The alarming escalation of multidrug-resistant (MDR) bacterial strains has grown into a severe global threat to human health, creating an urgent and mandatory requirement for the structural design and development of novel scaffold-based antimicrobial agents. Phenazine and imidazole heterocyclic cores represent privileged, highly effective structures in medicinal chemistry due to their exceptionally broad biological profiles, which include potent DNA-intercalating capabilities, cellular respiration disruption, and specific bacterial enzyme-inhibiting functions. In the present study, a novel series of 4,11-dimethoxy-1H-imidazo(4,5-b)phenazin-2(5H)-one derivatives 3(a–d) successfully and efficiently synthesized through a synthetic pathway starting from functionalized 3-bromo-1,4-dimethoxyphenazin-2(10H)-ones 1(a–d) and ortho-phenylenediamines 2(a-d). All the newly synthesized target hybrids were thoroughly and unambiguously structurally characterized using Fourier Transform Infrared (FT-IR), Proton Nuclear Magnetic Resonance (1H NMR), Carbon-13 Nuclear Magnetic Resonance (13C NMR), and Electrospray Ionization Mass Spectrometry (ESI-MS). The in vitro antibacterial efficacy of these tetracyclic chemical entities was systematically evaluated against a panel of clinically relevant Gram-negative bacterial strains Escherichia coli, Salmonella paratyphi, Klebsiella pneumoniae and Gram-positive bacterial strains Staphylococcus aureus, Micrococcus luteus, Bacillus cereus utilizing the standard agar disk diffusion bioassay. Among the screened chemical library, compounds bearing electron-withdrawing halogen substitutions, specifically the 7-chloro (3b) and 7-bromo (3c) variants, demonstrated exceptional antibacterial activity profiles, closely matching the standard reference antibiotic Streptomycin against several resistant strains. These definitive outcomes strongly highlight the novel 4,11-dimethoxy-1H-imidazo(4,5-b)phenazin-2(5H)-one framework as a highly promising lead molecular architecture for future anti-infective drug development and clinical optimization programs.