• Objectives
1- Ensure the quality and purity of all ingredients involved in
the pharmaceutical industry and their compliance to the
pharmacopeia.
2- Know the official methods for limit test.
3- Apply good laboratory practice and safety measures.
What is an impurity ?
Any material that affects the purity of
the drug or the desired substance.
Sources of impurities
1- Organic impurities such as
Intermediates :-
The compounds produced during the synthesis of the drug.
O OH O O O
[O] H2 N NH2
NH
O
O N
H
Benzoin Benzil Phenytoin
By-products :-
The unplanned compounds produced during the synthesis.
OH OH OCOCH3
(CH3CO)2O
2 +
NH2 NHCOCH3 NHCOCH3
p-Aminophenol Paracetamol By-product
Degradation products :-
The compounds produced due to decomposition of the drug Ex:
aspirin is known to undergo decomposition by hydrolysis into
salicylic acid.
COOH COOH
O CH3 OH
+ CH3COOH
O
Related products :-
The compounds with chemical structure similar to that of the
drug.
Ex. Enantiomers, E/Z isomers.
2- Inorganic impurities such as
e.g. Chloride, Sulphate, Iron, Lead and Cupper.
3- Residual solvents ???
Official methods carried out for impurities control
• Identification and determination of
physicochemical data (color, identity tests,
boiling/melting point, optical rotation,
……etc)
• Quantitative determination of pure
pharmaceutical substances
• Tests designed for impurities, e.g. limit tests
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Limit tests
●Semi-quantitative tests to identify and control
small quantities of impurities.
● All limit tests are designed for the impurities.
( NOT FOR THE DRUG )
● Are designed to show whether a drug
substance contain more or less than the
permitted level of specific impurity.
●All limit tests involve a direct comparison of a
sample experimental results against a standard
experimental results.
Limit tests can be performed by :
1. Nessler tubes method (Compare color or turbidity)
2. Chromatography : TLC & HPLC (Compare density of
spot and area under the peak of standard and impurity
in sample)
3. Spectrophotometry (Compare absorbance of standard
and impurity in sample)
Nessler tubes method
● Involve converting the impurity to a
detectable Opalescense/Turbidity or Color.
Then visual comparison against a standard
treated in the same manner.
For Color For turbidity comparison
comparison
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Nessler tubes
● Are cylindrical flat bottomed glass tubes.
● The two nessler tubes are identical in:-
[Link].
[Link] of marks.
[Link] of glass.
● Nessler tubes have a specific rack
So, remember never put them in a
stand position.
Limit Test for Salicylic Acid In
Aspirin Tablets (USP2007)
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Structure :
COOH COOH
O CH3 OH
Aspirin Salicylic acid
(Acetyl Salicylic acid)
Sources of Salicylic Acid Impurities :
1- Incomplete conversion to aspirin during manufacture.
COOH
COOH
OCOCH 3
OH
+ CH 3 COCl
aspirin
salicylic acid
2- Hydrolysis of aspirin on storage (under unfavorable
conditions).
COOH COOH
OCOCH 3 OH
hydrolysis
+ CH 3 COOH
aspirin salicylic acid
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Principle :
The test depends on the formation of violet colored
ferric salicylic acid complex by the reaction of
salicylic acid impurities with ferric ammonium
sulphate.
COOH COOH_
OH O
+3
Fe
+ FeNH4(SO4)2
3
Violet Color
The color produced is compared with that of standard
containing specified amount of salicylic acid treated
in the same manner.
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Preparation of sample
Mix well
Complete to 100 mL by Dist. Water
Shake for 5 min.
20 mL Ethanol
100 mL
0.200 g of powdered aspirin
tablets (sample)
Filter
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Filtration
1- Folding of filter paper.
2- Filter small amount then discard
after washing the conical flask (repeat twice).
3- Continue filtration till sufficient amount
of filtrate is collected.
Standard Sample
1mL Ferric amm.
4 sulphate 3
3 Complete water to 50 mL 2
with distilled water
2 2 mL Ethanol
1 3 mL freshly prepared 1 10.0 ml of
0.01 % w/v salicylic acid filtrate
Mix thoroughly then Compare the color within 5 mins Not
After. 17
Conclusion of the test :
● If the color of the sample is
much heavier than that of Above the limit
the standard, the present
impurity is called …
● If the color of the sample
is equal to or less than that Within the limit
of the standard, the present
impurity is called …
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Calculations :
USP 2007 Standard : 3.0 mL of 0.01 % w/v Salicylic acid
0.01 g of Salicylic acid 100 mL of Solution
X g of Salicylic acid 3 mL of Solution
Allowed amount (X) in 10 mL sample = 0.01 x 3 / 100 = 0.0003 g
= 0.3 mg Salicylic acid
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Note
• From USP 2008 until now , limit test of salicylic
acid in aspirin tablets is done by HPLC.
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