Traditional Korean medicines with antioxidant properties are believed to have potential therapeutic effects against oxidative stress in cerebral ischemia. In this study, the antioxidant effects of Joongpoongtang 05 (JP05) were evaluated in rat middle cerebral artery occlusion (MCAO). Male Sprague-Dawley rats treated with JP05 (100 and 200 mg/kg body weight, p.o.) for 5 days showed a reduction in infarct sizes and a decrease in oxidants after MCAO. Furthermore, a significant inhibition of enzymatic antioxidants in the ischemic cerebral cortex was observed. Treatment with JP05 reversed the inhibition to normal levels. In the glutathione system, the reduced glutathione (GSH) level was decreased while oxidized glutathione (GSSG) was increased in MCAO rats. Treatment with JP05 increased the GSH/GSSG ratio and redox index by enhancing the synthesis of GSH from GSSG. Moreover, in vitro studies showed that JP05 has strong antioxidant activity in the Trolox equivalent antioxidant capacity, oxygen radical absorbance capacity and 1,1-diphenyl-2-picrylhydrazyl radical assays. The present findings suggest that JP05 treatment can contribute to a protective effect on brain ischemia through antioxidant mechanisms.