Design, Synthesis, Characterization, in Vitro Biological Evaluation, and in Silico Investigation of 4-Substituted Arylidene Pyrazolones Through Docking, Molecular Dynamics Simulations, and DFT Studies

Abstract

A new series of six 4-substituted arylidene pyrazol-3-one derivatives was successfully designed, synthesized, and evaluated for their antioxidant and antibacterial properties using both experimental and in silico approaches. The pyrazolone nucleus, namely 5-methyl-2-phenyl-4H-pyrazol-3-one (edaravone, Pyr 1) was synthesized via the Knorr reaction between ethyl acetoacetate and phenylhydrazine, followed by Knoevenagel condensation with various substituted benzaldehydes to yield the target 4-substituted arylidene pyrazol-3-one. The structures of the target compounds were characterized using FT-IR and NMR spectroscopy, whereas their thermal behavior was assessed via thermogravimetric analysis (TGA). Antioxidant activity, evaluated through the DPPH radical scavenging assay, revealed that all compounds exhibit significant potential, with Pyr 4 showing the highest activity (98% inhibition, IC₅₀ = 0.25 mg/mL). Antibacterial activity, evaluated using the agar well diffusion method against Escherichia coli, Klebsiella pneumoniae, Listeria monocytogenes, and Acinetobacter baumannii, showed Pyr 5 as the most effective, with inhibition zone of 29 mm against A. baumannii and 28 mm against K. pneumoniae, comparable to or exceeded those of standard antibiotics such as gentamicin. Additionally, in silico studies provided insight into the electronic properties, binding modes, and stability of the ligand–target complexes. ADMET analysis further supported the drug-likeness and favorable pharmacokinetics of selected candidates. Overall, these findings validate the synthesized pyrazolones as promising antioxidant and antibacterial agents.