Flow cytometry as an alternative method to evaluate genotoxicity of natural cosmetic actives

Abstract

Alternative in vitro methods are important, as there is a call to ban the use of animals in cosmetics research.

Aim: To suggest the expansion of the use of in vitro safety techniques recommended by the OECD guidelines and to propose the use of the automation of the in vitro mammalian micronucleus test method by flow cytometry to assess the genotoxic potential of Centella asiatica, Horse Chestnut, Witch Hazel, Blend, Ecoblend, and Caffeine extracts due to their widespread use in commercial products.

Methods: Flow cytometer analysis was performed using the Accuri™ C6 equipment and analyzed using the FlowJo software. Cytotoxicity tests followed OECD 129 guidelines and Phototoxicity followed OECD/GD 432 guidelines.

Results: The results showed that the cytotoxicity assay presented a decrease in cell viability when cells were exposed to Centella asiatica from a concentration of 5.0%, horse chestnut 2.5%, Witch hazel 2.5%, Blend 3.13%, and Caffeine 3%, while Ecoblend at the tested concentrations did not show cytotoxicity. In the phototoxicity test, the samples at the tested concentrations showed a PIF <2 being considered potentially non-phototoxic. Finally, in the genotoxicity automated assay, samples were considered potentially non-genotoxic.

Conclusion: In vitro methods are of paramount importance for the development of pre-clinical tests and the use of test automation helps to reduce the time for analysis and dissemination of results, being a determining factor for the prospect of new compounds.

Keywords: cell culture; flow cytometry; in vitro test; safety testing.