Bioconcentration factors (BCF) for regulatory purposes are usually determined by fish flow-through tests according to technical guidance document OECD 305. Fish bioconcentration studies are time consuming, expensive, and use many laboratory animals. The aim of this study was to investigate whether the freshwater amphipod Hyalella azteca can be used as an alternative test organism for bioconcentration studies. Fourteen substances of different hydrophobicity (log Kow 2.4–7.6) were tested under flow-through conditions to determine steady state and kinetic bioconcentration factors (BCFss and BCFk). The results were compared with fish BCF estimates for the same substances described in the literature to show the relationship between both values. Bioconcentration studies with the freshwater amphipod H. azteca resulted in BCF estimates which show a strong correlation with fish BCF values (r2 = 0.69). Hyalella BCF values can be assessed in accordance with the regulatory B criterion (BCF > 2000, i.e., REACH) and thereby enable the prediction of B or non-B classification in the standard fish test. Therefore, H. azteca has a high potential to be used as alternative test organism to fish for bioconcentration studies.Electronic supplementary materialThe online version of this article (10.1007/s11356-018-3677-4) contains supplementary material, which is available to authorized users.
Biotransformation plays a crucial role in regulating the bioaccumulation potential and toxicity of organic compounds in organisms but is, in general, poorly understood for emerging contaminants. Here, we have used diclofenac as a model compound to study the impact of biotransformation on the bioaccumulation potential and toxicity in two keystone aquatic invertebrates: Gammarus pulex and Hyalella azteca. In both species, diclofenac was transformed into several oxidation products and conjugates, including two novel products, that is, diclofenac taurine conjugate (DCF-M403) and unexpected diclofenac methyl ester (DCF-M310.03). The ratios of biotransformation products to parent compound were 12−17 for DCF-M403 and 0.01−0.7 for DCF-M310.03 after 24 h exposure. Bioconcentration factors (BCFs) of diclofenac were 0.5 and 3.2 L kg ww −1 in H. azteca and G. pulex, respectively, whereas BCFs of DCF-M310.03 was 164.5 and 104.7 L kg ww −1, respectively, representing a 25-to 110-fold increase. Acute toxicity of DCF-M310.03 was also higher than the parent compound in both species, which correlated well with the increased bioconcentration potential. The LC 50 of diclofenac in H. azteca was 216 mg L −1 , while that of metabolite DCF-M310.03 was reduced to only 0.53 mg L −1 , representing a 430-fold increase in acute toxicity compared to diclofenac. DCF-M403 is less toxic than its parent compound toward H. azteca, which may be linked to its slightly lower hydrophobicity. Furthermore, the transformation of diclofenac to its methyl ester derivative was explored in crude invertebrate extracts spiked with an S-adenosylmethionine cofactor, revealing possible catalysis by an S-adenosylmethionine-dependent carboxylic acid methyltransferase. Methylation of diclofenac was further detected in fish hepatocytes and human urine, indicating a broader relevance. Therefore, potentially methylated metabolites of polar contaminants should be considered for a comprehensive risk assessment in the future.
Bioaccumulation assessment predominantly relies on the bioconcentration factor (BCF) as the sole decisive metric. The test guideline 305 by the Organisation for Economic Co‐operation and Development (OECD) provides the standard procedure for deriving this in vivo fish BCF, which is not only expensive and labor‐intensive, but also requires many animals. Accordingly, there is a great need for and interest in alternative methods that can help to reduce, replace, and refine vertebrate tests, as described in the 3R principles. Two alternative approaches have been developed: the bioconcentration test with the freshwater amphipod Hyalella azteca and the OECD test guideline 319 which provides a method to determine experimentally derived in vitro metabolism rates that can then be incorporated into in silico prediction models for rainbow trout BCF calculation. In the present study both alternative methods were applied to 5 substances of different physicochemical characteristics. The results were compared with literature values of fish in vivo BCFs and additional BCFs obtained with the alternative methods, if available. Potential differences between the results of the test methods are discussed utilizing information such as in vivo metabolism rates. The currently available data set suggests that these 2 alternative methods pose promising alternatives to predict bioaccumulation in fish, although defined applicability domains have yet to be determined. Environ Toxicol Chem 2020;39:1813–1825. © 2020 The Authors. Environmental Toxicology and Chemistry published by Wiley Periodicals LLC on behalf of SETAC
Background The high production volume of engineered nanomaterials (ENMs) may lead to high pressure on the environment, and a scientific assessment of ENMs that bioaccumulate in organisms and biomagnify in the food web is necessary. Within the regulation of chemicals in several jurisdictions, such as the European regulation REACH, the bioconcentration factor is the standard endpoint. The bioconcentration factor is mostly determined by flow-through fish tests. However, nanomaterials tend to agglomerate, which may lead to sedimentation in aquatic environments. The bioavailability of the tested nanomaterials may be thus impaired for pelagic species, including fish, in comparison to benthic or filtrating species. Several risk assessment regulations allow the usage of data gained during tests using invertebrates and such data may allow a waiver of further tests using vertebrates. The aim of this study was to elucidate the potential of different freshwater invertebrate species to be used in laboratory bioaccumulation studies on ENMs and to give some guidance for the use of bioaccumulation endpoints derived from studies using aquatic invertebrate species in the risk assessment process for ENMs. Results The existing literature related to the testing of nanomaterial bioaccumulation with freshwater invertebrates was screened and reviewed to find suitable test species with regard to their ecology and physiology, as well as laboratory test systems allowing to investigate the bioavailability/bioaccumulation of nanomaterials with the respective species. Bivalvia, gastropoda, isopoda, amphipoda, and branchiopoda were reviewed and their suitability for bioaccumulation testing was assessed. Amphipods and bivalves represent worst-case scenarios and show clear advantages to be used as test organisms. However, only amphipods allow the examination of two clearly independent exposure pathways (water and diet). Conclusion Amphipods are suitable test organisms for bioaccumulation testing of ENMs. The results from amphipod bioconcentration and biomagnification tests can be included in a tiered assessment suggested at the end of this study allowing a clear grading of the tested nanomaterials as “bioaccumulative” or “non bioaccumulative.” Due to the worst-case scenario of the amphipod test, this approach may allow a waiver of further vertebrate tests.
Background The European Water Framework Directive (WFD) requires the monitoring of biota—preferably fish—to check the compliance of tissue concentrations of priority substances (PS) against substance-specific environmental quality standards (EQSs). In monitoring programs, different fish species are covered, which often are secondary consumers with a trophic level (TL) of about 3. For harmonization, a normalization of monitoring data to a common trophic level is proposed, i.e., TL 4 (predatory fish) in freshwaters, so that data would be sufficiently protective. For normalization, the biomagnification properties of the chemicals can be considered by applying substance-specific trophic magnification factors (TMFs). Alternatively, TL-corrected biomagnification factors (BMFTLs) may be applied. Since it is impractical to derive site-specific TMFs or BMFTLs, often data from literature will be used for normalization. However, available literature values for TMFs and BMFTLs are quite varying. In the present study, the use of literature-derived TMFs and BMFTLs in data normalization is studied more closely. Results An extensive literature evaluation was conducted to identify appropriate TMFs for the WFD PS polybrominated diphenyl ethers (PBDE), hexachlorobenzene, perfluorooctane sulfonate (PFOS), dioxins and dioxin-like compounds (PCDD/F + dl-PCB), hexabromocyclododecane, and mercury. The TMFs eventually derived were applied to PS monitoring data sets of fish from different trophic levels (chub, bream, roach, and perch) from two German rivers. For comparison, PFOS and PBDE data were also normalized using literature-retrieved BMFTLs. Conclusions The evaluation illustrates that published TMFs and BMFTLs for WFD PS are quite variable and the selection of appropriate values for TL 4 normalization can be challenging. The normalized concentrations partly included large uncertainties when considering the range of selected TMFs, but indicated whether an EQS exceedance at TL 4 can be expected. Normalization of the fish monitoring data revealed that levels of substances accumulating in the food web (TMF or BMF > 1) can be underestimated when relying on fish with TL < 4 for EQS compliance assessment. The evaluation also revealed that TMF specifically derived for freshwater ecosystems in Europe would be advantageous. Field-derived BMFTLs seemed to be no appropriate alternative to TMFs, because they can vary even stronger than TMFs.
Background The trophic magnification factor (TMF) is a metric that describes the average trophic magnification of a chemical through a food web. TMFs may be used for the risk assessment of chemicals, although TMFs for single compounds can vary considerably between studies despite thorough guidance available in the literature to eliminate potential sources of error. The practical realization of a TMF investigation is quite complex and often only a few chemicals can be investigated due to low sample masses. This study evaluated whether a pragmatic approach involving the large-scale cryogenic sample preparation practices of the German Environmental Specimen Bank (ESB) is feasible. This approach could provide sufficient sample masses for a reduced set of samples allowing screenings for a broad spectrum of substances and by that enabling a systematic comparison of derived TMFs. Furthermore, it was assessed whether plausible TMFs can be derived with the ‘Food web on ice’ approach via a comparison with literature TMF values. Results This investigation at Lake Templin near Potsdam is the first TMF study for a German freshwater ecosystem and aimed to derive TMFs that are appropriate for regulatory purposes. A set of 15 composite biota samples was obtained and analyzed for an extended set of benchmark chemicals such as persistent organic pollutants, mercury and perfluoroalkyl substances. TMFs were calculated for all substances that were present in > 80% of the biota samples. For example, in the case of polychlorinated biphenyls, TMFs from 1.7 to 2.5 were determined and comparisons to literature TMFs determined in other freshwater ecosystems showed similarities. We showed that 32 out of 35 compounds analyzed had TMFs significantly above 1. In the remaining three cases, the correlations were not statistically significant. Conclusions The derived food web samples allow for an on-demand analysis and are ready-to-use for additional investigations. Since substances with non-lipophilic accumulation properties were also included in the list of analyzed substances, we conclude that the ‘Food web on ice’ provides samples which could be used to characterize the trophic magnification potential of substances with unknown bioaccumulation properties in the future which in return could be compared directly to the benchmarking patterns provided here.
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