Rational design of phenyl 2,4,5-trichlorobenzenesulfonate based thiosemicarbazones as α-glucosidase and α-amylase inhibitors: integrating enzymatic evaluation and molecular modeling

The present study aimed to investigate the antidiabetic potential of a new series of thiosemicarbazone derivatives through integrated <i>in vitro</i> enzymatic assays and <i>in silico</i> molecular modeling. The synthesized compounds were evaluated for their inhibitory activities against α-glucosidase (α-Glu) and α-amylase (α-Amy) enzymes. Among the tested derivatives, compound 16 (2-chlorophenyl-substituted) demonstrated the most potent dual inhibition with IC₅₀ values of 14.58 nM (α-Glu) and 88.37 nM (α-Amy), surpassing the reference drug acarbose in potency. Molecular docking analyses revealed that compound 16 formed stable interactions with Asn-214, Glu-276, Phe-157, and Tyr-71 in the α-Glu and Asp-197, Glu-233, and Lys-200 in α-Amy's active site. These key interactions were further supported by 250 ns molecular dynamics simulations, confirming the conformational stability of both complexes with average RMSD values below 2.0 Å and minimal ligand fluctuations. Energy decomposition analysis indicated that van der Waals and electrostatic interactions were the major contributors to the overall binding free energy. <i>In silico</i> ADME profiling predicted favorable pharmacokinetic properties, including high gastrointestinal absorption, good oral bioavailability, and compliance with Lipinski's rule of five, while no significant blood-brain barrier penetration was observed. The combined <i>in vitro</i> and <i>in silico</i> findings highlight compound 16 as a promising lead candidate for further optimization and development as a dual α-Glu and α-Amy inhibitor for the management of type 2 diabetes mellitus.