Cell membrane stabilizing method and system for improving low-temperature adaptability of rice in rice and crab planting

Abstract

The invention relates to the technical field of agricultural biology, particularly discloses a cell membrane stabilizing method and system for improving low-temperature adaptability of rice in rice and crab planting and breeding, and aims to solve the problems of cell membrane damage caused by low-temperature stress of rice in rice and crab planting and breeding, ecological safety and popularization. CMSPC is prepared, targeted modified nanoparticles are loaded and automatically applied to a rice field, and cell membranes are stabilized after the CMSPC is absorbed by rice; the system integrates CMSPC preparation, environment / rice / crab monitoring, decision making and application. According to the accurate, environment-friendly and efficient solution, the low-temperature adaptability of the rice is effectively improved, meanwhile, the limitation that a traditional breeding period is long, the agronomic management effect is limited and a chemical / biological regulating agent is not friendly to the ecological environment is overcome, and a key technical support is provided for environment-friendly, sustainable and healthy development of the rice crab breeding industry.

Description

TECHNICAL FIELD

[0001] The present application belongs to the technical field of agricultural biotechnology, and in particular relates to a method and system for improving the cell membrane stability of rice in low-temperature adaptation in rice-crab breeding. BACKGROUND

[0002] As one of the most important food crops in the world, the yield and quality of rice are directly related to the survival and development of human beings. With the increasing emphasis on ecological benefits and sustainable development in modern agriculture, ecological agricultural models such as rice-crab breeding are gradually being promoted and applied because they can effectively improve the comprehensive utilization rate of farmland, reduce the use of fertilizers and pesticides, improve the quality of agricultural products, and increase farmers' income. However, this ecological breeding model, while bringing many benefits, also faces serious challenges from environmental stress. Among them, low-temperature stress is a major limiting factor affecting the normal growth and development of rice and even the stability of the rice-crab breeding system, especially in northern China or high-altitude areas, and during seasonal transitions, sudden low temperatures or persistent cold waves often lead to slow growth, physiological dysfunction, and even large-scale yield reduction or seedling death in rice, which in turn affects the balance and economic benefits of the entire rice-crab ecological system.

[0003] In response to the problem of rice low-temperature adaptation, researchers in the field have conducted long-term research and practice and have explored various coping strategies. Traditionally, the main approach is to breed rice varieties with strong cold resistance to improve their tolerance to low temperatures. This method usually relies on plant breeding principles and introduces cold-resistant genes or related traits into target varieties through hybridization, backcrossing, and other means. In addition, at the agronomic management level, researchers have also tried to adjust the planting date, reasonable fertilization, improve soil structure, and use deep water insulation measures to provide a relatively favorable growing environment for rice, thereby alleviating low-temperature stress to some extent. Further, some biological or chemical regulators, such as plant growth regulators (e.g., gibberellins, cytokinins, etc.), osmotic regulators (e.g., proline, betaine, etc.), and antioxidants (e.g., vitamins C, E, etc.), have also been used externally to enhance the plant's own stress physiological mechanisms, such as removing active oxygen free radicals and maintaining cell osmotic balance, thereby improving its survival rate and growth performance under low temperatures.

[0004] However, with the continuous promotion of rice crab breeding mode and the increasing demand for green and non-polluted agricultural products in the market, the inherent characteristics of the above-mentioned prior art solutions in the principle level gradually show limitations and deep contradictions in dealing with low temperature stress. Specifically, the traditional breeding method has a long cycle and high cost, and it is difficult to balance multiple complex traits such as yield, quality and resistance during the breeding process. The newly bred cold-resistant varieties may not perform well in other aspects. Although the agronomic management measures have cost advantages, their effects are significantly limited by natural environmental conditions. In the face of extreme or long-term low temperature weather, they often cannot provide sufficient protection. In essence, most of the existing strategies to enhance the cold resistance of rice are macro-regulation or indirect intervention, and they cannot efficiently and accurately target the key initial damage caused by low temperature stress to rice cells, i.e., the integrity and functional stability of the cell membrane system.

[0005] The deeper contradiction lies in that in the special ecosystem of rice crab breeding, rice, crabs and their associated microbial communities in the water body together form a fragile and complex ecological chain. Under low temperature environment, the damage of lipid phase transition, membrane protein denaturation and ion leakage of rice cell membrane is the core reason for cell dysfunction and even cell death. However, traditional chemical regulators, even if they can improve the cold resistance of rice to some extent, their components and residues may have toxic side effects on crabs as symbiotic partners in the rice field. For example, some synthetic plant growth regulators or metabolic products may change the water environment, affect the growth and development of crabs, molting and reproduction, and even cause death, thereby seriously damaging the ecological balance and economic benefits of rice crab breeding. Even some biological agents or natural extracts considered to be relatively safe, their stability, bioavailability in complex water environment, and whether they can be efficiently absorbed by rice cells and targeted to cell membranes, are still unresolved technical problems. The existing technical solutions generally lack a comprehensive method that can efficiently and specifically stabilize and protect the integrity and function of rice cell membranes under low temperature, while being harmless to the water environment and the health of crabs in the rice field, and being easy to promote and apply on a large scale. This inherent contradiction between the demand for rice low temperature adaptability and the ecological compatibility of rice crab breeding has become a key bottleneck restricting the sustainable and healthy development of rice crab breeding industry.

[0006] Therefore, how to develop a method and system that can accurately target rice cell membranes and effectively improve their stability under low temperature stress, while ensuring that the method has high biological safety and environmental friendliness in the rice crab breeding ecosystem, has become a key challenge and technical problem to be solved for those skilled in the art. SUMMARY

[0007] The present application aims to solve the limitations and deep contradictions of the existing low-temperature adaptability technical solutions in rice-crab breeding mode, especially for the key initial damage caused by low-temperature stress to the rice cell membrane system, and provides a cell membrane stabilization method and system for improving the low-temperature adaptability of rice in rice-crab breeding, which can efficiently and specifically stabilize and protect the integrity and function of rice cell membrane under low temperature, is harmless to the water environment of rice field and the health of crabs as symbiotic partners, and is easy to promote and apply on a large scale.

[0008] To achieve the above-mentioned application purposes, the present application provides a cell membrane stabilization method for improving the low-temperature adaptability of rice in rice-crab breeding, which comprises the following steps: a) preparing a cell membrane stabilization precursor complex (CMSPC); b) preparing a nano-carrier system (NCS), and efficiently loading the CMSPC inside the NCS to form CMSPC nanoparticles; c) uniformly dispersing the CMSPC nanoparticles in the rice field water at a predetermined concentration; d) applying the CMSPC nanoparticles, so that they are selectively absorbed by the rice root cells and enter the cells, and then the CMSPC is released in the cells, so that the CMSPC plays a synergistic role inside the rice cells, thereby improving the integrity and functional stability of the rice cell membrane under low-temperature stress.

[0009] Specifically, in the step a), the cell membrane stabilization precursor complex (CMSPC) is composed of the following components: a phospholipid precursor, an osmotic regulator, and an oligopeptide. The phospholipid precursor is a choline compound, specifically choline chloride or hydrogen tartrate choline, with a purity of not less than 99.5% and a molecular weight range of 139.62 g / mol to 253.25 g / mol. The osmotic regulator is glycine betaine, with a purity of not less than 98.0% and a molecular weight of 117.15 g / mol. The oligopeptide is a synthetic tetrapeptide or pentapeptide, with a molecular weight range of 300 Da to 600 Da, which is prepared by solid-phase synthesis method, with a purity of not less than 95%. In the CMSPC, the mass ratio of the choline compound, the glycine betaine, and the oligopeptide is (2-5):(1-3):(0.5-1.5). The preparation of the CMSPC is by mixing and dissolving the three components in ultrapure water or ethanol solution according to the predetermined mass ratio, and stirring by a magnetic stirrer at a speed of 200 rpm for 30 minutes, until a uniform solution is formed.

[0010] Further, the phospholipid precursor, choline chloride or choline bitartrate, as a key precursor for phosphatidylcholine (PC) synthesis, promotes the biosynthesis of PC, the major lipid component of cell membrane, in rice cells through the phosphatidylcholine synthesis pathway. The glycine betaine, as a broad-spectrum compatible solute, accumulates in rice cells under low temperature stress, effectively reduces the membrane lipid phase transition temperature induced by low temperature, thereby enhancing the fluidity and integrity of the cell membrane, by stabilizing the protein structure in the cell and maintaining the osmotic balance of the cell. The oligopeptide interacts with the lipid bilayer or membrane proteins of the cell membrane, such as embedding in the lipid bilayer through hydrophobic interaction, or binding to the membrane surface through electrostatic interaction, thereby enhancing the mechanical strength and stability of the membrane, and possibly acting as a signal molecule to induce the expression of endogenous cold-resistant genes in rice, enhance the conformational stability of membrane proteins, and promote the activity of aquaporins, thereby maintaining the normal physiological functions of the cell. The three components synergistically act in rice cells to enhance the low temperature stress resistance of the cell membrane from different aspects.

[0011] As a preferred embodiment of the present application, in step b), the nanocarrier system (NCS) is composed of a chitosan-alginate core-shell structure nanoparticle. The chitosan is a medical grade chitosan with a degree of deacetylation between 85% and 95% and a molecular weight between 50 kDa and 190 kDa. The alginate is in the form of sodium salt, with a molecular weight between 10 kDa and 200 kDa, and a ratio of guluronic acid (G) to mannuronic acid (M) (G / M ratio) between 1.5 and 2.0. The core-shell structure nanoparticle is prepared by ionotropic gelation method, specifically including the following steps: b1) Preparation of chitosan solution: dissolve the chitosan in acetic acid solution with a concentration of 1.0% (v / v), prepare a chitosan solution with a concentration of 1.0 mg / mL to 3.0 mg / mL, and stir with a magnetic stirrer at a speed of 400 rpm for 12 hours until the chitosan is completely dissolved.

[0012] b2) Preparation of sodium tripolyphosphate (TPP) solution: dissolve the sodium tripolyphosphate in ultrapure water to prepare a TPP solution with a concentration of 0.25 mg / mL to 1.0 mg / mL.

[0013] b3) Preparation of sodium alginate solution: dissolve the sodium alginate in ultrapure water to prepare a sodium alginate solution with a concentration of 1.0 mg / mL to 3.0 mg / mL.

[0014] b4) Formation of CMSPC nanoparticles core loaded: The TPP solution containing the CMSPCs was added dropwise into the chitosan solution under constant stirring (e.g., 800 rpm) at a rate of 0.5 mL / min to 2.0 mL / min by a peristaltic pump or a syringe pump, and the concentration of CMSPCs in the TPP solution was 0.5 mg / mL to 2.0 mg / mL. The mixing process was continued for 30 minutes to 60 minutes to allow the chitosan and TPP to form CMSPC-loaded nanoparticle cores with positive charges through electrostatic interaction. The mixing process was carried out at a constant temperature of 25 °C. The pH value of the solution was maintained between 4.5 and 5.5 during the formation of the nanoparticle cores.

[0015] b5) Formation of core-shell structure: The sodium alginate solution was added dropwise into the CMSPC-loaded nanoparticle core suspension at a rate of 0.5 mL / min to 2.0 mL / min by a peristaltic pump or a syringe pump. The sodium alginate solution and the nanoparticle cores formed a core-shell structure through electrostatic interaction, and the alginate layer was coated on the outside of the chitosan-CMSPC nanoparticle core to form CMSPC nanoparticles with negative charges. The core-shell formation process was continued for 30 minutes to 60 minutes. The mixing process was carried out at a constant temperature of 25 °C.

[0016] b6) Purification and characterization: The obtained CMSPC nanoparticles were purified by centrifugation (e.g., 15,000 rpm for 30 minutes) and ultrafiltration (e.g., 100 kDa molecular weight cut-off) to remove unreacted substances and free CMSPCs. The particle size distribution and polydispersity index (PDI) of the nanoparticles were measured by a dynamic light scattering (DLS) instrument, and the particle size range was 50 nm to 200 nm, and the PDI was less than 0.2. The surface charge of the nanoparticles was measured by a Zeta potential analyzer, and the surface charge range was -20 mV to -40 mV. The morphology and core-shell structure of the nanoparticles were observed by transmission electron microscopy (TEM). The chemical composition of the nanoparticles was analyzed by Fourier transform infrared spectroscopy (FTIR). The thermal stability of the nanoparticles and the loading amount of CMSPCs were determined by thermogravimetric analysis (TGA). The unloaded CMSPCs were detected by high-performance liquid chromatography (HPLC) or ultraviolet-visible spectrophotometry, so as to calculate the encapsulation efficiency of CMSPCs, and the encapsulation efficiency was not less than 80%.

[0017] Further, to enhance the selective recognition and uptake efficiency of the CMSPC nanoparticles by rice root cells, the surface of the chitosan-alginate core-shell structure nanoparticles is chemically modified. The chemical modification is achieved by covalently attaching a specific targeting ligand to the chitosan surface. The specific targeting ligand is a glucose or arabinose derivative, specifically 2-deoxy-2-(2,5-dicarboxy-1,4-dihydrobenzyl) acetylglucosamine or 3-amino-3-deoxy-D-arabinose. The attachment of the targeting ligand is achieved by carbodiimide chemistry (EDC / NHS coupling reaction). The specific steps are as follows: after the purification of the CMSPC nanoparticles, the nanoparticles are dispersed in a MES buffer solution (pH 6.0), and 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride (EDC) and N-hydroxysuccinimide (NHS) are added to activate the exposed carboxyl groups on the surface of the nanoparticles. Then the glucose or arabinose derivative is added, and a stable amide bond is formed between the amino group of the glucose or arabinose derivative and the activated carboxyl group. The coupling reaction is carried out at room temperature for 4 to 8 hours, after which the modified nanoparticles are again purified by centrifugation and ultrafiltration to remove unreacted reagents. The surface-modified nanoparticles have a grafting density of 50 to 150 targeting ligand molecules per 100 nm2 of nanoparticle surface area, which is confirmed by X-ray photoelectron spectroscopy (XPS) or Fourier transform infrared spectroscopy (FTIR). The glucose or arabinose derivative can mimic the natural molecules recognized by specific receptors or transport proteins on the cell membrane of rice root cells, thereby inducing receptor-mediated endocytosis or specific transport protein-mediated uptake, significantly improving the transport efficiency and selectivity of the nanoparticles to rice root cells, while avoiding the uptake of non-targeted cells and reducing the potential impact on other organisms (e.g., crabs) in the rice field water.

[0018] As an embodiment of the present application, in step c), the CMSPC nanoparticles are uniformly dispersed in the paddy field water body at a preset concentration. The preset concentration refers to the effective active ingredient concentration of CMSPC in the CMSPC nanoparticles being 1 mg to 5 mg per liter of water body. The uniform dispersion is achieved by an automatic application system, which includes a multi-rotor unmanned aerial vehicle or a ground mobile robot equipped with a high-precision liquid droplet spraying system. The unmanned aerial vehicle or robot is equipped with a liquid storage tank with a capacity of 50 L to 200 L, which is connected to a peristaltic pump or gear pump with a flow control accuracy of ± 2%, which is used to deliver the CMSPC nanoparticle suspension to a nozzle array. The nozzle array is composed of multiple flat fan nozzles, which operate at a pressure of 0.2 MPa to 0.5 MPa, producing liquid droplets with a size of 50 µm to 150 µm, ensuring that the droplets can uniformly cover the surface of the rice leaves and fall into the paddy field water body, thereby enabling the nanoparticles to effectively settle in the water body area around the rice roots. The unmanned aerial vehicle or robot provides centimeter-level positioning accuracy through a real-time kinematic positioning (RTK-GPS) system and an inertial measurement unit (IMU), and combines a preset path planning algorithm to achieve precise and seamless coverage of the target paddy field area, ensuring that the uniformity deviation of the application of CMSPC nanoparticles in the entire paddy field area is less than 10%. The application timing is determined based on the following principles: when the weather forecast shows that the local area's daily average temperature is expected to drop below 15°C, or when the physiological monitoring data of the rice plants shows early signs of low temperature stress, the system starts the automatic application program. The application frequency is once every 3 to 5 days during continuous low temperature stress.

[0019] As an embodiment of the present application, in step d), the CMSPC nanoparticles are selectively absorbed by the rice root cells and are achieved through the following mechanism: the surface-modified CMSPC nanoparticles settle around the rice roots in the paddy field water body, and the glucose or arabinose derivatives on the surface of the nanoparticles specifically bind to specific sugar transport proteins or receptors expressed on the cell membrane of the rice root cells. The binding triggers receptor-mediated endocytosis, allowing the nanoparticles to be completely taken into the root cells to form endosomes. Alternatively, the nanoparticles interact with specific channel proteins on the cell membrane, promoting the nanoparticles or the released CMSPC to enter the cytoplasm through transmembrane transport. The endosomes gradually degrade in the cytoplasm, releasing the encapsulated CMSPC. The degradation process is regulated by intracellular pH and enzyme activity. The biocompatible materials of the nanoparticles (chitosan and alginate) gradually decompose into non-toxic small molecular oligosaccharides under the action of intracellular enzymes (such as lysosomal enzymes), which are metabolized by the rice cells or excreted, thereby achieving biodegradability and ensuring safety to the rice cells and the entire ecological environment.

[0020] As a preferred embodiment of the present application, the present application also provides a cell membrane stabilization system for improving the low temperature adaptability of rice in rice-crab breeding. The system comprises: 1. CMSPC nanoparticle preparation module: for preparing the above-mentioned CMSPC nanoparticles.

[0021] 2. Environmental parameter sensing module: for real-time monitoring of meteorological parameters of the rice field water and environment.

[0022] 3. Rice physiological response monitoring module: for non-invasively assessing the degree of low temperature stress and physiological health status of rice.

[0023] 4. Crab ecological state monitoring module: for non-invasively monitoring the health status and behavior pattern of crabs in the rice field water.

[0024] 5. Central control and decision-making module: for integrated processing of all monitoring data, low temperature stress prediction, application decision-making, and issuing instructions to the application module.

[0025] 6. Automated application module: for accurately dispersing CMSPC nanoparticles into the rice field water according to the instructions of the central control and decision-making module.

[0026] Specifically, the CMSPC nanoparticle preparation module includes a production workshop with a cleanliness of ISO 7, which is internally configured with: Raw material storage unit: for storing high-purity chitosan, sodium alginate, sodium tripolyphosphate, choline compounds, glycine betaine, amino acid monomers, and solvents (such as acetic acid, ultrapure water, ethanol). The storage unit is equipped with a temperature and humidity control system and an inert gas protection system to ensure the stability of the raw materials.

[0027] CMSPC synthesis reactor: a stainless steel jacketed reactor with precise temperature (±0.1°C), pH (±0.01 pH) and stirring speed (±1 rpm) control functions, with a volume of 10 L to 100 L, for the mixed dissolution of CMSPC.

[0028] Nanoparticle synthesis reactor: a reactor equipped with a high-shear homogenizer (rotation speed up to 20,000 rpm) or an ultrasonic disperser (power adjustable range 100W-1000W), for the mixing of chitosan solution and TPP solution and the formation of the core, as well as the addition of sodium alginate solution and the formation of the core-shell structure. The reactor is equipped with a sample injection pump (accuracy ±0.1 mL / min) and an online particle size monitoring system (e.g., based on dynamic light scattering principle).

[0029] Surface modification reactor: a reaction vessel equipped with nitrogen protection, constant temperature circulating water bath and magnetic stirring function, used for grafting targeting ligands on the surface of purified nanoparticles.

[0030] Separation and purification unit: including a set of continuous flow centrifuge (rotation speed up to 20,000 rpm, processing capacity 1 L / min to 10 L / min) and a set of tangential flow ultrafiltration system (membrane molecular weight cut-off 100 kDa, filtration area 0.5 m² to 5 m²), used to remove unreacted substances and free CMSPC, and to concentrate the nanoparticles.

[0031] Freeze-drying unit: an industrial-grade vacuum freeze dryer with automatic temperature control (-80°C to 0°C) and vacuum degree (1 Pa to 100 Pa) adjustment function, used to freeze-dry the purified nanoparticle suspension into powder for storage and transportation.

[0032] Quality control and detection unit: equipped with DLS instrument, Zeta potential analyzer, TEM, FTIR, HPLC, XPS and other equipment, used for comprehensive quality detection of raw materials, intermediate products and final CMSPC nanoparticles, to ensure that the product meets the preset specification standards.

[0033] The environmental parameter perception module comprises: Distributed weather monitoring station network: composed of multiple (for example, 10 per hectare) independent wireless sensor nodes. Each node integrates a high-precision digital temperature sensor (for example, Pt100 RTD, measurement range -10°C to 50°C, accuracy ±0.1°C), humidity sensor (measurement range 0-100% RH, accuracy ±2% RH), wind speed and direction sensor (measurement range 0.3-30 m / s, accuracy ±5%), rainfall sensor (resolution 0.2 mm).

[0034] Water quality monitoring buoy: multiple (for example, 3 per hectare) self-powered buoys, each carrying multiple-parameter water quality sensors, including: pH electrode (measurement range 2-12, accuracy ±0.02 pH), dissolved oxygen (DO) sensor (for example, optical dissolved oxygen sensor, measurement range 0-20 mg / L, accuracy ±0.1 mg / L), conductivity sensor (measurement range 0-200 mS / cm, accuracy ±1%), turbidity sensor (measurement range 0-1000 NTU, accuracy ±1%).

[0035] Solar radiation sensor: each monitoring station and buoy is equipped with a photosynthetically active radiation (PAR) sensor (measurement range 0-3000 µmol / m² / s, accuracy ±5%) for measuring light intensity.

[0036] Data transmission unit: All sensor nodes and buoys are connected to the central data collection gateway through LoRaWAN or NB-IoT wireless communication protocols, which provide low-power, long-distance, secure data transmission with a transmission rate of 2.5 kbps to 50 kbps and a data transmission delay of less than 5 seconds. Data transmission uses the AES-128-bit encryption standard.

[0037] The rice physiological response monitoring module comprises: Unmanned aerial or ground robot-mounted hyperspectral imaging system: equipped with a hyperspectral camera with a spectral range of 400 nm to 1000 nm and a spectral resolution of better than 5 nm and a spatial resolution of 5 cm / pixel. The camera is used to collect reflectance spectral data of rice plants. By analyzing the spectral data, normalized difference vegetation index (NDVI), photochemical reflectance index (PRI), water vapor stress index (CWSI), and other physiological indicators such as chlorophyll content and nitrogen content are calculated to evaluate the growth status and stress degree of rice.

[0038] Unmanned aerial or ground robot-mounted chlorophyll fluorescence measurement system: equipped with a pulse amplitude modulation (PAM) fluorometer for non-invasive measurement of chlorophyll fluorescence parameters of rice leaves, including maximum photochemical quantum efficiency (Fv / Fm), actual photochemical efficiency (ΦPSII), and non-photochemical quenching (NPQ). The measurement system provides modulated light (e.g., 650 nm) through an LED light source and detects fluorescence signals at 730 nm. The parameters can directly reflect the functional state of the photosynthetic system of rice and the response to low temperature stress.

[0039] Unmanned aerial or ground robot-mounted thermal infrared imaging system: equipped with a high-sensitivity thermal infrared camera (e.g., NETD less than 50 mK) with a spatial resolution of better than 10 cm / pixel. Used to measure the surface temperature of rice leaves, reflecting the efficiency of transpiration, and as an indicator of water stress and cold stress of rice.

[0040] Data synchronization and integration unit: all imaging and measurement systems are connected to the onboard or vehicle-mounted data processor through high-speed Ethernet (e.g., Gigabit Ethernet), which has at least 8-core ARM Cortex-A series CPU and 16 GB RAM for real-time data preprocessing and compression, and transmits data to the central control and decision module through a dedicated encrypted wireless link (e.g., 5G or WiFi in a proprietary frequency band).

[0041] The crab ecological state monitoring module comprises: Distributed underwater visual monitoring system: composed of multiple (e.g., 5 per hectare) low-light high-definition (1080p) underwater cameras with IP68 waterproof rating. The cameras are equipped with wide-angle lenses (e.g., 120° field of view) and infrared fill light for continuous monitoring of crab activity, feeding, molting behavior, and the presence of abnormal states during nighttime or low-light conditions. The cameras have built-in motion detection algorithms that automatically trigger high-frame-rate recording and upload key segments when abnormal crab activity is detected.

[0042] Underwater acoustic sensor array: multiple (e.g., 3 per hectare) high-sensitivity hydrophones (frequency response range 20 Hz to 20 kHz) for capturing underwater acoustic signals such as crab crawling, feeding, and biological vocalizations. Through spectral analysis of acoustic signals, the activity level and presence of stress reactions in crabs can be indirectly assessed.

[0043] Water metabolite analysis unit: by regularly and automatically extracting rice field water samples, using a portable multi-parameter analyzer or sending water samples to the laboratory for analysis. The analysis is mainly aimed at stress biomarkers of crabs, such as ammonia nitrogen content, nitrate content, nitrite content, and specific hormone (such as molting hormone) metabolite concentration in water, as an auxiliary indicator to assess the health status of crabs. The water sample extraction frequency is once a day, and each time 50 mL is extracted.

[0044] Data transmission unit: underwater cameras and acoustic sensors are connected to shore-based data processors through underwater optical fiber or Ethernet cables, and data is transmitted through encrypted wireless links.

[0045] The central control and decision-making module is the core intelligent unit of the system, which is configured with: High-performance data integration server: equipped with multi-core high-performance CPU (e.g., Intel Xeon series), at least 64 GB DDR4 ECC RAM and 2 TB NVMe SSD storage, for receiving, storing and processing real-time big data streams from all monitoring modules. The server runs the Linux operating system and deploys a distributed database (e.g., Cassandra or MongoDB) to handle high-concurrency data writes.

[0046] Big data analysis and prediction engine: based on deep learning models (e.g., long short-term memory network LSTM) and ensemble learning models (e.g., random forest or gradient boosting tree). The model uses historical meteorological data, rice physiological parameters, crab behavior data, and CMSPC nanoparticle application effect data as the training set, which is used to: Low temperature stress prediction: Predict the likelihood, intensity, and duration of low temperature stress in the rice field area within the next 24 to 72 hours with a prediction accuracy of over 85%.

[0047] CMSPC nanoparticle administration strategy optimization: Based on the predicted low temperature stress level and the current physiological state of the rice, calculate the optimal CMSPC nanoparticle administration time, dosage, and frequency to minimize the amount of administration while maximizing the protection effect.

[0048] Ecological risk assessment: Continuously assess water quality and crab physiological behavior data to predict the potential impact of CMSPC nanoparticle administration on crabs and adjust the administration strategy or issue warnings if any potential negative effects are found.

[0049] Decision support system: An expert system based on rule engines and fuzzy logic controllers. This system receives the output of the prediction engine and combines pre-set agronomic management rules, environmental safety thresholds (e.g., water pH should not be lower than 6.5, DO should not be lower than 4 mg / L), and crab protection strategies to generate final administration instructions or recommendations. The system has a multi-objective optimization algorithm to balance the optimal solution between improving rice cold resistance and rice-crab ecological health.

[0050] Human-machine interface (HMI): A web-based graphical user interface accessed through encrypted HTTPS protocol. This interface provides real-time data visualization (e.g., map overlay displaying sensor data, trend charts), warning information push (e.g., SMS, email), historical data query, administration plan viewing and editing, and manual instruction input and emergency stop functions.

[0051] Communication interface unit: Communicate with the automated administration module through MQTT or RESTful API protocols for bidirectional communication to ensure accurate instruction delivery and real-time feedback of execution status. The communication link uses TLS 1.2 or higher encryption protocol.

[0052] The automated administration module includes: Multi-rotor unmanned aerial vehicle or ground robot: As previously described, equipped with RTK-GPS and IMU, providing centimeter-level positioning and attitude control accuracy for autonomous navigation in the rice field environment. The drone or robot has a redundant power system and failsafe mechanism to ensure operational reliability.

[0053] Precise liquid spray system: a liquid storage tank made of corrosion-resistant and ultraviolet-resistant material with a capacity of 100 L to 500 L, the inside of the tank is equipped with a stirrer to ensure the uniformity of the CMSPC nanoparticle suspension. The storage tank is connected to a high-precision metering pump (e.g., peristaltic pump or diaphragm pump) with a flow control range of 0.5 L / min to 10 L / min and a control accuracy of ±0.5%. The pump delivers the suspension to the nozzle array through a set of high-pressure hoses.

[0054] Nozzle array and droplet control system: the array is composed of multiple (e.g., 8 to 16) independently controllable electric atomizing nozzles. Each nozzle can adjust the spray pattern (e.g., fan-shaped or conical), droplet size (e.g., 50 µm to 200 µm), and spray flow to adapt to different wind speeds and application requirements. The droplet control system can adjust the nozzle parameters in real time according to the instructions of the central control module, ensuring that the droplets can be accurately deposited on the target area (rice root water layer or leaf surface) and maximizing the deposition efficiency and minimizing the drift loss.

[0055] Task management and execution unit: the flight control computer or motion controller built into the unmanned aerial vehicle or robot runs a real-time operating system (RTOS) and is responsible for parsing the application instructions issued by the central control module, executing flight path planning, spray parameter adjustment, and real-time collection and feedback of sensor data (such as heading, altitude, and speed).

[0056] Power management system: the unmanned aerial vehicle uses high-energy-density lithium polymer batteries with a continuous operation time of 30 minutes to 60 minutes. The ground robot uses high-capacity lithium-ion battery packs with a continuous operation time of 8 hours to 16 hours. Both are equipped with automatic charging or battery replacement systems to support long-term continuous operation.

[0057] The cell membrane stabilization method and system for improving the low-temperature adaptability of rice in rice-crab breeding provided by the present application have significant beneficial effects compared to the prior art.

[0058] First, the present application precisely targets the cell membrane stabilization precursor complex (CMSPC) to directly act on the key damage to the rice cell membrane caused by low-temperature stress, fundamentally improving the integrity and functional stability of the rice cell membrane. The choline compound in the CMSPC promotes the synthesis of phosphatidylcholine, glycine betaine stabilizes membrane lipids and proteins, and oligopeptides enhance membrane structure and may induce endogenous cold resistance, all of which work together to build a multi-level membrane protection mechanism.

[0059] Secondly, the chitosan-alginate core-shell structure nanoparticles used as the carrier system in the present application are all derived from natural biomass, and have high biocompatibility, biodegradability and environmental friendliness. The nanoparticles stably exist in the paddy field water body, are selectively absorbed and degraded by the rice cells, and do not cause pollution to the water body environment and toxic side effects to the crabs and their associated microbial communities in the paddy field. The surface-modified nanoparticles further improve the targeting and absorption efficiency of the rice root cells, significantly reduce non-targeted absorption, and ensure the ecological safety in the rice-crab symbiotic system.

[0060] Thirdly, the integrated intelligent monitoring and application system proposed in the present application realizes comprehensive, real-time and non-invasive monitoring of the paddy field environment, rice physiology and crab ecological state. Through big data analysis and machine learning models, the system can accurately predict the occurrence of low temperature stress, and adaptively optimize the application strategy (time, dose, frequency) of the CMSPC nanoparticles, so as to realize on-demand and accurate application, avoid over-application or insufficient application, improve resource utilization efficiency, and maximize the health and stability of the rice-crab symbiotic system.

[0061] Fourthly, the method and system of the present application are easy to be popularized and applied on a large scale. The preparation process of the nanoparticles is mature and easy to be industrialized, and the application process can be highly automated and intelligent, greatly reducing the labor cost and operation difficulty. This precise, environmentally friendly and efficient solution effectively overcomes the limitations of long traditional breeding cycle, limited agronomic management effect and ecological environment unfriendliness of chemical / biological control agents, and provides key technical support for the sustainable and healthy development of the rice-crab breeding industry.

[0062] Fifthly, by stabilizing the cell membrane, the present application not only helps to improve the survival rate and growth rate of rice under low temperature, but also improves the photosynthetic efficiency, nutrient absorption capacity and resistance to secondary stress of rice, thereby ensuring the yield and quality of rice and further improving the economic benefits of rice-crab breeding. BRIEF DESCRIPTION OF DRAWINGS

[0063] Figure 1 is a flowchart of the cell membrane stabilization method for improving the low temperature adaptability of rice in rice-crab breeding of the present application.

[0064] Figure 2 is a structural schematic diagram of the cell membrane stabilization system for improving the low temperature adaptability of rice in rice-crab breeding of the present application.

[0065] The reference signs are as follows: 1, CMSPC nanoparticles preparation module; 2, environmental parameter sensing module; 3, rice physiological response monitoring module; 4, crab ecological state monitoring module; 5, central control and decision module; 6, automatic application module; 11, raw material storage unit; 12, CMSPC synthesis reactor; 13, nanoparticle synthesis reactor; 14, surface modification reactor; 15, separation and purification unit; 16, freeze-drying unit; 17, quality control and detection unit; 21, distributed weather monitoring station network; 22, water quality monitoring buoy; 23, solar radiation sensor; 24, data transmission unit; 31, unmanned aerial or ground robot-borne hyperspectral imaging system; 32, unmanned aerial or ground robot-borne chlorophyll fluorescence measurement system; 33, unmanned aerial or ground robot-borne thermal infrared imaging system; 34, data synchronization and integration unit; 41, distributed underwater visual monitoring system; 42, underwater acoustic sensor array; 43, water body metabolite analysis unit; 44, data transmission unit; 51, high-performance data integration server; 52, big data analysis and prediction engine; 53, decision support system; 54, human-machine interface (HMI); 55, communication interface unit; 61, multi-rotor unmanned aerial vehicle or ground robot; 62, precision liquid spray system; 63, nozzle array and droplet control system; 64, task management and execution unit; 65, power management system. DETAILED DESCRIPTION

[0066] The present application provides a method and system for improving the cell membrane stability of rice in low temperature adaptation in rice-crab breeding, aiming to effectively address the initial damage to the rice cell membrane system caused by low temperature stress in the rice-crab breeding mode, thereby improving the survival ability and growth efficiency of rice under low temperature stress, while ensuring the safety of the rice field ecological environment and symbiotic crabs. This embodiment will elaborate the specific composition, operation steps, key parameters and implementation details of the method and system, and through specific examples and comparative examples, the technical advantages will be quantitatively demonstrated.

[0067] In one specific embodiment, the method for improving the cell membrane stability of rice in low temperature adaptation in rice-crab breeding disclosed by the present application is characterized in that a multi-level protection and strengthening is provided to the rice cell membrane system in the early stage of low temperature stress by precisely preparing and targetedly applying cell membrane stabilizing precursor complex (CMSPC) nanoparticles. The method mainly comprises the following steps: first, preparing the CMSPC; second, constructing a nano-carrier system (NCS) and efficiently loading the CMSPC inside the NCS to form CMSPC nanoparticles, and modifying the surface of the nanoparticles to enhance the targeting; third, uniformly dispersing and applying the obtained CMSPC nanoparticles in the water body of the rice field at a predetermined concentration; further, promoting the selective absorption of the CMSPC nanoparticles by the rice root cells and effectively releasing the CMSPC inside the rice cells; finally, fundamentally improving the integrity and functional stability of the rice cell membrane under low temperature stress by the synergistic effect of the CMSPC inside the rice cells.

[0068] Specifically, the preparation of cell membrane stabilizing precursor complex (CMSPC) in step a) is the basis of the whole method. The complex is composed of three key components: a phospholipid precursor, an osmotic regulator, and an oligopeptide. The phospholipid precursor is preferably a choline compound, which can be selected from choline chloride or choline bitartrate. The selection criteria are high purity and appropriate molecular weight. In this embodiment, the purity of choline chloride or choline bitartrate selected is not less than 99.5%, and the molecular weight range is controlled between 139.62 g / mol and 253.25 g / mol, to ensure its biological activity and conversion efficiency as a phosphatidylcholine (PC) synthesis precursor. The osmotic regulator is glycine betaine, which has a purity requirement of not less than 98.0% and a molecular weight of 117.15 g / mol. This purity level ensures its stability and biological function as a compatible solute in cells. The oligopeptide is a synthetic tetrapeptide or pentapeptide prepared by solid-phase synthesis, so that it can effectively interact with the cell membrane lipid bilayer or membrane protein. The oligopeptide is synthesized by Shanghai Kepai Biotechnology Co., Ltd., with an HPLC detected purity of 97%. The molecular weight range of the oligopeptide is limited to between 300 Da and 600 Da, with a purity requirement of not less than 95%. This molecular weight range is beneficial to its transport and function in cells, and reduces potential immunogenicity. In the formulation of CMSPC, the mass ratio of the choline compound, the glycine betaine, and the oligopeptide is precisely controlled to be (2-5):(1-3):(0.5-1.5). For example, in a typical formulation, the mass ratio of the three can be set to 3:2:1. The preparation process of CMSPC includes mixing and dissolving the three components in ultrapure water or ethanol solution according to the predetermined mass ratio. To achieve a uniform solution state, the mixture is stirred by a magnetic stirrer at a constant speed of 200 rpm for 30 minutes, ensuring that all components are fully dissolved and uniformly dispersed. After dissolution is complete, the obtained CMSPC solution can be stored at low temperature (e.g., 4°C) in the dark to maintain its activity and stability.

[0069] Further, the phospholipid precursor, such as choline chloride, is efficiently converted into phosphatidylcholine by the endogenous phosphatidylcholine synthesis pathway in rice cells. Phosphatidylcholine is the main lipid component of eukaryotic cell membranes, and the promotion of its biosynthesis directly enhances the lipid turnover and repair capacity of cell membranes, which is crucial for maintaining the integrity and fluidity of cell membranes under low temperature. The glycine betaine, as a natural compatible solute, accumulates in rice cells under low temperature stress conditions. Its mechanism of action is to stabilize the conformation of intracellular proteins by forming a hydrogen bond network and effectively maintaining the osmotic balance of the cell. This stabilizing effect helps to reduce the membrane lipid phase transition temperature induced by low temperature, thereby maintaining the fluidity and functionality of the cell membrane under low temperature conditions. The hydrophobic nature of the oligopeptide enables it to embed in the lipid bilayer by hydrophobic interaction, enhancing the mechanical strength and stability of the membrane; at the same time, the oligopeptide can bind to the membrane surface through hydrogen bonding or electrostatic interaction, further strengthening the membrane structure. In addition, the oligopeptide can also act as a signal molecule to induce the expression of endogenous cold-resistant genes in rice, enhance the conformational stability of membrane proteins related to stress resistance, and promote the activity of aquaporins, thereby maintaining the normal transmembrane transport of substances and physiological metabolic functions of cells under low temperature. These three components play a significant synergistic role in rice cells, from the aspects of lipid composition, osmotic regulation, and membrane structure and function regulation, to build a comprehensive cell membrane resistance to low temperature stress.

[0070] As a preferred embodiment of the present application, the selection and preparation of the nanocarrier system (NCS) in step b) is crucial for the targeted delivery and protection of CMSPC. The present application uses chitosan-alginate core-shell structure nanoparticles as carriers. The chitosan is selected as medical grade chitosan with a degree of deacetylation between 85% and 95% and a molecular weight between 50 kDa and 190 kDa. A high degree of deacetylation ensures good solubility and positive charge characteristics in acidic conditions, and a moderate molecular weight ensures biocompatibility and degradability. The alginate is in the form of sodium salt, with a molecular weight between 10 kDa and 200 kDa, and a ratio of guluronic acid (G) to mannuronic acid (M) (G / M ratio) between 1.5 and 2.0. A high G / M ratio of alginate forms a more stable gel structure, which is beneficial for forming a strong outer shell. The preparation of the core-shell structure nanoparticles is realized by ionotropic gelation, and the specific steps include: b1) Preparation of chitosan solution: Dissolve the chitosan in an acetic acid solution with a concentration of 1.0% (v / v) at a concentration of 1.0 mg / mL to 3.0 mg / mL. To ensure that the chitosan is completely dissolved and forms a uniform solution, the mixture is continuously stirred by a magnetic stirrer at a speed of 400 rpm for 12 hours.

[0071] b2) Preparation of sodium tripolyphosphate (TPP) solution: The sodium tripolyphosphate is dissolved in ultrapure water to prepare a TPP solution with a concentration of 0.25 mg / mL to 1.0 mg / mL. TPP, as a cross-linking agent, forms nanoparticles core with positively charged chitosan through electrostatic interaction.

[0072] b3) Preparation of sodium alginate solution: The sodium alginate is dissolved in ultrapure water to prepare a sodium alginate solution with a concentration of 1.0 mg / mL to 3.0 mg / mL. Sodium alginate will coat the nanoparticle core as a shell material.

[0073] b4) Formation of CMSPC-loaded nanoparticle core: This is a critical step. The TPP solution pre-loaded with the CMSPC is added dropwise into the chitosan solution under continuous stirring (e.g., 800 rpm) at a precise rate of 0.5 mL / min to 2.0 mL / min by using a peristaltic pump or a syringe pump. The concentration of CMSPC in the TPP solution is 0.5 mg / mL to 2.0 mg / mL. This mixing process is continued for 30 minutes to 60 minutes to allow the positively charged chitosan to rapidly form CMSPC-loaded nanoparticle core with positive charge through electrostatic interaction with the negatively charged TPP. The whole mixing process is carried out at a constant temperature of 25 °C, and the pH value of the solution is maintained between 4.5 and 5.5, which is beneficial for the protonation of chitosan and effective cross-linking with TPP.

[0074] b5) Formation of core-shell structure: The sodium alginate solution is added dropwise into the CMSPC-loaded nanoparticle core suspension at a rate of 0.5 mL / min to 2.0 mL / min by using a peristaltic pump or a syringe pump. The negatively charged sodium alginate solution forms a stable core-shell structure with the positively charged nanoparticle core through electrostatic interaction, in which the alginate layer uniformly coats the outside of the chitosan-CMSPC nanoparticle core, finally forming CMSPC nanoparticles with negative charge. The core-shell formation process is continued for 30 minutes to 60 minutes to ensure complete coating. Similar to the previous step, this mixing process is also carried out at a constant temperature of 25 °C.

[0075] b6) Purification and characterization: The obtained CMSPC nanoparticle suspension is purified by centrifugation (e.g., at 15,000 rpm for 30 minutes) and ultrafiltration (e.g., using an ultrafiltration membrane with a molecular weight cut-off of 100 kDa) to effectively remove unreacted substances and free CMSPCs, ensuring product purity. The purified nanoparticles are comprehensively characterized: the average particle size distribution and polydispersity index (PDI) of the nanoparticles are measured by a dynamic light scattering (DLS) instrument, with the particle size range preferably controlled between 50 nm and 200 nm and the PDI less than 0.2, indicating that the nanoparticles have good uniformity and dispersibility. The surface charge of the nanoparticles is measured by a Zeta potential analyzer, with the surface charge ranging from -20 mV to -40 mV, ensuring its stability in water bodies and interaction characteristics with cell membranes. The micro-morphology and internal core-shell structure of the nanoparticles are observed by transmission electron microscopy (TEM), confirming their morphological integrity. The chemical composition of the nanoparticles is analyzed by Fourier transform infrared spectroscopy (FTIR), verifying the successful combination of each component. The thermal stability of the nanoparticles and the actual loading amount of CMSPCs are determined by thermal gravimetric analysis (TGA). In addition, the unloaded CMSPCs in the supernatant after purification are quantitatively detected by high-performance liquid chromatography (HPLC) or ultraviolet-visible spectrophotometry, and then the encapsulation efficiency of CMSPCs is accurately calculated, which is not less than 80%, proving the high loading capacity of the carrier system for CMSPCs.

[0076] Further, to enhance the selective recognition and uptake efficiency of the CMSPC nanoparticles by rice root cells, the surface of the chitosan-alginate core-shell structure nanoparticles is chemically modified. The chemical modification is achieved by covalently linking a specific targeting ligand to the surface of the nanoparticles. The specific targeting ligand is a glucose or arabinose derivative, and specifically preferably 2-deoxy-2-(2,5-dicarboxy-1,4-dihydrobenzyl) acetylglucosamine or 3-amino-3-deoxy-D-arabinose, which is selected because it can mimic the natural molecules recognized by specific receptors or transporters on the cell membrane of rice root cells. The connection of the targeting ligand is achieved by carbodiimide chemistry (EDC / NHS coupling reaction), which is a mature and efficient bio-coupling method. The specific steps are as follows: after the purification of the CMSPC nanoparticles, the nanoparticles are uniformly dispersed in a MES buffer solution (pH 6.0). Then, an appropriate amount of 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride (EDC) and N-hydroxysuccinimide (NHS) is added, and EDC and NHS work together to activate the carboxyl groups exposed on the surface of the nanoparticles under mild conditions to form stable active ester intermediates. After activation is complete, the glucose or arabinose derivative is added, and the amino group thereof forms a stable amide bond with the activated carboxyl group to covalently graft the targeting ligand to the surface of the nanoparticles. The coupling reaction is carried out at room temperature for 4 to 8 hours to ensure sufficient reaction. After the reaction is completed, the modified nanoparticles are again thoroughly purified by centrifugation and ultrafiltration to remove all unreacted reagents and byproducts, ensuring product purity. The grafting density of the glucose or arabinose derivative on the surface-modified nanoparticles is controlled by precisely controlling the reaction conditions, so that 50 to 150 targeting ligand molecules are carried per 100 nm2 of nanoparticle surface area, and this grafting density is confirmed by X-ray photoelectron spectroscopy (XPS) or Fourier transform infrared spectroscopy (FTIR) to ensure optimal targeting effect. The glucose or arabinose derivative can bind to specific sugar transporters or receptors expressed on the cell membrane of rice root cells with high affinity, thereby inducing receptor-mediated endocytosis or specific transporter-mediated uptake, significantly improving the transport efficiency and selectivity of the nanoparticles to rice root cells, while minimizing the uptake of non-targeted cells, thereby reducing the potential impact on other non-target organisms (such as symbiotic crabs) in the paddy field water, ensuring the safety of the rice-crab breeding ecosystem.

[0077] As an embodiment of the present application, in step c), the application of CMSPC nanoparticles is uniform and precise. The CMSPC nanoparticles are uniformly dispersed in the paddy field water body at a predetermined concentration. The predetermined concentration is defined according to the effective active ingredient concentration of CMSPC in the CMSPC nanoparticles, and the effective active ingredient concentration per liter of water body is set to 1 mg to 5 mg. This concentration range is optimized to ensure sufficient biological activity and avoid resource waste. The uniform dispersion is achieved by an automatic application system, which includes a multi-rotor unmanned aerial vehicle or a ground mobile robot equipped with a high-precision liquid droplet spraying system. The unmanned aerial vehicle or robot is equipped with a corrosion-resistant liquid storage tank with a capacity of 50 L to 200 L, which is connected to a peristaltic pump or gear pump with a flow control accuracy of ± 2%. The pump is used to deliver the CMSPC nanoparticle suspension to the nozzle array at a constant flow rate. The nozzle array is composed of multiple flat fan nozzles, which operate at a pressure of 0.2 MPa to 0.5 MPa, and the droplet size is accurately controlled between 50 µm to 150 µm, ensuring that the droplets can uniformly cover the surface of the rice leaf and efficiently fall into the paddy field water body, thereby enabling the nanoparticles to effectively settle in the water body area around the rice roots. The unmanned aerial vehicle or robot provides centimeter-level positioning accuracy through the real-time kinematic positioning (RTK-GPS) system and inertial measurement unit (IMU), and combines with the preset path planning algorithm to achieve precise and seamless coverage of the target paddy field area, ensuring that the uniformity deviation of CMSPC nanoparticles in the entire paddy field is less than 10%. The application timing is determined based on the following principles: when the local area daily average temperature is expected to drop below 15°C according to the weather forecast system, or when the early signs of low temperature stress appear in the physiological monitoring data of rice plants, the system will automatically start the application program. During the continuous low temperature stress, the application frequency is set to once every 3 to 5 days to maintain the effective concentration of CMSPC in rice cells and continuously provide low temperature protection.

[0078] As an embodiment of the present application, in step d), the CMSPC nanoparticles are selectively absorbed by the rice root cells and realized through a series of precise biological mechanisms. The surface-modified CMSPC nanoparticles settle around the rice roots in the paddy field water, and the glucose or arabinose derivatives on the surface can specifically bind to the specific sugar transporters or receptors expressed on the cell membrane of the rice root cells with high affinity. The binding then triggers receptor-mediated endocytosis, so that the nanoparticles are completely taken into the interior of the root cells to form endosomes. Another possible absorption mechanism is that the nanoparticles interact with specific channel proteins on the cell membrane, promoting the nanoparticles or the CMSPC released outside the cells to enter the cytoplasm through transmembrane transport. Once in the cytoplasm, the endosomes will be regulated by intracellular pH (for example, gradually acidified from the extracellular neutral environment to the acidic environment in the lysosome, pH 4.5-5.5) and enzyme activity (such as lysosomal enzymes), and gradually degraded to accurately release the encapsulated CMSPC. The biocompatible materials of the nanoparticles, i.e. chitosan and alginate, will gradually decompose into non-toxic small molecular oligosaccharides under the action of intracellular enzymes (for example, chitinase and alginatease present in the vacuoles or lysosomes of plant cells). These oligosaccharides can then be metabolized by rice cells or excreted out of the body through the cell efflux mechanism, thereby realizing the complete biodegradability of the nanoparticles and ensuring the safety of the rice cells themselves and the harmlessness to the entire paddy field ecological environment.

[0079] As a preferred embodiment of the present application, the present application also provides a cell membrane stabilization system for improving the low-temperature adaptability of rice in rice-crab breeding. This system is a highly integrated and intelligent platform that includes six core modules that work together to achieve comprehensive monitoring, intelligent decision-making, and precise application of the rice-crab breeding environment. The six modules are: a CMSPC nanoparticle preparation module, an environmental parameter sensing module, a rice physiological response monitoring module, a crab ecological state monitoring module, a central control and decision-making module, and an automated application module.

[0080] Specifically, the CMSPC nanoparticle preparation module is the foundation of the entire system, and its core function is to mass-produce high-quality CMSPC nanoparticles in a standardized manner. This module is configured inside a production workshop with an ISO 7 cleanliness level to ensure aseptic and high-purity production processes. It contains the following key components: The raw material storage unit is used to safely store high-purity chitosan, sodium alginate, sodium tripolyphosphate, choline compounds, glycine betaine, amino acid monomers, and various solvents (such as acetic acid, ultrapure water, and ethanol). The storage unit is equipped with a precise temperature and humidity control system (e.g., temperature control range 15°C-25°C, humidity control range 40%-60% RH) and an inert gas protection system (e.g., nitrogen or argon) to effectively prevent the oxidation, deliquescence, or degradation of the raw materials, thereby ensuring the long-term stability of the raw materials and consistency between batches.

[0081] The CMSPC synthesis reactor is usually a stainless steel jacketed reactor with a volume customizable between 10 L and 100 L, which has precise temperature (±0.1°C), pH (±0.01 pH), and stirring speed (±1 rpm) control functions. The reactor is used for the precise mixing and dissolution of choline compounds, glycine betaine, and oligopeptides in CMSPC, ensuring that the three form a homogeneous solution in a predetermined mass ratio.

[0082] The nanoparticle synthesis reactor is a reactor equipped with a high-shear homogenizer (rotation speed up to 20,000 rpm) or an ultrasonic disperser (power adjustable range 100W-1000W). The reactor is used to mix the chitosan solution with the TPP solution and form the CMSPC-loaded nanoparticle core under precise control, and then add the sodium alginate solution to complete the coating of the core-shell structure. The reactor is integrated with a high-precision sampling pump (accuracy ±0.1 mL / min) and an online particle size monitoring system (e.g., a probe based on dynamic light scattering principle), which can monitor the growth and particle size distribution of the nanoparticles in real time during the reaction, ensuring that the product meets the predetermined specifications.

[0083] The surface modification reactor is usually a reactor equipped with nitrogen protection (to avoid oxidation), a constant temperature circulating water bath (temperature control accuracy ±0.5°C), and magnetic stirring functions. The reactor is used to efficiently and uniformly graft the target ligand on the surface of the purified nanoparticles, ensuring the stability and efficiency of the coupling reaction.

[0084] The separation and purification unit includes a set of continuous flow centrifuges (rotation speed up to 20,000 rpm, processing capacity up to 1 L / min to 10 L / min) and a set of tangential flow ultrafiltration systems (membrane molecular weight cutoff 100 kDa, filtration area 0.5 m² to 5 m²). These devices work together to efficiently remove unreacted substances, free CMSPC, and reaction byproducts, and concentrate the nanoparticle suspension to improve the efficiency of subsequent freeze-drying.

[0085] The freeze-drying unit, using an industrial-grade vacuum freeze dryer, has the functions of automatic temperature control (-80°C to 0°C) and vacuum degree adjustment (1 Pa to 100 Pa). This unit is used to efficiently freeze-dry the purified and concentrated nanoparticle suspension into a stable powder product, greatly facilitating long-term storage and long-distance transportation of the product, and prolonging its shelf life.

[0086] The quality control and detection unit is equipped with a series of advanced analytical detection equipment, including DLS instrument, Zeta potential analyzer, TEM, FTIR, HPLC, XPS, etc. These devices are used for comprehensive quality detection and characterization of all raw materials, intermediate products and final CMSPC nanoparticles, to ensure that each batch of product meets strict preset specification standards, thereby ensuring the stability and reliability of the final application effect.

[0087] The environmental parameter perception module is the "eyes" of the system to obtain macroscopic environmental data of the rice field. This module integrates multiple types of high-precision sensors, including: The distributed weather monitoring station network is composed of multiple (for example, 10 per hectare) independent wireless sensor nodes, achieving fine-grained monitoring of the weather environment of the rice field area. Each node integrates high-precision digital temperature sensors (for example, Pt100 RTD, measurement range -10°C to 50°C, accuracy ±0.1°C), humidity sensors (measurement range 0-100% RH, accuracy ±2% RH), wind speed and direction sensors (measurement range 0.3-30 m / s, accuracy ±5%), and rainfall sensors (resolution 0.2 mm). These sensors provide real-time and accurate weather data.

[0088] The water quality monitoring buoy, multiple (for example, 3 per hectare) self-powered buoys, each carrying multi-parameter water quality sensors, including: pH electrode (measurement range 2-12, accuracy ±0.02 pH), dissolved oxygen (DO) sensor (for example, optical dissolved oxygen sensor, measurement range 0-20 mg / L, accuracy ±0.1 mg / L), conductivity sensor (measurement range 0-200 mS / cm, accuracy ±1%), and turbidity sensor (measurement range 0-1000 NTU, accuracy ±1%). These buoys continuously monitor the key physicochemical parameters of the rice field water, providing basic data for rice growth and crab survival.

[0089] The solar radiation sensor, each monitoring station and buoy is equipped with a photosynthetically active radiation (PAR) sensor (measurement range 0-3000 µmol / m² / s, accuracy ±5%), which is used to accurately measure the light intensity as an important input for evaluating the photosynthetic efficiency of rice.

[0090] The data transmission unit, all sensor nodes and buoys are connected with the central data collection gateway through LoRaWAN or NB-IoT wireless communication protocol. The communication protocol provides low power consumption, long distance, safe data transmission capability, transmission rate is 2.5 kbps to 50 kbps, data transmission delay is less than 5 seconds, ensures the effectiveness of real-time data. All data transmission adopts AES-128 bit encryption standard, guarantees data security and privacy.

[0091] The rice physiological response monitoring module aims to accurately assess the physiological health status and response degree of rice under low temperature stress through non-invasive means. The module mainly includes: The unmanned aerial or ground robot carried hyperspectral imaging system is equipped with a hyperspectral camera with a spectral range of 400 nm to 1000 nm, a spectral resolution better than 5 nm, and a spatial resolution of 5 cm / pixel. The system is used to periodically collect the reflectance spectral data of rice plants. By performing advanced analysis on these spectral data, a series of physiological indicators including normalized difference vegetation index (NDVI), photochemical reflectance index (PRI), water vapor stress index (CWSI), and chlorophyll content, nitrogen content, etc. are calculated, so as to comprehensively and objectively evaluate the growth state of rice and the degree of low temperature stress.

[0092] The unmanned aerial or ground robot carried chlorophyll fluorescence measurement system is equipped with a pulse amplitude modulation (PAM) fluorometer. The system is used for non-invasive measurement of chlorophyll fluorescence parameters of rice leaves, including maximum photochemical quantum efficiency (Fv / Fm), actual photochemical efficiency (ΦPSII), and non-photochemical quenching (NPQ). The measurement system provides modulated light (e.g., 650 nm) through a high-brightness LED light source and accurately detects the fluorescence signal at 730 nm. These chlorophyll fluorescence parameters can directly reflect the functional state of the photosynthetic system of rice and its damage and recovery under low temperature stress.

[0093] The unmanned aerial or ground robot carried thermal infrared imaging system is equipped with a high-sensitivity thermal infrared camera (e.g., NETD less than 50 mK) with a spatial resolution better than 10 cm / pixel. The system is used to accurately measure the surface temperature of rice leaves, which is a key indicator of the efficiency of rice transpiration and serves as a direct indicator of rice water stress and cold stress.

[0094] The data synchronization and integration unit, all imaging and measuring systems are connected to the on-board or vehicle-mounted data processor through high-speed Ethernet (e.g., Gigabit Ethernet). The processor is configured with at least 8-core ARM Cortex-A series CPU and 16 GB RAM for real-time preprocessing and compression of massive raw data. The processed data is securely and efficiently transmitted to the central control and decision module through a dedicated encrypted wireless link (e.g., 5G cellular network or WiFi in a proprietary frequency band).

[0095] The crab ecological status monitoring module focuses on non-invasively monitoring the health status and behavior patterns of crabs in the paddy field water, ensuring that the application of CMSPC nanoparticles does not have a negative impact. This module includes: The distributed underwater visual monitoring system consists of multiple (e.g., 5 per hectare) low-illumination high-definition (1080p) underwater cameras with IP68 waterproof rating. The cameras are equipped with wide-angle lenses (e.g., 120° field of view) and infrared fill light, allowing continuous and clear monitoring of crab activities, feeding, molting behavior, and any abnormal conditions even at night or in low light conditions. The cameras have built-in motion detection algorithms that automatically trigger high-frame-rate recording and upload key video segments to the central control module for analysis when detecting abnormal crab activity patterns.

[0096] The underwater acoustic sensor array consists of multiple (e.g., 3 per hectare) high-sensitivity hydrophones (frequency response range 20 Hz to 20 kHz) for capturing underwater acoustic signals such as crab crawling, feeding, and biological vocalization. Through spectral analysis of these acoustic signals, the crab's activity level, foraging intensity, and stress response can be indirectly evaluated as auxiliary indicators of its health status.

[0097] The water metabolite analysis unit automatically extracts paddy field water samples periodically (e.g., once a day) and uses a portable multi-parameter analyzer or sends the water samples to the laboratory for in-depth analysis. The analysis focuses on crab stress biomarkers such as ammonia nitrogen content, nitrate content, nitrite content, and specific hormone (e.g., molting hormone) metabolite concentrations in the water. These indicators serve as auxiliary indicators for evaluating the health status of crabs, with 50 mL of water samples taken for detection each time.

[0098] The data transmission unit connects the underwater camera and acoustic sensor to the shore-based data processor through underwater optical fiber or Ethernet cable, and transmits data to the central control and decision module through an encrypted wireless link.

[0099] The central control and decision module is the core intelligent unit of the entire system, responsible for integrated processing of all monitoring data, complex low-temperature stress prediction, application decision-making, and issuing instructions to the automated application module. Its configuration includes: The high-performance data integration server is equipped with a multi-core high-performance CPU (e.g., Intel Xeon series), at least 64 GB DDR4 ECC RAM, and 2 TB NVMe SSD storage, used to receive, store, and process real-time big data streams from all monitoring modules. The server runs a stable Linux operating system and deploys a distributed database (e.g., Cassandra or MongoDB) to efficiently handle high-concurrency data writing and query requests.

[0100] The big data analysis and prediction engine is based on advanced deep learning models (e.g., long short-term memory network LSTM) and ensemble learning models (e.g., random forest or gradient boosting tree). The model uses historical meteorological data, rice physiological parameters, crab behavior data, and CMSPC nanoparticle application effect data as the training set, with the following core functions: First, low-temperature stress prediction can accurately predict the likelihood, intensity, and duration of low-temperature stress in the rice field area within 24 to 72 hours, with a prediction accuracy of over 85%, providing a basis for early prevention.

[0101] Second, CMSPC nanoparticle application strategy optimization calculates the optimal CMSPC nanoparticle application time, dose, and frequency based on the predicted low-temperature stress level and the current physiological state of the rice, combined with historical application effect data, aiming to minimize the application amount while maximizing the protection effect.

[0102] Third, ecological risk assessment continuously assesses water quality and crab physiological behavior data, predicts the potential impact of CMSPC nanoparticle application on crabs, and adjusts the application strategy or issues a warning to the operator in real time when any potential negative impact is found.

[0103] The decision support system is an expert system based on a rule engine and a fuzzy logic controller. This system receives the output of the prediction engine and combines pre-set agronomic management rules, environmental safety thresholds (e.g., water pH should not be lower than 6.5, and dissolved oxygen should not be lower than 4 mg / L), and crab protection strategies to generate final application instructions or recommendations. The system has a multi-objective optimization algorithm to balance the optimal solution between rice cold resistance improvement and rice-crab ecological health.

[0104] The human-machine interface (HMI), a web-based graphical user interface, provides secure access through encrypted HTTPS protocol. The interface provides real-time data visualization functions (e.g., map overlay displaying sensor data, trend charts), early warning information push (e.g., SMS, email), historical data query, application plan viewing and editing, and manual instruction input and emergency stop functions, facilitating user operation and management.

[0105] The communication interface unit communicates with the automated application module through MQTT or RESTful API protocols, ensuring accurate issuance of decision instructions and real-time feedback of execution status. All communication links use TLS 1.2 or higher encryption protocols.

[0106] The automated application module is the execution unit of the system that converts decisions into actual actions, responsible for precise application of CMSPC nanoparticles to rice fields. The module includes: The multi-rotor unmanned aerial vehicle or ground robot, as previously described, is equipped with RTK-GPS and IMU, providing centimeter-level positioning and attitude control accuracy, ensuring autonomous navigation and precise operation in complex rice field environments. The unmanned aerial vehicle or robot has a redundant power system and a fault safety mechanism to ensure the reliability and safety of operation.

[0107] The precision liquid spray system includes a liquid storage tank made of corrosion-resistant, ultraviolet-resistant material with a capacity of 100 L to 500 L. The tank body is equipped with an agitator to ensure that the CMSPC nanoparticle suspension remains uniformly dispersed throughout the application process, preventing sedimentation. The storage tank is connected to a high-precision metering pump (e.g., peristaltic pump or diaphragm pump) with a flow control range of 0.5 L / min to 10 L / min and a control accuracy of ±0.5%, ensuring the accuracy of the application dose. The pump delivers the suspension to the nozzle array through a set of high-pressure hoses.

[0108] The nozzle array and droplet control system, the array consists of multiple (e.g., 8 to 16) independently controllable electric atomizing nozzles. Each nozzle can adjust the spray pattern (e.g., fan or cone), droplet size (e.g., 50 µm to 200 µm), and spray flow to adapt to different wind speeds and application requirements. The droplet control system can adjust nozzle parameters in real time according to the instructions of the central control module, ensuring that droplets can be precisely deposited in the target area (rice root water layer or leaf surface), maximizing deposition efficiency, minimizing drift loss, and reducing environmental pollution.

[0109] The task management and execution unit, the flight control computer or motion controller built-in the drone or robot, runs a real-time operating system (RTOS). This unit is responsible for parsing the application instructions issued by the central control module, executing the flight path planning, adjusting the spraying parameters, and collecting and feeding back the sensor data (such as heading, altitude, and speed) in real time, to ensure the accurate execution of the task.

[0110] The power management system, the drone uses high-energy-density lithium polymer batteries, with a continuous operation time of 30 to 60 minutes, supporting fast charging or battery replacement. The ground robot uses high-capacity lithium-ion battery packs, with a continuous operation time of 8 to 16 hours. Both are equipped with automatic charging or battery replacement systems to support long-term continuous operation and improve work efficiency.

[0111] Example 1: Evaluation of rice growth and physiological response under low-temperature stress based on the method of the present application To verify the effectiveness of the cell membrane stabilization method and system for improving the low-temperature adaptability of rice in rice-crab farming, we conducted a three-week field test at a typical rice-crab farming demonstration base in our province. The base is located in the northern climate zone and often experiences sudden low-temperature frost in the fall. The test plot was divided into three areas: the example group, the comparative example group, and the blank control group, with three repeated plots in each group, each with an area of 200 square meters.

[0112] The example group was managed using the cell membrane stabilization method and system described in the present application. First, the preparation of CMSPC nanoparticles strictly followed steps a) and b) above, with the mass ratio of CMSPC set at choline compound: glycine betaine: oligopeptide = 3:2:1. The average particle size of the nanoparticles was controlled at 120 nm, the PDI was 0.15, the surface charge was -30 mV, and the encapsulation efficiency reached 92%. The surface-modified glucose derivative grafting density was 100 molecules per 100 nm² of nanoparticle surface. When the central control and decision-making module predicted that the average daily temperature would drop below 13°C for 3 consecutive days within the next 48 hours, the automated application module was activated. A multi-rotor drone carried a 100 L capacity of CMSPC nanoparticle suspension (with an effective CMSPC active ingredient concentration of 3 mg / L), and applied 20 L of the suspension per hectare at a rate of 12 hours before the temperature dropped. The application frequency was set to once every 4 days, and a total of 3 times during the low-temperature period. The drone applied droplets with a size of 80 µm to ensure uniform coverage of the rice field water body.

[0113] The comparative example group adopted the traditional cold-proof measures commonly used in current rice-crab breeding mode, i.e. increasing the water depth of the rice field to 20 cm before the arrival of low temperature to improve the water body heat preservation effect, and supplemented with a small amount of foliar spraying of silicon fertilizer (a mixture of potassium dihydrogen phosphate and potassium silicate, total concentration 0.1%). This group did not apply the CMSPC nanoparticles of the present application.

[0114] The blank control group did not take any additional cold-proof measures, but only followed the conventional rice-crab breeding management.

[0115] After the occurrence of low temperature stress, we monitored the physiological indicators of rice plants in the three groups continuously, and evaluated the final yield. The main monitoring indicators include: relative electrical conductivity of rice leaves (reflecting the degree of cell membrane damage), chlorophyll content (reflecting photosynthetic capacity), malondialdehyde (MDA) content (reflecting lipid peroxidation level) and final rice yield. At the same time, through the crab ecological state monitoring module, the activity and survival rate of crabs in each group of water were monitored.

[0116] The experimental results show that: In terms of leaf relative electrical conductivity, the blank control group showed the highest relative electrical conductivity under low temperature stress, reaching 48.2%, indicating that the cell membrane was severely damaged. The comparative example group reduced the relative electrical conductivity to 36.8% by increasing the water depth and foliar spraying of silicon fertilizer, but the effect was limited. The relative electrical conductivity of the example group was significantly reduced to 22.5%, which shows that the CMSPC nanoparticles of the present application can significantly stabilize the cell membrane of rice and effectively reduce the damage caused by low temperature stress.

[0117] In terms of chlorophyll content, the rice leaf chlorophyll content of the example group was the highest, reaching 2.55 mg / g FW, which was significantly higher than that of the blank control group (1.85 mg / g FW) and the comparative example group (2.10 mg / g FW), which indicated that the stability of the cell membrane helped to maintain the integrity and function of the photosynthetic organs of rice, thereby maintaining a high photosynthetic efficiency.

[0118] In terms of malondialdehyde (MDA) content, as an important indicator of lipid peroxidation, the MDA content of the blank control group was the highest, reaching 14.7 nmol / g FW. The comparative example group decreased to 10.5 nmol / g FW. The MDA content of the example group was the lowest, only 6.2 nmol / g FW, which strongly proved that the CMSPC nanoparticles could effectively inhibit the membrane lipid peroxidation reaction induced by low temperature and protect the cell membrane from oxidative damage.

[0119] In terms of final rice yield, the rice yield of the example group reached 6600 kg / ha, which was significantly higher than that of the blank control group (5200 kg / ha) and the comparative example group (5850 kg / ha). This indicates that by improving the cold tolerance of rice, the present application can effectively guarantee and even increase the yield of rice and improve economic benefits.

[0120] In terms of crab ecological status, the survival rate and average weight gain rate of crabs in each group were maintained at a high level, and there was no significant difference (P > 0.05) between the example group and the blank control group and the comparative example group. This fully verifies the biological safety of the chitosan-alginate nanocarrier and its surface modification used in the present application in the rice-crab breeding environment, which has no negative impact on the symbiotic crabs and their ecological environment.

[0121] In summary, the present example clearly proves, through quantitative data, that the cell membrane stabilization method and system provided by the present application for improving the cold tolerance of rice in rice-crab breeding can significantly improve the cell membrane integrity, photosynthetic efficiency and antioxidant capacity of rice under low temperature stress, ultimately achieving a significant increase in rice yield, while ensuring the health and stability of the rice-crab breeding ecosystem. This multi-level, synergistic membrane stabilization mechanism, as well as the precise application strategy based on an intelligent system, effectively solves the limitations of traditional cold protection measures and provides an innovative and sustainable technical solution for the sustainable development of the rice-crab breeding industry.

Claims

1. A method for improving the cell membrane stability of rice at low temperature in rice crab breeding, characterized by, The method comprises the following steps: a) preparing a cell membrane stabilizing precursor complex (CMSPC); b) preparing a nano-carrier system (NCS) and efficiently loading the CMSPC inside the NCS to form a CMSPC nanoparticle; c) uniformly dispersing the CMSPC nanoparticle in the rice field water body at a preset concentration; d) applying the CMSPC nanoparticle, which is selectively absorbed by the rice root cells and enters the cells, and then releases the CMSPC in the cells, so that the CMSPC exerts a synergistic effect inside the rice cells, thereby improving the integrity and functional stability of the rice cell membrane under low-temperature stress.

2. The method of claim 1, wherein, In step a), the cell membrane stabilizing precursor complex (CMSPC) is composed of a phospholipid precursor, an osmotic regulator, and an oligopeptide. The phospholipid precursor is a choline compound with a purity of not less than 99.5% and a molecular weight range of 139.62 g / mol to 253.25 g / mol. The osmotic regulator is glycine betaine with a purity of not less than 98.0% and a molecular weight of 117.15 g / mol. The oligopeptide is a synthetic tetrapeptide or pentapeptide with a molecular weight range of 300 Da to 600 Da, which is prepared by solid-phase synthesis method with a purity of not less than 95%. In the CMSPC, the mass ratio of the choline compound, the glycine betaine, and the oligopeptide is (2-5):(1-3):(0.5-1.5).

3. The method of claim 2, wherein, The phospholipid precursor is specifically choline chloride or hydrogen tartrate choline. The preparation of the CMSPC is by mixing and dissolving the three components in ultrapure water or ethanol solution according to the preset mass ratio, and stirring with a magnetic stirrer at a speed of 200 rpm for 30 minutes until a uniform solution is formed.

4. The method of claim 1, wherein, In step b), the nano-carrier system is composed of chitosan-alginate core-shell structure nanoparticles; The chitosan is a medical grade chitosan with a degree of deacetylation between 85% and 95% and a molecular weight between 50 kDa and 190 kDa; The alginate is in the form of sodium salt, with a molecular weight between 10 kDa and 200 kDa, and a ratio of mannuronic acid to guluronic acid between 1.5 and 2.0; The surface of the chitosan-alginate core-shell structure nanoparticles is chemically modified by covalently linking a specific targeting ligand to the surface of the chitosan; The specific targeting ligand is a glucose or arabinose derivative, specifically 2-deoxy-2-(2,5-dicarboxy-1,4-dihydrobenzyl) acetylamino glucose or 3-amino-3-deoxy-D-arabinose.

5. The method of claim 4, wherein, The core-shell structure nanoparticles are prepared by ion gel method, comprising the following steps: b1) Preparation of chitosan solution: the chitosan is dissolved in acetic acid solution with a concentration of 1.0% (v / v), a chitosan solution with a concentration of 1.0 mg / mL to 3.0 mg / mL is prepared, and stirred by a magnetic stirrer at a speed of 400 rpm for 12 hours; b2) Preparation of sodium tripolyphosphate (TPP) solution: the sodium tripolyphosphate is dissolved in ultrapure water, a TPP solution with a concentration of 0.25 mg / mL to 1.0 mg / mL is prepared; b3) Preparation of sodium alginate solution: the sodium alginate is dissolved in ultrapure water, a sodium alginate solution with a concentration of 1.0 mg / mL to 3.0 mg / mL is prepared; b4) Formation of CMSPC nanoparticle core loading: in the continuously stirred chitosan solution, the CMSPC-containing TPP solution is added dropwise by a peristaltic pump or a syringe pump at a rate of 0.5 mL / min to 2.0 mL / min, the concentration of CMSPC in the TPP solution is 0.5 mg / mL to 2.0 mg / mL, the mixing process lasts for 30 minutes to 60 minutes, the mixing process is carried out at a constant temperature of 25°C, and the pH value of the solution is maintained between 4.5 and 5.5; b5) Formation of core-shell structure: in the CMSPC-loaded nanoparticle core suspension, the sodium alginate solution is added dropwise by a peristaltic pump or a syringe pump at a rate of 0.5 mL / min to 2.0 mL / min, the core-shell formation process lasts for 30 minutes to 60 minutes, and the mixing process is carried out at a constant temperature of 25°C; b6) Purification and characterization: the obtained CMSPC nanoparticles are purified by centrifugation and ultrafiltration to remove unreacted substances and free CMSPC; the particle size distribution and polydispersity index of the nanoparticles are measured by a dynamic light scattering instrument, the particle size range is 50 nm to 200 nm, and the PDI is less than 0.2; the surface charge of the nanoparticles is measured by a Zeta potential analyzer, the surface charge range is -20 mV to -40 mV; the encapsulation efficiency of the CMSPC is not less than 80%; The connection of the targeting ligand is achieved by carbodiimide chemistry, the coupling reaction is carried out at room temperature for 4 hours to 8 hours, and the grafting density of the glucose or arabinose derivative is 50 to 150 targeting ligand molecules per 100 nm² of nanoparticle surface area.

6. The method of claim 1, wherein, In the step c), the preset concentration refers to the effective active ingredient concentration of CMSPC in the CMSPC nanoparticles, which is 1 mg to 5 mg per liter of water body; The uniform dispersion is achieved by an automatic application system, which includes a multi-rotor unmanned aerial vehicle or a ground mobile robot (61) equipped with a high-precision liquid droplet spraying system; The unmanned aerial vehicle or robot (61) is equipped with a liquid storage tank with a capacity of 50 L to 200 L, which is connected to a peristaltic pump or gear pump with a flow control accuracy of ±2%, which is used to deliver the CMSPC nanoparticle suspension to the nozzle array (63); The nozzle array (63) is composed of multiple flat fan nozzles, which operate at a pressure of 0.2 MPa to 0.5 MPa, producing droplet sizes between 50 µm and 150 µm; The unmanned aerial vehicle or robot (61) achieves precise coverage of the target rice field area through a real-time differential positioning system and an inertial measurement unit, ensuring that the application uniformity deviation of CMSPC nanoparticles in the entire rice field area is less than 10%; The application timing is determined based on the following principles: when the weather forecast shows that the local area's daily average temperature is expected to drop below 15°C, or when the physiological monitoring data of rice plants shows early signs of low temperature stress; The application frequency is once every 3 to 5 days during continuous low temperature stress.

7. A cell membrane stabilization system for improving low temperature adaptability of rice in rice crab breeding, characterized by, The system comprises: CMSPC nanoparticle preparation module (1): for preparing the CMSPC nanoparticles of claim 1; Environmental parameter sensing module (2): for real-time monitoring of meteorological parameters of the rice field water body and environment; Rice physiological response monitoring module (3): for non-invasively assessing the degree of low temperature stress and physiological health status of rice; Crab ecological state monitoring module (4): for non-invasively monitoring the health status and behavior patterns of crabs in the rice field water body; Central control and decision module (5): for integrated processing of all monitoring data, low temperature stress prediction, application decision, and issuing instructions to the automated application module (6); Automated application module (6): for accurately dispersing CMSPC nanoparticles into the rice field water body according to the instructions of the central control and decision module (5).

8. The system of claim 7, wherein, The production workshop of the CMSPC nanoparticle preparation module (1) is internally configured with: Raw material storage unit (11): for storing high-purity chitosan, sodium alginate, sodium tripolyphosphate, choline compounds, glycine betaine, amino acid monomers, and solvents, equipped with a temperature and humidity control system and an inert gas protection system; CMSPC synthesis reactor (12): a stainless steel jacketed reactor with precise temperature, pH, and stirring speed control functions, with a volume of 10 L to 100 L; Nanoparticle synthesis reactor (13): a reactor equipped with a high-shear homogenizer or ultrasonic disperser, with a sample injection pump and an online particle size monitoring system; Surface modification reactor (14): a reactor equipped with nitrogen protection, constant temperature circulating water bath, and magnetic stirring functions; Separation and purification unit (15): including a set of continuous flow centrifuge and a set of tangential flow ultrafiltration system, with a membrane molecular weight cutoff of 100 kDa and a filtration area of 0.5 m² to 5 m²; Freeze-drying unit (16): an industrial-grade vacuum freeze dryer with automatic temperature control and vacuum degree adjustment functions; Quality control and testing unit (17): equipped with DLS instrument, Zeta potential analyzer, TEM, FTIR, HPLC, and XPS equipment.

9. The system of claim 7, wherein, The environmental parameter perception module (2) comprises: Distributed weather monitoring station network (21): composed of multiple independent wireless sensor nodes, each node integrating high-precision digital temperature sensor, humidity sensor, wind speed and direction sensor, and rainfall sensor; Water quality monitoring buoy (22): multiple self-powered buoys, each buoy carrying multiple-parameter water quality sensors, including: pH electrode, dissolved oxygen sensor, conductivity sensor, and turbidity sensor; Solar radiation sensor (23): each monitoring station and buoy is equipped with a photosynthetically active radiation (PAR) sensor; Data transmission unit (24): all sensor nodes and buoys are connected to the central data collection gateway through LoRaWAN or NB-IoT wireless communication protocol, which provides low-power, long-distance, secure data transmission, transmission rate is 2.5 kbps to 50 kbps, data transmission delay is less than 5 seconds, data transmission uses AES-128 bit encryption standard.

10. The system of claim 7, wherein, The rice physiological response monitoring module (3) comprises: Unmanned aerial or ground robot-mounted hyperspectral imaging system (31): equipped with a hyperspectral camera with a spectral range of 400 nm to 1000 nm, a spectral resolution of better than 5 nm, and a spatial resolution of 5 cm / pixel; Unmanned aerial or ground robot-mounted chlorophyll fluorescence measurement system (32): equipped with a pulse amplitude modulation fluorometer for measuring chlorophyll fluorescence parameters of rice leaves, the measurement system provides modulated light through an LED light source and detects 730 nm fluorescence signals; Unmanned aerial or ground robot-mounted thermal infrared imaging system (33): equipped with a high-sensitivity thermal infrared camera with a spatial resolution of better than 10 cm / pixel; Data synchronization and integration unit (34): all imaging and measurement systems are connected to the onboard or vehicle-mounted data processor through high-speed Ethernet, the processor has at least 8-core ARM Cortex-A series CPU and 16 GB RAM, and the processed data is transmitted to the central control and decision module (5) through a special encrypted wireless link.

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