Masked mycotoxins in food and feed: challenges and analytical approaches

Mycotoxins are among the most abundant contaminants in food and feed worldwide. Therefore, in the EU maximum levels are established, e.g. for the frequently occurring Fusarium toxins deoxynivalenol (DON) and zearalenone (ZEA). Additional to DON and ZEA, modified mycotoxins are present in naturally contaminated grain products contributing significantly to the exposure of humans and animals with mycotoxins. Up to now data on the spatial distribution of many (masked) mycotoxins in the kernels of wheat are missing. The aim of the present study was to investigate the amounts of DON and ZEA as well as their most abundant derivatives DON-3-glucoside (DON-3G), 3- and 15-acetyl-DON, ZEA-14- and 16-glucoside and ZEA-14-sulphate (ZEA-14S) in mill fractions of naturally contaminated wheat batches using HPLC-MS/MS. The investigated distribution pattern in ten milling fractions is comparable among the three investigated different wheat batches. Interestingly, DON and DON-3G were found to be present to similar amounts in all fractions. In bran, the levels were only slightly higher than in the endosperm. By contrast, for ZEA and ZEA-14S a significantly higher amount of toxin is located in the fibre-rich fractions. The relative mass proportion of DON-3G comprises for only between 2.9 and 11.2% of the free DON, while the relative mass proportion of ZEA-14S is estimated to even exceed the amount of free ZEA in certain fractions. Acetylated DON derivatives and ZEA-glucosides were only detected in low amounts. The experimental results show that a significant reduction of the ZEA and ZEA-14S level in wheat flour is feasible by applying milling technology strategies. However, the almost evenly distribution of DON and DON-3G in all fractions does not allow for the technological removal of relevant toxin amounts. Furthermore, the relative share of masked forms was higher for ZEA derivatives than for the DON conjugates in the investigated wheat lots.

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Trace element speciation in food: State of the art of analytical techniques and methods

Pure and Applied Chemistry ◽

10.1351/pac-con-11-08-14 ◽

2012 ◽

Vol 84 (2) ◽

pp. 169-179 ◽

Cited By ~ 14

Author(s):

Rola Bou Khouzam ◽

Joanna Szpunar ◽

Michel Holeman ◽

Ryszard Lobinski

Keyword(s):

High Resolution Mass Spectrometry ◽

State Of The Art ◽

Analytical Techniques ◽

Metal Species ◽

Recent Developments ◽

Food And Feed ◽

Element Speciation ◽

Analytical Approaches

Some elements in food are notoriously toxic, whereas others are considered essential for human health. Information on the exact chemical form in which an element is present in food is of paramount importance to determine the safety and nutritional quality of food. This critical review discusses the state of the art of analytical approaches to speciation of trace elements in food products. The topics addressed include (i) responding to regulations concerning some toxic elements (As, Hg, Sn); (ii) quality control of food and feed supplements; and (iii) characterization, in terms of element speciation, of nutritional plants (natural and genetically modified) and food supplements produced by biotechnology. The maturity of analytical techniques allowing the determination of individual well-defined metal species is highlighted. On the other hand, the recent developments of multidimensional hyphenated techniques and the democratization of electrospray high-resolution mass spectrometry (Orbitrap) start permitting fine characterization of element speciation in natural products.

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Occurrence and within field variability of Fusarium mycotoxins and their masked forms in maize crops in Belgium

World Mycotoxin Journal ◽

10.3920/wmj2013.1608 ◽

2014 ◽

Vol 7 (1) ◽

pp. 91-102 ◽

Cited By ~ 16

Author(s):

M. De Boevre ◽

S. Landschoot ◽

K. Audenaert ◽

P. Maene ◽

Diana Di Mavungu ◽

...

Keyword(s):

Fusarium Species ◽

Natural Infection ◽

Ear Rot ◽

Fusarium Mycotoxins ◽

Masked Mycotoxins ◽

Maize Varieties ◽

Food And Feed ◽

Feed Safety ◽

Field Variability ◽

Maize Ear Rot

Maize ear rot caused by several Fusarium species is an important fungal disease. Apart from yield losses, ear rot fungi can produce mycotoxins and masked forms in infected grains. Masked mycotoxins have received increased attention in view of their bioavailability and potential toxicity in animals and humans, but their presence and relevance in the field still remain undisclosed. To get a better insight, the present study assessed the presence of various Fusarium parent and masked mycotoxins, i.e. deoxynivalenol, 3-acetyldeoxynivalenol, 15-acetyldeoxynivalenol, deoxynivalenol-3-glucoside, zearalenone, α-zearalenol, β-zearalenol, zearalenone-14-glucoside, zearalenone-14-sulfate, α-zearalenol-14-glucoside, β-zearalenol-14-glucoside, T-2 and HT-2 toxin, in various commercial maize varieties grown under natural infection conditions in Flanders, Belgium. The results showed that the maize varieties were co-contaminated with both parent and masked mycotoxins. Moreover, a positive correlation between these forms was established. A higher contamination with a particular mycotoxin appeared to be coupled with an elevated load of another (masked) mycotoxin. The results highlight the importance to screen for multiple mycotoxins, both parent and masked, to guarantee food and feed safety. Furthermore, analysis was carried out to elucidate the distribution of the various mycotoxins in the field. The maize variety did not significantly influence mycotoxin accumulation, except for deoxynivalenol. Subdivisions in the field with higher mycotoxin levels for deoxynivalenol and its derivatives, zearalenone and its derivatives, and the sum of T-2 and HT-2 toxin were observed.

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Simultaneous determination of masked forms of deoxynivalenol and zearalenone after oral dosing in rats by LC-MS/MS

World Mycotoxin Journal ◽

10.3920/wmj2012.1411 ◽

2012 ◽

Vol 5 (3) ◽

pp. 303-318 ◽

Cited By ~ 41

Author(s):

A. Veršilovskis ◽

J. Geys ◽

B. Huybrechts ◽

E. Goossens ◽

S. De Saeger ◽

...

Keyword(s):

Hydrolytic Enzymes ◽

Uv Spectra ◽

In Vivo Metabolism ◽

Masked Mycotoxins ◽

Food And Feed ◽

Full Scan ◽

Indirect Analysis ◽

First Time

In vivo metabolism of masked or conjugated mycotoxins is poorly documented as standards are not commercially available and indirect analysis using hydrolytic enzymes is difficult to validate and cumbersome. We synthesised zearalenone-14-glucoside (ZEA-14G) chemically. Deoxynivalenol-3-glucuronide (DON-3GlcA) and glucuronides of 3- and 15-acetyl-deoxynivalenol (3- and 15-ADON-GlcAs), de-epoxydeoxynivalenol, zearalenone (ZEA), α- and β-zearalenol (α- and β-ZOL) were synthesised using rat microsomes. For the first time three ADON-GlcAs were synthesised: two 3-ADON-GlcAs and one 15-ADON-GlcA. After purification, the masked mycotoxin and the metabolites were characterised by NMR (DON-3GlcA, ZEA-14G) or by full scan MS, MS/MS fragmentation, UV-spectra, β-glucosidase and β-glucuronidase treatment. In a first experiment, rats were fed orally DON-3-glucoside (DON-3G) and ZEA-14G, together with 13C-DON and 13C-ZEA and were sacrificed after 55 minutes. A total of 21 masked metabolites, metabolites and parent mycotoxins were quantified in rat organs. Whereas DON-3G was hardly hydrolysed in the stomach, ZEA was clearly formed from ZEA-14G. In a second experiment, 3- and 15-ADON were given orally to rats. The acetylated forms of DON were hydrolysed in the stomach, in contrast to DON-3G. Rats can directly glucuronidate ADONs without deacetylation. Neither DOM, α- or β-ZOL nor their glucuronides could be quantified. Glucuronidated 3-ADON accumulated in the small intestines, together with DON-3GlcA in rats fed orally with 3- and 15-ADON. These differences in masked mycotoxins metabolism can be important in risk analysis of masked mycotoxins in food and feed.

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Cyclodextrins Can Entrap Zearalenone-14-Glucoside: Interaction of the Masked Mycotoxin with Cyclodextrins and Cyclodextrin Bead Polymer

Biomolecules ◽

10.3390/biom9080354 ◽

2019 ◽

Vol 9 (8) ◽

pp. 354 ◽

Cited By ~ 5

Author(s):

Zelma Faisal ◽

Eszter Fliszár-Nyúl ◽

Luca Dellafiora ◽

Gianni Galaverna ◽

Chiara Dall’Asta ◽

...

Keyword(s):

Aqueous Solution ◽

Molecular Modeling ◽

Fluorescence Spectroscopy ◽

Masked Mycotoxins ◽

Chemically Modified ◽

Promising Tool ◽

Food And Feed ◽

Neutral Conditions ◽

Feed Samples ◽

Analytical Detection

Zearalenone (ZEN) is a Fusarium-derived xenoestrogenic mycotoxin. In plants, zearalenone-14-O-β-d-glucoside (Z14G) is the major conjugated metabolite of ZEN, and is a masked mycotoxin. Masked mycotoxins are plant-modified derivatives, which are not routinely screened in food and feed samples. Cyclodextrins (CDs) are cyclic oligosaccharides built up from D-glucopyranose units. CDs can form stable host–guest type complexes with lipophilic molecules (e.g., with some mycotoxins). In this study, the interaction of Z14G with native and chemically modified β- and γ-CDs was examined employing fluorescence spectroscopy and molecular modeling. Furthermore, the removal of Z14G from aqueous solution by insoluble β-CD bead polymer (BBP) was also tested. Our results demonstrate that Z14G forms the most stable complexes with γ-CDs under acidic and neutral conditions (K ≈ 103 L/mol). Among the CDs tested, randomly methylated γ-CD induced the highest increase in the fluorescence of Z14G (7.1-fold) and formed the most stable complexes with the mycotoxin (K = 2 × 103 L/mol). Furthermore, BBP considerably reduced the Z14G content of aqueous solution. Based on these observations, CD technology seems a promising tool to improve the fluorescence analytical detection of Z14G and to discover new mycotoxin binders which can also remove masked mycotoxins (e.g., Z14G).

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Determination of T-2 and HT-2 toxins in food and feed: an update

World Mycotoxin Journal ◽

10.3920/wmj2013.1605 ◽

2014 ◽

Vol 7 (2) ◽

pp. 131-142 ◽

Cited By ~ 22

Author(s):

R. Krska ◽

A. Malachova ◽

F. Berthiller ◽

H.P. van Egmond

Keyword(s):

Mass Spectrometry ◽

Matrix Effects ◽

Certified Reference Materials ◽

Animal Health ◽

Rapid Screening ◽

Chromatography Method ◽

Food And Feed ◽

Immunochemical Methods ◽

Analytical Approaches

Based on the recent scientific opinion of the European Food Safety Authority (EFSA) Panel on Contaminants in the Food Chain on the risks to human and animal health related to the presence of T-2 and HT-2 toxins in food and feed that was published by EFSA in the EFSA Journal, this article provides an update on the determination of these Fusarium mycotoxins. After a brief introduction into the chemistry of these toxins, both chromatographic and immuno-analytical methods are discussed for the determination of these type A trichothecenes. During the last decade, liquid chromatography with (tandem) mass spectrometry has become the most frequently used method for the determination of T-2 and HT-2 toxins, often within a multi-analyte approach. However, complex matrices and the resulting signal suppression effects, as observed particularly in electrospray-mass spectrometry methods owing to matrix effects, may require careful optimisation of clean-up, usage of matrix matched standards, or e.g. the use of internal standards. For specific purposes where extremely low limits of quantification are needed, e.g. for the analysis of duplicate diets, a dedicated gas chromatography method with multistage mass spectrometry has become available. Other novel analytical approaches to determine T-2 and HT-2 toxins in food and feed include biosensor-based methods in surface plasmon resonance and electrochemical formats, as well as DNA microchip assays. For rapid screening, several immunochemical methods (mostly ELISAs) have become available and some are sold as commercial test kits. Whereas these methods work fast, cross-reactivities with other trichothecenes can have an undesired effect on their accuracy. While proficiency tests including T-2 and HT-2 toxins have been carried out, none of the chromatographic or immunochemical methods have been formally validated in interlaboratory validation studies. There are no certified reference materials available for T-2 and HT-2 toxins.

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Rapid enzymatic hydrolysis of masked deoxynivalenol and zearalenone prior to liquid chromatography mass spectrometry or immunoassay analysis

World Mycotoxin Journal ◽

10.3920/wmj2013.1662 ◽

2014 ◽

Vol 7 (2) ◽

pp. 107-113 ◽

Cited By ~ 4

Author(s):

M.W.F. Nielen ◽

C.A.G.M. Weijers ◽

J. Peters ◽

L. Weignerová ◽

H. Zuilhof ◽

...

Keyword(s):

Mass Spectrometry ◽

Liquid Chromatography ◽

Daily Intake ◽

Cross Reactivity ◽

Enzymatic Pretreatment ◽

Masked Mycotoxins ◽

Food And Feed ◽

Set Up ◽

Rapid Hydrolysis ◽

Hydrolysis Of

Recently it has been shown that conjugates (‘masked mycotoxins’) may contribute to the total daily intake of hazardous mycotoxins. Therefore, there is an urgent need for rapid analysis methods that assess the level of both free and masked mycotoxins in food and feed. However, the analysis of masked mycotoxins by either immunoassays or instrumental methods, such as liquid chromatography tandem mass spectrometry (LC-MS/MS), is severely hindered by the lack of standards and the unpredictable cross-reactivity profiles of the available antibodies. In this work, 26 enzymes were explored for rapid hydrolysis of masked mycotoxins using deoxynivalenol-3-glucoside (DON-3G) as model compound. Following initial screening, the most promising enzyme, a fungal 1,3-β-glucanase (laminarinase), was investigated in detail and found to be fit-for-purpose, providing complete conversions in minutes rather than hours according to LC-MS/MS analyses. As a proof of concept, the enzymatic pretreatment was applied to an extract of beer containing DON-3G. In addition, the feasibility of a fully automated enzymatic pretreatment of masked mycotoxin standards in an autosampler was demonstrated in an imaging surface plasmon resonance immunoassay set-up. Such an automated pretreatment was found to be equally applicable to other mycotoxin conjugates, as shown by the conversion of zearalenone-14-β-D-glucopyranoside and zearalenone-14-sulphate, in the latter case using a sulphatase enzyme. It is envisaged that laminarinase could be useful for other masked mycotoxins as well.

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