Determination of Thiabendazole in Fruit Juices by a New Monoclonal Enzyme Immunoassay

Abstract

Abstract A competitive, indirect enzyme-linked immunosorbent assay (ELISA) for thiabendazole has been developed and applied to the analysis of fruit juices spiked with this fungicide. The immunoassay is based on a new monoclonal antibody derived from a hapten functionalized at the nitrogen atom in the 1-position of the thiabendazole structure. To our knowledge, such a structure has not been previously used to obtain antibodies to thiabendazole. The l50 value and the detection limit of the ELISA for standards were 0.2 and 0.05 ng/mL, respectively. Fruit juices were analyzed by diluting samples in assay buffer, without extraction or cleanup. Samples were not even centrifuged or filtered to remove fruit pulp. Under these conditions, the immunoassay was able to accurately determine thiabendazole down to 1 ng/mL in orange and grapefruit juices, down to 5 ng/mL in banana juice, and down to 20 ng/mL in apple and pear juices. Sensitivity differences of the ELISA were caused by the minimum dilution required by each juice to minimize matrix effects: 1/10 for orange and grapefruit juices, 1/50 for banana juice, and 1/100 for apple and pear juices. In an attempt to further increase the sensitivity of the immunoassay for matrixes showing the strongest interferences, apple and pear juices spiked with thiabendazole at low levels (1–20 ng/mL) were extracted with ethyl acetate before analysis. This simple procedure entailed a significant reduction of matrix effects, which in fact allowed us to determine accurately as low as 5 ng/mL thiabendazole in apple and pear juices. Irrespective of whether samples were analyzed by the direct dilution method or after extraction, the simplicity, sensitivity, and sample throughput of this monoclonal immunoassay makes it a very convenient method for the routine monitoring of thiabendazole residues in fruit juices.