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Monitoring strategies for veterinary drugs in products of animal origin are shifting towards a more risk-based approach. Such strategies not only target a limited number of predefined .substances but also facilitate detection of unexpected substances. By combining the use of archive matrices such as feather meal with suspect-screening methods, early detection of new hazards in the food and feed industry can be achieved. Effective application of such strategies is hampered by complex data interpretation and therefore, targeted data analysis is commonly applied. In this study, the performance of a suspect-screening data processing workflow using a suspect list or the online spectral database mzCloudTM was explored to facilitate detection of veterinary drugs in archive matrices. Data evaluation parameters specifically investigated for application of a suspect list were mass tolerance and the addition or omission of retention times. Application of a mass tolerance of 1.5 ppm leads to an increase in the number of false positives, as does omission of retention times in the suspect list. Different acquisition modes yielding different qualities of MS2 data were studied and proved to be a critical factor, where data-dependent acquisition is preferred when matching to the mzCloudTM database. Using this approach, it is possible to search for compounds on a dedicated suspect list based on the exact mass and retention times and, at the same time, detect unexpected compounds without a priori information. A pilot study was conducted and fourteen different antibiotics were detected (and confirmed by MS/MS). Three of these antibiotics were not included in the suspect list. The optimised suspect-screening method proved to be fit for the purpose of finding veterinary drugs in feather meal, which are not in the scope of the current monitoring methods and therefore, it gives added value in the perspective of a risk-based monitoring.

Pyrrolizidine alkaloids (PAs) and their and the corresponding N-oxides (PAs-ox) are genotoxic plant metabolites which can be present as unwanted contaminants in food products of herbal origin like tea and food supplements. PAs and PAs-ox come in a wide variety of molecular structures including many structural isomers. For toxicity assessment it is important to determine the composition of a sample and to resolve all isomeric PAs and PAs-ox, which is currently not possible in one liquid or gas chromatographic (LC or GC) run. In this study an online two dimensional liquid chromatography quadrupole time-of-flight mass spectrometry (2D-LC QToF-MS) method was developed to resolve isomeric PAs and PAs-ox. After comprehensive column and mobile phase selection a polar endcapped C18 column was used at pH 3 in the first dimension, and a cross-linked C18 column at pH 10 in the second dimension. Injection solvents, column IDs, flow rates and temperatures were carefully optimized. The method with column selection valve switching described in this study was able to resolve and visualize 20 individual PAs/PAs-ox (6 sets of isomers) in one 2D-LC QToF-MS run. Moreover, it was shown that all isomeric PAs/PAs-ox could be unambiguously annotated. The method was shown to be applicable for the determination and quantification of isomeric PAs/PAs-ox in plant extracts and could be easily extended to include other PAs and PAs-ox.

Prenylated flavonoids and isoflavonoids are phytochemicals with remarkable antibacterial activity. In this study, 30 prenylated (iso)flavonoids were tested against Listeria monocytogenes and Escherichia coli (the latter in combination with an efflux pump inhibitor). Minimum inhibitory concentrations of the most active compounds ranged between 6.3–15.0 µg/mL. Quantitative structure-activity relationships (QSAR) analysis was performed and linear regression models were proposed with R2 between 0.77–0.80, average R2m between 0.70–0.75, Q2LOO between 0.66–0.69, and relatively low amount of descriptors. Shape descriptors (related to flexibility and globularity), together with hydrophilic/hydrophobic volume and surface area descriptors, were identified as important molecular characteristics related to activity. A 3D pharmacophore model explaining the effect of the prenyl position on the activity of compounds was developed for each bacterium. These models predicted active compounds with an accuracy of 71–88%. With regard to the mode of action, good antibacterial prenylated (iso)flavonoids with low relative hydrophobic surface area caused remarkable membrane permeabilization, whereas those with higher relative hydrophobic surface area did not. Based on the QSAR and membrane permeabilization studies, the mode of action of antibacterial prenylated (iso)flavonoids was putatively rationalized.

Het voorkomen, de risico’s en de mogelijkheden voor emissiereductie van humane geneesmiddelen in water krijgen de afgelopen jaren steeds meer aandacht. Naar diergeneesmiddelen gaat tot op heden minder aandacht uit dan naar humane geneesmiddelen. In 2017 is daarom onderzoek verricht naar de lotgevallen van diergeneesmiddelen in drijfmest uit de intensieve veehouderij die toegediend wordt op het land. Het onderzoek richtte zich op de kalver- en varkenshouderij en met name op het risico van uitspoeling. Voor het onderzoek werden vijf op zandgrond gelegen bedrijven in Gelderland geselecteerd uit iedere sector. Voorafgaand aan de mesttoediening werd een monster van de mest zelf genomen en op diverse tijdstippen voor en na de mestinjectie werden de bodem, het grondwater en het oppervlaktewater en sediment van kavelsloten naast de bemeste percelen bemonsterd. In al deze monsters werden de concentraties van een groot aantal antibiotica, antiparasitaire middelen, coccidiostatica en natuurlijke hormonen bepaald middels chemische analyse.

Item does not contain fulltext Six prenylated (iso)flavonoids were purified from a licorice root extract and subjected to competition experiments with six commercially available (iso)flavonoids. The agonistic and antagonistic activities of these compounds towards both hERalpha (human estrogen receptor alpha) and hERbeta were determined. Differences in the modes of action (agonist or antagonist) were observed for the various compounds tested. In general, each compound had the same mode of action towards both ERs. In silico modeling was performed in order to study the differences in estrogenicity observed between the compounds. It is suggested that prenyl chains fit into a hydrophobic pocket present in the hER, resulting in an increased agonistic activity. In addition, it was shown that an increase in length ( approximately 1.7 A) of pyran prenylated isoflavonoids resulted in an antagonistic mode of action. This might be caused by collision of the pyran ring with helix 11 in the ligand binding cavity of the hER. 01 december 2015

The isoflavonoid composition of an ethanolic extract of fungus-treated soybean sprouts was strongly altered by a combined acid/heat treatment. UHPLC-MS analysis showed that 6a-hydroxy-pterocarpans were completely converted to their respective, more stable, 6a,11a-pterocarpenes, whereas other isoflavonoids, from the isoflavone and coumestan subclasses, were affected to a much lesser extent (loss of ~15%). Subsequently, mixtures enriched in prenylated 6a-hydroxy-pterocarpans (pools of glyceollin I/II/III and glyceollin IV/VI) or prenylated 6a,11a-pterocarpenes (pools of dehydroglyceollin I/II/III and dehydroglyceollin IV/VI) were purified, and tested for activity on both human estrogen receptors (ERa and ERß). In particular, the response toward ERa changed, from agonistic for glyceollins to antagonistic for dehydroglyceollins. Toward ERß a decrease in agonistic activity was observed. These results indicate that the introduction of a double bond with the concomitant loss of a hydroxyl group in 6a-hydroxy-pterocarpans extensively modulates their estrogenic activity.

Seven prenylated 6a-hydroxy-pterocapans and five prenylated 6a,11a-pterocarpenes with different kinds of prenylation were purified from an ethanolic extract of fungus-treated soybean sprouts. The activity of these compounds toward both human estrogen receptors (hERα and hERβ) was determined in a yeast bioassay and the activity toward hERα was additionally tested in an U2-OS based hERα CALUX bioassay. In the yeast bioassay, compounds with chain prenylation showed in general an agonistic mode of action toward hERα, whereas furan and pyran prenylation led to an antagonistic mode of action. Five of these antagonistic compounds had an agonistic mode of action in the U2-OS based hERα CALUX bioassay, implying that these compounds can act as SERMs. The yeast bioassay also identified 8 ER subtype-selective compounds, with either an antagonistic mode of action or no response toward hERα and an agonistic mode of action toward hERβ. The ER subtype-selective compounds were characterized by 6a-hydroxy-pterocarpan or 6a,11a-pterocarpene backbone structure. It is suggested that either the extra D-ring or the increase in length to 12-13.5 Å of these compounds is responsible for an agonistic mode of action toward hERβ and, thereby, inducing ER subtype-selective behavior.

In vitro liver metabolism of 11 prenylated flavonoids and isoflavonoids was investigated by determining their phase I glucuronyl and sulfate metabolites using pork liver preparations. One hundred metabolites were annotated using RP-UHPLC-ESI-MS(n). A mass spectrometry-based data interpretation guideline was proposed for the tentative annotation of the position of hydroxyl groups, considering its relevance for estrogenic activity. To relate structure to metabolism, compounds were classified on the basis of three criteria: backbone structure (isoflavene, isoflavan, or flavanone), number of prenyl groups (0, 1, or 2), and prenyl configuration (chain or pyran). Glucuronidation was most extensive for isoflavenes and for unprenylated compounds (yield of 90-100%). Pyran and chain prenylation gave more complex hydroxylation patterns with 4 or more than 6 hydroxyl isomers, respectively, as compared to unprenylated compounds (only 1 hydroxyl isomer). Moreover, the number of hydroxyl isomers also increased with the number of prenyl groups.