Faculty of Life and Environmental Sciences, University of Tsukuba, Tsukuba, Ibaraki, 305-8572, Japan.
Combustion of Thermo and Fluid Dynamics, Department of Fundamental Technology, Tokyo Gas Co. Ltd., Yokohama, Kanagawa, 230-0045, Japan.
Sci Rep. 2020 Jun 30;10(1):10385. doi: 10.1038/s41598-020-66810-3.
Shake-flask cultures of microorganisms involve flame sterilization during sampling, which produces combustion gas with high CO concentrations. The gaseous destination has not been deeply analyzed. Our aim was to investigate the effect of flame sterilization on the headspace of the flask and on the shake-flask culture. In this study, the headspace CO concentration was found to increase during flame sterilization ~0.5-2.0% over 5-20 s empirically using the Circulation Direct Monitoring and Sampling System. This CO accumulation was confirmed theoretically using Computational Fluid Dynamics; it was 9% topically. To evaluate the influence of CO accumulation without interference from other sampling factors, the flask gas phase formed by flame sterilization was reproduced by aseptically supplying 99.8% CO into the headspace, without sampling. We developed a unit that can be sampled in situ without interruption of shaking, movement to a clean bench, opening of the culture-plug, and flame sterilization. We observed that the growth behaviour of Escherichia coli, Pelomonas saccharophila, Acetobacter pasteurianus, and Saccharomyces cerevisiae was different depending on the CO aeration conditions. These results are expected to contribute to improving microbial cell culture systems.
摇瓶培养微生物时，在取样过程中需要进行火焰灭菌，这会产生高浓度 CO 的燃烧气体。目前尚未对这种气态产物进行深入分析。我们的目的是研究火焰灭菌对瓶内空间和摇瓶培养的影响。在这项研究中，我们发现使用 Circulation Direct Monitoring and Sampling System 进行经验性测试时，火焰灭菌过程中瓶内空间的 CO 浓度会在 5-20 秒内增加约 0.5-2.0%。使用计算流体动力学对 CO 的积累进行了理论验证，其值为 9%。为了评估 CO 积累对培养的影响，而不受其他采样因素的干扰，我们通过将 99.8%的 CO 无菌注入瓶内空间，而不进行采样，重现了火焰灭菌时形成的瓶内气相。我们开发了一种可以在不中断摇瓶、不移动到超净工作台、不打开培养塞和不进行火焰灭菌的情况下进行原位采样的装置。我们观察到大肠杆菌、糖单胞菌、巴氏醋酸杆菌和酿酒酵母的生长行为因 CO 曝气条件而异。这些结果有望有助于改善微生物细胞培养系统。
