Giant Rice Is a Unique Candidate for the Sustainable Phytoremediation of Cadmium-Contaminated Paddy Fields

Abstract

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The limited availability of ideal cadmium (Cd) hyperaccumulating plants remains a significant challenge for phytoremediating Cd-contaminated soils. This study aims to find out if the high biomass giant rice can be a suitable phytoremediation candidate for Cd-contaminated paddy fields and to elucidate physiological mechanisms relative to low or high Cd accumulation. Soil column and lysimeter tests were conducted with giant and normal species of rice under different water conditions. Cd distribution in different parts of root and shoots were determined. Under sustained flooding in column tests or keeping soil moist after the tillering stage in lysimeter tests, Cd contents in the grain of giant rice were very low. The Cd bioconcentration factor (BCF) of grain of giant rice (0.015–0.034) were significantly lower than those of normal rice (0.088–0.099), while BCF of root was in the reversed trend. Of the Cd in giant rice roots, 23.5% was trapped by the iron plaque on the root surface, compared 11.7% for normal rice. More Fe–O, –COOH functional groups in giant rice roots were confirmed by FTIR spectra. Under semi-dry condition, Cd contents in giant rice were very significantly increased, BCF straw in regenerated giant rice reached 13.4. Cd in shoots were accumulated in basal stems of the giant rice, the phytoextraction rate reached 18.7% of total Cd in topsoil, compared to 9.6% for giant Napier grass (Pennisetum hybridum). Therefore, the giant rice is a unique crop who can highly produce safe rice under wet conditions in the first half year and meanwhile by regenerating it can efficiently phytoextract Cd in the soil under semi-dry condition with a low labor input and carbon emission.

Introduction

Croplands affected by toxic metals pollutions are estimated to be 14–17% globally and between 0.9 and 1.4 billion people live in regions of heightened public health and ecological risks (Hou et al. 2025). Among toxic metals, increased cadmium (Cd) levels in arable soil have caused widespread concern about Cd pollution in foods (Jin et al. 2021; McLaughlin et al. 2021). Rice (Oryza sativa L.), which is the primary food crop for over half the world’s population, is particularly important to human Cd exposure (Kong et al. 2023). Unfortunately, paddy soils may present a higher Cd bioavailability (27% DTPA-extractable of the total Cd in soil) than upland soils (18%) (Liu et al. 2023). Up to 10% of marketable rice exceeds Cd standard limit in China (Chen et al. 2018). The predicted Cd exceedance rates in the major rice export countries (India, Pakistan and Thailand) are even higher (10−60%) (Hou et al. 2025). Rice is the primary source of dietary Cd intake in Asian countries, for example contributing an average of 56% to total dietary Cd intake in China (Song et al. 2017). Cd in rice grain has relatively high bioavailability within the gastrointestinal tract (Gu et al. 2020), resulting in various adverse health effects (Huang et al. 2019). Therefore, global agricultural soil pollution by heavy metal(loid)s represents one of the biggest challenges to sustainable development, particularly in developing countries (Hou et al. 2020).

Changes in water conditions can alter physical and chemical properties of soil, affecting the soil Cd bioavailability (Yan et al. 2021). Under flooding, Cd in soil exists in insoluble forms (such as CdS), reducing its availability to plants. On the contrary, in the soil under the oxidation condition, Cd exists as soluble forms, increasing its availability to plants. Draining water at maximum tillering stage could promote Cd accumulation in rice shoot (Ibaraki et al. 2009).

There are significant differences in the uptake and accumulation of soil Cd among different rice varieties (Wu et al. 1999; Wang et al. 2021). Phyto-exclusion is a strategy to reduce plant Cd uptake and transport (Wang et al. 2021). Studies have shown that low-Cd rice effectively reduced the uptake and accumulation of Cd (Guo et al. 2019; Sun et al. 2025). However, they are only valid under prolonged irrigations in the fields with inter-mediate levels of Cd pollution (Duan et al. 2017).

Amendments often decreases soil Cd bioavailability and Cd accumulation in rice plants (Ge et al. 2024), however they may not enough to harvest rice grain conforming to food standards in some acid soils in south China (Liang et al. 2022). This is why Deng et al. (2025) pre-excluded rice in the screening of practical low-accumulating crops in Guangdong Province, China. To removal heavy metals from soil, phytoextraction is an important green technique for the clean-up of contaminated soils (Huang et al. 2020a; Rylott and Bruce 2011). However, the practical phytoextraction with non-crop hyperaccumulating plants is still underway due to the low biomass produced or/and the lack of appropriate planting and harvesting machines (Xia et al. 2025). Removing rice straw with common rice varieties has little effect on reducing Cd in the soil (Zhao et al. 2021). High-Cd accumulating rice has been used for the remediation of low to moderate level Cd-contaminated paddy fields (Takahashi et al. 2021), such as cultivar Lu527-8 (Tang et al. 2016), Akita 119 (Takahashi et al. 2020). However, the operation cost is still too high compared to the application of obtained Cd phytoextraction rates without agricultural income for local farmers.

The Giant rice was developed by the Institute of Subtropical Agriculture, Chinese Academy of Sciences (Qu et al. 2020). It has been authenticated as a novel rice germplasm after rice gene chip fingerprint detection by the Ministry of Agriculture of China (Tang 2022). Giant rice stands tall with long ears and abundant grains, boasting an average height of 1.8 m (highest to 2.2 m), more than 150% biomass than traditional rice varieties. The most distinctive feature is its high yield (12000–15000 kg/ha), together with the advantages of tall stalks, thick stems, lush leaves, and robust regeneration ability (Tang 2022). Giant rice is frequently used in polyculture with aquatic animals (Ding 2019; Liang et al. 2020). These polycultures may facilitate to keep sustained flooding, minimize the uptake and accumulation of Cd by rice. Therefore, giant rice is expected to be a good material for Cd contaminated soil via phyto-exclusion, even phytoextraction with its high biomass. However, current research on giant rice primarily involves cultivation techniques (Ding 2019; Luo et al. 2021) and nutrient utilization (Liu et al. 2021a), but no published report on its Cd accumulation characteristics.

Because early drainage of paddy field or semi-dry condition can significantly increase the level of Cd in rice, and giant rice has rather high shoot biomass, combining these two factors may realize effective phytoextraction. Therefore, Cd accumulation characteristics under different water conditions of giant rice were studied by soil column and lysimeter tests. Based on the results of the first year, we proposed a seasonal combination called “flooded growth of early rice and drained regrowth of late rice” with giant rice during a year to verify it’s low grain Cd characteristic of early rice under sustained flooding and efficient Cd phytoextraction by late rice under semi-dry condition, and also to elucidate which barrier mechanism in roots involved in high Cd retention and which parts of rice stem and leave to accumulate Cd when giant rice accumulates high amount of Cd under semi-dry condition.

Results

The Growth and Yield of Rice

Table 1 presents the growth status of rice at the mature stage in the first soil column experiment. Under identical water condition, giant rice exhibited significantly superior growth in terms of root length, plant height, and biomass compared to Simiao rice. The biomass of grain, straw, root in topsoil and root in subsoil of giant rice were 1.24, 1.50, 1.82 and 3.25 times of that for Simiao rice, respectively. The longest single root length of giant rice exceeded 70 cm, which was 1.36 times compared to that of Simiao rice, suggesting that giant rice had well-developed roots and increase C fixation in soil, in particular in subsoil. There was no significant difference in the growth indexes of giant rice under two water conditions. This indicated that drying after the tillering do not impact the normal growth of giant rice.

Table 1 Plant biomass of giant and Simiao (normal) rice at the maturity stage in column test-1

The lysimeter test provided a more favorable condition for giant rice growth, with a measure of 187.3 cm for early rice and 195.5 cm for late rice at maturity (Fig. 1). The average height of giant rice was 1.59 times of that for Simiao rice, The dry weights per-hectare of giant rice grain and straw for a single crop were 4428–7200 kg and 9148–11,452 kg, significantly higher than those of Simiao rice cultivated under the same conditions. The growth parameters of giant rice also demonstrated superior in late rice compared to early rice, suggesting that giant rice was suitable to be cultivated in late crop for subtropical regions.

Fig. 1

Plant growth and yield of giant and Simiao (normal) rice at the maturity stage in lysimeter test for early (spring-summer) and late (summer-autumn) seasons. (Different letters indicate significant difference (P < 0.05) according to Duncan’s test.)

Cd Concentration in Rice

The Cd concentration in each part of matured rice is shown in Table 2. In the first soil column experiment under sustained flooding, the Cd concentrations in giant rice and Simiao rice grains were 0.0079 and 0.0105 mg·kg⁻¹, respectively, which were significantly lower than the Cd limit (0.2 mg·kg⁻¹) stipulated in the Chinese food safety standard (GB 2762−2022). There was no significant difference in Cd concentrations between these two types of rice. Drained after the tillering stage, the Cd concentrations in giant and Simiao rice grain were significantly increased compared to those after sustained flooding, reached 0.5556 and 0.5386 mg·kg⁻¹, respectively, and exceeded the Cd standard limit for rice. The Cd concentrations in straw and roots in topsoil and subsoil of giant rice were significantly higher than those of Simiao rice, indicating that more Cd was intercepted by the roots and straw of giant rice.

In lysimeter test, taking early rice as an example, the Cd concentrations in giant rice and Simiao rice grains were found to be 0.0175 and 0.1126 mg·kg⁻¹, indicating a low accumulation of Cd. This phenomenon was consistently observed in late rice as well. On the whole, giant rice exhibited superior characteristics of low Cd accumulation in grain compared to Simiao rice. The Cd concentrations in grain and straw of giant rice were significantly lower than those of Simiao rice, while the Cd concentration in root showed an inverse trend. This implied that the Cd retention capacity of giant rice roots was stronger than that of Simiao rice.

Table 2 Concentrations of cd in different parts of giant and normal (Simiao) rice (mg·kg⁻¹ dry weight)

Bioconcentration and Translocation of Cd in Rice

The Bio-concentration factors (BCF) and translocation factors (TF) of Cd in different parts of giant rice and Simiao rice are presented in Table 3. In soil column test, under sustained flooding, there was no significant difference in BCF between two types of rice, with the BCF of grains being less than 0.01. Under semi-dry condition, the BCFs of different parts of rice significantly increased. Furthermore, the BCF_Root and BCF_Straw of giant rice were significantly higher than those of Simiao rice, indicating a stronger ability to enrich Cd. Under two irrigation conditions in soil column test, the TF_Root−Grain of giant rice was less than that of Simiao rice, suggesting that Cd is less likely to be transferred to grains in giant rice. Under semi-dry condition, TF_{Straw−Grain} of giant rice was also significantly lower than that of Simiao rice, indicating a stronger retention ability for Cd in the root system and straw of giant rice compared to that of Simiao rice. The proportion of Cd translocated in grains were 1.67–4.13% for giant rice under sustained flooding and semi-dry condition, while they were 2.94–12.22% for Simiao rice (Fig. S1).

In the early rice of lysimeter test, the BCF_Grain and BCF_Straw of giant rice were significantly lower, while BCF_Root was significantly higher than those of Simiao rice (Table 3). Similar results were obtained for late rice. Both TF_Root−Grain and TF_Root−Straw of giant rice were significantly lower than those of Simiao rice (early and late rice), while there was no significant difference in TF_{Straw−grain}, indicating that the root fixation ability of giant rice played a key role.

Cd Phytoextraction Ability of Rice

In column test-1, under sustained flooding, the amount of Cd extracted by giant rice was low, only 26.88 µg·pot⁻¹, and the phytoextraction rate of shoots and roots were 0.218% and 0.261%, respectively (Table 4). The extraction rate of giant rice was higher than that of Simiao rice, but the difference was not significant. In lysimeter test under the moist condition after tillering, giant rice had a higher biomass than Simiao rice. However, the Cd concentration in the shoot was lower (Table 3), the Cd phytoextraction rates of giant and Simiao rice were all very low, less than 0.042% by the shoots (the data are not presented).

Table 3 Bio-concentration factors (BCF) and translocation factors (TF) of cd for different parts of giant and normal (cv. Simiao) rice

Table 4 Phytoextraction amounts and rates of cd by giant and Simiao rice in column test-1

In the column test-2, low Cd contents in grain and straw were obtained together with a very low Cd phytoextraction amounts for giant rice and Simao rice under sustained flooding condition in the 1 st half year (Table 5, Plant-1). In the 2nd half year under semi-dry condition, Cd contents in grain and straw of the two types of rice increased substantially (Table 5, Plant-2). Cd contents in the straw of giant rice were higher than 10 mg·kg⁻¹, increased by 564-fold compared to the early giant rice under sustained flooding, and the highest value was recorded in the regenerated giant rice (16.03 mg·kg⁻¹) with a BCF = 13.4. The Cd phytoextractions by the shoots of the second crops were significantly different and followed a decreasing order: Regenerated giant rice > Giant rice > P. hybridum > Simiao rice. Compared to Cd amount in the 0–20 cm topsoil, the phytoextraction rate of the regenerated giant rice reached 18.7%, followed by giant rice (13.0%) and P. hybridum (9.6%).

Based on soil Cd measurements after phytoremediation, the total amounts of Cd in topsoil were not significantly decreased after the first rice crops under sustained flooding. However in the second crops, the concentrations of soil Cd were decreased from about 1.20 mg·kg⁻¹ to about 0.97 mg·kg⁻¹ in giant rice or P. hybridum, to about 0.86 mg·kg⁻¹ in regenerated giant rice (Table S1). The soil Cd removal rate was 18%, 19% and 28% respectively by planting giant rice, P. hybridum and regenerated giant rice; in contrast, the removal rate was only 9.6% by planting Simiao rice. This result was consistent with that in the column test-1.

Table 5 Dry biomass, cd contents and phytoextractions of plants with different plant combinations in column test-2

Cd Subcellular Distribution in Rice Roots

Figure 2 illustrates the subcellular distribution of Cd in the roots of giant rice and Simiao rice under different water conditions. When considering the absolute quantity fixed, the amount of Cd fixed on the root surface and in the cell wall of giant rice was obviously greater than that of Simiao rice. In terms of percentage of Cd in the giant rice root, 37.93% was retained on the root surface, 44.51% was fixed in the root cell wall, and only 17.56% entered the endoplast as shown in the lysimeter test (moistened after the tillering stage) (Fig. 2a). In the column test (drained after the tillering stage), the proportions of these three parts were 31.09%, 59.35%, 9.56%, respectively (Fig. 2b). This meant that giant rice could fix more Cd by cell wall when higher Cd stress occurs, however, Simiao rice did not show the enhanced barrier effect of root cell wall under high Cd stress.

Fig. 2

Cd distribution in roots of giant and Simiao (normal) rice under different water conditions. a moistened after the tillering stage, b drained after the tillering stage

In order to understand whether the high Cd fixation capability on giant rice root surface was associated with the iron plaque, we conducted measurements of the iron plaque in corresponding to its Cd content on the root surface of giant rice. As depicted in Fig. 3a, keeping soil moist after the tillering stage, the iron plaque content on the root surface of giant rice was 108 mg per kg dry root, markedly superior to that of Simiao rice (81 mg per kg dry root). The Cd contents within the iron plaque of giant and Simiao rice constituted 63.21% and 38.80% of their total root surface Cd, respectively. The Cd fixed by iron plaque of giant rice represented 23.5% of the total Cd in root, compared to 11.7% for normal rice. Similar results were observed under the semi-dry condition (drained after the tillering stage) (Fig. 3b). These findings suggest that in comparison to Simiao rice, the root surface of giant rice has the potential to generate a more important iron plaque, thereby adsorbing and immobilizing a higher amount of Cd.

Fig. 3

Contents of iron plaque and its Cd on the root surface (mg per kg dry roots) of giant and Simiao (normal) rice under different water conditions. a moistened after the tillering stage, b drained after the tillering stage

Functional Groups in Rice Roots

FTIR spectra in Fig. 4 shows abundant functional groups in rice roots, especially oxygen-containing functional groups such as hydroxyl, carboxyl, C–O, P–O, Fe–O, Mn–O and Si–O. Compared to Simiao rice, giant rice presented obviously higher intensity of hydroxyl (3400 cm⁻¹), carboxyl (1417 cm⁻¹), P–O (1031 cm⁻¹), Mn-O (590 cm⁻¹) and Fe-O (525 cm⁻¹) in rice roots under the semi-dry water condition. The increases in intensity of these functional groups explained the greater binding ability on its roots of giant rice than those of Simiao rice.

Fig. 4

FTIR spectra of rice roots of giant and Simiao (normal) rice in the soil column test-2 (late rice under semi-dry condition)

To eliminate possible systematic errors, the relative abundance of target functional groups based on C–O–C (Das et al. 2024) was calculated (Table 6). Results showed that more COOH, Fe–O and Si–O–Si groups in giant rice roots than those of Simiao rice were verified, however, the others (–OH, P–O, Mn–O) were not ensured.

Table 6 The relative abundance of target functional groups based on C-O-C of rice roots of two rice varieties

Cd Retention in Stems and Leaves

The location map of different organs is figured out in Fig. 5a, the Cd contents of giant rice and Simiao (normal) rice under the semi-dry condition are given in Fig. 5b. Cd contents in stem tubes and nodes were generally higher than in leaves. The proportions of Cd translocated in the basal two nodes (III, IV) and the basal two stem tubes (III, IV) were more important for giant rice than that for Simiao rice, in contrast, the proportion of top node (I), stem tube (I) and leaf (I) were all less important for giant rice than that for Simiao rice. These indicated that the barrier effect of basal nodes and stems was more important for giant rice than that for Simiao rice, resulting in less Cd near grains.

Fig. 5

Cd distribution in shoots of giant rice and Simiao (normal) rice under semi-dry condition. a The location map of different organs, b Cd contents distribution in shoots of rice

Discussion

Growth Advantages of Giant Rice

Compared to the pot experiment, column test has subsoil layer and limited less the root growth, this is particularly good for high plants such as giant rice and to observe more appropriately the root length (Table 1). Column test also facilitates the water control for semi-dry conditions, permeating more important water layer beneath the soil surface in the container. Lysimeter test has much more soil than those of pot/column test and approaches to the real field test, can verify the Cd accumulation characteristics of giant rice under conditions similar to real fields. However, the number of lysimeters is limited (only 9), only one water condition (keeping soil moist) was tested.

Giant rice represents a novel variety of rice characterized by its tall stature, substantial biomass, and robust tillering capacity (Tang 2022). In this study, the average height of giant rice at maturity was observed in the lysimeter tests to be 195.5 cm, and 1.59 times of that of Simiao rice, thereby underscoring its notable “giant” character. The maximum dry weight yield of giant rice grain and straw in a single harvest was recorded at 7200 and 11,452 kg·ha⁻¹ respectively in the lysimeter tests, highlighting that giant rice as a high-biomass crop, in concordance with the grain yield of giant rice, can be 1.6-fold compared to those of conventional rice (Meng et al. 2022). This study also revealed that giant rice possessed a highly developed fibrous root system, capable to form a root network in a relatively short period and reach the 20–40 cm subsoil to additionally fix carbon compared to normal rice. Typically, the impact of a plant root on soil is confined to the rhizosphere area, extending only a few millimeters to 1–2 cm from the root surface (Yang et al. 2009). Obviously, the root system of giant rice, by virtue of its ability to come into contact with a larger volume of soil particles, can potentially influence and absorb more heavy metals in the soil. In this study, the growth of giant rice remained unaffected under dry condition after the tillering stage, suggesting it performed a drought resistance. Contrast to the typical Cd hyperaccumulator Sedum alfredii, which is a small plant and not easy to be cropped in field (Guo et al. 2016), giant rice presented distinct growth advantages for soil remediation and carbon fixation in paddy fields contaminated by Cd.

Application Potential of Giant Rice in Efficient and Sustainable Phytoremediation

Water conditions significantly affect the accumulation of Cd in rice (Huang et al. 2022). This is why the Cd accumulation characteristics of giant rice should be studied under different water conditions. Under sustained flooding, the Cd concentration in giant rice grain did not significantly differ from that in Simiao rice, which was a relatively low Cd cultivar (Wu et al. 1999), indicating a favorable trait of low Cd accumulation in grain of giant rice. Under moist condition after tillering in the lysimeter experiments, giant rice outperformed Simiao rice in terms of low grain Cd accumulation. Therefore, the tested giant rice cultivar (Judao-8) can be classified as a low-Cd cultivar under flooding or moist conditions. This is verified by the preliminary results from on-going field experiment on a mining-affected paddy field (Table S2). This allows farmers continuing rice production on moderately contaminated paddy fields and is favorable to maintain economic and social stability.

Drainage and oxidation conditions are conducive to increase the redox potential and reducing pH, thereby to promote the dissolution of soil Cd and its transformation into a more active form, making soil Cd more easily absorbed by plants (Ibaraki et al. 2009). Under a semi-dry condition, the Cd concentration in straw of giant rice was very significantly increased, and significantly higher than those of Simiao rice (Tables 2 and 5), providing a simple solution (only drainage at maximum tillering stage and no more irrigation) to increase Cd phytoextraction by high biomass plant without chemical additives. We calculated root/straw ratios and Cd absorption by unit mass of roots (Table S3) and fund that under semi-dry conditions, giant rice increased its roots in topsoil, root/straw ratio increased from 0.093 to 0.110, by contrast, Simiao rice decreased its root/straw ratio from 0.093 to 0.072. Furthermore, Cd absorption ability by unit mass of roots of Giant rice was more important than those of Simiao rice, and increased by 26.20 times from sustained flooding to semi-dry, also higher than those of Simiao rice (21.75 times). Because much more Cd were absorbed by giant rice (777.7 µg·pot^-1) (Table 4) than that by Simiao rice (218.8 µg·pot^-1) under semi-dry condition, even the most important part was retained in roots (343.2 µg·pot^-1) in topsoil, still much Cd (249.8 µg·pot^-1) translocated to giant rice straw and resulted in a larger BCF of straw than those of Simiao rice. In column test-2, Cd phytoextraction rate was 13.08% for giant rice and higher than that in column test-1 (5.03%). This might be because the column test-2 was conducted in the second half year, and less precipitations both in amount and in frequency in the second half year than those in the first half year in south China, resulted in the water level being lower and soil oxidation being higher in column test-2 than that in column test-1.

Research has shown that high-Cd rice “YangDao-6” can extract 7.2% of Cd from the soil (Liu et al. 2021b). After a year of extensive field trials, the soil Cd concentration in plots remediated by the high-Cd rice “Akita 110” decreased by 15.5% (Takahashi et al. 2016). Furthermore, rice has performed strong climate adaptability, its cultivation techniques are mature with a high degree of mechanization, making it a suitable candidate for phytoremediation in Cd-contaminated soils (Takahashi et al. 2021). Xie et al. (2020) showed that, in the cultivation of 12 phytoremediating plants, labor costs accounted for an average of 81.7% of the total costs, with rice having the lowest labor cost. Giant rice exhibits a similar even higher Cd extraction capability to perform as the high Cd accumulating rice varieties and thus it could be a suitable candidate for Cd phytoextraction.

The high regeneration ability of giant rice (Liang et al. 2020) may further facilitate the application of giant rice in the phytoextraction of Cd from paddy soils, because the regeneration of rice significantly decreases the labor costs of planting rice, reduces carbon footprints (Xu et al. 2022), and furthermore enhances soil quality (Zhang et al. 2023). The column test-2 also showed that the regenerated giant rice had higher Cd contents in aerial parts than the normally planted giant rice. The regenerated giant rice has higher root/shoot ratio and then increases Cd uptake. The similar trend was appeared in the later cuttings of ryegrass with higher Cd contents than those for the first cutting on Cd contaminated soils (Wu et al. 1993). Secondly, Cd stored in the old roots can also be translocated to the shoots of the regenerated rice in addition to the newly absorbed Cd. The Cd BCF_straw of the regenerated rice reached 13.4, this was even higher than those of other reported high-Cd rice varieties (BCF = 4.81–9.66) (Liu et al. 2021b; Shao et al. 2017), and obviously higher than a reported high biomass plant (king grass) with NH₄Cl enhancement (BCF = 0.595 ± 0.001) (Chen et al. 2017). With its high biomass, Cd phytoextraction rate of giant rice reached 18.7% and a Cd removal rate was 28.3% from topsoil in this study. With the hyperaccumulator Sedum alfredii, Cd phytoextraction rate was 13.3% in a pot experiment with a soil containing less Cd (0.69 mg/kg) and lower pH (4.43) (Sun et al. 2014). In a pot experiment to assess the Cd accumulation in peanut cultivars, the phytoextraction rate was estimated as being 9.9% (121.5 g/ha) by the selected high-accumulating cultivar (with soil Cd = 2.05 mg/kg, pH = 4.55) (Zhang et al. 2024). Furthermore, regenerating giant rice saves labor/machine input than normal planting, generates less carbon emission. These indicated that the regenerated giant rice was an effective and economic crop for phytoextracting soil Cd in the second half year after a normal early rice harvest, providing a sustainable way to remediate Cd-contaminated paddy fields.

Mechanisms of Cd Fixation by Giant Rice Roots

The root is the primary organ for Cd accumulation, exhibiting the highest Cd concentration in normal plants (Wu et al. 2020). The apoplastic barrier can mitigate the translocation of Cd from the root to the shoot via the apoplastic pathway, thereby diminishing Cd accumulation in the shoot of the rice (Qi et al. 2020). Previous study has indicated that iron plaque serves as a natural barrier against the intrusion of heavy metals into plants (Li et al. 2019; Qi et al. 2020). The strengthening of this iron plaque can augment the fixation of Cd in root surface of rice, thereby offering a mechanism for the external exclusion of Cd from the soil (Huang et al. 2020b). This experiment further revealed that giant rice roots can form a higher amount of iron plaque, which in turn adsorbed much more Cd (1.06 mg·kg⁻¹ dry root) than Simiao rice (0.19 mg·kg⁻¹ dry root) (Fig. 3a), thereby amplifying the function of root surface fixation of Cd. Compared to Simiao rice, more Fe–O and Mn–O functional groups in giant rice roots as showed by FTIR spectra (Fig. 4) could further confirm the remarkable barrier of Fe/Mn plaque on the root surface of giant rice.

The root cell wall has been considered to be a key protective barrier, significantly contributed to Cd detoxification and accumulation (Wang et al. 2015). The results of this study showed that the cell wall of giant rice retained significantly higher amount of Cd than that of Simiao rice, however the proportion of restricted Cd was not higher under flooding condition (Fig. 2a). On the other hand, under the semi-dry condition, the proportion of restricted Cd by cell wall of giant rice was enhanced to be higher than that of Simiao rice (Fig. 2b), showing that the potential Cd retention by cell wall of giant rice was also high, therefore played an important role to barrier Cd entering endoplast when Cd stress was high. Proteins and polysaccharides in cell wall can provide many potential ligands (including hydroxyl, carboxyl, aldehyde, and amino groups) to participate in various reactions with Cd, such as adsorption, ion exchange, complexation, and precipitation (Li et al. 2020), thereby retaining a large amount of Cd. Compared to Simiao rice, giant rice roots obviously contained more hydroxyl and carboxyl under the semi-dry condition as showed by FTIR spectra (Fig. 4).

Limitations of Giant Rice and Further Studies Needed

It is to be acknowledged that giant rice is generally not resistant to strong winds, though the proposed variety has thick stems, this rice is not suitable for coastal zones where strong cyclones or typhoons occur.

The reported phytoextraction data of giant rice were obtained with column experiments under controlled conditions, field experiments are absolutely necessary to obtain the real effectiveness, and to optimize the planting and harvesting parameters. In our column test, the rice shoots were cut at about 5 cm, the Vth stem (nearest to soil) were partly (about 1 cm/6 cm) harvested in our experiment. This should overestimate somewhat the phytoextraction, because the Vth stem (nearest to soil) of rice is generally not harvested by machine, though this part of basal stem is not important by mass. Because more cadmium accumulation in the basal stems of giant rice, we should suggest to harvest rice stems as more as possible when harvesting phytoextraction rice in the field. The real phytoextraction rate needs to be assessed by field experiment using machine and manual harvesting.

Using high biomass giant rice for phytoremediation, the rice straw will not be returned to the field, which may exacerbate the loss of trace minerals in the soil. Measuring the contents of these trace minerals such as manganese, zinc, iron in straw should be necessary in further studies in order to replenish appropriately these depleted micronutrients.

For regenerated giant rice, the old roots have stored Cd and can translocate this Cd to the shoots of the regenerated rice shoots in addition to the newly absorbed Cd. The contributions of these two kinds of Cd supplies is interesting to be elucidated in the further studies. Because Cd is concentrated in roots and basal stems of giant rice, separation and characterization of different components (cellulose, semi- cellulose, pectin, etc.) of cell wall of roots and basal stems are needed to well understand the molecular mechanisms of Cd retention by giant rice.

Conclusions

Under conditions of sustained flooding or moist after tillering, giant rice exhibits low Cd accumulation in grains. Under semi-dry condition, the roots and straw of giant rice demonstrate an impressive ability to accumulate Cd, in particular the straw of the regenerated giant rice effectively phytoextracted Cd from soil. Cd in shoots were highly concentrated in basal stems. The primary mechanisms by which giant rice retains Cd within its root system involve fixation at the root surface and binding with the cell wall. Giant rice can form more important iron plaque at root surface and intercept a higher amount of Cd than normal Simiao rice. Therefore, giant rice is a unique plant who is a low-Cd rice cultivar under wet conditions and meanwhile is an efficient Cd phytoextracting plant under semi-dry condition. Field experiments are necessary before extension of giant rice to the real phytoremediation of Cd-contaminated paddy fields in areas without strong cyclone or typhoon.

Materials and methods

Experimental Materials

The soil samples collected from a paddy field (Fe-accumuli-Stagnic Anthrosols)in Shantou City, Guangdong Province, China, were polluted by electronic waste disassembling. The chemical properties and Cd contents are summarized in Table 7. According to the classification of soil contamination from the National Soil Contamination Survey of China (MEPC 2014), the topsoil used is mediumly contaminated by Cd.

Table 7 Chemical properties and cd contents of the tested soils

Seeds of normal rice (Oryza sativa L., cv. Simiao) were provided by the Agricultural Science Research Institute of Zengcheng City, Guangdong Province. Simiao cultivar is a traditional (not hybrid) and still popular variety in Guangdong Province, China. Seeds of giant rice (Oryza sativa L., cv. Judao-8) were provided by the Institute of Subtropical Agriculture Ecology, Chinese Academy of Sciences. Seed stems of hybrid giant Napier grass (Pennisetum hybridum) were provided by the ecological farm of South China Agricultural University (Guangzhou, Guangdong Province, China). This plant is a typical high biomass plant cropped in highlands and mining sites for animal feed and bio-energy, recently used in the farmland phytoremediation due to its high biomass and easy-cropping (Chen et al. 2023).

Experimental Design

The column test was conducted in a mesh greenhouse for protecting against insect but permeating sunlight and rain at the experimental farm of South China Agricultural University. PVC column with a 15-cm inner diameter and 50-cm height was prepared and filled with the following materials from bottom to top: 5 cm of quartz sand, 20 cm of uncontaminated soil (subsoil), and 20 cm of Cd-contaminated soil (topsoil). The soil bulk density is 1.25 g·cm⁻³, meaning each layer has 4.5 kg air-dried soil.

The experiment was set up with two irrigation conditions: sustained flooding and Semi-dry (drained after the tillering stage). Three treatments were set under each irrigation condition: blank controls (no plant), planted with giant rice, and with Simiao rice, all treatments were repeated four times. For each irrigation condition, 12 soil columns were set up and accommodated within a plastic water tank (800 L, 150 cm×120 cm×60 cm). The soil within these columns was thoroughly saturated using tap water at the beginning of the experiment.

For the sustained flooding treatment, the water level was consistently maintained at a height of 1–2 cm above the soil surface. The semi-dry treatment was initially submerged under 1–2 cm of water during the early planting stage, and after the tillering stage, and the water level 10–15 cm beneath the soil surface was maintained. Following a growth period of 4 weeks in plastic incubators, the rice seedlings were transplanted into soil columns. Then, three seedlings were planted in each soil column. The growth period was from April to July 2022. Each rice crop was fertilized at a rate of 10 g N per m² with 15–15–15 (N–P₂O₅–K₂O) compound fertilizer according to the need of local normal rice. The crops were harvested when giant rice was matured.

Lysimeter test were conducted with Six 1 m×1 m×1 m sided lysimeters in a three-layer format. The base layer consisted of 50 cm of uncontaminated local soil, followed by a 20 cm layer of uncontaminated subsoil. The surface layer was composed of 20 cm of Cd-contaminated topsoil. To avoid the mixture of different soils, a black nylon net was deployed between different soil layers. Prior to initiating the experiment, the soil was thoroughly saturated via tap water irrigation.

Two types of rice (giant and normal rice), each with three replicates, were randomly assigned to six lysimeter pools. The cultivation of giant rice was at a density of 12 plants·m^–2, while the normal rice at a density of 16 plants·m^–2. In the early stages of planting, the soil was maintained at a flooded state with a depth of 1 ~ 2 cm water on the soil surface. After the tillering stage, the soil was kept moist (without standing water) by drip irrigation. The experiment was conducted from April to July for early rice crop and from middle August to the beginning of December for late rice crop using the same rice cultivars.

The Column test-2 was set up after column test of the first year to further verify the Cd accumulation characteristics under sustained flooding for the 1 st half year and semi-dry conditions for the 2nd half year. Four treatments were designed: normal rice (cv. Simiao) control (T1); giant rice normal planting (T2); giant rice regeneration in the second half year (T3); and P. hybridum in the second half year after early giant rice planting, in order to compare the phytoextraction efficiency among different high biomass plants under a semi-dry condition. The details of these treatments are presented in Fig. 6 and all treatments were repeated three times.

Fig. 6

Plant combination (plant-1 for safe production, plant-2 for phytoextraction) and water condition for different treatments in the soil column test-2

The facilities of this column test-2 are similar to the column test-1. The rice planting and the sustained flooding condition were the same as that of the column test-1, but different under the semi-dry condition. The water level of rice seedlings was drained to a lower level (5 cm lower) than in the column test-1 after the tillering stage, meaning a water level 15–20 cm being maintained beneath the soil surface. Concerning P. hybridum, one stem seedling was transplanted to each column when rice seedlings were transplanted. The growth period of early rice was from April to July 2023, and the growth period of subsequent crops was from middle August to the beginning of December.

Sample Collection, Preparation and Analysis

The plant height and biomass of the rice were quantified when giant rice was matured. In column test, rice plants were separated into grain, straw, root (in both topsoil and subsoil). After cutting shoots at about 5 cm above the soil surface, the whole soil column was taken-out and placed on a clean plastic cloth, the maximum single root length was recorded with the main root system. For collecting rice roots, the topsoil layer of each column was separated and placed into a clean plastic container, discarded the bulk soil from rice roots with a woody fork. The roots with adhered soils were placed into a 5 mm nylon sieve, rinsed carefully with tap water to eliminate soil particles, the roots were then collected into a plastic bag. The collection of roots in the subsoil layer of each column was similarly to that for the topsoil layer. The straw and roots were rinsed twice with tap water and also twice with distilled water, and left to drop water droplet in laboratory. Due to operational difficulties, root length and subsoil root biomass were not measured in the lysimeter test. Plant samples placed into new paper envelops were dried in an oven set at 70 °C for 30 min and then at 45 °C until a constant weight was achieved. These dried plant samples were pulverized using a stainless-steel grinder, passed through a 0.125 mm nylon sieve, and stored in sealed plastic bags for subsequent analysis.

Soil samples (both topsoil and subsoil) were air-dried and ground with an agate mortar, passed through 1 mm and 0.125 mm nylon sieves, and then stored in sealed plastic bags at 0–4 °C prior to analysis.

Plant Cd were analyzed using microwave digestion-graphite furnace atomic absorption photometry (HITACHI, Z-2700) based on the Chinese national standard method (GB 5009.15–2014). Test results were verified using Chinese national standard materials for plants (GBW(E)100348a).

For subcellular distribution of the root Cd, one group of fresh rice root samples was pulverized using a stainless-steel grinder and digested with a mixture of concentrated HNO₃-HClO₄ (4:1, v/v), and then used to determine the total Cd content (GB 5009.15–2014). Another group of fresh root samples was soaked in 20 mmol·L^–1 Na₂-EDTA to remove the Cd adsorbed at the root surface (Taylor and Crowder 1983). After filtration, the Cd concentration in the solution was determined and used to calculate the Cd content at the root surface. Finally, the remaining Cd content in the soaked root samples was determined using the same method for the analysis of root total Cd.

The Cd adsorbed by the root surface was removed as described previously, after which a methanol-chloroform mixture (2:1, v/v) was used to remove cell inclusions based on the method described by Hart et al. (1992).This left morphologically intact root cell walls, which were then analyzed for Cd content using the same method that was used for analysis of root total Cd (GB 5009.15–2014).

The iron plaque was extracted from the fresh root surface by referring to the DCB method developed by Taylor and Crowder (1983). 0.5 g fresh root was weighed and placed in a 50 ml conical flask containing 10 ml of 0.03 mol·L⁻¹ sodium citrate and 1.25 ml of 1 mol·L⁻¹ sodium bicarbonate. Subsequently, 0.75 g of sodium disulfite was added, and the mixture was shaken at 25℃ for 3 h. The filtrates of Cd and Fe were determined using ICP-MS (Agilent Technologies, 7700X) to calculate the concentrations of Cd and Fe on the root surface.

For FTIR spectroscopy of roots, the root samples were ground and homogenized as previously described for other roots samples but after air-dried. 2.00 mg dry root power were mixed with 100.0 mg KBr and pressed to get KBr pellet (Yu et al. 2023). FTIR spectra were collected by KBr pellet method using a Vertex 70 spectrometer (Bruker, USA) (Li et al. 2020). Data were acquired in the range of 400–4000 cm⁻¹ with 4 cm⁻¹ nominal resolution and 64 coadded scans.

Soil total Cd were analyzed using microwave digestion-graphite furnace atomic absorption photometry (HITACHI, Z-2700) based on the Chinese environmental standard method (HJ 832–2017). Test results were verified using Chinese national standard materials for soil (GBW07405a).

Data Analysis

Data processing and graphing were completed using Microsoft Excel 365 (Microsoft Corp., Redmond, WA, USA) and Origion 2021 (Origin Lab Corporation, Northampton, MA, USA). One-way analysis of variance (ANOVA) and Duncan’s multiple range tests were used to identify statistically significant differences between treatments (p<0.05) using SPSS 21.0 (IBM Corp., Armonk, NY, USA).

The following methods were used to calculate the factors related to heavy metal migration in the soil-plant system:

• Bioconcentration factor (BCF) = Cd concentration in a plant part/initial total Cd concentration in soil.

• Translocation factor (TF _a−b) = Cd concentration in organ b/Cd concentration in organ a.

• Phytoextraction rate = Cd extraction amount by plant shoots/initial total Cd amount in topsoil (0–20 cm).

• Removal rate = Decrease in soil total Cd content after planting/initial total Cd content in soil.