Nitrogen-doped bamboo-based carbon quantum dots and preparation method and application thereof

Abstract

The application belongs to the technical field of agricultural fertilizer and waste resource utilization, and particularly relates to nitrogen-doped bamboo base carbon quantum dots, a preparation method and application thereof. The application provides a preparation method of nitrogen-doped bamboo base carbon quantum dots, which comprises the following steps: mixing bamboo, a nitrogen source and water, and performing hydrothermal reaction to obtain the nitrogen-doped bamboo base carbon quantum dots. The preparation method provided by the application uses agricultural waste bamboo as a single carbon source, uses urea as a nitrogen source, and prepares functionalized nitrogen-doped bamboo base carbon quantum dots through one-step hydrothermal in-situ doping. The method does not need to add strong acid, strong base or expensive chemical reagents, and the process is simple and green. The method not only effectively solves the problem of resource utilization of bamboo waste, but also greatly reduces the preparation cost of carbon quantum dots, so that the carbon quantum dots meet the demand of large-scale agricultural application.

Description

Technical Field

[0001] This invention belongs to the field of agricultural fertilizer and waste resource utilization technology, specifically relating to a nitrogen-doped reed-based carbon quantum dot, its preparation method and application. Background Technology

[0002] Microalgae, as single-celled photosynthetic microorganisms, are rich in proteins, amino acids, plant hormones, and various trace elements, making them a high-quality raw material for preparing bio-fertilizers. However, the large-scale application of microalgae bio-fertilizers is still limited by their production efficiency and cost. The natural proliferation rate of microalgae is mainly limited by light energy conversion efficiency. During large-scale propagation, as the density of algal cells increases, a severe "self-shading effect" occurs within the culture system. This leads to a significant reduction in the penetration depth of light in the culture medium, hindering photosynthesis in deeper algal cells due to insufficient light exposure, thus significantly slowing down the accumulation rate of biomass. This bottleneck forces an extension of the cultivation cycle, significantly increasing the energy consumption and time costs of lighting and temperature control equipment.

[0003] Carbon quantum dots, as an emerging zero-dimensional carbon nanomaterial, possess excellent water solubility, low toxicity, and unique optical properties (such as upconversion luminescence). Studies have shown that carbon quantum dots can enhance plant photosynthesis by improving photosynthetic electron transport, thereby promoting seed germination and root growth. However, the preparation of high-performance carbon quantum dots currently relies heavily on chemical reagents such as citric acid or expensive carbon sources, and the processes are complex and costly, making it difficult to meet the needs of low-cost, large-scale application in agriculture.

[0004] Currently, there are reports on using carbon quantum dots or microalgae to promote plant growth. For example, CN120964777A discloses the preparation of carbon quantum dots using citric acid / formamide as precursors to promote microalgae growth. However, the cost of carbon quantum dots promoting microalgae growth is relatively high, mainly due to the use of chemical reagents (citric acid / formamide) as raw materials. Therefore, there is an urgent need for a simple and low-cost carbon quantum dot preparation process. Summary of the Invention

[0005] The purpose of this invention is to provide a nitrogen-doped arisaema-based carbon quantum dot, its preparation method, and its application. The preparation method of the nitrogen-doped arisaema-based carbon quantum dot of this invention is simple and uses arisaema as raw material, which can reduce costs and enable large-scale application.

[0006] This invention provides a method for preparing nitrogen-doped arundinaceous carbon quantum dots, comprising the following steps: mixing arundinaceous plant, a nitrogen source, and water, and carrying out a hydrothermal reaction to obtain the nitrogen-doped arundinaceous carbon quantum dots;

[0007] The nitrogen source includes urea and / or ethylenediamine.

[0008] As a preferred embodiment, the mass ratio of the reed to the nitrogen source is 1:0.1 to 1:0.5.

[0009] As a preferred embodiment, the temperature of the hydrothermal reaction is 150~220℃; the time of the hydrothermal reaction is 6~8 h.

[0010] The present invention also provides nitrogen-doped arundinaceous carbon quantum dots prepared by the preparation method described above.

[0011] The present invention also provides applications of nitrogen-doped arundinaceous carbon quantum dots prepared by the preparation method described above, the applications including at least one of the following: (1) improving the photosynthetic efficiency of microalgae; (2) promoting the growth of microalgae; (3) increasing the production of IAA by microalgae.

[0012] The present invention also provides a compound fertilizer comprising nitrogen-doped arundinaceous carbon quantum dots and microalgae prepared by the preparation method described above; the nitrogen-doped arundinaceous carbon quantum dots are anchored on the surface of microalgal cells to form a carbon quantum dot-microalgae artificial photosynthetic hybrid system.

[0013] As a preferred embodiment, the microalgae concentration in the compound fertilizer is ≥10. 6 cell / mL.

[0014] The present invention also provides a method for preparing the compound fertilizer described above, comprising the following steps: inoculating microalgae into a microalgae culture medium for co-cultivation to obtain the compound fertilizer;

[0015] The microalgae culture medium contains nitrogen-doped arundinaceous carbon quantum dots prepared by the preparation method described above.

[0016] As a preferred embodiment, the content of nitrogen-doped arundinaceous carbon quantum dots in the microalgae culture medium is 0.1~1 mg / L.

[0017] The present invention also provides the application of the compound fertilizer described in the above scheme or the compound fertilizer prepared by the above preparation method in promoting plant growth.

[0018] Beneficial Effects: This invention provides a method for preparing nitrogen-doped reed-based carbon quantum dots, comprising the following steps: mixing reed, a nitrogen source, and water, and carrying out a hydrothermal reaction to obtain the nitrogen-doped reed-based carbon quantum dots; the nitrogen source includes urea and / or ethylenediamine. The preparation method of this invention uses agricultural waste reed as a single carbon source and urea and / or ethylenediamine as a nitrogen source, and prepares functionalized nitrogen-doped reed-based carbon quantum dots through a one-step hydrothermal in-situ doping process. This method does not require the addition of strong acids, strong bases, or expensive chemical reagents, and the process is simple and environmentally friendly. This not only effectively solves the problem of resource utilization of reed waste but also significantly reduces the preparation cost of carbon quantum dots, making it suitable for large-scale agricultural applications.

[0019] The present invention also provides nitrogen-doped arundinaceous carbon quantum dots (N-CQDs) prepared by the above preparation method. The nitrogen-doped arundinaceous carbon quantum dots have a surface rich in amino functional groups and have excellent water solubility and optical properties, such as upconversion luminescence characteristics, which can convert low-energy long-wavelength light (such as near-infrared) into high-energy short-wavelength light (such as visible / ultraviolet), thereby providing usable light energy for plant photosynthesis.

[0020] This invention also provides applications of the nitrogen-doped arundinaceous carbon quantum dots prepared by the aforementioned method, including at least one of the following applications: (1) improving the photosynthetic efficiency of microalgae; (2) promoting microalgae growth; and (3) increasing the production of IAA by microalgae. This invention utilizes the upconversion fluorescence characteristics and surface charge effect of N-CQDs, and by introducing them into the microalgae culture system, it can effectively improve the light energy transfer efficiency in the culture medium and promote electron transfer in microalgae photosynthesis. Experimental results show that N-CQDs can significantly shorten the growth lag phase of microalgae, enabling them to reach the growth plateau phase more quickly, thereby solving the industry pain points of long production cycles and low efficiency of microalgae biofertilizers. Simultaneously, N-CQDs can also induce microalgae to secrete endogenous plant hormones (IAA).

[0021] This invention also provides a compound fertilizer comprising nitrogen-doped arundinaceous carbon quantum dots and microalgae; the positive charge on the surface of the nitrogen-doped arundinaceous carbon quantum dots can electrostatically attract the negative charge of the microalgae cell wall, and the nitrogen-doped arundinaceous carbon quantum dots are anchored to the surface of the microalgae cells using electrostatic self-assembly technology to construct an artificial photosynthetic hybrid system of "nanomaterials-microalgae", which can realize a cascade enhancement mechanism at the material-cell level.

[0022] This invention also provides a method for preparing a compound fertilizer, comprising the following steps: inoculating microalgae into a microalgae culture medium for co-cultivation to obtain the compound fertilizer; wherein the microalgae culture medium contains nitrogen-doped arundinaceae-based carbon quantum dots. This invention adds N-CQDs to the microalgae culture medium, utilizing the electrostatic attraction between the positive charge on the surface of the N-CQDs and the negative charge of the Scenedessella cell wall, anchoring them to the surface of the microalgae cells. N-CQDs can act as photosynthetic promoters, inducing enhanced metabolism in microalgae cells and the secretion of endogenous auxins and extracellular polysaccharides, constructing a bioactive system with living microalgae as the core carrier and carbon quantum dots as functional ligands. Furthermore, no solid-liquid separation is required; the culture medium obtained containing highly active microalgae cells and their metabolites is the compound fertilizer.

[0023] This invention also provides the application of compound fertilizers in promoting plant growth. The compound fertilizer of this invention is a live microbial preparation in which microalgal cells maintain physiological activity, and carbon quantum dots are anchored to the microalgal surface through interfacial assembly, forming an active complex with continuous photosynthetic carbon fixation and rhizosphere colonization capabilities. Applying it to the soil can significantly improve the rhizosphere microecology, promote plant growth, and increase yield. The compound fertilizer of this invention achieves high-value utilization of agricultural waste and provides a new approach for developing low-cost, novel green bio-fertilizers with metabolic regulation functions. Attached Figure Description

[0024] To more clearly illustrate the technical solutions in the embodiments of the present invention or the prior art, the accompanying drawings used in the embodiments will be briefly described below.

[0025] Figure 1 The fluorescence spectrum of nitrogen-doped arundinaceous carbon quantum dots;

[0026] Figure 2 The growth curve of *Scenedesmus* over 21 days in Application Example 2 is shown; where A is the *Scenedesmus* group; B is the *Scenedesmus* + *Phyllostachys edulis* carbon quantum dot group; and C is the *Scenedesmus* + undoped *Phyllostachys edulis* carbon quantum dot group.

[0027] Figure 3 The amount of IAA produced by Scenedesmus in Application Example 2;

[0028] Figure 4 This is a growth diagram of the chili pepper used in Example 3;

[0029] Figure 5 The graph shows the results of pepper plant height in each treatment group in Example 3.

[0030] Figure 6 The graph shows the results of pepper stem diameter in each treatment group in Example 3.

[0031] Figure 7 The SPAD results of peppers in each treatment group in Application Example 3 are shown in the figure.

[0032] Figure 8 The graph shows the results of root activity of peppers in each treatment group in Example 3.

[0033] In the figure, ns indicates that the data show no significant difference; express p <0.05; express p <0.01; express p <0.001. Detailed Implementation

[0034] This invention provides a method for preparing nitrogen-doped arundinaceous carbon quantum dots, comprising the following steps: mixing arundinaceous plant, a nitrogen source, and water, and carrying out a hydrothermal reaction to obtain the nitrogen-doped arundinaceous carbon quantum dots;

[0035] The nitrogen source includes urea and / or ethylenediamine.

[0036] This invention mixes *Arundinaria lobata* with a nitrogen source to obtain a reaction precursor. Currently, the preparation of high-performance carbon quantum dots largely relies on chemical reagents such as citric acid or expensive carbon sources, and the processes are complex and costly, making it difficult to meet the needs of low-cost, large-scale application in agriculture. Furthermore, *Arundinaria lobata*, a fast-growing, high-biomass perennial grass, is currently mostly considered a weed or used only for combustion power generation, and its high-value-added utilization pathways have not been effectively developed. As one implementation method, the nitrogen source includes urea and / or ethylenediamine. This invention uses biomass *Arundinaria lobata* as the carbon source and urea and / or ethylenediamine as the nitrogen source, and through a one-step hydrothermal in-situ doping process, functionalized nitrogen-doped *Arundinaria lobata*-based carbon quantum dots (N-CQDs) can be prepared. The method described in this invention can effectively alleviate the problems of high raw material costs and low resource utilization in traditional carbon quantum dot preparation processes, achieving high-value utilization of agricultural waste.

[0037] In one embodiment, the reed includes reed powder; the preparation of the reed powder includes: drying and pulverizing the reed to obtain reed powder; this invention does not specifically limit the source of the reed, nor does it specifically limit the part of the reed used. In a specific embodiment of this invention, the reed can be the whole reed plant, or any part of the reed, such as roots, stems, or leaves. In one embodiment, the drying temperature is 60℃; the pulverization method includes grinding; after grinding, the ground reed is further passed through a 25-60 mesh sieve to obtain the undersize material, which is the reed powder. This invention uses reed powder for hydrothermal reaction, which can increase the specific surface area of ​​the raw materials, improve the reaction contact area, and enhance the preparation efficiency of carbon quantum dots.

[0038] In one embodiment, the mass ratio of *Arundinaria lobata* to the nitrogen source is 1:0.1 to 1:0.5. In specific embodiments of the present invention, the mass ratio of *Arundinaria lobata* to the nitrogen source can be any ratio within the range of 1:0.1 to 1:0.5, such as 1:0.1, 1:0.2, 1:0.3, 1:0.4, or 1:0.5. The present invention uses biomass *Arundinaria lobata* as the carbon source and urea and / or ethylenediamine as the nitrogen source to prepare functionalized nitrogen-doped *Arundinaria lobata*-based carbon quantum dots via a one-step hydrothermal in-situ doping method. Urea, as the nitrogen source, has the advantages of high nitrogen content, good water solubility, and low cost and availability, enabling efficient in-situ nitrogen doping during the one-step hydrothermal process. The present invention limits the ratio of *Arundinaria lobata* to the nitrogen source, which ensures the introduction of appropriate and structurally stable nitrogen-containing functional groups (such as amino, pyridine nitrogen, pyrrole nitrogen, etc.) into the carbon framework of the carbon quantum dots, significantly improving not only the water solubility, surface charge density, and active sites of the carbon quantum dots, but also optimizing their optical and electronic transport properties. When used as a component of compound fertilizers, these carbon quantum dots, rich in surface functional groups, can better synergize and chelate with soil nutrients or other components in fertilizers (such as microalgae), promoting the transport and absorption of nutrients within plants, thereby effectively improving the photosynthetic efficiency and stress resistance of crops. If the proportion of nitrogen source (urea and / or ethylenediamine) added is too low, the nitrogen doping degree of the carbon quantum dots will be insufficient, limiting the number of surface hydrophilic functional groups (such as amino groups). This will not only reduce the water dispersibility and stability of the carbon quantum dots but also weaken their chelating ability for mineral elements in fertilizers, leading to a decrease in their physiological activity and preventing them from fully exerting their stimulating and growth-promoting effects on plants. If the proportion of nitrogen source added is too high: excessive nitrogen will damage the graphitized framework of the carbon quantum dot core, resulting in too many carbon structural defects and even generating non-target byproducts (such as polymerized carbon nitride impurities), reducing the purity and yield of the carbon quantum dots. In addition, excessively high concentrations of nitrogen residues may also cause local osmotic pressure stress on plant roots or symbiotic micro-ecosystems, which is not conducive to their subsequent application as a safe and efficient fertilizer.

[0039] After obtaining the reaction precursor, it is mixed with water to obtain a reaction precursor solution. As one embodiment, the mass-to-volume ratio of the reaction precursor to water is 1 g:5 mL to 1 g:10 mL. In specific embodiments of the present invention, the mass-to-volume ratio of the reaction precursor to water can be any ratio within the range of 1 g:5 mL to 1 g:10 mL, for example, 1 g:5 mL, 1 g:6 mL, 1 g:7 mL, 1 g:8 mL, 1 g:9 mL, or 1 g:10 mL. The suitable mass-to-volume ratio range of the reaction precursor to water in this invention provides a better heat and mass transfer medium environment for the hydrothermal reaction, ensuring the complete hydrolysis of macromolecules such as cellulose and hemicellulose in the reed powder, as well as the complete dissolution of urea. More importantly, this ratio ensures that the concentration of the active intermediates generated by dehydration condensation in the reaction system is within the ideal "supersaturation" range, perfectly meeting the thermodynamic conditions for uniform nucleation and controlled growth of carbon quantum dots. This allows for the high-yield production of carbon quantum dots with small particle sizes (typically below 10 nm), uniform size distribution, and excellent water solubility. These uniformly sized nanomaterials, when subsequently applied as fertilizer, more easily penetrate plant cell walls, efficiently promoting growth and nutrient transfer. However, excessive water addition leads to over-dilution of the precursor concentration in the reactor, significantly weakening the reaction kinetics. On one hand, the probability of active intermediates colliding and nucleating at low concentrations drops sharply, resulting in extremely low carbon quantum dot yields. On the other hand, processing large amounts of water requires more heating energy, not only wasting water resources but also significantly increasing the energy consumption and time cost of a single hydrothermal reaction, severely deviating from the original intention of low-cost, large-scale agricultural production. If the amount of water added is too small: on the one hand, the viscosity of the reaction system will be too high, leading to uneven material dispersion and localized mass transfer obstruction, which can easily cause localized overheating in the reactor; on the other hand, excessively high intermediate concentrations can easily disrupt the stable growth mechanism of nanoparticles, causing severe overpolymerization and cross-linking carbonization, resulting in the rapid aggregation of the generated carbon nanoparticles, ultimately forming large, blocky carbonaceous precipitates (hydrothermal carbon) instead of the target nanoscale carbon quantum dots, thus completely losing the unique nanoscale size effect and high physiological activity of carbon quantum dots. In addition, the high solids content of the reaction system under high-temperature, closed conditions produces a large amount of gas, which can easily lead to excessive pressure in the hydrothermal reactor, posing an uncontrollable safety hazard.

[0040] The precursor solution is subjected to a hydrothermal reaction to obtain the hydrothermal reaction product. As one embodiment, the hydrothermal reaction temperature is 150-220°C; in a specific embodiment of the invention, the hydrothermal reaction temperature can be any value within the range of 150-220°C, such as 150, 160, 170, 180, 190, 200, 210, or 220°C. As one embodiment, the hydrothermal reaction time is 6-8 h; in a specific embodiment of the invention, the hydrothermal reaction time can be any value within the range of 6-8 h, such as 6, 7, or 8 h. This invention performs a one-step hydrothermal carbonization reaction, utilizing the high temperature and high pressure environment during the hydrothermal process to achieve in-situ doping of nitrogen in the carbon framework. Using the aforementioned reaction temperature of 150-220°C and reaction time of 6-8 h, under these thermodynamic conditions, this invention can further provide precise activation energy for the dehydration, cleavage, and aromatization of macromolecules such as cellulose and lignin in *Arundo donax* powder, while simultaneously promoting the moderate decomposition of the nitrogen source to provide sufficient active nitrogen, thus achieving efficient in-situ nitrogen doping. Carbon quantum dots synthesized under this process window not only possess highly regular carbon cores and uniform nanoscale dimensions, but also retain abundant and structurally stable hydrophilic functional groups (such as carboxyl, hydroxyl, and amino groups) on their surface. This excellent surface chemical property endows carbon quantum dots with excellent water solubility and biocompatibility, enabling them to efficiently chelate soil nutrients when used as fertilizer components. Furthermore, they exhibit significant synergistic effects with microalgae and other biological agents, thereby greatly enhancing root vitality and stress resistance in crops such as chili peppers in complex environments like saline-alkali soils. However, if the hydrothermal reaction temperature is too low (below 150℃) or the time is too short (less than 6 h), the energy within the reaction system is insufficient to effectively break the chemical bonds of biomass macromolecules, resulting in incomplete carbonization of the reed powder. Simultaneously, the nitrogen source cannot fully decompose and participate in the doping reaction. This leads to products consisting mostly of macromolecular polymer fragments or unreacted intermediates, resulting in extremely low carbon quantum dot yields and a lack of sufficient surface active sites, rendering them ineffective in nutrient delivery and plant physiological stimulation during agricultural application. If the hydrothermal reaction temperature is too high (above 220℃) or the reaction time is too long (greater than 8 hours), the continuous excessive heat input will induce deep carbonization and excessive cross-linking, causing the valuable oxygen- and nitrogen-containing functional groups on the surface of carbon quantum dots to undergo thermal desorption and detachment. This not only severely damages its water solubility, causing nanoparticles to agglomerate into large-sized hydrothermal carbon particles with no biological activity, but also completely destroys its ability to coordinate with other effective components in fertilizers. In addition, excessively harsh reaction conditions will produce a large amount of by-product gases, leading to a sharp increase in pressure inside the reactor, posing a safety hazard to the equipment, and the significantly increased energy consumption also violates the original intention of low-cost, large-scale production of agricultural fertilizers. The method described in this invention does not require the addition of strong acids, strong alkalis, or expensive chemical reagents, and the process is simple and green; this not only effectively solves the problem of resource utilization of reed waste, but also significantly reduces the preparation cost of carbon quantum dots, making it meet the needs of large-scale agricultural applications.

[0041] After obtaining the hydrothermal reaction product, the process further includes separation and purification. As one embodiment, the separation includes centrifuging the hydrothermal reaction product to obtain a supernatant liquid. As one embodiment, the centrifugation speed is 8000 rpm; the centrifugation time is 10 min. The centrifugation of this invention can remove large particulate residues. As one embodiment, the purification includes placing the supernatant liquid in a dialysis bag for dialysis purification to obtain purified nitrogen-doped arundinaceous carbon quantum dots; the molecular weight cutoff of the dialysis bag is 500 Da; the dialysis time is 1 day. The purified nitrogen-doped arundinaceous carbon quantum dots of this invention are substances with a molecular weight > 500 Da in the dialysis bag; the purification can remove unreacted precursors and small molecule byproducts. Obtaining the purified nitrogen-doped arundinaceous carbon quantum dots further includes lyophilization. As one embodiment, the lyophilization temperature is -65℃.

[0042] This invention also provides nitrogen-doped arundinaceous carbon quantum dots prepared by the method described above. The nitrogen-doped arundinaceous carbon quantum dots of this invention have a surface rich in amino functional groups and possess upconversion properties, enabling them to convert low-energy, long-wavelength light (such as near-infrared) into high-energy, short-wavelength light (such as visible / ultraviolet), thereby providing usable light energy for plant photosynthesis.

[0043] This invention also provides applications of nitrogen-doped arundinaceous carbon quantum dots prepared by the above-described preparation method, the applications including at least one of the following: (1) improving the photosynthetic efficiency of microalgae; (2) promoting microalgae growth; (3) increasing the production of IAA by microalgae. As one embodiment, the microalgae include Scenedesmus. This invention utilizes the upconversion fluorescence characteristics and surface charge effect of nitrogen-doped arundinaceous carbon quantum dots to significantly improve the photosynthetic electron transfer efficiency of microalgae and induce them to secrete endogenous plant hormones. Furthermore, the nitrogen-doped arundinaceous carbon quantum dots of this invention can absorb ultraviolet light that plants cannot absorb and utilize, converting it into light that plants can utilize. The results of the embodiments show that using nitrogen-doped arundinaceous carbon quantum dots can improve the photosynthetic efficiency of microalgae, thereby promoting microalgae growth.

[0044] This invention also provides a compound fertilizer comprising nitrogen-doped *Phyllostachys edulis*-based carbon quantum dots and microalgae prepared by the method described above; the nitrogen-doped *Phyllostachys edulis*-based carbon quantum dots are anchored on the surface of microalgal cells, forming a carbon quantum dot-microalgae artificial photosynthetic hybrid system. The nitrogen-doped *Phyllostachys edulis*-based carbon quantum dots of this invention have a positive charge on their surface, while the microalgae cell wall has a negative charge. The positive charge on the surface of the nitrogen-doped *Phyllostachys edulis*-based carbon quantum dots can electrostatically attract the negative charge of the microalgae cell wall. Using electrostatic self-assembly technology, the nitrogen-doped *Phyllostachys edulis*-based carbon quantum dots are anchored on the surface of microalgae cells, constructing a "nanomaterial-microalgae" (i.e., carbon quantum dot-microalgae) artificial photosynthetic hybrid system.

[0045] The present invention also provides a method for preparing a compound fertilizer, comprising the following steps: inoculating microalgae into a microalgae culture medium for co-cultivation to obtain the compound fertilizer;

[0046] The microalgae culture medium contains nitrogen-doped arundinaceous carbon quantum dots prepared by the preparation method described above.

[0047] In one embodiment, the content of nitrogen-doped arundinaceous carbon quantum dots in the microalgae culture medium is 0.1~1 mg / L. In a specific embodiment of the present invention, the content of nitrogen-doped arundinaceous carbon quantum dots in the microalgae culture medium can be any value within the range of 0.1~1 mg / L, such as 0.1, 0.2, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, or 1 mg / L. The carbon quantum dot content of the present invention is within the optimal physiological window for microalgae to respond to nanomaterial stimulation. At this concentration range, the carbon quantum dots are uniformly monodispersed, which can better exert the role of "micro-antennae" and optimize the light field distribution inside the culture system without causing any nanotoxicity. At the same time, an appropriate amount of carbon quantum dots can moderately stimulate the antioxidant enzyme system of microalgae cells and promote transmembrane electron transfer, thereby maximizing the acceleration of microalgae cell division and significantly increasing the synthesis and accumulation of their extracellular metabolites (such as IAA and other plant growth hormones), laying a material basis for the subsequent preparation of high-efficiency compound fertilizers. If the amount of carbon quantum dots added is too low (less than 0.1 mg / L), the absolute number of carbon quantum dots in the culture system is insufficient, and the resulting light scattering and light frequency conversion effects are too weak to effectively penetrate the deep culture medium, thus failing to substantially solve the "self-shading effect" in the later stages of high-density microalgae culture. On the other hand, at low concentrations, the probability of collision and interaction between carbon quantum dots and microalgae cells is extremely low, failing to trigger the growth-promoting metabolic pathways within the microalgae, resulting in no statistically significant difference in growth-promoting effect compared to the control group without added carbon quantum dots. If the amount of carbon quantum dots added is too high (greater than 1 mg / L), it will produce significant "nanotoxicity" and physical stress on microalgae. First, excessively high concentrations of nanoparticles are prone to agglomeration in the culture medium, not only losing the unique optical activity of carbon quantum dots, but the agglomerates will also directly attach to or encapsulate the surface of algal cells, physically blocking the absorption of light and nutrients, thus exacerbating the shading effect. Secondly, high concentrations of carbon quantum dots induce excessive reactive oxygen species (ROS) production within microalgal cells, triggering severe lipid peroxidation and cell membrane damage, leading to algal cell rupture or premature apoptosis, ultimately resulting in a precipitous decline in biomass and the yield of bioactive metabolites. Furthermore, excessive addition increases unnecessary compound fertilizer production costs. This invention does not specifically limit the type of microalgal culture medium; any conventional microalgal culture medium in the art can be used. In one specific embodiment of this invention, the microalgal culture medium is BG11 medium.

[0048] In one embodiment, the inoculum amount of microalgae is 0.15~0.9 g / L; in a specific embodiment of the present invention, the inoculum amount of microalgae can be any value in the range of 0.15~0.9 g / L, such as 0.15, 0.2, 0.25, 0.3, 0.35, 0.4, 0.45, 0.5, 0.55, 0.6, 0.65, 0.7, 0.75, 0.8, 0.85 or 0.9 g / L.

[0049] In one implementation, the co-culture condition is light culture. In another implementation, the light intensity of the light culture can be any value between 2000 and 5000 lx, for example, 2000, 2500, 3000, 3500, 4000, 4500, or 5000 lx; ​​the light-dark cycle (L:D) of the light culture can be any value between 12 h:12 h and 14 h:10 h, for example, 12 h:12 h, 13 h:11 h, or 14 h:10 h; the culture temperature of the light culture can be any value between 25 and 28°C, for example, 25, 26, 27, or 28°C. In one implementation method, the co-culture time is 10-30 days; in a specific embodiment of the present invention, the co-culture time can be any value within 10-30 days, such as 10, 11, 13, 15, 16, 18, 20, 22, 23, 25, 27, 29, or 30 days. Subsequently, carbon quantum dots are used as photosynthetic promoters to induce enhanced metabolism in microalgal cells and the secretion of endogenous auxin (IAA), constructing a bioactive system with living microalgae as the core carrier and carbon quantum dots as the functional ligand; after the culture is completed, no solid-liquid separation is required, and the culture medium containing highly active microalgal cells and their metabolites obtained is the compound fertilizer.

[0050] As one implementation method, the microalgae concentration in the compound fertilizer is ≥10. 6 The microalgae concentration described in this invention can ensure effective root colonization and niche competition, guarantee the continuous release of growth-promoting metabolites, and maximize the synergistic effect with carbon quantum dots.

[0051] This invention also provides the application of the compound fertilizer prepared by the above-described method in promoting plant growth. Currently, carbon quantum dots or microalgae are often used alone to promote plant growth. Previous studies have found that while carbon quantum dots can promote plant nutrient absorption, the lack of active microorganisms prevents improvement of the self-regulating ability of the rhizosphere microecology; furthermore, single microalgae fertilizers suffer from limited photosynthetic efficiency, short rhizosphere survival period, and slow nutrient release. This invention utilizes nitrogen-doped reed-based carbon quantum dots and microalgae to construct a carbon quantum dot-nano artificial photosynthetic hybrid system. The two work synergistically: nitrogen-doped reed-based carbon quantum dots enhance the photosynthetic efficiency of microalgae through upconversion fluorescence properties, while directly promoting plant nutrient absorption and root development; microalgae can colonize the rhizosphere to optimize the microecology, compensating for the lack of biological regulatory function in carbon quantum dots, resulting in a significantly better growth-promoting effect than single carbon quantum dot or single microalgae fertilizers.

[0052] In one embodiment, the plants include peppers, tomatoes, cauliflower, and corn. In one embodiment, promoting plant growth includes at least one of the following: (1) increasing plant height; (2) increasing stem diameter; (3) increasing SPAD value; and (4) increasing root vitality.

[0053] As one implementation method, the method for promoting plant growth includes: applying the compound fertilizer to the soil; the application rate of the compound fertilizer is 200-500 mL / mu. In a specific embodiment of the present invention, the application rate of the compound fertilizer can be any value within the range of 200-500 mL / mu, such as 200, 220, 230, 250, 270, 290, 300, 350, 380, 400, 420, 450, 460, 480, or 500 mL / mu. As one implementation method, the method further includes: diluting the compound fertilizer before application; the dilution ratio is 10-20 times. In a specific embodiment of the present invention, the dilution ratio can be any value within the range of 10-20 times, such as 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20 times. The present invention's method of diluting the compound fertilizer before application avoids osmotic pressure stress and "seedling burn" phenomena. In one implementation method, the application time is 7-10 days after plant transplanting, for example, 7, 8, 9, or 10 days after transplanting. This invention applies the compound fertilizer to the soil, allowing live microalgae to colonize the plant rhizosphere, continuously improving the rhizosphere microecological environment, and enhancing the growth rate of microalgae through carbon quantum dots. The release of IAA synergistically promotes plant root vitality.

[0054] To further illustrate the present invention, the technical solutions provided by the present invention will be described in detail below with reference to the accompanying drawings and embodiments, but these should not be construed as limiting the scope of protection of the present invention.

[0055] Example 1

[0056] A method for preparing nitrogen-doped arundinaceous carbon quantum dots, comprising the following steps:

[0057] (1) Pretreatment of Reed: Dry Reed at 60°C to constant weight, grind through a 25-mesh sieve to obtain Reed powder.

[0058] (2) Hydrothermal reaction: Mix Reed powder and urea at a mass ratio of 10 g: 1 g, and record it as the reaction precursor. Place it in a hydrothermal reactor. Then add water at a mass volume ratio of 1 g: 10 mL for the reaction precursor and water. Hydrothermal carbonize at 160℃ for 6 hours to obtain the hydrothermal reaction product.

[0059] (3) Separation: The hydrothermal reaction product obtained in step (2) is centrifuged at 8000 rpm for 10 min to separate the upper liquid.

[0060] (4) Purification: The upper liquid obtained in step (3) was dialyzed in a 500 Da dialysis bag for one day, and the substance in the dialysis bag was freeze-dried at -65°C to obtain nitrogen-doped arum-based carbon quantum dot powder.

[0061] Example 2

[0062] A method for preparing nitrogen-doped arundinaceous carbon quantum dots, comprising the following steps:

[0063] (1) Pretreatment of Reed: The operation is the same as step (1) in Example 1.

[0064] (2) Hydrothermal reaction: Mix Reed powder and urea at a mass ratio of 10 g: 5 g, and record it as the reaction precursor. Place it in a hydrothermal reactor. Then add water at a mass volume ratio of 1 g: 10 mL for the reaction precursor and water. Perform hydrothermal carbonization at 180℃ for 8 hours.

[0065] (3) Separation ~ (4) Purification steps are the same as steps (3) ~ (4) in Example 1.

[0066] Example 3

[0067] A method for preparing nitrogen-doped arundinaceous carbon quantum dots, comprising the following steps:

[0068] (1) Pretreatment of Reed: The operation is the same as step (1) in Example 1.

[0069] (2) Hydrothermal reaction: Mix Reed powder and urea at a mass ratio of 10 g: 5 g, and record it as the reaction precursor. Place it in a hydrothermal reactor. Then add water at a mass volume ratio of 1 g: 10 mL for the reaction precursor and water. Hydrothermal carbonize at 200℃ for 8 hours.

[0070] (3) Separation ~ (4) Purification steps are the same as steps (3) ~ (4) in Example 1.

[0071] Comparative Example 1

[0072] A method for preparing aromatic carbon quantum dots, the steps are the same as in Example 1, except that urea is omitted in step (2) to obtain undoped aromatic carbon quantum dots.

[0073] Example 4

[0074] A method for preparing a compound fertilizer, comprising the following steps:

[0075] (1) The nitrogen-doped arundinaceous carbon quantum dots prepared in Example 1 were added to BG11 liquid culture medium at a concentration of 0.5 mg / L to obtain nitrogen-doped arundinaceous carbon quantum dots-BG11 composite liquid culture medium.

[0076] (2) The Scenedesmus was inoculated into the nitrogen-doped Reed-based carbon quantum dot-BG11 composite liquid culture medium in step (1) at a concentration of 0.15 g / L; and cultured in a 27℃ light incubator for 21 days to obtain nitrogen-doped Reed-based carbon quantum dot-Scenedesmus composite fertilizer.

[0077] Example 5

[0078] A method for preparing a compound fertilizer, comprising the following steps:

[0079] (1) The nitrogen-doped arundinaceous carbon quantum dots prepared in Example 2 were added to BG11 liquid culture medium at a concentration of 0.2 mg / L to obtain nitrogen-doped arundinaceous carbon quantum dots-BG11 composite liquid culture medium.

[0080] (2) The Scenedesmus was inoculated into the nitrogen-doped Reed-based carbon quantum dot-BG11 composite liquid culture medium in step (1) at a concentration of 0.9 g / L; and cultured in a 27℃ light incubator for 21 days to obtain nitrogen-doped Reed-based carbon quantum dot-Scenedesmus composite fertilizer.

[0081] Example 6

[0082] A method for preparing a compound fertilizer, comprising the following steps:

[0083] (1) The nitrogen-doped arundinaceous carbon quantum dots prepared in Example 3 were added to BG11 liquid culture medium at a concentration of 0.6 mg / L to obtain nitrogen-doped arundinaceous carbon quantum dots-BG11 composite liquid culture medium.

[0084] (2) The Scenedesmus was inoculated into the nitrogen-doped Reed-based carbon quantum dot-BG11 composite liquid culture medium in step (1) at a concentration of 0.5 g / L; and cultured in a 27℃ light incubator for 21 days to obtain nitrogen-doped Reed-based carbon quantum dot-Scenedesmus composite fertilizer.

[0085] Comparative Example 2

[0086] A method for preparing a microalgae fertilizer, the steps are the same as in Example 4, except that Scenedesmus is inoculated into BG11 liquid culture medium.

[0087] Comparative Example 3

[0088] A method for preparing a microalgae fertilizer, the steps are the same as in Example 4, except that in step (1), the undoped arundinaceous carbon quantum dots prepared in Comparative Example 1 are added to BG11 liquid culture medium at a concentration of 0.5 mg / L.

[0089] Application Example 1

[0090] The performance of nitrogen-doped arundinaceous carbon quantum dots prepared in Example 1 was analyzed. The fluorescence spectra of the nitrogen-doped arundinaceous carbon quantum dots were detected using a fluorescence spectrophotometer (Shanghai Lingguang Technology Co., Ltd., F97 Pro). The operation was performed according to the instruction manual. The results are as follows: Figure 1 As shown.

[0091] according to Figure 1 It can be seen that the emission wavelength gradually redshifts with increasing excitation wavelength. The fluorescence intensity reaches its maximum when the excitation wavelength is 350 nm. This indicates that nitrogen-doped arundinaceous carbon quanta can absorb ultraviolet light that plants cannot absorb and utilize, and convert it into light that plants can use.

[0092] Application Example 2

[0093] The growth curves of *Scenedesmus* in Examples 4, 2, and 3 over 21 days were determined by microscopic counting. The method was described in *Experimental Microbiology* (edited by Zhou Deqing, Higher Education Press, 3rd edition, 2013). The results are as follows: Figure 2 As shown in Table 1, the data in Table 1 represent the cell density of Scenedesmus at different culture days.

[0094] Table 1. Growth curve of *Scenedesmus* after 21 days

[0095]

[0096] The results showed that the growth rate of *Scenedesmus* was faster when nitrogen-doped *Arundinella*-based carbon quantum dots were added, exceeding that of *Arundinella* without carbon quantum dots and *Arundinella*-based carbon quantum dots without nitrogen doping. Furthermore, the density of *Scenedesmus* was higher when the plateau phase (around 15 days) was reached. This indicates that nitrogen-doped *Arundinella*-based carbon quantum dots can promote the growth of *Scenedesmus*. In typical large-scale propagation culture of microalgae (such as *Scenedesmus*), the "self-shading effect" usually becomes significant around days 7-10 after inoculation (i.e., the mid-to-late logarithmic growth phase of the microalgae). At this time, with the rapid increase in algal cell concentration in the culture medium, the color of the culture medium deepens, and the light transmittance drops sharply. In traditional culture systems, insufficient light for deep cells during this period leads to a significant decrease in photosynthetic efficiency, resulting in a slowdown in biomass accumulation. In the later stages of cultivation using the compound fertilizer in Example 4 of this invention, the growth did not prematurely stagnate due to high density; instead, it significantly improved the final algal cell density at the plateau phase (around 15 days). This macroscopic phenotype, which "breaks the original environmental capacity limit and can still maintain photosynthesis at extremely high densities," is direct evidence that the self-shading effect is effectively weakened. The reason why nitrogen-doped arundinaceous carbon quantum dots can overcome the self-shading limitation in high-density microalgae cultivation is mainly reflected in the following three synergistic aspects: First, nitrogen-doped arundinaceous carbon quantum dots have excellent photoluminescence (PL) properties. They can absorb wavelengths that are difficult for microalgae to utilize in the culture system (such as ultraviolet light or some green light) and convert them into red or blue-violet light, which are most easily absorbed by chlorophyll a and chlorophyll b in microalgae, through fluorescence emission. In high-density deep culture media where "self-shading" occurs, these uniformly dispersed nano-carbon quantum dots act like countless "miniature light antennas" or "internal light sources," providing additional photosynthetically active radiation (PAR) to algal cells in low-light or dark areas, thereby activating the photosynthetic activity of deep cells. Second, due to the extremely small particle size of carbon quantum dots (usually less than 10 nm) and their good water solubility and dispersibility, they form a uniform nanosol system in the culture medium. When external light enters the culture medium, carbon quantum dots induce strong light scattering, altering the linear propagation path of photons in the liquid and increasing diffuse reflection and penetration depth within the culture medium. This weakens the light-blocking effect of surface algal cells from a physical optics perspective. Thirdly, compared to undoped carbon quantum dots, the in-situ nitrogen doping provided by urea not only increases the hydrophilic groups on the surface of the carbon quantum dots (making them more stable in the microalgae culture medium), but more importantly, the lone pair electrons of the nitrogen atoms alter the electron cloud distribution of the carbon nucleus, significantly improving the fluorescence quantum yield of the carbon quantum dots. Furthermore, some ultrasmall carbon quantum dots that enter the microalgae cells or attach to the cell surface, with their excellent electron conduction capabilities, can promote the efficiency of the electron transport chain in the microalgae photosystem II (PSII), physiologically improving the light energy conversion rate under weak light stress.

[0097] Meanwhile, the amount of IAA produced by Scenedesmus in Example 4 and Comparative Example 2 was determined by ultraviolet spectrophotometry, according to the method described in [Gordon, SA, & Weber, RP (1951). Colorimetric estimation of indoleaceticacid]. Plant Physiology , 26(1), 192-195】The results are as follows Figure 3 As shown in the figure. The results indicate that the addition of nitrogen-doped arundinaceous carbon quantum dots increased the IAA production of *Scenedesmus* by 12.45% compared to *Scenedesmus*.

[0098] Application Example 3

[0099] Select healthy chili seedlings with uniform growth and randomly divide them into 3 groups:

[0100] Nitrogen-doped Reed-based carbon quantum dot-microalgae compound fertilizer group: The nitrogen-doped Reed-based carbon quantum dot-microalgae compound fertilizer from Example 4 was applied at a rate of 200 mL / mu. The compound fertilizer was diluted 10 times with irrigation water during application. The first application was 7-10 days after transplanting. The application frequency was once every 2 weeks. This group was designated as the Reed-based carbon quantum dot-microalgae compound fertilizer group.

[0101] Scenedesmus group: The operation was the same as that of the Reed-based carbon quantum dot-microalgae compound fertilizer group, except that the microalgae fertilizer in Comparative Example 2 was applied.

[0102] Blank group: Irrigation with clean water.

[0103] Forty-five days after the first application of fertilizer, the plant height, stem diameter, SPAD (chlorophyll content), and root activity of peppers in each treatment group were measured. The methods were described in *Principles and Techniques of Plant Physiology and Biochemistry Experiments* (edited by Li Hesheng, Higher Education Press, 2000) and *Experimental Guide to Plant Physiology* (edited by Zou Qi, China Agriculture Press, 2000). Root activity of peppers was measured using the TTC reduction method. Results are shown in Figures 2 and 3. Figures 4-8 As shown.

[0104] Table 2. Detection indicators of chili peppers in each treatment group

[0105]

[0106] The results showed that the pepper plants grew more vigorously after applying nitrogen-doped reed-based carbon quantum dot-Scenedesmus compound fertilizer. The pepper plant height in the nitrogen-doped reed-based carbon quantum dot-Scenedesmus compound fertilizer group was 19.60% and 8.32% higher than the control group and the Scenedesmus group, respectively; the stem diameter in the nitrogen-doped reed-based carbon quantum dot-Scenedesmus compound fertilizer group was 28.57% and 9.76% higher than the control group and the Scenedesmus group, respectively. The leaf SPAD in the nitrogen-doped reed-based carbon quantum dot-Scenedesmus compound fertilizer group was 8.78% and 3.64% higher than the control group and the Scenedesmus group, respectively. The root activity in the nitrogen-doped reed-based carbon quantum dot-Scenedesmus compound fertilizer group was 58.70% and 29.29% higher than the control group and the Scenedesmus group, respectively.

[0107] Application Example 4

[0108] Healthy tomato seedlings, cauliflower seedlings, and corn seedlings with uniform growth were selected and randomly divided into two groups:

[0109] Nitrogen-doped Reed-based carbon quantum dot-microalgae compound fertilizer group: The nitrogen-doped Reed-based carbon quantum dot-microalgae compound fertilizer from Example 4 was applied at a rate of 300 mL / mu. The compound fertilizer was diluted 12 times with irrigation water during application. The first application was 7-10 days after the transplanting of tomato seedlings, cauliflower seedlings, and corn seedlings. The application frequency was once every 2 weeks. This group was designated as the Reed-based carbon quantum dot-microalgae compound fertilizer group.

[0110] Blank group: Irrigation with clean water.

[0111] Plant height was measured in each treatment group 45 days after the first application of fertilizer. The results are shown in Table 3.

[0112] Table 3 Plant height in each treatment group

[0113]

[0114] The results showed that after applying nitrogen-doped reed-based carbon quantum dot-Scenedesmus compound fertilizer, the plant height of tomatoes, cauliflower, and corn increased by 14.94%, 9.83%, and 11.54% respectively compared with the control group.

[0115] In summary, the nitrogen-doped arundinaceous carbon quantum dots prepared by the method described in this invention can improve the photosynthetic efficiency of Scenedesmus and promote the growth of microalgae; the nitrogen-doped arundinaceous carbon quantum dot-microalgae compound fertilizer prepared therefrom can effectively promote plant growth.

[0116] Although the above embodiments have provided a detailed description of the present invention, they are only some embodiments of the present invention, and not all embodiments. People can obtain other embodiments based on these embodiments without creative effort, and these embodiments all fall within the protection scope of the present invention.

Claims

1. A method for preparing nitrogen-doped arundocarpus-based carbon quantum dots, characterized in that, Includes the following steps: Arundo donax, a nitrogen source, and water were mixed and subjected to a hydrothermal reaction to obtain the nitrogen-doped arundo donax-based carbon quantum dots. The nitrogen source is urea; The mass ratio of reed to nitrogen source is 1:0.1 to 1:0.5; The hydrothermal reaction temperature is 150~220℃; the hydrothermal reaction time is 6~8h; The reed is reed powder; the preparation of the reed powder includes: drying and pulverizing the reed, and passing it through a 25-60 mesh sieve to obtain reed powder; The mass-to-volume ratio of the reed, nitrogen source, and water is 1g:5mL to 1g:10mL.

2. The nitrogen-doped arundinaceous carbon quantum dots prepared by the method of claim 1.

3. The application of the nitrogen-doped arundinaceous carbon quantum dots prepared by the method of claim 1, characterized in that, The application includes at least one of the following: (1) improving the photosynthetic efficiency of microalgae; (2) promoting the growth of microalgae; and (3) increasing the production of IAA by microalgae.

4. A compound fertilizer, characterized in that, The compound fertilizer comprises nitrogen-doped arundinaceous carbon quantum dots and microalgae prepared by the preparation method of claim 1; the nitrogen-doped arundinaceous carbon quantum dots are anchored on the surface of microalgal cells to form a carbon quantum dot-microalgae artificial photosynthetic hybrid system.

5. The compound fertilizer according to claim 4, characterized in that, The microalgae concentration in the compound fertilizer is ≥10 6 cell / mL.

6. The method for preparing the compound fertilizer according to claim 4 or 5, characterized in that, Includes the following steps: Microalgae were inoculated into a microalgae culture medium and co-cultured to obtain the compound fertilizer; The microalgae culture medium contains nitrogen-doped arundinaceous carbon quantum dots prepared by the method described in claim 1.

7. The preparation method according to claim 6, characterized in that, The content of nitrogen-doped arundinaceous carbon quantum dots in the microalgae culture medium is 0.1~1 mg / L.

8. The application of the compound fertilizer according to claim 4 or 5 or the compound fertilizer prepared by the preparation method according to claim 6 or 7 in promoting plant growth.

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