GB1201727A - A new antibiotic - Google Patents

Abstract

1,201,727. The antibiotic tuberactin. TOYO JOZO K.K. 15 Aug., 1968 [17 Aug., 1967], No. 39032/68. Heading C2A. The new antibiotic tuberactin, having antibacterial activity especially against tuberculosis bacilli, has the following characteristics: (a) a white crystalline basic peptide substance, the hydrochloride salt having an elementary analysis of C = 33.95%, H = 5.66%, N = 22.69%, Cl = 12.40%, O (by difference) = 25.30%; (b) molecular weight = 473 (cryoscopically) or 486 (by titration); (c) empirical formula of the HCl salt: C 16 H 31 N 9 O 9 .2HCl; (d) melting point of the HCl salt: 244-264 degrees (decomposition) ; (e) optical rotation (HCl salt): [α] D <SP>25</SP>= -31À5 degrees (C = 1%, HcO); (f) U.V. absorp tion peaks: #max = 268 mÁ, E<SP>1% </SP> 1cm = 330 (in H 2 O), max= 268.5 5 mÁ, E<SP>1%</SP> 1cm = 313 (in O.IN.HCI), # max = 285.5 mÁ, E<SP>1%</SP> 1cm = 206-5 (in 0À1 N. NaOH), and having a U.V. absorption spectrum as shown in Fig. 1 above; (g) I.R. absorption peaks (HC1 salt) at wave numbers 3250, 1660, 1495, 1225, 1154 and 1045 cm.-<SP>1</SP> and an I.R. absorption spectrum as shown in Fig. 2 above; (h) the HCl salt is soluble in water, slightly soluble in MeOH, D.M.F. and 2-methoxy-ethanol, and insoluble or almost insoluble in ether, CHCl 3 , n-butanol, EtAc, dioxan, EtOH, pyridine, acetone and benzene; (i) a positive reaction to the ninhydrin, Sakaguchi and biuret tests, a negative reaction to the isatin, Pauli, Molisch and Elson-Morgan tests, and a yellow colour in the Erlich test; and has a basicity of pKa 1 = 7.2, pKa 2 = 10À3. Tuberactin is produced by aerobically cultivating Streptomyces griseoverticillatus var. tuberacticus, NRRL, 3482 or a tuberactin-producing mutant thereof in an aqueous nutrient medium of conventional composition. Crude tuberactin is isolated from the filtered medium by (a) precipitation with a sulphonic acid dye such as methyl orange, eriochrome violet or Alizarin Red S, and either suspending the tuberactin-dye salt in an organic non-solvent and adding an acid salt of triethylamine to precipitate the corresponding tuberactin-acid salt, or dissolving the dye salt in aqueous acid, extracting the dye with water-immiscible solvent, concentrating the solution and adding a miscible non- solvent to precipitate the tuberactin acidaddition salt; or (b) adsorption on a cation exchange resin and elution with dilute aqueous acid or alkali, followed by neutralization and concentration of the eluate from which tuberactin is precipitated with a non-solvent. Alternatively the filtered medium may be purified by dialysis and gel-filtration, concentrated and the tuberactin precipated with a non-solvent. The crude material is purified by re-crystallization from aqueous-alkanol, chromatography on a column of cation-exchange resin, silica-gel or cellulose, dissolution in water followed by precipitation with triethylamine sulphate and a water-miscible non-solvent, or thin layer chromatography on silica gel with 10% aqueous ammonium acetate/acetone/ 10% aqueous ammonia, 9 : 10 : 0.5 (v/v).