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.

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 Ki :0.493 ± 0.155 µM and IC50: 1.638 ± 0.44 µM. For α-glucosidase, compound 1h showed the highest potency (Ki :1.341 ± 0.181 µM), while compound 1j demonstrated strong α-amylase inhibition (IC50: 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 Ki 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 Ki 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.