This paper mainly introduced the preparation and anticoagulant activity of sulfated Schizophyllan polysaccharides （S-SPG）.The single-factor and response surface method were used to optimize the preparation of polysaccharides-SPG with the sulfur trioxide-pyridine method.The anticoagulant activity was evaluated through APTT，TT，and PT assays.The results show that when the mass ratio of sulfur trioxide-pyridine complex to Schizophyllan polysaccharides （SPG） is 4，the reaction temperature is 70℃，and the reaction time is 3h，the degree of substitution of S-SPG was the highest （1.85）.The FT-IR and 13C NMR spectroscopy data proved that the sulfated group was successfully inserted into schizophyllan.Anticoagulant assays show that sulfated Schizophyllan polysaccharides exhibits strong anticoagulant activity and prolongs APTT and TT，but has no effect on PT.The anticoagulant acti-vity is positively correlated with the concentration and degree of substitution.S-SPG3 with the highest degree of substitution shows the strongest anticoagulant activity.When the mass concentration is 0.2g/L，the APTT values of S-SPG3 is 184.35s，which is 1.86 times of S-SPG2 and 2.49 times of S-SPG1 under the same concentration.Besides，the TT values of S-SPG3 was over 90 s at the mass concentration of 0.2g/L.