Limit Test

Limit Test

Limit Test 

Limit test is defined as quantitative or semi quantitative test designed to identify and control small quantities of impurity which is likely to be present in the substance.

Limit test is generally carried out to determine the inorganic impurities present in compound.

Limit test is nothing but to identify the impurities present in the substance and compare it with standard.

Limit = a value or amount that is likely to be present in a substance

Test = to examine or to investigate

Impurities = a foreign matter present in a compound

Importance of Limit tests

To find out the harmful amount of impurities

To find out the avoidable/unavoidable amount of impurities.

Limit test for Chlorides

Principle:

Limit test of chloride is based on the reaction of soluble chloride with silver nitrate in presence of dilute nitric acid to form silver chloride, which appears as solid particles (Opalescence) in the solution.

Procedure:  

Test sample

Standard compound

Specific weight of compound is dissolved in water or solution is prepared as directed in the pharmacopoeia and transferred in Nessler cylinder Take 1ml of 0.05845 % W/V solution of sodium chloride in Nessler cylinder

Add 1ml of nitric acid

Add 1ml of nitric acid

Dilute to 50ml in Nessler cylinder

Dilute to 50ml in Nessler cylinder

Add 1ml of AgNO3 solution

Add 1ml of AgNO3 solution

Keep aside for 5 min

Keep aside for 5 min

Observe the Opalescence/Turbidity

Observe the Opalescence/Turbidity

 

Observation:

The opalescence produce in sample solution should not be greater than standard solution.

If opalescence produces in sample solution is less than the standard solution, the sample will pass the limit test of chloride and visa versa.

Reasons:

Nitric acid is added in the limit test of chloride to make solution acidic and helps silver
chloride precipitate to make solution turbid at the end of process.

Limit test for Sulphates

Principle:

Limit test of sulphate is based on the reaction of soluble sulphate with barium chloride in presence of dilute hydrochloric acid to form barium sulphate which appears as solid particles (turbidity) in the solution.

Procedure:

Test sample

Standard compound

Specific weight of compound is dissolved in water or solution is prepared as directed in the pharmacopoeia and transferred in Nessler cylinder

Take 1ml of 0.1089 % W/V solution of potassium sulphate in Nessler cylinder

Add 2ml of dilute hydrochloric acid

Add 2ml of dilute hydrochloric acid

Dilute to 45 ml in Nessler cylinder

Dilute to 45 ml in Nessler cylinder

Add 5ml of barium sulphate reagent

Add 5ml of barium sulphate reagent

Keep aside for 5 min

Keep aside for 5 min

Observe the Turbidity

Observe the Turbidity

 

Barium sulphate reagent contains barium chloride, sulphate free alcohol and small amount of potassium sulphate.

Observation:

The turbidity produce in sample solution should not be greater than standard solution. If turbidity produces in sample solution is less than the standard solution, the sample will pass the limit test of sulphate and vice versa.

Reasons:

Hydrochloric acid helps to make solution acidic. Potassium sulphate is used to increase the sensitivity of the test by giving ionic concentration in the reagent. 

Alcohol helps to prevent super saturation.

Limit test for Iron

Principle:

Limit test of Iron is based on the reaction of iron in ammonical solution with thioglycollic acid in presence of citric acid to form iron thioglycolate which is pale pink to deep reddish
purple in colour.

Procedure:

Test sample

Standard compound

Sample is dissolved in specific amount of water and then volume is made up to 40 ml

2 ml of standard solution of iron diluted with water upto 40ml

Add 2 ml of 20 % w/v of citric acid (iron free)

Add 2 ml of 20 % w/v of citric acid (iron free)

Add 2 drops of thioglycollic acid

Add 2 drops of thioglycollic acid

Add ammonia to make the solution alkaline and adjust the volume to 50 ml

Add ammonia to make the solution alkaline and adjust the volume to 50 ml

Keep aside for 5 min

Keep aside for 5 min

Color developed is viewed vertically and compared with standard solution

Color developed is viewed vertically and compared with standard solution

Earlier aamoniumthiocyanate reagent was used for the limit test of iron. Since thioglycolic acid is more sensitive reagent, it has replaced ammonium thiocyanate in the test.

Observation:

The purple color produce in sample solution should not be greater than standard solution. If purple color produces in sample solution is less than the standard solution, the sample will pass the limit test of iron and vice versa.

Reasons:

Citric acid helps precipitation of iron by ammonia by forming a complex with it.

Limit test for Heavy Metals

Principle:

Limit test of heavy metals is based on the reaction of metallic impurities with hydrogen sulfide in acidic medium to form brownish colour solution. Metals that response to this test are lead, mercury, bismuth, arsenic, antimony, tin, cadmium, silver, copper, and molybdenum.

The metallic impurities in substances are expressed as parts of lead per million parts of the substance. The usual limit as per Indian Pharmacopoeia is 20 ppm

Procedure:

The Indian Pharmacopoeia has adopted three methods for the limit test of heavy metals.

Method I: Use for the substance which gives clear colorless solution under the specific condition.

Test sample

Standard compound

Solution is prepared as per the monograph and 25 ml is transferred in Nesslers cylinder

Take 2 ml of standard lead solution and dilute to 25 ml with water

Adjust the pH between 3 to 4 by adding dilute acetic acid ‘Sp’ or dilute ammonia solution ‘Sp’

Adjust the pH between 3 to 4 by adding dilute acetic acid ‘Sp’ or dilute ammonia solution ‘Sp’

Dilute with water to 35 ml

Dilute with water to 35 ml

Add freshly prepared 10 ml of hydrogen sulphide solution

Add freshly prepared 10 ml of hydrogen sulphide solution

Dilute with water to 50 ml

Dilute with water to 50 ml

Allow to stand for five minutes

Allow to stand for five minutes

View downwards over a white surface

View downwards over a white surface

 

Observation:

The color produce in sample solution should not be greater than standard solution. If color produces in sample solution is less than the standard solution, the sample will pass the limit test of heavy metals and vice versa.

Limit test for Lead

Lead is a most undesirable impurity in medicacompounds and comes through use of sulphuric acid, lead lined apparatus and glass bottles use for storage of chemicals.

Principle:

Limit test of lead is based on the reaction of lead and diphenylthiocabazone (dithizone) in alkaline solution to form lead dithizone complex which is read in color.

Dithizone is green in color in chloroform and leaddithizone complex is violet in color, so the resulting color at the end of process is red.

Procedure:

Test sample

Standard compound

A known quantity of sample solution is transferred in a separating funnel

A standard lead solution is prepared equivalent to the amount of lead permitted in the sample under examination

Add 6ml of ammonium citrate

Add 6ml of ammonium citrate

Add 2 ml of potassium cyanide and 2 ml of hydroxylamine hydrochloride

Add 2 ml of potassium cyanide and 2 ml of hydroxylamine hydrochloride

Add 2 drops of phenol red

Add 2 drops of phenol red

Make solution alkaline by adding ammonia solution.

Make solution alkaline by adding ammonia solution.

Extract with 5 ml of dithizone until it becomes green

Extract with 5 ml of dithizone until it becomes green

Combine dithizone extracts are shaken for 30 mins with 30 ml of nitric acid and the chloroform layer is discarded

Combine dithizone extracts are shaken for 30 mins with 30 ml of nitric acid and the chloroform layer is discarded

To the acid solution add 5 ml of standard dithizone solution

To the acid solution add 5 ml of standard dithizone solution

Add 4 ml of ammonium cyanide

Add 4 ml of ammonium cyanide

Shake for 30 mins

Shake for 30 mins

Observe the color

Observe the color

 

Observation:

The intensity of the color of complex, is depends on the amount of lead in the solution.

The color produce in sample solution should not be greater than standard solution.  

If color produces in sample solution is less than the standard solution, the sample will pass the limit test of lead and vice versa.

Reasons:

Ammonium citrate, potassium cyanide, hydroxylamine hydrochloride is used to make pH optimum so interference and influence of other impurities have been eliminated.

Pharmaceutical Impurities and Limit Test Notes PDF

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