VII. ESTIMATION 455 



to 4-wcck period. The rates of gain were compared with that ot)tained on 

 measured doses of vitamin Bio.^* 



A procedure for the assay t)f vitamin B12 with rats was de.scribed hy 

 Frost and coworkers.'^ The diet had the following composition: 



g./lOOg. 



mg./lOO g. 



Weanling rats of both sexes were placed on the basal diet for a depletion 

 period of 7 to 14 days, following which six rats were used in each assay 

 group. The average weight gain during the 2-week assay period was used 

 for the assay of vitamin B12. The authors noted that mortality was high 

 on the basal diet during the assay period and that death was generally 

 preceded by the appearance of bloody encrustations about the nose and 

 paws. Vitamin B12 prevented these signs. 



D. MICROBIOLOGICAL 



Shorb reported as early as January 1947 that Lactobacillus lactis Dorner 

 (8000) required an unidentified factor which was named the LLD factor^^ 

 and which was present in refined liver extracts. Later Shorb demonstrated^^ 

 that the LLD activity of refined liver extracts closely paralleled the U.S.P. 



'5 U. J. Lewis, U. D. Register, and C. A. Elvehjem, Proc. Soc. Exptl. Biol. Med. 



71, 509 (1949). 

 '« D. V. Frost, H. H. Fricke, and H. C. Spruth, Proc. Soc. Expll. Biol. Med. 72, 102 



(1949). 

 "M. S. Shorb, J. Bacteriol. 53, 669 (1947). 

 "M. S. Shorb, J. Biol. Chem. 169, 455 (1947). 



