Synthesis and evaluation of novel triazole–Schiff base derivatives bearing a morpholine moiety as potent urease inhibitors: Molecular docking and molecular dynamics studies

Abstract Urease, a binuclear nickel metalloenzyme central to microbial survival and pathogenicity, is a key protein target for mechanism‐guided inhibitor design. In this study, nine morpholine‐containing 1,2,4‐triazole‐based N ‐Mannich derivatives ( mb1–9 ) were synthesized through sequential Schiff‐base formation and Mannich functionalization and structurally characterized by FT‐IR, 1 H/ 13 C‐NMR, and HRMS analyses. In vitro urease‐inhibition assays revealed marked structure‐dependent activity, with mb4 exhibiting the highest potency (IC 50 = 5.55 μM), followed by mb7 (12.92 μM), both outperforming the reference inhibitor thiourea (IC 50 = 20.84 μM). The experimental activity profile was rationalized by molecular docking and molecular dynamics simulations, which demonstrated that productive inhibition is associated with persistent accommodation of the ligands within the catalytic pocket, coordination of the morpholine oxygen toward the binuclear Ni center, and additional stabilization involving the triazole carbonyl and Cys321. Although the calculated docking scores were relatively close across the series, mb4 adopted a particularly favorable binding geometry that simultaneously preserved metal‐centered interactions and active‐site stabilization, providing a structural basis for its superior experimental potency. Consistently, MD simulations showed that the more active aromatic derivatives, particularly mb4 , mb5 , and mb7 , maintained comparatively stable binding conformations with limited structural fluctuations, whereas less active and more conformationally flexible analogs exhibited greater mobility within the active‐site environment. MM/GBSA analysis further supported favorable binding of mb4 , while diffusion‐map analysis identified a dominant, well‐defined low‐energy conformational basin, indicating persistent ligand retention under dynamic conditions. Membrane‐permeation simulations and ADME predictions additionally suggested physiologically compatible passive diffusion and favorable drug‐like properties. The convergence of experimental inhibition data with metal coordination, active‐site interaction patterns, and dynamic stability identifies mb4 as the most promising mechanistically supported urease‐inhibitory scaffold within the series.