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Ph.D. Dissertation Defense Announcement - Nuno Dos Santos Pinto - Dec. 8th

Date: Sunday, Dec. 8th, 2013

Time: 10:00 am
Location: TRC 206

Light refreshments will be served at 9:45 am

Dissertation title: Novel hybrid chromatofocusing methods for protein purification


The efforts made to-date to alleviate the downstream challenges faced by the biopharmaceutical industry have been mainly focused on developing novel chromatographic column packings with either higher ligand densities to accommodate larger production capacity requirements or novel ligand groups that exhibit more than one interaction mode to increase selectivity. However, despite this earlier work, there are still unexploited interactions associated with the functional groups present on the column packings that, if optimized, may lead to novel chromatographic separation techniques. Thus, the rational behind the studies investigated here is to provide innovative separation methods based on hybrid chromatofocusing techniques and employ these unexploited interactions which may be useful in protein purification process development for the biopharmaceutical industry.

A comprehensive optimization method capable of exploiting the synergetic effects both the pH and ionic strength on ion-exchange column packing has not yet been developed. Consequently, one primary research objective of this study is to establish the usefulness of employing combined pH and ionic strength gradients to obtain elements of “orthogonal” two-dimension chromatography in one ion-exchange column that is suitable for the preparative purification of proteins in both dilute and non-dilute regimes. Another main objective is to take the concepts developed in this study for ion-exchange chromatography and apply them in affinity and mixed-mode chromatography, where the ligands on the column packings may exhibit electrostatic interactions as well as hydrophobic, hydrogen bonding and/or affinity interactions, and where pH gradients play a major role in the protein adsorption/desorption process. Lastly, this work aims to increase the understanding of the technique of chromatofocusing based on the use of modern theoretical and experimental tools, and to use this understanding to develop novel hybrid chromatofocusing methods. For this purpose, the development of a computer-aided optimization methodology was also performed which allows efficient chromatographic system identification and optimized design. In this way, the computer simulation methods described here go well beyond any previous attempts at simulations in this area.