Toxicokinetic and toxicodynamic mixture effects of plant protection products: A case study

Authorisation of ready to use plant protection products (PPPs) usually relies on the testing of acute and local toxicity only. This is in stark contrast to the situation for active substances where the mandatory data set comprises a most comprehensive set of studies. While the combination of certain active ingredients and co-formulants may nevertheless result in increased toxicity of the final product such combinations have never been evaluated systematically for complex and long-term toxicological endpoints. We therefore investigated the effect of three frequently used co-formulants on the toxicokinetic and toxicodynamic of the representative active substance combination of tebuconazol (Teb) and prothioconazol (Pro) or of cypermethrin (Cpm) and piperonyl butoxide (Pip), respectively. With all four active substances being potential liver steatogens, cytotoxicity and triglyceride accumulation in HepaRG were used as primary endpoints. Concomitantly transcriptomics and biochemical studies were applied to interrogate for effects on gene expression or inhibition of CYP3A4 as key enzyme for functionalization. Some of the tested combinations clearly showed more than additive effects, partly due to CYP3A4 enzyme inhibition. Other effects comprised the modulation of the expression and activity of steatosis-related nuclear key receptors. Altogether, the findings highlight the need for a more systematic consideration of toxicodynamic and toxicokinetic mixture effects during assessment of PPPs.