Single but Not Combined In Vitro Exposure to Bisphenol A and Nanoplastics Affects the Cholinergic Function of the Ascidian Ciona robusta

Nanoplastics are known to represent a threat to marine ecosystems. Their combination with other contaminants of emerging concerns (CECs) may amplify ecotoxic effects, with unknown impacts on marine biodiversity. This study investigates the effects, single and combined, of bisphenol A (BPA)-one of the most hazardous CECs-and polystyrene nanoparticles (PS NPs)-as a proxy for nanoplastics, being among the most commonly found asmarine debris-on cholinesterase (ChE) activities of the ascidian Ciona robusta. ChE activity was first measured in the siphons, tunic, and viscera of wild-caught adult specimens and exposed in vitro to BPA (0.01, 0.21, 0.69 mM) and PS NPs (0.0096-0.096 mM; 8.096 × 10⁹-10¹⁰ particles, respectively) alone and combined for 15 min of incubation. PS NPs' behavior in milliQ water and in the ChE assay reaction buffer was characterized alone, combined with BPA, and analyzed through ζ-potential measurements via Dynamic Light Scattering. The results revealed that ChE activity was predominant in the viscera and siphons of C. robusta; PS NPs did not affect the ChE activity alone or combined, while BPA caused a concentration-dependent inhibition of ChE activity in the viscera. No changes in ζ-potential were observed for PS NPs alone or combined with BPA in the ChE buffer, suggesting no interaction. Further investigations are needed to understand the potential neurotoxic consequences for C. robusta and ecological risk scenarios due to exposure to BPA and nanoplastics in marine coastal waters.