The differences of the metabolism related to vitamin B6-dependent enzymes were investigated using germ-free and conventional rats. There was a significant difference in the body weight gain between vitamin B6-deficient germ-free and conventional rats after about 30 days of the experiment, and the body weight gain was much less in the deficient germ-free rats than in the deficient conventional ones. Urinary excretion of xanthurenic acid was higher in the deficient germ-free rats than in the deficient conventional ones after 18 days. There was a significant difference in the activities of kynurenine aminotransferase in mitochondrial fractions of germ-free rats, but not in mitochondrial fractions of conventional ones. The activities of aspartate and alanine aminotransferases, with or without pyridoxal phosphate, significantly decreased in the deficient germ-free rats, but not in the deficient conventional ones. These findings indicate that the degree of vitamin B6 deficiency was more severe in the deficient germ-free rats than in the deficient conventional ones, and also suggest that intestinal microflora may have some effects on vitamin B6-deficient conventional rats.
The influence of vitamin B6 deficiency on the levels of several water-soluble vitamins and on acetyl-coenzyme A carboxylase activity was investigated using of germ-free and conventional rats. Judging from the vitamin B6 levels in tissues and the percent of decrease, the degree of vitamin B6 deficiency was more severe in the tissues of deficient germ-free rats than in deficient conventional rats. Nicotinic acid, pantothenic acid and biotin levels per wet weight significantly decreased in the liver of vitamin B6-deficient germ-free rats, and nicotinic acid levels per wet weight significantly decreased in the liver of deficient conventional rats. In the kidney of vitamin B6-deficient germ-free rats, a significant decrease in riboflavin and biotin levels was observed, although there was no observable difference in riboflavin, nicotinic acid, biotin and pantothenic acid levels in the kidney of deficient conventional rats. From an enzymatic standpoint, acetyl-coenzyme A carboxylase activity was especially significantly decreased in both germ-free and conventional rats fed a vitamin B6-deficient diet, and the percent of decrease was more in germ-free rats than in conventional ones. These findings suggest that vitamin B6 deficiency had stronger effects on the levels of water-soluble vitamins in germ-free rats compared with conventional rats.
The transition of zinc and iron metabolism in vitamin B6 deficiency was investigated using germ-free and conventional rats. In contrast to previous reports, a decrease in zinc content was not observed in the liver, pancreas, kindney, spleen, lung or testes of vitamin B6-deficient conventional and germ-free rats, but we found an increase in zinc content in the kidney of conventional rats and in the liver and spleen of germ-free rats. Vitamin B6-deficient conventional and germ-free rats retained more iron in their tissues than the control animals did, except for the spleen of germ-free rats. The deposit of iron was more evident in vitamin B6-deficient germ-free rats than in vitamin B6-deficient conventional rats, and is possibly proportional to the degree of vitamin B6 deficiency. It is possible that the deposit of iron in the organs had some influence on metabolic disorders in vitamin B6-deficient rats.
The effects of d, l-alpha-tocopheryl nicotinate (EN) on model hypertension in rats were studied in comparison with d, l-alpha-tocopheryl acetate (EA). The progress of hypertension in young SHR during the 9th to 15th weeks after birth was markedly accelerated by replacing their drinking water with 1% saline. The highly-developed hypertension in old SHR (9 months of age) was further advanced by salt-loading. Oral administration of 20 or 100mg/kg of EN or 88mg/kg of EA, once a day, delayed the progress of hypertension in young SHR and reduced advanced hypertension in old SHR. An antihypertensive effect of tocopheryl esters was also found in DOCA-salt hypertensive rats. The treatment with EN or EA definitely reduced the incidence of pathological changes accompanying model hypertension such as suppressed weight gain, pulmonary edema, myocardial fibrosis, cerebral hemorrhage and protected the animals from death. In antihypertensive effect. EN was about 5 times more active than EA in molecular base, and the effects of EN protecting from pathological changes associated with model hypertension were more definite than those of EA. The treatment with EN or EA reduced water and sodium retention in the DOCA-salt hypertensive animals. This fact may suggest the implication of a mechanism through electrolyte metabolism in the antihypertensive action of these tocopheryl esters.
Transketolase, sedoheptulose-7-phosphate: D-glyceraldehyde-3-phosphate glycolaldehyde-transferase [EC 126.96.36.199], was extracted from pig liver and purified 96-fold by ammonium sulfate fractionation, followed by column chromatography using DEAE-cellulose and a Sephadex G-200. Transketolase from pig liver was stable at pH 6.0 and above, whereas it was unstable at lower pH values. It could be resolved into apoenzyme and thiamine pyrophosphate in an acidic medium, in contrast to baker's or brewer's yeast transketolase which resolved in an alkaline solution. All the activity of pig liver transketolase was lost upon incubation at pH 5.0 for two hours even at 0°C but about 40% of the original activity could be restored by the addition of excess thiamine pyrophosphate and CaCl2. Restoration of the activity was achieved effectively at pH 7.6-8.0.
The present study was undertaken to determine the nutritive value of L-, DL- and D-tryptophan in chicks. Day-old chicks were fed commercial chick starter ration for one week and then they were given an experimental diet containing zein as a protein source for three weeks. The experimental results were analyzed by a slope ratio technique (weight gain vs. tryptophan intake). The relative biological utilization of DL- and D-tryptophan compared to L-tryptophan was approximately 55 and 15%, respectively. At the end of the experiment, the chicks were sacrificed and the concentration of free tryptophan in plasma was measured. The free tryptophan concentration in plasma corresponded closely with the increment level of tryptophan in diet when there was a normal level of dietary protein. But there was less correspondence when chicks were fed a diet low in protein.
In order to investigate the relationship between dietary amino acids and protein, and activities of intestinal sucrase [EC 188.8.131.52] and leucine aminopeptidase [EC 184.108.40.206, LAPase] in rats, the effect of supplementation of amino acids into a protein-free diet and a low casein diet containing sucrose as the carbohydrate source on these enzyme activities was studied. The segmental weights of the small intestine and its mucosa of rats fed the protein-free diet supplemented with L-methionine or with L-methionine and L-threonine at 0.1 or 0.2% levels were significantly higher than those of rats fed the protein-free diet or one supplemented with L-glutamic acid, but there was no difference in the segmental activities of the sucrase and LAPase among rats fed these diets. On the other hand, the supplementation of methionine or methionine plus threonine to the 5% or 10% casein diet produced remarkable increases in the segmental weights of the small intestine and its mucosa as well as in the segmental activities of the sucrase and LAPase. There was no difference between the segmental sucrase activity of rats fed the 10% casein diet supplemented with 0.2% methionine ad libitum and that of rats fed this diet under restricted feeding conditions, although the segmental LAPase activity was affected by the amount of food consumed.
Rats were fed on three kinds of diets for two weeks: (I) basal diet, (II) containing 0.1% cholate and (III) containing 0.1% cholesterol and 0.1% cholate. Each dietary group was further divided into subgroups to whose diet was added 0, 5 or 10% (dry weight) of minced oyster (Callocorchina) or clam (Tapes japonica). The serum and liver cholesterol levels of the rats fed the basal diet were reduced by feeding oyster or clam. The serum and liver triglyceride levels of all dietary groups were lowered markedly by feeding oyster or clam. The activities of glucose-6-phosphate dehydrogenase, malic enzyme and acetyl-CoA carboxylase were markedly reduced in the basal groups fed oyster or clam. These effects were observed in 5 and 10% shellfish feeding. These shellfish may be considered hypolipidemic foods.
The effects of voluntary exercise on the growth, glycogen of muscle and lipid contents of the liver and serum of mice fed different levels of dietary protein were investigated. In both the exercise and non-exercise groups, body weight gains were significantly greater in the 20% and 30% protein diet groups than in the 6% and 4% protein diet groups. After 6 weeks of age, it was shown that the amount of voluntary exercise by the 6% and 4% protein diet groups was greater than that by the 20% and 30% protein diet groups. As for hematological status, the raising of hemoglobin levels due to increasing dietary protein levels was further exaggerated by voluntary exercise. Hematocrit values rose with the increase in dietary protein levels. However the effect of exercise on hematocrit values was not clear. Liver glycogen levels, which were elevated with the increase in dietary protein levels, rose further due to exercise, though no changes were observed in muscle glycogen due to dietary protein levels and exercise. The lipid contents of the liver in all groups tended to be lower in exercise groups compared with non-exercise groups and it was observed that the high levels of dietary protein depressed the increase in liver lipids. Liver triglyceride levels of all groups fed the dietary protein levels except for the 20% casein diet group decreased due to voluntary exercise, and liver triglyceride levels were also lowered as dietary protein levels increased. The levels of serum triglyceride of all groups decreased due to voluntary exercise. This phenomenon was most remarkable in rats fed a 6% casein diet. The tendency for serum cholesterol levels to decrease due to increasing dietary levels of protein was further intensified by voluntary exercise. However it was not further influenced by voluntary exercise in the 4% casein diet group.
In liver homogenates of rats fed a low-level diet of Wheat-, Rice- or Miyazaki-pattern amino acid mixture, some enzymes such as glucose-6-phosphate dehydrogenase, ATP citrate lyase; fatty acid-synthesizing enzymes, malic enzyme and L-α-glycerophosphate dehy-drogenase, whose activities are indicators of lipogenesis have been determined from the viewpoint of the mechanisms producing fatty liver. In the early experimental period, malic enzyme activity increased more markedly in rats fed low amino acid mixture diets than in the control group, and L-α-glycerophosphate dehydrogenase activity in the liver increased slightly. Conversely, ATP citrate lyase and fatty acid-synthesizing enzyme activities remained almost at control levels, or glucose-6-phosphate dehydrogenase activity tended to decrease. These results suggest that some other associated factors, such as depression of the lipid transfer system in the liver rather than accelerated lipogenesis itself, may be the main cause of the fatty livers produced under these nutritional conditions.
An investigation was made of the effect of maternal protein and/or energy deficiency during pregnancy on developmental changes in the levels of catecholamines and serotonin in fetal brain. Pregnant rats were fed on a 20%, 6% or 0% casein diet from day 1 of pregnancy to the day of autopsy (day 18, 20 or 22 of pregnancy). In the control group, the catecholamine content of the brain increased during pregnancy, being 21ng on day 18, 48ng on day 20 and 52ng on day 22. A similar increase was found in the group on a 6% casein diet. In contrast, with complete protein deprivation there was no developmental increase in catecholamine. A slight increase in serotonin and a marked increase in 5-hydroxyindole acetic acid occurred during late pregnancy, irrespective of the maternal diet. At term, the levels of norepinephrine and dopamine and the tyrosine hydroxylase activity in the forebrain, cerebellum and brain stem of the fetuses in the group on a 0% casein diet were significantly less than those in the groups on 20% and 6% casein diets. The free tyrosine concentrations (μmol/g) in the brain of fetuses in the groups on 20%, 6% and 0% casein diets were 0.701, 0.213 and 0.661, respectively. From the above results it is concluded that the low catecholamine content of the brain in fetuses in the group on a 0% casein diet was due to disturbance of the system for catecholamine synthesis, rather than to deficiency of precursors.
A trypsin inhibitor was isolated and purified from the bran of rice, Oryza sativa, by extraction with 1% sodium chloride, heat treatment, ammonium sulfate precipitation, ion-exchange chromatog-raphy on a CM-Sephadex C-25 and gel filtration on a Sephadex G-75. The final preparation was homogeneous by electrophoretic analysis. Rice bran trypsin inhibitor (RBTI) had a molecular weight of about 14, 500 and an isoelectric point of 8.07. The amino acid composition was characterized by high contents of basic amino acids, aspartic acid, glutamic acid, proline and cystine. BRTI inhibited bovine trypsin at an inhibitor-enzyme molar ratio of 1:1.6. It displayed, however, nobility to inhibit α-chymotrypsin, pepsin, papain and subtilisin BPN'.