Miniature bioreactors for automated high‐throughput bioprocess design (HTBD): reproducibility of parallel fed‐batch cultivations with Escherichia coli

Abstract

To verify the reproducibility of cultivations of Escherichia coli in novel millilitre-scale bioreactors, fully automated fed-batch cultivation was performed in seven parallel-operated ml-scale bioreactors with an initial volume of 10 ml/reactor. The process was automatically controlled by a liquid-handling system responsible for glucose feeding, titration and sampling. Atline analysis (carried out externally of the reaction vessel with a short time delay) comprised automated pH and attenuance measurements. The partial pressure of oxygen (pO2) was measured online by a novel fluorimetric sensor block measuring the fluorescence lifetime of fluorophors immobilized inside the millilitre-scale bioreactors. Within a process time of 14.6 h, the parallel cultivation yielded a dry cell weight of 36.9+/-0.9 g.l(-1). Atline pH measurements were characterized by an S.D. of <1.1% throughout the process. Computational-fluid-dynamics simulation of single-phase flow yields a mean power input of 21.9 W.l(-1) at an impeller speed of 2800 rev./min corresponding to a power number (NP) of 3.7.

Scale-down and parallel operation of the riboflavin production process with Bacillus subtilis

“…In the latter case, the required oxygen transfer rate should be compensated by an augmented addition of oxygen to the gas flow, if this does not influence the biological system negatively. Regarding the shear stress, the mean volumetric power input in the milliliter reactors is 21.9 W L -1 at 2800 rpm for 11.2 mL according to simulations using computational fluid dynamics (Puskeiler, 2005b). This is similar to the mean power input in stirred tank reactors at laboratory scale.…”

“…Cultivations can be carried out in batch or in fully automated intermittent fedbatch mode with automated parallel compensation of pH changes and evaporation loss (Puskeiler et al, 2005b). However, the discontinuous liquid addition implies modifications compared to the standard continuous fed-batch operation and causes concentration dynamics, which can possibly affect the physiology of the used microorganism and thereby the overall process performance.…”

“…, increasing with rising stirrer speed and decreasing liquid volume, assessed for liquid volumes between 7 and 14 mL (Puskeiler, 2004 at an impeller speed of 2800 rpm (Puskeiler et al, 2005b). The setup is automated with a laboratory robot for intermittent liquid additions to realize feeding and pH adjustment and for parallel sampling from the reactors into microtiter plates for at-line analyses.…”

An automated workflow for enhancing microbial bioprocess optimization on a novel microbioreactor platform

“…The miniaturized stirred reactors can be handled automatically e.g. with an integration of the vessels in liquid-handling platforms [15,20,21]. With such a setup, repeated sampling during cultivations for offline biomass and product quantification for example in an adjacent spectrophotometer as described by Knorr et al [22] can be carried out.…”

Miniaturization in Biocatalysis

“…Generation of detailed predictive models, representative of the dynamics of flow, which can incorporate reaction data, has been made possible with computational fluid dynamics (CFD). CFD allows therefore for the design and analysis of a given system, as well as for the simulation of fluid flow, (bio)chemical reaction, mass and heat transfer [19,62,87,128,129]. The models developed within the scope of CFD methods are typically anchored in the Navier–Stokes equations and in the convection-diffusion equation [19,130].…”