A tea saponin aglycone thiazole derivative, and a preparation method and application thereof

Abstract

This invention discloses a tea saponin thiazole derivative, its preparation method, and its application. The preparation method includes: protecting the hydroxyl group on the tea saponin with tert-butyldimethylchlorosilane to ensure the correct site for subsequent reactions, obtaining a first intermediate; then reducing the aldehyde group on the first intermediate to a hydroxyl group using sodium triacetoxyborohydride, obtaining a second intermediate; subsequently reacting the reduced hydroxyl group with a thiazole-4-carboxylic acid derivative to obtain a third intermediate; and finally removing the tert-butyldimethylchlorosilane from the third intermediate using tetrabutylammonium fluoride, ultimately obtaining the tea saponin thiazole derivative. The tea saponin thiazole derivative prepared by this invention can be used to prepare anti-root-knot nematode drug formulations, exhibiting good control effects against root-knot nematodes, with its anti-root-knot nematode activity being more than 10 times higher than that of tea saponin.

Description

Technical Field

[0001] This invention relates to the field of biopesticide technology for controlling root-knot nematodes in vegetables, and particularly to a tea saponin thiazole derivative, its preparation method, and its application. Background Technology

[0002] Vegetable root-knot nematode disease has led to a significant reduction in vegetable yields, causing enormous economic losses and severely restricting the development of the vegetable planting industry. Among the control methods, resistance breeding has a long cycle and low success rate; high-temperature killing is difficult to implement on large areas of land; biological control is not very effective; and chemical control has problems such as environmental pollution, pesticide residues, and nematode resistance.

[0003] Tea saponin is the main active ingredient in camellia seed cake, possessing surface activity and antibacterial activity. Reports indicate that tea saponin has a certain inhibitory effect on root-knot nematodes, showing some effect over a long period, but the effect is weak in the short term (Chen Lianghong. Study on the nematicidal activity of tea saponin against southern root-knot nematodes [J]. Agricultural Disaster Research, 2018, 8(01): 1-5.). This suggests that it has significant development potential, but there are no reports on how to improve its effect on vegetable root-knot nematodes through structural modification. Summary of the Invention

[0004] In order to overcome the above-mentioned shortcomings and deficiencies of the prior art, the purpose of this invention is to provide a tea saponin thiazole derivative, which has more than 10 times higher activity against root-knot nematodes than tea saponin.

[0005] Another objective of this invention is to provide a method for preparing the above-mentioned tea saponin thiazole derivative, which is simple, can be industrialized, and is easy to promote and apply.

[0006] Another object of the present invention is to provide the application of the above-mentioned tea saponin thiazole derivative in the control of root-knot nematodes in vegetables.

[0007] The objective of this invention is achieved through the following technical solution:

[0008] This invention provides a tea saponin thiazole derivative having the molecular structure shown in formula (I):

[0009] ;

[0010] Formula (I);

[0011] Wherein, R is F, Cl or trifluoromethyl.

[0012] This invention also provides a method for preparing a tea saponin thiazole derivative, comprising the following steps:

[0013] (1) Dissolve tea saponin and tert-butyldimethylchlorosilane in N,N-dimethylformamide (DMF), and add imidazole and catalyst 4-dimethylaminopyridine (DMAP) to react. After the reaction is completed, distilled water is added to dilute the mixture, and it is extracted with ethyl acetate. The organic layer is washed, dehydrated, and the ethyl acetate is evaporated to obtain intermediate 1.

[0014] (2) Intermediate 1 was dissolved in dichloromethane and reacted with sodium triacetoxyborohydride. After the reaction was completed, distilled water was added for dilution, and the mixture was extracted with ethyl acetate. The organic layer was washed, dehydrated, and the ethyl acetate was evaporated to obtain intermediate 2.

[0015] (3) Intermediate 2 was dissolved in pyridine with thiazol-4-carboxylic acid or a thiazol-4-carboxylic acid derivative, and 4-dimethylaminopyridine (DMAP) and 1-ethyl-(3-dimethylaminopropyl)carbodiimide hydrochloride (EDC.HCl) were added to react. After the reaction was completed, distilled water was added to dilute the mixture, and the mixture was extracted with ethyl acetate. The organic layer was washed, dehydrated, and the ethyl acetate was evaporated to obtain intermediate 3.

[0016] (4) Intermediate 3 was dissolved in anhydrous THF solution and reacted. After the reaction was completed, distilled water was added for dilution, and the mixture was extracted with ethyl acetate. The organic layer was washed, dehydrated, and the ethyl acetate was evaporated to obtain the tea saponin thiazole derivative.

[0017] Preferably, the organic layer is washed and dehydrated, specifically as follows:

[0018] The organic layer was washed with a saturated sodium chloride solution and then dehydrated with anhydrous sodium sulfate.

[0019] Preferably, in step (1):

[0020] The amount of tert-butyldimethylchlorosilane used is 1.5-2 times the mass of tea saponin, and the reaction temperature with tea saponin is 20-30℃, and the reaction time is 10-14h.

[0021] The volume-to-mass ratio of DMF to tea saponin is 5-15:1 mL / g; the amount of imidazole is 1.5-2 times the mass of tea saponin; and the amount of DMAP is 0.02-0.05 times the mass of tea saponin.

[0022] The amount of distilled water and ethyl acetate used is 1-3 times the volume of DMF; the amount of saturated sodium chloride solution used is 1 / 4-1 / 2 times the volume of ethyl acetate; the amount of anhydrous sodium sulfate used is 1 / 2-1 times the mass of tea saponin; the dehydration time is 8-12 hours; and the evaporation temperature is 50-60℃.

[0023] Preferably, in step (2):

[0024] The amount of sodium triacetoxyborohydride used is 0.2-0.4 times the mass of intermediate 1, and the reaction temperature with intermediate 1 is 20-30℃, and the reaction time is 1-3h.

[0025] The volume-to-mass ratio of the dichloromethane solvent to intermediate 1 is 5-15:1 mL / g.

[0026] The amount of distilled water and ethyl acetate used is 1-3 times the volume of dichloromethane; the amount of saturated sodium chloride solution used is 1 / 4-1 / 2 times the volume of ethyl acetate; the amount of anhydrous sodium sulfate used is 1 / 2-1 times the mass of intermediate 1; the dehydration time is 8-12 hours; and the evaporation temperature is 50-60℃.

[0027] Preferably, in step (3):

[0028] The thiazole-4-carboxylic acid derivative is one of 2-(4-chlorophenyl)thiazole-4-carboxylic acid, 2-(4-fluorophenyl)thiazole-4-carboxylic acid, and 2-(4-(trifluoromethyl)phenyl)thiazole-4-carboxylic acid; its amount is 0.2-0.3 times the mass of intermediate 2, the reaction temperature is 30-50℃, and the reaction time is 12-36h.

[0029] The volume-to-mass ratio of pyridine to intermediate 2 is 5-15:1 mL / g; the amount of DMAP used is 0.1-0.2 times the mass of intermediate 2; and the amount of EDC.HCl used is 0.15-0.25 times the mass of intermediate 2.

[0030] The amount of distilled water and ethyl acetate used is 1-3 times the volume of pyridine; the amount of saturated sodium chloride solution used is 1 / 4-1 / 2 times the volume of ethyl acetate; the amount of anhydrous sodium sulfate used is 1 / 2-1 times the mass of intermediate 2; the dehydration time is 8-12 hours; and the evaporation temperature is 50-60℃.

[0031] Preferably, in step (4):

[0032] The volume-to-mass ratio of TBAF to intermediate 3 is 0.1-0.4:1 mL / g, the volume-to-mass ratio of anhydrous THF solvent to intermediate 3 is 3-5:1 mL / g, the reaction temperature is 20-30℃, and the reaction time is 1-3 h.

[0033] The amount of distilled water and ethyl acetate used is 1-3 times the volume of THF; the amount of saturated sodium chloride solution used is 1 / 4-1 / 2 times the volume of ethyl acetate; the amount of anhydrous sodium sulfate used is 1 / 2-1 times the mass of intermediate 3; the dehydration time is 8-12 hours; and the evaporation temperature is 50-60℃.

[0034] Preferably, the tea saponin is a compound obtained by acid and alkali hydrolysis and extraction of tea oil saponin. The preparation method is as follows: tea oil saponin is dissolved in a methanol aqueous solution with a volume fraction of 80-90% (g / mL) at a mass-to-volume ratio of 1:15-25; the pH of the solution is adjusted to 1-3 with 2-4 mol / L hydrochloric acid; the mixture is refluxed at 70-85℃ for 5-6 h; 5-8 times the volume of distilled water is added; and the mixture is allowed to stand for 12-24 h. The precipitate is then dissolved in a methanol aqueous solution with a volume fraction of 80-90% (g / mL) at a mass-to-volume ratio of 1:15-25; the pH of the solution is adjusted to 11-13 with 5-10% sodium hydroxide aqueous solution; the mixture is refluxed at 70-85℃ for 5-6 h; 5-8 times the volume of distilled water is added; and the mixture is allowed to stand for 12-24 h. h; The precipitate was extracted with chloroform at a mass-to-volume ratio of 1:5-10 (g / mL), and the solvent was evaporated to obtain tea saponin.

[0035] The present invention also provides the application of the above-mentioned tea saponin thiazole derivative in the control of root-knot nematodes in vegetables; the tea saponin thiazole derivative is used to prepare a drug preparation for preventing root-knot nematodes; the drug preparation is a single or compound preparation containing the tea saponin thiazole derivative.

[0036] Compared with the prior art, the present invention has the following advantages and beneficial effects:

[0037] (1) The present invention prepared a tea saponin thiazole derivative by structural derivatization. Due to the synergistic effect of multiple targets, the tea saponin thiazole derivative has a more than 10-fold increase in anti-root-knot nematode activity compared with tea saponin.

[0038] (2) The method for preparing the tea saponin thiazole derivative of the present invention involves protecting the hydroxyl group on the tea saponin with tert-butyldimethylchlorosilane to ensure the correct site for subsequent reactions and obtain a product with uniform structure, quality and activity, thus obtaining intermediate 1; then reducing the aldehyde group on intermediate 1 with sodium triacetoxyborohydride to make it a hydroxyl group, thus obtaining intermediate 2; then reacting the thiazole-4-carboxylic acid derivative with the reduced hydroxyl group to obtain intermediate 3; finally removing the tert-butyldimethylchlorosilane on intermediate 3 with tetrabutylammonium fluoride to obtain the tea saponin thiazole derivative.

[0039] (3) The preparation method of the present invention is simple, can be industrialized and prepared, and is easy to promote and apply. Attached Figure Description

[0040] Figure 1 Infrared spectra of tea saponin and the tea saponin derivative of Example 1 of the present invention.

[0041] Figure 2 This is the hydrogen nuclear magnetic resonance spectrum of tea saponin.

[0042] Figure 3 The image shows the hydrogen nuclear magnetic resonance spectrum of the tea saponin derivative obtained in Example 1 of this invention.

[0043] Figure 4 This is a microscopic image of root-knot nematodes before treatment with tea saponin derivatives, as shown in Example 1 of the present invention.

[0044] Figure 5 This is a microscopic image of root-knot nematodes treated with tea saponin derivatives according to Example 1 of the present invention.

[0045] Figure 6 This is an electron micrograph of root-knot nematodes before treatment with tea saponin derivatives in Example 1 of the present invention.

[0046] Figure 7 This is an electron micrograph of root-knot nematodes treated with tea saponin derivatives according to Example 1 of the present invention. Detailed Implementation

[0047] The present invention is further described below through specific embodiments, but the scope of protection of the present invention is not limited thereto.

[0048] In this embodiment, the camellia saponin was produced by Jiangxi Xinzhongye Tea Industry Technology Co., Ltd.; 2-(4-chlorophenyl)thiazole-4-carboxylic acid (CAS: 17228-98-7) and 2-(4-fluorophenyl)thiazole-4-carboxylic acid (CAS: 863668-07-9) were purchased from Beijing Mairuida Technology Co., Ltd.; and 2-(4-(trifluoromethyl)phenyl)thiazole-4-carboxylic acid (CAS: 144061-16-5) was purchased from Shanghai Bide Pharmaceutical Technology Co., Ltd.

[0049] Preparation of tea saponins:

[0050] 100g of tea saponin was dissolved in 2000mL of 85% methanol aqueous solution. The pH of the solution was adjusted to 2 with 3mol / L hydrochloric acid, and the mixture was refluxed at 80℃ for 5.5h. 12L of distilled water was added, and the mixture was allowed to stand for 18h. 30g of the precipitate was dissolved in 600mL of 85% methanol aqueous solution. The pH of the solution was adjusted to 12 with 8% sodium hydroxide aqueous solution, and the mixture was refluxed at 80℃ for 5.5h. 3.6L of distilled water was added, and the mixture was allowed to stand for 18h. 20g of the precipitate was extracted with 200mL of chloroform, and the solvent was evaporated to obtain 18g of tea saponin.

[0051] Example 1

[0052] (1) Dissolve 10 g of tea saponin and 20 g of tert-butyldimethylchlorosilane in 150 mL of DMF, add 20 g of imidazole and 0.5 g of DMAP, react at 30 °C for 10 h, after the reaction is completed, dilute with 150 mL of distilled water, extract with 150 mL of ethyl acetate, wash the obtained organic phase with 75 mL of saturated NaCl aqueous solution, dehydrate with 5 g of anhydrous Na2SO4 for 12 h, and evaporate ethyl acetate at 50 °C to obtain 26 g of intermediate 1.

[0053] (2) Dissolve 10 g of intermediate 1 and 4 g of sodium triacetoxyborohydride in 150 mL of dichloromethane and react at 30 °C for 1 h. After the reaction is complete, dilute with 150 mL of distilled water and extract with 150 mL of ethyl acetate. Wash the organic phase with 75 mL of saturated NaCl aqueous solution and dehydrate with 5 g of anhydrous Na2SO4 for 12 h. Ethyl acetate is evaporated at 50 °C to obtain 12 g of intermediate 2.

[0054] (3) 10 g of intermediate 2 and 3 g of 2-(4-chlorophenyl)thiazole-4-carboxylic acid were dissolved in 150 mL of pyridine, 2 g of DMAP and 2.5 g of EDC.HCl were added, and the reaction was carried out at 50 °C for 12 h. After the reaction was completed, 150 mL of distilled water was added to dilute the mixture, and 150 mL of ethyl acetate was used for extraction. The resulting organic phase was washed with 75 mL of saturated NaCl aqueous solution, dried with 5 g of anhydrous Na2SO4 for 12 h, and ethyl acetate was evaporated at 50 °C to obtain 12.5 g of intermediate 3.

[0055] (4) Dissolve 10 g of intermediate 3 and 4 mL of TBAF in 50 mL of anhydrous THF solution and react at 30 °C for 1 h. After the reaction is complete, dilute with 50 mL of distilled water and extract with 50 mL of ethyl acetate. Wash the organic phase with 25 mL of saturated NaCl and dehydrate with 5 g of anhydrous Na2SO4 for 12 h. Ethyl acetate is evaporated at 50 °C to obtain 7.5 g of tea saponin derivative.

[0056] Example 2

[0057] (1) Dissolve 10 g of tea saponin and 15 g of tert-butyldimethylchlorosilane in 50 mL of DMF, and add 15 g of imidazole and 0.2 g of DMAP. React at 20 °C for 14 h. After the reaction is complete, dilute with 150 mL of distilled water and extract with 150 mL of ethyl acetate. Wash the obtained organic phase with 50 mL of saturated NaCl aqueous solution, dehydrate with 10 g of anhydrous Na2SO4 for 8 h, and evaporate ethyl acetate at 60 °C to obtain 23 g of intermediate 1.

[0058] (2) Dissolve 10 g of intermediate 1 and 2 g of sodium triacetoxyborohydride in 50 mL of dichloromethane and react at 20 °C for 3 h. After the reaction is complete, dilute with 150 mL of distilled water and extract with 150 mL of ethyl acetate. Wash the organic phase with 50 mL of saturated NaCl aqueous solution and dehydrate with 10 g of anhydrous Na2SO4 for 8 h. Ethyl acetate is evaporated at 50 °C to obtain 11 g of intermediate 2.

[0059] (3) Dissolve 10 g of intermediate 2 and 2 g of 2-(4-fluorophenyl)thiazole-4-carboxylic acid in 50 mL of pyridine, add 1 g of DMAP and 1.5 g of EDC.HCl and react at 30 °C for 36 h. After the reaction is complete, dilute with 150 mL of distilled water and extract with 150 mL of ethyl acetate. Wash the obtained organic phase with 50 mL of saturated NaCl aqueous solution and dry with 10 g of anhydrous Na2SO4 for 8 h. Ethyl acetate is evaporated at 50 °C to obtain 11.5 g of intermediate 3.

[0060] (4) Dissolve 10 g of intermediate 3 and 1 mL of TBAF in 30 mL of anhydrous THF solution and react at 20 °C for 3 h. After the reaction is complete, dilute with 90 mL of distilled water and extract with 90 mL of ethyl acetate. Wash the organic phase with 30 mL of saturated NaCl and dehydrate with 10 g of anhydrous Na2SO4 for 8 h. Ethyl acetate is removed at 50 °C to obtain 6 g of tea saponin derivative.

[0061] Example 3

[0062] (1) Dissolve 10 g of tea saponin and 16 g of tert-butyldimethylchlorosilane in 100 mL of DMF, and add 16 g of imidazole and 0.4 g of DMAP. React at 25 °C for 12 h. After the reaction is complete, dilute with 200 mL of distilled water and extract with 200 mL of ethyl acetate. Wash the obtained organic phase with 70 mL of saturated NaCl aqueous solution and dehydrate with 8 g of anhydrous Na2SO4 for 10 h. Ethyl acetate is evaporated at 60 °C to obtain 25 g of intermediate 1.

[0063] (2) Dissolve 10 g of intermediate 1 and 3 g of sodium triacetoxyborohydride in 120 mL of dichloromethane and react at 25 °C for 2 h. After the reaction is complete, dilute with 240 mL of distilled water and extract with 240 mL of ethyl acetate. Wash the organic phase with 60 mL of saturated NaCl aqueous solution and dehydrate with 8 g of anhydrous Na2SO4 for 10 h. Ethyl acetate is evaporated at 55 °C to obtain 12 g of intermediate 2.

[0064] (3) 10 g of intermediate 2 and 2.5 g of 2-(4-(trifluoromethyl)phenyl)thiazole-4-carboxylic acid were dissolved in 120 mL of pyridine, 1.5 g of DMAP and 2 g of EDC.HCl were added, and the reaction was carried out at 40 °C for 20 h. After the reaction was completed, 240 mL of distilled water was added for dilution, and 240 mL of ethyl acetate was used for extraction. The organic phase was washed with 60 mL of saturated NaCl aqueous solution, dried with 8 g of anhydrous Na2SO4 for 10 h, and ethyl acetate was evaporated at 55 °C to obtain 12 g of intermediate 3.

[0065] (4) Dissolve 10 g of intermediate 3 and 3 mL of TBAF in 40 mL of anhydrous THF solution and react at 25 °C for 2 h. After the reaction is complete, dilute with 80 mL of distilled water and extract with 80 mL of ethyl acetate. Wash the organic phase with 20 mL of saturated NaCl and dehydrate with 8 g of anhydrous Na2SO4 for 10 h. Ethyl acetate is evaporated at 55 °C to obtain 7 g of tea saponin derivative.

[0066] Comparative Example 1

[0067] The difference between this comparative example and Example 1 is that the preparation in step (3) is omitted, and intermediate 3 in step (4) is replaced with intermediate 2, while other conditions remain unchanged, resulting in 6.3 g of product.

[0068] Comparative Example 2

[0069] The difference between this comparative example and Example 1 is that the reaction time in step (3) was changed to 10 h, while other conditions remained unchanged. The result was 5.8 g of intermediate 3. This indicates that insufficient reaction time significantly reduced the yield, and that the reaction time of this invention is appropriate.

[0070] Comparative Example 3

[0071] The difference between this comparative example and Example 1 is that the reaction temperature in step (3) was changed to 20°C, while other conditions remained unchanged. The result was 5.7g of intermediate 3. This indicates that a reaction temperature that is too low is detrimental to the reaction, significantly reducing the yield. The reaction temperature of this invention is appropriate.

[0072] Comparative Example 4

[0073] The difference between this comparative example and Example 1 is that 2-(4-chlorophenyl)thiazole-4-carboxylic acid in step (3) is replaced with thiazole-2-carboxylic acid (CAS: 14190-59-1, purchased from Guangzhou Hewei Pharmaceutical Technology Co., Ltd.), while other conditions remain unchanged, and the resulting product is 6.1g.

[0074] Performance testing:

[0075] Test 1

[0076] Structural characterization of tea saponin thiazole derivatives

[0077] The infrared and proton spectra of tea saponin and the tea saponin thiazole derivative of Example 1 were analyzed using a Fourier transform infrared spectrometer (Thermo Scientific Nicolet iS10) and a nuclear magnetic resonance spectrometer (Burker Avance III).

[0078] The results are as follows Figure 1 , Figure 2 , Figure 3 As shown. Compare the infrared spectra of tea saponins ( Figure 1 ), tea saponin thiazole derivative at 1256.01 cm -1 An asymmetric stretching vibration peak of the ester bond appeared at 838.82 cm⁻¹. -1 There is an out-of-plane bending vibration of the aromatic ring CH bond at 776.43 cm⁻¹. -1 Vibrational peaks of the aromatic ring and thiazole ring skeleton are present at 668.32 cm⁻¹. -1 The presence of C-Cl stretching vibration indicates that the tea saponin has successfully reacted, yielding the corresponding tea saponin derivative.

[0079] The hydrogen spectral data of tea saponins are as follows: 1 H NMR ((400 MHz, DMSO) δ 9.25 (s, 1H), 6.88(s, 1H), 5.49 (t, J = 26.4 Hz, 1H), 5.17 (s, 2H), 4.66 (s, 1H), 4.28 (s, 1H),3.66-3.97 (m, 1H), 3.58 (d, J = 7.9 Hz, 2H), 3.28-3.24 (m, 3H), 1.99 (t, J =21.2 Hz, 1H), 1.29-1.96 (m, 14H), 1.27 (t, J = 21.3 Hz, 1H), 1.23 (s, 3H),1.02 (t, J = 21.7Hz, 1H), 0.84–0.89 (m, 15H). The 1H NMR data for tea saponin thiazole derivatives are as follows: 1H NMR (500 MHz, DMSO): δ 9.27 (s, 1H), 8.08 (t, J = 7.9 Hz, 2H), 7.57 (t,J = 9.8 Hz, 2H), 6.86 (s, 1H), 5.29 (s, 2H), 5.19 (t, J = 8.9 Hz, 1H), 4.66(s, 1H), 4.22 (s, 1H), 3.95 (s, 2H), 3.80 (d, J = 8.9 Hz, 2H), 3.60 – 3.57(m, 1H), 3.18 – 3.15 (m, 3H), 2.36 (t, J = 10.7 Hz, 1H), 1.81 – 1.16 (m, 19H), 0.95 – 0.77 (m, 15H). Comparison of the 1H NMR spectra of the two studies shows that the aldehyde characteristic peak at δ 9.25 of the tea saponin thiazole derivative disappears, while characteristic peaks of the substituted benzene ring appear at δ 8.08–δ 7.57. Characteristic peaks of hydrogen on the thiazole ring appear at δ 9.27, and an ester group peak appears at δ 3.95. This indicates that 2-(4-chlorophenyl)thiazole-4-carboxylic acid successfully reacted and linked with tea saponin.

[0080] Elemental analysis was performed on tea saponin and the tea saponin thiazole derivative of Example 1, and the results are shown in Table 1. Based on the mass fraction of each element, the atomic ratio in tea saponin was calculated to be C:H:O=30:48:6, and the atomic ratio in the tea saponin thiazole derivative of Example 1 was C:H:O:N:Cl:S=40:54:7:1:1:1, both of which are consistent with the theoretical values.

[0081] Table 1. Elemental content of tea saponins and tea saponin thiazole derivatives

[0082]

[0083] Test 2

[0084] Anti-root-knot nematode activity of the tea saponin thiazole derivatives and comparative products prepared in Examples 1-3.

[0085] The tea saponin thiazole derivatives and comparative products prepared in Examples 1-3 were dissolved in 5% DMSO to prepare a solution of 20.56 mg / mL. This solution was then diluted sequentially to 11 gradients: 1:1, 1:2, 1:4, 1:8, 1:16, 1:32, 1:64, 1:128, 1:256, 1:512, and 1:1024. 150 μL of each solution was mixed with 100 μL of a suspension of 2000 second-instar root-knot nematode larvae. A solvent without the drug solution was used as a negative control, and avermectin as a positive control. The mixture was incubated at 27°C for 72 h, and the drug concentration at which 90% mortality of the root-knot nematodes was determined. Microscopic observation indicated death by stiffness and immobility, while worms exhibiting bending or wriggling movements were considered alive.

[0086] Results: The tea saponin thiazole derivative exhibited good killing effect on second-instar larvae of root-knot nematodes. Compared with tea saponin, the tea saponin thiazole derivative of this invention significantly reduced the killing concentration of root-knot nematodes and increased the activity by more than 10 times. The insecticidal effects of the products of Comparative Examples 1, 2, and 3 were all weak, indicating that the synthesis reaction in step (3) of this invention is indispensable. In Comparative Examples 2 and 3, due to the adjustment of reaction temperature and reaction time, there were more unreacted substances in the products, resulting in a decrease in their activity, indicating that the reaction conditions of this invention have been optimized. The weak insecticidal effect of the product of Comparative Example 4 indicates that if the derivative group is replaced with a similar structural group, the different carboxyl group positions of the derivative will produce steric hindrance, affecting the interaction with the nematode target and failing to achieve the effect of this invention.

[0087] Live nematodes and dead nematodes after drug treatment were observed under microscopes and scanning electron microscopes, respectively. The results showed that the live root-knot nematodes that were not treated with tea saponin thiazole derivatives had intact structures. Figure 4 , Figure 6 After treatment with tea saponin thiazole derivatives, root-knot nematodes shrank and ruptured. Figure 5 , Figure 7 This indicates that the tea saponin thiazole derivative of the present invention has a good anti-root-knot nematode effect. The nematodes treated with the comparative derivative showed no significant changes.

[0088] Table 2. Killing effect of tea saponin thiazole derivatives on root-knot nematodes.

[0089]

[0090] Test 3

[0091] Inhibitory effect of tea saponin thiazole derivatives on superoxide dismutase in root-knot nematodes.

[0092] The tea saponin thiazole derivatives and comparative products prepared in Examples 1-3 were dissolved in 5% DMSO to prepare a solution of 0.62 mg / ml, which was then diluted sequentially to three ratios: 1:1, 1:2, and 1:4. 150 μL of this solution was mixed with 100 μL of a suspension of 2000 second-instar root-knot nematode larvae. A solvent without the drug solution was used as a negative control. The mixture was incubated at 27°C for 72 h. After 72 h, the nematodes were washed three times with sterile water, then centrifuged at 5000 rpm for 5 min to remove the supernatant. 200 μL of lysis buffer was added, and the mixture was lysed at 0°C for 40 min. The lysate was then sonicated at 40% power for 3 min. The homogenate was collected, and superoxide dismutase (SOD) activity was tested and calculated according to the kit. The inhibition rate of SOD activity by each drug group was calculated with the negative control enzyme activity at 100%.

[0093] The results are shown in Table 3. The tea saponin thiazole derivative exhibits significant inhibitory activity against SOD in second-instar larvae of root-knot nematodes. Compared to tea saponin, the tea saponin thiazole derivative of this invention significantly improves the inhibition rate of SOD in root-knot nematodes. The products of Comparative Examples 1, 2, and 3 all showed weak inhibitory activity against enzyme activity, indicating that step (3) of the synthesis reaction in this invention is indispensable and the reaction conditions have been optimized. The weak inhibitory activity against enzyme activity in Comparative Example 4 indicates that replacing the derivatized group with a similar structural group would not achieve the same effect as this invention. The weak inhibitory activity of avermectin against SOD indicates that the tea saponin thiazole derivative of this invention has a different mechanism of action than avermectin.

[0094] Table 3. Inhibition rate (%) of tea saponin thiazole derivatives on SOD enzyme activity in root-knot nematodes

[0095]

[0096] Example 4

[0097] Take 30g of the tea saponin thiazole derivative prepared in Example 1, and mix it with 5g of dispersant sodium lignin sulfonate, 2g of disintegrant anhydrous sodium sulfate, 1g of wetting agent sodium dodecyl sulfate, and 62g of excipient kaolin. Add water and knead in a mixer, then granulate by extrusion, dry, and pass through a 40-60 mesh sieve to obtain the water-dispersible granules of the tea saponin thiazole derivative.

[0098] Application method: Dilute this formulation with water 500-800 times and apply it to the root zone soil of the crop by root drenching or furrow application. For transplanted crops (such as tomatoes and cucumbers), apply 0.5-1 liter of solution per plant by root drenching at the time of transplanting or at the early stage of disease occurrence, ensuring that the solution fully penetrates into the main root layer. Depending on the severity of the infestation, the application can be repeated every 10-15 days. This method is effective in controlling root-knot nematodes in vegetables.

[0099] Example 5

[0100] Take 10 g of the tea saponin thiazole derivative prepared in Example 2, and pulverize it separately with 5 g of the synergistic dispersant polyvinylpyrrolidone K30 and 85 g of the carrier anhydrous glucose through a 100-mesh sieve. Mix the above materials in a three-dimensional mixer for 30 min until homogeneous to obtain a soluble powder of the tea saponin thiazole derivative.

[0101] Application method:

[0102] Application: Before sowing or transplanting, mix 1-2 kg of this formulation with an appropriate amount of dry, fine soil and spread it evenly on the soil surface. Then, till it into the soil. This can effectively control root-knot nematodes in vegetables.

[0103] Application in planting holes: When transplanting, apply 0.5-1.0 grams of this formulation directly to each planting hole, mix it slightly with the soil, and then plant the seedling. Water thoroughly immediately after application to promote diffusion of the pesticide in the root zone, which can effectively control root-knot nematodes in vegetables.

[0104] Those skilled in the art will readily understand that the above description is merely an embodiment of the present invention and is not intended to limit the present invention. Any modifications, equivalent substitutions, and improvements made within the spirit and principles of the present invention should be included within the scope of protection of the present invention.

Claims

1. A process for the preparation of a tea saponin aglycone thiazole derivative, characterized by, Includes the following steps: (1) Dissolve tea saponin and tert-butyldimethylchlorosilane in N,N-dimethylformamide (DMF), and add imidazole and catalyst 4-dimethylaminopyridine (DMAP) to react. After the reaction is completed, distilled water is added to dilute the product, and the product is extracted with ethyl acetate. The organic layer is washed, dehydrated, and the ethyl acetate is evaporated to obtain the first intermediate. (2) The first intermediate was reacted with sodium triacetoxyborohydride in dichloromethane. After the reaction was completed, distilled water was added for dilution, and the mixture was extracted with ethyl acetate. The organic layer was washed, dehydrated, and the ethyl acetate was evaporated to obtain the second intermediate. (3) The second intermediate is dissolved in pyridine with thiazole-4-carboxylic acid or a thiazole-4-carboxylic acid derivative, and 4-dimethylaminopyridine (DMAP) and 1-ethyl-(3-dimethylaminopropyl)carbodiimide hydrochloride (EDC.HCl) are added to react. After the reaction is completed, distilled water is added to dilute the mixture, and the mixture is extracted with ethyl acetate. The organic layer is washed, dehydrated, and the ethyl acetate is evaporated to obtain the third intermediate. The reaction temperature is 30~50℃ and the reaction time is 12~36h. (4) The third intermediate was dissolved in anhydrous THF solution and reacted with tetrabutylammonium fluoride (TBAF). After the reaction was completed, distilled water was added for dilution, and the mixture was extracted with ethyl acetate. The organic layer was washed, dehydrated, and the ethyl acetate was evaporated to obtain the tea saponin thiazole derivative. The tea saponin thiazole derivative has the molecular structure shown in formula (I): ； Formula (I) Wherein, R is F, Cl or trifluoromethyl.

2. The process for preparing tea saponin aglycone thiazole derivatives as claimed in claim 1, wherein, In step (1), the amount of tert-butyldimethylchlorosilane used is 1.5-2 times the mass of tea saponin, and the reaction temperature with tea saponin is 20-30℃, and the reaction time is 10-14h. The amount of imidazole used is 1.5-2 times the mass of tea saponin; the amount of 4-dimethylaminopyridine used is 0.02-0.05 times the mass of tea saponin.

3. The method for preparing the tea saponin thiazole derivative according to claim 1, characterized in that, In step (2), the amount of sodium triacetoxyborohydride is 0.2-0.4 times the mass of the first intermediate, and the reaction temperature between it and the first intermediate is 20-30°C, and the reaction time is 1-3h.

4. The method for preparing the tea saponin thiazole derivative according to claim 1, characterized in that, In step (3), the thiazole-4-carboxylic acid derivative is one of 2-(4-chlorophenyl)thiazole-4-carboxylic acid, 2-(4-fluorophenyl)thiazole-4-carboxylic acid, and 2-(4-(trifluoromethyl)phenyl)thiazole-4-carboxylic acid, and the amount of the thiazole-4-carboxylic acid derivative is 0.2-0.3 times the mass of the second intermediate.

5. The method for preparing the tea saponin thiazole derivative according to claim 1, characterized in that, In step (3), the amount of 4-dimethylaminopyridine (DMAP) is 0.1-0.2 times the mass of the second intermediate; the amount of 1-ethyl-(3-dimethylaminopropyl)carbodiimide hydrochloride (EDC.HCl) is 0.15-0.25 times the mass of the second intermediate.

6. The method for preparing the tea saponin thiazole derivative according to claim 1, characterized in that, In step (4), the volume-to-mass ratio of the tetrabutylammonium fluoride (TBAF) to the third intermediate is 0.1-0.4:1 mL / g; the reaction temperature is 20-30℃ and the reaction time is 1-3 h.

7. The method for preparing the tea saponin thiazole derivative according to claim 1, characterized in that, The tea saponin is a compound obtained by acid and alkali hydrolysis and extraction of tea oil saponin.

8. A tea saponin thiazole derivative, characterized in that, It has a molecular structure as shown in formula (I): ； Formula (I) Wherein, R is F, Cl or trifluoromethyl.

9. The application of the tea saponin thiazole derivative according to claim 8 in the control of root-knot nematodes in vegetables.

10. The application according to claim 8, wherein the tea saponin thiazole derivative is used to prepare a pharmaceutical preparation for treating root-knot nematodes.

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