How D-amino acids embedded in the protein sequence modify its digestibility: Behaviour of digestive enzymes tested on a model peptide used as target.

D-amino acids can affect the action of digestive enzymes, hence the protein digestion. In this work the behaviour of the main stomach and gut digestive enzymes (pepsin, trypsin, and chymotrypsin) in the presence of D-amino acids in the protein chain was monitored over time using a model peptide, Ac-LDAQSAPLRVYVE-NH2 (belonging to ß-lactoglobulin, position 48-60), where L-amino acids were systematically substituted by D-amino acids. The results showed several changes in the behaviour of digestive enzymes, not only when the D-amino acids are inserted at the specific cleavage sites (after Val-57), but in some cases also when in distant positions. The effect seemed more pronounced in the case of pepsin rather than the gut enzymes, possibly indicating a better resilience of the upper gut phase of digestion to racemization. These results demonstrated that racemization could impair nutritional value by slowing down digestibility and has different effects according to the enzyme/amino acids involved.

Evaluation of in vitro whey protein digestibility in a protein-catechins model system mimicking milk chocolate: Interaction with flavonoids does not hinder protein bioaccessibility.

Flavonoids are largely present in plant food such as cocoa and derived products. These compounds can interact with proteins inherently contained in the food matrix and/or the proteolytic enzymes involved in gastrointestinal digestion. The flavonoid/protein interaction might hamper protein bioaccessibility and digestibility, affecting the nutritional quality. However, information on the digestion fate of proteins in food matrices containing both proteins and flavonoids is limited. The aim of this work was to evaluate the interaction between proteins and flavonoids and verify the potential effects of this interaction on protein digestibility. Taking milk chocolate as model, first a simple whey proteins/catechins mixed system was evaluated, and then the effects on digestibility were also verified in a real sample of commercial milk chocolate. The effects of the catechins/whey proteins interaction in the model system were evaluated by optical and chiro-optical spectroscopy, outlining a slight protein structure modification upon interaction with catechins. The digestibility of the protein fraction both in the model system, with and without catechins, and also in milk chocolate, was then determined by the application of INFOGEST in vitro digestion method: the bioaccessibility was evaluated in terms of protein hydrolysis and protein solubilisation, and major peptides generated by the digestion were also determined by LC/HR-MS. Despite the slight interaction with proteins, flavonoids were found to not hinder nor modify protein solubilization, protein hydrolysis and peptide profile by digestive enzymes. Also protein digestibility in milk chocolate, evaluated by SDS-PAGE, was found to be complete. The present data clearly indicate that the interaction of the proteins with the flavonoids present in the cocoa matrix does not to affect protein bioaccessibility during digestion.

Molecular composition of lipid and protein fraction of almond, beef and lesser mealworm after in vitro simulated gastrointestinal digestion and correlation with the hormone-stimulating properties of the digesta.

The current production of meat presents many disadvantages for the environment and much research focuses on alternative protein sources. Insects are novel protein sources highly valued for their nutritional and sustainable potential. However, many aspects concerning biological and nutritional properties of the insects after digestion, in comparison with other protein sources, are still overlooked. In this work, a comparative study on three different protein sources, namely almond, lean beef and insect Alphitobius diaperinus (lesser mealworm), was performed after in vitro simulated gastrointestinal digestion. An in-depth characterization of the chemical composition of the solubilized protein and lipid fractions of the digesta was performed by applying different analytical techniques, including chromatographic methods coupled to mass spectrometry and 1H NMR spectroscopy. Beef and insect were proven to be very similar in amino acid composition and protein solubilization after digestion, when considering the proper corrections for the chitin content. Lipid fraction from insects was solubilized during digestion as the one of almonds, but with a fastest kinetics. Thus, lesser mealworms are a good source of both lipids and highly nutritional proteins. Then, the amino acid composition of raw and digested protein fraction from the three sources was related to the PYY, ghrelin, GLP-1 and CCK release and rats' food intake. The composition of amino acids in insect digesta was found to be related to specific effects on enterohormone release, and the modulation of food intake in rats.

Structural and chemical changes induced by temperature and pH hinder the digestibility of whey proteins.

In the food and feed industry, protein extraction is commonly performed under acid or basic conditions, combined with heat, in order to increase the extraction yield. Under severe processing conditions, proteins may undergo molecular modifications. Here, the effects of heating (30, 60, 90 °C) at different pH values (2, 7, 9, 11, 13) were evaluated on commercial whey proteins, used as a simplified protein model. The main structure and chemical modifications concerning protein aggregation, hydrolysis, insolubilization, amino acid degradation and racemization were investigated in detail. Using in vitro static models, the degree of protein hydrolysis and the released peptides were determined after the digestive process. Accumulation of molecular modifications was mostly observed after basic pH and high temperatures treatments, together with a marked decrease and modification of the digestibility profile. Instead, protein digestibility increased in neutral and acidic conditions in a temperature-dependent manner, even if some modification in the structure occurs.

Beneficial effects of a T. monococcum wheat cultivar on diabetes incidence evaluated in non-obese diabetic mice and after in vitro simulated gastroduodenal digestion.

Wheat consumption can represent one of the nutritional factors involved in the onset of diabetes. We specifically investigated the potential diabetogenic effects of Hammurabi, a T. monococcum wheat cultivar, in non-obese diabetic (NOD) mice and analysed the levels of resistant starch in pasta manufactured with Hammurabi after in vitro gastroduodenal digestion. NOD mice were fed with Hammurabi, bread wheat or rice flour to evaluate diabetes incidence and insulitis score. An enzymatic method was applied to compare the content of resistant starch in Hammurabi pasta and durum wheat pasta (control). In NOD mice, the Hammurabi-based diet significantly delayed diabetes onset (p = 0.0042) and reduced insulitis score compared to rice or wheat-based diet. Furthermore, the resistant starch value following in vitro digestion of Hammurabi pasta was significantly higher (4.08%) than that of durum wheat pasta (2.28%). Taken together, these results highlighted the potential positive effects of the Hammurabi-based diet on diabetes incidence.