Photodynamic Inactivation of Bacteria with Porphyrin Derivatives: Effect of Charge, Lipophilicity, ROS Generation, and Cellular Uptake on Their Biological Activity In Vitro

Abstract

Resistance of microorganisms to antibiotics has led to research on various therapeutic strategies with different mechanisms of action, including photodynamic inactivation (PDI). In this work, we evaluated a cationic, neutral, and anionic meso-tetraphenylporphyrin derivative’s ability to inactivate the Gram-negative and Gram-positive bacteria in a planktonic suspension under blue light irradiation. The spectroscopic, physicochemical, redox properties, as well as reactive oxygen species (ROS) generation capacity by a set of photosensitizers varying in lipophilicity were investigated. The theoretical calculations were performed to explain the distribution of the molecular charges in the evaluated compounds. Moreover, logP partition coefficients, cellular uptake, and phototoxicity of the photosensitizers towards bacteria were determined. The role of a specific microbial efflux pump inhibitor, verapamil hydrochloride, in PDI was also studied. The results showed that E. coli exhibited higher resistance to PDI than S. aureus (3–5 logs) with low light doses (1–10 J/cm2). In turn, the prolongation of irradiation (up to 100 J/cm2) remarkably improved the inactivation of pathogens (up to 7 logs) and revealed the importance of photosensitizer photostability. The PDI potentiation occurs after the addition of KI (more than 3 logs extra killing). Verapamil increased the uptake of photosensitizers (especially in E. coli) due to efflux pump inhibition. This effect suggests that PDI is mediated by ROS, the electrostatic charge interaction, and the efflux of photosensitizers (PSs) regulated by multidrug-resistance (MDR) systems. Thus, MDR inhibition combined with PDI gives opportunities to treat more resistant bacteria.

Photoinactivation of Pseudomonas aeruginosa Biofilm by Dicationic Diaryl-Porphyrin

“…The high yield of binding of cationic diaryl-porphyrins (80-100%) could be ascribable to the electrostatic force displayed between negative-charged lipopolysaccharides on the outer layer of the outer membrane and positively charged PSs. As previously reported by other authors, one or more positive charges are required on the PS structure for a good interaction with Gram-negative bacteria [43]. The mechanism of binding of cationic PSs with bacteria is the so-called "self-promoted uptake" pathway.…”

“…The factor that could affect the potential activity of PSs is the capacity of PSs to bind and penetrate the cell wall and, possibly, reach cytoplasmic targets. In this regard, Sulek reported that cationic TMPyP was not efficient in photoinactivating E. coli cells, and it was necessary to administer verapamil, an efflux pump inhibitor, to potentiate the antibacterial effect [43]. The inhibitor could increase PS's accumulation in the bacteria cell of molecules attached to cell wall via electrostatic interactions.…”

“…This pathway involves the binding of the cationic molecules to LPS that results in the progressive displacement of divalent cations (Ca 2+ , Mg 2+ ) electrostatically bound to the LPS, thereby weakening the outer membrane. The destabilization of the LPS coat results in the formation of "cracks" in the permeability barrier, and divalent cations neutralize the negative character of cell membrane and eliminate electrostatic repulsion between PS and the bacterial envelope [43]. Furthermore, a partial hydrophobic feature associated to C4 and C8 substituents of P12 and P13 could facilitate the cross of the cytoplasmic membrane.…”

“…Furthermore, a partial hydrophobic feature associated to C4 and C8 substituents of P12 and P13 could facilitate the cross of the cytoplasmic membrane. A certain degree of amphiphilicity was reported as peculiar and advantageous for porphyrins in photodynamic applications [43]. Neutral PSs that do not interact with P. aeruginosa, neither affect its viability upon dark incubation.…”

Advances in the Chemistry of Porphyrins and Related Macrocycles

“…This class of compounds is especially versatile, being engaged in a wide multitude of today’s core applications ranging from biology to medicine and material science. The Special Issue entitled “Advances in the Chemistry of Porphyrins and Related Macrocycles“ collects featured research papers providing an overview of the recent developments and advances related to some topic aspects of porphyrin and porphyrinoid chemistry, including biological and biomedical applications [ 1 , 2 , 3 , 4 , 5 , 6 , 7 ], photophysics [ 8 ], supramolecular chemistry [ 9 ], and sensing materials [ 10 , 11 , 12 ]. The Special Issue welcomed the submission of original full research articles [ 1 , 3 , 4 , 6 , 7 , 8 , 9 , 10 , 11 ], as well as some review articles [ 2 , 5 , 12 ] aiming to provide a comprehensive foundation on some of the above-mentioned research field, highlighting the current state of knowledge.…”