Capillary electrophoresis solutions

First, solutes with larger electrophoretic mobilities (in the same direction as the electroosmotic flow) have greater efficiencies thus, smaller, more highly charged solutes are not only the first solutes to elute, but do so with greater efficiency. Second, efficiency in capillary electrophoresis is independent of the capillary s length. Typical theoretical plate counts are approximately 100,000-200,000 for capillary electrophoresis. 

Selectivity In chromatography, selectivity is defined as the ratio of the capacity factors for two solutes (equation 12.11). In capillary electrophoresis, the analogous expression for selectivity is... 

The basic instrumentation for capillary electrophoresis is shown in Figure 12.41 and includes a power supply for applying the electric field, anode and cathode compartments containing reservoirs of the buffer solution, a sample vial containing the sample, the capillary tube, and a detector. Each part of the instrument receives further consideration in this section. 

Injecting the Sample The mechanism by which samples are introduced in capillary electrophoresis is quite different from that used in GC or HPLC. Two types of injection are commonly used hydrodynamic injection and electrokinetic injection. In both cases the capillary tube is filled with buffer solution. One end of the capillary tube is placed in the destination reservoir, and the other is placed in the sample vial. 

A means of concentrating solutes in capillary electrophoresis after their injection onto the capillary column. 

Detectors Most of the detectors used in HPLC also find use in capillary electrophoresis. Among the more common detectors are those based on the absorption of UV/Vis radiation, fluorescence, conductivity, amperometry, and mass spectrometry. Whenever possible, detection is done on-column before the solutes elute from the capillary tube and additional band broadening occurs. 

Solutes that do not absorb UV/Vis radiation or undergo fluorescence can be detected by other detectors. Table 12.8 provides a list of detectors used in capillary electrophoresis along with some of their important characteristics. 

A form of capillary electrophoresis in which separations are based on differences in the solutes electrophoretic mobilities. 

Capillary Zone Electrophoresis The simplest form of capillary electrophoresis is capillary zone electrophoresis (CZE). In CZE the capillary tube is filled with a buffer solution and, after loading the sample, the ends of the capillary tube are placed in reservoirs containing additional buffer solution. Under normal conditions, the end of the capillary containing the sample is the anode, and solutes migrate toward... 

Weber, P. L. Buck, D. R. Capillary Electrophoresis A Past and Simple Method for the Determination of the Amino Acid Composition of Proteins, /. Chem. Educ. 1994, 71, 609-612. This experiment describes a method for determining the amino acid composition of cyctochrome c and lysozyme. The proteins are hydrolyzed in acid, and an internal standard of a-aminoadipic acid is added. Derivatization with naphthalene-2,3-dicarboxaldehyde gives derivatives that absorb at 420 nm. Separation is by MEKC using a buffer solution of 50 mM SDS in 20 mM sodium borate. 

Capillary Electrophoresis. Capillary electrophoresis (ce) is an analytical technique that can achieve rapid high resolution separation of water-soluble components present in small sample volumes. The separations are generally based on the principle of electrically driven ions in solution. Selectivity can be varied by the alteration of pH, ionic strength, electrolyte composition, or by incorporation of additives. Typical examples of additives include organic solvents, surfactants (qv), and complexation agents (see Chelating agents). 

P. D. Grossman, J. C. Colburn, H. H. Lauer, R. G. Nielsen, R. M. Riggin, G. S. Sittampalam and E. C. Rickard, Application of free-solution capillary electrophoresis to the analytical scale separation of proteins and peptides . Anal. Chem. 61 1186-1194 (1989). 

Electropherograms of a urine sample (8 ml) spiked with non-steroidal anti-inflammatory drugs (10 p-g/ml each) after direct CE analysis (b) and at-line SPE-CE (c). Peak identification is as follows I, ibuprofen N, naproxen K, ketoprofen P, flurbiprofen. Reprinted from Journal of Chromatography, 6 719, J. R. Veraait et al., At-line solid-phase exti action for capillary electrophoresis application to negatively charged solutes, pp. 199-208, copyright 1998, with permission from Elsevier Science. 

Kim, Y Morris, MD, Pulsed Field Capillary Electrophoresis of Multikilobase Length Nucleic Acids in Dilute Methyl Cellulose Solutions, Analytical Chemistry 66, 3081, 1994. 

High performance capillary electrophoresis was introduced originally as an analytical tool. Now that instruments are equipped with automated fraction collection, however, capillary electrophoresis can be used for micropreparative collection of individual peaks separated from a mixture. Using the fraction collection feature, nanomolar amounts of solute such as proteins, peptides, oligonucleotides can be collected in amounts sufficient for microsequencing. An intersample washing procedure and use of well-formed capillaries aid in the prevention of artifacts.44... 

In the previously described electrophoretic methods, the capillary was filled with electrolytes only. Another mode of operation in capillary electrophoresis involves filling the capillary with gel or viscous polymer solutions. If desired, a column can be packed with particles and equipped with a frit.68 This mode of analysis has been favorably used for the size determination of biologically important polymers, such as DNA, proteins, and polysaccharides. The most frequently used polymers in capillary gel electrophoresis are cross-linked or linear polyacrylamide,69 cellulose derivatives,70-75 agarose,76 78 and polyethylene glycols. 

Bullock, J.A. and Yuan, L.-C., Free solution capillary electrophoresis of basic proteins in uncoated fused silica capillary tubing, ]. Microcol. Sep., 3,241,1991. 

Tran, A. D., Park, S., Lisi, P. J., Huynh, O. T., Ryall, R. R., and Lane, P. A., Separation of carbohydrate-mediated microheterogeneity of recombinant human erythropoietin by free solution capillary electrophoresis. Effects of pH, buffer type and organic additives,. Ckromatogr., 542, 459, 1991. 

Bocek, P. and Chrambach, A., Capillary electrophoresis in agarose solutions extension of size separations to DNA of 12 kb in length, Electrophoresis, 13, 31, 1992. 

Grossman, P. D., Colburn, J. C., and Lauer, H. H., A semiempirical model for the electrophoretic mobilities of peptides in tree-solution capillary electrophoresis, Anal. Biochem., 179, 28, 1989. 

Lee, K.-J. and Heo, G. S., Free solution capillary electrophoresis of proteins using untreated fused-silica capillaries, ]. Chromatogr., 559, 317, 1991. 

Shimizu, T. and Kenndler, E., Capillary electrophoresis of small solutes in linear polymer solutions Relation between ionic mobility, diffusion coefficient and viscosity, Electrophoresis, 20, 3364, 1999. 

Hoagland, D.A., Arvanitidou E., and Welch C., Capillary Electrophoresis measurements of the free solution mobility for several model polyelectrolyte systems, Macromolecules, 32, 6180, 1999. 

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