Current issues in determining dietary protein and amino-acid requirements.

Pregnancy and the first two years of life are periods of rapid growth and yet the knowledge of requirements for protein and dietary indispensable amino acids is very limited. The development of carbon oxidation methods opens the way to studies that should fill these important gaps in knowledge.

Glucose kinetics and pregnancy outcome in Indian women with low and normal body mass indices.

BACKGROUND/OBJECTIVES: Fetal energy demands are met from the oxidation of maternally supplied glucose and amino acids. During the fasted state, the glucose supply is thought to be met by gluconeogenesis. Underweight women with low body mass index (BMI) might be unable to adequately supply amino acids to satisfy the demands of gluconeogenesis. SUBJECTS/METHODS: Glucose kinetics were measured during the first and second trimesters of pregnancy in 10 low-BMI and 10 normal-BMI pregnant women at the 12th hour of an overnight fast using a primed 6 h U-(13)C glucose infusion and was correlated to maternal dietary and anthropometric variables and birth weight. RESULTS: Low-BMI mothers consumed more energy, carbohydrates and protein, had faster glucose production (R (a)) and oxidation rates in the first trimester. In the same trimester, dietary energy and carbohydrate correlated with glucose production, glycogenolysis and glucose oxidation in all women. Both groups had similar rates of gluconeogenesis in the first and second trimesters. Glucose R (a) in the second trimester was weakly correlated with the birth weight (r=0.4, P=0.07). CONCLUSIONS: Maternal energy and carbohydrate intakes, not BMI, appear to influence glucose R (a) and oxidation in early and mid pregnancy.

In vivo arginine production and nitric oxide synthesis in pregnant Indian women with normal and low body mass indices.

BACKGROUND/OBJECTIVES: Nitric oxide (NO) has been proposed as a mediator of vascular expansion during pregnancy. Inability to increase NO synthesis and/or production of its precursor, arginine, may be a contributor to pregnancy-induced hypertension or preeclampsia. Because maternal weight is associated with blood pressure and risk of preeclampsia during pregnancy, it may also influence arginine and/or NO production. The purpose of this study was to determine the in vivo arginine production and NO synthesis rate in pregnant women with normal (n=10) and low (n=10) body mass indices (BMIs). SUBJECTS/METHODS: Arginine flux and NO synthesis rate were measured in the postabsorptive state with constant infusions of 15N2-arginine and 13C,2H4-citrulline. Plasma concentrations of arginine and NO metabolites were also measured. Kinetic parameters were correlated to maternal variables, gestational age, birth weight and blood pressure. RESULTS: Endogenous arginine flux was significantly faster in the low-BMI compared with normal-BMI women in the first trimester (63.1+/-3.4 vs 50.2+/-2.0 micromol/kg per h, P<0.01), but not in the second. Plasma NO concentration was higher (44.7+/-5.3 vs 30.4+/-1.9 micromol/l, P=0.03) and its rate of synthesis trended faster in the low-BMI compared with normal-BMI group in the second trimester. Maternal weight and BMI were negatively correlated with arginine flux in both trimesters and NO synthesis in the second trimester. CONCLUSIONS: These findings suggest, but do not prove, that maternal BMI may be a factor in the ability to produce NO during pregnancy and may be one way by which BMI influences blood pressure during pregnancy.

Synthesis of hepatic secretory proteins in normal adults consuming a diet marginally adequate in protein.

The plasma concentration and hepatic synthesis rates of albumin, transthyretin, very low-density lipoprotein apolipoprotein B-100 (VLDL-apoB-100), high-density lipoprotein apolipoprotein A-1, fibrinogen, alpha1-antitrypsin, and haptoglobin were measured in six normal adults before and after consuming a protein intake of 0.6 g. kg body wt(-1). day(-1) for 7 days. The synthesis of hepatic proteins was measured from the incorporation of [(2)H(5)]- phenylalanine, following prime/continuous infusion, using plasma VLDL-apoB-100 isotopic enrichment to represent the precursor pool. Synthesis of albumin declined by 50% (P < 0.001) following the lower-protein diet, VLDL-apoB-100 declined by 20% (P < 0.001), and apoA-1 declined by 16% (P < 0.05). By contrast, synthesis increased for fibrinogen (50%, P < 0.05) and haptoglobin (90%, P < 0.001). This pattern of change, with decreased synthesis of nutrient transport proteins and increased formation of acute-phase proteins, suggestive of a low-grade inflammatory response, was accompanied by increased plasma concentration of the inflammatory cytokine interleukin 6 (30%, P < 0.05). The pattern of change in the synthesis of hepatic secretory proteins following 7 days on the low-protein diet may be of functional relevance for lipid transport and the capacity to cope with stress.

Amino acid, glucose, and lipid kinetics after palliative resection in a patient with glucagonoma syndrome.

Glucagon excess causes catabolic changes, including enhanced glucose production, lipolysis, and amino acid oxidation. In this study, we evaluate the metabolic effects of debulking surgery on a patient with glucagon-producing tumor. Stable isotope tracer methods were used to measure glucose, glycerol, and alpha-ketoisocaproic acid (alpha KICA) rates of appearance (Ra) into plasma. Measurements were obtained 25 days after surgery in the basal state and during hormonal suppression of glucagon production by infusing somatostatin with insulin replacement. Basal plasma glucagon concentration (14,100 pg/mL) remained high after debulking surgery. Somatostatin infusion decreased plasma glucagon concentration to 6,735 pg/mL and basal substrate kinetics (alpha-KICA Ra from 1.97 to 1.48 micromol/kg/min; glucose Ra from 16.89 to 11.56 micromol/kg/min; and glycerol Ra from 3.33 to 2.74 micromol/kg/min). We conclude that debulking surgery fails to adequately suppress glucagon production and the alterations in substrate metabolism associated with excess glucagon. In these patients, somatostatin therapy can be an effective method to suppress secretion of glucagon and help attenuate its catabolic effects.

Do critically ill surgical neonates have increased energy expenditure?

BACKGROUND/PURPOSE: Adult metabolic studies suggest that critically ill patients have increased energy expenditures and thus require higher caloric allotments. To assess whether this is true in surgical neonates the authors utilized a validated, gas leak-independent, nonradioactive, isotopic technique to measure the energy expenditures of a stable postoperative group and a severely stressed cohort. METHODS: Eight (3.46+/-1.0 kg), hemodynamically stable, total parenteral nutrition (TPN)-fed, nonventilated, surgical neonates (5 with gastroschisis, 2 with intestinal atresia, and 1 with intestinal volvulus) were studied on postoperative day 15.5+/-11.9. These were compared with 10 (BW = 3.20+/-0.2 kg), TPN-fed, extracorporeal life support (ECLS)-dependent neonates, studied on day of life 7.0+/- 2.8. Energy expenditure was obtained using a primed, 3-hour infusion of NaH(13)CO(3'), breath (13)CO(2) enrichment determination by isotope ratio mass spectroscopy, and the application of a standard regression equation. Interleukin (IL)-6 levels and C-reactive protein (CRP) concentrations were measured to assess metabolic stress. Comparisons between groups were made using 2 sample Student's t tests. RESULTS: The mean energy expenditure was 53+/-5.1 kcal/kg/d (range, 45.6 to 59.8 kcal/kg/d) for the stable cohort and 55+/-20 kcal/kg/d (range, 32 to 79 kcal/kg/d) for the ECLS group (not significant, P =.83). The IL-6 and CRP levels were significantly higher in the ECLS group (29 +/-11.5 v 0.7+/-0.6 pg/mL [P<.001], and 31+/-22 v 0.6+/-1.3 mg/L [P<.001], respectively). Mortality rate was 0% for the stable postoperative patients and 30% for the ECLS group. CONCLUSIONS: Severely stressed surgical neonates, compared with controls, generally do not show increased energy expenditures as assessed by isotopic dilution methods. These data suggest that the routine administration of excess calories may not be warranted in critically ill surgical neonates and support the hypothesis that neonates obligately redirect energy, normally used for growth, to fuel the stress response. This is a US government work. There are no restrictions on its use.

Glutathione in disease.

Altered glutathione metabolism in association with increased oxidative stress has been implicated in the pathogenesis of many diseases. However, whether strategies aimed at restoring glutathione concentration and homeostasis are effective in ameliorating or modifying the natural history of these states is unknown. In this review we discuss the pathogenic role for altered glutathione metabolism in such diseases as protein energy malnutrition, seizures, Alzheimer's disease, Parkinson's disease, sickle cell anaemia, chronic diseases associated with ageing and the infected state. In addition, we discuss the efficacy of glutathione precursors in restoring glutathione homeostasis both in vitro and in vivo.

Ligation of a patent ductus arteriosus under fentanyl anesthesia improves protein metabolism in premature neonates.

BACKGROUND/PURPOSE: Although surgical ligation effectively reverses the cardiopulmonary failure associated with patent ductus arteriosus (PDA), previous findings have suggested that such surgery itself elicits a catabolic response in premature neonates. Therefore, the authors sought to quantitatively assess whether PDA ligation under fentanyl anesthesia aggravated or improved the protein metabolism of premature neonates. METHODS: Seven ventilated, premature neonates (birth weight 815 +/- 69 g) underwent PDA ligation with standardized fentanyl anesthesia (15 microg/kg) on day-of-life 8.4 +/- 1.2 and were studied immediately pre- and 16 to 24 hours postoperatively while receiving continuous total parenteral nutrition (TPN). Whole-body protein kinetics were calculated using intravenous 1-[13C]leucine, and skeletal muscle protein breakdown was measured from the urinary 3-methylhistidine to creatinine ratio (MH:Cr). RESULTS: Whole-body protein breakdown (10.9 +/- 1.2 v8.9 +/- 0.8 g/kg/d, P < .05), turnover (17.4 +/- 1.2 v 15.4 +/- 0.8 g/kg/d, P< .05), and MH:Cr (1.95 +/- 0.20 v 1.71 +/- 0.16 micromol:mg, P< .05) decreased significantly after operation. This resulted in a 60% improvement in protein balance (1.6 +/- 0.8 v 2.6 +/- 0.6 g/kg/d, P = 0.08) postoperatively. CONCLUSIONS: Because of decreased whole-body protein breakdown, whole-body protein turnover, skeletal muscle protein breakdown, and increased protein accrual, surgical PDA ligation under fentanyl anesthesia promptly improves the protein metabolism of premature neonates enduring the stress of a PDA.

Validation of a [13C]bicarbonate tracer technique to measure neonatal energy expenditure.

The use of a stable isotope-labeled [13C]bicarbonate infusion to measure energy expenditure is advantageous, as a complete collection of expired air is not required. This technique allows for facile measurements of energy expenditure in intubated neonates. The aim of the present study was to determine the accuracy of energy expenditure estimates in postsurgical neonates by using the [13C]bicarbonate method compared with the current standard, indirect calorimetry. Eight neonates who were receiving total parenteral nutrition [98 +/- 21 (SD) kcal x kg(-1) x d(-1); 3.1 +/- 0.7 (SD) protein g x kg(-1) x d(-1)] were studied on postoperative d 15.5 +/- 11.9. A primed continuous 3-h intravenous infusion of NaH13CO3 and indirect calorimetry were performed simultaneously. Energy expenditure was calculated separately from the Weir equation and from the dilution of 13CO2 in the breath in combination with the individual energy equivalents of CO2 from the diet. The rate of CO2 appearance and energy expenditure calculated from the bicarbonate method (0.725 +/- 0.021 mol x kg(-1) x d(-1); 89.5 +/- 2.5 kcal x kg(-1) x d(-1)) highly correlated (r = 0.94 and 0.98, respectively) with the CO2 excretion and energy expenditure determined by indirect calorimetry (0.489 +/- 0.016 mol x kg(-1) x d(-1); 60.2 +/- 2.0 kcal x kg(-1) x d(-1)) when analyzed nonproportionately to weight. Bland-Altman analysis demonstrated the 95% confidence interval to be +/- 8.2 kcal x kg(-1) x d(-1). Linear regression analysis revealed a highly statistically significant equation relating the two energy expenditures: Indircal (kcal/d) = -9.341 + [0.705 x Bicarb (dcal/d)]; p < 0.001, r2 = 96.4%. We conclude that energy expenditure in neonates can be accurately determined using the [13C]bicarbonate method and a regression equation. Therefore, the bicarbonate method may be useful for determining energy expenditure in neonates not readily accessible to indirect calorimetry, such as those being mechanically ventilated or on extracorporeal life support.

In vivo urea cycle flux distinguishes and correlates with phenotypic severity in disorders of the urea cycle.

Urea cycle disorders are a group of inborn errors of hepatic metabolism that result in often life-threatening hyperammonemia and hyperglutaminemia. Clinical and laboratory diagnosis of partial deficiencies during asymptomatic periods is difficult, and correlation of phenotypic severity with either genotype and/or in vitro enzyme activity is often imprecise. We hypothesized that stable isotopically determined in vivo rates of total body urea synthesis and urea cycle-specific nitrogen flux would correlate with both phenotypic severity and carrier status in patients with a variety of different enzymatic deficiencies of the urea cycle. We studied control subjects, patients, and their relatives with different enzymatic deficiencies affecting the urea cycle while consuming a low protein diet. On a separate occasion the subjects either received a higher protein intake or were treated with an alternative route medication sodium phenylacetate/benzoate (Ucephan), or oral arginine supplementation. Total urea synthesis from all nitrogen sources was determined from [(18)O]urea labeling, and the utilization of peripheral nitrogen was estimated from the relative isotopic enrichments of [(15)N]urea and [(15)N]glutamine during i.v. co-infusions of [5-(amide)(15)N]glutamine and [(18)O]urea. The ratio of the isotopic enrichments of (15)N-urea/(15)N-glutamine distinguished normal control subjects (ratio = 0.42 +/- 0.06) from urea cycle patients with late (0.17 +/- 0.03) and neonatal (0.003 +/- 0.007) presentations irrespective of enzymatic deficiency. This index of urea cycle activity also distinguished asymptomatic heterozygous carriers of argininosuccinate synthetase deficiency (0. 22 +/- 0.03), argininosuccinate lyase deficiency (0.35 +/- 0.11), and partial ornithine transcarbamylase deficiency (0.26 +/- 0.06) from normal controls. Administration of Ucephan lowered, and arginine increased, urea synthesis to the degree predicted from their respective rates of metabolism. The (15)N-urea/(15)N-glutamine ratio is a sensitive index of in vivo urea cycle activity and correlates with clinical severity. Urea synthesis is altered by alternative route medications and arginine supplementation to the degree that is to be expected from theory. This stable isotope protocol provides a sensitive tool for evaluating the efficacy of therapeutic modalities and acts as an aid to the diagnosis and management of urea cycle patients.

Lipolysis and lipid oxidation in cirrhosis and after liver transplantation.

On the basis of the finding that plasma glycerol concentration is not controlled by clearance in healthy humans, it has been proposed that elevated plasma free fatty acid (FFA) and glycerol concentrations in cirrhotic subjects are caused by accelerated lipolysis. This proposal has not been validated. We infused 10 volunteers, 10 cirrhotic subjects, and 10 patients after orthotopic liver transplantation (OLT) with [1-(13)C]palmitate and [(2)H(5)]glycerol to compare fluxes (R(a)) and FFA oxidation. Cirrhotic subjects had higher plasma palmitate (52%) and glycerol (33%) concentrations than controls. Palmitate R(a) was faster (1.45+/-0.18 vs. 0.85+/-0.17 micromol x kg(-1) x min(-1)) but glycerol R(a) and clearance slower (1.20+/-0.09 vs. 1.90+/-0.24 micromol x kg(-1) x min(-1) and 21.2+/-1.2 vs. 44.7+/- 4.9 ml x kg(-) x h(-1), respectively) than in controls. After OLT, plasma palmitate and glycerol concentrations and palmitate R(a) did not differ, but glycerol R(a) (1.16+/-0.11 micromol x kg(-1) x min(-1)) and clearance (26.7+/-2.4 ml x kg(-1) x h(-1)) were slower than in controls. We conclude that 1) impaired reesterification, not accelerated lipolysis, elevates FFA in cirrhotic subjects; 2) normalized FFA after OLT masks impaired reesterification; and 3) plasma glycerol concentration poorly reflects lipolytic rate in cirrhosis and after OLT.

In vivo rates of erythrocyte glutathione synthesis in children with severe protein-energy malnutrition.

Although the compromised GSH status of children with edematous protein-energy malnutrition (PEM) has been documented, the in vivo kinetic mechanism(s) responsible for this is not known. To determine if decreased synthesis contributes to the alteration of GSH homeostasis, the fractional and absolute rates of synthesis of erythrocyte GSH were determined shortly after admission (study 1), approximately 9 days postadmission (study 2), and at recovery (study 3) in seven children with edematous PEM and seven children with nonedematous PEM. Children with edematous PEM had significantly lower erythrocyte GSH and slower absolute rates of GSH synthesis than children with nonedematous PEM both shortly after admission, when they were both malnourished and infected, and approximately 9 days later, when the infection had resolved but they were still malnourished. At these times, the edematous group also had significantly lower erythrocyte GSH concentrations and absolute rates of synthesis than at recovery. Plasma and erythrocyte-free cysteine concentrations of the edematous group were significantly lower at studies 1 and 2 than at recovery. In contrast, erythrocyte GSH concentrations, rates of GSH synthesis, and plasma and erythrocyte free cysteine concentrations of the nonedematous group were similar at all three time points and greater at studies 1 and 2 than in the edematous group. These results confirm that GSH deficiency is characteristic of edematous PEM and suggest that this is due to a reduced rate of synthesis secondary to a shortage in cysteine.

Intestinal glutamate metabolism.

Although it is well known that the intestinal tract has a high metabolic rate, the substrates that are used to generate the necessary energy remain poorly established, especially in fed animals. Under fed conditions, the quantification of substrate used by the gut is complicated by the fact that potential oxidative precursors are supplied from both the diet and the arterial circulation. To circumvent this problem, and to approach the question of the compounds used to generate ATP in the gut, we combined measurements of portal nutrient balance with enteral and intravenous infusions of [U-(13)C]substrates. We studied rapidly growing piglets that were consuming diets based on whole-milk proteins. The results revealed that 95% of the dietary glutamate presented to the mucosa was metabolized in first pass and that of this, 50% was metabolized to CO(2). Dietary glucose was oxidized to a very limited extent, and arterial glutamine supplied no >15% of the CO(2) production by the portal-drained viscera. Glutamate was the single largest contributor to intestinal energy generation. The results also suggested that dietary glutamate appeared to be a specific precursor for the biosynthesis of glutathione, arginine and proline by the small intestinal mucosa. These studies imply that dietary glutamate has an important functional role in the gut. Furthermore, these functions are apparently different from those of arterial glutamine, the substrate that has received the most attention.

Protein kinetics in callipyge lambs.

The objectives for this experiment were to determine the effect of the callipyge phenotype on protein kinetics. We studied callipyge and normal lambs (n = 37) at 5, 8, and 11 wk of age (n = 4 to 7/ group) to determine how protein kinetics are altered by this trait. Total protein, DNA, and RNA and calpastatin activity were measured in five skeletal muscles and in the heart, kidneys, and liver, and protein accretion rates were calculated. At 8 wk, the fractional synthesis rates of proteins in these tissues were measured in vivo using a primed, continuous 8-h infusion of [2H5]phenylalanine. Fractional rates of protein degradation were estimated by differences. At 5 wk of age, muscle weights, protein mass, protein:DNA, RNA:DNA, and calpastatin activity were higher (P < .05) for callipyge, and protein mass differences continued to increase through 11 wk. At 8 wk, fractional rates of protein synthesis and degradation were lower (P < .05) in callipyge than in normal lambs. The organs of callipyge lambs exhibited reduced growth at 11 wk. Thus, enhanced muscle growth seems to be maintained in callipyge lambs by reduced protein degradation rather than increased protein synthesis. However, we cannot exclude the possibility that the initial onset of the callipyge condition may be caused by an increase in the fractional rate of protein synthesis.

Methods for measuring glutathione concentration and rate of synthesis.

Altered gamma-glutamylcysteinylglycine homeostasis has been implicated in a wide variety of human diseases. The measurement of the rates of synthesis or loss of gamma-glutamylcysteinylglycine is necessary in order to make meaningful inferences about changes in gamma-glutamylcysteinylglycine concentration in these diseased states. In this review, we discuss methods for measuring gamma-glutamylcysteinylglycine concentration in biological samples as well as how improvements in the sensitivity of gas chromatography-mass spectrometric analyses have permitted the development of new and convenient stable isotope tracer methods for the in-vivo measurement of gamma-glutamylcysteinylglycine kinetics.

Deficiency in peripheral glutamine production in pediatric patients with burns.

Plasma glutamine levels decrease in association with severe injury, which suggests that the consumption of glutamine exceeds the production of glutamine or possibly represents a deficit in the release of glutamine from skeletal muscle. The goal of this study was to assess the peripheral glutamine kinetic response to prolonged stress in children with critical injuries. To accomplish this purpose, we quantitated peripheral glutamine kinetics in vivo with the use of 5N15 glutamine in 5 children with severe burns (total body surface area, 74%+/-14%; mean +/- SEM) and 3 children who underwent elective scar reconstruction. In the children with severe burns, leg blood flow was significantly elevated (16.2+/-2.1 vs 7.5 +/-0.3 mL/min/100 mL leg volume, P < .02) and the arterial concentration of glutamine was significantly reduced (0.31+/-0.04 vs 0.84+/-0.05 mmol/L, P < .001). The rate of glutamine turnover within the leg was significantly reduced in the patients with acute burns, whereas the net efflux of glutamine was similar between the 2 groups. These findings suggest that plasma glutamine concentrations decrease during severe stress as a result of a deficit in peripheral glutamine release in conjunction with an increased central consumption. This preliminary study supports the notion that exogenous glutamine supplementation in pediatric patients with severe injuries may be needed because of this inadequate skeletal muscle response.

Ethnicity affects the postprandial regulation of glycogenolysis.

We investigated the effect of nutrient intake on glucose metabolism in normal Mexican-Americans (n = 6) and European-Americans (n = 6). Subjects were studied after an 18-h fast and after 5-6 h of ingestion of hourly meals that supplied 6.35 or 12.75 micromol glucose. kg(-1). min(-1). Endogenous glucose production (EGP), gluconeogenesis (GNG), and glycogenolysis (GLY) were estimated by mass isotopomer analysis with [U-(13)C]glucose infusions. Fasting EGP, GNG, and GLY did not differ between the groups. Food ingestion lowered the molar rate of GNG by only 31%. However, while consuming the lower quantity of nutrients, Mexican-Americans had higher plasma glucose (P < 0.05), a 39% higher rate of EGP (P < 0.05), and a 68% (P < 0.025) higher rate of GLY than the European-Americans. At the higher intake, EGP and GLY were suppressed completely in both groups. There was a linear relationship between insulin concentrations, EGP, and GLY in both groups, but the slope of the line was significantly (P < 0.05) greater in the European-Americans. We conclude that the sensitivity of GLY to nutrient intake differs between ethnic groups and that this may play a role in the increased predisposition of Mexican-Americans to type II diabetes.

The determinants of protein catabolism in neonates on extracorporeal membrane oxygenation.

BACKGROUND/PURPOSE: Protein catabolism appears to be markedly elevated among neonates on extracorporeal membrane oxygenation (ECMO). The aim of this study was to determine the effect of dietary caloric intake on protein catabolism in neonates on ECMO to help construct therapies that may promote anabolism. METHODS: Twelve total parenteral nutrition (TPN)-fed (88.1 +/- 5.0 [SE] kcal/kg/d; range, 60 to 113 kcal/kg/d; 2.3 +/- 0.2 g/kg/d protein) neonates were studied on ECMO at day of life 7.2 +/- 0.8 d. Protein kinetics were determined using infusions of NaH13CO3 and 1-[13C]leucine. RESULTS: As expected, C-reactive protein levels were significantly elevated compared with normal controls (44.0 +/- 7.6 mg/L v 1.9 +/- 1.1 mg/L; P < .001). Negative protein balance (-2.3 +/- 0.6 g/kg/d; range, 1 to -6.4 g/kg/d) highly correlated (r = -0.88, P < .001) with total protein turnover. Increased dietary caloric intake correlated with increased amino acid oxidation (r = 0.85, P < .001), increased total protein turnover (r = 0.73, P < .01), continued negative protein balance (r = 0.72, P < .01), increased whole-body protein breakdown (r = 0.66, P < .05), and increased CO2 production rate (r = 0.73, P < .01). CONCLUSIONS: A surplus of dietary caloric intake does not improve protein catabolism and merely increases CO2 production in these highly stressed neonates. Thus, judicious caloric supplementation is warranted.

A minimally invasive tracer protocol is effective for assessing the response of leucine kinetics and oxidation to vaccination in chronically energy-deficient adult males and children.

In disadvantaged populations, recurrent infections lead to a loss of body nitrogen and worsen nutritional status. The resulting malnutrition, in its turn, produces a greater susceptibility to infection. This study aimed to examine the ability of a new minimally invasive tracer protocol to measure leucine oxidation, and then to use it to quantify the effect of vaccination on leucine kinetics and oxidation. Undernourished men (n = 5; body mass index 16.3 +/- 0.9 kg/m(2)) and children (n = 9; age 4.1 +/- 0.6 y; weight-for-age Z-score -2.3 +/- 0.7) underwent metabolic studies 6 d before and 1 d after vaccination with diphtheria, pertussis and tetanus (DPT). The tracer protocol was performed in the fed state and involved two 3-h sequential periods of frequent (20 min) oral doses of NaH(13)CO(3) or [1-(13)C] leucine. Frequent breath samples and urine collections were made. Blood samples were obtained from the men and used for the determination of the isotopic enrichment of alpha-ketoisocaproic acid. The prevaccination oxidation of leucine (percentage of dose +/- SD) was 18.1 +/- 2.3 (men) and 16.7 +/- 3.8 (children). One day after vaccination, these values had risen to 19. 9 +/- 1.9 (P < 0.05) in the men and to 19.5 +/- 4.6 (P < 0.01) in the children. In the adults, vaccination was associated with a rise in whole-body protein breakdown [mg protein/(kg.h)] from 200 +/- 40 to 240 +/- 10 (P < 0.05). A minor simulated infection increases leucine catabolism in undernourished humans and this new, minimally invasive protocol is sufficiently sensitive to measure these changes.