Lewis D. Stegink

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.

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.

Infusion of protein hydrolysates in the newborn infant: plasma amino acid concentrations.

Protein hydrolysate infusions commonly administered to young infants unable to sustain adequate protein intake contain large amounts of glutamate and aspartate. Neurotoxic effects have been reported in other species and were attributed to high doses of these amino acids. Plasma glutamate and aspartate levels in infants treated with such preparations were within normal limits, but other amino acids were markedly below fasting levels. Plasma amino acid levels quickly reflected the amino acid composition of the hydrolysate and some degree of amino acid imbalance resulted when either of the 2 products employed in this study constituted the sole source of protein intake.

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.

Blood methanol concentrations in normal adult subjects administered abuse doses of aspartame

Blood methanol concentrations were measured in 30 normal adult subjects administered aspartame, a dipeptide methyl ester. The doses studied included the 99th percentile of projected daily ingestion (34 mg/kg body weight) and three doses considered to be in the abuse range (100, 150, and 200 mg/kg body weight). Methanol concentrations were below the level of detection (0.4 mg/dl) in the blood of the 12 normal subjects who ingested aspartame at 34 mg/kg. They were significantly elevated (p less than or equal to 0 .001) after ingestion of each abuse dose, with the mean peak blood methanol concentrations and the areas under the blood methanol concentration-time curve increasing in proportion to dose. Mean (+/- SD) peak blood methanol concentrations were 1.27 +/- 0.48 mg/dl at the 100 mg/kg dose, 2.14 +/- 0.35 mg/dl at the 150 mg/kg dose, and 2.58 +/- 0.78 mg/dl at the 200 mg/kg dose. Blood methanol concentrations returned to predosing levels by 8 h after administration of the 100 mg/kg dose. Methanol was still detected in the blood 8 h after the subjects had ingested aspartame at 150 or 200 mg/kg. Blood formate analyses were carried out in the 6 subjects who ingested aspartame at 200 mg/kg, since recent studies indicate that the toxic effects of methanol are due to formate accumulation. No significant increase in blood formate concentrations over predosing concentrations was noted. No changes were noted in any of the blood chemistry profile parameters measured 24 h after aspartame ingestion, compared to values noted before administration. Similarly, no differences were noted in ophthalmologic examinations carried out before and after aspartame loading.

Placental transfer of glutamate and its metabolites in the primate

When radioactive glutamate was infused into pregnant rhesus monkeys, 69 to 88 per cent of radioactivity in the maternal plasma remained in association with glutamate while 10 to 22 per cent was converted to glucose. In the fetal plasma, glucose and lactate accounted for more than 80 per cent of radioactivity, with less than 2 per cent of the label found in glutamate. Maternal glutamate infusions resulting in a ten- to twenty-fold increase in maternal plasma glutamate levels (60 to 100 mumoles per 100 ml.) had no effect upon fetal glutamate levels. Infusions producing maternal glutamate levels 70 times normal (280 mumoles per 100 ml.) did result in some transfer of glutamate to the fetal circulation. Labeled glutamate administered to the fetus at 1.5 to 2.4 Gm. per kilogram of fetal weight did not result in glutamate transfer to the maternal circulation. Infusion of glutamate to the fetus at 5 Gm. per kilogram of fetal weight increased fetal plasma glutamate levels to 2, 000 mumoles per 100 ml. and resulted in some transfer of glutamate to maternal circulation. Glutamate metabolites (lactate and glucose) were readily transferred across the placenta in either direction. These studies indicate that the primate placenta is virtually impermeable to glutamate unless extreme elevations of plasma glutamate are induced.

Plasma and erythrocyte concentrations of free amino acids in adult humans administered abuse doses of aspartame

Plasma and erythrocyte concentrations of amino acids were measured in 18 fasting adult subjects (9 male, 9 female) administered abuse doses of aspartame (100, 150, and 200 mg/kg body weight) dissolved in 500 ml orange juice. Six subjects were studied at each dose. Plasma aspartate concentrations increased significantly (p less than or equal to 0.05) over baseline values after ingestion of each dose. However, the increase was small in each case, and maximal levels observed were below those noted postprandially in formula-fed infants. No significant changes (p greater than 0.05) were noted in erythrocyte glutamate, or erythrocyte aspartate concentrations after any dose. Plasma phenylalanine concentrations increased significantly over fasting concentrations (p less than 0.01) from 15 min to 6 h after each dose, and the increase was proportional to dose. Mean (+/- SD) peak plasma phenylalanine concentrations were 20.3 +/- 2.03, 35.1 +/- 11.3, and 48.7 +/- 15.5 mumol/dl, respectively, after aspartame doses of 100, 150, and 200 mg/kg. Erythrocyte phenylalanine concentrations showed similar changes. Although these phenylalanine concentrations are considerably above the normal postprandial range (12 +/- 3 mumol/dl), they are below values associated with toxic findings. These data indicate little risk to normal subjects from excessive aspartate or phenylalanine levels after ingestion of single abuse loads of aspartame.

Stimulation of Pituitary Hormone Secretion by Neurotransmitter Amino Acids in Humans

The effects of several neurotransmitter amino acids on pituitary hormone secretion were examined in normal humans. Oral administration of 10 g of glutamic acid stimulated the secretion of prolactin (PRL) and cortisol to approximately twice baseline values, with no effect on GH, TSH or LH. Aspartic acid (10 g), taurine (5 g), and cysteine (5 or 10 g) had no consistent effect on any hormone measured, although the lack of effect of aspartic acid may relate to the modest increments in serum concentration achieved. Glutamic acid may be an important modulator of PRL and ACTH secretion in humans.

Repeated ingestion of aspartame-sweetened beverage: Effect on plasma amino acid concentrations in normal adults

Aspartame (APM) is a dipeptide sweetener (L-aspartyl-L-phenylalanine methyl ester). It has been suggested that excessive use of the product might elevate plasma aspartate and phenylalanine concentrations. Eight normal adults (four male, four female) ingested three successive 12-oz servings of APM-sweetened beverage at two-hour intervals. The study was carried out in two parts in a randomized cross-over design. In one study the beverage was not sweetened. In the other, the beverage provided 10 mg APM/kg body weight per serving. Plasma amino acid concentrations were measured throughout the six-hour study period. The addition of APM to the beverage had no significant effect on plasma aspartate concentration. APM addition did increase plasma phenylalanine levels 1.64 to 2.05 mumol/dL above baseline values (5.09 +/- 0.82 mumol/dL) 30 to 45 minutes after each dose. However, plasma phenylalanine levels did not exceed normal postprandial values at any time. The data indicate ready metabolism of APMs amino acid content when administered at levels likely to be ingested by individuals who are heavy users of such beverages.

Toxicity of protein hydrolysate solutions: correlation of glutamate dose and neuronal necrosis to plasma amino acid levels in young mice.

Abstract Casein and fibrin hydrolysate preparations currently used for human total parenteral alimentation therapy contain varying quantities of glutamate and aspartate. The administration of large quantities of these amino acids to the neonatal mouse either orally or by injection, produces a variety of neurotoxic effects, the most marked of which is an acute hypothalamic neuronal necrosis. Olney et al. have recently reported a dose-related incidence of such neuronal necrosis in infant mice injected with these protein hydrolysates, suggesting a possible hazard to the young human infant infused with such preparations. Since the neonatal mouse is acutely sensitive to glutamate-induced neuronal damage, we have measured plasma amino acid levels following injection of these protein hydrolysates in an attempt to determine the threshold plasma glutamate level which first results in neuronal damage. Casein and fibrin hydrolysates were administered subcutaneously to 9- to 11-day-old mice at 3 dose levels (20, 50, and 100 μl/g body weight), and plasma amino acid levels were determined at appropriate time intervals thereafter. Control animals were injected with isotonic saline at 50 μl/g body weight. Comparison of the maximal glutamate levels obtained in this study with the extent of neuronal necrosis reported by Olney et al. , indicated that no neuronal damage would be expected at plasma glutamate levels ranging from 24 to just under 50 μmoles/dl. This comparison indicates that plasma glutamate levels reached 50 to 52 μmoles/dl (normal, 6–12 μmoles/dl) during the hydrolysate injection reported to produce the smallest lesion, and thus approximate a minimal threshold value for the neonatal mouse. The data of Olney et al. indicate that most animals (11 of 12) were able to sustain this level without neuronal damage. Older mice (25 days) showed a marked improvement in ability to metabolize glutamate following injection when compared to 9- to 11-day-old animals. The latter phenomenon likely accounts for the decreased glutamate susceptibility noted in the adult mouse. Since plasma glutamate and aspartate levels remain within normal limits in the human infant during parenteral alimentation, and since the acutely sensitive neonatal mouse tolerates blood glutamate levels of at least 24 μmoles/dl, there appears to be little danger to the human infant during parenteral alimentation therapy.