Practical and theoretical considerations of the effects of ionic liquids on the separation properties of phenyl-based stationary phases in reversed-phase liquid chromatography

Abstract

The use of ionic liquids (ILs) as mobile phase additives in reversed-phase liquid chromatography (RP-LC) can influence the dynamic coating of the stationary phases and even allow them to change their chromatographic properties. In this study, the interaction between thirteen different ILs added to 0.1% HCOOH (mobile phase) and the functional groups on phenyl and pentafluorophenyl stationary phases was investigated via RP-LC analysis and the fluorescence detection (FL) of four basic aromatic analytes (anthracycline antibiotics). In addition, an octadecyl column was included in the experiments to investigate potential differences in how the aromatic and alkyl stationary phases interacted with the tested ILs. Changes in behaviors of the studied analytes in the absence and presence of tested ILs were evaluated based on retention factor (k), peak area (A), number of theoretical plates (NA) and tailing factor (Tf). Furthermore, hierarchical cluster analysis (HCA) and k-means approach were used to better visualize found relationships between the chromatographic parameters of anthracyclines for each column. The obtained results indicated that π-π interactions occurring on the phenyl-based stationary phases resulted in more pronounced effects from the ILs compared to those occurring on the alkyl stationary phase. Different subclusters of the k, A, NA, and Tf parameters detected by the HCA confirmed that the addition of ILs leads to various changes in the chromatographic parameters of the studied analytes. These observations were additionally confirmed by k-means calculations. Overall, our findings show that selecting the appropriate IL for addition to the mobile phase can significantly reduce analysis time and improve peak shape and column performance when using aromatic stationary phases.