Separation of carbohydrates using hydrophilic interaction liquid chromatography

• Carbohydrate separation using HILIC was systematically evaluated.
• Column temperature, selectivity and surface charge property played critical roles.
• Complementary selectivity among different columns was beneficial to separation.
• Buffer salt with proper pH and concentration was necessary to ionic carbohydrates.
• A guidance for analysts during method development for carbohydrates’ separation.

A strategy was developed to rapidly evaluate chromatographic properties of hydrophilic interaction chromatography (HILIC) columns for separating carbohydrates. Seven HILIC columns (Silica, Diol, TSK Amide-80, XAmide, Click Maltose, Click β-CD, and Click TE-Cys columns) were evaluated by using three monosaccharide and seven disaccharides as probes. The influence of column temperature on the peak shape and tautomerization of carbohydrates, as well as column selectivity were investigated. The influence of surface charge property on the retention was also studied by using glucose, glucuronic acid, and glucosamine, which indicated that buffer salt concentration and pH value in mobile phase was necessary to control the ionic interactions between ionic carbohydrates and HILIC columns. According to evaluation results, the XAmide column was selected as an example to establish experimental schemes for separation of complex mixtures of oligosaccharide.

A strategy was developed to rapidly evaluate chromatographic properties of hydrophilic interaction chromatography (HILIC) columns for separation of carbohydrates by using three monosaccharide and seven disaccharides as probes.Figure optionsDownload as PowerPoint slide