Purification and identification of antioxidant activity and angiotensin converting enzyme (ACE) inhibitory activity peptide of peanut protein based on ultrafiltration (UF) membrane separation

Peanut protein isolates (PPI) were extracted from defatted peanut flour as a byproduct of peanut seed oil extraction. PPI treated by high pressure (HP) at 300 MPa was hydrolyzed by Alcalase, and the resulting peanut protein hydrolysates (PPH) were further fractionated into four fractions by ultrafiltration (UF) membrane with a molecular weight (Mw) cut-off ranging between 3 kDa and 5 kDa. The fractionated hydrolysates were designated UF-I (whole hydrolysates), UF-II (Mw of more than 5 k Da), UF-III (Mw of 3-5 kDa), and UF-IV (Mw of less than 3 kDa), respectively. The molecular weight distribution, amino acid composition, antioxidant activity and angiotensin-converting enzyme (ACE) activity of the various ultrafiltration fractions were analyzed. The maximum membrane flux was obtained when the ultrafiltration conditions were 120 min of hydrolysis time, 100 rpm of rotor speed and 4% of hydrolysate concentration, separately. The content of hydrophobic amino acids, polar amino acids, basic amino acids, aromatic amino acids and branched amino acids in the different fractions from highest to lowest was UF-IV, UF-III, UF-I, and UF-II, respectively. Based on the evaluation of the antioxidant and ACE inhibitory activities of the various components, a novel peptide exhibiting both antioxidant and ACE inhibitory activities was isolated. The amino acid sequence Tyr-Cys-Ala-Asp, with Mw of 471 Da, was identified. The findings suggest that the UF membrane is capable of effectively separating protein hydrolysates based on their molecular weight, amino acid composition and the sequence of bioactive peptides, which significantly enhances the bioactivity of peptides.