Estimation of exocrine pancreatic insufficiency in children with acute pancreatitis using the 13C mixed triglyceride breath test.

BACKGROUND: /Objective: The extent of exocrine pancreatic insufficiency (EPI) in the paediatric population with acute pancreatitis (AP) is unknown. The primary objective was to use a 6 h stable-isotope breath test to determine the prevalence of EPI in children with AP. The secondary objective was to determine the diagnostic ability of a 4 h abbreviated breath test in the detection of EPI. METHODS: 13C-mixed triglyceride (MTG) breath test was used to measure fat digestibility in 12 children with AP and 12 normal children. EPI was diagnosed based on a cumulative dose percentage recovery (cPDR) cut-off value < 26.8% present in literature. To reduce the test burden, the diagnostic accuracy of an abbreviated 4 h test was evaluated, using a cPDR cut-off that was the 2.5th percentile of its distribution in control children. RESULTS: The cPDR of cases was significantly lower than that of controls (27.71 ± 7.88% vs 36.37 ± 4.70%, p = 0.005). The cPDR during acute illness was not significantly different to that at 1 month follow up (24.69 ± 6.83% vs 26.98 ± 11.10%, p = 0.52). The 4 h and 6 h breath test results correlated strongly (r = 0.93, p < 0.001) with each other. The new 4 h test had 87.5% sensitivity and 93.8% specificity for detecting EPI. CONCLUSION: Two-thirds (66.7%) of this sample of children with AP had EPI during admission, which persisted at 1 month follow up. The 4 h abbreviated 13C-MTG breath test has good diagnostic ability to detect EPI in children and may improve its clinical utility in this age group.

The Systemic Availability of Indispensable Amino Acids from Orally Ingested Algal and Legume Protein in Young Children at Risk of Environmental Enteric Dysfunction.

BACKGROUND: The digestion and absorption of ingested protein may be reduced in children with environmental enteric dysfunction (EED), reducing systemic amino acid availability for protein synthesis with resultant growth faltering. This has not been directly measured in children with EED and associated growth faltering. OBJECTIVES: To evaluate the systemic availability of algal (spirulina) and legume (mung bean) indispensable amino acids (IAAs) in children with EED. METHODS: Indian children (18-24 mo) from urban slums were assigned to EED (n = 24) or no-EED (control, n = 17) groups based on the lactulose rhamnose test, where the lactulose rhamnose ratio cutoff for diagnosing EED (≥0.068) was the mean + 2SD of its distribution in healthy, age-, and sex-matched children of high socioeconomic status. Fecal biomarkers of EED were also measured. Systemic IAA availability was calculated from the plasma: meal IAA enrichment ratio for each protein. True ileal mung bean IAA digestibility was measured by the dual isotope tracer method using spirulina protein as reference. Co-administration of free 13C6-phenylalanine allowed for estimating true ileal phenylalanine digestibility of both proteins, and a phenylalanine absorption index. RESULTS: There was no significant difference (independent t-test) in the systemic IAA availability from spirulina or mung bean protein between EED and no-EED groups. There was no between-group difference in true ileal phenylalanine digestibility and its absorption index, or in mung bean IAA digestibility. CONCLUSIONS: The systemic IAA availability of algal and legume protein, or the latter's IAA/phenylalanine digestibility, is not significantly reduced in children with EED and does not correlate with linear growth. This study was registered in Clinical Trials Registry of India (CTRI) with registration number: CTRI/2017/02/007921.

Measurement of True Indispensable Amino Acid Digestibility by the Dual Isotope Tracer Technique: A Methodological Review.

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 (2H or 15N-labeled) test protein and (13C-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-15N and α-2H 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 15N or 2H 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.

The effect of l-arginine supplementation on amelioration of oxygen support in severe COVID-19 pneumonia.

BACKGROUND & AIMS: l-Arginine (l-Arg) has been shown to help reduce respiratory support requirements in coronavirus disease 2019 (COVID-19), in an Italian study. We investigated the effect of l-Arg supplementation on the reduction in respiratory support for patients with severe COVID-19 pneumonia in an Indian population. METHODS: A parallel-group, triple-blinded, randomized controlled trial (RCT) was conducted on patients with severe COVID-19 pneumonia on oxygen (O2) support. Patients received either 3 g of oral l-Arg or placebo, daily under supervision, until they were off O2 support, or for a maximum of 10 days, whichever was earlier. The primary outcome was cessation in O2 support. Other outcomes were time to cessation of O2 support, duration of hospitalization, and incidence of adverse thrombotic events. RESULTS: We did an intention-to-treat analysis on 74 patients who were randomized into l-Arg (n = 38) or placebo group (n = 36). There were no significant differences between the two groups in the outcomes. At end of the study, 28 patients (73.6%) in l-Arg and 26 patients (72.2%) in the placebo group were weaned off oxygen support. The median number of days to the cessation of O2 support estimated using Kaplan Meir survival analysis, was 3 days in the l-Arg group (95% confidence interval [CI]: 1.2, 4.7) and 5 days in the placebo group (95% CI, 4.1,5.8); P = 0.27. CONCLUSION: In this group of patients with severe COVID-19 pneumonia, l-Arg supplementation did not show any significant difference in outcomes when compared to placebo supplementation.

Tryptophan oxidation in young children with environmental enteric dysfunction classified by the lactulose rhamnose ratio.

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.

Evaluation of Protein Quality in Humans and Insights on Stable Isotope Approaches to Measure Digestibility - A Review.

The recent Food and Agricultural Organization/World Health Organization/United Nations University expert consultations on protein requirements and quality have emphasized the need for the new Digestible Indispensable Amino Acid Score (DIAAS), as a measure of protein quality. This requires human measurements of the true ileal digestibility of individual indispensable amino acids (IAAs) until the end of the small intestine. Digestibility is measured using standard oro-ileal balance methods, which can only be achieved by an invasive naso-ileal intubation in healthy participants or fistulation at the terminal ileum. Significant efforts have been made over the last 2 decades to develop noninvasive or minimally invasive methods to measure IAA digestibility in humans. The application of intrinsically labeled (with stable isotopes like 13C, 15N, and 2H) dietary proteins has helped in circumventing the invasive oro-ileal balance techniques and allowed the differentiation between endogenous and exogenous protein. The noninvasive indicator amino acid oxidation (IAAO) technique, which is routinely employed to measure IAA requirements, has been modified to estimate metabolic availability (a sum of digestibility and utilization) of IAA in foods, but provides an estimate for a single IAA at a time and is burdensome for participants. The recently developed minimally invasive dual isotope tracer method measures small intestinal digestibility of multiple amino acids at once and is suitable for use in vulnerable groups and disease conditions. However, it remains to be validated against standard oro-ileal balance techniques. This review critically evaluates and compares the currently available stable isotope-based protein quality evaluation methods with a focus on the digestibility and metabolic availability measurements in humans. In view of building a reliable DIAAS database of various protein sources and subsequently supporting protein content claims in food labeling, a re-evaluation and harmonization of the available methods are necessary.

Goat milk protein digestibility in relation to intestinal function.

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.

True ileal digestibility of legumes determined by dual-isotope tracer method in Indian adults.

BACKGROUND: Good-quality plant protein sources are important for protein adequacy in a balanced diet. Legumes are known to be a source of good quality plant protein, but the true ileal digestibility of indispensable amino acids (IAAs) of commonly consumed legumes is not known in humans. OBJECTIVES: In this study we measured the true ileal IAA digestibility of 2H-intrinsically labeled chickpea, yellow pea, and mung bean (hulled and dehulled) protein, using the dual-isotope tracer technique referenced to a standard protein ([U-13C] spirulina). The study also aimed to validate the use of [U-13C] spirulina as a reference protein in this method. METHODS: 2H-intrinsically labeled legumes, obtained by watering plants with deuterium oxide (2H2O), were administered in a plateau feeding method to healthy Indian adults to measure their true ileal IAA digestibility with the dual-isotope tracer technique, using [U-13C] spirulina protein or a 13C-algal IAA mixture as the standard. RESULT: The true ileal IAA digestibilities (mean ± SD) of chickpea, yellow pea, and mung bean were 74.6 ± 0.8%, 71.6 ± 1.3%, and 63.2 ± 1.5%, respectively. The true mean ileal IAA digestibility of mung bean when referenced to [U-13C] spirulina protein or a 13C-algal IAA mixture did not differ significantly (63.2 ± 1.5% versus 64.0 ± 2.4%, P > 0.05). The true ileal IAA digestibility of mung bean improved to 70.9 ± 2.1% after dehulling. CONCLUSIONS: The true mean ileal IAA digestibility of legumes in healthy Indian adults was lower than expected. Traditional processing techniques such as dehulling improve protein digestibility by about 8%. This study was registered in the Clinical Trials Registry of India (CTRI): CTRI/2017/11/010468 (http://ctri.nic.in, accessed on 28/03/2019).

Co-ingestion of Black Tea Reduces the Indispensable Amino Acid Digestibility of Hens' Egg in Indian Adults.

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.

Protein-quality evaluation of complementary foods in Indian children.

BACKGROUND: The types of food in complementary feeding of infants and young children are important for growth and development. Food protein quality, as measured by the Digestible Indispensable Amino Acid Score (DIAAS), requires the determination of true ileal digestibility of indispensable amino acids (IAAs) in children. OBJECTIVES: First, the aim of this study was to measure the true ileal IAA digestibility of 4 (rice, finger millet, mung bean, and hen egg) commonly consumed complementary foods in children aged <2 y using the dual-isotope tracer method. Second, we calculated the DIAAS of complementary feeding diets and their relation to stunting in a representative Indian rural population. DESIGN: Rice, finger millet, and mung bean were intrinsically labeled with deuterium oxide (2H2O), whereas egg was labeled through oral dosing of hens with a uniformly 2H-labeled amino acid mixture. True ileal IAA digestibility was determined by the dual-isotope tracer technique. The DIAAS of complementary food protein was calculated in children aged 1-3 y from a nationally representative survey to evaluate its relation with stunting. RESULTS: True ileal IAA digestibility was lowest in mung bean (65.2% ± 7.1%), followed by finger millet (68.4 %± 5.3%) and rice (78.5% ± 3.5%), and was highest for egg (87.4% ± 4.0%). There was a significant inverse correlation of complementary food DIAAS with stunting in survey data (r = -0.66, P = 0.044). The addition of egg or milk to nationally representative complementary diets theoretically improved the DIAAS from 80 to 100. CONCLUSIONS: The true ileal IAA digestibility of 4 foods commonly consumed in complementary diets showed that the DIAAS was associated with stunting and reinforces the importance of including animal source food (ASF) in diets to improve growth. This trial was registered at http://ctri.nic.in/clinicaltrials/login.php as CTRI/2017/02/007921.

Ileal digestibility of intrinsically labeled hen's egg and meat protein determined with the dual stable isotope tracer method in Indian adults.

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.