Soil Immobilization of Heavy Metal Using Soil Amendments in a Greenhouse Study

ABSTRACT

The application of soil amendments to immobilize heavy metals is a promising technology to meet the requirements for environmentally sound and cost-effective remediation. The present work was carried out to evaluate the effect of phosphogypsum (PG) used alone and in combination with compost (CP) at a mix ratio of 1:1 wet weight ratio (PG+CP) at 10 and 20 g dry weight kg^?1 dry soil, on heavy metal immobilization in contaminated soil, and on canola growth (Brassica napus). The results were then compared with untreated soil (control). The results revealed that the Pb, Cd, and Zn uptake of canola plants was reduced by the application of PG alone and when it was mixed with CP. At an application rate of 10 g dry weight kg^?1 dry soil of (PG+CP), the dry weight of canola plants increased by 66.8%, which was increased in comparison on its weight in the untreated soil (control). The addition of PG alone resulted in more pronounced immobilization of heavy metals as compared to PG mixed with CP. Plant growth was improved with CP addition but heavy metals immobilization was greatest in PG alone treatments. Results suggest that PG may be useful for the immobilization of heavy metals in contaminated soils.     

Spectroscopic analyses to study the effect of biochar and compost on dry mass of canola and heavy metal immobilization in soil

This research was conducted to use spectroscopic analysis to evaluate the effect of biochar and compost application on heavy metal immobilization during the canola plants grown in the contaminated soil using X-Ray Diffraction (XRD), Energy Dispersive spectroscopy by X-rays (EDX) and Fourier Transmission Infrared Spectroscopy (FTIR). The results showed that the cadmium (Cd) and lead (Pb) concentrations in the root and shoot of canola plants significantly decreased with the addition of rice straw compost (RC) and biochar (RB) to contaminated soil. The use of spectroscopic analysis observed the precipitation, inner-sphere complex reaction, and electrostatic attraction are the dominating mechanisms for heavy metals immobilization with organic amendments. Our results indicate that the rice straw biochar and compost can immobilize heavy metals and improve dry weight of canola plants. Moreover, the spectroscopic analysis is a simple method and can be effective to understand the mechanism of heavy metals immobilization.     

Silicon alleviates the toxicity of cadmium and zinc for maize (Zea mays L.) grown on a contaminated soil

Although silicon (Si) is not an essential element, it presents a close relationship with the alleviation of heavy‐metal toxicity to plants. This work was carried out to evaluate the effects of Si application to soil on the amelioration of metal stress to maize grown on a contaminated soil amended with Si (0, 50, 100, 150, and 200 mg kg^–1) as calcium silicate (CaSiO₃). Additionally, the cadmium (Cd) and zinc (Zn) bioavailability as well as their distribution into soil fractions was also studied. The results showed that adding Si to a Cd‐ and Zn‐contaminated soil effectively diminished the metal stress and resulted in biomass increase in comparison to metal‐contaminated soil not treated with Si. This relied on Cd and Zn immobilization in soil rather than on the increase of soil pH driven by calcium silicate application. Silicon altered the Cd and Zn distribution in soil fractions, decreasing the most bioavailable pools and increasing the allocation of metals into more stable fractions such as organic matter and crystalline iron oxides.     

Lead and Zinc Extraction Potential of Two Common Crop Plants, Helianthus Annuus and Brassica Napus

Phytoextraction is a remediation technology that uses plants to remove heavy metals from soil. The success of a phytoextraction process depends on adequate plant yield (aerial parts) and high metal concentrations in plant shoots. A pot experiment was conducted to investigate the combination effects of plants [sunflower (Helianthus annuus) and canola (Brassica napus)] with soil treatments (manure, sulfuric acid and DTPA). Treatments, including two plants and seven soil treatments, which applied according to completely randomized factorial design with three replications. The largest shoot dry weight biomass production occurred in manure treatments for both plants. The maximum shoot concentrations of Pb and Zn were 234.6 and 1364.4 mg kg^?1 respectively in three mmoles DTPA kg^?1 treatment of sunflower. Furthermore the results showed that sunflower had a higher extracting potential for removal of Pb and Zn from polluted soil.     

Rice Straw Composting and Its Effect on Soil Properties

In Egypt, recycling rice straw and organic wastes is of great concern as well as improvement of soil properties. Rice straw compost could improve both organic waste recycling and soil quality. The aim of this study was to evaluate the effect of the rice straw compost, with or without water treatment residuals (WTR), on soil chemical properties and dry weight of canola. The results showed that the addition of rice straw compost and WTR decreased soil salinity and increased Ca⁺², K⁺ and organic matter. The addition of compost and WTR (2:1 wet weight ratio) at level of 10 g dry weight kg^?1 dry soil gave the best reduction in soil salinity compared to compost or WTR only or for level 10 g dry weight kg^?1 dry soil. The results also showed that the available P decreased with the application of WTR while it increased with the application of compost. The study demonstrated that the dry weight and relative increase (R.I %) of dry weight of canola plants increased with the application of WTR and compost to soil. Combinations of WTR and compost to soils had a greater effect on increasing yield and improved the efficiency of WTR on soil properties.     

Fraction Distributions of Lead,Cadmium, Copper,and Zinc in Metal‐Contaminated Soil before and after Extraction with Disodium Ethylenediaminetetraacetic Acid

Abstract: The fraction distributions of heavy metals have attracted more attention because of the relationship between the toxicity and their speciation. Heavy‐metal fraction distributions in soil contaminated with mine tailings (soil A) and in soil irrigated with mine wastewater (soil B), before and after treatment with disodium ethylenediaminetetraacetic acid (EDTA), were analyzed with Tessier's sequential extraction procedures. The total contents of lead (Pb), cadmium (Cd), copper (Cu), and zinc (Zn) exceeded the maximum permissible levels by 5.1, 33.3, 3.1, and 8.0 times in soil A and by 2.6, 12.0, 0.2, and 1.9 times in soil B, respectively. The results showed that both soils had high levels of heavy‐metal pollution. Although the fractions were found in different distribution before extraction, the residual fraction was found to be the predominant fraction of the four heavy metals. There was a small amount of exchangeable fraction of heavy metals in both contaminated soils. Furthermore, in this study, the extraction efficiencies of Pb, Cd, and Cu were higher than those of Zn. After extraction, the concentrations of exchangeable Pb, Cd, Cu, and Zn increased 84.7 mg·kg^?1, 0.3 mg·kg^?1, 4.1 mg·kg^?1, and 39.9 mg·kg^?1 in soil A and 48.7 mg·kg^?1, 0.6 mg·kg^?1, 2.7 mg·kg^?1, and 44.1 mg·kg^?1 in soil B, respectively. The concentrations of carbonate, iron and manganese oxides, organic matter, and residue of heavy metals decreased. This implies that EDTA increased metal mobility and bioavailability and may lead to groundwater contamination.     

Reduction of soil heavy metal bioavailability by nanoparticles and cellulosic wastes improved the biomass of tree seedlings

The purpose of this study was to use zero‐valent iron nanoparticles (nZVI) and cellulosic wastes to reduce bioavailability of lead (Pb) and cadmium (Cd), and to establish Persian maple seedlings (Acer velutinum Bioss.) in contaminated soil. One‐year‐old seedlings were planted in pots filled with unpolluted soil. Lead [Pb(NO₃)₂] and Cd [Cd(NO₃)₂] were added with concentrations of 0 (Control), 100 (Pb100), 200 (Pb200), and 300 (Pb300) mg kg⁻¹ and 10 (Cd10), 20 (Cd20), and 30 (Cd30) mg kg⁻¹. Cellulosic wastes were mixed with soil at the same time of planting [four levels: 0, 10 (W1), 20 (W2), 30 (W3) g 100 g⁻¹ soil]. The nZVI was prepared by reducing Fe³⁺ to Fe⁰ and injected to pots [four levels: 0, 1 (N1), 2 (N2), and 3 (N3) mg kg⁻¹]. Height, diameter, biomass, tolerance index of seedlings, bioavailability of heavy metals in soil, and removal efficiency of amendments were measured. The highest values of seedling characteristics were observed in N3. The highest removal efficiency of Pb (Pb100: 81.95%, Pb200: 75.5%, Pb300: 69.9%) and Cd (Cd10: 92%, Cd20: 73.7%, Cd30: 68.5%) was also observed in N3. The use of nZVI and cellulosic waste could be a proper approach for seedling establishment in forests contaminated with heavy metals.     

Removal of Lead from Contaminated Soils by Typha Angustifolia

A greenhouse study was demonstrated for removal of lead (Pb) from contaminated soil by the narrow — leaved cattail, Typha angustifolia. The plants were grown in sandy loam soil containing various concentrations of Pb(NO₃)₂ (53.3, 106.7, 160, 213.3, and 266.7 mg Pb kg^-1 soil). Most lead was accumulated in roots and then transported to leaves. In soil contaminated with 266.7 mg kg^-1 of lead, the plants accumulated 7492.6 mg Pb kg^-1 dry weight in the roots and 167 mg Pb kg^-1 dry weight in the leaves. Yet, no growth retardation from lead was detected. T. angustifolia has high potential as a plant to clean up lead contaminated soil due to its vigorous growth, high biomass productivity, and because it is a perennial in nature. Further work is required to study on the iron plaque formation and its role in metal immobilization.     

Effect of phosphogypsum application and bacteria co-inoculation on biochemical properties and nutrient availability to maize plants in a saline soil

Phosphogypsum (PG), which contains Ca, P and S and has an acidic effect, may be applied to manage soil constraints such as alkalinity and salinity. For increasing nutrients bioavailability, biofertilizers are commonly applied. Therefore, the aim of this study was to assess PG effect either alone or in combination with the mixed co-inoculation of plant growth promoting rhizobacteria on a saline soil. In a greenhouse pot experiment with maize (Zea mays L.), the inoculated and non-inoculated saline soils were treated with PG at 10 g kg^?1 (PG10), 30 g kg^?1 (PG30), and 50 g kg^?1 (PG50). The soil pH, electrical conductivity (EC_e), and macro-(NPK) and micronutrients (Fe, Mn, Zn, and Cu) availability to mays were examined. Applying PG reduced soil pH and co-inoculation induced significant decreases in soil EC_e. Applying PG increased significantly soil available P. Applying PG combined with co-inoculation effectively increased the soil available K. The soil available micronutrients decreased significantly with PG. However, the inoculated maize treated with PG showed significant higher dry weight (82.1–127.4%) and nutrients uptake than the control. It could be concluded that PG along with co-inoculation may be an important approach for alleviating negative effects of salinity on plant growth.     

Optimization strategies for Cd and Pb immobilization in soil using meta-analysis combined with numerical modeling

Chemical immobilization is one of the most effective technologies for remediating sites with heavy metals,but the selection of proper immobilization material and determination of its dose ratio is a challenge that limits the remediation efficiency.In this study,we conducted a meta-analysis of 489 independent observations on the immobilization of heavy metals,in which the immobilization materials were divided into biochar,phosphate,lime,metal oxides,and clay minerals.The statistical analysis of t...     

Effect of Organic Acids on Heavy-Metal Uptake and Growth of Canola Grown in Contaminated Soil

Heavy-metal pollution of soils causes many environmental, animal, and human health problems. Phytoextraction of heavy metals from contaminated soils is an effective and economic technique. Humic acids are naturally occurring phenol body polymerisates, which form chelate compounds with heavy metals. In the present study the influence of soil- applied humic, citric, and malic acids on the lead (Pb), cadmium (Cd), and chromium (Cr) uptake from a contaminated soil by canola plant was examined in a greenhouse experiment. The experiment was arranged in factorial design based on randomized complete blocks with three replications. The factors of experiment included three organic acids (humic, citric, and malic acid) as first factor and five concentrations [0, 0.001, 0.002, 0.003, and 0.004 (v/v)] as second factor. The results showed that increase in organic acid concentration significantly increased heavy-metal uptake by canola plant, which accumulated heavy metals in different parts of the plant. In addition, crop growth representing by plant height and plant dry weight as well as seed production significantly decreased. Based on these results, canola can be considered as effective crop for phytoextraction of heavy metals from contaminated soils.     

Organobentonite for Sorption and Degradation of Phenol in the Presence of Heavy Metals

An experimental study was performed to determine the feasibility of usingorganobentonite modified with quarternary ammonium cations (QACs) as a reactive medium in immobilization and biodegradation barriers for mixed contaminants in the subsurface soil. Various factors, including interactions between heavy metals, organic contaminants, and soil microorganisms, were investigated when they coexisted with untreated bentonite and organobentonite. Batch sorption tests for cadmium and lead were conducted to quantify sorption selectivity of these metals on untreated bentonite and organobentonite. Metal concentrations of 50 × 10^-6 M slightly reduced the growth of soil microbes and partially interfered with the biodegradation of phenol. Soil microorganisms tested with untreated bentonite grew after approximately 25 hr of lag period and degraded phenol completely within 350 hr. The results from this study demonstrate that organobentonite could be used as a reactive medium for immobilization and biodegradation of organic contaminants in the presence of heavy metals in the subsurface soil.     

Application of Organic Wastes in a Soil Polluted with Different Rates of Cr-Pb: Effects on Soil Biological Properties

An experiment was performed to determine the effects of adding municipal solid waste (MSW) and poultry manure (PM) to a soil polluted with chromium (Cr), lead (Pb), and Cr + Pb on the biological parameters of the soil. Soil was mixed with two solutions of Cr(NO₃)₃ and/or Pb(NO₃)₂ to give three concentrations (0, 100, and 250 mg Cr kg^?1 soil and 0, 100, and 250 mg Pb kg^?1 soil) and treated with MSW or PM. When the soil was contaminated with the metals without combining, the greatest adenosine triphosphate (ATP), urease, and phosphatase inhibition percentages occurred for 250 mg Pb kg^?1 soil. When the heavy metals were mixed, the inhibition of the biochemical parameters increased. The application of MSW and PM decreased the inhibition of the biochemical parameters and microbial population in the polluted soils. The inhibition percentage was greater for the soil amended with MSW than with PM, possibly due to the high humic acid concentration.     

Transfer of Cd, Pb, Ra and U from Phosphogypsum Amended Soils to Tomato Plants

About 170 million tons of phosphogypsum (PG) are annually generated worldwide as a by-product of phosphoric acid factories. Agricultural uses of PG could become the main sink for this waste, which usually contains significant radionuclide (from the ²³⁸U-series) and toxic metals concentrations. To study PG effects on pollutant uptake by crops, a completely randomised greenhouse experiment was carried out growing Lycopersicum esculentum Mill L. on a reclaimed marsh soil amended with three PG rates (treatments), corresponding to zero (control without PG application), one, three and ten times the typical PG rates used in SW Spain (20 Mg ha^?1). The concentrations of Cd, Pb, U (by inductively coupled plasma mass spectroscopy) and ²²⁶Ra and ²¹⁰Po (by γ-spectrometry and α-counting, respectively) were determined in soil, vegetal tissue and draining water. Cadmium concentrations in fruit increased with PG rates, reaching 44?±?7 μg kg^?1 formula weight with ten PG rates (being 50 μg kg^?1 the maximum allowed concentration by EC 1881/2006 regulation). Cd transfer factors in non-edible parts were as high as 4.8?±?0.5 (dry weight (d.w.)), two orders of magnitude higher than values found for lead, lead, uranium and radium concentrations in fruit remained below the corresponding detection limits—0.5 and 0.25 mg kg^?1 and 0.6 mBq kg^?1, respectively (in a d.w. basis). ²³⁸U (up to 7 μg kg^?1 d.w.) and ²¹⁰Po (up to 0.74 Bq kg^?1 d.w.) could be measured in some fruit samples by α-spectrometry. Overall, the concentrations of these metals and radionuclides in the draining water accounted for less than 1% of the amount applied with PG.     

Ecological implications of motor oil pollution: Earthworm survival and soil health

The nontarget effects of fresh and used motor oil were studied in a soil test system involving such criteria as earthworm survival, response of soil dehydrogenase and urease, and nitrification. When earthworms were exposed to motor oil-contaminated soil for 4 weeks, the observed median lethal concentrations (LC₅₀) were 40.33 and 3.88 g kg⁻¹ soil for fresh and used oil, respectively. Only fresh motor oil application increased earthworms' body weight even at the higher dose of 19 g kg⁻¹ soil. Gas chromatography/mass spectrometry revealed that used motor oil contained more of aromatic hydrocarbons and heavy metals than fresh oil. This disparity in the chemical composition might be the factor responsible for the significant toxicity of used motor oil towards earthworms. Activities of soil dehydrogenase and urease were significantly enhanced in presence of both the motor oils, while there was a significant inhibition in nitrification by the used motor oil even at a low concentration of 0.2 g kg⁻¹ soil. This study clearly demonstrated that earthworm survival and nitrification could serve as suitable indices to assess motor oil pollution in soil.     

Bioremediation of Co-contamination of Crude Oil and Heavy Metals in Soil by Phytoremediation Using Chromolaena odorata (L) King & H.E. Robinson

The capability of Chromolaena odorata (L) to grow in the presence of different concentrations of three heavy metals in crude oil-contaminated soil and its capability to remediate the contaminated soil was investigated using pot experiments. C. odorata plants were transplanted into contaminated soil containing 50,000 mg kg^?1 crude oil and between 100 and 2,000 mg kg^?1 of cadmium, nickel, and zinc and watered weekly with water containing 5% NPK fertilizer for 180 days. C. odorata did not show any growth inhibition in 50,000 mg kg^?1 crude oil. Plants in experiments containing 2,000 mg kg^?1 Cd showed little adverse effect compared to those in Zn-treated soil. Plants in 1,000 and 2,000 mg kg^?1 Ni experiments showed more adverse effects. After 180 days, reduction in heavy metals were: 100 mg kg^?1 experiments, Zn (35%), Cd (33%), and Ni (23%); 500 mg kg^?1, Zn (37%), Cd (41%), and Ni (25%); 1,000 mg kg^?1, Zn (65%), Cd (55%), and Ni (44%); and 2,000 mg kg^?1, Zn (63%), Cd (62%), and Ni (47%). The results showed that the plants accumulated more of the Zn than Cd and Ni. Accumulation of Zn and Cd was highest in the 2,000 mg kg^?1 experiments and Ni in the 500 mg kg^?1 experiments. Crude oil was reduced by 82% in the experiments that did not contain heavy metals and by up to 80% in the heavy metal-treated soil. The control experiments showed a reduction of up to 47% in crude oil concentration, which was attributed to microbial action and natural attenuation. These results show that C. odorata (L) has the capability of thriving and phytoaccumulating heavy metals in contaminated soils while facilitating the removal of the contaminant crude oil. It also shows that the plant??s capability to mediate the removal of crude oil in contaminated soil is not significantly affected by the concentrations of metals in the soil.     

Phytoextraction of heavy metal contaminated soils with Thlaspi goesingense and Amaranthus hybridus: Rhizosphere manipulation using EDTA and ammonium sulfate

Selection of appropriate plant species and rhizosphere manipulation to enhance metal uptake are considered key factors in the development of phytoextraction technologies. A pot trial was conducted with two contaminated soils to investigate the effect of EDTA and ammonium sulfate on the accumulation of heavy metals into shoots of the low‐biomass hyperaccumlator Thlaspi goesingense Hálácsy (Brassicaceae) and the high‐biomass non‐hyperaccumulating plant Amaranthus hybridus (Amaranthaceae). Upon application of 1 g EDTA (kg soil)^—1 metal extractability with 1 M NH₄NO₃ increased substantially, whereas the application of (NH₄)₂SO₄ was less effective. The EDTA treatment increased the heavy metal concentrations in both plant species, however, the difference to the control was larger for A. hybridus. EDTA enhanced shoot concentrations in A. hybridus grown on soil Arnoldstein from 32.7 mg kg^—1 to 1140 mg kg^—1 for Pb and from 3.80 mg kg^—1 to 10.3 mg kg^—1 for Cd. Cd concentrations in shoots of T. goesingense were also increased by EDTA application, however, a slight decrease was observed for Pb. T. goesingense accumulated 2840 mg Pb kg^—1 without any treatment. This is the first report of Pb hyperacumulation by T. goesingense. A decrease of shoot Pb concentration was observed in T. goesingense upon treatment with ammonium sulfate. Although metal concentrations in the shoots were rather large and significantly increased upon application of EDTA, plant growth and heavy metal removal were still too small to obtain reasonable extraction rates in soils heavily polluted by metals. It should be also noted that metal lability largely increased in EDTA‐treated soils and this lability persisted for several weeks after the application of the chelating agent, which is likely to be associated with the risk of groundwater contamination.     

Influence of acetamiprid on soil enzymatic activities and respiration

The effect of a new pesticide, acetamiprid, applied at normal field concentration (0.5 mg kg⁻¹ dried soil) and at high concentration (5 and 50 mg kg⁻¹ dried soil), on soil enzyme activities and soil respiration in upland soil was studied. The results showed that acetamiprid had a strong negative influence on soil respiration and phosphatase activity, and the enzyme activities in soil treated with 5 and 50 mg kg⁻¹ dry soil were significantly (P < 0.05) lower than the CK over the course of incubation. The 7-, 14-, and 35-day EC10 for phosphatase were 11, 15, and 11 mg kg⁻¹ dry soil, respectively. The 21-day EC10 and EC50 for soil respiration was 0.005 and 83 mg kg⁻¹ dry soil. The activity of dehydrogenase was enhanced after acetamiprid application for 2 weeks and the enzyme activities in samples treated with 0.5, 5 and 50 mg kg⁻¹ dry soil was about 2.5-, 1.5- and 2-fold to that of the control on sample day 28. Variance of urease and catalase had no distinct relationship with the application concentration. The activity of proteinase was not significantly affected within the first 2 weeks but inhibited from the fourth week after acetamiprid application and was only 0.45-fold to that of the control on sample day 28. Overall, acetamiprid at normal field dose would not pose a toxicological threat to soil enzymes, but a certain potential threat to soil respiration.     

Role of soil constituents in fixation of soluble Zn, Cu, Ni and Cd added to soils

Slow immobilization of trace metals in soil, termed ‘fixation’, affects their natural attenuation but it is still unclear which reactions occur. Twenty‐eight soils were selected to assess the role of Fe oxides and carbonates on fixation of Cu, Cd, Zn and Ni. Soils included samples from 2 toposequences (Vietnam, Spain) and 13 European topsoils with different soil characteristics (pH 3.4–7.7). Samples were amended with 250 mg Zn kg⁻¹, 100 mg Cu kg⁻¹, 80 mg Ni kg⁻¹ and 2.5 mg Cd kg⁻¹ as metal salts and incubated for 850 days. Fixation was measured as the increase of the fraction of added metals that were not isotopically exchangeable. Fixation increased with time and was, averaged over all the soils, 43% (Cu), 41% (Zn), 41% (Ni) and 28% (Cd) after 850 days. Metal fixation within samples from each toposequence was generally positively related to total Fe oxide concentration (Fe_d) for Zn, Ni and Cd. However, the fixation of Cd, Zn and Ni was mainly explained by pH and not by Fe_d when considering all soils. Fixation of Zn and Cd in soils with pH >7.0 increased with increasing concentrations of carbonates at initial ageing times. Fixed fractions of Zn, Ni and Cd were significantly released when experimentally removing 50% of carbonates by acidification. Fixation of Cu was most poorly related to soil properties. Our data suggest that fixation of Cd, Zn and Ni is related to a pH‐dependent diffusion into oxides and that of Cd and Zn also to diffusion and/or coprecipitation in carbonates. Fixation of Ni at neutral pH may also be related to stabilization of precipitates that form readily in soil.     

Schwermetallanreicherung und Schwermetallverfügbarkeit im Einsickerungsbereich von Stammablaufwasser in Buchenwäldern (Fagus sylvatica) des Wienerwaldes

Heavy metal enrichment and mobility in the infiltration zone of stemflow from beech (Fagus sylvatica) in the Vienna woods Filtering action of beech crowns in a polluted atmosphere produces acidic stemflow enriched with heavy metals. Series of soil profiles through the infiltration zone of stemflow waters were analyzed in three old growth beech stands on pseudogley (aqualf) soils of the northeastern Vienna Woods to study soil acidification and heavy metal accumulation and mobility: + In the infiltration zone of stemflow soil pH can be less than pH 3 and thus more than 2 pH units lower than in profiles more distant from the stem base. + Heavy metal contents of the soil peaked near the stembase. Maximum values for lead were 500 to 1.200 mg·kg^?1 soil, for copper 70 to 150 mg·kg^?1 soil and for zinc 250 to 500 mg·kg^?1 soil. + Manganese was not enriched but leached out from the soil profile. + The proportion of AED-complexable lead and copper to total lead and copper decreased under stemflow influence in the topsoil but increased in the subsoil. These results indicate that significant amounts of heavy metals are deposited with stemflow in beech stands of the Vienna Woods. Strong soil acidification enhances heavy metal solubility and thus promotes leaching into deeper soil strata.