Innovative means and methods were tested to develop an economical, pragmatic and environmentally sustainable soil remediation process for heavy metal contaminated soils. An unsaturated-flow soil washing procedure was devised to dissolve the soil-bound toxic heavy metals; the latter were extracted by a chemical washing solution that percolated through the soil matrix. Subsequently, the leached toxic heavy metals were selectively concentrated, by a chemical precipitation process, into a solid waste. Thereby, a fraction of the spent ethylenediaminetetraacetic acid (EDTA), within the washing and rinsing leachate, was theoretically regenerated and recycle-ready. / The unsaturated-flow washing procedure was perfected by applying different treatments to a soil from a secure landfill. This soil was contaminated with Cd, Co, Cr, Cu, Fe, Mn, Ni, Pb, S and Zn. The major contaminants were Fe, Pb, Zn, S, Cu and Mn, making up 25, 1.9, 1.0, 0.4, 0.4 and 0.2%wt of the soil. The extraction responses of the contaminants and those of Al, Ca, Mg and P were established for citric acid (0.5 M) and different molarities of diammonium EDTA ((NH4)2EDTA). The DOW Chemical Company supplied the (NH4)2EDTA (i.e. VERSENE), a 1.37M industrial cleaner, which roughly costs $1.85kg-1 in bulk. The affordability of VERSENE was a pre-condition for hoping to satisfy the economical feasibility of remediating trace metal contaminated soils. / Ultimately, the developed unsaturated-flow washing procedure was tested in a pilot-scale experiment, for its ability to remediate a soil from an abandoned car battery recycling facility. The latter soil was severely contaminated with Pb (3.9%wt). Drip irrigation was used to apply (NH4) 2EDTA and water-rinsing solutions to the surface of soil heaps that rested atop an impermeable barrier, which permitted the retrieval of the leachate. A cumulative EDTA input to the soil of 10.6% wt extracted 49.4% of the total Pb content of the soil. Alternatively, readily biodegradable citric acid barely extracted 2.2% of the total Pb content of the soil, for a cumulative input of 18.1% weight of soil. Different treatments were tested for their effectiveness in concentrating the leached toxic heavy metals into a solid waste. The Pb was best precipitated with Na2S alone, as it provided the most concentrated solid toxic waste. / The environmental sustainability of remediating trace metal contaminated soils was thoroughly examined, as per the amounts of chemical entrants and toxic waste by-products, and per the post-treatment leaching of toxic levels of the remaining and potentially toxic trace metals. (Abstract shortened by UMI.)

The overall goal of this study was to investigate the possibility of reducing NO3-- concentration in the lower soil horizon by promoting denitrification. The study looked at an inexpensive remediation practice for subsurface-drained fields in order to degrade N0j' and consequently diminish NO3-- pollution. The experiments were conducted on a corn field at the Macdonald Campus Farm. The field was composed of sandy loam soil underlined by a clay layer. / In this study, sucrose was injected at a concentration of 20 mg L --1 within the subirrigation water. Furthermore, the water table was maintained at approximately 70 cm from the soil surface in order to create an anaerobic environment that allows for denitrification. The purpose was to furnish dissolved organic carbon (DOC) to the resident microorganisms in order to carryout a higher amount of denitrification. / The NO3-- levels in ground water were monitored weekly. Gas samples were taken from the field to observe whether N2O emissions increased as a result of the treatment This was considered to be important since N2O is a greenhouse gas. / It was found that the addition of sucrose significantly resulted in a faster rate of denitrification. The higher biological activity severely damped the NO3-- peaks in the subsoil, which arose following major rainfall events. / The addition of sucrose at 20 mg L--1 was found to be significant in lowering the amounts of released N2O. Therefore, adding DOC in the subirrigation water not only helped remediate water contamination but also served as a remedy to atmospheric pollution. / In addition, the hydraulic conductivity of the soil was monitored to ensure that no bioclogging arose from an anticipated rise in the bacterial population due to the addition of sucrose. It was found that the addition of sucrose at 20 mg L--1 did not contribute in anyway to reduce the soil's hydraulic conductivity.

A major factor complicating the cleanup at many sites is co-contamination by both organic compounds and heavy metals. Whereas much research has focused on the removal of either organic compounds or metals, relatively few studies have investigated simultaneous removal of organic and inorganic pollutants from soil. / The studies reported in this thesis have evaluated a novel technique for the simultaneous mobilization of polychlorinated biphenyl (PCB) compounds and heavy metals (HMs) from a field contaminated soil. Soil extraction with washing aids {surfactants/cyclodextrin in combination with chelating reagent(s)} was optimized for mobilization efficiency, recovery/recycle of washing additives, and in parallel detoxification of mobilized contaminants. PCB extraction efficiencies were determined with a method that converted all the PCB congeners to dicyclohexyl by hydrogenation over palladium. Studies demonstrated that 10 minutes of ultrasonic mixing of field contaminated soil with a combination of surfactant (30 mL L-1) or cyclodextrin (100 g L-1) and a sparing quantity (2 mmoles) of EDTA, simultaneously mobilized appreciable quantities of PCBs and most analyte metals (Cd, Cu, Mn, Pb, Zn, Ni, Cr). / Relative to individual reagents, combinations of surfactant (Brij 98, Triton X-301, or Triton XQS-20) or cyclodextrin (RAMEB or HPCD) with EDTA did not influence PCB extraction efficiencies perceptibly. The presence of surfactant or cyclodextrin in admixture with EDTA did not appreciably change the efficiency of mobilization of most heavy metals (Al, Cd, Cr, Fe, Mn, Ni, and Zn) but did increase the recovery of Cu and Pb with nonionic surfactant and cyclodextrin. When coupled with PCB removal by hexane back-extraction and precipitation of the HMs (mediated by hydrolysis of zero-valent magnesium (Mg0)}, aqueous washing suspension was regenerated and recycled twice to mobilize more contaminants from the soil. Three sonication-washes with the same charge of reagent mobilized appreciable quantities of PCBs (68 - 83%) and virtually all of the available Cd, Cu, Mn, and Pb and lesser amounts of the Zn (56%), Ni (59%), and Cr (50%) but only small quantities of Al (28%) and Fe (30%). / The release of EDTA from heavy metals complexes was efficient for most metals (99%) but was influenced by the nature of surfactant. EDTA recovery (62-65%) post three cycles of soil washing, hexane back-extraction, and Mg 0 treatment was similar for all reagent combinations. Among surfactants and cyclodextrin, only anionic surfactants suffered losses to Mg0 treatment.

This study describes a laboratory scale development of an in-situ bioremediation method, which uses a water table management system to supply nutrients to subsoil microorganisms, for biostimulation and subsequent biodegradation of pollutants such as fertilizer-nitrate and hydrocarbons (e.g., diesel oils), in the unsaturated zone of the soil. The study, which was divided into two parts: first nitrate bioremediation and secondly diesel biodegradation, was carried out on packed soil columns. / For the nitrate study, different levels of glucose were introduced into packed soil columns, 1,000 mm long x 200 mm, diameter, via subirrigation in order to supplement the organic carbon levels in the soil. Two sandy soils were used, with 1.6% and 3.4% organic matter content, respectively; and the water table in the soil columns was maintained at a depth of 350 mm below the surface. Fertilizer-nitrate was applied to the soil surface at a rate of 180 kg/ha nitrate-N. Simulated rainfall was used to leach nitrates to lower depths. The efficacy of using the subirrigation system, as a method for nutrient delivery in the bioremediation of leached nitrate, was monitored with time and with reference to the nitrate residue, redox potential of the soil solution, and solubilized Fe and Mn. / Leached nitrate was denitrified to less than 10 mg/L nitrate-N, which is the limit permitted in drinking water. The ideal organic carbon range was considered to be the glucose level (20 mg/L glucose-C) that reduced mom nitrate and gave redox potential and soluble Fe and Mn levels, similar to the control soil solution, when subjected to 96 days of subirrigation. Successful delivery of nutrient for the bioremediation of nitrate, within the farm boundaries, will be considered a "break through" toward nitrate residue control if this novel approach to nitrate control is demonstrated in the field. The delivery method will offer a technical solution to on-farm nitrate pollution. It is inexpensive, easy to adopt, and does not require major changes in the current farm practices. / In the second part of the study, a diesel contaminated sandy soil was packed in columns, 2,000 nun long x 200 nun diameter. The subirrigation method was used to supply two different combinations of treatments to the microorganisms in the soil for the biodegradation of the diesel namely: air, water and nutrients (N, P etc.), and air and water. The success of using subirrigation, to deliver nutrients to the soil in the columns, was monitored by measuring the trend in the reduction of soil diesel-TPH (diesel-total petroleum hydrocarbon) residue with time. Results obtained from the treated columns were compared with each other, and with the control columns undergoing passive biodegradation. / The study showed that subirrigation can be used as a method of nutrient delivery in the -bioremediaton of diesel contaminated soil. The TPH in the contaminated soil decreased, from an initial 670 mg diesel TPH/kg soil to an acceptable level of 40 mg diesel TPH/kg soil, in 82 days in the columns subjected to a combination of nutrient, air and water treatments. If this method of delivering biostimulants to the subsoil microbial population is demonstrated in the field, it will be invaluable to in-situ bioremediation of contaminated soils.

The efficacies ofsoil extraction methods, namely, Soxhlet, sonication, agitation, alkaline digestion and the ethyl acetate micro-method, for monitoring soil bioremediation were evaluated using three soil types, Swartland, Rensburg and Hutton, encompassing the mineralogical range prevalent in Kwa Zulu Natal. Phenol, atrazine and the BTEX component of petrol were the molecules used in this study and were extracted under different spiking concentrations, after prolonged ageing times up to 21 days and after changing the composition of the spiking solution. It was concluded that extraction methods must be validated for the specific conditions under which they would be used, taking into consideration, soil type, spiking solutions, moisture content, weathering times and the analyte(s) in question. A preliminary appraisal of atrazine degradation in a Hutton soil was then made under the conditions of sterilized, fertilized/non-fertilized and non-sterilized, fertilized/nonfertilized soils. The predominant pathway of atrazine degradation was deemed to be chemically/abiotically mediated due to the soil pH and the presence of iron and aluminium oxides as well as the high levels of manganese in the soil. The results obtained prompted further study into atrazinecatabolism using soil-slurry reactors, under the conditions of carbon-limitation, nitrogen limitation, carbon/nitrogen non-limitation and carbon/nitrogen limitation. A comparison was made between inoculated and non-inoculated bioreactors. The ability of the indigenous microbial population to return the Hutton soil to its original pristine state was confirmed. The expense of inoculation and culture maintenance could be avoided since carbon and nitrogen supplementation would be as equally effective as inoculation. / Thesis (Ph.D.)-University of Natal, Pietermaritzburg, 2002.

Most natural products and xenobiotic molecules, irrespective of their molecular or structural complexity, are degradable by some microbial species/associations within particular environments. Atrazine- and selected petroleum hydrocarbon (benzene, toluen~ and 0-, m- and p-xylene (BTX))-degrading associations were enriched and isolated"trom atrazine- and petroleum hydrocarbon (PHC)-contaminated KwaZuluNatal loamy and sandy soils, respectively. In total, eight pesticide- and forty BTXcatabolizing associations were isolated. Electron microscopy revealed that, numerically, rods constituted the majority of the populations responsible for both atrazine and PHC catabolism. Cocci and, possibly, spores or fungal reproductive bodies were observed also. For the BTX-catabolizing associations, the population profiles appeared to be dependent on the enrichment pH and the molecule concentration. After combining selected associations, to ensure that all the isolated species were present, batch cultures were made to determine the optimum pH and temperature for growth; With an atrazine concentration of 30 mgr1, the highest specific growth rates, as determined by biomass (OD) changes, were recorded at 30DC and pH 4 although the rate§ at 25DC and pH 5 were comparable. For the BTX (50 mgr1)-catabolizing associations, the highest growth rates were recorded at pH 4 for the four temperatures (15, 20, 25 and 30DC) examined. The sole exception was p-xylene with the highest specific growth rate recorded at pH 5 and 30De. Batch and continuous (retentostat) cultivations in the presence/absence of methanol and under C- and N-limited conditions were used to investigate the impacts of the solvent and the catabolic potentials of a combined atrazine-catabolizing culture (KRA30). In general, different degradation rates were recorded for the culture in response to element limitation. Addition of citrate as the primary carbon source / effected atrazine (100 mg!"l) degradation rates comparable to that of Pseudomonas sp. strain ADP while succinate addition effected herbicide co-metabolism. Carbon supplementation may, therefore, be considered for site amelioration practices. To complement conventional culture-based microbiological procedures, molecular techniques were employed to explore the diversities and analyze the structures of the microbial communities. In parallel, anaerobic microbial associations which targeted atrazine were also characterized. The soil DNA isolation/characterization protocol adopted consisted of a clean-up step followed by the polymerase chain reaction (peR) and 16S rDNA fingerprinting by denaturing-gradient gel electrophoresis (DGGE). The preliminary results suggested that despite different, but chemically similar, petroleum hydrocarbon molecules, the common selection pressures of the primary enrichments effected the isolation of similar and complex aerobic microbial associations. Some similar numerically-dominant bands characterized the aerobic and anaerobic atrazine-catabolizing associations although distinct differences were also recorded on the basis of the enrichment/isolation pH value and the concentration of the herbicide. Cloning and sequencing were then used to identify some of the numerically-dominant and non-dominant association members. Community-level physiological profiling (CLPP) for physiological fingerprinting was made with Biolog EcoPlates and highlighted the differences in the isolated aerobic atrazine-catabolizing associations depending on the enrichment pH and molecule concentration. Logarithmic-phase cultures of the combined atrazine- and BTX-catabolizing associations were used to explore the association profiles following pH and temperaiure optImIzation. Although some common numerically-dominant components were maintained, differences in numerical and, possibly, activity dominance were observed in the 16S rDNA profiles in response to changes in pH and temperature. This indicated that environmental parameter optimization and characterization of catabolic association structure must precede bioaugmentation so that control of key variables will facilitate maintenance of the dominant site-specific species. Following KRA30 cultivation in the presence/absence of methanol and under carbon and nitrogen-limited conditions, the population fingerprints showed that the presence of methanol effected shifts in species numerical dominance and, possibly, changes in atrazine catabolic capacity. Also, Coulter counter results, optical density readings and 16S rDNA characterization by DGGE indicated that degradation rate changes were accompanied by shifts in species numerical/activity dominance within the association. Although N-limitation effected the highest rates of herbicide catabolism, a potential versatility of the combined association for bioaugmented and/or biosupplemented remediation with acceptable rates regardless of any elemental limitation was recorded. To determine if the contaminated and pristine source soils contained comparable catabolic populations and, thus, offered potential for intrinsic bioremediation, PCRDGGE was used to characterize the populations in comparison with the enriched/isolated associations. Some similar dominant bands characterized the contaminated soils and the enriched/isolated associations. The significance of this, in relation to a possible correlation between numerical and activity dominance in the component species, is discussed with respect to the use of PCR-DGGE to identify natural attenuation potential and monitor sustained intrinsic and enhanced (bioaugmented and biosupplemented) bioremediation. / Thesis (Ph.D.)-University of Natal, Pietermaritzburg, 2001.

The central aim of this study was to determine if indigenous fungi of an oil-contaminated soil could be effectively used in a low-cost bioremediation of the soil. Since some of the contaminant had been present at the site for over two decades, the indigenous microbial species had been subjected to specific selection pressures for a protracted period, thus facilitating key enzymatic capabilities for hydrocarbon degradation. Analysis of the pertinent influential parameters of soil bioremediation indicated that an ex situ technique, utilising the catabolic activities of the indigenous soil fungi, was a feasible low-cost option. Fungi were isolated from the contaminated soil through a variety of techniques. The abilities of these isolates to degrade the contaminant oil and a range of representative hydrocarbon molecules was evaluated by a systematic screening programme. Sixty-two isolates were initially examined for their growth potential on hydrocarbon-supplemented agar. A bioassay, utilising hydrocarbon-impregnated filter paper discs, was then used to examine the abilities of 17 selected isolates to catabolise three representative hydrocarbon molecules (hexadecane, phenanthrene and pristane) in different concentrations. In the same bioassay, the influence of a co-metabolite (glucose) on growth potential was also examined. Eight fungal species: Trichophyton sp.; Mucor sp.; Penicillium sp.; Graphium sp.; Acremoniwn sp.; Chaetomium sp.; Chrysosporium sp.; and an unidentified basidiomycete were then selected. Liquid batch cultures with a hydrocarbon mixture of hexadecane, phenanthrene, pristane and naphthalene facilitated quantitative analysis (HPLC) of the hydrocarbon catabolic abilities of the selected isolates. Ex situ bioremediation was evaluated at laboratory-scale by both bioaugmentation and biostimulation in soil microcosm trials. During the course of the study, total petroleum hydrocarbon (TPH) concentration (U.S. EPA Method 418.1) was used as a simple and inexpensive parameter to monitor hydrocarbon disappearance in response to soil treatments. Soil microbial activities were estimated by use of a fluorescein diacetate hydrolysis bioassay. This was found to be a reliable and sensitive method to measure the activity of respiring heterotrophs as compared with the unreliable data provided by plate counts. In the bioaugmentation trial, the eight selected isolates were individually used to inoculate (30% v/v) the contaminated soil. The highest rate of biodegradation (50.5% > than the non-sterile control) was effected by an Acremonium species after 50 days incubation (25°C). The second highest rate of biodegradation (47% > than the non-sterile control) was achieved with a soil treatment of sterile barley/beer waste only. Comparable rates of hydrocarbon degradation were achieved in simple biostimulation trials. Thus, due to its lower cost, biostimulation was the preferred remediation strategy and was selected for further laboratory investigation. Common agricultural or industrial lignocellulosic wastes such as: wood chips; straw; manure; beer brewery waste; mushroom compost; and spent mushroom substrate were used as soil treatments, either alone or in combination. The effect of the addition of a standard agricultural fertiliser was also examined. The highest level of biodegradation (54.4% > the non-sterile control) was recorded in a microcosm supplemented (40% v/v) with chicken manure. Finally, an ex situ bioremediation technique was examined in a pilot-scale field trial. Wood chips and chicken manure were co-composted with the contaminated soil in a low-cost, low-maintenance bioremediation system know as passive thermal bio venting. Extensive monitoring of the thermal environment within the biopile was made as an indirect measure of microbial activity. These data were then used to optimise the composting process. Three-dimensional graphical representations of the internal temperatures, in time and space, were constructed. From these graphs, it was determined that an inner core region of approximately 500 cm3 provided a realistic simulation of conditions within a full-scale biopile. During this trial a TPH reduction of 68% was achieved in 130 days. The findings of this research suggested that the utilisation of fungal catabolism is applicable to soils contaminated with a wide range of hydrocarbon contaminants. Passive thermal bioventing offers a bioremediation strategy which is highly suitable for South African conditions in terms of its low level of technological sophistication, low maintenance design and, most importantly, its relatively low cost. / Thesis (M.Sc.)-University of Natal, Pietermaritzburg, 1997.

Extractive processes can permanently and significantly reduce the volume, toxicity and mobility of contaminated materials at affected sites. Ethylenediaminetetraacetic acid (EDTA), N-2 (acetamido)iminodiacetic acid (ADA), pyridine2,6-dicarboxylic acid (PDA), and hydrochloric acid (HC1) were evaluated in batch studies for their ability to remove lead (Pb) and cadmium (Cd) from a Superfund soil. The extraction of Pb as a function of time was limited and the order of Pb removal was EDTA > ADA > PDA > HC1. Repeated extractions did not treat the soil below the Pb regulatory limit (1,000 mg/kg); however, the Pb remaining occurs in an immobile form. All extractants treated the soil below the proposed Cd regulatory limit (40 mg/kg) within 1 h. Lead recovery from solution was accomplished by hydroxide precipitation in the presence of excess calcium, and recovery at pH 11 was 70%, 98%, and 97% from the EDTA, ADA, and PDA complexes, respectively. / Department of Natural Resources and Environmental Management

Permeable reactive barriers are considered among the most promising technologies for contaminated soil and groundwater remediation. Zero-valent iron (ZVI), hydroxyapatite (HA), and organic compost, with (OM) and without (OMx) dextrose/sulfate were assessed in column studies for their ability to attenuate chromium (Cr) or cadmium (Cd). PVC columns were packed with the reactive media and Cr or Cd solutions were pumped through the columns at concentrations of 5, 50 and 200 mg/l. These media were also assessed for their abilities to attenuate Cr and Cd from a contaminated soil. The order of Cr removal was: ZVI > OMx > OM > HA. The ZVI treatment maintained a removal rate of > 95% throughout the study. All treatments used for Cd removal had a removal rate of 98% across all treatments. The ZVI was the only treatment capable of retaining any of the mobile soil Cr and Cd from the contaminated soil. / Department of Natural Resources and Environmental Management

While the extent of grasslands in Southern Ontario has been greatly reduced, urban and suburban areas provide numerous potential sites for their restoration. Grassland restoration in cities can provide ecological and cultural benefits, but soil conditions may be less than optimal for native species recovery. This thesis explores the use of soil amendments in order to address nutrient deficiency on old-field meadow restoration sites. Five treatments were tested, namely the addition of (1) nitrogenous fertilizer, (2) native legume species, (3) biochar, (4) a combination of the previous three, and (5) an unaltered control. Each treatment was replicated four times on two different test plots in Waterloo Region, Ontario, Canada (Huron Natural Area and Springbank Farm), for a total of 40 subplots. The experimental plots were tilled in the fall of 2011, a randomly selected treatment was applied to each subplot, and then all were seeded with a mix of five native meadow species (2 grass, 2 forb, 1 sedge). Soil samples were taken from each subplot both before treatment application and also at the end of the growing season in 2012, and tested for nutrient levels (N, P, K), pH and organic matter. Species richness, as well as soil temperature and moisture, were regularly monitored over the growing season. In the fall of 2012, above-ground vegetation was harvested to assess accrued biomass. In order to detect differences in means, results were tested using one-way and repeated-measures ANOVAs, where appropriate. Pearson???s product-moment correlations were also employed to test for linear dependence between variables. There were no significant differences between treatments in terms of soil nutrients or pH at either site. At Huron Natural Area, post-treatment biochar-treated subplots had slightly higher levels of organic matter than controls (p=0.095). Values for species richness, above-ground biomass, soil temperature and soil moisture did not vary significantly between treatments. Species richness at Huron Natural Area was positively correlated with 2011 N (r=0.42; p=0.07) and organic matter (r=0.52; p=0.02) levels, while at Springbank Farm it was negatively correlated with 2012 N levels (r=-0.67; p<0.001). Above-ground biomass at Huron Natural Area was positively correlated with 2011 and 2012 P levels (both r=0.52; p=0.02), while at Springbank Farm it was positively correlated with 2011 N, P, K and organic matter, and 2012 N, P and K (all r>0.44; p<0.05). At Huron Natural Area, above-ground biomass was negatively correlated with soil temperature (r=-0.64; p<0.0001) and positively correlated with soil moisture (r=0.38; p=0.1). This study uncovered a strong, but variable, relationship between N concentration and species richness in old-field meadows. Furthermore, productivity was tightly correlated with different soil nutrient concentrations at each study site. The results demonstrate the need for restoration approaches to address local soil conditions on order to be effective. To date, there have been very few studies on meadow restoration, particularly in North America. More, and longer-term, multivariate studies are needed in order to test the effectiveness of different techniques.