COMBINED FORMS—EXTRACTION 31 



inhibition studies, found evidence that thiamine itself may form a rela- 

 tively undissociable linkage with protein, which possesses enzymatic 

 activity. Tatum and Bell/ 1 using the same inhibiting agent (pyrithi- 

 amine) on mutant strains of Neurospora, interpret their somewhat similar 

 findings to indicate that endogenous thiamine is more effectively utilized 

 than exogenous thiamine. In a subsequent study of the dissociation of 

 carboxylases in different yeasts and yeast preparations, Parve and Wes- 

 tenbrink 12 concluded that the various phenomena could be explained on 

 the supposition that cocarboxylase combines with proteins other than the 

 unaltered apoenzyme. 



There are a number of other suggestive findings with respect to the 

 extraction of thiamine from various sources ; the need for further research 

 is indicated. Of great importance in this connection is the fact that plants 

 often contain very little of their thiamine in the form of cocarboxylase, 

 and many unexplained irregularities occur. 13 - 14 Rice polishings yield free 

 thiamine without enzymatic treatment. Wheat does not contain cocar- 

 boxylase, although an enzyme is present which is capable of catalyzing 

 its hydrolysis. 15 Jackbeans and soybeans both contain relatively large 

 and comparable amounts of thiamine; jackbeans are rich in carboxylase, 16 

 but soybeans are said to have none. Under strongly acid conditions com- 

 mercial phosphatase preparations were found effective in releasing thia- 

 mine from wheat embryos, but were wholly ineffective in releasing the 

 vitamin from brewers' yeast. 17 In a wholly different experiment in a 

 different laboratory, one out of three samples of wheat germ yielded a 

 slightly increased amount of thiamine upon enzymatic treatment. 14 The 

 availability of the thiamine present in different yeasts varies widely, sug- 

 gesting differences in the method of combination. In the case of three 

 brewers' yeast, 93-100 per cent utilization by rats was reported, 18 whereas 

 in the case of some bakers' type yeasts the utilization by human subjects 

 is said to be as low as 17 per cent (see p. 291 ). 19 Utilization by rats in 

 these cases was also low. 20 



Unexplained irregularities have also been observed in connection with 

 animal tissues. In liver, brain and kidney a very large percentage of the 

 total thiamine is present as cocarboxylase, while in muscle the concentra- 

 tion of free thiamine may nearly equal to (or in some cases even exceed) 

 that of cocarboxylase. 21, 22 In milk, only 50-60 per cent of the thiamine 

 is in the free state, and the rest is combined in nondialyzable form, which 

 is not released by phosphatases but requires a proteolytic agent such as 

 papain. 23 - 24 Melnick and co-workers found that thiamine is apparently 

 destroyed 50-90 per cent when it is incubated with bile, but that subse- 

 quent treatment with a special enzyme from yeast caused a recovery of 

 a substantial part of the lost activity. The precursor of this recovered 



