Introduction Separation of enantiomers is of utmost importance in drug research, because of the requirement for high purity grade ingredients for pharmaceutical formu- lations. Among the various techniques for enantioseparation, chromatographic techniques, such as high-performance liquid chromatography (HPLC), play an important role. Enantiomers interact differently and selectively with a chiral stationary phase (CSP), therefore exhib- iting distinct retention times (Figure 1). Stationary phases often consist of porous silica particles modified by complex bind- ing groups (selectors), which are respon- sible for the selective interaction with the analytes (selectands), thus providing separation. The selectands are injected in the column and eluted with the mobile phase (solvent), which has opposite polarity to that of the CSP. In modern instruments, the separated analytes are identified typically using a UV detector, the main reason being the excellent detection limit of the method. Infrared spectroscopy is also a potent liquid chro- matographic detection method 1 and a number of pertinent techniques have been developed in the last two decades. 2 The advantage of infrared spectroscopy over UV methods is that a direct molec- ular identification of the analyte can be achieved, given the fact that the IR spec- trum of an organic compound provides a unique fingerprint, which makes it distin- guishable from other compounds. It is obvious that the interactions occur- ring at the solid–liquid interface between the CSP and the selectands dissolved in the mobile phase are the essence of an HPLC experiment. The separation of a racemic mixture is based on the differ- ent energies of the two diastereomeric complexes that are formed upon interac- tion between the CSP and the enantiom- ers. The larger the difference in the free energy between the two diastereomeric analyte–CSP complexes is, the higher is the separation capacity. To success- fully separate enantiomers, at least three interactions (three-point model) have to be generated between one enanti- omer and the CSP, including attractive (ionic, hydrogen bonding, π–π interac- tion, van der Waals) and repulsive (steric hindrance) interactions. The rational design of a new CSP requires the understanding of the nature of its interaction with the analytes. At this point in situ techniques come into play. The determination of crucial inter- actions manifesting at solids in contact with liquids is the goal of our research and we pursue this goal applying infra- red spectroscopy of the solid–liquid interface, during, for example, the chro- matographic event. Hence, we report here on an analytical technique based on the combination of attenuated total reflection infrared (ATR-IR) spectroscopy 3 and modulation excitation (ME), 4 which enables the investigation of the inter- 8 SPECTROSCOPYEUROPE ARTICLE ARTICLE www.spectroscopyeurope.com VOL. 19 NO. 1 (2007) Probing chiral recognition in liquid chromatography by absolute configuration modulation ATR-IR spectroscopy Ronny Wirz, a Davide Ferri, a* Thomas Bürgi b and Alfons Baiker a a Institute for Chemical and Bioengineering, ETH Zurich, Wolfgang-Pauli-Strasse 10, HCI, CH–8093 Zurich, Switzerland. E-mail: davide.ferri@chem.ethz.ch b Institute of Microtechnology, University of Neuchâtel, Rue Emile-Argand 11, CH–2009 Neuchâtel, Switzerland Figure 1. In chiral liquid chromatography a racemic mixture is separated because of the different adsorption strength of the enantiomers on the chiral stationary phase. The less strongly bound enantiomer is eluted faster and shows a shorter retention time.