International Journal of Phytoremediation
ISSN: 1522-6514 (Print) 1549-7879 (Online) Journal homepage: http://www.tandfonline.com/loi/bijp20
Enhanced phytoremediation of cadmium and/or benzo(a)pyrene contaminated soil by hyperaccumlator Solanum nigrum L. Ran Han, Huiping Dai, Chuanjie Yang, Shuhe Wei, Lei Xu, Wei Yang & Xuekai Dou To cite this article: Ran Han, Huiping Dai, Chuanjie Yang, Shuhe Wei, Lei Xu, Wei Yang & Xuekai Dou (2018) Enhanced phytoremediation of cadmium and/or benzo(a)pyrene contaminated soil by hyperaccumlator Solanum nigrum L., International Journal of Phytoremediation, 20:9, 862-868, DOI: 10.1080/15226514.2018.1438357 To link to this article: https://doi.org/10.1080/15226514.2018.1438357
Published online: 06 Jun 2018.
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INTERNATIONAL JOURNAL OF PHYTOREMEDIATION 2018, VOL. 20, NO. 9, 862–868 https://doi.org/10.1080/15226514.2018.1438357
Enhanced phytoremediation of cadmium and/or benzo(a)pyrene contaminated soil by hyperaccumlator Solanum nigrum L. Ran Hana,c, Huiping Daib, Chuanjie Yanga, Shuhe Weia, Lei Xua,c, Wei Yanga,c, and Xuekai Doua,c a Pollution Ecology and Environmental Engineering, Institute of Applied Ecology, Chinese Academy of Sciences, Shenyang, P.R. China; bShaanxi Province Key Laboratory of Bio-resources, Shaanxi University of Technology, Hanzhong, China; cUniversity of Chinese Academy of Sciences, Beijing, P.R. China
ABSTRACT
KEYWORDS
The role of same amendment on phytoremediating different level contaminated soils is seldom known. Soil pot culture experiment was used to compare the strengthening roles of cysteine (CY), EDTA, salicylic acid (Sa), and Tween 80 (TW) on hyperaccumulator Solanum nigrum L. phytoremediating higher level of single cadmium (Cd) or Benzo(a)pyrene (BAP) and their co-contaminated soils. Results showed that the Cd capacities (ug pot¡1) in shoots of S. nigrum in the combined treatment T0.1EDTAC0.9CY were the highest for the 5 and 15 mg kg¡1 Cd contaminated soils. When S. nigrum remediating co-contaminated soils with higher levels of Cd and BAP, that is, 5 mg kg¡1 Cd C 1 mg kg¡1 BAP and 15 mg kg¡1 Cd C 2 mg kg¡1 BAP, the treatment T0.9CYC0.9SaC0.3TW showed the best enhancing remediation role. This results were different with co-contaminated soil with 0.771 mg kg¡1 Cd C 0.024 mg kg¡1 BAP. These results may tell us that the combine used of CY, SA, and TW were more useful for the contaminated soils with higher level of Cd and/ or BAP. In the combined treatments of SaCTW, CY was better than EDTA.
cadmium (cd); benzo(a) pyrene (bap); solanum nigrum L
Introduction Cd (Cadmium) is one of heavy metals to be mainly concerned in environmental science. PAHs (polycyclic aromatic hydrocarbons) is another important organic pollutant and Benzo(a)pyrene (BAP) is its typical representative. Single Cd or PAHs contaminated soil is normal in environment but their co-pollution is not quite often. Usually, combustion of coal, oil and wood, waste incineration and automotive exhaust emission, wastewater irrigation, sludge applications, solid waste disposal, and industrial activities may cause Cd and PAHs co-contaminated soil (Pitts and Pitts 2000; Yang et al. 2011; Zeng et al. 2015). The remediation of Cd and PAHs co-contaminated soil is very difficult, especially for the agricultural field with low concentration and very huge area. Phytoremediation which mainly use some special plants to remove pollutants provides a reliable and cost-effective method due to plant is easy to be observed and operated (Wei et al. 2011; Liu et al. 2016; Tao et al. 2016; Wang et al. 2017). As for heavy metal and organic pollutant co-contaminated soil, two remediating pathways are important. One is phytoextraction, that is, hyperaccumulator was used to extremely accumulate heavy metal from soil, then the accumulated heavy metal was removed accompany with the plant biomass taken away from the field. Another pathway of phytoremediation is the degradation of contaminants in rhizosphere through the chemical and/or microbial activity in rhizospherial zone of plant (Shaw and Burns 2003). However, the capacities (accumulation content) of phytoextraction and rhizosphere degradation were not very strong due to the lower bioavailability of heavy metal and organic CONTACT Shuhe Wei
[email protected]
© 2018 Taylor & Francis Group, LLC
pollutant in soil. Some chelators such as EDTA, EGTA, EDDS, and NTA were used to enhance heavy metal accumulation in shoot and thus to achieve a higher removal rate from contaminated soil. Though some chelators showed toxic effect on plant growth, the strengthen roles of them were really significant through increased heavy metal available speciation in soil. Usually, EDTA was typical and quite often used among of these chelators (Martha 2005; Zaier et al. 2010). The degradation of PAHs is very difficult in soil due to its low bioavailability. The environmental condition in rhizosphere is more active than bulk soil due to the increase of chemical matter and diverse community of microbes caused by root secretion (Lee et al. 2008). The addition of surfactant like Tween 80 (TW) can enhance the solubility of PAHs in rhizosphere and then increased its degradation rates (Zhu et al. 2008). S. nigrum was discovered and confirmed as a Cd hyperaccumulator since 2005 (Wei et al. 2005; Xu et al. 2009; Gao et al. 2010; Xiao et al. 2010; Ji et al., 2011; Wan et al. 2012). Benzo(a) pyrene (BAP) is a typical representative of PAHs. Yang et al. studied the co-phytoremediation effects of S. nigrum on CdPAHs contaminated soil with the improvements of some reagents (Yang et al. 2011). The results showed that the strengthening roles of ethylenediaminetetraacetic acid (EDTA), cysteine (CY), salicylic acid (Sa), and Tween 80 (TW80) for S. nigrum phytoremediating low levels of Cd-PAHs co-contaminated soils were significant. However, what will happen for S. nigrum when Cd or BAP concentration in soil is very high? We suppose that the strengthen roles of these reagents may be different. Thus, this study was used to compare the strengthen roles of CY, EDTA, Sa, and TW on hyperaccumulator
72, Wenhua Road, Shenhe Shenyang, 110016 P.R. China.
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S. nigrum phytoremediating higher level of single Cd or BAP and their co-contaminated soils.
Materials and methods Soil and plant seedling preparation Soil samples collected from top layer (0–20 cm) of a farmland in the suburb of Shenyang city, China, was meadow brown soil with pH 6.9, an organic matter content of 1.79%, soil soluble organic carbon (SOC) of 45.7 ng kg¡1, a total N, P, and K content of 1.32, 1.39, and 0.14 g kg¡1. The results showed that Cd and 16 polycyclic aromatic hydrocarbons (PAHs) from the US EPA priority pollutant list were 0.771 and 0.569 mg kg¡1 (BAP was 0.024 mg kg¡1), which were low levels of Cd and PAHs co-contaminated soil compared to the National Soil-Environmental Quality Standards of China (GB 15618, 1995), the remediation standard of PAH-contaminated soil in the Netherlands, and local pollution level in Shenyang (Yang et al. 2011). Seeds of S. nigrum were sterilized by 0.5% NaClO for 10 min, then sowed in soil tray for germinating. When the seedlings were with 3–4 leaves and 10 cm height, they are prepared to be used in different treatments.
Soil pot experiment design Air-dried collected soil samples were sieved through 1 mm nylon mesh, then transferred into plastic pots with 18 cm in diameter and 15 cm in height, yielding a total dry weight of 1.0 kg of soil per pot. At the meantime, 1 g (NH4)2SO4 as fertilizer was added in each pot. Some reagents of CdCL22.5H2O and BAP were spiked to different pots according to the experiment design and all pots were kept in a control room for two months. Reagents of CY, EDTA, Sa, and TW were spiked after all seedlings of S. nigrum transplanted to pots in accordance with the designed plan. There were three different experiment designs. According to the environmental conditions in local field, the Cd concentrations were added at 5 and 15 mg kg¡1, and BAP were 1 and 2 mg kg¡1, which were higher levels of contaminated soils compared to the National Soil-Environmental Quality Standards of China (GB 15618, 1995), the remediation standard of PAHcontaminated soil in the Netherlands, and local pollution level in Shenyang (Yang et al. 2011). The concentrations of strengthening reagents of EDTA, CY, TW and Sa were added at 0.1, 0.9, 0.3, and 0.9 mmol Kg¡1, respectively, based on the best remediation results of published (Yang et al. 2011). The first experiment was phytoremediation of single higher level of Cd contaminated soil, that is, 5 and 15 mg kg¡1 Cd were added respectively. There were nine treatments as shown in Table 1. The second experiment was phytoremediation of single high level of BAP polluted soil. There were 10 treatments as shown in Table 2. The third experiment was phytoremediation of high levels of Cd and BAP co-contaminated soil. There were 20 treatments as shown in Table 3.
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Table 1. Phytoremediation of single higher level of Cd-contaminated soil. Treatment CK CK11 T11 (T0.1EDTA) T12 (T0.9CY) T13 (T0.1EDTAC0.9CY) CK12 T14 (T0.1EDTA) T15 (T0.9CY) T16 (T0.1EDTAC0.9CY)
Detail treatment information S. nigrum grown in original soil S. nigrum grown in original soil with 5 mg kg¡1 Cd added S. nigrum grown in original soil with 5 mg kg¡1 Cd and 0.1 mmol Kg¡1 EDTA added S. nigrum grown in original soil with 5 mg kg¡1 Cd and 0.9 mmol Kg¡1 CY added S. nigrum grown in original soil with 5 mg kg¡1 Cd, 0.1 mmol Kg¡1 EDTA and 0.9 mmol Kg¡1 CY added S. nigrum grown in original soil with 15 mg kg¡1 Cd added S. nigrum grown in original soil with 15 mg kg¡1 Cd and 0.1 mmol Kg¡1 EDTA added S. nigrum grown in original soil with 15 mg kg¡1 Cd and 0.9 mmol Kg¡1 CY added S. nigrum grown in original soil with 15 mg kg¡1 Cd, 0.1 mmol Kg¡1 EDTA and 0.9 mmol Kg¡1 CY added
Plant growth and management Prepared seedlings of S. nigrum were transplanted to every treated pot of experiment 1, 2, and 3 with two uniform plants each. All treatments were repeated for three times and pots were randomly lined on the culture shelves in a control room with 10 h photoperiod (08:00–18:00) daily, day temperature about 25 § 4 C and night temperature about 19 § 3 C and a photo synthetic photon flux density about 350–450 mmol m¡2s¡1 (Yang et al. 2011). Strengthening reagents of EDTA, CY, TW, and Sa were spiked to different pots based on the designs of experiment 1, 2, and 3 when tap water was made up. All strengthening reagents were spiked for three times, that is, every other 15 days after plant seedlings transplanted for half month. Tap water was used to replenish the losses of the pot soils to 80% soil water-holding capacity. When S. nigrum was at maturity stage (80 days), all plants were harvested and rhizosphere soil samples were collected either. Cd and BAP concentrations in the rhizospherial soils were determined to calculate the Cd capacity and BAP degradation rate after phytoremediation. The rhizosphere soil was collected by shaking method (Yang et al. 2011). Table 2. Phytoremediation of single high level of BAP-polluted soil. Treatment CK21 CK22 T21 (T0.9Sa) T22 (T0.3TW) T23 (T0.3TWC0.9Sa) CK23 CK24 T24 (T0.9Sa) T25 (T0.3TW) T26 (T0.3TWC0.9Sa)
Detail treatment information Original soil with 1 mg kg¡1 BAP added S. nigrum grown in original soil with 1 mg kg¡1 BAP added S. nigrum grown in original soil with 1 mg kg¡1 BAP and 0.9 mmol Kg¡1 Sa added S. nigrum grown in original soil with 1 mg kg¡1 BAP and 0.3 mmol Kg¡1 TW added S. nigrum grown in original soil with 1 mg kg¡1 BAP, 0.3 mmol Kg¡1 TW and 0.9 mmol Kg¡1 Sa added original soil with 2 mg kg¡1 BAP added S. nigrum grown in original soil with 2 mg kg¡1 BAP added S. nigrum grown in original soil with 2 mg kg¡1 BAP and 0.9 mmol Kg¡1 Sa added S. nigrum grown in original soil with 2 mg kg¡1 BAP and 0.3 mmol Kg¡1 TW added S. nigrum grown in original soil with 2 mg kg¡1 BAP, 0.3 mmol Kg¡1 TW and 0.9 mmol Kg¡1 Sa added
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Table 3. Phytoremediation of Cd and BAP co-contaminated soil. Treatment CK31 CK32 T33 (T0.1EDTAC0.9Sa) T34 (T0.1EDTAC0.3TW) T35 (T0.9CYC0.9Sa) T36 (T0.9CYC0.3TW) T37 (T0.1EDTAC0.9CYC0.9Sa) T38 (T0.1EDTAC0.9CYC0.3TW) T39 (T0.1EDTAC0.3TWC0.9Sa) T40 (T0.9CYC0.3TWC0.9Sa) CK31 CK32 T43 (T0.1EDTAC0.9Sa) T44 (T0.1EDTAC0.3TW) T45 (T0.9CYC0.9Sa) T46 (T0.9CYC0.3TW) T47 (T0.1EDTAC0.9CYC0.9Sa) T48 (T0.1EDTAC0.9CYC0.3TW) T49 (T0.1EDTAC0.3TWC0.9Sa) T50 (T0.9CYC0.3TWC0.9Sa)
Detail treatment information Original soil with 5 mg kg¡1 Cd and 1 mg kg¡1 BAP added S. nigrum grown in original soil with 5 mg kg¡1 Cd and 1 mg kg¡1 BAP added S. nigrum grown in original soil with 5 mg kg¡1 Cd, 1 mg kg¡1 BAP, 0.1 mmol Kg¡1 EDTA and 0.9 mg kg¡1 Sa added S. nigrum grown in original soil with 5 mg kg¡1 Cd, 1 mg kg¡1 BAP, 0.1 mmol Kg¡1 EDTA and 0.3 mmol Kg¡1 TW added S. nigrum grown in original soil with 5 mg kg¡1 Cd, 1 mg kg¡1 BAP, 0.9 mmol Kg¡1 CY and 0.9 mmol Kg¡1 Sa added S. nigrum grown in original soil with 5 mg kg¡1 Cd, 1 mg kg¡1 BAP, 0.9 mmol Kg¡1 CY and 0.3 mmol Kg¡1 TW added S. nigrum grown in original soil with 5 mg kg¡1 Cd, 1 mg kg¡1 BAP, 0.1 mmol Kg¡1 EDTA, 0.9 mmol Kg¡1 CY and 0.9 mmol Kg¡1 Sa added S. nigrum grown in original soil with 5 mg kg¡1 Cd, 1 mg kg¡1 BAP, 0.1 mmol Kg¡1 EDTA, 0.9 mmol Kg¡1 CY and 0.3 mmol Kg¡1 TW added S. nigrum grown in original soil with 5 mg kg¡1 Cd, 1 mg kg¡1 BAP, 0.1 mmol Kg¡1 EDTA, 0.3 mmol Kg¡1 TW and 0.9 mmol Kg¡1 Sa added S. nigrum grown in original soil with 5 mg kg¡1 Cd, 1 mg kg¡1 BAP, 0.9 mmol Kg¡1 CY, 0.3 mmol Kg¡1 TW and 0.9 mmol Kg¡1 Sa added Original soil with 5 mg kg¡1 Cd and 2 mg kg¡1 BAP added S. nigrum grown in original soil with 5 mg kg¡1 Cd and 2 mg kg¡1 BAP added S. nigrum grown in original soil with 5 mg kg¡1 Cd, 2 mg kg¡1 BAP, 0.1 mmol Kg¡1 EDTA and 0.9 mg kg¡1 Sa added S. nigrum grown in original soil with 5 mg kg¡1 Cd, 2 mg kg¡1 BAP, 0.1 mmol Kg¡1 EDTA and 0.3 mmol Kg¡1 TW added S. nigrum grown in original soil with 5 mg kg¡1 Cd, 2 mg kg¡1 BAP, 0.9 mmol Kg¡1 CY and 0.9 mmol Kg¡1 Sa added S. nigrum grown in original soil with 5 mg kg¡1 Cd, 2 mg kg¡1 BAP, 0.9 mmol Kg¡1 CY and 0.3 mmol Kg¡1 TW added S. nigrum grown in original soil with 5 mg kg¡1 Cd, 2 mg kg¡1 BAP, 0.1 mmol Kg¡1 EDTA, 0.9 mmol Kg¡1 CY and 0.9 mmol Kg¡1 Sa added S. nigrum grown in original soil with 5 mg kg¡1 Cd, 2 mg kg¡1 BAP, 0.1 mmol Kg¡1 EDTA, 0.9 mmol Kg¡1 CY and 0.3 mmol Kg¡1 TW added S. nigrum grown in original soil with 5 mg kg¡1 Cd, 2 mg kg¡1 BAP, 0.1 mmol Kg¡1 EDTA, 0.3 mmol Kg¡1 TW and 0.9 mmol Kg¡1 Sa added S. nigrum grown in original soil with 5 mg kg¡1 Cd, 2 mg kg¡1 BAP, 0.9 mmol Kg¡1 CY, 0.3 mmol Kg¡1 TW and 0.9 mmol Kg¡1 Sa added
Sample determination and Data processing Plants in the pots were separated into roots and shoots (stem, leaf, and fruit) to determine biomasses and Cd concentrations. Tap water was firstly used to rinse all plant samples, and then deionized water finally washed all. After oven-dried at 105 C for 5 min, and then at 70 C for about 2 days till constant dry weight, plant samples were ground to powder and passed through a 0.3 mm sieve. Powdered plant samples were digested using concentrated nitric acid and perchlorate (87% HNO3/13% HClO4). The obtained solutions were analyzed for Cd by atomic absorption spectrophotometry (AAS, WFX-120A with a 1.3 nm spectral bandwidth). For QA/QC, measured values of Cd were verified by using certified standard reference materials (NIST SRM 1547, peach leaves) (Yang et al. 2011; Wei et al. 2016). pH was measured with a pH meter and electrode (PHS-3B) in soil slurries of a soil: water ratio of 1:2.5. Basic soil properties were determined by normal method introduced by Lu et al. (Lu 2000). EDTA, CY, TW, and Sa were all purchased from the DuPont De Nemours & Co., USA. A standard of BAP was obtained from the Chem Service Inc. (West Chester, USA). All solvents such as n-hexane, dichloromethane, and cyclohexane used for sample processing were of analytical grade. Acetonitrile used for HPLC analysis was of HPLC grade. BAP concentration in 5.0 g of air-dried soil sample was
determined by HPLC (Waters 1525, USA) after extraction. The HPLC system comprised a Multi λ Fluorescence Detector and Dual λ Absorbance Detector, equipped with an Agilent ZORBAX Eclipse PAH column (4.6 £ 250 mm, 5 mm) (Yang et al. 2011; Wei et al. 2016). Microsoft EXCEL was used to process and calculate Data and standard deviation. The treatment responses were analyzed by one-way ANOVA and LSD multiple range tests to separate means using the SPSS software. Differences were considered significant at the p < 0.05 level (Wei et al. 2016).
Result and discussion Strengthen roles of reagents on S. nigrum remediation of single Cd polluted soil The accumulation capacity (ug plant¡1) reflects the remediation efficacy of hyperaccumulator removing heavy metal from contaminated soil (Shaw et al. 2003). Higher accumulation capacity means higher remediation efficacy. Usually, the roots of hyperaccumulator are very difficult to be removed from contaminated soil and easily kept in soil. Thus, higher accumulation capacity in shoots means higher remediation efficacy of hyperaccumulator. The accumulation capacity in shoots is equal to the product of its biomass and heavy metal concentration. Thus, shoot biomass is very important (Zhou and Song 2004).
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paper by Yang et al., the treatment T0.1EDTAC0.9CY (T13 and T16) was the best for S. nigrum accumulating Cd at 5 and 15 mg kg¡1 in soils in this experiment (Yang et al. 2011; Figure 1). This result showed that the combined use of EDTA and CY is needed when S. nigrum phytoremediating higher level of Cd-contaminated soil.
Strengthening roles of reagents on S. nigrum remediating single BAP polluted soil Usually, organic pollutant like BAP is degraded by some reactions in rhizoshperial zone of plant and the accumulation of biomass contributed seldom (Lee et al. 2008). Shoot biomass reflects root growth, that is, higher shoot biomass is often with higher root biomass (Yang and Zheng 1989; Wei et al. 2011). Thus, shoot biomass also showed the effect of BAP on S. nigrum growth through its role on root. The biomasses of CK22 (1 mg kg¡1 BAP spiked) and CK24 (2 mg kg¡1 BAP spiked) were 3.12 g plant¡1 and 3.02 g plant¡1, which were not significantly decreased (p < 0.05) compared to the control CK (3.60 g plant¡1, initial soil). There were not significantly differences (p < 0.05) among of shoot biomasses of S. nigrum in treatments of Ck22, T21, T22, T23 and CK24, T24, T25, T26, indicating the addition of 1 and 2 mg kg¡1 BAP, and spiked with reagents of TW and Sa didn’t impact on the growth of S. nigrum. BAP degradation rate was listed in Figure 2. When 1 mg kg¡1 BAP was added, the degradation rates of BAP in treatments of CK21 and CK22 were 3.4% and 3.1%. As for higher BAP pollution with 2 mg kg¡1 BAP addition, its degradation rates were 6.1% and 6.2% (CK23 and CK24). Compared to the control CK21 and CK23 (without plants), the growth of S. nigrum in CK22 and CK24 (without reagents added) were not significantly increased (p < 0.05) the degradation rates of BAP, indicating the concentration of BAP in shoot or root was negligible. Thus, the BAP concentration in plant was not determined. The same result was approved by Moutaz et al. (Moutaz et al. 2015). When reagents of TW, Sa, and their compound were spiked, BAP degradation rates were significantly increased (p < 0.05) by 17.7%, 16.4%, and 29.5% for treatments of T21, T22, and T23, and 26.7%, 33.3%, and 31.5% for
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The shoot biomass of S. nigrum in treatment of CK (initial soil without Cd addition) was 3.60 g plant¡1. The shoot biomasses of S. nigrum in CK11 and CK12 were 3.55 and 2.79 g plant¡1, which were not significantly decreased (p < 0.05) compared to the control CK, indicating strong tolerance of S. nigrum to Cd. Basically, the addition of EDTA, CY, and their compound reagents showed seldom impact on shoot biomasses of S. nigrum, suggesting their very weak toxic under these concentration conditions to plant either (T11, T12, T13, T14, T15, and T16). Cd concentration (mg kg¡1) and capacity (ug plant¡1) in shoots of S. nigrum were showed in Figure 1. Under the condition of 5 mg kg¡1 of Cd added, the addition of EDTA, CY and their compound reagents were all significantly increased (p < 0.05) Cd accumulation in shoots of S. nigrum, indicating their available strengthening roles. The treatment of compound reagents (T3) remediation effect was better compared to the control CK11, T1, and T2, that is, Cd capacity in shoot (117.5 ug plant¡1) was significantly increased (p < 0.05) by 39.6% than CK11 (Figure 1). Similarly, the addition of EDTA, CY and their compound reagents also increased Cd concentration and capacity in shoots of S. nigrum when 15 mg kg¡1 Cd in soils (CK12, T14, T15, and T16). Particularly, the addition of compound reagents of EDTA and CY obtained the highest remediation effect, that is, Cd capacity (164.77 ug plant¡1) in shoots of S. nigrum was increased (p < 0.05) by 34.1% (Figure 1). Compared to the treatment T0.9CY improved S. nigrum phytoextracting 0.771 mg kg¡1Cd contaminated soil in published
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Figure 1. Cd concentration and capacity in shoot of S. nigrum remediating single Cd polluted soil. (The bars represent standard deviation. In different treatment, data followed by same letter over column bar are not significantly different (p < 0.05).)
Figure 2. Degradation rate of S. nigrum remediating single BAP polluted soil. (The bars represent standard deviation. In different treatment, data followed by same letter over column bar are not significantly different (p < 0.05).)
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treatments of T24, T25, and T26. Obviously, the strengthening role of compound reagents of TW and Sa were the highest in the treatments of 1 and 2 mg kg¡1 BAP added (Figure 2). Compared to the treatment T0.3TW or T0.9Sa improved S. nigrum phytoremediating 0.024 mg kg¡1 BAP contaminated soil in published paper by Yang et al., the treatment T0.3TWC0.9Sa (T23 and T26) was the best for S. nigrum degradating BAP at 1 and 2 mg kg¡1 in soils in this experiment (Yang et al. 2011; Figure 2). This result showed that the combined use of TW and Sa is needed when S. nigrum phytoremediating higher level of BAP-contaminated soil. Strengthening roles of reagents on S. nigrum remediating Cd and BAP copolluted soil
Shoot Cd capacity (ug plant-1)
In the treatments of Cd and BAP coadded soil, the shoots biomasses of S. nigrum in CK32 (5 mg kg¡1 Cd and 1 mg kg¡1 BAP added, without other reagents) and CK42 (15 mg kg¡1 Cd and 2 mg kg¡1 BAP added, without other reagents) were 2.71 and 2.69 g plant¡1, respectively, which were significantly decreased (p < 0.05) compared to the CK (initial soil without Cd and BAP addition), suggesting that Cd or BAP toxicity was increased when they were added together. The addition of two or three compound reagents like CY, Sa, TW, or EDTA in the treatments of T35, T36, T37, T38, T39, and T40 significantly increased (p < 0.05) the shoot biomasses of S. nigrum compared to the control of CK32, but their biomasses significantly decreased (p < 0.05) compared to CK42 when added Cd and BAP concentration were increased from 5 and 1 mg kg¡1 to 15 and 2 mg kg¡1 (T45, T46, T47, T48, T49, and T50).
As shown in Figure 3, Cd concentration and capacity in shoots of S. nigrum in control CK32 (5 mg kg¡1 Cd and 1 mg kg¡1 BAP added, without other reagents addition) were 29.30 mg kg¡1 and 41.59 ug plant¡1, respectively. When the addition of Cd and BAP were increased to 15 mg kg¡1 Cd and 2 mg kg¡1, Cd concentration and capacity in shoots were 28.7 mg kg¡1 and 38.58 ug plant¡1 (CK42). Compared to the control CK32 and CK42, the addition of compound Sa, TW, and CY (T40 and T50) showed the best strengthening role on shoot accumulation of Cd by S. nigrum (Cd in shoot capacities were 123.06 ug plant¡1 for T40 and 149.21 ug plant¡1 for T50) (Figure 3). BAP degradation in rhizosphere of S. nigrum rate in control CK32 and CK42 (plants were transplanted without reagents added) were 2.3% and 1.1%, which were not significantly increased (p < 0.05) compared to the treatments of CK31 and CK41 without plant growth (Figure 4). When 5 mg kg¡1 Cd and 1 mg kg¡1 BAP were added, the degradation rates of BAP in treatments of T35 and T40 were the highest (p < 0.05) with 20.7% and 18.1% (Figure 4). However, BAP degradation rates were significantly increased (p < 0.05) in treatments of T45, T49, and T50 even when 15 mg kg¡1 Cd and 2 mg kg¡1 BAP were spiked, that is, BAP degradation rates were 39.2%, 36.3%, and 33.6%, respectively (Figure 4). These results showed that the strengthening roles of compound reagents were better. Actually, the treatments of T40 and T50 were the same reagents (the compound Sa, TW, and CY), which was the best strengthening reagent both in 5 mg kg¡1 Cd plus 1 mg kg¡1 BAP and 15 mg kg¡1 Cd plus 2 mg kg¡1 combine polluted soils (Figure 4). a
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