CN108576064B

CN108576064B - Jackfruit extract and application thereof

Info

Publication number: CN108576064B
Application number: CN201810257424.4A
Authority: CN
Inventors: 牛晓庆; 吕朝军; 宋薇薇
Original assignee: Coconut Research Institute of Chinese Academy of Tropical Agricultural Sciences
Current assignee: Coconut Research Institute of Chinese Academy of Tropical Agricultural Sciences
Priority date: 2018-03-27
Filing date: 2018-03-27
Publication date: 2020-11-03
Anticipated expiration: 2038-03-27
Also published as: CN108576064A

Abstract

The invention discloses application of a jackfruit extract, which is particularly applied to preventing and treating plant diseases. Through indoor bioassay and greenhouse seedling tests, the invention provides the application of the ethanol extract of the jack fruit leaves in preventing and treating coconut diarrhea pathogenic fusca mirabilis paradoxa. The invention provides a new idea for the new application of the jack fruit and the extract thereof in preventing and treating plant diseases, provides a new resource for developing environment-friendly plant source bactericides, and has great significance for solving the resistance problem of pathogenic bacteria to chemical agents and the fruit safety problem.

Description

Jackfruit extract and application thereof

Technical Field

The invention belongs to the field of plant protection, and relates to a jackfruit extract for efficiently preventing and treating coconut bloody diarrhea germs and application thereof.

Background

Coconut Stem diarrhoea (Stem greening Disease of Coconut) caused by sexual generation of Rhinocosoma paradoxa (Ceratophysis paradoxa) of Rhinocosoma mirabilis (Thielavissis paradoxa) is one of the main diseases of Coconut, and the pathogen can cause Coconut Stem rot and Coconut fruit rot. The disease is commonly caused in coconut planting countries and generally invades from wounds, if no prevention measures are taken, diseased plants die within 3-4 months after symptoms appear, if coconut stems are rotten in rainy days, fruits can fall in advance, and serious loss is caused to the coconut industry. Meanwhile, the bacterium is an important pathogenic bacterium of the palmaceae plants, almost all the palmaceae plants can be damaged, and symptoms such as stem rot, bud rot, bloody discharge and the like can be caused. Meanwhile, the fertilizer can also harm important tropical and subtropical economic crops such as bananas, pineapples and sugarcanes, and brings great economic loss to production. The disease is reported to be one of the most devastating diseases in andela, india, with an incidence of up to 20%. The losses of the young palmaceae plantations of liberia due to the damage of the bacteria are as high as 50%. The incidence of pineapple disease caused by harm to pineapples can reach 40 percent at most in pineapple production areas, and the market supply and processing production of pineapples are seriously influenced. As the biggest tropical area in China, Hainan province has large planting area and multiple varieties of palmaceae plants, and is also a main planting area of important tropical and subtropical economic crops such as bananas, pineapples and sugarcanes, once rotting diseases of coconut stems caused by moniliforme are epidemic, damage to the hot-area crops cannot be predicted.

The current method for effectively preventing and treating the coconut diarrhea disease uses a large amount of chemical bactericides. The used medicaments comprise iprodione, chlorothalonil, dimethomorph, thiophanate-methyl, oxadiumzinc, prochloraz manganese salt, carbendazim, mancozeb, tridemorph, eugenol and the like, wherein the eugenol, the carbendazim and the prochloraz manganese salt have strong inhibition effect on C.paradoxa hyphae, and other medicaments have common effects and some medicaments have drug resistance. However, long-term use of these chemical bactericides can easily affect the quality of coconut and other fruits, can easily cause the drug resistance of germs, only increase the dosage of the bactericide to control diseases, but increase the production cost and the pesticide residue, affect the ecological environment and destroy the ecological balance. In fact, people are more and more worried about the influence of chemical residues in the food chain on human health, and the dosage of fruit and vegetable products is required to be reduced. Therefore, it is urgent to find a new method for safely, non-toxic and effectively controlling plant (fruit and vegetable) diseases, and the use of botanical fungicides for controlling plant diseases is an important research direction for solving the negative effects of the current chemical fungicides.

The plant is a treasure house of natural product pesticides, new bactericidal and bacteriostatic active substances are searched from the plant, and a novel botanical fungicide is researched and developed, so that the plant is a hotspot for developing new pesticides in the world. For many years, researchers at home and abroad have conducted researches on insecticidal substances of various plants such as Lauraceae plant Indian avocado Persea indica, Meliaceae plant Agalia plant, Acorus calamus, Araceae plant and the like, but the researches on plant-derived bacteriostatic and bactericidal active substances are relatively few, and actually 1340 plants are reported to be possible to be bactericides, and antitoxin, flavonoid, protein related to morbid diseases, organic acid, phenolic compounds and the like in the plants have bactericidal and antibacterial activities. Plant source bactericidal substances are mostly composite bacteriostatic active ingredients and have various action sites and action mechanisms, so that plants are considered as the best development resources of chemical synthetic bactericide substitutes.

With the social development and the ecological environment requirement, the research and the popularization of the biological pesticide are paid unprecedented attention, especially the botanical pesticide is widely concerned by the society, and the development of the botanical pesticide becomes a research hotspot. China is a big agricultural country, and pesticides play an important role in agricultural production. With the improvement of health consciousness of people, most countries pay great attention to the safety of agricultural products, and the limitation on pesticide residues is very strict. After China adds WTO, the export of agricultural products faces a very severe green barrier, wherein the overproof pesticide residue is a problem which is often encountered, and the competitiveness of agricultural products in China in the international market is seriously influenced. In order to reduce the amount of residual agricultural chemicals, efforts to develop novel agricultural chemicals have become urgent.

The botanical pesticide is derived from nature, can be degraded in the nature, generally cannot pollute the environment and agricultural products, has low possibility of accumulating toxicity in the environment and human body, is relatively safe to people and livestock, has the characteristics of low toxicity and low residue, can keep the high quality of the agricultural products, has low use cost and the like, and has the advantages of being more and more valued and favored by people. In the world, the plant-derived pesticide which is safe, nontoxic, wide in source and low in cost is researched and developed more deeply and more widely at present, and has important economic significance, ecological significance and social significance.

Jack fruit (Latin's name: Artocarpus heterophyllus Lam.) is an evergreen arbor of Moraceae, the genus Artocarpus. The pulp can be eaten fresh or processed into canned food, preserved fruit and fruit juice. The seeds are rich in starch and can be cooked; the sap and the leaves are used for treating swelling and detoxifying; the pulp has effects of quenching thirst, promoting lactation, invigorating middle warmer and invigorating qi. The insecticidal activity of the jackfruit extract is reported, the egg laying repellent rate and the food refusal rate of the diamondback moth are high, but the bactericidal activity of the jackfruit extract is not reported yet, so that the study on the bacteriostatic activity of the jackfruit extract on plant pathogenic bacteria is expected to be applied to the prevention and the treatment of plant diseases in the future, and the jackfruit extract has important significance for solving the increasingly prominent resistance problem of the pathogenic bacteria to medicaments. According to the current situation, the invention initially discusses the research of inhibiting the coconut bloodletting disease by the jack fruit leaf extract, and aims to provide a theoretical basis for preventing and controlling the coconut and other fruit and vegetable diseases caused by pathogenic longrosthorny shell bacteria.

Disclosure of Invention

The technical problem to be solved by the invention is as follows: aiming at the defects of the prior art, the control effect of the jackfruit extract on plant pathogenic bacteria is provided.

In order to solve the technical problems, the technical scheme adopted by the invention is as follows: use of a jackfruit extract in the control of coconut diarrhoea, the jackfruit extract being an ethanol extract of jack fruit leaves

The above-mentioned jack fruit extract is the extract of the leaf of jack fruit, adopt Soxhlet's extraction method to extract, wash the leaf, sun-dry, grind into powder, leach with ethanol, the filtrate is evaporated and concentrated to the paste and get to get final product, specifically:

A. cleaning and air drying the jack fruit leaves, putting the jack fruit leaves into a baking oven at 45-50 ℃ until the leaves are crisp, crushing the leaves into fine powder of 75-95 meshes by a plant high-speed crusher (DWF-100), weighing, sealing and storing.

B. Weighing 50g of dry jackfruit leaf powder, putting the dry jackfruit leaf powder into a Soxhlet extractor, adding 250mL of ethanol, analyzing and purifying, and performing reflux extraction for 24 hours in a water bath at 90 ℃.

C. Vacuum filtering the obtained extractive solution, concentrating under reduced pressure in rotary evaporator (R-210) at 42 deg.C and 247Mpa to obtain paste, transferring into brown jar, weighing, labeling, and storing in refrigerator at 4 deg.C.

The above-mentioned extract of jackfruit can be prepared by adopting existent technology, and said invention only adopts one of its preparation methods to illustrate its feasibility.

The jackfruit extract has an inhibiting effect on the coconut diarrhea germs by inhibiting colony growth, conidium germination and other modes, and meanwhile, the jackfruit extract has a certain control effect on seedlings inoculated with the coconut diarrhea germs and can be further developed into a new botanical fungicide. The extraction method of the extract is convenient and fast, and is easy to operate and popularize; the materials required for separation have wide sources, are easy to purchase, have low cost, do not pollute the environment, and are safe and environment-friendly to people and livestock.

Description of the attached drawings

FIG. 1 is a graph showing the effect of the pineapple honey extract of the present invention on the growth inhibition of hyphae of bloody bacteria of coconut;

wherein: 1 represents an extract of jackfruit; 2 represents the control agent eugenol; and 3 represents blank control sterile water.

FIG. 2 is a graph showing the effect of the pineapple honey extract of the present invention on the control of coconut seedlings inoculated with the coconut diarrhea bacterium;

Detailed Description

The technical solution of the present invention will be further described below with reference to specific tests and embodiments.

The jack fruit leaves used as the raw materials are collected in the grand garden of Wenchang coconut, and the raw materials are sufficiently supplied and are convenient and easy to obtain.

The pathogen, chaconia mirabilis paradoxa, used in the examples was supplied by the group of phytopathology subjects of the coconut institute. Activation of the strain: scraping fusca preserved in potato glucose agar slant culture medium by using inoculating ring, inoculating potato glucose agar plate culture medium, and culturing at 25 deg.C for 48-96 hr.

Example 1

Cleaning jackfruit leaves, airing the surface water, putting the jackfruit leaves into a baking oven at 45-50 ℃ until the leaves are crisp, and crushing the leaves into fine powder of 75-95 meshes by a high-speed plant crusher (DWF-100) to obtain jackfruit powder; weighing 50g of dry powder of the jack-fruit leaves, putting the dry powder into a Soxhlet extractor, adding 250mL of ethanol analytically pure, performing reflux extraction for 24h in a water bath at 90 ℃, performing reduced pressure suction filtration on the obtained extract, putting the extract into a rotary evaporator (R-210), performing reduced pressure concentration at 42 ℃ and 247Mpa to obtain a paste, obtaining a tan jack-fruit extract, transferring the tan jack-fruit extract into a brown wide-mouth bottle, weighing and marking the tan jack-fruit extract, and putting the tan jack-fruit extract into a refrigerator at 4 ℃ for storage for later use.

Example 2 inhibitory Effect of Bromelain extract on Hydnocarpus mirabilis

Weighing the prepared jackfruit extract, filling the jackfruit extract into a triangular flask, preparing the medicaments into solutions of 1.25-50 mg/mL respectively, sucking 2mL of the solutions into culture dishes respectively, adding 18mL of the non-solidified PDA culture medium into the culture dishes containing the medicaments, and uniformly mixing and solidifying the solutions. The pathogenic bacteria dish with diameter of 5mm is placed in the center of the medicated plate, 1% ethanol culture medium is added as control, each treatment is repeated three times, the plate is placed in an incubator at 25-30 ℃ for culture, and after 4 days, the diameter of the bacterial colony is measured according to a cross method. And calculating the relative inhibition rate of the medicament on the growth of hyphae. Solving toxicity regression equation according to least square methodCalculating EC₅₀。

The hypha growth inhibition ratio (%) [ (control colony diameter-treatment colony diameter)/(control colony diameter-5) ] × 100%.

TABLE 1 inhibitory Effect of the extract of jackfruit according to the invention on the growth of colonies of Corallotrichum mirabilis

Note: different lower case letters indicate significant differences (p <0.05, different upper case letters indicate very significant differences (p < 0.01).

Results and analysis: the jackfruit extract has certain inhibition effect on the fusca mirabilis. The results of the indoor bacteriostasis test show that: after 4 days of culture, the inhibition rate of the jackfruit extract on the fusca kirilowii is 85.40 percent and is slightly lower than the inhibition effect of a reference medicament eugenol. The analysis of variance results show that: the difference between the treatment of the jackfruit extract and the treatment of a contrast medicament eugenol is obvious, which indicates that the jackfruit extract can be used for preventing and treating the fusca mirabilis and has the potential of further developing plant-derived bactericides.

Example 3 the effect of jackfruit extract on the germination of conidia of fusca mirabilis was examined using a slide germination method. Firstly, diluting the jackfruit extract with sterile water to be 1.25-50 mg/mL solution, meanwhile, placing the fusca mirabilis cake on a potato glucose agar culture medium plate for culturing for 4 days, and diluting with sterile water to be 1 multiplied by 10 after conidia grow out⁵Taking an equal amount of conidium suspension, mixing the conidium suspension with the diluted solution, dripping the mixture on a glass slide, taking eugenol as a control medicament, setting a blank control, repeating for 3 times, performing moisture preservation culture in a culture dish or a groove glass slide for 10-15 h, observing the conidium germination condition under a microscope, and calculating the spore germination rate, wherein the result is shown in the following table 2.

TABLE 2 Effect of extract of jackfruit on spore germination

Note: different lower case letters indicate significant differences (p <0.05), different upper case letters indicate very significant differences (p <0.01)

Results and analysis: the germination of conidia can be strongly inhibited by both the jackfruit extract and the eugenol, wherein the germination rate of the jackfruit extract treated fusca mirabilis spores is far lower than that of the jackfruit extract which is not treated, but is slightly higher than that of the reference medicament eugenol. The analysis of variance results show that: the difference between the treatment with the jackfruit extract and the treatment with the reference medicament eugenol is significant. Because the conidium can be transmitted by rainwater, anemophily and the like to cause pathogen re-infection, thereby causing disease occurrence and causing great yield reduction, the pineapple honey extract is applied to inhibit the conidium germination, so that the re-infection source can be reduced, and the disease occurrence can be reduced or stopped.

EXAMPLE 4 Effect of Artocarpus mirabilis extract on in vitro leaf control of coconut diarrhea caused by Carcinoma mirabilis

Picking up adjacent cracked leaves on the same coconut tree and the same petiole under the same growth condition, and ensuring that the picked leaves do not have any disease and pest invasion symptoms. Cutting a plurality of leaves with basically consistent sizes, cleaning floating dust on the surfaces of the leaves by using sterile water, then sterilizing the surfaces of the leaves by using 75% alcohol for 10s, repeatedly washing away residual alcohol by using the sterile water, and placing the leaves in a ventilation hood for natural air drying. Making wound with sterilizing needle (each wound is pricked 10 times), respectively taking equal amount of fructus Ananadis Comosi extract, eugenol, and sterile water, uniformly applying to wound of coconut leaf, naturally air drying, and taking 5mm diameter bacteria dish with concentration of 1 × 10⁵20ul of spore solution per mL are evenly smeared on the wound of the leaf, the leaf is placed in a culture dish after the smearing, the moisture preservation and the culture are carried out for 3 days at the temperature of 25-30 ℃, 3 leaves are treated and repeated for 3 times, the diameter of the lesion of the coconut leaf is measured by adopting a cross method, the prevention and treatment effect is calculated, and the result is shown in the following table 3.

Control effect (%) < i [ (control colony diameter-treated colony diameter)/(control colony diameter-5) ] × 100%

Results and analysis: the lesion diameter of treated fusca mirabilis infected with the jackfruit extract of table 3 was significantly smaller than that of control-treated fusca mirabilis, but slightly larger than that of control-agent eugenol. Meanwhile, the diameter of the disease spot is measured by a cross method, and the prevention and treatment effect is calculated and found as follows: the effect of the jackfruit extract on the control of the fusca mirabilis is lower than that of the reference medicament eugenol. The analysis of variance results show that: the difference between the treatment of the jackfruit extract and the treatment of the reference medicament eugenol is obvious. The jackfruit extract can be used for preventing and treating diseases caused by the fusca mirabilis.

TABLE 3 in vitro leaf prevention and treatment effects of the Artocarpus communis extract on coconut diarrhea

EXAMPLE 4 prevention and treatment Effect of Bromelain extract on seedlings of coconut diarrhea caused by Cordyrhynchus mirabilis

Seedlings of 6 months old coconuts were planted in a plastic greenhouse, three coconut heartleaves were selected for each coconut tree, and three treatments were performed for each leaf. Cleaning floating dust on the surface of the leaves with sterile water, sterilizing the surface of the leaves with 75% alcohol for 10s, pricking the surfaces of the leaves with a sterilizing needle, and naturally drying the leaves. Then respectively and uniformly smearing the same amount of jackfruit extract, eugenol and sterile water on the wound of coconut leaf, naturally drying, and after 1h, uniformly smearing 100ul of 1 × 10 concentration on each wound⁵And (3) covering the leaves with a larger transparent fresh-keeping bag, and regularly spraying moisture to the inner surface of the transparent bag. In the experiment, 3 coconuts were treated in each treatment, and three heartleaves were taken from each coconut. Spraying tap water as a control, and setting eugenol as a control. 2. 4 and 6d, observing the disease condition, recording the temperature and the disease spot size of the disease degree as a unit. Finally, the lesion area of 6d is determined as a comparison object, the lesion area is measured by the homogenization of rectangular area, the longitudinal longest length is the length, the transverse widest width is the width, and the unit is in cm²And (6) counting. The results are shown in Table 4

Area of lesion (cm)²) Length of lesion x width of lesion

Control effect (%) < control lesion area-treated lesion area/control lesion area x 100%

TABLE 4 prevention and treatment effects of the pineapple honey extract of the present invention on coconut seedling pieces for treating diarrhea of coconut

Results and analysis: 2. 4 and 6d observation results show that the pineapple honey extract and the eugenol treated leaf scab have almost no expansion phenomenon any more, so that the result of the 6d is taken as a standard for counting the area of the scab. From fig. 2, it can be seen that the diameter of the lesion formed by the infection of the treated fusca mirabilis by the jackfruit extract is significantly smaller than that of the lesion formed by the infection of the control-treated fusca mirabilis, but slightly larger than that of the control medicament eugenol. Meanwhile, the diameter of the disease spot is measured by a cross method, and the prevention and treatment effect is calculated and found as follows: the effect of the jackfruit extract on the control of the fusca mirabilis is lower than that of the reference medicament eugenol. The analysis of variance results show that: the difference between the treatment of the jackfruit extract and the treatment of the reference medicament eugenol is obvious. The jackfruit extract can be used for preventing and treating diseases caused by the fusca mirabilis.

Claims

1. The application of the jackfruit extract in preventing and treating the coconut diarrhea is characterized in that the jackfruit extract is an ethanol extract of jackfruit and is prepared by the following steps:

A. cleaning and air drying the jack fruit leaves, putting the jack fruit leaves into a baking oven at 45-50 ℃ until the leaves are crisp, crushing the jack fruit leaves into fine powder by a plant high-speed crusher DWF-100, weighing, sealing and storing;

B. weighing 50g of dry jackfruit leaf powder, putting the dry jackfruit leaf powder into a Soxhlet extractor, adding 250mL of ethanol, analyzing and extracting for 24 hours in a pure water bath under reflux;

C. vacuum filtering the obtained extractive solution, concentrating under reduced pressure in a rotary evaporator R-210 at 42 deg.C and 247Mpa to obtain paste, transferring into brown jar, weighing, labeling, and storing in a refrigerator at 4 deg.C; the concentration of the jackfruit extract is 25mg/mL paste concentration and is used for preventing and treating coconut diarrhea.

2. Use according to claim 1, characterized in that: the jack fruit leaves are crushed into fine powder of 75-95 meshes.

3. Use according to claim 1, characterized in that: adding ethanol into the dry powder of the jack fruit leaves, and refluxing for 24h under the condition of a water bath at 90 ℃.

4. An application of a jackfruit extract in inhibiting coconut diarrhea pathogenic longcoracoid fungus is characterized in that: the jackfruit extract with a certain concentration inhibits the growth of hyphae and the germination of conidia; the jackfruit extract is an ethanol extract of jackfruit and is prepared by the following steps:

C. vacuum filtering the obtained extractive solution, concentrating under reduced pressure in rotary evaporator R-210 at 42 deg.C and 247Mpa to obtain paste, transferring into brown jar, weighing, labeling, and storing in refrigerator at 4 deg.C.

5. Use of an extract of jackfruit according to claim 4 for the inhibition of the coconut diarrhoea pathogen fusca mirabilis, characterized in that: the concentration of the extract of jackfruit used was 7.5 mg/mL.

6. Use of an extract of jackfruit according to claim 4 for the inhibition of the coconut diarrhoea pathogen fusca mirabilis, characterized in that: in an experiment for observing the inhibition of the germination rate of conidia, a mixed solution of a medicament and a spore liquid is subjected to moisture preservation culture for 10-15 hours in a culture dish or a groove glass slide.

7. Use of an extract of jackfruit according to claim 4 for the inhibition of the coconut diarrhoea pathogen fusca mirabilis, characterized in that: the jackfruit extract has inhibitory effect on the extension of lesion spots of in-vitro diseased leaves, and the result is observed after 3 days.

8. Use of an extract of jackfruit according to claim 4 for the inhibition of the coconut diarrhoea pathogen fusca mirabilis, characterized in that: the pineapple honey extract has an inhibiting effect on the lesion extension of the diseased seedlings of the living coconuts, and the observation and recording time is 6 days.