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Purpose of Review: The colon is an effective bioreactor with auxotrophic microbiota that can serve by hydrolyzing dietary and endogenous protein, as well as by synthesizing essential amino acids through nitrogen salvage. Due to assumed negligible amino acid absorption, this colonic contribution was thought to be minimal, but this may not be true., Recent Findings: Several studies that examined the colonic environment in health and disease, show the presence of proteases in the colonic lumen, which are of both host and microbial origin, along with indirect evidence of amino acid transporters in the colonic epithelium. There are also amino acid biosynthetic pathways in the microflora, and the contribution of colonic amino acid to host amino acid nutrition has been shown in wild animals. Yet, current direct and quantitative evidence on amino acid absorption in human colon is minimal., Summary: Although amino acid absorption in colon is not very well established, current studies show that substantial amounts of amino acid could possibly be contributed to the host by the colon. There is a need for assessing this contribution quantitatively using direct isotopic methods under different nutritional conditions, dietary intakes, and clinical conditions., (Copyright © 2024 Wolters Kluwer Health, Inc. All rights reserved.)

The digestible indispensable amino acid score uses ileal digestibility of each indispensable amino acid (IAA) of a dietary protein to calculate its protein quality. However, true ileal digestibility, which is the exclusive sum of digestion and absorption of a dietary protein up to the terminal ileum, is difficult to measure in humans. It is traditionally measured using invasive oro-ileal balance methods but can be confounded by endogenous secreted protein in the intestinal lumen, although the use of intrinsically labeled protein corrects for this. A recent, minimally invasive dual isotope tracer technique is now available to measure true IAA digestibility of dietary protein sources. This method involves simultaneous ingestion of 2 intrinsically but differently (stable) isotopically labeled proteins, a ( 2 H or 15 N-labeled) test protein and ( 13 C-labeled) reference protein whose true IAA digestibility is known. Using a plateau-feeding protocol, the true IAA digestibility is determined by comparing the steady state ratio of blood to meal test protein IAA enrichment to the similar reference protein IAA ratio. The use of intrinsically labeled protein also distinguishes between IAA of endogenous and dietary origin. The collection of blood samples makes this method minimally invasive. As the α- 15 N and α- 2 H atoms of AAs of the intrinsically labeled protein are prone to label loss because of transamination, underestimation of digestibility, appropriate correction factors need to be employed when using 15 N or 2 H labeled test protein. The true IAA digestibility values of highly digestible animal protein by the dual isotope tracer technique are comparable to that measured by direct oro-ileal balance measurements, but no data are yet available for proteins with lower digestibility. A major advantage is that the minimally invasive method allows for true IAA digestibility measurement in humans across different age groups and physiological conditions., (Copyright © 2022 American Society for Nutrition. Published by Elsevier Inc. All rights reserved.)

Background: Ultra-processed foodstuffs have been replacing traditional beans with tortillas in the Mexican diet in the last decades. Therefore, scientific support is needed to promote a return to good-quality traditional dishes., Objectives: This study aims to evaluate the amino acid digestibility and score of pinto beans (Phaseolus vulgaris) consumed with corn tortillas and guacamole in adults using the dual-tracer method., Methods: The pinto beans were intrinsically labeled using 250 mL of 2H2O (99.8%) per 19 L pot with 3 plants. A paste of cooked beans on toasted corn tortillas and guacamole topping were administered to 3 male and 3 female adults (21-25 years old; BMI, 19-23.5 kg/m2). The protocol was plateau feeding given along with U-[13C]-spirulina protein to evaluate indispensable amino acid (IAA) digestibility using the dual-tracer method. Blood samples were taken in the plateau state. The digestibility of each IAA of the bean protein was calculated by the ratio of its enrichment in the beans to the spirulina in the meal and its appearance in plasma collected in the plateau state, as a percentage corrected by spirulina digestibility. Additionally, the digestible IAA score (DIAAS) was calculated., Results: The 2H enrichment of IAA in the pinto beans was 471 parts per million excess. The isotopic enrichment of 2H and 13C in IAA at 5-8 hours presented plateau states with mean CVs of 12.2% and 13.3%, respectively. The mean digestibility of IAA from pinto beans was 77% ± 1.6%, with the lowest value for threonine. The DIAAS calculated with respect to the pattern requirement for children older than 3 years, adolescents, and adults was 83%, with methionine and cysteine being the limiting amino acids., Conclusions: A Mexican dish of pinto beans, tortillas, and guacamole is a good source of protein as evaluated in adults and could be promoted as a nutritious snack. The assay is registered with the Ethical Committee of the Centro de Investigación en Alimentación y Desarrollo, A.C. as CE/015/2019., (© The Author(s) 2021. Published by Oxford University Press on behalf of the American Society for Nutrition.)

Background: Milk is an important high-quality animal protein source in low- and middle-income countries (LMICs). Although the true ileal digestibility and absorption of milk has been shown to be high in French adults, this may be lower in individuals from LMICs who are at risk of environmental enteropathy., Objective: To determine the true ileal indispensable amino acid (IAA) digestibility of intrinsically labeled goat milk protein in South Indian women of reproductive age (WRA), using the dual-isotope tracer technique, and to measure intestinal absorption of amino acid and inert sugar in the same participants using L-allo-isoleucine and a dual-sugar assay., Methods: Milk with 2H-labeled protein collected from a lactating goat fed intrinsically 2H-labeled fodder (maize and cowpea) was spray dried. Labeled milk protein was administered in a plateau feeding protocol to WRA with normal BMI, in whom urinary lactulose and mannitol recovery and the lactulose/mannitol ratio (LMR) were measured, to determine its true ileal IAA digestibility by the dual-isotope tracer technique with a reference U-13C-amino acid mixture. A phenylalanine absorption index was calculated from the plasma to meal ratio of 13C9 phenylalanine within the digestibility protocol. On a separate day, the allo-isoleucine absorption index was estimated from the ratio of plasma allo-isoleucine enrichments after oral 13C6-15N-L- and intravenous 2H10-L-allo-isoleucine administration., Results: The means ± SDs of true ileal IAA digestibility of goat milk protein, lactulose and mannitol recovery, LMR, allo-isoleucine and phenylalanine absorption index were 94.0 ± 2.9%, 0.09 ± 0.03%, 7.9 ± 2.3%, 0.012 ± 0.004, 88.4 ± 3.8% and 24.5 ± 1.6%, respectively. The LMR correlated with the allo-isoleucine absorption index (rs = -0.93, P = 0.008)., Conclusion: The true ileal digestibility of goat milk protein in South Indian WRA with normal intestinal absorptive function and integrity was comparable to earlier estimates in healthy French adults., (© The Author(s) 2021. Published by Oxford University Press on behalf of the American Society for Nutrition.)

Background: Legumes are an excellent plant source of the limiting indispensable amino acid (IAA) lysine in vegetarian, cereal-based diets. However, their digestibility is poor largely because of their antiprotease content. Extrusion can enhance digestibility by inactivating trypsin inhibitors and thus potentially improve the protein quality of legumes., Objective: We measured the digestibility of extruded chickpea and yellow pea protein with use of a dual stable isotope method in moderately stunted South Indian primary school children., Methods: Twenty-eight moderately stunted children (height-for-age z scores <-2.0 SD and >-3.0 SD) aged 6-11 y from low to middle socioeconomic status were randomly assigned to receive a test protein (extruded intrinsically [2H]-labeled chickpea or yellow pea) along with a standard of U-[13C]-spirulina protein to measure amino acid (AA) digestibility with use of a dual stable isotope method. Individual AA digestibility in the test protein was calculated by the ratios of AA enrichments in the test protein to the standard protein in the food and their appearance in blood plasma collected at 6 and 6.5 h during the experiment, representing a plateau state., Results: The mean AA digestibility of extruded chickpea and yellow pea protein in moderately stunted children (HAZ; -2.86 to -1.2) was high and similar in both extruded test proteins (89.0% and 88.0%, respectively, P = 0.83). However, lysine and proline digestibilities were higher in extruded chickpea than yellow pea (79.2% compared with 76.5% and 75.0% compared with 72.0%, respectively, P < 0.02)., Conclusion: Extruded chickpea and yellow pea protein had good IAA digestibility in moderately stunted children, which was 20% higher than an earlier report of their digestibility when pressure-cooked, measured by the same method in adults. Higher digestibility of lysine and proline highlights better retention of these AA in chickpea during extrusion-based processing. Extrusion might be useful for developing high-quality protein foods from legumes. This trial was registered at www.ctri.nic.in as CTRI/2018/03/012439., (Copyright © The Author(s) 2020.)

Background: Tea, a commonly consumed beverage, contains high amounts of polyphenols that can impair protein digestibility, as demonstrated in vitro. There are no human studies examining the inhibitory influence of tea polyphenols (TPP) on high-quality protein digestibility., Objective: The aim of this study was to determine the effect of black tea on the true indispensable amino acid (IAA) digestibility of whole boiled egg protein, in healthy adult humans, through use of a dual isotope tracer approach., Methods: The effect of black TPP (4.6 mg/mL, ingested as a beverage with the meal) on 2H-labeled whole boiled egg protein, administered with ghee rice and tomato curry, was measured with reference to 13C-spirulina protein in healthy Indian adults aged 20-27 y of both sexes with BMI of 22.0 ± 2.8 kg/m2. The results were then compared to previously determined whole egg mean IAA digestibility measured by the same method, without black tea, in the same subjects (n = 5). To correct for any independent effect of TPP on spirulina protein (used as a standard protein), the true IAA digestibility of 13C-spirulina protein was independently measured with reference to a 2H-amino acid mixture, with and without co-ingestion of black tea, in 3 of the same subjects., Results: The true IAA digestibility of whole boiled egg protein significantly decreased by 17% when co-ingested with black tea. However, there was no significant reduction in the true IAA digestibility of spirulina protein when co-ingested with black tea., Conclusions: TPP protein interactions reduced whole egg digestibility in healthy Indian adults but had minimal effect on spirulina protein digestibility. In populations who are at risk of dietary quality protein inadequacy, the consumption of tea during or after a meal can further increase the risk of inadequacy. This trial was registered at Clinical Trials Registry of India (http://ctri.nic.in) as CTRI/2018/03/012265., (Copyright © American Society for Nutrition 2019.)

The relevance of protein and its constituent amino acids (AAs) in the structure and function of the human body is well known. Accumulating evidence has conferred specific functional and regulatory roles for individual AAs, adding relevance to their requirements across different age groups. The methods for measuring AA requirements have progressed from the classical nitrogen balance to the current stable isotope-based AA balance methods. Requirements of most of the indispensable AA (IAA) have been estimated in healthy Indian population by the best available balance method and has shown to be higher than earlier 1985 WHO/FAO/UNU (World Health Organization/Food and Agriculture Organization/United Nations University) recommendations. In addition, potential changes in the requirement, through adaptation to chronic undernutrition or to infection, have also been evaluated. In 2007, the WHO/FAO/UNU released a recommendation that increased the daily IAA requirement, based on primary evidence from Indian balance studies. This meant that to ensure that the new IAA requirements were met, individual foods or mixed diets needed to be assessed for their protein quality, or their ability to deliver the required amount of IAA. The recent FAO report on protein quality evaluation recommends the use of a new chemical AA score, the digestible IAA score (DIAAS), to replace the earlier protein digestibility corrected AA score. The DIAAS requires the determination of individual AA digestibility at the ileal level. A minimally invasive dual stable isotope tracer-based approach has been developed in India and has been used to determine digestibility of various foods in Indian adults and children. The increase in IAA requirements and subsequent protein quality requirements have implications for national regulatory frameworks, growth and development, and in turn, for economic and agricultural policy., Competing Interests: None

Background: Protein quality assessment through the Digestible Indispensable Amino Acid Score requires accurate measurements of true ileal protein and amino acid digestibility, for which a dual isotope technique was recently developed. However, the ileal digestibility of indispensable amino acids (IAA) in humans from high-quality proteins is not well known., Objective: The aim of this study was to intrinsically label hen's egg and meat protein by the use of uniformly 2H-labeled amino acids, and to measure their true ileal indispensable amino acid (IAA) digestibility via the dual isotope method in humans., Design: 2H-labeled lyophilized boiled egg white protein, whole boiled egg, and cooked meat were obtained from layer hens (BV-300) administered a uniformly 2H-labeled amino acid mix orally for 35 d with their daily feed. The ileal IAA digestibility of these proteins was determined with reference to digestibility of previously characterized [U-13C]spirulina in a dual tracer method in healthy Indian subjects whose intestinal health was measured by the plasma kynurenine-to-tryptophan (KT) ratio., Results: All subjects had normal KT ratios. The mean ± SD true ileal IAA digestibility of 2H-labeled egg white protein, whole boiled egg, and cooked meat was 86.3% ± 4.6%, 89.4% ± 4.5%, and 92.0% ± 2.8%, respectively. Leucine digestibility correlated with the KT ratio (r = -0.772; P = 0.009)., Conclusions: Uniformly 2H-labeled hen's egg and meat protein can be used to measure ileal IAA digestibility by the dual isotope tracer approach in humans. The mean IAA digestibility values for these high-quality proteins in the healthy Indians studied were similar to values obtained in earlier human and animal experiments. Leucine digestibility in these meal matrices correlated with the KT ratio, but this aspect needs further evaluation. This trial was registered at the Clinical Trials Registry of India (http://ctri.nic.in) as CTRI/2018/03/012265.

The Protein Digestibility Corrected Amino Acid Score (PDCAAS) has been adopted for assessing protein quality in human foods since 1991, and the shortcomings of using the PDCAAS have been recognized since its adoption. The 2011 FAO Expert Consultation recognized that the Digestible Indispensable Amino Acid Score (DIAAS) was superior to the PDCAAS for determining protein quality. However, there were insufficient human data on amino acid digestibility before adopting the DIAAS. More human data were needed before DIAAS could be implemented. In 2014, FAO convened an expert working group to propose and agree on research protocols using both human-based assays and animal models to study ileal amino acid digestibility (metabolic availability) of human foods. The working group identified 5 research protocols for further research and development. A robust database of protein digestibility of foods commonly consumed worldwide, including those consumed in low-income countries, is needed for an informed decision on adopting the DIAAS. A review on the impacts of using the DIAAS on public health policies is necessary. It would be advantageous to have a global coordinating effort to advance research and data collection. Collaboration with international and national agriculture institutes is desirable. Opportunities should be provided for young researchers, particularly those from developing countries, to engage in protein-quality research for sustainable implementation of DIAAS. To conclude, the DIAAS is a conceptually preferable method compared with the PDCAAS for protein and amino acid quality evaluation. However, the complete value of the DIAAS and its impact on public health nutrition cannot be realized until there are sufficient accumulated ileal amino acid digestibility data on human foods that are consumed in different nutritional and environmental conditions, measured by competent authorities. A future meeting may be needed to evaluate the size and quality of the data set and to determine the timeline for full adoption and implementation of the DIAAS., (© 2016 American Society for Nutrition.)

Background: In India, the prevalence of low birth weight is high in women with a low body mass index (BMI), suggesting that underweight women are not capable of providing adequate energy and protein for fetal growth. Furthermore, as pregnancy progresses, there is increased need to provide methyl groups for methylation reactions associated with the synthesis of new proteins and, unlike normal-BMI American women, low-BMI Indian women are unable to increase methionine transmethylation and remethylation rates as pregnancy progresses from trimester 1 to 3. This also negatively influences birth weight., Objective: The aim was to determine the effect of dietary supplementation with energy and protein from 12 ± 1 wk of gestation to time of delivery compared with no supplement on pregnancy outcomes, protein kinetics, and the fluxes of the methyl group donors serine and glycine., Methods: Protein kinetics and serine and glycine fluxes were measured by using standard stable isotope tracer methods in the fasting and postprandial states in 24 pregnant women aged 22.9 ± 0.7 y with low BMIs [BMI (in kg/m(2)) ≤18.5] at 12 ± 1 wk (trimester 1) and 30 ± 1 wk (trimester 3) of gestation. After the first measurement, subjects were randomly assigned to either receive the supplement (300 kcal/d, 15 g protein/d) or no supplement., Results: Supplementation had no significant effect on any variable of pregnancy outcome, and except for fasting state decreases in leucine flux (125 ± 7.14 compared with 113 ± 5.06 μmol ⋅ kg(-1) ⋅ h(-1); P = 0.04) and nonoxidative disposal (110 ± 6.97 compared with 101 ± 3.69 μmol ⋅ kg(-1) ⋅ h(-1); P = 0.02) from trimesters 1 to 3, it had no effect on any other leucine kinetic variable or urea, glycine, and serine fluxes., Conclusion: We conclude that in Indian women with a low BMI, supplementation with energy and protein from week 12 of pregnancy to time of delivery does not improve pregnancy outcome, whole-body protein kinetics, or serine and glycine fluxes., (© 2016 American Society for Nutrition.)

Prevention of malnutrition in infants and children is multifaceted and requires the following: access to and intake of nutritious food starting at birth with exclusive breastfeeding for the first 6 mo of life, continued breastfeeding in combination with complementary foods from 6-24 mo of age, access to clean drinking water and sanitation, and access to preventive and curative health care (including prenatal). Nutrient-dense complementary foods can improve nutritional status and have long-term benefits; however, in a review of plant-based complementary foods in developing countries, most of them failed to meet many micronutrient requirements. There is need to provide other cost-effective alternatives to increase the quality of the diet during the complementary feeding stage of the lifecycle. This paper provides an overview of the development, testing, efficacy and effectiveness of the delivery of KOKO Plus on the growth and nutritional status of infants 6-24 mo of age.

Over a third of all deaths of children under the age of five are linked to undernutrition. At a 90% coverage level, a core group of ten interventions inclusive of infant and young child nutrition could save one million lives of children under 5 y of age (15% of all deaths) (Lancet 2013). The infant and young child nutrition package alone could save over 220,000 lives in children under 5 y of age. High quality proteins (e.g. milk) in complementary, supplementary and rehabilitation food products have been found to be effective for good growth. Individual amino acids such as lysine and arginine have been found to be factors linked to growth hormone release in young children via the somatotropic axis and high intakes are inversely associated with fat mass index in pre-pubertal lean girls. Protein intake in early life is positively associated with height and weight at 10 y of age. This paper will focus on examining the role of protein and amino acids in infant and young child nutrition by examining protein and amino acid needs in early life and the subsequent relationship with stunting.

Background: Evidence suggests that in women with a normal to high body mass index (BMI; in kg/m(2)), the extra amino acids needed during pregnancy are met through reduced oxidation. It is not known whether a woman with a low BMI can make this adaptation successfully., Objective: The objective was to measure and compare leucine kinetic parameters and alanine-nitrogen, glutamine amide-nitrogen, and glycine and cysteine fluxes in Indian women with a low and normal BMI in early and midpregnancy., Design: Fasted- and fed-state kinetics were measured by infusing 1-[(13)C]leucine, [(2)H(2)]cysteine, [(2)H(2)]glycine, [5-(15)N]glutamine, and [(15)N]alanine in groups of 10 women with a low BMI (<18.5) and 10 women with a normal BMI (18.5-25) in the first and second trimesters of pregnancy., Results: Leucine, glutamine, glycine, and cysteine fluxes were faster in women with a low BMI in both trimesters, but there was no difference in alanine flux between groups. This difference was explained in the first trimester by a higher proportion of fat-free mass in low-BMI women. Leucine oxidation and percentage of dietary leucine oxidized were higher in low-BMI women in both trimesters, but nonoxidative disposal was not different between groups., Conclusions: Although they use dietary protein less efficiently, low-BMI women maintain net protein synthesis at the same rate as do normal-BMI women and produce similar quantities of labile nitrogen for the de novo synthesis of other dispensable amino acids such as glycine and cysteine. The extra amino acids required for increased maternal protein synthesis during pregnancy are provided by an overall decrease in amino acid catabolism in women with normal or low BMI.

We recently reported the lysine requirement of school-aged children living in Canada consuming a mixed diet to be 35 mg.kg(-1).d(-1). Because the majority of children in the world live on cereal-based diets in developing countries, we measured the daily lysine requirement in healthy children living in India and consuming a mostly cereal-based diet. Our objective in this study was to determine the lysine requirement in healthy, school-aged children in the developing world by using the indicator amino acid oxidation method with l-[1-(13)C] phenylalanine. Six healthy, school-aged children consumed 7 levels of lysine (5, 15, 25, 35, 50, 65, and 80 mg.kg(-1).d(-1)) each in a random order along with an amino acid mixture providing energy and protein intakes of 1.7 x resting energy expenditure and 1.5 g.kg(-1).d(-1), respectively. The mean lysine requirement was determined by applying a 2-phase linear regression crossover analysis on tracer oxidation (F(13)CO(2)) data, which identified a breakpoint (requirement) at minimal F(13)CO(2) in response to the graded lysine intakes. The mean lysine requirements with the upper 95% CI for children were determined to be 33.5 and 46.6 mg.kg(-1).d(-1), respectively, by breakpoint analysis of the F(13)CO(2) data. The mean lysine requirements of Indian children were almost identical to that of Canadian children (35 mg.kg(-1).d(-1)). There is no evidence for any adaptation in lysine requirements in children from developing countries such as India.

Nutrition and infection interact with each other in a synergistic vicious cycle, leading to an adverse nutritional status and increased susceptibility to infection. Infectious episodes result in hypermetabolism and a negative nitrogen balance which is modulated by hormones, cytokines and other pro-inflammatory mediators, and is compounded by a reduced food intake. The extent of the negative nitrogen balance varies with the type of infection and its duration; however, it is reasonable to suggest that the loss of body protein could be minimized by the provision of dietary nitrogen, although anorexia will limit this. Further, distinctions need to be made about the provision of nutrients or protein during the catabolic and anabolic or recovery phase of the infection, since the capacity of the body to retain protein is enhanced in the anabolic recovery phase. Meeting the increased requirement for protein (and other nutrients) in infection does not imply a complete therapeutic strategy. Infections need to be treated appropriately, with nutrition as an adjunct to the treatment. Prior undernutrition could also impair the body's response to infection, although the weight of the evidence would suggest that this happens more particularly in oedematous undernutrition. In general, the amount of extra protein that would appear to be needed is of the order of 20-25 per cent of the recommended intake, for most infections. In acute infections, this is particularly relevant during the convalescence period. Community trials have suggested that lysine supplementation to the level required for normal daily nutriture, in predominantly wheat eating or potentially lysine deficient communities, improves immune function among other functional nutritional parameters; however, there is as yet insufficient evidence to suggest a specific requirement for amino acids in infections over and above the normal daily requirement as based on recent evidence. Some clinical studies that have showed benefits with specific amino acids through selected clinical outcomes, however, these do not provide enough evidence for a firm recommendation.

Background: We previously questioned the validity of the 1985 FAO/WHO/UNU upper requirement value for threonine (7 mg x kg(-1) x d(-1)) and proposed a tentative mean requirement of 15 mg x kg(-1) x d(-1)., Objective: In this study we used a 24-h indicator amino acid oxidation and balance technique, with [1-(13)C]leucine as the indicator amino acid, to assess threonine adequacy at 6 test intakes (7, 11, 15, 19, 22, and 27 mg x kg(-1) x d(-1)) with a 6-d dietary adaptation phase in healthy, well-nourished Indian men., Design: Sixteen men were randomly allocated to 3 of 6 test intakes and were studied after 6 d of adaptation to the experimental diets. Diets were based on an L-amino acid mixture in which the threonine content was varied. At 1800 on day 6, a 24-h intravenous [(13)C]leucine tracer infusion protocol was conducted to assess 24-h leucine oxidation and daily leucine balances., Results: Leucine balances differed significantly (P = 0.02) between the different intakes of threonine. Two-phase linear regression analysis from 12-h and 24-h leucine oxidation and 24-h leucine balance gave a breakpoint at a threonine intake of 15 mg x kg(-1) x d(-1), with 95% CIs ranging from 11 to 27 mg x kg(-1) x d(-1). There was no significant effect of threonine intake on 24-h leucine flux., Conclusion: The results of the 24-h indicator amino acid oxidation and balance experiments indicate that the current FAO/WHO/UNU threonine recommendation of 7 mg x kg(-1) x d(-1) is inadequate. A mean threonine intake of 15 mg x kg(-1) x d(-1) is sufficient to achieve the indicator (leucine) amino acid balance in healthy Indian men.

Background & aimsStunting in children is a comorbid condition in undernutrition that may be ameliorated by the provision of high-quality foods that provide protein and micronutrients. Addressing this problem in lower social economic environments requires, in part, affordable and scalable food-based solutions with efficacious food products. Towards this end, biochemical/metabolic indicators for fast-throughput screening of foods and their components are desired. A highly acceptable and economical micronutrient-fortified food product with different levels of legume protein was provided to stunted Indian children for one month, to determine change in their linear growth and explore associated biochemical, metabolomic and microbiome indicators.MethodsA randomized controlled pilot trial was conducted with 100 stunted children (6-10 years of age) to elucidate metabolic and microbiome-based biomarkers associated with linear growth. They were randomized into 4 groups receiving 6, 8, 10 or 12 g of legume-based protein for one month. Anthropometry, blood biochemistry, aminoacidomics, acylcarnitomics and fecal microbiome were measured before and after feeding.ResultsNo significant differences were observed between groups in height, height-for-age Z-score (HAZ) or BMI-for-age Z-score (BAZ); however, 38 serum metabolites were altered significantly (Bonferroni adjusted P  0.2, P = 0.02), while serine and ornithine (ρ  1.5) were significantly correlated with change in height.ConclusionsIn this pilot trial, a number of fasting serum metabolomic and fecal microbiome signatures were associated with linear growth after a short-term dietary intervention. The alterations of these markers should be validated in long-term dietary intervention trials as potential screening indicators towards the development of food products that favor growth. This trial was registered at www.ctri.nic.in as CTRI/2016/12/007564.

Background In young children, associations between linear growth faltering, environmental enteric dysfunction (EED), and the plasma kynurenine (Kyn)/tryptophan (Trp) ratio (KTR) have led to the proposal that higher Trp catabolism in response to intestinal/systemic inflammation limits Trp availability for protein synthesis, resulting in impaired growth. Objectives We sought to estimate the Trp oxidation rate and the Trp conversion rate to Kyn in young children with and without EED. Methods Children aged 18–24 mo, from urban slums, were assigned to EED (n  = 19) or no-EED (n  = 26) groups on the basis of a urinary lactulose/rhamnose ratio (LRR) cutoff based on mean + 2 SDs of LRR (≥0.068) in normal age- and sex-matched, high–socioeconomic status children. Plasma KTR and fecal biomarkers of EED were measured. Trp oxidation in the fed state was measured using 13C1-Trp in an oral plateau feeding protocol. Results The median (quartile 1, quartile 3) fasted KTR was 0.089 (0.066, 0.110) in children with EED compared with 0.070 (0.050, 0.093) in children with no EED (P  = 0.077). However, there was no difference in fed-state Trp oxidation [median (quartile 1, quartile 3) 3.1 (1.3, 5.8) compared with 3.9 (1.8, 6.0) µmol/kg FFM/h, respectively, P  = 0.617] or Trp availability for protein synthesis [42.6 (36.5, 45.7) compared with 42.5 (37.9, 46.9) µmol/kg FFM/h, respectively, P  = 0.868] between the groups. In contrast, the median (quartile 1, quartile 3) fractional synthesis rates of Kyn [12.5 (5.4, 20.0) compared with 21.3 (16.1, 24.7) %pool/h, P  = 0.005] and the fraction of Ala derived from Trp [0.007 (0.005, 0.015) compared with 0.012 (0.008, 0.018), P  = 0.037], respectively, in the plasma compartment were significantly slower in the EED group. Fecal biomarkers of EED did not differ between the groups. Conclusions The static plasma KTR value is not a good indicator of the dynamic Trp flux down its oxidative pathway. In a poor sanitary environment, children without EED actually have a faster Kyn synthesis rate, which might be beneficial, because of the cytoprotective and anti-inflammatory functions of downstream metabolites. This study was registered in the Clinical Trials Registry of India as CTRI/2017/02/007921. [ABSTRACT FROM AUTHOR]

Background: Lysine rich foods such as milk and legumes serve as important food additions to the lysine deficient cereal-based diets of vegetarian populations in low- and middle-income countries (LMICs) to alleviate the risk of quality corrected dietary protein inadequacy. Dietary protein quality can be determined by estimating the metabolic availability (MA) of lysine.Objectives: The study aimed to estimate the MA of lysine in spray-dried cow milk powder (SMP), heat-treated spray-dried cow milk powder (HSMP), and a habitually consumed cereal-legume based vegetarian meal (VM), using the indicator amino acid oxidation (IAAO) slope-ratio method.Methods: The MA of lysine in SMP, HSMP, and VM was estimated in 7 healthy young men aged 19-24 y with BMI of 21.5 ± 0.5 kg/m2 in a repeated measures design. The IAAO response slopes with 2 graded lysine intakes (10.5 and 15.0 mg·kg-1·d-1) from the SMP and VM were compared with the response slope generated with 3 graded crystalline lysine intakes (6.0, 10.5, and 15.0 mg·kg-1·d-1) at the subrequirement level. To produce HSMP, pasteurized cow milk was heat treated and spray dried. The MA of lysine in HSMP was tested at a single level of lysine intake (15 mg·kg-1·d-1). A total of 8 IAAO experiments were conducted on each participant in randomized order. The IAAO slopes were estimated using a linear mixed-effect regression model.Results: The MA of lysine in SMP, HSMP, and VM was 91.9%, 69.9%, and 86.6% respectively.Conclusions: Heat treatment reduced the MA of lysine by 22% in HSMP compared with SMP in healthy Indian adults. The lysine MA estimates can be used to optimize lysine limited cereal-based diets, with the addition of appropriately processed legumes and milk powder, to meet the protein requirement. This trial was registered at Clinical Trials Registry of India (http://ctri.nic.in) as CTRI/2019/08/020568. [ABSTRACT FROM AUTHOR]

The FAO of the UN convened an Expert Working Group meeting to provide recommendations related to protein quality evaluation of Follow-up Formula for Young Children (FUF-YC) and Ready-to-Use Therapeutic Foods (RUTFs). The protein and amino acid (AA) scoring patterns for the target age groups were defined and recommendations provided on the use of currently available protein and indispensable AA digestibility data. For FUF-YC, an age category of 1-2.9 y was identified, and a matching protein requirement of 0.86 g · kg-1 · d-1 with corresponding AA requirements were recommended. For RUTF, the protein requirement recommended was 2.82 g · kg-1 · d-1, to achieve a catch-up weight gain of 10 g · kg-1 · d-1 in children recovering from severe acute malnutrition. The AA requirements were factorially derived based on the adult protein requirement for maintenance and tissue AA composition. A flowchart was proposed for the best available methods to estimate digestibility coefficients (of either protein or AAs), in the following order: human, growing pig, and rat true ileal AA digestibility values. Where this is not possible, fecal protein digestibility values should be used. The Expert Working Group recommends the use of the Protein Digestibility Corrected Amino Acid Score (PDCAAS), with existing protein digestibility values, or the Digestible Indispensable Amino Acid Score provided that individual AA digestibility values are available for protein quality evaluation using the latter score. The Group also recommends the use of ileal digestibility of protein or of AAs for plant-based protein sources, recognizing the possible effects of antinutritional factors and impaired gut function. A PDCAAS score of ≥90% can be considered adequate for these formulations, whereas with a score <90%, the quantity of protein should be increased to meet the requirements. Regardless of the protein quality score, the ability of formulations to support growth in the target population should be evaluated. Future research recommendations are also proposed based on the knowledge gaps identified. [ABSTRACT FROM AUTHOR]

Background: Recent evaluations of the risk of dietary protein defi- ciency have indicated that protein digestibility may be a key limiting factor in the provision of indispensable amino acids (IAAs), particularly for vulnerable populations living in challenging environments where intestinal dysfunction may exist. Since the digestion of protein occurs only in the small intestine, and the metabolic activity of colonic bacteria confounds measurements at the fecal level, there is a need to develop noninvasive protein digestibility measurements at the ileal level. Objective: We used a dual-tracer method with stable isotopes to characterize the digestibility of uniformly labeled [13C]-spirulina protein as a standard protein, in comparison to a mixture of 2Hlabeled crystalline amino acids, and then demonstrated the use of this standard protein to measure the digestibility of selected legumes (chick pea and mung bean) through the use of proteins that were intrinsically labeled with ²H. Design: The digestibility of uniformly labeled [13C]-spirulina was first measured in 6 healthy volunteers (3 males and 3 females) by feeding it along with a standard mixture of ²H-labeled amino acids, in a dual-tracer, plateau-fed test meal approach. Next, intrinsically labeled legume protein digestibility was studied with a similar dualtracer approach, with uniformly labeled [13C]-spirulina as the standard, when processed differently before consumption. Results: The average digestibility of IAA in spirulina protein was 85.2%. The average IAA digestibility of intrinsically ²H-labeled chick pea and mung bean proteinwas 56.6% and 57.7%, respectively. Dehulling of mung bean before ingestion increased the average IAA digestibility by 9.9% in comparison to whole mung bean digestibility. Conclusions: An innovative, minimally invasive "dual-stableisotope" method was developed to measure protein digestibility, in which the ingestion of an intrinsically ²H-labeled test protein along with a 13C-labeled standard protein of known digestibility allows for an accurate measure of digestion and absorption of the intrinsically labeled protein. This minimally invasive method is critical to redefining protein quality and will aid in revisiting human protein requirements in different settings and in vulnerable populations. This trial was registered at Clinical Trials Registry--India as CTRI/2017/11/010468. [ABSTRACT FROM AUTHOR]

Background: Methionine loading test (MLT) has been used primarily to identify defects in transsulfuration of homocysteine in cystathionine beta synthase deficiency. It may not be as useful to evaluate remethylation pathway, in vitamin B-12 and folate deficiencies. Objective: We used tracer isotope labelled MLT to interrogate transsulfuration and remethylation independently in vitamin B-12 deficiency. Design: We studied vitamin B-12 deficient women with a tracer labelled MLT before and eleven months after treatment with vitamin B-12. The fractional contribution of [13C]homocysteine to breath CO2 was used as a measure of transsulfuration, and difference in the intracellular enrichment of [13C]methionine and that of [C2H3]methionine as a measure of remethylation of homocysteine. Combined pre- and post-treatment results were analyzed to investigate the association between plasma vitamin B-12 concentrations and measures of homocysteine metabolism. Results: The subjects were 17 years old, with a BMI of 19.4 kg/m2. Treatment with vitamin B-12, 2µg/day increased plasma B-12 from 93 (78.7, 106.2) [median (25th, 75th centiles)] to 161.5 (125.5, 226.2) pmol/L; 44% were below <150pmol/L after treatment. Fasting homocysteine concentration was significantly lower and that of cysteine higher in subjects with B-12 levels >150pmol/L. The tracer estimated transsulfuration of homocysteine was lower and remethylation higher with B-12 levels >150pmol/L when compared with those <150pmol/L. Conclusions: The tracer labelled MLT in combination with fasting parameters is a robust way to estimate parameters of methionine metabolism and can be used in the field where prime-constant rate infusion studies cannot be done efficiently. [ABSTRACT FROM AUTHOR]

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 a 13C-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. [ABSTRACT FROM AUTHOR]

There is now an expanding body of evidence to recommend, in the case of adult humans, the use of revised indispensable amino acid requirement values; these are ∼2 to 3 times higher than the current international recommendations. The earlier methodologies for determining amino acid requirements, based on nitrogen balance, were criticized because of their design and the associated high energy intakes. The 1985 World Health Organization/Food & Agriculture Organization!United Nations University requirement for leucine has been demonstrated to be too low by short- and long-term (24-h) tracer-derived estimates of leucine oxidation and balance. The best values for leucine requirements come from 24-h direct amino acid oxidation (DAAO) and direct amino acid balance (DAAB) studies. Finally, we also collated all available data from studies on fed-state leucine oxidation with an adequate dietary adaptation period to assess the inflection on the leucine oxidation-leucine intake curve. The mean requirements for leucine, valine, and isoleucine are likely to be 40, 17-25, and 19 mg· kg-1· d-1, respectively. This adds up to a total of ∼84mg·kg-1· d-1, which is much lower than the lowest estimate of the total BCAA requirement of ∼110 mg·kg-1·d-1 made by the short-term indicator amino acid oxidation (IAAO) method, which determined the BCAA requirement from the pattern of oxidation of an indicator amino acid (phenylyalanine) at different levels of BCAA intake. An additional estimate of the leucine requirement was also made by a meta-analysis of all available 24-h DAAO!DAAB data from different studies. This resulted in a higher value for the leucine requirement than that obtained by the specific studies that utilized the 24-h DAAO/DAAB approach; however, even adding this value to the total BCAA requirement does not account for the difference in the total BCAA requirement estimates and the summed individual BCAA estimates. [ABSTRACT FROM AUTHOR]

Background: In an earlier study, using a modification of the indicator amino acid oxidation approach, we concluded that the 1985 FAO/WHO/UNU-proposed lysine requirement of 12 mg ⋅ kg-1 ⋅ d-1 is likely inadequate to maintain body amino acid homeostasis in apparently healthy south Asian subjects and that our proposed requirement of 30 mg ⋅ kg-1 ⋅ d-1 is more appropriate. Objective: We assessed the lysine requirement in a similar population by using 4 test lysine intakes (12, 20, 28, and 36 mg ⋅kg-1 ⋅ d-1) with an indicator amino acid balance approach. Design: Sixteen healthy male Indians were studied during each of 2 randomly assigned 8-d L-amino acid diets that supplied either 12 and 28 or 20 and 36 mg lysine. At 1800 on day 8, a 24-h intravenous [13C]leucine tracer-infusion protocol was conducted to assess leucine oxidation and daily leucine balance at each lysine intake. Results: Mean 24-h leucine oxidation rates decreased significantly (P = 0.005) across different lysine intakes and were 104.1, 97.8, 87.3, and 87.3 mg ⋅ kg-1 ⋅ d-1 at intakes of 12, 20, 28, and 36 mg ⋅ kg-1 ⋅ d-1, respectively; mean 24-h leucine balances were 3.3, 9.1, 19.7, and 20.7 mg ⋅ kg-1 ⋅ d-1, respectively (P = 0.015, mixed-model analysis of variance). Oxidation and balances differed significantly between the lower and higher lysine intakes but were not significantly different between the 12- and 20-mg and 28- and 36-mg test intakes. Two-phase regression analysis indicated a mean breakpoint at 29 mg lysine ⋅ kg-1 ⋅ d-1 in the relation between lysine intake and leucine oxidation or balance. Conclusion: We propose a mean lysine requirement of 30 mg ⋅ kg-1 ⋅ d-1 for healthy Indian adults, which is the same amount we proposed previously for Western populations. [ABSTRACT FROM AUTHOR]

The factorial approach is used to measure the dietary indispensable amino acid (IAA) requirements in children, although recent measurements based on the indicator amino acid oxidation (IAAO) method have begun to generate more direct evidence. Difficulties with the factorial method are that it depends on accurate estimates of the maintenance protein requirement, as well as of protein deposition during growth. Also, a value for the efficiency of utilizing dietary protein for deposition has to be selected, based on published Nitrogen (N) balance studies. In the recent 2007 WHO/FAO/UNU report, the amino acid requirement pattern for infants was taken to be similar to the amino acid composition of breast milk. For pre-school and older children, the factorial method gave values for the scoring pattern of protein that were fairly close to the earlier 1985 WHO/FAO/UNU report for children, since growth progressively became a smaller component of the factorial calculation as age progressed. However, given that there are several assumptions in the derivation of factorial estimates, direct experimental measurements of the amino acid requirement are desirable. The IAAO method, as it is non-invasive, as made it possible to measure the IAA requirements in children. Over the last decade, some of the IAA requirements have been determined by using the IAAO method in healthy school age children; however, the data on IAA requirements in developing country populations are still being conducted. In the elderly, there are not enough data to make a separate recommendation for IAA requirements from that of adults. [ABSTRACT FROM PUBLISHER]

Indian diets derive almost 60 % of their protein from cereals with relatively low digestibility and quality. There have been several surveys of diets and protein intakes in India by the National Nutrition Monitoring Board (NNMB) over the last 25 years, in urban and rural, as well as in slum dwellers and tribal populations. Data of disadvantaged populations from slums, tribals and sedentary rural Indian populations show that the protein intake (mainly from cereals) is about 1 gm/kg/day. However, the protein intake looks less promising in terms of the protein digestibility corrected amino acid score (PDCAAS), using lysine as the first limiting amino acid, where all populations, particularly rural and tribal, appear to have an inadequate quality to their protein intake. The protein: energy (PE) ratio is a measure of dietary quality, and has been used in the 2007 WHO/FAO/UNU report to define reference requirement values with which the adequacy of diets can be evaluated in terms of a protein quality corrected PE ratio. It is likely that about one third of this sedentary rural population is at risk of not meeting their requirements. These levels of risk of deficiency are in a population with relatively low BMI populations, whose diets are also inadequate in fruits and vegetables. Therefore, while the burden of enhancing the quality of protein intake in rural India exists, the quality of the diet, in general, represents a challenge that must be met. [ABSTRACT FROM PUBLISHER]