Use of beta-caryophyllene in the knockdown of b. tabaci

Abstract

The application belongs to the technical field of insect control, and discloses application of beta-caryophyllene in contact killing of Bemisia tabaci. It is found that beta-caryophyllene can play a contact killing role on Bemisia tabaci, and can cause rapid death of Bemisia tabaci MED cryptic species adults in a short time. The beta-caryophyllene is prepared into a beta-caryophyllene liposome preparation, and has good stability and dispersibility, thereby improving the control effect on Bemisia tabaci.

Description

TECHNICAL FIELD

[0001] The application belongs to the technical field of insect control, and particularly relates to application of beta-caryophyllen in contact killing of Bemisia tabaci. BACKGROUND

[0002] Bemisia tabaci is a polyphagous insect, and the host plant species are up to 600, which is a worldwide agricultural pest. The pest mainly lives on the back of plant leaves, feeds on plant sap, transmits various plant viruses, affects plant growth, and greatly reduces agricultural yield. At present, spraying chemical pesticides is the main method to control the number of Bemisia tabaci to protect crops. However, the overuse of pesticides can cause pests to develop resistance, and also cause great harm to humans and the environment. Natural plant source products such as plant volatile components and extracts have the characteristics of safety, greenness, environmental protection and the like. Therefore, in the replacement of chemical pesticides and the control of agricultural pests, natural plant source products have good application prospects.

[0003] Beta-caryophyllen (BCP) is a plant compound of bicyclic sesquiterpene, and is one of the main active ingredients in volatile oils of basil, black pepper, lavender, rosemary, oregano, hops, cinnamon leaf oil, clove leaf oil, vitex oil, and orange peel oil. Previous pharmacological studies have shown that BCP has local anesthetic, anti-inflammatory, mosquito-repelling, anti-anxiety, and anti-depression effects. Since beta-caryophyllen is relatively stable in volatility, it belongs to natural equivalent spices and artificial spices, and existing studies have focused on the repellent effect of volatile substances contained in facility crops on Bemisia tabaci, and it has been found that beta-caryophyllen has a natural repellent effect on Bemisia tabaci. Liu Yuchen et al. have shown that beta-caryophyllen is an important insecticidal active ingredient in Vitex trifolia seed extract, and beta-caryophyllen has contact killing and repellent effects on Aphis gossypii. Since different insects have different sensitivities to insecticides, the same drug can have completely different effects on insects. The prior art does not disclose the contact killing activity of beta-caryophyllen on Bemisia tabaci. SUMMARY

[0004] The purpose of the present application is to provide an application of beta-caryophyllen in contact killing of Bemisia tabaci, and to provide a new control method for Bemisia tabaci.

[0005] The technical solution of the present application is as follows:

[0006] The present application provides an application of beta-caryophyllen in contact killing of Bemisia tabaci. It is found that beta-caryophyllen can have a contact killing effect on Bemisia tabaci, and can cause rapid death of Bemisia tabaci MED cryptic adult in a short time.

[0007] As an implementable mode, the beta-caryophyllene is a beta-caryophyllene liposome. Since the beta-caryophyllene has defects of difficult solubility and poor dispersibility, the beta-caryophyllene liposome can be prepared to improve the dispersibility and increase the insecticidal effect. The skilled person in the art can also prepare the beta-caryophyllene into other pesticide formulations with good dispersing effect.

[0008] The application also provides a beta-caryophyllene liposome, which solves the problem of difficult dispersion of beta-caryophyllene in the application process. The beta-caryophyllene liposome comprises lecithin, cholesterol and beta-caryophyllene, and the ratio is 0.05-0.25 g:0.01-0.03 g:50-250 μL, and further preferably 0.1-0.2 g:0.015-0.025 g:100-200 μL. 。

[0009] Preferably, the beta-caryophyllene liposome further comprises a stabilizer.

[0010] Further preferably, the stabilizer comprises Tween 80 and / or chitosan.

[0011] As an implementation mode, in the beta-caryophyllene liposome, the ratio of beta-caryophyllene and Tween 80 is 50-250 μL:0.1-0.5 ml, and further 100-200 μL:0.2-0.4 ml.

[0012] As an implementation mode, in the beta-caryophyllene liposome, the ratio of beta-caryophyllene and chitosan is 50-250 μL:1-6 mg, and further 100-200 μL:2-4 mg.

[0013] The application also provides a preparation method of the above beta-caryophyllene liposome. The beta-caryophyllene, lecithin and cholesterol are ultrasonically dissolved with ethanol, and after adding the stabilizer, stirring and dispersing, rotary evaporation is performed to remove ethanol, and a proper amount of deionized water is added to dissolve the solute, so as to obtain the beta-caryophyllene liposome. The application uses nanoliposome as a release carrier, and uses the ultrasonic dispersion method-ethanol injection method to prepare the beta-caryophyllene liposome, so as to improve the dispersibility and insecticidal effect of the beta-caryophyllene, thereby effectively preventing and treating the whitefly.

[0014] Preferably, the stirring is performed by using a magnetic stirrer, and the setting parameters of the magnetic stirrer are 220 rpm and 10-20 min.

[0015] Preferably, the temperature of the rotary evaporation is below 40℃.

[0016] The application also includes the application of the above beta-caryophyllene liposome in preventing and treating the whitefly.

[0017] Preferably, the beta-caryophyllene liposome has a touch-killing effect on the whitefly.

[0018] Compared with the prior art, the present application has the following technical effects:

[0019] The present application provides an application of beta-caryophyllene in the knockdown of Bemisia tabaci. Beta-caryophyllene can play a knockdown role on Bemisia tabaci, and can cause rapid death of Bemisia tabaci MED cryptic adult in a short time. The mortality of Bemisia tabaci after 30 min of treatment with 0.125%, 0.25% and 0.5% beta-caryophyllene is 73.33%, 80.33% and 86.67%, respectively.

[0020] The beta-caryophyllene liposome provided by the present application has good combination of beta-caryophyllene and the slow-release carrier, the encapsulation rate can reach 70-80%, can delay the action time of caryophyllene to a certain extent, and improve the control effect on Bemisia tabaci. The control effect on Bemisia tabaci can reach 97±3.6%. BRIEF DESCRIPTION OF DRAWINGS

[0021] Figure 1 It is the knockdown effect of beta-caryophyllene on Bemisia tabaci 30 min after spraying.

[0022] Figure 2 It is the transmission electron microscope (TEM) image of the beta-caryophyllene liposome of Example 5.

[0023] Figure 3 It is the Fourier infrared spectrum (FTIR) of the beta-caryophyllene liposome.

[0024] Figure 4 It is the knockdown effect of beta-caryophyllene liposome preparation on Bemisia tabaci 30 min after spraying. DETAILED DESCRIPTION

[0025] The technical solutions of the present application will be further described in detail below in combination with specific embodiments. The technical solutions of the present application include but are not limited to the following embodiments.

[0026] The raw material components used in the following embodiments are all commercially available products. Among them, beta-caryophyllene is purchased from Shanghai Maikelin Reagent Co., Ltd., and the purity is >80%.

[0027] Example 1

[0028] Knockdown effect of beta-caryophyllene on Bemisia tabaci

[0029] Prepare 0.125%, 0.25% and 0.5% beta-caryophyllene liquid medicine: use 0.3 ml Tween 80 as an emulsifier, dilute beta-caryophyllene in 100 ml deionized water to a final concentration of 0.125, 0.25 and 0.5 μL / mL, respectively, and divide into 150 mL polyethylene sprayers with pump nozzles.

[0030] Method of application: The contact toxicity of 0.125%, 0.25% and 0.5% β-caryophyllene solution to Bemisia tabaci was determined. The solution was sprayed with a 150 mL sprayer, sprayed 3 times in a petri dish to form a uniform film, and the solution in the center of the petri dish was wiped clean with filter paper to form a 1 cm Bemisia tabaci release area. After 30 min, the survival of Bemisia tabaci in the petri dish was observed, and Bemisia tabaci that did not move or could not move normally was considered dead. The number of Bemisia tabaci in each group was 50, and the experiment was repeated 6 times, with sterile water as a control.

[0031] The mortality rates of Bemisia tabaci after 30 min of treatment with 0.125%, 0.25% and 0.5% β-caryophyllene solution were 73.33%, 80.33% and 86.67%, respectively, indicating that β-caryophyllene has contact toxicity to Bemisia tabaci.

[0032] Example 2

[0033] β-caryophyllene-containing liposomes (B-NPS)

[0034] 0.1 g of lecithin and 0.02 g of cholesterol were weighed, 150 uL of β-caryophyllene was measured into a conical flask, 20 mL of anhydrous ethanol was added, the beaker was sealed with a sealing film, and the mixture was ultrasonicated in an ultrasonic cleaner until the solute was fully dissolved. The ethanol was removed by rotary evaporation (40°C), and the solute was dissolved in an appropriate amount of deionized water to obtain β-caryophyllene-containing liposomes (B-NPS).

[0035] Example 3

[0036] β-caryophyllene and Tween 80-containing liposomes (BT-NPS)

[0037] 0.1 g of lecithin and 0.02 g of cholesterol were weighed, 150 uL of β-caryophyllene was measured into a conical flask, 20 mL of anhydrous ethanol was added, the beaker was sealed with a sealing film, and the mixture was ultrasonicated in an ultrasonic cleaner until the solute was fully dissolved. 0.2 mL of Tween 80 was added, and the solute was uniformly dispersed by a magnetic stirrer (220 rpm, 15 min). The ethanol was removed by rotary evaporation (40°C), and the solute was dissolved in an appropriate amount of deionized water to obtain a milky white β-caryophyllene liposome solution (BT-NPS).

[0038] Example 4

[0039] β-caryophyllene and chitosan-containing liposomes (H-B-NPS)

[0040] Take 0.1 g of lecithin, 0.02 g of cholesterol, and 150 uL of β-caryophyllene in a conical flask, add 20 mL of anhydrous ethanol, seal the beaker with a sealing film, and put it into an ultrasonic cleaner for ultrasonic, until the solute is fully dissolved. Add 0.2 mL of Tween 80, and uniformly disperse the solute by a magnetic stirrer (220 rpm, 15 min). Remove ethanol by rotary evaporation method (40°C), and add an appropriate amount of deionized water to dissolve the solute, to obtain a milky white solution (BT-NPS). Prepare a 0.2 mg / mL chitosan aqueous solution, and mix it with the BT-NPS solution at a volume ratio of 1:2. Obtain a β-caryophyllene liposome solution (H-BT-NPS) by ultrasonic dispersion method (360 w, ultrasonic for 10 min).

[0041] Example 5

[0042] β-caryophyllene, chitosan and Tween 80 containing liposome (H-BT-NPS)

[0043] Take 0.1 g of lecithin, 0.02 g of cholesterol, and 150 uL of β-caryophyllene in a conical flask, add 20 mL of anhydrous ethanol, seal the beaker with a sealing film, and put it into an ultrasonic cleaner for ultrasonic, until the solute is fully dissolved. Add 0.2 mL of Tween 80, and uniformly disperse the solute by a magnetic stirrer (220 rpm, 15 min). Remove ethanol by rotary evaporation method (40°C), and add an appropriate amount of deionized water to dissolve the solute, to obtain a milky white solution (BT-NPS). Prepare a 0.2 mg / mL chitosan aqueous solution, and mix it with the BT-NPS solution at a volume ratio of 1:2. Obtain a β-caryophyllene liposome solution (H-BT-NPS) by ultrasonic dispersion method (360 w, ultrasonic for 10 min).

[0044] Example 6

[0045] β-caryophyllene and Tween 80 containing liposome

[0046] Take 0.15 g of lecithin, 0.03 g of cholesterol, and 150 uL of β-caryophyllene in a conical flask, add 20 mL of anhydrous ethanol, seal the beaker with a sealing film, and put it into an ultrasonic cleaner for ultrasonic, until the solute is fully dissolved. Add 0.4 mL of Tween 80, and uniformly disperse the solute by a magnetic stirrer (220 rpm, 20 min). Remove ethanol by rotary evaporation method (40°C), and add an appropriate amount of deionized water to dissolve the solute, to obtain a milky white β-caryophyllene liposome solution.

[0047] Example 7

[0048] β-caryophyllene and chitosan containing liposome

[0049] Take 0.2 g of lecithin, 0.01 g of cholesterol, 150 uL of β-caryophyllene in a conical flask, add 20 mL of anhydrous ethanol, seal the beaker with a sealing film, and put it into an ultrasonic cleaner for ultrasonic, until the solute is fully dissolved. Remove ethanol by rotary evaporation method (40℃), add appropriate amount of deionized water to dissolve the solute, and obtain the β-caryophyllene-containing liposome (B-NPS). 6 mg of chitosan (HACC) is dissolved in 10 mL of sterile water to obtain a 0.6 mg / mL chitosan aqueous solution, then the HACC solution is mixed with the B-NPS solution at a volume ratio of 1:2. The β-caryophyllene liposome solution is obtained by ultrasonic dispersion method (360w, ultrasonic 10min).

[0050] Example 8

[0051] β-caryophyllene, chitosan and Tween80-containing liposome

[0052] Take 0.2 g of lecithin, 0.01 g of cholesterol, 150 uL of β-caryophyllene in a conical flask, add 20 mL of anhydrous ethanol, seal the beaker with a sealing film, and put it into an ultrasonic cleaner for ultrasonic, until the solute is fully dissolved. Add 0.3 mL of Tween80, and uniformly disperse the solute by magnetic stirrer (220 rpm, 15 min). Remove ethanol by rotary evaporation method (40℃), add appropriate amount of deionized water to dissolve the solute, and obtain a milky white solution (BT-NPS). Prepare a 1 mg / mL chitosan aqueous solution, and mix it with the BT-NPS solution at a volume ratio of 1:2. The β-caryophyllene liposome solution is obtained by ultrasonic dispersion method (360w, ultrasonic 10min).

[0053] Example 9

[0054] The particle size, zeta potential, encapsulation efficiency and other morphology of the β-caryophyllene liposome in Examples 2-5 are determined by transmission electron microscopy (TEM), Fourier transform infrared spectrometer (FTIR), nanoparticle size analyzer, gas chromatograph-mass spectrometer (GC-MS).

[0055] The morphology of the β-caryophyllene liposome preparation is spherical ( Figure 2 ), and the combination of β-caryophyllene and the sustained-release carrier is good ( Figure 3 ).

[0056] As can be seen from Table 1, the particle size of the β-caryophyllene-containing liposome preparation is large, and after adding the stabilizer, the particle size of the liposome is greatly reduced. The particle size of H-BT-NPS is 174.22±2.59 nm, which increases slightly after 14 days; the particle size of BT-NPS is 176.61±4.32, which remains basically unchanged after 14 days.

[0057] Table 1 Particle size of β-caryophyllene liposome preparation

[0058] Formulation 1 day (particle size / nm) 14 days (particle size / nm) NPS 233.43±2.93 248.39±4.21 B-NPS 1290.17±319.20 2691.11±698.48 BT-NPS 176.61±4.32 179.92±4.32 H-B-NPS 554.92±133.47 483.66±33.41 H-BT-NPS 174.22±2.59 201.28±4.09

[0059] As can be seen from Table 2, the dispersion (PDI) of the β-caryophyllene liposome preparation is improved after the addition of the stabilizer, and the dispersion can be maintained for 14 days. The dispersion of the β-caryophyllene liposome preparation added with chitosan and Tween 80 is 0.43±0.02.

[0060] Table 2 Dispersion of β-caryophyllene liposome preparation

[0061] Formulation 1 day (PDI) 14 days (PDI) NPS 0.26±0.01 0.28±0.01 B-NPS 1.00±0.01 0.84±0.15 BT-NPS 0.47±0.02 0.44±0.02 H-B-NPS 0.48±0.10 0.65±0.24 H-BT-NPS 0.43±0.02 0.45±0.02

[0062] As can be seen from Table 3, the potential of the β-caryophyllene liposome preparation is in the range of 39-60 mV after the addition of the stabilizer, and the potential stability is maintained for 14 days.

[0063] Table 3 Potential of β-caryophyllene liposome preparation

[0064]

[0065]

[0066] As can be seen from Table 4, the encapsulation efficiency of the β-caryophyllene liposome preparation prepared in the present application is 60-80%, and the encapsulation efficiency is not changed much for 14 days, and the content of β-caryophyllene in the preparation is maintained well.

[0067] Table 4 Encapsulation efficiency of β-caryophyllene liposome preparation

[0068] Formulation 1 day (%) 14 days (%) B-NPS 40.30±1.40 9.19±1.48 BT-NPS 75.80±0.36 53.26±1.33 H-B-NPS 60.26±0.84 38.82±0.89 H-BT-NPS 80.76±0.41 77.00±0.28

[0069] As can be seen from Table 5, after 14 days, the content of β-caryophyllene in B-NPS is decreased from 133.39±4.88 mg / L to 34.37±5.47 mg / L, and the content of β-caryophyllene in H-BT-NPS is decreased from 763.62±5.97 mg / L to 257.46±2.39 mg / L, which to some extent prolongs the action time of caryophyllene.

[0070] Table 5 Content of β-caryophyllene in β-caryophyllene liposome preparation

[0071] Formulation 1 day (mg / L) 14 days (mg / L) B-NPS 133.39±4.88 34.37±5.47 BT-NPS 709.04±1.44 205.09±5.96 H-B-NPS 79.03±2.84 22.03±0.64 H-BT-NPS 763.62±5.97 257.46±2.39

[0072] Example 7

[0073] Determination of the contact killing effect of β-caryophyllene liposome on Bemisia tabaci by drug film method

[0074] Method of application: The contact toxicity of two commercial chemical pesticides and β-caryophyllene liposome against B. tabaci was determined. Thiamethoxam (4 g / L) and imidacloprid (4 g / L) were prepared, and the β-caryophyllene liposome (Example 5) was prepared at a concentration of 763.62 ± 5.97 mg / L. Sterile water was used as a control. A 150 mL sprayer was used to spray the drug solution three times in a culture dish to form a uniform drug film. Filter paper was used to wipe the drug solution from the center of the culture dish to form a 1 cm B. tabaci release area. After 30 min, the survival of B. tabaci in the culture dish was observed. B. tabaci was considered dead if it was immobile or could not move normally. The number of B. tabaci in each group was 50, and the experiment was repeated six times.

[0075] In the drug film method test ( Figure 4 ), the mortality of B. tabaci MED cryptic species adults treated with thiamethoxam and imidacloprid for 30 min was 15.67% and 10.00%, respectively, indicating that B. tabaci MED cryptic species adults had developed high resistance. The insecticidal effect of β-caryophyllene liposome preparation B-NPS was 89.67%, and H-BT-NPS could reach 97 ± 3.6%, which was 23.67% higher than the use of 0.125% β-caryophyllene alone ( Figure 1 and 4 ). Compared with thiamethoxam and imidacloprid, β-caryophyllene liposome preparation reduced the amount of drug used and had good contact toxicity against B. tabaci.

[0076] The above only describes the preferred embodiments of the present application, and it should be noted that for those skilled in the art, without departing from the principles of the present application, several improvements and refinements can be made, and these improvements and refinements should also be considered within the scope of protection of the present application.

Claims

1. Application of β-caryophyllene liposomes in contact killing of whiteflies; The β-caryophyllene liposomes are characterized in that... It includes lecithin, cholesterol, and β-caryophyllene in a ratio of 0.05-0.25g: 0.01-0.03g: 50-250μL; The β-caryophyllene liposomes also include stabilizers; the stabilizers include Tween 80 and chitosan; the ratio of β-caryophyllene to Tween 80 is limited to 50-250 μL: 0.1-0.5 mL, and the ratio of β-caryophyllene to chitosan is 50-250 μL: 1-6 mg.

2. The application according to claim 1, characterized in that, The preparation method of the β-caryophyllene liposomes is as follows: β-caryophyllene, lecithin and cholesterol are dissolved in ethanol by ultrasonication, the stabilizer is added and stirred to disperse, the ethanol is removed by rotary evaporation, and an appropriate amount of deionized water is added to dissolve the solute to obtain β-caryophyllene liposomes.

3. The application according to claim 2, characterized in that, The stirring is performed using a magnetic stirrer, with the magnetic stirrer set to 220 rpm for 10-20 minutes.

4. The application according to claim 2, characterized in that, The rotary evaporation temperature is below 40°C.

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