Treatments of fused-silica capillaries and their influence on the electrophoretic characteristics of these columns before and after coating

Abstract

Making coated columns in capillary electrophoresis (CE) is a laborious task involving many preparation steps such as etching, leaching, dehydration, silylation and coating of the inner wall of fused-silica capillaries. In this work we demonstrate, by testing more than 250 columns, that it is possible to follow up the influence of the different steps on both the electrophoretic behavior of CE columns and the reproducibility of their preparation. This study was done by carrying out triplicate measurements of electroosmotic flow values from columns at four different pH values after each step. The effectiveness of the coatings was also investigated by injecting a test-group of basic proteins. It is demonstrated that etching the columns with sodium hydroxide followed by a leaching treatment with hydrochloric acid provides higher reproducibility than by either leaching or etching alone. Dehydration of the capillary tubing affects the yield of the subsequent silylation reaction while the best results seem to be obtained by dehydrating the columns overnight at 160°C. The silylation degree achieved is demonstrated to also depend on the time of reaction and the concentration of silyl reagent. Moreover, a conclusive demonstration about the effect of both silylation and polymerization reactions on the final coating performance is given.