Preparation method of bougainvillea spectabilis extract and application of bougainvillea spectabilis extract in plant diseases

Abstract

According to the invention, extracts of three representative varieties of bougainvillea speetabilis, such as pink leopard, red sparrow and polygonum orientale, are adopted to carry out antibacterial test research on anise anthracnose, bitter gourd fusarium wilt, tomato gray mold, apple tree canker and corresponding pathogenic bacteria, such as colletotrichum gloeosporioides, fusarium oxysporum bitter gourd specialized type, botrytis cinerea and black beancurd skin fungus. It is found that when the concentration of the bougainvillea spectabilis extract is 5 mg / mL, the antibacterial effect on colletotrichum gloeosporioides is optimal, and a new thought is provided for prevention and treatment of illicium verum anthracnose.

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 flowers in preventing and controlling plant diseases. Background Art

[0002] Plants are a natural treasure trove of bioactive compounds, producing over 400,000 types of secondary metabolites, primarily including terpenes, alkaloids, flavonoids, phenols, steroids, amino acids, and polysaccharides, all of which possess antimicrobial and insecticidal activity. Plant secondary metabolites or plant extracts are widely used in biomedicine, agriculture, and environmental fields due to their rapid onset of action, ease of degradation, resistance to drug resistance, safety against non-target organisms, and environmental friendliness.

[0003] Colletotrichum gloeosporioides is a common plant pathogen. Its lesions on leaves connect to each other, causing them to wither and fall, which in turn affects photosynthesis. Affected fruits are prone to rot, losing their edible and commercial value. Ulcers form on the stems, secreting toxins and enzymes that damage the plant's vascular system, impairing water and nutrient absorption and transport, and leading to cortical necrosis. It is particularly noteworthy that it weakens the plant's own defenses by disrupting its cell structure and physiological functions, thinning its cell walls and increasing cell membrane permeability, thereby weakening the plant's physical defenses. Furthermore, the pathogen inhibits the expression of defense-related genes and the synthesis of defense substances, reducing the plant's chemical defenses. Ultimately, it severely impacts crop yield and quality, causing farmers to suffer economic losses in the form of reduced yields, decreased quality, and increased control costs. The average annual incidence of Colletotrichum gloeosporioides on fruit trees, including apple trees, pear trees, grapevines, citrus, jujube, persimmon trees and strawberries, can reach about 40%. Among them, Colletotrichum gloeosporioides has become the third largest disease restricting the development of China's strawberry industry after strawberry gray mold and strawberry powdery mildew; the annual incidence of Colletotrichum gloeosporioides on vegetables, including peppers and watermelons, is about 30%-50% and 20%-40% respectively; the annual incidence of Colletotrichum gloeosporioides on flowers, including orchids and roses, can reach 10%-30%; in addition, Colletotrichum gloeosporioides can also harm rubber trees and other forest crops.

[0004] Currently, the primary method for controlling Colletotrichum gloeosporioides is the use of chemical fungicides. Although these agents are effective in suppressing the occurrence of Colletotrichum gloeosporioides, their widespread use can lead to a series of problems, including the development of drug resistance in Colletotrichum gloeosporioides, increased costs, chemical pesticide residues in food, and threats to the environment and human health.

[0005] For these reasons, the present invention aims to control Colletotrichum gloeosporioides through biological methods, reducing the environmental and health impacts of chemical fungicides. The fungicidal components in plant extracts are derived from nature and are more easily degraded than chemical fungicides, making them more environmentally friendly. Therefore, the present invention seeks to develop a plant extract with superior control effects against anthracnose in star anise. Summary of the Invention

[0006] The invention provides a preparation method and application of an extract for preventing and treating Colletotrichum gloeosporioides. The extract has a good inhibitory effect on Colletotrichum gloeosporioides and can prevent and treat anise anthracnose.

[0007] The present invention provides a use of a bougainvillea bract extract for preparing a preparation for inhibiting Colletotrichum gloeosporioides or preventing and treating anthracnose. The bougainvillea bract extract is prepared by the following method:

[0008] a) taking fresh Bougainvillea flower samples and drying 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 three bougainvillea varieties—Pink Panther, Red Cardinal, and Red Saffron—to conduct antibacterial tests against anise anthracnose, bitter melon wilt, tomato gray mold, and apple tree rot, respectively. The corresponding pathogens are Colletotrichum gloeosporioides, Fusarium oxysporum (specific for bitter melon), Botrytis cinerea, and Coriolus nigricans. It was found that the Bougainvillea flower extract at a concentration of 5 mg / mL had the best antibacterial effect against Colletotrichum gloeosporioides, providing a new approach for the prevention and treatment of anise anthracnose. BRIEF DESCRIPTION OF THE DRAWINGS

[0017] Figure 1 The inhibitory effect of Bougainvillea flower extract on Colletotrichum gloeosporioides

[0018] Figure 2 The toxicity curve of the extract of Bougainvillea bougainvillea on the mycelial growth of Colletotrichum gloeosporioides

[0019] Figure 3 The inhibitory effect of Bougainvillea flower extract on black rot fungus

[0020] Figure 4 The toxicity curve of the Bougainvillea flower extract on the mycelial growth of the black rot fungus

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

[0022] Figure 6 The toxicity curve of the extract of Bougainvillea bougainvillea against the mycelial growth of Fusarium oxysporum genus

[0023] Figure 7 The inhibitory effect of Bougainvillea flower extract on Botrytis cinerea Figure 3 The inhibitory effect of Cercidiphyllum extract on different pathogens

[0024] Figure 8 The toxicity curve of Bougainvillea flower extract on the growth inhibition of Botrytis cinerea hyphae

[0025] Figure 9 This is the inhibitory effect of Bougainvillea flower extract on different pathogens

[0026] Figure 10 The inhibitory effect of Bougainvillea bract 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 flower extract

[0029] Step S1: fresh samples of three representative varieties of Bougainvillea, namely, Pink Panther, Red Cardinal, and Red Flower, were taken and quickly dried in an oven (superimposed blast 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 step S2 is mixed with 95% ethanol by volume in a ratio of 1:2, and then ultrasonically extracted at 30° C. for 30 minutes. After filtration, the powder is soaked again, and the above steps are repeated three times. Finally, the filtrate is combined.

[0032] Step S4: Pour the filtrate from S3 into a vacuum distillation concentrator (IKA, Shanghai Yarong Biochemical Instrument Factory) and concentrate it at 45° C. to 100 mg / mL, which is then 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, adding deionized water to 1000 mL; and steam sterilizing at 121° C. and 0.1 MPa for 20 min.

[0036] Step S2 is activated by the test bacteria (Colletotrichum gloeosporioides, black rot crust fungus, Fusarium oxysporum bitter melon special type, Botrytis cinerea, 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.), the melted PDA culture medium is poured into a culture dish near the flame of an alcohol lamp. The inoculation needle and the punch are placed in the outer flame of the alcohol lamp and burned repeatedly. After it cools, the punch is used to punch along the edge of the colony, and then the inoculation needle is used to place the punched bacterial cake in the center of the solidified PDA culture medium. After sealing, it is placed in a constant temperature incubator at 25°C and cultured 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 borer to punch a bacterial cake onto the activated target strain culture medium. Use an inoculating needle to pick a bacterial cake and inoculate it onto the center of the drug-containing plate in step S7. Seal the inoculated drug-containing plate and incubate it upside down in a 25°C incubator for 5-7 days. When the colony in the blank control grows to more than two-thirds of the plate, 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]

[0041] 2.2 Experimental results:

[0042] From the toxicity test of Bougainvillea extract on the tested pathogens in Table 1 below, it can be seen that the inhibitory effect of Bougainvillea extract on the tested pathogens is in the following order: Colletotrichum gloeosporioides > Coriolus nigricans > Botrytis cinerea > Fusarium oxysporum type 2, and the corresponding EC 50 The values ​​are: 1.6170mg / mL, 1.8546mg / mL, 2.7946mg / mL, 3.1700mg / mL. Figures 1-9 (a~f are CK, 5mg / mL, 2.5mg / mL, 1.25mg / mL, 0.625mg / mL, and 0.3125mg / mL, respectively) The figure shows that the bougainvillea flower extract has the best antibacterial effect of 100% on Botrytis cinerea, the causative agent of tomato gray mold, at a concentration of 5mg / mL; the antibacterial effect on the pathogenic fungus Cortex Moraxella nigra, the causative agent of apple tree rot, and Colletotrichum gloeosporioides, the causative agent of anise anthracnose, is second best, reaching 96.67% and 95.14% respectively; the inhibition rate on the pathogenic agent Fusarium oxysporum, the causative agent of bitter melon wilt, is relatively low, reaching 71.73%; comprehensively judging, the bougainvillea flower extract has the best inhibitory effect on Colletotrichum gloeosporioides.

[0043] Table 1 Toxicity test of Bougainvillea flower extract to the tested pathogens

[0044]

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

[0046] The bracts and leaves of Bougainvillea were extracted using the method in Example 1, and the antibacterial experiment was carried out using the method in Example 2.

[0047] Experimental results: From the following table 2, Figure 10 and Figure 11 It can be seen from the results that only the Bougainvillea flower extract of the present invention can achieve excellent inhibitory effect on Colletotrichum gloeosporioides.

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

[0049]

[0050]

Claims

1. A use of a Bougainvillea flower extract for preparing a preparation for inhibiting Colletotrichum gloeosporioides or preventing and treating Colletotrichum octagonalis, characterized in that: The bougainvillea bract extract is prepared by the following method: a) taking fresh Bougainvillea flower samples and drying them; b) crushing the dried material; c) ethanol extraction; 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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