Optimized polymer-based glucose release in microtiter plates for small-scale E. coli fed-batch cultivations

Journal of Biological Engineering, vol. 14, pp. 24

Abstract

Abstract Background Small-scale cultivation vessels, which allow fed-batch operation mode, become more and more important for fast and reliable early process development. Recently, the polymer-based feeding system was introduced to allow fed-batch conditions in microtiter plates. Maximum glucose release rates of 0.35 mg/h per well (48-well-plate) at 37 °C can be achieved with these plates, depending on the media properties. The fed-batch cultivation of fluorescent protein-expressingE. coliat oxygen transfer rate levels of 5 mmol/L/h proved to be superior compared to simple batch cultivations. However, literature suggests that higher glucose release rates than achieved with the currently available fed-batch microtiter plate are beneficial, especially for fast-growing microorganisms. During the fed-batch phase of the cultivation, a resulting oxygen transfer rate level of 28 mmol/L/h should be achieved. Results Customization of the polymer matrix enabled a considerable increase in the glucose release rate of more than 250% to up to 0.90 mg/h per well. Therefore, the molecular weight of the prepolymer and the addition of a hydrophilic PDMS-PEG copolymer allowed for the individual adjustment of a targeted glucose release rate. The newly developed polymer matrix was additionally invariant to medium properties like the osmotic concentration or the pH-value. The glucose release rate of the optimized matrix was constant in various synthetic and complex media. Fed-batch cultivations ofE. coliin microtiter plates with the optimized matrix revealed elevated oxygen transfer rates during the fed-batch phase of approximately 28 mmol/L/h. However, these increased glucose release rates resulted in a prolonged initial batch phase and oxygen limitations. The newly developed polymer-based feeding system provides options to manufacture individual feed rates in a range from 0.24–0.90 mg/h per well. Conclusions The optimized polymer-based fed-batch microtiter plate allows higher reproducibility of fed-batch experiments since cultivation media properties have almost no influence on the release rate. The adjustment of individual feeding rates in a wide range supports the early process development for slow, average and fast-growing microorganisms in microtiter plates. The study underlines the importance of a detailed understanding of the metabolic behavior (through online monitoring techniques) to identify optimal feed rates.

Authors 4

1. Timm Keil Aachen AVT‑Biochemical Engineering

   RWTH Aachen University

   Affiliation as printed

   AVT - Biochemical Engineering, RWTH Aachen University, Forckenbeckstraße 51, 52074 Aachen, Germany

   AVT - Biochemical Engineering, RWTH Aachen University, Forckenbeckstraße 51, 52074, Aachen, Germany

2. Barbara Dittrich Aachen DWI - Leibniz Institute for Interactive Materials

   DWI – Leibniz Institute for Interactive Materials

   Affiliation as printed

   DWI - Leibniz Institute for Interactive Materials, Forckenbeckstraße 50, 52074 Aachen, Germany

   DWI – Leibniz Institute for Interactive Materials, Forckenbeckstraße 50, 52074, Aachen, Germany

3. Clemens Lattermann
   Affiliation as printed

   Kuhner Shaker GmbH, Kaiserstraße 100, 52134 Herzogenrath, Germany

   Kuhner Shaker GmbH, Kaiserstraße 100, 52134, Herzogenrath, Germany

4. Jochen Büchs Aachen AVT‑Biochemical Engineering

   RWTH Aachen University

   Affiliation as printed

   AVT - Biochemical Engineering, RWTH Aachen University, Forckenbeckstraße 51, 52074 Aachen, Germany

   AVT - Biochemical Engineering, RWTH Aachen University, Forckenbeckstraße 51, 52074, Aachen, Germany