Unnatural amino acid compounds as potent multi-target inhibitors of aldose reductase, α-glucosidase, and α-amylase: integrated in vitro, SAR, and molecular dynamics insights

Abstract Diabetes mellitus is a multifactorial metabolic disorder in which sustained post-prandial hyperglycaemia and aberrant activation of the polyol pathway contribute to disease progression and long-term complications. Simultaneous modulation of digestive enzymes and aldose reductase (ALR2) therefore represents a rational multitarget therapeutic strategy. In this study, a series of previously reported aryl-substituted unnatural N -methoxysulfonyl β-ketoester derivatives were investigated for their inhibitory potential against ALR2, α-glucosidase, and α-amylase. Compound 1i exhibited the strongest ALR2 inhibition with K i :0.493 ± 0.155 µM and IC 50 : 1.638 ± 0.44 µM. For α-glucosidase, compound 1h showed the highest potency (K i :1.341 ± 0.181 µM), while compound 1j demonstrated strong α-amylase inhibition (IC 50 : 1.361 ± 0.26 µM). The compounds were evaluated through in vitro enzyme inhibition assays supported by comprehensive structure–activity relationship (SAR) analysis. Several derivatives displayed pronounced inhibitory activity, achieving sub-micromolar K i values against ALR2 and low-micromolar inhibition of α-glucosidase and α-amylase, in some cases surpassing reference inhibitors. SAR analysis revealed that ALR2 inhibition is strongly governed by planar aromatic expansion and conformational rigidity, whereas α-glucosidase and α-amylase preferentially accommodate compact, hydrophobically enriched substituents, highlighting distinct steric and electronic requirements across targets. A strong correlation was observed between experimental and docking-derived K i values, supporting the consistency of the computational protocol. Subsequent 100 ns molecular dynamics simulations confirmed the stability of the most active ligand–enzyme complexes under physiological conditions. Overall, this integrated experimental and computational evaluation identifies unnatural N -methoxysulfonyl β-ketoester derivatives as promising multitarget antidiabetic lead scaffolds.