Mechanistic in vitro evaluation of surfactant-induced skin irritation: Correlating micellar physicochemistry with 3D reconstructed human epidermis, zein, and ecotoxicity endpoints

The dermal irritation potential of representative anionic, non-ionic, amphoteric, and ethoxylated surfactants was evaluated using three complementary in vitro approaches: the zein protein solubilization assay, reconstructed human epidermis (RhE) cell viability testing, and the Vibrio fischeri bioluminescence inhibition assay to assess ecotoxicity. Surfactants forming small and highly mobile micelles generally tended to exhibit higher zein numbers, reduced RhE viability, and lower EC₅₀ values, indicating increased irritant and toxic potential. In contrast, non-ionic surfactants forming larger and less mobile aggregates showed lower protein solubilization, higher cell viability, and reduced bacterial toxicity. Multivariate principal component analysis demonstrated that micellar diffusivity, with additional contributions from electrostatic character and hydrophilic-lipophilic balance, represents a major physicochemical axis associated with biological variability, whereas micellar size plays a secondary structural role. This integrated framework provides mechanistic insight into surfactant-induced irritation and supports the rational selection and design of safer, more sustainable surfactant systems.