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Migration behaviour of weakly retained, charged analytes in voltage-assisted micro-high performance liquid chromatography

Channer, B. and Skellern, G.G. and Euerby, M.R. and McKeown, A.P. and Rathore, A.S. (2005) Migration behaviour of weakly retained, charged analytes in voltage-assisted micro-high performance liquid chromatography. Journal of Chromatography A, 1095 (1-2). pp. 172-179. ISSN 0021-9673

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The application of voltage in micro-high performance liquid chromatography (micro-HPLC) creates a system where separation is governed by a hybrid differential migration process, which entails the features of both HPLC and capillary zone electrophoresis (CZE), i.e., chromatographic retention and electrophoretic migration. In this paper, we use our previously published approach to decouple these two mechanisms via analysis of the input data for estimation of electrokinetic parameters, such as conductivity, equivalent lengths, mobilities and velocities. Separation of weakly retained, charged analytes was performed via voltage-assisted micro-HPLC. Contrary to conclusions from data analysis using the conventional definitions of the retention factor, it is shown that our approach allows us to isolate the “chromatographic retention” component and thus, investigate the “modification” of the retention process upon application of voltage in micro-HPLC. It is shown that the traditional approaches of calculating retention factor would erroneously lead to conclusion that the retention behavior of these analytes changes upon application of voltage. However, the approach suggested here demonstrates that under the conditions investigated, most of the charged analytes do not show any significant retention on the columns and that all the changes in their retention times can be attributed to their electrophoretic migration.