C26 | Emerging mycotoxins in the pig production chain: development and validation of a multi-residue UHPLC-HRMS method for detection in pork, liver and pig feed

P. Lorusso¹, A. Manfredi¹, L.M. Chiesa², S. Panseri², M. Nobile², S. Ghidini², A. Di Pinto¹, E. Bonerba¹. | ¹Dipartimento di Medicina Veterinaria, Università degli Studi di Bari Aldo Moro, Valenzano; ²Dipartimento di Medicina Veterinaria e Scienze Animali, Università degli Studi di Milano, Lodi.

Purpose. This study is part of the growing scientific interest in emerging mycotoxins, a group of contaminants that are not yet regulated but are increasingly detected in food and feed. The incidence of these secondary metabolites, produced by different fungal species, is increasing due to climatic and environmental changes that favour their formation and co-presence with known mycotoxins, leading to synergistic toxicological interactions that are still poorly characterised. The European Commission recently adopted EU Recommendation 2022/553 on the monitoring of Alternaria mycotoxins in food, which urges Member States to monitor these toxins in food of plant origin. However, the presence of emerging mycotoxins in food of animal origin is still documented in few cases, mainly due to the lack of validated analytical methods for their identification and quantification. The aim of the study was therefore to develop and validate an analytical method for the simultaneous determination of seven emerging mycotoxins in pork, liver and pig feed, in order to improve monitoring and risk assessment activities in the pig production chain. Methods. A method based on ultra-high-performance liquid chromatography coupled with high-resolution mass spectrometry (UHPLC-HRMS) was developed for the simultaneous quantitative determination of: beauvericin, enniatin A, A1, B, B1, alternariol and alternariol monomethyl ether. The clean-up phase from meat, liver and complete feed for fattening pigs was carried out by solid-liquid extraction to optimise the efficiency, time and cost of the analysis, while ensuring effective removal of matrix interferents. The method was validated according to the criteria established by Implementing Regulation (EU) 2023/2782, including the definition of the limit of quantification (LOQ), the percentage recovery and intra-day precision (repeatability) and inter-day precision (reproducibility). Results. The LOQ is 1 ng g⁻¹ for all mycotoxins analysed, both in pork meat and liver and in feed. The percentage recoveries obtained from fortified samples ranged from 72% to 88% for pork meat and liver and from 73% to 120% for feed. The intra- and inter-day precision values are less than 14% for all matrices investigated. The results obtained confirm the reliability and robustness of the method for the simultaneous analysis of emerging mycotoxins in complex matrices, both of animal and plant origin. Conclusions. The development of a sensitive, selective and fully validated analytical method for the simultaneous detection of emerging mycotoxins in matrices of animal origin represents a significant step forward in the monitoring and characterisation of risk along the pig production chain. This method provides robust and reliable data on the presence and distribution of toxins that are still few in study, supporting the development of regulatory strategies based on solid scientific evidence and clarifying the potential carry-over in the food chain. The future extension of the method's application to other species and matrices could further enhance risk assessment and contribute to the development of safer, more sustainable and resilient agri-food systems.