The Role of Emulsification and Composition in Modulating Cell Type-Dependent Cytotoxicity in o/w nanoemulsion

Background Nanoemulsions (NEs) are emerging as effective drug delivery systems for poorly water-soluble drugs, enhancing solubility and bioavailability. However, concerns regarding their cytotoxicity persist, and systematic evaluations across different cell types remain limited. Methods NE–Et and NE–PEG were prepared with identical compositions except for the cosurfactant. Cytotoxicity was evaluated using standard viability assays at full-strength (100%) and at actual NE concentrations (NE%). Individual components, binary and ternary mixtures, and complete NEs were tested. The effect of dilution medium (deionized water vs. DMEM) on NE cytotoxicity was examined. Results Both NEs displayed concentration-dependent cytotoxicity. In Caco-2 cells, individual surfactants and almond oil exhibited high cytotoxicity, whereas complete NEs demonstrated higher cell viability (~ 47–64% at NE%), indicating the protective effects of emulsification. Ethanol- and PEG400-based mixtures showed composition-dependent responses. In L929 cells, surfactant-containing mixtures were most toxic, while oil–PEG and oil–water systems maintained moderate to high viability (~ 60–70% at NE%). No significant differences in cytotoxicity were observed between deionized water (DI) and DMEM-diluted NEs. Conclusions Emulsification mitigates the cytotoxicity of individual NE components. Both the type of cosurfactant and formulation composition critically affect cell-specific responses. These results highlight the importance of rational cosurfactant selection and component organization in designing safe and biocompatible NE-based drug delivery systems.