Human blood samples were investigated in vitro to observe the antithrombotic effect of water extract of Gynostemma pentaphyllum (GP). The results showed that GP could inhibit significantly the platelet aggregation induced by ADP and compound agonists (P < 0.05), accelerate obviously the disaggregation (P < 0.05) and inhibit effectively the experimental thrombosis (P < 0.05). The delayed effects of GP on KPTT, PT, TT, AT, RVV-RT, RVV-CT suggested that this drug could decrease the activity of multiple coagulation factors. And it showed that GP could accelerate the erythrocyte electrophoresis rate. This study revealed that GP is an antithrombotic agent affecting the links of thrombotic chain which is worthwhile to be studied further.