Design and Synthesis of Aromatic Sulfonohydrazides as Antibacterial Agents: Integrated In Vitro , Biofilm, Efflux Pump Inhibition, and Computational Studies

In this study, a series of N’-phenylmethylidene benzene-sulfonohydrazide derivatives (C1–C17) was designed and synthesized. The antibacterial activities of these compounds were systematically evaluated in vitro against selected Gram-positive and Gram-negative bacterial strains using the broth microdilution method. Among the synthesized derivatives, compounds C1, C14, and C17 exhibited significant inhibitory activity against Streptococcus pneumoniae MTCC 655, with minimum inhibitory concentrations (MICs) ranging between 9.93 and 14.60 µg/mL, in comparison to ciprofloxacin (MIC: 6.85 µg/mL). In addition, compound C13 exhibited notable activity against Pseudomonas aeruginosa, with MIC of 4.30 µg/mL Minimum bactericidal concentration (MBC) assays further confirmed that compounds C1 and C14 exerted bactericidal effects on S. pneumoniae MTCC 655 at both MIC and sub-MIC concentrations after 12 h of exposure. Consistent with these findings, time–kill kinetic studies demonstrated that compounds C1, C14, and C17 achieved complete eradication of S. pneumoniae MTCC 655 within 24 h. Additionally, compounds C1–C17 was evaluated for cytotoxicity to assess their safety profile. Three compounds C1, C14, and C17 were also screened for their ability to inhibit biofilm formation and efflux pump activity. Docking results suggest that coordination with Mg460 at the N-terminal uridyltransferase active site is crucial for the binding of compounds C1-C17 in the S. pneumoniae GlmU catalytic pocket. In silico predicted ADME profile of compounds C1–C17 indicates no CNS activity, high oral absorption, no HERG-related cardiotoxicity, and no violation of Lipinski’s rule of five. Collectively, C1, C14, and C17 emerged as promising scaffolds for the design of broad-spectrum antibacterial agents.