Patulin is one of the most important mycotoxins that contaminates fruit-derived products and causes acute or chronic toxicity in humans. In the present study, a novel patulin-degrading enzyme preparation was developed by taking a short-chain dehydrogenase/reductase and covalently linking it to dopamine/polyethyleneimine co-deposited magnetic Fe₃O₄ particles. Optimum immobilization provided 63% immobilization efficiency and 62% activity recovery. Moreover, the immobilization protocol substantially improved thermal and storage stabilities, proteolysis resistance, and reusability. Using reduced nicotinamide adenine dinucleotide phosphate as a cofactor, the immobilized enzyme exhibited a detoxification rate of 100% in phosphate-buffered saline and a detoxification rate of more than 80% in apple juice. The immobilized enzyme did not cause adverse effects on juice quality and could be magnetically separated quickly after detoxification to ensure convenient recycling. Moreover, it did not exhibit cytotoxicity against a human gastric mucosal epithelial cell line at a concentration of 100 mg/L. Consequently, the immobilized enzyme as a biocatalyst had the characteristics of high efficiency, stability, safety, and easy separation, establishing the first step in building a bio-detoxification system to control patulin contamination in juice and beverage products.