Date of Award
Doctor of Philosophy in Chemical Engineering - (Ph.D.)
Chemical Engineering, Chemistry and Environmental Science
Kamalesh K. Sirkar
Robert G. Luo
Barbara B. Kebbekus
Biomolecules obtained from fermentation and cell-culture processes are present in low concentrations in a complex medium/broth. For intracellular products, cells are first harvested. Subsequently, cell lysis and homogenization are undertaken to produce a homogenate. The biomolecules are next separated from the cell debris by lysate clarification. For extracellular products, the biomolecules are separated from the whole cells by clarification. The overall yield is reduced with each additional step. As the cost of bioseparation process accounts for 50-80% of the final product cost, innovations in and improvement of bio-downstream processing are greatly needed.
In this research, a cyclic process integrating a commercially available hollow fiber ultrafiltration (UF) cartridge and chromatographic resin beads was developed to reduce the number of devices and steps in bioseparation. This device consisted of ion exchange beads carefully packed on the shell side of a hollow fiber UF module. Loading of proteins on the beads on the shell side was carried out for a period of 10 - 30 minutes from the UF permeate on the shell side produced from a tube-side feed containing protein mixtures in buffer or in a cellular suspension. The eluent was next introduced either from the shellside inlet or tube-side inlet. The chromatographic fractions were collected from the shellside outlet. Continuous cyclic processes were also studied by repeatedly loading, washing and eluting protein(s) without applying cleaning agents to the membrane cartridge at the beginning of each cycle except the first one. A yeast-based cellular suspension containing a binary protein mixture was also applied to this device. The target proteins were recovered and purified successfully by the continuous cyclic process.
A general mathematical model has been developed to predict pressure, flow rate and flux profiles in a bead-filled UF/MF hollow-fiber membrane module. The model was studied for a variety of operational modes in such modules, e.g., UF/MF, permeate flow control, Starling flow, elution, backflushing, etc. By using this model, preliminary mathematical descriptions for protein loading and elution behaviors in the bead-filled device have also been developed.
Dai, Xiaoping, "An integrated membrane-based chromatographic process for biomolecule isolation and purification" (2000). Dissertations. 400.