Determination of competitive adsorption isotherms for modeling large-scale separations in liquid chromatography

Abstract

Preparative and process-scale high-performance liquid chromatography has assumed an increasingly important role in the production of highly purified substances, such as proteins expressed by recombinant DNA technology for use as human pharmaceuticals. The theory for modeling chromatographic separations is well-developed, but requires data on the competitive adsorption behavior of all mixture components for accurate predictions and process design. This paper describes two new methods for determining competitive adsorption isotherms from multi-component frontal chromatography experiments. The first new method reported here complements a method described previously that employed the theory of chromatography to estimate Langmuir isotherm parameters from the breakthrough volumes in frontal chromatography. The new method estimates Langmuir parameters from the experimentally determined compositions of the breakthrough zones, rather than from retention volumes, and so provides a check on the magnitudes of these parameters, but also yields values that may more accurately predict the concentrations of zones in a chromatogram than the Langmuir parameters estimated by other methods. The second method is based on both the mass balance and composition velocity approaches to analysis of the profile obtained in frontal chromatography. The method does not require measurement of concentrations in the profile, instead, concentrations are estimated according to the theory of chromatography from the breakthrough volumes of zones of different composition. The resulting method enables calculation of data points for mobile and stationary phase concentrations at equilibrium, yet it eliminates the need for a tandem rapid analytical high-performance liquid chromatographic unit to monitor effluent compositions.