Using computer simulation to assist in the robustness analysis of an ion-exchange chromatography step.

This paper presents a methodology to gain process knowledge and assist in the robustness analysis of an ion-exchange step in a protein purification process using a model-based approach. Factorial experimental design is common practice in industry today to obtain robustness characterization of unit operations with respect to variations in process parameters. This work aims at providing a better insight into what process variations affect quality and to further reduce the experimental work to the regions of process variation that are of most interest. This methodology also greatly increases the ability to predict process performance and promotes process understanding. The model calibration part of the methodology involves three consecutive steps to calibrate a steric mass action (SMA) ion-exchange chromatography model. Firstly, a number of gradient elution experiments are performed. Secondly, experimental breakthrough curves have to be generated for the proteins if the adsorption capacity of the medium for each component is not known. Thirdly, a multi-component loading experiment is performed to calibrate the multi-component effects that cannot be determined from the single-component experiments. The separation process studied in this work is the separation of polyclonal IgG from a mixture containing IgG, myoglobin and BSA. The calibrated model is used to simulate six process variations in a full factorial experiment. The results of the simulations provide information about the importance of the different process variations and the simulations are also used to determine the crucial points for the process parameter variations. The methodology can be used to assist in the robustness analysis normally performed in the pharmaceutical industry today as it is able to predict the impact on process performance resulting from variations in salt concentration, column load, protein concentration and flow rate.

Influence of bioreactor scale and complex medium on probing control of glucose feeding in cultivations of recombinant strains of Escherichia coli.

The objective of this work was to evaluate the performance of a feedback glucose control strategy (the probing strategy) in production relevant bioreactors with complex and mineral media. Experimental results from fed-batch cultivations with two recombinant Escherichia coli constructs expressing two different human therapeutic proteins were used to assess the performance and limitations of the glucose probing technique. Even though the performance of the probing strategy was affected by scale and complex media, this methodology rapidly identified a glucose feed protocol similar to an experimentally derived feed regime. This methodology may serve as a powerful tool for industrial process development and in optimization of glucose feed regimes when transferring process technology from one bioreactor system to another.