Capillary electrophoresis separation modes

Table 1 Common Capillary Electrophoresis Separation Modes for the Characterization of Biopharmaceuticals...

Application 2 Capillary Gel Electrophoresis. Recently, Karger and co-workers demonstrated the use of polyacrylamide gel-filled capillaries to separate peptide/protein (SDS PAGE) Qfi) and oligonucleotide mixtures (12,22) by capillary electrophoresis. This mode of CE operation may prove to couple well with on-line radioisotope detection. The results of several preliminary capillary electrophoresis separations using gel-filled capillaries and on-line radioisotope detection using the coincidence unit described here are presented below. 

Capillary electrophoresis (CE) has several unique advantages compared to HPLC, snch as higher efficiency dne to non-parabolic fronting, shorter analytical time, prodnction of no or much smaller amounts of organic solvents, and lower cost for capillary zone electrophoresis (CZE) and fused-silica capillary techniques. However, in CZE, the most popular separation mode for CE, the analytes are separated on the basis of differences in charge and molecular sizes, and therefore neutral compounds snch as carotenoids do not migrate and all co-elute with the electro-osmotic flow. 

Suzuki, S., Kakehi, K., and Honda, S., Two-dimensional mapping of N-gly-cosidically linked asialo-oligosaccharides from glycoproteins as reductively pyridylaminated derivatives using dual separation modes of high-performance capillary electrophoresis, Anal. Biochem., 205, 227, 1992. 

Fan et al. [106] developed a high performance capillary electrophoresis method for the analysis of primaquine and its trifluoroacetyl derivative. The method is based on the mode of capillary-zone electrophoresis in the Bio-Rad HPE-100 capillary electrophoresis system effects of some factors in the electrophoretic conditions on the separation of primaquine and trifluoroacetyl primaquine were studied. Methyl ephedrine was used as the internal standard and the detection was carried out at 210 nm. A linear relationship was obtained between the ratio of peak area of sample and internal standard and corresponding concentration of sample. The relative standard deviations of migration time and the ratio of peak area of within-day and between-day for replicate injections were <0.6% and 5.0%, respectively. 

Identification and quantification of natural dyes need high performance analytical techniques, appropriate for the analysis of materials of complicated matrices containing a small amount of coloured substances. This requirement perfectly fits coupling of modern separation modules (usually high performance liquid chromatography in reversed phase mode, RPLC, but also capillary electrophoresis, CE) with selective detection units (mainly mass spectrometer). 

Also, specific chapters deal with the use of CL reactions as detection mode in FIA (Chapter 12), in separational techniques, such as liquid chromatography (LC) (Chapter 14) or capillary electrophoresis (CE) (Chapter 15), in immunoassay (Chapter 18), and in the development of sensors (Chapter 20). The recent use of this technique for the analysis of DNA (Chapter 19) and a photosensitized CL mode for medical routine and industrial applications (Chapter 17) are also considered in this book. 

This section deals briefly with classical methods based on conventional mixing of the sample and reagents such as the batch mode and low-pressure flow mixing methods, as well as the use of CL detection in continuous separation techniques such as liquid chromatography and capillary electrophoresis for comparison with the unconventional mixing mode. 

Several modes of capillary electrophoretic separation are available ordinary CE, capillary zone electrophoresis, capillary electrokinetic chromatography, capillary gel electrophoresis, capillary electrochromatography, capillary isota-chophoresis, and capillary isoelectric focusing. The different separation mechanisms make it possible to separate a wide variety of substances depending on their mass, charge, and chemical nature.53... 

Reversed-phase HPLC can be compared in terms of its utility with other modes of chromatography and with other separation techniques such as gel electrophoresis, capillary electrophoresis (CE), and capillary electrochromatography. 

One of the major advantages of CE as a separation technique is the wide variety of separation modes available. Analytes can be separated on the basis of charge, molecular size or shape, pi, or hydrophobicity. The same CE instrument can be used for zone electrophoresis, IEF, sieving separations, isotachophoresis, and chromatographic techniques such as MEKC and capillary electrokinetic chromatography. This section provides a brief description of each separation mode. Zone electrophoresis, IEF, and sieving are the primary modes used for protein separations, and these will be discussed in detail in the following sections. 

Chiral separation of drng molecules and of their precursors, in the case of synthesis of enantiomerically pure drugs, is one of the important application areas of HPLC in pharmaceutical analysis. Besides HPLC, capillary electrophoresis (CE) is another technique of choice for chiral separations. Chapter 18 provides an overview of the different modes (e.g., direct and indirect ones) of obtaining a chiral separation in HPLC and CE. The direct approaches, i.e., those where the compound of interest is not derivatized prior to separation, are discussed in more detail since they are cnrrently the most frequently used techniques. These approaches require the use of the so-called chiral selectors to enable enantioselective recognition and enantiomeric separation. Many different molecnles have been nsed as chiral selectors, both in HPLC and CE. They can be classified into three different groups, based on their... 

Subsequently four different CE modes are described in the sections Capillary Zone Electrophoresis, Capillary Gel Electrophoresis, Capillary Isoelectric Focussing, and Micellar Electrokinetic Chromatography (MEKC), respectively. The fundamental principles of the specific separation modes are briefly explained, using appropriate equations where required. In Table 3 all equations are listed. In addition, the influence of both instrumental parameters and electrolytic solution parameters on the optimization of separations is described. 

Capillary zone electrophoresis (CZE) is the most common electrophoretic separation technique due to its simplicity of operation and its flexibility. It is the standard mode for drug analysis, identification of impurities, and pharmacokinetic studies. Other separation modes, such as capillary isotachopho-resis (CITP), micellar electrokinetc chromatography (MEKC), capillary electrochromatography (CEC), capillary gel electrophoresis (CGE), capillary isoelectric focusing, and affinity capillary electrophoresis (ACE), have then-advantages in solving specific separation problems, since the separation mechanism of each mode is different. 

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