Release of angiotensin converting enzyme-inhibitor peptides during in vitro gastrointestinal digestion of Parmigiano Reggiano PDO cheese and their absorption through an in vitro model of intestinal epithelium

The occurrence of 8 bovine casein-derived peptides (VPP, IPP, RYLGY, RYLG, AYFYPEL, AYFYPE, LHLPLP, and HLPLP) reported as angiotensin converting enzyme-inhibitors (ACE-I) was investigated in the 3-kDa ultrafiltered water-soluble extract (WSE) of Parmigiano Reggiano (PR) cheese samples by ultra-performance liquid chromatography coupled to high-resolution mass spectrometry via an electrospray ionization source. Only VPP, IPP, LHLPLP, and HLPLP were revealed in the WSE, and their total amount was in the range of 8.46 to 21.55 mg/kg of cheese. Following in vitro static gastrointestinal digestion, the same ACE-I peptides along with the newly formed AYFYPEL and AYFYPE were found in the 3 kDa WSE of PR digestates. Digestates presented high amounts (1,880-3,053 mg/kg) of LHLPLP, whereas the remaining peptides accounted for 69.24 to 82.82 mg/kg. The half-maximal inhibitory concentration (IC50) values decreased from 7.92 ± 2.08 in undigested cheese to 3.20 ± 1.69 after in vitro gastrointestinal digestion. The 3-kDa WSE of digested cheeses were used to study the transport of the 8 ACE-I peptides across the monolayers of the Caco-2 cell culture grown on a semipermeable membrane of the transwells. After 1 h of incubation, 649.20 ± 148.85 mg/kg of LHLPLP remained in the apical compartment, whereas VPP, IPP, AYFYPEL, AYFYPE, and HLPLP accounted in total for less than 36.78 mg/kg. On average, 0.6% of LHLPLP initially present in the digestates added to the apical compartment were transported intact to the basolateral chamber after the same incubation time. Higher transport rate (2.9%) was ascertained for the peptide HLPLP. No other intact ACE-I peptides were revealed in the basolateral compartment. For the first time, these results demonstrated that the ACE-I peptides HLPLP and LHLPLP present in the in vitro digestates of PR cheese are partially absorbed through an in vitro model of human intestinal epithelium.