In Vivo and Bioinformatics Evaluation of Nine Traditional Chinese Medicine Compounds Targeting Acetylcholinesterase and Butyrylcholinesterase Enzymes in Alzheimer's Disease.

Background: Alzheimer's disease (AD) is characterized by cholinergic dysfunction and oxidative stress, creating a need for the development of new therapeutic agents. Traditional Chinese medicine (TCM) compounds, such as hainanolidol and norwogonin, have shown potential neuroprotective activity.

Objectives: This study investigates the inhibitory action of these compounds on cholinesterase enzymes and their possible therapeutic application in an AD-like rat model generated by lead acetate (PbAc), a well-known neurotoxicant that mimics AD-associated oxidative damage and cognitive decline.

Methods: The cholinesterase inhibitory activity of nine TCM compounds was assessed in vitro against acetylcholinesterase (AChE) and butyrylcholinesterase (BuChE), with donepezil and tacrine as controls. Hainanolidol and norwogonin, identified as the most potent inhibitors, were further evaluated in a PbAc-induced AD rat model to assess their neuroprotective effects. Oxidative stress biomarkers malondialdehyde (MDA), glutathione (GSH), cholinesterase activity, and in silico molecular interactions were analyzed, including docking studies, molecular dynamics (MD) simulations, and ADME-toxicity profiling.

Results: Hainanolidol and norwogonin showed strong nanomolar-range inhibition of both AChE and BuChE, with considerable IC₅₀ values superior to those of standard inhibitors. In the PbAc-induced AD model, both compounds significantly reduced MDA levels and increased GSH levels, indicating oxidative stress mitigation.The level of cholinesterase activity inhibition was as effective as, or more effective than, the suppression achieved by standard treatments, particularly regarding AChE, thus suggesting enhanced therapeutic potential compared to donepezil. Molecular docking and MD simulations confirmed stable binding interactions with key catalytic residues of AChE and BuChE, reinforcing their inhibitory mechanisms. ADME-toxicity analysis further demonstrated favorable pharmacokinetics and safety profiles.

Conclusions: This study concludes that both hainanolidol and norwogonin are worthy of being regarded as dual cholinesterase inhibitors with antioxidant properties, which may serve as alternative therapeutics for AD. The use of PbAc as an AD model underscores the role of environmental neurotoxins in disease pathogenesis, offering insights into novel intervention strategies. Advanced preclinical and clinical studies are needed for further validation.