Abstract: An aqueous two-phase system composed of 20% PEG600 and 20% (NH4)2SO4 was used to extract total protein from grape seeds.The effects of mass fraction of NaCl,pH value,extraction temperature and leaching solution volume on the extraction rates and distribution coefficient were explored by one-factor-at-a-time and central composite design(CCD)experiments.The results showed that:extraction temperature and leaching solution volume were major affecting factors,mass fraction of NaCl and pH value exhibited less effect on the extraction efficiency of total protein .The best extraction condition were NaCl mass fraction 0.06%,pH value 7.05,temperature 25℃,volume ratio of leaching solution to aqueous two-phase system was 3.97∶10.00.On the optimization condtition,the extraction rates of protein and dist ribution coefficient reached 77.52% and 4.61.