Antagonistic properties against Fusarium sporotrichioides and glycosylation of HT-2 and T-2 toxins by selected Trichoderma strains.

The present study assessed the ability of Trichoderma to combat F. sporotrichioides, focusing on their antagonistic properties. Tests showed that Trichoderma effectively inhibited F. sporotrichioides mycelial growth, particularly with T. atroviride strains. In co-cultures on rice grains, Trichoderma almost completely reduced the biosynthesis of T-2 and HT-2 toxins by Fusarium. T-2 toxin-α-glucoside (T-2-3α-G), HT-2 toxin-α-glucoside (HT-2-3α-G), and HT-2 toxin-ß-glucoside (HT-2-3ß-G) were observed in the common culture medium, while these substances were not present in the control medium. The study also revealed unique metabolites and varying metabolomic profiles in joint cultures of Trichoderma and Fusarium, suggesting complex interactions. This research offers insights into the processes of biocontrol by Trichoderma, highlighting its potential as a sustainable solution for managing cereal plant pathogens and ensuring food safety.

Fusarium sporotrichioides Produces Two HT-2-α-Glucosides on Rice.

Fusarium is a genus that mostly consists of plant pathogenic fungi which are able to produce a broad range of toxic secondary metabolites. In this study, we focus on a type A trichothecene-producing isolate (15-39) of Fusarium sporotrichioides from Lower Austria. We assessed the secondary metabolite profile and optimized the toxin production conditions on autoclaved rice and found that in addition to large amounts of T-2 and HT-2 toxins, this strain was able to produce HT-2-glucoside. The optimal conditions for the production of T-2 toxin, HT-2 toxin, and HT-2-glucoside on autoclaved rice were incubation at 12 °C under constant light for four weeks, darkness at 30 °C for two weeks, and constant light for three weeks at 20 °C, respectively. The HT-2-glucoside was purified, and the structure elucidation by NMR revealed a mixture of two alpha-glucosides, presumably HT-2-3-O-alpha-glucoside and HT-2-4-O-alpha-glucoside. The efforts to separate the two compounds by HPLC were unsuccessful. No hydrolysis was observed with two the alpha-glucosidases or with human salivary amylase and Saccharomyces cerevisiae maltase. We propose that the two HT-2-alpha-glucosides are not formed by a glucosyltransferase as they are in plants, but by a trans-glycosylating alpha-glucosidase expressed by the fungus on the starch-containing rice medium.

Debunking the Myth of Fusarium poae T-2/HT-2 Toxin Production.

Fusarium poae is commonly detected in field surveys of Fusarium head blight (FHB) of cereal crops and can produce a range of trichothecene mycotoxins. Although experimentally validated reports of F. poae strains producing T-2/HT-2 trichothecenes are rare, F. poae is frequently generalized in the literature as a producer of T-2/HT-2 toxins due to a single study from 2004 in which T-2/HT-2 toxins were detected at low levels from six out of forty-nine F. poae strains examined. To validate/substantiate the observations reported from the 2004 study, the producing strains were acquired and phylogenetically confirmed to be correctly assigned as F. poae; however, no evidence of T-2/HT-2 toxin production was observed from axenic cultures. Moreover, no evidence for a TRI16 ortholog, encoding a key acyltransferase shown to be necessary for T-2 toxin production in other Fusarium species, was observed in any of the de novo assembled genomes of the F. poae strains. Our findings corroborate multiple field-based and in vitro studies on FHB-associated Fusarium populations which also do not support the production of T-2/HT-2 toxins with F. poae and therefore conclude that F. poae should not be generalized as a T-2/HT-2 toxin producing species of Fusarium.

Co-occurrence of T-2 and HT-2 Mycotoxins and α and ß Anomers of Their Glucosides in Wheat and Oat Grains.

The aim of this study was to simultaneously determine T-2 and HT-2 toxins and the α and ß anomers of their glucosides to assess their content in wheat and oat grains harvested in Poland (2020-2022). Of 298 wheat samples, only 14 (5%) contained the sum of the T-2 and HT-2 toxins (average 34.2 µg/kg; 10.6-67.7 µg/kg). In oat (n = 129), these compounds were detected much more frequently (70% of samples) at an average level of 107.5 µg/kg (6.9-949.1 µg/kg). The sum of T-2 and HT-2 glucosides was detectable in 3% of the wheat (average 16.3 µg/kg; 7.1-39.4 µg/kg) and 65% of the oat samples (average 35.1 µg/kg; 4.0-624.1 µg/kg). Following the study, T-2-3-α-glucoside was identified as the only naturally occurring anomer, while both anomers of HT-2-3-glucosides were detected with higher contents and occurrence rates of HT-2-3-ß-glucoside than the α anomer of this compound.

Determination of mycotoxins by HPLC, LC-MS/MS and health risk assessment of the mycotoxins in bee products of Turkey.

The aim of this study is to determine the levels of deoxynivalenol (DON), HT-2 toxin (HT2), T-2 toxin (T2), and ochratoxin A (OTA) in bee products (bee pollen, propolis, honey and royal jelly) available in Turkey. In addition, exposure and health risk assessments were performed to identify the potential health risk of these mycotoxins. The mycotoxins were analyzed by high-performance liquid chromatography (HPLC) with a UV detector and positive samples were confirmed with liquid chromatography-tandem mass spectrometry (LC-MS/MS). The most common mycotoxins in all bee products were DON and T-2 toxin, with mean concentrations of 1.601 and 0.704 µg/per kg dry sample, respectively, followed by OTA and HT-2 toxin. It was determined that the mycotoxins taken as a result of consuming bee products in specified amounts do not pose a risk to health.

Searching for the Fusarium spp. Which Are Responsible for Trichothecene Contamination in Oats Using Metataxonomy to Compare the Distribution of Toxigenic Species in Fields from Spain and the UK.

The contamination of oats with Fusarium toxins poses a high risk for food safety. Among them, trichothecenes are the most frequently reported in European oats, especially in northern countries. The environmental conditions related to the climate change scenario might favour a distribution shift in Fusarium species and the presence of these toxins in Southern European countries. In this paper, we present an ambitious work to determine the species responsible for trichothecene contamination in Spanish oats and to compare the results in the United Kingdom (UK) using a metataxonomic approach applied to both oat grains and soil samples collected from both countries. Regarding T-2 and HT-2 toxin producers, F. langsethiae was detected in 38% and 25% of the oat samples from the UK and Spain, respectively, and to the best of our knowledge, this is the first report of the detection of this fungus in oats from Spain. The relevant type B trichothecene producer, F. poae, was the most frequently detected Fusarium species in oats from both origins. Other important trichothecene producers, such as the Fusarium tricinctum species complex or Fusarium cerealis, were also frequently detected in oat fields. Many Fusarium toxins, including T-2 and HT-2 toxins, deoxynivalenol, or nivalenol, were detected in oat samples. The results obtained in this work revealed a clear change in the distribution of trichothecene producers and the necessity to establish the potential of these species to colonize oats and their ability to produce mycotoxins.

Different Resistance to DON versus HT2 + T2 Producers in Nordic Oat Varieties.

Over recent decades, the Norwegian cereal industry has had major practical and financial challenges associated with the occurrence of Fusarium head blight (FHB) pathogens and their associated mycotoxins in cereal grains. Deoxynivalenol (DON) is one of the most common Fusarium-mycotoxins in Norwegian oats, however T-2 toxin (T2) and HT-2 toxin (HT2) are also commonly detected. The aim of our study was to rank Nordic spring oat varieties and breeding lines by content of the most commonly occurring Fusarium mycotoxins (DON and HT2 + T2) as well as by the DNA content of their respective producers. We analyzed the content of mycotoxins and DNA of seven fungal species belonging to the FHB disease complex in grains of Nordic oat varieties and breeding lines harvested from oat field trials located in the main cereal cultivating district in South-East Norway in the years 2011-2020. Oat grains harvested from varieties with a high FHB resistance contained on average half the levels of mycotoxins compared with the most susceptible varieties, which implies that choice of variety may indeed impact on mycotoxin risk. The ranking of oat varieties according to HT2 + T2 levels corresponded with the ranking according to the DNA levels of Fusarium langsethiae, but differed from the ranking according to DON and Fusarium graminearum DNA. Separate tests are therefore necessary to determine the resistance towards HT2 + T2 and DON producers in oats. This creates practical challenges for the screening of FHB resistance in oats as today's screening focuses on resistance to F. graminearum and DON. We identified oat varieties with generally low levels of both mycotoxins and FHB pathogens which should be preferred to mitigate mycotoxin risk in Norwegian oats.

Determination of T-2 and HT-2 Toxins in Seed of Milk Thistle [Silybum marianum (L.) Gaertn.] Using Immunoaffinity Column by UPLC-MS/MS.

Milk thistle [Silybum marianum (L.) Gaertn.] achieved a significant increase in interest over the past few years from local and foreign pharmaceutical corporations. The silymarin complex of constituents extracted from milk thistle achenes provides compelling health benefits primarily thanks to antioxidant activities and hepatoprotective effects. However, consuming mycotoxin-contaminated plant material can cause immunosuppression and hepatotoxic problems. The aim of this study was to develop and validate a method for the determination of mycotoxin content in milk thistle. Fusarium toxins as T-2 and HT-2 toxins in grown milk thistle harvested from a breeding station in the Czech Republic during 2020-2021 were studied. The analysis of T-2 and HT-2 toxins was performed by UPLC-MS/MS after immunoaffinity columns EASI-EXTRACT® T-2 & HT-2 clean up. All analysed samples of milk thistle were contaminated with T-2 toxin and HT-2 toxin. The content of T-2 toxin in the samples from 2020 was in the range of 122.7-290.2 µg/kg and HT-2 toxin 157.0-319.0 µg/kg. In 2021, the content of T-2 toxin was in the range of 28.8-69.9 µg/kg and HT-2 toxin was 24.2-75.4 µg/kg. The results show that the climatic conditions of the year of harvesting have a highly statistically significant effect on the content of T-2 and HT-2 toxins in milk thistle.

T-2 and HT-2 toxins in harvested oat grains and their prevalence in whole grain flour during storage.

Mycotoxin contamination of food is a constant global concern. There has been a scientific debate in Europe on the validation of accredited detection methods for type A trichothecenes T-2 and HT-2 and the restriction on dangerous concentrations. The issue is of great importance as this type of mycotoxin is frequently found in spring cereals grown in Lithuania. The aim of this study was to optimise and validate a method for the determination of T-2/HT-2 toxin concentrations in oats harvested in 2015-2018 and to observe the changes in the concentrations of both toxins in oat flour during 3- and 6-week storage at different temperatures and increased relative air humidity. All of the oat grain samples (100%) collected in 2015-2018 tested positive for contamination with type A trichothecenes. The highest mean co-contamination by T-2 + HT-2 (260.4 ± 140.9 µg/kg) and the highest concentration (594.6 µg/kg) were determined in 2018 when warm and wet weather conditions prevailed during oat flowering. The effect of long-term storage (6 weeks) on T-2 and HT-2 toxin production manifested itself only when the samples had been stored under cooler conditions (8 °C). The most important factors which impacted the variation of the concentrations of type A trichothecenes in flour were ambient temperature and storage time.

Accumulation of HT-2 toxin from contaminated mushroom compost by edible Agaricus bisporus.

Wheat straw is commonly used as a cellulose source in mushroom compost and could be a secondary source of mycotoxin contamination in the food chain. We cultivated edible Agaricus bisporus and Pleurotus ostreatus on T-2/HT-2 artificially-contaminated mushroom compost and developed and in-house validated an UHPLC-MS/MS method for determination of T-2, HT-2, T2-triol and T2-tetraol in mushroom compost and mushroom basidiocarp. A rapid phase I metabolization of T-2 and HT-2 in mushroom compost was observed. In Agaricus bisporus, basidiocarps 8-15 µg kg-1 accumulation of HT-2 calculated on wet weight was measured. No detectable mycotoxins were found in Pleurotus ostreatus basidiocarp.

Reduction of deoxynivalenol, T-2 and HT-2 toxins and associated Fusarium species during commercial and laboratory de-hulling of milling oats.

Oats (Avena sativa L.) are well known for their nutritional properties but are susceptible to the growth of different Fusarium fungi resulting in mycotoxin contamination of harvested oats. In this study, oat samples from harvest years 2011 to 2017 were preselected for their suitability as milling oats for food purposes with DON contents below 1750 µg/kg. The reduction of DON, T-2 and HT-2 toxins during the commercial de-hulling process was analysed. While the average reduction for the sum of T-2 and HT-2 toxins in large oat kernels was 85%, the reduction for thin kernels was 66%. The reduction for DON was about 60% and did not differ for the two kernel fractions. In laboratory de-hulling experiments, milling oat samples and de-hulled oat kernels with known DON, T-2 and HT-2 toxin content were correlated with the associated DNA amount of Fusarium graminearum, Fusarium culmorum and Fusarium langsethiae. The reduction of the Fusarium DNA amount after de-hulling was comparable to the reduction of the associated mycotoxins. Notably, the correlation between F. langsethiae DNA amounts and the sum of T-2 and HT-2 toxin contents was R2 = 0.69 in milling oats and it rose to R2 = 0.85 in de-hulled oat kernels. In laboratory tests, at least one third of the initial levels of DON and the sum of T-2 and HT-2 toxins could be removed by polishing off the first parts of the outer layers; two thirds remained in the polished oat kernels. These observations indicate that de-hulling alone may not be completely sufficient to remove mycotoxin contamination in oats. These findings are of high importance in the discussion of determining legal maximum levels for DON or the sum of T-2 and HT-2 toxins in intermediate and final products.

Potential Role of Individual and Combined Effects of T-2 Toxin, HT-2 Toxin and Neosolaniol on the Apoptosis of Porcine Leydig Cells.

T-2 toxin usually co-occurs with HT-2 toxin and neosolaniol (NEO) in the grains and feed. Our previous studies found that T-2 toxin and its metabolites' binary or ternary combination exposure to porcine Leydig cells (LCs) displayed synergism in certain range of dosage and cannot be predicted based on individual toxicity. However, the possible mechanism of these mycotoxins' combined exposure to cell lesions remains unknown. Based on 50% cell viability, the mechanism of apoptosis in porcine Leydig cells was investigated after exposure to T-2, HT-2, NEO individual and binary or ternary combinations. Compared with control, the adenosine triphosphate (ATP) content decreased, reactive oxygen species (ROS) level increased, and mitochondrial membrane potential (MMP) decreased in all treated groups. Additionally, the cell apoptosis rates were significantly increased in test groups (p < 0.05), and the B-cell lymphoma 2 (Bcl-2) Associated X (Bax)/Bcl-2 ratio and the expression of caspase 3, caspase 8, cytochrome c (Cytc) in the treated group are all significantly higher than the control group. Moreover, the expression of Cytc and caspase 8 gene in NEO and T-2+NEO groups was significantly higher than that in other individual and combined groups. It can be concluded that the toxicities of T-2, HT-2, and NEO individually and in combination can induce apoptosis related to the oxidative stress and mitochondrial damage, and the synergistic effect between toxins may be greater than a single toxin effect, which is beneficial for assessing the possible risk of the co-occurrences in foodstuffs to human and animal health.

Human Biomonitoring of T-2 Toxin, T-2 Toxin-3-Glucoside and Their Metabolites in Urine through High-Resolution Mass Spectrometry.

The metabolic profile of T-2 toxin (T-2) and its modified form T-2-3-glucoside (T-2-3-Glc) remain unexplored in human samples. Therefore, the present study aimed to investigate the presence of T-2, T-2-3-Glc and their respective major metabolites in human urine samples (n = 300) collected in South Italy through an ultra-high performance liquid chromatography (UHPLC) coupled to Q-Orbitrap-HRMS methodology. T-2 was quantified in 21% of samples at a mean concentration of 1.34 ng/mg Crea (range: 0.22-6.54 ng/mg Crea). Almost all the major T-2 metabolites previously characterized in vitro were tentatively found, remarking the occurrence of 3'-OH-T-2 (99.7%), T-2 triol (56%) and HT-2 (30%). Regarding T-2-3-Glc, a low prevalence of the parent mycotoxin (1%) and its metabolites were observed, with HT-2-3-Glc (17%) being the most prevalent compound, although hydroxylated products were also detected. Attending to the large number of testing positive for T-2 or its metabolites, this study found a frequent exposure in Italian population.

Comparative Analysis of Machine Learning Methods to Predict Growth of F. sporotrichioides and Production of T-2 and HT-2 Toxins in Treatments with Ethylene-Vinyl Alcohol Films Containing Pure Components of Essential Oils.

The efficacy of ethylene-vinyl alcohol copolymer films (EVOH) incorporating the essential oil components cinnamaldehyde (CINHO), citral (CIT), isoeugenol (IEG), or linalool (LIN) to control growth rate (GR) and production of T-2 and HT-2 toxins by Fusarium sporotrichioides cultured on oat grains under different temperature (28, 20, and 15 °C) and water activity (aw) (0.99 and 0.96) regimes was assayed. GR in controls/treatments usually increased with increasing temperature, regardless of aw, but no significant differences concerning aw were found. Toxin production decreased with increasing temperature. The effectiveness of films to control fungal GR and toxin production was as follows: EVOH-CIT > EVOH-CINHO > EVOH-IEG > EVOH-LIN. With few exceptions, effective doses of EVOH-CIT, EVOH-CINHO, and EVOH-IEG films to reduce/inhibit GR by 50%, 90%, and 100% (ED50, ED90, and ED100) ranged from 515 to 3330 µg/culture in Petri dish (25 g oat grains) depending on film type, aw, and temperature. ED90 and ED100 of EVOH-LIN were >3330 µg/fungal culture. The potential of several machine learning (ML) methods to predict F. sporotrichioides GR and T-2 and HT-2 toxin production under the assayed conditions was comparatively analyzed. XGBoost and random forest attained the best performance, support vector machine and neural network ranked third or fourth depending on the output, while multiple linear regression proved to be the worst.

Water and temperature relations of Fusarium langsethiae strains and modelling of growth and T-2 and HT-2 mycotoxin production on oat-based matrices.

In the UK and Northern Europe, ripening oats can become contaminated with T-2 and HT-2 mycotoxins, produced mainly by Fusarium langsethiae. There are indicative levels related to the maximum limits for oat grain for these toxins. The objectives of this study were to examine the effect of interacting conditions of temperature (10-30 °C) and water activity (aw, 0.995-0.90) on (a) lag times prior to growth, (b) growth and (c) T-2 and HT-2 toxins by two strains of F. langsethiae isolated from oats in the UK and compare this with the type strain (Fl201059) which has been genomically sequenced, and (d) develop (and validated with published data) a probabilistic models for impacts of temperature × aw on growth and toxin production. All three strains had an optimum aw range and temperature of 0.995-0.98 and 25 °C for growth. For T-2 + HT-2 production these were 0.995 aw and 20 °C. Overall, the type strain produced higher amounts of T-2 + HT-2 with a HT-2/T-2 ratio of up to 76. Using this study data sets and those from the literature, probabilistic models were developed and validated for growth and T-2 + HT-2 toxin production in relation to temperature × aw conditions. These models, when applied in stored oats, will be beneficial in determining the conditions on the relative level of risk of contamination with these two toxins in the context of the EU indicative maximum levels.

In-utero exposure to HT-2 toxin affects meiotic progression and early oogenesis in foetal oocytes by increasing oxidative stress.

HT-2 toxin (HT-2), a mycotoxin produced by Fusarium species, is detected in a variety of cereal grain-based human food and animal feed. Apart from its well-established immunotoxicity and haematotoxicity, it also causes reproductive disorders. In the present study, we revealed the adverse effects of HT-2 on early oogenesis at the foetal stage. Pregnant mice were orally administered with HT-2 for 3 days at mid-gestation. Oocytes from female foetuses exposed to HT-2 displayed defects in meiotic prophase, including unrepaired DNA damage, elevated recombination levels, and reduced expression of meiotic-related genes. Subsequently, increased oxidative stress was observed in the foetal ovaries exposed to HT-2, along with the elevated levels of reactive oxygen species, malondialdehyde, catalase, and superoxide dismutase 1/2, thereby resulting in impaired mitochondrial membrane potential and cell apoptosis. Furthermore, pre-treatment with urolithin A, a natural compound with antioxidant activities, partially reversed the delayed meiotic process by alleviating oxidative stress. Since early oogenesis is essential to determine female fertility in adult life, this study indicated that brief maternal exposure to HT-2 toxin may compromise the fertility of a developing female foetus.

Reduction of T-2 and HT-2 mycotoxins by atmospheric cold plasma and its impact on quality changes and germination of wheat grains.

Wheat (Triticum aestivum) is susceptible to mycotoxin contamination, which can result in significant health risks and economic losses. This research examined the ability of air atmospheric cold plasma (air-ACP) treatment to reduce pure and spiked T-2 and HT-2 mycotoxins' concentration on wheat grains. This study also evaluated the effect of ACP treatment using different gases on wheat grain germination parameters. The T-2 and HT-2 mycotoxin solutions applied on round cover-glass were placed on microscopy slides and wheat grains (0.5 g) were individually spiked with T-2 and HT-2 on their surfaces. Samples were then dried at room temperature (∼24 °C) and treated by air-ACP for 1 to 10 min. Ten minutes of air-ACP treatment significantly reduced pure T-2 and HT-2 concentrations by 63.63% and 51.5%, respectively. For mycotoxin spiked on wheat grains, 10 min air-ACP treatment significantly decreased T-2 and HT-2 concentrations up to 79.8% and 70.4%, respectively. No significant change in the measured quality and color parameters was observed in the ACP-treated samples. Wheat grain germination parameters were not significantly different, when treated with ACP using different gases. Air-ACP treatment and ACP treatment using 80% nitrogen + 20% oxygen improved the germination of wheat grains by 10% and 6%, respectively. This study demonstrated that ACP is an innovative technology with the potential to improve the safety of wheat grains by reducing T-2/HT-2 mycotoxins with an additional advantage of improving their germination. PRACTICAL APPLICATION: Atmospheric cold plasma (ACP) technology has a huge potential to degrade mycotoxins in food grains. This study evaluated the efficacy of ACP to reduce two major mycotoxins (T-2 and HT-2 toxins) in wheat grains. The results of this study will help to develop and scale-up the ACP technology for mycotoxin degradation in grains.

HT-2 toxin affects cell viability of goat spermatogonial stem cells through AMPK-ULK1 autophagy pathways.

HT-2 toxin is widely found in moldy crops and is the major metabolite of T-2 toxin, which has been shown to exert various toxic effects in farm animals. However, little is known about the effects of HT-2 toxin on male reproduction, particularly spermatogenesis. This study aims to investigate the toxic effects of HT-2 toxin on goat spermatogonial stem cells (SSCs) and related autophagy-regulated mechanisms. Our results showed that HT-2 toxin exposure resulted in decreased cell viability and proliferation, disrupted SSCs self-renewal, and reduced germ cell-related gene expression. HT-2 toxin exposure also induced oxidative stress and cell apoptosis, as shown by ROS accumulation, increased antioxidant enzyme activity levels, decreased the mitochondrial membrane potential, and increased caspase-9 mRNA and Bcl/bax protein levels. Additionally, HT-2 toxin exposure increased the expression of the autophagy-inducing genes Atg5, Atg7 and Beclin1 and the number of autophagosomes, which indicated that HT-2 toxin induced autophagy in the goat SSCs. Moreover, we also examined a possible mechanism by which HT-2 toxin exposure induced higher expression of AMPK, mTOR and ULK at both the mRNA and protein levels. our results indicated that HT-2 toxin caused apoptosis and autophagy by activating AMPK-mTOR-ULK1 pathway, which further affected SSCs viability.

A Two-Year Occurrence of Fusarium T-2 and HT-2 Toxin in Croatian Cereals Relative of the Regional Weather.

To investigate into the T-2 and HT-2 toxin occurrence, 240 samples of unprocessed cereals (maize, wheat, barley, and oats) were sampled from different fields located in three Croatian regions during 2017-2018. In all samples, sum concentrations of T-2/HT-2 toxin were determined using the ELISA method, while the LC-MS/MS was used as a confirmatory method for both mycotoxins in positive samples (>LOD) and the establishment of T-2 over HT-2 toxin ratios. The results showed oats to be the most contaminated cereal, with T-2/HT-2 toxins detected in 70.0% of samples, followed by barley (40.9%), maize (26.8%) and wheat (19.2%), with the mean T-2/HT-2 ratio ranging from 1:2.7 in maize to 1:4.4 in oats. Sum T-2/HT-2 concentrations in two maize samples were higher than the indicative level recommended by the European Commission, necessitating subsequent investigations into the conditions under which these poorly investigated mycotoxins are produced. Statistically significantly (p < 0.05) higher concentrations of T-2/HT-2 toxin were determined in oats throughout study regions as compared to those found in wheat, but not maize and barley, while the concentrations of these mycotoxins were related to the regional weather in Croatia.

Development and validation of a liquid chromatography tandem mass spectrometry multi-method for the determination of 41 free and modified mycotoxins in beer.

A fast high performance liquid chromatography tandem mass spectrometry multi-method based on an ACN-precipitation extraction was developed for the analysis of 41 (modified) mycotoxins in beer. Validation according to the performance criteria defined by the European Commission (EC) in Commission Decision no. 657/2002 revealed good linearity (R2 > 0.99), repeatability (RSDr < 15%), reproducibility (RSDR < 15%), and recovery (79-100%). Limits of quantification ranging from 0.04 to 75 µg/L were obtained. Matrix effects varied from -67 to +319% and were compensated for using standard addition. In total, 87 beer samples, produced worldwide, were analyzed for the presence of mycotoxins with a focus on modified mycotoxins, whereof 76% of the samples were contaminated with at least one mycotoxin. The most prevalent mycotoxins were deoxynivalenol-3-glucoside (63%), HT-2 toxin (15%), and tenuazonic acid (13%). Exposure estimates of deoxynivalenol and its metabolites for German beer revealed no significant contribution to intake of deoxynivalenol.