A top flower bench fruit alkaloid compound with insecticidal activity and a preparation method and application thereof

Abstract

This invention belongs to the field of pesticide and phytochemistry technology, specifically relating to an insecticidal alkaloid compound from *Prunus persica* var. *pinnata*, its preparation method, and its application. The structure of the *Prunus persica* var. *pinnata* alkaloid compound is shown below. This alkaloid compound, isolated from *Prunus persica* var. *pinnata*, is designated as compound Pactermine X. It is a novel alkaloid isolated from *Prunus persica* var. *pinnata*. Its stomach poison and repellency activities against pea aphids were determined. The results showed that compound Pactermine X exhibited strong LC50 activity against pea aphids at 48 h. 50 The concentration was 0.272 mg / mL. At concentrations of 0.5 mg / mL, 0.25 mg / mL, 0.125 mg / mL, and 0.0625 mg / mL, the avoidance rates over 12 hours were 100%, 72.5%, 42.5%, and 30%, respectively.

Description

Technical Field

[0001] This invention belongs to the field of pesticide and phytochemistry technology, specifically relating to an insecticidal alkaloid compound from the fruit of the top flower (Benjamina stenoptera), its preparation method, and its application. Background Technology

[0002] With the continuous improvement of people's living standards, the quality and safety of agricultural products are receiving increasing attention. The safety issues arising from pesticide residues and resistance cannot be ignored. Therefore, the research and development of environmentally friendly plant-derived pesticides has garnered more attention and is one of the hot topics and effective approaches in the research and creation of new pesticides. Currently, plant-derived pesticides with matrine, azadirachtin, rotenone, and pyrethroids as their main active ingredients have shown promising application potential, providing a basis for further exploration of plant-derived pesticides with application value.

[0003] Currently, compounds used to control plant diseases include chemically synthesized insecticides (such as pyrethroids, organophosphates, and carbamates) and plant-derived insecticides (such as pyrethrin, matrine, and azadirachtin). Although these compounds can control the damage caused by piercing-sucking pests such as pea aphids to some extent, they have problems such as insufficient stomach poison activity and repellency activity against piercing-sucking pests like pea aphids, and difficulty in exerting effective inhibitory effects during the feeding and colonization stages of pests. Summary of the Invention

[0004] To address the problem that existing compounds used in the prevention and control of plant diseases have insufficient stomach poison and repellent activity against piercing-sucking pests such as pea aphids, and are difficult to effectively inhibit pests during their feeding and colonization stages, this invention provides an insecticidal alkaloid compound from the fruit of the pea pod, its preparation method, and its application.

[0005] To achieve the above objectives, the present invention adopts the following technical solution.

[0006] This invention provides an alkaloid compound of *Berberis vulgaris* with insecticidal activity, the structure of which is shown below: .

[0007] This invention isolates a novel alkaloid compound from the pea aphid, designated as Pactermine X, or simply Pactermine X. Experimental studies have shown that Pactermine X exhibits stomach poisoning activity against the pea aphid, with an LC50 value of [missing information]. 50The concentration was 0.272 mg / mL, and it also had certain inhibitory activity. At concentrations of 0.5 mg / mL, 0.25 mg / mL, 0.125 mg / mL and 0.0625 mg / mL, the inhibitory rates after 12 hours were 100%, 72.5%, 42.5% and 30%, respectively.

[0008] Among them, the top flower bench fruit Pachysandra terminalis Siebold Zucc. is an evergreen plant belonging to the genus Siebold Zucc. in the Buxaceae family, widely distributed in my country and Japan. The chemical components isolated from Siebold Zucc. mainly include alkaloids, triterpenes, and volatile oils. It has the effects of clearing heat and detoxifying, reducing swelling and relieving pain. Modern pharmacological studies have shown that it has anti-tumor, antioxidant, antibacterial, and insecticidal activities. The Siebold Zucc. described above was collected in September 2020 from the Taibai Mountain area of ​​Baoji, Shaanxi Province, and dried in the shade at room temperature before use.

[0009] The alkaloid compound of the fruit (compound Pactermine X (C)) is found in the fruit. 14 H 25 NO); Physicochemical properties: yellowish-brown oily substance; Specific rotation: [ α ] 20 D +12.0 ( c 0.1, MeOH), High-resolution mass spectrometry: HR-ESI-MS: m / z 224.2016 [M + H] + Infrared spectrum: IR (KBr) νmax: 2964, 2877, 1678, 1470, 1425, 1196, 1132, 813, 720 cm⁻¹ -1 ; 1 H-NMR spectrum and 13 The C-NMR spectral data are shown in Table 1, and the spectra are shown in the attached figure.

[0010] The alkaloid compound of *Pactermine X* provided by this invention has good insecticidal activity, especially showing significant stomach poison and repellent activity against pea aphids. It can play an inhibitory role during the feeding and colonization stages of pests. It has high stomach poison and repellent activity against piercing-sucking pests such as pea aphids, thus solving the problem that existing compounds used to control plant diseases have insufficient stomach poison and repellent activity against piercing-sucking pests such as pea aphids, and are difficult to play an effective inhibitory role during the feeding and colonization stages of pests.

[0011] Preferably, the alkaloid compound of the fruit is obtained by separating it from the n-butanol phase of the ethanol extract of the fruit.

[0012] The present invention also provides a pharmaceutical composition using the alkaloid compound of the fruit of the top flower (Benjamina stenoptera) or a pharmaceutically acceptable salt thereof as the active ingredient.

[0013] The present invention also provides a plant-based insecticide, comprising the alkaloid compound of the fruit of the top flower or the pharmaceutical composition.

[0014] Preferably, it also includes a pesticide-acceptable carrier.

[0015] Preferably, the carrier is selected from any one or more of diatomaceous earth, attapulgite, bentonite, silica, kaolin, talc, and light calcium carbonate.

[0016] The present invention also provides the application of the alkaloid compound of the top-flowering fruit, the pharmaceutical composition, or the plant insecticide in the prevention and control of plant diseases.

[0017] Preferably, the plant diseases include those caused by the pea aphid.

[0018] This invention also provides a method for preparing the alkaloid compounds from the fruit of the top-flowering vine, comprising the following steps: Take the dried fruit of the top flower bench and extract it with 75%~85% ethanol by volume as the extraction solvent. Concentrate the extract to obtain an ethanol extract. Dissolve the ethanol extract in water and extract it sequentially with petroleum ether, dichloromethane and n-butanol. Collect the n-butanol extract and concentrate it to obtain the n-butanol phase of the ethanol extract of the top flower bench fruit.

[0019] Dissolve the n-butanol phase of the ethanol extract of the top-flowered jujube in water, acidify with dilute hydrochloric acid, extract with petroleum ether, discard the petroleum ether layer; alkalize the aqueous layer, extract with chloroform to obtain a total alkaloid extract.

[0020] The total alkaloid extract was separated and purified by semi-preparative liquid chromatography to obtain the alkaloid compounds of the fruit of the top flower, *Dendrobium nobile*.

[0021] Preferably, the separation and purification steps are as follows: The total alkaloid extract was subjected to a first separation by semi-preparative liquid chromatography, and the fraction Fr.3 with a retention time of 25 min to 40 min was collected. The chromatographic conditions for the first separation were as follows: ShimNex UP C18 column 5 μm, 21.2 mm × 250 mm, mobile phase was a solution of methanol and 0.1% trifluoroacetic acid (methanol-0.1% trifluoroacetic acid aqueous solution) with a volume ratio of 60:40, and flow rate was 5 mL / min.

[0022] The fraction Fr.3 was subjected to a second separation by semi-preparative liquid chromatography, and the fraction Fr.3-2 with a retention time of 30 min to 50 min was collected. The chromatographic conditions for the second separation were as follows: ShimNex UP C18 column 5 μm, 21.2 mm × 250 mm, mobile phase was a solution of methanol and 0.1% trifluoroacetic acid in a volume ratio of 55:45 (methanol-0.1% trifluoroacetic acid aqueous solution), and flow rate was 5 mL / min.

[0023] The fraction Fr.3-2 was purified sequentially by gel chromatography and semi-preparative liquid chromatography, and the fraction with a retention time of 14 min to 19 min was collected to obtain the alkaloid compounds of the fruit of the top flower. The chromatographic conditions for purification by semi-preparative liquid chromatography were as follows: Nucifera C30C column 10 μm, 10 mm × 250 mm, mobile phase was a solution of acetonitrile and water with a volume ratio of 25:75 (acetonitrile-water solution), and flow rate was 3 mL / min.

[0024] Compared with the prior art, the present invention has the following beneficial effects: This invention provides an alkaloid compound from the pea aphid (compound Pactermine X), which exhibits good insecticidal activity, specifically demonstrating significant stomach poison and repellent activity against the pea aphid. This is because the invention identifies the insecticidal active ingredient in the pea aphid as a novel alkaloid monomer compound with a defined structure, thereby enabling the active ingredient to directly act on the target pest and exert an inhibitory effect during the pest's feeding and colonization stages. The experimental results in the instruction manual show that the alkaloid compound of *Prunus persica* has an LC50 of 0.272 mg / mL against pea aphids at 48 h, and the repellency rates at 12 h are 100%, 72.5%, 42.5%, and 30% at concentrations of 0.5 mg / mL, 0.25 mg / mL, 0.125 mg / mL, and 0.0625 mg / mL, respectively. This indicates that the alkaloid compound of *Prunus persica* not only has a toxic effect but also effectively inhibits pest attraction and feeding. Therefore, it solves the problems of resistance risk, large pesticide residues, and insufficient environmental compatibility caused by long-term use of chemical pesticides in current technologies, as well as the limited sources and unclear composition of plant-derived insecticidal active ingredients. Compared to existing plant-derived pesticides with matrine, azadirachtin, rotenone, and pyrethrin as the main active ingredients, or technical solutions that only focus on crude plant extracts or particulate extracts, the alkaloid compound from *Berberis vulgaris* provided by this invention is a new monomeric alkaloid compound derived from *Berberis vulgaris*. This alkaloid compound not only has clearly defined active ingredients and a well-defined structure, making it suitable for development as a new plant-derived insecticidal lead compound, but also facilitating subsequent quality control, stable preparation, and formulation research. Furthermore, it exerts an inhibitory effect during the feeding and colonization stages of pests, exhibiting high stomach poison and repellency activity against piercing-sucking pests such as pea aphids. This solves the problem that existing compounds used for plant disease control have insufficient stomach poison and repellency activity against piercing-sucking pests such as pea aphids, and are unable to effectively inhibit pests during their feeding and colonization stages. Attached Figure Description

[0025] Figure 1 This is the infrared spectrum of compound 1 in this invention.

[0026] Figure 2 This is the hydrogen spectrum of compound 1 in this invention.

[0027] Figure 3 This is the carbon spectrum of compound 1 in this invention.

[0028] Figure 4 This is the DEPT spectrum of compound 1 in this invention.

[0029] Figure 5 This is the HSQC spectrum of compound 1 in this invention.

[0030] Figure 6 Compound 1 in this invention 1 H- 1 H COSY spectrum.

[0031] Figure 7 This is the HMBC spectrum of compound 1 in this invention.

[0032] Figure 8 This is the NOESY spectrum of compound 1 in this invention.

[0033] Figure 9 This is the HRESIMS spectrum of compound 1 in this invention.

[0034] Figure 10 This is the calculated ECD spectrum of compound 1 in this invention.

[0035] Figure 11 This is a graph showing the relationship between the 12-hour repellency rate and concentration of compound 1 in this invention.

[0036] Figure 12 This is a comparison of the insecticidal effect of compound 1 on pea aphids after 48 hours of treatment in this invention; where a: upward position before PactermineX treatment; b: leftward position before PactermineX treatment; c: downward position before PactermineX treatment; d: rightward position before PactermineX treatment; e: upward position before PactermineX treatment; f: upward position after PactermineX treatment for 48 hours; f~j: different directions after PactermineX treatment for 48 hours; PactermineX is compound 1. Detailed Implementation

[0037] The present invention will now be described in detail with reference to the accompanying drawings and specific embodiments, but this should not be construed as limiting the invention. Unless otherwise specified, the technical means used in the following embodiments are conventional means well known to those skilled in the art, and the materials, reagents, etc. used in the following embodiments are commercially available unless otherwise specified.

[0038] The compound Pactermine X of this invention was isolated from the ethanol extract of *Pactermine arguta* var. *arguta*. This *Pactermine arguta* var. *arguta* was collected from Taibai Mountain, Taibai County, Baoji City, Shaanxi Province, People's Republic of China.

[0039] pea aphid ( Acyrthosiphon pisum ) is a widely distributed agricultural pest, especially harmful to leguminous crops such as peas ( Pisum sativumPea aphids are notorious for their harmful effects. As an insect with piercing-sucking mouthparts, the pea aphid relies on sucking plant sap to obtain the nutrients it needs. This behavior not only directly weakens the plant's growth potential but also induces a series of physiological damages, specifically manifested as leaf curling, malformation, and slow growth. These damages further reduce the crop's photosynthetic efficiency, affecting its nutrient absorption and ultimately causing a significant decrease in crop yield. Common chemical pesticides include organophosphates, carbamates, neonicotinoids, and insect growth regulators. These insecticides rapidly reduce aphid populations and achieve control by interfering with the aphid's nervous system and hindering its growth and development. However, long-term use of chemical pesticides has drawbacks such as resistance and residues. Compared to chemical pesticides and traditional biological control methods, traditional Chinese medicine has shown many advantages in controlling pea aphids. For example, its active ingredients are mostly low in toxicity and environmentally friendly, giving it a unique advantage in reducing pesticide residues and protecting the ecological environment. Overall, with the increasing demand for green and sustainable agricultural solutions, traditional Chinese medicine (TCM) has broad application prospects in agricultural pest control, especially in controlling pests such as pea aphids, where it possesses unique advantages. As the research and development of TCM pesticides deepens and market demand continues to expand, their application in global agricultural pest control will become increasingly widespread. Therefore, discovering effective agricultural insecticides from TCM has broad application prospects.

[0040] Example 1: Extraction method, identification, determination of antitumor activity, and application of compound Pactermine X (an alkaloid compound from the fruit of Pactermine X). 1. Experimental materials (1) Top-flowered Bench Fruit: Fresh top-flowered bench fruit is dried in the shade at room temperature before use. Among them, the top-flowered bench fruit was collected in September 2020 from Taibai Mountain, Taibai County, Baoji City, Shaanxi Province.

[0041] (2) Extraction of extract: 80% ethanol (v / v) was refluxed to extract the top flower of the fruit, and the extraction was repeated 3 times. The extract was concentrated to obtain 1 kg of extract (ethanol extract or ethanol extract of the top flower of the fruit).

[0042] (3) Reagents and instruments: Commonly used organic solvents: dichloromethane, chloroform, methanol, petroleum ether, and ethyl acetate are all industrial reagents and are used after redistillation. Organic solvents for chromatography: methanol, acetonitrile, trifluoroacetic acid, dilute hydrochloric acid, and ultrapure water are analytical grade or chromatographic grade reagents. Unless otherwise specified, all reagent volumes below are volume ratios.

[0043] Commonly Used Instruments: AVANCE400 NMR Spectrometer: Burker GmbH, Switzerland; Applied Photophysics Circular Dichroism Spectrometer: Applied Photophysics Ltd., UK; MCP 300 Polarimeter: Anton Paar GmbH, Germany; Fourier Transform Infrared Spectrometer: Shanghai Ruhai Optoelectronic Technology Co., Ltd.; Shimadzu LC-6AD Semi-Preparative Liquid Chromatograph: Beijing Beyond Future Technology Development Co., Ltd.; HB10digital Rotary Evaporator: IKA GmbH, Germany; HH-2 Thermostatic Water Bath: Changzhou Guohua Electric Co., Ltd.; DLSBseries Low-Temperature Condensing Circulating Pump: Zhengzhou Great Wall Science & Industry Trade Co., Ltd.; ZF-1 Three-Purpose Ultraviolet Analyzer: Shanghai Jinpeng Analytical Instrument Co., Ltd.; Shumei KQ-500DE Ultrasonic Cleaner: Kunshan City Ultrasonic Instruments Co., Ltd.; Cell Culture Incubator: NU-5800E, Nuaire, USA; Centrifuge: CR412, Giant, France; Autoclave: YXQ-LS-50SⅡ, Shanghai Boxun Industrial Co., Ltd.; Ultrasonic Cleaner: KQ-250E, Kunshan Ultrasonic Instruments Co., Ltd.; Electronic Analytical Balance: BT-125D, Sartorius, Germany; Ultra-Low Temperature Freezer: U410-85PRIME, NBS, USA; Electric Thermostatic Water Bath: DK-824, Shanghai Jinghong Experimental Equipment.

[0044] 2. Specific extraction and separation of compound Pactermine X The above-mentioned extract was dissolved in water and extracted sequentially with petroleum ether, dichloromethane, and n-butanol. The extract was separated into three types according to polarity. The n-butanol extract was collected and concentrated to obtain the n-butanol phase (n-butanol phase or n-butanol extract) of the ethanol extract of the fruit. In this invention, only the n-butanol phase was separated. The specific operation is as follows:

[0045] Dry the n-butanol extract in a cool place. Dissolve the n-butanol extract in 6 times its volume of water, acidify it to pH 1 with dilute hydrochloric acid (4%, v / v), extract the acidic aqueous solution with petroleum ether to obtain 25g of petroleum ether extract, adjust the pH of the lower acidic aqueous solution to 10 with 4% (v / v) sodium hydroxide solution, and extract with chloroform to obtain 120g of total alkaloid extract (total alkaloid extract obtained by chloroform extraction).

[0046] The total alkaloid extract obtained by chloroform extraction was first separated by semi-preparative liquid chromatography with isocratic elution. Fractions with the same retention time were combined to obtain four fractions Fr.1 to Fr.4: Fr.1 (tR=12min~18min, 8.4g), Fr.2 (tR=18min~25min, 9.5g), Fr.3 (tR=25min~40min, 39.1g), and Fr.4 (tR=40min~60min, 59.8g). The fraction Fr.3 with a retention time of 25min~40min was collected.

[0047] The chromatographic conditions for the first separation were as follows: a ShimNex UP C18 column of 5 μm, 21.2 mm × 250 mm, a mobile phase of an aqueous solution of methanol and 0.1% (v / v) trifluoroacetic acid at a volume ratio of 60:40 (methanol-0.1% trifluoroacetic acid aqueous solution), and a flow rate of 5 mL / min.

[0048] Fraction Fr.3 was subjected to a second separation using a semi-preparative liquid phase, followed by isocratic elution. Fractions with the same retention time were combined to obtain three fractions: Fr.3-1 to Fr.3-3. Fr.3-1 (tR = 15 min ~ 30 min, 4.0 g), Fr.3-2 (tR = 30 min ~ 50 min, 15.0 g), and Fr.3-3 (tR = 50 min ~ 60 min, 1.9 g). Fraction Fr.3-2, with a retention time of 30 min ~ 50 min, was collected.

[0049] The chromatographic conditions for the second separation were as follows: a ShimNex UP C18 column of 5 μm, 21.2 mm × 250 mm, a mobile phase of an aqueous solution of methanol and 0.1% (v / v) trifluoroacetic acid in a volume ratio of 55:45 (methanol-0.1% trifluoroacetic acid aqueous solution), and a flow rate of 5 mL / min.

[0050] The fraction Fr.3-2 was purified by gel chromatography using a Sephadex LH-20 (MeOH) column, followed by purification by semi-preparative liquid chromatography. Isocratic elution was performed, and the fraction with a retention time of 14-19 min was collected to obtain the alkaloid compound from the fruit of *Pactermine X*. t R =14min~19min, 7.5g), denoted as compound 1; wherein, the chromatographic conditions for purification are: chromatographic column Nucifera C30C 10μm, 10mm×250mm, mobile phase is a solution of acetonitrile and water with a volume ratio of 25:75 (acetonitrile-water solution), flow rate 3mL / min.

[0051] 3. Specific physicochemical properties of compound Pactermine X were tested. The nuclear magnetic resonance spectrometer was used to determine the composition of compound 1 at room temperature using CD3OD as the solvent. 1 H NMR (400MHz) and 13 C10 NMR (100MHz) spectrum, with chemical shifts using tetramethylsilane (TMS) as an internal standard.

[0052] 1 H-NMR spectrum and 13 C-NMR spectral data are shown in Table 1, and NMR and mass spectrometry data are shown in Table 2. Figures 1-10 .

[0053] Table 1. Compound 1 1 H (400MHz) and 13 C (100Hz) NMR data (CD3OD) Note: "—" indicates that the quaternary carbon has no proton hydrogen signal.

[0054] The physicochemical properties of the alkaloid compound 1 (compound Pactermine X) of the present invention are as follows: Pactermine X, C 14 H 25 NO, yellowish-brown oily substance; physicochemical properties: yellowish-brown oily substance; specific rotation: [α]20D +12.0 (c 0.1, MeOH); high-resolution mass spectrometry: HR-ESI-MS: m / z 224.2016 [M + H]+; infrared spectrum: IR (KBr) νmax: 2964, 2877, 1678, 1470, 1425, 1196, 1132, 813, 720 cm⁻¹ -1 .

[0055] 4. Tests on the stomach poison and repellency activities of compound Pactermine X against pea aphids. Following the artificial feed-medication method, specifically referring to "Geyter ED, Smagghe G, Rahbe Y, Geelen D. Triterpene saponins of Quillaja saponaria show strong aphicidal and deterrent activity against the pea aphid Acyrthosiphon pisum. Pest Management Science, 2012; 68:164–169," the stomach poison and repellent activities of compound Pactermine X were tested using the pea aphid as the test pest. The pea aphid population was collected from an alfalfa field in Dongzhang Village, Qindu District, Xianyang City, Shaanxi Province, and subsequently cultured artificially in the laboratory.

[0056] The results showed that compound Pactermine X had a 48-hour LC50 response to pea aphids. 50 The concentration was 0.272 mg / mL. At concentrations of 0.5 mg / mL, 0.25 mg / mL, 0.125 mg / mL, and 0.0625 mg / mL, the avoidance rates over 12 hours were 100%, 72.5%, 42.5%, and 30%, respectively. (See attached table for avoidance rates.) Figure 11 and Figure 12 .

[0057] As can be seen from the above, the novel compound 1 (compound Pactermine X) of the present invention exhibits good stomach poison activity and repellency activity against the hemiptera pest, the pea aphid.

[0058] In conclusion, alkaloid compounds can be developed as potential lead compounds for the control of pea aphids.

[0059] The preferred embodiments of this disclosure have been described in detail above with reference to the accompanying drawings. However, this disclosure is not limited to the specific details of the above embodiments. Within the scope of the technical concept of this disclosure, various simple modifications can be made to the technical solutions of this disclosure, and these simple modifications all fall within the protection scope of this disclosure.

[0060] 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, this disclosure will not describe the various possible combinations separately.

[0061] Furthermore, various different embodiments of this disclosure can be combined in any way, as long as they do not violate the spirit of this disclosure, they should also be regarded as the content disclosed in this disclosure.

[0062] This invention addresses the technical problems of existing aphid control methods, such as the long-term reliance on chemical pesticides leading to resistance, high pesticide residues, and insufficient environmental compatibility. It provides a novel alkaloid compound (compound Pactermine X) derived from the pea aphid, along with its preparation method and applications. Compared to existing methods that primarily rely on traditional chemical pesticides or only utilize crude plant extracts or particulate extracts, this invention identifies the insecticidal active ingredient in the pea aphid as a structurally defined monomeric compound, thus making the active ingredient more clearly defined and facilitating subsequent quality control, stable preparation, and formulation development. Furthermore, experimental results in this specification demonstrate that compound Pactermine X exhibits clear stomach poison and repellent activity against the pea aphid, with a 48-h LC50... 50The concentration was 0.272 mg / mL, and the repellency effect increased with increasing concentration at different concentrations, indicating that the compound Pactermine X not only has a toxic effect on the target pests but also inhibits their feeding and colonization stages. For piercing-sucking pests like the pea aphid, this dual effect helps reduce their sustained damage to plants. Therefore, the technical solution provided by this invention can effectively address and solve the problems in the background art regarding the lack of green and efficient insecticidal active ingredients, the large environmental burden of existing control methods, and the unclear composition of plant-derived active substances, and has good application prospects.

[0063] It should be noted that when numerical ranges are involved in this invention, it should be understood that the two endpoints of each numerical range and any value between the two endpoints can be selected. To avoid redundancy, this invention describes preferred embodiments.

[0064] Although preferred embodiments of the invention have been described, those skilled in the art, upon learning the basic inventive concept, can make other changes and modifications to these embodiments, all of which fall within the scope of the invention.

Claims

1. An alkaloid compound of the fruit *Prunus persica* with insecticidal activity, characterized in that, The structure of the alkaloid compound from the top-flowering bench fruit is shown below: 。 2. A pharmaceutical composition, characterized in that, The active ingredient is the alkaloid compound of the fruit of the top flower as described in claim 1 or its pharmaceutically acceptable salt.

3. A plant-based insecticide, characterized in that, It includes the alkaloid compound of the fruit of the top flower as described in claim 1 or the pharmaceutical composition as described in claim 2.

4. The plant-based insecticide according to claim 3, characterized in that, It also includes pesticide-acceptable carriers.

5. The plant-based insecticide according to claim 4, characterized in that, The carrier is selected from any one or more of diatomaceous earth, attapulgite, bentonite, silica, kaolin, talc, and light calcium carbonate.

6. The application of the alkaloid compound of the top-flowering fruit according to claim 1, the pharmaceutical composition according to claim 2, or the plant insecticide according to any one of claims 3 to 5 in the prevention and control of plant diseases.

7. The application according to claim 6, characterized in that, The plant diseases mentioned include those caused by the pea aphid.

8. The method for preparing the alkaloid compound of the fruit of the top-flowering vine as described in claim 1, characterized in that, Includes the following steps: Take the dried fruit of the top flower of the Bench Fruit and extract it with 75%~85% ethanol by volume as the extraction solvent. Concentrate the extract to obtain an ethanol extract. Dissolve the ethanol extract in water and extract it sequentially with petroleum ether, dichloromethane and n-butanol. Collect the n-butanol extract and concentrate it to obtain the n-butanol phase of the ethanol extract of the top flower of the Bench Fruit. The n-butanol phase of the ethanol extract of the top-flowered bench fruit was dissolved in water, acidified with dilute hydrochloric acid, and then extracted with petroleum ether. The petroleum ether layer was discarded. The aqueous layer was alkalized and then extracted with chloroform to obtain a total alkaloid extract. The total alkaloid extract was separated and purified by semi-preparative liquid chromatography to obtain the alkaloid compounds of the fruit of the top flower, *Dendrobium nobile*.

9. The preparation method according to claim 8, characterized in that, The separation and purification steps are as follows: The total alkaloid extract was subjected to a first separation by semi-preparative liquid chromatography, and the fraction Fr.3 with a retention time of 25 min to 40 min was collected. The chromatographic conditions for the first separation were as follows: ShimNex UP C18 column 5 μm, 21.2 mm × 250 mm, mobile phase was a solution of methanol and 0.1% trifluoroacetic acid with a volume ratio of 60:40, and flow rate was 5 mL / min. The fraction Fr.3 was subjected to a second separation by semi-preparative liquid chromatography, and the fraction Fr.3-2 with a retention time of 30 min to 50 min was collected. The chromatographic conditions for the second separation were as follows: ShimNex UP C18 column 5 μm, 21.2 mm × 250 mm, mobile phase was a solution of methanol and 0.1% trifluoroacetic acid with a volume ratio of 55:45, and flow rate was 5 mL / min. The fraction Fr.3-2 was purified sequentially by gel chromatography and semi-preparative liquid chromatography, and the fraction with a retention time of 14 min to 19 min was collected to obtain the alkaloid compounds of the fruit of the top flower. The chromatographic conditions for purification by semi-preparative liquid chromatography were as follows: Nucifera C30C column 10 μm, 10 mm × 250 mm, mobile phase was a solution of acetonitrile and water with a volume ratio of 25:75, and flow rate was 3 mL / min.

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