Assessment of casein phosphopeptide profile in in vitro digestates of Trentingrana PDO cheese

Two different in vitro static gastrointestinal digestion (GID) protocols were applied to investigate the profile of casein phosphopeptides (CPPs) released from protected designation of origin (PDO) Trentingrana cheese. CPPs in the gastrointestinal digestates were selectively precipitated and identified by ultra-performance liquid chromatography coupled to high-resolution mass spectrometry. Digestion parameters adopted during gastric and intestinal phases of the two GID procedures were pivotal in the resulting CPP patterns. A standardized digestive protocol mimicking in vivo conditions, made available half of the CPPs identified following a simpler procedure based only on the action of pepsin and pancreatin. Independently of the GID protocol used, the mono-phosphorylated CPPs were the most abundant (>55%) followed by bi- (23-29%) and tri-phosphorylated (8-18%) peptides. Alfas1-casein was the parent fraction mainly contributing to CPP release. The number of peptides containing the biofunctional cluster sequence SerP-SerP-SerP-Glu-Glu almost overlapped in the digestates from the two GID models. About a half of these peptides was common between the two GID models, and a large part of them derived from alfas2- and beta-caseins. Overall, results of this investigation proved that CPP profiles of in vitro digestates of Trentingrana were similar to those previously reported for Grana Padano PDO using the same digestion protocols.