Cross-reactivity of some commercially available deoxynivalenol (DON) and zearalenone (ZEN) immunoaffinity columns to DON- and ZEN-conjugated forms and metabolites

Seven commercially available deoxynivalenol (DON) and zearalenone (ZEN) immunoaffinity columns (IACs) were tested for cross-reactivity to conjugated forms (3-acetyl-deoxynivalenol, 15-acetyl-deoxynivalenol, DON-3-glucoside, DON-3-glucuronide, ZEN-glucosides, ZEN-glucuronide) and metabolites (de-epoxydeoxynivalenol, α-zearalenol, β-zearalenol) and nivalenol (NIV), using a semi-quantitative multi-mycotoxin ultra-performance liquid chromatography-tandem mass spectrometry method. The DON IACs showed cross-reactivity for nearly all DON derivatives tested. The ZEN IACs showed limited cross-reactivity to some of the ZEN derivatives. The IACs were evaluated for their potential use as sample clean-up for mycotoxins in serum.     

Cross-reactivity of some commercially available deoxynivalenol (DON) and zearalenone (ZEN) immunoaffinity columns to DON- and ZEN-conjugated forms and metabolites

Seven commercially available deoxynivalenol (DON) and zearalenone (ZEN) immunoaffinity columns (IACs) were tested for cross-reactivity to conjugated forms (3-acetyl-deoxynivalenol, 15-acetyl-deoxynivalenol, DON-3-glucoside, DON-3-glucuronide, ZEN-glucosides, ZEN-glucuronide) and metabolites (de-epoxydeoxynivalenol, α-zearalenol, β-zearalenol) and nivalenol (NIV), using a semi-quantitative multi-mycotoxin ultra-performance liquid chromatography-tandem mass spectrometry method. The DON IACs showed cross-reactivity for nearly all DON derivatives tested. The ZEN IACs showed limited cross-reactivity to some of the ZEN derivatives. The IACs were evaluated for their potential use as sample clean-up for mycotoxins in serum.     

Analysis of deoxynivalenol and deoxynivalenol-3-glucosides content in Canadian spring wheat cultivars inoculated with Fusarium graminearum

Contamination of wheat grains with Fusarium mycotoxins and their modified forms is an important issue in wheat industry. The objective of this study was to analyse the deoxynivalenol (DON) and deoxynivalenol-3-glucosides (D3G) content in Canadian spring wheat cultivars grown in two locations, inoculated with a mixture of 3-acetyldeoxynivalenol (3-ADON)-producing Fusarium graminearum strains and a mixture of 15-acetlyldeoxynivalenol (15-ADON)-producing F. graminearum strains. According to the analysis of variance, significant differences were observed among the cultivars for Fusarium head blight (FHB) disease index, Fusarium-damaged kernel percentage (%FDK), DON content and D3G content. When the effect of chemotype was considered, significant differences were observed for FHB disease index, FDK percentage and DON content. The D3G content and D3G/DON ratio were not significantly different between the chemotypes, except for D3G content at the Winnipeg location. The Pearson correlation coefficient between DON and D3G was 0.84 and 0.77 at Winnipeg and Carman respectively. The highest D3G/DON ratio was observed in cultivars Carberry (44%) in Carman and CDC Kernen (63.8%) in Winnipeg. The susceptible cultivars showed lower D3G/DON ratio compared with the cultivars rated as moderately resistant and intermediate. The current study indicated that Canadian spring cultivars produce D3G upon Fusarium infection.     

Analysis and occurrence of deoxynivalenol (DON) in cocoa

The primary objective of this study was to establish a current situation assessment of the possible occurrence of deoxynivalenol in cocoa and cocoa products. Since there was no analytic method for determining DON in cocoa and cocoa products, a special method was developed. The applicability and consistency of the method was confirmed by performing recovery assays on various cocoa products. A special post-column derivatisation procedure was used to increase selectivity and raise sensitivity by a factor of 80. The method’s limit of detection (LOD) was thereby reduced to 7 μg/kg; the limit of quantification (LOQ) was 14 μg/kg. The method was used to test 230 samples for possible DON content, ranging from cocoa beans to cocoa bean shells, nibs, cocoa liquor and cocoa powders through to finished cocoa-based products. In the case of cocoa beans and cocoa bean shells, DON content close to the detection limit was only determined in isolated cases. No DON content was detected in nibs, cocoa liquor, cocoa powders and finished cocoa-based products. Analogous to ochratoxin A and aflatoxins, the results show DON is more likely to be found in cocoa bean shells. Separation of shells during cocoa processing can reduce potential DON contents. Since no DON was determined in the fractions relevant for chocolate production, these assays show it does not represent a considerable issue for the cocoa and chocolate industry.     

粮油食品中呕吐毒素危害及风险分析

该文分析呕吐毒素性质,在自然界和粮油食品中存在情况,及对人类和动物危害,并进行风险评估,同时提出预防呕吐毒素危害建议。     

Stability of the mycotoxin deoxynivalenol (DON) during the production of flour-based foods and wheat flake cereal

Deoxynivalenol (DON) is a mycotoxin found in cereal grains and cereal-based foods. DON concentrations in finished products are reduced under some processing conditions, but not others. DON concentrations in flour, wheat and selected foods made from them under commercially relevant conditions were compared by GC with electron capture detection. Average concentrations (n?=?9/item) in cookies, crackers and pretzels ranged from 61% (cookies) to 111% (pretzels) compared with flour (100%?=?0.46?µg?g^?1). Lesser amounts were found in donuts and bread: their respective DON concentrations were 44% and 30% that of flour. Mass balance estimates for DON (µg?g^?1 flour equivalents) ranged from 50% (bread?=?0.23?µg?g^?1 flour equivalents) to 120% (donuts), indicating that dilution by recipe ingredients contributed to DON reductions in bread and accounted for all of the apparent reduction in donuts. Mass balance estimates averaged 76% (crackers) to 107% (pretzels) for the other flour products. DON concentrations were higher in cereal flakes (0.55?µg?g^?1 in the finished product and 0.58?µg?g^?1 on a mass balance basis) than in wheat (0.40?µg?g^?1), suggesting that DON concentrations might increase during processing of wheat cereals under some conditions. In summary, DON concentrations of finished food products were reduced?≥50% only in bread and donuts. Reduction in bread resulted from a combination of DON ‘loss’ and dilution by recipe ingredients whereas the reduction in donuts was due entirely to dilution. These results are further evidence of DON stability during the preparation of popular flour or wheat-based products.     

Survey of deoxynivalenol and its conjugates deoxynivalenol-3-glucoside and 3-acetyl-deoxynivalenol in 374 beer samples

Beer is one of the most popular beverages worldwide. Malted cereal grains are among the basic ingredients and hence mycotoxin contamination might occur. Previous studies reported the presence of the Fusarium mycotoxins deoxynivalenol (DON) and 3-acetyl-deoxynivalenol (3ADON), as well as of the masked mycotoxin deoxynivalenol-3-glucoside (D3G) in beer. In the present survey, 374?beer samples from 38?countries with a focus on Austrian (156) and German (64) beers were analysed for the presence of D3G, DON and 3ADON. Beers were assigned to the following six categories: pale (217), wheat (46), dark (47), bock (20), nonalcoholic beers (19) and shandies (25). In total, 348 and 289 beers (93 and 77%, respectively) contained D3G and DON at the levels above the limit of detection, whereas 3ADON was not detected in any of the samples. Average concentrations of all beers were 6.9?µg?L^?1 for D3G and 8.4?µg?L^?1 in the case of DON. Nonalcoholic beers and shandies showed the lowest contaminations, 1.5 and 3.2?µg?L^?1 for D3G and 2.7 and 4.4?µg?L^?1 for DON, respectively. In bock beers characterised by a higher gravity, a significant trichothecene load of 14.8?µg?L^?1 D3G and 12.4?µg?L^?1 DON was found. The highest contamination (81?µg?L^?1 D3G, 89?µg?L^?1 DON) was detected in a pale beer from Austria, underlining the importance of this study for food safety. The molar D3G to DON ratio ranged between 0.11 and 1.25 and was 0.56 on average. Concluding, the average contamination of beer is not of toxicological concern for moderate beer drinkers. However, in the case of heavy beer drinkers, beer consumption may considerably contribute to the overall intake of DON, which might even lead to exceeding the maximum tolerable limits established for this Fusarium toxin.     

Assessment of deoxynivalenol (DON) adsorbents and characterisation of their efficacy using complementary in vitro tests

Deoxynivalenol (DON) is a prevalent and resistant mycotoxin found in cereals and related products. Adsorbents appear to provide an opportunity to decrease DON absorption in animals but, due to their specificity, it is very difficult to evaluate their actual efficacy. It is pointless to extrapolate results obtained with one mycotoxin to another and even to extrapolate results obtained in vitro in buffer to an in vivo situation. We carried out experiments to characterize the properties of potential DON adsorbents. Initial tests in buffer pH 7 allowed us to focus on six adsorbents: activated charcoal, cholestyramin, Saccharomyces cerevisiae mannans, algal β-glycan, fungal β-glycan and leguminous plant. The use of equilibrium sorption models suggested a non-saturated phenomenon and involved variable mechanisms according to the specific material. Subsequent tests with a Caco-2 cell model showed a high reduction in DON cytotoxicity on proliferative intestinal cells and DON absorption by differentiated intestinal cells when adsorbent was added (except for cholestyramin). Otherwise, values were not always in accordance with those obtained in buffer. Our work allowed us to identify five potential DON adsorbents and to propose a complementary in vitro test allowing improved determination of adsorbent properties.     

Residues of deoxynivalenol (DON) in pig tissue after feeding mash or pellet diets containing low concentrations

Residues of deoxynivalenol (DON) and its metabolite de-epoxy-DON (DOM) were analyzed in specimens of pigs fed diets containing 0, 25, and 50% contaminated wheat (2.5 mg DON/kg) fed as mash or pellets over the final growing period of 11 wk. Median DON concentrations decreased from bile > kidney > serum > liver = muscle, while DOM was only detected in bile and kidney. Maximum carry over rates were 0.0319 for kidney, 0.0064 for liver, and 0.0043 for muscle, demonstrating that the contribution of animal derived food to the consumers' exposure is very low. The high interindividual variation of DON concentrations in all analyzed specimen of pigs fed diets containing similar concentrations of DON does not allow a diagnostic differentiation of animals fed diets containing DON concentrations of approximately 61% of the guidance level of 0.9 mg DON/kg, and those fed diets containing 137% of this concentration. The different feed forms did not affect residue concentrations in any of the investigated specimens.     

Advantages and drawbacks of immunoaffinity columns in analysis of mycotoxins in food

A number of countries are setting legislations on mycotoxins. In order to reduce dispute between importing and exporting countries, the analytical data should be as comparable as possible, especially when levels are close to the regulatory limits. The present trend in the analysis of mycotoxins is to use immunoaffinity column (IAC) as a clean-up and enrichment technique, and Association of Official Analytical Chemists and European Union have validated methods which address a few food commodities. This study describes our experience using both conventional and IAC approaches in the analysis of three mycotoxins. Aflatoxins (AFs): Aflatoxin G1 has been detected by liquid-liquid partitioning methods with HPLC detection as false-positive in some maize. On IACs, this compound behaves as an AF, lowering the amount of the AFs trapped. The problem was solved using either TLC or HPLC with detection in the Kobra cell. Depending on the additives to food during the processing and cooking, the AFs might appear as an opened ring not recognised by the antibody. Fumonisins (FB): Compounds interfering with the FB's antibodies were also observed while analysing breakfast cereals leading to underestimation of FB. Ochratoxin A (OTA): Depending on the food composition and extraction techniques, OTA is underestimated with IAC in some breakfast cereals and coffee. These data strengthen the necessity to validate methods using IAC for each complex matrix.     

Effects of oral exposure of pigs to deoxynivalenol (DON) sulfonate (DONS) as the non-toxic derivative of DON on tissue residues of DON and de-epoxy-DON and on DONS blood levels

The Fusarium toxin deoxynivalenol (DON) is of outstanding importance in pig nutrition because of its frequent occurrence in cereal grains at levels high enough to cause adverse effects such as a decrease in feed intake and impairment of the immune system. Thus, simple decontamination procedures would be useful. The present study aimed to examine the effects of wet preservation of triticale contaminated with DON and zearalenone (ZON) with sodium metabisulphite (SBS) on the treatment-related non-toxic derivative of DON (DON-sulfonate, DONS), and on ZON and its metabolites in blood and various physiological specimens of piglets. The uncontaminated control triticale (CON) and the DON-contaminated triticale (FUS) were included in the diets either untreated or SBS treated (CON-SBS, FUS-SBS) and fed to piglets for 28 days starting from weaning. The diet concentrations for DON were 0.156, 0.084, 2.312 and 0.275 mg kg^?1, for DONS were <0.05, <0.05, <0.05 and 1.841 mg kg^?1, and for ZON were <0.001, 0.006, 0.017, and 0.016 mg kg^?1 for each of CON, CON-SBS, FUS and FUS-SBS, respectively. DONS was present in the blood of piglets fed the FUS-SBS at a median concentration of 15.5 ng ml^?1 (3–67 ng ml^?1), while the median DON concentration amounted to 2 ng ml^?1 (0–5 ng ml^?1) at the same time. The median DON concentration in the blood of piglets fed the FUS diet reached a median concentration of 10.5 ng ml^?1 (5–17 ng ml^?1). Moreover, the relative differences between the DON concentrations in other physiological specimens (muscle, liver, kidney, bile and urine) in piglets fed the FUS-SBS and the FUS diet were comparable with the blood DON concentration differences. Although these differences can be taken as an indication for DONS stability after absorption and distribution further studies examining DONS in these other physiological specimens directly are necessary to substantiate this conclusion. Moreover, ZON and α-zearalenol could only be detected in bile and urine where their levels were not influenced by the SBS treatment.     

Influence of temperature and pH on the degradation of deoxynivalenol (DON) in aqueous medium: comparative cytotoxicity of DON and degraded product

Deoxynivalenol (DON), a toxic fungal metabolite, is stable under different processing conditions; however, its stability in aqueous medium at different temperatures and low pH (1–2) (present in the gastrointestinal tract) has not been investigated. In the present study, DON standard was used to study the influence of temperature and pH on DON stability in aqueous medium, the characterisation of the degraded product, and the comparative toxicity profile of the degraded and the parent compound. The results suggest that standard DON was unstable at 125–250°C showing 16–100% degradation whereas DON at pH 1–3 had 30–66% degradation, with a concomitant increase in the formation of a degraded product. Further ESI-MS characterisation of the dominant precursor ion of the HPLC eluate of the DON-degraded product was found to be m/z 279, resembling the known metabolite DOM-1. The degraded product of DON was reconfirmed as DOM-1 by comparison with standard DOM-1 and both gave a similar λ_max at 208 nm. Comparative studies of both standard DOM-1 and the degraded product of DON showed no cytotoxicity up to 6400 ng ml^–1 while significant cytotoxicity was observed for DON (400 ng ml^–1). The results suggest that a highly acidic environment (pH 1–2) could be responsible for the de-epoxydation of DON leading to the formation of DOM-1.     

Occurrence and risk assessment of population exposed to deoxynivalenol in foods derived from wheat flour in Brazil

Deoxynivalenol (DON) is the most important of the trichothecenes in terms of amounts and occurrence in wheat. This compound was shown to be associated with a glomerulonephropathy involving an increase of immunoglobulin A in humans. This study assessed the occurrence of DON in wheat flour and the exposure of Brazilian teenagers, adults and elderly to this mycotoxin due to intake of wheat flour-based products. DON extraction in wheat flour was carried out by solid phase extraction and the quantification was performed by ultra-high proficiency liquid chromatography with diode-array detection. A total of 77.9% of all samples were positive for DON, with concentrations ranging from 73.50 to 2794.63 µg kg^?1. The intake was calculated for the average and 90th percentile of the contamination levels of DON in foods based-wheat for teenagers, adults and elderly in Brazil, and compared with the provisional maximum tolerable daily intakes (PMTDI). Females of all age groups were exposed to DON at higher levels when compared to males in regard of consumption of breads and pastas. Teenagers were the main consumers of foods derived from wheat flour, with maximum probable daily intakes of 1.28 and 1.20 µg kg^?1 b.w. day^?1 for females and males, respectively. This population is at an increased risk of exposure to DON due to consumption of wheat flour-based foods in Brazil.     

Fate of deoxynivalenol,T-2 and HT-2 toxins and their glucoside conjugates from flour to bread: an investigation by high-performance liquid chromatography high-resolution mass spectrometry

Deoxynivalenol, T-2 and HT-2 toxins are mycotoxins frequently occurring in cereals and cereal-based products along with their conjugated forms. In this paper, we provide insights into the fate of deoxynivalenol, T-2 and HT-2 toxins and their glucoside derivatives during bread making, using naturally contaminated wheat flour. High-resolution mass spectrometry was used to assess the extent of degradation of the three mycotoxins during bread baking and to identify some glucoside conjugates, namely deoxynivalenol, T-2 and HT-2 mono-glucosides, detected both in the flour and in the respective breads. Our findings show deoxynivalenol's levels markedly increased upon baking, whereas those of HT-2 and T-2 toxins were decreased in the final bread with special regard to the T-2 toxin.     

Occurrence of mycotoxins (ochratoxin A, deoxynivalenol) and toxigenic fungi in Moroccan wheat grains: impact of ecological factors on the growth and ochratoxin A production

The aim of the present work was to evaluate the contamination of some samples, taken from Moroccan wheat grains, by ochratoxin A (OTA), deoxynivalenol (DON) and the associated toxigenic fungi. Moreover, we focused on the influence of environmental factors on both the growth and OTA production by three strains of Aspergillus. The results showed that only few samples were contaminated by the two mycotoxins (2 samples for OTA and 7 for DON). The main isolated fungi belong to the Aspergillus, Penicillium and Fusarium genus; 74 Aspergillus and 28 Penicillium isolates were tested for their ability to produce OTA. Only 2 A. alliaceus and 14 A. niger were able to synthesize OTA. However, none of Penicillium isolates can produce this toxin under the conditions mentioned. In respect of the effects of the temperature and water activity (aw), the optimal conditions for the growth and OTA production were different. While the optimal conditions of growth for A. alliaceus and A. terreus are 30 degrees C and 0.98 aw, A. niger preferred 0.93-0.95 aw at 25 degrees C, whereas the optimal production of OTA was observed at 30 degrees C for both A. alliaceus and A. niger at 0.93 and 0.99 aw, respectively.     

Use of immunoaffinity columns for clean-up of diarrhetic toxins (okadaic acid and dinophysistoxins) extracts from shellfish prior to their analysis by HPLC/fluorimetry

Diarrhetic Shellfish Poisoning (DSP) is a severe gastro-intestinal disease caused by consumption of seafood contaminated by microalgal toxins, mainly okadaic acid (OA) and structurally related toxins, dinophysistoxins (DTXs). Regulatory monitoring is generally based on rodent bioassays which, however, present some technical and ethical disadvantages. The most promising technique of analysis of these toxins involves an HPLC separation with spectrofluorimetric detection after derivatization of the toxins with a fluorescent reagent. The lack of specificity of the extraction procedure (liquid-liquid partition), and the presence of interfering compounds in the matrix, does not allow the determination and the quantification of low amounts of toxins in seafood. In this paper, the authors report the development and the characterization of immunoaffinity columns (IAC), which were elaborated using anti-okadaic acid monoclonal antibodies, for a specific retention of the OA group of toxins. The coupling yield and the stability of these columns were investigated as well as their capacity to remove interfering compounds. Cross-reactivity was observed between the antibodies and the DTX-1 and the DTX-2, allowing the detection of the different toxins in a single analysis. Different spiked (1mugOA/ g) or naturally-contaminated (mussel digestive gland: 2mugOA/g; algae: 165mugOA/g) matrices were tested. The recovery for OA varied from 55 to 95% according to the matrices. The IAC purification was then included as a step of a global [IAC/HPLC/spectrofluorimetric detection] method and the performance of the method was evaluated. Estimations of the linearity and the accuracy (percentages of the presumptive response for OA in the range + 101% to + 114%) were satisfactory in accordance with the method validation criteria.     

Micelle mediated extraction and simultaneous spectrophotometric determination of vanadium(V) and molybdenum(VI) in plant foodstuff samples

A micelle-mediated extraction method for preconcentration of trace quantities of V(V) and Mo(VI) as a prior step to their simultaneous spectrophotometric determination has been developed. Bromopyrogallol red, cetyltrimethylammonium bromide (CTAB) (cationic surfactant) and KI were used as chelating, extraction and co-extraction agents, respectively.     

Rapid and non-invasive analysis of deoxynivalenol in durum and common wheat by Fourier-Transform Near Infrared (FT-NIR) spectroscopy

Fourier transform near-infrared spectroscopy (FT-NIR) was used for rapid and non-invasive analysis of deoxynivalenol (DON) in durum and common wheat. The relevance of using ground wheat samples with a homogeneous particle size distribution to minimize measurement variations and avoid DON segregation among particles of different sizes was established. Calibration models for durum wheat, common wheat and durum + common wheat samples, with particle size <500 µm, were obtained by using partial least squares (PLS) regression with an external validation technique. Values of root mean square error of prediction (RMSEP, 306–379 µg kg^–1) were comparable and not too far from values of root mean square error of cross-validation (RMSECV, 470–555 µg kg^–1). Coefficients of determination (r ²) indicated an “approximate to good” level of prediction of the DON content by FT-NIR spectroscopy in the PLS calibration models (r ² = 0.71–0.83), and a “good” discrimination between low and high DON contents in the PLS validation models (r ² = 0.58–0.63). A “limited to good” practical utility of the models was ascertained by range error ratio (RER) values higher than 6. A qualitative model, based on 197 calibration samples, was developed to discriminate between blank and naturally contaminated wheat samples by setting a cut-off at 300 µg kg^–1 DON to separate the two classes. The model correctly classified 69% of the 65 validation samples with most misclassified samples (16 of 20) showing DON contamination levels quite close to the cut-off level. These findings suggest that FT-NIR analysis is suitable for the determination of DON in unprocessed wheat at levels far below the maximum permitted limits set by the European Commission.     

A comparison of solid-phase microextraction (SPME) with simultaneous distillation-extraction (SDE) for the analysis of volatile compounds in heated beef and sheep fats

A comparison has been made on the application of SPME and SDE for the extraction of volatile compounds from heated beef and sheep fats with separation and measurement by gas chromatography-mass spectrometry. As far as we know, this report represents the first time that such a comparison has been made for the measurement of volatile compounds in heated sheep fat. Approximately 100 compounds (in relatively high abundance) were characterised in the volatile profiles of heated beef and sheep fats using both techniques. Differences were observed in the volatile profiles obtained from each technique, independent of compound class. Rather than rate one technique as superior to another, the techniques can be regarded as complementary to each other.