Nitroaromatic explosives have contaminated millions of acres of soil and water across the globe since World War II with known mutagenic, carcinogenic, and ecotoxicological effects. Recently, the U.S. Army initiated a shift away from traditional explosive compounds, such as trinitrotoluene (TNT) and hexahydrotrinitrotriazine (RDX), towards new, insensitive high explosive formulations. The new formulations approved for use include “IMX-101” and “IMX-104,” which contain 2,4-dinitroanisole (DNAN), 3-nitro-1,2,4-triazol-5-one (NTO), and nitroguanidine (NQ). These mixtures are less prone to accidental detonation making storage, transport, and implementation of these formulations safer for soldiers. Furthermore, initial research indicates that these compounds are less toxic than the older analogues. Despite the apparent benefits, the new explosives have higher solubility (approximately 3-300 times) than the compounds they are replacing, and NTO and NQ are fairly recalcitrant to aerobic biodegradation. The refractory nature and high solubility of the compounds raises concerns about leaching and water contamination considering the previous scale of environmental contamination from production and use of legacy explosives, while feasible strategies for cleaning up the new chemicals from soil and water have not been developed. Therefore, there is a critical need for understanding of the mechanisms of biodegradation these compounds will undergo in the environment and in engineered systems. In addition, a number of questions remain about the photochemistry of the compounds and how they may transform in sunlit surface water. Accordingly, this thesis examines biological transformations of DNAN and NTO in vegetative, fungal, and bacterial organisms, as well as photolysis of NTO and NQ in aqueous solution and DNAN in plant leaves. I identified 34 novel biotransformation products of DNAN using stable-isotope labeled DNAN and high resolution mass spectrometry. Most identified biotransformation products were the result of a nitro-group reduction as the first metabolic step. Arabidopsis plants, a Rhizobium bacterium, and a Penicillium fungus all further metabolized DNAN to produce large, conjugated compounds, and no mineralization was observed in the systems studied. All three organisms reduced both para- and ortho-nitro groups of DNAN, with a dramatic preference for ortho reduction. I found that photodegradation of DNAN and its plant metabolites within Arabidopsis leaves could impact the phytoremediation of DNAN and other contaminants. Soil slurries acclimated to nitroaromatic wastewater degraded DNAN with and without carbon and nitrogen amendments and NTO with added carbon. Organisms capable of degrading DNAN and NTO were isolated, and NTO was transformed to urea, amino-triazolone, and hydroxyl-triazolone. Photolysis of NTO sensitized singlet oxygen formation and yielded hydroxyl-triazolone, nitrite, nitrate, and ammonium. The rate of photolysis of NTO increased over the neutral pH range, and natural organic matter quenched the photolysis of NTO. An unknown volatile product accumulated in the headspace of sealed reactors after NTO photolysis. Singlet oxygen degraded NTO and formed nitrite in stoichiometric yield. Photolysis of NQ produced nitrite and nitrate, but at high pH, the reaction occurred much faster than at neutral pH, and the mass balance of inorganic nitrogen was much lower. Further work should be done to investigate the mechanisms of and products from NTO and NQ photolysis.

Poplar (Populus deltoides x nigra DN34) tissue cultures removed 2,4,6-trinitrotoluene (TNT) from an aqueous solution in five days, reducing the toxicity of the solution from highly toxic Microtox® EC value to that of the control. 1,3,5-trinitro-1,3,5-triazacyclohexane (RDX) was taken up by the plant tissue cultures more slowly, but toxicity reduction of the solution was evident. The measurement of toxicity reduction of aqueous solutions containing TNT and RDX was performed using a novel methodology developed for use with the Microtox® testing system. Radiolabeled TNT and RDX were used to confirm removal of explosives from hydroponic solutions containing plant tissue cultures and to verify that toxicity did not change in solutions where no plant cultures were present (positive controls). High Performance Liquid Chromatography (HPLC) and Liquid Scintillation Counter (LSC) measurements confirmed removal of TNT and RDX from solutions containing poplar plant tissue cultures and constancy of the plant-free controls. In addition, metabolites were identified in remediated solutions by HPLC, confirming the mechanism by which plants can remediate groundwater, surface water, and soil solutions. Using an Affymetrix® microarray, poplar trees were exposed to TNT over 48 hours. General patterns, as well as significant downregulated and upregulated genes were studied. We identified several new genes that were implicated in the detoxification and metabolism of TNT by Populus. In particular, our results support the "green liver" model of different gene families being expressed during the time course experiments. This suggests an alignment with Phase I transformation, Phase II conjugation, and Phase III compartmentation processes. Many of the genes identified in this study were related to those significantly expressed in previous Arabidopsis studies, supporting the comparison between these two plants. Gene families represented as significant in this study were glucosyltransferases, glycosyltransferases, cytochromes, and phosphofructokinases. We saw patterns in the areas of respiration, citric acid cycle, shikimatic pathway, and toxic responses.

The energetic compounds TNT (2,4,6-trinitrotoluene), RDX (hexahydro-1,3,5-trinitro-1,3,5-triazine), and HMX (octahydro-1,3,5,7-tetranitro-1,3,5,7-tetrazocine) contaminate military testing ranges worldwide yet are known to be degraded by plants and microbes in the laboratory. However, these contaminants remain persistent in the environment and represent a health threat to both humans and ecosystems. The use of traditional soil remediation technologies, such as landfilling or incineration, require large excavation costs and disrupt the ecology of the site. Phytoremediation, the use of green plants for the in situ treatment of contaminants, may be the most appropriate means of treating energetic residues present at military testing ranges. Eglin Air Force Base (EAFB), located near Niceville, FL, is one of the largest military installations in the world and holds many plant and animal species which are threatened or endangered. The use of explosives during training exercises on firing ranges at EAFB has resulted in contamination of energetics on range soils. In an effort to increase range sustainability with respect to explosives contamination, EAFB has been established as the site where phytoremediation processes will be explored for this research.

<p>Mercury (Hg) pollution is a global environmental problem. Numerous Hg-contaminated sites exist in the world and new techniques for remediation are urgently needed. Phytoremediation, use of plants to remove pollutants from the environment or to render them harmless, is considered as an environment-friendly method to remediate contaminated soil <i>in-situ</i> and has been applied for some other heavy metals. Whether this approach is suitable for remediation of Hg-contaminated soil is, however, an open question. The aim of this thesis was to study the fate of Hg in terrestrial plants (particularly the high biomass producing willow, <i>Salix spp</i>.) and thus to clarify the potential use of plants to remediate Hg-contaminated soils.</p><p>Plants used for phytoremediation of Hg must tolerate Hg. A large variation (up to 30-fold difference) was detected among the six investigated clones of willow in their sensitivity to Hg as reflected in their empirical toxicity threshold (TT_95b), the maximum unit toxicity (UTmax) and EC50 levels. This gives us a possibility to select Hg-tolerant willow clones to successfully grow in Hgcontaminated soils for phytoremediation.</p><p>Release of Hg into air by plants is a concern when using phytoremediation in practice. No evidence was found in this study that Hg was released to the air via shoots of willow, garden pea (<i>Pisum sativum</i> L. cv Faenomen), spring wheat (<i>Triticum aestivum</i> L. cv Dragon), sugar beet (<i>Beta vulgaris </i>L. cv Monohill), oil-seed rape (<i>Brassica napus</i> L. cv Paroll) and white clover (<i>Trifolium repens</i> L.). Thus, we conclude that the Hg burden to the atmosphere via phytoremediation is not increased.</p><p>Phytoremediation processes are based on the ability of plant roots to accumulate Hg and to translocate it to the shoots. Willow roots were shown to be able to efficiently accumulate Hg in hydroponics, however, no variation in the ability to accumulate was found among the eight willow clones using CVAAS to analyze Hg content in plants. The majority of the Hg accumulated remained in the roots and only 0.5-0.6% of the Hg accumulation was translocated to the shoots. Similar results were found for the five common cultivated plant species mentioned above. Moreover, the accumulation of Hg in willow was higher when being cultivated in methyl-Hg solution than in inorganic Hg solution, whereas the translocation of Hg to the shoots did not differ.</p><p>The low bioavailability of Hg in contaminated soil is a restricting factor for the phytoextraction of Hg. A selected tolerant willow clone was used to study whether iodide addition could increase the plant-accumulation of Hg from contaminated soil. Both pot tests and field trials were carried out. Potassium iodide (KI) addition was found to mobilize Hg in contaminated soil and thus increase the bioavailability of Hg in soils. Addition of KI (0.2–1 mM) increased the Hg concentrations up to about 5, 3 and 8 times in the leaves, branches and roots, respectively. However, too high concentrations of KI were toxic to plants. As the majority of the Hg accumulated in the roots, it might be unrealistic to use willow for phytoextraction of Hg in practice, even though iodide could enhance the phytoextraction efficiency.</p><p>In order to study the effect of willow on various soil fractions of Hg-contaminated soil, a 5-step sequential soil extraction method was used. Both the largest Hg-contaminated fractions, i.e. the Hg bound to residual organic matter (53%) and sulphides (43%), and the residual fraction (2.5%), were found to remain stable during cultivations of willow. The exchangeable Hg (0.1%) and the Hg bound to humic and fulvic acids (1.1%) decreased in the rhizospheric soil, whereas the plant accumulation of Hg increased with the cultivation time. The sum of the decrease of the two Hg fractions in soils was approximately equal to the amount of the Hg accumulated in plants. Consequently, plants may be suitable for phytostabilization of aged Hg-contaminated soil, in which root systems trap the bioavailable Hg and reduce the leakage of Hg from contaminated soils.</p>

Mercury (Hg) pollution is a global environmental problem. Numerous Hg-contaminated sites exist in the world and new techniques for remediation are urgently needed. Phytoremediation, use of plants to remove pollutants from the environment or to render them harmless, is considered as an environment-friendly method to remediate contaminated soil in-situ and has been applied for some other heavy metals. Whether this approach is suitable for remediation of Hg-contaminated soil is, however, an open question. The aim of this thesis was to study the fate of Hg in terrestrial plants (particularly the high biomass producing willow, Salix spp.) and thus to clarify the potential use of plants to remediate Hg-contaminated soils. Plants used for phytoremediation of Hg must tolerate Hg. A large variation (up to 30-fold difference) was detected among the six investigated clones of willow in their sensitivity to Hg as reflected in their empirical toxicity threshold (TT95b), the maximum unit toxicity (UTmax) and EC50 levels. This gives us a possibility to select Hg-tolerant willow clones to successfully grow in Hgcontaminated soils for phytoremediation. Release of Hg into air by plants is a concern when using phytoremediation in practice. No evidence was found in this study that Hg was released to the air via shoots of willow, garden pea (Pisum sativum L. cv Faenomen), spring wheat (Triticum aestivum L. cv Dragon), sugar beet (Beta vulgaris L. cv Monohill), oil-seed rape (Brassica napus L. cv Paroll) and white clover (Trifolium repens L.). Thus, we conclude that the Hg burden to the atmosphere via phytoremediation is not increased. Phytoremediation processes are based on the ability of plant roots to accumulate Hg and to translocate it to the shoots. Willow roots were shown to be able to efficiently accumulate Hg in hydroponics, however, no variation in the ability to accumulate was found among the eight willow clones using CVAAS to analyze Hg content in plants. The majority of the Hg accumulated remained in the roots and only 0.5-0.6% of the Hg accumulation was translocated to the shoots. Similar results were found for the five common cultivated plant species mentioned above. Moreover, the accumulation of Hg in willow was higher when being cultivated in methyl-Hg solution than in inorganic Hg solution, whereas the translocation of Hg to the shoots did not differ. The low bioavailability of Hg in contaminated soil is a restricting factor for the phytoextraction of Hg. A selected tolerant willow clone was used to study whether iodide addition could increase the plant-accumulation of Hg from contaminated soil. Both pot tests and field trials were carried out. Potassium iodide (KI) addition was found to mobilize Hg in contaminated soil and thus increase the bioavailability of Hg in soils. Addition of KI (0.2–1 mM) increased the Hg concentrations up to about 5, 3 and 8 times in the leaves, branches and roots, respectively. However, too high concentrations of KI were toxic to plants. As the majority of the Hg accumulated in the roots, it might be unrealistic to use willow for phytoextraction of Hg in practice, even though iodide could enhance the phytoextraction efficiency. In order to study the effect of willow on various soil fractions of Hg-contaminated soil, a 5-step sequential soil extraction method was used. Both the largest Hg-contaminated fractions, i.e. the Hg bound to residual organic matter (53%) and sulphides (43%), and the residual fraction (2.5%), were found to remain stable during cultivations of willow. The exchangeable Hg (0.1%) and the Hg bound to humic and fulvic acids (1.1%) decreased in the rhizospheric soil, whereas the plant accumulation of Hg increased with the cultivation time. The sum of the decrease of the two Hg fractions in soils was approximately equal to the amount of the Hg accumulated in plants. Consequently, plants may be suitable for phytostabilization of aged Hg-contaminated soil, in which root systems trap the bioavailable Hg and reduce the leakage of Hg from contaminated soils.

Phytoremediation is a relatively new remediation technology that may be useful in removing organic and inorganic pollutants from soils. Much research has focused on this type of remediation in the past few years due to its potential as an efficient and cost effective technology.<p>The purpose of this project was to extensively monitor phytoremediation of diesel-contaminated field soils in the laboratory under simulated field conditions. The main objectives were: to examine petroleum hydrocarbon (PHC) transfer and degradation processes involved in phytoremediation of contaminated field soils; to compare phytoremediation of contaminated field soils with intrinsic bioremediation; and, to develop a rationally-based model that could be used as a starting point for a quantitative prediction of the rate of PHC removal.<p>To realize these objectives a series of laboratory scale experiments were designed and carried out. The experiments reproduced pole planting of hybrid poplars into diesel contaminated field soils from a former bulk fuel station. The experiments were conducted in a closed and controlled environment over a 215-230 day period with numerous aspects of the system being monitored including volatilization of PHC from the tree and soil, and microbial activity of the soil.<p>Monitoring data indicated that microbial degradation of the contaminant was by far the most influential monitored degradation pathway, accounting for 96.3 to 98.7% of the mass removed for soils containing poplars. The monitoring data also indicated a significant difference in the mass of contaminant removed from the soil for soils containing poplars compared to those without. The total estimated mass of contaminant removed varied between 8.3 and 27.7% of the initial mass for soils containing poplars and between 6.0 and 6.1% of the initial mass for soils without poplars. Lastly, using the monitoring data and the below ground biomass of the poplars from each of the experimental test cells, a rationally-based model was developed to be used as a starting point for quantitative prediction of the rate of PHC removal.

Phytoremediation is a relatively new remediation technology that may be useful in removing organic and inorganic pollutants from soils. Much research has focused on this type of remediation in the past few years due to its potential as an efficient and cost effective technology.<p>The purpose of this project was to extensively monitor phytoremediation of diesel-contaminated field soils in the laboratory under simulated field conditions. The main objectives were: to examine petroleum hydrocarbon (PHC) transfer and degradation processes involved in phytoremediation of contaminated field soils; to compare phytoremediation of contaminated field soils with intrinsic bioremediation; and, to develop a rationally-based model that could be used as a starting point for a quantitative prediction of the rate of PHC removal.<p>To realize these objectives a series of laboratory scale experiments were designed and carried out. The experiments reproduced pole planting of hybrid poplars into diesel contaminated field soils from a former bulk fuel station. The experiments were conducted in a closed and controlled environment over a 215-230 day period with numerous aspects of the system being monitored including volatilization of PHC from the tree and soil, and microbial activity of the soil.<p>Monitoring data indicated that microbial degradation of the contaminant was by far the most influential monitored degradation pathway, accounting for 96.3 to 98.7% of the mass removed for soils containing poplars. The monitoring data also indicated a significant difference in the mass of contaminant removed from the soil for soils containing poplars compared to those without. The total estimated mass of contaminant removed varied between 8.3 and 27.7% of the initial mass for soils containing poplars and between 6.0 and 6.1% of the initial mass for soils without poplars. Lastly, using the monitoring data and the below ground biomass of the poplars from each of the experimental test cells, a rationally-based model was developed to be used as a starting point for quantitative prediction of the rate of PHC removal.

The objectives of this study are to use phytoremediation ecotechnology to improve the long-term soil pollution contaminated by petroleum and its refined products, and to explore the influence of environmental factors to the effective degradation of TPH.This study is divided into three stages.First, we selected the biofuel crops seeds to test their diesel fuel pollution tolerance.The crops include soybean¡Bsunflower¡Bcanola and corn.This four Taiwanese common energy crops were selected to manually configure three levels of diesel fuel pollution(1000¡B5000¡B10000 mg kg-1)in soil test the seed tolerance experiments.The experimental results in the first stage exhibited that the best energy crop species and non-edible crop(Jatropha),are selected in second phase for contaiminated soil degradation experiment to explore the possible influence of enviromented factors,such as soil moisture¡BpH¡Btotal plate¡Ketc,and to explore the applied fertilizer to increase soil nutrients,whether it will affect the degradation of diesel pollution.Finally, in the third phase, the energy crops were used in the oil-contaminated site to assess their decontamination efficiency. From June 2011 to November 2011, the experimental results shown in the first phase of seed tolerance test, for a period of 30 days showed that the soybean diesel-contaminated soil presented the best tolerance.Although the germination rate was increased with the concentration from 80% to 27%, it showed the best growth conditions.Therefore, in the second phase of test run, the speices of soybean and jatropha were selected prepared with concentrations of 1745¡B6271 and 10072 mg kg-1 dry soil. After 90 day for phytoremediation, soybean group(S) were found that the residual concentrations in soil were measured equal to 524¡B809 and 1913 mg kg-1 dry soil,with the removal rates of 69.97%¡B87.09% and 81.01% respectively.The concentration level of 10000 ppm was found not reach our control standard of 1000 ppm. The soil planted by jatropha(J) showed that residual concentration in soils equial to 303¡B1864 and 4837 mg kg-1 dry soil, with removal rates equal to 82.61%¡B70.27% and 51.98% respectively.Through statistical regression analytical results, the soybean can handle up to a concentration of 5300 mg/kg for diesel, while jatropha can handle up to 2170 mg/kg in this system. Except for the concentration level of 1000 ppm can reach our control standards, the other two groups were found below the control standard. To improve the removal efficieneies, it was suggested that phyto remediation time can be extended.

Due to several reasons, treatment methods for a certain waste are oftennot available locally in the waste handling and management industry. This is especially true for regions which are not densely populated. This requires transports, the majority of which consumes fossil fuel. To avoid this, local waste treatment methods need to be developed. In this work it is investigated how treatment of one hazardous waste is done; water and sludge containing oil. Based on sustainability criteria three novel methods are presented that can be conducted locally; mycoremediation, phytoremediation and electrocoagulation. The methods are evaluated in a case study of a recycling company. Mycoremediation and electrocoagulation were found to be suitable in the case study, as long as some criteria are fulfilled. In addition it is shown what barriers exist in law, policies and practices that hinder local treatment of water and sludge containing oil.

The utilisation of a willow vegetation filter for the treatment of landfill leachate is an environmentally and economically appealing solution for landfill operators. Investigations into the design and efficacy of the system, the effects of landfill leachate irrigation on soil ecology, soil chemistry and willow growth were undertaken. Two low cost, high density polyethylene-lined experimental willow plots (25x50 m2) were installed at Cranford landfill, Northamptonshire, UK, and irrigated with landfill leachate between June 2001 and October 2005. During the growing season, leachate volume was often reduced to zero. On other occasions, maximum removal efficiencies of between 33 % and 75 % for total Kjeldahl nitrogen, chemical oxygen demand and sodium, potassium and chloride ions were determined in landfill leachate effluent samples. The addition of landfill leachate produced no negative effects on both soil and foliar macronutrients, which were found to be in the range for sufficient or optimum growth and where additional fertilisers would not bring about a further increase in yields. The effects of landfill leachate application on soil microbial communities were explored and were found to be significantly higher for dehydrogenase activity and ammonium oxidising bacteria in the plot receiving a higher rate of leachate application. An economic analysis was carried out to demonstrate the financial viability of a willow vegetation filter as a treatment for landfill leachate. Willow vegetation filters could provide a desirable alternative to conventional treatment systems, such as sequencing batch reactors, as they incur lower capital expenses and potentially similar operational costs. This study also identified additional revenue benefits in the region of £94 per hectare for wood chip heat/energy production. The results from the willow vegetation filter under investigation in this study demonstrated that this type of system can be effective, in terms of volume reduction and removal efficiency in landfill leachate, with no detrimental effect upon the trees or surrounding environment