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Risk assessment of chemicals is usually conducted for individual chemicals whereas mixtures of chemicals occur in the environment. Considering that neuroactive chemicals are a group of contaminants that dominate the environment, it is then imperative to understand the combined effects of mixtures. The commonly used models to predict mixture effects, namely concentration addition (CA) and independent action (IA), are thought to be suitable for mixtures of similarly or dissimilarly acting components, respectively. For mixture toxicity prediction, one important challenge is to clarify whether to group neuroactive substances based on similar mechanisms of action, e.g., same molecular target or rather similar toxicological response, e.g., hyper- or hypoactivity (effect direction). We addressed this by using the spontaneous tail coiling (STC) of zebrafish embryos, which represents the earliest observable motor activity in the developing neural network, as a model to elucidate the link between the mechanism of action and toxicological response. Our objective was to answer the following two questions: (1) Can the mixture models CA or IA be used to predict combined effects for neuroactive chemical mixtures when the components share a similar mode of action (i.e., hyper- or hypoactivity) but show different mechanism of action? (2) Will a mixture of chemicals where the components show opposing effect directions result in an antagonistic combined effect? Results indicate that mixture toxicity of chemicals such as propafenone and abamectin as well as chlorpyrifos and hexaconazole that are known to show different mechanisms of action but similar effect directions were predictable using CA and IA models. This could be interpreted with the convergence of effects on the neural level leading to either a collective activation or inhibition of synapses. We also found antagonistic effects for mixtures containing substances with opposing effect direction. Finally, we discuss how the STC may be used to amend risk assessment.
Afolarin Ogungbemi; Riccardo Massei; Rolf Altenburger; Stefan Scholz; Eberhard Küster. Assessing Combined Effects for Mixtures of Similar and Dissimilar Acting Neuroactive Substances on Zebrafish Embryo Movement. Toxics 2021, 9, 104 .
AMA StyleAfolarin Ogungbemi, Riccardo Massei, Rolf Altenburger, Stefan Scholz, Eberhard Küster. Assessing Combined Effects for Mixtures of Similar and Dissimilar Acting Neuroactive Substances on Zebrafish Embryo Movement. Toxics. 2021; 9 (5):104.
Chicago/Turabian StyleAfolarin Ogungbemi; Riccardo Massei; Rolf Altenburger; Stefan Scholz; Eberhard Küster. 2021. "Assessing Combined Effects for Mixtures of Similar and Dissimilar Acting Neuroactive Substances on Zebrafish Embryo Movement." Toxics 9, no. 5: 104.
Risk assessment of chemicals is usually conducted for individual chemicals whereas mixtures of chemical are occurring in the environment. Considering that neuroactive chemicals are a group of contaminants that dominate in the environment, it is then imperative to understand the combined effects from mixtures. The commonly used models to predict mixture effects, namely concentration addition (CA) and independent action (IA), are thought suitable for mixtures of similarly or dissimilarly acting components, respectively. For mixture toxicity prediction, one important challenge is to clarify whether to group neuroactive substances based on similar mechanisms of action, e.g. same molecular target or rather similar toxicological response, e.g. hyper- or hypoactivity (effect direction). We addressed this by using the spontaneous tail coiling (STC) of zebrafish embryos, which represents the earliest observable motor activity in the developing neural network, as a model to elucidate the link between mechanism of action and toxicological response. Two questions were asked: 1.) Can the mixture models CA or IA be used to predict combined effects for neuroactive chemical mixtures when the components share a similar mode of action (i.e. hyper- or hypoativity) but show different mechanism of action? 2.) Will a mixture of chemicals where the components show opposing effect directions result in an antagonistic combined effect? Results indicate that mixture toxicity of chemicals such as propafenone and abamectin as well as chlorpyrifos and hexaconazole that are known to show different mechanisms of action but similar effect directions were predictable using CA and IA models. This could be interpreted with the convergence of effects on the neural level leading to either a collective activation or inhibition of synapses. We also found antagonistic effects for mixtures containing substances with opposing effect direction. Finally, we discuss how the STC may be used to amend risk assessment.
Afolarin Olaposi Ogungbemi; Riccardo Massei; Rolf Altenburger; Stefan Scholz; Eberhard Küster. Assessing Combined Effects for Mixtures of Similar and Dissimilar Acting Neuroactive Substances on Zebrafish Embryo Movement. 2021, 1 .
AMA StyleAfolarin Olaposi Ogungbemi, Riccardo Massei, Rolf Altenburger, Stefan Scholz, Eberhard Küster. Assessing Combined Effects for Mixtures of Similar and Dissimilar Acting Neuroactive Substances on Zebrafish Embryo Movement. . 2021; ():1.
Chicago/Turabian StyleAfolarin Olaposi Ogungbemi; Riccardo Massei; Rolf Altenburger; Stefan Scholz; Eberhard Küster. 2021. "Assessing Combined Effects for Mixtures of Similar and Dissimilar Acting Neuroactive Substances on Zebrafish Embryo Movement." , no. : 1.
Neuroactive chemicals are frequently detected in the environment. At sufficiently high concentrations or within mixtures, they could provoke neurotoxic effects and neurological diseases to organisms and humans. Fast identification of such neuroactive compounds in the environment could help in hazard assessment and risk mitigation. Behavior change is considered as an important endpoint and might be directly or indirectly connected to a neuroactive mode of action. For a fast evaluation of environmental samples and pure substances, we optimized the measurement of a behavioral endpoint in zebrafish embryos - the spontaneous tail coiling (STC). Evaluation of results is automated via the use of a workflow established with the KNIME® software. Analysis duration and developmental stage were optimized to 1 min and 25 ± 1 hpf respectively during measurement. Exposing the embryos in a group of 10 or 20 and acclimatizing for 30 min at room temperature proved to be reliable. The optimized method was used to investigate neurotoxic effects of 18 substances with different modes of action (MoA). The STC test accurately detected the effect of 8 out of 11 neuroactive substances (chlorpyrifos, chlorpyrifos-oxon, diazinon, paraoxon-methyl, abamectin, carbamazepine, propafenone and diazepam). Aldicarb and nicotine showed subtle effects which were considered to be conditional and imidacloprid showed no effect. For substances with unknown neuroactive MoA, 3 substances did not provoke any effect on the STC (pyraclostrobin, diuron and daunorubicin-hydrochloride) while 4 other substances provoked an increased STC (hexaconazole, aniline, dimethyl-sulfoxide and 3,4-dichloroaniline). Such unexpected effects indicate possible neuroactive side effects or unknown mechanisms of action that impact on the STC. In conclusion, the optimized STC parameters and the automated analysis in KNIME® indicate opportunities for the harmonization of the STC test and further development for prospective and diagnostic testing.
Afolarin O. Ogungbemi; Elisabet Teixido; Riccardo Massei; Stefan Scholz; Eberhard Küster. Optimization of the spontaneous tail coiling test for fast assessment of neurotoxic effects in the zebrafish embryo using an automated workflow in KNIME®. Neurotoxicology and Teratology 2020, 81, 106918 .
AMA StyleAfolarin O. Ogungbemi, Elisabet Teixido, Riccardo Massei, Stefan Scholz, Eberhard Küster. Optimization of the spontaneous tail coiling test for fast assessment of neurotoxic effects in the zebrafish embryo using an automated workflow in KNIME®. Neurotoxicology and Teratology. 2020; 81 ():106918.
Chicago/Turabian StyleAfolarin O. Ogungbemi; Elisabet Teixido; Riccardo Massei; Stefan Scholz; Eberhard Küster. 2020. "Optimization of the spontaneous tail coiling test for fast assessment of neurotoxic effects in the zebrafish embryo using an automated workflow in KNIME®." Neurotoxicology and Teratology 81, no. : 106918.
Tests with zebrafish embryos have gained wide acceptance as an alternative test model for drug development and toxicity testing. In particular, the behavioral response of the zebrafish embryo is currently seen as a useful endpoint to diagnose neuroactive substances. Consequently, several behavioral test methods have been developed addressing various behavioral endpoints such as spontaneous tail coiling (STC), photomotor response (PMR), locomotor response (LMR) and alternating light/dark-induced locomotor response (LMR-L/D). Although these methods are distinct in their application, most of their protocols differ quite strongly in the use of experimental parameters and this is usually driven by different research questions. However, if a single mode of action is to be diagnosed, then varying experimental parameters may cause incoherent behavioral responses (hypo- or hyperactivity) of zebrafish during toxicity assessment. This could lead to inconclusiveness of behavioral test results for use within a prospective and diagnostic risk assessment framework. To investigate the influence of these parameters, we conducted a review of existing behavioral assays to address the following two questions: (1) To what extent do varying experimental parameters influence observed effects in published behavioral test methods? (2) Is the observed behavior change (hypo- or hyperactivity) of zebrafish embryos consistent with the expected mode of action of a chemical? We compiled a set of 18 substances which are anticipated to be neuroactive. We found that behavioral changes are not only affected by chemicals but also variation in the use of experimental parameters across studies seems to have a high impact on the outcome and thus comparability between studies. Four parameters, i.e., exposure concentration, exposure duration, endpoint parameter and developmental stage were the most influential parameters. Varying combinations of these parameters caused a non-reproducible outcome for the hyperactivity expected for the organophosphates; chlorpyrifos and diazinon. We highlighted that the STC test shows a higher capacity to predict the hyperactivity of organophosphates, while PMR and LMR-L/D were more suitable to predict the hypoactivity expected for anticonvulsants. We provide a list of recommendations which, when implemented, may help to exclude the risk of bias due to experimental parameters if similar goals are desired.
Afolarin Ogungbemi; David Leuthold; Stefan Scholz; Eberhard Küster. Hypo- or hyperactivity of zebrafish embryos provoked by neuroactive substances: a review on how experimental parameters impact the predictability of behavior changes. Environmental Sciences Europe 2019, 31, 1 -26.
AMA StyleAfolarin Ogungbemi, David Leuthold, Stefan Scholz, Eberhard Küster. Hypo- or hyperactivity of zebrafish embryos provoked by neuroactive substances: a review on how experimental parameters impact the predictability of behavior changes. Environmental Sciences Europe. 2019; 31 (1):1-26.
Chicago/Turabian StyleAfolarin Ogungbemi; David Leuthold; Stefan Scholz; Eberhard Küster. 2019. "Hypo- or hyperactivity of zebrafish embryos provoked by neuroactive substances: a review on how experimental parameters impact the predictability of behavior changes." Environmental Sciences Europe 31, no. 1: 1-26.
Cathepsins have been proposed as biomarkers of chemical exposure in the zebrafish embryo model but it is unclear whether they can also be used to detect sublethal stress. The present study evaluates three cathepsin types as candidate biomarkers in zebrafish embryos. In addition to other functions, cathepsins are also involved in yolk lysosomal processes for the internal nutrition of embryos of oviparous animals until external feeding starts. The baseline enzyme activity of cathepsin types H, C and L during the embryonic development of zebrafish in the first 96 h post fertilisation was studied. Secondly, the effect of leupeptin, a known cathepsin inhibitor, and four embryotoxic xenobiotic compounds with different modes of action (phenanthrene—baseline toxicity; rotenone—an inhibitor of electron transport chain in mitochondria; DNOC (Dinitro-ortho-cresol)—an inhibitor of ATP synthesis; and tebuconazole—a sterol biosynthesis inhibitor) on in vivo cathepsin H, C and L total activities have been tested. The positive control leupeptin showed effects on cathepsin L at a 20-fold lower concentration compared to the respective LC50 (0.4 mM) of the zebrafish embryo assay (FET). The observed effects on the enzyme activity of the four other xenobiotics were not or just slightly more sensitive (factor of 1.5 to 3), but the differences did not reach statistical significance. Results of this study indicate that the analysed cathepsins are not susceptible to toxins other than the known peptide-like inhibitors. However, specific cathepsin inhibitors might be identified using the zebrafish embryo.
Eberhard Küster; Stefan Kalkhof; Silke Aulhorn; Martin Von Bergen; Ulrike Gündel. Effects of Five Substances with Different Modes of Action on Cathepsin H, C and L Activities in Zebrafish Embryos. International Journal of Environmental Research and Public Health 2019, 16, 3956 .
AMA StyleEberhard Küster, Stefan Kalkhof, Silke Aulhorn, Martin Von Bergen, Ulrike Gündel. Effects of Five Substances with Different Modes of Action on Cathepsin H, C and L Activities in Zebrafish Embryos. International Journal of Environmental Research and Public Health. 2019; 16 (20):3956.
Chicago/Turabian StyleEberhard Küster; Stefan Kalkhof; Silke Aulhorn; Martin Von Bergen; Ulrike Gündel. 2019. "Effects of Five Substances with Different Modes of Action on Cathepsin H, C and L Activities in Zebrafish Embryos." International Journal of Environmental Research and Public Health 16, no. 20: 3956.
The toxicological characterization of sediments is an essential task to monitor the quality of aquatic environments. Many hazardous pollutants may accumulate in sediments and pose a risk to the aquatic community. The present study provides an attempt to integrate a diagnostic whole mixture assessment workflow based on a slightly modified Danio rerio embryo acute toxicity test with chemical characterization. Danio rerio embryos were directly exposed to sieved sediment (≤ 63 μm) for 96 h. Sediment samples were collected from three polluted sites (Kramfors, Sundsvall and Örnsköldsvik) in the Gulf of Bothnia (Sweden) which are characterized by a long history of pulp and paper industry impact. Effect data were supported by chemical analyses of 237 organic pollutants and 30 trace elements. The results show that malformations and neurotoxic compounds are the main drivers of differentiation in chemical and effects analyses, respectively. Specific spinal cord malformations and delayed hatching were observed only in sediments from Kramfors while light hyperactivity was seen only after exposure to sediments from Sundsvall. Our experiments demonstrate that specific chemical profiles lead to specific effect patterns in Danio rerio embryos. In fact, behavioral endpoints could help detect the exposure to neurotoxins, and the observation of body malformations seems to be a potential tool for the identification of site-specific pollutants as polychlorinated biphenyl (PCBs), brominated flame retardants (BFRs) and several pesticides. Overall, results show the suitability of Danio rerio embryos for the fast screening of sediment samples.
Riccardo Massei; Henner Hollert; Martin Krauss; Wolf Von Tümpling; Cindy Weidauer; Peter Haglund; Eberhard Küster; Christine Gallampois; Mats Tysklind; Werner Brack. Toxicity and neurotoxicity profiling of contaminated sediments from Gulf of Bothnia (Sweden): a multi-endpoint assay with Zebrafish embryos. Environmental Sciences Europe 2019, 31, 8 .
AMA StyleRiccardo Massei, Henner Hollert, Martin Krauss, Wolf Von Tümpling, Cindy Weidauer, Peter Haglund, Eberhard Küster, Christine Gallampois, Mats Tysklind, Werner Brack. Toxicity and neurotoxicity profiling of contaminated sediments from Gulf of Bothnia (Sweden): a multi-endpoint assay with Zebrafish embryos. Environmental Sciences Europe. 2019; 31 (1):8.
Chicago/Turabian StyleRiccardo Massei; Henner Hollert; Martin Krauss; Wolf Von Tümpling; Cindy Weidauer; Peter Haglund; Eberhard Küster; Christine Gallampois; Mats Tysklind; Werner Brack. 2019. "Toxicity and neurotoxicity profiling of contaminated sediments from Gulf of Bothnia (Sweden): a multi-endpoint assay with Zebrafish embryos." Environmental Sciences Europe 31, no. 1: 8.
Wastewaters contain complex mixtures of chemicals, which can cause adverse toxic effects in the receiving environment. In the present study, the toxicity removal during wastewater treatment at seven municipal wastewater treatment plants (WWTPs) was investigated using an effect-based approach. A battery of eight bioassays was applied comprising of cytotoxicity, genotoxicity, endocrine disruption and fish embryo toxicity assays. Human cell-based CALUX assays, transgenic larval models and the fish embryo toxicity test were particularly sensitive to WWTP effluents. The results indicate that most effects were significantly reduced or completely removed during wastewater treatment (76-100%), while embryo toxicity, estrogenic activity and thyroid disruption were still detectable in the effluents suggesting that some harmful substances remain after treatment. The responsiveness of the bioassays was compared and the human cell-based CALUX assays showed highest responsiveness in the samples. Additionally, the fish embryo toxicity test and the transgenic larval models for endocrine disrupting effects showed high responsiveness at low sample concentrations in nearly all of the effluent samples. The results showed a similar effect pattern among all WWTPs investigated, indicating that the wastewater composition could be rather similar at different locations. There were no considerable differences in the toxicity removal efficiencies of the treatment plants and no correlation was observed with WWTP characteristics, such as process configuration or sludge age. This study demonstrated that a biotest battery comprising of multiple endpoints can serve as a powerful tool when assessing water quality or water treatment efficiency in a holistic manner. Rather than analyzing the concentrations of a few selected chemicals, bioassays can be used to complement traditional methods of monitoring in the future by assessing sum-parameter based effects, such as mixture effects, and tackling chemicals that are present at concentrations below chemical analytical detection limits.
Pia Välitalo; Riccardo Massei; Ilse Heiskanen; Peter Behnisch; Werner Brack; Andrew Tindall; David Du Pasquier; Eberhard Küster; Anna Mikola; Tobias Schulze; Markus Sillanpää. Effect-based assessment of toxicity removal during wastewater treatment. Water Research 2017, 126, 153 -163.
AMA StylePia Välitalo, Riccardo Massei, Ilse Heiskanen, Peter Behnisch, Werner Brack, Andrew Tindall, David Du Pasquier, Eberhard Küster, Anna Mikola, Tobias Schulze, Markus Sillanpää. Effect-based assessment of toxicity removal during wastewater treatment. Water Research. 2017; 126 ():153-163.
Chicago/Turabian StylePia Välitalo; Riccardo Massei; Ilse Heiskanen; Peter Behnisch; Werner Brack; Andrew Tindall; David Du Pasquier; Eberhard Küster; Anna Mikola; Tobias Schulze; Markus Sillanpää. 2017. "Effect-based assessment of toxicity removal during wastewater treatment." Water Research 126, no. : 153-163.
Not much is known about the biotransformation capability of zebrafish (Danio rerio) embryos. For understanding possible toxicity differences to adult fish, it might be crucial to understand the biotransformation of chemicals in zebrafish embryos i.e. as part of toxicokinetics. The biotransformation capabilities were analysed for two different stages of zebrafish embryos in conjunction with the internal concentrations of a xenobiotic. Zebrafish embryos of the late cleavage/early blastula period (2-26 hpf) and the early pharyngula period (26-50 hpf) were exposed for 24 h to the AhR binding compound benz[a]anthracene (BaA). Time dependent changes in cyp transcription (cyp1a, cyp1b1, cyp1c1 and cyp1c2) as well as concentration & time-dependent courses of BaA in the fish embryo and the exposure medium were analysed. Additionally, the CYP mediated formation of biotransformation products was investigated. We found correlations between transcriptional responses and the internal concentration for both exposure types. These correlations were depending on the start of the exposure i.e. the age of the exposed embryo. While no significant induction of the examined gene transcripts was observed in the first 12 h of exposure beginning in the blastula period a correlation was apparent when exposure started later i.e. in the pharyngula period. A significant induction of cyp1a was detected already after 1.5 h of BaA exposure. Gene transcripts for cyp1b1, cyp1c1 and cyp1c2 showed expressions distinctly different from cyp1a and were, in general, less inducible by BaA in both exposure windows. The toxicokinetic analysis showed that the biotransformation capability was fivefold higher in the older fish embryos. Biotransformation products of phase I reactions were found between 32 hpf and 50 hpf and were tentatively identified as benz[a]anthracene-phenol and benz[a]anthracene-dihydrodiol-epoxide. In conclusion, not only duration but also onset of exposure in relation to the developmental stage of zebrafish embryos is important in the analysis and interpretation of effects due to different biotransformation capabilities.
Agnes Kühnert; Carolina Vogs; Bettina Seiwert; Silke Aulhorn; Rolf Altenburger; Henner Hollert; Eberhard Küster; Wibke Busch. Biotransformation in the zebrafish embryo –temporal gene transcription changes of cytochrome P450 enzymes and internal exposure dynamics of the AhR binding xenobiotic benz[ a ]anthracene. Environmental Pollution 2017, 230, 1 -11.
AMA StyleAgnes Kühnert, Carolina Vogs, Bettina Seiwert, Silke Aulhorn, Rolf Altenburger, Henner Hollert, Eberhard Küster, Wibke Busch. Biotransformation in the zebrafish embryo –temporal gene transcription changes of cytochrome P450 enzymes and internal exposure dynamics of the AhR binding xenobiotic benz[ a ]anthracene. Environmental Pollution. 2017; 230 ():1-11.
Chicago/Turabian StyleAgnes Kühnert; Carolina Vogs; Bettina Seiwert; Silke Aulhorn; Rolf Altenburger; Henner Hollert; Eberhard Küster; Wibke Busch. 2017. "Biotransformation in the zebrafish embryo –temporal gene transcription changes of cytochrome P450 enzymes and internal exposure dynamics of the AhR binding xenobiotic benz[ a ]anthracene." Environmental Pollution 230, no. : 1-11.
The time-resolved uptake of 17 nonionic and ionic polar compounds (logD ≤ 2) with a diversity of functional groups into zebrafish embryos (ZFE) was studied over 96 h of exposure. Among them were pharmaceuticals, pesticides and plant active ingredients. Uptake rates for the diffusion controlled passive uptake through the ZFE membrane ranged from 0.02 to 24 h–1 for the nonionic compounds and were slower for ionic compounds (90% of the initial amount taken up into the ZFE. For six compounds internal concentrations remained very low (rel. int. conc. < 0.2). Besides biotransformation (sulfamethoxazole), poor membrane permeability (cimetidine, colchicine) and also affinity to efflux transporters (atropine and chloramphenicol) are the likely reasons for these low internal concentrations. This study outlines that the uptake of polar compounds into ZFE is influenced by their physicochemical properties. However, biological processes, biotransformation and, likely, efflux can strongly affect the internal concentrations already in early developmental stages of the ZFE. This should be considered in future toxicokinetic modeling. The evaluation of the toxicity of chemicals by ZFE requires toxicokinetic studies of the test compounds and their TPs to increase comparability to effects in fish.
Stephan Brox; Bettina Seiwert; Eberhard Küster; Thorsten Reemtsma. Toxicokinetics of Polar Chemicals in Zebrafish Embryo (Danio rerio): Influence of Physicochemical Properties and of Biological Processes. Environmental Science & Technology 2016, 50, 10264 -10272.
AMA StyleStephan Brox, Bettina Seiwert, Eberhard Küster, Thorsten Reemtsma. Toxicokinetics of Polar Chemicals in Zebrafish Embryo (Danio rerio): Influence of Physicochemical Properties and of Biological Processes. Environmental Science & Technology. 2016; 50 (18):10264-10272.
Chicago/Turabian StyleStephan Brox; Bettina Seiwert; Eberhard Küster; Thorsten Reemtsma. 2016. "Toxicokinetics of Polar Chemicals in Zebrafish Embryo (Danio rerio): Influence of Physicochemical Properties and of Biological Processes." Environmental Science & Technology 50, no. 18: 10264-10272.
The zebrafish embryo (ZFE) is increasingly used in ecotoxicology research but detailed knowledge of its metabolic potential is still limited. This study focuses on the xenobiotic metabolism of ZFE at different life-stages using the pharmaceutical compound clofibric acid as study compound. Liquid chromatography with quadrupole-time-of-flight mass spectrometry (LC-QToF-MS) is used to detect and to identify the transformation products (TPs). In screening experiments, a total of 18 TPs was detected and structure proposals were elaborated for 17 TPs, formed by phase I and phase II metabolism. Biotransformation of clofibric acid by the ZFE involves conjugation with sulfate or glucuronic acid, and, reported here for the first time, with carnitine, taurine, and aminomethanesulfonic acid. Further yet unknown cyclization products were identified using non-target screening that may represent a new detoxification pathway. Sulfate containing TPs occurred already after 3 h of exposure (7 hpf), and from 48 h of exposure (52 hpf) onwards, all TPs were detected. The detection of these TPs indicates the activity of phase I and phase II enzymes already at early life-stages. Additionally, the excretion of one TP into the exposure medium was observed. The results of this study outline the high metabolic potential of the ZFE with respect to the transformation of xenobiotics. Similarities but also differences to other test systems were observed. Biotransformation of test chemicals in toxicity testing with ZFE may therefore need further consideration.
Stephan Brox; Bettina Seiwert; Nora Haase; Eberhard Küster; Thorsten Reemtsma. Metabolism of clofibric acid in zebrafish embryos ( Danio rerio ) as determined by liquid chromatography–high resolution–mass spectrometry. Comparative Biochemistry and Physiology Part C: Toxicology & Pharmacology 2016, 185-186, 20 -28.
AMA StyleStephan Brox, Bettina Seiwert, Nora Haase, Eberhard Küster, Thorsten Reemtsma. Metabolism of clofibric acid in zebrafish embryos ( Danio rerio ) as determined by liquid chromatography–high resolution–mass spectrometry. Comparative Biochemistry and Physiology Part C: Toxicology & Pharmacology. 2016; 185-186 ():20-28.
Chicago/Turabian StyleStephan Brox; Bettina Seiwert; Nora Haase; Eberhard Küster; Thorsten Reemtsma. 2016. "Metabolism of clofibric acid in zebrafish embryos ( Danio rerio ) as determined by liquid chromatography–high resolution–mass spectrometry." Comparative Biochemistry and Physiology Part C: Toxicology & Pharmacology 185-186, no. : 20-28.
Three water contaminants were selected to be tested in the zebrafish embryo toxicity test (DarT) in order to investigate the sensitivity of the zebrafish embryo toxicity test with respect to mixture effect detection. The concentration-response curves for the observed effects lethality and hypo-pigmentation were calculated after an exposure of the embryos for 96 h with a fungicide (carbendazim), a plasticizer or propellent precursor (2,4-DNT: 2,4- dinitrotoluene) and an aromatic compound (AαC: 2-amino-9H-pyrido[2,3-b]indol), respectively. Follow-up mixture tests were based on the calculated LC50 or EC50 of the single compounds and combined effects were predicted according to the mixture concepts of concentration addition (CA) and independent action (IA). The order of toxicity for the single substances was carbendazim (LC50 = 1.25 μM) < AαC (LC50 = 8.16 μM) < 2,4-DNT (LC50 = 177.05 μM). For AαC and 2,4 DNT hypo-pigmentation was observed in addition (AαC EC50 = 1.81 μM; 2,4-DNT EC50 = 8.81 μM). Two binary and one ternary mixture were studied on lethality and one on hypo-pigmentation: 2,4-DNT/AαC (LC50 = 119.21 μM, EC50 = 5.37 μM), carbendazim/AαC (LC50 = 4.49 μM) and AαC/Carbendazim/2,4 DNT (LC50 = 108.62 μM). Results showed that the effects were in agreement with the CA model when substances were tested in mixtures. Therefore, in a reasonable worst case scenario substance combination effects in fish embryos were at maximum only prone to overestimation when using CA as the mixture concept.
Susanne Schmidt; Wibke Busch; Rolf Altenburger; Eberhard Küster. Mixture toxicity of water contaminants-effect analysis using the zebrafish embryo assay (Danio rerio). Chemosphere 2016, 152, 503 -512.
AMA StyleSusanne Schmidt, Wibke Busch, Rolf Altenburger, Eberhard Küster. Mixture toxicity of water contaminants-effect analysis using the zebrafish embryo assay (Danio rerio). Chemosphere. 2016; 152 ():503-512.
Chicago/Turabian StyleSusanne Schmidt; Wibke Busch; Rolf Altenburger; Eberhard Küster. 2016. "Mixture toxicity of water contaminants-effect analysis using the zebrafish embryo assay (Danio rerio)." Chemosphere 152, no. : 503-512.
Morphology and physiology of fish embryos undergo dramatic changes during their development until the onset of feeding, supplied only by endogenous yolk reserves. For obtaining an insight how these restructuring processes are reflected by body mass related parameters, dry weights (dw), contents of the elements carbon and nitrogen and lipid and protein levels were quantified in different stages within the first four days of embryo development of the zebrafish (Danio rerio). The data show age dependent changes in tissue composition. Dry weights decreased significantly from 79μgdw/egg at 0hours post fertilization (hpf) to 61 μgdw/egg after 96 hpf. The amounts of total carbon fluctuated between 460 mg g-1 and 540 mg g-1 dw, nitrogen was at about 100 mg g-1 dw and total fatty acids were between 48–73 mg g-1 dw. In contrast to these parameters that remained relatively constant, the protein content, which was 240 mg g-1 at 0 hpf, showed an overall increase of about 40%. Comparisons of intact eggs and dechorionated embryos at stages prior to hatching (24, 30, 48 hpf) showed that the differences seen for dry weight and for carbon and nitrogen contents became smaller at more advanced stages, consistent with transition of material from the chorion to embryo tissue. Further, we determined the effect of 2,4-dinitrophenol at a subacutely toxic concentration (14 μM, LC10) as a model chemical challenge on the examined body mass related parameters. The compound caused significant decreases in phospholipid and glycolipid fatty acid contents along with a decrease in the phospholipid fatty acid unsaturation index. No major changes were observed for the other examined parameters. Lipidomic studies as performed here may thus be useful for determining subacute effects of lipophilic organic compounds on lipid metabolism and on cellular membranes of zebrafish embryos.
Nancy Hachicho; Sarah Reithel; Anja Miltner; Hermann J. Heipieper; Eberhard Küster; Till Luckenbach. Body Mass Parameters, Lipid Profiles and Protein Contents of Zebrafish Embryos and Effects of 2,4-Dinitrophenol Exposure. PLOS ONE 2015, 10, e0134755 .
AMA StyleNancy Hachicho, Sarah Reithel, Anja Miltner, Hermann J. Heipieper, Eberhard Küster, Till Luckenbach. Body Mass Parameters, Lipid Profiles and Protein Contents of Zebrafish Embryos and Effects of 2,4-Dinitrophenol Exposure. PLOS ONE. 2015; 10 (8):e0134755.
Chicago/Turabian StyleNancy Hachicho; Sarah Reithel; Anja Miltner; Hermann J. Heipieper; Eberhard Küster; Till Luckenbach. 2015. "Body Mass Parameters, Lipid Profiles and Protein Contents of Zebrafish Embryos and Effects of 2,4-Dinitrophenol Exposure." PLOS ONE 10, no. 8: e0134755.
The chorion and the perivitelline space which surround unhatched zebrafish embryos (ZFE, Danio rerio) may affect the determination of internal concentrations of study compounds taken up in early life-stages of ZFE. Internal concentration-time profiles were gathered for benzocaine, caffeine, clofibric acid, metribuzin and phenacetin as study compounds over 96 h of exposure starting with ZFE at 4h post-fertilization. Liquid chromatography coupled to tandem-mass spectrometry (LC-MS/MS) was used to determine the concentration of the study compounds from intact (i.e. unhatched), dechorionated and from hatched ZFE. The mass of the study compounds per ZFE was 5-30 ng higher for intact ZFE compared to dechorionated ones. Thus, internal concentrations were overestimated if only intact ZFE were analyzed. Dechorionation of unhatched ZFE after their exposure is proposed to determine the true internal concentration in the embryo. For the compounds studied here the mass of the study compounds determined in unhatched ZFE after a short term (5 min) exposure provided a reasonable estimate of the mass taken up by the chorion and the PVS. This mass can be subtracted from the total mass found in unhatched ZFE to calculate the true internal mass. Estimating the mass in the chorion and the PVS from the concentration of the study compound in the external exposure medium and the volume of the PVS provided no reasonable results.
Stephan Brox; Axel P. Ritter; Eberhard Küster; Thorsten Reemtsma. Influence of the perivitelline space on the quantification of internal concentrations of chemicals in eggs of zebrafish embryos (Danio rerio). Aquatic Toxicology 2014, 157, 134 -140.
AMA StyleStephan Brox, Axel P. Ritter, Eberhard Küster, Thorsten Reemtsma. Influence of the perivitelline space on the quantification of internal concentrations of chemicals in eggs of zebrafish embryos (Danio rerio). Aquatic Toxicology. 2014; 157 ():134-140.
Chicago/Turabian StyleStephan Brox; Axel P. Ritter; Eberhard Küster; Thorsten Reemtsma. 2014. "Influence of the perivitelline space on the quantification of internal concentrations of chemicals in eggs of zebrafish embryos (Danio rerio)." Aquatic Toxicology 157, no. : 134-140.
The toxic potency of chemicals is determined well by using the internal effect concentration accounting for differences in toxicokinetic processes and mechanisms of toxic action. This present study examines toxicokinetics of specifically acting and reactive chemicals in the green algae Scenedesmus vacuolatus by using an indirect method. Concentration depletion in the exposure medium was measured for chemicals of lower (log Kow < 3: isoproturon, metazachlor, paraquat) and moderate (log Kow 4‐5: irgarol, triclosan, N‐phenyl‐2‐naphtylamine) hydrophobicity at seven time points over 240 min or 360 min. Uptake and overall elimination rates were estimated by fitting a toxicokinetic model to the observed concentration depletions. The equilibrium of exposure concentrations was reached within minutes to hours or was even not observed within the exposure time. Kinetics of bioconcentration cannot be explained by the chemical's hydrophobicity only, but influencing factors such as the ionization of chemicals, the ion‐trapping mechanism or the potential susceptibility for biotransformation are discussed. Internal effect concentrations associated to 50% inhibition of Scenedesmus vacuolatus reproduction were predicted by linking the bioconcentration kinetics to the effect concentrations and ranged from 0.048 to 7.61 mmol/kg wet weight for specifically acting and reactive chemicals. Knowing the time‐course of the internal effect concentration may help to understand toxicity processes like delayed toxicity, carry‐over toxicity or mixture toxicity in future studies. Environ Toxicol Chem © 2014 SETAC
Carolina Vogs; Agnes Kühnert; Christine Hug; Eberhard Küster; Rolf Altenburger. A toxicokinetic study of specifically acting and reactive organic chemicals for the prediction of internal effect concentrations inScenedesmus vacuolatus. Environmental Toxicology and Chemistry 2014, 34, 100 -111.
AMA StyleCarolina Vogs, Agnes Kühnert, Christine Hug, Eberhard Küster, Rolf Altenburger. A toxicokinetic study of specifically acting and reactive organic chemicals for the prediction of internal effect concentrations inScenedesmus vacuolatus. Environmental Toxicology and Chemistry. 2014; 34 (1):100-111.
Chicago/Turabian StyleCarolina Vogs; Agnes Kühnert; Christine Hug; Eberhard Küster; Rolf Altenburger. 2014. "A toxicokinetic study of specifically acting and reactive organic chemicals for the prediction of internal effect concentrations inScenedesmus vacuolatus." Environmental Toxicology and Chemistry 34, no. 1: 100-111.
An analytical method using high-performance liquid chromatography-tandem mass spectrometry was developed to determine internal concentrations of 34 test compounds such as pharmaceuticals and pesticides in zebrafish embryos (ZFE), among them, cimetidine, 2,4-dichlorophenoxyacetic acid, metoprolol, atropine and phenytoin. For qualification and quantification, multiple reaction monitoring mode was used. The linear range extends from 0.075 ng/mL for thiacloprid and metazachlor and 7.5 ng/mL for coniine and clofibrate to 250 ng/mL for many of the test compounds. Matrix effects were strongest for nicotine, but never exceeded ±20 % for any of the developmental stages of the ZFE. Method recoveries ranged from 90 to 110 % from an analysis of nine pooled ZFE. These findings together with the simple sample preparation mean this approach is suitable for the determination of internal concentrations from only nine individual ZFE in all life stages up to 96 h post-fertilization. Exemplarily, the time course of the internal concentrations of clofibric acid, metribuzin and benzocaine in ZFE was studied over 96 h, and three different patterns were distinguished, on the basis of the speed and extent of uptake and whether or not a steady state was reached. Decreasing internal concentrations may be due to metabolism in the ZFE.
Stephan Brox; Axel P. Ritter; Eberhard Küster; Thorsten Reemtsma. A quantitative HPLC–MS/MS method for studying internal concentrations and toxicokinetics of 34 polar analytes in zebrafish (Danio rerio) embryos. Analytical and Bioanalytical Chemistry 2014, 406, 4831 -4840.
AMA StyleStephan Brox, Axel P. Ritter, Eberhard Küster, Thorsten Reemtsma. A quantitative HPLC–MS/MS method for studying internal concentrations and toxicokinetics of 34 polar analytes in zebrafish (Danio rerio) embryos. Analytical and Bioanalytical Chemistry. 2014; 406 (20):4831-4840.
Chicago/Turabian StyleStephan Brox; Axel P. Ritter; Eberhard Küster; Thorsten Reemtsma. 2014. "A quantitative HPLC–MS/MS method for studying internal concentrations and toxicokinetics of 34 polar analytes in zebrafish (Danio rerio) embryos." Analytical and Bioanalytical Chemistry 406, no. 20: 4831-4840.
Tests with vertebrates are an integral part of environmental hazard identification and risk assessment of chemicals, plant protection products, pharmaceuticals, biocides, feed additives and effluents. These tests raise ethical and economic concerns and are considered as inappropriate for assessing all of the substances and effluents that require regulatory testing. Hence, there is a strong demand for replacement, reduction and refinement strategies and methods. However, until now alternative approaches have only rarely been used in regulatory settings. This review provides an overview on current regulations of chemicals and the requirements for animal tests in environmental hazard and risk assessment. It aims to highlight the potential areas for alternative approaches in environmental hazard identification and risk assessment. Perspectives and limitations of alternative approaches to animal tests using vertebrates in environmental toxicology, i.e., mainly fish and amphibians, are discussed. Free access to existing (proprietary) animal test data, availability of validated alternative methods and a practical implementation of conceptual approaches such as the Adverse Outcome Pathways and Integrated Testing Strategies were identified as major requirements towards the successful development and implementation of alternative approaches. Although the focus of the article is on European regulations, the considerations and conclusions are of global relevance.JRC.I.5-Systems Toxicolog
Stefan Scholz; Erika Sela; Ludek Blaha; Thomas Braunbeck; Malyka Galay-Burgos; Mauricio Garcia-Franco; Joaquin Guinea; Nils Klüver; Kristin Schirmer; Katrin Tanneberger; Marysia Tobor-Kapłon; Hilda Witters; Scott Belanger; Emilio Benfenati; Stuart Creton; Mark Cronin; Rik I.L. Eggen; Michelle Embry; Drew Ekman; Anne Gourmelon; Marlies Halder; Barry Hardy; Thomas Hartung; Bruno Hubesch; Dirk Jungmann; Mark Lampi; Lucy Ej Lee; Marc Léonard; Eberhard Küster; Adam Lillicrap; Till Luckenbach; Albertinka J. Murk; José M Navas; Willie Peijnenburg; Guillermo Repetto; Edward Salinas; Gerrit Schüürmann; Horst Spielmann; Knut Erik Tollefsen; Susanne Walter-Rohde; Graham Whale; James Wheeler; Matthew Winter. A European perspective on alternatives to animal testing for environmental hazard identification and risk assessment. Regulatory Toxicology and Pharmacology 2013, 67, 506 -530.
AMA StyleStefan Scholz, Erika Sela, Ludek Blaha, Thomas Braunbeck, Malyka Galay-Burgos, Mauricio Garcia-Franco, Joaquin Guinea, Nils Klüver, Kristin Schirmer, Katrin Tanneberger, Marysia Tobor-Kapłon, Hilda Witters, Scott Belanger, Emilio Benfenati, Stuart Creton, Mark Cronin, Rik I.L. Eggen, Michelle Embry, Drew Ekman, Anne Gourmelon, Marlies Halder, Barry Hardy, Thomas Hartung, Bruno Hubesch, Dirk Jungmann, Mark Lampi, Lucy Ej Lee, Marc Léonard, Eberhard Küster, Adam Lillicrap, Till Luckenbach, Albertinka J. Murk, José M Navas, Willie Peijnenburg, Guillermo Repetto, Edward Salinas, Gerrit Schüürmann, Horst Spielmann, Knut Erik Tollefsen, Susanne Walter-Rohde, Graham Whale, James Wheeler, Matthew Winter. A European perspective on alternatives to animal testing for environmental hazard identification and risk assessment. Regulatory Toxicology and Pharmacology. 2013; 67 (3):506-530.
Chicago/Turabian StyleStefan Scholz; Erika Sela; Ludek Blaha; Thomas Braunbeck; Malyka Galay-Burgos; Mauricio Garcia-Franco; Joaquin Guinea; Nils Klüver; Kristin Schirmer; Katrin Tanneberger; Marysia Tobor-Kapłon; Hilda Witters; Scott Belanger; Emilio Benfenati; Stuart Creton; Mark Cronin; Rik I.L. Eggen; Michelle Embry; Drew Ekman; Anne Gourmelon; Marlies Halder; Barry Hardy; Thomas Hartung; Bruno Hubesch; Dirk Jungmann; Mark Lampi; Lucy Ej Lee; Marc Léonard; Eberhard Küster; Adam Lillicrap; Till Luckenbach; Albertinka J. Murk; José M Navas; Willie Peijnenburg; Guillermo Repetto; Edward Salinas; Gerrit Schüürmann; Horst Spielmann; Knut Erik Tollefsen; Susanne Walter-Rohde; Graham Whale; James Wheeler; Matthew Winter. 2013. "A European perspective on alternatives to animal testing for environmental hazard identification and risk assessment." Regulatory Toxicology and Pharmacology 67, no. 3: 506-530.
This article introduces ‘Tox-Box’, a joint research project designed to develop a holistic approach towards a harmonized testing strategy for exposure- and hazard-based risk management of anthropogenic trace substances in drinking water to secure a long-term drinking water supply. The main task of the Tox-Box consortium is to enhance the existing health-related indicator value concept (German: GOW-Konzept - Gesundheitlicher Orientierungswert) through development and prioritization of additional end point-related testing strategies for genotoxicity, neurotoxicity, germ cell damage, and endocrine effects. In this context, substance-specific modes of action will be identified and characterized. Toxicological data collected by the 12 Tox-Box subprojects will be evaluated and weighted to structure a hierarchical testing strategy for an improved risk assessment. A technical guidance document for exposure and hazard-based risk management of anthropogenic trace substances in drinking water will eventually be prepared.
Tamara Grummt; Jochen Kuckelkorn; Arnold Bahlmann; Christa Baumstark-Khan; Werner Brack; Thomas Braunbeck; Sebastian Feles; Stefan Gartiser; Hansruedi Glatt; Rita Heinze; Christine E Hellweg; Henner Hollert; Ralf Junek; Martina Knauer; Birgit Kneib-Kissinger; Meike Kramer; Martin Krauss; Eberhard Küster; Sibylle Maletz; Walter Meinl; Abu Noman; Eva-Maria Prantl; Elke Rabbow; Regine Redelstein; Petra Rettberg; Walter Schadenboeck; Carsten Schmidt; Tobias Schulze; Thomas-Benjamin Seiler; Luis Spitta; Daniel Stengel; Petra Waldmann; Alexander Eckhardt. Tox-Box: securing drops of life - an enhanced health-related approach for risk assessment of drinking water in Germany. Environmental Sciences Europe 2013, 25, 27 .
AMA StyleTamara Grummt, Jochen Kuckelkorn, Arnold Bahlmann, Christa Baumstark-Khan, Werner Brack, Thomas Braunbeck, Sebastian Feles, Stefan Gartiser, Hansruedi Glatt, Rita Heinze, Christine E Hellweg, Henner Hollert, Ralf Junek, Martina Knauer, Birgit Kneib-Kissinger, Meike Kramer, Martin Krauss, Eberhard Küster, Sibylle Maletz, Walter Meinl, Abu Noman, Eva-Maria Prantl, Elke Rabbow, Regine Redelstein, Petra Rettberg, Walter Schadenboeck, Carsten Schmidt, Tobias Schulze, Thomas-Benjamin Seiler, Luis Spitta, Daniel Stengel, Petra Waldmann, Alexander Eckhardt. Tox-Box: securing drops of life - an enhanced health-related approach for risk assessment of drinking water in Germany. Environmental Sciences Europe. 2013; 25 (1):27.
Chicago/Turabian StyleTamara Grummt; Jochen Kuckelkorn; Arnold Bahlmann; Christa Baumstark-Khan; Werner Brack; Thomas Braunbeck; Sebastian Feles; Stefan Gartiser; Hansruedi Glatt; Rita Heinze; Christine E Hellweg; Henner Hollert; Ralf Junek; Martina Knauer; Birgit Kneib-Kissinger; Meike Kramer; Martin Krauss; Eberhard Küster; Sibylle Maletz; Walter Meinl; Abu Noman; Eva-Maria Prantl; Elke Rabbow; Regine Redelstein; Petra Rettberg; Walter Schadenboeck; Carsten Schmidt; Tobias Schulze; Thomas-Benjamin Seiler; Luis Spitta; Daniel Stengel; Petra Waldmann; Alexander Eckhardt. 2013. "Tox-Box: securing drops of life - an enhanced health-related approach for risk assessment of drinking water in Germany." Environmental Sciences Europe 25, no. 1: 27.
In ecotoxicity assessment, the ambient exposure concentration is typically applied to quantify the toxic potential of xenobiotic substances. However, exposure and organism-related differences in bioconcentration often cause a considerable variability of toxicity data. This can be minimized by using the internal organism concentration, because toxicokinetic modifying factors are considered implicitly. In the present study, the relationship between ambient and internal concentration-time profiles was investigated for zebrafish (Danio rerio) embryos. The aim was to gain a better understanding and interpretation of exposure-based methods using this model organism. For this purpose, a simple and effective approach to determine the internal concentration was developed. Embryos were exposed to a series of 4 neutral organic substances (naphthalene, fluorene, fluoranthene, benz[a]anthracene) of different hydrophobicity for 72 h. The internal and ambient concentrations were measured at 8 to 9 time points. Kinetics of uptake and elimination were modeled using a first-order 1-compartment model. Biotransformation processes appeared to influence the internal concentrations of fluoranthene and benz[a]anthracene after 48 h. The bioconcentration factors (BCFs) obtained are in excellent agreement with those determined in previous studies using radiolabeled substances. The method demonstrated in the present study is a further step toward a refined ecotoxicity assessment using fish embryos, which links toxicity to the chemical concentration within the organism. This system may also be considered as an alternative to animal testing for BCF determination.
Agnes Kühnert; Carolina Vogs; Rolf Altenburger; Eberhard Küster. The internal concentration of organic substances in fish embryos-A toxicokinetic approach. Environmental Toxicology and Chemistry 2013, 32, 1819 -1827.
AMA StyleAgnes Kühnert, Carolina Vogs, Rolf Altenburger, Eberhard Küster. The internal concentration of organic substances in fish embryos-A toxicokinetic approach. Environmental Toxicology and Chemistry. 2013; 32 (8):1819-1827.
Chicago/Turabian StyleAgnes Kühnert; Carolina Vogs; Rolf Altenburger; Eberhard Küster. 2013. "The internal concentration of organic substances in fish embryos-A toxicokinetic approach." Environmental Toxicology and Chemistry 32, no. 8: 1819-1827.
Ecotoxicological bioassays, e.g. based on Danio rerio teratogenicity (DarT) or the acute luminescence inhibition with Vibrio fischeri, could potentially lead to significant benefits for detecting on site contaminations on qualitative or semi-quantitative bases. The aim was to use the observed effects of two ecotoxicological assays for estimating the extent of a Benzene groundwater contamination plume. We used a Maximum Entropy (MaxEnt) method to rebuild a bivariate probability table that links the observed toxicity from the bioassays with Benzene concentrations. Compared with direct mapping of the contamination plume as obtained from groundwater samples, the MaxEnt concentration map exhibits on average slightly higher concentrations though the global pattern is close to it. This suggest MaxEnt is a valuable method to build a relationship between quantitative data, e.g. contaminant concentrations, and more qualitative or indirect measurements, in a spatial mapping framework, which is especially useful when clear quantitative relation is not at hand.
Agung Wahyudi; Mariana Bartzke; Eberhard Küster; Patrick Bogaert. Maximum entropy estimation of a Benzene contaminated plume using ecotoxicological assays. Environmental Pollution 2012, 172, 170 -179.
AMA StyleAgung Wahyudi, Mariana Bartzke, Eberhard Küster, Patrick Bogaert. Maximum entropy estimation of a Benzene contaminated plume using ecotoxicological assays. Environmental Pollution. 2012; 172 ():170-179.
Chicago/Turabian StyleAgung Wahyudi; Mariana Bartzke; Eberhard Küster; Patrick Bogaert. 2012. "Maximum entropy estimation of a Benzene contaminated plume using ecotoxicological assays." Environmental Pollution 172, no. : 170-179.
Concentration-response experiments, based on the testing of less replicates in favour of more exposure concentrations, represent the typical design of choice applied in toxicological and ecotoxicological effect assessment studies using traditional endpoints such as lethality. However, to our knowledge this concept has not found implementation in the increasingly applied OMICS techniques studying thousands of molecular endpoints at the same time. The present study is among the first applying the concentration-response concept for an ecotoxicoproteomics study. The effects of six different concentrations in the low effect range (
Ulrike Gündel; Stefan Kalkhof; Dimitar Zitzkat; Martin von Bergen; Rolf Altenburger; Eberhard Küster. Concentration–response concept in ecotoxicoproteomics: Effects of different phenanthrene concentrations to the zebrafish (Danio rerio) embryo proteome. Ecotoxicology and Environmental Safety 2012, 76, 11 -22.
AMA StyleUlrike Gündel, Stefan Kalkhof, Dimitar Zitzkat, Martin von Bergen, Rolf Altenburger, Eberhard Küster. Concentration–response concept in ecotoxicoproteomics: Effects of different phenanthrene concentrations to the zebrafish (Danio rerio) embryo proteome. Ecotoxicology and Environmental Safety. 2012; 76 (2):11-22.
Chicago/Turabian StyleUlrike Gündel; Stefan Kalkhof; Dimitar Zitzkat; Martin von Bergen; Rolf Altenburger; Eberhard Küster. 2012. "Concentration–response concept in ecotoxicoproteomics: Effects of different phenanthrene concentrations to the zebrafish (Danio rerio) embryo proteome." Ecotoxicology and Environmental Safety 76, no. 2: 11-22.