抄録
A rapid and sensitive tandem immunoaffinity column (IAC) cleanup and high performance liquid chromatography (IAC-HPLC) for the determination of deoxynivalenol (DON) in wheat and corn is described. A monoclonal antibody specific to DON conjugated to the CNBr- activated Sepharose 4B gel was used in construction of the IAC with a capacity of 2.5 μg DON per column. In the analysis, ground sample was extracted with acetonitrile-water (84+16) to which 4 ml (or less) were withdrawn and evaporated. The residue was dissolved in 0.01 M phosphate buffered saline, and subjected to the IAC. After washing with 10% methanol, DON was eluted from the IAC with 100% methanol, which was evaporated, redissolved in acetonitrile-water (2+8) and then subjected to HPLC using a reversed-phase C18 column and diode array detection (DAD). The analysis was completed in 6 min after injection of each sample with a detection limit of 0.03 μg DON/g. The chromatogram and the DAD spectrum for DON peak showed no interference substances in the sample treated with IAC. The average recoveries of DON added to wheat and corn extracts at levels from 0.5 to 3 μg/g were found to be 94.4 and 87.1%, respectively. Analysis of 6 naturally contaminated wheat samples (0.7 to 12 μg DON/g) showed good agreement between the IAC-HPLC data and those from direct competitive enzyme linked immunosorbent assay (dc-ELISA) (r=0.9970; slope=0.8533).
