Taking the euglena as the raw material, the extraction of euglena polysaccharide by ultrasonic-assisted enzymatic method and the optimal extraction conditions were studied, from the ultrasonic power, ultrasonic time, enzymolysis time, and enzyme concentration by response surface analysis(RSA) based on single factor tests. The results showed that the optimal process parameters were: ultrasonic power 360 W, ultrasonic time 19 min, neutral protease addition 3.7%, enzymatic hydrolysis time 3.5 h. Under the conditions, the yield of euglena polysaccharide reached 3.68%. The euglena polysaccharide (EGPS) was separated to three different ranges of molecular weight by ultrafiltration membrane, which were 35, 510 and ＞10 kDa, and the antioxidant activity of three EGPSs were analyzed. The results showed that, 35 kDa has better hydroxyl radical scavenging ability, superoxide anion scavenging ability, Fe2+ chelation ability, reducing power, and more than 10 kDa has better lipid peroxidation inhibition ability, none of the EGPSs has DPPH freedom.