Analysis of porous breathable stopper and development of PID control for gas phase during shake-flask culture with micro
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APPLIED MICROBIAL AND CELL PHYSIOLOGY
Analysis of porous breathable stopper and development of PID control for gas phase during shake-flask culture with microorganisms Masato Takahashi 1
&
Hideki Aoyagi 1
Received: 1 June 2020 / Revised: 6 August 2020 / Accepted: 17 August 2020 # Springer-Verlag GmbH Germany, part of Springer Nature 2020
Abstract We evaluated the ventilation ability of two types (plug-type and cap-type) of culture-stoppers having standard air permeability. The culture-stoppers were evaluated using the circulation direct monitoring and sampling system with CO2 concentration in the gas phase of a shake-flask culture as an index. The half-lives of CO2 in the headspace of the shake flask with the plug-type and cap-type stoppers were about 51.5 min and about 30.3 min, respectively. Based on these half-lives, we formulated a model equation to simulate the behaviour of CO2 with different culture-stoppers. After validating the model equation by shake-flask culture with Saccharomyces cerevisiae, we investigated the effect of different ventilation abilities of the culture-stoppers on the growth of Pelomonas saccharophila and Escherichia coli: the sensitivity of the culture-stopper to the ventilation ability was dependent on the microorganism species. In the case of P. saccharophila, when the plug-type culture-stopper was combined with controlled CO2 concentration (6%) in the flask, the maximum yield increased by twofold compared to that of the control. This study shows the importance of ventilation in headspace and conventional culture-stoppers during the shake-flask culture of microorganisms. The problems that may occur between the conventional shake-flask culture approach using a breathable culturestopper and the next-generation shake-flask culture without a conventional culture-stopper were clarified from the evaluation of gas-permeable culture-stoppers. The importance of controlled gaseous phase in the headspace during shake-flask culture of the microorganisms was also elucidated. Key points • Ventilation capacity of culture-stoppers was evaluated using the CO2 half-life concentration. • Behaviour of microorganisms varies with the type of culture-stopper. • Developed a PID system for control of CO2 in flask gas phase to enhance the shake-flask culture. Keywords Breathable culture-stopper . Carbon dioxide . Monitoring device . PID control . Shake-flask culture
Introduction Shake-flask culture was developed as an alternative to surface culture of Aspergillus flavus, which was the conventional method for aerobic cultures (Kluyver and Perquin 1932). Shake-flask culture is one of the most popular Electronic supplementary material The online version of this article (https://doi.org/10.1007/s00253-020-10847-x) contains supplementary material, which is available to authorized users. * Hideki Aoyagi [email protected] 1
Faculty of Life and Environmental Sciences, University of Tsukuba, Tsukuba, Ibaraki 305-8572, Japan
submerged culture methods used for various cells, such as microorganism, a
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