title: Quantification of the Fabry marker lysoGb3 in human plasma by tandem mass spectrometry.
abstract: Morbus Fabry is a hereditary metabolic disorder with low prevalence and late clinical manifestation. A defect in the α-galactosidase gene leads to lysosomal accumulation of the glycolipid globotriaosylceramide (Gb3). Gb3 may be used for monitoring of enzyme replacement therapy (ERT), but diagnostic sensitivity is limited. Recently, globotriaosylsphingosine (lysoGb3) was introduced as a promising new marker with significantly better sensitivity. For Fabry diagnosis, clinical studies and possible therapy monitoring, we established a fast and reliable LC-MS/MS assay for quantification of lysoGb3 in human plasma. Protein precipitation and glycolipid extraction from EDTA plasma was performed using acetone/methanol. Samples were analyzed by UPLC-MS/MS in MRM mode. In contrast to HPLC with fluorescence detection, the LC-MS/MS method requires no derivatization, less sample preparation and less instrument analysis time (<3 min). As internal standard (ISTD), a glycine derivative of lysoGb3 was synthesized, and the product was purified by HPLC. ISTD properties such as polarity (affecting extraction and elution), ionization and fragmentation pathway were almost identical compared to the analyte. The new LC-MS/MS assay for the Fabry marker lysoGb3 shows good performance and allowed for better discrimination between Fabry patients and controls than Gb3.
authors.affiliation: Institute of Clinical Chemistry and Laboratory Medicine, Medical Center of the Johannes Gutenberg University, Mainz, Germany. ralf.krueger@mri.bund.de
authors.forename: Ralf
authors.initials: R
authors.lastname: Krüger
authors.forename: Andreas
authors.initials: A
authors.lastname: Tholey
authors.forename: Thomas
authors.initials: T
authors.lastname: Jakoby
authors.forename: Rita
authors.initials: R
authors.lastname: Vogelsberger
authors.forename: René
authors.initials: R
authors.lastname: Mönnikes
authors.forename: Heidi
authors.initials: H
authors.lastname: Rossmann
authors.forename: Michael
authors.initials: M
authors.lastname: Beck
authors.forename: Karl J
authors.initials: KJ
authors.lastname: Lackner
mesh_terms: D000328:Adult; D015415:Biomarkers / Q000097:blood; D002240:Carbohydrate Sequence; D016022:Case-Control Studies; D005591:Chemical Fractionation; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D000795:Fabry Disease / Q000097:blood*; D005260:Female; D006017:Glycolipids / Q000097:blood*; D006801:Humans; D016014:Linear Models; D008297:Male; D008969:Molecular Sequence Data; D015203:Reproducibility of Results; D013107:Sphingolipids / Q000097:blood*; D053719:Tandem Mass Spectrometry / Q000379:methods*; D014281:Trihexosylceramides / Q000097:blood
chemical_list: D015415:Biomarkers; D006017:Glycolipids; D013107:Sphingolipids; D014281:Trihexosylceramides; C063288:globotriaosyl lysosphingolipid; C018549:globotriaosylceramide