Abstract: In order to obtain Hypophthalmichthys molitrix antioxidant peptides, two-step-hydrolysis methodology was used.First of all, Alcalase was chosen as the enzyme for the first-step-hydrolysis from Alcalase, Flavourzyme, Trypsin and Neutrase.Degree of hydrolysis (DH) was tested by pH-stat method.Results indicated that the optimal processing conditions for hydrolyzing Hypophthalmichthys molitrix protein hydrolysate of Alcalase were protein concentrations 15%, pH9.0, 50℃, ratio of Alcalase to silver carp protein with 24AU/kg.Then Trypsin was chosen from Flavourzyme, Trypsin and Neutrase as the enzyme for the second-step-hydrolysis.DPPH free radical scavenging ratio as response values was analyzed with response surface methodology.With the factors such as product cost and process requirement considered, optimum conditions to achieve the maximum antioxidant ability were as follows:Alcalase enzymolysis time 59.12min, Trypsin enzymolysis time 82.24min, Alcalase to Hypophthalmichthys molitrix protein with 24AU/kg, Trypsin to silver carp protein with 35.5AU/kg.The results of analyzing components showed that Hypophthalmichthys molitrix antioxidant peptides made under the same hydrolysis conditions contained 70.84% of soluble N, 69.38% of peptides, 4.21% of water and 25.67% of ash.DPPH free radical scavenging ratio was 19.33% when the peptide concentration was 2mg/mL.