Effect of individual amino acids on plasma cholesterol level in rats fed a high cholesterol diet

The effect of individual amino acids added to a cholesterol containing casein diet on the plasma cholesterol level of rats was systematically investigated with objective of confirming the possibility that specific amino acids are responsible for the regulation of plasma cholesterol level by dietary proteins. Half the amino acids tested were effective to some degree in influencing the plasma cholesterol level when amino acids were added singly to the diet at a 5% level. However, only sulfur containing amino acids had significant effects when added to the diet at a 1% level; Met enhanced and Cys reduced the plasma cholesterol level. In addition, the results of an experiment with amino acid mixture diets with different Met, Cys, and/or Gly contents indicated the importance of sulfur amino acids and also of Gly in plasma cholesterol regulation. The results obtained here support the notion that the plasma cholesterol level of rats can be influenced by specific amino acids or amino acid compositions of dietary proteins, at least under the experimental conditions used.     

The effect of amino-acid-supplemented wheat gluten on cholesterol metabolism in the rat

The effect of lysine- and threonine-supplemented wheat gluten on cholesterol metabolism was studied using male weanling rats. Animals were fed on cholesterol-free diets containing 100 or 200 g gluten/kg with or without amino acid supplementation, and compared with animals given 50, 100 and 200 g casein/kg diets, for 3 weeks. A hypocholesterolaemic effect observed with the wheat gluten-fed rats, compared with the animals given 100 and 200 g casein/kg diets, was accompanied by increased turnover of cholesterol as expressed by enhanced cholesterol biosynthesis and increased faecal excretion of cholesterol and bile acids. This effect was not abolished by lysine and threonine supplementation. Low levels of blood cholesterol were also observed in the rats fed on the 50 g casein/kg diet. However, a different mechanism, related to impairment of cholesterol transport from the liver, was most likely responsible for the hypocholesterolaemia found in these protein-malnourished animals. The effect on cholesterol metabolism produced by dietary wheat gluten was independent of the low quality of the protein and of its specific deficiency in lysine and threonine.     

Sulfur amino acid requirements of the growing rat fed eight percent dietary protein

Methionine and methionine plus cystine requirements of the growing rat fed equivalent to 8% dietary protein were determined. Diets formulated to be adequate in all nutrients except sulfur amino acids (SAA) and providing 4% protein from ANRC casein and equivalent to 4% casein protein from amino acids (excluding methionine and cystine) were supplemented with graded levels of L-methionine. The diet providing 0.34% total SAA (0.32% L-methionine + 0.02% L-cystine) gave the best feed/gain ratio, relative net protein ratio (RNPR), liver protein utilization (LPU) and plasma amino acid acid parameters. When similar ANRC casein + crystalline amino acid diets providing 0.44% total SAA but varying in ratio (by weight) by methionine/cystine were fed to rats, optimal feed/gain ratio, RNPR and plasma amino acid parameters were obtained when cystine replaced 33 to 60% of dietary L-methionine. Inclusion of L-cystine at the expense of L-methionine in these 8% protein diets improved overall rat performance and utilization of dietary methionine. Supplementation of the casein + amino acid basal diet with methionine sulfoxide, methionine sulfone (oxidized forms of methionine) or cysteic acid (oxidized form of cysteine/cystine) to provide 0.34% total SAA, indicated that relative to methionine (100), methionine sulfoxide was completely available and methionine sulfone (68) and cysteic acid (55) were less available. Correction for methionine and cystine digestibilities in ANRC casein suggests that the SAA requirements for the growing rat are 0.33% of diet or 4.1% of dietary protein when 8% protein diets are fed.     

Characteristic properties of isolated soy protein in the metabolic changes due to dietary polychlorinated biphenyls

Feeding polychlorinated biphenyls (PCB) to rats causes an increase in activity of liver microsomal drug-metabolizing enzyme system, plasma level of cholesterol and urinary ascorbic acid. The influences of different types of dietary protein, including whole-egg protein, isolated soy protein and wheat gluten, on these metabolic responses were studied. In the animals not receiving PCB, the activity of drug-metabolizing enzymes and plasma cholesterol were well correlated with the quality of dietary protein as shown by growth rate and nitrogen balance. In the animals fed 200 ppm of PCB containing diets, aniline hydroxylase activity still correlated with the protein quality, but in the animals fed isolated soy protein diets containing PCB, aminopyrine N-demethylase activity, cytochrome P-450 content, plasma cholesterol and urinary ascorbic acid were less than those expected from the biological value. However, supplementation of methionine to isolated soy protein diet containing PCB caused a marked increase in urinary ascorbic acid and plasma cholesterol. This suggests that more sulfurcontaining amino acids are required for the maximum increase in these metabolic parameters than those for the growth rate and nitrogen balance.     

Effects of dietary proteins on lipogenic enzymes in rat liver

When fasted rats were fed fat-free diets containing various sources of protein for 3 d, the activities of liver glucose-6-phosphate dehydrogenase, malic enzyme, acetyl-CoA carboxylase and fatty acid synthetase were markedly lower in rats fed soybean protein or gluten than in those fed casein or fish protein. Since malic enzyme mRNA activity was not low in the soybean protein-fed animals, the translation of malic enzyme appears to be suppressed by dietary soybean protein. The incorporation of tritiated water into liver fatty acids was significantly lower in animals fed soybean protein than in those fed casein. The triglyceride levels in plasma and especially in liver were also lower in the groups fed soybean and gluten than in the groups fed casein and fish. In addition, when dietary soybean protein was replaced with amino acids to simulate casein or soybean protein, the effects on the levels of lipogenic enzymes were still found but were not as great. Thus, some effects can be ascribed to the protein itself and some to the amino acid composition of the diet.     

Hepatic cysteine dioxygenase activity and sulfur amino acid metabolism in rats: possible indicators in the evaluation of protein quality

Experiments were carried out to examine the possibility that the sulfur amino acid metabolism of rats may be an indicator of the nutritional value of dietary protein. Rats were fed diets containing 8, 16 or 24% of gluten, soy protein or casein for 3 wk. Hepatic cysteine dioxygenase activity, hepatic concentration of glutathione, cysteine and taurine and urinary taurine were examined. In addition, the sulfur amino acid metabolism of rats fed these diets fortified with the appropriate first limiting amino acid for 7 d was also examined. High urinary taurine excretion was observed in the three gluten groups, whereas very low urinary taurine excretion was observed with up to 24% soy protein or up to 16% casein. The hepatic hepatic cysteine dioxygenase activities of the gluten diet groups were higher than those of corresponding soy protein or casein diet groups, except that of rats fed the 24% casein diet. The hepatic concentrations of both glutathione and cysteine in gluten diet groups were also higher than those of corresponding soy protein or casein diet groups, except 24% soy protein and 16 and 24% casein diet groups. In rats fed the casein or soy protein diets urinary taurine excretion and hepatic cysteine dioxygenase activity increased with increasing methionine supplementation, the first limiting amino acid. Conversely, in rats fed the gluten diet both urinary taurine excretion and hepatic cysteine dioxygenase activity decreased with increasing lysine supplementation, the first limiting amino acid. These findings suggest that urinary taurine excretion and hepatic cysteine dioxygenase activity may be useful as sensitive indicators of the nutritional value of dietary protein.     

Induction of lysine imbalance in rats: relation to competition for lysine transport into the brain in vitro

The ability of amino acids inhibitory of lysine transport into brain slices to induce lysine imbalance was determined by feeding wheat gluten or casein diets with additions of such amino acids. Lysine transport was only moderately inhibited by amino acids; the most effective were basic amino acids or mixtures of indispensable (IAA) or branched chain amino acid (BCAA). Only mild depressions in growth and food intake occurred during a 10-day period when male, 60--65 g rats of the Sprague-Dawley strain were fed lysine-limiting, 18% wheat gluten diets with additions of these amino acids. The effects were prevented by added lysine. Rats allowed a choice between the lysine-imbalanced or non-protein diets selected the imbalanced, wheat gluten diets (in severe imbalances rats will choose the non-protein diet). Growth depression, prevented by added lysine, occurred in rats fed a 6% casein diet supplemented with IAA; individual amino acids were ineffective. Growth depressions also occurred when rats were fed a basal diet containing 6% case in + 5% of an equimolar mixture of nine IAA and supplemented with arginine or more IAA; BCAA were less effective. Additional lysine completely prevented the growth depressions, but growth of rats fed the diets containing arginine and BCAA was greater than that of those fed the extra IAA. It is difficult to induce a severe lysine imbalance; this is consistent with the failure of amino acids to cause under our conditions strong inhibition of lysine transport into brain.     

The quality and quantity of dietary protein affect brain protein synthesis in rats.

The influence of the amino acid supply in diets with different quality and quantity of protein on the rate of protein synthesis in the brain was investigated. Amino acid concentrations in serum and brain altered in accordance with the amino acid levels of the diets, with the exception of some amino acids such as aspartic acid, glutamic acid, glycine and threonine. When rats were fed various levels of dietary casein (0, 5 and 20%), the aggregation of ribosomes increased and the fractional rate of protein synthesis tended to increase with the increase in dietary protein. When rats were fed a 20% casein diet, greater aggregation of brain ribosomes and protein synthesis rate were observed compared with those in rats fed 20% wheat gluten or gelatin diets. The RNA activity was related to the degree of the aggregation of brain ribosomes and the fractional rate of protein synthesis.     

Effects of dietary animal and plant proteins on the cholesterol metabolism in immature and mature rats

Three- and 9-mo-old rats were fed purified diets that contained either casein, cottonseed or soybean protein for 28 d, and plasma total and high density lipoprotein (HDL) cholesterol, lecithin cholesterol acyltransferase (LCAT) activity and excretion of fecal neutral sterols were measured. These analyses were performed in order to examine how various dietary proteins from animal and plant sources fed in a purified diet influence the changes in the cholesterol metabolism of the young and old rats. Both immature (3-mo-old) and mature (9-mo-old) rats fed purified diet containing casein maintained significantly higher plasma total and HDL cholesterol levels than their counterparts fed the same diets but containing plant proteins (soybean and cottonseed). The fractional rate of esterification (FR) of plasma free cholesterol in mature casein-fed rats was lower than that in immature rats. The FR was also lower in immature rats fed casein than in those fed plant protein. The net turnover rate (NR) of plasma cholesteryl esters (CE) tended to be higher in mature rats and in general was not affected by the dietary protein source. The rate of fecal excretion of neutral sterols was significantly higher in immature rats than in mature rats and in animals fed plant proteins at both ages than in those fed casein.     

Effects of casein or soybean protein on plasma cholesterol level in jejunectomized or ileectomized rats

The contribution of enterohepatic circulation of cholesterol and bile acids in the hypocholesterolemic activity of soybean protein isolate (SPI) was compared with the activity of casein. Intact, sham-operated, jejunectomized or ileectomized adult rats were fed a cholesterol-free, purified diet containing either 20% casein or 20% SPI for 7 or 10 d. For the subsequent 7 d the diets were reversed. In intact rats the plasma cholesterol concentration (p-chol) was significantly higher when the casein diet was fed than when the SPI diet was fed. Within 3 d after the diet crossover, p-chol in each group of intact rats reached rapidly the same level as that in its previous counterpart. These rapid inherent responses of p-chol to the casein and SPI diets remained unchanged even when the jejunum or the ileum was resected, and p-chol in jejunectomized or ileectomized rats was similar to that found in sham-operated rats. The extent of the change in p-chol induced by exchanging dietary proteins was almost the same among intact, sham-operated, jejunectomized and ileectomized rats. These findings indicate that the interruption of enterohepatic circulation of cholesterol and bile acids is not the major factor involved in the differential effects of dietary proteins on p-chol.     

Biliary steroid excretion in rats fed soybean protein and casein or their amino acid mixtures

Male Wistar rats were fed 1% fat, cholesterol-free semipurified diets containing soybean protein isolate and casein or amino acid mixtures simulating these proteins for 28-30 days. The animals then underwent surgery for biliary diversion, and bile was collected for 2 hours. The rate of bile flow was not influenced by the type of dietary protein. The concentration and output of biliary cholesterol in rats fed soybean protein were significantly higher than in those fed casein, while biliary bile acid excretion was comparable. The group given the soy protein-type amino acid mixture also tended to excrete more biliary cholesterol relative to the casein-type amino acid mixture, although the difference was not significant, while biliary bile acid output was comparable. When 5% fat diets containing 0.5% cholesterol were given to rats, concentrations of both biliary cholesterol and bile acid were elevated significantly on a vegetable protein diet. In these experiments, there was a significant negative correlation between serum cholesterol and biliary cholesterol levels. Stimulation of the biliary excretion of cholesterol seems at least relevant to the cholesterol-lowering action of soybean protein.     

Nitrogen source primarily supplied by amino acids and the efficacy for maximal growth of rats.

The effect of osmotic pressure of diets on be food intake of young rats was investigated by comparing the nutritional effect of casein with the corresponding amino acid mixture at 3.2% nitrogen level. Changes in the osmolarity of diets due to the type of dietary carbohydrate had a little effect on food intake, whereas partial substitution of casein for amino acids resulted in significant increase in food intake. Weight gain and food intake of rats fed the 25% replaced amino acid diet were compared with those of rats fed the casein diet. Food intake was lower at 12.5% replacement than at 25% replacement. These observation suggest that a well-balanced amino acid mixture supported maximal growth of rats when 25% of the amino acid mixture was replace with casein. This suggestion could be experimentally confirmed by using an amino acid mixture based on the amino acid composition of whole-egg protein.     

Effects of addition of arginine, cystine, and glycine to the bovine milk-simulated amino acid mixture on the level of plasma and liver cholesterol in rats

The effects of the addition of varying levels of arginine (Arg), cystine (Cys), and glycine (Gly) to the bovine milk-simulated amino acid mixture on the levels of plasma and liver cholesterol were investigated in rats. The diets containing a high amount of Cys lowered significantly the level of plasma cholesterol as the amount of Cys in diets increased. The serum high density lipoprotein (HDL)-cholesterol level and fecal excretion of acidic steroids were higher in rats fed the Cys diets than in those fed the Arg and Gly diets. No differences, however, was observed in the content of liver cholesterol. Liver triglyceride contents elevated significantly on feeding the Arg and Cys diets. Furthermore, liver phospholipid contents elevated significantly on feeding the Arg diet but lowered on feeding the Cys diet. Therefore, these results indicated that the feeding of a high amount of Cys lowers the plasma cholesterol levels as the result of the enhanced conversion of cholesterol to bile acids.     

Effect of cholesterol supplementation on plasma and liver cholesteryl ester fatty acids in rats fed casein or soy protein

The effect of supplementation with cholesterol on plasma and liver cholesteryl ester fatty acids was examined in rats fed diets containing different protein sources. Weanling male rats were fed a semi-purified diet containing 20% casein or 20% soy protein for seven weeks and then 1% (by weight) of cholesterol was added for three more weeks. Rats fed soy protein grew consistently slower than those fed casein. The plasma and liver cholesterol contents were not significantly different between the two protein groups in the unsupplemented rats; however, they were significantly increased in cholesterol-fed animals, particularly in casein-fed rats. Analysis of the cholesteryl ester fatty acid composition in plasma and liver demonstrated that cholesterol-feeding significantly increased the levels of saturated fatty acids, monounsaturated fatty acids and linoleic acid in casein-fed rats as compared to those fed soy protein. The level of cholesteryl arachidonate in liver, which was low initially, increased slightly in the liver of casein-fed rats but did not significantly change in the plasma of either protein group.     

Dietary animal proteins and cholesterol metabolism in rats

The effects on cholesterol metabolism in rats of diets containing various animal proteins or soy protein were studied. The animal proteins tested were casein, whey protein, fish protein, hemoglobin, plasma proteins, ovalbumin, egg-yolk protein, beef protein and chicken-meat protein. The semi-purified diets were isonitrogenous and balanced for residual fat and cholesterol in the protein preparations. The nature of the dietary protein had no effect on serum cholesterol concentration. Group mean liver cholesterol concentration was increased and fecal excretion of bile acids was decreased by all animal proteins when compared with soy protein. This study suggests that carefully balancing diets for components other than protein in the protein preparations prevents protein effects on serum cholesterol in rats but not on liver cholesterol and bile acid excretion.     

Effects of dietary methionine, cystine and potassium sulfate on serum cholesterol and urinary ascorbic acid in rats fed PCB

Liver weight, liver and urinary ascorbic acid levels and serum cholesterol concentration were higher in rats fed polychlorinated biphenyls (PCB) than in controls. The influences of methionine, cystine and potassium sulfate on these metabolic responses were studied. Methionine or equivalent moles of cystine or potassium sulfate were added to a basal diet containing 10% soy protein isolate. The basal diet contained 0.3% of total sulfur-containing amino acids (S-AAs). When methionine was added to the basal diet, maximum gain in body weight was obtained with 0.5% of dietary S-AAs, while the highest values in serum cholesterol and urinary ascorbic acid were obtained with 0.8% of dietary S-AAs in rats fed PCB. Dietary addition of cystine had little effect on body weight gain. Nevertheless, in rat fed PCB, urinary ascorbic acid and serum cholesterol were significantly higher in rats fed the cystine-supplemented diet than in those fed the unsupplemented diet. Addition of potassium sulfate had no effect on body weight gain, urinary ascorbic acid or serum cholesterol. These results suggest that more S-AAs are required for the highest metabolic response to PCB than for maximum growth, and the higher requirement for S-AAs cannot be replaced by inorganic sulfate.     

Dietary fish protein alters blood lipid concentrations and hepatic genes involved in cholesterol homeostasis in the rat model

It is known that various dietary plant proteins are capable of influencing the lipid metabolism of human subjects and animals when compared with casein. Less, however, is known about the effects of fish protein on the cholesterol and triacylglycerol metabolism. Therefore, two experiments were conducted in which rats were fed diets containing 200 g of either fish protein, prepared from Alaska pollack fillets, or casein, which served as control, per kilogram, over 20 and 22 d, respectively. As parameters of lipid metabolism, the concentrations of cholesterol and triacylglycerols in the plasma and liver, the faecal excretion of bile acids and the hepatic expression of genes encoding proteins involved in lipid homeostasis were determined. In both experiments, rats fed fish protein had higher concentrations of cholesteryl esters in the liver, a lower concentration of cholesterol in the HDL fraction (rho > 1.063 kg/l) and lower plasma triacylglycerol concentrations than rats fed casein (P < 0.05). The gene expression analysis performed in experiment 2 showed that rats fed fish protein had higher relative mRNA concentrations of sterol regulatory element-binding protein (SREBP)-2, 3-hydroxy-3-methylglutaryl coenzyme A reductase, LDL receptor, apo AI, scavenger receptor B1 and lecithin-cholesterol-acyltransferase in their liver than did rats fed casein (P < 0.05). The faecal excretion of bile acids and the mRNA concentrations of cholesterol 7alpha-hydroxylase, SREBP-1c and corresponding target genes were not altered. These findings show that fish protein had multiple effects on plasma and liver lipids that were at least in part caused by an altered expression of the hepatic genes involved in lipid homeostasis.     

Effects of addition of sulfur-containing amino acids and glycine to soybean protein and casein on serum cholesterol levels of rats

The effects of the supplementation of methionine (Met), cystine (Cys), and glycine (Gly) to soybean protein or casein on serum and liver lipid levels were studied in rats. Rats were fed cholesterol-free diets containing 25% soybean protein or casein supplemented with 0.75% Met, 2.5% Gly, or a combination of these two for 4 weeks. The addition of Met to soybean protein caused a significant increase in serum cholesterol and this was slightly ameliorated when Gly was given simultaneously. In rats fed casein diets, serum cholesterol tended to decrease when Gly, or Met and Gly were added. A simultaneous supplementation of Met and Gly to casein resulted in a reduction of hepatic cholesterol. Cystine added at the 0.6% level did not cause demonstrable changes in lipid concentrations except for a drop in serum triglyceride of the casein group. When 2.0% Gly was added to cholesterol-enriched diets containing 20% protein, serum cholesterol decreased significantly only when the protein source was casein and the level attained was comparable to that observed in rats fed soybean protein. Liver cholesterol was also markedly decreased by the addition of Gly to casein. The results suggest a possible role of Gly in the regulation of serum cholesterol levels by dietary protein.     

Protein utilization in rats receiving a low-protein diet with various limiting amino acids

1. A study was made of protein utilization in rats given a variety of low-protein diets containing (g/kg) egg albumin 18, casein 49, gluten 50, or mixtures of either maize protein and gelatine 58, or casein and gelatin 37, each with supplemental methionine. The diets were limiting in leucine, tryptophan or lysine, or in both tryptophan and the sulphur-containing amino acids. 2. Values obtained for net protein utilization (NPU) at these low levels of nitrogen intake were markedly higher than the amino acid score calculated for the different test proteins, except with casein for which the two values were similar. The NPU values agreed more closely with chemical score values based on the content of S-amino acids or isoleucine in the dietary protein.     

The effect of dietary sulfur-containing amino acids on the activity of drug-metabolizing enzymes in rat-liver microsomes.

Male Wistar rats were fed either a balanced diet whose protein source was a mixture of amino acids (diet 1), or a similar diet which differed only in having 48% less the quantity of the sulfur-containing amino acids methionine and cystine (diet 2). The diets were given either continuously for 1 month or for 15 days after a protein-free diet. Both diets 1 and 2 permitted good growth of rats and relatively stable microsomal protein content. Protein depletion decreased the total proteins, total phospholipids, and cytochrome P-450 content, and it strongly increased UDP-glucuronosyltransferase activity in rat-liver microsomes. Repletion with diet 1 restored those values, to the level found in control rats. However, diet 2, given continuously induced an increase in UDP-glucuronosyltransferase activity and in the cytochrome P-450 concentration. Since high UDP-glucuronosyltransferase activity was related to lessened amounts of sulfur-containing amino acids in the diet, we discuss the possible effect of methionine and cystine on the regulation of glucuronoconjugation in relation to sulfoconjugation.