Simultaneous determination of pesticides, mycotoxins, and metabolites as well as other contaminants in cereals by LC-LC-MS/MS

Abstract

A 2D LC-MS/MS method for the simultaneous determination of 350 pesticides, 16 mycotoxins as well as the growth regulators Chlormequat and Mepiquat was developed. The method is applicable to cereals and products thereof. Attention should be paid to the simultaneous analysis of the cereal-relevant mycotoxins aflatoxin B1, B2, G1 and G2, ochratoxin A, deoxynivalenol and zearalenone. Moreover, the tropane alkaloids atropine/scopolamine could be integrated into the final method. The samples were extracted with a mixture of acetonitrile/water (80:20), diluted with acetonitrile and injected into an LC-LC-MS/MS system. There were no further manual clean-up steps. The automatic online clean-up took place during the HILIC-separation in the first dimension (YMC-Pack Diol; 2.1 × 100 mm; 5 μm, 120 Å). Here, polar matrix compounds were retained, while the majority of the analyte scope eluted in a fraction at the beginning of the analytical run. This fraction was transferred to the second dimension by a packed loop interface (Agilent Zorbax SB-C8; 4.6 × 12.5 mm; 5 μm; 80 Å). On the second column (Phenomenex Synergi Fusion RP C18; 2 × 100 mm; 2.5 μm; 100 Å), the majority of the scope was separated by a typical RP-gradient. Only some of the polar pesticides could not be transferred to the second column. They eluted directly after the transfer step from the HILIC-column to the MS/MS. The final method was sensitive enough to meet all the regulated maximum levels for pesticides in cereals according to EU Regulation 396/2005 and those for contaminants according to EU Regulation 1881/2006. Above all, the method was so robust and accurate that nearly 90% of the pesticides and all the tested mycotoxins, growth regulators and tropane alkaloids fulfilled the validation criteria of the SANTE guideline document, although the demanding criteria are only applicable to pesticides. For the verification, eight proficiency tests were passed successfully: three for the pesticide analysis, three for the mycotoxin analysis, and two for the analysis of the tropane alkaloids. In addition to the already mentioned contaminants, the six most important ergot alkaloids (e.g. ergotamine/ergotaminine) and two modified mycotoxins (deoxynivalenol-3-glucoside and zearalenone-sulfate, also known as masked mycotoxins) were detected during the routine analysis of rye and corn samples.