基于毛细管电泳-质谱联用技术的代谢/蛋白质组学分析

doi:10.3724/SP.J.1123.2020.02025

摘要/Abstract

摘要：

毛细管电泳-质谱（CE-MS）联用技术具有高灵敏度、高分析效率、低样品损耗等优点，在强极性和带电荷的物质分析中具有明显优势，已广泛应用于生命科学、医学、药学等多个领域。在过去的十几年，影响其应用的主要因素包括系统的稳定性、实验的可重复性、数据的准确性等。为解决现有问题，进一步拓展其应用，研究人员在技术设计和改进等方面做了大量工作。医学和分析化学领域的相关研究证明了CE-MS在代谢组学和蛋白质组学中的实用性。这篇文章综述了2015年以来，CE-MS在技术和应用方面的最新进展，为未来的技术发展及应用提供借鉴。为提高CE-MS的分析效率和数据可比性，该文对多个方面的研究进行了讨论，包括涂层与样品的相互作用、接口技术、运行参数和数据处理方法。文中关于复杂样品（组织、细胞、体液等）代谢组学/蛋白质组学的综述研究，使癌症病理分析、药物开发和疾病监测等分析数据更加可视化，为CE-MS临床分析应用提供借鉴。除了对CE-MS的最新发展进行综合评述外，还提出未来应加强3个方面的研究：（i）从样品前处理和分离技术方面优化分析方法；（ii）从毛细管涂层和接口技术方面调整分析技术；（iii）从临床研究和数据分析方面开发新思路。

关键词: 毛细管电泳-质谱联用, 技术改进, 代谢组学, 蛋白质组学, 综述

Abstract:

Capillary electrophoresis-mass spectrometry (CE-MS) has the advantages of higher sensitivity, higher efficiency, and less sample consumption. Moreover, it possesses obvious advantages during the analysis of strongly charged and highly polar samples. CE-MS has been widely applied in life sciences, medicine, and pharmacology. In the past ten years, the main factors affecting its application were system stability, reproducibility, and data accuracy. In order to solve the existing problems of CE-MS, researchers have invested significant effort in technology innovation to further expand CE-MS application. In the fields of medicine and analytical chemistry, substantial research indicates that CE-MS is superior compared to other metabolomic and proteomic approaches.

This study aims at reviewing the latest methods and applications developed in the fields of medicine and analytical chemistry since 2015. Furthermore, it also aims at enhancing the technology development-related application value of CE-MS and serving as a reference for future development. Further development of the CE-MS technology is discussed from the aspects of coating-sample interaction, interface types, and data processing methods. Concerning the coating types, neutral coatings had been applied extensively in CE-MS and there should be no limitation to the charge of the analyte. The coating decreased sample adsorption on the inner wall by covering the surface charge, greatly reducing the electroosmotic flow (EOF). A charged capillary coating could modify such an EOF direction. The cationic coating could reduce the hydrophobic interaction between the sample and the capillary column, resulting in higher EOF. If it is applied to the sheathless interface, the resolution could be improved by extending the capillary length. Anionic coatings are predominant among the anionic compounds, shortening the separation time by reducing the interaction between the anionic compounds and the capillary. The coating type should be chosen relative to the analyte characteristics. Concerning the interface technology, all interfaces should be simple, practical, and non-dependent on sheath liquid and background electrolytes. As far as data processing methods are concerned, it is necessary to design and develop a practical method for span space data comparison and processing.

The optimized experimental conditions have effectively improved separation efficiency and data comparison analysis. Furthermore, they established a solid foundation for its application development. CE-MS analysis of complex samples in the fields of metabolomics and proteomics (e. g., of tissues, cells, body fluids, etc.) could provide a visualization method for future clinical analysis. It contributes to the development of cancer pathological analysis, drug development, disease surveillance, etc. The characteristic analysis of small molecule metabolites and protein biomarkers directly reflects on enzymatic activity in the biological systems. It could be associated with the development of various diseases/complications. Omics analysis also has an important directive to disease detection and surveillance with obvious advantages in disease diagnosis, staged treatment, drug development, and patient treatment progress. CE-MS is useful in detecting complications and promoting personalized medicine. It provides technical support for future clinical developments.

In addition to a comprehensive review of the recent advances of CE-MS research, this paper also indicates the development directions of CE-MS. In order to avoid the problem of omics analysis and obtain the optimized analysis results, future analysis should be improved from the following three aspects:(ⅰ) The analysis conditions should be optimized concerning sample preparation methods and separation techniques. (ⅱ) The analytic techniques should be supported to adjust to capillary coating and interface technology. (ⅲ) New ideas should be developed in the fields of clinical research and statistical analysis.

Key words: capillary electrophoresis-mass spectrometry (CE-MS), technical improvement, metabolomics, proteomics, review

王芳, 王松, 丛海林, 于冰. 基于毛细管电泳-质谱联用技术的代谢/蛋白质组学分析[J]. 色谱, 2020, 38(9): 1013-1021.

WANG Fang, WANG Song, CONG Hailin, YU Bing. Analysis of metabolomics and proteomics based on capillary electrophoresis-mass spectrometry[J]. Chinese Journal of Chromatography, 2020, 38(9): 1013-1021.

图/表 6

图1 不同锥型接口所得的紫外(UV)与电喷雾电离(ESI)对比图

Fig. 1 Comparison of ultraviolet (UV) and electrospray ionization (ESI) from different cone interfaces Reproduced from Ref. [22] with permission.

图2 (a) 无鞘CE-ESI-MS界面示意图与(b) CE-MS在不同界面下对代谢产物的分析

Fig. 2 (a) Schematic of the sheathless CE-ESI-MS interface and (b) analysis of m/z of metabolites by CE-MS at different interfaces For Fig. 2a: 1. plastic plate; 2. electrodialysis membrane; 3. separation capillaries; 4. platinum electrodes; 5. buffer reservoirs. For Fig. 2b: black line, unsheathed; grey line, sheath flow. Reproduced from Ref. [30] with permission.

图3 一组标准化合物在不同分析平台上的迁移时间, 迁移电泳率转化对比

Fig. 3 Comparison of migration time and migration electrophoresis rate of a set of standard compounds on different analysis platforms Reproduced from Ref. [34] with permission.

图4 微探针取样与CE-ESI-MS联用对双离子代谢物进行分析

Fig. 4 Microprobe sampling combined with CE-ESI-MS for analysis of dual ion metabolites Reproduced from Ref. [47] with permission.

图5 (a) 阳离子模式下的CE-MS酸性化合物代谢谱分析与(b)样品预浓缩处理后核苷酸谱图

Fig. 5 (a) Metabolic analysis of CE-MS acidic compounds in cation mode and (b) nucleotide profile after sample preconcentration Reproduced from Ref. [49] with permission.

图6 用纳米LC-CZE-MS分析蛋白质混合物

Fig. 6 Analysis of protein mix by nano-LC-CZE-MS The mix contained RNase B (100 μg/mL), cytochrome c (80 μg/mL), lysozyme c (80 μg/mL), and myoglobin (50 μg/mL). First dimension (a): nano-LC chromatogram (214 nm detection wavelength) of the separation of different proteins. Second dimension (b): CZE-MS electropherogram obtained after a heart-cut of RNase B peak (transfer volume: 20 nL). Reproduced from Ref. [55] with permission.

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