A fibrinolytic protease was purified from a Chinese herb (Spirodela polyrhiza). The protease has a molecular mass of 145 kDa and 70 kDa in gel filtration and SDS-polyacrlamide gel electrophoresis (PAGE), respectively, implying it is a dimer. Its optimum pH was 4.5-5.0. The enzyme was stable below 42 degrees C and after lyophilization. The enzyme activity was inhibited significantly by leupeptin and aprotinin. The protease hydrolyzed not only fibrin but also fibrinogen, cleaving Aalpha and Bbeta without affecting the gamma chain of fibrinogen. It preferentially cleaved the peptide bond of Arg or Lys of synthetic substrates (P1 position). The enzyme had an anticoagulating activity measured with activated partial thromboplastin time (APTT), thrombin time (TT), and prothrombin time (PT) tests. It delayed APTT, TT, and PT two times at the concentration of 36, 39, and 128 nM, respectively and this was drastically reduced after heat treatment.