VITAMINS

The authors proved that the green fluorescent substance found in the culture filtrate of Cl.acetobutylicum was identical with G-substance (8-N-ribityl-6,7-dimethyllumazine) (I) found in Er.ashbyii by Kuwada and Masuda. Using the enzyme preparations of several microorganisms, the authors succeeded to synthesize riboflavin enzymatically from this compound (I) in the presence of adequate 4C-donor and cofactors. It was also proved that (I) was synthesized enzymatically from 4-ribitylamino-5-aminouracil (II) and these two enzyme systems, (II)→(I) and (I) →riboflavin, were differed each other.

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It was found that FAD or FMN formed complexes equimolecularly with metal ions such as Ca^<++>, Mg^<++>, Ba^<++>, Cd^<++>, Mn^<++>, Co^<++> and Ni^<++>. The stability constants for these complexes were determined at pH 7.2 and 0.1 of ionic strength. It was shown that the stability constants for FMN complexes containing transition metals such as Mn^<++>, Co^<++>, Ni^<++> were greater than those for the other metal complexes. However, these relations were not observed in the stability constants for FAD complexes.

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A chromatographical study was made on possibility for existence of nucleotides in FAD preparations as the impurities which were hardly eliminated from the preparation. The results were as follows : cyclic-FMN and two other substances were detected as the impurities in FAD with 75% purity. It was found that the latter substances absorbed ultraviolet rays at 260mμ. This fact suggested that the substances were nucleotide relating compounds. However, a very slight amount of cyclic-FMN was only detected in more purified FAD preparations. It was previously reported that cyclic-FMN was produced from FAD during preservation. In this paper, it was, furthermore, confirmed that occurrence of the substance was observed also in the course of purification procedure of FAD.

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Determination of vitamin A in blood serum and serum protein of healthy human subjects showed that about 90% of the vitamin in serum was present in protein fractions. Fractional determination by salting-out with varying sodium sulfite concentrations revealed that about 70% of the vitamin in protein fractions occurred in globulin and 30% in albumin. Determination of vitamin A in serum, serum protein and serum globulin by salting-out of the serum taken 3 hours after orally administering 1 to 12 mg vitamin A palmitate to healthy adult subjects, when the serum vitamin A levels reached the maximum values, showed that the increase of the vitamin in total body was calculated to be about 11 to 50% of the amount administered, the percentage recoveries being the less, the more the vitamin was administered. Determination of vitamin A in serum protein fractions of normal healthy subjects by zone electrophosesis showed that about 1/4 of the total protein vitamin A was albumin vitamin A and 3/4 globulin vitamin A, of which α_1-vitamin A was greatest and α_2-vitamin A smallest. The ratios of vitamin A alcohol and vitamin A ester in each protein fraction were about the same. The serum protein vitamin A values in patients with kidney disturbances were about twice those of the normal subjects. The values of patients with liver disturbances were about the same as those of normal subjects. The ratios of the vitamin values of each fraction were somewhat different in kidney disturbances. Incubation of 3 ml serum with varying amounts (5 to 100 μg) of vitamin A palmitate resulted in the increase of the vitamin of each protein fraction by 1 to 8%, whereby the recoveries were the less, the more the amounts added. When vitamin A palmitate equivalent to 20 μg of vitamin A alcohol was added to 3 ml of dialyzed serum incubated for varying periods, and the vitamin contents in protein fractions were determined, the values rose gradually with time, reaching the maximum in two hours. The ratios of the vitamin values of each protein fraction varied somewhat, but it remained roughly constant after two hours. After adding vitamin A alcohol, vitamin A aldehyde, β-carotene, vitamin D_2,D_3,α-tocopherol, vitamin K_3,respectively, to dialyzed serum and incubated at 37℃ for varying periods, the vitamin values of the protein fractions became maximum after one hour. The ratios of the vitamin in each protein fraction varied somewhat according to the kinds of fat-soluble vitamins, whereby about 1 to 12% of the vitamin added was recovered. Following incubation of riboflavin, FMN, FAD, respectively, to dialyzed serum, about 10% of the vitamin added was detected in protein fraction. Each vitamin added to the serum became associated with each protein fraction without showing any special affinity to a certain protein fraction.

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