Electrophoretic mobilization in capillary isoelectric focusing by a weak acid or an acidic ampholyte as catholyte assessed by computer simulation.

CIEF with cationic electrophoretic mobilization induced via replacing the catholyte with the anolyte or a solution of another acid or amino acid was investigated by computer simulation for a wide range pH gradient bracketed between two amphoteric spacers and short electrode vials with a higher id than the capillary. Dynamic simulations provide insight into the complexity of the mobilizing process in a hitherto inaccessible way. The electrophoretic mobilizing process begins with the penetration of the mobilizing compound through the entire capillary, is followed by a gradual or steplike decrease of pH, and ends in an environment with a non-homogenous solution of the mobilizer. Analytes do not necessarily pass the point of detection in the order of decreasing pI values. Cationic mobilization encompasses an inherent zone dispersing and refocusing process towards the capillary end. This behavior is rather strong with phosphoric acid and citric acid, moderate with aspartic acid, glutamic acid, formic acid and acetic acid, and less pronounced in absence of the cathodic spacer. The data reveal that optical detectors should not be placed before 90 % of capillary length. Aspartic acid, glutamic acid, formic acid and acetic acid provide an environment with a continuously decreasing pH which explains their successful use in optimized two-step CIEF protocols. This article is protected by copyright. All rights reserved.