Development of an analytical method for detecting nitrofurans in bee pollen by liquid chromatography-electrospray ionization tandem mass spectrometry

Bee pollen collected by honeybees, which is in powdered form, is a good nutritional supplement. Nitrofuran antibiotics are assumed not to be present in bee pollen, which is important as the level of antibiotics in bee pollen is strongly regulated in many countries. A liquid chromatography-tandem mass spectrometry (LC-MS/MS) method to detect nitrofurans in honey has been developed, but this method is not suitable for bee pollen because of it being in powdered form. During preparation of bee pollen samples, the dispersal of powder particles in an aqueous solution often makes them susceptible to forming an emulsion with solvent components such as hexane and ethyl acetate. This may reduce the reproducibility and sensitivity of analyses of nitrofuran levels in bee pollen. Therefore, we attempted to optimize the sample preparation conditions to detect nitrofurans in bee pollen by determining three nitrofuran residues, namely, 3-amino-2-oxazolidinone (AOZ), 3-amino-5-methyl-morpholino-2-oxazolidinone (AMOZ), and 1-aminohydantoin (AHD), using LC-MS/MS. The optimized method prevented the formation of powder-induced emulsion. To verify the reproducibility and sensitivity of this method, it was validated using nitrofuran-free bee pollen spiked with analytes with different side chains at 1.0, 2.0, and 5.0 μg kg−1. The accuracy levels were 94.1%-104.0% and the coefficients of variation were less than 12%. The limits of detection for AOZ, AMOZ, and AHD were 0.18, 0.25, and 0.30 μg kg−1, respectively, while their limits of quantitation were 0.59, 0.83, and 1.00 μg kg−1. The LC-MS/MS method developed to analyze nitrofuran in bee pollen should contribute to the quality control of bee pollen and food safety.