Metabolism of nivalenol and nivalenol-3-glucoside in rats.

Plant-derived mycotoxin conjugates like deoxynivalenol-3-glucoside can be partly hydrolyzed to their aglycones in vivo, albeit to different extent depending on the mycotoxin conjugate and on the animal species. The aim of this work was to investigate the metabolization of the trichothecene mycotoxin nivalenol (NIV) and the fate of its modified form NIV-3-glucoside (NIV3G) in rats. To that end, 350 µg/kg body weight of NIV and the equimolar dose of NIV3G were administered to six rats by gavage in a 5 × 6 design and excreta were collected for 2 days after each treatment. For further analysis of NIV and NIV3G metabolites in rat urine and feces, seven novel NIV- and NIV3G metabolites including NIV sulfonates (NIVS) 1, 2 and 3, deepoxy-NIV (DNIV), DNIV sulfonate 2, NIV3G sulfonate (NIV3GS) 2 and NIV-3-glucuronide were produced, isolated and characterized. Subsequently, LC-MS/MS based methods for determination of NIV, NIV3G and their metabolites in excreta samples were developed, validated and applied. The biological recoveries of administered toxins in the form of their fecal and urinary metabolites were 57 ± 21% for NIV and 94 ± 36% for NIV3G. The majority of NIV and NIV3G metabolites was excreted into feces, with DNIV and NIVS 2 as major NIV metabolites and NIV3GS 2 and DNIV as major metabolites of NIV3G. Only 1.5% of the administered NIV3G was recovered in urine, with NIV3G itself as major urinary metabolite. The biological recovery of free NIV in urine was approximately 30 times lower after treatment with NIV3G than after administration of NIV, indicating that exposure of rats to NIV3G results in lower toxicity than exposure to NIV.

Optimised extraction methods for the determination of trichothecenes in rat faeces followed by liquid chromatography-tandem mass spectrometry.

The mycotoxin deoxynivalenol (DON) and some of its derivatives, such as 3­acetyl­deoxynivalenol (3AcDON), 15­acetyl­deoxynivalenol (15AcDON), deoxynivalenol­3­glucoside (DON3G) and de-epoxy deoxynivalenol (DOM-1), are commonly found in food and/or biological samples. However, literature does not present suitable methodologies for detecting and quantifying these mycotoxins at very low levels, which would be especially useful when they are present in biological samples. The main goal of the present paper was to evaluate different extraction techniques for the determination of these mycotoxins in rat faecal samples, in order to reduce the interferences present in the matrix and be able to quantify the mycotoxins at low concentration levels. Using diverse extraction methodologies such as QuEChERS (Quick, Easy, Cheap, Effective, Rugged and Safe) and pressurised liquid extraction (PLE), the clean-up strategy was optimised. QuEChERS extraction followed by a dispersive solid-phase extraction (dSPE) clean-up step with activated carbon was the method with the best extraction recovery results, ranging between 78% and 83% (except for DON3G). The matrix effect values were from -2% to -20% which supposed a reduction in comparison with the other tested strategies. These results enabled low quantification limits to be achieved, from 0.2 µg kg-1 to 3.4 µg kg-1. In view of the results, it was possible to quantify the natural presence of DON and DOM-1 in the tested faecal samples at low concentration levels.

Monitoring and evaluation of the interaction between deoxynivalenol and gut microbiota in Wistar rats by mass spectrometry-based metabolomics and next-generation sequencing.

Published evidence has demonstrated the several toxic characteristics of mycotoxins and their considerable risk to human and animal health. One of the most common uncertainties regards whether if very low concentrations of the mycotoxin deoxynivalenol (DON), easily consumed within the Mediterranean Diet, can cause metabolic alterations; some of them produced by the interaction between DON and gut microbiota. Accordingly, faecal samples were collected from Wistar rats that had consumed the mycotoxin DON at low levels (60 and 120 µg kg-1 body weight of DON per day), and were analysed by ultra-high performance liquid chromatography coupled with tandem mass spectrometry detection, in order to monitor the mycotoxin DON and its metabolite de-epoxy deoxynivalenol (DOM-1). The obtained results showed an evolution in DON excretion and the metabolite DOM-1 which has less toxic properties, over the course of the days of the study. To elucidate whether intestinal microbiota had a role in the observed detoxification process, the changes in microbial gut biodiversity were explored through 16s rRNA high throughput sequencing. No main changes were detected but significant increase in Coprococcus genus relative abundance was found. Further studies are needed to confirm if intestinal microbiota composition and function are affected by low mycotoxin concentrations.

Determination of mycotoxins in plant-based beverages using QuEChERS and liquid chromatography-tandem mass spectrometry.

A method was developed for the simultaneous determination of 11 mycotoxins in plant-based beverage matrices, using a QuEChERS extraction followed by ultra-high performance liquid chromatography coupled to tandem mass spectrometry detection (UHPLC-(ESI)MS/MS). This multi-mycotoxin method was applied to analyse plant-based beverages such as soy, oat and rice. QuEChERS extraction was applied obtaining suitable extraction recoveries between 80 and 91%, and good repeatability and reproducibility values. Method Quantification Limits were between 0.05µgL-1 (for aflatoxin G1 and aflatoxin B1) and 15µgL-1 (for deoxynivalenol and fumonisin B2). This is the first time that plant-based beverages have been analysed, and certain mycotoxins, such as deoxynivalenol, aflatoxin B1, aflatoxin B2, aflatoxin G1, aflatoxin G2, ochratoxin A, T-2 toxin and zearalenone, were found in the analysed samples, and some of them quantified between 0.1µgL-1 and 19µgL-1.