抄録
From our laboratory, a differential assay method of vitamin B_6 group was already reported, involving the separation of PIN (pyridoxine), PAL (pyridoxal) and PAM (pyridoxamine) using a cation exchange resin (Duolite-C 60), and microbiological assay procedure with Saccharomyces carlsbergensis. But this resin has not been commercially available, so the reexamination of new resin and a differential assay method of vitamin B_6 group containing the phosphoric esters in addition to free forms have been reported in this paper. The general method is as follows. First, the extracts of natural products are added on the anion exchange resin, Dowexlx8 (AcO^-), the phosphoric esters of vitamin B_6 are adsorbed on the resin, while free types are not. The latter is then added to the column of the cation exchange resin, Amberlite IRC CG-50 (H^+, 100〜200 mesh). PIN and PAL are eluted with acetate buffer, pH 4.6,while PAM is retained. The fraction containing PIN and PAL are divided in two parts, and PAL in one part is inactivated by acetone-alkali treatment. PAM adsorbed on column is eluted with acetic acid. Each content of PIN, PAM and sum of PIN and PAL are determined by the microbiological assay with Sacch. carlsbergensis. On the other hand, the phosphoric esters adsorbed on the Dowex-1 are eluted with acetic acid, hydrolyzed to the free forms and treated as described above. The expriments with authentic vitamin B_6 samples and some natural products (chick liver, baker's yeast etc.) under this method gave the satisfactory result. The effect of salts and limiting concentration of vitamin B_6 compounds in this assay are also showed.
