Bioaccessibility and Antioxidant Activity of Faba Bean Peptides in Comparison to those of Pea and Soy after In Vitro Gastrointestinal Digestion and Transepithelial Transport across Caco-2 and HT29-MTX-E12 Cells.

In this study, the transepithelial transport of bioactive peptides derived from faba bean flour gastrointestinal digestates was investigated, in vitro, using a Caco-2 and HT29-MTX-E12 coculture monolayer, in comparison to those of pea and soy. The profile of transported peptides was determined by mass spectrometry, and the residual antioxidant activity was assessed. The ORAC value significantly (p < 0.05) decreased after transepithelial transport (24-36% reduction) for all legumes, while the antioxidant activity in ABTS assay significantly (p < 0.05) increased, as shown by the EC50 decrease of 26-44%. Five of the nine faba bean peptides that crossed the intestinal cell monolayer exhibited antioxidant activity. Two of these peptides, TETWNPNHPEL and TETWNPNHPE, were further hydrolyzed by the cells' brush border peptidases to smaller fragments TETWNPNHP and TWNPNHPE. These metabolized peptides were synthesized, and both maintained high antioxidant activity in both ABTS (EC50 of 1.2 ± 0.2 and 0.4 ± 0.1 mM, respectively) and ORAC (2.5 ± 0.1 and 3.4 ± 0.2 mM of Trolox equivalent/mM, respectively) assays. These results demonstrated for the first time the bioaccessibility of faba bean peptides produced after in vitro gastrointestinal digestion and how their bioactive properties can be modulated during transepithelial transport.

Antioxidant and Angiotensin-Converting Enzyme Inhibitory Activity of Faba Bean-Derived Peptides After In Vitro Gastrointestinal Digestion: Insight into Their Mechanism of Action.

Faba bean flour, after in vitro gastrointestinal digestion, showed important antioxidant and angiotensin-converting enzyme (ACE) inhibitory activities. In the present study, 11 faba bean- derived peptides were synthesized to confirm their bioactivities and provide a deeper understanding of their mechanisms of action. The results revealed that 7 peptides were potent antioxidants, namely, NYDEGSEPR, TETWNPNHPEL, TETWNPNHPE, VIPTEPPH, VIPTEPPHA, VVIPTEPPHA, and VVIPTEPPH. Among them, TETWNPNHPEL had the highest activity in the ABTS (EC50 = 0.5 ± 0.2 mM) and DPPH (EC50 = 2.1 ± 0.1 mM) assays (p < 0.05), whereas TETWNPNHPE had the highest activity (p < 0.05) in the ORAC assay (2.84 ± 0.08 mM Trolox equivalent/mM). Synergistic and/or additive effects were found when selected peptides (TETWNPNHPEL, NYDEGSEPR, and VVIPTEPPHA) were combined. Four peptides were potent ACE inhibitors, where VVIPTEPPH (IC50 = 43 ± 1 µM) and VVIPTEPPHA (IC50 = 50 ± 5 µM) had the highest activity (p < 0.05), followed by VIPTEPPH (IC50 = 90 ± 10 µM) and then VIPTEPPHA (IC50 = 123 ± 5 µM) (p < 0.05). These peptides were noncompetitive inhibitors, as supported by kinetic studies and a molecular docking investigation. This study demonstrated that peptides derived from faba beans have multifunctional bioactivities, making them a promising food-functional and nutraceutical ingredient.

Improved in vitro gastrointestinal digestion protocol mimicking brush border digestion for the determination of the Digestible Indispensable Amino Acid Score (DIAAS) of different food matrices.

The Digestible Indispensable Amino Acid Score (DIAAS) is the new gold standard method for the assessment of protein nutritional quality. The DIAAS is evaluated with in vivo models, that are complex, constraining and costly. There is still no established method to assess it in vitro. In this study, we proposed to add a jejunal-ileal digestion phase to the standardized in vitro gastrointestinal digestion protocol developed by the International Network of Excellence on the Fate of Food in the Gastrointestinal Tract (INFOGEST protocol) to mimic brush border digestion and to enable DIAAS assessment in vitro in a more physiologically relevant manner. This jejunal-ileal digestion phase was performed with a porcine intestinal aminopeptidase as an alternative to brush border membrane extract, which is more difficult to obtain in a standardized way. This modified INFOGEST protocol was applied to various food matrices (faba bean, pea and soy flours, whey protein isolate and caseins) and the results were compared to published in vivo data to assess the model's physiological relevance. The addition of the jejunal-ileal digestion phase lead to a significant (p < 0.05) increase of 31 and 29 % in free and total amino acid digestibility, respectively, and of 83 % on average for the in vitro DIAAS values for all food matrices. Although the in vitro DIAAS remained underestimated compared to the in vivo ones, a strong correlation between them was observed (r = 0.879, p = 0.009), stating the relevance of this last digestion phase. This improved digestion protocol is proposed as a suitable alternative to evaluate the DIAAS in vitro when in vivo assays are not applicable.