CN103125502B - Purpose of (-)-sumatrol serving as agricultural fungicide - Google Patents
Purpose of (-)-sumatrol serving as agricultural fungicide Download PDFInfo
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- CN103125502B CN103125502B CN201110396842.XA CN201110396842A CN103125502B CN 103125502 B CN103125502 B CN 103125502B CN 201110396842 A CN201110396842 A CN 201110396842A CN 103125502 B CN103125502 B CN 103125502B
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Abstract
The invention first separates amorpha fruticosa seeds to obtain (-)-sumatrol with agricultural fungicide activity and discloses (-)-sumatrol having strong restrain and disinfection functions on phytopathogen. The (-)-sumatrol can be prepared into agricultural fungicide, widens using ranges of leguminous plant amorpha fruticosa and provides reliable technical base for development of plant pesticide.
Description
Technical field
The invention belongs to pesticides application field, relate to the comprehensive utilization of leguminous plant shrubby flase indigo, be specifically related to the purposes of a kind of active substance (-)-sumatrol that separation and purification obtains from shrubby flase indigo seed as disinfectant use in agriculture.
Background technology
Shrubby flase indigo (Amorpha fruticosa L.) is pulse family (Leguminosae) Amorpha fallen leaves bush.Approximately 25 kinds, the Amorpha plant whole world, originates in North America, and the shrubby flase indigo that China introduces cultivation only has one.China's the South and the North all has the cultivation of shrubby flase indigo, and its applicability is strong, have a very wide distribution, and there is well windproof, the function of fixing the sand, conserve water and soil, northern China particularly desert area be mainly used in water and soil conservation.In addition, cultivation can, for viewing and admiring, be again nectariferous plant, and branch can be for braiding, and leaf can be made green manure.
In the last few years, people started the medical value of shrubby flase indigo to study.Its root, stem, leaf can be done medicinal, and the volatile oil in fruit can be used as desirable natural perfume material, in root, trunk, bark, leaf and the fruit and seed of shrubby flase indigo, all contain the chemical composition that some insect is had to toxic action, and plant resources is very abundant.Zheng Min is handsome at " vetiver and the Primary Study of shrubby flase indigo leaching liquor to legume crop allelopathy " Chinese master's best paper database, has reported in shrubby flase indigo acrial part leaching liquor in 2008 and has contained multiple allelochemical, and the sprouting of seed is had to inhibitory action; Bai Liping is at " shrubby flase indigo fruit pharmacognostical study " Chinese master's best paper database, in 2004, reported the pharmacologically active of shrubby flase indigo extract, its have antitumor, antiviral, phosphodiesterase suppresses, protect the liver, malicious fish, desinsection and insect inhibitory action.WO 9815269A1 has reported the Medical sterilization activity of (-)-sumatrol, can be used for treating the microbial disease of helicobacter pylorus; Dae Sik Jang etc. are at " Potential cancer chemopreventive flavonoids from the stems of Tephrosia toxicaria " Journal of Natural Products, 2003,66 (9), in 1166-1170 page, report that (-)-sumatrol has potential cancer chemoprevention effect.Seiferle etc. are at " Insecticides derived from plants " Journal of Industrial and Engineering Chemistry, 1948,40 (4), in 683-691 page, report the agricultural insecticidal activity of (-)-sumatrol.Ermias Dagne etc. are at " Flavonoids and isoflavonoids from Tephrosia fulvinervis and Tephrosia pentaphylla ", Phytochemistry, 28 (11), the chemical constitution of having reported (-)-sumatrol in 3207-3210 page with and H spectrum ownership, C.T.T.BLATT etc. are at " Cytotoxic Flavonoids from the Stem Bark of Lonchocarpus aff.fluvialis ", Phytotherapy Research, 16, the chemical constitution of having reported (-)-sumatrol in 320-325 page with and C spectrum ownership.But from shrubby flase indigo seed, do not separate so far the report that obtains (-)-sumatrol, also have no (-)-sumatrol has the report of agricultural bactericidal activity.
Summary of the invention
The object of the invention is to from shrubby flase indigo seed to separate the active substance that obtains can be used as disinfectant use in agriculture, and determine its structure, for expand shrubby flase indigo comprehensive utilization, develop new botanical pesticide technical basis be provided.
In order to realize object of the present invention, the present invention to shrubby flase indigo seed (commercially available) carry out ethyl acetate extraction, silicagel column rough segmentation, silicagel column segmentation, high speed adverse current chromatogram separates and high speed adverse current chromatogram secondary separation for the first time, from shrubby flase indigo seed, separate first and obtain having the extract of agricultural bactericidal active substance and proved that its structure is for (-)-sumatrol.Concrete process for separation and purification is as follows:
1. Soxhlet is extracted
First using miniature plant sample cracker is pulverized shrubby flase indigo seed (buying by oneself), be crushed to after 40-80 order, according to shrubby flase indigo seed meal quality (Kg) and solvent volume (L) 1: 4-1: 6 ratio, adopt Soxhlet extraction equipment, under reflux temperature, extract, until the solvent under refluxing from apparatus,Soxhlet's extraction tube is not with color, extraction time is approximately 4-6h.Solvent used is respectively benzinum, ethyl acetate, ethanol, respectively extract is concentrated to the ligroin extraction, ethyl acetate extract and the ethanol extract that obtain shrubby flase indigo seed.Three parts of extracts are carried out respectively to desinsection (killing mite), sterilization and Herbicidal.Insecticidal activity assay selects diamond-back moth, beet armyworm and black peach aphid etc. as supplying examination insect, acaricidal activity selects Tetranychus cinnabarinus as supplying examination mite, bactericidal activity is using Botrytis cinerea, Pyricularia oryzae, tomato late blight bacterium, wheat powdery mildew, cucumber downy mildew, corn rust and cucumber anthracnose etc. as supplying examination germ, and activity of weeding selects barnyard grass and pepper grass as examination material.Life is surveyed result of the test and is shown, the ethyl acetate extract of shrubby flase indigo seed has certain pesticide activity.
2. silica gel column chromatography separates for the first time
The ethyl acetate extract that upper part is had to the shrubby flase indigo seed of certain pesticide activity carries out silicagel column rough segmentation.Taking 100-140 order silica gel as fixing phase, carry out wash-out taking benzinum, carrene, ethyl acetate, acetone, methyl alcohol as mobile phase respectively, the variation according to colour band on pillar, initial gross separation obtains 5 elution fraction.The elution fraction that separation is obtained is carried out respectively biological activity determination (method is with for the first time), the active result of the second secondary survey shows that the eluent ethyl acetate part pesticide activity after silicagel column rough segmentation is best, therefore it is carried out next step silica gel column chromatography secondary separation.
3. silica gel column chromatography secondary separation
Ethyl acetate part that first separation obtains to silicagel column is proceeded silica gel column chromatography secondary separation, eluant, eluent is selected benzinum, benzinum and ethyl acetate (v: v, down together) (10: 1), benzinum and ethyl acetate (5: 1), benzinum and ethyl acetate (1: 1), benzinum and ethyl acetate (1: 2), benzinum and ethyl acetate (1: 5), benzinum and ethyl acetate (1: 10), ethyl acetate, ethanol carry out wash-out, and collect each part of elution fraction.The elution fraction that separation is obtained is carried out respectively biological activity determination (method is with for the first time), biologically active result shows the benzinum after silicagel column secondary separation for the third time: ethyl acetate (1: 1) part elution fraction bactericidal activity is best, and the high speed adverse current chromatogram that therefore it is carried out to next step separates.
4. high speed adverse current chromatogram (HSCCC) separates for the first time
The benzinum that secondary separation is collected to silicagel column: ethyl acetate (1: 1) part carries out high speed adverse current chromatogram and separates for the first time, solvent system selection n-hexane: ethyl acetate: ethanol: water=1: separate at 0.8: 1.3: 1, flow velocity 2ml/min, applied sample amount 300mg, positive rotating speed 850r/min, the fast 600r/min that reverses, 30 DEG C of temperature.Carry out the collection of elution fraction according to the absworption peak of the UV-detector of instrument configuration (254nm wave band), in forward elution process, obtain altogether 11 duplicate samples, in reversion elution process, obtain altogether 4 duplicate samples.The sample obtaining is given birth to respectively test and is tested (method is the same), and result shows that wherein forward the 10th duplicate samples has stronger bactericidal activity.This duplicate samples detects through high performance liquid chromatography, and purity, only in 50% left and right, therefore should further be purified.Next step separates and continues to select high speed adverse current chromatogram to carry out.
5. high speed adverse current chromatogram (HSCCC) secondary separation
High speed adverse current chromatogram is separated to the 10th duplicate samples that forward receives for the first time to carry out high speed adverse current chromatogram and again separates, solvent system selection n-hexane: ethyl acetate: ethanol: water=(1: 2: 1: 1) separate, flow velocity 2ml/min, applied sample amount 300mg, positive rotating speed 850r/min, the fast 600r/min that reverses, 30 DEG C of temperature.Collect according to the absworption peak of the UV-detector of instrument configuration (254nm wave band), in forward elution process, obtain altogether 3 duplicate samples, in reversion elution process, obtain altogether 1 duplicate samples.Detect through high performance liquid chromatography, wherein forward the 3rd duplicate samples purity, more than 95%, has good bactericidal activity.Above-mentioned separator detects and proves that this duplicate samples composition is (-)-sumatrol through nuclear-magnetism, mass spectrum etc.
Adopt said method, from shrubby flase indigo seed, extracting and separating (-)-sumatrol outward appearance obtaining is light yellow solid, and structure is suc as formula shown in (I):
(I) (-)-sumatrol
ESI-MS (-) m/z 409[M-H]
-, ESI-MS (+) m/z 411[M+H]
+, relative molecular mass is 410.
1h NMR (300MHz, CDCl
3): δ 12.44 (1H, s, 11-OH), 6.86 (1H, s, H-1), 6.46 (1H, s, H-4), 6.01 (1H, s, H-10), 5.20 (1H, t, H-2 '), 5.06 and 4.93 (2H, s, 4 '-CH
2), 4.88 (1H, t, H-6a), 4.59 (1H, dd, H-6), 4.16 (1H, d, H-6), 3.84 (1H, d, H-12a), 3.79 and 3.82 (each 3H, s, 2,3-OMe), 3.23 (1H, q, H-1 '), 2.85 (1H, q, H-1 '), 1.75 (3H, s, 5 '-CH
3).H spectrum data and " Flavonoids and isoflavonoids from Tephrosia fulvinervis and Tephrosia pentaphylla ", Phytochemistry, 28 (11), the H spectrum data consistent of (-)-sumatrol of reporting in 3207-3210 page.
13c NMR (75MHz, CDCl
3): 110.3 (C-1), 104.6 (C-1a), 142.9 (C-2), 149.7 (C-3), 101.0 (C-4), 147.2 (C-4a), 66.0 (C-6), 71.7 (C-6a), 156.3 (C-7a), 104.1 (C-8), 166.1 (C-9), 91.9 (C-10), 169.1 (C-11), 101.2 (C-11a), 193.6 (C-12), 43.7 (C-12a), 30.5 (C-1 '), 88.1 (C-2 '), 144.0 (C-3 '), 112.7 (C-4 '), 17.0 (C-5 '), 55.9 and 56.3 (2, 3-OMe).C spectrum data and C.T.T.BLATT etc. are at " Cytotoxic Flavonoids from the Stem Bark of Lonchocarpus aff.fluvialis ", Phytotherapy Research, the C spectrum data consistent of (-)-sumatrol of reporting in 16,320-325 page.
Detect and prove through biologically active, the present invention extracts (-)-sumatrol of obtaining first from shrubby flase indigo seed to be had and suppresses and killing action phytopathogen, should serve as the active component of the disinfectant use in agriculture of a kind of efficient, low toxicity, environmental protection.Therefore, the present invention has found the purposes of (-)-sumatrol as disinfectant use in agriculture first, the comprehensive utilization of having expanded leguminous plant shrubby flase indigo, has not only realized object of the present invention, and provides reliable technical basis for the exploitation of botanical pesticide.
Embodiment
Following examples are used for further describing the present invention.(-) that extract from shrubby flase indigo seed according to the method described above-sumatrol is carried out to agricultural bactericidal active testing.
The mensuration of embodiment 1 (-)-sumatrol bactericidal activity
Use (-)-sumatrol sample extracting from shrubby flase indigo seed to carry out the test of live body prolection to the fungal disease of plant.Control cucumber downy mildew protection effect test method is as follows:
Select the consistent potted plant cucumber seedling of growth, cut off growing point and retain two true leaves, as test material.Carry out foliar spray processing according to design concentration with extract (-)-sumatrol, separately establish the blank of spray clear water, repeat for 3 times.Process latter second day inoculation cucumber downy mildew sporangia suspension, then be positioned over phytotron (temperature: 25 DEG C of daytimes, 20 DEG C of nights, relative moisture: 95-100%) moisturizing cultivation, after 24 hours, be positioned over greenhouse (25+2 DEG C) normal management, 5 days " Invest, Then Investigate " control efficiency.Disease scale, with reference to National Standard of the People's Republic of China " pesticide field efficacy medicine test criterion ", calculates control efficiency with disease index.
Result of the test shows that (-)-sumatrol has stronger bactericidal activity, while being 250mg/L, the preventive effect of cucumber downy mildew has been reached to 98% as (-)-sumatrol concentration.
Claims (4)
1. (-)-sumatrol is as the purposes of disinfectant use in agriculture.
2. purposes according to claim 1, is characterized in that: described (-)-sumatrol separates and obtains from shrubby flase indigo seed.
3. purposes according to claim 2, described (-)-sumatrol adopts following methods separating-purifying from shrubby flase indigo seed to obtain: 1) Soxhlet is extracted, solvent used is ethyl acetate, 2) silica gel column chromatography separates for the first time, collect eluent ethyl acetate component, 3) silica gel column chromatography secondary separation, collect the elution fraction that benzinum and ethyl acetate are 1:1, 4) high speed adverse current chromatogram separates for the first time, dicyandiamide solution is n-hexane: ethyl acetate: ethanol: water=1:0.8:1.3:1, 5) high speed adverse current chromatogram secondary separation, solvent system selection n-hexane: ethyl acetate: ethanol: water=1:2:1:1, it is characterized in that: the absworption peak that UV-detector is 254nm wave band at wavelength is collected, high speed adverse current chromatogram is separated to the 10th duplicate samples that forward receives for the first time and carry out high speed adverse current chromatogram and again separate, the 3rd duplicate samples obtaining in forward elution process is (-)-sumatrol.
4. purposes according to claim 3, is characterized in that: the chemical constitution that the process for separation and purification described in adopting separates (-)-sumatrol obtaining from shrubby flase indigo seed is as follows:
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| CN103125502B true CN103125502B (en) | 2014-06-18 |
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| CN110742079B (en) * | 2019-10-29 | 2021-10-08 | 四川国光农化股份有限公司 | Pesticide composition for promoting root growth and increasing yield of edible leguminous crops, preparation and application |
| CN111903718A (en) * | 2020-07-16 | 2020-11-10 | 海南欧兰德农业高科发展有限公司 | Low-pollution pesticide for green soybean planting |
| CN113200995A (en) * | 2021-04-27 | 2021-08-03 | 河北大学 | Compound Sterigmatocystin, preparation method thereof and application thereof in pesticide |
Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO1998015269A1 (en) * | 1996-10-08 | 1998-04-16 | Mitsubishi Chemical Corporation | Drugs containing rotenoids |
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Patent Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO1998015269A1 (en) * | 1996-10-08 | 1998-04-16 | Mitsubishi Chemical Corporation | Drugs containing rotenoids |
Non-Patent Citations (3)
| Title |
|---|
| Dae Sik Jang,et al.Potential Cancer Chemopreventive Flavonoids from the stems of tephrosia toxicaria.《Journal of Natural Products》.2003,第66卷(第9期),第1166-1170页. |
| Potential Cancer Chemopreventive Flavonoids from the stems of tephrosia toxicaria;Dae Sik Jang,et al;《Journal of Natural Products》;20030820;第66卷(第9期);第1166-1170页 * |
| 纪明山,等.紫穗槐种子杀蚜活性初探.《江苏农业科学》.2011,第39卷(第2期),第208-210页. * |
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