Abstract: In this study, Inonotus hispidus was used as a raw material, extraction rate of polysaccharide was used as the index, the single factor experiments and Box Behnken experiment design were used to study the preparation process of Inonotus hispidus polysaccharide (IHP) by ultrasound and microwave synergistic extraction, and the effects of several extraction methods on the extraction percentage of IHP and the bile acid binding capacity in vitro were compared and analyzed. The results showed that, the optimal process parameters for the extraction of IHP were: Material-to-water ratio 1:33 g:mL, microwave time 50 s, microwave power 500 W, ultrasonic time 51 min, ultrasonic power 200 W, the extraction percentage of polysaccharide under this condition was 85.61%. Compared with ultrasonic assisted extraction and hot water extraction, the extraction rate increased by 24.87% and 36.38%, respectively. The IHP had significant binding ability to sodium taurocholate and sodium glycocholate, which was positively correlated with the dosage of polysaccharide, and the binding ability of IHP to sodium taurocholate was higher than sodium glycocholate. And under the same mass concentration conditions, the binding ability of polysaccharides prepared by the three extraction methods to bile salts was ultrasonic and microwave synergistic extraction>ultrasonic assisted extraction>hot water extraction, and the binding rates for sodium glycocholate and sodium taurocholate were 30.93% and 32.13%, respectively. This study showed that the ultrasonic and microwave synergistic extraction method could significantly improve the extraction effect of IHP and the ability to bind cholate in vitro, and would provide a theoretical basis for the preparation and development and utilization of highly active IHP.