Abstract: The extraction condition of polyphenols from evening primrose defatted seed by ultrasonic-asswasted extraction was optimized. The effects of solvent concentration, solid-liquid ratio, extraction time, extraction temperature, p H value of solvent, and dosage of extraction adjuvant on extraction efficiency are investigated by single-factor experiment coupled with orthogonal experiment. The results showed that the optimum extraction condition was extraction solvent p H3, extraction adjuvant ( sodium hexametaphosphate) mass concentration0.2%, volume percentage of methanol 80%, solid-liquid ratio 1∶30 (g/m L) , extraction time 60 min, and extraction temperature 40℃. Under thwas condition, the yield of polyphenols was 6.70%. The crude extract was further separated by macroporous resin. The polyphenols content of each fraction was analyzed. And the antioxidant activity of each fraction was investigated by autoxidation of pyrogallol method and DPPH assay, respectively.The results showed that polyphenols were mainly concentrated in fraction Ⅱ with the mass percent of 50.09%in crude extract. The mass fraction of polyphenols in fraction Ⅱ was 79.49%. The antioxidant activity of fractionⅡ and fraction Ⅲ were similar and significantly stronger than the other fractions.