Electrophoresis separation

Diet soft drinks contain appreciable quantities of aspartame, benzoic acid, and caffeine. What is the expected order of elution for these compounds in a capillary zone electrophoresis separation using a pH 9.4 buffer solution, given that aspartame has pJC values of 2.964 and 7.37, benzoic acid s pfQ is 4.2, and the pfQ for caffeine is less than 0. 

Fig. 2. Zone electrophoresis separation where S, F, S, and F are different materials.

The difference between paper electrophoresis and paper chromatography is that electrophoresis separates by charge whereas chromatography... 

Describe clearly the challenges of interfacing electrochemical detectors to capillary electrophoresis separation systems. How can these challenges be overcome ... 

Locke, BR Carbonell, RG, A Theoretical and Experimental Study of Counteracting Chromatographic Electrophoresis, Separation and Purification Methods 18, 1, 1989. 

SDS gel electrophoresis separation in total denaturing conditions was carried out on the protein of culture filtrates and proteins of known molecular mass. The four dark bands (Figure 2) which appear in the gel between 45 and 36 kDa of the standards were assumed to be PG based on the gel filtration results for PG activity and total protein. The relative molecular mass of the four protein bands were estimated as 45 kDa, 42 kDa, 39 kDa and 36 kDa. It was calculated that about 85% of total protein secreted into the culture medium by K. marxianus consisted of PG. 

Wu, S. and Dovichi, N. J., Capillary zone electrophoresis separation and laser-induced fluorescence detection of zeptomole quantities of fluorescein thiohy-dantoin derivatives of amino acids, Talanta, 39, 173, 1992. 

Guttman, A. and Pritchett, T., Capillary gel electrophoresis separation of high-mannose type oligosaccharides derivatized by l-aminopyrene-3,6,8-trisul-fonic acid, Electrophoresis, 16, 1906, 1995. 

Guttman, A., Cooke, N., and Star, C. M., Capillary electrophoresis separation of oligosaccharides. I. Effect of operational variables, Electrophoresis, 15,1518, 1994. 

McKillop, A.G., Smith, R.M., Rowe, R.C., and Wren, S.A.C., Modeling and prediction of electrophoretic mobilities in capillary electrophoresis separation of alkylpyridines, Anal. Chem. 71, 497, 1999. 

The second step in 2D electrophoresis is to separate proteins based on molecular weight using SDS-PAGE. Individual proteins are then visualized by Coomassie or silver staining techniques or by autoradiography. Because 2D gel electrophoresis separate proteins based on independent physical characteristics, it is a powerful means to resolve complex mixtures proteins (Fig. 2.1). Modem large-gel formats are reproducible and are the most common method for protein separation in proteomic studies. 

Pietrogrande, M.C., Marchetti, N., Dondi, F., Righetti, P.G. (2002). Spot overlapping in two-dimensional polyacrylamide gel electrophoresis separations a statistical study of complex protein maps. Electrophoresis 23, 283. 

Hoffmann, R, Ji, H., Moritz, R. L., Connolly, L. M., Frecklington, D. F., Layton, M. J., Eddes, J. S., Simpson, R. J. (2001). Continuous free-flow electrophoresis separation of cytosolic proteins from the human colon carcinoma cell line LIM 1215 a non two-dimensional gel electrophoresis-based proteome analysis strategy. Proteomics 1(7), 807. 

FIGURE 15.1 One-dimensional capillary electrophoresis separation of a protein homogenate prepared from the hTERT cell line. Both separations were preformed in 30 pm ID, 145 pm OD, 20 cm long capillaries at 20,000 V. (a) Micellar electrokinetic chromatography performed with a 100 mM CHES, 100 mM Tris, and 15 mM SDS buffer at pH 8.7. Sample is electro-kinetically injected with 0.25 kV for 1 s (b) Capillary sieving electrophoresis performed in 5% Dextran (513 kDa), 100 mM CHES, 100 mM Tris, 3.5 mM SDS, pH 8.7. 

This instrumentation has exquisite sensitivity, which allows the analysis of single cancer cells (Hu et al., 2004). Our earlier work employed slow separation conditions and a rather primitive photodetection system. Our current system takes roughly 1 h to complete the two-dimensional capillary electrophoresis separation and employs state-of-the-art photodetectors. 

Issaq, H.J., Chan, K.C., Cheng, S.L., Qingho, L. (2001). Multidimensional high performance liquid chromatography-capillary electrophoresis separation of a protein digest an update. Electrophoresis 22, 1133-1135. 

FIGURE 18.2 Capillary gel electrophoresis separation of an octylphenol ethoxylate sulfate (with an ethylene oxide chain length from 1 to 8). Run conditions pH 8.3 (100 mM tris-borate, 7 M urea) 50 pm x 75 cm J W polyacrylamide gel capillary (PAGE-5, 5%T, and 5%C) run at 20 kV with a 5kV injection for 5 s UV detection at 260nm. 

The reagent is similar to another maleimide-containing biotinylation reagent, 3-(N-maleimi-dopropionyl) biocytin, a compound used to detect sulfhydryl-containing molecules on nitrocellulose blots after SDS-electrophoresis separation (Bayer et al., 1987). Biotin-BMCC should be useful in similar detection procedures. 

C.S. Effenhauser, A. Manz, and M. Widmer, Glass chips for high-speed capillary electrophoresis separations with submicrometer plate heights. Anal. Chem. 65, 2637-2642 (1993). 

Capillary electrophoresis separates by differences in charge while reversed-phase HPLC separates on the basis of hydrophobicity. It turns out that these proteins have wide differences in hydrophobicity. 

Capillary electrophoresis systems are also likely to play an increasingly prominent analytical role in the QC laboratory (Figure 7.2). As with other forms of electrophoresis, separation is based upon different rates of protein migration upon application of an electric field. 

Wang C., Oleschuk R., Ouchen F., Li J., Thibault R, and Harrison D.J. (2000), Integration of immobilized trypsin bead beds for protein digestion within a micro-fluidic chip incorporating capillary electrophoresis separations and an electrospray mass spectrometry interface, Rapid Commun. Mass Spectrom. 14(15), 1377-1383. 

Strege, M.A., Huff, B.E., and Risley, D.S., Evaluation of macrocyclic antibiotic A82846B as a chiral selector for capillary electrophoresis separations, LC-GC, 14, 144, 1996. 

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. 

Zhou SY, Zuo H, Stobaugh JE, Lunte CE, Lunte SM. 1995. Continuous in vivo monitoring of amino acid neurotransmitters by microdialysis sampling with on-line derivatization and capillary electrophoresis separation. Anal Chem 67(3) 594-599. 

Miller, J. H. McB., and Rose, U. (2001). Comparison of chiral liquid chromatographic methods and capillary electrophoresis, separation of the enantiomers of ephedrine hydrochloride. Pharmeuropa 13(1), 3-7. 

Gong, Z. L., Zhang, Y., Zhang, H., and Cheng, J. K. (1999). Capillary electrophoresis separation and permanganate chemiluminescence on-line detection of some alkaloids with beta-cyclodextrin as an additive.. Chromatogr. A 855, 329—335. 

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