Synchrotron radiation based confocal micro-XRF was employed to unravel the tissue-specific three-dimensional (3D) distribution of metals down to trace concentration levels in a non-destructive manner within the crustacean Daphnia magna, an ecotoxicological model organism. Next to the analytical characterization of the employed confocal micro-XRF set-up, specific areas of metal accumulation in different cross-sections of interest within the organism were investigated. The use of a fast dynamic (continuous) scanning approach delivered 3D information on major, minor and trace element distributions in specific sub-regions within Daphnia magna. Coupling the obtained elemental information with microscopic morphological data obtained by laboratory absorption microtomography, full 3D element-to-tissue correlation could be derived, allowing a more detailed interpretation of the obtained results with respect to metal accumulation within this model organism.

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In the field of environmental toxicology, the study on the effects of the presence of transition metals such as Cu, Ni and Zn on the health of pelagic and benthic invertebrates is an important research topic. Fast dynamic 2D micro-XRF scanning and dynamic XRF micro-CT experiments on D. magna, a frequently used ecotoxicological model organism, allow the quantitative investigation of the accumulation of metals within specific organs with microscopic resolution. K-means clustering allows comparison of the concentrations of the elements of interest between different tissues/organs. Principal component analysis allows most of the significant statistical variations in these large and complex multivariate data sets to be explained. Also, by combining SR-XRF and absorption CT data sets using appropriate software packages, it becomes possible to unravel the tissue-specific 2D/3D distribution of metals in-situ within delicate organic samples on the 3-15 mu m resolution level in an essentially non-destructive manner

Recent studies have suggested that exposure of the freshwater invertebrate Daphnia magna to dietary Zn may selectively affect reproduction without an associated increase of whole body bioaccumulation of Zn. The aim of the current research was therefore to investigate the hypothesis that dietary Zn toxicity is the result of selective accumulation in tissues that are directly involved in reproduction. Since under field conditions simultaneous exposure to both waterborne and dietary Zn is likely to occur, it was also tested if accumulation and toxicity under combined waterborne and dietary Zn exposure is the result of interactive effects. To this purpose, D. magna was exposed during a 16-day reproduction assay to Zn following a 5 × 2 factorial design, comprising five waterborne concentrations (12, 65, 137, 207, and 281 μg Zn/L) and two dietary Zn levels (49.6 and 495.9 μg Zn/g dry wt.). Tissue-specific Zn distribution was quantified by synchrotron radiation based confocal X-ray fluorescence (XRF). It was observed that the occurrence of reproductive inhibition due to increasing waterborne Zn exposure (from 65 μg/L to 281 μg/L) was accompanied by a relative increase of the Zn burdens which was similar in all tissues considered (i.e., the carapax, eggs, thoracic appendages with gills and the cluster comprising gut epithelium, storage cells and ovaries). In contrast, the impairment of reproduction during dietary Zn exposure was accompanied by a clearly discernible Zn accumulation in the eggs only (at 65 μg/L of waterborne Zn). During simultaneous exposure, bioaccumulation and toxicity were the result of interaction, which implies that the tissue-specific bioaccumulation and toxicity following dietary Zn exposure are dependent on the Zn concentration in the water. Our findings emphasize that (i) effects of dietary Zn exposure should preferably not be investigated in isolation from waterborne Zn exposure, and that (ii) XRF enabled us to provide possible links between tissue-specific bioaccumulation and reproductive effects of Zn.