Method for repairing cadmium contaminated soil by synergistic effect of double-layer microspheres and EDDS / NTA and bidens pilosa

Abstract

This invention provides a method for remediating cadmium-contaminated soil with Bidens trifoliata using bilayer microspheres in synergistic EDDS / NTA enhancement. The method includes preparing bilayer biodegradable microspheres consisting of a humic acid shell and a PHA core loaded with citric acid and nicotinamide; applying the microspheres to cadmium-contaminated soil and sowing Bidens trifoliata; applying EDDS and NTA aqueous solutions in stages during the 30th to 50th day of plant growth; and harvesting the above-ground parts on the 60th day of growth to remove cadmium. This invention promotes plant growth by continuously releasing humic acid from the shell, pre-activates soil cadmium by releasing citric acid from the core, and, combined with the staged application of biodegradable chelating agents EDDS and NTA, improves the bioavailability and plant translocation efficiency of cadmium. This invention is simple to operate, environmentally friendly, and can effectively shorten the remediation cycle of moderately and severely cadmium-contaminated farmland soil in arid and semi-arid regions, showing promising prospects for widespread application.

Description

Technical Field

[0001] This invention relates to the field of phytoremediation technology for heavy metal contaminated soil, specifically to a method for enhancing the remediation of cadmium-contaminated soil with Bidens trifoliata using bilayer microspheres in synergistic EDDS / NTA enhancement. Background Technology

[0002] Cadmium (Cd), a heavy metal, has become one of the major pollutants in farmland soil in my country due to its high toxicity, easy migration, and difficulty in degradation. Cd can be transferred through the food chain in soil, posing a serious threat to the ecological environment and human health. Currently, remediation technologies for Cd-contaminated soil mainly include physical remediation, chemical remediation, and bioremediation. However, physical and chemical remediation are often costly, prone to secondary pollution or damage to soil structure, and difficult to meet the needs of large-scale remediation of large areas of farmland.

[0003] Bioremediation technology has garnered widespread attention due to its environmental friendliness, relatively low cost, and lack of soil ecosystem disruption. Bidens trifoliata is a widely recognized Cd hyperaccumulator; its roots efficiently absorb Cd from the soil and transport it to the aboveground parts (stems and leaves). Regular harvesting can gradually reduce soil Cd concentration, achieving in-situ remediation. However, this technology faces significant efficiency bottlenecks in practical farmland applications. These bottlenecks primarily stem from the low biomass of Bidens trifoliata under natural conditions, resulting in a limited harvestable plant body per season and insufficient total Cd removal from the soil per unit time. Furthermore, Cd in the soil exists mainly in low-bioavailability forms such as carbonate-bound and iron-manganese oxide-bound forms, making direct absorption by plant roots difficult. The transport of Cd from roots to aboveground parts is limited by the plant's own physiological mechanisms; even if roots absorb Cd, efficient transfer to harvestable aboveground parts is challenging. These factors collectively lead to lengthy remediation cycles and overall low efficiency, failing to meet the urgent need for moderately or severely polluted farmland to reach safe utilization standards within a reasonable timeframe.

[0004] To improve phytoremediation efficiency, researchers have attempted to add artificial chelating agents to the soil to convert insoluble Cd into water-soluble Cd chelates through complexation, thereby enhancing Cd bioavailability. However, existing chelating agent enhancement technologies still have the following shortcomings: excessive single-dose chelating agents can easily cause toxicity to plants; chelating agents degrade rapidly in the soil, making it difficult to maintain an activation effect that matches the plant growth cycle; and long-term use of chelating agents alone may disrupt the soil microecology. Therefore, developing a synergistic remediation method that can continuously provide plant nutrition and pre-activate soil Cd while precisely replenishing chelating agents in stages to enhance absorption and translocation is of great significance for overcoming the bottleneck of phytoremediation efficiency and promoting the practical application of Cd pollution control in farmland. Summary of the Invention

[0005] The technical problems to be solved: Addressing the shortcomings of existing phytoremediation methods for cadmium-contaminated soil, such as low plant biomass, poor cadmium bioavailability in the soil, low cadmium translocation efficiency, and the mismatch between the activation effect of single chelating agent enhancement methods and the plant growth cycle, this paper proposes a method for enhancing the remediation of cadmium-contaminated soil with Bidens pilosa using bilayer microspheres in synergistic EDDS / NTA. This method utilizes bilayer biodegradable microspheres to continuously release humic acid to promote plant growth and release citric acid to pre-activate cadmium in the soil. During the mid-to-late stages of plant growth, the biodegradable chelating agents EDDS and NTA are applied in stages to achieve efficient cadmium absorption and removal, shorten the remediation cycle of cadmium-contaminated soil, and is environmentally friendly.

[0006] Technical Solution: A method for remediating cadmium-contaminated soil using Bidens trifoliata with synergistic effects of double-layer microspheres and EDDS / NTA, comprising the following steps: S1. Prepare bilayer biodegradable microspheres, the microspheres comprising a shell and a core, the shell containing humic acid, and the core containing polyhydroxyalkanoate and citric acid and nicotinamide loaded therein; S2. Soak the seeds of Bidens trifoliata in NaClO solution for disinfection, then rinse thoroughly with clean water and air dry for later use; S3. Apply the double-layer biodegradable microspheres obtained in step S1 into cadmium-contaminated soil and sow disinfected Bidens trifoliata seeds. S4. Apply EDDS and NTA aqueous solutions to the soil in several applications between the 30th and 50th day of growth of Bidens trifoliata; S5. Harvest the above-ground parts of the Bidens trifoliata around day 60 of its growth and remove the cadmium from the field.

[0007] A preferred method for preparing bilayer biodegradable microspheres includes the following steps: S11. Dissolve polyhydroxy fatty acid ester in dichloromethane to obtain a polymer solution, add citric acid and nicotinamide to it, and stir at high speed to form a uniform suspension; S12. The suspension was added dropwise to the PVA solution, the solvent was evaporated by stirring, the suspension was collected by centrifugation, washed, and freeze-dried to obtain core microspheres; S13. Disperse humic acid powder in water, add KOH to adjust the pH to 8.0-9.0 to obtain potassium humate solution, dissolve sodium carboxymethyl cellulose in water, heat to dissolve, cool and mix with potassium humate solution to obtain composite coating solution; S14. Immerse the core microspheres obtained in S12 into the composite coating solution in S13, stir to adsorb, leach, and then vacuum dry to obtain bilayer microspheres.

[0008] Preferably, in step S11, the concentration of the polymer solution is 5-15 wt%, the amount of citric acid added is 10-15 wt% of the dry weight of the polymer, and the amount of nicotinamide added is 0.5-2.0 wt% of the dry weight of the polymer.

[0009] Preferably, in step S12, the concentration of the PVA solution is 0.5~2wt%, the stirring speed for evaporating the solvent is 300~600rpm, and the particle size of the core microspheres is 50~200μm.

[0010] Preferably, in step S13, the concentration of potassium humate solution is 2-3 wt%, the concentration of sodium carboxymethyl cellulose solution is 1-1.5 wt%, and the volume ratio of potassium humate solution to sodium carboxymethyl cellulose solution after mixing is 2:1.

[0011] Preferably, in step S14, the impregnation and adsorption time is 10–30 min, the vacuum drying temperature is 30–40 °C, and the drying is carried out to constant weight. The outer shell thickness of the bilayer microspheres is 20–50 μm.

[0012] Preferably, in step S3, the amount of double-layer biodegradable microspheres applied is 1-5g per kilogram of soil.

[0013] Preferably, in step S4, EDDS and NTA are applied three times, with an interval of 10 days between each application, and the total volume of EDDS and NTA solution applied each time is 1 L / m³. 2 The concentrations were all 0.2~2 mmol / L.

[0014] Beneficial effects: The method of using bilayer microspheres in synergistic EDDS / NTA enhancement of Bidens trifoliata for remediation of cadmium-contaminated soil has the following advantages: This invention encapsulates potassium humate in a coating, utilizing the biodegradability of sodium carboxymethyl cellulose to achieve its continuous release into the soil. Potassium humate has good water solubility, forming a homogeneous system in the coating solution, ensuring its uniform distribution within the coating. After application to the rhizosphere, it transforms into humic acid in a slightly acidic environment. Its rich carboxyl and phenolic hydroxyl groups promote root elongation and lateral root proliferation in Bidens pilosa, increasing the root absorption surface area. The potassium ions released by potassium humate enhance the plant's resistance to cadmium stress and synergistically induce the plant to increase photosynthetic pigment content and antioxidant enzyme activity, thereby enhancing the plant's growth performance under cadmium stress.

[0015] This invention encapsulates citric acid and nicotinamide within the core of a phosphoric acid (PHA). Utilizing the slow degradation of PHA in soil over 8-12 weeks, both are continuously released during the later stages of plant growth. Citric acid, a low-molecular-weight organic acid, undergoes a complexation reaction with insoluble cadmium in the soil, including carbonate-bound and iron-manganese oxide forms, through its carboxyl group, forming a soluble citric acid-cadmium complex. This transforms cadmium from a solid phase into a form absorbable by plants, thus pre-activating it. Nicotinamide, once absorbed by the plant, promotes the transport of cadmium from the roots to the aboveground parts, increasing the cadmium transport coefficient.

[0016] This invention involves applying EDDS and NTA in stages during the later stages of plant growth. Simultaneously, citric acid released from the PHA kernel pre-converts some of the insoluble cadmium into an exchangeable state, providing a more readily complexing substrate for the chelating agent. Both EDDS and NTA are biodegradable green chelating agents; multiple carboxyl and amino groups in their molecules can react with Cd in the soil solution. 2+ It forms stable water-soluble complexes, effectively reducing the adsorption affinity of soil solids for cadmium, promoting the conversion of non-exchangeable cadmium such as carbonate-bound and iron-manganese-bound cadmium into exchangeable forms, and improving the absorption efficiency of cadmium by the roots of Bidens trifoliata. Attached Figure Description

[0017] Figure 1 This is a schematic diagram of phytoremediation technology. Figure 2 Biomass of Bidens trifoliata sampled after 60 days of growth; Figure 3 The percentage of different forms of Cd in the soil. Detailed Implementation

[0018] The present invention will be further described below with reference to embodiments. These embodiments are illustrative of the present invention, but the present invention is not limited to these embodiments: In both the embodiment and the comparative example of this invention, the plants were grown in flowerpots of uniform size, and the soil used was simulated Cd-contaminated soil. The preparation method was as follows: farmland soil and nutrient soil were mixed in a 1:1 ratio, with 1.5 kg of soil added to each pot. The Cd concentration in the soil was adjusted to the target pollution level (15 mg / kg) using a CdCl2·2.5H2O solution. -1 ); Till the prepared soil to a depth of 10-20cm to enhance soil looseness and aeration, and to distribute pollutants evenly on the surface; after tilling, level the soil surface, maintain the soil moisture content at about 60-70%, and let it stand for three weeks to mature, in order to stabilize the soil structure and create a physical environment conducive to seed germination.

[0019] Example 1

[0020] The preparation of bilayer biodegradable microspheres includes the following steps: S11. Dissolve 10.0g of polyhydroxy fatty acid ester in 90g of dichloromethane to obtain a polymer solution, add 1.0g of citric acid and 0.05g of nicotinamide to it, and stir at high speed to form a uniform suspension; S12. The suspension was added dropwise to a 2.0 wt% PVA solution, the solvent was evaporated by stirring at 600 rpm, the suspension was collected by centrifugation, washed, and freeze-dried to obtain core microspheres with a particle size of 50 μm. S13. Disperse 2.0g of humic acid powder in 100mL of water, add KOH to adjust the pH to 8.0, dissolve 1.0g of sodium carboxymethyl cellulose in 100mL of water, heat to dissolve, cool, and then mix the potassium humate solution and sodium carboxymethyl cellulose solution at a volume ratio of 2:1 to obtain a composite coating solution; S14. Immerse the core microspheres obtained in S12 into the composite coating solution in S13, stir and adsorb for 10 min, leach out and vacuum dry at 30℃ to obtain bilayer microspheres with a shell thickness of 20 μm.

[0021] A method for enhancing the remediation of cadmium-contaminated soil with Bidens trifoliata using bilayer microspheres in synergistic EDDS / NTA enhancement includes the following steps: S1. Preparation of bilayer biodegradable microspheres; S2. Soak Bidens trifoliata seeds in a 1% NaClO solution for 10 minutes to disinfect them, then rinse them thoroughly with clean water and air dry them for later use. S3. Apply 3g of double-layer microspheres per kilogram of soil to cadmium-contaminated soil and sow sterilized Bidens trifoliata seeds. When sowing, scatter the seeds evenly on the soil surface and cover them with about 2cm of Cd-contaminated simulated soil. During the germination period, maintain the temperature at 20±5℃ and the light at 14h per day to promote successful seed germination. After the seedlings emerge, thin them out to maintain a density of about 15 plants per pot. S4. Prepare NTA aqueous solution with a concentration of 1 mmol / L and EDDS aqueous solution with a concentration of 1 mmol / L respectively. On the 30th day of the growth of Bidens trifoliata, apply the EDDS and NTA aqueous solutions evenly into the soil by root irrigation. Then, apply them a second and a third time on the 40th and 50th days of the growth of Bidens trifoliata, respectively. The application method is the same as the first treatment each time.

[0022] S5. Harvest the above-ground parts of the Bidens trifoliata around day 60 of its growth and remove the cadmium from the field.

[0023] Example 2

[0024] The preparation of bilayer biodegradable microspheres includes the following steps: S11. Dissolve 15g of polyhydroxy fatty acid ester in 85g of dichloromethane to obtain a polymer solution, add 2.25g of citric acid and 0.3g of nicotinamide to it, and stir at high speed to form a uniform suspension; S12. The suspension was added dropwise to a 0.5 wt% PVA solution, the solvent was evaporated by stirring at 300 rpm, the suspension was collected by centrifugation, washed, and freeze-dried to obtain core microspheres with a particle size of 200 μm. S13. Disperse 3g of humic acid powder in 100mL of water, add KOH to adjust the pH to 8.0-9.0, dissolve 1.5g of sodium carboxymethyl cellulose in 100mL of water, heat to dissolve, cool, and then mix the potassium humate solution and sodium carboxymethyl cellulose solution at a volume ratio of 2:1 to obtain a composite coating solution; S14. Immerse the core microspheres obtained in S12 into the composite coating solution in S13, stir and adsorb for 30 min, leach out and vacuum dry at 40℃ to obtain bilayer microspheres with a shell thickness of 50 μm.

[0025] A method for enhancing the remediation of cadmium-contaminated soil with Bidens trifoliata using bilayer microspheres in synergistic EDDS / NTA enhancement includes the following steps: S1. Preparation of bilayer biodegradable microspheres; S2. Soak the seeds of Bidens trifoliata in a 1% NaClO solution for 10 minutes to disinfect them, then rinse them thoroughly with clean water and air dry them for later use. S3. Apply 3g of double-layer microspheres per kilogram of soil to cadmium-contaminated soil and sow sterilized Bidens trifoliata seeds. When sowing, scatter the seeds evenly on the soil surface and cover them with about 2cm of Cd-contaminated simulated soil. During the germination period, maintain the temperature at 20±5℃ and the light at 14h per day to promote successful seed germination. After the seedlings emerge, thin them out to maintain a density of about 15 plants per pot. S4. Prepare NTA aqueous solution with a concentration of 0.4 mmol / L and EDDS aqueous solution with a concentration of 0.4 mmol / L respectively. On the 30th day of the growth of Bidens trifoliata, apply the EDDS and NTA aqueous solutions evenly into the soil by root irrigation. Then, apply them for the second and third time on the 40th and 50th days of the growth of Bidens trifoliata, respectively. The application method is the same as the first treatment each time.

[0026] S5. Harvest the above-ground parts of the Bidens trifoliata around day 60 of its growth and remove the cadmium from the field.

[0027] Example 3

[0028] The preparation of bilayer biodegradable microspheres includes the following steps: S11. Dissolve 10.0g of polyhydroxy fatty acid ester in 90g of dichloromethane to obtain a polymer solution, add 1.2g of citric acid and 0.1g of nicotinamide to it, and stir at high speed to form a uniform suspension; S12. The suspension was added dropwise to a 1.0 wt% PVA solution, the solvent was evaporated by stirring at 450 rpm, the suspension was collected by centrifugation, washed, and freeze-dried to obtain core microspheres with a particle size of 100 μm. S13. Disperse 2.5g of humic acid powder in 100mL of water, add KOH to adjust the pH to 8.0, dissolve 1.2g of sodium carboxymethyl cellulose in 100mL of water, heat to dissolve, cool, and then mix the potassium humate solution and sodium carboxymethyl cellulose solution at a volume ratio of 2:1 to obtain a composite coating solution; S14. Immerse the core microspheres obtained in S12 into the composite coating solution in S13, stir and adsorb for 20 min, leach out and vacuum dry at 35℃ to obtain bilayer microspheres with a shell thickness of 30 μm.

[0029] A method for enhancing the remediation of cadmium-contaminated soil with Bidens trifoliata using bilayer microspheres in synergistic EDDS / NTA enhancement includes the following steps: S1. Preparation of bilayer biodegradable microspheres; S2. Soak Bidens trifoliata seeds in a 1% NaClO solution for 10 minutes to disinfect them, then rinse them thoroughly with clean water and air dry them for later use. S3. Apply 1.5g of double-layer microspheres per kilogram of soil to cadmium-contaminated soil and sow sterilized Bidens trifoliata seeds. When sowing, scatter the seeds evenly on the soil surface and cover them with about 2cm of Cd-contaminated simulated soil. During the germination period, maintain the temperature at 20±5℃ and the light at 14h per day to promote successful seed germination. After the seedlings emerge, thin them out to maintain a density of about 15 plants per pot. S4. Prepare NTA aqueous solution with a concentration of 0.33 mmol / L and EDDS aqueous solution with a concentration of 0.33 mmol / L respectively. On the 30th day of the growth of Bidens trifoliata, apply the EDDS and NTA aqueous solutions evenly into the soil by root irrigation. Subsequently, apply them a second and a third time on the 40th and 50th days of the growth of Bidens trifoliata, respectively. The application method is the same as the first treatment each time.

[0030] S5. Harvest the above-ground parts of the Bidens trifoliata around day 60 of its growth and remove the cadmium from the field.

[0031] Example 4

[0032] The preparation of bilayer biodegradable microspheres includes the following steps: S11. Dissolve 12.0g of polyhydroxy fatty acid ester in 88g of dichloromethane to obtain a polymer solution, add 1.56g of citric acid and 0.18g of nicotinamide to it, and stir at high speed to form a uniform suspension; S12. The suspension was added dropwise to a 0.8wt% PVA solution, the solvent was evaporated by stirring at 350 rpm, the suspension was collected by centrifugation, washed, and freeze-dried to obtain core microspheres with a particle size of 150 μm. S13. Disperse 2.8g of humic acid powder in 100mL of water, add KOH to adjust the pH to 8.0, dissolve 1.4g of sodium carboxymethyl cellulose in 100mL of water, heat to dissolve, cool, and then mix the potassium humate solution and sodium carboxymethyl cellulose solution at a volume ratio of 2:1 to obtain a composite coating solution; S14. The core microspheres obtained in S12 are immersed in the composite coating solution in S13, stirred and adsorbed for 25 min, leached out and vacuum dried at 35℃ to obtain bilayer microspheres with a shell thickness of 40 μm.

[0033] A method for enhancing the remediation of cadmium-contaminated soil with Bidens trifoliata using bilayer microspheres in synergistic EDDS / NTA enhancement includes the following steps: S1. Preparation of bilayer biodegradable microspheres; S2. Soak Bidens trifoliata seeds in a 1% NaClO solution for 10 minutes to disinfect them, then rinse them thoroughly with clean water and air dry them for later use. S3. Apply 1.5g of double-layer microspheres per kilogram of soil to cadmium-contaminated soil and sow sterilized Bidens trifoliata seeds. When sowing, scatter the seeds evenly on the soil surface and cover them with about 2cm of Cd-contaminated simulated soil. During the germination period, maintain the temperature at 20±5℃ and the light at 14h per day to promote successful seed germination. After the seedlings emerge, thin them out to maintain a density of about 15 plants per pot. S4. Prepare NTA aqueous solution with a concentration of 0.33 mmol / L and EDDS aqueous solution with a concentration of 0.33 mmol / L respectively. On the 30th day of the growth of Bidens trifoliata, apply the EDDS and NTA aqueous solutions evenly into the soil by root irrigation. Subsequently, apply them a second and a third time on the 40th and 50th days of the growth of Bidens trifoliata, respectively. The application method is the same as the first treatment each time.

[0034] S5. Harvest the above-ground parts of the Bidens trifoliata around day 60 of its growth and remove the cadmium from the field.

[0035] Example 5

[0036] The preparation of bilayer biodegradable microspheres includes the following steps: S11. Dissolve 10.0g of polyhydroxy fatty acid ester in 90g of dichloromethane to obtain a polymer solution, add 1.2g of citric acid and 0.12g of nicotinamide to it, and stir at high speed to form a uniform suspension; S12. The suspension was added dropwise to a 1.2wt% PVA solution, the solvent was evaporated by stirring at 500 rpm, the suspension was collected by centrifugation, washed, and freeze-dried to obtain core microspheres with a particle size of 80 μm. S13. Disperse 2.5g of humic acid powder in 100mL of water, add KOH to adjust the pH to 8.0, dissolve 1.2g of sodium carboxymethyl cellulose in 100mL of water, heat to dissolve, cool, and then mix the potassium humate solution and sodium carboxymethyl cellulose solution at a volume ratio of 2:1 to obtain a composite coating solution; S14. Immerse the core microspheres obtained in S12 into the composite coating solution in S13, stir and adsorb for 20 min, leach out and vacuum dry at 35℃ to obtain bilayer microspheres with a shell thickness of 30 μm.

[0037] A method for enhancing the remediation of cadmium-contaminated soil with Bidens trifoliata using bilayer microspheres in synergistic EDDS / NTA enhancement includes the following steps: S1. Preparation of bilayer biodegradable microspheres; S2. Soak Bidens trifoliata seeds in a 1% NaClO solution for 10 minutes to disinfect them, then rinse them thoroughly with clean water and air dry them for later use. S3. Apply 1.5g of double-layer microspheres per kilogram of soil to cadmium-contaminated soil and sow sterilized Bidens trifoliata seeds. When sowing, scatter the seeds evenly on the soil surface and cover them with about 2cm of Cd-contaminated simulated soil. During the germination period, maintain the temperature at 20±5℃ and the light at 14h per day to promote successful seed germination. After the seedlings emerge, thin them out to maintain a density of about 15 plants per pot. S4. Prepare NTA aqueous solution with a concentration of 0.33 mmol / L and EDDS aqueous solution with a concentration of 0.33 mmol / L respectively. On the 30th day of the growth of Bidens trifoliata, apply the EDDS and NTA aqueous solutions evenly into the soil by root irrigation. Subsequently, apply them a second and a third time on the 40th and 50th days of the growth of Bidens trifoliata, respectively. The application method is the same as the first treatment each time.

[0038] S5. Harvest the above-ground parts of the Bidens trifoliata around day 60 of its growth and remove the cadmium from the field.

[0039] Comparative Example 1 The difference between Comparative Example 1 and Example 5 is that the soil used in Comparative Example 1 is uncontaminated soil, and Bidens trifoliata seeds are directly sown on the uncontaminated soil without applying double-layer biodegradable microspheres or EDDS and NTA chelating agents.

[0040] Comparative Example 2 The difference between Comparative Example 2 and Example 5 is that Comparative Example 2 did not apply double-layer biodegradable microspheres or EDDS and NTA chelating agents when sowing Bidens trifoliata seeds.

[0041] Comparative Example 3 The difference between Comparative Example 3 and Example 5 is that Comparative Example 3 only applied EDDS (1 mmol / L) once on the 30th day of growth, without applying NTA.

[0042] Comparative Example 4 The difference between Comparative Example 4 and Example 5 is that in the Comparative Example, 0.33 mmol / L EDDS aqueous solution was evenly applied to the soil by root irrigation on the 30th, 40th and 50th days of growth of Bidens trifoliata, without the application of NTA.

[0043] Comparative Example 5 The difference between Comparative Example 5 and Example 5 is that Comparative Example 5 only applied NTA (1 mmol / L) once on the 30th day of growth, and did not apply EDDS.

[0044] Comparative Example 6 The difference between Comparative Example 6 and Example 5 is that Comparative Example 6 applied a 0.33 mmol / L NTA aqueous solution evenly into the soil by root irrigation on the 30th, 40th and 50th days of growth of Bidens trifoliata, without applying EDDS.

[0045] Comparative Example 7 The difference between Comparative Example 7 and Example 5 is that Comparative Example 7 applied 1.5g of double-layer microspheres per kilogram of soil to cadmium-contaminated soil before sowing Bidens trifoliata seeds, and did not apply EDDS and NTA during growth.

[0046] Comparative Example 8 The difference between Comparative Example 8 and Example 5 is that citric acid and nicotinamide are not added in step S11 of preparing bilayer biodegradable microspheres in Comparative Example 8, while the remaining steps are the same as in Example 5.

[0047] Comparative Example 9 The difference between Comparative Example 9 and Example 5 is that in step S13 of preparing bilayer biodegradable microspheres in Comparative Example 9, humic acid is not added, and only sodium carboxymethyl cellulose is used as the coating solution. The remaining steps are the same as in Example 5.

[0048] Figure 1 This is a schematic diagram illustrating the principle of phytoremediation technology.

[0049] Test Example 1: Samples were taken from each group on the 60th day of seedling growth. Soil samples were air-dried, ground, and sieved, and then pretreated with acid digestion. The Cd content in the digestion solution was determined by atomic absorption spectrophotometry. The remediation efficiency was calculated based on the total Cd content of the soil before and after remediation. Remediation efficiency (%) = (total Cd in the soil before remediation - total Cd in the soil after remediation) / total Cd in the soil before remediation × 100%.

[0050] Table 1 Repair Results

[0051] Referring to Table 1, the remediation efficiency of the embodiments of the present invention is significantly higher than that of the comparative examples. This is because the continuous release of humic acid from the outer shell of the microspheres promotes the root development and aboveground biomass accumulation of *Bidens trifoliata*; the citric acid in the core converts insoluble cadmium in the soil into an exchangeable form, providing a more easily complexed substrate for subsequent chelating agents; nicotinamide enhances the efficiency of cadmium translocation from the roots to the aboveground parts; and the applied EDDS and NTA further chelate the activated cadmium into a water-soluble complex, which is efficiently absorbed by the plants. These three factors synergistically achieve efficient cadmium removal. Comparative Example 1 is a cadmium-free blank control with no remediation target. Comparative Example 2 did not apply microspheres or chelating agents, relying solely on natural absorption by *Bidens trifoliata*. Cadmium in the soil mainly exists in low-availability forms such as carbonate-bound forms, and plant biomass is inhibited by cadmium stress, resulting in the lowest remediation efficiency. Comparative Examples 3 and 4 involved the application of EDDS without NTA. Comparative Example 3 was a single application. While EDDS could activate some carbonate-bound cadmium, it lacked the synergistic activation effect of NTA on iron-manganese oxide-bound cadmium. Furthermore, without microsphere pre-activation and growth-promoting assistance, the remediation efficiency was low. Comparative Example 4, with EDDS applied in three stages, showed slight improvement compared to a single application, but still lagged behind the combined use of EDDS and NTA. Comparative Examples 5 and 6 involved the application of NTA without EDDS. NTA's ability to activate cadmium alone was weaker than EDDS, especially in the conversion of carbonate-bound cadmium, where the effect was not significant. Even with three stages of application, the remediation efficiency of Comparative Example 6 was significantly lower than the EDDS+NTA combined group. Comparative Example 7, although using intact microspheres, did not apply any chelating agent. Pre-activation relied solely on the humic acid and citric acid released by the microspheres, lacking the potent chelation extraction effect of EDDS / NTA, resulting in a failure to continuously improve the bioavailability of cadmium in the mid-to-late stages. In Comparative Example 8, the microsphere core lacked citric acid and nicotinamide, thus lacking pre-activation and transport-promoting functions. When EDDS / NTA directly acted on insoluble cadmium, the complexation efficiency decreased, and cadmium transport to the aboveground parts was hindered, resulting in a significantly lower remediation efficiency compared to the intact microsphere examples. In Comparative Example 9, the microsphere shell lacked humic acid, thus lacking the functions of promoting root growth and enhancing plant stress resistance. The Bidens trifoliata had a lower biomass, weakened overall cadmium absorption capacity, and a significantly lower remediation efficiency compared to the examples.

[0052] Test Example 2: Samples were taken from Bidens trifoliata on the 60th day of growth. The entire plant was removed from the pot and the roots and above-ground parts were repeatedly rinsed with deionized water to remove soil particles and dust adsorbed on the surface. After absorbing the surface moisture with filter paper, the plant was divided into above-ground and underground parts. The fresh weight of the above-ground and underground parts was then weighed using an analytical balance. The parts were then dried in an oven at 105°C until constant weight was achieved. The dry weight of the above-ground and underground parts was then weighed separately.

[0053] Reference Figure 2 As shown, the *Bidens trifoliata* plant in this embodiment exhibited the highest biomass. This was attributed to the continuous release of humic acid from the microsphere shell, whose active functional groups promoted root elongation and lateral root proliferation, thus expanding the absorption surface area. Simultaneously, potassium ions released from potassium humate enhanced the plant's resistance to cadmium stress, increasing photosynthetic pigment content and antioxidant enzyme activity. Furthermore, citric acid and nicotinamide released from the core, along with the fractionally applied EDDS / NTA, mitigated cadmium toxicity, collectively promoting biomass accumulation. Comparative Example 1 was a cadmium-free blank control; plant growth was unaffected by any stress, resulting in high biomass, but without remediation significance. Comparative Example 2, subjected only to cadmium stress without any enhancement measures, showed a significant decrease in plant biomass, demonstrating the strong inhibitory effect of cadmium on growth. Comparative Examples 3 and 4, treated only with EDDS, partially alleviated cadmium stress due to the chelating effect of EDDS, resulting in increased biomass compared to Comparative Example 2, but lacked the synergistic effect of NTA and the continuous growth-promoting effect of the microspheres. Comparative Examples 5 and 6 only applied NTA. NTA's chelating ability for cadmium is weaker than EDDS, resulting in limited stress relief and a small increase in biomass. Comparative Example 7, although using intact microspheres, did not apply any chelating agent, lacking the strong chelating effect of EDDS / NTA. Cadmium accumulated in the rhizosphere, causing root toxicity and hindering growth. In Comparative Example 8, the microsphere core lacked citric acid and nicotinamide, missing the acidifying and chelating pre-activation effects of citric acid on the rhizosphere microenvironment, and also lacking the regulatory effect of nicotinamide on plant metabolism. Cadmium toxicity was not effectively alleviated, and biomass decreased. In Comparative Example 9, the microsphere shell lacked humic acid. The lack of humic acid's root-promoting effect and potassium ion stress-inducing effect led to reduced biomass. These data fully demonstrate that this invention, through the three-layer synergy of humic acid in the microsphere shell, citric acid and nicotinamide in the core, and the chelating agents EDDS / NTA, can significantly improve the growth biomass of Bidens trifoliata in cadmium-contaminated soil, laying a material foundation for efficient remediation.

[0054] Test Example 3: After harvesting, soil samples were air-dried, ground, and sieved. The Tessier continuous extraction method was used to separate and extract cadmium in different bound forms in the soil step by step. After centrifugation and filtration, the content of each form of cadmium was determined by atomic absorption spectrophotometry or inductively coupled plasma mass spectrometry, and the percentage of each form in the total cadmium was calculated.

[0055] Reference Figure 3 As shown, the proportion of exchangeable cadmium in the soil of this embodiment of the invention is significantly increased, while the proportion of insoluble cadmium such as residual, carbonate-bound, and iron-manganese oxide-bound cadmium is significantly reduced. This is because the citric acid released from the core of the microspheres pre-converts some of the insoluble cadmium into the exchangeable form. Subsequently, the EDDS and NTA applied in stages further convert non-exchangeable cadmium such as carbonate-bound and iron-manganese-bound cadmium into water-soluble chelates through complexation, thereby continuously improving the bioavailability of cadmium. Comparative Example 2 had no treatment, and cadmium mainly existed in low-availability forms such as residual cadmium. Comparative Examples 3-6 only applied EDDS or NTA, which had a certain activation effect, but the cadmium form conversion was incomplete, and the increase in exchangeable cadmium was limited; among them, only the EDDS treatment had a certain effect on carbonate-bound cadmium, while the NTA treatment was weaker. Comparative Example 7 only applied microspheres without chelating agents, relying solely on citric acid pre-activation, lacking subsequent strong chelation, and the increase in exchangeable cadmium was insufficient. Comparative Example 8 did not contain citric acid and nicotinamide in the core, and EDDS / NTA directly acted on the insoluble cadmium, resulting in a decrease in activation efficiency. Comparative Example 9 showed that the plant's outer shell lacked humic acid, resulting in poor plant growth and weak root secretion and absorption capacity, which indirectly affected the transformation of cadmium forms.

[0056] Obviously, the above embodiments are merely illustrative examples for clear explanation and are not intended to limit the implementation. Those skilled in the art will recognize that other variations or modifications can be made based on the above description. It is neither necessary nor possible to exhaustively list all possible implementations here. However, obvious variations or modifications derived therefrom are still within the scope of protection of this invention.

Claims

1. A method for synergistically enhancing the remediation of cadmium-contaminated soil with Bidens trifoliata using bilayer microspheres and EDDS / NTA, characterized in that, Includes the following steps: S1. Prepare bilayer biodegradable microspheres, the microspheres comprising a shell and a core, the shell comprising humic acid, and the core comprising polyhydroxyalkanoate and citric acid and nicotinamide loaded thereon; S2. Soak the seeds of Bidens trifoliata in NaClO solution for disinfection, then rinse thoroughly with clean water and air dry for later use; S3. Apply the double-layer biodegradable microspheres obtained in step S1 into cadmium-contaminated soil and sow disinfected Bidens trifoliata seeds. S4. Apply EDDS and NTA aqueous solutions to the soil in several applications between the 30th and 50th day of growth of Bidens trifoliata; S5. Harvest the above-ground parts of the Bidens trifoliata around day 60 of its growth and remove the cadmium from the field.

2. The method for remediating cadmium-contaminated soil with Bidens trifoliata using bilayer microspheres in synergistic EDDS / NTA enhancement according to claim 1, characterized in that, The method for preparing the bilayer biodegradable microspheres includes the following steps: S11. Dissolve polyhydroxy fatty acid ester in dichloromethane to obtain a polymer solution, add citric acid and nicotinamide to it, and stir at high speed to form a uniform suspension; S12. The suspension was added dropwise to the PVA solution, the solvent was evaporated by stirring, the suspension was collected by centrifugation, washed, and freeze-dried to obtain core microspheres; S13. Disperse humic acid powder in water, add KOH to adjust the pH to 8.0-9.0 to obtain potassium humate solution, dissolve sodium carboxymethyl cellulose in water, heat to dissolve, cool and mix with potassium humate solution to obtain composite coating solution; S14. Immerse the core microspheres obtained in S12 into the composite coating solution in S13, stir to adsorb, leach, and then vacuum dry to obtain bilayer microspheres.

3. The method for remediating cadmium-contaminated soil with Bidens trifoliata using bilayer microspheres in synergistic EDDS / NTA enhancement according to claim 2, characterized in that, In step S11, the concentration of the polymer solution is 5-15 wt%, the amount of citric acid added is 10-15 wt% of the dry weight of the polymer, and the amount of nicotinamide added is 0.5-2.0 wt% of the dry weight of the polymer.

4. The method for remediating cadmium-contaminated soil with Bidens trifoliata using bilayer microspheres in synergistic EDDS / NTA enhancement according to claim 2, characterized in that, In step S12, the concentration of the PVA solution is 0.5~2wt%, the stirring speed for evaporating the solvent is 300~600rpm, and the particle size of the core microspheres is 50~200μm.

5. The method for remediating cadmium-contaminated soil with Bidens trifoliata using bilayer microspheres in synergistic EDDS / NTA enhancement according to claim 2, characterized in that, In step S13, the concentration of potassium humate solution is 2-3 wt%, the concentration of sodium carboxymethyl cellulose solution is 1-1.5 wt%, and the volume ratio of potassium humate solution to sodium carboxymethyl cellulose solution after mixing is 2:

1.

6. The method for remediating cadmium-contaminated soil with Bidens trifoliata using bilayer microspheres in synergistic EDDS / NTA enhancement according to claim 2, characterized in that, In step S14, the impregnation and adsorption time is 10–30 min, the vacuum drying temperature is 30–40 °C, and the drying is carried out to constant weight. The outer shell thickness of the bilayer microspheres is 20–50 μm.

7. The method for remediating cadmium-contaminated soil with Bidens trifoliata using bilayer microspheres in synergistic EDDS / NTA enhancement according to claim 1, characterized in that, In step S3, the amount of double-layer biodegradable microspheres applied is 1-5g per kilogram of soil.

8. The method for remediating cadmium-contaminated soil with Bidens trifoliata using bilayer microspheres in synergistic EDDS / NTA enhancement according to claim 1, characterized in that, In step S4, EDDS and NTA are applied three times, with an interval of 10 days between each application. The total volume of EDDS and NTA solution applied each time is 1 L / m³. 2 The concentrations were all 0.2~2 mmol / L.

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