Abstract: Nine modification conditions were designed to sulfate epimedium polysaccharide (EPS) by chlorsulfonic acid-pyridine method as orthogonal test, and nine sulfated EPS (sEPS) were obtained. The effects of the nine sEPS on IBDV infecting chick embryo fibroblast (CEF) was compared by MTT assay. The results showed that modified sEPS had significant enhanced resisting effect of CEF on IBDV infection in comparison with non-sulfated EPS especially sEPS2 and sEPS5 presented better effects. The enhanced resisting effects of sEPS have certain relationships with the degree of sulfation (DS) and carbohydrate content. The optimum modification conditions were the reaction temperature of 80℃, the ratio of chlosulfonic acid to pyridine of 1∶8 and the reaction time of 2 h.