Fluorescence polarization immunoassay for rapid detection of T-2 and HT-2 toxins in cereals and cereal-based products

T-2 toxin (T-2) and HT-2 toxin (HT-2) are type A trichothecene mycotoxins produced by several Fusarium species, mainly Fusarium sporotrichioides, Fusarium langsethiae and Fusarium poae. Generally, these Fusarium species can grow on cereals and produce T-2 and HT-2 under moist cool conditions already prior to harvesting [1]. Among cereals, oats, wheat, rye and derived products are key dietary sources of T-2 and HT-2 exposure [1]. Fluorescence polarization (FP) immunoassay is a homogeneous technique that is getting attention as a screening tool in food safety control due to its simplicity, rapidity, cheapness and reliability. A rapid, sensitive and reliable FP immunoassay has been recently reported for the determination of the sum of T-2 and HT- 2 toxins in wheat [2]. The aim of present work was to evaluate the applicability of the above-mentioned FP immunoassay to other unprocessed cereals, such as rye, and cereal-based products, such as oats crispbread, for the quantitative determination of the total content of T-2 and HT-2. The concept of determining the total content of T-2 and HT-2 in cereal samples for both official control purposes and risk assessment studies results in line with the EC Recommendation [3]. No purification step of extracts was required, although in order to reduce the matrix effect a dilution step with NaCl solution (4 % for rye and 1% for oats crispbread), in a ratio 1:5 (v/v), was necessary to let precipitation of proteins and matrix interfering compounds. For the optimized FP immunoassay, LOD of 0.20 ng/mL (equivalent to 20 ?g/kg in rye and oats crispbread) was calculated. Overall mean recoveries of the optimized FP immunoassay were 105 and 107% for rye and oats crispbread, respectively, with relative standard deviations lower than 4%. The analytical performances of the optimized FP immunoassay in terms of accuracy and precision fulfill the criteria established by the European Commission [4]. In addition, a comparative analysis of the levels of contamination in spiked and blank samples was performed by both FP immunoassay and UHPLC method. In particular, a total of 30 rye and oats crispbread samples, of which 20 spiked samples at levels from 50 to 700 ?g/kg and 10 uncontaminated samples, were analyzed. The proposed method coupled performances in terms of sensitivity, accuracy and precision comparable to those of a chromatographic technique with rapidity (20 min), costs and simplicity typical of a high-throughput screening method and can be used as a valid alternative to more expensive and time-consuming LC methods for quantitative determination in rye and oats crispbread [5]. ACKNOWLEDGMENTS This work has been supported by the Italian Ministry of Education, University and Research (MIUR) project no. CTN01_00230 CL.A.N. Cluster Tecnologici Nazionali - SAFE&SMART project "New enabling technologies for food safety and food chain integrity within a global scenario". REFERENCES [1] EFSA Panel on Contaminants in the Food Chain Scientific opinion on the risks for animal and public health related to the presence of T-2 and HT-2 toxin in food and feed, vol. 12, pp. 2481-2668, 2011 (available online: www.efsa.europa.eu/ efsajournal). [2] Lippolis V, Pascale M, Valenzano S, Pluchinotta V, Baumgartner S, Krska R, Visconti A, "A rapid fluorescence polarization immunoassay for the determination of T-2 and HT-2 toxins in wheat", Analytical and Bioanalytical Chemistry, vol. 401, pp. 2561-2571, Sept. 2011. [3] European Commission, "Commission recommendation 165/2013/ UE of 27 March 2013 on the presence of T-2 and HT-2 toxin in cereals and cereal products", Official Journal of the European Union L91:12-15, March 2013. [4] European Commission, "Commission Regulation 401/2006/EC of 23 February 2006 laying down the methods of sampling and analysis for the official control of the levels of mycotoxins in foodstuffs", Official Journal of the European Union, L70:12-34, Feb. 2006. [5] Porricelli ACR, Lippolis V, Valenzano S, Cortese M, Suman M, Zanardi S, Pascale M, "Optimization and validation of a fluorescence polarization immunoassay for rapid detection of T-2 and HT-2 toxins in cereals and cereal-based products", Food Analytical Methods, DOI 10.1007/s12161-016-0527-1.