Ultrasound-assisted extractionof Cissus repenspolysaccharides (CRPs) was optimized through response surface methodology (RSM) based on Box-Behnken design (BBD).The maximum CRPsyield (16.18%) was achieved under the optimum extraction conditions: extraction time 72 min, extraction temperature 74 ℃, extraction power 240 W.Then three-phase partitioning (TPP) method combined with gradient alcohol precipitationwas used to obtained CRP20,CRP40, CRP60 and CRP80 from CRPs, and CRP80 has a higher purity than others.Theprimary chemical and structural characteristics of CRP80 were investigated by UV, FT-IR, high-performance liquid chromatography (HPLC) and high-performance gel-permeation chromatography (HPGPC).CRP80 is mainly composed of glucose, galactose, arabinose and mannose, with a molecularweights of approximately 2.95kDa. Furthermore, the antioxidant activity and hypoglyceamicactivity of CRP80 in vitrowere evaluated. The results showed that CRP80 had strong scavenging activities on ABTS, hydroxyl and DPPH radicals, as well as high scavenging activities on α-glucosidase andα-amylase. Our research providedan efficient method for the extraction of polysaccharidesfrom C. repensandCRP80 haspotential as a promising source of natural antioxidants and hypoglycemic agent for the functional food and medicinal industries.