Extract of cleomea breviscapa with insect antifeedant activity, preparation method and application thereof

Abstract

The present invention discloses a short-rib feather moss extract with insect antifeedant activity, as well as its preparation method and application. The method comprises the following steps: subjecting fresh material to reflux extraction with ethanol several times, combining the extracts; rotary evaporating the extract to an extract paste, dissolving the extract paste, and performing liquid separation extraction, wherein the extraction solvent is petroleum ether, ethyl acetate, n-butanol, or water. After extraction, a short-rib feather moss extract is obtained, and evaporation is performed to obtain a short-rib feather moss extract, which is a short-rib feather moss extract with insect antifeedant activity. The present invention discloses for the first time a preparation method for a short-rib feather moss extract with insect antifeedant activity and the application of the compound in insect antifeedant activity; the present invention separates and extracts the compound from plants, is low-toxic, residue-free, and pollution-free to economic crops such as fruits and vegetables, is more conducive to environmental protection, and can achieve the effect of preventing insect pests; the short-rib feather moss extract has antifeedant activity, a more significant antifeedant effect, and has good application prospects.

Description

Technical Field

[0001] The invention belongs to the technical field of pest repellents, and particularly relates to a cleometrium breviscapum extract having insect repellent activity, a preparation method thereof, and application of the extract in preparing the pest repellent. Background Art

[0002] The information disclosed in this background technology section is only intended to enhance some understanding of the overall background of the invention and should not be necessarily regarded as an admission or any form of suggestion that the information constitutes the prior art already known to those skilled in the art.

[0003] The diamondback moth larvae are a worldwide pest of cruciferous vegetables, particularly cabbage, kale, cauliflower, and rapeseed. Their rapid reproduction and strong pesticide resistance make them difficult to control, severely impacting vegetable yield and quality. Currently, organic pesticides dominate pest control, but their harmful effects on humans and animals are becoming increasingly prominent, necessitating the development of natural insecticides. Developing insect repellent compounds from plants, thereby creating green pesticides, is a key approach.

[0004] Insect antifeedant activity refers to the numerous secondary metabolites with defensive functions produced by plants during their coordinated evolution with insects. my country has abundant resources of the family Pleurotus, but there are currently no reports on the application of Pleurotus brevis extracts for insect antifeedant activity. Summary of the Invention

[0005] In view of the shortcomings of the prior art, the present invention provides a short-costed lichen extract with insect antifeedant activity, and its preparation method and application. The extract was used to study insect antifeedant activity and found to have antifeedant activity against Plutella xylostella larvae.

[0006] The technical solution adopted in the present invention is as follows:

[0007] In a first aspect of the present invention, a method for preparing a Psoralea corylifolia extract having insect antifeedant activity is provided, the method comprising the following steps:

[0008] The fresh material - short-rib feather moss is subjected to reflux extraction with ethanol for several times, and the extracts are combined; the extracts are rotary evaporated to form an extract paste, the extract paste is dissolved, and separation extraction is performed. The extraction solvent is petroleum ether, ethyl acetate, n-butanol or water. After extraction, a short-rib feather moss extract is obtained, and the short-rib feather moss extract is evaporated to obtain a short-rib feather moss extract, which is a short-rib feather moss extract with insect antifeedant activity.

[0009] In one or more preferred embodiments of the present invention, the extraction solvent is petroleum ether. In leaf disc assays for insect antifeedant activity, high concentrations of petroleum ether exhibited the most pronounced inhibitory effect on insect feeding. This indicates that high-concentration petroleum ether extracts from P. brevis contain highly potent ecological activity, providing a theoretical basis for the development of green pesticides.

[0010] In one or some preferred embodiments of the present invention, the extract paste is dissolved in water, and the ratio of the extract paste to water is (0.1~1) g:150 mL.

[0011] In one or some preferred embodiments of the present invention, the volume ratio of the dissolved liquid to the extraction solvent is 1:(1-1.5). Preferably, the volume ratio of the dissolved liquid to the extraction solvent is 1:1.

[0012] In one or some preferred embodiments of the present invention, the number of separation extractions is 2 to 3 times. Preferably, the number of separation extractions is 3 times.

[0013] In a second aspect of the present invention, a Pseudostellaria breviscapina extract prepared by the above method is provided.

[0014] In a third aspect of the present invention, there is provided use of the extract in preparing a pest repellent.

[0015] In one or some embodiments of the present invention, the pest repellent has insect antifeedant activity.

[0016] In one or some embodiments of the present invention, the pests repelled by the pest repellent are larvae of Plutella xylostella.

[0017] In a fourth aspect of the present invention, a pest repellent is provided, which is prepared by the following method: dissolving the Pseudostellaria brevis extract in methanol at a concentration of 1 to 100 mg / mL to prepare the pest repellent.

[0018] Compared with the related art known to the inventors, one of the technical solutions of the present invention has the following beneficial effects:

[0019] The present invention discloses for the first time a preparation method of a short-rib feather moss extract having insect antifeedant activity and the application of the compound in insect antifeedant activity; the present invention separates and extracts the extract from plants, has low toxicity, no residue, and is pollution-free to economic crops such as fruits and vegetables, is more conducive to environmental protection, and can achieve the effect of preventing insect pests; the short-rib feather moss extract has antifeedant activity, has a significant antifeedant effect, and has good application prospects. BRIEF DESCRIPTION OF THE DRAWINGS

[0020] The accompanying drawings, which constitute a part of the specification of the present invention, are used to provide a further understanding of the present invention. The exemplary embodiments of the present invention and their descriptions are used to explain the present invention and do not constitute improper limitations on the present invention.

[0021] Figure 1 : Leaf disc area after insect feeding in petroleum ether phase for 24 h.

[0022] Figure 2 : Leaf disc area after insect feeding in petroleum ether phase for 48 h.

[0023] Figure 3 : Histogram of feeding area of ​​petroleum ether leaf butterfly.

[0024] Figure 4 : Leaf disc area after insect feeding in ethyl acetate phase for 24 h.

[0025] Figure 5 : Leaf disc area after insect feeding in ethyl acetate phase for 48h.

[0026] Figure 6 : Histogram of feeding area of ​​Ethylactylophora acetophylla.

[0027] Figure 7 : Leaf disc area after 24h n-butanol feeding.

[0028] Figure 8 : Leaf disc area after insect feeding in the n-butanol phase for 48 h.

[0029] Figure 9 : Histogram of feeding area of ​​n-butanol phase leaf butterfly.

[0030] Figure 10 : Leaf disc area after 24 h of water-phase insect feeding.

[0031] Figure 11 : Leaf disc area after 48h of feeding by water-phase insects.

[0032] Figure 12 : Histogram of feeding area of ​​water leaf butterfly. DETAILED DESCRIPTION

[0033] It should be noted that the following detailed descriptions are exemplary and intended to provide further explanation of the present invention. Unless otherwise specified, all technical and scientific terms used herein have the same meaning as commonly understood by those skilled in the art to which the present invention belongs.

[0034] It should be noted that the terms used herein are only for describing specific embodiments and are not intended to limit the exemplary embodiments according to the present invention. As used herein, unless the context clearly indicates otherwise, the singular form is intended to include the plural form. In addition, it should be understood that when the terms "comprise" and / or "include" are used in this specification, they indicate the presence of features, steps, operations and / or combinations thereof.

[0035] In order to enable those skilled in the art to more clearly understand the technical solution of the present invention, the technical solution of the present invention will be described in detail below with reference to specific embodiments.

[0036] Example 1

[0037] Take 40 g of brevipes serrata, grind it, and reflux it with ethanol 4-5 times in a heating reflux apparatus to obtain a brevipes serrata extract. After filtration, the extract is rotary evaporated to a paste using a rotary evaporator to obtain an extract (450 mg) with an extract yield of 1.125%. Dissolve the extract in 150 ml of purified water. Because brevipes serrata is poorly water-soluble, an ultrasonic cleaner can be used to aid dissolution. Once the extract is completely dissolved, prepare for separation and extraction.

[0038] Separative extraction method: Extraction in a 1:1 equal volume ratio. First, place 150ml of petroleum ether and 150ml of purified water dissolved with the extract of the wormwood wormwood into a 500ml separatory funnel. After tightening the stopper, invert the funnel five times to thoroughly mix the water and petroleum ether. Place the funnel on a metal stand and let it stand. Once the petroleum ether and water phases are clearly separated and clear, drain the aqueous phase from the bottom of the funnel and pour the petroleum ether phase from the top. Take another 150ml of petroleum ether and place it together with the aqueous phase into a 500ml separatory funnel. Repeat this process twice to obtain the petroleum ether phase. Take 150ml of ethyl acetate and extract it with the aqueous phase after extraction with petroleum ether. Follow the same steps as for the petroleum ether phase, extracting it three times to obtain the ethyl acetate phase. Take 150ml of n-butanol and extract it with the aqueous phase after extraction with ethyl acetate. Follow the same steps as for the petroleum ether phase, extracting it three times to obtain the n-butanol phase and the aqueous phase.

[0039] The petroleum ether phase, ethyl acetate phase, n-butanol phase and aqueous phase obtained after extraction were respectively evaporated into extracts using a rotary evaporator, as shown in Table 1.

[0040] Table 1 Extract quality

[0041]

[0042] The four-phase extract obtained by extraction and rotary evaporation was dissolved in methanol and transferred to a penicillin bottle, and then the methanol was blown dry with a hair dryer to obtain the sample powder of the four phases of short-ribbed feather moss, which was sealed and stored in a refrigerator at 4°C for use.

[0043] Example 2

[0044] Antifeedant activity of the extract: Through comparative research, it was found that the feeding rate of Plutella xylostella larvae on rape leaves was the highest. A hole punch with a diameter of 1.2 cm was used to punch holes in rape leaves, and a total of 390 pieces were punched. Every 30 pieces were placed in a round Petri dish, marked, and waited for treatment.

[0045] The leaf disc method is generally used in the study of insect antifeedant activity. The specific method generally adopted is to put the leaf discs treated with the agent and the blank control into Petri dishes respectively. After a period of time, the feeding area of the insects is determined by measuring the area of the leaf discs, so as to measure their selective antifeedant rate or non-selective antifeedant rate.

[0046] Dissolve 1 ml of methanol in the four-phase medicine of Brachythecium salebrosum, and divide it into three different concentrations: high, medium, and low. The specific concentration data are shown in Table 2:

[0047] Table 2 Concentrations of the medicine added to the insect antifeedant leaves

[0048] <00 extraordinariest00116>Add the medicine to the round Petri dish containing the leaves, and use a small brush to make all 30 round leaves in the Petri dish (10 leaves in each group of experiments, with three parallel experiments) soaked with the medicine, and then dry them. After the methanol on the leaves has dried, put them into a plastic Petri dish lined with absorbent filter paper. Put ten leaves in each Petri dish, and put one third-instar Plutella xylostella larva on each leaf. Drill holes in the cover of the Petri dish to ensure that the hole diameter is not large enough for the larva to pass through, but also ensure that the larva is not in a completely enclosed space. Put the Petri dish containing the larva into an environment with moist air and conduct dark treatment. Measure the leaf area after 24 h and 48 h.

[0050] Experimental results:

[0051] Figures 1 to 12 These are the data results of the petroleum ether phase, ethyl acetate phase, n-butanol phase, and water phase, which are respectively the leaf disc diagram after 24 h of insect feeding, the leaf disc diagram after 48 h of insect feeding, the column chart of the average insect feeding area, and the insect antifeedant rate.

[0052] Among them, in the significance difference test, P(F<=f)>0.05 indicates that the difference is not significant; 0.01<P(F<=f)<0.05 indicates that the difference is significant; P(F<=f)<0.01 indicates that the difference is extremely significant. There are also F values and F crit values. If F>F crit, it indicates that the difference is significant. If F<F crit, it indicates that there is no obvious difference between the two groups of data, which are represented by a, b, c, and d.

[0053] Feed refusal rate = (blank group feeding area - treatment group feeding area) / blank group feeding area * 100%, where the total feeding area of ​​ten leaves is taken into the formula for calculation.

[0054] 1. Petroleum ether phase

[0055] Conclusion: Histogram ( Figure 3 ) It can be seen that the medium and high concentrations of petroleum ether phase extracts of Plutella xylostella have obvious anti-insect and antifeedant effects. The feeding area of ​​diamondback moth on leaf discs is significantly lower than that of the blank group ( Figure 1 and Figure 2 ).

[0056] Table 3 Petroleum ether phase rejection rate

[0057]

[0058] 2. Ethyl acetate phase

[0059] Conclusion: Histogram ( Figure 6 ) It can be seen that in the ethyl acetate phase, the medium and high concentrations inhibited the activity of diamondback moth more significantly, while the low concentration had some inhibition, but the inhibitory effect was weak. The feeding area of ​​diamondback moth on the leaf disc was significantly lower than that of the blank group ( Figure 4 and Figure 5 ).

[0060] Table 4 Ethyl acetate phase rejection rate

[0061]

[0062] 3. n-butanol phase

[0063] Conclusion: Histogram ( Figure 9 ) It can be seen that in the n-butanol phase of the short-rib moth extract, the low concentration has a weak inhibitory effect on the feeding of the diamondback moth, but the high concentration has a more obvious inhibitory effect on the feeding of the diamondback moth ( Figure 7 and Figure 8 ).

[0064] Table 5 n-Butanol phase rejection rate

[0065]

[0066] Conclusion: Histogram ( Figure 12 ) It can be seen that the aqueous extract of Plutella xylostella inhibited the feeding activity of Plutella xylostella at various concentrations, among which the low concentration was more obvious, while the high concentration had a weaker anti-insect and anti-feeding effect ( Figure 10 and Figure 11 ).

[0067] 4. Water phase

[0068] Conclusion: The aqueous extract of P. breviscapus inhibited the feeding activity of Plutella xylostella at all concentrations, with low concentration being more obvious and high concentration having weaker anti-insect and anti-feeding effect.

[0069] Table 6 Water phase refusal rate

[0070]

[0071] Among the different extracts obtained from the ethanol treatment, the petroleum ether and n-butanol extracts of P. brevis showed a significant inhibitory effect on feeding by third-instar diamondback moth larvae. The repellency rate also increased significantly with increasing concentration, indicating that the petroleum ether and n-butanol extracts possessed the greatest insect repellent activity. The large variance in the ethyl acetate and aqueous phases resulted in a significant degree of randomness, making the results inaccurate and inconclusive. However, the histograms show that both phases clearly exhibited significant inhibitory and repellent effects on feeding by P. brevis. These results demonstrate that the ethanol extracts of P. brevis do contain active insect repellent compounds, providing a theoretical basis for the development of green insecticides.

[0072] Experimental conclusion: In this example, the extract was studied for insect antifeedant activity, and it was found that Plutella xylostella had antifeedant activity against the larvae of the diamondback moth.

[0073] In summary, the above examples provide the plant source, isolation and extraction method, insect antifeedant activity and application of the extract. Insect antifeedant activity was studied using the extract.

[0074] 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 considered as equivalent replacement methods and are included in the scope of protection of the present invention.

Claims

1. Use of a Psoralea corylifolia extract having insect repellent activity in the preparation of an insect repellent, characterized in that: The preparation method of the short-costal lichen extract comprises the following steps: The fresh material is subjected to reflux extraction with ethanol for several times, and the extracts are combined, wherein the fresh material is a short-ribbed feather moss; the extract is rotary evaporated to obtain an extract paste, the extract paste is dissolved, and the extract is separated and extracted, and the extraction solvent is petroleum ether, ethyl acetate, n-butanol or water to obtain a short-ribbed feather moss extract after extraction, and the short-ribbed feather moss extract is evaporated to obtain a short-ribbed feather moss extract, which is a short-ribbed feather moss extract with insect antifeedant activity; The pests repelled by the pest repellent are larvae of the diamondback moth.

2. The use according to claim 1, characterized in that: The extraction solvent is petroleum ether.

3. The use according to claim 1, characterized in that: The extract paste is dissolved in water, and the ratio of the extract paste to water is (0.1~1) g:150 mL.

4. The use according to claim 1, characterized in that: The volume ratio of the dissolved liquid to the extraction solvent is 1: (1~1.5).

5. The use according to claim 4, characterized in that: The volume ratio of the dissolved liquid to the extraction solvent is 1:

1.

6. The use according to claim 1, characterized in that: The number of separation extractions is 2 to 3 times.

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