Capillary electrophoresis-inductively coupled plasma-mass spectrometry: a report on technical principles and problem solutions, potential, and limitations of this technology as well as on examples of application

This paper summarizes some basic principles of capillary electrophoresis (CE), inductively coupled plasma-mass spectrometry (ICP-MS), and coupling of both devices. Especially the interfacing is described in detail. A special focus is drawn to various interface developments reported in literature and technical problems, i.e., requirements to the interface setup and respective solutions. Nowadays, typically sheath flow-based interfaces are used. The sheath flow fulfills two requirements of hyphenation, (i) the closing of the electrical circuit of CE and (ii) the feeding of the used nebulizer with an adequate flow rate. In the beginning of CE-ICP-MS coupling predominantly home-made interface-nebulizer constructions were developed and tested for various speciation problems. Now increasingly such laboratory-constructed interfaces are left. Mostly commercial nebulizers are employed being combined with commercially available tee or cross fittings to connect the CE capillary to the electrode, the additional sheath flow, and the nebulizer. Due to the low sample amounts and low flow rates from CE, such nebulizers are typically low-flow nebulizers like, e.g., the microconcentric nebulizer (MCN) and the direct injection nebulizer (DIN). However, there are also reports on couplings using standard Meinhard systems. Still the control and reduction of a siphoning sucting flow and sufficient detection limits are the major problems in hyphenating CE to ICP-MS. Different solutions are reported on these problems and summarized here. Finally numerous applications are reported. Mostly, applications are performed on speciation of selenium, arsenic, metallothionein isoforms, mercury, or cobalt.     

Capillary electrophoresis-inductively coupled plasma-mass spectrometry: an attractive complementary technique for elemental speciation analysis

Some basic and practical aspects of interfacing capillary electrophoresis to inductively coupled plasma-mass spectrometry (CE-ICP-MS) are reviewed in this article with emphasis on the use of this hyphenated technique for elemental speciation analysis. The principles behind the techniques of both CE and ICP-MS are introduced. The interfacing of CE to ICP-MS is discussed including several devices and nebulizers reported in literature. A brief account of their advantages and limitations is given. The various CE-ICP-MS applications for elemental speciation analysis are also reviewed. Some issues concerning the future of CE-ICP-MS for the elemental speciation analyses are discussed.     

Recent advances in capillary electrophoresis‐mass spectrometry: Instrumentation,methodology and applications

Capillary electrophoresis (CE) offers fast and high‐resolution separation of charged analytes from small injection volumes. Coupled to mass spectrometry (MS), it represents a powerful analytical technique providing (exact) mass information and enables molecular characterization based on fragmentation. Although hyphenation of CE and MS is not straightforward, much emphasis has been placed on enabling efficient ionization and user‐friendly coupling. Though several interfaces are now commercially available, research on more efficient and robust interfacing with nano‐electrospray ionization (ESI), matrix‐assisted laser desorption/ionization (MALDI) and inductively coupled plasma mass spectrometry (ICP) continues with considerable results. At the same time, CE‐MS has been used in many fields, predominantly for the analysis of proteins, peptides and metabolites. This review belongs to a series of regularly published articles, summarizing 248 articles covering the time between June 2016 and May 2018. Latest developments on hyphenation of CE with MS as well as instrumental developments such as two‐dimensional separation systems with MS detection are mentioned. Furthermore, applications of various CE‐modes including capillary zone electrophoresis (CZE), nonaqueous capillary electrophoresis (NACE), capillary gel electrophoresis (CGE) and capillary isoelectric focusing (CIEF) coupled to MS in biological, pharmaceutical and environmental research are summarized.     

An overview of LC–MS interfacing systems with selected applications

Advantages and limitations are described for the different LC–MS interfacing systems (moving belt; direct liquid introduction; thermospray; atmospheric pressure ionization with heated pneumatic nebulizer, electrospray, or high flow ion spray; particle beam; and continuous flow fast atom bombardment). Some comments are also made about interfacing capillary zone electrophoresis (CZE). The peculiarities of the various interfaces are described, as are liquid chromatographic requirements prior to mass spectrometry using the different ionization techniques. Selected biological and environmental applications are given.     

Capillary electrophoresis mass spectrometry and its application to the analysis of biological mixtures

Capillary electrophoresis (CE) mass spectrometry (MS), with its ability to separate compounds present in extremely small volume samples rapidly, with high separation efficiency, and with compound identification capability based on molecular weight, is an extremely valuable analytical technique for the analysis of complex biological mixtures. The highest sensitivities and separation efficiencies are usually achieved by using narrow capillaries (5-50 micro m i.d.) and by using sheathless CE-to-MS interfaces. The difficulties in CE-to-MS interfacing and the limited loadability of these narrow columns, however, have prevented CE-MS from becoming a widely used analytical technique. To remedy these limitations, several CE-MS interfacing techniques have recently been introduced. While electrospray ionization is the most commonly used ionization technique for interfacing CE-to-MS, matrix assisted laser desorption ionization has also been used, using both on-line and off-line techniques. Moreover, the high concentration detection limit of CE has been addressed by development of several sample concentration and sample focusing methods. In addition, a wide variety of techniques such as capillary zone electrophoresis, capillary isoelectric focusing, and on-column transient isotachophoresis have now been interfaced to MS. These advances have resulted in a rapid increase in the use of CE-MS in the analysis of complex biological mixtures. CE-MS has now been successfully applied to the analysis of a wide variety of compounds including amino acids, protein digests, protein mixtures, single cells, oligonucleotides, and various small molecules relevant to the pharmaceutical industry.     

Coupling of a large-size capillary column with an electrospray mass spectrometer. A reliable and sensitive sheath flow capillary electrophoresis-mass spectrometry interface

The concept of interfacing a large-size column for capillary electrophoresis (CE) to electrospray ionization mass spectrometry (ESI-MS) for robust and automatic CE-MS operation is reported. Both standard ionspray interface and microionspray interface have been modified to operate in a sheath flow pattern to overcome the common stability problem in CE-MS coupling. To make the interface sensitive, a step-down stainless steel tube with smaller inner diameter and tapered tip was incorporated onto a larger tube embracing the CE column via cold soldering. The devices were evaluated for quantitative analysis of nucleotides at femtomole level and stable analytical performance in peptide profiling.     

Capillary electrophoresis-mass spectrometry for the analysis of intact proteins

Developments in the fields of protein chemistry, proteomics and biotechnology have increased the demand for suitable analytical techniques for the analysis of intact proteins. In 1989, capillary electrophoresis (CE) was combined with mass spectrometry (MS) for the first time and its potential usefulness for the analysis of intact (i.e. non-digested) proteins was shown. This article provides an overview of the applications of CE-MS within the field of intact protein analysis. The principles of the applied CE modes and ionization techniques used for CE-MS of intact proteins are shortly described. It is shown that separations are predominantly carried out by capillary zone electrophoresis and capillary isoelectric focusing, whereas electrospray ionization (ESI) and matrix-assisted laser desorption ionization (MALDI) are the most popular ionization techniques used for interfacing. The combination of CE with inductively coupled plasma (ICP) MS for the analysis of metalloproteins is also discussed. The various CE-MS combinations are systematically outlined and tables provide extensive overviews of the applications of each technique for intact protein analysis. Selected examples are given to illustrate the usefulness of the CE-MS techniques. Examples include protein isoform assignment, single cell analysis, metalloprotein characterization, proteomics and biomarker screening. Finally, chip-based electrophoresis combined with MS is shortly treated and some of its applications are described. It is concluded that CE-MS represents a powerful tool for the analysis of intact proteins yielding unique separations and information.     

Hydrogen/deuterium exchange using a coaxial sheath-flow interface for capillary electrophoresis/mass spectrometry

The interfacing of capillary electrophoresis (CE) with mass spectrometry (MS) is well established and may be accomplished by use of either a coaxial arrangement or by employing a liquid T-junction. In both these interfaces a make-up flow is introduced. This is required because of the mismatch in flow rates for capillary electrophoresis approximately nL/min and 'true' electrospray approximately 2-10 microL/min. Electrical connectivity may also be established where the liquid flows meet (the introduction of nanospray renders the use of make-up flow unnecessary). Hydrogen/deuterium (H/D) exchange occurs in solution when there are labile hydrogen atoms present in a molecule. The establishment of the presence and the number of such exchangeable hydrogen atoms may be of importance in the identification and differentiation of compounds. It may also be an aid in the structural elucidation of unknown materials. We have investigated the feasibility of carrying out H/D exchange via a CE/MS interface. This involved the addition of D2O to the sheath flow and our preliminary results showing the separations of drug substances, subsequently undergoing exchange, are presented.     

Cover Picture: Electrophoresis 21'2010

Issue no. 21 is a regular issue with "Emphasis on Nucleic Acids" and comprises 17 contributions distributed over 4 distinct parts. Part I is on nucleic acids and has 8 research articles on various aspects of nucleic acid research including an automated instrument for human STR identification, a high resolution chip‐CE assay for rapid detection of gene mutations and amplifications, a fluorescence‐based sequence‐specific primer PCR, pulsed field CE for RNA separation, methylated DNA immunoprecipitation sequencing, PCR‐SSCP, DNA methylation, and analysis of SNPs. Part II has 4 research articles dealing with studies on protein and proteomics analysis, e.g., microchip IEF, CE analysis of microheterogeneous glutelin subunits in rice, protein precipitation methods and prefractionation of plasma for proteomic analysis. Part III has 2 contributions that describe the separation of the enantiomers of DL‐isocitric acid by CE and the analysis of trace degradation products of S‐adenosylmethionine, which are decarboxylated diastereomers, by CE. The last 3 articles in this issue (Part IV) are on novel methodologies for facile interfacing of CE with MS, electrokinetic chromatography for the determination of contaminants and impurities of heparin samples, and particle electrophoresis. As such, issue no. 21 is packed with the latest developments and innovations in the field. Featured articles include: An automated instrument for Human STR identification: design, characterization, and experimental validation (doi: 10.1002/elps.201000305 )) Miniature flowing atmospheric‐pressure afterglow ion source for facile interfacing of capillary electrophoresis with mass spectrometry (doi: 10.1002/elps.201000350 )) Capillary electrophoresis for analysis of microheterogeneous glutelin subunits in rice (Oryza sativa L.). (doi: 10.1002/elps.201000333 ))     

Capillary electrophoresis and electrochromatography of pesticides and metabolites

Synthetic pesticides are important chemicals since they are widely used to control many types of weeds, insects and other pests in a wide variety of agricultural and nonagricultural settings. This review article is aimed at describing the recent progress made in capillary electrophoresis (CE) and capillary electrochromatography (CEC) of pesticides and their metabolites. The various electrophoretic systems and detection schemes that have been introduced so far for the CE and CEC of pesticides are discussed. Also included in this review article are the various approaches for trace enrichment that are involved in the analysis of dilute pesticide samples.     

CE determination of small ions: methods and techniques

This paper provides an overview on the current status of capillary electrophoresis (CE) in the analysis of inorganic and charged small organic species. The various CE strategies used to improve the separation of ionic analytes are summarized. Technical developments in the design of improved detection systems are described. A brief account of their advantages and limitations is given. The potential use of these devices for miniaturized CE systems is also described. Finally, special attention is focused on the on-capillary preconcentration techniques developed in attempts to overcome the poor detectability of CE. Recent review articles are frequently cited to provide readers with a source of information about pioneering work, theoretical treatments, and specific applications.     

Recent developments in electrokinetically driven analysis on microfabricated devices

This review is devoted to the rapid developments in the field of microfluidic separation devices in which the flow is electrokinetically driven, and where the separation element forms the heart of the system, in order to give an overview of the trends of the last three years. Examples of microchip layouts that were designed for various application areas are given. Optimization of mixing and injection strategies, designs for the handling of multiple samples, and capillary array systems show the enormous progress made since the first proof-of-concept papers about lab-on-a-chip devices. Examples of functional elements for on-chip preconcentration, filtering, DNA amplification and on-chip detection indicate that the real integration of various analytical tasks on a single microchip is coming into reach. The use of materials other than glass, such as poly(dimethylsiloxane) and polymethylmethacrylate, for chip fabrication and detection methods other than laser-induced fluorescence (LIF) detection, such as mass spectrometry and electrochemical detection, are described. Furthermore, it can be observed that the separation modes known from capillary electrophoresis (CE) in fused-silica capillaries can be easily transferred to the microchip platform. The review concludes with an overview of applications of microchip CE and with a brief outlook.     

Capillary electrophoresis and electrochromatography of pesticides and metabolites

Synthetic pesticides are important chemicals since they are widely used to control many types of weeds, insects, and other pests in a wide variety of agricultural and nonagricultural settings. This review article is aimed at describing the recent progress made in capillary electrophoresis (CE) and capillary electrochromatography (CEC) of pesticides and metabolites. The various electrophoretic systems and detection schemes that were introduced during the period extending from the second half of 1999 to the first half of 2001 for the CE and CEC of pesticides are discussed. Also included in this review article are the various approaches for trace enrichment that are involved in the analysis of dilute pesticide samples.     

Electrophoretic separations on chips with hydrodynamically closed separation systems

This review focuses on capillary electrophoretic separations performed on capillary electrophoresis chips (CE chips) with hydrodynamically closed separation systems in a context with transport processes (electroosmotic flow (EOF)) and hydrodynamic flow (HDF)) that may accompany the separations in these devices. It also reflects some relevant works dealing with conventional CE operating under such hydrodynamic conditions. The use of zone electrophoresis (ZE), isotachophoresis (ITP) and their on-line combination (ITP-ZE) on the single-column and column-coupling CE chips with the closed separation systems and related problems are key topics of the review. Some attention is paid to sample pretreatment in the separations performed on the CE chips. Here, mainly potentialities of the ITP-ZE combination in trace analysis applications of the miniaturized systems are discussed in a broader extent. Links between the ZE separation and detection provide a frame for the discussion of current status of the detection on the CE chips. Analytical applications illustrate potentialities of the CE chips operating with the closed separation systems (suppressed HDF and EOF) to the determination of small ions present in various matrices by ZE, ITP and ITP-ZE.     

Flow/sequential injection sample treatment coupled to capillary electrophoresis. A review

Various flow sample handling approaches coupled to capillary electrophoresis (CE) are reviewed, covering the research in this field in the 12 years since the milestone year of 1997, when practical interfaces to on-line couple flow injection (FI) and capillary electrophoresis were first developed independently by two research groups. Some previous attempts are also presented. Since 1997 a plethora of ingenious coupled systems have been developed. Although several reviews are available on various aspects of the topic, we have opted for a comprehensive overview of all FI-CE systems, as well as related and similar systems. This coupling has thus also led to the development of systems based on hybrids between the classical and microchip approaches. Truly microchip FI-CE systems are also included in this review.The developed systems have been used for various sample treatments, including on-line membrane-assisted sample treatment, column-based preconcentration, on-line derivatization and monitoring, to name just a few. The utility of coupling flow sampling to CE has been demonstrated in various practical applications that are discussed in detail. The current state-of-the-art and foreseeable future developments are also discussed.     

Recent developments in the separation of inorganic and small organic ions by capillary electrophoresis

Which method should I use for ion analysis, ion chromatography (IC) or capillary electrophoresis (CE)? In terms of actual theoretical plates CE has a clear-cut advantage. The separation ability of IC is adequate for many sample types, and many separation scientists feel that IC offers greater reliability and confidence than CE. However, IC is a more mature technique and there has been more time to solve problems such as peak tailing and to improve reproducibility. The two techniques should be viewed as complementary. A number of recent developments in ion analysis by CE are discussed. These include some simple ways to control electroosmotic flow and improve reproducibility, separation of isotopes, improved methods of indirect photometric detection, a new contactless conductivity detector, separation of ions at low pH, and in solutions of high salt content. Progress in a new technique called IC-CE will be described in which a soluble ion-exchange polymer is added to the capillary electrolyte to separate anions based on differences in both electrophoretic mobility and ion-exchange interactions.     

Effects of nebulizing and drying gas flow on capillary electrophoresis/mass spectrometry

This study was focused on examining the influence of gas flow parameters on capillary electrophoresis/mass spectrometry (CE /MS) performance using sheath-liquid CE /MS interfaces. The effects of nebulizing and drying gas velocity and drying gas temperature on CE separation and MS detection sensitivity were systematically determined. Nebulizing gas velocity was observed to be a critical parameter in the optimization of CE /MS method, since it affected both MS detection sensitivity, and also CE separation efficiency for one interface design tested. Better detection sensitivity was obtained when the nebulizing gas velocity was increased. However, high velocity of the nebulizing gas flow can cause a hydrodynamic bulk flow inside the CE capillary, thus clearly increasing the apparent mobility and decreasing the resolution obtained for the compounds studied. Increasing the drying gas velocity or temperature did not affect the apparent mobility or the separation efficiency and the temperature could be increased to achieve the optimal detection sensitivity in the CE /MS analysis. For comparison, the effects of nebulizing gas flow were studied using a different design of the coaxial sheath-liquid CE /MS interface, and in this case better detection sensitivity but no effect on CE separation efficiency was observed with increased nebulizing gas velocity. These different effects of nebulizing gas flow on the CE bulk flow were concluded to result from pressure differences at the tip of the CE capillaries for the different CE /MS interface arrangements. It is therefore recommended that the cross-sectional dimensions of the fused-silica and steel capillaries, and the gas streamlines, should be optimized when CE /MS interfaces are built. Moreover, the effect of gas flow on CE separation should be studied when optimizing the CE /MS operation parameters.     

A flow injection-capillary electrophoresis system with high-voltage contactless conductivity detection for automated dual opposite end injection

The system comprises two flow injection-capillary electrophoresis interfaces into which the opposite ends of the separation capillary are inserted. The electrolyte solution flows through both interfaces by use of hydrostatic pressure. The injection of the samples into the electrolyte flow is accomplished by a rotary-type chromatographic valve at the grounded side and by a pinch-valve injector at the high-voltage side that provides sufficient isolation from the high electric field. The system allows a fully automated dual-injection sequence of samples from both capillary ends and simultaneous electrophoretic separation of anions and cations in the samples. The analytes are detected by a high-voltage contactless conductometric detector positioned approximately in the middle of the separation capillary. The parameters of the system were evaluated. The repeatability of the flow injection-capillary electrophoresis system for the simultaneous determination of anions and cations was evaluated for ten consecutive injections and relative standard deviation (RSD) values for peak areas were better than 1.0%. The sample throughput for total ionic analysis was estimated to be 25 samples per hour. The system was used for automated simultaneous analysis of anions and cations in various real samples. Using a short separation capillary, rapid total ionic analysis in less then 1 min is demonstrated.     

Separation of natural and synthetic heparin fragments by high-performance capillary electrophoresis.

The application of capillary electrophoresis (CE) for the analysis of natural and synthetic low-molecular-mass heparin fragments at low pH is described. It is demonstrated that under the applied conditions the separation is based on charge, charge distribution and molecular mass of the heparin molecules, yielding a high resolution. It is shown that the presence of sodium chloride in the sample solution has hardly any effect on the CE performance. However, the pH of the electrophoresis buffer is a critical parameter. The resolutions obtained with CE and high-performance anion-exchange chromatography (HPAEC) are compared for various heparin fragments and it is concluded that, at least for this type of molecule, CE forms an attractive alternative to HPAEC.