Influence of silicon treatment on antimony uptake and translocation in rice genotypes with different radial oxygen loss

Ecotoxicology and Environmental Safety 144 (2017) 572–577

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Influence of silicon treatment on antimony uptake and translocation in rice genotypes with different radial oxygen loss

MARK

⁎

Liping Zhanga,1, Qianqian Yanga,1, Shiliang Wanga,b, , Wanting Lia, Shaoqing Jiangc, Yan Liua a b c

School of Geography and Tourism, Qufu Normal University, Rizhao 276826, China School of Environment, Tsinghua University, Beijing 100084, China Changshushi Middle School, Changshu 215500, China

A R T I C L E I N F O

A B S T R A C T

Keywords: Antimony Silicon Rice Radial oxygen loss

Antimony (Sb) pollution in soil may have a negative impact on the health of people consuming rice. This study investigated the effect of silicon (Si) application on rice biomass, iron plaque formation, and Sb uptake and speciation in rice plants with different radial oxygen loss (ROL) using pot experiments. The results demonstrated that Si addition increased the biomass of straw and grain, but had no obvious impact on the root biomass. Indica genotypes with higher ROL underwent greater iron plaque formation and exhibited more Sb sequestration in iron plaque. Silicon treatments increased iron levels in iron plaque from the different genotypes but decreased the total Sb concentration in root, straw, husk, and grain. In addition, Si treatment reduced the inorganic Sb concentrations but slightly increased the trimethylantimony (TMSb) concentrations in rice straw. Moreover, rice straw from hybrid genotypes accumulated higher concentrations of TMSb and inorganic Sb than that from indica genotypes. The conclusions from this study indicate that Sb contamination in rice can be efficiently reduced by applying Si treatment and selecting genotypes with high ROL.

1. Introduction Antimony (Sb) is an element in group VA. Although its concentration is relatively low in the natural environment, large quantities of Sb contaminants have been released into the environment by anthropogenic activities such as smelting, mining, shooting, and burning of fossil fuels (Filella et al., 2002; Wilson et al., 2010), which measurably increases the Sb concentration in soil. The increased risk of Sb toxicity has gained attention, especially in industrial zones, near roads, and at mining sites (Filella et al., 2002; Feng et al., 2013). Antimony is toxic to most organisms and is a potential carcinogen for humans (Filella et al., 2002). Therefore, antimony and its compounds have been listed as priority controlled contaminants by the European Union (EU) and the Environmental Protection Agency of the United States. China possesses the largest amount of Sb ore reserves in the world, and Xikuangshan in the Hunan Province is the largest Sb mine in the world. Soil around Sb smelting and mining areas has been heavily polluted by Sb (He and Yang, 1999; He, 2007). Antimony is not necessary for plants, but it can be adsorbed from soil and accumulated by numerous plants. For example, the Sb concentration reached 1367 and 1105 mg/kg in the flowers and leaves of Achillea ageratum, respectively,

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1

and 1150 and 1164 mg/kg in the root and stems of Silene vulgaris, respectively, grown near the Sb mine (Baroni et al., 2000). The Sb concentration ranged from 3.92 to 143.69 mg/kg in plants from Sb mining and smelting areas in the Hunan Province (Qi et al., 2011). Therefore, a serious risk will ensue from Sb entering the food chain through plants growing in Sb-contaminated soil (Vaculík et al., 2013). Excess Sb in soil can retard root elongation and decrease plant biomass (Feng et al., 2013). Previous studies of seed germination and pot experiments indicated that both Sb(III) and Sb(V) measurably inhibited seed germination and root development, and the grain yield and biomass of rice also decreased with increasing Sb levels in soil (He and Yang, 1999). However, information about the tolerance and accumulation of Sb in plants is very limited, and few investigations have been conducted on the effect of Sb on plant physiology and metabolism. Silicon (Si) is the second most abundant element in the Earth's lithosphere. This element is important and beneficial for the growth and development of plants. For example, it can increase the growth and yield of plants and improve their resistance to biotic and abiotic stresses (Ma and Yamaji, 2006). Previous studies have demonstrated that Si application to soil showed positive effects on plant growth in soil contaminated by various heavy metals and toxic elements, such as lead

Corresponding author at: School of Geography and Tourism, Qufu Normal University, Rizhao 276826, China. E-mail address: [email protected] (S. Wang). Both authors contributed equally to this work.

http://dx.doi.org/10.1016/j.ecoenv.2017.06.076 Received 20 November 2016; Received in revised form 27 June 2017; Accepted 30 June 2017 0147-6513/ © 2017 Elsevier Inc. All rights reserved.

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of the soil sample used in this study was carried out, and the soluble and exchangeable, reducible, oxidizable, and residual fractions of Sb in soil were sequentially and selectively extracted, and the concentrations of these four fractions were 0.92, 2.26, 3.25, and 7.17 mg/kg, respectively. The collected soil samples were taken to the laboratory and airdried at room temperature, then slightly ground and sieved to < 2 mm. Fertilizers (including P as CaH2PO4·H2O (at 0.15 g/kg P2O5), K as KCl (at 0.20 g/kg K2O), and N as CO (NH2)2 (at 0.20 g/kg N)) were thoroughly mixed with the above soil samples for the growth of rice seedlings. Antimony solution (potassium hexahydroxoantimonate (V): KSb (OH)6) were applied at 50 mg/kg to all treatments with exception of the control. Silicon was then added as a SiO2 gel. All treatments with Sb and Si addition in this study were as follows: Control, no Si and no Sb addition; Treatment A, Sb only (Si0); Treatment B, Sb and 10 mg Si/kg (Si10); Treatment C, Sb and 20 mg Si/kg (Si20); Treatment D, Sb and 50 mg Si/kg (Si50). All treatments were mixed thoroughly and equilibrated for 20 days. A series of polyethylene pots (20 cm in height, 20 cm in diameter) were filled with 5.0 kg of soil. Three rice seedlings were planted per pot. All treatments were conducted in triplicate. Under the waterlogged conditions of 2 cm water depth above the soil surface, the rice seedlings in the pots were grown in a greenhouse (with a photoperiod of 12 h light (25 °C)/12 h night (20 °C) and a relative humidity of 70%), and a sodium lamp (1200 PAR) supplied natural light. Rice plants were not harvested until the mature stage (90 days after rice seedings transplantation).

(Pb), cadmium (Cd), and arsenic (As) (Vatehová et al., 2012; Fleck et al., 2013). However, little information is known about the impact of Si on Sb toxicity in rice. To our knowledge, this is the first study to investigate the effects of Si application to soil on the Sb toxicity in rice plants. So as to acclimate the oxygen deficient environment, adventitious roots of some wetland plants such as rice (Oryza sativa L.) have developed plentiful aerenchyma and induction of a barrier to radial oxygen loss (ROL), defined as the oxygen transfer from aerenchyma to the rhizosphere (Armstrong, 1979). ROL is important and essential for the detoxification of phytotoxins (McDonald et al., 2001). ROL of rice roots was relevant to As tolerance and accumulation in rice (Wu et al., 2011). In addition, some studies have verified that the rhizosphere oxygenation induced by microbial activities and the plant roots oxygenation induced by ROL can convert Fe2+ to Fe3+, which lead to the iron plaque formation on the root surface (Colmer, 2003). As the main components, ferric hydroxides, goethite, and minor concentrations of siderite accounts for 63%, 32%, and 5% of the iron plaque, respectively (Liu et al., 2004). Therefore, the structure of iron plaque is characterized as amorphous or crystalline iron (oxyhydr) oxides (Liu et al., 2004). Some studies have found that iron plaque can sequester metalloids (e.g. As and Sb), metals, and anions including silicate and carbonate on rice roots (Liu et al., 2004; Liu and Zhu, 2005). Studies have demonstrated that iron plaque has an important effect on As accumulation and toxicity in rice plants (Ultra et al., 2009; Wu et al., 2012). Iron plaque serves as a barrier to prevent As translocation from roots to shoots (Liu et al., 2014). Some previous studies investigate the impact of ROL on As tolerance and uptake (Wu et al., 2011) and the impact of Si on As uptake by rice plants (Li et al., 2009b; Seyfferth and Fendorf, 2012), but investigation on the impact of Si on Sb accumulation and speciation of rice genotypes with different ROL is very limited. The aim of this study is to investigate the effect of Si on iron plaque formation on the root surface of different genotypes of rice, on Sb sequestration by iron plaque, and on Sb distribution and accumulation in different genotypes of rice.

2.3. Extraction analysis of iron plaque Iron plaque on the surface of the rice root was examined by dithionite-citrate-bicarbonate (DCB) extraction, as detailed by Liu et al. (2004). The DCB solution contained sodium citrate (Na3C6H5O7·2H2O, 0.03 M), sodium bicarbonate (NaHCO3, 0.125 M), and sodium dithionite (NaS2O4, 0.06 M). Fresh root from the rice plants (1 g) was soaked in the DCB solution (30 mL) for 1 h at room temperature (25 °C). After that, the roots were rinsed three times with deionized water, and the rinsed water was added to the above DCB extracts. The final extraction solution was increased to 100 mL by the addition of deionized water prior to analysis. The Fe and Sb concentrations in the above DCB-extraction solutions were analyzed. The Fe concentration in the extract was determined by atomic absorption spectroscopy (AAS, TAS-990, Beijing Puxi Instruments Co., P.R. China). The antimony concentration was determined by hydride generation atomic fluorescence spectrometry (HG-AFS, AFS-8230, Beijing Haiguang Instruments Co., China).

2. Materials and methods 2.1. Preparation and treatment of rice seeds Four genotypes of rice (O. sativa L.), including hybrid subspecies Xiangfengyou9 (HX9) and T-you207 (HT207) and indica subspecies Xiangwanxian17 (IX17) and Xiangwanxian12 (IX12), were used in our study. According to the results of a previous study, the radial oxygen loss (ROL) of HX9, HT207, IX17, and IX12 were 9.55, 15.4, 19.7, and 27.0 µmol O2/g root dry weight/h, respectively (Wu et al., 2015). Seeds from these four rice genotypes were purchased from the Chinese Academy of Agricultural Sciences. After disinfection in a hydrogen peroxide (H2O2) solution (30%, w/w) for 20 min, the rice seeds were washed by deionized water, germinated in moist perlite, and then cultured for 15 days in a PVC pot (14 cm in height and 7.5 cm in diameter) with 500 mL of nutrient solution (containing macronutrients (in mM) including KH2PO4, 1.3; MgSO4·7H2O, 1.5; CaCl2, 4.00; K2SO4, 2.0; NH4NO3, 5.0; and micronutrients (in µM) including Na2MoO4·2H2O, 0.5; H3BO4, 10.0; Fe(II)-EDTA, 50.0; ZnSO4·7H2O, 1.0; CuSO4·H2O, 1.0; CoSO4·7H2O,0.2; MnSO4·H2O, 5.0). During the experiment, the nutrient solution was changed twice per week, and the pH of the solution was adjusted to 5.5 with KOH and HCl.

2.4. Analysis of the total Sb in rice plants After being harvested, the rice plants were washed cleanly and divided into root, straw, husk, and grain. These four sections were dried and crushed using a mechanical mill. Then, the milled sample (0.5 g) was weighed and put into a conical flask (100 mL) with 10 mL of concentrated nitric acid. The samples were digested using an electric hot plate (120 °C). After digestion, the solution was increased to 20 mL with ultrapure water. The Sb concentration in the digested solutions from the root, straw, husk, and grain was measured by hydride generation atomic fluorescence spectrometry (HG-AFS, afs-8230, Beijing Haiguang Instruments Co., China). A certified reference material (CRM) (bush twigs and leaves) was used for quality control, and the recovery ratios of Sb were in the range of 85–103%.

2.2. Pot experiments

2.5. Analysis of Sb speciation

The soil sample used in this study was the surface (0–20 cm depth) soils of a paddy field near the mine area of Xikuangshan, located in Lengshuijiang, Hunan Province, China. It had a pH of 6.8 and contained 13.6 mg/kg total Sb. In this study, based on the standard BCR protocol of the European Commission (Rauret et al., 1999), sequential extraction

Cultivars (HX9 and IX12) with the highest and lowest ROL rates were selected to determine the Sb speciation. Milled samples (1.0 g) of straw were put into 50 mL centrifuge tubes and digested with nitric acid 573

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Table 1 Biomass of rice plants from the four genotypes after Si treatment (g/pot; data = mean ± SD). Genotypes

Treatment

Root

Straw

Grain

Exposure concentration (mg/kg soil)

HX9

HT207

IX17

IX12

Control Si0 Si10 Si20 Si50 Control Si0 Si10 Si20 Si50 Control Si0 Si10 Si20 Si50 Control Si0 Si10 Si20 Si50

Si

Sb

（g/pot）

（g/pot）

（g/pot）

0 0 10 20 50 0 0 10 20 50 0 0 10 20 50 0 0 10 20 50

0 50 50 50 50 0 50 50 50 50 0 50 50 50 50 0 50 50 50 50

13.2 ± 1.25 14.4 ± 3.62 13.2 ± 2.54 14.3 ± 1.55 16.3 ± 5.21 8.8 ± 0.94 10.2 ± 4.51 11.3 ± 1.95 11.5 ± 3.42 10.9 ± 2.51 10.3 ± 1.04 9.4 ± 4.36 10.1 ± 0.89 9.7 ± 3.42 10.2 ± 1.68 8.5 ± 0.75 8.7 ± 2.85 9.2 ± 1.21 8.8 ± 3.54 8.9 ± 0.78

20.5 ± 5.47 19.4 ± 2.42 20.6 ± 6.38 25.5 ± 2.68 24.7 ± 7.58 18.1 ± 1.54 19.2 ± 2.75 22.5 ± 3.28 26.9 ± 8.24 24.1 ± 4.02 27.9 ± 3.28 25.8 ± 5.81 26.9 ± 1.94 30.8 ± 7.62 34.1 ± 4.21 16.9 ± 2.04 15.4 ± 1.73 15.6 ± 1.22 15.8 ± 2.41 17.2 ± 3.98

8.1 ± 0.78 9.6 ± 1.78 10.3 ± 0.67 14.8 ± 3.59 16.2 ± 5.84 10.5 ± 1.32 9.6 ± 1.65 10.1 ± 2.62 16.4 ± 6.84 14.9 ± 3.65 14.5 ± 1.57 13.1 ± 0.87 14.2 ± 1.76 15.6 ± 2.01 17.1 ± 5.47 6.9 ± 0.87 6.8 ± 1.61 10.1 ± 0.79 10.6 ± 2.57 11.5 ± 1.43

p < 0.05 NS NS

p < 0.001 p < 0.05 NS

p < 0.05 p < 0.05 p < 0.05

Analysis of variance Genotype (G) Si G × Si

Compared with Si0 treatment, supplementing Si did not significantly increase the rice root biomass. For example, the greatest growth rate of root biomass only was 12.75% for HT207 after Si20 treatment and 13.19% for HX9 after Si50 treatment. However, supplementing Si dramatically increased the straw biomass from the four rice genotypes. For example, the straw biomass increased by 40.10% for HT207 after Si20 treatment and 32.17% for IX17 after Si50 treatment compared with Si0 treatment, revealing the greatest growth rate. Moreover, with increasing supplementary Si, the straw biomass increased gradually for IX17 and IX12, although no significant increase was observed for IX12, but fluctuated for HX9 and HT207. Therefore, adding Si obviously increased the straw biomass, especially for HX9 and HT207. The grain biomass from the four rice genotypes also showed significant growth after Si addition (p < 0.05). Compared with Si0 treatment, the grain biomass increased by 68.75% for HX9 and 69.12% for IX12 after Si50 treatment, which was higher than that of other two genotypes. Moreover, the grain biomass of HX9, HT207, and IX12 after Si20 treatment increased by more than 50% compared with that after Si0 treatment. From Si10 to Si50 treatment, the grain biomass increased gradually for HX9, IX17, and IX12, while it fluctuated for HT207. In all, adding Si to the soil sample significantly increased the grain biomass (p < 0.05) of the four rice genotypes. Previous studies have found that rice is an Si-accumulating plant and that Si treatment has positive impacts on the health and yield of rice because it can negate the negative effects on rice growth from various biotic and abiotic stress factors, including heavy metals and toxic elements (Seyfferth and Fendorf, 2012). For example, previous studies have demonstrated that the biomass and shoot length of rice seedlings in hydroponic solutions containing As can be promoted by Si treatment (Guo et al., 2007). Adding Si also significantly increases the grain and straw yield of rice (Li et al., 2009a, b; Fleck et al., 2013). Moreover, supplementing Si can promote the growth and biomass production of maize exposed to Cd, Pb, As, Mn, and Al (Líang et al., 2007). Previous studies also indicated that Si treatment significantly increases not only the dry weight of shoot in hydroponic experiments (Guo et al., 2005) but also the grain and straw yield in pot experiments

(1%, 20 mL) at 95 °C for 2 h. The solution was cooled to 25 °C and centrifuged (5000 r/min, 10 min). Then, the supernatant was filtered through a 0.22 µm membrane filter. The concentrations of Sb were measured by high performance liquid chromatography coupled with hydride generation atomic fluorescence spectrometry (HPLC-HG-AFS) (HPLC, Shimadzu LC-15C Suzhou Instruments Co., China; HG-AFS, AFS8230, Beijing Haiguang Instruments Co., China). 2.6. Data analysis All data including rice biomass, Fe and Sb concentrations in different rice genotypes and Si treatments were subjected to two-way analysis of variance (ANOVA) by SPSS 19.0. 3. Results and discussion 3.1. Variations in rice growth The results of the root, straw, and grain biomass from the four genotypes that underwent the different Sb treatments are indicated in Table 1. It could be observed from the Si0 treatments that antimony treatment (50 mg/kg) without silicon had no obvious influence on the biomass of root, straw, or grain from the four rice genotypes, showing the negligible toxicity of Sb on rice growth at this concentration. Although Sb is a non-essential element for plants, it can be adsorbed and accumulated by rice from soil. A certain concentration of Sb in soil limits root growth and decreases the rice biomass (Feng et al., 2013). For example, the results observed from soil and pot cultivation experiments revealed that when the concentration of Sb (III) and Sb (V) in soil was in the range of 300–1000 µg/g, rice plants showed yellow leaves and sparse, short and brown roots, as well as reduced yields; thus, the growth was obviously inhibited (He and Yang, 1999). Significant differences in the biomass of root (p < 0.05), straw (p < 0.001), and grain (p < 0.001) were observed among the rice genotypes (Table 1). Compared with Si0 treatment, adding Si to the soil significantly enhanced the biomass of straw and grain (p < 0.05) but didn’t increased the root biomass for each rice genotype. 574

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L. Zhang et al.

Control

(a)

Si0

Si10

Si20

a'

a'

2000

A a' A'

1500

a*

1000

a' AB

A'

b a*

ab*

AB

60

Si50 Sb concentration (mg/kg)

Fe concentration (mg/kg)

2500

B B'

A'

b* b

b

a 500

Si0

(b)

Si10

Si20

Si50

a'b'

50

b' 40

a' a'

30

A

a

a

a

a A

A'

A

A A'

A'

A'

20 10 0

0 HX9

HT207

IX17

HX9

IX12

G enotype

HT207

IX17

IX12

G enotype

Fig. 1. Concentration variation of Fe (a) and Sb (b) in iron plaque on the surface of rice root after the addition of different amounts of silicon (mg/kg; data = mean ± SD; n = 3). Significant differences (p < 0.05) for the different rice genotypes in the same treatment are indicated by the different letters on the bars. The legend meaning is as follows: Control, no Si and no Sb addition; Si0, Sb only; Si10, Sb and 10 mg Si/kg; Si20, Sb and 20 mg Si/kg; Si50, Sb and 50 mg Si/kg.

degree of variation was different (Fig. 2). Silicon addition decreased root Sb concentrations of rice but not significantly except of Si50 treatment in HX9 (p < 0.05). With Si application amount increasing, root Sb concentrations in these four rice genotypes decreased and reached the lowest contents in Si50 treatment (Fig. 2a). For straw Sb contents, genotypes didn’t show significant impact (p < 0.05). However, application of Si decreased straw Sb contents in rice plants of four genotypes, significantly in IX17 and IX12 (p < 0.05) (Fig. 2b). In addition, significant genotypic differences were revealed (p < 0.01) and application of Si decreased husk Sb contents, significantly in rice genotypes IX17 and IX12 (p < 0.05) (Fig. 2c). Significant genotypic differences were observed for grain Sb contents (p < 0.001). Application of Si significantly decreased grain Sb contents in genotypes HX9, HT207, and IX12 (p < 0.05) but not significantly in IX17 (Fig. 2d). Previous studies have found that wetland plants with higher ROL contain relatively high As levels on their root surfaces (Li et al., 2011), and rice plants with higher ROL displayed a strong ability to reduce As content in shoots (Wu et al., 2011; Mei et al., 2012). The results obtained from this study showed that the Sb content in straw, husk, and grain from rice genotypes with higher ROL was lower than that from genotypes with lower ROL. However, the rice root with higher ROL possessed higher Sb content than that with lower ROL. Therefore, it can be concluded that Sb may be adsorbed on iron plaque due to the formation of iron plaque on the surface of the rice root, which may be enhanced in rice genotypes with higher ROL. This inference is consistent with the results of this study, that the Sb concentration in the iron plaque from rice genotypes with higher ROL (IX17 and IX12) is higher than that from the genotypes with lower ROL (HX9 and HT207) (Fig. 2), which could explain the observation in this study that higher total Sb concentrations in root were observed for genotypes with higher ROL rather than those with lower ROL. The predominant species in waterlogged soils is Sb (III) due to the absence of O2 (He, 2007), which can be taken up into the rice root through the Si transport channel (Huang et al., 2012). However, Si competes with Sb for uptake and translocation into the rice root. Therefore, Si addition impacted the uptake of Sb(III) into rice roots and, as a result, impacted the translocation of Sb(III) and decreased the total Sb content in root, straw, husk and grain. The analysis results of this study also revealed that Si addition decreased the Sb content in the straw, husk, and grain of rice with lower ROL more than in that with higher ROL. For example, the Sb concentration in straw decreased by 31.34% for HX9 and 10.38% for IX12 after Si10 treatment and decreased by 44.03% for HX9 and 22.64% for IX12 after Si40 treatment. Previous studies demonstrated that supplementing Si increases the barrier against ROL on the root surface of rice by contributing to a suberized exodermis and lignified sclerenchyma cells, which can inhibit the process of ROL (Kotula and Steudle, 2008; Fleck et al., 2011). Therefore, a barrier would develop on the root surface of the rice (Armstrong, 1979). This barrier against ROL is

(Li et al., 2009a), which is consistent with the results of this study. 3.2. Variations in iron plaque formation In all control samples, the Sb concentration in iron plaque was below the detection limit, and the Fe concentration was in the order of HX9 < HT207 < IX17 < IX12. However, adding Si significantly increased the Fe concentration in iron plaque (p < 0.01) (Fig. 1a). For example, compared with Si0 treatment, the Fe concentration in iron plaque for HX9 and IX12 increased by 86.42% and 69.77%, respectively, after Si10 treatment; by 128.86% and 71.76%, respectively, after Si20 treatment; and by 94.06% and 78.56%, respectively, after Si50 treatment. However, the Sb concentration in iron plaque initially increased and then decreased after Si application (Fig. 1b). In addition, the levels of Fe and Sb in iron plaque for indica genotypes with high ROL were obviously higher than those for hybrid genotypes with low ROL (Fig. 1). These results revealed that adding Si to soil influenced the sequestration of Sb on iron plaque for these four rice genotypes. Numerous studies have observed that iron plaque can form on the root surface of common aquatic plants. Certain factors, including pH, Eh, microorganisms, genotype, and ROL, can impact the formation of iron plaque (Emerson et al., 1999). Silicon can combine with iron plaque to form iron silicate and deposit on the root surface (Liu and Zhu, 2005). These conclusions are consistent with the results obtained in this study, that supplementing Si in soil significantly promoted the formation of iron plaque and increased the Fe concentration in iron plaque. In addition, iron plaque is considered to be a “barrier” to the uptake and accumulation of elements such as nitrogen, phosphorous, and heavy metals in rice (Liu et al., 2004; Huang et al., 2012). Oxides, mainly ferrihydrite and goethite, are the main components of iron plaque (Liu et al., 2004). Therefore, iron plaque on the root surface of rice can adsorb metals such as Sb and As on ferrihydrite, but silicon acid limits their adsorption by occupying the adsorption sites (Luxton et al., 2008), which may explain the decrease in Sb observed in DCB-extracts with increasing Si concentrations (Fig. 1b). 3.3. Variations in Sb in rice seedlings Antimony in different parts of the rice plants (including root, straw, husk, and grain) from the four rice genotypes was not detected in the control samples (Fig. 2). The results of this study indicated that the Sb concentration in the root of rice with lower ROL was lower than that in the root of rice with higher ROL, while the Sb concentration in the straw, husk, and grain of rice with lower ROL was higher than in that with higher ROL. For example, the total Sb concentration in the root of HX9 was lower 105 mg/kg than that of IX12, and the total Sb concentration in straw was 13.4 mg/kg for HX9 and 10.6 mg/kg for IX12. In addition, silicon treatment obviously decreased the Sb concentration in rice plants from the four rice genotypes (p < 0.05), although the 575

Ecotoxicology and Environmental Safety 144 (2017) 572–577

L. Zhang et al.

1000

(a)

A' A'

AB B AB

a'

A'

a

A

800

a

16

a' a'

a

Sb concentration (mg/kg dw)

Sb concentration (mg/kg dw)

1200

a'

A'

a

600 400 200 0

(b)

A

a

B'

14

b'

A

12

a a

A A

10 8 6 4 2

HT207

IX17

IX12

HX9

HT207

3

(c) a A

6

a a

A

(d)

B Sb concentration (mg/kg dw)

A

B'

a

b' A' A' A'

A

IX17

IX12

G enotype

G enotype

Sb concentration (mg/kg dw)

a' a' a'b'

0

HX9

9

a

A'B' A'B'A'

a'b' a'

3

a'

0

Si0

b A 2

A

A

ab

Si10

Si20

Si50

A' A'

ab

A'

a

A'

a'b'

b' a'

a'

1

0

HX9

HT207 IX17 G enotype

IX12

HX9

HT207

IX17

IX12

G enotype

Fig. 2. Total antimony (Sb) concentration in root (a), straw (b), husk (c), and grain (d) from the four rice genotypes (HX9, HT207, IX17, and IX12) cultivated under different silicon treatments (mg/kg; data = mean ± SD; n = 3). Significant differences (p < 0.05) for the same rice genotypes undergoing different silicon treatments are indicated by the letters on the bars. The legend meaning is as follows: Si0, Sb only; Si10, Sb and 10 mg Si/kg; Si20, Sb and 20 mg Si/kg; Si50, Sb and 50 mg Si/kg.

consistent with conclusions obtained in previous studies (Zhu et al., 2008a, b). In addition, the results of this study revealed that the inorganic Sb content decreased gradually with increasing supplementary Si (Table 2). For example, compared with Si0 treatment, the inorganic Sb concentration (sum of Sb(III) and Sb(V)) in the straw of HX9 decreased by 25.38%, 32.38%, and 38.49% after Si10, Si20, and Si50 treatment, respectively. The results can be explained by the fact that the translocation and uptake of Sb in rice seedlings can be inhibited by Si because of the competition between Si and Sb in the silicic acid transport system. A previous study also found that Si addition decreased inorganic As accumulation in rice grain (Li et al., 2009a). In addition, the inorganic Sb content in straw from rice genotypes with lower ROL was generally higher than that from higher ROL genotypes (Table 2). Moreover, compared with the Si0 sample, the inorganic Sb concentration in straw from rice genotypes with lower ROL decreased more than that from higher ROL genotypes, which is consistent with the variation in the total Sb concentration in straw. Therefore, the effect of Si on the total Sb concentration in rice genotypes with different ROL can be explained by the impact of Si on the uptake of inorganic Sb, especially that of Sb(III).

present in basal parts of the root (Colmer, 2003). In general, suberin deposition, densely packed cells, and lignification in the outer cell layers provide important contributions to barrier formation (Sorrell, 1994), while the barrier against ROL is attributed to a suberized exodermis with casparian bands and lignified sclerenchyma cells (Kotula and Steudle, 2008). However, silicon acid can enhance the formation of casparian bands in the exodermis and endodermis (Fleck et al., 2011; Vaculikova et al., 2014, 2016). Therefore, supplementing Si can weaken the ROL and the effects of ROL on Sb uptake in rice, and higher ROL genotypes may be weakened more. Thus, the Sb concentration in rice plants from lower ROL genotypes decreased more than that from higher ROL genotypes after Si addition. 3.4. Variations in Sb speciation in rice straw In this study, Sb species, including Sb(III), Sb(V), and trimethylantimony (TMSb), were detected in the straw of two rice genotypes, HX9 and IX12, and the analysis results are shown in Table 2. The percentage of inorganic Sb, Sb(III) and Sb(V), was in the range of 89.42–96.71% and was the predominant species. This result is

Table 2 Concentration of antimony species in straw from two rice genotypes (HX9 and IX12) cultivated under different silicon treatments (mg/kg, data = mean ± SD). Genotypes

Treatment

Total Sb (mg/kg)

Inorganic Sb (mg/kg)

TMSb (mg/kg)

Inorganic Sba (%)

Recoveryb (%)

HX9

Si0 Si10 Si20 Si50 Si0 Si10 Si20 Si50

13.4 ± 1.25 10.3 ± 1.68 9.5 ± 1.36 8.9 ± 0.86 10.6 ± 2.15 9.2 ± 1.24 8.7 ± 2.15 8.1 ± 2.43

11.49 ± 3.15 10.04 ± 1.06 8.41 ± 2.45 7.87 ± 0.51 10.72 ± 1.32 8.86 ± 0.52 6.63 ± 0.98 6.43 ± 1.45

0.44 ± 0.09 0.57 ± 0.11 0.71 ± 0.15 0.94 ± 0.19 0.41 ± 0.06 0.52 ± 0.08 0.59 ± 0.07 0.62 ± 0.13

96.71 94.42 92.49 89.42 96.12 94.31 93.19 92.30

89 103 96 99 105 102 83 87

IX12

a b

Inorganic Sb (%) = [(inorganic Sb)/(species sum)] ×100. Recovery (%) = [(species sum)/(total Sb)] ×100.

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Ecotoxicology and Environmental Safety 144 (2017) 572–577

L. Zhang et al.

period of germination and growth and antimony concentration in rice tissue. Sci. Total Environ. 243, 149–155. He, M.C., 2007. Distribution and phytoavailability of antimony at an antimony mining and smelting area, Hunan, China. Environ. Geochem. Health 29, 209–219. Huang, Y.Z., Zhang, W.Q., Zhao, L.J., 2012. Silicon enhances resistance to antimony toxicity in the low-silica rice mutant lsi1. Chem. Ecol. 28, 341–354. Kotula, L., Steudle, E., 2008. Measurements of oxygen permeability coefficients of rice (Oryza sativa L.) roots using a new perfusion technique. J. Exp. Bot. 60, 567–580. Li, R.Y., Ago, Y., Liu, W.J., Mitani, N., Feldmann, J., McGrath, S.P., Ma, J.F., Zhao, F.J., 2009a. The rice aquaporin lsi1 mediates uptake of methylated arsenic species. Plant Physiol. 150, 2071–2080. Li, R.Y., Stroud, J.L., Ma, J.F., McGrath, S.P., Zhao, F.J., 2009b. Mitigation of arsenic accumulation in rice with water management and silicon fertilization. Environ. Sci. Technol. 43, 3778–3783. Li, H., Ye, Z.H., Wei, Z.J., Wong, M.H., 2011. Root porosity and radial oxygen loss related to arsenic tolerance and uptake in wetland plants. Environ. Pollut. 159, 30–37. Líang, Y., Sun, W., Zhu, Y.G., Christie, P., 2007. Mechanisms of silicon-mediated alleviation of abiotic stresses in higher plants: a review. Environ. Pollut. 147, 422–428. Liu, W.J., Zhu, Y.G., Smith, F.A., Smith, S.E., 2004. Do phosphorus nutrition and iron plaque alter arsenate (As) uptake by rice seedlings in hydroponic culture? New Phytol. 162, 481–488. Liu, W.J., Zhu, Y.G., 2005. Iron and Mn plaques on the surface of roots of wetland plants. Acta Ecol. Sin. 25, 358–363. Liu, C.P., Wei, L., Zhang, S.R., Xu, X.H., Li, F.B., 2014. Effects of nanoscale silica sol foliar application on arsenic uptake, distribution and oxidative damage defense in rice (Oryza sativa L.) under arsenic stress. RSC Adv. 4, 57227. Luxton, T.P., Eick, M.J., Rimstidt, D.J., 2008. The role of silicate in the adsorption/desorption of arsenite on goethite. Chem. Geol. 252, 125–135. Ma, J.F., Yamaji, N., 2006. Silicon uptake and accumulation in higher plants. Trends Plant Sci. 11, 392–397. McDonald, M.P., Galwey, N.W., Colmer, T.D., 2001. Waterlogging tolerance in the tribe Triticeae: the adventitious roots of Cristesion marinum have a relatively high porosity and a barrier to radial oxygen loss. Plant Cell Environ. 24, 585–596. Mei, X.Q., Wong, M.H., Yang, Y., Dong, H.Y., Qiu, R.L., Ye, Z.H., 2012. The effects of radial oxygen loss on arsenic tolerance and uptake in rice and on its rhizosphere. Environ. Pollut. 165, 109–117. Qi, C.C., Wu, F.C., Deng, Q.J., Liu, G.J., Mo, C.L., Liu, B.J., Zhu, J., 2011. Distribution and accumulation of antimony in plants in the super-large Sb deposit areas, China. Microchem. J. 97, 44–51. Rauret, G., López-Sánchez, J.F., Sahuquillo, A., Rubio, R., Davidson, C., Ure, A., 1999. Improvement of the BCR three-step sequential extraction procedure prior to the certification of new sediment and soil reference materials. J. Environ. Monit. 1, 57–61. Seyfferth, A.L., Fendorf, S., 2012. Silicate mineral impacts on the uptake and storage of arsenic and plant nutrients in rice (Oryza sativa L.). Environ. Sci. Technol. 46, 13176–13183. Sorrell, B.K., 1994. Airspace structure and mathematical modeling of oxygen diffusion, aeration and anoxia in Eleocharis sphacelata R. Br. roots. Aust. J. Mar. Freshw. Res. 45, 1529–1541. Ultra Jr., V.U., Nakayama, A., Tanaka, S., Kang, Y., Sakurai, K., Iwasaki, K., 2009. Potential for the alleviation of arsenic toxicity in paddy rice using amorphous iron(hydr) oxide amendments. Soil. Sci. Plant. Nutr. 55, 160–169. Vaculikova, M., Vaculik, M., Simkova, L., Fialova, I., Kochanova, Z., Sedlakova, B., Luxova, M., 2014. Influence of silicon on maize roots exposed to antimony-Growth and antioxidative response. Plant Physiol. Biochem. 83, 279–284. Vaculikova, M., Vaculik, M., Tandy, S., Luxova, M., Schulin, R., 2016. Alleviation of antimonate (SbV) toxicity in maize by silicon (Si). Environ. Exp. Bot. 128, 11–17. Vatehová, Z., Kollárová, K., Zelko, I., Richterová-Kučerová, D., Bujdoš, M., Lišková, D., 2012. Interaction of silicon and cadmium in Brassica juncea and Brassica napus. Biologia 67, 498–504. Vaculík, M., Jurkovič, Ĺ., Matejkovič, P., Molnárová, M., Lux, A., 2013. Potential risk of arsenic and antimony accumulation by medicinal plants naturally growing on old mining sites. Water Air Soil Pollut. 224, 1–16. Wilson, S.C., Lockwood, P.V., Ashley, P.M., Tighe, M., 2010. The chemistry and behaviour of antimony in the soil environment with comparisons to arsenic: a critical review. Environ. Pollut. 158, 1169–1181. Wu, C., Ye, Z.H., Shu, W.S., Zhu, Y.G., Wong, M.H., 2011. Arsenic accumulation and speciation in rice are affected by root aeration and variation of genotypes. J. Exp. Bot. 62, 2889–2898. Wu, C., Ye, Z., Li, H., Wu, S., Deng, D., Zhu, Y., Wong, M., 2012. Do radial oxygen loss and external aeration affect iron plaque formation and arsenic accumulation and speciation in rice? J. Exp. Bot. 63, 2961–2970. Wu, C., Zou, Q., Xue, S., Mo, J., Pan, W., Pan, W., Lou, L., Wong, M.H., 2015. Effects of silicon (Si) on arsenic (As) accumulation and speciation in rice (Oryza sativa L.) genotypes with different radial oxygen loss (ROL). Chemosphere 138, 447–453. Zhao, F.J., Zhu, Y.G., Meharg, A.A., 2013. Methylated arsenic species in rice: geographical variation, origin, and uptake mechanisms. Environ. Sci. Technol. 47, 3957–3966. Zhu, Y.G., Williams, P.N., Meharg, A.A., 2008a. Exposure to inorganic arsenic from rice: a global health issue? Environ. Pollut. 154, 169–171. Zhu, Y.G., Sun, G.X., Lei, M., Teng, M., Liu, Y.X., Chen, N.C., Wang, L.H., Carey, A.M., Deacon, C., Raab, A., Meharg, A.A., Williams, P.N., 2008b. High percentage inorganic arsenic content of mining impacted and nonimpacted Chinese rice. Environ. Sci. Technol. 42, 5008–5013.

However, with increasing supplementary Si, the concentration of TMSb also weakly increased in straw from these two rice genotypes. Some studies have found that adding Si can measurably increase the dimethylarsinate (DMA) concentration in rice grain (Li et al., 2009a). To date, no study has reported that Sb can be methylated in rice plants. Studies have confirmed that rice plants cannot methylate As, and DMA is mainly derived from methylation by microorganisms in soil (Zhao et al., 2013). Therefore, TMSb may be derived from the rice rhizosphere in soil. It can be concluded from the results of this study that the uptake of TMSb into rice root may increase upon Si addition. When increasing the amount of Si, the TMSb content in the straw indeed increased gradually, which verifies our above inference. The mechanism of Sb methylation and its uptake by rice plants will be systematically investigated in future research. 4. Conclusions The biomass of root, straw, husk, and grain from different rice genotypes increased upon Si addition. Indica genotypes with higher ROL produced more iron plaque than hybrid genotypes and sequestered greater amounts of Sb. Adding Si to soil measurably enhanced the formation of iron plaque and impacted the Fe and Sb levels in iron plaque from different rice genotypes. Antimony levels in the root, straw, husk, and grain of hybrid genotypes were higher than those of indica genotypes. Moreover, Si addition reduced the Sb concentration in rice plants. In addition, inorganic antimony accounted for more than 85% of the total Sb content. Hybrid genotypes accumulated higher levels of inorganic Sb than indica genotypes. Supplementing Si decreased the inorganic Sb content in the rice straw of hybrid and indica genotypes. The results of this study provide a foundation for understanding Sb uptake mechanisms of rice and mitigating the risks of Sb pollution in paddy fields. Acknowledgements This study is supported by the Project of National Natural Science Foundation of Youth Science Foundation (No. 41301532), the Encouraging Foundation for Outstanding Youth Scientists of Shandong Province (No. BS2011SW012), and the Qufu Normal University National Undergraduate Innovation and Entrepreneurship Training Program (No. 201610446088). References Armstrong, W., 1979. Aeration in higher plants. In: Woolhouse, H.W., (Ed.), Academic Press, London, pp. 225–332. Baroni, F., Boscagli, A., Protano, G., Riccobono, F., 2000. Antimony accumulation in Achillea ageratum, Plantago lanceolata and Silene vulgaris growing in an old Sb-mining area. Environ. Pollut. 109, 347–352. Colmer, T.D., 2003. Aerenchyma and an inducible barrier to radial oxygen loss facilitate root aeration in upland, paddy and deep-water rice (Oryza sativa L.). Ann. Bot. 91, 301–309. Emerson, D., Weiss, J.V., Megonigal, J.P., 1999. Iron-oxidizing bacteria are associated with ferric hydroxide precipitates (Fe-plaque) on the roots of wetland plants. Appl. Environ. Microb. 65, 2758–2761. Feng, R.W., Wei, C.Y., Tu, S.X., Ding, Y.Z., Wang, R.G., Guo, J.K., 2013. The uptake and detoxification of antimony by plants: a review. Environ. Exp. Bot. 96, 28–34. Filella, M., Belzile, N., Chen, Y.W., 2002. Antimony in the environment: a review focused on natural waters I. occurrence. Earth Sci. Rev. 57, 125–176. Fleck, A.T., Nye, T., Repenning, C., Stahl, F., Zahn, M., Schenk, M.K., 2011. Silicon enhances suberization and lignification in roots of rice (Oryza sativa). J. Exp. Bot. 62, 2001–2011. Fleck, A.T., Mattusch, J., Schenk, M.K., 2013. Silicon decreases the arsenic level in rice grain by limiting arsenite transport. J. Plant Nutr. Soil Sci. 176, 785–794. Guo, W., Hou, Y.L., Wang, S.G., Zhu, Y.G., 2005. Effect of silicate on the growth and arsenate uptake by rice (Oryza sativa L.) seedlings in solution culture. Plant Soil 272, 173–181. Guo, W., Zhu, Y.G., Liu, W.J., Liang, Y.C., Geng, C.N., Wang, S.G., 2007. Is the effect of silicon on rice uptake of arsenate (AsV) related to internal silicon concentrations, iron plaque and phosphate nutrition? Environ. Pollut. 148, 251–257. He, M.C., Yang, J.R., 1999. Effects of different forms of antimony on rice during the

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