BUTYRIC ACID-BUTANOL FERMENTATIONS 35 



Johnson. 19 They found that when a cell paste of CI. 

 butylicum was kept frozen for a week or more and then 

 allowed to thaw, a soluble enzyme preparation was obtained 

 that catalyzed the conversion of pyruvate to acetate, carbon 

 dioxide, and hydrogen. Phosphate was required for pyru- 

 vate decomposition, and later 20 evidence was obtained for 

 the formation of acetyl phosphate (equation 6) . This pro- 



CH3COCOOH + HOPO3H2 — -> 



CH3COOPO3H2 + C0 2 + H 2 (6) 



vided an indication that an acetyl compound might be 

 a precursor of butyrate. Actually the extracts used by 

 Koepsell and Johnson were unable to form butyrate from 

 pyruvate under the conditions used. However, they did 

 catalyze a reaction by which acetyl phosphate and butyrate 

 were apparently converted to acetate and butyryl phosphate 

 (reaction 7), thus opening the possibility that the latter 



Acetyl phosphate + Butyrate — > 



Acetate + Butyryl phosphate (7) 



compound was involved in butyrate or butanol synthesis. 

 The origin of the hydrogen, which is a conspicuous product 

 of most butyric fermentations, was somewhat clarified by 

 this work. Extracts that produced hydrogen readily from 

 pyruvate (equation 6) were found to be unable to decom- 

 pose formate. This eliminated the theory that formate is 

 an obligatory intermediate in the formation of hydrogen 

 from pyruvate. 



The success of Koepsell and Johnson in obtaining enzy- 

 matically active extracts from CI. butylicum suggested that 

 similar methods might be used with other species. Conse- 

 quently Stadtman and I 21 tried preparing cell-free extracts 

 of CI. kluyveri by drying the cells and then extracting with 



