Isomer separation and effect of the polymerization degree on the gas-phase structure of chondroitine sulfate oligosaccharides analyzed by ion mobility-tandem mass spectrometry

International audienceChondroitin sulfate (CS) glycosaminoglycans (GAG) are bioactive sulfated polysaccharides comprising 40-200 sulfated repeating units of D-glucuronic acid (D-GlcA) and 2-acetamido-2-deoxy-D-galactose (D-GalNac) sulfated at various positions. D-GalNac may contain a 6-O-sulfate group and/or a 4-O-sulfate group, and D-GlcA may be sulfated at the 2- or 3-O position, leading thus to a huge diversity of the CS sequence. Being present in the brain, articulations, placenta, and immune system, CS play critical roles in various physiological processes. In this context, development of efficient analytical methods for the investigation of CS sulfation patterns is of high necessity to associate these biological functions with precisely sulfated CS structures.Due to their high negative charge density and heterogeneous sulfation pattern, CS oligosaccharides are fragile and challenging to analyze. In this context, MS characterization of synthetics oligosaccharides of chondroitin sulfated in position 4 (CS-A), 6 (CS-C) or in positions 4 and 6 (CS-E) of the D-GalNac unit was carried out. Isomer separation of di-, tetra- and hexa-CS-A and CS-C was investigated using both MS/MS and IMS-MS. The effects on the gas-phase structure of the polymerization degree and number of sulfate moiety were studied. Diagnostic ion products were identified from di-, tetra and hexa-CS-A and CS-C alkali adduct isomers allowing a non-ambiguous identification of these compounds. By contrast, MS/MS spectra of deprotonated species of CS-A and CS-C did not show specific ion product. Isomers differentiation by IMS-MS was achieved only for hexa-CS-A and CS-C from the [CS−6H+3Cs]3− species. Finally, influence of polymerization degree and sulfate group position on the gas phase conformation was investigated. For the same charge state mono- and di-sulfated species linearly evolved in function of the side chain. Increase in CCS values with polymerization degree is more important for mono-sulfated species than for CS-E when the charge density increases. These data can be integrated in biological phenomenon involving CS