Isovanillin-derived bis-hydrazones as dual cholinesterase and carbonic anhydrase inhibitors: synthesis, enzymatic profiling, and computational insights from molecular docking and dynamics

Abstract

To develop isovanillin-based bis-hydrazones as multitarget inhibitors of acetylcholinesterase (AChE), butyrylcholinesterase (BChE), and human carbonic anhydrase I/II ( h CA I/II). Twelve bis-hydrazones ( 4a–4l ) were synthesized in two steps and evaluated by spectrophotometric enzyme assays, Lineweaver–Burk kinetics, molecular docking, MM-GBSA, molecular dynamics simulations, and in silico ADME/Tox profiling. All compounds showed nanomolar inhibition. Compound 4d was the most potent AChE/BChE inhibitor ( K I = 10.46 and 3.56 nM), while 4a and 4j led the h CA I/II panel ( K I = 3.46 and 16.12 nM). Docking, MM-GBSA, and molecular dynamics supported dual-site cholinesterase engagement and non-zinc, peripherally anchored h CA inhibition. Isovanillin-based bis-hydrazones, particularly 4d, 4a , and 4j , represent promising multitarget leads for cholinergic and h CA-linked disorders.