Oxidative stress is implicated in the pathogenesis of ischemic brain injury. Flavonoids from various herbal extracts have been shown to be neuroprotective in experimental models of cerebral ischemia/reperfusion (I/R). The present study was designed to investigate the neuroprotective effect of the biflavone rich fraction from Araucaria bidwillii Hook (ABH) (Family: Araucariaceae) in I/R induced oxidative stress. The I/R was induced by occluding bilateral common carotid arteries (BCCAO) for 30 min, followed by 24 h reperfusion. BCCAO caused significant depletion in superoxide dismutase (SOD), catalase (CAT), glutathione (GSH) and significant increase in lipid peroxidation (LPO) in various brain regions. The neurological deficit and sensory motor function were also decreased significantly by BCCAO group as compared to sham group animals. All the alteration induced by cerebral ischemia was significantly attenuated by 7 days’ pretreatment with biflavone fraction (BFR) at the dose of 100 and 200 mg/kg, comparable to that given by Vitamin E (200 mg/kg). Consistent with neurobehavioral deficits, pretreatment with biflavones at higher doses significantly reduced ischemia-induced neuronal loss of the brain. In conclusion the biflavone rich fraction from A. bidwillii was found to protect rat brain against I/R induced oxidative stress, and attributable to its antioxidant properties.