Abstract: Enzymatic hydrolysates derived from pumpkin seed protein was separated through ultrafiltration membranes of which the cut-off molecular weight were 10, 4ku.Optimum conditions of the two separation processes were obtained from the study of the effect of pressure, concentration of hydrolysate, pH and time on ultrafiltration permeate flux and membrane efficiency, and the antioxidant activity of hydrolysates before and after ultrafiltration also was studied.The result showed that the optimum conditions of the cut-off molecular weight of 10ku were pressure 0.25MPa, concentration of hydrolysate 3.0%, pH7 and time 60min.And the optimum conditions of the cut-off molecular weight of 4ku were pressure 0.20MPa, concentration of hydrolysate 2.0%, pH7 and time 60min.After ultrafiltration components with a molecular weight below 4ku had stronger free radical scavenging capacity, and reducing power, the IC 50 value of diphenyl picryl-hydrazyl radical (DPPH.) , hydroxyl radical (.OH) , and superoxide anion radical (O 2.) respectively was 2.76, 2.91, 23.58mg/mL, and the concentration of the components was 9.43mg/mL when the absorbance was 0.5.