Abstract: The fibrinolytic enzyme was separated and purified from protein crude extracts, of Trichosanthes kirilowii by using affinity chromatography method and its enzymatic properties were studied. The results showed that the apparent molecular weight of the fibrinolytic enzyme was 66.0 kD, the optimal pH value was 9.0, and the enzyme activities had no significant changes under the temperature of 4-40℃ and was inactivate completely when temperature was more than 78℃. In the hydrolysis reaction with the casein as substrate, the effects of 4 kinds of factors such as temperature, enzyme substrate ratio, pH value and reaction time on the hydrolysis reaction were in order as follows： temperature〉enzyme substrate ratio〉reaction time 〉 pH value; the Km value was 1.89 mg/mL and Vmax was 555.56μmol/（ L·min）.