Characteristics and mechanism of glutamine-dipeptide absorption in human intestine.

Using in vivo and in vitro techniques, the mechanism by which intestinal mucosa obtains glutamine from luminal oligopeptides was investigated in humans. The rate of hydrolysis by mucosal brush border membrane was more than threefold greater for alanylglutamine than for glycylglutamine. Despite this difference, rates of dipeptide and amino acid disappearance during intestinal perfusion were greater from test solutions containing glycylglutamine than alanylglutamine. Furthermore, rates of intraluminal appearance of products of hydrolysis during the infusion of two dipeptides were similar and less than 5% of the disappearance rate of the parent dipeptide. In contrast to free glutamine, uptake of peptide-bound glutamine by brush border membrane vesicles was not inhibited by deletion of sodium or addition of free amino acids to the incubation medium but was inhibited by other oligopeptides and stimulated by a proton gradient. Inhibition constants for the saturable uptake of glycylglutamine and alanylglutamine by vesicles were not significantly different, suggesting similar affinities for the peptide transporter. It is concluded that in human intestine the predominant mechanism for assimilation of glutamine-dipeptides is absorption as intact dipeptide rather than hydrolysis.

Oligopeptides: mechanism of renal clearance depends on molecular structure.

We have investigated the relative contribution of hydrolysis, intact transport and urinary excretion to the renal clearance of Gly-Sar, Gly-Sar-Sar, and Gly-Gly-Sar in fed and starved rats. The results obtained from isolated kidney perfusion studies are summarized as follows: 1) clearance was fastest for Gly-Gly-Sar and slowest for Gly-Sar-Sar, 2) urinary excretion of Gly-Sar-Sar exceeded that of Gly-Gly-Sar or Gly-Sar, 3) there was accumulation of products of hydrolysis of Gly-Gly-Sar in the perfusate but not of Gly-Sar or Gly-Sar-Sar, 4) isolated brush-border and basolateral membranes of renal tubular cells lacked hydrolytic activity against Gly-Sar and Gly-Sar-Sar but possessed hydrolytic activity against Gly-Gly-Sar, 5) an excess amount of Gly-Sar-Sar reduced the rate of clearance of Gly-Gly-Sar by approximately 40% and significantly increased urinary excretion of this peptide, 6) the nonfiltering kidney cleared Gly-Gly-Sar at a rate which was 50% of that of the filtering kidney but did not clear Gly-Sar, and 7) starvation for 96 h was without a significant effect on the renal clearance of either Gly-Sar or Gly-Sar-Sar but significantly reduced the renal clearance of Gly-Gly-Sar and the brush-border membrane hydrolase activity against this peptide. We conclude that the molecular structure determines the affinity of oligopeptides for the membrane transport and hydrolytic systems, which, in turn, determines their efficiency for clearance by the kidney.

Determinants of substrate affinity for the oligopeptide/H+ symporter in the renal brush border membrane.

We and others have shown previously the existence of high and low affinity systems for oligopeptide transport in kidney brush border membrane vesicles (BBMV). In the present study we investigated the relationship between the structure of substrates and their affinity for interaction with the high-affinity oligopeptide/H+ transporter in kidney BBMV. Based on competition experiments using [3H]Gly-Gln as a probe we determined the Ki values for more than 60 selected peptides. For a high-affinity interaction with the carrier site the following structural features of substrates are required: (a) both a free amino and carboxyl terminus; (b) the amino group and peptide bond nitrogen located in the alpha-position; (c) a trans peptide bond rather than the cis configuration; (d) L-alpha-amino acid isomers in both COOH and NH2 termini, although D-isomers of hydrophobic amino acids are acceptable in the NH2 terminus; and (e) a backbone of less than 3 amino acid residues. A striking finding of the present study is that, for peptides satisfying these minimal structural requirements, the primary determinant of affinity is hydrophobicity. The fact that there is a highly significant (p less than 0.001) correlation between Ki and hydrophobicity allows the prediction of the affinity for any di- or tripeptide composed of alpha-amino acids in the L-form.

Glycylglutamine: metabolism and effects on organ balances of amino acids in postabsorptive and starved subjects.

The present study was designed to investigate the metabolism of glycylglutamine and its effects on organ balances of amino acids during intravenous infusion of this dipeptide (100 mumol.h-1.kg-1) in postabsorptive and briefly starved (84-86 h) human subjects. Arterial concentrations of glycylglutamine were not significantly different in postabsorptive (265 +/- 18 microM) and starved (241 +/- 13 microM) subjects. Among the organs examined, kidney predominated in clearance of glycylglutamine from plasma. Moreover, renal clearance of glycylglutamine was reduced by starvation (87 +/- 7 vs. 52 +/- 5 mumol/min, P less than 0.01), whereas neither splanchnic nor muscle clearance was significantly affected. Infusion of glycylglutamine raised plasma concentrations of glycine and glutamine by increasing renal release of these amino acids. In postabsorptive subjects the infusion significantly increased splanchnic balances of glycine and glutamine with little or no effect on the muscle balances; the opposite was found in starved subjects. As far as other amino acids are concerned, the infusion decreased the muscle release of alanine and increased renal release of serine. We conclude that the amino acid residues of glycylglutamine are largely metabolized by the splanchnic organs in postabsorptive subjects and by peripheral organs in starved subjects. The latter results in selective inhibition of muscle release of amino acids.

The high and low affinity transport systems for dipeptides in kidney brush border membrane respond differently to alterations in pH gradient and membrane potential.

The principal aim of the present study was to investigate the effects of variation in proton gradient and membrane potential on the transport of glycyl-L-glutamine (Gly-Gln) by renal brush border membrane vesicles. Under our conditions of transport assay, Gly-Gln was taken up by brush border membrane vesicles almost entirely as intact dipeptide. This uptake was mediated by two transporters shared by other dipeptides and characterized as the high affinity (Kt = 44.1 +/- 11.2 microM)/low capacity (Vmax = 0.41 +/- 0.03 nmol/mg protein/5 s) and low affinity (Kt = 2.62 +/- 0.50 mM)/high capacity (Vmax 4.04 +/- 0.80 nmol/mg protein/5 s) transporters. In the absence of a pH gradient, only the low affinity system was operational, but with a reduced transport capacity. Imposing a pH gradient of 1.6 pH units increased the Vmax of both transporters. Kinetic analysis of the rates of Gly-Gln uptake as a function of external pH revealed Hill coefficients of close or equal to 1, indicating that transporters contain only one binding site for the interaction with external H+. The effects of membrane potential on Gly-Gln uptake were investigated with valinomycin-induced K+ diffusion potentials. The velocity of the high affinity system but not of the low affinity system increased linearly with increasing inside-negative K+ diffusion potentials (p less than 0.01). The Kt of neither system was affected by alterations in either pH gradient or membrane potential. We conclude that (a) the high affinity transporter is far more sensitive to changes in proton gradient and membrane potential than the low affinity transporter and (b) in the presence of a pH gradient, transport of each dipeptide molecule requires cotransport of one hydrogen ion to serve as the driving force.

Mechanism of clearance of dipeptides by perfused hindquarters: sarcolemmal hydrolysis of peptides.

The objective of the present experiment was to investigate the mechanism of clearance of a load of dipeptides (10 mumols) by perfused hindquarters of rats. The clearance was progressive over 60 min and was significantly (P less than 0.01) greater for glycylleucine than for glycylglycine (99 vs. 58% disappearance from the medium). Insulin had no significant effect on clearance of these dipeptides but stimulated the net uptake of their constituent amino acids. Investigation of the fate of peptides considered resistant to membrane hydrolysis showed a modest (24%) clearance for glycylsarcosine but a substantial one (89%) for glycylproline. Investigation of hydrolysis by sarcolemmal vesicles of skeletal muscle showed hydrolase activity against glycylglycine and glycylleucine but none against glycylsarcosine and glycylproline. Investigation of hydrolysis in the medium previously used to perfuse hindquarters for 60 min showed considerable activity against glycylleucine and glycylproline but none against glycylglycine and glycylsarcosine. These activities were entirely abolished by p-hydroxymercuribenzoate, an inhibitor of cytoplasmic peptide hydrolases. In conclusion, our data show that the mechanism of clearance of dipeptides by the perfused hindquarters is largely by hydrolysis, and the site of this hydrolysis differs for different dipeptides; hydrolysis is mediated either by plasma membrane enzymes, cytoplasmic enzymes released into the medium, or a combination of both.

Uptake and metabolism of dipeptides by human red blood cells.

A function of the abundant cytoplasmic peptidases in red blood cells could be hydrolysis of oligopeptides circulating in plasma. To investigate whether human red blood cells actively transport dipeptides for this purpose, these cells were incubated with 14C-labelled glycylproline, glycylsarcosine, glycine, proline and alanine. There was uptake of each dipeptide, as indicated by their recovery as dipeptides in the cell cytoplasm. However, after a brief time (1-2 min) uptake of dipeptides abruptly ceased, while that of amino acids continued. As a result, after 30 min red blood cell uptake of amino acids was 5-13-fold greater than that of any dipeptide. Investigation of intracellular contents after 1 min of incubation revealed different metabolism for different dipeptides. The composition of intracellular radioactivity was 19-71% as intact dipeptides, 0-20% as free amino acids and 8-77% as neither dipeptides nor constituent amino acids. Investigation of the mechanism of dipeptide uptake by red blood cells showed: (1) a lack of hydrolysis by the plasma membrane, (2) no non-specific binding to the plasma membrane, and (3) a lack of saturation over a wide range of concentrations (0.05-50 mM). The data suggest that the mechanism of uptake of trace amounts of dipeptides by human red blood cells is either by simple diffusion or by a carrier system which has a very weak affinity for dipeptides. Upon entry, depending on the molecular structure, dipeptides are either hydrolysed or transformed into new compounds. The red blood cell uptake, however, does not appear to play any appreciable role in clearance of dipeptides from the plasma in the human.

Splanchnic, renal, and muscle clearance of alanylglutamine in man and organ fluxes of alanine and glutamine when infused in free and peptide forms.

The present study was designed to investigate organ metabolism of intravenously (IV) infused (100 mumol.h-1.kg-1) alanylglutamine and its amino acid constituents in a group of healthy subjects. The dipeptide clearance (mumol/min) by kidney (51 +/- 3) was significantly (P less than .01) greater than the clearance by either splanchnic organs (19 +/- 6) or skeletal muscle (21 +/- 8). Infusion of alanylglutamine significantly (P less than .01) increased arterial plasma concentrations of free alanine (260 +/- 31 v 330 +/- 38 mumol/L) and free glutamine (620 +/- 66 v 764 +/- 65 mumol/L) when compared with the baseline period. Concurrently, splanchnic uptake of alanine and glutamine increased and muscle release of alanine ceased. However, muscle release of glutamine remained unaffected. Renal balances of alanine and glutamine changed from neutral to negative (net release) and from positive (net uptake) to neutral, respectively. Infusion of a corresponding mixture of alanine and glutamine had similar effects on arterial plasma concentrations and splanchnic and muscle balances of alanine and glutamine, but had no effect on renal balances of these amino acids. From these studies in man, we conclude that kidney predominates over other organs in clearance of alanylglutamine from plasma and that this may account for the different effect of infusion of alanine and glutamine in free and peptide forms on renal fluxes of these amino acids.

Possible sources of glutamine for parenteral nutrition: impact on glutamine metabolism.

Due to its instability, glutamine is not included in solutions for parenteral solution. This problem can be obviated by providing glutamine as acetyl-, glycyl-, or alanylglutamine. Using an organ balance technique in conscious dogs, we investigated metabolism of these three sources of glutamine. Liver, gut, kidney, and muscle participated in clearance of glycyl- and alanylglutamine from plasma, but among these organs only kidney cleared acetylglutamine. Furthermore, there was a large urinary excretion for acetylglutamine (38 +/- 6% of amount infused) but only a trace amount for either dipeptide. The infusion of glutamine-dipeptides resulted in similar increases in blood level of free glutamine. The main source of this increase appeared to be hydrolysis of dipeptides by kidney and release of free glutamine to circulation. During the infusion of both dipeptides, glutamine balance (free and dipeptide forms) was always positive (net uptake) across liver, gut, and kidney but was neutral across muscle. Liver or gut glutamine balances were not significantly different during the infusion of dipeptides, but kidney glutamine balance was twofold greater during the infusion of glycyl- than alanylglutamine. We conclude that among these three sources of glutamine, acetylglutamine is least desirable for use in parenteral nutrition. Glycylglutamine may be preferable over alanylglutamine if the objective is to target glutamine for kidney.

Specificity and mechanism of influence of amino acid residues on hepatic clearance of oligopeptides.

We have investigated the influence of amino acid residues on hepatic clearance of oligopeptides by determining the rate of disappearance (nmol.(min.g liver)-1) of selective oligopeptides from the medium during isolated rat liver perfusion. (a) N terminus: the rate of disappearance of Ala-Leu was greater (p less than 0.01) than those of Gly-Leu, Phe-Leu, and Arg-Leu (208 +/- 13, 135 +/- 13, 116 +/- 12, and 127 +/- 12, respectively). (b) C terminus: the rate of disappearance of Leu-Ala (244 +/- 18) was significantly greater (p less than 0.01) than that of Leu-Gly (145 +/- 16). (c) Number of residues: with each increase in the number of alanine residues (2-4) there was a significant increase in the rate of peptide disappearance, and conversely, with each increase in the number of glycine residues (2-6) there was a significant decrease in the rate of peptide disappearance. Further studies showed no peptide transport by isolated liver plasma membrane vesicles and no significant correlation between the rates of peptide disappearance and hydrolase activities of the perfusion medium but highly significant correlation with hydrolase activity of plasma membrane. We conclude that certain amino acid residues, such as alanine, enhance hepatic clearance of oligopeptides by increasing their affinity as substrates for plasma membrane peptide hydrolases.

Metabolism of dipeptides and their constituent amino acids by liver, gut, kidney, and muscle.

Oligopeptides may enter the bloodstream from endogenous and exogenous sources. Using an organ-balance technique in conscious dogs, we investigated the role of individual organs in removal of two model oligopeptides (glycylleucine and glycylglycine) from plasma under steady-state conditions. Despite an identical infusion rate, arterial concentration of glycylglycine was twofold greater than that of glycylleucine. This appeared to be a result of greater fractional extraction of glycylleucine than glycylglycine by organs. Although all of the organs examined participated in removal of dipeptides from plasma, their roles varied. Liver, kidney, muscle, and gut accounted for the disappearance of 25, 24, 12, and 10% of the infused amount of glycylleucine, respectively. With glycylglycine as the substrate, disappearance across kidney accounted for 37% of the infused amount, whereas muscle, liver, and gut accounted for 18, 15, and 11%, respectively. Finally, we investigated glycine and leucine balances across organs with infusion of these amino acids in free and dipeptide forms. Glycine and leucine balances were uniquely more positive across muscle during the infusion of glycylleucine than the corresponding amino acid mixture. The possible mechanisms included release of products of glycylleucine hydrolysis by all organs except muscle. We conclude that molecular structure influences the organ extraction of dipeptides; if extraction, particularly by the liver, is not sufficiently rapid, kidney assumes a greater role than other organs in dipeptide removal from plasma.

Toward a theory of therapy with cultic victims.

A theoretical framework for treating cultic victims is presented. Data from over 70 cases treated with a conjoint model focuses on the value of this methodology. The specific and special treatment needs of these clients and the nature of their imposed pathology are discussed. A model for freeing the victims of this pathology is delineated.

Metabolism of glycylleucine and its constituent amino acids by liver, muscle, kidney and gut in conscious dogs.

The role of liver, muscle, kidney and gut in the assimilation of intravenously administered glycylleucine was investigated in 8 mongrel dogs. The rates of disappearance of glycylleucine during its passage across liver, muscle, kidney and gut were 1487 +/- 80, 740 +/- 216, 1436 +/- 115, and 602 +/- 103 mumol/(min x kg B.W.), respectively. The infusion of glycylleucine greatly altered the fluxes of glycine and leucine across these organs. The major alterations included increases in the uptake of glycine by the liver and that of leucine by the muscle, and increases in release of leucine by liver and kidney. We conclude that all organs are involved in the assimilation of intravenously administered glycylleucine, but with varying importance. Although liver and kidney appear to be the dominant organs for the assimilation of glycylleucine, the metabolism of glycine is chiefly accomplished in the liver, and that of leucine is chiefly accomplished in the muscle.

Mechanism of hepatic assimilation of dipeptides. Transport versus hydrolysis.

To investigate dipeptide assimilation by the liver, a series of interrelated experiments were performed in rats. Partial hepatectomy prolonged the plasma half-life (min) of Gly-Ala (3.42 +/- 0.22 versus 4.90 +/- 0.35, p less than 0.05) but had no significant effect on plasma half-life of Gly-Leu, Gly-Pro, or Gly-Sar. We then investigated the rate of disappearance (mumol X (g liver X h)-1) of the above four dipeptides (initial concentration = 1 mM) from the medium during isolated liver perfusion. The order of dipeptide disappearance was: Gly-Leu (8.75 +/- 0.65) greater than Gly-Ala (3.36 +/- 0.46) greater than Gly-Pro (1.29 +/- 0.54) greater than Gly-Sar (0.35 +/- 0.12). This order of dipeptide disappearance corresponded exactly to the order of the rates of glycine accumulation in the medium during liver perfusion with the four dipeptides. Addition of glucagon had no effect on the disappearance rate of Gly-Ala from the medium, but reduced accumulation rates of glycine (3.39 +/- 0.30 versus 1.42 +/- 30, p less than 0.01) and alanine (4.42 +/- 0.66 versus 1.35 +/- 0.39, p less than 0.01). Finally, we found that hydrolysis by the liver plasma membranes and/or perfusion medium accounted for disappearance of dipeptides. In conclusion, the liver does not appear to have a transport system for dipeptides, but assimilates dipeptides by extracellular hydrolysis. Hydrolysis is achieved by enzymes either located on the plasma membranes or released from the cytosol. The amino acid residues released as the result of dipeptide hydrolysis are then taken up by the liver.

Influence of molecular structure on half-life and hydrolysis of dipeptides in plasma: importance of glycine as N-terminal amino acid residue.

To investigate the effect of molecular structure on plasma disappearance and metabolism of dipeptides, rats were injected intravenously with individual dipeptides, and at various intervals after injection, dipeptide and amino acid concentrations were measured in plasma, tissues, and urine. In addition, plasma hydrolase activity against individual dipeptides was investigated. The half-lives of Ala-Leu, Ala-Tyr, and Ala-Gln were shorter than those of dipeptides with glycine substituting for alanine. Furthermore, the increases in plasma concentrations of leucine, tyrosine, and glutamine and rates of dipeptide hydrolysis by plasma enzymes were far greater with alanyl than glycyl dipeptides. In fact, Ala-Leu behaved like a mixture of corresponding free amino acids in raising the plasma concentration of leucine while Gly-Leu did not. There was no significant difference in either plasma half-life or hydrolysis when Leu-Gly and Leu-Ala were used as substrates, but both had rapid rates of hydrolysis in plasma. In comparison to Gly-Leu, Phe-Leu and Arg-Leu had shorter half-lives and greater rates of hydrolysis in plasma. On the other hand, Asp-Leu had a slower rate of plasma hydrolysis than Gly-Leu, but its excretion in the urine was much greater than that of Gly-Leu. In contrast to Gly-Leu and Ala-Leu, Gly-Pro was detected intracellularly in liver, muscle, and particularly, kidney. In fact, the intracellular concentration of Gly-Pro in kidney was either equal to or greater than Gly-Pro concentration in plasma. Increases in intracellular amino acid concentration after injection of individual dipeptides were considerably greater in the kidney than in either liver or muscle.(ABSTRACT TRUNCATED AT 250 WORDS)

Effect of starvation on amino acid and peptide transport and peptide hydrolysis in humans.

Jejunal disappearance rates of glycine (a model for neutral amino acid absorption), triglycine (a model for peptide transport), and tetraglycine (a model for brush-border membrane hydrolysis) were investigated by an in situ perfusion technique before and after 2 wk of starvation in seven obese volunteers. The three test solutions of glycine, triglycine, and tetraglycine were equivalent in glycine content. Before starvation glycine absorption was greatest from the triglycine solution and smallest from the glycine solution. Starvation significantly decreased glycine absorption from both glycine and triglycine solutions, but not from the tetraglycine solution. However, glycine absorption was still significantly greater from the triglycine and tetraglycine solutions than from the glycine solution. Starvation had no significant effect on the disappearance rate of triglycine, but it increased the disappearance rate of tetraglycine. We conclude that a) starvation has different effects on functions of mucosal brush-border membrane, for example, it reduces amino acid absorption but enhances peptide hydrolysis; and b) the greater amino acid absorption from peptides is maintained even after 2 wk of starvation, suggesting that peptides are superior to free amino acids as the nitrogen source for enteral nutrition if employed in malnutrition.

Effect of dietary fat, carbohydrate, and protein on branched-chain amino acid catabolism during caloric restriction.

To assess the effect of each dietary caloric source on the catabolism of branched-chain amino acids, we investigated the rate of leucine oxidation before and after obese volunteers consumed one of the following diets for one week: (a) starvation, (b) 300 or 500 cal of fat/d, (c) 300 or 500 cal of carbohydrate/d, (d) 300 or 500 cal of protein/d, (e) a mixture of carbohydrate (300 cal/d) and fat (200 cal/d), or (f) a mixture of carbohydrate (300 cal/d) and protein (200 cal/d). Starvation significantly increased the rate of leucine oxidation (1.4 +/- 0.11 vs. 1.8 +/- 0.16 mmol/h, P less than 0.01). The same occurred with the fat and protein diets. In sharp contrast, the 500-cal carbohydrate diet significantly decreased the rate of leucine oxidation (1.3 +/- 0.13 vs. 0.6 +/- 0.09 mmol/h, P less than 0.01). The same occurred when a portion of the carbohydrate diet was isocalorically replaced with either fat or protein. The cumulative nitrogen excretion during the fat diet and starvation was not significantly different. As compared with the fat diets, the carbohydrate diets on the average reduced the urinary nitrogen excretion by 12 g/wk. Nitrogen balance was positive during the consumption of the 500-cal protein diet, but negative during the consumption of carbohydrate-protein diet. The fat diets, like the protein diets and starvation, greatly increased plasma leucine (119 +/- 13 vs. 222 +/- 15 microM, P less than 0.01) and beta-hydroxybutyrate (0.12 +/- 0.02 vs. 4.08 +/- 0.43 mM, P less than 0.01) concentrations, and significantly decreased plasma glucose (96 +/- 4 vs. 66 +/- 3 mg/dl, P less than 0.01) and insulin (18 +/- 4 vs. 9 +/- 1 microU/ml, P less than 0.05) concentrations. These changes did not occur, or were greatly attenuated, when subjects consumed carbohydrate alone or in combination with fat or protein. We conclude that during brief caloric restriction, dietary lipid and protein, unlike carbohydrate, do not diminish the catabolism of branched-chain amino acids and the decrease in branched-chain amino acid oxidation is associated with protein sparing.

Enrichment of glycine pool in plasma and tissues by glycine, di-, tri-, and tetraglycine.

Very little information is available on metabolism of oligopeptides in vivo. The present studies were performed to investigate the metabolic fate of diglycine, triglycine, and tetraglycine when injected into a central vein in rats. These peptides disappeared rapidly from plasma without any significant loss in urine. Plasma and tissue concentrations of glycine were measured when the same amount of glycine was injected in free or peptide form. Two minutes after the injection of glycine (1.l0 mumol/g body wt), there was over a tenfold increase in plasma glycine concentration. This increase was diminished when diglycine instead of glycine was injected. Each increase in the number of glycine residues resulted in further reduction in the initial rise in plasma glycine concentration was increased by each injection. This was more pronounced in the kidney than in the liver. Injection of triglycine and tetraglycine resulted in greater glycine concentration in the kidney than injection of either glycine or diglycine. Furthermore, unlike liver and muscle, each increase in the number of glycine residues resulted in greater recovery of glycine peptides from the kidney. These results suggest that with each increase in the number of glycine residues a greater amount of injected glycine peptide is taken up by the kidney for hydrolysis to glycine.

Effect of exercise on rates of oxidation, turnover, and plasma clearance of leucine in human subjects.

We have previously hypothesized that increased muscle oxidation of leucine in starvation is an adaptive response to fuel deficiency in this tissue. To investigate this hypothesis further, we have measured the rates of oxidation, turnover, and plasma clearance of [1-14C]leucine in six obese subjects at rest and during 2 h of mild leg exercise. This experimental design was based on the fact that exercise has the greatest impact on energy expenditure in muscle, the principal site for leucine oxidation. Exercise produced a fourfold increase in oxygen consumption. The rate of alpha-decarboxylation of leucine was increased twofold by leg exercise, whereas there were modest decreases (13%) in the rates of turnover and plasma clearance of this amino acid. The plasma concentrations of lactate and alanine increased twofold during exercise, but plasma concentrations of leucine and other amino acids, glucose, beta-hydroxybutyrate, and insulin remained unaltered. Our data suggest that during exercise oxidation of leucine as an energy source increases, whereas the utilization of this amino acid as a substrate for protein synthesis decreases.