CN110452170A - A kind of isoquinoline alkaloids bases compound and its preparation method and application - Google Patents
A kind of isoquinoline alkaloids bases compound and its preparation method and application Download PDFInfo
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- CN110452170A CN110452170A CN201910810833.7A CN201910810833A CN110452170A CN 110452170 A CN110452170 A CN 110452170A CN 201910810833 A CN201910810833 A CN 201910810833A CN 110452170 A CN110452170 A CN 110452170A
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- A—HUMAN NECESSITIES
- A01—AGRICULTURE; FORESTRY; ANIMAL HUSBANDRY; HUNTING; TRAPPING; FISHING
- A01N—PRESERVATION OF BODIES OF HUMANS OR ANIMALS OR PLANTS OR PARTS THEREOF; BIOCIDES, e.g. AS DISINFECTANTS, AS PESTICIDES OR AS HERBICIDES; PEST REPELLANTS OR ATTRACTANTS; PLANT GROWTH REGULATORS
- A01N43/00—Biocides, pest repellants or attractants, or plant growth regulators containing heterocyclic compounds
- A01N43/34—Biocides, pest repellants or attractants, or plant growth regulators containing heterocyclic compounds having rings with one nitrogen atom as the only ring hetero atom
- A01N43/40—Biocides, pest repellants or attractants, or plant growth regulators containing heterocyclic compounds having rings with one nitrogen atom as the only ring hetero atom six-membered rings
- A01N43/42—Biocides, pest repellants or attractants, or plant growth regulators containing heterocyclic compounds having rings with one nitrogen atom as the only ring hetero atom six-membered rings condensed with carbocyclic rings
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- C—CHEMISTRY; METALLURGY
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- C07D217/00—Heterocyclic compounds containing isoquinoline or hydrogenated isoquinoline ring systems
- C07D217/12—Heterocyclic compounds containing isoquinoline or hydrogenated isoquinoline ring systems with radicals, substituted by hetero atoms, attached to carbon atoms of the nitrogen-containing ring
- C07D217/14—Heterocyclic compounds containing isoquinoline or hydrogenated isoquinoline ring systems with radicals, substituted by hetero atoms, attached to carbon atoms of the nitrogen-containing ring other than aralkyl radicals
- C07D217/16—Heterocyclic compounds containing isoquinoline or hydrogenated isoquinoline ring systems with radicals, substituted by hetero atoms, attached to carbon atoms of the nitrogen-containing ring other than aralkyl radicals substituted by oxygen atoms
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Abstract
The invention discloses a kind of isoquinoline alkaloids bases compounds and its preparation method and application.The compound obtains after ethyl acetate extraction, silica gel column chromatography and high pressure liquid chromatography isolate and purify using Asia-Europe grass of meadow rue complete stool as raw material.Its molecular formula is C12H11NO2, there is following structural formula:Compound nomenclature are as follows: 1- (6- hydroxyl -7- methylisoquinolinium -1- base) ethyl ketone.The compound is significantly better than agricultural streptomycin to the inhibiting effect of Phytophthora nicotianae Breda, reaches (72.2 ± 3.3) % to tobacco black shank control efficiency.Raw materials for production of the present invention distribution is wide, biological yield is big, alkaloid is high, and compound preparation process is simple, and production cost is low, industrial application easy to accomplish.
Description
Technical field
The invention belongs to biological pesticide technical fields, and being specifically related to one kind, extraction separates for the first time from Asia-Europe grass of meadow rue
The isoquinoline alkaloids bases compound arrived.Meanwhile the invention further relates to the preparation methods of the compound and the compound in cigarette
Application in careless balck shank prevention and treatment.
Background technique
Thalictrum (Classification system: Thalictrum L.) belongs to Ranunculaceae herbaceos perennial.The platymiscium whole world
There are about 200 kinds, wherein China is there are about 67 kinds, majority is distributed in the west and south.In global thalictrum spp, have chemistry at
Dividing the platymiscium of report has more than 90, and main active has the types such as alkaloid, saponin, flavones.The category more than 43 kinds
Plant has Medicinal record in China.Asia-Europe grass of meadow rue (Classification system: Thalictrum minus) is under Thalictrum
A kind of herbaceous plant, plant is all hairless, and stem is 28-55 centimetres high, and stem life four times three goes out pinnate compound leaf.Inflorescence pyramid type；Sepal
4, pistac；Stamen is most, filigree silk shape.Achene is narrow oval spherical, slightly flat.The 6-7 month blooms.It is height above sea level 1400-2700 meters raw
Between mountainous region Cao Po, Tian Bian, in shrubbery or in woods.It is distributed widely in Asia, Europe.Sichuan, Yun Nanxi are distributed mainly in China
The areas such as portion, Qinghai, Xinjiang, Gansu, Shanxi.The plant be Yunnan, the civil common Chinese medicine in Sichuan, for treat dysentery and
Diarrhea.Domestic and international fitochemical studies early period show Asia-Europe grass of meadow rue rich in alkaloids active constituent.
Tobacco black shank is one of most destructive disease, also known as tobacco epidemic disease on tobacco leaf production, and tobacco grower is known as " black bar
It is crazy ", " black root ", " rhizome of Chinese monkshood disease " etc..Each main production cigarette district of China has different degrees of generation, and wherein Anhui, Shandong, Henan Province are
Historical grave illness area；The Main Tobacco-growing Regions In Souths such as Yunnan, Guizhou, Sichuan, Hunan, Guangdong, Guangxi, Fujian occur also fairly common.Mesh
The prevention and treatment of preceding balck shank mainly passes through the methods of rotation cropping, kind improvement of genes, biological pesticide and realizes prevention and treatment.Wherein with biology
Pesticide control is the most frequently used and is easiest to the method realized.
It is always biological both at home and abroad that the new active substance of control of plant disease effect is found to have from resources of medicinal plant
The hot spot of pesticide research.The present invention is extracted isolated by research to Asia-Europe grass of meadow rue chemical component and screening active ingredients
Relevant report is not yet seen in a kind of new isoquinoline alkaloids bases compound, the compound, it is worth mentioning at this point that the chemical combination
Object has the function of significant resisting tobacco black shank virus activity.
Summary of the invention
The purpose of the present invention is to provide a kind of new isoquinoline alkaloids bases compounds.
It is a further object to provide a kind of methods for preparing the isoquinoline alkaloids bases compound.
The object of the invention is also to provide the isoquinoline alkaloids bases compound answering in tobacco black shank prevention and treatment
With.
The purpose of the present invention is achieved by the following technical programs.
Unless otherwise indicated, percentage employed in the present invention is mass percent.
A kind of isoquinoline alkaloids bases compound, molecular formula C12H11NO2, there is following structural formula:
The Compound nomenclature are as follows: 1- (6- hydroxyl -7- methylisoquinolinium -1- base) ethyl ketone, English name are as follows: 1- (6-
hydroxy-7-methylisoquinolin-1-yl)ethanone。
A method of preparing the isoquinoline alkaloids bases compound, comprising the following steps:
(1) medicinal extract extracts: taking Asia-Europe grass of meadow rue complete stool to dry, is crushed to 35~60 mesh, smashed sample is placed in glass
In glass reaction kettle, with 95% alcohol reflux extract 2 times, merge twice extracting solution be concentrated to small size, then with 3% winestone
Acid solution dilution, then be extracted with ethyl acetate 2 times；Water phase is saturated with sodium carbonate after having extracted, and 2 are extracted with ethyl acetate again
It is secondary, merge the ethyl acetate phase of extraction, alkaloid position medicinal extract is concentrated under reduced pressure to obtain；
(2) silica gel column chromatography: the pure methanol or straight alcohol or pure C that medicinal extract obtained by step (1) is measured with 1.5~3 times of weight ratio
After ketone dissolution, with 0.8~2.5 times of weight ratio of 80~100 mesh silica gel mixed samples, drying；Dress column silica gel is 150~200 mesh, dosage
It is 3~6 times of medicinal extract weight；Ladder is carried out with the chloroform of weight ratio 20:1,9:1,8:2,7:3,6:4 and 5:5-acetone soln
Degree elution collects gradient eluent, concentration, monitors through TLC, merge identical part；
(3) high pressure liquid chromatography isolates and purifies: the part 6:4 of column chromatographic grade eluent being taken to carry out high pressure liquid chromatography point
From purifying: with Zorbax PrepHT GF, 21.2mm × 25cm reversed-phase column of Agilent company, being with 40% methanol aqueous solution
Mobile phase, flow velocity 20mL/min, UV detector Detection wavelength are 292nm, collect the chromatographic peak of 22.7min, repeatedly cumulative
It is evaporated afterwards to get the isoquinoline alkaloids bases compound is arrived.
In step (3), the compound obtained after high pressure liquid chromatography isolates and purifies is dissolved with pure methanol again, and with pure first
Alcohol is mobile phase, and with sephadex column chromatography for separation, obtained yellow jelly is pure compounds.
The Structural Identification of compound:
The compound of the present invention is yellow jelly, and HRESI-MS shows that its quasi-molecular ion peak is 224.0680 [M+Na]+,
In conjunction with1H and13C H NMR spectroscopy determines that molecular formula is C12H11NO2, it is alkaloid compound.Its infrared spectroscopy, which is shown in compound, to be had
Hydroxyl (3389cm-1), carbonyl (1668cm-1) and aromatic ring (1615,1547 and 1442cm-1) signal, ultraviolet spectra 212,258,
292 and 334nm has absorption maximum to also confirm that there are aromatic ring structures in compound.Compound1H、13C-NMR and DEPT composes (table-
1) show that it contains 12 carbon and 11 hydrogen.It include: isoquinolin parent nucleus (C-1~C-10 that a 1,6,7- replaces；H-3,H-4,
H-5 and H-8), an acetyl group (- CO-CH3；C-1 ' and C-2 '；H3- 2 '), a methyl (C-3 '；H3- 2 ') and a phenol hydroxyl
Base (δH10.75s).The presence of isoquinolin parent nucleus can pass through H-3 in compound and C-1, C-4, C-10；H-4 and C-3, C-5, C-
9,C-10；H-5 and C-4, C-9, C-10；And H-8 (figure -3) related to the HMBC of C-1, C-9, C-10 is confirmed.
After the parent nucleus of compound determines, by further analyzing its HMBC Correlated Spectroscopy, it may be determined that remaining substituent group (acetyl
Base, methyl and phenolic hydroxyl group) position.Pass through acetonyl hydrogen H3-2′(δHAnd C-1 (δ 2.46)C156.5) HMBC is related,
It can determine that acetyl group is substituted in C-1；Pass through aromatic ring methyl hydrogen H3-3′(δHAnd C-6 (δ 2.30)C 160.1)、C-7(δC
And C-8 (δ 154.0)CAnd H-8 (δ 127.5)HAnd C-3 ' (δ 8.49)C15.2) HMBC correlation can determine that the methyl takes
In generation, is at C-7；Phenolic hydroxyl group is substituted in C-6 can be by phenolic hydroxyl group hydrogen (δHAnd C-5 (δ 10.75)C 112.4)、C-6(δC
160.1)、C-7(δC132.2) HMBC correlation is confirmed.So far the structure of compound is confirmed, the Compound nomenclature
Are as follows: 1- (6- hydroxyl -7- methylisoquinolinium -1- base) ethyl ketone.
Table -1, compound1H and13C NMR data (C5D5N, 500 and 125MHz)
Infrared, the ultraviolet and mass spectrometric data of compound: UV (CH3OH)λmax(logε)212(4.04)、258(3.36)、292
(3.15),334(3.32)nm；IR(KBr)νmax 3389、3060、2961、1668、1615、1547、1442、1376、1242、
1168、1072、938、826cm-1；13C-NMR and1H-NMR data (C5D5N, 125 and 500MHz), it is shown in Table -1；ESIMS m/z
224[M+Na]+；HRESIMS m/z 224.0680[M+Na]+(calculated value C12H11NNaO2, 224.0687).
The application of isoquinoline alkaloids bases compound of the invention in tobacco black shank prevention and treatment.
1, tobacco black shank is caused by being disseminated by Phytophthora nicotianae Breda, and measurement the compounds of this invention inhibits phytophthora activity
Ability, comprising the following steps:
(1) preparation of oatmeal agar culture medium: oatmeal adds water 1000mL, heats 1 hour on boiling water bath, gauze mistake
After filter plus water supplies 1000mL, then sugaring and agar, after heating is completely melt agar, while hot with gauze (centre plus degreasing
Cotton) filtering is in triangular flask, and 121 DEG C, 15 pounds of sterilizing 20min take out and are cooled to 45 DEG C or so, in ammonia is added on aseptic operating platform
Parasiticin (5mg/100mL), is poured into plate after mixing, and 28 DEG C are cultivated 48 hours, sterile rear stand-by on inspection.
(2) bacteriostatic experiment: taking the circular filter paper of diameter 5mm, be put into culture dish, is placed in 15 pounds of high pressure sterilization 30min, dries
It is immersed respectively after dry in 20 μM of compound, 75% ethanol solution and sterile purified water.In using aseptic straw on aseptic operating platform
The fresh bacterium solution of 0.2mL Phytophthora nicotianae Breda is drawn respectively on oatmeal agar culture medium flat plate.Stick coating is applied with triangular glass
Uniformly, filter paper is gently affixed on corresponding plate respectively with tweezers, sets 28 DEG C of culture observation experiment results, measurement inhibition zone is big
It is small.Meanwhile using agricultural streptomycin as positive control.
Test result shows: isoquinoline alkaloids bases compound antibacterial circle diameter of the invention is 14.6 ± 1.2mm, positive
The antibacterial circle diameter for compareing agricultural streptomycin is 12.2 ± 1.0mm.Illustrate that the compounds of this invention inhibits the effect of Phytophthora nicotianae Breda
It is significantly better than positive control agricultural streptomycin, there is inhibition balck shank activity outstanding.
2, control efficiency of the compounds of this invention to tobacco black shank:
Tobacco seedlings are transplanted to diameter 10cm, in the flowerpot of high 10cm, cultivation matrix are as follows: sterile soil, turf, perlite culture
(2:2:1), 1 plant of every basin.10g/ plants of bacterium paddy are added in root after transplanting slow seedling, tobacco seedlings are placed in artificial climate room and are cultivated, it is white
It 30 DEG C, 28 DEG C of night, illumination: dark (12h:12h), relative humidity 95% make tobacco seedlings fall ill.It is used before the onset of balck shank
20 μM of the compounds of this invention carries out root irrigation to tobacco seedlings, and every plant is poured 10mL；It pours altogether 2 times, each 10 plants of processing is repeated 3 times,
Incidence is investigated after 14d, calculates disease index.The result shows that: the compounds of this invention is to tobacco black shank control efficiency
(72.2 ± 3.3) % has significant tobacco black shank control efficiency.
Compared with prior art, the invention has the following advantages that
(1) present invention extracts a kind of isolated isoquinoline alkaloids bases compound for the first time from Asia-Europe grass of meadow rue, right
The inhibiting effect of Phytophthora nicotianae Breda is significantly better than agricultural streptomycin, reaches (72.2 ± 3.3) % to tobacco black shank control efficiency.
The application of the compound will provide efficient, safe novel biopesticide molecular structure to black shank of tobacco prevention and treatment.
(2) raw materials for production (Asia-Europe grass of meadow rue) distribution of the present invention is wide, biological yield is big, alkaloid is high, compound system
Standby simple process, production cost is low, industrial application easy to accomplish.
Detailed description of the invention
Fig. 1 is the carbon-13 nmr spectra of isoquinoline alkaloids bases compound of the present invention；
Fig. 2 is the nuclear magnetic resonance spectroscopy of isoquinoline alkaloids bases compound of the present invention；
Fig. 3 is the main HMBC correlation figure of isoquinoline alkaloids bases compound of the present invention.
Specific embodiment
The present invention is described in further detail with reference to the accompanying drawings and examples, but drawings and examples are not pair
The restriction of technical solution of the present invention, it is all based on present invention teach that made variation or equivalent replacement, should belong to the present invention
Protection scope.
Embodiment 1
Prepare isoquinoline alkaloids bases compound C12H11NO2, including medicinal extract extraction, silica gel column chromatography, high pressure liquid chromatography
Separation specifically uses following steps:
(1) medicinal extract extracts: taking Asia-Europe grass of meadow rue complete stool to dry, is crushed to 35~60 mesh.After weighing 3.5~5.2kg crushing
Sample, be placed in the glass reaction kettle of 20L, 95% 8~12L of ethyl alcohol be added, 35~60min of refluxing extraction filters out extraction
Liquid；95% 8~12L of ethyl alcohol is added into filter residue again, 35~60min of refluxing extraction filters out extracting solution.Merging is extracted twice
Liquid is concentrated to small size, is then diluted with 3% 4~6L of tartaric acid solution, then is extracted 2 times with the ethyl acetate of 4~6L.Extraction
Water phase is saturated with sodium carbonate after complete, is extracted 2 times with the ethyl acetate of 4~6L again, and the ethyl acetate phase of extraction is merged, and is depressurized dense
Contract to obtain alkaloid position 85.6~128.5g of medicinal extract.
(2) it silica gel column chromatography: after the pure methanol or straight alcohol or pure acetone that medicinal extract is measured with 1.5~3 times of weight ratio dissolve, uses
120~280g (80-100 mesh) thick silica gel mixed sample, drying, with 260~440g silica gel (150-200 mesh) column chromatography, chloroform:
Acetone (20:1,9:1,8:2,7:3,6:4,5:5) gradient elution, is divided into 6 parts.
(3) high pressure liquid chromatography isolates and purifies: choosing 6:4 elution fraction progress HPLC and further separates: is public with Agilent
Zorbax PrepHT GF (21.2mm × 25cm) reversed-phase column of department, using 40% methanol aqueous solution as mobile phase, flow velocity 20mL/
Min, UV detector Detection wavelength are 292nm, collect the chromatographic peak of 22.7min, are evaporated after repeatedly adding up, it is thick to obtain compound
Product.The dissolution of crude product again with methanol, using methanol as mobile phase, with sephadex column purification, obtains pure compounds.
Asia and Europe grass of meadow rue raw material used in the present invention not by area and kind limited, the present invention may be implemented, below since
The present invention will be further described for Asia-Europe grass of meadow rue raw material derived from Yunnan:
Embodiment 2
Asia-Europe grass of meadow rue sample source is in Dali.It takes Asia-Europe grass of meadow rue complete stool to dry, is crushed to about 45 mesh.It weighs
The smashed sample of 3.8kg, is placed in the glass reaction kettle of 20L, and 95% ethyl alcohol 8L is added, and refluxing extraction 38min is filtered out and mentioned
Take liquid；95% ethyl alcohol 8L, refluxing extraction 40min is added into filter residue again, filters out extracting solution, merges extracting solution twice and is concentrated
To small size.Then it is diluted with 3% tartaric acid solution 6L, is extracted 2 times with the ethyl acetate of 6L.Water phase carbonic acid after having extracted
Sodium saturation, is extracted 2 times with the ethyl acetate of 6L again, merges the ethyl acetate phase of extraction, and the leaching of alkaloid position is concentrated under reduced pressure to obtain
Cream 86.4g.Medicinal extract 120g (80-100 mesh) thick silica gel mixed sample, drying, with 280g silica gel (150-200 mesh) column chromatography, trichlorine
Methane: acetone (20:1,9:1,8:2,7:3,6:4,5:5) gradient elution is divided into 6 parts.6:4 elution fraction is chosen to carry out
HPLC is further separated: with Zorbax PrepHT GF (21.2mm × 25cm) reversed-phase column of Agilent company, with 40% methanol
Aqueous solution is mobile phase, and flow velocity 20mL/min, UV detector Detection wavelength is 292nm, collects the chromatographic peak of 22.7min,
It is evaporated after repeatedly adding up, obtains crude compound.The dissolution of crude product again with methanol, it is net with sephadex column using methanol as mobile phase
Change, obtains pure compounds.
Embodiment 3
Asia-Europe grass of meadow rue sample source is in Dongzhou Period in Chuxiong.It takes Asia-Europe grass of meadow rue complete stool to dry, is crushed to 55 mesh.It weighs
The smashed sample of 4.6kg, is placed in the glass reaction kettle of 20L, and 95% ethyl alcohol 12L is added, and refluxing extraction 25min is filtered out
Extracting solution；95% ethyl alcohol 12L, refluxing extraction 30min is added into filter residue again, filters out extracting solution, merges extracting solution twice
It is concentrated to small size, is then diluted with 3% tartaric acid solution 10L, is extracted 2 times with the ethyl acetate of 10L.Water phase after having extracted
It is saturated with sodium carbonate, is extracted 2 times with the ethyl acetate of 10L again, merge the ethyl acetate phase of extraction, alkaloid is concentrated under reduced pressure to obtain
Position medicinal extract 116.8g.Medicinal extract 200g (80-100 mesh) thick silica gel mixed sample, drying, with 380g silica gel (150-200 mesh) column color
Spectrum, chloroform: acetone (20:1,9:1,8:2,7:3,6:4,5:5) gradient elution is divided into 6 parts.Choose 6:4 elution portion
Point carrying out HPLC further separates: with Zorbax PrepHT GF (21.2mm × 25cm) reversed-phase column of Agilent company, with
40% methanol aqueous solution is mobile phase, and flow velocity 20mL/min, UV detector Detection wavelength is 292nm, collects 22.7min's
Chromatographic peak is evaporated after repeatedly adding up, obtains crude compound.The dissolution of crude product again with methanol, it is solidifying with glucan using methanol as mobile phase
Rubber column gel column purification, obtains pure compounds.
Embodiment 4
Asia-Europe grass of meadow rue sample source is in Jianchuan.It takes Asia-Europe grass of meadow rue complete stool to dry, is crushed to 38 mesh.It weighs
The smashed sample of 5.0kg, is placed in the glass reaction kettle of 20L, and 95% ethyl alcohol 12L is added, and refluxing extraction 30min is filtered out
Extracting solution；95% ethyl alcohol 10L, refluxing extraction 50min is added into filter residue again, filters out extracting solution, merges extracting solution twice
It is concentrated to small size, is then diluted with 3% tartaric acid solution 10L, is extracted 2 times with the ethyl acetate of 10L.Water phase after having extracted
It is saturated with sodium carbonate, is extracted 2 times with the ethyl acetate of 8L again, merge the ethyl acetate phase of extraction, alkaloid is concentrated under reduced pressure to obtain
Position medicinal extract 124.7g.Medicinal extract 200g (80-100 mesh) thick silica gel mixed sample, drying, with 440g silica gel (150-200 mesh) column color
Spectrum, chloroform: acetone (20:1,9:1,8:2,7:3,6:4,5:5) gradient elution is divided into 6 parts.Choose 6:4 elution portion
Point carrying out HPLC further separates: with Zorbax PrepHT GF (21.2mm × 25cm) reversed-phase column of Agilent company, with
40% methanol aqueous solution is mobile phase, and flow velocity 20mL/min, UV detector Detection wavelength is 292nm, collects 22.7min's
Chromatographic peak is evaporated after repeatedly adding up, obtains crude compound.The dissolution of crude product again with methanol, it is solidifying with glucan using methanol as mobile phase
Rubber column gel column purification, obtains pure compounds.
Embodiment 5
The identification of --- --- compound structure
Isoquinoline alkaloids bases compound prepared by Example 2, is measured by the following method.Compound is yellow
Jelly, HRESI-MS show that its quasi-molecular ion peak is 224.0680 [M+Na]+, in conjunction with1H and13C H NMR spectroscopy determines molecular formula
For C12H11NO2, it is alkaloid compound.Its infrared spectroscopy, which is shown in compound, hydroxyl (3389cm-1), carbonyl (1668cm-1) and aromatic ring (1615,1547 and 1442cm-1) signal, ultraviolet spectra has absorption maximum also to demonstrate,prove in 212,258,292 and 334nm
There are aromatic ring structures in real compound.Compound1H、13C-NMR and DEPT spectrum (table -1) shows that it contains 12 carbon and 11
Hydrogen.It include: isoquinolin parent nucleus (C-1~C-10 that a 1,6,7- replaces；H-3, H-4, H-5 and H-8), acetyl group (-
CO-CH3；C-1 ' and C-2 '；H3- 2 '), a methyl (C-3 '；H3- 2 ') and a phenolic hydroxyl group (δH10.75s).Isoquinolin is female
The presence of core can pass through H-3 in compound and C-1, C-4, C-10；H-4 and C-3, C-5, C-9, C-10；H-5 and C-4, C-9, C-
10；And H-8 (figure -3) related to the HMBC of C-1, C-9, C-10 is confirmed.
After the parent nucleus of compound determines, by further analyzing its HMBC Correlated Spectroscopy, it may be determined that remaining substituent group (acetyl
Base, methyl and phenolic hydroxyl group) position.Pass through acetonyl hydrogen H3-2′(δHAnd C-1 (δ 2.46)C156.5) HMBC is related,
It can determine that acetyl group is substituted in C-1；Pass through aromatic ring methyl hydrogen H3-3′(δHAnd C-6 (δ 2.30)C 160.1)、C-7(δC
And C-8 (δ 154.0)CAnd H-8 (δ 127.5)HAnd C-3 ' (δ 8.49)C15.2) HMBC correlation can determine that the methyl takes
In generation, is at C-7；Phenolic hydroxyl group is substituted in C-6 can be by phenolic hydroxyl group hydrogen (δHAnd C-5 (δ 10.75)C 112.4)、C-6(δC
160.1)、C-7(δC132.2) HMBC correlation is confirmed.So far the structure of compound is confirmed, the Compound nomenclature
Are as follows: 1- (6- hydroxyl -7- methylisoquinolinium -1- base) ethyl ketone.
Embodiment 6
Compound prepared by Example 3 is yellow jelly.Measuring method is same as Example 5, confirms embodiment 3
The compound of preparation is the isoquinoline alkaloids bases compound --- 1- (6- hydroxyl -7- methylisoquinolinium -1- base) ethyl ketone.
Embodiment 7
Compound prepared by Example 4 is yellow jelly.Measuring method is same as Example 5, confirms embodiment 4
The compound of preparation is described 1- (6- hydroxyl -7- methylisoquinolinium -1- base) ethyl ketone.
Embodiment 8
Any isoquinoline alkaloids bases compound of Example 1-4 preparation carries out resisting tobacco black shank activity test, examination
It is as follows to test situation:
(1) oatmeal adds water 1000mL, heats 1 hour on boiling water bath, and after filtered through gauze plus water supplies 1000mL, then
Sugaring and agar, after heating is completely melt agar, while hot with gauze (centre plus absorbent cotton) filtering in triangular flask, 121 DEG C,
15 pounds of sterilizing 20min, taking-up are cooled to 45 DEG C or so, in ampicillin (5mg/100mL) is added on aseptic operating platform, mix
After be poured into plate, 28 DEG C are cultivated 48 hours, stand-by after sterile on inspection.
(2) circular filter paper for taking diameter 5mm, is put into culture dish, is placed in 15 pounds of high pressure sterilization 30min, after drying respectively
It immerses in 20 μM of compound, 75% ethanol solution and sterile purified water.In being drawn respectively on aseptic operating platform with aseptic straw
The fresh bacterium solution of 0.2mL Phytophthora nicotianae Breda is on oatmeal agar culture medium flat plate.Stick coating is applied uniformly with triangular glass, will be filtered
The scraps of paper are gently affixed on corresponding plate respectively with tweezers, set 28 DEG C of culture observation experiment results, measure inhibition zone size.Meanwhile
Using agricultural streptomycin as positive control.
(3) test result shows: isoquinoline alkaloids bases compound antibacterial circle diameter of the invention is 14.6 ± 1.2mm,
The antibacterial circle diameter of positive control agricultural streptomycin is 12.2 ± 1.0mm.Illustrate that the compounds of this invention inhibits Phytophthora nicotianae Breda effect
Fruit is significantly better than positive control agricultural streptomycin, has inhibition balck shank activity outstanding.
(4) tobacco seedlings are transplanted to diameter 10cm, in the flowerpot of high 10cm, cultivation matrix are as follows: sterile soil, turf, perlite
It cultivates (2:2:1), 1 plant of every basin.10g/ plants of bacterium paddy are added in root after transplanting slow seedling, tobacco seedlings are placed in artificial climate room and are trained
It supports, 30 DEG C of daytime, 28 DEG C of night, illumination: dark (12h:12h), relative humidity 95%., tobacco seedlings is made to fall ill.It falls ill in balck shank
Preceding to carry out root irrigation to tobacco seedlings using 20 μM of the compounds of this invention, every plant is poured 10mL；It pours altogether 2 times., each 10 plants of processing,
It is repeated 3 times, incidence is investigated after 14d, calculate disease index.The result shows that: the compounds of this invention prevents and treats tobacco black shank
Effect is (72.2 ± 3.3) %, has significant tobacco black shank control efficiency.
Claims (4)
1. a kind of isoquinoline alkaloids bases compound, molecular formula C12H11NO2, there is following structural formula:
The Compound nomenclature are as follows: 1- (6- hydroxyl -7- methylisoquinolinium -1- base) ethyl ketone, English name are as follows: 1- (6-hydroxy-7-
methylisoquinolin-1-yl)ethanone。
2. the preparation method of isoquinoline alkaloids bases compound described in claim 1, comprising the following steps:
(1) medicinal extract extracts: taking Asia-Europe grass of meadow rue complete stool to dry, is crushed to 35~60 mesh, it is anti-that smashed sample is placed in glass
It answers in kettle, is extracted 2 times with 95% alcohol reflux, merge extracting solution twice and be concentrated to small size, it is then molten with 3% tartaric acid
Liquid dilution, then be extracted with ethyl acetate 2 times；Water phase is saturated with sodium carbonate after having extracted, and is extracted with ethyl acetate again 2 times, is closed
And the ethyl acetate phase extracted, alkaloid position medicinal extract is concentrated under reduced pressure to obtain；
(2) silica gel column chromatography: the pure methanol or straight alcohol or pure acetone that medicinal extract obtained by step (1) is measured with 1.5~3 times of weight ratio are molten
Xie Hou, with 0.8~2.5 times of weight ratio of 80~100 mesh silica gel mixed samples, drying；Dress column silica gel is 150~200 mesh, and dosage is leaching
3~6 times of cream weight；Gradient is carried out with the chloroform of weight ratio 20:1,9:1,8:2,7:3,6:4 and 5:5-acetone soln to wash
It is de-, gradient eluent, concentration are collected, is monitored through TLC, merges identical part；
(3) high pressure liquid chromatography isolates and purifies: taking the part 6:4 of column chromatographic grade eluent to carry out high pressure liquid chromatography separation pure
Change: being flowing with 40% methanol aqueous solution with Zorbax PrepHT GF, 21.2mm × 25cm reversed-phase column of Agilent company
Phase, flow velocity 20mL/min, UV detector Detection wavelength are 292nm, collect the chromatographic peak of 22.7min, are steamed after repeatedly adding up
It does to get the isoquinoline alkaloids bases compound is arrived.
3. preparation method according to claim 2, it is characterised in that: in step (3), isolated and purified through high pressure liquid chromatography
The compound obtained afterwards is dissolved with pure methanol again, and using pure methanol as mobile phase, with sephadex column chromatography for separation, is obtained
Yellow jelly is pure compounds.
4. application of the isoquinoline alkaloids bases compound described in claim 1 in tobacco black shank prevention and treatment.
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