Article pubs.acs.org/est
In Situ Metal Imaging and Zn Ligand-Speciation in a Soil-Dwelling Sentinel: Complementary Electron Microprobe and Synchrotron Microbeam X‑ray Analyses A. J. Morgan,†,* J. F. W. Mosselmans,‡ J. M. Charnock,§ A. Bennett,∥ C. Winters,† M. O’Reilly,† P. Fisher,⊥ J. Andre,† M. Turner,† P. Gunning,# and P. Kille† †
Cardiff School of Biosciences, Cardiff University, Park Place, Cardiff CF10 3US, Wales, U.K. Diamond Light Source Ltd., Harwell Science and Innovation Campus, Didcot, U.K. § School of Earth, Atmospheric and Environmental Sciences, University of Manchester, Williamson Building, Oxford Road, Manchester, M13 9PL ∥ Science & Technology Facilities Council, Daresbury Laboratory, Daresbury Science & Innovation Campus, Warrington WA4 4AD, Cheshire, U.K. ⊥ School of Earth & Ocean Sciences, Cardiff University, Main Building, Park Place, Cardiff CF10 3AT, U.K. # Smith & Nephew, Heslington, York Science Park, York YO10 5DF, U.K. ‡
ABSTRACT: Understanding the relationships between accumulated metal speciation in cells and tissues of ecologically significant taxa such as earthworms will improve risk assessments. Synchrotron-based μ-focus X-ray spectroscopy was used to detect, localize, and determine ligand-speciation of Zn and Pb in thin sections of two epigeic earthworm species collected from a Pb/Zn-mine soil. The findings indicated that Zn and Pb partition predominantly as typical hard acids (i.e., strong affinities for O-donors) within liverlike chloragocytes. Moreover, Zn speciation was very similar in the chloragog and intestinal epithelia but differed subtly in the kidneylike nephridial tubules; neither Zn nor Pb was detectable in the ventral nerve cord. High resolution X-ray mapping of high pressure-frozen, ultrathin, freeze-substituted sections in a transmission electron microscope (TEM), combined with conventional TEM structural analysis, identified a new cell type packed with highly organized rough endoplasmic reticulum and containing deposits of Cd (codistributed with S); there was no evidence that these cells are major depositories of Zn or Pb. These data may be used in a systems biology approach to assist in the interpretation of metal-evoked perturbations in whole-worm transcriptome and metabolome profiles.
1. INTRODUCTION The total accumulated burden of a metal in an organism’s tissues is of less toxicological consequence than are estimates of the proportion of the burden, which is bioreactive or free to interfere with biochemical targets. Chemical state (valence, identity, and properties of binding ligands, i.e. speciation) influences metal toxicokinetics and toxicodynamics1 but is seldom directly assessed in the biotic subjects of ecotoxicology. Better descriptions of the relationships between metal speciation and toxicity would not only improve current understanding of modes of toxicity in ecologically significant taxa but will ultimately lead to improved risk assessments. Such considerations have probably motivated studies to use differential centrifugation of tissue homogenates to divide the accumulated metal contents of aquatic2,3 and terrestrial 4 invertebrates, including earthworms,5−7 into operationally © 2012 American Chemical Society
defined detoxified- and nondetoxified subcellular metal compartments. Ironically, cellular fractionation of metals in tissue homogenates does not and cannot provide information about metal speciation in morphologically identified cell types. Several in situ physicochemical techniques are available that provide spatially resolved images of metal distribution and, to different degrees, speciation. These include laser ablation inductively coupled mass spectrometry (LA-ICP-MS), nanoscale secondary ion mass spectrometry (NanoSIMS), electron and nuclear microprobe analyses, and synchrotron-based microfocus methods.8 X-ray absorption spectroscopy (XAS) Received: Revised: Accepted: Published: 1073
June 29, 2012 November 23, 2012 December 2, 2012 December 2, 2012 dx.doi.org/10.1021/es302633f | Environ. Sci. Technol. 2013, 47, 1073−1081
Environmental Science & Technology
Article
components of the present study. Although Zn is an essential metal with fundamental and wide-ranging catalytic, structural, and regulatory roles in living systems,20 its toxic effects are known to limit the distributions of earthworms21 and other organisms in mixture-contaminated field sites.
for example, using synchrotron radiation generated by particle accelerators, can now provide detection sensitivity in the micromolar to nanomolar range9,10 and a sufficient spatial resolution (
