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.