The kinetic plot method applied to gradient chromatography: theoretical framework and experimental validation.

The kinetic plot method, originally developed for isocratic separations, was extended to the practically much more relevant case of gradient elution separations. A set of explicit as well as implicit data transformation expressions has been established. These expressions can readily be implemented in any calculation spread-sheet program, and allow to directly turn any experimental data set representing the relation between the separation efficiency and the flow rate measured on a single column into the kinetic performance limit curve of the tested separation medium. Since the kinetic performance limit curve is based on an extrapolation to columns with a different length, it should be realized that the curve is only valid under the assumption that the gradient time and the delay time (if any) are adapted such that the analytes are subjected to the same relative mobile phase history when the column length is changed. Both experimental and numerical data are presented to corroborate the fact that the kinetic performance limit curves that are obtained using the proposed expressions are indeed independent of the column length the experimental data were collected in. Deviations might arise if excessive viscous heating occurs in columns with a pronounced non-adiabatic thermal behaviour.