Abstract: The preparation and antioxidant activity of protein from defatted Chinese chive seed powder by phosphate buffer extraction method were studied. With the leek seed protein extraction ratio, DPPH radical scavenging rate and·OH radical scavenging rate ability as index, it explored the effect of extraction temperature, extraction time, pH and solid-liquid ratio for each three levels on polypeptide material extraction of leek seed and maked L9 (34) orthogonal test to determine the optimal preparation conditions. The optimal technological conditions were as follows:extraction temperature 48℃, extraction time 1.5h, pH7.5 and the ratio of solid-liquid1∶10 and the largest amount of extracted protein could reach 9.87%. When the leek seed protein concentration were 1mg/mL and 2mg/mL, the OH· and DPPH· removal rate were 73.37% and 25.2% respectively. The protein from defatted Chinese chive seed with the phosphate buffer solution had antioxidant activity.