A Retention-Matching Strategy for Method Transfer in Supercritical Fluid Chromatography: Introducing the Isomolar Plot Approach

Abstract

A strategy to match any retention shifts due to increased or decreased pressure drop during supercritical fluid chromatography (SFC) method transfer is presented. The strategy relies on adjusting the co-solvent molarity without the need to adjust the back-pressure regulator. Exact matching can be obtained with minimal changes in separation selectivity. To accomplish this, we introduce the isomolar plot approach, which shows the variation in molar co-solvent concentration depending on the mass fraction of co-solvent, pressure, and temperature, here exemplified by CO 2 –methanol. This plot allowed us to unify the effects of the co-solvent mass fraction and density on retention in SFC. The approach, which was verified on 12 known empirical retention models for each enantiomer of six basic pharmaceuticals, allowed us to numerically calculate the apparent retention factor for any column pressure drop. The strategy can be implemented either using a mechanistic approach if retention models are known or empirically by iteratively adjusting the co-solvent mass fraction. As a rule of thumb for the empirical approach, we found that the relative mass fraction adjustment needed is proportional to the relative change in the retention factor caused by a change in the pressure drop. Different proportionality constants were required to match retention in the case of increasing or decreasing pressure drops.

Exploring the application limits of different hold-up time markers in supercritical fluid chromatography

“…For this purpose, a series of measurements were carried out with mobile phases containing 5, 10, 15 and 20% of methanol as organic modifier. The literature provides accurate methods aimed to determine the exact mass or volume fraction of the organic modifier in the mobile phase, as shown by the group of Fornstedt [ 5 ]. This time, the exact composition had no important role in, e.g., numerical calculations, therefore, only the nominal composition provided by the instrument pumps was considered for the sake of observation and comparison.…”

“…However, it can be aided by the use of isopycnic lines in order to better understand the behavior of the physical properties of carbon dioxide as shown by Tarafder et al [ 1 – 4 ]. Method transfer and scalability introduce additional challenges to SFC practitioners; however, more recent solutions provide new tools to overcome the obstacles [ 5 ].…”

Effect of the injection of water-containing diluents on band broadening in analytical supercritical fluid chromatography

“…A non-linear density increase and a pressure gradient along the run at fixed BPR pressure (here 140 bars) results in a dependency of retention behavior not only to gradient conditions (initial composition, slope) but also to the operating conditions [17,18]. For any rigorous method transfer of a linear gradient, solute retention should be reported via composition at elution expressed as co-solvent molarity [19,20]. Nonetheless, for easiness of instrumental setup, the apparent composition at elution (Eq.…”

“…Models expressed as volumetric fractions are easier to use but they are valid only for given operating conditions, here 40 °C, 140 bars at BPR. Any transfer of models would require translation of volumetric fraction into mass fraction of co-solvent [20]. The retention data were fitting a HILIC mixedmode model for the three columns (Figure S3).…”