title: On-line nano-HPLC/ESI QTOF MS and tandem MS for separation, detection, and structural elucidation of human erythrocytes neutral glycosphingolipid mixture.
abstract: A superior approach involving nano-high-performance liquid chromatography (nano-HPLC) in on-line conjunction to electrospray ionization quadrupole time-of-flight mass spectrometry (ESI QTOF MS) and tandem MS for screening and structural characterization of complex mixtures of neutral glycosphingolipids (GSLs) is here described. Neutral GSLs purified from human erythrocytes were efficiently separated according to the differences in carbohydrate chain length by an optimized nano-HPLC protocol and flow-through detected by ESI QTOF MS at the low femtomole level. Additionally, GSL species were accurately distinguished from the accompanying lipids in the mixture, thus permitting the determination of detailed structural characteristics by data-dependent analysis for identification of GSL constitution within single experiments. An alternative nano-HPLC/ESI QTOF MS approach was designed for dissection of unsaturation/saturation degree of the ceramide moieties defining the hydrophobic portion of GSLs and subsequent localization by nano-HPLC/ESI QTOF MS/MS of the -CH=CH- within the ceramide regions. The method is fast, highly sensitive, and high-throughput amenable and is highlighted as a new and valuable analytical dimension in glycolipidomics.
authors.affiliation: Institute for Medical Physics and Biophysics, University of Münster, Robert Koch Strasse 31, D-48149 Münster, Germany.
authors.forename: Stephan
authors.initials: S
authors.lastname: Kirsch
authors.forename: Mostafa
authors.initials: M
authors.lastname: Zarei
authors.forename: Mario
authors.initials: M
authors.lastname: Cindrić
authors.forename: Johannes
authors.initials: J
authors.lastname: Müthing
authors.forename: Laura
authors.initials: L
authors.lastname: Bindila
authors.forename: Jasna
authors.initials: J
authors.lastname: Peter-Katalinić
mesh_terms: D002240:Carbohydrate Sequence; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D002855:Chromatography, Thin Layer; D004912:Erythrocytes / Q000737:chemistry / Q000378:metabolism*; D006028:Glycosphingolipids / Q000097:blood* / Q000737:chemistry*; D006801:Humans; D006863:Hydrogen-Ion Concentration; D008969:Molecular Sequence Data; D049329:Nanostructures / Q000737:chemistry*; D009862:Online Systems; D021241:Spectrometry, Mass, Electrospray Ionization / Q000379:methods*; D053719:Tandem Mass Spectrometry / Q000379:methods*
chemical_list: D006028:Glycosphingolipids