Non-ferrous smelter emissions have prompted revegetation efforts near the border towns of Flin Flon, Manitoba and Creighton, Saskatchewan to facilitate regrowth of the surrounding boreal forest. Previously, several soil amendments were tested for plant response in soils from the smelter-impacted area and one amendment, a pyrolyzed meat and bone meal (MBM) biochar, was of particular interest because of its potential to immobilize Zn. Hydroxyapatite (HAP), with a small degree of carbonate substitution, was identified as the major component of the MBM biochar. The solubility of this Ca-mineral was pH dependent, with dissolution occurring at pH 6.3; consequently, adsorption experiments were performed at a slightly lower pH (6.1 ± 0.1). Zinc adsorption kinetics were bi-phasic and could be modeled using the Elovich equation, suggesting a diffusion limited reaction likely related to material aggregation. Adsorption also was modeled using the Langmuir equation, which indicated a moderate affinity of the biochar for Zn and an adsorption maximum of 0.650 mmol Zn g-1. Synchrotron-based Zn K-edge x-ray absorption spectroscopy (XAS) indicated an inner-sphere monodentate, tetrahedral bonding of Zn to phosphate groups in the MBM biochar. This was consistent with Zn adsorbed to a HAP standard, indicating that the same sorption mechanism was involved. The ability of MBM biochar to affect Zn speciation in soils was investigated using four soils from four locations in the smelter-impacted region around Flin Flon. A 1:10 (w/w) mixture of the MBM and soil was suspended in 200-mL deionized water (pH 6.1 ± 0.1) and equilibrated for 30 d. Whereas all the soils showed a decrease in extractable Zn following equilibration, only one exhibited a change in Zn speciation—with ca. 25% of the Zn adsorbed onto the MBM biochar. Ore-derived minerals were present in all soils and strong backscattering made identification of minor Zn species difficult. However, using microprobe-based x-ray absorption near edge structure (XANES) spectroscopy, several minor Zn species were identified; including hopeite, a ZnPO4 mineral. The presence of both hopeite and adsorbed Zn are indicative of a direct Zn-phosphate reaction. These results indicate that, under certain condition, MBM biochar can be an effective soil amendment.

The chemical speciation of trace metals controls their potential bioavailability and therefore toxicity to exposed organisms. Despite previous studies demonstrating the ameliorative effects of dissolved organic carbon (DOC) on metal toxicity, the effectiveness of ligands from varying sources and of potentially variable composition in controlling speciation has not been studied in detail in estuarine waters. In addition, the effect of DOC on radionuclide contaminants in combination with trace metals has not been investigated in any waters. This is of particular interest in the estuarine environment, where both anthropogenic and natural ligands, and contaminants that pose a potential threat to ecosystem health, can be present. Competitive ligand exchange adsorptive cathodic stripping voltammetry (CLE-AdCSV) with complexation capacity titrations was employed to determine the speciation of dissolved Cu and Zn, two metals that possess revised environmental quality standards (EQS) which now account for potential metal bioavailability. Dissolved metal concentrations in the < 0.4 and < 0.2 μm filter fractions of samples from the Tamar Estuary were determined during seasonal transects made over a calendar year. Samples were taken over a full salinity range (0-35) and from locations thought to contain DOC from a variety of sources (e.g. terrigenous, biogenic, sewage). No seasonal trends in metal speciation were identified, but a semi-quantitative assessment of DOC type using 3-D fluorimetry showed domination of humic and fulvic type ligands in the upper estuary, and biogenic-type ligands in the lower estuary, the former appearing the most important in controlling Cu and Zn complexation. Filter size fraction differences showed a major portion of the dissolved metal is associated with the 0.2 ≥ 0.4 μm fraction, indicating an importance of larger molecule ligands in controlling potentially bioavailable metal. Sample ligand concentrations ([L_x]) ranged from 1-372 nM (Cu) and 3-412 nM (Zn), and metal-ligand conditional stability constants (log K_(ML_x )) from 10.5-13.5 (Cu) and 7.5-10 (Zn), which are similar to reported literature. Calculated free metal ion concentrations ([M2+]) of 0.3 – 109 nM (Zn) and 1.4 x 10-13 – 7.3 x 10-11 M (Cu) compared well (92% showed no significant differences (P = 0.02)) with direct measurements of [Zn2+] made for the first time in estuarine waters using “Absence of Gradients and Nernst Equilibrium Stripping” (AGNES) after optimisation for estuarine waters. AGNES fully complements CLE-AdCSV in terms of analytical capability and shows that methods are now available that are capable of directly determining [Zn2+] in estuarine waters for use in environmental monitoring studies. Calculations made using the chemical equilibrium speciation programme Visual MINTEQ (VM) showed [Cu2+] and [Zn2+] could be predicted to within one order of magnitude of measured values when log K_(ML_x ) and [L_x] are determined and input into the model. This was in contrast to poor agreement between measured and predicted [M2+] when VM was used with the NICA-Donnan complexing model, which assumes a set portion of the total DOC concentration input is fulvic acid that actively complexes metals. These results corroborate a lack of identification of a relationship between metal speciation in the Tamar samples and DOC concentration, highlighting that knowledge of DOC type, log K_(ML_x )and L_x are important when assessing environmental risk, setting EQSs and for accurate modeling of [Cu2+]. Finally, a combined chemical and biological study investigating the effects of mixtures of DOC, Zn and the radionuclide tritium (3H) on the marine mussel presents the first evidence of a protective effect of Zn on DNA damage caused by 3H. The association of 3H with DOC remains elusive and an assessment of DOC type is recommended for future research, but the study emphasises the importance of investigating mixture effects in order to avoid inaccurate risk assessment and potentially costly site remediation.

abstract: The discovery and development of novel antibacterial agents is essential to address the rising health concern over antibiotic resistant bacteria. This research investigated the antibacterial activity of a natural clay deposit near Crater Lake, Oregon, that is effective at killing antibiotic resistant human pathogens. The primary rock types in the deposit are andesitic pyroclastic materials, which have been hydrothermally altered into argillic clay zones. High-sulfidation (acidic) alteration produced clay zones with elevated pyrite (18%), illite-smectite (I-S) (70% illite), elemental sulfur, kaolinite and carbonates. Low-sulfidation alteration at neutral pH generated clay zones with lower pyrite concentrations pyrite (4-6%), the mixed-layered I-S clay rectorite (R1, I-S) and quartz. Antibacterial susceptibility testing reveals that hydrated clays containing pyrite and I-S are effective at killing (100%) of the model pathogens tested (E. coli and S. epidermidis) when pH (< 4.2) and Eh (> 450 mV) promote pyrite oxidation and mineral dissolution, releasing > 1 mM concentrations of Fe2+, Fe3+ and Al3+. However, certain oxidized clay zones containing no pyrite still inhibited bacterial growth. These clays buffered solutions to low pH (< 4.7) and oxidizing Eh (> 400 mV) conditions, releasing lower amounts (< 1 mM) of Fe and Al. The presence of carbonate in the clays eliminated antibacterial activity due to increases in pH, which lower pyrite oxidation and mineral dissolution rates. The antibacterial mechanism of these natural clays was explored using metal toxicity and genetic assays, along with advanced bioimaging techniques. Antibacterial clays provide a continuous reservoir of Fe2+, Fe3+ and Al3+ that synergistically attack pathogens while generating hydrogen peroxide (H2O¬2). Results show that dissolved Fe2+ and Al3+ are adsorbed to bacterial envelopes, causing protein misfolding and oxidation in the outer membrane. Only Fe2+ is taken up by the cells, generating oxidative stress that damages DNA and proteins. Excess Fe2+ oxidizes inside the cell and precipitates Fe3+-oxides, marking the sites of hydroxyl radical (•OH) generation. Recognition of this novel geochemical antibacterial process should inform designs of new mineral based antibacterial agents and could provide a new economic industry for such clays. / Dissertation/Thesis / Doctoral Dissertation Geological Sciences 2015

abstract: Due to analytical limitations, thermodynamic modeling is a lucrative alternative for obtaining metal speciation in chemically complex systems like life. However, such modeling is limited by the lack of equilibrium constant data for metal-complexation reactions, particularly for metal-organic species. These problems were ameliorated estimating these properties from 0-125°C for ~18,000 metal complexes of small molecules, proteins and peptides. The estimates of metal-ligand equilibrium constants at 25°C and 1 bar were made using multiple linear free energy relationships in accordance with the metal-coordinating properties of ligands such as denticity, identity of electron donor group, inductive effects and steric hindrance. Analogous relationships were made to estimated metal-ligand complexation entropy that facilitated calculation of equilibrium constants up to 125°C using the van’t Hoff equation. These estimates were made for over 250 ligands that include carboxylic acids, phenols, inorganic acids, amino acids, peptides and proteins. The stability constants mentioned above were used to obtain metal speciation in several microbial growth media including past bioavailability studies and compositions listed on the DSMZ website. Speciation calculations were also carried out for several metals in blood plasma and cerebrospinal fluid that include metals present at over micromolar abundance (sodium, potassium, calcium, magnesium, iron, copper and zinc) and metals of therapeutic or toxic potential (like gallium, rhodium and bismuth). Metal speciation was found to be considerably dependent on pH and chelator concentration that can help in the selection of appropriate ligands for gallium & rhodium based anticancer drugs and zinc-based antidiabetics. It was found that methanobactin can considerably alter copper speciation and is therefore a suitable agent for the treatment of Wilson Disease. Additionally, bismuth neurotoxicity was attributed to the low transferrin concentration in cerebrospinal fluid and the predominance of aqueous bismuth trihydroxide. These results demonstrate that metal speciation calculations using thermodynamic modeling can be extremely useful for understanding metal bioavailability in microbes and human bodily fluids. / Dissertation/Thesis / Doctoral Dissertation Biochemistry 2019

The carbon dioxide (CO2) system is the primary buffer in seawater which controls oceanic pH. Changes in the marine CO2 system affect a number of processes such as metal speciation, mineral saturation states, auditory responses in fish, and primary productivity rates. Increased atmospheric concentrations of CO2 from human activities (e.g. burning of fossil fuels, deforestation, and cement production) has led to a global decrease in surface ocean pH termed anthropogenic ocean acidification. One particular concern in response to increased oceanic CO2 is a substantial decrease in the calcium carbonate (CaCO3) saturation states, ΩCaCO3. The long-term physiological effects of ocean acidification and decreased ΩCaCO3 on marine biota are currently subjects of intensive global investigation. Consequently improved methods are needed to facilitate evaluations of the evolving CO2 system chemistry and the responses of marine organisms to those changes. Currently two of four measureable chemical parameters (pH, dissolved inorganic carbon, fugacity of CO2, and total alkalinity) are required for full characterization of the inorganic CO2 system; carbonate ion concentrations ([CO3 2-]) can, for example, be calculated from paired measurements of pH-DIC and pH-TA. The primary objective of this dissertation is to refine a method for directly determining [CO3 2-] using a single measurement, the distinctive ultraviolet absorbance spectra of Pb(II) species in seawater. The technique is fast, methodologically simple, and suitable for routine use in laboratory and shipboard studies. It is, as well, suitable for analyses using autonomous instrumentation. My studies began with an investigation of lead carbonate (PbCO3 0) complexation in synthetic media (at 25 °C between 0.001 to 5.0 molal ionic strength) to evaluate factors that control Pb(II) speciation, and thereby Pb(II) spectra, in seawater. Since laboratory investigations of Pb(II) speciation in seawater require potentiometric measurements of seawater pH, my dissertation includes development of a novel spectrophotometric method for calibrating pH electrodes directly in seawater. This spectrophotometric electrode calibration enables improved assessment of the extent to which electrode behavior is Nernstian and the influence of salinity on electrode calibrations. In addition, for the first time at sea, [CO3 2-] was directly determined in the Arctic and the Eastern Pacific Oceans using the Pb(II) method. These field studies allowed assessment of the consistency between direct [CO3 2-] determinations and carbonate determined using conventional CO2 system measurements. Finally, using techniques from my evaluation of lead speciation, as well as my electrode calibration development and field studies, additional laboratory studies were used to increase carbonate measurement sensitivity and applicability over a wider range of salinity.

The preservation of freshwater aquifers is one important task of managing natural resources today. Intrusion of saltwater from the sea into the groundwater might occur in coastal-near areas and can affect the freshwater quality negatively. One such risk area is the Adriatic coast line of Albania, a country predominantly rich in freshwater resources due to the numerous rivers flowing from the mountains in the east towards the sea. The coastal areas are mainly used for agricultural activities were groundwater is an important resource for irrigation purposes. This study was therefore partly conducted in Albania where samples of groundwater and river waters were collected from the river deltas of Drin and Vjosa in order to investigate the origin of the salt and to assess the bioavailable metal concentrations of nickel and zinc. Both on-site tests and off-site tests, in laboratories, were conducted. In the laboratory, the samples were analysed for oxygen and deuterium isotope data. The results were used to assess the origin of the oxygen content as the method of investigating the origin of the salt uses the oxygen isotope as a tracer. Anion and cation concentrations were analysed as well as the concentrations of metals. These were used as input data to Visual Minteq, a model used to calculate the metal speciation of waters, and a Biotic Ligand Model used to calculate the bioavailable concentrations and the HC5 concentrations. The isotope analysis yielded results similar to previous studies conducted in the nearby area indicating that the salt does not have oceanic origin but meteorological. The bioavailable concentrations were in most sample sites lower than the calculated HC5 concentrations indicating that there is a sufficient buffering system present even though there are several mining activities upstream of the sample sites that could have affected the water quality. This study concludes that the origin of salt is of meteoric origin and not from seawater intrusion. The bioavailable concentrations do not pose a threat as the concentrations are under the HC5 concentrations for all sites but one. In order to deepen the understanding and knowledge of the water qualities in these areas, future studies should focus on a continuous measuring period lasting for example over the whole year minimizing any seasonal variations of the sampled data. Other methods such as geophysical measures could also contribute to a wider analysis of the groundwater condition. / Grundvatten är en av de viktigaste naturresurserna vilket innebär att för att säkerställa en långvarig försörjning krävs en god förvaltning av befintliga akvifärer. I kustnära områden kan intrång av saltvatten från havet till grundvattnet påverka sötvattenkvaliteten negativt. Ett sådant riskområde är den Adriatiska kustlinjen i Albanien, ett land som är övervägande rikt på sötvattenresurser tack vare de många floder som rinner från bergen i öst mot havet i väst. Kustområdena används huvudsakligen för jordbruksverksamhet, där grundvatten är en viktig resurs vid bevattning. Denna studie genomfördes därför delvis i Albanien där prov av grundvatten och flodvatten hämtades från Drins och Vjosas floddeltan för att undersöka saltets ursprung och för att bedöma de biotillgängliga metalkoncentrationerna av nickel och zink. Vattenprover togs på ett urval av platser och testades direkt för omgivningsberoende parametrar samt analyserades vidare i laboratorier. I laboratoriet analyserades proverna med avseende på fördelning av syre- och deuteriumisotoper. Resultaten användes för att bedöma syreinnehållets ursprung, eftersom metoden för att undersöka saltets ursprung använder syreisotopen som spårämne. Anjon- och katjonkoncentrationer analyserades såväl som koncentrationerna av metaller. Dessa användes som inmatningsdata till Visual Minteq, en modell som används för att beräkna metallsammansättningen i vatten och en Biotic ligand model som användes för att beräkna de biotillgängliga koncentrationerna samt HC5-koncentrationerna. Isotopanalysen gav resultat som liknar tidigare studier utförda i det närliggande området vilket indikerar att saltet inte har oceaniskt ursprung utan meteorologiskt. De biotillgängliga koncentrationerna av nickel och zink var i de flesta fall lägre än de beräknade HC5-koncentrationerna, vilket indikerar att det finns ett buffertsystem, trots att gruvaktiviteter uppströms provplatserna skulle kunna ha påverkat vattenkvaliteten negativt.   Studien slutsats är att saltet i grundvattnet och flodvattnet är av meteoriskt ursprung och inte ett resultat av saltvatteninträngning från det Adriatiska havet. De biotillgängliga halterna av nickel och zink understiger HC5 halterna i samtliga fall utom ett.   För att fördjupa förståelsen och kunskapen om vattenkvaliteten i dessa områden bör framtida studier fokusera på en kontinuerlig mätperiod som till exempel sträcker sig över hela året vilket skulle minimera eventuella säsongsvariationer av insamlad data. Andra metoder så som geofysiska mätningar kan också bidra till en bredare analys av tillståndet i grundvattnet.

The objective of this research was to investigate lead speciation in the soil/sediment-water environment and to better understand how the species affect lead mobility under different environmental conditions. The research involved both field soil and sediment samples as well as standard lead solutions. Field samples were fully characterized and extracted by aqueous and organic solvents. The results were compared and evaluated with the metal speciation model, MINTEQA2. Hydrolytic polymerization and organic complexation studies were conducted with standard lead solutions under controlled experimental conditions. Results of the field samples showed that pH, dissolved cations, ionic strength, dissolved organic matter, and nature of the soil/sediment matrix play major roles in the distribution and mobility of lead (Pb) from contaminated sites. In the aqueous equilibration experiment, the magnitude of Pb2+ solubilization was in the order of pH4>pH7>pH9. The results were in good agreement with MINTEQA2 predictions. An important finding of the research is the detection of Pb polymerization species under controlled experimental conditions. At pH 5.22, Pb polymeric species were formed at rate of 0.03 per day. The role of Pb complexation with organic matter was evaluated in both field and standard samples. Different methodologies showed three types of organically bound Pb. A very small fraction of Pb, in the ppb range, was extractable from the contaminated soil by polar organic solvents. Sequential extractions show that 16.6±1.4 % of the Pb is organically complexed. Complexation of Pb with fulvic acid provided new information on the extent of Pb association with soluble organic matter. The overall results of this research have provided new and useful information regarding Pb speciation in environmental samples. The results, in several instances, have provided verification of MINTEQA2 model's prediction. They also revealed areas of disagreement between the models prediction and the experimental results. A positive note regarding the experimental work done in the research is the verification of the mass balance in all the repeated experiments.

Foram conduzidos dois experimentos em casa-de-vegetação com solo contaminado acidentalmente com metais pesados. No primeiro deles objetivou-se avaliar o potencial das espécies vegetais Brassica juncea, Raphunus sativus L., Hybiscus cannabinus e Amaranthus crentus em absorver, translocar e acumular zinco, cobre, manganês, chumbo e boro na parte aérea e assim atuarem na recuperação do solo estudado. No segundo experimento avaliou-se a aplicação dos materiais orgânicos: Solomax, turfa e concentrado húmico mineral como amenizantes de toxidez dos elementos citados para Brassica juncea. Em um terceiro experimento em laboratório, avaliou-se a capacidade de retenção de zinco pelos materiais orgânicos estudados. As variáveis avaliadas foram: produção de material vegetal; quantidades absorvida e acumulada de Zn, Cu, Mn, Pb e B; índices de translocação e remoção desses elementos pelas plantas. No solo foram determinados os teores total e disponível (DTPA e CaCl2 0,01 mol L-1) de Zn, Cu, Mn e Pb. No extrato de saturação foram determinados os teores solúvel (ICP-OES) e livre (eletroforese capilar) dos elementos citados, que também foram determinados na parte aérea da Brassica juncea. Resultados das determinações analíticas também foram introduzidos no programa de especiação iônica MINTEQ. Foram avaliados ainda os efeitos dos materiais orgânicos na distribuição dos metais pesados nas frações do solo. No ajuste dos dados do experimento de retenção de zinco foi a empregado o modelo de Freundlich e calculado o coeficiente de distribuição Kd. Embora as espécies tenham sido capazes de acumular quantidades elevadas dos elementos na parte aérea, elas não puderam ser consideradas hiperacumuladoras. O Amaranthus crentus e a Brassica juncea apresentaram os maiores índices de remoção para Zn, Mn e B. Os materiais orgânicos apresentaram valores elevados de Kd refletindo a alta afinidade do zinco pela fase sólida e elevada capacidade dos materiais orgânicos em imobilizar o elemento. Os elevados coeficientes de correlação obtidos para as isotermas de adsorção indicam que o modelo Freundlich pode ser utilizado para descrever a adsorção do Zn pelos materiais. A quantidade de Zn removida pelos materiais, em termos percentuais variou com o material e com o pH, sendo favorecida nos valores mais elevados. A turfa e o concentrado húmico reduziram os teores de metais extraídos por DTPA e CaCl2 sendo esta redução refletida nos teores acumulados pela Brassica juncea, contudo esta redução não foi suficiente para impedir os efeitos fitotóxicos dos elementos. A especiação dos metais mostrou que o Zn e o Mn encontraram-se principalmente na forma livre, enquanto que o Cu e o Pb apresentaram-se complexados à matéria orgânica dissolvida. Não foram observadas correlações significativas entre os teores dos metais determinados diretamente por eletroforese capilar e indiretamente pelo programa de especiação. O fracionamento seqüencial dos metais indicou que grande parte dos metais está ligada a frações pouco disponíveis do solo. Conclui-se que a aplicação da turfa e do concentrado húmico mineral juntamente com o cultivo do Amaranthus crentus e da Brassica juncea podem auxiliar na recuperação a longo prazo de solos contaminados com Zn, Mn Pb e B. / Two pot trials were carried out using a heavy metal accidentally contaminated soil. In the first one it was evaluated the plant absorbing, translocating and accumulating power for zinc, copper, manganese, lead and boron by the following vegetal species: Brassica juncea, Raphunus sativus L., Hybiscus cannabinus and Amaranthus crentus, so they could help in the rehabilitation of the studied soil. In the second pot trial it was evaluated the application of the following organic materials: Solomax, peat, and humic mineral concentrate for heavy metal toxicity amelioration for Brassica juncea. In a laboratory experiment it was evaluated the zinc retention capacity for the above mentioned organic materials. The measured variables were plant dry matter yield, absorbed and accumulated amounts of Zn, Cu, Mn, Pb and B; the translocation index and the removal index of metal by the mentioned plant species. The total and available (DTPA and CaCl2 001 mol L-1) content of Zn, Cu, Mn and Pb, were determined in soil samples. Concerning the soil saturation extract, the soluble and free forms of the mentioned elements were determined by ICPOES and capillary electrophoresis respectively. The same metals were also determined in the plant tissue of Brassica juncea. Analytical results were introduced in the chemical equilibrium program MINTEQ in order to calculate the concentration of the free forms of Zn, Cu, Mn, and Pb. A fractioning scheme for metals in soil was carried out in order to evaluate the effect of adding sources of organic carbon. The Freundlich model was used to account for zinc adsorption by the studied organic materials and the distribution coefficient Kd was also calculated. Although the plant species were able to accumulate high amounts of metal in the aerial part, that could not be considered as a hipper-accumulation process. The highest metal removal indexes were observed for Brassica juncea and Amaranthus crentus. Organic materials showed high values of Kd, reflecting its high affinity for zinc and a great capacity of zinc immobilization as well. Freundlich model was very effective in describing the adsorption isotherms as indicated by the high determination coefficients. Zinc retention was affected by the organic material type and pH, being favored by the highest studied pH values. Peat and humic mineral concentrate reduced the amounts of metals extracted by DTPA and CaCl2 in soil but the reduction was not great enough to avoid toxic effects in plants. Zn and Mn were present mostly as free cations in soil saturation extract while Pb and Cu were complexed by dissolved organic matter. It was not detected any significant correlation between free metal contents directly determined by capillary electrophoresis and those estimated by MINTEQ. Metal sequential extraction showed that most of them occurred in the least available fractions of soil. Peat and humic mineral concentrate application to soil associated with Amaranthus crentus and Brassica juncea cultivation may be used in long term amelioration of soils contaminated with Zn, Mn, Pb and B.

Foram conduzidos dois experimentos em casa-de-vegetação com solo contaminado acidentalmente com metais pesados. No primeiro deles objetivou-se avaliar o potencial das espécies vegetais Brassica juncea, Raphunus sativus L., Hybiscus cannabinus e Amaranthus crentus em absorver, translocar e acumular zinco, cobre, manganês, chumbo e boro na parte aérea e assim atuarem na recuperação do solo estudado. No segundo experimento avaliou-se a aplicação dos materiais orgânicos: Solomax, turfa e concentrado húmico mineral como amenizantes de toxidez dos elementos citados para Brassica juncea. Em um terceiro experimento em laboratório, avaliou-se a capacidade de retenção de zinco pelos materiais orgânicos estudados. As variáveis avaliadas foram: produção de material vegetal; quantidades absorvida e acumulada de Zn, Cu, Mn, Pb e B; índices de translocação e remoção desses elementos pelas plantas. No solo foram determinados os teores total e disponível (DTPA e CaCl2 0,01 mol L-1) de Zn, Cu, Mn e Pb. No extrato de saturação foram determinados os teores solúvel (ICP-OES) e livre (eletroforese capilar) dos elementos citados, que também foram determinados na parte aérea da Brassica juncea. Resultados das determinações analíticas também foram introduzidos no programa de especiação iônica MINTEQ. Foram avaliados ainda os efeitos dos materiais orgânicos na distribuição dos metais pesados nas frações do solo. No ajuste dos dados do experimento de retenção de zinco foi a empregado o modelo de Freundlich e calculado o coeficiente de distribuição Kd. Embora as espécies tenham sido capazes de acumular quantidades elevadas dos elementos na parte aérea, elas não puderam ser consideradas hiperacumuladoras. O Amaranthus crentus e a Brassica juncea apresentaram os maiores índices de remoção para Zn, Mn e B. Os materiais orgânicos apresentaram valores elevados de Kd refletindo a alta afinidade do zinco pela fase sólida e elevada capacidade dos materiais orgânicos em imobilizar o elemento. Os elevados coeficientes de correlação obtidos para as isotermas de adsorção indicam que o modelo Freundlich pode ser utilizado para descrever a adsorção do Zn pelos materiais. A quantidade de Zn removida pelos materiais, em termos percentuais variou com o material e com o pH, sendo favorecida nos valores mais elevados. A turfa e o concentrado húmico reduziram os teores de metais extraídos por DTPA e CaCl2 sendo esta redução refletida nos teores acumulados pela Brassica juncea, contudo esta redução não foi suficiente para impedir os efeitos fitotóxicos dos elementos. A especiação dos metais mostrou que o Zn e o Mn encontraram-se principalmente na forma livre, enquanto que o Cu e o Pb apresentaram-se complexados à matéria orgânica dissolvida. Não foram observadas correlações significativas entre os teores dos metais determinados diretamente por eletroforese capilar e indiretamente pelo programa de especiação. O fracionamento seqüencial dos metais indicou que grande parte dos metais está ligada a frações pouco disponíveis do solo. Conclui-se que a aplicação da turfa e do concentrado húmico mineral juntamente com o cultivo do Amaranthus crentus e da Brassica juncea podem auxiliar na recuperação a longo prazo de solos contaminados com Zn, Mn Pb e B. / Two pot trials were carried out using a heavy metal accidentally contaminated soil. In the first one it was evaluated the plant absorbing, translocating and accumulating power for zinc, copper, manganese, lead and boron by the following vegetal species: Brassica juncea, Raphunus sativus L., Hybiscus cannabinus and Amaranthus crentus, so they could help in the rehabilitation of the studied soil. In the second pot trial it was evaluated the application of the following organic materials: Solomax, peat, and humic mineral concentrate for heavy metal toxicity amelioration for Brassica juncea. In a laboratory experiment it was evaluated the zinc retention capacity for the above mentioned organic materials. The measured variables were plant dry matter yield, absorbed and accumulated amounts of Zn, Cu, Mn, Pb and B; the translocation index and the removal index of metal by the mentioned plant species. The total and available (DTPA and CaCl2 001 mol L-1) content of Zn, Cu, Mn and Pb, were determined in soil samples. Concerning the soil saturation extract, the soluble and free forms of the mentioned elements were determined by ICPOES and capillary electrophoresis respectively. The same metals were also determined in the plant tissue of Brassica juncea. Analytical results were introduced in the chemical equilibrium program MINTEQ in order to calculate the concentration of the free forms of Zn, Cu, Mn, and Pb. A fractioning scheme for metals in soil was carried out in order to evaluate the effect of adding sources of organic carbon. The Freundlich model was used to account for zinc adsorption by the studied organic materials and the distribution coefficient Kd was also calculated. Although the plant species were able to accumulate high amounts of metal in the aerial part, that could not be considered as a hipper-accumulation process. The highest metal removal indexes were observed for Brassica juncea and Amaranthus crentus. Organic materials showed high values of Kd, reflecting its high affinity for zinc and a great capacity of zinc immobilization as well. Freundlich model was very effective in describing the adsorption isotherms as indicated by the high determination coefficients. Zinc retention was affected by the organic material type and pH, being favored by the highest studied pH values. Peat and humic mineral concentrate reduced the amounts of metals extracted by DTPA and CaCl2 in soil but the reduction was not great enough to avoid toxic effects in plants. Zn and Mn were present mostly as free cations in soil saturation extract while Pb and Cu were complexed by dissolved organic matter. It was not detected any significant correlation between free metal contents directly determined by capillary electrophoresis and those estimated by MINTEQ. Metal sequential extraction showed that most of them occurred in the least available fractions of soil. Peat and humic mineral concentrate application to soil associated with Amaranthus crentus and Brassica juncea cultivation may be used in long term amelioration of soils contaminated with Zn, Mn, Pb and B.

Les sols enrichis en éléments traces métalliques, encore appelés sols métallifères, constituent un modèle original pour l’étude des processus écologiques et évolutifs opérant au sein de la végétation qui y est associée. Au Sud-Est de l’Afrique centrale, dans l’ex-province du Katanga, une succession d’affleurements naturels de roches enrichies en cuivre (Cu) et en cobalt (Co), uniques en leur genre à la surface de la terre, forment le très célèbre « Arc Cuprifère Katangais ». De véritables « collines de cuivre », isolées géographiquement dans une matrice de forêt claire, s’étendent sur plus de 300 Km et constituent des écosystèmes remarquables attirant l’attention de chercheurs depuis plus d’un demi-siècle.Les conditions écologiques extrêmes des collines de Cu et de Co, liées principalement à des concentrations métalliques élevées dans les sols (1 000 à 10 000 fois supérieures aux sols normaux), ont sélectionné des végétations hautement originales et uniques composées de métallophytes, aussi appelées cupro-cobaltophytes, pouvant toutefois présenter des niches écologiques très variables. Certaines de ces plantes possèdent même la fascinante particularité d’accumuler le Cu et/ou le Co dans leurs tissus foliaires à des niveaux de concentrations extrêmement toxiques pour des plantes normales, et sont qualifiées arbitrairement d’hyperaccumulatrices. À ce jour, de fortes variations d’accumulation du Cu et du Co encore mal comprises sont observées chez ces cupro-cobaltophytes. Celles-ci pourraient être expliquées par une variation de la disponibilité du Cu et du Co dans les sols, par une variabilité des mécanismes de tolérance au Cu et au Co, et par une variation de la capacité à accumuler ces métaux.Certaines cupro-cobaltophytes sont restreintes de ces habitats métallifères hors-normes, et d’autres se distribuent plus largement, possédant également des populations au sein d’habitats non-métallifères, en savane ou en forêt, sur sol normal. Ces dernières, à plus large distribution écologique, sont qualifiées de cupro-cobaltophytes facultatives, et constituent des modèles biologiques hautement intéressants pour étudier les processus écologiques et évolutifs liés à la tolérance et l’accumulation du Cu et du Co.La biogéochimie, l’écologie et l’évolution de la tolérance et de l’accumulation du Cu et du Co chez les cupro-cobaltophytes facultatives restent à ce jour peu connues, surtout pour le Co. La stratégie de recherche repose sur une approche interdisciplinaire (biogéochimie, écologie fonctionnelle et écologie évolutive) qui s’articule autour de travaux réalisés à la fois sur le terrain et en conditions contrôlées :sur un sol et en hydroponie. Chez une métallophyte facultative modèle à large amplitude écogéographique :Anisopappus chinensis (Asteraceae), la thèse s’attache à :(i) caractériser la variation phénotypique de l’accumulation du Cu et du Co et connaître les facteurs édaphiques qui l’influence, pour mieux comprendre la disponibilité du Cu et du Co dans les sols métallifères, (ii) tester l’influence de facteurs chimiques du sol sur la mobilité du Cu et du Co dans les sols et sur l’accumulation foliaire de Cu et de Co, (iii) étudier la réponse adaptative de populations métallicoles et non-métallicoles provenant d’habitats très contrastés, et leur réponse plastique au Co, (iv) examiner la variabilité génétique de la capacité à tolérer et à accumuler le Cu et le Co entre des populations métallicoles et non-métallicoles.Les variations d’accumulation de Cu et de Co observées au sein des populations métallicoles d’A. chinensis seraient en grande partie influencées par la teneur en matière organique et les concentrations totales en manganèse et en fer des sols de la zone racinaire. Le Cu et le Co potentiellement dans la solution du sol ne sembleraient pas être les seules fractions expliquant les variations d’accumulation. Les concentrations de Cu et de Co liées respectivement aux oxydes de manganèse et à la matière organique pourraient également représenter une concentration significativement disponible pour les plantes. En culture, une variation dans la mobilité du Cu et du Co dans le sol n’expliquait pas nécessairement une variation dans les concentrations foliaires mesurées chez A. chinensis. La disponibilité du Cu et du Co en milieu métallifère est un concept difficile à appréhender, élément- et espèce-dépendant, qui serait la résultante d’interactions biogéochimiques complexes à l’échelle de la rhizosphère, impliquant les microorganismes.Il existe une réponse adaptative à la diversité d’habitats chez A. chinensis traduite par des hauteurs de plantes et surfaces de feuilles plus faibles chez les populations métallicoles étudiées. Malgré des sols très contrastés chimiquement entre habitats métallifères et non-métallifères, très peu de variations intraspécifiques de la surface foliaire spécifique et des concentrations foliaires en nutriments ont été observées. La très faible réponse plastique au Co chez les populations étudiées d’A. chinensis semble mettre en évidence une homéostasiedes traits fonctionnels foliaires mesurés, qui pourrait expliquer la large niche écologique de l’espèce.La tolérance au Cu n’est pas un attribut vérifié à l’échelle de l’espèce chez A. chinensis. Celle-ci semblerait s’exprimer dans des conditions édaphiques bien particulières sur les sols métallifères, et pourrait être le fruit de processus rhizosphériques impliquant les microorganismes. Une différenciation génétique de la tolérance au Co a été observée chez les populations métallicoles des sols enrichis en Co. Une relation positive entre le degré de tolérance au Co et le niveau de concentration en Co dans le sol natif existerait. Pour la première fois chez une métallophyte, une variation génétique de l’accumulation de Co a été mise en évidence. L’hyperaccumulation du Cu et du Co chez les métallophytes, à de faibles concentrations disponibles dans les sols, n’existerait pas. Anisopappus chinensis provenant de sols enrichis en Co constitue un matériel végétal remarquable à valoriser, puisque s’exprimant comme une véritable accumulatrice de Co en conditions contrôlées. / Soils enriched in trace metal elements (TE) (i.e. metalliferous soils) constitute original model systems to study ecological and evolutionary processes occurring among their associated vegetation. In Southeastern central Africa (Katanga), a unique succession of natural copper (Cu) and cobalt (Co) outcrops occurs; the so-called “Katangan Copperbelt”. Here, scattered over 300 Km, geographically isolated “copper hills” form remarkable ecosystems that strongly contrast with the surrounding clear forest.Soil Cu and Co concentrations of those hills can be 1000- to 10 000-fold higher than in normal soils. These extreme ecological conditions have selected a unique vegetation of metallophytes, also called cupro-cobaltophytes. Some plants have the fascinating peculiarity to accumulate Cu and Co in their tissues up to extremely phytotoxic concentrations and are called “hyperaccumulators”. High misunderstood inter and intraspecific variations of Cu and Co accumulation are observed within this flora. These variations could be explained by variations in Cu and Co availability in soils, but also by inter and intraspecific variations in Cu and Co tolerance mechanisms and capacity to accumulate Cu and Co.Some cupro-cobaltophytes are restricted to metalliferous habitats (i.e. endemic metallophytes) and some are widely distributed, with populations on normal soils (i.e. facultative metallophytes). These latter are of high interest to study the ecology and evolution of Cu and Co tolerance and accumulation.Biogeochemistry, ecology and evolution of Cu and Co tolerance and accumulation in facultative cupro-cobaltophytes remains poorly understood, especially for Co. Research strategy was to develop a transdisciplinary approach (biogeochemistry, functional ecology and evolutionary ecology) based on field works and experiments (using soil or nutrient solution). For Anisopappus chinensis (Asteraceae), a broad-niched and geographically widespread facultative metallophyte chosen as model species, this PhD thesis aims at: (i) investigate the phenotypic variation of Cu and Co accumulation, and the influencing soil chemical factors, to understand better Cu and Co availability in metalliferous soils, (ii) test the influence of soil chemical factors on Cu and Co mobility in soil and accumulation, (iii) studyadaptive response of metallicolous (M) and non-metallicolous (NM) populations from contrasted habitats, and their plastic response to Co, (iv) examine the genetic variation in Cu and Co tolerance and accumulation between M and NM populations.Copper and Co accumulation variations among M populations of A. chinensis are clearly influenced by organic matter content and soil total manganese and iron concentrations in the rooting zone. Mobile Cu and Co concentrations, potentially in the soil solution, would not only explain Cu and Co accumulation variations. Copper and Co bound to respectively manganese oxides and organic matter could also represent Cu and Co available concentrations for plants. In experimental conditions, variations in Cu and Co mobility would not necessary explained variations in foliar Cu and Co concentrations in A. chinensis. Copper and Co availability is a complex element- and species-specific mechanism, closely related to all biogeochemical processes that occur in the rhizosphere. Important role of microorganisms is expected.Adaptive response to habitats has been highlighted for A. chinensis. Metallicolous plants had consistently lower height and leaf size than NM plants. Despite strong contrast in the soil chemistry between metalliferous and non-metalliferous habitats, very few variations in specific leaf area and leaf nutrient concentrations was observed between M and NM populations. The low plastic response to Co seems to reveal homeostasis of the studied functional leaf traits, which might explain the broad ecological niche of the species.Tolerance to Cu is not constitutive of A. chinensis and would be express under specific growth conditions in nature. Expression of Cu tolerance could be the result of specific soil-plant-microorganisms processes. Genetic differentiation in Co tolerance has been demonstrated in M populations from Co-enriched soils. Positive relationship between the level of tolerance to Co and the concentration of Co in the native soil may exist. Genetic variability of Co accumulation has been demonstrated for the first time in a metallophyte. Cu and Co hyperaccumulation at low available concentration in the soil would not exist in metallophytes. Anisopappus chinensis form Co-enriched soils expressed as a genuine Co accumulator and thus, constitute an interesting valuable biological model. / Doctorat en Sciences / info:eu-repo/semantics/nonPublished