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.

Kashyap S ，Bellam HRR ，Preston T ，Devi S ，Kurpad AV ... -  《-》

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

Kashyap S ，Shivakumar N ，Varkey A ，Duraisamy R ，Thomas T ，Preston T ，Devi S ，Kurpad AV ... -  《-》

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.

Bandyopadhyay S ，Kashyap S ，Calvez J ，Devi S ，Azzout-Marniche D ，Tomé D ，Kurpad AV ，Gaudichon C ... -  《-》

Values for the Digestibility of Pea Protein Isolate or Casein Amino Acids Determined using the Dual Isotope Method Are Not Similar to Those Derived with the Standard Ileal Balance Method in Healthy Volunteers.

Guillin FM ，Gaudichon C ，Guérin-Deremaux L ，Lefranc-Millot C ，Airinei G ，Khodorova N ，Benamouzig R ，Pomport PH ，Martin J ，Calvez J ... -  《-》

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

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. 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. 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. 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. 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).

Kashyap S ，Varkey A ，Shivakumar N ，Devi S ，Reddy B H R ，Thomas T ，Preston T ，Sreeman S ，Kurpad AV ... -  《-》