Factors Controlling the Geochemical Partitioning of Trace Metals in Estuarine Sediments

The geochemical partitioning of trace metals in sediments is of great importance in risk assessment and remedial investigation. Selected factors that may control the partitioning behavior of Cu, Pb and Zn in non-sulfidic, estuarine sediments were examined with the use of combined sorption curve—sequential extraction analysis. This approach, which has not been previously used to examine estuarine sediments, allowed determination of sorption parameters for Cu, Pb and Zn partitioning to individual geochemical fractions. Partitioning behavior in sulfidic sediments was also determined by sequentially extracting Cu, Pb, and Zn from synthetic sulfide minerals and from natural sediment and pure quartz sand after spiking with acid-volatile sulfide (AVS). Trace metal sorption to the “carbonate” fraction (pH 5, NaOAc extraction) increased with metal loading due to saturation of sorption sites associated with the “Fe-oxide” (NH₂OH·HCl extraction) and “organic” (H₂O₂ extraction) fractions in non-sulfidic sediments. Freundlich parameters describing sorption to the “Fe-oxide” and “organic” fractions were controlled by the sediment Fe-oxide and organic carbon content, respectively. Sequential extraction of Cu from pure CuS, AVS-spiked sediment and AVS-spiked quartz sand showed that AVS-bound Cu was quantitatively recovered in association with the “organic” fraction. However, some AVS-bound Pb and Zn were recovered by the NH₂OH·HCl step (which has been previously interpreted as “Fe-oxide” bound metals) in the sequential extraction procedure used in this study. This indicates that the sequential extraction of Pb and Zn in sulfidic sediments may lead to AVS-bound metals being mistaken as Fe-oxide bound species. Caution should therefore be exercised when interpreting sequential extraction results for Pb and Zn in anoxic sediments.     

Long-term assessment of natural attenuation: statistical approach on soils with aged PAH contamination

Natural attenuation processes valorization for PAH-contaminated soil remediation has gained increasing interest from site owners. A misunderstanding of this method and a small amount of data available does not encourage its development. However, monitored natural attenuation (MNA) offers a valuable, cheaper and environmentally friendly alternative to more classical options such as physico-chemical treatments (e.g., chemical oxidation, thermal desorption). The present work proposes the results obtained during a long-term natural attenuation assessment of historically contaminated industrial soils under real climatic conditions. This study was performed after a 10 year natural attenuation period on 60 off-ground lysimeters filled with contaminated soils from different former industrial sites (coking industry, manufactured gas plants) whose initial concentration of PAH varied between 380 and 2,077 mg kg^?1. The analysed parameters included leached water characterization, soil PAH concentrations, evaluation of vegetation cover quality and quantity. Results showed a good efficiency of the PAH dissipation and limited transfer of contaminants to the environment. It also highlighted the importance of the fine soil fractions in controlling PAH reactivity. PAH dissipation through water leaching was limited and did not present a significant risk for the environment. This PAH water concentration appeared however as a good indicator of overall dissipation rate, thereby illustrating the importance of pollutant availability in predicting its degradation potential.     

Methyl-tertiary Butyl Ether Natural Attenuation Case Studies

The occurrence of methyl-tertiary butyl ether (MtBE), a gasoline additive in ground water and surface water, is causing regulatory agencies, owner/operators, environmental professionals, and researchers to reevaluate remediation strategies at sites where gasoline containing this additive has been released. Over the last 5 to 10 years, monitored natural attenuation has been applied at petroleum hydrocarbon-impacted sites with increasing frequency. The efficacy of this remediation method for releases containing MtBE is now coming under increased scrutiny. The question of natural attenuation efficacy stems from uncertainty about MtBE biodegradability and behavior in the subsurface. Researchers and applied environmental scientists have completed and are continuing studies concerning MtBE biodegradability and behavior. This article briefly summarizes the history of MtBE, its physicochemical properties, its behavior in the environmental, and the applicability of monitored natural attenuation as a remediation tool. Case studies representing past and current research are then presented and followed by a brief discussion. Results from the documented research reviewed show that MtBE does biodegrade in the laboratory and at actual release sites. “Plumathon” studies document MtBE concentration and mass reduction and/or plume stabilization over time. MtBE concentrations in monitoring wells may show declining concentrations indicating natural attenuation. The data reviewed from past and current research suggest that natural attenuation may be the appropriate remediation strategy at some release sites. Yet, the data also indicate that care must be exercised in determining the efficacy of applying monitored natural attenuation at sites impacted with MtBE.     

Monitored Natural Attenuation of Explosives

Explosives are subject to several attenuation processes that potentially reduce concentrations in groundwater over time. Some of these processes are well defined, while others are poorly understood. The objective of the project was to optimize data collection and processing procedures for evaluation and implementation of monitored natural attenuation of explosives. After conducting experiments to optimize data quality, a protocol was established for quarterly monitoring of thirty wells over a 2-year period at a former waste disposal site. Microbial biomarkers and stable isotopes of nitrogen and carbon were explored as additional approaches to tracking attenuation processes. The project included a cone penetrometry sampling event to characterize site lithology and to obtain sample material for biomarker studies. A three-dimensional groundwater model was applied to conceptualize and predict future behavior of the contaminant plume. The groundwater monitoring data demonstrated declining concentrations of explosives over the 2 years. Biomarker data showed the potential for microbial degradation and provided an estimate of the degradation rate. Measuring stable isotopic fractions of nitrogen in TNT was a promising method of monitoring TNT attenuation. Overall, results of the demonstration suggest that monitored natural attenuation is a viable option that should be among the options considered for remediation of explosives-contaminated sites.     

SEQUENCE Visualization of Natural Attenuation Trends at Hill Air Force Base, Utah

For monitored natural attenuation to be considered as an acceptable remedial approach, the proponent must clearly document converging lines of evidence that illustrate the effectiveness of this measure. SEQUENCE, a visualization tool based on a modified radial diagram approach, is ideally suited for evaluating spatial and temporal trends that provide supporting evidence for the efficacy of monitored natural attenuation. SEQUENCE was applied to evaluate the natural attenuation of benzene, toluene, ethylbenzene, and total xylene (BTEX) concentrations observed in groundwater at Hill Air Force Base, Utah. SEQUENCE-BTEX maps provided an efficient means of documenting the declining BTEX concentrations downgradient from the source area. SE-QUENCE-Redox maps were used to facilitate a correlation between elevated BTEX concentrations; decreasing electron acceptor concentrations (oxygen, nitrate, and sulfate); and elevated metabolic byproduct concentrations (iron(II) and methane) providing a second line of evidence that suggests BTEX compounds are being destroyed through biodegradation processes downgradient from the source area. Site-specific guidelines for interpolating representative data sets for use with the SEQUENCE approach are discussed.     

The selectivity of a sequential extraction procedure for the determination of iron oxyhydroxides andiron sulfides in lake sediments

A popular sequential extraction procedure (Tessier et al. 1979) designed t o extract metals partitioned in various sediment phases, was evaluated for its selectivity. Amorphous FeOOH, FeS, and FeS₂ were added separately to natural lake sediments and sequentially extracted. The selectivity of the sequential procedure for the added solid phases was evaluated by determining the difference in the mass of Fe extracted from treated and control sediments. In the experiments where sulfide minerals were added, total S was measured in the residual solids in order to confirm selectivity of the method. Concentrations of total carbon remaining in the solid phase after each extraction step were also measured to determine the selectivity of the sequential procedure for carbon. The procedure was moderately selective for Fe added as FeOOH; a mean of 77 ± 12% (p < 0.05) of the Fe added was extracted in the step designed to reduce Fe-Mn oxyhydroxides. In experiments where FeS was added, a mean of 69 ± 11% (p < 0.05) of the Fe added as FeS was extracted in the fraction designed to oxidize sulfides and organic matter. Approximately 25% of the Fe added as FeS may have been extracted prematurely. Although less precise, total S analyses confirmed that much of the FeS was extracted in the oxidation step, yielding 104 ± 87% (p < 0.05) of the S added as FeS. The procedure was highly selective for FeS₂; 92 ± 14% (p < 0.05) of the Fe added as pyrite was extracted in the sulfide extraction step. Extraction of 80 ± 54% (p < 0.05) of S added as pyrite confirmed that FeS₂ were selectively extracted in the sulfide extraction step. Carbon in the sediments was also selectively extracted in the oxidation step (77 ± 2.4% of total C; p < 0.05). The applications and limitations of sequential extraction procedures as limnological research tools are discussed in light of our results. Request for offprints     

Effects of Chronic Waterborne and Dietary Metal Exposures on Gill Metal-Binding: Implications for the Biotic Ligand Model

The biotic ligand modeling (BLM) approach has gained recent widespread interest among the scientific and regulatory communities because of its potential for developing ambient water quality criteria (AWQC), which are site-specific, and in performing aquatic risk assessment for metals. Currently, BLMs are used for predicting acute toxicity (96?h LC50 for fish) in any defined water chemistry. The conceptual framework of the BLM has a strong physiological basis because it considers that toxicity of metals occurs due to the binding of free metal ions at the physiologically active sites of action (biotic ligand, e.g., fish gill) on the aquatic organism, which can be characterized by conditional binding constants (log K) and densities (B_max). At present, these models assume that only water chemistry variables such as competing cations (e.g., Na⁺, Ca²⁺, Mg²⁺, and H⁺), inorganic ligands (e.g., hydroxides, chlorides, carbonates), and organic ligands (dissolved organic matter) can influence the bioavailability of free metal ions and thereby the acute toxicity of metals. Current BLMs do not consider the effects of chronic history of the fish in modifying gill-metal binding characteristics and acute toxicity. Here, for Cu, Cd, and Zn, we review a number of recent studies on the rainbow trout that describe significant modifying effects of chronic acclimation to waterborne factors (hardness and chronic metal exposure) and dietary composition (metal and essential ion content) on gill metalbinding characteristics (on both log K and B_max) and on acute toxicity. We conclude that the properties of gill-metal interaction and toxicological sensitivity appear to be dynamic rather than fixed, with important implications for further development of both acute and chronic BLMs. Now that the initial framework of the BLM has been established, future research needs a more integrative approach with additional emphasis on the dynamic properties of the biotic ligand to make it a successful tool for ecological risk assessment of metals in the natural environment.     

Bioremediation of Diesel-contaminated Soil by Composting with Locally Generated Bulking Agents

Herein, we conducted a study toward understanding the impact of composting of the diesel-contaminated soil with some locally available bulking agents (rice husks (RHs), sawdust (SD), and wood chips (WCs)). In order to ascertain the effectiveness of petroleum degradation by the process assayed, we compared the protocols with monitored natural attenuation (MNA). The overall degradation pattern was modeled with non-linear regression by comparing the experimental data with first and second-order kinetic equations. At the end of the six-week study, the amount of total petroleum hydrocarbon removed from contaminated soil was 98.26 ± 1.33% (amendment with SD + RHs + WCs), 96.89 ± 1.20 (RHs amendment), 96.55 ± 1.29% (amendment with SD), 90.01 ± 0.22% (WCs amendment), and 85.02 ± 0.21% (MNA). The degradation of TPH trends followed a second-order kinetic model for all the four compost treatments while the MNA was found to follow a first-order (slower) degradation pattern. In general, the results of the parameter estimate showed that amendment with mixture of the three bulking agents was 1.08 (8%) slower (k₂ = (1.289 ± 0.16) × 10^?5 (g mg^?1 d^?1), r² = 0.991) than SD amendment alone (k₂ = (1.392 ± 0.14) × 10^?5 (g mg^?1 d^?1), r² = 0.995). However, the mixture of the bulking agents was found to be 1.67, 1.41, and 2.4 times faster than amendments with WCs, rice, and MNA, respectively. The phytotoxicity test revealed that all the compost treatments except WCs resulted in germination index of ≥80% after six weeks of bioremediation tests. The outcome of the current investigation confirms the effectiveness of bulking agents (especially when combined) in the supply of nutrients for the bioremediation of diesel-impacted soil.     

Solid-phase distribution of heavy metals as affected by single reagent extraction in dredged sediment derived surface soils

ABSTRACT

EDTA is useful to assess mobile metal pools in polluted soils and sediments. There is a need to enhance our understanding of the significance of metal fractions released. The impact of single reagent extraction with 0.05 mol L^?1 EDTA on the solid phase distribution of trace metals in surface soils sampled from confined dredged sediment disposal sites was investigated. Not extracted and EDTA extracted soils were subjected to sequential extraction to fractionate the total contents into: (1) easily exchangeable and carbonate bound fraction; (2) reducible fraction; (3) oxidisable fraction; and (4) residual fraction. With EDTA, significant portions of metals associated with the acid extractable and reducible fractions were released. The oxidisable and residual fractions remained unaffected for most of the investigated metals except for the organic matter associated metals (Cu and Pb). A decrease in the residual fraction after EDTA-extraction for Cu and Pb was attributed to artifacts of the sequential extraction procedure.     

Differentiating Natural and Anthropogenic Sources of Metals to the Environment

Since natural and anthropogenic sources can contribute to elevated levels of metals at remote and background sites, identifying the source of a metal is an important step in environmental risk assessment. Various source apportionment procedures are available to identify metal sources, and have been used extensively to determine sources in urban settings and to a lesser extent at remote sites. However, measuring metals at remote or background sites presents unique challenges with respect to experimental design. The state of the science in monitoring techniques and source apportionment procedures is discussed in terms of limitations and applicability to remote sites, and recommendations are made on maximizing information recovery through source apportionment procedures by incorporating appropriate experimental design.     

Bioremediation strategies for diesel and biodiesel in oxisol from southern Brazil

Leaks and spillages during the extraction, transport and storage of petroleum and its derivatives may result in environmental contamination. Biodiesel is an alternative energy source that can contribute to a reduction in environmental pollution. The aim of the present work was to evaluate biodegradation of diesel, biodiesel, and a 20% biodiesel-diesel mixture in oxisols from southern Brazil, using two bioremediation strategies: natural attenuation and bioaugmentation/biostimulation. Fuel biodegradation was monitored over 60 days by dehydrogenase activity, CO₂ evolution and gas chromatography. The bacterial inoculum employed for bioaugmentation/biostimulation consisted of Bacillus megaterium, Bacillus pumilus, Pseudomonas aeruginosa, and Stenotrophomonas maltophilia and PCR-DGGE using 16S RNAr primers showed that some members of this consortium survived in the soil after 60 days. The biodegradation of pure biodiesel was higher for bioaugmentation/biostimulation than for natural attenuation, suggesting that the addition of the microbial consortium, together with adjustment of the macronutrient ratio, increased biodiesel degradation. The results of dehydrogenase and respiratory activity, together with GC analysis, suggested that the presence of biodiesel may, by stimulating general microbial degradative metabolism, increase the biodegradation of petroleum diesel. The microbial community was altered by both treatments, with natural attenuation producing a lower diversity index than the amended soil. The bioaugmentation/biostimulation strategy was showed to have a high potential for cleaning up soils contaminated with diesel and biodiesel blends.     

Natural Attenuation Rate Clarifications: The True Picture is in the Details

The term “Natural Attenuation” (NA) has been defined as naturally occurring processes in soil and groundwater environments that act without human intervention to reduce the mass, toxicity, mobility, volume, or concentration of contaminants in those media. Monitored natural attenuation (MNA) protocols generally involve the collection of biogeochemical data from groundwater monitoring wells at sites. The data are correlated in time and space with the various chemicals of concern (COC's) to establish predominant biodegradation mechanisms. Modelers using the first-order decay expression typically use the rate coefficient as a calibration parameter and adjust it until the transport model results match field data. With this approach, uncertainties with a number of parameters (e.g., dispersion, sorption, biodegradation, etc.) are lumped together in a single calibration parameter. The problems associated with the lumped parameter approach are illustrated using two commonly used models, BIOSCREEN and Buscheck/Alcantar Analytical Solution, in a variety of practical examples. The natural attenuation decay rate estimated using the lumped parameter approach is distinguished from a biodegradation rate established by isolating processes and examining biodegradation lines of evidence. The half-life determined from empirical data using the lumped parameter approach is often mistakenly interchanged with a biodegradation half-life when it is an all-encompassing half-life based on the interaction of numerous processes. Isolation of the processes, as they are represented in the governing transport equation, and a rationale approach at parameter estimation to avoid the potential pitfalls of the all-inclusive “attenuation rate,” are provided. In closing, it is imperative to implement the following steps to dissern lumped process degradation rates from biodegradation half-lives: (a) be sure the rate/half-life processes are clarified as to what they encompass, (b) establish exactly how the rate/half-life was determined, (c) make certain other processes, such as dispersion, were estimated correctly, and (d) if the half-life is presented as a first-order biodegradation rate, examine the available lines of evidence to substantiate it.     

Challenges in Monitoring the Natural Attenuation of Spatially Variable Plumes

Monitored natural attenuation may be applied as a risk-based remediation strategy if it can be established that contaminants are or will be reduced to some acceptable level at or before a compliance point. Contaminant attenuation is often attributed to intrinsic biodegradation, which in some circumstances may occur only at the plume fringes where electron acceptors from the surrounding uncontaminated zones mix by dispersion and diffusion with the plume. However, due to the common spatial and temporal variability exhibited by many plumes, the centreline monitoring approaches advocated in many natural attenuation protocols may be unable to detect natural attenuation occurring primarily by fringe processes. Snapshot data from a multilevel sampling well transect across an MTBE plume at Vandenberg Air Force Base, CA, USA, illustrate the difficulty of centreline monitoring and the challenge of providing sufficient detail to detect attenuation processes that may be occurring primarily at plume fringes. In a study of a phenols plume in Wolverhampton, UK, high-resolution multilevel wells demonstrated that the key biodegradation processes were restricted spatially to the upper fringe of the plume and were rate-limited by transverse dispersion and diffusion of electron acceptors into the plume. Thus the overall extent of biodegradation was considerably less than suggested by a plume-scale analysis of total electron acceptor and contaminant budgets. These examples indicate that more robust and cost-effective MNA assessments can be obtained using monitoring strategies that focus on the location of key biodegradation processes.     

Multiple Stressor Differential Tolerances: Possible Implications at the Population Level

The probability of the most sensitive genotypes being eliminated from a population due to a contaminant pulse–genetic erosion–is negatively associated to the within-genotype variation. A sensitive genotype with a small phenotypic variation would be more prone to be lost–a critically sensitive genotype. Furthermore, natural populations inhabiting contaminated sites are usually exposed to several pollutants. Such co- or sequential exposure can have severe effects if at least some tolerant clonal lineages surviving one contaminant are sensitive to the others. Such an inverse relationship coupled with a low within-genotype variation potentially enhances genetic erosion. Accordingly, this study evaluated co-tolerance and the occurrence of clonal lineages critically sensitive to 48-hours lethal exposures of copper, zinc, cobalt, and chromium among eight clonal lineages of the cladocerans Daphnia longispina. Median lethal concentrations (LC₅₀) of each metal were found to have the potential to provoke genetic erosion. Pairwise comparisons of LC₅₀, from the eight clonal lineages, revealed neither negative nor positive correlations (r ≤ |0.56|; p ≥ 0.18), but inversely sensitive clonal lineages were found for all pairs of metals. Therefore, besides having the potential to eliminate critically sensitive clonal lineages in a first intermediately lethal pulse, all tested metals may provoke further losses of clonal lineages in an already genetically eroded population.     

Plasmid pU29, a vehicle for mutagenesis of the photosynthetic puf operon in Rhodopseudomonas capsulata

Plasmid pU21, which carries the reaction center and light-harvesting genes (puf operon) of Rhodopseudomonas capsulata, has been redesigned by site-specific mutagenesis. Five restriction sites have been removed and three unique restriction sites have been introduced into this 11,589-bp pBR322 derivative. The modifications divide the puf structural genes into four regions separated by five unique and nonmutagenic restriction sites. These four fragments have been subcloned into the M13-mp series of vectors to facilitate oligonucleotide-mediated site-specific mutagenesis experiments on the photosynthetic apparatus structural genes. The inserts can then be returned from the M13 replicative form to the redesigned pU21 derivative. The modified plasmid, pU29, greatly facilitates in vitro mutagenesis experiments since previously described techniques and screening procedures are more efficient with M13 derivatives carrying smaller inserts. Additionally, tandem homologous sequences (the reaction center L and M subunits) within the puf operon are now separated on different phage vectors, eliminating problems encountered in the targeting of mutagenic oligonucleotides to only one of the two homologous sites.     

The Importance of Nickel Phytoavailable Chemical Species Characterization in Soil for Phytoremediation Applicability

In this work Ni speciation in natural and spiked soils (with similar total concentration) was studied. Spiked soils were prepared by addition of NiSO₄.6H₂O to obtain concentration similar to the one in natural soils. In soils mobile species were determined with a simplified sequential extraction as follows: H₂O for water-soluble metal, KNO₃ for exchangeable species, DTPA for complexed/adsorbed species. Results show that in spiked soils the exchangeable and adsorbed Ni concentrations are considerably higher than in natural soils. A study of plant uptake was carried out in order to evaluate the relation between mobile species and phyto-availability. Alfalfa (Medicago sativa L.), even though it is not a hyperaccumulator, was selected for its tolerance to high metal concentrations in soil. Preliminary results show a very high correlation between Ni mobile species and Ni uptake by alfalfa. Significant differences were found between spiked and natural soils. In the latter, high levels of total Ni did not correspond to relevant uptake as in the case of spiked soil. Results stress the importance of evaluating preliminarly heavy metal speciation in soil before planning phytoremediation procedures.     

Site-specific gene integration technologies for crop improvement

Targeted integration of foreign genes into plant genomes is a much sought-after technology for engineering precise integration structures. Homologous recombination-mediated targeted integration into native genomic sites remained somewhat elusive until made possible by zinc finger nuclease-mediated double-stranded breaks. In the meantime, an alternative approach based on the use of site-specific recombination systems has been developed which enables integration into previously engineered genomic sites (site-specific integration). Follow-up studies have validated the efficacy of the site-specific integration technology in generating transgenic events with a predictable range and stability of expression through successive generations, which are critical features of reliable and practically useful transgenic lines. Any DNA delivery methods can be used for site-specific integration; however, best efficiency is mostly obtained with direct DNA delivery methods such as particle bombardment. Although site-specific integration approach provides unique advantages for producing transgenic plants, it is still not a commonly used method. The present article discusses barriers and solutions for making it readily available to both academic research and applicative use.     

Characterization of a single glycosylated asparagine site on a glycopeptide using solid-phase Edman degradation

The characterization of site-specific glycosylation is traditionally dependent on the availability of suitable proteolytic cleavage sites between each glycosylated residue, so that peptides containing individual glycosylation sites are recovered. In the case of heavily glycosylated domains such as theO-glycosylated mucins, which have no available protease sites, this approach is not possible. Here we introduce a new method to gain site-specific compositional data on the oligosaccharides attached to a single amino acid. Using a model glycopeptide from a mutant human albumin Casebrook, glycosylated PTH-Asn was recovered after sequential solid-phase Edman degradation, subjected to acid hydrolysis and the sugars were identified by high performance anion exchange chromatography with pulsed amperometric detection. The PTH-Asn(Sac) derivative was further characterized by ionspray mass spectrometry. Comparison between an endoproteinase Glu-C glycopeptide and a tryptic glycopeptide showed that the oligosaccharide attached to Asn494 was stable after at least 10 cycles of Edman degradation.     

Rapid intrinsic biodegradation of benzene, toluene, and xylenes at the boundary of a gasoline-contaminated plume under natural attenuation

A groundwater plume contaminated with gasoline constituents [mainly benzene, toluene, and xylenes (BTX)] had been treated by pumping and aeration for approximately 10 years, and the treatment strategy was recently changed to monitored natural attenuation (MNA). To gain information on the feasibility of using MNA to control the spread of BTX, chemical and microbiological parameters in groundwater samples obtained inside and outside the contaminated plume were measured over the course of 73 weeks. The depletion of electron acceptors (i.e., dissolved oxygen, nitrate, and sulfate) and increase of soluble iron were observed in the contaminated zone. Laboratory incubation tests revealed that groundwater obtained immediately outside the contaminated zone (the boundary zone) exhibited much higher potential for BTX degradation than those in the contaminated zone and in uncontaminated background zones. The boundary zone was a former contaminated area where BTX were no longer detected. Denaturing gradient gel electrophoresis (DGGE) analysis of polymerase chain reaction (PCR)-amplified bacterial 16S rRNA gene fragments revealed that DGGE profiles for groundwater samples obtained from the contaminated zone were clustered together and distinct from those from uncontaminated zones. In addition, unique bacterial rRNA types were observed in the boundary zone. These results indicate that the boundary zone in the contaminant plumes served as a natural barrier for preventing the BTX contamination from spreading out.     

Effect of plants on the bioavailability of metals and other chemical properties of biosolids in a column study

The effects of metal-accumulating plants (Salix x reichardtii and Populus balsamifera) on the chemical properties and dynamics of metals in biosolids were investigated using different techniques including diffusive gradients in thin films (DGT), sequential extraction procedures and partitioning coefficient (K(d)). Plants could effectively extract Cd, Ni, and Zn and decreased dissolved organic carbon (DOC). The presence of plants increased the potential bioavailability of these metals, as assessed by an increase in the ratio of metal measured by DGT and metals in the solution. The plants affected the Cd, Ni, and Zn pools (soluble/exchangeable; Fe/Mn oxide and organic matter bound) characterised by sequential extraction and K(d) but did not reduce the total metals in either substrate. However, plants had no effect on Cu, presumably because of the effective buffering of available Cu by organic matter in both solution and solid phases. A high density of plant roots was associated with increased leaching of metals.