Capillary zone electrophoresis of biological substances with fused silica capillaries having zero or constant electroosmotic flow

Abstract

A series of capillary surface modifications entailing multilayered coatings were introduced and evaluated in capillary zone electrophoresis of biological substances, e.g., proteins, peptides, oligosaccharides and nucleotides. In one set of surface modifications, large molecular weight hydroxypropyl cellulose afforded “zero” flow capillaries, which were used as precursors for developing anodal flow capillaries. When “zero” flow capillaries were further functionalized with a charged polyethyleneimine layer to which a top polyether layer was covalently attached, the resulting anodal electroosmotic flow was relatively weak due to the high viscosity of the coated wall imparted by the hydroxypropyl cellulose layer. Capillaries with relatively strong and constant anodal electroosmotic flow were best achieved when the inner capillary surface was first chemically derivatized with methylated (i.e., quaternarized) polyethyleneimine hydroxyethylated. This hydroxylated and permanently charged polymeric coating yielded constant anodal flow regardless of electrolyte pH. The hydroxyl groups of the charged polymeric coating permitted the covalent attachment of polyether chains, which minimized electrostatic interaction between the positive charges of the polymeric layer and oppositely charged biopolymers. Under these conditions, rapid transport of acidic biopolymers past the detection point in the separation capillary could be achieved, and relatively high plate counts were obtained.