Exploration of In Situ Extraction for Enhanced Triterpenoid Production by Saccharomyces cerevisiae

Microbial Biotechnology, vol. 17, pp. e70061

Abstract

Plant-derived triterpenoids are in high demand due to their valuable applications in cosmetic, nutraceutical, and pharmaceutical industries. To meet this demand, microbial production of triterpenoids is being developed for large-scale production. However, a prominent limitation of microbial synthesis is the intracellular accumulation, requiring cell disruption during downstream processing. Destroying the whole-cell catalyst drives up production costs and limits productivity and product yield per cell. Here, in situ product extraction of triterpenoids into a second organic phase was researched to address this limitation. An organic solvent screening identified water-immiscible isopropyl myristate as a suitable in situ extractant, enabling extraction of up to 90% of total triterpenoids from engineered Saccharomyces cerevisiae. Combining isopropyl myristate and β-cyclodextrins improved extraction efficiency. In a first configuration, repeated batch fermentation with sequential product extraction and cell recycling resulted in 1.8 times higher production than a reference fermentation without in situ product extraction. In the second configuration, yeast cells were in contact with the second organic phase throughout a fed-batch fermentation to continuously extract triterpenoids. This resulted in 90% product extraction and an extended production phase. Further improvement of triterpenoid production was not achieved due to microbial host limitations uncovered through omics analyses.

Authors 7

1. Mariam Dianat Aachen Institute of Applied Microbiology

   RWTH Aachen University

   Affiliation as printed

   Institute of Applied Microbiology (iAMB), Aachen Biology and Biotechnology (ABBt) RWTH Aachen University Aachen Germany

2. Sarah Luise Straaten Aachen Institute of Applied Microbiology

   RWTH Aachen University

   Affiliation as printed

   Institute of Applied Microbiology (iAMB), Aachen Biology and Biotechnology (ABBt) RWTH Aachen University Aachen Germany

3. Aldo Maritato Aachen Institute of Applied Microbiology

   RWTH Aachen University

   Affiliation as printed

   Institute of Applied Microbiology (iAMB), Aachen Biology and Biotechnology (ABBt) RWTH Aachen University Aachen Germany

4. Daniel Wibberg

   Forschungszentrum Jülich · Bielefeld University

   Affiliation as printed

   Center for Biotechnology (CeBiTec) Bielefeld University Bielefeld Germany

   Center for Biotechnology (CeBiTec), Bielefeld University Institute of Bio, and Geosciences ‐ Computational Metagenomics (IBG‐5), Forschungszentrum Jülich GmbH ‐ Branch Office Bielefeld University Forschungszentrum Jülich GmbH Bielefeld Germany

5. Tobias Busche

   Bielefeld University

   Affiliation as printed

   Center for Biotechnology (CeBiTec) Bielefeld University Bielefeld Germany

6. Lars M. Blank corresponding Aachen Institute of Applied Microbiology

   RWTH Aachen University

   Affiliation as printed

   Institute of Applied Microbiology (iAMB), Aachen Biology and Biotechnology (ABBt) RWTH Aachen University Aachen Germany

7. Birgitta E. Ebert corresponding

   The University of Queensland

   Affiliation as printed

   Australian Institute for Bioengineering and Nanotechnology The University of Queensland St. Lucia Queensland Australia