LC–MS/MS multi-method for mycotoxins after single extraction, with validation data for peanut, pistachio, wheat, maize, cornflakes, raisins and figs

Abstract

Mycotoxin analysis is usually carried out by high performance liquid chromatography after immunoaffinity column cleanup or in enzyme-linked immunosorbent assay tests. These methods normally involve determination of single compounds only. EU legislation already exists for the aflatoxins, ochratoxin A and patulin in food, and legislation will come into force for deoxynivalenol, zearalenone and the fumonisins in 2007. To enforce the various legal limits, it would be preferable to determine all mycotoxins by routine analysis in different types of matrices in one single extract. This would also be advantageous for HACCP control purposes. For this reason, a multi-method was developed with which 33 mycotoxins in various products could be analysed simultaneously. The mycotoxins were extracted with an acetonitrile/water mixture, diluted with water and then directly injected into a LC-MS/MS system. The mycotoxins were separated by reversed-phase HPLC and detected using an electrospray ionisation interface (ESI) and tandem MS, using MRM in the positive ion mode, to increase specificity for quality control. The following mycotoxins could be analysed in a single 30-min run: Aflatoxins B1, B2, G1 and G2, ochratoxin A, deoxynivalenol, zearalenone, T-2 toxin, HT-2 toxin, alpha-zearalenol, alpha-zearalanol, beta-zearalanol, sterigmatocystin, cyclopiazonic acid, penicillic acid, fumonisins B1, B2 and B3, diacetoxyscirpenol, 3- and 15-acetyl-deoxynivalenol, zearalanone, ergotamin, ergocornin, ergocristin, alpha-ergocryptin, citrinin, roquefortin C, fusarenone X, nivalenol, mycophenolic acid, alternariol and alternariol monomethyl ether. The limit of quantification for the aflatoxins and ochratoxin A was 1.0 microg kg(-1) and for deoxynivalenol 50 microg kg(-1). The quantification limits for the other mycotoxins were in the range 10-200 microg kg(-1). The matrix effect and validation data are presented for between 13 and 24 mycotoxins in peanuts, pistachios, wheat, maize, cornflakes, raisins and figs. The method has been compared with the official EU method for the determination of aflatoxins in food and relevant FAPAS rounds. The multi-mycotoxin method has been proven by the detection of more than one mycotoxin in maize, buckwheat, figs and nuts. The LC-MS/MS technique has also been applied to baby food, which is subject to lower limits for aflatoxin B1 and ochratoxin A, ergot alkaloids in naturally contaminated rye and freeze-dried silage samples.

Human exposure to mycotoxins and their masked forms through cereal-based foods in Belgium

“…T-2 (1 mg, solid standard) and DON3G (50.2 ng µL -1 , in acetonitrile) were purchased from Biopure Referenzsubstanzen GmbH (Tulln, Austria). All mycotoxin solid standards were dissolved in methanol (1 mg mL -1 ) and storable for a maximum of 1 year at -18 °C (Spanjer et al, 2008). The DON3G-solution was kept at 4 °C.…”

Fusarium head blight and associated mycotoxin occurrence on winter wheat in Luxembourg in 2007/2008

“…Mycotoxin analysis was based on the procedure described by Spanjer et al (2008), but modified as follows. For mycotoxin assays, 500 g of wheat grains per field were dried for 48 h at 30 C for humidity standardization and aliquots (200 g) were obtained by milling with a Cyclotec TM 1093 (Foss, Belgium).…”

Scientific Opinion on the risk for public and animal health related to the presence of sterigmatocystin in food and feed

“…Liquid chromatography-tandem mass spectrometry (LC-MS/MS) multi-mycotoxin methods that include STC are frequently based on a single extraction step for all investigated mycotoxins (e.g., acetonitrile/water mixture) followed by the direct analysis of the diluted crude extract (Sulyok et al, 2007;Spanjer et al, 2008). This so-called dilute-and-shoot approach requires very sensitive analytical instruments and often suffers from considerable matrix effects.…”