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Environmental Measurements Methods
Quantification of Mercury Bioavailability for Methylation Using Diffusive Gradient in Thin-Film Samplers Udonna Ndu, Geoff A. Christensen, Nelson A. Rivera, Caitlin M Gionfriddo, Marc A. Deshusses, Dwayne A Elias, and Heileen Hsu-Kim Environ. Sci. Technol., Just Accepted Manuscript • DOI: 10.1021/acs.est.8b00647 • Publication Date (Web): 19 Jun 2018 Downloaded from http://pubs.acs.org on June 21, 2018
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Environmental Science & Technology
Quantification of Mercury Bioavailability for Methylation Using Diffusive Gradient in Thin-Film Samplers
Udonna Ndu1, Geoff A. Christensen2, Nelson A. Rivera1, Caitlin M. Gionfriddo2, Marc A. Deshusses1, Dwayne A. Elias2, and Heileen Hsu-Kim1,* 1
Department of Civil and Environmental Engineering, Duke University, Box 90287, Durham, North Carolina, 27708, United States 2
Biosciences Division, Oak Ridge National Laboratory, P.O. Box 2008, MS-6036, Oak Ridge, Tennessee 37831-6036, United States
*Corresponding author: Heileen Hsu-Kim (email:
[email protected]; phone: +1-919-660-5109).
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Abstract Mercury (Hg)-contaminated sediment and water contain various Hg species, with a small
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fraction available for microbial conversion to the bioaccumulative neurotoxin
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monomethylmercury (MeHg). Quantification of this available Hg pool is needed to prioritize
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sites for risk management. This study compared the efficacy of diffusive gradient in thin-film
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(DGT) passive samplers to a thiol-based selective extraction method with glutathione (GSH) and
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conventional filtration (0.2 µm) of nanocrystalline FeS (0.95 µmol 199Hg per g FeS) (Figure S1), and
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196
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Preparation of these stock solutions are described in the Supporting Information (SI).
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Measurements of 198Hg were used to track Hg(II) and MeHg derived from the ambient or
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“native” mercury of the original sediment. Dissolved stock solutions of 202Hg(II) and Me202Hg
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were used as internal standards to correct for method extraction efficiencies for total Hg and
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MeHg analysis. The measured isotopic compositions of all Hg stocks are shown in Table S1.
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Each DGT sampler entailed a 25-mm circular plastic housing (DGT Research Ltd,
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Hg2+, nanoparticulate 200HgS (39 nm average hydrodynamic diameter), 199Hg(II) sorbed to Hg(II) bound to dissolved Suwannee humic acid (5 µmol 196Hg per mg of organic C).
Lancaster, UK) layered with a 0.45 µm nitrocellulose filter, an agarose gel diffusion layer, and a
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resin binding layer of thiolated silica beads supported on a polyacrylamide gel, as described
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previously33 and in the SI.
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Sediment and surface water samples from two different locations were used for laboratory
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microcosms. The first location was a mesohaline tidal salt marsh near the Rhodes River
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(Edgewater, Maryland, USA). Sediment (top 15 cm, 172 ± 7 ng g-1 Hg dry weight basis) and
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surface water were collected on 30th September 2015, were stored at 4 °C, and used for the
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experiment 1 month after collection. Slurries made from these samples were designated the
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‘mesohaline’ microcosm for this study. The second site is a small (approximately 8000 m2)
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retention pond of Sandy Creek (Durham, North Carolina, USA), a freshwater stream located in
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an urbanized watershed and known to receive moderate amounts of legacy Hg, possibly
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originating from historical application of turf grass pesticides in upland areas37. The sediments
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(49.6 ± 6 ng Hg g-1) were collected by hand from the edge of this pond and were used for the
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“freshwater” slurry microcosms within 2 days of collection on 3rd March 2017.
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Slurry Preparation and Experimental Design. We selected batch anaerobic sediment slurries
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(instead of more complex laboratory or field experiments) to test our hypothesis. Reasons for this
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approach were to: 1) Control for a range of Hg bioavailability and methylation potentials; 2)
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Incorporate multiple replicates across multiple time points (from 0.5 to 7 days) and microbial
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communities; and 3) Focus on Hg methylation while minimizing confounding factors associated
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with MeHg degradation.
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Two sediment slurry microcosm experiments (i.e., the mesohaline and freshwater
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microcosms) were performed independently. In both cases, the slurries were prepared by placing
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80 g (wet weight) of sediment and 100 mL of surface water in 0.2 L glass jars. The mesohaline
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slurries (20 individual jars) were amended with 5 µM sodium pyruvate and the freshwater
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slurries (15 jars) were amended with 1 mM sodium pyruvate as a means to stimulate the
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methylating microbial community in these slurries. The slurries were sealed with gas-tight screw
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caps and incubated at room temperature in the dark for 4 days. An additional replicate slurry
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with a resazurin indicator (0.001%) changed from dark pink to colorless over this time,
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indicating the development of anoxic conditions in the jars. After this pre-incubation period, all
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further manipulations of the slurry were performed in an anaerobic chamber with a 3% H2/97%
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N2 atmosphere (Coy Labs).
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Each anaerobic jar was amended with four forms of inorganic Hg (i.e., “endmembers”):
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dissolved 204Hg2+, 196Hg(II)-humic acid, nanoparticulate 200HgS, and 199Hg(II) sorbed to FeS. The
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target Hg content for each endmember was within the range of 50-200 ng g-1 d.w. (The amount
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of each isotope spike was verified later by analysis of total Hg content in the slurry). The jars
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were shaken end-over-end for a few seconds between each Hg addition. After all Hg spike
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addtions, a DGT sampler was added to each jar, and the jars were stored under static conditions
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in the dark (to avoid photodegradation of MeHg). During this incubation, the jars were mixed
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end-over-end once per day. Replicate jars (n = 3 or 4) were sacrificed periodically from < 0.5 d
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and up to 7 d after Hg isotope addition.
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We note that results between the mesohaline and freshwater slurry experiments were not
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designed to be similar to each other (e.g., they utilized sediment-water from different sites and
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were stimulated to different extents with organic carbon). Rather, we view these slurries as
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distinct microbial community mixtures and geochemical compositions for us to test our
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Environmental Science & Technology
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hypothesis regarding the efficacy of DGTs as indicators for Hg bioavailability and methylation
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potential.
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Microcosm Sampling. At the designated incubation time point, each jar was mixed end-over-end
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and held static for 10 minutes to allow for large particles to settle prior to subsampling in the
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anaerobic chamber. Porewater was collected by filtering 15 mL of the overlying water through a
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0.2 µm polyethersulfone filter (VWR). This filtered sample was apportioned into samples
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designated for total Hg analysis, major cations including Fe, sulfate and dissolved organic carbon
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(DOC) analyses.
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The whole slurry was then mixed end-over-end again and aliquots of the unfiltered slurry
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were immediately collected and frozen for later analysis of MeHg, total Hg contents, the GSH-
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extractable Hg fraction, dry-wet mass ratio, microbial community composition, and abundance
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of methylating microorganisms. The pH values of the slurries were measured at the time of
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sampling.
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Chemical Analyses. Procedures for all chemical analyses are described in the SI. Isotopic-
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specific MeHg and Hg contents were quantified by cold vapor inductively coupled plasma mass
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spectrometry (ICP-MS). Other cations were determined by conventional ICP-MS. Dry-wet mass
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ratio was determined by the gravimetric mass of a whole slurry sample before and after heating
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in an oven overnight at 105 oC.
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The glutathione (GSH)-extractable fraction of Hg(II) for each sample was determined as
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described previously.17 In brief, 1 mL of whole slurry (corresponding to 0.13 gdw sediment) was
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added to 10 mL of deionized water spiked with dissolved GSH to a final concentration of 1 mM
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and mixed end-over-end for 1 h. The supernatant was then collected and subjected to
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centrifugation (
