Separation of complex sugar mixtures on a hydrolytically stable bidentate urea-type stationary phase for hydrophilic interaction near ultra high performance liquid chromatography.

A stationary phase bearing both bridged bis-ureido and free amino groups (USP-HILIC-NH2 -2.5SP) for high-speed hydrophilic interaction liquid chromatography separations was prepared using a one-pot two-step procedure starting from 2.5 μm totally porous silica particles. Highly polar compounds, such as polyols, hydroxybenzoic acids, and sugars, were successfully analyzed in shorter times and with higher peak efficiency, when compared to results obtained with a bidentate urea-type column packed with 5 μm particles. Increased sugarophilicity and better peak shape were attested for the USP-HILIC-NH2 -2.5SP column (100 × 3.2 mm id) when compared with two commercially available UHPLC columns, namely an acquity BEH amide packed with totally porous 1.7 μm microparticles and a HILIC Kinetex column packed with core-shell 2.6 μm particles. Finally, the new column was employed in the separation of complex mixture of sugars (mono-, di-, and oligosaccharides) and in the analysis of beer samples. The resulting chromatograms showed good selectivity and overall resolution, while the catalyzing effect of the free amino moieties resulted in excellent peak shapes and in the absence of split peaks due to sugar anomerization phenomena.