Plant-derived attractant for curculio of camellia oleifera and application thereof

Abstract

The application discloses a plant source attractant for Camptocerus aequatus and application thereof, and belongs to the technical field of plant disease and pest control. The plant source attractant comprises palmitic acid and / or plant alcohol. The palmitic acid and / or plant alcohol have significant and efficient attractant activity on the Camptocerus aequatus, and are suitable for detecting the population trend of the Camptocerus aequatus and efficient attraction and prevention and the like. Compared with the traditional chemical prevention and treatment method for the Camptocerus aequatus at the present stage, the prevention and treatment method for the Camptocerus aequatus by using the plant source attractant is more moderate, efficient, green, environment-friendly and non-toxic and side-effect, and can provide important value for market application and use popularization.

Description

Technical Field

[0001] This invention belongs to the field of plant disease and pest control technology, specifically relating to a plant-derived attractant for the camellia weevil and its application. Background Technology

[0002] Camellia oleifera Abel is a major woody oilseed crop in my country, and one of the four major woody oilseed plants in China and the world. Its main product is camellia oil, which, due to its various functional components, has broad market prospects across various industries. In the chemical industry, camellia oil is a raw material for biodiesel, possessing good combustion performance and environmentally friendly characteristics. Extracts such as camellia saponins can be used to produce surfactants, detergents, and other chemical products. In the pharmaceutical industry, camellia oil extracts, as well as some triterpenoids (camellia saponins) and phenolic compounds, have various pharmacological effects such as antibacterial and anti-inflammatory properties, and can be used in the development and utilization of pharmaceuticals and health products. In the health care industry, camellia oil is rich in unsaturated fatty acids such as oleic acid and linoleic acid, which have good health benefits for cardiovascular and cerebrovascular diseases and are often used as a nutritional supplement. In the catering industry, camellia oil is a high-quality edible oil, known as "Oriental olive oil," and is widely used in food processing. In recent years, with the economic development brought about by the camellia oil industry to various fields, the planting area has gradually expanded due to its needs, resulting in increasingly serious camellia oil pests and diseases. Therefore, a solution needs to be sought for the contradiction between the large-scale demand for camellia oil and camellia oil pests and diseases.

[0003] The fruit of the camellia oleifera plant is rich in nutrients during its growing season, and its fat content makes it an important food source for animals. Among the pests that damage camellia oleifera fruits is the camellia weevil (Curculio chinensis Chevrolat), also known as the tea seed weevil, belonging to the family Curculionidae in the order Coleoptera. It damages the fruit by feeding and laying eggs. Adults pierce and suck the juice from the camellia fruit, causing it to die. Larvae, having laid eggs on the adults, bore into the fruit and feed on the kernels, causing the fruit to fall off. Because the larvae hide inside the fruit during their larval stage, and the surrounding environment provides suitable conditions for the breeding and spread of the camellia weevil, traditional chemical control methods are ineffective.

[0004] Therefore, based on chemical ecology, researching and developing a milder, more efficient, green, environmentally friendly, and non-toxic camellia weevil control technology is an urgent technical problem to be solved in this field. Summary of the Invention

[0005] In view of this, the present invention provides a plant-derived attractant for the camellia weevil and its application. The plant-derived attractant has significant and efficient attraction activity for the camellia weevil and can be used to detect the population movement of the camellia weevil and for efficient attraction and control.

[0006] To achieve the above objectives, the present invention adopts the following technical solution:

[0007] The first objective of this invention is to provide a plant-derived attractant for the camellia weevil, said plant-derived attractant comprising palmitic acid and / or phytosterols.

[0008] Preferably, the concentration of palmitic acid is 55-222 ng / μL. More preferably, the concentration of palmitic acid is 55-111 ng / μL.

[0009] Preferably, the concentration of the phytosterol is 12.5-276 ng / μL. More preferably, the concentration of the phytosterol is 69-138 ng / μL.

[0010] Preferably, the palmitic acid and phytosterol are derived from tropist plants.

[0011] Preferably, the tropist plant includes *Paulownia tomentosa*.

[0012] The second objective of this invention is to provide the application of the plant-derived attractant of the Camellia oleifera weevil in the field of green pest control.

[0013] The third objective of this invention is to provide a method for using the plant-derived attractant for the camellia weevil, wherein the plant-derived attractant is added to a slow-release bottle and placed in an area where the camellia weevil is concentrated for trapping and killing.

[0014] Compared with the prior art, the present invention has the following beneficial effects:

[0015] This invention explores the attraction activities of two monomeric compounds, palmitic acid and / or phytol, on the camellia weevil using GC-MS and a Y-type selector. The effect of monomeric compound B, phytol, on the behavior of the camellia weevil is investigated using antennal potentiometry and a captive selection experiment, resulting in a plant-derived attractant with highly efficient attraction activity for the camellia weevil.

[0016] The plant-derived attractant provided by this invention has significant and efficient attraction activity for both female and male adult camellia weevils, and is suitable for detecting the population movement of camellia weevils, as well as for efficient attraction and control.

[0017] Compared to traditional chemical control methods, the plant-derived attractant method for controlling the camellia weevil provided by this invention is milder, more efficient, environmentally friendly, and has no toxic side effects, making it of great value in market application and promotion. Attached Figure Description

[0018] Figure 1 This is a total ion flux diagram of the plant crude extract from Example 1 of the present invention.

[0019] Figure 2 This is a comparison chart of the number of choices made by female and male adult camellia weevils regarding crude plant extracts and the control group in Example 2 of this invention.

[0020] Figure 3 This is a comparison diagram of the dwell time of female and male adult camellia weevils on both arms in Embodiment 2 of the present invention.

[0021] Figure 4 This is a comparison chart of the number of choices made by female and male adult camellia weevils and the control group for four monomeric compounds in Example 3 of this invention.

[0022] in, Figure 2 and Figure 3 In the table, “***” indicates significance for P < 0.001, “**” indicates significance for P < 0.01, “*” indicates significance for P < 0.05, and “ns” indicates no significance for P > 0.05.

[0023] Figure 5 This is a comparison chart of the residence time of female and male adult camellia weevils in four monomeric compounds and the control in Example 3 of the present invention.

[0024] Figure 6 This is a comparison chart of the number of selections made by female and male adult camellia weevils for three mixtures and a control in Example 3 of the present invention.

[0025] Figure 7 This is a comparison chart of the residence time of female and male adult camellia weevils in three compounds and the control in Example 3 of the present invention.

[0026] Figure 8 This is the antennal potential response of compound B at different concentrations in Example 4 of the present invention (significant differences between treatments with different letter representations, p<0.05).

[0027] Figure 9 This is the behavioral response of compound B at different concentrations in Example 5 of the present invention (significant differences between treatments with different letter representations, p<0.05). Detailed Implementation

[0028] Example 1: GC-MS Isolation and Identification of Crude Extracts from Leaves of Thygroscopic Plants

[0029] S1. Cold maceration extraction method for extracting crude plant extracts

[0030] The leaves of the tropist plant *Paulownia tomentosa*, after being air-dried, were dried in an oven at 60 °C until brittle. They were then removed, ground into powder, and stored. Using a cold maceration extraction method, the leaf powder and anhydrous ethanol were mixed in a 1:6 ratio and placed in a 1 L wide-mouth bottle. After thorough stirring, the mixture was extracted twice at 20–25 °C for 24 h each time. Both extracts were filtered through four layers of gauze, then through qualitative filter paper, and collected. The collected extract was then concentrated under reduced pressure in a rotary evaporator at 40 °C until no solvent dripped out, yielding a crude extract of the plant in an oily form. This crude extract was stored in a refrigerator at 4 °C for later use.

[0031] S2. Analysis and identification of components in crude plant extracts

[0032] The components and identification of the crude plant extract prepared in S1 were performed using gas chromatography-mass spectrometry (Varian 450-GC / 320-MS GC-MS), under the following conditions:

[0033] (1) Chromatographic conditions: Column: Rxi-50 column (30×0.25 mm 0.25 Micron); Injector temperature: 280℃; Carrier gas: high-purity helium; Carrier gas flow rate: 1 mL / min; Split ratio: 0; Vaporization temperature: 300℃.

[0034] (2) Mass spectrometry conditions: Ion source: EI; electron energy: 70 eV; ion source temperature: 240 ℃; interface temperature: 280 ℃; column pressure: 100 kPa; scanning voltage range: 33~500 m / z.

[0035] (3) GC heating program: The initial temperature is 80 ℃ and held for 1 min; the temperature is increased to 240 ℃ at a heating rate of 20 ℃ / min and held for 4 min; the temperature is increased to 280 ℃ at a heating rate of 20 ℃ / min and held for 5 min; 1 μL of the sample is injected after being diluted with anhydrous ethanol; the split ratio is 1:10.

[0036] S3. The results showed that, based on the analysis and identification of the components of the crude plant extract, and according to the ion flux of GC-MS... Figure 1 As shown, 21 compound peaks were identified through analysis. The crude extract contained 11 terpenoids, 9 aliphatic compounds, and 1 aromatic compound (see Table 1). The four most abundant monomeric compounds were hexamethylenetetramine, compound A (palmitic acid), compound B (phytosterol), and farnesol. Compound B (phytosterol) and farnesol are terpenoids; compound A (palmitic acid) and hexamethylenetetramine are aliphatic compounds.

[0037] Table 1

[0038]

[0039] Example 2: Y-shaped olfactory instrument test to verify the tropism behavior of crude plant extracts on the Camellia oleifera weevil.

[0040] (1) Concentration preparation: The crude plant extract was prepared into flavor source solutions of different concentrations (1000 ng / μL, 500 ng / μL, 250 ng / μL, 125 ng / μL, 62.5 ng / μL) with anhydrous ethanol as the control. The attraction activity of the crude plant extract to Camellia oleifera weevil was verified by a Y-type olfactometer.

[0041] (2) Experimental setup: The glass tube “Y”-shaped olfactory apparatus has two arms with a length of 20 cm each, a main arm with a length of 10 cm, and an angle of 75° between the two arms. The two arms are connected to the flavor source bottle, humidification bottle, activated carbon, glass rotor flow meter, and air delivery device in order from near to far using Teflon tubes. A 1.5×1.5 mm square filter paper is placed in each flavor source bottle. 30 μL of flavor source liquid of different concentrations is dropped onto one side of the filter paper as a treatment, and 30 μL of solvent is dropped onto the other side as a control. The air pump flow rate is controlled by the glass rotor flow meter and stabilized at 200 mL / min. Air is then continued for 1 min to ensure that the “Y”-shaped olfactory apparatus is filled with gas. The olfactory apparatus is placed under a parallel light source to ensure that the light intensity is evenly distributed on both arms of the “Y”-shaped olfactory apparatus, and to ensure that the experimental environment is consistent with the breeding environment.

[0042] (3) Measurement standard: One tea oil weevil that has been starved overnight is placed at the main arm of the "Y"-shaped tube. The behavior of the adult beetle is continuously observed and recorded within 10 minutes. Timing begins when the adult beetle reaches one end of the side wall of the olfactory instrument and ends when it leaves the side wall. When the adult beetle reaches 1 / 3 of the side wall of the olfactory instrument within 10 minutes and stays for more than 1 minute, it is recorded as tactic attraction and the dwell time is obtained. Conversely, if it does not reach the side arm within 5 minutes and stays for more than 1 minute, it is recorded as no selection and is used as the identification time. The dwell time of each adult beetle is recorded separately until 30 female and 30 male beetles each respond. In order to avoid the position effect on the selection results and the interference of residual odor substances on the two selection arms, the position of the two arms of the "Y"-shaped olfactory instrument is changed every 5 adults tested. The inner wall of the tube is cleaned with 75% ethanol solution and dried with a hair dryer. The odor source is changed and a new "Y"-shaped olfactory instrument is used every 10 adults tested.

[0043] Percentage of dwell time = (Dwell time on the lateral arm / Total observation time (10 min)) × 100%

[0044] The results showed that, through observation and testing of tactic behavior on both arms, different concentrations of crude extract solutions (1000 ng / μL, 500 ng / μL, 250 ng / μL, 125 ng / μL, 62.5 ng / μL) demonstrated differences in the attraction of adult female and male *Camellia oleifera* weevils. Higher concentrations resulted in a greater number of adults and a longer residence time in the treated arms; lower concentrations resulted in fewer adults and a shorter residence time. Except for the 62.5 ng / μL concentration, the number of tactic insects at other concentrations was higher than that in the control group. Specifically, the number of tactic insects in male *Camellia oleifera* weevils at concentrations (500 ng / μL, 250 ng / μL, 125 ng / μL) was higher than that in females (see...). Figure 2-3 ).

[0045] Example 3: Detection of the attraction activity of a “Y”-type olfactory instrument on four monomeric compounds and their mixtures.

[0046] Based on the relative contents of the first four monomeric compounds in the crude plant extract in Example 1 and the tactic behavior test of the Camellia oleifera weevil in Example 2, the number of Camellia oleifera weevils induced by the four monomeric compounds under different concentration settings and different mixtures of the four monomeric compounds prepared with anhydrous ethanol were counted using a Y-type olfactometer (see Table 2) and compared with the control.

[0047] Table 2

[0048]

[0049] like Figure 4-7 As shown, compounds A (palmitic acid) and B (phytol), the mixture of compounds A (palmitic acid) and B (phytol), and the mixture of all four compounds significantly attracted both male and female Camellia weevil. Conversely, the other two compounds, farnesol and acetamiprid, showed no significant attraction. However, at 38 ng / μL, the females remained in the farnesol-treated group longer than the control group, indicating the onset of a tactic response. Furthermore, the attraction effects of different concentrations of the compounds on male and female Camellia weevil varied. Compound A at 55 ng / μL only significantly attracted males, with the attraction effect at higher concentrations being greater for males than females. Compound B at 69 ng / μL only significantly attracted females, with the attraction effect at higher concentrations being greater for females than males. Therefore, the two active attractant components screened from the four compounds primarily act on males (palmitic acid) and females (phytol).

[0050] Example 4: Measurement of the antennal potential response of compound B (phytol) to the camellia weevil using an insect antennal potentiometer.

[0051] The antennal electrophysiological responses of the camellia weevil to different concentrations of compound B (phytosterol) were tested using an insect antennal electrophysiometer (IDAC-2, Syntech, Kirchzarten) in order to screen for the optimal dose-response effect.

[0052] Antennae potential settings: Y-Ampl 0.5mV, time base 15 s, stimulating gas flow rate 300 mL, stimulation time 0.2 s, stimulation interval 60 s. First, six different concentrations of test liquids (200 ng / μL, 100 ng / μL, 50 ng / μL, 25 ng / μL, 12.5 ng / μL, and 6.25 ng / μL) were prepared using compound B standard with liquid paraffin and stored in sealed volumetric flasks. Next, camellia weevils were selected, and their antennae were cut off at the base. The two ends of the cut antennae were attached to electrodes with conductive adhesive. Liquid paraffin served as the control. The six different concentrations of test liquids were tested sequentially from low to high. Only one antenna was used per adult insect, and each antenna was measured eight times.

[0053] Antennae potential relative response value = (response value after induction by test substance - response value after induction by control substance) / (response value after induction by reference substance - response value after induction by control substance) × 100%

[0054] The results showed that, at the selected six concentrations of compound B, different concentrations could induce electrophysiological responses in the antennae of the camellia weevil (see...). Figure 8 Starting from a concentration of 12.5 ng / μL, the antennal potential of the camellia weevil is positively correlated with its concentration, reaching its maximum at the highest concentration of 200 ng / μL. Its reaction intensity increases with the increase of the concentration of compound B.

[0055] Example 5: Envelopment selection experiment to test the behavioral response of the Camellia weevil to compound B (phytosterol).

[0056] Based on Example 4, a 200 ng / μL solution of compound B was dripped onto absorbent cotton and placed at both ends of a trap. Twenty overnight starved camellia weevils were placed in the center of the trap. After 30 hours, the behavioral responses of the camellia weevils were observed and recorded. Their selective behavior was determined based on the adults' feeding behavior on camellia fruits near the odor source.

[0057] Experimental results are as follows Figure 9 As shown, different concentrations of compound B exhibit varying degrees of attraction to the camellia weevil, with the attraction rate increasing with increasing concentration. At concentrations of 100 ng / μL and 200 ng / μL, the behavioral response of the camellia weevil was similar but significantly better than that of the solvent. Therefore, compound B at 100 ng / μL demonstrates the optimal attraction effect for the camellia weevil.

Claims

1. A plant-derived attractant for the camellia weevil, characterized in that, The plant-derived attractants include palmitic acid and phytosterols; The concentration of palmitic acid is 55-222 ng / μL; The concentration of the phytosterol is 12.5-276 ng / μL; The palmitic acid and phytol are derived from tropist plants; The tropist plants include the white-backed tung tree.

2. The plant-derived attractant for the camellia weevil according to claim 1, characterized in that, The concentration of palmitic acid is 55-111 ng / μL.

3. The plant-derived attractant for the camellia weevil according to claim 1, characterized in that, The concentration of the phytosterol is 69-138 ng / μL.

4. The application of a plant-derived attractant of the Camellia oleifera weevil as described in any one of claims 1-3 in the field of green pest control.

5. A method of using the plant-derived attractant for the Camellia oleifera weevil as described in any one of claims 1-3, characterized in that, The plant-derived attractant was added to a slow-release bottle and placed in an area where camellia weevils were concentrated for trapping and killing.

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