Monitoring of CO2 and O2 concentrations in the headspace of Sakaguchi flasks during liquid culture of microorganism
CO2 and O2 in the Sakaguchi flask headspace during culture was monitored via circulation direct monitoring and sampling system (CDMSS), a device with circulation bypass system. In static culture with Saccharomyces cerevisiae (circulation rate, 50 mL/min), a vertical CO2 concentration gradient (maximum gap ~ 2% (v/v) [height from the bottom of flask 45 mm, 7%; 155 mm, 5%]) in the Sakaguchi flask headspace was observed; no concentration O2 gradient was observed. However, shake flask culture showed vertical gradient concentrations for both CO2 and O2 (maximum gap of CO2 and O2 concentrations: 2 and 4% [heights from the bottom of flask 115 mm, 6.0 and 9.5%; 175 mm, 4.0 and 13.5%], respectively). When the CDMSS circulation rate in the Sakaguchi flask headspace was 300 or 400 mL/min, the gaseous environment was uniformly distributed so that no vertical gradient concentration was observed. In shaking culture with Escherichia coli under these conditions, CO2 was accumulated at high concentrations in the headspace and culture broth (maximum values 8%, in the headspace; 120 mg/L, in the culture broth). Most of the accumulated CO2 in the headspace could be removed by inserting a column packed with CO2 adsorbent at the bypass port of the CDMSS gaseous circulation. Thus, dissolved CO2 was maintained at a lower concentration, and the final UOD (unit optical density) value of culture was increased compared with that of the control. This study is the first to demonstrate that vertical gradients of CO2 and O2 concentrations exist in the headspace of Sakaguchi flask during culture.
KeywordsBatch culture Carbon dioxide Monitoring device Oxygen Sampling operation Shake-flask culture
The manuscript has been carefully edited by native English-speaking professional editors from Editage, a division of Cactus Communications.
This work was supported by a grant-in-aid from the Japan Society for the Promotion of Science (JSPS) (fellowship 16J00800). This work was a funded by a JSPS KAKENHI (grant number 15H04569), JSPS KAKENHI Challenging Research [Exploratory] (grant number 17K19218), and The Sumitomo Electric Industries Group Corporate Social Responsibility Foundation.
Compliance with ethical standards
Conflict of interest
The authors declare that they have no conflict of interest.
This article does not contain any studies with human participants or animals performed by any of the authors.
Masato Takahashi (Postdoctoral researcher; Research fellow of Japan Society for the Promotion of Science). Faculty of Life and Environmental Sciences, University of Tsukuba, Tsukuba, Ibaraki 305–8572, Japan.
Hideki Aoyagi (Professor and Principal investigator). Faculty of Life and Environmental Sciences, University of Tsukuba, Tsukuba, Ibaraki 305–8572, Japan.
MT and HA designed the research. HA supervised the research. MT created the CDMSS. MT performed all culture experiments. MT and HA analysed the data. MT and HA wrote the manuscript. All authors read and approved the final manuscript.
- Ali M, Ismaini, Depamede SN, Setyono BDH, Mukhlis A, Amin M, Ashari M (2015) Stirred bioreactor for the robustness production of recombinant GST.VP28 in fed-batch cultivation of Escherichia coli. Sci Study Res Chem Chem Eng Biotechnol Food Ind 16:245–252Google Scholar
- Kato I, Tanaka H (1998) Influence of CO2 ventilation on microbial cultivation in shake-flasks. Biotechnol Tech 12:325–328Google Scholar
- Kluyver A, Perquin L (1933) Zur methodik der schimmel-stoffwechseluntersuchung. Biochem Z 266:68–81Google Scholar
- Shiota H, Sakaguchi K (1950) Studies on the production of mold protease in the submerged culture. Part I. J Agric Chem Soc Japan 23:426–429Google Scholar
- Takahashi M, Aoyagi H (2018a) Effect of intermittent opening of breathable culture plugs and aeration of headspace on the structure of microbial communities in shake-flask culture. J Biosci Bioeng. https://doi.org/10.1016/j.jbiosc.2018.01.009