10. limit tests

Introduction
Limit Test for Chlorides
Limit Test for sulphates
Limit Test for Heavy metals
Limit Test for Iron
Limit Test for Arsenic
Limit Test for Lead
Reference

1. Limit tests Prepared by G. Nikitha, M.Pharmacy Assistant Professor Department of Pharmaceutical Chemistry Sree Dattha Institute of Pharmacy Hyderabad Subject Name: Pharmaceutical Inorganic chemistry Year: Pharm -D Ist year

2. contents Introduction Limit Test for Chlorides Limit Test for sulphates Limit Test for Heavy metals Limit Test for Iron Limit Test for Arsenic Limit Test for Lead

3. Introduction 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 . Nessler Cylinder

4. 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. The silver chloride produced in the presence of dilute nitric acid makes the test solution turbid., the extent of turbidity depending upon the amount of chloride present in the substance is compared with a standard opalescence produced by addition of silver nitrate to a standard solution having a known amount of chloride and the same volume of dilute nitric acid as used in the test solution. If turbidity from the sample has been less than the standard turbidity, the sample will pass the limit test.

5. Method for the limiting test for chlorides Specific weight of the substance is dissolved in water or the solution and transferred to the Nessler cylinder A. To the solution 1ml of nitric acid is added and volume made up to 50ml with water. Then 1ml of silver nitrate solution is added and the solution stirred and kept aside for 5 minutes. Simultaneously for standard opalescence, place 1ml of 0.05845 per cent w/v solution of sodium chloride in Nessler cylinder B and add 1ml of dilute nitric make up the volume to 50ml with water add 1ml of silver nitrate solution, stirr with glass rod and aside for 5 minutes. The opalescence produced by the sample should not be grater than standard opalescence. If the opalescence from the sample has been less than the standard opalescence the sample will pass the limit test.

6. Test sample standard compound Specific weight of compound (1g) + 10ml of water. 1ml of 0.05845 % W/V solution of sodium chloride Add 1ml of nitric acid, Dilute to 50ml in Nessler cylinder A + 1ml of AgNO 3  solution Add 1ml of nitric acid, Dilute to 50ml in Nessler cylinder B + 1ml of AgNO 3  solution Observe the Opalescence/Turbidity Observe the Opalescence/Turbidity 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.

7. Limit Test for sulphates

8. 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.

9. Method for the limiting test for sulphate A solution of specified quantity of a substance is prepared in water or solution in Nessler’s cylinder and 2ml of dilute Hcl is added. Dilute to 45ml with water add 5ml of barium sulphate reagent, stirr immediately with the glass rod and a side for 5 minutes. Simultaneously for standard opalescence, place 1ml of 0.1089 % w/v solution of potassium sulphate and 2ml of dilute Hcl. Dilute to 45ml with water add 5ml of barium sulphate reagent, stirr immediately with the glass rod and a side for 5 minutes. The opalescence produced by the sample should not be grater than standard opalescence. If the opalescence from the sample has been less than the standard opalescence the sample will pass the limit test.

10. Test sample standard compound Specific weight of compound (1g) + water or solution. 1ml of 0.1089 % W/V solution of potassium sulphate Add 2ml of dilute hydrochloric acid, Dilute to 45 ml in Nessler cylinder, Add 5ml of barium sulphate reagent Add 2ml of dilute hydrochloric acid, Dilute to 45 ml in Nessler cylinder, Add 5ml of barium sulphate reagent Observe the Opalescence/Turbidity Observe the Opalescence/Turbidity 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 .

11. Limit Test for Heavy metals

12. 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 

13. The Indian pharmacopeia has adopted three methods for limit test for heavy metals. Method-I: It is used for the substance which gives a clear colourless solution under specified conditions. Method-II: It is used for the substance which do not give clear colourless solution under the test conditions specified for Method-I Method-III: It is used for the substance that give colourless solution in sodium hydroxide medium. The reagents like acetic acid, ammonia, Hcl, nitric acid, potassium cyanide and sulphuric acid should be lead free

14. Method-I: This method is applicable for the samples which give clear colourless solution under specified conditions of test. 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. Test sample (A) Standard compound (B) Solution is prepared as per the monograph and 25 ml is transferred in Nessler’s 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’. Dilute with water to 35 ml Adjust the pH between 3 to 4 by adding dilute acetic acid ‘Sp’ or dilute ammonia solution ‘Sp’. Dilute with water to 35 ml Add freshly prepared 10 ml of hydrogen sulphide solution, Dilute with water to 50 ml Add freshly prepared 10 ml of hydrogen sulphide solution, Dilute with water to 50 ml Allow to stand for five minutes View downwards over a white surface Allow to stand for five minutes View downwards over a white surface

15. Method II: Use for the substance which do not give clear colorless solution under the specific condition . Test sample Standard compound Weigh specific quantity of test substance, moisten with sulphuric acid and ignite on a low flame till completely charred.  Add few drops of nitric acid and heat to 500 °C. Allow to cool and add 4 ml of hydrochloric acid and evaporate to dryness. Moisten the residue with 10 ml of hydrochloric acid and digest for twominutes Neutralize with ammonia solution and make just acid with acetic acid. Take 2 ml of standard lead solution and dilute to 25 ml with water Adjust the pH between 3 to 4 and filter if necessary. Dilute with water to 35 ml. Add freshly prepared 10 ml of hydrogen sulphide solution. Dilute with water to 50 ml Adjust the pH between 3 to 4 and filter if necessary. Dilute with water to 35 ml. Add freshly prepared 10 ml of hydrogen sulphide solution. Dilute with water to 50 ml

16. Test sample Standard compound Allow to stand for five minutes Allow to stand for five minutes View downwards over a white surface View downwards over a white surface 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.

17. Method III: Use for the substance which gives clear colorless solution in sodium hydroxide solution. Test sample Standard compound Solution is prepared as per the monograph and 25 ml is transferred in Nessler’s cylinder or weigh specific amount of substance and dissolve in 20 ml of water and add 5 ml of dilute sodium hydroxide solution Take 2 ml of standard lead solution Make up the volume to 50 ml with water Add 5 ml of dilute sodium hydroxide solution and make up the volume to 50 ml with water Add 5 drops of sodium sulphide solution. Mix and set aside for 5 min. Add 5 drops of sodium sulphide solution. Mix and set aside for 5 min. View downwards over a white surface View downwards over a white surface 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 .

18. Limit Test for Iron

19. 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 color. The colour produced from a specified amount of substance from the test is compared by viewing vertically with a standard. If the colour from test solution is less dark than standard, then the sample passes the test.

20. Preparation of Standard solution of Iron: It is prepared by adding 0.173g of ferric ammonium sulphate to 1.5ml of HCl and adding sufficient water to produce 1000ml. Each ml of solution contain 0.02mg of iron.

21. Test sample Standard compound 1. Sample is dissolved in specific amount of water and then volume is made up to 40 ml 1. 2 ml of standard solution of iron diluted with water upto 40ml 2. Add 2 ml of 20 % w/v of citric acid (iron free ). Add 2 drops of thioglycollic acid 2. Add 2 ml of 20 % w/v of citric acid (iron free ). Add 2 drops of thioglycollic acid 3. Add ammonia to make the solution alkaline and adjust the volume to 50 ml. Keep aside for 5 min 3. Add ammonia to make the solution alkaline and adjust the volume to 50 ml. Keep aside for 5 min 4. Color developed is viewed vertically and compared with standard solution 4. Color developed is viewed vertically and compared with standard solution 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.

22. Limit Test for Lead

23. Limit Test for Lead Principle: Limit test of lead is based on the reaction of lead and diphenyl thiocabazone ( dithizone ) in alkaline solution to form lead  dithizone complex which is read in color.  Dithizone is green in color in chloroform and lead- dithizone complex is violet in color, so the resulting color at the end of process is red.

24. 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 2 ml of potassium cyanide and 2 ml of hydroxylamine hydrochloride. Add 2 drops of phenol red Add 6ml of ammonium citrate. Add 2 ml of potassium cyanide and 2 ml of hydroxylamine hydrochloride. Add 2 drops of phenol red Make solution alkaline by adding ammonia solution . Extract with 5 ml of dithizone until it becomes green Make solution alkaline by adding ammonia solution . . 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. Add 4 ml of ammonium cyanide To the acid solution add 5 ml of standard dithizone solution. Add 4 ml of ammonium cyanide Shake for 30 mins . Observe the color Shake for 30 mins . Observe the color 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 .

25. Limit Test for Arsenic

26. Limit Test for Arsenic Principle : ( It is also called as Gutzeit   test) Limit test of Arsenic is based on the reaction of arsenic gas with hydrogen ion to form yellow stain on mercuric chloride paper in presence of reducing agents like potassium iodide. It is also called as Gutzeit test and requires special apparatus . Arsenic, present as arsenic acid in the sample is reduced to arsenious acid by reducing agents like potassium iodide, stannous acid, zinc, hydrochloric acid, etc. Arsenious acid is further reduced to arsine (gas) by hydrogen and reacts with mercuric chloride paper to give a yellow stain.   H 3 AsO 4  + H 2 SnO 2  → H 3 AsO 3  + H 2 SnO 3   Arsenic acid            Arsenious acid   H 3 AsO 3  + 3H 2  → AsH 3  + 3H 2 O    Arsenious acid    Arsine The depth of yellow stain on mercuric chloride paper will depend upon the quality of arsenic present in the sample.

27. apparatus

28. It is havi n g a wide mouthed glass bottle of 120 ml capacity having mouth of about 2.5cm in diameter. The bottle is fitted with a rubber bung through which passes a glass tube 20cm long having an external diameter of about 0.8cm and inter diameter of 0.65cm. The tube is constructed at its lower end extremity to about 1mm diameter and there is blown a hole not less 2mm in diameter in the side of tube near the constricted part. The upper end of the glass tube has been fitted with two rubber bungs each having a hole bored centrally and exactly 6.5 mm in diameter. One of the bungs has been fitted to the upper end of the tube while the second bung has to be fitted upon the first bung in such a way that the mercuric chloride paper gets exactly sandwiched between the central perforation of two. The bungs are kept in close contact by using string clip in such a manner that gas evolved from bottle must have to pass through the 0.65mm internal circle of mercuric chloride paper.

29. Procedure: The glass tube is lightly packed with cotton wool, previously moistened with lead acetate   solution and dried, so that the upper surface of the cotton wool is not less than 25 mm below the top of the tube. The upper end of the tube is then inserted into the narrow end of one of the pair of rubber bungs, to a depth of l0 mm (the tube must have a rounded-off end). A piece of mercuric chloride paper is placed flat on the top of the bung and the other bung placed over it and secured by means of the spring clip in such a manner that the holes of the two bungs meet to form a true tube 6.5 mm diameter interrupted by a diaphragm of mercuric chloride paper .

30. The test solution prepared as specified, is placed in the wide-mouthed bottle, 1 g of KI ( AsT ) and 10 g of Zn ( AsT ) are added, and the prepared glass tube is placed quickly in position. The reaction is allowed to proceed for 40 minutes. The yellow stain that is produced on the HgCl 2  paper if As is present is compared by daylight with the standard stains obtained by performing in an identical manner with known quantities of dilute arsenic solution ( AsT ). The comparison of the stains is made immediately at the completion of the test. By matching the intensity and depth of colour with standard stains, the proportion of arsenic in the substance may be estimated. Thus, a stain equivalent to the 1 ml standard stain obtained by performing on l0 g of a substance implies that the proportion of As is 1 part per million.

31. Reference Pharmaceutical Chemistry -Inorganic Volume-1 by G. R. Chatwal . Essentials of Inorganic Chemistry by Katja A. Strohfeldt . Indian Pharmacopoeia. M.L Schroff , Inorganic Pharmaceutical Chemistry. P. GunduRao , Inorganic Pharmaceutical Chemistry, 3rd Edition A.I. Vogel, Text Book of Quantitative Inorganic analysis. Bentley and Driver's Textbook of Pharmaceutical Chemistry .

32. Thank you