Metabolomics Integrated with Mass Spectrometry Imaging Reveals Novel Action of Rb1 in Ischemic Stroke

, has demonstrated neuroprotective activity, but its metabolic mechanisms remain incompletely defined. Here, we combined lipid metabolomics with matrix-assisted laser desorption/ionization mass spectrometry imaging (MALDI-MSI) to investigate the spatially resolved metabolic effects of Rb1 in a rat model of cerebral ischemia/reperfusion injury. Rb1 treatment significantly reduced infarct volume and improved neurological outcomes. Metabolomic profiling revealed that Rb1 reversed ischemia-induced disturbances in the glycerophospholipid and amino acid metabolism, while MSI demonstrated recovery of phosphatidic acid, phosphatidylcholine, phosphatidylethanolamine, and phosphatidylserine distributions in the ischemic cortex. These metabolic improvements were strongly correlated with reduced levels of pro-inflammatory cytokines (TNF-α, IL-6, IL-1β) and oxidative stress marker malondialdehyde, along with increased superoxide dismutase activity. Rb1 also preserved blood-brain barrier integrity by enhancing the expression of tight-junction proteins ZO-1 and Occludin. Together, these findings indicate that Rb1 confers neuroprotection through metabolic reprogramming linked to anti-inflammatory and antioxidant actions, and they highlight the value of integrating metabolomics with MSI to elucidate spatially defined drug mechanisms in the brain.