Chlorophyll-Loaded Castor Oil Nanoemulsions Exhibit Photodynamic Therapy Efficacy Against B16-F10 Melanoma with Low Cytotoxicity Toward HaCaT Keratinocytes.

Background: Photodynamic therapy (PDT) is a promising minimally invasive approach for melanoma; however, many photosensitizers lose activity in aqueous media due to aggregation-induced quenching effects. Objectives: The aim of this study was to develop and characterize castor oil-based nanoemulsions containing chlorophyll (NFs-Chl) and to evaluate their in vitro photodynamic potential against melanoma cells (B16-F10), as well as their selectivity compared with human keratinocytes (HaCaT). Methods: NFs-Chl were prepared by spontaneous emulsification. Physicochemical characterization was carried out using dynamic light scattering (DLS), UV-Vis spectroscopy, FTIR, and Raman spectroscopy. In vitro assays included MTT for cell viability (IC50 determination), real-time cell proliferation (RealTime-Glo™), and cell migration analysis (scratch assay). All photodynamic treatments were performed under irradiation at 660 nm. Results: NFs-Chl exhibited homogeneous nanometric sizes (≈24-31 nm) and a low polydispersity index (≈0.25-0.40), indicating a narrow size distribution. UV-Vis spectra confirmed the preservation of the characteristic absorption peaks of chlorophyll after encapsulation. In B16-F10 cells, NFs-Chl associated with PDT significantly reduced cell viability and metabolic activity over 48 h. Furthermore, NFs-Chl inhibited the migratory capacity of B16-F10 cancer cells. Cell migration assays revealed a clear inhibition of B16-F10 cell migration following treatment with NFs-Chl + PDT. Conclusions: Encapsulation of chlorophyll into castor oil nanoemulsions protected the photosensitizer, improved its cellular delivery, and enhanced its photodynamic cytotoxic effect against melanoma cells, while relatively preserving normal keratinocytes in vitro.