Estrogenic activity of aromatic compounds present in mineral oil used in printing inks

Mineral oils (MO) used as ingredients for many printing inks are complex mixtures of hydrocarbons. Although migration of MO from paperboard food packaging made of recycled paper into food has been shown, the toxicological relevance of these findings is elusive. Therefore we investigated the estrogenic potential of mineral oil using different in vitro assays. Proliferative effects of MO in the estrogen responsive human breast cancer cell line MCF7 were assessed in the E-screen, and endogenous estrogen responsive gene expression was quantified by real-time PCR. Transactivation assays were performed in HeLa9903 containing the human estrogen receptor α (ERα) and an estrogen responsive element (ERE)-driven luciferase. MO primarily consist of mineral oil saturated hydrocarbons (MOSH) and mineral oil aromatic hydrocarbons (MOAH). These two fractions were separated by preparative HPLC and also assessed in the bioassays. MO in a thousand-fold dilution stimulated MCF7 proliferation but did not induce reporter gene expression in HeLa9903 cells. Furthermore, expression of the estrogen responsive genes PGR, TFF1 and GREB1 in MCF7 cells was induced after 24 h exposure to total MO. The estrogenic effects were derived from the MOAH fraction, as exposure of MCF7 cells to this fraction induced gene expression and increased proliferation. Estrogenic activity of 14 more tested MO correlated with the percentage of the MOAH fraction. In conclusion estrogenic action of aromatic substances derived from MO could be shown in vitro.