Antibiotic-resistant pathogens, such as methicillin resistant Staphylococcus aureus (MRSA), pose significant challenges in managing chronic wound infections. Antibacterial photodynamic inactivation (aPDI) has emerged as an alternative to antibiotics by offering localized and resistance-independent bacterial elimination. Natural compounds with photosensitizing properties are particularly attractive due to their biocompatibility and sustainability. Therefore, this study investigates the potential of aPDI using riboflavin (RB), a vitamin and vital micronutrient commonly found in various animals and plants. RB is known to be a promising natural photosensitizer, capable of causing bacterial cell damage by reactive oxygen species (ROS) generation. Additionally, the potential synergistic combinations with the commonly used wound antiseptics, namely octenidine dihydrochloride (OCT) and polyhexamethylene biguanide (PHMB) against S. aureus clinical strain (MJMC568-B: MRSA) was assessed. The approach aimed to improve antiseptic effiency by reducing the necessary concentrations. The antibacterial efficacy of RB, OCT and PHMB were determined independently by minimum inhibitory concentration (MIC) and minimum bactericidal concentration (MBC). The photodynamic activity of RB was assessed by irradiating with blue light (420 nm) using a light-emitting diode (LED) system and quantification through colony-forming unit (CFU) analysis. An evaluation of the potential synergistic interactions of RB-mediated aPDI, with OCT and PHMB was conducted by a disk diffusion assay, and a checkerboard assay. The results highlight the potential of combining aPDI with antiseptics to achieve enhanced antibacterial effects against S. aureus. This dual approach could offer a promising solution for managing chronic wound infections, reducing the reliance on traditional antibiotics, and mitigating the risk of resistance development.

Acknowledgments: This work was supported by:Project InnovAntiBiofilm(ref. 101157363) financed by European Commission(Horizon-Widera 2023-Acess-02/Horizon-CSA); LEPABE, UIDB/00511/2020((DOI: 10.54499/UIDB/00511/2020)) and UIDP/00511/2020(DOI: 10.54499/UIDP/00511/2020); ALiCE, LA/P/0045/2020(DOI: 10.54499/LA/P/0045/2020); funded by national funds through the FCT/MCTES(PIDDAC; Lisbon, Portugal). Lília S. Teixeira acknowledges individual PhD fellowship from FCT(2023.04774.BDANA).