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Optimisation du suivi de l'exposition aux hydrocarbures aromatiques polycycliques pendant la grossesse avec le développement et la validation de méthodes analytiques et biologiques

Abstract : Polycyclic Aromatic Hydrocarbons (PAHs) are mainly emitted by human activities and are one of the main pollutant groups in air, soils, waters, and food. Therefore, they are subject to atmospheric, environmental and food monitoring. This study focuses on the particular case of exposure of pregnant women and their fetus, which are at risks. Indeed, PAH exposure can lead to fetal malformations, prematurity and even disorders in children and adult. To date, only a few PAHs regulated by US-EPA were searched and quantified in maternal or umbilical cord blood. Moreover, analytical performance data are often lacking. Hence, this study aims for the first time in France, at developing analytical and biological methods for the monitoring of PAH exposure during pregnancy. In a first part, the simultaneous determination of the 24 regulated PAHs in sera from maternal and umbilical cord bloods was developed. First, the analysis of the 24 PAHs by liquid chromatography and UV and fluorescence detections (HPLC-UV-FD) and by gas chromatography hyphenated to mass spectrometry (GC-MS) were optimized. The sample pretreatment of the plasma samples was next developed. The precipitation of the proteins present in the plasma was optimized with a Design of Experiment to disrupt all the interactions between them and the PAHs. A solid phase extraction (SPE) protocol, with a C18-based sorbent, was next optimized to extract and concentrate the PAHs. To improve the enrichment factors, the elution SPE fractions were next evaporated and 2 approaches were evaluated: a total or a partial evaporation ("last drop" method). The final analytical protocol involving the sample pretreatment followed by HPLC-UV-FD or GC-MS was successfully validated with spiked pooled sera from maternal and umbilical cord bloods with the accuracy profile approach. Then, sera samples from maternal and umbilical cord bloods were analyzed by LC/UV-FD and GC/MS. Finally, the previously developed analytical methods were used for a study with the ex vivo human cotyledon perfusion model. Perfusions were carried out for the first time with a mix of 14 US-EPA PAHs and their placental transfer was demonstrated. In addition, human trophoblast cell cultures with 1 µM with 14 US-EPA PAHs were carried out to evaluate the impact of PAH exposure on placental functions. The dysfunction of certain placental functions such as trophoblast fusion and differentiation, endocrine functions and detoxification functions was demonstrated.
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Stéphanie Swiha. Optimisation du suivi de l'exposition aux hydrocarbures aromatiques polycycliques pendant la grossesse avec le développement et la validation de méthodes analytiques et biologiques. Gynécologie et obstétrique. Université Sorbonne Paris Cité, 2019. Français. ⟨NNT : 2019USPCB020⟩. ⟨tel-02538673⟩

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