The effects of various vitamins on lipid peroxidation and the suppression of DNA synthesis induced by adriamycin (ADR) in vitro using Ehrlich ascites carcinoma (EAC) cells were studied. ADR produced a concentration-dependent stimulation of lipid peroxidation in EAC cells. α-Tocopherol and coenzyme Q10 inhibited ADR-induced lipid peroxida-tion to about the same extent and these effects were the greatest for all antioxidants added. The inhibitory effect of riboflavin 2', 3', 4', 5'-tetrabu-tyrate was greater than that of riboflavin 5'-phosphate. On measuring incorporation of [3H]thymidine into EAC cells, these vitamins did not alter appreciably the magnitude of the ADR-induced suppression of DNA synthesis in EAC cells.
Male Wistar rats received intraperitoneal injections of adriamycin (4mg/kg body weight/day) and/or riboflavin-butyrate (20mg/kg body weight/day) for 6 consecutive days. Cardiac mitochondria were then prepared for our present experiment. The combined use of riboflavin-butyrate with adriamycin was evaluated for reduction of lipid peroxide formation in rat cardiac mitochondria. In order to find the mechanism of the effect of riboflavin-butyrate, the glutathione peroxidase-glutathione reductase system was examined. Adriamycin reduced the glutathione reductase activity in rat cardiac mitochondria, but did not affect the glutathione peroxidase activity. The mitochondrial content of flavin adenine dinucleotide, a prosthetic group of glutathione reductase, was greatly reduced and apoprotein of glutathione reductase also de-creased. The administration of riboflavin-butyrate with adriamycin in-creased flavin adenine dinucleotide and glutathione reductase activity. These results suggest that exogenous administration of riboflavin-butyrate is capable of reducing lipid peroxide by both enzymatic detoxification through glutathione reductase and non-enzymatic detoxification due to direct reaction with lipid peroxide.
The blood level of [14C]coenzyme Q10 and the redox levels of [14C]coenzyme Q10 in the liver and heart were measured after in-travenous injection of [14C]coenzyme Q10 solubilized in multilamellar liposomes into guinea pigs. The blood level of radioactivity declined biexponentially with half-lives of 11.5 min and 15.6 h in the first and second phases, respectively. The levels of reduced [14C]coenzyme Q10 in the liver and heart reached 55.8 and 46.4%, respectively, of the labeled compound in the tissues at 30min after the injection. Coenzyme Q10-reducing activity in cytosol, microsomes and mitochondria was also investigated. This activity was found in all the fractions. The total activity was the highest in the liver cytosol. Moreover, the results of experiments using a purified enzyme suggested that one of the coenzyme Q10-reducing enzymes was NAD(P)H : quinone oxidoreductase [EC 220.127.116.11, DT-di-aphorase]. These results are discussed in relation to the protective effect of reduced coenzyme Q10 against lipid peroxidation in membranes.
A urinary metabolite of nicotinic acid, single- and double-labeled N1-methylnicotinic acid (trigonelline: Tg) were isolated as urinary excretion products of rats following portal vein injection of [carboxyl-14C]nicotinic acid (14C-NiA) and/or S-adenosyl-L-[methyl-3H]methionine (3H-SAM). The labeled compounds produced were identified as Tg by comparison of ultraviolet absorption pattern and chromatographic be-havior with those of the authentic compound, and by cocrystallization with the authentic carrier. It was demonstrated that enzymatic synthesis of double-labeled Tg is accomplished by reaction between 14C-NiA and 3H-SAM, and the enzymatic product was identified using the above methods. Approximately 1.5% respectively of the radioactivity of 14C-NiA and 3H-SAM was incorporated into Tg by enzymatic reaction. Furthermore, Tg from normal rat urine was isolated by ion-exchange chromatographic procedures and was crystallized from ethyl alcohol. The isolated com-pound was identified as Tg by comparison of ultraviolet spectra, infrared spectra, gas-mass spectra, elemental analysis, melting point and thin layer chromatogram behavior with those of the authentic compound. From these results, it was clearly demonstrated that Tg was one of the metabolites of nicotinic acid in the rat.
Changes in plasma and RBC α-tocopherol after intravenous and oral administration of dl-α-tocopheryl acetate were investigated using vitamin E-deficient rats and were compared with those following the administration of d-α-tocopherol, which is the most active and natural form of tocopherol. 1. Intravenous administration: By the administration of dl-tocopheryl acetate, the elevated plasma tocopherol levels were higher at 6h than those elevated by d tocopherol, but the RBC tocopherol was lower 6 h after the infusion of dl-tocopheryl acetate than after d tocopherol infusion. The tocopheryl acetate incorporated into the RBCs remained for 24h after the administration of tocopheryl acetate, the acetate then becoming undetect-able in plasma. 2. Oral administration: The tocopherol elevated in plasma and RBC was only the α-form and not tocopheryl acetate. In both plasma and RBCs, tocopherol levels were higher after the administration of d tocopherol than after dl-tocopheryl acetate administration, and more was found in RBCs.
Purified γ-glutamyltransferase from hog small intestine was competitively inhibited by glutathione in vitro when L-γ-glutamyl -p-nitroanilide was used as a substrate. An S-acetyldextran derivative of glutathione inhibited the enzyme as well as glutathione, although dextran had no inhibitory effect. γ-Glutamyltransferase in the rat small intestine could utilize in situ L-γ-glutamyl-p-nitroanilide circulated in the lumen, and was inhibited by the impermeable derivative of glutathione which was on the luminal side. These data suggested that the active site of intestinal γ-glutamyltransferase faced the luminal side of the brush border membrane.
The effects of protein and/or energy deficiency on long bone growth were studied using rats of an average weight 100g. Four groups of Wistar rats were respectively fed on a normal diet, a low-energy diet, a low protein-diet, and a low-protein and low-energy diet for 45 days. Both energy and protein deficiency restricted gains of body weight and femur length and the activity of alkaline phosphatase in the tibia to the same extent. The epiphyseal growth plates of the femur in protein-deficient rats were as thin as those in energy-deficient rats. On the other hand, femur width and activity of acid phosphatase in the tibia were lower in protein-deficient animals than those in energy-deficient ones. Serum calcium concentrations and calcium content of bone were the same among all groups.
Hematological values and serum amino-acid concentrations were measured in 17 healthy male adult Papua New Guinea highlanders who live on a sweet-potato staple diet. Hematological values were within the normal range, except for a low serum urea concentration. The concentrations of serum threonine, valine, isoleucine, leucine and tyrosine were significantly lower, and those of arginine, glutamate, glycine and alanine were significantly higher, than in Japanese controls. These amino acid patterns in the serum of Papua New Guinea highlanders are an indication of low protein nutrition and adequate energy supply. Some essential amino acid and urea concentrations in the serum of nine Papua New Guinea subjects fed on an adequate protein diet (1.3g/kg body weight, about twice their habitual diet) for 13 days were significantly increased but were still significantly lower than those of Japanese subjects. Serum alanine decreased on an adequate protein diet. These results show that amino acid uptake and utilization by peripheral tissues may be accelerated on an adequate protein diet. Blood status and serum amino acid concentrations did not show any change except for urea and some amino acids, when five Papua New Guinea highlanders were fed on a low protein diet (0.6g/kg) consecutively for 13 days.
Digestibility of α- and β-cyclodextrin (CD) and nutritional consequences of α-CD and a CD mixture (n-dextrin, α-, β- and γ-CDs =50, 30, 15 and 5% by weight) were investigated in rats. In contrast with β-CD, α-CD was revealed to be indigestible. Growing rats were fed on diets supplemented with the CD mixture at 19.5, 39, 58.5 and 78% levels for 110 days, resulting in smaller weight gain and body fat deposition when they were fed on a higher CD diet. Rates of weight loss during the restricted feeding were faster in rats fed on a higher CD diet. These were due to food efficiency lowered by CD. Reduced serum and liver triacylglycerol (TG) levels were noted during a 110-day period of feeding of the CD diets, and the former was revealed due to a reduced hepatic-intestinal TG secretion rate. Rats fed on a 78% CD diet, which contained α-CD at the 24% level, showed abnormal symptoms such as poor appetite and constipation with gas accumulation in the large intestine, and some of them died during the first 2-week feeding period. However, the surviving animals showed adaptation to the diet in the later period of the 110-day feeding. These results suggest that a-CD may be classified as dietary fiber which can modulate lipid metabolism in rats. Furthermore, the CD mixture may be available as a calorie substitute for weight control, which may owe mostly to α-CD.
The inhibitory effect of dietary taurine on cholesterol gallstone formation was studied using male mice of Jcl : ICR strain. Mice were provided three kinds of semi-purified diet; a cholesterol-free diet (standard), a lithogenic diet containing 0.5% cholesterol and 0.25 sodium cholate (C-CA), and a lithogenic diet supplemented with 5 taurine. The changes of total cholesterol in serum, cholesterol mass in the liver and the gall bladder, and gallstone incidence were studied as a function of time. Gallstone formation was observed only in the mice fed on the C-CA diet for more than 3 weeks. The changes of serum cholesterol concentration were not consistent with gallstone formation. The choles-terol mass of the liver in taurine-supplemented mice decreased after the 3rd week. Cholesterol content of the gall bladder increased with choles-terol gallstone formation. Therefore, the inhibitory effect of dietary taurine on cholesterol gallstone formation may be related to the decrease in cholesterol content of the liver.
Three groups of two-month-old swine were fed on basal diets supplemented with either oleic acid-rich safflower oil, lard or hydro-genated soybean oil in order to monitor the atherogenecity of various dietary fatty acids. The level of plasma triglyceride was highest in the safflower oil group and the level of plasma cholesterol was highest in the lard group. The degree of intimal thickening of the coronary arteries was most severe in the safflower oil group and least severe in the hydrogenated fat group. Both the lard- and safflower oil-supplemented groups displayed lipid-rich coronary arterial lesions. The thickened intima of these two groups contained numerous activated smooth muscle cells, degenerated cells with or without stainable lipid and abundant cell debris. Cellular changes were less conspicuous in the coronary arteries from the hy-drogenated fat group than in those from the other two groups.
Some properties of luminal sucrase4somaltase complex and the effect of luminal fluid on their complex were studied in rat small intestine. Luminal contents were collected by flushing the small intestine with the buffered solution. The enzyme activity was observed in luminal contents and intestinal mucosa. Sucrase and Isomaltase activities were located mainly in the intestinal mucosa. However, approximately 20% of sucrase and 10% of isomaltase activities of total small intestine were found in the luminal contents. A significant amount of sucrase without isomaltase activity, the molecular weight of which was estimated at about 140, 000 daltons, was found in the luminal supernatant of the distal intestine in addition to the complexed form of sucrase and isomaltase. The luminal sucrase and sucrase-isomaltase complex had similar properties such as Km values, optimal pH, molecular weights and antigenicity against anti sucrase-isomaltase antibody compared with brush border membrane-bound sucrase-isomaltase complex. Furthermore, the supernatant of the luminal contents of the ileum had a degradative effect on the isomaltase moiety of the purified sucrase-isomaltase complex and a free sucrase without isomaltase also appeared in vitro as observed in vivo. These results suggest that the sucrase-isomaltase complex is released into the intestinal lumen from the brush border membrane and that a luminal factor affects the degradation step of this enzyme as well as the biosynthesis of su-crase-isomaltase complex.