The effects of co-exposure to methyl paraben and dibutyl phthalate on cell line derived from human skin.

Data on the cumulative effects of chemical substances are necessary for the proper risk assessment, but their availability is still insufficient. The aim of the study was to evaluate the cytotoxic effect of methyl paraben (MePB) and dibutyl phthalate (DBP) on the cells of the skin line (A431) and to compare the cytotoxic effects of the tested substances after single application to A431 cells with the effects of an equimolar/equitoxic (1:1) binary mixture of these compounds as well as their mixtures in ratio 1:3: and 3:1. On the basis of the obtained results, it was found that there were interactions between the tested compounds in terms of cytotoxic effect on A431, assessed on the basis of metabolic activity of cells (MTT test) and integrity of their cell membranes (NRU test). The obtained values of synergy coefficients (SI) and isobolographic analysis indicate that between the tested chemicals in a two-component equimolar mixture (1:1) there is a synergism of action, which, at a high DBP content in the mixture (> 50%) turned into antagonism. Observations using a holotomographic microscope show morphological changes in A431 cells after exposure to both DBP and MePB separately and binary mixtures of these compounds, compared to untreated cells. The observed changes in cell morphology seem to be more pronounced when the cells are exposed to the binary mixtures of DBP and MePB than when exposed to these substances individually, which may confirm the synergy of cytotoxic activity between them (this phenomenon was observed for the higher of the tested concentrations in all tested proportions). It is important to consider such effects when considering the effects of cumulative exposure in the risk assessment in order not to underestimate the risk of adverse effects associated with exposure to chemical mixtures.