Biocatalytic Detoxification of Ochratoxins A/B by a Fungal Dye-Decolorizing Peroxidase: Mechanistic Insights and Toxicity Assessment

Mycotoxin contamination in agricultural products poses severe global health risks, with ochratoxins (particularly OTA and OTB) exhibiting marked nephrotoxicity and classified as Group 2B carcinogens by IARC. Conventional physical/chemical detoxification methods often impair food nutritional quality, highlighting the need for enzymatic alternatives. Herein, we systematically investigated the degradation mechanisms of ochratoxin A (OTA) and ochratoxin B (OTB) using Pleurotus ostreatus dye-decolorizing peroxidase (PoDyP4) coupled with redox mediators. Remarkably, hydroxybenzotriazole (HBT) enhanced degradation efficiency 26.7-fold for OTA and 10.6-fold for OTB compared to mediator-free systems, establishing it as the optimal catalytic enhancer. Through LC-MS/MS analysis, we identified five key degradation products, including 6-OH-OTA and OTB-quinone, elucidating a putative oxidative degradation pathway. In vitro cytotoxicological evaluation in HK-2 cells demonstrated that PoDyP4-treated ochratoxins significantly attenuated cytotoxicity, reducing malondialdehyde (MDA) levels by 48.7% (OTA) and 42.3% (OTB) (p < 0.01) and suppressing ROS generation. Molecular docking revealed strong binding affinities between PoDyP4 and ochratoxins, with calculated binding energies of -7.6 kcal/mol (OTA) and -8.6 kcal/mol (OTB), stabilized by hydrogen bond networks (1.9-3.4 Å). These findings position PoDyP4 as a promising biocatalyst for mycotoxin mitigation in food systems, offering a sustainable alternative to traditional detoxification methods.