190 CHEMICAL NATURE OF ANTIBIOTIC SUBSTANCES 



matogram slowly developed. The column was then washed with a solu- 

 tion of 1 5 parts acetone to 85 parts benzene until the yellow-orange band 

 approached the bottom of the column. The elution of the pigment from 

 the column was accomplished finally by further washing with 30 per 

 cent acetone in benzene until the eluate was faintly yellow in color. The 

 later eluates were found by assays to contain all the active pigment, 

 whereas all previous eluates, as well as the fractions remaining on the 

 adsorbent, showed no bacteriostatic or bactericidal activity. 



Pure actinomycin was obtained by concentrating the 30 per cent 

 acetone-benzene eluates to dryness, then recrystallizing the red solid 

 residue from acetone-ether mixtures or from ethyl acetate. From these 

 solvents, the pigment separated as vermilion-red platelets which 

 melted at 250° C, with slow decomposition. The pigment is readily 

 soluble in chloroform, benzene, and ethanolj moderately in acetone 

 and hot ethyl acetate j and slightly in water and ether. The color of the 

 solid pigment depends on its state of subdivision j when ground very 

 fine, its color is orange-red (910). 



Actinomycin is optically active, a solution of 5 mg. in 2 cc. ethanol 

 in a I dm. tube having a rotation — i.6o°j [a]'^ = —320° ± 5. Its 

 molecular weight was found to be around 1,000. Cryoscopic measure- 

 ments in cyclohexanol and in phenol gave molecular weights of 768 to 

 780 and 813, respectively. The approximate molecular formula was 

 found to be C41H56O11N8. Actinomycin exhibits characteristic ab- 

 sorption in the visible and ultraviolet regions. In ethyl alcohol, it shows 

 strong absorption at 450 (E| ^ = 200) and between 230 and 250. 



Actinomycin is not soluble in dilute aqueous alkali or in dilute min- 

 eral acids. It is soluble in 10 per cent hydrochloric acid and appears to 

 be regenerated by diluting such solutions with water. With strong alco- 

 holic alkali a purple color is formed, which rapidly disappears. Actino- 

 mycin is readily reduced by sodium hydrosulfite and by stannous 

 chloride, but is unaffected by sodium bisulfite. With sodium hydro- 

 sulfite the reduction is characterized by a change in color from red to 

 pale yellow. The color change is reversed by exposing the reduced pig- 

 ment to air. The same reversibility of color occurs when the pigment 

 is subjected to catalytic hydrogenation in the presence of platinum 

 oxide. The pigment has one or more functional groups capable of re- 



