Electrophoretic separations in PMMA capillaries with uniform and discontinuous buffers

Abstract

The basic phenomena occurring within polymethylmethacrylate (PMMA, Plexiglass) capillaries under the conditions of capillary zone electrophoresis (CZE), isotachophoresis (CITP), and isoelectric focusing (CIEF) have been explored in our laboratory and are compared to those characteristic for untreated fused-silica (FS) capillaries. The origin of charge at the inner wall of the PMMA capillary is different compared to that of FS, this having a significant impact on the magnitude of the electroosmotic flow. While both column materials are characterized with a negative surface charge, electroosmosis in PMMA is considerably smaller over the entire pH range. Furthermore, electroosmosis in PMMA and FS is shown to follow different dependencies on the ionic strength (I). Using a set of experimentally determined values, the electroosmotic mobility (μ_EO) of a PMMA capillary is best described using the relationship μ_EO=a+b/√I. For a FS capillary, μ_EO follows the expression μ_EO=a+b log(I). A dynamic capillary electrophoresis simulation model comprising the determined pH and ionic strength dependent wall titration data as input for calculation of electroosmosis is shown to provide CZE and CITP electropherograms that qualitatively agree well with those obtained experimentally. In contrast to FS and glass, electroosmosis at any pH in PMMA is too weak to perform bidirectional (i.e., simultaneous cationic and anionic) CZE and CITP analyses with a detector placed toward the cathodic capillary end. Furthermore, the same is true for the performance of CIEF with electroosmotic zone displacement. Imposed flow is demonstrated to provide the required net buffer flow. © 2001 John Wiley & Sons, Inc. J Micro Sep 13: 69–83, 2001