514 VITAMIN Bi2 



Bi2. It probably escaped major attention in nutritional studies with rats 

 for many years because of its pronounced tendency to be carried over in 

 the offspring from the well-supplemented diets which were fed to the mother 

 rats and also because of the apparent tendency of the vitamin to adhere 

 to casein which was thought to be vitamin-free. Both Coward and Gary 

 were concerned with obtaining diets which were as low as possible in vi- 

 tamin A. The approach to this problem involved extracting casein with 

 hot alcohol to remove the last traces of vitamin A. This also removed vi- 

 tamin Bi2, so that a deficiency of this factor was developed in the offspring 

 of mother rats which received the basal diet. 



A deficiency of an unknown growth factor was shown by Coward and 

 coworkers''' "^^ to occur in rats on purified diets containing 8 % of dried yeast. 

 The factor was present in some samples of casein but not in others and was 

 termed the "light white casein factor." The factor was removed from casein 

 by boiling with alcohol. Similar studies were reported by Mapson,^^ who 

 used the name "physin" for a growth factor needed by rats and occurring 

 in liver. Mapson found that female rats when fed liver transmitted the 

 effect of "physin" to their offspring which had not received the supplement. 

 The basal diet contained 10 % wheat germ and 8% yeast, which are good 

 sources of B-complex factors, except vitamin B12. In a subsequent article''^ 

 Mapson described the concentration of physin by extraction of a papain 

 digest of fresh beef liver with 90 % alcohol and 90 % acetone. The final solu- 

 tion contained 380 mg. of solids from 100 g. of fresh liver and showed ac- 

 tivity when fed in doses equivalent to 1 to 2 g. of original liver. Manganese 

 was ineffective. Mapson^"* obtained negative results with Lilly's extract 

 No. 343 and a liver extract ("B.D.H."), which presumably could be re- 

 garded as sources of the anti-pernicious anemia factor, and the trail was 

 lost. It was picked up again by Cary, Hartman, and their coworkers, who 

 carried ovit investigations during 1941 to 1949 which were described only 

 briefly and usually in the form of abstracts. In 1941, they reported^" that 

 a vitamin A-free diet, similar to that of Sherman, plus cod liver oil in ample 

 amounts was inadequate for optimum growth rate with weanling rats when 

 the casein was extracted with hot alcohol. Growth was increased when the 

 ration was supplemented with a liver extract. The deficiency could be 

 demonstrated with weanling rats whose mothers' diets were changed at 

 parturition to the basal diet. 



Further studies were described in 1946.'^ Rats at weaning ordinarily con- 

 tained a still unidentified factor (X) which alfected their growth and de- 

 velopment on a diet (A) adequate in all known mitrients. The young could 

 be depleted of X by feeding their mothers a diet deficient in X. Casein pre- 

 pared by centrifugation from milk was a good soiu'ce of X, and commercial 

 and various purified caseins contained different amounts of X; but, when 

 20 % or 40 % casein C, prepared by ten G-hour extractions with hot alcohol, 



