Zearalenone In Grains Research Articles

Immunoenzymatic evaluation for aflatoxin, ochratoxin, and zearalenone in grains stored

Immunoenzymatic evaluation for aflatoxin, ochratoxin, and zearalenone in grains stored

Quantitative detection of zearalenone in wheat grains based on near-infrared spectroscopy

Zearalenone (ZEN) can easily contaminate wheat, seriously affecting the quality and safety of wheat grains. In this study, a near-infrared (NIR) spectroscopy detection method for rapid detection of ZEN in wheat grains was proposed. First, the collected original near-infrared spectra were denoised, smoothed and scatter corrected by Savitzky-Golay smoothing (SG-smoothing) and multiple scattering correction (MSC), and then normalized. Three wavelength variable selection algorithms were used to select variables from the preprocessed NIR spectra, which were random frog (RF), successive projections algorithm (SPA), least absolute shrinkage and selection operator (LASSO). Finally, based on the feature variables extracted by the above algorithms, support vector machine (SVM) models were established respectively to realize the quantitative detection of the ZEN in wheat grains. Eventually, the prediction effect of the LASSO-SVM model was the best, the prediction correlation coefficient (RP) was 0.99, the root mean square error of prediction (RMSEP) was 2.1 μg·kg−1, and the residual prediction deviation (RPD) was 6.0. This research shows that the NIR spectroscopy can be used for high-precision quantitative detection of the ZEN in grains, and the research gives a new technical solution for the in-situ detection of mycotoxins in stored grains.

A simple mesoporous silica nanoparticle-based fluorescence aptasensor for the detection of zearalenone in grain and cereal products.

Zearalenone (ZEN) is a type of estrogenic mycotoxin commonly occurring in cereals. The aim of this study was to design a simple, rapid, inexpensive and ultrasensitive fluorescence assay for the determination of ZEN. Here, amino-modified mesoporous silica nanoparticles (MSNs-NH2) were synthesized to be the positive charge-rich reactor. A 6-carboxy-fluorescein-labeled aptamer (aptamer-FAM) was designed as the signal probe, ZEN-capture probe and negative charge reactor. In the absence of ZEN, the negatively charged aptamer-FAM combined with the positively charged MSNs-NH2 in an electrostatic manner. In the presence of ZEN, the fluorescence intensity in the supernatant increased significantly because the aptamer-FAM could bind to ZEN instead of MSNs-NH2. Under the optimal experimental conditions, this assay exhibited excellent specificity, repeatability and a wide linearity range of 0.005-150ng/mL, with a detection limit of 0.012ng/mL. Additionally, it showed high recovery (83.3-101.5%) for the spiked samples. There was no statistically significant difference in the ZEN concentrations detected by the proposed assay and HPLC in naturally contaminated samples. Overall, this design provides a new strategy for the rapid, inexpensive and sensitive detection of ZEN, and it could be applied to develop fluorometric assays for different targets by the selection of appropriate aptamers. Graphical abstract.

Preparation of magnetic molecularly imprinted polymers for the identification of zearalenone in grains.

This study was based on the specific binding ability of magnetic molecularly imprinted polymers (MMIPs) combined with a high-performance liquid chromatography-fluorescence detector (HPLC-FLD) for the rapid determination of zearalenone (ZEN) in cereals. A novel magnetic molecularly imprinted polymer was prepared by surface imprinting technology. Warfarin was used as a virtual template, 3-aminopropyl triethoxysilane (APTES) was used as the functional monomer, and tetraethyl orthosilicate (TEOS) was used as the cross-linking agent. Analysis by a vibrating sample magnetometer (VSM), Fourier transform infrared spectroscopy (FT-IR), X-ray diffraction (XRD), thermogravimetric analysis (TGA), scanning electron microscopy (SEM), and transmission electron microscopy (TEM) showed that MMIPs were prepared with a particle size about 450nm, the imprinted molecular layer accounting for 10.7% of the total mass, and saturation magnetization of about 34.54emu/g. The maximum adsorption capacity (Qmax) of the thermodynamic and kinetic adsorption experiments were 13.90mg/g and 8.71mg/g, respectively. The Langmuir model showed that the binding sites were uniformly distributed on the surface of the MMIPs. The Scatchard analysis showed that MMIPs had two types of binding sites with Qmax of 8.22mg/g and 15.37mg/g, respectively. In actual sample detection, the limit of detection (LOD) and limit of quantification (LOQ) were 0.4ng/kg and 0.9ng/kg, respectively. The sample recovery rate was 90.56-99.96%, the daytime stability was 1.35-2.87%. These results showed that MMIPs had good performance in selectively identifying ZEN and were suitable for determining ZEN in cereals.

Quantum dot nanobead-based fluorescent immunochromatographic assay for simultaneous quantitative detection of fumonisin B1, dexyonivalenol, and zearalenone in grains

The mycotoxins in grains is a serious threat to the health of humans and animals. In this study, we demonstrated an approach for development of a fast and sensitive fluorescent immunochromatographic assay for fumonisin B1 (FB1), deoxynivalenol (DON) and zearalenone (ZEN) detection using quantum dots nanobeads (QDNBs, 100 nm). Amplification of the fluorescence signal of the quantum dot by nanosphere coating can significantly improve the sensitivity of immunochromatographic test strip. The detection probes (FB1-McAb@QDNBs, DON-McAb@QDNBs, and ZEN-McAb@QDNBs) were prepared using the QDNBs with 615 nm emissions coupling with three monoclonal antibodies (McAb) against FB1, DON, and ZEN, respectively, in a competitive immunochromatography platform. The surface of the QDNBs is modified with a carboxyl functional group, which facilitates the coupling of the antibody and significantly improves the sensitivity of test strip. Compared with traditional ELISA, the immunoassay based on the McAb@QDNBs is more sensitive, with IC50 of 12.66 ng/mL (FB1), 2.97 ng/mL (DON), and 0.87 ng/mL (ZEN), of which the linear range were from 2.295 ng/mL to 69.867 ng/mL (FB1), 0.465 ng/mL to 16.19 ng/mL (DON), and 0.295 ng/mL to 2.575 ng/mL (ZEN), respectively. The accuracy, repeatability, and specificity of the established test strip was evaluated by testing the spiked and natural samples. Furthermore, a high agreement was observed between the developed test strips and LC-MS/MS in simultaneously and quantitative detection of three mycotoxins. In brief, the developed approach was feasible and could be employed for simultaneously and quantitative mycotoxins detection in agricultural products.

Occurrence of zearalenone in grains and its reduction by gamma radiation

ABSTRACTPurpose: The purpose of the present study was to examine the reduction of Zearalenone (ZEA) in grains by using gamma radiation (γ- ray) to minimise exposure hazards and of economic and health consequence.Materials and Methods: A total of 150 samples of food grains (wheat, yellow corn, and white corn) were collected from Egyptian retail markets from five governorates (Cairo, Alexandria, Giza, Qena and Gharbia) to determine the ZEA contamination levels by high performance liquid chromatography (HPLC). To study reduction by γ- ray, twelve uncontaminated samples were spiked with ZEA at the maximum permitted limit (100 µg/kg) and irradiated with different energies of γ-rays (5, 10, 15 and 20 kGy), and then the reduction rates were evaluated.Results: Only two samples out of 150 randomly collected samples were found to be contaminated with ZEA. In the case of the effect of γ-rays on grain ZEA contents, the reduction in ZEA ranged between 49.0 ± 4.7 and 97.0 ± 1.5% in wheat samples, and between 24.8 ± 4.1 and 51.1 ± 3.5% in yellow corn samples, between 20.3 and 59.9% in white corn samples. Finally, at 20 kGy, the reductions were 97.0 ± 1.5, 51.1 ± 3.5 and 59.9 ± 3.2% in wheat, yellow corn and white corn, respectively.Conclusions: ZEA was more easily degraded in wheat than in yellow corn and white corn. Moisture content played a vital role in ZEA degradation. γ- Ray exposure was used as an effective method to reduce ZEA in wheat, yellow corn and white corn. The reduction rate increased relative to the irradiation dose. Further study is recommended to determine the reduction yields and toxicities of ZEA that may be achieved post γ- ray radiation.

One-step rapid detection of fumonisin B1, dexyonivalenol and zearalenone in grains

Mycotoxins are a class of secondary metabolites produced by filamentous fungi that are harmful to humans and animals. Here, a multiplex immunochromatographic assay was reported, which is based on 25 nm colloidal gold for simultaneous qualitative detection of fumonisin B1 (FB1), deoxynivalenol (DON) and zearalenone (ZEN) in wheat and corn samples. After conditions optimization, the immunochromatographic strip has high sensitivity. The visual detection limits of the immunochromatographic test strips for FB1, DON, and ZEN reached 60, 12.5 and 6 ng/mL, respectively. In addition, the immunochromatographic strip has high specificity, no reaction to each other, or no cross-reactivity with ochratoxin A (OTA) and aflatoxin B1 (AFB1). Furthermore, eighteen samples (including corn and wheat) were tested using the test strips and verified by liquid chromatography-mass spectrometry (LC-MS/MS). The results of strips were found to be consistent with those of liquid chromatography-mass spectrometry (LC-MS/MS). In short, this work demonstrates that the spherical colloidal gold-based immunochromatographic test strip we developed can be used to quickly and simultaneously monitor the existence of multiple mycotoxins in agricultural food samples.

Anti-idiotypic nanobody-phage display-mediated real-time immuno-PCR for sensitive, simultaneous and quantitative detection of total aflatoxins and zearalenone in grains

An anti-idiotypic nanobody-phage display-mediated immuno-polymerase chain reaction (PD-IPCR) method was developed for simultaneous quantitative detection of total aflatoxins and zearalenone in cereals. Two phages, displaying the variable domain of the heavy chain anti-idiotypic nanobody that binds aflatoxin- or zearalenone-specific monoclonal antibody (1C11 or 2D3), were used as competitors for corresponding analytes. Specific DNA sequences encoding anti-idiotypic nanobodies were used to design the primers for PCR amplification. The results indicated that detection limits for total aflatoxins and zearalenone in a sample were 0.03 and 0.09 ng mL−1, respectively. Recoveries of spiked aflatoxins and zearalenone were 80–118% and 76.7–111%, respectively. Validation results were in good agreement with the gold-standard high-performance liquid chromatography method. This report is the first to describe PD-IPCR for simultaneous quantitative detection of total aflatoxins and zearalenone in cereals.

Progress in bio-degradation of mycotoxin zearalenone

Zearalenone (ZEN) and its derivatives are non-steroidal estrogenic mycotoxins mainly produced by Fusarium species. They are widely distributed in grain feeds originated from maize, barley, wheat and sorghum, causing serious harm to animal and human health. Currently, there is a pressing need of an efficient technology for ZEN degradation and detoxification. Because traditional physical and chemical methods could not effectively detoxify ZEN in grains, and might also affect the grain nutrients and food taste, and even result in secondary pollution, the biological technologies are developed to detoxify ZEN and its derivatives. In this paper, we reviewed the structure of ZEN and its derivatives, the fungi and bacteria species with ability of degradation of ZEN. In addition, the characterization, protein sequences and conformation of currently identified ZEN degrading enzymes, the only solved ZHD structure from Clonostachys rose were analyzed and compared, and the enzymes heterologous expression and application were also reviewed. This review will provide reference for reducing the cost of ZEN degrading enzymes by biological technologies such as enzyme engineering and fermentation engineering.

RNA-seq based gene expression analysis of ovarian granulosa cells exposed to zearalenone in vitro: significance to steroidogenesis.

Zearalenone (ZEA) is a natural contaminant of various food and feed products representing a significant problem worldwide. Since the occurrence of ZEA in grains and feeds is frequent, the present study was carried out to evaluate the possible effects of ZEA on steroid production and gene expression of porcine granulosa cells, using RNA-seq analysis. Porcine granulosa cells were administered 10 μM and 30 μM ZEA during 72 h of culture in vitro. Following ZEA treatment the gene expression profile of control and exposed granulosa cells was compared using RNA-seq analysis. The results showed that in the exposed granulosa cells ZEA significantly altered the transcript levels, particularly steroidogenesis associated genes. Compared with the control group, 10 μM and 30 μM ZEA treatment significantly increased the mRNA expression of EDN1, IER3, TGFβ and BDNF genes and significantly reduced the mRNA expression of IGF-1 and SFRP2 genes. In particular, ZEA significantly decreased the expression of genes essential for estrogen synthesis including FSHR, CYP19A1 and HSD17β in granulosa cells. Furthermore, Q-PCR and Western-blot analysis also confirmed reduced expression of these genes in ZEA exposed granulosa cells. These effects were associated with a significant reduction of 17β-estradiol concentrations in the culture medium of granulosa cells. Collectively, these results demonstrated a concretely deleterious effect of ZEA exposure on the mRNA expression of steroidogenesis related genes and the production of steroid hormones in porcine ovarian granulosa cells in vitro.

Simultaneous determination of trichothecene micotoxins, ochratoxin A, and zearalenone in grain and products of its processing, feed premixes, and meat by gas chromatography

A rapid and simple method is developed for the simultaneous determination of ochratoxin A, toxin T-2, deoxynivalenol, and zearalenone by gas chromatography with electron capture detection; the method is applicable to grain, products of its processing, feed premixes, and meat in concentration ranges 0.05–5 mg/kg (0.01–2 mg/kg for ochratoxin A and 0.03–3 mg/kg for T-2). The micotoxins were extracted from samples with acetonitrile; the extracts were purified by the QuEChERS method; and derivatives of target analytes were obtained with trifluoroacetic anhydride. The overall duration of analysis was 1–1.5 h; the relative standard deviation of the results did not exceed 8%.

Highly sensitive electrochemical immunoassay for zearalenone in grain and grain-based food

We have developed a highly sensitive electrochemical immunoassay for the quantitation of zearalenone (ZEN), a mycotoxin produced by Fusarium species. In this enzyme linked immunosorbent assay, the enzymatic conversion of the substrate p-nitrophenylphosphate is detected by a microplate reader and the signal subsequently converted into an electrical signal. The concentrations of coating antigen (ZEN-ovalbumin), of monoclonal antibody, and of goat anti-mouse antibody labeled with alkaline phosphatase were optimized. In terms of electrochemical detection, the types and pH values of the buffers, the conditions for agitating, and scanning frequency were optimized. The effective detection range of this immunoassay is quite wide (0.004 to 9.5 ng mL−1), and the limit of detection is 2 pg mL−1. ZEN-free corn, wheat, and grain-based food samples were spiked with ZEN and analyzed by this method, and recoveries were found to range from 91.6 % to 113.0 %. Unlike previously described electrochemical methods, this method is both highly sensitive and has a wide working range. The method is fast and thus provides a platform for high-throughput analysis that meets the current need to monitor trace levels of analytes in grain and grain-based food.

A fluorescence polarization immunoassay for the detection of zearalenone in corn

The aim of this study was to develop a fluorescence polarization immunoassay (FPIA) that is based on the change in fluorescence polarization of fluorescently labeled small antigen when bound by a specific antibody, for use as a screening test for zearalenone (ZEN) in cereals and their products. Syntheses of fluorescein-labeled ZEN tracers containing three linkers of different lengths (2, 3 and 6-carbon bridge), ethylenediamine, 1,2-diaminopropane and hexamethylenediamine, were explored and their binding response with ZEN-specific antibody was evaluated. A fluoresceinthiocarbamyl hexamethylenediamine-labeled ZEN conjugate (ZEN-HMDF), which contain a 6-carbon bridge, was found to be the most sensitive FPIA for detection of ZEN. When tested on corn matrix the FPIA using the ZEN-HMDF tracer showed a detection range for ZEN of 150–1000 μg kg −1 with a detection limit of 137 μg kg −1, and required less than 2 min per sample to carry out, excluding extraction time. The average recovery from spiked corn samples was 106.4 ± 12.5%. Comparative analyses of 70 naturally occurring cereals and their products using FPIA, enzyme-linked immunosorbent assay (ELISA) and high-performance liquid chromatography (HPLC) showed that the coefficients of determination ( r 2) between FPIA and ELISA, and between FPIA and HPLC were 0.76 and 0.72, respectively. These results suggest that the ZEN-HMDF tracer is suitable for FPIA, which has potential as a screening tool for ZEN in grains without the need for a complicated clean-up procedure.

Simultaneous Determination of Aflatoxins, Ochratoxin A, and Zearalenone in Grains by New Immunoaffinity Column/Liquid Chromatography

The simultaneous determination of mycotoxins was performed in 3 steps: extraction, cleanup, and detection. For extraction, a mixture of acetonitrile-water (60 + 40, v/v) was proved appropriate. For cleanup, a new Afla-Ochra-Zea immunoaffinity column was used. After derivatization with trifluoroacetic acid, the mycotoxins aflatoxins, ochratoxin A (OTA), and zearalenone (ZEA) were determined simultaneously by liquid chromatography with fluorescence detection. The detection limits in different matrixes after cleanup with the new immunoaffinity column were very low: aflatoxins, 0.002-0.7 microg/kg; OTA, 0.07-0.25 microg/kg; ZEA, 1-3 microg/kg. The limits of determination were: aflatoxins, 0.25 microg/kg; OTA, 0.5 microg/kg; ZEA, 5 microg/kg. The recovery rates for aflatoxins, OTA, and ZEA for rye and rice were between 86 and 93% when a 0.5 g sample matter per immunoaffinity column was used.

Determination of zearalenone in grains by high-performance liquid chromatography–tandem mass spectrometry after solid-phase extraction with RP-18 columns or immunoaffinity columns

In this paper a robust, sensitive and selective LC–MS–MS method for the determination of zearalenone (ZON) in several cereals is described. Sample preparation was performed by extraction of the commodities with a mixture of acetonitrile and water followed by solid-phase extraction with RP-18 columns or immunoaffinity columns. The selective determination of ZON was achieved with an atmospheric pressure chemical ionization interface. Using the negative ion mode a detection limit of 0.5 μg/kg and a determination limit of 1 μg/kg grain was achieved, which is by a factor of 100 more sensitive than the positive ion mode. Zearalanone (ZAN), which does not occur in nature, was used as internal standard for quantification. A linear working range from 1.0 μg/kg to 1000 μg/kg could be achieved in grains with a standard deviation of 4% and recovery rates around 100%. All these results were independent from the grain matrices (maize, barley, oats, wheat) when ZAN was used as internal standard. Sample preparation with RP-18 and immunoaffinity materials gave comparable results. In addition, the method was successfully used for the investigation of naturally contaminated maize samples in the course of an interlaboratory comparison test.

Isolation and characterization of zearalenone sulfate produced by Fusarium spp

A water-soluble compound related to zearalenone was isolated from a culture of Fusarium graminearum 30 grown in rice. The structure of the novel metabolite was determined to be zearalenone-4-sulfate on the basis of fast-atom-bombardment mass spectrometry, proton nuclear magnetic resonance, UV spectroscopy, and by chemical and enzymatic reactions. Strains representing Fusarium equiseti, Fusarium sambucinum, and Fusarium roseum produced the sulfate conjugate as well. In the rat uterus enlargement bioassay, the metabolite or its hydrolysis product was found to retain the estrogenic activity characteristic of zearalenone. Natural occurrence of this novel metabolite might be significant because analytical methods devised for zearalenone in grain cannot detect the conjugate but the conjugate retains the biological properties of the mycotoxin when ingested by animals.

Liquid Chromatographic Determination of Aflatoxins, Ochratoxin A, and Zearalenone in Grains, Oilseeds, and Animal Feeds by Post-Column Derivatization and On-Line Sample Cleanup

A liquid chromatographic method using on-line sample cleanup, reverse flow analytical column loading, gradient elution, and postcolumn derivatization with iodine permits direct, rapid determination of aflatoxins B1, B2, G1, and G2, as well as ochratoxin A and zearalenone. Limits of quantitation are 5 ppb for the aflatoxins and ochratoxin A and 30 ppb for zearalenone. This procedure performs well as a multimycotoxin screen for cereal grains and oilseeds, with more limited success in complete animal feeds.

Zearalenone in cereal grains

AbstractZearalenone, a secondary metabolite with estrogenic properties, is produced by severalFusarium species that colonize cereal grains in the field and in storage. Recently, there have been reports of zearalenone contamination in corn, oats, barley, wheat, and grain sorghum. Corn and grain sorghum were examined for contamination due to obvious mold damage. Wheat, corn, and sorghum have been examined to determine the incidence of zearalenone in grains moving through commercial channels and stored on farms and at country elevators. Other grains such as oats and barley were analyzed because of associated estrogenic disturbances in farm animals. Stepsin procedures for the determination of zearalenone are extraction of a representative sample, partial purification of the extract by column chromatography, alkali treatment, or liquid‐liquid partitioning, and subsequent measurement of the isolated toxin. Zearalenone is measured in partially purified extracts by thin layer chromatography (TLC), gas liquid chromatography (GLC), and high pressure liquid chromatography (HPLC). Confirmation of zearalenone contamination can be accomplished by gas chromatography‐mass spectroscopy (GC‐MS). Multitoxin screening procedures have been deveoped for zearalenone in combination with one or more of the following mycotoxins: aflatoxin, T‐2 toxin, diacetoxyscirpenol, patulin, ochratoxin, penicillic acid, citrinin, penitrem A, and sterigmatocystin.