Determination of Isomeric Glycan Structures by Permethylation and Liquid Chromatography–Mass Spectrometry (LC-MS)

Byeong Gwan Cho, Alireza Banazadeh, Wenjing Peng, Jingfu Zhao, Mona Goli, Sakshi Gautam, Ahmed Hussein, Yehia Mechref

Research output: Chapter in Book/Report/Conference proceedingChapterpeer-review

Abstract

The existence of glycans in isomeric forms is responsible for the multifariousness of their properties and biological functions. Their altered expression has been associated with various diseases and cancers. Analysis of native glycans is not very sensitive due to the low ionization efficiency of glycans. These facts necessitate their comprehensive structural studies and establishes a high demand for sensitive and reliable techniques. In this chapter, we discuss the strategies for effective separation and identification of permethylated isomeric glycans. The sample preparation for permethylated glycans derived from model glycoproteins and complex biological samples, analyzed using LC-MS/MS, is delineated. We introduce protein extraction and release of glycans, followed by strategies to purify the released glycans, which are reduced and permethylated to improve ionization efficiency and stabilize sialic acid residues. High-temperature LC-based separation on PGC (porous graphitized carbon) column is conducive to isomeric separation of glycans and allows their sensitive identification and quantification using MS/MS.

Original languageEnglish
Title of host publicationMethods in Molecular Biology
PublisherHumana Press Inc.
Pages281-301
Number of pages21
DOIs
StatePublished - 2021

Publication series

NameMethods in Molecular Biology
Volume2271
ISSN (Print)1064-3745
ISSN (Electronic)1940-6029

Keywords

  • Glycomics
  • Isomeric separation
  • LC-MS
  • Permethylation
  • Porous graphitized carbon

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