Institut für Bioverfahrenstechnik, Universität Stuttgart, Allmandring 31, 70569 Stuttgart, Germany.
Biotechnol Bioeng. 1997 Jul 20;55(2):305-16. doi: 10.1002/(SICI)1097-0290(19970720)55:2<305::AID-BIT8>3.0.CO;2-M.
The goal of this work was to obtain rapid sampling technique to measure transient metabolites in vivo. First, a pulse of glucose was added to a culture of the yeast Saccharomyces cerevisiae growing aerobically under glucose limitation. Next, samples were removed at 2 to 5 s intervals and quenched using methods that depend on the metabolite measured. Extracellular glucose, excreted products, as well as glycolytic intermediates (G6P, F6P, FBP, GAP, 3-PG, PEP, Pyr) and cometabolites (ATP, ADP, AMP, NAD(+), NADH) were measured using enzymatic or HPLC methods. Significant differences between the adenine nucleotide concentrations in the cytoplasm and mitochondria indicated the importance of compartmentation for the regulation of the glycolysis. Changes in the intra- and extracellular levels of metabolites confirmed that glycolysis is regulated on a time scale of seconds. (c) 1997 John Wiley & Sons, Inc. Biotechnol Bioeng 55: 305-316, 1997.
本工作旨在获得快速采样技术以测量体内的瞬态代谢物。首先，向在葡萄糖限制下有氧生长的酵母酿酒酵母的培养物中添加葡萄糖脉冲。然后，每隔 2 到 5 秒取出样品，并根据要测量的代谢物使用依赖于代谢物的方法进行淬灭。使用酶或 HPLC 方法测量细胞外葡萄糖、分泌产物以及糖酵解中间产物（G6P、F6P、FBP、GAP、3-PG、PEP、Pyr）和共代谢物（ATP、ADP、AMP、NAD(+)、NADH）。细胞质和线粒体中腺嘌呤核苷酸浓度的显著差异表明区室化对于糖酵解的调节很重要。代谢物的细胞内外水平的变化证实了糖酵解在秒的时间尺度上受到调节。（c）1997 年 John Wiley & Sons，Inc.《生物工艺学与生物工程》55：305-316，1997 年。
