Capillary isotachophoresis applications

The hyphenation of CE and NMR combines a powerful separation technique with an information-rich detection method. Although compared with LC-NMR, CE-NMR is still in its infancy it has the potential to impact a variety of applications in pharmaceutical, food chemistry, forensics, environmental, and natural products analysis because of the high information content and low sample requirements of this method [82-84]. In addition to standard capillary electrophoresis separations, two CE variants have become increasingly important in CE-NMR, capillary electrochromatography and capillary isotachophoresis, both of which will be described later in this section. 

Other applications include the online coupling of capillary isotachophoresis and CZE for the quantitative determination of flavonoids in Hypericum perforatum (Guttiferae) leaves and flowers. This method involved the concentration and preseparation of the flavonoid fraction before introduction into the CZE capillary. The limit of detection for quercetin 3-0-glycosides was 100 ng/ml. ... 

Among the electrophoretic methods of chiral resolution, various forms of capillary electrophoresis such as capillary zone electrophoresis (CZE), capillary isotachophoresis (CIF), capillary gel electrophoresis (CGE), capillary isoelectric focusing (CIEF), affinity capillary electrophoresis (ACE), and separation on microchips have been used. However, in contrast to others, the CZE model has been used frequently for this purpose [44]. On the other hand, drawbacks associated with the electrophoretic technique due to lack of development of modem chiral phases have limited the application of these methods. Moreover, the electrophoretic techniques cannot be used at the preparative scale, which represents an urgent need of chiral separation science. 

A range of electrophoretic techniques use the capillary prindple capillary zone electrophoresis (CZE), capillary isotachophoresis (CITP), capillary gel electrophoresis (CGE), capillary isoelectric focusing (CIEF) and micellar electrokinetic capillary chromatography (MECC). Some of these techniques, particularly CZE and CGE, have already established themselves as important analytical tools others, notably MECC, may open new approaches in analytical biochemistry. Table 4-11 summarises the areas of application of the various techniques in what follows we shall focus on CZE, CGE and MECC. 

The first applications of CDs as chiral selectors in CE were reported in capillary isotachophoresis (CITP) [2] and capillary gel electrophoresis (CGE) [3]. Soon thereafter, Fanali described the application of CDs as chiral selectors in free-solution CE [4] and Terabe used the charged CD derivative for enantioseparations in the capillary electrokinetic chromatography (CEKC) mode [5]. It seems important to note that although the experiment in the CITP, CGE, CE, and CEKC is different, the enantiomers in all of these techniques are resolved based on the same (chromatographic) principle, which is a stereoselective distribution of enantiomers between two (pseudo) phases with different mobilities. Thus, enantioseparations in CE are commonly based on an electrophoretic migration principle and on a chromatographic separation principle [6]. 

A15. Arlinger, L., Preparative capillary isotachophoresis—principle and some applications. /. Chromatogr. 119, 9-24 (1976). 

D6. Delmotte, P., Monitoring of protein-protein interaction by capillary isotachophoresis. In Biochemical and Biological Applications of Isotachophoresis (A. Adam and C. Schots, eds.), pp. 259-265. Elsevier, Amsterdam, 1980. 

Capillary isotachophoresis was well established before the introduction of capillary electrophoresis, but was quickly overshadowed by the rapid development of the latter. Current use is limited for analytical applications [387-389] with capillary electrophoresis being preferred in most cases. Renewed interest in capillary isotachophoresis as a sample concentration and preseparation technique for capillary electrophoresis is responsible for a somewhat limited revival. The self-sharpening and concentration characteristics of capillary isotachophoresis make it more suitable than capillary electrophoresis for preparative scale separations, where single run and continuous flow instalments have been described for the isolation of milligram to gram quantities of material [392-394]. Capillary isotachophoresis is also suitable for the determination of values for effective ion mobility [395,396]. 

Capillary Isotachophoresis CIT is a moving-boundary technique. The sample is sandwiched between a leading electrolyte with mobility higher than any of the sample components, and a terminating electrolyte with mobility lower than any of the sample components. Upon application of the electrical field, the sample ions are separated into bands according to their electrophoretic mobility. Once the band is formed, all ions that are separated migrate at the same velocity. 

Over the last two decades, capillary electrophoresis (CE) has been developed as a powerful separation technique for complex mixtures. Its advantages include a high separation efficiency, short analysis time, small sample requirement, and applicability to a wide range of analytes. The basic modes of CE that are presently being exploited include capillary zone electrophoresis (CZE), micellar electrokinetic chromatography (MEKC), capillary isotachophoresis, capillary isoelectric focusing, and capillary gel electrophoresis. 

Gebauer, P. Bocek, P. Recent application and developments of capillary isotachophoresis. Electrophoresis 1997, 18, 2154. 

Capillary isotachophoresis (cITP) uses capillaries with pH gradients for separation of macromolecular species with different isoelectric points (pi). On application of an electric field, ions move until they reach the zone with the pH eorresponding to their pi value, at whieh point their overall electric charge is null. This mode is particularly useful in the eharaeterization of protein samples. 

A variety of formats and options for different types of applications are possible in CE, such as micellar electrokinetic chromatography (MEKC), isotachophoresis (ITP), and capillary gel electrophoresis (CGE). The main applications for CE concern biochemical applications, but CE can also be useful in pesticide methods. The main problem with CE for residue analysis of small molecules has been the low sensitivity of detection in the narrow capillary used in the separation. With the development of extended detection pathlengths and special optics, absorbance detection can give reasonably low detection limits in clean samples. However, complex samples can be very difficult to analyze using capillary electrophoresis/ultraviolet detection (CE/UV). CE with laser-induced fluorescence detection can provide an extraordinarily low LOQ, but the analytes must be fluorescent with excitation peaks at common laser wavelengths for this approach to work. Derivatization of the analytes with appropriate fluorescent labels may be possible, as is done in biochemical applications, but pesticide analysis has not been such an important application to utilize such an approach. 

Miiller et al., 2000 Her et al., 2003). Emphasis has been placed on (a) the use of gel chromatography or gel permeation chromatography for the fractionation of DOM on the basis of molecular size differences and (b) the application of electrophoretic separation methods (Perminova et al., 1998,2003 Specht and Frimmel, 2000), including electrophoresis, capillary electrophoresis (CE), isotachophoresis, isolelectric focusing,polyacrylamide gel electrophoresis (PAGE), and capillary zone electrophoresis (CEZ) (De Nobili et al., 1989,1998 Schmitt-Kopplin et al., 1998). 

Two kinds of conductivity detector are distinguished contact detectors and contactless detectors. Both types were originally developed for isotachophoresis in 0.2-0.5-mm-inner diameter (i.d.) PTFE tubes. Contactless detectors are based on the measurement of high-frequency cell resistance and, as such, inversely proportional to the conductivity. The advantage is that electrodes do not make contact with the buffer solution and are, therefore, outside the electric field. As these types of detectors are difficult to miniaturize down to the usual 50-75-jU.m capillar inner diameter, their actual application in capillary electrophoresis (CE) is limited. 

Electrophoretic methods are widely used alternatives for the analytical determination of the enantiomeric purity of chiral compounds [194]. Due to the high elTi-ciency of capillary electrophoresis, separations can be achieved even when very low selectivities are observed. At a preparative scale, these methods are well established for the purification of proteins and cells [195] but there is very little published on enantioselective separations. Only recently, some interest in chiral preparative applications has been manifested. Separation of the enantiomers ofterbu-taline [196] and piperoxan [197] have been reported by classical gel electrophoresis using sulfated cyclodextrin as a chiral additive, while the separation of the enantiomers of methadone could be successfully achieved by using free-fluid isotachophoresis [198] and by applying a process called interval-flow electrophoresis [199]. 

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