Ramifications and Insights on the Role of Water in Chiral Sub/Supercritical Fluid Chromatography.

More than 40 cosolvents have been used with carbon dioxide to alter its solvation strength. Among the most interesting systems is the subcritical/supercritical CO2/alkanol eluents. Using small amounts of water in CO2/MeOH is known to be beneficial in chiral subcritical/supercritical chromatography. However, the ramifications of introducing water as a cosolvent component is not entirely understood. In this work, we demonstrate important aspects of the CO2/MeOH/H2O system on nine chiral stationary phases with very different surface chemistries, encompassing derivatized polysaccharides, macrocyclic glycopeptides, iso-butylmercaptoquinine, isopropyl macrocyclic oligosaccharides, and π-electron acceptor/π-electron donor phases. A hydrophilicity scale has been shown to be useful in predicting if a given chiral column chemistry would show a significant enhancement in separation efficiency in the presence of water in the CO2/MeOH system. We demonstrate up to 8-fold enhancements in plate counts of chiral separations with a concomitant decrease in retention times, as predicted by the qualitative test. The same chiral analysis can now be completed in almost a third of the time with the addition of small amounts of water, thereby decreasing organic solvent consumption by a considerable amount. Hydrophobic stationary phases show a minimal increase in efficiency and decrease in analysis times and optimized separations show much larger reduced plate heights, compared to more hydrophilic stationary phases. Furthermore, the presence of water can alter the nature of the adsorption isotherm under nonlinear conditions. Small amounts of water can be used to tune nonlinear tailing peaks into fronting ones, significantly improving preparative enantiomeric separations.