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Effect-directed analysis of endocrine-disrupting compounds in multi-contaminated sediment: identification of novel ligands of estrogen and pregnane X receptors

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Abstract

Effect-directed analysis (EDA)-based strategies have been increasingly used in order to identify the causative link between adverse (eco-)toxic effects and chemical contaminants. In this study, we report the development and use of an EDA approach to identify endocrine-disrupting chemicals (EDCs) in a multi-contaminated river sediment. The battery of in vitro reporter cell-based bioassays, measuring estrogenic, (anti)androgenic, dioxin-like, and pregnane X receptor (PXR)-like activities, revealed multi-contamination profiles. To isolate active compounds of a wide polarity range, we established a multi-step fractionation procedure combining: (1) a primary fractionation step using normal phase-based solid-phase extraction (SPE), validated with a mixture of 12 non-polar to polar standard EDCs; (2) a secondary fractionation using reversed-phase-based high-performance liquid chromatography (RP-HPLC) calibrated with 33 standard EDCs; and (3) a purification step using a recombinant estrogen receptor (ER) affinity column. In vitro SPE and HPLC profiles revealed that ER and PXR activities were mainly due to polar to mid-polar compounds, while dioxin-like and anti-androgenic activities were in the less polar fractions. The overall procedure allowed final isolation and identification of new environmental PXR (e.g., di-iso-octylphthalate) and ER (e.g., 2,4-di-tert-butylphenol and 2,6-di-tert-butyl-α-methoxy-p-cresol) ligands by using gas chromatography coupled with mass spectrometry with full-scan mode acquisition in mid-polar fractions. In vitro biological activity of these chemicals was further confirmed using commercial standards, with di-iso-octylphthalate identified for the first time as a potent hPXR environmental agonist.

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Acknowledgments

This study was supported by the French Ministry of Ecology (P190-ECOPI) and by a doctoral fellowship to NC (contract No. CIFRE703/2008). The authors wish to thank Dr. Marina Grimaldi for her help with the purification on the hERα column and two anonymous reviewers for helping to improve the quality of the manuscript.

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Authors and Affiliations

  1. Unité Écotoxicologie In Vitro et In Vivo, INERIS, Parc ALATA, BP2, 60550, Verneuil-en-Halatte, France

    Nicolas Creusot, Saïd Kinani, Jean-Marc Porcher & Selim Aït-Aïssa

  2. Université de Bordeaux, EPOC, UMR 5805, 33405, Talence, France

    Nicolas Creusot & Hélène Budzinski

  3. CNRS, EPOC, UMR 5805, 33405, Talence, France

    Nicolas Creusot & Hélène Budzinski

  4. INSERM U896, IRCM-UM1-CRLC Val d’Aurelle, 34298, Montpellier, France

    Patrick Balaguer

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  1. Nicolas Creusot
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  2. Hélène Budzinski
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Correspondence to Selim Aït-Aïssa.

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Creusot, N., Budzinski, H., Balaguer, P. et al. Effect-directed analysis of endocrine-disrupting compounds in multi-contaminated sediment: identification of novel ligands of estrogen and pregnane X receptors. Anal Bioanal Chem 405, 2553–2566 (2013). https://doi.org/10.1007/s00216-013-6708-5

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  • DOI: https://doi.org/10.1007/s00216-013-6708-5

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