Enhanced phytoextraction of Cu, Pb, Zn and Cd with EDTA and EDDS

Chemically enhanced phytoextraction has been proposed as an effective approach to removing heavy metals from contaminated soil through the use of high biomass plants. Using pot experiments, the effects of the application of EDTA, EDDS and citric acid on the uptake of Cu, Pb, Zn and Cd by corn (Zea mays L. cv. Nongda 108) and bean (Phaseolus vulgaris L. white bean) plants were studied. The results showed that EDDS was more effective than EDTA at increasing the concentration of Cu in corn and beans. The application of 5 mmol kg-1 soil EDDS to soil significantly increased concentrations of Cu in shoots, with maximum levels of 2060 and 5130 mg kg-1 DW in corn and beans, respectively, which were 45- and 135-fold higher than that in the corresponding control plants to which chelate had not been applied. Concentrations of Zn in shoots were also higher in the plants treated with EDDS than in those treated with EDTA. For Pb and Cd, EDDS was less effective than EDTA. The maximum Cu phytoextraction was found with the EDDS treatment. The application of EDTA and EDDS also significantly increased the shoot-to-root ratios of the concentrations of Cu, Pb, Zn and Cd in both plant species. The results of metal extraction with chelates showed that EDDS was more efficient at solubilizing Cu and Zn than EDTA, and that EDTA was better at solubilizing Pb and Cd than EDDS.     

Biodegradation and speciation of residual SS-ethylenediaminedisuccinic acid (EDDS) in soil solution left after soil washing

This paper aims to investigate the degradation and speciation of EDDS-complexes (SS-ethylenediaminedisuccinic acid) in soil following soil washing. The changes in soil solution metal and EDDS concentrations were investigated for three polluted soils. EDDS was degraded after a lag phase of 7-11 days with a half-life of 4.18-5.60 days. No influence of EDDS-speciation on the reaction was observed. The decrease in EDDS resulted in a corresponding decrease in solubilized metals. Changes in EDDS speciation can be related to (1) initial composition of the soil, (2) temporarily anoxic conditions in the soil slurry after soil washing, (3) exchange of EDDS complexes with Cu even in soils without elevated Cu and (4) formation of NiEDDS. Dissolved organic matter is important for metal speciation at low EDDS concentrations. Our results show that even in polluted soils EDDS is degraded from a level of several hundred micromoles to below 1 microM within 50 days.     

柠檬酸浸出土壤中铜、锌的优化设计

以某工业废弃地土壤为供试材料,采用批次实验方法,研究了柠檬酸对重金属Cu、Zn的浸出效果。并在单因素实验基础上,探讨不同pH值、柠檬酸浓度、固液比和浸提时间等对浸出效果的影响,进而对实验条件进行正交优化组合,确定优化工艺参数为：pH值为3.2,柠檬酸浓度为0.4 mol/L,壤土固液比为1/10,土壤中Cu、Zn接近平衡的时间分别为1 h和1.5 h。结果表明,在此条件下,对这2种金属来说,Cu较Zn易释放,Cu、Zn的最大浸出率分别为74.8%和65.3%。柠檬酸浸提前后,土壤中有机质含量下降范围为0.61%~3.16%。单从土壤有机质含量来看,柠檬酸对土壤质量影响较小,具有较好的应用前景。     

Simultaneous extraction of Cr(VI) and Cu(II) from humic acid with new synthesized EDTA derivatives

Soil washing is one of the few permanent treatment alternatives for removing metal contaminants. Ethylenediaminetetraacetic acid and its salts (EDTA) is very effective at removing cationic metals and has been utilized globally. However it is ineffective for anionic metal contaminants or metals bound to soil organic matter. The simultaneous removal of cationic and anionic metal contaminants by soil washing is difficult due to differences in their properties. The present study evaluated the potential of a washing process using two synthesized EDTA-derivatives, C₆HEDTA (2,2′-((2-((carboxymethyl)(2-(hexanoyloxy)ethyl)amino)ethyl)azanediyl)diacetic acid) and C₁₂HEDTA (2,2′-((2-((carboxymethyl) (2-(dodecanoyloxy)ethyl)amino)ethyl)azanediyl)diacetic acid), which consist of a hydrophilic polycarboxylic moiety and a hydrophobic moiety with a monoalkyl ester group. A series of equilibrium batch experiments at room temperature were conducted to investigate the efficacy of C₆HEDTA and C₁₂HEDTA as extractants for both oxyanion Cr(VI) and cationic Cu(II). Results showed that either C₆HEDTA or C₁₂HEDTA can extract both Cr(VI) and Cu(II) from humic acid simultaneously. However, C₆HEDTA was less effective for Cr(VI) probably because it has no surface activities to increase solubility of humic acid, like C₁₂HEDTA. Extraction of Cr(VI) was mainly attributed to the decreased surface tension and enhanced solubility of organic matter. Extraction of Cu(II) was attributed to both the Cu(II) chelation and enhanced solubility of humic acid. It was demonstrated that the hydrophilic polycarboxylic moiety of C₁₂HEDTA chelates cations while the monoalkyl ester group produces surface active properties that enhance the solubility of humic acid.     

The use of NTA and EDDS for enhanced phytoextraction of metals from a multiply contaminated soil by Brassica carinata

The potential of nine different species to grow in the presence of metals (As, Cd, Cu, Pb and Zn) and to accumulate them in the shoots was assessed for each metal separately by germination and root length tests, and successively by hydroponic experiments. Of the nine species tested, Brassica carinata was the species that accumulated the highest amounts of metals in shoots without suffering a significant biomass reduction. To further evaluate the potential of B. carinata for chelant-enhanced phytoextraction of a natural, multiply metal-polluted soil (As, Cd, Cu, Pb and Zn), both hydroponic and pot experiments were carried out with nitrilotriacetic acid (NTA) or (S,S)-ethylenediamine disuccinic acid (EDDS) as complexing agents. The hydroponic study with solutions containing the five metals together showed that accumulation of Cd, Cu, Pb and Zn in shoots was higher following EDDS addition compared to NTA. EDDS was more effective than NTA in desorbing Cu, Pb and Zn from the soil, whereas As and Cd were poorly extracted. B. carinata plants were grown for 4 weeks in the multiply metal-contaminated soil and then the soil was amended with 5 mmol kg(-1) NTA or EDDS. All plants were harvested 1 week after amendment. In comparison to NTA, EDDS was more effective in enhancing the concentrations of Cu, Pb and Zn in B. carinata shoots (2- to 4-fold increase compared to the control). One week after chelant addition, the DTPA-extractable metal concentrations in the polluted soil were lower in the EDDS treatment in comparison with the NTA amendment. Even though B. carinata showed a reduced growth and a relatively low metal uptake, it demonstrated the ability to survive and tolerate the presence of more metals simultaneously.     

Significance of metal exchange in EDDS-flushing column experiments

Chelating agents have been widely studied for extracting heavy metals from contaminated soils, and the effectiveness of EDDS ([S,S]-ethylene-diamine-disuccinic acid) has aroused extensive attention because of its biodegradability in the natural environment. However, in the course of EDDS-flushing, metal exchange of newly extracted metal-EDDS complexes with other sorbed metals and mineral cations may result in metal re-adsorption on the soil surfaces. Therefore, this study investigated the relative significance of metal exchange under different travel distances of chelant complexes, characteristics of soil contamination, and solution pH in the column experiments. As a result of metal exchange, the elution of Zn and Pb was retarded and the cumulative extraction was lower than those of Ni and Cu, especially over a longer travel distance. Compared with the field-contaminated soils, the effects of metal exchange were even more substantial in the artificially contaminated soil because of a greater amount of extractable metals and a larger proportion of weakly bound fractions. By contrast, metal exchange was insignificant at pH 8, probably due to less adsorption of metal-EDDS complexes. These findings highlight the conditions under which metal exchange of metal-EDDS complexes and the resulting impacts are more significant during EDDS-flushing.     

Ryegrass uptake of soil Cu/Zn induced by EDTA/EDDS together with a vertical direct-current electrical field

This paper summarizes a study on the application of vertical electric direct-current (DC) to control the migration of metal complexes in soil columns when EDTA/[S,S]-ethylenediaminedisuccinic acid (EDDS) is used to enhance ryegrass uptake of Cu/Zn from contaminated soil. The results show that application of EDTA/EDDS significantly increased ryegrass uptake of Cu/Zn when compared with no EDTA/EDDS application and the Cu/Zn concentrations in all soil solutions sampled at three different levels of the soil column, i.e. 15, 30 and 50 cm. Application of vertical DC electrical field (1.0 V cm(-1)) through top anode (close to the surface) and bottom cathode caused redistribution of Cu/Zn concentrations. Copper/Zn concentrations significantly decreased in soil pore fluid that were sampled in the bottom sections of the column, suggesting an efficient control of the leaching risk of the Cu/Zn complexes when the vertical electrical field is applied. The shoot Cu concentration of ryegrass in the treatments with EDTA/EDDS and electrical field together was 0.46/0.61 times higher than that in treatments without electrical fields.     

Effect of composting on characterization and leaching of copper, manganese, and zinc from swine manure.

This research was conducted to study the influence of composting on the concentrations, water solubility, and phase association of Cu, Mn, and Zn from swine (Sus domesticus) feces. Composting of separated swine manure was performed in two piles for 122 days. The metal concentrations increased rapidly during the first 49 days and leveled off thereafter. All metal concentrations increased approximately 2.7-fold in the final compost due to decomposition of organic matter. A sequential extraction protocol was used to evaluate the humification process and partition metals into water-soluble, exchangeable, organically complexed, organically bound, solid particulate, and residual fractions. Temporal changes in the water-soluble fractions of Cu, Mn, and Zn were reflected by water-soluble organic C concentrations, which rapidly increased to a maximum at Day 18 and declined thereafter. An increase in the humic acid/fulvic acid ratio in Na4P2O7 or NaOH extracts at various stages of composting represented the humification process. During composting, the major portions of Cu, Mn, and Zn were in the organically-bound, solid particulate, and organically complexed fractions, respectively. Metal distributions in different chemical fractions were generally independent of composting age and, thus, independent of respective total metal concentrations in the composts.     

Potential availability of heavy metals to phytoextraction from contaminated soils induced by exogenous humic substances

Effective phytoremediation of soils contaminated by heavy metals depends on their availability to plant uptake that, in turn, may be influenced by either the existing soil humus or an exogenous humic matter. We amended an organic and a mineral soil with an exogenous humic acid (HA) in order to enhance the soil organic carbon (SOC) content by 1% and 2%. The treated soils were further enriched with heavy metals (Cu, Pb, Cd, Zn, Ni) to a concentration of 0, 10, 20, and 40 microg/g for each metal and allowed to age at room temperature for 1 and 2 months. After each period, they were extracted for readily soluble and exchangeable (2.5% acetic acid), plant-available (DTPA, Diethylentriaminepentaacetic acid), and occluded (1 N HNO(3)) metal species. Addition of HA generally reduced the extractability of the soluble and exchangeable forms of metals. This effect was directly related to the amount of added HA and increased with ageing time. Conversely, the potentially plant-available metals extracted with DTPA were generally larger with increasing additions of exogenous HA solutions. This was attributed to the formation of metal-humic complexes, which ensured a temporary bioavailability of metals and prevented their rapid transformation into insoluble species. Extractions with 1 N HNO(3) further indicated that the added metals were present in complexes with HA. The observed effects appeared to also depend on the amount of native SOC and its structural changes with ageing. The results suggest that soil amendments with exogenous humic matter may accelerate the phytoremediation of heavy metals from contaminated soil, while concomitantly prevent their environmental mobility.     

Sequential extraction of heavy metals during composting of sewage sludge

The major limitation of soil application of sewage sludge compost is the total heavy metal contents and their bioavailability to the soil-plant system. This study was conducted to determine the heavy metal speciation and the influence of changing the physico-chemical properties of the medium in the course of composting on the concentrations, bioavailability or chemical forms of Cu, Zn, Pb and Ni in sewage sludge. Principal physical and chemical properties and FTIR spectroscopical characterization of sludge compost during treatment show the stability and maturity of end product. The total metal contents in the final compost were much lower than the limit values of composts to be used as good soil fertilizer. Furthermore, it was observed by using a sequential extraction procedure in sludge compost at different steps of treatment, that a large proportion of the heavy metals were associated to the residual fraction (70-80%) and more resistant fractions to extraction X-NaOH, X-EDTA, X-HNO3 (12-29%). Less than 2% of metals bound to bioavailable fractions X-(KNO3+H2O). Heavy metal distribution and bioavailability show some changes during composting depending on the metal itself and the physico-chemical properties of the medium. Bioavailable fractions of all elements tend to decrease except Ni-H2O. Zn and mainly Cu present more affinity to organic and carbonate fractions. In contrast, Pb is usually preferentially bound to sulfide forms X-HNO3. Nickel shows a significant decrease of organic form. Significant degrees of correlation were found between heavy metal fractions and changes of some selected variables (e.g. pH, ash, organic matter, humic substance) during the course of composting. Mobile fractions of metals are poorly predictable from the total content. The R2 value was significantly increased by the inclusion of other variables such as the amount of organic matter (OM) and pH.     

Metal extractability patterns to evaluate (potentially) mobile fractions in periurban calcareous agricultural soils in the Mediterranean area—analytical and mineralogical approaches

A set of periurban calcareous agricultural Mediterranean soils was spiked with a mixture of Cd, Cu, Pb and Zn at two levels within the limit values proposed by current European legislation, incubated for up to 12 months, and subjected to various one-step extraction procedures to estimate mobile (neutral salts) and potentially mobile metal fractions (complexing and acidic extraction methods). The results obtained were used to study metal extractability patterns according to the soil characteristics. The analytical data were coupled with mineralogical investigations and speciation modelling using the Visual Minteq model. The formation of soluble metal-complexes in the complexing extracts (predicted by the Visual Minteq calculations) led to the highest extraction efficiency with complexing extractants. Metal extractability patterns were related to both content and composition of carbonate, organic matter, Fe oxide and clay fractions. Potentially mobile metal fractions were mainly affected by the finest soil fractions (recalcitrant organic matter, active lime and clay minerals). In the case of Pb, scarce correlations between extractable Pb and soil constituents were obtained which was attributed to high Pb retention due to the formation of 4PbCO₃·3PbO (corroborated by X-ray diffraction). In summary, the high metal proportion extracted with complexing agents highlighted the high but finite capacity to store potentially mobilizable metals and the possible vulnerability of these soils against environmental impact from metal accumulation.     

Metal extraction by Arabidopsis halleri grown on an unpolluted soil amended with various metal-bearing solids: a pot experiment.

Most studies dealing with phytoremediation have considered metal extraction efficiency in relation to metal concentration of bulk soil samples or metal concentration of the soil solution. However, little is known about the effect of various metal-bearing solids on plant growth and metal extraction of hyperaccumulators. In this study, we investigated the ability of Arabidopsis halleri to grow and extract metals from different substrates consisting in an unpolluted soil amended with various metal-bearing solids collected in soils around a Zn smelter complex. The metal-bearing solids used as amendments were: fresh and decomposing organic residues in the soil, a soil clay fraction and two waste slags. Pure mono-metallic salt (ZnSO4) was also used. Two series of substrates were produced, one moderately polluted, and the other highly polluted. An additional substrate was formed by the unamended soil, and used as an unpolluted control. Zn, Cd, Cu, and Pb were measured in the substrates, and in the roots and shoots of A. halleri. The dry matter yield of A. halleri was shown not to depend on the nature of the metal-bearing solid used, except when Cu-toxicity was suspected. On highly-polluted substrates, Zn extraction by A. halleri depended on the nature of metal-bearing solids used, showing the following trend: pure mono-metallic salt > waste slags and soil clay fraction > fresh and decomposing organic matter. We explained these differences by the high solubility of Zn in the mono-metallic salt, whereas in the mineral metal-bearing solids and in both fresh and decomposing organic matter, Zn release required mineral weathering or organic matter mineralization, respectively. This work clearly showed that phytoremediation studies have to consider the nature of metal-bearing solids in contaminated soils to better predict the efficiency of plant extraction.     

The solid-solution partitioning of heavy metals (Cu,Zn, Cd,Pb) in upland soils of England and Wales

Ninety-eight surface soils were sampled from the uplands of England and Wales, and analysed for loss-on-ignition (LOI), and total and dissolved base cations, Al, Fe, and trace heavy metals (Cu, Zn, Cd, Pb). The samples covered wide ranges of pH (3.4-8.3) and LOI (9-98%). Soil metal contents measured by extraction with 0.43 mol l-1 HNO3 and 0.1 mol l-1 EDTA were very similar, and generally lower than values obtained by extraction with a mixture of concentrated nitric and perchloric acids. Total heavy metal concentrations in soil solution depend positively upon soil metal content and [DOC], and negatively upon pH and LOI, values of r2 ranging from 0.39 (Cu) to 0.81 (Pb). Stronger correlations (r2=0.76-0.95) were obtained by multiple regression analysis involving free metal ion (Cu2+, Zn2+, Cd2+, Pb2+) concentrations calculated with the equilibrium speciation model WHAM/Model VI. The free metal ion concentrations depend positively upon MHNO3 and negatively upon pH and LOI. The data were also analysed by using WHAM/Model VI to describe solid-solution interactions as well as solution speciation; this involved calibrating each soil sample by adjusting the content of "active" humic matter to match the observed soil pH. The calibrated model provided fair predictions of total heavy metal concentrations in soil solution, and predicted free metal ion concentrations were in reasonable agreement with the values obtained from solution-only speciation calculations.     

Soil solution dynamics of Cu and Zn in a Cu- and Zn-polluted soil as influenced by gamma-irradiation and Cu-Zn interaction

A pot experiment was conducted to study soil solution dynamics of Cu and Zn in a Cu/Zn-polluted soil as influenced by gamma-irradiation and Cu-Zn interaction. A slightly acid sandy loam was amended with Cu and Zn (as nitrates) either singly or in combination (100 mg Cu and 150 mg Zn kg(-1) soil) and was then gamma-irradiated (10 kGy). Unamended and unirradiated controls were included, and spring barley (Hordeum vulgare L. cv. Forrester) was grown for 50 days. Soil solution samples obtained using soil moisture samplers immediately before transplantation and every ten days thereafter were used directly for determination of Cu, Zn, pH and absorbance at 360 nm (A360). Cu and Zn concentrations in the solution of metal-polluted soil changed with time and were affected by gamma-irradiation and metal interaction. gamma-Irradiation raised soil solution Cu substantially but generally decreased soil solution Zn. These trends were consistent with increased dissolved organic matter (A360) and solution pH after gamma-irradiation. Combined addition of Cu and Zn usually gave higher soil solution concentrations of Cu or Zn compared with single addition of Cu or Zn in gamma-irradiated and non-irradiated soils, indicating an interaction between Cu and Zn. Cu would have been organically complexed and consequently maintained a relatively high concentration in the soil solution under higher pH conditions. Zn tends to occur mainly as free ion forms in the soil solution and is therefore sensitive to changes in pH. The extent to which gamma-irradiation and metal interaction affected solubility and bioavailability of Cu and Zn was a function of time during plant growth. Studies on soil solution metal dynamics provide very useful information for understanding metal mobility and bioavailability.     

Comparison of EDTA and EDDS as potential soil amendments for enhanced phytoextraction of heavy metals

Phytoextraction has been proposed as an alternative remediation technology for soils polluted with heavy metals or radionuclides, but is generally conceived as too slow working. Enhancing the accumulation of trace pollutants in harvestable plant tissues is a prerequisite for the technology to be practically applicable. The chelating aminopolycarboxylic acid, ethylene diamine tetraacetate (EDTA), has been found to enhance shoot accumulation of heavy metals. However, the use of EDTA in phytoextraction may not be suitable due to its high environmental persistence, which may lead to groundwater contamination. This paper aims to assess whether ethylene diamine disuccinate (EDDS), a biodegradable chelator, can be used for enhanced phytoextraction purposes. A laboratory experiment was conducted to examine mobilisation of Cd, Cu, Cr, Ni, Pb and Zn into the soil solution upon application of EDTA or EDDS. The longevity of the induced mobilisation was monitored for a period of 40 days after application. Estimated effect half lives ranged between 3.8 and 7.5 days for EDDS, depending on the applied dose. The minimum observed effect half life of EDTA was 36 days, while for the highest applied dose no decrease was observed throughout the 40 day period of the mobilisation experiment. Performance of EDTA and EDDS for phytoextraction was evaluated by application to Helianthus annuus. Two other potential chelators, known for their biodegradability in comparison to EDTA, were tested in the plant experiment: nitrilo acetic acid (NTA) and citric acid. Uptake of heavy metals was higher in EDDS-treated pots than in EDTA-treated pots. The effects were still considered insufficiently high to consider efficient remediation. This may be partly due to the choice of timing for application of the soil amendment. Fixing the time of application at an earlier point before harvest may yield better results. NTA and citric acid induced no significant effects on heavy metal uptake.     

Monometal and competitive adsorption of heavy metals by sewage sludge-amended soil

Sewage sludge-amended soils may alter their ability to adsorb heavy metals over time, due to the decomposition of sludge-borne organic matter. Thus, we studied Cd, Ni, and Zn adsorption by a sewage sludge-amended soil (Typic Xerofluvent) before and after one-year incubation in both monometal and competitive systems. In the monometal system, the order of decreasing sorption was Zn>Cd>Ni. Competition significantly reduced metal K(d), especially that of Cd which decreased by nearly 50%. Over the course of the incubation there was a 31% reduction of soil organic matter content. At the same time, in competitive systems Cd K(d) significantly decreased, while Zn K(d) significantly increased, and Ni K(d) remained unaffected. This study shows that sewage sludge-amended soils may change in their ability to sorb heavy metals over time at high metal concentrations. The data suggest that Cd is likely to be of most environmental significance in such soils, since it exhibited decreased sorption under competitive conditions and as the organic matter content of the soil was reduced. The potential for long-term release of metals should be considered in the risk assessment associated with sewage sludge addition to soils, particularly in climates where degradation of organic matter is likely to be enhanced.     

The influence of EDDS on the uptake of heavy metals in hydroponically grown sunflowers

Phytoextraction is an environmentally friendly in situ technique for cleaning up metal contaminated land. Unfortunately, efficient metal uptake by remediation plants is often limited by low phytoavailability of the targeted metals. Chelant assisted phytoextraction has been proposed to improve the efficiency of phytoextraction. Phytoremediation involves several subsequent steps: transfer of metals from the bulk soil to the root surfaces, uptake into the roots and translocation to the shoots. Nutrient solution experiments address the latter two steps. In this context we investigated the influence of the biodegradable chelating agent SS-EDDS on uptake of essential (Cu and Zn) and non-essential (Pb) metals by sunflowers from nutrient solution. EDDS was detected in shoots and xylem sap for the first time, proving that it is taken up into the above ground biomass of plants. The essential metals Cu and Zn were decreased in shoots in the presence of EDDS whereas uptake of the non-essential Pb was enhanced. We suggest that in the presence of EDDS all three metals were taken up by the non-selective apoplastic pathway as the EDDS complexes, whereas in the absence of EDDS essential metal uptake was primarily selective along the symplastic pathway. This shows that synthetic chelating agents do not necessarily increase uptake of heavy metals, when soluble concentrations are equal in the presence and absence of chelates.     

Plant accumulation of potentially toxic elements in sewage sludge as affected by soil organic matter level and mycorrhizal fungi.

Leek (Allium ameloprasum) was grown in pot trials in two clay loams of contrasting organic contents, with and without indigenous mycorrhizal propagules. Sewage sludges containing varying levels of Cd, Cu and Zn were added. Extractable soil metals, plant growth, major nutrient content and accumulation of metals, and soil microbial indices were investigated. The aim was to establish whether soil organic content and mycorrhizal status affected plant and microbial exposure to these metals. Extractable metals were higher and responses to inputs more pronounced in the arable, lower organic matter soil, although only Cd showed a soil difference in the CaCl2 fraction. There were no metal toxic effects on plants and some evidence to suggest that they promoted growth. Uptake of each metal was higher in the larger plants of the grassland, higher organic matter soil. Inoculation with arbuscular mycorrhizal fungi increased root Cd and Zn concentrations. With the exception of Cd (roots) and Zn (shoots), higher inputs of sludge metals did not increase plant metals. Zn and Cu, but not Cd, concentrations were higher in roots than in shoots.     

Fractionation of heavy metals and distribution of organic carbon in two contaminated soils amended with humic acids

The effects of humic acids (HAs) extracted from two different organic materials on the distribution of heavy metals and on organic-C mineralisation in two contaminated soils were studied in incubation experiments. Humic acids isolated from a mature compost (HAC) and a commercial Spaghnum peat (HAP) were added to an acid soil (pH 3.4; 966 mg kg(-1) Zn and 9,229 mg kg(-1) Pb as main contaminants) and to a calcareous soil (pH 7.7; 2,602 mg kg(-1) Zn and 1,572 mg kg(-1) Pb as main contaminants) at a rate of 1.1g organic-C added per 100g soil. The mineralisation of organic-C was determined by the CO(2) released during the experiment. After 2, 8 and 28 weeks of incubation the heavy metals of the soils were fractionated by a sequential extraction procedure. After 28 weeks of incubation, the mineralisation of the organic-C added was rather low in the soils studied (<8% of TOC in the acid soil; <10% of TOC in the calcareous soil). Both humic acids caused significant Zn and Pb immobilisation (increased proportion of the residual fraction, extractable only with aqua regia) in the acid soil, while Cu and Fe were slightly mobilised (increased concentrations extractable with 0.1M CaCl(2) and/or 0.5M NaOH). In the calcareous soil there were lesser effects, and at the end of the experiment only the fraction mainly related to carbonates (EDTA-extractable) was significantly increased for Zn and decreased for Fe in the humic acids treated samples. However, HA-metal interactions provoked the flocculation of these substances, as suggested by the association of the humic acids with the sand fraction of the soil. These results indicate that humic acid-rich materials can be useful amendments for soil remediation involving stabilisation, although a concomitant slight mobilisation of Zn, Pb and Cu can be provoked in acid soils.