Abstract: To investigated the activity in vivo and in vitro of polysaccharide of Polygala tenuifolia Willd., the exhaustive animal exercise model was designed. Allmice were administrated with low, medium and high doses ofpolysaccharide extract(0.10, 0.20, 0.40 mg/g·d) by gavage. After this, weight-loaded swimming time, and the levels of liver glycogen(LG), muscle glycogen(MG), blood ureanitrogen(BUN), blood lactic acid(BLA), serum lactate dehydrogenase(LDH) of different groups of animals were measured as well as the determination of antioxidant activity in vitro. The results showed that compared with the blank control group, the polysaccharide extract groups at low, middle and high dose prolonged weight-loaded swimming time by 96.6 s(P<0.05), 254.4 s(P<0.01) and 421.8 s(P<0.01), increased LG concentration by 13.1%(P<0.05), 37.4%(P<0.01) and 56.4%(P<0.01), increased MG concentration by 10.8%(P<0.05), 31.2%(P<0.01) and 42.0%(P<0.01), decreased BLA concentration after sport by 4.6%(P<0.05), 8.6%(P<0.01) and 14.5%(P<0.01), decreased BUN concentration after sport by 1.5%(P<0.05), 3.9%(P<0.01) and 7.5%(P<0.01), enhanced LDH activity by 10.4%(P<0.01), 14.0%(P<0.01) and 19.9%(P<0.01). When the concentration of polysaccharide was 4 mg/mL, the scavenging rate of hydroxyl free radical and DPPH free radical was 61.3% and 79.5%. Therefore, the polysaccharide of Polygala tenuifolia Willd. would have contribute to relieve fatigue in body, and the anti-oxidantfunction in vitro would be better.