A new series of compounds containing 4-thiazolidinone pharmacophore 5(a-d) have been synthesized. The chemical structure of the intermediate and final compounds was characterized and confirmed by using FT-IR and 1H-NMR spectroscopy. All final compounds were tested against Gram-positive and Gram-negative bacteria using a well-diffusion technique for their ability as antimicrobial agents. The 5b and 5c compounds tested showed comparable antibacterial activity against Gram-negative bacteria and Gram-positive bacteria, such as Escherichia coli, Acinetobacter bumannii, Staphylococcus aureus, and Streptococcus pyougenes, as a standard drug compared to Trimethoprim. Simulations of the molecular docking of the molecular core were studied. The results have come from an in vitro-based docking of the most active compounds into the active protein site of the bacteria.