Trichothecenes In Cereals Research Articles

An accurate and efficient method for simultaneous determination of 16 trichothecenes in maize and wheat by UHPLC-Q-Orbitrap-HRMS combined one-step pretreatment with a novel isotope multipoint calibration strategy

Trichothecenes are mycotoxins generated by Fusarium species that are toxic to humans and animals and frequently contaminate cereals. Herein, we present an UHPLC-Q-Orbitrap-HRMS method using one-step pretreatment for simultaneous detection of 16 type A and B trichothecenes in maize and wheat. A calibration strategy based on natural isotopic distribution was adopted and validated to replace traditional time-consuming and laborious multipoint external calibration curves. Various conditions influencing the method were studied. The recoveries of spiked analytes at three different concentrations ranged from 74.2% to 105.3% and the inter-day relative standard deviations were 0.5–15.6%. The limits of quantitation were 4.4–108.8 μg/kg. Moreover, the current methodology was applied to the evaluation of type A and B trichothecenes present in samples from major wheat production areas in China. This study provides a facile methodology for type A and B trichothecenes analysis and presents promising applications for exposure risk assessment of trichothecenes in cereals.

The mode of phytotoxic action of trichothecenes during plant-<i>Fusarium</i> interaction; our approach to detoxification of trichothecenes in cereals.

The mode of phytotoxic action of trichothecenes during plant-<i>Fusarium</i> interaction; our approach to detoxification of trichothecenes in cereals.

Simultaneous determination of type A, B and D trichothecenes and their occurrence in cereals and cereal products

A sensitive LC–MS/MS method for the simultaneous determination of type A, B and D trichothecenes in cereals is presented. The limits of detection ranged between 0.1 and 0.7 µg kg−1 for all analytes. The method was applied to 289 representatively drawn samples of wheat, rye and oat products. Ninety-four percent of the wheat samples (n = 130), 95% of the rye samples (n = 61) and 100% of the oat samples (n = 98) were contaminated with the type A trichothecenes T-2 and HT-2 toxin. Median levels of T-2/HT-2 (sum of the toxins) were 0.91, 0.53 and 8.2 µg kg−1, respectively. Highest levels were found in wheat bran (24 µg kg−1), rye kernels (3.1 µg kg−1) and oat flakes (85 µg kg−1). All wheat and rye samples and 75% of the oat samples were contaminated with the type B trichothecene deoxynivalenol. Median levels of this toxin were 23, 15 and 0.53 µg kg−1, respectively. Highest levels were found in wheat bran (1160 µg kg−1), rye kernels (288 µg kg−1) and oat flakes (55 µg kg−1). The type B trichothecene nivalenol was detected in 67% of the wheat samples, in 3% of the rye samples and in 24% of the oat samples with highest levels in wheat bran (96 µg kg−1), rye kernels (1.8 µg kg−1) and in oat flakes (17 µg kg−1), respectively. Levels of other type A and B trichothecenes played a minor role, although the rates of contamination were often high. Neither macrocyclic type D trichothecenes (satratoxin G and H, verrucarin A, roridin A) nor diacetylverrucarol and verrucarol (type A trichothecenes), were detected in any of the samples.

Multitoxin extraction and detection of trichothecenes in cereals: an improved LC‐MS/MS approach

AbstractBACKGROUND: Many multiresidual methods to evaluate natural occurrence of Fusarium toxins are already reported in the scientific literature but a new rapid, reliable, cost‐efficient and high‐sensitivity method for the simultaneous determination of several fusariotoxins is always welcome. Nivalenol (NIV), deoxynivalenol, fusarenon‐X (FUS‐X), 3‐acetyldeoxynivalenol, diacetoxyscirpenol (DAS), HT‐2 toxin, T‐2 toxin, neosolaniol (NEO), zearalanone and zearalenone (ZON) belong to the most common mycotoxins in food matrix grains, e.g., wheat and maize. The proposed method is a multitoxin analytical method that combines high‐performance liquid chromatography (HPLC), atmospheric pressure chemical ionization (APCI), triple‐quadrupole tandem mass spectrometry (LC‐MS/MS) under the selected reaction monitoring (SRM) mode, and it is focused on the optimization of the sample preparation without the need for any cleanup.RESULTS: Three different methods for sample preparation and for the simultaneous extractions of the above‐mentioned fusariotoxins were tested: two of these were followed by a different cleanup step for comparison, while the extraction method proposed in this work, which uses an 84% (v/v) acetonitrile aqueous solution, sample homogenization and subsequent filtration, was validated without any further cleanup step.CONCLUSION: Calibration curves for all analytes are linear, except DAS, HT‐2 and ZON, over the working range of 10–1000 µg kg−1. The calibration curve of DAS was linear between 10 and 500 µg kg−1, although the curves of HT‐2 and ZON were linear in the range 10–250 µg kg−1. Squared correlation coefficients (R2) were in the range 0.995–0.998 for the all point calibration curves. The lowest limits of detection (LOD) were found for DON and ZAN with 0.5 and 0.2 µg kg−1, respectively, while the highest LODs were obtained for NIV, FUS‐X and NEO, with 3.3 µg kg−1 for each toxin. Copyright © 2009 Society of Chemical Industry

Determination of trichothecenes in cereals and cereal-based products by liquid chromatography–tandem mass spectrometry

A sensitive, accurate and precise method for the simultaneous determination of nivalenol (NIV), deoxynivalenol (DON), T-2 toxin (T-2) and HT-2 toxin (HT-2) in different food matrices, including wheat, maize, barley, cereal-based infant foods, snacks, biscuits and wafers, has been developed. The method, using liquid chromatography coupled with atmospheric pressure chemical ionization triple quadrupole mass spectrometry (LC–APCI–MS/MS), allowed unambiguous identification of the selected trichothecenes at low µg per kg levels in such complex food matrices. A clean-up procedure, based on reversed phase SPE Oasis® HLB columns, was used, allowing good recoveries for all studied trichothecenes. In particular, NIV recoveries significantly improved compared to those obtained by using Mycosep® #227 columns for clean-up of the extracts. Limits of detection in the various investigated matrices ranged 2.5–4.0 µg kg−1 for NIV, 2.8–5.3 µg kg−1 for DON, 0.4–1.7 µg kg−1 for HT-2 and 0.4–1.0 µg kg−1 for T-2. Mean recovery values, obtained from cereals and cereal products spiked with NIV, DON, HT-2 and T-2 toxins at levels from 10 to 1000 µg kg−1, ranged from 72 to 110% with mean relative standard deviation lower than 10%. A systematic investigation of matrix effects in different cereals and cereal products was also carried out by statistically comparing the slopes of standard calibration curve with matrix-matched calibration curve for each of the four toxins and the eight matrices tested. For seven of the eight matrices tested, statistically significant matrix effects were observed, indicating that, for accurate quantitative analysis, matrix-matched calibration was necessary. The method was applied to the analysis of 57 samples of ground wheat originated from South Italy and nine cereal food samples collected from retail markets.

A new solid phase extraction clean‐up method for the determination of 12 type A and B trichothecenes in cereals and cereal‐based food by LC‐MS/MS

A new reliable and cost-efficient solid phase extraction-based clean-up method for the determination of 12 type A and B trichothecenes [deoxynivalenol (DON), nivalenol, 3-acetyldeoxynivalenol, 15-acetyldeoxynivalenol, fusarenon-X, T-2 toxin, HT-2 toxin, neosolaniol, monoacetoxy-scirpenol, diacetoxyscirpenol, T-2 triol and T-2 tetraol] in cereals and cereal-based food is presented. Furthermore, the suitability for the simultaneous determination of zearalenone is examined. Toxins were extracted from cereal samples using ACN/water (80/20, v/v), purified by means of a new Bond Elut Mycotoxin column and analyzed via liquid chromatography-electrospray ionization tandem mass spectrometry. Limits of detection were calculated for the matrix wheat and ranged from 0.3 to 5 ng/g, depending on the toxin. Average recovery rates for the tested compounds in seven cereal-based matrices have been determined ranging from 65 to 104%. The relative standard deviations of the complete method ranged from 2.67 (DON, wheat) to 20.0% (T-2 toxin, oats).

Determination of 12 Type A and B Trichothecenes in Cereals by Liquid Chromatography−Electrospray Ionization Tandem Mass Spectrometry

A new sensitive method for the simultaneous determination of 12 trichothecenes (deoxynivalenol, nivalenol, 3-acetyldeoxynivalenol, 15-acetyldeoxynivalenol, fusarenon X, T-2 toxin, HT-2 toxin, neosolaniol, monoacetoxyscirpenol, diacetoxyscirpenol, T-2 triol, and T-2 tetraol) by liquid chromatography-electrospray ionization tandem mass spectrometry (LC-ESI-MS/MS) is presented. The development of the method and investigations on the matrix influence on the MS signal are described in particular. The matrix effect was thereby minimized by using an internal standard, a special mobile phase, and specific fragmentation parameters. The sample was extracted with acetonitrile/water (84:16, v/v), and the extract was cleaned up with a MycoSep 227 column. Quantification was based on the internal standard de-epoxy-deoxynivalenol. Calibration curves were linear between 16 and 1600 ng/g, and the limits of detection ranged from 0.18 to 5.0 ng/g. The developed method was applied for the determination of trichothecenes in 120 naturally contaminated wheat and oat samples.

Simultaneous detection of type A and type B trichothecenes in cereals by liquid chromatography–electrospray ionization mass spectrometry using NaCl as cationization agent

A LC/MS method for the simultaneous determination of both type A and type B trichothecenes by using an electrospray ionization (ESI) interface in the positive ionization mode with a single quadrupole analyzer is described. In order to enhance the ionization of both groups of trichothecenes, the sodium ion was used as cationization agent by adding sodium chloride to the eluent. All LC/MS parameters were optimized. The newly developed LC/ESI-MS method was applied to the analysis of a wheat reference material and cereal-based foods and feeds.

Simultaneous detection of type A and type B trichothecenes in cereals by liquid chromatography–electrospray ionization mass spectrometry using NaCl as cationization agent

A LC/MS method for the simultaneous determination of both type A and type B trichothecenes by using an electrospray ionization (ESI) interface in the positive ionization mode with a single quadrupole analyzer is described. In order to enhance the ionization of both groups of trichothecenes, the sodium ion was used as cationization agent by adding sodium chloride to the eluent. All LC/MS parameters were optimized. The newly developed LC/ESI-MS method was applied to the analysis of a wheat reference material and cereal-based foods and feeds.

Toxicological evaluation of trichothecenes in animal feed

Trichothecenes are mycotoxins commonly found in cereals world-wide. Fusarium fungi are the main producers of trichothecenes in cereals. Trichothecenes are rapidly excreted from animals and residues of trichothecenes in animal-derived food products are not considered to pose any threat to consumers. The toxins are toxic to all tested species, but the sensitivity varies considerably between toxins and between species. Available feeding studies with the trichothecenes deoxynivalenol (DON), nivalenol (NIV) and T-2 toxin in feed to production animals have been reviewed. There are not sufficient available data about the effects of trichothecenes in ruminant feed to allow a scientifically-based risk assessment. The available studies of the metabolism of trichothecenes in cattle indicate that trichothecenes to a large extent are transformed to the much less toxic de-epoxide metabolite in the rumen before absorption. Furthermore, no effect has been found on milk production, feed intake or other parameters measured at levels used in the studies. It is concluded that trichothecenes are not likely to cause any harm in ruminants unless fed visibly damaged feed and no guideline value is probably needed. Poultry are more sensitive to trichothecenes than ruminants. Levels from 9 mg DON/kg feed have lead to negative effects of chickens, while no effect was found in chicken fed 5 mg DON/kg feed, and a guideline value of 2.5 mg DON/kg feed is proposed. The available information about NIV does not allow a guideline limit to be set, but the finding of minor pathological changes in chicken fed 1 mg NIV/kg feed indicates that NIV may be more toxic to poultry than DON. Oral lesions are observed in chickens and hens fed 1 mg T-2 toxin/kg feed. Other effects, including a reduction in feed intake, are found with increasing levels of T-2 toxin. A guideline limit of 0.5 mg T-2 toxin/kg is proposed. Pigs are more sensitive to trichothecenes than other farm animals. The effects occurring at the lowest levels of trichothecenes are reduced feed intake and weight gain, normally occurring at levels from 0.6 mg DON/kg feed in naturally contaminated feed. Pigs fed 0.5 mg T-2 toxin/kg feed reduced their feed intake. Impairment of the immune system has also been observed in pigs at this level of T-2 toxin in the feed. Guidelines of 0.3 mg DON/kg feed and 0.2 mg T-2 toxin/kg feed is proposed in pig feed.

Sources of variation in the analysis of trichothecenes in cereals by gas chromatography-mass spectrometry.

The sources of variation In the analysis of trichothecenes in cereals by gas chromatography mass spectrometry (GC-MS) were studied and Identified. Ways to decrease some of the variations identified are presented and discussed. The method Is validated for deoxynivalenol, nivalenol, 3-acetyldeoxynivalenol, fusarenon-X, diacetoxyscirpenol, HT-2 toxin and T-2 toxin. The general variability in the performance of the GC-MS apparatus itself, as well as derivatisation, matrix effect, injection and quantification were identified to be the sources of variation. The use of internal standards was studied in order to decrease the variation due to problems with derivatisation and the overall sample preparation. Two derivatisation reagents were compared, one of which was found to be more effective for decreasing this variation. The calibrants prepared in the cereal extract reduced the variation due to the matrix effect and thus improved the quantification. "Constant Standards" were introduced in order to detect and decrease the variation caused by injection and the apparatus itself. The validation study proved that this analytical method for trichothecenes was adequately reliable and sensitive. The relative standard deviation varied between 3-9% and the recovery between 46-90% for the different trichothecenes determined. The limit of detection for a range of trichothecenes varied from 5 μg/kg to 15 μg/kg.

Revisión: Implicaciones toxicológicas y nutricionales de la toxina T-2 / Review: Toxicological and nutritional implications of T-2 toxin

Trichothecenes are mycotoxins produced by species of the genus Fusarium. These toxins are associated with health problems in humans and animals. The most common trichothecenes in cereals are deoxynivalenol, HT-2 toxin, diacetoxyscirpenol, nivalenol, neosolaniol and T-2 toxin; the latter is the most widely studied because it is easy to produce in the laboratory. The effects of T-2 toxicosis include dermatonecrosis, reduced body weight and efficiency of food utilization, severe diarrhoea, haemorrhage, necrosis of the upper alimentary tract, anaemia, immuno suppression ; and in birds, poor feathering. This paper reviews the latest information about the occurrence, chemical characteristics, toxicity, metabolic alterations, biotransformation and detoxifi cation methods of the T-2 toxin.

Determination of Type A and B Trichothecenes in Cereals by Gas Chromatography with Electron Capture Detection

Abstract A quantitative method has been developed for determination of nonmacrocyclic trichothecenes in cereals. The mycotoxins are extracted with acetonitrile- ethyl acetate-water, and the extracts are quickly defatted with hexane and purified on a Sep- Pak Florisil cartridge. The trichothecenes are then silylated with Tri-Sil/TBT and quantitated by capillary gas chromatography with electron capture detection. High recoveries of 13 tested trichothecenes were achieved in experiments on wheat at the 250 μg/kg level. The method was also tested on barley, maize, oats, and rye with good results

Determination of trichothecenes in cereals by gas chromatography with ion trap detection

A method for determination of the trichothecene toxins, deoxynivalenol, 3α-acetyl-deoxynivalenol, nivalenol, T-2 toxin, HT-2 toxin and diacetoxyscirpenol in cereals (wheat, barley, oats, corn) is described. Extraction was performed according to Tanaka et al. (J. Chromatogr.328, 271 (1985)) [33], derivatization by trifluoroacylation with trifluoroacetic acid anhydride. For quantitation and confirmation a capillary gas chromatograph combined with a selective mass detector (ion trap) working in CI-mode with methanol as reagent gas was used. The quantitation limit for the complete method is 1–5 μg/kg, depending on the chemical characteristics of each toxin and cleanness of the extracts. Recoverics from spiked cereals were 78–89%.