Photoisomerization of all-trans-retinoic acid and the geo-metrical isomers [9-cis-retinoic acid, 11-cis-retinoic acid, 13-cis-retinoic acid and 9, 13-di-cis-retinoic acid] in ethanol and their biological effects on F9 teratocarcinoma cells were analyzed. The rates of photoisomerization of the retinoic acids illuminated by fluorescent lamps (1, 200 lx) increased in inverse proportion to their concentrations. When the ethanolic solution of all-trans-retinoic acid (10-5 M) was kept under illuminated condition, the equilibrium mixture of the geometrical isomers of retinoic acid [all-trans-retinoic acid 25%, 9-cis-retinoic acid 10%, 11-cis-retinoic acid 10%, 13-cis-retinoic acid 30%, 9, 13-di-cis-retinoic acid 5% and uniden-tified compound 20%] formed at around 30 min. The apparent velocity of the photoisomerization was approximately 8×10-7 mol/L .min. Equi-librium mixtures with similar compositions were obtained by the photo-isomerization of other geometrical isomers. The geometrical isomers produced by the photoisomerization possessed significantly different bio-logical effects in the induction of differentiation of F9 cells into parietal endoderm-like cells: activities of 9-cis-retinoic acid (ED50, 8×10-7 M), 11-cis-retinoic acid (EDS0, 8×10-7 M), and 13-cis-retinoic acid (ED50, 8×10-7 M) were approximately 1/10 of all-trans-retinoic acid (ED50, 8×10-8 M), and activity of 9, 13-di-cis-retinoic acid (ED50, 1×10-5 M) was 1/100 of the level of all-trans-retinoic acid. Further, the retinoic acids acted with each other additively on F9 cells.
Cobalamin-dependent methionine synthase was purified from rat liver. The enzyme activity was separated into two peaks upon Mono-Q column chromatography. Peaks I and II of the enzyme, eluted in this order, were purified 18, 000- and 44, 000-fold in overall yields of 0.7 and 1.8%, respectively. Peak II methionine synthase, the major fraction, was homogeneous as judged by SDS-polyacrylamide gel electrophoresis. The enzyme was a large monomeric protein with an apparent molecular weight of 143, 000 Da. Interconversion of the enzyme between the two peaks was not observed during purification procedures. The enzyme required S adenosylmethionine and a reducing system for activity. Ap-parent Km values of the peak II enzyme for 5-methyltetrahydrofolate and homocysteine were 75 and 1.7μM, respectively.
In order to investigate the optimal fat content for total parenteral nutrition (TPN) solutions, male Wistar rats were subjected to 70% hepatectomy and then placed, for five days, on one of five TPN regimens in which fat represented 0%, 10%, 20%, 30% and 40%, respectively, of the total calorie content. As serum triglyceride levels in the fat-treated groups were lower than those in the non-treated normal rats, it was concluded that the administered fat was sufficiently hydro-lyzed. The greater the fat content, the higher the regeneration rate of the remnant liver. Significant differences were found between the 0%-fat group and 20%-plus fat groups. Hepatic triglyceride level was signifi-cantly lower in the 20%-fat group. Hepatic protein level was significantly elevated in all fat-treated groups. Serum phospholipids and total choles-terol due to the lecithin contained in fat emulsion were significantly elevated in the 30 and 40%-fat groups, indicating that fat content of 30 and 40% was excessive. The results suggest that TPN containing fat is superior to fat-free TPN for liver regeneration after partial hepatectomy, and that optimal fat content is estimated to be about 20% of total calorie content in the case of this fat emulsion.
We studied the effects of whey protein (WP) from cow's milk on calcium and bone metabolism in ovariectomized (OVX) rats. Six-week-old female Sprague-Dawley rats were ovariectomized and fed a low-calcium diet (0.03% Ca, 0.3% P) for 4 weeks. The OVX rats were divided into three groups and subjected to two experiments: Exp. 1, Cont group (20% casein, 0.3% Ca), WP (1%) group (19% casein, 1 % whey protein, 0.3% Ca) and Low-Ca group (20% casein, 0.03% Ca); and Exp. 2, Cont group (20% casein, 0.3% Ca), WP (1%) group (19% casein, 1 % whey protein, 0.3% Ca) and WP (2%) group (18% casein, 2% whey protein, 0.3% Ca). The rats were fed each experimental diet for 4 weeks. The final body weight, weight gain, food intake and food efficiency showed no significant difference between the Cont and WP (1%, 2%) groups in Exps. 1 and 2. There were no significant differences in the calcium balance, serum ALP or serum calcitonin levels between the Cont and WP groups in Exp. 1. But the breaking energies of the WP (1%, 2%) groups were higher than those of the Cont groups in Exps. 1 and 2. As for the amount of calcium, phosphorus and magnesium in the femur, there were no significant differences between the Cont and WP (1%, 2%) groups; however, the amounts of total amino acids in the femur of the WP (1%, 2%) groups were higher than those of the Cont groups in Exps. 1 and 2. The amounts of proline and hydroxyproline in the femur of the WP (1%, 2%) groups were also higher than those of the Cont groups in Exps. 1 and 2. These data indicate that the milk whey protein influence in OVX rats is an increase in bone proteins such as collagen and enhanced bone-breaking energy.
Stroke-prone spontaneously hypertensive rats (SHRSP) were fed a diet containing docosahexaenoic acid (DHA)-enriched Euglena glacilis (DHA-Euglena) as the protein source from 5 weeks of age. The effects on endothelial functions were investigated by perfusion experimentation using mesenteric vasculature, and compared with the effects of antihypertensive drugs. (1) At 13 weeks of age, SHRSP fed the DHA-Euglena diet showed an average blood pressure of 220 mmHg, which was 20 mmHg lower (p<0.05) than that in the control group, while SHRSP of the captopril-treated group (angiotensin I converting enzyme inhibitor: 200 mg/L drinking water) and hydralazine-treated group (vasodilator: 60 mg/L drinking water) showed marked hypotensive effects with blood pressures of 150-160 mmHg and 180-190 mmHg, respectively. (2) The constriction response to norepinephrine (NE) was lower (p<0.01) in the mesenteric vasculature isolated from the DHA-Euglena-treated SHRSP than in that from the control group. (3) When the mesenteric vasculature isolated from 13-week-old SHRSP fed the DHA-Euglena diet was perfused with an acetylcholine solution (10-6 M) in the presence of NE (8×10-6 M), the relaxation rate was 81%, which was higher (p<0.01) than that in the control group (61%). Among the antihypertensive-treated groups, the captopril-treated group gave nearly the same relaxation rate as the DHA-Euglena diet group, while the hydralazine-treated group indicated a slightly lower rate (65%). At 18 weeks of age, the endothelium-dependent relaxation of SHRSP in the control group was further reduced (28%), but in both the DHA-Euglena diet group and antihypertensive-treated groups, the relaxation rates were not substantially different from the levels at 13 weeks of age. Reduction of the endothelium function in SHRSP occurs due to aging and blood pressure elevation. However, by improving nutritional conditions by the feeding of a DHA-Euglena diet, the endothelial functions were protected without a fall in blood pressure unlike antihypertensive drugs. It is hence considered that nutritional improvement helps maintain a sound architecture for the vascular wall, thereby leading to the suppression and delay of onset of cerebrovascular diseases, and subsequently to the prolongation of life-span.
The effects of high-fat diets containing fish oil or lard on blood glucose and plasma insulin after oral glucose loading were com-pared in genetically diabetic (db/db) mice, one of the animal models of non-insulin-dependent diabetes mellitus (NIDDM) with hyperinsuline-mia. The blood glucose levels were significantly decreased in 27% of the mice fed high-fat diets containing 20% fish oil 30 and 60 min after the oral administration of glucose (both;p<0.05). Conversely, the plasma insulin levels were significantly increased 30 min after the glucose loading as compared to 27% of the mice fed high-fat diets containing 20% lard (p<0.01). In addition, a significant hypoglycemic effect was observed 60 min after the subcutaneous administration of insulin to mice on the fish oil diet (p<0.05), whereas no effect was demonstrated in the case of those on the lard diet. The average body weight of the fish oil-treated mice was not significantly different from that of the lard-treated mice. The fish oil diet has a beneficial effect on glucose tolerance by increasing the insulin secretory capacity from pancreatic β cells and also ameliorating insulin resistance.
The change of tryptophan-niacin metabolism in D-galactos-amine (D-galN) injected rats was investigated. Rats fed with niacin-free diets containing 40% casein for 11 days were injected with D-galN (0.8 g/kg body weight). The urinary excretions of nicotinamide and its metabolites, and the activity of liver α-amino-β-carboxymuconate-E-semialdehyde decarboxylase (ACMSD) (EC 188.8.131.52), a key enzyme of tryptophan-niacin metabolism, were assayed. As the result, the urinary excretions of N1-methylnicotinamide (MNA), N1-methyl-2-pyridone-5-carboxamide (2-Py), N1-methyl-4-pyridone-3-carboxamide (4-Py) and their sum (nicotinamide+MNA+2-Py+4-Py) were higher in the D-galN-injected group than in the control group. Hepatic ACMSD activity in the D-galN-injected group was lower than that of the control group. These results suggest that the increase in urinary excretion of nicotin-amide and its metabolites after the injection of D-galN is considered to be attributable to a decrease in liver ACMSD activity.
An anti-thrombin substance (M2) was isolated from a culture broth of Rhizopus javanicus. Accumulation of M2 reached a maximum peak after 14 to 15 days of incubation and then decreased. The yield of M2 was 500 mg from 1 L of culture broth. M2 inhibited thrombin activity, and its 50% inhibition concentration in a reaction mixture containing 50μL of 12.5 NIH unit/mL thrombin and 200μL of 0.33% bovine fibrinogen was 63μM. M2 had a specific activity for thrombin, but it was less responsive to plasmin, tissue-type plasminogen activator, urokinase, plasma kallikrein and glandular kallikrein. The structure of M2 was identified as fumaric acid by elementary analysis, FAB/MS, 1H-NMR, 13C-NMR and IR spectra.
Recently, a new potent antioxidant was isolated from Tempeh (a traditional fermented soybean food in Indonesia) and was identified as 3-hydroxyanthranilic acid (HAA). This study deals with the antioxidant mechanism of HAA under biological systems and the cytokill-ing function of HAA to human malignant cells. HAA eliminated free radicals and inhibited the formation of fatty acid hydroperoxide in vitro, suggesting that HAA would serve as an antioxidant in the initial reaction in lipid oxidation systems. Actually, HAA inhibited the formation of the dominant product of membrane lipids, 12-hydroxyeicosatetraenoic acid (12-HETE) at a high concentration, while HAA accelerated 12-HETE formation at a low concentration in mammalian tissue. HAA oxidized glutathione and inhibited superoxide dismutase in vitro. Furthermore, HAA inhibited cell growth and induced apoptosis to HuH-7, a human hepatoma-derived cell line. As long as HAA is taken as a component of Tempeh, and not in large doses as a chemical, it may possibly act as a prooxidant rather than an antioxidant in vivo.
The effects of cabbage leaf protein concentrate (CLPC) on serum and liver lipid concentrations were determined in rats fed choles-terol-enriched and cholesterol-free diets. In rats fed the cholesterol-enriched diet with CLPC, total cholesterol, triacylglycerol and phospholi-pid concentrations in both the serum and liver, as well as the atherogenic index diet were significantly lower than those of the rats fed a casein diet. A supplement of methionine to the CLPC diet raised serum HDL-cholesterol and body weight gain, indicating that the addition of methio-nine to the CLPC diet is not only available to improve the nutritive value of CLPC but also to lower the atherogenic index. In rats fed the cholesterol-free diet, the liver total cholesterol and triacylglycerol concen-trations of the CLPC-fed rats also showed lower values than those of the easein-fed rats, however, the serum total cholesterol concentration of the CLPC-fed rats did not differ from that of the casein-fed rats.