Synthesis of cysteine from methionine in normal adult subjects: effect of route of alimentation.

Parenteral alimentation solutions free of cysteine, probably an essential amino acid for premature infants, were administered continuously to eight healthy men through catheters in the superior vena cava and through nasogastric tubes. When the preparation was administered parenterally, the plasma cystine concentration dropped markedly. When feeding was switched to the oral route, the concentration rose immediately, but returned to baseline only when a cystine-containing diet was fed. These studies indicate that the synthesis of cysteine from methionine is limited, even in the adult subject, when cystine-free diets are administered parenterally.

The different effects on the serum lipids and fecal steroids of high carbohydrate diets given orally or intravenously.

The hypothesis that diets high in carbohydrate produce hyperlipidemia in man was tested in new experiments which provided all calories either by the intravenous route or orally. After a base-line general diet, eight healthy men were fed fat-free diets consisting of 80% of the calories from glucose and 20% from an amino acid hydrolysate. The calories were adequate to maintain body weight. The solutions (1 cal/ml) were infused by constant drip over a 24 h period through either a superior vena cava catheter or a nasogastric tube. Each feeding was for 12 days in sequence but assigned in random order. The high CHO diet given orally, as expected, increased the mean base-line serum triglyceride level from 176+/-29 (SE) to 274+/-47. The identical diet given intravenously (i.v.) failed to produce hypertriglyceridemia; triglyceride levels were not significantly changed, 154+/-37, nor were blood glucose levels. Serum insulin levels were higher during the intravenous feeding. In contrast, both i.v. and oral feedings greatly lowered mean serum cholesterol concentration from the base-line value of 220+/-13 mg/100 ml to 135+/-11 and 151+/-13, respectively. However, the serum cholesterol level was significantly lower (P < 0.01) with the intravenous feeding than with the oral feeding. In addition, the fecal excretion of both neutral sterols and bile acids diminished greatly during the period of intravenous feeding. The fecal mass was likewise decreased. The bacterial conversion of cholesterol to conprostanol did not occur with either intravenous or oral feeding, but with both regimens secondary bile acids predominated, as usual, in the bile acid fraction of the stool. These results emphasize the key role of the intestinal mucosa in the etiology of carbohydrate-induced hypertriglyceridemia and as a direct or indirect contributor to plasma triglyceride and cholesterol levels in the absence of dietary lipids. When the gut mucosa was bypassed, carbohydrate-induced hypertriglyceridemia did not occur and both serum triglyceride and serum cholesterol levels decreased greatly at a time when the excretion of steroids in the stool was also reduced.

Amino- and carboxyl-terminal analyses of hepatoma lactate dehydrogenase isozymes.

The M4 isozyme of lactate dehydrogenase was purified to homogeneity from normal rat liver and from two Morris hepatomas (7777 and 7793). Amino-terminal analyses with fluorodinitrobenzene failed to detect the presence of free amino-terminal residues in each enzyme studied. Each enzyme contained between 3.7 and 4.1 moles of protein-bound acetyl groups per mole of enzyme. The amino-terminal peptide, characterized as N-acetylalanylalanine, was isolated from Pronase digests of each isozyme preparation, and quantitative recovery experiments indicated that all acetyl residues were bound at the amino termini. Carboxylterminal analyses demonstrated phenylalanine to be the carboxyl-terminal residue in each enzyme studied. These data indicate no differences in either amino- or carboxyl-terminal regions of the hepatoma M4 isozymes compared to normal liver M4 isozyme.

Rabbit muscle pyruvate kinase. Amino- and carboxyl-terminal studies.

Amino-terminal analysis of rabbit muscle pyruvate kinase (ATP:pyruvate 2-O-phosphotransferase, EC 2.7.1.40) failed to detect the presence of any free amino-terminal residues. Acetyl group analysis demonstrated the presence of between 3.7 and 4.0 mol of acetyl groups per mol of enzyme. The acetylated amino-terminal residue was isolated from pronase digests of the enzyme and identified as N-acetylserine. Quantitative recovery experiments indicated that all acetyl residues are found at the amino termini. Carboxyl-terminal analyses using the tritium exchange method suggested the presence of a blocked carboxyl-terminal residue, supporting previous hydrazinolysis and carboxypeptidase studies.

Effect of insulin and oral glutathione on glutathione levels and superoxide dismutase activities in organs of rats with streptozocin-induced diabetes.

The effect of insulin or glutathione treatment on glutathione content of liver and jejunal mucosa and on superoxide dismutase (SOD) activity of liver, kidney, and erythrocytes was investigated in pair-fed animals with streptozocin (STZ)-induced diabetes. Diabetes lowered hepatic glutathione concentration, but glutathione concentration of the jejunal mucosa was not affected. Insulin, but not oral glutathione, restored hepatic glutathione concentration to normal levels. Diabetes depressed activity of the cytosolic form of SOD in liver, kidney, and erythrocyte. Treatment of diabetic rats with oral glutathione or intramuscular insulin increased cytosolic SOD activity of renal cortex and liver (but not erythrocytes) to control levels. These results suggest a link between glutathione metabolism and cytosolic SOD activity in diabetes.

Aspartame metabolism in normal adults, phenylketonuric heterozygotes, and diabetic subjects.

This study reviews clinical studies testing the effects of various doses of aspartame on blood levels of phenylalanine, aspartate, and methanol in normal subjects and known phenylketonuric heterozygotes. The effect of aspartame on the phenylalanine-to-large neutral amino acid ratio under various feeding situations is shown. The clinical studies of aspartame in diabetic subjects are limited to observations of its effects on blood levels of glucose, lipids, insulin, and glucagon. These studies clearly demonstrate the safety of this high-intensity sweetener for use by humans.

Pregnancy increases guanosine 3',5'-monophosphate in the myometrium independent of nitric oxide synthesis.

The mechanism for myometrial quiescence during pregnancy is unknown. cGMP plays an integral role in the relaxation of smooth muscle, and nitric oxide (NO) is the most important endogenous activator of soluble guanylate cyclase. The purpose of this study was to determine the effect of gestational age on myometrial cGMP and NO synthase (NOS) activity in the guinea pig. Myometrial cGMP content (measured by RIA) rose slowly until 0.49 (fraction of pregnancy completed) gestation before abruptly increasing to 200 times the non-pregnant control value. It then declined precipitously after 0.87 gestation. Of the known isoenzymes of NOS, the messenger RNAs coding for both endothelial and neuronal NOS could be amplified from the myometrium of pregnant and nonpregnant animals using reverse transcriptase-polymerase chain reaction, but inducible NOS messenger RNA was not found. Myometrial calcium-dependent NOS activity (measured by the conversion of L-[U-14C]arginine to [U-14C]citrulline) declined slowly with advancing gestation (r2 = 0.096; slope = -0.34; P = 0.01), but never differed significantly from the activity in nonpregnant animals [31.1 +/- 11 (term pregnancy) vs. 56.9 +/- 16 (nonpregnant) pmol/min.g; P = NS]. Calcium-independent activity declined shortly after conception, and then rose toward the nonpregnant level (r2 = 0.19; slope = 0.45; P = 0.0009). However, at no time was it significantly different from that in the nonpregnant animal. Pregnancy had no effect on myometrial L-arginine and L-citrulline content. The administration of L-nitro-arginine methyl ester (200 mg/kg) to inhibit NOS dramatically increased blood pressure and reduced fetal renal NOS activity, but had no effect on the myometrial cGMP content. Estradiol (500 micrograms/kg for 5 days) modestly increased cGMP, but in contrast to many tissues in which estradiol increases NOS, it had no effect on myometrial NOS activity. We conclude that pregnancy dramatically increases cGMP by a mechanism independent of NOS. The stimulus remains to be identified. The temporal change in cGMP concentration is consistent with the hypothesis that cGMP mediates myometrial quiescence during pregnancy.

The aspartame story: a model for the clinical testing of a food additive.

Toxicology is based on the premise that all compounds are toxic at some dose. Thus, it is not surprising that very large doses of aspartame (or its components--aspartate, phenylalanine, and methanol) produce deleterious effects in sensitive animal species. The critical question is whether aspartame ingestion is potentially harmful to humans at normal use and potential abuse levels. This paper reviews clinical studies testing the effects of various doses of aspartame upon blood levels of aspartate, phenylalanine, and methanol. These studies demonstrate that blood levels of these compounds are well below levels associated with adverse effects in sensitive animal species.

Placental transfer of glucose-amino acid complexes present in parenteral solutions.

Intravenous infusion of glucose-amino acid complexes, formed during heat sterilization of mixtures of glucose and amino acids or protein hydrolysates, has been associated with mild dehydration in infants, and with excessive trace metal ion excretion in both infants and adults. When parenteral solutions containing these glucose-amino acid complexes were infused into pregnant rhesus monkeys, the compounds accumlated in the maternal plasma and were transported to the fetal circulation. Although all of the compounds studied crossed the placenta, fetal levels were consistently lower than maternal levels. Amniotic fluid concentrations of these compounds increased progressively with the length of maternal infusion, presumably through fetal urination into the amniotic sac. In animals infused with solutions not heat sterilized, levels of these compounds could not be detected in maternal plasma and urine, fetal plasma and urine, or in amniotic fluid. In view of possible toxicity of the glucose-amino acid complexes, parenteral solutions containing these compounds should probably be avoided in the intravenous nutrition of the pregnant woman.

Effect of diet on plasma aminograms of low birth weight infants.

The composition of nutrient mixtures for the low birth weight infant is a matter of considerable concern, and questions have been raised about the adequacy of the cystine and tyrosine contents of available preparations. Low birth weight infants were fed isonitrogenous isocaloric formulas whose content of tyrosine and cystine varied 3- and 7-fold, respectively, for 3-day periods in a Latin Square design. Two-hour postprandial plasma aminograms indicate a statistically significant difference (P = 0.05) between plasma cystine levels noted in infants fed the formula containing cystine at 6 mg/100 ml and those fed the formula containing cystine at 28 mg/100 ml. No significant differences were noted between other formula groupings. Plasma tyrosine concentrations were rapidly reduced whenever tyrosine intake was less then 50 mg/kg of body weight. Such a dietary approach may be of value in reduction of the elevated plasma tyrosine levels seen in infants with transient tyrosinemia of prematurity. Postprandial concentrations of plasma amino acids for the low birth weight infant are a useful reference standard for evaluation of the response of the low birth weight infant to new therapeutic feeding mixtures, particularly parental or jejunal feedings.

Effect of aspartame plus monosodium L-glutamate ingestion on plasma and erythrocyte amino acid levels in normal adult subjects fed a high protein meal.

It has been suggested that aspartame addition to meals already containing large amounts of monosodium L-glutamate would result in an early rapid rise in plasma glutamate and/or aspartate concentrations and increase the potential for dicarboxylic amino acid-induced toxicity. Six normal adult subjects were fed hamburger and milk shake meals providing protein at 1 g/kg body weight in a randomized cross-over design. One meal had no additions while the other contained added monosodium L-glutamate and aspartame (each present at 34 mg/kg body weight). The addition of aspartame plus glutamate had little effect on either plasma or erythrocyte concentrations of glutamate or aspartate beyond those arising from the meal itself. Plasma phenylalanine concentrations were significantly higher (p less than 0.05, paired t test) after ingestion of meals containing aspartame plus glutamate reflecting the increased phenylalanine load.

Plasma glutamate concentrations in adult subjects ingesting monosodium L-glutamate in consomme.

The effect of MSG ingestion in consomme on the plasma glutamate concentration response was studied in normal adult subjects. In the first study nine subjects ingested three different consomme servings (providing 0, 25 and 50 mg/kg body weight MSG) in a Latin square design. Plasma glutamate concentrations were not significantly increased over baseline (3.69 +/- 1.08 mumol/dl) when no added MSG was present. However, mean peak plasma glutamate levels increased proportional to dose when MSG was added (10.2 +/- 2.00 and 17.0 +/- 8.06 mumol/dl at 25 and 50 mg/kg body weight respectively). Since six of the nine subjects in this study reported an idiosyncratic symptom response when tested with MSG at 150 mg/kg body weight, nine additional subjects were also studied. They ingested consomme providing MSG at 0 and 50 mg/kg body weight. No significant differences in plasma amino acid responses were noted between the two groups of subjects.

Modulating effect of Sustagen on plasma glutamate concentration in humans ingesting monosodium L-glutamate.

It has been suggested that monosodium L-glutamate (MSG) addition to meals would significantly increase plasma glutamate concentrations compared to values noted after ingestion of protein-bound glutamate. To test this hypothesis, plasma amino acid concentrations were measured in six normal adults ingesting a ready-to-feed liquid meal (Sustagen) containing added MSG at 0, 100, and 150 mg/kg body weight (Latin square design), and compared to plasma values noted after ingestion of 150 mg/kg body weight MSG in water. The mean (+/- SD) peak plasma glutamate concentrations after ingestion of meals providing 0, 100, and 150 mg/kg body weight MSG were 6.64 +/- 1.99, 11.2 +/- 4.89 and 10.8 +/- 3.10 mumol/dl, respectively. Erythrocyte glutamate concentrations were unchanged after each meal. Peak plasma glutamate concentrations after ingestion of meals with added MSG were similar to those noted in normal adults ingesting a similar quantity of protein-bound glutamate. In contrast, ingestion of MSG in water (150 mg/kg body weight) markedly increased the mean (+/- SD) peak plasma glutamate concentration to 71.8 +/- 35.7 mumol/dl. Similarly, the area under the plasma glutamate concentration-time-curve was significantly higher. MSG ingestion with meals results in lower plasma glutamate concentrations than ingestion of equivalent doses in water.

Absence of the biochemical symptoms of essential fatty acid deficiency in surgical patients undergoing protein sparing therapy.

The biochemical symptoms of essential fatty acid deficiency appear within 7 to 10 days of fat-free total parenteral nutrition using glucose-amino acid mixtures. The linoleic acid (C18:2W6) content of all plasma lipid fractions decreases greatly and plasma eicosatrienoic acid (C20:3W9) increases. We have measured the fatty acid composition of the plasma lipid fractions in six surgical patients receiving parenteral nutritional solutions containing only amino acids, and completely free of glucose, before and after 10 to 14 days of such therapy. Biochemical symptoms of essential fatty acid deficiency did not occur. The fatty acid composition of all plasma lipid fractions remained unchanged after this time period. Mobilization of linoleic acid from the adipose tissue occurred with this treatment in contrast to the inhibition of lipolysis of adipose tissue triglyceride produced by continuous infusions of hypertonic glucose-amino acid mixtures.

Effect of aspartame and sucrose loading in glutamate-susceptible subjects.

It has been postulated that individuals reporting an idiosyncratic symptom response after glutamate ingestion might also experience such symptoms after aspartame ingestion. Such sensitive subjects might have been missed in earlier studies of aspartame. In the present study, six subjects reporting various symptoms after glutamate ingestion, but not after placebo, were administered aspartame (34 mg/kg body weight) or sucrose (1 g/kg body weight) dissolved in orange juice in a randomized, cross-over, double-blind study. No subject reported symptoms typical of a glutamate response after either sucrose or aspartame loading. One subject reported slight nausea approximately 1.5 h after aspartame ingestion, but indicated that the symptoms were not those of a glutamate response. Plasma phenylalanine and aspartate levels were similar to those noted in normal subjects administered identical doses of aspartame. The data indicate no effect of aspartame loading in glutamate-susceptible subjects.

Effect of carbohydrate on plasma and erythrocyte glutamate levels in humans ingesting large doses of monosodium L-glutamate in water.

In previous studies, plasma glutamate concentration was lower when equivalent doses of monosodium L-glutamate (MSG) were given with a ready-to-feed liquid formula meal (Sustagen; 0.4 g protein, 1.1 g carbohydrate, 0.06 g fat, 6.6 kcal energy/kg body weight) rather than in water. This difference was suggested to reflect a carbohydrate effect on mucosal cell glutamate metabolism. To test this hypothesis, a large dose of monosodium L-glutamate (150 mg/kg body weight) dissolved in water, with or without added carbohydrate, was administered to eight healthy adult subjects. Carbohydrate was administered at 1.1 g/kg body weight in the form of partially hydrolyzed corn starch (Polycose). In the absence of carbohydrate, the mean (+/- SD) peak plasma glutamate concentration was 59.4 +/- 46.5 mumol/dl, and the incremental area under the plasma glutamate concentration time curve was 3391 +/- 2360 mumol/(dl x min). The addition of carbohydrate to the glutamate solution significantly decreased (p = 0.001) both the mean peak plasma glutamate concentration (7.18 +/- 3.48 mumol/dl) and the incremental area under the plasma glutamate concentration-time-curve (451 +/- 20.8 mumol/(dl x min). Erythrocyte glutamate and aspartate concentrations were not affected by glutamate loading in either test. Delayed gastric emptying did not account for the carbohydrate effect. Carbohydrate is postulated to serve as a pyruvate source for mucosal cells, facilitating the transamination of glutamate and its subsequent metabolism. This process would reduce the release of glutamate to the peripheral circulation.

Effect of sampling site on plasma amino acid concentrations of infants: effect of skin amino acids.

Plasma taurine, aspartate, threonine, serine, glycine, alanine, valine, leucine, tyrosine, phenylalanine, tryptophan, lysine, histidine, and ornithine concentrations are significantly greater (p less than 0.05, "Student's" t test) in blood samples obtained by conventional heel skin puncture techniques from 1-yr-old infants than values in venous plasma. Differences in plasma concentrations of taurine, aspartate, serine, glycine, and ornithine were particularly striking, with levels in plasma collected from the heel being 1.6 to 6.7 times higher than levels in venous plasma. These increased plasma amino acid concentrations were shown to result primarily from contamination of the plasma with amino acids present on the skin surface. Thorough washing and stimulation of blood flow to the heel by warming prior to skin puncture reduced observed differences. Plasma amino acid concentrations of blood samples obtained by conventional heel skin puncture procedures can be "normalized" to venous values through the use of data on the amino acid composition of heel skin washings.

Oligosaccharides as an intravenous energy source in postsurgical patients: utilization when infused with glucose, amino acids, and lipid emulsion.

Utilization of intravenously administered oligosaccharides was evaluated in postsurgical patients by infusing oligosaccharides simultaneously with glucose, amino acids, and lipid emulsion for 4 d postoperatively. Seven patients were infused with a nutritional regimen providing glucose, amino acids, lipid emulsion, and oligosaccharides and seven patients received a similar regimen without oligosaccharides. Patients infused with oligosaccharides received an overall mean (+/- SD) of 144 +/- 41.0 g oligosaccharides per day. The mean overall excretion of total glucose (free plus oligosaccharide-bound) was significantly greater in patients infused with oligosaccharides (65.1 +/- 33.2 g/d) than in controls (1.83 +/- 1.55 g/d). Overall oligosaccharide utilization for the 4-d period was 48.7 +/- 10.1%. Plasma oligosaccharide concentrations increased from a baseline value of 2.43 +/- 1.90 mg/dL to 58.1 +/- 42.3 mg/dL after 4 d of oligosaccharide infusion, suggesting accumulation.

Placental transfer of taurine in the rhesus monkey.

Maternal to fetal transfer of taurine was examined in 10 pregnant monkeys infused with taurine. In four animals infused at 15 mg/kg body weight, maternal plasma taurine concentration increased from preinfusion values of 8.82 +/- 2.61 to 28.0 +/- 6.00 mumol/dl (mean +/- SD) by the end of the 2-h infusion period. Mean (+/- SD) fetal plasma taurine concentration increased concomitantly, rising from preinfusion values of 13.3 +/- 2.61 to 36.6 +/- 17.8 mumol/dl at the end of the infusion, maintaining the normal fetal to maternal plasma ratio of 1.3 to 1.5. Maternal infusion of larger quantities of taurine increased both maternal and fetal plasma taurine levels; however, the placenta was unable to maintain the normal fetal to maternal gradient. In four monkeys infused with taurine at 25 mg/kg body weight over a 2-h period, mean (+/- SD) maternal plasma taurine concentration increased from 10.0 +/- 1.33 to 43.5 +/- 11.0 mumol/dl. Fetal plasma concentration increased from 13.6 +/- 1.05 to 34.9 +/- 5.89 mumol/dl. However, the maternal to fetal plasma taurine ratio fell from 1.36 to 0.80. Similar results were noted in single animals infused with taurine at 50 and 250 mg/kg body weight. Maternal and fetal plasma concentrations of most other amino acids were not affected by taurine infusion. Fetal, but not maternal, plasma alanine concentration increased after taurine infusion. These data indicate that taurine is efficiently concentrated to the fetal circulation at postprandial maternal plasma taurine concentrations.

Effect of sucrose ingestion on plasma glutamate concentrations in humans administered monosodium L-glutamate.

Plasma glutamate concentrations in human subjects are markedly lower when monosodium L-glutamate (MSG) is ingested in consomme with starch than when ingested in consomme alone. This study investigated whether sucrose had a similar effect. Six normal adult subjects (three male, three female) ingested two servings of beef consomme each providing 50 mg MSG/kg body weight in a randomized crossover design. One serving of consomme contained no added carbohydrate; the other provided 0.5 g sucrose/kg body weight. Ingestion of the consomme without sucrose significantly (p less than 0.05) increased the mean plasma glutamate concentration from baseline (4.44 +/- 0.97 mumol/dl) to a peak value of 18.1 +/- 6.99 mumol/dl 30 min after dosing. The area under the plasma glutamate concentration-time curve was 553 +/- 238 mumol/dl X min. When the consomme contained 0.5 g sucrose/kg body weight, both the mean peak plasma glutamate concentration (5.48 +/- 2.19 mumol/dl) and the area under the curve (105 +/- 46 mumol/dl X min) were significantly lower. These data confirm that metabolizable carbohydrate has a significant effect on plasma glutamate concentration response after MSG loading.