Abstract:

Deep eutectic solvents (DESs) were used as a new kind of mobile phase modifier in hydrophilic interaction liquid chromatography (HILIC). In our experiment, a SiO₂ column (150 mm×4.6 mm, 3 μm) was selected to separate several nucleobases and nucleosides by using the mixed solution of acetonitrile and DES (choline chloride-ethylene glycol (1:3, mol/mol)) as mobile phase. Subsequently, the concentrations of DESs in acetonitrile and the column temperature on the effect of separation were investigated. According to the experimental results, better separation of nucleobases and nucleosides was obtained by using acetonitrile and DESs mixed solution as mobile phase than that using traditional water-based solution. For example, a baseline separation between cytosine and cytidine cannot be achieved by HILIC with water-based mobile phase, however, greater improvement was gained by HILIC with modified DES-acetonitrile mobile phase. Meanwhile, the retention times of nucleobases and nucleosides decreased as the proportion of DESs in acetonitrile increased, the most significant decrease of which was with cytidine. Similar retention behavior took place with the effect of column temperature. Decreased retention times of the analytes were observed as column temperature increased. The experimental results indicated that this new method may solve some separation difficulties in traditional water-based HILIC, which also successfully verify the feasibility of DESs as mobile phase modifiers.

Key words: deep eutectic solvents (DESs), hydrophilic interaction liquid chromatography (HILIC), mobile phase, modifier, nucleobases, nucleosides