Objective: To improve the availability of cherry seed, cherry seed oil was extracted by the aqueous enzymatic method. Methods: the ratio of mixed enzymes was optimized by mixture design on the basis of single factor experiment. The basic indexes of oil such as fatty acid composition and physical and chemical properties were detected. Results: the optimum extraction conditions were a 2.0% addition of mixed enzyme comprising cellulase, pectinase and acid protease （0.67∶0.10∶0.23）, liquid to material ratio of 10∶1 (mL/g), enzymolysis temperature of 45 ℃, pH of 4.0 and time of 4.0 h. The recovery of oil was 93.18% and the actual extraction rate was 28.66%. The oil from aqueous enzymatic technology was rich in oleic acid and linoleic acid, with the color of oil light-colored and transparent. Acid value and peroxide value conformed to the hygiene standard for edible vegetable oil. Conclusion: The mixture design assisted aqueous enzymatic extraction of cherry seed oil is feasible.