202 PTEROYLGLUTAMIC ACID 



twice as much leucovorin as PGA is required to produce half-maximum 

 growth. 



It is e\4dent, then, that in the past when aS. faccalis or L. casei has been 

 employed as the assay organism to determine the pteroylglutamic acid 

 content of natural materials, these assays have given a figure representing 

 both PGA and CF. It should be possible to determine with Le. citrovorum 

 that proportion of the "free PGA" which exists as CF, since this organism 

 for all practical purposes does not respond to PGA. Before such differential 

 assays are performed, consideration should be given to the existence of 

 bound forms of PGA. There is now considerable evidence that the CF in 

 fresh liver, and presumably in most other tissues, exists as a conjugate 

 which behaves in all respects as a polyglutamate analogous to that of 

 pteroylglutamic acid. Thus Hill and Scott^^^ reported that a sample of dried 

 brewer's yeast which contained initially 1200 CF units per gram was found 

 to contain 15,300 CF units per gram after incubation overnight at 37° with 

 a hog kidney enzyme preparation. Moreover this hog kidney preparation 

 was found to have similar properties to the PGA conjugase known to be 

 present in hog kidney. Other sources of "CF conjugase" include chicken 

 pancreas and the enzymes normally present in certain tissues that are 

 active during autolysis. ^^^' ^^^ There is no evidence at present to indicate 

 whether "CF conjugase" is identical with PGA conjugase, although it seems 

 logical that it would be. 



Another interesting microbiological property of leucovorin is that, al- 

 though it appears to be only half as active as PGA for growth of S. faecalis 

 and L. casei (cf. Table X), when a folic acid antagonist such as 4-amino 

 PGA is incorporated in the medium, leucovorin is markedly more effective 

 than PGA in reversing growth inhibition.^^^ In the presence of an eciui- 

 molar amount of leucovorin or PGA about eight times as much 4-amino 

 PGA is required to reduce growth of S. faecalis to half-maximum in the 

 presence of leucovorin as is needed in the presence of PGA. Such data im- 

 ply that CF may be more closely related to the biologically active coen- 

 zyme than PGA itself. 



C. IN MAN 

 FRANK H. BETHELL 



The most conspicuous effect of PGA deficiency in man is macrocytic 

 anemia associated with megaloblastic erythropoiesis. Normal gro\\'th and 



"« C. H. Hill and M. L. Scott, Federation Proc. 10, 197 (1951). 



"7 M. E. Swendseid, F. II. Botholl, and W. W. Ackcrmaim, J. Biol. Chem. 190, 791 



(1951). 

 "8 L. S. Dietrich, W. J. Monson, H. Gwoh, and C. A. Elvehjem, /. Biol. Chem. 194, 



549 (1952). 



