We investigated pharmacological effects of rutin isolated form Lonicera japonica on H2O2-induced cell death in H9c2 cells in vitro and rat myocardial ischemia-reperfusion injury model in vivo. Western blot analysis showed that H2O2 increased expression of cleaved form of caspase-3 and proapoptotic Bax protein, but decreased antiapoptotic Bcl-2 protein in H9c2 cell. However, treatment with rutin decreased expression of both cleaved from of caspase-3 and increased Bcl-2/Bax ratio in H9c2 cells. The protective effect of rutin was inhibited not by JNK inhibitor or p38 MAPK inhibitor but by PI3K inhibitor or ERK inhibitor. Rutin increased phosphorylation of ERK and Akt in H9c2 cells. These anti-apoptotic effects of rutin were confirmed both by annexin-V and TUNEL assay. Furthermore, rutin improved I/R-induced myocardial contractile function and reduced infarct size. Rutin administration also inhibited apoptosis in myocardial tissues in I/R rats by increasing Bcl-2/bax ratio and decreasing active caspase-3 expression. These results suggest that rutin reduced oxidative stress-mediated myocardial damage in vitro model and in vivo model, which might be useful in treatment of myocardial infarction.