SUBSTANCES PRODUCED BY ACTINOMYCETES 195 



passes through the column less rapidly, since it is less soluble than the 

 chloride in methanol. 



The various streptomycin fractions obtained from the column are 

 concentrated and lyophilized, giving white amorphous powders. The 

 most active fractions range from 600 to 900 jjg/mg., and amount to 

 approximately 80 per cent of the total. Satisfactory results are obtained 

 only if the crude streptomycin has an activity of about 200 Mg/mg. or 

 higher. Preparations of lesser purity contain substances which interfere 

 with the development of the chromatogram. The chloride is soluble in 

 methanol, less soluble in ethanol, practically insoluble in butyl alcohol, 

 acetic acid, and pyridine. The sulfate is only slightly soluble in metha- 

 nol and practically insoluble in the other solvents. 



Streptomycin gives a positive Sakaguchi test, the presence of a guani- 

 dine group being indicated by the fact that alkaline hydrolysis results 

 in the formation of ammonia and the disappearance of the Sakaguchi 

 test. Streptomycin also gives a positive test for an hydroxyl group. 

 Negative tests are obtained in the amino nitrogen, Hopkins-Cole, Mil- 

 Ion, xanthoproteic, biuret, and Pauly diazo tests. The presence of a 

 carboxyl group is considered as questionable, since the streptomycin 

 chloride, obtained by precipitation from methanolic hydrogen chloride 

 with ether, gives approximately neutral solution. The ultraviolet spec- 

 trum of streptomycin showed only end-absorption below 230 my, 

 which makes improbable the presence of an aromatic ring or conjugated 

 double bonds. 



Streptomycin is inactivated rapidly by o.i N sodium hydroxide at 

 room temperature. It is relatively stable over a -pH range of i to 10 but 

 is inactivated by i N hydrochloric acid. 



Streptomycin was first crystallized as the reineckate salt from water, 

 in the form of thin plates which decomposed at 162°-! 64° C, the basic 

 component being (CioHi907_8N3)^. The antibiotic potency of pure 

 streptomycin lies between 800 and 910 Mg per mg. (299). Streptomycin 

 can also be isolated (537a) by the method described above for strep- 

 tothricin. This includes charcoal adsorption, elution with methanolic 

 formic acid, precipitation with picric acid, conversion to the hydro- 

 chloride, chromatography with aluminum oxide, and final conversion 

 to the crystalline helianthate. 



