title: Direct coupling of a flow-flow electromembrane extraction probe to LC-MS.
abstract: A fully integrated and automated electromembrane extraction LC-MS (EME-LC-MS) system has been developed and characterized. Hyphenation of a flow-flow EME probe to LC-MS was accomplished by using an in-built 10-port switching valve of the LC-MS system. The 10-port switching valve decoupled the high pressure of the UHPLC-system from the low pressure required for operation of the EME-probe by automated switching between a sample extraction/analysis and a sample load position. In the sample load position the extracted analytes were loaded into a HPLC sample loop. By switching the valve to the sample extraction/analysis position the setup allowed simultaneous analysis of previously loaded analytes while extracting a new sample. Performance of the system was characterized with respect to precision and linearity (RSD < 2.5%, R(2): 0.998) and the setup was applied for studying the in-vitro metabolism of methadone by rat liver microsomes. As the metabolic reaction proceeded, methadone and its metabolites were extracted and analyzed in parallel by LC-MS using either isocratic or gradient elution. Compared to a conventional in-vitro metabolism analysis based on protein precipitation followed by LC-MS analysis the fully automated EME-LC-MS system offers a significant time saving and in addition demonstrates increased sensitivity as the analytes were automatically enriched during the extraction process. The experiment revealed 6 to 16 times higher S/N ratios of the EME-LC-MS method compared to protein precipitation followed by LC-MS and thus concomitantly lower LOD and LOQ. The setup integrates a complete analytical workflow of rapid extraction, enrichment, separation and detection of analytes in a fully automated manner. These attributes make the developed system a powerful alternative approach for a wide range of analytical applications.
authors.affiliation: Department of Pharmacy, Faculty of Health and Medical Sciences, University of Copenhagen, Universitetsparken 2, 2100 Copenhagen, Denmark.
authors.forename: David
authors.initials: D
authors.lastname: Fuchs
authors.affiliation: Department of Pharmacy, Faculty of Health and Medical Sciences, University of Copenhagen, Universitetsparken 2, 2100 Copenhagen, Denmark.
authors.forename: Charlotte
authors.initials: C
authors.lastname: Gabel-Jensen
authors.affiliation: Department of Pharmacy, Faculty of Health and Medical Sciences, University of Copenhagen, Universitetsparken 2, 2100 Copenhagen, Denmark.
authors.forename: Henrik
authors.initials: H
authors.lastname: Jensen
authors.affiliation: Department of Pharmacy, Faculty of Health and Medical Sciences, University of Copenhagen, Universitetsparken 2, 2100 Copenhagen, Denmark.
authors.forename: Kasper D
authors.initials: KD
authors.lastname: Rand
authors.affiliation: Department of Pharmacy, Faculty of Health and Medical Sciences, University of Copenhagen, Universitetsparken 2, 2100 Copenhagen, Denmark; School of Pharmacy, University of Oslo, P.O. Box 1068, Blindern, 0316 Oslo, Norway.
authors.forename: Stig
authors.initials: S
authors.lastname: Pedersen-Bjergaard
authors.affiliation: Department of Pharmacy, Faculty of Health and Medical Sciences, University of Copenhagen, Universitetsparken 2, 2100 Copenhagen, Denmark.
authors.forename: Steen Honoré
authors.initials: SH
authors.lastname: Hansen
authors.affiliation: Department of Pharmacy, Faculty of Health and Medical Sciences, University of Copenhagen, Universitetsparken 2, 2100 Copenhagen, Denmark. Electronic address: nickolaj.petersen@sund.ku.dk.
authors.forename: Nickolaj Jacob
authors.initials: NJ
authors.lastname: Petersen
mesh_terms: D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D013058:Mass Spectrometry / Q000379:methods*; D008567:Membranes, Artificial
chemical_list: D008567:Membranes, Artificial
keywords: EME-LC-MS; Electromembrane exraction (EME); In-vitro drug metabolism; Methadone metabolism; On-line/in-line sample preparation; Supported liquid membrane (SLM)