TY - CONF A1 - Herter, Sven-Oliver A1 - Haase, Hajo A1 - Koch, Matthias T1 - Semi-synthesis of isotopic labeled ergot alkaloids: New reference standards N2 - We developed a two-step semi-synthesis for the preparation of isotopically labeled EAs, starting from native EAs. This universal strategy enabled the successful synthesis of all isotopically labeled priority EAs. The structure of the isotopically labeled EAs was confirmed by HPLC-HR-MS/MS using native, unlabeled EAs as a reference standard. The next step will be the implementation of the isotopically labeled standards in the European standard procedure EN 17425 to improve the quantification of EAs in foodstuffs. T2 - 45th Mycotoxin Workshop CY - Vienna, Austria DA - 02.06.2024 KW - Reference Material KW - HPLC-MS/MS KW - Mycotoxins KW - Isotop standards KW - Organic Synthesis KW - Ergot alkaloids PY - 2024 AN - OPUS4-60239 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - CONF A1 - Riedel, Soraya A1 - Hülagü, Deniz A1 - Bennet, Francesca A1 - Carl, Peter A1 - Flemig, Sabine A1 - Schmid, Thomas A1 - Schenk, Jörg A1 - Hodoroaba, Vasile-Dan A1 - Schneider, Rudolf T1 - Application of 3,3’,5,5’-Tetramethylbenzidine (TMB) in Amperometric Immunoassays for Mycotoxin Detection N2 - Electrochemical methods make great promise to meet the demand for user-friendly on-site devices for monitoring important parameters. Food industry often runs own lab procedures, e.g., for mycotoxin analysis, but it is a major goal to simplify analysis, linking analytical methods with miniaturized technologies. Enzyme-linked immunosorbent assays, with photometric detection of the horseradish peroxidase (HRP) substrate 3,3’,5,5’-tetramethylbenzidine (TMB), form a good basis for sensitive detection. To provide a straight-forward approach for the miniaturization of the detection step, we have studied the pitfalls of the electrochemical TMB detection. By cyclic voltammetry, it could be shown that TMB electrochemistry is strongly dependent on the pH and the electrode material. It was found that screen-printed gold electrodes and a very low pH value (pH 1) are well-suited to perform the electrochemical detection of TMB, due to the reversible character of the redox reaction under these conditions. Under these conditions, a good signal stability over several measuring cycles is achieved, providing the basis for analyzing multiple samples. In contrast to this, for carbon screen-printed electrodes, it was found that the signal response has changed after the electrochemical reaction with TMB at pH 1. At moderately acidic conditions (pH 4), neither with carbon nor with gold electrodes a reproducible electrochemical detection of TMB could be achieved. Based on these findings, we created a smartphone-based, electrochemical, immunomagnetic assay for the detection of ochratoxin A (OTA) and ergometrine in food samples. A competitive assay is performed on magnetic beads using HRP and TMB/H2O2 to generate the signal. Enzymatically oxidized TMB is quantified after addition of H2SO4 by amperometry with screen-printed gold electrodes in a custom-made wall-jet flow cell. The results are in good correlation with the established photometric detection method, providing a solid basis for sensing of further analytes in HRP-based assays using the newly developed miniaturized smartphone-based, electrochemical, immunomagnetic assay. T2 - 73rd Annual Meeting of the ISE CY - Online meeting DA - 12.09.2022 KW - Cylic Voltammetry KW - Immunoassay KW - Mycotoxins KW - Amperometry KW - Electrochemistry PY - 2022 N1 - Geburtsname von Riedel, Soraya: Höfs, S. - Birth name of Riedel, Soraya: Höfs, S. AN - OPUS4-55793 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - CONF A1 - Riedel, Soraya A1 - Bayram, Rabia A1 - Döring, Sarah A1 - Schneider, Rudolf A1 - Konthur, Zoltán T1 - Tracing toxic fumonisins - Fumonisin sensing by Aspergillus niger Fumonisin Amine Oxidase (AnFAO) and amperometric hydrogen peroxide detection N2 - Fumonisins are a class of toxic secondary metabolites produced by various Fusarium species. The two most important producers of fumonisins are F. verticillioides and F. proliferatum but also Aspergillus niger is known to produce fumonisins. Most frequently they occur on maize, but also other grains can be contaminated with this group of mycotoxins. Exposure to fumonisins by dietary intake can have serious health effects on farm animals such as equine leukoencephalomalacia and porcine pulmonary oedema and is associated with neural tube defects and esophageal cancer in humans. Thus, the European Commission sets legal limits for fumonisins in foodstuffs. The detection of fumonisins is frequently performed in laboratories by chromatographic methods, which are costly and require trained personnel. Simplifying the analysis is therefore a major goal using portable detection systems. Electrochemical enzymatic biosensors offer great promise to meet this demand. Here we report for the first time an enzymatic fumonisin sensing approach with amperometric detection. For this purpose, an Aspergillus niger fumonisin amine oxidase (AnFAO) catalyzing the oxidative deamination of fumonisins, producing hydrogen peroxide, was recombinantly produced in E. coli. For the first time, the specific enzyme activity of AnFAO was determined using a horseradish peroxidase-based fluorescence assay. It was found that the specific activity of AnFAO using 20 μM Fumonisin B1 as substrate is higher than for 20 μM Fumonisin B2 with 0.122 U mg-1 and 0.058 U mg-1, respectively. It was possible to show a dependence of enzyme activity with enzyme – and substrate-concentration. For fumonisin B1 detection, the enzyme was coupled covalently to magnetic particles and the enzymatically produced H2O2 was detected amperometrically in a flow injection system using Prussian blue carbon electrodes. The developed method allows to quantify fumonisin B1 concentrations down to 1.5 µM and demonstrates that the recombinantly produced AnFAO was able to deaminate different concentrations of fumonisin even in immobilized form. Thus, this enzyme is well suited to develop an enzyme based electrochemical biosensor for fumonisin contaminated food and feed. T2 - Posterschau Adlershofer Forschungsforum CY - Berlin, Germany DA - 11.11.2022 KW - Biosensor KW - Amperometry KW - Mycotoxins KW - Electrochemical sensor KW - Recombinant protein expression PY - 2022 N1 - Geburtsname von Riedel, Soraya: Höfs, S. - Birth name of Riedel, Soraya: Höfs, S. AN - OPUS4-56248 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - CONF A1 - Riedel, Soraya T1 - How to trace a cereal killer? N2 - How to trace the "cereal" killer? Fungal secondary metabolites, aka mycotoxins, pose a severe health risk for humans and animals. Since their occurrence in food, such as cereal products, is inevitable, the detection is of considerable importance. I want to demonstrate how the trace analysis of mycotoxins in foodstuff can be performed outside the laboratory. For this purpose, sensitive, antibody-based methods are combined with electrochemical smartphone-based detection. T2 - Bionnale 2021 CY - Online meeting DA - 12.05.2021 KW - Mycotoxins KW - Electrochemistry KW - Ochratoxin A KW - Food safety PY - 2021 N1 - Geburtsname von Riedel, Soraya: Höfs, S. - Birth name of Riedel, Soraya: Höfs, S. AN - OPUS4-52658 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - CONF A1 - Hoffmann, A. A1 - Lischeid, G. A1 - Koch, Matthias A1 - Lentzsch, P. A1 - Müller, M.E.H. T1 - Priority effects influence the production of mycotoxins of Fusarium and Alternaria N2 - Mycotoxigenic fungal pathogens Fusarium and Alternaria are a leading cause of loss in cereal production. On wheat-ears, they are confronted by bacterial antagonists such as pseudomonads. Studies on these groups’ interactions often neglect the infection process’s temporal aspects and the associated priority effects. In the present study, the focus was on how the first colonizer affects the subsequent ones. In a climate chamber experiment, wheat-ears were successively inoculated with two different strains (Alternaria tenuissima At625, Fusarium graminearum Fg23, or Pseudomonas simiae Ps9). Over three weeks, microbial abundances and mycotoxin concentrations were analyzed and visualized via Self Organizing Maps with Sammon Mapping (SOM-SM). All three strains revealed different characteristics and strategies to deal with co-inoculation: Fg23, as the first colonizer, suppressed the establishment of At625 and Ps9. Nevertheless, primary inoculation of At625 reduced all of the Fusarium toxins and stopped Ps9 from establishing. Ps9 showed priority effects in delaying and blocking the production of the fungal mycotoxins. The SOM-SM analysis visualized the competitive strengths: Fg23 ranked first, At625 second, Ps9 third. Our findings of species-specific priority effects in a natural environment and the role of the mycotoxins involved are relevant for developing biocontrol strategies. T2 - 42. Mycotoxin Workshop CY - Online meeting DA - 30.05.2021 KW - Microbe interactions KW - Antagonists KW - Mycotoxins PY - 2021 AN - OPUS4-52815 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - CONF A1 - Kuner, Maximilian A1 - Koch, Matthias A1 - Kühn, S. A1 - Haase, H. T1 - Measuring the sum – A novel screening method for ergot alkaloids in food N2 - Ergot alkaloids play a significant role in human history for more than 1000 years and are part of the most common contaminants of food and feed worldwide. Their high toxicity against humans and livestock even at low concentrations causes a high demand for quick and reliable analytics even though no European limits for ergot alkaloids have been determined yet. Currently the most common methods for the quantitation of the six major ergot alkaloids and their corresponding stereoisomers are HPLC based, using either fluorescence or mass spectrometric detection. Whereas these conventional detection methods measure each compound individually, a novel approach is to transfer all ergot alkaloids to one basic structure, which could be measured as a sum parameter. Since all ergots contain a lysergic acid amide moiety and a differing peptide component, cleaving the molecule into a simple lysergic acid derivative, which could be quantified via either HPLC-FLD or MS/MS, is intended. To clean up the cleavage reaction mixture molecularly imprinted polymers (MIPs) are a simple and effective way to separate the desired structure. Due to the selectivity of MIPs, an improved matrix separation is expected, which results in fewer interferences in the FLD and the possibility to measure samples with more complex matrices. When fully developed, the novel method could overcome some major drawbacks of the conventional detection methods. Higher throughput and the need for less well-trained personnel are just two advantages, that should lead to a quick and cheap quantitation of ergot alkaloids. First results of this project will be presented, funded by the German ZIM program (Zentrales Innovationsprogramm Mittelstand) of the Federal Ministry for Economic Affairs and Energy. T2 - WMFmeetsIUPAC 2019 CY - Belfast, UK DA - 14.10.2019 KW - Ergot KW - Mycotoxins KW - Molecularly imprinted polymers PY - 2019 AN - OPUS4-49436 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - CONF A1 - Kuner, Maximilian A1 - Koch, Matthias A1 - Haase, H. A1 - Kühn, S. T1 - Measuring the sum – A novel screening method for ergot alkaloids in food N2 - Ergot alkaloids play a significant role in human history for more than 1000 years and are part of the most common contaminants of food and feed worldwide. Their high toxicity against humans and livestock even at low concentrations causes a high demand for quick and reliable analytics even though no European limits for ergot alkaloids have been determined yet. Currently the most common methods for the quantitation of the six major ergot alkaloids and their corresponding stereoisomers are HPLC based, using either fluorescence or mass spectrometric detection. Whereas these conventional detection methods measure each compound individually, a novel approach is to transfer all ergot alkaloids to one basic structure, which could be measured as a sum parameter. Since all ergots contain a lysergic acid amide moiety and a differing peptide component, cleaving the molecule into a simple lysergic acid derivative, which could be quantified via either HPLC-FLD or MS/MS, is intended. To clean up the cleavage reaction mixture molecularly imprinted polymers (MIPs) are a simple and effective way to separate the desired structure. Due to the selectivity of MIPs, an improved matrix separation is expected, which results in fewer interferences in the FLD and the possibility to measure samples with more complex matrices. When fully developed, the novel method could overcome some major drawbacks of the conventional detection methods. Higher throughput and the need for less well-trained personnel are just two advantages, that should lead to a quick and cheap quantitation of ergot alkaloids. First results of this project will be presented, funded by the German ZIM program (Zentrales Innovationsprogramm Mittelstand) of the Federal Ministry for Economic Affairs and Energy. T2 - WMFmeetsIUPAC 2019 CY - Belfast, UK DA - 14.10.2019 KW - Ergot KW - Mycotoxins KW - Molecularly imprinted polymers PY - 2019 AN - OPUS4-49437 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - CONF A1 - Maul, Ronald A1 - Borzekowski, Antje A1 - Drewitz, Tatjana A1 - Keller, Julia A1 - Pfeifer, Dietmar A1 - Kunte, Hans-Jörg A1 - Koch, Matthias A1 - Rohn, S. T1 - Biosynthesis of zearalenone conjugates by fungi N2 - Zearalenone (ZEN) and its sulfate and glucoside conjugates have been detected in (a broad variety of) food and feed commodities1. Both conjugated derivatives are formed as part of fungal or plant secondary metabolism and thus, belong to the group of modified mycotoxins2. After consumption of contaminated foodstuff, the conjugates can be hydrolyzed by human intestinal microbiota leading to liberation of ZEN that implies an underestimation of the true ZEN exposure. In order to include ZEN conjugates in routine analysis, as well as for toxicological investigation reliable standards are needed. The objective of the present study was to develop a simple and economic method for biosynthesis of ZEN conjugates. Preceding experiments on the biotransformation of ZEN by Rhizopus and Aspergillus species showed a mixed metabolite formation3. Therefore, these known ZEN conjugating fungal strains were screened for their potential to selectively synthesize the ZEN derivatives ZEN-14-sulfate (Z14S), ZEN-14-glucoside (Z14G) and ZEN-16-glucoside (Z16G). The screening was conducted by adding ZEN to liquid fungal cultures. Cultivation conditions and ZEN incubation time were varied. All media samples were analyzed for metabolite formation by HPLC-MS/MS. Z14S was exclusively formed by A. oryzae. Under optimized conditions a specific biosynthesis of Z14G by R. oryzae and Z16G by R. oligosporus was achieved. After liquid-liquid-extraction and preparative chromatographic cleanup 1H-NMR purities of ≥ 73% for Z14S, ≥ 82% for Z14G and ≥ 50% for Z16G were obtained. In addition, a consecutive biosynthesis was developed by first using Fusarium graminearum for ZEN biosynthesis on rice based liquid medium. After inactivation of Fusarium the subsequent conjugation reaction was conducted utilizing Aspergillus and Rhizopus species under the various optimized conditions. In this study an easy and cost-efficient biosynthesis for Z14S, Z14G and Z16G was developed. The developed biosynthesis could be also used for other metabolites like ZEL conjugates. Our results of the in vitro screening indicate also the formation of a ZEL-glucoside and α ZEL-sulfate as major metabolites by R. oryzae. In sum, under optimized cultivation conditions fungi can be easily utilized for a targeted and stereospecific synthesis of ZEN conjugates. T2 - 10th World Mycotoxin Forum Conference CY - Amsterdam, The Netherlands DA - 12.03.2018 KW - Mycotoxins KW - Food safety KW - Analytical standards PY - 2018 AN - OPUS4-44547 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - CONF A1 - Sebald, M. A1 - Gebauer, J. A1 - Koch, Matthias T1 - Under Investigation: Novel Approach towards the Synthesis of Deuterium-labelled Alternariol- and Alternariol monomethylether-Standards for the HPLC-MS/MS-Analysis in Food & Feed N2 - Alternariol (AOH) and Alternariol monomethylether (AME) are two secondary metabolites of Alternaria fungi which can be found in various foodstuffs like tomatoes, nuts and grains. Due to their toxicity and potential mutagenic activity the need for the development of high-throughput methods for the supervision of AOH- and AME-levels is of increasing interest. As the availability of both native and labelled AOH and AME analytical standards is very limited we herein wish to present a novel concise approach towards their synthesis employing a ruthenium-catalyzed ortho-arylation4 as the key step. T2 - 13th International Symposium on the Synthesis and Applications of Isotopes and Isotopically Labelled Compounds CY - Prague, Czech Republic DA - 03.06.2018 KW - Mycotoxins KW - Mass Spectrometry PY - 2018 AN - OPUS4-45268 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - CONF A1 - Keller, Julia A1 - Borzekowski, Antje A1 - Haase, H. A1 - Rueß, L. A1 - Menzel, R. A1 - Koch, Matthias T1 - Using the model organism Caenorhabditis elegans for the toxicity testing of citrinin, zearalenone and zearalenone-14-sulfate N2 - To keep up with emerging mycotoxins and their transformation products fast and reliable toxicity tests are needed. Toxicity testing of mycotoxins is carried out usually by performing in vitro assays or is evaluated by using laboratory animals like mice, rats or chicken in in vivo studies. Settled between classical in vitro approaches and in vivo studies with higher animals are tests with the nematode Caenorhabditis elegans. Since Sydney Brenner described 1974 the cultivation and handling of C. elegans, this worm is widely used as model organism in developmental biology and neurology. Due to many benefits like easy and cheap cultivation, a completely sequenced genome and short generation time, it also plays an important role in toxicological research. Finally, the high number of conserved genes between human and C. elegans make the worm an ideal candidate for toxicological investigations. In this study we used C. elegans to assess the toxic effects of the relevant food mycotoxin citrinin (CIT), the mycoestrogen zearalenone (ZEN) and the modified mycotoxin ZEN-14-sulfate (ZEN-14-S) on different lifetable parameters including reproduction, thermal and oxidative stress tolerance and lifespan. All tested mycotoxins significantly decreased the amount of offspring. In case of ZEN and CIT also significant negative effects on stress tolerance and lifespan were observed compared to the control group. Moreover, metabolization of mycotoxins in the worms was investigated by using LC MS/MS. Extraction of the worms treated 5 days with mycotoxin-containing and UVC-killed bacteria showed metabolization of ZEN to α-ZEL and β-ZEL (ZEL = zearalenol, ratio about 3:2). ZEN 14-S was reduced to ZEL 14-S and CIT was metabolized to mono hydroxylated CIT. T2 - 40th Mycotoxin Workshop CY - Munich, Germany DA - 11.06.2018 KW - Mycotoxins KW - Caenorhabditis elegans KW - Toxicity testing PY - 2018 AN - OPUS4-45170 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER -