434 THIAMINE 



Krcbs also demonstrated that acetoacetate formation is somewhat re- 

 duced in livers of thiamine-deficient pigeons and that addition of the vita- 

 min restored this formation. 



Furthermore Smyth/"* as a collaborator of Krebs, prepared cultures of 

 Staphylococcus aureus or S. albus that were deficient in thiamine. The pyru- 

 vate metabolism of these microorganisms was much lower than that of 

 vitamin-saturated cells. Added thiamine greatly enhanced this pyruvate 

 metabolism. In this respect thiamine could be replaced by oxalacetate. This 

 effect could not be obtained in Staphylococci grown in vitamin-sufficient 

 media. So these experiments are in accordance with the assumptions that 

 thiamine catalyzes the formation of oxalacetate and that this substance 

 acts as a hydrogen carrier in the dismutation of pyruvate. At the end of 

 his article Krebs drew attention to the work of Ruben and Kamen/^ who 

 were able to demonstrate, with the aid of radioactive carbon, that animal 

 tissues were able to assimilate carbon dioxide. In the following years several 

 investigators firmly established the fact of the assimilation of carbon diox- 

 ide by animal tissues (e.g., Evans and Slotin,^^ Solomon et al.,^^ and Utter 

 and Wood^^). The role of thiamine pyrophosphate in this process is not 

 definitely proved, however, and there are several experimental data which 

 cannot be brought into line with this unitarian view of Krebs. Barron ct 

 al.,^"^ in contradiction to the results of Smyth with bacteria, stated that the 

 condensation reactions of pyruvate in tissues of avitaminotic animals were 

 accelerated by thiamine but not by oxalacetate. The results of Green with 

 the purified carboxylase preparations are not easily reconciled with the 

 Krebs' theory. 



2. Coupling of Pyruvic Acid Oxidation with Phosphorylation 



The coupling of oxidations with phosphorylations with high-energy and 

 low-energy phosphate bonds cuts the flow of energy of the oxidation into 

 fractions (Lipmann^^). In this way it might bethought that the difference 

 between oxidizing and non-oxidizing enzymes becomes less important. 

 Lipmann-*' ^^ studied the oxidation of pyruvate by an extract from Bacillus 

 delhriickii (Bad. acidificans longissimum). The pyruvic acid is oxidized to 

 acetic acid and carbon dioxide. 



CH3COCOOH + 10., -^ CH3COOH -f C().2 



The reaction requires inorganic phosphate and adenylic acid. The inorganic 



«3 S. Ruben and M. D. Kamen, Proc. Natl. Acad. Sci. U. S. 26, 418 (1940). 



6^ E. A. Evans, Jr. and L. Slotin, J. Biol. Chem. 141, 439 (1941). 



*^ A. K. Solomon, B. Vennesland, F. W. Klemporer, J. M. Buchanan, and A. B. Hastings, 



/. Biol. Chem. 140, 171 (1941). 

 66 M. F. Utter and H. G. Wood, /. Biol. Chem. 164, 455 (1946). 

 6^ F. Lipmann, Advances in Enzymol. 1, 99 (1941). 



