Food and Chemical Toxicology 48 (2010) 3051–3057

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Food and Chemical Toxicology

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Evaluating the accumulation of arsenic in maize (Zea mays L.) plants from itsgrowing media by cloud point extraction

Jameel Ahmad Baig a,b, Tasneem Gul Kazi a,⇑, Abdul Qadir Shah a, Mohammad Balal Arain a,Hassan Imran Afridi a, Sumaira Khan a, Ghulam Abbas Kandhro a, Naeemullah a, Abdul Sattar Soomro a

a Centre of Excellence in Analytical Chemistry, University of Sindh, Jamshoro 76080, Pakistanb Young Welfare Society, Mughalabad, Taluka Thari Mirwah District, Khairpur Mir’s, Pakistan

a r t i c l e i n f o

Article history:Received 11 May 2010Accepted 28 July 2010

Keywords:ArsenicCloud point extractionMaizeElectrothermal atomic absorption

spectrometry

0278-6915/$ - see front matter � 2010 Elsevier Ltd. Adoi:10.1016/j.fct.2010.07.043

⇑ Corresponding author. Tel.: +92 022 2771379; faxE-mail addresses: jab_mughal@yahoo.com (J.A

(T.G. Kazi), aqshah07@yahoo.com (A.Q. Shah), bilalku2hassanimranafridi@yahoo.com (H.I. Afridi), skhangakandhro@yahoo.com (G.A. Kandhro), naeemullah43sattarsoomro2009@yahoo.com (A.S. Soomro).

a b s t r a c t

A cloud point extraction has been developed for the determination of As in maize (Zea mays L.) andadjoining agriculture soil. The different parts of maize (grains, shoots and roots) and soil were subjectedto microwave assisted digestion in a mixture of nitric acid and hydrogen peroxide (2:1 ratio). The traceamounts of As was pre-concentrated by cloud point extraction, as prior step to its determination by elec-trothermal atomic absorption spectrometry. The main factors affecting cloud point extraction efficiency,such as pH of sample solution, concentration of ammonium pyrrolidine dithiocarbamate and Triton X-114, equilibration temperature and time period for shaking were investigated in detail. For validationof proposed method a certified reference material of whole meal flour BCR 189 was used. No significantdifference p > 0.05 was observed between the experimental results and the certified values of CRM(paired t-test). The translocation of total As in different parts of maize were found in the order of,Root > Shoot > Grain.

� 2010 Elsevier Ltd. All rights reserved.

1. Introduction

Arsenic (As) was ranked as the No. 1 of hazardous element andhas serious effects on plants, animals and human health (Baig et al.,2009a,b,c; International Programme on Chemical Safety (IPCS),2001; Agency for Toxic Substances and Disease Registry (ASTDR),2005). The elevated As levels in agricultural soil is due to irrationalapplication of commercial fertilizers, manures and sewage sludge(Shemirani et al., 2005; Jamali et al., 2008; Arain et al., 2009).The As can be taken up by plants and vegetables through agricul-tural soil and irrigated water than subsequently enters into foodchains (Meharg and Hartley-Whitaker, 2002). It is reported thatrice straw grown in soil, irrigated by As contaminated groundwater have As uptake up to 92 mg kg�1 (Abedin et al., 2002).McLaughlin et al. (2000) had studied that plant available/bioavail-able toxic metals concentration in soils is important than total con-centrations of toxic metals (McLaughlin et al., 2000; Rodriguezet al., 2003). Therefore, determination of total As and plant avail-able As (aqueous extractable) in terrestrial plants and adjoining

ll rights reserved.

: +92 022 2771560.. Baig), tgkazi@yahoo.com004@yahoo.com (M.B. Arain),zai@gmail.com (S. Khan),3@yahoo.com (Naeemullah),

soil, helps us to understand the distribution, and transfer to thefood chain and organisms (Requejo and Tena, 2006).

For quantitative determination of As at trace levels in environ-mental and biological samples were conducted by several spectros-copy and hyphenated techniques (Zhang et al., 2007; Dias et al.,2002; Tuzen et al., 2009a; Boutakhrit et al., 2005; Cava-Montesinoset al., 2003; Morita et al., 2007). However, spectroscopy techniqueshave low limit of detection, whereas, hyphenated techniques wereexpensive, required high running cast and using toxic reagent.Therefore, pre-concentration method is most preferable for preciseand accurate determination of As (Shemirani et al., 2005; Zhanget al., 2004). There are several pre-concentration methods i.e., solidphase extraction, cloud point extraction, co-precipitation and ionexchange, chromatography were reported (Elçi et al., 2008; Ghaediet al., 2008; Khan et al., 2009; Baig et al., 2009c; Baig et al., 2010a,b;Ghaedi et al., 2009a,b; Kazi et al., 2010; Uluozlu et al., 2010; Tuzenet al., 2010). Cloud point extraction (CPE) is an outstanding alterna-tive to conventional solvent extraction method because it produceshigh extraction efficiencies and pre-concentration factors, by usinginexpensive and non-toxic reagents (Tuzen et al., 2009b; Daveigaet al., 2003; Pedro et al., 2008; Shemirani et al., 2005; Manzooriand Bavili-Tabrizi, 2002).

In the present work, we proposed a CPE method for the deter-mination of As in maize plant and its growing media by electro-thermal atomic absorption spectroscopy (ETAAS) with a modifierPd(NO3)2. For maximum extraction of As different experimental

3052 J.A. Baig et al. / Food and Chemical Toxicology 48 (2010) 3051–3057

variables i.e., complexing reagent (APDC), non-ionic surfactant(Triton X-114), pH, equilibrium time and temperature were stud-ied in detail. The accuracy of proposed CPE method was checkedby simultaneously analyzing the certified reference material ofwhole meal flour BCR 189. The proposed method was applied ondifferent tissues of maize plant and adjoining soil of two sub-dis-tricts of Khairpur Mir’s (Sindh, Pakistan).

2. Experimental work

2.1. Sampling

Soil samples were collected from agricultural sites, situated inKhairpur and Kot Diji sub-district of Khairpur Mir’s Pakistan(Southern areas of Pakistan) and classified as soil irrigated withtube well water (SIT) and soil irrigated with canal water (SIC).The samples were collected from the surface layer of sub soil (0–25 cm) with an auger made of stainless steel. The understudieddistrict lies in between Latitude 26� 00–27� 450 and Longitude 68�00–70� 150 (Baig et al., 2009). On returning to the laboratory, thesoil samples were spread on the plastic trays in fume cupboards,air dried for eight days at room temperature.

The grain crops (maize) grown on under study agriculturalsoil, irrigated with canal water and tube well water were col-lected simultaneously. The plants (each sample consisted ofthree sub-samples having a distance of approximately 300 cmbetween each other) were approximately 50–55 days old. Allsamples were collected from agricultural fields by hands pro-tected with vinyl gloves, were kept in separate plastic bags,stored in a cold box at 4 �C and transferred to laboratory onthe same day. Different parts of each plant (root, shoot andgrain) were separated with a quartz knife. At first, the crop tis-sues were put through a three step washing sequence, which in-volved agitating and rinsing first with distilled water followed bythree separate washes in ultrapure water. The soil samples werespread on plastic trays in fume cupboards and allowed to dry atambient temperature for 8 days. The each part of maize plantsand soil samples were air-dried, then placed in an electric ovenat 70 �C for 48–72 h depending on the sample size. The driedsamples were homogenized by grinding in an Agate mortarand sieved through a nylon sieve (<75 lm mesh size). The finalsamples were kept in labeled polypropylene containers at ambi-ent temperature before analysis.

Table 1Physicochemical property of the agriculture soils of two sub-districts of KhairpurMir’s irrigated by tube well and canal water.

Parameters TW soil CW soil

pH 7.62 ± 0.35 7.12 ± 0.39Organic matter (%) 71.0 ± 3.50 63.0 ± 3.20Organic carbon (%) 26.5 ± 1.80 24.8 ± 1.75Silica (%) 14.5 ± 1.28 12.9 ± 1.65CEC (mequiv./100 g) 15.3 ± 1.60 13.5 ± 1.70Total nitrogen (mg/kg) 2950 ± 39.2 2769 ± 25.3

2.2. Reagents and materials

The ultrapure water obtained form ELGA lab water system (Buc-hs, UK) was used throughout the work. A 0.1% (w/v) of Ammoniumpyrrolidine dithiocarbamate (APDC), Fluka Kamica (Bushs, Switzer-land) solution was prepared by dissolving in ultrapure water. Cal-ibrations were prepared for each analytical session using certifiedstock standard solution of As 1000 lg L�1, (Fluka). Working stan-dard solutions were prepared by stepwise dilution of stock stan-dard solutions prior to use. The chemical modifiers Mg(NO3)2

stock standard solution, 2000 mg L�1, prepared from Mg(NO3)2

Merck (Darmstadt, Germany), and Pd stock standard solution,3000 mg L�1, was prepared from Pd 99.999% Aldrich (Milwaukee,WI, USA). The standard reference material of human hair BCR397 (Brussels, Belgium) was used. The pH of the sample solutionwas adjusted with 0.1 M HCl (Merck). Nitric acid (HNO3) �16 Mand 30% hydrogen peroxide (H2O2) were obtained from (Merck).The certified reference materials, whole meal flour BCR 483was purchased from the bureau of references of Europeancommunities.

2.3. Apparatus

A thermostatic water bath (Gallankamp, Germany) was used tostudy temperature effects on cloud point extraction. The phaseseparation was assisted with a centrifuge ROWKA Laboratoryjnatype WE-1, nr-6933 (Mechanika Phecyzyjna, Poland). A PELdomestic microwave oven (Osaka, Japan), programmable for timeand microwave power from 100 to 900 W, was used for total diges-tion of samples. A pH meter (Ecoscan Ion 6, Kuala Lumpur, Malay-sia) was employed for pH adjustments. A programmable ultrasonicwater bath model No. SC-121TH (Sonicor, Deep Park, NY, USA) wasused for incubation with temperature ranging from 0 to 80 �C atintensification frequency of 35 kHz. The As in micelle mediatedphase was determined by a double beam Perkin Elmer model AAnalyst 700 (Norwalk, CT, USA) atomic absorption spectrophotom-eter, equipped with a graphite furnace HGA-400, autosampler AS-800 and deuterium lamp for background correction. Single elementhollow cathode lamp of As operated at 7.5 mA was used as energysource. The As signal was isolated at 193.7 nm with a spectralbandwidth of 0.7 nm and atomization was achieved in a pyrocoat-ed graphite tube with integrated platform. The WinLab 32 softwarewas used to acquire and process analytical data. The graphite fur-nace heating program was set for As analysis as: drying, ashing,atomization and cleaning at temperature range �C/(ramp time inseconds/holding time in seconds) (80–120/1/30, 300–600/10/20,2200–2400/0/5 and 2400–2800/0/2), respectively. For all determi-nations, argon (200 mL min�1) was used as a purge gas except dur-ing the atomization step. The 10 lL of (standard or sample volume)and 10 lL modifier (5 lg Mg (NO3)2 + 3 lg Pd) were injected toelectrothermal graphite atomizer. The peak height was used forquantitation of As (Baig et al., 2009,b,c).

2.4. Physico-chemical studies of soil

The physicochemical parameters of each batch of SIT and SICsamples i.e., pH, organic matter, organic carbon, silica, cation ex-change capacity and total nitrogen were determined according tothe official methods of soil analysis. Detail was reported in our pre-vious work (Jamali et al., 2008; Baig et al., 2009). The soil and ultra-pure water (1:2.5) suspension was prepared and kept overnight atroom temperature, for pH measurement (Buurman et al., 1996).Organic matter content was obtained by ashing duplicate samplesof each batch of soil in muffle furnace at 540 �C for 6 h. The differ-ence between the dry weight of soil samples before and after ash-ing was used for the calculation of organic matter content (Soltner,1988). The organic carbon was measured by the Walkeley–Blackmethod (Jamali et al., 2007). It was oxidised to carbon dioxide withpotassium dichromate in the presence of sulphuric acid. The unre-acted potassium dichromate was titrated with iron (II) sulphate.Total nitrogen level was measured by Kjeldahl method (Jamaliet al., 2006). All analyses were performed in triplicate. Blanks wererun simultaneously. Selected parameters of each soil are presentedin Table 1.

J.A. Baig et al. / Food and Chemical Toxicology 48 (2010) 3051–3057 3053

2.5. Sample preparation

2.5.1. Aqueous extractable arsenic in soilTriplicate 2 g samples of air-dried SIT and SIC, kept in 50 ml

polyethylene bottles, were added with ultra pure water (1:10 sub-stratum:water) and kept in the mechanical shaker at 30 rpm for6 h. To separate the extract from the residue, the mixture was thencentrifuged for 10 min at 4000 rpm. Blanks were analysed in paral-lel of each batch, using the same extracting reagents.

2.5.2. Total arsenic in maize and soilA microwave assisted acid digestion procedure was carried out

to obtain total As in samples of maize plant and its growingmedia. The six replicates samples of CRM and triplicate of differ-ent tissues of maize and soil samples (0.2 g) were directlyweighed into PTFE flasks (50 mL in capacity). Two ml of a freshlyprepared mixture of concentrated HNO3 and H2O2 (2:1, v/v) wasadded to each flask and was kept for 10 min at room temperature.The flasks were sealed and subjected to microwave heatingprogram (Kazi et al., 2009). Following digestion, samples weretransferred to 20 ml volumetric flask and the volume was made-up with de-ionized water.

The aqueous extractable and digested solutions were further di-vided into two set, one set of digested solution was subjected toETAAS with modifier for TAs and Asaqueous determination, whileother set was subjected to cloud point extraction of TAs andAsaqueous,prior to subjecting ETAAS.

2.5.3. Enrichment procedureAliquots of 10 mL of standard solutions containing As in the

range of 10 lg L�1, replicate six samples of 10 mL of digestedCRM, and triplicate acid digested solutions and aqueous extractof soil samples, and acid digested tissues of maize were taken ingraduated centrifuge tubes (25 mL in capacity). Added 30% KIand 5% ascorbic acid or L-cysteine (50 g L�1) to reduce AsV to AsIII.Then added 0.1–1.2 mL (0.61 � 10�4–7.3 � 10�4 mol L�1) of APDCsolution (3.05 � 10�3 mol L�1), 2.0 mL of Triton X-114 solution(0.5–2%, v/v) and pH of solutions were adjusted (1–10) by adding0.1 mol L�1 of HCl/NaOH. The tubes were kept in an ultrasonic bathat 20–60 �C for 2–10 min. After different time intervals, separationof the two phases was achieved by centrifuging for 5 min at3500 rpm. After cooling in ice-bath, the surfactant-rich phasebecame viscous and the upper aqueous phase decanted. To de-crease the viscosity of extracts added acidic ethyl alcohol(0.1 mol L�1 HNO3) and introduced into ETAAS with modifier. Ablank submitted to the same procedure was measured parallel tothe calibration solutions of standards, whole meal flour BCR 483and real samples.

2.6. Statistical analysis

Data processing and statistical analysis were conducted byusing computer program EXCEL (XP 2002; Microsoft Corp., Red-mond, WA) and Minitab 13.2 Minitab Inc., State (College, PA) soft-ware packages. Normally distributed data were expressed asMeans ± SD, Student’s t-test and Mann–Whitney test were usedto assess the significance of the differences between the variablesinvestigated in proposed method.

Fig. 1. Effect of pH on the CPE of 10 lg L�1 As. Other CPE conditions:4.3 � 10�4 mol L�1 APDC, 0.12% concentration of Triton X-114, equilibrationtemperature 35 �C, equilibration time 10 min.

3. Results and discussion

For the optimization of CPE, five factors were selected to beexamined, pH, mass of complexing agent, amount of surfactant,equilibrium time and temperature.

3.1. Effect of pH

It is known that the pH of the sample solution on the aqueous-organic extraction process is very important because the complex-ation of metals with organic ligands mostly depends on its form ata particular pH (Stalikas, 2002; Scaccia and Frangini, 2004). In or-der to evaluate the effects of this important parameter, pH valuesof sample solutions were adjusted in the range of 1.0–10.0 withHCl or NaOH (Fig. 1). The complex formation was begin at pH 2.5and started to decrease at pH 6, showed a plateau at pH values3.5–5.5. The possible reasons are that the stable complex formedbetween AsIII and APDC at pH > 3 and <6 in organic aqueous phase(Tang et al., 2005). Hence, APDC is selective for the formation ofcomplex with reducing form of As (AsIII) at pH ranged 3–6. Asthe signal for As increases after pH 3.5 and becomes decreasedafter pH 6. Thus, pH 4.5 was chosen as an adequate pH value forfurther studies.

3.2. Effect of APDC concentration

For current study, APDC was used as the chelating agent due toits highly hydrophobic nature for metal/metalloid complexformation. The optimization of APDC concentration is importantparameter for maximum extraction efficiency of As by CPE inenvironmental and biological samples. To check the fluctuationduring complex formation at different concentration levels ofAPDC is given in Fig. 2, ranged from 0.61 � 10�4 to7.3 � 10�4 mol L�1. It was observed that the extraction efficiencyof CPE for As was enhanced rapidly as the concentration of APDCincreased from 3.0 � 10�4 to 6.1 � 10�4 mol L�1. No significantenhancement was observed on the analytical signal with furtherincrease in APDC concentration upto7.3 � 10�4 mol L�1. Therefore,for further experiments, 4.3 � 10�4 mol L�1 of APDC concentrationof was employed. The stoichiometry of the As-PDC complex wasobserved 1:3 ratios.

3.3. Effect of Triton X-114

There are several non-ionic surfactants (Triton X-114, and X-100 etc.) were used for CPE. But, Triton X-114 was chosen for thisstudy because of its higher extraction efficiency as well as its lowercloud point temperature, which facilitates the phase separation bycentrifugation as compared with other tested surfactants (Silvaet al., 2006; Shah et al., 2010). The Fig 3 shows the variation inextraction efficiency of As with APDC complex range of 0.01–0.25% was observed. The 60–70% recovery was observed at 0.05%of Triton X-114, while the extraction efficiency reached a maxi-mum in the concentration of 0.12%. So, a concentration of 0.12%

Fig. 2. Effect of concentration of APDC on the CPE of 10 lg L�1 As. Other CPEconditions: 0.12% (v/v) concentration of Triton X-114, pH 4.5, equilibrationtemperature 35 �C, equilibration time 10 min.

Fig. 3. Effect of concentration of Triton X-114 on the CPE of 10 lg L�1 As. Other CPEconditions: 4.3 � 10�4 mol L�1 APDC, pH 4.5, equilibration temperature 35 �C,equilibration time 10 min.

3054 J.A. Baig et al. / Food and Chemical Toxicology 48 (2010) 3051–3057

was chosen as the optimum surfactant concentration in order toachieve the highest possible extraction recovery of As for stan-dards, CRM samples, while <0.12% the extraction efficiency of com-plexes is low probably because of the inadequacy of the assembliesto entrap the hydrophobic complex quantitatively. At volume high-er than 0.12% (v/v), the signals decrease because of the incrementin the volumes and the viscosity of the surfactant phase. To de-crease the viscosity of extracts acidic ethyl alcohol 0.1 mol L�1

was added.

Table 2The results for tests of addition/recovery for Asaqueous and TAs determination in soil samp

Species Add

Asaqueous 0.02.55.010.

Total As 0.02.55.010.

Validation for total As (mg kg�1)Element Indicative value (whole meal flour BCR 189) Fou

Total As 0.018 ± 0.0005 0.0tcertical = 2.57 at 95% confidence limit, (n = 6)

a %Recovery ¼ Cafter spikedCinitialþCspiked

� 100.

3.4. Effects of equilibration temperature and time

It was desirable to employ the shortest equilibration time andthe lowest possible equilibration temperature, as a compromisebetween completion of extraction and efficient separation ofphases. It was found that 35 �C is adequate for these analyses.The dependence of extraction efficiency upon equilibration timewas studied for a time span of 5–20 min. An equilibration time of10 min was chosen for the quantitative extraction.

3.5. Interferences

The effects of the matrix ions were investigated for efficientextraction of As by CPE. About 1 lg of As model solutions(100 mL) containing matrix ions were used in this study. The re-sults showed that Se4+, Pb2+, Ni2+, Co2+, Mn2+ and Fe2+ (up to theconcentration level of 100 mg L�1), Na+ (up to 1000 mg L�1), Mg2+

and K+ (up to 500 mg L�1) did not cause any significant interfer-ence on the CPE of As. Therefore, the proposed method has beenshown good selectivity.

3.6. Analytical performance

Calibration graphs were obtained by pre-concentrations of10 mL of samples in the presence of 1% Triton X-114 at pH 4.2.The linear range of the calibration curve for As reached from thedetection limit up to 0.2 and 20 lg L�1. The regression equationwas: y = (0.238) (As) + 0.142, R2 = 0.999, where y is integratedabsorbance and the concentration of As in SH samples is expressedas lg g�1. The limits of detection (LOD) and limits of quantification(LOQ) for As was calculated as under, LOD = 3 � s/m and LOD =10 � s/m respectively, where s is the standard deviation of tenmeasurements of the blank and m is the slope of the calibrationgraph. The LOD and LOQ of As was 0.025 and 0.083 g g g�1, respec-tively. The enhancement factor of about 50 was obtained by pre-concentration a 10 ml of sample solutions. The results indicatedthat the method has good precision. The method was assured bythe analysis of triplicate samples, reagent blank, procedural blanksand standard reference material. In order to validate the methodfor accuracy and precision, a certified reference material of wholemeal flour BCR 483 was analyzed with As content of 0.018 ±0.0005 lg g�1 (indicative value) (Jamali et al., 2008). The % recov-ery of As with CPE was observed 98.2 ± 0.3% (Table 1). The preci-sion of the methods, expressed as the relative standard deviation(RSD) of 6 independent analyses of the same CRM sample withCPE was 1.70%. The paired t-test was calculated for (n�1 = 5) de-grees of freedom, the value of tExperiment (0.12) was less than thetcertical (2.57) at 95% confidence interval (Table 2), indicating no

les by CPE (n = 6).

ed Conc. (lg L�1) Mean ± SD (lg g�1) a% Recovery

0 0.85 ± 0.02 –3.26 ± 0.05 97.35.75 ± 0.07 98.3

0 10.69 ± 0.18 98.5

0 2.90 ± 0.03 –17.3 ± 0.18 97.719.8 ± 0.21 98.0

0 24.9 ± 0.32 98.8

nd values x� ts=ffiffiffi

np

% recovery (% RSD) tExperiment

177 ± 0.0003 98.2 (1.70) 0.27

Table 3Comparative data of analytical characteristics of the CPE method for As (lg/L).

Complexing reagent/surfactant Technique ar LOD/LOQ Precision EFb Reference

APDC/Triton X-114 CPE/ETAAS 0.998 0.04/0.13 3.0 (n = 11) 36 Tang et al. (2005)APDC/Triton X-114 CPE/ETAAS 0.999 0.03/0.11 <2.3% (n = 6) 40 Baig et al.(2009)O,O-Diethyldithiophosphate/Triton X-114 Ultrasonic nebulization inductively

coupled plasma mass spectrometry0.993 0.006/– 3.8% (n = 8) 42 da Silva et al. (2000)

APDC-Activated Carbon GFAAS – 0.05/– 4.1% (n = 9) 50 Jiang et al. (2008)Molybdate/sulfuric acid CPE/ETAAS 0.999 0.011/0.037 5.00% (n = 5) 52 Shemirani et al. (2005)APDC/Triton X-114 CPE/ETAAS 0.999 0.025/0.083 1.70% (n = 6) 50 This work

a Correlation coefficient.b Enrichment factor.

J.A. Baig et al. / Food and Chemical Toxicology 48 (2010) 3051–3057 3055

difference between found values and indicative value. Due to thelack of reference material for As speciation, the validity of analyti-cal method was performed on replicate three sub-samples of SIC,spiking with three concentration levels of Asaqueous and TAs thenapplied both methods. The recoveries for Asaqueous and TAs weregenerally greater than 98% (Table 2). A good agreement was ob-tained between the added and measured analytes concentration.

The CPE of different forms of As using different organic ligandsare shown in Table 3. The comparative data of analytical character-istics shows that the characteristic parameters of CPE viz. correla-tion coefficient (r), LOD/LOQ, precision and enrichment factor ofpresent study was comparatively better than previously reportedworks (da Silva et al., 2000; Tang et al., 2005; Baig et al., 2009).While the CPE characteristic parameters of our study were con-sisted with those reported by Shemirani et al. (2005) and Jianget al., 2008.

3.7. Application

The accumulation of toxic metals in food crops has been recog-nized as an issue of high priority by many governmental agenciesaround the world (Organization for Economic Co-operation andDevelopment (OECD), 2003; United States EnvironmentalProtection Agency (USEPA), 1996). The methods related withphytotoxicity should be enhanced in assessing the impacts ofchemicals on terrestrial ecosystem. The food crop serves as animportant pathway for human exposure to toxic elements includ-ing As (Cheng et al., 2006). Thus current study has documented theconcentration of TAs and Asaqueous in soil samples and TAs concen-tration in different parts of maize plants (grain, shoot and root).The developed CPE method was successfully applied for the analy-sis of As contents in understudied agricultural soils and differentparts of maize. The results shown in Table 4 indicated that a highTAs concentration in SIT of two sub-districts Khairpur and Kot Dijiwas found in the range of 25.0–36.3 and 18.7–54.0 lg g�1, respec-tively. The TAs level in SIC of both sub-districts were obtained inthe range of 20.5–31.0 and 15.2–48.2 lg g�1, respectively. The

Table 4Total As (TAs) and water extractable As (Asaqueous) concentrations in soil (lg g�1) byCPE.

Khairpur Mir’s Kot Diji

aTWIS bCWIS aTWIS bCWIS

TAs Range 25.0–36.3 20.5–31.0 15.2–48.2 18.7–54.0Mean 29.6 25.0 30.5 32.7Median 12.5 5.90 15.8 9.30

Asaqueous

(Din-test)Range 0.85–1.95 0.90–1.40 0.71–2.88 0.71–2.15Mean 1.52 1.13 1.50 1.24Median 1.48 1.05 1.42 1.21

a Tube well water irrigated soil.b Canal water irrigated soil.

Asaqueous in SIT samples of both sub-districts were ranged 0.85–1.95 and 0.71–2.88 lg g�1, respectively and the level of Asaqueous

in SIC samples of both district was 0.90–1.40 and 0.71–2.15 lg g�1,respectively. The concentration of TAs and Asaqueous in SIT were sig-nificantly higher than those observed in SIC (P < 0.001). It is be-cause of high As concentration in ground water (>50 lg L�1), ascompares to surface water (<10 lg L�1) (Baig et al., 2010). The highlevel of As in ground water (tube well water used for irrigation pur-poses) is due to dissolution of As compounds coming from Hima-laya through the Indus river and settled down over the years andthen introduced into groundwater by geothermal, geo-hydrologi-cal and bio-geo-chemical factors (Baig et al., 2010; Smedleyet al., 2002; Singh, 2006). Moreover, it might be due to the As con-taining insecticides and herbicides used for agriculture purposesand seepages from hazardous waste sites (Smedley and Kinni-burgh, 2002).

The TAs contents in different parts of maize plant i.e., grain,shoot and root grown in soils (SIT and SIC) understudied sub-dis-tricts shown in Table 5. The mean TAs concentrations in root, shootand grain of maize (n = 76), grown in SIT of Khairpur Mir’s were2.05 ± 0.66, 0.406 ± 0.09 and 0.302 ± 0.05 lg g�1, respectively andin same tissues of maize (n = 83) grown in SIC were found to be0.50 ± 0.0.9 0.202 ± 0.08 and 0.151 ± 0.06, respectively. These re-sults are consistent with the study conducted in Bangladesh byDas et al. (2004) found 2.4 lg g�1 As in rice roots, 0.73 lg g�1 inshoots and 0.14 lg g�1 in grain. Whereas, the lower mean TAs con-tents in root, shoots and grains of maize crop were observed1.74 ± 0.53, 0.365 ± 0.1 and 0.280 ± 0.04 lg g�1 and 0.542 ± 0.08,0.27 ± 0.10 and 0.171 ± 0.07 lg g�1, respectively grown in SIT andSIC of Kot Diji. The TAs in different parts of maize crops grown inSIT and SIC is shown in Table 5. The TAs uptake by maize plantsfrom both studied soil of two sub-districts were observed in theincreasing order: grain < shoot < root. This is according to reportedstudy as the roots accumulate more toxicants, and is more sensi-tive, than shoot or grain (United States Environmental ProtectionAgency (USEPA), 1996; An, 2004). The concentration of As in stemsof plants were considerably lower as compared to the root system,which proved that As movement along the plants conductive sys-tem was strongly limited.

Table 5Concentration of total As in different part of maize with CPE (lg g�1) andcontamination factor (CF).

Area Parts Maize samples grownin TWIS

Maize samples grown inCWIS

Mean ± SD CF Mean ± SD CF

Khairpur Mir’s Grain 0.302 ± 0.05 0.10 0.151 ± 0.06 0.03Shoot 0.406 ± 0.09 0.202 ± 0.08Root 2.05 ± 0.66 0.50 ± 0.0.9

Kot Diji Grain 0.280 ± 0.04 0.07 0.171 ± 0.07 0.034Shoot 0.365 ± 0.1 0.27 ± 0.10Root 1.74 ± 0.53 0.542 ± 0.08

3056 J.A. Baig et al. / Food and Chemical Toxicology 48 (2010) 3051–3057

The ratio between plant and soil concentrations of elements(contamination factor ‘‘CF”) is an index of soil–plant transfer thatfavors the understanding of plant uptake characteristics (Chamber-lain, 1981) and it is widely used in biomonitoring studies (Mingo-rance et al., 2005). Ratios > 1 indicate that plants are enriched withelements (accumulator), ratios around 1 indicates that plants arenot influenced by elements (indicator) and ratios < 1 shows thatplants exclude the elements from uptake (Baker, 1981). Resultsof CF (Table 5) display that both species exhibited the same behav-ior. The CF values of this study were <1 for both SIT and SIC ofunderstudied sub-districts, indicating a low translocation from soilto plant shoots in all sampling sites. These findings indicate thatdifferences in soil accumulation rates in SIT can occur not only lat-erally but also between years. They point to the need for morestudy sites covering a wider range of environmental conditionsand for relevant parameters to be monitored over a period of years.

4. Conclusions

This study provided a safe alternative method based on CPEmethod for the pre-concentration of As in maize crop and adjoin-ing soil samples and determination by ETAAS. The proposed meth-od has the following advantages; is a simple, rapid, sensitive,inexpensive, non-polluting technique with high enhancement fac-tor. The complexing agent, APDC, has enough hydrophobicity to beused in the proposed procedure, being quite selective and stable athigh acid concentrations, which is very convenient, since the watersamples are usually preserved with acids. The maximum extrac-tion efficiency was achieved at optimum levels of 4.5 pH,4.3 � 10�4 mol L�1 of APDC concentration, 0.12% of Triton X-114amount, 35 �C of equilibration temperature and 10 min equilibra-tion temperature, for As in soil and grain crop samples. The exper-imental results showed that the CPE was a successful method fordetermination of As in maize and adjoining soils irrigated by tubewell and canal water in two sub-districts of Pakistan with satisfac-tory recoveries. These findings urged more work on As controlledand exposed grain crops and vegetables in detail and should takeinto consideration variations in uptake between different species,cropping history, the levels of metals present in the atmosphere(quantified) and the difference in maize uptake between soilsand foliar mechanisms.

Conflict of interest

None.

Acknowledgements

This work was supported by Higher Education Commission(HEC), Islamabad (Grant No. 20-1246/R&D/9).

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