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Vekiru E.,Christian Doppler Laboratory | Hametner C.,Vienna University of Technology | Mitterbauer R.,Christian Doppler Laboratory | Mitterbauer R.,Sandoz GmbH | And 5 more authors.
Applied and Environmental Microbiology | Year: 2010

Zearalenone (ZON) is a potent estrogenic mycotoxin produced by several Fusarium species most frequently on maize and therefore can be found in food and animal feed. Since animal production performance is negatively affected by the presence of ZON, its detoxification in contaminated plant material or by-products of bioethanol production would be advantageous. Microbial biotransformation into nontoxic metabolites is one promising approach. In this study the main transformation product of ZON formed by the yeast Trichosporon mycotoxinivorans was identified and characterized by liquid chromatography- tandem mass spectrometry (LCMS/MS) and LC-diode array detector (DAD) analysis. The metabolite, named ZOM-1, was purified, and its molecular formula, C 18H24O7, was established by time of flight MS (TOF MS) from the ions observed at m/z 351.1445 [M-H]- and at m/z 375.1416 [M-HNa]+. Employing nuclear magnetic resonance (NMR) spectroscopy, the novel ZON metabolite was finally identified as (5S)-5-({2,4-dihydroxy-6- [(1E)-5-hydroxypent-1-en1-yl]benzoyl}oxy)hexanoic acid. The structure of ZOM-1 is characterized by an opening of the macrocyclic ring of ZON at the ketone group at C6′. ZOM-1 did not show estrogenic activity in a sensitive yeast bioassay, even at a concentration 1,000-fold higher than that of ZON and did not interact with the human estrogen receptor in an in vitro competitive binding assay. Copyright © 2010, American Society for Microbiology. All Rights Reserved.

Rampler E.,University of Natural Resources and Life Sciences, Vienna | Rampler E.,Biomin Research Center | Dalik T.,University of Natural Resources and Life Sciences, Vienna | Stingeder G.,University of Natural Resources and Life Sciences, Vienna | And 2 more authors.
Journal of Analytical Atomic Spectrometry | Year: 2012

For the first time, quantitative analysis of the proteinogenic sulfur containing amino acids methionine and cysteine was performed by LC-ICP-MS. A dedicated sample preparation procedure was implemented consisting of two steps, i.e. (1) protection of the redox sensitive amino acids by controlled oxidation of methionine, cystine and cysteine to methionine sulfone and cysteic acid, respectively, and (2) subsequent protein hydrolysis. Anion exchange chromatography enabled the separation of all relevant sulfur species within 10 min. Sulfur was detected on m/z 32S 16O using O 2 as the reaction gas. Absolute limits of detection in the pmol range were achieved for methionine sulfone and cysteic acid. The method offered the possibility of protein quantification. Absolute amounts of 2 μg of hydrolyzed protein (on column) were investigated by LC-ICP-MS. Both oxidized forms of amino acids showed excellent recoveries from lysozyme and myoglobin standards, enabling accurate quantification. A repeatability of <10% (n = 6 independently prepared samples) was found without the application of isotope dilution strategies. The limits of detection of <1 μM protein were comparable to the limits of detection achieved by spectroscopy based protein quantification assays. Moreover, the validity of the approach was shown by implementing HPLC in combination with fluorescence detection as a reference method for the quantification of proteinogenic amino acids in yeast. Both methods were in good agreement and met the theoretical value in a yeast reference material certified for the methionine content. © 2012 The Royal Society of Chemistry.

Grenier B.,Purdue University | Grenier B.,Biomin Research Center | Applegate T.J.,Purdue University
Toxins | Year: 2013

Mycotoxins are secondary metabolites of fungi that can cause serious health problems in animals, and may result in severe economic losses. Deleterious effects of these feed contaminants in animals are well documented, ranging from growth impairment, decreased resistance to pathogens, hepato- and nephrotoxicity to death. By contrast, data with regard to their impact on intestinal functions are more limited. However, intestinal cells are the first cells to be exposed to mycotoxins, and often at higher concentrations than other tissues. In addition, mycotoxins specifically target high protein turnover- and activated-cells, which are predominant in gut epithelium. Therefore, intestinal investigations have gained significant interest over the last decade, and some publications have demonstrated that mycotoxins are able to compromise several key functions of the gastrointestinal tract, including decreased surface area available for nutrient absorption, modulation of nutrient transporters, or loss of barrier function. In addition some mycotoxins facilitate persistence of intestinal pathogens and potentiate intestinal inflammation. By contrast, the effect of these fungal metabolites on the intestinal microbiota is largely unknown. This review focuses on mycotoxins which are of concern in terms of occurrence and toxicity, namely: aflatoxins, ochratoxin A and Fusarium toxins. Results from nearly 100 published experiments (in vitro, ex vivo and in vivo) were analyzed with a special attention to the doses used. © 2013 by the authors; licensee MDPI, Basel, Switzerland.

Nagl V.,University of Natural Resources and Life Sciences, Vienna | Woechtl B.,University of Veterinary Medicine Vienna | Schwartz-Zimmermann H.E.,University of Natural Resources and Life Sciences, Vienna | Hennig-Pauka I.,University of Veterinary Medicine Vienna | And 3 more authors.
Toxicology Letters | Year: 2014

Plants can metabolize the Fusarium mycotoxin deoxynivalenol (DON) by forming the masked mycotoxin deoxynivalenol-3-β-d-glucoside (D3G). D3G might be cleaved during digestion, thus increasing the total DON burden of an individual. Due to a lack of in vivo data, D3G has not been included in the various regulatory limits established for DON so far. The aim of our study was to contribute to the risk assessment of D3G by determination of its metabolism in pigs. Four piglets received water, D3G (116. μg/kg b.w.) and the equimolar amount of DON (75. μg/kg b.w.) by gavage on day 1, 5 and 9 of the experiment, respectively. Additionally, 15.5. μg D3G/kg b.w. were administered intravenously on day 13. Urine and feces were collected for 24. h and analyzed for DON, D3G, deoxynivalenol-3-glucuronide (DON-3-GlcA), deoxynivalenol-15-GlcA (DON-15-GlcA) and deepoxy-deoxynivalenol (DOM-1) by UHPLC-MS/MS. After oral application of DON and D3G, in total 84.8. ±. 9.7% and 40.3. ±. 8.5% of the given dose were detected in urine, respectively. The majority of orally administered D3G was excreted in form of DON, DON-15-GlcA, DOM-1 and DON-3-GlcA, while urinary D3G accounted for only 2.6. ±. 1.4%. In feces, just trace amounts of metabolites were found. Intravenously administered D3G was almost exclusively excreted in unmetabolized form via urine. Data indicate that D3G is nearly completely hydrolyzed in the intestinal tract of pigs, while the toxin seems to be rather stable after systemic absorption. Compared to DON, the oral bioavailability of D3G and its metabolites seems to be reduced by a factor of up to 2, approximately. © 2014 The Authors.

Mountzouris K.C.,Agricultural University of Athens | Tsitrsikos P.,Agricultural University of Athens | Palamidi I.,Agricultural University of Athens | Arvaniti A.,Agricultural University of Athens | And 3 more authors.
Poultry Science | Year: 2010

The aim of this work was to investigate the effect of inclusion levels of a 5-bacterial species probiotic in broiler nutrition. Five hundred twenty-five 1-d-old male Cobb broilers were allocated in 5 experimental treatments for 6 wk. The experimental treatments received a corn-soybean coccidiostat-free basal diet and depending on the addition were labeled as follows: no addition (C), 108 cfu probiotic/kg of diet (P1), 109 cfu probiotic/kg of diet (P2), 1010 cfu probiotic/kg of diet (P3), and 2.5 mg of avilamycin/kg of diet (A). Each treatment had 3 replicates of 35 broilers each. Treatment effects on broiler growth performance and biomarkers such as ileal and total tract nutrient digestibility, plasma Ig concentration, and cecal micro-flora composition were determined. Differences among treatments were considered significant when P ≤ 0.05. Overall BW gain was significantly higher in treatment P1 (2,293 g) compared with P2 (2,163 g), C (2,165 g), and P3 (2,167 g), with A (2,230 g) being intermediate and not different from P1. Overall feed conversion ratio values were similar and significantly better for P1 (1.80) and A (1.80) compared with P2 (1.87), C (1.89), and P3 (1.92). Ileal apparent digestibility coefficients (ADC) of CP and ether extract were higher in A. Generally, treatments A and P1 showed an improved total tract ADC for DM, organic matter, ash, ether extract, and AMEn values. The total tract ADC of CP was higher in P1, C, and P2. There were no differences between treatments regarding plasma Ig in 14-and 42-d-old broilers. Treatments P2 and P3 were effective at beneficially modulating cecal microflora composition. In particular, the lower cecal coliform concentration (log cfu/g of wet digesta) was seen in P2 (6.12) and P3 (4.90) in 14-and 42-d-old broilers, respectively, whereas at 42 d, P3 and P2 had the highest Bifido-bacterium (8.31; 8.08) and Lactobacillus concentrations (8.20; 7.86), respectively. It is concluded that probiotic inclusion level had a significant effect on broiler growth responses, nutrient ADC, AMEn, and cecal microflora composition. © 2010 Poultry Science Association Inc.

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