In this study, the extraction conditions of polysaccharides from Hippocampus mohnikei were optimized by single factor test and response surface test, and the monosaccharide composition and antioxidant activity of Hippocampus mohnikei polysaccharides were determined. The results showed that the optimum extraction condition were ultrasonic time 47 min, extraction temperature 87℃, and solvent-to-solid ratio of 23 mL/g. Under these conditions, the extraction yield of crude polysaccharide HP was 9.91%. Two kinds of polysaccharides, HP-1 and HP-2, were obtained after being purified by column chromatography. HP-1 was mainly composed of glucose, galactose, mannose in a molar ratio of 11.32:6.39:5.64, with a small amount of arabinose, xylose, fucose and rhamnose. HP-2 was mainly composed of glucosamine, glucuronic acid, galacturonic acid, galactose, glucose, mannose in a molar ratio of 29.03:18.18:8.48:6.30:4.88:2.70, with a small amount of fucose, and xylose. HP-1, HP-2 had the ability of scavenging DPPH free radical, among which HP-2 had the best effect. Furthermore, HP, HP-1 and HP-2 had reducing power, indicating that Hippocampus mohnikei polysaccharides had better antioxidant activity.