Glycosaminoglycans (GAGs) are linear anionic polysaccharides that play important roles in modulating biological functions in most mammalian organisms. The characterization of GAGs has been the subject of numerous analytical techniques over the years, allowing researchers to gain a better understanding of the functions of GAG molecules. Nonetheless, highly sulfated GAGs pose an analytical challenge because of a high level of heterogeneity inherent from their non-template biosynthesis, and also because of the chemical lability of the sulfation modifications that are the target of these analyses. In this study, mass spectrometry-based techniques, tandem mass spectrometry (MS/MS) and capillary zone electrophoresis-mass spectrometry (CZE-MS), have allowed for GAG analysis in a fast, accurate, and reliable manner. The stereochemistry of the C5 carbon of GAGs was distinguished by different types of MS/MS methods and multivariate statistical analysis. Workflows have been evaluated to examine GAG biomarkers with multivariate statistical analysis. In addition, applications of our methods have been explored, such as the high-throughput analysis of clinical GAG samples (e.g., complex and/or low levels of GAGs), synthetic GAG oligosaccharides, and CRISPR/Cas9-mediated GAG biosynthetic products.
