Fluorescently Labeled Glycans Analyzed by Ion Mobility-Mass Spectrometry
Marko Grabarics1, Christian Manz2, Alexandra Stuckmann2, Johanna Hofmann1, Weston Struwe3 and Kevin Pagel1,2
1Molekülphysik, FHI der Max-Planck-Gesellschaft, Faradayweg 4-6, 14195, Berlin, GER
2Institut für Chemie und Biochemie, Freie Universität Berlin, Takustr. 3, 14195, Berlin, GER
3Department of Biochemistry, University of Oxford, South Parks Road, OX1 3QU, Oxford, GBR
Complex carbohydrates, also referred to as glycans, are ubiquitous in nature and form one of the major classes of biopolymers. Glycans can adopt highly branched structures with a complex regio- and stereochemistry, resulting in isomers that are often indistinguishable by available analytical approaches.
LC-MS analysis of fluorescently labeled glycans – one of the most widespread methods in analytical glycosciences – often struggles with the aforementioned problem: the in-depth characterization of very similar isomeric species. The hyphenation of ion mobility spectrometry (IMS) with liquid chromatography (LC) and mass spectrometry (MS) offers the possibility to overcome this limitation. To exploit the full potential of the resulting three-dimensional coupling, a comprehensive investigation of the influence of fluorescent labels on the mobility behaviour of glycans is essential. Therefore, we present a systematic study on the effects of various reducing end modifications on the mobility separation of glycans.
For this purpose, the Lewis antigen tri- and tetrasaccharide system was chosen as a representative set of model oligosaccharide structures. Ions of native and derivatized species labeled with procainamide, 2-aminobenzamide and 2-aminobenzoic acid were generated by electrospray ionization, and analyzed in a drift tube IM-MS instrument. Our data show that ion mobility separation is strongly dependent on the specific labels and on the nature of adduct ions. While it is often not feasible to differentiate glycan isomers in their underivatized form, we demonstrate that certain fluorescent labels can lead to significantly enhanced separation of isomeric species.