:-;5() 



DESCRIPTIONS OF ANTIBIOTICS 



separated into four fractions by paper chromatog- 

 raphy (3 per cent ammonium chloride-1 per cent 

 ascorbic acid). The complex can also be separated 

 into four components l)y countercurrent distribu- 

 tion (methanol-0.01 A' HCl-benzene-petroleimi 

 ether, 10:5:15:5) if 100 transfers are used: rifo- 

 mycin E = tubes 1 to 10; rifomycin D = tubes 40 to 

 50; rifomycin C = tubes 60 to 70; rifomycin A = 

 tubes 90 to 100. Rifomycin C and D: Brown amor- 

 phous substances. Soluble in methanol, acetone, 

 chloroform, and ethyl acetate. Slightly soluble in 

 neutral or acidic water. Soluble (2 per cent maxi- 

 mum) in buffer solutions at pH 9 to 10 in presence 

 of ascorbic acid to prevent rapid destruction. 

 Light-absorption maxima at 240, 305, 385, and 461 

 niju in methanol or ethyl acetate. The peak at 461 

 in^ shifts to 422 m^ in the presence of a reducing 

 agent. Positive Tollen. Fehling, and FeClg tests. 

 Negative ninhydrin test. Rifomycin C gives a 

 faintly positive ninhydrin test only after strong 

 acid hydrolysis. Rifomycin C: C' = 61.52%; H = 

 6.73%; N = 4.21%,. Rifomycin I): C = 62.17%; 

 H = 6.58%; N = 3.53%. Rifomycin E: Maximal 

 light at)sorption at 400 m/j. Rifomycin B: Brilliant 

 yellow prismatic needles. If a warm saturated 

 solution is slowly cooled, very long, thin needles 

 are formed. Decomposes at 160-164°C and does 

 not melt until 300°C. Solubility; water, 0.027%; 

 methanol, 2.62%; ethanol, 0.44%; acetone, 0.31%; 

 chloroform, 0.34%; benzene, 0.018%; ethyl ace- 

 tate, 0.19%; carbon tetrachloride, 0.0011%; ethyl 

 ether, 0.005%; petroleum ether, <0.005%. Dibasic 

 acid. Equivalent weight, 765. C = 61.75%; H = 

 6.72%; N = 1.88%; O = 29.22%. Proposed em- 

 pirical formula: C:s9HmNOi4 . Molecular weight 

 (Rast) 750. [a]5S9 = —11° (c = 1 per cent in 

 methanol). Light -absorption maxima in phos- 

 phate buffer (pH 7.3) at 223, 304, and 425 m^. 

 Infrared light-absorption spectrum given in refer- 

 ence 1. Crystals are very stable. Increased biologi- 

 cal activity of solutions at pH 4.0, resulting from 

 conversion of the antibiotic into a more potent 

 substance. Stability studies carried out by re- 

 covering the crystalline antibiotic from solutions 

 showed that at 37°C, after 160 hours at pH 4.0, 

 12 per cent of the antibiotic is left , and at pH 7.2, 

 79 per cent of the antibiotic remains undegraded 

 (1). 



Biological activity: Rifomycins D and C have 

 more biological activity than rifomycins E and A. 

 Rifomycin B: Active in vitro against gram-positive 

 bacteria, including mycobacteria ; inactive against 

 gram-negative bacteria and fungi. With Staph, 

 aureus, no cross-resistance observed between 

 rifomycin B and penicillin, erythromycin, novo- 



biocin, oleandomycin, streptomycin, chloram- 

 phenicol, and tetracycline. Serum does not affect 

 the antibiotic activity. Upon standing, solutions 

 show an increase in biological activity, because 

 of the formation of a derivative which is more 

 potent biologically than rifomycin B and which 

 can be separated by paper chromatography. Ac- 

 tive in experimental infections (mice) caused by 

 Streptococcus hemolyticus, D. pneumoniae, and 

 Staph, aureus (2). Very active in vivo against 

 freshly isolated strains of Staph, aureus (4). 



Toxicity: Rifomycin B: LD50 (mice) 2040 mg per 

 kg intravenously, and >3000 mg per kg intraperi- 

 toneally, subcutaneously, and orally. LD50 (rats) 

 1680 mg per kg intravenously, and >3000 mg per 

 kg intraperitoneal!}', sul)cutaneously, and orally. 

 LDoo (dogs) about 1200 mg per kg intravenously. 

 LD50 (guinea pigs) about 3000 mg per kg intra- 

 peritoneally. Further ])harmacological data in 

 reference 3. Not absorbed orally (4). 



Utilization: Rif(»nycin B: Active intramuscu- 

 larly against a numlier of infections in human 

 beings. 



References: 



1. Sensi, P. e/ o/. Antiltiotics Ann. 1959-1960, 



PI). 262-270. 



2. Timbal,M. T. Antil)iotics Ann. 1959-1960, 



pp. 271-276. 



3. Maffii, G. and Timbal, AI. T. Antil)iotics 



Ann. 1959-1960, pp. 277-284. 



4. Furesz, S. and Scotti, R. Antibiotics Ann. 



1959-1960, pp. 285-292. 



KiiiKK'idin 



Produced by: Strcptomyces ri)iiosus (1, 9). This 

 culture also produces oxytetracycline (9). 



Synonyin: Antibiotic PA 85 (9). 



Method of extraction: I. Broth-filtrate ex- 

 tracted with butanol at pH 9.0. Butanol extracted 

 with dilute mineral acid to remove oxytetracy- 

 cline. Butanol: (aj treated with active carbon, 

 washed with tlilute acids and alkali, then washed 

 with a i)e1r()l<Mun fraction (l).p. 30-50°C) to sepa- 

 rate out an aciueous layer containing rimocidin; 

 (b) concentrated in vacuo at 30-50°C, then treated 

 with petroleum ether. Purified by passage through 

 a column containing an IR-100-IR4B mixture. 

 Crystallized from an alcoholic solution containing 

 triethylamine sulfate ui)on addition of acetone, 

 then recrystallized from methanol-water (9). 

 II. Adsorbed from broth on activated carbon 

 and eluted with aqueous low molecular weight 

 alcohols or pyridine. Forced into the aciueous 

 layer on addition of hexane or ethyl ether (9). 



Chemical and physical properties: Amphoteric 



