volumetric flask.
c. Add deionized water to the flask and mix to dissolve the standard.
d. Dilute to the mark with deionized water. Mix well.
2. Use the test procedure to measure the concentration of the nitrogen standard.
Calculate the percent recovery as follows:
% recovery = [(measured concentration)/120] x 100
Note: The minimum expected % recovery for each standard is 95%.

Nitrogen, Total, Persulfate Digestion TNT Method (150 mg/L) 5

Standard additions method (sample spike)
Use the standard additions method (for applicable instruments) to validate the test
procedure, reagents and instrument and to find if there is an interference in the sample.
Items to collect:
• Ammonia Nitrogen Standard Solution, 1000-mg/L as NH3–N
• Ampule breaker
• Pipet, TenSette®, 0.1–1.0 mL and tips
• 25-mL mixing cylinders (3)

1. Use the test procedure to measure the concentration of the sample, then keep the
(unspiked) sample in the instrument.
2. Go to the Standard Additions option in the instrument menu.
3. Select the values for standard concentration, sample volume and spike volumes.
4. Open the standard solution.
5. Prepare three spiked samples: use the TenSette pipet to add 0.1 mL, 0.2 mL and
0.3 mL of the standard solution, respectively, to three 25-mL portions of fresh sample.
Mix well.
6. Use the test procedure to measure the concentration of each of the spiked samples.
Start with the smallest sample spike. Measure each of the spiked samples in the
instrument.
7. Select Graph to compare the expected results to the actual results.
Note: If the actual results are significantly different from the expected results, make sure that
the sample volumes and sample spikes are measured accurately. The sample volumes and
sample spikes that are used should agree with the selections in the standard additions menu. If
the results are not within acceptable limits, the sample may contain an interference.

Standard solution method

Use the standard solution method to validate the test procedure, reagents and
instrument.
Items to collect:
• 100-mg/L ammonia nitrogen standard solution

1. Use the test procedure to measure the concentration of the standard solution.
2. Compare the expected result to the actual result.
Note: The factory calibration can be adjusted slightly with the standard adjust option so that the
instrument shows the expected value of the standard solution. The adjusted calibration is then
used for all test results. This adjustment can increase the test accuracy when there are slight
variations in the reagents or instruments.

Method performance
The method performance data that follows was derived from laboratory tests that were
measured on a spectrophotometer during ideal test conditions. Users may get different
results under different test conditions.
Program Standard Precision (95% Confidence Interval) Sensitivity
Concentration change per 0.010 Abs change
394 100 mg/L NH3–N 98–102 mg/L N 0.5 mg/L N

Summary of method
An alkaline persulfate digestion converts all forms of nitrogen to nitrate. Sodium
metabisulfite is added after the digestion to eliminate halogen oxide interferences. Nitrate
then reacts with chromotropic acid under strongly acidic conditions to form a yellow
complex. The measurement wavelength is 410 nm for spectrophotometers or 420 nm for
colorimeters.

6 Nitrogen, Total, Persulfate Digestion TNT Method (150 mg/L)

Consumables and replacement items
Required reagents

Description Quantity/test Unit Item no.

Nitrogen, Total, Test 'N Tube™ Reagent Set 50 vials 2714100

Required apparatus

Description Quantity/test Unit Item no.

DRB200 Reactor, 110 V, 15 x 16 mm 1 each LTV082.53.40001

OR
DRB200 Reactor, 220 V, 15 x 16 mm 1 each LTV082.52.40001
Funnel, micro, poly 1 each 2584335
®
Pipet, TenSette , 0.1- to 1.0-mL 1 each 1970001
Pipet Tips, for TenSette Pipet 1970001 2 50/pkg 2185696
Pipet, TenSette 1.0–10.0 mL 1 each 1970010
Pipet Tips, for TenSette Pipet 1970010 varies 50/pkg 2199796
Test tube rack 1 each 1864100
Finger cots 2 2/pkg 1464702

Recommended standards and apparatus

Description Unit Item no.

Nitrogen Ammonia Standard Solution, 1000-mg/L as NH3–N 1L 2354153

Nitrogen Ammonia Standard Solution, 100-mg/L as NH3–N 500 mL 2406549
Balance, analytical, 80 g x 0.1 mg 100-240 VAC each 2936701
Cylinder, mixing, 25-mL each 2088640
Flask, volumetric, Class A, 1000-mL each 1457453
Pipet tips for TenSette Pipet 1970001 1000/pkg 2185628
Pipet tips for TenSette Pipet 1970010 250/pkg 2199725
Kjeldahl Nitrogen Primary Standard Set set of 3 2277800
Sodium Hydroxide, 5 N 50 mL 245026
Sulfuric Acid, ACS 500 mL 97949
Wastewater Influent Standard, Mixed Parameter, for NH3-N, NO3-N, PO4, COD, SO4,
500 mL 2833149
TOC
Water, deionized 500 mL 27249
Water, organic-free 500 mL 2641549
Paper, for weighing, 100 x 100 mm 500/pkg 1473885
®
PourRite Ampule Breaker, 2-mL each 2484600
®
Ampule Breaker, Voluette ampules each 2196800
®
Nitrogen Ammonia Standard Solution, 10-mL Voluette Ampule, 50-mg/L NH3–N 16/pkg 1479110
Nitrogen Ammonia Standard Solution, 150-mg/L NH3–N, 10-mL Voulette® Ampules 16/pkg 2128410
®
Nitrogen Ammonia Standard Solution, 2-mL PourRite Ampules, 50 mg/L 20/pkg 1479120
Nitrogen, Ammonia Standard Solution, 10-mg/L NH3-N 500 mL 15349

Nitrogen, Total, Persulfate Digestion TNT Method (150 mg/L) 7

 FOR TECHNICAL ASSISTANCE, PRICE INFORMATION AND ORDERING: HACH COMPANY
In the U.S.A. – Call toll-free 800-227-4224 WORLD HEADQUARTERS
Outside the U.S.A. – Contact the HACH office or distributor serving you. Telephone: (970) 669-3050
On the Worldwide Web – www.hach.com; E-mail – [email protected] FAX: (970) 669-2932

© Hach Company/Hach Lange GmbH, 1989–2013. All rights reserved. 04/2013, Edition 8
Nitrogen, Total DOC316.53.01086

Persulfate Digestion Method Method 10071

0.5 to 25.0 mg/L N (LR) Test ‘N Tube™ Vials
Scope and application: For water and wastewater.

Test preparation

Instrument specific information

The table in this section shows all of the instruments that have the program for this test.
Table 1 shows adapter and light shield requirements for the instruments that use them.
To use the table, select an instrument, then read across to find the corresponding
information for this test.
Table 1 Instrument-specific information for test tubes
Instrument Adapters Light shield
DR 6000 — —
DR 5000 — —
DR 900 4846400 Cover supplied with the instrument
DR 3900 — LZV849
DR 3800 — LZV646
DR 2800 —

Before starting
Install the instrument cap on the DR 900 cell holder before ZERO or READ is pushed.
DR 3900, DR 3800, DR 2800 and DR 2700: Install the light shield in Cell Compartment #2 before this test is started.
Digestion is required for total nitrogen determinations.
The vials must be mixed carefully for accurate results. Start each vial inversion with the vial in the vertical position, with the
cap on the top. Turn the vial upside-down and wait for all of the solution to flow down to the cap. Return the vial to the
vertical position and wait for all of the solution to flow down to the bottom of the vial. This mixing method equals one
inversion.
If the test result is over-range, dilute a fresh portion of sample and repeat the complete test procedure. The digestion must
be repeated for accurate results.
Use the deionized water that is supplied in the reagent set or organic-free water for the blank vial and for the preparation of
standard solutions.
Review the Safety Data Sheets (MSDS/SDS) for the chemicals that are used and use any recommended personal protective
equipment.
Dispose of reacted solutions according to local, state and federal regulations. Use the Safety Data Sheets for disposal
information for unused reagents. Consult the environmental, health and safety staff for your facility and/or local regulatory
agencies for further disposal information.

Items to collect
Description Quantity

Test 'N Tube LR Total Nigtrogen Reagent Set 1

DRB200 Reactor 1
Finger cots 2

1
Items to collect (continued)
Description Quantity

Funnel, micro 1
Light shield or adapter (For information about sample cells, adapters or light shields, refer to
1
Instrument specific information on page 1.)
®
Pipet, TenSette , 0.1- to 1.0-mL, with pipet tips 1
Test tube rack 1 to 3

Refer to Consumables and replacement items on page 7 for reorder information.

Sample collection and storage

• Collect samples in clean glass or plastic bottles.
• Analyze the samples as soon as possible for best results.
• To preserve samples for later analysis, adjust the sample pH to less than 2 with
concentrated sulfuric acid (about 2 mL per liter). No acid addition is necessary if the
sample is tested immediately.
• Keep the preserved samples at or below 6 °C (43 °F) for up to 28 days.
• Let the sample temperature increase to room temperature before analysis.
• Before analysis, adjust the pH to 7 with 5.0 N sodium hydroxide standard solution.
• Correct the test result for the dilution from the volume additions.

Persulfate digestion for Test 'N Tubes

1. Start the 2. Use a funnel to add the 3. Prepare the sample: 4. Prepare the blank: Add
DRB200 Reactor. Set the contents of one Total Add 2 mL of sample to one 2 mL of deionized water
temperature to 105 °C. Nitrogen Persulfate Reagent of the vials. (included in the kit) to the
Powder Pillow to each of second vial.
two Total Nitrogen Use only water that is free of
Hydroxide Digestion all nitrogen-containing
Reagent vials. species as a substitute for
Make sure to clean any the provided deionized
reagent that gets on the lip water.
of the vials or on the vial
threads.

2 Nitrogen, Total, Persulfate Digestion TNT Method (25.0 mg/L)

 Start

5. Put the caps on both 6. Put the vials in the 7. At 30 minutes, use finger 8. Start program 350 N,
vials. Shake vigorously for reactor and close the lid. cots to immediately remove Total LR TNT. For
at least 30 seconds to mix. Leave the vials in the the vials from the reactor. information about sample
Undissolved powder will not reactor for exactly Let the vials cool to room cells, adapters or light
affect the accuracy of the 30 minutes. temperature. shields, refer to Instrument
test. specific information
on page 1.
Note: Although the program
name may vary between
instruments, the program
number does not change.

9. Add the contents of one 10. Put the caps on both 11. Start the instrument 12. After the timer expires,
Total Nitrogen (TN) Reagent vials. Shake for 15 seconds. timer. A 3-minute reaction remove the caps from the
A Powder Pillow to each time starts. vials. Add one TN Reagent
vial. B Powder Pillow to each
vial.

13. Put the caps on both 14. Start the instrument 15. Prepared sample: 16. Blank: When the timer
vials. Shake for 15 seconds timer. A 2-minute reaction When the timer expires, use expires, use a pipet to put
to mix. The reagent will not time starts. a pipet to put 2 mL of the 2 mL of the digested,
dissolve completely. digested, treated prepared treated blank into the
Undissolved powder will not sample into one TN second TN Reagent C vial.
affect the accuracy of the Reagent C vial.
test.
The solution will start to turn
yellow.

Nitrogen, Total, Persulfate Digestion TNT Method (25.0 mg/L) 3

17. Put the caps on both 18. Start the instrument 19. When the timer expires, 20. Insert the blank vial into
vials. Invert 10 times to mix. timer. A 5-minute reaction clean the blank vial. the 16-mm cell holder.
Use slow, deliberation time starts.
inversions for complete The yellow color will
recovery. The vials will be intensify.
warm to the touch.

Zero Read

21. Push ZERO. The 22. Clean the sample vial. 23. Insert the sample vial 24. Push READ. Results
display shows 0.0 mg/L N. into the 16-mm cell holder. show in mg/L N.
Multiple samples can be
measured after "zero" is set
with the blank.

Blanks for colorimetric measurement

The reagent blank can be used for up to 7 days for measurements that use the same lot
of reagents. Keep the reagent blank in the dark at room temperature (18–25 °C). If a
small amount of white floc appears within a week, discard the reagent blank and prepare
a new one.
Interferences
The substances in the Table 2 have been tested and found not to interfere up to the
indicated levels (in mg/L). Interfering substances that resulted in a concentration change
of ±10% appear in the Table 3.
Table 2 Non-interfering substances
Interfering substance Interference level
Barium 2.6 mg/L
Calcium 300 mg/L
Chromium (3+) 0.5 mg/L
Iron 2 mg/L
Lead 6.6 µg/L
Magnesium 500 mg/L
Organic Carbon 150 mg/L
Phosphorus 100 mg/L

4 Nitrogen, Total, Persulfate Digestion TNT Method (25.0 mg/L)

 Table 2 Non-interfering substances (continued)
Interfering substance Interference level
Silica 150 mg/L
Silver 0.9 mg/L
Tin 1.5 mg/L

Table 3 Interfering substances

Interfering substance Interference level
Bromide > 60 mg/L; positive interference
Chloride > 1000 mg/L; positive interference

This test performed with standard nitrogen solutions prepared from the following
compounds obtained 95% recovery:
• Ammonium chloride
• Ammonium sulfate
• Ammonium acetate
• Glycine
• Urea
Ammonium chloride or nicotinic-PTSA spikes in domestic influent, effluent and the ASTM
standard specification for substitute wastewater (D 5905-96) also resulted in ≥ 95%
recovery.
The large amounts of nitrogen-free organic compounds in some samples may decrease
digestion efficiency by consuming some of the persulfate reagent. Samples known to
contain high levels of organics should be diluted and re-run to verify digestion efficiency.
Accuracy check
Digestion method
For proof of accuracy use Primary Standards for Kjeldahl Nitrogen. This method generally
gives 95–100% recovery on organic nitrogen standards. Analysts have found Ammonia-
PTSA (p-Toluenesulfonate) to be the most difficult to digest. Other compounds may yield
different percent recoveries.
Items to collect:
• Primary Standard for Kjeldahl Nitrogen (Ammonia-PTSA, Glycine-PTSA or Nicotinic-
PTSA)
• 1-L volumetric flask, Class A
• Deionized water (use the deionized water supplied in the reagent set or water that is
free of all organic and nitrogen-containing species)

1. Prepare a 25-mg/L N equivalent standard.

a. Weigh the applicable standard:
• Ammonia-PTSA: 0.3379 g
• Glycine-PTSA: 0.4416 g
• Nicotinic-PTSA: 0.5274 g
b. Use a funnel to add the standard to the volumetric flask.
c. Add deionized water to the flask and mix to dissolve the standard.
d. Dilute to the mark with deionized water. Mix well.
2. Use the test procedure to measure the concentration of the nitrogen standard.
Calculate the percent recovery as follows:
% recovery = [(measured concentration)/25] x 100
Note: The minimum expected % recovery for each standard is 95%.

Nitrogen, Total, Persulfate Digestion TNT Method (25.0 mg/L) 5

Standard additions method (sample spike)
Use the standard additions method (for applicable instruments) to validate the test
procedure, reagents and instrument and to find if there is an interference in the sample.
Items to collect:
• Ammonia Nitrogen Standard Solution, 1000-mg/L as NH3–N
• Ampule breaker
• Pipet, TenSette®, 0.1–1.0 mL and tips
• 50-mL mixing cylinders (3)

1. Use the test procedure to measure the concentration of the sample, then keep the
(unspiked) sample in the instrument.
2. Go to the Standard Additions option in the instrument menu.
3. Select the values for standard concentration, sample volume and spike volumes.
4. Open the standard solution.
5. Prepare three spiked samples: use the TenSette pipet to add 0.1 mL, 0.2 mL and
0.3 mL of the standard solution, respectively, to three 50-mL portions of fresh sample.
Mix well.
6. Use the test procedure to measure the concentration of each of the spiked samples.
Start with the smallest sample spike. Measure each of the spiked samples in the
instrument.
7. Select Graph to compare the expected results to the actual results.
Note: If the actual results are significantly different from the expected results, make sure that
the sample volumes and sample spikes are measured accurately. The sample volumes and
sample spikes that are used should agree with the selections in the standard additions menu. If
the results are not within acceptable limits, the sample may contain an interference.

Standard solution method

Use the standard solution method to validate the test procedure, reagents and
instrument.
Items to collect:
• 10-mg/L ammonia nitrogen standard solution

1. Use the test procedure to measure the concentration of the standard solution.
2. Compare the expected result to the actual result.
Note: The factory calibration can be adjusted slightly with the standard adjust option so that the
instrument shows the expected value of the standard solution. The adjusted calibration is then
used for all test results. This adjustment can increase the test accuracy when there are slight
variations in the reagents or instruments.

Method performance
The method performance data that follows was derived from laboratory tests that were
measured on a spectrophotometer during ideal test conditions. Users may get different
results under different test conditions.
Program Standard Precision (95% Confidence Interval) Sensitivity
Concentration change per 0.010 Abs change
350 10 mg/L NH3–N 9.6–10.4 mg/L N 0.5 mg/L N

Summary of method
An alkaline persulfate digestion converts all forms of nitrogen to nitrate. Sodium
metabisulfite is added after the digestion to eliminate halogen oxide interferences. Nitrate
then reacts with chromotropic acid under strongly acidic conditions to form a yellow
complex. The measurement wavelength is 410 nm for spectrophotometers or 420 nm for
colorimeters.

6 Nitrogen, Total, Persulfate Digestion TNT Method (25.0 mg/L)

Consumables and replacement items
Required reagents

Description Quantity/test Unit Item no.

Nitrogen, Total, LR, Test 'N Tube™ Reagent Set 50 vials 2672245

Required apparatus

Description Quantity/test Unit Item no.

DRB200 Reactor, 110 V, 15 x 16 mm 1 each LTV082.53.40001

OR
DRB200 Reactor, 220 V, 15 x 16 mm 1 each LTV082.52.40001
Funnel, micro, poly 1 each 2584335
®
Pipet, TenSette , 0.1- to 1.0-mL 1 each 1970001
Pipet Tips, for TenSette Pipet 1970001 2 50/pkg 2185696
Test tube rack 1 each 1864100
Finger cots 2 2/pkg 1464702

Recommended standards

Description Unit Item no.

Nitrogen, Ammonia Standard Solution, 1000-mg/L NH3-N 1L 2354153

Nitrogen Ammonia Standard Solution, 10-mg/L NH3–N 500 mL 15349
Kjeldahl Nitrogen Primary Standard Set set of 3 2277800
Wastewater Influent Standard, Mixed Parameter, for NH3-N, NO3-N, PO4, COD, SO4,
500 mL 2833149
TOC
Water, deionized 500 mL 27249
Water, organic-free 500 mL 2641549

Optional reagents and apparatus

Description Unit Item no.

Balance, analytical, 80 g x 0.1 mg 100-240 VAC each 2936701

Cylinder, mixing, 50-mL each 2088641
Flask, volumetric, Class A, 1000-mL each 1457453
Sodium Hydroxide, 5 N 50 mL 245026
Sulfuric Acid, ACS 500 mL 97949
®
Ampule Breaker, PourRite ampules each 2484600
®
Ampule Breaker, Voluette ampules each 2196800
Nitrogen Ammonia Standard Solution, 1.0-mg/L NH3–N 500 mL 189149
Nitrogen, Ammonia Standard Solution, 100-mg/L NH3-N 500 mL 2406549
Nitrogen, Ammonia Standard Solution, 2-mL PourRite Ampule, 50 mg/L 20/pkg 1479120
Nitrogen, Ammonia Standard Solution, 10-mL Voluette Ampules, 50 mg/L 16/pkg 1479110
®
Nitrogen, Ammonia Standard Solution, 10-mL Voluette Ampules, 150 mg/L 16/pkg 2128410
®
Pipet, TenSette , 1.0 to 10.0 mL each 1970010

Nitrogen, Total, Persulfate Digestion TNT Method (25.0 mg/L) 7

Consumables and replacement items (continued)
Description Unit Item no.

Pipet tips for TenSette Pipet 1970010 50/pkg 2199796

Pipet tips for TenSette Pipet 1970010 250/pkg 2199725
Pipet tips for TenSette Pipet 1970001 1000/pkg 2185628

FOR TECHNICAL ASSISTANCE, PRICE INFORMATION AND ORDERING: HACH COMPANY

In the U.S.A. – Call toll-free 800-227-4224 WORLD HEADQUARTERS
Outside the U.S.A. – Contact the HACH office or distributor serving you. Telephone: (970) 669-3050
On the Worldwide Web – www.hach.com; E-mail – [email protected] FAX: (970) 669-2932

© Hach Company/Hach Lange GmbH, 1989–2013. All rights reserved. 04/2013, Edition 8
Organic Carbon, Total DOC316.53.01093

Direct Method1 Method 10129

0.3 to 20.0 mg/L C (LR) Test ‘N Tube™ Vials
Scope and application: For water, drinking water and wastewater
1 U.S. Patent 6,368,870

Test preparation

Instrument specific information

The table in this section shows all of the instruments that have the program for this test.
Table 1 shows adapter and light shield requirements for the instruments that use them.
To use the table, select an instrument, then read across to find the corresponding
information for this test.
Table 1 Instrument-specific information for test tubes
Instrument Adapters Light shield
DR 6000 — —
DR 5000 — —
DR 900 4846400 Cover supplied with the instrument
DR 3900 — LZV849
DR 3800 — LZV646
DR 2800 —

Before starting
Install the instrument cap on the DR 900 cell holder before ZERO or READ is pushed.
DR 3900, DR 3800, DR 2800 and DR 2700: Install the light shield in Cell Compartment #2 before this test is started.
A reagent blank is required for each series of samples.
To test for higher concentrations of TOC, use Method 10173 or method 10128.
Review the Safety Data Sheets (MSDS/SDS) for the chemicals that are used and use any recommended personal protective
equipment.
Dispose of reacted solutions according to local, state and federal regulations. Use the Safety Data Sheets for disposal
information for unused reagents. Consult the environmental, health and safety staff for your facility and/or local regulatory
agencies for further disposal information.

Items to Collect
Description Quantity

Total Organic Carbon Direct Method Low Range Test 'N Tube Reagent Set 1
Cylinder, graduated, 10-mL 1
DRB200 Reactor 1
Flask, Erlenmeyer, 50-mL 1
Light shield and adapter (For information about sample cells, adapters or light shields, refer to
1
Instrument specific information on page 1.)
Magnetic stirrer 1

1
Items to Collect (continued)
Description Quantity

Paper, pH 1
®
Pipet, TenSette , 0.1- to 1.0-mL, with pipet tips 1
®
Pipet, TenSette , 1.0- to 10.0-mL, with pipet tips 1
Stir bar, magnetic 1
Test tube rack 1
Water, organic-free 3.0 mL
Wipes, disposable 1

Refer to Consumables and replacement items on page 7 for reorder information.

Sample collection
• Collect samples in clean glass bottles.
• Homogenize samples that contain solids to get a representative sample.
• Rinse the sample bottle several times with the sample to be collected.
• Fill the bottle completely full, then tighten the cap on the bottle.
• Analyze the samples as soon as possible for best results.
• Acid preservation is not recommended.

Test 'N Tube procedure

1. Start the 2. Add 10 mL of sample to 3. Add 0.4 mL of Buffer 4. Put the flask on a stir
DRB200 Reactor. Select the a 50-mL Erlenmeyer flask. Solution to the Erlenmeyer plate. Stir at a moderate
TOC program. Put the stir bar in the flask, pH 2.0. Use pH paper speed for 10 minutes.
Erlenmeyer flask. to make sure that the
sample pH is 2.

2 Organic Carbon, Total, Direct TNT Method (20.0 mg/L)

5. Put a label that says 6. Prepare the blank: Add 7. Prepare the sample: 8. Use deionized water to
"Reagent Blank" on one 3.0 mL of organic-free water Add 3.0 mL of sample from rinse two blue Low Range
Low Range Acid Digestion to the blank vial. the Erlenmeyer flask to the Indicator Ampules. Clean
vial. Put a lable that says sample vial. the ampules with a wipe. Do
"Sample" on a second Low not touch the sides of the
Range Acid Digestion vial. ampules after they are
Add the contents of one clean. Hold the ampules by
TOC Persulfate Powder the top.
Pillow to each Acid
Digestion Vial.

9. Put one unopened 10. Close the vials tightly. 11. Close the reactor. Let 12. After two hours, remove
ampule into each Acid Insert them into the reactor. the vials react for 2 hours at the vials from the reactor.
Digestion Vial. Snap the top 103 to 105 °C. Put them in a test tube rack
off of the ampule when the to cool for one hour. Make
score aligns with the top of sure that the vials stay in an
the vial. Let the ampules upright position at all times.
drop into the vials. The liquid in the blank
Do not invert or tilt the should show a dark blue
vials after the ampule is color.
inside.

Organic Carbon, Total, Direct TNT Method (20.0 mg/L) 3

 Start Zero

13. Start program 14. Clean the blank vial. 15. Insert the blank vial into 16. Push ZERO. The
427 Organic Carbon LR. the 16-mm cell holder. display shows 0.0 mg/L C.
For information about
sample cells, adapters or
light shields, refer to
Instrument specific
information on page 1.
Note: Although the program
name may vary between
instruments, the program
number does not change.

Read

17. Clean the sample vial. 18. Insert the sample vial 19. Push READ. Results
into the 16-mm cell holder. show in mg/L C.

Interferences
If the sample contains more than 600 mg/L CaCO3 alkalinity, add sulfuric acid to lower
the sample pH to less than 7, then start the test procedure.
Most sample turbidity is either dissolved during the digestion stage or settled during the
cooling period. Sample turbidities up to 50 NTU have been tested without interference.
The table that follows shows the substances that were tested for interference and did not
interfere up to the levels shown.
Interfering substance Interference level
Aluminum 10 mg/L Al
Ammonia Nitrogen 1000 mg/L as N
ASTM Wastewater No effect
Bromide 500 mg/L Br–
Bromine 25 mg/L Br2
Calcium 2000 mg/L as CaCO3
Chloride 500 mg/L Cl–
Chlorine 10 mg/L Cl2
Chlorine Dioxide 6 mg/L ClO2

4 Organic Carbon, Total, Direct TNT Method (20.0 mg/L)

Interfering substance Interference level
Copper 10 mg/L Cu
Cyanide 10 mg/L CN–
Iodide 50 mg/L I–
Iron (II) 10 mg/L Fe2+
Iron (III) 10 mg/L Fe3+
Magnesium 2000 mg/L as CaCO3
Manganese (VII) 1 mg/L Mn
Monochloramine 14 mg/L NH2Cl as Cl2
Nitrite 500 mg/L NO2-
Ozone 2 mg/L O3
Phosphate 3390 mg/L PO43-
Silica 100 mg/L SiO2
Sulfate 5000 mg/L SO42-
Sulfide 20 mg/L S2-
Sulfite 50 mg/L SO32-
Zinc 5 mg/L Zn

Reagent blank water

Water that is used for the reagent blank must contain less than 0.05 mg/L carbon. If the
organic-free water container is left open for extended periods, the water can absorb
carbon dioxide (CO2) from the atmosphere and contaminate the blank. To remove the
dissolved CO2 from the organic-free water, acidify the water and stir for 10 minutes as in
the test procedure.
Generally, water that is stored in plastic containers is not suitable for low-range TOC
blanks. Water that is stored in plastic can become contaminated with organic compounds
from the container walls. These leached organic compounds usually cannot be removed
by the acid-sparge process.
Accuracy check
Standard additions method (sample spike)
Use the standard additions method (for applicable instruments) to validate the test
procedure, reagents and instrument and to find if there is an interference in the sample.
Items to collect:
• TOC Standard Solution Ampule, 1000 mg/L C
• 100-mL volumetric flask, Class A
• 15-mL volumetric pipet and pipet filler
• Organic-free water
• Pipet, TenSette®, 0.1–1.0 mL and tips

1. Prepare a 150-mg/L total organic carbon standard solution as follows:

a. Use a pipet to add 15.00 mL of a 1000 mg/L TOC standard solution into the
volumetric flask.
b. Dilute to the mark with organic-free water. Mix well. Prepare this solution daily.
2. Use the test procedure to measure the concentration of the sample, then keep the
(unspiked) sample in the instrument.
3. Go to the Standard Additions option in the instrument menu.

Organic Carbon, Total, Direct TNT Method (20.0 mg/L) 5

 4. Select the values for standard concentration, sample volume and spike volumes.
5. Prepare three spiked samples: use the TenSette pipet to add 0.1 mL, 0.2 mL and
0.3 mL of the prepared standard solution, respectively, to three Acid Digestion Vials.
6. Add the contents of one TOC Persulfate Powder Pillow to each vial.
7. Add 3.0 mL of sample to each vial. Swirl to mix.
8. Use the test procedure to measure the concentration of each of the spiked samples.
Start with the smallest sample spike. Measure each of the spiked samples in the
instrument.
9. Select Graph to compare the expected results to the actual results.
Note: If the actual results are significantly different from the expected results, make sure that
the sample volumes and sample spikes are measured accurately. The sample volumes and
sample spikes that are used should agree with the selections in the standard additions menu. If
the results are not within acceptable limits, the sample may contain an interference.

Standard solution method

Use the standard solution method to validate the test procedure, reagents and
instrument.
Items to collect:
• TOC Standard Solution Ampule 1000 mg/L C
• 1-L volumetric flask, Class A
• 10-mL volumetric pipet, Class A and pipet filler
• Organic-free reagent water

1. Prepare a 10.0 mg/L C standard solution as follows:

a. Use a pipet to add 10.00 mL of 1000 mg/L total organic carbon standard solution
into the volumetric flask.
b. Dilute to the mark with organic-free reagent water. Mix well. Prepare this solution
daily.
2. Use the test procedure to measure the concentration of the prepared standard
solution.
3. Compare the expected result to the actual result.
Note: The factory calibration can be adjusted slightly with the standard adjust option so that the
instrument shows the expected value of the standard solution. The adjusted calibration is then
used for all test results. This adjustment can increase the test accuracy when there are slight
variations in the reagents or instruments.

Method performance
The method performance data that follows was derived from laboratory tests that were
measured on a spectrophotometer during ideal test conditions. Users may get different
results under different test conditions.
Program Standard Precision (95% Confidence Interval) Sensitivity
Concentration change per 0.010 Abs change
427 10.0 mg/L C 9.1–10.9 mg/L C 0.2 mg/L C

Summary of method
The total organic carbon (TOC) concentration is determined by first sparging the sample
under slightly acidic conditions to remove the inorganic carbon. In the outside vial,
organic carbon in the sample is digested by persulfate and acid to form carbon dioxide.
During digestion, the carbon dioxide diffuses into a pH indicator reagent in the inner
ampule. The absorption of carbon dioxide into the indicator forms carbonic acid. Carbonic
acid changes the pH and thus the color of the indicator solution. The amount of color
change is related to the original amount of carbon present in the sample. The
measurement wavelengths are 598 and 430 nm for spectrophotometers or 610 nm for
colorimeters.

6 Organic Carbon, Total, Direct TNT Method (20.0 mg/L)

Consumables and replacement items
Required reagents

Description Quantity/test Unit Item no.

pH Paper 1 5/pkg 39133

Water, Organic-free 3.0 mL 500 mL 2641549
Reagent Set, Total Organic Carbon Direct Method Low Range Test 'N
— 50 vials 2760345
Tube™
Includes:
Acid Digestion Solution Vials, Low Range TOC (not sold separately) 1 50/pkg —
Buffer Solution, Sulfate (not sold separately; see alternate size below) 0.4 mL 25 mL 45233
Funnel, micro, poly 1 each 2584335
Indicator Ampule, Low Range TOC (not sold separately) 1 10/pkg —
TOC Persulfate Powder Pillows (not sold separately) 1 50/pkg —

Required apparatus

Description Quantity/test Unit Item no.

Cylinder, graduated, 10-mL 1 each 50838

DRB200 Reactor, 110 V, 15 x 16 mm 1 each LTV082.53.40001
OR
DRB200 Reactor, 220 V, 15 x 16 mm 1 each LTV082.52.40001
Flask, Erlenmeyer, 50-mL 1 each 50541
Magnetic Stirrer 1 each 2881200
®
Pipet, TenSette , 0.1- to 1.0-mL 1 each 1970001
Pipet Tips, for TenSette Pipet 1970001 2 50/pkg 2185696
Pipet, TenSette 1.0–10.0 mL 1 each 1970010
Pipet Tips, for TenSette Pipet 1970010 varies 50/pkg 2199796
Stir bar, magnetic 1 each 4531500
Test tube rack 1 each 1864100
Wipes, Disposable 1 280/pkg 2097000

Recommended standards

Description Unit Item no.

TOC Standard Solution Ampule (KHP Standard, 1000-mg/L C) 5/pkg 2791505

Optional reagents and apparatus

Description Unit Item no.

Balance, analytical, 80 g x 0.1 mg 100-240 VAC each 2936701

Buffer Solution, Sulfate pH 2.0 500 mL 45249
Flask, volumetric, Class A, 1000-mL each 1457453
Flask, volumetric, Class A, 100-mL each 1457442
Paper, for weighing, 100 x 100 mm 500/pkg 1473885

Organic Carbon, Total, Direct TNT Method (20.0 mg/L) 7

Consumables and replacement items (continued)
Description Unit Item no.

Pipet filler, safety bulb each 1465100

Pipet tips for TenSette Pipet 1970001 1000/pkg 2185628
Pipet tips for TenSette Pipet 1970010 250/pkg 2199725
Pipet, volumetric Class A, 15-mL each 1451539
Pipet, volumetric, Class A, 10-mL each 1451538
Potassium Acid Phthalate, ACS 500 g 31534
Sulfuric Acid Standard Solution, 5.25 N 100 mL 244932

FOR TECHNICAL ASSISTANCE, PRICE INFORMATION AND ORDERING: HACH COMPANY

In the U.S.A. – Call toll-free 800-227-4224 WORLD HEADQUARTERS
Outside the U.S.A. – Contact the HACH office or distributor serving you. Telephone: (970) 669-3050
On the Worldwide Web – www.hach.com; E-mail – [email protected] FAX: (970) 669-2932

© Hach Company/Hach Lange GmbH, 1989–2013. All rights reserved. 04/2013, Edition 8
Organic Carbon, Total DOC316.53.01094

Direct Method1 Method 10173

15 to 150 mg/L C (MR) Test ‘N Tube™ Vials
Scope and application: For water, drinking water and wastewater
1 U.S. Patent 6,368,870

Test preparation

Instrument specific information

The table in this section shows all of the instruments that have the program for this test.
Table 1 shows adapter and light shield requirements for the instruments that use them.
To use the table, select an instrument, then read across to find the corresponding
information for this test.
Table 1 Instrument-specific information for test tubes
Instrument Adapters Light shield
DR 6000 — —
DR 5000 — —
DR 900 4846400 Cover supplied with the instrument
DR 3900 — LZV849
DR 3800 — LZV646
DR 2800 —

Before starting
Install the instrument cap on the DR 900 cell holder before ZERO or READ is pushed.
DR 3900, DR 3800, DR 2800 and DR 2700: Install the light shield in Cell Compartment #2 before this test is started.
A reagent blank is required for each series of samples.
To test for higher concentrations of TOC, use Method 10128. To test for lower concentrations of TOC, use Method 10129.
Review the Safety Data Sheets (MSDS/SDS) for the chemicals that are used and use any recommended personal protective
equipment.
Dispose of reacted solutions according to local, state and federal regulations. Use the Safety Data Sheets for disposal
information for unused reagents. Consult the environmental, health and safety staff for your facility and/or local regulatory
agencies for further disposal information.

Items to collect
Description Quantity

Total Organic Carbon Direct Method Mid Range Test 'N Tube Reagent Set 1
Cylinder, graduated, 10-mL 1
DRB200 Reactor 1
Flask, Erlenmeyer, 50-mL 1
Light shield and adapter (For information about sample cells, adapters or light shields, refer to
1
Instrument specific information on page 1.)
Magnetic stirrer 1

1
Items to collect (continued)
Description Quantity

Paper, pH 1
®
Pipet, TenSette , 0.1- to 1.0-mL, with pipet tips 1
®
Pipet, TenSette , 1.0- to 10.0-mL, with pipet tips 1
Stir bar, magnetic 1
Test tube rack 1
Water, organic-free 3.0 mL
Wipes, disposable 1

Refer to Consumables and replacement items on page 7 for reorder information.

Sample collection
• Collect samples in clean glass bottles.
• Homogenize samples that contain solids to get a representative sample.
• Rinse the sample bottle several times with the sample to be collected.
• Fill the bottle completely full, then tighten the cap on the bottle.
• Analyze the samples as soon as possible for best results.
• Acid preservation is not recommended.

Test 'N Tube procedure

1. Start the 2. Add 10 mL of sample to 3. Add 0.4 mL of Buffer 4. Put the flask on a stir
DRB200 Reactor. Select the a 50-mL Erlenmeyer flask. Solution to the Erlenmeyer plate. Stir at a moderate
TOC program. Put the stir bar in the flask, pH 2.0. Use pH paper speed for 10 minutes.
Erlenmeyer flask. to make sure that the
sample pH is 2.

2 Organic Carbon, Total, Direct TNT Method (150 mg/L)

5. Add the contents of one 6. Prepare the blank: Add 7. Prepare the sample: 8. Use deionized water to
TOC Persulfate Powder 1.0 mL of organic-free water Add 1.0 mL of sample from rinse two blue Mid/High
Pillow to each Acid to the blank vial. the Erlenmeyer flask to the Range Indicator Ampules.
Digestion Vial. Put a label sample vial. Clean the ampules with a
that says "Reagent Blank" wipe. Do not touch the sides
on one Mid Range Acid of the ampules after they
Digestion vial. Put a lable are clean. Hold the ampules
that says "Sample" on a by the top.
second Mid Range Acid
Digestion vial.

9. Put one unopened 10. Close the vials tightly. 11. Close the reactor. Let 12. After two hours, remove
ampule into each Acid Insert them into the reactor. the vials react for 2 hours at the vials from the reactor.
Digestion Vial. Snap the top 103 to 105 °C. Put them in a test tube rack
off of the ampule when the to cool for one hour. The
score aligns with the top of liquid in the blank should
the vial. Let the ampules show a dark blue color.
drop into the vials.
Do not invert or tilt the vials
after the ampule is inside.

Organic Carbon, Total, Direct TNT Method (150 mg/L) 3

 Start Zero

13. Start program 14. Clean the blank vial. 15. Insert the blank vial into 16. Push ZERO. The
425 Organic Carbon MR. the 16-mm cell holder. display shows 0 mg/L C.
For information about
sample cells, adapters or
light shields, refer to
Instrument specific
information on page 1.
Note: Although the program
name may vary between
instruments, the program
number does not change.

Read

17. Clean the sample vial. 18. Insert the sample vial 19. Push READ. Results
into the 16-mm cell holder. show in mg/L C.

Interferences
If the sample contains more than 1000 mg/L CaCO3 alkalinity, add sulfuric acid to lower
the sample pH to less than 7, then start the test procedure.
Most sample turbidity is either dissolved during the digestion stage or settled during the
cooling period. Sample turbidities up to 50 NTU have been tested without interference.
The table that follows shows the substances that were tested for interference and did not
interfere up to the levels shown.
Interfering substance Interference level
Aluminum 10 mg/L Al
Ammonia Nitrogen 1000 mg/L as N
ASTM Wastewater No effect
Bromide 500 mg/L Br–
Bromine 25 mg/L Br2
Calcium 2000 mg/L as CaCO3
Chloride 1500 mg/L Cl–
Chlorine 10 mg/L Cl2
Chlorine Dioxide 6 mg/L ClO2

4 Organic Carbon, Total, Direct TNT Method (150 mg/L)

Interfering substance Interference level
Copper 10 mg/L Cu
Cyanide 10 mg/L CN–
Iodide 50 mg/L I–
Iron (II) 10 mg/L Fe2+
Iron (III) 10 mg/L Fe3+
Magnesium 2000 mg/L as CaCO3
Manganese (VII) 1 mg/L Mn
Monochloramine 14 mg/L NH2Cl as Cl2
Nitrite 500 mg/L NO2-
Ozone 2 mg/L O3
Phosphate 3390 mg/L PO43-
Silica 100 mg/L SiO2
Sulfate 5000 mg/L SO42-
Sulfide 20 mg/L S2-
Sulfite 50 mg/L SO32-
Zinc 5 mg/L Zn

Reagent blank water

Water that is used for the reagent blank must contain less than 0.05 mg/L carbon. If the
organic-free water container is left open for extended periods, the water can absorb
carbon dioxide (CO2) from the atmosphere and contaminate the blank. To remove the
dissolved CO2 from the organic-free water, acidify the water and stir for 10 minutes as in
the test procedure.
Generally, water that is stored in plastic containers is not suitable for low-range TOC
blanks. Water that is stored in plastic can become contaminated with organic compounds
from the container walls. These leached organic compounds usually cannot be removed
by the acid-sparge process.
Accuracy check
Standard additions method (sample spike)
Use the standard additions method (for applicable instruments) to validate the test
procedure, reagents and instrument and to find if there is an interference in the sample.
Items to collect:
• TOC Standard Solution Ampule, 1000 mg/L C
• 50-mL volumetric flask, Class A
• 15-mL volumetric pipet and pipet filler
• Organic-free water
• Pipet, TenSette®, 0.1–1.0 mL and tips

1. Prepare a 300-mg/L total organic carbon standard solution as follows:

a. Use a pipet to add 15.00 mL of a 1000 mg/L TOC standard solution into the
volumetric flask.
b. Dilute to the mark with organic-free water. Mix well. Prepare this solution daily.
2. Use the test procedure to measure the concentration of the sample, then keep the
(unspiked) sample in the instrument.
3. Go to the Standard Additions option in the instrument menu.

Organic Carbon, Total, Direct TNT Method (150 mg/L) 5

 4. Select the values for standard concentration, sample volume and spike volumes.
5. Prepare three spiked samples: use the TenSette pipet to add 0.1 mL, 0.2 mL and
0.3 mL of the prepared standard solution, respectively, to three Acid Digestion Vials.
6. Add the contents of one TOC Persulfate Powder Pillow to each vial.
7. Add 1.0 mL of sample to each vial. Swirl to mix.
8. Use the test procedure to measure the concentration of each of the spiked samples.
Start with the smallest sample spike. Measure each of the spiked samples in the
instrument.
9. Select Graph to compare the expected results to the actual results.
Note: If the actual results are significantly different from the expected results, make sure that
the sample volumes and sample spikes are measured accurately. The sample volumes and
sample spikes that are used should agree with the selections in the standard additions menu. If
the results are not within acceptable limits, the sample may contain an interference.

Standard solution method

Use the standard solution method to validate the test procedure, reagents and
instrument.
Items to collect:
• TOC Standard Solution Ampule 1000 mg/L C
• 50-mL volumetric flask, Class A
• 5-mL volumetric pipet, Class A and pipet filler
• Organic-free reagent water

1. Prepare a 100 mg/L C standard solution as follows:

a. Use a pipet to add 5.00 mL of 1000 mg/L total organic carbon standard solution
into the volumetric flask.
b. Dilute to the mark with organic-free reagent water. Mix well. Prepare this solution
daily.
2. Use the test procedure to measure the concentration of the prepared standard
solution.
3. Compare the expected result to the actual result.
Note: The factory calibration can be adjusted slightly with the standard adjust option so that the
instrument shows the expected value of the standard solution. The adjusted calibration is then
used for all test results. This adjustment can increase the test accuracy when there are slight
variations in the reagents or instruments.

Method performance
The method performance data that follows was derived from laboratory tests that were
measured on a spectrophotometer during ideal test conditions. Users may get different
results under different test conditions.
Program Standard Precision (95% Confidence Interval) Sensitivity
Concentration change per 0.010 Abs change
425 70 mg/L C 68–72 mg/L C 1.2 mg/L C

Summary of method
The total organic carbon (TOC) concentration is determined by first sparging the sample
under slightly acidic conditions to remove the inorganic carbon. In the outside vial,
organic carbon in the sample is digested by persulfate and acid to form carbon dioxide.
During digestion, the carbon dioxide diffuses into a pH indicator reagent in the inner
ampule. The absorption of carbon dioxide into the indicator forms carbonic acid. Carbonic
acid changes the pH and thus the color of the indicator solution. The amount of color
change is related to the original amount of carbon in the sample. The measurement
wavelengths are 598 and 430 nm for spectrophotometers or 610 nm for colorimeters.

6 Organic Carbon, Total, Direct TNT Method (150 mg/L)

Consumables and replacement items
Required reagents

Description Quantity/test Unit Item no.

pH Paper 1 5/pkg 39133

Water, Organic-free 3.0 mL 500 mL 2641549
Reagent Set, Total Organic Carbon Direct Method Mid Range Test 'N
— 50 vials 2815945
Tube™
Includes:
Acid Digestion Solution Vials, High Range TOC (not sold separately) 1 50/pkg —
Buffer Solution (not sold separately; see alternate size below) 0.4 mL 25 mL —
Funnel, micro, poly 1 each 2584335
Indicator Ampule, MR/HR TOC (not sold separately) 1 10/pkg —
TOC Persulfate Powder Pillows (not sold separately) 1 50/pkg —

Required apparatus

Description Quantity/test Unit Item no.

Cylinder, graduated, 10-mL 1 each 50838

DRB200 Reactor, 110 V, 15 x 16 mm 1 each LTV082.53.40001
OR
DRB200 Reactor, 220 V, 15 x 16 mm 1 each LTV082.52.40001
Flask, Erlenmeyer, 50-mL 1 each 50541
Magnetic Stirrer 1 each 2881200
®
Pipet, TenSette , 0.1- to 1.0-mL 1 each 1970001
Pipet Tips, for TenSette Pipet 1970001 2 50/pkg 2185696
Pipet, TenSette 1.0–10.0 mL 1 each 1970010
Pipet Tips, for TenSette Pipet 1970010 varies 50/pkg 2199796
Stir bar, magnetic 1 each 4531500
Test tube rack 1 each 1864100
Wipes, Disposable 1 280/pkg 2097000

Recommended standards

Description Unit Item no.

TOC Standard Solution Ampule (KHP Standard, 1000-mg/L C) 5/pkg 2791505

Optional reagents and apparatus

Description Unit Item no.

Balance, analytical, 80 g x 0.1 mg 100-240 VAC each 2936701

Buffer Solution, Sulfate pH 2.0 500 mL 45249
Flask, volumetric, 50-mL each 1457441
Flask, volumetric, Class A, 1000-mL each 1457453
Pipet tips for TenSette Pipet 1970001 1000/pkg 2185628

Organic Carbon, Total, Direct TNT Method (150 mg/L) 7

Consumables and replacement items (continued)
Description Unit Item no.

Pipet tips for TenSette Pipet 1970010 250/pkg 2199725

Paper, for weighing, 100 x 100 mm 500/pkg 1473885
Pipet, volumetric Class A, 15-mL each 1451539
Pipet, volumetric, Class A, 5.00-mL each 1451537
Pipet filler, safety bulb each 1465100
Potassium Acid Phthalate, ACS 500 g 31534
Sulfuric Acid Standard Solution, 5.25 N 100 mL 244932

FOR TECHNICAL ASSISTANCE, PRICE INFORMATION AND ORDERING: HACH COMPANY

In the U.S.A. – Call toll-free 800-227-4224 WORLD HEADQUARTERS
Outside the U.S.A. – Contact the HACH office or distributor serving you. Telephone: (970) 669-3050
On the Worldwide Web – www.hach.com; E-mail – [email protected] FAX: (970) 669-2932

© Hach Company/Hach Lange GmbH, 1989–2013. All rights reserved. 04/2013, Edition 8
Organic Carbon, Total DOC316.53.01095

Direct Method1 Method 10128

100 to 700 mg/L C (HR, spectrophotometers) Test ‘N Tube™ Vials
20 to 700 mg/L C (HR, colorimeters)
Scope and application: For wastewater and industrial water
1 U.S. Patent 6,368,870

Test preparation

Instrument specific information

The table in this section shows all of the instruments that have the program for this test.
Table 1 shows adapter and light shield requirements for the instruments that use them.
To use the table, select an instrument, then read across to find the corresponding
information for this test.
Table 1 Instrument-specific information for test tubes
Instrument Adapters Light shield
DR 6000 — —
DR 5000 — —
DR 900 4846400 Cover supplied with the instrument
DR 3900 — LZV849
DR 3800 — LZV646
DR 2800 —

Before starting
Install the instrument cap on the DR 900 cell holder before ZERO or READ is pushed.
DR 3900, DR 3800, DR 2800 and DR 2700: Install the light shield in Cell Compartment #2 before this test is started.
A reagent blank is required for each series of samples.
To test for lower concentrations of TOC, use Method 10128 or Method 10129.
Review the Safety Data Sheets (MSDS/SDS) for the chemicals that are used and use any recommended personal protective
equipment.
Dispose of reacted solutions according to local, state and federal regulations. Use the Safety Data Sheets for disposal
information for unused reagents. Consult the environmental, health and safety staff for your facility and/or local regulatory
agencies for further disposal information.

Items to collect
Description Quantity

Total Organic Carbon Direct Method High Range Test 'N Tube Reagent Set 1
Cylinder, graduated, 10-mL 1
DRB200 Reactor 1
Flask, Erlenmeyer, 50-mL 1
Light shield or adapter (For information about sample cells, adapters or light shields, refer to
1
Instrument specific information on page 1.)
Magnetic stirrer 1

1
Items to collect (continued)
Description Quantity

Paper, pH 1
®
Pipet, TenSette , 0.1- to 1.0-mL, with pipet tips 1
®
Pipet, TenSette , 1.0- to 10.0-mL, with pipet tips 1
Stir bar, magnetic 1
Test tube rack 1
Water, organic-free 0.3 mL
Wipes, disposable 1

Refer to Consumables and replacement items on page 7 for reorder information.

Sample collection
• Collect samples in clean glass bottles.
• Homogenize samples that contain solids to get a representative sample.
• Rinse the sample bottle several times with the sample to be collected.
• Fill the bottle completely full, then tighten the cap on the bottle.
• Analyze the samples as soon as possible for best results.
• Acid preservation is not recommended.

Organic Carbon Total HR

1. Start the 2. Add 10 mL of sample to 3. Add 0.4 mL of Buffer 4. Put the flask on a stir
DRB200 Reactor. Select the a 50-mL Erlenmeyer flask. Solution to the Erlenmeyer plate. Stir at a moderate
TOC program. Put the stir bar in the flask, pH 2.0. Use pH paper speed for 10 minutes.
Erlenmeyer flask. to make sure that the
sample pH is 2.

2 Organic Carbon, Total, Direct TNT Method (700 mg/L)

5. Add the contents of one 6. Prepare the blank: Add 7. Prepare the sample: 8. Use deionized water to
TOC Persulfate Powder 0.3 mL of organic-free water Add 0.3 mL of sample from rinse two blue Mid/High
Pillow to each Acid to the blank vial. the Erlenmeyer flask to the Range Indicator Ampules.
Digestion Vial. Put a label sample vial. Clean the ampules with a
that says "Reagent Blank" wipe. Do not touch the sides
on one High Range Acid of the ampules after they
Digestion vial. Put a label are clean. Hold the ampules
that says "Sample" on a by the top.
second High Range Acid
Digestion vial.

9. Put one unopened 10. Close the vials tightly. 11. Close the reactor. Let 12. After two hours, remove
ampule into each Acid Insert them into the reactor. the vials react for 2 hours at the vials from the reactor.
Digestion Vial. Snap the top 103 to 105 °C. Keep the vials in an upright
off of the ampule when the position at all times. Put
score aligns with the top of them in a test tube rack to
the vial. Let the ampules cool for one hour. The liquid
drop into the vials. in the blank should show a
Do not invert or tilt the vials dark blue color.
after the ampule is inside.

Organic Carbon, Total, Direct TNT Method (700 mg/L) 3

 Start Zero

13. Start program 14. Clean the blank vial. 15. Insert the blank vial into 16. Push ZERO. The
426 Organic Carbon HR. the 16-mm cell holder. display shows 0 mg/L C.
For information about
sample cells, adapters or
light shields, refer to
Instrument specific
information on page 1.
Note: Although the program
name may vary between
instruments, the program
number does not change.

Read

17. Clean the sample vial. 18. Insert the sample vial 19. Push READ. Results
into the 16-mm cell holder. show in mg/L C.

Interferences
If the sample contains more than 1000 mg/L CaCO3 alkalinity, add sulfuric acid to lower
the sample pH to less than 7, then start the test procedure.
Most sample turbidity is either dissolved during the digestion stage or settled during the
cooling period. Sample turbidities up to 50 NTU have been tested without interference.
The table that follows shows the substances that were tested for interference and did not
interfere up to the levels shown.
Interfering substance Interference level
Aluminum 10 mg/L Al
Ammonia Nitrogen 1000 mg/L as N
ASTM Wastewater No effect
Bromide 500 mg/L Br–
Bromine 25 mg/L Br2
Calcium 2000 mg/L as CaCO3
Chloride 5000 mg/L Cl–
Chlorine 10 mg/L Cl2
Chlorine Dioxide 6 mg/L ClO2

4 Organic Carbon, Total, Direct TNT Method (700 mg/L)

Interfering substance Interference level
Copper 10 mg/L Cu
Cyanide 10 mg/L CN–
Iodide 50 mg/L I–
Iron (II) 10 mg/L Fe2+
Iron (III) 10 mg/L Fe3+
Magnesium 2000 mg/L as CaCO3
Manganese (VII) 1 mg/L Mn
Monochloramine 14 mg/L NH2Cl as Cl2
Nitrite 500 mg/L NO2-
Ozone 2 mg/L O3
Phosphate 3390 mg/L PO43-
Silica 100 mg/L SiO2
Sulfate 5000 mg/L SO42-
Sulfide 20 mg/L S2-
Sulfite 50 mg/L SO32-
Zinc 5 mg/L Zn

Reagent blank water

Water that is used for the reagent blank must contain less than 0.05 mg/L carbon. If the
organic-free water container is left open for extended periods, the water can absorb
carbon dioxide (CO2) from the atmosphere and contaminate the blank. To remove the
dissolved CO2 from the organic-free water, acidify the water and stir for 10 minutes as in
the test procedure.
Generally, water that is stored in plastic containers is not suitable for low-range TOC
blanks. Water that is stored in plastic can become contaminated with organic compounds
from the container walls. These leached organic compounds usually cannot be removed
by the acid-sparge process.
Accuracy check
Standard additions method (sample spike)
Use the standard additions method (for applicable instruments) to validate the test
procedure, reagents and instrument and to find if there is an interference in the sample.
Items to collect:
• TOC Standard Solution Ampule, 1000 mg/L C
• 50-mL volumetric flask, Class A
• 15-mL volumetric pipet and pipet filler
• Organic-free water
• Pipet, TenSette®, 0.1–1.0 mL and tips

1. Prepare a 300-mg/L total organic carbon standard solution as follows:

a. Use a pipet to add 15.00 mL of a 1000 mg/L TOC standard solution into the
volumetric flask.
b. Dilute to the mark with organic-free water. Mix well. Prepare this solution daily.
2. Use the test procedure to measure the concentration of the sample, then keep the
(unspiked) sample in the instrument.
3. Go to the Standard Additions option in the instrument menu.

Organic Carbon, Total, Direct TNT Method (700 mg/L) 5

 4. Select the values for standard concentration, sample volume and spike volumes.
5. Prepare three spiked samples: use the TenSette pipet to add 0.1 mL, 0.2 mL and
0.3 mL of the prepared standard solution, respectively, to three Acid Digestion Vials.
6. Add the contents of one TOC Persulfate Powder Pillow to each vial.
7. Add 0.3 mL of sample to each vial. Swirl to mix.
8. Use the test procedure to measure the concentration of each of the spiked samples.
Start with the smallest sample spike. Measure each of the spiked samples in the
instrument.
9. Select Graph to compare the expected results to the actual results.
Note: If the actual results are significantly different from the expected results, make sure that
the sample volumes and sample spikes are measured accurately. The sample volumes and
sample spikes that are used should agree with the selections in the standard additions menu. If
the results are not within acceptable limits, the sample may contain an interference.

Standard solution method

Use the standard solution method to validate the test procedure, reagents and
instrument.
Items to collect:
• TOC Standard Solution Ampule 1000 mg/L C
• 50-mL volumetric flask, Class A
• 15-mL volumetric pipet, Class A and pipet filler
• Organic-free reagent water

1. Prepare a 300 mg/L C standard solution as follows:

a. Use a pipet to add 15.00 mL of 1000 mg/L total organic carbon standard solution
into the volumetric flask.
b. Dilute to the mark with organic-free reagent water. Mix well. Prepare this solution
daily.
2. Use the test procedure to measure the concentration of the prepared standard
solution.
3. Compare the expected result to the actual result.
Note: The factory calibration can be adjusted slightly with the standard adjust option so that the
instrument shows the expected value of the standard solution. The adjusted calibration is then
used for all test results. This adjustment can increase the test accuracy when there are slight
variations in the reagents or instruments.

Method performance
The method performance data that follows was derived from laboratory tests that were
measured on a spectrophotometer during ideal test conditions. Users may get different
results under different test conditions.
Program Standard Precision (95% Confidence Interval) Sensitivity
Concentration change per 0.010 Abs change
426 350 mg/L C 337–363 mg/L C 4 mg/L C

Summary of method
The total organic carbon (TOC) concentration is determined by first sparging the sample
under slightly acidic conditions to remove the inorganic carbon. In the outside vial,
organic carbon in the sample is digested by persulfate and acid to form carbon dioxide.
During digestion, the carbon dioxide diffuses into a pH indicator reagent in the inner
ampule. The absorption of carbon dioxide into the indicator forms carbonic acid. Carbonic
acid changes the pH and thus the color of the indicator solution. The amount of color
change is related to the original amount of carbon in the sample. The measurement
wavelengths are 598 and 430 nm for spectrophotometers or 610 nm for colorimeters.

6 Organic Carbon, Total, Direct TNT Method (700 mg/L)

Consumables and replacement items
Required reagents

Description Quantity/test Unit Item no.

pH Paper 1 5/pkg 39133

Water, Organic-free 3.0 mL 500 mL 2641549
Total Organic Carbon Direct Method High Range Test ’N Tube™
— 50 vials 2760445
Reagent Set
Includes:
Acid Digestion Solution Vials, High Range TOC (not sold separately) 1 50/pkg —
Buffer Solution, Sulfate (not sold separately; see alternate size below) 0.4 mL 25 mL —
Funnel, micro, poly 1 each 2584335
Indicator Ampules, MR/HR TOC (not sold separately) 1 10/pkg —
TOC Persulfate Powder Pillows (not sold separately) 1 50/pkg —

Required apparatus

Description Quantity/test Unit Item no.

Cylinder, graduated, 10-mL 1 each 50838

DRB200 Reactor, 110 V, 15 x 16 mm 1 each LTV082.53.40001
OR
DRB200 Reactor, 220 V, 15 x 16 mm 1 each LTV082.52.40001
Flask, Erlenmeyer, 50-mL 1 each 50541
Magnetic Stirrer 1 each 2881200
®
Pipet, TenSette , 0.1- to 1.0-mL 1 each 1970001
Pipet Tips, for TenSette Pipet 1970001 2 50/pkg 2185696
Pipet, TenSette 1.0–10.0 mL 1 each 1970010
Pipet Tips, for TenSette Pipet 1970010 varies 50/pkg 2199796
Stir bar, magnetic 1 each 4531500
Test tube rack 1 each 1864100
Wipes, Disposable 1 280/pkg 2097000

Recommended standards

Description Unit Item no.

TOC Standard Solution Ampule (KHP Standard, 1000-mg/L C) 5/pkg 2791505

Optional reagents and apparatus

Description Unit Item no.

Balance, analytical, 80 g x 0.1 mg 100-240 VAC each 2936701

Buffer Solution, Sulfate pH 2.0 500 mL 45249
Flask, volumetric, 50-mL each 1457441
Flask, volumetric, Class A, 1000-mL each 1457453
Pipet, volumetric Class A, 15-mL each 1451539

Organic Carbon, Total, Direct TNT Method (700 mg/L) 7

Consumables and replacement items (continued)
Description Unit Item no.

Pipet filler, safety bulb each 1465100

Pipet tips for TenSette Pipet 1970001 1000/pkg 2185628
Pipet tips for TenSette Pipet 1970010 250/pkg 2199725
Potassium Acid Phthalate, ACS 500 g 31534
Sulfuric Acid Standard Solution, 5.25 N 100 mL 244932
Paper, for weighing, 100 x 100 mm 500/pkg 1473885

FOR TECHNICAL ASSISTANCE, PRICE INFORMATION AND ORDERING: HACH COMPANY

In the U.S.A. – Call toll-free 800-227-4224 WORLD HEADQUARTERS
Outside the U.S.A. – Contact the HACH office or distributor serving you. Telephone: (970) 669-3050
On the Worldwide Web – www.hach.com; E-mail – [email protected] FAX: (970) 669-2932

© Hach Company/Hach Lange GmbH, 1989–2013. All rights reserved. 04/2013, Edition 8
Oxygen Demand, Chemical DOC316.53.01099

USEPA1 Reactor Digestion Method2 Method 8000

0.7 to 40.03 mg/L COD (ULR); 3 to 150 mg/L COD (LR); 20 to 1500 mg/L
COD (HR); 200 to 15,000 mg/L COD (HR Plus)
Scope and application: For water and wastewater. Digestion is required.
1 Ranges 3 to 150 mg/L COD and 20 to 1500 mg/L COD are USEPA approved for wastewater analyses (Standard Method 5220 D), Federal
Register, April 21, 1980, 45(78), 26811-26812.
2 Jirka, A.M.; Carter, M.J., Analytical Chemistry, 1975, 47(8), 1397.
3 The ULR is only available with spectrophotometers that can measure at a wavelength of 350 nm.

Test preparation

Instrument specific information

The table in this section shows all of the instruments that have the program for this test.
Table 1 shows adapter and light shield requirements for the instruments that use them.
To use the table, select an instrument, then read across to find the corresponding
information for this test.
Table 1 Instrument-specific information for test tubes
Instrument Adapters Light shield
DR 6000 — —
DR 5000 — —
DR 900 4846400 Cover supplied with the instrument
DR 3900 — LZV849
DR 3800 — LZV646
DR 2800 —

Before starting
Install the instrument cap on the DR 900 cell holder before ZERO or READ is pushed.
DR 3900, DR 3800, DR 2800 and DR 2700: Install the light shield in Cell Compartment #2 before this test is started.
The reagent that is used in this test is corrosive and toxic. Use protection for eyes and skin and be prepared to flush any
spills with running water.
The reagents that are used in this test contain mercury. Collect the reacted samples for proper disposal.
Review the Safety Data Sheets (MSDS/SDS) for the chemicals that are used and use any recommended personal protective
equipment.
Run one blank with each set of samples. Run all tests (the samples and the blank) with the same lot of vials. The lot number
is on the container label. Refer to Blanks for colorimetric determination on page 4.
Store unused (light sensitive) vials in a closed box.
If the samples contain high concentrations of chloride, refer to the Alternate reagents section.
Dispose of reacted solutions according to local, state and federal regulations. Use the Safety Data Sheets for disposal
information for unused reagents. Consult the environmental, health and safety staff for your facility and/or local regulatory
agencies for further disposal information.

1
Items to collect
Description Quantity

Beaker, 250-mL 1
Blender 1
COD Digestion Reagent vials varies
DRB200 Reactor 1
Light shield or adapter (For information about sample cells, adapters or light shields, refer to
1
Instrument specific information on page 1.)
Magnetic stirrer and stir bar 1
Opaque shipping container for storage of unused, light-sensitive reagent vials varies
Pipet, TenSette, 0.1- to 1.0-mL, with pipet tips (for use with the 200–15,000 mg/L range) 1
Pipet, volumetric, 2.00-mL 2
Pipet filler safety bulb 1
Test tube rack 2

Refer to Consumables and replacement items on page 7 for reorder information.

Sample collection and storage

• Collect samples in clean glass bottles. Use plastic bottles only if they are known to be
free of organic contamination.
• Test biologically active samples as soon as possible.
• Homogenize samples that contain solids to get a representative sample.
• To preserve samples for later analysis, adjust the sample pH to less than 2 with
concentrated sulfuric acid (about 2 mL per liter). No acid addition is necessary if the
sample is tested immediately.
• Keep the preserved samples at or below 6 °C (43 °F) for up to 28 days.
• Correct the test result for the dilution from the volume additions.

Reactor digestion procedure

1. Put 100 mL of sample in 2. For the 200–15,000 mg/L 3. Set the DRB200 Reactor 4. Prepare the sample:
a blender. Blend for range or to improve power to on. Preheat to Remove the cap from a vial
30 seconds or until accuracy and reproducibility 150 °C. for the selected range. Hold
homogenized. of the other ranges, pour the Refer to the DRB200 User the vial at an angle of
For samples with large homogenized sample into a Manual for selecting pre- 45 degrees. Use a clean
amounts of solids, increase 250-mL beaker and gently programmed temperature pipet to add 2.00 mL of
the homogenization time. If stir with a magnetic stir applications. sample to the vial.
the sample does not contain plate. For 250–15,000 mg/L vials:
suspended solids, omit Use a TenSette Pipet to add
steps 1 and 2. 0.20 mL of sample to the
vial.

2 Oxygen Demand, Chemical, Dichromate Method (multi-range: 40.0, 150, 1500, 15,000 mg/L)
5. Prepare the blank: 6. Close the vials tightly. 7. Hold the vials by the cap, 8. Put the vials in the
Remove the cap from a Rinse the vials with water over a sink. Invert gently preheated DRB200 reactor.
second vial for the selected and wipe with a clean paper several times to mix. Close the lid.
range. Hold the vial at an towel. The vials get very hot
angle of 45 degrees. Use a during mixing.
clean pipet to add 2.00 mL
of deionized water to the
vial.
For 250–15,000 mg/L vials:
Use a TenSette Pipet to add
0.20 mL of deionized water
to the vial.

9. Heat the vials for 10. Set the reactor power to 11. Invert each vial several 12. Put the vials in a tube
2 hours. off. Let the vials cool in the times while it is still warm. rack to cool to room
reactor for about temperature.
20 minutes. The vials should
cool to 120 °C or less.

Oxygen Demand, Chemical, Dichromate Method (multi-range: 40.0, 150, 1500, 15,000 mg/L) 3
Colorimetric procedure

Start Zero

1. Start program 431 COD 2. Clean the blank. 3. Insert the blank into the 4. Push ZERO. The display
ULR, 430 COD LR or cell holder. shows 0 or 0.0 mg/L COD.
435 COD HR. For
information about sample
cells, adapters or light
shields, refer to Instrument
specific information
on page 1.
Note: Although the program
name may vary between
instruments, the program
number does not change.

Read

5. Clean the prepared 6. Insert the prepared 7. Push READ. Results 8. If using High Range Plus
sample. sample into the cell holder. show in mg/L COD. COD digestion reagent
vials, multiply the result by
10. For the most accurate
results with samples near
1500 or 15,000 mg/L COD,
repeat the analysis with a
diluted sample.

Blanks for colorimetric determination

The blank vial can be used again and again for measurements that use the same lot of
reagent vials. Measure the absorbance of the blank vial over time and prepare a new
blank vial when the absorbance changes.

1. Put the instrument in the absorbance mode at the applicable wavelength. Refer to
Table 3 on page 7.
2. Add 5 mL of deionized water into an empty vial.
3. Put the vial in the instrument and zero the instrument.
4. Put the blank vial that is used in the test procedure into the instrument and record the
absorbance value.
5. Keep the blank vial in the dark.
6. Prepare a new blank when the absorbance has changed by approximately
0.01 absorbance units.

4 Oxygen Demand, Chemical, Dichromate Method (multi-range: 40.0, 150, 1500, 15,000 mg/L)
Interferences
Chloride is the primary interference in this test procedure. Each COD vial contains
mercuric sulfate that removes chloride interference to the level specified in Column 1 of
Table 2. Dilute samples that have higher chloride concentrations to the level given in
Column 2.
Note: For best results, use the low range and ultra-low range vials for samples that have high
chloride concentrations (near maximum concentration) and low COD concentrations.
If sample dilution causes the COD concentration to be too low for accurate
measurements, add 0.50 g of mercuric sulfate (HgSO4) to each COD vial before the
sample is added. The additional mercuric sulfate will increase the maximum chloride
concentration to the level given in Column 3.
Note: Bromide interference is not removed with mercuric sulfate.
Table 2 Chloride concentration limits in the sample
Vial range Column 1 (maximum mg/L Column 2 (mg/L Cl– for Column 3 (maximum mg/L
Cl–) diluted samples) Cl– with mercuric sulfate)
ULR1 (0.7–40.0 mg/L) 2000 1000 N/A
LR (3–150 mg/L) 2000 1000 8000
HR (20–1500 mg/L) 2000 1000 4000
HR Plus (200–15,000 mg/L) 20,000 10,000 40,000
1 The ULR is only available for spectrophotometers that can measure at a wavelength of 350 nm.

Accuracy check
Standard solution method
Items to collect:
• 1000 mg/L COD standard solution
• 100-mL volumetric flask, Class A
• Volumetric pipets, Class A and pipet filler
• Deionized water
• Potassium acid phthalate (KHP), dried overnight at 120 °C (HR Plus only)
0.7 to 40.0 mg/L ULR
1. Prepare a 30-mg/L COD standard solution as follows:
a. Use a pipet to add 3.00 mL of the 1000 mg/L standard solution into a 100-mL
volumetric flask.
b. Dilute to the mark with deionized water. Mix well.
2. Use the test procedure to measure the concentration of the standard solution.
3. Compare the expected result to the actual result.
Note: The factory calibration can be adjusted slightly with the standard adjust option so that the
instrument shows the expected value of the standard solution. The adjusted calibration is then
used for all test results. This adjustment can increase the test accuracy when there are slight
variations in the reagents or instruments.

Oxygen Demand, Chemical, Dichromate Method (multi-range: 40.0, 150, 1500, 15,000 mg/L) 5
 3 to 150 mg/L LR
1. Prepare a 100-mg/L COD standard solution as follows:
a. Use a pipet to add 10 mL of the 1000 mg/L standard solution into a 100-mL
volumetric flask.
b. Dilute to the mark with deionized water. Mix well.
2. Use the test procedure to measure the concentration of the standard solution.
3. Compare the expected result to the actual result.
Note: The factory calibration can be adjusted slightly with the standard adjust option so that the
instrument shows the expected value of the standard solution. The adjusted calibration is then
used for all test results. This adjustment can increase the test accuracy when there are slight
variations in the reagents or instruments.

20 to 1500 mg/L HR
1. Use the test procedure with a 300-mg/L, 800 mg/L or 1000 mg/L COD standard
solution to measure the concentration of the standard solution.
2. Compare the expected result to the actual result.
Note: The factory calibration can be adjusted slightly with the standard adjust option so that the
instrument shows the expected value of the standard solution. The adjusted calibration is then
used for all test results. This adjustment can increase the test accuracy when there are slight
variations in the reagents or instruments.

200 to 15,000 mg/L HR Plus

1. Prepare a 10,000 mg/L COD standard solution as follows:
a. Dissolve 8.500 g of dried KHP in 1000-mL of organic-free deionized water.
2. Use the test procedure to measure the concentration of the standard solution.
3. Compare the expected result to the actual result.
Note: The factory calibration can be adjusted slightly with the standard adjust option so that the
instrument shows the expected value of the standard solution. The adjusted calibration is then
used for all test results. This adjustment can increase the test accuracy when there are slight
variations in the reagents or instruments.

Alternate reagents
Mercury-free COD2 Reagents are available as a mercury-free alternative. These
reagents are fully compatible with test procedures and stored programs in the
instruments. Chloride and ammonia determinations are recommended for accurate
results.

NOTICE
COD2 reagents are not approved for USEPA reporting purposes. Because COD2 reagents do not
contain mercury as a masking agent, they exhibit a positive interference from chloride. More
information is available for use with specific applications.

Method performance
The method performance data that follows was derived from laboratory tests that were
measured on a spectrophotometer during ideal test conditions. Users may get different
results under different test conditions.
Program Standard Precision (95% Confidence Interval) Sensitivity
Concentration change per 0.010 Abs change
431 (ULR) 30 mg/L COD 28.8–31.2 mg/L COD 0.5 mg/L COD
430 (LR) 80 mg/L COD 77–83 mg/L COD 3 mg/L COD
435 (HR) 800 mg/L COD 785–815 mg/L COD 23 mg/L COD
435 (HR Plus) 8000 mg/L COD 7850–8150 mg/L COD 230 mg/L COD

6 Oxygen Demand, Chemical, Dichromate Method (multi-range: 40.0, 150, 1500, 15,000 mg/L)
Summary of method
The results in mg/L COD are defined as the milligrams of O2 consumed per liter of
sample under the conditions of this procedure. The sample is heated for 2 hours with
sulfuric acid and a strong oxidizing agent, potassium dichromate. Oxidizable organic
compounds react, reducing the dichromate ion (Cr2O72–) to green chromic ion (Cr3+).
When the 0.7–40.0 or the 3–150 mg/L colorimetric method is used, the amount of Cr6+
that remains is measured. When the 20–1500 mg/L or 200–15,000 mg/L colorimetric
method is used, the amount of Cr3+ that is produced is measured. The COD reagent also
contains silver and mercury ions. Silver is a catalyst, and mercury is used to complex
chloride interferences.
Test results are measured at the wavelengths that are specified in Table 3.
Table 3 Range-specific test wavelengths
Range in mg/L COD Wavelength
0.7 to 40.0 mg/L 350 nm (for applicable instruments)
3 to 150 mg/L 420 nm
20 to 1500 620 nm (610 nm for colorimeters)
2000 to 15,000 mg/L 620 nm (610 nm for colorimeters)

Pollution prevention and waste management

Reacted samples contain mercury, silver and chromium and must be disposed of as a
hazardous waste. Dispose of reacted solutions according to local, state and federal
regulations. Users in the United States can use the ez COD Recycling Service for
disposal of COD vials. Refer to Consumables and replacement items on page 7.
Consumables and replacement items
Required reagents

Description Quantity/test Unit Item no.

COD, Ultra Low Range, 0.7 to 40 mg/L 1-2 vials 25/pkg 2415825
COD, Low Range, 3 to 150 mg/L 1-2 vials 25/pkg 2125825
COD, High Range, 20 to 1500 mg/L 1-2 vials 25/pkg 2125925
COD, High Range Plus, 200 to 15,000 mg/L 1-2 vials 25/pkg 2415925
Water, deionized varies 4L 27256

Alternate reagents and package sizes

Description Quantity/test Unit Item no.

COD2, Low Range, 0 to 150 mg/L COD 1-2 vials 25/pkg 2565025
COD2, High Range, 0 to 1500 mg/L COD 1-2 vials 25/pkg 2565125
COD2, High Range, 0 to 1500 mg/L COD 1-2 vials 150/pkg 2565115
COD2, High Range Plus, 0 to 15,000 mg/L COD 1-2 vials 25/pkg 2834325
COD Digestion Reagent Vials, 3 to 150 mg/L COD 1-2 vials 150/pkg 2125815
COD Digestion Reagent Vials, 200 to 1500 mg/L COD 1-2 vials 150/pkg 2125915
COD Digestion Reagent Vials, ULR 0.7-40.0 mg/L 1-2 vials 150/pkg 2415815
COD Digestion Reagent Vials, HR plus,200-25,000 mg/L 1-2 vials 150/pkg 2415915

Oxygen Demand, Chemical, Dichromate Method (multi-range: 40.0, 150, 1500, 15,000 mg/L) 7
Required apparatus

Description Quantity/test Unit Item no.

Blender, 2-speed, 120 VAC 1 each 2616100

OR
Blender, 2-speed, 240 VAC 1 each 2616102
DRB200 Reactor, 110 V, 15 x 16 mm 1 each LTV082.53.40001
OR
DRB200 Reactor, 220 V, 15 x 16 mm 1 each LTV082.52.40001
Pipet filler, safety bulb 1 each 1465100
Pipet, volumetric, Class A, 2.00-mL 1 each 1451536

Recommended standards and apparatus

Description Unit Item no.

Beaker, 250-mL each 50046H

COD Standard Solution, 300-mg/L 200 mL 1218629
COD Standard Solution, 300-mg/L 500mL 1218649
COD Standard Solution, 800-mg/L 200 mL 2672629
COD Standard Solution, 1000-mg/L 200 mL 2253929
Oxygen Demand Standard (BOD, COD, TOC), 10-mL ampules 16/pkg 2833510
®
Pipet, TenSette , 0.1–1.0 mL each 1970001
Pipet tips for TenSette Pipet 1970001 50/pkg 2185696
Pipet tips for TenSette Pipet 1970001 1000/pkg 2185628
Potassium Acid Phthalate, ACS 500 g 31534
Stir bar, octagonal each 2095352
Stirrer, electromagnetic, 120 VAC, with electrode stand each 4530001
Stirrer, electromagnetic, 230 VAC, with electrode stand each 4530002
Test tube rack each 1864100
Wipes, disposable 70/pkg 2096900

Optional reagents and apparatus

Description Unit Item no.

Balance, analytical, 80 g x 0.1 mg 100-240 VAC each 2936701

Flask, volumetric, Class A, 1000-mL each 1457453
Flask, volumetric, Class A, 100-mL each 1457442
Mercuric Sulfate 28 g 191520
Pipet, volumetric, Class A, 3-mL each 1451503
Pipet, volumetric, Class A, 10-mL each 1451538
Sulfuric Acid, ACS 500 mL 97949
Wastewater Influent Standard, Mixed Parameter, for NH3-N, NO3-N, PO4, COD, SO4,
500 mL 2833149
TOC

8 Oxygen Demand, Chemical, Dichromate Method (multi-range: 40.0, 150, 1500, 15,000 mg/L)
Consumables and replacement items (continued)
Description Unit Item no.

EZ COD™ Recycling Service with 5-gal bucket-mail back option (For US customers only.
each 2895405
20 and 55 gallon sizes are also available. )
EZ COD™ Recycling Service with 5-gal bucket- pick up option. (For US customers only.
each 2895405P
20 and 55 gallon sizes are also available. )
Finger cots 2/pkg 1464702
Gloves, chemical resistant, size 9-9.5 pair 24101041
Paper, for weighing, 100 x 100 mm 500/pkg 1473885
Safety goggles, vented each 2550700
Wastewater, Effluent Inorganics, for NH3-N, NO3-N, PO4, COD, SO4, TOC 500 mL 2833249
1 Other sizes available

Oxygen Demand, Chemical, Dichromate Method (multi-range: 40.0, 150, 1500, 15,000 mg/L) 9
 FOR TECHNICAL ASSISTANCE, PRICE INFORMATION AND ORDERING: HACH COMPANY
In the U.S.A. – Call toll-free 800-227-4224 WORLD HEADQUARTERS
Outside the U.S.A. – Contact the HACH office or distributor serving you. Telephone: (970) 669-3050
On the Worldwide Web – www.hach.com; E-mail – [email protected] FAX: (970) 669-2932

© Hach Company/Hach Lange GmbH, 1989–2013. All rights reserved. 04/2013, Edition 8
Oxygen Demand, Chemical DOC316.53.01101

Manganese III Reactor Digestion Method (with chloride removal) Method 10067
30 to 1000 mg/L COD Mn Test ‘N Tube™ Vials
Scope and application: For water and wastewater

Test preparation

Instrument specific information

The table in this section shows all of the instruments that have the program for this test.
Table 1 shows adapter and light shield requirements for the instruments that use them.
To use the table, select an instrument, then read across to find the corresponding
information for this test.
Table 1 Instrument-specific information for test tubes
Instrument Adapters Light shield
DR 6000 — —
DR 5000 — —
DR 900 4846400 Cover supplied with the instrument
DR 3900 — LZV849
DR 3800 — LZV646
DR 2800 —

Before starting
Install the instrument cap on the DR 900 cell holder before ZERO or READ is pushed.
DR 3900, DR 3800, DR 2800 and DR 2700: Install the light shield in Cell Compartment #2 before this test is started.
To find if the sample contains chloride, use Quantab® Titrator Strips for low range chloride.
If the sample COD is expected to exceed 1000 mg/L, dilute the sample. Refer to Multiplication factors for sample dilutions
on page 6.
Run one blank with each set of samples. Run all tests (the samples and the blank) with the same lot of vials. The lot number
is on the container label.
The reagent blank vial can be used for multiple tests. Fill a clean COD vial with deionized water and use this vial to zero the
instrument, then measure the absorbance of the reagent blank vial. The absorbance value should be approximately 1.41–
1.47. Prepare a new reagent blank vial when the absorbance is outside of this range.
To oxidize resistant organics, samples can be digested for up to four hours. Digest the blank for the same time period as the
samples.
Make sure that the filter disc is not in the center of the vial during the Zero and Read steps. Make sure that the filter disc is
more than 20 mm (0.8 in.) or less than 10 mm (0.4 in.) from the bottom of the vial. If necessary, move the filter disk by gently
swirling or by lightly tapping the vial on a table top.
The Chloride Removal Cartridge can be used only once.
If the sample boils during the digestion, the vial is not properly sealed. Test results will be invalid.
Spilled reagent will affect test accuracy and is hazardous. Do not run tests with spilled vials.
The maximum range of the VPD gauge is 40 inches of water; it will not indicate the full vacuum level obtained. Full vacuum
is 20–25 inches of mercury; this can be measured at the vacuum pump with a gauge calibrated for inches of mercury.

1
Review the Safety Data Sheets (MSDS/SDS) for the chemicals that are used and use any recommended personal protective
equipment.
Dispose of reacted solutions according to local, state and federal regulations. Use the Safety Data Sheets for disposal
information for unused reagents. Consult the environmental, health and safety staff for your facility and/or local regulatory
agencies for further disposal information.

Items to collect
Description Quantity

Blender 1
DRB200 Reactor 1
Forceps, extra fine 1
Light shield and adapter (For information about sample cells, adapters or light shields, refer to
1
Instrument specific information on page 1.)
Manganese III COD Reagent Vials, 20 to 1000 mg/L COD 1
Pipet, TenSette, 0.1- to 1.0-mL, with pipet tips 1
Pipet, TenSette, 10.0 to 10.0-mL, with pipet tips 1
Sulfuric Acid, concentrated ACS 1 mL
Test tube rack 1
Vacuum pretreatment device 1
Vacuum pump 1
Vial, glass, for sample and acid 2
Water, deionized varies

Refer to Consumables and replacement items on page 7 for reorder information.

Sample collection and storage

• Collect samples in clean glass bottles. Use plastic bottles only if they are known to be
free of organic contamination.
• Test biologically active samples as soon as possible.
• Homogenize samples that contain solids to get a representative sample.
• To preserve samples for later analysis, adjust the sample pH to less than 2 with
concentrated sulfuric acid (about 2 mL per liter). No acid addition is necessary if the
sample is tested immediately.
• Keep the preserved samples at or below 6 °C (43 °F) for up to 28 days.
• Correct the test result for the dilution from the volume additions.

2 Oxygen Demand, Chemical, Manganese III Method with chloride removal (1000 mg/L)
Acidified sample preparation

1. Set the DRB200 Reactor 2. Put 100 mL of sample in 3. Prepare the blank: Use 4. Prepare the sample:
power to on. Preheat to a blender. Blend for a pipet to add 9.0 mL of Use a pipet to add 9.0 mL of
150 °C or set to the COD 30 seconds or until deionized water to a glass homogenized sample to a
program. homogenized. mixing cell. second glass mixing cell.
If suspended solids are
present, continue to mix the
sample while the sample is
moved to the mixing cell for
the prepared sample.

5. Use a pipet or dispenser 6. Close the mixing cells

to add 1.0 mL of tightly. Invert several times.
concentrated Sulfuric Acid The cells get hot during
to both mixing cells. mixing. Let the mixing cells
cool to room temperature.
Go to the Vacuum
pretreatment procedure
on page 4.

Oxygen Demand, Chemical, Manganese III Method with chloride removal (1000 mg/L) 3
Vacuum pretreatment procedure

1. Attach the vacuum 2. Write a sample identifier 3. Put the VPD top on the 4. Start the vacuum pump.
pretreatment device (VPD) on each vial. Insert the Mn base. Insert a fresh Chloride Adjust the vacuum regulator
to a vacuum pump that can III COD vials in the Removal Cartridge (CRC) valve on top of the VPD until
create a vacuum of 20– numbered holes in the VPD directly above each Mn III the internal gauge reads
25 inches of mercury. base. Remove the caps COD Reagent Vial. Close 20 inches of water.
Do not use an aspirator type from the vials. any open holes in the VPD
vacuum. with the supplied stoppers.

5. Prepare the blank: Use 6. Prepare the sample: 7. Close the vacuum 8. Open the VPD regulator
a pipet to add 0.60 mL of Use a pipet to add 0.60 mL regulator valve completely to release the vacuum. Turn
acidified blank into the CRC. of each acidified sample into to achieve full vacuum. After the pump off. Remove the
It should take 30– the CRC. one minute of full vacuum, VPD top and set it aside.
45 seconds to pull the liquid move the VPD back and Dispose of the used
through the CRC into the forth several times to Chloride Removal
vial. remove drops that cling to Cartridges.
Note: If the liquid does not the CRC. Go to Sample preparation
flow through the CRC, and measurement
increase the vacuum until on page 5.
the flow starts, then reduce
the vacuum back to
20 inches of water.

4 Oxygen Demand, Chemical, Manganese III Method with chloride removal (1000 mg/L)
Sample preparation and measurement

1. Use forceps to remove 2. Insert each filter into the 3. Remove the Mn III COD 4. Insert the vials in the
the filter from the top of the corresponding Mn III COD vial from the vacuum DRB200 Reactor at 150 °C.
CRC. vial. Use the numbers on chamber. Put the original Close the cover. Digest the
If the sample does not the VPD as a guide. Use a caps on and close tightly. samples for one hour.
contain suspended solids, it clean towel or deionized Invert the vials several times
is not necessary to transfer water to clean the forceps to mix.
the filter to the digestion vial. between samples.

Start

5. After an hour, remove 6. Let the vials cool to room 7. Start program 432 COD 8. Clean the blank vial.
the vials from the DRB200. temperature in a cool water Mn III. For information about
Let the vials cool in a rack bath, or hold under running sample cells, adapters or
for two minutes. tap water for several light shields, refer to
If the solution develops a minutes. Instrument specific
colorless upper layer and a information on page 1.
purple lower layer, invert the Note: Although the program
vials several times to mix. name may vary between
instruments, the program
number does not change.

Zero

9. Insert the blank vial into 10. Push ZERO. The 11. Clean the sample vial. 12. Insert the sample vial
the 16-mm cell holder. Make display shows 0 mg/L COD into the 16-mm cell holder.
sure that the filter disc does Mn. Make sure that the filter disc
not block the instrument does not block the
light beam. Refer to Before instrument light beam. Refer
starting on page 1. to Before starting on page 1.

Oxygen Demand, Chemical, Manganese III Method with chloride removal (1000 mg/L) 5
 Read

13. Push READ. Results

show in mg/L COD Mn.

Multiplication factors for sample dilutions

If the sample COD is expected to exceed 1000 mg/L, dilute the sample as shown in
Table 2. For other dilutions not shown, add the sample volume and the deionized water
and divide by the sample volume to obtain the multiplication factor. All dilutions require
that the ratio of sample to sulfuric acid remain at 9:1.
Note: Mixing concentrated sulfuric acid and water is not additive. Adding 1.0 mL of concentrated
sulfuric acid to 9.0 mL of sample does not result in a final volume of 10.0 mL. This factor is built into
the calibration curve.
Table 2 Multiplication factors
Sample (mL) Deionized water (mL) Range (mg/L COD) Multiplication factor
6.0 3.0 30–1500 1.5
3.0 6.0 60–3000 3
1.0 8.0 180–9000 9
0.5 8.5 360–18,000 18

For best results, use a minimum of 0.5 mL sample for the dilution. If the sample values
exceed 18,000 mg/L COD, use a separate sample dilution, then start the sample chloride
removal procedure.
Example: Dilute the sample to a range of 90–4500 mg/L COD.
Sample Volume (2.0 mL) + Deionized water (7.0 mL) = Total Volume (9.0 mL)
Multiplication factor = (total volume)/(sample volume) = 9.0 mL/2.0 mL = 4.5
Standard test range is 50 to 1000 mg/L COD.
Example test range = 4.5(50) to 4.5(1000) = 225 to 4500 mg/L COD
Interferences
Inorganic materials may also be oxidized by trivalent manganese and constitute a positive
interference when present in significant amounts. Chloride is the most common
interference and is removed by sample pretreatment with the Chloride Removal
Cartridge. If chloride is known to be absent or present in insignificant levels, the
pretreatment can be omitted. A simple way to examine if chloride will affect test results is
to run routine samples with and without the chloride removal, then compare results. Other
inorganic interferences (i.e., nitrite, ferrous iron, sulfide) are not usually present in
significant amounts. If necessary, these interferences can be corrected after finding their
concentrations with separate methods and adjusting the final COD test results
accordingly.
Ammonia nitrogen is known to interfere in the presence of chloride; it does not interfere if
chloride is absent.

6 Oxygen Demand, Chemical, Manganese III Method with chloride removal (1000 mg/L)
Accuracy check
Standard solution method
Use the standard solution method to validate the test procedure, reagents and
instrument.
Items to collect:
• COD standard solution, 800 mg/L (use 0.60 mL in place of the sample)

1. Use the test procedure to measure the concentration of the standard solution.
2. Compare the expected result to the actual result.
Note: The factory calibration can be adjusted slightly with the standard adjust option so that the
instrument shows the expected value of the standard solution. The adjusted calibration is then
used for all test results. This adjustment can increase the test accuracy when there are slight
variations in the reagents or instruments.

Method performance
The method performance data that follows was derived from laboratory tests that were
measured on a spectrophotometer during ideal test conditions. Users may get different
results under different test conditions.
Program Standard Precision (95% Confidence Interval) Sensitivity
Concentration change per 0.010 Abs change
432 600 mg/L COD 576–624 mg/L COD 8 mg/L COD

Estimated detection limit

The EDL for program 432 is 4 mg/L COD. The EDL is the calculated lowest average
concentration in a deionized water matrix that is different from zero with a 99% level of
confidence.
Summary of method
Chemical Oxygen Demand (COD) is defined as a measure of the oxygen equivalent of
the organic matter content of a sample that is susceptible to oxidation by a strong
chemical oxidant (APHA Standard Methods, 19th ed., 1995). Trivalent manganese is a
strong, non-carcinogenic chemical oxidant that changes quantitatively from purple to
colorless when it reacts with organic matter. It typically oxidizes about 80% of the organic
compounds. Studies have shown that the reactions are highly reproducible and test
results correlate closely to Biochemical Oxygen Demand (BOD) values and hexavalent
chromium COD tests. None of the oxygen demand tests provide 100% oxidation of all
organic compounds.
A calibration is provided which is based on the oxidation of Potassium Acid Phthalate
(KHP). A different response may be seen in analyzing various wastewaters. The KHP
calibration is adequate for most applications. The highest degree of accuracy is obtained
when test results are correlated to a standard reference method such as BOD or one of
the chromium COD methods. Special waste streams or classes will require a separate
calibration to obtain a direct mg/L COD reading or to generate a correction factor for the
precalibrated KHP response. The sample digestion time can be extended up to four hours
for samples that are difficult to oxidize. Test results are measured at 510 nm in
spectrophotometers and 520 nm in colorimeters.
Consumables and replacement items
Required reagents

Description Quantity/test Unit Item no.

Manganese III COD Reagent Vials, 20–1000 mg/L COD 1 25/pkg 2623425
Chloride Removal Cartridge (CRC) 1 25/pkg 2661825

Oxygen Demand, Chemical, Manganese III Method with chloride removal (1000 mg/L) 7
Consumables and replacement items (continued)
Description Quantity/test Unit Item no.

Sulfuric Acid, concentrated, ACS 75 mL 2.5 L 97909

Water, deionized varies 4L 27256

Required apparatus

Description Quantity/test Unit Item no.

Blender, 2-speed, 120 VAC 1 each 2616100

Cap, with inert Teflon liner, for mixing bottle varies 12/pkg 2401812
Cell, mixing 2 each 2427700
DRB200 Reactor, 110 V, 15 x 16 mm 1 each LTV082.53.40001
DRB200 Reactor, 220 V, 15 x 16 mm 1 each LTV082.52.40001
Forceps, extra fine point 1 each 2669600
®
Pipet, TenSette , 1.0- to 10.0-mL 1 each 1970010
Pipet Tips, for TenSette Pipet 1970010 varies 50/pkg 2199796
®
Pipet, TenSette , 0.1- to 1.0-mL 1 each 1970001
Pipet Tips, for TenSette Pipet 1970001 2 50/pkg 2185696
Test tube rack 1 each 1864100
Vacuum Pretreatment Device (VPD) 1 each 4900000
Vacuum Pump, 1.2 CFM 115 V 1 each 2824800

Recommended standards

Description Unit Item no.

COD Standard Solution, 800-mg/L 200 mL 2672629

Oxygen Demand Standard (BOD, COD, TOC), 10-mL ampules 16/pkg 2833510
Potassium Acid Phthalate, ACS 500 g 31534
Wastewater Influent Standard, Mixed Parameter, for NH3-N, NO3-N, PO4, COD, SO4,
500 mL 2833149
TOC

Optional reagents and apparatus

Description Unit Item no.

Dispenser, automatic, 1.0–5.0 mL each 2563137

®
Titrator Strips, Quantab , for low range chloride 40 tests 2744940
Finger cots 2/pkg 1464702
Paper, pH, 0–14 pH range 100/pkg 2601300
COD Standard Solution, 300-mg/L 200 mL 1218629
COD Standard Solution, 300-mg/L 500mL 1218649
COD Standard Solution, 1000-mg/L 200 mL 2253929
Standard Methods Book, most current edition each 2270800
Pipet tips for TenSette Pipet 1970010 250/pkg 2199725
Pipet tips for TenSette Pipet 1970001 1000/pkg 2185628

8 Oxygen Demand, Chemical, Manganese III Method with chloride removal (1000 mg/L)
Oxygen Demand, Chemical, Manganese III Method with chloride removal (1000 mg/L) 9
 FOR TECHNICAL ASSISTANCE, PRICE INFORMATION AND ORDERING: HACH COMPANY
In the U.S.A. – Call toll-free 800-227-4224 WORLD HEADQUARTERS
Outside the U.S.A. – Contact the HACH office or distributor serving you. Telephone: (970) 669-3050
On the Worldwide Web – www.hach.com; E-mail – [email protected] FAX: (970) 669-2932

© Hach Company/Hach Lange GmbH, 1989–2013. All rights reserved. 04/2013, Edition 8
Oxygen, Dissolved DOC316.53.01096

HRDO Method Method 8166

®
0.3 to 15.0 mg/L O2 (HR) AccuVac Ampuls
Scope and application: For water and wastewater

Test preparation

Instrument-specific table
The tables in this section show all of the instruments that have the program for this test.
Table 2 shows sample cell and adapter requirements for AccuVac Ampul tests.
To use the table, select an instrument, then read across to find the corresponding
information for this test.
Table 1 Instrument-specific information for AccuVac Ampuls
Instrument Adapter Sample cell
DR 6000 — 2427606
DR 5000
DR 900
DR 3900 LZV846 (A)
DR 3800 LZV584 (C) 2122800
DR 2800
DR 2700

Before starting
Samples must be analyzed immediately after collection and cannot be preserved for later analysis.
Install the instrument cap on the DR 900 cell holder before ZERO or READ is pushed.
Review the Safety Data Sheets (MSDS/SDS) for the chemicals that are used and use any recommended personal protective
equipment.
Dispose of reacted solutions according to local, state and federal regulations. Use the Safety Data Sheets for disposal
information for unused reagents. Consult the environmental, health and safety staff for your facility and/or local regulatory
agencies for further disposal information.

Items to collect
Description Quantity
®
High Range Dissolved Oxygen AccuVac Ampuls 1
Polypropylene beaker, 50-mL 1
Stoppers, for 18-mm tubes and AccuVac Ampuls 1
Sample cells (For information about sample cells, adapters or light shields, refer to Instrument-
1
specific table on page 1.)

Refer to Consumables and replacement items on page 4 for reorder information.

1
Sample collection
Good sample collection and handling technique are important to get meaningful results.
The dissolved oxygen content of the water that is tested can change with depth,
turbulence, temperature, sludge deposits, light, microbial action, mixing, travel time and
other factors. A single dissolved oxygen test rarely reflects the accurate overall condition
of a body of water. Several samples taken at different times, locations and depths are
recommended for most reliable results.
The main consideration with sample collection is to prevent contamination of the sample
with atmospheric oxygen.
• Samples must be analyzed immediately after collection, although only a small error
results if the reading on a capped ampule is taken several hours later. The
absorbance will decrease by approximately 3% during the first hour and will not
change significantly afterwards.
• Make sure to put the cap on the ampule before the ampule is removed from the
sample.
®
AccuVac Ampul procedure

Start

1. Start program 2. Prepare the blank: Fill 3. Fill a blue Ampul cap 4. Prepare the sample:
445 Oxygen, Dis HR AV. the sample cell with 10 mL with sample. Collect at least 40 mL of
For information about of sample. sample in a 50-mL beaker.
sample cells, adapters or Fill the AccuVac Ampul with
light shields, refer to sample. Keep the tip
Instrument-specific table immersed while the Ampul
on page 1. fills completely.
Note: Although the program
name may vary between
instruments, the program
number does not change.

5. Hold the Ampul with the 6. Shake the Ampul for 7. Start the instrument 8. When the timer expires,
tip down. Immediately put 30 seconds. timer. A 2-minute reaction shake the Ampul for
the Ampul into the Ampul A small amount of time starts. 30 seconds.
cap. undissolved reagent will not The oxygen that has Let all of the bubbles
The cap prevents affect results. degassed during aspiration dissipate before the next
contamination from dissolves again and reacts. step.
atmospheric oxygen.

2 Oxygen, Dissolved, HRDO Method (15.0 mg/L)

 Zero

9. Clean the blank. 10. Insert the blank into the 11. Push ZERO. The 12. Clean the AccuVac
cell holder. display shows 0.0 mg/L O2. Ampul.

Read

13. Insert the prepared 14. Push READ. Results

sample AccuVac Ampul into show in mg/L O2.
the cell holder.

Interferences
Interfering substance Interference level
Cr3+ More than 10 mg/L
Cu2+ More than 10 mg/L
Fe2+ More than 10 mg/L
Mg2+ Magnesium is commonly present in seawater and causes a negative interference. If the sample
contains more than 50% seawater, the oxygen concentration obtained by this method will be 25%
less than the true oxygen concentration. If the sample contains less than 50% seawater, the
interference will be less than 5%.
Mn2+ More than 10 mg/L
Ni2+ More than 10 mg/L
NO2- More than 10 mg/L

Accuracy check
Comparison method
To validate the test results, measure the concentration of the same sample with a
dissolved oxygen meter or with a titrimetric method.
Method performance
The method performance data that follows was derived from laboratory tests that were
measured on a spectrophotometer during ideal test conditions. Users may get different
results under different test conditions.
Program Standard Precision (95% Confidence Interval) Sensitivity
Concentration change per 0.010 Abs change
445 6.7 mg/L O2 6.2–7.3 mg/L O2 0.09 mg/L O2

Oxygen, Dissolved, HRDO Method (15.0 mg/L) 3

Summary of method
The High Range Dissolved Oxygen AccuVac Ampul contains reagent vacuum-sealed in a
glass Ampul. When the AccuVac Ampul is opened in a sample that contains dissolved
oxygen, the solution forms a yellow color which turns purple. The purple color
development is proportional to the concentration of dissolved oxygen. The measurement
wavelength is 535 nm for spectrophotometers or 520 nm for colorimeters.
Consumables and replacement items
Required reagents

Description Quantity/test Unit Item no.

High Range Dissolved Oxygen AccuVac® Ampuls 1 25/pkg 2515025

Required apparatus

Description Quantity/test Unit Item no.

Beaker, polypropylene, 50-mL, low form 1 each 108041

Optional reagents, apparatus and meters

Description Unit Item no.

®
AccuVac Snapper each 2405200
®
AccuVac Sampler each 2405100
®
AccuVac vials for sample blanks 25/pkg 2677925
Stoppers for 18-mm tubes and AccuVac Ampuls 6/pkg 173106
HQ30d Meter with Standard LDO Dissolved Oxygen Probe (1 meter cable; additional
each HQ30d53301000
probes and cable lengths are available)
HQ40d Meter with Standard LDO Probe (1 meter cable; additional probes and cable
each HQ40d53301000
lengths are available)

FOR TECHNICAL ASSISTANCE, PRICE INFORMATION AND ORDERING: HACH COMPANY

In the U.S.A. – Call toll-free 800-227-4224 WORLD HEADQUARTERS
Outside the U.S.A. – Contact the HACH office or distributor serving you. Telephone: (970) 669-3050
On the Worldwide Web – www.hach.com; E-mail – [email protected] FAX: (970) 669-2932

© Hach Company/Hach Lange GmbH, 1989–2013. All rights reserved. 04/2013, Edition 8
Oxygen, Dissolved DOC316.53.01098

Indigo Carmine Method Method 8316

®
6 to 800 µg/L O2 (LR, spectrophotometers) AccuVac Ampuls
10 to 1000 µg/L O2 (LR, colorimeters)
Scope and application: For boiler feedwater

Test preparation

Instrument-specific table
The tables in this section show all of the instruments that have the program for this test.
Table 2 shows sample cell and adapter requirements for AccuVac Ampul tests.
To use the table, select an instrument, then read across to find the corresponding
information for this test.
Table 1 Instrument-specific information for AccuVac Ampuls
Instrument Adapter Sample cell
DR 6000 — 2427606
DR 5000
DR 900
DR 3900 LZV846 (A)
DR 3800 LZV584 (C) 2122800
DR 2800
DR 2700

Before starting
Samples must be analyzed immediately after collection and cannot be preserved for later analysis.
Install the instrument cap on the DR 900 cell holder before ZERO or READ is pushed.
The dissolved oxygen reading is only stable for 30 seconds. After 30 seconds, the Ampul solution will absorb oxygen from
the air.
Review the Safety Data Sheets (MSDS/SDS) for the chemicals that are used and use any recommended personal protective
equipment.
Dispose of reacted solutions according to local, state and federal regulations. Use the Safety Data Sheets for disposal
information for unused reagents. Consult the environmental, health and safety staff for your facility and/or local regulatory
agencies for further disposal information.

Items to collect
Description Quantity
®
Low Range Dissolved Oxygen AccuVac Ampuls 1
Polypropylene beaker, 50-mL 1
Stoppers, for 18-mm tubes and AccuVac Ampuls 1
Sample cells (For information about sample cells, adapters or light shields, refer to Instrument-
1
specific table on page 1.)

Refer to Consumables and replacement items on page 3 for reorder information.

1
Sample collection
The main consideration with sample collection is to prevent contamination of the sample
with atmospheric oxygen.
• Samples must be analyzed immediately after collection and cannot be preserved for
later analysis.
• For best results, collect the sample from a stream of water that is hard-plumbed to the
sample source.
• Use a funnel to maintain a continual flow of sample and yet collect enough sample to
immerse the Ampul.
• Do not introduce air in place of the sample.
• Rubber tubing, if used, will introduce unacceptable amounts of oxygen into the
sample unless the length of tubing is minimized and the flow rate is maximized.
• Flush the sampling system with sample for at least 5 minutes.
®
AccuVac Ampul procedure

Start

1. Start program 2. Prepare the blank: Fill 3. Clean the blank. 4. Insert the blank into the
446 Oxygen, Dis LR AV. the sample cell with 10 mL cell holder.
For information about of sample.
sample cells, adapters or
light shields, refer to
Instrument-specific table
on page 1.
Note: Although the program
name may vary between
instruments, the program
number does not change.

Zero Read

5. Push ZERO. The display 6. Prepare the sample: 7. Immediately clean and 8. Push READ. Results
shows 0 µg/L O2. Open the AccuVac Ampul. then insert the Ampul into show in µg/L O2.
Fill the Ampul with sample. the cell holder.
For best results, collect the
sample from a stream of
water that is hard-plumbed
to the sample source. Refer
to Sample collection
on page 2.

2 Oxygen, Dissolved, Indigo Carmine Method (800 µg/L)

Interferences
Excess amounts of thioglycolate, ascorbate, ascorbate + sulfite, ascorbate + cupric
sulfate, nitrite, sulfite, thiosulfate and hydroquinone will not reduce the oxidized form of
the indicator and do not cause significant interference.
Interfering substance Interference level
Hydrazine 100,000 fold excess will begin to reduce the oxidized form of the indicator solution.
Sodium hydrosulfite Reduces the oxidized form of the indicator solution and will cause a significant interference.

Accuracy check
Reagent blank measurement
A reagent blank for this test can be measured as follows:

1. Fill a 50-mL beaker with sample.

2. Add one sodium hydrosulfite powder pillow and mix.
3. Fill a Low Range Dissolved Oxygen AccuVac Ampul with this sample.
4. Measure the dissolved oxygen concentration as shown in the test procedure. The
result should be 0 ± 6 µg/L O2.

Method performance
The method performance data that follows was derived from laboratory tests that were
measured on a spectrophotometer during ideal test conditions. Users may get different
results under different test conditions.
Program Standard Precision (95% Confidence Interval) Sensitivity
Concentration change per 0.010 Abs change
446 N/A not determined 6 µg/L O2

Summary of method
The Low Range Dissolved Oxygen AccuVac Ampul contains reagent vacuum-sealed in
an Ampul. When the AccuVac Ampul is broken open in a sample containing dissolved
oxygen, the yellow solution will turn blue. The blue color development is proportional to
the concentration of dissolved oxygen. Test results are measured at 610 nm.
Consumables and replacement items
Required reagents

Description Quantity/test Unit Item no.

®
Low Range Dissolved Oxygen AccuVac Ampuls 1 25/pkg 2501025

Required apparatus

Description Quantity/test Unit Item no.

Beaker, polypropylene, 50-mL, low form 1 each 108041

Recommended standards

Description Unit Item no.

Hydrosulfite Reagent Powder Pillows 100/pkg 2118869

Oxygen, Dissolved, Indigo Carmine Method (800 µg/L) 3

Optional reagents and apparatus

Description Unit Item no.

®
AccuVac Snapper each 2405200
®
AccuVac Sampler each 2405100
®
AccuVac vials for sample blanks 25/pkg 2677925
Stoppers for 18-mm tubes and AccuVac Ampuls 6/pkg 173106

FOR TECHNICAL ASSISTANCE, PRICE INFORMATION AND ORDERING: HACH COMPANY

In the U.S.A. – Call toll-free 800-227-4224 WORLD HEADQUARTERS
Outside the U.S.A. – Contact the HACH office or distributor serving you. Telephone: (970) 669-3050
On the Worldwide Web – www.hach.com; E-mail – [email protected] FAX: (970) 669-2932

© Hach Company/Hach Lange GmbH, 1989–2013. All rights reserved. 04/2013, Edition 8
Oxygen Scavengers DOC316.53.01105

Iron Reduction Method Method 8140

3 to 450 µg/L DEHA; 5 to 600 µg/L carbohydrazide; 9 to 1000 µg/L Powder Pillows
hydroquinone; 13 to 1500 µg/L iso-ascorbic acid (ISA); 15 to 1000 µg/L
methylethyl ketoxime (MEKO)
Scope and application: For testing residual corrosion inhibitors (oxygen scavengers) in boiler feed water or
condensate

Test preparation

Instrument specific information

The table in this section shows all of the instruments that have the program for this test.
Table 1 shows sample cell and orientation requirements for reagent addition tests, such
as powder pillow or bulk reagent tests.
To use the table, select an instrument, then read across to find the corresponding
information for this test.
Table 1 Instrument-specific information for powder pillows
Instrument Sample cell orientation Sample cell
DR 6000 The fill line is to the right. 2495402
DR 3800
DR 2800
DR 2700
DR 5000 The fill line is toward the user.
DR 3900
DR 900 The orientation mark is toward the user. 2401906

Before starting
Samples must be analyzed immediately after collection and cannot be preserved for later analysis.
Install the instrument cap on the DR 900 cell holder before ZERO or READ is pushed.
The sample temperature should be 25 ± 3 °C (77 ± 5 °F).
Clean all glassware with 6.0 N (50%) hydrochloric acid, then rinse thoroughly with deionized water to remove iron
contaminants.
To measure the ferrous iron concentration, repeat the test procedure but do not add the DEHA Reagent 2. To automatically
subtract the ferrous iron concentration from the test results, use the reagent blank adjust option. Use the ferrous iron
concentration as the reagent blank value.
Review the Safety Data Sheets (MSDS/SDS) for the chemicals that are used and use any recommended personal protective
equipment.
Dispose of reacted solutions according to local, state and federal regulations. Use the Safety Data Sheets for disposal
information for unused reagents. Consult the environmental, health and safety staff for your facility and/or local regulatory
agencies for further disposal information.

1
Items to collect
Description Quantity

Bottle, glass mixing, with 25-mL mark 2

DEHA Reagent 1 Powder Pillows 2
DEHA Reagent 2 Solution 1 mL
Dropper, 0.5 and 1.0 mL marks 1
Hydrochloric acid, 1:1, 6.0 N varies
Water, deionized 25 mL
Sample cells (For information about sample cells, adapters or light shields, refer to Instrument
2
specific information on page 1.)

Refer to Consumables and replacement items on page 4 for reorder information.

Sample collection
• Samples must be analyzed immediately after collection and cannot be preserved for
later analysis.
• Collect samples in clean, dry glass or plastic bottles with tight-fitting caps.
• Rinse the container several times with the sample before collection.
• Prevent agitation of the sample or exposure to sunlight or air.
• Fill the bottle completely and let the sample overflow. Immediately tighten the cap so
that there is no headspace above the sample.

Powder pillow procedure

Start

1. Start program 180 O 2. Prepare the blank: Fill a 3. Prepare the sample: Fill 4. Add the contents of one
Scav-Carbohy, 181 O mixing bottle with 25 mL of a second mixing bottle with DEHA Reagent 1 Powder
Scav-DEHA, 182 O Scav- deionized water. 25 mL of sample. Pillow to each mixing bottle.
Hydro, 183 O Scav-ISA or To measure oxygen
184 O Scav-MEKO. For scavenders that react
information about sample quickly with oxygen at room
cells, adapters or light temperature, close the
shields, refer to Instrument bottle.
specific information
on page 1.
Note: Although the program
name may vary between
instruments, the program
number does not change.

2 Oxygen Scavengers, Iron Reduction Method (450 µg/L DEHA)

5. Swirl to mix. 6. Use a pipet to add 7. Swirl to mix. 8. Start the instrument
0.5 mL of DEHA Reagent Put both mixing bottles in a timer. A 10-minute (2-minute
2 Solution to each bottle. dark location. A purple color for hydroquinone) reaction
shows if an oxygen time starts.
scavender is present in the Keep the mixing bottles in
sample. the dark during the reaction
period.

Zero

9. When the timer expires, 10. Clean the blank. 11. Insert the blank into the 12. Push ZERO. For
transfer the blank and cell holder. greater accuracy, read the
prepared samples to the result immediately after the
sample cells. timer expires.

Read

13. Clean the prepared 14. Insert the prepared 15. Push READ. Results
sample. sample into the cell holder. show in µg/L.

Interferences
Substances which reduce ferric iron will interfere. Substances which complex iron
strongly may also interfere.
Interfering substance Interference level
Borate (as Na2B4O7) More than 500 mg/L
Cobalt More than 0.025 mg/L
Copper More than 8.0 mg/L
Ferrous Iron All levels. Measure and subtract (refer to Before starting on page 1)

Oxygen Scavengers, Iron Reduction Method (450 µg/L DEHA) 3

Interfering substance Interference level
Hardness (as CaCO3) More than 1000 mg/L
Light Light may interfere. Keep sample cells in the dark during color development.
Lignosulfonates More than 0.05 mg/L
Manganese More than 0.8 mg/L
Molybdenum More than 80 mg/L
Nickel More than 0.8 mg/L
Phosphate More than 10 mg/L
Phosphonates More than 10 mg/L
Sulfate More than 1000 mg/L
Temperature Sample temperatures below 22 °C or above 28 °C (72 °F or 82 °F) may affect test accuracy.
Zinc More than 50 mg/L

Method performance
The method performance data that follows was derived from laboratory tests that were
measured on a spectrophotometer during ideal test conditions. Users may get different
results under different test conditions.
Program Standard Precision (95% Confidence Interval) Sensitivity
Concentration change per 0.010 Abs change
180 299 µg/L 295–303 µg/L 4 µg/L
181 226 µg/L 223–229 µg/L 3 µg/L
182 600 µg/L 591–609 µg/L 8 µg/L
183 886 µg/L 873–899 µg/L 12 µg/L
184 976 µg/L 962–990 µg/L 14 µg/L

Summary of method
Diethylhydroxylamine (DEHA) or other oxygen scavengers in the sample react with ferric
iron in DEHA Reagent 2 Solution to produce ferrous ion in an amount that is equivalent to
the DEHA concentration. This solution then reacts with DEHA 1 Reagent, which forms a
purple color with ferrous iron that is proportional to the concentration of the oxygen
scavenger. This method reacts with all oxygen scavengers and does not differentiate
when the sample contains more than one type of oxygen scavenger. The measurement
wavelength is 562 nm for spectrophotometers or 560 nm for colorimeters.
Consumables and replacement items
Required reagents

Description Quantity/test Unit Item no.

Hydrochloric Acid, 6.0 N varies 500 mL 88449

Water, deionized varies 4L 27256
Oxygen Scavenger Reagent Set — — 2446600
Includes:
DEHA Reagent 1 Powder Pillows 2 100/pkg 2167969
DEHA Reagent 2 Solution 1 mL 100 mL 2168042

4 Oxygen Scavengers, Iron Reduction Method (450 µg/L DEHA)

Required apparatus

Description Quantity/test Unit Item no.

Bottle, square, with 25 mL mark 1 each 1704200

Dropper, measuring, 0.5 and 1.0 mL plastic 2 20/pkg 2124720
Sample cell, 10 mL square, matched pair 2 2/pkg 2495402

Optional apparatus

Description Unit Item no.

Thermometer, non-mercury, -10 to +225 °C each 2635700

Oxygen Scavengers, Iron Reduction Method (450 µg/L DEHA) 5

 FOR TECHNICAL ASSISTANCE, PRICE INFORMATION AND ORDERING: HACH COMPANY
In the U.S.A. – Call toll-free 800-227-4224 WORLD HEADQUARTERS
Outside the U.S.A. – Contact the HACH office or distributor serving you. Telephone: (970) 669-3050
On the Worldwide Web – www.hach.com; E-mail – [email protected] FAX: (970) 669-2932

© Hach Company/Hach Lange GmbH, 1989–2013. All rights reserved. 04/2013, Edition 8
Ozone DOC316.53.01106

Indigo Method Method 8311

®
0.01 to 0.25 mg/L O3 (LR), 0.01 to 0.75 mg/L O3 (MR), AccuVac Ampuls
0.01 to 1.50 mg/L O3 (HR)
Scope and application: For water.

Test preparation

Instrument-specific information
The table in this section shows all of the instruments that have the program for this test.
Table 1 shows the adapter requirement for AccuVac Ampul tests.
To use the table, select an instrument, then read across to find the corresponding
information for this test.
Table 1 Instrument-specific information for AccuVac Ampuls
Instrument Adapter
DR 6000 —
DR 5000
DR 900
DR 3900 LZV846 (A)
DR 3800 LZV584 (C)
DR 2800
DR 2700

Before starting
Samples must be analyzed immediately after collection and cannot be preserved for later analysis.
Install the instrument cap on the DR 900 cell holder before ZERO or READ is pushed.
Use tap water or deionized water for the blank (ozone-free water).
In this method, the instrument is intentionally zeroed on the sample, not the blank.
Review the Safety Data Sheets (MSDS/SDS) for the chemicals that are used and use any recommended personal protective
equipment.
Dispose of reacted solutions according to local, state and federal regulations. Use the Safety Data Sheets for disposal
information for unused reagents. Consult the environmental, health and safety staff for your facility and/or local regulatory
agencies for further disposal information.

Items to collect
AccuVac Ampuls

Description Quantity
®
Ozone AccuVac Ampules, 0-0.25 mg/L 2
®
Ozone AccuVac Ampules, 0-0.75 mg/L 2
®
Ozone AccuVac Ampules, 0-1.5 mg/L 2
Beaker, 50 mL 1
Stoppers, for 18-mm tubes and AccuVac Ampuls 2
Water, ozone-free varies

1
Refer to Consumables and replacement items on page 3 for reorder information.

Sample collection
• Samples must be analyzed immediately after collection and cannot be preserved for
later analysis.
• The most important consideration during sample collection is to prevent the escape of
ozone from the sample.
• Collect the sample gently and analyze immediately. Do not shake or stir the sample
or allow the sample temperature to increase.
• Do not transfer the sample from one container to another unless absolutely
necessary.

AccuVac Ampul procedure

Note: For this procedure, the zero step is done on the prepared sample, and the read step on the
blank.

Start

1. Start program 2. Prepare the blank: Pour 3. Prepare the sample: 4. Quickly invert the Ampuls
454 Ozone LR AV, at least 40 mL of ozone-free Pour at least 40 mL of several times to mix.
455 Ozone MR AV or water in a 50-mL beaker. Fill sample in a 50-mL beaker. Some of the blue color will
456 Ozone HR AV. For an Indigo Ozone Reagent Fill an Indigo Ozone be bleached if ozone is
information about sample AccuVac Ampul with the Reagent AccuVac Ampul present.
cells, adapters or light ozone-free water. Keep the with the sample. Keep the
shields, refer to Instrument- tip immersed while the tip immersed while the
specific information Ampul fills fully. Ampul fills fully.
on page 1.
Note: Although the program
name may vary between
instruments, the program
number does not change.

Zero

5. Clean the prepared 6. Insert the prepared 7. Push ZERO. The display 8. Clean the blank AccuVac
sample AccuVac Ampul. sample AccuVac Ampul into shows 0.00 mg/L O3. Ampul.
the cell holder.

2 Ozone, Indigo Method (multi-range: 0.25, 0.75, 1.50 mg/L)

 Read

9. Insert the blank AccuVac 10. Push READ. Results

Ampul into the cell holder. show in mg/L O3.

Reagent stability
The indigo reagent is light-sensitive. Keep the unused AccuVac Ampuls in the dark. The
indigo solution decomposes slowly under room light after the AccuVac Ampul is filled.
The filled blank Ampul can be used for multiple measurements during the same day.
Method performance
The method performance data that follows was derived from laboratory tests that were
measured on a spectrophotometer during ideal test conditions. Users may get different
results under different test conditions.
Program Standard Precision (95% Confidence Interval) Sensitivity
Concentration change per 0.010 Abs change
454 0.15 mg/L O3 0.14–0.16 mg/L O3 0.01 mg/L O3
455 0.45 mg/L O3 0.43–0.47 mg/L O3 0.01 mg/L O3
456 1.00 mg/L O3 0.97–1.03 mg/L O3 0.01 mg/L O3

Summary of method
The reagent formulation adjusts the sample pH to 2.5 after the Ampule has filled. The
indigo reagent reacts immediately and quantitatively with ozone. The blue color of indigo
is bleached in proportion to the amount of ozone present in the sample. Other reagents in
the formulation prevent chlorine interference. No transfer of sample is needed in the
procedure, therefore ozone loss due to sampling is eliminated. The measurement
wavelength is 600 nm for spectrophotometers or 610 nm for colorimeters.
Consumables and replacement items
Required reagents

Description Quantity/test Unit Item no.

®
Ozone AccuVac Ampules, 0-0.25 mg/L 2 25/pkg 2516025
®
Ozone AccuVac Ampules, 0-0.75 mg/L 2 25/pkg 2517025
®
Ozone AccuVac Ampules, 0-1.5 mg/L 2 25/pkg 2518025

Required apparatus

Description Quantity/test Unit Item no.

AccuVac Snapper 1 each 2405200

Beaker, 50-mL 1 each 50041H
Stoppers for 18-mm tubes and AccuVac Ampuls 2 6/pkg 173106
Beaker, polypropylene, 50-mL, low form 1 each 108041

Ozone, Indigo Method (multi-range: 0.25, 0.75, 1.50 mg/L) 3

Optional reagents and apparatus

Description Unit Item no.

Water, deionized 4L 27256

SpecCheck™ Gel Secondary Standard Kit, Ozone, 0–0.75 mg/L set each 2708000

FOR TECHNICAL ASSISTANCE, PRICE INFORMATION AND ORDERING: HACH COMPANY

In the U.S.A. – Call toll-free 800-227-4224 WORLD HEADQUARTERS
Outside the U.S.A. – Contact the HACH office or distributor serving you. Telephone: (970) 669-3050
On the Worldwide Web – www.hach.com; E-mail – [email protected] FAX: (970) 669-2932

© Hach Company/Hach Lange GmbH, 1989–2013. All rights reserved. 04/2013, Edition 8
Phosphonates DOC316.53.01109

Persulfate UV Oxidation Method1 Method 8007

Multiple ranges from 0.02 to 125 mg/L PO43– Powder Pillows
Scope and application: For boiler and cooling water, wastewater and seawater.
1 Adapted from Blystone, P., Larson, P., A Rapid Method for Analysis of Phosphate Compounds, International Water Conference,
Pittsburgh, PA. (Oct 26-28, 1981)

Test preparation

Instrument specific information

The table in this section shows all of the instruments that have the program for this test.
Table 1 shows sample cell and orientation requirements for reagent addition tests, such
as powder pillow or bulk reagent tests.
To use the table, select an instrument, then read across to find the corresponding
information for this test.
Table 1 Instrument-specific information for reagent addition
Instrument Sample cell orientation Sample cell
DR 6000 The fill line is to the right. 2495402
DR 3800
DR 2800
DR 2700
DR 5000 The fill line is toward the user.
DR 3900
DR 900 The orientation mark is toward the user. 2401906

Before starting
Install the instrument cap on the DR 900 cell holder before ZERO or READ is pushed.
Clean all glassware with 6.0 N (50%) hydrochloric acid, then rinse thoroughly with deionized water to remove contaminants.
Do not use a detergent that contains phosphate to clean glassware. The phosphate in the detergent will contaminate the
sample.
Wear UV safety goggles while the UV lamp is on.
Do not touch the UV lamp surface with bare fingers. Fingerprints can damage the glass. Rinse the lamp and wipe with a soft,
clean tissue between tests.
The UV digestion in this procedure is normally complete in less than 10 minutes. However, high-organic loaded samples or a
weak lamp can cause incomplete phosphate conversion. To check conversion efficiency, use a longer digestion time and
make sure the readings do not increase.
Review the Safety Data Sheets (MSDS/SDS) for the chemicals that are used and use any recommended personal protective
equipment.
Dispose of reacted solutions according to local, state and federal regulations. Use the Safety Data Sheets for disposal
information for unused reagents. Consult the environmental, health and safety staff for your facility and/or local regulatory
agencies for further disposal information.

1
Items to collect
Description Quantity

Bottle, square, with 25-mL mark 1

Cylinder, graduated mixing, 50-mL 1
Goggles, UV safety 1
Pipet, serological, 10-mL 1
®
PhosVer 3 Phosphate Reagent Powder Pillows, 10-mL 2
Potassium Persulfate Powder Pillow for Phosphonate 1
Pipet filler, safety bulb 1
Sample cells (For information about sample cells, adapters or light shields, refer to Instrument
2
specific information on page 1.)
Water, deionized varies
UV lamp with power supply 1

Refer to Consumables and replacement items on page 7 for reorder information.

Sample collection and storage

• Collect samples in clean glass or plastic bottles that have been cleaned with
1:1 hydrochloric acid and rinsed with deionized water.
• Do not use a commerical detergent to clean the sample bottles. The phosphate in the
detergent will contaminate the sample.
• To preserve samples for later analysis, adjust the sample pH to less than 2 with
concentrated sulfuric acid (about 2 mL per liter). No acid addition is necessary if the
sample is tested immediately.
• Keep the preserved samples at or below 6 °C (43 °F) for up to 24 hours.
• Let the sample temperature increase to room temperature before analysis.
• Before analysis, adjust the pH to 7 with 5.0 N sodium hydroxide standard solution.
• Correct the test result for the dilution from the volume additions.

Persulfate UV oxidation for powder pillows

Start

1. Start program 2. Select the sample size 3. Prepare the blank: Fill a 4. Prepare the digested
501 Phosphonates. For from Select the sample sample cell to the 10-mL sample: Fill a mixing bottle
information about sample volume and multiplier mark with the diluted sample to the 25-mL mark with the
cells, adapters or light on page 4. Use a pipet to from step 2. diluted sample from step 2.
shields, refer to Instrument add the correct volume of
specific information sample into a 50-mL
on page 1. graduated cylinder.
Note: Although the program If necessary, dilute the
name may vary between sample to 50-mL with
instruments, the program deionized water and mix
number does not change. well.

2 Phosphonates, Persulfate UV Oxidation Method (125.0 mg/L)

5. Add the contents of one 6. Swirl to mix. 7. Put on the UV safety 8. Put the ultraviolet lamp
Potassium Persulfate for goggles. into the mixing bottle. Turn
Phosphonate Powder Pillow on the UV lamp.
to the 25-mL sample.

9. Start the instrument 10. When the timer expires, 11. Prepare the sample: 12. Add the contents of one
timer. A 10-minute reaction turn off the UV lamp. Fill a second sample cell to PhosVer 3 Phosphate
time starts. Remove the UV lamp from the 10-mL mark with the Reagent Powder Pillow to
Phosphonates are the sample. digested sample. both the blank and the
converted to prepared sample.
orthophosphate in this step.

13. Immediately swirl both 14. Start the instrument 15. When the timer expires, 16. Insert the blank into the
cells vigorously for 20– timer. A 2-minute reaction clean the blank. cell holder.
30 seconds to mix. Some time starts. Complete the rest of the
powder may not dissolve. If the sample is colder than steps in this procedure
A blue color shows if 15 °C, wait» four minutes for within three minutes.
phosphate is present. Both color development.
the sample and the blank
may show color.

Phosphonates, Persulfate UV Oxidation Method (125.0 mg/L) 3

 Zero Read

17. Push ZERO. The 18. Clean the prepared 19. Insert the prepared 20. Push READ. Results
display shows 0.00 mg/L sample. sample into the cell holder. show in mg/L PO43– .
PO43–.

21. Multiply the results with

the appropriate multiplier for
the phosphonate
concentration. Refer to
Convert phosphate to
phosphonate on page 4.

Select the sample volume and multiplier

Use the expected phosphonate concentration to select a sample volume (refer to
Table 2). Use the multiplier to adjust the test result (in mg/L PO4) for the sample volume
that was used.
Table 2 Expected phosphonate range with multiplier
Expected range (mg/L phosphonate) Sample volume (mL) Multiplier
0–2.5 50 0.1
0–5 25 0.2
0–12.5 10 0.5
0–25 5 1
0–125 1 5

Convert phosphate to phosphonate

To convert the final test result from mg/L PO43– to active phosphonate, multiply the final
test result by the applicable conversion factor in Table 3.
Table 3 Conversion factors by phosphonate type
Phosphonate type Conversion factor
PBTC 2.84
NTP 1.05
HEDPA 1.085

4 Phosphonates, Persulfate UV Oxidation Method (125.0 mg/L)

 Table 3 Conversion factors by phosphonate type (continued)
Phosphonate type Conversion factor
EDTMPA 1.148
HMDTMPA 1.295
DETPMPA 1.207
HPA 1.49

Interferences
Interference levels decrease as the sample size increases. For example, copper does not
interfere at or below 100 mg/L for a 5.00 mL sample. If the sample volume is increased to
10 mL, copper will begin to interfere above 50 mg/L.
Interfering substance Interference level (5 mL sample)
Aluminum 100 mg/L
Arsenate Interferes at all levels
Benzotriazole 10 mg/L
Bicarbonate 1000 mg/L
Bromide 100 mg/L
Calcium 5000 mg/L
CDTA 100 mg/L
Chloride 5000 mg/L
Chromate 100 mg/L
Copper 100 mg/L
Cyanide 100 mg/L (Increase the UV digestion to 30 minutes.)
Diethanoldithiocarbamate 50 mg/L
EDTA 100 mg/L
Iron 200 mg/L
Nitrate 200 mg/L
NTA 250 mg/L
Orthophosphate 15 mg/L
Phosphites and organophosphorus compounds Reacts quantitatively. Meta- and polyphosphates do not interfere.
Silica 500 mg/L
Silicate 100 mg/L
Sulfate 2000 mg/L
Sulfide Interferes at all levels
Sulfite 100 mg/L
Thiourea 10 mg/L
Highly buffered samples or extreme sample pH Can prevent the correct pH adjustment of the sample by the reagents.
Sample pretreatment may be necessary.

Phosphonates, Persulfate UV Oxidation Method (125.0 mg/L) 5

Accuracy check
Digestion method
To validate the full procedure with the digestion, prepare a solution that has a known
concentration of a phosphonate compound. Use the test procedure to measure the
concentration of the phosphonate solution.

Standard solution method

To validate the colorimetric portion of the procedure (without digestion), use a phosphate
standard solution for the sample and deionized water for the blank. Add the PhosVer
3 reagent directly to 10 mL of the phosphate standard solution and to the blank. The
expected result is 10 times the value of the standard solution due to a built-in dilution
factor of 10 in the calibration.
Items to collect:
• Phosphate Standard Solution, 1 mg/L (the expected result is 10 mg/L if 10 mL is
used)

1. Use the test procedure to measure the concentration of the standard solution.
2. Compare the expected result to the actual result.
Note: The factory calibration can be adjusted slightly with the standard adjust option so that the
instrument shows the expected value of the standard solution. The adjusted calibration is then
used for all test results. This adjustment can increase the test accuracy when there are slight
variations in the reagents or instruments.

Method performance
The method performance data that follows was derived from laboratory tests that were
measured on a spectrophotometer during ideal test conditions. Users may get different
results under different test conditions.
Program Standard Precision (95% Confidence Interval) Sensitivity
Concentration change per 0.010 Abs change
501 2.00 mg/L PO43– 1.97–2.03 mg/L PO43– Refer to Sensitivity on page 6.

Sensitivity
The sensitivity depends on the sample volume. Sensitivity is expressed as PO43– in
Table 4. To express as a specific phosphonate, refer to Table 3 on page 4.
Table 4 Sensitivity per sample volume
Range (mg/L phosphonate) Sample volume (mL) Concentration change per 0.010 Abs change
0–2.5 50 0.02 mg/L PO43–
0–5 25 0.04 mg/L PO43–
0–12.5 10 0.10 mg/L PO43–
0–25 5 0.20 mg/L PO43–
0–125 1 1.00 mg/L PO43–

Summary of method
This method is directly applicable to boiler and cooling tower samples. The procedure is
based on a UV-catalyzed oxidation of phosphonate to orthophosphate. The
orthophosphate reacts with the molybdate in the PhosVer 3 reagent to form a mixed
phosphate/molybdate complex. This complex is reduced by the ascorbic acid in the
PhosVer 3, which gives a blue color that is proportional to the amount of phosphonate in
the original sample. The orthophosphate in the original sample is removed when the
blank is used to set the zero concentration. The measurement wavelength is 880 nm for
spectrophotometers or 610 nm for colorimeters.

6 Phosphonates, Persulfate UV Oxidation Method (125.0 mg/L)

Consumables and replacement items
Required reagents

Description Quantity/test Unit Item no.

Water, deionized varies 4L 27256

Phosphonate Reagent Set, 10-mL 1 100 tests 2429700
Includes:
®
PhosVer 3 Phosphate Reagent Powder Pillow, 10-mL 1 100/pkg 2106069
Potassium Persulfate Powder Pillow for Phosphonate 1 100/pkg 2084769

Required apparatus

Description Quantity/test Unit Item no.

Bottle, square, with 25 mL mark 1 each 1704200

Beaker, polypropylene, 50-mL, low form 1 each 108041
Cylinder, graduated mixing, 50-mL, with glass stopper 1 each 189641
UV safety goggles 1 each 2113400
Pipet, serological, graduated, 10-mL 1 each 53238
Pipet filler, safety bulb 1 each 1465100
UV lamp with power supply, 115 VAC 1 each 2082800
OR
UV lamp with power supply, 230 VAC 1 each 2082802

Recommended standards

Description Unit Item no.

Phosphate Standard Solution, 1-mg/L as PO4 500 mL 256949

Optional reagents and apparatus

Description Unit Item no.

Hydrochloric Acid, 1:1, 6N 500 mL 88449

Sulfuric Acid, concentrated, ACS 500 mL 97949
Thermometer, non-mercury, -10 to +225 °C each 2635700
Paper, pH, 0–14 pH range 100/pkg 2601300
®
Ampule Breaker, Voluette ampules each 2196800
PhosVer 3 Phosphate Reagent Powder Pillows, 10 mL 1000/pkg 2106028
UV Lamp, shortwave, pencil type each 2671000
Power supply, 115 V/60 Hz each 2670700
Power supply, 220 V/50 Hz each 2670702
Phosphate Standard Solution, 3-mg/L as PO4 946 mL 2059716
Phosphate Standard Solution, 10-mg/L 946 mL 1420416
Phosphate Standard Solution, 15-mg/L 100 mL 1424342
Phosphate Standard Solution, 30-mg/L 946 mL 1436716

Phosphonates, Persulfate UV Oxidation Method (125.0 mg/L) 7

Consumables and replacement items (continued)
Description Unit Item no.
®
Phosphate Standard Solution, 50-mg/L, 10-mL Voluette Ampules 16/pkg 17110
Phosphate Standard Solution, 100-mg/L 100 mL 1436832
Phosphate Standard Solution, 10-mL Ampule, 500 mg/L 16/pkg 1424210
Phosphate Standard Solution, 500-mg/L 100 mL 1424232

FOR TECHNICAL ASSISTANCE, PRICE INFORMATION AND ORDERING: HACH COMPANY

In the U.S.A. – Call toll-free 800-227-4224 WORLD HEADQUARTERS
Outside the U.S.A. – Contact the HACH office or distributor serving you. Telephone: (970) 669-3050
On the Worldwide Web – www.hach.com; E-mail – [email protected] FAX: (970) 669-2932

© Hach Company/Hach Lange GmbH, 1989–2013. All rights reserved. 04/2013, Edition 8
Phosphorus, Acid Hydrolyzable DOC316.53.01110

USEPA1 Acid Digestion Method2 Method 8180

Scope and application: For water, wastewater, and seawater.

1 USEPA Accepted for wastewater analyses.
2 Adapted from Standard Methods for the Examination of Water and Wastewater 4500-P B & E.

Test preparation

Before starting
Clean all glassware with 6.0 N (50%) hydrochloric acid, then rinse thoroughly with deionized water to remove contaminants.
The results of a reactive phosphorus test after acid-hydrolyzable digestion includes the combination of orthophosphate and
the acid-hydrolyzable (condensed) phosphate. To find the condensed phosphate concentration, subtract the result of an
orthophosphate test without digestion from the result with digestion. Make sure that both results are in the same units, either
mg/L PO43– or mg/L P. To find the organic phosphorus concentration, subtract the result of a acid hydrolyzable test from the
result of a total phosphorus test.
Review the Safety Data Sheets (MSDS/SDS) for the chemicals that are used and use any recommended personal protective
equipment.
Dispose of reacted solutions according to local, state and federal regulations. Use the Safety Data Sheets for disposal
information for unused reagents. Consult the environmental, health and safety staff for your facility and/or local regulatory
agencies for further disposal information.

Items to collect
Description Quantity

Sodium Hydroxide Solution, 5.0 N 2 mL

Sulfuric Acid Solution, 5.25 N 2 mL
Water, deionized varies
Cylinder, graduated, 25-mL 1
Flask, Erlenmeyer, 125-mL 1
Hot plate 1

Refer to Consumables and replacement items on page 3 for reorder information.

Sample collection and storage

• Collect samples in clean glass or plastic bottles that have been cleaned with
1:1 hydrochloric acid and rinsed with deionized water.
• Do not use a detergent that contains phosphate to clean the sample bottles. The
phosphate in the detergent will contaminate the sample.
• Analyze the samples as soon as possible for best results.
• If prompt analysis is not possible, immediately filter and keep the samples at or below
6 °C (43 °F) for a maximum of 48 hours.
• Let the sample temperature increase to room temperature before analysis.

1
Acid digestion

1. Use a graduated cylinder 2. Add 2.0 mL of 5.25 N 3. Boil the sample gently for 4. Let the sample cool to
to add 25-mL of sample into Sulfuric Acid Solution to the 30 minutes. Do not let the room temperature. Add
the 125-mL Elenmeyer flask. flask boil dry. 2.0 mL of 5.0 N Sodium
flask. For the best recovery, Hydroxide Solution to the
concentrate the sample to flask.
less than 20 mL. After
concentration, maintain the
volume of sample near
20 mL by adding small
amounts of deionized water.
Do not exceed 20 mL.

Start

5. Swirl to mix. 6. Pour the sample into a 7. Proceed with a reactive

25-mL graduated cylinder. phosphorous test of the
Adjust the volume to 25 mL expected acid hydrolyzable
by rinsing the flask with phosphorous concentration
deionized water and pouring range.
the rinse water into the
cylinder. • 480 P React. Mo
• 482 P React. Mo AV
• 485 P React. Amino
• 490 P React. PV
• 492 P React. PV AV
• 535 P React. PV TNT
• 540 P React. HT TNT

Extend the color

development time to
10 minutes for the PhosVer
3 method.

2 Phosphorus, Acid Hydrolyzable, Acid Digestion

Interferences
Interfering substance Interference level
Alkaline or highly If the pH of the sample after the acid is added is not below pH 1, add additional acid.
buffered samples
Turbidity Use 50 mL of sample and double the reagent quantities. Use a portion of the digested sample to
zero the instrument in the reactive phosphorus procedure. This compensates for any color or
turbidity destroyed by this procedure.

Summary of method
Phosphates present in condensed inorganic forms (meta-, pyro- or other polyphosphates)
must be converted to reactive orthophosphate before analysis. Pretreatment of the
sample with acid and heat hydrolyzes the condensed inorganic forms to orthophosphate.
This procedure must be followed by one of the reactive phosphorus (orthophosphate)
analysis methods to determine the phosphorus content of the sample. If the ascorbic acid
(PhosVer 3) method is used to measure the reactive phosphorus, this method is USEPA
accepted for NPDES reporting.
Consumables and replacement items
Required reagents

Description Quantity/test Unit Item no.

100 mL
Sodium Hydroxide Solution, 5.0 N 2 mL 245032
MDB
100 mL
Sulfuric Acid Solution, 5.25 N 2 mL 244932
MDB
Water, deionized varies 4L 27256

Required apparatus

Description Quantity/test Unit Item no.

Cylinder, graduated, 25-mL. 1 each 50840

Flask, Erlenmeyer, 125-mL 2 each 50543
Hot plate, 7 inches x 7 inches, digital, 120 VAC 1 each 2881500
Hot plate, stirrer, 220 - 240 VAC 1 each 2881602

Required apparatus (field applications)

Description Quantity/test Unit Item no.

Heatab cookit, with 1 box Heatabs — each 220600

Heatab replacements — 21/pkg 220700

Optional reagents and apparatus

Description Unit Item no.

Sodium Hydroxide Standard Solution, 5.0 N 1L 245053

Sulfuric Acid, concentrated, ACS 500 mL 97949
Paper, pH, 0–14 pH range 100/pkg 2601300
Filter paper, folded, 12.5-cm 100/pkg 69257
Funnel, poly, 65-mm each 108367

Phosphorus, Acid Hydrolyzable, Acid Digestion 3

Consumables and replacement items (continued)
Description Unit Item no.

Thermometer, non-mercury, -10 to +225 °C each 2635700

Hydrochloric Acid, 6.0 N 1:1, 50% 500 mL 88449
Bottle, sampling, with cap, low density polyethylene, 250-mL 12/pkg 2087076

FOR TECHNICAL ASSISTANCE, PRICE INFORMATION AND ORDERING: HACH COMPANY

In the U.S.A. – Call toll-free 800-227-4224 WORLD HEADQUARTERS
Outside the U.S.A. – Contact the HACH office or distributor serving you. Telephone: (970) 669-3050
On the Worldwide Web – www.hach.com; E-mail – [email protected] FAX: (970) 669-2932

© Hach Company/Hach Lange GmbH, 1989–2013. All rights reserved. 04/2013, Edition 8
Phosphorus, Acid Hydrolyzable DOC316.53.01111

PhosVer® 3 with Acid Hydrolysis Method Method 8180

0.06 to 3.50 mg/L PO43– Test ‘N Tube™ Vials
Scope and application: For water, wastewater and seawater.

Test preparation

Instrument specific information

The table in this section shows all of the instruments that have the program for this test.
Table 1 shows adapter and light shield requirements for the instruments that use them.
To use the table, select an instrument, then read across to find the corresponding
information for this test.
Table 1 Instrument-specific information for test tubes
Instrument Adapters Light shield
DR 6000 — —
DR 5000 — —
DR 900 4846400 Cover supplied with the instrument
DR 3900 — LZV849
DR 3800 — LZV646
DR 2800 —

Before starting
Install the instrument cap on the DR 900 cell holder before ZERO or READ is pushed.
DR 3900, DR 3800, DR 2800 and DR 2700: Install the light shield in Cell Compartment #2 before this test is started.
For best results, measure the reagent blank value for each new lot of reagent. Replace the sample with deionized water in
the test procedure to get the reagent blank value. Subtract the reagent blank value from the sample results automatically
with the reagent blank adjust option.
Clean all glassware with 6.0 N (50%) hydrochloric acid, then rinse thoroughly with deionized water to remove contaminants.
The reagent that is used in this test is corrosive. Use protection for eyes and skin and be prepared to flush any spills with
running water.
Review the Safety Data Sheets (MSDS/SDS) for the chemicals that are used and use any recommended personal protective
equipment.
Dispose of reacted solutions according to local, state and federal regulations. Use the Safety Data Sheets for disposal
information for unused reagents. Consult the environmental, health and safety staff for your facility and/or local regulatory
agencies for further disposal information.

Items to collect
Description Quantity

Total and Acid Hydrolyzable Phosphorus Reagent Set 1

DRB200 Reactor 1
Funnel, micro 1
Light shield or adapter (For information about sample cells, adapters or light shields, refer to
1
Instrument specific information on page 1.)

1
Items to collect (continued)
Description Quantity
®
Pipet, TenSette , 1.0- to 10.0-mL, with pipet tips 1
Test tube rack 1
Water, deionized varies

Refer to Consumables and replacement items on page 5 for reorder information.

Sample collection and storage

• Collect samples in clean glass or plastic bottles that have been cleaned with
1:1 hydrochloric acid and rinsed with deionized water.
• Do not use a detergent that contains phosphate to clean the sample bottles. The
phosphate in the detergent will contaminate the sample.
• Analyze the samples as soon as possible for best results.
• If prompt analysis is not possible, immediately filter and keep the samples at or below
6 °C (43 °F) for a maximum of 48 hours.
• Let the sample temperature increase to room temperature before analysis.

Acid hydrolysis, Test 'N Tube procedure

Start

1. Start the 2. Start program 536 P 3. Add 5.0 mL of sample to 4. Insert the vial into the
DRB200 Reactor. Preheat Total/AH PV TNT. For the Total and Acid reactor. Close the reactor.
to 150 °C. Refer to the information about sample Hydrolyzable Test Vial.
DRB200 manual. cells, adapters or light Close the vial and mix.
shields, refer to Instrument
specific information
on page 1.
Note: Although the program
name may vary between
instruments, the program
number does not change.

2 Phosphorus, Acid Hydrolyzable, PhosVer 3 TNT Method (3.50 mg/L)

5. Start the instrument 6. When the timer expires, 7. Add 2 mL of 1.00 N 8. Clean the vial.
timer. A 30-minute reaction carefully remove the vial Sodium Hydroxide to the
time starts. from the reactor. Set the vial vial. Cap the vial and shake
in a test tube rack. Let the to mix.
vial cool to room
temperature.

Zero

9. Insert the vial into the 16- 10. Push ZERO. The 11. Add the contents of one 12. Put the cap on the vial.
mm cell holder. display shows 0.00 mg/L PhosVer 3 Powder Pillow to Shake for 20 to 30 seconds.
PO43–. the vial. The powder will not
completely dissolve .

Read

13. Start the instrument 14. Clean the vial. 15. Insert the vial into the 16. Push READ. Results
timer. A 2-minute reaction 16-mm cell holder. show in mg/L PO43–.
time starts.
Read the results within two
to eight minutes after adding
the PhosVer 3 reagent.

Interferences
Interfering substance Interference level
Aluminum More than 200 mg/L
Arsenate Interferes at any level.
Chromium More than 100 mg/L
Copper More than 10 mg/L

Phosphorus, Acid Hydrolyzable, PhosVer 3 TNT Method (3.50 mg/L) 3

Interfering substance Interference level
Sulfide More than 9 mg/L sulfide. Remove sulfide interference as follows:

1. Measure 25 mL of sample into a 50-mL beaker.

2. Add Bromine Water drop-wise with constant swirling until a permanent yellow color remains.
3. Add Phenol Solution drop-wise with constant swirling until the yellow color just disappears.

Use this sample in the test procedure.

Iron More than 100 mg/L
Nickel More than 300 mg/L
Highly buffered samples Can prevent the correct pH adjustment of the sample by the reagents. Sample pretreatment may
or extreme sample pH be necessary.
Silica More than 50 mg/L
Silicate More than 10 mg/L
Turbidity or color May cause inconsistent results. The acid in the powder pillow can dissolve some of the
suspended particles and the desorption of orthophosphate from the particles can vary. For highly
turbid or colored samples, add the contents of one Phosphate Pretreatment Powder Pillow to
25 mL of sample. Mix well. Use this solution to zero the instrument.
Zinc More than 80 mg/L

Accuracy check
Standard additions method (sample spike)
Use the standard additions method (for applicable instruments) to validate the test
procedure, reagents and instrument and to find if there is an interference in the sample.
Items to collect:
• Phosphate 2-mL Ampule Standard, 50-mg/L as PO43–
• Ampule breaker
• Pipet, TenSette®, 0.1–1.0 mL and tips
• Mixing cylinders, 25-mL (3)

1. Use the test procedure to measure the concentration of the sample, then keep the
(unspiked) sample in the instrument.
2. Go to the Standard Additions option in the instrument menu.
3. Select the values for standard concentration, sample volume and spike volumes.
4. Open the standard solution.
5. Prepare three spiked samples: use the TenSette pipet to add 0.1 mL, 0.2 mL and
0.3 mL of the standard solution, respectively, to three 25-mL portions of fresh sample.
Mix well.
6. Use the test procedure to measure the concentration of each of the spiked samples.
Start with the smallest sample spike. Measure each of the spiked samples in the
instrument.
7. Select Graph to compare the expected results to the actual results.
Note: If the actual results are significantly different from the expected results, make sure that
the sample volumes and sample spikes are measured accurately. The sample volumes and
sample spikes that are used should agree with the selections in the standard additions menu. If
the results are not within acceptable limits, the sample may contain an interference.

Standard solution method

Use the standard solution method to validate the test procedure, reagents and
instrument.

4 Phosphorus, Acid Hydrolyzable, PhosVer 3 TNT Method (3.50 mg/L)

 Items to collect:
• 3.0-mg/L phosphate standard solution

1. Use the test procedure to measure the concentration of the standard solution.
2. Compare the expected result to the actual result.
Note: The factory calibration can be adjusted slightly with the standard adjust option so that the
instrument shows the expected value of the standard solution. The adjusted calibration is then
used for all test results. This adjustment can increase the test accuracy when there are slight
variations in the reagents or instruments.

Method performance
The method performance data that follows was derived from laboratory tests that were
measured on a spectrophotometer during ideal test conditions. Users may get different
results under different test conditions.
Program Standard Precision (95% Confidence Interval) Sensitivity
Concentration change per 0.010 Abs change
536 3.00 mg/L PO43– 2.93–3.07 mg/L PO43– 0.06 mg/L PO43–

Summary of method
Phosphates in condensed inorganic forms (meta-, pyro-, or other polyphosphates) are
converted to reactive orthophosphate before measurement. The sample is pretreated with
acid and heat to hydrolyze the condensed inorganic forms to orthophosphate.
Orthophosphate reacts with molybdate in an acid medium to produce a mixed
phosphate/molybdate complex. Ascorbic acid then reduces the complex, which gives an
intense molybdenum blue color. The measurement wavelength is 880 nm for
spectrophotometers or 610 nm for colorimeters.
Pollution prevention and waste management
Reacted samples contain molybdenum and must be disposed of as a hazardous waste.
Dispose of reacted solutions according to local, state and federal regulations.
Consumables and replacement items
Required reagents

Description Quantity/test Unit Item no.

Water, deionized varies 100 mL 27242

Total and Acid Hydrolyzable Phosphorus Reagent Set — 50 tests 2742745
Includes:
®
PhosVer 3 Phosphate Reagent Powder Pillow, 10-mL 1 50/pkg 2106046
Potassium Persulfate Powder Pillows 1 pillow 50/pkg 2084766
Sodium Hydroxide, 1.54 N varies 100 mL 2743042
Sodium Hydroxide Standard Solution, 1.00 N 2 mL 100 mL 104542
Total and Acid Hydrolyzable Test Vials (not sold separately) 1 50/pkg —

Required apparatus

Description Quantity/test Unit Item no.

DRB200 Reactor, 110 V, 15 x 16 mm 1 each LTV082.53.40001

DRB200 Reactor, 220 V, 15 x 16 mm 1 each LTV082.52.40001
Funnel, micro, poly 1 each 2584335
Pipet, volumetric, Class A, 2.00-mL 1 each 1451536

Phosphorus, Acid Hydrolyzable, PhosVer 3 TNT Method (3.50 mg/L) 5

Consumables and replacement items (continued)
Description Quantity/test Unit Item no.

Pipet, volumetric, Class A, 5.00-mL 1 each 1451537

Pipet filler, safety bulb 1 each 1465100
®
Pipet, TenSette , 1.0- to 10.0-mL 1 each 1970010
Pipet Tips, for TenSette Pipet 1970010 2 250/pkg 2199725
Test tube rack 1 each 1864100

Recommended standards

Description Unit Item no.

Drinking Water Standard, Mixed Parameter, Inorganic for F-, NO3, PO4, SO4 500 mL 2833049
®
Phosphate Standard Solution, 50-mg/L, 10-mL Voluette Ampules 16/pkg 17110
Phosphate Standard Solution, 1-mg/L as PO4 500 mL 256949
Phosphate Standard Solution, 3-mg/L as PO4 946 mL 2059716
Wastewater, Effluent Inorganics, for NH3-N, NO3-N, PO4, COD, SO4, TOC 500 mL 2833249

Optional reagents and apparatus

Description Unit Item no.

Bromine Water, 30 g/L 29 mL 221120

Cylinder, mixing, 25-mL each 189640
Hydrochloric Acid, 6.0 N 1:1, 50% 500 mL 88449
Phenol Solution, 30-g/L 29 mL 211220
Paper, pH, 0–14 pH range 100/pkg 2601300
Filter paper, folded, 12.5-cm 100/pkg 69257
Funnel, poly, 65-mm each 108367
Thermometer, non-mercury, -10 to +225 °C each 2635700
Bottle, sampling, with cap, low density polyethylene, 250-mL 12/pkg 2087076
Pipet tips for TenSette Pipet 1970010 50/pkg 2199796
Beaker, 50-mL each 50041H

Optional standards

Description Unit Item no.

®
Ampule Breaker, Voluette ampules each 2196800
Phosphate Standard Solution, 3-mg/L as PO4 946 mL 2059716
Phosphate Standard Solution, 10-mg/L as PO4 946 mL 1420416
Phosphate Standard Solution, 15-mg/L as PO4 100 mL 1424342
Phosphate Standard Solution, 30-mg/L as PO4 946 mL 1436716
®
Phosphate Standard Solution, 50-mg/L, 10-mL Voluette Ampules 16/pkg 17110
Phosphate Standard Solution, 100-mg/L as PO4 100 mL 1436832
Phosphate Standard Solution, 10-mL Ampule, 500 mg/L as PO4 16/pkg 1424210
Phosphate Standard Solution, 500-mg/L as PO4 100 mL 1424232

6 Phosphorus, Acid Hydrolyzable, PhosVer 3 TNT Method (3.50 mg/L)

Phosphorus, Acid Hydrolyzable, PhosVer 3 TNT Method (3.50 mg/L) 7
 FOR TECHNICAL ASSISTANCE, PRICE INFORMATION AND ORDERING: HACH COMPANY
In the U.S.A. – Call toll-free 800-227-4224 WORLD HEADQUARTERS
Outside the U.S.A. – Contact the HACH office or distributor serving you. Telephone: (970) 669-3050
On the Worldwide Web – www.hach.com; E-mail – [email protected] FAX: (970) 669-2932

© Hach Company/Hach Lange GmbH, 1989–2013. All rights reserved. 04/2013, Edition 8
Phosphorus, Reactive DOC316.53.01113

(Orthophosphate)
Amino Acid Method1 Method 8178
0.23 to 30.00 mg/L PO43– Reagent Solution
Scope and application: For water, wastewater and seawater.
1 Adapted from Standard Methods for the Examination of Water and Wastewater.

Test preparation

Instrument specific information

The table in this section shows all of the instruments that have the program for this test.
Table 1 shows sample cell and orientation requirements for reagent addition tests, such
as powder pillow or bulk reagent tests.
To use the table, select an instrument, then read across to find the corresponding
information for this test.
Table 1 Instrument-specific information for reagent solution
Instrument Sample cell orientation Sample cell
DR 6000 The fill line is to the right. 2495402
DR 3800
DR 2800
DR 2700
DR 5000 The fill line is toward the user.
DR 3900
DR 900 The orientation mark is toward the user. 2401906

Before starting
Install the instrument cap on the DR 900 cell holder before ZERO or READ is pushed.
For best results, measure the reagent blank value for each new lot of reagent. Replace the sample with deionized water in
the test procedure to get the reagent blank value. Subtract the reagent blank value from the sample results automatically
with the reagent blank adjust option.
The contents of one Amino Acid Reagent Powder Pillow can be used as an alternative to the 1 mL of Amino Acid Reagent
Solution in the test procedure.
Review the Safety Data Sheets (MSDS/SDS) for the chemicals that are used and use any recommended personal protective
equipment.
Dispose of reacted solutions according to local, state and federal regulations. Use the Safety Data Sheets for disposal
information for unused reagents. Consult the environmental, health and safety staff for your facility and/or local regulatory
agencies for further disposal information.

1
Items to collect
Description Quantity

Amino Acid Reagent 1 mL

Cylinder, 25-mL, graduated mixing 1
Molybdate Reagent 1 mL
Sample cells (For information about sample cells, adapters or light shields, refer to Instrument
2
specific information on page 1.)

Refer to Consumables and replacement items on page 5 for reorder information.

Sample collection and storage

• Collect samples in clean glass or plastic bottles that have been cleaned with
1:1 hydrochloric acid and rinsed with deionized water.
• Do not use a detergent that contains phosphate to clean the sample bottles. The
phosphate in the detergent will contaminate the sample.
• Analyze the samples as soon as possible for best results.
• If prompt analysis is not possible, immediately filter and keep the samples at or below
6 °C (43 °F) for a maximum of 48 hours.
• Let the sample temperature increase to room temperature before analysis.

Powder pillow procedure

Start

1. Start program 485 P 2. Prepare the sample: Fill 3. Prepare the sample: 4. Add 1 mL of Amino Acid
React. Amino. For a mixing cylinder to the 25- Add 1 mL of Molybdate Reagent Solution.
information about sample mL line with sample. Reagent.
cells, adapters or light
shields, refer to Instrument
specific information
on page 1.
Note: Although the program
name may vary between
instruments, the program
number does not change.

2 Phosphorus, Reactive, Amino Acid Method (30.00 mg/L)

5. Close the cylinder. Invert 6. Start the instrument 7. Prepare the blank: Fill a 8. When the timer expires,
the cylinder several times to timer. A 10-minute reaction sample cell with 10 mL of clean the blank.
mix. time starts. untreated sample.
A blue color shows if Prepare the blank while the
phosphate is present in the timer is counting down.
sample.

Zero

9. Insert the blank into the 10. Push ZERO. The 11. Fill a second sample 12. Clean the prepared
cell holder. display shows 0.00 mg/L cell with 10-mL of the sample.
PO43–. prepared sample.

Read

13. Insert the prepared 14. Push READ. Results

sample into the cell holder. show in mg/L PO43–.

Interferences
Interfering Interference level
substance
Calcium More than 10,000 mg/L as CaCO3
Chloride More than 150,000 mg/L Cl–
Colored samples Add 1 mL of 10 N Sulfuric Acid Standard Solution to another 25-mL sample. Use this instead of
untreated sample as the blank to zero the instrument. Use a pipet and pipet filler to measure the
sulfuric acid standard.
High salt levels (Na+) May cause low results. To eliminate this interference, dilute the sample until two successive dilutions
give about the same result.
Magnesium More than 40,000 mg/L as CaCO3

Phosphorus, Reactive, Amino Acid Method (30.00 mg/L) 3

Interfering Interference level
substance
Nitrite (NO2–) Bleaches the blue color. Remove nitrite interference by adding 0.05 g of sulfamic acid to 25 mL
sample. Swirl to mix. Use this treated sample in the test procedure.
Phosphates, high As the concentration of phosphate increases, the color changes from blue to green, then to yellow
levels (PO43–) and finally to brown. The brown color may suggest a concentration as high as 100,000 mg/L PO43–. If
a color other than blue is formed, dilute the sample and retest.
Sulfide (S2–) Sulfide interferes. For samples with sulfide concentration less than 5 mg/L sulfide interference may
be removed by oxidation with Bromine Water as follows:

1. Measure 50 mL of sample into an Erlenmeyer flask.

2. Add Bromine Water drop-wise with constant swirling until permanent yellow color develops.
3. Add Phenol Solution by drops until the yellow color just disappears. Use this treated sample in
the test procedure.

Temperature For best results, sample temperature should be 21 ±3 °C (70 ±5 °F).

Turbidity May give inconsistent results for two reasons. Some suspended particles may dissolve because of
the acid used in the test. Also, desorption of orthophosphate from particles may occur. For highly
turbid samples, add 1 mL of 10 N Sulfuric Acid Standard Solution to another 25-mL sample. Use this
instead of untreated sample as the blank to zero the instrument. Use a pipet and pipet filler to
measure the sulfuric acid standard.
Highly buffered Can prevent the correct pH adjustment of the sample by the reagents. Sample pretreatment may be
samples or extreme necessary.
sample pH

Accuracy check
Standard additions method (sample spike)
Use the standard additions method (for applicable instruments) to validate the test
procedure, reagents and instrument and to find if there is an interference in the sample.
Items to collect:
• Phosphate 2-mL Ampule Standard, 500-mg/L PO43–
• Ampule breaker
• Pipet, TenSette®, 0.1–1.0 mL and tips
• Mixing cylinders, 25-mL (3)

1. Use the test procedure to measure the concentration of the sample, then keep the
(unspiked) sample in the instrument.
2. Go to the Standard Additions option in the instrument menu.
3. Select the values for standard concentration, sample volume and spike volumes.
4. Open the standard solution.
5. Prepare three spiked samples: use the TenSette pipet to add 0.1 mL, 0.2 mL and
0.3 mL of the standard solution, respectively, to three 25-mL portions of fresh sample.
Mix well.
6. Use the test procedure to measure the concentration of each of the spiked samples.
Start with the smallest sample spike. Measure each of the spiked samples in the
instrument.
7. Select Graph to compare the expected results to the actual results.
Note: If the actual results are significantly different from the expected results, make sure that
the sample volumes and sample spikes are measured accurately. The sample volumes and
sample spikes that are used should agree with the selections in the standard additions menu. If
the results are not within acceptable limits, the sample may contain an interference.

Standard solution method

Use the standard solution method to validate the test procedure, reagents and
instrument.

4 Phosphorus, Reactive, Amino Acid Method (30.00 mg/L)

 Items to collect:
• 10-mg/L Phosphate Standard Solution

1. Use the test procedure to measure the concentration of the standard solution.
2. Compare the expected result to the actual result.
Note: The factory calibration can be adjusted slightly with the standard adjust option so that the
instrument shows the expected value of the standard solution. The adjusted calibration is then
used for all test results. This adjustment can increase the test accuracy when there are slight
variations in the reagents or instruments.

Method performance
The method performance data that follows was derived from laboratory tests that were
measured on a spectrophotometer during ideal test conditions. Users may get different
results under different test conditions.
Program Standard Precision (95% Confidence Interval) Sensitivity
Concentration change per 0.010 Abs change
485 10.00 mg/L PO43– 9.86–10.14 mg/L PO43– 0.20 mg/L PO43–

Summary of method
In a highly acidic solution, ammonium molybdate reacts with orthophosphate to form
molybdophosphoric acid. This complex is then reduced by the amino acid reagent to yield
an intensely colored molybdenum blue compound. The measurement wavelength is
530 nm for spectrophotometers or 520 nm for colorimeters.
Consumables and replacement items
Required reagents

Description Quantity/test Unit Item no.

High Range Reactive Phosphorus Reagent Set — 100 tests 2244100

Includes:
100 mL
Amino Acid Reagent 1 mL 193432
MDB
100 mL
Molybdate Reagent 1 mL 223632
MDB

Required apparatus

Description Quantity/test Unit Item no.

Cylinder, graduated mixing, 25-mL 1 each 189640

Recommended standards and apparatus

Description Unit Item no.

Phosphate Standard Solution, 10-mg/L as PO4 946 mL 1420416

®
Phosphate Standard Solution, 2-mL PourRite Ampule, 500-mg/L PO43– 16/pkg 1424220
Wastewater, Effluent Inorganics, for NH3-N, NO3-N, PO4, COD, SO4, TOC 500 mL 2833249
Wastewater Influent Standard, Mixed Parameter, for NH3-N, NO3-N, PO4, COD, SO4,
500 mL 2833149
TOC
Water, deionized 4L 27256
®
Ampule Breaker, PourRite ampules each 2484600

Phosphorus, Reactive, Amino Acid Method (30.00 mg/L) 5

Optional reagents and apparatus

Description Unit Item no.

Amino Acid Reagent Powder Pillow 100/pkg 80499

Bromine Water, 30 g/L 29 mL 221120
Flask, Erlenmeyer, 125-mL each 50543
Hydrochloric Acid, 6.0 N 1:1, 50% 500 mL 88449
Phenol Solution, 30-g/L 29 mL 211220
Sulfamic Acid, 454 g each 234401
Sulfuric Acid Standard Solution, 10 N 1000 mL 93153
®
Pipet, TenSette , 0.1–1.0 mL each 1970001
Pipet tips for TenSette Pipet 1970001 50/pkg 2185696
Pipet tips for TenSette Pipet 1970001 1000/pkg 2185628
Paper, pH, 0–14 pH range 100/pkg 2601300
Filter paper, folded, 12.5-cm 100/pkg 69257
Funnel, poly, 65-mm each 108367
Thermometer, non-mercury, -10 to +225 °C each 2635700
Bottle, sampling, with cap, low density polyethylene, 250-mL 12/pkg 2087076

Optional standards

Description Unit Item no.

®
Ampule Breaker, Voluette ampules each 2196800
Phosphate Standard Solution, 3-mg/L as PO4 946 mL 2059716
Phosphate Standard Solution, 15-mg/L as PO4 100 mL 1424342
Phosphate Standard Solution, 30-mg/L as PO4 946 mL 1436716
®
Phosphate Standard Solution, 50-mg/L, 10-mL Voluette Ampules 16/pkg 17110
Phosphate Standard Solution, 100-mg/L as PO4 100 mL 1436832
Phosphate Standard Solution, 10-mL Ampule, 500 mg/L as PO4 16/pkg 1424210
Phosphate Standard Solution, 500-mg/L as PO4 100 mL 1424232

FOR TECHNICAL ASSISTANCE, PRICE INFORMATION AND ORDERING: HACH COMPANY

In the U.S.A. – Call toll-free 800-227-4224 WORLD HEADQUARTERS
Outside the U.S.A. – Contact the HACH office or distributor serving you. Telephone: (970) 669-3050
On the Worldwide Web – www.hach.com; E-mail – [email protected] FAX: (970) 669-2932

© Hach Company/Hach Lange GmbH, 1989–2013. All rights reserved. 04/2013, Edition 8
Phosphorus, Reactive DOC316.53.01115

(Orthophosphate)
Molybdovanadate Method1 Method 8114
®
0.3 to 45.0 mg/L PO43– Reagent Solution or AccuVac Ampuls
Scope and application: For water and wastewater.
1 Adapted from Standard Methods for the Examination of Water and Wastewater.

Test preparation

Instrument-specific information
The tables in this section show all of the instruments that have the program for this test.
Table 1 shows sample cell and orientation requirements for reagent addition tests, such
as powder pillow or bulk reagent tests. Table 2 shows sample cell and adapter
requirements for AccuVac Ampul tests.
To use either table, select an instrument, then read across to find the corresponding
information for this test.
Table 1 Instrument-specific information for reagent solution
Instrument Sample cell orientation Sample cell
DR 6000 The fill line is to the right. 2495402
DR 3800
DR 2800
DR 2700
DR 5000 The fill line is toward the user.
DR 3900
DR 900 The orientation mark is toward the user. 2401906

Table 2 Instrument-specific information for AccuVac Ampuls

Instrument Adapter Sample cell
DR 6000 — 2427606
DR 5000
DR 900
DR 3900 LZV846 (A)
DR 3800 LZV584 (C) 2122800
DR 2800
DR 2700

Before starting
Install the instrument cap on the DR 900 cell holder before ZERO or READ is pushed.
For best results, the sample temperature should be 20–25 °C (68–77 °F).

1
Review the Safety Data Sheets (MSDS/SDS) for the chemicals that are used and use any recommended personal protective
equipment.
Dispose of reacted solutions according to local, state and federal regulations. Use the Safety Data Sheets for disposal
information for unused reagents. Consult the environmental, health and safety staff for your facility and/or local regulatory
agencies for further disposal information.

Items to collect
Reagent solution

Description Quantity

Molybdovanadate reagent 1.0 mL

Sample cells (For information about sample cells, adapters or light shields, refer to Instrument-
2
specific information on page 1.)

Refer to Consumables and replacement items on page 6 for reorder information.

AccuVac Ampuls

Description Quantity
®
Molybdovanadate reagent AccuVac Ampuls 2
Beaker, 50-mL 1
Stoppers for 18 mm tubes and AccuVac Ampuls 2

Refer to Consumables and replacement items on page 6 for reorder information.

Sample collection and storage

• Collect samples in clean glass or plastic bottles that have been cleaned with
1:1 hydrochloric acid and rinsed with deionized water.
• Do not use a detergent that contains phosphate to clean the sample bottles. The
phosphate in the detergent will contaminate the sample.
• Analyze the samples as soon as possible for best results.
• If prompt analysis is not possible, immediately filter and keep the samples at or below
6 °C (43 °F) for a maximum of 48 hours.
• Let the sample temperature increase to room temperature before analysis.

2 Phosphorus, Reactive, Molybdovanadate Method (45.0 mg/L)

Reagent solution procedure

Start

1. Start program 480 P 2. Prepare the blank: Fill a 3. Prepare the sample: Fill 4. Add 0.5 mL of
React. Mo. For information sample cell with 10 mL of a second sample cell with Molybdovanadate reagent to
about sample cells, deionized water. 10 mL of sample. each cell.
adapters or light shields,
refer to Instrument-specific
information on page 1.
Note: Although the program
name may vary between
instruments, the program
number does not change.

5. Swirl to mix. 6. Start the instrument 7. When the timer expires, 8. Insert the blank into the
timer. A 7-minute reaction clean the blank. cell holder.
time starts.
If the sample concentration
is greater than 30 mg/L
PO43–, read at exactly
7 minutes or make a
1:1 dilution of the sample
and repeat the test.

Zero Read

9. Push ZERO. The display 10. Clean the prepared 11. Insert the prepared 12. Push READ. Results
shows 0.0 mg/L PO43–. sample. sample into the cell holder. show in mg/L PO43–.

Phosphorus, Reactive, Molybdovanadate Method (45.0 mg/L) 3

AccuVac Ampuls procedure

Start

1. Start program 482 P 2. Prepare the blank: Pour 3. Prepare the sample: 4. Start the instrument
React. Mo. AV. For at least 40 mL of deionized Collect at least 40 mL of timer. A 7-minute reaction
information about sample water into a 50-mL beaker. sample in a 50-mL beaker. time starts.
cells, adapters or light Fill an AccuVac Ampul with Fill the AccuVac Ampul with If the sample concentration
shields, refer to Instrument- deionized water. Keep the sample. Keep the tip is greater than 30 mg/L
specific information tip immersed while the immersed while the Ampul PO43–, read at exactly
on page 1. Ampul fills completely. fills completely. 7 minutes or make a
Note: Although the program 1:1 dilution of the sample
name may vary between and repeat the test.
instruments, the program
number does not change.

Zero

5. When the timer expires, 6. Insert the blank AccuVac 7. Push ZERO. The display 8. Clean the AccuVac
clean the blank AccuVac Ampul into the cell holder. shows 0.0 mg/L PO43–. Ampul.
Ampul.

Read

9. Insert the prepared 10. Push READ. Results

sample AccuVac Ampul into show in mg/L PO43–.
the cell holder.

4 Phosphorus, Reactive, Molybdovanadate Method (45.0 mg/L)

Interferences
Table 3 shows the interferences and interference levels. Table 4 shows the substances
that do not interfere at or below the indicated levels.
Table 3 Interfering substances
Interfering substance Interference level
Arsenate Causes a positive interference if the sample is warm when the reagent is added. The sample can
be gently warmed to room temperature without interference.
Iron, ferrous Causes a blue color which interferes at more than 100 mg/L.
Molybdate Causes a negative interference at more than 1000 mg/L.
Silica Causes a positive interference if the sample is warm when the reagent is added. The sample can
be gently warmed to room temperature without interference.
Sulfide Causes a negative interference. Correct for this interference as follows:

1. Measure 50 mL of sample into an Erlenmeyer flask.

2. Add Bromine Water drop-wise with constant swirling until a permanent yellow color remains.
3. Add Phenol Solution drop-wise until the yellow color just disappears.

Use this sample in the test procedure.

Highly buffered samples Can prevent the correct pH adjustment of the sample by the reagents. Sample pretreatment may
or extreme sample pH be necessary. The pH should be approximately 7.
Fluoride, thorium, Causes a negative interference.
bismuth, thiosulfate or
thiocyanate
Temperature Temperatures below 18 °C (64 °F) cause a negative interference. Temperatures above 25 °C
(77 °F) cause a positive interference. The sample can be gently warmed to room temperature
without interference.

Table 4 Substances that do not interfere at less than 1000 mg/L

Pyrophosphate Tetraborate Selenate Benzoate
Citrate Oxalate Lactate Tartrate
Formate Salicylate Al3+ Fe3+
Mg2+ Ca2+ Ba2+ Sr2+
Li+ Na+ K+ NH4+
Cd2+ Mn2+ NO3– NO2–
SO42– SO32– Pb2+ Hg+
Hg2+ Sn2+ Cu2+ Ni2+
Ag+ U4+ Zr4+ AsO3–
Br– CO32– ClO4– CN–
IO3– SiO44– — —

Accuracy check
Standard additions method (sample spike)
Use the standard additions method (for applicable instruments) to validate the test
procedure, reagents and instrument and to find if there is an interference in the sample.
Items to collect:
• Phosphate standard solution, 500 mg/L PO43– ampule
• Ampule breaker
• Pipet, TenSette®, 0.1–1.0 mL and tips

Phosphorus, Reactive, Molybdovanadate Method (45.0 mg/L) 5

 • Mixing cylinders, 25-mL (3)

1. Use the test procedure to measure the concentration of the sample, then keep the
(unspiked) sample in the instrument.
2. Go to the Standard Additions option in the instrument menu.
3. Select the values for standard concentration, sample volume and spike volumes.
4. Open the standard solution.
5. Prepare three spiked samples: use the TenSette pipet to add 0.1 mL, 0.2 mL and
0.3 mL of the standard solution, respectively, to three 25-mL portions of fresh sample.
Mix well.
®
Note: For AccuVac Ampuls, add 0.1 mL, 0.2 mL and 0.3 mL of the standard solution to three
25-mL portions of fresh sample.
6. Use the test procedure to measure the concentration of each of the spiked samples.
Start with the smallest sample spike. Measure each of the spiked samples in the
instrument.
7. Select Graph to compare the expected results to the actual results.
Note: If the actual results are significantly different from the expected results, make sure that
the sample volumes and sample spikes are measured accurately. The sample volumes and
sample spikes that are used should agree with the selections in the standard additions menu. If
the results are not within acceptable limits, the sample may contain an interference.

Standard solution method

Use the standard solution method to validate the test procedure, reagents and
instrument.
Items to collect:
• 10 mg/L phosphate standard solution

1. Use the test procedure to measure the concentration of the standard solution.
2. Compare the expected result to the actual result.
Note: The factory calibration can be adjusted slightly with the standard adjust option so that the
instrument shows the expected value of the standard solution. The adjusted calibration is then
used for all test results. This adjustment can increase the test accuracy when there are slight
variations in the reagents or instruments.

Method performance
The method performance data that follows was derived from laboratory tests that were
measured on a spectrophotometer during ideal test conditions. Users may get different
results under different test conditions.
Program Standard Precision (95% Confidence Interval) Sensitivity
Concentration change per 0.010 Abs change
480 30.0 mg/L PO43– 29.6–30.4 mg/L PO43– 0.3 mg/L PO43–
482 30.0 mg/L PO43– 29.7–30.3 mg/L PO43– 0.3 mg/L PO43–

Summary of method
In the molybdovanadate method, orthophosphate reacts with molybdate in an acid
medium to produce a mixed phosphate/molybdate complex. In the presence of vanadium,
yellow molybdovanadophosphoric acid is formed. The intensity of the yellow color is
proportional to the phosphate concentration. The measurement wavelength is 430 nm for
spectrophotometers or 420 nm for colorimeters.
Consumables and replacement items
Note: Product and Article numbers may vary for some selling regions. Contact the appropriate
distributor or refer to the company website for contact information.

6 Phosphorus, Reactive, Molybdovanadate Method (45.0 mg/L)

Required reagents

Description Quantity/Test Unit Item no.

100 mL
Molybdovanadate Reagent 1.0 mL 2076032
MDB
OR
®
Molybdovanadate Reagent AccuVac Ampul 2 25/pkg 2525025
Water, deionized varies 4L 27256

Required apparatus

Description Quantity/Test Unit Item no.

AccuVac Snapper 1 each 2405200

Beaker, 50-mL 1 each 50041H
Stoppers for 18-mm tubes and AccuVac Ampuls 2 6/pkg 173106

Recommended standards and apparatus

Description Unit Item no.

®
Ampule Breaker, Voluette ampules each 2196800
Phosphate Standard Solution, 10-mg/L as PO4 946 mL 1420416
Phosphate Standard Solution, 10-mL Ampule, 500 mg/L as PO4 16/pkg 1424210
Wastewater Influent Standard, Mixed Parameter, for NH3-N, NO3-N, PO4, COD, SO4,
500 mL 2833149
TOC

Optional reagents and apparatus

Description Unit Item no.

®
AccuVac Drainer each 4103600
Bottle, sampling, with cap, low density polyethylene, 250-mL 12/pkg 2087076
Bromine Water, 30-g/L 29 mL 221120
Cylinder, mixing, 25-mL each 2088640
Hydrochloric Acid, 6.0 N 1:1, 50% 500 mL 88449
Paper, pH, 0–14 pH range 100/pkg 2601300
Phenol Solution, 30-g/L 29 mL 211220
Phosphate Standard Solution, 3-mg/L as PO4 946 mL 2059716
Phosphate Standard Solution, 15-mg/L as PO4 100 mL 1424342
Phosphate Standard Solution, 30-mg/L as PO4 946 mL 1436716
®
Phosphate Standard Solution, 50-mg/L, 10-mL Voluette Ampules 16/pkg 17110
Phosphate Standard Solution, 100-mg/L as PO4 100 mL 1436832
Phosphate Standard Solution, 500-mg/L as PO4 100 mL 1424232
®
Pipet, TenSette , 0.1–1.0 mL each 1970001
®
Pipet, TenSette , 1.0 to 10.0 mL each 1970010
Pipet tips for TenSette Pipet 1970001 50/pkg 2185696

Phosphorus, Reactive, Molybdovanadate Method (45.0 mg/L) 7

Consumables and replacement items (continued)
Description Unit Item no.

Pipet tips for TenSette Pipet 1970001 1000/pkg 2185628

Thermometer, non-mercury, -10 to +225 °C each 2635700

FOR TECHNICAL ASSISTANCE, PRICE INFORMATION AND ORDERING: HACH COMPANY

In the U.S.A. – Call toll-free 800-227-4224 WORLD HEADQUARTERS
Outside the U.S.A. – Contact the HACH office or distributor serving you. Telephone: (970) 669-3050
On the Worldwide Web – www.hach.com; E-mail – [email protected] FAX: (970) 669-2932

© Hach Company/Hach Lange GmbH, 1989–2013. All rights reserved. 04/2013, Edition 8
Phosphorus, Reactive DOC316.53.01116

(Orthophosphate)
Molybdovanadate Method1 Method 8114
1.0 to 100.0 mg/L PO43– (HR) Test ‘N Tube™ Vials
Scope and application: For water and wastewater.
1 Adapted from Standard Methods for the Examination of Water and Wastewater.

Test preparation

Instrument specific information

The table in this section shows all of the instruments that have the program for this test.
Table 1 shows adapter and light shield requirements for the instruments that use them.
To use the table, select an instrument, then read across to find the corresponding
information for this test.
Table 1 Instrument-specific information for test tubes
Instrument Adapters Light shield
DR 6000 — —
DR 5000 — —
DR 900 4846400 Cover supplied with the instrument
DR 3900 — LZV849
DR 3800 — LZV646
DR 2800 —

Before starting
Install the instrument cap on the DR 900 cell holder before ZERO or READ is pushed.
DR 3900, DR 3800, DR 2800 and DR 2700: Install the light shield in Cell Compartment #2 before this test is started.
The blank vial that is prepared in the test procedure can be used more than once. At room temperature, the reagent blank is
stable for a maximum of three weeks. Prepare a new blank vial when a new lot of reagent is used.
The 7-minute reaction time in the test procedure is for samples that are at 23 °C (73 °F). If the sample temperature is 13 °C
(55 °F), wait 15 minutes. If the sample temperature is 33 °C (91 °F), wait 2 minutes.
The reagent that is used in this test is corrosive. Use protection for eyes and skin and be prepared to flush any spills with
running water.
Review the Safety Data Sheets (MSDS/SDS) for the chemicals that are used and use any recommended personal protective
equipment.
Dispose of reacted solutions according to local, state and federal regulations. Use the Safety Data Sheets for disposal
information for unused reagents. Consult the environmental, health and safety staff for your facility and/or local regulatory
agencies for further disposal information.

Items to collect
Description Quantity

High Range Reactve Phosphorus Test 'N Tube Vials 1

Light shield or adapter (For information about sample cells, adapters or light shields, refer to
1
Instrument specific information on page 1.)

1
Items to collect (continued)
Description Quantity
®
Pipet, TenSette , 1.0- to 10.0-mL, with pipet tips 1
Test tube rack 1
Water, deionized 5 mL

Refer to Consumables and replacement items on page 5 for reorder information.

Sample collection and storage

• Collect samples in clean glass or plastic bottles that have been cleaned with
1:1 hydrochloric acid and rinsed with deionized water.
• Do not use a detergent that contains phosphate to clean the sample bottles. The
phosphate in the detergent will contaminate the sample.
• Analyze the samples as soon as possible for best results.
• If prompt analysis is not possible, immediately filter and keep the samples at or below
6 °C (43 °F) for a maximum of 48 hours.
• Let the sample temperature increase to room temperature before analysis.

Molybdovanadate method for Test 'N Tubes

Start

1. Start program 540 P 2. Prepare the blank: Add 3. Put the cap on the vial. 4. Prepare the sample:
React. HR TNT. For 5.0 mL of deionized water to Invert to mix. Add 5.0 mL of sample to a
information about sample a Reactive High Range second Reactive High
cells, adapters or light Phosphorus Test 'N Tube Range Phosphorus Test 'N
shields, refer to Instrument Vial. Tube Vial.
specific information
on page 1.
Note: Although the program
name may vary between
instruments, the program
number does not change.

2 Phosphorus, Reactive, Molybdovanadate TNT Method (100.0 mg/L)

5. Put the cap on the vial. 6. Start the instrument 7. When the timer expires, 8. Insert the blank into the
Invert to mix. timer. A 7-minute reaction clean the blank. cell holder.
time starts.
Measure the prepared
sample within two minutes
after the timer expires.

Zero Read

9. Push ZERO. The display 10. Clean the sample vial. 11. Insert the sample vial 12. Push READ. Results
shows 0.0 mg/L PO43–. into the 16-mm cell holder. show in mg/L PO43–.

Interferences
Table 2 shows the interferences and interference levels. Table 3 shows the substances
that do not interfere at or below the indicated levels.
Table 2 Interfering substances
Interfering substance Interference level
Arsenate Causes a positive interference if the sample is warm when the reagent is added. The sample can
be gently warmed to room temperature without interference.
Iron, ferrous Causes a blue color which interferes at more than 100 mg/L.
Molybdate Causes a negative interference at more than 1000 mg/L.
Silica Causes a positive interference if the sample is warm when the reagent is added. The sample can
be gently warmed to room temperature without interference.
Sulfide Causes a negative interference. Correct for this interference as follows:

1. Measure 50 mL of sample into an Erlenmeyer flask.

2. Add Bromine Water drop-wise with constant swirling until a permanent yellow color remains.
3. Add Phenol Solution drop-wise until the yellow color just disappears.

Use this sample in the test procedure.

Highly buffered samples Can prevent the correct pH adjustment of the sample by the reagents. Sample pretreatment may
or extreme sample pH be necessary. The pH should be approximately 7.

Phosphorus, Reactive, Molybdovanadate TNT Method (100.0 mg/L) 3

 Table 2 Interfering substances (continued)
Interfering substance Interference level
Fluoride, thorium, Causes a negative interference.
bismuth, thiosulfate or
thiocyanate
Temperature Temperatures below 18 °C (64 °F) cause a negative interference. Temperatures above 25 °C
(77 °F) cause a positive interference. The sample can be gently warmed to room temperature
without interference.

Table 3 Substances that do not interfere at less than 1000 mg/L

Pyrophosphate Tetraborate Selenate Benzoate
Citrate Oxalate Lactate Tartrate
Formate Salicylate Al3+ Fe3+
Mg2+ Ca2+ Ba2+ Sr2+
Li+ Na+ K+ NH4+
Cd2+ Mn2+ NO3– NO2–
SO42– SO32– Pb2+ Hg+
Hg2+ Sn2+ Cu2+ Ni2+
Ag+ U4+ Zr4+ AsO3–
Br– CO32– ClO4– CN–
IO3– SiO44– — —

Accuracy check
Standard additions method (sample spike)
Use the standard additions method (for applicable instruments) to validate the test
procedure, reagents and instrument and to find if there is an interference in the sample.
Items to collect:
• 10-mL Voluette® Ampule of Phosphate Standard Solution, 500-mg/L PO43–
• Ampule breaker
• Pipet, TenSette®, 0.1–1.0 mL and tips
• Mixing cylinders, 10-mL (3)

1. Use the test procedure to measure the concentration of the sample, then keep the
(unspiked) sample in the instrument.
2. Go to the Standard Additions option in the instrument menu.
3. Select the values for standard concentration, sample volume and spike volumes.
4. Open the standard solution.
5. Prepare three spiked samples: use the TenSette pipet to add 0.1 mL, 0.2 mL and
0.3 mL of the standard solution, respectively, to three 10-mL portions of fresh sample.
Mix well.
6. Use the test procedure to measure the concentration of each of the spiked samples.
Start with the smallest sample spike. Measure each of the spiked samples in the
instrument.
7. Select Graph to compare the expected results to the actual results.
Note: If the actual results are significantly different from the expected results, make sure that
the sample volumes and sample spikes are measured accurately. The sample volumes and
sample spikes that are used should agree with the selections in the standard additions menu. If
the results are not within acceptable limits, the sample may contain an interference.

4 Phosphorus, Reactive, Molybdovanadate TNT Method (100.0 mg/L)

Standard solution method
Use the standard solution method to validate the test procedure, reagents and
instrument.
Items to collect:
• 50-mg/L PO43– standard solution

1. Use the test procedure to measure the concentration of the standard solution.
2. Compare the expected result to the actual result.
Note: The factory calibration can be adjusted slightly with the standard adjust option so that the
instrument shows the expected value of the standard solution. The adjusted calibration is then
used for all test results. This adjustment can increase the test accuracy when there are slight
variations in the reagents or instruments.

Method performance
The method performance data that follows was derived from laboratory tests that were
measured on a spectrophotometer during ideal test conditions. Users may get different
results under different test conditions.
Program Standard Precision (95% Confidence Interval) Sensitivity
Concentration change per 0.010 Abs change
540 50.0 mg/L PO43– 49.1–50.9 mg/L PO43– 0.7 mg/L PO43–

Summary of method
Orthophosphate reacts with molybdate in an acid medium to produce a mixed
phosphate/molybdate complex. In the presence of vanadium, yellow
molybdovanadophosphoric acid forms. The intensity of the yellow color is proportional to
the phosphate concentration. Test results are measured at 420 nm.
Pollution prevention and waste management
Reacted samples contain molybdenum and must be disposed of as a hazardous waste.
Dispose of reacted solutions according to local, state and federal regulations.
Consumables and replacement items
Required reagents

Description Quantity/test Unit Item no.

High Range Reactive Phosphorus Test ’N Tube™ Reagent Set — 50 vials 2767345
Includes:
Reactive High Range Phosphorus Test ’N Tube Vials (not sold
1 50/pkg —
separately)
Water, deionized varies 100 mL 27242

Required apparatus

Description Quantity/test Unit Item no.

®
Pipet, TenSette , 1.0- to 10.0-mL 1 each 1970010
Pipet Tips, for TenSette Pipet 1970010 varies 50/pkg 2199796
Test tube rack 1 each 1864100

Phosphorus, Reactive, Molybdovanadate TNT Method (100.0 mg/L) 5

Recommended standards and apparatus

Description Unit Item no.

Phosphate Standard Solution, 50-mg/L as PO4 500 mL 17149

Phosphate Standard Solution, 10-mL Ampule, 500 mg/L as PO4 16/pkg 1424210
Wastewater Influent Standard, Mixed Parameter, for NH3-N, NO3-N, PO4, COD, SO4,
500 mL 2833149
TOC
®
Ampule Breaker, Voluette ampules each 2196800

Optional reagents and apparatus

Description Unit Item no.

Silica Standard Solution, 1-mg/L SiO2 500 mL 110649

Bromine Water, 30 g/L 29 mL 221120
Cylinder, mixing, 25-mL each 189640
Hydrochloric Acid, 6.0 N 1:1, 50% 500 mL 88449
Phenol Solution, 30-g/L 29 mL 211220
Dropper, LDPE, 0.5 –1.0 mL 20/pkg 2124720
Pipet tips for TenSette Pipet 1970010 250/pkg 2199725
Paper, pH, 0–14 pH range 100/pkg 2601300
Thermometer, non-mercury, -10 to +225 °C each 2635700
Bottle, sampling, with cap, low density polyethylene, 250-mL 12/pkg 2087076
Filter paper, folded, 12.5-cm 100/pkg 69257
Funnel, poly, 65-mm each 108367
Beaker, 50-mL each 50041H
Finger cots 2/pkg 1464702

Optional standards

Description Unit Item no.

Phosphate Standard Solution, 3-mg/L as PO4 946 mL 2059716

Phosphate Standard Solution, 10-mg/L as PO4 946 mL 1420416
Phosphate Standard Solution, 15-mg/L as PO4 100 mL 1424342
Phosphate Standard Solution, 30-mg/L as PO4 946 mL 1436716
®
Phosphate Standard Solution, 50-mg/L, 10-mL Voluette Ampules 16/pkg 17110
Phosphate Standard Solution, 100-mg/L as PO4 100 mL 1436832
Phosphate Standard Solution, 10-mL Ampule, 500 mg/L as PO4 16/pkg 1424210
Phosphate Standard Solution, 500-mg/L as PO4 100 mL 1424232

FOR TECHNICAL ASSISTANCE, PRICE INFORMATION AND ORDERING: HACH COMPANY

In the U.S.A. – Call toll-free 800-227-4224 WORLD HEADQUARTERS
Outside the U.S.A. – Contact the HACH office or distributor serving you. Telephone: (970) 669-3050
On the Worldwide Web – www.hach.com; E-mail – [email protected] FAX: (970) 669-2932

© Hach Company/Hach Lange GmbH, 1989–2013. All rights reserved. 04/2013, Edition 8
Phosphorus, Reactive DOC316.53.01119

(Orthophosphate)
®
USEPA1 PhosVer 3 (Ascorbic Acid) Method2 Method 8048
®
0.02 to 2.50 mg/L PO43– Powder Pillows or AccuVac Ampuls
Scope and application: For water, wastewater and seawater.
1 USEPA Accepted for reporting for wastewater analyses. Procedure is equivalent to USEPA and Standard Method 4500-P-E for
wastewater.
2 Adapted from Standard Methods for the Examination of Water and Wastewater.

Test preparation

Instrument-specific information
The tables in this section show all of the instruments that have the program for this test.
Table 1 shows sample cell and orientation requirements for reagent addition tests, such
as powder pillow or bulk reagent tests. Table 2 shows sample cell and adapter
requirements for AccuVac Ampul tests.
To use either table, select an instrument, then read across to find the corresponding
information for this test.
Table 1 Instrument-specific information for powder pillows
Instrument Sample cell orientation Sample cell
DR 6000 The fill line is to the right. 2495402
DR 3800
DR 2800
DR 2700
DR 5000 The fill line is toward the user.
DR 3900
DR 900 The orientation mark is toward the user. 2401906

Table 2 Instrument-specific information for AccuVac Ampuls

Instrument Adapter Sample cell
DR 6000 — 2427606
DR 5000
DR 900
DR 3900 LZV846 (A)
DR 3800 LZV584 (C) 2122800
DR 2800
DR 2700

1
Before starting
Install the instrument cap on the DR 900 cell holder before ZERO or READ is pushed.
For best results, measure the reagent blank value for each new lot of reagent. Replace the sample with deionized water in
the test procedure to get the reagent blank value. Subtract the reagent blank value from the sample results automatically
with the reagent blank adjust option.
Review the Safety Data Sheets (MSDS/SDS) for the chemicals that are used and use any recommended personal protective
equipment.
Dispose of reacted solutions according to local, state and federal regulations. Use the Safety Data Sheets for disposal
information for unused reagents. Consult the environmental, health and safety staff for your facility and/or local regulatory
agencies for further disposal information.

Items to collect
Powder pillows

Description Quantity
®
PhosVer 3 Phosphate Reagent powder pillow, 10-mL 1
Sample cells. (For information about sample cells, adapters or light shields, refer to Instrument-
2
specific information on page 1.)

Refer to Consumables and replacement items on page 6 for reorder information.

AccuVac Ampuls

Description Quantity
® ®
PhosVer 3 Phosphate Reagent AccuVac Ampul 1
Beaker, 50-mL 1
Sample cells (For information about sample cells, adapters or light shields, refer to Instrument-
1
specific information on page 1.)
Stopper for 18-mm tubes and AccuVac Ampuls 1

Refer to Consumables and replacement items on page 6 for reorder information.

Sample collection and storage

• Collect samples in clean glass or plastic bottles that have been cleaned with
1:1 hydrochloric acid and rinsed with deionized water.
• Do not use a detergent that contains phosphate to clean the sample bottles. The
phosphate in the detergent will contaminate the sample.
• Analyze the samples as soon as possible for best results.
• If prompt analysis is not possible, immediately filter and keep the samples at or below
6 °C (43 °F) for a maximum of 48 hours.
• Let the sample temperature increase to room temperature before analysis.

2 Phosphorus, Reactive, PhosVer 3 Method (2.50 mg/L)

Powder pillow procedure

Start

1. Start program 490 P 2. Prepare the sample: Fill 3. Add the contents of one 4. Immediately close the
React. PP. For information a sample cell with 10 mL of PhosVer 3 Phosphate sample cell. Shake
about sample cells, sample. Reagent Powder Pillow to vigorously for 30 seconds.
adapters or light shields, the cell.
refer to Instrument-specific
information on page 1.
Note: Although the program
name may vary between
instruments, the program
number does not change.

5. Start the instrument 6. Prepare the blank: Fill a 7. When the timer expires, 8. Insert the blank into the
timer. A 2-minute reaction second sample cell with clean the blank. cell holder.
time starts. 10 mL of sample.
If the sample was digested
using the Acid Persulfate
digestion, a 10-minute
reaction period is
necessary.

Zero Read

9. Push ZERO. The display 10. Clean the prepared 11. Insert the prepared 12. Push READ. Results
shows 0.00 mg/L PO43–. sample. sample into the cell holder. show in mg/L PO43–.

Phosphorus, Reactive, PhosVer 3 Method (2.50 mg/L) 3

AccuVac Ampul procedure

Start

1. Start program 492 P 2. Prepare the blank: Fill 3. Prepare the sample: 4. Close the AccuVac
React. PV AV. For the sample cell with 10 mL Collect at least 40 mL of Ampul. Shake for
information about sample of sample. sample in a 50-mL beaker. approximately 30 seconds.
cells, adapters or light Fill the AccuVac Ampul with Accuracy is not affected by
shields, refer to Instrument- sample. Keep the tip undissolved powder.
specific information immersed while the Ampul
on page 1. fills completely.
Note: Although the program
name may vary between
instruments, the program
number does not change.

Zero

5. Start the instrument 6. When the timer expires, 7. Insert the blank into the 8. Push ZERO. The display
timer. A 2-minute reaction clean the blank. cell holder. shows 0.00 mg/L PO43–.
time starts.
If the sample was digested
using the Acid Persulfate
digestion, a 10-minute
reaction period is
necessary.

Read

9. Clean the AccuVac 10. Insert the prepared 11. Push READ. Results
Ampul. sample AccuVac Ampul into show in mg/L PO43–.
the cell holder.

4 Phosphorus, Reactive, PhosVer 3 Method (2.50 mg/L)

Interferences
Interfering substance Interference level
Aluminum More than 200 mg/L
Arsenate Interferes at any level.
Chromium More than 100 mg/L
Copper More than 10 mg/L
Hydrogen Sulfide Interferes at any level.
Iron More than 100 mg/L
Nickel More than 300 mg/L
Highly buffered samples or Can prevent the correct pH adjustment of the sample by the reagents. Sample pretreatment
extreme sample pH may be necessary. A pH range of 2–10 is recommended.
Silica More than 50 mg/L
Silicate More than 10 mg/L
Turbidity or color May cause inconsistent results. The acid in the powder pillow can dissolve some of the
suspended particles and the desorption of orthophosphate from the particles can vary. For
highly turbid or colored samples, add the contents of one Phosphate Pretreatment Powder
Pillow to 25 mL of sample. Mix well. Use this solution to zero the instrument.
Zinc More than 80 mg/L

Accuracy check
Standard additions method (sample spike)
Use the standard additions method (for applicable instruments) to validate the test
procedure, reagents and instrument and to find if there is an interference in the sample.
Items to collect:
• Phosphate standard solution, 50 mg/L PO43– ampule
• Ampule breaker
• Pipet, TenSette®, 0.1–1.0 mL and tips
• Mixing cylinders, 25-mL (3)

1. Use the test procedure to measure the concentration of the sample, then keep the
(unspiked) sample in the instrument.
2. Go to the Standard Additions option in the instrument menu.
3. Select the values for standard concentration, sample volume and spike volumes.
4. Open the standard solution.
5. Prepare three spiked samples: use the TenSette pipet to add 0.1 mL, 0.2 mL and
0.3 mL of the standard solution, respectively, to three 10-mL portions of fresh sample.
Mix well.
®
Note: For AccuVac Ampuls, add 0.2 mL, 0.4 mL and 0.6 mL of the standard solution to three
50-mL portions of fresh sample.
6. Use the test procedure to measure the concentration of each of the spiked samples.
Start with the smallest sample spike. Measure each of the spiked samples in the
instrument.
7. Select Graph to compare the expected results to the actual results.
Note: If the actual results are significantly different from the expected results, make sure that
the sample volumes and sample spikes are measured accurately. The sample volumes and
sample spikes that are used should agree with the selections in the standard additions menu. If
the results are not within acceptable limits, the sample may contain an interference.

Phosphorus, Reactive, PhosVer 3 Method (2.50 mg/L) 5

Standard solution method
Use the standard solution method to validate the test procedure, reagents and
instrument.
Items to collect:
• 50 mg/L phosphate standard solution
• 100-mL volumetric flask, Class A
• 4-mL volumetric pipet, Class A and pipet filler
• Deionized water

1. Prepare a 2.00 mg/L phosphate standard solution as follows:

a. Use a pipet to add 4.00 mL of 50 mg/L phosphate standard solution into the
volumetric flask. (Alternately, use one of the available mixed parameter
standards. These standards contain 2.0 mg/L phosphate.)
b. Dilute to the mark with deionized water. Mix well. Prepare this solution daily.
2. Use the test procedure to measure the concentration of the prepared standard
solution.
3. Compare the expected result to the actual result.
Note: The factory calibration can be adjusted slightly with the standard adjust option so that the
instrument shows the expected value of the standard solution. The adjusted calibration is then
used for all test results. This adjustment can increase the test accuracy when there are slight
variations in the reagents or instruments.

Method performance
The method performance data that follows was derived from laboratory tests that were
measured on a spectrophotometer during ideal test conditions. Users may get different
results under different test conditions.
Program Standard Precision (95% Confidence Interval) Sensitivity
Concentration change per 0.010 Abs change
490 2.00 mg/L PO43– 1.98–2.02 mg/L PO43– 0.02 mg/L PO43–
492 2.00 mg/L PO43– 1.98–2.02 mg/L PO43– 0.02 mg/L PO43–

Summary of method
Orthophosphate reacts with molybdate in an acid medium to produce a mixed
phosphate/molybdate complex. Ascorbic acid then reduces the complex, giving an
intense molybdenum blue color. The measurement wavelength is 880 nm for
spectrophotometers or 610 nm for colorimeters.
Consumables and replacement items
Required reagents

Description Quantity/Test Unit Item no.

®
PhosVer 3 Phosphate Reagent Powder Pillow, 10-mL 1 100/pkg 2106069
OR
® ®
PhosVer 3 Phosphate Reagent AccuVac Ampul 1 25/pkg 2508025

Required apparatus

Description Quantity/Test Unit Item no.

Beaker, 50-mL 1 each 50041H

Stoppers for 18-mm tubes and AccuVac Ampuls 2 6/pkg 173106

6 Phosphorus, Reactive, PhosVer 3 Method (2.50 mg/L)

Recommended standards

Description Unit Item no.

®
Phosphate Standard Solution, 10-mL Voluette Ampul, 50-mg/L as PO4 16/pkg 17110
Phosphate Standard Solution, 50-mg/L as PO4 500 mL 17149
Phosphate Standard Solution, 1-mg/L as PO4 500 mL 256949
Drinking Water Standard, Mixed Parameter, Inorganic for F-, NO3, PO4, SO4 500 mL 2833049
Wastewater, Effluent Inorganics, for NH3-N, NO3-N, PO4, COD, SO4, TOC 500 mL 2833249
Water, deionized 4L 27256

Optional reagents and apparatus

Description Unit Item no.

®
AccuVac Drainer each 4103600
®
AccuVac Snapper each 2405200
®
AccuVac vials for sample blanks 25/pkg 2677925
®
Ampule Breaker, Voluette ampules each 2196800
Bottle, sampling, with cap, low density polyethylene, 250-mL 12/pkg 2087076
Cylinder, mixing, 50-mL each 189641
Flask, volumetric, Class A, 100-mL each 1457442
Hydrochloric Acid, 6.0 N 1:1, 50% 500 mL 88449
Paper, pH, 0–14 pH range 100/pkg 2601300
Phosphate Treatment Powder Pillow 100/pkg 1450199
Phosphate Standard Solution, 10-mg/L as PO4 946 mL 1420416
Phosphate Standard Solution, 15-mg/L as PO4 100 mL 1424342
Phosphate Standard Solution, 100-mg/L as PO4 100 mL 1436832
Phosphate Standard Solution, 10-mL Ampule, 500 mg/L as PO4 16/pkg 1424210
Phosphate Standard Solution, 500-mg/L as PO4 100 mL 1424232
®
Pipet, TenSette , 0.1–1.0 mL each 1970001
®
Pipet, TenSette , 1.0 to 10.0 mL each 1970010
Pipet tips for TenSette Pipet 1970001 50/pkg 2185696
Pipet tips for TenSette Pipet 1970001 1000/pkg 2185628
Pipet tips for TenSette Pipet 1970010 50/pkg 2199796
Pipet tips for TenSette Pipet 1970010 250/pkg 2199725
Pipet, volumetric, Class A, 4.00-mL each 1451504

Phosphorus, Reactive, PhosVer 3 Method (2.50 mg/L) 7

 FOR TECHNICAL ASSISTANCE, PRICE INFORMATION AND ORDERING: HACH COMPANY
In the U.S.A. – Call toll-free 800-227-4224 WORLD HEADQUARTERS
Outside the U.S.A. – Contact the HACH office or distributor serving you. Telephone: (970) 669-3050
On the Worldwide Web – www.hach.com; E-mail – [email protected] FAX: (970) 669-2932

© Hach Company/Hach Lange GmbH, 1989–2013. All rights reserved. 04/2013, Edition 8
Phosphorus, Reactive DOC316.53.01118

(Orthophosphate)
USEPA PhosVer® 3 Method1 Method 8048
0.06 to 5.00 mg/L PO43– (0.02 to 1.60 mg/L P) Test ‘N Tube™ Vials
Scope and application: For water, wastewater and seawater.
1 USEPA accepted for reporting wastewater analysis. Procedure is equivalent to USEPA and Standard Method 4500-P E for wastewater.

Test preparation

Instrument specific information

The table in this section shows all of the instruments that have the program for this test.
Table 1 shows adapter and light shield requirements for the instruments that use them.
To use the table, select an instrument, then read across to find the corresponding
information for this test.
Table 1 Instrument-specific information for test tubes
Instrument Adapters Light shield
DR 6000 — —
DR 5000 — —
DR 900 4846400 Cover supplied with the instrument
DR 3900 — LZV849
DR 3800 — LZV646
DR 2800 —

Before starting
Install the instrument cap on the DR 900 cell holder before ZERO or READ is pushed.
DR 3900, DR 3800, DR 2800 and DR 2700: Install the light shield in Cell Compartment #2 before this test is started.
For best results, measure the reagent blank value for each new lot of reagent. Replace the sample with deionized water in
the test procedure to get the reagent blank value. Subtract the reagent blank value from the sample results automatically
with the reagent blank adjust option.
Review the Safety Data Sheets (MSDS/SDS) for the chemicals that are used and use any recommended personal protective
equipment.
Dispose of reacted solutions according to local, state and federal regulations. Use the Safety Data Sheets for disposal
information for unused reagents. Consult the environmental, health and safety staff for your facility and/or local regulatory
agencies for further disposal information.

Items to collect
Description Quantity
®
PhosVer 3 Reagent Powder Pillow 1
Reactive Phosphorus Test 'N Tube Vial 1
Funnel, micro 1
Light shield or adapter (For information about sample cells, adapters or light shields, refer to
1
Instrument specific information on page 1.)

1
Items to collect (continued)
Description Quantity
®
Pipet, TenSette , 1.0- to 10.0-mL, with pipet tips 1
Test tube rack 1

Refer to Consumables and replacement items on page 4 for reorder information.

Sample collection and storage

• Collect samples in clean glass or plastic bottles that have been cleaned with
1:1 hydrochloric acid and rinsed with deionized water.
• Do not use a detergent that contains phosphate to clean the sample bottles. The
phosphate in the detergent will contaminate the sample.
• Analyze the samples as soon as possible for best results.
• If prompt analysis is not possible, immediately filter and keep the samples at or below
6 °C (43 °F) for a maximum of 48 hours.
• Let the sample temperature increase to room temperature before analysis.

Test 'N Tube procedure

Start

1. Start program 535 P 2. Add 5.0 mL of sample to 3. Put the cap on the vial. 4. Clean the vial.
React. PV TNT. For a Reactive Phosphorus Test Invert to mix.
information about sample 'N Tube Vial.
cells, adapters or light
shields, refer to Instrument
specific information
on page 1.
Note: Although the program
name may vary between
instruments, the program
number does not change.

Zero

5. Insert the vial into the 16- 6. Push ZERO. The display 7. Add the contents of one 8. Put the cap on the vial.
mm cell holder. shows 0.00 mg/L PO43–. PhosVer 3 Phosphate Shake for at least
Powder Pillow. 20 seconds. The powder will
not dissolve completely.

2 Phosphorus, Reactive, PhosVer 3 TNT Method (5.00 mg/L)

 Read

9. Start the instrument 10. When the timer expires, 11. Insert the vial into the 12. Push READ. Results
timer. A 2-minute reaction clean the vial. 16-mm cell holder. show in mg/L PO43–.
time starts.
Measure the sample
between two and eight
minutes after adding the
PhosVer 3 reagent.

Interferences
Interfering substance Interference level
Aluminum More than 200 mg/L
Arsenate Interferes at any level.
Chromium More than 100 mg/L
Copper More than 10 mg/L
Sulfide More than 6 mg/L. Remove sulfide interference as follows:

1. Measure 25 mL of sample into a 50-mL beaker.

2. Swirl continuously and add bromine water by drops until a permanent yellow color is seen.
3. Swirl continuously and add phenol solution by drops only until the yellow color is removed.
Use this treated sample in the test procedure.

Iron More than 100 mg/L

Nickel More than 300 mg/L
Highly buffered samples Can prevent the correct pH adjustment of the sample by the reagents. Sample pretreatment may
or extreme sample pH be necessary.
Silica More than 50 mg/L
Silicate More than 10 mg/L
Turbidity or color May cause inconsistent results. The acid in the powder pillow can dissolve some of the
suspended particles and the desorption of orthophosphate from the particles can vary. For highly
turbid or colored samples, add the contents of one Phosphate Pretreatment Powder Pillow to
25 mL of sample. Mix well. Use this solution to zero the instrument.
Zinc More than 80 mg/L

Accuracy check
Standard additions method (sample spike)
Use the standard additions method (for applicable instruments) to validate the test
procedure, reagents and instrument and to find if there is an interference in the sample.
Items to collect:
• Phosphate 2-mL Ampule Standard, 50-mg/L as PO43–
• Ampule breaker
• Pipet, TenSette®, 0.1–1.0 mL and tips

Phosphorus, Reactive, PhosVer 3 TNT Method (5.00 mg/L) 3

 • Mixing cylinders, 25-mL (3)

1. Use the test procedure to measure the concentration of the sample, then keep the
(unspiked) sample in the instrument.
2. Go to the Standard Additions option in the instrument menu.
3. Select the values for standard concentration, sample volume and spike volumes.
4. Open the standard solution.
5. Prepare three spiked samples: use the TenSette pipet to add 0.1 mL, 0.2 mL and
0.3 mL of the standard solution, respectively, to three 25-mL portions of fresh sample.
Mix well.
6. Use the test procedure to measure the concentration of each of the spiked samples.
Start with the smallest sample spike. Measure each of the spiked samples in the
instrument.
7. Select Graph to compare the expected results to the actual results.
Note: If the actual results are significantly different from the expected results, make sure that
the sample volumes and sample spikes are measured accurately. The sample volumes and
sample spikes that are used should agree with the selections in the standard additions menu. If
the results are not within acceptable limits, the sample may contain an interference.

Standard solution method

Use the standard solution method to validate the test procedure, reagents and
instrument.
Items to collect:
• 3.0-mg/L phosphate standard solution

1. Use the test procedure to measure the concentration of the standard solution.
2. Compare the expected result to the actual result.
Note: The factory calibration can be adjusted slightly with the standard adjust option so that the
instrument shows the expected value of the standard solution. The adjusted calibration is then
used for all test results. This adjustment can increase the test accuracy when there are slight
variations in the reagents or instruments.

Method performance
The method performance data that follows was derived from laboratory tests that were
measured on a spectrophotometer during ideal test conditions. Users may get different
results under different test conditions.
Program Standard Precision (95% Confidence Interval) Sensitivity
Concentration change per 0.010 Abs change
535 3.00 mg/L PO43– 2.94–3.06 mg/L PO43– 0.06 mg/L PO43–

Summary of method
Orthophosphate reacts with molybdate in an acid medium to produce a mixed
phosphate/molybdate complex. Ascorbic acid then reduces the complex, giving an
intense molybdenum blue color. The measurement wavelength is 880 nm for
spectrophotometers or 610 nm for colorimeters.
Consumables and replacement items
Required reagents

Description Quantity/test Unit Item no.

Reactive Phosphorus Test ’N Tube™ Reagent Set — 50 tests 2742545

Includes:
®
PhosVer 3 Phosphate Reagent Powder Pillow, 10-mL 1 50/pkg 2106046
Reactive Phosphorus Test ‘N Tube Dilution Vials (not sold separately) 1 50/pkg —

4 Phosphorus, Reactive, PhosVer 3 TNT Method (5.00 mg/L)

Required apparatus

Description Quantity/test Unit Item no.

Funnel, micro, poly 1 each 2584335

®
Pipet, TenSette , 1.0- to 10.0-mL 1 each 1970010
Pipet Tips, for TenSette Pipet 1970010 varies 50/pkg 2199796
Test tube rack 1 each 1864100

Recommended standards and apparatus

Description Unit Item no.

®
Phosphate Standard Solution, PourRite Ampule, 50-mg/L as PO43–, 2-mL 20/pkg 17120H
Phosphate Standard Solution, 50-mg/L as PO4 500 mL 17149
Phosphate Standard Solution, 1-mg/L as PO4 500 mL 256949
Phosphate Standard Solution, 3-mg/L as PO4 946 mL 2059716
Drinking Water Standard, Mixed Parameter, Inorganic for F-, NO3, PO4, SO4 500 mL 2833049
Wastewater, Effluent Inorganics, for NH3-N, NO3-N, PO4, COD, SO4, TOC 500 mL 2833249
®
Ampule Breaker, PourRite ampules each 2484600

Optional reagents and apparatus

Description Unit Item no.

Bromine Water, 30-g/L 29 mL 221120

Hydrochloric Acid, 6.0 N 1:1, 50% 500 mL 88449
Phenol Solution, 30-g/L 29 mL 211220
®
Pipet, TenSette , 0.1–1.0 mL each 1970001
Pipet tips for TenSette Pipet 1970001 50/pkg 2185696
Pipet tips for TenSette Pipet 1970001 1000/pkg 2185628
Filter paper, folded, 12.5-cm 100/pkg 69257
Funnel, poly, 65-mm each 108367
Thermometer, non-mercury, -10 to +225 °C each 2635700
Bottle, sampling, with cap, low density polyethylene, 250-mL 12/pkg 2087076
Beaker, 50-mL each 50041H

Optional standards

Description Unit Item no.

®
Ampule Breaker, Voluette ampules each 2196800
Phosphate Standard Solution, 10-mg/L as PO4 946 mL 1420416
Phosphate Standard Solution, 15-mg/L as PO4 100 mL 1424342
®
Phosphate Standard Solution, 50-mg/L, 10-mL Voluette Ampules 16/pkg 17110
Phosphate Standard Solution, 100-mg/L as PO4 100 mL 1436832
Phosphate Standard Solution, 10-mL Ampule, 500 mg/L as PO4 16/pkg 1424210
Phosphate Standard Solution, 500-mg/L as PO4 100 mL 1424232

Phosphorus, Reactive, PhosVer 3 TNT Method (5.00 mg/L) 5

 FOR TECHNICAL ASSISTANCE, PRICE INFORMATION AND ORDERING: HACH COMPANY
In the U.S.A. – Call toll-free 800-227-4224 WORLD HEADQUARTERS
Outside the U.S.A. – Contact the HACH office or distributor serving you. Telephone: (970) 669-3050
On the Worldwide Web – www.hach.com; E-mail – [email protected] FAX: (970) 669-2932

© Hach Company/Hach Lange GmbH, 1989–2013. All rights reserved. 04/2013, Edition 8
Phosphorus, Total, Digestion DOC316.53.01112

USEPA1 Acid Persulfate Digestion Method2 Method 8190

Scope and application: For water, wastewater and seawater.

1 USEPA Accepted for wastewater analyses when used with the ascorbic acid (PhosVer 3) method.
2 Adapted from Standard Methods for the Examination of Water and Wastewater 4500-P B & E.

Test preparation

Before starting
Clean all glassware with 6.0 N (50%) hydrochloric acid, then rinse thoroughly with deionized water to remove contaminants.
Review the Safety Data Sheets (MSDS/SDS) for the chemicals that are used and use any recommended personal protective
equipment.
Dispose of reacted solutions according to local, state and federal regulations. Use the Safety Data Sheets for disposal
information for unused reagents. Consult the environmental, health and safety staff for your facility and/or local regulatory
agencies for further disposal information.

Items to collect
Description Quantity

Potassium Persulfate Powder Pillow 1

Sodium Hydroxide Solution, 5.0 N 2 mL
Sulfuric Acid Solution, 5.25 N 2 mL
Water, deionized varies
Cylinder, graduated, 25-mL 1
Flask, Erlenmeyer, 125-mL 1
Hot plate 1

Refer to Consumables and replacement items on page 3 for reorder information.

Sample collection and storage

• Collect samples in clean glass or plastic bottles.
• Analyze the samples as soon as possible for best results.
• To preserve samples for later analysis, adjust the sample pH to less than 2 with
concentrated sulfuric acid (about 2 mL per liter). No acid addition is necessary if the
sample is tested immediately.
• Keep the preserved samples at or below 6 °C (43 °F) for up to 28 days.
• Let the sample temperature increase to room temperature before analysis.
• Before analysis, adjust the pH to 7 with 5.0 N sodium hydroxide standard solution.
• Correct the test result for the dilution from the volume additions.

1
Acid persulfate digestion procedure

1. Use a graduated cylinder 2. Add the contents of one 3. Swirl to mix. 4. Add 2.0 mL of 5.25 N
to add 25-mL of sample into Potassium Persulfate Sulfuric Acid Solution to the
the 125-mL Erlenmeyer Powder Pillow. flask.
flask.

5. Boil the sample gently for 6. Let the sample cool to 7. Swirl to mix. 8. Pour the sample into a
30 minutes. Do not let the room temperature. Add 25-mL graduated cylinder.
flask boil dry. 2.0 mL of 5.0 N Sodium Adjust the volume to 25 mL
For the best recovery, Hydroxide Solution to the by rinsing the flask with
concentrate the sample to flask. deionized water and pouring
less than 20 mL. After the rinse water into the
concentration, maintain the cylinder.
volume of sample near
20 mL by adding small
amounts of deionized water.
Do not exceed 20 mL.

2 Phosphorus, Total, Acid Persulfate Digestion

 Start

9. Proceed with a reactive

phosphorous test of the
expected acid hydrolyzable
phosphorous concentration
range.

• 480 P React. Mo
• 482 P React. Mo AV
• 485 P React. Amino
• 490 P React. PV
• 492 P React. PV AV
• 535 P React. PV TNT
• 540 P React. HT TNT

Extend the color

development time to
10 minutes for the PhosVer
3 method.

Interferences
Interfering substance Interference level
Alkaline or highly If the pH of the sample after the acid is added is not below pH 1, add additional acid.
buffered samples
Turbidity Use 50 mL of sample and double the reagent quantities. Use a portion of the digested sample to
zero the instrument in the reactive phosphorus procedure. This compensates for any color or
turbidity destroyed by this procedure.

Summary of method
Phosphates in organic and condensed inorganic forms (meta-, pyro- or other
polyphosphates) must be converted to reactive orthophosphate before analysis.
Pretreatment of the sample with acid and heat provides the conditions for hydrolysis of
the condensed inorganic forms. Organic phosphates are converted to orthophosphate by
heating with acid and persulfate. Organically bound phosphates are thus determined
indirectly by subtracting the result of an acid hydrolyzable phosphorus test from the total
phosphorus result. This procedure must be followed by one of the reactive phosphorus
(orthophosphate) analysis methods for determining the phosphorus content of the
sample. If the ascorbic acid (PhosVer 3) method is used to measure the reactive
phosphorus, this method is USEPA accepted for NPDES reporting.
Consumables and replacement items
Required reagents

Description Quantity/test Unit Item no.

Potassium Persulfate Powder Pillows 1 100/pkg 245199

100 mL
Sodium Hydroxide Solution, 5.0 N 2 mL 245032
MDB

Phosphorus, Total, Acid Persulfate Digestion 3

Consumables and replacement items (continued)
Description Quantity/test Unit Item no.

100 mL
Sulfuric Acid Solution, 5.25 N 2 mL 244932
MDB
Water, deionized varies 4L 27256

Required apparatus

Description Quantity/test Unit Item no.

Cylinder, graduated, 25-mL. 1 each 50840

Flask, Erlenmeyer, 125-mL 2 each 50543
Hot plate, 7 inches x 7 inches, digital, 120 VAC 1 each 2881500
Hot plate, stirrer, 220 - 240 VAC 1 each 2881602

Optional reagents and apparatus

Description Unit Item no.

Sodium Hydroxide Standard Solution, 5.0 N 1L 245053

Sulfuric Acid, concentrated, ACS 500 mL 97949
Paper, pH, 0–14 pH range 100/pkg 2601300
Thermometer, non-mercury, -10 to +225 °C each 2635700
Hydrochloric Acid, 6.0 N 1:1, 50% 500 mL 88449
Bottle, sampling, with cap, low density polyethylene, 250-mL 12/pkg 2087076

FOR TECHNICAL ASSISTANCE, PRICE INFORMATION AND ORDERING: HACH COMPANY

In the U.S.A. – Call toll-free 800-227-4224 WORLD HEADQUARTERS
Outside the U.S.A. – Contact the HACH office or distributor serving you. Telephone: (970) 669-3050
On the Worldwide Web – www.hach.com; E-mail – [email protected] FAX: (970) 669-2932

© Hach Company/Hach Lange GmbH, 1989–2013. All rights reserved. 04/2013, Edition 8
Phosphorus, Total DOC316.53.01123

Molybdovanadate with Acid Persulfate Digestion Method1 Method 10127

1.0 to 100.0 mg/L PO43– (HR) Test ‘N Tube™ Vials
Scope and application: For water and wastewater.
1 Adapted from Standard Methods for the Examination of Water and Wastewater, (4500 B-C).

Test preparation

Instrument specific information

The table in this section shows all of the instruments that have the program for this test.
Table 1 shows adapter and light shield requirements for the instruments that use them.
To use the table, select an instrument, then read across to find the corresponding
information for this test.
Table 1 Instrument-specific information for test tubes
Instrument Adapters Light shield
DR 6000 — —
DR 5000 — —
DR 900 4846400 Cover supplied with the instrument
DR 3900 — LZV849
DR 3800 — LZV646
DR 2800 —

Before starting
Install the instrument cap on the DR 900 cell holder before ZERO or READ is pushed.
DR 3900, DR 3800, DR 2800 and DR 2700: Install the light shield in Cell Compartment #2 before this test is started.
Reagent blanks can be used more than once, but should not be used more than one day.
The reagent that is used in this test is corrosive. Use protection for eyes and skin and be prepared to flush any spills with
running water.
Review the Safety Data Sheets (MSDS/SDS) for the chemicals that are used and use any recommended personal protective
equipment.
Dispose of reacted solutions according to local, state and federal regulations. Use the Safety Data Sheets for disposal
information for unused reagents. Consult the environmental, health and safety staff for your facility and/or local regulatory
agencies for further disposal information.

Items to collect
Description Quantity

Total High Range Phosphorus Test 'N Tube Reagent Set 1

DRB200 Reactor 1
Funnel, micro 1
Light shield or adapter (For information about sample cells, adapters or light shields, refer to
1
Items to collect on page 1.)
®
Pipet, TenSette , 1.0- to 10.0-mL, with pipet tips 1

1
Items to collect (continued)
Description Quantity

Test tube rack 1

Water, deionized varies

Refer to Consumables and replacement items on page 6 for reorder information.

Sample collection and storage

• Collect samples in clean glass or plastic bottles that have been cleaned with
1:1 hydrochloric acid and rinsed with deionized water.
• Analyze the samples as soon as possible for best results.
• Do not use a detergent that contains phosphate to clean the sample bottles. The
phosphate in the detergent will contaminate the sample.
• To preserve samples for later analysis, adjust the sample pH to less than 2 with
concentrated sulfuric acid (about 2 mL per liter). No acid addition is necessary if the
sample is tested immediately.
• Keep the preserved samples at or below 6 °C (43 °F) for up to 28 days.
• Let the sample temperature increase to room temperature before analysis.
• Before analysis, adjust the pH to 7 with 5.0 N sodium hydroxide standard solution.
• Correct the test result for the dilution from the volume additions.

Molybdovanadate method, acid persulfate digestion

Start

1. Start the 2. Start program 542 P 3. Prepare the blank: Add 4. Prepare the sample:
DRB200 Reactor. Preheat Total HR TNT. For 5.0 mL of deionized water to Add 5.0 mL of sample to a
to 150 °C. Refer to the information about sample a Total Phosphorus Test Total Phosphorus Test Vial.
DRB200 manual. cells, adapters or light Vial.
shields, refer to Items to
collect on page 1.
Note: Although the program
name may vary between
instruments, the program
number does not change.

2 Phosphorus, Total, Molybdovanadate TNT Method (100.0 mg/L)

5. Add the contents of one 6. Put the cap on the vial. 7. Insert the vial into the 8. Start the instrument
Potassium Persulfate Shake to dissolve the reactor. Close the reactor. timer. A 30-minute reaction
Powder Pillow to each vial. powder. time starts.

9. When the timer expires, 10. Add 2 mL of 1.54 N 11. Put the cap on the vial. 12. Use a polyethylene
carefully remove the hot Sodium Hydroxide Standard Invert to mix. dropper to add 0.5 mL of
vials from the reactor. Set Solution to each vial. Molybdovanadate Reagent
the vials in a test tube rack. to each vial.
Let the vials cool to room
temperature.

13. Put the cap on the vial. 14. Start the instrument 15. Clean the blank vial. 16. Insert the vial into the
Invert to mix. timer. A 7-minute reaction 16-mm cell holder.
time starts.
Measure the sample
between seven and nine
minutes after the addition of
the Molybdovanadate
Reagent.

Phosphorus, Total, Molybdovanadate TNT Method (100.0 mg/L) 3

 Zero Read

17. Push ZERO. The 18. Clean the sample vial. 19. Insert the vial into the 20. Push READ. Results
display shows 0.0 mg/L 16-mm cell holder. show in mg/L PO43–.
PO43–.

Interferences
Table 2 shows the interferences and interference levels. Table 3 shows the substances
that do not interfere at or below the indicated levels.
Table 2 Interfering substances
Interfering substance Interference level
Arsenate Causes a positive interference if the sample is warm when the reagent is added. Let the
sample temperature decrease to room temperature after the digestion.
Iron, ferrous Causes a blue color which interferes at more than 100 mg/L.
Molybdate Causes a negative interference at more than 1000 mg/L.
Silica Causes a positive interference if the sample is warm when the reagent is added. Let the
sample temperature decrease to room temperature after the digestion.
Highly buffered samples or Can prevent the correct pH adjustment of the sample by the reagents. Sample pretreatment
extreme sample pH may be necessary.
Fluoride, thorium, bismuth, Causes a negative interference.
thiosulfate or thiocyanate
Turbidity Sample turbidity can cause inconsistent results because the acid in the reagents can dissolve
some of the suspended particles and because of variable desorption of orthophosphate from
the particles.
Temperature Temperatures below 18 °C (64 °F) cause a negative interference. Temperatures above 25 °C
(77 °F) cause a positive interference. Let the sample temperature decrease to room
temperature after the digestion.

Table 3 Substances that do not interfere at less than 1000 mg/L

Pyrophosphate Tetraborate Selenate Benzoate
Citrate Oxalate Lactate Tartrate
Formate Salicylate Al3+ Fe3+
Mg2+ Ca2+ Ba2+ Sr2+
Li+ Na+ K+ NH4+
Cd2+ Mn2+ NO3– NO2–
SO42– SO32– Pb2+ Hg+
Hg2+ Sn2+ Cu2+ Ni2+
Ag+ U4+ Zr4+ AsO3–
Br– CO32– ClO4– CN–
IO3– SiO44– — —

4 Phosphorus, Total, Molybdovanadate TNT Method (100.0 mg/L)

Accuracy check
Standard additions method (sample spike)
Use the standard additions method (for applicable instruments) to validate the test
procedure, reagents and instrument and to find if there is an interference in the sample.
Items to collect:
• Phosphate 10-mL Ampule Standard, 500-mg/L as PO43–
• Ampule breaker
• Pipet, TenSette®, 0.1–1.0 mL and tips
• Mixing cylinders, 10-mL (3)

1. Use the test procedure to measure the concentration of the sample, then keep the
(unspiked) sample in the instrument.
2. Go to the Standard Additions option in the instrument menu.
3. Select the values for standard concentration, sample volume and spike volumes.
4. Open the standard solution.
5. Prepare three spiked samples: use the TenSette pipet to add 0.1 mL, 0.2 mL and
0.3 mL of the standard solution, respectively, to three 10-mL portions of fresh sample.
Mix well.
6. Use the test procedure to measure the concentration of each of the spiked samples.
Start with the smallest sample spike. Measure each of the spiked samples in the
instrument.
7. Select Graph to compare the expected results to the actual results.
Note: If the actual results are significantly different from the expected results, make sure that
the sample volumes and sample spikes are measured accurately. The sample volumes and
sample spikes that are used should agree with the selections in the standard additions menu. If
the results are not within acceptable limits, the sample may contain an interference.

Standard solution method

Use the standard solution method to validate the test procedure, reagents and
instrument.
Items to collect:
• 50-mg/L phosphate standard solution

1. Use the test procedure to measure the concentration of the standard solution.
2. Compare the expected result to the actual result.
Note: The factory calibration can be adjusted slightly with the standard adjust option so that the
instrument shows the expected value of the standard solution. The adjusted calibration is then
used for all test results. This adjustment can increase the test accuracy when there are slight
variations in the reagents or instruments.

Method performance
The method performance data that follows was derived from laboratory tests that were
measured on a spectrophotometer during ideal test conditions. Users may get different
results under different test conditions.
Program Standard Precision (95% Confidence Interval) Sensitivity
Concentration change per 0.010 Abs change
542 50 mg/L PO43– 49.4–50.6 mg/L PO43– 0.7 mg/L PO43–

Summary of method
Phosphates present in organic and condensed inorganic forms (meta-, pyro- or other
polyphosphates) must be converted to reactive orthophosphate before analysis.
Pretreatment of the sample with acid and heat provides the conditions for hydrolysis of
the condensed inorganic forms. Organic phosphates are converted to orthophosphates
by heating with acid and persulfate. Orthophosphate reacts with molybdate in an acid

Phosphorus, Total, Molybdovanadate TNT Method (100.0 mg/L) 5

 medium to produce a mixed phosphate/molybdate complex. In the presence of vanadium,
yellow molybdovanadophosphoric acid forms. The intensity of the yellow color is
proportional to the phosphate concentration. Test results are measured at 420 nm.
Pollution prevention and waste management
Reacted samples contain molybdenum and must be disposed of as a hazardous waste.
Dispose of reacted solutions according to local, state and federal regulations.
Consumables and replacement items
Required reagents

Description Quantity/test Unit Item no.

Total High Range Phosphorus Test ’N Tube™ Reagent Set — 50 vials 2767245
Includes:
Molybdovanadate Reagent (not sold separately) 0.5 mL 25 mL —
Potassium Persulfate Powder Pillows 1 pillow 50/pkg 2084766
Sodium Hydroxide, 1.54 N varies 100 mL 2743042
Total Phosphorus Test Vials (not sold separately) 1 50/pkg —
Water, deionized varies 100 mL 27242

Required apparatus

Description Quantity/test Unit Item no.

DRB200 Reactor, 110 V, 15 x 16 mm 1 each LTV082.53.40001

DRB200 Reactor, 220 V, 15 x 16 mm 1 each LTV082.52.40001
Dropper, measuring, 0.5 and 1.0 mL plastic 2 20/pkg 2124720
Funnel, micro, poly 1 each 2584335
Light shield, DR 3900 1 each LZV849
Light shield, DR 3800, DR 2800, DR 2700 1 each LZV646
®
Pipet, TenSette , 1.0- to 10.0-mL 1 each 1970010
Pipet Tips, for TenSette Pipet 1970010 2 250/pkg 2199725
Test tube rack 1 each 1864100

Recommended standards and apparatus

Description Unit Item no.

Phosphate Standard Solution, 10-mL Ampule, 500 mg/L as PO4 16/pkg 1424210
Phosphate Standard Solution, 50-mg/L as PO4 500 mL 17149
Wastewater Influent Standard, Mixed Parameter, for NH3-N, NO3-N, PO4, COD, SO4,
500 mL 2833149
TOC
®
Ampule Breaker, Voluette ampules each 2196800

Optional reagents and apparatus

Description Unit Item no.

Hydrochloric Acid, 6.0 N 1:1, 50% 500 mL 88449

Sodium Hydroxide Standard Solution, 5.0 N 1L 245053
Sulfuric Acid, concentrated, ACS 500 mL 97949

6 Phosphorus, Total, Molybdovanadate TNT Method (100.0 mg/L)

Consumables and replacement items (continued)
Description Unit Item no.

Molybdovanadate Reagent 100 mL 2076032

Pipet tips for TenSette Pipet 1970010 50/pkg 2199796
®
Pipet, TenSette , 0.1–1.0 mL each 1970001
Pipet tips for TenSette Pipet 1970001 50/pkg 2185696
Pipet tips for TenSette Pipet 1970001 1000/pkg 2185628
Paper, pH, 0–14 pH range 100/pkg 2601300
Water, deionized 4L 27256
Thermometer, non-mercury, -10 to +225 °C each 2635700
Finger cots 2/pkg 1464702
Bottle, sampling, with cap, low density polyethylene, 250-mL 12/pkg 2087076
Cylinder, mixing, 25-mL each 189640
Funnel, micro each 2584335

Optional standards

Description Unit Item no.

Phosphate Standard Solution, 30-mg/L as PO4 946 mL 1436716

Phosphate Standard Solution, 100-mg/L as PO4 100 mL 1436832
®
Phosphate Standard Solution, 10-mL Voluette Ampul, 50-mg/L as PO4 16/pkg 17110
Phosphate Standard Solution, 500-mg/L as PO4 100 mL 1424232

Phosphorus, Total, Molybdovanadate TNT Method (100.0 mg/L) 7

 FOR TECHNICAL ASSISTANCE, PRICE INFORMATION AND ORDERING: HACH COMPANY
In the U.S.A. – Call toll-free 800-227-4224 WORLD HEADQUARTERS
Outside the U.S.A. – Contact the HACH office or distributor serving you. Telephone: (970) 669-3050
On the Worldwide Web – www.hach.com; E-mail – [email protected] FAX: (970) 669-2932

© Hach Company/Hach Lange GmbH, 1989–2013. All rights reserved. 04/2013, Edition 8
Phosphorus, Total DOC316.53.01121

USEPA1 PhosVer® 3 with Acid Persulfate Digestion Method Method 8190

0.06 to 3.50 mg/L PO43– (0.02 to 1.10 mg/L P) Test ‘N Tube™ Vials
Scope and application: For water, wastewater and seawater.
1 USEPA accepted for reporting wastewater analyses (Standard Methods 4500-P E).

Test preparation

Instrument specific information

The table in this section shows all of the instruments that have the program for this test.
Table 1 shows adapter and light shield requirements for the instruments that use them.
To use the table, select an instrument, then read across to find the corresponding
information for this test.
Table 1 Instrument-specific information for test tubes
Instrument Adapters Light shield
DR 6000 — —
DR 5000 — —
DR 900 4846400 Cover supplied with the instrument
DR 3900 — LZV849
DR 3800 — LZV646
DR 2800 —

Before starting
Install the instrument cap on the DR 900 cell holder before ZERO or READ is pushed.
DR 3900, DR 3800, DR 2800 and DR 2700: Install the light shield in Cell Compartment #2 before this test is started.
For best results, measure the reagent blank value for each new lot of reagent. Replace the sample with deionized water in
the test procedure to get the reagent blank value. Subtract the reagent blank value from the sample results automatically
with the reagent blank adjust option.
The test range for total phosphate is limited to 0.06 to 3.5 mg/L PO43–. Test results that are more than 3.5 mg/L can be used
to estimate dilution ratios, but should NOT be used for reporting purposes. If the test result is more than 3.5 mg/L, dilute the
sample and repeat the digestion and the colorimetric test.
Clean all glassware with 6.0 N (50%) hydrochloric acid, then rinse thoroughly with deionized water to remove contaminants.
The reagent that is used in this test is corrosive. Use protection for eyes and skin and be prepared to flush any spills with
running water.
Review the Safety Data Sheets (MSDS/SDS) for the chemicals that are used and use any recommended personal protective
equipment.
Dispose of reacted solutions according to local, state and federal regulations. Use the Safety Data Sheets for disposal
information for unused reagents. Consult the environmental, health and safety staff for your facility and/or local regulatory
agencies for further disposal information.

Items to collect
Description Quantity

Total Phosphorus Test 'N Tube Reagent Set 1

DRB200 Reactor 1

1
Items to collect (continued)
Description Quantity

Funnel, micro 1
Light shield or adapter (For information about sample cells, adapters or light shields, refer to
1
Instrument specific information on page 1.)
®
Pipet, TenSette , 1.0- to 10.0-mL, with pipet tips 1
Test tube rack 1
Water, deionized varies

Refer to Consumables and replacement items on page 5 for reorder information.

Sample collection and storage

• Collect samples in clean glass or plastic bottles that have been cleaned with
1:1 hydrochloric acid and rinsed with deionized water.
• Analyze the samples as soon as possible for best results.
• Do not use a detergent that contains phosphate to clean the sample bottles. The
phosphate in the detergent will contaminate the sample.
• To preserve samples for later analysis, adjust the sample pH to less than 2 with
concentrated sulfuric acid (about 2 mL per liter). No acid addition is necessary if the
sample is tested immediately.
• Keep the preserved samples at or below 6 °C (43 °F) for up to 28 days.
• Let the sample temperature increase to room temperature before analysis.
• Before analysis, adjust the pH to 7 with 5.0 N sodium hydroxide standard solution.
• Correct the test result for the dilution from the volume additions.

Acid persulfate digestion for Test 'N Tubes

Start

1. Start the 2. Start program 536 P 3. Add 5.0 mL of sample to 4. Add the contents of one
DRB200 Reactor. Preheat Total/AH PV TNT. For the Total Phosphorus Test Potassium Persulfate
to 150 °C. Refer to the information about sample Vial. Powder Pillow for
DRB200 manual. cells, adapters or light Phosphonate to the vial.
shields, refer to Instrument
specific information
on page 1.
Note: Although the program
name may vary between
instruments, the program
number does not change.

2 Phosphorus, Total, PhosVer 3 TNT Method (3.50 mg/L)

5. Put the cap on the vial. 6. Insert the vial into the 7. Start the instrument 8. When the timer expires,
Shake to dissolve the reactor. Close the reactor. timer. A 30-minute reaction carefully remove the vial
powder. time starts. from the reactor. Set the vial
in a test tube rack. Let the
vial cool to room
temperature.

9. Add 2 mL of 1.54 N 10. Put the cap on the vial. 11. Clean the vial. 12. Insert the vial into the
Sodium Hydroxide Standard Invert to mix. 16-mm cell holder.
Solution to the vial.

Zero

13. Push ZERO. The 14. Add the contents of one 15. Put the cap on the vial. 16. Start the instrument
display shows 0.00 mg/L PhosVer 3 Powder Pillow to Shake to mix for 20– timer. A 2-minute reaction
PO43–. the vial. 30 seconds. The powder will time starts.
not dissolve completely. Measure the sample within
two to eight minutes after
the timer expires.

Phosphorus, Total, PhosVer 3 TNT Method (3.50 mg/L) 3

 Read

17. Clean the vial. 18. Insert the vial into the 19. Push READ. Results
16-mm cell holder. show in mg/L PO43–.

Interferences
Interfering substance Interference level
Aluminum More than 200 mg/L
Arsenate Interferes at any level.
Chromium More than 100 mg/L
Copper More than 10 mg/L
Sulfide More than 90 mg/L
Iron More than 100 mg/L
Nickel More than 300 mg/L
Highly buffered samples or Can prevent the correct pH adjustment of the sample by the reagents. Sample pretreatment
extreme sample pH may be necessary.
Silica More than 50 mg/L
Silicate More than 10 mg/L
Turbidity or color May cause inconsistent results. The acid in the powder pillow can dissolve some of the
suspended particles and the desorption of orthophosphate from the particles can vary. For
highly turbid or colored samples, add the contents of one Phosphate Pretreatment Powder
Pillow to 25 mL of sample. Mix well. Use this solution to zero the instrument.
Zinc More than 80 mg/L

Accuracy check
Standard additions method (sample spike)
Use the standard additions method (for applicable instruments) to validate the test
procedure, reagents and instrument and to find if there is an interference in the sample.
Items to collect:
• Phosphate 10-mL Ampule Standard, 50-mg/L as PO43–
• Ampule breaker
• Pipet, TenSette®, 0.1–1.0 mL and tips
• Mixing cylinders, 25-mL (3)

1. Use the test procedure to measure the concentration of the sample, then keep the
(unspiked) sample in the instrument.
2. Go to the Standard Additions option in the instrument menu.
3. Select the values for standard concentration, sample volume and spike volumes.
4. Open the standard solution.

4 Phosphorus, Total, PhosVer 3 TNT Method (3.50 mg/L)

 5. Prepare three spiked samples: use the TenSette pipet to add 0.1 mL, 0.2 mL and
0.3 mL of the standard solution, respectively, to three 25-mL portions of fresh sample.
Mix well.
6. Use the test procedure to measure the concentration of each of the spiked samples.
Start with the smallest sample spike. Measure each of the spiked samples in the
instrument.
7. Select Graph to compare the expected results to the actual results.
Note: If the actual results are significantly different from the expected results, make sure that
the sample volumes and sample spikes are measured accurately. The sample volumes and
sample spikes that are used should agree with the selections in the standard additions menu. If
the results are not within acceptable limits, the sample may contain an interference.

Standard solution method

Use the standard solution method to validate the test procedure, reagents and
instrument.
Items to collect:
• 3.0-mg/L phosphate standard solution

1. Use the test procedure to measure the concentration of the standard solution.
2. Compare the expected result to the actual result.
Note: The factory calibration can be adjusted slightly with the standard adjust option so that the
instrument shows the expected value of the standard solution. The adjusted calibration is then
used for all test results. This adjustment can increase the test accuracy when there are slight
variations in the reagents or instruments.

Method performance
The method performance data that follows was derived from laboratory tests that were
measured on a spectrophotometer during ideal test conditions. Users may get different
results under different test conditions.
Program Standard Precision (95% Confidence Interval) Sensitivity
Concentration change per 0.010 Abs change
536 3.00 mg/L PO43– 2.93–3.07 mg/L PO43– 0.06 mg/L PO43–

Summary of method
Phosphates present in organic and condensed inorganic forms (meta-, pyro- or other
polyphosphates) must be converted to reactive orthophosphate before analysis.
Pretreatment of the sample with acid and heat provides the conditions for hydrolysis of
the condensed inorganic forms. Organic phosphates are converted to orthophosphates
by heating with acid and persulfate. Orthophosphate reacts with molybdate in an acid
medium to produce a mixed phosphate/molybdate complex. Ascorbic acid then reduces
the complex, giving an intense molybdenum blue color. The measurement wavelength is
880 nm for spectrophotometers or 610 nm for colorimeters.
Pollution prevention and waste management
Reacted samples contain molybdenum and must be disposed of as a hazardous waste.
Dispose of reacted solutions according to local, state and federal regulations.
Consumables and replacement items
Required reagents

Description Quantity/test Unit Item no.

Water, deionized varies 100 mL 27242

Total Phosphorus Test ’N Tube™ Reagent Set 1 50 tests 2742645
Includes:
®
PhosVer 3 Phosphate Reagent Powder Pillow, 10-mL 1 50/pkg 2106046

Phosphorus, Total, PhosVer 3 TNT Method (3.50 mg/L) 5

Consumables and replacement items (continued)
Description Quantity/test Unit Item no.

Potassium Persulfate Powder Pillows 1 pillow 50/pkg 2084766

Sodium Hydroxide, 1.54 N varies 100 mL 2743042
Total and Acid Hydrolyzable Test Vials (not sold separately) 1 50/pkg —

Required apparatus

Description Quantity/test Unit Item no.

DRB200 Reactor, 110 V, 15 x 16 mm 1 each LTV082.53.40001

DRB200 Reactor, 220 V, 15 x 16 mm 1 each LTV082.52.40001
Funnel, micro, poly 1 each 2584335
Light shield, DR 3900 1 each LZV849
Light shield, DR 3800, DR 2800, DR 2700 1 each LZV646
®
Pipet, TenSette , 1.0- to 10.0-mL 1 each 1970010
Pipet Tips, for TenSette Pipet 1970010 2 250/pkg 2199725
Test tube rack 1 each 1864100

Recommended standards and apparatus

Description Unit Item no.

Drinking Water Standard, Mixed Parameter, Inorganic for F-, NO3, PO4, SO4 500 mL 2833049
®
Phosphate Standard Solution, 50-mg/L, 10-mL Voluette Ampules 16/pkg 17110
Phosphate Standard Solution, 1-mg/L as PO4 500 mL 256949
Phosphate Standard Solution, 3-mg/L as PO4 946 mL 2059716
Wastewater, Effluent Inorganics, for NH3-N, NO3-N, PO4, COD, SO4, TOC 500 mL 2833249
®
Ampule Breaker, Voluette ampules each 2196800

Optional reagents and apparatus

Description Unit Item no.

Cylinder, mixing, 25-mL each 189640

Pipet, volumetric, Class A, 2-mL each 1451536
Hydrochloric Acid, 6.0 N 1:1, 50% 500 mL 88449
Sodium Hydroxide Standard Solution, 5.0 N 1L 245053
Sulfuric Acid, concentrated, ACS 500 mL 97949
®
Pipet, TenSette , 0.1–1.0 mL each 1970001
Pipet tips for TenSette Pipet 1970001 50/pkg 2185696
Pipet tips for TenSette Pipet 1970001 1000/pkg 2185628
Bottle, sampling, with cap, low density polyethylene, 250-mL 12/pkg 2087076
Paper, pH, 0–14 pH range 100/pkg 2601300
Water, deionized 4L 27256
Thermometer, non-mercury, -10 to +225 °C each 2635700
Finger cots 2/pkg 1464702

6 Phosphorus, Total, PhosVer 3 TNT Method (3.50 mg/L)

Optional standards

Description Unit Item no.

Phosphate Standard Solution, 10-mg/L as PO4 946 mL 1420416

Phosphate Standard Solution, 15-mg/L as PO4 100 mL 1424342
Phosphate Standard Solution, 100-mg/L as PO4 100 mL 1436832
Phosphate Standard Solution, 10-mL Ampule, 500 mg/L as PO4 16/pkg 1424210
Phosphate Standard Solution, 500-mg/L as PO4 100 mL 1424232

Phosphorus, Total, PhosVer 3 TNT Method (3.50 mg/L) 7

 FOR TECHNICAL ASSISTANCE, PRICE INFORMATION AND ORDERING: HACH COMPANY
In the U.S.A. – Call toll-free 800-227-4224 WORLD HEADQUARTERS
Outside the U.S.A. – Contact the HACH office or distributor serving you. Telephone: (970) 669-3050
On the Worldwide Web – www.hach.com; E-mail – [email protected] FAX: (970) 669-2932

© Hach Company/Hach Lange GmbH, 1989–2013. All rights reserved. 04/2013, Edition 8
pH DOC316.53.01330

Phenol Red Method (colorimeters only) Method 10076

6.5 to 8.5 pH units
Scope and application: For water and wastewater

Test preparation

Instrument-specific table
The table in this section shows all of the instruments that have the program for this test.
Instrument specific information PP shows sample cell and orientation requirements for
reagent addition tests, such as powder pillow or bulk reagent tests.
Table 1 Instrument-specific information for reagent addition
Instrument Sample cell orientation Sample cell
DR 900 The orientation mark is toward the user. 2401906

Before starting
Install the instrument cap on the DR 900 cell holder before ZERO or READ is pushed.
The sample temperature must be between 21–29 °C (70–84 °F) for accurate results.
Review the Safety Data Sheets (MSDS/SDS) for the chemicals that are used and use any recommended personal protective
equipment.
Dispose of reacted solutions according to local, state and federal regulations. Use the Safety Data Sheets for disposal
information for unused reagents. Consult the environmental, health and safety staff for your facility and/or local regulatory
agencies for further disposal information.

Items to collect

Description Quantity

Phenol Red Indicator Solution, spec grade 1.0 mL

Dropper with 0.5 and 1.0 mL marks 1
Sample cells, with caps 2

Refer to Consumables and replacement items on page 3 for reorder information.

Sample collection
• Collect samples in clean glass or plastic bottles.
• Analyze the samples as soon as possible for best results.

1
Colorimetric procedure

Start

1. Start program 461 pH. 2. Prepare the blank: Fill 3. Clean the blank. 4. Insert the blank into the
the sample cell with 10 mL cell holder.
of sample.

Zero

5. Push ZERO. The display 6. Prepare the sample: Fill 7. Use a dropper to add 8. Close the prepared
shows 6.0 pH . a second sample cell with 1 mL of Phenol Red sample cell. Invert the
10 mL of sample. Indicator Solution to the prepared sample two times
prepared sample. to mix.

Read

9. Clean the prepared 10. Insert the prepared 11. Push READ. Results
sample. sample into the cell holder. show in pH units.

Interferences
Chlorine does not interfere at levels of 6 mg/L Cl2 or less. Salt water (seawater) interferes
and cannot be analyzed with this method.
Accuracy check
Standard solution method
Use the standard solution method to validate the test procedure, reagents and
instrument.

2 pH, Phenol Red Colorimetric Method (6.5–8.5 pH units)

 Items to collect:
• pH 7.0 buffer solution, clear

1. Use the test procedure to measure the concentration of the standard solution.
2. Compare the expected result to the actual result.
Note: The factory calibration can be adjusted slightly with the standard adjust option so that the
instrument shows the expected value of the standard solution. The adjusted calibration is then
used for all test results. This adjustment can increase the test accuracy when there are slight
variations in the reagents or instruments.

Method performance
The method performance data that follows was derived from laboratory tests that were
measured on a spectrophotometer during ideal test conditions. Users may get different
results under different test conditions.
Program Standard Precision (95% Confidence Interval) Sensitivity
Concentration change per 0.010 Abs change
461 pH 7.0 buffer solution < 0.1 pH units not applicable

Summary of method
This method uses a sulfonphthalein indicator (Phenol Red) to determine pH
colorimetrically. Test results are measured at 520 nm. This method is available for
colorimeters only.
Consumables and replacement items
Required reagents

Description Quantity/test Unit Item no.

Phenol Red Indicator Solution, spec grade 1.0 mL 50 mL 2657512

Sample Cells, 10-20-25 mL, w/ cap 2 6/pkg 2401906

Required apparatus

Description Quantity/test Unit Item no.

Dropper, measuring, 0.5 and 1.0 mL plastic 1 20/pkg 2124720

Optional reagents and apparatus

Description Unit Item no.

pH 7.0 Buffer Solution 500 mL 1222249

Thermometer, -20 to 110 °C, Non-Mercury each 2635702

pH, Phenol Red Colorimetric Method (6.5–8.5 pH units) 3

 FOR TECHNICAL ASSISTANCE, PRICE INFORMATION AND ORDERING: HACH COMPANY
In the U.S.A. – Call toll-free 800-227-4224 WORLD HEADQUARTERS
Outside the U.S.A. – Contact the HACH office or distributor serving you. Telephone: (970) 669-3050
On the Worldwide Web – www.hach.com; E-mail – [email protected] FAX: (970) 669-2932

© Hach Company/Hach Lange GmbH, 1989–2013. All rights reserved. 04/2013, Edition 8
Silica DOC316.53.01132

Heteropoly Blue Method1 Method 8186

0.010 to 1.600 mg/L SiO2 (LR, spectrophotometers) Powder Pillows
0.01 to 1.60 mg/L SiO2 (LR, colorimeters)
Scope and application: For boiler and ultrapure water.
1 Adapted from Standard Methods for the Examination of Water and Wastewater.

Test preparation

Instrument specific information

The table in this section shows all of the instruments that have the program for this test.
Table 1 shows sample cell and orientation requirements for reagent addition tests, such
as powder pillow or bulk reagent tests.
To use the table, select an instrument, then read across to find the corresponding
information for this test.
Table 1 Instrument-specific information for powder pillows
Instrument Sample cell orientation Sample cell
DR 6000 The fill line is to the right. 2495402
DR 3800
DR 2800
DR 2700
DR 5000 The fill line is toward the user.
DR 3900
DR 900 The orientation mark is toward the user. 2401906

Before starting
Install the instrument cap on the DR 900 cell holder before ZERO or READ is pushed.
The reaction times in the test procedure are for samples that are at 20 °C (68 °F). If the sample temperature is 10 °C (50 °F),
wait 8 minutes for the first (4-minute) reaction time and 2 minutes for the second (1-minute) reaction time. If the sample
temperature is 30 °C (86 °F), wait 2 minutes for the first (4-minute) reaction time and 30 seconds for the second (1-minute)
reaction time.
To test for very low concentrations, use the ULR rapid liquid method for the best results.
Review the Safety Data Sheets (MSDS/SDS) for the chemicals that are used and use any recommended personal protective
equipment.
Dispose of reacted solutions according to local, state and federal regulations. Use the Safety Data Sheets for disposal
information for unused reagents. Consult the environmental, health and safety staff for your facility and/or local regulatory
agencies for further disposal information.

1
Items to collect
Description Quantity

Amino Acid F Reagent powder pillows, 10-mL 1

Citric acid powder pillows, 10-mL 2
Molybdate 3 Reagent solution 1 mL
Sample cells (For information about sample cells, adapters or light shields, refer to Instrument
2
specific information on page 1.)

Refer to Consumables and replacement items on page 5 for reorder information.

Sample collection
• Collect samples in clean plastic bottles with tight-fitting caps. Do not use glass
bottles, which will contaminate the sample.
• Analyze the samples as soon as possible for best results.
• If prompt analysis is not possible, keep the samples at or below 6 °C (43 °F) for up to
28 days.
• Let the sample temperature increase to room temperature before analysis.

Powder pillow procedure

Start

1. Start program 651 Silica 2. Prepare the blank: Fill a 3. Prepare the sample: Fill 4. Add 14 drops of
LR. For information about sample cell with 10 mL of a second sample cell with Molybdate 3 reagent
sample cells, adapters or sample. 10 mL of sample. solution to each cell.
light shields, refer to
Instrument specific
information on page 1.
Note: Although the program
name may vary between
instruments, the program
number does not change.

2 Silica, Heteropoly Blue Method (1.600 mg/L)

5. Swirl to mix. 6. Start the instrument 7. After the timer expires, 8. Swirl to mix.
timer. A 4-minute reaction add the contents of one
time starts. Citric Acid Reagent powder
pillow to each sample cell.

9. Start the instrument 10. After the timer expires, 11. Swirl to mix. 12. Start the instrument
timer. A 1-minute reaction add the contents of one timer. A 2-minute reaction
time starts. Amino Acid F Reagent time starts.
The destruction of possible powder pillow to the A blue color shows if silica is
phosphate interference prepared sample cell. present.
occurs during this period. Blank: The sample without
the Amino Acid F Reagent is
the blank.

Zero

13. When the timer expires, 14. Insert the blank into the 15. Push ZERO. The 16. Clean the prepared
clean the blank. cell holder. display shows 0.000 mg/L sample.
SiO2.

Silica, Heteropoly Blue Method (1.600 mg/L) 3

 Read

17. Insert the prepared 18. Push READ. Results

sample into the cell holder. show in mg/L SiO2.

Interferences
Interfering Interference level
substance
Color Does not interfere when the original sample is used to zero the instrument.
Iron Large amounts of both ferrous and ferric iron interfere.
Phosphate Does not interfere at levels less than 50 mg/L PO4. At 60 mg/L PO4, an interference of –2% occurs. At
75 mg/L PO4, the interference is –11%.
Slow reacting forms Occasionally a sample contains silica which reacts very slowly with molybdate. The nature of these
of silica "molybdate-unreactive" forms is not known. A pretreatment with sodium bicarbonate, then sulfuric acid
will make these forms reactive to molybdate. The pretreatment is given in Standard Methods for the
Examination of Water and Wastewater under Silica-Digestion with Sodium Bicarbonate. A longer
reaction time with the sample and the molybdate and acid reagents (before the citric acid is added) can
help as an alternative to the bicarbonate pretreatment.
Sulfides Interfere at all levels.
Turbidity Does not interfere when the original sample is used to zero the instrument.

Accuracy check
Standard additions method (sample spike)
Use the standard additions method (for applicable instruments) to validate the test
procedure, reagents and instrument and to find if there is an interference in the sample.
Items to collect:
• Silica Standard Solution, 25 mg/L SiO2
®
• Pipet, TenSette , 0.1–1.0 mL
• Pipet tips

1. Use the test procedure to measure the concentration of the sample, then keep the
(unspiked) sample in the instrument.
2. Go to the Standard Additions option in the instrument menu.
3. Select the values for standard concentration, sample volume and spike volumes.
4. Open the standard solution.
5. Prepare three spiked samples: use the TenSette pipet to add 0.1 mL, 0.2 mL and
0.3 mL of the standard solution, respectively, to three 10-mL portions of fresh sample.
Mix well.
6. Use the test procedure to measure the concentration of each of the spiked samples.
Start with the smallest sample spike. Measure each of the spiked samples in the
instrument.
7. Select Graph to compare the expected results to the actual results.
Note: If the actual results are significantly different from the expected results, make sure that
the sample volumes and sample spikes are measured accurately. The sample volumes and

4 Silica, Heteropoly Blue Method (1.600 mg/L)

 sample spikes that are used should agree with the selections in the standard additions menu. If
the results are not within acceptable limits, the sample may contain an interference.

Standard solution method

Use the standard solution method to validate the test procedure, reagents and
instrument.
Items to collect:
• Silica Standard Solution, 1.00 mg/L SiO2

1. Use the test procedure to measure the concentration of the standard solution.
2. Compare the expected result to the actual result.
Note: The factory calibration can be adjusted slightly with the standard adjust option so that the
instrument shows the expected value of the standard solution. The adjusted calibration is then
used for all test results. This adjustment can increase the test accuracy when there are slight
variations in the reagents or instruments.

Method performance
The method performance data that follows was derived from laboratory tests that were
measured on a spectrophotometer during ideal test conditions. Users may get different
results under different test conditions.
Program Standard Precision (95% Confidence Interval) Sensitivity
Concentration change per 0.010 Abs change
651 1.000 mg/L SiO2 0.990–1.010 mg/L SiO2 0.012 mg/L SiO2

Summary of method
Silica and phosphate in the sample react with molybdate ion under acidic conditions to
form yellow silicomolybdic acid complexes and phosphomolybdic acid complexes.
Addition of citric acid destroys the phosphate complexes. An amino acid is then added to
reduce the yellow silicomolybdic acid to an intense blue color, which is proportional to the
silica concentration. The measurement wavelength is 815 nm for spectrophotometers or
610 nm for colorimeters.
Consumables and replacement items
Required reagents

Description Quantity/Test Unit Item no.

Silica Reagent Set, low range, includes: — 100 tests 2459300

Amino Acid F Reagent Powder Pillow, 10-mL 1 100/pkg 2254069
Citric Acid Powder Pillow, 10-mL 2 100/pkg 2106269
Molybdate 3 Reagent Solution 1 mL 50 mL 199526

Recommended standards and apparatus

Description Unit Item no.

Silica Standard Solution, 1-mg/L SiO2 500 mL 110649

Silica Standard Solution, 25-mg/L as SiO2 236 mL 2122531
Water, deionized 4L 27256

Silica, Heteropoly Blue Method (1.600 mg/L) 5

Optional reagents and apparatus

Description Unit Item no.

Sodium Bicarbonate 454 g 77601

Sulfuric Acid, 1.00 N 1000 mL 127053
®
Pipet, TenSette , 0.1–1.0 mL each 1970001
Pipet tips for TenSette Pipet 1970001 50/pkg 2185696
Pipet tips for TenSette Pipet 1970001 1000/pkg 2185628

FOR TECHNICAL ASSISTANCE, PRICE INFORMATION AND ORDERING: HACH COMPANY

In the U.S.A. – Call toll-free 800-227-4224 WORLD HEADQUARTERS
Outside the U.S.A. – Contact the HACH office or distributor serving you. Telephone: (970) 669-3050
On the Worldwide Web – www.hach.com; E-mail – [email protected] FAX: (970) 669-2932

© Hach Company/Hach Lange GmbH, 1989–2013. All rights reserved. 04/2013, Edition 8
Silica DOC316.53.01133

Silicomolybdate Method Method 8185

1 to 100 mg/L SiO2 (HR, spectrophotometers) Powder Pillows
1 to 75 mg/L SiO2 (HR, colorimeters)
Scope and application: For water and seawater.

Test preparation

Instrument specific information

The table in this section shows all of the instruments that have the program for this test.
Table 1 shows sample cell and orientation requirements for reagent addition tests, such
as powder pillow or bulk reagent tests.
To use the table, select an instrument, then read across to find the corresponding
information for this test.
Table 1 Instrument-specific information for powder pillows
Instrument Sample cell orientation Sample cell
DR 6000 The fill line is to the right. 2495402
DR 3800
DR 2800
DR 2700
DR 5000 The fill line is toward the user.
DR 3900
DR 900 The orientation mark is toward the user. 2401906

Before starting
Install the instrument cap on the DR 900 cell holder before ZERO or READ is pushed.
The sample temperature must be between 15–25 °C (59–77 °F) for accurate results.
Use the Standard Adjust option with each new lot of reagent for the best results.
Review the Safety Data Sheets (MSDS/SDS) for the chemicals that are used and use any recommended personal protective
equipment.
Dispose of reacted solutions according to local, state and federal regulations. Use the Safety Data Sheets for disposal
information for unused reagents. Consult the environmental, health and safety staff for your facility and/or local regulatory
agencies for further disposal information.

Items to collect
Description Quantity

High Range SIlica Reagent Set 1

Sample cells (For information about sample cells, adapters or light shields, refer to Instrument
2
specific information on page 1.)

1
Refer to Consumables and replacement items on page 4 for reorder information.

Sample collection
• Collect samples in clean plastic bottles with tight-fitting caps. Do not use glass
bottles, which will contaminate the sample.
• Analyze the samples as soon as possible for best results.
• If prompt analysis is not possible, keep the samples at or below 6 °C (43 °F) for up to
28 days.
• Let the sample temperature increase to room temperature before analysis.

Powder pillow procedure

Start

1. Start program 656 Silica 2. Prepare the sample: Fill 3. Add the contents of one 4. Swirl until the reagent is
HR. For information about a sample cell with 10 mL of Molybdate Reagent Powder completely dissolved.
sample cells, adapters or sample. Pillow for High Range Silica
light shields, refer to to the sample cell.
Instrument specific
information on page 1.
Note: Although the program
name may vary between
instruments, the program
number does not change.

5. Add the contents of one 6. Swirl to mix. 7. Start the instrument 8. When the timer expires,
Acid Reagent Powder Pillow timer. A 10-minute reaction add the contents of one
for High Range Silica. time starts. Citric Acid Powder Pillow to
A yellow color shows if silica the sample cell.
or phosphorus is present in Any yellow color caused by
the sample. phosphorous is removed
during this step.

2 Silica, Silicomolybdate Method (100 mg/L)

9. Start the instrument 10. Prepare the blank: Fill 11. When the timer expires, 12. Insert the blank into the
timer. A 2-minute reaction a second sample cell with clean the blank. cell holder.
time starts. 10 mL of the original
Complete the rest of the sample.
procedure within three
minutes after the timer
expires.

Zero Read

13. Push ZERO. The 14. Clean the prepared 15. Insert the prepared 16. Push READ. Results
display shows 0 mg/L SiO2. sample. sample into the cell holder. show in mg/L SiO2.

Interferences
Interfering Interference level
substance
Color Does not interfere when the original sample is used to zero the instrument.
Iron Large amounts of both ferrous and ferric iron interfere.
Phosphate Does not interfere at levels less than 50 mg/L PO4. At 60 mg/L PO4, an interference of –2% occurs. At
75 mg/L PO4, the interference is –11%.
Slow reacting forms Occasionally a sample contains silica which reacts very slowly with molybdate. The nature of these
of silica "molybdate-unreactive" forms is not known. A pretreatment with sodium bicarbonate, then sulfuric acid
will make these forms reactive to molybdate. The pretreatment is given in Standard Methods for the
Examination of Water and Wastewater under Silica-Digestion with Sodium Bicarbonate. A longer
reaction time with the sample and the molybdate and acid reagents (before the citric acid is added) can
help as an alternative to the bicarbonate pretreatment.
Sulfides Interfere at all levels.
Turbidity Does not interfere when the original sample is used to zero the instrument.

Accuracy check
Standard additions method (sample spike)
Use the standard additions method (for applicable instruments) to validate the test
procedure, reagents and instrument and to find if there is an interference in the sample.
Items to collect:
• Silica Standard Solution, 1000 mg/L

Silica, Silicomolybdate Method (100 mg/L) 3

 • Pipet, TenSette®, 0.1–1.0 mL
• Pipet tips

1. Use the test procedure to measure the concentration of the sample, then keep the
(unspiked) sample in the instrument.
2. Go to the Standard Additions option in the instrument menu.
3. Select the values for standard concentration, sample volume and spike volumes.
4. Open the standard solution.
5. Prepare three spiked samples: use the TenSette pipet to add 0.1 mL, 0.2 mL and
0.3 mL of the standard solution, respectively, to three 10-mL portions of fresh sample.
Mix well.
6. Use the test procedure to measure the concentration of each of the spiked samples.
Start with the smallest sample spike. Measure each of the spiked samples in the
instrument.
7. Select Graph to compare the expected results to the actual results.
Note: If the actual results are significantly different from the expected results, make sure that
the sample volumes and sample spikes are measured accurately. The sample volumes and
sample spikes that are used should agree with the selections in the standard additions menu. If
the results are not within acceptable limits, the sample may contain an interference.

Standard solution method

Use the standard solution method to validate the test procedure, reagents and
instrument.
Items to collect:
• Silica Standard Solution, 50 mg/L

1. Use the test procedure to measure the concentration of the standard solution.
2. Compare the expected result to the actual result.
Note: The factory calibration can be adjusted slightly with the standard adjust option so that the
instrument shows the expected value of the standard solution. The adjusted calibration is then
used for all test results. This adjustment can increase the test accuracy when there are slight
variations in the reagents or instruments.

Method performance
The method performance data that follows was derived from laboratory tests that were
measured on a spectrophotometer during ideal test conditions. Users may get different
results under different test conditions.
Program Standard Precision (95% Confidence Interval) Sensitivity
Concentration change per 0.010 Abs change
656 50 mg/L SiO2 48–52 mg/L SiO2 1.0 mg/L SiO2

Summary of method
Silica and phosphate in the sample react with molybdate ion under acidic conditions to
form yellow silicomolybdic acid complexes and phosphomolybdic acid complexes.
Addition of citric acid destroys the phosphate complexes. Silica is then determined by
measuring the remaining yellow color. The measurement wavelength is 452 nm for
spectrophotometers or 420 nm for colorimeters.
Consumables and replacement items
Required reagents

Description Quantity/test Unit Item no.

Water, deionized varies 4L 27256

High Range Silica Reagent Set, 10-mL — 100 tests 2429600

4 Silica, Silicomolybdate Method (100 mg/L)

Consumables and replacement items (continued)
Description Quantity/test Unit Item no.

Includes:
Acid Reagent Powder Pillows for High Range Silica, 10-mL 1 100/pkg 2107469
Citric Acid Powder Pillow, 10-mL 2 100/pkg 2106269
Molybdate Reagent Powder Pillows for High Range Silica, 10-mL 1 100/pkg 2107369

Recommended standards

Description Unit Item no.

Silica Standard Solution, 50-mg/L as SiO2 200 mL 111729

Silica Standard Solution, 1000-mg/L as SiO2 500 mL 19449

Optional reagents and apparatus

Description Unit Item no.

Sodium Bicarbonate 454 g 77601

100 mL
Sulfuric Acid Standard Solution, 1 N 127032
MDB
Bottle, sampling, with cap, low density polyethylene, 250-mL 12/pkg 2087076
Thermometer, non-mercury, -10 to +225 °C each 2635700

Silica, Silicomolybdate Method (100 mg/L) 5

 FOR TECHNICAL ASSISTANCE, PRICE INFORMATION AND ORDERING: HACH COMPANY
In the U.S.A. – Call toll-free 800-227-4224 WORLD HEADQUARTERS
Outside the U.S.A. – Contact the HACH office or distributor serving you. Telephone: (970) 669-3050
On the Worldwide Web – www.hach.com; E-mail – [email protected] FAX: (970) 669-2932

© Hach Company/Hach Lange GmbH, 1989–2013. All rights reserved. 04/2013, Edition 8
Sulfate DOC316.53.01135

USEPA1 SulfaVer 4 Method2 Method 8051

®
2 to 70 mg/L SO42– Powder Pillows or AccuVac Ampuls
Scope and application: For water, wastewater and seawater.
1 USEPA accepted for reporting wastewater analyses. Procedure is equivalent to USEPA method 375.4 for wastewater.
2 Adapted from Standard Methods for the Examination of Water and Wastewater.

Test preparation

Instrument-specific information
The tables in this section show all of the instruments that have the program for this test.
Table 1 shows sample cell and orientation requirements for reagent addition tests, such
as powder pillow or bulk reagent tests. Table 2 shows sample cell and adapter
requirements for AccuVac Ampul tests.
To use either table, select an instrument, then read across to find the corresponding
information for this test.
Table 1 Instrument-specific information for powder pillow
Instrument Sample cell orientation Sample cell
DR 6000 The fill line is to the right. 2495402
DR 3800
DR 2800
DR 2700
DR 5000 The fill line is toward the user.
DR 3900
DR 900 The orientation mark is toward the user. 2401906

Table 2 Instrument-specific information for AccuVac Ampuls

Instrument Adapter Sample cell
DR 6000 — 2427606
DR 5000
DR 900
DR 3900 LZV846 (A)
DR 3800 LZV584 (C) 2122800
DR 2800
DR 2700

Before starting
Install the instrument cap on the DR 900 cell holder before ZERO or READ is pushed.
Use the Standard Adjust option with each new lot of reagent for the best results.

1
For best results, calibrate the instrument with each new lot of reagent. Refer to Calibration on page 6.
For best results, measure the reagent blank value for each new lot of reagent. Replace the sample with deionized water in
the test procedure to get the reagent blank value. Subtract the reagent blank value from the sample results automatically
with the reagent blank adjust option.
Filter samples that are turbid with filter paper and a funnel.
Do not use the Pour-Thru Cell or sipper module (for applicable instruments) with this test.
The reagents that are used in this test contain barium chloride. Collect the reacted samples for proper disposal.
An AccuVac Ampule for Blanks can be used to zero the instrument in the AccuVac test procedure.
Review the Safety Data Sheets (MSDS/SDS) for the chemicals that are used and use any recommended personal protective
equipment.
Dispose of reacted solutions according to local, state and federal regulations. Use the Safety Data Sheets for disposal
information for unused reagents. Consult the environmental, health and safety staff for your facility and/or local regulatory
agencies for further disposal information.

Items to collect
Powder pillows

Description Quantity
®
SulfaVer 4 Reagent Powder Pillows, 10-mL 1
Sample Cells (Refer to Instrument-specific information on page 1. ) 2

Refer to Consumables and replacement items on page 7 for reorder information.

AccuVac Ampuls

Description Quantity
® ®
SulfaVer 4 Reagent AccuVac ampuls 1
Beaker, 50-mL 1
Sample Cells (Refer to Instrument-specific information on page 1. ) 1
Stopper 1

Refer to Consumables and replacement items on page 7 for reorder information.

Sample collection and storage

• Collect samples in clean glass or plastic bottles.
• To preserve samples for later analysis, keep the samples at or below 6 °C (43 °F) for
up to 28 days.
• Let the sample temperature increase to room temperature before analysis.

2 Sulfate, SulfaVer 4 Method (70 mg/L)

SulfaVer 4 powder pillow procedure

Start

1. Start program 2. Prepare the sample: Fill 3. Add the contents of one 4. Swirl the sample cell to
680 Sulfate. For information a sample cell with 10 mL of powder pillow to the sample mix. Undissolved powder
about sample cells, sample. cell. will not affect accuracy.
adapters or light shields, White turbidity will form if
refer to Instrument-specific sulfate is present.
information on page 1.
Note: Although the program
name may vary between
instruments, the program
number does not change.

5. Start the instrument 6. Prepare the blank: Fill a 7. When the timer expires, 8. Insert the blank into the
timer. A five-minute reaction second sample cell with clean the blank. cell holder.
time starts. 10 mL of sample.
Do not disturb the cell
during this time.

Zero Read

9. Push ZERO. The display 10. Clean the prepared 11. Within five minutes after 12. Push READ. Results
shows 0 mg/L SO42–. sample. the timer expires, insert the show in mg/L SO42–.
prepared sample into the
cell holder.

Sulfate, SulfaVer 4 Method (70 mg/L) 3

13. Clean the sample cells
with soap and a brush.

®
SulfaVer 4 AccuVac Ampuls procedure

Start

1. Start program 2. Prepare the sample: 3. Close the Ampul and 4. Start the instrument
685 Sulfate AV. For Collect at least 40 mL of quickly invert the Ampul timer. A five-minute reaction
information about sample sample in a 50-mL beaker. several times to mix. time starts.
cells, adapters or light Fill the AccuVac Ampul with Undissolved powder will not Do not disturb the cell
shields, refer to Instrument- sample. Keep the tip affect accuracy. during this time.
specific information immersed while the Ampul White turbidity will form if
on page 1. fills completely. sulfate is present.
Note: Although the program
name may vary between
instruments, the program
number does not change.

Zero

5. Prepare the blank: Fill 6. When the timer expires, 7. Insert the blank AccuVac 8. Push ZERO. The display
the sample cell with 10 mL clean the blank AccuVac Ampul into the cell holder. shows 0 mg/L SO42–.
of sample. Ampul.

4 Sulfate, SulfaVer 4 Method (70 mg/L)

 Read

9. Clean the AccuVac 10. Within five minutes after 11. Push READ. Results
Ampul. the timer expires, insert the show in mg/L SO42–.
prepared sample AccuVac
Ampul into the cell holder.

Interferences
Interfering substance Interference level
Barium Interferes at all levels. The higher the barium concentration when compared to the sulfate
concentration, the higher the error. Samples with high barium concentrations will generally give a
result that is 20% lower than the actual sulfate concentration.
Calcium More than 20,000 mg/L as CaCO3
Chloride More than 40,000 mg/L as Cl–
Magnesium More than 10,000 mg/L as CaCO3
Silica More than 500 mg/L SiO2

Accuracy check
Standard additions method (sample spike)
Use the standard additions method (for applicable instruments) to validate the test
procedure, reagents and instrument and to find if there is an interference in the sample.
Items to collect:
• Sulfate Ampule Standard Solution, 2500 mg/L sulfate
• Ampule breaker
• Pipet, TenSette®, 0.1–1.0 mL and tips
• Mixing cylinders (3), 25

1. Use the test procedure to measure the concentration of the sample, then keep the
(unspiked) sample in the instrument.
2. Go to the Standard Additions option in the instrument menu.
3. Select the values for standard concentration, sample volume and spike volumes.
4. Open the standard solution.
5. Prepare three spiked samples: use the TenSette pipet to add 0.1 mL, 0.2 mL and
0.3 mL of the standard solution, respectively, to three 25-mL portions of fresh sample.
Mix well.
®
Note: For AccuVac Ampuls, add 0.2 mL, 0.4 mL and 0.6 mL of the standard solution to three
50-mL portions of fresh sample.
6. Use the test procedure to measure the concentration of each of the spiked samples.
Start with the smallest sample spike. Measure each of the spiked samples in the
instrument.
7. Select Graph to compare the expected results to the actual results.
Note: If the actual results are significantly different from the expected results, make sure that
the sample volumes and sample spikes are measured accurately. The sample volumes and

Sulfate, SulfaVer 4 Method (70 mg/L) 5

 sample spikes that are used should agree with the selections in the standard additions menu. If
the results are not within acceptable limits, the sample may contain an interference.

Standard solution method

Use the standard solution method to validate the test procedure, reagents and
instrument.
Items to collect:
• Sulfate standard solution, 1000 mg/L
• 100-mL volumetric flask, Class A
• 7-mL volumetric pipet, Class A and pipet filler
• Deionized water

1. Prepare a 70 mg/L sulfate standard solution as follows:

a. Use a pipet to add 7.0 mL of 1000 mg/L sulfate standard solution into the
volumetric flask.
b. Dilute to the mark with deionized water. Mix well. Prepare this solution daily.
2. Use the test procedure to measure the concentration of the prepared standard
solution.
3. Compare the expected result to the actual result.
Note: The factory calibration can be adjusted slightly with the standard adjust option so that the
instrument shows the expected value of the standard solution. The adjusted calibration is then
used for all test results. This adjustment can increase the test accuracy when there are slight
variations in the reagents or instruments.

Calibration
A calibration is recommended for the SulfaVer 4 method for the best accuracy. Complete
the following steps to enter a new calibration curve in the instrument. Perform this
procedure for each new lot of reagent.
Items to collect:
• Sulfate standard solution, 1000 mg/L
• 100-mL volumetric flasks (7), Class A
• 1–10 mL TenSette pipet and tips
• Deionized water

1. Prepare seven calibration standard solutions (10, 20, 30, 40, 50, 60 and 70 mg/L
SO42–) as follows:
a. Use a pipet to add 1, 2, 3, 4, 5, 6 and 7 mL of the 1000-mg/L sulfate standard
solution into seven different 100-mL volumetric flasks.
b. Dilute each flask to the mark with deionized water. Mix well.
2. Use the test procedure to measure the concentration of each standard solution.
3. Refer to the user manual for the instrument to enter the calibration into the instrument
as a user program.

Method performance
The method performance data that follows was derived from laboratory tests that were
measured on a spectrophotometer during ideal test conditions. Users may get different
results under different test conditions.
Program Standard Precision (95% Confidence Interval) Sensitivity
Concentration change per 0.010 Abs change
680 40 mg/L SO42– 30–50 mg/L SO42– 0.4 mg/L SO42–
685 40 mg/L SO42– 32–48 mg/L SO42– 0.7 mg/L SO42–

6 Sulfate, SulfaVer 4 Method (70 mg/L)

Summary of method
Sulfate ions in the sample react with barium in the SulfaVer 4 and form a precipitate of
barium sulfate. The amount of turbidity formed is proportional to the sulfate concentration.
The measurement wavelength is 450 nm for spectrophotometers or 520 nm for
colorimeters.
Pollution prevention and waste management
Reacted samples contain barium and must be disposed of as a hazardous waste.
Dispose of reacted solutions according to local, state and federal regulations.
Consumables and replacement items
Required reagents

Description Quantity/test Unit Item no.

®
SulfaVer 4 Reagent Powder Pillows, 10-mL 1 100/pkg 2106769
OR
® ®
SulfaVer 4 Sulfate Reagent AccuVac Ampuls 1 25/pkg 2509025

Required apparatus

Description Quantity/test Unit Item no.

AccuVac Snapper 1 each 2405200

Beaker, 50-mL 1 each 50041H
Sample cell, 10 mL round, 25 x 54 mm 1 each 2122800
Sample cell, 10 mL round, 25 x 60 mm 1 6/pkg 2427606
Sample cell, 10 mL square, matched pair 2 2/pkg 2495402
Stoppers for 18-mm tubes and AccuVac Ampuls 2 6/pkg 173106

Recommended standards

Description Unit Item no.

Sulfate Standard Solution, 1000-mg/L as SO4 500 mL 2175749

Sulfate Standard Solution, 2500-mg/L, 10-mL Ampules as SO4 16/pkg 1425210
Drinking Water Standard, Mixed Parameter, Inorganic for F -, NO3, PO4, SO4 500 mL 2833049

Optional reagents and apparatus

Description Unit Item no.

Cylinder, mixing, 25-mL each 189640

Cylinder, mixing, 50-mL each 189641
Blank AccuVac Ampules 25/pkg 2677825
®
Ampule Breaker, Voluette ampules each 2196800
®
Pipet, TenSette , 0.1–1.0 mL each 1970001
Pipet tips for TenSette Pipet 1970001 50/pkg 2185696
®
Pipet, TenSette , 1.0 to 10.0 mL each 1970010
Pipet tips for TenSette Pipet 1970010 50/pkg 2199796
Flask, volumetric, Class A, 100-mL each 1457442

Sulfate, SulfaVer 4 Method (70 mg/L) 7

 FOR TECHNICAL ASSISTANCE, PRICE INFORMATION AND ORDERING: HACH COMPANY
In the U.S.A. – Call toll-free 800-227-4224 WORLD HEADQUARTERS
Outside the U.S.A. – Contact the HACH office or distributor serving you. Telephone: (970) 669-3050
On the Worldwide Web – www.hach.com; E-mail – [email protected] FAX: (970) 669-2932

© Hach Company/Hach Lange GmbH, 1989–2013. All rights reserved. 04/2013, Edition 8
Sulfate DOC316.53.01316

SulfaVer 4 Method1 Method 10248

2 to 70, 20 to 700, 200 to 7000 mg/L SO42– Powder Pillows
Scope and application: For oil and gas field waters.
1 Adapted from Standard Methods for the Examination of Water and Wastewater.

Test preparation

Instrument specific information

The table in this section shows all of the instruments that have the program for this test.
Table 1 shows sample cell and orientation requirements for reagent addition tests, such
as powder pillow or bulk reagent tests.
To use the table, select an instrument, then read across to find the corresponding
information for this test.
Table 1 Instrument-specific information for powder pillows
Instrument Sample cell orientation Sample cell
DR 6000 The fill line is to the right. 2495402
DR 3800
DR 2800
DR 2700
DR 5000 The fill line is toward the user.
DR 3900
DR 900 The orientation mark is toward the user. 2401906

Before starting
Install the instrument cap on the DR 900 cell holder before ZERO or READ is pushed.
Use the Standard Adjust option with each new lot of reagent for the best results.
For best results, measure the reagent blank value for each new lot of reagent. Replace the sample with deionized water in
the test procedure to get the reagent blank value. Subtract the reagent blank value from the sample results automatically
with the reagent blank adjust option.
Filter samples that are turbid with filter paper and a funnel.
Do not use the Pour-Thru Cell or sipper module (for applicable instruments) with this test.
The reagents that are used in this test contain barium chloride. Collect the reacted samples for proper disposal.
Review the Safety Data Sheets (MSDS/SDS) for the chemicals that are used and use any recommended personal protective
equipment.
Dispose of reacted solutions according to local, state and federal regulations. Use the Safety Data Sheets for disposal
information for unused reagents. Consult the environmental, health and safety staff for your facility and/or local regulatory
agencies for further disposal information.

1
Items to collect
Description Quantity
®
SulfaVer 4 Reagent Powder Pillows, 10-mL 1
Sample cells (For information about sample cells, adapters or light shields, refer to Instrument
1
specific information on page 1.)

Refer to Consumables and replacement items on page 5 for reorder information.

Sample collection and storage

• Collect samples in clean glass or plastic bottles.
• To preserve samples for later analysis, keep the samples at or below 6 °C (43 °F) for
up to 28 days.
• Let the sample temperature increase to room temperature before analysis.

Powder pillow procedure

Start

1. Start program 2. Add the sample volume 3. If the sample volume is 4. Swirl to mix.
680 Sulfate. For information that is specified for the test less than 10-mL add
about sample cells, range to a sample cell: deionized water to the 10-
adapters or light shields, mL line.
refer to Instrument specific • 2–70 mg/L: 10 mL For the dilution factor, refer
information on page 1. • 20–700 mg/L: 1.0 mL to Set the dilution factor
Note: Although the program • 200–7,000 mg/L: on page 3.
name may vary between 0.1 mL
instruments, the program
Use a TenSette Pipet or
number does not change.
glass pipet to measure
0.1 mL or 1.0 mL.

Zero

5. Clean the blank. 6. Insert the blank into the 7. Push ZERO. The display 8. Add the contents of one
cell holder. shows 0 mg/L SO42–. SulfaVer 4 Reagent Powder
Pillow to the sample cell.
The sample will get cloudy if
sulfate is present in the
sample.

2 Sulfate, SulfaVer 4 (multi-range: 70, 700, 7000 mg/L)

9. Swirl the sample cell to 10. Start the instrument 11. Clean the prepared 12. Within five minutes after
mix. Undissolved powder timer. A 5-minute reaction sample. the timer expires, insert the
will not affect accuracy. time starts. prepared sample into the
Do not move the sample cell cell holder.
during the reaction period.

Read

13. Push READ. Results 14. Clean the sample cell

show in mg/L SO42–. immediately after each test
with soap, water and a
brush.

Interferences
Interfering substance Interference level
Barium Interferes at all levels. The higher the barium concentration when compared to the sulfate
concentration, the higher the error. Samples with high barium concentrations will generally give a
result that is 20% lower than the actual sulfate concentration.
Calcium More than 20,000 mg/L as CaCO3
Chloride More than 40,000 mg/L as Cl–
Magnesium More than 10,000 mg/L as CaCO3
Silica More than 500 mg/L SiO2

Set the dilution factor

Instruments that have a dilution factor option can include the dilution factor in the result
and show the concentration of the original, undiluted sample. For example, if the sample
is diluted by a factor of 10, the instrument multiplies the result by 10 and shows the
calculated result in the instrument display.

1. Select Options>More>Dilution factor from the instrument menu.

Note: Colorimeters include a dilution factor when the chemical form is set. Go to
Options>Advanced Options>Chemical Form and select LR, MR or HR.
2. Enter the dilution factor:
• 1 mL sample diluted to 10 mL: dilution factor is 10.
• 0.1 mL sample diluted to 10 mL: dilution factor is 100.

Sulfate, SulfaVer 4 (multi-range: 70, 700, 7000 mg/L) 3

 3. Push OK to confirm. Push OK again.
4. Push RETURN to go back to the measurement screen.

Accuracy check
Standard additions method (sample spike)
Use the standard additions method (for applicable instruments) to validate the test
procedure, reagents and instrument and to find if there is an interference in the sample.
Items to collect:
• Sulfate Ampule Standard Solution, 2500 mg/L sulfate
• Ampule breaker
• Pipet, TenSette®, 0.1–1.0 mL and tips
• Mixing cylinders (3), 25 mL

1. Use the test procedure to measure the concentration of the sample, then keep the
(unspiked) sample in the instrument.
2. Go to the Standard Additions option in the instrument menu.
3. Select the values for standard concentration, sample volume and spike volumes.
4. Open the standard solution.
5. Prepare three spiked samples: use the TenSette pipet to add 0.1 mL, 0.2 mL and
0.3 mL of the standard solution, respectively, to three 25-mL portions of fresh sample.
Mix well.
6. Use the test procedure to measure the concentration of each of the spiked samples.
Start with the smallest sample spike. Measure each of the spiked samples in the
instrument.
7. Select Graph to compare the expected results to the actual results.
Note: If the actual results are significantly different from the expected results, make sure that
the sample volumes and sample spikes are measured accurately. The sample volumes and
sample spikes that are used should agree with the selections in the standard additions menu. If
the results are not within acceptable limits, the sample may contain an interference.

Standard solution method

Use the standard solution method to validate the test procedure, reagents and
instrument.
Items to collect:
• Sulfate standard solution, 1000 mg/L
• 100-mL volumetric flask, Class A
• 5-mL volumetric pipet, Class A and pipet filler
• Deionized water

1. Prepare a 50 mg/L sulfate standard solution as follows:

a. Use a pipet to add 5.0 mL of 1000 mg/L sulfate standard solution into the
volumetric flask.
b. Dilute to the mark with deionized water. Mix well. Prepare this solution daily.
2. Use the test procedure to measure the concentration of the prepared standard
solution.
3. Compare the expected result to the actual result.
Note: The factory calibration can be adjusted slightly with the standard adjust option so that the
instrument shows the expected value of the standard solution. The adjusted calibration is then
used for all test results. This adjustment can increase the test accuracy when there are slight
variations in the reagents or instruments.

4 Sulfate, SulfaVer 4 (multi-range: 70, 700, 7000 mg/L)

Method performance
The method performance data that follows was derived from laboratory tests that were
measured on a spectrophotometer during ideal test conditions. Users may get different
results under different test conditions.
Program Standard Precision (95% Confidence Interval) Sensitivity
Concentration change per 0.010 Abs change
680 40 mg/L SO42– 30–50 mg/L SO42– 0.4 mg/L SO42–

Summary of method
Sulfate ions in the sample react with barium in the SulfaVer 4 and form a precipitate of
barium sulfate. The amount of turbidity formed is proportional to the sulfate concentration.
The measurement wavelength is 450 nm for spectrophotometers or 520 nm for
colorimeters.
Pollution prevention and waste management
Reacted samples contain barium and must be disposed of as a hazardous waste.
Dispose of reacted solutions according to local, state and federal regulations.
Consumables and replacement items
Required reagents

Description Quantity/test Unit Item no.

®
SulfaVer 4 Reagent Powder Pillows, 10-mL 1 100/pkg 2106769

Recommended standards

Description Unit Item no.

Sulfate Standard Solution, 1000-mg/L as SO4 500 mL 2175749

Sulfate Standard Solution, 2500-mg/L, 10-mL Ampules as SO4 16/pkg 1425210

Optional reagents and apparatus

Description Unit Item no.

Cylinder, mixing, 25-mL each 189640

Cylinder, mixing, 50-mL each 189641
®
Ampule Breaker, Voluette ampules each 2196800
Pipet, volumetric 5.00-mL each 1451537
Pipet Filler 1 1465000
®
Pipet, TenSette , 0.1–1.0 mL each 1970001
Pipet tips for TenSette Pipet 1970001 50/pkg 2185696
®
Pipet, TenSette , 1.0 to 10.0 mL each 1970010
Pipet tips for TenSette Pipet 1970010 50/pkg 2199796
Flask, volumetric, Class A, 100-mL each 1457442

Sulfate, SulfaVer 4 (multi-range: 70, 700, 7000 mg/L) 5

 FOR TECHNICAL ASSISTANCE, PRICE INFORMATION AND ORDERING: HACH COMPANY
In the U.S.A. – Call toll-free 800-227-4224 WORLD HEADQUARTERS
Outside the U.S.A. – Contact the HACH office or distributor serving you. Telephone: (970) 669-3050
On the Worldwide Web – www.hach.com; E-mail – [email protected] FAX: (970) 669-2932

© Hach Company/Hach Lange GmbH, 1989–2013. All rights reserved. 04/2013, Edition 8
Sulfide DOC316.53.01136

USEPA1 Methylene Blue Method2 Method 8131

5 to 800 µg/L S2– (spectrophotometers) Reagent Solution
0.01 to 0.70 mg/L (colorimeters)
Scope and application: For testing total sulfides, H2S, HS–, and certain metal sulfides in groundwater,
wastewater, brines and seawater.
1 USEPA approved for reporting wastewater analysis. Procedure is equivalent to Standard Method 4500-S2– D.
2 Adapted from Standard Methods for the Examination of Water and Wastewater.

Test preparation

Instrument specific information

The table in this section shows all of the instruments that have the program for this test.
Table 1 shows sample cell and orientation requirements for reagent addition tests, such
as powder pillow or bulk reagent tests.
To use the table, select an instrument, then read across to find the corresponding
information for this test.
Table 1 Instrument-specific information for reagent addition
Instrument Sample cell orientation Sample cell
DR 6000 The fill line is to the right. 2495402
DR 3800
DR 2800
DR 2700
DR 5000 The fill line is toward the user.
DR 3900
DR 900 The orientation mark is toward the user. 2401906

Before starting
Samples must be analyzed immediately after collection and cannot be preserved for later analysis.
Install the instrument cap on the DR 900 cell holder before ZERO or READ is pushed.
Some sulfide loss can occur if dilution is necessary.
Review the Safety Data Sheets (MSDS/SDS) for the chemicals that are used and use any recommended personal protective
equipment.
Dispose of reacted solutions according to local, state and federal regulations. Use the Safety Data Sheets for disposal
information for unused reagents. Consult the environmental, health and safety staff for your facility and/or local regulatory
agencies for further disposal information.

1
Items to collect
Description Quantity

Sulfide 1 Reagent 1–2 mL

Sulfide 2 Reagent 1–2 mL
Water, deionized 10–25 mL
Pipet, serological, 10-mL 1
Pipet Filler, safety bulb 1
Sample cells (For information about sample cells, adapters or light shields, refer to Instrument
2
specific information on page 1.)
Stoppers 2

Refer to Consumables and replacement items on page 5 for reorder information.

Sample collection
• Samples must be analyzed immediately after collection and cannot be preserved for
later analysis.
• Collect samples in clean glass or plastic bottles with tight-fitting caps. Fill the bottle
completely and immediately tighten the cap.
• Prevent agitation of the sample or exposure to air.

Methylene Blue Method

Start

1. Start program 2. Prepare the blank: Fill a 3. Prepare the sample: 4. Use a dropper to add
690 Sulfide. For information sample cell with deionized Use a pipet to add sample 0.5 mL Sulfide 1 Reagent to
about sample cells, water. Use 10 mL for to a second sample cell. each cell.
adapters or light shields, spectrophotometers and Use 10 mL for
refer to Instrument specific 25 mL for colorimeters. spectrophotometers and
information on page 1. 25 mL for colorimeters.
Note: Although the program Do not mix the sample more
name may vary between than necessary to prevent
instruments, the program sulfide loss.
number does not change.

2 Sulfide, Methylene Blue Method (800 µg/L)

5. Swirl to mix. 6. Use the dropper to add 7. Close the sample cell. 8. Start the instrument
Sulfide 2 Reagent to each Invert the sample cell to mix. timer. A five-minute reaction
cell. Use 0.5 mL for A pink color will develop time starts.
spectrophotometers and initially. If sulfide is present,
1.0 mL for colorimeters. the solution becomes blue.

Zero

9. When the timer expires, 10. Insert the blank into the 11. Push ZERO. The 12. Clean the prepared
clean the blank. cell holder. display shows 0 µg/L or sample.
0.00 mg/L S2–.

Read

13. Insert the prepared 14. Push READ. Results

sample into the cell holder. show in µg/L or mg/L S2–.

Soluble sulfides
To measure soluble sulfides, use a centrifuge to separate the solids. To make an
estimate of the amount of insoluble sulfides in the sample, subtract the soluble sulfide
concentration from the total (with solids) sulfide concentration.

1. Fill a centrifuge tube completely with sample and immediately cap the tube.
2. Put the tube in a centrifuge and run the centrifuge to separate the solids.
3. Use the supernatent as the sample in the test procedure.

Sulfide, Methylene Blue Method (800 µg/L) 3

Interferences
Interfering Interference level
substance
Barium Concentrations more than 20 mg/L react with the sulfuric acid in Sulfide 1 Reagent and form a BaSO4
(barite) precipitate. To correct for this interference:

1. Dilute the sample in the test procedure as follows:

• Spectrophotometers: use a 0.1-mL or 1.0-mL sample volume and add deionized water to the 10-
mL mark.
• Colorimeters: use a 0.25-mL or 2.5-mL sample volume and add deionized water to the 25-mL
mark.
2. Let the sample fully react with both reagents.
3. After the 5 minute reaction period, pour the sample into a 50-mL beaker.
4. Pull the sample into a Luer-Lock syringe (10 cc for spectrophotometers; 60 cc for colorimeters).
5. Put a 0.45-μm filter disc on the Luer-Lock tip and filter the sample into a clean sample cell for
measurement. Use deionized water to prepare the blank.

Strong reducing Prevent the full color development or reduce the blue color
substances such
as sulfite,
thiosulfate and
hydrosulfite
Sulfide, high High concentrations of sulfide can inhibit the full color development. Use a diluted sample in the test
levels procedure. Some sulfide loss can occur when the sample is diluted.
Turbidity Pre-treat the sample to remove sulfide, then use the pre-treated sample as the blank in the test procedure.
Prepare a sulfide-free blank as follows:

1. Measure 25 mL of sample into a 50-mL Erlenmeyer flask.

2. Add 30-g/L Bromine Water by drops with constant swirling until a yellow color remains.
3. Add 30-g/L Phenol Solution by drops with constant swirling until the yellow color is removed.
4. Use this solution to replace the deionized water blank in the test procedure.

Method performance
The method performance data that follows was derived from laboratory tests that were
measured on a spectrophotometer during ideal test conditions. Users may get different
results under different test conditions.
Program Standard Precision (95% Confidence Interval) Sensitivity
Concentration change per 0.010 Abs change
690 520 µg/L S2– 504–536 µg/L S2– 5 µg/L S2–

Summary of method
Hydrogen sulfide and acid-soluble metal sulfides react with N,N-dimethyl-p-
phenylenediamine sulfate to form methylene blue. The intensity of the blue color is
proportional to the sulfide concentration. High sulfide levels in oil field waters may be
determined after proper dilution. The measurement wavelength is 665 nm for
spectrophotometers or 610 nm for colorimeters.
Pollution prevention and waste management
Reacted samples contain hexavalent chromium and must be disposed of as a hazardous
waste. Dispose of reacted solutions according to local, state and federal regulations.

4 Sulfide, Methylene Blue Method (800 µg/L)

Consumables and replacement items
Required reagents

Description Quantity/test Unit Item no.

Water, deionized varies 4L 27256

Sulfide Reagent Set — — 2244500
Includes:
100 mL
Sulfide 1 Reagent 1–2 mL 181632
MDB
100 mL
Sulfide 2 Reagent 1 –2 mL 181732
MDB

Required apparatus

Description Quantity/test Unit Item no.

Pipet, serological, graduated, 10-mL 1 each 53238

Pipet filler, safety bulb 1 each 1465100
Stoppers for 18-mm tubes and AccuVac Ampuls 2 6/pkg 173106

Optional reagents and apparatus

Description Unit Item no.

Bromine Water, 30-g/L 29 mL 221120

Phenol Solution, 30-g/L 29 mL 211220
Stoppers for 18-mm tube 25/pkg 173125
Flask, Erlenmeyer, 50 mL each 50541

Sulfide, Methylene Blue Method (800 µg/L) 5

 FOR TECHNICAL ASSISTANCE, PRICE INFORMATION AND ORDERING: HACH COMPANY
In the U.S.A. – Call toll-free 800-227-4224 WORLD HEADQUARTERS
Outside the U.S.A. – Contact the HACH office or distributor serving you. Telephone: (970) 669-3050
On the Worldwide Web – www.hach.com; E-mail – [email protected] FAX: (970) 669-2932

© Hach Company/Hach Lange GmbH, 1989–2013. All rights reserved. 04/2013, Edition 8
Sulfide, HR DOC316.53.01319

Methylene Blue Method1 Method 10253

0.01 to 0.70, 0.1 to 7.0, 1 to 70 mg/L S2– Reagent Solution

Scope and application: For oil and gas field waters.
1 Adapted from Standard Methods for the Examination of Water and Wastewater.

Test preparation

Instrument specific information

The table in this section shows all of the instruments that have the program for this test.
Table 1 shows sample cell and orientation requirements for reagent addition tests, such
as powder pillow or bulk reagent tests.
To use the table, select an instrument, then read across to find the corresponding
information for this test.
Table 1 Instrument-specific information for reagent solution
Instrument Sample cell orientation Sample cell
DR 6000 The fill line is to the right. 2495402
DR 3800
DR 2800
DR 2700
DR 5000 The fill line is toward the user.
DR 3900
DR 900 The orientation mark is toward the user. 2401906

Before starting
Samples must be analyzed immediately after collection and cannot be preserved for later analysis.
Install the instrument cap on the DR 900 cell holder before ZERO or READ is pushed.
Some sulfide loss can occur if dilution is necessary.
Review the Safety Data Sheets (MSDS/SDS) for the chemicals that are used and use any recommended personal protective
equipment.
Dispose of reacted solutions according to local, state and federal regulations. Use the Safety Data Sheets for disposal
information for unused reagents. Consult the environmental, health and safety staff for your facility and/or local regulatory
agencies for further disposal information.

Items to collect
Description Quantity

Sulfide 1 Reagent 1–2 mL

Sulfide 2 Reagent 1–2 mL
Pipet or mechanical pipettor (appropriate sample and reagent size) 1

1
Items to collect (continued)
Description Quantity

Sample cells (For information about sample cells, adapters or light shields, refer to Instrument
2
specific information on page 1.)
Stoppers 2
Water, deionized 10 mL

Refer to Consumables and replacement items on page 5 for reorder information.

Sample collection
• Samples must be analyzed immediately after collection and cannot be preserved for
later analysis.
• Collect samples in clean glass or plastic bottles with tight-fitting caps. Fill the bottle
completely and immediately tighten the cap.
• Prevent agitation of the sample or exposure to air.

Methylene Blue method

Start

1. Start program 2. Prepare the blank: Fill a 3. Prepare the sample: 4. Spectrophotometers:
691 Sulfide HR. For sample cell with deionized Add the sample volume that Add deionized water to the
information about sample water. Use 10 mL for is specified for the test 10-mL line. Colorimeters:
cells, adapters or light spectrophotometers and range to a clean sample Add deionized water to the
shields, refer to Instrument 25 mL for colorimeters. cell. Refer to Table 2 25-mL line.
specific information on page 3. To prevent sulfide loss, do
on page 1. Use a pipet to measure not mix the sample more
Note: Although the program small volumes. than necessary.
name may vary between
instruments, the program
number does not change.

2 Sulfide, Methylene Blue Method (multi-range: 0.80, 8.00, 80.00 mg/L)

5. Add Sulfide 1 Reagent to 6. Swirl to mix. 7. Add Sulfide 2 Reagent to 8. Close both sample cells
each sample cell. Use each sample cell. Use with a stopper. Invert to mix.
0.5 mL of reagent for 0.5 mL of reagent for The solution shows pink and
spectrophotometers. Use spectrophotometers. Use then blue if sulfide is in the
1.0 mL of reagent for 1.0 mL of reagent for sample.
colorimeters. colorimeters.

Zero

9. Start the instrument 10. When the timer expires, 11. Insert the blank into the 12. Push ZERO. The
timer. A 5-minute reaction clean the blank. cell holder. display shows 0 mg/L S2–.
time starts.

Read

13. Clean the prepared 14. Insert the prepared 15. Push READ. Results
sample. sample into the cell holder. show in mg/L S2–.

Select a sample volume

Table 2 Sample volumes and ranges
Range Spectrophotometer volume Colorimeter volume
0.01–0.70 mg/L (LR) 10 mL 25 mL
0.1–7.0 mg/L (MR) 1.0 mL 2.5 mL
1–70 mg/L (HR) 0.1 mL 0.25 mL

Set the dilution factor

Instruments that have a dilution factor option can include the dilution factor in the result
and show the concentration of the original, undiluted sample. For example, if the sample

Sulfide, Methylene Blue Method (multi-range: 0.80, 8.00, 80.00 mg/L) 3

 is diluted by a factor of 10, the instrument multiplies the result by 10 and shows the
calculated result in the instrument display.

1. Select Options>More>Dilution factor from the instrument menu.

Note: Colorimeters include a dilution factor when the chemical form is set. Go to
Options>Advanced Options>Chemical Form and select LR, MR or HR.
2. Enter the dilution factor:
• 1 mL sample diluted to 10 mL: dilution factor is 10.
• 0.1 mL sample diluted to 10 mL: dilution factor is 100.
3. Push OK to confirm. Push OK again.
4. Push RETURN to go back to the measurement screen.

Soluble sulfides
To measure soluble sulfides, use a centrifuge to separate the solids. To make an
estimate of the amount of insoluble sulfides in the sample, subtract the soluble sulfide
concentration from the total (with solids) sulfide concentration.

1. Fill a centrifuge tube completely with sample and immediately cap the tube.
2. Put the tube in a centrifuge and run the centrifuge to separate the solids.
3. Use the supernatent as the sample in the test procedure.

Interferences
Interfering Interference level
substance
Barium Concentrations more than 20 mg/L react with the sulfuric acid in Sulfide 1 Reagent and form a BaSO4
(barite) precipitate. To correct for this interference:

1. Dilute the sample in the test procedure as follows:

• Spectrophotometers: use a 0.1-mL or 1.0-mL sample volume and add deionized water to the 10-
mL mark.
• Colorimeters: use a 0.25-mL or 2.5-mL sample volume and add deionized water to the 25-mL
mark.
2. Let the sample fully react with both reagents.
3. After the 5 minute reaction period, pour the sample into a 50-mL beaker.
4. Pull the sample into a Luer-Lock syringe (10 cc for spectrophotometers; 60 cc for colorimeters).
5. Put a 0.45-μm filter disc on the Luer-Lock tip and filter the sample into a clean sample cell for
measurement. Use deionized water to prepare the blank.

Strong reducing Prevent the full color development or reduce the blue color
substances such
as sulfite,
thiosulfate and
hydrosulfite
Sulfide, high High concentrations of sulfide can inhibit the full color development. Use a diluted sample in the test
levels procedure. Some sulfide loss can occur when the sample is diluted.
Turbidity Pre-treat the sample to remove sulfide, then use the pre-treated sample as the blank in the test procedure.
Prepare a sulfide-free blank as follows:

1. Measure 25 mL of sample into a 50-mL Erlenmeyer flask.

2. Add 30-g/L Bromine Water by drops with constant swirling until a yellow color remains.
3. Add 30-g/L Phenol Solution by drops with constant swirling until the yellow color is removed.
4. Use this solution to replace the deionized water blank in the test procedure.

4 Sulfide, Methylene Blue Method (multi-range: 0.80, 8.00, 80.00 mg/L)

Accuracy check
Standard solution method
Sulfide standard solutions are not stable and must be prepared by the user. Refer to
Standard Methods, 4500S2– for preparation and standardization instructions.
Method performance
The method performance data that follows was derived from laboratory tests that were
measured on a spectrophotometer during ideal test conditions. Users may get different
results under different test conditions.
Program Standard Precision (95% Confidence Interval) Sensitivity
Concentration change per 0.010 Abs change
691 0.52 mg/L S2– 0.50–0.54 µg/L S2– 0.005 mg/L S2–

Summary of method
Hydrogen sulfide and acid-soluble metal sulfides react with N,N-dimethyl-p-
phenylenediamine sulfate to form methylene blue. The intensity of the blue color is
proportional to the sulfide concentration. High sulfide levels in oil field waters may be
determined after proper dilution. The measurement wavelength is 665 nm for
spectrophotometers or 610 nm for colorimeters.
Pollution prevention and waste management
Reacted samples contain hexavalent chromium and must be disposed of as a hazardous
waste. Dispose of reacted solutions according to local, state and federal regulations.
Consumables and replacement items
Required reagents

Description Quantity/test Unit Item no.

Water, deionized varies 4L 27256

Sulfide Reagent Set — — 2244500
Includes:
100 mL
Sulfide 1 Reagent 1–2 mL 181632
MDB
100 mL
Sulfide 2 Reagent 1–2 mL 181732
MDB

Required apparatus

Description Quantity/test Unit Item no.

®
Pipet, TenSette , 0.1- to 1.0-mL 1 each 1970001
Pipet Tips, for TenSette Pipet 1970001 2 50/pkg 2185696
Pipet, TenSette 1.0–10.0 mL 1 each 1970010
Pipet Tips, for TenSette Pipet 1970010 varies 50/pkg 2199796
Pipet, 0.2 –1.0 mL 1 each BBP078
Pipet Tip for BBP078 2 100/pkg BBP079
Pipet, 1.0–5.0 mL 1 each BBP065
Pipet Tip for BBP065 1 75/pkg BBP068

Sulfide, Methylene Blue Method (multi-range: 0.80, 8.00, 80.00 mg/L) 5

Optional reagents and apparatus

Description Unit Item no.

Beaker, 50-mL each 50041H

Bromine Water, 30-g/L 29 mL 221120
Cylinder, graduated mixing, 10 mL 1 2088638
Flask, Erlenmeyer, 50 mL each 50541
Phenol Solution, 30-g/L 29 mL 211220
Pipet, serological, 10-mL 1 53238
Pipet filler, safety bulb each 1465100
Syringe, 10 cc, Luer-Lock tip 1 2202400
Syringe, 60 cc, Luer-Lock tip 1 2258700
Syringe filter, 0.45 µm, 33 mm PVDF 50/pkg 2513603

FOR TECHNICAL ASSISTANCE, PRICE INFORMATION AND ORDERING: HACH COMPANY

In the U.S.A. – Call toll-free 800-227-4224 WORLD HEADQUARTERS
Outside the U.S.A. – Contact the HACH office or distributor serving you. Telephone: (970) 669-3050
On the Worldwide Web – www.hach.com; E-mail – [email protected] FAX: (970) 669-2932

© Hach Company/Hach Lange GmbH, 1989–2013. All rights reserved. 04/2013, Edition 8
Surfactants, Anionic (Detergents) DOC316.53.01138

Crystal Violet Method1 Method 8028

0.002 to 0.275 mg/L as LAS (spectrophotometers)
0.020 to 0.300 mg/L as LAS (colorimeters)
Scope and application: For water, wastewater and seawater.
1 Adapted from Analytical Chemistry, 38, 791 (1966).

Test preparation

Instrument-specific information
The table in this section shows all of the instruments that have the program for this test.
Instrument specific information PP shows sample cell and orientation requirements for
reagent addition tests, such as powder pillow or bulk reagent tests.
To use the table, select an instrument, then read across to find the corresponding
information for this test.
Table 1 Instrument-specific information
Instrument Sample cell orientation Sample cell
DR 6000 The fill line is to the right. 2612602
DR 3800
DR 2800
DR 2700
DR 5000 The fill line is toward the user.
DR 3900
DR 900 The orientation mark is toward the user. 2401906

Before starting
Install the instrument cap on the DR 900 cell holder before ZERO or READ is pushed.
This procedure must be done in a well-ventilated area or fume hood.
Acetone can be used to clean benzene from glassware.
To prevent the formation of water droplets in the sample cells, use only dry sample cells and discard the first few mL of
benzene. Additionally, it can help to transfer the liquid from the funnel to a sample cell, let it sit for a few seconds and decant
to a second cell for the measurement.
Excessive shaking can cause an emulsion to form, which makes the phases separate more slowly. If this occurs, remove
most of the water layer, then gently mix the contents of the funnel with a clean Teflon®-coated rod or other inert tool.
Spilled reagent will affect test accuracy and is hazardous to the skin and other materials.
For best results, measure the reagent blank value for each new lot of reagent. Replace the sample with deionized water in
the test procedure to get the reagent blank value. Subtract the reagent blank value from the sample results automatically
with the reagent blank adjust option.
In bright light conditions (e.g., direct sunlight), close the cell compartment on spectrophotometers, if applicable, with the
protective cover during measurements.
Do not use the Pour-Thru Cell or sipper module (for applicable instruments) with this test.

1
Review the Safety Data Sheets (MSDS/SDS) for the chemicals that are used and use any recommended personal protective
equipment.
Dispose of reacted solutions according to local, state and federal regulations. Use the Safety Data Sheets for disposal
information for unused reagents. Consult the environmental, health and safety staff for your facility and/or local regulatory
agencies for further disposal information.

Items to collect
Description Quantity

Benzene, ACS 40 mL
Buffer Solution, sulfate-type 10 mL
Detergent Reagent Powder Pillows 1
Clippers (for powder pillows) 1
Cylinder, graduated, 25-mL 1
Cylinder, graduated, 50-mL 1
Cylinder, graduated, 500-mL 1
Funnel, separatory, 500-mL 1
Sample cells (For information about sample cells, adapters or light shields, refer to Instrument-
2
specific information on page 1.)
Support Ring, 4-inch 1
Support Ring Stand, 5 x 8 inch base 1

Refer to Consumables and replacement items on page 6 for reorder information.

Sample collection and storage

• Collect samples in clean glass or plastic bottles.
• Analyze the samples as soon as possible for best results.
• To preserve samples for later analysis, keep the samples at or below 6 °C (43 °F) for
up to 48 hours.
• Let the sample temperature increase to room temperature before analysis.

2 Surfactants, Anionic (Detergents), Crystal Violet Method (0.275 mg/L)

Powder pillow procedure

Start

1. Start program 2. Fill a clean 500-mL 3. Pour the sample from the 4. Add 10 mL of Sulfate
710 Surfactants. For graduated cylinder to the cylinder to a clean 500-mL Buffer Solution.
information about sample 300-mL mark with sample. separatory funnel.
cells, adapters or light
shields, refer to Instrument-
specific information
on page 1.
Note: Although the program
name may vary between
instruments, the program
number does not change.

5. Close the funnel. Shake 6. Add the contents of one 7. Close the funnel and 8. Add 30 mL of benzene to
the funnel for 5 seconds Detergent Reagent Powder shake until the powder the funnel.
Pillow to the funnel. dissolves completely. The
powder will dissolve slowly.

9. Close the funnel and 10. Put the funnel in the 11. Start the instrument 12. When the timer expires,
shake gently for one minute. support stand. timer. A 30-minute reaction remove the stopper and
time starts. drain the bottom water layer.
Collect this water for safe
disposal.

Surfactants, Anionic (Detergents), Crystal Violet Method (0.275 mg/L) 3

13. Prepare the sample: 14. Prepare the blank: Fill 15. Clean the blank. 16. Insert the blank into the
Drain the top benzene layer another sample cell to the cell holder.
into a clean sample cell. 10-mL mark with pure
Close the sample cell. benzene. Close the sample
Do not filter the benzene cell.
layer before color
measurement. Filtration
removes the blue color.

Zero Read

17. Push ZERO. The 18. Clean the prepared 19. Insert the prepared 20. Push READ. Results
display shows 0.000 mg/L sample. sample into the cell holder. show in mg/L LAS.
LAS.

Interferences
Interfering substance Interference level
Chloride High amounts of chloride, such as in brines and seawater, cause low results.
Perchlorate ions Interfere at all levels
Periodate ions Interfere at all levels

Accuracy check
Standard additions method (sample spike)
Use the standard additions method (for applicable instruments) to validate the test
procedure, reagents and instrument and to find if there is an interference in the sample.
Items to collect:
• Detergent Voluette® Ampule Standard, 60-mg/L LAS
• Ampule breaker
• Pipet, TenSette®, 0.1–1.0 mL and tips

1. Use the test procedure to measure the concentration of the sample, then keep the
(unspiked) sample in the instrument.
2. Go to the Standard Additions option in the instrument menu.
3. Select the values for standard concentration, sample volume and spike volumes.
4. Open the standard solution.

4 Surfactants, Anionic (Detergents), Crystal Violet Method (0.275 mg/L)

 5. Prepare three spiked samples: use the TenSette pipet to add 0.1 mL, 0.2 mL and
0.3 mL of the standard solution, respectively, to three 300-mL portions of fresh
sample. Mix well.
6. Use the test procedure to measure the concentration of each of the spiked samples.
Start with the smallest sample spike. Measure each of the spiked samples in the
instrument.
7. Select Graph to compare the expected results to the actual results.
Note: If the actual results are significantly different from the expected results, make sure that
the sample volumes and sample spikes are measured accurately. The sample volumes and
sample spikes that are used should agree with the selections in the standard additions menu. If
the results are not within acceptable limits, the sample may contain an interference.

Standard solution method

Use the standard solution method to validate the test procedure, reagents and
instrument.
Items to collect:
• Detergent Voluette® Ampule Standard, 60 mg/L LAS
• 1-L volumetric flask, Class A
• 3-mL volumetric pipet, Class A and pipet filler
• Deionized water

1. Prepare a 0.180 mg/L LAS standard solution as follows:

a. Use a pipet to add 3.0 mL of 60 mg/L LAS standard solution into the volumetric
flask.
b. Dilute to the mark with deionized water. Mix well. Prepare this solution daily.
2. Use the test procedure to measure the concentration of the prepared standard
solution.
3. Compare the expected result to the actual result.
Note: The factory calibration can be adjusted slightly with the standard adjust option so that the
instrument shows the expected value of the standard solution. The adjusted calibration is then
used for all test results. This adjustment can increase the test accuracy when there are slight
variations in the reagents or instruments.

Method performance
The method performance data that follows was derived from laboratory tests that were
measured on a spectrophotometer during ideal test conditions. Users may get different
results under different test conditions.
Program Standard Precision (95% Confidence Interval) Sensitivity
Concentration change per 0.010 Abs change
710 0.180 mg/L LAS 0.172–0.188 mg/L LAS 0.002 mg/L LAS

Summary of method
Detergents, ABS (alkyl benzene sulfonate) or LAS (linear alkylate sulfonate) are
determined by association with crystal violet dye and extraction of the ion-pair complex
into benzene. The measurement wavelength is 605 nm for spectrophotometers or 610 nm
for colorimeters.
Pollution prevention and waste management
Reacted samples contain benzene and must be disposed of as a hazardous waste.
Dispose of reacted solutions according to local, state and federal regulations.

Surfactants, Anionic (Detergents), Crystal Violet Method (0.275 mg/L) 5

Consumables and replacement items
Required reagents

Description Quantity/test Unit Item no.

Detergents Reagent Set — — 2446800

Includes:
Benzene, ACS 40 mL 4 liters 1444017
Buffer Solution, sulfate-type 10 mL 500 mL 45249
Detergent Reagent Powder Pillows 1 pillow 25/pkg 100868

Required apparatus

Description Quantity/test Unit Item no.

Clippers for solution pillows 1 each 96800

Cylinder, graduated, 25-mL. 1 each 50840
Cylinder, graduated , 50-mL 1 each 50841
Cylinder, graduated, 500-mL 1 each 50849
Funnel, separatory, 500-mL 1 each 52049
Support Ring, 4-inch 1 each 58001
Support, Ring Stand, 5 x 8 inch base 1 each 56300

Recommended standards

Description Unit Item no.

®
Detergent Standard Solution, 10-mL Voluette Ampule, 60-mg/L LAS 16/pkg 1427110

Optional reagents and apparatus

Description Unit Item no.

Acetone, ACS 500 mL 1442949

Beaker, 600 mL each 50052
Flask, volumetric, Class A, 1000-mL each 1457453
Pipet filler, safety bulb each 1465100
®
Pipet, TenSette , 0.1–1.0 mL each 1970001
Pipet tips for TenSette Pipet 1970001 50/pkg 2185696
Pipet, volumetric, Class A, 3-mL each 1451503

FOR TECHNICAL ASSISTANCE, PRICE INFORMATION AND ORDERING: HACH COMPANY

In the U.S.A. – Call toll-free 800-227-4224 WORLD HEADQUARTERS
Outside the U.S.A. – Contact the HACH office or distributor serving you. Telephone: (970) 669-3050
On the Worldwide Web – www.hach.com; E-mail – [email protected] FAX: (970) 669-2932

© Hach Company/Hach Lange GmbH, 1989–2013. All rights reserved. 04/2013, Edition 8
Suspended Solids DOC316.53.01139

Photometric Method1 Method 8006

5 to 750 mg/L TSS
Scope and application: For water and wastewater.
1 Adapted from Sewage and Industrial Wastes, 31, 1159 (1959).

Test preparation

Instrument specific information

The table in this section shows all of the instruments that have the program for this test.
Table 1 shows sample cell and orientation requirements for reagent addition tests, such
as powder pillow or bulk reagent tests.
To use the table, select an instrument, then read across to find the corresponding
information for this test.
Table 1 Instrument-specific information for reagent addition
Instrument Sample cell orientation Sample cell
DR 6000 The fill line is to the right. 2495402
DR 3800
DR 2800
DR 2700
DR 5000 The fill line is toward the user.
DR 3900
DR 900 The orientation mark is toward the user. 2401906

Before starting
Install the instrument cap on the DR 900 cell holder before ZERO or READ is pushed.
Do not use the Pour-Thru Cell or sipper module (for applicable instruments) with this test.

Items to collect
Description Quantity

Beaker, 600-mL, polypropylene 1

Blender 1
Cylinder, 500-mL polypropylene, graduated 1
Sample cells (For information about sample cells, adapters or light shields, refer to Instrument
2
specific information on page 1.)

Refer to Consumables and replacement items on page 3 for reorder information.

Sample collection and storage

• Collect samples in clean glass or plastic bottles.

1
 • To preserve samples for later analysis, keep the samples at or below 6 °C (43 °F) for
up to 7 days.
• Let the sample temperature increase to room temperature before analysis.

Photometric procedure

Start

1. Start program 2. Blend 500 mL of sample 3. Pour the blended sample 4. Prepare the sample:
630 Suspended Solids. For in a blender at high speed into a 600-ml beaker. Stir the sample and
information about sample for exactly two minutes. immediately pour 10 mL of
cells, adapters or light the blended sample into a
shields, refer to Instrument sample cell.
specific information
on page 1.
Note: Although the program
name may vary between
instruments, the program
number does not change.

Zero

5. Prepare the blank: Fill a 6. Clean the blank. 7. Insert the blank into the 8. Push ZERO. The display
second sample cell with cell holder. shows 0 mg/L TSS.
10 mL of tap water or
deionized water.

Read

9. Swirl the prepared 10. Clean the prepared 11. Insert the prepared 12. Push READ. Results
sample to remove any gas sample. sample into the cell holder. show in mg/L TSS.
bubbles and uniformly
suspend any residue.

2 Suspended Solids, Photometric Method (750 mg/L)

Interferences
Samples that absorb strongly at the measurement wavelength, such as blue dyes, may
give false, high-bias readings. A user-entered calibration is advised for these samples.
Accuracy check
Standard solution method
Calibration for this test is based on the gravimetric technique with parallel sewage
samples from a municipal sewage plant. For most samples, this calibration supplies
satisfactory results. When higher accuracy is required, run parallel spectrophotometric
and gravimetric determinations with portions of the same sample. Make the new
calibration on the particular sample using a gravimetric technique as a basis.
Summary of method
This method of determining total suspended solids (TSS) is a simple, direct measurement
which does not require the filtration or ignition/weighing steps that gravimetric procedures
do. The USEPA specifies the gravimetric method for solids determinations, while this
method is often used for checking in-plant processes. The measurement wavelength is
810 nm for spectrophotometers or 610 nm for colorimeters.
Consumables and replacement items
Required apparatus

Description Quantity/test Unit Item no.

Beaker, 600-mL, polypropylene 1 each 108052

Blender, 2-speed, 120 VAC 1 each 2616100
Blender, 2-speed, 240 VAC 1 each 2616102
Cylinder, graduated, 500-mL, polypropylene 1 each 108149

Suspended Solids, Photometric Method (750 mg/L) 3

 FOR TECHNICAL ASSISTANCE, PRICE INFORMATION AND ORDERING: HACH COMPANY
In the U.S.A. – Call toll-free 800-227-4224 WORLD HEADQUARTERS
Outside the U.S.A. – Contact the HACH office or distributor serving you. Telephone: (970) 669-3050
On the Worldwide Web – www.hach.com; E-mail – [email protected] FAX: (970) 669-2932

© Hach Company/Hach Lange GmbH, 1989–2013. All rights reserved. 04/2013, Edition 8
Tannin and Lignin DOC316.53.01140

Tyrosine Method1 Method 8193

0.1 to 9.0 mg/L Tannins (as Tannic Acid) Reagent Solution
Scope and application: For water, wastewater and boiler water.
1 Adapted from Kloster, M.B., Journal American Water Works Association, Vol. 66, No. 1, p. 44 (1974).

Test preparation

Instrument specific information

The table in this section shows all of the instruments that have the program for this test.
Table 1 shows sample cell and orientation requirements for reagent addition tests, such
as powder pillow or bulk reagent tests.
To use the table, select an instrument, then read across to find the corresponding
information for this test.
Table 1 Instrument-specific information for reagent addition
Instrument Sample cell orientation Sample cell
DR 6000 The fill line is to the right. 2495402
DR 3800
DR 2800
DR 2700
DR 5000 The fill line is toward the user.
DR 3900
DR 900 The orientation mark is toward the user. 2401906

Before starting
Install the instrument cap on the DR 900 cell holder before ZERO or READ is pushed.
Measure the volume of the reagent accurately. Use a pipet if possible.
Filter samples that are turbid with filter paper and a funnel. The test results are then mg/L soluble tannic acid.
The Pour-Thru Cell can be used (for applicable instruments) if rinsed well with deionized water between the blank and the
prepared samples.
Review the Safety Data Sheets (MSDS/SDS) for the chemicals that are used and use any recommended personal protective
equipment.
Dispose of reacted solutions according to local, state and federal regulations. Use the Safety Data Sheets for disposal
information for unused reagents. Consult the environmental, health and safety staff for your facility and/or local regulatory
agencies for further disposal information.

Items to collect
Description Quantity

Tannin and Lignin Reagent Set:

Sodium Carbonate Solution 10 mL

1
Items to collect (continued)
Description Quantity

TanniVer™ 3 Tannin-Lignin Reagent 1 mL

Cylinder, graduated mixing, 25-mL 2
Pipet Filler 1
Pipet, volumetric Class A, 5.0-mL 1
Pipet, volumetric Class A, 0.5-mL 1
Sample cells (For information about sample cells, adapters or light shields, refer to Instrument
2
specific information on page 1.)
Water, deionized 25 mL

Refer to Consumables and replacement items on page 4 for reorder information.

Sample collection
• Collect samples in clean glass or plastic bottles.

Tyrosine procedure

Start

1. Start program 2. Prepare the blank: 3. Prepare the sample: 4. Pipet 0.5 mL of
720 Tannin & Lignin. For Fill a 25-mL mixing cylinder Fill a second 25-mL mixing TanniVer™ 3 Tannin-Lignin
information about sample to the 25-mL mark with cylinder to the 25-mL mark Reagent into each cylinder.
cells, adapters or light deionized water. with sample.
shields, refer to Instrument
specific information
on page 1.
Note: Although the program
name may vary between
instruments, the program
number does not change.

2 Tannin and Lignin, Tyrosine Method (9.0 mg/L)

5. Close the two cylinders. 6. Add 5.0 mL of Sodium 7. Close the two cylinders. 8. Pour 10 mL of each
Invert both cylinders several Carbonate Solution into Invert both cylinders several solution into two sample
times to mix. each cylinder. times to mix. cells.
A blue color will develop if
tannins and/or lignins are
present.

Zero

9. Start the instrument 10. When the timer expires, 11. Insert the blank into the 12. Push ZERO. The
timer. A 25-minute reaction clean the blank. cell holder. display shows 0.0 mg/L
time starts. Tannins (as Tannic Acid).

Read

13. Clean the prepared 14. Insert the prepared 15. Push READ. Results
sample. sample into the cell holder. show in mg/L Tannins (as
Tannic Acid).

Interferences
Interfering Interference level
substance
Ferrous iron Causes a positive interference. (2 mg/L of ferrous iron produces a color equivalent to about 1 mg/L of
tannic acid.) To remove interference of ferrous iron up to 20 mg/L, add one 0.2 g scoop of Sodium
Pyrophosphate to the sample before the test.
Sulfite To remove sulfite interference, add 1 mL of formaldehyde1 to the sample before the sample test.
1 Refer to Consumables and replacement items on page 4 for reorder information.

Tannin and Lignin, Tyrosine Method (9.0 mg/L) 3

Accuracy check
Standard solution method
Use the standard solution method to validate the test procedure, reagents and
instrument.
Items to collect:
• 0.200 g tannic acid, analytical grade
• 1-L volumetric flask, Class A
• 500-mL volumetric flask, Class A
• 15-mL volumetric pipet, Class A and pipet filler
• Deionized water

1. Prepare a 200-mg/L tannic acid stock solution as follows:

a. Add 0.200 g of tannic acid into a 1-L volumetric flask.
b. Dilute to the mark with deionized water. Mix well. Prepare the stock solution each
month.
2. Prepare a 6.0 mg/L tannic acid standard solution as follows:
a. Use a pipet to add 15.00 mL of the 200-mg/L tannic acid stock solution into a
500-mL volumetric flask.
b. Dilute to the mark with deionized water. Mix well. Prepare the standard solution
each day.
3. Use the test procedure to measure the concentration of the prepared standard
solution.
4. Compare the expected result to the actual result.
Note: The factory calibration can be adjusted slightly with the standard adjust option so that the
instrument shows the expected value of the standard solution. The adjusted calibration is then
used for all test results. This adjustment can increase the test accuracy when there are slight
variations in the reagents or instruments.

Method performance
The method performance data that follows was derived from laboratory tests that were
measured on a spectrophotometer during ideal test conditions. Users may get different
results under different test conditions.
Program Standard Precision (95% Confidence Interval) Sensitivity
Concentration change per 0.010 Abs change
720 6.0 mg/L tannic acid 5.8–6.2 mg/L tannic acid 0.1 mg/L tannic acid

Summary of method
This test measures all hydroxylated aromatic compounds, including tannin, lignin, phenol
and cresol. This method produces a blue color proportional to the amount of these
compounds in the sample. The results are reported as total tannin and lignin and
expressed as mg/L tannic acid. The measurement wavelength is 700 nm for
spectrophotometers or 610 nm for colorimeters.
Consumables and replacement items
Required reagents

Description Quantity/test Unit Item no.

Water, deionized varies 4L 27256

Up to
Tannin and Lignin Reagent Set 1 2244600
100 tests

4 Tannin and Lignin, Tyrosine Method (9.0 mg/L)

Consumables and replacement items (continued)
Description Quantity/test Unit Item no.

Includes:
Sodium Carbonate Solution 10 mL 500 mL 67549
TanniVer™ 3 Tannin-Lignin Reagent 1 mL 100 mL 256032

Required apparatus

Description Quantity/test Unit Item no.

Cylinder, graduated mixing, 25 mL with stopper 2 each 2088640

Pipet filler, safety bulb 1 each 1465100
Pipet, volumetric, Class A, 5.00-mL 1 each 1451537
Pipet, volumetric, Class A, 0.5-mL 1 each 1451534

Recommended standards

Description Unit Item no.

Tannic Acid, Analytical Grade 113 g 79114

Optional reagents and apparatus

Description Unit Item no.

Flask, volumetric, Class A, 1000-mL each 1457453

Flask, volumetric, Class A, 500-mL each 1457449
100 mL
Formaldehyde, ACS 205932
MDB
Pipet, volumetric Class A, 15-mL each 1451539
Sodium Pyrophosphate 50 g 1429525
®
Pipet, TenSette , 0.1–1.0 mL each 1970001
Pipet tips for TenSette Pipet 1970001 1000/pkg 2185628
Balance, analytical, 80 g x 0.1 mg 100-240 VAC each 2936701
Paper, for weighing, 100 x 100 mm 500/pkg 1473885
Spatula, micro each 1225600

Tannin and Lignin, Tyrosine Method (9.0 mg/L) 5

 FOR TECHNICAL ASSISTANCE, PRICE INFORMATION AND ORDERING: HACH COMPANY
In the U.S.A. – Call toll-free 800-227-4224 WORLD HEADQUARTERS
Outside the U.S.A. – Contact the HACH office or distributor serving you. Telephone: (970) 669-3050
On the Worldwide Web – www.hach.com; E-mail – [email protected] FAX: (970) 669-2932

© Hach Company/Hach Lange GmbH, 1989–2013. All rights reserved. 04/2013, Edition 8
Toxicity DOC316.53.01141

ToxTrak™ Method1, 2 Method 10017

0 to 100% inhibition Test ‘N Tube™ Vials
Scope and application: For drinking water, wastewater and natural waters.
1 Liu, D., Bull. Environ. Contm. Toxicol. 26, 145-149 (1981)
2 Environmental Technology Verification ETV Program evaluated, November, 2003

Test preparation

Instrument specific information

The table in this section shows all of the instruments that have the program for this test.
Table 1 shows adapter and light shield requirements for the instruments that use them.
To use the table, select an instrument, then read across to find the corresponding
information for this test.
Table 1 Instrument-specific information for test tubes
Instrument Adapters Light shield
DR 6000 — —
DR 5000 — —
DR 900 4846400 Cover supplied with the instrument
DR 3900 — LZV849
DR 3800 — LZV646
DR 2800 —

Before starting
Install the instrument cap on the DR 900 cell holder before ZERO or READ is pushed.
DR 3900, DR 3800, DR 2800 and DR 2700: Install the light shield in Cell Compartment #2 before this test is started.
Do not leave the tubes in the instrument during incubation. Make sure that all samples and control cells have similar
conditions of temperature and light during the reaction.
If the samples contain chlorine, add two drops of sodium thiosulfate to each blank and sample before the test is started.
Review the Safety Data Sheets (MSDS/SDS) for the chemicals that are used and use any recommended personal protective
equipment.
Dispose of reacted solutions according to local, state and federal regulations. Use the Safety Data Sheets for disposal
information for unused reagents. Consult the environmental, health and safety staff for your facility and/or local regulatory
agencies for further disposal information.

Items to collect
Description Quantity

Bacterial Count Broth Tube 1 tube

Pipet, transfer, sterile 2
Test 'N Tube, with cap 1
Sodium Thiosulfate varies
ToxTrak™ Reagent Powder Pillows 2
ToxTrak™ Accelerator Solution 4 drops

1
Items to collect (continued)
Description Quantity

Water, deionized varies

Clippers 1
Incubator 1
Pipet, volumetric, Class A, 5.00 mL and pipet filler 1

Refer to Consumables and replacement items on page 6 for reorder information.

Sample collection
• Collect samples in clean glass or plastic bottles.
• If the sample is drinking water, take the control sample from a reservoir of tap water
that is known to be free of toxins if possible.

Inoculum development with indigenous biomass

1. Use one of the supplied 2. Incubate the tube

dropper pipets to add contents at 35 °C (95 °F)
1.0 mL of source culture until the broth is visibly
(indigenous biomass) to a turbid (approximately
Total Bacteria Count Broth 12 hours).
Tube. The culture can be kept for
Commercial sources of several days in the
freeze-dried bacteria may incubator or at room
also be used. temperature. Use before
72 hours for best results.

2 Toxicity, ToxTrak Method (% Inhibition)

Reaction tube colorimetric procedure

Start

1. Push Single 2. Prepare the blank: Fill 3. Clean the blank. 4. Insert the blank into the
Wavelength and enter the an empty Test 'N Tube vial cell holder.
wavelength. Refer to to the top of the label with
Summary of method deionized water.
on page 6 and
Instrument specific
information on page 1.

Zero

5. Push ZERO. The display 6. Write "control" on one 7. For each sample or 8. Add 5.0 mL of deionized
shows 0.000 Abs. Test 'N Tube vial. Then, dilution, write the sample water to the Test 'N Tube
open one ToxTrak Reagent number on each Test 'N vial. Use deionized water
Powder Pillow and add the Tube vial. Then, open one that is free of toxicity or
contents to the empty tube. ToxTrak Reagent Powder another water source that
Pillow and add the contents represents baseline toxicity.
to the empty sample vial.

9. Add 5.0 mL of sample (or 10. Add two drops of 11. Close the tube and 12. Add 0.5 mL of the
dilution) to each sample vial. Accelerator Solution to each shake to mix. inoculum (previously
Refer to Interpreting results vial. Shake to fully oxygenate the prepared) to each tube.
on page 5 to find the samples, so that the oxygen
approximate threshold level concentration does not
of toxicity for a sample. affect the respiration rate.

Toxicity, ToxTrak Method (% Inhibition) 3

 Read

13. Close the tube and 14. Clean the "control" vial. 15. Insert the "control" into 16. Push READ. Record
invert to mix. the cell holder. the absorbance value.
Repeat step 14 and 15 for
all samples and dilutions.
Record all of the
absorbance values.

17. Let the solutions in the 18. Remove the "control" 19. Clean the blank. 20. Insert the blank into the
tubes react until the from the cell holder after the cell holder.
absorbance of the "control" absorbance of the control
has decreased by 0.60 (± has decreased by 0.60 (±
0.10) Abs. This takes 45– 0.10) Abs.
75 minutes. Invert
occasionally.
The reaction time varies
according to temperature,
age of the culture, bacteria
concentrations, etc.

Zero Read

21. Push ZERO. The 22. Clean the "control" vial. 23. Insert the "control" into 24. Push READ.
display shows 0.000 Abs. the cell holder. Record the absorbance
value.

4 Toxicity, ToxTrak Method (% Inhibition)

 Read

25. Clean each sample or 26. Insert each sample or 27. Push Read. 28. Calculate the %
dilution vial. dilution into the cell holder. Record all absorbance Inhibition. Refer to Calculate
values. the Inhibition on page 5.

Calculate the Inhibition

Calculate the % Inhibition:
% I = 1–(ΔAsample/ΔAcontrol) x 100
where: Δ A = Initial absorbance value–Final absorbance value
Example:
Absorbance of control: initial = 1.500 abs, final = 0.900 abs; ΔAcontrol = 0.600
Absorbance of sample: initial = 1.700 abs, final = 1.300 abs; ΔAsample = 0.400
% I = 1–(0.400/0.600) x 100 = 33%
Interpreting results
The results as percent inhibition (% I) are a relative measurement. The results do not
represent a true quantitative measurement of toxic concentration. The percent inhibition
does not necessarily increase in direct proportion to the concentration of toxins.
Results below 10% are not reliable, but can be used to make an estimate of toxicity when
the results are consistent. If a sample shows less than 10% inhibition, repeat the test
several times. Look at the series of data points to find the likelihood of toxicity. Refer to
Table 2.
Some toxins will increase respiration and give a negative percent inhibition on this and all
other respiration-based toxicity tests. After repeated testing, samples that always give a
percent inhibition that is more negative than –10% should be considered toxic.
Table 2 Interpreting results that are less than 10% inhibition
Data points: percent inhibition Conclusion
7%, 9%, 5%, 8%, 5% May be slightly toxic
7%, -4%, -5%, 5%, 1% Most likely not toxic
-7%, -9%, -5%, -8%, -5% May be slightly toxic

Lowest observable effect concentration (LOEC)

Due to the many variables involved in the test, the limit of detection is approximately 10%
inhibition. This correlates to the Lowest Observable Effect Concentration (LOEC).
No observed effect concentration (NOEC)
To determine the minimum inhibition concentration of a toxin:

1. Dilute 1 mL of sample to 10 mL with deionized water.

2. Run the test and find the percent inhibition for the dilution.
3. Dilute 1 mL of the sample dilution from step 1 to 10 mL with deionized water.
4. Run the test and find the percent inhibition for the dilution.
5. Continue to make serial 1:10 dilutions of the sample (1:10, 1:100, 1:1000, etc.) until a
level is reached that gives 0% inhibition in the final calculation.

Toxicity, ToxTrak Method (% Inhibition) 5

 When 0% inhibition is found, the dilution represents the approximate threshold level
of toxicity for a sample. This is the No Observed Effect Concentration (NOEC).

Disposal of test cultures

Use one of the methods that follow to dispose of active bacterial cultures:
• Autoclave used test containers at 121 °C (250 °F) for 15 minutes at 15 pounds of
pressure. Once the containers are sterile, pour the contents down the drain with
running water. The reaction tubes may be washed and reused.
• Sterilize test containers with a 1:10 dilution of commercial laundry bleach. Pour the
test container contents and test containers into the bleach solution. Allow 10–
15 minutes of contact time with the bleach solution. Then pour the liquid down the
drain and wash the reaction tubes for reuse.

Summary of method
This method is based on the reduction of resazurin, a redox-active dye, by bacterial
respiration. When it is reduced, resazurin changes color from blue to pink. Toxic
substances can inhibit the rate of resazurin reduction. A chemical accelerant has been
added to shorten the reaction time. The measurement wavelength is 603 nm for
spectrophotometers or 610 nm for colorimeters.
Consumables and replacement items
Required reagents

Description Quantity/test Unit Item no.

Water, deionized varies 500 mL 27249

ToxTrak™ Reagent Set 1 25/set 2597200
Includes:
Media Set, Total Bacteria Count Tubes 1 15/pkg 2277700
Pipet, transfer, sterile 1 15/pkg 2232512
Sodium Thiosulfate Standard Solution, 0.0246 N varies 100 mL 2409232
ToxTrak™ Reagent Powder Pillows 2 50/pkg 2560766
15 mL
ToxTrak™ Accelerator Solution 4 drops 2560836
SCDB
Tubes, glass, 16 mm x 100 mm 1 6/pkg 2275806
Caps, white, Teflon lining, for 16-mm tubes 2 6/pkg 2241106

Required apparatus

Description Quantity/test Unit Item no.

Clippers 1 each 93600

Dropper, measuring, 0.5 and 1.0 mL plastic 2 20/pkg 2124720
Forceps, flat square tip 1 each 1453700
Incubator, Dri-Bath, 12 well, 120 VAC 1 each 2281400
Pipet, volumetric, Class A, 5.00-mL 1 each 1451537
Pipet filler, safety bulb 1 each 1465100

6 Toxicity, ToxTrak Method (% Inhibition)

Optional reagents and apparatus

Description Unit Item no.

®
Pipet, TenSette , 0.1–1.0 mL each 1970001
Pipet tips for TenSette Pipet 1970001 50/pkg 2185696
®
Pipet, TenSette , 1.0 to 10.0 mL each 1970010
Pipet tips for TenSette Pipet 1970010 50/pkg 2199796
Test tube rack each 1864100
Pipet tips for TenSette Pipet 1970001 1000/pkg 2185628
Pipet tips for TenSette Pipet 1970010 250/pkg 2199725
BOD seed (polyseed) 50/pkg 2918700
Laboratory pen, permanent marker each 2092000

Toxicity, ToxTrak Method (% Inhibition) 7

 FOR TECHNICAL ASSISTANCE, PRICE INFORMATION AND ORDERING: HACH COMPANY
In the U.S.A. – Call toll-free 800-227-4224 WORLD HEADQUARTERS
Outside the U.S.A. – Contact the HACH office or distributor serving you. Telephone: (970) 669-3050
On the Worldwide Web – www.hach.com; E-mail – [email protected] FAX: (970) 669-2932

© Hach Company/Hach Lange GmbH, 1989–2013. All rights reserved. 04/2013, Edition 8
Turbidity DOC316.53.01332

Absorptometric Method1 Method 8237

21 to 1000 FAU
Scope and application: For water, wastewater and seawater.
1 Adapted from FWPCA Methods for Chemical Analysis of Water and Wastes, 275 (1969).

Test preparation

Instrument-specific table
The table in this section shows all of the instruments that have the program for this test.
Instrument specific information PP shows sample cell and orientation requirements for
reagent addition tests, such as powder pillow or bulk reagent tests.
Table 1 Instrument-specific information for reagent addition
Instrument Sample cell orientation Sample cell
DR 900 The orientation mark is toward the user. 2401906

Before starting
Install the instrument cap on the DR 900 cell holder before ZERO or READ is pushed.
For samples with high color or turbidity, use a filtered portion of sample instead of the deionized water for the blank.
Review the Safety Data Sheets (MSDS/SDS) for the chemicals that are used and use any recommended personal protective
equipment.
Dispose of reacted solutions according to local, state and federal regulations. Use the Safety Data Sheets for disposal
information for unused reagents. Consult the environmental, health and safety staff for your facility and/or local regulatory
agencies for further disposal information.

Items to collect
Description Quantity

Sample cells 2

Refer to Consumables and replacement items on page 3 for reorder information.

Sample collection and storage

• Collect samples in clean glass or plastic bottles.
• To preserve samples for later analysis, keep the samples at or below 6 °C (43 °F) for
up to 48 hours.
• Let the sample temperature increase to room temperature before analysis.

1
Absorbtometric method

Start

1. Start program 745 FAU. 2. Prepare the blank: Fill a 3. Clean the blank. 4. Insert the blank into the
sample cell with 10 mL of cell holder.
deionized water.

Zero

5. Push ZERO. The display 6. Prepare the sample: Fill 7. Clean the prepared 8. Insert the prepared
shows 0 FAU. a second sample cell with sample. sample into the cell holder.
10 mL of sample.
Mix the sample well before it
is added to the sample cell.

Read

9. Push READ. Results

show in Formazin
Attenuation Units (FAU).

Interferences
Interfering substance Interference level
Air bubbles Interfere at all levels. Use the Degassing Kit or an ultrasonic bath to degas the samples.
Color Interferes if the color absorbs light at the measurement wavelength.
Temperature extremes May interfere by changing the turbidity of the sample. Analyze samples as soon as possible after
collection. Analyze at the same temperature as the original sample.

2 Turbidity, Absorptometric Method (1000 FAU)

Accuracy check
Standard solution method
Use the standard solution method to validate the test procedure, reagents and
instrument.
Items to collect:
• 4000 NTU Formazin Stock Solution
• 100-mL volumetric flask, Class A
• 5-mL volumetric pipet, Class A and pipet filler
• Deionized water

1. Prepare a 200 FAU formazin standard solution as follows:

a. Use a pipet to add 5.00 mL of 4000 NTU formazin standard solution into the
volumetric flask. As an alternative, use a 200 NTU StablCal™ Standard Solution.
b. Dilute to the mark with deionized water. Mix well. Prepare this solution daily.
2. Use the test procedure to measure the concentration of the prepared standard
solution.
3. Compare the expected result to the actual result.
Note: The factory calibration can be adjusted slightly with the standard adjust option so that the
instrument shows the expected value of the standard solution. The adjusted calibration is then
used for all test results. This adjustment can increase the test accuracy when there are slight
variations in the reagents or instruments.

Method performance
The method performance data that follows was derived from laboratory tests that were
measured on a spectrophotometer during ideal test conditions. Users may get different
results under different test conditions.
Program Standard Precision (95% Confidence Interval) Sensitivity
Concentration change per 0.010 Abs change
745 200 FAU 195–205 FAU 21 FAU

Summary of method
This turbidity test measures an optical property of the sample which results from
scattering and absorption of light by particles in the sample. The amount of turbidity
measured depends on variables such as the size, shape, color and refractive properties
of the particles. This procedure is calibrated using formazin turbidity standards and the
readings are in terms of Formazin Attenuation Units (FAU). This test cannot be used for
USEPA reporting purposes, but it may be used for daily in-plant monitoring. One FAU is
equivalent to one Nephelometric Turbidity Unit (NTU) of Formazin. However, the optical
method of measurement for FAU is very different from the nephelometric (NTU) method
(1 NTU = 1 FTU = 1 FAU when traced to formazin primary standards). Test results are
measured at 520 nm.
Consumables and replacement items
Recommended standards and apparatus

Description Quantity/test Unit Item no.

Formazin Stock Solution, 4000 NTU 1 500 mL 246149

Silicone Oil 1 15 mL DB 126936
StablCal Stabilized Turbidity Standard, 200 NTU 1 500 mL 2660449
Water, deionized varies 4L 27256

Turbidity, Absorptometric Method (1000 FAU) 3

Optional reagents and apparatus

Description Unit Item no.

Bath, ultrasonic each 2489500

Bottle, wash, 250 mL each 62031
Flask, volumetric, Class A, 100-mL each 1457442
Flask, filter, 500 mL each 54649
Filter holder each 1352900
Filter pump, aspirator each 213100
Oiling cloth, for applying silicone oil each 2687300
Pipet filler, safety bulb each 1465100
Pipet, volumetric 5.00-mL each 1451537
Sample Degassing Kit each 4397500
Stopper, rubber, one-hole, No. 7 6/pkg 211907
3.66 m
Tubing, rubber, 5/16-in. inside diameter 56019
(12 ft)
Tweezers, plastic each 1428200

FOR TECHNICAL ASSISTANCE, PRICE INFORMATION AND ORDERING: HACH COMPANY

In the U.S.A. – Call toll-free 800-227-4224 WORLD HEADQUARTERS
Outside the U.S.A. – Contact the HACH office or distributor serving you. Telephone: (970) 669-3050
On the Worldwide Web – www.hach.com; E-mail – [email protected] FAX: (970) 669-2932

© Hach Company/Hach Lange GmbH, 1989–2013. All rights reserved. 04/2013, Edition 8
Volatile Acids DOC316.53.01144

Esterification Method1 Method 8196

27 to 2800 mg/L (as acetic acid) Reagent Solution
Scope and application: For digestor sludges.
1 Adapted from The Analyst, 87, 949 (1962).

Test preparation

Instrument specific information

The table in this section shows all of the instruments that have the program for this test.
Table 1 shows sample cell and orientation requirements for reagent addition tests, such
as powder pillow or bulk reagent tests.
To use the table, select an instrument, then read across to find the corresponding
information for this test.
Table 1 Instrument-specific information for reagent solution
Instrument Sample cell orientation Sample cell
DR 6000 The fill line is to the right. 2495402
DR 3800
DR 2800
DR 2700
DR 5000 The fill line is toward the user.
DR 3900
DR 900 The orientation mark is toward the user. 2401906

Before starting
Install the instrument cap on the DR 900 cell holder before ZERO or READ is pushed.
Review the Safety Data Sheets (MSDS/SDS) for the chemicals that are used and use any recommended personal protective
equipment.
Dispose of reacted solutions according to local, state and federal regulations. Use the Safety Data Sheets for disposal
information for unused reagents. Consult the environmental, health and safety staff for your facility and/or local regulatory
agencies for further disposal information.

Items to collect
Description Quantity

Centrifuge 1
Centrifuge tubes and caps 2
Cylinder, 10-mL graduated 1
Ethylene Glycol 3 mL
Ferric Chloride-Sulfuric Acid Solution 20 mL
Funnel and filter paper varies

1
Items to collect (continued)
Description Quantity

Hot Plate 1
Hydroxylamine Hydrochloride Solution, 100-g/L 1 mL
Pipet filler 1
Pipet, 2-mL 1
Pipet, Class A volumetric, 0.50-mL1 1
Pipet, Class A volumetric, 10-mL 1
Sample cells (For information about sample cells, adapters or light shields, refer to Instrument
2
specific information on page 1.)
Sodium Hydroxide Standard Solution, 4.5 N 4 mL
Sulfuric Acid Standard Solution, 19.2 N 0.4 mL
Water bath and rack 1
Water, deionized 20.5 mL
1 A TenSette Pipet can be used in place of individual pipets in this procedure.

Refer to Consumables and replacement items on page 5 for reorder information.

Sample collection and storage

• Collect samples in clean glass or plastic bottles.
• To preserve samples for later analysis, keep the samples at or below 6 °C (43 °F) for
up to 24 hours.
• Let the sample temperature increase to room temperature before analysis.

Esterification method

Start

1. Start program 2. Prepare the blank: Use 3. Use a filter or a 4. Prepare the sample:
770 Volatile Acids. For a pipet to add 0.5 mL of centrifuge to separate Use a pipet to add 0.5 mL of
information about sample deionized water to a sample 10 mL of sample. the filtrate or supernatant to
cells, adapters or light cell. a second sample cell.
shields, refer to Instrument
specific information
on page 1.
Note: Although the program
name may vary between
instruments, the program
number does not change.

2 Volatile Acids, Esterification Method (2800 mg/L)

5. Add 1.5 mL of Ethylene 6. Swirl to mix. 7. Add 0.2 mL of 19.2 N 8. Swirl to mix.
Glycol to each sample cell. Sulfuric Acid to each sample
cell.

9. Put both cells into a 10. Start the instrument 11. When the timer expires, 12. Add 0.5 mL of
boiling water bath. timer. A 3-minute reaction use a cold water bath to Hydroxylamine
The sample cells can also time starts. cool the samples to 25 °C Hydrochloride Solution to
be boiled in a 500-mL (the cells will feel cold). each sample cell.
beaker.

13. Swirl to mix. 14. Add 2.0 mL of 4.5 N 15. Swirl to mix. 16. Add 10 mL of Ferric
Sodium Hydroxide Standard Chloride Sulfuric Acid
Solution to each cell. Solution to each sample
cell.

17. Swirl to mix. 18. Add 10 mL of deionized 19. Close both cells. Invert 20. Transfer 10 mL of the
water to each sample cell. both cells to mix. blank solution to a clean
sample cell.

Volatile Acids, Esterification Method (2800 mg/L) 3

21. Transfer 10 mL of the 22. Start the instrument 23. When the timer expires, 24. Insert the blank into the
prepared sample solution to timer. A second 3-minute clean the blank. cell holder.
a clean sample cell. reaction time starts.
Set the instrument to zero
during this reaction time.

Zero Read

25. Push ZERO. The 26. Clean the prepared 27. Insert the prepared 28. Push READ. Results
display shows 0 mg/L sample. sample into the cell holder. show in mg/L HOAC.
HOAC.

Accuracy check
Standard additions method (sample spike)
Use the standard additions method (for applicable instruments) to validate the test
procedure, reagents and instrument and to find if there is an interference in the sample.
Items to collect:
• Volatile Acid Voluette® Ampule Standard, 62,500-mg/L as acetic acid
• Ampule breaker
• Pipet, TenSette®, 0.1–1.0 mL and tips
• Mixing cylinders, 25-mL (3)

1. Use the test procedure to measure the concentration of the sample, then keep the
(unspiked) sample in the instrument.
2. Go to the Standard Additions option in the instrument menu.
3. Select the values for standard concentration, sample volume and spike volumes.
4. Open the standard solution.
5. Prepare three spiked samples: use the TenSette pipet to add 0.1 mL, 0.2 mL and
0.3 mL of the standard solution, respectively, to three 25-mL portions of fresh sample.
Mix well.
6. Use the test procedure to measure the concentration of each of the spiked samples.
Start with the smallest sample spike. Measure each of the spiked samples in the
instrument.
7. Select Graph to compare the expected results to the actual results.
Note: If the actual results are significantly different from the expected results, make sure that
the sample volumes and sample spikes are measured accurately. The sample volumes and

4 Volatile Acids, Esterification Method (2800 mg/L)

 sample spikes that are used should agree with the selections in the standard additions menu. If
the results are not within acceptable limits, the sample may contain an interference.

Standard solution method

Use the standard solution method to validate the test procedure, reagents and
instrument.
Items to collect:
• Volatile Acid Voluette® Ampule Standard, 62,500-mg/L as acetic acid
• 500-mL volumetric flask, Class A
• 4-mL volumetric pipet, Class A and pipet filler
• Deionized water

1. Prepare a 500 mg/L volatile acids standard solution as follows:

a. Use a pipet to add 4.00 mL of 62,500-mg/L as acetic acid standard solution into
the volumetric flask.
b. Dilute to the mark with deionized water. Mix well. Prepare this solution daily.
2. Use the test procedure to measure the concentration of the prepared standard
solution.
3. Compare the expected result to the actual result.
Note: The factory calibration can be adjusted slightly with the standard adjust option so that the
instrument shows the expected value of the standard solution. The adjusted calibration is then
used for all test results. This adjustment can increase the test accuracy when there are slight
variations in the reagents or instruments.

Method performance
The method performance data that follows was derived from laboratory tests that were
measured on a spectrophotometer during ideal test conditions. Users may get different
results under different test conditions.
Program Standard Precision (95% Confidence Interval) Sensitivity
Concentration change per 0.010 Abs
change
770 1350 mg/L as acetic acid 1218–1482 mg/L as acetic acid 27 mg/L as acetic acid (HOAC)
(HOAC) (HOAC)

Summary of method
The volatile acids test is designed specifically to determine volatile acids in digestor
sludges. The method is based on esterification of the carboxylic acids present in the
sample and subsequent determination of the esters by the ferric hydroxamate reaction.
All volatile acids present are reported as their equivalent mg/L as acetic acid. The
measurement wavelength is 495 nm for spectrophotometers or 520 nm for colorimeters.
Consumables and replacement items
Required reagents

Description Quantity/test Unit Item no.

Water, deionized varies 4L 27256

Volatile Acid Reagent Set 1 90 tests 2244700
Includes:
Ethylene Glycol 3 mL 1000 mL 203953
Ferric Chloride-Sulfuric Acid Solution 20 mL 1000 mL 204253
Hydroxylamine Hydrochloride Solution, 100-g/L 1 mL 100 mL 81842

Volatile Acids, Esterification Method (2800 mg/L) 5

Consumables and replacement items (continued)
Description Quantity/test Unit Item no.

Sodium Hydroxide Standard Solution, 4.5 N 4 mL 1000 mL 204053

Sulfuric Acid Standard Solution, 19.2 N 0.4 mL 1000 mL 203832

Required apparatus

Description Quantity/test Unit Item no.

Centrifuge, 115 VAC, 6 x 15 mL 1 each 2676500

Centrifuge tubes, 15-mL 2 10/pkg 2278739
Centrifuge tube caps 2 20/pkg 2585220
Cylinder, graduated, 10-mL 1 each 50838
Filter paper, folded, 12.5-cm 1 100/pkg 189457
Funnel, poly, 65-mm 1 each 108367
Hot plate, 7 inches x 7 inches, digital, 120 VAC 1 each 2881500
Hot Plate, 7-inch digital, 240 VAC 1 each 2881502
Pipet filler, safety bulb 1 each 1465100
Pipet, serological, 2-mL 1 each 53236
Pipet, volumetric, Class A, 0.5-mL 1 each 1451534
Pipet, volumetric, Class A, 10.00-mL 1 each 1451538
Cell holder assembly 1 each 4788000
Evaporating dish, 125 mm x 65 mm 1 each 2764700
Sample cell, 10 mL square, matched pair 2 2/pkg 2495402

Recommended standards and apparatus

Description Unit Item no.

®
Volatile Acids Standard Solution, 10-mL Voluette Ampule, 62,500-mg/L as HOAC 16/pkg 14270-10
®
Ampule Breaker, Voluette ampules each 2196800

Optional reagents and apparatus

Description Unit Item no.

Cylinder, mixing, 25-mL each 189640

Water bath and rack each 195555
®
Pipet, TenSette , 1.0 to 10.0 mL each 1970010
Pipet tips for TenSette Pipet 1970010 250/pkg 2199725
Pipet tips for TenSette Pipet 1970010 50/pkg 2199796
®
Pipet, TenSette , 0.1–1.0 mL each 1970001
Pipet tips for TenSette Pipet 1970001 50/pkg 2185696
Pipet tips for TenSette Pipet 1970001 1000/pkg 2185628
Flask, volumetric, Class A, 500-mL each 1457449
Finger cots 2/pkg 1464702

6 Volatile Acids, Esterification Method (2800 mg/L)

Volatile Acids, Esterification Method (2800 mg/L) 7
 FOR TECHNICAL ASSISTANCE, PRICE INFORMATION AND ORDERING: HACH COMPANY
In the U.S.A. – Call toll-free 800-227-4224 WORLD HEADQUARTERS
Outside the U.S.A. – Contact the HACH office or distributor serving you. Telephone: (970) 669-3050
On the Worldwide Web – www.hach.com; E-mail – [email protected] FAX: (970) 669-2932

© Hach Company/Hach Lange GmbH, 1989–2013. All rights reserved. 04/2013, Edition 8
Zinc DOC316.53.01145

USEPA1 Zincon Method2 Method 8009

0.01 to 3.00 mg/L Zn Powder Pillows
Scope and application: For water and wastewater. Digestion is required for a total zinc analysis.
1 USEPA approved for wastewater analyses 3500 Zn B: Federal Register, 45(105) 36166 (May 29, 1980).
2 Adapted from Standard Methods for the Examination of Water and Wastewater.

Test preparation

Instrument specific information

The table in this section shows all of the instruments that have the program for this test.
Table 1 shows sample cell and orientation requirements for reagent addition tests, such
as powder pillow or bulk reagent tests.
To use the table, select an instrument, then read across to find the corresponding
information for this test.
Table 1 Instrument-specific information for powder pillows
Instrument Sample cell orientation Sample cell
DR 6000 The fill line is to the right. 2495402
DR 3800
DR 2800
DR 2700
DR 5000 The fill line is toward the user.
DR 3900
DR 900 The orientation mark is toward the user. 2401906

Before starting
Install the instrument cap on the DR 900 cell holder before ZERO or READ is pushed.
Clean all glassware with 6.0 N (50%) hydrochloric acid, then rinse thoroughly with deionized water to remove contaminants.
Use only glass-stoppered mixing cylinders in this procedure.
Make sure that the dropper that is used in this procedure is plastic. Droppers that have rubber bulbs can contaminate the
reagent.
The reagents that are used in this test contain potassium cyanide. Keep cyanide solutions at pH > 11 to prevent
exposure to hydrogen cyanide gas. Collect the reacted samples for proper disposal.
Review the Safety Data Sheets (MSDS/SDS) for the chemicals that are used and use any recommended personal protective
equipment.
Do not use the Pour-Thru Cell or sipper module (for applicable instruments) with this test.
Dispose of reacted solutions according to local, state and federal regulations. Use the Safety Data Sheets for disposal
information for unused reagents. Consult the environmental, health and safety staff for your facility and/or local regulatory
agencies for further disposal information.

1
Items to collect
Description Quantity

Cyclohexanone 0.5 mL
ZincoVer® 5 Reagent Powder Pillow, 20-mL 1
Cylinder, graduated mixing, 25-mL 1
Sample cells (For information about sample cells, adapters or light shields, refer to Instrument
2
specific information on page 1.)

Refer to Consumables and replacement items on page 6 for reorder information.

Sample collection and storage

• Collect samples in clean glass or plastic bottles that have been cleaned with
1:1 hydrochloric acid and rinsed with deionized water.
• To preserve samples for later analysis, adjust the sample pH to less than 2 with
concentrated nitric acid (about 2 mL per liter). No acid addition is necessary if the
sample is tested immediately.
• Keep preserved samples at room temperature for a maximum of 6 months.
• Before analysis, adjust the pH to 4–5 with 5.0 N sodium hydroxide standard solution.
Do not exceed pH 5 as zinc can precipitate.
• Correct the test result for the dilution from the volume additions.

Powder pillow procedure

Start

1. Start program 780 Zinc. 2. Fill a 25-mL graduated 3. Add the contents of one 4. Close the cylinder. Invert
For information about mixing cylinder with 20 mL ZincoVer 5 Reagent Powder the cylinder several times to
sample cells, adapters or of sample. Pillow to the mixing cylinder. dissolve the powder
light shields, refer to completely. Inconsistent
Instrument specific readings may result if all the
information on page 1. particles are not dissolved.
Note: Although the program The sample should be
name may vary between orange. If the sample is
instruments, the program brown or blue, the zinc
number does not change. concentration is too high or
an interfering metal is
present. Dilute the sample
and repeat the test.

2 Zinc, Zincon Method (3.00 mg/L)

5. Blank preparation: Pour 6. Prepared sample: Use a 7. Start the instrument 8. During the reaction
10 mL of the solution into a plastic dropper to add timer. A 30-second reaction period, close the mixing
sample cell. 0.5 mL of cyclohexanone to time starts. cylinder and vigorously
the solution that is still in the shake the prepared sample.
mixing cylinder. The sample shows reddish-
orange, brown or blue,
depending on the zinc
concentration.

9. Start the instrument 10. Pour the prepared 11. When the timer expires, 12. Insert the blank into the
timer. A 3-minute reaction sample solution from the clean the blank. cell holder.
time starts. mixing cylinder into a
During the reaction period, second sample cell.
complete the next step.

Zero Read

13. Push ZERO. The 14. Clean the prepared 15. Insert the prepared 16. Push READ. Results
display shows 0.00 mg/L Zn. sample. sample into the cell holder. show in mg/L Zn.

Interferences
Interfering substance Interference level
Aluminum More than 6 mg/L
Cadmium More than 0.5 mg/L
Copper More than 5 mg/L
Iron (ferric) More than 7 mg/L
Manganese More than 5 mg/L

Zinc, Zincon Method (3.00 mg/L) 3

Interfering substance Interference level
Nickel More than 5 mg/L
Organic Material Large amounts may interfere. Pretreat the sample with a mild digestion.
Highly buffered samples or Can prevent the correct pH adjustment of the sample by the reagents. Sample
extreme sample pH pretreatment may be necessary. Adjust the pH to 4–5.
Amino-tri(methylene phosphonic Samples that contain AMP cause a negative interference. Digest the sample to remove
acid) (AMP) this interference (use the total phosphorus hot plate digestion, Method 8190).
Note: Be sure to adjust the pH of the sample after the digestion to pH 4–5 with sodium hydroxide
before the zinc analysis.

Digestion
For total zinc determinations, the sample must be digested with heat and acid to make
sure that all forms of the metal are measured. The steps that follow can be used for a
mild digestion.
Note: The following procedure is the USEPA mild digestion procedure. Refer to the Water Analysis
Guide for more digestion procedures.

1. Add concentrated nitric acid to the sample with a glass serological pipet and pipet
filler:
• If the sample was acidified for preservation, add 3 mL of nitric acid to 1 liter of the
preserved sample.
• If the sample was not acidified for preservation, add 5 mL of nitric acid to 1 liter of
sample.
2. Transfer 100 mL of acidified sample to a 250-mL Erlenmeyer flask.
3. Add 5 mL of 1:1 hydrochloric acid.
4. Put the sample on a hot plate at 95 °C (203 °F) until 15–20 mL of the sample
remains. Make sure that the sample does not boil.
5. Put the cooled sample through a 0.45-µm filter to remove any insoluble material.
6. Adjust the pH of the digested sample to pH 4–5 with 5.0 N sodium hydroxide. Do not
exceed pH 5 as zinc may precipitate.
7. Quantitatively transfer the sample to a 100-mL volumetric flask and dilute to the mark
with deionized water.

Accuracy check
Standard additions method (sample spike)
Use the standard additions method (for applicable instruments) to validate the test
procedure, reagents and instrument and to find if there is an interference in the sample.
Items to collect:
• 25 mg/L Zinc Voluette® Ampule Standard Solution
• Ampule breaker
• Pipet, TenSette®, 0.1–1.0 mL and tips
• Mixing cylinders, 25-mL (3)

1. Use the test procedure to measure the concentration of the sample, then keep the
(unspiked) sample in the instrument.
2. Go to the Standard Additions option in the instrument menu.
3. Select the values for standard concentration, sample volume and spike volumes.
4. Open the standard solution.
5. Prepare three spiked samples: use the TenSette pipet to add 0.1 mL, 0.2 mL and
0.3 mL of the standard solution, respectively, to three 20-mL portions of fresh sample.
Mix well.

4 Zinc, Zincon Method (3.00 mg/L)

 6. Use the test procedure to measure the concentration of each of the spiked samples.
Start with the smallest sample spike. Measure each of the spiked samples in the
instrument.
7. Select Graph to compare the expected results to the actual results.
Note: If the actual results are significantly different from the expected results, make sure that
the sample volumes and sample spikes are measured accurately. The sample volumes and
sample spikes that are used should agree with the selections in the standard additions menu. If
the results are not within acceptable limits, the sample may contain an interference.

Standard solution method

Use the standard solution method to validate the test procedure, reagents and
instrument.
Items to collect:
• 100 mg/L zinc standard solution
• 1-L volumetric flask, Class A
• 10-mL volumetric pipet, Class A and pipet filler
• Deionized water

1. Prepare a 1.00 mg/L zinc standard solution as follows:

a. Use a pipet to add 10.00 mL of 100 mg/L zinc standard solution into the
volumetric flask.
b. Dilute to the mark with deionized water. Mix well. Prepare this solution daily.
2. Use the test procedure to measure the concentration of the prepared standard
solution.
3. Compare the expected result to the actual result.
Note: The factory calibration can be adjusted slightly with the standard adjust option so that the
instrument shows the expected value of the standard solution. The adjusted calibration is then
used for all test results. This adjustment can increase the test accuracy when there are slight
variations in the reagents or instruments.

Method performance
The method performance data that follows was derived from laboratory tests that were
measured on a spectrophotometer during ideal test conditions. Users may get different
results under different test conditions.
Program Standard Precision (95% Confidence Interval) Sensitivity
Concentration change per 0.010 Abs change
780 1.00 mg/L Zn 0.97–1.03 mg/L Zn 0.013 mg/L Zn

Summary of method
Zinc and other metals in the sample are complexed with cyanide. Adding cyclohexanone
causes a selective release of zinc. The zinc reacts with 2-carboxy-2'-hydroxy-5'-
sulfoformazyl benzene (zincon) indicator to form a blue-colored species. The blue color is
masked by the brown color from the excess indicator. The intensity of the blue color is
proportional to the amount of zinc present. The measurement wavelength is 620 nm for
spectrophotometers or 610 nm for colorimeters.
Pollution prevention and waste management
Reacted samples contain potassium cyanide and must be disposed of as a hazardous
waste. Dispose of reacted solutions according to local, state and federal regulations.

Zinc, Zincon Method (3.00 mg/L) 5

Consumables and replacement items
Required reagents

Description Quantity/test Unit Item no.

Zinc Reagent Set, 20-mL sample size, includes: — 100 tests 2429300
100 mL
Cyclohexanone 0.5 mL 1403332
MDB
®
ZincoVer 5 Reagent Powder Pillow, 20-mL 1 100/pkg 2106669

Required apparatus

Description Quantity/test Unit Item no.

Cylinder, graduated mixing, 25 mL with stopper 1 each 2088640

Recommended standards

Description Unit Item no.

Water, deionized 4L 27256

Zinc Standard Solution, 100-mg/L 100 mL 237842
®
Zinc Standard Solution, 10-mg/L Voluette Ampule, 25-mL as Zn 16/pkg 1424610
Zinc Standard Solution, 1000-mg/L 100 mL 1417742

Optional reagents and apparatus

Description Unit Item no.

Flask, Erlenmeyer, 250-mL each 50546

Hot plate, 120 V each 1206701
Hydrochloric Acid, 6.0 N 1:1, 50% 500 mL 88449
Nitric Acid, concentrated 500 mL 15249
®
Pipet, TenSette , 0.1–1.0 mL each 1970001
Pipet tips for TenSette Pipet 1970001 50/pkg 2185696
®
Ampule Breaker, Voluette ampules each 2196800
Pipet, volumetric, Class A, 10-mL each 1451538
Pipet filler, safety bulb each 1465100
Flask, volumetric, Class A, 1000-mL each 1457453

FOR TECHNICAL ASSISTANCE, PRICE INFORMATION AND ORDERING: HACH COMPANY

In the U.S.A. – Call toll-free 800-227-4224 WORLD HEADQUARTERS
Outside the U.S.A. – Contact the HACH office or distributor serving you. Telephone: (970) 669-3050
On the Worldwide Web – www.hach.com; E-mail – [email protected] FAX: (970) 669-2932

© Hach Company/Hach Lange GmbH, 1989–2013. All rights reserved. 04/2013, Edition 8