468 THE BIOCHEMISTRY OF B VITAMINS 



analogue in the absence of an exogenous aspartic acid supplement; but the 

 inhibition index determined in the presence of both aspartic and oleic 

 acids varies from 1,000,000 to 10,000,000. Often when the inhibition index 

 is relatively low (1,000,000), cfs-aconitic acid and related tricarboxylic 

 acids increase the inhibition index several fold. However, the effect of 

 the tricarboxylic acids is not always obtained; consequently, even if the 

 tricarboxylic acids are essential products, presumably derived from 

 cc-ketoglutaric acid, additional enzymatic reactions involving biotin are 

 essential for growth of Lactobacillus arabinosus. The implications of 

 these data are indicated in Figure 5, which shows that the biotin 

 analogue presumably prevents the formation of a coenzyme which is 

 either directly or indirectly concerned with the carboxylation of pyruvic 

 acid to form aspartic acid, the biosynthesis of oleic acid or an analogous 

 product, and at least one additional product. 5, 20 



The carboxylation of pyruvic acid as a function of biotin was inde- 

 pendently and almost simultaneously discovered in three different 

 laboratories. 21, 20, 22, 2:i An interrelationship of aspartic acid and biotin 

 had been established for yeast much earlier, 24 and was extended to 

 bacteria. 25 The ability of oleic acid to replace the nutritional requirement 

 of some microorganisms for biotin had been previously reported. 26 



Homobiotin, which inhibits the utilization of biotin by Lactobacillus 

 casei at a relatively low inhibition index, does not inhibit the growth of 

 that organism in a medium containing oleic acid, 27 even at relatively 

 high concentrations. Since analogues of biotin do not usually inhibit the 

 utilization of biotin synthesized by an organism, it is possible that oleic 

 acid as a limiting product reduces the biotin requirement of Lactobacillus 

 casei to such a point that the organism is capable of synthesis of the small 

 amount of the vitamin needed for other enzyme systems. The possibility 

 of complete replacement of a vitamin by all the products of the enzyme 

 systems in which it functions must be considered in this case. In view 

 of the results with Lactobacillus arabinosus, it appears that synthesis 

 of the small biotin requirement by the organism may be the most logical 

 explanation. 



Desthiobiotin competitively prevents the toxicity of 2-oxo-4-imidazol- 

 idinecaproic acid for Escherichia coli. 28 The inhibition index is approxi- 

 mately 100. Since biotin, at slightly greater concentration than the lowest 

 giving any response, prevents the toxicity of even relatively high con- 

 centrations of the desthiobiotin analogue, it appears that the analogue 

 prevents the formation of biotin from a metabolite identical with or 

 similar to desthiobiotin. A supplement of either glutamic acid or a-keto- 

 glutaric acid prevents the toxicity of the inhibitor in such a manner as 

 to increase the inhibition index to 300. Either glutamic acid or cc-keto- 



