Investigating the role of biotransformation in the toxicokinetics of ionizable organic compounds, using LC-HRMS: Zebrafish (Danio rerio) embryos exposed to ibuprofen as a case study.

Pharmaceuticals and their metabolites are increasingly detected in aquatic ecosystems, raising concerns about their environmental impact. Most of them, including ibuprofen (IBU), are ionizable organic compounds (IOCs). Their chemical speciation depends on the pH of the surrounding environment, with neutral species having greater permeability and potentially higher toxicity. Τhe pH-dependent toxicity of IBU has been observed, with the lowest LC50 values at pH 5. Also, the effect of pH on the uptake and bioaccumulation of IBU in zebrafish (Danio rerio) embryos (ZFE) was investigated, highlighting the importance of considering pH in toxicokinetic studies. Slight variations in internal concentrations of IBU (Cint) were detected among samples at different pH values, whereas bioconcentration factors (BCFs) were higher under acidic conditions. Biotransformation of IBU was investigated through suspect and non-target screening workflows, utilizing liquid chromatography quadrupole-time-of-flight mass spectrometry (LC-QTOF-MS). This is the first comprehensive study investigating the biotransformation of IBU in ZFE, and hydroxylation and taurine-conjugation were identified as primary pathways. Biotransformation affects the Cint of IBU since variations in Cint among the samples were further reduced when the concentrations of biotransformation products (bio-TPs) were considered (Cint-total). Future studies could greatly benefit from considering the biotransformation of IOCs for a better interpretation of toxicity and bioaccumulation in aquatic organisms.