Development of a sensitive indirect competitive enzyme-linked immunosorbent assay for high-throughput detection and risk assessment of aflatoxin B1 in animal-derived medicines.

Animal-derived medicine is an important part of traditional Chinese medicine (TCM). Studies have shown that many animal-derived medicinal products are susceptible to contaminate of aflatoxins, nevertheless, the rapid detection for animal-derived medicine is prone to be ignored. Here we developed a sensitive indirect competitive enzyme-linked immunosorbent assay (icELISA) for rapid screening of aflatoxin B1 (AFB1) in ground beetle, cockroach, silkworm and earthworm. The sensitivity of the icELISA method was significantly enhanced. The IC50 for the four animal-derived medicinal samples ranged from 0.092 to 0.135 ng·mL-1; the limit of detection (LOD) was 0.008 - 0.020 ng·mL-1. To obtain high accuracy, the extraction solution and time were evaluated. By using this method, a total of 138 samples were investigated, and the detection rates of AFB1 in ground beetle and earthworm samples were 26.6% and 16.7%, respectively. The result was validated by liquid chromatography combined with tandem mass spectrometry, and an excellent correlation was observed between the two datasets, with a R2 value of 0.999. Our results indicate that the proposed method can be used for the rapid detection of AFB1 in animal-derived medicine. Furthermore, the quantitative risk assessment was conducted for ground beetle and earthworm based on the results, demonstrating that the intake of AFB1 in ground beetle had a slight threat to the risk of cancer.