Patent GB-912339-A (GB)
Title: Process for the manufacture of the antibiotic pilosomycin, its components and salts thereof
IPC: C12P1/06
Filed: 1959-07-17
Abstract: An antibiotic pilosomycin (identical with antibiotic A9578 of Specification 893,880) or its components A and B are produced by the aerobic and preferably submerged aerobic cultivation of a pilosomycin-producing strain of Streptomyces galilaeus n.sp. especially NRRL 2722 in an aqueous medium containing assimilable carbon, nitrogen and inorganic salt and, if desired, a growth-promoting substance.  Sources of carbon are glucose, saccharose, lactose, mannitol, starch and glycerol.  Sources of nitrogen are amino acids, peptides and proteins, peptone, tryptone, meat extracts, distillers solubles, grain solubles, ammonium salts and nitrates.  Inorganic salts are chlorides, carbonates and sulphates of alkali metals, alkaline earth metals magnesium, iron, zinc and manganese.  Cultivation is at 18-40 DEG C. for 1 1/2 to 5 days.  Pilosomycin is isolated from the clarified broth which is separated from the mycelium at pH 3 to 5. Isolation and purification of pilosomycin is obtained by a combination of one or more of the following steps:-(A) adsorption on active carbon, activated earth or a resin such as a copolymer of m-phenylenediamine and formaldehyde and elution with aqueous acid e.g. acetic or a mixture of water and a water-miscible organic solvent e.g. aqueous pyridine or an aqueous mixture of such organic solvents.  (B) adsorption on a cation exchange resin e.g. "Amberlite 1RC50" (Registered Trade Mark) in p acid, sodium or mixed acid-sodium form, and elution with a dilute acid.  (C) Precipitation as a salt with an organic acid especially the picrate.  Other salts are produced by treating the picrate with a salt of an organic base e.g. triethylammonium sulphate or a dilute acid in an aqueous medium e.g. aqueous methanol.  (D) Addition of excess water-miscible solvent e.g. acetone or dioxane to an aqueous or alcoholic aqueous solution of a salt of the antibiotic.  (E) Extracting an aqueous solution of the antibiotic with a solution of phenol in chloroform, the pH of the aqueous solution and the phenol content of the chloroform solution being varied, the antibiotic being recovered from the organic phase. (F) Adsorption chromatography on carbon, alumina, magnesium silicates, silica gel, calcium sulphate and ion exchange resins. (G) Distribution chromatography on cellulose, starch, silica gel and "Celite" (Registered Trade Mark) especially using a mixture of butanol/glacial acetic acid/water (4:1:5).  (H) Countercurrent distribution between two immiscible solvent phases e.g. N/10 ammonium acetate buffer of pH 4.6 and 10% phenol in chloroform. Pilosomycin has two components A and B of Rf value 0.34 and 0.58 respectively in watersaturated butanol containing 2% p-toluene sulphonic acid.  Salts are formed with inorganic and organic acids e.g. hydrochloric, sulphuric, phosphoric, acetic, propionic, valeric, palmitic, oleic, succinic, citric, mandelic, pantotheric and glutamic acids.  The antibiotic is put up into pharmaceutical compositions (see Group VI).ALSO:A pharmaceutical preparation for enteral, parenteral or local administration comprises the antibiotic pilosomycin, its component A, its component B, or salt thereof with an organic or inorganic acid, and a carrier. Pilosomycin (Group IV (b)) is said to be identical with antibiotic A9578 of Specification 893,880.