Abstract: To study the protective effects of Roudoukou-8 San extract on hypoxia/reoxygenation injury of cardiac myocytes and its relationship with tyrosine protein kinase 2(JAK2)/signal transducer and activator 3(STAT3)signaling pathway, hypoxia/reoxygenation injury model of H9c2 cells was built by sodium dithionite(Na₂S₂O₄). The vitality of the cells was tested by CCK-8; the contents of lactate dehydrogenase(LDH), creatine phosphate kinase(CK)and aspartate aminotransferase(AST)in cell culture medium were tested by fully automatic biochemical analyzer; the contents of catalase(CAT), superoxide dismutase(SOD)and glutathione peroxidase(GSH-Px)in cells were tested by kits; cell apoptosis degree was tested by hoechst staining. And the protein expressions of JAK2, p-JAK2, STAT3, p-STAT3, Bcl-2 and Bax in myocardial cells of each group were tested by Western blot. Hypoxia for 1 hour and reoxygenation for 3 hours of 2 mmol/L Na₂S₂O₄ caused damage of about 50% H9c2 cells. Compared with the model group, the extracts of Roudoukou-8 San with different concentrations could increase the viability of cells. Besides, contents of CK, LDH and AST in cell culture medium were decreased significantly, while contents of CAT, SOD and GSH-Px were increased significantly. At the same time, the expression of p-JAK2, p-STAT3 and Bcl-2 were significantly increased and that of Bax was significantly decreased. The effects of Roudoukou-8 San extract could bereduced by AG490 blocker. Therefore, Roudoukou-8 San extract possesses protective effects on Na₂S₂O₄ induced-hypoxia/reoxygenation injury of cardiac myocytes through JAK2/STAT3 signaling pathway.