The increasing interest in clinical bacterial photodynamic inactivation has led to the search for photosensitizers with higher bactericidal efficiency and less side effects on the surrounding tissues. We present a novel nonionic porphyrin, the 5,10,15-tris(2,6-dichlorophenyl)-20-[4-N-(6-amino-hexyl)sulfonamido)phenyl]-porphyrin (ACS769F4) with substantial improvements in the efficiency of nonionic sensitizers. This porphyrin causes eradication of both Escherichia coli and Staphylococcus aureus by the photodynamic effect but in higher concentrations compared with 5,10,15,20-tetrakis (4-N,N,N-trimethylammoniumphenyl)-porphyrin p-tosylate (TTAP4+), a known bactericidal tetracationic porphyrin. More important, under such conditions, ACS769F4 proved to be harmless to two mammalian cells lines (human embryonic and baby hamster kidney), causing no reduction in their viability or negative impact on their cytoskeleton, despite its accumulation in cellular structures. On the contrary, TTAP4+ is shown to accumulate in the nucleus of mammalian cells, in association to DNA, causing chromatin condensation after exposure to light. Furthermore, dark incubation with TTAP4+ was shown to have a deleterious effect on the microtubule network. Based on its bactericidal efficiency, also observed without exposure to light, and on the low tendency to be harmful or genotoxic to mammalian cells, ACS769F4 should be looked at as an interesting photosensitizer to be evaluated for clinical purposes.