Use of High Ionic Strength Buffers for the Separation of Proteins and Peptides with Capillary Electrophoresis

The objective of this study was to prepare a coated capillary to achieve rapid analysis of peptides and amino acids (AAs) by CE-ESI-MS. The coated capillary was prepared by chemical modification of the fused silica (FS) capillary with 2-acrylamido-2-methyl-1-propanesulfonic acid (AMPS) via a silanization reaction followed by an on-column in situ graft polymerization. In preparing capillary coating, AMPS was introduced for the following two purposes: first, to achieve strong and stable EOF in a wide pH range, and second, to obtain good repeatability of separations of peptides and AAs by reducing their adsorption to the FS capillary. The inner wall morphology, the characteristics of EOF, and the performances of coated capillary including separation ability, repeatability, and stability were investigated in detail and evaluated by analyses of four peptides (D-Ala-D-Ala, Gly-Gly-L-Leu, Gly-D-Phe, and L-Leu-D-Leu) and three AAs (glycine, glutamic acid, and phenylalanine), respectively. Under the optimized conditions, the analysis times of four peptides and three AAs in the coated capillary were 5.6 and 6.0 min, respectively, i.e. much shorter than those in the bare-FS capillary, 20 and 26 min, respectively. These coated capillaries were shown to be well suitable for CE coupling with ESI-MS to analyze peptides and AAs thanks to high separation power, short analysis times, good repeatability and high stability.

Section: Introductionmentioning

confidence: 99%

Rapid analysis of peptides and amino acids by CE‐ESI‐MS using chemically modified fused‐silica capillaries

Zhang

et al. 2009

“…In CE, a commonly solution to this is a use of a low pH [11] or high ionic strength buffer, sometimes with an organic modifier [12,13], and a covalent (static) [14] or dynamic coating [14][15][16][17] modification of the capillary surface. Recently, there have been many reviews on this subject [18,19].…”

Section: Introductionmentioning

confidence: 99%

Cationic amylopectin derivatives as additives for analysis of proteins in capillary electrophoresis

Kato

Sakai‐Kato

et al. 2006

Positively charged amylopectin, which is a major constituent of cationic starch, was used to modify the inner surface of fused-silica capillaries by addition to the running solution, which was subsequently employed in CE. Capillaries filled with cationic amylopectin derivatives were shown to generate a stable reversed EOF in the investigated range of pH 4-8. Among the additives studied, quaternary ammonium amylopectin derivatives with high amino and low hydroxypropyl groups showed fast electroosmotic mobility and very effectively suppressed the adsorption of proteins. The run-to-run and batch-to-batch repeatability of the procedures were satisfactory with RSDs of 0.5% and 2.4%, respectively. A basic protein, alpha-chymotrypsinogen, migrated within 6 min and the theoretical plate number of it reached 560 000 plates/m.

“…For this reason, DOI-CE is practically limited to charged analytes, and only when the EOF was not completely suppressed the determination of neutral compounds would be possible which would be moved by the low EOF together with ionic compounds with a higher and opposite sign electrophoretic mobility than the EOF. In order to allow the electrophoretic mobility of the analyte to dominate, some researchers have reduced the EOF by increasing the ionic strength of the buffer [34,35]. Brechtel et al [36] first reported the use of high-charged metal salts for this purpose.…”

Section: Foundations and Requirements Of Doi-cementioning

confidence: 99%

Dual injection capillary electrophoresis: Foundations and applications

Priego-Capote

Castro

2004

The state of the art of capillary electrophoresis (CE) approaches based on dual injection is here reported. Dual injection strategies have been proposed with three main objectives: (i) to provide information about reaction kinetics and/or related parameters, (ii) to perform in-capillary derivatization for improving separation and/or determination, (iii) to develop electrophoretic methods for the simultaneous analysis of anionic and cationic compounds. For the first two purposes, dual injection, which involves sample and reagent, can be realized either from the same end of the capillary (electrophoretically mediated microanalysis, EMMA) or from the two ends of the capillary (electroinjection analysis, EIA). The third objective, with dual injection of sample from the two ends of the capillary, takes advantage of moving cationic and anionic compounds with opposite directions. The foundations of each alternative, conditions necessary for working with them, restrictions, applications as well as perspectives are reviewed in order to establish the advantages, shortcomings, and convenience or no of their use in comparison to conventional CE.