Abstract: In this study, ultrasonic-assisted enzymatic hydrolysis, ultrafiltration and ?KTA chromatography were sequentially used to separate and purify angiotensin-converting enzyme (ACE) inhibitory peptides from an enzymatic hydrolysate of soybean meal, and the molecular mass distribution of peptides was investigated. Then, the amino acid sequence of the ACE inhibitory peptides was identified by mass spectrometry (MS) and molecular docking. A consensus peptide sequence was synthesized by a solid-phase method to determine its ACE inhibitory activity and explore its underlying mechanism by molecular docking. The results showed that after ultrafiltration, the molecular mass distribution of peptides in the hydrolysate was mainly under 6 000 Da. The peptides GVRP and IIVTP, selected for their lowest free energy (–8.44 and –9.04 kcal/mol correspongdingly) predicted by molecular docking, could inhibit ACE activity, with half-maximum inhibitory concentration (IC50) values of (84 ± 0.06) and (77 ± 0.08) μmol/L, respectively. The molecular docking results revealed that GVRP and IIVTP could form hydrogen bonds with the S1, S1′ and S2 pockets of ACE. The shared amino acid residues in close contacts (within 3.5 ?) with ACE were His513, Ala354 and Glu384. Based on the?LC-MS/MS?and the molecular docking technology, ?a method to rapidly identify and select active ACE inhibitory peptides from peptides mixture was established. The stable binding of active peptides to ACE and the inhibitory mechanism of its ACE activity were also reflected in this study. The finding of this study may provide further references for studies on ACE inhibitory peptides.

Key words: soybean meal; angiotensin-converting enzyme inhibitory peptides; isolation and purification; molecular?docking; active site pockets