Chapter 2 - Reversed-phase and hydrophobic interaction chromatography of carbohydrates and glycoconjugates

Abstract

Reversed-phase chromatography (RPC) and hydrophobic interaction chromatography (HIC) share in common nonpolar interactions between solutes and stationary phase ligands. However, the separation conditions are very different regarding the nature of the mobile and stationary phases used in both techniques. In fact, RPC uses highly nonpolar stationary phases consisting mainly of long alkyl chains covalently bonded to the solid support, while the stationary phase used in HIC consists of a highly hydrophilic organic layer to which widely spaced, short alkyl or small aryl functions are attached. On these bases, HIC has been exclusively used in glycoproteins analyses, while RPC has been applied successfully to small and large glycoconjugates separations. To the best of our knowledge, this chapter constitutes the most comprehensive treatment of the field in all its aspects including the basic principles of solute’ retention and separation by RPC and HIC and a variety of applications with 333 references discussed.