Plastic nanoparticles amplify endocrine activity of paracetamol and ibuprofen mixtures at environmentally relevant concentrations

Abstract

Paracetamol and ibuprofen are among the most widely used pharmaceuticals and are increasingly relevant aquatic co-contaminants. Their hazard assessment is further complicated by co-exposure to plastic nanoparticles (PNPs), which can adsorb chemicals and modify mixture effects. Here, we investigated the endocrine activity of paracetamol (PARA), ibuprofen (IBU), their binary mixtures, and their co-mixtures with polystyrene, polypropylene, and polyethylene nanoparticles (PS-NPs, PP-NPs and PE-NPs, respectively) under in vitro conditions using environmentally relevant, non-cytotoxic concentrations. Estrogen receptor (ER) activation and steroidogenesis responses were evaluated to resolve both receptor-mediated and downstream endocrine effects. Neither PARA nor IBU alone showed ER agonistic activity under the tested conditions. In contrast, low-concentration PARA–IBU mixtures elicited concentration- and ratio-dependent synergistic or antagonistic responses, indicating emergent mixture behaviour not predictable from the individual compounds. Co-exposure to PNPs further enhanced ER agonistic activity. In mixtures containing PE-NPs and PP-NPs, the magnitude of the response was driven predominantly by PNPs concentration, whereas this was governed mainly by drug concentration in PS-NP-containing mixtures. For steroidogenesis assays, IBU shifted hormone production towards upstream intermediates, increasing progesterone and 17-hydroxyprogesterone while decreasing estradiol and testosterone, whereas PARA increased the final steroid products. PS-NPs potentiated drug-specific steroidogenic effects in mixture with both pharmaceuticals. These findings demonstrate that environmentally relevant pharmaceutical mixtures can gain endocrine activity in the presence of PNPs, and that the outcome depends on both mixture composition and nanoplastics identity. Overall, our study highlights the need to incorporate pharmaceutical–nanoplastic co-exposure into environmental hazard assessment and mixture risk evaluation.