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Eingeladener Vortrag
- nein (22)
The drug salinomycin (SAL) is a polyether antibiotic and used in veterinary Medicine as coccidiostat and growth promoter. Recently, SAL was suggested as a potential anticancer drug.
However, transformation products (TPs) resulting from metabolic and environmental degradation of SAL are incompletely known and structural information is missing. In this study, we therefore systematically investigated the formation and identification of SAL derived TPs using electrochemistry (EC) in an electrochemical reactor and rat and human liver microsome incubation (RLM and HLM) as TP generating methods. Liquid chromatography (LC) coupled to high-resolution mass spectrometry (HRMS) was applied to determine accurate masses in a suspected target analysis to identify TPs and to deduce occurring modification reactions of derived TPs. A total of 14 new, structurally different TPs were found (two EC-TPs, five RLM-TPs, and 11 HLM-TPs). The main modification reactions are decarbonylation for EC-TPs and oxidation (hydroxylation) for RLM/HLM-TPs. Of particular interest are potassium-based TPs identified after liver microsome incubation because these might have been overlooked or declared as oxidated sodium adducts in previous, non-HRMS-based studies due to the small mass difference between K and O + Na of 21 mDa. The MS fragmentation pattern of TPs was used to predict the position of identified modifications in the SAL molecule. The obtained knowledge regarding transformation reactions and novel TPs of SAL will contribute to elucidate SAL-metabolites with regards to structural prediction.
Polycyclic aromatic hydrocarbons (PAHs) are a large group of priority organic pollutants, which contaminate environmental compartments, food, and consumer products as well. Due to their frequent occurrence associated with elevated Levels of PAHs, plastic and rubber parts of consumer products and toys are particular sources of exposure. Although European maximum levels exist for eight carcinogenic PAHs in consumer products and toys according to REACH Regulation (EC) No. 1907/2006, certified reference materials (CRM) are still not available. To overcome this lack, the first CRM for the determination of PAHs in rubber toys (BAM-B001) was developed according to the requirements of ISO Guide 35. The whole process of CRM development including preparation, homogeneity and stability studies, and value assignment is presented.
The assignment of the certified mass fractions was based upon in-house study at BAM using stable isotope Dilution analysis (SIDA) gas chromatography mass spectrometry (GC–MS). The obtained values were confirmed by the results of two interlaboratory comparison (ILC) studies with more than 50 expert laboratories from Germany and China. The mass fractions of 14 PAHs including all REACH and GS mark regulated compounds were certified ranging between 0.2 and 15.4 mg/
kg accompanied by expanded uncertainties (coverage factor k = 2). In addition, informative values were determined for 4 PAHs, mainly due to higher uncertainties and/or lack of ILC data for confirmation. BAM-B001 is intended for analytical quality control particularly based on the AfPS GS 2019:01 PAK method and contributes to improve the chemical safety of consumer products including toys.
The ubiquitous Fusarium genus is responsible for the spoilage of vast amounts of cereals and fruits.
Besides the economic damage, the danger to human and animal health by the concomitant exposure to mycotoxins represents a serious problem. A large number of Fusarium species produce a variety of different mycotoxins of which the class of trichothecenes are of particular importance due to their toxicity.
Being identified as the common volatile precursor during the biosynthesis of trichothecenes, (−)-trichodiene (TD) is considered to be a biomarker for the respective mycotoxin content in food samples. We postulated that the development of a non-invasive, on-site GC-IMS method for the quantification of (−)-trichodiene supplemented with a stationary SIDA headspace GC-MS reference method would allow circumventing the laborious and expensive analyses of individual trichothecenes in large cereal samples.
In this work we present the syntheses of the required native calibration standard and an isotope labeled (TD-D3) internal standard.
Abstract Alternariol (AOH) and alternariol-9-monomethyl ether (AME) are two secondary metabolites of Alternaria fungi which can be found in various foodstuff 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 labeled 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-arylation as the key step. Finally, we demonstrate their suitability as internal standards in stable-isotope dilution assay (SIDA)-HPLC-MS/MS analysis commonly used for the quantification of the natural products in food and feed.
The aim of this study was to identify specific truffle marker substances within the truffle aroma. The aroma profile of different truffle species was analyzed using static headspace sampling with gas chromatography mass spectrometry analysis (SHS/GC-MS). Possible marker substances were identified, taking the additional literature into account. The selected marker substances were tested in an experiment with 19 truffle dogs. The hypothesis “If trained truffle dogs recognize the substances as supposed truffles in the context of an experiment, they can be regarded as specific” was made. As it would be nearly impossible to investigate every other possible emitter of the same compounds to determine their specificity, this hypothesis was a reasonable approximation. We were interested in the question of what it is the dogs actually search for on a chemical level and whether we can link their ability to find truffles to one or more specific marker substances. The results of the dog experiment are not as unambiguous as could have been expected based on the SHS/GC-MS measurements. Presumably, the truffle aroma is mainly characterized and perceived by dogs by dimethyl sulfide and dimethyl disulfide. However, as dogs are living beings and not analytical instruments, it seems unavoidable that one must live with some degree of uncertainty regarding these results.
Rapid, cost-efficient, and eco-friendly methods are desired today for routine analysis of the Fusarium mycotoxin zearalenone (ZEN) in edible vegetable oils. Liquid chromatography with fluorescence detection (HPLC-FLD) is commonly used to reliably control the specified ZEN maximum levels, which requires efficient sample clean-up to avoid matrix interferences. Therefore, a highly selective extraction and clean-up method based on reversible covalent hydrazine chemistry (RCHC) using hydrazine-functionalized silica was developed. This efficient solid-phase extraction (SPE) involves reversible hydrazone formation of ZEN with the hydrazine moiety covalently bound to a solid phase. Optimal conditions were achieved with 1 mL SPE cartridges filled with 400 mg of hydrazine-functionalized silica. The developed RCHC-SPE method was validated in an interlaboratory comparison study (ILC) with twelve participants analyzing six edible vegetable oils with a focus on maize oils. The derived method parameters (ZEN recovery 83%, repeatability 7.0%, and reproducibility 18%) meet the performance criteria of Commission Regulation (EC) No 401/2006. The developed RCHC-SPE-based HPLC-FLD method allows the reliable quantification of ZEN in the range of 47–494 μg/kg for different types of edible vegetable oils, also for matrix-reach native oils. Due to the high efficiency, the significantly reduced matrix load helps to extend the lifetime of analytical equipment. Furthermore, the re-useability of the RCHC-SPE cartridges contributes to an eco-friendly approach and reduced analysis costs. To our knowledge, this is the first report on ZEN quantification in edible vegetable oils based on manual RCHC-SPE cartridges. Due to its high performance, the developed RCHC-SPE method is a promising alternative to the current European standard method EN 16924:2017 (HPLC-FLD part).
Analysis of electrochemical and liver microsomal transformation products of lasalocid by LC/HRMS
(2022)
Rationale: Lasalocid (LAS), an ionophore, is used in cattle and poultry farming as feed additive for its antibiotic and growth-promoting properties. Literature on transformation products (TP) resulting from LAS degradation is limited. So far, only hydroxylation is found to occur as the metabolic reaction during the LAS degradation. To investigate potential TPs of LAS, we used electrochemistry (EC) and liver microsome (LM) assays to synthesize TPs, which were identified using liquid chromatography high-resolution mass spectrometry (LC/HRMS).
Methods: Electrochemically produced TPs were analyzed online by direct coupling of the electrochemical cell to the electrospray ionization (ESI) source of a Sciex Triple-TOF high resolution mass spectrometer. Then, EC-treated LAS solution was collected and analyzed offline using LC/HRMS to confirm stable TPs and improve their annotation with a chemical structure due to informative MS/MS spectra. In a complementary approach, TPs formed by rat and human microsomal incubation were investigated using LC/HRMS. The resulting data were used to investigate LAS modification reactions and elucidate the chemical structure of obtained TPs.
Results: The online measurements identified a broad variety of TPs, resulting from modification reactions like (de-)hydrogenation, hydration, methylation, oxidation as well as adduct formation with methanol. We consistently observed different ion complexations of LAS and LAS-TPs (Na+; 2Na+ K+; NaNH4 +; KNH4 +). Two stable methylated EC-TPs were found, structurally annotated, and assigned to a likely modification reaction. Using LM incubation, seven TPs were formed, mostly by oxidation/hydroxylation. After the identification of LM-TPs as Na+-complexes, we identified LM-TPs as K+-complexes.
Conclusion: We identified and characterized TPs of LAS using EC- and LM-based methods. Moreover, we found different ion complexes of LAS-based TPs. This knowledge, especially the different ion complexes, may help elucidate the metabolic and environmental degradation pathways of LAS.
Volatile organic compounds (VOCs) are of interest in many different fields. Among them are food and fragrance analysis, environmental and atmospheric research, industrial applications, security or medical and life science. In the past, the characterization of these compounds was mostly performed via sample collection and off-site analysis with gas chromatography coupled to mass spectrometry (GC-MS) as the gold standard. While powerful, this method also has several drawbacks such as being slow, expensive, and demanding on the user. For decades, intense research has been dedicated to find methods for fast VOC analysis on-site with time and spatial resolution. We present the working principles of the most important, utilized, and researched technologies for this purpose and highlight important publications from the last five years. In this overview, non-selective gas sensors, electronic noses, spectroscopic methods, miniaturized gas chromatography, ion mobility spectrometry and direct injection mass spectrometry are covered. The advantages and limitations of the different methods are compared. Finally, we give our outlook into the future progression of this field of research.
The present study investigates the in vitro phase I metabolism of cis-zearalenone (cis-ZEN) in rat liver microsomes and human liver microsomes. cis-ZEN is an often ignored isomer of the trans-configured Fusarium mycotoxin zearalenone (trans-ZEN). Upon the influence of (UV-) light, trans-ZEN isomerizes to cis-ZEN. Therefore, cis-ZEN is also present in food and feed. The aim of our study was to evaluate the in vitro phase I metabolism of cis-ZEN in comparison to that of trans-ZEN. As a result, an extensive metabolization of cis-ZEN is observed for rat and human liver microsomes as analyzed by HPLC-MS/MS and high-resolution MS. Kinetic investigations based on the substrate depletion approach showed no significant difference in rate constants and half-lives for cis- and trans-ZEN in rat microsomes. In contrast, cis-ZEN was depleted about 1.4-fold faster than trans-ZEN in human microsomes. The metabolite pattern of cis-ZEN revealed a total of 10 phase I metabolites. Its reduction products, α- and β-cis-zearalenol (α- and β-cis-ZEL), were found as metabolites in both species, with α-cis-ZEL being a major metabolite in rat liver microsomes. Both compounds were identified by co-chromatography with synthesized authentic standards. A further major metabolite in rat microsomes was monohydroxylated cis-ZEN. In human microsomes, monohydroxylated cis-ZEN is the single dominant peak of the metabolite profile. Our study discloses three metabolic pathways for cis-ZEN: reduction of the keto-group, monohydroxylation, and a combination of both. Because these routes have been reported for trans-ZEN, we conclude that the phase I metabolism of cis-ZEN is essentially similar to that of its trans isomer. As trans-ZEN is prone to metabolic activation, leading to the formation of more estrogenic metabolites, the novel metabolites of cis-ZEN reported in this study, in particular α-cis-ZEL, might also show higher estrogenicity.
Trichodiene is the first and only volatile intermediate in the biosynthesis of Fusarium mycotoxins and its detection in the gas-phase might therefore be of potential interest as a marker for food safety analysis. We herein present an improved diastereoselective synthesis of trichodiene which can be used as an analytical standard for a headspace gas chromatography / mass spectrometry method to be developed.
Cylindrospermopsin (CYN) is a cyanobacterial toxin associated with human and animal poisonings. Due to its toxicity in combination with its widespread occurrence, the development of reliable methods for selective, sensitive detection and accurate quantification is mandatory. Liquid chromatography tandem mass spectrometry (LC-MS/MS) analysis using stable isotope dilution analysis (SIDA) represents an ideal tool for this purpose. U-[15N5]-CYN was synthesized by culturing Aphanizomenon flos-aquae in Na15NO3-containing cyanobacteria growth medium followed by a cleanup using graphitized carbon black columns and mass spectrometric characterization. Subsequently, a SIDA-LC-MS/MS method for the quantification of CYN in freshwater and Brassica matrices was developed showing satisfactory performance data. The recovery ranged between 98 and 103 %; the limit of quantification was 15 ng/L in freshwater and 50 µg/kg dry weight in Brassica samples. The novel SIDA was applied for CYN determination in real freshwater samples as well as in kale and in vegetable mustard exposed to toxin-containing irrigation water. Two of the freshwater samples taken from German lakes were found to be CYN-contaminated above limit of quantification (17.9 and 60.8 ng/L). CYN is systemically available to the examined vegetable species after exposure of the rootstock leading to CYN mass fractions in kale and vegetable mustard leaves of 15.0 µg/kg fresh weight and 23.9 µg/kg fresh weight, respectively. CYN measurements in both matrices are exemplary for the versatile applicability of the developed method in environmental analysis.
Biotransformation of the mycotoxin zearalenone by fungi of the genera Rhizopus and Aspergillus
(2014)
Zearalenone (ZEN) is a nonsteroidal estrogenic mycotoxin biosynthesized by various Fusarium fungi. These fungal species frequently infest grains; therefore, ZEN represents a common contaminant in cereal products. The biotransformation of ZEN differs significantly from species to species, and several metabolites are known to be formed by animals, plants, and microorganisms. The aim of the present study was to investigate the microbial conversion of ZEN by species of the genera Rhizopus and Aspergillus representing relevant fungi for food processing (e.g. fermentation). To monitor the ZEN metabolism, ZEN was added to liquid cultures of the different fungal species. After a period of 3 days, the media were analyzed by HPLC-MS/MS for metabolite formation. Two Aspergillus oryzae strains and all seven Rhizopus species were able to convert ZEN into various metabolites, including ZEN-14-sulfate as well as ZEN-O-14- and ZEN-O-16-glucoside. Microbial transformation of ZEN into the significantly more estrogenic α-zearalenol (α-ZEL) was also observed. Additionally, a novel fungal metabolite, α-ZEL-sulfate, was detected. Semi-quantification of the main metabolites indicates that more than 50% of initial ZEN may be modified. The results show that fungal strains have the potential to convert ZEN into various metabolites leading to a masking of the toxin, for example in fermented food.
Fusarium mycotoxins in foodstuffs at levels in violation of allowed legal limits belong to the most frequently occurring type of food contamination. Out of the large variety of known mycotoxins, deoxynivalenol (DON), zearalenone (ZEN), and the T-toxins are the most prevalent and are responsible for extensive and recurring economic damage. For ensuring consumer safety, it is required to continually identify faster, simpler and more reliable analytical methodologies. In the present work, miniaturized centrifugation (spin) columns for the extraction of mycotoxins from grain samples were investigated, with a focus on matrix removal, applicability of different extraction solvents, and recovery of mycotoxin. The method was further extended to the extraction of naturally occurring DON and ZEN conjugates (masked mycotoxins). The spin column method was observed to routinely provide a total matrix reduction of >90% from typical wheat samples in a single centrifugation step. The use of H2O/ACN (20/80; v/v) as an extraction solvent provided non-masked DON, ZEN and T-toxin recoveries of greater than 80%. For the masked DON and ZEN conjugates (DON-3-glucoside and ZEN-14-sulfate), recoveries were lower. Overall, the evaluated miniaturized spin column procedure enabled rapid loss-free extraction of unconjugated mycotoxins from conventional grain matrix and a significantly reduced matrix load in the extracts.
Deacetyl tenuazonic acid
(2009)
Tenuazonic acid (TA) is a major Alternaria mycotoxin. In the present work a novel approach for the detection of TA in cereals by liquid chromatographyion-trap multistage mass spectrometry after derivatization with 2,4-dinitrophenylhydrazine is described. The product of the derivatization reaction and its major MS2 fragments were characterised by Fourier transform-ion cyclotron resonance tandem mass spectrometry. Without preconcentration, the established method features a limit of detection of 10 µg/kg using 2 g of sample in a rapid workup procedure. Accuracy, precision and linearity were evaluated in the working range of 505000 µg/kg. TA was detected in 13 and quantified in 3 out of 27 cereal samples obtained from a local supermarket, the average content being 49 µg/kg (highest incidence: 851 ± 41 µg/kg).
Certified reference materials (CRMs) are a versatile tool for quality assurance in the chemical laboratory. In the case of acrylamide analysis, the availability of appropriate materials was rather limited. This lack of acrylamide matrix CRMs has now been overcome by the development of two European reference materials (ERM) for the determination of acrylamide in food (crispbread, ERM-BD272, and rusk, ERM-BD274). This article describes the preparation of the materials, provides the results of the homogeneity and stability studies, and presents and discusses the outcome of the certification studies. Expert laboratories from different European countries took part in the certification studies using various analytical methods. The acrylamide mass fractions were certified to 980 µg kg-1 for crispbread and 74 µg kg-1 for rusk.
The stability of the Alternaria mycotoxins alternariol, alternariol monomethyl ether, and altenuene upon bread baking was investigated by model experiments using a spiked wholemeal wheat flour matrix. For alternariol and alternariol monomethyl ether, but not for altenuene, degradation products, formed through a sequence of hydrolysis and decarboxylation, could be identified in pilot studies. The simultaneous quantification of alternariol, alternariol monomethyl ether, altenuene, and the degradation products was achieved by a newly developed high performance liquid chromatography tandem mass spectrometry (HPLC-MS/MS) multimethod. The obtained quantitative data indicate that the Alternaria mycotoxins are barely degraded during wet baking, while significant degradation occurs upon dry baking, with the stability decreasing in the order alternariol monomethyl ether > alternariol > altenuene. The novel degradation products could be detected after the wet baking of flour spiked with alternariol and in a sample survey of 24 commercial cereal based baking products.
Sulfur dioxide (SO2) or sulfites are the most common preservatives used in winemaking. The level of total SO2 is subject to regulation. Currently, the regulatory determination of total SO2 (including sulfites) is done by the optimized Monier–Williams (OMW) method, which includes time-consuming distillation and titration steps. This paper describes the development and application of an alternative, rapid, straightforward, and reliable method for the determination of total sulfite in wine. In this method, a simple oxidation step using alkaline hydrogen peroxide (H2O2) solution is followed by ion chromatographic (IC) analysis of sulfate coupled with conductometric detection. Thirteen wines were analyzed in order to compare the in-sample oxidation method with the OMW-procedure. A t-test revealed satisfying compliance regarding sample preparation, i.e., alkaline H2O2 treatment and acidic distillation (OMW method). Comparable results were also obtained between IC analysis and acid/base titration. Our results indicate that the novel method (limit of quantification: 4 mg SO2 L-1) is well suited for the cost-efficient monitoring of regulatory limits.
Tenuazonic acid (TA) is a major water soluble Alternaria mycotoxin. In the present work, a method for the quantification of TA in beer by liquid chromatography-ion-trap multistage mass spectrometry after derivatization with 2,4-dinitrophenylhydrazine is described. The method is based on a rapid workup procedure and features a LOD of 2 µg/kg without preconcentration using 400 mg of sample. Validation was performed for a working range of 8-500 µg/kg.
A total of 43 beers of different brewing styles (pilsener, wheat beer, bock beer, dark beer and alcohol free beer) was analysed. TA was detected in 37 samples, 16 samples were above the LOQ. An average content of 11 µg/kg was found, the highest incidence being 175 ± 13 µg/kg. To our knowledge, this is the first report on the occurrence of TA in beer and beverages in general.
The degradation kinetics of the Alternaria mycotoxin tenuazonic acid (l-TA) in aqueous buffer were studied over a period of 4 months at different pH levels (3.5 and 7.0) and temperatures (4, 25 and 40°C). l-TA and its degradation products were quantified by newly developed high-performance liquid chromatography methods with UV or electrospray multistage mass spectrometry detection. At pH 3.5, significant degradation occurred at 25 and 40°C, the respective l-TA half-lives being 73.8±0.4 and 14.0±0.1 days. Two degradation processes, epimerization and hydrolysis, were evaluated kinetically. The hydrolytically formed iso-deacetyl TA (iso-DTA, epimeric mixture) was found to be the stable end product of l-TA degradation under the conditions of this study. This indicates that iso-DTA as well as the l-TA epimer u-TA are formed in aqueous beverage matrices.
Überwachung pflanzlicher Rohstoffe - Trichothecen-Analytik in Getreide und Getreideerzeugnissen
(2010)
Established maximum levels for the mycotoxin zearalenone (ZEN) in edible oil require monitoring by reliable analytical methods. Therefore, an automated SPE-HPLC online system based on dynamic covalent hydrazine chemistry has been developed. The SPE step comprises a reversible hydrazone formation by ZEN and a hydrazine moiety covalently attached to a solid phase. Seven hydrazine materials with different properties regarding the resin backbone, pore size, particle size, specific surface area, and loading have been evaluated. As a result, a hydrazine-functionalized silica gel was chosen. The final automated online method was validated and applied to the analysis of three maize germ oil samples including a provisionally certified reference material. Important performance criteria for the recovery (70–120 %) and precision (RSDr <25 %) as set by the Commission Regulation EC 401/2006 were fulfilled: The mean recovery was 78 % and RSDr did not exceed 8 %. The results of the SPE-HPLC online method were further compared to results obtained by liquid&–liquid extraction with stable isotope dilution analysis LC-MS/MS and found to be in good agreement. The developed SPE-HPLC online system with fluorescence detection allows a reliable, accurate, and sensitive quantification (limit of quantification, 30 µg/kg) of ZEN in edible oils while significantly reducing the workload. To our knowledge, this is the first report on an automated SPE-HPLC method based on a covalent SPE approach.
The role of mycotoxins in the microbial competition in an ecosystem or on the same host plant is still unclear. Therefore, a laboratory study was conducted to evaluate the influence of mycotoxins on growth and mycotoxin production of Fusarium and Alternaria fungi. Fusarium culmorum Fc13, Fusarium graminearum Fg23 and two Alternaria tenuissima isolates (At18 and At220) were incubated on wheat kernels supplemented with alternariol (AOH), tetramic acid derivates (TeA), deoxynivalenol (DON) and zearalenone (ZEA) in an in vitro test system. Fungal biomass was quantified by determining ergosterol content. Three Fusarium toxins (DON, nivalenol and ZEA) and three Alternaria toxins (AOH, alternariol methyl ether (AME) and altenuene) were analysed by HPLC-MS/MS. If Alternaria strains grew in wheat kernels spiked with Fusarium mycotoxins, their growth rates were moderately increased, their AOH and AME production was enhanced and they were simultaneously capable of degrading the Fusarium mycotoxins DON and ZEA. In contrast, both Fusarium strains behaved quite differently. The growth rate of Fc13 was not distinctly influenced, while Fg23 increased its growth in wheat kernels spiked with AOH. TeA depressed the ergosterol content in Fc13 as well as in Fg23. The DON production of Fc13 was slightly depressed, whereas the ZEA production was significantly increased. In contrast, Fg23 restricted its ZEA production. Both Fusarium strains were not capable of degrading the Alternaria mycotoxin AOH. Mycotoxins might play an important role in the interfungal competitive processes. They influence growth rates and mycotoxin production of the antagonistic combatants. The observed effects between phytopathogenic Alternaria and Fusarium strains and their mycotoxins aid the understanding of the complexity of microbial competitive behaviour in natural environments.
Die Prozesskontaminante Acrylamid steht im Verdacht krebserregend und erbgutverändernd zu sein, weshalb große Anstrengungen unternommen werden, um die Acrylamidgehalte in Lebensmitteln auf möglichst niedrige Werte zu reduzieren. Durch Anwendung des dynamischen Minimierungskonzepts konnte über die letzten Jahre in Deutschland eine deutliche Verringerung der mittleren täglichen Acrylamidaufnahme pro kg Körpergewicht erreicht werden, wobei die durch häusliche Zubereitung (Braten, Backen, Frittieren) entstehenden als auch die durch den Verzehr weiterer relevanter Lebensmittel wie Kakao und Kakaoerzeugnisse aufgenommenen Acrylamidmengen nicht oder nur marginal berücksichtigt werden.
Für die Ermittlung der Acrylamidbelastungen in Schokoladen unterschiedlicher Kakaogehalte sowie in reinem Kakaopulver wurden insgesamt 140 Proben aus deutschen Einzelhandelsmärkten untersucht. Die quantitative Bestimmung erfolgte unter Verwendung eines schnellen, kostengünstigen und validierten HPLC-MS/MS-basierten Analysenverfahrens in Kombination mit der Stabilisotopenverdünnungsanalyse (SIVA). In allen untersuchten Proben konnte Acrylamid nachgewiesen werden, wobei Gehalte im Bereich von 9,0-1747 µg kg-1 mit relativen Standardabweichungen zwischen 0,4 % und 10,8 % ermittelt wurden.
Zearalenone and its cis-isomer, cis-zearalenone, are nonsteroidal mycotoxins that elicit an estrogenic response upon binding to the estrogen receptor. This study compares the estrogenicity of eleven congeners including novel metabolites as 15-OH-zearalenone, zearalenone-14-sulfate, α-cis-zearalenol and β-cis-zearalenol using the E-Screen assay. Overall, a change in the configuration from trans to cis retains significant estrogenic activity. In contrast, alterations of the aromatic moiety including hydroxylation and sulfation showed a markedly decreased estrogenicity when compared to zearalenone.
A novel method for the determination of phomopsin A (1) in lupin flour, pea flour, and bean flour as well as whole lupin plants was established based on stable isotope dilution assay (SIDA) LC-MS/MS using 15N6-1 as an isotopically labeled internal standard. Artificially infected samples were used to develop an optimized extraction procedure and sample pretreatment. The limits of detection were 0.5–1 µg/kg for all matrices. The limits of quantitation were 2–4 µg/kg. The method was used to analyze flour samples generated from selected legume seeds and lupin plant samples that had been inoculated with Diaporthe toxica and two further fungal strains. Finally, growing lupin plants infected with D. toxica were investigated to simulate a naturally in-field mycotoxicosis. Toxin levels of up to 10.1 µg/kg of 1 were found in the pods and 7.2 µg/kg in the stems and leaves.
The mycotoxin deoxynivalenol (DON), formed by Fusarium species, is one of the most abundant mycotoxins contaminating food and feed worldwide. Upon ingestion, the majority of the toxin is excreted by humans and animal species as glucuronide conjugate. First in vitro data indicated that DON phase II metabolism is strongly species dependent. However, kinetic data on the in vitro metabolism as well as investigations on the specific enzymes responsible for DON glucuronidation in human are lacking. In the present study, the DON metabolism was investigated using human microsomal fractions and uridine-diphosphoglucuronyltransferases (UGTs) as well as liver microsomes from five animal species. Only two of the twelve tested human recombinant UGTs led to the formation of DON glucuronides with a different regiospecificity. UGT2B4 predominantly catalyzed the formation of DON-15-O-glucuronide (DON-15GlcA), while for UGT2B7 the DON-3-O-glucuronide (DON-3GlcA) metabolite prevailed. For human UGTs, liver, and intestinal microsomes, the glucuronidation activities were low. The estimated apparent intrinsic clearance (Clapp,int) for all human UGT as well as tissue homogenates was <1 mL/min mg protein. For the animal liver microsomes, moderate Clapp,int between 1.5 and 10 mL/min mg protein were calculated for carp, trout, and porcine liver. An elevated glucuronidation activity was detected for rat and bovine liver microsomes leading to Clapp,int between 20 and 80 mL/min mg protein. The obtained in vitro data points out that none of the animal models is suitable for estimating the human DON metabolism with respect to the metabolite pattern and formation rate.
Ergopeptides, like ergocornine and a-ergocryptine, exist in an S- and in an R-configuration. Kinetic experiments imply that certain configurations are preferred depending on the solvent. The experimental methods are explained in this article. Furthermore, computational methods are used to understand this configurational preference. Standard quantum chemical methods can predict the favored configurations by using minimum energy calculations on the potential energy landscape. However, the explicit role of the solvent is not revealed by this type of methods. In order to better understand its influence, classical mechanical molecular simulations are applied. It appears from our research that 'folding' the ergopeptide molecules into an intermediate state (between the S- and the R-configuration) is mechanically hindered for the preferred configurations.