The inhibition of N-acetyl-beta-d-glucosaminase activity by 2-acetamido-2-deoxy-d-gluconolactone was examined. Separation of the (1-->4)- and (1-->5)-lactones was achieved by using paper chromatography or countercurrent distribution and identification was obtained by examination of the relative stability of the components of separated material. Quantitative measurement of the two lactones was by kinetic titration or by a colorimetric method based on their reaction with hydroxylamine. It was shown that only the (1-->5)-lactone acted as an inhibitor of N-acetyl-beta-d-glucosaminase.

THE use of polarography to assay streptomycin has received limited attention. Levy, Schwed and Sackett (1946) have reported on its application at streptomycin concentrations of greater than 200 units/ml. using partially purified material. Bricker and Vail (1951) have used it to study the alkaline degradation of streptomycin and have concluded that the independent determination of streptomycin and mannosidostreptomycin in mixtures of the two is impracticable. The existence of two tautomeric forms of streptomycin has been demonstrated by this technique Heuser, Dolliver and Stiller (1953), whilst Tsukamoto andTachi (1952) have shown that the height of the reduction wave increased with pH increase, and recommend N sodium hydroxide as a base electrolyte. Much of the work on the polarography of streptomycin is summarked by Brezina and Zuman (1958). The present paper describes a development of the polarographic method mainly for the assay of streptomycin fermenter broth and associated recovery stages. EXPERIMENTAL AND RESULTS Assay of Solutions of Pure Streptomycin Sulphate (International Standardof 780 urrits/mg.)The work described was carried out using a Tinsley Mark 19 pen recording polarograph, with mercury capillaries of drop times between 2 and 3 sec. at a 50cm. height. As an alternative to tetramethylammonium hydroxide used by Levy and his co-workers (1946) the experiments described below used lithium hydroxide as base electrolyte (Wise, unpublished). Its advantage over sodium hydroxide (Tsukamoto and Tachi, 1952) was that it had a more negative half-wave potential.Initial experiments with pure streptomycin showed that polarograms from solutions with concentrations below 200 units/ml. were difficult to measure. This difficulty was caused by the poor resolution of the slopes due to the residual, limiting and diffusion currents, and also, in less pure solutions, by additional reduction waves. Use of the derivative circuit reduced the overall sensitivity but improved the resolution so that it became possible to measure polarograms from solutions as dilute as 10 units/ml. The streptomycin response moreover was rectilinear and proportional up to at least 300 units/ml. Variation of the lithium hydroxide concentration over the range 0.025 to 0 . 5~ (pH 12.4 to 13.3) had little effect on the diffusion current of the main peak. The concentration of 0 . 0 5~ was finally chosen as most suitable for the assay of fermenter broth. T

A new paper chromatographic method for streptomycin and similar compounds is described. It is used to detect the products of reaction between streptomycin and ammonia in low purity streptomycin; the presence of such compounds causes the microbiological and maltol methods of streptomycin assay to give different results.