Protein adsorption and transport in agarose and dextran-grafted ion-exchange media.

摘要

Ion-exchange chromatography continues to be dominant in the recovery, separation, and purification of proteins, particularly on the industrial process scale. As a result, there is considerable interest in improving the design and efficiency of ion-exchange adsorbents suitable for preparative and process protein chromatography. Recent advances in this area include the development of novel matrices based on dextran-grafted agarose. These materials exhibit enhanced binding capacities and rapid mass transfer rates crucial to chromatographic protein separation applications. However, a fundamental understanding of protein partitioning and transport in these materials remains elusive. Also, variations in media properties, such as dextran content and charge density, greatly affect protein capacity and transport. Firstly, this dissertation investigates the adsorptive behavior of proteins on these materials by characterizing matrices of varying dextran and charge composition. On one hand, experimental measurements are carried out to determine independent contributions of adsorption equilibrium, morphology and diffusion. On the other, a modeling effort is undertaken in parallel to the experimental measurements to correlate the experimental behavior with dominant mass transfer mechanisms. Secondly, this dissertation presents a novel technique based on the use of radioactive tracers for the measurement of protein adsorption kinetics in adsorbent particles. This method allows the measurements of protein mass transfer rates under gradient and tracer diffusion conditions as well as mass transfer of individual proteins in complex mixtures. Mass transfer measurements are obtained for a number of representative chromatography matrices and at different salt concentrations. The different rates of mass transfer obtained are correlated to structural and chemical differences in the different materials.