The impact of extraction methodologies on the toxicity of sediments in the zebrafish (Danio rerio) embryo test
- 380 Downloads
Traditionally, methods for sediment extractions are characterised using chemical analyses. However, in order to evaluate sediment extracts with regard to biological effects and, thus, bioaccessibility, extraction methods have to be compared to effect data obtained from experiments with in situ exposure scenarios, i.e., sediment contact tests. This study compares four extraction methods and sediment contact test data from a previous project with respect to predictive power in the fish embryo test with zebrafish (Danio rerio).
Materials and methods
A natural and an artificial sediment spiked with a mixture of six organic pollutants (2,4-dinitrophenol, diuron, fluoranthene, nonylphenol, parathion and pentachlorophenol) were extracted using (a) membrane dialysis extraction (MDE), (b) a Soxhlet procedure, (c) hydroxypropyl-β-cyclodextrin (HPCD) or (d) Tenax®-TA. Whereas the former two are regarded being exhaustive with respect to non-covalently bound contaminants, the latter two are considered to predict bioaccessibility. Resulting extracts were tested in the fish embryo assay with D. rerio for embryotoxic and teratogenic potential.
Results and discussion
Mortalities caused by organic extracts from Soxhlet extraction and MDE were high. However, HPCD extracts turned out to be at least as effective as extracts obtained with these two methods. One possible reason might be short ageing time of the spiked sediments. Only Tenax®-TA extracts gave results comparable to the sediment contact assay for natural sediment, but revealed low reproducibility. Significant differences between natural and artificial sediment were found for extracts obtained with techniques using native (i.e., non-freeze-dried) sediments, i.e., HPCD and Tenax®-TA. In contrast, MDE and Soxhlet extracts used freeze-dried sediment and did not differentiate between natural and artificial sediment. Therefore, freeze-drying has likely altered and equalised sediment properties that influence accessibility, such as composition of bacterial communities and organic matter quality.
Four extraction methods were successfully characterised with respect to their stringency and predictiveness for bioaccessibility. MDE was confirmed as an alternative to Soxhlet extraction. High mortalities induced by HPCD extracts underline the need to include ageing into consideration when assessing sediments. Although Tenax®-TA may basically be used to predict bioaccessibility in the fish embryo test, the high variability observed warrants further investigation of the relation between effect and extractability. Apparently, freeze-drying can severely affect sediment properties, potentially eliminating individual properties of natural sediments.
KeywordsCyclodextrin Extraction Fish embryo test Membrane dialysis extraction Tenax Sediment Soxhlet Zebrafish
- Braunbeck T, Boettcher M, Hollert H, Kosmehl T, Lammer E, Leist E, Rudolf M, Seitz N (2005) Towards an alternative for the acute fish LC50 test in chemical assessment: The fish embryo toxicity test goes multi-species—an update. Altex 22:87–102Google Scholar
- DIN (2001) DIN 38415-6: Deutsche Einheitsverfahren zur Wasser-, Abwasser- und Schlammuntersuchung: Suborganismische Testverfahren (Gruppe T) Teil 6: Giftigkeit gegenüber Fischen: Bestimmung der nicht akut giftigen Wirkung von Abwasser auf die Entwicklung von Fischeiern über Verdünnungsstufen (T 6). Deutsches Institut für Normung e. V., Berlin, 14 S.Google Scholar
- EP/EC (2008) Directive 2008/105/EC of the european parliament and of the council of 16 December 2008 on environmental quality standards in the field of water policy, amending and subsequently repealing Council Directives 82/176/EEC, 83/513/EEC, 84/156/EEC, 84/491/EEC, 86/280/EEC and amending Directive 2000/60/EC of the European Parliament and of the Council. http://eur-lex.europa.eu/LexUriServ/LexUriServ.do?uri=OJ:L:2008:348:0084:0097:EN:PDF
- Feiler U, Ahlf W, Fahnenstich C, Gilberg D, Hammers-Wirtz M, Höss S, Hollert H, Melbye K, Meller M, Neumann-Hensel H, Ratte HT, Seiler TB, Spira D, Weber J, Heininger (2009a) Definition von Referenzbedingungen, Kontrollsedimenten und Toxizitätsschwellenwerten für limnische Sedimentkontakttetst (SeKT) - Final project report BfG1614, Federal Institute of Hydrology, Koblenz, Germany, ISBN 978-3-940247-01-8, 251 p.Google Scholar
- Fleming RJ, Holmes D, Nixon SJ (1998) Toxicity of permethrin to Chironomus riparius in artificial and natural sediments. Environ Toxicol Chem 17:1332–1337Google Scholar
- Höss S, Ahlf W, Fahnenstich C, Gilberg D, Hollert H, Melbye K, Meller M, Hammers-Wirtz M, Heininger P, Neumann-Hensel H, Ottermanns R, Ratte HT, Seiler TB, Spira D, Weber J, Feiler U (2010) Variability of freshwater sediment contact tests in sediments with low anthropogenic contamination—determination of toxicity thresholds. Environ Pollut. doi:10.1016/j.envpol.2010.05.013 Google Scholar
- Huckins JN, Tubergen MW, Lebo JA, Gale RW, Schwartz TR (1990) Polymeric film dialysis in organic-solvent media for cleanup of organic contaminants. J Assoc Off Ana Chem 73:290–293Google Scholar
- ISO (International Organization for Standardization) (1996) ISO 7346/3: Water quality—Determination of the acute lethal toxicity of substances to a freshwater fish [Brachydanio rerio, Hamilton-Buchanan (Teleostei, Cyprinidae)]—part 3: flow-through method. Iso GuidelineGoogle Scholar
- Lammer E, Carr GJ, Wendler K, Rawlings JM, Belanger SE, Braunbeck T (2009) Is the fish embryo toxicity test (FET) with the zebrafish (Danio rerio) a potential alternative for the fish acute toxicity test? Comp Biochem Phys C 149:196–209Google Scholar
- Nagel R (2002) DarT: The embryo test with the zebrafish Danio rerio—a general model in ecotoxicology and toxicology. Altex 19:38–48Google Scholar
- OECD (2004) OECD guideline 218: sediment-water chironomid toxicity using spiked sediment. Organisation for Economic Cooperation and Development, BerlinGoogle Scholar
- Reichenberg F, Mayer P (2006) Two complementary sides of bioavailability: accessibility and chemical activity of organic contaminants in sediments and soils. Environ Sci Technol 25:1239–1245Google Scholar
- Seiler TB (2010) Total or biomimetic extracts or direct contact exposure? - Comparative research towards a realistic ecotoxicological characterisation of sediments. PhD thesis, Ruprecht-Karls University Heidelberg, 340 p., http://archiv.ub.uni-heidelberg.de/volltextserver/volltexte/2010/11171/pdf/20100701_Dissertation_Seiler_veroeff.pdf
- Wölz J, Engwall M, Maletz S, Takner HO, van Bavel B, Kammann U, Klempt M, Weber R, Braunbeck T, Hollert H (2008) Changes in toxicity and Ah receptor agonist activity of suspended particulate matter during flood events at the rivers Neckar and Rhine—a mass balance approach using in vitro methods and chemical analysis. Environ Sci Poll Res 15:536–553CrossRefGoogle Scholar
- Wölz J, Cofalla C, Hudjetz S, Roger S, Brinkmann M, Schmidt B, Schaffer A, Kammann U, Lennartz G, Hecker M, Schuttrumpf H, Hollert H (2009) In search for the ecological and toxicological relevance of sediment re-mobilisation and transport during flood events. J Soils Sediments 9:1–5CrossRefGoogle Scholar