The preparation of background electrolytes in capillary zone electrophoresis: Golden rules and pitfalls

Abstract

In this article the methodology of the design of suitable background electrolytes (BGEs) in capillary zone electrophoresis (CZE) is described. The principal aspects of the role of a BGE in CZE are discussed with respect to an appropiate migration behavior of analytes, including the transport of the electric current, the buffering of pH, the Joule heat, the electro-endosmotic flow (EOF) and the principal migration and detection modes. The impact of the composition of the BGE upon migration and detection is discussed. It is shown that the total concentration of the BGE is a principal factor and the adjustment of migrating analyte zones according to the Kohlrausch regulating function (KRF) is the principal effect in most of the sample stacking techniques. The number of co-ions and their properties are of key importance for peak shapes of the analyte peaks and for the existence of system zones. The detection of UV-transparent analytes may advanteously be done in the indirect UV mode, by using UV-absorbing co-ions, however, both peaks and dips may be expected in the UV trace in case of multiple co-ionic BGEs. Properties of BGEs can be predicted applying mathematical models and it is shown that with SystCharts, predictions can be given concerning the existence of system zones, detection modes and the peak shapes of analytes for a given BGE. Practical examples of methodological considerations are given in the design of suitable BGEs for four principal combinations of migration and detection modes. The properties of the BGEs selected are exemplified with experimental results. Golden rules are summarized for the preparation of suitable BGEs in CZE.

System zones in capillary zone electrophoresis

“…The appearance of the "hydrogen" or "hydroxide" system peak is often explained as a consequence of the fact that H and OH 2 ions in significant concentration behave as a "co-ion", which causes an additional system zone [10]. We will show later in the text that this is not an entirely correct approach.…”

“…The number of system zones is a topic often discussed [5][6][7][8][9][10]13] and various authors still have a different opinion. Nevertheless, it comes out clearly from the linearized model of electromigration [5,13] that the number of eigenvalues of the system, i.e., the number of eigenmobilities, is the same as the number of constituents, and also the number of the eigenzones is the same.…”

“…This is due to rather complex equations describing electromigration, which are inherently nonlinear. Although papers by Beckers, Gebauer, and Boč ek [8][9][10] reviewing the behavior of system zones were published recently, we feel that a progress in the mathematical description of electromigration [11][12][13][14] allows a deeper understanding of the behavior of system zones now. This mathematical model unifies the classification of the system zones and even allows to predict some interesting phenomena overlooked so far, such as the existence of BGEs having no stationary injection zone.…”

Capillary electrophoresis of inorganic ions: An update

“…An extensive book chapter on CEC published by Remcho et al [54] and a similar review article by Kok and coauthors [55] contain excellent mixes of inorganic and other appli- [9,52] cations. Beckers and Boč ek [5] have extensively treated the methodology of the design of suitable background electrolytes (BGEs) in CE. Concluded with a summary of the most important basic principles (golden rules) how to prepare a BGE for a good migration/detection, this work would be particularly useful to facilitate the task of method development by those just beginning to work in the field.…”

Salt removal during Off-Gel? electrophoresis of protein samples

“…Concerning electromigration phenomena, numerous mathematical and numerical studies have been presented in the literature, as for example [29][30][31][32][33][34][35][36][37], to describe the different modes of capillary electrophoresis or IEF processes. In a first approximation, the gel is considered as a supporting (i.e., background) electrolyte, ensuring the initial current in the gel and the electroneutrality condition when the ions from the chamber (i.e., K…”