There has been growing effort in the scientific community to develop new antibiotics to address the major threat of bacterial resistance. One promising approach is the use of metal complexes that provide broader opportunities. Among these systems, polypyridine-ruthenium(II) complexes have received particular attention as drug candidates. Here, we prepared two new ruthenium(II) complexes with the formulation [Ru(DFO)(phtpy-R)Cl](PF6), where phtpy = 4'-phenyl-2,2':6',2''-terpyridine; R = -H(MPD1), -CH3(MPD2); and DFO = 4,5-diazafluoren-9-one, and investigated their chemical, biochemical and antibacterial activities. These compounds exhibit photoreactivity and produce reactive oxygen species (ROSs). Photogeneration of singlet oxygen (1O2) was measured in acetonitrile with significant quantum yields using blue light, Φ = 0.40 and 0.39 for MDP1 and MPD2, respectively. Further studies have shown that MPD1 and MPD2 can generate superoxide radicals. Antibacterial assays demonstrated a significant enhancement in MIC (minimum inhibitory concentration) upon blue light irradiation (>32-fold), with MICs of 15.6 μg mL-1 (S. aureus, ATCC 700698) and 3.9 μg mL-1 (S. epidermidis, ATCC 35984) for both metal complexes. Interestingly, an MIC of 15.6 μg mL-1 for MPD1 and MPD2 was observed against S. epidermidis ATCC 12228 under red light irradiation. The latter results are encouraging, considering that red light penetrates deeper into the skin. In addition, no significant cytotoxicity was observed in some mammalian cells, even upon light irradiation, supporting their potential safety. Altogether, these data show evidence of the potential use of these compounds as antimicrobial photodynamic therapeutic agents, enriching our arsenal to combat this worldwide bacterial threat.