Hydrophilic interaction liquid chromatography with methanol-water eluent on a zeolite

Hydrophilic interaction chromatography (HILIC) works with organic solvent-water mixtures as eluent and is based on the formation of a water enriched liquid phase on the surface of a hydrophilic stationary phase. Hydrophilic solutes are retained on that stagnant water-rich film depending on the difference of solvation compared to the mobile phase composition. However, the enhancement of selectivity by increasing the fraction of organic cosolvent is coupled with a limitation the analyte solubility, and the improvement of the HILIC principle by new hydrophilic stationary phases is the remaining option.

Y-zeolite (faujasite, FAU type) in the Na⁺-form with an average particle diameter of 5 μm was used as packing material in a 125 mm long HPLC column. The chromatographic response of the column was tested in methanol-water mixtures as eluent after injection of several aliphatic alcohols, polyols and monosaccharides with eluent conditions where no separation occurs on diol functionalized silica. On the zeolite the retention time increases according to ethylene glycol < glycerol < erythritol < sorbitol < inositol. The separation principle is explained to be superposed by two effects: firstly, a partition equilibrium between the water-rich phase in the zeolite micropores exists, and secondly, selective interactions with the inner crystalline pore surface and fixed-position Na⁺ ions, both serving to enhance the selectivity. Furthermore, arabinose and fructose monosaccharides could be separated into their tautomeric forms. Only upon increasing the temperature from 20 to 60 °C the tautomeric pattern merges into a single peak.

Instead of the stagnant water rich surface layer, zeolite micropores now take over that function. As a result, the selectivity among polyols and between α/β-arabinopyranose and β-fructopyranose/β-fructofuranose tautomers is extraordinary superior towards conventional hydrophilic interaction liquid chromatography (HILIC).