Department of Chemistry, University of British Columbia, Vancouver, British Columbia, Canada.
Department of Urology, University of California, Los Angeles, Los Angeles, California, USA.
Electrophoresis. 2022 Aug;43(15):1626-1637. doi: 10.1002/elps.202100365. Epub 2022 Jun 1.
Migration time fluctuation strongly affects peak alignment and identification of unknown compounds, making migration time correction an essential step in capillary electrophoresis (CE)-based metabolomics. To obtain more reliable information, metabolites with different apparent mobilities are analyzed by tandem mass spectrometry. Applying a small pressure is a common practice for reducing the analysis time of anions in a positive mode CE, known as the pressure-assisted CE. However, applying pressure may reduce the separation efficiency and can be undesirable for cation analysis. A simple way to address this issue is to increase the pressure after a certain time, during the separation. We term this practice as dual pressure CE. However, changing the pressure during the CE separation complicates migration time correction. Previous migration time correction methods were established based on a consistent electroosmotic flow and a constant pressure-driven bulk-flow velocity. We proposed a new correction method to support the peak alignment when dual pressure CE is used. A Python-based script was developed to implement dual pressure CE migration time correction for semi-targeted metabolomics study performed by a multiple reaction monitoring-based method. This script can help select suitable endogenous metabolites as correction markers, perform migration time correction, and conduct peak alignment. A case study showed that migration time precision of 156 metabolites in 32 samples can be improved from 4.8 to 11.4%RSD (relative standard deviation) to less than 1.8%RSD.
迁移时间波动会强烈影响峰对齐和未知化合物的识别，因此在基于毛细管电泳（CE）的代谢组学中，迁移时间校正成为必不可少的步骤。为了获得更可靠的信息，采用串联质谱法分析具有不同表观迁移率的代谢物。在正模式 CE 中，施加较小的压力是减少分析阴离子所需时间的常用方法，称为压力辅助 CE。然而，施加压力可能会降低分离效率，并且对于阳离子分析可能不理想。解决此问题的一种简单方法是在分离过程中的某个时间后增加压力。我们将这种做法称为双压 CE。然而，在 CE 分离过程中改变压力会使迁移时间校正变得复杂。以前的迁移时间校正方法是基于恒定的电渗流和恒定的压力驱动的整体流速建立的。当使用双压 CE 时，我们提出了一种新的校正方法来支持峰对齐。我们开发了一个基于 Python 的脚本，用于执行基于多重反应监测的半靶向代谢组学研究中的双压 CE 迁移时间校正。该脚本可以帮助选择合适的内源性代谢物作为校正标记，进行迁移时间校正，并进行峰对齐。案例研究表明，32 个样本中 156 种代谢物的迁移时间精度可以从 4.8%RSD（相对标准偏差）提高到 11.4%RSD 以下，至小于 1.8%RSD。
