Application of Houttuynia cordata extract in the prevention and control of potato cyst nematodes

Abstract

The present application mainly relates to the technical field of pest control, specifically the application of Houttuynia cordata extract in the prevention and control of potato cyst nematodes, wherein the Houttuynia cordata extract is Houttuynia cordata volatile oil, which is extracted from the roots of Houttuynia cordata by steam distillation; the prevention and control of potato cyst nematodes refers to the poisoning of the second-instar larvae of potato cyst nematodes; the specific application method is any one or more of the following: 1) using Houttuynia cordata volatile oil to fumigate soil or crops; 2) applying Houttuynia cordata volatile oil to soil and irrigating crops; the present invention has the characteristics of excellent nematicidal activity, good permeability and diffusivity, easy availability of raw materials, and environmental friendliness.

Description

Technical Field

[0001] The present invention mainly relates to the technical field of pest control, and specifically relates to the application of Houttuynia cordata extract in preventing and controlling potato cyst nematodes. Background Art

[0002] Potato cyst nematodes, which include the potato golden wireworm and the potato white wireworm, are a major quarantine disease of potatoes. After invading the host's root system, the plant's ability to absorb water and nutrients is reduced, resulting in symptoms such as stunting, yellowing, and water and fertilizer shortages in the aboveground parts. This leads to a reduction in potato yields, which can reach 80% in severe areas. Due to their protective and hard cysts, potato cyst nematodes can survive in the soil for decades and have the ability to evade environmental self-healing. Therefore, complete eradication is extremely challenging, and a sustainable and reusable means of control is urgently needed.

[0003] The potato cyst nematode primarily hosts plants in the Solanaceae family, such as potato, tomato, and eggplant. However, over 170 species of Solanaceae are considered potential hosts of the nematode. Currently, highly toxic chemical nematicides remain the primary method for nematode control. However, their long-term and extensive use has led to increasing resistance in nematodes, posing significant risks to the environment and humans. Consequently, biological control techniques are gaining increasing attention and are being used for sustainable plant-parasitic nematode management, such as crop rotation, intercropping, deep tillage, planting disease-resistant varieties, and the application of biogenic nematicides.

[0004] Plant volatile compounds are volatile compounds with low boiling points and strong odors. They can attract insects for pollination, repel pests, or communicate with other organisms. These compounds play a vital role in plant growth, development, and adaptation to the environment. Thousands of plant volatiles have been identified, and some highly volatile compounds can attract animals or insects for pollination or attract natural enemies of herbivores, providing a defensive effect. Some volatile compounds also function as communication agents with other plants or organisms, playing a crucial role in plant defense and plant interactions. Numerous studies have found that volatile compounds released by commercial crops, such as mustard, exhibit significant nematicidal activity against the southern root-knot nematode. Extracts from garlic leaves, hemp, and volatile compounds from mustard and fennel have demonstrated significant inhibitory activity against both root-knot nematodes and soybean cyst nematodes. However, there is little research on the use of cash crops to control potato cyst nematodes. Currently, only Chinese patent publication number CN118160718A discloses the use of 1-octen-3-ol in controlling potato cyst nematodes. Specifically, 1-octen-3-ol is used as a fumigant to fumigate soil or crops to achieve the purpose of controlling potato cyst nematodes. However, 1-octen-3-ol is insoluble in water and can only be dissolved in organic solvents. It is also slightly soluble in ethyl acetate and acetonitrile. Therefore, the extraction of 1-octen-3-ol is costly and prone to organic solvent pollution. Therefore, it is urgent to find a crop that can effectively use plant volatile compounds to control nematodes for potato cyst nematodes. Summary of the Invention

[0005] In view of the deficiencies in the prior art, the present invention proposes the use of Houttuynia cordata extract in preventing and controlling potato cyst nematodes.

[0006] This is achieved specifically through the following technical solutions:

[0007] The invention relates to an application of a houttuynia cordata extract in preventing and controlling potato cyst nematodes. The houttuynia cordata extract is a houttuynia cordata volatile oil, which is obtained by extracting the houttuynia cordata root by using a steam distillation method.

[0008] Furthermore, the Houttuynia cordata volatile oil includes any one or more of decanoic acid, 2-tridecanone, n-nonanol, palmitic acid, n-decyl alcohol, and dodecanoic acid in any ratio.

[0009] Furthermore, the control of potato cyst nematodes refers to poisoning the second-instar larvae of potato cyst nematodes.

[0010] The insecticidal concentration of the Houttuynia cordata volatile oil is 30 µ g / mL-200 µ g / mL.

[0011] Furthermore, the insecticidal concentration LC of the decanoic acid is 50 The value is 46.13 µ g / mL.

[0012] Furthermore, the insecticidal concentration LC of the 2-tridecanone is 50 The value is 45.37 µ g / mL.

[0013] Furthermore, the insecticidal concentration of the n-nonanol is LC 50 The value is 61.63 µ g / mL.

[0014] Furthermore, the insecticidal concentration LC of the palmitic acid is 50 The value is 58.85 µ g / mL.

[0015] Furthermore, the insecticidal concentration LC of the n-hexyl alcohol is 50 The value is 31.52 µ g / mL.

[0016] Furthermore, the insecticidal concentration LC of the lauric acid is 50 The value is 37.03 µ g / mL.

[0017] The application of the Houttuynia cordata extract in preventing and controlling potato cyst nematodes is specifically carried out in any one or more of the following ways: 1) fumigating the soil or crops with Houttuynia cordata volatile oil; 2) applying Houttuynia cordata volatile oil to the soil and irrigating the crops.

[0018] Furthermore, the crop is potato.

[0019] Beneficial effects:

[0020] 1. Excellent nematicidal activity. The in vitro nematicidal activity test on potato cyst nematodes was conducted using Houttuynia cordata volatile oil or six compounds in its volatile oil. The results showed that Houttuynia cordata volatile oil can effectively kill potato cyst nematodes, thus achieving green prevention and control of potato cyst nematode disease. µ The nematicidal activity at the concentration of 100 g / mL was 100%, and the concentration was reduced to 50 µ g / mL, its nematicidal activity can still be greater than 50%. Six volatile compounds of Houttuynia cordata are further selected, namely, n-decanoic acid, 2-tridecanone, n-nonanol, palmitic acid, n-decyl alcohol, and dodecanoic acid (structures shown in Table 7). Their nematicidal activity can still be greater than 50% at 50 µ g / mL concentration is greater than 50%, and its LC 50 The values ​​are 46.13 µ g / mL, 45.37µ g / mL, 61.63 µ g / mL, 58.85 µ g / mL, 31.52 µ g / mL, 37.03 µ g / mL, among which n-decyl alcohol showed the best nematicidal activity.

[0021] 2. Good permeability and diffusion. The six volatile compounds or Houttuynia cordata volatile oils used to control potato cyst nematodes have good permeability and diffusion, and can be used as a fumigant for nematode treatment or for root irrigation of crops to reduce potato cyst nematode disease. Potato cyst nematodes treated with Houttuynia cordata volatile oils at 12.5 µ g / mL concentration, the number of potato root nematodes and the number of cysts in the soil were different from those in the control. µ g / mL, the number of hatched nematodes was significantly different from that of the control. After treatment with the compound n-decanol, the number of nematodes invading the root system, the number of hatched nematodes and the number of cysts were all reduced, and the inhibitory effect was always better than that of volatile oil, and the inhibitory effect was equivalent to that of avermectin.

[0022] 3. The raw materials are easily available. Houttuynia cordata is distributed in Shaanxi, Gansu and various places south of the Yangtze River Basin, and has a wide range of sources. In the present invention, the volatile oil can be obtained by steam distillation. The preparation method is simple, green and environmentally friendly, and no organic solvent is required. The present invention first extracts the volatile oil of Houttuynia cordata and tests whether the volatile oil of Houttuynia cordata has good toxicity to potato cyst nematodes. Then, a series of compounds of Houttuynia cordata are obtained by GC-MS analysis and comparison of the differences in the components and insecticidal activity of Houttuynia cordata in multiple regions. Then, the nematicidal activity of 45 available compounds is determined, and finally 6 volatile compounds of Houttuynia cordata roots are selected to have good nematicidal activity in preventing and controlling potato cyst nematode disease.

[0023] 4. Environmentally friendly. The Houttuynia cordata volatile oil of the present invention is safe to non-target organisms and is not prone to drug resistance. As a natural ingredient that is both medicinal and edible, the volatile oil produced by Houttuynia cordata is environmentally friendly. Its use as a bionematicide has lower ecological risks compared to traditional chemical nematicides, and it does not cause soil acidification, heavy metal residues, and other problems. BRIEF DESCRIPTION OF THE DRAWINGS

[0024] The supplementary drawings can more clearly describe and analyze the specific implementation results.

[0025] Figure 1 This is the total ion chromatogram of Houttuynia cordata essential oil determined by GC-MS at six locations across China; H1-H6 are Kunming, Yunnan, Liangping, Chongqing, Leshan, Sichuan, Yichang, Hubei, Xinhuang, Hunan, and Guiyang, Guizhou, respectively.

[0026] Figure 2 Statistical chart of the number of root-invading nematodes, the number of hatched nematodes, and the number of soil cysts after root irrigation with Houttuynia cordata volatile oil, the compound with the best in vitro nematicidal activity, and avermectin; A: the number of nematodes invading potato roots under different concentrations and three treatments; B: the number of soil nematodes under different concentrations and three treatments; C: the number of soil cysts under different concentrations and three treatments. DETAILED DESCRIPTION

[0027] The specific embodiments of the present invention are further described in detail below, but the present invention is not limited to these embodiments. Any improvement or replacement based on the basic spirit of the present embodiment still falls within the scope of protection required by the claims of the present invention.

[0028] Example 1

[0029] The present invention will be further described in detail below with reference to specific examples. It should be noted that the following examples are only for illustration and are not intended to limit the scope of the present invention in any way.

[0030] The potato variety used in the following experiments is Qingshu No. 9, and the Houttuynia cordata used in the examples is a commercially available wild type of Houttuynia cordata, as follows:

[0031] Example 1: In order to determine whether the volatile oil of Houttuynia cordata has nematicidal activity, Houttuynia cordata from Guiyang, Guizhou Province was selected for extraction and in vitro nematicidal activity test.

[0032] After washing and air-drying, Houttuynia cordata was chopped and distilled in a distillation apparatus for 8 h to collect the volatile oil, which was then dried over anhydrous Na2SO4 and stored in a light-proof glass bottle at 4°C for further analysis. The purified volatile oil was dissolved in 0.3% Tween 20, and then potato cyst nematodes were transferred to the volatile oil containing different concentrations (12.5, 25, 50, 100, and 200 μ g / mL) in 48-well microtiter plates, with Tween 20 as a control. The plates were sealed with parafilm and incubated at 23°C for 24 and 48 hours. After exposure, the nematodes were observed under an inverted microscope and their activity was recorded. Nematodes were considered dead when their bodies straightened or remained motionless after being touched with a needle. The experiment was repeated three times, with three replicates each time. The corrected mortality rate was calculated using the Schneider-Orellis equation:

[0033] Mortality rate (%) = number of dead nematodes in treatment group / number of test nematodes in treatment group × 100;

[0034] Adjusted mortality rate (%) = (mortality rate of treatment group – mortality rate of control group) / (1 – mortality rate of control group) × 100.

[0035] Experimental results

[0036] The results are shown in Table 1. The volatile oil of Houttuynia cordata has a certain nematicidal activity. μ The nematicidal activity at the concentration of 50 μg / mL can reach 100% in 24 hours. μ g / mL, the insecticidal activity can still reach 51.2%.

[0037] Table 1 Nematicidal activity of Houttuynia cordata essential oil at different concentrations a

[0038]

[0039] Note: a Each experiment was repeated three times

[0040] Example 2: Insecticidal activity test of Houttuynia cordata volatile oil and analysis of its active ingredients in six locations across China (Kunming, Yunnan, Liangping, Chongqing, Leshan, Sichuan, Yichang, Hubei, Xinhuang, Hunan, and Guiyang, Guizhou).

[0041] The specific operation method is: Houttuynia cordata collected from 6 places across China, and extracted and tested its nematicidal activity in vitro according to the operation method shown in Example 1. At the same time, the volatile oil of Houttuynia cordata collected from 6 places was analyzed by GC-MS (precision oil extraction 1 µ L thermal desorption injection. Initial temperature: 50°C (hold for 2 min), ramped to 180°C at 5°C / min, then to 310°C at 10°C / min (hold for 20 min), run time: 61 min; vaporizer temperature: 250°C; carrier gas: high-purity He (99.999%); column head pressure: 7.6522 psi, carrier gas flow rate: 1.0 mL / min, split: 20:1, solvent delay time: 6 min. ion source: EI; ion source temperature: 230°C; quadrupole temperature: 150°C; electron energy: 70 eV; emission current: 34.6 μ A; multiplier voltage 1422 V; interface temperature 280°C; mass range 29–500 amu. Volatile chemical components were identified by retrieval of peaks in the total ion current (TIC) using a mass spectrometry computer data system and comparison with the NIST20 standard mass spectrum. The relative mass fractions of each chemical component were determined using peak area normalization.

[0042] Experimental results

[0043] According to the GC-MS test results, Figure 1 As shown in Table 2, the volatile oil of Houttuynia cordata in different regions varies in composition. µThe 24-hour adjusted mortality rate against potato cyst nematodes was greater than 92% at 100 μg / mL. Houttuynia cordata essential oils from Liangping, Chongqing; Leshan, Sichuan; Xinhuang, Hunan; and Guiyang, Guizhou achieved 100% mortality. This indicates that Houttuynia cordata essential oils from different regions exhibited a strong toxic effect against potato cyst nematodes, and that the toxicity of Houttuynia cordata against potato cyst nematodes varied from region to region.

[0044] Table 2 Volatile oil of Houttuynia cordata in different regions µ g / mL nematicidal activity a

[0045]

[0046] By determining the components of Houttuynia cordata volatile oil from different regions, a total of 94 compounds were detected in 6 regions (Table 3). 45 commercially available compounds were selected and determined in 200 µ g / mL concentration, there are 14 compounds with nematicidal activity greater than 50% at this concentration (Table 4). Therefore, these 14 compounds were tested at lower concentrations and found that at 50 µ g / mL concentration, the nematicidal activity of the six compounds was still 50% (Table 5), so the LC 50 values ​​(Table 6), among which the LC values ​​of decanoic acid, 2-tridecanone, nonanol, palmitic acid, hemp alcohol, and dodecanoic acid 50 The values ​​are 46.13 µ g / mL, 45.37 µ g / mL, 61.63 µ g / mL, 58.85 µ g / mL, 31.52 µ g / mL, 37.03 µ g / mL. Table 7 shows the structures of these six compounds.

[0047] Table 3 Chemical components of Houttuynia cordata essential oils from six locations across China determined by GC-MS

[0048]

[0049]

[0050]

[0051] a H1-H6 are Kunming, Yunnan, Liangping, Chongqing, Leshan, Sichuan, Yichang, Hubei, Xinhuang, Hunan, and Guiyang, Guizhou.

[0052] Table 4 45 compounds in 200 μIn vitro nematicidal activity at 100 μg / mL

[0053]

[0054] Table 5 14 compounds at 50 μ In vitro nematicidal activity at 100 μg / mL

[0055]

[0056] Table 6 LC values ​​of 6 compounds against nematodes at 48 h 50 value

[0057]

[0058] Table 7 Names and structures of 6 compounds

[0059]

[0060] Example 3: Root irrigation experiment of Houttuynia cordata volatile oil and compound n-heptyl alcohol

[0061] The specific operation is as follows: potato Qingshu No. 9 with uniform buds was planted in a greenhouse for natural growth, and after the potato seedlings emerged from the soil, potatoes with uniform growth were selected for root irrigation. The purified Houttuynia cordata volatile oil, the obtained n-houttuynia cordata and the positive control drug avermectin were prepared with 0.3% Tween 20 to a concentration of 12.5 µ g / mL, 25 µ g / mL, 50 µ g / mL, 100 µ g / mL, 200 µ g / mL root irrigation solution was applied 3 and 7 days after potato seedlings emerged from the soil, with 200 mL per irrigation. A negative control group was treated with Tween 20. After root irrigation, the number of hatched nematodes in the soil and the number of nematodes invading the potato roots were counted on the 14th day after potato seedlings emerged from the soil. The number of cysts in the soil was counted on the 112th day after potato seedlings emerged from the soil, the final stage of cyst formation.

[0062] Nematode counts were calculated by root staining and soil incubation. Potato pots were turned upside down, and soil was gently loosened. The soil from the potato root system was collected into labeled sample bags. Using a Baermann funnel, 100 g of each soil sample was weighed for nematode collection. The potato roots were cleaned, treated with 5.25% sodium hypochlorite solution, and then rinsed twice with distilled water. The roots were then heat-stained with an acidic dye. The stained roots were observed under a microscope for infection, and the number of nematodes per gram of root was counted.

[0063] Experimental results

[0064] like Figure 2As shown in the figure, the number of potato cyst nematodes invading the root system and hatching in the soil decreased after treatment with each concentration. µ g / mL, the number of potato root nematodes and the number of cysts in the soil were different from those in the control, 25 µ g / mL, the number of hatched nematodes was significantly different from that of the control. After treatment with the compound n-decanol, the number of nematodes invading the root system, the number of hatched nematodes and the number of cysts were all reduced, and the inhibitory effect was always better than that of volatile oil, and the inhibitory effect was equivalent to that of avermectin.

[0065] The embodiments of the present application have been described above. The above description is illustrative and not exhaustive, and is not limited to the disclosed embodiments. Many modifications and variations will be apparent to those skilled in the art without departing from the scope and spirit of the described embodiments. The terminology used herein is selected to best explain the principles of the embodiments, their practical applications, or improvements to the technology in the market, or to enable other persons skilled in the art to understand the embodiments disclosed herein.

Claims

1. The application of Houttuynia cordata extract in preventing and controlling potato cyst nematodes, characterized in that: The houttuynia cordata extract is houttuynia cordata volatile oil, which is extracted from the root of houttuynia cordata by steam distillation; The Houttuynia cordata volatile oil comprises any one or more of decanoic acid, 2-tridecanone, n-nonanol, palmitic acid, n-decyl alcohol, and dodecanoic acid in any ratio; The insecticidal concentration of the Houttuynia cordata volatile oil is 50 µ g / mL-200 µ g / mL; The application method is any one or more of the following: 1) fumigating the soil or crops with Houttuynia cordata volatile oil; 2) applying Houttuynia cordata volatile oil to the soil and irrigating the crops.

2. The use of the Houttuynia cordata extract in preventing and controlling potato cyst nematodes according to claim 1, characterized in that: The insecticidal concentration LC of the decanoic acid 50 The value is 46.13 µ g / mL, the insecticidal concentration LC of the 2-tridecanone 50 The value is 45.37 µ g / mL, the insecticidal concentration of n-nonanol is LC 50 The value is 61.63 µ g / mL, the insecticidal concentration of the palmitic acid LC 50 The value is 58.85 µ g / mL, the insecticidal concentration of the n-hexanediol LC 50 The value is 31.52 µ g / mL, the insecticidal concentration LC of the lauric acid 50 The value is 37.03 µ g / mL.

3. The use of the Houttuynia cordata extract in preventing and controlling potato cyst nematodes according to claim 1, characterized in that: The crop is potato.

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