ESTIMATION 



were destroyed by oxidising agents such as nitric acid at ioo° C or 

 potassium permanganate or hydrogen peroxide at room temperature.^' ^ 



References to Section 6 



1. M. Hochberg, D. Melnick and B. L. Oser, /. Biol. Chem., 1944. 



155, 129. 



2. E. Cunningham and E. E. Snell, ibid., 1945, 158, 491. 



3. D. Melnick, M. Hochberg, H. W. Himes and B. L. Oser, ibid., 



1945. 160, I. 



4. M. Hochberg, D. Melnick, L. Siegel and B. L. Oser, ibid., 1943. 



148, 253. 



5. H. C. Epley, Amer. J. Pharm., 1945, 117, 265. 



7. ESTI2VIATION OF VITAMIN Bt 

 Biological Assay 



The biological estimation of pyridoxine by means of rats depends 

 largely on finding a diet complete in all the vitamins except vitamin 

 Bg ; and probably much of the earlier work was unsatisfactory because 

 the diets used were not completely free from traces of the vitamin. 

 Edgar et al.^ described a method of assaying " eluate factor " (pyri- 

 doxine) and " filtrate factor " (pantothenic acid) based on the growth 

 response of rats to graded doses of each of the factors, but they did 

 not claim that their method was completely satisfactory, whilst 

 R. C. Bender and G. C. Supplee ^ stated that the growth rate of rats 

 was not sufficiently specific for use in the estimation of vitamin Bg, 

 and used the onset of acrodynia as the basis of an assay method ; 

 they devised a basal diet that produced acrodynia in 100 % of their 

 animals in six to eight weeks. They reported that in order to obtain 

 optimal growth with vitamin Bg, another factor, " factor II " (pre- 

 sumably pantothenic acid), had to be added to the diet, thus con- 

 firming the work of Edgar et al. 



T. W. Conger and C. A. Elvehjem ^ used a synthetic diet consisting 

 of sucrose and casein, supplemented by aneurine, riboflavine, nico- 

 tinic acid, choline and pantothenic acid with a fuller's earth filtrate 

 from a butanol extract of liver to supply other members of the vitamin 

 B complex. They claimed to obtain satisfactory results when the 

 growth of rats was used as the criterion of response. Satisfactory 

 results were also reported by M. F. Clarke and M. Lechycka,* using a 

 similar method ; the dose-response curve obtained by plotting the 

 logarithm of the dose against the gain in weight was linear with 

 amounts of pyridoxine ranging from i to i-8 /^g. Even the best 

 biological method, however, takes at least a month to carry out.^ 



309 



