280 



DESCRIPTIONS OF ANTIBIOTICS 



pH 7.4 and stirred with 4 per cent acidic clay. 

 Clay filtered off and eluted with methanol -acetone 

 (1:1). Eluate concentrated in vacuo. Extraction 

 of residue with ethyl acetate at pH 7.5 to 8.0. The 

 solvent layer extracted with acidic water and 

 back-extracted with ethyl acetate at an alkaline 

 reaction. The solvent extract is passed through an 

 alumina column and eluted with a mixture of 

 methanol and ethyl acetate (1:1). Concentration 

 of eluate in vacuo and extraction of residue with an 

 ethyl acetate-ether (1:10) mixture. Crystallization 

 (needles) occurs in a few hours at room tempera- 

 ture . 



Chemical and physical properties: Basic sub- 

 stance. Soluble in water, ethyl acetate, butyl 

 alcohol, and chloroform; slightly soluble in ether; 

 insoluble in petroleum ether, benzene, toluene, 

 and ligroin. Ultraviolet absorption maxima at 255 

 and 320 m/x in water. Melting point of free grisa- 

 mine is 165-170°C, and of its sulfate 175-180°C. 

 Negative ninhydrin, biuret, Fehling, Sakaguchi, 

 and FeCls tests. Weakly positive p-dimethylamine 

 aldehyde and sodium nitroprusside tests. C-jsNe- 

 HssOio : C = 55.62%; H = 6.33%; N = 11.58%. 



Biological activity: Active on M. tuberculosis 

 strains 607, BCG, and H-2 (human type), and M. 

 phlei; very slightly active on B. subtilis and Staph, 

 aureus. Not active on gram-negative bacteria, 

 yeasts, or fungi tested. 



Toxicity: 150 mg per kg intravenously is toler- 

 ated by mice; 160 mg per kg causes death within 

 1 week. 



Reference: 1. Sawazaki, T. et al. J. Antibiotics 

 (Japan) 8A: 39-41, 1955. 



Grisein 



Produced by: Streptomyces griseus strains (1-4, 

 12) and S. subtropicus (8). 



Synonyms: Antibiotic 3510 (3), antil)iotic A 1787 

 (12), albomycin (8, 14, 15). All are identical to, or 

 very closely related to grisein. 



Remarks: Zahner et al. (15) recently called 

 sideromycins antibiotics that show cross resistance 

 with grisein and are antagoni?;ed in their action 

 upon gram-po.sitive (but not gram-negative) bac- 

 teria by ferrioxamines. They included in this group 

 of antibiotics grisein, albomycin, antibiotic 1787, 

 LA 5352, LA 5937, ferrimycin, and antibiotic 22765; 

 the first three are active upon both gram -positive 

 and gram -negative bacteria and the last four only 

 upon gram-positive bacteria. Ferrioxamines, pro- 

 duced by streptomycetes, form a new group of iron- 

 containing metabolites, with growth-stimulating 

 properties for a number of microorganisms. Bickel 

 et al. (16) proposed to include them, together with 



some known substances, like ferrichrome, copro- 

 gen, and the terregens, which either contain or bind 

 iron, in a new class of growth factors, the sider- 

 amines. This biological property of the sideramines 

 is counteracted by iron-containing antibiotics 

 from streptomycetes, the sideromycins. 



Method of extraction: Adsorption from broth- 

 filtrate on activated carbon at pH 7.5 to 8.0. Elu- 

 tion with aqueous pyridine, a-picoline, or ethanol 

 (5, 6). Eluate evaporated in vacuo and methanol 

 added to precipitate impurities. Addition of ether 

 precipitates grisein. Precipitate leached with 

 methanol, then treated with silver oxide to remove 

 impurities. Distributed between water and phenol- 

 chloroform mixtures l)y altering the pH of the 

 aqueous phase and the phenol concentration. 

 Reprecipitated from aqueous phase on addition of 

 isopropyl alcohol or ether. Purification by chroma- 

 tography on silica gel from a solution in 34 per 

 cent phenol-chloroform saturated with pH 4.6, 0.1 

 M citrate buffer; the solution is diluted in half 

 with chloroform before chromatography. Elution 

 with 17 per cent phenol -chloroform (6, 7). 



Chemical and physical properties: Complex with 

 neutral or weakly acidic components (6, 11). Red 

 powder (1, 8). Insoluble in ether, chloroform, 

 absolute acetone and ethanol, and benzene. 

 Slightly soluble in acetone and 95 per cent ethanol. 

 Soluble in water and phenol (1, 6). Ultraviolet 

 absorption spectrum maxima at 265 m^u (-Biem 108) 

 and 420 m/x (^1^28.9) (6). Positive biuret test (8). 

 Negative Brady, Fehling, Tollen, Liebermann, 

 Sakaguchi, Alillon, Pauly, Molisch, Seliwanoff', 

 orcinol, phloroglucinol, and ninhydrin tests (6, 

 8). Mild deamination results in loss of antibac- 

 terial activity (8). Heat -stable (1). Moderately 

 stable to aqueous acid solution; inactivated by 

 methanolic HCl (6). C4oH6iNioO-2oSFe: N = 

 13.28%; amino N = 1.34% (6, 8) ; Fe = 4.50% 

 (11). Molecvdar weight, 1300 (8, 13) or 1090 (6). 

 Removal of iron causes a loss of color, the ultra- 

 violet peak at 420 ni/x, and reduction in biological 

 activity. These changes are reversed on re-addi- 

 tion of Fe to the molecule. Contains four compo- 

 nents, A, B, C, and D, with Rf values of 1.0, 0.75, 

 0.60, and 0.28 on paper chromatography (butanol- 

 acetic acid-water, 4:1:5). Component C breaks 

 down during purification with concomitant ap- 

 pearance of more A (10). Albomycin was first re- 

 ported to contain three active components and 

 two inactive ones which were degradation prod- 

 ucts of the first three (11). Electrophoretic studies, 

 however, showed the presence of one major com- 

 ponent making up 85 per cent of the complex, and 

 three other trace components (13). Grisein was 



