(438c) Process Development of a Continuous Precipitation and Filtration Unit Operation for the Capture of Biotherapeutics

The biopharmaceutical industry is currently facing several challenges in the downstream purification of recombinant proteins of diverse and significant therapeutic values. The advancement in genetic engineering has enabled a ten-fold increase of target protein titers in bacteria or mammalian cell cultures. However, this improvement in productivity has brought a bottleneck to the subsequent purification steps. This bottleneck refers to the decline in performance of the first crude purification step that relies heavily on chromatography, where the desired proteins are selectively separated by passing through a packed column. One appealing alternative option is target protein precipitation by the addition of reagents. It is an efficient and selective bulk purification technique that takes advantage of the higher titer because it enhances supersaturation which is the driving force for precipitation. Therefore, we propose a continuous precipitation-filtration coupled unit operation to serve as the capture step in the protein purification train. We have identified that the combination of polyethylene glycol (PEG) and ZnCl₂ is efficient and effective in inducing target protein precipitation. Equilibrium and kinetic information on this synergistic precipitation was obtained with human immunoglobulin (hIgG) and bovine serum albumin (BSA) in a batch setting. In addition, study of the critical flux during tangential flow microfiltration of BSA showed that the critical flux increases with increasing ZnCl₂ concentration but reaches a maximum at an intermediate PEG concentration. In this work, we couple a tubular reactor with static mixer for precipitation and tangential flow filtration for the subsequent solid-liquid separation to construct a continuous unit operation for the capture step in downstream processing. Using this laboratory-scale system, we test and demonstrate its applicability to industrially relevant clarified cell culture fluid (CCCF) and show process optimization based on fundamental solubility, kinetics and precipitate particle filtration behavior to fulfill purity and yield requirements for the target therapeutic.