PHAR1018

Drug Structure
Practical 1: Calibrations,
Standardisations and Titrations

Student Name

Student ID

Practical Group
Overview
This practical is split into two sessions:
 Morning Session (09:30 am – 12:30 pm)
Practical 1a. Calibration of glassware
In this experiment you will calibrate pipettes, burettes and volumetric
flasks.
Practical 1b. Standardisation of HCl using borax
In this experiment you will standardise a solution of HCl for use in
Practical 1d.
 Afternoon Session (2 pm – 5 pm)
Practical 1c. Standardisation of NaOH using HCl
In this experiment you will standardise a solution of NaOH for use in
part 1d, using your standardised solution of HCl.
Practical 1d. Assay of aspirin tablets by back-titration
In this experiment you will quantify the amount of aspirin in tablets,
using a ‘back-titration’ method.

Aims
 To develop key practical skills including an appreciation of COSHH and
SOP’s
 To become proficient in the use of pipettes, burettes, volumetric flasks and
balances
 To understand the terms: accuracy, precision and tolerance
 To become proficient in performing titrations, including the associated
calculations
 To carry out a British Pharmacopoeia method for the assay of aspirin tablets

Safety
Personal Protective Equipment (PPE)
 You must always wear your safety glasses and lab coat
 Wear gloves where appropriate
 Long hair must be tied back
 No open-toed shoes

Acids and Bases

 Take care using acids and bases
 Report any spillages to a member of staff
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  Be aware of the location of the emergency eye wash station

Pipettes and pipette fillers

 Choose the right size pipette and pipette filler
 Check that there are no chips in the glassware
 Gently push the pipette into the pipette filler – do not force
 Keep the filler on the pipette to expel the contents

Balances
 Do not use if the pan is dirty
 Do not attempt to clean the balance if you do not know what the spillage is
 Clean up your own spillages after use

Glassware
 Do not use glassware with chips or cracks
 Take care rinsing out glassware – check for cracks
 Do not pick up broken glass, ask a member of staff to help

Thermometers
 Should not be used for stirring
 If a mercury thermometer breaks, inform a member of staff immediately

COSHH

Substance Pictogram Hazard Statement

Aspirin  Harmful if swallowed

 May be corrosive to metals

0.5M NaOH  Causes severe skin burns and eye
damage
 Causes severe skin burns and eye
0.5M HCl
damage

 Causes serious eye irritation

Borax  May damage fertility
 May damage the unborn child
 Toxic if swallowed
Barium chloride
 Causes serious eye irritation
solution
 Harmful if inhaled
Methyl red, phenol
red, bromophenol
 Highly flammable liquid and vapor
blue,
phenolphthalein

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Practical 1a: Calibration of glassware

 In this experiment you will calibrate pipettes, burettes and volumetric flasks.
The accurate delivery and measurement of liquids within Pharmaceutical Analysis is
crucial. This practical experiment re-introduces the safe and accurate practice of
measuring volumes and introduces the concept of accuracy in glassware.
Grade B volumetric glassware is satisfactory for the assay of most pharmaceutical
substances, but if very great accuracy is required all pipettes and burettes should be
calibrated. This is achieved by determination of the weight of deionised water they
deliver, and subsequent calculation of the volume delivered.

Accuracy
How closely a measurement represents the true value of what is being measured.

Precision
How well a set of results agree with each other.

Tolerance
The maximum permissible error in a measurement determined by a particular
piece of equipment.

Meniscus
Water adheres to the sides of glass containers, and therefore has a meniscus – a
curve at its surface. When using volumetric glassware, readings are always taken
from the bottom of the meniscus.

Pipettes
Volumetric pipettes are designed to accurately measure one specific volume.
They have a single graduation mark to indicate this specific volume.
Graduated pipettes have multiple graduations at regular intervals, and are a lot
like burettes.

Burettes
Burettes are used to dispense variable quantities of solutions using the stopcock.
The graduations start at 0.00 mL at the top of the burette, and increase moving
down the burette. An initial reading is taken, the liquid is dispensed, then a final
reading is taken – the difference between the two readings is the amount of liquid
dispensed.

Volumetric Flask
Volumetric flasks are designed to accurately measure one specific volume. They
have a single graduation line.

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Method
1. Fill a 400 mL beaker with deionised water.
2. Measure and record the temperature of the water.

Temperature of water / °C

3. The density of water varies with temperature. Use the following table to
determine the density of your deionised water.

Density / Density /
Temperature / °C Temperature / °C
g mL−1 g mL−1
15 0.99793 18 0.99751
16 0.99780 19 0.99735
17 0.99766 20 0.99718

Density of water / g mL-1

10 mL volumetric pipette
4. Measure and record the mass of a 100 mL conical flask to the nearest 0.1
mg (use 4 d.p. balance).

Mass of conical flask / g

5. Pipette 10.0 mL of water into the flask using a volumetric pipette:

a. Rinse the pipette twice with the liquid you are about to dispense, then
dispose of that liquid appropriately.
b. Squeeze the pipette filler bulb, and gently place it on the end of the
pipette – you do not need to force it.
c. Place the pipette tip in the solution, then gently release the bulb,
drawing liquid into the pipette.
d. Fill the pipette above the calibration line.
e. Remove the pipette from the liquid and wipe off any excess water from
the outside of the pipette/tip.
f. Carefully lower the level of the liquid to the point that the bottom of the
meniscus is level with the calibration line.
g. Touch the tip of the pipette to the side of the original container to
remove any remaining drops.
h. Place the tip of the pipette into the receiving vessel and dispense.
i. Touch the tip of the pipette to the side of the vessel.
j. Rinse the pipette with deionised water (unless you are going to pipette
again with the same solution).
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 6. Record the mass of conical flask + water dispensed to the nearest 0.1 mg.
7. Repeat step 5 until the mass of water delivered is ± 50 mg each time
(concordant). There is no need to empty the conical flask each time, as you
can calculate the mass of water added from the previous mass.

Mass of water dispensed from 10 mL

Mass of conical flask + water / g
pipette / g
1
2
3
4
5

8. Calculate the mean of your concordant readings.

Mean mass of water dispensed from 10 mL pipette / g

9. Use your mean mass and the density of water (determined earlier) to
calculate the mean volume of water dispensed.
mass
density =
volume

Mean volume of water dispensed

from 10 mL pipette / mL

10. Dispose of the contents of the conical flask appropriately.

25 mL volumetric pipette
11. Measure and record the mass of a 100 mL conical flask to the nearest 0.1
mg.

Mass of conical flask / g

12. Pipette 25.0 mL of water into the flask using a volumetric pipette (see step 5
for method).

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13. Rinse the pipette with deionised water (unless you are going to pipette again
with the same solution).
14. Record the mass of conical flask + water dispensed to the nearest 0.1 mg.
15. Repeat step 12 until the mass of water delivered is ± 100 mg each time
(concordant). There is no need to empty the conical flask each time, as you
can calculate the mass of water added from the previous mass.

Mass of water dispensed from 25 mL

Mass of conical flask + water / g
pipette / g
1
2
3
4
5

16. Calculate the mean of your concordant readings.

Mean mass of water dispensed from 25 mL pipette / g

17. Use your mean mass and the density of water (determined earlier) to
calculate the mean volume of water dispensed.
mass
density =
volume

Mean volume of water dispensed

from 25 mL pipette / mL

18. Dispose of the contents of the conical flask appropriately.

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10 mL burette aliquots
19. Measure and record the mass of a 100 mL conical flask to the nearest 0.1
mg.

Mass of conical flask / g

20. Dispense a 10.0 mL aliquot of deionised water into the flask using a burette:
a. Rinse the burette twice with the liquid you are about to dispense, then
dispose of that liquid appropriately.
b. Clamp the burette vertically.
c. Using a funnel, fill the burette with the liquid to be dispensed, note that
you should not ‘aim’ for a particular number – it does not matter how full
the burette is (as long as it is on the scale, and is enough to be
dispensed).
d. Record the initial burette reading to the nearest 0.05 mL.
e. Slowly open the stopcock to allow the liquid to drain into the collection
vessel. When approaching the desired volume, dispense dropwise.
f. Close the stopcock once the desired volume has been dispensed.
g. Gently tap the burette to the side of the collection flask to collect drops.
h. Record the final burette reading to the nearest 0.05 mL.
i. Touch the tip of the burette to the side of the original container to
remove any remaining drops.
21. Record the mass of the conical flask + water dispensed to the nearest 0.1
mg.
22. Repeat step 20 until the mass of water delivered is ± 100 mg each time
(concordant). There is no need to empty the conical flask each time, as you
can calculate the mass of water added from the previous mass.

Initial burette Final burette Mass of flask + Mass dispensed /

reading / mL reading / mL water / g g

1
2
3
4
5

23. Calculate the mean of your concordant readings.

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 Mean mass of water dispensed from burette / g

24. Use your mean mass and the density of water (determined earlier) to
calculate the mean volume of water dispensed.
mass
density =
volume

Mean volume of water dispensed from burette / mL

25. Dispose of the contents of the conical flask appropriately.

50 mL volumetric flask
26. Measure and record the mass of a 50 mL volumetric flask (and cap) to the
nearest 0.1 mg.

Mass of volumetric flask / g

27. Fill the volumetric flask with 50.0 mL deionised water.

a. Rinse the volumetric flask twice with the liquid you are about to
dispense, then dispose of that liquid appropriately.
b. Fill the flask until the level of the liquid is 1 - 2 cm below the graduation
line.
c. Pause to allow any liquid to run down the inside walls of the volumetric
flask.
d. Add the remainder of the liquid dropwise using a Pasteur pipette, until
the bottom of the meniscus is level with the graduation line.
e. Insert the cap into the volumetric flask.
28. Ensure the outside of the flask is dry, then record the mass of the capped
volumetric flask + water dispensed to the nearest 0.1 mg.
29. Repeat step 27 until the mass of water delivered is ± 150 mg each time
(concordant).

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 Mass of 50 mL volumetric flask + Mass of water contained in 50 mL
water / g volumetric flask / g
1
2
3
4
5

30. Calculate the mean of your concordant readings.

Mean mass of water contained in

50 mL volumetric flask / g

31. Use your mean mass and the density of water (determined earlier) to
calculate the mean volume of water dispensed.
mass
density =
volume

Mean volume of water contained in

50 mL volumetric flask / mL

32. Dispose of the contents of the volumetric flask appropriately.

33. Using the tables below, compare the results you have obtained with the
acceptable tolerances for grade A, and grade B volumetric glassware.

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Pipettes

Total Volume / mL 2 10 20 25 50 100

Grade A tolerance / mL 0.01 0.02 0.02 0.03 0.04 0.06

Grade B tolerance / mL 0.02 0.04 0.05 0.06 0.08 0.12

Burettes

Total Volume / mL 2 10 50 100

Grade A tolerance / mL 0.01 0.02 0.06 0.10

Grade B tolerance / mL 0.02 0.04 0.10 0.20

Volumetric Flasks

Total Volume / mL 10 20 25 50 100

Grade A tolerance / mL 0.02 0.04 0.04 0.06 0.10

Grade B tolerance / mL 0.05 0.06 0.06 0.10 0.20

Results
Equipment Grade A or Grade B?

10 mL pipette
25 mL pipette
Burette
50 mL volumetric flask

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Practical 1b: Standardisation of HCl using borax
 In this experiment you will calculation the accurate concentration of a supplied
solution of hydrochloric acid, by titrating it against a standard solution of borax.
When performing a titration, it is clearly important to be using solutions of accurately
determined concentration.

Borax
Sodium tetraborate decahydrate, Na2B4O7.10H2O (borax) does not decompose under
normal storage and can easily be obtained in high purity. These properties make
borax a useful compound for standardising other solutions.
In aqueous solution, borax dissociates by the following equation:

Then the B4O72- ion is hydrolysed:

The hydroxide ions (OH-) released can be used to titrate against strongly acidic
solutions, such as hydrochloric acid. For every 1 mole of borax in aqueous solution,
2 moles of hydroxide ion will be released.
Therefore, a standard solution of borax can be used to accurately determine the
concentration of a solution of hydrochloric acid by titration.

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Method
1. You are provided with approximately 8 g of borax in a glass vial.
a. Accurately record the mass of the borax, vial + lid.
b. Pour approximately one third of the borax (approx. 2.5 g) directly into a
250 mL conical flask.
c. Accurately record the mass of the borax, vial + lid.
d. Determine the mass of borax in the conical flask by difference.

Mass of stock Mass of stock Mass of borax in conical

vial before / g vial after / g flask / g

2. Calculate the number of moles of borax in the conical flask. The molar mass
of borax is 381.37 g mol-1.
3. Calculate the number of moles of HCl required to neutralise the borax.
4. Assuming that the HCl solution provided is 0.500 M, calculate the
approximate volume of HCl which you will be dispensing from the burette in
the upcoming titration.

Amount of borax in Amount of HCl needed to Volume of 0.500 M HCl

conical flask / mol neutralise / mol needed to neutralise / mL

5. Dissolve the borax in the conical flask using 80 mL of deionised water –

encourage dissolution by carefully heating where necessary. Do not boil, and
ensure the solution has cooled to room temperature before titration.
6. Rinse the burette, first with deionised water, then with the 0.5 M HCl solution.
Collect waste in a beaker.
7. Correctly fill your burette with the provided solution of approximately 0.5 M
hydrochloric acid.
a. Ensure the burette is vertical (from all sides).
b. Ensure the tap is closed.
c. Using a small funnel, carefully fill the burette with approx. 0.5 M HCl, to
just above the 0.00 mL line.
d. Remove the funnel.
e. Place a beaker underneath the burette, open the tap to ensure there is
liquid in the part of the burette below the tap.

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 f. Remove any air bubbles by gentle tapping of the burette, or
manipulation of the tap.
g. Ensure the level of liquid is below the 0.00 mL mark at the top of the
burette.
h. Leave to stand for 30 seconds, then take an initial reading from the
burette. Record all burette readings to the nearest 0.02 mL.
8. Add three drops of methyl red indicator solution to the borax solution.
9. Using your approximate volume calculated in step 4, titrate your borax
solution with the 0.5 M HCl solution. When approaching the endpoint, add
dropwise. Keep swirling the flask throughout. Repeat until concordant results
are obtained.

Initial burette reading / Final burette reading / Volume of HCl

mL mL dispensed / mL
1
2
3

10. Calculate the mean value of your concordant titres, and use this to calculate
the accurate concentration of the HCl. Remember to consider the
stoichiometry of the neutralisation.

Mean amount of borax in Mean of titres of HCl / mL Concentration of HCl / M

conical flask / mol

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Practical 1c: Standardisation of NaOH using HCl
 In this exercise we are attempting to determine the strength of a solution of
sodium hydroxide by titrating it with the hydrochloric acid that has been
standardized above.

Dissolution of CO2 in aqueous solutions

Aqueous solutions have a small amount of dissolved carbon dioxide. CO 2 in water
forms carbonic acid and therefore the pH of water exposed to the atmosphere is
below 7.
Sodium hydroxide solutions exposed to the air contain sodium carbonate.
In this experiment, we need to remove the sodium carbonate from the solution so
that we only titrate the sodium hydroxide. This is done by precipitating out the
carbonate with barium.
Once all of the hydroxide has been titrated and the endpoint found, we can continue
to add acid to titrate the carbonate.
Our titration has two endpoints:
1. The amount of acid added to reach the first endpoint equates to the amount of
hydroxide present.
2. The amount of acid added after the first endpoint to reach the second
endpoint equates to the amount of carbonate present.

Precipitating carbonate using barium

The addition of barium chloride quantitatively precipitates barium carbonate:

Provided this suspension is titrated slowly and with continuous thorough mixing, at
the phenolphthalein endpoint, only the OH- reacts:

By further similar titration to the bromophenol blue endpoint (pH 3.7), the precipitate
of BaCO3 also reacts with acid:

The overall titration measures the total alkali present; the difference between the
endpoints measures the carbonate.

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Method
1. Accurately pipette 25 mL of the NaOH solution into a conical flask.
2. Add 5 mL BaCl2 solution.
3. Add 2 drops of phenolphthalein solution.
4. Titrate the suspension against your standardised HCl solution. Use the table
below to record your burette readings.
5. Record the titre required to reach the phenolphthalein endpoint.
6. Add 5 drops of bromophenol blue solution.
7. Continue the titration to the bromophenol blue endpoint.
8. Calculate:
a. The moles of HCl which neutralised the NaOH.
b. The concentration of NaOH in the solution.
c. The percentage w/v of sodium carbonate, Na 2CO3 in the solution (ie.
how many grams per 100 mL).

Initial burette Final burette Volume of HCl Indicator

reading / mL reading / mL dispensed / mL
1 Phenolphthalein

2 Bromophenol blue

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Practical 1d: Assay of aspirin tablets
 In this experiment we will determine the amount of aspirin in aspirin tablets by a
British Pharmacopoeia method.

British Pharmacopoeia Standards

Because of the way they are made, tablets are liable to vary in weight and in content
of active ingredient.
Among the standards laid down (see B.P. 1993, Vol II, page 753) for tablets are the
following:
1. Uniformity of weight. Normally a sample of 20 tablets, randomly selected, is
used to obtain the average weight per tablet. For the tablets assayed in this
exercise (250 mg), not more than two of the individual weights deviate from
the average weight by more than 5% and none deviate more than twice this
amount (10%).
2. Content of active ingredient. Normally a sample of 20 tablets is used to
ascertain the weight of active ingredient in one tablet of average weight. The
weight of active ingredient is then expressed as a percentage of its stated
weight. The percentage limits are stated in the appropriate Pharmacopoeial
monograph and reflect all permissible variations in the active component and
in the tabletting process.

Aspirin Assay - British Pharmacopoeia

The phenolic ester in acetylsalicylic acid (aspirin) can be hydrolysed by heating with
an excess of NaOH:

One equivalent of hydroxide hydrolyses the ester, and a subsequent equivalent of

hydroxide will neutralise the carboxylic acid group in salicylic acid to give salicylate.
During this hydrolysis, a small amount of alkali reacts with the glassware. To correct
for this, a ‘blank’ titration is performed under identical conditions but in the absence
of the sample. Provided equal quantities of identical strength alkali are used in both
the blank and the sample determinations, neither the precise strength nor the volume

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of the alkali need be known. The volume of NaOH which reacted with the aspirin will
simply be the difference between the blank titration and the aspirin titration.

Method
See B.P. (1993) Vol II, page 778
1. You are provided with powdered aspirin tablets equivalent to three tablets.
a. Record the mass of the ‘three’ powdered aspirin tablets.
b. Weigh accurately (by difference), approximately half of the powder into
a 250 mL conical flask.
c. Add 25 mL of standardised NaOH solution.
d. Insert a funnel into the neck of the conical flask to act as a spray trap.
e. Boil the mixture gently in the flask for 10 minutes.
f. Cool.
g. Rinse the funnel.
h. Titrate the solution against a standardised solution of HCl using phenol
red as an indicator until concordant titres obtained.
2. Repeat step 1c-h as a ‘blank’.
3. If you have time (check with a member of staff), you can repeat the sample
titration with the remaining aspirin powder.
4. The molar mass of aspirin is 180.16 g mol-1. Calculate:
a. The amount of NaOH which reacted with the glassware.
b. The amount of NaOH which reacted with aspirin.
c. The amount of aspirin in the conical flask.
d. The mass of aspirin in the conical flask.
e. The mass of aspirin per powdered tablet.
f. The mass of aspirin per powdered tablet as a percentage of the stated
amount (250 mg).
5. Is your sample of aspirin tablets within the specification of the British
Pharmacopoeia?

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