Department of Chemical Engineering, California Institute of Technology, Pasadena, California 91125, USA.
Biotechnol Bioeng. 1993 Jun 20;42(2):215-21. doi: 10.1002/bit.260420209.
An experimental system has been constructed which enables on-line measurements of phosphorus-31 ((31)P) nuclear magnetic resonance (NMR) spectra for growing bacterial suspensions under anaerobic or aerobic conditions. A sample stream from a laboratory bioreactor is circulated to the NMR sample chamber in a gas exchange system which permits maintenance of aerobic conditions for high-cell-density cultures. (31)P NMR spectra with resolution comparable with those obtained traditionally using dense, concentrated, nongrowing cell suspensions can be obtained at cell densities above 25 g/L with acquisition times ranging from 14 to 3 minutes which decline as cell density increases. This system has been employed to characterize the changes in intracellular state of a stationary phase culture which is subjected to a transition from aerobic to anaerobic conditions. Both intracellular NTP level and cytoplasmic pH are substantially lower under anaerobic conditions. Also, the system has been employed to observe the response of a growing culture to external addition of acetate. Cells are able to maintain pH difference across the cytoplasmic membrane at extracellular acetate concentrations of 5 and 10 g/L. However, acetate concentrations of 20 g/L cause collapse of the transmembrane DeltapH and sharp reduction of the growth rate of the culture. The experimental configuration described should also permit NMR observations of many other types of microbial cultures and of other nuclei.
已经构建了一个实验系统，可以在厌氧或需氧条件下在线测量生长细菌悬浮液的磷-31（31P）核磁共振（NMR）谱。来自实验室生物反应器的样品流在气体交换系统中循环到 NMR 样品室，该系统允许为高细胞密度培养物维持需氧条件。在细胞密度高于 25 g/L 时，可以获得与传统方法相比分辨率相当的 31P NMR 谱，采集时间从 14 分钟到 3 分钟不等，随着细胞密度的增加而减少。该系统已用于表征从需氧到厌氧条件过渡的静止相培养物的细胞内状态变化。在厌氧条件下，细胞内的 NTP 水平和细胞质 pH 值都显著降低。此外，该系统还用于观察生长培养物对外加乙酸的响应。当细胞外乙酸浓度为 5 和 10 g/L 时，细胞能够维持细胞质膜两侧的 pH 值差异。然而，20 g/L 的乙酸浓度会导致跨膜 DeltapH 的崩溃和培养物生长速率的急剧下降。所描述的实验配置还应该允许对许多其他类型的微生物培养物和其他核进行 NMR 观察。
