Comparison of the Practical Resolving Power of One- and Two-Dimensional High-Performance Liquid Chromatography Analysis of Metabolomic Samples

详细信息    查看全文

• 作者：Dwight R. Stoll ; Xiaoli Wang ; Peter W. Carr
• 刊名：Analytical Chemistry
• 出版年：2008
• 出版时间：January 1, 2008
• 年：2008
• 卷：80
• 期：1
• 页码：268 - 278
• 全文大小：205K
• 年卷期：v.80,no.1(January 1, 2008)
• ISSN：1520-6882

文摘

Two-dimensional liquid chromatography (2DLC) has become a mainstay of proteomics research due to its higherpeak capacity compared to one-dimensional LC (1DLC).Because of the long analysis times typically associatedwith 2DLC (tens of hours) and its use primarily inproteomics applications, 2DLC in the context of generalHPLC has been regarded as a niche technique for use inanalysis of mixtures containing hundreds to thousandsof components compared to the far more common techniques of isocratic and gradient elution 1DLC. A significant next step in the analytical development of 2DLC isto consider using its higher resolving power to reduce theanalysis time of rather "simple" mixtures, in the range ofonly tens to hundreds of chemical constituents. The chiefobjective of this paper is to provide guidance to practitioners who need to decide whether 1DLC or 2DLC givesthe superior separation in a given analysis time. Conditional peak capacities are predicted for fully optimized1DLC and practical 2DLC separations of the low molecular weight constituents of an extract of corn seed atseveral analysis times using a model based on thechromatographic properties of compounds that are representative of real mixtures of lower molecular weightspecies. Two important corrections to the ideal 2DLCpeak capacity are made to account for both incompleteusage of the separation space and the serious effect offirst-dimension undersampling; this allows, we believe forthe first time, a fair comparison of the resolving power of1D- and 2DLC under realistic conditions. The predictedoptimum conditions are then used to carry out experimental separations of low molecular weight corn seedextract, and peaks are counted in each 1D- and 2DLCchromatogram. Based on comparisons of both the predicted peak capacities and number of peaks observed inexperimental chromatograms, we believe that practical2DLC will be superior to fully optimized gradient 1DLCfor separations lasting more than only about 10 min. Thiscrossover time is much shorter than intuitively expected,and we believe this finding will inevitably have a majorimpact on the practice of 2DLC in liquid-phase separations in general.