(472c) Hybrid Shortcut-Modell for Estimating Partition Coefficients in Aqueous Two-Phase Extraction of Therapeutic Proteins

The development of industrial scale bioprocesses has come into high scientific and industrial interest over the last decades. Especially in the red and white biotechnology, process costs are often dominated by 50 - 80 % by the downstream processing. First investigated in the 1896, aqueous two-phase systems (ATPS) were shown to have an enormous potential for the extraction of these biomolecules. In general ATPS show a high biocompatibility and are cheap in comparison to chromatographic methods. The most common ATPS consist of a hydrophilic polymer and a salt serving as phase-forming components. The distribution of the target protein within these systems can either be influenced by the choice of the phase forming components, and additionally by a choice of a displacement salt. The displacement salt influences both the solubility and the distribution of the protein in the two phases. Thus, knowledge of the influence of the different salts potentially useful to enhance the distribution coefficient is necessary.

One promising and fast measure accounting for complex interactions of biological macromolecules is the second osmotic virial coefficient (B₂₂) characterizing the interactions between the biological macromolecules in solution and the cross virial coefficient (B₂₃) characterizing interactions of the biological macromolecules with the salts / impurities. It combines the different influence factors such as pH, Temperature, solvent, salt and salt concentration within one measure.

B₂₂ as well as B₂₃ data was measured using composition gradient multi angle light scattering (GC-MALS) for different proteins and salts in aqueous solution. The results were then used to characterize the distribution behavior of the target protein in the ATPS dependent on the displacement salt. The results point to the fact, that based on B₂₂ and B₂₃ the suitability of a salt as displacer for proteins in aqueous two-phase systems can be investigated. One major advantage using this approach is the fact, that the partition coefficient can be estimated using a hybrid ShortCut Modell based on a minimal set of experimental data. Furthermore the influence of the displacer (salt) on the solubility of the protein can be judged qualitatively serving as an ideal first measure in the development of aqueous two-phase extraction systems. The results will help to improve the development and optimization of ATPS used for efficient separation of processes.