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An LC-MS/MS method was developed and validated for the simultaneous determination of deoxynivalenol, zearalenone, T-2-toxin, HT-2-toxin and metabolites, including 3-acetyldeoxynivalenol, 15-acetyldeoxynivalenol, deoxynivalenol-3-glucoside, α-zearalenol, β-zearalenol, zearalenone-4-glucoside, α-zearalenol-4-glucoside, β-zearalenol-4-glucoside and zearalenone-4-sulfate in maize, wheat, oats, cornflakes and bread. Extraction was performed with acetonitrile/water/acetic acid (79/20/1, v/v/v) followed by a hexane defatting step. After filtration, the extract was evaporated and the residue was redissolved in mobile phase for injection. The mobile phase, which consisted of a mixture of methanol and water with 10 mM ammonium acetate, was adjusted to pH 3 with glacial acetic acid. A sample clean-up procedure was not included because of the low recoveries of free and masked mycotoxins and their differences in polarity. The method allowed the simultaneous determination of 13 Fusarium mycotoxins in a one-step chromatographic run using a Waters Acquity UPLC system coupled to a Quattro Premier XE mass spectrometer. The method was validated for several parameters such as linearity, apparent recovery, limit of detection, limit of quantification, precision, expanded measurement uncertainty and specificity. The limits of detection varied from 5 to 13 ng g−1; those for the limit of quantification from 10 to 26 ng g−1. The results of the performance characteristics of the developed LC-MS/MS method were in good agreement with the criteria mentioned in Commission Regulation (EC) No. 401/2006. Thirty samples of a variety of food and feed matrices were sampled and analysed between July 2010 and January 2011. [ABSTRACT FROM PUBLISHER]

A collection of 84 cereal-based food products in 25 composites, including beer, was screened for the presence of deoxynivalenol, zearalenone, and their respective metabolites deoxynivalenol-3-glucopyranoside, 3-acetyl-deoxynivalenol, zearalenol-4-glucopyranoside, α-zearalenol, β-zearalenol, α-zearalenol-4-glucopyranoside, β-zearalenol-4-glucopyranoside, and zearalenone-4-sulfate. The most abundant analyte was zearalenone-4-sulfate, which was found in 13 composites, albeit in low concentrations. Furthermore, deoxynivalenol was detected in eight, zearalenone in seven, and deoxynivalenol-3-glucopyranoside in two composites. None of the remaining six analytes was found in any matrices, which suggests that, if at all present, the concentrations of these latter metabolites are very low and, hence, do not impose any danger to consumers. The highest mycotoxin content was found in bran flakes with 254 ng g−1 deoxynivalenol, 6 ng g−1 zearalenone-4-sulfate, and 44 ng g−1 zearalenone. [ABSTRACT FROM AUTHOR]

Deoxynivalenol-3-β-D-glucoside (D3G), a phase II plant metabolite of the mycotoxin deoxynivalenol (DON), occurs in naturally Fusarium-contaminated cereals. In order to investigate the frequency of occurrence as well as the relative and absolute concentrations of D3G in naturally infected cereals, 23 wheat samples originating from fields in Austria, Germany and Slovakia as well as 54 maize samples from Austrian fields were analysed for DON and D3G by liquid chromatography-tandem mass spectrometry (LC-MS/MS). Both analytes were detected in all the 77 field samples. DON was found at levels from 42 to 4130 ng g-1 (977 ± 1000 ng g-1 on average). The D3G concentrations in all cereal samples were in the range 10-1070 ng g-1 (216 ± 253 ng g-1 on average), corresponding to about 5-46 mol% of their DON concentrations (15 ± 8 mol% on average). [ABSTRACT FROM AUTHOR]

Glucosides of several Fusarium mycotoxins occur in naturally infected cereals and may contribute to an increased content to the total mycotoxin load of food and feed. The paper presents the results of a fermentation procedure to produce zearalenone-4O-β-D-glucopyranoside from zearalenone using an engineered Saccharomyces cerevisiae strain, expressing the Arabidopsis thaliana UDP-glucosyltransferase UGT73C6. About 24 mg of zearalenone-4O-β-D-glucopyranoside was obtained from 50 mg of zearalenone and further purified. A total of 10 mg of the glucoside were reduced with sodium borohydride, yielding 4.1 mg α-zearalenol-4O-β-D-glucopyranoside and 4.5 mg β-zearalenol-4O-β-D-glucopyranoside at purities higher than 99%. To confirm the identities of the three produced glucosides, MS and MS/MS spectra were acquired using negative electrospray ionization. Besides the deprotonated ions at m/z 479 or 481, respectively, in full-scan mode, fragments, adducts, and dimers were recorded and assigned. MS/MS spectra of the glucosylated substances yielded the deprotonated ions of the mycotoxins zearalenone, α-zearalenol, β-zearalenol and their fragments, respectively. Unambiguous structural assignment of the three substances was achieved using two-dimensional NMR methods. This way, the glucose attachment to position C-4, the β-configuration of the sugar unit and the stereo-chemical assignment of the zearalenol hydroxyl group at C-6' were proven. [ABSTRACT FROM AUTHOR]

Concentrations of fungal metabolites were measured in 32 wheat grain samples from north-eastern Poland in 2003. The samples originated from fields cultivated conventionally (but varying in chemical protection level) or cultivated organically. Concentrations of Fusarium toxins (HT-2, DON, 3-AcDON, NIV), trichodiene, microbial biomass indicators (fungal ergosterol and general adenosine 5'-triphosphate (ATP)) and seed vigour were assessed. A large variation between samples was observed, depending on their origin. Seed from organic farms contained similar amounts of Fusarium toxins but more ergosterol and ATP than conventionally grown and chemically protected seed. The highest levels of toxins and ergosterol were detected in samples from conventional cultivation lacking chemical protection. Intensive agronomic practices (including complete chemical protection) significantly lowered the levels of ergosterol, ATP and trichodiene, as compared with other cultivation systems. [ABSTRACT FROM AUTHOR]

The biotransformation products of zearalenone, a Fusarium mycotoxin, were elucidated using the model plant Arabidopsis thaliana. After treatment of plant seedlings with 50 µM zearalenone, both the liquid media and the plant extracts were analysed by liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS). An array of 17 different metabolites, most prominently glucosides, malonylglucosides, di-hexose- and hexose–pentose disaccharides of zearalenone, and α- and β-zearalenol, were detected in the samples. Time courses for the different zearalenone metabolites were recorded and they give a closer insight into the metabolism kinetics. A scheme proposing the zearalenone metabolism in A. thaliana is given. The aspect of food safety regarding the (potential) occurrence of masked mycotoxins in agricultural commodities is discussed. [ABSTRACT FROM AUTHOR]

The powerful combination of liquid chromatography and mass spectrometry (MS) is often limited by matrix effects during ionization in the MS ion source. The use of fully isotope-substituted (13C15)-deoxynivalenol ((13C15)-DON) as an internal standard (IS) corrects matrix effects and improves the accuracy of analytical methods using mass spectrometry for the quantitative determination of the Fusarium mycotoxin deoxynivalenol (DON). The IS was characterized with respect to its chromatographic purity by liquid chromatography-ultraviolet light and its isotope distribution by time-of-flight mass spectrometry. Its low-energy collision-induced dissociation behaviour was compared with DON. Moreover, this work describes the successful application of (13C15)-DON as IS for the determination of DON in maize using high-performance liquid chromatography (HPLC) electrospray (ESI) with tandem mass spectrometry. The results demonstrate that the IS can successfully correct for fluctuations during extraction and clean-up of the sample as well as the ionization of DON in the MS ion source. Random variations in ionization affect the IS in the same way as the analyte. Recoveries for DON in maize of 76% ± 1.9% (external calibration) or 101% ± 2.4% (internal calibration) were reached, respectively, after sample clean-up. [ABSTRACT FROM AUTHOR]

This article covers the latest activities in mycotoxin analysis and the advances of its respective quality assurance. The majority of mycotoxin analyses carried out in the laboratories is still based on physicochemical methods, which are continually improved. For example, immunoaffinity columns and multifunctional clean–up columns have become of increasing importance and in some areas of mycotoxin analysis they have more or less displaced conventional liquid–liquid partitioning or column chromatography during clean–up. The need for rapid yes/no decisions on the other hand has led to a number of new screening methods. In particular, rapid and easy–to–use test kits based on immunoanalytical principles or the generation of artificial macromolecular receptors employed in molecularly imprinted polymers (MIPs) have made good progress. Further research in mycotoxin analysis is pursued in the field of biosensors and also the potential of infrared spectroscopic techniques as screening method has been demonstrated. In the area of multi mycotoxin analysis the most promising development was observed in mass spectrometry. At the same time, several interlaboratory studies in the field of mycotoxin analysis revealed problems proven by high between laboratory standard deviation and non–traceable results. This not only shows the necessity of reliable methods and well defined performance characteristics but also the need for appropriate calibrants of defined concentration and stated purity. A certified zearalenone (ZON) calibrant is already available and a certified calibrant containing various trichothecenes is currently under development. (Certified) reference materials are available for aflatoxins in a number of commodities, ochratoxin A (OTA) in wheat, deoxynivalenol (DON) in maize and wheat, and ZON in maize. With these measures important steps towards traceability of results in mycotoxin analysis have been achieved. [ABSTRACT FROM AUTHOR]

The performance of the newly developed Mycosep® 229 Ochra and Multisep® 229 Ochra clean-up columns for ochratoxin A (OTA) determination was evaluated. OTA was subsequently analysed using RP-HPLC with fluorescence detection. Recoveries for frequently contaminated commodities, like cereals, red wine, raisins and green coffee, were estimated. The recoveries obtained for the Mycosep 229 Ochra column were in the range from 87.9 ± 12.5% ( n = 6) for wheat to 99.4 ± 2.7% ( n = 24) for raisins. For Multisep 229 Ochra, recoveries from 76.5 ± 8.0% ( n = 6) for barley to 86.4 ± 1.4% ( n = 24) for raisins were achieved. Limits of detection for all matrices investigated (maize, wheat, rice, barley, raisins, green coffee beans, red wine) were in the range 0.4-2.4 μg kg -1 . The trueness of the method was tested using a certified reference material. [ABSTRACT FROM AUTHOR]