毛细管电泳分析中的富集技术及其应用

毛细管电泳（CE）具有分析速度快、分离效率高、样品消耗少、成本低廉等优点,已被应用于无机离子、有机小分子、蛋白质、核酸及细胞等的分析中。CE中最常用的检测方式是紫外检测（UV）,但由于常规进样样品体积小、检测光程短,CE-UV的灵敏度往往不能满足复杂样品中痕量物质直接分析的要求。CE中的在柱富集技术包括堆积、动态pH界面、吹扫和瞬间等速电泳等,可在很大程度上提高CE-UV的检测灵敏度;另外,固相和液相微萃取技术及其与在柱富集技术相结合应用在CE中也能净化样品基质,进一步提高富集倍数,改善分析灵敏度,从而拓宽了CE-UV在复杂样品分析中的应用范围。     

Sweeping: concentration mechanism and applications to high-sensitivity analysis in capillary electrophoresis

Sweeping in capillary electrophoresis (CE) involves the interaction of a pseudostationary phase (PS) in the separation solution and a sample in the matrix that is free of the PS used. The PS includes not only the PSs employed in electrokinetic chromatography, but also complexation reagents such as borate. The sample matrix could have a lower, similar, or higher conductance than the separation solution. Thus, the basic condition for sweeping is a sample matrix free of the additive. The accumulation of analyte molecules during the interaction makes this interesting phenomenon very useful as an on-line preconcentration method for CE. Preconcentration occurs due to chromatographic partitioning, complexation, or any interaction between analytes and PS. Contact between analyte and PS is facilitated by the action of electrophoresis and is independent of electroosmosis. The analyte, PS, or both should have electrophoretic velocities when an electric field is applied. The extent of preconcentration is dictated by the strength of the interaction involved. From tens to several thousand-fold improvements in detector response for many neutral and charged analytes have been achieved with this technique, suggesting sweeping as a general approach to on-line preconcentration in CE. The mechanism and applications of the sweeping phenomenon under different experimental conditions are discussed in this review, with particular emphasis on a better understanding of the sweeping mechanism under reduced electric field (high conductivity) in the sample zone.     

Chiral separations by capillary electrophoresis: Recent developments and applications

This paper provides an overview of the different classes of chiral selectors that are used in CE. The main properties of every class are described, together with the mechanism of enantioseparation. Newly introduced selectors are also discussed. Pharmaceutical and biomedical applications published from January 2004 till March 2005 are summarized.     

Sixteen capillary electrophoresis applications for viral vaccine analysis

A broad range of CE applications from our organization is reviewed to give a flavor of the use of CE within the field of vaccine analyses. Applicability of CE for viral vaccine characterization, and release and stability testing of seasonal influenza virosomal vaccines, universal subunit influenza vaccines, Sabin inactivated polio vaccines (sIPV), and adenovirus vector vaccines were demonstrated. Diverse CZE, CE-SDS, CGE, and cIEF methods were developed, validated, and applied for virus, protein, posttranslational modifications, DNA, and excipient concentration determinations, as well as for the integrity and composition verifications, and identity testing (e.g., CZE for intact virus particles, CE-SDS application for hemagglutinin quantification and influenza strain identification, chloride or bromide determination in process samples). Results were supported by other methods such as RP-HPLC, dynamic light scattering (DLS), and zeta potential measurements. Overall, 16 CE methods are presented that were developed and applied, comprising six adenovirus methods, five viral protein methods, and methods for antibodies determination of glycans, host cell-DNA, excipient chloride, and process impurity bromide. These methods were applied to support in-process control, release, stability, process- and product characterization and development, and critical reagent testing. Thirteen methods were validated. Intact virus particles were analyzed at concentrations as low as 0.8 pmol/L. Overall, CE took viral vaccine testing beyond what was previously possible, improved process and product understanding, and, in total, safety, efficacy, and quality.     

Thirty years of capillary electrophoresis in food analysis laboratories: potential applications

CE has generated considerable interest in the research community since instruments were introduced by different trading companies in the 1990s. Nowadays, CE is popular due to its simplicity, speed, highly efficient separations and minimal solvent and reagent consumption; it can also be included as a useful technique in the nanotechnology field and it covers a wide range of specific applications in different fields (chemical, pharmaceutical, genetic, clinical, food and environmental). CE has been very well evaluated in research laboratories for several years, and different new approaches to improve sensitivity (one of the main drawbacks of CE) and robustness have been proposed. However, this technique is still not well accepted in routine laboratories for food analysis. Researching in data bases, it is easy to find several electrophoretic methods to determine different groups of analytes and sometimes they are compared in terms of sensitivity, selectivity, precision and applicability with other separation techniques. Although these papers frequently prove the potential of this methodology in spiked samples, it is not common to find a discussion of the well-known complexity of the matrices to extract analytes from the sample and/or to study the interferences in the target analytes. Summarizing, the majority of CE scientific papers focus primarily on the effects upon the separation of the analytes while ignoring their behavior if these analytes are presented in real samples.     

Pesticide analysis by capillary electrophoresis

In this work, a critical and updated revision of the current situation of the analysis of pesticides by Capillary Electrophoresis (CE) is presented. The review has been written in two main sections. The first one presents a thorough revision of the various offline and on-line sample preconcentration procedures that have been used in conjunction with CE to analyze these compounds. The second part reviews the various detection strategies (i.e., UV, LIF, MS, and electrochemical) and CE modes that have been applied to the analysis of pesticides. Future trends that can be expected from this hot research area are also discussed.     

Dual injection capillary electrophoresis: foundations and applications

The state of the art of capillary electrophoresis (CE) approaches based on dual injection is here reported. Dual injection strategies have been proposed with three main objectives: (i) to provide information about reaction kinetics and/or related parameters, (ii) to perform in-capillary derivatization for improving separation and/or determination, (iii) to develop electrophoretic methods for the simultaneous analysis of anionic and cationic compounds. For the first two purposes, dual injection, which involves sample and reagent, can be realized either from the same end of the capillary (electrophoretically mediated microanalysis, EMMA) or from the two ends of the capillary (electroinjection analysis, EIA). The third objective, with dual injection of sample from the two ends of the capillary, takes advantage of moving cationic and anionic compounds with opposite directions. The foundations of each alternative, conditions necessary for working with them, restrictions, applications as well as perspectives are reviewed in order to establish the advantages, shortcomings, and convenience or no of their use in comparison to conventional CE.     

Pharmacokinetic applications of capillary electrophoresis

This review briefly discusses the use of capillary electrophoretic (CE) methods for the investigations of different aspects of pharmacokinetics. In most investigations, CE was the method of choice because of its unique features, including high resolving power for chiral or metabolite separation, small sample volume for pediatric pharmacokinetics or for cell-based investigations, in situ microdialysis sampling for rapid eliminations, low UV wavelength detection for nonderivatized analytes, fast and simplified sample processing for existing methods that require tedious sample preparation, or as a second method for verifications. Moreover, instrumental aspects of CE-based assays for pharmacokinetic studies, such as different modes of CE methods for analyzing biological samples, sample stacking for increasing detection sensitivity, and coupling techniques with microdialysis and mass spectrometry, are also discussed in this review. Furthermore, the advantages and limitations of CE methods as well as the future outlook for pharmacokinetic studies are summarized.     

High quality drug screening by capillary electrophoresis: A review

High quality assays are needed in drug discovery to reduce the high attrition rate of lead compounds during primary screening. Capillary electrophoresis (CE) represents a versatile micro-separation technique for resolution of enzyme-catalyzed reactions, including substrate(s), product(s), cofactor(s) and their stereoisomers, which is needed for reliable characterization of biomolecular interactions in free solution. This review article provides a critical overview of new advances in CE for drug screening over the past five years involving biologically relevant enzymes of therapeutic interest, including transferases, hydrolases, oxidoreductases, and isomerases. The basic principles and major configurations in CE, as well as data processing methods needed for rigorous characterization of enzyme inhibition are described. New developments in functional screening of small molecules that modulate the activity of disease-related enzymes are also discussed. Although inhibition is a widely measured response in most enzyme assays, other important outcomes of ligand interactions on protein structure/function that impact the therapeutic potential of a drug will also be highlighted, such as enzyme stabilization, activation and/or catalytic uncoupling. CE offers a selective platform for drug screening that reduces false-positives while also enabling the analysis of low amounts of complex sample mixtures with minimal sample handling.     

Energy drink analysis by capillary electrophoresis

Caffeine and vitamins C, PP, and B6 have been determined in energy drinks by capillary electrophoresis. Its advantages and disadvantages over high-performance liquid chromatography have been considered and the influence of analysis conditions on the error of analysis and error sources in capillary electrophoresis have been estimated.     

Element speciation analysis by capillary electrophoresis

An overview of recent developments in the application of capillary electrophoresis to simultaneous separation and determination of different chemical forms of an inorganic element is presented, with particular emphasis placed on metal speciation analysis. Examples of species analysis are addressed, covering metal ions in different oxidation states, metal complexes with inorganic and organic ligands, metalloid oxoanions, organometallic compounds, ionic non-metal species, etc. The speciation performance of capillary electrophoresis is illustrated by a number of practically relevant applications. The method's strengths and current limitations with regard to chemical speciation studies are critically discussed.     

Cyclodextrins in capillary electrophoresis enantioseparations--recent developments and applications

Capillary EKC has been established as a versatile and robust CE method for the separation of enantiomers. Within the chiral selectors added to the BGE CDs continue as the most widely used selectors due to their structural variety and commercial availability. This is reflected in the large number of practical applications of CDs to analytical enantioseparations that have been reported between January 2006 and January 2008, the period of time covered by this review. Most of these applications cover aspects of life sciences such as drug analysis, bioanalysis, environmental analysis, or food analysis. Moreover, new CD derivatives have been developed in an attempt to achieve altered enantioselectivities and to further broaden the application range. Finally, efforts will be summarized that aim at an understanding of the molecular level of the chiral recognition between CDs and the analytes.     

Bioanalytical applications of fast capillary electrophoresis

Capillary electrophoresis is unique among liquid-phase separations in its utility for fast separations. Development of technology such as optical-gating, flow-gating, and microfabrication has allowed separations on the millisecond time scale to be developed. The fast separation times place great demands on the detector systems, frequently requiring detection limits below 1 amol to be practical. The development of such fast separations has opened many new applications not previously feasible for separations-based methods. This has included real time chemical monitoring, detection of short-lived species such as protein conformers or non-covalent complexes, and rapid multi-dimensional separations. Other applications currently being developed include high-throughput assays for clinical laboratories or screening combinatorial libraries. This review covers recent developments in the instrumentation for fast CE and some of the applications.     

Recent applications of capillary electrophoresis-electrospray ionisation-mass spectrometry in drug analysis

A critical review of applications for the period 2000-2004, taken from the Web of Knowledge database, of the technique capillary electrophoresis-electrospray ionisation-mass spectrometry (CE-ESI-MS) in drug analysis is presented. The review is concerned with molecules of mass less than 500 Da, chosen according to selected structural classes in which they give ESI signals primarily as [M+H](+) ions although other ions, such as [M-H](-), [M+Na](+), and [M+NH(4)](+), are also reported. These structural classes are drugs with amine-containing side chains, drugs with N-containing saturated ring structures, 1,4-benzodiazepines, other heterocyclic hypnotics, carbohydrates, sulphonylureas, anthracyclines, sulphonamides, penicillins, cephalosporins, tetracyclines, nitrocatechols, steroids, flavonoids/polyphenols, cannabinols, and miscellaneous molecules. Details are given on the fragmentations, where available, that these ionic species exhibit in-source and in ion-trap, triple quadrupole, and time of flight-mass spectrometers. The review gives a critical evaluation of these recent CE-ESI-MS analytical methods in drug analysis. Analytical information on, for example, sample concentration techniques, CE separation conditions, recoveries from biological media and limits of detection (LODs) are provided. Potential applications of CE-MS to particular drugs or drug classes are also briefly discussed in the text.     

Separation of drug stereoisomers by capillary electrophoresis with cyclodextrins

Using capillary electrophoresis, the enantiomers and isomers of several chiral drug molecules were resolved with cyclodextrins. Parameters affecting the resolution between (+)- and (−)-epinephrine, such as pH, cyclodextrin concentration, buffer concentration, and capillary dimensions were investigated. In addition to this, the effect of cyclodextrin type (β and several derivatized β-cyclodextrins) on resolution between stereoisomers of several chiral drug was also investigated. This study showed that the structural features of the molecule, the derivative groups on the cyclodextrin, the buffer composition and the capillary dimensions influence resolution. The chiral drugs used in this study were propranolol, atenolol, betaxolol, dipivefrin, AL03152 (an aldose reductase inhibitor), AL03363 (an oxidation product of AL03152) and the cis/trans isomers of pilocarpine.     

Analysis of the antiepileptic drug keppra by capillary electrophoresis

A simple and rapid method for determination of the new antiepileptic drug keppra (levetiracetam) by capillary electrophoresis in borate buffer containing sodium dodecyl sulfate is described. The serum was injected without any treatment. The method compared well to high performance liquid chromatography. The mean of keppra in the serum of 35 patients was 25 mg/l (range 7-77 mg/l).     

Determination of organic acid drug counter-ions by capillary electrophoresis

Summary A capillary electrophoresis method is described for the novel application of quantifying levels of the simple organic acid counter-ions of a variety of basic drugs. These counter-ions are organic acids such as succinic and maleic. The method uses indirect UV detection and an electroosmotic flow modifier. Acceptable precision and detector linearity were obtained using internal standards. Method validation was completed and acceptable data was generated. The method is now in routine use for this type of testing.