When neuroblastoma N 18 cells, precultured in vitamin B_1-and B_6-deficient Dulbecco's modified Eagle's minimum essential medium (MEM), were cultured in Eagle's MEM deficient in the same vitamin, cell proliferation was depressed to 75〜78% of that observed with the regular Dulbecco's and Eagle's media. Under the same culture conditions, the addition of thiamine+PL+CN-B_<12> or thiamine tetrahydrofurfuryl disulfide (TTFD)+PLP+OH-B_<12> to the deficient Eagle's MEM restored the impaired cell proliferation. The restorative effect of the latter three was more remarkable than the former three. Among the constituents of Vitaneurin^[○!R], TTFD returned the growth to 99% of that of the normal cells, whereas PLP or OH-B_<12> alone showed no appreciable improvement. On the other hand, the combined addition of these three vitamins showed a restoration of 106%, exceeding that of the cells growing in the regular media.

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Neuroblastoma N 18,precultured in Dulbecco's modified Eagle's minimum essential medium (MEM), were cultured in Eagle's MEM and incorporation of either ^3H-[CH_3]-choline or ^3H-[2]-myoinositol was studied on 48-hour cultured cells. Of the cellular radioactivity, about 72% and 22% were identified as phosphatidyl choline and phosphoryl choline, respectively, while about 79% was recovered in phosphatidyl inositol. The incorporation of choline into phosphatidyl choline was depressed when the cells, precultured in vitamins B_1-and B_6-deficient Dulbecco's MEM, were cultured in the same deficient Eagle's MEM. Addition of thiamine+PL+CN-B_<12> or thiamine tetrahydrofurfuryl disulfide (TTFD)+PLP+OH-B_<12> to the deficient Eagle's MEM restored the impaired incorporation. The restorative effect of the latter three was more remarkable than that of the former three. Of the latter three, TTFD was most effective, while PLP or OH-B_<12>alone was less effective. A combination of the three vitamins showed an incorporation which was comparable to a simple summation of the respective effects. The stimulating action of TTFD, PLP and OH-B_<12> was more effective when the incorporation was studied on synchronized cells by using a pulse-label method. In contrast to ^3H-[CH_3]-choline, the cellular incorporation of ^3H-[2]-myoinositol was not significantly affected by the addition or omission of the vitamins.

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The effect of penicillamine (PeA) administration on the distribution of B_6 vitamers in mouse liver was investigated. Both DL-and D-PeA reduced markedly the contents of PLP and PMP. Effect of D-form was weaker than DL-form. The PeA administration induced an appearance of thiazolidine derivative of PL (PL-T) in response to the reduction of PLP and PMP, but the total amount of B_6 vitamers was significantly decreased. When PeA was injected along with PN, the treatment induced a marked increase of PL-T, thiazolidine derivative of PLP (PLP-T), and PNP. From the present results, it may be postulated that a depletion of PLP and PMP by the PeA treatment is attributed to the conversion from PLP to PLP-T in liver, and that the derivative is dephosphorylated to PL-T, followed by elimination of PL-T from the liver tissue.

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Effects of dietary vitamin B_6-deficiency on rat liver glycolytic and glyconeogenetic enzymes, and on fatty acid synthesizing enzymes were investigated. B_6-deficiency of rats was confirmed by the observations of growth retardation, increase in urinary excreted amounts of xanthurenate and microcytosis. In the liver of B_6-deficient rats compared to controls, the levels of activities of glycogen phosphorylase, pyruvate kinase and pyruvate dehydrogenation complex were significantly lowered, while the activities of hexokinase and fructosediphosphate aldolase were increased. The difference of glyceraldehyde-3-phosphate dehydrogenase activity was not significant. Citrate synthase and malate dehydrogenase activities were accelerated and aconitate hydratase activity was decelerated, and citrate was slightly accumulated in deficient rat liver. The activities of phosphoenolpyruvate carboxykinase, triosephosphate isomerase and glycerol-3-phosphate dehydrogenase in deficient group were higher than those in controls. There was no difference in the activity of glycerol kinase between the two groups of rats. The activities of acetyl-CoA carboxylase and fatty acid synthetase were significantly increased by B_6-deficiency. Urinary excreted amounts of acetoacetate in deficient rats were less than in controls.

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