A series of mutant strains of Lactobacillus fermenti resistant to the growthinhibitory properties of amprolium were isolated by serial transfers of the original strain in media which contained increasing concentrations of amprolium. Using three different resistance levels of strains (resistant strain A, B and C), the mechanism of amprolium resistance was investigated. Through the present studies, the relation between the degree of resistance and the thiamine uptake potential was elucidated.

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A hydroxymethylpyrimidine-requiring mutant of E. coli, 70-17, does not respond to mono- or pyrophosphate ester of the pyrimidine in the growth medium of Davis and Mingioli which contains excess amount of inorganic phosphate that represses the synthesis of alkaline phosphatase. A number of mutant strains which respond to mono- or pyrophosphate ester of the pyrimidine have been isolated from the strain, 70-17, following the mutagenesis of N-methyl-N′-nitro-N-nitrosoguanidine, although they require the phosphate esters for growth in much more amounts than the free form of the pyrimidine in the minimal medium at the short time incubation. One of these strains, 70-17-15, has been identified as a hvdroxymethyl-pyrimidine-requiring, alkaline phosphatase constitutive mutant whose synthetic ability for alkaline phosphatase has no longer been repressed by excess amount of inorganic phosphate present in the medium. Experimental evidence have been obtained that mono- or pyrophosphate ester of hydroxymethylpyrimidine is dephosphorylated by this enzyme, and resulting free form of hydroxymethylpyrimidine is utilized for the thiamine biosynthesis in this organism. The strain is useful for the bioautographic identification and quantitative assay, for mono- and pyrophosphates of hydroxymethylpyrimidine from biological materials.

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Vitamin D₃ action on uridine-5-³H incorporation into rat intestinal mucosal RNA was studied. The uridine was administered 1 hour after 2, 000 iu vitamin D₃ treatment. Specific activity of RNA in nuclear and supernatant fractions was increased by vitamin D₃ at 15 minutes after uridine-5-³H administration. Two hours after uridine-5-³H administration, specific activity of RNA in nuclei, mitochondria and supernatant were also increased, but it was decreased in microsome.
Mucosal RNA was further analyzed by methylated albumin-kieselguhr column. It was found that uridine-5-³H incorporation into sRNA was increased and that into rRNA was decreased with vitamin D₃ administration.
DNA dependent RNA polymerase activities were studied in isolated nuclei obtained from rat intestinal mucosal cell. Mg²⁺-activated DNA dependent RNA polymerise was not influenced at 1 hour after vitamin D₃ administration, however, decreased at 3 hours and 16 hours after vitamin D₃ treatment. These decreases of activity would be corresponded with decreases of uridine-5-³H incorporation into rRNA. Mn²⁺ and (NH₄)₂SO₄-activated DNA dependent RNA polymerase was not influenced by vitamin D₃.

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Vitamin D₃ binding protein has been identified in the cytoplasm and nuclear fractions of rat intestinal mucosa. Each protein binds in vitro directly with vitamin D₃-1, 2-³H. Cytoplasmic binding protein has little content of DNA, and nuclear binding protein contained DNA and a small amount of RNA. Its sedimentation constant was calculated as about 13S and 10S respectively by sucrose density gradient ultracentrifugation. Their different behaviours were also observed by the polyacrylamide gel disc electrophoresis. However, in the both cases the vitamin D₃ binding was not affected with equal amount of cholesterol, 7-dehydrocholesterol, 17-β-estradiol and testosterone. Vitamin D₃ transport from cytoplasm into nuclei was observed at a temperature of 37°, and not at the 2°. The vitamin D₃ binding protein would be participated in the intracellular translocation of vitamin D₃ of rat intestinal mucosa.

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To elucidate the biosynthetic pathway of thiamine in plants, the nature of the activation and condensation reactions of pyrimidine (2-methyl-4-amino-5-hydroxymethylpyrimidine) and thiazole (4-methyl-5, β-hydroxyethylthiazole) moieties were studied with cell-free extracts prepared from acetone powders of Japanese Radish and Rape. Investigations of the effects of preliminary incubation and of the early stage of the reactions using phosphorylated substrates suggested that both pyrimidine and thiazole had to be activated by ATP in the presence of Mg²⁺ before their condensation to form thiamine-P (thiamine monophosphate). The rate of thiamine-P synthesis in the presence of ATP and Mg²⁺ increased in the order of pyrimidine-PP>pyrimidine-P>pyrimidine in respect of pyrimidine moieties, and thiazole-P was more active substrate than thiazole. The nature of thiamine synthesized was determined as thiamine-P by the bioautographic method using a thiamine-less mutant of Lactobacillus fermenti. From these results it was concluded that thiamine-P in plants is synthesized through the pathway proposed in the cases of microorganisms.
It was found that thiaminephosphate pyrophosphorylase (EC 2.5.1.3) activity was inhibited by ATP, ADP, and EDTA, and that the inhibition was reversed almost completely by the addition of Mg²⁺ of about the equimolar concentration as each of them.

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De novo formation of N⁵-formyl-5, 6, 7, 8-tetrahydropteroyl-glutamic acid and rhizopterin, N¹⁰-formylpteroic acid, by Rhizopus oligosporus in a synthetic medium free of folate was observed by the method of bioautography. It was shown that both compounds converted to the expected acids by deformylation with alkaline treatment.
These folate compounds were also identified by comparison of their fluoro spectra and absorption spectra with those of authentic samples.

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The influence of vitamin B₁₂ and glucose cycloacetoacetate hydrolysate on oxidative phosphorylation in liver mitochondria of rats fed an atherogenic diet has been studied. Feeding of the atherogenic diet containing high content of cholesterol to animals caused characteristic alterations in the P/O ratios of liver mitochondria. With citrate and pyruvate as substrates, the P/O ratios of the liver mitochondria in the atherogenic diet-fed group are found to be much lower as compared to the normal group. Subcutaneous administration of vitamin B₁₂ or GCAh to atherogenic diet-fed rats resulted in an appreciable increase in the P/O ratios.

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The effect of feeding Katha (Acacia catechu) on niacin, riboflavin and ascorbic acid status in rats was investigated. The riboflavin content of blood and liver and its urinary excretion are not affected. Initially, the excretion and tissue content of ascorbic acid increases slightly, but eventually it returns to normal. The feeding of katha to rats is accompanied by a significant decrease in the niacin content of blood and liver as well as its excretion.

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Ethyldeoxythiamine having a CH₃CH₂-radical in place of CH₃ at C-2 position of pyrimidine moiety of deoxythiamine was synthesized by applying the method of ethylthiamine synthesis. The following chemical and physical properties were similar to those of deoxythiamine and thiamine: stability to heat, formation of thiochrome form with BrCN. The biological activities of this compound was first tested by two kinds of microorganisms which require thiamine, Lactobacillus fermenti and Kloeckera apiculata. Ethyldeoxythiamine showed thiamine antagonistic activity at the same level as deoxythiamine to both organisms. Inhibition indices of ethyldeoxythiamine for growth and for thiamine uptake were as follows: growth 280 (L. fermenti), 380 (Kl. apiculata); thiamine uptake 10 (L. fermenti), 50 (Kl. apiculata).

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1. Electron microscopic observations have shown that when dietary fatty liver is induced in rats, the intracellular structure of the liver parenchymal cells is considerably deformed due to depositions of fat droplets in the cytoplasm. Such changes are considerably recovered by treating rats for a week with myoinositol; the reduction of the size and the number of fat droplets is evident and the lamellar structure of the rough-surfaced endoplasmic reticulum becomes distinct.
2. Histological comparison of the liver sections before and after the perfusion of the isolated dietary fatty liver indicated that myoinositol, when added to the perfusate, accelerates the movement of fats from the liver to the artificial blood stream. This was confirmed by chemical analysis of the blood before and after the perfusion.
3. Similar acceleration of the removal of fats by this cyclitol was observed in the hepatic cellular level regardless of the presence of infiltrated fats in the cytoplasm.

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