Two‐dimensional separation system of coupling capillary liquid chromatography to capillary electrophoresis for analysis of Escherichia coli metabolites
Two‐dimensional separation system of coupling capillary liquid chromatography to capillary electrophoresis for analysis of Escherichia coli metabolites
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DOI：
10.1002/elps.200500308
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发表时间：
2005-09
期刊：
影响因子：
2.9
通讯作者：
L. Jia;N. Tanaka;S. Terabe
中科院分区：
文献类型：
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作者：
L. Jia;N. Tanaka;S. Terabe
建立了一种用于大肠杆菌代谢产物分析的色谱-电泳耦合二维分离系统。采用硅胶-十八烷基硅胶整体柱（500×0.2 mm ID）的毛细管液相色谱（LC）作为第一维，通过毛细管电泳（CE）作为第二维进一步分析流出液馏分。场增强堆叠被选择性地用作连接两个维度的浓缩策略，这被证明有利于代谢物的检测。使用含有118个标准品的人工样品来优化二维分离系统，其中一些标准品缺乏发色团或具有弱的紫外吸收。在最佳条件下，在254 nm处有吸光度的人工样品中63个组分可被很好地分辨和检测。通过对E.大肠杆菌代谢产物。与我们在Anal. 2004，76，1419-1428中，在第一维度上使用较长的整体柱提高了分离效率，并提供了在不损害分离效率的情况下增加注射体积的可能性。在第二维，场增强堆积用于提高代谢物的浓度灵敏度，并且E.大肠杆菌细胞提取物进行检测和鉴定，使用开发的2-D分离系统。此外，还通过在毛细管LC和CE技术中使用挥发性缓冲液，对未来的CE-质谱耦合进行了初步研究。
A two‐dimensional (2‐D) separation system of coupling chromatography to electrophoresis was developed for profiling Escherichia coli metabolites. Capillary liquid chromatography (LC) with a monolithic silica‐octadecyl silica column (500×0.2 mm ID) was used as the first dimension, from which the effluent fractions were further analyzed by capillary electrophoresis (CE) acting as the second dimension. Field‐enhanced stacking was selectively employed as a concentration strategy to interface the two dimensions, which proved to be beneficial for the detection of metabolites. An artificial sample containing 118 standards, some of which lack chromophores or have weak UV absorbance, was used to optimize the 2‐D separation system. Under the optimum conditions, 63 components in the artificial sample having absorbance at 254 nm could be well resolved and detected. The utility of the system was demonstrated by comprehensive analysis of E. coli metabolites. Comparing with the previous 2‐D separation system we published in Anal. Chem. 2004, 76, 1419–1428, using a longer monolithic column in the first dimension improved the separation efficiency and offered the possibility of increasing the injection volume without compromising the separation efficiency. In the second dimension, field‐enhanced stacking was used to improve the concentration sensitivity of the metabolites, and more metabolites in E. coli cell extract were detected and identified using the developed 2‐D separation system. In addition, preliminary investigation for future CE‐mass spectrometry coupling was also made in the study by using volatile buffers in the capillary LC and CE techniques.