International standards and international reference preparations: amphotericin B, vancomycin, capreomycin, cefalotin, demethylchlortetracycline, gentamycin, gramicidin S, kanamycin and kanamycin B, lincomycin, lymecycline, methacycline, paromomycin, rifamycin SV, ristocetin and ristocetin B, spiramycin, and triacetyloleandomycin.

Each of the preparations described here was obtained and evaluated at the request of a WHO Expert Committee on Biological Standardization. Unless otherwise stated, a standard procedure was used to distribute the material into individual ampoules. The procedure was as follows. Upon receipt by the National Institute for Medical Research (NIMR), London, materials were stored temporarily in the dark at a temperature of -10 degrees C or lower, and protected from moisture. At a convenient time they were brought back to room temperature, mixed, and distributed into individual neutral glass ampoules so that each ampoule contained 50-100 mg of powder. If it was known that the material was light-sensitive non-actinic glass ampoules were used. After exhaustive drying in vacuum over phosphorus(V) oxide, the ampoules were either constricted (up to 1963) or fitted with capillary leak plugs, dried for a further period under the same conditions, filled with dry nitrogen, and sealed by fusion of the glass. The total drying period varied from 8 to 38 days according to the nature of the material. After they had been tested for leaks, the ampoules were stored in the dark at -20 degrees C.

The international reference preparation of gramicidin.

The National Institute for Medical Research, London, was requested by the WHO Expert Committee on Biological Standardization to establish an International Reference Preparation of Gramicidin. This preparation was needed to standardize preparations of gramicidin containing predominantly gramicidin A, B and C, for which purpose the International Reference Preparation of Gramicidin S cannot be used. A batch of 100 g of crystalline gramicidin obtained in 1963 was distributed into ampoules in 55 mg amounts and dried in vacuo; the ampoules were then filled with dried nitrogen and sealed. The proposed international reference preparation was assayed biologically against the Master Standard of Gramicidin of the US Food and Drug Administration in 7 laboratories in 6 countries by turbidimetric methods.Significant curvature of the dose-response lines was found for most assays; no single transformation improved the linearity of assays from all laboratories. Although significant heterogeneity of potencies was obtained in 5 laboratories the mean potency ratios for all laboratories only varied over a range of 5% to 6%. The composition of the material is 7% gramicidin B, 50% gramicidin A and 25% gramicidin C; preparations of gramicidin containing appreciably higher concentrations of gramicidin B can be expected to give invalid assays against the international reference preparation.The material has been established as the International Reference Preparation of Gramicidin with a defined potency of 1000 IU/mg. The International Unit of Gramicidin is defined as the activity of 0.001 mg of the International Reference Preparation of Gramicidin.