Human Amino Acid Requirements Research Articles

Post-prandial tracer studies of protein and amino acid utilisation: what can they tell us about human amino acid and protein requirements?

Nitrogen-balance, the principal methodology used to derive recommendations for human protein and amino-acid requirements, has been widely criticised, and calls for increased protein and amino-acid requirement recommendation have been made, often on the basis of post-prandial amino-acid tracer-kinetic studies of muscle protein synthesis, or of amino-acid oxidation. In each case, for such studies to inform about requirements over 24hrs or longer, it is arguable that their design and interpretation should take into account an understanding of the impact of varying protein intakes on homeostatic regulation of the FFM throughout the diurnal cycle of feeding and fasting. This narrative review considers our knowledge of this regulation and what can and has been learnt from post-prandial amino-acid tracer studies, about amino-acid and protein requirements. It appears that the FFM in well fed weight-stable adults with healthy lifestyles is fixed at a phenotypic level within a wide range of habitual protein intakes, with muscle-mass likely to be unresponsive to increased intakes. However homeostatic regulation occurs in response to variation in habitual protein intake, with adaptive changes in amino-acid oxidation which influence the magnitude of diurnal losses and gains of body protein. Post-prandial amino-acid tracer studies have been introduced as an alternative to the logistically complex, expensive and difficult to execute 24h [13C-1] amino-acid balance studies, the recognised valid alternative to nitrogen-balance for assessment of amino acid requirements. However, a detailed examination of the most widely used methodology, indicator amino-acid oxidation studies of human amino-acid and protein requirements, shows that their design and interpretation takes no account of the homeostatic regulation of amino acid oxidation in response to varying habitual protein intakes and also involves design factors and analyses which seriously constrain their ability to provide reliable values. New ideas and a much more critical approach to existing work is needed if real progress is to be made in the area.

Beyond the Bowl: Understanding Amino Acid Requirements and Digestibility to Improve Protein Quality Metrics for Dog and Cat Foods.

The determination of amino acid (AA) requirements for mammals has traditionally been done through nitrogen (N) balance studies, but this technique underestimates AA requirements in adult animals. There has been a shift toward researchers using the indicator amino acid oxidation (IAAO) technique for the determination of AA requirements in humans, and recently in dogs. However, the determination of AA requirements specific to adult dogs and cats at maintenance is lacking and the current requirements outlined by the National Research Council are based on a dearth of data and are likely underreporting the requirements of indispensable AA (IAA) for the population. To ensure the physiological requirements of our cats and dogs are met, we need methods to accurately and precisely measure digestibility. In vivo methods, such as ileal cannulation, are most commonly used, however, due to ethical considerations, we are moving away from animal models and toward in vitro methods. Harmonized static digestion models have the potential to replace in vivo methods but work needs to be done to have these methods more accurately represent the gastrointestinal tract (GIT) of cats and dogs. The Digestible IAA Score (DIAAS) is one metric that can help define protein quality for individual ingredients or mixed diets that uses AA SID estimates and ideally those can be replaced with in vitro AA digestibility estimates. Finally, we need accurate and reliable laboratory AA analyses to measure the AA present in complete diets, especially those used to quantify methionine (Met) and cysteine (Cys), both often limiting AAs in cat and dog diets. Together, this will guide accurate feed formulation for our companion animals to satisfy requirements while avoiding over-supplying protein, which inevitably contributes to excess N excretion, affecting both the environment and feed sustainability.

Options for substantiating protein content claims for conventional foods.

In Canada and the United States, front-of-package protein content claims require data to support the quality of the protein. In general, protein quality reflects the product of the amino acid composition of the food protein relative to human amino acid requirements and a measure of digestibility. The currently accepted method in both jurisdictions is the protein digestibility-corrected amino acid score (PDCAAS) that requires the measurement of true fecal protein (nitrogen) digestibility. The latter must be measured in vivo using a rat model. This requirement for animal testing is inconsistent with international efforts to reduce the usage of animals in testing for regulatory purposes. The current commentary positions four options to remove the need to use animal testing for determining protein quality, when considering protein content claim substantiation. These options include (i) a focus on protein quantity alone; (ii) the use of the amino acid score alone, with no correction for digestibility; (iii) the use of a fixed digestibility coefficient to estimate protein quality; and (iv) the use of in vitro methods to measure protein and/or amino acid digestibility. The relative merits and deficiencies of the options are positioned with the goal of encouraging dialogue within the regulatory agencies to move towards alternative approaches for substantiating protein content claims on foods, including those derived from plant-based sources.

Pattern of Amino Acid Requirements in Humans: An Interspecies Comparison using Published Amino Acid Requirement Recommendations

We undertook an interspecies comparison of essential (indispensable) amino acid requirements, in relation to those for total protein, to examine whether the current, internationally proposed human amino acid requirement patterns are significantly different from those of other animals. Data were compiled in their original form and then expressed as mg amino acid/g total dietary protein (N × 6.25) required (amino acid requirement pattern). Patterns of requirements within the various species were organized according to four, arbitrary age-developmental groupings (very young, early growth, growth and mature), and these were compared with the requirements for (a) human infants, (b) 2–5 year old children, (c) 10–12 year old children and (c) adults, respectively. Statistical comparisons determined if significant differences existed between humans and other species, at apparently similar age-development stages, for both the total and for individual indispensable amino acids. The sum of the specific indispensable amino acid requirements for humans was significantly different than that for other species at infancy, growth and adulthood; the greatest differences, however, were for the amino acid requirement pattern in adulthood. The change between the very young and adult was the greatest for humans. Some of this difference might be due to experimental factors, including the biological status of a species within the arbitrary groupings described and the criteria used to determine “requirements.” Nevertheless, it is difficult to escape the conclusion that the current international human amino acid requirements, for adults in particular, appear to be anomalous when judged against data for other animal species.

Indicator Amino Acid Oxidation Does Not Require a Lengthy Period of Adaptation When Estimating Amino Acid Requirements in Healthy Young Men

ObjectivesThe objective of the current study was to determine whether 3 or 7 days, compared to 8 hours of adaptation to the test level of threonine intake alters the threonine requirement in healthy young men.Methods11 men (19–35 years) were studied at 6 levels of threonine intake (5, 10, 15, 20, 25 and 35 mg.kg−1.d−1), in random order. Participants were first habituated to a diet containing intact milk protein (1.0 g.kg−1.d−1) and adequate calories for 2 days. Subjects then received an experimental diet containing the test threonine intakes for 7 days, with IAAO studies performed on days 1, 3, and 7. A total of 162 IAAO studies were completed; 54 for each length of adaptation. The rate of release of 13CO2 from the oxidation of L-[1–13C]phenylalanine (F13CO2) was measured. Phenylalanine flux and kinetics were determined from urinary enrichment of L-[1–13C]phenylalanine. The threonine requirement was determined by applying mixed effect change-point regression in R to the F13CO2 data. The 95% CI of each breakpoint was calculated using parametric bootstrap and the threonine requirement estimates measured on days 1, 3, and 7 were compared using ANOVA. The effect of threonine intake and days of adaptation on phenylalanine flux and kinetics were assessed using marginal linear mixed models and ANOVA.ResultsThe mean threonine requirement (upper, lower 95% CI) for days 1, 3, and 7 was 10.2 (5.4, 15), 10.6 (8, 13.3), and 12.1(9.5, 14.7) mg.kg−1.d−1 respectively, and were not statistically different from each other (p = 0.502). Phenylalanine flux was not affect by threonine intake, however flux significantly decreased with increasing length of adaptation (p = 0.000). The interaction between threonine intake and days of adaptation on phenylalanine balance was significant (p = 0.0015); decreased with increasing length of adaptation at intakes below 10 mg.kg−1.d−1 threonine, and increased with increasing length of adaptation above 15 mg.kg−1.d−1.ConclusionsWe have demonstrated that the short, 8 hour IAAO protocol is valid for the determination of amino acid requirements in humans. This length of adaptation is better positioned to study the amino acid needs of vulnerable populations such as infants, children, and older adults.Funding SourcesThis study was funded by The Canadian Institute of Health Research.

The Nutritional Profiles of Five Important Edible Insect Species From West Africa-An Analytical and Literature Synthesis.

Background: Undernutrition is a prevalent, serious, and growing concern, particularly in developing countries. Entomophagy—the human consumption of edible insects, is a historical and culturally established practice in many regions. Increasing consumption of nutritious insect meal is a possible combative strategy and can promote sustainable food security. However, the nutritional literature frequently lacks consensus, with interspecific differences in the nutrient content of edible insects generally being poorly resolved.Aims and methods: Here we present full proximate and fatty acid profiles for five edible insect species of socio-economic importance in West Africa: Hermetia illucens (black soldier fly), Musca domestica (house fly), Rhynchophorus phoenicis (African palm weevil), Cirina butyrospermi (shea tree caterpillar), and Macrotermes bellicosus (African termite). These original profiles, which can be used in future research, are combined with literature-derived proximate, fatty acid, and amino acid profiles to analyse interspecific differences in nutrient content.Results: Interspecific differences in ash (minerals), crude protein, and crude fat contents were substantial. Highest ash content was found in H. illucens and M. domestica (~10 and 7.5% of dry matter, respectively), highest crude protein was found in C. butyrospermi and M. domestica (~60% of dry matter), whilst highest crude fat was found in R. phoenicis (~55% of dry matter). The fatty acid profile of H. illucens was differentiated from the other four species, forming its own cluster in a principal component analysis characterized by high saturated fatty acid content. Cirina butyrospermi had by far the highest poly-unsaturated fatty acid content at around 35% of its total fatty acids, with α-linolenic acid particularly represented. Amino acid analyses revealed that all five species sufficiently met human essential amino acid requirements, although C. butyrospermi was slightly limited in leucine and methionine content.Discussion: The nutritional profiles of these five edible insect species compare favorably to beef and can meet human requirements, promoting entomophagy's utility in combatting undernutrition. In particular, C. butyrospermi may provide a source of essential poly-unsaturated fatty acids, bringing many health benefits. This, along with its high protein content, indicates that this species is worthy of more attention in the nutritional literature, which has thus-far been lacking.

Effect of harvest season on the nutritional value of bee pollen protein.

Bee pollen is a natural product that has valuable nutritional and medicinal characteristics and has recently garnered increasing attention in the food industry due to its nutritive value. Here, we harvested pollen loads from the Al-Ahsa oasis in eastern Saudi Arabia during spring, summer, autumn, and winter in 2018/2019 to compare the nutritional value of bee pollen protein with the amino acid requirements of honeybees and adult humans. Based on the nutritional value of bee pollen protein, the optimal season for harvesting bee pollen was determined. The composition of the bee pollen showed the highest contents of crude protein, total amino acids, leucine, glutamic acid, valine, isoleucine, threonine, and glycine in samples collected in spring. The highest contents of lysine, phenylalanine, threonine, tryptophan, arginine, tyrosine, and cysteine were observed in samples collected in winter. The highest contents of histidine, methionine, and serine were in samples collected in autumn. Moreover, the highest levels of aspartic acid, proline, and alanine were in samples collected in summer. Leucine, valine, lysine, histidine, threonine, and phenylalanine (except in autumn bee pollen) contents in pollen from all four seasons were above the requirements of honeybees. Leucine, valine, histidine, isoleucine (except in autumn bee pollen), lysine (except in spring and summer bee pollen), and threonine (except in winter and spring bee pollen) in all tested samples were above the requirements of adult humans. In comparison with the minimal amino acid requirements of adult humans and honeybees, the 1st limiting amino acid in bee pollen collected during the different seasons was methionine. Bee pollen collected during spring (March-May) and winter (December-February) can be considered a nutritive food source for adult humans and honeybees.

Production of a high-nutritional-value functional food, the Update1 bread, with the supplementation of the wheat flour with high-protein-content raw food materials

AbstractDuring our research, we added extracted soya bean meal, egg-white powder, gluten, wheat sourdough, and bamboo fibre to wheat flour in order to increase the quantity of the essential amino acid and the biological value of the wheat protein, producing such a functional, health-protecting, health-preservative food product which is suitable to satisfy the essential amino acid requirements of humans, assuming normal nutrition. Furthermore, we could produce such a food, which, on the one hand, was suitable to confine or prevent the essential amino acid’s malnutrition symptoms, while, on the other hand, when applied alone, to meet the consumers’ needs. During our work, we determined the protein content and amino acid composition of the wheat flour, of the additives used in bread baking, and in the bread both baked with supplementation (Update1 bread) and without supplementation (normal bread), as well as the quantity of the Maillard reaction products (hydroxymethylfurfural). We calculated the biological value of the protein of different breads and evaluated the sensory characteristics of the produced functional food and the fortified bread, supplemented with high essential-amino-acid-containing additives.

Dietary Protein and the Health-Nutrition-Agriculture Connection in India.

The connection between the production, availability, and consumption of high-quality food is becoming very important in developing countries. The requirement of protein is linked to its quality, or its digestibility and ability to meet human indispensable amino acid requirements. This requirement is particularly relevant in India, where commonly consumed cereal-based diets and cereal-based food subsidy programs offer low-quality protein and therefore pose a risk of quality protein deficiency. The production of and access to sustainable sources of high-quality protein will be important parts of mitigating risks to human health. Although milk production has risen in India, its consumption by the poor remains low. On the other hand, leguminous grain production, which has greater climate resilience and soil improvement properties, has fallen, yet this can help resource-poor farmers increase their intake of quality food. Nonetheless, concerns about the nutritional quality of plant protein exist and may be more relevant in settings where environmental enteric dysfunction already affects nutrient absorption. With the use of nationally representative household protein consumption data in India, the percentage of the population at risk of quality protein deficiency was found to vary between 4% and 26% among different age groups and between the urban or rural sector. Mitigating these risks requires a greater intake of high-quality proteins, such as pulses and milk, and that food subsidy policies move beyond cereals and become more quality conscious.

Human Protein and Amino Acid Requirements.

Human protein and amino acid nutrition encompasses a wide, complex, frequently misunderstood, and often contentious area of clinical research and practice. This tutorial explains the basic biochemical and physiologic principles that underlie our current understanding of protein and amino acid nutrition. The following topics are discussed: (1) the identity, measurement, and essentiality of nutritional proteins; (2) the definition and determination of minimum requirements; (3) nutrition adaptation; (4) obligatory nitrogen excretion and the minimum protein requirement; (5) minimum versus optimum protein intakes; (6) metabolic responses to surfeit and deficient protein intakes; (7) body composition and protein requirements; (8) labile protein; (9) N balance; (10) the principles of protein and amino acid turnover, including an analysis of the controversial indicator amino acid oxidation technique; (11) general guidelines for evaluating protein turnover articles; (12) amino acid turnover versus clearance; (13) the protein content of hydrated amino acid solutions; (14) protein requirements in special situations, including protein-catabolic critical illness; (15) amino acid supplements and additives, including monosodium glutamate and glutamine; and (16) a perspective on the future of protein and amino acid nutrition research. In addition to providing practical information, this tutorial aims to demonstrate the importance of rigorous physiologic reasoning, stimulate intellectual curiosity, and encourage fresh ideas in this dynamic area of human nutrition. In general, references are provided only for topics that are not well covered in modern textbooks.

Dietary phenylalanine requirement of adult Labrador Retrievers using the direct oxidation approach

Historically, the determination of amino acid (AA) requirements in dogs have been done in growing dogs and have been invasive in nature, requiring long periods of confinement and exposure to diets deficient in AA, which result in a reduction in the health and well being of dogs during the study period. Therefore, we were interested in examining whether carbon oxidation methods, with short periods of exposure to diets deficient in a single AA, could be successfully applied to determine AA requirements in dogs. Four adult Labrador Retrievers consumed 7 levels of phenylalanine (Phe), and excess tyrosine (Tyr) in random order and in combination with kibble that was formulated to be deficient in Phe and adequate in dietary energy (3730 kcal/kg as fed) and fed at ~13.5 g kg−1 BW d−1. Dogs were fed test diets for two days prior to conducting the tracer studies. The Phe requirement was determined by applying a 2‐phase linear regression crossover model to tracer oxidation (F13CO2). The mean Phe requirement was 3.27 g/kg diet (upper 95% CI= 4.96 g/kg diet) on a dry matter basis. The current NRC estimate of the adequate intake for Phe is 4.50 g/kg diet and based on growth and nitrogen balance in growing dogs. This magnitude of difference is similar to that seen when estimates of AA requirements in humans using nitrogen balance vs. carbon oxidation methods are compared.

Recent advances in determining protein and amino acid requirements in humans

During the past 25 years a significant amount of research has been conducted to determine amino acid requirements in humans. This is primarily due to advancements in the application of stable isotopes to examine amino acid requirements. The indicator amino acid oxidation (IAAO) method has emerged as a robust and minimally invasive technique to identify requirements. The IAAO method is based on the concept that when one indispensable dietary amino acid (IDAA) is deficient for protein synthesis, then the excess of all other IDAA, including the indicator amino acid, will be oxidized. With increasing intakes of the limiting amino acid, IAAO will decrease, reflecting increasing incorporation into protein. Once the requirement for the limiting amino acid is met there will be no further change in the indicator oxidation. The IAAO method has been systematically applied to determine most IDAA requirements in adults. The estimates are comparable to the values obtained using the more elaborate 24h-indicator amino acid oxidation and balance (24h-IAAO/IAAB) model. Due to its non-invasive nature the IAAO method has also been used to determine requirements for amino acids in neonates, children and in disease. The IAAO model has recently been applied to determine total protein requirements in humans. The IAAO method is rapid, reliable and has been used to determine amino acid requirements in different species, across the life cycle and in disease. The recent application of IAAO to determine protein requirements in humans is novel and has significant implications for dietary protein intake recommendations globally.

Available versus digestible amino acids – new stable isotope methods

The nutritive value of food protein sources is dependent on the amino acid composition and the bioavailability of the nutritionally indispensable amino acids. Traditionally the methods developed to determine amino acid bioavailability have focused on intestinal absorption or digestibility, which is calculated as the percent of amino acid intake that does not appear in digesta or faeces. Traditional digestibility based methods do not always account for gut endogenous amino acid losses or absorbed amino acids which are unavailable due to the effect of heat processing and the presence of anti-nutritional factors, though methods have been developed to address these issues. Furthermore, digestibility based methods require the use of animal models, thus there is a need to develop in vivo methods that can be applied directly in human subjects to identify the proportion of dietary amino acids which is bioavailable, or metabolically available to the body for protein synthesis following digestion and absorption. The indicator amino acid oxidation (IAAO) method developed in our laboratory for humans has been systematically applied to determine almost all indispensable amino acid requirements in adult humans. Oxidation of the indicator amino acid is inversely proportional to whole body protein synthesis and responds rapidly to changes in the bioavailability of amino acids for metabolic processes. Using the IAAO concept, we developed a new in vivo method in growing pigs, pregnant sows and adult humans to identify the metabolic availability of amino acids in foods. The stable isotope based metabolic availability method is suitable for rapid and routine analysis in humans, and can be used to integrate amino acid requirement data with dietary amino acid availability of foods.

Methods to assess amino acid requirements in humans

There are now different stable isotopic methods to measure minimum daily human indispensable amino acid (IAA) requirements. There has been debate on the appropriateness of statistical methods used to define the nature of change in the response. Current isotopic methods measure daily amino acid oxidation and balance of the IAA under test, or of a selected indicator amino acid, to graded intakes of the test IAA. A key concern is how response curves of oxidation/balance are analyzed to find the inflection point (breakpoint) at which the intake requirement is identified. Evaluating the pattern of the response to identify a breakpoint by a two-phase regression appears best. The indicator amino acid oxidation method has also been shortened and developed into a noninvasive protocol suitable for different populations and age groups. The indicator amino acid oxidation and balance method might be considered the best approach currently available, but it is challenging. The short-term indicator oxidation method is noninvasive and nondemanding. IAA requirements, based on these methods, have implications for the quality of protein in the dietary intake of populations. Methods that assess the IAA requirements to support optimal body function also need to be developed.

Biomarkers for optimal requirements of amino acids by animals and humans

Amino acids are building blocks of proteins and key regulators of nutrient metabolism in cells. However, excessive intake of amino acids can be toxic to the body. Therefore, it is important to precisely determine amino acid requirements by organisms. To date, none of the methods is completely satisfactory to generate comprehensive data on amino acid requirements of animals or humans. Because of many influencing factors, amino acid requirements remain a complex and controversial issue in nutrition that warrants further investigations. Benefiting from the rapid advances in the emerging omics technologies and bioinformatics, biomarker discovery shows great potential in obtaining in-depth understanding of regulatory networks in protein metabolism. This review summarizes the current approaches to assess amino acid requirements of animals and humans, as well as the recent development of biomarkers as potentially functional parameters for recommending requirements of individual amino acids in health and disease. Identification of biomarkers in plasma or serum, which is a noninvasive approach, holds great promise in rapidly advancing the field of protein nutrition.

The Study on The Properties of The Yellowfin Tuna (Thunnus albacares) Visceral Protein Hydrolysates Using Commercial Enzymes

In current study, protein hydrolysates were produced using microbial proteases of Alcalase, Protamex and Flavourzyme from Yellowfin tuna viscera. The results indicated that, protein hydrolysate from Alcalase, had significantly higher protein content, nitrogen recovery and degree of hydrolysis than the others (P 0.05). The results of amino acids composition revealed that all hydrolysates were almost similar in amino acids composition. In addition, the protein efficiency ratio (PER) showed that all hydrolysates had high nutritional value. The chemical score results showed that all hydrolysates could fulfill human amino acid requirements. While, in comparison with common carp requirements, the amino acids of methionine, lysine and phenyl alanine, were the limiting amino acids in all hydrolysates. According to the results of this study, Alcalase is the most affective enzyme. Also, Yellowfin tuna hydrolysates have high nutritional value, that can be used as protein source in diets. Key words: Tuna fish, Enzymatic hydrolysis, Commercial enzymes, Nutritional value

Chemical composition and amino acid profile ofPleurotus djamorandPleurotus ostreatuscultivated in Mexico

Pleurotus djamor from Yucatan State, Mexico was compared with a local variety of P. ostreatus , in terms of chemical composition and amino acid profile. No differences were found (P=0.55) in crude protein content of P. djamor and P. ostreatus (20.2±0.15 vs. 20.3±0.16 g/100 g DM., respectively). P. ostreatus had a lower fiber and fat content (P 0.05). P. ostreatus has a higher proportion (P<0.05) of valine, isoleucine, phenylalanine, aspartate and alanine while P. djamor had higher proportion (P<0.05) of proline and glutamate, no difference was found in the remaining amino acids. Both species had a high proportion of aspartate and glutamate. In general, the human amino acid requirements for adults are satisfied, however, leucine and lysine might become limiting. It was concluded that P. djamor can be included in human diet in similar fashion as P. ostreatus .

A kinetic approach to the determination of human amino acid requirements.

A kinetic approach to the determination of human amino acid requirements.

Amino acid requirements in humans: with a special emphasis on the metabolic availability of amino acids

Due to advances made in the development of stable isotope based carbon oxidation methods, the determination of amino acid requirements in humans has been an active area of research for the past 2 decades. The indicator amino acid oxidation (IAAO) method developed in our laboratory for humans has been systematically applied to determine almost all indispensable amino acid requirements in adult humans. Nutritional application of experimentally derived amino acid requirement estimates depends upon the capacity of food proteins to meet the amino acid requirements in humans. Therefore, there is a need to know the proportion of dietary amino acids which are bioavailable, or metabolically available to the body for protein synthesis following digestion and absorption. Although this concept is widely applied in animal nutrition, it has not been applied to human nutrition due to lack of data. We developed a new in vivo method in growing pigs to identify the metabolic availability of amino acids in foods using the IAAO concept. This metabolic availability method has recently been adapted for use in humans. As this newly developed IAAO based method to determine metabolic availability of amino acids in foods is suitable for rapid and routine analysis in humans, it is a major step forward in defining the protein quality of food sources and integrating amino acid requirement data with dietary amino acid availability of foods.

Intestinal Microbial Contribution to Metabolic Leucine Input in Adult Men

New estimates of the indispensable amino acid requirements of adult humans are much higher than previously thought and questions the adequacy of cereal-based diets of low protein quality. However, dietary amino acid requirements may be supplemented by contributions from the intestinal microbiota. This study measured the contribution of intestinal microbes to leucine input in healthy adult men. Fourteen adult men were studied during each of 2 11-d periods (before and after intestinal antimicrobial treatment), in which leucine was supplied at 1.25 times the estimated average requirement (EAR) (d 1–7) and at 2.5 times the EAR (d 8–11) providing an L-amino acid diet. We estimated fasting- and fed-state leucine oxidation on d 7 and d 11 using a13C-leucine tracer infusion. The microbial contribution to body leucine input was calculated from the relationship of leucine oxidation to leucine intake and the reduction in leucine oxidation after antimicrobial treatment. Antimicrobial treatment did not affect the slope of the relationship of leucine oxidation to leucine intake. Mean and fed-state leucine oxidation declined by ∼13 and 20%, respectively (both P < 0.05) after antimicrobial treatment with the 1.25 EAR diet, but not with the 2.5 EAR diet. The contribution of the intestinal microbiota to body leucine input was estimated to be between 19 and 22% at the 1.25 EAR diet. The contribution of the intestinal microbiota to body amino acid homeostasis may be significant at maintenance intakes, but its long-term nutritional importance remains to be determined.