Development and validation of a rapid multiplex ELISA for pyrrolizidine alkaloids and their N-oxides in honey and feed
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Pyrrolizidine alkaloids (PAs) are a group of plant secondary metabolites with carcinogenic and hepatotoxic properties. When PA-producing plants contaminate crops, toxins can be transferred through the food chain and cause illness in humans and animals, most notably hepatic veno-occlusive disease. Honey has been identified as a direct risk of human exposure. The European Food Safety Authority has recently identified four groups of PAs that are of particular importance for food and feed: senecionine-type, lycopsamine-type, heliotrine-type and monocrotaline-type. Liquid or gas chromatography methods are currently used to detect PAs but there are no rapid screening assays available commercially. Therefore, the aim of this study was to develop a rapid multiplex ELISA test for the representatives of three groups of alkaloids (senecionine, lycopsamine and heliotrine types) that would be used as a risk-management tool for the screening of these toxic compounds in food and feed. The method was validated for honey and feed matrices and was demonstrated to have a detection capability less than 25 μg/kg for jacobine, lycopsamine, heliotrine and senecionine. The zinc reduction step introduced to the extraction procedure allows for the additional detection of the presence of N-oxides of PAs. This first multiplex immunoassay for PA detection with N-oxide reduction can be used for the simultaneous screening of 21 samples for >12 PA analytes. Honey samples (n = 146) from various origins were analysed for PA determination. Six samples were determined to contain measurable PAs >25 μg/kg by ELISA which correlated to >10 μg/kg by LC-MS/MS.
KeywordsMultiplex ELISA Pyrrolizidine alkaloids Food and feed Honey Screening assay
The authors would like to thank the CONffIDENCE coordinator Dr. Jacob de Jong from RIKILT-Institute of Food Safety (Wageningen, the Netherlands) for his support of this research. The authors also want to acknowledge the support of Ir. Albert Swinkels from MasterLab B.V. a Nutreco company (Boxmeer, The Netherlands) CONffIDENCE partner, who provided the blank feed and honey, Dr Simon Haughey for feed samples and the many collaborators to Queen’s University who provided honey samples. CONffIDENCE is the acronym of the EU FP7 project “CONtaminants in Food and Feed: Inexpensive Detection for Control of Exposure” financially supported by the European Commission (grant agreement number 211326—collaborative project).
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