Design, Synthesis and Biological Evaluation of Rhodanine-Based Hydrazide Derivatives as Antibacterial, Antifungal, and Antitubercular Agents

Background/Objectives: The rapid emergence of antimicrobial resistance has created an urgent need for new chemotypes with activity against bacterial, fungal, and mycobacterial pathogens. Rhodanine-based compounds are recognized as privileged scaffolds in medicinal chemistry because of their broad spectrum of biological activities, making them attractive candidates for the development of novel antimicrobial agents. Methods: A library of thirty-two novel rhodanine-based hydrazide derivatives (21–52) was synthesized by condensation of 5-ethoxymethylidenerhodanine derivatives with aromatic acid hydrazides and characterized by IR, 1H NMR, and 13C NMR spectroscopy. Their drug-likeness and pharmacokinetics properties were evaluated using SwissADME, while antimicrobial activity was assessed against Gram-positive and Gram-negative bacteria, Candida spp., and both drug-susceptible and drug-resistant Mycobacterium tuberculosis strains using minimum inhibitory concentration (MIC) assays. Results: The synthesized compounds demonstrated selective activity against Gram-positive bacteria, whereas only limited activity was observed against Gram-negative species. The most potent derivatives (36, 40, 46, and 50) inhibited Micrococcus luteus with MIC values as low as 31.3 mg/L. Several compounds also demonstrated noteworthy antifungal activity, particularly against Candida parapsilosis (MIC = 15.6–31.3 µg/mL). Pyridyl-containing analogues displayed moderate antitubercular activity and, importantly, compounds 49 and 52 retaining activity against both drug-susceptible and drug-resistant M. tuberculosis strains. In silico analysis indicated favorable drug-like properties for most compounds, with good compliance with Lipinski’s rule of five and predicted oral bioavailability. Preliminary structure–activity relationship analysis revealed that a 3-phenylrhodanine core combined with para-halogen or para-nitro substituents significantly enhanced antimicrobial potency. Conclusions: The present study identifies rhodanine-based hydrazide derivatives as promising multifunctional antimicrobial scaffolds with activity against Gram-positive bacteria, pathogenic yeasts, and M. tuberculosis. The identified structure–activity relationships provide a rational basis for further lead optimization and support continued development of this chemotype as a potential source of new antimicrobial agents.