A brief review has been made of the studies carried out by the author on intestinal absorption of vitamin B_<12>. The absorption of vitamin B_<12> is of a dual mechanism ; intrinsic factor of Castle is involved in the absorption of physiological doses and absorption of supraphysiological doses is effected by concentration gradient not involving intrinsic factor. The former mechanism may consist of at least three steps of a) formation of intrinsic factor-B_<12> complex, b) mucosal adsorption of this complex, and c) transfer of vitamin B_<12> from the complex through the mucosl barrier. No experimental evidence is as yet available to verify that these steps actually occur in the absorption of food vitamin B_<12> in the gut. In addition to these intraluminal factors. vitamin B_<12> absorption is subjected to humoral regulation which acts through the mucosa, as in thyroid disorders and pregnancy. Intestinal flora is another factor to be considered. The significance of this elaborate absorption system is discussed from a nutritional and physiological points of view.

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Producing chronic liver damage by intraperitoneal injection of 0.8 g/kg/week of CCl_4 to mature rats of Wistar strain, we observed the influences of pyridoxal phosphate (PAL-P) (5 mg/kg/day), administered with the diet. Increase of neutral lipid in liver (fatty degeneration), rise of liver cholesterol ester ratio and decrease of iodine number of fatty acid in each lipid fraction in liver were observed in the control group. All these changes were found remarkably reduced in PAL-P treated group. Decrease of liver protein and fall of paraphenylene diamine oxidase (copper-protein enzyme) activity were also reduced in PAL-P treated group. Histologically, vacuolar degeneration of liver cells, proliferation of collagenous fibres and decreases of PAS-positive substances and pyroninophile substance (ribonucleic acid) were observed markedly improved in PAL-P treated group. All these effects are considered to be due to pyridoxal, because PAL-P is thought to be absorbed after the dephosphorylation in intestine.

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Administering orally 5 mg/kg/day of pyridoxal phosphate (PAL-P) to chronically CCl_4-intoxicated rats, we observed the increases of diamine oxidase (DAO) activities in liver and in brain and the improvement of decrease of ornithine carbamyl transferase activity in liver. Pal-P caused the increase of the activity of DAO in liver in vitro also. This effect is more remarkable in case of CCl_4-damaged liver than normal liver. Clinically, PAL-P is found to activate the electroencephalogram, that is, to decrease the slow wave, when injected intravenously to patients of liver diseases. From these facts, PAL-P is supposed to improve the retentions of ammonia and amines in liver damage and is expected to be effective upon hepatic encephalopathy.

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The efficiency of S-adenosylmethionine and methylthioadenosine as sulfur source in the biosynthesis of biotin from desthiobiotin by washed cells of Saccharomyces cerevisiae, which were deficient in biotin and sulfur, was investigated. In the liquid medium containing desthiobiotin, Saccharomyces cerevisiae grew enough in response to S-adenosylmethionine and methionine, whereas it did not grow in the case of methylthioadenosine. S-Adenosylmethionine and methionine were incorporated into the cells by the addition of glucose better than without glucose. But 5'-methylthioadenosine was not incorporated, irrespective of the presence or absence of glucose. S-Adenosylmethionine and methionine had the almost same efficiency as sulfur source in the biosynthesis of biotin.

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The procedure for the isolation of biotin-related compounds from yeast cells using avidin of egg white was investigated. Three μg of biotin was almost completely inactivated by an addition of 60ml of 10% egg white solution, 0.2M ammonium carbohate solution was found to be useful for separation of biotin-avidin complex by dialysis. When the biotin-avidin complex was heated at 100℃ for 30 to 60 minutes, biotin was completely released. The biotin-related compounds were completely extracted with 95% ethanol from the concentrate. Biosynthesis of biotin from desthiobiotin by washed cells of Saccharomyces cerevisiae, deficient in biotin and sulfur, was investigated. It was elucidated by bioautography that the biotin-related compounds in yeast cells was composed of biotin, biotin-d-sulfoxide, biocytin and biocytin sulfoxide.

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Sulfur source in the biosynthesis of biotin from desthiobiotin by washed cells of Saccharomyces cerevisiae, deficient in biotin and sulfur, was investigated. It was elucidated that the radioactive-S of methionine was incorporated into biotin. Biotin-^<35>S was concentrated from the reaction mixture containing 140μc of methionine -^<35>S by the procedure using avidin, described in the previous report. This biotin concentrate contained 0.26μc of ^<35>S. It was found by radioautography, bioautography and paper chromatoscanning that 70% of radioactivity in the biotin concentrate was located in biotin-ralated compounds. The radioactivity was found in biotin, biotin-d-sulfoxide, biotin-l-sulfoxide (?), biocytin and biocytin sulfoxide.

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The purified glutamic-oxoalacetic apotransaminase extracted from soluble fraction of pig heart was incubated with the phosphopyridoxal oxime. Then the enzyme was precipitated by adding ammonium sulfate to the solution. At higher concentration of the enzyme, it was found that from its ultraviolet absorption spectrum and its pyridoxal phosphate content the oxime combined with apoenzyme remained nearly intact even after the enzymic reaction. At lower concentration, where the reaction rate increased liniearly with an amount of the enzyme, the sodium borohydride was used as the means to see the state of the phosphopyridoxal oxime in the enzyme, and it was found that the oxime in the enyme was cleaved after the transformation with the substrates.

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Treatment of anhydrous formamide and pyridine solution of a mixture of thiamine-5-phosphate and adenosine-5-phosphate with dicyclohexylcarbodiimide at room temperature for one day gave linear thiamine adenosine diphosphate (I) as a main products. By a similar procedure the thiamine inosine diphosphate (II) was synthesized from thiamine-phosphate with inosine-5-phosphate. II was obtained also from I by deamination with nitrous acid. Purification was achieved in each by the rechromaography on anion- and week cathion exchange resins.

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5-Deoxy-derivatives of pyridoxal, pyridoxamine and pyridoxine were prepared by the following processes. Pyridoxamine was reduced catalytically to 5-deoxypyridoxamine, which was converted to oxime of deoxypyridoxal by MnO_2 and NH_2OH, leading to deoxypyridoxal by HCl and AgNO_2. Deoxypyridoxine was obtained by deaminization of deoxypyridoxamine.

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In the 15th paper of this series it was reported that conversion of hydroxocobalamin to DBCC in a concentration as high as 50μg/ml was achieved by cell suspension method using Propionibacterium shermanii. This paper deals with the improvements on the culture conditions and the constituents of the media to cultivate the bacteria from the economical stand point. Replacement of the half volume of the tap water with distillage of acetone butanol fermentation broth spared the yeast extract in the culture medium. Employment of the solidculture process shortened the incubation period of the liquid-culture process to 3 days.

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The present paper describes the thiamine metabolism after anesthesia. After the subcutaneous injection of 20 mg of thiamine, fluctuations of total and bound thiamine were examined in the blood and of urinary thiamine in 3 hours. Simultaneously pyruvic acid in the blood and liver were also determined. As for the ether anesthesia, a slight change was found in the young and middle ages with normal liver function. However, in some persons of old ages with latent abnormality of thiamine metabolism before the operation, free thiamine in blood and 3 hours urine were increased after the operation. As for the nitrous oxide anesthesia, no change of thiamine metabolism was detected after operation even in old age. As for the fluothane anesthesia, disturbance of thiamine utilization was detected under 1 to 2% concentration with 1l/min, oxygen, but it was improved by an increase of oxygen to 4l/min. and employment of the semi-closed circle method before operation. A parallel relation was seen between thamine metabolism and liver function by the BSP test. The BSP test and thiamine metabolism returned to normal about one week after the general anesthesia. But thiamine metabolism in patients with disturbed liver function was very complicated and disturbance remained for a long period in many cases. From the experimental results and referencial consideration, circulated blood amount in the liver should be a great factor of abnormal thiamine metabolism after the general anesthesia. In order to supply a sufficient oxygen, anoxia under anethesia must be avoided, and administration of thiamine diphosphate with ATP or some thiamine derivatives should be recommended.

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