The optimal enzymolysis conditions of preparing antioxidant peptide from Corbicula fluminea were optimized. On the basic of single factor experiment such as kinds of proteases, dosage of protease, the ratio of material to liquid, enzymatic hydrolysis time, enzymatic hydrolysis temperature and pH, the effects of different factors and level on hydroxyl radical scavenging were researched by Box-Behnken center combination experiment and response surface analysis methods. The optimal hydrolysis conditions of preparing antioxidant peptide from Corbicula fluminea were that adding 0.94% neutral protease（by weight of Corbicula fluminea meat）, material to liquid ratio of 1∶2(m∶V), pH of 2.6，enzymatic hydrolysis temperature of 54.70 ℃ and enzymatic hydrolysis time of 3.91 h. Hydroxyl radical scavenging of hydrolysates prepared under these conditions was 81.63%, which was in accordance with the value estimated by RSM (83.34%). The conditions of enzymatic preparation of antioxidant peptide from Corbicula fluminea could accurately predicted by Box-Behnken center combination experiment.