Radial flow chromatography with applications to bioseparation

Radial flow chromatography (RFC) has been found to be efficient for quick and simple purification of biomolecules such as plasma proteins and monoclonal antibodies. In this work, the principle and application of radial flow chromatography in bioseparation has been studied both theoretically and experimentally. On the theoretical aspect, both impulse (zonal) analysis and frontal analysis have been considered. Mathematical models of RFC were derived and solutions were obtained both analytically and numerically. A unified approach for moment expression in chromatography via series expansion has been developed. Based on this approach, the impulse responses in both radial-flow and axial-flow chromatographic systems can be obtained by a simple method. For the purpose of scale-up of RFC, design equations have been developed. By extension of the concept of plate theory, the novel equations for scale-up of the chromatographic system have been obtained on the basis of bed volume. The resulting equations of the Volume Equivalent Theoretical Stage (VETS) are governed by the effects of the system factors that contribute to dispersion and mass transfer resistances. Experiments have been designed and tested for verification of these design equations. A systematic study of the nonlinear effects on the determination of the binding constant in a radial-flow affinity chromatographic system has been undertaken. The criteria for the applicable range of linear theory have been derived for both zonal and frontal analyses. Equations of elution volume derived in this study provide an alternative way for determining the binding constant in the RFC from frontal elution experiments. Applications of RFC for trypsin purification were studied experimentally. A para-aminobenzamidine Zetaffinity cartridge was used and breakthrough experiments were conducted. This radial-flow affinity chromatographic system had proved to be efficient for fast purification. The parameters characterizing the performance of the purification system were evaluated from the breakthrough curves fitting the proposed model. These breakthrough curves from frontal elution experiments also provide valuable information for the design of operating conditions