UPLC-ESI-qTOF-MS metabolomic characterization and in vivo neuroprotective activity of Diplotaxis harra ethanolic extract through antioxidant, anti-cholinesterase, and anti-amyloid mechanisms.

Aluminum chloride (AlCl3)-induced neurotoxicity is a widely used model of Alzheimer's disease, featuring oxidative stress, cholinergic dysfunction, amyloid deposition, and cognitive deficits. This study investigated the neuroprotective effects of Diplotaxis harra ethanolic extract (DH-EtOH) in AlCl3-treated rats and linked them to its phytochemical profile. UPLC-ESI-qTOF-MS analysis revealed a chemically rich extract dominated by phenolic acids, flavonol glycosides (∼40% of metabolites), glucosinolates, amino acid derivatives, phospholipids, and oxidized fatty acids, supporting a multitarget mode of action. Oral DH-EtOH (75-150 mg/kg) improved spatial memory in T-maze tests, with the higher dose restoring near-normal performance. Biochemically, hippocampal acetylcholinesterase activity was significantly inhibited, glutathione levels were restored, and malondialdehyde levels decreased by ∼35% versus AlCl3 controls. Histology showed preserved hippocampal architecture and reduced GFAP immunoreactivity, indicating attenuation of astrocyte activation. Pathological markers Tau and Aβ1-42 were also significantly diminished in a dose-dependent manner. These results demonstrate that DH-EtOH exerts multitarget neuroprotection through synergistic actions of diverse metabolite classes. Its chemical complexity underpins efficacy in restoring cognitive function, reducing oxidative stress, and mitigating Alzheimer-like pathology, highlighting the potential of this medicinal plant as a natural, multifaceted cognitive enhancer.