A novel series of 4-(arylideneamino)-N'-((2-chloro-8-methylquinolin-3-yl)methylene)-3-phenyl-2-thioxo-2,3-dihydrothiazole-5-carbohydrazides (5a-o) bearing biologically active 2-chloroquinoline-3-carbaldehyde and 2-thioxothiazole ring have been synthesized. The structures of synthesized compounds were established by spectroscopic techniques (IR, 1H NMR, 13C NMR and mass spectrometry). Antimicrobial screening of compounds (5a-o) was done against Gram-positive bacteria (Staphylococcus aureus and Streptococcus pyogenes), Gram-negative bacteria (Escherichia coli and Pseudomonas aeruginosa) and fungi (Candida albicans, Aspergillus niger and Aspergillus clavatus) using serial broth dilution method. SAR for the series was developed by comparing their MIC values with ampicillin and griseofulvin. Some of the compounds from the series like 5o were found to be most active at MIC 12.5 μg/mL against P. Aeruginosa, and compound 5k was found to be most active at MIC 12.5 μg/mL against A. clavatus.