In this study, in-depth observation and investigation of blood-brain barrier permeability and neuroprotective effect of Trichosanthes kirilowii cassia twig particles on rats with cerebral ischemia-reperfusion injury were performed. Focal cerebral ischemia-reperfusion injury model was established by middle cerebral artery occlusion method, reperfusion was implemented 2 hours after ischemia; qualitative analysis and investigation of trichosanthes kirilowii cassia twig particles in plasma, brain tissue and cerebrospinal fluid in normal and middle cerebral artery occlusion (MCAO) rats were done by high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS); changes in neurological deficits, cerebral infarction stereometry, blood-brain barrier permeability and histopathological changes of MCAO model rats were observed. Qualitative analysis by HPLC-MS/MS results showed that ingredients, paeoniflorin, albiflorin, liquiritin in Trichosanthes kirilowii cassia twig particles can reach the brain through the blood-brain barrier. In the model group, glycyrrhizin and glycyrrhizic acid can be detected in brain tissue or cerebrospinal fluid. In addition, Trichosanthes kirilowii cassia twig particles can significantly lower neurological deficits of rats in middle cerebral artery occlusion model, reduce the Evans blue penetration, contract infarct size, and reduce pathological tissue injury of cerebral ischemia reperfusion. The ingredients of Trichosanthes kirilowii cassia twig particles can reach the brain tissue through the blood-brain barrier and play a role in neuroprotection of rats with cerebral ischemia-reperfusion injury, which has important research significance and brings scientific experimental, theoretical basis for clinical drug use.