A Dual Column pH Switchable Water Stationary Phase System for Separation Control in Supercritical Fluid Chromatography

A dual column system comprised of a pH switchable water stationary phase column and a conventional non-polar capillary column is introduced for use in Supercritical Fluid Chromatography (SFC). By removing or adding NH₄OH to the system hydration source, the water stationary phase pH can be rapidly switched between acidic (measured at pH∼3) and basic (measured at pH∼9) in seconds, while the operating character of the conventional column is unchanged. This switch modulates the velocity of ionizable analytes about 20-fold in the system, whereas non-ionizable analytes are not affected. In this way, the retention time of acids and/or bases can be reproducibly altered (<1% RSD; n = 3) in SFC separations. As a result, analyte selectivity and resolution can be readily controlled during analyses. For example, a selectivity reversal (alpha from 0.4 to 1.6) and a resolution increase (from 0 to 13) are demonstrated. Rapid stationary phase pH switching also allows multiple acids, bases, and/or neutral analytes to be determined simultaneously. Applications demonstrate that this method can greatly simplify complex mixture analysis in SFC by helping to separate target analytes from interfering matrix components.