Natural Occurrence of Beauvericin and Enniatins in Corn- and Wheat-Based Samples Harvested in 2017 Collected from Shandong Province, China.
ABSTRACT: Totals of 158 corn and corn-based samples and 291 wheat and wheat-based samples from Shandong province, China in 2017 were analyzed for five mycotoxins including beauvericin (BEA), enniatin A (ENA), enniatin A? (ENA?), enniatin B (ENB), and enniatin B? (ENB?) by high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS). BEA was the predominant toxin detected, followed by ENB, ENA?, ENA, and ENB?. Corn and corn-based samples were more easily contaminated by BEA with an average concentration of 65.26 µg/kg, compared with that in wheat and wheat-based samples (average = 0.41 µg/kg). Concentrations of BEA, ENA, and ENB? in corn kernels, flours, and flakes were significantly different (Kruskal?Wallis Test, p < 0.05), as well as for BEA, ENA, ENB, and ENB? in wheat kernels, flours, and noodles (Kruskal?Wallis test, p < 0.05). Furthermore, 59.5% (94/158) and 59.8% (174/291) corn- and wheat-based samples were co-contaminated by at least two mycotoxins, respectively. Positive correlations in concentrations were observed in corn between levels of ENA and ENB?, ENA and ENB, ENA? and ENB?, as well as in wheat between BEA and ENA, BEA and ENA?, BEA and ENB, BEA and ENB?, ENA and ENA?, ENA and ENB, ENA and ENB?, ENA? and ENB, ENA? and ENB?, and ENB and ENB?. These results demonstrate that co-contamination of BEA and enniatins (ENNs) in corn- and wheat-based samples from Shandong, China is very common. More data on the contamination of five mycotoxins in cereal and cereal-based samples nationwide are needed.
Project description:A total of 470 edible vegetable oil samples including peanut, soybean, rapeseed, sesame seed, corn, blend, and others collected from eight provinces of China were analyzed for the concentrations of beauvericin (BEA), enniatin A (ENA), A1 (ENA1), B (ENB), and B1 (ENB1) by ultraperformance liquid chromatography/electrospray ionization tandem mass spectrometry (UPLC/ESI-MS/MS). Concentrations of BEA, ENB, and ENB1 (average = 5.59 μg/kg, 5.16 μg/kg, and 4.61 μg/kg) in all positive samples were higher than those for ENA and ENA1 (average = 0.85 μg/kg and 1.88 μg/kg). Frequencies of BEA and ENNs in all analyzed samples were all higher than 50% with the exception of ENA1 (36.6%, 172/470). Levels of BEA and ENNs in all analyzed samples varied based on their sample types and geographical distributions (Kruskal–Wallis test, p < 0.05). The soybean and peanut oil samples were found to be more easily contaminated by BEA and ENNs than other oil samples. Concentrations of BEA and ENNs in samples obtained from Heilongjiang, Shandong and Guizhou were higher than those found in samples from other provinces. Besides, frequencies of mycotoxin co-contaminations were high and their co-contamination types also varied by oil types. BEA-ENA-ENA1-ENB-ENB1 was the most commonly found toxin combination type, almost in one third of the analyzed samples (30%, 141/470). Overall, these results indicate that co-occurrence of BEA and ENNs in analyzed Chinese edible vegetable oil samples is highly common, and it is vital to monitor them, both simultaneously and on a widespread level.
Project description:In this study, a rapid multi-mycotoxin approach was developed for biomonitoring and quantification of 27 important mycotoxins and mycotoxin metabolites in human blood samples. HPLC-MS/MS detection was used for the analysis of dried serum spots (DSS) and dried blood spots (DBS). Detection of aflatoxins (AFB<sub>1</sub>, AFB<sub>2</sub>, AFG<sub>1</sub>, AFG<sub>2</sub>, AFM<sub>1</sub>), trichothecenes (deoxynivalenol, DON; DON-3-glucoronic acid, DON-3-GlcA; T-2; HT-2; and HT-2-4-GlcA), fumonisin B<sub>1</sub> (FB<sub>1</sub>), ochratoxins (OTA and its thermal degradation product 2'R-OTA; OT?; 10-hydroxychratoxin A, 10-OH-OTA), citrinin (CIT and its urinary metabolite dihydrocitrinone, DH-CIT), zearalenone and zearalanone (ZEN, ZAN), altenuene (ALT), alternariols (AOH; alternariol monomethyl ether, AME), enniatins (EnA, EnA<sub>1</sub>, EnB, EnB<sub>1</sub>) and beauvericin (Bea) was validated for two matrices, serum (DSS), and whole blood (DBS). HPLC-MS/MS analysis showed signal suppression as well as signal enhancement due to matrix effects. However, for most analytes LOQs in the lower pg/mL range and excellent recovery rate were achieved using matrix-matched calibration. Besides validation of the method, the analyte stability in DBS and DSS was also investigated. Stability is a main issue for some analytes when the dried samples are stored under common conditions at room temperature. Nevertheless, the developed method was applied to DBS samples of a German cohort (n?=?50). Besides positive findings of OTA and 2'R-OTA, all samples were positive for EnB. This methodical study establishes a validated multi-mycotoxin approach for the detection of 27 mycotoxins and metabolites in dried blood/serum spots based on a fast sample preparation followed by sensitive HPLC-MS/MS analysis. Graphical Abstract ?.
Project description:Aromatic and medicinal plants (AMPs), as herbal material, are subjected to contamination by various mycotoxin-producing fungi, either free and conjugated. Such a problem is associated with poor storage practices, and lack of adopting good agricultural practices and good harvesting practices. Nevertheless, AMPs are poorly investigated. The purpose of this study was to investigate the co-occurrence of 15 mycotoxins (four aflatoxins (AFB1, AFB2, AFG1, and AFG2), ochratoxin A (OTA), beauvericin (BEA), four enniatins (ENA, ENA1, ENB, and ENB1), zearalenone (ZEN), alternariol (AOH), tentoxin (TENT), T-2, and HT-2 toxins) in 40 samples of AMPs frequently consumed in Morocco by using liquid chromatography tandem mass spectrometry. Evaluation of conjugated mycotoxins and their identification using liquid chromatography coupled to time-of-flight mass spectrometry with ion mass exact was also carried out. Results showed that 90% of the analyzed samples presented at least one mycotoxin, and 52% presented co-occurrence of them. Mycotoxins detected were: AOH (85%), ZEN (27.5%), ?-ZEL (22%), AFG1 (17.5%), TENT (17.5%), ENB (10%), AFG2 (7.5%), ?-ZEL (5%), ENA1 (2.5%), and HT-2 (2.5%), while the conjugated mycotoxins were ZEN-14-Glc (11%) and ZEN-14-Sulf (9%). The highest observed level was for AOH, with 309 ng/g. Ten samples exceeded the recommended levels set by the European Pharmacopoeia for AF mycotoxins in plant material (4 ng/g), and three samples exceeded the maximum limits for AFs (10 ng/g) in species established by the European Commission. Although the co-occurrence of several mycotoxins in AMP samples was observed, the dietary exposure assessment showed that the intake of mycotoxins through the consumption of AMP beverages does not represent a risk for the population.
Project description:Beauvericin (BEA) and enniatins (ENNs) are cyclic peptide mycotoxins produced by a wide range of fungal species, including pathogenic Fusaria. Amounts of BEA and ENNs were quantified in individual rice cultures of 58 Fusarium strains belonging to 20 species, originating from different host plant species and different geographical localities. The species identification of all strains was done on the basis of the tef-1? gene sequence. The main aim of this study was to analyze the variability of the esyn1 gene encoding the enniatin synthase, the essential enzyme of this metabolic pathway, among the BEA- and ENNs-producing genotypes. The phylogenetic analysis based on the partial sequence of the esyn1 gene clearly discriminates species producing exclusively BEA from those synthesizing mainly enniatin analogues.
Project description:Consumption of fruit juice is becoming trendy for consumers seeking freshness and high vitamin and low caloric intake. Mycotoxigenic moulds may infect fruits during crop growth, harvest, and storage leading to mycotoxin production. Many mycotoxins are resistant to food processing, which make their presence in the final juice product very likely expected. In this way, the presence of 30 mycotoxins including aflatoxin B1 (AFB1), aflatoxin B2 (AFB2), aflatoxin G1 (AFG1), aflatoxin G2 (AFG2), alternariol (AOH), alternariol monomethyl ether (AME), Ochratoxin A (OTA), fumonisin B1 (FB1), fumonisin B2 (FB2), enniatin A (ENNA), enniatin A1 (ENNA1), enniatin B (ENNB), enniatin B1 (ENNB1), beauvericin (BEA), sterigmatocystin (STG), zearalenone (ZEA), ?-zearalanol (?-ZAL), ?-zearalanol (?-ZAL), ?-zearalenol (?-ZOL), ?-zearalenol (?-ZOL), deoxynivalenol (DON), 3-acetyl-deoxynivalenol (3-ADON), 15-acetyl-deoxynivalenol (15-ADON), diacetoxyscirpenol (DAS), nivalenol (NIV), fusarenon-X (FUS-X), neosolaniol (NEO), patulin (PAT), T-2 toxin and HT-2 toxin was evaluated in 80 juice samples collected from Valencia retail Market. An efficient Dispersive Liquid-Liquid Microextraction method (DLLME) was carried out before their trace level determination by chromatographic techniques coupled to tandem mass spectrometry. The results obtained revealed the presence of nine mycotoxins namely AOH, AME, PAT, OTA, AFB1, AFB2, AFG2, ?-ZAL, and HT2 in the analyzed samples, with incidences ranging from 3 to 29% and mean contents between 0.14 and 59.52 µg/L. Considerable percentages of TDIs were reached by children when 200 mL was considered as daily fruit juice intake.
Project description:Members of the fungal genus Fusarium can produce numerous secondary metabolites, including the nonribosomal mycotoxins beauvericin (BEA) and enniatins (ENNs). Both mycotoxins are synthesized by the multifunctional enzyme enniatin synthetase (ESYN1) that contains both peptide synthetase and S-adenosyl-l-methionine-dependent N-methyltransferase activities. Several Fusarium species can produce ENNs, BEA or both, but the mechanism(s) enabling these differential metabolic profiles is unknown. In this study, we analyzed the primary structure of ESYN1 by sequencing esyn1 transcripts from different Fusarium species. We measured ENNs and BEA production by ultra-performance liquid chromatography coupled with photodiode array and Acquity QDa mass detector (UPLC-PDA-QDa) analyses. We predicted protein structures, compared the predictions by multivariate analysis methods and found a striking correlation between BEA/ENN-producing profiles and ESYN1 three-dimensional structures. Structural differences in the ? strand's Asn789-Ala793 and His797-Asp802 portions of the amino acid adenylation domain can be used to distinguish BEA/ENN-producing Fusarium isolates from those that produce only ENN.
Project description:The levels of 26 mycotoxins were determined in 147 samples of the grain of cereals cultivated in five regions of Poland during the 2014 growing season. The HPLC-HRMS (time-of-flight) analytical technique was used. An analytical procedure to simultaneously determine 26 mycotoxins in grain was developed, tested and verified. Samples from eastern and southern Poland were more contaminated with mycotoxins than the samples from northern and western Poland. Toxins produced by Fusarium fungi were the main contaminants found. Some deoxynivalenol (DON) was found in 100% of the tested samples of wheat (Osiny, Borusowa, Werbkowice), triticale, winter barley and oats, while the maximum permissible DON level (as defined in the EU Commission Regulation No. 1881/2006) was exceeded in 10 samples. Zearalenone (ZEN), DON metabolites and enniatins were also commonly found. The presence of mycotoxins in grain reflected the prevailing weather conditions during the plant flowering/earing stages, which were favorable for the development of blight. Among all investigated wheat genotypes, cv. Fidelius was the least contaminated, while Bamberka, Forkida and Kampana were the most contaminated. However, the single-factor ANOVA analysis of variance did not reveal (at a statistical significance level ? = 0.05) any differences between levels of mycotoxins in individual genotypes. Triticale was the most contaminated grain among all of the tested varieties. ZEN, DON and the sum of 3-acetyldexynivalenol and 15-acetyldeoxynivalenol (3- and 15-ADON) were found in 100% of the tested triticale samples at concentrations within the 4-86, 196-1326 and 36-374 µg·kg(-1) range, respectively. Of particular concern was the fact that some "emerging mycotoxins" (enniatins) (in addition to commonly-known and legally-regulated mycotoxins) were also found in the tested triticale samples (enniatin B (Enn-B), enniatin B1 (Enn-B1), enniatin A-1 (Enn-A1), 100% of samples, and enniatin A (Enn-A), 70% of samples). Depending on the toxin, they were found at levels between 8 and 3328 µg·kg(-1).
Project description:In this study, the occurrence of multiple fungal metabolites including mycotoxins was determined in four different winter wheat varieties in a field experiment in Croatia. One group was naturally infected, while the second group was inoculated with a Fusarium graminearum and F. culmorum mixture to simulate a worst-case infection scenario. Data on the multiple fungal metabolites including mycotoxins were acquired with liquid chromatography with mass spectrometry (LC-MS/MS) multi-(myco)toxin method. In total, 36 different fungal metabolites were quantified in this study: the Fusarium mycotoxins deoxynivalenol (DON), DON-3-glucoside (D3G), 3-acetyldeoxynivalenol (3-ADON), culmorin (CULM), 15-hydroxyculmorin, 5-hydroxyculmorin, aurofusarin, rubrofusarin, enniatin (Enn) A, Enn A1, Enn B, Enn B1, Enn B2, Enn B3, fumonisin B1, fumonisin B2, chrysogin, zearalenone (ZEN), moniliformin (MON), nivalenol (NIV), siccanol, equisetin, beauvericin (BEA), and antibiotic Y; the Alternaria mycotoxins alternariol, alternariolmethylether, altersetin, infectopyron, tentoxin, tenuazonic acid; the Aspergillus mycotoxin kojic acid; unspecific metabolites butenolid, brevianamid F, cyclo(L-Pro-L-Tyr), cyclo(L-Pro-L-Val), and tryptophol. The most abundant mycotoxins in the inoculated and naturally contaminated samples, respectively, were found to occur at the following average concentrations: DON (19,122/1504 µg/kg), CULM (6109/1010 µg/kg), 15-hydroxyculmorin (56,022/1301 µg/kg), 5-hydroxyculmorin (21,219/863 µg/kg), aurofusarin (43,496/1266 µg/kg). Compared to naturally-infected samples, Fusarium inoculations at the flowering stage increased the concentrations of all Fusarium mycotoxins, except enniatins and siccanol in Ficko, the Aspergillus metabolite kojic acid, the Alternaria mycotoxin altersetin, and unspecific metabolites brevianamid F, butenolid, cyclo(L-Pro-L-Tyr), and cyclo(L-Pro-L-Val). In contrast to these findings, because of possible antagonistic actions, Fusarium inoculation decreased the concentrations of the Alternaria toxins alternariol, alternariolmethylether, infectopyron, tentoxin, tenuazonic acid, as well as the concentration of the nonspecific metabolite tryptophol.
Project description:Mycotoxins are secondary metabolites produced by a variety of fungi that contaminate food and feed resources, and are capable of inducing a wide range of toxicity. Here, we studied the developmental and behavioral toxicity in zebrafish (Danio rerio) embryos and larvae exposed to three mycotoxins: beauvericin (BEA), Enniatin A (ENN A), and Ennitain B (ENN B). Zebrafish embryos were collected after fertilization, treated individually from 1 to 6 dpf with BEA at 8, 16, 32 and, 64 μM and for both enniatins at 3.12, 6.25, 12.5 and, 25 μM. Mixture of mycotoxins were assayed as follows: i) for BEA + ENN A and BEA + ENN B at [32 + 12.5] μM and [16 + 6.25] μM; ii) for ENN A + ENN B at [12.5 + 12.5] μM and [6.25 + 6.25] μM and, iii) for BEA + ENN A + ENN B at [32 + 12.5 + 12.5] μM and [16 + 6.25 + 6.25] μM. Response was collected after a white light-flash intermittent coming on for 5 s during 2 h with a imaging platform. Outcomes measured were: time to death, response to light, and circadian rhythm. This last outcome was measured in a plate where embryos had evolved in natural intervals of light and dark until day 7 or in a plate maintained in darkness. Images of all stages and evolution were collected. Results indicated that mycotoxins induced toxicity at the concentrations tested. All exposed zebrafish induced developmental defects, specifically hatching time and motion activity. After exposure, fish showed enhanced baseline activity but they lost their responsiveness to light.
Project description:The exposure to mycotoxins of Swedish adolescents is currently unknown. The aim of the present study was to investigate the exposure to mycotoxins and their association with food intake, and background characteristics in adolescents of a national dietary survey. About 3000 school students (1000 from the 5th, 8th and 11th school years) were recruited for the survey. The participants completed Web-based questionnaires on food propensity, sociodemography and health, and a Web-based dietary recall. Spot urine and blood samples were collected from 1105 of the participants for mycotoxin biomarker analysis. Mycotoxins were analysed with multibiomarker methods in urine (HPLC-MS/MS) and serum (HPLC-MS/MS). Of the 35 different analytes in urine, the frequency of positive samples were the following: deoxynivalenol (DON, 4.8%), DON-15-?-D-O-glucuronide (DON-15GlcA, 9.1%), dihydro-citrinone (DH-CIT, 0.5%), HT-2-glucuronide (HT-2-3-GlcA, 0.1%) and ochratoxin A (OTA, 0.1%). Of the 27 different analytes in serum, OTA was detected in all samples, while 2'R-ochratoxin A (2'R-OTA) was found in 8.3% and enniatin B (EnB) in 99.2% of the samples. Exposure assessment calculations were performed on OTA from the serum concentration and on DON equivalents (DON eqv) from the urine concentration. All probable daily intake (PDI) estimates were below tolerable daily intakes, except for 1.6% of the participants for DON. The maximum PDI was 4.3 ?g DON eqv/kg body weight and day. Consumption of cereal grain commodities was associated with levels of DON, EnB or OTA in biofluids. Serum OTA was also associated with intakes of raisins and coffee. Furthermore, coffee consumption correlated well with 2'R-OTA concentration in serum. In conclusion, exposure to mycotoxins in Swedish adolescents is common, but fortunately, high exposure was rare.