Pancreatic hydrolysis of bovine casein: Peptide release and time-dependent reaction behavior

Pancreatic hydrolysis of bovine casein was carried out at pH 8.0 in a batch stirred tank reactor. The resulting peptides were identified by liquid chromatography–tandem mass spectrometry and their release kinetics were tracked. During 24 h of digestion, a total of 297 peptides consisting of 109, 48, 126, and 14 peptides, from αs1-, αs2-, β-, and κ-caseins (αs1-, αs2-, β-, and κ-CN), respectively, were identified by LC–MS–MS. We found that pancreatin attacked different caseins with varying hydrolysis rates (β-CN > αs1-CN > αs2-CN > κ-CN). Based on our analysis of the peptide fragments produced, we determined that pancreatin preferentially cleaved the N-terminal phosphorylated region, the C-terminal hydrophobic region and the middle region of β-casein, the terminal regions of αs1-casein, and both the terminals and the middle region of αs2-casein. The hydrolysis of κ-casein occurred at the slowest rate, releasing peptides only after 30 min of hydrolysis.