VITAMINS

The binding of ^3H-d-α-tocopherol to isolated rat liver nuclei was investigated. 1) ^3H-d-α-Tocopherol rapidly bound to isolated nuclei and there seem to be specific binding sites for d-α-tocopherol in rat liver nuclei. 2) Nuclear membrane accounted for 90% of the total nuclear radioactivity found after the incubation of ^3H-d-α-tocopherol with rat liver nuclei. Subfractionation of nuclei revealed that nuclear matrix was the preferable binding sites of ^3H-d-α-tocopherol. 3) Dissociation of ^3H-d-α-tocopherol from nuclei by protease digestion suggested that α-tocopherol binds to proteins. 4) Subfractionation of chromatin further revealed that ^3H-d-α-tocopherol is associated with non-histone proteins having a high affinity for DNA.

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The fractional determination of ascorbic acid 2-sulfate (AsS), ascorbic acid (AsA) and dehydroascorbic acid (DAsA) was examined by hydrazine method. By using bromine as oxidizing agent of AsS on hydrazine method, the formation of DAsA and inorganic sulfate was confirmed quantitatively, but stabled to 2,6-dichlorophenolindophenol (DCIP). When each value by hydrazine method from tissues extract oxidized with DCIP and bromine were numerically almost equal, in these tissues, determination of added or intubated AsS in the tissues were following. Half volume of AsS solution extracted with metaphosphoric acid solution was oxidized with bromine at 37℃ for 10 min. Bromine was expelled by N_2 gas after reaction. The remainder extract was oxidized with DCIP at room temperature for 5 min. Both oxidized solutions were treated by hydrazine method and determined photometrically at 530 nm, respectively. The AsS levels in AsS treated tissues were obtained from following equation, AsS=bromine value (AsS+AsA+DAsA)-DCIP value (AsA+DAsA). The detective percent of additional AsS in tissues were 90〜100 in this method.

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The effect of dietary Ca and vitamin D_3 on the duodenal Cd transport was observed in the rat using everted gut sac technique in vitro. 1. Duodenal Cd transport was significantly increased in vitamin D deficient rat, however, it was not influenced by dietary Ca. The duodenal tissue binding and/or uptake of Cd from mucosal incubation medium was neither influenced by vitamin D_3 nor dietary Ca. 2. The transported Cd to serosal medium was very little compared to the Cd binding and/or uptake of duodenal tissue, suggesting two steps mechanism for intestinal Cd transport. The first step is the transfer from mucosal medium to duodenal mucosa and the second step is from duodenal mucosa to serosal medium. The second step was influenced by vitamin D. These results suggested that a significant increase in hepatic Cd accumulation of vitamin D deficient rat could be ascribed to the increase in the intestinal Cd absorption.

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Effect of nicotinic acid on the formation of catecholamine and the causes of decrease in catecholamine formation has been studied in the nicotinic acid-deficient rats. 1) Concentrations of nicotinic acid in whole brain and liver of rats fed the nicotinic acid-deficient diet were significantly lower than the animals fed the nicotinic acid-supplemented diet. And the levels of NAD and NADH in the brain were significantly lower in the nicotinic acid-deficient rats than those receiving the nicotinic acid-supplemented diet. 2) The activity of tyrosine hydroxylase in whole brain homogenate was increased in the nicotinic acid-deficient rats. No remarkable difference of the enzyme activity in soluble enyzme preparation was observed between the nicotinic acid-deficient and the nicotinic acid-supplemented group. In particulate fractions, however, the specific activity of the enzyme in the nicotinic acid-deficient group was higher than that of the nicotinic acid-supplemented group. When the particle-bound enzyme was solubilized with Triton X-100,no difference of the enzyme activity was observed between the two groups. There was no difference in the Km values for tyrosine between the enzymes purified by ammonium sulfate fractionation of the nicotinic acid-deficient and the nicotinic acid-supplemented groups. Therefore, the increased enzyme activity in the crude enzyme preparation from the nicotinic acid-deficient rat was not due to change of tyrosine hydroxylase, but appeared to be related to endogenous inhibitors. 3) No remarkable difference of the activity of dihydropteridine reductase was observed between the nicotinic acid deficient and the nicotinic acid supplemented group. However, as the concentration of pyridine nucleotide was decreased in the nicotinic acid-deficient rat, it was supposed that the pyridine nucleotide-mediated reduction of 6,7-dihydrobiopterin to the tetrahydro form might not catalyse well. 4) The concentrations of biopterin in the nicotinic acid-deficient rats were significantly lower than the animals fed the nicotinic acid-supplemented diet. The concentrations of biopterin in brain was very smaller than the Km values of tyrosine hydroxylase for biopterin. The studies reported here showed that the decreased formation of catecholamine of the nicotinic acid-deficient rat was due to the decreased concentration of biopterin in brain.

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