244 PYRIDOXINE AND RELATED COMPOUNDS 



metric method has been adapted to pure solutions, but as yet has not been 

 used successfully with biological materials. 



Vacher and Faucquembergue'^ utilized the change in the shape of the 

 ultraviolet absorption curve of pyridoxine with pH as the basis for their 

 assay. They analyzed for pyridoxine in the presence of other substances by 

 obtaining ultraviolet absorption curves at two or more levels of pH. 



2. POLAROGRAPHIC 



Pyridoxine produces a double w^ave when it is reduced at the dropping 

 mercury electrode, indicating a reduction in two 2-electron steps.^ Although 

 reduction was difficult, satisfactory results were obtained when tetramethyl- 

 ammonium bromide was the supporting electrolyte. This method, how- 

 ever, has not yet been used for the estimation of pyridoxine in foods. 



B. CHEMICAL METHODS 

 HENRY SHERMAN 



Most of the colorimetric tests that have been used for the assay of the 

 vitamin Be group were originally devised when pyridoxine was believed to 

 be the only member of this biologically active complex. Since pyridoxal 

 and pyridoxamine are now considered important members of the vitamin 

 Be complex and since they are present in natural materials along Avith other 

 biologically inactive phenolic compounds which often react similarly, many 

 of the original concepts concerning the various colorimetric tests have had 

 to be revised. It seems unlikely that any simple colorimetric test can be ap- 

 plied toward the assay of all forms of the vitamin Be complex. At present, 

 three separate and distinct compounds possess vitamin Be activity, and 

 there is some evidence that even these three compounds do not account for 

 all the biological activity.^ The colorimetric procedures which are now avail- 

 able are most accurate when applied to analytical samples, such as pharma- 

 ceutical preparations, which contain only pyridoxine. However, if these tests 

 are employed to assay natural products in which p3'ridoxine does not con- 

 stitute the major portion of the vitamin Be content, considerable caution 

 must be exercised in interpreting the results. 



One class of color reactions includes the formation of cj^anine dyestuffs 

 and phthaleins. 



1. Cyanine Dye Test^ 



The cyanine dye test is specific for cc-picoline compounds of which the 

 vitamin Be group is the only naturally occurring member. It has been ap- 



6 M. Vacher and D. Faucquembergue, Bull. soc. chim. biol. 31, 1419 (1949). 

 ^ J. J. Lingane and O. L. Davis, J. Biol. Chem. 137, 567 (1941). 

 8 R. Kuhn and I. Low, Ber. 72, 1453 (1939). 



