The Wonderfontein Cave is located within the Witwatersrand Basin (Gauteng province, South Africa) and is associated with the river banks of the Wonderfontein Spruit. This cave system has for many years been subjected to the influx of polluted mine water. Since subterranean environments remain poorly studied, it is unknown what the effect of this might be on the associated ecosystem. Furthermore, water that enters the Wonderfontein Cave poses a severe health threat as it drains into the underlying aquifer, which is abstracted for human and animal use. The general aim of this study was to determine the extent of metal pollution (enrichment), as well as to study the toxicity hazard potential of the soils and sediments associated with the Wonderfontein Cave. The objectives of this study were to (1) quantify the extent of anthropogenic metal pollution of water, soils and sediments associated with the Wonderfontein Cave (2) and assessing the toxicity hazard potential of these substrates; (3) determining whether nematode taxa and C. gariepinus individuals represent isolated communities and a population within the Wonderfontein Cave, respectively; (4) measuring the effect of mining-associated pollutants on the soil and sediment health of the Wonderfontein Cave by making use of nematodes to serve as bioindicators; (5) evaluating and comparing biomarker responses to metal bioaccumulation in C. gariepinus populations associated with the Wonderfontein Cave and epigean (surface) environments and lastly (6) comparing the results of the above identified assessments over both a temporal and spatial scale. Sampling was undertaken during April (1st sampling interval) and September (2nd sampling interval) 2013, which respectively represented the end of the high and low flow periods. Also, sampling of the subterranean (Wonderfontein Cave) and associated surface (Wonderfontein Spruit) environments were undertaken. The findings of this study suggested that especially the sediments associated with the Wonderfontein Cave have been subjected to severe nickel, copper, zinc, cobalt, aluminium, cadmium, lead and uranium enrichment. Also, the concentrations of many of the studied metals exceeded the respective water, soil and sediment environmental quality guidelines. Thus, also taking into consideration that most of the sediments were classified as being toxic, a severe threat is posed to the health of the associated biota. Although 60 nematode genera were identified from soil and sediments samples collected from the respective sampling sites associated with the Wonderfontein Cave and Spruit, it was concluded that most of these genera were likely only temporary residents of the subterranean environment. Even though plant-parasitic and nonparasitic nematodes were present, most of the collected soil and sediment samples were dominated by bacterivores (non-parasitic nematodes). Zero genetic divergence was recorded between the C. gariepinus populations associated with the Wonderfontein Cave and Stoffels Dam (Wonderfontein Spruit). However, significant temporal and spatial variation was observed in some bioaccumulated metals and biomarker responses within and between the respective C. gariepinus populations. Furthermore, the metal bioaccumulation levels present in both these fish populations pose a substantial threat to human health and are thus not fit for consumption. Also, no significant fish condition differences were observed between the C. gariepinus populations associated with the Wonderfontein Cave and Spruit. This study served as an initiative to create awareness and promote the conservation of Africa’s karst landscapes. / MSc (Environmental Sciences), North-West University, Potchefstroom Campus, 2015

The Wonderfontein Cave is located within the Witwatersrand Basin (Gauteng province, South Africa) and is associated with the river banks of the Wonderfontein Spruit. This cave system has for many years been subjected to the influx of polluted mine water. Since subterranean environments remain poorly studied, it is unknown what the effect of this might be on the associated ecosystem. Furthermore, water that enters the Wonderfontein Cave poses a severe health threat as it drains into the underlying aquifer, which is abstracted for human and animal use. The general aim of this study was to determine the extent of metal pollution (enrichment), as well as to study the toxicity hazard potential of the soils and sediments associated with the Wonderfontein Cave. The objectives of this study were to (1) quantify the extent of anthropogenic metal pollution of water, soils and sediments associated with the Wonderfontein Cave (2) and assessing the toxicity hazard potential of these substrates; (3) determining whether nematode taxa and C. gariepinus individuals represent isolated communities and a population within the Wonderfontein Cave, respectively; (4) measuring the effect of mining-associated pollutants on the soil and sediment health of the Wonderfontein Cave by making use of nematodes to serve as bioindicators; (5) evaluating and comparing biomarker responses to metal bioaccumulation in C. gariepinus populations associated with the Wonderfontein Cave and epigean (surface) environments and lastly (6) comparing the results of the above identified assessments over both a temporal and spatial scale. Sampling was undertaken during April (1st sampling interval) and September (2nd sampling interval) 2013, which respectively represented the end of the high and low flow periods. Also, sampling of the subterranean (Wonderfontein Cave) and associated surface (Wonderfontein Spruit) environments were undertaken. The findings of this study suggested that especially the sediments associated with the Wonderfontein Cave have been subjected to severe nickel, copper, zinc, cobalt, aluminium, cadmium, lead and uranium enrichment. Also, the concentrations of many of the studied metals exceeded the respective water, soil and sediment environmental quality guidelines. Thus, also taking into consideration that most of the sediments were classified as being toxic, a severe threat is posed to the health of the associated biota. Although 60 nematode genera were identified from soil and sediments samples collected from the respective sampling sites associated with the Wonderfontein Cave and Spruit, it was concluded that most of these genera were likely only temporary residents of the subterranean environment. Even though plant-parasitic and nonparasitic nematodes were present, most of the collected soil and sediment samples were dominated by bacterivores (non-parasitic nematodes). Zero genetic divergence was recorded between the C. gariepinus populations associated with the Wonderfontein Cave and Stoffels Dam (Wonderfontein Spruit). However, significant temporal and spatial variation was observed in some bioaccumulated metals and biomarker responses within and between the respective C. gariepinus populations. Furthermore, the metal bioaccumulation levels present in both these fish populations pose a substantial threat to human health and are thus not fit for consumption. Also, no significant fish condition differences were observed between the C. gariepinus populations associated with the Wonderfontein Cave and Spruit. This study served as an initiative to create awareness and promote the conservation of Africa’s karst landscapes. / MSc (Environmental Sciences), North-West University, Potchefstroom Campus, 2015

The environmental bioavailability of contaminants, rather than their total concentrations in the soil compartment play a decisive role for the risks associated with contaminated sites. Various soil constituents and abiotic conditions have strong influence on bioavailability, which may vary substantially between different locations. It is therefore necessary to site-specifically use tools that reflect the fractions of contaminants that are available to biota and pose the highest potential environmental risks. Bioassays provide integrated toxic responses which include effects from unknown contaminants or combinatory toxic effects from mixtures of contaminants. Thus, biological effect data greatly contribute to establish more realistic exposure and risk-scenarios at contaminated sites. The work underlying this thesis presents possible techniques for high capacity screening for site-specific hazards at contaminated areas. By combining rapid water extractions and cell-based in vitro designs measures of the toxic potential in soils was obtained. Toxicologically bioavailable fractions of mixed metal pollution, including arsenic, were primarily investigated in this thesis. In two of the studies, environmental availability and toxicological bioavailability of arsenic was explored in CCA-contaminated soils. Application of cell-based in vitro screening techniques was also conducted at a metal contaminated industrial site to obtain spatial distribution of toxicity. Multivariate association techniques were employed in the interpretation of environmental exposure and cytotoxicity data. It was shown that cell-based in vitro systems for both basal cytotoxicity and specific end-points targeting arsenic could assess the toxic potential from extracts obtained by several water-based extraction techniques including Pressurised Liquid Extraction (PLE). The cell-based in vitro systems were found to add important information on the site-specific differences in arsenics genotoxic potential from CCA-contaminated soils. The results highlight the importance of taking speciation and toxicological bioavailability into account in the risk analysis, rather than to base risk estimates on total load of contaminants. The presented screening approach was successfully applied at a metal polluted industrial site where spatial distribution of toxicity was obtained. PLE extraction also provided means for combined toxicological and chemical screening of explosives in soils from live-fire training ranges. Multivariate association techniques highly facilitated the interpretation of complex environmental data. The PLE was found to be a rapid extraction technique that has sufficient environmental relevance to be used in environmental impact analyses. It was also concluded that other cell-based in vitro systems that target specific toxic effects have large potential for being used in screening for a variety of environmental chemicals. Keywords: Environmental availability, Environmental bioavailability, Toxicological bioavailability, mixture toxicity, hazard screening, contaminated soils, heavy metals, arsenic, CCA, explosives, soil extraction, water extracts, cell-based in vitro tests, cytotoxicity, genotoxicity, PLE, MVDA, PCA, PLS. / Föroreningars biotillgänglighet snarare än deras totala koncentration i markmiljön styr den risk som kan förknippas med förorenade områden. Biotillgängligheten är ofta långt från 100% p.g.a. en rad olika bindningsytor och processer i jorden som reducerar biotillgängligheten. Således kan biotillgängligheten variera kraftigt mellan olika förorenade platser och även inom samma plats till följd av de specifika förhållanden som råder på respektive plats. Tillämpning av biologiska indikatorer som ger ett mått på den samlade giftigheten från biotillgängliga föroreningar är därför viktiga verktyg i platsspecifika exponerings- och farobedömningar. Många biologiska tester är ofta laborativt intensiva och dyra och lämpar sig mindre väl i testning av ett stort antal prover vilket är önskvärt om en tillräcklig geografisk täckning ska uppnås över ett förenat område. Testsystem som har kapacitet att hantera många prover till en rimlig kostnad är därför mycket användbart för screening i ett inledande skede av en miljöriskanalys av ett förorenat område. Föreliggande avhandlingsarbete presenterar möjliga lösningar i att kombinera snabb vattenextraktionsmetodik med cellbaserade in vitro system för platsspecifik toxikologisk faroscreening av metallförorenade områden. Metodiken erbjuder hög kapacitet för många jordprover. Tillämpning av metodiken har gjorts mot huvudsakligen metallföroreningar, inklusive arsenik. I två delarbeten studerades två modelljordar från CCA-förorenade fastigheter avseende tillgänglighet och giftighet av framför allt arsenik. Vidare studerades om det med applicerad metodik gick att illustrera geografisk utbredning av toxicitet, mätt i cellbaserade in vitro system, som biotillgängliga föroreningar uppvisar på ett metallförorenat industriområde. Slutligen studerades lämpligheten i att använda PLE för kombinerad kemisk och toxikologisk screening av jordar från militära skjutfält som var förorenade med explosivämnen. Cellbaserade in vitro system för mätning av både generell toxicitet och mer specifika effektmarkörer för arsenik visade sig användbara vid mätning från flera vattenbaserade extraktionsmetoder, inklusive PLE (trycksatt vätskeextraktion). Resultaten visade på PLEs tillämplighet som en snabb extraktionsmetod med bibehållen relevans för miljöanalyser. Applikation av cellbaserade in vitro system på vattenextrakt från förorenad jord gav värdefull information bl.a. om platsspecifik genotoxisk potential där specieringen av arsenik hade avgörande betydelse i en fallstudie med CCA-förorenade jordar. Vattenextraktion av jordprover kombinerat med cellbaserade in vitro system kunde också ge en geografisk bild av den omedelbara faran från biotillgängliga metallföroreningar inom ett industriområde. Vattenextraktion med PLE visade sig även användbart för screening av explosivämnen där extrakten direkt kunde användas för såväl kemisk karakterisering som för toxikologisk analys. Även andra typer av in vitro system än de som användes i detta arbete har stor framtida potential för tillämpning i faroscreening av ett stort antal olika typer av miljöföroreningar.

Magister Scientiae - MSc / The aim of this study was to use plants to determine the degree of heavy metal contamination in water and sediments in order to effectively monitor and provide possible recommendation to improve the water quality in the aquatic ecosystem of the Bottelary River.

Polygonum hydropiper L. was cultivated on alluvial soil (Pb = 2.6 mg/kg, dry weight) and Pb contaminated soil (Pb = 1,380 mg/kg dry weight) without and with amendment of 2 g organic fertilizer/kg soil and 2.5; 5.0; 10.0 g NPK fertilizer/1kg soil. After 45 days of cultivation, the growth in height and biomass of P. hydropiper in Pb contaminated soil without amendment of fertilizer was lower than that in alluvial soil, but the Pb content in the above-ground part of the P. hydropiper was higher. In the formula 4, on Pb contaminated soil (Pb = 1,380 mg/kg dry weight), when amending 2 g of microbiological organic fertilizer + 5 g NPK fertilizer per 1 kg of soil (with the total nutrients amended were: N = 0.25, P2O5= 0.52, K2O = 0.15, and organic matter = 0.21 g/kg soil), growth of P. hydropiper was optimal (its height and biomass were up to 244.0% and 284.9% in comparison to that of before experiment) and their Pb extraction potential was promoted to the highest level among the formulae used. The average level of Pb accumulated in the above-ground part of P. hydropiper cultivated at formula 4 was 1,098.3 mg/kg dry weight (DW) after 45 days of cultivation that was 1.6 time higher than the one of formula 2 without fertilizer amendment (687.8 mg/kg DW). The potential of Pb extracted and stored in the above-ground part of P. hydropiper cultivated at formula 4 after 45 days was 479.2 g/ha that was 2.85 time higher than the one of formula 2 without fertilizer amendment (168.02 g/ha). / Nghể răm Polygonum hydropiper L. được trồng theo 5 công thức trên đất phù sa không ô nhiễm chì (Pb = 2,6 mg/kg khô) và đất ô nhiễm chì (Pb = 1380 mg/kg khô)không bón phân và có bón phân với các liều lượng 2 g phân hữu cơ vi sinh/1 kg đất và 2,5; 5,0; 10,0 g phân NPK /1 kg đất. Sau 45 ngày thí nghiệm trồng cây, tăng trưởng về chiều cao và sinh khối của cây trên đất ô nhiễm Pb không bón phân thấp hơn trên đất phù sa, nhưng hàm lượng Pb trong phần trên mặt đất của cây cao hơn. Ở công thức (CT) 4, trên đất ô nhiễm chì (Pb = 1380 mg/kg khô) khi bón phân với liều lượng 2 g hữu cơ + 5 g NPK/1 kg đất (với tổng hàm lượng dinh dưỡng được bón là: N = 0,25, P2O5= 0,52, K2O = 0,15, và chất hữu cơ = 0,21 g/kg đất) thì tăng trưởng của câynghể răm đạt tối ưu (chiều cao và khối lượng đạt 244,0 % và 284, 9% so với trước thí nghiệm) và tiềm năng hút thu Pb của chúng cũng được thúc đẩy cao nhất trong số các công thức được sử dụng. Lượng Pb trung bình tích lũy trong phần trên mặt đất của nghể răm ở CT 4 đạt 1.098,3 mg/kg khô sau 45 ngày trồng, cao gấp 1,6 lần so với cây ở công thức 2 không bón phân (687,8 mg/kg khô). Khả năng loại bỏ Pb từ đất ô nhiễm của nghể răm khi được bón phân ở CT4 đạt 479,2 g/ha sau 45 ngày trồng, cao gấp 2,85 lần so với cây ở CT 2 không bón phân (168,02 g/ha).

Thesis (PhD)--Stellenbosch University, 2011. / ENGLISH ABSTRACT: The terrestrial Oligochaete species Eisenia andrei and Enchytraeus doerjesi were exposed to different concentration series of Cd and Zn, both separately and in mixtures for 28 days in artificial OECD soil at 15, 20 and 25°C. At the end of the four week exposure period, survival and reproduction were assessed in E. doerjesi and survival, reproduction, biomass change, metal uptake and biomarker responses (MTT and comet assays) in E. andrei. Survival results for both E. andrei and E. doerjesi indicated that the lethality of Cd increased at higher temperatures, whereas the opposite was observed for Zn.Cadmium LC50 values were the highest at 15°C and the lowest at 25°C. In the Zn exposures LC50 increased with increasing temperature. Mixture results in both test organisms indicated that mixtures were less lethal than the metals separately. Effects of mixtures on survival, nonetheless, increased with increasing temperature. In the Cd experiment, reproduction in E. andrei was only recorded in the control treatments at the three temperatures investigated. However, the deleterious effect of Zn on the reproduction of both E. andrei and E. doerjesi decreased with increasing temperature and Zn EC50 for reproduction increased with increasing temperature. Results for exposures to mixtures indicated in both test organisms that the interaction between Cd and Zn were antagonistic. In both E. andrei and E. doerjesi, the effect of mixture exposures on reproduction decreased with increasing temperatures. The highest mixture EC50 values for reproduction were found at higher temperature. In E. doerjesi (using reproduction results in MixToxModules) Cd and Zn interactions were dose level dependent at the three temperatures investigated. Antagonism was the predominant interaction at lower mixture concentrations whereas synergism occurred at mixture concentrations equal to or higher than the mixtures’ EC50 values. Biomass loss increased with increasing temperature in the Cd exposures (p 9 0.05) but not in the Zn exposures in E. andrei. In this species mixture results indicated antagonistic interactions between Cd and Zn at all temperatures investigated. The deleterious effect of mixtures on the biomass of E. andrei increased with increasing temperature. When Cd and Zn interactions were further investigated in E. andrei (using biomass results in MixToxModules) it was found that they were dose level dependent at the three temperatures investigated. Antagonism was the predominant interaction at lower mixture concentrations whereas synergism occurred at mixture concentrations higher than the mixtures EC50 values. The assessment of metal uptake in E. andrei revealed a temperature dependent Cd uptake with higher Cd body burdens occurring at higher exposure concentrations and temperatures (p ≤ 0.05). In the case of Zn, although uptake was lower at higher temperature, there was no statistical difference in uptake between exposure concentrations and between temperatures. Mixture results however indicated that in mixture exposures less Cd was accumulated by E. andrei than in single Cd exposures (p ≤ 0.05). Inversely, in mixture exposures more Zn was accumulated by E. andrei than in single Zn exposures (p ≤ 0.05). Biomarker studies revealed that Cd and Zn were both cytotoxic and genotoxic whether in single or mixture exposures. Factorial ANOVA analyses of the effects of temperature and metals on the reduction of MTT by E. andrei indicated that temperature rather than the metals was the most important factor controlling mitochondrial activity (p < 0.001). In both Cd and Zn exposures significant deleterious metal effects on mitochondrial processes were found to increase with temperature (p ≤ 0.01). Mixture exposures indicated decreasing cytotoxicity with increasing temperature (p ≤ 0.05) and possible antagonism between Cd and Zn at cellular level. Results of the comet assay showed that the genotoxic profile of Cd was the opposite of the genotoxic profile of Zn. Cd was less genotoxic at lower temperature and increasingly deleterious at higher temperature while Zn was more genotoxic at lower than higher temperature (p ≤ 0.05). The results of mixture exposures indicated decreasing mixture genotoxicity with increasing temperature and suggested that the interactions between Cd and Zn at molecular level were probably antagonistic. / AFRIKAANSE OPSOMMING: Eisenia andrei en Enchytraeus doerjesi is aan verskillende konsentrasiereekse van Cd en Zn, afsonderlik en in mengsels, vir 28 dae in OECD kunsmatige grond onderskeidelik by 15, 20 en 25°C blootgestel Die volgende eindpunte is aan die einde van die vier weke blootstellingsperiode gemeet: oorlewing en voortplanting (by E. doerjesi) en oorlewing, voortplanting, biomassaverandering, metaalopname, MTT en komeettoetse (by E. andrei). Oorlewingsresultate by beide E. andrei en E. doerjesi het getoon dat toenemende temperatuur die letale toksisiteit van Cd laat toeneem terwyl die teenoorgestelde waar was vir Zn. By die Cd blootstellings was die LK50 waardes die hoogste by 15°C en die laagste by 25°C. By die blootstellings aan Zn het die LK50 waardes toegeneem by hoër temperatuur. Resultate by die mengsels by beide toestspesies het aangetoon dat die mengsels minder letaal was as die afsonderlike metale. Effekte van mengsels op oorlewing het nietemin toegeneem met toenemende temperatuur. By die Cd blootstellings is voortplanting slegs by die kontroles en in die geval van E. andrei by die drie onderskeie temperature ondersoek. Die nadelige uitwerking van Zn op voortplanting by beide E. andrei en E. doerjesi het afgeneem met stygende temperatuur en die Zn LK50 vir voortplanting het toegeneem met toenemende temperatuur. Resultate van die blootstellings aan mengsels het getoon dat die wisselwerking tussen Cd en Zn by beide spesies antagonisties was. By beide spesies het die invloed van die mengsels op voortplanting afgeneem met stygende temperatuur. Die hoogste mengsel LK50 waardes vir voortplanting is by hoër temperature gevind. By E. doerjesi was Cd en Zn wisselwerkings by blootstelling aan mengsels (voortplantingsresultate ondersoek deur van MixToxModules gebruik te maak) dosisvlak verwant by die drie temperature wat ondersoek is. Antagonisme was die oorwegende wisselwerking by laer mengsel konsentrasies terwyl sinergisme voorgekom het by mengsel konsentrasies gelyk aan of hoër as die LK50 waardes van die mengsels. In die geval van die Cd blootstellings by E. andrei het biomassaverlies toegeneem met toenemende temperatuur (p ≤ 0.05) maar nie by die Zn blootstellings nie. Resultate van blootstellings aan mengsels het getoon dat die uitwerking van mengsels op die biomassa van E. fetida toegeneem het met toenemende temperatuur. By die verdere ondersoek van Cd en Zn wisselwerkings, waar gekyk is na dosis verhouding of dosisvlak antagonisme (deur van biomassa resultate in MixToxModules gebruik te maak), is gevind dat Cd en Zn wisselwerkings dosisvlak afhanklik was by die drie temperature wat ondersoek is. Antagonisme was die oorwegende wisselwerking by laer mengselkonsentrasies terwyl sinergisme voorgekom het by mengselkonsentrasies hoër as die mengsel EK50 konsentrasies. Die bepaling van metaalopname deur E. andrei het ’n temperatuurafhanklike opname van Cd getoon met hoër Cd liggaamskonsentrasies by hoër blootstellingskonsentrasies en temperature (p ≤ 0.05). Alhoewel Zn opname laer was by hoër temperatuur was daar geen statisties betekenisvolle verskille in opname tussen blootstellingskonsentrasies of temperature nie. Die bepaling van metaalopname by wurms wat aan mengsels blootgestel is, het getoon dat minder Cd deur E. andrei opgeneem is as waneer die wurms aan Cd as enkelmetaal blootgestel is (p ≤ 0.05), Daarteenoor het die teenoorgestelde gebeur in die geval van Zn, Meer van die metaal is opgeneem wanneer E. andrei aan mengsels blootgestel is as aan die enkelmetaal. Biomerkerstudie het getoon dat Cd en Zn beide sito- en genotoksies kan wees ongeag of dit as enkelmetale of in mengsels toegedien is. Faktoriale ANOVA analises van die effekte van temperatuur en metale op die verlaging van MTT by E. andrei het getoon dat temperatuur ’n belangriker faktor was as metaalbesoedeling by die kontrole van mitochondriale aktiwiteit. (p ≤ 0.001). By beide Cd en Zn blootstellings was daar in elk geval statisties betekenisvolle metaaleffekte op mitochondriale prosesse met toename in temperatuur (P ≤ 0.01). By blootstellings aan mengsels is gevind dat sitotoksisiteit afgeneem het met toenemende temperatuur (p ≤ 0.05) asook ’n moontlike antagonisme tussen Cd en Zn op sellulêre vlak. Resultate van die komeettoets het getoon dat die genotoksiese profiel van Cd die teenoorgestelde was as die van Zn. Cd was minder genotoksies by laer temperature en meer en meer skadelik by hoër temperature terwyl Zn meer genotoksies was by laer as by hoër temperature (p ≤ 0.05). Die resultate van blootstlling aan mengsels het laer genotoksisiteit getoon met toename in temperatuur. Dit dui daarop dat wisselwerkings tussen Cd en Zn op molekulêre vlak moontlik antagonisties was.

M. Tech. (Department of Chemistry, Faculty of Applied and Computer Sciences), Vaal University of Technology. / Removal of wastewater pollutants is urgent as they are continuously defiling the limited freshwater resources, affecting the ecosystem, aquatic and terrestrial life. Carbon nanotubes-based adsorbent materials are effective for removal of wastewater pollutants owing to their large specific surface area. Surface modification of carbon nanotubes (CNTs) can mediate specific pollutant adsorption and increase CNTs colloidal stability and chemical reactivity. Heavy metal pollution of wastewater is one of the major threats, as this metals can be toxic to humans when present at certain concentrations in drinking water. This study report the synthesis of chlorine functionalized and nitrogen doped carbon nanotubes (ClNCNTs) loaded with iron oxide nanoparticles and their use as adsorbents for Pb2+ ions in aqueous solutions. Carbon nanomaterials that are functionalized with chlorine and doped with nitrogen were successfully synthesized. This was done through pyrolysis of a mixture of dichlorobenzene and acetonitrile (in a 1:1 volume ratio) over 10% Fe-Co/CaCO3 bi-metallic catalyst via chemical vapour deposition (CVD) method. Addition of chlorine and nitrogen to the CNTs was to enable defect and disorder creation on the surface of the nanotubes which is envisaged to create nucleation sites on the their surface for better adhesion of the iron oxide nanoparticles. Different loadings of magnetite (Fe3O4) nanoparticles on the surface of the ClNCNTs was achieved using a co-precipitation method. The synthesized materials were charaterized by Raman spectroscopy, Transmission electron microscopy (TEM), Powder X-ray diffraction (PXRD) spectroscopy, Thermal gravimetric analysis (TGA), Brunauer Emmett and Teller (BET) and X-ray photoelectron spectroscopy (XPS). Highly defected CNTs, some with hollow and others with bamboo-compartments due to nitrogen inclusion were obtained. The effect of metal salt concentration in wt.% (10, 20, 30 and 53 wt.%) was investigated. The increase in wt.% loading has resulted in an increase in surface area, and a decrease in thermal stability as a result of defected Fe3O4/ClNCNTs. In addition, agglomeration was observed at 30 and 53 wt.% loading, due to large amount of iron present. The identity of the Fe3O4 nanoparticles was confirmed by PXRD and XPS with two iron peaks deconvoluted at 725.6 eV and 721 eV respectively. The percentage loading of the Fe3O4 nanoparticles at the surface of the ClNCNTs was affirmed by TGA analysis, where the residual mass obtained from TGA were closely related to the mass percentages added. Different nitrogen environments namely, the quatenary, pyridinic, pyrollic and nitrogen oxides were also observed, whilst chlorine could not be deconvoluted because it was present in very limited amount probably it was masked by the iron oxide nanoparticle. Thus, a 20 wt.% Fe3O4/ClNCNTs was chosen as an optimum, due to uniform distribution of spherical nanopaticles observed along the radial length of ClNCNTs that had an average size of 10 ± 4.5 nm. The synthesized ClNCNTs and a nanocomposite made from a 20 wt.% Fe3O4/ClNCNTs were applied in the removal of Pb2+ ions from aqueous solution. The results obtained showed that a nanocomposite made from a 20 wt.% Fe3O4/ClNCNTs had a better adsorption capacity of 17.0 mg/g as compared with 14.8 mg/g for ClNCNTs.

In Freiland- und Gefäßversuchen sollten Aufnahme der Schwermetalle (SM) und Wuchsverhalten an Kulturpflanzen im Hinblick auf die Wirkung von arbuskulären Mykorrhizapilzen (AM) Glomus intraradices, des Bakterienstammes Pseudomonas fluorescens (RA56) und als Dualinokulation auf Schwermetall belasteten Böden untersucht werden. Als Versuchspflanzen wurden Mais (Zea maize L.) und Weidelgras (Lolium multiflorum L.) und zusätzlich die Sonnenblume (Helianthus annuus L.) in Gefäßversuchen eingesetzt. Die Schwermetallkonzentrationen in den Böden lagen zwischen 0,5 und 56 mg Cd/kg TS, 17 und 650 mg Cu/kg TS und 110 und 1870 mg Zn/kg TS. Der Sonnenblumenertrag konnte im Gefäßversuch auf dem hoch belasteten Boden mit 36 mg Cd/kg TS, 556 mg Cu/kg TS und 1270 mg Zn/kg TS durch den AM-Pilz um 220 % und in der Dualinokulation um 348 % signifikant zur Kontrolle gesteigert werden. Gleichzeitig erhöhte die AM die Cd-Aufnahme um 29 % und die Dualinokulation um 66 %. Trotz der hohen Cd-Konzentrationen wiesen die Pflanzen mit Pilzbehandlungen keine Schäden auf. Der Maisertrag wurde durch AM und Bakterien im Feldversuch und als Dualinokulation in den ersten beiden Versuchsjahren bis zu 47 % gefördert. Im dritten Versuchsjahr kam es zu einer Ausbreitung der Mykorrhiza auf der gesamten Fläche, so dass eine Wuchsförderung auch auf den Kontrollparzellen statt fand. Durch die Variabilität der Cd -Gesamtgehalte auf der Versuchsfläche war der Vergleich der Aufnahmeraten zwischen den Varianten nur über den Cd-Transferfaktor möglich. In den oberirdischen Pflanzenteilen des Maises zeichneten deutlich eine Abnahme und in die Wurzeln eine Zunahme des Cd-Transfers ab. Gleichzeitig wurden die Mykorrhizainfektionen an den Maiswurzeln erhöht. Durch die Anwendung der Mykorrhiza in Kombination mit den Pseudomonaden können im Mais über mehrere Anbaujahre das Cd in das Erntegut reduziert werden mit gleichzeitiger Ertragserhöhung. / Influence of Mycorrhiza fungi and associate bacteria on the availability of nutrients and heavy metal in culture plants on heavy metal soils The heavy metal uptake and the influence on plant growing in culture plants is analysed in this study. Field- and pot experiments were done. An emphasis of this study was focused on the effect of the arbuscular Mycorrhiza (AM) fungi Glomus intraradices, the bacteria Pseudomonas fluorescens (RA56) and as dual inoculation. Maize (Zea maize L.) and Italien rye (Lolium multiflorum L.) were used and in addition sunflowers (Helianthus annuus L.) in the pot experiments. The concentrations of heavy metals in the soil were measured between 0.5 and 56 mg Cd/kg dry weight (d.w.), 17 and 650 mg Cu/kg d.w. and 110 and 1870 mg Zn/kg d.w. The yield of sunflowers was increased significantly up to 220% with AM fungi in the pot experiments. Cd-absortion increased by 29% in the soil with 36 mg Cd/kg d.w., 556 mg Cu/kg d.w. and 1270 mg Zn/kg d.w. in those pot experiments. With AM and Pseudomonades the yield was increased significantly up to 348% compared to the control group, the Cd-absortion increased by 66%. In spite of high Cd-concentrations, the plants showed no damages. The yield of maize was increased up to 47% due to AM fungi and Pseudomonades in singular and dual inoculation in the field experiments during the first and second year. In the third year, the AM fungi were spread over the whole area and no differences of yield were found. Because of the heterogeneous concentrations of Cd in the field area the comparison of Cd assimilation was allowed only by analysis of the transfer factor in the variants. Within the experimental years, the AM infection rate and the Cd-transfer was increased in the maize roots. On the other hand, the Cd translocation rate was decreased in the shoots. The reduction of Cd rate in monocotyledonous plants and an increase of yield are estimated for long term application of the AM fungi with Pseudomonades.

Les activités des anciennes industries métallurgiques spécialisées dans la production et la transformation de plomb, de zinc ou d’autres métaux ont engendré une pollution métallique importante des sols selon la localisation de ces derniers. Des études actuelles proposent ainsi des solutions de requalification de ces sols via des processus de phytoremédiation afin de valoriser au maximum les plantes cultivées sur ces sols contaminés. Ce projet de thèse a ainsi été réalisé dans le but de valoriser des écocatalyseurs hétérogènes régénérables. Ces derniers sont obtenus à partir des plantes cultivées sur des sols contaminés dans la synthèse organique pour la production de composés à haute valeur ajoutée. Les sols contaminés utilisés dans ce projet proviennent de la région Hauts-de-France (Auby et Noyelles-Godault). Dans cette optique, quatre types de réactions ont été testés pour valider l’efficacité et la recyclabilité des écocatalyseurs : La réaction d’aminolyse, La réaction de Friedel-Crafts, La transformation d’aza-Michael, et enfin La réaction de transestérificationDes écocatalyseurs riches en zinc (EcoZn), obtenus à partir de la biomasse végétale de ray-grass (Lolium perenne L.), ont été caractérisés par des méthodes physico-chimiques puis validités leurs critères d’efficacité et de recyclabilité. Ceux-ci ont ainsi permis de synthétiser plusieurs séries de composés à activité anti-inflammatoires connus (piroxicam, méloxicam, idrocilamide) et de nouveaux, des inhibiteurs de la farnésyltransférase (à visée anticancéreuse ou pour combattre des maladies orphelines telles que la progéria) ou encore de découvrir un nouvel accepteur d’aza-Michael. / The activities of the former metallurgy industries specialized in the production and processing of lead, zinc or other metals have resulted in significant metallic pollution of soils depending on their location. Current studies propose solutions for the requalification of these soils through the phytoremediation processes in order to maximize the value of plants grown on these contaminated soils. This PhD thesis project was thus realized with the aim of valorizing regenerative heterogeneous ecocatalysts. The latter were obtained from the plants cultivated on contaminated soils in organic synthesis for the production of high value added molecules. The contaminated soils used in this project came from the Hauts-de-France region (Auby and Noyelles-Godault). In this light, four types of reactions have been tested to validate the efficiency and recyclability of ecocatalysts: The aminolysis reaction The Friedel-Crafts reaction The aza-Michael transformation The transesterification reactionThe zinc-rich ecocatalysts (EcoZn), obtained from the plant biomass of ryegrass (Lolium perenne L.), were characterized by physicochemical methods and validated their efficiency and recyclability criteria. The latter were successfully used to reach various series of compounds with anti-inflammatory activity known (piroxicam, meloxicam, idrocilamide) and new, inhibitors of farnesyltransferase (for anticancer purpose or to fight orphan diseases such as progeria) or to discover a new aza-Michael acceptor.

The present thesis aims to evaluate the use of antioxidant enzyme activities (AEA) of biofilm communities as biomarkers of metal pollution (mainly Zn) in fluvial ecosystems. In order to test AEA as biomarkers of metal pollution as well as their responses to environmental variables, several ecotoxicological experiments have been performed. A zoom from field studies (with high ecological realism) to microcosm experiments (under controlled conditions) has been done to fill the gap between both approaches and understand AEA responses under a multiple-stress (environmental and metallic) scenario. The field studies were carried out in the Riera d’Osor, a tributary of the Ter, located in the region of La Selva (Girona province) / L’objectiu d’aquesta tesi és avaluar l’ús de les activitats enzimàtiques antioxidants (AEA) dels biofilms com a biomarcadors de contaminació metàl·lica (principalment per zinc (Zn)) en els ecosistemes fluvials. Per tal d’estudiar la resposta de les AEA a la contaminació metàl•lica, així com també la resposta a altres canvis ambientals (situacions d’estrès múltiple), s’han realitzat diversos experiments ecotoxicològics fent un zoom des d’estudis de camp (amb un elevat realisme ecològic) fins a un estudi de laboratori utilitzant microcosmos (amb condicions controlades). Els estudis de camp s’han dut a terme a la riera d’Osor, afluent del Ter, situada a la comarca de la Selva (província de Girona)