ADSA® -EAAP PhD Student Travel Award Presentation: Bioactivities of milk proteins evaluated after in vitro digestion and peptidomic/proteomic profile

The health-promoting functions of milk peptides are related to their structures, which depend on the parent protein composition. A crucial issue in this field is the demonstration of a cause-effect relationship, from the ingested form to the bioactive form. Thus, the aim our study was to in vitro digest intact whey proteins (WPI) and casein proteins (CP), mimicking in vivo digestion, to investigate their bioactive effects. WPI and CP were digested using a pepsin/pancreatin protocol and ultrafiltered (3kDa cut-off membrane). A permeate (mimicking absorbed fraction) and a retentate (mimicking intestinal fraction) were obtained. Soya protein was included as control (CTR). Antioxidant (AOX) and angiotensin-I-converting enzyme inhibitory activity (ACEI-i) of the WPI and CP fractions were assessed and compared with undigested sample (UND) and CTR. Further, the effects of WPI and CP retentate on proliferation and on mucus production (MUC5AC gene expression and mucin secretion) were assessed in intestinal goblet HT29-MTX cells. Finally, permeate has been characterized by LC-nano ESI MS/ MS using a shotgun-peptidomic approach while retentate has been further digested with trypsin and analyzed by mass spectrometry with a shotgun-proteomic approach and the potentially bioactive peptides and polypeptides have been identified. WPI and CP permeate showed a significant (P < 0.05) increase in AOX up to 1.58 ± 0.04 and 1.61 ± 0.02 µmol trolox antioxidant equivalents /mg protein (TrAOX equ/ mg P), compared with UND (<0.6 µmol TrAOX equ/mg P) and CTR permeate (1.37 ± 0.04% µmol TrAOX equ/mg P). WPI and CP permeate exhibited a higher (P < 0.05) ACEI-i (49.20 ± 0.64 and 23.91 ± 0.64%, respectively) compared with UND (<19%) and with CTR permeate (10.40 ± 1.07%). In HT29-MTX cells, WPI and CP retentate, at specific concentrations, were able to modulate (P < 0.05) cell proliferation; MUC5AC expression was increased (+20%; P < 0.05) by CP retentate and unaltered (P > 0.1) by WPI retentate. CP retentate evoked mucinsecretory effect. Our results confirm that milk proteins may be rich sources of bioactive compounds with a greatest beneficial potential of CP at intestinal goblet cell level.