Small-sized selenium nanoparticles reduce the bioavailability of selenium in rice (Oryza sativa L.) by stimulating the formation of more iron plaque

Abstract

Nano-enabled agricultural technologies, particularly the application of small-sized selenium nanoparticles (Se NPs, <100 nm), demonstrate significant potential for stimulating crop growth and enhancing Se biofortification efficiency in dryland farming systems. However, the influence of Se NP size on bioavailability in flooded systems remains poorly understood. In this study, a rice (Oryza sativa L.) cultivation system was used to explore the relationship between the Se NP size (30 and 110 nm, 22 μm) and Se bioavailability under waterlogged conditions. Interestingly, 110 nm Se NPs significantly enhanced rice biomass, evidenced by a 19.4% increase in root fresh weight, and improved Se bioavailability in plants compared to both smaller (30 nm) and larger particles (2 μm). Mechanistically, smaller Se NPs (30 nm) appeared to enhance radial oxygen loss (ROL) and stimulate antioxidant enzyme activity (superoxide dismutase [SOD], peroxidase [POD], and catalase [CAT]). This physiological response promoted Fe(II) oxidation and subsequent iron plaque (IP) deposition on root surfaces, with DCB-extractable Fe levels showing a 29.1% increase compared to those of the 110 nm NP treatment group. The resulting increase in Se adsorption by the IP reduced Se translocation to aerial tissues, thereby decreasing its bioavailability in the smaller Se NP treatment group. Full life cycle experiments further confirmed that 110 nm Se NPs exhibited significantly higher Se accumulation in grains, an 85.3% increase compared to 30 nm NPs. These findings underscore the critical role of nanoparticle size and IP sequestration in determining Se bioavailability in rice grains. This study provides valuable insights for optimizing nano-Se fertilizers to improve Se biofortification in flooded agricultural systems.

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Environmental significance

Rice (Oryza sativa L.) serves as a primary food source for more than 3.5 billion individuals, with improving the bioavailability of selenium (Se) in grains being crucial for combating malnutrition in flooded agricultural areas. This research revealed that selenium nanoparticles (Se NPs) measuring 110 nm significantly enhanced Se accumulation in grains by 85.3% compared to 30 nm particles, while also promoting plant growth. The smaller 30 nm Se NPs led to excessive iron plaque (IP) formation on roots, which hindered Se absorption and decreased its transportation to grains. In contrast, the 110 nm Se NPs reduced IP accumulation, facilitating effective Se uptake and distribution to aboveground plant parts. These results offer a sustainable approach for optimizing nano-Se fertilizers in flooded agroecosystems, promoting precision agriculture for producing nutrient-rich rice and bolstering global food security and nutritional well-being.

1. Introduction

Nano-enabled agricultural technologies have gained increasing attention due to their high use efficiency, beneficial effects on crop growth, and superior biocompatibility.^1,2 Among them, selenium nanoparticles (Se NPs) have emerged as a promising tool in agriculture, owing to their effectiveness in promoting plant growth, enhancing crop yield and quality, and increasing Se content.^3–5 However, the bioavailability of NPs is a critical factor that determines their biological impact.^6–8 Previous studies have identified particle size as a key determinant of Se NP absorption, transformation, and bioavailability in soil–plant systems.^9–11 For instance, Lyu et al. reported that larger Se NPs significantly improved both Se content and utilization efficiency in wheat (Triticum aestivum L.) grains, thereby enhancing overall bioavailability in the wheat–soil system.⁹ Conversely, Cheng et al. demonstrated that smaller Se NPs markedly increased Se content in the rhizosphere and aerial tissues of B. chinensis—by 319.4% and 220.2%, respectively—and also stimulated photosynthesis and yield, indicating the pivotal role of particle size in Se biofortification.¹⁰ Despite these advances, most studies have focused on dryland crops, leaving a knowledge gap regarding the impact of NP size on Se bioavailability in flooded agricultural systems.

Rice (Oryza sativa L.), one of the world's most important cereal crops, is the dietary staple and primary nutrient source for approximately 3.5 billion individuals worldwide.^12,13 It plays a vital role in global food security and nutritional stability.^14,15 As a typical flooded crop, rice develops roots under anaerobic, waterlogged conditions.^16,17 Oxygen released through root aerenchyma oxidizes Fe(II) to Fe(III) at the root surface, where Fe(III) oxides precipitate and form iron plaque (IP).¹⁸ There are two types of IP on rice roots: structural iron and adsorbed iron. Structural iron, primarily composed of amorphous ferrihydrite, has a strong adsorption capacity.^19,20 Yu et al. suggested that IP strongly adsorbs arsenic (As), thereby reducing its mobility.²¹ Similarly, Li et al. demonstrated that IP can adsorb phosphorus through electrostatic interactions and form chemical bonds through coordination exchange reactions, thus affecting phosphorus bioavailability.²² Moreover, nanoparticles have been shown to affect IP formation. For example, Bao et al. observed that rice exposed to 14 nm and 25 nm CeO₂ NPs exhibited size-dependent differences in IP accumulation.²³ However, the relationship between the Se NP size and IP formation remains unclear. We hypothesize that Se NPs of varying sizes differentially influence IP formation, leading to variable adsorption capacities and, consequently, differing bioavailability outcomes.

To test this hypothesis, we conducted hydroponic and full life-cycle cultivation experiments under flooded conditions to examine how the Se nanoparticle (NP) size affects Se bioavailability. We addressed three specific aims: (1) assess the impact of Se NPs of different sizes on rice biomass and Se concentration; (2) elucidate the mechanisms by which Se NPs modulate iron plaque (IP) formation on rice roots; and (3) determine how the Se NP size influences bioavailability in rice–soil systems. These results are intended to advance understanding of Se NP bioavailability in semi-wetland crops and to inform Se biofortification strategies for flooded agricultural systems.

2. Materials and methods

2.1 Chemical synthesis and characterization of Se NPs

Se NPs with different particle sizes were synthesized by adjusting the concentration of the reducing agent, according to the method of Cheng et al.¹⁰ Briefly, 1.644 g Na₂S₂O₃ was dissolved in 20 mL of sodium dodecylsulfate (SDS, 0.01 M) to form solution A. Solution B was prepared by dissolving 11.54 mg SeO₂ in 20 mL of SDS solution (0.01 M). Se NPs (30 nm) were obtained by adding solution A (0.2 mL) to solution B (20 mL), followed by ultrasonication and centrifugation (8000 rpm, 10 min). Similarly, Se NPs (110 nm) were obtained by adding solution A (0.8 mL) to solution B (20 mL), followed by ultrasonication and centrifugation (8000 rpm, 10 min). For comparison, commercially available Se NPs (2 μm) were purchased from Nanjing Jike Biotechnology. Transmission electron microscopy (TEM, JEM-2100, Nippon Electronics Co., Japan) was used to characterize the morphology and size of the Se NPs. X-ray diffraction (XRD, XD-2, Purkinje General, China) was used to assess the crystal structure. X-ray photoelectron spectroscopy (XPS, Thermo K-Alpha, Thermo Fisher Scientific, USA) was employed to analyze the valence state of the synthesized Se NPs.

2.2 Plant cultivation and Se NPs treatment

Rice seeds (Oryza sativa L., Nanjing 9180) were acquired from the Jiangsu Academy of Agricultural Sciences. Following the method of Huang et al., seeds were disinfected with 30% (v/v) H₂O₂ for 15 min, rinsed three times with deionized water, and pre-soaked for 24 h.⁴⁶ They were then placed on moist gauze and germinated in the dark for 5 days at 28 °C and 85% relative humidity. Uniform seedlings were transferred to hydroponic pots containing 2 L of half-strength nutrient solution for 7 days, followed by cultivation in full-strength nutrient solution for an additional 21 days. The composition of the half-strength nutrient solution is listed in Table S1. The nutrient solution pH was adjusted to 5.6 with HCl and NaOH prior to initial use and upon each replacement. Plants were grown under controlled conditions: 14 h light per day, 26 °C/21 °C day/night temperatures, 65% relative humidity, with nutrient solution renewal every 3 days. Previous experiments have shown that adding 150 μg Se per L is the best option for hydroponics.^24,42 Hence, after 3 weeks, Se NPs (30 nm and 110 nm) and Se microparticles (Se MPs, 2 μm) were added to the culture solution at 150 μg Se per L. Control groups received no Se treatment. Each treatment included five replicates and three parallel samples.

After 10 days of hydroponic cultivation, the seedlings were transplanted into pots containing soil from Jiangsu Province during the regular rice growing season. Soil properties are shown in Table S2. Previous experiments have shown that adding 0.1 mg kg⁻¹ is the best option for pot experiments.²⁵ Therefore, the soil was pre-mixed with Se NPs (30 nm and 110 nm) or Se MPs (2 μm) at 0.1 mg kg⁻¹. The control soil received no Se supplementation. Each pot (22 cm high, 25 cm top diameter, 16 cm bottom diameter) contained 4.5 kg of soil. Each treatment had 3 replicates, with 2 rice plants per pot.

2.3 Determination of antioxidant enzyme activity, O₂·−, H₂O₂ and malondialdehyde (MDA) in rice seedling roots

Fresh root samples were homogenized in 0.1 M sodium phosphate buffer (pH 6.8) and centrifuged at 4000 rpm for 10 min.²⁶ SOD activity was measured based on the inhibition of nitroblue tetrazole (NBT) reduction by superoxide radicals. One unit of SOD activity was defined as the amount of enzyme required to inhibit photochemical reduction by 50%, measured at 560 nm using a microplate reader (Varioskan LuX, Thermo, USA).²⁷ POD activity was determined by monitoring the increase in absorbance at 470 nm due to guaiacol oxidation at 25 °C. One unit of POD activity was represented as the amount of enzyme that produced a change of 0.01 absorbance per min.²⁸ CAT activity was analyzed by the decrease in absorbance at 240 nm as H₂O₂ was decomposed. One unit of CAT activity was determined as the amount of enzyme that caused a reduction of 0.1 absorbance per minute.²⁹ Detection of superoxide radicals (O₂·−) and hydrogen peroxide (H₂O₂) are described in detail in Text S1 and the method for the determination of malondialdehyde MDA are described in detail in Text S2.

2.4 Determination of radial oxygen loss (ROL) in roots

ROL of rice seedlings was determined according to the method described by Kludze et al.³⁰ Briefly, roots were gently washed and stems were coated with paraffin wax. Each root was placed in a test tube containing 50 mL deionized water and dioxygen (DO). A 5 mL Ti³⁺-citrate solution, prepared in HCl solution under argon atmosphere, was injected promptly into each test tube using a plastic syringe, followed by a 20 mm coating layer of paraffin oil to prevent oxygen exchange. After incubation at 25 °C for 6 h, the absorbance of the partially oxidized Ti³⁺-citrate solution was measured at 527 nm with a microplate reader (Varioskan LuX, Thermo, USA). The ROL rate was calculated using the following equation:

Rate of ROL = c(y − z)/gt

where c (L) is the initial volume of Ti³⁺ solution, y (μmol L⁻¹) is the concentration of Ti³⁺ in the control group, z (μmol L⁻¹) is the final concentration of Ti³⁺ in the solution after 6 h treatment, g (g) is the root dry weight of rice seedlings, and t (h) is the time.

2.5 Extraction and characterization of iron plaque on the root surface

IP was extracted using the dithionite–citrate–bicarbonate (DCB) method. Briefly, fresh roots were thoroughly washed and soaked in an extractant containing 40 mL of 0.3 M Na₃C₆H₅O₇, 5 mL of 1 M NaHCO₃, and 1 g Na₂S₂O₄. The mixture was shaken at 220 rpm and 25 °C for 3 h, then diluted to 100 mL with deionized water. Fe and Se concentrations in the extract were measured using inductively coupled plasma mass spectrometry (ICP-MS, Thermo Fisher, Germany). The morphology and composition of the plaque were analyzed by scanning electron microscopy (SEM, SU8010, Hitachi, Japan), energy dispersive spectroscopy (EDS, X-MAX80, Oxford Instruments, UK), and XRD.

2.6 Determination of Se and Fe in plant tissues

Fe and Se contents in roots, shoots, and various tissues (roots, stems, leaves, and grains) of mature plants were determined using ICP-MS. Briefly, dried samples (25 mg) were digested in a microwave digestion system (Mars 6, USA) using 3 mL deionized water and 3 mL nitric acid (65–68%, GR grade). The digested samples were filtered with a microfiltration membrane (0.22 μm) and diluted to 50 mL. Each treatment had 5 replicates. Calibration was performed using multi-element standards (GRINM, gnm-m203814-2013), with Ge as the internal standard element (GSB 04-2826-2011, GRINM). Element recovery ranged from 84.9% to 102.9%.

2.7 Statistical analysis

All results are expressed as mean ± standard deviation (n = 3, P < 0.05). One-way analysis of variance (ANOVA) was conducted to evaluate differences among treatments, followed by Fisher's least significant difference (LSD) test for multiple comparisons.

3. Results and discussion

3.1 Characterization of Se NPs

Se NPs of different sizes were successfully synthesized and characterized. TEM images revealed that both types of Se NPs were spherical and uniformly dispersed, with average diameters of 35 ± 7 nm and 110 ± 13 nm, respectively, while commercial Se MPs exhibited a diameter of 2 ± 0.12 μm (Fig. 1a and S1a). The added SDS, a surfactant, ensured the activity and prevented the homogeneous agglomeration of Se NPs.³¹ Thus, good monodispersity of the synthesized Se NPs was ensured. These sizes fall within the range reported by Hu et al., who showed that Se NPs in the range of 40–140 nm could be effectively absorbed by crops.³² XRD pattern showed peaks at the (100), (101), (110), (102), and (111) planes, consistent with the standard Se crystal structure (JCPDS NO. 06-0362), indicating the successful synthesis of crystalline Se NPs (Fig. 1b and S1b). Applying the Debye–Scherrer equation,³³ the crystallite sizes were estimated to be approximately 30.8 nm and 110.6 nm, aligning closely with the TEM results. XPS analysis confirmed the presence of Se along with the C, N, and O elements (Fig. 1c), attributed to the organic stabilizers used during synthesis. High-resolution XPS spectra of Se 3d exhibited two types of characteristic peaks for Se: Se 3d_5/2 at 55.15 and 55.40 eV, and Se 3d_3/2 at 56.11 and 56.38 eV for the 30 and 110 nm NPs, respectively (Fig. 1d). These values are consistent with the literature reports, indicating that Se in the synthesized NPs was in the elemental (zero-valent) state.³⁴

Fig. 1 (a) TEM images of Se NPs. (b) Powder XRD patterns, (c) XPS survey and (d) high-resolution Se 3d XPS spectra of Se NPs.

3.2 Effects on growth and Se content of rice seedlings upon exposure to different-sized Se NPs

Exposure to Se NPs significantly influenced the growth and Se accumulation in rice seedlings. Compared with the control (CK), root fresh weights increased by 23.2% and 19.4% after treatment with 30 and 110 nm Se NPs, respectively (Fig. 2a). Shoot fresh weight was significantly enhanced when treated with 110 nm Se NPs but remained statistically unchanged under 30 nm Se NP and 2 μm Se MP treatments. This pattern was mirrored in the dry weight measurements of roots and shoots (Fig. 2b). Fresh and dry weights of rice at maturity under different treatments showed a similar trend to that of hydroponic seedlings (Fig. S2). These findings align with earlier studies, including Hussain et al., who observed a 25.4% increase in rice biomass after foliar application of Se NPs,³⁵ and Cheng et al., who showed size-dependent growth promotion in Brassica chinensis.¹⁰ However, in this study, 110 nm Se NPs proved more effective at promoting growth than the smaller 30 nm Se NPs, suggesting that the growth-promoting effect of Se NPs in rice is not strictly dependent on size.

Fig. 2 Biomass and Se content of rice seedlings under different treatments: (a) fresh weight, (b) dry weight, (c) Se content in shoots, and (d) Se content in roots.

In terms of Se uptake, treatments with Se NPs markedly enhanced Se accumulation in rice tissues. Compared to CK, Se content in rice shoots increased by 79.8, 98.0, and 23.9 fold under 30 nm, 110 nm, and 2 μm treatments, respectively (Fig. 2c). Root Se content showed a similar trend, with increases of 24.1, 29.9, and 6.6 fold (Fig. 2d), respectively. Among the treatments, the 110 nm Se NP group exhibited the highest bioavailability of Se, with shoot and root Se contents 22.9% and 24.0% higher than those in the 30 nm treatment group, respectively. These observations align with findings from Bao et al., who reported greater Se accumulation in rice under larger-sized NPs.²³ However, the results contrast with those of Cheng et al., who found greater Se accumulation in Brassica chinensis treated with smaller-sized NPs.¹⁰ This discrepancy may be attributed to the water-logged conditions of rice cultivation, which could reduce the mobility and uptake efficiency of smaller particles. Overall, the application of Se NPs—particularly the 110 nm variant—not only promoted biomass accumulation but also enhanced Se uptake in rice seedlings, highlighting their potential for improving crop Se nutrition under hydroponic conditions.

3.3 Response of IP formation to different sizes of Se NPs

IP, a protective barrier formed on the surface of rice roots, plays a crucial role in preventing the uptake of external contaminants, including heavy metals such as arsenic and cadmium.^36–38 In the present study, IP formation increased significantly upon exposure to Se NPs of 30 and 110 nm, compared to CK. In contrast, no significant difference in IP formation was observed between the Se MP (2 μm) treatment and CK (Fig. 3a). This enhancement in IP formation is further confirmed by quantifying the concentration of DCB–Fe on the root surface (Fig. 3b). The DCB–Fe content of the 30 nm and 110 nm Se NPs treatments was 4.3 and 3.4 times higher than CK, respectively. Compared to the Se MP treatment, DCB–Fe levels increased by 92.8% and 49.4% under the 30 nm and 110 nm Se NP treatments, respectively. Notably, the DCB–Fe content was 29.1% higher in the 30 nm treatment group than that in the 110 nm group. These findings suggest a size-dependent effect, where smaller Se NPs more effectively stimulate IP formation. A similar observation was reported by Bao et al., who found enhanced IP formation in rice roots treated with smaller CeO₂ NPs.²³ Further evidence was provided by SEM and EDS analyses (Fig. 4). SEM–EDS mapping showed strong signals for both the Se and Fe elements in root surfaces treated with 30 nm and 110 nm Se NPs, whereas weak or negligible signals were observed in the CK and Se MP treatment groups (Fig. 4c). The 30 nm Se NPs, in particular, included a rougher root surface and showed stronger Fe and Se element signals (Fig. 4b), confirming that smaller Se NPs are more effective in promoting IP deposition.

Fig. 3 Formation of IP on rice roots under different Se NP treatments: (a) image of rice roots, and (b and c) content of IP formed on rice roots as a function of Se particle size.

Fig. 4 Characterization of IP under different treatments: (a) SEM images of rice root surfaces, (b) elemental distribution of Se and Fe, and (c) EDS spectra.

In addition to Fe, Se content in the IP also increased significantly with decreasing particle size (Fig. 3c). Specifically, the Se content in the IP formed by 30 nm and 110 nm Se NPs was 14.4 fold and 10.2 fold higher, respectively, compared to that in CK. This parallels the observed trend in DCB–Fe content. The enhanced Se content in the IP can be attributed to the high surface area and abundant functional groups of iron (hydr)oxides, which facilitate Se adsorption via complexation and co-precipitation, as previously described by Huang et al.³⁹ Zhang et al. also showed that the application of Fe(II) significantly increased Se concentration in DCB extracts, which is consistent with our results.⁴⁰ Therefore, the promotion of IP formation by smaller Se NPs enhances Se adsorption on rice root surfaces, thereby reducing Se translocation into root tissues and, subsequently, into aerial parts. This size-dependent effect on IP dynamics likely contributes to reduced Se bioavailability in edible parts such as rice grains. Overall, these findings reveal a strong correlation between the Se NP size, IP formation, and the regulation of Se accumulation in rice tissues, suggesting a dual role of Se NPs in promoting biomass while modulating Se uptake through IP-mediated pathways.

3.4 Mechanism of IP formation regulated by the Se NPs size

IP formation on rice root surfaces is primarily driven by ROL, which enhances oxygen availability in the rhizosphere and facilitates the oxidation of Fe(II) to Fe(III).^41,42 Consequently, ROL is considered to be the most significant physiological process underpinning IP formation.^43,44 In the present study, ROL rates were 18.1, 15.1, and 10.1 μmol O₂ g⁻¹ DW h⁻¹ following exposure to 30 nm and 110 nm Se NPs, and 2 μm Se MPs, respectively (Fig. 5a). These results indicate that smaller Se NPs promote significantly higher oxygen release from roots. Ando et al. reported that three-week-old rice seedlings released 9 times more O₂ from their roots than that required for root respiration, suggesting that root-secreted O₂ is a primary contributor to IP formation.⁴⁵ Similarly, Huang et al. observed that Se application enhanced ROL and thereby accelerated IP formation in hydroponically grown rice.⁴⁶ In this study, a positive correlation was found between ROL and Fe content in IP (Fig. S3), further supporting the critical role of ROL in the formation of IP. Specifically, smaller Se NPs stimulated greater oxygen release into the rhizosphere, enhancing the oxidation of Fe(II) to Fe(III) and thereby promoting more extensive IP formation.

Fig. 5 Mechanism of IP formation regulated by the Se NP size: (a) radial oxygen loss (ROL) in rice roots under different treatments, (b–d) activities of CAT, POD, and SOD in roots under different treatments, (e) fluorescence histochemical staining of roots for superoxide radical (O₂·−) (red) and hydrogen peroxide (H₂O₂) (green).

Previous research revealed two primary mechanisms known to enhance ROL in rice roots: (1) translocation of O₂ from the shoots to the roots and (2) local enzymatic generation of O₂ within root tissues.^47,48 The latter involves the enzymatic oxidation of organic acids—including glyoxylic, oxalic, and formic acids—producing carbon dioxide (CO₂) and hydrogen peroxide (H₂O₂).⁴⁹ H₂O₂ can subsequently be enzymatically decomposed into water and molecular O₂, thus increasing local oxygen availability around the roots.⁵⁰ In this study, exposure to 30 nm Se NPs significantly elevated antioxidant enzyme activities in rice roots. Specifically, CAT, POD, and SOD activities increased by 6.3%, 11.4%, and 18.4%, respectively, compared with CK (Fig. 5b–d). These increases were also higher than those observed under 110 nm Se NP and 2 μm Se MP treatments. Similarly, antioxidant enzyme (SOD) activity in rice roots at maturity was highest under the 30 nm Se NP treatment (Fig. S4). A similar pattern was reported by Zahedi et al., who discovered that foliar application of 10 nm Se NPs in pomegranate trees notably enhanced CAT, APX, SOD, and POD activities compared to larger Se NPs (50 nm), which produced more modest effects.⁵¹ Furthermore, changes in superoxide radicals and hydrogen peroxide in rice roots further demonstrate that small-sized Se NPs are more effective in enhancing the activity of antioxidant enzymes. Compared to Se NPs (110 nm) and CK, superoxide radical (O₂·−) and hydrogen peroxide (H₂O₂) accumulation were significantly reduced under 30 nm Se NP treatment (Fig. 5e and S5). Similarly, the same trend was observed in malondialdehyde (MDA) content (Fig. S6). Therefore, the enhanced enzyme activity induced by smaller Se NPs (30 nm) may lead to an increase in oxygen production within the root system by degrading O₂·− and H₂O₂ in rice roots, thereby increasing the ROL level (Fig. 5a), and subsequently promoting IP formation (Fig. 3b and 4). This mechanistic insight underscores the importance of Se NP size in modulating oxidative metabolism and oxygen dynamics in rice roots, ultimately influencing IP deposition and the rhizosphere's physicochemical environment.

3.5 The effects of IP formation on the Se distribution in mature rice plants

SEM images revealed that the root surface exhibited a non-uniform, porous, or flocculent structure following exposure to Se NPs, particularly in the 30 nm treatment group (Fig. 6a and b). These morphological features suggest substantial IP formation on the root surface after exposure to Se NPs (30 and 110 nm), with more pronounced effects observed in the 30 nm group. Quantitative analysis showed that the DCB–Fe content was highest in the 30 nm Se NP treatment group. A clear size-dependent trend was observed, where smaller NPs resulted in higher DCB–Fe accumulation (Fig. 6c). Specifically, the DCB–Fe content was increased by 29.1% in the treatment with 30 nm Se NPs compared to that in the 110 nm Se NP treatment (Fig. 6c). Similarly, DCB–Se content increased by 42.7% upon exposure to 30 nm Se NPs compared with that to the 110 nm Se NP treatment group (Fig. 6d). The EDS results further confirmed the presence of large amounts of Fe and Se on the root surface, which also indicated that IP has a high adsorption capacity for Se. Specifically, stronger elemental Fe and Se signals were detected on the root surface under the Se NP (30 nm) treatment, suggesting greater effectiveness in enhancing IP deposition (Fig. 6b and S7). XRD also showed strong adsorption of Se by the iron plaque under different treatments, especially under 30 nm Se NP treatment (Fig. S8). These results suggest that the IP acts as an effective adsorbent for Se, and its abundance and composition significantly affect the bioavailability of Se NPs. As the IP content increases, more Se is retained on the root surface, potentially limiting its translocation to aerial parts of the plant.

Fig. 6 Formation of IP and Se distribution in the mature rice plant. (a) Image of rice roots, (b) SEM images and EDS elemental maps showing Se and Fe under different treatments, (c and d) content of IP formed on rice roots as a function of Se NP size, and (e) Se content in rice organs under different treatments.

Elemental analysis of mature rice plants further confirmed this pattern. Compared with that of CK, Se content in roots, stems, leaves, and grains significantly increased upon exposure to Se NPs (Fig. 6e). Interestingly, the 110 nm Se NP treatment group exhibited the highest Se accumulation across all plant tissues, followed by the 30 nm group. In contrast, Se MP and CK treatments showed the lowest Se accumulation. Specifically, compared with the 30 nm Se NP treatment group, the 110 nm Se NP treatment group resulted in the Se content increasing by 39.2%, 27.9%, 52.7%, and 85.3% in roots, stems, leaves, and grains, respectively (Fig. 6d). This result can be explained by the enhanced IP formation induced by smaller Se NPs, which promote greater Se adsorption onto the IP and subsequently reduce Se uptake and translocation within the plant. Although the 2 μm Se MPs did not promote the deposition of IP on the root surface, their bioavailability was also low, likely due to the limited mobility of larger particles. Collectively, these results suggest that while smaller Se NPs strongly stimulate IP formation and Se sequestration on the root surface, this process decreases Se bioavailability in the plant. Conversely, larger Se NPs (110 nm) are less prone to IP-mediated immobilization and demonstrate superior translocation to aboveground tissues, thereby increasing Se accumulation in edible plant parts. Moreover, rice growth and rice panicle quality were improved under the treatment of Se NPs (110 nm) (Fig. S9).

4. Conclusion

This study provides compelling evidence that the bioavailability of selenium nanoparticles (Se NPs) in rice is strongly influenced by particle size. Small-sized Se NPs (e.g., 30 nm) were found to significantly enhance radial oxygen loss (ROL) in rice roots by stimulating oxidase activity, leading to increased iron plaque (IP) formation on the root surface. This IP acts as a barrier that adsorbs and retains Se at the root–soil interface, thereby limiting Se translocation to aerial plant parts and reducing overall bioavailability.

In contrast, larger Se NPs (e.g., 110 nm) induced less IP formation, resulting in lower Se retention on the root surface and enhanced translocation within the plant. Consequently, rice plants exposed to larger Se NPs accumulated significantly more Se in roots, stems, leaves, and grains, along with increased biomass production, indicating greater agronomic effectiveness.

Together, these findings elucidate a clear, size-dependent mechanism of Se NP behavior in soil–plant systems. The study highlights the trade-off between enhanced root-level Se sequestration via IP formation and systemic Se bioavailability, governed by nanoparticle size. These insights offer valuable guidance for the design and application of nano-Se fertilizers in different crop tillage systems (flood system or dryland system), enabling tailored strategies for maximizing nutrient uptake and improving crop nutrition.

Author contributions

Bingxu Cheng: writing – original draft, methodology, investigation, formal analysis, and data curation. Bo Chen: writing – review & editing, and conceptualization. Jing Liu: writing – review & editing, and formal analysis. Jiangshan Zhang: methodology, investigation, and data curation. Yubo Lu: investigation. Chuanxi Wang: writing – review & editing, supervision, funding acquisition, and conceptualization. Zhenyu Wang: writing – review & editing, and formal analysis.

Conflicts of interest

The authors report no conflict of interest to this work.

Data availability

Data will be availability on request. Supplementary information (SI): SI includes detection methods for superoxide radicals, hydrogen peroxide, and MDA, nutrient solution formulas, and soil physical and chemical properties. Fig. S1–S9 are respectively the characterization of Se NPs (2 μm), the biomass of rice at the mature stage, the relationship between Fe and Se content in the iron plaque (IP) and radial oxygen loss (ROL) of rice, SOD enzyme activity, the characterization of superoxide radicals and hydrogen peroxide in rice roots, the content of MDA, the characterization of iron plaque on the root surface after Se NPs (2 μm) treatment, the XRD characterization of iron plaque on the root surface, and the effect photos of rice at the mature stage. See DOI: https://doi.org/10.1039/d5en00639b.

Acknowledgements

This research was supported by the National Natural Science Foundation of China (42307352, 42421005, and 42322705) and funded by the Basic Research Program of Jiangsu (BK20230044) and the Fundamental Research Funds for the Central Universities (JUSRP202501065).

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Footnote

† Bingxu Cheng and Bo Chen contributed equally to this work.

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