Structural modification based on l-proline and its synthetic method and application

Abstract

This invention relates to a bioactivator based on L-proline structural modification, its synthesis method, and its application, belonging to the field of agricultural chemistry. The activator employs a biomimetic molecular design strategy, prepared through the splicing of active structural units. L-proline's unique natural source advantage and the important active unit fragment N,N-diethylethanolamine from DA-6 undergo an esterification reaction to yield 2-(diethylamino)ethyl-L-proline ester. Bioactivity experiments were conducted on wheat and soybean, respectively. The results showed that this compound can promote plant growth at low concentrations, exhibiting significantly better growth effects than the blank control in terms of germination rate, root length, and stem length. Furthermore, the preparation process of this invention is simple, low-cost, and easily scalable for industrial production, and it possesses good environmental compatibility. Its unique natural source advantage provides a new option for activator development and has significant potential for agricultural development.

Description

Technical Field

[0001] This invention relates to the field of agrochemical technology, specifically to bioactivators based on L-proline structure modification, their synthesis methods, and applications. Background Technology

[0002] Bioactivators are a class of functional products derived from natural biological resources, such as amino acids, microbial metabolites, and plant extracts, prepared through biosynthesis, extraction and purification, or structural modification. Their core function is to synergistically regulate plant growth and development, enhance nutrient absorption efficiency, and improve stress resistance, while simultaneously reducing reliance on chemical pesticides and fertilizers, thus aligning with the needs of green agriculture. Their research background involves multiple fields, including plant physiology, molecular biology, agricultural science, and ecology.

[0003] L-proline is a naturally occurring cyclic imine that can be widely distributed in plants in its free state. Its solubility in aqueous solution is greater than that of any other amino acid, making it not only an ideal osmotic regulator but also playing a crucial role in plant growth, development, and stress resistance. However, unoptimized natural amino acids are easily degraded by plant enzymes, resulting in a short-lived effect of only 3-7 days, requiring frequent application. Further optimization is needed in terms of controllability, diversity, and efficiency. Currently, 2-(diethylamino)ethyl-L-proline esters have been designed and synthesized using the important active fragment of N,N-diethylaminoethanol in DA-6 to study its plant regulatory activity. However, no relevant literature reports the biological activity of this type of compound. Summary of the Invention

[0004] This invention addresses the problems of existing technologies where the natural amino acid L-proline has not undergone structural optimization, is easily degraded rapidly by plant enzymes, has a short duration of action, requires frequent application, and needs further improvement in controllability, diversity, and efficiency. This invention aims to synthesize a novel compound through a biomimetic molecular design strategy to enrich the molecular library of plant activators and provide a new candidate drug molecule.

[0005] Another objective of this invention is to provide a simple method for synthesizing the above-mentioned compounds, as well as their application parameters in wheat and soybeans, to provide technical support for the development of green agrochemical products.

[0006] To achieve the above-mentioned technical objectives, the technical solution of the present invention is as follows: A bioactivator based on L-proline structure modification, with the following structural formula: ; The synthesis method of bioactivators based on L-proline structure modification is as follows: ; Prepared via the following steps: N,N-diethylethanolamine and L-proline were mixed and dissolved in toluene. The catalyst was then added to the reaction system, followed by heating. Water produced in the reaction system was separated using a water separator. After the reaction was completed, the heating was turned off, and the reaction system was allowed to cool to room temperature. The organic phase was filtered to remove insoluble matter, and the solvent was removed by vacuum distillation. The crude product was purified by silica gel column chromatography to obtain the target product 2-(diethylamino)ethyl-L-proline ester.

[0007] The catalyst is one of concentrated H2SO4, KH2PO4, p-toluenesulfonic acid, and titanium oxysulfate.

[0008] The molar ratio of N,N-diethylethanolamine, L-proline and catalyst is 1:(0.8-1.2):(0.04-0.06).

[0009] The heating reaction is carried out at a temperature of 110-130℃ for 4-8 hours.

[0010] The application of bioactivators based on L-proline structure modification allows them to be used as active or activating components to regulate plant growth activity.

[0011] Preferably, it is used as an activator for wheat and soybeans.

[0012] Preferably, when used as a plant activator, the appropriate concentration for promoting the germination and growth of wheat and soybeans is 5-60 ppm.

[0013] Compared with the prior art, the present invention has the following beneficial effects: 1. This invention employs a biomimetic molecular design strategy, preparing 2-(diethylamino)ethyl-L-proline ester through the splicing of active structural units. Leveraging the unique natural source advantage of L-proline, it undergoes an esterification reaction with N,N-diethylethanolamine, an important active unit fragment in DA-6, to yield 2-(diethylamino)ethyl-L-proline ester. The synthesized 2-(diethylamino)ethyl-L-proline ester is a novel compound, enriching the molecular library of this class and providing a promising drug candidate molecule for exploring novel activators.

[0014] 2. The 2-(diethylamino)ethyl-L-proline ester of the present invention has good water solubility, with a solubility of 26 g / L at 25°C, making it convenient for practical production applications.

[0015] 3. The 2-(diethylamino)ethyl-L-proline ester of the present invention has a good effect on the germination and growth of wheat and soybean, and significantly improves the efficiency of plant regulatory activity. Detailed Implementation

[0016] The technical solution of the present invention will be clearly and completely described below with reference to specific embodiments. Obviously, the described embodiments are only some embodiments of the present invention, and not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of the present invention. Example 1

[0017] This embodiment provides a method for synthesizing a bioactivator based on L-proline structure modification, which is achieved through the following preparation steps: L-proline (0.4 mol), N,N-diethylethanolamine (0.4 mol), toluene (100 mL), and concentrated H₂SO₄ catalyst (0.02 mmol, 0.05 eq) were added to a 250 mL two-necked flask. The mixture was then heated to 120 °C and reacted for 6 h. Water produced in the reaction system was separated using a water separator. After the reaction was complete, the heating was turned off, and the system was allowed to cool to room temperature. The organic phase was filtered to remove insoluble matter, and the solvent was removed by vacuum distillation. The crude product was purified by silica gel column chromatography (n-hexane / ethyl acetate = 1:1) to obtain the target product 2-(diethylamino)ethyl-L-proline ester in 75% yield. Example 2

[0018] This embodiment provides a method for synthesizing a bioactivator based on L-proline structure modification, which is achieved through the following preparation steps: L-proline (0.32 mol), N,N-diethylethanolamine (0.4 mol), toluene (100 mL), and catalyst KH₂PO₄ (0.016 mol, 0.04 eq) were added to a 250 mL two-necked flask. The mixture was then heated to 110 °C and reacted for 8 h. Water produced in the reaction system was separated using a water separator. After the reaction was complete, the heating was turned off, and the system was allowed to cool to room temperature. The organic phase was filtered to remove insoluble matter, and the solvent was removed by vacuum distillation. The crude product was purified by silica gel column chromatography (n-hexane / ethyl acetate = 1:1) to obtain the target product 2-(diethylamino)ethyl-L-proline ester in 80% yield. Example 3

[0019] This embodiment provides a method for synthesizing a bioactivator based on L-proline structure modification, which is achieved through the following preparation steps: L-proline (0.48 mol), N,N-diethylethanolamine (0.4 mol), toluene (100 mL), and the catalyst p-toluenesulfonic acid (0.023 mol, 0.06 eq) were added to a 250 mL two-necked flask. The mixture was then heated to 130 °C and reacted for 4 h. Water produced in the reaction system was separated using a water separator. After the reaction was complete, the heating was turned off, and the system was allowed to cool to room temperature. The organic phase was filtered to remove insoluble matter, and the solvent was removed by vacuum distillation. The crude product was purified by silica gel column chromatography (n-hexane / ethyl acetate = 1:1) to obtain the target product 2-(diethylamino)ethyl-L-proline ester in 85% yield. Example 4

[0020] This embodiment provides a method for synthesizing a bioactivator based on L-proline structure modification, which is achieved through the following preparation steps: L-proline (0.4 mol), N,N-diethylethanolamine (0.4 mol), toluene (100 mL), and the catalyst titanium oxysulfate (0.02 mol, 0.05 eq) were added to a 250 mL two-necked flask. The mixture was then heated to 120 °C and reacted for 6 h. Water produced in the reaction system was separated using a water separator. After the reaction was complete, the heating was turned off, and the system was allowed to cool to room temperature. The organic phase was filtered to remove insoluble matter, and the solvent was removed by vacuum distillation. The crude product was purified by silica gel column chromatography (n-hexane / ethyl acetate = 1:1) to obtain the target product 2-(diethylamino)ethyl-L-proline ester in 96% yield.

[0021] 1H NMR spectrum: 1 H NMR (400 MHz, Deuterium Oxide) d 4.64–4.51 (m, 3H), 3.55 (dd, J = 5.5, 4.6 Hz, 2H), 3.46–3.37 (m, 2H), 3.32–3.23 (m, 4H), 2.46 (ddt, J = 13.2, 8.7, 6.7 Hz, 1H), 2.18 (ddt, J = 13.0, 8.5, 7.2 Hz, 1H), 2.10–2.01 (m, 2H), 1.28 (td, J = 7.3, 1.4 Hz, 6H). Carbon NMR spectrum: 13 C NMR (101 MHz, DeuteriumOxide) d 169.0, 60.5, 59.2, 49.8, 47.9, 46.2, 27.9, 23.2, 7.9. High-resolution mass spectrometry: HRMS (positive ESI) m / z: calcd for C11 H 23 N2O2 + [M + H] + , 215.1754; found, 215.1760.

[0022] Effect Examples Bioactivity Test

[0023] Preparation of 2-(diethylamino)ethyl-L-proline ester activator: 0.5 g of the activator (prepared in Example 1) and DA-6 were weighed and diluted with tap water to 500 mL to obtain a 1 g / L activator stock solution. These solutions were then diluted to concentrations of 1 ppm, 5 ppm, 10 ppm, 20 ppm, 40 ppm, 60 ppm, and 80 ppm. Tap water treatment served as the control (CK).

[0024] b. Seedling cultivation: Wheat and soybean seeds of uniform size and plumpness were selected and placed in 25 mL beakers, with 30 seeds in each beaker. The prepared activator was poured into the beakers containing the seeds, and the seeds were soaked for 6 hours. After soaking, the seeds were evenly arranged on a moist paper bed for germination until the taproot showed white and the lateral roots just began to emerge. During this process, 1.0 g of agar was accurately weighed using an analytical balance and placed in a beaker containing 2000 mL of water. The water was heated in a microwave oven until completely dissolved, cooled, and then poured evenly into four 500 mL beakers until solidified, preparing the agar medium. The germinated seeds were then planted on the solidified agar and cultured in a 27 ℃ incubator. When the taproot of the control group (CK group) seeds touched the bottom of the beaker, all seedlings were removed and measured.

[0025] c. Measurement Indicators and Methods Seed germination rate: Number of germinated seeds / Total number of seeds Seedling sampling method: All seedlings were randomly sampled, with 10 seedlings randomly selected from each treatment as test samples, and the samples were repeated 3 times to detect the morphological and physiological indicators of the seedlings.

[0026] Plant height: The standard measurement is the length from the highest point of the leaf to the hypocotyl.

[0027] Root length: The measurement method is to select the three longest roots from a seedling and take their average value.

[0028] Dry weight of above-ground and underground parts: The measurement method is to wash the seedlings clean, absorb the surface moisture with filter paper, and divide them into above-ground and underground parts. They are blanched at 105 ℃ for 20 minutes, and then dried at 80 ℃ to constant weight. The dry weight is then measured.

[0029] The results of the bioactivity tests of 2-(diethylamino)ethyl-L-proline ester are shown in Tables 1 and 2.

[0030] ;

[0031] The compound was tested for bioactivity on wheat and soybean. The results showed that: 1. The appropriate concentration for promoting the germination and growth of wheat was 5-60 ppm, and the appropriate concentration for promoting the germination and growth of soybean was 5-60 ppm. The germination and growth effect was much better than that of the blank control and DA-6 group.

[0032] In conclusion, 2-(diethylamino)ethyl-L-proline ester can significantly improve the growth of various crops (such as wheat and soybeans) at appropriate concentrations. Its unique natural source advantage avoids the problem of residual degradation in the soil from the source, and its green and environmentally friendly advantages provide a new option for the development of activators. We have reason to believe that 2-(diethylamino)ethyl-L-proline ester can effectively improve plant growth and lay a certain foundation for improving the yield and quality of agricultural products.

[0033] The above description is only a preferred embodiment of the present invention. It should be noted that for those skilled in the art, several improvements and modifications can be made 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 bioactivator based on L-proline structural modification, characterized in that, Its structural formula is:

2. The method for synthesizing the bioactivator based on L-proline structure modification as described in claim 1, characterized in that... Prepared via the following steps: N,N-diethylethanolamine and L-proline were mixed and dissolved in toluene. The catalyst was then added to the reaction system, followed by heating. Water produced in the reaction system was separated using a water separator. After the reaction was completed, the heating was turned off, and the reaction system was allowed to cool to room temperature. The organic phase was filtered to remove insoluble matter, and the solvent was removed by vacuum distillation. The crude product was purified by silica gel column chromatography to obtain the target product 2-(diethylamino)ethyl-L-proline ester.

3. The method for synthesizing the bioactivator based on L-proline structure modification according to claim 2, characterized in that, The catalyst is one of concentrated H2SO4, KH2PO4, p-toluenesulfonic acid, and titanium oxysulfate.

4. The method for synthesizing the bioactivator based on L-proline structure modification according to claim 2, characterized in that, The molar ratio of N,N-diethylethanolamine, L-proline and catalyst is 1:(0.8-1.2):(0.04-0.06).

5. The method for synthesizing the bioactivator based on L-proline structure modification according to claim 2, characterized in that, The heating reaction is carried out at a temperature of 110-130℃ for 4-8 hours.

6. The application of the bioactivator based on L-proline structure modification as described in claim 1, characterized in that, It can be used as an active ingredient or activator to regulate plant growth activity.

7. The application of the bioactivator based on L-proline structure modification according to claim 6, characterized in that, It is used as an activator for wheat and soybeans.

8. The application of the bioactivator based on L-proline structure modification according to claim 6, characterized in that, When used as a plant activator, the appropriate concentration for promoting the germination and growth of wheat and soybeans is 5-60 ppm.

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