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1.6 TEST SOLUTIONS

Acetic Anhydride-Pyridine TS Dissolve 25 ml of acetic anhydride in anhydrous pyridine and dilute to 100 ml with anhydrous pyridine.

Prepare freshly before use.

Acetic-Bromine TS Dissolve 100 g of potassium acetate in glacial acetic acid, and add 4 ml of bromine and sufficient glacial acetic acid to produce 1000 ml.

Alizarin Complexone Dihydrate TS Dissolve 38 mg of alizarin complexone dihydrate in a solution of 30 mg of sodium hydroxide in 25 ml of water. Dilute with water to about 150 ml and add 50 mg of sodium acetate and 2 M hydrochloric acid until a thin layer of the solution is pale pink in colour. Add sufficient water to produce 200 ml.

Aminophenazone TS (Aminopyrazolone TS) A 0.1 per cent w/v solution of 4-aminophenazone in borate buffer pH 9.0.

Ammonium Iron(III) Sulfate TS A freshly prepared 10 per cent w/v solution of ammonium iron(III) sulfate.

Gives a deep red colour with ammonium thiocyanate in acid solutions.

Ammonia TS It contains 9.5 to 10.5 per cent w/w of NH₃. Prepare by diluting 400 ml of strong ammonia solution with water to make 1000 ml.

Store in well-closed containers, at a temperature not exceeding 20º.

Ammonium Chloride TS A 10.7 per cent w/v solution of ammonium chloride.

Ammonium Mercurithiocyanate TS Dissolve 30 g of ammonium thiocyanate and 27 g of mercury(II) chloride in sufficient water to produce 1000 ml.

Ammonium Molybdate TS A 10.0 per cent w/v solution of ammonium molybdate.

Ammonium Molybdate with Ascorbic Acid TS Mix in the following order 1 volume of a 2.5 per cent w/v solution of ammonium molybdate, 1 volume of a 10 per cent w/v solution of ascorbic acid and 1 volume of 3 M sulfuric acid and add 2 volumes of water.

Use within 1 day.

Ammonium Sulfide TS Saturate ammonia TS with hydrogen sulfide by bubbling hydrogen sulfide gas through the solution for 1 minute. The solution is not rendered turbid either by magnesium sulfate TS or by calcium chloride TS. This solution is unstable for use if an abundant precipitate of sulfur is present.

Prepare freshly before use.

Ammonium Thiocyanate TS Dissolve 8 g of ammonium thiocyanate in water to make 100 ml.

Anisaldehyde TS Mix, in order, 0.5 ml of anisaldehyde, 10 ml of glacial acetic acid, 85 ml of methanol and 5 ml of sulfuric acid.

Barium Chloride TS Dissolve 10 g of barium chloride in water to make 100 ml.

Bromelains TS A 1.0 per cent w/v solution of bromelains in a mixture of 1 volume of mixed phosphate buffer pH 5.5 and 9 volumes of saline TS.

Bromine TS (Bromine Water) A saturated solution of bromine, prepared by agitating 2 to 3 ml of bromine with 100 ml of cold water to a glass-stoppered bottle, the stopper of which should be lubricated with petrolatum.

Store in a cool place, protected from light.

Bromophenol Blue TS Dissolve 100 mg of bromophenol blue in 100 ml of ethanol (50 per cent), and filter if necessary.

Bromophenol Blue TS, Aqueous Dissolve 50 mg of bromophenol blue with gentle heating in 3.73 ml of 0.02 M sodium hydroxide and dilute to 100 ml with water.

Cadmium Iodide TS A 5.0 per cent w/v solution of cadmium iodide.

Calcium Chloride TS Dissolve 7.5 g of calcium chloride in water to make 100 ml.

Calcium Hydroxide TS A freshly prepared saturated solution which may be prepared in the following manner. Shake 10 g of calcium hydroxide repeatedly with 1000 ml of water and allow to stand until clear.

Cephalin TS Solvents used to prepare this solution should contain a suitable antioxidant such as butylated hydroxyanisole at a concentration of 0.1 mM.

          To 0.5 to 1 g of acetone-dried ox brain add 20 ml of acetone and leave for 2 hours. Centrifuge for 2 minutes at 500 × g and decant the supernatant liquid. Dry the residue at a pressure of 2 kPa (about 15 Torr) and extract the dried material with 20 ml of chloroform for 2 hours, shaking the mixture frequently. After removal of the solid material by filtration or centrifugation, evaporate the chloroform from the extract at a pressure of 2 kPa (about 15 Torr). Suspend the residue in 5 to 10 ml of saline TS. This stock emulsion may be stored frozen or freeze-dried for 3 months.

Cerium(III) Nitrate TS Dissolve 220 mg of cerium(III) nitrate in 50 ml of water, add 0.1 ml of nitric acid and 50 mg of hydroxylamine hydrochloride and dilute with water to 1000 ml.

Chlorine TS (Chlorine Water) A freshly prepared, saturated solution of chlorine in water.

Copper(II) Sulfate TS Dissolve 12.5 g of copper(II) sulfate in water to make 100 ml.

Copper-Citric TS Dissolve 25 g of copper(II) sulfate, 50 g of citric acid and 144 g of anhydrous sodium carbonate in sufficient water to produce 1000 ml. Diazobenzenesulfonic Acid TS Heat 200 mg of sulfanilic acid with 20 ml of 1 M hydrochloric acid until dissolved, cool to about 4º, and add dropwise, with continuous swirling, 2.2 ml of a 4 per cent w/v solution of sodium nitrite. Allow to stand in ice water for 10 minutes and add 1 ml of a 5 per cent w/v solution of sulfamic acid.

Diphenylamine TS A 1.0 per cent w/v solution of diphenylamine in sulfuric acid. The solution must becolourless.

Store protected from light.

Diphenylamine TS1 Dissolve 1 g of diphenylamine in 100 ml of glacial acetic acid and add 2.75 ml of sulfuric acid.

Use immediately after preparation.

Dithizone TS Dissolve 25.6 mg of dithizone in 100 ml of ethanol.

Store in a cold place, and use within 2 months.

Eosin Y TS A 0.5 per cent w/v solution of eosin Y.

Fast Blue B Salt TS Dissolve 50 mg of fast blue B salt in 10 mL of water

Prepare freshy before use.

Guaiacol TS A 5 per cent w/v solution of guaiacol in ethanol.

Store protected from light.

Hydrogen Peroxide TS (10 volumes) (Dilute Hydrogen Peroxide Solution)

Dilute hydrogen peroxide TS (20 volumes) with an equal volume of water.

Hydrogen Peroxide TS (20 volumes)

Use analytical reagent grade of commerce containing about 6 per cent w/v of H₂O₂ or dilute 1 volume of strong hydrogen peroxide solution with 4 volumes of water.

DESCRIPTION Colourless liquid.

WEIGHT PER MILLILITRE About 1.02 g (Appendix 4.9).

Hydrogen Peroxide TS (100 volumes)

Use Strong hydrogen Peroxide Solution (See under “Reagents”).

Hydrogen Sulfide TS A freshly prepared, saturated solution of hydrogen sulfide in water. It contains about 0.45 per cent w/v of H2S at 20º.

Hydroxylamine TS Dissolve 1 g of hydroxylamine hydrochloride in 50 ml of water, add 50 ml of ethanol and 1 ml of bromophenol blue TS, and then add 0.1 M sodium hydroxide until the solution becomes green.

Hydroxylamine in Ethanol (60 per cent) TS Dissolve 3.5 g of hydroxylamine hydrochloride in 95 ml of ethanol (60 per cent), add 0.5 ml of a 0.2 per cent w/v solution of methyl orange in ethanol (60 per cent), and then add 0.5 M potassium hydroxide in ethanol (60 per cent) until the full yellow colour is obtained. Add sufficient ethanol (60 per cent) to produce 100 ml. The solution complies with the following test.

          To 10 ml add 0.05 ml of 0.5 M potassium hydroxide in ethanol (60 per cent): no change in colour is produced. To a further 10 ml add 0.05 ml of 0.5 M hydrochloric acid: the colour changes slightly towards orange.

Hydroxylamine in Ethanol (90 per cent) TS Dissolve 7.0 g of hydroxylamine hydrochloride in 90 ml of ethanol (90 per cent), warming gently if necessary, and add 1.6 ml of dimethyl yellow TS and sufficient 1 M potassium hydroxide in ethanol (90 per cent) to give a full yellow colour. Dilute with ethanol (90 per cent) to 100 ml.

Hypophosphorous TS Dissolve, by heating gently, 10 g of sodium hypophosphite in 20 ml of water and dilute to 100 ml with hydrochloric acid. Allow to settle and decant or filter through glass wool.

Iodine TS Use 0.05 M iodine VS.

Iodine and Potassium Iodide TS Dissolve 500 mg of iodine in a solution of 1.5 g of potassium iodide in 5 ml of water, and dilute with water to 25 ml.

Store protected from light.

Iodine Bromide TS (Iodobromide TS) A 2 per cent w/v solution of iodine monobromide in glacial acetic acid.

Store in glass containers, protected from light.

Iodine Chloride TS (Iodochloride TS) Dissolve 1.65 g of iodine monochloride in 100.0 ml of glacial acetic acid.

Store protected from light.

Iron(III) Chloride TS Dissolve 9 g of iron(III) chloride in water to make 100 ml.

Iron(III) Chloride-Sulfamic Acid TS A solution containing 1.0 per cent w/v of iron(III) chloride and 1.6 per cent w/v of sulfamic acid.

Lead Acetate TS A 9.5 per cent w/v solution of lead(II) acetate in carbon dioxide-free water.

Store in well-closed containers.

Magenta TS, Decolorized (Fuchsin TS, Decolorized) Dissolve 0.1 g of basic magenta in 60 ml of water and add 1 g of anhydrous sodium sulfite dissolved in 10 ml of water. Slowly add 2 ml of hydrochloric acid, stirring continuously, and dilute to 100 ml with water. Allow to stand protected from light for at least 12 hours. Shake with sufficient decolorizing charcoal (0.2 to 0.3 g) to remove the colour and then filter immediately. If the solution becomes cloudy, filter before use. If on standing the solution becomes violet, decolorize again by adding decolorizing charcoal. The solution complies with the following test.

SENSITIVITY TO FORMALDEHYDE To 1.0 ml of the solution add 1.0 ml of water and 0.1 ml of aldehyde-free ethanol. Add 0.2 ml of a solution containing 0.01 per cent w/v of formaldehyde. A pale pink colour is produced within 5 minutes.

Store protected from light.

Magnesium Sulfate TS A 12.3 per cent w/v solution of magnesium sulfate.

Mercuric-Potassium Iodide TS (Mayer’s Reagent) Dissolve 1.358 g of mercury(II) chloride in 60 ml of water. Dissolve 5 g of potassium iodide in 10 ml of water. Mix the two solutions, and dilute with water to 100 ml.

Mercuric-Potassium Iodide TS, Alkaline (Nessler’s Reagent) Dissolve 143 g of sodium hydroxide in 700 ml of water. Dissolve 50 g of red mercury(II) iodide and 40 g of potassium iodide in 200 ml of water. Pour the iodide solution into the hydroxide solution, and dilute with water to 1000 ml. Allow to settle, and use the clear supernatant.

Mercury(I) Nitrate TS Dissolve 15 g of mercury(I) nitrate in a mixture of 90 ml of water and 10 ml of dilute nitric acid.

Store in dark, amber-coloured bottles in which a small globule of mercury has been placed.

Mercury(II) Sulfate TS (Denigès’ Reagent) Mix 5 g of yellow mercury(II) oxide with 40 ml of water, and, while stirring slowly, add 20 ml of sulfuric acid; add 40 ml of water and stir until completely dissolved.

2-Naphthol TS Dissolve 5 g of 2-naphthol, freshly recrystallized, in 40 ml of 2 M sodium hydroxide and add sufficient water to produce 100 ml.

Prepare immediately before use.

1-Naphthol TS, Dilute Dissolve 100 mg of 1-naphthol in 3 ml of a 15 per cent w/v solution of sodium hydroxide and dilute to 100 ml with water.

Prepare freshly before use.

Ninhydrin TS (Triketohydrindene Hydrate TS) Dissolve 200 mg of ninhydrin in water to make 10 ml.

Prepare freshly before use.

Nitro-vanado-molybdic TS Solution A Dissolve 10 g of ammonium molybdate in water, add 1 ml of 10 M ammonia and dilute to 100 ml with water.

Solution B Dissolve 0.5 g of ammonium metavanadate in hot water, add 2.8 ml of nitric acid and dilute to 100 ml with water.

          To 100 ml of solution A and 100 ml of solution B add 96 ml of nitric acid and dilute to 500 ml with water.

Oxalic Acid TS Dissolve 6.3 g of oxalic acid in water to make 100 ml.

Prepare freshly before use.

Potassium Chromate TS A 5.0 per cent w/v solution of potassium chromate. Gives a red precipitate with silver nitrate in neutral solutions.

Potassium Dichromate TS Dissolve 7.5 g of potassium dichromate in water to make 100 ml.

Potassium Hexacyanoferrate(II) TS A 5.3 per cent w/v solution of potassium hexacyanoferrate(II).

Potassium Hexacyanoferrate(III) TS Wash about 1 g of potassium hexacyanoferrate(III), in crystals, with a little water, and dissolve the washed crystals in 100 ml of water. Gives a blue colour with solutions of iron(II) salts.

Prepare freshly before use.

Potassium Hydroxide TS, Ethanolic Dissolve 3 g of potassium hydroxide in 5 ml of water and add sufficient aldehyde-free ethanol to produce 100 ml. Allow the solution to stand for 1 hour and decant the clear solution.

Potassium Iodide TS Dissolve 16.5 g of potassium iodide in water to make 100 ml.

Store protected from light.

Potassium Iodide and Starch Solution Dissolve 750 mg of potassium iodide in 100 ml of water, heat to boiling and add, while stirring, a solution of 500 mg of soluble starch in 35 ml of water. Boil for 2 minutes and allow to cool.

SENSITIVITY TO IODINE To 15 ml of the solution add 0.05 ml of glacial acetic acid and 0.30 ml of 0.00050 M iodine. A blue colour is produced.

Prepare freshly before use.

Potassium Iodobismuthate TS Dissolve 10 g of tartaric acid in 40 ml of water and add 850 mg of bismuth oxynitrate. Shake during 1 hour, add 20 ml of a 40 per cent w/v solution of potassium iodide, and shake well. Allow to stand for 24 hours and filter.

Store protected from light.

Potassium Iodobismuthate TS, Acetic Dissolve 8 g of potassium iodide in 20 ml of water and add the solution to a mixture of 850 mg of bismuth oxynitrate, 40 ml of water and 10 ml of glacial acetic acid.

Potassium Iodobismuthate TS, Dilute Dissolve 10 g of tartaric acid in 50 ml of water and add 5 ml of potassium iodobismuthate TS.

Potassium Iodoplatinate TS Add 50 ml of a 5 per cent w/v solution of chloroplatinic(IV) acid to 45 ml of potassium iodide TS and dilute to 100 ml with water.

Store in amber glass containers.

Potassium Permanganate TS Use 0.02 M potassium permanganate VS.

Potassium Permanganate and Phosphoric Acid TS Dissolve 3 g of potassium permanganate in a mixture of 15 ml of phosphoric acid and 70 ml of water and add sufficient water to produce 100 ml.

Pyridylazonaphthol TS A 0.1 per cent w/v solution of pyridylazonaphthol in absolute ethanol.

Resorcinol with Copper(II) Sulfate TS To 80 ml of hydrochloric acid add 10 ml of a 2 per cent w/v solution of resorcinol and 0.25 ml of a 2.5 per cent w/v solution of copper(II) sulfate and dilute to 100 ml with water.

Prepare the solution at least 4 hours before use. Store at 2º to 8º and use within 1 week.

Saline TS A 0.9 per cent w/v solution of sodium chloride, sterilized by “Heating in an Autoclave” (Appendix 12).

Saline TS, Pyrogen-free Saline TS which complies with the requirements of “Pyrogen Test” (Appendix 8.2).

Silver Nitrate TS Use 0.1 M Silver Nitrate VS.

Silver Diethyldithiocarbamate TS Dissolve 1 g of silver diethyldithiocarbamate in 200 ml of a freshly opened bottle or recently distilled pyridine.

Store in light-resistant containers and use within 1 month.

Simulated Gastric Fluid TS Dissolve 2.0 g of sodium chloride and 3.2 g of pepsin in 7.0 ml of hydrochloric acid and sufficient water to make 1000 ml. This test solution has a pH of about 1.2.

Sodium Hexanitrocobaltate(III) TS Dissolve 5 g of cobalt(II) nitrate in water, add 1.5 ml of nitric acid and dilute to 100 ml with water. Dissolve 30 g of sodium nitrite in water and dilute to 100 ml with water. Mix the two solutions, allow to stand and use the clear solution. 

Prepare immediately before use.

Sodium Hydroxide TS Dissolve 4.0 g of sodium hydroxide in water to make 100 ml (about 1 M).

Sodium Hypochlorite TS Use Sodium Hypochlorite Solution (see under “Reagents”).

Sodium Hypochlorite TS, Dilute Dilute 35 ml of sodium hypochlorite solution with water to 100 ml.

Prepare immediately before use.

Simulated Intestinal Fluid TS Dissolve 6.8 g of potassium dihydrogenphosphate in 250 ml of water, mix, and add 77 ml of 0.2 M sodium hydroxide and 500 ml of water. Add 10.0 g of pancreatin, mix, and adjust the resulting solution with either 0.2 M sodium hydroxide or 0.2 M hydrochloric acid to a pH of 6.8±0.1. Dilute with water to 1000 ml.

Sodium Metabisulfide TS Dissolve 10 g of sodium metabisulfite to make 30 ml.

Prepare freshly before use.

Sodium Molybdotungstophosphate TS Boil under a reflux condenser 350 ml of water, 50 g of sodiumt tungstate, 12 g of phosphomolybdic acid and 25 ml of phosphoric acid for 2 hours. Cool and add sufficient water to produce 500 ml.

Store at 2º to 8º, protected from light.

Sodium Phosphate-Sodium Chloride Buffer TS Dissolve 90.0 g of sodium chloride, 10.9 g of anhydrous disodium hydrogenphosphate and 2.45 g of monosodium dihydrogenphosphate dihydrate in sufficient water to produce 1000.0 ml.

Store in a cold place. Any crystals produced should be dissolved by warming. Shake the solution before use.

Sodium Nitrite TS A freshly prepared 10 per cent w/v solution of sodium nitrite.

Prepare immediately before use.

Sodium Sulfide TS Dissolve 1 g of sodium sulfide in water to make 10 ml.

Prepare freshly before use.

Sodium Sulfide TS1 Dissolve, with heating, 12 g of sodium sulfide in 45 ml of a mixture of 10 volumes of water and 29 volumes of glycerol (85 per cent), allow to cool and dilute to 100 ml with the same mixture. It should be colourless.

Sodium Tetraphenylborate TS Dissolve 1.2 g of sodium tetraphenylborate in water to make 200 ml. If necessary, filter before use.

Starch TS Triturate 1 g of starch or soluble starch with 5 ml of water and add with continuous stirring to 100 ml of boiling water, and boil for 1 minute. Cool and use only the clear solution.

Prepare freshly before use.

Sulfuric Acid-Formaldehyde TS Mix 2 ml of formaldehyde TS with 100 ml of sulfuric acid.

Tetrazolium Blue TS, Alkaline Immediately before use mix 1 volume of a 0.2 per cent w/v solution of tetrazolium blue in methanol with 3 volumes of a 12 per cent w/v solution of sodium hydroxide in methanol.

Tin(II) Chloride TS, Stronger Acid Dissolve 40 g of tin(II) chloride in 100 ml of hydrochloric acid.

Store in glass containers and use within 3 months.

Trinitrophenol TS Add 0.25 ml of 10 M sodium hydroxide to 100 ml of a saturated solution of 2,4,6-trinitrophenol in water.