Molecular modeling of methylene blue photosensitizer and its <i>in vivo</i> evaluation against canine fibrosarcoma under light irradiation

Abstract Methylene blue is a widely used photosensitizer in biophotonics, particularly in photodynamic therapy (PDT). In this work, we conducted a comprehensive theoretical investigation of its molecular and electronic properties including frontier molecular orbital energies, electron spin density, theoretical electronic transitions, and reactivity indices to better understand its biomedical potential. These theoretical insights were closely integrated with experimental analyses, including UV–Vis spectral measurements and a preliminary PDT application. Theoretical calculations revealed distinct redox characteristics: the cationic form ([MB]+) exhibited high electron affinity and pronounced electrophilicity, while the neutral form (MB) showed low polarizability and minimal electrophilicity. These electronic features provide a mechanistic basis for methylene blue’s photochemical reactivity and its interaction profile in biological systems. Experimental UV–Vis spectra corroborated the absorption bands predicted by the theoretical models, demonstrating strong agreement between computational and experimental data. Furthermore, after 90 d of PDT treatment, significant clinical improvements were observed in the treated canine subjects, including enhanced wound healing, increased mobility, and improved sociability. The convergence of theoretical and experimental findings reinforces methylene blue’s role as an effective photosensitizer, supporting its potential for safe and impactful therapeutic applications.