INTRODUCTION 



OX liver concentrate prepared by a method previously described by 

 W. B. Emery and L. F. J. Parker.* Better yields were obtained 

 from proteolysed liver extracts than from non-proteolysed extracts. 

 One pigment appeared to be produced from the other by proteolysis. 

 In the early stages of the fractionation, clinical tests with pernicious 

 anaemia patients in relapse were used to follow the course of purification, 

 but in the later stages the colour of the fractions was used for this 

 purpose. The most active preparation gave a response with a dose 

 containing only 0-3 mg. of total solids and, in addition to being anti- 

 anaemic, was effective in sub-acute combined degeneration of the cord. 

 The product was not pure and was not homogeneous when examined 

 in the Tiselius apparatus. It contained neither folic acid nor xantho- 

 pterine, and had a molecular weight of about 3000. 



Rickes et al} claimed to have isolated the anti-pernicious anaemia 

 factor in the pure state and gave it the name vitamin B12. They gave 

 no information about its properties or method of isolation beyond the 

 fact that the substance formed red needles which did not melt below 

 300° C. and contained cobalt. Vitamin B^g was showTi to be identical 

 with one of two unidentified growth factors required by Lactobacillus 

 lactis Dorner ; both of these were present in refined liver extracts.^ 

 Crystalline vitamin B^^g g^-ve a positive response in three cases of 

 Addisonian pernicious anaemia following single intramuscular injec- 

 tions of 3, 6 and 150 ^g. respectively.^ 



Subsequently, E. L. Smith ^ crystallised the anti-pernicious 

 anaemia factor from liver, and found it to contain 4 % of cobalt and, 

 assuming this to represent one atom per molecule, three atoms of 

 phosphorus per molecule. The phosphorus content of Smith's factor 

 apparently differentiates it from the vitamin B^g of other workers, for 

 both Ellis et al.^ and Brink et al.^ stated that the Co : P ratio was 

 I : I and not 1:3. This discrepancy may perhaps be explained by 

 the existence of several forms of vitamin B^g, all clinically active. A 

 second form, known as vitamin B^ga, was obtained by catalytic hydro- 

 genation of vitamin B^^ '^^ it was somewhat less active than the latter 

 and had a similar but not identical absorption spectrum. A third 

 form, vitamin B^2b> was isolated, together with vitamin B^g, from liver 

 and a culture of Streptomyces aureofaciens ; ^^ it likewise differed from 

 vitamin B^^ ^^ its absorption spectrum. Vitamin B^g has been isolated 

 from 5. griseus, which is now an important commercial source of this 

 factor. 12 



It has also been claimed ^^ that vitamin B^g exists in certain sub- 

 stances in the form of conjugates, inactive in pernicious anaemia until 

 they have been digested with hogs' stomach mucosa or with pan- 

 creatic enzyme extracts. These conjugates, it is suggested, may be 

 Castle's extrinsic factor (see page 498). 



531 



