Mass Spectrometry Resource, Department of Biochemistry, Boston University School of Medicine, 670 Albany Street, Boston, Massachusetts 02118, USA.
Anal Chem. 2010 Jan 15;82(2):516-22. doi: 10.1021/ac901706f.
Heparan sulfate (HS) and heparin are linear, heterogeneous carbohydrates of the glycosaminoglycan (GAG) family that are modified by N-acetylation, N-sulfation, O-sulfation, and uronic acid epimerization. HS interacts with growth factors in the extracellular matrix, thereby modulating signaling pathways that govern cell growth, development, differentiation, proliferation, and adhesion. High-performance liquid chromatography (HPLC)-chip-based hydrophilic interaction liquid chromatography/mass spectrometry has emerged as a method for analyzing the domain structure of GAGs. However, analysis of highly sulfated GAG structures decasaccharide or larger in size has been limited by spray instability in the negative-ion mode. This report demonstrates that addition of postcolumn makeup flow to the amide-HPLC-chip configuration permits robust and reproducible analysis of extended GAG domains (up to degree of polymerization 18) from HS and heparin. This platform provides quantitative information regarding the oligosaccharide profile, degree of sulfation, and nonreducing chain termini. It is expected that this technology will enable quantitative, comparative glycomics profiling of extended GAG oligosaccharide domains of functional interest.
硫酸乙酰肝素 (HS) 和肝素是糖胺聚糖 (GAG) 家族的线性、不均一的碳水化合物，通过 N-乙酰化、N-硫酸化、O-硫酸化和糖醛酸差向异构化进行修饰。HS 与细胞外基质中的生长因子相互作用，从而调节控制细胞生长、发育、分化、增殖和黏附的信号通路。基于高效液相色谱 (HPLC)-芯片的亲水性相互作用液相色谱/质谱法已成为分析 GAG 结构域的一种方法。然而，由于在负离子模式下喷雾不稳定，对高度硫酸化 GAG 结构（十糖或更大）的分析受到限制。本报告证明，在酰胺-HPLC-芯片构型中添加柱后补流可以稳健、可重复地分析 HS 和肝素中的延伸 GAG 结构域（聚合度高达 18）。该平台提供有关寡糖谱、硫酸化程度和非还原链末端的定量信息。预计这项技术将能够对功能相关的延伸 GAG 寡糖结构域进行定量、比较糖组学分析。
