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Anion-Exchange Displacement Centrifugal Partition Chromatography

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Anal. Chem. 2004, 76, 6179-6186 Articles Anion-Exchange Displacement Centrifugal Partition Chromatography Alexandre Maciuk,† Jean-Hugues Renault,*,† Rodolphe Margraff,‡ Philippe Tre ́ buchet,§ Monique Ze` ches-Hanrot,† and Jean-Marc Nuzillard† FRE CNRS 2715 Isolement, Structures, Transformations et Synthe`ses des Substances Naturelles, IFR 53 Biomole ́cules, Universite ́ de Reims Champagne-Ardenne, Baˆt. 18, Moulin de la Housse, F-51687 Reims Cedex 2, France, Partus Technologies, 2 alle ́e Albert Caquot, F-51100 Reims, France, and INRIA, 2004 route des Lucioles, F-06902 Sophia Antipolis, France Ion-exchange displacement chromatography has been adapted to centrifugal partition chromatography. The use of an ionic liquid, benzalkonium chloride, as a strong anion-exchanger has proven to be efficient for the pre- parative separation of phenolic acid regioisomers. Multi- gram quantities of a mixture of three hydroxycinnamic acid isomers were separated using iodide as a displacer. The displacement process was characterized by a trap- ezoidal profile of analyte concentration in the eluate with narrow transition zones. By taking advantage of the partition rules involved in support-free liquid-liquid chromatography, a numerical separation model is pro- posed as a tool for preliminary process validation and further optimization. The concept of countercurrent chromatography (CCC) is now widely used to refer to a solid support-free liquid-liquid chroma- tography which uses two immiscible solvents (or solutions) prepared from an equilibrated liquid-liquid biphasic system. Since its invention by Ito in the late 1960s, many applications and developments have been proposed.1-3 Centrifugal partition chro- matography (CPC) is one of the few technical solutions to the challenge of maintaining a liquid phase stationary while another one is pumped through it. It is based on partition cells radially engraved in a disk connected to each other by capillary ducts (Figure 1). The centrifugal force field resulting from disk rotation causes decantation in each cell, thus allowing for a continuous process. One of the most striking aspects of this concept is the ability the *Corresponding author. Fax: +3 26 91 35 96. E-mail: jh.renault@ univ-reims.fr. † Universite ́ de Reims Champagne-Ardenne. ‡ Partus Technologies. § INRIA. (1) Mandava, N. B., Ito, Y., Eds. Countercurrent Chromatography - Theory and Practice; Chromatographic Science Series 44; Marcel Dekker: New York, 1988. (2) Foucault, A. P., Ed. Centrifugal Partition Chromatography; Chromatographic Science Series 68; Marcel Dekker: New York, 1995. (3) Berthod, A., Ed. Countercurrent Chromatography - The Support-Free Liquid Stationary Phase; Wilson & Wilson’s Comprehensive Analytical Chemistry 38; Elsevier Science B.V.: Amsterdam, 2002. 10.1021/ac049499w CCC: $27.50 © 2004 American Chemical Society Published on Web 09/24/2004 Figure 1. Kromaton Technologies FCPC 200 mL rotor (left) and partition disk (right). Note the connecting ducts centered on the bottom and the top of each cell. The upper and lower cell walls consist of the interdisk Teflon gaskets. user has to customize the mobile and stationary phases to obtain a wide range of chromatographic behaviors. Consequently, all of the development modes used in liquid chromatography can be applied to CCC and CPC, especially the displacement mode. The latter was proposed for the first time by Tiselius in 1943 and is mainly used today for preparative separations in the biotechnology industry.4,5 Basically, displacement chromatography in CCC can be performed by dissolving a displacer in the mobile phase and a retainer or an ion-exchanger in the stationary phase.6 By adding an acid or a base in the stationary phase as a retainer, Ito introduced the pH-zone-refining mode.7 For the first time in CCC and CPC, isotachic rectangularly shaped blocks of analytes separated by steep boundaries, the so-called shock layers, were observed.8 This protocol is restricted to solutes showing a dramatic (4) Cramer, S. M.; Subramanian, G. Sep. Purif. Methodol. 1990, 19, 31-91. (5) Gorshkov, V.; Muraviev, D.; Warshawsky, A. Solvent Extr. Ion Exch. 1998, 16, 1-73. (6) Billardello, B.; Berthod, A. In Countercurrent Chromatography - The Support- Free Liquid Stationary Phase; Wilson & Wilson’s Comprehensive Analytical Chemistry 38; Berthod, A., Ed.; Elsevier Science B.V.: Amsterdam, 2002; pp 177-200. (7) Ito, Y.; Ma, Y. J. Chromatogr., A 1996, 753, 1-36. (8) Zhu, J.; Ma, Z.; Guiochon, G. Biotechnol. Prog. 1993, 9, 421-428. Analytical Chemistry, Vol. 76, No. 21, November 1, 2004 6179

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