Biosynthesis of niacin from 3-hydroxyanthranilic acid (3-OHAA) in normal and streptozotocin diabetic rats was studied in vivo and in vitro. Streptozotocin (SZ) diabetic rats were found to excrete lesser quantities of 3-OHAA, quinolinic acid, niacin and N1-methyl nicotinamide (NMN) in their urines following 3-OHAA administration than corresponding normal rats. In vitro studies indicated that SZ diabetic livers form less quinolinic acid from 3-OHAA than normal livers. It appears that this may be due to elevated picolinic carboxylase activity in the livers of SZ diabetic rats.
An acute dose (10mg/100g body weight) of biotin at the post-implantation stage (day 14 and 15) inhibited the fetal and placental growth, and in few rats it also caused resorption of fetuses and placentae. The maintenance of pregnancy with normal fetal and placental growth was effected with estrogen therapy, but progesterone failed to correct the biotin-induced effect. The uterine and placental glycogen, RNA and protein levels, as well as, glucose-6-phosphate dehydrogenase activity in the ovary, liver and uterus showed a reduction following biotin treatment. Estrogen therapy under such conditions corrected these adverse effects of biotin overdose, while progesterone had no significant effects. The study suggests that the acute dose of biotin at an advance stage of pregnancy may cause adverse effects on the physiological regulation of gestation, pos sibly by creating deficiency of estrogen and gestagen. The possible role of estrogen in the fetal and placental growth and regulation of gestagen secretion is discussed.
A brush border preparation from rat intestine was incubated with rat intrinsic factor-vitamin B12 complex in 0.01 M Tris-HCl buffer, pH 7.4. The 57Co-B12 uptake to brush borders was proportional to the amount of protein or to alkaline phosphatase activity in the preparations. The uptake increased with time of incubation. At 37°C, the uptake after incubation for 15 min was 80-85% of that for one hr. The uptake at 4°C was approximately 70% of that at 37°C. There was no difference as a result of adding glucose to the incubation medium. The uptake was observed in the alkaline environment above pH 6.3. Maximum uptake occurred at pH 8.0. Brush borders washed with Krebs-Ringer bicarbonate buffer (pH 7.4) exhibited no difference in B12 uptake, whether in the presence or absence of calcium ion. But brush borders washed with ethylenediaminetetraacetate exhibited no uptake when incubated in calcium-free medium. The uptake reached a maximum by addition of calcium ion at a concentration of 0.3 mM, and was not alter up to 10 mM. Addition of magnesium ion exhibited no uptake. Calcium-dependent B12 uptake was markedly inhibited by manganese ion. Magnesium ion seemed to slightly inhibit the calcium-dependent uptake.
The effects of L-ascorbate 2-sulfate (AAS) on the lipid metabolism were examined in Triton-induced hyperlipemic mice, hyper cholesterolemic and normal rats, the following results being obtained. 1) In Triton-induced hyperlipemic mice, AAS (300mg/kg) significantly decreased the serum cholesterol level, while L-ascorbate (AA, 175mg/kg) was found ineffective. 2) In hypercholesterolemic rats fed 0.5 % cholester ol diet, the consecutive administration of AAS decreased the level of serum cholesterol and liver triacylglycerols. AA only slightly affected these levels. However, both AAS and AA prevented the unordinal increase in the liver weight caused by cholesterol feeding. 3) In normal rats, the administration of AAS over a 4-week period decreased the level of serum cholesterol and liver triacylglycerols.
Gas-liquid chromatographic (GLC) determination of vitamin D in multivitamin preparations containing excess amounts of vitamin E (a more than 2, 500 weight ratio of dl-α-tocopheryl acetate to vitamin D) was investigated and a simplified routine method was esta blished because the method reported previously (l) could not be applied to such special preparations. After applying the unsaponifiable matters of a sample to a phosphate-treated alumina column chromatography pre pared according to MULDER et al. (2), the eluate was evaporated and the residue was subjected to thin-layer chromatography (TLC) and GLC. When this method was applied to model preparations made by mixing vitamin D2 and dl-α-tocopheryl acetate (excess amounts), good results were obtained. Since the results on a commercial multivitamin prepara tion containing excess amounts of vitamin E were also satisfactory, it was confirmed that the proposed method could be used for simplified routine determinations.
Gas-liquid chromatographic (GLC) determination of vitamin D3 in tuna liver and vitamin D3 resin oils was investigated and a routine method slightly modified from the previously reported methods (1, 2) was established. Since both tuna liver and vitamin D3 resin oils contained large amounts of sterols, digitonin-Celite column chromato graphy according to SHEPPARD et al. (4) was used to remove the sterol influence from the unsaponifiable matters of the oils. After collecting the eluate and evaporating the solvent, the residue was subjected to thin-layer chromatography (TLC) using Kieselgel GF254 as an adsorbent and a mix ture of n-hexane-ethyl acetate (4:1) as a developing solvent. The scraped zones corresponding to vitamin D3 and pre-D3 were trimethylsilylated and then applied to the GLC using 1.5% OV-17 packed on Shimalite W (80 100 mesh) as a stationary phase. Trimethylsilylation of the gas chro matograms was an essential procedure, because the peaks of unknown substances in tuna liver oils and lumisterol in vitamin D3 resin oils could not be separated from the peak of pyro-D3 without trimethylsilylation. When the proposed method was applied to the samples, satisfactory results were obtained.
Standardized and reliable methods have been devised to assay preparations of the glutamic oxaloacetic transaminase from human erythrocytes by the CAS principle, i.e., the unsaturation and saturation of the coenzyme (pyridoxal 5'-phosphate) of a Coenzyme-Apoenzyme System. This dual enzymic assay is effective to detect and measure human deficiencies of vitamin B6. A preferred procedure has been developed, particularly for individuals and for limited groups, but is also applicable to large groups. A simplified procedure, which emphasizes the statistical significance of difference between large groups was also developed. Lyophilized erythrocyte hemolysates appear stable for up to 7 days for simplified transportation.
The dihydrofolate synthetase (EC 6. 3. 2. 12) responsible for catalyzing the synthesis of dihydrofolic acid from dihydropteroic acid and L-glutamic acid was purified about 130-fold from extracts of Serratia indica IFO 3759 by ammonium sulfate fractionation, DEAE-Sephadex column chromatography, Sephadex G-200 gel filtration, and DEAE-cellulose column chromatography. The enzyme preparation obtained was shown to be homogeneous by DEAE-cellulose column chromatography and ultracentrifugal analysis. The sedimentation coefficient of this enzyme was 3.9 S, and the molecular weight was determined to be about 47, 000 by Sephadex G-100. The optimum pH for the reaction was 9.0. The enzymatic reaction required dihydropteroate, γ-glutamate and ATP as substrates, and Mg2+ and K+ as cofactors. γ-L-GlutamylL-glutamic acid cannot replace L-glutamic acid as the substrate. Neither pteroic acid nor tetrahydropteroic acid can be used as the substrate. ATP was partially replaced by ITP or GTP. The enzyme reaction was inhibited by the addition of ADP, but not by AMP. One mole of dihydrofolate, l mole of ADP and 1 mole of orthophosphate were produced from each 1 mole of dihydropteroic acid, L-glutamic acid, and ATP by the following equation: 7, 8-Dihydropteroic acid+L-Glutamic acid+ATP Mg2+, K+→Dihydrofolic acid+ADP+Pi These results suggest that the systematic name for the dihydrofolate synthetase is 7, 8-dihydropteroate: L-glutamate ligase (ADP).
A cortisol-binding protein exists in the lens of cynomolgus monkeys as in other species. The cortisol-binding capacity decreased markedly with a diet either moderately or severely depleted of protein, even if the diet was later changed to a normal diet during infancy. This suggests that malnutrition may cause insufficient synthesis of the cortisol binding protein in the lens and that malnutrition is one of the important factors controlling the inactivation of steroid hormone in the lens.
n-Alkane assimilating yeast, Candida tropicalis YO-148, was grown on an n-alkane-containing medium. A synthetic diet containing 6.8% of dried yeast was fed to rats. The fatty acid composition of adipose tissue and liver fats was determined after a two week feeding period. The percentage of odd-numbered acids increased in the animals fed the yeast diet. Furthermore, it was shown that heptadecenoic acid, the major odd-numbered acid in yeast, was accumulated in neutral lipid fraction of adipose tissue fat. Fatty acid composition of protein isolate prepared from yeast cells had a profile similar to that of the original cells.