QUANTITATIVE ANALYSIS OF THE BACILLUS CEREUS EMETIC TOXIN, CEREULIDE, USING MICELLAR ELECTROKINETIC CHROMATOGRAPHY‐CAPILLARY ELECTROPHORESIS

Abstract

ABSTRACT A micellar electrokinetic chromatography capillary electrophoresis (MEKC‐CE) method was developed for quantitative analysis of cereulide, the emetic toxin produced by Bacillus cereus. Valinomycin, a highly hydrophobic cyclic dodecadepsipeptide with a chemical structure similar to that of cereulide, was used as a surrogate for method development. A series of buffer systems and electrophoretic conditions were investigated and optimized. The final method, yielding good peak resolution and reproducibility of analysis, utilized 20 mM borate buffer, pH 8.5, containing 75 mM SDS. The capillary had a 50 µm internal diameter and 36.4 cm effective length, and was maintained at 25C throughout column preparation and analysis. The sample compartment was maintained at 22C. A constant voltage of 15 kV was applied, after hydrodynamic injection of the sample at 34.5 KPa for 2 s. Detection used ultraviolet light, at 195 nm. A calibration curve, using peak area rather than peak height, was obtained with purified cereulide, with linearity between 2 and 30 ng/µL.PRACTICAL APPLICATIONSDetection of the emetic toxin cereulide, as well as enumeration of B. cereus, is important for determining the risk of emetic food poisoning. The novel capillary electrophoresis method developed in this study, employing the principles of MEKC‐CE, can be used for the quantitative analysis of cereulide. MEKC‐CE gives results quickly and simply, and offers advantages over reversed phase high performance liquid chromatography (RP‐HPLC), through simple sample handling and preparation, less sample required and lower cost of operation. After calibration with purified cereulide was obtained, the applicability of the method to model and real food samples was evaluated with rice samples inoculated with an emetic B. cereus strain and samples from an outbreak of emetic food poisoning. The results showed that the method can be successfully applied to both semi‐purified toxin extracts of cultures and food samples.