M. Cron, O. B. Fardig, D. Johnson
Sep 1, 1958
Citations
0
Influential Citations
24
Citations
Journal
Annals of the New York Academy of Sciences
Abstract
Kanamycin, discovered by Umezawa and co-workers,l is a water-soluble basic antibiotic active against Mycobateria and Gram-positive and Gramnegative organisms. I t is best isolated as the crystalline sulfate after adsorption on and elution from a carboxylic cation exchange resin. Characterization of the crystalline base, sulfate, N-acetylated derivative, and A' ,0-acetylated derivative,2 as well as Schiff base derivatives3 prepared from kanamycin base led to the formulation of kanamycin as C1~HasN4011. The crystalline sulfate, ClgH36N4011.H2S04, exists as the monohydrate from which water is removed with difficulty. The presence of four primary amine groups was indicated by Van Slyke nitrogen determinations and by the formation of a tetra-N-acetate and Schiff bases containing four moles of aromatic aldehyde per mole of kanamycin. Kanamycin sulfate is insoluble in the common alcohols and nonpolar solvents and is soluble in water to the extent of more than 25 per cent at pH 7. It is a very stable compound, exhibiting less than 10 per cent loss after autoclaving an aqueous solution for 1 hour a t 120" C. Kanamycin base is strongly dextrorotatory, [a]i4 + 146 (c 1, N/10 HSSOd), and gives no characteristic ultraviolet absorption spectrum. I t gives positive Molisch, ninhydrin, and Elson-Morgan tests and negative reducing sugar, Sakaguchi, and maltol tests. Treatment with 40 per cent sulfuric acid for 100 min. at 100" yields a product with an ultraviolet spectrum similar to that of furfural. Paper chromatography of kanamycin preparations revealed a second antibiotic, designated kanamycin B, which has been isolated and ~haracterized.~ The two antibiotics are best separated in Peterson's n-butanol-water-2 per cent p-toluenesulfonic acid system5 on Schleicher and Schuell (S & S) 589 Blue Ribbon or Whatman No. 1 papers. With S & S 589 Blue Ribbon Paper, kanamycin has an R f of about 0.35, and kanamycin B has an Rf of about 0.6. In this system the presence of impurities or contaminating salts interferes markedly with the paper chromatography of the kanamycins. The infrared spectra of kanamycin and kanamycin B are similar. Each is typical of a polyhydroxy, polyamino compound. No carbonyl or carbon-carbon double bond absorption is evident. By boiling for 15 min. in 4 N HCl, kanamycin was hydrolyzed to 3 major ninhydrin-positive materials that could be separated on Whatman 52 paper with n-butanol-acetic acid-water 4: 1 : 5 , giving Rf values of 0.02, 0.06, and 0.10. The 2 faster moving materials also gave positive reducing tests, suggesting the presence in kanamycin of 2 amino sugars glycosidically linked to a hydroxylated base. The material of Rr 0.02 was obtained in a pure crystalline state by 6 N HCl hydrolysis for 45 min. at loo", followed by concentration, precipitation with