Characterization of polyethylene glycol-modified proteins by semi-aqueous capillary electrophoresis

A new program to characterize polyethylene glycol-modified (PEGylated) prot eins is outlined using capillary zone electrophoresis (CZE). PEGylated ribo nuclease A and lysozyme were selected as examples. Five separation procedur es were compared to select out the mixed buffer of acetonitrile-water (1:1, v/v) at pH 2.5 as the best to characterize the PEGylated proteins without sample pretreatment. Polyethylene oxide (PEO) with a high molecular mass of 8X10(6) was applied to rinse the capillary to form a dynamic coating which would decrease the undesirable proteins adsorbed to the inner wall of the silica. The electroosmotic flow (EOF) mobility of the five procedures was d etermined, respectively. It is found that acetonitrile is mainly responsibl e for the good resolution of PEGylated proteins with the help of PEO coatin g in the semi-aqueous system. The low EOF mobility and current in the semi- aqueous system might also have some responsibility for the high resolution. The semi-aqueous procedure described in this paper also demonstrates highe r resolution of natural proteins than aqueous ones. (C) 2001 Elsevier Scien ce B.V. All rights reserved.