Experimental kS estimation: A comparison of methods for Corynebacterium glutamicum from lab to microfluidic scale

Biotechnology and Bioengineering, vol. 120, pp. 1288–1302

Abstract

Abstract Knowledge about the specific affinity of whole cells toward a substrate, commonly referred to as kS, is a crucial parameter for characterizing growth within bioreactors. State‐of‐the‐art methodologies measure either uptake or consumption rates at different initial substrate concentrations. Alternatively, cell dry weight or respiratory data like online oxygen and carbon dioxide transfer rates can be used to estimate kS. In this work, a recently developed substrate‐limited microfluidic single‐cell cultivation (sl‐MSCC) method is applied for the estimation of kS values under defined environmental conditions. This method is benchmarked with two alternative microtiter plate methods, namely high‐frequency biomass measurement (HFB) and substrate‐limited respiratory activity monitoring (sl‐RA). As a model system, the substrate affinity kS of Corynebacterium glutamicum ATCC 13032 regarding glucose was investigated assuming a Monod‐type growth response. A kS of <70.7 mg/L (with 95% probability) with HFB, 8.55 ± 1.38 mg/L with sl‐RA, and 2.66 ± 0.99 mg/L with sl‐MSCC was obtained. Whereas HFB and sl‐RA are suitable for a fast initial kS estimation, sl‐MSCC allows an affinity estimation by determining tD at concentrations less or equal to the kS value. Thus, sl‐MSCC lays the foundation for strain‐specific kS estimations under defined environmental conditions with additional insights into cell‐to‐cell heterogeneity.

Authors 8

1. Heiko Steinhoff

   Bielefeld University

   Affiliation as printed

   Center for Biotechnology (CeBiTec) Bielefeld Germany

   Multiscale Bioengineering Bielefeld University Bielefeld Germany

   Center for Biotechnology (CeBiTec), Bielefeld, Germany

   Multiscale Bioengineering, Bielefeld University, Bielefeld, Germany

2. Maurice Finger Aachen AVT‑Biochemical Engineering

   RWTH Aachen University

   Affiliation as printed

   AVT – Biochemical Engineering RWTH Aachen University Aachen Germany

3. Michael Osthege Aachen Institute of Biotechnology

   Forschungszentrum Jülich · RWTH Aachen University

   Affiliation as printed

   Institute of Biotechnology RWTH Aachen University Aachen Germany

   Institute of Bio‐ and Geoscience IBG‐1: Biotechnology Jülich Germany

   Institute of Biotechnology, RWTH Aachen University, Aachen, Germany

4. Corinna Golze

   Bielefeld University

   Affiliation as printed

   Multiscale Bioengineering Bielefeld University Bielefeld Germany

   Multiscale Bioengineering, Bielefeld University, Bielefeld, Germany

5. Simone Schito

   Forschungszentrum Jülich

   Affiliation as printed

   Institute of Bio‐ and Geoscience IBG‐1: Biotechnology Jülich Germany

6. Stephan Noack

   Forschungszentrum Jülich

   Affiliation as printed

   Institute of Bio‐ and Geoscience IBG‐1: Biotechnology Jülich Germany

7. Jochen Büchs Aachen AVT‑Biochemical Engineering

   RWTH Aachen University

   Affiliation as printed

   AVT – Biochemical Engineering RWTH Aachen University Aachen Germany

8. Alexander Grünberger corresponding

   Karlsruhe Institute of Technology · Bielefeld University

   Affiliation as printed

   Center for Biotechnology (CeBiTec) Bielefeld Germany

   Microsystems in Bioprocess Engineering, Institute of Process Engineering in Life Sciences Karlsruhe Institute of Technology Karlsruhe Germany

   Multiscale Bioengineering Bielefeld University Bielefeld Germany

   Center for Biotechnology (CeBiTec), Bielefeld, Germany

   Microsystems in Bioprocess Engineering, Institute of Process Engineering in Life Sciences, Karlsruhe Institute of Technology, Karlsruhe, Germany

   Multiscale Bioengineering, Bielefeld University, Bielefeld, Germany