PHARMACEUTICAL SCIENCES (PHARMACEUTICAL ANALYSIS)

UNIT -1
The pharmaceutical analysis is a branch of chemistry, which involves the series of process for
the identification, determination, quantitation, and purification. This is mainly used for
the separation of the components from the mixture and for the determination of the structure
of the compounds.
The different pharmaceutical agents are as follows:
1. Plants
2. Microorganisms
3. Minerals
4. Synthetic compounds
Scope:
 It is used to determine the identity of the drug in the formulated product correctly.
 It is used to calculate the percentage of the stated content of a drug present in a
formulation.
 Does this formulation contain solely the active ingredient, or are additional impurities
present/not present?
 To ascertain the stability of a drug in the formulation and hence the shelf-life of the
product.
 To determine the rate at which the drug is released from its formulation so that it can be
absorbed by the body.
 It can determine if the identity and purity of a pure drug substance used in the preparation
of a formulation meet specifications.
 To define the concentrations of specified impurities in the pure drug substance.
 To confirm the concentration of the drug in a sample of tissue or biological fluid.
 To perform the pKa values, partition coefficients, solubilities, and stability of a drug
substance under development.
Based upon the determination type, there are mainly two types of analytical methods. They
are as follows:
1. Qualitative analysis: This method is used for the identification of chemical compounds.
2. Quantitative analysis: This method is used for the determination of the amount of the
sample.

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Non-Instrumental methods of Analysis Instrumental methods of

Analysis.
Non -Instrumental Methods of Analysis
S.No Name of the Method Applications
Gravimetry Analysis
1 Ex: Gravimetry, Electro gravimetry, Quantitative Estimation of Weight
Thermo gravimetry
Titrimetric Analysis
Ex:
2 Detection of Equivalent point.
Acidimetry, Alkalimetry,
N.A.T(Non-Aqueous Titration)
Oxidation -Reduction titration
Ex:
Transfer of electrons between two
Permanganometry
species i.e., determine the
Potassium iodate
3 concentration of a given analyte by
Potassium bromates
causing a redox reaction between the
Cerimetry
titrant and analyte.
Iodimetry
Iodometry
Precipitation titration
Ex:
Mohr’s method Determination of halide ions the
4
Volhard Method solution
Faja n’s Method
Gay -Loussac Method

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5 Complexometric Titrations Determination of the metal ions

6 Gasometry Measurement of volume of gases
Miscellaneous methods a. comparison of the inhibition of the
a. Microbiological methods growth of bacteria by measured
b. biological methods concentration of antibiotics.
Ex: cylinder plate (cup plate) and
7
turbimetric (tube assay) method.
b. sample and standard preparation
are tested under identical conditions.
Ex: Response of drug.
Instrumental Analysis
Optical methods
Ex:
a. Spectrophotometry (UV Visible) Study of drugs/ analytes with
1 b.IR Electromagnetic radiation, i.e.,
c. atomic absorption spectroscopy ABSORPTION
d. Refractometry

Electrical methods
Electrical potential
a.Potentiometry
b.Conductometry Electrical conductance
2
c.Voltametry Electrical current
d.Coulometry
e.Polarography Electrical current
Emission methods
Emission of radiation
a.Emission spectroscopy
3
b.Fluorimetry Emission of fluorescence
c.flame photometry Flame Emission spectroscopy
Other techniques
Diffraction of solids
4 a.X-ray spectroscopy
b.Mass spectroscopy Charge to mass ratio

Titrimetric Analysis Principle

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An analyte is chemically reacted with a standard solution of a reagent of precisely known

concentration or with a concentration that can be precisely determined. The amount of a
standard solution required to completely react with all of the samples is used to estimate the
purity of the sample.
Applications
 Provide standard pharmacopeial methods for the assay of unformulated drugs and
excipients and some formulated drugs, e.g., those that lack a strong chromophore.
 Used for standardizations of raw materials and intermediates used in drug synthesis
in industry. Suppliers of raw materials may provide these materials at a specified purity
which has been assayed titrimetrically to a pharmacopoeia standard.
 Certain specialist titrations, such as the Karl Fischer titration used to estimate water
content, are widely used in the pharmaceutical industry.
Advantages
Capable of a higher degree of precision and accuracy than instrumental analysis methods,
with precisions of ca 0.1% being achievable.
 The methods are generally robust.
 Analyses can be automated.
 Cheap to perform and do not require specialised apparatus.
 They are absolute methods and are not dependent on the calibration of an instrument.
Limitations
 Non-selective.
 Time-consuming if not automated and require a greater level of operator skill than routine
instrumental methods.
 Require large amounts of sample and reagents.
 Reactions of standard solutions with the analyte should be rapid and complete
Fundamentals of Volumetric Analysis

Volumetric Analysis

It’s one of the useful analytical techniques. It’s fairly rapid and very good accuracy can be
obtained.

Titration:

Volumetric analysis is carried out by a process known as titration.

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Titrant: The Reagent Know Concentration.

Titrate: The substance being titrated.

1. The reaction should be simple & expressed by a well-defined chemical reaction/equation.

2. The reaction should be rapid.
3. The reaction must proceed to completion when an equivalent amount of standard solution
has been added. This gives satisfactory end point detection.
4. There must be some sharp change in either physical or chemical properties of the solution
at the equivalent point.
5. Reaction should have some simple method for the detection of end point or equivalence
point of titration.
Classification of Volumetric chemical reactions
1. Neutralization (Aqueous -acid base) titrations
2. Non aqueous titrations
3. Precipitation titrations
4. Complexometric titrations
5. Redox titrations

Neutralization titrations

It involves neutralisation reaction in presence of water as solvent.

Non-aqueous titrations

It involves neutralisation reaction between acid & base in presence of non-aqueous.

Precipitation titrations

It involves the reaction leading to formation of precipitation.it includes the methods where
the reacting substance & standard solution react to yield a precipitate or a slightly soluble salt
as primary reaction product.

NaCl + AgNO3 →NaNO3 + AgCl

Complexometric Titration

Where in the reacting substance and the standard solution react to form a soluble but very
slightly dissociated complex substance.

It is based on complex formation reaction mainly EDTA titrations.

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Redox titration

Involves simultaneous oxidation reduction reactions. It includes all the methods wherein
reacting substance is oxidized or reduced by the standard solution.

PRIMARY STANDARDS
Primary standards are stable chemical compounds that are available in high purity and which
can be used to standardize the standard solutions used in titrations. Titrants such as sodium
hydroxide or hydrochloric acid cannot be considered primary standards since their purity is
quite variable.

Property Primary Secondary

High
Purity Low
Ex: 99.95 -100.5%
Stability Stable Unstable
Oxidation by air No Yes
Hygroscopic No Yes
Molecular weight High Low
Solubility Good Poor
Heat Condition 110 -120º C ….

Primary Standard Uses

Potassium hydrogen
Standardization of sodium hydroxide solution
phthalate
Potassium hydrogen
Standardization of acetous perchloric acid
phthalate
Standardization of sodium thiosulphate solution through the
Potassium iodate
generation of iodine
Anhydrous sodium
Standardization of hydrochloric acid
carbonate
Zinc metal, CaCo3, Mg
Standardisation of EDTA solution
SO4

EDTA: Ethylene diamine tetra acetic acid.

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Secondary Standards:
a chemical that has been standardized against a primary standard. In the example that we
discussed, the NaOH was standardized against a primary standard. Once the exact
concentration of the NaOH is determined and is used as a titrant in another experiment, it
becomes a secondary standard. Secondary standards are usually used to calibrate analytical
methods.
Ex: HCl, NaOH
Sodium Hydroxide; absorbs CO2 from atmosphere
HCl: produce chlorine gas in reactions and liberate hydrogen when exposed to air
Nitric acid: can act as an oxidising agent interfere ring with reactions
Sulphuric acid: absorb water from the air
Solubility Condition
Depends on 1. Temperature2. Pressure 3. Type of bond and forces between the particle.
Description ml/g of solute
Very soluble <1 part of solvent
Freely soluble From 1 to 10
Soluble From 10 to 30
Sparingly soluble From 30 t0 100
Slightly soluble From 100 to 1000
Very slightly soluble From 1000 to 10,000
Practically insoluble or insoluble More than 10,000
Supersaturation:
The solubility of a chemical substance at any given temperature in a given solution is
the amount of the substance dissolved by known weight of the solvent when substance
is in equilibrium with the solvent. A super saturated solution is one that contains a
greater concentration of solute than that corresponds to the equilibrium solubility at
given temperature.

Methods of Concentration Expression

Formality or Formal Concentration (F):
Some substances do not exist in molecular form whether in solid or solution form. They
remain in ionic form in solid state as well as in solution.

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The formality, of a solution expresses the number of gram- formula weights of solute
present in one liter of solution.
𝑁𝑜. 𝑔𝑟𝑎𝑚 𝑓𝑜𝑟𝑚𝑢𝑙𝑎 𝑚𝑎𝑠𝑠 𝑜𝑓 𝑠𝑜𝑙𝑢𝑡𝑒
𝐹=
volume of solution in litre

OR

𝐺𝑖𝑣𝑒𝑛 𝑤𝑒𝑖𝑔ℎ𝑡 𝑜𝑓 𝑠𝑜𝑙𝑢𝑡𝑒

Formality =
Molecular weight of solute x Volume of solution in litre
Example:
5.85 g of NaCl dissolved in 250ml solution.
Weight of Nacl -5.85 g
Mwt of Nacl = 58.5 g/mol
Volume of solution – 250 ml
5.85
Formality =
58.5g mol−1 x 0.250L
Formality = 0.4 F

Mole Concept:
The mole is the unit of measurement in the International System of Units (SI) for amount
of substance. It is defined as the amount of a chemical substance that contains as many
elementary entities (e.g., atoms, molecules, ions, electrons, or photons). This number is
expressed by the Avogadro constant, which has a value of 6.022140857×1023 mole−1. The
mole is one of the base units of the SI, and has the unit symbol mol.
Example:
Calculate the mass of 6.022 × 1023 molecule of Calcium carbonate (CaCO3).
Solution- -
Molar mass (Molecular mass in gram) of CaCO3 = 40+12+3×16 = 100g
No. of moles of CaCO3
= No. of molecules/Avogadro constant
= 6.022 × 1023/ 6.022 × 1023
= 1 mole
Mass of CaCO3
= No. of moles × molar mass
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= 1 × 100 g = 100 g.
Example:
Calculate the mass of 12.044 × 1023 carbon atoms.
Solution -
No. of moles of Carbon atoms
= No. of atoms/Avogadro constant
= 12.044 × 1023/6.022 × 1023
= 2 moles
Mass of carbon atoms
= No. of moles × atomic mass
= 2 × 12
= 24 g.
Molarity / Molar concentration (M):
The number of gram-molecular weights (or moles) of a solute in one litre of solution.
𝑀𝑜𝑙𝑒𝑠 𝑜𝑓 𝑆𝑜𝑙𝑢𝑡𝑒
Molarity =
Litres of solution

For small amounts of solute may expressed, i.e., mM.

1mM = 1 X 10-3M

Normality (N):
The number of equivalents of dissolved solute in one liter of solution.
𝑊𝑒𝑖𝑔ℎ𝑡 𝑜𝑓 𝑆𝑜𝑙𝑢𝑡𝑒
Normality =
Equivalent Weight of solute X Vol. of solution

Equivalent Weight = Mwt/ neither H+/OH-

a). Acids equal a no. of H+ ions that react.
1. Inorganic acids n = no. of H in formula (HCl=1; HNO3=1; H3PO4=3; H2SO4=2;
H2CO3=2).
2. Organic acids n = no. of –COOH in formula (CH3COOH=1; HOOC-CH2-CH2-
COOH=2).
b). Bases equal a n =no. of OH-ions that react (NaOH =1; NH4OH=1; Al (OH)3=3; Ba
(OH)2=2).
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c). Salts equal a no. of metals ion multiplied by the number of its oxidants: - {NaCl, KI,
AgNO3=1}; {Na2CO3, Ca (NO3)2, BaSO4, CaHPO4 = 2}; {Na3PO4, Al (NO3)3, Na3AsO4 =
3}; Ca3 (PO4)2= 6; Al2(SO4)3 =6; NaHCO3 = 1
d). Redox reagents equal a no. of electron moles in reaction or the change in oxidation
number of reagent:
Fe +3 + e- → Fe +2 n=1
Sn +2 → Sn +4 + 2e- n=2
e). Ions equal a no. of ion charge: (Na+, NO3-, HCO3-, H2PO4-, K+ =1), (Ca2+, Mg2+, SO42-,
CO32- =2) (PO43-, Al3+ = 3)
For all reactions, from chemical equation balanced equal the number of moles of the
substance interacting with it:
H3PO4+ NaOH→NaH2PO4+H2O n=1
H3PO4+2NaOH→Na2HPO4+2H2O n=2
H3PO4+3NaOH→Na3PO4+3H2O n=3
5C2O4 = + 2MnO4 - + 16H+ 10CO2 + 2Mn+2 + 8H2O
n of KMnO4 = 5; n of C2O4 = 2
Molality(m):
The number of moles of solute in 1000gm of solvent.
𝑚𝑜𝑙𝑒𝑠 𝑖𝑛 𝑔𝑟𝑎𝑚
Molality =
Molecular Weight
Note: Concentration is used for the study of thermodynamic properties of solutions.
Expression of Strengths
Number of grams of solute in 100g of product
1 %W/W
Ex: H2SO4 -98%, CH3COOH 33% W/W etc.
Number of grams of solute in 100ml of product
2 %W/V
Ex: H2O2 , 5-7% w/v, Bacl2 solution 10% w/v etc.
Number of ml of solute in 100ml of product
3 %V/V
Ex: Alcohol 95% v/v
4 %V/W Number of ml of solute in 100gof product
106 i.e., parts per million
5 PPM
Ex: Limit tests- Chloride, Sulphate, Iron, Lead, Arsenic, Heavy metals.

Preparation & Standardization of molar and Normal solutions

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Common Formulas for calculation of Mass by Molar/ Normal

Molar Solution(s) Normal Solution(s)
𝑴𝒐𝒍𝒆𝒔 𝒐𝒇 𝒔𝒐𝒍𝒖𝒕𝒆 𝑴𝒐𝒍𝒆𝒄𝒖𝒍𝒂𝒓 𝑾𝒆𝒊𝒈𝒉𝒕
M = 𝑳𝒊𝒕𝒆𝒓𝒆𝒔 𝒐𝒇 𝑺𝒐𝒍𝒖𝒕𝒊𝒐𝒏 EWT = 𝑵𝒖𝒏𝒃𝒆𝒓 𝒐𝒇 𝒓𝒆𝒑𝒍𝒂𝒄𝒆𝒃𝒍𝒆 𝒂𝒕𝒐𝒎(𝒙)

X --- H+ or OH- atoms

𝑴𝒐𝒍𝒆𝒄𝒖𝒍𝒂𝒓 𝑾𝒆𝒊𝒈𝒉𝒕 𝑿 𝒙 𝒓𝒆𝒒𝒖𝒊𝒓𝒆𝒅 𝑴𝒐𝒍𝒂𝒓 𝒙 𝑽𝒐𝒍 (𝒎𝒍) 𝑬𝒒𝒖𝒊𝒗𝒂𝒍𝒆𝒏𝒕 𝒘𝒆𝒊𝒈𝒉𝒕 𝑿 𝒙 𝑵𝒐𝒓𝒎𝒂𝒍𝒊𝒕𝒚 𝒙 𝑽𝒐𝒍 (𝒎𝒍)
Mass = N=
𝑳𝒊𝒕𝒆𝒓𝒆𝒔 𝒐𝒇 𝑺𝒐𝒍𝒖𝒕𝒊𝒐𝒏 𝒊.𝒆.,𝟏𝟎𝟎𝟎𝒎𝒍 𝑳𝒊𝒕𝒆𝒓𝒆𝒔 𝒐𝒇 𝑺𝒐𝒍𝒖𝒕𝒊𝒐𝒏 𝒊.𝒆.,𝟏𝟎𝟎𝟎𝒎𝒍

Standardization:
Standardization is the process of determining the exact concentration
(molarity) of a solution. Titration is one type of analytical procedure often
used in standardization.
Equivalent Weight Calculation for Acids
𝑀𝑜𝑙𝑒𝑐𝑢𝑙𝑎𝑟 𝑊𝑒𝑖𝑔ℎ𝑡
Equivalent Weight =
No.of replaceble H atom

Hydrochloric Acid:
= 36.47/1
= 36.47 g
Sodium Carbonate:
Na2CO3 + 2HCl 2 NaCl +H2O +CO2
106 gm Na2CO3 = 2000 ml N
53 gm Na2CO3 = 1000ml N HCl
106
Equivalent Weight = = 53 𝑔𝑚
2
Sulphuric acid:
98
Equivalent Weight = = 49 𝑔𝑚
2
Oxalic Acid:
Molecular Formula: C2H2O4

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126
Equivalent Weight = = 63 𝑔𝑚
2
Equivalent Weight Calculation for Bases:
𝑀𝑜𝑙𝑒𝑐𝑢𝑙𝑎𝑟 𝑊𝑒𝑖𝑔ℎ𝑡
Equivalent Weight =
No. of replaceble OH atom
Sodium Hydroxide:
Molecular Weight = 40
40
Equivalent Weight = = 40 𝑔𝑚
1
Barium Hydroxide:
315.47
Equivalent Weight = = 157.7 𝑔𝑚
2
SODIUM HYDROXIDE(NaOH)
Oxalic Acid (Primary Standard)
Molecular Formula: C2H2O4. 2H2O
Molecular Weight: 126 g/mol-1
Mass Calculation of oxalic acid:
12 x 2 = 24 (C atoms 2) 24 + 6+ 96 = 126
1 x 6 = 6 (H atoms 6)
16 x 6 =96 (O atoms 6)
Preparation and standardisation of 0.1N NaoH solution
Principle:
Sodium hydroxide is an example of strong base but has secondary standard property
because of hygroscopic property and absorb moisture and contains impurities like Cl
and So4.So that concentration of it has not maintain pure state. Standardise the NaoH using
Primary standard as oxalic acid and detect end point is phenolphthalein (weak acid)
indicator solution.
Primary Standards:

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OR

Method -I
Standardisation Procedure:
Transfer 10 ml of Oxalic Acid solution & 1-2 drops of phenolphthalein solution were
in a conical flask. To that solution, NaOH solution was added until the solution
became just pink from burette. The titration was performed 3 times along with 3 blank
titrations. Volume of NaOH solution used in blank titration was subtracted from the
titration value of NaOH to obtain the volume of NaOH solution used against Oxalic
acid.
Method -2
1. Weigh accurately between 0.3 and 0.4 g of KHP into a tared plastic weighing dish.
2. Transfer it quantitatively to the clean 250-mL Erlenmeyer flask.
3. Add enough water to bring the total volume to about 50 to 75 mL.
4. Swirl the flask and rinse down the sides of the flask to dissolve the sample. Add
magnetic stirring bar to mechanically stir the solution.
5. Add 2 or 3 drops of phenolphthalein indicator to the flask.
6. Titrate the sample of KHP until the faint, pink endpoint is reached.
Reagent Preparations
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Preparation of 0.1N NaOH Solution:

4g of NaoH in 1litre water.
Preparation of 0.1N oxalic acid Solution:
6.3g of oxalic acid in 1 litre solution.
Calculation
N NaOH V NaOH = N Oxalic Acid V Oxalic Acid Here
V NaOH = volume used in titration (A) - volume used in blank titration (B) = N Oxalic
Acid = 0.1 V Oxalic Acid = 10
Thus, N NaOH = N Oxalic Acid V Oxalic Acid = 0.1 X 10 = ……. N V NaOH …….
HYDROCHLORIC ACID(HCL)(Mwt:36.5,Specific Gravity:1.18)
Preparation and standardisation of 0.1MHCl solution
Principle:
HCl is gaseous form at room temperature condition so that there is no stable
concentration. This is standardised against primary stand as anhydrous sodium carbonate
for determination of its strength. Methyl orange (Weak acid) is indicator (weak base).
Na2 CO3 + 2 HCl NaCl + CO2 + H2O
Standardisation Procedure:
Take 0.53g of sodium carbonate and dissolve in 100ml water and transfer 10ml this
resultant solution into conical flask and add 2-3 drops of methyl orange indicator. Titrate
against 0.1M hydrochloric acid solution.
Each ml of 0.1M Hcl = 0.005299 g of Na2 CO3
Equivalent factor
53 g of Na2 CO3 = 1000ml of Na2 CO3
Each ml of 0.1M Hcl = 0.005299 g of Na2 CO3
𝑊𝑒𝑖𝑔ℎ𝑡 𝑜𝑓 𝑁𝑎2 𝐶𝑂3 𝑥 𝐸𝑥𝑝𝑒𝑐𝑡𝑒𝑑 𝑀𝑜𝑙𝑎𝑟𝑖𝑡𝑦
Molarity of Hcl= 𝑇𝑖𝑡𝑟𝑒 𝑉𝑜𝑙𝑢𝑚𝑒 𝑥 𝐸𝑞𝑢𝑖𝑣𝑎𝑙𝑒𝑛𝑡 𝑓𝑎𝑐𝑡𝑜𝑟

Reagent Preparation
Preparation of 0.1M HCl Solution:
8.5 ml Hcl in 1 litre water.
Explanation
% Purity of Hcl – 36.5
Specific gravity = 1.18
36.5 X 1.18 = 43.07 g /100mL
1000……...?
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1000 𝑥 43.07
= 𝟒𝟑𝟎. 𝟕 𝒎𝑳
100
𝑾𝒆𝒊𝒈𝒉𝒕 𝒐𝒇 𝒔𝒐𝒍𝒖𝒕𝒆
𝑴𝒐𝒍𝒂𝒓𝒊𝒕𝒚 𝒐𝒇 𝒔𝒐𝒍𝒖𝒕𝒊𝒐𝒏 =
𝐌𝐨𝐥𝐞𝐜𝐮𝐥𝐚𝐫 𝐖𝐞𝐢𝐠𝐡𝐭
430.7
= = 𝟏𝟏. 𝟖𝒎𝑳
36.5
1000
= = 𝟖𝟓𝒎𝑳
11.8
Short Cut Method:
𝑀𝑜𝑙𝑒𝑐𝑢𝑙𝑎𝑟 𝑊𝑒𝑖𝑔ℎ𝑡
= 𝑋 100
𝑆𝑝𝑒𝑐𝑖𝑓𝑖𝑐 𝐺𝑟𝑎𝑣𝑖𝑡𝑦 𝑥 𝑃𝑢𝑟𝑖𝑡𝑦
Explanation
36.5
= 𝑥 100
1.18 𝑥 36.5
3650
= = 𝟖𝟒. 𝟕𝟒 𝒎𝑳
43.07
Preparations of 0.1M Sodium carbonate Solution:
Dissolve 10.6 g of Sodium carbonate in 1 litre water.
PREPARATION & STANDARDIZATION OF 0.1 N SULPHURIC ACID (Mwt:98,
Specific gravity:1.8)
PRINCIPLE:
This is direct acid base titration& Alkalimetry in which sulphuric acid (diprotic acid)
reacts with sodium carbonate to form CO2 and water. As CO2 formed in reaction can
change colour of the indicator before equivalence point it is advisable to boil solution to
perform the errorless titration.
Na2CO3 + H2SO4--------- >Na2SO4+CO2 +H2O
PROCEDURE:
PREPARATION OF 0.1N SULPHURIC ACID (Mwt:98.079, Specific gravity:1.8)
Add slowly, with stirring, 0.56ml of sulphuric acid to about 100ml of distilled water allow
to cool 25 ̊ and standardize.
SAFETY TIP
1. Sulphuric acid in higher concentration forms acidic mist. Both the mist and solution
have a corrosive effect on tissues with the potential to damage respiratory organs, eyes,
skin and intestine.
2. While preparing Sulphuric acid solution, take water in a beaker and add the acid drop by
drop with mixing. If you do the reverse it will burst.
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STANDARDISATION PROCEDURE:
Weigh accurately about 0.53g of anhydrous sodium carbonate previously heated at about
270̊ for one hour. Dissolve it in 100ml of water and add 0.1ml of methyl orange solution.
Add the acid slowly from a burette, with constant stirring until the solution become faintly
pink. Heat the solution to boiling, cool and continue the titration. Heat again to boiling and
titrate further as necessary until the red colour is no longer affected by continued boiling.
Calculation
N1V1 = N2V2
N1 = 0.1 N = Normality of Na2CO3 Solution, N2 = ? = Normality of H2SO4
V1 = 10 ml = Volume of Na2CO3 Solution, V2 = x ml = Volume of H2SO4

PREPARATION AND STANDARDISATION OF 0.1M SODIUM THIOSULPHATE

Principle:
Sodium thiosulphate is a secondary standard which is standardized against the primary
standard potassium dichromate. Potassium dichromate in acid solution is reduced by
potassium iodide to liberate an equivalent amount of iodine which in turn reacts with
sodium thiosulphate. Free iodine forms blue colour with starch. At the end point all the
iodine reacts with thiosulphate which causes disappearance of blue colour. In this titration
the dichromate ion gets converted to green chromic salt which gives green colour at the
end point.
REACTION:
K2Cr2O7 + 6KI + 14HCl →2CrCl3 + 3I2 + 8KCl + 7H2O
I2 +2Na2S2O3 → Na2S2O6 + NaI
PROCEDURE:
Preparation of 0.1M Sodium thiosulphate pentahydrate (Mwt:248.18 g/mol)
Dissolve 2.5 g of sodium thiosulphate and 0.02 g of sodium carbonate in carbon dioxide-
free water and dilute to 100 ml with the same solvent.
Note:
Sodium thiosulphate is a reducing agent and efflorescent in nature. If the solution of
sodium thiosulphate is prepared by using distilled water, the excess carbon dioxide present
in the distilled water may decompose the thiosulphate with the formation of sulphur. This
decomposition also caused by bacterial action.
Remedy: Use freshly boiled and cooled water to expel carbon dioxide

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Add a small amount of sodium carbonate to maintain pH between 9 and 10 at which

bacterial activity is least.
Standardization of 0.1M sodium thiosulphate
Weigh accurately 1.2 g of potassium dichromate (294.185g/mol-1) and dissolve in 250 ml
of distilled water. From the solution, pipette out 25 ml and transfer into an iodine flask.
Dilute the solution with 50 ml of water and add 2 g of KI, 5 ml of concentrated
hydrochloric acid. Stopper the flask and allow to stand for 5 minutes. Dilute the contents
with 50 ml of water and titrate with sodium thiosulphate solution until the colour becomes
light yellow. Further add 1 ml of starch solution. The solution turns to blue colour.
Continue the titration until the blue colour disappears green colour appears.
PREPARATION AND STANDARDISATION OF 0.02M
POTASSIUMPERMANGANATE (Neutral Salt) Molecular Weight:158.034g/mol-1
Theory:
Potassium permanganate often contains a small proportion of manganese dioxide;
volumetric solutions must be made up approximately and then standardized. The intense
colour of the solution makes difficult the detection of undissolved solid. The use of
heat in the preparation of potassium permanganate solutions is also undesirable, since
traces of other contaminants on the glass vessels used can catalyse its decomposition.
Potassium permanganate may be standardized with either sodium oxalate or oxalic acid.
The former is preferred because it is readily available to higher standard of purity (99.95%)
and unlike oxalic acid, it is available in the anhydrous state. Redox titration is based on
change in electrode potential of sample during redox titration, i.e., gain of electrons or
decrease in oxidation number.
Principle
Potassium permanganate acts as a powerful oxidising agent. Although KMnO4 acts as an
oxidising agent in alkaline medium also, for quantitative analysis mostly acidic medium is
used. The oxidising action of KMnO4 in the acidic medium can be represented by the
following equation:
MnO4 - + 8H+ + 5e- →Mn2+ +4H2O
The acid used in this titration is dilute sulphuric acid. Nitric acid is not used as it is itself
an oxidising agent and hydrochloric acid is usually avoided because it reacts with
KMnO4 according to the equation given below to produce chlorine and chlorine which is
also an oxidising agent in the aqueous solution.
2KMnO4 - + 16HCl →2KCl +2MnCl2 +5Cl2 +5H2O
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Since, oxalic acid acts as a reducing agent, it can be titrated against potassium
permanganate in the acidic medium according to the following equation:

PROCEDURE:
PREPARATION OF 0.02 M POTASSIUM PERMANGANATE SOLUTIONS:
Dissolve 3.2g of potassium permanganate in 1000ml of water, heat on a water bath for 1
hour; allow standing for 2 days and filtering through glass wool. Store protected from light.
STANDARDISATION PROCEDURE:
METHOD A: With SODIUM OXALATE:
Weigh out sodium oxalate(134g/mol-1) about 6.7g accurately into a Liter graduated flask,
dissolve in water and make up to the volume. Pipette out 20ml of this solution add
concentrated sulphuric acid about (5ml) and warm to about 70ͦ. Add the potassium
permanganate solution from the burette. The first few drop results in a pink colour
persisting for about 20 seconds. Wait until the colour disappears and then continue the
titration. Formation of a brown colour during the titration is caused by insufficient acid,
by using too high a temperature or by the use of dirty flask. The end point is reached when
a faint pink colour persists for about 30 seconds upon the shaking the flask. 1ml of
0.02M potassium permanganate is equivalent to 0.0067g of sodium oxalate.
METHOD B: with SODIUM THIO SULPHATE:
PREPARATION OF 0.1M SODIUM THIOSULPHATE:
Dissolve 25g of sodium thiosulfate and 2.0g of sodium carbonate in CO 2 free water and
dilute to 1000ml with the same solvent. Standardize the solution in the following manner:
STANDARDISATION PROCEDURE:
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The standardization depends upon the reactions expressed as follows:

Principle Chemical Reaction:
8 H2SO4 + 2 KMnO4 + 5 Na2C2O4 → 2 MnSO4 + 10 CO2 + K2SO4 + 5 Na2SO4 + 8 H2O
STANDARDISATION OF KMnO4 PROCEDURE:
To 25.0ml of the solution in a glass-stoppered flask add 2g of potassium iodide, followed
by 10ml of 1M sulphuric acid. Titrate the liberated iodine with 0.1M sodium thiosulphate,
using 3 ml of starch solution, added towards the end of the titration, as indicator. Perform a
blank determination and make necessary correction.
1ml of 0.1M sodium thiosulphate is equivalent to 0.003161 g of KMnO4.
PREPARATION AND STANDARDISATION OF 0.1 M CERRIC AMMONIUM
SULPHATES (Mwt:632.55g/mol-1)
Principle
Cerium sulphate is a power full oxidizing agent in acid solution (1-8N). It is bright yellow
colour and corresponding cerium salt form by reduction is colourless strong solutions are
self-indicating. However, since dilute solutions are used hence indicators are used for
observation of end point. Arsenic trioxides used as primary standard in presence of
sulphuric acid and osmic acid using ferroin sulphate as an indicator for standardization of
the solution. The standardization depends on the reactions expressed by the following
equations.

PROCEDURE:
Preparation of 0.1M ceric ammonium sulphate:
Dissolve 65g of ceric ammonium sulphate, with the aid of gentle heat, in a mixture of 30ml
of sulphuric acid and 500ml of water. Cool, filter the solution, if turbid, and dilute to
1000ml with water. Standardize the solution in the following manner.
Standardization of 0.1M Ceric ammonium sulphate:
Weigh accurately about 0.2 gm arsenic trioxide of previously dried at 105ºC for 1 hour and
transfer to a 50 ml conical flask. Add 25 ml of 8% w/v sodium hydroxide, swirl and
dissolve contents in 100ml of water and add 30ml of dilute sulphuric acid ,0.15ml osmic

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acid, 0.1 ml of ferroin sulphate solution and slowly titrate with the ceric ammonium
sulphate solution until the pink colour is changed to pale blue colour,
Each ml of 0.1M Ceric Ammonium sulphate is equivalent to 0.004946g of As2O3.
SOURCE OF ERRORS
Error: Defined as deviation or numerical difference from the absolute value or from the
true average of large number of results.
Types of errors:
1.Determinate (CONSTANT ERRORS) 2. Indeterminate
Determinate (Constant/ Systemic Errors):
These errors can be avoided or whose magnitude can be determined.
i) Operational and Personal Errors:
These kinds of errors related to personally (experimenter) responsible. Do not depend on
the SOP /Methodology. They are arising from the erratic personal judgement, prejudice,
bias or inability.
Example:
Colour solution
Judgement of the colour of solution at the end point in titration.
Analytical balance
Estimation of the pointer position of a balance between two scale divisions.
Meniscus Level
Judgement of the level of meniscus with respect to graduation on a burette or a pipette.
Gravimetry
Mechanical loss of a material in gravimetric exercise in its various steps:
1. Under washing of precipitate
2. Excess washing of precipitate
3. Ignition of precipitates at incorrect temperature
4. Insufficient cooling of crucibles before weighing
5. Excess use of reagent than required
6. Allowing hygroscopic substance to absorb moisture before or during weighing etc.,
ii) Instrumental And Reagent Errors:
Arise from the imperfections in the measuring device.
Example:
Balance

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Unadjusted chemical balance and use of uncalibrated weights.

Glassware
Graduated glass ware and other instruments.
Chemical reaction
Reaction with glass ware Ex: caustic alkalis
Temperature:
Volatilization of platinum
Low grade reagent usage & instruments prone to determinate errors because of fluctuations
in voltage etc.
iii) Method Errors:
Arise from the incorrect sampling and incomplete reactions involved in a determination.
Ex: Gravimetry.
Solubility of precipitates, co-precipitation, post precipitation, decomposition or
volatilization of weighing substance on ignition.
Volumetric Analysis:
Due to failure of a reaction to proceed to completion, occurrence of side and induced
reactions etc.
Ex: Iodometric determination of copper, iodine gets adsorbed on the starch particles,
which is difficult to remove completely.
Further classification of determinate errors and proportional errors
Constant Errors:
It is independent of the magnitude of the measured quantity and becomes less significant as
the magnitude increases.
Ex:
0.10 ml this indicates a relative error of 1% for 10 ml titrant and 0.2% for 50ml of titrant.
Note: constants would increase with the decreasing quantity of the substance being
measured.
Proportional Errors:
Arise from sample (interfering impurities). This is varying with sample size.
Ex: Iodometric titration
Copper -II is contaminated with impurity of Fe3+. It will also liberate I2 from KI along with
copper. Hence the analysis of sample would yield a relatively high value of the percentage
of copper.
b) Indeterminate Errors:
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Intermediate or random errors arise from uncertainties which are associated with very
physical or chemical measurements. Such an error cannot be attributed to any known
cause. They are led to both high and low results with equal probability. these cannot be
eliminated or corrected and are the ultimate limitation on the measurement.
Minimisation of Errors
1. Calibration of apparatus and application of corrections:
Need to be conduct periodic calibration of the instrument and apparatus.
Ex: weights (fractional and complete) flasks, pipettes and burettes.
2. Analysis of standard samples: method can be checked by carrying out the
analysis of standard sample prepared in such a way that its composition is exactly
the same as that of the material to be analysed.
Standards constituents available from National Bureau of standards.
3. Running a blank determination:
It helps us to make correction for the excess reagent(titrant) added in titration to
bring about a change in the colour of indicator at the end point.
4. Independent analysis:
Accuracy of a determination may be established by carrying out the analysis in
different manner.
Ex: IRON, HCl, AgNO3
Gravimetrically: Fe3+
Volumetrically: Fe2+.
Significant Figures
The numbers which express the results of a measurement such that only the last digit is in
doubt are called significant figures OR Number of digits in a figure that express the
precision of a measurement.
Significant Figure -Rules
1) ALL non-zero numbers (1, 2, 3, 4, 5, 6, 7, 8, 9) are ALWAYS significant.
2) ALL zeroes between non-zero numbers are ALWAYS significant.
3) ALL zeroes which are SIMULTANEOUSLY to the right of the decimal point
AND at the end of the number are ALWAYS significant.
4) ALL zeroes which are to the left of a written decimal point and are in a number >= 10 are
ALWAYS significant.

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Computation rules:
Rule 1:
In expressing an experiment measurement, never retain more than one doubtful digit.
Eliminate all digits that are not significant.
Examples:
76.8437 i.e.,76.8
All the numbers following the last figure to be retained is less than 5, the last digit
retained is not changed.
59.873 i.e., 59.9
All the numbers following the last figure to be retained is greater than 5, the last digit
retained is increased.
69.752 i.e.,69.8
if the figure following the last digit to be retained is 5 and at least one of the other digits is
non-zero, the last digit retained is increases by one.
68.20 i.e., 68.2 and 64.35 i.e., 64.4
if the figure following the last digit to be retained is 5 and all the other digits is to be
removed are zero, the last digit retained not changed if it is even but is increased by 1 if is
odd.
2.Addition and Subtraction:
In addition, or subtraction, there should be in each number only as many significant
figures as there are in the least accurately known number.

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Example:168.11 + 7.045 + 0.6832 should be written

168.11
7.05
0.68
175.84

Example: +15.23+9.145 – 4.6750 +121.4 =141.09

15.23
9.145
121.4
145.775
-4.675
141.1

3.Multiplication and division:

The numbers of significant figures in the results is the same as in the least reliable
measurement.
Example:
0.165m3 x 10.487 kg m-3=1.730355 Kg = 1.73 kg.
4. Logarithms and Significant figures:
Logarithm made up of two parts
1. A whole number – Characteristic 2. Decimal fraction – mantissa
Accuracy (How close a particular measure is to the true or correct value)
It expresses the closeness of agreement between the value which is accepted either as a
conventional true value or an accepted reference value and the value found.
OR
Agreement between the data and true value or it refers to the closeness of a single
measurement to its true value.
Possible ways of determination of accuracy
1.Absolute method i.e., difference between experimental value and true value.
𝐴𝑏𝑠𝑜𝑙𝑢𝑡𝑒 𝑒𝑟𝑟𝑜𝑟 = 𝐸𝑎𝑏𝑠 = 𝑥 − 𝑇
Absolute error can have either a positive or negative value.
2.Comparative method
Example:
20.54 – 20.44 =0.10%

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20.34-20.44 = -0.10%
Negative sign indicates that experimental value is smaller than true value.
Precision (a measure of how reproducible or how close replicate measurements become):
The precision of an analytical procedure expresses the closeness of agreement (degree of
scatter) between a series of measurements obtained from multiple sampling of the same
homogeneous sample under the prescribed conditions.
Precision may be considered at three levels: repeatability, intermediate precision and
reproducibility.
Precision should be investigated using homogeneous, authentic samples. However, if it is not
possible to obtain a homogeneous sample it may be investigated using artificially prepared
samples or a sample solution. The precision of an analytical procedure is usually expressed as
the variance, standard deviation or coefficient of variation of a series of measurements.
Standard deviation: Measures the spread of the experimental values and gives a good
indication of how close the values are to each other.

√Σ(𝑥𝑖 − 𝜇)2
𝜎=
n
𝜎 = 𝑠𝑡𝑎𝑛𝑑𝑎𝑟𝑑 𝑑𝑒𝑣𝑖𝑎𝑡𝑖𝑜𝑛
𝑥𝑖 = 𝑖𝑛𝑑𝑖𝑣𝑖𝑑𝑢𝑎𝑙 𝑠𝑎𝑚𝑝𝑙𝑒 𝑣𝑎𝑙𝑢𝑒𝑠
𝜇 = 𝑡𝑟𝑢𝑒 𝑚𝑒𝑎𝑛
n = total population of samples
For calculating the standard deviation of a group of assays.

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Range:
Difference between the largest and smallest observation.
PHARMACOPOEIA
PHARMACOPOEIA / FORMULARIES / COMPENDIA
The books containing the standards for drugs and other related substances are known as
pharmacopoeia and formularies - collectively these books are known as the drug compendia.
The pharmacopoeias or formularies contain a list of drugs and other related substances
regarding their source, descriptions, standards, tests, formulae for preparing the same,
action and uses, doses, storage conditions etc. These books are prepared under the authority
of the Government of the respective countries. The word “pharmacopoeia” is derived from
the Greek words ‘pharmacon’ meaning ‘drug’ and ‘poieo’ means ‘make’. Literally it means
that it is a list of medicinal substances, crude drugs and formulae for making preparations
from them. These books are revised from time to time so as to introduce the latest
information available as early as possible after they become established. In order to keep the
size of book within reasonable limit it becomes necessary to omit certain less frequently used
drugs and pharmaceutical adjuvants from each new edition of the book. Therefore, in each
new edition of these books certain new monographs are added while the older ones are
deleted. For the preparation of these books the expert opinion of medical practitioners,
teachers and pharmaceutical manufacturers are obtained.
CLASSIFICATION
The drug-compendia are classified as:
(i) Official compendia
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(ii) Non-official compendia

OFFICIAL COMPENDIA
Official compendia are the compilations of drugs and other related substances which are
recognized as legal standards of purity, quality and strength by a government agency of
respective countries of their origin.
Ex:
British Pharmacopoeia (BP)
British Pharmaceutical Codex (BPC)
Indian Pharmacopoeia (IP)
United States Pharmacopoeia (USP)
National Formulary (NF)
The State Pharmacopoeia of USSR and
Pharmacopoeias of other countries
NON-OFFICIAL COMPENDIA
The book other than official drug compendia which are used as secondary reference sources
for drugs and other related substances are known as non-official drug compendia.
EX:
Merck Index
Extra Pharmacopoeia (Martindale)
United States Dispensatory etc.
Indian Pharmacopoeia
History
The historical developments of Pharmacopoeia in India traces back to 1563 and the credit
goes to Garcia da Orta a Portuguese physician-cum-teacher.
The idea of indigenous Indian Pharmacopoeia was conceived in 1837 which bore fruits in
1841 in the shape of Bengal Pharmacopoeia and Conspectus of Drugs.
The Hindustani version in Bengali and Hindi of London Pharmacopoeia was made available
in India from 1901 onwards.
The Indian Pharmacopeial List, published in 1946 formed the seeding for the true Official
Indian Pharmacopoeia published in 1955.
The first edition of Indian Pharmacopoeia was published in 1955, but actually the process
was started as early as 1944. In 1944 Government of India asked the Drugs Technical
Advisory Board to prepare the list of drugs used, in India, having sufficient medicinal value
to justify their inclusion in official pharmacopoeia.
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The Indian Pharmacopeial List, 1946.

The list of drugs both included and not included in the British Pharmacopoeia along with
standards to secure their usefulness, tests for identity and purity was prepared by the
committee and was published by the Government of India under the name ‘The Indian
Pharmacopoeial List 1946’.
The committee constituted under the chairmanship of Col. Sir R.N.Chopra along with other
nine members, prepared the list of drugs with the following details:
Substances included in the British Pharmacopoeia for crude drugs, chemicals and their
preparations.
Substances not included in the British pharmacopoeia
a) Drugs of plant origin
b) Drugs of animal origin
c) biological products
d) Insecticides
e) Colouring agents
f) Synthetics
g) Miscellaneous
h) Drugs for veterinary use.
The Indian Pharmacopoeial List 1946 was prepared by Department of Health, Govt. of India
in 1946.
The history of development of Indian Pharmacopoeia:

Year Event
1946 The Govt. of India published the Indian Pharmacopeial L
1948 The Govt. of India constituted a permanent Indian Pharmacopoeia Committee. This
committee was assigned the task of preparing Indian Pharmacopoeia and to keep it
up-to-date.
1955 The first edition of Indian Pharmacopoeia (IP) was published.
1960* Supplement of IP 1955 was published
N.B. The work of revision of the Indian Pharmacopoeia as well as compilation of
new edition was taken up simultaneously under the chairmanship of Dr. B.N.Ghosh,
who died in 1958. After Dr. B.N.Ghosh, Dr. B.Mukherjee, the Director of Central
Drug Research Institute was appointed as the chairman of Indian Pharmacopoeia

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committee.
1966* The second edition of IP was published.
1975 A supplement of IP 1966 was published.
1978 The Indian Pharmacopoeia Committee was reconstituted by the Govt. of India,
Ministry of Health and Family Welfare, under the chairmanship of Dr. Nitya Nand,
Director, Central Drug Research Institute, Lucknow
1985 The third edition of IP was published in two volumes, Volume-I and Volume II by
the Controller of Publications, on behalf of Govt. of India, Ministry of Health and
Family Welfare.
Volume-I contains
Legal Notices, Preface, Acknowledgments, Introduction, General Notices, and
Monographs from A to P.
Volume-II contains:
Monographs from Q to Z, Appendices, Contents of Appendices and Index.
1989 Addendum (I) to IP 1985 was published.
1991 Addendum (II) to IP 1985 was published
1996* The fourth edition of IP was published

For the preparation of Pharmacopoeia of India, the pharmacopoeias of other countries, like
British, Europe, United States, USSR, Japan, the National Formulary (USA) and Merck
Index were consulted. The persons working in pharmaceutical industry, drug control
laboratories, research and teaching institutions also actively participated. Under the Drugs
and Cosmetics Act 1940, the Indian Pharmacopoeia is an official book which contains the
standards for drugs and other related substances included in the pharmacopoeia. The drugs
and other related substances prepared by pharmaceutical manufacturers must comply with
these standards.
VARIOUS OFFICIAL PUBLICATIONS RELATED TO PHARMACY PROFESSION
IN INDIA
1. NATIONAL FORMULARY OF INDIA
For the guidance of medical practitioners, medical students and pharmacists in
hospitals and in sales departments National Formulary of India has been formulated.
1960 First edition was published by Govt. of India, Ministry of Health.
1966 Second edition was published.
1979 Third edition was published.
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It contains information about drug interaction, resistance, cumulative effects, drug

dependence, prescription writing etc.
2. THE INDIAN PHARMACOPOEIA
Under the Drugs and Cosmetics Act 1940, the Indian Pharmacopoeia is an official
book which contains the standards for drugs and other related substances included in
the Pharmacopoeia. The drugs and other related substances prepared by
pharmaceutical manufacturers must comply with these standards.
1946 Indian Pharmacopoeial List was published by Govt. of India.
1955 First edition of Indian Pharmacopoeia was published.
1960 Supplement of IP 1955 was published.
1966 Second edition of IP was published.
1975 Supplement of IP 1966 was published.
1985 Third edition of IP was published.
1989 Addendum-I to IP 1985 was published.
1991 Addendum-II to IP 1985 was published.
1996 Fourth edition of IP was published.
2000 addendum, supplement 2000 for Veterinary Products, addendum 2002 and
addendum 2005;
2007 Fifth edition, followed by addendum 2008;
2010 - Six edition with DVD followed by its addendum 2012;
2014 – Seventh edition with DVD followed by its addendum 2015 and addendum
2016
2018 with DVD - Eighth edition
2022 - Ninth edition
Under each monograph chemical structures, molecular weight, physical description,
solubility, identification tests, standards, assay method, storage etc. are given. Indian
Pharmacopoeia is published by the Controller of Publications, Delhi on behalf of
Govt. of India, Ministry of Health and Family Welfare.
3. THE BRITISH PHARMACOPOEIA (BP)
Under the Medical Act 1858 the General Council of Medical Education and
Registration was empowered to alter, amend and republish the British
Pharmacopoeia (BP) as often as necessary. The first BP was published in 1964.
1864 The first BP was published.

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1926 Committee of Civil Research recommended that a Pharmacopoeia Commission

be formed and it should be entrusted the work of new editions of BP and also
recommended that BP be revised and reissued at an interval of ten years.
1932 New edition of BP was published according to the above recommendation.
1968 Medicines Act 1968 gave the responsibility of preparing the BP to the
Medicines Commission. Medicines Commission reconstituted the British
Pharmacopoeia Commission and gave the responsibility to British Pharmacopoeia
Committee.
1980 The thirteenth edition of BP was published.
1988 The 14th edition of BP was published.
1993 The 15th edition of BP was published.
BP 1988 contains two volumes with 2100 monographs:
Vol-I contains monographs on medicinal and pharmaceutical substances along with
Infrared (IR) reference spectra.
Vol-II contains formulated preparations, blood products, immunological products,
radiopharmaceutical preparations, surgical materials and appendices.
BP is the source of standards of drugs in United Kingdom and other parts of
Common Wealth Countries.
4. BRITISH PHARMACEUTICAL CODEX (BPC)
It was in 1903 that the council of Pharmaceutical Society of Great Britain decided
to prepare a reference book for the use of medical practitioners and dispensing
pharmacists. The first edition of BPC was published in 1907.
On the request of British Pharmacopoeia Commission, the Council of the
Pharmaceutical Society agreed in 1959 for the publication of Codex to coincide with
that of the BP, so that BP and BPC should come into effect on the same date.
The BPC differs from BP in that:
a) It contains many more drugs and preparations some may be included in advance to
the pharmacopoeia while other drugs may have been included in the former editions
of pharmacopoeia but now, they are retained in the Codex because they are still
commonly used.
b) It provides information on the actions and uses of drugs, their undesirable effects,
precautions and the treatment of poisoning.
c) It contains formulae, method of preparation, container and storage conditions of
most of the preparations which are still extemporaneously prepared in the pharmacy.
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5. THE UNITED STATES PHARMACOPOEIA (USP)

The USP was originally published in 1820 under the authority of United States
Pharmacopeial Convention. The National Formulary (NF) was published in 1888
under the guidance of American Pharmaceutical Association. In 1974 the NF was
purchased by the United States Pharmacopeial Convention and from 1980 onwards
only one official book of drug standards was published under the heading The United
States Pharmacopoeia and The National Formulary (USP-NF).
6. EXTRA PHARMACOPOEIA
The Extra Pharmacopoeia was first produced in 1883 by William Martindale and is
still known as ‘Martindale’. This is an authorized reference book on drugs and is
used throughout the world. It provides all sorts of latest information on drugs and
medicines. It is published by the direction of the Council of the Royal
Pharmaceutical Society of Great Britain and prepared in the Society’s Department of
Pharmaceutical Sciences.
7. THE MERCK INDEX
It is an encyclopaedia of chemicals, drugs and biologicals. The first edition was
published in 1989 and the eleventh edition was published in 1989 by Merck & Co.,
Inc. Rahway, New Jersy, USA.
8. THE INTERNATIONAL PHARMACOPOEIA
The International Pharmacopoeia is published by the World Health Organization and
is particularly used in developing countries. The first edition was published in 1951
(Volume - I) and in 1955 (Volume-II).
The object of this was to provide a uniform list which would avoid the confusion
caused by different national standards, strengths and names especially for the use of
travellers who might need to use the same prescription in different countries.
IMPURITIES
The impurities in drug are unwanted chemicals that remain with the Active pharmaceutical
Ingredients (API), developed during formulation and upon ageing of API/drug products. The
presence of these unwanted chemicals even in trace amount may influence the efficacy and
safety of pharmaceutical products.
The control of pharmaceutical impurities is currently a critical issue to the pharmaceutical
industry. The International Conference on Harmonization (ICH) has formulated a workable
guideline regarding the control of impurities.
Classification of Impurities
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A. According to ICH guidelines, impurities associated with API’s are classified into:
1. Organic Impurities
2. Inorganic impurities
3. Residual solvents
B. United States Pharmacopoeia (USP):
1. Impurities in Official Articles
2. Ordinary Impurities
3. Organic volatile Impurities
Organic Impurities
Organic impurities arise during the manufacturing process or storage of drug substance i.e.,
starting material, byproducts, intermediates, degradation products, reagents, catalysts and
enantiomeric impurities.
Arise during synthesis, purification and storage of drug substances.
a) By product impurities
In synthetic chemistry getting a single and product with 100% yield is very rare as there is
always a chance of some by-product with desired end product.

b) Degradation Products
Degradation products resulting from storage of formulation to different dosage forms
or aging are common impurities in the medicines.
For e.g. Degradation of penicillin and cephalosporin, Penicillin reacts with moisture
to form Penillic acid, Penicillo aldehyde and Penicillamine etc.
c) Intermediate Products
These products from within the reaction and sometimes those intermediates do not get
converted into the product and remains as such as the impurity with the end product.

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d) Reagents, ligands and catalysts:

These are the chemicals used to carry out various reactions, these impurities are less
commonly found However, in some cases they may produce a problem as impurities.
e) Enantiomeric Impurities:
Stereoisomers (enantiomers and Dia stereoisomers) are related products similar to
drug substance, which may also act as impurity.
For e.g. (R)- and (S)- naproxen are enantiomers. (S)-naproxen is used to treat arthritis
pain, but (R)-naproxen causes liver poisoning.

INORGANIC IMPURITIES
Inorganic impurities involve reagents, ligands, catalysts, heavy metals, or other
residual metals, inorganic salts, filter-aids, charcoal etc.
a) Reagents, ligands and catalysts:

The precipitation of calcium carbonate is washed to remove excess of Na 2CO3 and CaCl2.
If the precipitate is not properly washed, it may remain as an impurity.
b) Heavy metals:
The main source of heavy metals is the water used in the processes and the reactors (if
stainless steel reactors are used), where acidification and acid hydrolysis take place. May
avoided by using demineralized water and glass-lined reactors.
c) Other materials (e.g., filter aids, charcoal):
Activated Carbon, Filters and filtering aids such as centrifuge bags are used in drug
manufacturing, fibres and black particles in bulk drug manufacturing is essential to avoid.
Residual Solvents

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Solvents are organic volatile chemicals used during the manufacturing process or
generated during the production.
Residual solvents are classified into three classes
Class I Solvents: Benzene (1-2 ppm), carbon tetrachloride (1-4 ppm) is avoided because
of their carcinogenic and toxic effect
Class II Solvents: methyl chloride, methanol, pyridine, toluene, acetonitrile, Most
commonly used
Class III Solvents: acetic acid, acetone, isopropyl alcohol, butanol, ethanol, ethyl acetate
permitted daily exposure 50 mg or less per day.
Other Impurities
a) Excipient impurities:
Excipients such as peroxide, aldehydes, heavy metals, alkaline residue may render as
an impurity in final product.
b) Elemental impurities:
Brought about as excipient impurities or during manufacturing as catalyst for
oxidation and hydrolysis. Ex: As, Al, Ca, Na, Pb.
c) Leachable and extractable substances from primary packaging material form reaction
products (impurities) with drug substances. Ex: Alkaline oxide from glass (Na2O,
SiO2,MgO, Ca O).
EFFECTS OF IMPURITIES
 Impurities which are toxic can be injurious when present above certain limit.
 Impurities can cause incompatibility with other substances. Impurities may cause
a change in physical and chemical properties of substances.
 Impurities present in large proportion that the active strength of the substance get
lower, its therapeutic effect gets decrease.
 Impurities though harmless in nature may cause change in odour, colour, taste etc.
Impurities may decrease the shelf life of the product.
SOURCES OF IMPURITIES
 A compound is said to be impure if it is having foreign matter i.e., impurities. The
substance employed in the pharmaceutical field must be almost pure so that they
can be used safely.
 A list of the possible impurities can be readily complied from knowledge of the
raw material used for method of manufacture and the stability of the product. The

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type and number of impurities present in the chemical or pharmaceutical

substance depends upon several factors.
 There are several sources of impurities in pharmaceutical substance:

1.Raw materials employed in manufacturing:

If impurities are present in the raw material (ores, metals etc.) which are used in the
preparation of pharmaceutical chemicals then these may be carried through during the
manufacturing process to the final product. Thus, final compound may be
contaminated with these impurities.
Examples:
a) Zinc sulphate is prepared by either zinc oxide or zinc metal with sulphuric acid.

Both zinc and zinc oxide contain aluminium (Al), copper (Cu), magnesium (Mg),
manganese (Mn), nickel (Ni), arsenic (As) and iron (Fe) as impurities.
b) Copper sulphate may be prepared by the action of sulphuric acid on copper
turnings.

Copper turnings may contain iron and arsenic as impurities.

2. Reagents used in manufacturing process:
If reagents used in the manufacturing process are not completely removed by
washing, these may find entry into the final products. Examples:
a) Ammoniated mercury may be prepared by adding a solution of mercuric chloride
to dilute ammonia solution.

The NH2HgCl (ammoniated mercury precipitate) completely washed to remove

ammonium hydroxide. If it is not removed completely by washing with water, the
final product may contain ammonium hydroxide as impurity.

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b) When calcium chloride is react with sodium carbonate precipitated calcium

carbonate is formed.

The precipitate of calcium carbonate is washed to excess of sodium carbonate and

calcium chloride.
3. Intermediate products in the manufacturing process:
There are some intermediates which are produced during the manufacturing process.
Sometimes, these intermediates may be carried by the final product.
Examples:
6KOH + 3I2 → 5KI + KIO3 +3H2O
The resulting solution is first evaporated to dryness and heated with charcoal.
KIO3 + 3C → KI + 3CO
Potassium iodide so formed is an intermediate product and if it not converted into
potassium iodide, it may be present as impurity in the final product.
4. Defects in manufacturing process:
Defects such as imperfect mixing, incompleteness of reaction, non-adherence
to proper temperature, pressure, pH or reaction condition etc. may results in the
production of chemical compounds with impurities in them.
Examples:
Method of manufacturing zinc oxide involves heating metallic zinc to bright
redness in a current of air. The vapours of the metal burn to form zinc oxide which is
collected as a fine white powder.
2ZN + O2 → 2ZNO
If zinc metal is not completely converted into zinc oxide (due to lesser heat or air) a
small amount of zinc metal may still remain as impurity in final product.
5. Solvents
Water is a cheapest solvent and is commonly used especially in the manufacturing of
inorganic chemicals.
Tap water contains chloride, sulphate, bicarbonate, magnesium, and calcium etc as
impurities though in very small amounts. Hence traces of such impurities may still

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remain in final products, to remove these impurities from water either distilled water
or deionized water should be used.
6. Action of solvent and reagent on reacting vessel:
Some reagent and solvent may react with the container in which they are stored or
processed.
Example:
Strong acid leads out alkali from borosilicate glass, copper and zinc vessels react with
slightly acidic substance etc. hence a great care must be taken in selection of suitable
vessels for manufacturing process or for the purpose of storage.
7. Atmospheric contamination:
In industrial areas, atmosphere is contaminated with dust particles (aluminium oxide,
silica glass particles, porcelain particles, plastic fragments, etc.) and some gases like
hydrogen sulphide, sulphur dioxide and black smoke. During the manufacture or
purification of pharmaceutical products, these impurities enter the final products as
impurities.
Example:
Sodium hydroxide absorbs atmospheric carbon dioxide (contamination) to form
sodium carbonate and bicarbonate
2NaOH + CO2 → Na2CO3 + H2O
Therefore, NaOH should not be exposed to atmosphere for a long time during its
manufacturing.
8. Defective storage of final products:
Some pharmaceutical chemicals undergo chemical decomposition if these are not
properly stored, may also change their physicochemical properties.
Example:
When ferrous sulphate is exposed to moist air ferric sulphate is formed. Potassium
iodide gets liquefied if it is exposed to moist air for a long time.
Limit Tests
They are designed to detect and limit / control small quantities of impurities present in
the substance. Comparison of standard turbidity/ colour with that of the sample under
test.
Limit Test for Chlorides
Principle: It is based upon the chemical reaction between silver nitrate and
soluble chlorides in presence of dilute nitric acid to give opalescence of silver
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chloride. If the Opalescence produced is compared with the standard solution.

If the opalescence in the sample is less than the standard, it passes the test. If it
is more than the standard, it fails the test.

Procedure:
Take two 50 ml Nessler Cylinders. Label one as “Test” and the other as ‘Standard’.
Test Standard
Dissolve the specified quantity of the Place 1ml of 0.05845% w/v solution of
substance in distilled water and transfer NaCl and transfer to Nessler cylinder.
Add 10 ml of dil. Add 10ml of dil. Add 10ml of dil. HNO3. Dilute to 50ml
HNO3. Dilute to 50ml with water and add with water and add 1 ml of silver nitrate
1 ml of silver nitrate solution. Stir solution. Stir immediately with a glass
immediately with glass rod and allow to rod and allow to stand for 5 minutes.
stand for 5 minutes.

Limit Test for Sulphates

Principle:
It is based upon the chemical reaction between Barium chloride and soluble
sulphate in presence of dilute Hydrochloric acid. The turbidity produced is
compared with the standard solution. Barium sulphate reagent contains barium
chloride, sulphate – free alcohol and small quantity of potassium sulphate. The
inclusion of the small quantity of potassium sulphate in the reagent increase the
sensitivity of the test. Alcohol prevents super saturation and more uniform turbidity
develops. If the turbidity produced in the test is more intense than the standard
turbidity it fails the test. otherwise, it passes the test.

Procedure: Take two 50 ml Nessler Cylinders. Label one as “Test” and the other as
‘Standard’
Test Standard
Dissolve the specified quantity of the Place 1ml of 0.1089% w/v solution

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substance in distilled water and transfer to Potassium sulphate and 2 ml of dil HCl in
Nessler cylinder. Add 2ml of dil. HCl. Nessler cylinder. Add 10ml of dil. HNO3.
Dilute to 45ml with water and add 5 ml of Dilute to 45ml with water and add 5 ml
barium chloride reagent. Stir immediately of barium sulphate reagent. Stir
with glass rod and allow to stand for 5 immediately with a glass rod and allow to
minutes. stand for 5 minutes.

Limit Test for IRON

Principle:
The test depends upon the reaction between ferrous iron and
thioglycollic acid in the presence of ammonia and citric acid When a pale
pink to deep reddish-purple colour is produced. Ferric iron is reduced to
ferrous iron by the thioglycolic acid and the compound produced is ferrous
thioglycolate. Citric acid forms a soluble complex with iron and prevents its
precipitation by ammonia as ferrous hydroxide. Ferrous thioglycollate is
colourless in neutral or acid solutions. The colour develops only in the presence of
alkali. It is stable in the absence of air but fades when exposed to air due to oxidation
to the ferric compound. Therefore, the colours should be compared immediately after
the time allowed for full development of colours is over.

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Procedure: Take two 50ml Nessler cylinders. Label one as ‘Test’ and the others as
‘standard’.
Test Standard
Dissolve the specified quantity of the Dilute 2ml of standard iron solution
substance in 20ml distilled water and with 20 ml of water in Nessler cylinder.
transfer to Nessler cylinder. Add 2ml of Add 2ml of a 20%W/V solution of iron -
20% W/V solution of iron -free citric acid ferric acid and 0.1ml of thioglycolic acid
and 0.1ml of thioglycolic acid and mix. and mix. Make alkaline with iron free
Make alkaline with iron -free ammonia ammonia solution. Dilute to 50ml with
solution. Dilute to50ml with water and water and allow to stand for 5 minutes.
allow to stand for 5 minutes.
Limit Test for Arsenic

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Arsenic is harmful due to its toxic nature Pharmacopoeia method is based on ‘Gutzeit
Method’. Concentration of arsenic beyond 0.01 mg/L in pollutant by the World
Health Organization (WHO).
Principle:
Limit Test for Arsenic (As) is based on the fact that Arsenic is easily reduced into
Arsine gas (AsH3), which on mercuric chloride paper gives yellow stain.
Arsenic (As) may be present as As3+(Trivalent) or As5+ (Pentavalent).

The solution is treated with a reducing agent (stannous Chloride) to convert the
pentavalent arsenic acid into trivalent arsenious acid.

The arsenious acid is then converted into gaseous arsenious hydride (arsine gas) with
the help of nascent hydrogen, which is produced by Zn + HCl.

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Arsine gas is carried through the tube by the stream of hydrogen and out through the
mercuric chloride paper.

This results in the formation of yellow or brown stain on the mercuric chloride paper.
The intensity of the colour is proportional to the quantity of arsenic.

Procedure:
Test Standard
1.Dissolve 10g sodium chloride 1.Transfer 1.0ml of arsenic standard
in 50 ml of distilled water solution and dilute it to 50ml with
distilled water.
2.Add 12 ml of stagnated
hydrochloric acid AsT 2.Add 10 ml of stannated hydrochloric
acid AsT.
3.Add 5 ml of 1 M of potassium 3.Add 5 ml of 1 M of potassium iodide
iodide and 10g of zinc AsT and 10g of zinc AsT.
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Limit Test for Heavy Metals

Principle:
It is based on the reaction between the solution Heavy metals and a saturated solution
of Hydrogen sulphides. In acidic media, it produces reddish / black colour with
Hydrogen sulphide which is compared with standard lead nitrate solution.
Pb + H2S →Pbs + H2
Procedure:
Take two 50 ml Nessler Cylinders. Label one as “Test” and the other as ‘Standard’.
Test Standard
Place 25ml of the solution and adjust pH 2ml of standard lead solution is taken in
3-4 by using dilute acetic acid or dilute Nessler cylinder. And dilute to 25ml with
ammonia. Dilute to 35ml with water and water. Adjust the pH 3-4 by using dilute
mix. Add 10ml of freshly prepared by ammonia. Dilute to 35ml with water and
H2S solution. Dilute to50ml with water mix. Add 10 ml of freshly H2S solution.
and allow to stand for 5 minutes. Dilute to 50ml with water. Mix and allow
to stand for 5 minutes.

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UNIT-II
ACID-BASE THEORY
Reason: To control acids in oils.
PROPERTIES
PROPERTY ACID BASE
Color Colourless Colourless
Taste Sour Bitter
Touch Tingling Sensation or A Sharp, Bitting Almost Slippery
Painful Sensation When in Excess
Solubility Water Water
PH < 7 and turn blue litmus red >7
Example Lemons, oranges, Vinegar (acetic acid), sulfuric Soap, toothpaste,
acid (car batteries), tartaric acid (baking) and cleaning agents, lime
Urine water

Classification of Acids and Bases

ACID -BASE THEORIES

Arrhenius Theory Bronsted Theory Lowis Theory
Acid- Produces H+ Acid- Donate a Proton Acid- accept a pair
of electrons
Base-Produces OH- Base- accept a Proton Base- donate a pair
Dissociation Proton transfer of electrons
Electron pair
transfer

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KEY POINTS OF THEORIES

Conjugate Acid: acid becomes the 1. Complex ion/ionic
Limitations
conjugate base after it donates the proton. compound
Conjugate Base: The base becomes the
Drawbacks 2. Amphoterism
conjugate acid after it accepts the proton

Arrhenius Theory:
Svante Arrhenius attributed the properties of acidity to hydrogen ions (H+) in 1884.
Acids which produce hydrogen ions (H+)
Bases are substances that produce hydroxide ions (OH-) in solution.
H+ Ions form the hydronium ion (H3O+) when they combine with water molecules.
Example: Acid
HCl(aq)+H2O(l)→H3O+(aq)+Cl−(aq)
Example: Gas
HBr(g)+H2O(l)→H3O+(aq)+Br−(aq)
The Complete dissociation of H br gas into water results generates Free H3O+ Ions.
Example: Base
NaOH(aq)→Na+(aq)+OH−(aq)
Creation of Hydronium ion:

High charge density of the proton strongly attracts it to any part of a nearby atom or molecule
in which there is an excess of negative charge.

Example: Water (H2o -amphiprotic)

Lone pair of electrons of the oxygen atom, the tiny proton will be occupied within the lone
pair electron and will form a shared-electron (coordinate) bond with it, creating
a hydronium ion, H3O+. In a sense, H2O is acting as a base here, and the product H3O+ is the
conjugate acid of water.

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Limitations
1. The Arrhenius definitions of acidity and alkalinity are restricted to aqueous solutions and
refer to the concentration of the solvated ions.
2. Under this definition, pure H2SO4 or HCl dissolved in toluene is not acidic, even though
both of these acids will donate a proton to toluene. In addition, under the Arrhenius
definition, a solution of sodium amide (NaNH2) in liquid ammonia is not alkaline, even
though the amide ion (NH2) will readily deprotonate ammonia. Thus, the Arrhenius
definition can only describe acids and bases in an aqueous environment. Some acids do
not contain H+ ions and some bases do not contain OH ions. But behave as acids and
bases respectively.
3. Possible only in aqueous solution. However, acid-base reactions can also occur in non-
aqueous solution—NH4NO3 in Liq. NH3 acts as acid though it does not give H+ ions. NH3 is
a base though it does not give OH ions.
4. It cannot explain why AlCl3 is acidic.
5. It cannot explain the neutralization reaction in the absence of water.
The theory suggests that for a substance to release either H+ or OH- ions, it must contain that
particular ion. However, this does not explain the weak base ammonia (NH3) which
releases hydroxide ions into solution in the presence of water but does not contain OH- itself.
Similar reactions occur.
NaOH(aq)+HCl(aq)→NaCl(aq)+H2O(l)
NH3(aq)+HCl(aq)→NH4Cl(aq) …For Gases – Arrhenius Explanation Failure.
In the sodium hydroxide case, hydrogen ions from the acid react with hydroxide ions from
the sodium hydroxide. However, in the ammonia case, there are no hydroxide ions.
In this case, there are not any hydrogen ions or hydroxide ions in solution - because there isn't
any solution.
NH3(aq)+H2O(l)⇌NH+4(aq)+OH−
When the ammonia reacts with the water, it is dissolved in to produce ammonium ions and
hydroxide ions.
Bronsted Lowry Theory
In 1923, chemists Johannes Nicolaus Brønsted and Thomas Martin Lowry independently
developed definitions of acids and bases based on the compounds' abilities to either donate
or accept protons (H+ ions).
Acids Bases

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Proton donors Proton acceptors

The compounds act as both acid and base together is called Amphoteric.
HCl(aq)+NH3(aq) →NH+4 (aq)+Cl−(aq)
The hydrochloric acid (HCl) "donates" a proton (H+) to ammonia (NH3) which
"accepts" it, forming a positively charged ammonium ion (NH4+) and a negatively charged
chloride ion (Cl-). Therefore, HCl is a Brønsted-Lowry acid (donates a proton) while the
ammonia is a Brønsted-Lowry base (accepts a proton). Also, Cl- is called the conjugate
base of the acid HCl and NH4+ is called the conjugate acid of the base NH3.
According to theory
An acid is a substance that can release a proton (like in the Arrhenius theory) and a base is
a substance that can accept a proton. A basic salt, such as Na+ F-, generates OH- ions in
water by taking protons from water itself (to make HF).
F−(aq)+H2O(l)⇌HF (aq)+OH−
When a Brønsted acid dissociates, it increases the concentration of hydrogen ions in the
solution, [H+]; conversely, Brønsted bases dissociate by taking a proton from the solvent
(water) to generate [OH−].
Ionisation:
The process in which an electron is given enough energy to break away from an atom is
called ionisation. This process results in the formation of two charged particles or ions: the
molecule with a net positive charge, and the free electron with a negative charge
Acid dissociation:
HA (aq)⇌A−(aq)+H+(aq)
Acid Ionization Constant
Ka=[A−][H+]/[HA]
Base dissociation:
B(aq)+H2O(l)⇌HB+(aq)+OH−
Base Ionization Constant
Kb =[HB+] [OH−]/[B]
STRONG ACIDS: HCl, HNO3, H2SO4, HBr, HI, HClO4
WEAK ACIDS: All other acids, such as HCN, HF, H2S, HCOOH
Strong acids such as HCl dissociate to produce spectator ions such as Cl− as conjugate
bases, whereas weak acids produce weak conjugate bases. This is illustrated below for acetic

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acid and its conjugate base, the acetate anion. Acetic acid is a weak acid (Ka = 1.8 x 10-5) and
acetate is a weak base (Kb = Kw/Ka = 5.6 x 10-10).
STRONG BASES: The hydroxides of the Group I and Group II metals such as LiOH,
NaOH, KOH, RbOH, CsOH
WEAK BASES: All other bases, such as NH3, CH3NH2, C5H5N
The strength of a conjugate acid/base varies inversely with the strength or weakness of its
parent acid or base. Any acid or base is technically a conjugate acid or conjugate base also;
these terms are simply used to identify species in solution (i.e acetic acid is the conjugate acid
of the acetate anion, a base, while acetate is the conjugate base of acetic acid, an acid).
pH Scale:
Since acids increase the amount of H+ ions present and bases increase the amount of OH -
ions, under the pH scale, the strength of acidity and basicity can be measured by its
concentration of H+ ions. This scale is shown by the following formula.
pH = -log[H+]
The pH scale is often measured on a 1 to 14 range.

pH:
one measure of acidity & indicates the concentration of free hydrogen ions(H +)
present in solution.
Examples:
PH of

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 1-is very acid (Conc.H4PO4 i.e., metaphosphoric acid)

 7-is neutral (pure water)
 14 - is very basic (Conc.NaOH)
pH(potential of Hydrogen):
That is the power value of the hydrogen ion concentration presented as a positive number.
This explains why an increase in acidity, or hydrogen ion concentration ([H+]), results in a
lower pH value.
i) [H+] = 0.001 M = 1/1000 M = 1/103 M = 10 -3 M = pH 3.00

(ii) [H+] = 0.01 M = 1/100 M = 1/102 M = 10 -2 M = pH 2.00

Lewis Theory
Lewis' theory used electrons instead of proton transfer and specifically stated that an acid is a
species that accepts an electron pair while a base donates an electron pair.
The Lewis theory of acids (electrophile-attract) and bases (nucleophile -attack) states that
acids act as electron pair acceptors and bases act as electron pair doners. This definition
doesn't mention anything about the hydrogen atom at all, unlike the other definitions.

A coordinate covalent bond is just a type of covalent bond in which one reactant gives it
electron pair to another reactant. In this case the Lewis base donates its electrons to the
Lewis acid. When they do react this way, the resulting product is called an addition
compound, or more commonly an adduct.
Lewis Acid: a species that accepts an electron pair (i.e., an electrophile) and will have vacant
orbitals

Lewis Base: a species that donates an electron pair (i.e., a nucleophile) and will have lone-
pair electrons.
LEWIS ACIDS
Lewis’s acids accept an electron pair. Lewis Acids are Electrophilic meaning that they are
electron-attracting. When bonding with a base the acid uses its lowest unoccupied
molecular orbital or LUMO.
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Examples: Cu2+, Fe2+, Fe3+ and BF3, AlF3 i.e., incomplete octet of electron, SiBr4, SiF4 i.e.,
More than 8 valance shell electrons, and CO2, SO2i.e., multiple bonds.
LEWIS BASES
Lewis Bases donates an electron pair. Lewis Bases are Nucleophilic meaning that they
“attack” a positive charge with their lone pair. They utilize the highest occupied molecular
orbital or HOMO.
e.g., OH−, CN−, CH3COO−, NH3, H2O, CO

LUMO: LUMO stands for lowest unoccupied molecular orbital. These molecular orbitals
can receive electrons from HOMO. As its name implies, these orbitals are unoccupied;
thus, contains no electrons. This is because the energy of these orbitals is very high and
electrons tend to occupy in low energy levels first. Apart from that, these molecular
orbitals are characteristic for electrophilic substances.
HOMO:
The electrons in these molecular orbitals can be donated to the LUMO type molecular
orbitals. This is because these molecular orbitals contain weakly attached electrons.
These molecular orbitals are the most available form for covalent chemical bonding. The
presence of these molecular orbitals is characteristic for nucleophilic substances.
Example:
a reaction between ammonia (NH3) and boron trifluoride (BF3). Since there is no transfer of
hydrogen atoms here, it is clear that this is a Lewis acid-base reaction. In this reaction,
NH3 has a lone pair of electrons and BF3 has an incomplete octet, since boron doesn't have
enough electrons around it to form an octet.

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Because boron only has 6 electrons around it, it can hold 2 more. BF3 can act as a Lewis acid
and accept the pair of electrons from the nitrogen in NH 3, which will then form a bond
between the nitrogen and the boron.

This is considered an acid-base reaction where NH3 (base) is donating the pair of electrons to
BF3. BF3 (acid) is accepting those electrons to form a new compound, H 3NBF3.

Complex Ion / Coordination Compounds

Complex ions are polyatomic ions, which are formed from a central metal ion that has
other smaller ions joined around it. While Brønsted theory can't explain this reaction
Lewis acid-base theory can help. A Lewis Base is often the ligand of a coordination
compound with the metal acting as the Lewis Acid (see Oxidation States of Transition
Metals).

Al3++6H2O⇌ [Al (H2O)6]3+

The aluminium ion is the metal and is a cation with an unfilled valence shell, and it is a Lewis
Acid. Water has lone-pair electrons and is an anion, thus it is a Lewis Base.

Fig. Aluminium ion acts as a Lewis acid and accepts the electrons from water, which is
acting as a Lewis base. This helps explain the resulting Hexa aqua aluminum (III) ion.

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The Lewis Acid accepts the electrons from the Lewis Base which donates the electrons.
Another case where Lewis’s acid-base theory can explain the resulting compound is the
reaction of ammonia with Zn2+.

Zn2++4NH3→ [Zn (NH3)4]4+

Similarly, the Lewis Acid is the zinc Ion and the Lewis Base is NH3. Note how
Brønsted Theory of Acids and Bases will not be able to explain how this reaction occurs
because there are no H+ or OH− ions involved. Thus, Lewis Acid and Base Theory allow us
to explain the formation of other species and complex ions that do not ordinarily contain
hydronium or hydroxide ions. One can expand the definition of an acid and a base via the
Lewis Acid and Base Theory. The lack of H+ or OH− ions in many complex ions can make it
harder to identify which species is an acid and which is a base. Therefore, by defining a
species that donates an electron pair and a species that accepts an electron pair, the definition
of an acid and base is expanded.

Amphoterism

Acids and bases are distinguished as two separate things however some substances can be
both an acid and a base. Noticed this with water, which can act as both an acid or a base. This
ability of water to do this makes it an amphoteric molecule. Water can act as an acid by
donating its proton to the base and thus becoming its conjugate acid, OH-. However, water
can also act as a base by accepting a proton from an acid to become its conjugate base, H 3O+.

Water acting as an Acid:

H2O+NH3→NH+4 +OH−

Water acting as a Base:

H2O+HCl→Cl−+H3O+

You may have noticed that the degree to which a molecule acts depends on the
medium in which the molecule has been placed in. Water does not act as an acid in an acid
medium and does not act as a base in a basic medium. Thus, the medium which a molecule is
placed in has an effect on the properties of that molecule. Other molecules can also act as
either an acid or a base. For example,
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Al (OH)3+3H+→Al3++3H2O

Where Al (OH)3 is acting as a Lewis Base.

Al (OH)3+OH−→Al (OH)−4

Where Al (OH)3 is acting as a Lewis Acid.

Acidic and basic strength and pKa
The pka value of a compound is defined as: pKa =-logKa.
Application(s):
A measure of acidic/ basic strength.
For Acids
The smaller the pKa value i.e., Strong acid, and the Higher the pKa value i.e., Weak Acid
For base
Larger the pKa value i.e., the Stronger base and the weaker base the low the pKa value.

Henderson -Hasselback equation

Derive an equation by considering ionized species proton (H+) and Conjugate base (A-) of a
weak acid (HA),
HA + H2O ↔ H+ + A-(aq)
The ionization constant (Ka) represents the ionised species of weak acid and is written as:
[𝐴−][𝐻+]
𝐾𝑎 = [𝐻𝐴]

Rearranged substituting pH for -log[H+] and pKa for -log Ka to give:

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𝐼𝑜𝑛𝑖𝑧𝑒𝑑 𝑑𝑟𝑢𝑔 𝑐𝑜𝑛𝑐𝑒𝑛𝑡𝑟𝑎𝑡𝑖𝑜𝑛

𝑝𝐻 = 𝑝𝐾𝑎 + 𝑙𝑜𝑔
𝑈𝑛𝑖𝑜𝑛𝑖𝑧𝑒𝑑 𝑑𝑟𝑢𝑔 𝑐𝑜𝑛𝑐𝑒𝑛𝑡𝑟𝑎𝑡𝑖𝑜𝑛
In the case of acid
[𝐴−]
𝑝𝐻 = 𝑝𝐾𝑎 + 𝑙𝑜𝑔
[𝐻𝐴]
10𝑃𝐻−𝑃𝑘𝑎
𝑃𝑒𝑟𝑐𝑒𝑛𝑡𝑎𝑔𝑒 𝑜𝑓 𝑑𝑟𝑢𝑔 𝑖𝑜𝑛𝑖𝑧𝑒𝑑 = 𝑋 100
1 + 10𝑃𝐻−𝑃𝑘𝑎
In the case of base
𝑈𝑛𝑖𝑜𝑛𝑖𝑧𝑒𝑑 𝑑𝑟𝑢𝑔 𝑐𝑜𝑛𝑐𝑒𝑛𝑡𝑟𝑎𝑡𝑖𝑜𝑛
𝑝𝐻 = 𝑝𝐾𝑎 + 𝑙𝑜𝑔
𝐼𝑜𝑛𝑖𝑧𝑒𝑑 𝑑𝑟𝑢𝑔 𝑐𝑜𝑛𝑐𝑒𝑛𝑡𝑟𝑎𝑡𝑖𝑜𝑛
10𝑃𝑘𝑎−𝑃𝐻
𝑃𝑒𝑟𝑐𝑒𝑛𝑡𝑎𝑔𝑒 𝑜𝑓 𝑑𝑟𝑢𝑔 𝑖𝑜𝑛𝑖𝑧𝑒𝑑 = 𝑋 100
1 + 10𝑃𝑘𝑎−𝑃𝐻

Significance:
The pH of a buffer solution can be calculated from the initial concentrations of
the weak acid and the salt provided Ka is given.
However, the Henderson-Hasselbalch equation for a basic buffer will give pOH
and its pH can be calculated as (14 - pOH).
The dissociation constant of a weak acid (or weak base) can be determined by
measuring the pH of a buffer solution containing equimolar concentrations of
the acid (or base) and the salt.
Since, [salt] = [acid], log ([salt]/[acid]) = log 1 =0
pKa = pH
The measured pH, therefore, gives the value of pKa of the weak acid.

Likewise, we can find the pKb of a weak base by determining the pOH of
equimolar basic buffer.
A buffer solution of desired pH can be prepared by adjusting the concentrations
of the salt and the acid added for the buffer.

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It is noteworthy that buffer solutions are most effective when the concentrations
of the weak acid (or weak base) and the salt are about equal. This means that pH
is close to the value of pKa of the acid (or pKb of the base).
Buffers:
A buffer solution is a solution where the pH does not change significantly even on dilution
or even if an acid or base is added at constant temperature. Its pH changes very little
when a small amount of strong acid or base is added to it.
Buffers are prepared from any weak acid or weak base and are used to maintain the pH of a
solution in a narrow range. this is important in living systems:
Types of Buffers
Acid buffer: A buffer solution containing large amounts of a weak acid, and its salt with a
strong base, is termed as an acid buffer. Such buffer solutions have pH on the acidic side i.e.,
pH is less than 7 at 298 K. The pH of an acid buffer is given by the equation. CH3COOH and
CH3COONa.
[𝑆𝑎𝑙𝑡]
𝑝𝐻 = 𝑝𝐾𝑎 + 𝑙𝑜𝑔
[𝐴𝑐𝑖𝑑]

Basic Buffer: A buffer solution containing relatively large amounts of a weak base and its
salt with a strong acid is termed as a basic buffer. Such buffers have pH on the alkaline side
i.e., pH is higher than 7 at 298 K. e.g.: NH4OH and NH4Cl.
[𝑆𝑎𝑙𝑡]
𝑝𝑂𝐻 = 𝑝𝐾𝑏 + 𝑙𝑜𝑔
[𝐴𝑐𝑖𝑑]
Mechanism
acidic buffer solution consisting the acetic acid (CH3COOH) and its salt, sodium acetate
(CH3COONa). Acetate buffers are used in estimation of biochemical enzymes to prevent pH
changes that might affect the biochemical activity of enzymes.
Acetic acid, a weak acid is partially ionized into acetate ions (Ac¯) and proton ions (H+ ) and
undissociated acetic acid in an aqueous solution (aq).
CH3COOH (aq) ↔H+ + CH3COO¯ (aq)
Sodium acetate is a strong conjugate base of acetic acid that is ionized completely into
Sodium ions (Na+) and acetate ions in an aquoues solution (aq). The buffer solution has large
amount of acetate ions.
CH3COO-Na+ (aq)↔ Na+ + CH3COO¯ (aq)

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The ratio of ionized species of acetic acid and sodium acetate is represented by equilibrium
constant or dissociation constant (Ka).
BUFFER-CAPACITY AND BUFFER-RANGE
The effectiveness of any buffer is described in terms of its buffer capacity.
It is defined as, 'the number of equivalents of a strong acid (or a strong base) required to
change the pH of one litre of a buffer solution by one unit, keeping the total amount of the
acid and the salt in the buffer constant.
The buffer capacity of a buffer is maximum when acid to salt or base to salt ratio is equal to
1 i.e., it contains equal number of moles of acid (or base) and the salt.
All buffer solutions remain effective over a small pH range: this pH-range is characteristic of
the buffer and is termed as the buffer-range.
Example: human plasma is buffered at pH 7.4 by carbonic acid/bicarbonate buffer system.
Application(s):
Used in number of areas of analytical chemistry such as
Mobile phase for chromatography and extraction of drugs from aqueous phase.
Many chemical reactions are affected by the acidity of the solution in which they occur. In
order for a particular reaction to occur or to occur at an appropriate rate, the pH of the
reaction medium must be controlled.
Such control is provided by buffer solutions, which are solutions that maintain a particular
pH. Biochemical reactions are especially sensitive to pH.
Most biological molecules contain groups of atoms that may be charged or neutral depending
on pH, and whether these groups are charged or neutral has a significant effect on the
biological activity of the molecule.
In all multi cellular organisms, the fluid within the cell and the fluids surrounding the cells
have a characteristic and nearly constant pH.
This pH is maintained in a number of ways, and one of the most important is through buffer
systems.
Zwitterionic buffers
A zwitterion is a molecule that carry equal number of positively and negatively charged
functional groups and the entire charge of zwitterions becomes zero. Amino acids are
generally good example of zwitterionic molecule as they bear positive charged amine group
and negatively charged carboxyl group. The opposite charged groups cancel each other and
an amino acid become neutral. Hence, they are used for preparation of zwitterionic buffers.
When acids and bases added in a zwitterionic buffer solution, they neutralize protons and
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hydroxyl ions. These buffers fulfil most of the important properties of good buffers which are
widely used in Biochemistry and molecular Biology research.

The important properties of a goods buffer:

1) Solubility: A buffer should be highly soluble in water as most the of cellular reactions
occur in an aqueous environment.
2) Permeability: A buffer should not permeable via plasma membrane.
3) pKa value: A buffer should be highly efficient to maintain pKa range between 6 and 8 as
most of the biochemical experiment have optimum pH range of 6-8.
4) Ionic strength: A buffer should be influenced minimum by ionic composition of medium,
temperature, concentration. They should maintain physiological ionic strength.
5) UV absorption: Buffer should not absorb visible or ultraviolet radiations.
6) Nonreactive: Buffer components should have minimum interaction with the biological
components i.e. DNA, metal ions or enzymes.
7) Non-toxic: Any components of a buffer should not toxic to the test system (cells).
8) Purity: A buffer should be available in high degree of purity or They should free be from
any impurities which may interfere with analysis.
9) Buffer preparation: They should be easily prepared in the laboratory.

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Water Dissociation Constant:

Water is an amphoteric substance. The dissociation of water is an equilibrium reaction. It
means the rate of the forward reaction is equal to the rate of the reverse reaction and the
concentration of the reactants and products do not change at equilibrium. The molar
concentration of H3O+ represented as [H3O+] is equal to 10-7 M in a pure water sample at
25 oC, where M is in moles/Liter. The molar concentration of OH - represented as [OH-] is
equal to the molar concentration of H3O+ in pure water, i.e., [H3O+] = [OH-] = 10-7 M.
The product of the molar concentration of H3O+ and OH- in water is a constant called water
dissociation constant Kw equal to 10-14 at 25 o C, i.e.:

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Kw=[H3O+] [OH−]
= (10−7) (10−7)
=10−14

Calculations of [H3O+] and [OH-] based on Kw

The water dissociation constant remains the same whether the aqueous solution is neutral,
acidic, or basic, i.e.:

Kw=[H3O+] [OH−]
= (10−7) (10−7)
=10−14 at 25ͦ C
Therefore, if the molar concentration of hydronium ions [H3O+] is known, the molar
concentration of hydroxide ions [OH -] can be calculated using the following formula:
𝐾𝑤 10−14
[𝐻3 O +] = =
[𝑂𝐻−] [𝑂𝐻−]

Similarly, if the molar concentration of hydroxide ions [OH -] is known, the molar
concentration of hydronium ions [H3O+] can be calculated using the following formula:

𝐾𝑤 10−14
[OH −] = =
[𝐻3 𝑂+] [𝐻3 𝑂−]
When a strong acid like HCl dissolves in water, it dissociates ~100% into ions. Therefore, the
[H3O+] is equal to the molar concentration of the acid. The amount H3O+ added by
dissociation of water molecules is very small compared to that coming from the dissociation
of a strong acid and can be neglected. Similarly, when a strong base like NaOH dissolves in
water, it dissociates ~100% into ions. Therefore, the [OH-] is equal to the molar concentration
of the base.
10-14
THE LAW OF MASS ACTION
Stated by guldberg and wage in 1867.
The rate of reaction is proportional to the active masses of the reacting substances.
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Ideal state
Active mass represented by the concentration of reacting species i.e., gram molecules or gram
ions per litre. The constant of proportionality is known as velocity constant.
Simple reaction
AB,
The rate of reaction = k [A] is concentration of A and k is the velocity constant.
Reversible reaction
A+ B ↔ C+D
vf = K1 [A]. [B] and vb = K2 [C] [D]
vf = velocity of the forward reaction; vb = velocity of the backward reaction
[A] molar concentration of A and k1 and k2 are constants.
At equilibrium vf = vb
therefore k2 [C] .[D] = K1 [A] .[B]
𝑘1 [𝐶 ]. [𝐷]
=
𝑘2 [𝐴]. [𝐵]
Where K = the equilibrium constant of reaction (constant at given temperature).
Applications
 Determination of electrolytes strength i.e., weak electrolyte or strong electrolyte.
 Determination of water dissociation constant.
 Determination of hydrogen ion exponent (pH).
 Dissociation of weak acids and bases.
 The hydrolysis of salts.
 Preparation of buffer solution
Common ion Effect:
The degree of ionization of an electrolyte is suppressed by the addition of a strong electrolyte
containing common ion. This effect is known common ion effect.
In other words: The phenomenon of lowering the degree of ionization of a weak electrolyte
by adding a solution of a strong electrolyte having a common ion is called common ion
effect. OR
The common ion effect is an effect that suppresses the ionization of an electrolyte when
another electrolyte (which contains an ion which is also present in the first electrolyte, i.e. a
common ion) is added. It is considered to be a consequence of Le Chatlier’s principle (or the
Equilibrium Law).
The common ion effect is seen when weak and strong electrolytes are mixed.

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Since NH4OH is a weak electrolyte and NH4Cl is a strong electrolyte, the common ion effect
is seen.
Example of common ion effect:
Solutions containing both Sodium chloride NaCl and Silver chloride AgCl also contain a
common ion, Cl- ion.
The equilibrium is as follows:
AgCl ↔Ag++Cl-
The equilibrium will be moved to the left to generate more solid AgCl.
As a result, the solubility of AgCl will decrease.
Hydrogen sulfide (H2S) is a weak electrolyte. It is partially ionized when in aqueous
solution.therefore there exists an equilibrium between un-ionized molecules and constituent
ions in an aqueous medium as follows.
H2S ⇌ H+ + HS−
By applying the law of mass action, we have
[𝐻+][𝐻𝑆−]
𝐾𝑎 =
[𝐻2𝑆]

Hydrochloric acid (HCl) is strong electrolyte, which nearly completely ionizes as

HCl→ H+ + HS-
if HCl is added to the H2S solution, H+ is a common ion and creates a common ion effect.
Due to the increase in concentration of H+ ions from the added HCl, the equilibrium of the
dissociation of H2S shifts to the left and keeps the value of Ka constant. Thus, the dissociation
of H2S decreases, the concentration of un-ionized H2S increases, and as a result, the
concentration of sulphide ions decreases.
Types of acid base titrations
Acidimetry:
An acid-base titration in which an alkali is titrated with a standard solution of acid.
It is a direct or residual volumetric analysis of base with a standard acid.
Organic substances:
Urea, sodium salicylate, diphenhydramine, emetine hydrochloride, meprobamate,
paramethadione, pyrazinamide.
Inorganic Substances:
Sodium bicarbonate, milk of magnesia, ammonium chloride, calcium hydroxide, lithium
carbonate, zinc oxide.
Direct titration:
Burette - Standard acid and conical flask –base being analysed, Indicator: Methyl Orange.
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Example: Sodium Carbonate

Principle:
2NaHCO3 + H2SO4 → Na2SO4 + 2H2O + 2CO2
Sodium Sulpjhate

Residual Titration or back titration:

Reasons:
 one of the reactants is volatile, for example ammonia.
 an acid or a base is an insoluble salt, for example calcium carbonate
 a particular reaction is too slow
 direct titration would involve a weak acid - weak base titration
(The end-point of this type of direct titration is very difficult to observe)
Case I: when a chemical reaction proceeds rather slowly or sluggishly
Case II: when the substance under determination fails to give a sharp and distinctly visible
end-point with an indicator by direct titration.
When the rate of reaction between basic compounds with an acid is slow. In this, the solution
of the base is treated with an excess of accurately measured standard acid and excess
acid is subsequently titrated with standard base.
Example: assay of zinc oxide.
ZNO + H2SO4 → ZNSO4 + 2H2O
Zinc Sulpjhate

Alkalimetry
An acid-base titration in which an acid is titrated with a standard solution of an alkali.
or
It is estimation of acid or acidic drugs by titration with standard alkali.
Direct titration
Standard base solution is taken into burette and acid being assayed taken in to conical flask.
Example: assay of boric acid.
Principle:
H3BO3 + 2NaOH → Na2BO4 + 2H2O
Boric acid Sodium borate
Residual Titration:
When the rate of reaction between acidic compounds with a base is slow. In this, the solution
of the acid is treated with an excess of accurately measured standard base and excess base is
subsequently titrated with standard acid.

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Example: Assay of aspirin.

Difference
Blank Titration Back Titration
Definition: Ex: Acetyl salicylic acid in Aspirin
A blank titration is a titration without an
Many reactions are slow or un favourable for
analyte being present, only the solvent used
direct titration. Aspirin is a weak acid that
in the analyte solution.
also undergoes slow hydrolysis; i.e., each
Use:
aspirin molecule reacts with two hydroxide
The titration error can be reduced by using a
ions. To overcome this problem, a known
blank titration because in a blank titration the
excess amount of base (NaOH) is added to
quantity of titrant required to reach the
the sample solution and HCl titration is
endpoint in the absence of analyte can be
carried out to determine the amount of
subtracted from the quantity of titrant
unreacted base. This is subtracted from the
required to reach the endpoint in the presence
initial amount of base to find the amount of
of an analyte. This reduces the titration error.
base that reacted with the aspirin and hence
the quantity of aspirin in the analyte can
be known

INDICATORS
Principally organic dyes, inorganic substances, compounds capable of fluorescence and
chemiluminescent systems may act as acid-base indicators. The most important group of
these indicators is the one of organic dyes.
Classification
1.Azo dyes EX: Methyl orange, methyl red, Congo red, chyrsoidine
2.pthaleins Ex: Phenolphthalein’s, Eosin, Fluorescein
3.sulphonapthalein Ex: Bromocresol green, purple, blue, cresol red, phenol red, thymol blue,
bromophenol blue
4. Triphenyl methane dyes Ex: crystal violet, rosaniline(magenta)
Indicator pH range Colour change
Methyl orange 3.2-4.5 Pink to yellow
Methyl red 4.4-6.5 Red to yellow
Litmus 5.5-7.5 Red to blue

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Phenol red 6.8-8.4 Yellow to red

Phenolphthalein 8.3-10.5 Colourless to pink

Theories
1. Oswald theory
2. Quinoid theory
Ostwald theory
According to Ostwald these indicators are such weak acids (HI) or bases (IOH) whose
colour is different from that of the indicator-ion formed by their dissociation.
For instance, methyl orange, an indicator-base, is present as a yellow undissociated
indicator molecule in alkaline medium. Due to the neutralization of the base-molecule in
acidic medium a completely dissociated salt will be produced. The indicator-cation thus
formed is of a red colour.
Indicator-acids HI dissociate in aqueous solution as follows:
H I ↔ H + + I-
Applying the law of mass action to this dissociation:

[𝐻+][𝐼−]
=KA
[𝐻𝐼]

From which
[𝐻𝐼] [𝐼−]
[H+] = KA [𝐼−] or pH = pK A + log [𝐻𝐼]

Whose colour is called "acid colour" while [I-] denotes the concentration of the indicator
anions, the colour of which is called "alkaline colour"; KA is the dissociation constant of the
indicator-acid and pKA represents the negative logarithm of the same.
The colour of the indicator is defined at a given pH value by the concentration ratio of the
acid (HI) and alkaline (I~) forms. The equilibrium of the indicator system is shifted by
decreasing the pH in the direction of more HI-formation, while the increase of the pH favours
the formation of the indicator-anion I". The colour of the indicator is consequently a function
of pH. When the indicator is dissociated in about 50%, i.e. [HI] « [I~], the colour is
transitional. The corresponding pH value at 50% ionization may be called "transition point".
pH = -log KA = pK1
In this equation pK represents the indicator exponent.
The indicator bases may be characterized similarly to the indicator acids:
I O H ↔ I+ + OH-
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[𝐼+][𝑂𝐻−]
=KB
[𝐼𝑂𝐻]

Taking the ion product of water into consideration:

[𝐼+]𝐾𝑤
=KB
[𝐻+][𝐼𝑂𝐻]
𝐾𝑤 [𝐼+]
[H+] = [𝐾𝐵] [𝐼𝑂𝐻]

[𝐼𝑂𝐻]
pH=14 - pKB +log [𝐼+]

Where Kw represents the ion product of water, KB denotes the dissociation constant of the
Indicator base, and pKB the negative logarithm of it; IOH means the undissociated indicator
base, the colour of which is the alkaline colour; the acid colour is due to the I + ion.
The Colour change effected by pH may be interpreted similarly to the colour change of
indicator acids. At the transition point:
𝐾𝑤
[H+] = [𝐾𝐵] OR pH = 14 - pKB

The expression 14 - pKB is constant and characteristic of the indicator base, so it may be
denoted as pK1, therefore:
pH = pK1
Example: Phenolpthein (weak acid, pKa -9.4) and methyl orange (weak base)
It can be represented as HPh. It ionises in solution to a small extent as:
HPh ↔ H+ + Ph
Colourless Pink
Applying law of mass action
K = [H+] [Ph-]/[HpH]
The undissociated molecules of phenolphthalein are colourless while Ph- ions are pink in
colour. In presence of an acid the ionisation of HPh is practically negligible as the
equilibrium shifts to left hand side due to high concentration of H + ions. Thus, the solution
would remain colourless. On addition of alkali, hydrogen ions are removed by OH - ions in
the form of water molecules and the equilibrium shifts to right hand side. Thus, the
concentration of Ph- ions increases in solution and they impart pink colour to the solution.
Quinonoid theory:
Friedlander and witt, the term chromophore was applied to the system responsible for
imparting colour to a compound.
Chromophore – Greek word -colour carrier.

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a)The acid-base indicators exist in two tautomeric forms having different structures. Two
forms are in equilibrium. One form is termed benzenoid form and the other quinonoid
form.

(b) The two forms have different colours. The colour changes in due to the inter
conversation of one tautomeric form into other.
(c) One form mainly exists in acidic medium and the other in alkaline medium.
Thus, during titration the medium changes from acidic to alkaline or vice-versa.
The change in pH converts one tautomeric form into other and thus, the colour change
occurs.
P - Nitrophenol: In alkaline solution yellow colour and while in acid solution is colourless
due to benzenoid form.

Phenolphthalein has benziod form in acidic medium and thus, it is colour.

Chemically: 3,3-bis(4-hydroxyphenyl)-2-benzofuran-1-one

Methyl orange (Weak base): chemically:

sodium;4-[[4-(dimethylamino)phenyl] diazenyl] benzenesulfonate
it has quinonoid form in acidic solution and benzenoid form in alkaline solution. The colour
of benzenoid form is yellow while that of quinoniod form is red.

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Methyl orange is a weak base and is yellow in colour in the unionised form.

TYPES
1. Strong acid –Strong base Ex: HCl, H2SO4, Perchloric Acid, HNO3.
2. Weak acid –Strong base Ex: Aspirin, Acetic acid
3. Strong acid –Weak base Ex: NH4OH, HCl
4. Weak acid –Weak base Ex: NH4OH, Acetic acid
Direct acid-base titrations (aqueous phase)
Strong acid vs strong base:
The titration curve obtained from the titration of a strong acid with a strong base. The
pH remains low until just before the equivalence point, when it rises rapidly to a high value.
In many titrations a coloured indicator is used, although electrochemical methods of end-
point detection are also used. An indicator is a weak acid or base that changes colour
between its ionised and un-ionised forms; the useful range for an indicator is 1 pH either side
of its pKa value. For example, phenolphthalein (PP) pKa 9.4 (colour changes between pH
8.4 and pH 10.4) undergoes a structural rearrangement as a proton is removed from one
of its phenol groups when the pH rises, and this causes the colour change. Methyl orange
(MO) pKa 3.7 (colour changes between pH 2.7 and pH 4.7) undergoes a similar pH-
dependent structural change. Both these indicators fall within the range of the inflection of
the strong acid/strong base titration curve.
HCl + NaOH →NaCl + H2O

Examples:

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Strong acid/strong base titrations are used in pharmacopeial assays of: perchloric acid,
hydrochloric acid, sulphuric acid and thiamine hydrochloride.
Weak acid/strong base and weak base/strong acid titration
On addition of a small volume of the strong acid or strong base to a solution of the weak base
or weak acid, the pH rises or falls rapidly to about 1 pH unit below or above the pKa value of
the acid or base. Often a water-miscible organic solvent such as ethanol is used to dissolve
the analyte prior to the addition of the aqueous titrant. Figure shows a plot of pH when 1 M
NaOH is added to 25 ml of a 1 M solution of the weak acid aspirin. In the case of aspirin, the
choice of indicator is restricted by where the inflection in its titration curve lies; PP is suitable
as an indicator whereas MO is not. In the example of the titration of quinine with
hydrochloric acid (Fig), MO is a suitable indicator because it falls within the inflection of the
titration curve.

Weak Acid –Aspirin Weak Base-Quinine

Weak acid/strong base titration is used in the pharmacopeial assays of: benzoic acid, citric
acid, chlorambucil injection, mustine injection, nicotinic acid tablets and undecanoic
acid.
Additional Notes:
Indirect titrations (aqueous medium):
These can be of the strong acid/strong base, weak acid/strong base or weak base/strong acid
type. The more common examples are weak acid/strong base.
Estimation of esters by back titration Excess of sodium hydroxide is added to the ester. The
following reaction occurs:
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RCOOR’+ XSNaOH → RCOONa + R’ OH

The XSNaOH is back titrated with HCl using PP as an indicator. This procedure is used in
pharmacopoeial assays of: benzyl benzoate, dimethyl phthalate, ethyl oleate, methyl
salicylate, cetostearyl alcohol, emulsifying wax, castor oil, arachis oil, cod liver oil and
coconut oil.
Saponification value:
The assay of fixed oils provides a special case of ester hydrolysis since they are triesters of
glycerol. The saponification value for a fixed oil is the number of mg of potassium hydroxide
(KOH) equivalent to 1 g of oil. A high value means rancidity, a low value possible
adulteration with mineral oil. Almost all edible oils have a saponification value between 188
and 196. Hydrolysis of the fixed oil is carried out with ethanolic KOH.
This procedure is used in the pharmacopoeial assays of: castor oil, cod liver oil, cotton seed
oil, almond oil and sesame seed oil.
Acid values are also determined for fixed oils. The acid value for a substance is the number
of mg KOH required to neutralized 1g of the substance when titrated with 0.1 M ethanolic
KOH to a PP end-point. This value is quoted for many fixed oils in order to eliminate rancid
oils, which contain large amounts of free fatty acid. Typically, acid values for fixed oils are in
the range of 1–2.
Estimation of alcohols and hydroxyl values by reaction with acetic anhydride (AA):
Alcohols can be determined by reaction with excess acetic anhydride (AA). This is a useful
titrimetric method because the alcohol group is difficult to estimate by any other means

The excess AA and acetic acid may be back titrated with NaOH using PP as an indicator. In a
related assay, a hydroxyl value is determined for a fixed oil. A 1:3 mixture of AA in pyridine
is used in the determination; the pyridine is present as a catalyst. The hydroxyl value may be
defined as: The number of mg of KOH required to neutralise a blank titration of the reagents
– the number of mg KOH required to neutralise excess AA + acetic acid after reaction with 1
g of the test substance.
NEUTRALIZATION CURVES
A titration curve is the plot of the pH of the analyte solution versus the volume of the
titrant added as the titration progresses.

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S.NO TYPES Example

1 Strong Acid- Strong Base HCl and NaOH
2 Weak Acid -Strong Base Acetic acid, Ethanoic acid
and NaOH
3 Strong Acid -Weak Base HCl and NH3
4 Weak Acid-Weak base Ethanoic acid and NH3

In a titration, the equivalence point is the point at which exactly the same number of moles of
hydroxide ions have been added as there are moles of hydrogen ions. In a titration, if the base
is added from the burette and the acid has been accurately measured into a flask. The shape of
each titration curve is typical for the type of acid-base titration.

The pH does not change in a regular manner as the acid is added. Each curve has horizontal
sections where a lot of bases can be added without changing the pH much. There is also a
very steep portion of each curve except for weak acid and the weak base where a single drop
of base changes the pH by several units. There is a large change of pH at the equivalence
point even though this is not centred on pH 7. This is relevant to the choice of indicators for
each type of titration.
Strong Acid and Strong Base;

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1)HCL + H2O → H3O + Cl- (The pH of the analyte is low (it predominantly contains
H3O from dissociation of HCl).
2) HCL + NaOH → NaCl + H2O
NaOH is added dropwise, H3O slowly starts getting consumed by OH - produced by
dissociation of NaOH. Analyte is still acidic due to predominance of H 3O ions. the pH
recorded at a time point just before complete neutralization takes place. addition of NaOH
continues, pH starts becoming basic because HCl has been completely neutralized and now
excess of OH- ions are present in the solution (from dissociation of NaOH).

Titration of a weak acid with a strong base

The pH of the analyte is low (it predominantly contains H 3O from dissociation of

CH3COOH). But acetic acid is a weak acid, so the starting pH is higher than i case 1 where
had a strong acid (HCl).
CH3COOH +H2O ↔CH3COO- + H3O
As NaOH is added dropwise, H3Oslowly starts getting consumed by OH- (produced by
dissociation of NaOH). But analyte is still acidic due to predominance of H 3Oions.
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This is the pH recorded at a time point just before complete neutralization takes place
CH3COOH +NaOH ↔CH3COONa- + H2O
Note: The pH is not neutral at the equivalence point. The solution is basic (pH ~ 9) at the
equivalence point. at the equivalence point the solution contains CH3COONa salt. This
dissociates into acetate ions CH3COO-and sodium ions Na+ CH3COO-is relatively a strong
base (weak acid CH3COOH has a strong conjugate base), and will thus react with H 2O to
produce hydroxide ions (OH-) thus increasing the pH to ~ 9 at the equivalence point.

Beyond the equivalence point (when sodium hydroxide is in excess) the curve is identical to
HCl-NaOH titration curve.
Titration of a strong acid with a weak base
Principle Reaction:
HCl + NH3 → NH4 + Cl
Analyte is hydrochloric acid HCl (strong acid) and the titrant is ammonia NH 3 (weak base).
HCl + H2O → H3O + Cl-
NH3 is added dropwise, H3O slowly starts getting consumed by OH - produced by dissociation
of NH3.
NH3 + H3O ↔ NH4 + H2O
Record the pH.
This is the equivalence point (halfway up the steep curve). At this point, moles of NH 3 added
= moles of HCl in the analyte.
the pH is not neutral at the equivalence point. The solution is in fact acidic (pH ~ 5.5) at the
equivalence point.
At the equivalence point, the solution only has ammonium ions NH 4 and Cl-. But the
ammonium ion NH4 is the conjugate acid of the weak base NH3. So NH4 is a relatively strong
acid (weak base NH3 has a strong conjugate acid), and thus NH4 will react with H2O to
produce hydronium ions making the solution acidic.
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After the equivalence point, NH3 addition continues and is in excess, so the pH
increases. NH3is a weak base so the pH is above 7, but is lower than what we saw with a
strong base NaOH.

Titration of a weak base with a weak acid

Analyte is NH3(weak base) and the titrant is acetic acid CH3COOH (weak acid).

NH3 + CH3 COOH → CH3 COO- + NH4+

There is just what we call a ‘point of inflexion’ at the equivalence point. Lack of any steep
change in pH throughout the titration renders titration of a weak base versus a weak acid
difficult.

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