TY - JOUR A1 - Schmidt, Stephan A1 - Hoffmann, Holger A1 - Garbe, L.-A. A1 - Schneider, Rudolf T1 - Liquid chromatography-tandem mass spectrometry detection of diclofenac and related compounds in water samples N2 - A frequently studied environmental contaminant is the active substance diclofenac, which is removed insufficiently in sewage treatment plants. Since its inclusion in the watch list of the EU Water Framework Directive, the concentrations in surface waters will be determined throughout Europe. For this, still, more precise analytical methods are needed. As a reference, HPLC-MS is frequently employed. One of the major metabolites is 4’-hydroxydiclofenac (4’-OH-DCF). Also, diclofenac lactam is important for assessing degradation and transformation. Aceclofenac (ACF), the glycolic acid ester of diclofenac is used as a drug, too, and could potentially be cleaved to yield diclofenac again. In various sewage treatment plant influent samples, diclofenac, 4’-OH-DCF, DCF lactam and ACF could be determined with detection limits of 3 µg/L, 0.2 µg/L, 0.17 µg/L and 10 ng/L, respectively. KW - Diclofenac KW - Festphasenextraktion KW - HPLC-MS KW - Probenvorbereitung KW - Kläranlage KW - Abwasser KW - Zyklisierung KW - Diclofenac KW - Solid-phase extraction KW - Sewage treatment plant KW - Cyclization KW - Sample treatment KW - HPLC–MS PY - 2018 U6 - https://doi.org/10.1016/j.chroma.2018.01.037 SN - 0021-9673 SN - 1873-3778 VL - 1538 SP - 112 EP - 116 PB - Elsevier B.V. CY - Amsterdam, NL AN - OPUS4-44183 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - JOUR A1 - Baldofski, Stefanie A1 - Canitz, C. A1 - Garbe, L.-A. A1 - Schneider, Rudolf T1 - Studies on the development of antibodies for the highly hydrophobic plasticizers DINCH and DEHT N2 - Diisononylcyclohexane-1,2-dicarboxylate (DINCH) and di-2-ethylhexyl terephthalate (DEHT), two of the most important substitutes for phthalate plasticizers, are used for a wide range of applications. Consequently, an increasing occurrence in urine and environmental samples is reported. Reliable and fast analytical methods for the quantification of these plasticizers are needed. So far, mainly GC-MS or LC-MS methods are used. We aimed to develop the first antibodies and immunoassays allowing for high-throughput analysis of samples. We designed two DINCH hapten structures and one DEHT hapten structure and employed hapten-protein conjugates for the immunization of rabbits. Sensitive competitive enzyme-linked immunosorbent assays (ELISAs) against each hapten using the produced polyclonal antibodies were established. Yet, binding of DINCH to the respective antibodies was not observed in neither direct nor indirect assay formats, even when using protein conjugates with the heterologous haptens and different carrier proteins in the indirect format. The use of surfactants and solvents in the sample buffer did not result in recognition of the plasticizers. Also, no binding of DEHT in ELISA employing the respective antibodies was detected. We speculate that the production of antibodies against these highly hydrophobic molecules is not possible via our route, however a different hapten design could overcome this obstacle. KW - Weichmacher KW - Phthalate KW - Antikörper KW - Selektivität KW - Ersatzstoffe KW - Polyclonal antibodies KW - Phthalate substitutes KW - Antibody production for hydrophobic molecules KW - DEHT KW - DINCH KW - Hapten design PY - 2018 U6 - https://doi.org/10.1016/j.ab.2017.12.006 SN - 0003-2697 VL - 543 SP - 90 EP - 96 PB - Elsevier Inc. CY - Amsterdam, NL AN - OPUS4-44015 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - JOUR A1 - Oberleitner, Lidia A1 - Dahmen-Levison, U. A1 - Garbe, L.-A. A1 - Schneider, Rudolf T1 - Application of fluorescence polarization immunoassay for determination of carbamazepine in wastewater N2 - Carbamazepine is an antiepileptic drug that can be used as a marker for the cleaning efficiency of wastewater treatment plants. Here, we present the optimization of a fast and easy on-site measurement system based on fluorescence polarization immunoassay and the successful application to wastewater. A new monoclonal highly specific anti-carbamazepine antibody was applied. The automated assay procedure takes 16 min and does not require sample preparation besides filtration. The recovery rates for carbamazepine in wastewater samples were between 60.8 and 104% with good intra- and inter-assay coefficients of variations (less than 15 and 10%, respectively). This automated assay enables for the onsite measurement of carbamazepine in wastewater treatment plants. KW - On-site measurement KW - Wastewater treatment plant KW - Fluorescence polarization immunoassay KW - Assay optimization KW - Mix-and-read assay PY - 2017 U6 - https://doi.org/10.1016/j.jenvman.2017.01.063 SN - 0301-4797 SN - 1095-8630 VL - 193 SP - 92 EP - 97 PB - Elsevier Ltd. AN - OPUS4-39152 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - JOUR A1 - Schneider, Rudolf A1 - Schmidt, S. A1 - Hanelt, S. A1 - Canitz, C. A1 - Hoffmann, H. A1 - Garbe, L.-A. T1 - Synthetic Strategies for the Modification of Diclofenac N2 - For many heterogeneous sensor applications as well as the synthesis of hapten antigens to produce antibodies, protein conjugates of the target substance are essential. A requirement is that the target substance already offers or is modified to contain a functionality that allows for coupling to a protein, that is, an amino acid residue. Ideally, to avoid shielding of the compound by the carrier protein, a sufficient distance to the protein surface should be provided. With its carboxyl function diclofenac (DCF) allows for direct binding to lysine residues after in situ synthesis of the NHS ester. One problem is that diclofenac as free acid tends to autocondensation, which results in low yields. Here we describe the 'insertion' of a C6 spacer via synthesis of the amide with 6-aminohexanoic acid. To carry out the reaction in solution, first the methyl ester of the amino acid had to be produced. Due to otherwise low yields and large cleaning efforts, solid-phase synthesis on Fmoc Ahx Wang resin is recommended. The crude product is mainly contaminated by cleavage products from the resin which were removed by chromatography. The structure of the highly pure hapten was completely determined by nuclear magnetic resonance (NMR) spectroscopy. KW - Diclophenac KW - Hapten synthesis KW - Solid-phase synthesis PY - 2017 U6 - https://doi.org/10.1055/s-0036-1588858 SN - 0936-5214 VL - 28 IS - 15 SP - 1984 EP - 1989 PB - Georg Thieme Verlag AN - OPUS4-43316 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - JOUR A1 - Baldofski, Stefanie A1 - Hoffmann, Holger A1 - Lehmann, Andreas A1 - Breitfeld, Stefan A1 - Garbe, L.-A. A1 - Schneider, Rudolf T1 - Enzyme-linked immunosorbent assay (ELISA) for the anthropogenic marker isolithocholic acid in water N2 - Bile acids are promising chemical markers to assess the pollution of water samples with fecal material. This study describes the optimization and validation of a direct competitive enzyme-linked immunosorbent assay for the bile acid isolithocholic acid (ILA). The quantification range of the optimized Assay was between 0.09 and 15 mg/L. The assay was applied to environmental water samples. Most studies until now were focused on bile acid fractions in the particulate phase of water samples. In order to avoid tedious sample preparation, we undertook to evaluate the dynamics and significance of ILA levels in the aqueous phase. Very low concentrations in tap and surface water samples made a pre-concentration step necessary for this matrix as well as for wastewater treatment plant (WWTP) effluent. Mean recoveries for spiked water samples were between 97% and 109% for tap water and WWTP influent samples and between 102% and 136% for WWTP effluent samples. 90th percentiles of intra-plate and inter-plate coefficients of variation were below 10% for influents and below 20% for effluents and surface water. ILA concentrations were quantified in the range of 33-72 mg/L in influent, 21-49 ng/L in effluent and 18-48 ng/L in surface water samples. During wastewater Treatment the ILA levels were reduced by more than 99%. ILA concentrations of influents determined by ELISA and LC-MS/MS were in good agreement. However, findings in LC-ELISA experiments suggest that the true ILA levels in concentrated samples are lower due to interfering effects of matrix compounds and/or cross-reactants. Yet, the ELISA will be a valuable tool for the performance check and comparison of WWTPs and the localization of fecal matter input into surface waters. KW - ELISA KW - LC-MS/MS KW - Bile acids KW - Fecal marker KW - Surface water KW - Wastewater PY - 2016 U6 - https://doi.org/10.1016/j.jenvman.2016.08.023 SN - 0301-4797 VL - 182 SP - 612 EP - 619 AN - OPUS4-37668 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - JOUR A1 - Oberleitner, Lidia A1 - Garbe, L.-A. A1 - Dahmen-Levison, Ursula A1 - Schneider, Rudolf T1 - Improved strategies for selection and characterization of new monoclonal anticarbamazepine antibodies during the screening process using feces and fluorescence polarization immunoassay N2 - Immunoassays are suitable tools for high-throughput screenings. The prerequisite for accurate determinations by these methods is the selection of an excellent antibody. The production and selection of monoclonal antibodies is usually a tedious process. In this study, new strategies for improving antibody production and characterization were applied. This includes the monitoring of the immunization progress in mice through antibodies extracted from feces, which allows a time-resolved and animal-friendly monitoring of the immune response. Additionally, fluorescence polarization immunoassay (FPIA) could be successfully applied for fast and easy examination of cell culture supernatants and the investigation of antibody/antigen interactions including kinetics and fluorescence properties. These methods simplify the selection of the optimal antibody. As a target analyte, carbamazepine was chosen. This is a widely used antiepileptic drug which also frequently occurs in the environment. The new antibody enables CBZ determination in the concentration range 0.66–110 µg L-1 within 10 min using a high-throughput microtiter plate-based FPIA, and between 1.4 and 79 µg L-1 within 5 min applying an automated cuvette-based FPIA instrument, and from 0.05–36 µg L-1 using ELISA. The measurements were performed at a non-equilibrium state which improved the sensitivity and selectivity of the assays. Due to low cross-reactivity especially towards the main CBZ metabolite and other pharmaceuticals (<1%), this antibody gives the opportunity for application in medical and environmental analyses. KW - Antikörper KW - Emerging Pollutants KW - Schadstoffe KW - Monoklonal KW - Pharmazeutika KW - Arzneistoffe KW - Assay KW - Pesticide parathion-methyl KW - Waste-water KW - Samples KW - Pharmaceuticals KW - Quantification PY - 2016 U6 - http://nbn-resolving.de/urn/resolver.pl?urn:nbn:de:kobv:b43-392473 SN - 1759-9660 VL - 8 SP - 6883 EP - 6894 PB - Royal Society of Chemistry CY - London AN - OPUS4-39247 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - JOUR A1 - Gärtner, Stefanie A1 - Carvalho, J.J. A1 - Emmerling, Franziska A1 - Garbe, L.-A. A1 - Schneider, Rudolf T1 - Polyclonal murine and rabbit antibodies for the bile acid isolithocholic acid N2 - Bile acids are relevant markers for clinical research. This study reports the production of antibodies for isolithocholic acid, the isomer of the extensively studied lithocholic acid. The IgG titer and affinity maturation were monitored during the immunizations of three mice and two rabbits. In both animal models, polyclonal antibodies with a high selectivity and affinity were produced. The development of a direct competitive ELISA with a test midpoint of 0.69 ± 0.05 µg/L and a measurement range from 0.09–15 µg/L is reported. Additionally, the crystal structure of isolithocholic acid is described for the first time. KW - ELISA KW - Crystal structure KW - Competitive assay KW - Immunization monitoring KW - 3ß-hydroxy-5ß-cholanic acid PY - 2015 U6 - https://doi.org/10.1080/15321819.2014.924419 SN - 1532-1819 SN - 1532-4230 VL - 36 IS - 3 SP - 233 EP - 252 PB - Taylor & Francis CY - Philadelphia, Pa. AN - OPUS4-31836 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - JOUR A1 - Drzymala, Sarah A1 - Weiz, S. A1 - Heinze, J. A1 - Marten, S. A1 - Prinz, Carsten A1 - Zimathies, Annett A1 - Garbe, L.-A. A1 - Koch, Matthias T1 - Automated solid-phase extraction coupled online with HPLC-FLD for the quantification of zearalenone in edible oil N2 - 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. KW - Dynamic covalent hydrazine chemistry (DCHC) KW - Maize germ oil KW - Food KW - Mycotoxin KW - ZEA KW - Isomerisation KW - Occurrence KW - Edible oil KW - SIDA KW - LC-MS/MS KW - Quantification PY - 2015 U6 - https://doi.org/10.1007/s00216-015-8541-5 SN - 1618-2642 SN - 1618-2650 VL - 407 IS - 12 SP - 3489 EP - 3497 PB - Springer CY - Berlin AN - OPUS4-33067 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - JOUR A1 - Drzymala, Sarah A1 - Binder, J. A1 - Brodehl, Antje A1 - Penkert, M. A1 - Rosowski, M. A1 - Garbe, L.-A. A1 - Koch, Matthias T1 - Estrogenicity of novel phase I and phase II metabolites of zearalenone and cis-zearalenone N2 - 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. KW - Zearalenone KW - Mycotoxin KW - MCF-7 KW - E-screen assay KW - Estrogenicity KW - Isomerization PY - 2015 U6 - https://doi.org/10.1016/j.toxicon.2015.08.027 SN - 0041-0101 SN - 1879-3150 VL - 105 SP - 10 EP - 12 PB - Elsevier CY - Oxford [u.a.] AN - OPUS4-34167 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER - TY - JOUR A1 - Oberleitner, Lidia A1 - Eremin, S.A. A1 - Lehmann, Andreas A1 - Garbe, L.-A. A1 - Schneider, Rudolf T1 - Fluorescence polarization immunoassays for carbamazepine - comparison of tracers and formats N2 - For the antiepileptic drug and anthropogenic marker carbamazepine (CBZ), a fast and cost-effective immunoassay based on fluorescence polarization (FPIA) was developed. The required fluorophore conjugates were synthesized from different fluorescein and CBZ derivatives. The most suitable tracer was CBZ–triglycine–5-(aminoacetamido)fluorescein. Additionally, the applicability of the assay in tubes and on microtiter plates was tested. The first format can be performed in a portable instrument and therefore can be applied in field measurements. The measurement of an individual sample can be carried out within 4 min. This assay shows a measurement range of 2.5–1000 µg L-1 and a test midpoint (or IC50) of 36 µg L-1. The FPIA performed on microtiter plates is useful for the assay development and is suitable for a very high throughput (up to 24 samples in 20 min). The test midpoint of this assay is 13 µg L-1 and the measurement range is 1.5–300 µg L-1. Furthermore, this assay requires smaller sample volumes and less reagents, including the crucial amount of antibody. The applicability of both assays to spiked surface water samples was evaluated. The recovery rates vary between 66–110% on microtiter plates and 81–140% in tubes. PY - 2015 U6 - https://doi.org/10.1039/c5ay00617a SN - 1759-9660 SN - 1759-9679 VL - 7 IS - 14 SP - 5854 EP - 5861 PB - RSC Publ. CY - Cambridge AN - OPUS4-33821 LA - eng AD - Bundesanstalt fuer Materialforschung und -pruefung (BAM), Berlin, Germany ER -