title: A micellar electrokinetic chromatography-mass spectrometry approach using in-capillary diastereomeric derivatization for fully automatized chiral analysis of amino acids.
abstract: In the context of bioanalytical method development, process automatization is nowadays a necessity in order to save time, improve method reliability and reduce costs. For the first time, a fully automatized micellar electrokinetic chromatography-mass spectrometry (MEKC-MS) method with in-capillary derivatization was developed for the chiral analysis of d- and l-amino acids using (-)-1-(9-fluorenyl) ethyl chloroformate (FLEC) as labeling reagent. The derivatization procedure was optimized using an experimental design approach leading to the following conditions: sample and FLEC plugs in a 2:1 ratio (15s, 30mbar: 7.5s, 30mbar) followed by 15min of mixing using a voltage of 0.1kV. The formed diastereomers were then separated using a background electrolyte (BGE) consisting of 150mM ammonium perfluorooctanoate (APFO) (pH=9.5) and detected by mass spectrometry (MS). Complete chiral resolution was obtained for 8 amino acids, while partial separation was achieved for 6 other amino acid pairs. The method showed good reproducibility and linearity in the low micromolar concentration range. The applicability of the method to biological samples was tested by analyzing artificial cerebrospinal fluid (aCSF) samples.
authors.affiliation: Laboratory for the Analysis of Medicines, Department of Pharmacy, Faculty of Medicine, CIRM, University of Liège, Avenue Hippocrate 15, B36-+3-T4, 4000 Liège, Belgium; Department of Analytical Chemistry and Instrumental Analysis, Faculty of Pharmacy, "Iuliu Hațieganu" University of Medicine and Pharmacy Cluj-Napoca, 4 Louis Pasteur Street, 400349 Cluj-Napoca, Romania.
authors.forename: Radu-Cristian
authors.initials: RC
authors.lastname: Moldovan
authors.affiliation: Department of Analytical Chemistry and Instrumental Analysis, Faculty of Pharmacy, "Iuliu Hațieganu" University of Medicine and Pharmacy Cluj-Napoca, 4 Louis Pasteur Street, 400349 Cluj-Napoca, Romania.
authors.forename: Ede
authors.initials: E
authors.lastname: Bodoki
authors.affiliation: Department of Analytical Chemistry and Instrumental Analysis, Faculty of Pharmacy, "Iuliu Hațieganu" University of Medicine and Pharmacy Cluj-Napoca, 4 Louis Pasteur Street, 400349 Cluj-Napoca, Romania.
authors.forename: Timea
authors.initials: T
authors.lastname: Kacsó
authors.affiliation: Laboratory for the Analysis of Medicines, Department of Pharmacy, Faculty of Medicine, CIRM, University of Liège, Avenue Hippocrate 15, B36-+3-T4, 4000 Liège, Belgium.
authors.forename: Anne-Catherine
authors.initials: AC
authors.lastname: Servais
authors.affiliation: Laboratory for the Analysis of Medicines, Department of Pharmacy, Faculty of Medicine, CIRM, University of Liège, Avenue Hippocrate 15, B36-+3-T4, 4000 Liège, Belgium.
authors.forename: Jacques
authors.initials: J
authors.lastname: Crommen
authors.affiliation: Department of Analytical Chemistry and Instrumental Analysis, Faculty of Pharmacy, "Iuliu Hațieganu" University of Medicine and Pharmacy Cluj-Napoca, 4 Louis Pasteur Street, 400349 Cluj-Napoca, Romania.
authors.forename: Radu
authors.initials: R
authors.lastname: Oprean
authors.affiliation: Laboratory for the Analysis of Medicines, Department of Pharmacy, Faculty of Medicine, CIRM, University of Liège, Avenue Hippocrate 15, B36-+3-T4, 4000 Liège, Belgium. Electronic address: marianne.fillet@ulg.ac.be.
authors.forename: Marianne
authors.initials: M
authors.lastname: Fillet
mesh_terms: D000596:Amino Acids / Q000134:cerebrospinal fluid / Q000737:chemistry* / Q000302:isolation & purification; D002210:Caprylates / Q000737:chemistry; D020374:Chromatography, Micellar Electrokinetic Capillary / Q000379:methods; D005449:Fluorenes / Q000737:chemistry; D005466:Fluorocarbons / Q000737:chemistry; D013058:Mass Spectrometry / Q000379:methods; D015203:Reproducibility of Results; D013237:Stereoisomerism
chemical_list: D000596:Amino Acids; D002210:Caprylates; D005449:Fluorenes; D005466:Fluorocarbons; C052812:1-(9-fluorenyl)ethyl chloroformate; C023036:perfluorooctanoic acid
keywords: Amino acids; Chiral separation; FLEC; In-capillary derivatization