03/10/2016
Determination of Phomopsin A in Lupin-Containing Food, Using UHPLC-MS/MS
Eric Eichhorn , Stefanie Marschik , Chemisches und Veterinäruntersuchungsamt (CVUA) Stuttgart
Phomopsin A, a secondary metabolite of the mold Diaporthe toxica, showed severe toxic effects in animal studies with potential relevance for human health. This lead to the establishment of a legal maximum amount for phomopsin A of 5 µg/kg in lupin seeds and their products by the Australian and New Zealand Food Authority (2001) as well as by the Food and Agriculture Organisation (2004).
We therefore developed a sensitive method for the determination of phomopsin A in lightly processed matrices like lupin flour with a limit of quantification (LOQ) of 1.4 µg/kg. The method involves a simple acetonitrile/methanol/water-extraction (3/1/1, v/v/v) without any clean-up step ("dilute & shoot"), followed by UHPLC-MS/MS analysis. Five matrices (lupin flour, lupin steak, lupin coffee, soy flour and wheat flour) were validated in accordance with the criteria set forth in EU-guideline SANCO/12495/2011 for the presented method. Sufficient recoveries and variation coefficients (cv) for lupin flour at the spiking levels of 1.4 µg/kg (LOQ) with 73 % (cv 4.9 %) and 7.1 µg/kg (5-fold LOQ) with 75 % (cv 4.3 %) were obtained. Wheat flour also showed sufficient results regarding the recovery and the LOQ, while unsatisfactory LOQ's were obtained for lupin steak, lupin coffee and soy flour.
A market survey of twenty-five relevant foods showed that no current contamination of European food with phomopsin A seems to be detectable.