A long-acting brassinolide compound and its application

Abstract

This invention provides a long-acting brassinolide compound and its application, the structure of which is shown in Formula I or Formula II. The long-acting brassinolide compound provided by this invention, when used in tea cultivation, can effectively increase tea yield and improve tea quality, exhibiting superior technical effects compared to other reagents.

Description

Technical Field

[0001] This invention belongs to the agricultural field, specifically relating to a long-acting brassinolide compound and its applications. Background Technology

[0002] Brassinolides are a class of highly active plant endogenous hormones with a steroidal skeleton. They were isolated from rapeseed by the U.S. Department of Agriculture in 1979, and their structure was determined by XRD analysis, leading to their name, brassinolides. At the 16th International Congress of the Plant Growth Substances Society in 1998, brassinolides were formally recognized as the sixth class of plant hormones, following auxins, gibberellins, and cytokinins. Their chemical structure is similar to that of animal steroids, a characteristic that distinguishes them from other plant growth hormones.

[0003] Brassinolides possess extremely strong physiological activity, promoting plant growth and division even at low concentrations, and are virtually ubiquitous within plants. They function throughout the entire growth and developmental stage of plants, participating in physiological processes such as seed germination and flowering. Furthermore, brassinolides are essential for resisting certain external stressors, such as cold, drought, soil salinization, and heavy metals (aluminum, nickel). Research has also revealed that the effects of brassinolide steroids are not limited to plants; these steroids have shown potential medicinal effects in animal models, such as neuroprotective effects, antioxidant properties, anticancer activity, and cholesterol-lowering effects. Simultaneously, brassinolides are recognized as harmless to plants, the environment, humans, and livestock, and are highly beneficial agents for producing high-quality agricultural products and increasing yields. However, currently registered brassinolide compounds are mainly 24-epibrassinolide and 28-homobrassinolide. With their widespread application in agriculture, some problems have also been exposed in their application. For example, synthetically produced 24-epibrassinolide and 28-homobrassinolide work well after spraying, but their effectiveness diminishes quickly. This is advantageous for crops that require rapid application, but for crops that need long-term application, multiple sprays are necessary, increasing both usage and labor costs.

[0004] In 1993, Takatsuto (WO9428011A1) reported a brassinolide compound with a long-lasting effect, named propionyl brassinolide. This compound significantly improved upon the short duration of action of brassinolides in agricultural applications. Studies of existing bioactivity reports of brassinolide compounds revealed that the absolute configuration at the C2 and C3 positions is crucial. Therefore, highly selective generation of chiral configurations at the C2 and C3 positions is essential in the synthetic route, specifically the dihydroxylation step in the synthesis of the dihydroxy compound (22E,24S)-2α,3α-dihydroxy-5α-stigmaster-22-en-6-one.

[0005]

[0006] However, the current problem is that there are few synthetic routes for propionyl brassinolide (related patents: CN1217338A, CN108070017A), and the existing synthetic routes basically use expensive and toxic osmium tetroxide as an oxidant, which will pollute the environment and greatly limit its application. Therefore, there is an urgent need to improve its process and find a preparation method more suitable for industrial production.

[0007] Furthermore, the current brassinolide industry has relatively few types of long-acting brassinolides (related patents: CN101434637A, CN1962688A, CN115716862A), which cannot meet the usage requirements of different regions and different types of crops. Therefore, it is very necessary to develop new long-acting brassinolide drugs, test their activity, and develop their applications in the agricultural field. Summary of the Invention

[0008] To address the shortcomings of existing technologies, the present invention aims to provide a long-acting brassinolide compound and its applications. The long-acting brassinolide compound provided by the present invention, when used in tea cultivation, can effectively improve tea yield and quality, exhibiting superior technical effects compared to other reagents.

[0009] To achieve this objective, the present invention adopts the following technical solution:

[0010] In a first aspect, the present invention provides a long-acting brassinolide compound, the structure of which is shown in Formula I or Formula II:

[0011]

[0012] Wherein, R is selected from any one of substituted or unsubstituted C1-C12 alkyl, substituted or unsubstituted C2-C12 alkenyl, substituted or unsubstituted C2-C12 alkynyl, substituted or unsubstituted C6-C12 aryl, and substituted or unsubstituted C6-C12 heteroaryl.

[0013] C1-C12 respectively represent structures containing one carbon atom, two carbon atoms, three carbon atoms, and so on, without further explanation. Other similar expressions have similar meanings.

[0014] Compared to reagents such as brassinolide, which have a short duration of action in agricultural applications and require frequent application (e.g., daily spraying for several days) to achieve significant effects, the aforementioned long-acting brassinolide compounds with specific structures have a long-lasting effect in tea cultivation. The efficacy of the drug can last for several days after application, and one or several applications can effectively increase tea yield and improve tea quality, demonstrating superior technical effects compared to other reagents.

[0015] Preferably, the substituted group is selected from any one of halogen, hydroxyl, amino, nitro, mercapto, aldehyde, ester, acyl, C1-C6 alkyl, C2-C6 alkenyl or C2-C6 alkynyl.

[0016] Preferably, the substituted group is selected from any one of halogen, hydroxyl, amino, nitro, mercapto, aldehyde, ester, acyl, and C1-C6 alkyl.

[0017] Preferably, R is selected from any one of substituted or unsubstituted C1-C12 alkyl groups and substituted or unsubstituted C6-C12 aryl groups.

[0018] Preferably, R is selected from any one of substituted or unsubstituted methyl, substituted or unsubstituted ethyl, substituted or unsubstituted isopropyl, substituted or unsubstituted phenyl, and substituted or unsubstituted naphthyl.

[0019] Preferably, the long-acting brassinolide compound is selected from any one of the following compounds:

[0020]

[0021] Secondly, the present invention provides the application of the long-acting brassinolide compounds as described above in the preparation of plant growth regulators.

[0022] Thirdly, the present invention also provides the application of the long-acting brassinolide compounds described above in tea production.

[0023] Compared with the prior art, the present invention has the following beneficial effects:

[0024] This invention provides a long-acting brassinolide compound with a specific structure, which can effectively increase tea yield when used in tea cultivation and has superior technical effects compared to other reagents. Detailed Implementation

[0025] To further illustrate the technical means and effects of the present invention, the following describes the technical solution of the present invention in conjunction with preferred embodiments of the present invention. However, the present invention is not limited to the scope of the embodiments.

[0026] Example 1

[0027] This embodiment provides a method for preparing long-acting brassinolide compounds BR03 and BR04, the specific route of which is as follows:

[0028]

[0029] Synthesis of (3S,8S,9S,10R,13R,14S,17R)-17-((2R,5S,E)-5-ethyl-6-methylhept-3-en-2-yl)-10,13-dimethyl-2,3,4,7,8,9,10,11,12,13,14,15,16,17-tetradecano-1H-cyclopentadiene[a]phenanthrene-3-ylmethanesulfonate (5):

[0030] Stigmasterol (5.00 g, 12.1 mmol) was dissolved in dichloromethane (60 mL), and triethylamine (2.45 g, 24.2 mmol) was added. The mixture was cooled to 5 °C, and then methanesulfonyl chloride (2.08 g, 18.1 mmol) was added dropwise. After stirring at the same temperature for 30 min, the stigmasterol was completely consumed. The reaction solution was extracted with saturated brine, and the combined organic phases were washed with saturated sodium bicarbonate solution and saturated brine solution, respectively, and dried over anhydrous sodium sulfate. The solvent was removed under vacuum (40 °C) to obtain the crude product, which was used directly in the next step without purification.

[0031] Synthesis of (1aR,3aR,3bS,5aR,6R,8aS,8bS,10R,10aR)-6-((2R,5S,E)-5-ethyl-6-methylhept-3-en-2-yl)-3a,5a dimethylhexahydrocyclopentadiene[a]cyclopropyl[2,3]cyclopentadiene[1,2-f]naphthalene-10-ol (6):

[0032] Compound 5 was dissolved in 1,4-dioxane (60 mL) and water (20 mL), followed by the addition of potassium bicarbonate (2.42 g, 24.2 mmol). The reaction mixture was heated under reflux (external temperature: 120 °C) for 2.5 h. After cooling, most of the 1,4-dioxane reagent was removed by vacuum concentration, followed by extraction with dichloromethane. The combined organic phases were dried over anhydrous sodium sulfate and concentrated under vacuum. The resulting residue was dissolved in anhydrous ethanol (15 mL) and heated to 65 °C. The mixture was then slowly cooled to ambient temperature with stirring, and then the temperature was lowered to 0 °C. After stirring overnight at 0 °C, the mother liquor was filtered and dried under reduced pressure at 45 °C to obtain a white solid product (4.13 g, yield 80%). 1 H NMR (500MHz, DMSO-d) 6)δ5.14(dd,J=15.1,8.7Hz,1H),5.02(dd,J=15.1,8.7Hz,1H),4.09(s,1H),3.04(s,1H),2.07–1.9 9(m,1H),1.91(d,J=12.5Hz,1H),1.80–1.71(m,1H),1.63(dd,J=10.3,6.7Hz,3H),1.56–1.20(m, 10H),1.20–1.08(m,3H),1.05–0.90(m,9H),0.94–0.89(m,1H),0.82(d,J=6.1Hz,3H),0.77(q,J= 5.2,3.4Hz,7H),0.69(s,3H),0.41(t,J=3.9Hz,1H),0.23(dd,J=7.4,5.1Hz,1H).HRMS(ESI),m / z forC 29 H 49 O + [M+H] + ,Calcd:413.3778,found413.3747.

[0033] Synthesis of (1aR,3aR,3bS,5aR,6R,8aS,8bS,10aR)-6-((2R,5S,E)-5-ethyl-6-methylhept-3-en-2-yl)-3a,5a dimethyltetradecylcyclopentadiene[a]cyclopropyl[2,3]cyclopentadiene[1,2-f]naphthyl-10(1H)-one (7):

[0034] Compound 6 (4.13 g, 10.0 mmol) and silica gel (200-300 mesh, 7.53 g) were reacted in dichloromethane (156 mL) and cooled to 0 °C. Pyridium dichromate (7.53 g, 20.0 mmol) was added in portions over 0.5 h. After addition, the reaction mixture was slowly heated to room temperature. After stirring overnight, the mixture was filtered (with diatomaceous earth added), the filter cake was washed with dichloromethane, and the mother liquor was concentrated under vacuum. The resulting residue was dissolved in anhydrous ethanol (20 mL) and heated to 60 °C. The mixture was slowly cooled to room temperature with stirring, and then the temperature was lowered to 0 °C. After stirring overnight at 0 °C, the solid was collected by filtration and washed with a pre-cooled (0 °C) solution of anhydrous ethanol (5 mL). The solid was dried under reduced pressure at 45 °C to obtain a white solid (3.00 g, 66% yield). 1H NMR(500MHz,Chloroform-d)δ5.12(dd,J=15.2,8.6Hz,1H),5.00(dd,J=15.2,8.6H z,1H),2.42–2.36(m,1H),2.06–1.97(m,2H),1.92–1.82(m,3H),1.81–1.72(m,1H) ,1.72–1.61(m,3H),1.57–1.44(m,5H),1.41(dd,J=28.2,3.6Hz,1H),1.33–1.11(m ,7H),1.10–0.92(m,8H),0.79(dd,J=24.7,6.5Hz,9H),0.70(s,4H).HRMS(ESI),m / z for C 29 H 47 O + [M+H] + ,Calcd:411.3621,found411.3629.

[0035] Synthesis of (5S,8S,9S,10R,13R,14S,17R)-17-((2R,5S,E)-5-ethyl-6-methylhept-3-en-2-yl)-10,13-dimethyl-1,4,5,7,8,9,10,11,12,13,14,15,16,17-tetradecano-6H-cyclopentadien[a]phenanthrene-6-one (3):

[0036] Compound 7 (3.00 g, 7.3 mmol), lithium bromide (634 mg, 7.3 mol), and p-toluenesulfonic acid hydrate (PTSA) (2.48 g, 2.9 mmol) were heated under reflux (external temperature: 155 °C) overnight in N,N-dimethylformamide (49 mL). Extraction was performed with methyl tert-butyl ether, and the organic phases were combined. Most of the methyl tert-butyl ether was concentrated, and the remaining N,N-dimethylformamide was washed with saturated brine. The organic phase was then concentrated to dryness. The resulting residue was dissolved in anhydrous ethanol (2 mL) and heated to 65 °C. The mixture was slowly cooled to room temperature with stirring, and then the temperature was lowered to 0 °C. After stirring overnight at 0 °C, the solid was collected by filtration and washed with a pre-cooled (0 °C) solution of anhydrous ethanol (10.0 mL). The solid was dried under reduced pressure at 45 °C. A white solid product (1.37 g, yield 46%) was given. 1H NMR(500MHz,Chloroform-d)δ5.68(d,J=9.5Hz,1H),5.56(d,J=9.5Hz,1H),5.14(dd,J=15.1,8.6Hz,1H ),5.02(dd,J=15.1,8.7Hz,1H),2.34(dt,J=11.6,4.2Hz,2H),2.30–2.19(m,1H),2.09–1.83(m,6H),1. 73(s,2H),1.65–1.56(m,1H),1.53(dt,J=11.8,6.2Hz,3H),1.47–1.37(m,2H),1.35–1.14(m,6H),1.02 (d,J=6.6Hz,4H),0.84(d,J=6.2Hz,3H),0.80(d,J=9.5Hz,6H),0.67(d,J=10.2Hz,6H).HRMS(ESI),m / z for C 29 H 47 O + [M+H] + ,Calcd:411.3621,found 411.3624.

[0037] Synthesis of (2R,3S,5S,8S,9S,10R,13R,14S,17R)-17-((2R,5S,E)-5-ethyl-6-methylhept-3-en-2-yl)-2,3-dihydroxy-10,13-dimethylhexadecylhydro-6H-cyclopenta[a]phenanthrene-6-one (4):

[0038] Compound 3 (7.00 g, 17.0 mmol), N-methylmorpholine-N-oxide (4.99 g, 42.6 mmol), (DHQD)2-PHAL (265 mg, 0.3 mmol), and potassium osmium tetroxide dihydrate (188 mg, 0.5 mmol) were dissolved in a mixture of acetone (189 mL) and water (42 mL). The reaction mixture was stirred vigorously overnight at room temperature. The reaction mixture was extracted with 10% methanol / dichloromethane, and the combined organic phases were dried over anhydrous sodium sulfate and then concentrated under vacuum. The resulting residue was dissolved in anhydrous ethanol (20 mL) and heated to 80 °C. The mixture was then slowly cooled to room temperature with stirring, and then the temperature was lowered to 0 °C. After stirring overnight at 0 °C, the solid was collected by filtration and washed with a pre-cooled (0 °C) solution of anhydrous ethanol (6 mL). The solid was recrystallized with anhydrous ethanol and dried under reduced pressure at 45°C to obtain a white pure solid product (4.47 g, yield 59%, dr>15:1). 1H NMR(500MHz,Chloroform-d)δ5.13(dd,J=15.1,8.6Hz,1H),5.01(dd,J=15.0,8.6Hz,1H),4.04(s,1H ),3.76(d,J=11.0Hz,1H),2.67(d,J=10.9Hz,1H),2.29(dd,J=13.1,4.2Hz,1H),2.08–1.87(m,5H),1 .81–1.61(m,6H),1.52(dd,J=11.0,5.5Hz,4H),1.44–1.32(m,3H),1.31–1.11(m,6H),1.02(d,J=6.7 Hz,3H),0.83(d,J=6.0Hz,3H),0.79(dd,J=9.4,6.6Hz,6H),0.75(s,3H),0.67(s,3H).HRMS(ESI),m / z for C 29 H 49 O3 + [M+H] + ,Calcd:445.3676,found445.3675.

[0039] Synthesis of (3aS,5S,6R,7aR,7bS,9aS,10R,12aS,12bS)-5,6-dihydroxy-7a,9a-dimethyl-10-((1S)-1-(3-((S)-2-methylpent-3-yl)oxacyclohexane-2-yl)ethyl)hexadecylhydro-3H-benzo[c]indeno[5,4-e]oxacyclopropane-3-one (8):

[0040] 2,2,2-Trifluoroacetic anhydride (1.91 g, 9.1 mmol) was dissolved in chloroform (18 mL) and cooled to 0 °C. A 30% aqueous solution of hydrogen peroxide (761 mg, 6.7 mmol) was added dropwise to the solution. The mixture was stirred at 0 °C for 1 h, and then a chloroform solution of compound 4 (300 mg, 0.7 mmol) (15 mL) was added dropwise, maintaining the temperature of the reaction mixture below 5 °C. After the addition, the reaction mixture was heated to room temperature and stirred for 4 h, then cooled again at 0 °C. Freshly prepared peroxytrifluoroacetic acid (synthesized according to the above method) was added again. The reaction mixture was stirred at room temperature for 24 h. Upon completion of the reaction, sodium thiosulfate pentahydrate was added to quench the hydrogen peroxide solution in the reaction. After the absence of oxidizing properties was detected by starch-potassium iodide test paper, saturated sodium carbonate and saturated sodium bicarbonate were added for washing, followed by extraction with dichloromethane. The combined organic phases were concentrated to dryness. The crude product was subjected to silica gel column chromatography (PE / EA1 / 1) to obtain the product (200 mg, yield 63%). 1H NMR(500MHz,Chloroform-d)δ4.09(s,2H),4.01(s,1H),3.70(d,J=10.6Hz,1H),3.11(d,J=12 .1Hz,1H),2.73(d,J=5.4Hz,1H),2.49(s,2H),2.38–2.24(m,1H),2.13(ddd,J=14.9,12.2,2.4 Hz,1H),2.03–1.91(m,3H),1.86(dd,J=13.1,5.1Hz,2H),1.7–1.62(m,5H),1.55(s,2H),1.46 –1.36(m,2H),1.32(s,2H),1.26(d,J=15.0Hz,4H),1.06–0.84(m,15H),0.69(d,J=5.6Hz,3H).

[0041] Synthesis of (3aS,5S,6R,7aR,7bS,9aS,10R,12aS,12bS)-7a,9a-dimethyl-1-((2R,3R)-3-((S)-2-methylpentan-3-yl)epoxyethylene-2-yl)ethyl)-3-oxohexadecyl-1H-benzo[c]indeno[5,4-e]oxacyclopentan-5,6-diylbis(2-(naphth-1-yl)acetate) (1, Compound BR04):

[0042] Compound 8 (152 mg, 0.3 mmol) was dissolved in dichloromethane (4 mL), and 2-naphthaleneacetic acid (118 mg, 0.6 mmol), dicyclohexylcarbodiimide (198 mg, 1.0 mmol), and the catalyst 4-dimethylaminopyridine (2.0 mg, 0.004 mmol) were added. The reaction mixture was then stirred at 0 °C for 4 h. Most of the dicyclohexylcarbodiimide byproduct was removed by filtration, the mother liquor was evaporated to dryness, and the crude product was purified by silica gel column chromatography to give a white solid product (135 mg, 52% yield). 1H NMR (500MHz, CDCl3) δ7.94 (dd, J=14.0, 8.0Hz, 2H), 7.87 (d, J=8.2Hz, 1H), 7. 81(d,J=8.1Hz,1H),7.74(d,J=8.1Hz,1H),7.56(m,J=25.9,15.5,7.8Hz,4H), 7.48–7.41(m,2H),7.34(dd,J=37.7,6.5Hz,1H),7.30(d,J=7.3Hz,1H)7.16(s ,1H),5.16(s,1H),4.66(d,J=10.2Hz,1H),4.03(d,J=3.0Hz,2H),3.95–3.67( m,4H),2.92(d,J=3.4Hz,1H),2.77(d,J=7.4Hz,1H),2.59(dd,J=13.2,5.3Hz, 2H),2.12–1.99(m,1H),1.96–1.82(m,3H),1.72–1.57(m,3H),1.43(s,3H),1. 39–1.32(m,3H),1.29–1.20(m,3H),1.08–1.01(m,7H),0.98(dd,J=17.6,8.3H z,8H),0.86(t,J=6.8Hz,1H),0.66(s,3H),0.57(d,J=6.7Hz,3H).MS(ESI)m / z 813.58 [M+H] + 835.59 [M+Na] + .

[0043] Synthesis of (3aS,5S,6R,7aR,7bS,9aS,10R,12aS,12bS)-10-((2S,5S)-5-ethyl-3,4-dihydroxy-6-methylheptane-2-yl)-7a,9a-dimethyl-3-oxohexadecylhydro-1H-benzo[c]indeno[5,4-e]oxacyclopenten-5,6-diylbis(2-(naphth-1-yl)acetate) (2, Compound BR03):

[0044] Compound BR04 (106.0 mg, 0.1 mmol) was dissolved in a mixed solvent of 1,4-dioxane and water (v / v 1.0 mL / 0.5 mL), and concentrated sulfuric acid (13.1 mg, 0.13 mmol) was added dropwise. The mixture was stirred at 60 °C and reacted overnight. After the reaction was complete, the pH was adjusted to 8 with saturated sodium bicarbonate solution, and the mixture was extracted with ethyl acetate. The combined organic phases were dried over anhydrous sodium sulfate and purified by silica gel column chromatography (PE / EA1 / 1) to give the target product (29 mg, yield 28%). 1H NMR(500MHz,Chloroform-d)δ7.93(s,2H),7.87(d,J=7.5Hz,1H),7.81(d,J=5.0Hz,1H),7.73(d,J=7.1Hz,1H),7.64–7.48(m,4H),7.43(s,2H),7.3 7(s,1H),7.30(s,1H),7.16(s,1H),5.16(s,1H),4.66(d,J=11.1Hz,1H), 4.12(d,J=7.0Hz,1H),4.02(s,2H),3.91(d,J=17.2Hz,1H),3.74(d,J=14. 6Hz,2H),2.97–2.88(m,1H),2.61(d,J=9.3Hz,1H),2.04(d,J=3.3Hz,2H) ,1.90(t,J=13.8Hz,2H),1.62(d,J=26.8Hz,5H),1.52–1.44(m,2H),1.44– 1.31(m,5H),1.27(d,J=11.8Hz,5H),1.13(dd,J=11.5,7.0Hz,3H),1.00( tt,J=21.0,8.6Hz,12H),0.66(s,3H),0.60(d,J=16.2Hz,3H).MS(ESI)m / z 831.55[M+H] + 853.56 [M+Na] + .

[0045] Example 2

[0046] This embodiment provides a long-acting brassinolide compound BR05, which is prepared using the same synthesis process as BR04 in Example 1. The difference between this embodiment and Example 1 is that the compound 2-naphthaleneacetic acid in Example 1 is replaced with an equimolar amount of isobutyric acid.

[0047] The characterization data are as follows:

[0048] 1H NMR(500MHz, CDCl3)δ5.37(d,J=8.7Hz,1H),4.92–4.83(m,1H,),4.17–3.98(m,2H),2.99(dd,J=12.3,4.4Hz,1H) ,2.73(dd,J=7.3,2.1Hz,1H),2.66–2.56(m,1H),2.53–2.41(m,2H),2.29(t,J=14.7Hz,1H),2.04–1.87(m,4H),1 .85–1.66(m,4H),1.66–1.56(m,2H),1.48–1.33(m,5H),1.32–1.23(m,4H),1.21(dd,J=10.7,7.0Hz,6H),1.12(d ,J=7.0Hz,6H),δ1.02–0.97(m,7H),0.96–0.92(m,6H).0.91(d,J=5.6Hz,3H),0.70(d,J=6.5Hz,3H).MS(ESI)m / z 617.54[M+H] + 639.48 [M+Na] + .

[0049]

[0050] Example 3

[0051] This embodiment provides a long-acting brassinolide compound BR01, which is prepared by referring to the synthesis process of BR03 in Example 1. The difference from Example 1 is that the compound 2-naphthaleneacetic acid in Example 1 is replaced with an equimolar amount of propionic acid.

[0052] The characterization data are as follows:

[0053] 1H NMR (500MHz, CDCl3) δ5.38 (s, 1H), 4.89 (dd, J = 8.3, 2.7Hz, 1H), 4.08 (dd, J = 41.8, 12.3, 9.8Hz, 2H), 3.59 (dd, J = 12.2, 5.2Hz,2H),2.99(dd,J=12.3,4.4Hz,1H),2.40–2.33(m,2H),2.33–2.20(m,3H),2.27–2.17(m,2H),2.16–2.01(m,2H), 1.96–1.86(m,2H),1.82–1.58(m,6H),1.49–1.32(m,4H),1.26(dd,J=10.9,4.3Hz,6H),1.17(t,J=7.5Hz,3H),1.10(t ,J=7.5Hz,3H),1.02(d,J=6.9Hz,3H),0.98(s,3H),0.97–0.92(m,6H),0.87(d,J=6.8Hz,3H),0.74(s,3H).MS(ESI)m / z 607.48[M+H] + 629.45 [M+Na] + .

[0054]

[0055] Example 4

[0056] This embodiment provides a long-acting brassinolide compound BR02, which is prepared using the same synthesis process as BR03 in Example 1. The difference between this embodiment and Example 1 is that the compound 2-naphthaleneacetic acid in Example 1 is replaced with an equimolar amount of isobutyric acid.

[0057] The characterization data are as follows:

[0058] 1H NMR(500MHz,Chloroform-d)δ5.36(s,1H),4.90–4.85(m,1H),4.17–4.00(m,2H),3.62–3.53(m,2H),2.98(dd,J=12 .3,4.4Hz,1H),2.62–2.57(m,1H),2.45(d,J=7.0Hz,1H),2.33–2.24(m,1H),2.13–1.87(m,6H),1.83–1.55(m,6H), 1.49–1.40(m,2H),1.37(s,1H),1.33(s,1H),1.30–1.23(m,6H),1.20(dd,J=10.2,7.0Hz,6H),1.12(d,J=7.0Hz,6H ),1.02(d,J=6.8Hz,3H),0.99(s,3H),0.95(dd,J=9.9,7.2Hz,6H),0.87(d,J=6.8Hz,3H),0.74(s,3H).MS(ESI)m / z 635.39[M+H] + 657.51[M+Na] + .

[0059]

[0060] Effect test:

[0061] The effects of the above compounds BR01-BR05 were tested using the following methods:

[0062] Sample Treatment: The tea garden experiment was conducted in mid-July at the Yingde Base of the Tea Research Institute of Guangdong Academy of Agricultural Sciences (23°N, 113°E). Prior to treatment, the tea plants in the sample plots underwent fertilization, pest control, weed control, and pruning. No insecticides, fungicides, or herbicides were applied during the entire treatment period, and no tea leaves were harvested. Other management practices remained consistent with field management. The compound application concentration and dosage were based on published literature and the effective dosage of commercial propionyl BR, with a comprehensive analysis yielding a concentration of 0.02 mg / L per 667 m². 2 The amount of pesticide sprayed was 50 kg. The area of ​​each treatment group in this experiment was approximately 3 m². 2The compound was applied at a concentration of 0.02 mg / L, with each application consisting of 250 mL of solution. The compound was sprayed during the tea budding stage, once on day 1 (July 12th), once on day 4 (July 16th), and once on day 7 (July 19th), for a total of three applications. Samples were collected on day 21 (August 2nd) to analyze relevant indicators of the tea leaves. The experiment included six treatment groups and two control groups. The experimental groups received 250 mL of water with 50 μL of the novel BR compound (0.1 mg / mL, DMSO for dissolution). Control group CK1 received 250 mL of water with 50 μL of DMSO (final concentration 0.02%), and control group CK2 received 250 mL of water with 50 μL of BR (brassinolide, 0.1 mg / mL, DMSO for dissolution).

[0063] Bud density: The survey was conducted 21 days after the first spraying, with 5 square feet (33cm × 33cm, 0.1m) randomly selected from each treatment. 2 ), to investigate the bud density of tea trees.

[0064] 100-bud weight: Tea branches from the sample plots were harvested 21 days after the first spraying. Five points were randomly selected from each treatment, each measuring 33cm × 33cm (0.1m). 2 The branches are pruned into uniform shapes of one bud and three leaves according to harvesting standards, and then weighed using an electronic balance. The formula for calculating the weight of 100 tea buds is as follows:

[0065]

[0066] Fresh leaf yield: Based on the weight of 100 buds and bud density counted on day 21, the tea yield (fresh weight) is estimated. Then, based on the tea moisture content, the dry weight yield can be estimated. The calculation formula is as follows:

[0067]

[0068] Twenty-one days after exogenous application of propionyl brassinolide, the yield of fresh tea leaves was counted. The yield of fresh leaves was estimated based on the bud density and weight of 100 buds in the quadrat plots. Data are expressed as mean ± standard deviation (n = 5). T1: BR01 compound treatment; T2: BR02 compound treatment; T3: BR03 compound treatment; T4: BR04 compound treatment; T5: BR05 compound treatment; T6: propionyl BR (from Tian J, Lin Y, Han J, et al. Practical synthesis of epocholeone, a plant growth-promoting steroid with long-lasting activity[J]. Tetrahedron Letters, 2024, 140. DOI:10.1016 / j.tetlet.2024.155024, Epocholeone) treatment; CK1: 0.02% DMSO treatment; CK2: BR treatment. Significant differences between the treatment group and the control group (CK1) were calculated using the Student's t-test, expressed as *p<0.05 and **p<0.01. The results are as follows:

[0069] Table 1. Effects of exogenous application of propionyl brassinolide on tea yield

[0070]

[0071] Table 2. Changes in the content of non-ester catechin compounds in tea twigs after exogenous application of propionyl brassinolide

[0072]

[0073] The results above show that the long-acting brassinolide compounds provided by this invention can effectively increase the yield of fresh tea leaves and the catechin content of the product compared with other reagents, demonstrating excellent technical effects.

[0074] The applicant declares that this invention illustrates the long-acting brassinolide compounds and their applications through the above embodiments, but this invention is not limited to the above embodiments, that is, it does not mean that this invention must rely on the above embodiments to be implemented. Those skilled in the art should understand that any improvements to this invention, equivalent substitutions of the raw materials of the product of this invention, addition of auxiliary components, and selection of specific methods, etc., all fall within the protection scope and disclosure scope of this invention.

[0075] The preferred embodiments of the present invention have been described in detail above. However, the present invention is not limited to the specific details in the above embodiments. Within the scope of the technical concept of the present invention, various simple modifications can be made to the technical solution of the present invention, and these simple modifications all fall within the protection scope of the present invention.

[0076] It should also be noted that the various specific technical features described in the above specific embodiments can be combined in any suitable manner without contradiction. In order to avoid unnecessary repetition, the present invention will not describe the various possible combinations separately.

Claims

1. A long-acting brassinolide compound, characterized in that, The long-acting brassinolide compound is selected from any one of the following compounds: 、 、 、 。 2. The application of a long-acting brassinolide compound according to claim 1 in the preparation of plant growth regulators.

3. The application of a long-acting brassinolide compound according to claim 1 in tea production.

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