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.

BACKGROUND: The measurement of ileal amino acid (AA) digestibility is invasive and inappropriate when applied to vulnerable populations. The dual isotope method has been developed over the past 5 y as an alternative method. OBJECTIVE: The aim of this work was to compare the indispensable amino acid (IAA) digestibility values of 2 different proteins obtained using the dual isotope and the standard ileal balance methods in the same subjects. METHODS: Fifteen healthy adults completed the study. Over 4 h, they ingested 9 successive portions of mashed potatoes containing the test protein (pea protein or casein) labeled intrinsically with 15N and 2H, and a 13C-free AA mixture as a reference for the dual isotope method. Plasma was sampled regularly over the 8-h postprandial period, whereas the ileal digesta was collected continuously via a naso-ileal tube. Isotopic enrichments (15N and 13C) were measured in the digesta for the direct determination of ileal IAA digestibility, whereas plasma enrichments (2H and 13C) were measured to determine IAA digestibility using the dual isotope method. RESULTS: The 4-h repeated meal procedure enabled the almost complete digestion of test proteins at 8 h and the attainment of a plasma isotopic plateau between 2.5 and 4 h. These conditions were necessary to perform the ileal balance and dual isotope methods simultaneously. For pea protein, the mean IAA digestibility was similar between the 2 methods, but significant differences (from 10% to 20%) were observed for individual IAA values. For casein, IAA digestibility was significantly lower with the dual isotope method for all the IAA analyzed. CONCLUSIONS: Under our experimental conditions, the degree of agreement between the dual isotope and ileal balance methods varied among AAs and depended on the protein source. Further research is needed to validate the dual isotope method. This study was registered at clinicaltrials.gov as NCT04072770.

Compared with Milk Protein, a Wheat and Pea Protein Blend Reduces High-Fat, High-Sucrose Induced Metabolic Dysregulations while Similarly Supporting Tissue Protein Anabolism in Rats.

BACKGROUND: Plant proteins (PPs) have been associated with better cardiovascular health than animal proteins (APs) in epidemiological studies. However, the underlying metabolic mechanisms remain mostly unknown. OBJECTIVES: Using a combination of cutting-edge isotopic methods, we aimed to better characterize the differences in protein and energy metabolisms induced by dietary protein sources (PP compared with AP) in a prudent or western dietary context. METHODS: Male Wistar rats (n = 44, 8 wk old) were fed for 4.5 mo with isoproteic diets differing in their protein isolate sources, either AP (100% milk) or PP (50%:50% pea: wheat) and being normal (NFS) or high (HFS) in sucrose (6% or 15% kcal) and saturated fat (7% or 20% kcal), respectively. We measured body weight and composition, hepatic enzyme activities and lipid content, and plasma metabolites. In the intestine, liver, adipose tissues, and skeletal muscles, we concomitantly assessed the extent of amino acid (AA) trafficking using a 15N natural abundance method, the rates of macronutrient routing to dispensable AA using a 13C natural abundance method, and the metabolic fluxes of protein synthesis (PS) and de novo lipogenesis using a 2H labeling method. Data were analyzed using ANOVA and Mixed models. RESULTS: At the whole-body level, PP limited HFS-induced insulin resistance (-27% in HOMA-IR between HFS groups, P < 0.05). In the liver, PP induced lower lipid content (-17%, P < 0.01) and de novo lipogenesis (-24%, P < 0.05). In the different tissues studied, PP induced higher AA transamination accompanied by higher routings of dietary carbohydrates and lipids toward dispensable AA synthesis by glycolysis and ß-oxidation, resulting in similar tissue PS and protein mass. CONCLUSIONS: In growing rats, compared with AP, a balanced blend of PP similarly supports protein anabolism while better limiting whole-body and tissue metabolic dysregulations through mechanisms related to their less optimal AA profile for direct channeling to PS.

Caecal digestibility as an approximation of ileal protein digestibility evaluation in rats.

The rat model can be used to assess ileal protein digestibility rapidly and in first intention, but no standardised method exists. Our objective was to compare methods to assess protein digestibility, depending on collection site (ileum/caecum) and use of a non-absorbable marker. A meal containing either casein, gluten or pea protein and chromium oxide as non-absorbable marker was given to male Wistar rats and the entire digestive content was collected 6 h later. Total chromium recovery was incomplete and variable, depending on protein source. We observed no significant difference in digestibility between the methods for any of the protein sources tested. Although none of the methods tested is optimal, our results suggest that caecal digestibility can be used as a proxy of ileal digestibility in rats without using a non-absorbable marker. This simple method makes it possible to evaluate protein digestibility of new alternative protein sources for human consumption.

Real ileal amino acid digestibility of pea protein compared to casein in healthy humans: a randomized trial.

BACKGROUND: It is necessary to propose plant alternatives to animal proteins that are of good nutritional quality. Pea is a good candidate owing to its high protein content and its well-balanced amino acid (AA) profile. OBJECTIVES: This study aimed to assess the real ileal AA and nitrogen digestibility (RIDAA and RIDN) of pea protein isolate as compared to milk casein in humans. It also aimed to evaluate their nutritional quality through calculation of the digestible indispensable amino acid score (DIAAS) and to determine the net postprandial protein utilization (NPPU). METHODS: Fifteen healthy volunteers were included in a randomized, single-blinded, 2-arm, parallel-design trial. They were equipped with a naso-ileal tube. They ingested the test meals, which consisted of 9 successive portions of mashed potatoes containing either pea protein or casein, intrinsically labeled with nitrogen 15. Ileal content, plasma, and urine samples were collected regularly over an 8-h postprandial period. RESULTS: The mean RIDAA values were 93.6% ± 2.9% for pea protein and 96.8% ± 1.0% for casein, with no difference between the sources (P = 0.22). Leucine, valine, lysine, and phenylalanine were significantly less digestible in pea than in casein. The RIDN values were 92.0% ± 2.7% and 94.0% ± 1.7% for pea protein and casein, respectively, and were not different (P = 0.11). The DIAAS was 1.00 for pea protein and 1.45 for casein. The NPPU was 71.6% ± 6.2% and 71.2% ± 4.9% for pea protein and casein, respectively (P = 0.88). CONCLUSIONS: Although some AAs are less digestible in pea protein than in casein, the real ileal digestibility and the NPPU were not different. The DIAAS of 1.00 obtained for pea protein demonstrated its ability to meet all AA requirements. This study shows the potential of pea isolate as a high-quality protein. This study was registered at clinicaltrials.gov as NCT04072770.

Co-ingestion of NUTRALYS® pea protein and a high-carbohydrate beverage influences the glycaemic, insulinaemic, glucose-dependent insulinotropic polypeptide (GIP) and glucagon-like peptide-1 (GLP-1) responses: preliminary results of a randomised controlled trial.

PURPOSE: Plant-based proteins may have the potential to improve glycaemic and gastrointestinal hormone responses to foods and beverages. The aim of this study was to investigate the effect of two doses of pea protein on postprandial glycaemic, insulinaemic, glucose-dependent insulinotropic polypeptide (GIP) and glucagon-like peptide-1 (GLP-1) response following a high-carbohydrate beverage intake in healthy individuals. METHODS: In a single-blind, randomised, controlled, repeat measure, crossover design trial, thirty-one participants were randomly assigned to ingest 50 g glucose (Control), 50 g glucose with 25 g pea protein (Test 1) and 50 g glucose with 50 g pea protein (Test 2) on three separate days. Capillary blood samples (blood glucose and plasma insulin measurements) and venous blood samples (GIP and GLP-1 concentrations) were taken before each test and at fixed intervals for 180 min. The data were compared using repeated-measures ANOVA or the Friedman test. RESULTS: Glucose incremental Area under the Curve (iAUC180) was significantly lower (p < 0.001) after Test 2 compared with Control (- 53%), after Test 1 compared with Control (- 31%) and after Test 2 compared with Test 1 (-32%). Insulin iAUC 180 was significantly higher (p < 0.001) for Test 1 (+ 28%) and Test 2 (+ 40%) compared with Control and for Test 2 (+ 17%) compared with Test 1 (p = 0.003). GIP and GLP-1 release showed no clear difference between Control and Pea protein drinks. CONCLUSION: The consumption of pea protein reduced postprandial glycaemia and stimulated insulin release in healthy adults with a dose-response effect, supporting its role in regulating glycaemic and insulinaemic responses.

Multi-criteria assessment of pea protein quality in rats: a comparison between casein, gluten and pea protein alone or supplemented with methionine.

The objective of this study was to assess the nutritional quality of pea protein isolate in rats and to evaluate the impact of methionine (Met) supplementation. Several protein diets were studied: pea protein, casein, gluten, pea protein-gluten combination and pea protein supplemented with Met. Study 1: Young male Wistar rats (n 8/group) were fed the test diets ad libitum for 28 d. The protein efficiency ratio (PER) was measured. Study 2: Adult male Wistar rats (n 9/group) were fed the test diets for 10 d. A protein-free diet group was used to determine endogenous losses of N. The rats were placed in metabolism cages for 3 d to assess N balance, true faecal N digestibility and to calculate the Protein Digestible-Corrected Amino Acid Score (PDCAAS). They were then given a calibrated meal and euthanised 6 h later for collection of digestive contents. The true caecal amino acid (AA) digestibility was determined, and the Digestible Indispensable Amino Acid Score (DIAAS) was calculated. Met supplementation increased the PER of pea protein (2·52 v. 1·14, P < 0·001) up to the PER of casein (2·55). Mean true caecal AA digestibility was 94 % for pea protein. The DIAAS was 0·88 for pea protein and 1·10 with Met supplementation, 1·29 for casein and 0·25 for gluten. Pea protein was highly digestible in rats under our experimental conditions, and Met supplementation enabled generation of a mixture that had a protein quality that was not different from that of casein.

Explorative Screening of Bioactivities Generated by Plant-Based Proteins after In Vitro Static Gastrointestinal Digestion.

The gastrointestinal digestion of food proteins can generate peptides with a wide range of biological activities. In this study, we screened various potential bioactivities generated by plant-based proteins. Whey protein as an animal protein reference, five grades of pea protein, two grades of wheat protein, and potato, fava bean, and oat proteins were submitted to in vitro SGID. They were then tested in vitro for several bioactivities including measures on: (1) energy homeostasis through their ability to modulate intestinal hormone secretion, to inhibit DPP-IV activity, and to interact with opioid receptors; (2) anti-hypertensive properties through their ability to inhibit ACE activity; (3) anti-inflammatory properties in Caco-2 cells; (4) antioxidant properties through their ability to inhibit production of reactive oxygen species (ROS). Protein intestinal digestions were able to stimulate intestinal hormone secretion by enteroendocrine cells, to inhibit DPP-IV and ACE activities, to bind opioid receptors, and surprisingly, to decrease production of ROS. Neither pro- nor anti-inflammatory effects have been highlighted and some proteins lost their pro-inflammatory potential after digestion. The best candidates were pea, potato, and fava bean proteins.

Effects of Whey and Pea Protein Supplementation on Post-Eccentric Exercise Muscle Damage: A Randomized Trial.

This randomized trial compared pea protein, whey protein, and water-only supplementation on muscle damage, inflammation, delayed onset of muscle soreness (DOMS), and physical fitness test performance during a 5-day period after a 90-min eccentric exercise bout in non-athletic non-obese males (n = 92, ages 18-55 years). The two protein sources (0.9 g protein/kg divided into three doses/day) were administered under double blind procedures. The eccentric exercise protocol induced significant muscle damage and soreness, and reduced bench press and 30-s Wingate performance. Whey protein supplementation significantly attenuated post-exercise blood levels for biomarkers of muscle damage compared to water-only, with large effect sizes for creatine kinase and myoglobin during the fourth and fifth days of recovery (Cohen's d > 0.80); pea protein versus water supplementation had an intermediate non-significant effect (Cohen's d < 0.50); and no significant differences between whey and pea protein were found. Whey and pea protein compared to water supplementation had no significant effects on post-exercise DOMS and the fitness tests. In conclusion, high intake of whey protein for 5 days after intensive eccentric exercise mitigated the efflux of muscle damage biomarkers, with the intake of pea protein having an intermediate effect.

Intrinsic immunomodulatory effects of low-digestible carbohydrates selectively extend their anti-inflammatory prebiotic potentials.

The beneficial effects of carbohydrate-derived fibers are mainly attributed to modulation of the microbiota, increased colonic fermentation, and the production of short-chain fatty acids. We studied the direct immune responses to alimentary fibers in in vitro and in vivo models. Firstly, we evaluated the immunomodulation induced by nine different types of low-digestible fibers on human peripheral blood mononuclear cells. None of the fibers tested induced cytokine production in baseline conditions. However, only one from all fibers almost completely inhibited the production of anti- and proinflammatory cytokines induced by bacteria. Secondly, the impact of short- (five days) and long-term (three weeks) oral treatments with selected fibers was assessed in the trinitrobenzene-sulfonic acid colitis model in mice. The immunosuppressive fiber significantly reduced levels of inflammatory markers over both treatment periods, whereas a nonimmunomodulatory fiber had no effect. The two fibers did not differ in terms of the observed fermentation products and colonic microbiota after three weeks of treatment, suggesting that the anti-inflammatory action was not related to prebiotic properties. Hence, we observed a direct effect of a specific fiber on the murine immune system. This intrinsic, fiber-dependent immunomodulatory potential may extend prebiotic-mediated protection in inflammatory bowel disease.

Pea proteins oral supplementation promotes muscle thickness gains during resistance training: a double-blind, randomized, Placebo-controlled clinical trial vs. Whey protein.

BACKGROUND: The effects of protein supplementation on muscle thickness and strength seem largely dependent on its composition. The current study aimed at comparing the impact of an oral supplementation with vegetable Pea protein (NUTRALYS®) vs. Whey protein and Placebo on biceps brachii muscle thickness and strength after a 12-week resistance training program. METHODS: One hundred and sixty one males, aged 18 to 35 years were enrolled in the study and underwent 12 weeks of resistance training on upper limb muscles. According to randomization, they were included in the Pea protein (n = 53), Whey protein (n = 54) or Placebo (n = 54) group. All had to take 25 g of the proteins or placebo twice a day during the 12-week training period. Tests were performed on biceps muscles at inclusion (D0), mid (D42) and post training (D84). Muscle thickness was evaluated using ultrasonography, and strength was measured on an isokinetic dynamometer. RESULTS: Results showed a significant time effect for biceps brachii muscle thickness (P < 0.0001). Thickness increased from 24.9 ± 3.8 mm to 26.9 ± 4.1 mm and 27.3 ± 4.4 mm at D0, D42 and D84, respectively, with only a trend toward significant differences between groups (P = 0.09). Performing a sensitivity study on the weakest participants (with regards to strength at inclusion), thickness increases were significantly different between groups (+20.2 ± 12.3%, +15.6 ± 13.5% and +8.6 ± 7.3% for Pea, Whey and Placebo, respectively; P < 0.05). Increases in thickness were significantly greater in the Pea group as compared to Placebo whereas there was no difference between Whey and the two other conditions. Muscle strength also increased with time with no statistical difference between groups. CONCLUSIONS: In addition to an appropriate training, the supplementation with pea protein promoted a greater increase of muscle thickness as compared to Placebo and especially for people starting or returning to a muscular strengthening. Since no difference was obtained between the two protein groups, vegetable pea proteins could be used as an alternative to Whey-based dietary products. TRIAL REGISTRATION: The present trial has been registered at ClinicalTrials.gov (NCT02128516).

Ethological comparison of the effects of a bovine alpha s1-casein tryptic hydrolysate and diazepam on the behaviour of rats in two models of anxiety.

A bovine alpha s1-casein tryptic hydrolysate was previously demonstrated to display an anxiolytic-like activity in the conditioned defensive burying and in the elevated plus-maze models when i.p. injected. The present study assessed the anxiolytic-like effects of this tryptic hydrolysate after an oral administration in rats faced to the same behavioural situations using diazepam as a reference. In a first experiment, the behavioural effects of the hydrolysate in the conditioned defensive burying test were investigated at doses ranging 5-50 mg/kg. The results showed that the minimal dose required to elicit an anxiolytic-like activity is 15 mg/kg. In a second experiment, the alpha s1-casein tryptic hydrolysate (15 mg/kg, p.o.) was demonstrated to display an anxiolytic-like activity similar to diazepam (3 mg/kg, p.o.) in the conditioned defensive burying test and the elevated plus-maze. However, the ethological analysis of behaviour indicated that this hydrolysate has a different activity compared to diazepam. While diazepam induced a disinhibition state in rats, possibly related to the risk-taking behaviour observed after a benzodiazepine ingestion in humans, the tryptic hydrolysate did not display such a side effect. These results suggest that the mechanism of action of the bovine alpha s1-casein tryptic hydrolysate may differ from that of diazepam.

A tryptic hydrolysate from bovine milk alphaS1-casein improves sleep in rats subjected to chronic mild stress.

The putative effects of a tryptic bovine alphaS1-casein hydrolysate on stress-induced sleep disorders were investigated and their possible link with typical blood stress parameters such as plasma corticosterone concentrations and glycaemia was assessed. Rats were subjected to chronic stress in the form of environmental disturbances, while receiving an oral administration of the alphaS1-casein hydrolysate (CH). Chronic stress significantly reduced sleep duration in control rats during the first 2 days of the stress period, but stress-induced sleep disturbance was prevented in CH-treated rats. Indeed, CH administration allowed the maintenance of slow wave sleep (SWS) duration and even a slight increase in paradoxical sleep (PS) duration in treated rats. Results on plasma corticosterone concentrations and on glycemia values were inconclusive with respect to the implication of the HPA axis in this study. However, the protective effect of the alphaS1-casein hydrolysate on sleep during exposure to our chronic mild stress conditions may be mediated by modulation of the central adrenergic response.

Effects of a tryptic hydrolysate from bovine milk alphaS1-casein on hemodynamic responses in healthy human volunteers facing successive mental and physical stress situations.

BACKGROUND: Preclinical results in rats have demonstrated anxiolytic-like effects of a tryptic bovine alphaS1-casein hydrolysate. AIM OF THE STUDY: We investigated the putative effects of this tryptic hydrolysate on systolic (SBP), diastolic (DBP) blood pressures, heart rate (HR) values and plasma cortisol concentrations (CC) in human healthy volunteers facing successive stress situations. METHODS: The subjects were (double blind) randomly allocated to ingest three times, 12 hours apart, two capsules containing either 200 mg of alphaS1-casein hydrolysate (TS) or bovine skimmed milk powder as a placebo (CS). On the morning of the test day, a first blood sample for baseline measurement of CC was taken before the subjects were submitted to the Stroop test (ST) and, after a 30-min rest, to a Cold Pressor test (CPT). SBP, DBP, and HR were continuously recorded for 5 min before the ST and during each stress situation. A second blood sample was taken 15 min after the end of the CPT condition. RESULTS: ST and ST + CPT combined test situations increased SBP, DBP and HR. The significant "Treatment x SBP" and "Treatment x DBP" interactions indicated the lower percentage changes in SBP and DBP of the TS. In addition, the results showed a significant decrease of the CC in the TS but not in the CS throughout the ST + CPT combined stress tests. HR remained stable in TS between the initial rest period and the CPT unlike what happened in CS. CONCLUSION: On the basis of blood pressure and cortisol changes, these results suggest an antistress profile of this alphaS1-casein hydrolysate in human subjects.