Abstract: OBJECTIVE To separate the fibrinolytic enzyme which is highly effective, specific, security and few side effects, to lay the foundation for the thrombolytic drugs. METHODS The crude fibrinolytic enzyme from peach kernel was precipitated with saturated ammonium sulfate and purified by affinity chromatography. The specific activity of the crude fibrinolytic enzyme was detected. Compared the inhibitory action of different proteinase inhibitors, separated and purified the crude fibrinolytic enzyme with the method of affinity chromatography. RESULTS A single active component was isolated from crude fibrinolytic enzyme with the method of affinity chromatography. The specific activity of the component was 89.08 U·mg^-1, which was 7.37 times higher than that before purified. This component belongs to the serine proteinase family. CONCLUSION Using the inhibitor of soybean trypsin as a ligand, a single active component could be obtained by affinity chromatography. And the specific activity increased relatively.