Resolution capillary zone electrophoresis

C 2 C jj alkyl ether sulfates average E = 1 to 3 partial separation by decreasing alkyl chain length incomplete resolution Capillary zone electrophoresis (CZE) Silica, 50 pm x 49 cm 0.1 M boric acid/0.005 M borax buffer, 0.005 M in naphthalene sulfonate pH 8 30% CH,CN added Cathodic, indirect U V, 274 nm 25... 

Biomolecule Separations. Advances in chemical separation techniques such as capillary zone electrophoresis (cze) and sedimentation field flow fractionation (sfff) allow for the isolation of nanogram quantities of amino acids and proteins, as weU as the characterization of large biomolecules (63—68) (see Biopolymers, analytical techniques). The two aforementioned techniques, as weU as chromatography and centrifugation, ate all based upon the differential migration of materials. Trends in the area of separations are toward the manipulation of smaller sample volumes, more rapid purification and analysis of materials, higher resolution of complex mixtures, milder conditions, and higher recovery (69). 

Weiss, GH Garner, M Yarmola, E Bocek, P Chrambach, A, A Comparison of Resolution of DNA Fragments Between Agarose Gel and Capillary Zone Electrophoresis in Agarose Solutions, Electrophoresis 16, 1345, 1995. 

CE was recently used for anthocyanin analysis because of its excellent resolution. This technique has different modes capillary zone electrophoresis (CZE), capillary gel electrophoresis (CGE), micellar electrokinetic chromatography (MEKC), capillary electrochromatography (CEC), capillary isoelectric focusing (CIEE), and capillary isotachophoresis (CITP)."° CZE is the most popular method for anthocyanin... 

Tsuda, T., Modification of electro-osmotic flow with cetyltrimethylammonium bromide in capillary zone electrophoresis, /. High Resolut. Chromatogr., 10,622, 1987. 

Mosher, R. A., The use of metal ion-supplemented buffers to enhance the resolution of peptides in capillary zone electrophoresis, Electrophoresis, 11, 765, 1990. 

Fanali, S., Ossicini, L., Foret, F., and Bocek, R, Resolution of optical isomers by capillary zone electrophoresis study of enantiomeric and distereoisomeric cobalt (III) complexes with ethylenediamine and amino acid ligands, /. Microcol. Sep., 1, 190, 1989. 

Kuhn, R., Emi, F., Bereuter, T., and Hausler, J., Chiral recognition and enantiomeric resolution based on host-guest complexation with crown ethers in capillary zone electrophoresis, Anal. Chem., 64, 2815, 1992. 

Capillary zone electrophoresis, an up-to-date high resolution separation method useful for proteins and peptides, has been shown to be a useful method for determining electrophoretic mobilities and diffusion coefficients of proteins [3], Diffusion coefficients can be measured from peak widths of analyte bands. The validity of the method was demonstrated by measuring the diffusion coefficients for dansylated amino acids and myoglobin. 

Table 1 summarizes several of the experimental methods discussed in this chapter. A need exists for new or revised methods for transport experimentation, particularly for therapeutic proteins or peptides in polymeric systems. An important criterion for the new or revised methods includes in situ sampling using micro techniques which simultaneously sample, separate, and analyze the sample. For example, capillary zone electrophoresis provides a micro technique with high separation resolution and the potential to measure the mobilities and diffusion coefficients of the diffusant in the presence of a polymer. Combining the separation and analytical components adds considerable power and versatility to the method. In addition, up-to-date separation instrumentation is computer-driven, so that methods development is optimized, data are acquired according to a predetermined program, and data analysis is facilitated. 

We therefore sought to evaluate reproducibility of shotgun proteomics in studies of archival FFPE tissue. Because FFPE samples are more complex than non-cross-linked samples, we evaluated FFPE human liver for analytical reproducibility and confidence in protein assignments.20 This complexity strengthens the argument for using high-resolution separations to maximize analyte concentration and minimize matrix effects. In this case, we used transient capillary isotachophoresis/capillary zone electrophoresis (cITP/cZE) in place of IEF to help address this effect. cITP/cZE has a resolution superior even to cIEF (90% of identified peptides in 1 fraction, 95% in 2 fractions or less for cITP/cZE, vs. 75% and 80%, respectively, for cIEF). 

Amino acids derivatized with 9-fluorenylmethyl chloroformate (90) were separated by CE and determined by LIF with a pulsed laser LOD 0.5 nM (SNR 2)322. A sensitive technique for amino acids is capillary zone electrophoresis (CZE) combined with LIF of their fluorescein isothiocyanate (133) derivatives. Not all amino acids give good resolution. LOD for proline and arginine were 0.3 and 0.5 nM, respectively323. 

Mccomb, M. E., Krutchinsky, A. N., Ens, W., Standing, K. G., and Perreault, H. (1998). Sensitive high-resolution analysis of biological molecules by capillary-zone-electrophoresis coupled with reflecting time-of-flight mass-spectrometry. /. Chromatogr. A 800, 1 — 11. 

Capillary zone electrophoresis is a separation technique that benefits from very high efficiency, not selectivity. This is in contrast to chromatography, for which the converse is true. Differences in mobility in the range of 0.01% can be enough for complete resolution of neighboring peaks. The resolution R is defined as... 

J Cai, JT Smith, Z El Rassi. Determination of the ionization constants of weak electrolytes by capillary zone electrophoresis. J. High Resolut. Chromaogr. 15 30-32 (1992). 

S Fanali, G Caponecchi, Z Aturki. Enantiomeric resolution by capillary zone electrophoresis use of pepsin for separation of chiral compounds of pharmaceutical interest. J Microcolumn Sep 9 9—14, 1997. 

Acid hydrolysis under standard conditions (6M HC1, 110 °C, 24 h) leads to partial decomposition of selenocystine and selenocysteine derivatives, thus making quantification of this amino acid by amino acid analysis difficult. Similarly, acid hydrolysis of 5e-[2-(4-pyr-idinyl)ethyl]selenocysteine peptides, obtained by reduction of the selenocystine peptides with NaBH4 and reaction with 4-vinylpyridine, results in partial decomposition. This de-rivatization, however, is useful for the enantiomeric resolution of the acid hydrolysates by capillary zone electrophoresis by applying host-guest complexation with crown ethers.11" 22 ... 

The application of high-performance capillary zone electrophoresis (HP-CZE) in its various selectivity modes has become a very valuable adjunct to HPLC for the analysis of peptides. For synthetic peptides, in particular, both HPLC and HP-CZE now form essential components of the analytical characterization of these molecules. Increasingly, zonal, micellar, or (biospecific) affinity-based HP-CZE procedures with open tubular capillary systems are adapted to allow resolution with extremely high separation efficiencies (e.g., >105 plates per meter) of synthetic or naturally occurring peptides as part of the determination of their structural, biophysical, or functional properties. Illustrative of these capabilities are the results shown in Figure 19 for the separation of several peptides with different charge and Stokes radius characteristics by HP-CZE. 

Capillary Electrophoresis. Capillary electrophoresis (ce) or capillary zone electrophoresis (cze), a relatively recent addition to the arsenal of analytical techniques (20,21), has also been demonstrated as a powerful chiral separation method. Its high resolution capability and lower sample loading relative to hplc makes it ideal for the separation of minute amounts of components in complex biological mixtures (22,23). 

Nitroaromatic explosives and other nitrated organic explosives are under the normal conditions neutral compounds and therefore cannot be separated directly by capillary zone electrophoresis (CZE) technique. Another separation vector must be introduced in order to achieve the resolution between the solutes. Micellar electrokinetic chromatography (MEKC) is typically employed on microchip scene for separation of nitroaromatic explosives. 

Capillary electrophoresis (CE) coupled to MS has the advantage of high resolution and soft ionization for biomolecules, which may be used to differentiate post-translational modifications and variants of intact proteins and oligonucleotides. Different modes of CE (capillary zone electrophoresis, capillary isoelectric focusing, capillary electrochromatography, micellar electrokinetic chromatography, nonaqueous capillary electrophoresis) to MS as well as online preconcentration techniques (transient capillary isotachophoresis, solid-phase extraction, membrane preconcentration) are used to compensate for the restricted detection sensitivity of the CE methodology [77, 78]. 

Capillary zone electrophoresis (CZE) is the most simple and widely used mode in CE. Separations take place in an open-tube, fused silica capillary under the influence of an electric field. The velocity of the analytes is modified by controlling the pH, viscosity, or concentration of the buffer, or by changing the separation voltage. The electroosmotic flow is often used in this mode to improve resolution or to shorten analysis times. 

Although in some instances it is possible to determine low concentrations of inorganic anions by capillary zone electrophoresis [24], problems arise when the concentrations of the sample constituents vary considerably. This is due to the fact that the determination of microconstituents may require the sample load to have an impact on both the migration velocities and the resolution of analytes [25], The use of indirect detection, as preferred in the case of inorganic anions [24,26] is also less capable of achieving adequate load capacities. 

Capillary zone electrophoresis is another technique which has been used to separate products such as organic acids.26 Separation is based on differences in the mobility of analytes exposed to an electric field. Resolution and separation time in such systems depends on factors including electroosmotic flow (EOF), and a number of approaches for adjusting the EOF have been examined. While some of the approaches (pretreatment of capillaries) are not useful as means of process control, adjusting buffer pH and the electric field27 seem to be possible handles for true feedback control of the separation, although closed-loop operation does not seem to have been attempted. 

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