Journal
JOURNAL OF CHROMATOGRAPHY A
Volume 1610, Issue -, Pages -Publisher
ELSEVIER
DOI: 10.1016/j.chroma.2019.460548
Keywords
Glycosaminoglycan; Heparan sulfate; Capillary liquid chromatography; HILIC-WAX; Salt gradient
Funding
- New National Excellence Program of the Ministry of Human Capacities [UNKP-18-3-II-BME-230]
- National Research Development and Innovation Office [OTKA PD 121187, OTKA 119459]
- Janos Bolyai Research Scholarship of the Hungarian Academy of Sciences
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The analysis of heparan sulfate disaccharides poses a real challenge both from chromatographic and mass spectrometric point of view. This necessitates the constant improvement of their analytical methodology. In the present study, the chromatographic effects of solvent composition, salt concentration, and salt type were systematically investigated in isocratic HILIC-WAX separations of heparan sulfate disaccharides. The combined use of 75% acetonitrile with ammonium formate had overall benefits regarding intensity, detection limits, and peak shape for all salt concentrations investigated. Results obtained with the isocratic measurements suggested the potential use of a salt gradient method in order to maximize separation efficiency. A 3-step gradient from 14 mM to 65 mM ammonium formate concentration proved to be ideal for separation and quantitation. The LOD of the resulting method was 0.8-1.5 fmol for the individual disaccharides and the LOQ was between 2.5-5 fmol. Outstanding linearity could be observed up to 2 pmol. This novel combination provided sufficient sensitivity for disaccharide analysis, which was demonstrated by the analysis of heparan sulfate samples from porcine and bovine origin. (C) 2019 Elsevier B.V. All rights reserved.
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