Elsevier

Chemosphere

Volume 175, May 2017, Pages 275-285
Chemosphere

Mechanisms of Fe biofortification and mitigation of Cd accumulation in rice (Oryza sativa L.) grown hydroponically with Fe chelate fertilization

https://doi.org/10.1016/j.chemosphere.2017.02.053Get rights and content

Highlights

  • Compared to EDDHAFe(III), EDTANa2Fe(II) exhibit better mitigation of Cd accumulation in rice.

  • EDTANa2Fe(II) generate EDTANa2Cd(II) in solution and decrease net Cd influx rate in root to reduce Cd uptake in rice.

  • Addition of Fe chelates reduce Cd transport in rice by inhibiting gene expressions of Fe- and Cd-related xylem and phloem.

Abstract

Cadmium contaminated rice from China has become a global food safety issue. Some research has suggested that chelate addition to substrates can affect metal speciation and plant metal content. We investigated the mitigation of Cd accumulation in hydroponically-grown rice supplied with EDTANa2Fe(II) or EDDHAFe(III).

A japonica rice variety (Nipponbare) was grown in modified Kimura B solution containing three concentrations (0, 10, 100 μΜ) of the iron chelates EDTANa2Fe(II) or EDDHAFe(III) and 1 μΜ Cd. Metal speciation in solution was simulated by Geochem-EZ; growth and photosynthetic efficiency of rice were evaluated, and accumulation of Cd and Fe in plant parts was determined. Net Cd fluxes in the meristematic zone, growth zone, and maturation zone of roots were monitored by a non-invasive micro-test technology. Expression of Fe- and Cd-related genes in Fe-sufficient or Fe-deficient roots and leaves were studied by QRT-PCR.

Compared to Fe deficiency, a sufficient or excess supply of Fe chelates significantly enhanced rice growth by elevating photosynthetic efficiency. Both Fe chelates increased the Fe content and decreased the Cd content of rice organs, except for the Cd content of roots treated with excess EDDHAFe(III). Compared to EDDHAFe(III), EDTANa2Fe(II) exhibited better mitigation of Cd accumulation in rice by generating the EDTANa2Cd complex in solution, decreasing net Cd influx and increasing net Cd efflux in root micro-zones. Application of EDTANa2Fe(II) and EDDHAFe(III) also reduced Cd accumulation in rice by inhibiting expression of genes involved in transport of Fe and Cd in the xylem and phloem.

The ‘win-win’ situation of Fe biofortification and Cd mitigation in rice was achieved by application of Fe chelates. Root-to-stem xylem transport of Cd and redistribution of Cd in leaves by phloem transport can be regulated in rice through the use of Fe chelates that influence Fe availability and Fe-related gene expression. Fe fertilization decreased Cd influx and increased Cd efflux in rice roots.

Introduction

Rice is a staple food for more than 1.3 billion Chinese people, so it is vital to utilize paddy soils safely and sustainably in China. However, in recent years, incidents of Cd contamination of rice have occurred frequently in China's major rice grain production areas. The causes of this contamination have included paddy irrigation with sewage in Shenyang city, irrigation with Cd-contaminated river water in Hunan Province, and industrial waste contamination of so-called ‘cancer villages’ in Guangdong Province. The primary source of human Cd intake in Asia is consumption of rice containing Cd (Arao et al., 2009, Cheng et al., 2014, Zhao et al., 2015). As China is a major exporter of rice to other Asian countries, the widespread distribution and consumption of Cd-contaminated rice poses a potential health hazard throughout the region. Hence, the application of rice Cd mitigation technologies has been a focus of research both in China and abroad (Larson, 2014). Much research has shown that rice Cd uptake and transportation processes are primarily affected by external environmental factors such as soil pH, Eh and organic matter (Kashem and Singh, 2001, Liu et al., 2013, Honma et al., 2016). Since much of this research has employed hydroponic culture, it is important to consider how the differences between soil culture and hydroponic culture affect Cd uptake and transportation in the rice plant. In particular, to prevent precipitation of ionic Fe salts in hydroponic solution, which would affect Fe phytoavailability, researchers often use more soluble Fe chelates such as EDTA-Fe and EDDHA-Fe instead of ionic Fe salts. Such chelators also affect the speciation and phytoavailability of other metals (e.g., Cd, Zn, and Cu). There are many contradictory results in the literature on the effect of chelators on metal phytoavailability. Some studies have found that dissociation of metal-chelate complexes can greatly promote uptake of metal ions into plant roots (Chaney et al., 1972, Degryse et al., 2006, Degryse et al., 2012, Panfili et al., 2009, Wang et al., 2009). In contrast, other studies have shown that application of chelates can reduce metal uptake by plant roots and mitigate accumulation of toxic metals in rice (Lin et al., 2014, Custos et al., 2014). Additionally, some crop plants (Phaseolus vulgaris, Brassica juncea, Hordeum vulgare and Solanum tuberosum) can directly absorb chelate-metal complexes (e.g., EDTA-Pb, -Zn, -Cd, and -Fe), which have been identified in xylem sap in studies utilizing hydroponic or soil culture (Sarret et al., 2001, Collins et al., 2002, Schaider et al., 2006). It has been proposed that plant uptake of these metal complexes occurs passively (Nowack et al., 2006). We hypothesize that addition of synthetic chelates to a substrate will result in ligand substitution reactions with natural metal compounds, producing metal complexes that are more stable and less phytoavailable than are the natural metal compounds. Plant roots have no known transport systems for these synthetic metal complexes as they do for natural metal-phytosiderophore complexes (Von Wirén et al., 1996). Thus, further research on how plants are affected by these synthetic metal complexes is needed.

Mitigation mechanisms based on amendment of soil are generally focused on affecting external environmental factors, such as increasing soil pH to reduce the availability of heavy metals in substrates or enhance metal compartmentalization in the plant. Examples of this approach include application of lime, silicon fertilizer, fly ash, steel slag, red mud, or iron oxides to reduce Cd accumulation in rice (Arao et al., 2010, Gu et al., 2011, Gu et al., 2012, Suda and Makino, 2016). Soil application of some amendments may result in secondary environmental pollution. The 10-Chapter Soil Pollution Prevention Action Plan, whose goal is to assure security of agricultural production, prohibits the direct application to soil of urban wastes, sewage sludge, and industrial waste as fertilizers (State Council, 2016). Thus, amendments made of the above waste materials cannot be applied to soil. Nevertheless, the use of functional metal-antagonist fertilizers is likely to become increasingly important in the future. EDTANa2Fe(II) and EDDHAFe(III) are two common and efficient Fe chelate fertilizers that have been widely used for Fe biofortification. However, there has been little research on the efficiency of Fe chelate fertilization for Cd mitigation in rice or on the mechanisms involved. The physiological mechanisms of uptake of Cd by rice roots and its root-to-shoot translocation are associated with several chemically related metal ions, most notably Fe ions (Uraguchi and Fujiwara, 2012). There is molecular evidence that Cd can be transported into rice via Fe metabolic systems that are affected by Fe status in substrates (Ishimaru et al., 2006, Nakanishi et al., 2006; Takahashi et al., 2011; Sasaki et al., 2012). It has been reported that certain rice genes are responsible for Fe and Cd xylem loading and phloem redistribution between rice plant parts. For instance, absorption of Cd from the rhizosphere into root cells is mediated by OsIRT1/2 and OsNRAMP5 (Nakanishi et al., 2006, Sasaki et al., 2012), and OsHMA3 plays a critical role in Cd compartmentalization into vacuoles in root cells (Miyadate et al., 2011, Sasaki et al., 2014). Takahashi et al. (2012) reported that OsHMA2 increased Cd xylem loading in roots for translocation to shoots but decreased Cd phloem loading for storage in the grain sink. In addition, OsLCT1 and OsHMA2 mediate xylem-to-phloem transfer at nodes (Uraguchi et al., 2011, Takahashi et al., 2012). However, the influence of Fe chelates on regulation of Fe- and Cd-related genes is unclear and is worthy of investigation.

In the work presented here, the Fe chelates EDTANa2Fe(II) and EDDHAFe(III) were added to Cd-contaminated hydroponic systems used for growing rice seedlings, with the goal of evaluating how fertilization with Fe chelates affects mitigation of Cd accumulation in rice. Although indirect evidence has shown that Fe plays a role in reducing Cd toxicity in the rhizosphere of rice, direct physiological evidence of Fe-induced Cd tolerance at the level of root micro-zones (the meristematic, growth, and maturation zones) remains very limited. Thus, a non-invasive micro-test technology (NMT) was used for real-time monitoring of net Cd influx and efflux in root micro-zones. In addition, we discuss the absorption pathways of Fe chelates in rice roots in combination with Fe- and Cd-related gene expression.

Section snippets

Plant materials

A low Cd-accumulating japonica cultivar (Nipponbare) was used for the hydroponic experiment. Rice seeds were germinated in a plastic box (102 × 102 × 5 cm) divided into 100 compartments (1 × 1 × 5 cm), with two seeds per compartment. Sufficient 0.5 mM CaCl2 solution was added to maintain a flooded layer 3 cm deep, and the box was placed in darkness to accelerate germination. After 10 days of germination, the seedlings were separated into two groups and transplanted to 1-L plastic hydroponic

Speciation of Cd and Fe in hydroponic solutions

Geochem-EZ software was used to simulate and calculate the distribution of Cd species in hydroponic solutions containing a range of Fe chelate concentrations (0 μM–100 μM). Simulation results for EDDHAFe(III) showed that regardless of the chelate concentration, dissolved Cd maintained the same speciation ratio as in the no-Fe condition (0 μM). Free Cd ion (91.51%) was the major Cd species in these solutions, followed by inorganic complexes of Cd with SO4[2-] (8.22%), PO4[3-] (0.17%), NO3[-]

Discussion

The hypothesis of this study is based on the observation that in the hydroponic environment, the addition of chelates can affect the chemical speciation of Fe and Cd in the nutrient solution, which in turn affect the bioavailability of these metals to plants. However, there have been conflicting reports of how chelates affect the phytoavailability of heavy metals. Some research has shown that the introduction of chelating agents promotes the absorption and transport of metals in plants, whilst

Conclusion

The physiological mechanisms of uptake of Cd by rice roots and its root-to-shoot translocation are associated with several chemically related metal ions, most notably Fe ions. Fertilization of hydroponically grown rice with synthetic Fe chelates affected not only the speciation of Cd and Fe in solution but also the uptake, transport and redistribution of Cd and Fe in the rice plant, as well as the expression of rice genes involved in transport of Fe and Cd. Our results demonstrated that the

Acknowledgments

We thank Shao Guosheng for providing EDTANa2Fe(II); we appreciate for Dai Yangshuo and Wang Fengzhu for their help in QRT-PCR measurements and NMT test. This research was financially supported by the National Natural Science Foundation of China (No. 41225004), the Guangdong Provincial Natural Science Foundation of China (No. 2014A030313200), National Key Technology R&D Program of the Ministry of Science and Technology of China (No. 2015BAD05B05), Science and Technology Program of Guangzhou,

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