Preparation and evaluation of antioxidant peptides from ethanol-soluble proteins hydrolysate of Sphyrna lewini muscle

Abstract To get high yield of ethanol-soluble proteins (EP) and the antioxidant peptides from Sphyrna lewini muscle, orthogonal experiments (L 9 (3) 4 ) were applied to optimize the best extraction conditions and enzyme hydrolysis conditions. The yield of EP reached 5.903 ± 0.053% under the optimum conditions of ethanol concentration 90%, solvent to material ratio 20:1, extraction temperature of 40 °C and extraction time of 80 min. The antioxidant SEPH (EP hydrolysate of S. lewini muscle) was prepared by using papain under the optimum conditions of enzymolysis time 2 h, total enzyme dose 1.2%, enzymolysis temperature 50 °C and pH 6, and its DPPH radical scavenging activity reached 21.76 ± 0.42% at the concentration of 10 mg/ml. Two peptides (F42-3 and F42-5) were isolated from SEPH by using ultrafiltration, anion-exchange chromatography, gel filtration chromatography and RP-HPLC. The structures of F42-3 and F42-5 were identified as Trp-Asp-Arg and Pro-Tyr-Phe-Asn-Lys with molecular weights of 475.50 Da and 667.77 Da, respectively. F42-3 and F42-5 exhibited good scavenging activity on hydroxyl radical (EC 50 0.15 mg/ml and 0.24 mg/ml), ABTS radical (EC 50 0.34 mg/ml and 0.12 mg/ml), and superoxide anion radical (EC 50 0.09 mg/ml and 0.11 mg/ml), but moderate DPPH radical (EC 50 3.63 mg/ml and 4.11 mg/ml). F42-3 and F42-5 were also effectively against lipid peroxidation in the model system and peroxyl free radical scavenging in β-carotene linoleic acid assay. Their high activities were due to the smaller size and the presence of antioxidative amino acids within the peptide sequences.