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Category Antibacterial.

      Streptomycin Sulfate contains not less than 650 μg and not more than 850 μg of C₂₁H₃₉N₇O₁₂ per mg.

Description White or almost white powder; hygroscopic.

Solubility Very soluble in water; practically insoluble in ethanol and in ether.

Stability It is hygroscopic, but it is relatively stable in the dry state. In aqueous solutions, it may become darkened when stored at room temperature, but its antibacterial potency is still retained for several weeks. Therefore, the aqueous solutions should be refrigerated.

Contra-indication; Warning; Precaution See under Gentamicin Sulfate, p. 111.

Additional information Intravenous administration should be avoided.

      See also under Gentamicin Sulfate, p. 111.

Packaging and storage Streptomycin Sulfate shall be kept in tightly closed containers and stored at a temperature not exceeding 30º.

Labelling The label on the container states (1) the number of μg of activity per mg; (2) storage condition; (3) parenteral grade.

Identification

      A. Carry out the test as described in the “Thin-layer Chromatography” (Appendix 3.1), using silica gel G as the coating substance and a mixture of 1 volume of chloroform, 3 volumes of methanol and 2 volumes of strong ammonia solution as the mobile phase. Apply separately to the plate, 1 μl of each of the two solutions containing (A) 1 mg per ml of the test substance and (B) 1 mg per ml of Streptomycin Sulfate RS and at a third point, apply 1 μl of a mixture of equal volumes of solutions (A) and (B). After removal of the plate, allow it to dry in air, spray with a 1 per cent w/v solution of ninhydrin in 1-butanol, and heat at 105º for 2 minutes: the principal red spot in the chromatogram obtained from solution (A) corresponds to that in the chromatogram obtained from solution (B) and the principal red spot in the third chromatogram appears as a single compact spot.

      B. Dissolve 5 to 10 mg in 4 ml of water and add 1 ml of 1 M sodium hydroxide. Heat for 4 minutes in a waterbath. Add a slight excess of 2 M hydrochloric acid and 2 drops of iron(III) chloride TS: a violet colour is produced.

      C. Dissolve 100 mg in 2 ml of water and add 1 ml of dilute 1-naphthol TS and 2 ml of a solution prepared by diluting 1 ml of sodium hypochlorite solution to 10 ml with water: a red colour is produced.

      D. It yields the reactions characteristic of sulfates (Appendix 5.1).

Clarity of solution Allow a 25 per cent w/v solution to stand protected from light, at a temperature of about 20º for 24 hours; it is not more opalescent than reference suspension II (Appendix 4.1).

pH 4.5 to 7.0, in a 25.0 per cent w/v solution (Appendix 4.11).

Loss on drying Not more than 5.0 per cent w/w after drying at 60º at a pressure not exceeding 0.7 kPa (about 5 Torr) for 3 hours (Appendix 4.15).

Sulfated ash Not more than 1.0 per cent w/w (Appendix 5.3).

Methanol Not more than 0.3 per cent w/w. Carry out the test as described in the “Gas Chromatography (Appendix 3.4).

      Test solution Dissolve 1.00 g of the test substance in water and dilute to 25.0 ml with the same solvent.

      Standard solution Dilute 12.0 mg of methanol to 100.0 ml with water.

      Chromatographic system The chromatographic procedure may be carried out using (a) a glass column (1.5 to 2.0 m × 2 to 4 mm) packed with ethylvinylbenzene-divinylbenzene copolymer (150 to 180 μm), (b) nitrogen as the carrier gas at a constant flow rate of 30 to 40 ml per minute, and (c) a flame ionization detector. Maintain the column at a constant temperature between 120º and 140º and the injection port and the detector at a temperature of at least 50º higher than that of the column.

      Procedure Separately inject equal volumes of Standard solution and Test solution into the chromatograph. The area of the peak corresponding to methanol in the chromatogram obtained from Test solution is not more than that in the chromatogram obtained from Standard solution.

Streptomycin B Not more than 3.0 per cent w/w. Carry out the test as described in the “Thin-layer Chromatography” (Appendix 3.1), using silica gel 60 precoated plate and a mixture of 25 volumes of glacial acetic acid, 25 volumes of methanol and 50 volumes of toluene as the mobile phase.

      Diluting solution A freshly prepared mixture of 3 volumes of sulfuric acid and 97 volumes of methanol.

      Test solution Dissolve 200 mg of the test substance in Diluting solution and dilute to 5 ml with the same solvent. Heat under a reflux condenser for 1 hour, cool, rinse the condenser with methanol, and dilute to 20.0 ml with Diluting solution (1 per cent w/v solution).

      Standard solution Dissolve 36 mg of D-mannose in Diluting solution and dilute to 5 ml with the same solvent. Heat under a reflux condenser for 1 hour, cool, rinse the condenser with methanol, and dilute to 50.0 ml with Diluting solution. Dilute 5.0 ml of the solution to 50.0 ml with methanol (0.03 per cent w/v solution expressed as streptomycin B; 1 mg of D-mannose is equivalent to 4.13 mg of streptomycin B).

      Procedure Apply separately to the plate 10 μl of each solution. After removal of the plate, allow it to dry in air and spray with a freshly prepared mixture of equal volumes of a 0.2 per cent w/v solution of 1,3- dihydroxynaphthalene in ethanol and a 20 per cent v/v solution of sulfuric acid and heat at 110º for 5 minutes. Any spot corresponding to streptomycin B in the chromatogram obtained from Test solution is not more intense than that in the chromatogram obtained from Standard solution.

Colourimetric test Dry the test substance and Streptomycin Sulfate RS at 60º over phosphorus pentoxide desiccant at a pressure not exceeding 0.1 kPa (about 0.7 Torr) for 24 hours. Dissolve 100.0 mg of the dried test substance in water and dilute to 100.0 ml with the same solvent. Prepare a standard solution in the same manner using 100.0 mg of the dried Streptomycin Sulfate RS. Place 5.0 ml of each solution separately in two volumetric flasks and in a third flask place 5 ml of water. To each flask add 5.0 ml of 2.0 M sodium hydroxide and heat for exactly 10 minutes in a water-bath. Cool in ice for exactly 5 minutes, add 3.0 ml of a 1.5 per cent w/v solution of ammonium iron(III) sulfate in 0.5 M sulfuric acid, dilute to 25.0 ml with water and mix. Exactly 20 minutes after the addition of the ammonium iron(III) sulfate solutions, measure the absorbances (Appendix 2.2) of the test solution and the standard solution in a 2-cm cell at the maximum at 525 nm, using as compensation liquid the solution prepared from 5.0 ml of water. The absorbance of the test solution is not less than 90.0 per cent of that of the standard solution.

Assay Carry out the microbiological assay of Streptomycin Sulfate according to the “Microbiological Assay of Antibiotics” (Appendix 6.10).

Other requirements Streptomycin Sulfate intended for parenteral administration complies with the following additional requirements.

      Bacterial endotoxins When tested as described in the “Test for Bacterial Endotoxins” (Appendix 8.5), it contains not more than 0.25 Endotoxin Unit per mg of streptomycin.

      Sterility Complies with the “Sterility Test” (Method I, Appendix 10.1).