Maximizing hypocholesterolemic peptides from an olive byproduct by enzymatic hydrolysis

Novel food proteases (Maxipro A, Maxipro B, Maxipro C, and Maxipro D) with broad specificity and different effective pH range (from 2.5 to 9.0) were evaluated to produce hypocholesterolemic peptides from olive seeds proteins. Box-Behnken experimental design was employed to evaluate the effect of the hydrolysis time, pH, temperature, and enzyme:substrate ratio on the amount of released peptides. The production of peptides, under optimal conditions, was higher with Maxipro C and lower with Maxipro B. The capability of extracts to reduce the absorption of exogenous cholesterol and the production of endogenous cholesterol was also evaluated. All hydrolysates showed a higher capacity to bind bile acids and inhibit cholesterol esterase enzyme than the obtained with usual Alcalase enzyme. Proteomic analysis using high resolution RP-HPLC-ESI-QTOF with database searching and de novo identification was employed to identify peptides in hydrolysates. Some sequences with hypocholesterolemic features were detected among the 45 peptides identified in Maxipro A hydrolysate, the 56 peptides in that of Maxipro C, and the 50 in the Maxipro B one. This work demonstrates that Maxipro A enzyme can hydrolyze olive seeds proteins, under acidic pH, releasing peptides with multifunctional and higher hypocholesterolemic capacity than the obtained with Alcalase, under alkaline conditions. •New enzymes yielded hydrolysates with hypocholesterolemic ability from olive seeds.•Hydrolysate from Maxipro A showed high capacity to reduce cholesterol solubility.•It promoted bile acid binding and cholesterol esterase and HMG-CoA R inhibition.•Some peptides in Maxipro enzymes showed hypocholesterolemic features.•Maxipro A, using acid pH, is an alternative to Alcalase, under alkaline conditions.