Class 2018
PhD Grant: PD/BD/142943/2018
PhD thesis
Strategies to integrate and intensify the manufacturing of biological products
Supervisor
• Ana M. Azevedo, Professor @ IST/ULisboa, PT
Collaborators
• Raquel Aires-Barros, Professor @ IST/ULisboa, PT
• Alois Jungbauer, Professor @ BOKU, Vienna, Austria
• Nico Ling, Professor @ BOKU, Vienna, Austria
PhD Degree complete: Discussion on July 17, 2024
THESIS ABSTRACT
The biotechnology industry has been based on batch processing, which is normally associated with low productivities, but moving to continuous processing enables process intensification and brings strong advantages.
Aqueous two-phase extraction and precipitation are low-cost unit operations that have been widely studied in batchwise mode, but they lack an efficient alternative for continuous processing. An oscillatory flow reactor (OFR) is a tubular reactor that has been used in multiple chemical engineering processes. One of the most important features of this reactor is the uniform mixing that is provided by the combination of periodically spaced restrictions and the oscillatory motion of the fluid, making it the answer for a more cost-competitive process. In this work, the partition of a commercial industrial enzyme – α-amylase – in different polyethylene glycol (PEG)/phosphate systems was studied in batch, and the compositions presenting the highest yield of partition to the top phase were transferred to this reactor, where different conditions for frequency and amplitude of oscillation, and total mass flow were evaluated. It was performed PEG/zinc chloride (ZnCl2) precipitation of antibodies from an artificial mixture with fetal bovine serum, mimicking the supernatant of antibody-producing cells. PEG/ZnCl2 precipitation was first optimized in batch and further implemented in the reactor under continuous operation, where the influence of mixing intensity was investigated.
With the expansion of the biopharmaceuticals market, it is important to improve the processes that lead to high purity products. To obtain secretory immunoglobulin A (sIgA), two affinity chromatography processes, using Capture Select IgA and Capture Select IgA-CH1 resins, were built to verify if it would be possible to develop an efficient purification of this innovative biopharmaceutical from clarified Chinese hamster ovary cells supernatant.
RESUMO DA TESE
A indústria biotecnológica tem baseado a sua produção em processos descontínuos, normalmente associados a baixas produtividades. Neste sentido, uma mudança para um processamento contínuo irá permitir a intensificação de processo, trazendo fortes vantagens.
Extração com duas fases aquosas e precipitação são processos de baixo custo que têm sido estudados em modo descontínuo, carecendo de uma alternativa eficiente para processamento contínuo. O reator de fluxo oscilatório (RFO) é um tipo de reator tubular que tem sido usado em vários processos da industria química. Uma das características mais importantes deste tipo de reator é a mistura uniforme que é proporcionada pela combinação de restrições periodicamente espaçadas e pelo movimento oscilatório do fluido, tornando-o a resposta para um processo com custo mais competitivo. Neste trabalho, a partição de uma enzima industrial comercial – α-amilase – em diferentes sistemas polietilenoglicol (PEG)/fosfato foi estudada em modo descontínuo, e as composições que apresentavam maior rendimento de partição para a fase superior foram transferidas para este reator, onde diferentes condições de frequência e amplitude de oscilação e fluxo de massa total foram avaliadas. Este reactor foi igualmente usado na precipitação de anticorpos com PEG/ZnCl2 a partir de uma mistura artificial com soro fetal bovino, mimetizando o sobrenadante de células produtoras de anticorpos. A precipitação foi primeiro otimizada em descontínuo e posteriormente implementada no reator em operação contínua.
Com a expansão do mercado biofarmacêutico, torna-se necessário melhorar os processos que levam a produtos de alta pureza. Para a obtenção da imunoglobulina A secretora (sIgA), foram construídos dois processos de cromatografia de afinidade, utilizando duas resinas cromatográficas diferentes, com vista ao desenvolvimento de um processo de purificação eficiente para este biofármaco inovador, a partir dum sobrenadante clarificado de células de ovário de hamster Chinês.
PUBLICATIONS
Papers
Ferreira-Faria, D., Domingos-Moreira, F., Aires-Barros, M. R., Ferreira, A., Azevedo, A.M., “Continuous precipitation of antibodies using oscillatory flow reactor: A proof of concept”, Sep. Purif. Technol., 317 (2023) 123924 (doi.org/10.1016/j.seppur.2023.123924)
Ferreira-Faria, D., Aires-Barros, M.R., Azevedo, A.M., Continuous aqueous two-phase extraction: From microfluidics to integrated biomanufacturing, Fluid Phase Equilibria, 508 (2020) 112438 (https://doi.org/10.1016/j.fluid.2019.112438)
Diogo F. Faria, D.F., Silva, T.P., Aires-Barros, M.R., Azevedo, A.M., “A Chronology of the Development of Aqueous Two-Phase Systems as a Viable Liquid-Liquid Extraction for Biological Products”, Reference Module in Chemistry, Molecular Sciences and Chemical Engineering, Elsevier,2019 (https://doi.org/10.1016/B978-0-12-409547-2.14393-8)
Oral Communications
Ferreira-Faria, D., Aires-Barros, M.R., Ferreira, A., Azevedo, A.M., Oscillatory Flow Reactor: a solution for continuous biomanufacturing, ISPPP 2021, Porto, Portugal, November 2021
Azevedo, A.M., Faria-Ferreira, D., Bras, E., Silva, D., Conde; J.P., M.R. Aires-Barros, M.R., “Continuous extraction platform using aqueous two-phase systems for the integrated purification of biological products”, ESBES 2020 – virtual meeting, September, 2020
Poster Communications
Ferreira-Faria, D., Azevedo, A.M., Aires-Barros, M.R., Ferreira, A., Continuous Aqueous Two-Phase Extraction using Oscillatory Flow Reactor, ESBES 2021 – virtual meeting, May 2021
Ferreira-Faria, D., Azevedo, A.M., Aires-Barros, M.R., Ferreira, A., Continuous Aqueous Two-Phase Extraction using Oscillatory Flow Reactor, MicroBiotec, Coimbra, Portugal, December 2019
Ferreira-Faria, D., Azevedo, A.M., Aires-Barros, M.R., Ferreira, A. Continuous Aqueous Two-Phase Extraction using Oscillatory Flow Reactor, Integrated Continuous Biomanufacturing | ICB IV (ECI Engineering Conferences International), session on “Validation of ICB processes”, Cape Cod (MA, USA), October 6-10, 2019
DOCTORAL PROGRAM (36 ECTS)
Curricular Units:
• General Doctoral Training (6 ECTS)
• Advanced Experimental Techniques and methodologies (6 ECTS)
• Bioentrepreneurship (6 ECTS)
• Outreach and Teaching Skills (6 ECTS)
• Chemical Engineering Reactions (6 ECTS)
• Nanobiotechnology (6 ECTS)
Advanced courses
10th Course on Animal Cell Technology organized by the European Society of Animal Cell Technology, performed in virtual mode, from 21st to 25th September 2020
Current Position
Consultant at Avoras – Business Technology Advisory , Basel, CH