Objective:To purify non hemorrhagic fibrinolytic enzyme(NHFLE) from Agkistrodon acutus venom and study their physicochemical properties .Methods:We purify non hemorrhagic fibrinolytic enzyme by filtration and anion exchange chromatography and determined fibrinolytic activity by fibrin plate,calculated molecular weight by SDS PAGE;and measured heorrhagic activity on animals.Result:By means of DEAE Sepharose CL 6B ,Sephadex G 75 chromatography and TSK GEL G-2000SW high performance gel filtration,a purified protein with fibrinolytic activity was obtained from the venom of Agkistrodon acutus.The protein was homogeneous as judged by electrophresis.Its molecular weight was estimated to be 25 000 without any hemorrhagic activity. The NHFLE had directly fibrinolytic activity and The specific fibrinolytic activity of the protein was 3.2 time higher than that of crude venom.Conclusion:The NHFLE from Agkistrodon acutus venom has directly fibrinolytic activity. Its molecular weight is estimated to be 25 000 without any hemorrhagic activity.