CN101543214B - Application of isoliquiritin - Google Patents
Application of isoliquiritin Download PDFInfo
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- CN101543214B CN101543214B CN200910038353XA CN200910038353A CN101543214B CN 101543214 B CN101543214 B CN 101543214B CN 200910038353X A CN200910038353X A CN 200910038353XA CN 200910038353 A CN200910038353 A CN 200910038353A CN 101543214 B CN101543214 B CN 101543214B
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Abstract
The invention discloses an application of isoliquiritin, particularly the application of isoliquiritin to plant disease prevention. Through indoor biological measurement and pot experiment, the invention provides a better preventing and treating application of isoliquiritin which is active ingredient in liquorice methyl alcohol extract to common plant disease such as Peronophythora Chen, Phytophthora capsici Leonian, Cladosporium LK. ex Fr, Sclerotinia Colletotrichum Corda and the like. The invention provides a new design for liquorice and extract thereof applied to plant disease prevention, provides a new development resource for plant substitution chemical synthesis bactericide, and has important significance for solving the problem that pathogenicbacteria has increasingly remarkable resistance to drug.
Description
Technical field
The invention belongs to the bactericide technical field, be specifically related to the new purposes of active ingredient isoliquiritin in controlling plant diseases in a kind of licorice.
Background technology
In recent years, along with the continuous development of related disciplines such as plant pathology, biology, the research of bactericide has got into a brand-new period in the agricultural chemicals.The appearance of efficient systemic fungicide such as Strobilurn series bactericidal agent makes the control of plant disease obtain breakthrough on the one hand, also causes pathogen that the resistance problem of medicament is become increasingly conspicuous on the other hand.It is good to seek Environmental compatibility, and to the public nuisance-free agricultural chemicals that pathogen has lasting effect, important seemed in today that pollution-free food is much accounted of day by day.
Natural products often shows many biologically actives, and widely people are used for hundreds of years.Wherein, the research and development of plant resource fungicide active ingredient is a research direction that solves the many negative effects of current chemical bactericide.
Plant is the huge natural treasure-house of bioactive compound, and the secondary metabolite of its generation surpasses 400000 kinds, it is reported, in tellurian 350,000 various plants, has 2400 plant species to have the activity of control pest approximately now.For many years; Researcher has carried out the research of insect killing substance to Aglaia Aglaia plant, Lauraceae Lauraceae plant Indian avocado Persea indica, Araeceae Araceae, plant calamus etc. in various plants such as the Meliaceae Meliacea plant both at home and abroad; And plant resource is antibacterial, the research of Fungicidal active substance is less relatively; In fact it is reported that 1389 plant species might be as bactericide, the antitoxin in the plant, flavonoids, fall ill relevant protein, organic acid and phenolic compound etc. all have sterilization and antibacterial activity.The plant resource Fungicidal substance is compound antibacterial active ingredient mostly, and has multiple action site and mechanism of action, thereby plant is considered to best the exploiting natural resources of chemosynthesis bactericide substitute.
Radix Glycyrrhizae is the pulse family glycyrrhiza genus, and its root and rhizome are the most frequently used Chinese medicine, and main product is in the Inner Mongol and Xinjiang Uygur Autonomous Regions, except that as the medicine, also extensively be used in food, brewage with cosmetics industry on, more and more wider application prospect is arranged.In recent years, the activity research of Radix Glycyrrhizae had become the focus of concern.On clinical medicine; This platymiscium is widely used in aspects such as anti-inflammatory, analgesia, antibechic, antiulcer, anti-hepatitis, detoxifcation, treatment endocrine system disease; Promptly have effects such as antitumor action, anti-inflammatory and anti-allergy action, resisting pathogenic microbes effect, immunoregulation effect, can also be applied in the preservation and antisepsis aspect of food as the resisting pathogenic microbes medicine.
In glycyrrhiza genus; People have carried out The Chemical Constituents to Ural Radix Glycyrrhizae, glycyrrhiza inflate bat, glycyrrhiza glabra, yellow Radix Glycyrrhizae, coarse wool Radix Glycyrrhizae etc., like the contrast of enoxolone (Clycyrrhetinic aci) content in the different places of production not of the same race, of the same race, plant different parts, the different tissues structure.In fact, from glycyrrhiza genus, isolated about 170 surplus kind of compound, its main compound is flavonoids and triterpene compound, also has a spot of alkaloid, lignin, coumarin and several amino acids; On the composition kind of research, mainly contain glycyrrhizin (Glycyrrhizin), licorice polysaccharide class (Glycyrrhiza polpsacchride), licoflavone class (Glycyrrhizaflavonoides), studied their character and purposes respectively.
The bactericidal activity of Radix Glycyrrhizae is found in report medically, and does not appear in the newspapers about the bacteriostatic activity of licorice to phytopathogen, especially to specifically being that any active ingredient has bactericidal activity and do not see that technology report is arranged in the Radix Glycyrrhizae.
The research licorice is applied to licorice matter in the control of plant disease the bacteriostatic activity of phytopathogen, to solving pathogen the resistance problem that medicament becomes increasingly conspicuous is had great importance.And specifically study active component in the licorice specific aim and the raising that the bacteriostatic activity of phytopathogen can improve control prevented and treated efficient, have important application prospects, do not see the correlation technique report at present.
Summary of the invention
The objective of the invention is to overcome the deficiency of bactericide technology, the new application of licorice active component isoliquiritin aspect sterilization is provided.
The object of the invention is achieved through following technical scheme:
The application of isoliquiritin (isoliquiritin) in controlling plant diseases is provided.
Said isoliquiritin (isoliquiritin) is for being active component in the Radix Glycyrrhizae methanolic extract.
The preparation method of said isoliquiritin (isoliquiritin) gathers the Radix Glycyrrhizae complete stool, and preferred licorice places 50 ℃ of electric heating constant temperature air dry ovens to dry; Cracker is pulverized, and crosses 40 mesh sieves, and the gained licorice powder is preserved to be equipped with to extract with sealed bag sealing lucifuge place and used.Adopt existing routine techniques that the licorice powder powder is extracted.The licorice powder methanolic extract carries out separation, purification and the structure of active ingredient again and identifies that the affirmation active component is isoliquiritin (isoliquiritin).
Isoliquiritin (isoliquiritin) is mixed with certain density confession reagent liquid, adopts mycelial growth rate method, sporangium germ tube sprouting method, excised leaf method that the frequently seen plants disease of white Phytophthora (Peronophythora Chen), Phytophthora (Phytophthora capsici Leonian), Blastocladia (Cladosporium LK.ex Fr), Sclerotinia (Sclerotinia), colletotrichum genus such as (Colletotrichum Corda) has been carried out Toxicity Determination and carried out the greenhouse pot culture test.Result of the test shows that isoliquiritin (isoliquiritin) has good inhibition effect to supplying the examination pathogen, can develop its new purposes in controlling plant diseases.
As a preferred version, the invention provides said isoliquiritin (isoliquiritin), to be applied to prevent and treat the capsicum Phytophthora capsici with the dry powder concentration of 400 μ g/mL sick.
The invention has the beneficial effects as follows:
The present invention breaks through the restriction that traditional Radix Glycyrrhizae only is applied to medicine and pharmacology, the application of Radix Glycyrrhizae has been proposed new thinking, and confirmed that licorice active component isoliquiritin (isoliquiritin) is used for the technical scheme of controlling plant diseases.For plant instead of chemical synthesizing fungicide provides new exploiting natural resources, the resistance problem that medicament becomes increasingly conspicuous is had great importance to solving pathogen.
Embodiment
Come further explain the present invention below in conjunction with concrete embodiment, but content of the present invention is not limited thereto.
The preparation of embodiment 1 isoliquiritin (isoliquiritin)
Gather licorice, place 50 ℃ of electric heating constant temperature air dry ovens to dry; Cracker is pulverized, and crosses 40 mesh sieves.Gained plant powder is preserved to be equipped with to extract with sealed bag sealing lucifuge place and is used.
The present invention adopts existing conventional ultrasonic extraction to prepare licorice, also can join with existing other routine techniques and extract preparation.The methanolic extract for preparing is obtained isoliquiritin (isoliquiritin) through separation, purification and structure evaluation.
Embodiment 2: isoliquiritin (isoliquiritin) is to the inhibitory action of white Phytophthora (Peronophythora Chen) germ
, and then be mixed with the PDA medium and be with malicious medium as the dissolution with solvents extract with 75% methyl alcohol, measure the inhibitory action of variable concentrations extract the growth of peronophythora litchi silk with the mycelial growth rate method.Concrete operations are:
Adopt the mycelial growth rate method: as solvent isoliquiritin (isoliquiritin) is mixed with certain density confession reagent liquid with 75% methyl alcohol; Accurately drawing the 1mL soup joins in the culture dish of pouring the bacterium of going out after (about 50 ℃) mix in the PDA medium that 49mL melts into (diameter 9cm); Evenly pour in 3 culture dishes for every bottle, be mixed with the band poison medium of desired concn.Sterile water or 75% methyl alcohol to sneak into equal volume are done contrast.Supply examination germ peronophythora litchi bacterium in the culture dish of diameter 9cm, to use about 17mLPDA medium culture; Cultivating after 5 days uses card punch to buy the bacterium cake of cut-off footpath as 0.6cm at the vigorous place of colony edge mycelial growth; Be inoculated in each culture dish central authorities respectively, have one of mycelia to face down.Each culture dish is put a bacterium cake, and three repetitions are done in each processing, cultivate a couple of days in 25 ℃ of incubators after, adopt the right-angled intersection method to measure colony diameter (cm), calculate bacteriostasis rate by following formula, experimental result is seen table 1:
Table 1 isoliquiritin (isoliquiritin) is to the inhibitory action (7 days) of peronophythora litchi bacterium mycelial growth
Annotate: 1) result is the mean value of three repetitions in the table;
2) same columns according to after indicate same letter person and be illustrated on 5% level difference not significantly (DMRT method).
Experiment shows; Isoliquiritin (isoliquiritin) to the inhibiting rate of peronophythora litchi bacterium mycelial growth apparently higher than processing; Inhibiting rate reaches 89.443%, 80.683%, 63.830% when 10mg/mL, 5mg/mL, 1mg/mL dry powder concentration, demonstrates very strong bacteriostasis.Embodiment 3: isoliquiritin (isoliquiritin) is to the inhibitory action of white Phytophthora (Peronophythora Chen) germ
Adopt slide method (Fang Zhongda; 1998) measure: the peronophythora litchi bacterium that will on the PDA medium, cultivate 10 days washes sporangium with sterile water; Add soup and be prepared into the sporangia suspension that contains various concentration liquids, sporangium concentration is adjusted into about 40～60 sporangiums in every visual field (10 * 10 times), gets 50 μ L sporangia suspensions and drops on the slide; Again slide is placed in the 9cm culture dish of putting a filter paper; And on filter paper, drip small amount of aseptic water to preserve moisture, and cover culture dish, place 28 ℃ of following illumination cultivation to observe sporangium after 4 hours and sprout situation.Be calculated as follows the sporangium germination rate of chemicals treatment and the inhibition percentage that medicament is sprouted sporangium.
It is active to the inhibition that lichee frost epidemic disease mould sporangium germ tube is sprouted 1.3,1.0,0.7,0.5,0.3, under the 0.15mg/mL to adopt the slide method to measure isoliquiritin (isoliquiritin).Experimental result shows: handled back 4 hours; Microscopically is observed, along with the rising of concentration of treatment, and the corresponding reduction of sporangium germination rate; Inhibiting rate is followed successively by 97.84%, 86.45%, 67.27%, 52.67%, 30.21%, 13.26%, and concrete experimental result is seen table 2.
Average inhibiting rate value is carried out percentage-probability conversion, is abscissa with probability probability value, and the logarithm value of concentration of treatment is an ordinate, asks for regression curve, obtains virulence regression equation y=3.0474x+5.9922, coefficient R=0.9288, EC
50Be 0.47mg/mL, 95% confidence limit 0.45～0.49.
The inhibition active (4 hours) that table 2 isoliquiritin (isoliquiritin) is sprouted peronophythora litchi bacterium sporangium germ tube
Annotate: 1) germination rate, inhibiting rate result are respectively the mean value that repeats for five times in the table;
2) same columns according to after indicate same letter person and be illustrated on 5% level difference not significantly (DMRT method).
Embodiment 4: isoliquiritin (isoliquiritin) to Phytophthora (Phytoph thora capsiciLeonian) germ greenhouse pot culture test
The preparation of Phytophthora capsici germ sporangium liquid: adopt MSS solwution method (Chang Caitao etc., 1995) to induce the Phytophthora capsici germ to produce a large amount of sporangiums, be about on PSA, cultivate 7 days mycelia piece; Be inoculated in the triangular flask that contains the PS liquid nutrient medium; Cultivation is after 48～60 hours down at 25～27 ℃, and the elimination liquid nutrient medium cleans the refunds triangular flask with 20mL MSS liquid with the mycelia on the filter paper;, get culture bacteria liquid and be used to inoculate the capsicum seedling after 24 hours 25～27 ℃ of following illumination cultivation.
Agent prescription is following:
(1) MSS formula of liquid:
MgSO
4·7H
2O 0.004mol/L
，KNO
3 0.05mol/L
Fe-EDTA 1mL
Distilled water 1000mL
(2) prescription of Fe-EDTA:
EDTA-Na
2 14.86g
FeSO
4·7H
2O 24.9g
Distilled water 1000mL
Pot experiment is carried out in plastic tunnel.Each plastic basin alms bowl dress natural air drying vegetable garden soil 0.5kg, healthy and strong capsicum seedling of the 4 leaf phases of transplanting.3 repetitions of every processing, each repeats 5 basins, every basin 4 strains.Long during seedling to 4～5 true leaves, first spraying liquid, contrast spray running water, and establish the metalaxyl contrast, spray connects the Phytophthora capsici germ sporangia suspension for preparing behind the 2h, and the covered with plastic film 24h that preserves moisture immediately.2,4,7d observes incidence respectively, the record temperature, occurring degree one strain is a unit, by 0～4 grade grade scale investigation:
0 grade: anosis;
1 grade: 1 leaf morbidity;
2 grades: 2 leaf morbidities;
3 grades: 3 leaf morbidities;
4 grades: 4 above leaf morbidities or stem are injured
Experimental result is seen shown in the table 3:
The potted plant control efficiency that table 3 isoliquiritin (isoliquiritin) is sick to Phytophthora capsici
Annotate: 1) the many Mils of 53% thunder are treated to 500 times of liquid of dilution;
2) data are the mean value of 3 repetitions in the table; Indicate same letter person after the data and be illustrated on 5% level difference not significantly (DMRT method).
The indoor pot test result shows that isoliquiritin (isoliquiritin) has the better prevention effect to the Phytophthora capsici disease, and control efficiency improves with the rising of handling concentration.When supplying examination concentration to be 50,100,200,300,400 μ g/mL, handle that preventive effect is followed successively by 65.54%, 72.40%, 86.22%, 85.66%, 96.57 after 2 days; Handle that preventive effect is followed successively by 44.44%, 74.61%, 82.53%, 84.13%, 88.90% after 5 days; Handle that preventive effect is followed successively by 34.38%, 50.00%, 68.75%, 75.00%, 86.46% after 10 days.Result also display process concentration is that the preventive effect of 400 μ g/mL is superior to contrasting many Mils of medicament thunder MnZn.
Embodiment 5: isoliquiritin (isoliquiritin) is to the bacteriostatic activity of 3 kinds of pathogens
Adopt the mycelial growth rate method to measure the inhibition effect of isoliquiritin (isoliquiritin) to 3 kinds of peanut black pointss that belong to Blastocladia (Cladosporium LK.ex Fr), Sclerotinia (Sclerotinia), colletotrichum (Colletotrichum Corda), stripe disease of barley, banana anthracnose fungal pathogens.Under aseptic condition, preparation variable concentrations medicament is poured in the medium (temperature is about 60 ℃) that has melted again, fully shakes up, and pours into while hot in the culture dish of 90cm and processes the pastille culture medium flat plate.Flat board solidifies cooling back and meets the various plant pathogenic fungi bacterium cakes that into size is consistent, growth is consistent (Φ=5.0mm) with transfer needle; Every ware connects 1 bacterium cake; Growth has one of bacterium colony to face down, and clear water blank, the contrast of 1% methyl alcohol are established in 3 repetitions of every processing simultaneously.Place biochemical incubator, cultivated 5 days under 25 ℃ of conditions, measure the size that supplies examination fungus colony diameter with the right-angled intersection method again, calculate bacteriostasis rate, and then calculate chemicals treatment concentration (EC in effective inhibition of different germs by following formula
50) and correlation coefficient (r) value.
Bacteriostatic activity is measured the result and is seen table 4.
Table 4 isoliquiritin is to the bacteriostatic activity of 3 plant species disease funguses
Claims (2)
1. isoliquiritin in the application aspect the controlling plant diseases, is characterized in that said plant disease comprises white Phytophthora, Blastocladia, Sclerotinia and/or colletotrichum plant disease as unique active component.
2. isoliquiritin in the application aspect the controlling plant diseases, is characterized in that said isoliquiritin is applied to prevent and treat the Phytophthora capsici disease with the concentration of 400 μ g/mL as unique active component.
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| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN200910038353XA CN101543214B (en) | 2009-03-31 | 2009-03-31 | Application of isoliquiritin |
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|---|---|---|---|
| CN200910038353XA CN101543214B (en) | 2009-03-31 | 2009-03-31 | Application of isoliquiritin |
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| CN101543214B true CN101543214B (en) | 2012-06-27 |
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| CN110742076B (en) * | 2019-10-24 | 2021-05-14 | 福建省农业科学院植物保护研究所 | Application of liquiritin in preventing and treating crop oomycetes and fungal diseases |
Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2009010567A2 (en) * | 2007-07-18 | 2009-01-22 | Trifolio-M Gmbh | A process for the production of a storage stable fungicidal extract of glycyrrhiza glabra for the control of phytopathogenic fungi and other plant diseases |
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Patent Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2009010567A2 (en) * | 2007-07-18 | 2009-01-22 | Trifolio-M Gmbh | A process for the production of a storage stable fungicidal extract of glycyrrhiza glabra for the control of phytopathogenic fungi and other plant diseases |
Non-Patent Citations (1)
| Title |
|---|
| 薛伟等.抗菌植物的研究新进展.《农药》.2005,第44卷(第6期),241-246. * |
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