Patent GB-923612-A (GB)
Title: A new antibiotic actinomycin z and components thereof and a process for making these substances
IPC: G01N33/487
Filed: 1959-07-27
Abstract: <PICT:0923612/IV(a)/1> <PICT:0923612/IV(a)/2> <PICT:0923612/IV(a)/3> A new antibiotic designated actinomycin Z having 6 components, Actinomycin Z0, Z1, Z2, Z3, Z4 and Z5, is produced by cultivating an Actinomycin Z producing strain of Streptomyces fradiae especially C. fradiae NRRL 2765 under aerobic conditions in an aqueous nutrient medium containing one or more inorganic salts, a source of carbon and a source of nitrogen.  Specified inorganic salts are chlorides, nitrates, carbonates and sulphates of alkali metals, alkaline earth metals, Fe, Zn or Mn.  Specified sources of carbon are carbohydrates such as glucose, saccharose, lactose and starch.  Specified sources of nitrogen are amino acids and mixtures thereof, peptides, proteins and their hydrosylates such as peptone or tryptone, meat extracts, water soluble constituents, of cereal grains such as maize <PICT:0923612/IV(a)/4> and wheat, of distillation residues in the manufacture of alcohol, of yeast, of beans especially soya beans and of seeds of e.g. cotton plants.  Submerged aerobic cultivation at 18 DEG  to 40 DEG C. for 24 to 96 hours is preferred.  The antibiotic is isolated from the clarified broth (a) by solvent extraction with a water immiscible organic solvent, e.g. ethyl acetate, chloroform, butanol or diethyl ether, (b) absorption on active carbon, alumina, or activated earths, e.g. fuller's earth or floridin and elution with an at least partially water miscible solvent, e.g. acetone, butanol or methylethyl ketone.  Impurities are removed by extraction with first aqueous acid of pH <5 and then with aqueous alkali pH >8.  The actinomycin Z is precipitated as an amorphous red powder by the addition of a non solvent such as petroleum ether, pentane or hexane. The crude actinomycin Z is further purified using aluminium oxides as absorbent.  The 6 components can be isolated by (a) chromatography using aluminium oxide or cellulose, (b) distribution between two immiscible or partially miscible phases, (c) the counter current method using an aqueous solution of sodium naphthalene b  sulphonate and isopropylether as solvent system.  Actinomycin Z is active against a number of organisms including influenza virus.  On acid hydrolysis actinomycin Z and its components give N-methyl valine, valine N-methyl alanine, sarcosine and threonine and are orange to red in colour.  Actinomycin Z has a melting point at 260 DEG -264 DEG C., an optical rotation [a ]22D = -314 DEG  (c = 0.246 in chloroform), elementary analysis c = 54.88%, H = 6.42%, N = 12.25%, O (calc.) = 26.45%, U.V. maxima at 242, 429 and 443 mm  in ethanol (see Fig. 4) and an I.R. Spectrum as shown in Fig. 1. Actinomycin Zo decomposes at 250 DEG C., has U.V. maxima at 236 and 437 mm  and has Rf value 0.35 in the system di-n-butyl ether : n-butyl-acetate (1:3) on paper impregnated with 10% aqueous sodium metacresotinate, against Actinomycin C2 1.00. Actinmoycin Z1 has a melting point at 256-260 DEG C., optical rotation [a ]22D = - 362 DEG  (c = 0.185 in chloroform), elementary analysis C = 53.97%, H = 6.79%, N = 12.30%, O = 26.94%, CH3(N) = 7.48% and CH3O = 0%, U.V. maxima at 240, 427 and 442 mm  and an I.R. spectrum as shown in Fig. 2.  Actinomycin Z5 has a melting point at 261-267 DEG C., optical rotation [a ]22D = -284 DEG  (c = 0.244 in chloroform), elementary analysis C = 55.71%, H = 6.44%, N = 12.25%, U.V. maxima at 240, 428 and 443 mm  and an I.R. spectrum as shown in Fig. 3.  A mixture of Actinomycins Z2, Z3 and Z4 decomposes at 260 DEG C., has an optical rotation [a ]22D = -296 DEG (c = 0.257 in chloroform) and U.V. maxima at 240, 428 and 441 mm . Actinomycin Z and its components may be used in pharmaceutical preparations (see Group VI).ALSO:Pharmaceutical preparations for enteral, parenteral or local administration comprise the antibiotic Actinomycin Z and its components Actinomycin Z0, Z1, Z2, Z3, Z4 and Z5 or mixtures thereof (see Group IV (b)) in admixture with an organic or inorganic carrier.  Specified carriers are gelatine lactose, starch, magnesium stearate, talc, vegetable oils, benzyl alcohols gums polyalkylene glycols, white petroleum jelly, cholesterol or other medicaments.  The preparations may be in the form of a tablet, dragee, powder, salve, cream, suppository, solution, suspension or emulsion and may also contain preserving, stabilizing, wetting or emulsifying agents.