Limit Tests for Non-Metallic Impurities

LIMIT TESTS FOR NON-METALLIC IMPURITIES

Non-metallic impurities, such as boron, free halogens (I₂, Br₂ and Cl₂) and selenium in pharmaceutical substances usually contribute untoward reactions, skin manifestations and are found to be toxic to healthy tissues.

A few typical examples are described below which essentially contains the above cited nonmetallic impurities :

1. Boron

A. Salbutamol Sulphate : Boron shows its presence in the above compound as a result of the use of sodium borohydride (NaBH₄) in the manufacturing process. The estimation depends upon the conversion of boron to borate and the organic matter is subsequently destroyed by ignition with anhydrous sodium carbonate. The quantity of boron is finally determined by colorimetric assay.

Materials Required : Salbutamol sulphate 50 mg ; solution of an equimolar mixture of anhydrous sodium carbonate and potassium carbonate (3% w/v in DW) 5.0 ml ; Solution of curcumin (0.125% w/v in glacial acetic acid) 3.0 ml ; mixture of H₂SO₄ and glacial CH₃COOH (5 ml : 5 ml) 3.0 ml ; ethanol (96%) 100 ml ; solution of boric acid (dissolve 5 g of boric acid in a mixture of 20 ml DW and 20 ml absolute ethanol and dilute to 250 ml with absolute ethanol) : 100 ml.

Procedure : To 50 mg of substance add 5 ml of a 3% w/v solution of an equimolar mixture of anhydrous Na₂CO₃ and K₂CO₃, evaporate to dryness on a water-bath and dry at 120°C. Ignite the residue rapidly until the organic matter has been destroyed, allow to cool and add 0.5 ml DW and 3 ml freshly prepared 0.125% w/v soln. of curcumin in glacial acetic acid. Warm gently to effect solution, allow to cool and add 3 ml of a mixture of H₂SO₄, with stirring, to 5 ml of glacial acetic acid. Mix and allow to stand for 30 minutes. Add sufficient ethanol (96%) to produce 100 ml, filter and measure the absorbance of the filtrate at the maximum of 555 nm. Calculate the content of boron from a reference curve prepared from the absorbance obtained by treating suit-able aliquots of a solution of boric acid in the same manner.

Prescribed Limits : Not more than 50 ppm.

2. Free Halogens

A few typical examples of pharmaceutical chemicals in which free halogens like Iodine, Bromine, Fluo-rine and Chlorine are present as non-metallic impurities are given below.

A. Clioquinol : (Free Iodine)

Materials Required : Clioquinol 1.0 g ; potassium iodide 1.0 g ; H₂SO₄ (1 M) 1.0 ml ; chloroform 2.0 ml ; sodium thiosulphate (0.005 M) 0.1 ml.

Procedure : Shake 1.0 g with a solution of 1 g potassium iodide in 20 ml DW for 30 seconds, allow to stand for 5 minutes and filter. To 10 ml of the filtrate add 1 ml 1 M H₂SO₄ and 2 ml chloroform and shake.

Prescribed Limits : Any colour in the chloroform layer is discharged on the addition of 0.1 ml of 0.005 M sodium thiosulphate.

B. Diethylpropion Hydrochloride : (Free Bromine)

Test : Place 0.05 ml of a 10% w/v solution on starch-iodide paper.

Prescribed Limit : No colour is produced.

C. Doxycycline Hydrochloride : (Free Fluorine)

Materials Required : Doxycyline Hydrochloride : 0.30 g ; oxygen-combustion flask ; 1 L capacity;

Nessler cylinder 100 ml ; zirconyl alizarin solution* : 5.0 ml ; fluoride standard solution (10 ppm F) (dilute 5.0 ml of a 0.0442 % w/v soln. of sodium fluoride, previously dried at 300°C for 12 hours, to 100 ml with DW) : 3.0 ml.

Procedure : Burn 0.30 g, in three equal portions, by the method for oxygen-flask combustion (BP), using a 1 Litre flask and a separate 20 ml portion of DW as the absorbing liquid for each combustion, shaking the flask vigorously for about 15 minutes and transferring to the same 100 ml Nessler cylinder. Add 5 ml of acid zirconyl alizarin solution to the combined liquids, adjust the volume to 100 ml with DW and allow to stand for 1 hour.

Prescribed Limit : The colour of the resulting solution is greater than that obtained by repeating the operation with no substance enclosed in the successive portions of filter paper burnt in the method for oxygen flask combustion, but adding 3.0 ml of fluoride standard solution (10 ppm F) to the combined absorption liquids before adding the acid zirconyl alizarin solution.

D. Chloroform : (Free Chlorine)

Materials Required : Chloroform 10.0 ml ; cadmium iodide solution (5.0% w/v in DW) 1.0 ml ; starch mucilage 0.1 ml.

Procedure : Shake 10 ml of chloroform with 20 ml of freshly boiled and cooled DW for 3 minutes and allow to separate. To the aqueous layer add 1 ml cadmium iodide soln. and 0.1 ml of 10 ml of starch mucilage.

Prescribed Limit : No blue colour is produced.

E. Tetrachloroethylene (Free Chlorine)

Perform the limit test as stated under chloroform. No blue colour is produced.

3. Selenium

A. Selenium Sulphide

Theory : Selenium is very toxic and its contamination is usually controlled by an absorptiometric method after destruction of the organic compound with fuming nitric acid. The latter converts selenium (Se) as selenous acid (H₂SeO₃), which on subsequent treatment with 3,3′-diaminobenzidine under controlled experimental pa-rameters, results into the formation of a highly coloured compound known as 3,4-diaminophenylpiazselenol. The latter is consequently extracted with toluene after making the aqueous solution alkaline, and the colour compared with a standard prepared likewise from a known amount of selenium. The various reactions involved may be expressed as follows :

Materials Required : Selenium sulphide : 10.0 g ; formic acid (2.5 M) : 2.0 ml ; 3,3′-diaminobenzedine tetrahydrochloride solution (0.5% w/v in DW) : 2.0 ml ; ammonia (5 M) : 20 ml ; selenium standard solution (1 ppm Se) (Dilute 2.5 ml of a 0.00654% w/v solution of selenous acid to 100 ml with DW) : 5.0 ml.

Procedure : To 10 g of selenium sulphide add 100 ml DW, mix well, allow to stand for 1 hour with frequent shaking and filter. To 10 ml of the filtrate, add 2 ml of 2.5 M formic acid, dilute to 50 ml with DW, adjust the pH to 2.0 to 3.0 with 2.5 M formic acid, add 2.0 ml of a 3,3′-diaminobenzidine tetrahydrochloride in DW, allow to stand for 45 minutes and adjust the pH to 6.0 to 7.0 with 5 M ammonia. Shake the solution for 1 minute with 10 ml of toluene and allow to separate. Measure the absorbance at 420 nm.

Prescribed Limit : The measured absorbance at 420 nm is not greater than that of a solution prepared by treating 5 ml of selenium standard solution (1 ppm Se) in the same manner (5 ppm, calculated as Se).