For a long time Rhus verniciflua Stokes (RVS) has traditionally been used as a herbal plant. It is known to contain various biological activities. Previously, a crude ethanol extract from RVS was reported to have antioxidant effects, and antiproliferative activities, on human cancer cell lines. In this report, we prepared a highly purified ethanol extract from RVS, which did not contain the urushiol derivatives, named REEE-1 (thus ethanol eluted extract-1), to investigate the mechanisms of the scavenging activity of hydroxyl radicals using mouse thymocytes. The results from the deoxyribose, DNA nicking, and glucose/glucose oxidase enzyme assays showed that REEE-1 contained a strong scavenging activity of oxygen free radicals, especially of hydroxyl radicals. However, interestingly, REEE-1 also showed cytotoxicity against the thymocytes, although the effect was variable, depending on the concentrations and times of treatment. The REEE-1-mediated cytotoxicity against thymocytes, which has been used as one of the well-characterized models for apoptosis studies, was verified to be apoptotic. This was proven by the following: the appearance of DNA laddering, increases in DNA fragmentation, low fluorescence intensity in the nuclei after propidium iodide staining, and positive Annexin V staining of the cells. These results suggested that REEE-1 had both antioxidative activity and cytotoxicity against the thymocytes, although the effect of the cytotoxicity was variable, depending on the dose and time of the treatment.