André De Villiers 1 Gerd Vanhoenacker 2 Fréderic Lynen 1 Pat Sandra 1, 2 1 University of Stellenbosch, Department of Chemistry, CENSSUS, Matieland, South Africa 2 Research Institute for Chromatography, Kortrijk, Belgium Stir bar sorptive extraction-liquid desorption applied to the analysis of hop-derived bitter acids in beer by micellar electrokinetic chromatography Stir bar sorptive extraction-liquid desorption (SBSE-LD) has been applied as an effi- cient sample preparation method for the analysis of beer bitter acids. Extracts free of almost all interfering compounds were obtained, allowing simultaneous analysis of iso-a-acids and reduced iso-a-acids. A robust micellar electrokinetic chromatography (MEKC) method was developed that enables fast separation of iso-a-acids and reduced iso-a-acids. Quantitative data are in good agreement with results obtained by high-performance liquid chromatography (HPLC) using direct beer injection. Keywords: Beer / Hop bitter acids / Liquid desorption / Micellar electrokinetic chromatography / Stir bar sorptive extraction DOI 10.1002/elps.200305758 1 Introduction The bitter taste of beer is derived from hop (Humulus lupus L.) or hop extracts added to the wort during brew- ing [1]. In the boiling process the hop a-acids or humu- lones are converted into the iso-a-acids or isohumu- lones. These products are not only responsible for the bitter taste of beer but also exhibit bacteriostatic prop- erties and furthermore play an essential role in the foam stability of beer as well as in the formation of off-flavors like the sunstruck flavor [2–4]. Reduced iso-a-acids such as rho-iso-a-acids and tetrahydro-iso-a-acids are nowa- days often added in the brewing process to enhance the light and foam stability. HPLC is intensively used for the analysis of a- and b-acids in hops [5–7] and iso-a-acids in beer [2, 8]. Problems related with interaction of the solutes with trace metals in the chromatographic system resulting in poor resolution and quantitation have been reported [2, 8]. Electrodriven separation techniques such as micellar electrokinetic chro-matography (MEKC) [9, 10], microemulsion electrokinetic chromatography (MEEKC) [11], and capillary electrochromatography (CEC) [12] have been evaluated for the analysis of the hop acids while the iso-a-acids were successfully separated by MEKC [13, 14]. For the analysis of the iso-a-acids in beer, a preconcentration step is normally performed, although direct injection of beers with stacking has been described [15]. Enrichment is commonly carried out by liquid-liquid extraction or by solid-phase extraction on reversed-phase material [13, 15, 16]. Recently, a new solventless extrac- tion method named stir bar sorptive extraction (SBSE) has been described [17]. SBSE is commonly combined with thermal desorption (TD)-capillary GC (cGC) analysis. The sensitivity and simplicity of SBSE has been demon- strated for the analysis of volatiles and semivolatiles in a variety of sample matrices like water [18], beverages [19], biological fluids [20], fruits and vegetables [21], etc. SBSE, however, can also be combined with liquid desorption (LD) and this opens perspectives for the analysis of thermally labile compounds. Popp et al. [22] used SBSE-LD for the determination by HPLC with fluorescence detection of polyaromatic hydrocarbons (PAHs) in water samples, Sandra et al. [23] used SBSE-LD to confirm SBSE-TD- cGC results for the determination of fungicides in wine by LC-MS, and Harms et al. [24] presented the first applica- tion of SBSE-LD for the profiling of iso-a-acids in beer. The features of SBSE have recently been reviewed [25]. In this contribution, the use of SBSE-LD for quantita- tive analysis of iso-a-acids and for the elucidation of reduced iso-a-acids in various beers by MEKC is de- scribed. The structures of the investigated compounds are presented in Fig. 1. The SBSE sorption and liquid de- sorption processes are discussed. The iso-a-acid content of several beers was determined and compared with results obtained with HPLC using direct beer injection. 2 Materials and methods 2.1 Chemicals and reagents Na 2 HPO 4 , HCl, and NaOH were from Merck (Darmstadt, Germany), sodium dodecyl sulfate (SDS) from BDH Chemicals (Poole, UK), and MgSO 4 from Saarchem Correspondence: Professor Dr. Pat Sandra, Research Institute for Chromatography, Kennedypark 20, B-8500 Kortrijk, Belgium E-mail: pat.sandra@richrom.com Fax: 132-(0)-56-204859 Abbreviations: LD, liquid desorption; PDMS, polydimethylsilox- ane; SBSE, stir bar sorptive extraction 664 Electrophoresis 2004, 25, 664–669  2004 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim