Robert L. Peck

CHROMATOGRAPHY IN THE STREPTOMYCIN PROBLEM

Streptomycin has become increasingly important since its discovery was first reported, in 1944, by Schatz, Bugie, and Waksman.l The isolation of pure streptomycin and the application of streptomycin to clinical practice in less than three years after its discovery represent a significant achievement. To a considerable extent, this accomplishment is due to the development of chromatographic methods for purification of the active substance. The present paper is concerned with this phase of the streptomycin problem. The object of the reported work on the chromatographic purification of streptomycin was essentially the preparation of pure material for degradative and other studies, rather than the careful investigation of chromatographic behavior. I t will be understandable, therefore, that there remains much to be desired concerning knowledge of the chromatographic behavior of streptomycin. The problem was the separation of a complex active substance which was soluble in water and in methanol, but nearly insoluble in the common organic solvents, from large amounts of inactive impurities possessing very similar solubility properties. Chromatography appeared to be the only practicable approach, since neither purification by extraction from water into organic solvents and back into water, nor direct crystallization of the active substance was possible. The solubility of streptomycin concentrates was such that the solvent for chromatography was necessarily strongly polar (water, methanol, or mixtures largely composed of these with miscible solvents). Examination of adsorption behavior led to the general choice of alumina as the adsorbent for columns. Darco G-60 carbon was also found applicable. The colorless nature and similarity in properties of streptomycin and many of the impurities present in the concentrates made it difficult to follow the chromatographic purification of streptomycin on the columns by visual observation. Although colored salts of streptomycin have been made:) their chromatography does not appear to have been investigated to any great extent. The “liquid chromatogram”, discussed by Zechmeister and Cholnoky,4 has been employed, therefore, for the chromatographic work thus far described in detail.