BACKGROUND: Platelets play important roles in several physiological and pathological processes. Multiple antiplatelet drugs have been developed for clinical practice. The active components of traditional Chinese medicine with antithrombotic effects are promising drugs to modulate platelet function. In our study, the antiplatelet effect of isoliquiritigenin (ILTG) and its mechanisms were examined. METHODS: Human platelet-rich plasma and a washed platelet suspension were prepared. Platelets were stimulated using collagen, thrombin, or adenosine diphosphate (ADP). The platelet lumi-aggregometer was applied to detect the aggregation of platelets and the release of adenosine triphosphate (ATP). The expression of P-selectin and the activation of integrin αIIbβ3 were detected using flow cytometry. The spreading of platelets on a fibrinogen-coated surface was visualized using immunofluorescent staining. The mechanisms of the antiplatelet effect were investigated using Western blotting. RESULTS: In this study, ILTG inhibited collagen- and thrombin-induced platelet aggregation, the release of dense granules and α-granules, and the activation of integrin αIIbβ3 in a dose-dependent manner. In addition, ILTG suppressed the spreading of platelets on immobilized fibrinogen. In collagen-activated platelets, ILTG markedly inhibited the expression of phosphorylation of phospholipase C gamma-2 (PLCγ2) and protein kinase B (Akt). CONCLUSIONS: These results indicated that ILTG could inhibit the collagen- and thrombin-induced platelet aggregation and granule release via the glycoprotein VI-mediated signal pathway .