Reactive oxygen species can play an important role in the pathogenesis of anoxia-reoxygenation injury. Sasanquasaponin (SQS) is a biologically active ingredient extracted from the Chinese medicinal plant Camellia oleifera Abel. Some studies have shown that SQS possesses potent antioxidant activities. However, it has not been elucidated whether SQS diminishes reactive oxygen species stress induced by anoxia-reoxygenation injury in cardiomyocytes. In this work, neonatal rat cardiomyocytes pretreated with the test compound were subjected to anoxia-reoxygenation. The extent of cellular damage was accessed by cell viability and the amount of released lactate dehydrogenase (LDH). Superoxide dismutase, catalase and glutathione peroxidase activities, reduced (GSH) and oxidized glutathione (GSSG) levels, and malondialdehyde contents were measured by a colorimetric method. The levels of intracellular reactive oxygen species and calcium were determined by flow cytometry. The results showed that SQS reduced LDH release and increased cell viability in a dose-dependent manner up to 10 microM and concomitantly decreased malondialdehyde and GSSG contents, while significantly increased GSH contents and the activities of superoxide dismutase, catalase and glutathione peroxidase. Moreover, treatment with SQS decreased intracellular reactive oxygen species levels and alleviated calcium accumulation in cardiomyocytes undergoing anoxia-reoxygenation. It is suggested that SQS could protect cardiomyocytes against oxidative stress induced by anoxia-reoxygenation by attenuating reactive oxygen species generation and increasing activities of endogenous antioxidants.