Preparation method of bougainvillea bract extract and its use in treating plant diseases

Abstract

This study used extracts from three representative bougainvillea varieties—Pink Panther, Red Cardinal, and Red Saffron—to conduct antibacterial tests against anise anise anthracnose, bitter melon wilt, tomato gray mold, and apple rot, respectively. The corresponding pathogens were Colletotrichum gloeosporioides, Fusarium oxysporum (specific for bitter melon), Botrytis cinerea, and Coriolus fusca. The results showed that the bougainvillea flower extract at a concentration of 5 mg / mL had the best inhibitory effect against Coriolus fusca, the pathogen of apple rot, reaching 100%. This study provides a new approach for preventing and controlling apple rot and Coriolus fusca.

Description

Technical Field

[0001] The invention belongs to the technical field of natural medicines for preventing and controlling plant diseases, and particularly relates to use of an alcohol extract of bougainvillea bracts in preventing and controlling plant diseases. Background Art

[0002] Botanical pesticides are pesticides that are extracted and processed from plant parts or their metabolites to control pests and diseases. With increasing awareness of environmental protection and agricultural product quality and safety, botanical pesticides, as a green and environmentally friendly type of pesticide, have garnered widespread attention and research. As a green and environmentally friendly pesticide, botanical pesticides offer numerous advantages and hold significant application value and development potential in pest control. For example, a 20% horseradish extract emulsion in water achieves 100% efficacy against Fusarium spp. in soil fungi and exhibits significant efficacy against Pythium, Aspergillus, and Penicillium. At low concentrations, an 80% horseradish extract biofumigant can inhibit the growth of Penicillium, Aspergillus flavus, and Botrytis cinerea, and even kill spores.

[0003] Black rot fungus ( Valsa sordida ) is the primary pathogen causing fruit tree rot. It primarily harms apples, pears, peaches, apricots, plums, and other fruit trees, causing significant damage. After infecting the branches and trunks of fruit trees, the pathogen spreads within the cortex, forming ulcers. Initially, the lesions appear reddish-brown, water-soaked, slightly raised, and with soft tissue. Pressing with fingers produces a yellowish-brown sap with a wine-dregs odor. Later, the lesions gradually shrink and become sunken, with small black dots forming on their surface. These are the pathogen's pycnidia. In humid conditions, the pycnidia release yellow, filamentous conidia, which can be transported by wind, rain, insects, and other factors, leading to further infection. As the disease progresses, the lesions expand, encircling the branches and trunks. This causes the upper branches and leaves to gradually wither and die due to a lack of nutrient and water supply. Severe damage to the trunk or branches of a fruit tree can significantly weaken the entire tree, affecting both yield and quality. Fruit trees chronically infected with rot suffer reduced fruit bearing capacity, resulting in smaller fruit, poorer color, and inferior taste, significantly reducing the commercial value of the fruit. In orchards severely affected by the disease, large numbers of trees die due to the pathogen infecting branches and trunks, causing significant economic losses to fruit growers. In orchards generally affected, the disease rate can reach 10%-30%, resulting in yield losses of approximately 10%-20%. In severely affected orchards, the disease rate can reach as high as 50%-80%, with yield losses reaching 30%-50% or even higher. For example, in apple-producing areas with poor management and widespread disease, rot has led to significant yield losses, with some orchards experiencing yield reductions exceeding 50%.

[0004] Currently, the primary method for controlling black rot fungi is chemical agents. However, the long-term, continuous use of a single chemical fungicide can easily lead to the development of resistance in black rot fungi. For example, after long-term use of benzimidazole fungicides such as carbendazim, black rot fungi in some areas have developed high levels of resistance, significantly reducing the effectiveness of these agents. To address this resistance, it is necessary to continuously switch between fungicides with different mechanisms of action or adopt combination formulations. This results in multiple fungicides remaining in soil and water for extended periods of time, potentially affecting soil microbial communities, harming aquatic life, and negatively impacting the ecological environment. Furthermore, pesticide use can be transmitted through the food chain, posing a potential threat to human health. Furthermore, the excessive use of chemical agents can cause environmental pollution. Regarding some impacts on non-target organisms, chemical control not only kills black rot fungi but can also harm non-target organisms such as beneficial microorganisms and insects in orchards. For example, some pesticides may accidentally kill pollinating insects such as bees, affecting the pollination and fruiting of fruit trees; while fungicides may inhibit the activities of some beneficial microorganisms in the soil, affecting the soil's nutrient cycle and ecological balance.

[0005] Therefore, it is urgent to find a new method for preventing and controlling black rot fungus to reduce the impact of chemical fungicides on the environment and health. Summary of the Invention

[0006] The invention provides a preparation method and application of an extract for preventing and treating black rot fungus on fruit trees, which can achieve a 100% inhibitory effect on the black rot fungus and can prevent and treat apple tree rot.

[0007] The present invention provides a use of a bougainvillea bract extract for preparing a preparation for inhibiting black rot fungus or preventing and treating apple tree rot. The bougainvillea bract extract is prepared by the following method:

[0008] a) Take fresh samples of Bougainvillea bracts and dry them;

[0009] b) crushing the dried material;

[0010] c) ethanol extraction;

[0011] d) concentrating the extract.

[0012] Furthermore, in the above step c) ethanol extraction, the volume ratio of the powder obtained in step b) to ethanol is 1:2.

[0013] Furthermore, in the above step d), the extract is concentrated to 100 mg / mL.

[0014] Furthermore, the above also includes pharmaceutically acceptable excipients.

[0015] Furthermore, the concentration of the Bougainvillea bract extract in the above preparation is 5 mg / mL.

[0016] The present invention uses extracts from the bracts of three bougainvillea varieties—Pink Panther, Red Cardinal, and Water Red—to conduct antibacterial tests against anise anthracnose, bitter melon wilt, tomato gray mold, and apple rot, respectively. The corresponding pathogens are Colletotrichum gloeosporioides, Fusarium oxysporum (specific for bitter melon), Botrytis cinerea, and Coriolus fusca. It was found that the bougainvillea bract extract at a concentration of 5 mg / mL had the best antibacterial effect against Coriolus fusca, the pathogen of apple rot, reaching 100%. This provides a new approach for preventing and controlling apple rot or Coriolus fusca. BRIEF DESCRIPTION OF THE DRAWINGS

[0017] Figure 1 The inhibitory effect of Bougainvillea bract extract on black rot fungus

[0018] Figure 2 The toxicity curve of the extract from Bougainvillea bracts against the mycelial growth of the black rot fungus

[0019] Figure 3 The inhibitory effect of Bougainvillea bract extract on Colletotrichum gloeosporioides

[0020] Figure 4 The toxicity curve of the extract from Bougainvillea bracts against the mycelial growth of Colletotrichum gloeosporioides

[0021] Figure 5 The inhibitory effect of Bougainvillea bract extract on Fusarium oxysporum var.

[0022] Figure 6 The toxicity curve of the extract from Bougainvillea bracts against the mycelial growth of Fusarium oxysporum strain Momordica charantia

[0023] Figure 7 The inhibitory effect of Bougainvillea bract extract on Botrytis cinerea

[0024] Figure 8 The toxicity curve of the extract from Bougainvillea bracts against the mycelial growth of Botrytis cinerea

[0025] Figure 9 The inhibitory effect of Bougainvillea bract extract on different pathogens

[0026] Figure 10 This is the inhibitory effect of Bougainvillea flower extract on different pathogens

[0027] Figure 11 The inhibitory effect of Bougainvillea leaf extract on different pathogens DETAILED DESCRIPTION

[0028] Example 1 Preparation method of Bougainvillea bract extract

[0029] Step S1: take three representative varieties of Bougainvillea, Pink Panther ( Bougainvillea spectabilis 'Pink Panther'), Cardinal (Bougainvillea × spectoglabra 'Tmato Red'), Water Red ( Bougainvillea × buttiana Fresh bracts of 'Miss Manila' were quickly dried in an oven (superimposed forced air drying oven, model DHG-9120L-3) at 28°C;

[0030] Step S2: the material of S1 is crushed in a crusher (Yuzaki Co., Ltd.);

[0031] In step S3, the powder obtained in S2 is mixed with 95% ethanol by volume in a ratio of 1:2, and then ultrasonically extracted at 30°C for 30 min. After filtration, the powder is soaked again, and the above steps are repeated three times. Finally, the filtrate is combined.

[0032] In step S4, the filtrate from S3 was poured into a vacuum distillation concentrator (IKA, Shanghai Yarong Biochemical Instrument Factory) and concentrated at 45° C. to 100 mg / mL, which was used as the technical drug of the bougainvillea flower extract.

[0033] Example 2 Antibacterial effect experiment

[0034] 2.1 Experimental Procedure

[0035] Step S1: Prepare potato dextrose agar (PDA) culture medium by weighing 200.0 g peeled potato pieces, 20.0 g glucose, and 15 g agar, and add deionized water to 1000 mL; steam sterilize at 121°C and 0.1 MPa for 20 min.

[0036] Step S2: Activation of the test bacteria (source: provided by the Key Laboratory of Plant Pathology, College of Plant Protection, Shanxi Agricultural University). In a clean bench (Shanghai Boxun Medical Biological Instrument Co., Ltd.), melted PDA culture medium was poured into a Petri dish near the flame of an alcohol burner. An inoculating needle and a punch were repeatedly burned in the outer flame of the alcohol burner. After cooling, a hole was punched along the edge of the colony using the punch. The resulting bacterial cake was then placed in the center of the solidified PDA culture medium using the inoculating needle. The plate was sealed and incubated in a 25°C constant-temperature incubator for 7 days.

[0037] Step S3: Preparation of drug-containing plates: The original drug obtained in S4 was added to sterilized PDA culture medium at 45°C. The drug and culture medium were mixed to prepare drug-containing plates with concentrations of 5 mg / mL, 2.5 mg / mL, 1.25 mg / mL, 0.625 mg / mL, and 0.3125 mg / mL. A blank control was prepared by adding an equal volume of sterile water to the culture medium. Each treatment was repeated three times.

[0038] In step S4, use a 0.7 cm punch to punch a bacterial cake on the activated target strain culture medium. Use an inoculation needle to pick up the bacterial cake and inoculate it into the center of the drug-containing plate in step S7. Seal the drug-containing plate with bacteria and place it in an incubator at 25°C for 5-7 days. When the colony in the blank control grows to more than two-thirds of the culture dish, measure the colony diameter.

[0039] Step S5: Data processing. When the colonies in the blank control grew to more than two-thirds of the culture dish, the colony diameters were measured using the cross-hatch method and photographed to record the effect of the drug-containing plates on fungal colony morphology. Inhibition rates were calculated using Excel software, and virulence analysis was performed using IBM SPSS Statistics 26.

[0040] Mycelial growth inhibition rate (100%) ×100%

[0041] 2.2 Experimental Results

[0042] From the toxicity test of the bract extract of Bougainvillea against the tested pathogens in Table 1, it can be seen that the inhibitory effect of the bract extract of Bougainvillea against the tested pathogens is in the following order: Cordyceps sinensis > Colletotrichum gloeosporioides > Fusarium oxysporum > Botrytis cinerea, and the corresponding EC 50 The values ​​were: 1.2520 mg / mL, 2.3973 mg / mL, 3.1025 mg / mL, 3.8427 mg / mL; Figures 1-9 (a~f are CK, 5 mg / mL, 2.5 mg / mL, 1.25 mg / mL, 0.625 mg / mL, and 0.3125 mg / mL, respectively) It can be seen that when the concentration of Bougainvillea bract extract is 5 mg / mL, it has the best antibacterial effect on the pathogen of apple tree rot, Cortex Moraxella, reaching 100%; the antibacterial effect on the pathogen of star anise anthracnose, Colletotrichum gloeosporioides, is second, with an inhibition rate of 75.28%; the inhibition rates on the pathogen of bitter melon wilt, Fusarium oxysporum, and the pathogen of tomato gray mold, Botrytis cinerea, are relatively low, reaching 64.65% and 58.38%, respectively.

[0043] Table 1 Toxicity test of Bougainvillea bract extract against the tested pathogens

[0044]

[0045] Example 3 Antibacterial effect experiment of extracts from different parts of Bougainvillea

[0046] The method in Example 1 was used to extract Bougainvillea flowers and Bougainvillea leaves, and the method in Example 2 was used to conduct an antibacterial experiment.

[0047] Experimental results: From the following table 2, Figure 10 and Figure 11 It can be seen from the results that only the Bougainvillea bract extract of the present invention can achieve a 100% inhibitory effect on the black rot fungus.

[0048] Table 2 Inhibitory effects of extracts from three different parts of Bougainvillea on four bacteria

[0049]

Claims

1. A use of a Bougainvillea bract extract for preparing a preparation for inhibiting Valsa sordida, characterized in that: The bougainvillea bract extract is prepared by the following method: Step a) taking fresh samples of Bougainvillea bracts and drying them; Step b) crushing the dried material; Step c) ethanol extraction; Step d) concentrating the extract.

2. The use according to claim 1, characterized in that: In the step c) ethanol extraction, the volume ratio of the powder obtained in step b) to ethanol is 1:

2.

3. The use according to claim 1, characterized in that: In the step d), the extract is specifically concentrated to 100 mg / mL.

4. The use according to any one of claims 1 to 3, characterized in that: The preparation also includes pharmaceutically acceptable excipients.

5. The use according to claim 4, characterized in that: The concentration of the Bougainvillea bract extract in the preparation is 5 mg / mL.

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