Abstract
A procedure was developed for rapid screening of xenobiotic biotransformation products (bioTPs) in single zebrafish (ZF; Danio rerio) embryos. Exposure was carried out from 0–120 hours post fertilization (hpf) to 6 different concentrations of the model compound propranolol (PPL). Following in-plate extraction and non-target instrumental analysis by high resolution mass spectrometry, suspected bioTPs were identified using custom data filtration scripts and matching to in silico structural predictions. A total of eight PPL bioTPs were identified (five at a level 1 confidence and one at a level 2–3 confidence). These findings supplement previously generated toxicometabolomic models derived from the same dataset, and were obtained without conducting additional exposure experiments. In addition to facilitating assessments of inter-individual variability in bioTP production in ZF embryos, we demonstrate that bioTPs can be elucidated using extremely small quantities of biomass (i.e. ∼200 μg). To the best of our knowledge, this is the first time bioTP elucidation has been carried out in single ZF embryos.
Highlights
Determination of pharmaceutical biotransformation is of great importance from both a pharmacotoxicological and environmental toxicological perspective
In pharmacotoxicology it is essential to establish that a drug does not generate toxic biotransformation products within patients, to which end the European Medicinal Agency (EMA) and the US Food and Drug Administration (FDA) have both issued regulatory documents underlining the importance of detection and quanti cation of drug bioTPs during development of pharmaceuticals.[1,2]
Two analyses were performed per sample, utilizing either a BEH amide hydrophilic interaction liquid chromatography (HILIC) column (Waters, USA) with the MS operated in positive electrospray ionization (ESI) mode, or a T3 Atlantis C18 column (Waters) with the MS operated in negative Electronic supplementary information (ESI) mode.[36,37]
Summary
Determination of pharmaceutical biotransformation is of great importance from both a pharmacotoxicological and environmental toxicological perspective. In this study we developed a rapid screening procedure for xenobiotic biotransformation in single ZF embryos using existing non-target data from a toxicometabolomic study involving PPL.[36] By utilizing in silico bioTP structural predictions together with in-house R-scripts, we ltered out PPL bioTPs and con rmed their structures by matching to MS2 data (both in silico predictions and authentic standards). To the best of our knowledge, this is the rst time bioTP elucidation has been carried out in single ZF embryos and highlights the potential for re-purposing non-target metabolomics data for bioTP identi cation and for extracting information on bioTPs from extremely small sample sizes. Two analyses were performed per sample, utilizing either a BEH amide hydrophilic interaction liquid chromatography (HILIC) column (Waters, USA) with the MS operated in positive electrospray ionization (ESI) mode, or a T3 Atlantis C18 column (Waters) with the MS operated in negative ESI mode.[36,37]
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
