title: Rapid Screening of Acetylcholinesterase Inhibitors by Effect-Directed Analysis Using LC × LC Fractionation, a High Throughput in Vitro Assay, and Parallel Identification by Time of Flight Mass Spectrometry.
abstract: Effect-directed analysis (EDA) is a useful tool to identify bioactive compounds in complex samples. However, identification in EDA is usually challenging, mainly due to limited separation power of the liquid chromatography based fractionation. In this study, comprehensive two-dimensional liquid chromatography (LC × LC) based microfractionation combined with parallel high resolution time of flight (HR-ToF) mass spectrometric detection and a high throughput acetylcholinesterase (AChE) assay was developed. The LC × LC fractionation method was validated using analytical standards and a C18 and pentafluorophenyl (PFP) stationary phase combination was selected for the two-dimensional separation and fractionation in four 96-well plates. The method was successfully applied to identify AChE inhibitors in a wastewater treatment plant (WWTP) effluent. Good orthogonality (>0.9) separation was achieved and three AChE inhibitors (tiapride, amisulpride, and lamotrigine), used as antipsychotic medicines, were identified and confirmed by two-dimensional retention alignment as well as their AChE inhibition activity.
authors.affiliation: Institute for Environmental Studies (IVM), VU University Amsterdam , De Boelelaan 1087, 1081 HV Amsterdam, The Netherlands.
authors.forename: Xiyu
authors.initials: X
authors.lastname: Ouyang
authors.affiliation: Institute for Environmental Studies (IVM), VU University Amsterdam , De Boelelaan 1087, 1081 HV Amsterdam, The Netherlands.
authors.forename: Pim E G
authors.initials: PE
authors.lastname: Leonards
authors.affiliation: Environmental Institute (EI) , Okruzna 784/42, 972 41 Kos, Slovak Republic.|Faculty of Science, Masaryk University , RECETOX, Kamenice 753/5, 625 00 Brno, Czech Republic.
authors.forename: Zuzana
authors.initials: Z
authors.lastname: Tousova
authors.affiliation: Environmental Institute (EI) , Okruzna 784/42, 972 41 Kos, Slovak Republic.
authors.forename: Jaroslav
authors.initials: J
authors.lastname: Slobodnik
authors.affiliation: Institute for Environmental Studies (IVM), VU University Amsterdam , De Boelelaan 1087, 1081 HV Amsterdam, The Netherlands.
authors.forename: Jacob
authors.initials: J
authors.lastname: de Boer
authors.affiliation: Institute for Environmental Studies (IVM), VU University Amsterdam , De Boelelaan 1087, 1081 HV Amsterdam, The Netherlands.
authors.forename: Marja H
authors.initials: MH
authors.lastname: Lamoree
mesh_terms: D000110:Acetylcholinesterase / Q000378:metabolism*; D002800:Cholinesterase Inhibitors / Q000032:analysis* / Q000737:chemistry* / Q000494:pharmacology; D002851:Chromatography, High Pressure Liquid / Q000295:instrumentation; D057166:High-Throughput Screening Assays / Q000295:instrumentation / Q000379:methods*; D006801:Humans; D013058:Mass Spectrometry / Q000295:instrumentation / Q000379:methods*; D013997:Time Factors
chemical_list: D002800:Cholinesterase Inhibitors; D000110:Acetylcholinesterase