Sesquiterpene compound containing three-membered ring structure, preparation method of sesquiterpene compound and application of sesquiterpene compound in preparation of agricultural insecticide
The tri-ring structure sesquiterpene compound isolated from the marine fungus Penicillium sp.JX-2-1 solved the problem of insufficient anti-worm activity of marine terpenes, and prepared a highly efficient agricultural pesticide. In order to control Asian citrus psyllia, it has achieved a 95% mortality rate and provides a solution to green pesticides.
Patent Information
- Application Number
- CN202411380237.7
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- Filing Date
- 2024-09-30
- Publication Date
- 2025-01-03
AI Technical Summary
In the prior art, the anti-worm activity of terpenes from marine sources is rarely reported, especially the control effect of Asian citrus psyllium used to prevent and treat citrus Huanglong disease is poor, and the existing pesticides have environmental pollution and health risks, and lack green killing. Insect agent solutions.
A sesquiterpene compound containing trimetallic ring structure was isolated from the fermentation broth of marine fungus Penicillium sp.JX-2-1, and the compound was prepared by specific culture and extraction methods for the preparation of agricultural pesticides. Its high mortality rate has an insecticidal effect on Asian citrus psylliae.
The lethality rate of this compound to Asian citrus psyllia at a concentration of 1000 mg/kg can reach 95%, providing a natural and effective agricultural insecticide with significant insecticide effects and potential green pesticide application potential.
Smart Images
Figure CN119241484A_ABST
Abstract
Description
Technical Field
[0001] The invention belongs to the technical field of small molecule compounds, and specifically relates to a sesquiterpene compound containing a three-membered ring structure, a preparation method thereof, and an application thereof in the preparation of agricultural pesticides. Background Art
[0002] Terpenoid natural products are a large family of natural products, accounting for about 60% of known natural products. They have a wide range of biological activities such as anti-cancer, anti-inflammatory, antioxidant and anti-malarial. For example, paclitaxel with anti-cancer activity and the new antimalarial drug artemisinin are both terpenoid compounds in structure, attracting the research of many domestic scholars.
[0003] Due to its unique living environment: high salt, high pressure, low temperature and hypoxia, marine organisms have formed unique metabolic pathways and physical and chemical defense mechanisms, making it an important source of terpene small molecules. In particular, through the discovery of marine-derived terpene natural products published in recent years, most compounds have anti-tumor, antibacterial, anti-inflammatory and other activities, but there are relatively few reports on anti-insect activity, and most of the compounds reported to have anti-insect activity are halogenated compounds, such as: 8R-8-bromo-10-epi-β-snyderol isolated from red algae has an active effect on the reproduction of Plasmodium falciparum, and its IC50 is less than 10μg / mL.
[0004] Natural products have more significant structural diversity and complexity than synthetic compounds, and have a higher screening rate in terms of biological activity. Marine microbial secondary metabolites show high novelty and significant biological activity, such as antibacterial, anti-tumor, immunomodulatory, anti-inflammatory, enzyme inhibition, etc., and have become an important resource for the research and development of marine medicinal natural products.
[0005] In addition, the literature survey also found that citrus Huanglongbing is the fastest-spreading, most dangerous, most difficult and most destructive disease in global citrus production, and is also a scientific and technological problem that has not yet been overcome internationally. It can only be eliminated. At present, an extremely important and key measure for the prevention and control of citrus Huanglongbing is to suppress the risk of primary transmission and secondary transmission of Asian citrus psyllid.
[0006] Therefore, it is of great significance to screen out natural compounds with insecticidal properties and develop green pesticides. Summary of the invention
[0007] In order to overcome the shortcomings and deficiencies in the prior art, the primary purpose of the present invention is to provide a sesquiterpene compound containing a three-membered ring structure.
[0008] Another object of the present invention is to provide a method for preparing the above-mentioned sesquiterpene compound containing a three-membered ring structure.
[0009] Another object of the present invention is to provide the use of the above-mentioned sesquiterpene compound containing a three-membered ring structure in the preparation of agricultural pesticides.
[0010] The purpose of the present invention is achieved through the following technical solutions:
[0011] A sesquiterpene compound containing a three-membered ring structure, the compound having a structure as shown in the following formula 1:
[0012]
[0013] The compound is isolated from the fermentation liquid of the marine fungus Penicillium sp. JX-2-1 with a deposit number of GDMCC No: 60525.
[0014] The above-mentioned method for preparing a sesquiterpene compound containing a three-membered ring structure comprises the following steps:
[0015] S1. Inoculate the marine fungus Penicillium sp. JX-2-1 into the seed culture medium and culture it in a shaking table to obtain a seed culture solution;
[0016] S2. The seed culture solution is inoculated into a solid fermentation medium, and the culture is allowed to stand to obtain a fermentation product;
[0017] S3. The fermentation product is manually crushed, ultrasonically extracted with ethyl acetate three times, the extract is concentrated to obtain an extract, and then column chromatography separation technology is used to obtain a compound with a structural formula as shown in Formula 1.
[0018] The formula of the seed culture medium in step S1 is: on average, each liter of the seed culture medium includes 200 grams of potatoes, 20 grams of glucose, 33.3 grams of sodium chloride, and the balance is water.
[0019] The shaking culture conditions in step S1 are shaking speed of 200 rpm and culture at 25-30° C. for 3-7 days.
[0020] The formula of the solid fermentation medium in step S2 is: on average, per liter of the fermentation solid medium, the fermentation solid medium includes 1000 grams of rice, 333 grams of sea salt, and the balance is water. (Compared with the solid fermentation medium of the prior art, the proportion of sea salt is changed. The present invention uses high salt, preferably natural sea salt)
[0021] The static culture conditions in step S2 are 25-30° C. for 25-50 days.
[0022] The column chromatography separation technology described in step S3 includes silica gel column chromatography, gel column chromatography and C-18 reverse phase column chromatography. The specific operation is: the extract is subjected to column chromatography separation using a silica gel column, and gradient elution is performed using petroleum ether-ethyl acetate with a volume ratio of 95:5 to 60:40 as eluent, and the petroleum ether-ethyl acetate eluate with a volume ratio of 90:10 is collected, and then repeated column chromatography separation is performed using a reverse phase ODS silica gel column; the obtained eluate is then chromatographed on a C-18 reverse phase liquid chromatography column, and prepared and collected with a methanol aqueous solution with a volume percentage concentration of 84%, to obtain a sesquiterpene compound containing a three-membered ring structure.
[0023] The application of the above-mentioned sesquiterpene compound containing a three-membered ring structure in the preparation of agricultural insecticides is an agent for killing Asian citrus psyllids.
[0024] Compared with the prior art, the present invention has the following advantages and beneficial effects:
[0025] The present invention provides a sesquiterpene compound containing a three-membered ring structure, which is a compound derived from marine fungi. The compound has the activity of killing Asian citrus psyllids, and the lethality rate can reach 95% at a concentration of 1000 mg / kg. The compound can be used to prepare natural insecticides. Therefore, the compound derived from marine fungi provided by the present invention has the potential for application as an agricultural insecticide. BRIEF DESCRIPTION OF THE DRAWINGS
[0026] Figure 1 This is the high-resolution mass spectrum (HR-ESI-EI) of compound 1.
[0027] Figure 2 Compound 1 1 H NMR spectra.
[0028] Figure 3 Compound 1 13 C NMR spectrum. DETAILED DESCRIPTION
[0029] The present invention is further described in detail below in conjunction with embodiments and drawings, but the embodiments of the present invention are not limited thereto.
[0030] Example 1: Isolation and identification of fungus Penicillium sp. JX-2-1
[0031] 1. The strain was isolated from the leaves of the mangrove plant Kandelia candel and identified as Penicillium sp. by gene sequencing. The Penicillium sp. JX-2-1 strain of GDMCC No. 60525 of marine fungi was deposited in Guangdong Microbiological Culture Collection Center on January 4, 2019, with the deposit number of GDMCC No. 60525.
[0032] Example 2: Preparation of compounds
[0033] 1. Isolate and prepare compounds from the metabolites of the fungus Penicillium sp. JX-2-1, the steps are as follows:
[0034] (1) Seed culture of Penicillium sp. JX-2-1:
[0035] Seed culture medium composition: 200 grams of potatoes, 20 grams of glucose, 33.3 grams of sodium chloride, add water to 1L (specific preparation method: mix and dissolve each component and shake well).
[0036] After the culture medium is sterilized at high temperature, the strain is picked up with an inoculation loop and inoculated, and cultured in a constant temperature shaker at 25°C for 3 days to obtain the seed culture solution.
[0037] (2) Fermentation culture of fungus Penicillium sp. JX-2-1:
[0038] The solid fermentation medium formula is 1000 grams of rice, 333 grams of sea salt, add deionized water to 1L, stir until completely dissolved, divide into 500ml triangular bottles, and sterilize at 121℃ for 20 minutes.
[0039] Transfer the strain in the seed solution into a conical flask filled with culture medium, with an inoculation volume of 8-20 ml, and always control the temperature at 28°C. The culture time is 30 days.
[0040] (3) The fermentation product of step (2) is manually crushed, ultrasonically extracted, extracted and concentrated with ethyl acetate; the concentrated extract is separated by column chromatography, and gradient elution is performed with petroleum ether-ethyl acetate in a volume ratio of 95:5 to 60:40 as eluent, and the petroleum ether-ethyl acetate eluate in a volume ratio of 90:10 is collected, and then repeated column chromatography separation is performed using a reversed-phase ODS silica gel column; the obtained eluate is then chromatographed and separated using a C-18 reversed-phase liquid chromatography column, and a methanol aqueous solution with a volume percentage concentration of 84% is prepared and collected to obtain a sesquiterpene compound containing a three-membered ring structure.
[0041] During separation, a 95:5---60:40 petroleum ether-ethyl acetate gradient elution was used respectively, and the 90:10 petroleum ether-ethyl acetate gradient eluate was combined and collected; the 90:10 eluate was collected and then separated by reverse phase ODS silica gel column chromatography and prepared by semi-preparative reverse phase C-18 reverse phase column, and finally compound 1 was obtained.
[0042] Example 3: Structural analysis of compound 1
[0043] The compound 1 obtained in Example 2 was subjected to structural analysis to obtain the following experimental data: high resolution mass spectrum (HR-ESI-EI), 1 1 H NMR spectra and 1 13 C NMR spectra are shown in Figure 1 , Figure 2 and Figure 3 As shown:
[0044] Compound 1: white powder, HR-ESI-MS m / z 287.1254[M+Na] + (Theoretical value 287.1257, C 15 H 20 O4Na + ). One-dimensional NMR data: 1 H NMR 1.07(s),1.31(m),1.40(s),1.61(dt,14.0,4.0),1.69(td,14.0,4.0),1.84(m),2.01(overlapped),2.07
[0045] (overlapped),2.47(m),2.60(m),3.78(d,2.7),4.66(dd,16.4,2.6),4.74(dt,16.4,2.6),6.59(s); 13 C NMR 22.1, 25.7, 26.0, 30.6, 34.3, 38.4, 40.1, 71.9, 74.2, 75.2, 114.7, 126.7, 152.7, 166.4, 175.3. After identification, the structural formula of compound 1 is shown in Formula 1:
[0046]
[0047] Example 4: Compound Activity Test
[0048] Experiment on the inhibitory activity of compound 1 obtained in Example 2 against Asian citrus psyllid:
[0049] 1. Materials:
[0050] 1.1 Dimethyl sulfoxide (Mpbio, USA); Avermectin (Qilu Pharmaceutical);
[0051] 1.2 Insecticidal activity test of compounds:
[0052] This experiment adopted the residual exposure method, which was as follows: Compound 1 and avermectin were dissolved in dimethyl sulfoxide (100%) to a concentration of 1 mg / mL to obtain compound solution and avermectin solution, which were placed in a refrigerator for standby use; during the experiment, the compound solution and avermectin solution were sprayed on citrus leaves, and after 2 hours, the solution evaporated to dryness, and 20 Asian citrus psyllids were placed on the citrus leaves sprayed with the compound solution and avermectin solution, and the activity of Asian citrus psyllids was observed after 24 hours, and then the mortality rate was calculated. The blank group was sprayed with pure water for testing.
[0053] 2. Test results
[0054] The test results show that compound 1 has a significant inhibitory effect on Asian citrus psyllids. The inhibitory effect of compound 1 on Asian citrus psyllids is shown in Table 1.
[0055] Table 1 Mortality of compounds to Asian citrus psyllids
[0056]
[0057] The above embodiments are preferred implementation modes of the present invention, but the implementation modes of the present invention are not limited to the above embodiments. Any other changes, modifications, substitutions, combinations, and simplifications that do not deviate from the spirit and principles of the present invention should be equivalent replacement methods and are included in the protection scope of the present invention.
Claims
1. A sesquiterpene compound containing a three-membered ring structure, characterized in that: The compound has a structure shown in Formula 1:
2. A sesquiterpene compound containing a three-membered ring structure according to claim 1, characterized in that: The compound is isolated from the fermentation liquid of the marine fungus Penicillium sp. JX-2-1 with a deposit number of GDMCC No: 60525.
3. The method for preparing a sesquiterpene compound containing a three-membered ring structure according to claim 1, characterized in that The steps include: S1. Inoculate the marine fungus Penicillium sp. JX-2-1 into the seed culture medium and culture it in a shaking table to obtain a seed culture solution; S2. The seed culture solution is inoculated into a solid fermentation medium, and the culture is allowed to stand to obtain a fermentation product; S3. The fermentation product is manually crushed, ultrasonically extracted with ethyl acetate three times, the extract is concentrated to obtain an extract, and then column chromatography separation technology is used to obtain a compound with a structural formula as shown in Formula 1.
4. The preparation method according to claim 3, characterized in that: The formula of the seed culture medium in step S1 is: on average, each liter of the seed culture medium includes 200 grams of potatoes, 20 grams of glucose, 33.3 grams of sodium chloride, and the balance is water.
5. The preparation method according to claim 3, characterized in that: The shaking culture conditions in step S1 are shaking speed of 200 rpm and culture at 25-30° C. for 3-7 days.
6. The preparation method according to claim 3, characterized in that: The formula of the solid fermentation medium in step S2 is: on average, each liter of the fermentation solid medium includes 1000 grams of rice, 333 grams of sea salt, and the remainder is water.
7. The preparation method according to claim 3, characterized in that: The static culture conditions in step S2 are 25-30° C. for 25-50 days.
8. The preparation method according to claim 3, characterized in that: The column chromatography separation technology described in step S3 includes silica gel column chromatography, gel column chromatography and C-18 reverse phase column chromatography. The specific operation is: the extract is subjected to column chromatography separation using a silica gel column, and gradient elution is performed using petroleum ether-ethyl acetate with a volume ratio of 95:5 to 60:40 as eluent, and the petroleum ether-ethyl acetate eluate with a volume ratio of 90:10 is collected, and then repeated column chromatography separation is performed using a reverse phase ODS silica gel column; the obtained eluate is then chromatographed on a C-18 reverse phase liquid chromatography column, and prepared and collected with a methanol aqueous solution with a volume percentage concentration of 84%, to obtain a sesquiterpene compound containing a three-membered ring structure.
9. Use of a sesquiterpene compound containing a three-membered ring structure in the preparation of an agricultural pesticide according to claim 1, characterized in that: The insecticide is an agent for killing Asian citrus psyllids.