Immunoassays of Selected Mycotoxins in Hay, Silage and Mixed Feed

Abstract

Immunoassays, including radioimmunoassay (RIA) and direct competitive enzyme-linked immunosorbent assays (dc-ELISAs) were used for the analysis of the mycotoxins, produced by Alterneria alternata , AAL TA toxin (AAL), cyclopiazonic acid (CPA), deoxynivalenol (DON), fumonisin B1 (FmB1), PR toxin (PRT) and zearalenone (ZEA) in hay, silage and mixed feed. In the analysis, 10 g well-ground feed samples in 100 ml of 84% acetonitrile in water were shaken in an orbital shaker for 1 h. After filtration, the filtrates were diluted with buffer and directly subjected to dc-ELISA for AAL toxin, CPA, FmB1, PRT and ZEA. For total DON, including 15-acetyldeoxynivalenol (15-AcDON) and 3-acetyldeoxynivalenol (3-AcDON), the extracts were air-dried, acetylated, and subjected to both RIA and dc-ELISA using the Ridascreen DON enzyme immunoassay (EIA) kit. The average recoveries of toxins added to the hay at 0.3 and 1.0 ppm levels were found to be 77.5, 111, 95.8, 85.7, 95.5 and 138% for AAL toxin, DON, CPA, FmB1, PRT and ZEA, respectively. When the sample extracts were subjected to a C-18 reversed-phase cartridge treatment prior to dc-ELISA, the recovery of ZEA at 0.1 and 1.0 ppm levels was found to be 111 and 75%, respectively. Analysis of 63 samples (25 hay and 38 others, including corn silage and mixed feed) showed that the frequencies of AAL toxin, DON, CPA, FmB1, PRT and ZEA in the feed were 96.8, 100, 87.3, 36.5, 76.2, and 3.2%, respectively. The overall average concentrations of toxins in the feed samples, excluding the nondetectable, were 0.56, 0.73, 0.34, 0.28, 0.13 and 0.22 microg g -1 (ppm) for AAL toxin, DON, CPA, FmB1, PRT and ZEA, respectively. For hay alone, the frequencies were 100% (average level of 0.72 ppm), 100% (average level of 0.61 ppm), 80% (0.39 ppm), 52% (0.12 ppm) and 80% (0.15 ppm) for AAL, DON, CPA, FmB1 and PRT, respectively. ZEA was not found in the hay samples. Linear regression analysis between the results of DON in 30 feed samples analyzed simultaneously with RIA and EIA showed agreement, with a correlation coefficient of 0.80 and a slope of 1.34.