Background <p>Alzheimer’s disease (AD) is an irrevocable neurodegenerative disorder that has caught global attention as the root cause of dementia. <i>Polyalthia suberosa</i> (Roxb.) Thwaites was reported to exhibit acetylcholinesterase inhibitory activity. To date, its potential protective effects against memory impairment have not been explored.</p> Methods <p>This study focused on evaluating the neuroprotective effect of <i>Polyalthia suberosa</i> leaf methanol extract (PSTE) and its dichloromethane soluble fraction (PSDF) against scopolamine (SCO)-induced memory impairment rat model. Additionally, metabolic profiling of PSTE and PSDF was carried out using Ultra-Performance Liquid Chromatography-Electrospray Ionization Mass Spectrometry (UPLC-ESI-MS/MS). Anticholinesterase activity of PSTE and PSDF was evaluated in vitro. <i>An in vivo</i> study was conducted. Behavioral testing was performed using the elevated plus maze (EPM) and novel object recognition (NOR) tests. Additionally, histopathological examination, acetylcholinesterase activity, and levels were assessed.</p> Results <p>Results showed that both PSTE and PSDF displayed promising anticholinesterase activity in vitro, which was also confirmed by in vivo results where PSTE and PSDF at a dose 150&#xa0;mg/kg were able to normalize the elevated acetylcholinesterase enzyme (AChE) level and activity in hippocampus tissue and minimized neurodegeneration in the brain tissue histopathologically assessed. The low dose of PSDF exhibited noticeable amelioration to SCO-induced memory deficit as evidenced by the significant elevation in the discrimination index and inflexion ratio in both NOR and EPM tests. Moreover, Metabolic profiling of PSTE and PSDF resulted in the tentative identification of 69 compounds, with alkaloids being the major class.</p> Conclusion <p>This unprecedented study highlights the substantial use of <i>Polyalthia suberosa</i> leaves as a natural neuroprotective adjuvant against AD.</p>