Bioactive peptides from milk proteins: focusing on peptides displaying immunomodulatory activity

Milk-derived peptides are milk components able to influence specific physiologicalfunctions, finally acting on body health condition. At present, the bioactivitiesdescribed for milk-derived peptides include opiate, antithrombotic, antihypertensive, immunomodulating, antioxidative, antimicrobial, anticancer, mineral-carrying andgrowth-promoting activities. In this thesis, special attention has been given to bioactive peptides with ACE-inhibitory and immunomodulatory activities. In the Experiment 1 Enterococcus faecalis TH563 (E. faecalis TH563) and Lactobacillus delbrueckii subsp. bulgaricus LA2 (L. delb. bulgaricus LA2), two bacterial strains isolated from traditional North Eastern Italy dairy products, have been evaluated for their ability to produce fermented milk rich in ACE-inhibitory and immunomodulatory activities. The preliminary results obtained from this experiment demonstrated that E. faecalis TH563 produced a fermented milk with high ACEinhibitory activity while L. delb. bulgaricus LA2 showed an immunomodulatory activity on bovine lymphocytes. To better understand the mechanisms underlying the immunomodulatory activity of fermented milks, in the Experiment 2 the immunomodulatory effects of the milkderived bioactive tri-peptide YGG have been examined. YGG could be generated during milk fermentation from a–lactalbumin hydrolysis operated by bacterial enzymes, so it could be present in milk fermented by L. delb. bulgaricus LA2. YGG has been administered to purified peripheral blood lymphocytes in different culture conditions (presence/absence of activators of lymphocyte proliferation, different concentration of newborn calf serum) and its effects on lymphocyte proliferation and cytokine RNA expression (IL2 and INFg) have been analyzed. YGG modulated lymphocytes proliferation, in a manner dependent from culture conditions but its effects did not seem mediated by the modulation of IL2 or INFg RNA expression. An important limiting factor of the large-scale diffusion of food carrying potential bioactivities is the bioavailability of the peptides responsible of such bioactivities. The main factors influencing the bioavailability of peptides are the resistance to digestion enzymes of and the absorption by the intestinal epithelium. In the Experiments 3 and 4 the sensitivity to gastrointestinal enzymes and the mechanisms of absorption of the peptide b-CN (193-209) have been evaluated. b-CN (193-209) is a long hydrophobic peptide derived from b-casein that has been already isolated and identified from fermented milks and yogurt and displayed immunomodulatory properties. The pattern of digestion and the mechanisms of absorption have been evaluated in well-known in vitro models for the intestinal epithelium, as the brush border membrane vesicles (BBMV) and the Caco-2 cell line. The results of these studies demonstrated that the b-CN (193-209) peptide is absorbed intact by the Caco-2 monolayer, probably via a vesicles-mediated mechanism. In conclusion, the main contribution of this PhD thesis was to provide new knowledge about milk-derived products with bioactivities. In particular, original contributions are in relation to the mechanisms by which milk-derived bioactive peptides are generated, express their bioactivities, and their fate in the gastrointestinal tract. As a result, new questions have arisen on this area that could constitute the objective of further research programs in the future.