【Objcetive】 The objective of the study is to research the extracellular proteinase of Ascospheare apis which plays a key role for the pathogen to infect honeybee. 【Mehtod】 The extracellular enzyme of A. apis was induced, purified and concentrated from an artificial medium. Then different substrates were used to determine the activity of caseinase,azocaseinase,collagenase and elastase trypsin. 【Result】 The results showed that caseinase, azocaseinase and collagenase were secreted by A. apis. Moreover, the optimum reaction temperature of three extracellular pretenase was all around 40℃, the optimum pH was all around 7. They were stable in middle pH buffer system under low temperature but quickly lost their activity under high temperature. 【Conclusion】 Multiple extracellular proteinase could be secreted by A. apis. Their optimum temprature was around 40℃ and the optimum pH was around 7, which was coincident with the physiological parameter of bee larvae midgut.