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(1) Background: Aflatoxin contamination in food and grain poses serious problems both for economic development and public health protection, thus leading to a focus on an effective approach to control it; (2) Methods: Aflatoxin B₁ (AFB₁) degrading bacteria were isolated using a medium containing coumarin as the sole carbon source, and the biodegradation of AFB₁ by the isolate was examined by high performance liquid chromatography, and liquid chromatography mass spectrometry; (3) Results: a bacterial strain exhibiting strong AFB 1 degradation activity (91.5%) was isolated and identified as Bacillus  velezensis DY3108. The AFB₁ degrading activity was predominantly attributed to the cell-free supernatant of strain DY3108. Besides, it was heat-stable and resistant to proteinase K treatment but sensitive to sodium dodecyl sulfate treatment. The optimal temperature for the maximal degradation of AFB₁ was 80 °C. Even more notable, the supernatant showed a high level of activity over a broad pH (4.0 to 11.0) and exhibited the highest degradation (94.70%) at pH 8.0. Cytotoxicity assays indicated that the degradation products displayed significantly ( p &lt; 0.05) lower cytotoxic effects than the parent AFB₁; (4) Conclusions: B.  velezensis DY3108 might be a promising candidate for exploitation in AFB₁ detoxification and bioremediation in food and feed matrices.

Biological Degradation of Aflatoxin B1 by Cell-Free Extracts of Bacillus velezensis DY3108 with Broad PH Stability and Excellent Thermostability