PhD Thesis Presentation - Process Intensification of Protein Crystallization through Innovative Process Equipment and Operation

Crystallization is an alternative separation and purification technique to conventional chromatography for proteins due to specific characteristics, such as good scalability, low cost, and the attainable stability of the crystalline products. However, protein crystallization often suffers from long batch time due to the slow crystallization kinetics. Furthermore, protein crystallization is conventionally conducted in the batch mode of operation, which has potential disadvantages compared to continuous operation. These challenges of protein crystallization are addressed in this thesis by employing process intensification principles. The work presented in this thesis will focus on two typical process intensification domains, i.e., the space and time domains involving equipment structurization and continuous operation. 3D printing is used as a tool to achieve process intensification through structurization, because of its ability for rapid fabrication of customized process equipment with complex geometries. Novel crystallizer types and process configurations are investigated with the aid of 3D printing as the fabrication technique.

First, a workflow that includes various steps in the fabrication of crystallizers with 3D printing is presented. The design and fabrication of the crystallizers used in this thesis is also included. The workflow aims to facilitate the adoption of 3D printing as a tool to achieve process intensification of crystallization processes through structurization.

Second, the performance of a pneumatically agitated airlift crystallizer (ALC) is characterized and compared to a mechanically agitated stirred tank crystallizer (STC) for the crystallization of the model protein lysozyme. Process intensification is achieved in this work through structurization by the adoption of an ALC for protein crystallization resulting in higher throughput, along with larger and unagglomerated crystals with high activity.

Third, the 3D printed ALC and STC are characterized for continuous crystallization of lysozyme with a focus on the attainable solid-state forms from a mixed-suspension mixed-product-removal (MSMPR) crystallizer. This work presents the first experimental study on protein crystallization in an MSMPR crystallizer, which reveals a dramatic influence of the mode of operation on the attainable solid-state form of a protein.

Finally, three 3D printed tubular crystallizers, a standard Kenics static mixer (sKTC), a novel gapped Kenics static mixer (gKTC), and a hollow tube (HTC), are designed and characterized numerically and experimentally. This study shows that the novel gKTC outperforms the HTC in terms of mixing length, residence time distribution (RTD), and solid suspension and the sKTC in terms of pressure drop, solids suspension, and lower shear while achieving the same RTD and mixing length, which has important implications for practical applications of continuous protein crystallization. In this thesis, process intensification is achieved through structurization by employing the airlift crystallizer and the static mixer-based tubular crystallizer and continuous operation by MSMPR crystallizer and plug flow crystallizer.