A novel milliliter-scale chemostat system for parallel cultivation of microorganisms in stirred-tank bioreactors

Schmideder, Andreas; Severin, Timm Steffen; Cremer, Johannes H.; Weuster‐Botz, Dirk

doi:10.1016/j.jbiotec.2015.06.402

Cited by 35 publications

(39 citation statements)

References 29 publications

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“…2017, 17,[1215][1216][1217][1218][1219][1220] the mass-transfer boundaries of the bioreactor. The classical approach for investigations of the productivity at different dilution rates is to perform a series of chemostat experiments, which is very tedious and has been recently scaled down to minibioreactors [12]. Another option for accelerated data collection from continuous cultures is the change-stat technique.…”

Section: Introductionmentioning

confidence: 99%

Detection of growth rate‐dependent product formation in miniaturized parallel fed‐batch cultivations

Glauche

Glazyrina

Bournazou

et al. 2017

Engineering in Life Sciences

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Saccharomyces cerevisiae is a popular expression system for recombinant proteins. In most cases, production processes are performed as carbon-limited fed-batch cultures to avoid aerobic ethanol formation. Especially for constitutive expression systems, the specific product formation rate depends on the specific growth rate. The development of optimal feeding strategies strongly depends on laboratory-scale cultivations, which are time and resource consuming, especially when continuous experiments are carried out. It is therefore beneficial for accelerated process development to look at alternatives. In this study, S. cerevisiae AH22 secreting a heterologous endo-polygalacturonase (EPG) was characterized in microwell plates with an enzyme-based fed-batch medium. Through variation of the glucose release rate, different growth profiles were established and the impact on EPG secretion was analyzed. Product formation rates of 200-400 U (g x h) −1 were determined. As a reference, bioreactor experiments using the change-stat cultivation technique were performed. The growth-dependent product formation was analyzed over dilution rates of D = 0.01-0.35 with smooth change of D at a rate of 0.003 h −2 . EPG production was found to be comparable with a q p of 400 U (g x h) −1 at D = 0.27 h −1 . The presented results indicate that parallel miniaturized fed-batch cultures can be applied to determine product formation profiles of putative production strains. With further automation and parallelization of the concept, strain characterization can be performed in shorter time.

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Section: Introductionmentioning

confidence: 99%

Detection of growth rate‐dependent product formation in miniaturized parallel fed‐batch cultivations

Glauche

Glazyrina

Bournazou

et al. 2017

Engineering in Life Sciences

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“…After batch phase, the growth rate could be maintained at 0.154 h −1 (µ set = 0.15 h −1 ). The growth of E. coli was clearly limited by glucose after depletion of the batch substrate, because the measured glucose concentrations (< 25 mg L −1 ) were below the substrate affinity constant of 90 mg L −1 of glucose for the applied E. coli strain . The induction was carried out at a CDW of approximately 29 g L −1 .…”

Section: Resultsmentioning

confidence: 99%

“…The speed of the gas‐inducing impellers was set to 3000 rpm and the headspace of each bioreactor was rinsed with 0.1 L min −1 of sterile air. Losses in liquid volume by evaporation were prevented by pre‐saturating the inlet air with water at room temperature and setting the headspace cooling of the reactors to 20°C . The inlet and efflux of medium was realized by multi‐channel peristaltic pumps (MP8, DASGIP–an Eppendorf company, Jülich, Germany).…”

Section: Methodsmentioning

confidence: 99%

“…The at‐line measurement of the optical density at 600 nm (OD 600 ) and the cell dry weight (CDW) at milliliter and liter‐scale was carried out as described before …”

Section: Methodsmentioning

confidence: 99%

“…Hence, innovative methods for the timesaving and cost‐reducing identification of suitable reaction conditions for recombinant protein production processes in fed‐batch mode would be of great benefit. A highly useful tool to investigate the influence of different reaction conditions on the metabolism of microorganisms are chemostats, which are frequently employed for the identification of kinetic parameters of non‐induced growing cells . However, continuous operation results in time‐consuming and expensive experiments .…”

Section: Introductionmentioning

confidence: 99%

See 2 more Smart Citations

Parallel steady state studies on a milliliter scale accelerate fed‐batch bioprocess design for recombinant protein production with Escherichia coli

Schmideder

Weuster‐Botz

2016

Biotechnology Progress

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In general, fed-batch processes are applied for recombinant protein production with Escherichia coli (E. coli). However, state of the art methods for identifying suitable reaction conditions suffer from severe drawbacks, i.e. direct transfer of process information from parallel batch studies is often defective and sequential fed-batch studies are time-consuming and cost-intensive. In this study, continuously operated stirred-tank reactors on a milliliter scale were applied to identify suitable reaction conditions for fed-batch processes. Isopropyl β-d-1-thiogalactopyranoside (IPTG) induction strategies were varied in parallel-operated stirred-tank bioreactors to study the effects on the continuous production of the recombinant protein photoactivatable mCherry (PAmCherry) with E. coli. Best-performing induction strategies were transferred from the continuous processes on a milliliter scale to liter scale fed-batch processes. Inducing recombinant protein expression by dynamically increasing the IPTG concentration to 100 µM led to an increase in the product concentration of 21% (8.4 g L ) compared to an implemented high-performance production process with the most frequently applied induction strategy by a single addition of 1000 µM IPGT. Thus, identifying feasible reaction conditions for fed-batch processes in parallel continuous studies on a milliliter scale was shown to be a powerful, novel method to accelerate bioprocess design in a cost-reducing manner. © 2016 American Institute of Chemical Engineers Biotechnol. Prog., 32:1426-1435, 2016.

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Continuous Bioprocess Development: Methods for Control and Characterization of the Biological System

Neubauer

Bournazou

2017

Continuous Biomanufacturing ‐ Innovative Technologies and Methods

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A novel milliliter-scale chemostat system for parallel cultivation of microorganisms in stirred-tank bioreactors

Cited by 35 publications

References 29 publications

Detection of growth rate‐dependent product formation in miniaturized parallel fed‐batch cultivations

Detection of growth rate‐dependent product formation in miniaturized parallel fed‐batch cultivations

Parallel steady state studies on a milliliter scale accelerate fed‐batch bioprocess design for recombinant protein production with Escherichia coli

Continuous Bioprocess Development: Methods for Control and Characterization of the Biological System

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