Meriem Mokaddem 1 Anne Varenne 1 Jamel-Eddine Belgaied 2 Cécile Factor 3 Pierre Gareil 1 1 Laboratoire d’Electrochimie et Chimie Analytique, UMR 7575 CNRS-ENSCP-Paris 6, Paris, France 2 Laboratoire de Chimie Analytique et d’Electrochimie, (04UR12-01) INSAT, B.P. 676, Tunisie, France 3 Guerbet, BP 50400, Roissy, France Received March 7, 2007 Revised May 3, 2007 Accepted May 18, 2007 Research Article Single-run separation of cationic, anionic, and polyanionic compounds by CE-ESI-MS A method for a single-run separation of cationic, anionic, and polyanionic compounds by CE hyphenated to ESI MS (CE-ESI-MS) is described. One of the main issues for coupling CE to MS with an ESI source consists in maintaining an electric contact for the electro- phoretic separation. This condition is only performed if a liquid flow arising from the separation capillary is directed to the needle, making it coupling-compatible. This latter situation is incompatible with the separations of polyanionic compounds of higher elec- trophoretic mobility (in absolute value) than the electroosmotic mobility, performed in bare fused-silica capillaries under a negative polarity. In this study, several alternative approaches were evaluated to circumvent this difficulty, and applied to the setup of the CE-MS separa- tion of a mixture containing both cationic and polyanionic compounds, which are synthesis intermediates of contrast agents for medical imaging. Eventually, the detection of the cati- onic and anionic compounds in a single run could be obtained by either using neutrally coated polymethylsiloxane (DB-1) capillaries and simultaneously applying a negative volt- age polarity and a pressure allowing to compensate for the residual cathodic EOF or by dynamically modifying the inner wall of a bare fused-silica capillary with a polycationic polymer (hexadimethrine bromide) and using it afterwards under negative voltage polarity. Keywords: CE-ESI-MS / CE-MS DOI 10.1002/elps.200700176 3070 Electrophoresis 2007, 28, 3070–3077 1 Introduction For about 15 years, the hyphenation between CE and MS has received increasing attention. Whereas CE provides low sample consumption, short analysis time, high separation efficiency, various separation modes, and ease of operation and automation, MS detection brings unambiguous identi- fication and m/z separation capability. For coupling CE to MS, the ESI interface is the most commonly employed for its sensitivity, versatility, relative easiness of use, and its suit- ability for the analysis of polar compounds, thereby showing a good match with the type of analytes usually separated by CE [1]. This hyphenation is widely used in a variety of fields, such as biomolecules [2–8] pharmaceuticals [9] and metabo- lites [10], food analysis [11], environmental analysis [12, 13], and investigation of technical products [14]. However, the major limiting difficulty for coupling CE to MS through an ESI interface consists in maintaining an electric contact for the electrophoretic separation. When the outlet electrode and vial are replaced by the interfacing system (interface needle), the electric connexion is provided if a liquid flow arising from the separation capillary is directed to the needle, mak- ing it coupling compatible. If not, electrolyte depletion at the outlet end of the capillary hinders this contact, resulting in a current drop. This latter situation is encountered for the separation of polyanionic compounds of higher electropho- retic mobility (in absolute value) than the electroosmotic mobility, performed in bare fused-silica capillaries under negative CE voltage polarity. For this reason, few papers on anion analysis have been reported by CE-ESI-MS using this configuration. A method for the determination of anionic metabolites based on CE-ESI-MS was developed by Soga et al. [15], requiring inversion of EOF and CE voltage polarity by using a cationic polymer-coated capillary. Various types of monoanionic metabolites could be analyzed in a short time with a relatively high sensitivity, selectivity, and reproduci- bility. For the case of polyanions, the same authors proposed using a neutral-polymer capillary coating to avoid wall adsorption, and applying pressure during separation to pro- duce constant liquid flow toward the anode on the MS side [16]. Other groups developed a new method allowing the simultaneous separation of some anions and polyanions using EOF suppression and an air pressure gradient [17, 18]. However, for a more complex mixture of both cationic, anio- Correspondence: Dr. Anne Varenne, Laboratoire d’Electrochimie et Chimie Analytique, UMR 7575 CNRS-ENSCP-Paris 6, 11, rue Pierre et Marie Curie, F-75231 Paris Cedex 05, France E-mail: anne-varenne@enscp.fr Fax: 133-1-44-27-67-50 Abbreviation: SMIL, successive multiple ionic layer  2007 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim www.electrophoresis-journal.com