A natural heparin-like compound was isolated from swim bladder and its anticoagulant activities were determined. The response surface analysis based on Box-Behnken design was adopted to examine effects of protease consumption, extracting time and temperature on heparin-like compound extraction. The anticoagulant activity of swim bladder heparin-like compound was evaluated by APTT， PT and TT method. Consequently, the optimum conditions were as follows. The concentration of protease from Bacillus licheniformis was 5.4 mg/mL, extracted at 50 ℃ for 20 h. Under the control of these conditions, the predicted extraction rate by mathematical model was (1.79±0.05)%, and the standard error was 3.24%. In addition, the heparin-like compound from swimming bladder (HSB) showed negligible in vitro anticoagulant activity, and played a role through the exogenous coagulation pathway and the common coagulation pathway.