Patent GB-846801-A (GB)
Title: Antibiotic 6798rp and process for its preparation
Filed: 1958-05-27
Abstract: <PICT:0846801/IV(b)/1> A new antibiotic designated 6798 R.P. is produced by growing streptomyces peruviensis NRRL2757 Nov. Spec. in an aqueous nutrient medium containing assimiliable sources of carbon, nitrogen and mineral salts under aerobic and preferably submerged aerobic conditions.  The initial pH is 6,0 to 7,5, the temperature 25 to 35 DEG C. and the duration 3 to 4 days.  Specified sources of (a) carbon are glucose, saccharose, dextrose, lactose, molasses, mannitol and glycerol, (b) nitrogen are ammonium salts, e.g. chloride, acetate, lactate and tartrate, meat extracts, fish flours, soya bean meal, peanut flour, corn syrup, yeast extracts and casein hydrolysates, and (c) calcium or magnesium carbonate, alkali metal and alkaline earth metal phosphates, chlorides, sulphates, zinc, cobalt, copper, manganese and iron salts. Antibiotic 6798 R.P. is isolated from the clarified broth by (A) solvent extraction (B) adsorption.  In the case of (A) there is used a water-immiscible aliphatic alcohol of 4 to 5 carbon atoms, e.g. n-butyl-alcohol or an aromatic alcohol such as benzyl alcohol at pH 2 to 9 and preferably 3 to 4., preferably in the presence of an inorganic salt such as ammonium sulphate.  The antibiotic is recovered by (a) removing the solvent or (b) re-extracting the extract with (1) a base at pH 7 to 9, preferably in the presence of a poor solvent, e.g. benzene and recovering the so formed salt, or (2) a complex forming salt such as sodium salicylate.  In the case of (B) adsorption may be on charcoal, magnesium silicate earths or ion-exchange resins, e.g. Amberlite 1RC-50.  Elution is by aqueous methanol or ethanol containing hydrochloric acid or ammonia, or by an aqueous organic base such as aqueous pyridine.  Further purification is by means of (a) chromatography (b) solvent extraction (c) crystallisation of the antibiotic as a complex with an alkali metal salt of an aromatic organic acid such as salicylic, p-hydroxy benzoic, cresstinic, toluene-p-sulphonic, mandelic or sulphanilic acids.  The antibiotic base and the salt are mixed with water and the alkali metal base of the salt added to pH 7 to 9, the complex crystallises on adjusting the pH to 4,5 to 5,5 by addition of acetic acid.  Alkali metal salts are prepared by suspending the antibiotic complex in water, adding alkali metal hydroxide to pH 8,0, removing the organic acid of the complex by adsorption on an ion exchange resin, adjusting to pH 8,0 by means of an ion exchange resin and adding a non-solvent to precipitate the alkali metal salt.  Salts with organic bases such as methylamine, ethylamine and N-methyl glucamine are prepared in a similar manner.  The alkaline earth metal salts are prepared by double decomposition between the alkali metal salts of the antibiotic and alkaline earth metal salts.  Antibiotic 6798 R.P. is active against mycobacteria, especially streptomycin and neomycin resistant strains thereof, and certain Gram positive cocci and is of low toxicity.  It is a white amphoteric substance forming salts and complexes (see above); the base is weakly soluble in water, aqueous alcohols and dimethylformamide insoluble in acetone, benzene and chloroform; contains C=55,25 to 55,35%, H=5,9 to 6,1%, O=23,05 to 23,25% and N=15,60 to 15,65%, probably (C42H54N13N10)n; melts at 240-245 DEG C; [a ]20D= -21 DEG (C=1% in N10 sodium hydroxide); red colouration with cone.  HCl turning to green; positive reaction for sugar and phenolic hydroxyl; Rf values (Table III not shown); antibacterial spectrum (Table IV not shown); ultraviolet absorption max. 222-223 mm  E1%1cm=519, min. 237,5mm  E1%1cm=183, max. 257,5mm E1%1cm=235; infra red absorption spectrum (Fig. 1) at 3370, 1660, 1550, 1492, 1463, 1388, 1345, 1280, 1260, 1220, 1184, 1127, 1106, 1064, 1014, 1001, 961, 878, 812, 782, 749 and 699 cm.-1