Functional regulator for enhancing the waterlogging resistance of plants and its preparation method

Abstract

This invention discloses a functional regulator for enhancing plant resistance to waterlogging and its preparation method, belonging to the field of agricultural biotechnology. The functional regulator of this invention is a compound of plant growth regulators, osmotic regulators, nano-silica particles, and adjuvants. The plant growth regulators contain at least melatonin and 2,4-epibrassinolide. This invention comprehensively enhances plant waterlogging resistance through multi-target synergistic effects, addressing four aspects: antioxidant defense, osmotic regulation, photosynthetic recovery, and root protection. The product of this invention can significantly reduce malondialdehyde content in plant leaves under waterlogging stress, increase superoxide dismutase and peroxidase activity, increase proline and soluble sugar content, enhance chlorophyll content and net photosynthetic rate, strengthen root vitality and hydraulic conductivity, and simultaneously exhibit excellent resistance to rainwater erosion. The preparation method of this invention is simple and flexible in application, suitable for both pre-disaster preventative spraying and post-disaster recovery spraying, and applicable to various crops.

Description

Technical Field

[0001] This invention belongs to the field of agricultural biotechnology, specifically relating to a functional regulator that enhances the waterlogging resistance of plants and its preparation method. Background Technology Waterlogging stress is one of the major abiotic stressors limiting agricultural production globally. Statistics show that approximately one-third of the world's arable land suffers periodically from waterlogging, resulting in annual crop yield reductions of 15%-30%, and in severe cases, complete crop failure.

[0002] Under waterlogging stress, soil pores become saturated with water, leading to root hypoxia. Hypoxia inhibits aerobic respiration in roots, while anaerobic respiration intensifies, resulting in the accumulation of toxic substances such as acetaldehyde, ethanol, and lactic acid. Simultaneously, a surge in reactive oxygen species causes membrane lipid peroxidation damage, chlorophyll degradation, decreased photosynthetic efficiency, and inhibited plant growth, ultimately leading to wilting, yellowing, and even death. Therefore, developing products and technologies that effectively enhance plant waterlogging tolerance is of great significance for ensuring safe agricultural production.

[0003] However, existing products and technologies generally suffer from drawbacks such as a single mode of action, poor synergy, weak resistance to rainwater runoff, and a narrow application window. Waterlogging is often accompanied by continuous rainfall, and the active ingredients applied through foliar spraying are easily washed away, significantly reducing their effectiveness. Therefore, there is an urgent need to develop a novel waterlogging control agent that is multi-target synergistic, rainwater erosion resistant, and easy to use. Summary of the Invention

[0004] In view of this, the purpose of this invention is to provide a functional regulator that enhances the plant's resistance to waterlogging and its preparation method. This invention scientifically combines plant growth regulators, osmotic regulators, nano-silicon particles, and adhesion enhancers to construct a synergistic system integrating "antioxidant defense—osmotic regulation—photosynthetic recovery—root protection." Simultaneously, it improves leaf retention rate through anti-erosion adjuvants, significantly enhancing the crop's overall tolerance to waterlogging stress.

[0005] To achieve the above objectives, the present invention provides the following technical solution: This invention provides a functional regulator that enhances the resistance of plants to waterlogging, composed of the following raw materials in parts by weight: Plant growth regulator 0.1-5.5 parts; 5-20 parts of osmotic conditioning substance; 0.5-5 parts of nano-silicon particles; 1-10 parts of adhesion enhancer; Additives: 0.5-5 parts; Solvent 950-990.

[0006] Preferably, the plant growth regulator is selected from at least two of melatonin, 2,4-epibrassinolide, spermidine, chlorocholine-naphthaleneacetic acid, abscisic acid, gibberellin, cytokinin, and methyl jasmonate. More preferably, it is a combination of melatonin and 2,4-epibrassinolide. Melatonin, as a potent antioxidant, can directly scavenge reactive oxygen species and upregulate the expression of antioxidant enzyme genes; 2,4-epibrassinolide can stabilize chloroplast structure, improve photosynthetic efficiency, and promote the accumulation of osmotic regulatory substances. The two work synergistically to enhance waterlogging tolerance through two pathways: antioxidant and photosynthetic recovery.

[0007] More preferably, the plant growth regulator contains 0.1-5 parts by weight of melatonin and 0.01-0.5 parts by weight of 2,4-epibrassinolide. If the melatonin content is too low, the antioxidant effect is not significant; if the content is too high, it may inhibit growth. 2,4-epibrassinolide can exert a highly efficient regulatory effect even at extremely low concentrations.

[0008] Preferably, the osmotic regulator is selected from at least one of γ-polyglutamic acid, trehalose, proline, and betaine; more preferably, it is γ-polyglutamic acid. γ-polyglutamic acid has extremely strong hydrophilicity and water retention capacity, which can effectively maintain cell osmotic potential, and at the same time, it can act as a biostimulant to promote root development.

[0009] Preferably, the average particle size of the nano-silicon particles is 20-200 nm. The nano-silicon particles can enter the plant through leaf stomata, promoting the formation of root aerenchyma, increasing the number of root tips and the expression of aquaporin genes, and improving root water conductivity; simultaneously, they can enhance leaf mechanical strength and reduce reactive oxygen species damage.

[0010] Preferably, the adhesion synergist is a biodegradable short-fiber suspension or a plant-based wax nanoemulsion. This synergist can form a micron-scale fiber network or wax film on the leaf surface, anchoring the active ingredients and significantly improving resistance to rainwater erosion.

[0011] Preferably, the additives include one or more of the following: dispersants (such as sodium dodecyl sulfate, sodium lignosulfonate), emulsifiers (such as Tween-80, Span-80), and pH adjusters (such as citric acid, disodium hydrogen phosphate). The solvent is water or a mixture of water and ethanol (volume ratio 10:1-5:1).

[0012] The present invention also provides a method for preparing the aforementioned functional regulator, comprising the following steps: (1) Dissolve the plant growth regulator in an organic solvent to obtain the regulator stock solution; (2) Add the nano-silicon particles to an aqueous solution containing a dispersant and ultrasonically disperse for 10-60 min to obtain a nano-dispersion. (3) Dissolve the osmotic conditioning substance in water to obtain the osmotic conditioning mother liquor; (4) Mix the solutions obtained in steps (1), (2) and (3), add the adhesion enhancer, and homogenize for 10-20 min; (5) Adjust the pH to 5.5-6.5 with a pH adjuster, filter (80-200 mesh) to obtain the product.

[0013] Preferably, the organic solvent in step (1) is ethanol, and the amount used is 2-5 times the total weight of the plant growth regulator. In step (2), the ultrasonic power is 200-500 W and the frequency is 20-40 kHz. In step (4), the homogenization rotation speed is 5000-10000 rpm.

[0014] This invention also provides the application of the aforementioned functional regulator in improving the waterlogging resistance of plants. Specific application methods include: 1-2 days before waterlogging occurs or within 24 hours after it occurs, dilute the product 200-500 times and spray it evenly on both sides of the leaves until the leaves are moist but not dripping; alternatively, seed soaking treatment (diluted 100-200 times, soaked for 4-8 hours) or root irrigation treatment (diluted 300-500 times, 50-200 mL per plant) can be used.

[0015] Compared with the prior art, the present invention has at least the following beneficial technical effects: This invention is the first to systematically combine four types of active ingredients—melatonin, 2,4-epibrassinolide, γ-polyglutamic acid, and nano-silicon particles—targeting the antioxidant defense system, photosynthetic system, osmotic regulation system, and root aeration system respectively, forming a network-based protective effect with significant synergistic effects, far superior to single components or combinations of two. By introducing short fiber or plant wax-based adhesion enhancers, the retention rate of active ingredients on leaves under waterlogged and rainy conditions is increased by more than three times, solving the problem of traditional foliar fertilizers easily becoming ineffective during the rainy season. It can be used for both pre-disaster prevention and post-disaster emergency treatment; it can be applied as a foliar spray, or by soaking seeds or drenching roots. All components are biodegradable materials, safe for crops, and non-polluting to soil and the environment. It can be produced at room temperature or low temperature, without the need for high temperature and high pressure, making it suitable for industrial production. Detailed Implementation

[0016] Various exemplary embodiments of the present invention will now be described in detail. This detailed description should not be considered as a limitation of the present invention, but rather as a more detailed description of certain aspects, features, and embodiments of the present invention.

[0017] It should be understood that the terminology used in this invention is merely for describing particular embodiments and is not intended to limit the invention. Furthermore, with respect to numerical ranges in this invention, it should be understood that each intermediate value between the upper and lower limits of the range is also specifically disclosed. Every smaller range between any stated value or intermediate value within a stated range, and any other stated value or intermediate value within said range, is also included in this invention. The upper and lower limits of these smaller ranges may be independently included or excluded from the range.

[0018] Unless otherwise stated, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art. While only preferred methods and materials have been described herein, any methods and materials similar or equivalent to those described herein may be used in the implementation or testing of this invention. All references to this specification are incorporated by way of citation to disclose and describe methods and / or materials associated with those references. In the event of any conflict with any incorporated reference, the content of this specification shall prevail.

[0019] Various modifications and variations can be made to the specific embodiments described in this specification without departing from the scope or spirit of the invention, as will be apparent to those skilled in the art. Other embodiments derived from this specification will also be obvious to those skilled in the art. This application specification and embodiments are merely exemplary.

[0020] The terms “include,” “including,” “have,” “contain,” etc., used in this article are all open-ended terms, meaning that they include but are not limited to.

[0021] Unless otherwise specified, "room temperature" and "normal temperature" in this invention refer to 25±2℃.

[0022] Unless otherwise specified, all raw materials or instruments used in the following embodiments of the present invention are commercially available.

[0023] Example 1 This embodiment provides a functional regulator to enhance the ability of plants to resist waterlogging, with the following formula (total weight 1000 parts): 2 parts melatonin, 0.05 parts 2,4-epibrassinolide, 10 parts γ-polyglutamic acid, 2 parts nano-silica particles (average particle size 50 nm), 5 parts adhesion synergist (biodegradable polylactic acid short fiber suspension, fiber length 100-300 μm, solid content 5%), 1 part dispersant (sodium lignosulfonate), 0.5 parts emulsifier (Tween-80), 10 parts ethanol, and deionized water to make up to 1000 parts.

[0024] Preparation method: (1) Melatonin and 2,4-epibrassinolide were added to 10 parts of ethanol and stirred at room temperature until completely dissolved to obtain the conditioning mother liquor. (2) Nano-silicon particles and sodium lignosulfonate were added to 200 parts of deionized water and ultrasonically dispersed for 30 min using an ultrasonic cell disruptor (power 300 W, frequency 25 kHz) to obtain a uniform nano-dispersion. (3) γ-polyglutamic acid was dissolved in 200 parts of deionized water and stirred to obtain the osmotic conditioning mother liquor. (4) The solutions obtained in steps (1), (2) and (3) were combined, and the adhesion synergist and Tween-80 were added. The mixture was homogenized for 15 min using a high-speed homogenizer (8000 rpm). (5) The pH was adjusted to 6.0 with 1 mol / L citric acid solution, and deionized water was added to a total weight of 1000 parts. The mixture was passed through a 120-mesh sieve and packaged to obtain the finished product. The product is a milky white suspension with a pH of 6.0±0.2 and an average particle size of ≤200 nm.

[0025] Example 2 Following the method in Example 1, adjust the allocation ratio of each group. 5 parts melatonin, 0.2 parts 2,4-epibrassinolide, 15 parts γ-polyglutamic acid, 4 parts nano-silicon particles, 8 parts adhesion enhancer, 1.5 parts dispersant, 0.8 parts emulsifier, 15 parts ethanol, and deionized water to make up to 1000 parts.

[0026] Example 3 Following the method in Example 1, adjust the allocation ratio of each group. Melatonin 0.5 parts, 2,4-epibrassinolide 0.02 parts, γ-polyglutamic acid 6 parts, nano-silicon particles 1 part, adhesion enhancer 2 parts, dispersant 0.8 parts, emulsifier 0.3 parts, ethanol 8 parts, deionized water to make up to 1000 parts.

[0027] Example 4 Following the method in Example 1, adjust the allocation ratio of each group. 5 parts melatonin, 5 parts adhesion enhancer, 1 part dispersant, 0.5 parts emulsifier, 10 parts ethanol, and deionized water to make up to 1000 parts.

[0028] Experimental Example 1 Pot experiment to verify the effect The experimental material was the maize variety "Zhengdan 958". After disinfection and germination, the seeds were sown in plastic pots with a substrate of garden soil:vermiculite = 3:1, and 3 seedlings were kept in each pot. Conventional management was carried out until the three-leaf stage.

[0029] The experiment was set up with six treatment groups: The control group (CK) received normal water management (soil moisture content 70%–80%) and was sprayed with water on the leaves. The waterlogging group (WL) placed the pot in the pool with the water level 2 cm above the soil surface and waterlogged for 7 days. On the day the waterlogging started, the leaves were sprayed with clean water. The waterlogging treatment group (WL+FA) of this invention is the same as that of WL. On the day the waterlogging begins, the functional regulator prepared in Example 1 is diluted 300 times and sprayed on the leaves, with 10 mL sprayed per pot. Positive control group 1 (WL+MT) was treated with waterlogging and sprayed with 200 μmol / L melatonin aqueous solution; Positive control group 2 (WL+EBR) was treated with waterlogging and sprayed with 0.1 mg / L 2,4-epibrassinolide solution; Positive control group 3 (WL+PG) was treated with waterlogging and sprayed with 0.2 g / L γ-polyglutamic acid solution; Positive control group 4 (WL+Si) was treated with waterlogging and sprayed with 200 mg / L nano-silica suspension. Each group consisted of 5 pots, with 3 replicates.

[0030] Physiological indicators were measured on the second leaf after waterlogging treatment on the 3rd and 7th days, and root activity was measured on the 7th day. Plant phenotype was also observed.

[0031] The results showed that after 7 days of waterlogging stress, the chlorophyll content (SPAD value) of the treatment group (WL+FA) was 38.4, significantly higher than that of the WL group (18.6), and also higher than that of the WL+MT group (28.3), WL+EBR group (31.5), WL+PG group (20.2), and WL+Si group (24.5). Regarding malondialdehyde (MDA) content, the WL+FA group had 7.5 μmol / g FW, far lower than that of the WL group (18.7 μmol / g FW), and also lower than that of the positive control groups (WL+MT group 11.2, WL+EBR group 12.8, WL+PG group 16.5, WL+Si group 14.2). Regarding superoxide dismutase (SOD) activity, the WL+FA group had 268 U / g FW, significantly higher than the WL group's 112 U / g FW and all positive control groups (WL+MT group 225, WL+EBR group 198, WL+PG group 130, WL+Si group 168). Regarding proline content, the WL+FA group had 178 μg / g FW, between the WL group's 210 and the CK group's 85, but not inferior to any of the positive control groups. Regarding root activity, the WL+FA group had 265 μg / g·h, significantly higher than the WL group's 98 μg / g·h and all positive control groups (WL+MT group 152, WL+EBR group 140, WL+PG group 112, WL+Si group 178). Meanwhile, plant phenotypic photographs showed that in the WL+FA group, only the lower, older leaves showed slight yellowing, while the central leaves remained green, and the plants stood upright; whereas in the WL group, most leaves wilted and turned yellow, and the plants lodged. These results indicate that the product of this invention significantly alleviates waterlogging damage in corn compared to single-component treatments, demonstrating a clear synergistic effect.

[0032] Experiment Example 2 Rainwater erosion resistance test The product prepared in Example 1 was diluted 300 times and sprayed evenly onto the leaves of maize at the three-leaf stage, then allowed to air dry for 1 hour. Subsequently, a simulated rainfall device (nozzle diameter 0.5 mm, water pressure 0.02 MPa, rainfall intensity 20 mm / h) was used for continuous spraying for 15 minutes. Leaves that were not rinsed after spraying were used as a control. The retention rates of the effective components (melatonin as a marker, determined by high-performance liquid chromatography) on the leaf surface were measured before and after rinsing. A control product without the adhesion synergist (all other components were identical to that in Example 1, only the adhesion synergist was removed) was also tested using the same simulated rainfall rinsing method.

[0033] The experimental results showed that the product of Example 1 with added adhesion synergist maintained a melatonin retention rate of 82.5% on the leaf surface after simulated rainfall rinsing, with a standard deviation of 3.2%. In contrast, the control product without the adhesion synergist maintained only 24.3% melatonin under the same rinsing conditions, with a standard deviation of 2.8%. The difference between the two groups was statistically significant (P<0.01) according to the t-test. These results indicate that the adhesion synergist used in this invention can significantly improve the product's resistance to rainwater erosion under waterlogged and rainy conditions, ensuring long-term adhesion of the active ingredients to the leaf surface, thereby solving the problem of traditional foliar fertilizers being easily washed away and becoming ineffective during the rainy season.

[0034] Experimental Example 3 Effects of application on different crops Following the waterlogging treatment method in Example 3, pot experiments were conducted on soybeans (variety "Zhonghuang 13", four-leaf stage), tomatoes (variety "Fenguan", five-leaf stage), and peppers (variety "Xiangyan 15", six-leaf stage). On the day of waterlogging treatment, a 300-fold diluted solution of the product prepared in Example 1 was sprayed, 10 mL per pot, and the waterlogging lasted for 7 days. A control group treated only with water was used. Each treatment consisted of 5 pots, with 3 plants per pot, and 3 replicates. After the waterlogging treatment, the aboveground fresh weight (grams per plant) and plant survival rate (%) were measured.

[0035] In soybean trials, the average aboveground fresh weight of the control group under waterlogging was 2.85 grams per plant, with a survival rate of 73.3%; while the average aboveground fresh weight of the treatment group of this invention was 5.92 grams per plant, with a survival rate of 96.7%. In tomato trials, the average aboveground fresh weight of the control group under waterlogging was 8.43 grams per plant, with a survival rate of 66.7%; while the average aboveground fresh weight of the treatment group of this invention was 15.86 grams per plant, with a survival rate of 93.3%. In pepper trials, the average aboveground fresh weight of the control group under waterlogging was 3.12 grams per plant, with a survival rate of 60.0%; while the average aboveground fresh weight of the treatment group of this invention was 7.45 grams per plant, with a survival rate of 90.0%. These results indicate that the product of this invention significantly improved biomass and survival rate under waterlogging stress in three different crops: soybean, tomato, and pepper, demonstrating its broad applicability across a wide range of crops. This invention is also applicable to rice. From a mechanistic perspective, although rice is a wetland plant, it is still intolerant of prolonged and complete flooding. Under waterlogging stress, root hypoxia leads to ethanol accumulation, reactive oxygen species (ROS) bursts, and premature leaf senescence. The melatonin in this invention can effectively remove excessive ROS accumulated in rice roots and leaves; 2,4-epibrassinolide can stabilize chloroplast ultrastructure and delay waterlogging-induced leaf senescence; and nano-silicon particles can promote the development of root aerenchyma and adventitious root formation, thereby improving oxygen transport efficiency. This multi-target synergistic effect comprehensively enhances rice's tolerance to waterlogging stress from three levels: antioxidant defense, photosynthetic system protection, and enhanced root aeration.

[0036] The above description is only a preferred embodiment of the present invention. It should be noted that those skilled in the art can make several improvements and modifications without departing from the principle of the present invention, and these improvements and modifications should also be considered within the scope of protection of the present invention.

Claims

1. A functional regulator that enhances the resistance of plants to waterlogging, characterized in that, It consists of the following raw materials in parts by weight: Plant growth regulator 0.1-5.5 parts; 5-20 parts of osmotic conditioning substance; 0.5-5 parts of nano-silicon particles; 1-10 parts of adhesion enhancer; Additives: 0.5-5 parts; Solvent 950-990.

2. The functional regulator according to claim 1, characterized in that, The plant growth regulator is selected from at least two of the following: melatonin, 2,4-epibrassinolide, spermidine, chlorocholine-naphthaleneacetic acid, abscisic acid, gibberellin, cytokinin, and methyl jasmonate.

3. The functional regulator according to claim 2, characterized in that, The plant growth regulator is a combination of melatonin and 2,4-epibrassinolide; the weight parts of melatonin are 0.1-5 parts, and the weight parts of 2,4-epibrassinolide are 0.01-0.5 parts.

4. The functional regulator according to claim 1, characterized in that, The osmotic regulating substance is selected from at least one of γ-polyglutamic acid, trehalose, proline, and betaine.

5. The functional regulator according to claim 1, characterized in that, The average particle size of the silicon nanoparticles is 20-200 nm.

6. The functional regulator according to claim 1, characterized in that, The adhesion enhancer is a biodegradable short-fiber suspension or a plant-based wax nanoemulsion.

7. The functional regulator according to claim 1, characterized in that, The additives include one or more of dispersants, emulsifiers, and pH adjusters; the solvent is water or a mixture of water and ethanol.

8. A method for preparing the functional regulator according to any one of claims 1-7, characterized in that, Includes the following steps: (1) Dissolve the plant growth regulator in an organic solvent to obtain the regulator stock solution; (2) Add the nano-silicon particles to an aqueous solution containing a dispersant and ultrasonically disperse for 10-60 min to obtain a nano-dispersion. (3) Dissolve the osmotic conditioning substance in water to obtain the osmotic conditioning mother liquor; (4) Mix the solutions obtained in steps (1), (2) and (3), add the adhesion enhancer, and homogenize for 10-20 min; (5) Adjust the pH to 5.5-6.5 with a pH adjuster, filter, and the product is obtained.

9. The preparation method according to claim 8, characterized in that, The organic solvent mentioned in step (1) is ethanol, and the amount used is 2-5 times the total weight of the plant growth regulator.

10. The application of the functional regulator according to any one of claims 1-7 in improving the waterlogging resistance of plants, characterized in that, The applications include foliar spraying, seed soaking, or root irrigation. When foliar spraying, dilute 200-500 times and spray evenly on both sides of the leaves 1-2 days before waterlogging or within 24 hours after waterlogging.