Two derivatives of pyridoxine monooctanoate were synthesized. α^4-O-Octanoyl pyridoxine (PIN-4MO) and α^5-O-octanoyl pyridoxine (PIN-5MO) were hydrolyzed in 10% aqueous solution of acetone at pH 1〜3 following an apparent first-order reaction. At the pH range of 1〜7,PIN-4MO was hydrolyzed 10 times as fast as PIN-5MO. At pH 4〜12,an interconversion between the two derivatives was found. At the pH range higher than 9,their hydrolysis was enhanced with an increase of pH and the total amount recovered as PIN and PIN-MO was 60〜80% for each derivative. The residue fraction was found to be water-soluble and reactive with imide reagent. The structure of this fraction was left unknown in the present study. The treatment of PIN-MO in 50% aqueous ethanol or methanol solution for 48 hours at pH 7 and 60℃ resulted in 4-ethoxy PIN and 4-methoxy PIN dimer.

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[in Japanese]

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Utilization of a chemically synthesized pyridoxine-β-glucoside by vitamin B_6-deficient rats was examined in terms of its effects on the urinary excretion of xanthurenic acid and on the activation of vitamin B_6-enzymes, glutamate-pyruvate transaminase and cysteine desulfhydrase. Oral administration of pyridoxine-β-glucoside (30 μg per animal per day) for 12 days has led to a complete restoration of the urinary excretion of xanthurenic acid to a normal level. The levels of the enzymic activities in liver and erythrocytes recovered with statistical significance (P<0.01) to those in the "positive" control rats administered with authentic pyridoxine. The "β-glucosidase" catalyzing hydrolysis of pyridoxine-β-glucoside was found in small intestine at a significant level, and to a somewhat lesser level, in liver and blood. It thus appears that pyridoxine-β-glucoside in vivo avails as vitamin B_6 by enzymic conversion to free pyridoxine either before or after absorption in small intestine. The post-absorptive conversion is supported by permeation of pyridoxine-β-glucoside into serosal side with everted sacs and by the high effectiveness as vitamin B_6 of intravenously injected pyridoxine-β-glucoside.

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Pyruvate dehydrogenase [EC 1.2.4.1] was isolated from the pyruvate dehydrogenase complex and its molecular weight was estimated to be about 150,000 by the sedimentation equilibrium methods. The enzyme was dissociated into two subunits (α and β), with estimated molecular weights of 41,000 (α) and 36,000 (β), respectively, by sodium dodecyl sulfatepolyacrylamide gel electrophoresis. The subunits were isolated by the phosphocellulose column chromatography and their chemical properties were examined. The subunit structure was assigned as α_2β_2. The enzyme contained no thiamine pyrophosphate (TPP), therefore its dehydrogenase activity was completely dependent on added TPP and partially dependent on added Mg^2+ or Ca^2+. The K_m value of pyruvate dehydrogenase for TPP was estimated to be 6.5×10^-5M in the presence of Mg^2+ or Ca^2+. The enzyme showed highly specific activity for both pyruvate and α-ketobutyrate, but it also showed some activity with α-ketovalerate, α-ketoisocaproate, and α-ketoisovalerate. The pyruvate dehydrogenase was strongly inhibited by sulfhydryl inhibitors ; and it contained 27 moles of 5,5'-dithiobis (2-nitrobenzoic acid)-reactive sulfhydryl group and a total of 36 moles of sulfhydryl group. The function of sulfhydryl group was examined and discussed. The reconstitution of the complex by the reassociation of pyruvate dehydrogenase and the yellow fraction is also demonstrated.

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