Among the modern ionization techniques for the analysis of biomolecules, Electrospray Ionization (ESI) has proven the most effective for the mass spectrometry (MS) analysis of sulfated carbohydrates, called Glycosaminoglycans (GAGs). GAGs are acidic molecules made up of repeating disaccharide units with numerous sulfate groups. GAGs are easily ionized and produce abundant negative ions. Sulfates are the most labile functional groups and are more fragile than peptides and less acidic glycans. Due to sulfate lability, GAGs can be difficult to analyze by ESI mass spectrometry without using finely tuned ESI parameters. Optimization of the ESI tuning parameters will result in little or no in-source fragmentation of sulfated GAGs.
Highly sulfated heparin hexasaccharides with defined structure can be used to optimize the ESI tuning parameters in various mass spectrometers. Extent and position of sulfation can lead to varying degrees of lability for every oligosaccharide. The lability of the sulfate groups increases as the size of the heparin oligosaccharide increases. Successful tuning parameters are more easily achieved using a highly sulfated hexasaccharide standard as compared to a disaccharide standard.