Biomarker-Based Evaluation of a Zearalenone-Degrading Enzyme in Broilers and Piglets Across Multiple Biological Matrices

Zearalenone (ZEN) is an estrogenic mycotoxin that impairs animal health and productivity, necessitating effective mitigation strategies in livestock production. This study evaluated the efficacy of the ZEN lactonase ZenA, an enzyme that converts ZEN to non-estrogenic hydrolyzed ZEN (HZEN) and decarboxylated HZEN (DHZEN). Broilers were fed either uncontaminated feed, feed contaminated with 1500 µg ZEN/kg, or ZEN-contaminated feed supplemented with 20 U ZenA/kg for 35 days. Piglets received 200 µg ZEN/kg feed, with or without 10 U ZenA/kg, for 43 days. ZEN biomarkers (ZEN, α-zearalenol, β-zearalenol, HZEN, and DHZEN) were quantified in plasma, urine, feces/excreta, and gastrointestinal contents using liquid chromatography–tandem mass spectrometry. While performance parameters remained unaffected, ZenA supplementation significantly reduced ZEN concentrations (by 19.6–66.2%) in all matrices and at all time points in both species. In addition, significant formation of HZEN was observed in gastrointestinal samples. Thus, in the present study, ZenA efficiently degraded ZEN in both broilers and piglets. Biomarker analysis in multiple matrices provided complementary insights: gastrointestinal samples confirmed the enzyme’s mode of action, while plasma and urine data showed a marked reduction in systemic ZEN exposure. Finally, the results reinforce that performance parameters are insufficient for assessing the efficacy of mycotoxin-detoxifying feed additives and support biomarker-based evaluation approaches.