Abstract: In order to prepare corn oligo-peptide efficiently, corn gluten meal was pretreated by ultrasonic.Using the yield of oligo-peptide as the evaluation indicator, Alcalase was screened out from the following six kinds of protease-Protamex, Alcalase, Flavourzyme, Protex7L, Trypsin and Papain. Single factor experiment and response surface methodology were used for optimizing the enzymatic hydrolysis conditions in this study. The optimum hydrolysis conditions were designated as enzyme concentration of 2660U/g, substrate concentration8.0g/100mL, Enzymolysis temperature of 57.0℃, pH 9.0, and enzymolysis time of 3h. Under these conditions, yield of corn oligo-peptide could reached up to 34.34%±0.22%, which coincided with predictive values with the relative error of less than ±1%.