Effects of milling and baking technologies on levels of deoxynivalenol and its masked form deoxynivalenol-3-glucoside.

The co-occurrence of the major Fusarium mycotoxin deoxynivalenol (DON) and its conjugate deoxynivalenol-3-glucoside (DON-3-Glc) has been documented in infected wheat. This study reports on the fate of this masked DON within milling and baking technologies for the first time and compares its levels with those of the free parent toxin. The fractionation of DON-3-Glc and DON in milling fractions was similar, tested white flours contained only approximately 60% of their content in unprocessed wheat grains. No substantial changes of both target analytes occurred during the dough preparation process, i.e. kneading, fermentation, and proofing. However, when bakery improvers enzymes mixtures were employed as a dough ingredient, a distinct increase up to 145% of conjugated DON-3-Glc occurred in fermented dough. Some decrease of both DON-3-Glc and DON (10 and 13%, respectively, compared to fermented dough) took place during baking. Thermal degradation products of DON, namely norDON A, B, C, D, and DON-lactone were detected in roasted wheat samples and baked bread samples by means of UPLC-Orbitrap MS. Moreover, thermal degradation products derived from DON-3-Glc were detected and tentatively identified in heat-treated contaminated wheat and bread based on accurate mass measurement performed under the ultrahigh mass resolving power. These products, originating from DON-3-Glc through de-epoxidation and other structural changes in the seskviterpene cycle, were named norDON-3-Glc A, B, C, D, and DON-3-Glc-lactone analogically to DON degradation products. Most of these compounds were located in the crust of experimental breads.

The study of deoxynivalenol and its masked metabolites fate during the brewing process realised by UPLC-TOFMS method.

The co-occurrence of deoxynivalenol (DON) and deoxynivalenol-3-glucoside (DON-3-Glc) has been recently reported in malt and beer. In this study, the concentration changes were monitored within the brewing process of four beer brands: light, dark tap and two lagers, produced from ground malt mixtures differing in composition, and also mycotoxins content. A simple and rapid method employing DON-dedicated immunoaffinity columns (IAC) for the selective pre-concentration, followed by ultra-performance liquid chromatography coupled to a time-of-flight mass spectrometer (UPLC-TOFMS) system for the reliable quantification at (ultra)trace levels, was validated for all experimental matrices. The results document the key role of the malt contamination nature. While in the first monitoring period a significant increase of both DON and DON-3-Glc occurred (up to 250% and 450%, respectively), fairly different trends were observed when new malts were used for identical technological processing (in some beers a decrease of DON and only a small increase of DON-3-Glc occurred). Worth noticing, that the outcome of the brewing process was surprisingly reproducible for a particular malt mixture. In the final phase, a small monitoring study comparing Czech and Austrian alcohol-free and conventional beers was carried out.

Occurrence of deoxynivalenol and its major conjugate, deoxynivalenol-3-glucoside, in beer and some brewing intermediates.

Since deoxynivalenol (DON), the main representative of Fusarium toxic secondary metabolites, is a relatively common natural contaminant in barley, its traces can be detected in many commercial beers. Our previous study reporting for the first time the occurrence of relatively high levels of DON-3-glucoside (DON-3-Glc) in malt and beer prepared from relatively "clean" barley (semiscale experimental conditions) induced a follow-up investigation focused on this DON conjugate in commercial beers. The current survey involving in total 176 beers, representing different brands, and collected at various markets, has documented a ubiquitous occurrence of DON-3-Glc in this product. Its levels even exceeded that of free DON in some samples; the highest level found was 37 microg/L. In addition to glucosylated DON, its acetylated forms (ADONs) were also common contaminants in most of the beers. Generally, stronger beers (higher alcohol content) tended to contain higher levels of DON and its conjugates. No distinct relationship between the contamination of malt and beer was observed in samples collected from several breweries. Attention was also paid to comparison of data on malts obtained by LC-MS/MS and ELISA DON-dedicated kits. The latter provided apparently higher levels of DON, the most distinct difference being observed for malts processed at higher temperatures (caramel and roasted malts). The nature of this phenomenon has not yet been explained; in addition to cross-reacting species, other factors, such as the higher content of dark pigment, can also be the cause.

Deoxynivalenol and its conjugates in beer: a critical assessment of data obtained by enzyme-linked immunosorbent assay and liquid chromatography coupled to tandem mass spectrometry.

Enzyme-linked immunosorbent assays (ELISAs) are often employed for the control of deoxynivalenol (DON) in barley and other intermediates involved in beer production chain. Because of the occurrence of high levels of DON-3-glucoside (DON-3-Glc) in malt and beer that have been reported for the first time in our earlier study, research focused on the accuracy of DON determination by immunoassays in cereal-based matrices has been initiated. DON-3-Glc was strongly cross-reacting in all examined commercial ELISA test kits (Ridascreen) DON (R-Biopharm), Veratox 5/5 DON) (Neogen Corporation), Deoxynivalenol EIA (Euro-Diagnostica), and AgraQuant) DON Assay 0.25/5.0 Test Kit (Romer Labs). The highest overestimation in beer analysis, up to 1000%, when taking the DON content determined by liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS) as a reference method, was obtained by AgraQuant assay. Besides of DON-3-Glc and 3- and 15-acetyldeoxynivalenol (ADONs), also other, not known yet, matrix components contributed to false positive results. Similar phenomenon, although in a lesser extent due to lower content of these substances, was observed for using ELISA in the analysis of wheat. The relationship between a way of sample handling and DON overestimation was demonstrated; higher ELISA response was measured in an aqueous extract compared to that prepared by acetonitrile-water (84:16, v/v). Most of cross-reacting co-extracts were removed by MycoSep# 226 cartridge, what leads us to the hypothesis on the presence of currently unknown cross-reactive species.