Separation of Monosaccharide Anomers on Photo-Click Cysteine-Based Stationary Phase: The alpha/beta Interconversion Process Studied by Dynamic Hydrophilic Liquid Chromatography

In High-Performance Liquid Chromatography (HPLC), the separation of reducing sugars can typically show three possible typologies of chromatographic profiles (i.e., single peak, two resolved peaks and two peaks interconnected by a plateau) due to the rate at which the relevant alpha/beta anomers interconversion (anomerization) can take place in relation to their elution-time. By analyzing these chromatographic profiles, thermodynamic and kinetic properties of anomerization phenomenon can be extrapolated. In this work we studied the anomerization of some monosaccharides by using a recently developed photo-click cysteine-based stationary phase through dynamic hydrophilic interaction liquid chromatography (D-HILIC) conditions. In the 5-25 degrees C temperature range, the DG(#) (alpha ->beta) and DG(#) (beta ->alpha) barriers were found to achieve values within the interval 21.1/22.2 kcal/mol for glucose, with differences between alpha ->beta and beta ->alpha reactions of about 0.4 kcal/mol. For xylose, in the same temperature range, the DG(#) (alpha ->beta) and DG(#) (beta ->alpha) barriers are between 20.7 to 21.5 kcal/mol, with differences between alpha ->beta and beta ->alpha reactions of about 0.2 kcal/mol. The experimental data are in agreement with those reported in literature, confirming the this new stationary phase using HILIC conditions is a robust platform to measure kinetic and thermodynamic properties of the isomerization reaction.