Long-term efficiency of soil stabilization with apatite and Slovakite: the impact of two earthworm species (Lumbricus terrestris and Dendrobaena veneta) on lead bioaccessibility and soil functioning.

Remediation soil is exposed to various environmental factors over time that can affect the final success of the operation. In the present study, we assessed Pb bioaccessibility and microbial activity in industrially polluted soil (Arnoldstein, Austria) stabilized with 5% (w/w) of Slovakite and 5% (w/w) of apatite soil after exposure to two earthworm species, Lumbricus terrestris and Dendrobaena veneta, used as model environmental biotic soil factors. Stabilization resulted in reduced Pb bioaccessibility, as assessed with one-step extraction tests and six-step sequential extraction, and improved soil functioning, mirrored in reduced ß-glucosidase activity in soil. Both earthworm species increased Pb bioaccessibility, thus decreasing the initial stabilization efficacy and indicating the importance of considering the long-term fate of remediated soil. The earthworm species had different effects on soil enzyme activity, which can be attributed to species-specific microbial populations in earthworm gut acting on the ingested soil.

Immobilization of potentially toxic metals using different soil amendments.

The in situ stabilization of potentially toxic metals (PTMs), using various easily available amendments, is a cost-effective remediation method for contaminated soils. In the present study, we investigated the effectiveness of apatite and a commercial mixture of dolomite, diatomite, smectite basaltic tuff, bentonite, alginite and zeolite (Slovakite) on Pb, Zn, Cu and Cd stabilization by means of decreasing their bioavailability in contaminated soil from an old lead and zinc smelter site in Arnoldstein, Austria. We also investigated the impact of 5% (w/w) apatite and Slovakite applications on soil functionality and quality, as assessed by glucose-induced soil respiration, dehydrogenase, acid and alkaline phosphatase and ß-glucosidase activity. Both amendments resulted in increased soil pH and decreased PTM potential bioavailability assessed by diethylenetriamine pentaacetic acid extraction and by sequential extractions in the water-soluble and exchangeable fractions. The efficiency of stabilization was reflected in the soil respiration rate and in enzymatic activity. The ß-glucosidase activity assay was the most responsive of them.

Influence of ozonation on extractability of Pb and Zn from contaminated soils.

The effect of soil ozonation on Pb and Zn extraction with EDTA, bioavailability (Ruby's Physiologically Based Extraction Test, PBET) and mobility (Toxicity Characteristic Leaching Procedure, TCLP) of Pb was studied on contaminated soils taken from 7 different locations in the Mezica Valley, Slovenia. EDTA extraction (40 mmol kg(-1)) removed from 27.4+/-1.5% to 64.8+/-1.5% of Pb, and from 1.9+/-0.2% to 22.4+/-2.0% of Zn from tested soils, and significantly reduced soil Pb bioavailability (PBET) and mobility (TCLP). Pretreatment of tested soils with ozone before EDTA extraction enhanced EDTA extractability of Pb for 11.0 to 28.9%, but had no effect on the extractability of Zn. In most of the soils, ozonation had no statistically significant effect on bioavailability and mobility of Pb, residual after EDTA extraction. Using linear regression analysis we found a significant increase (p<0.01) in EDTA extractability of Pb after soil ozonation in soils with a higher initial Pb content. EDTA extractability of Pb after soil ozonation was also significantly higher for soils with a lower Pb extractability when treated with EDTA alone. We found no correlation between soil organic matter content and the percentage of the Pb fraction bound to soil organic matter (where from 25.6+/-1.3% to 73.2+/-0.6% of Pb reside in tested soils) and Pb extractability with EDTA after soil ozonation.

Heap leaching of lead contaminated soil using biodegradable chelator [S,S]-ethylenediamine disuccinate.

The feasibility of heap leaching Pb contaminated soil using biodegradable chelator [S,S]-ethylenediamine disuccinate ([S,S]-EDDS) in concentrations ranging from 10 to 40 mmol kg(-1) was assessed in a laboratory study. Treatment with 40 mmol kg(-1) [S,S]-EDDS was the most efficient, removing 31.3% of soil Pb and producing a waste washing solution with a peak Pb concentration slightly exceeding 10 mM. The removal of Pb from simulated washing solution containing 10 mM Pb by biodegradation of the Pb-[S,S]-EDDS complex and sorption of the released Pb on a vermiculite/soya meal/Slovakite based permeable bed was completed in 7.5 days. Lead recovery from washing solutions with higher molarity was significantly slower. Lead fractionation and bioavailability were assessed 6-months after soil treatment. Heap leaching removed Pb proportionally from carbonates (approx. 25% Pb) and organic matter (> 50% Pb), the two main Pb-bearing soil fractions,and increased the small pool of Pb soluble in the soil solution. The latter had no effect on Pb phytoavailability. Accumulation of Pb in leaves of the reported hyperaccumulator species Thlaspi goesingense was < 20 mg kg(-1) in all treatments, comparable to the Pb concentration in lettuce (Latuca sativa). Soil treatment with 40 mmol kg(-1) [S,S]-EDDS reduced concentrations of Pb orally-accessible in the stomach and intestine phase from initial 234.0 and 52.5 mg kg(-1) to 157.8 and 29.7 mg kg(-1), respectively, according to Ruby's test. Our results indicate that heap leaching of Pb with [S,S]-EDDS is not a viable remediation option for the tested soil and conditions used.

Washing of Pb contaminated soil using [S,S] ethylenediamine disuccinate and horizontal permeable barriers.

The feasibility of in situ washing of soil contaminated with Pb (6.83 mmol kg(-1)) using biodegradable chelator, [S,S] stereoisomere of ethylenediamine disuccinate ([S,S]-EDDS) and horizontal permeable barriers was examined in soil columns. After 4-cycles of 10 mmol kg(-1) soil [S,S]-EDDS applications, followed by irrigation, 24.7% of total initial Pb was washed from the contaminated soil and accumulated into the barrier. Sequential extractions indicated that washing removed most of the Pb from the organic soil fraction. Barriers were positioned 20 cm deep in the soil and consisted of a 2 cm layer of nutrient enriched vermiculite. Barriers reduced leaching of Pb in the first cycle of [S,S]-EDDS addition by more than 500-times compared to columns with no barrier. After four cycles of chelator addition, a total of 0.24% of the initial Pb was leached from the columns with barriers. Four cycles of in situ soil washing in soil columns were less effective than simulated ex situ soil washing with 40 mmol kg(-1) [S,S]-EDDS, where 51.0% of the Pb was removed after 48-h extraction. Ex situ soil washing with 10 mmol kg(-1) [S,S]-EDDS was equally effective as the first cycle of in situ soil washing (15.5% and 14.5% of removed Pb, respectively).

Ethylenediaminedissuccinate as a new chelate for environmentally safe enhanced lead phytoextraction.

Using a soil column experiment, we compared the effect of a single dose and weekly additions of ethylenediaminetetraacetic acid (EDTA) and ethylenediaminedissuccinate (EDDS) on the uptake of Pb, Zn, and Cd by Chinese cabbage [Brassica rapa L. subsp. pekinensis (Lour.) Hanelt], and on the leaching of heavy metals through the soil profile. The analysis of plant material revealed that both chelates increased the concentrations of Pb and, to a lesser extent, also of Zn and Cd in the leaves of the test plant. The most effective applications were single doses of 10 mmol EDTA and EDDS kg(-1) soil, which caused the concentrations of Pb in the shoots to increase 94.2- and 102.3-fold, respectively, relative to the control. The same dose of EDTA increased the concentration of Zn and Cd in the leaves 4.3- and 3.8-fold and of EDDS 4.7- and 3.5-fold, respectively. In treatments with weekly additions and lower concentrations of both chelates, EDTA was more effective than EDDS in increasing the plant uptake of Pb. In soil columns treated with weekly additions of 10 mmol kg(-1) EDTA, on average 22.7, 7.0, and 39.8% of initial total Pb, Zn, and Cd in the soil were leached through the soil profile. The same amount of EDDS caused much lower leaching of Pb and Cd--only 0.8 and 1.5% of initial total concentrations. Leaching of Zn, 6.2% of the total concentration, was comparable with the EDTA treatment. A biotest with red clover (Trifolium pratense L.) indicated a greater phytotoxic effect of EDTA than EDDS addition. EDDS was also less toxic to soil fungi, as determined by phospholipid fatty acid (PLFA) analysis, and caused less stress to soil microorganisms, as indicated by the trans to cis PLFA ratio. Chelate addition did not prevent the development of arbuscular mycorrhiza on red clover.

Development of fungal inocula for bioaugmentation of contaminated soils.

This report describes novel fungal inocula for bioaugmentation of soils contaminated with hazardous organic compounds. The inocula are in the form of pelleted solid substrates coated with a sodium alginate suspension of fungal spores or mycelial fragments and incubated until overgrown with the mycelium of selected lignin-degrading fungi. The organisms evaluated were Phanerochaete chrysosporium (BKM F-1767, ATCC 42725), P. sordida (HHB-8922-Sp), Irpex lacteus (Mad-517, ATCC 11245), Bjerkandera adusta (FP-135160-Sp, ATCC 62023), and Trametes versicolor (MD-277). The pelleted fungal inocula resisted competition and proliferation from indigenous soil microbes, were lower in moisture content than current fungal inocula, and had sufficient mechanical strength to allow handling and introduction into the soil without a change in the mechanical consistency of the pellets. Inoculated at a rate of 3% in artificially contaminated nonsterile soil, I. lacteus, B. adusta, and T. versicolor removed 86, 82, and 90%, respectively, of the pentachlorophenol in 4 weeks. A mathematical model was developed to explain moisture distribution in a hydrogel-coated pelleted substrate.

Physiological Aspects of Biosynthesis of Lignin Peroxidases by Phanerochaete chrysosporium.

Methods based on UV-visible diffuse reflectance spectroscopy were used to study the physiological aspects of lignin-peroxidase biosynthesis by Phanerochaete chrysosporium. Here we introduce the use of cytochrome aa(3) as an indicator of active fungal biomass and of its redox state to calculate the oxygen mass transport coefficient between the growth medium and the fungal cell interior. When lignin peroxidase biosynthesis was enhanced by the addition of Tween 80 or Tween 20 to the growth medium, a higher proportion of reduced cytochrome aa(3) and a higher oxygen diffusion barrier were observed compared with control cultures. In cultures supplemented with Tween 80 or Tween 20, a higher oxygen mass transport coefficient between the growth medium and the interior of the fungal cell was also found. The beginning of the lignin peroxidase activity in these cultures was found to coincide with a temporary cessation in the dry biomass increase and a reduction in the relative active-biomass concentration. During the lignin peroxidase activity, a decrease in the intracellular pH and an increase in the growth medium pH were determined in cultures supplemented with Tween 80.

Analysis of Fungal Pellets by UV-Visible Spectrum Diffuse Reflectance Spectroscopy.

The application of the UV-visible spectrum diffuse reflectance spectroscopy for the determination of intracellular pH in vivo, for determination of cytochrome content, and for the noninvasive in vivo detection of the redox state of fungal mitochondrial cytochromes in filamentous fungi is introduced. The time course of the intracellular pH values, mitochondrial cytochromes, and CO-binding pigments content and the correlations between the actual redox state of cytochrome aa(3) and saturation of growth medium with oxygen in pellets of the basidiomycete Phanerochaete chrysosporium were determined. As the test microorganism, the yeast Saccharomyces cerevisiae was used. UV-visible spectrum diffuse reflectance spectroscopy proved to be a promising method for the quick and simple analysis of light-impermeable biological structures for which the classical transmittance spectrophotometric methods are difficult to implement.

Amino acids in chitin - glucan complex ofAspergillus niger.

DeproteinatedA. niger biomass contains several covalently bound amino acids. The most abundant are arginine, serine, and proline in molar ratio of 3:2:2. One order of magnitude less is the amount of valine, phenylalanine, leucine and glycine. On deacetylation and separation of chitosan from glucan, the main three amino acids remain bound predominantly to chitosan, whereas the hydrophobic amino acids accompany mainly glucan. The presence of arginine could be the cause of stronger basicity of fungal chitosan compared to polyglucosamine.