Ilex pubescens is commonly used in traditional Chinese medicine to treat cardiovascular and cerebrovascular diseases, such as coronary artery disease and stroke. However, the underlying mechanisms remain to be fully elucidated. The aim of the present study was to investigate the effects of Ilex pubescens total flavonoids (IPTF) on neuroprotection and the potential mechanisms in a rat model of focal cerebral ischemia/reperfusion (I/R) injury. Rats were pretreated with intragastric administration of IPTF at doses of 200 and 100 mg/kg for 5 days; middle cerebral artery occlusion surgery was then performed to induce cerebral I/R injury. Neurological deficits were determined using the 5‑point neurological function score evaluation system, brain infarct sizes were determined by 2,3,5‑triphenyltetrazolium chloride staining and alterations in brain histology were determined by hematoxylin and eosin staining. The neurological deficit score, the infarcted area and the brain tissue pathological injury were significantly reduced when the rats were pretreated with IPTF. In addition, inflammatory mediators and neurotrophic factors in the brain were investigated. IPTF pretreatment decreased the activities of total nitric oxide synthase (TNOS), induced NOS (iNOS) and constitutive NOS (cNOS), and the levels of nitric oxide (NO), interleukin‑1β (IL‑1β) and tumor necrosis factor‑α (TNF‑α), however, it increased the levels of IL‑10 in brain tissues. Furthermore, pretreatment with IPTF also increased the protein expressions of brain‑derived neurotrophic factor, glial cell‑derived neurotrophic factor and vascular endothelial growth factor, when compared with the model group. In conclusion, the results of the present study demonstrated that IPTF has a neuroprotective effect against focal cerebral I/R injury in rats. The mechanism may be associated with the decreased production of certain proinflammatory cytokines including NO, IL‑1β, TNF‑α, TNOS, iNOS and cNOS, the increased production of the anti‑inflammatory cytokine IL‑10 and the increased secretion of neurotrophic factors.