62 THE BIOCHEMISTRY OF B VITAMINS 



extract, for example) which need to be added to the cultures are small 

 enough so that extraneous growth stimulants are not likely to interfere. 

 This is particularly true if the lower portion of the "growth curve" is 

 utilized. 144 Turbidity and color of extracts for the same reason usually 

 offer no problem whatever. This method has been used extensively in the 

 Cornell 145 and Texas laboratories and elsewhere. Hofmann 146 character- 

 ized it as "simple, accurate and fast," and as "the most satisfactory 

 technique." In view of the striking effect which amino acids were later 

 found to have on yeast growth, when the necessary nutrilites are sup- 

 plied, 147 Hertz improved the basal medium by the addition of a casein 

 digest and by the obvious substitution of calcium pantothenate for 

 ^-alanine. 148 



The yeast test is not applicable for the determination of biotin in the 

 presence of desthiobiotin, biotin diaminocarboxylic acid, desthiobiotin 

 diaminocarboxylic acid, 149 oxybiotin, 150 biotin sulfoxide methyl ester, 151 

 and a few less active compounds because they too possess biotin activity 

 for yeast. However, as these are synthetic substances and are not encoun- 

 tered in many types of investigations, the limitation on the method is 

 often not serious. Other microbiological procedures are limited to some 

 extent in the same way, but yeast possesses great synthetic powers and 

 can utilize some compounds in place of biotin which lactic acid bacteria, 

 for example, cannot. 152 



In applying the yeast test to metabolism studies, Oppel 153 found that 

 only a portion of the total biotin (as measured by the yeast test) of dog, 

 rabbit, rat and human urines was avidin-combinable, but that "only 

 minute amounts of the non-avidin-combining biotin were found in diets 

 and stool specimens." In the case of one subject who consumed 35-55 /xg 

 of biotin on alternate days but no raw egg white, the "non-combinable 

 biotin" of the urine was about 35-40 per cent of the total, whereas when 

 large amounts of egg white and biotin were both consumed, the "non- 

 combinable" constituted 75-85 per cent of the total urinary excretion. 



Burk and Winzler, 154 however, reported that uncombinable biotin is 

 abundant and widespread constituting 90 to 100 per cent of that in the 

 urine of rats and mice fed avidin and in Squibb urease, 30 to 50 per cent 

 in "Vitab" (rice bran concentrate) and unhydrolyzed beer, and 1 to 10 

 per cent in rat lung, spleen, testes, adrenals, lymph nodes, skin, intestinal 

 tract contents, feces, polished rice, dried yeast and food mixtures. Various 

 active substances possessing biotin activity were designated by these 

 investigators as "miotin," "tiotin" and "rhiotin." 



Chu and Williams 144 corroborated the findings of Oppel in that they 

 found substantial amounts of "uncombinable biotin" in urine but nowhere 

 else. They were unable to find evidence for the existence of "uncombinable 



