Patent GB-764198-A (GB)
Title: The antibiotic vengicide and processes for the preparation of the antibiotics oxytetracycline and vengicide
Filed: 1954-10-21
Abstract: <PICT:0764198/IV(b)/1> <PICT:0764198/IV(b)/2> Oxytetracycline and an antifungal antibiotic vengicide are produced by cultivating Streptomyces vendargensis (deposited in Centraal Bureau voor Schimmel-Cultuur in Boorn, Holland), or a mutant thereof, in an aqueous nutrient medium containing a source of carbon and of nitrogen under aerobic conditions.  The temperature of fermentation is 20-35 DEG  C., preferably 26-28 DEG  C., the pH of the medium is 5 to 8 and preferably 6.5 to 7.3 and the duration of cultivation 72 to 96 hours. Specified sources of carbon are soluble and insoluble carbohydrates, e.g. glucose, saccharose, lactose, malt extract and starch; sugar alcohols, e.g. glycerol.  Specified nitrogen sources are hydrolysed or non-hydrolysed casein corn steep liquor, yeast, peptone, extract of meat, extract of fish, fish meal, soybean meal, peanut meal, distillers' solubles and inorganic nitrates, e.g. ammonium nitrate. Other additions to the culture medium are mineral salts, e.g. sodium chloride, potassium dihydrogen phosphate, magnesium sulphate and ferrous sulphate; trace metals and buffers, e.g. calcium carbonate. The antibiotics are isolated from the broth, clarified after adjusting pH to 2.0 to 2.5 by centrifugation and/or filtration, by (a) adsorption on activated carbon or a weakly-acid ion-exchanger, or (b) extraction with a butanol or an amyl alcohol at pH 8-11.  For adsorption on carbon the pH is 6.5 to 7.5 and elution is with an alcohol partially miscible with water, e.g. butanol at pH 1.5. Preferred is a mixture of butanol, water and phenol.  After removal of solvents, the residue is extracted with a partially water-miscible alcohol at pH 8 to 11 and the extract so produced with aqueous acid of pH 1.5 to 2.5.  Oxytetracycline is precipitated from the aqueous extract as a complex with a quaternary ammonium salt and a metal salt of a magnesium or an alkaline earth metal at a pH of 9. Vengicide is isolated from the filtrate by extraction with alcohol, e.g. butanol at pH 9, followed by aqueous extraction of the extract so produced and adjustment of the pH to 4.0 to 5.0. For adsorption on an ion-exchange resin, clarified broth of pH 2.5 is passed down a column of the phenolic resin "Duolite S30," the oxytetracycline is adsorbed and the vengicide remains in solution from which it is recovered by adsorption on carbon or solvent extraction.  For direct extraction of the antibiotics from the clarified broth, a butanol or an amyl alcohol is used at pH 9, the alcoholic extract being extracted with aqueous acid of pH 2 from which the tetracycline is precipitated as a quaternary ammonium complex, the vengicide being recovered from the filtrate. Vengicide is active against yeasts and fungi; it is a white crystalline amphoteric substance, slightly soluble in water and in organic solvents; melting point 241.5-243 DEG  C.; [a ]D20-51.6 DEG in 0.1N. hydrochloric acid; empirical formula C24H29O9N10 carbon 47.05 per cent, hydrogen 4.85 per cent, nitrogen 23.25 per cent and oxygen 24.85 per cent; has ultraviolet absorption spectrum maxima at 233.5 mm  and 273.5 mm  when dissolved in 0.05N hydrochloric acid; and in suspension in mineral oil and in hexachlorbutadiene exhibits characteristic absorption in the infra red at the following frequencies expressed in reciprocal centimeters: 2920, 2240, 1650, 1580, 1485, 1450, 1365, 1320, 1250, 1200, 1040, 985, 910 and 885. Specifications 684,417 and 718,032 are referred to.