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Online coupling of immunoextraction, digestion, and microliquid chromatography-tandem mass spectrometry for the analysis of sarin and soman-butyrylcholinesterase adducts in human plasma
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Edité par CCSD ; Springer Verlag -
International audience. Organophosphorus nerve agent (OPNA) adducts formed with human butyrylcholinesterase (HuBuChE) can be used as biomarker of OPNAs exposure. Indeed, intoxication by OPNAs can be confirmed by the LC/MS 2 analysis of a specific HuBuChE nonapeptide on which OPNAs covalently bind. A fast, selective and highly sensitive on‐line method was developed to detect sarin and soman‐adducts in plasma, including immunoextraction by anti‐HuBuChE antibodies, pepsin digestion on Immobilized Enzyme Reactors (IMER) and microLC‐MS 2 analysis of the OPNA‐adducts. The potential of three different monoclonal antibodies, covalently grafted on sepharose, were compared for the extraction of HuBuChE. The on‐line method developed with the most promising antibodies allowed to extract up to 100% of HuBuChE contained in plasma and to digest 45% of it in less than 40 min. Moreover, OPNA‐HuBuChE adducts, aged OPNA‐adducts and unadducted HuBuChE could be detected (with S/N> 2000), even in plasma spiked with low concentration of OPNA (10 ng.mL ‐1). Finally, the potential of this method was compared to approaches involving other affinity sorbents, already described for HuBuChE extraction.
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