As a specific angiogenesis inhibitor, angiostatin can inhibit new vascular formation, and it has potential clinical practical value. Human plasminogen was purified from human plasma by affinity chromatograph, and then in situ digestion of plasminogen by elastase was carried out to produce angiostatin fragments. After washing, angiostain was eluted specifically by 0.2 mol/L 6-aminocaproic acid solution. This improvement made this method be simpler, rapider and more efficient. In vitro and in vivo experiments indicated that this purified protein has potent antiangiogenic activity.