Abstract: The purpose of this study was to optimize the preparation condition of the antioxidant peptide from Enteromorpha clathrata protein by emzymatic method and to test its digestive stability. Using DPPH free radical scavenging rate and DH as indicators,the preparation conditions of antioxidant peptides were optimized by single factor experiment and response surface analysis methodology. The hydrolysate was separated into three components with different molecular weight by three-stage membrane filtration system. Digestive stability of the components were tested by simulated gastrointestinal digestion system in vitro.The results showed that the optimum enzymatic hydrolysis conditions were:Temperature 47℃,pH7.4,enzyme amount 3300 U/g pro,solid-liquid ratio 1:20 g/mL,and duration time 3.3 h. Under this condition,DPPH radical scavenging rate was 84.33%±1.78%,The predicted values of the model was 85.53%. T test showed that there was no significant difference between the actual and the predicted values,which indicated that the model was reliable.The two components with molecular weight less 10 kDa had strong antioxidant activity and resistance to gastrointestinal proteases in vitro. The antioxidant activity of the components with molecular weight over 10 kDa was significantly lower,and was significantly reduced after digestive enzyme digestion. In conclusion,peptides with antioxidant activity and digestive stability could be obtained from Enteromorpha clathrata protein by neutral protease hydrolysis. The process model was reliable,and the hydrolysate had potential application value in food.