Journal of Hydrology xxx (2012) xxx–xxx

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Journal of Hydrology

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Arsenic bioaccessibility and speciation in the soils amendedwith organoarsenicals and drinking-water treatment residuals basedon a long-term greenhouse study

Rachana Nagar a, Dibyendu Sarkar b, Konstantinos C. Makris c, Rupali Datta d,⇑a Weiss Associates, 2200 Powell Street, Suite 925, Emeryville, CA 94608, USAb Department of Earth and Environmental Studies, Montclair State University, Montclair, NJ, USAc Cyprus International Institute for the Environment and Public Health in Association with the Harvard School of Public Health, 5 Iroon Street, 1105 Nicosia, Cyprusd Department of Biological Sciences, Michigan Technological University, 1400 Townsend Drive, Houghton, MI 49931, USA

a r t i c l e i n f o s u m m a r y

Article history:Available online xxxx

Keywords:OrganoarsenicalsDimethylarsinic acid (DMA)BioaccessibilitySpeciationDrinking-water treatment residual (WTR)

0022-1694/$ - see front matter � 2012 Elsevier B.V. Ahttp://dx.doi.org/10.1016/j.jhydrol.2012.10.013

⇑ Corresponding author. Tel.: +1 906 487 1783; faxE-mail address: rupdatta@mtu.edu (R. Datta).

Please cite this article in press as: Nagar, R., et atreatment residuals based on a long-term green

Although organoarsenical pesticides are no longer applied to agricultural fields in the US, their wide-spread use until recently, toxicity, and potential transformation to inorganic arsenic has raised seriousconcern. Drinking-water treatment residuals (WTRs) have been proposed as a low-cost amendment forremediation of organoarsenical pesticide contaminated soils. A long-term greenhouse study was initiatedto evaluate the effect WTR application on bioaccessibility, geochemical partitioning, and speciation of theDimethylarsinic acid (DMA). Two soils (Immokalee and Orelia series) were spiked with DMA(1500 mg As kg�1) and amended with an Al- and Fe-based WTR at two rates (5% and 10% by wt.). Soilsampling was done immediately after spiking (time zero) and after 0.25, 0.5, 1, and 3 (time final) yearsof equilibration and subjected to bioaccessibility test and sequential extraction. Results showed thatcompared to the unamended (no WTR) control, As bioaccessibility in the WTR-amended soils signifi-cantly (p < 0.001) decreased by 40–70% in 3 years. The Fe-WTR was more effective than Al-WTR indecreasing soil As bioaccessibility. The in vitro and water-extracted samples were subjected to As speci-ation at time zero and time final. Results showed transformation of DMA into inorganic As, irrespective ofWTR amendments. The Orelia soil showed significantly (p < 0.001) higher transformation than the Immo-kalee soil.

� 2012 Elsevier B.V. All rights reserved.

1. Introduction

Until recently, the use of organoarsenical pesticides were al-lowed for agricultural purposes under the assumption that theywere non-carcinogenic and less toxic compared to inorganic As(i-As). Organoarsenicals monomethylarsonate (MMAV) and dime-thylarsinate (DMAV), mainly used as pesticides/herbicides, intro-duced >1000 metric tons of As into the environment each year(Pongratz, 1988; Bendar et al., 2002). Recent toxicological data re-ported that these methyl As compounds can be as toxic as the inor-ganic species, and have several genotoxic and clastogenic effects(Yamamoto et al., 1995; Mass et al., 2001; Petrick et al., 2000;Salim et al., 2003; Schwerdle et al., 2003). Health concerns havebeen further exacerbated by potential biotransformation oforganoarsenicals to the more toxic inorganic forms in agriculturalfields and golf courses (Bendar et al., 2002; Feng et al., 2005; Huanget al., 2007; Yin et al., 2011). Recent studies by Shimizu et al.(2011) demonstrated demethylation of DMA to As(V) over 1 year

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l. Arsenic bioaccessibility andhouse study. J. Hydrol. (2012),

of incubation in aerobic environment. Lately, USEPA has bannedthe re-registration of organic arsenical pesticides (USEPA, 2006).However, vast stretches of agricultural lands have already beencontaminated. The phenomenon of urban sprawl has raised therisk of human contact with As-contaminated agricultural soils,and incidental hand-to-mouth activity by children playing in As-contaminated soils has become an issue of concern (Datta andSarkar, 2004). A key parameter determining the risk of As iningested soils is bioavailability: the relative amount of As thatis freely available for biological action (Subaczi et al., 2007).Therefore, it is imperative to develop cost-effective remediationmethods to minimize the risk of human exposure to As contami-nated soil.

Some studies have investigated the sorption of MMA and DMAon alumina, hematite, quartz, iron filing, and hydrous ferric oxidesin aqueous media (Xu et al., 1991; Cheng et al., 2005; Lafferty andLoeppert, 2005; Ramesh et al., 2007). Compared to MMA and i-Asspecies, DMA weakly bound to Fe/Al containing phases and showeda decreased affinity in the following order: i-As > MMAV > DMAV

(Cheng et al., 2005; Xu et al., 1991). However, a study by Coxand Ghosh (1994) showed the formation of strong inner-sphere

speciation in the soils amended with organoarsenicals and drinking-waterhttp://dx.doi.org/10.1016/j.jhydrol.2012.10.013

2 R. Nagar et al. / Journal of Hydrology xxx (2012) xxx–xxx

complexes of organoarsenicals on the surfaces of Fe oxides andalumina. Shimizu et al. (2011) showed that MMA and DMA formbidentate binuclear complexes with amorphous aluminum oxide.Results also indicated that increased methyl group substitutiondecreased As sorption. Another Extended X-ray Absorption FineStructure (EXAFS) study showed that both MMA and DMAform bidentate binuclear inner-sphere complexes with Al/Fe-oxyhydroxide surfaces of soil (Shimizu et al., 2011).

The present study proposes the use of a waste by-product gen-erated from the drinking water treatment process (i.e. the drink-ing-water treatment residual, or WTR) as a cost-effective methodfor immobilization of DMA from contaminated soils. Earlier batchexperiments in our laboratory demonstrated the huge affinity ofboth Al- and Fe-hydroxide based WTR in immobilizing i-As fromsoil and water (Nagar et al., 2009a, b and Nagar et al., 2010; Makriset al., 2006; Sarkar et al., 2007a). Water treatment residual wasalso effective in immobilizing organic As species (such as Roxar-sone and DMA) from poultry litter (PL) aqueous suspensions, asindicated by a recent study in our laboratory (Makris et al.,2008). Subaczi et al. (2007) showed decreasing As bioaccessibilitywith iron amendments (such as metallic Fe and soluble Fe(II)-and Fe(III)-halide salts) in As(V) contaminated soils. A recent re-search study by our group, using in vitro and in vivo experiments,has indicated the effectiveness of WTR in significantly (p < 0.001)decreasing soil As(V) bioavailability compared to unamended (noWTR) soil (Nagar et al., 2009a, b). However, to the best of ourknowledge, no work has been done so far on the long-term effec-tiveness of WTR in decreasing soil As bioaccessibility in DMA-amended agricultural soils. Because the chemical architecture ofMMAV and DMAV is similar to that of arsenate (Cheng et al.,2005), we hypothesized that these organic species could also beimmobilized by an As geosorbent (WTR) which might result in de-creased As availability in organoarsenical pesticide contaminatedsoils. A dynamic column study involving interactions between soil,water, WTR, and plants was carried out in a temperature-andhumidity-controlled greenhouse setting. Two types of soils, withvarying physico-chemical properties, were spiked with DMAV,which is the most commonly used organoarsenic pesticide (Caiet al., 2002), and amended with two types of WTR (Fe- and Al-based). Rice (Oryza sativa) was used as the test crop.

The objectives of the present study were (i) to assess the long-term (3-year) effect of two types of WTR (Fe- and Al-based) on Asbioaccessibility in two soils amended with DMAV; (ii) to evaluatethe effect of WTR on As uptake by rice (Oryza sativa) and As con-centration in leachates; (iii) to identify the geochemical forms thatinfluence As reactivity and bioaccessibility; and (iv) to examine thechanges in As oxidation states (speciation) in in vitro and water ex-tracts at the end of 3 years (time final) in WTR-amended and una-mended soils.

2. Materials and methods

2.1. Greenhouse study

2.1.1. Study design and soil amendmentsSurface soils (0–15 cm) of Immokalee and Orelia series were

used for the current greenhouse study. The Immokalee series soilwas collected from the Southwest Florida Research and EducationCenter in Immokalee, Florida and the Orelia soil was collected fromCorpus Christi, Texas. The Fe- and Al-based WTRs were obtainedfrom the drinking-water treatment plants in Tampa, FL andBradenton, FL, USA, respectively. All the samples were allowed toair-dry and were subsequently passed through a 2-mm sieve be-fore being subjected to characterization and sorption experiments.Soil and WTR samples were characterized for several physico-chemical properties as discussed earlier in Nagar et al. (2009a, b).

Please cite this article in press as: Nagar, R., et al. Arsenic bioaccessibility andtreatment residuals based on a long-term greenhouse study. J. Hydrol. (2012),

A three-year greenhouse study was initiated by preparing 40columns. The columns were made of PVC (0.38 m high � 0.15 minternal diameter) pipes. An outlet nozzle was fitted at the bottomand connected with a tube to collect the leachate in a clean, 1 Lplastic bottle. To prevent soil leaching from the column, a fine ny-lon mesh (8 lm mesh size) was placed at the bottom of the columnabove the outlet nozzle. Each column was filled with 0.18 m heightof play sand, followed by 0.15 m of soil.

Both soils (Immokalee and Orelia series) were amended with anorganoarsenical, cacodylic acid sodium salt or DMAV (CH3)2AsO2H-MP Biomedics Inc., Germany, CAS #105645) – to a final As load of1500 mg kg�1. Soils were also mixed thoroughly with Al- or Fe-WTR at two rates (5% and 10% w/w) (Sarkar et al., 2007b). Soilspiked with DMAV but not WTR was included as a control. Columnswere filled with the soil-pesticide-WTR mixture after thoroughmixing. For samples spiked with DMA, there were a total of 32 potswith WTR treatments (two soils � two types of WTR � two rates ofWTR � four replicates). In addition, eight control columns withoutWTR (two soils � four replicates) were also prepared. The soil col-umns were maintained at optimum soil moisture conditions (70%of water holding capacity). The pots were arranged in a random-ized block design and were rotated periodically to account for vari-ances in temperature and sunlight within the greenhouse. Rice wasused as the test crop.

Soil samples were collected by composite sampling from thetop 10-cm surface. The first soil sampling was done immediatelyafter spiking (time zero), and then after 0.25 and 0.5 year of equil-ibration time. The columns were over-watered twice (after 0.25and 0.5 year) to induce leaching. The leachate was collected foreach column and analyzed for soluble As. Plant samples were har-vested at 0.5 year. These samples were digested and analyzed forAs according to Carbonell et al., 1998. Columns were retained forthe remaining time without grass cover to understand the effectof soil/WTR aging on As fractionation and bioaccessibility. After0.5 year, soils were sampled at the end of 1 and 3 years. After eachsampling, soil samples were digested using the USEPA 3050Bmethod (USEPA, 2000), and analyzed using an ICP-MS (Perkin El-mer Elan 9000 model). Samples were also subjected to in vitroand sequential extraction schemes after each sampling using thefollowing protocols.

2.1.2. Sequential extraction procedureThe sequential extraction of As was performed using the

method by Chunguo and Zhui (1988) with certain modifications(Datta et al., 2007) for the following operationally defined Asforms: water-soluble, NH4Cl-extractable (exchangeable), NaOH-extractable (amorphous Fe/Al-bound), H2AsO4-extractable (Ca/Mg-bound), H2O2-extractable (organic/sulfides-bound), and HNO3-extractable (residual) As. Extracts were filtered and analyzed fortotal soluble As by ICP-MS (Perkin Elmer Elan 9000 model).

2.1.3. In vitro extraction schemeBioaccessible As in the stomach phase was determined using

the in vitro method (IVG) by Rodriguez et al. (1999) with certainmodifications by Sarkar and Datta (2003). We considered onlythe gastric phase, because further extending the gastric to theintestinal phase has not been shown to improve the predictiveability of the in vitro models (Pouschat and Zagury, 2006;Rodriguez et al., 1999). One gram of soil was added to 150 mL gas-tric solution. The gastric solution consisted of 0.15 M NaCl and 1%porcine pepsin (Acros Organics, New Jersey, USA). Gastric solutionpH was adjusted to1.8 with trace metal grade HCl following theaddition of soil. Samples were incubated for 1 hour (h) underanaerobic conditions, which was maintained by purging argongas through the gastric solution. After 1 h, 10 mL of supernatantwas withdrawn, centrifuged at 4000 g for 20 min, passed through

speciation in the soils amended with organoarsenicals and drinking-waterhttp://dx.doi.org/10.1016/j.jhydrol.2012.10.013

R. Nagar et al. / Journal of Hydrology xxx (2012) xxx–xxx 3

0.22 lm nylon filters, and analyzed for bioaccessible As byinductively-coupled plasma mass-spectrometry (ICP-MS).

2.1.4. Arsenic speciation protocolAt time zero and time final (year 3), in vitro and water-ex-

tracted (1:10 – soil:dH2O) samples were subjected to As speciationanalysis. First, filtered samples were subjected to a liquid/liquid (L/L) extraction to remove high molecular weight organic compoundspresent in samples, and then passed through an anion column(Makris et al., 2008). This extraction has been successfully usedin poultry litter water extracts amended with WTR (Makris et al.,2008). In brief, 1 mL of the filtered sample was mixed with toluene(2 mL) and 100 mM L�1 sodium perchlorate (1 mL) in a glass vial,shaken for 2 h at 120 rpm, and subsequently frozen for another2 h. Finally, the supernatant toluene was discarded, and the samplewas thawed and analyzed for As species. Samples were quantifiedfor As species (As(III), As(V), DMA, and MMA) according to the Asspeciation protocol by Jackson and Bertsch (2001) using High per-formance liquid chromatography coupled to an inductively-cou-pled plasma mass spectrometer (LC-ICPMS). Cacodylic acidsodium salt (DMAV) (MP Biomedics Inc., Germany, CAS #105645)and monomethyl arsenic acid (MMA) (Chem Service, USA) wereused to prepare the As stock solution at 100 mg L�1 in d-H2O. Stockstandard solutions for As(III) and As(V) (Spex Certiprep, NJ) werediluted to 100 mg L�1. The mobile phase for As speciation was10 mM L�1 NaH2PO4 in 1% CH3OH (adjusted to pH of 7.2). The mo-bile phase program was: 0 to 2.59 min – 10% of 10 mM L�1 NaH2-

PO4 in 1% CH3OH and 80% d-H2O in 1% CH3OH at a flow rate of1 mL min�1; 3–8.53 min: 100% 10 mM L�1 NaH2PO4 in 1% CH3OHat a flow rate of 1.7 mL min�1. The HPLC (Prostar 210, Varian,Inc., Palo Alto, CA) was coupled with ICPMS (Elan 9000, Perkin–El-mer). A 100 lL of sample was delivered to a Dionex column IonPacAS14 (4 � 50 mm) and its respective guard column by HPLC.

2.2. Statistical analysis

Statistical analysis was performed using JMP IN version 5.1 (Sallet al., 2005). Factorial (three-way ANOVA) analysis of variance wasperformed to examine the effect of WTR on As bioaccessibility(IVG-S) and geochemical As forms as a function of contact time,type- and rate-WTR for both soils. Pearson correlation coefficientswere generated using multivariate analysis to examine the rela-tionship between soil As fractionation and bioaccessibility. A TukeyHSD analysis was performed to evaluate the difference among thetreatment means for the amount of As in leachates. A student t-testwas done to examine the difference among the amount of As spe-cies fraction (%) in in vitro and water-extracted samples.

3. Results and discussion

3.1. Soil arsenic bioaccessibility: WTR effect

3.1.1. Immokalee soilThe two soils used in this study were selected based on their

distinct physico-chemical properties. Table 1 shows the physico-chemical properties of soils and WTR. Immokalee soil has loweramorphous Fe/Al content (0.04 g kg�1) when compared to Oreliasoil (0.8 g kg�1). Being sandy and lacking positively charged sur-faces (amorphous Fe/Al oxides), Immokalee soil is likely to haveminimal As retention capacity (Pierce and Moore, 1982). As dis-cussed earlier in Nagar et al. (2010), the organic matter for Al-and Fe-based WTR were 330 and 400 g kg�1, respectively, higherthan that of both soils (Elliott et al., 2002).

At time zero, no significant (p > 0.05) difference was found in Asbioaccessibility (as measured by IVG-S) between the unamended

Please cite this article in press as: Nagar, R., et al. Arsenic bioaccessibility andtreatment residuals based on a long-term greenhouse study. J. Hydrol. (2012),

and the Al-WTR-amended soil. This trend remained unchanged un-til 0.5 year for both application rates, i.e. 5% and 10% (Fig. 1A). Thiswas due to lack of soil WTR equilibration. The methyl substitutionmight lower the adsorption of DMA on oxide surfaces by decreasingspatial compatibility. Also, electrostatic attraction between posi-tively charged Al-hydroxides and negatively charged As species isexpected to be the highest for As V and the lowest for DMA (Laffertyand Loeppert, 2005; Shimizu et al., 2010). After 0.5 year of soilequilibration, the effect of Al-WTR amendment on soil As bioacces-sibility became significant (p < 0.001), with a decrease in bioacces-sibility to 80% compared to the unamended control (Fig. 1A). Theseresults are in accordance with a previous study by Ghosh and Yuan(1987), showing a significant removal of DMA by alumina (alumi-num oxide). Studies by Shimizu et al. (2010) showed formation ofbidentate binuclear complexes between MMA/DMA and aluminumoxides. A significant (p < 0.001) effect of the Al-WTR applicationrates on soil As bioaccessibility was observed only after 1 year,where As bioaccessibility decreased to 58% and 48% at 5% and10% application rates, respectively (Fig. 1A). These results are inagreement with a previous study on As(V) indicating that soil agingplays a major role in decreasing As dissolution from WTR particlesin an acidic gastric environment (Sarkar et al., 2007b; Nagar et al.,2009a, b). No significant (p > 0.05) decrease in As bioaccessibilitywas observed after 1 year, indicating that the WTR-amended soilsmight have reached equilibration (Fig. 1A).

In the case of Fe-WTR-amended soils, there was a significant(p < 0.001) decrease in As bioaccessibility (as measured by IVG-S)compared to unamended (no WTR) soil at time zero (Fig. 1A). Pre-vious studies showed a significant affinity of DMAV on the surfacesof hydrous ferric oxide by the possible formation of inner-spherecomplexes (Ghosh and Yuan, 1987; Lafferty and Loeppert, 2005).An EXAFS study by Shimizu et al. (2011) showed formation ofbidentate binuclear inner-sphere complexes between MMA/DMAand Fe-oxides surfaces of soil. Contrary to Al-WTR amended soil,Fe-WTR amended soils showed a significant (p < 0.001) effect ofapplication rates on decreasing soil As bioaccessibility at time zero(Fig. 1A). Arsenic bioaccessibility decreased to 80% and 70% at 5%and 10% application rates, respectively, compared to the una-mended control (Fig. 1A). These results were similar to previousstudies on As(V), which showed the greater effectiveness of Fe-WTR in resisting acidic conditions of the environment, and hence,in decreasing soil As bioaccessibility compared to Al-WTR (Sarkaret al., 2007b; Nagar et al., 2009a, b). Arsenic bioaccessibility furthersignificantly (p < 0.001) decreased with an increase in time, andreached 30% (at 10% application rate) compared to unamended soil(95%) after 1 year of equilibration (Fig. 1A). However, no significant(p > 0.05) decrease in bioaccessible As was reported after 1 year.

3.1.2. Orelia soilCompared to unamended Immokalee soil, the unamended

Orelia soil showed significantly (p < 0.001) lower As bioaccessibil-ity in DMA amended soil, which can be explained by the higher Al/Fe content of Orelia soil (Table 1). Arsenic bioaccessibility de-creased to 80% in unamended Orelia soil at the end of the 3-yearstudy (Fig. 1B). However, similar to Immokalee soil, there was nosignificant (p > 0.05) difference in gastric As bioaccessibility be-tween the unamended and the Al-WTR amended soil at time zero(Fig. 1B). With an increase in contact time to 0.25 year, As bioacces-sibility significantly (p < 0.001) decreased to 85% in Al-WTR-amended soil compared to the control, i.e. 100% bioaccessibility(Fig. 1B). However, there was no significant (p > 0.01) effect ofAl-WTR application rates observed until the year 1 (Fig. 1B). After1 year of equilibration time, As bioaccessibility decreased to 50%and 40% at 5% and 10% application rates, respectively (Fig. 1B).The effect of WTR in decreasing soil As bioaccessibility was signif-icantly (p < 0.001) higher in Orelia soil (with its clayey nature and

speciation in the soils amended with organoarsenicals and drinking-waterhttp://dx.doi.org/10.1016/j.jhydrol.2012.10.013

Table 1Selected physico-chemical properties of soils (Immokalee and Orelia series) and WTRs (Al and Fe based). Numbers are the means of three replicates ± standard deviation.

Immokalee Orelia Al-WTR Fe-WTR

pH 4.05 ± 0.48 7.85 ± 0.12 5.10 ± 0.34 5.40 ± 0.45ECa (lS cm�1) 59.0 ± 5.0 203 ± 13 363 ± 12.3 164 ± 12.5OMb (g kg�1) 8.40 ± 0.18 23.9 ± 1.5 330 ± 8.78 400 ± 9.23Ca/Mg (g kg�1) Total 0.24 ± 0.08 13.1 ± 1.1 4.27 ± 0.63 6.01 ± 0.54Fe (g kg�1) Total 0.05 ± 0.002 4.09 ± 0.52 12.3 ± 0.86 268 ± 9.63

Oxalate 0.04 ± 0.003 0.73 ± 0.01 5.70 ± 0.65 78.7 ± 8.65Al (g kg�1) Total 0.08 ± 0.001 5.42 ± 0.46 113 ± 8.9 1.34 ± 0.23

Oxalate 0.02 ± 0.001 0.20 ± 0.005 82.0 ± 8.3 0.36 ± 0.89As (g kg�1) Total 0.015 ± 0.001 0.012 ± 0.001 0.005 ± 0.002 0.006 ± 0.003Sand (%) 99.68 ± 0.0 63.76 ± 0.0 Nd NdClay (%) 0.57 ± 0.05 21.91 ± 0.60 Nd Nd

Nd – not determined.a Electrical conductivity.b Organic matter.

Time Period (Year)

0.0 0.5 1.0 1.5 2.0 2.5 3.0 3.5

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Fig. 1. Changes in As bioaccessibility (measured by IVG-S) with equilibration time(years) in Immokalee (A) and Orelia (B) soils amended with DMA and two rates (5%and 10%) of Al- and Fe-WTR. Control was soil spiked with DMA without WTRamendment. Data were the average of four replicates ± one standard deviation.

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Fig. 2. Effect of WTR on soil arsenic fractionation for Immokalee soil amendedwith DMA and two rates (5% and 10%) of Al- and Fe-WTR. Data are presentedfor Immokalee soil at 0 (A), 0.5 (B), and 3 years (C). Control was soil amendedwith DMA (no WTR). Data were the average of four replicates ± one standarddeviation.

4 R. Nagar et al. / Journal of Hydrology xxx (2012) xxx–xxx

higher Fe/Al content) compared to Immokalee (Table 1). However,similar to Immokalee, there was no change in As bioaccessibilityafter 1 year, indicating that the system reached an equilibrium(Fig. 1B).

Similar to Immokalee soil, Orelia soil showed a significant(p < 0.001) effect of Fe-WTR amendment on As bioaccessibilitycompared to the unamended control immediately after spiking(Fig. 1B). After 3 years, As bioaccessibility decreased to 20% (for10% application rate) compared to the unamended control, i.e.80% (Fig. 1B).

3.2. Sequential arsenic fractionation: WTR effect

3.2.1. Immokalee soilAt time zero, most of the As in Immokalee soil was extracted in

the soluble (water-extractable) fraction (Fig. 2A). The soluble form

Please cite this article in press as: Nagar, R., et al. Arsenic bioaccessibility and speciation in the soils amended with organoarsenicals and drinking-watertreatment residuals based on a long-term greenhouse study. J. Hydrol. (2012), http://dx.doi.org/10.1016/j.jhydrol.2012.10.013

Fe 5% Fe 10% Al 5% Al 10% Control

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Fig. 3. Effect of WTR on soil As fractionation for Orelia soil amended with DMA andtwo rates (5% and 10%) of Al- and Fe-WTR. Data were presented for Immokalee soilat 0 (A), 0.5 (B), and 3 years (C). Control was soil spiked with DMA without WTRamendment. Data represents the average of four replicates ± one standard devia-tion.

R. Nagar et al. / Journal of Hydrology xxx (2012) xxx–xxx 5

likely represents the As fraction that can be lost through runoff andleaching or is available to plants (Martin and Ruby, 2003; Novakand Watts, 2004). For DMA-spiked, unamended Immokalee soil,nearly 90% of the total As was present in the soluble form, whichwas expected due to the sandy nature and low Fe/Al content ofImmokalee soil (Table 1). A previous study in our laboratoryshowed that 89% of the total As was in the soluble fraction inImmokalee soil amended with 450 mg DMA kg�1 soil at time zero(Datta et al., 2006; Quazi et al., 2011). Tlustos et al. (2002) showedthat DMA remained in soil in a highly concentrated, soluble formcompared to i-As species. However, the addition of WTR decreasedthe soluble fraction to 65% and 50% at 5% and 10% application rates,respectively (Fig. 2A). There was no significant (p > 0.05) differencebetween the type of WTR at time zero (Fig. 2A). A significantly(p < 0.001) higher amount of As was in Fe/Al-bound fraction forWTR-amended soil compared to the unamended control, confirm-ing the affinity of DMA for Fe/Al hydroxides surfaces (Fig. 2A). Onlya small portion (<1%) of As was found in the remaining fractions,namely the Ca/Mg bound, organic bound and residual fractions(Fig. 2A).

There was a significant (p < 0.001) decrease in the soluble frac-tion of As, paralleling an increase in Fe/Al and Ca/Mg fractions withsoil aging (Fig. 2). These results corroborate a previous study in ourlaboratory, showing a significant effect of soil aging in DMAV-spiked soils (Datta et al., 2006). The effect of time, however, wasmore pronounced for WTR-amended soil than the unamended con-trol. After 0.5 year of equilibration time, the water-soluble fractionsignificantly (p < 0.001) decreased to 65% in unamended soil(Fig. 2B). At the same time, in WTR-amended soil, the soluble frac-tion decreased to 10–15% with a significant (p < 0.001) increase inFe/Al-bound As to 60–65% (Fig. 2B). These results confirmed thatamorphous Fe and Al content of WTR acts as strong As scavengersfor DMA-contaminated soils (Cheng et al., 2005). There was also asignificant (p < 0.001) increase in Ca/Mg-bound As in WTR-amended soil (10–20%) compared to the unamended control(Fig. 2B). However, the remaining fractions – namely the organicbound and residual fractions (contributing < 1%) – were not signif-icantly changed with time or WTR application (Fig. 2B).

This trend continued up to the 1 year of equilibration time,when the soluble fraction decreased to 6% in WTR-amended soilcompared to the unamended control, i.e. 53% (data not shown).No further significant (p < 0.001) change in As distribution was ob-served after the 1 year, indicating that an equilibrium might havebeen reached for soil aging and WTR effect (Fig. 2C).

3.2.2. Orelia soilFor unamended Orelia soil, the water-extracted fraction was

significantly (p < 0.001) lower than the unamended Immokalee soil(Fig. 3A). These results are similar to previous studies by Dattaet al. (2007) and Quazi et al. (2011). This could be explained bythe clayey nature, high Fe/Al hydroxides, and Ca/Mg content of Or-elia soil compared to Immokalee (Table 1). For unamended Oreliasoil, the As fraction associated with Fe/Al amorphous oxyhydrox-ides contributed to 23% of total As (Fig. 3A). However, WTR amend-ment significantly (p < 0.001) increased the Fe/Al-bound fraction to33–43% (Fig. 3A). Similar to Immokalee soil, there was no signifi-cant (p > 0.05) effect of type of WTR at time zero (Fig. 3A). Theremaining As fractions (Ca/Mg-bound, organic-bound, and resid-ual) contributed a very small portion (<1%) of total As (Fig. 3A).There was a significant (p < 0.001) effect of soil aging on bothWTR-amended and unamended soil, although this effect was morepronounced for WTR-treated soils.

At time final (year 3), the soluble As fraction in WTR-amendedsoil became negligible (2%) compared to total As, although it wasstill significant (30%) in the unamended control (Fig. 3C). The Fe/Al and Ca/Mg-bound forms immobilized As, and therefore possibly

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decreased the weakly bound (readily bioavailable) fraction inWTR-amended and DMA-contaminated soil (Wragg et al., 2007).

3.3. Correlation between bioaccessibility and fractionation of soilarsenic

The correlation between in vitro As (measured as IVG-S) and theAs extracted by different steps of sequential extractions was per-formed at all incubation time points and with all treatments forImmokalee and Orelia soils and shown in Table 2. BioaccessibleAs in Immokalee soil was significantly and positively correlatedwith the soluble (r = 0.80, p < 0.001) and exchangeable (r = 0.64,p < 0.001) fractions. Because WTR is the sink of Fe/Al hydroxides,Immokalee soil amended with WTR showed a significant negativecorrelation between Fe/Al (r = �0.78, p < 0.001)-bound As andbioaccessible As (Table 2). This led us to speculate that As bound

speciation in the soils amended with organoarsenicals and drinking-waterhttp://dx.doi.org/10.1016/j.jhydrol.2012.10.013

Table 2Pearson correlation coefficients for in vitro (IVG-S) and sequential arsenic fractions as a function of incubation time and treatments for both Immokalee and Orelia soil.

Soluble Exchangeable Fe/Al Ca/Mg Organic Residual IVG-S

ImmokaleeSoluble 1.00Exchangeable 0.62a 1.00Fe/Al �0.98a �0.64a 1.00Ca/Mg �0.77a �0.67a 0.68a 1.00Organic �0.65a �0.61a 0.60a 0.61a 1.00Residual �0.74a �0.45b 0.70a 0.60a 0.68a 1.00IVG-S 0.80a 0.64a �0.78a �0.66a �0.64a �0.67a 1.00

OreliaSoluble 1.00Exchangeable 0.83a 1.00Fe/Al �0.92a �0.83a 1.00Ca/Mg �0.84a �0.67a 0.56a 1.00Organic �0.36c �0.37c 0.28c 0.36c 1.00Residual �0.08 0.003 0.008 �0.22 0.16 1.00IVG-S 0.76a 0.78a �0.67a �0.72a �0.24 0.03 1.00

a Data were significantly correlated at p < 0.001 using multivariate analysis.b Data were significantly correlated at p < 0.01 using multivariate analysis.c Data were significantly correlated at p < 0.05 using multivariate analysis.

6 R. Nagar et al. / Journal of Hydrology xxx (2012) xxx–xxx

to Fe/Al hydroxides was unaffected by acidic stomach conditions.These results are in accordance with previous studies in ourlaboratory, where partial resistance of As-containing particles ina gastric environment was observed in DMA-spiked soils (Dattaet al., 2006; Sarkar et al., 2005). The degree of determination waslower but still significant (r = �0.45, p < 0.01) for the Ca/Mg-boundAs fraction and bioaccessible As (Table 2). Similar to Immokaleesoil, Orelia soil showed a significant (p < 0.001) positive correlationbetween IVG-S and the soluble/exchangeable fraction with coeffi-cient values of 0.76 and 0.78, respectively (Table 2).

Additionally, As associated with Fe/Al oxyhydroxides showed asignificant (p < 0.001) negative correlation (Table 2). However, bio-accessible As in Orelia soil showed a stronger negative correlation(r = �0.72, p < 0.001) than that found in Immokalee soil (Table 2).

3.4. Total arsenic in soil, leachates and plants: WTR effect

Table 3 shows the distribution of As in soil, plant, and leachatewater in the Immokalee and Orelia soil amended with DMA. Theamount of As (mg) in leachates represents the total As measuredfrom the leachates collected after 0.25 and 0.5 years. For the una-mended Immokalee soil, with its low extractable Fe/Al contentand sandy nature, the majority of As (85% of total) was lost inthe leachates (Table 3). These results support the previous findingsthat a lower amount of DMA is immobilized via the adsorptionmechanism compared to i-As species due to increased methyl sub-stitution (Cheng et al., 2005; Tlustos et al., 2002; Shimizu et al.,2010). However, WTR amendment significantly (p < 0.001) de-creased the downward movement of As through leachates, byincreasing the amount of Fe/Al hydroxides in Immokalee soil (Ta-ble 3). A previous study by Makris et al. (2008) also showed theeffectiveness of WTR in immobilizing organic As species from PLsuspension. WTR rate and type had no significant effect (p > 0.05)in deceasing the amount of As in leachates (Table 3). A similartrend was observed for Orelia soil, where the amount of As inleachates significantly (p < 0.001) decreased after WTR amend-ment (Table 3).

The total amount of As (lg) in the rice plants was calculated bymultiplying plant biomass (g) and As concentration (lg g�1) inplant tissue (Table 3). For both soils, total As (lg) in rice plantswas minimal (<0.1%) compared to the amount of As present in soiland leachates (Table 3). This might be the result of phytotoxicityand decreased biomass caused by a very high initial As concentra-

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tion (1500 mg kg�1) in the form of DMA. A previous hydroponicsstudy by Marin et al. (1993) also showed the inhibitory effect ofhigher concentrations (1.6 mg/L) of DMA on biomass and photo-synthesis of rice plants. High As concentrations resulted in de-creased plant biomass in control (no WTR) soils (Fig. 4). The WTRapplication, however, decreased the soluble (potentially plant-available) form of As in soil, which resulted in lower As per gramweight of the plant compared to the control (Fig. 4). Al-WTR wasmore effective than Fe-WTR (i.e. 50 lg g�1) in decreasing the Asconcentration (20 lg g�1) in rice plants (Fig. 4). Therefore, totalAs (on a weight basis) was lower (�2 lg) for plants grown in bothsoils after Al-WTR amendment (Table 4). In addition, compared tothe control (no WTR) soil, WTR amendment increased plant bio-mass, and this effect was more pronounced for Fe-WTR (Fig. 4).Iron (a component of Fe-WTR) is essential for the functioning ofa number of redox proteins in the electron transport chains of res-piration and photosynthesis. The increased biomass resulted inhigher total As (lg) in Fe-WTR-amended soils (Table 4). Al-WTRtreatment to the DMA-spiked soils caused significantly(p < 0.001) low plant growth compared to Fe-WTR (Fig. 4), whichmight be due to partial Al-toxicity caused by Al-WTR application(Heil and Barbarick, 1989; Rengaswamy et al., 1980). The percentrecoveries (calculated from the sum total of As accumulated in rice,lost in the leachates, and remaining in the soil) of As in this studyranged between 76% and 91% for Immokalee and 71% and 83% forOrelia soil (Table 3).

3.5. Arsenic speciation studies: DMA transformation

Arsenic speciation analysis was performed for water as well asin vitro-extracted samples using LC-ICPMS. Results showed that forboth extractions, DMA transformed to the more toxic i-As(V) bythe end of 3 years, regardless of the presence of WTR (Fig. 5, Ta-ble 4). There was no significant (p < 0.001) difference in % As spe-cies released from water or in vitro-extracted samples (Table 4).This supported the previous findings in our laboratory that thein vitro experimental conditions did not change the As oxidationstate when As was added as DMA (Makris et al., 2008). DMA trans-formation to the more toxic i-As occurred on soil or WTR particlesurfaces, and microbial (biotic) degradation may be a possible sce-nario (Feng et al., 2005; Makris et al., 2008; Shimizu et al., 2011).DMA transformation into As(V) was observed in both soils,although Orelia soil showed a significantly (p < 0.001) higher trans-

speciation in the soils amended with organoarsenicals and drinking-waterhttp://dx.doi.org/10.1016/j.jhydrol.2012.10.013

Table 3Mass balance of arsenic (As) in Immokalee and Orelia series soils. The percent recoveries calculated from As remaining in the soil, As in the leachates, and As accumulated in riceplants. Values are mean (± standard deviation) of four replicates.

As amount (mg/soil column)

Soil type Soiltreatment

Time-zero Time-final (3rd Year)

Initial soil As– theoritical (mg)

Initial soil Asa

–expérimental (mg)As remainingin soila (mg)

As inrice (lg)

As inleachates (mg)

Sum (mg) Percentrecovery (%)

Immokalee Fe WTR-5% 5550 5594 ± 167 2767 ± 143 32.9 ± 4.4 1439 ± 204 BC 4229 75.6Fe WTR-10% 5550 5826 ± 418 3216 ± 241 7.25 ± 2.8 1219 ± 241C 4439 76.2Al WTR-5% 5550 5586 ± 75.5 2714 ± 340 3.35 ± 1.6 1444 ± 233 BC 4163 74.5Al WTR-10% 5550 5857 ± 181 3290 ± 243 7.19 ± 0.09 1114 ± 106 C 4409 75.30% WTR 5550 5741 ± 172 722.7 ± 70.9 11.29 ± 0.4 4188 ± 335A 4914 85.6

Orelia Fe WTR-5% 5550 5675 ± 195 3036 ± 506 24.38 ± 0.5 1425 ± 146 BC 4463 78.6Fe WTR-10% 5550 5948 ± 286 4012 ± 456 10.83 ± 0.4 942.3 ± 81.1C 4960 83.4Al WTR-5% 5550 5725 ± 189 2520 ± 238 2.95 ± 0.3 1617 ± 165 BC 4140 72.3Al WTR-10% 5550 5920 ± 366 2986 ± 394 4.64 ± 2.5 1478 ± 124 BC 4469 75.50% WTR 5550 5880 ± 331 846.5 ± 148 2.05 ± 1.1 3373 ± 197A 4221 71.8

Levels in the leachates data not connected with the same letter are significantly different.a Total digestion (EPA 3050B).

0

50

100

150

200

250

Fe5% Fe10% Al5% Al10% Control

Bio

mas

s (u

g)

0

50

100

150

200

250

300

As

conc

entr

atio

n (u

g/g)

biomass (ug)

As conc (ug/g)

0

50

100

150

200

250

300

Fe5% Fe10% Al5% Al10% Control

Bio

mas

s (u

g)

0

20

40

60

80

100

120

140

160

As

conc

entr

atio

n (u

g/g)

biomass (ug)

As conc (ug/g)

A

B

Fig. 4. Effect of WTR on plant biomass (lg) and As concentration (lg/g plant tissue), grown in both Immokalee (A) and Orelia (B) soils at the initial As concentration of1500 mg As kg�1. Data represented the average of four replicates ± one standard deviation.

R. Nagar et al. / Journal of Hydrology xxx (2012) xxx–xxx 7

formation than Immokalee soil (Table 4). For Orelia soil, almost allDMA (95%) transformed to As(V), while in Immokalee soil, 75% ofAs was in the As(V) form (Table 4). These results were consistentwith previous studies by Feng et al. (2005), showing higher organ-oarsenical transformation to As(V) in clayey soil than in sandy soil.

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The longer the As remained on the substrate, the more likely it wasto be biotically transformed to i-As (Feng et al., 2005). However,there was no overall significant (p < 0.001) effect of WTR on DMAtransformation. Therefore, studies with more sampling betweentime zero to 3 years are required to better understand the effect

speciation in the soils amended with organoarsenicals and drinking-waterhttp://dx.doi.org/10.1016/j.jhydrol.2012.10.013

Table 4Organoarsenical transformation in WTR-amended and unamended Immokalee and Orelia soil at time zero and time final (3 year). Data shown as % species fraction of total Asextracted by in vitro and water extractions.

Treatments Time 0 Time final

In vitro As (% fraction) Water-soluble As (% fraction) In vitro As (% fraction) Water-soluble As (% fraction)

Immokalee soil As(V) DMA As(V) DMA As(V) DMA As(V) DMA

Fe-WTR (5%) 0 100 0 100 88.0DE 12.0HI 83.6D 16.4H

Fe-WTR (10%) 0 100 0 100 75.5F 24.5G 76.3DF 23.7GH

Al-WTR (5%) 0 100 0 100 87.9DE 12.1HI 83.9D 16.1H

Al-WTR (10%) 0 100 0 100 85.6E 14.4H 82.7DE 17.3HI

No WTR 0 100 0 100 84.1E 15.9H 86.7CD 13.3IJ

Orelia soilFe-WTR (5%) 0 100 0 100 93.8BC 6.2JK 90.6BC 9.4JK

Fe-WTR (10%) 0 100 0 100 92.0BC 8.0JK 95.9AB 4.1KL

Al-WTR (5%) 0 100 0 100 98.1AB 1.9KL 95.3AB 4.7KL

Al-WTR (10%) 0 100 0 100 99.3A 1.7L 94.1AB 5.9KL

No WTR 0 100 0 100 96.0AB 4.0KL 98.0A 2.0L

Time 0

3rd Year

DMA

DMA

As (V)

Fig. 5. LC/ICP-MS chromatograms showing DMA transformation into As(V) for a sample (Orelia soil amended with 10% Fe-WTR) at time 0 and year 3. Sample injection –100 mL. Time (X-axis) in seconds.

8 R. Nagar et al. / Journal of Hydrology xxx (2012) xxx–xxx

of WTR on organoarsenical transformation. Still, the current resultsshowed that DMA transformation to more toxic As(V) occurredregardless of the DMA binding to soil, or WTR particles or beingdissolved in the soil solution.

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4. Conclusion

Previous studies in our laboratory showed that WTR was aneffective sorbent for i-As. The current study demonstrated that this

speciation in the soils amended with organoarsenicals and drinking-waterhttp://dx.doi.org/10.1016/j.jhydrol.2012.10.013

R. Nagar et al. / Journal of Hydrology xxx (2012) xxx–xxx 9

waste by-product (WTR) was similarly effective in significantly(p < 0.001) immobilizing As, hence decreasing As bioaccessibility,in DMA-contaminated soil over a 3-year study period. There wasa significant (p < 0.001) decrease in water-soluble As with timeaccompanied by an increase in Fe/Al and Ca/Mg fractions, showingthat these phases controlled the mobility of As. Repeated applica-tion of DMA to agricultural soils, especially those used for rice cul-tivation, may increase As buildup, which would lead to plantuptake and subsequent transfer of As to the human food chain orleaching to the groundwater. The current study, however, showedthat the amendment with WTR could minimize As mobility, there-fore reducing the risk associated with leaching and plant uptake ofAs. However, studies with lower As concentrations and WTR appli-cation rates are required to calibrate WTR use for optimum plantgrowth and decreased phytoavailability. DMA transformation intoAs(V) was observed in both soils, although Orelia soil showed sig-nificantly higher transformation than did Immokalee soil. Althoughthere was no overall significant effect of WTR amendment on DMAtransformation, further studies with more sampling between timeszero and year 3 are required.

Acknowledgements

The authors would like to thank NIH-SCORE, SALSI, and USEPA-STAR programs for funding this study.

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