Application of cyclic terpenoid natural products and derivatives thereof as agricultural bactericides in plant diseases
By using cyclic terpenoid natural products and their derivatives to target and inhibit bacterial biofilm formation, the problems of drug resistance and environmental safety of traditional fungicides have been solved, achieving highly efficient control of plant diseases.
Patent Information
- Application Number
- CN202510981196.5
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- Filing Date
- 2025-07-16
- Publication Date
- 2025-11-28
AI Technical Summary
Existing traditional fungicides have problems with drug resistance in the control of plant diseases and pose a threat to the environment and agricultural product safety. There is a need to develop agricultural fungicides with novel structures, unique mechanisms of action, high efficiency and low toxicity.
Cyclic terpenoid natural products and their derivatives are used as agricultural fungicides to target and inhibit bacterial biofilm formation, thereby preventing and controlling bacterial, fungal, and viral diseases in plants.
Cyclic terpenoid natural products and their derivatives exhibit good inhibitory effects on plant pathogenic bacteria, fungi and viruses, are green and safe, and can effectively prevent and control a variety of plant diseases and reduce the risk of drug resistance.
Smart Images
Figure BDA0005502982030000021 
Figure BDA0005502982030000031 
Figure BDA0005502982030000061
Abstract
Description
TECHNICAL FIELD
[0001] The present application relates to the field of plant disease control, in particular to the application of cyclic terpenoid natural products and derivatives thereof as agricultural fungicides in plant bacterial diseases. BACKGROUND
[0002] Plant diseases, as a major threat in the field of agricultural production and natural ecology, are of great importance and far-reaching harm. Plant fungi can cause various plant diseases, leading to plant growth and development inhibition, tissue necrosis, physiological disorder, and even whole plant death. They can cause crop yield reduction and quality decline by directly invading plant tissues, destroying cell structure and function, and interfering with key physiological processes such as photosynthesis, seriously affecting the economic benefits of agricultural production and food security. At the same time, the prevalence of fungal diseases can also disrupt the ecological balance and exacerbate the loss of biodiversity. The harm is throughout the whole growth period of plants, with diverse transmission routes and great difficulty in prevention and control, which is a major challenge in agricultural production and ecological system. Plant pathogenic bacteria have become the second largest plant pathogen after fungi, and almost every plant, whether it is a vegetable, fruit tree or other crop, can be affected by various bacterial diseases. Common bacterial diseases on rice include rice bacterial leaf blight and bacterial leaf streak, which can cause an area of 330,000-660,000 hm 2 2 per year, usually causing a reduction of about 10%; there are also many types of bacterial diseases on vegetables, such as cucumber bacterial leaf spot, tomato bacterial wilt, cruciferous vegetable soft rot and black rot, etc. Bacterial diseases are also an important reason for the reduction of melon and fruit crops; citrus canker can affect the appearance of fruit trees and fruits, and can cause massive defoliation, fruit drop, and even branch dieback, seriously affecting the yield and quality of citrus; after kiwi fruit infection, local ulceration and rotting can cause tree weakness, and the disease rate can be as high as 20%. In recent years, plant bacterial diseases have been increasing year by year, and there have been reports of new plant bacterial diseases, such as wheat black necrosis, corn bacterial stem rot, cucumber bacterial flow disease, and bacterial basal rot, which pose an increasingly serious threat to agricultural production.
[0003] The outbreak, prevalence, destructiveness and prevention of plant diseases make chemical control a necessary measure. Under the background of national agricultural green development, with the rapid development of agriculture and the increasing problems of pests and diseases, the prevention and control of plant diseases should comply with the development of the times, take into account both disease control and chemical pesticide reduction, and vigorously promote the green prevention and control technology guided by the principle of "prevention first and comprehensive prevention and control". Agricultural fungicides have become the most important means of controlling plant diseases due to their advantages of high efficiency and rapid effect, and are the main measures to ensure high and stable yield of crops in China. Agricultural fungicides can effectively control plant diseases and ensure the healthy growth of crops, thereby improving the yield and quality of crops and playing a crucial role in agricultural production. Although traditional fungicides can inhibit diseases to some extent, they have many shortcomings. Due to the frequent use of traditional fungicides and the single mode of antibacterial action, the problem of drug resistance of plant pathogens is becoming increasingly serious. In addition, some traditional fungicides have large residual amounts, which pose potential threats to the environment and agricultural product safety, and affect the export of agricultural products and the health of consumers. In summary, the drug resistance of plant pathogens has become a key scientific problem restricting the research and development of pesticides, so it is urgent to develop novel antibacterial drugs with novel structure, unique mechanism of action, high efficiency and low toxicity to replace traditional fungicides, and to strengthen the integration and innovation of disease control technology to improve the safety and sustainability of agricultural production.
[0004] Bacteria produce biofilms during growth and reproduction. Various major biological macromolecules such as proteins, polysaccharides, DNA, RNA, peptidoglycan, lipids and phospholipids exist in the biofilm. In addition, the formation of biofilm is a dynamic process, including the initial adhesion of bacteria, the development and maturation of biofilm, and the spread of biofilm. Targeting biofilm and other virulence factors can effectively avoid the problem of drug resistance caused by drug pressure. Therefore, by inhibiting the formation of bacterial biofilm, the process of bacterial infection is inhibited to achieve the effect of disease resistance, which has become an important direction for the development of new pesticides. Therefore, it is expected to accelerate the development of new pesticides by mining new biofilm inhibitors from natural compounds. SUMMARY
[0005] The purpose of the present application is to provide the application of cyclic terpenoid natural products and their derivatives as agricultural fungicides in plant diseases, in order to solve the problems existing in the prior art.
[0006] To achieve the above purpose, the present application provides the following solutions:
[0007] The present application provides the application of cyclic terpenoid natural products and their derivatives as agricultural fungicides in plant diseases.
[0008] Cyclic terpenoid natural products and their derivatives include: jasmone, cocaraldehyde, cuminaldehyde, β-ionone, perillaldehyde, citric acid, β-cyclocitral, carvone, perillaldehyde, juniperol, noprol, crocinaldehyde, damastone, naringenone, α-pinene, eugenol, sabinene, terpinene, γ-terpinene, p-cymene, and phellandrene, with the following structural formulas:
[0009]
[0010]
[0011] Among a series of cyclic terpenoid natural products and their derivatives screened, nobutol and citric acid aldehyde were preferred.
[0012] Preferably, the plant diseases include bacterial plant diseases, fungal plant diseases, and viral plant diseases. The bacterial plant diseases include pathogens of rice bacterial blight, *Xanthomonas oryzae* (rice leaf spot pathogen), *Xanthomonas carpetii*, *Xanthomonas brasiliensis* (rapeseed pathogen), *Pseudomonas syringae* (tomato leaf spot pathogen), *Pseudomonas syringae* (kiwifruit pathogen), *Ralstonia solanacearum*, and *Pseudomonas syringae* (cucumber leaf spot pathogen). Diseases caused by leaf spot pathogens, *Ralstonia solanacearum*, *Xanthomonas aeruginosa*, soybean pathogens, or *Corynebacterium micranthum*, the pathogen causing tomato canker; fungal diseases include cucumber gray mold, pepper wilt, rapeseed sclerotinia rot, wheat scab, potato late blight, blueberry root rot, grape coccidioidomyces, dragon fruit anthracnose, rice sheath blight, diseases caused by Fusarium SP or *Colletotrichum sorghum*; viral diseases include those caused by tobacco mosaic virus.
[0013] This invention also provides a method for preventing and controlling agricultural diseases, which involves spraying agricultural fungicides onto crops for disease control. The diseases include rice bacterial blight, rice bacterial leaf streak, citrus canker, cabbage black rot, tomato bacterial spot, kiwifruit canker, tobacco bacterial wilt, cucumber bacterial angular leaf spot, potato bacterial wilt, soybean bacterial spot or tomato canker, cucumber gray mold, pepper wilt, rapeseed sclerotinia rot, wheat scab, potato late blight, blueberry root rot, grape coccidioidomyces, dragon fruit anthracnose, rice sheath blight, diseases caused by Fusarium SP or sorghum spore-forming fungi, and tobacco mosaic virus.
[0014] The present invention discloses the following technical effects:
[0015] The application firstly studies the cyclo terpenes natural products and derivatives thereof as a fungicide to prevent and treat plant diseases. Compared with traditional fungicides, the cyclo terpenes natural products and derivatives thereof have the advantages of green, safety, and soil microbial degradation. The application verifies the in vitro antibacterial activity of the cyclo terpenes natural products and derivatives thereof on rice bacterial leaf blight through experiments. At a concentration of 100 μg / mL, the in vitro antibacterial activity of nopol on plant pathogenic bacteria Xoo, Xac and Psa is 97.18%, 74.81% and 25.24% respectively, and the in vitro antibacterial activity of citraflavonol on plant pathogenic bacteria Xoo, Xac and Psa is 98.99%, 86.01% and 62.74% respectively; at a concentration of 50 μg / mL, the in vitro antibacterial activity of nopol on plant pathogenic bacteria Xoo, Xac and Psa is 97.18%, 56.85% and 14.63% respectively, and at a concentration of 50 μg / mL, the in vitro antibacterial activity of citraflavonol on plant pathogenic bacteria Xoo, Xac and Psa is 98.54%, 79.65% and 30.63% respectively.
[0016] The experimental results prove that the cyclo terpenes natural products and derivatives thereof have good inhibitory effect on pathogenic bacteria, and preliminary exploration shows that the cyclo terpenes natural products and derivatives thereof can potentially affect the biofilm formation of plant pathogenic bacteria, wherein nopol, citraflavonol, round yuzuketone and γ-terpinene show excellent biofilm inhibition activity, and such natural products have extremely high research value in preventing and treating plant bacterial diseases. DETAILED DESCRIPTION
[0017] The various exemplary embodiments of the application will now be described in detail, which should not be considered as limiting the application, but should be understood as a more detailed description of certain aspects, characteristics and embodiments of the application.
[0018] It should be understood that the terms described in the application are only for describing the specific embodiments, and are not used to limit the application. In addition, for the numerical range in the application, it should be understood that each intermediate value between the upper limit and the lower limit of the range is also specifically disclosed. Each smaller range between the intermediate value in any stated value or stated range and any other stated value or intermediate value in the range is also included in the application. The upper limit and the lower limit of these smaller ranges can be independently included or excluded from the range.
[0019] Unless otherwise indicated, all technical and scientific terms used herein have the same meaning as those of a person of ordinary skill in the art to which the application pertains. Although methods and materials similar or equivalent to those described herein can be used in the practice or testing of the present application, preferred methods and materials are described. All documents mentioned herein are incorporated by reference to disclose and describe in full the methods and / or materials which are described therein. In case of conflict between the content of the specification and that of any document incorporated herein by reference, the content of the specification prevails.
[0020] Many modifications and variations of the present application described in the specification are possible without departing from the scope or spirit of the application. Other implementations of the application will be apparent to those skilled in the art from consideration of the specification and practice of the application. The specification and examples are illustrative only.
[0021] As used herein, the terms "comprises", "comprising", "includes", "including", "has", "having" and the like are open-ended terms that are intended to permit but not limit the inclusion of elements or the number of elements, as well as the possibility that one or more elements can be incorporated into the composition, method, or the like.
[0022] In the present application, crocetin (95%), cypressene (98%), γ-terpinene (95%), p-cymene (95%), α-pinene (98%), cuminic aldehyde (98%), citral (97%), β-ionone (97%), perilla alcohol (90%), nopol (97%) were purchased from Anhui Zesheng Science and Technology Co., Ltd. β-cyclocitral (95%), perillaldehyde (90%), carvone (98%), damascenone (97%), terpinolene (95%), phellandrene (85%), sabinene (75%), jasminone (98%), corchoone (94%), eugenol (98%) were purchased from Shanghai Bide Pharmaceutical Technology Co., Ltd. Xoo was provided by Professor Zhou Mingguo's research group of Nanjing Agricultural University; Xac was preserved by Professor Yang Song's research group; Psa was provided by Professor Zou Lifang's research group of Shanghai Jiaotong University.
[0023] Example 1
[0024] Biological activity test of cyclic terpenoid natural products and their derivatives on plant pathogenic bacteria.
[0025] The test objects were Xoo, Xac, and Psa. Zinc thiazole (30% wettable suspension) was used as the test control, and DMSO dissolved in the culture medium was used as the blank control.
[0026] 1.1 Xoo was inoculated into NB medium, respectively, and cultured in a constant temperature shaker at 28°C and 220 rpm to the logarithmic growth phase for standby.
[0027] 1.2 The selected cyclic terpenoid natural products and their derivatives, and thiazole zinc (30% wettable suspension) were respectively formulated into NB liquid medium containing different concentrations of compounds at 100 μg / mL, 50 μg / mL, and 5 mL of each was added to a test tube.
[0028] 1.3 To the test tube containing different concentrations of the medicament in step 1.2, 40 μL of NB liquid medium containing plant pathogenic bacteria was added, and the culture was incubated at 28°C, 220 rpm on a constant temperature shaker for 36 h for Xoo, 48 h for Psa, and 28 h for Xac.
[0029] 1.4 The OD value of each concentration of the bacterial solution was measured on a spectrophotometer, and the OD value of the corresponding concentration of sterile NB liquid medium containing the toxin was also measured. 595 595
[0030] Corrected OD value = OD value of the bacterial culture - OD value of the sterile culture
[0031] Inhibition rate % = [(corrected OD value of the control culture - corrected OD value of the culture containing the toxin) / corrected OD value of the control culture] x 100
[0032] The experimental results are shown in Table 1.
[0033] Table 1 In vitro antibacterial activity of the compounds against plant pathogenic bacteria Xoo, Psa, and Xac at concentrations of 100 μg / mL and 50 μg / mL
[0034]
[0035]
[0036] As can be seen from Table 1, the cyclic terpenoid natural products and their derivatives showed good in vitro antibacterial activity against plant pathogenic bacteria. Some of the compounds, such as nopol and citral, showed in vitro antibacterial activity against plant pathogenic bacteria Xoo, Xac, and Psa at a concentration of 100 μg / mL as follows: nopol, 97.18%, 74.81%, and 25.24%; and citral, 98.99%, 86.01%, and 62.74%, respectively. At a concentration of 50 μg / mL, nopol showed in vitro antibacterial activity against plant pathogenic bacteria Xoo, Xac, and Psa as follows: 97.18%, 56.85%, and 14.63%, respectively, and citral showed in vitro antibacterial activity against plant pathogenic bacteria Xoo, Xac, and Psa as follows: 98.54%, 79.65%, and 30.63%, respectively.
[0037] Example 2
[0038] Test of the biofilm inhibition activity of cyclic terpenes natural products and their derivatives on plant pathogenic bacteria.
[0039] Biofilm, also known as biofilm, refers to the secreted aggregates formed by bacteria and their adhesion to the surface of living or non-living objects, which plays a very important role in plant pathogenic bacteria resistance. Biofilm formation is studied by crystal violet staining method.
[0040] Resuspend Xoo culture to OD 595 = 0.6-0.8, dilute the resuspended Xoo bacterial solution with sterilized NB medium to adjust the OD value to 0.1-0.2, add 150 μL of bacterial suspension to a sterile 96-well plate. Add DMSO or compounds to the bacterial suspension, respectively, so that the concentration of farnesol in the bacterial suspension is 100 μg / mL and 50 μg / mL, respectively, and each treatment is repeated 3 times, and place it in a 28℃ incubator for 72h.
[0041] After incubation, gently remove the bacterial suspension with a 200 μL pipette, wash off the excess suspended cells with sterile water, and then air dry at room temperature for 4h. Add 0.1% crystal violet solution (w / v) 200 μL to each well and stain for 15min, then remove the crystal violet solution, wash off the excess crystal violet solution with sterile water, then dry in a 60℃ oven for 1h, then add 200 μL of 95% ethanol solution to completely dissolve the crystal violet, and finally determine the biofilm content by measuring the OD 570 .
[0042] Corrected OD value = OD value of bacterial culture medium - OD value of sterile culture medium
[0043] Inhibition rate % = [(corrected control ethanol dissolution solution OD value - corrected drug-containing ethanol dissolution solution OD value) / corrected control ethanol dissolution solution OD value] x 100
[0044] The experimental results are shown in Table 2.
[0045] Table 2. Biofilm inhibition activity of compounds on plant pathogenic bacteria Xoo at concentrations of 100 μg / mL and 50 μg / mL
[0046]
[0047] As can be seen from Table 2, cyclic terpenes natural products and their derivatives have good inhibitory effect on pathogenic bacteria, and preliminary exploration shows that the cyclic terpenes natural products and their derivatives can potentially affect the biofilm formation of plant pathogenic bacteria, among which nopol, citral, round yuzuketone and γ-terpinene show excellent biofilm inhibition activity. Such natural products have very high research value in the prevention and control of plant bacterial diseases.
[0048] Example 3
[0049] Biological activity test of cyclic terpenes natural products and their derivatives on plant pathogenic fungi.
[0050] The mycelial growth rate method, also known as the toxic medium method, is one of the conventional methods for determining the virulence of fungicides. The main principle is to mix the test agent with the culture medium, and the virulence of the agent is measured by the growth rate of the colonies on the toxic culture medium. In this example, wheat scab fungus, rice sheath blight fungus, and pepper fusarium wilt fungus were used as test objects, and DMSO was used as a blank control. The specific operation is as follows: 1) according to the test concentration, weigh an appropriate amount of drug, dissolve it with DMSO (the amount should not exceed 1% of the final toxic culture medium), then add 0.1% Tween 20 solution to make up to 10 mL, pour into 90 mL of melted PDA culture medium, mix well and pour into 9 Petri dishes for standby; 2) sterilize the puncher (inner diameter 5 mm) by burning, then cool it down, punch the mycelium at the edge of the pre-activated strain, and use the inoculation needle to paste the mycelium surface to the center of the toxic culture medium, then place them in a 25°C incubator after treatment; 3) when the colony diameter of the control group grows to 5.5-6.6 cm, measure the colony diameter of the control group and each drug treatment group using the cross method; 4) calculate the inhibition rate (%) using the following formula: inhibition rate (%) = (C-T) / (C-0.5) x 100. Where C is the colony diameter of the control group, T is the colony diameter of the drug treatment group, and 0.5 cm is the diameter of the inoculated fungus cake.
[0051] Table 3 Inhibitory activity of cyclic terpenes natural products and their derivatives on plant pathogenic fungi (25 μg / mL)
[0052]
[0053]
[0054] As can be seen from Table 3, cyclic terpenes natural products and their derivatives showed certain in vitro inhibitory activity on plant pathogenic fungi (such as wheat scab fungus, rice sheath blight fungus, and pepper fusarium wilt fungus). At a concentration of 25 μg / mL, the inhibition rates of nopol on wheat scab fungus, rice sheath blight fungus, and pepper fusarium wilt fungus were 15.45%, 20.12%, and 18.37%, respectively; the inhibition rates of citraflavon on rice sheath blight fungus and pepper fusarium wilt fungus were 16.45% and 17.41%, respectively.
[0055] Example 4
[0056] Biological activity test of cyclic terpenes natural products and their derivatives on plant pathogenic viruses.
[0057] The activity of the compounds on tobacco mosaic virus (TMV) in vivo was determined by the half-leaf spot method. 3 mg of the test compound was accurately weighed in a weighing bottle, and 60 μL of solvent DMSO was added to dissolve the compound. The compound solution was prepared to 100 mg / L with double-distilled water containing 1 wt% Tween 20. 250 μL of 2 wt% anisomycin aqueous solution was taken, 60 uL of solvent DMSO was added, and the anisomycin solution was prepared to 100 mg / L with double-distilled water containing 1 wt% Tween 20.
[0058] The in vivo treatment activity of the medicament on TMV infection. The heart leaves of tobacco with consistent growth were selected, and the virus liquid (concentration of 6 x 10 -3 mg / mL) was first taken with a brush, and was inoculated on the leaf blade with embedded corundum by artificial rubbing along the branch vein direction of the leaf surface (whole leaf). The inoculation force of the left and right leaf blades was kept consistent as much as possible, and the wood plate was used to support under the leaf blade. After the virus liquid was dried, the corundum on the leaf blade was washed away with running water. After the leaf blade was dried, the medicament was applied on the left half leaf, and sterile water was applied on the right half leaf as a control. 3 plants were set for each medicament treatment, and 3-4 leaves were set for each plant. Subsequently, the plants were placed in a light incubator for moisture culture, and the temperature was controlled at 23°C, and the light was controlled at 10000 LuX. After 2-4 days, the number of dry spots was observed and recorded. Each medicament was repeated 3 times according to the above method, and the inhibition rate (Y) was calculated according to the following formula:
[0059] Y (%) = (R-L) / R x 100%
[0060] Wherein: Y is the inhibition rate of the compound on tobacco mosaic virus; R is the number of dry spots of the control group (right half leaf); and L is the number of dry spots of the treatment group (left half leaf).
[0061] Table 4 Activity test of cyclic terpene natural products and derivatives thereof on plant pathogenic virus TMV (100 mg / L)
[0062]
[0063]
[0064] As can be seen from Table 4, in the in vivo test, part of the cyclic terpene natural products and derivatives thereof showed certain treatment activity and protective activity on the plant pathogenic virus TMV. Among them, at a concentration of 100 mg / L, the protective activity of nopol, citraflavon and round yuzuketone was 11.0%, 12.8% and 13.78% respectively; the treatment activity of nopol and citraflavon was 17.8% and 14.5% respectively.
[0065] In conclusion, through a series of biological activity tests, the results show that the cyclo terpenes natural product and the derivative thereof have good biological activity on plant pathogenic bacteria, and also have certain biological activity on plant pathogenic viruses and fungi, and have important research value and application prospect.
[0066] The above-described embodiments are merely preferred modes of the present application and are not intended to limit the scope of the present application, and various modifications and improvements to the technical solutions of the present application made by those skilled in the art without departing from the design spirit of the present application shall fall within the protection scope of the present application as defined by the claims.
Claims
1. Application of cyclic terpenoid natural products and their derivatives as agricultural fungicides in plant diseases.
2. The application of cyclic terpenoid natural products and their derivatives as agricultural fungicides in plant diseases according to claim 1, characterized in that: The cyclic terpenoid natural products and their derivatives are: jasmone, cocaraldehyde, cuminaldehyde, β-ionone, perillol, citric acid, β-cyclic citral, carvone, perillaldehyde, juniperol, noprol, crocinaldehyde, damastone, naringenone, α-pinene, eugenol, sabinene, terpinene, γ-terpinene, p-cymene, and phellandrene.
3. The application of cyclic terpenoid natural products and their derivatives as agricultural fungicides in plant diseases according to claim 2, characterized in that: The cyclic terpenoid natural products and their derivatives are nobutol or citric acid aldehyde.
4. The application of cyclic terpenoid natural products and their derivatives as agricultural fungicides in plant diseases according to claims 1-3, characterized in that: Plant diseases include bacterial diseases, fungal diseases, and viral diseases. Bacterial diseases include those caused by pathogens such as rice bacterial blight, *Xanthomonas oryzae* (rice leaf spot pathogen), *Xanthomonas carpetii*, *Xanthomonas brasiliensis* (rapeseed pathogen), *Pseudomonas syringae* (tomato leaf spot pathogen), *Pseudomonas syringae* (kiwifruit pathogen), *Ralstonia solanacearum*, *Pseudomonas syringae* (cucumber angular leaf spot pathogen), *Ralstonia solanacearum*, *Xanthomonas brasiliensis* (soybean pathogen), or *Corynebacterium micranthum* (tomato canker pathogen). Fungal diseases include those caused by cucumber gray mold, pepper wilt, rapeseed sclerotinia rot, wheat scab, potato late blight, blueberry root rot, *Gnaphalium affine*, dragon fruit anthracnose, rice sheath blight, Fusarium oxysporum SP, or *Colletotrichum sorghum*. Viral diseases include those caused by tobacco mosaic virus.
5. A method for preventing and controlling agricultural diseases, characterized in that: The above-mentioned cyclic terpenoid natural products and their derivatives are used for disease control, including rice bacterial leaf blight, rice bacterial leaf streak, citrus canker, cabbage black rot, tomato bacterial spot, kiwifruit canker, tobacco bacterial wilt, cucumber bacterial angular leaf spot, potato bacterial wilt, soybean bacterial spot or tomato canker, cucumber gray mold, pepper wilt, rapeseed sclerotinia rot, wheat scab, potato late blight, blueberry root rot, grape coccidioidomyces, dragon fruit anthracnose, rice sheath blight, diseases caused by Fusarium SP or sorghum spore spores, and tobacco mosaic virus.