Science of the Total Environment 438 (2012) 286–292

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Arsenic species in broiler (Gallus gallus domesticus) litter, soils, maize (Zea mays L.),and groundwater from litter-amended fields

Elisa D'Angelo a,⁎, Georgia Zeigler a,1, E. Glenn Beck b,2, John Grove a,3, Frank Sikora c,4

a University of Kentucky, N-122 Agricultural Science Building North, Lexington, KY 40546, United Statesb Kentucky Geological Survey, Western Kentucky Office, 1401 Corporate Court, Henderson, KY 42420, United Statesc University of Kentucky, Rm 135, 1600 University Drive, Lexington, KY 40546, United States

H I G H L I G H T S

► Broiler litter from different integrators contained divergent amounts and species of bioavailable arsenic.► The organoarsenical roxarsone was largely transformed to arsenate in litter.► Arsenate sorption in soils was strongly correlated to Fe and Al oxide contents.► Soil sorption reduced the mobility and bioavailability of arsenate in groundwater and maize.► The environmental fate of As depends on the As species in litter and chemical properties of soils.

⁎ Corresponding author. Tel.: +1 859 257 8651; fax:E-mail addresses: edangelo@uky.edu (E. D'Angelo), g

ebeck@uky.edu (E.G. Beck), jgrove@uky.edu (J. Grove),1 Tel.: +1 859 257 2859; fax: +1 859 257 3655.2 Tel.: +1 270 827 3414x23; fax: +1 270 827 1117.3 Tel.: +1 859 257 5852; fax: +1 859 257 3655.4 Tel.: +1 859 257 2785; fax: +1 859 257 7351.

0048-9697/$ – see front matter © 2012 Elsevier B.V. Allhttp://dx.doi.org/10.1016/j.scitotenv.2012.08.078

a b s t r a c t

a r t i c l e i n f o

Article history:Received 22 May 2012Received in revised form 21 August 2012Accepted 21 August 2012Available online 23 September 2012

Keywords:RoxarsoneArsenateSorption isothermsLysimeterBioavailability

Manure and bedding material (litter) generated by the broiler industry (Gallus gallus domesticus) often con-tain high levels of arsenic (As) when organoarsenical roxarsone and p-arsanilic acid are included in feed tocombat disease and improve weight gain of the birds. This study was conducted to determine As levels andspecies in litter from three major broiler producing companies, and As levels in soils, corn tissue (Zea maysL.), and groundwater in fields where litter was applied. Total As in litter from the three different integratorsranged between b1 and 44 mg kg−1. Between 15 and 20% of total As in litter consisted of mostly of arsenate,with smaller amounts of roxarsone and several transformation products that were extractable with phos-phate buffer. Soils amended with litter had higher levels of bioavailable As (extractable with Mehlich 3 solu-tion and taken up by corn leaves). Arsenic concentrations in plant tissue and groundwater, however, werebelow theWorld Health Organization thresholds, which was attributed to strong sorption/precipitation of ar-senate in Fe- and Al-rich soils. Ecological impacts of amending soils with As-laden litter depend on the Asspecies in the litter, and chemical and physical properties of soil that strongly affect As mobility and bioavail-ability in the environment.

© 2012 Elsevier B.V. All rights reserved.

1. Introduction

For several decades, roxarsone (4-hydroxy-3-nitrobenzenearsonicacid) has been approved for use in the feed of broilers (Gallus gallusdomesticus) and swine (Sus scrofa domesticus) to control coccidiosis,speed weight gain, and improve meat color (Chapman and Johnson,2002). As of July 2011, the use of roxarsone was banned in the UnitedStates, after it was discovered that potentially dangerous amounts ofAs accumulated in meat tissue, particularly in young broilers (up to

+1 859 257 3655.mzeig2@uky.edu (G. Zeigler),fsikora@uky.edu (F. Sikora).

rights reserved.

0.45 mg As kg−1) (Lasky et al., 2004). However, roxarsone is stillavailable for use in several other countries, including Argentina, Aus-tralia, Brazil, Canada, Chile, Indonesia, Jordan, Malaysia, Mexico, Paki-stan, Philippines, Venezuela, and Vietnam.

Although small amounts of As fed to broilers can accumulate in ani-mal tissue, larger amounts are excreted and accumulate in the litter,which is the floor material in broiler houses that consists of a mixtureof feed,manure, and beddingmaterial, such as sawdust,wood shavings,or rice hulls (Anderson and Chamblee, 2001; Jackson et al., 2003). In2010, broiler industries in the United States generated 5 to 6 millionMg of dry litter (Coufal et al., 2006; National Agricultural StatisticsService, 2011), which contained between 1 and 77 mg As kg−1

(Anderson and Chamblee, 2001; Jackson et al., 2003).One of the main challenges facing the broiler industry is how to

efficiently manage and dispose of large amounts of litter. Currently,the main strategy for disposing broiler litter is application to pastures

287E. D'Angelo et al. / Science of the Total Environment 438 (2012) 286–292

and crop lands that are located near the production facilities (Bolan etal., 2010). This practice has the added benefit of improving soil fertil-ity, since the broiler litter contains high concentrations of several keymacronutrients N, P, and K (Coufal et al., 2006). However, applicationof litter to soils can also have detrimental environmental effects, suchas eutrophication of surface water bodies when excess nutrients aremobilized in surface runoff. Moreover, amendment of soils with litterthat contains high levels of potentially toxic arsenicals can also bemobilized to nearby water bodies, or taken up by plants grown onthe litter amended-soils (Jones, 2007; Morrison, 1969; Walsh et al.,1977). These concerns seem justified in light of reports that showedconsiderable amounts of As in litter (35–75%) were readily extractedwith water or other weak salt solutions that could be mobilized inrainwater or taken up by plants once litter is distributed to soils(Han et al., 2004; Jackson et al., 2003; Rutherford et al., 2003). Onthe other hand, Arai et al. (2003) reported that b15% of As in littercould be desorbed with water at either pH 4.5 or 7, suggesting thatonly low amounts of As in the litter would be mobilized in the envi-ronment. One explanation for the wide range in As solubilities couldbe differences in As species present in the litter at the time that itwas collected. As mentioned previously, roxarsone fed to birds islargely excreted and accumulates in freshly-produced litter. In ashort period (less than one month), however, much of the roxarsonecan be converted to other organic and inorganic degradation interme-diates that can form insoluble phases and/or be strongly retained inthe litter (Arai et al., 2003; Garbarino et al., 2003; Jackson et al.,2003; Seiter, 2009). Since litter at commercial operations is usuallyrecycled and may not be entirely cleaned out for several years(Malone, 1992), it is likely that litter will contain a multitude of Asspecies with variable water solubilities that could affect their fateonce applied to soils.

Once litter is added to soils, As can undergo several additionalbiogeochemical processes, including transport to surface and ground-water bodies (Hancock et al., 2001; Rutherford et al., 2003), degrada-tion (Cortinas et al., 2006; Garbarino et al., 2003), and sorption/coprecipitation (Bothe and Brown, 1999; Bowell, 1994; Brown et al.,2005; Cornu et al., 2003; Grafe et al., 2002; Violante et al., 2006).Differences in the extent of these processes likely account for thewide range in As accumulation levels in soils that have been reported(Arai et al., 2003; Ashjaei et al., 2011; Morrison, 1969; Rutherford etal., 2003). Among these processes, sorption is particularly importantbecause it governs the concentration of As in the dissolved pool,and therefore largely dictates the extent of the other As fate process-es. In studies that have evaluated As sorption in soils, sorption wasfound to be most highly correlated with crystalline and non-crystalline Fe and Al oxides, organic matter, and clay contents(Brown et al., 2005; Sahu et al., 2011; Shimizu et al., 2011; Wasay etal., 2000; Wilson et al., 2010). Arsenic sorption also appears to beinfluenced by levels of several dissolved constituents in litter, suchas concentrations of H+, organic matter, phosphate, and Ca2+. Withincreasing pH, for example, arsenate sorption tends to be reduced(Fendorf et al., 1997). Therefore, the addition of broiler litter, whichtypically has an alkaline pH, would be expected to mobilize As insoils. High levels of dissolved organic matter and phosphate in litterwould also be expected to increase As bioavailability and mobilityin soils by competing with As for common sorption sites (Bauer andBlodau, 2006; Jackson et al., 2006; Wang and Mulligan, 2006). Theeffect of these constituents on arsenate mobility, however, could bepartially counteracted by elevated dissolved Ca2+, which can formseveral types of calcium arsenate precipitates, or sorb onto Al:Fehydroxide surfaces and reduce negative charges that would favorarsenate sorption (Wilkie and Hering, 1996; Bothe and Brown,1999; Cornu et al., 2003).

The primary goals of this research were to further knowledgeabout (i) the levels and species of As in litter from three major broilerproducing companies (integrators), (ii) determine the amounts of As

in soils, corn (Zea mays L.) leaves and grain, and groundwater inagricultural plots that have been amended with broiler litter, and(iii) determine relationships between soil properties and arsenatesorption constants in whole soils.

2. Materials and methods

2.1. Collection and analysis of broiler litter from different integratorfarms

Broiler litter was collected from 15 broiler houses on differentfarms that represented three major broiler integrators in Kentuckyin June 2006 (five houses from each of the three integrator compa-nies, designated as A, B, and C). From each house, samples werecollected from 20 randomly selected locations using a plastic shovelto a depth of 15 cm, which were then mixed in equal amounts toprepare one composite sample that represented litter from thathouse.

In the lab, litter samples were air-dried and ground to pass a twomm mesh in a Wiley Mill, and then evaluated for different forms ofAs and other elements to estimate their potential bioavailability andmobility. Total As, P, Fe, Al, Cu, Ca, Mg, Mn, K, and Zn were deter-mined by digesting 0.100 g litter with 3 mL HNO3 and heating over-night at 105 °C. After cooling, 3 mL of 50% H2O2 was added, and themixture was heated at 120 °C for 1 h. After cooling, distilled waterwas added to a volume of 35.00 mL, and the diluted samples werefiltered with Whatman No. 42 paper. Samples were analyzed formetals andmetalloids using a Varian Vista-PRO Inductively CoupledArgon Plasma (ICP) (Palo Alto, CA) at the University of KentuckyRegulatory Services Soil Testing Lab.

Mehlich 3 extractable elements in soils were also determinedbecause it more accurately reflects bioavailable concentrations ofmultiple elements in both acid and neutral soils (Mehlich, 1984).Mehlich 3 extractable As, Ca, Cu, Fe, K, Mg, Mn, P and Zn were deter-mined by extracting 2.00 cm3 soil with 20.00 mL of Mehlich 3 solu-tion, which contains 0.2 N acetic acid, 0.25 N NH4NO3, 0.015 NNH4F, 0.013 N HNO3, and 0.001 N EDTA. The soil-extract mixturewas shaken for 5 min, and immediately filtered through Whatman#2 filter paper. The filtrate was analyzed via ICP at the UK RegulatoryServices Soil Testing Lab. Concentrations were converted from soilvolume to dry soil mass basis (mg kg−1) using soil density.

Phosphate buffer extractable As species (e.g. As(III), As(V),monomethylarsonic acid (MMA), dimethylarsinic acid (DMA), roxarsone,p-arsanilic, phenylarsonic acid, 3-amino-4-hydroxyphenylarsonic acid,and 4-hydroxy phenylarsonic acid) were determined by extracting1.00 g litter (composited from the five integrator farms) with 25.00 mLof 0.1 M sodium phosphate in polypropylene tubes for 20 h, filteringextracts with a 0.45 μm membrane, and analyzing filtered extracts byion chromatography-ICP-DRC-MS, which was conducted by AppliedSpeciation and Consulting (Bothell, WA). Standards for calibrating theinstrument for the various As species included certified As standardNIST 1640 (National Institute of Standards and Technology, Gaithersburg,MD), as well as standards from Alfa Aeasar (Ward Hill, MA) for DMA,Thermo Fisher Scientific (Pittsburgh, PA) for As(III) and As(V), ChemService (West Chester, PA) for MMA, Acros (Waltham, MA) forp-arsanilic and roxarsone, Avocado Research Chemicals (Ward Hill, MA)for phenylarsonic acid, Pfaltz and Bauer (Waterbury, CT) for 3-amino-4-hydroxyphenylarsonic acid, and ABCR Chemicals (Mt. Pleasant, SC)for 4-hydroxy phenylarsonic acid. The minimum detection limits forthese species ranged between 0.01 and 0.04 mg kg−1 litter, as indicatedin Table 1.

Total carbon was determined using a C/N elemental analyzer(Elemantar Vario Max, Hanau, Germany) at the UK Regulatory ServicesSoil Testing Lab. Litter pH was determined by equilibrating 2.00 g litterwith 10.00 mL of water for 15 min andmeasuring pH of the supernatantwith a calibrated pH meter and electrode.

Table 1Selected chemical properties of broiler litter from three integrators, designated as A, B,and C. Except for the phosphate extractable As species, each value represents the meanof five samples±one standard deviation. Values in the same row followed by a differ-ent letter are significantly different at a p value of ≤0.05. Values for the phosphate ex-tractable As species represent amounts in composite samples from the five farms.

Property Integrator A(n=5)

Integrator B(n=5)

Integrator C(n=5)

pvalue

pH 8.0a±0.3 8.4ab±0.5 8.7b±0.1 0.034% C 31±4 28±3 29±2 0.283

Total elements (mg kg−1)As 7.4a±3.8 0.6a±0.7 44b±11 0.000Cu 410a±64 580ab±150 730b±120 0.004P 14000a±

130018000ab±260

19000b±4400

0.023

Mehlich III (mg kg−1)As 4.3a±2.3 0.5a±0.3 25b±9 0.000Cu 170a±15 180a±51 310b±43 0.000P 9500a±700 12000b±

170013000b±1500

0.002

Ca 6600b±1300

3800a±2100

9900c±2200 0.001

Fe 110±18 160±51 140±9 0.053Zn 370±36 360±51 360±46 0.942

Phosphate extractableAs species (mg kg−1)

Total 1.17 0.23 8.67DMA 0.11 0.05 0.51As(III) b0.02 b0.02 b0.02MMA 0.04 b0.01 0.20p-Arsanilic 0.03 b0.02 0.25Phenylarsonic acid 0.02 b0.02 0.10As(V) 0.84 0.18 5.023-Amino-4-hydroxyphenyl-arsonic acid

b0.02 b0.02 0.11

4-Hydroxyphenyl-arsonicacid

b0.02 b0.02 0.58

Roxarsone 0.14 b0.04 1.91

288 E. D'Angelo et al. / Science of the Total Environment 438 (2012) 286–292

2.2. Soil sampling and characterization

Field experiments were conducted on 20, 3.7 m by 9.1 m plots thatwere equipped with 0.6 m by 0.9 m stainless steel tension-free pan ly-simeters installed at a depth of 90 cm in the B horizon of the soil profiles.The soil series in the plots was Maury silt loam, which formed in thinloess over residuum of Ordovician phosphatic limestone. The plotswere located at the College of Agriculture Spindletop Agricultural Exper-iment Station Farm near Lexington, Kentucky. For a more detailed de-scription of the study site and lysimeter installation, see Stoddard et al.(1998).

The 20 plots were arranged in a randomized complete block de-sign with five replications and four treatments, including (i) no tillminus litter (NT−L), (ii) no till plus litter (NT+L), (iii) chiselplowing plus secondary disking minus litter (T−L), and (iv) chiselplowing plus litter (T+L). Litter from Integrator C, which wasfound to contain high levels of As, was hand-applied to the soil sur-face of +L treatments at a rate of 5 Mg ha−1 in Spring 2007, 2008,and 2009. In the tillage treatments, tillage was conducted one weekafter applying litter.

Soils from the 0–5, 5–10, 10–20, 20–30, 30–40, 40–50, 50–60,60–70, 70–80 and 80–90 cm depth increments were collected usinga tractor-mounted hydraulic soil probe in Fall, 2007. Soil sampleswere air-dried and ground using a Spex 8000M Mixer Mill (SPEXCertiPrep, Metuchen, NJ). Soils were evaluated for total and Mehlich3 extractable elements using the same procedures as broiler litterdescribed above.

2.3. Corn tissue sampling and analysis

Corn leaves, collected at the beginning of the reproductive stage(R1 stage), and grain were sampled from the experimental plots inthe Fall 2007 and 2008. From each plot, ten samples from differentplants were randomly collected and composited to represent onefor each of the 20 plots. Samples were air-dried and ground to2 mm using a Wiley Mill before analyzing for total As by acid diges-tion using the same procedure as used for the broiler litter.

2.4. Groundwater sampling and analysis

Water that leached through the soil profile was collected fromlysimeter pans after each rain event that resulted in water in thepans between July 2007 and April 2009 (28 events). Samples werecollected using a hand-held plastic rotary pump, which was rinsedwith the first 200 mL of leachate to decontaminate the sample collec-tion system. The samples were collected in 20 mL scintillation vialsand were preserved with 50 μL of 25% H3PO4 to achieve 10 mMH3PO4 in the final solution, and stored at 4 °C. Arsenic in water sam-ples was analyzed using an 8807 Varian Spectra Graphite FurnaceAtomic Absorption Spectrometer (Palo Alto, CA) that was calibratedwith certified As reference standard solutions from Fisher Scientific(Item number SA449-100, Pittsburgh, PA) at the University of Ken-tucky Environmental Research and Teaching Laboratory. The mini-mum detection limit of the instrument was 5 μg L−1.

2.5. Sorption isotherms

Batch isotherms were conducted by equilibrating soils (0.25 g)from the surface (0–5 cm) and subsurface (80–90 cm) of four plotswith 25.00 mL of arsenate solutions with 0, 1 and 5 mg As L−1

in poly-allomer tubes for 24 h in the dark on a horizontal shakerat 180 osc min−1. A no-soil control using the 5 mg As L−1 solutionwas included to account for As losses by sorption to tubes orother processes. After the equilibration period, the samples werecentrifuged at 9400 g for 15 min, and the supernatants were filteredwith a 0.45 micron syringe filter and stored at −20 °C until analysison a Varian Vista Pro ICP (Agilent Technologies, Inc., Santa Clara,CA). The amount of As sorbed by the soils was determined from thedifference in the amounts of As in the final and initial solutions. Sorp-tion constants were determined by best-fit linear regression analysisbetween the equilibrium concentrations of As in solution and sorbedphases.

2.6. Statistical analysis

Statistical differences in concentrations of As in litter, soils, planttissue, and water in the various treatments were determined by twoway analysis of variance with tillage and litter amendments as mainfactors, using a significance level of 0.05. When significant differenceswere found, data were evaluated using Tukeys Honestly SignificantDifference (HSD) test. All statistical analyses were performed usingSTATGRAPHICS Plus Version 5.0 by StatPoint Technologies, Inc.,Warrenton, Virginia.

3. Results and discussion

Total As in the litter from the different farms varied widely (0.6 to43.8 mg kg−1) (Table 1). The concentration range was similar to thatreported in litter collected from three other states in the southeasternUS (Jackson et al., 2003). Total As was strongly dependant on theintegrator farm it originated from: Integrator C farms contained thehighest amounts of total As, Integrator A farms had intermediatelevels of As, and Integrator B farms had the lowest amounts of As.This was attributed to different amounts of organoarsenicals included

289E. D'Angelo et al. / Science of the Total Environment 438 (2012) 286–292

in broiler feed by the different integrators. Litter from the variousfarms also had significantly different pH, Cu, P, and Ca, which also sig-nified differences in the feed composition (Table 1). Total As concen-trations in all litter samples were below the ceiling limit of75 mg As kg−1 for land application of biosolids (US EPA, 2010), butthe As level in litter from Integrator C farms exceeded the monthlyaverage limit of 41 mg As kg−1, signifying that cumulative As loadingrates would need to be closely monitored in fields receiving thismaterial.

In litter from Integrator A and C farms, considerable amounts of Ascould be extracted with phosphate buffer (15–20%) and Mehlich 3solution (56–58%) (Table 1). Although these two different extractantsremoved different amounts of As, results from both extractions indi-cated that considerable amounts of As in the litter could potentiallybe taken up by plants or mobilized once applied to soils. The fractionof total As that could be readily extracted with water or other weaksalt solutions reported in other studies was found to range from0 to >70% (Arai et al., 2003; Garbarino et al., 2003; Han et al., 2004;Jackson et al., 2003). A likely explanation for the wide range inamounts of readily extractable As observed in various studies couldbe differences in As species present in the litter when it was collected.In freshly produced litter, highly water soluble roxarsone would beexpected to be the dominant As species, since most of what is fed tothe birds is excreted in the manure (Garbarino et al., 2003). However,if litter is allowed to accumulate in the houses for very long, as it is inmost broiler production facilities (Malone, 1992), then much of theroxarsone can be transformed to arsenate and other As species(Garbarino et al., 2003; Jackson et al., 2003; Seiter, 2009). Resultsfrom this study showed that the major phosphate buffer extractableAs species in litter were arsenate (58–72%), roxarsone (11–22%),

0

15

30

45

60

75

90

0 2 4 6 8 10 12 14

Total As (mg kg-1)0 10000 20000 30000 4000

Total Ca (mg kg-1)

0

15

30

45

60

75

90

0 20000 40000 60000 80000

Total Fe (mg kg-1)0 2000 4000

Total Mg (mg kg-1)

0

15

30

45

60

75

90

0 2000 4000 6000

Total Mn (mg kg-1) Total Al (mg kg-1)0 20000 40000 6000

So

il D

epth

(cm

)

Fig. 1. Total element concentrations in the profile of soils in 2007 under four tillage and litt(NT,−L), till plus litter (T, +L) and till minus litter (T,−L). Each marker represents the meaDifference test.

DMA (6–9%), 4-hydroxyphenylarsonic acid (0–7%), p-arsanilic acid(3%),MMA (2–3%), phenylarsonic acid (1–2%), and 3-amino-4-hydroxyphenylarsonic acid (0–1%) (Table 1). Conversions of roxarsone to thesespecies could explain why a considerable fraction of total As in litter(42–85%) could not be extracted with phosphate buffer or Mehlich 3solution, since arsenate can strongly interact with Ca, Cu, and Fe (Araiet al., 2003; Bothe and Brown, 1999) or form arsenstruvite mineralwith the formula NH4MgAsO4·6H2O in litter (Seiter, 2009).

Litter contained considerable amounts of As and several otherelements (Table 1), so it was not surprising that soils amended withthis material would also have high amounts of these elements.Indeed, the concentrations of several elements, including total organ-ic C, Cu, Zn, and pH, and Mehlich 3 Ca, Cu, Fe, K, P, and Zn were signif-icantly higher in the surface of litter-amended soils (Figs. 1 and 2),which is in accordance to what was reported by others (Ashjaei etal., 2011; Codling et al., 2008; Daigh et al., 2009). Litter-amendedsoils, however, did not have significantly higher amounts of total Asthan the non-amended soils (Fig. 1), which was in line with resultsfrom some studies (Arai et al., 2003; Morrison, 1969), but not others(Ashjaei et al., 2011; Codling et al., 2008; Daigh et al., 2009; Han et al.,2004).

A likely explanation for our inability to detect significant increases intotal As in litter-amended soils was that native soils already containedconsiderable amounts of total As (between 7 and 13 mg As kg−1 drysoil) (Fig. 1). Background As in soils observed in this study were withinthe range of 1.6–32.3 mg As kg−1 that were reported for 103 surfaceand subsurface Kentucky soils (Vosnakis et al., 2009). Broadcast applica-tion of 5 Mg litter ha−1 with 44 mg As kg−1 to the top 3 cm of a soilwith a bulk density of 1.2 Mg m−3, for example, would increase totalAs in the surface horizon from a background level of 7 mg As kg−1 to

0 0 10 20 30 40

NT, +L

NT, -L

T, +L

T, -L

Total Cu (mg kg-1)0 2000 4000 6000

Total K (mg kg-1)

0 10000 20000 30000

Total P (mg kg-1)0 50 100 150

Total Zn (mg kg-1)

0

pH5 5.5 6 6.5

Total C (%)0 1 2 3

er management combinations, including no-till plus litter (NT, +L), no-till minus littern of five replications and error bars were determined using Tukeys Honestly Significant

0 1000 2000 3000

Meh3 Ca (mg kg-1)

0

15

30

45

60

75

90

0 50 100 150 200

Meh3 Mn (mg kg-1)

0 200 400

Meh3 Mg (mg kg-1)

0

15

30

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0 100 200

Meh3 Fe (mg kg-1)

So

il D

epth

(cm

)

0

15

30

45

60

75

90

0.00 0.05 0.10

Meh3 As (mg kg-1) 0 2 4 6 8

Meh3 Cu (mg kg-1)

0 200 400 600

Meh3 K (mg kg-1)

0 100 200 300 400

Meh3 P (mg kg-1)0 10 20

Meh3 Zn (mg kg-1)

NT, +L

NT, -L

T, +L

T, -L

Fig. 2.Mehlich 3 extractable elements in the profile of soils under four tillage and litter management combinations, including no-till plus litter (NT, +L), no-till minus litter (NT,−L),till plus litter (T, +L) and till minus litter (T,−L). Each marker represents the mean of five replications and error bars were determined using Tukeys Honestly Significant Differencetest.

Table 2Total arsenic in grain and leaves of corn grown in broiler litter amended andnon-amended soils under no till and tillage management in 2007 and 2008. Eachvalue represents the mean As in composite samples taken from five replicate plots.Values in a row followed by a different letter are significantly different at a p value of≤0.05.

Plant tissue Year Till No-till p value

+Litter −Litter +Litter −Litter

μg kg−1

Grain 2007 6.4 5.0 5.8 6.2 0.3292008 6.2 5.8 6.2 5.0 0.310

Leaf 2007 22b 17a 29c 22b 0.0242008 27c 24b 22b 19a 0.017

290 E. D'Angelo et al. / Science of the Total Environment 438 (2012) 286–292

about 7.6 mg As kg−1. Thus, several litter amendments might berequired to detect significant increases in total As in these soils.

In this study, total As increased with soil depth, and was stronglyrelated to total Fe, Al, Mn, Ca, and Mg in the soils (R2 between 0.42and 0.64, p-valueb0.0001), which suggested that As was associatedwith oxyhydroxides and carbonates of these cations in soil parentmaterial. Compared to the Ecological Soil Screening Level for totalAs (US EPA, 2005), total As concentrations in both the litter-amended and non-amended soils were below than those expectedto deleteriously affect plants (18 mg As kg−1) or other organisms.Additional work is probably warranted to establish more accuratepredictors of As bioavailability in soils than total As.

Unlike total As, Mehlich 3 As tended to be highest in the soilsurface and decreased with depth in the litter-amended soils, whichsuggested that litter amendments increased As bioavailability inthese soils (Fig. 2). Several constituents in broiler litter, in additionto As, could explain why As bioavailability was higher in litter-amended soils (Jackson et al., 2006; Rutherford et al., 2003; Wangand Mulligan, 2006). Elevated organic matter and phosphate inlitter-amended soils (Figs. 1 and 2) can increase As solubility by com-peting for common sorption sites (Bauer and Blodau, 2006; Peryea,1991; Wang and Mulligan, 2006). Organic matter can also supporthigh rates of microbial respiration, development of anaerobic condi-tions, biological reduction of iron oxyhydroxides, and subsequentrelease of Fe-bound As (Weber et al., 2010). High and low molecularweight organic acids in litter can complex Fe and Al and sorb ontomineral surfaces, thereby reducing As sorption by these soil constitu-ents (Cornu et al., 2003; Tessema and Kosmus, 2001). It is likely that acombination of these factors contributed to higher As bioavailabilityin the litter-amended soils in this study.

Increased amounts of As in the bioavailable pool in the surface oflitter-amended soils were supported by results from the corn leaf tis-sue analysis, which contained significantly greater amounts of Asthan those leaves of plants grown in non-amended soils in both2007 and 2008 (Table 2). The role of tillage on As levels in leaf tissue,however, was not consistent in the two years. Levels of As in the corngrain ranged between 5 and 6 μg As kg−1 in both years, and was notsignificantly different in the litter-amended and non-amended soils,or tilled and non-tilled soils (Table 2). These results are consistentwith those of Liebhardt (1976), who did not find significant differ-ences in As levels in the grain of corn grown in soils amended with lit-ter from broilers that were fed roxarsone. At the As concentrations ofgrain measured in this study, it is unlikely that consumption ofreasonable amounts would pose health risks, assuming a tolerabledaily intake level of 2 μg As kg−1 body weight d−1 (World HealthOrganization, 2010).

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Several biological and chemical processes could explain why theleaves and grain of corn, which has a high capacity to take up As(Parsons et al., 2008), had low levels of As in spite of elevated bio-available As in the soils. Arsenic translocation from roots to aboveground plant tissues can be restricted because of As sequestrationby Fe oxide plaques and thiol ligands on corn roots (Parsons et al.,2008). Arsenic accumulation in above ground corn tissues can alsobe restricted when roots are associated with arbuscular mycorrhizalfungi (Yu et al., 2009). On the other hand, arsenic mobility andbioavailability and uptake by corn plants can be increased by earth-worm (Eissennia foetida) activity (Bai et al., 2007; Covey et al.,2010). Due to the complexity of these interactions, further investiga-tions of As cycling processes in the rhizosphere of living and decayingplants grown in litter-amended soils seem warranted.

It was expected that As in litter and soils would be leached byrainfall and accumulate in groundwater in the litter-amended plots.However, As concentrations in groundwater were consistentlybelow the US EPA drinking water maximum contaminant level of0.01 mg As L−1 (US EPA, 2006), and remained below the methoddetection limit of 0.005 mg As L−1 during the course of the study(data not shown). These results signified that As in the native soilsand litter was very strongly retained by the soils.

To investigate the importance of arsenate sorption, batchisotherms were conducted using soil samples from the 0–5 and80–90 cm depths. In the arsenate concentration range of 0 to5 mg As L−1, sorption followed a linear model with constants rang-ing between 39 and 202 L kg−1 (Fig. 3). Arsenate was retained signif-icantly more by the subsurface soils (average sorption constant89 L kg−1) than surface soils (46 L kg−1), which was likely attribut-ed to differences in Fe and Al oxyhydroxides in the different depths,which are known to strongly retain arsenate through ligand exchangereactions (Fendorf et al., 1997; Ona-Nguema et al., 2005; Waychunaset al., 1993). The importance of crystalline Fe and Al interactions witharsenate was supported by strong relationships between arsenatesorption constants and amounts of total Fe and Al in the soils(Fig. 3), which were not observed with other forms of Fe and Al(oxalate or Mehlich 3 extractable), pH, organic matter, or any otherproperty measured in this study. Strong affinities of crystalline Feand Al for arsenate in these soils likely accounted for the low levelsof As in plant tissue and groundwater observed in this study.

y = 0.0032x - 40

R2 = 0.91

0

50

100

150

200

250

0 20000 40000 60000 80000

y = 0.0048x - 62

R2 = 0.68

0

50

100

150

200

250

0 10000 20000 30000 40000 50000

Line

ar s

orpt

ion

cons

tant

, L k

g-1

Total Fe, mg kg-1

Total Al, mg kg-1

0-5 cm

80-90 cm

p= 0.0002

p= 0.012

A

B

Fig. 3. Relationships between arsenate sorption constants and A. total Fe and B. total Alin soils from the 0–5 cm and 80–90 cm depth increments.

4. Conclusions

The concentration of As in litter varied widely (b1 to>40 mg As kg−1) depending on which the broiler company farmthat it originated from. Considerable amounts of total As in litterwas soluble in phosphate buffer (15–20%) and consisted mostly ofarsenate and several other roxarsone transformation intermediatesthat were produced during the time that litter accumulated in thebroiler houses. When litter containing As was amended to agricul-tural soils, very little accumulated in corn leaves, grain, or ground-water. This was likely attributed to rapid retention of arsenate tothe Fe and Al oxides in the soils, as evidenced by the high sorptionconstants and strong relationships between sorption constants andtotal Fe and Al in the soils. Because of this, it appears that the envi-ronmental and health risks associated with amending litter tothese agricultural soils would be low; this may not be the case, how-ever, in other situations where freshly-produced litter that containshigher concentrations of water soluble As species are applied to soilswith low Fe and Al contents and shallow water tables.

Acknowledgments

We would like to acknowledge the assistance of Colleen Steele ofthe University of Kentucky, College of Agriculture, who collectedwater samples for arsenic analysis from lysimeters, Tricia Coakley ofthe University of Kentucky, Environmental Research and TeachingLaboratory, who analyzed arsenic in water, plant and litter samples,and also financial support for the project from the Kentucky SenateBill 271 Water Quality Grant.

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