Development and validation of a liquid chromatography/tandem mass spectrometric method for the determination of 39 mycotoxins in wheat and maize
ABSTRACT This paper describes the first validated method for the determination of 39 mycotoxins in wheat and maize using a single extraction step followed by liquid chromatography with electrospray ionization triple quadrupole mass spectrometry (LC/ESI-MS/MS) without the need for any clean-up. The 39 analytes included A- and B-trichothecenes (including deoxynivalenol-3-glucoside), zearalenone and related derivatives, fumonisins, enniatins, ergot alkaloids, ochratoxins, aflatoxins and moniliformin. The large number and the chemical diversity of the analytes required the application of the positive as well as the negative ion ESI mode in two consecutive chromatographic runs of 21 min each. The solvent mixture acetonitrile/water/acetic acid 79 + 20 + 1 (v/v/v) has been determined as the best compromise for the extraction of the analytes from wheat and maize. Raw extracts were diluted 1 + 1 and were injected without any clean-up. Ion-suppression effects due to co-eluting matrix components were negligible in the case of wheat, whereas significant signal suppression for 12 analytes was observed in maize, causing purely proportional systematic errors. Method performance characteristics were determined after spiking blank samples on multiple levels in triplicate. Coefficients of variation of the overall process of <5.1% and <3.0% were obtained for wheat and maize, respectively, from linear calibration data. Limits of detection ranged from 0.03 to 220 microg/kg. Apparent recoveries (including both the recoveries of the extraction step and matrix effects) were within the range of 100 +/- 10% for approximately half of the analytes. In extreme cases the apparent recoveries dropped to about 20%, but this could be compensated for to a large extent by the application of matrix-matched standards to correct for matrix-induced signal suppression, as only a few analytes such as nivalenol and the fumonisins exhibited incomplete extraction. For deoxynivalenol and zearalenone, the trueness of the method was confirmed through the analysis of certified reference materials.
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ABSTRACT: Using micro high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS) a simple and fast method for the quantitative determination of 26 mycotoxins was developed. Sample preparation consists of a single extraction step and a dilute-and-shoot approach without further cleanup. With a total run time of 9 min and solvent consumption below 0.3 mL per chromatographic run, the presented method is cost-effective. All toxins regulated by the European Commission with maximum or guidance levels in grain products (fumonisins B1 and B2 (FB1 and FB2)); deoxynivalenol (DON); aflatoxins B1, G1, B2, and G2 (AFB1, AFG1, AFB2, and AFG2); ochratoxin A (OTA); T-2 and HT-2 toxins; and zearalenone (ZEN) can be quantified with this method. Furthermore, the enniatins B, B1, A, and A1 (EnB, EnB1, EnA, and EnA1); beauvericin (BEA); 3-acetyl-deoxynivalenol (3-AcDON); fusarin C (FusC); sterigmatocystin (STC); gliotoxin (GT); and the Alternaria toxins alternariol (AOH), alternariol monomethyl ether (AME), altenuene (ALT), tentoxin (TEN), and altertoxin I (ATX I) can also be quantified. For all regulated compounds, recoveries ranged between 76 and 120 %. For all other toxins, the recovery was at least 51 %. The method was applied for the analysis of 42 maize samples from field trials in South Africa.Mycotoxin Research 03/2015; 31(2). DOI:10.1007/s12550-015-0221-y
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ABSTRACT: Having entered the Genomic Era, it is now evident that the biosynthetic potential of filamentous fungi is much larger than was thought even a decade ago. Fungi harbor many cryptic gene clusters encoding for the biosynthesis of polyketides, non-ribosomal peptides, and terpenoids – which can all undergo extensive modifications by tailoring enzymes – thus potentially providing a large array of products from a single pathway. Elucidating the full chemical profile of a fungal species is a challenging exercise, even with elemental composition provided by high-resolution mass spectrometry (HRMS) used in combination with chemical databases (e.g., AntiBase) to dereplicate known compounds.This has led to a continuous effort to improve chromatographic separation in conjunction with improvement in HRMS detection. Major improvements have also occurred with 2D chromatography, ion-mobility, MS/MS and MS 3 , stable isotope labeling feeding experiments, classic UV/Vis, and especially automated data-mining and metabolomics software approaches as the sheer amount of data generated is now the major challenge. This review will focus on the development and implementation of dereplication strategies and will highlight the importance of each stage of the process from sample preparation to chromatographic separation and finally toward both manual and more targeted methods for automated dereplication of fungal natural products using state-of-the art MS instrumentation.Frontiers in Microbiology 02/2015; 6(71). DOI:10.3389/fmicb.2015.00071 · 3.94 Impact Factor
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ABSTRACT: A multi-mycotoxin method based on liquid chromatography/tandem mass spectrometry (LC-MS/MS) was used for a mycotoxin survey in 627 samples of processed cassava collected from different districts across Tanzania and Rwanda after the method performance for this matrix had been determined. Matrix effects as well as extraction efficiencies were found to be similar to most other previously investigated matrices with the exception of distinct matrix effects in the negative ionization mode for early eluting compounds. Limits of detection were far below the regulatory limits set in the EU for other types of commodities. Relative standard deviations were generally lower than 10% as determined by replicates spiked on two concentration levels. The sample-to-sample variation of the apparent recoveries was determined for 15 individually spiked samples during three different analytical sequences. The related standard deviation was found to be lower than 15% for most of the investigated compounds, thus confirming the applicability of the method for quantitative analysis. The occurrence of regulated mycotoxins was lower than 10% (with the exception of zearalenone) and the related limits were exceeded only in few samples, which suggests that cassava is a comparatively safe commodity as regards mycotoxins. The most prevalent fungal metabolites were emodin, kojic acid, beauvericin, tryptophol, 3-nitropropionic acid, equisetin, alternariol methylether, monocerin, brevianamide F, tenuazonic acid, zearalenone, chrysophanol, monilifomin, enniatins, apicidin and macrosporin. The related concentrations exceeded 1 mg/kg only in few cases. However, extremely high levels of cyanogenic plant toxins, which had been previously added to the method, were observed in few samples, pointing out the need for improved post-harvest management to decrease the levels of these compounds.Food Additives and Contaminants - Part A Chemistry, Analysis, Control, Exposure and Risk Assessment 10/2014; 32(4). DOI:10.1080/19440049.2014.975752 · 2.34 Impact Factor