A dynamic method for the investigation of induced state metabolic capacities as a function of temperature

Science-based recombinant bioprocess designs as well as the design of statistical experimental plans for process optimization (Design of Experiments, DoE) demand information on physiological bioprocess boundaries, such as the onset of acetate production, adaptation times, mixed feed metabolic capabilities or induced state maximum metabolic rates as at the desired cultivation temperature. Dynamic methods provide experimental alternatives to determine this information in a fast and efficient way. Information on maximum metabolic capabilities as a function of temperature is needed in case a reduced cultivation temperature is desirable (e.g. to avoid inclusion body formation) and an appropriate feeding profile is to be designed.

Here, we present a novel dynamic method for the determination of the specific growth rate as a function of temperature for induced recombinant bacterial bioprocesses. The method is based on the control of the residual substrate concentration at non-limiting conditions with dynamic changes in cultivation temperature. The presented method was automated in respect to information extraction and closed loop control by means of in-line Fourier Transformation Infrared Spectroscopy (FTIR) residual substrate measurements and on-line first principle rate-based soft-sensors. Maximum induced state metabolic capabilities as a function of temperature were successfully extracted for a recombinant E. coli C41 fed-batch bioprocess without the need for sampling in a time frame of 20 hours.

The presented method was concluded to allow the fast and automated extraction of maximum metabolic capabilities (specific growth rate) as a function of temperature. This complements the dynamic toolset necessary for science-based recombinant bacterial bioprocess design and DoE design.