Wahl S Aljoscha, Seifar Reza Maleki, Ten Pierick Angela, Ras Cor, van Dam Jan C, Heijnen Joseph J, van Gulik Walter M
Department of Biotechnology, Delft University of Technology, Julianalaan 67, 2628 BC, Delft, The Netherlands,
Methods Mol Biol. 2014;1191:91-105. doi: 10.1007/978-1-4939-1170-7_6.
Quantitative intracellular metabolite measurements are essential for systems biology and modeling of cellular metabolism. The MS-based quantification is error prone because (1) several sampling processing steps have to be performed, (2) the sample contains a complex mixture of partly compounds with the same mass and similar retention time, and (3) especially salts influence the ionization efficiency. Therefore internal standards are required, best for each measured compound. The use of labeled biomass, (13)C extract, is a valuable tool, reducing the standard deviations of intracellular concentration measurements significantly (especially regarding technical reproducibility). Using different platforms, i.e., LC-MS and GC-MS, a large number of different metabolites can be quantified (currently about 110).
细胞内代谢物的定量测量对于系统生物学和细胞代谢建模至关重要。基于质谱的定量容易出错，原因如下：（1）必须执行多个采样处理步骤；（2）样品包含部分具有相同质量和相似保留时间的化合物的复杂混合物；（3）尤其是盐会影响电离效率。因此，需要内标，最好是针对每种被测化合物。使用标记的生物质，即¹³C提取物，是一种有价值的工具，可显著降低细胞内浓度测量的标准偏差（特别是在技术可重复性方面）。使用不同的平台，即液相色谱 - 质谱联用仪（LC-MS）和气相色谱 - 质谱联用仪（GC-MS），可以对大量不同的代谢物进行定量（目前约为110种）。
