Capillary Electroseparations in Pharmaceutical Analysis of Basic Drugs and Related Substances

Enlund, A.M. 2001. Capillary Electroseparations in Pharmaceutical Analysis of Basic Drugs and Related Substances. Acta Universitatis Upsaliensis. Comprehensive Summaries of Uppsala Dissertations from the Faculty of Pharmacy 246, 49 pp. Uppsala. ISBN 91-554-4908-5 Capillary electroseparation methods are exciting new techniques with very broad application areas and vast potential in pharmaceutical and biomedical analysis. To improve the limit of detection (LOD) capillary zone electrophoresis (CZE) has been combined with isotachophoretic (ITP) preconcentration in a single capillary. Using the ITP-CZE combination the LOD can be improved at least 100-fold. Laser-induced fluorescence (LIF) detection is more sensitive and more selective than the most common detection technique, UV, and the intensity and focusing capability of LIF fits well with the small dimensions in CZE. The total sensitivity enhancement attained for a new acetylcholinesterase inhibitor, NXX-066, by using ITP-CZE-LIF was more than 5500-fold compared to CZE-UV. Capillary electrochromatography (CEC) combines the high separation efficiency of CZE with the vast possibilities to improve selectivity of HPLC. We have examined different ways to solve the problem of extensively tailing peaks and studied the influence of the mobile phase composition on the electrochromatographic performance for a number of tricyclic antidepressants and related quaternary ammonium compounds: (1) Adding aliphatic amines to the mobile phase in reversed phase CEC. The effect on the chromatographic performance was coupled to the hydrophobicity of the additive and the amine of our choice was dimethyloctylamine. (2) Silicabased cation exchangers with different pore size. The large-pore materials promoted pore flow, but this had no positive influence on the performance. The small-pore (highest surface area) particles gave the best selectivity. (3) Designing special continuous beds. As the bed is covalently attached to the capillary wall, problems related to retaining frits are avoided. The stationary phase most suitable for our analytes had a molar ratio of 1:80 between the functional ligands, vinyl sulphonic acid and isopropyl groups, respectively. The LOD was lowered 26000-fold by dissolving the sample in a low-conducting medium. Anna Maria Enlund, Department of Medicinal Chemistry, Division of Analytical Pharmaceutical Chemistry, Uppsala University Biomedical Centre, Box 574, SE-751 23 Uppsala, Sweden  Anna Maria Enlund 2001 ISSN 0282-7484 ISBN 91-554-4908-5 Printed in Sweden by Uppsala University, Tryck & Medier, Uppsala 2001 Table of contents Papers discussed.................................................................... ........6 Abbreviations........................................................................ ........7 1 Analytes used in the studies.................................................... ........8 Introduction.......................................................................... ......11 2 Basic principles and phenomena in capillary electrophoresis........ ......11 2.1 Electroosmosis........................................................... ......13 2.2 Separation principles.................................................. ......14 2.2.1 Capillary zone electrophoresis.................................... ......14 2.2.2 Micellar electrokinetic chromatography........................ ......14 2.2.3 Capillary electrochromatography.............................. ......14 3 Aim of studies....................................................................... ......15 4 Part one. Detectability improvements in capillary electrophoresis by isotachophoretic preconcentration........................................ ......15 4.1 On-line preconcentration methods for CE....................... ......16 4.1.1 Stacking.................................................................. ......16 4.1.2 Single capillary isotachophoretic preconcentration......... ......17 4.2 Laser-induced fluorescence.......................................... ......21 4.3 Conclusions part one................................................... ......23 5 Part two. Capillary electrochromatography and analysis of basic drugs.................................................................................... ......24 5.1 Aliphatic amines in the mobile phase to improve the chromatographic performance in reversed phase capillary electrochromatography................................................ ......26 5.2 Capillary electrochromatography on strong cation exchangers................................................................ ......31 5.3 Capillary electrochromatography on continuous beds...... ......38 5.4 Conclusions part two................................................... ......42 6 Future studies....................................................................... ......43 7 Acknowledgements................................................................ ......44 8 References............................................................................ ......46