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LEGUME-GRASS INTERCROPPINGPHYTOREMEDIATION OF PHTHALIC ACIDESTERS IN SOIL NEAR AN ELECTRONICWASTE RECYCLING SITE: A FIELD STUDYTing Ting Ma a b , Ying Teng a b , Yong Ming Luo a b & Peter Christie ca Key Laboratory of Soil Environment and Pollution Remediation ofthe Chinese Academy of Sciences, Institute of Soil Science, Nanjing,Chinab Graduate University of the Chinese Academy of Sciences, Beijing,Chinac Agri-Environment Branch, Agri-Food and Biosciences Institute,Newforge Lane, Belfast, United KingdomAccepted author version posted online: 29 May 2012.Publishedonline: 11 Sep 2012.

To cite this article: Ting Ting Ma , Ying Teng , Yong Ming Luo & Peter Christie (2013): LEGUME-GRASSINTERCROPPING PHYTOREMEDIATION OF PHTHALIC ACID ESTERS IN SOIL NEAR AN ELECTRONIC WASTERECYCLING SITE: A FIELD STUDY, International Journal of Phytoremediation, 15:2, 154-167

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International Journal of Phytoremediation, 15:154–167, 2013Copyright C© Taylor & Francis Group, LLCISSN: 1522-6514 print / 1549-7879 onlineDOI: 10.1080/15226514.2012.687016

LEGUME-GRASS INTERCROPPING PHYTOREMEDIATIONOF PHTHALIC ACID ESTERS IN SOIL NEAR ANELECTRONIC WASTE RECYCLING SITE: A FIELD STUDY

Ting Ting Ma,1,2 Ying Teng,1,2 Yong Ming Luo,1,2

and Peter Christie31Key Laboratory of Soil Environment and Pollution Remediation of the ChineseAcademy of Sciences, Institute of Soil Science, Nanjing, China2Graduate University of the Chinese Academy of Sciences, Beijing, China3Agri-Environment Branch, Agri-Food and Biosciences Institute, Newforge Lane,Belfast, United Kingdom

A field experiment was conducted to study the phytoremediation of phthalic acid esters(PAEs) by legume (alfalfa, Medicago sativa L.)-grass (perennial ryegrass, Lolium perenneL. and tall fescue, Festuca arundinacea) intercropping in contaminated agricultural soilat one of the largest e-waste recycling sites in China. Two compounds, DEHP and DnBP,were present in the soil and in the shoots of the test plants at much higher concentrationsthan the other target PAEs studied. Over 80% of ‘total’ (i.e., all six) PAEs were removedfrom the soil across all treatments by the end of the experiment. Alfalfa in monocultureremoved over 90% of PAEs and alfalfa in the intercrop of the three plant species containedthe highest shoot concentration of total PAEs of about 4.7 mg kg−1 DW (dry weight).Calculation of phytoextraction efficiency indicated that the most effective plant combinationsin eliminating soil PAEs were the three-species intercrop (1.78%) and the alfalfa monocrop(1.41%). Phytoremediation with alfalfa was effective in both monoculture and intercropping.High bioconcentration factors (BCFs) indicated the occurrence of significant extraction ofPAEs by plants from soil, suggesting that phytoremediation may have potential for theremoval of PAEs from contaminated soils.

KEY WORDS: phthalic acid esters, phytoremediation, intercropping, monoculture, biocon-centration factor

INTRODUCTION

Phthalic acid ester (PAE) compounds are the most extensively used plasticizersworldwide and have become ubiquitous pollutants with widespread distribution in the en-vironment (Zeng et al. 2008). PAEs can severely retard crop growth, lower the quality ofcrop products, and impact soil animals and microorganisms, and their toxicity in ecosys-tems can include adverse effects on reproductive development and other genetic effects in

Address correspondence to Yong Ming Luo, Key Laboratory of Soil Environment and Pollution Remediation,Institute of Soil Science, Chinese Academy of Sciences, P.O. Box 821, 71 East Beijing Road, Nanjing 210008,Jiangsu Province, China. E-mail: ymluo@issas.ac.cn

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mammals (Rhee 2002; Shono and Suita 2003; Takeuchi et al. 2005; Kondo et al. 2006; Fuet al. 2007; Kusu et al. 2008; Saito et al. 2010; Suzuki et al. 2010). PAEs interact withandrogen and thyroid receptors more than estrogen receptors (Shigeki et al. 2001). Theirpossible accumulation in edible vegetables from soils and potential harm to humans viathe food chain has also aroused public concern (Zeng et al. 2008). The remediation of PAEcompounds in contaminated soils is therefore an issue of active concern.

Uncontrolled e-waste recycling at Taizhou, Zhejiang province, one of the largeste-waste dismantling sites in China, has led to combined soil contamination by substancessuch as potentially toxic elements (including heavy metals), polychlorinated biphenyls(PCBs), polycyclic aromatic hydrocarbons (PAHs), polychlorinated dibenzo-p-dioxins/dibenzofurans (PCDD/Fs), and polybrominated diphenyl ethers (PBDEs), and this has ledto great public concern for the environment and human health (Wang et al. 2006; Chenet al. 2008; Li et al. 2008; Ni et al. 2008). PAEs can be found in the ash formed fromthe open burning of electric wires (Liu et al. 2009). PAEs tend to have high lipophicityand this accounts for their ready adsorption to organic matter in contaminated soil andsediment particles (Zheng et al. 2007; Xu and Li 2008). Furthermore, during the last twoyears the soil total concentrations of five PAE compounds were reported to range from12.6 to 46.7 mg kg−1, concentrations much higher than the allowable soil concentrationsin the United States (Liu et al. 2009; Liu et al. 2010; Zhang et al. 2010a,b). They havebeen nominated in the list of 129 prior pollutants in the United States and as environmentalendocrine disruptor chemicals by USEPA and we therefore selected them to be the targetpollutants in our investigation.

Potential advantages of phytoremediation compared with other remediation ap-proaches include the preservation of soil texture and structure, the utilization of solarenergy, the involvement of an active soil microbial biomass, and low cost (Schnoor et al.1995; Chekol et al. 2004; Huang et al. 2004). The use of higher plants for effective re-mediation has become a topic of increasing interest (Cunningham et al. 1996; McIntireand Lewis 1997). Environmentally friendly phytoremediation using plant species such asalfalfa, perennial ryegrass and tall fescue has been employed for PCBs, PAHs and othersoil pollutants (Chekol et al. 2004; Rezek et al. 2008; Cheema et al. 2009; Shen et al.2009; Cheema et al. 2010; Tang et al. 2010; Xu et al. 2010a) and in preliminary greenhousetests for PAEs in our laboratory. However, phytoremediation of phthalate compounds hasso far received little attention. The aims of the present work were therefore to compare theconcentrations and components of PAEs in an agricultural soil under different treatmentsin an experimental field area, to check the efficiency of phytoremediation to PAE targetpollutants and determine the concentration of total and individual PAE compounds in theshoots of different plant species, and to evaluate different plant species combinations fortheir suitability for larger scale remediation.

MATERIALS AND METHODS

Chemicals

A mixed standard solution of 6 PAE compounds (1 mg ml−1) was prepared, namelydimethyl phthalate (DMP), diethyl phthalate (DEP), butyl benzyl phthalate (BBP), di-n-butyl phthalate (DnBP), bis (2-Ethylhexyl) phthalate (DEHP), di-n-octyl phthalate (DnOP),and the internal standard benzyl benzoate (BB, 5 mg ml−1) which were all obtained fromAccuStandard, Inc., New Haven, CT, USA. The six target pollutants were selected for

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study because they were nominated as priority pollutants by USEPA. Certified referencematerial CRM 136–100 (Base Neutral Acids: BNAs – Clay 1) was purchased from RTCorp., Laramie, WY, one of the original Proficiency Test providers recognized by USEPA.

Analytical grade solvents (acetone and hexane) obtained from chemical reagentcompanies in Nanjing were re-distilled in an all-glass system to remove trace impuritiesbefore use. HPLC grade hexane was purchased from Tedia Company Inc., Fairfield, OH.Anhydrous sodium sulfate (NaSO4, reagent grade), neutral alumina (Al2O3, 400 mesh andreagent grade), neutral silica gel (100–200 mesh) and sulphuric acid (H2SO4, guaranteedreagent) were obtained from National Pharmaceutical Group Chemical Reagent Co., Ltd.,Shanghai, China. The three column packing materials were dried in a muffle furnace at400◦C for 6 h and stored in desiccators before use.

Plant Materials

Seeds of the three plant species alfalfa (Medicago sativa L.), perennial ryegrass(Lolium perenne L.) and tall fescue (Testuca arundinacea) were purchased from the Instituteof Agricultural Science and Technology, Jiangsu province, east China.

No plastic equipment was used during sample collection and preparation to mini-mize background contamination in PAE analysis. Glassware was washed by the followingprocedure prior to analysis. After immersion and washing with soapy water in a laboratoryultrasonic washer and air drying, glassware with tick marks was immersed in sulfuric acid(H2SO4, 98%, reagent grade) and washed with tap water, then rinsed with ultrapure waterbefore oven dry at 50◦C. Glassware without tick marks was baked for 6 h at 400◦C in amuffle furnace. All glassware was then thoroughly rinsed with acetone:hexane (1:1 v/v)before use.

Phytoremediation Experiment

The field experiment was located on agricultural land about 250 m from an electronicwaste dismantling site at Taizhou, Zhejiang province, east China for a period of two years.The average total concentrations of target PAEs were 1032.6 ± 311.8 µg kg−1 in October2008 and 2389.5 ± 370.4 µg kg−1 in October 2010 in the control treatments. The pH valueof the soil was 5.56, with an organic matter content of 36.5 g kg−1, and total nitrogen,phosphorus and potassium concentrations of 1.96, 0.56 and 23.1 g kg−1, respectively.

The planting regimes consisted of an unplanted control treatment (CK), alfalfa mono-culture (A), perennial ryegrass monoculture (P), tall fescue monoculture (T), alfalfa andperennial ryegrass intercrop (AP, seed ratio 1:1), alfalfa and tall fescue intercrop (AT, seedratio 1:1) and alfalfa, perennial ryegrass and tall fescue intercrop (APT, seed ratio 1:1:1).There were 4 replicate plots of each treatment, each 2.4 × 2.4 m, giving 28 plots in total ina fully randomized design. All three plant species were sown by broadcasting at the startof the experiment.

Sample Processing

Two years after the establishment of the experiment the soil and shoots of individualplant species in each plot were sampled. Soil samples were collected to 15 cm depth usinga soil corer. Five cores were collected from each plot and combined to give a compositesample. All of the plant shoots were collected by cutting just above ground level within each

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plot for biomass measurement and five random shoots were selected for further analysis.Fresh plant samples were taken to the laboratory, immediately washed with tap water andrinsed with distilled water and wiped dry with paper tissue before freeze-drying with thesoil samples in a Free Zone 2.5 Liter Freeze Dry System (Labconco Corp., Kansas City,MO, USA). Soil samples were air dried and sieved through a 60-mesh screen after dryingand plant samples were homogenized in liquid nitrogen prior to storage at –20◦C and furtheranalysis.

10.00 g of soil (or 1.00 g of plant sample) was placed in a clean glass centrifuge bottle,mixed on a vortex mixer for 1 min, and immersed in 30 ml of acetone:hexane (1:1 v/v)overnight. Spiked samples were prepared by adding 1 ml of 1 mg l−1 standard solution of6 target PAEs to the soil (or 0.1 ml to the plant sample) before analysis following the sameprocedure. Ultrasonic extraction was carried out the next day in a water bath at 25◦C for30 min at 100% power before centrifuging at 1500 rpm for 5 min. The three supernatantswere all filtered into a round bottom flask after another two extractions with 20 ml ofacetone: hexane (1:1 v/v) for 15 min each. About 70 ml of liquid in the flask was reducedby rotary evaporation to 1–2 ml (350 mbar, 40◦C water bath, 80 rpm). Hexane (3–4 ml)was added to the remaining solvent and rotary evaporation was continued to less than 1 mlbut not to dryness.

Column chromatography was performed in a glass column (1 × 26 cm) with 2 g ofNaSO4, 6 g of neutral Al2O3 and 12 g of neutral silica gel (from bottom to top). Pre-washingwas with 15 ml of hexane and 15 ml of acetone:hexane (1:4 v/v) mixture before sampleloading and elution with 40 ml of acetone:hexane (1:4 v/v). All the washing solutions werecollected and reduced to less than 1 ml by rotary evaporation as described above. Plantsamples were further washed with sulphuric acid (Meng et al. 1996) and concentrated toless than 1 ml. 10 µl of internal standard, BB, was added before hexane (HPLC grade) wasadded to bring the final volume to 1 ml. Samples were transferred to brown sample bottlesand stored at -20ºC before further analysis.

Instrumental Analysis and Quality Control

Analysis of individual PAEs in samples was performed by a modification of USEPAmethod 8270C (USEPA 1996) with an Agilent 7890GC-5975 MSD gas chromatograph-mass spectrometer. Samples were resolved on a DB-5 (30 m × 0.25 mm × 0.25 µm)fused-silica capillary column with helium (purity >99.999%) as carrier gas at 1.2 mlmin−1. The injector temperature was set at 250◦C. The GC temperature program featuredan initial column temperature of 50◦C which was held for 1 min, with a ramp of 15◦Cmin−1 to 200◦C which was held for 1 min, then 8◦C min−1 ramp to 280◦C which was heldfor 3 min. Post run was at 285◦C for 2 min. Under selected ion monitoring mode, non-pulseinjection with a volume of 1.0 µl each in split-less mode was carried out. The GC-MStransfer line was set at 280◦C.

During analysis, whole procedure blanks, soil matrix blanks, spiked soil matrixand parallel samples were all employed, together with analysis of the certified referencematerial. The recovery rates of spiked soils matrix at 100 µg kg−1 (DW) were between75.88% and 107.61%, with an RSD of 3.88–8.91. Instrument detection limits (IDLs) forthe 6 target compounds were 0.11–0.35 µg l−1, and method detection limits (MDLs) were68–135 µg kg−1. Linearity of response between 0.02 to 2 mg l−1 showed R2 (correlationcoefficient) values of the calibration curve >0.999. BB was used as the internal standard toallow high accuracy and sensitivity. Analysis of the CRM also showed the reliability of the

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results (Tables S1 and S2). For every 16 samples, 2 whole procedure blanks, 2 soil matrixblanks and 1 CRM 136–100 were analyzed.

Statistical Analysis

The removal rates of 6 PAEs were calculated as:

R% = (1 − Cat/Cac) × (Cac/Ctc),where, R is the removal rate of an individual PAE compound, Cat is the residual concentra-tion of an individual target compound under different treatments, Cac is the concentrationof an individual target compound in the control treatment, and Ctc is the total concentrationof the 6 target compounds in the control treatment.

All the data were processed with Microsoft Excel 2003 and the SPSS v.14.0 softwarepackage. The data were analyzed for significant differences from the control treatment orbetween treatments using one-way analysis of variance.

RESULTS AND DISCUSSION

Soil Concentrations and Composition of six Target PAE Compounds

PAE concentrations in soil samples under the different experimental treatments weredetermined by assaying for the six target pollutants remaining in the soil (Figure 1). Com-pared to the control treatment, PAE concentrations in all experimental treatments weresubstantially reduced with highly significant differences (p < 0.01), indicating that effec-tive remediation had been achieved by using the three test plant species. The concentrationsof the six PAE compounds were much lower than the soil allowable concentrations listed byUSEPA (Table 1) after remediation, even though the concentrations of individual PAEs inthe control were near the listed allowable concentrations (Zhang et al. 2010a). The highest

Figure 1 Concentrations of six PAE compounds in soil under the different treatments (µg kg−1 DW). Fortreatment abbreviations see the Phytoremediation Experiment section (Materials and Methods). Each point isthe mean of three replicates. Double asterisks in columns show highly significant difference at p < 0.01 levelcompared to control soil.

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Table 1 Soil allowable concentrations and cleanup objectives of the six PAE compounds in the US

Compound Allowable concentration (mg kg−1) Cleanup objective value (mg kg−1)

DMP 0.020 2.0DEP 0.071 7.1DnBP 0.081 8.1BBP 1.215 50.0DEHP 4.350 50.0DnOP 1.200 50.0

removal rate was observed in the alfalfa monoculture, with a total removal rate of over90%, and the removal rates of all the other treatments also exceeded 80% (Figure 2). Thereare a number of factors that might explain the greater effectiveness of alfalfa monoculturecompared with the other planting combinations in eliminating PAE pollutants. Alfalfa is alegume and is likely to form a nitrogen fixing symbiotic system with rhizobia in the soil.This may promote plant growth and accelerate microbial multiplication by increasing thenitrogen supply in the rhizosphere and thus contribute to phytoremediation. Root noduleswith a healthy appearance were observed on roots of alfalfa when the soil samples werecollected from the field plots, indicating that alfalfa was capable of dinitrogen fixation inour experiment. The large shoot biomass of alfalfa could also be an explanation becauseit removed 22.8 ± 0.67 mg total PAE per plot, the highest removal rate of the differentexperimental treatments investigated (Table 2). The PAE removal rates of the different treat-ments followed the order alfalfa monoculture > alfalfa/ryegrass > ryegrass monoculture >alfalfa/ryegrass/fescue > alfalfa/fescue > fescue monoculture. It has been reported thatintercrops of different remediating plant species have a distinct advantage in eliminatinghydrophobic pollutants such as PAHs compared with monocultures (Maila et al. 2005; Wuet al. 2007), a trend which was not apparent in the present study.

Figure 2 Percentage of six target PAE compounds removed from test soil under different treatments (µg kg−1DW). Each point is the mean of three replicates.

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Table 2 Shoot biomass of test plants under the different treatments

Shoot biomass, kg PAE removal by plant shoots,Treatment DM per plot mg per plot

A 6.78 ± 0.20a 22.78 ± 0.67P 2.58 ± 0.11c 8.01 ± 0.34T 1.49 ± 0.03d 4.55 ± 0.09AP-A 4.09 ± 0.04e 12.32 ± 0.12 15.07 ± 0.23AP-P 0.96 ± 0.04f 2.75 ± 0.11AT-A 2.28 ± 0.06g 7.41 ± 0.20 9.01 ± 0.25AT-T 0.67 ± 0.02b 1.60 ± 0.05APT-A 2.97 ± 0.09h 14.10 ± 0.43 18.04 ± 0.59APT-P 0.65 ± 0.02b 2.71 ± 0.08APT-T 0.32 ± 0.02i 1.23 ± 0.08

For treatment abbreviations see the Phytoremediation Experiment section (Materials and Methods).Each biomass value is the mean of four replicates ± standard derivation (SD).Different letters in columns denote significant difference at p< 0.05 level between treatments.

DEHP and DnBP were found to be major PAE compounds in all the soil samples.Their initial proportions in control soil were about 90% and 7%, respectively. DEHP wasthe compound most effectively removed of the six target pollutants, usually with over 60%total removal except in the control. This agrees with previous conclusions regarding therelationship between structure and characteristics of PAE compounds. Compounds withlonger alkyl chains are more readily adsorbed and accumulate as time proceeds. The sec-ond most effectively removed pollutant, DnBP, had total removal rates >5% (individualremoval of over 70%) in the various experimental treatments (other than the control), whichmight be regarded is an acceptable result for phytoremediation. Alfalfa monoculture andalfalfa/ryegrass intercrop were more competitive than the other treatments, especially inelimination of DEHP. The two grass species, perennial ryegrass and tall fescue, demon-strated little difference in the removal of DEHP and DnBP in this experiment. The grassesextracted slightly more DnOP than the legume even when they were intercropped with theleguminous species.

Our results indicate the feasibility of phytoremediation of the contaminated soil irre-spective of whether monoculture or intercropping was used, although the process requiresa longer time period, usually more than one year, than biodegradation by microorganismsand other remediation methods such as photocatalytic degradation within 24 h (Zeeb et al.2006; Xu et al. 2010). Alfalfa may be considered to be useful for the removal of both totalPAEs and typical PAE pollutants in the soil.

Concentrations of Six PAE Compounds in Plant Shoots

Figure 3 lists and compares the concentrations of the six target pollutants in plantshoots in the different treatments. Total concentrations of the six PAE pollutants in individualplant species were high in the three-species intercrop, alfalfa/ryegrass/fescue, with almostall present at >4 mg kg−1 (DW) in the individual species. However, alfalfa monocultureremoved the largest amount of total PAE per plot of 22.8 ± 0.67 mg (Table 2) with thehelp of its large shoot biomass. This is equivalent to a removal rate of about 1.7 ± 0.05 mgg ha−1 to a depth of 15 cm. Alfalfa shoots accumulated the highest concentrations of the

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target pollutants. DEHP and DnBP also comprised the bulk of the total concentrations,especially in alfalfa shoots across treatments.

PAE concentrations in plant roots were not included in the study because the largeroot systems are difficult to sample quantitatively without soil contamination after twoyears of plant growth. However, variation between individual plant species might be due,at least in part, to different capacities of their root systems to take up PAEs, as this hasbeen found to be true for PCBs (Schnoor et al. 1995). From a scientific perspective itwould be preferable to include the roots to elucidate the dynamics of the pollutants in thewhole soil-plant system. However, in practice the plant shoots are more important becausephytoremediation involves harvesting the plant shoots and removing them from the site.

The extent of assimilation in the plant shoots depends on both extraction from con-taminated soil and absorption from the atmosphere. Previous studies have suggested thatatmospheric absorption is a principal route of absorption in vegetables in agriculturalland near plastics manufacturing industries (Wang et al. 2010) and DEHP was absorbedmainly through the leaves (Schmitzer et al. 1988). Foliar absorption may therefore make acontribution to plant PAE absorption at our experimental site at Taizhou.

Bioconcentration Factors in Plant Shoots

Comparing the BCFs of PAE compounds in the shoots of the different plant speciesindicates that direct absorption and metabolism might be the main mechanisms by whichPAE-contaminated soil is phytoremediated. The sum of the BCFs of alfalfa in the three-species intercrop was found to be the largest among all the shoot samples (Figure 4). Thelarge biomass of alfalfa shoots may have increased the capability of this species to accumu-late PAE compounds to the aboveground parts (Table 2). An advantage of intercropping isindicated by the observation that alfalfa intercropped with the other species showed largerBCFs than did alfalfa in monoculture.

Figure 4 BCFs of six PAE compounds in shoots under different treatments (µg kg−1 DW). Each point is themean of three replicates.

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The BCF of DnBP was relatively low in the three-species intercrop whereas thatof DEHP was relatively low in monoculture treatments of the three plant species. Aninteresting observation is the large BCF of DnOP in all treatments, especially in alfalfain the three species intercrop. The large shoot biomass of alfalfa in this treatment mayhave promoted the accumulation of DnOP from the atmosphere as it is present at lowconcentration in the soil but high in the shoots.

The BCFs of the different compounds indicate that phytoremediation can be adaptedto local conditions, and alfalfa/ryegrass and alfalfa/ryegrass/fescue might be particularlyuseful combinations, especially for soil highly contaminated with DEHP and DnOP.

Phytoextraction Efficiency

Figure 5 indicates that the phytoextraction efficiency of the experimental treatmentsfollowed the order alfalfa/ryegrass/fescue > alfalfa monoculture > ryegrass monoculture> fescue monoculture > alfalfa/ryegrass > alfalfa/fescue and ranged from 1.18 to 1.78%.The three-species intercrop appeared to greatly increase the efficiency of phytoextractioncompared with the monocultures. The major finding is the relatively high capability ofalfalfa in removal of PAEs, but all species combinations showed some phytoremediationcapacity.

CONCLUSIONS

Both alfalfa in monoculture and the three-species intercrop gave satisfactory removalof PAEs from the contaminated soil and intercropping appeared to make alfalfa moreeffective. DEHP and DnBP were the two pollutants that were present at the highest con-centrations and gave the highest removal rates in soil and plant samples. Calculated BCFsindicate the substantial potential of these plant species to accumulate PAEs and the resultsalso indicate that the atmosphere was an important source of PAEs to the plants. The abilityof grasses and legumes to take up PAEs emphasizes the environmental risk from thesecompounds in terms of contamination of the human food chain and possible endocrinedisrupting effects.

ACKNOWLEDGMENTS

This research was supported by the Chinese National Environmental Protection Spe-cial Funds for Scientific Research on Public Causes (Projects 201109018 and 2010467016)and the Program of Innovative Engineering of the Chinese Academy of Sciences (ProjectKZCX2 –YW -Q02 -06 -02).

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SUPPORTING INFORMATION

Table S1 Analysis parameters of the six PAE target compounds

Compound IDL (µg l−1) MDL (mg kg−1) Linear Equation Correlation Coefficient (R2)

DMP 0.35 0.089 y = 136.97x + 47.33 0.9994DEP 0.19 0.068 y = 506.27x + 328.78 0.9993DnBP 0.17 0.097 y = −598.73x + 166.04 0.9995BBP 0.25 0.073 y = −2588x + 88.91 0.9990DEHP 0.21 0.118 y = −2913x + 139.23 0.9993DnOP 0.11 0.135 y = −245.2x + 163.77 0.9991

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Table S2 QA/QC report of PAE spiked matrix samples

Recovery of Given Confidence Determined ValueCompound EPA-8270 (%) Recovery (%)a Interval (mg kg−1) (mg kg−1) b

DMP 93.18 75.88 ± 4.30 2.85−3.4 2.72 ± 0.14DEP 95.75 78.09 ± 3.88 1.29−1.64 1.55 ± 0.22DnBP 90.77 81.20 ± 3.91 0.643−0.797 0.767 ± 0.19BBP 91.21 92.33 ± 4.57 6.61−8.32 7.32 ± 0.19DEHP 94.20 99.87 ± 8.91 0.821−0.961 0.927 ± 0.49DnOP 89.90 107.61 ± 7.47 4.78−5.71 5.62 ± 0.28

aMean value of recovery ± standard deviation (SD) at 0.1 mg kg−1; n = 7.bMean value of recovery ± SD; n = 6.

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