Simple and rapid formic acid sample treatment for the isolation of HgSe nanoparticles from animal tissues.

The present work explores for the first time the potential of formic acid on the extraction of tiemannite (HgSe) nanoparticles from seabird tissues, in particular giant petrels. Mercury (Hg) is considered one of the top ten chemicals of major public health concern. However, the fate and metabolic pathways of Hg in living organisms remain unknown. Methylmercury (MeHg), largely produced by microbial activity in the aquatic ecosystems is biomagnified in the trophic web. HgSe is considered the end-product of MeHg demethylation in biota and an increasing number of studies focuses on the characterization of this solid compound to understand its biomineralization. In this study, a conventional enzymatic treatment is compared with a simpler and environmentally friendly extraction by using formic acid (5 mL of = 50 % formic acid) as exclusive reagent. The analyses by spICP-MS of the resulting extracts from a variety of seabird biological tissues (liver, kidneys, brain, muscle) reveal comparable results by both extraction approaches in terms of nanoparticles stability and extraction efficiency. Therefore, the results included in this work demonstrate the good performance of employing organic acid as simple, cost effective and green procedure to extract HgSe nanoparticles from animal tissues. Moreover, an alternative consisting of a classical enzymatic procedure but with ultrasonic assistance reducing the extraction time from 12 h to 2 min is also described for the first time. The sample processing methodologies developed, combined with spICP-MS, have emerged as powerful tools for the rapid screening and quantification of HgSe nanoparticles in animal tissues. Finally, this combination allowed us to identify the possible occurrence of Cd particles and As particles associated with HgSe NPs in seabirds.

Early-life exposure to methylmercury induces reversible behavioral impairments and gene expression modifications in one isogenic lineage of mangrove rivulus fish Kryptolebias marmoratus.

Methylmercury (MeHg) is a ubiquitous bioaccumulative neurotoxicant present in aquatic ecosystems. It is known to alter behaviors, sensory functions and learning abilities in fish and other vertebrates. Developmental and early-life stages exposure to MeHg can lead to brain damage with immediate consequences on larvae behavior, but may also induce long term effects in adults after a detoxification period. However, very little is known about developmental origin of behavioral impairment in adults due to early exposure to MeHg. The aim of this study is to assess whether early-life MeHg exposure induces immediate and/or delayed effects on behaviors, related genes expression and DNA methylation (one of epigenetic mechanisms). To reach this goal, newly hatched larvae of mangrove rivulus fish, Kryptolebias marmoratus, were exposed to two sub-lethal concentrations of MeHg (90 µg/L and 135 µg/L) for 7 days, and immediate and delayed effects were assessed respectively in 7 dph (days post-hatching) and 90 dph fish. This species naturally produces isogenic lineages due to its self-fertilizing reproduction system, which is unique among vertebrates. It allows to study how environment stressors can influence organism's phenotype while minimizing genetic variability. As results, both MeHg exposures are associated with a decreased foraging efficiency and thigmotaxis, and a dose-dependent reduction in larvae locomotor activity. Regarding molecular analysis in larvae whole bodies, both MeHg exposures induced significant decreased expression of DNMT3a, MAOA, MeCP2 and NIPBL, and significant increase of GSS, but none of those genes underwent methylation changes in targeted CpGs. None of significant behavioral and molecular impairments observed in 7-dph larvae were found in 90-dph adults, which highlight a distinction between immediate and delayed effects of developmental MeHg exposure. Our results suggest implications of aminergic system and its neurotransmitters, redox/methylation trade-off and possibly other epigenetic mechanisms in MeHg neurotoxicity underlying behavioral alterations in rivulus.

Recent and historical pollution legacy in high altitude Lake Marboré (Central Pyrenees): A record of mining and smelting since pre-Roman times in the Iberian Peninsula.

We have analyzed potential harmful trace elements (PHTE; Pb, Hg, Zn, As and Cu) on sediment cores retrieved from lake Marboré (LM) (2612 m a.s.l, 42°41'N; 0° 2'E). PHTE variability allowed us to reconstruct the timing and magnitude of trace metal pollutants fluxes over the last 3000 years in the Central Pyrenees. A statistical treatment of the dataset (PCA) enabled us to discern the depositional processes of PHTE, that reach the lake via direct atmospheric deposition. Indeed, the location of LM above the atmospheric boundary layer makes this lake an exceptional site to record the long-range transport of atmospheric pollutants in the free troposphere. Air masses back-trajectories analyses enabled us to understand the transport pathways of atmospheric pollutants while lead isotopic analyses contributed to evaluate the source areas of metal pollution in SW Europe during the Late Holocene. PHTE variability, shows a clear agreement with the main exploitation phases of metal resources in Southern Europe during the Pre-Industrial Period. We observed an abrupt lead enrichment from 20 to 375 yrs CE mostly associated to silver and lead mining and smelting practices in Southern Iberia during the Roman Empire. This geochemical data suggests that regional atmospheric metal pollution during the Roman times rivalled the Industrial Period. PHTE also increased during the High and Late Middle Ages (10-15th centuries) associated to a reactivation of mining and metallurgy activities in high altitude Pyrenean mining sites during climate amelioration phases. Atmospheric mercury deposition in the Lake Marboré record mostly reflects global emissions, particularly from Almadén mines (central Spain) and slightly fluctuates during the last three millennia with a significant increase during the last five centuries. Our findings reveal a strong mining-related pollution legacy in alpine lakes and watersheds that needs to be considered in management plans for mountain ecosystems as global warming and human pressure effects may contribute to their future degradation.

Diagenetic production, accumulation and sediment-water exchanges of methylmercury in contrasted sediment facies of Lake Titicaca (Bolivia).

Monomethylmercury (MMHg) concentrations in aquatic biota from Lake Titicaca are elevated although the mercury (Hg) contamination level of the lake is low. The contribution of sediments to the lake MMHg pool remained however unclear. In this work, seven cores representative of the contrasted sediments and aquatic ecotopes of Lake Titicaca were sliced and analyzed for Hg and redox-sensitive elements (Mn, Fe, N and S) speciation in pore-water (PW) and sediment to document early diagenetic processes responsible for MMHg production and accumulation in PW during organic matter (OM) oxidation. The highest MMHg concentrations (up to 12.2 ng L-1 and 90% of THg) were found in subsurface PWs of the carbonate-rich sediments which cover 75% of the small basin and 20% of the large one. In other sediment facies, the larger content of OM restricted MMHg production and accumulation in PW by sequestering Hg in the solid phase and potentially also by decreasing its bioavailability in the PW. Diagenetically reduced S and Fe played a dual role either favoring or restricting the availability of Hg for biomethylation. The calculation of theoretical diffusive fluxes suggests that Lake Titicaca bottom sediments are a net source of MMHg, accounting for more than one third of the daily MMHg accumulated in the water column of the Lago Menor. We suggest that in the context of rising anthropogenic pressure, the enhancement of eutrophication in high altitude Altiplano lakes may increase these MMHg effluxes into the water column and favor its accumulation in water and biota.

Multiple regression analysis to assess the spatial distribution and speciation of mercury in surface sediments of a contaminated lagoon.

The concentrations of inorganic mercury (IHg) and methylmercury (MeHg) in surface sediments from the contaminated Aveiro Lagoon (Portugal) were determined by species-specific isotope dilution analysis. Different behaviour of IHg and MeHg was observed based upon multiple regression analysis, including geochemical characteristics of the surface sediments (fine fraction, concentrations of organic matter and metals) and non-Euclidean distances between sampling points. This data treatment method was valid over the entire concentration range of IHg and MeHg, allowing robust quantitative evaluation with respect to extrapolation. For IHg, there was statistical separation of the dispersion away from the contamination source and of Al concentration in the sediments. The MeHg concentrations followed those of IHg at high concentrations. The geochemical variables, such as concentrations of Ca (marine influence proxy), Mn and organic matter, were necessary to describe the behaviour of MeHg across the whole concentration range. The models for MeHg demonstrated that, close to the mouth of the lagoon, net production of MeHg was higher. In future, multiple regression analysis could be applied to separate and to evaluate quantitatively the effects of geochemistry and dispersion away from the contamination source in sediments contaminated with other substances.

Mercury Isotopic Fractionation during Pedogenesis in a Tropical Forest Soil Catena (French Guiana): Deciphering the Impact of Historical Gold Mining.

We used natural mercury (Hg) stable isotopes to investigate the Hg cycle in a rainforest soil catena (French Guiana) partially gold-mined during the early 1950s. Litterfall showed homogeneous Δ199Hg values [-0.18 ± 0.05‰, i.e., a modern gaseous elemental Hg (GEM) isotopic signature]. After litter decomposition, Hg bound to organic matter (OM) is mixed with Hg from pristine (-0.55 ± 0.22‰) or gold-mined (-0.09 ± 0.16‰) mineral materials. Negative Δ199Hg values in deep pristine mineral horizons (-0.60 ± 0.16‰) suggest the transfer of Hg bound to dissolved OM depleted in odd isotopes due to mass-independent fractionation during Hg abiotic reduction. Perennial palm tree leaves collected above gold-mined and pristine soil recorded contrasting Δ199Hg signatures likely resulting from GEM re-emission processes from soils and leaf surfaces. Upslope, soil δ202Hg signatures showed a negative shift (ε ∼ -1‰) with depth attributed to mass-dependent fractionation during Hg sorption and complexation onto iron oxides and dissolved OM. Downslope, higher δ202Hg values in soils resulted from hydromorphy [lower humification, greater Hg(II) reduction, etc.]. The unique Hg isotopic signatures of Amazonian soils probably result in multistep fractionation processes during pedogenesis (millions of years) and in a potentially different Hg isotopic signature of preanthropogenic background GEM.

Development of a large volume injection method using a programmed temperature vaporization injector - gas chromatography hyphenated to ICP-MS for the simultaneous determination of mercury, tin and lead species at ultra-trace levels in natural waters.

The current EU legislation lays down Environmental Quality Standards (EQS) for 45 priority substances in surface waters; among them levels for (organo)metallic species of Hg, Sn and Pb are set between ng L-1 (for Hg and Sn) and µg L-1 (for Pb). To date, only a few analytical methods can reach these very restrictive limits and there is thus a need for comprehensive methods able to analyze these species down to these levels in natural waters. The aim of this work was to develop an online automated pre-concentration method using large volume injections with a Programmed Temperature Vaporization (PTV) injector fitted with a sorbent packed liner coupled to GC-ICP-MS to further improve the detection limits associated to this well-established method. The influence of several parameters such as the PTV transfer temperature and time, carrier gas flow rate and amount of packing material was investigated. Finally, the maximum volume injected through single or multiple injection modes was optimized to obtain the best compromise between chromatographic resolution and sensitivity. After optimization, very satisfactory results in terms of absolute and methodological detection limits were achieved, down to the pg L-1 level for all species studied. The potential of the method was exemplified by determining the concentrations of organometallic compounds in unpolluted river waters samples from the Adour river basin (SW France) and results were compared with conventional (splitless) GC-ICP-MS. The strength of this analytical method lies in the low detection limits reached for the simultaneous analysis of a wide group of organometallic compounds, and the potential to transfer this method to other gas chromatographic applications with inherent lower sensitivity.

Mercury contamination level and speciation inventory in Lakes Titicaca & Uru-Uru (Bolivia): Current status and future trends.

Aquatic ecosystems of the Bolivian Altiplano (∼3800 m a.s.l.) are characterized by extreme hydro-climatic constrains (e.g., high UV-radiations and low oxygen) and are under the pressure of increasing anthropogenic activities, unregulated mining, agricultural and urban development. We report here a complete inventory of mercury (Hg) levels and speciation in the water column, atmosphere, sediment and key sentinel organisms (i.e., plankton, fish and birds) of two endorheic Lakes of the same watershed differing with respect to their size, eutrophication and contamination levels. Total Hg (THg) and monomethylmercury (MMHg) concentrations in filtered water and sediment of Lake Titicaca are in the lowest range of reported levels in other large lakes worldwide. Downstream, Hg levels are 3-10 times higher in the shallow eutrophic Lake Uru-Uru than in Lake Titicaca due to high Hg inputs from the surrounding mining region. High percentages of MMHg were found in the filtered and unfiltered water rising up from <1 to ∼50% THg from the oligo/hetero-trophic Lake Titicaca to the eutrophic Lake Uru-Uru. Such high %MMHg is explained by a high in situ MMHg production in relation to the sulfate rich substrate, the low oxygen levels of the water column, and the stabilization of MMHg due to abundant ligands present in these alkaline waters. Differences in MMHg concentrations in water and sediments compartments between Lake Titicaca and Uru-Uru were found to mirror the offset in MMHg levels that also exist in their respective food webs. This suggests that in situ MMHg baseline production is likely the main factor controlling MMHg levels in fish species consumed by the local population. Finally, the increase of anthropogenic pressure in Lake Titicaca may probably enhance eutrophication processes which favor MMHg production and thus accumulation in water and biota.

Occurrence and Fate of Organic and Organometallic Pollutants in Municipal Wastewater Treatment Plants and Their Impact on Receiving Waters (Adour Estuary, France).

To achieve a "Good Environmental Status by 2015," as demanded by the water framework directive, monitoring programs are needed to furnish data on target compounds. In this study, a first evaluation of influents and main emissions of 3 local wastewater treatment plants (WWTP) in the Adour estuary (southwest of France) was performed for 23 pollutants (10 musk fragrances, 5 alkylphenols, and 8 organometallics), as well as receiving estuarine water from the same area. High frequency of occurrence of these compounds was found in influents samples (musks: 22-100%; alkylphenols 11-100%; organometallics 0-100%) and effluents (musks: 0-100%; alkylphenols 0-100%; organometallics 0-100%). The removal efficiencies were calculated and varied from negative values up to 98% with the lowest values for synthetic musk compounds. Temporal variability of the target compounds also was studied, and a few tendencies were observed. Estimation of the daily output of each WWTP into the estuary also showed that galaxolide, nonylphenol, monobuthyltin, and inorganic mercury were the compounds discharged into the environment at the highest concentrations. Finally, the occurrence of these compounds in estuarine waters was evaluated; most of them were present at concentrations below the limits of quantification (musks: 0.53-41.5 ng/L; alkylphenols 3.4-410 ng/L; organometallics 0.02-0.70 ng/L) suggesting a low impact in the resulting receiving waters.

Synergistic effects of mining and urban effluents on the level and distribution of methylmercury in a shallow aquatic ecosystem of the Bolivian Altiplano.

Lake Uru Uru (3686 m a.s.l.) located in the Bolivian Altiplano region receives both mining effluents and urban wastewater discharges originating from the surrounding local cities which are under rapid development. We followed the spatiotemporal distribution of different mercury (Hg) compounds and other metal(oid)s (e.g., Fe, Mn, Sb, Ti and W) in both water and sediments during the wet and dry seasons along a north-south transect of this shallow lake system. Along the transect, the highest Hg and metal(oid) concentrations in both water and sediments were found downstream of the confluences with mining effluents. Although a dilution effect was found for major elements during the wet season, mean Hg and metal(oid) concentrations did not significantly differ from the dry season due to the increase in acid mine drainage (AMD) inputs into the lake from upstream mining areas. In particular, high filtered (<0.45 µm) mono-methylmercury (MMHg) concentrations (0.69 ± 0.47 ng L-1) were measured in surface water representing 49 ± 11% of the total filtered Hg concentrations (THgF) for both seasons. Enhanced MMHg lability in relation with the water alkalinity, coupled with abundant organic ligands and colloids (especially for downstream mining effluents), are likely factors favoring Hg methylation and MMHg preservation while inhibiting MMHg photodegradation. Lake sediments were identified as the major source of MMHg for the shallow water column. During the dry season, diffusive fluxes were estimated to be 227 ng m-2 d-1 for MMHg. This contribution was found to be negligible during the wet season due to a probable shift of the redox front downwards in the sediments. During the wet season, the results obtained suggest that various sources such as mining effluents and benthic or macrophytic biofilms significantly contribute to MMHg inputs in the water column. This work demonstrates the seasonally dependent synergistic effect of AMD and urban effluents on the shallow, productive and evaporative high altitude lake ecosystems which promotes the formation of natural organometallic toxins such as MMHg in the water column.

Multiple regression analysis to assess the role of plankton on the distribution and speciation of mercury in water of a contaminated lagoon.

Spatial and seasonal variation of mercury species aqueous concentrations and distributions was carried out during six sampling campaigns at four locations within Laranjo Bay, the most mercury-contaminated area of the Aveiro Lagoon (Portugal). Inorganic mercury (IHg(II)) and methylmercury (MeHg) were determined in filter-retained (IHgPART, MeHgPART) and filtered (<0.45µm) fractions (IHg(II)DISS, MeHgDISS). The concentrations of IHgPART depended on site and on dilution with downstream particles. Similar processes were evidenced for MeHgPART, however, its concentrations increased for particles rich in phaeophytin (Pha). The concentrations of MeHgDISS, and especially those of IHg(II)DISS, increased with Pha concentrations in the water. Multiple regression models are able to depict MeHgPART, IHg(II)DISS and MeHgDISS concentrations with salinity and Pha concentrations exhibiting additive statistical effects and allowing separation of possible addition and removal processes. A link between phytoplankton/algae and consumers' grazing pressure in the contaminated area can be involved to increase concentrations of IHg(II)DISS and MeHgPART. These processes could lead to suspended particles enriched with MeHg and to the enhancement of IHg(II) and MeHg availability in surface waters and higher transfer to the food web.

Diurnal variability and biogeochemical reactivity of mercury species in an extreme high-altitude lake ecosystem of the Bolivian Altiplano.

Methylation and demethylation represent major transformation pathways regulating the net production of methylmercury (MMHg). Very few studies have documented Hg reactivity and transformation in extreme high-altitude lake ecosystems. Mercury (Hg) species concentrations (IHg, MMHg, Hg°, and DMHg) and in situ Hg methylation (M) and MMHg demethylation (D) potentials were determined in water, sediment, floating organic aggregates, and periphyton compartments of a shallow productive Lake of the Bolivian Altiplano (Uru Uru Lake, 3686 m). Samples were collected during late dry season (October 2010) and late wet season (May 2011) at a north (NS) and a south (SS) site of the lake, respectively. Mercury species concentrations exhibited significant diurnal variability as influenced by the strong diurnal biogeochemical gradients. Particularly high methylated mercury concentrations (0.2 to 4.5 ng L(-1) for MMHgT) were determined in the water column evidencing important Hg methylation in this ecosystem. Methylation and D potentials range were, respectively, <0.1-16.5 and <0.2-68.3 % day(-1) and were highly variable among compartments of the lake, but always higher during the dry season. Net Hg M indicates that the influence of urban and mining effluent (NS) promotes MMHg production in both water (up to 0.45 ng MMHg L(-1) day(-1)) and sediment compartments (2.0 to 19.7 ng MMHg g(-1) day(-1)). While the sediment compartment appears to represent a major source of MMHg in this shallow ecosystem, floating organic aggregates (dry season, SS) and Totora's periphyton (wet season, NS) were found to act as a significant source (5.8 ng MMHg g(-1) day(-1)) and a sink (-2.1 ng MMHg g(-1) day(-1)) of MMHg, respectively. This work demonstrates that high-altitude productive lake ecosystems can promote MMHg formation in various compartments supporting recent observations of high Hg contents in fish and water birds.

Mercury speciation and dispersion from an active gold mine at the West Wits area, South Africa.

Total mercury (HgTOT), inorganic mercury (IHg), and methylmercury (MHg) were determined in dry season waters, sediments, and tailings from an active mine which has long history of gold exploitation. Although HgTOT in waters was generally low (0.03 to 19.60 ng L(-1)), the majority of the samples had proportions of MHg of at least 90 % of HgTOT which denotes a substantial methylation potential of the mine watersheds. Mercury was relatively high in tailing materials (up to 867 µg kg(-1)) and also in the mine sediments (up to 837 µg kg(-1)) especially in samples collected near tailing storage facilities and within a receiving water dam. Sediment profiles revealed mercury enrichment and enhanced methylation rate at deeper layers. The presence of IHg and decaying plants (organic matter) in the watersheds as well as the anoxic conditions of bulk sediments are believed to be some of the key factors favoring the mercury methylation at the site.

In-port derivatization coupled to different extraction techniques for the determination of alkylphenols in environmental water samples.

Large volume injection (LVI)-in port silylation coupled to gas chromatography-mass spectrometry (GC-MS) for the determination of alkylphenols (APs) in water samples applying four different extraction approaches was evaluated. Among the variables studied for in-port derivatization, vent time, cryo-focusing temperature and the ratio solvent volume/N,O-bis(trimethylsilyl)trifluoroacetamide (BSTFA) volume were optimized using an experimental design approach. Regarding the extraction techniques, different approaches previously optimized in the research group were tested. On the one hand different polymeric materials were tested: silicon rod (SR), polyethersulfone (PES) and polydimethylsiloxane (PDMS), the latter in the stir-bar sorptive extraction format (SBSE-PDMS). PES was chosen among the polymeric materials due to the higher recoveries (compared with SR) and lower price (compared to PDMS in the stir-bar sorptive extraction, SBSE-PDMS). Both MASE and PES protocols were selected at this point for further method validation and application to real samples. Finally, the developed methods were validated and applied to the determination of target analytes in various aqueous environmental matrices, including estuarine water and wastewater. Acceptable repeatability in the case of MASE (5-17%) and PES (7-21%) procedures and method detection limits (MDLs, 5-123 and 28-328 ng L(-1) for PES and MASE, respectively) were obtained for most analytes. In terms of apparent recoveries in the presence of matrix, estuarine and effluent samples showed no significant matrix effect (apparent recoveries in the 73-121% for PES and 74-128% for MASE), while a stronger matrix effect was observed for influent wastewater samples (98-132% for PES and 65-156% for MASE). Both MASE and PES extractions combined with LVI-in-port derivatization-GC-MS were applied to the determination of APs in the estuary of Bilbao (Gulf of Biscay, Spain).

Comparison between GC-MS and GC-ICPMS using isotope dilution for the simultaneous monitoring of inorganic and methyl mercury, butyl and phenyl tin compounds in biological tissues.

The aim of this work is to compare simultaneous isotope dilution analysis of organotin and organomercury compounds by gas chromatography-mass spectrometry (GC-MS) and gas chromatography-inductively coupled plasma mass spectrometry (GC-ICP/MS) on certified bivalve samples. These samples were extracted by microwave with tetramethylammonium hydroxide (TMAH). Derivatization with both NaBEt4 and NaBPr4 was evaluated, and analytical performances were compared. Two CRM materials, BCR-710 and CRM-477, were analyzed by both techniques to verify accuracy. A mixed spike containing (201)Hg-enriched methylmercury (MeHg), (199)Hg-enriched inorganic mercury (iHg), (119)Sn-enriched monobutyltin (MBT), dibutyltin (DBT), and tributyltin (TBT) as well as homemade (116)Sn-enriched monophenyltin (MPT), diphenyltin (DPT), and triphenyltin (TPT) was used for the isotope dilution analysis of samples. The two techniques studied were compared in terms of classic analytical parameters: linearity, precision or repeatability (i.e., percent relative standard deviation, RSD%), limit of detection (LOD), and limit of quantification (LOQ), showing excellent linearity, precision below 12% for all analytes, and LOQs of 0.06-1.45 pg for GC-MS and 0.02-0.27 pg for GC-ICP/MS.

Investigations into the differential reactivity of endogenous and exogenous mercury species in coastal sediments.

Stable isotopic tracer methodologies now allow the evaluation of the reactivity of the endogenous (ambient) and exogenous (added) Hg to further predict the potential effect of Hg inputs in ecosystems. The differential reactivity of endogenous and exogenous Hg was compared in superficial sediments collected in a coastal lagoon (Arcachon Bay) and in an estuary (Adour River) from the Bay of Biscay (SW France). All Hg species (gaseous, aqueous, and solid fraction) and ancillary data were measured during time course slurry experiments under variable redox conditions. The average endogenous methylation yield was higher in the estuarine (1.2 %) than in the lagoonal sediment (0.5 %), although both methylation and demethylation rates were higher in the lagoonal sediment in relation with a higher sulfate-reducing activity. Demethylation was overall more consistent than methylation in both sediments. The endogenous and exogenous Hg behaviors were always correlated but the exogenous inorganic Hg (IHg) partitioning into water was 2.0-4.3 times higher than the endogenous one. Its methylation was just slightly higher (1.4) in the estuarine sediment while the difference in the lagoonal sediment was much larger (3.6). The relative endogenous and exogenous methylation yields were not correlated to IHg partitioning, demonstrating that the bioavailable species distributions were different for the two IHg pools. In both sediments, the exogenous IHg partitioning equaled the endogenous one within a week, while its higher methylation lasted for months. Such results provide an original assessment approach to compare coastal sediment response to Hg inputs.

Identification of mercury and other metals complexes with metallothioneins in dolphin liver by hydrophilic interaction liquid chromatography with the parallel detection by ICP MS and electrospray hybrid linear/orbital trap MS/MS.

A novel analytical procedure for the identification of metal (Hg, Cd, Cu, Zn) complexes with individual metallothionein (MT) isoforms in biological tissues by electrospray MS/MS was developed. The sample preparation was reduced to three rapid steps: the two-fold dilution of the sample cytosol with acetonitrile, the recovery of the supernatant containing MT-complexes by centrifugation and its concentration under nitrogen flow. The replacement of reversed phase HPLC by hydrophilic interaction LC (HILIC) allowed the preservation of the unstable and low abundant metallothionein zinc-mercury mixed complexes (MT-Zn(6)Hg). The MT complexes eluted were detected by ICP MS and identified in terms of molecular mass by electrospray high resolution (100,000) MS. The identification was completed by on line demetallation and the determination of the molecular mass of the apoform, followed by amino acid sequencing in the top-down mode using high energy collision fragmentation (HCD). The method was applied to the identification of MT complexes in a white-sided dolphin (Lagenorhynchus acutus) liver homogenate. The Zn complex of the N-acetylated MT2 isoform was found to be predominant, the presence of mixed complexes with Cd, Cu and, for the first time ever, Hg, was demonstrated. The latter finding has the potential to shed new light on the mercury detoxification mechanism in marine organisms.

Phytoplankton distribution and productivity in a highly turbid, tropical coastal system (Bach Dang Estuary, Vietnam).

Phytoplankton diversity, primary and bacterial production, nutrients and metallic contaminants were measured during the wet season (July) and dry season (March) in the Bach Dang Estuary, a sub-estuary of the Red River system, Northern Vietnam. Using canonical correspondence analysis we show that phytoplankton community structure is potentially influenced by both organometallic species (Hg and Sn) and inorganic metal (Hg) concentrations. During March, dissolved methylmercury and inorganic mercury were important factors for determining phytoplankton community composition at most of the stations. In contrast, during July, low salinity phytoplankton community composition was associated with particulate methylmercury concentrations, whereas phytoplankton community composition in the higher salinity stations was more related to dissolved inorganic mercury and dissolved mono and tributyltin concentrations. These results highlight the importance of taking into account factors other than light and nutrients, such as eco-toxic heavy metals, in understanding phytoplankton diversity and activity in estuarine ecosystems.

An experimental approach to investigate mercury species transformations under redox oscillations in coastal sediments.

This work describes a laboratory experiment designed to unravel mercury species reactivity in superficial coastal sediments oscillating between oxic and anoxic conditions. The experimental set-up has been applied to a sediment slurry from the Arcachon Bay (France) to follow the evolution of both naturally occurring (i.e. endogenous) and isotopically enriched added mercury species (i.e. exogenous, ¹99IHg and ²°¹MMHg) at environmental levels. The transformation and partition between the different phases (aqueous, solid and gaseous) of the endogenous and exogenous mercury species (inorganic Hg (IHg), monomethyl Hg (MMHg), elemental Hg (Hg°) and dimethyl Hg (DMHg)) have been investigated by isotopic speciation methods throughout the experiment. The results demonstrate that the experimental approach is able to promote sediment redox oscillations and to simultaneously follow the biogeochemical fate of naturally occurring or added mercury species. Experimentally driven redox transition events were found to significantly enhance the aqueous Hg species concentrations, while the MMHg burden is not greatly affected. Indeed, during the anoxic-oxic transition, while aqueous endogenous IHg and MMHg exhibited a two-fold increase, aqueous exogenous IHg and MMHg increased 7 and 4 times, respectively. Transient increases of the net IHg methylation were recorded during the redox transitions with the largest increase of the MMHg contents (factor 1.8) observed during the oxic-anoxic transition. High resolution in situ redox experiments have not been performed up to now, therefore the developed experimental set-up provides novel insights in both the influence of redox conditions on Hg methylation/demethylation and adsorption/desorption processes and kinetics in superficial sediments.

Mercury methylation by a microbial community from sediments of the Adour Estuary (Bay of Biscay, France).

In order to study the influence of microorganisms on the mercury biogeochemistry, the metal content and the structure of microbial communities were determined in sediments from stations along the Adour Estuary. The comparison of the bacterial communities and their distribution in function of the environmental parameters by Canonical Correspondence Analysis (CCA) revealed the influence of metals on the bacterial communities structure. Sediments where the bacterial communities are mostly influenced by methylmercury were incubated in slurries with or without mercury, under oxic and anoxic conditions. Methylmercury production was detected in the anoxic biotic slurries with a net methylation yield of 0.3% after 24 h. CCA based on T-RFLP profiles revealed the impact of mercury addition on the bacterial communities structure. In addition, 17 bacterial strains, mainly sulphate-reducing bacteria involved in mercury methylation, were isolated and identified.