Phenazinones represent a foundational class of nitrogen-containing heterocyclic redox agents that are highly recognized for their exceptional antibiotic, antifungal, and cellular modulating characteristics. In this study, a novel series of substituted 3-bromo-1,4-dimethoxyphenazin-2(10H)-one derivatives 3(a-d) was successfully synthesized through a direct, base-catalyzed condensation approach. The synthesis involved the reaction of 2,5-dibromo-3,6-dimethoxy-1,4-benzoquinone with various substituted ortho-phenylenediamines in an ethanol medium, using anhydrous sodium acetate as a basic catalyst under reflux conditions for 3 hours. The target phenazinone compounds were isolated in high purity via silica gel column chromatography and characterized systematically. All the newly synthesized derivatives were thoroughly evaluated for their in vitro antibacterial efficacy against a selected panel of human pathogens, including four Gram-negative strains (Escherichia coli, Salmonella paratyphi, and Klebsiella pneumoniae) and three Gram-positive strains (Staphylococcus aureus, Micrococcus luteus and Bacillus cereus) using the agar disc diffusion methodology at multiple concentrations (400, 600, and 800 µg/ml). The biological assays revealed that the derivative carrying a chlorine substitution (3b) exhibited the most potent antibacterial profile, particularly demonstrating a remarkable zone of inhibition of 22–23 mm against Escherichia coli, which was highly comparable to the standard antibiotic Streptomycin. Other derivatives, including the unsubstituted (3a), bromo-substituted (3c), and methyl-substituted (3d) analogues, showed moderate to strong broad-spectrum inhibitory potential across both Gram-positive and Gram-negative classes. These findings reveal that the incorporation of halogen groups on the phenazinone core significantly boosts cell membrane penetration and redox-disrupting capabilities, establishing these scaffolds as promising leads for downstream antimicrobial drug development.