LABEL‐FREE BIO‐AFFINITY MASS SPECTROMETRY FOR SCREENING AND LOCATING BIOACTIVE MOLECULES

Despite the recent increase in the development of bioactive molecules in the drug industry, the enormous chemical space and lack of productivity are still important issues. Additional alternative approaches to screen and locate bioactive molecules are urgently needed. Label‐free bio‐affinity mass spectrometry (BA‐MS) provides opportunities for the discovery and development of innovative drugs. This review provides a comprehensive portrayal of BA‐MS techniques and of their applications in screening and locating bioactive molecules. After introducing the basic principles, alongside some application notes, the current state‐of‐the‐art of BA‐MS‐assisted drug discovery is discussed, including native MS, size‐exclusion chromatography‐MS, ultrafiltration‐MS, solid‐phase micro‐extraction‐MS, and cell membrane chromatography‐MS. Finally, several challenges and limitations of the current methods are summarized, with a view to potential future directions for BA‐MS‐assisted drug discovery. © 2019 John Wiley & Sons Ltd. Mass Spec Rev     

Neuropeptides in the crayfish stomatogastric nervous system

Neuropeptides are peptides with profound effects on the nervous system. The function of neuropeptides can be studied in detail in the stomatogastric nervous system (STNS). Neuropeptides are ubiquitously distributed in the STNS and it contains well-studied neural circuits that are strongly modulated by neuropeptides. The STNS controls the movements of the foregut in crustaceans and has been studied intensively in a variety of decapod crustaceans including crayfish. This article reviews our knowledge of neuropeptides in the crayfish STNS. Within crayfish, peptides reach the circuits of the STNS as neurohormones released by neurohaemal organs or by putative neurohemal zones located within the STNS. As transmitters, neuropeptides are present in identified motoneurons, interneurons, and sensory neurons (mainly shown by immunocytochemistry), indicating a multiple role of peptides in the plasticity of neural networks. Neuropeptides are not only present in varicosities within the neuropil of ganglia, but also in varicosities on muscles and within small neuropil patches along nerves. This suggests that the muscles of the stomach are under a more direct modulatory control than previously thought, and that information processing can also occur within nerves. In addition to anatomical studies, biochemical and electrophysiological methods were used. For example, MALDI-TOF MS (matrix-assisted laser desorption ionization time of flight mass spectrometry) revealed the presence of four different peptides of the orcokinin family within a single neuron, and electrophysiological experiments demonstrated that the networks of the STNS are not only under excitatory but also inhibitory peptidergic influence. Comparing the similarities and differences between the STNS of crayfish and that of other decapod crustaceans has already contributed to our knowledge about peptides and will further help to unravel peptide function in the plasticity of neural circuits. For example, the identified neurons in the STNS can be used to study co-transmission because neuropeptides are co-localized with classical transmitters, biogenic amines, or other peptides in these neurons.     

Tandem mass spectrometry in the study of fatty acids,bile acids,and steroids

Over the last 50 years, the mass spectrometry of lipids has evolved to become one of the most mature techniques in biomolecule analysis. Many volatile and non-polar lipids are directly amenable to analysis by gas-chromatography-mass spectrometry (GC-MS), a technique that combines the unsurpassed separation properties of gas-chromatography with the sensitivity and selectivity of electron ionization mass spectrometry. Less volatile and/or thermally labile lipids can be analyzed by GC-MS, following appropriate sample derivatization. However, many complex lipids are not readily analyzed by GC-MS, and it is these molecules that are the subject of the current review. Since the early 1970s, there have been three outstanding developments in mass spectrometry that are particularly appropriate in lipid analysis; i.e., the introduction of (i) fast atom bombardment (FAB); (ii) electrospray (ES); and (iii) tandem mass spectrometry (MS/MS). The FAB and ES ionization techniques will be discussed in relation to MS/MS, and examples of their application in biochemical studies will be presented. The review will concentrate on the analysis of fatty acids, bile acids, steroid conjugates, and neutral steroids.     

Direct-EI in LC-MS: towards a universal detector for small-molecule applications

This review article will give an up-to-date and exhaustive overview on the efficient use of electron ionization (EI) to couple liquid chromatography and mass spectrometry (LC-MS) with an innovative interface called Direct-EI. EI is based on the gas-phase ionization of the analytes, and it is suitable for many applications in a wide range of LC-amenable compounds. In addition, thanks to its operating principles, it prevents unwelcome matrix effects (ME). In fact, although atmospheric pressure ionization (API) methodologies have boosted the use of LC-MS, the related analytical methods are sometime affected by inaccurate quantitative results, due to unavoidable and unpredictable ME. In addition, API's soft ionization spectra always demand for costly and complex tandem mass spectrometry (MS/MS) instruments, which are essential to acquire an "information-rich" spectrum and to obtain accurate quantitative information. In EI a one-stage analyzer is sufficient for a qualitative investigation and MS/MS detection is only used to improve sensitivity and to cut chemical noise. The technology illustrated here provides a robust and straightforward access to classical, well-characterized EI data for a variety of LC applications, and readily interpretable spectra for a wide range of areas of research. The Direct-EI interface can represent the basis for a forthcoming universal LC-MS detector for small molecules.     

Analytical strategies for identifying drug metabolites

With the dramatic increase in the number of new chemical entities (NCEs) arising from combinatorial chemistry and modern high-throughput bioassays, novel bioanalytical techniques are required for the rapid determination of the metabolic stability and metabolites of these NCEs. Knowledge of the metabolic site(s) of the NCEs in early drug discovery is essential for selecting compounds with favorable pharmacokinetic credentials and aiding medicinal chemists in modifying metabolic "soft spots". In development, elucidation of biotransformation pathways of a drug candidate by identifying its circulatory and excretory metabolites is vitally important to understand its physiological effects. Mass spectrometry (MS) and nuclear magnetic resonance (NMR) have played an invaluable role in the structural characterization and quantification of drug metabolites. Indeed, liquid chromatography (LC) coupled with atmospheric pressure ionization (API) MS has now become the most powerful tool for the rapid detection, structure elucidation, and quantification of drug-derived material within various biological fluids. Often, however, MS alone is insufficient to identify the exact position of oxidation, to differentiate isomers, or to provide the precise structure of unusual and/or unstable metabolites. In addition, an excess of endogenous material in biological samples often suppress the ionization of drug-related material complicating metabolite identification by MS. In these cases, multiple analytical and wet chemistry techniques, such as LC-NMR, enzymatic hydrolysis, chemical derivatization, and hydrogen/deuterium-exchange (H/D-exchange) combined with MS are used to characterize the novel and isomeric metabolites of drug candidates. This review describes sample preparation and introduction strategies to minimize ion suppression by biological matrices for metabolite identification studies, the application of various LC-tandem MS (LC-MS/MS) techniques for the rapid quantification and identification of drug metabolites, and future trends in this field.     

Mass spectrometry imaging under ambient conditions

Mass spectrometry imaging (MSI) has emerged as an important tool in the last decade and it is beginning to show potential to provide new information in many fields owing to its unique ability to acquire molecularly specific images and to provide multiplexed information, without the need for labeling or staining. In MSI, the chemical identity of molecules present on a surface is investigated as a function of spatial distribution. In addition to now standard methods involving MSI in vacuum, recently developed ambient ionization techniques allow MSI to be performed under atmospheric pressure on untreated samples outside the mass spectrometer. Here we review recent developments and applications of MSI emphasizing the ambient ionization techniques of desorption electrospray ionization (DESI), laser ablation electrospray ionization (LAESI), probe electrospray ionization (PESI), desorption atmospheric pressure photoionization (DAPPI), femtosecond laser desorption ionization (fs‐LDI), laser electrospray mass spectrometry (LEMS), infrared laser ablation metastable‐induced chemical ionization (IR‐LAMICI), liquid microjunction surface sampling probe mass spectrometry (LMJ‐SSP MS), nanospray desorption electrospray ionization (nano‐DESI), and plasma sources such as the low temperature plasma (LTP) probe and laser ablation coupled to flowing atmospheric‐pressure afterglow (LA‐FAPA). Included are discussions of some of the features of ambient MSI for example the ability to implement chemical reactions with the goal of providing high abundance ions characteristic of specific compounds of interest and the use of tandem mass spectrometry to either map the distribution of targeted molecules with high specificity or to provide additional MS information on the structural identification of compounds. We also describe the role of bioinformatics in acquiring and interpreting the chemical and spatial information obtained through MSI, especially in biological applications for tissue diagnostic purposes. Finally, we discuss the challenges in ambient MSI and include perspectives on the future of the field. © 2012 Wiley Periodicals, Inc., Mass Spec Rev 32:218–243, 2013     

Laser-BasedAmbientIonizationMassSpectrometry

Electrospray-assisted laser desorption ionization (ELDI) and laser-induced acoustic desorption (LIAD) mass spectrometry (MS) are useful techniques for directly characterizing small chemical or large biological components in solid or liquid under ambient conditions. Sample pretreatment is usually unnecessary for both techniques. The analytes molecules in the samples are desorbed by the action of a pulsed laser without the assistance of organic matrix. The desorbed molecules are subsequently ionized in an electrospray plume generated by electrospraying an acidic methanol solution. Recently, we have modified the existing ELDI and LIAD sources so unique applications are performed: 1) High throughput TLC/MS analysis - the TLC plate was linearly scanned by the laser from either side of the plate. Typically the systems allow to continuously screening up to 400 TLC plates per day. 2) Molecular imaging analysis - with the assistance of a stepper motor for precisely moving the sample plate and a high spatial resolution laser, the distribution of the predominant chemical compounds on a particular sample surface were obtained. The molecular images based on volatile and non-volatile chemical compounds on dry fungus and plant slices were obtained. 3) Monitoring chemical reaction - the states of ongoing chemical reactions－nanoparticle catalyzed photodecomposition of dye molecules in different organic solvents, were continuously monitored. 4) Protein structure study - the change of protein structure was studied via H/D and D/H exchange. The amino acids, peptides, and proteins with different exchangeable H atoms were chosen to ascertain the number of H/D or D/H exchanges in a reactive-ELDI/MS system. In addition, a new configuration of the electrospray system was built for both ELDI and LIAD sources. A multiple electrosprayer system was used to generate an atmosphere containing homogenously charged species. The efficiency of post ionization of the desorbed analyte molecules in the new ion sources is greatly increased. Different solvent systems can be simultaneously used to produce electrospray plumes to ionize the analyte molecules with different polarities.     

Mass spectrometric methods for monitoring redox processes in electrochemical cells

Electrochemistry (EC) is a mature scientific discipline aimed to study the movement of electrons in an oxidation–reduction reaction. EC covers techniques that use a measurement of potential, charge, or current to determine the concentration or the chemical reactivity of analytes. The electrical signal is directly converted into chemical information. For in‐depth characterization of complex electrochemical reactions involving the formation of diverse intermediates, products and byproducts, EC is usually combined with other analytical techniques, and particularly the hyphenation of EC with mass spectrometry (MS) has found broad applicability. The analysis of gases and volatile intermediates and products formed at electrode surfaces is enabled by differential electrochemical mass spectrometry (DEMS). In DEMS an electrochemical cell is sampled with a membrane interface for electron ionization (EI)‐MS. The chemical space amenable to EC/MS (i.e., bioorganic molecules including proteins, peptides, nucleic acids, and drugs) was significantly increased by employing electrospray ionization (ESI)‐MS. In the simplest setup, the EC of the ESI process is used to analytical advantage. A limitation of this approach is, however, its inability to precisely control the electrochemical potential at the emitter electrode. Thus, particularly for studying mechanistic aspects of electrochemical processes, the hyphenation of discrete electrochemical cells with ESI‐MS was found to be more appropriate. The analytical power of EC/ESI‐MS can further be increased by integrating liquid chromatography (LC) as an additional dimension of separation. Chromatographic separation was found to be particularly useful to reduce the complexity of the sample submitted either to the EC cell or to ESI‐MS. Thus, both EC/LC/ESI‐MS and LC/EC/ESI‐MS are common. © 2013 The Authors. Mass Spectrometry Reviews published by Wiley Periodicals, Inc. Mass Spec Rev 34:64–92, 2015     

基于纳米毛细管的非变性质谱分析

非变性质谱利用电喷雾电离质谱技术从非变性缓冲溶液中研究自然折叠状态下的蛋白质、核酸及其复合物,是大型蛋白质组装体、蛋白质-药物复合物、核酸-配体复合物的结构、组成及其相互作用解析的重要工具。生化缓冲溶液中的非挥发性盐能提供足够的离子强度和适宜的pH值,以维持蛋白质、核酸分子折叠和结合状态,但通常与电喷雾电离质谱不兼容。基于纳米毛细管的电喷雾电离具备强大的基质耐受能力,为直接从生理或近生理缓冲溶液中完成蛋白质及其复合物的高效离子化提供了可能,极大地促进了近年来非变性质谱的应用研究。本文综述了纳米毛细管的制备与表征方法及其尺寸和界面性质导致的电离行为特性,回顾了近10年来纳米毛细管在非变性质谱中的应用研究进展。     

自动富集-单光子电离质谱在线测量氯酚的研究

氯酚类化合物是垃圾焚烧所产生的二噁英前生体之一,对二噁英的浓度有很好的指示作用。针对气相中氯酚类二噁英前生体的在线连续监测,自行研制了全自动富集进样-单光子电离-反射式飞行时间质谱仪。为实现低浓度氯酚的检测,设计了全自动的富集进样系统,利用3根阵列式Tenax TA吸附管,结合全自动控制的高温电磁阀,实现了气体样品连续自动的“富集 热解析-质谱进样”。研制的反射式单光子电离-飞行时间质谱仪采用10.6 eV真空紫外灯作为光电离源,对气体样品分子进行软电离,避免了碎片离子的产生,所获得的样品分子离子经反射式飞行时间质量分析器完成检测,仪器分辨率达到3 000（m/z 78）。以3种典型的二噁英前生体氯酚类化合物（邻氯苯酚、2,4-二氯苯酚、2,4,6-三氯苯酚）为检测对象,对该仪器性能进行考察。富集进样系统对3种氯酚类化合物的富集倍数分别达到60、249、503,检测限分别达到66、120、125 pg/L,动态线性范围分别为166 pg/L~16.6 ng/L、210 pg/L~21 ng/L、255 pg/L~25.5 ng/L。这些研究结果为今后开展垃圾焚烧烟气中二噁英的现场连续监测工作奠定了基础。     

Biomedical application of MALDI mass spectrometry for small‐molecule analysis

Matrix‐assisted laser desorption/ionization (MALDI) mass spectrometry (MS) is an emerging analytical tool for the analysis of molecules with molar masses below 1,000 Da; that is, small molecules. This technique offers rapid analysis, high sensitivity, low sample consumption, a relative high tolerance towards salts and buffers, and the possibility to store sample on the target plate. The successful application of the technique is, however, hampered by low molecular weight (LMW) matrix‐derived interference signals and by poor reproducibility of signal intensities during quantitative analyses. In this review, we focus on the biomedical application of MALDI‐MS for the analysis of small molecules and discuss its favorable properties and its challenges as well as strategies to improve the performance of the technique. Furthermore, practical aspects and applications are presented. © 2010 Wiley Periodicals, Inc., Mass Spec Rev 30:101–120, 2011     

Electrospray ionization mass spectrometry of oligonucleotide complexes with drugs, metals, and proteins.

I. Introduction 61 II. Binding of Small Molecules to DNA 62 A. Covalent Binding 62 B. Reversible (Noncovalent) DNA-Binding Agents 65 III. DNA-Metal Ion Complexes 67 A. Platinum Complexes 70 B. Other Metal Ions 73 IV. DNA-Protein Complexes 74 A. Introduction 74 B. ESI-MS of DNA-Protein Complexes 76 C. ESI-MS Analysis of Proteolytic Products of DNA-Protein Complexes 79 D. ESI-MS of Ternary DNA-Protein-Ligand Complexes 80 V. Conclusions 80 Abbreviations 81 References 81 --Interactions of DNA with drugs, metal ions, and proteins are important in a wide variety of biological processes. With the advent of electrospray ionization (ESI) and matrix-assisted laser desorption ionization (MALDI), mass spectrometry (MS) is now a well-established tool for the characterization of the primary structures of biopolymers. The gentle nature of the ESI process, however, means that ESI-MS is also finding application for the study of noncovalent and other fragile biomolecular complexes. We outline here the progress, to date, in the use of ESI-MS for the study of noncovalent drug-DNA and protein-DNA complexes together with strategies that can be employed to examine the binding of small molecules and metal complexes to DNA. In the case of covalent complexes with DNA, sequence information can be derived from ESI-MS used in conjunction with tandem mass spectrometry (MS/MS) and/or enzymatic digestion. MS/MS can also be used to probe the relative binding affinities of drugs that bind to DNA via noncovalent interactions. Overall, the work in this area, to date has demonstrated that ESI-MS and MS/MS will prove to be valuable complements to other structural methods, offering advantages in terms of speed, specificity, and sensitivity. (c) 2001 John Wiley & Sons, Inc.     

Collisional activation of peptide ions in FT-ICR mass spectrometry

In the last decade, the characterization of complex molecules, particularly biomolecules, became a focus of fundamental and applied research in mass spectrometry. Most of these studies utilize tandem mass spectrometry (MS/MS) to obtain structural information for complex molecules. Tandem mass spectrometry (MS/MS) typically involves the mass selection of a primary ion, its activation by collision or photon excitation, unimolecular decay into fragment ions characteristic of the ion structure and its internal excitation, and mass analysis of the fragment ions. Although the fundamental principles of tandem mass spectrometry of relatively small molecules are fairly well-understood, our understanding of the activation and fragmentation of large molecules is much more primitive. For small ions, a single energetic collision is sufficient to dissociate the ion; however, this is not the case for complex molecules. For large ions, two fundamental limits severely constrain fragmentation in tandem mass spectrometry. First, the center-of-mass collision energy-the absolute upper limit of energy transfer in a collision process-decreases with increasing mass of the projectile ion for fixed ion kinetic energy and neutral mass. Secondly, the dramatic increase in density of states with increasing internal degrees of freedom of the ion decreases the rate of dissociation by many orders of magnitude at a given internal energy. Consequently, most practical MS/MS experiments with complex ions involve multiple-collision activation (MCA-CID), multi-photon activation, or surface-induced dissociation (SID). This review is focused on what has been learned in recent research studies concerned with fundamental aspects of MCA-CID and SID of model peptides, with an emphasis on experiments carried out with Fourier transform ion cyclotron resonance mass spectrometers (FT-ICR MS). These studies provide the first quantitative comparison of gas-phase multiple-collision activation and SID of peptide ions. Combining collisional energy-resolved data with RRKM-based modeling revealed the effect of peptide size and identity on energy transfer in collisions-very important characteristics of ion activation from fundamental and the analytical perspectives. Finally, the combination of FT-ICR with SID was utilized to carry out the first time-resolved experiments that examine the kinetics of peptide fragmentation. This has lead to the discovery that the time-dependence of ion dissociation varies smoothly up to a certain collision energy, and then shifts dramatically to a time-independent, extensive dissociation. This near-instantaneous "shattering" of the ion generates a large number of relatively small fragment ions. Shattering of ions on surfaces opens up a variety of dissociation pathways that are not accessible with multiple-collision and multiphoton excitation.     

基质辅助激光解吸电离质谱用于生物组织的质谱成像应用进展

基质辅助激光解吸电离质谱(MALDI MS)用于分析组织切片已成为质谱学的一个新领域。质谱成像技术通过直接对组织切片表面的质谱扫描,可以快速直观地分析组织中的分子,如蛋白质、多肽、药物分子、代谢产物等及其空间分布信息。本工作综述了组织切片的质谱成像原理,方法学和相关应用。     

印迹膜电喷雾电离质谱法快速分析尿液中沙丁胺醇

沙丁胺醇(SAL)可促进动物生长,常被非法添加于畜禽饲料中,并可通过食物链进入人体,危害人体健康。目前,沙丁胺醇的分析方法已有很多报道,但对于复杂生物样本中沙丁胺醇的快速定量分析依然存在挑战。为实现这一目标,本研究采用印迹膜电喷雾电离质谱法(MIM-ESI MS)对尿液中的沙丁胺醇进行直接定性和定量分析。结果表明,该方法具有较高的灵敏度,尿液样本的检出限和定量限分别为5 ng/L和10 ng/L,在0.01~10 000 μg/L浓度范围内具有较好的线性关系。该方法无需样品前处理,其分子印迹膜材料具有特异性富集的功能,可实现生物复杂样本中痕量目标物的快速检测。     

Phospholipids as cancer biomarkers: Mass spectrometry‐based analysis

Lipids, particularly phospholipids (PLs), are key components of cellular membrane. PLs play important and diverse roles in cells such as chemical‐energy storage, cellular signaling, cell membranes, and cell–cell interactions in tissues. All these cellular processes are pertinent to cells that undergo transformation, cancer progression, and metastasis. Thus, there is a strong possibility that some classes of PLs are expected to present in cancer cells and tissues in cellular physiology. The mass spectrometric soft‐ionization techniques, electrospray ionization (ESI), and matrix‐assisted laser desorption/ionization (MALDI) are well‐established in the proteomics field, have been used for lipidomic analysis in cancer research. This review focused on the applications of mass spectrometry (MS) mainly on ESI‐MS and MALDI‐MS in the structural characterization, molecular composition and key roles of various PLs present in cancer cells, tissues, blood, and urine, and on their importance for cancer‐related problems as well as challenges for development of novel PL‐based biomarkers. The profiling of PLs helps to rationalize their functions in biological systems, and will also provide diagnostic information to elucidate mechanisms behind the control of cancer, diabetes, and neurodegenerative diseases. The investigation of cellular PLs with MS methods suggests new insights on various cancer diseases and clinical applications in the drug discovery and development of biomarkers for various PL‐related different cancer diseases. PL profiling in tissues, cells and body fluids also reflect the general condition of the whole organism and can indicate the existence of cancer and other diseases. PL profiling with MS opens new prospects to assess alterations of PLs in cancer, screening specific biomarkers and provide a basis for the development of novel therapeutic strategies. © 2016 Wiley Periodicals, Inc. Mass Spec Rev 37:107‐138, 2018     

A tutorial in small molecule identification via electrospray ionization‐mass spectrometry: The practical art of structural elucidation

The identification of unknown molecules has been one of the cornerstone applications of mass spectrometry for decades. This tutorial reviews the basics of the interpretation of electrospray ionization‐based MS and MS/MS spectra in order to identify small‐molecule analytes (typically below 2000 Da). Most of what is discussed in this tutorial also applies to other atmospheric pressure ionization methods like atmospheric pressure chemical/photoionization. We focus primarily on the fundamental steps of MS‐based structural elucidation of individual unknown compounds, rather than describing strategies for large‐scale identification in complex samples. We critically discuss topics like the detection of protonated and deprotonated ions ([M + H]⁺ and [M ? H]^?) as well as other adduct ions, the determination of the molecular formula, and provide some basic rules on the interpretation of product ion spectra. Our tutorial focuses primarily on the fundamental steps of MS‐based structural elucidation of individual unknown compounds (eg, contaminants in chemical production, pharmacological alteration of drugs), rather than describing strategies for large‐scale identification in complex samples. This tutorial also discusses strategies to obtain useful orthogonal information (UV/Vis, H/D exchange, chemical derivatization, etc) and offers an overview of the different informatics tools and approaches that can be used for structural elucidation of small molecules. It is primarily intended for beginning mass spectrometrists and researchers from other mass spectrometry sub‐disciplines that want to get acquainted with structural elucidation are interested in some practical tips and tricks.

     

3-氨基喹啉基体在MALDI-TOF/MS分析中对环状多糖分子的特殊解吸电离作用

This paper reports the special desorption/ionization effect of matrix 3-aminoquinoline(3-AQ) and binary matrix 2,5-dihydroxybenzoic acid/3-aminoquinoline(DHB/3-AQ) on the cyclic polysaccharide molecules in MALDI-TOF/MS analysis.     

Ionization techniques in capillary electrophoresis-mass spectrometry: principles, design, and application

A major step forward in the development and application of capillary electrophoresis (CE) was its coupling to ESI-MS, first reported in 1987. More than two decades later, ESI has remained the principal ionization technique in CE-MS, but a number of other ionization techniques have also been implemented. In this review the state-of-the-art in the employment of soft ionization techniques for CE-MS is presented. First the fundamentals and general challenges of hyphenating conventional CE and microchip electrophoresis with MS are outlined. After elaborating on the characteristics and role of ESI, emphasis is put on alternative ionization techniques including sonic spray ionization (SSI), thermospray ionization (TSI), atmospheric pressure chemical ionization (APCI), atmospheric pressure photoionization (APPI), matrix-assisted laser desorption ionization (MALDI) and continuous-flow fast atom bombardment (CF-FAB). The principle of each ionization technique is outlined and the experimental set-ups of the CE-MS couplings are described. The strengths and limitations of each ionization technique with respect to CE-MS are discussed and the applicability of the various systems is illustrated by a number of typical examples.