CN111978279A - Method for extracting compound methoxy ouabamectin from xanthium italicum and application thereof - Google Patents

Abstract

The invention discloses a method for extracting a compound methoxy ouabamectin from Xanthium italicum and application thereof, which comprises the steps of crushing fruits of Xanthium italicum in a mature period, soaking and crude extracting with ethanol, degreasing with petroleum ether, extracting with trichloromethane, purifying with a silica gel column, and purifying with a Sephadex LH-20 column to obtain the methoxy ouabamectin. The obtained methoxy-containing anthuridin is used as a herbicide to try 3 tested plants, namely monocotyledon bluegrass and pennisetum and dicotyledon amaranthus retroflexus, and the result shows that: the seedlings of monocotyledon annual bluegrass and pennisetum and dicotyledon amaranthus retroflexus are remarkably inhibited from growing, and the inhibition rate of the root length of the amaranthus retroflexus reaches 97 percent; the root length inhibition rate of pennisetum is 52 percent, and the rate of bluegrass is 43 percent. The methoxy-containing anthurium element is a natural component of plant source, can be rapidly decomposed in nature, and does not pollute the environment. The herbicide can be prepared into different concentrations as herbicide, can be used independently before or during seedling stage or mixed with other pesticides, is widely applied to development of various herbicides, and has potential commercial application value.

Description

Method for extracting compound methoxy ouabamectin from xanthium italicum and application thereof

Technical Field

The invention relates to a preparation method for extracting flavonoids from plants and the technical field of application of the flavonoids to herbicides, in particular to a preparation method for extracting a compound methoxy echinacea essence from harmful plant xanthium italicum and application of the compound methoxy echinacea essence as a herbicide.

Background

With the development of modern agriculture, more than 80 percent of weeds in agriculture, forestry and animal husbandry are favored to be controlled by chemical herbicides, and the advantages of labor saving, time saving, labor saving, low cost, simplicity, convenience and the like bring huge benefits for the economic development of agriculture. However, with the mass production and wide use of chemical herbicides, the chemical herbicides can cause serious harm to the natural environment and are killers of the ecological environment. Meanwhile, various artificially synthesized chemical herbicides are easy to remain in the nature for a long time due to lack of corresponding microorganisms for decomposing the herbicides, so that the yield and the quality of agriculture, forestry, animal husbandry and fishery are influenced, and even the health of human beings is harmed. In addition, some chemical herbicides, although capable of decomposing in soil, are gradually resistant to weeds due to their long-term use in large quantities, so that the efficacy of the herbicide is reduced or even lost, see documents 1 to 3. However, the secondary metabolite extracted from natural plants as herbicide is a new direction for the development of pesticides at home and abroad nowadays because it is safe to the environment, not easy to generate drug resistance and can be rapidly degraded. Under the condition, the research and development of the herbicide with novel action targets and action mechanisms has important application value. The extensive screening of secondary metabolites with herbicidal activity in plants, and the separation, purification and structural modification of the secondary metabolites have become the leading and hot fields of novel herbicide research. Some secondary metabolites have own herbicidal activity comparable to that of chemical herbicides, such as 1, 4-cineole, artemisinine, monolaurol and the like; there are also herbicides of natural origin which have a potency comparable to that of chemically synthesized herbicides and are thus processed and promoted to commercial herbicides such as phytotoxin nonanoic acid extracted from geraniaceae, and pulegone-modified sulcotrione isolated from cajeput, see document 4.

Methoxy shozurin, an English name Axillarin, is a flavonoid compound and is presented as yellow amorphous powder. Methoxy-containing anthurium elements are reported to have activities of resisting tumors, oxidation, inflammation and the like, but the growth inhibition effect on plants is not reported at home and abroad, and the reference is 5-8. The artificial chemical synthesis method has been reported, and is disclosed in reference 9.

The xanthium plants for extracting the methoxy oujudin have wide sources. The Xanthium plant is widely distributed in multiple provinces of China, and two exotic invasive Xanthium italicum (Xanthium italicum) plants and smooth Xanthium sibiricum contain the chemical components. The exogenous invasive xanthium sibiricum has strong ecological adaptability and large biomass, can compete with local crops for nutrients and space, has hooked spines on fruits, is difficult to effectively remove, and causes serious influence and harm to agriculture, forestry, animal husbandry and ecological systems of the invasive land, and is disclosed in documents 10-11. The method for extracting the methoxy oujujuin by using the Italian xanthium as a raw material not only explores a new way for the high-value utilization of the invasive plant, but also better controls the further spreading and diffusion of the invasive plant, and protects the surrounding ecological environment and the agricultural economic development.

Reference documents:

1. zhanglong.2012 chemical herbicide: ecological killer.eco-economy, 11:14-17.

2. Wangchunhua.2017. status quo and strategy of chemical herbicide application in China, Shandong pesticide information 01:19-20.

3. Shixiang, Chen also, Huangrong, 2016, State of the use of chemical herbicides and strategies in China, modern agricultural technology, 19: 133-.

4. Mowenyana, herowu.2005, research progress on plant-derived herbicidal actives, chemical and biological engineering, 08:7-9.

5、Park,E.J.；Kim,Y.；Kim,J.Acylated Flavonol Glycosides from the Flower of Inula britannica.JournaL of Natural Products.2000,63(1):34-36.

6、Forgo,P.；István Zupkó.；Judit MoLnár.Bioactivity-guided isolation of antiproliferative compounds from Centaurea jacea L.Fitoterapia.2012,83(5).

7、Juan,B.M.；Habtemariam,S.；Jackson,C.Antioxidant Principles of Tanacetum vulgare L.Aerial Parts.Natural Product Communications.2009,4(11):1561-1564.

8、MoscateLLi,V.；Hnatyszyn,O.；Acevedo,C.Flavonoids from Artemisia copa with anti-inflammatory activity.Planta Medica.2006,72(01):72-74.

9、Fukui,K.；Nakayama,M.；Horie,T.The syntheses of axillarin and its related compounds.Experientia.1968,24(8):769-770.

10. Wangfeng, Dengshitong, Qujing, and Huishan, 2017, research on chemical components of smooth Xanthium strumarium fruits, research and development of natural products, and (05):69-72.

11. Du pearl, xuwen and bin, Hades and Hades, Wang Shaoshan, Guo Yi Min 2012, Sinkiang Xanthium 3 exotic invasive new plants, Sinkiang agricultural sciences, 49(05) 879-.

Disclosure of Invention

The invention aims to provide a method for extracting a compound methoxy ouabamectin from Xanthium italicum and application thereof, the method comprises the steps of crushing fruits of Xanthium italicum in a mature period, soaking the crushed fruits in ethanol, filtering the soaked fruits, steaming the filtered fruits by using a rotary evaporator until no alcohol smell exists, degreasing petroleum ether, extracting trichloromethane, carrying out chromatography by using a 100-and 200-mesh silica gel column, eluting the chromatographic column, and carrying out Sephadex LH-20 column chromatography and elution on the chromatographic column to obtain the methoxy ouabamectin with the purity of more than 95%. The methoxy-containing senecio floridifolin obtained by the method of the invention is taken as a herbicide, can be degraded in the nature due to being derived from plants, does not cause pollution to the environment, simultaneously utilizes the waste of harmful plants, is suitable for preventing and removing farmland weeds, and is shown by experimental results after taking 3 tested plants, namely monocotyledon poa annua and pennisetum alopecuroides, and dicotyledon amaranthus retroflexus as examples: the seedling growth of monocotyledon annual bluegrass and pennisetum and dicotyledon amaranthus retroflexus is obviously inhibited, and the root length inhibition rate of the seedlings on the retroflexus reaches 97 percent; the root length inhibition rate of pennisetum is 52 percent, and the rate of blue grass is 43 percent. In addition, the roots of the plants were distorted and browned, indicating that they were more severely poisoned. High inhibition rate for seedlings: 93 percent of Amaranthus retroflexus, 40 percent of pennisetum alopecuroides and 15 percent of Poa pratensis. In addition, the seedlings also undergo distortion.

The method for extracting the compound methoxy ouabalin from the Italian xanthium comprises the following steps:

a. collecting Italian xanthium plants in mature period, separating fruits from other parts, air drying, and pulverizing for later use;

b. b, soaking the crushed Italian xanthium fruits in 5 times volume of 95% ethanol at room temperature for 7 days, repeating the operation for 3 times, filtering the soaking solution, removing residues, collecting filtrate, and concentrating and drying the filtrate by using a rotary evaporator to obtain an extract;

c. b, stirring the extract obtained in the step b by using distilled water, adding 2 times of volume of petroleum ether for degreasing, repeating the operation for 3 times, and collecting degreased suspension;

d. c, extracting the degreased suspension obtained in the step c by using 2 times of chloroform, repeating the operation for 3 times, combining the extract liquor, and concentrating by using a rotary evaporator to obtain an extract;

e. d, performing chromatography on the extract obtained in the step d by using a 100-200-mesh silica gel column, eluting by using a chloroform-methanol mixed solution with the volume ratio of 98:2, collecting eluent, concentrating and drying to obtain a yellow oily substance;

f. and e, purifying the yellow oily substance obtained in the step e by a Sephadex LH-20 column, eluting by a chloroform-methanol solution with the volume ratio of 1:1, collecting eluent, concentrating and drying to obtain the compound methoxy-containing echinacea essence with the purity of 95%.

The application of the compound methoxy ouabamectin obtained by the method in preparing herbicides.

The invention relates to a method for extracting a compound methoxy ouabamectin from xanthium italicum and application thereof, wherein the structural formula of the extracted compound methoxy ouabamectin is as follows:

the method for extracting the compound methoxy ouabalin from the Italian cocklebur and the application thereof, disclosed by the invention, can be used for preparing the methoxy ouabalin with different concentrations, and the methoxy ouabalin can be used alone or mixed with other pesticides before or during the seedling stage, so that the method has potential commercial application value. After being dissolved by common 95 percent ethanol or acetone, the herbicide is diluted by 10000-20000 times before use and is sprayed before or after emergence of seedlings according to the concentration of 1000-2000 mu g/ml.

The method for extracting the compound methoxy ouabalin from the Italian xanthium and the application have the beneficial effects that:

1. the methoxy ouabamectin extracted from Xanthium italicum provided by the invention can obviously inhibit the growth of seedlings of 3 tested plants, namely monocotyledon Poa pratensis and pennisetum alopecuroides and dicotyledon Amaranthus retroflexus when the concentration is 1000 mug/ml, and the inhibition rate of the methoxy ouabamectin on the root length of the Amaranthus retroflexus reaches 97%; the root length inhibition rate of pennisetum is 52 percent, and the rate of blue grass is 43 percent. In addition, the roots of the plants were distorted and browned, indicating that they were more severely poisoned. High inhibition rate for seedlings: 93 percent of Amaranthus retroflexus, 40 percent of pennisetum alopecuroides and 15 percent of Poa pratensis. In addition, the seedlings also undergo distortion.

2. The methoxy ouabamectin extracted from the Xanthium italicum provided by the invention is suitable for preventing and killing farmland weeds as a herbicide. Because the inhibition effect of the compound on the amaranthus retroflexus is far higher than that of other common weeds, the methoxy-containing anthrin is more suitable for controlling the amaranthus retroflexus. The methoxy ouabamectin extracted from the Xanthium italicum provided by the invention is a natural component of a plant source, can be quickly degraded in the nature, does not pollute the environment, is easy to be applied in large quantities, and has important potential application value for the sustainable development of agriculture. In addition, the compound can be extracted from the alien invasive harmful plants of Xanthium italicum and Xanthium strumarium, which are luxuriant in field growth. The height of the Italian cocklebur plant can reach more than two meters, the biomass is large, the natural herbicide extracted from the Italian cocklebur plant is beneficial to environmental protection, and the harmful plants are comprehensively utilized, so that the Italian cocklebur plant has two purposes.

Drawings

FIG. 1 is a graph showing the growth regulating effects of methoxy shogaetin on the root length of two monocotyledons, namely bluegrass and pennisetum and two dicotyledons, namely amaranthus retroflexus and alfalfa, wherein the action concentrations are 10, 50, 200 and 1000 mu g/ml respectively.

FIG. 2 is a graph showing the high growth regulating effect of methoxy showy chrysanthemin on two monocotyledon plants, namely bluegrass and pennisetum and two dicotyledon plants, namely Amaranthus retroflexus and alfalfa seedlings, wherein the acting concentrations are 10, 50, 200 and 1000 mu g/ml respectively.

FIG. 3 is a graph of growth inhibition by methoxy shozurine on the root length and seedling height of two monocot plants Poa annua and pennisetum alopecuroides and two dicot plants Amaranthus retroflexus and alfalfa.

Detailed Description

The present invention is further illustrated by the following examples, but is not limited thereto.

All materials, reagents and equipment selected for use in the present invention are well known in the art, and other reagents and equipment well known in the art are suitable for practice.

Example 1

Extracting methoxy shougu extract:

a. collecting Italian xanthium plants in mature period, separating fruits from other parts, air drying, and pulverizing for later use;

b. b, soaking the crushed Italian xanthium fruits in 95% ethanol with the volume 5 times that of the Italian xanthium fruits for 7 days at room temperature, filtering the soaking solution, removing residues, and steaming the filtrate by using a rotary evaporator until no alcohol smell exists to obtain an extract;

c. b, stirring the extract obtained in the step b with distilled water, adding petroleum ether with the same volume for degreasing, repeating for 3 times, and collecting supernate to obtain degreased suspension;

d. c, extracting the degreased suspension obtained in the step c for 3 times by using chloroform with the same volume, combining the extract liquor, and concentrating by using a rotary evaporator to obtain an extract;

e. d, performing chromatography on the extract obtained in the step d by using a 100-200-mesh silica gel column, eluting by using a chloroform-methanol mixed solution with the volume ratio of 98:2 to obtain an eluent, and concentrating the eluent to obtain a yellow oily substance;

f. and e, performing Sephadex LH-20 column chromatography on the yellow oily substance obtained in the step e, and eluting with chloroform and methanol in a volume ratio of 1:1 to obtain the compound methoxy-containing echinacea essence with the purity of 95%.

Example 2

The application of methoxy-echinacea essence in preparing herbicide comprises the following steps:

the methoxy-senecio chrysanthemi element obtained in the example 1 is firstly prepared into 1000 mu g/ml by 0.5 percent of acetone, then sequentially diluting with distilled water to concentration of 10 μ g/ml, 50 μ g/ml or 200 μ g/ml, sequentially adding 5ml of methoxyl Augustin solution with concentration of 10 μ g/ml, 50 μ g/ml, 200 μ g/ml and 1000 μ g/ml into culture dishes with diameter of 9cm, spreading 1 layer of filter paper in each culture dish, adding 20 full herba Medicaginis, radix Euphorbiae Fischerianae, herba Potentillae Anserinae and Amaranthus retroflexus seed in each culture dish, repeating for 3 times, sealing with sealing film, placing in an incubator at 25 deg.C, irradiating for 16 hr, the seedlings were cultured for 5 days in the dark for 8 hours, and then the height and root length of the seedlings in each dish were measured, using distilled water containing 0.5% acetone as a control;

the data statistical method comprises the following steps: processing the root growth and seedling height data of each plant by using software such as SPSS22.0, EXCEL2017 and the like, performing analysis of variance (ANOVA) by using one-factor analysis, and comparing the difference significance among different processing groups by using an LSD method;

as a result: the result of activity determination of the methoxy-containing Helichrysum solution on the growth of 4 receptor plant seedlings of alfalfa, Chinese pennisetum herb, bluegrass and Amaranthus retroflexus seeds shows that the methoxy-containing Helichrysum solution has an inhibiting effect on the growth of the tested plant seedlings, the higher the concentration is, the stronger the inhibiting effect is, under the action of the concentration of 200 mu g/ml, the growth of seedlings of dicotyledon Amaranthus retroflexus, monocotyledon bluegrass and Chinese pennisetum is influenced, and the influence degree is enhanced along with the increase of the concentration; it is well known in the art that the weeds alfalfa, bluegrass, pennisetum and amaranthus retroflexus are all well known in the art as being herbicidal targets that are generally difficult to eradicate with common herbicides; when the concentration is increased to 1000 mug/ml, the seedling heights and the root lengths of the bluegrass, the pennisetum and the amaranthus retroflexus are all significantly different from the control, the inhibition rates of the root lengths are 43 percent, 52 percent and 97 percent of the control in sequence, and the roots of the seedlings are obviously distorted and deformed to be blackish brown, which indicates that the seedlings are severely poisoned; the effect of the methoxy-containing anthurin on the seedling height of the tested plant is similar to the effect on the root length, but the effect strength on the seedling height is slightly weaker than the effect on the root length because the methoxy-containing anthurin directly acts on the root of the seedling, and the inhibition rates on the seedlings of the bluegrass, the pennisetum alopecuroides and the redroot amaranth are 15 percent, 40 percent and 93 percent of the control in sequence; see fig. 2 and 3; as can be seen from the inhibition rate of the methoxy-containing echinacea essence solution on the root length and seedling height of 4 plants, the methoxy-containing echinacea essence solution has the effect on the growth of 4 plant receptors, namely Amaranthus retroflexus, pennisetum setosum, Poa pratensis and alfalfa; because the inhibition effect of the compound on the amaranthus retroflexus is far higher than that of other common weeds, the methoxy-containing anthrin is more suitable for being developed into the amaranthus retroflexus herbicide.

And (4) conclusion:

except that the compound methoxy shojudin obtained by the method does not show the inhibition effect on dicotyledonous plant alfalfa, the compound methoxy shojudin can generate the inhibition effect on the growth of other 3 mono-and dicotyledonous plants pennisetum alopecuroides, bluegrass and amaranthus retroflexus seeds when the concentration is 200 mu g/ml; when the concentration reaches 1000 mug/ml, the root length inhibition rate of the compound preparation on dicotyledonous plant amaranthus retroflexus reaches 97%, the root length inhibition rate on monocotyledonous plant pennisetum is 52%, and the root length inhibition rate on bluegrass is 43%; in addition, the roots of the tested plants are distorted and deformed, and the color is browned, which indicates that the plants are subjected to more serious toxicity; the inhibition rate to the seedling height is as follows: 93% of Amaranthus retroflexus, 40% of pennisetum alopecuroides and 15% of Poa pratensis; the seedlings also have distortion; on the whole, the inhibition effect of the methoxy-containing anthurium androsaceus extract on the weed amaranthus retroflexus is more obvious.

The method for extracting the compound methoxy shoujujulin from the xanthium italicum and the application thereof provided by the invention have the advantages that the obtained methoxy shoujulin is used as a herbicide with different concentrations, and can be applied to intractable weeds such as amaranthus retroflexus, pennisetum alopecuroides and bluegrass, and can also obtain a good weeding effect on other common weeds.

The method for extracting the compound methoxy ouabamectin from the xanthium italicum and the application thereof are provided by the invention, and the extracted methoxy ouabamectin is suitable for being applied to preventing and removing weeds in a farm land; because the influence on the bluegrass and the like is far less than the effect on the amaranthus retroflexus, the methoxy-containing anthurium andraeanum element is more suitable for preventing and killing the amaranthus retroflexus weeds in farmlands. The methoxy shojudin is extracted from an exotic invasive harmful plant Xanthium italicum, the plant grows in the wild and is luxuriant, the height can reach more than two meters, the biomass is high, and the herbicide extracted from the plant is beneficial to environmental protection, comprehensive utilization of the harmful plant and prevention and control of the harmful plant. The methoxy-containing anthurium element is a plant-derived natural component, can be rapidly decomposed in nature, does not pollute the environment, and can be applied to weed control in organic agriculture. Is beneficial to developing sustainable agriculture and simultaneously utilizes the waste of harmful plants.

Claims (2)

1. A method for extracting a compound methoxy ouabamectin from Italian cocklebur is characterized by comprising the following steps:

a. collecting Italian xanthium plants in mature period, separating fruits from other parts, air drying, and pulverizing for later use;

b. b, soaking the crushed Italian xanthium fruits in 5 times volume of 95% ethanol at room temperature for 7 days, repeating the operation for 3 times, filtering the soaking solution, removing residues, collecting filtrate, and concentrating and drying the filtrate by using a rotary evaporator to obtain an extract;

c. b, stirring the extract obtained in the step b by using distilled water, adding 2 times of volume of petroleum ether for degreasing, repeating the operation for 3 times, and collecting degreased suspension;

d. c, extracting the degreased suspension obtained in the step c by using 2 times of chloroform, repeating the operation for 3 times, combining the extract liquor, and concentrating by using a rotary evaporator to obtain an extract;

e. d, performing chromatography on the extract obtained in the step d by using a 100-200-mesh silica gel column, eluting by using a chloroform-methanol mixed solution with the volume ratio of 98:2, collecting eluent, concentrating and drying to obtain a yellow oily substance;

f. and e, purifying the yellow oily substance obtained in the step e by a Sephadex LH-20 column, eluting by a chloroform-methanol solution with the volume ratio of 1:1, collecting eluent, concentrating and drying to obtain the compound methoxy-containing echinacea essence with the purity of 95%.

2. Use of the compound methoxy-echinacea obtained by the process according to claim 1 for the preparation of herbicides.

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