CN113185484B - Chromone compound and preparation method and application thereof - Google Patents
Chromone compound and preparation method and application thereof Download PDFInfo
- Publication number
- CN113185484B CN113185484B CN202110385027.7A CN202110385027A CN113185484B CN 113185484 B CN113185484 B CN 113185484B CN 202110385027 A CN202110385027 A CN 202110385027A CN 113185484 B CN113185484 B CN 113185484B
- Authority
- CN
- China
- Prior art keywords
- extract
- eluent
- organic solvent
- compound
- silica gel
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Active
Links
Images
Classifications
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07D—HETEROCYCLIC COMPOUNDS
- C07D311/00—Heterocyclic compounds containing six-membered rings having one oxygen atom as the only hetero atom, condensed with other rings
- C07D311/02—Heterocyclic compounds containing six-membered rings having one oxygen atom as the only hetero atom, condensed with other rings ortho- or peri-condensed with carbocyclic rings or ring systems
- C07D311/04—Benzo[b]pyrans, not hydrogenated in the carbocyclic ring
- C07D311/22—Benzo[b]pyrans, not hydrogenated in the carbocyclic ring with oxygen or sulfur atoms directly attached in position 4
-
- 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
- A01N43/02—Biocides, pest repellants or attractants, or plant growth regulators containing heterocyclic compounds having rings with one or more oxygen or sulfur atoms as the only ring hetero atoms
- A01N43/04—Biocides, pest repellants or attractants, or plant growth regulators containing heterocyclic compounds having rings with one or more oxygen or sulfur atoms as the only ring hetero atoms with one hetero atom
- A01N43/14—Biocides, pest repellants or attractants, or plant growth regulators containing heterocyclic compounds having rings with one or more oxygen or sulfur atoms as the only ring hetero atoms with one hetero atom six-membered rings
- A01N43/16—Biocides, pest repellants or attractants, or plant growth regulators containing heterocyclic compounds having rings with one or more oxygen or sulfur atoms as the only ring hetero atoms with one hetero atom six-membered rings with oxygen as the ring hetero atom
-
- Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
- Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
- Y02A—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE
- Y02A50/00—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE in human health protection, e.g. against extreme weather
- Y02A50/30—Against vector-borne diseases, e.g. mosquito-borne, fly-borne, tick-borne or waterborne diseases whose impact is exacerbated by climate change
Landscapes
- Chemical & Material Sciences (AREA)
- Organic Chemistry (AREA)
- Life Sciences & Earth Sciences (AREA)
- Agronomy & Crop Science (AREA)
- Pest Control & Pesticides (AREA)
- Plant Pathology (AREA)
- Health & Medical Sciences (AREA)
- Engineering & Computer Science (AREA)
- Dentistry (AREA)
- General Health & Medical Sciences (AREA)
- Wood Science & Technology (AREA)
- Zoology (AREA)
- Environmental Sciences (AREA)
- Heterocyclic Carbon Compounds Containing A Hetero Ring Having Oxygen Or Sulfur (AREA)
- Medicines Containing Plant Substances (AREA)
- Pharmaceuticals Containing Other Organic And Inorganic Compounds (AREA)
Abstract
The invention discloses a chromone compound, a preparation method and application thereof, wherein the chromone compound takes branches and leaves of dry cassia plants of Leguminosae as raw materials, and the raw materials are subjected to the steps of extract extraction, organic solvent extraction, silica gel column chromatography and high-pressure liquid chromatography separation. Through experiments on resisting tobacco mosaic virus, the inhibition rate of the compound reaches 28.7+/-4.1%, and the compound has good activity on resisting tobacco mosaic virus, and is similar to that of a positive control, namely naningmycin (32.2+/-3.2%). The test proves that the compound has good tobacco mosaic virus resistance, simple structure and good activity; can be used as lead compound of tobacco mosaic virus resisting medicine.
Description
Technical Field
The invention relates to the technical field of extraction of plant active ingredients, in particular to a chromone compound and a preparation method and application thereof.
Background
Chromone compounds widely exist in nature, have antibacterial and anti-inflammatory effects and are important components in antibacterial and anti-inflammatory drugs. Current chromone compounds are generally extracted from plants, such as the leguminous caragana genus of plants.
About 600 species of Cassia (Cassia) plants are distributed in tropical and subtropical regions of the world, and a few species are distributed in temperate regions; the original products of China are more than 10, including more than 20 of introduction and cultivation, and are widely distributed in various provinces of south and north. The cassia plant is one of the main resources of natural flavone (flavonoids) components. In addition to flavonoids, anthraquinone (anthraquinone) and polysaccharide (polysaccharide) compounds are characteristic components of plants of this genus and have various biological activities.
In the prior art, the research on the cassia plants is very few, and the report of extracting the chromone compounds from the cassia plants is also rare. In order to more effectively utilize the plant resources of the cassia genus in China, the active ingredients with development prospects are searched from the plant resources of the cassia genus, and the active ingredients are an important work of current cassia genus plant research.
Disclosure of Invention
In view of the above, it is necessary to provide a chromone compound, and a preparation method and application thereof, based on extraction and research of active ingredients of cassia plants.
In one aspect, the present invention provides a chromone compound having the following chemical structural formula:
the chromone compound is prepared from dried branches, leaves or fruits of Cassia plant by extracting with extract, extracting with organic solvent, subjecting to silica gel column chromatography, and separating with high pressure liquid chromatography, and has molecular formula of C 13 H 12 O 5 This compound was designated 7-hydroxy-5- (3-hydroxy-2-oxoopyl) -2-methyl-4H-chromen-4-one.
The invention also provides a preparation method of the chromone compound, which comprises the following steps:
A. extracting extract: pulverizing branches, leaves or fruits of cassia plants to 20-40 meshes, ultrasonically extracting for 2-4 times by using an organic solvent for 30-60 min each time, combining the extracting solutions, filtering the extracting solutions, concentrating the extracting solutions under reduced pressure to 1/4-1/2 of the volume, standing, filtering out precipitate, and concentrating into a first extract;
B. extracting with an organic solvent: adding water into the extract a, wherein the weight of the added water is 1-2 times of that of the extract, extracting for 3-5 times by using an organic solvent with the same volume as that of the water, combining the organic solvent extraction phases, and concentrating under reduced pressure to obtain an extract b;
C. silica gel column chromatography: dissolving the extract b by using an organic solvent with the weight of 1.5-3 times, then mixing the extract b with 200-300 meshes of silica gel with the weight of 0.8-1.2 times of the extract b, then loading the extract b on a silica gel column for chromatography, wherein the loading silica gel is 200-300 meshes, and the loading silica gel is 6-8 times of the weight of the extract b; gradient eluting with two mixed organic solvents at a volume ratio of 1:0-0:1, collecting gradient eluent, concentrating, monitoring by TLC, and mixing the same parts;
D. reversed phase column chromatography: c, loading the eluent obtained by combining the same parts in the step C on a reversed phase column for chromatography, wherein the reversed phase column is filled with reversed phase materials; gradient elution is carried out by using a methanol aqueous solution with the volume content of 20-100%, three solutions with sequentially increased methanol concentration are adopted in the gradient elution process, eluent of each part is collected and concentrated, TLC monitoring is carried out, and the same parts are combined to obtain 3 components;
E. gradient eluting the 1 st component of the eluent in the step D by using a normal phase silica gel column, wherein the eluent is petroleum ether-ethyl acetate solution, and 8 petroleum ether-ethyl acetate solutions with sequentially increased ethyl acetate concentration are adopted in the gradient elution process to obtain 8 components;
F. high performance liquid chromatography separation: separating and purifying the concentrated solution of the 5 th component of the eluent in the step E through high performance liquid chromatography to obtain the chromone compound 7-hydroxy-5- (3-hydroxy-2-oxypropyl) -2-methyl-4H-chromen-4-one.
In the invention, the cassia plant is researched, and the novel chromone compound is obtained through the separation and preparation of active ingredients in the cassia plant. The chromone compounds are identified by nuclear magnetic resonance and other spectroscopic methods of determination and characterized by the specific structure:
further, the preparation method of the chromone compound comprises the steps of separating and purifying the concentrated solution of the 5 th component of the eluent in the step E by high performance liquid chromatography, wherein the method comprises the following steps:
loading the concentrated solution of the 5 th component of the eluent in the step E on a high performance liquid chromatographic column, taking 45-65% methanol as a mobile phase, taking a reversed phase preparation column with the flow rate of 8-14 ml/min and 21.2mm multiplied by 25cm and 5 mu m as a stationary phase, detecting the wavelength of 254nm by an ultraviolet detector, feeding 45-60 mu L each time, collecting chromatographic peaks for 15-33 min, accumulating for multiple times, and evaporating to dryness to obtain the chromone compound.
Further, the preparation method of the chromone compound comprises the step A, wherein the organic solvent adopted in the extract extraction is 80-100% of acetone, ethanol or methanol.
In the step B, the organic solvent used for extracting the extract a is ethyl acetate, chloroform, diethyl ether, petroleum ether or benzene.
Further, the mixed organic solvent in the step C is n-hexane-acetone, chloroform-methanol, petroleum ether-acetone and petroleum ether-ethyl acetate.
Further, in the preparation method of the chromone compound, the volume ratio of the mixed organic solvent in the step C is 1:0, 20:1, 9:1, 8:2, 7:3, 6:4, 1:2 and 0:1.
The invention also provides application of the chromone compound in preparing a medicine for resisting tobacco mosaic virus. The anti-tobacco mosaic virus medicine takes the chromone compound as the only active component.
Through experiments on resisting tobacco mosaic virus, the inhibition rate of the compound reaches 28.7+/-4.1%, and the compound has good activity on resisting tobacco mosaic virus, and is similar to that of a positive control, namely naningmycin (32.2+/-3.2%). The test proves that the compound has good tobacco mosaic virus resistance, simple structure and good activity; can be used as lead compound of tobacco mosaic virus resisting medicine.
Drawings
FIG. 1 is a compound
Nuclear magnetic resonance carbon spectrum of 7-hydroxy-5- (3-hydroxy-2-oxoopyl) -2-methyl-4H-chromen-4-one 13 C NMR)；
FIG. 2 is a compound
Nuclear magnetic resonance hydrogen spectrum of 7-hydroxy-5- (3-hydroxy-2-oxoopyyl) -2-methyl-4H-chroman-4-one 1 H NMR)；
FIG. 3 is a compound
The main HMBC (& gtfwdarw) correlation of 7-hydroxy-5- (3-hydroxy-2-oxoopyl) -2-methyl-4H-chromen-4-one.
Detailed Description
The following detailed description of the present invention will provide further understanding of the objects, features and advantages of the present invention by way of example. Several embodiments of the invention are presented in the examples. This invention may, however, be embodied in many different forms and should not be construed as limited to the embodiments set forth herein. Rather, these embodiments are provided so that this disclosure will be thorough and complete.
Example 1
The preparation method of the chromone compound in the embodiment comprises the following steps:
a1, taking 6.5kg of dried branches, leaves and/or fruits of cassia plants, coarsely crushing the branches, leaves and/or fruits to 20 meshes, ultrasonically extracting the branches, leaves and/or fruits with 80% acetone for 4 times, each time for 30min, and combining the extracting solutions; filtering the extract, and concentrating under reduced pressure to 1/4 of the volume; standing, filtering out precipitate, and concentrating into 380g of extract a;
b1, adding 480g of water into the extract a, extracting for 5 times by chloroform with the same volume as the water, combining the extracted phases, and concentrating under reduced pressure to 154g of extract B;
c1, loading 1300g of 200-mesh silica gel into a column, adding 450g of acetone into the extract b for dissolution, then adding 180g of 100-mesh silica gel, stirring, and loading the mixture into the column after stirring; gradient eluting with dichloromethane-ethyl acetate mixed organic solvents with volume ratios of 1:0, 20:1, 9:1, 8:2, 7:3, 6:4, 1:2 and 0:1 respectively, collecting gradient eluates, concentrating, monitoring by TLC, and combining the same parts to obtain 8 components, wherein the eluent obtained by eluting the 5 th component, namely the dichloromethane-ethyl acetate mixed organic solvent with the volume ratio of 7:3 is 60g;
loading the column with a reversed phase material C-18, loading the eluent of the 5 th component obtained by eluting in the step C1 on the reversed phase column, carrying out gradient elution by using a methanol aqueous solution with the volume content of 20-100%, adopting solutions with three concentration ratios with sequentially increased methanol concentration in the gradient elution process, collecting and concentrating eluent of each part, monitoring by TLC, and merging the same parts to obtain 3 components; in the step, the three concentration ratios are sequentially 20%,40%,70% and 100%, the polarities of the obtained 3 components are different, and the chromone compound only exists in the 1 st component;
e1, carrying out gradient elution on the eluent of the 1 st component in the step D1 by using a normal phase silica gel column, wherein the eluent is petroleum ether-ethyl acetate solution, and the volume ratio of the petroleum ether-ethyl acetate solution in the gradient elution process is 1:0, 20:1, 9:1, 8:2, 7:3, 6:4, 1:2 and 0:1 to obtain 8 components;
f1, eluting with an organic solvent with a volume ratio of 7:3 in the step E1 to obtain an eluent, taking a methanol aqueous solution with a volume content of 45-60% as a mobile phase, taking a Zorbax PrepHT GF reversed phase preparation column with a flow rate of 8ml/min and a detection wavelength of 254nm and 5 mu m as a stationary phase, feeding 50 mu L each time, collecting chromatographic peaks for 21min, accumulating for multiple times, and evaporating to dryness to obtain the chromone compound 7-hydroxy-5- (3-hydroxy-2-oxypropyl) -2-methyl-4H-chromen-4-one.
Example 2
The preparation method of the chromone compound in the embodiment comprises the following steps:
a2, taking 3.3kg of dried branches, leaves and/or fruits of the cassia plant, coarsely crushing the branches, leaves and/or fruits to 20 meshes, ultrasonically extracting the branches, leaves and/or fruits with 100% ethanol for 2 times, each time for 30min, and combining the extracting solutions; filtering the extract, and concentrating under reduced pressure to 1/3 of the volume; standing, filtering out precipitate, and concentrating into 280g of extract a;
b2, adding 280g of water into the extract a, extracting for 3 times by using chloroform with the same volume as the water, combining the extracted phases, and concentrating under reduced pressure to 110g of extract B;
c2, filling columns with 680g of 160-mesh silica gel, adding 150g of acetone into the extract b for dissolution, then adding 90g of 80-mesh silica gel for sample mixing, and filling the columns after sample mixing; gradient eluting with petroleum ether-acetone mixed organic solvents with volume ratios of 1:0, 20:1, 9:1, 8:2, 7:3, 6:4, 1:2 and 0:1 respectively, collecting gradient eluates, concentrating, monitoring by TLC, and combining the same parts to obtain eluates with 8 components, wherein the eluent obtained by eluting the 5 th component, namely petroleum ether-acetone mixed organic solvent with volume ratio of 7:3 is 48g;
loading the column with a reversed phase material C-18, loading the eluent of the 5 th component in the step C2 on the reversed phase column, carrying out gradient elution by using a methanol aqueous solution with the volume content of 20-100%, adopting solutions with three concentration ratios with sequentially increased methanol concentration in the gradient elution process, collecting and concentrating eluent of each part, monitoring by TLC, and merging the same parts to obtain 3 components;
e2, carrying out gradient elution on the eluent of the 1 st component in the step D1 by using a normal phase silica gel column, wherein the eluent is petroleum ether-ethyl acetate solution, and the volume ratio of the petroleum ether-ethyl acetate solution in the gradient elution process is 1:0, 20:1, 9:1, 8:2, 7:3, 6:4, 1:2 and 0:1 to obtain 8 components;
f2, eluting with an organic solvent with a volume ratio of 7:3 in the step E2 to obtain an eluent, taking a methanol aqueous solution with a volume content of 40-60% as a mobile phase, taking a Zorbax PrepHT GF reversed phase preparation column with a flow rate of 8ml/min and a detection wavelength of 254nm and 5 mu m as a stationary phase, feeding 50 mu L each time, collecting chromatographic peaks for 22min, accumulating for multiple times, and evaporating to dryness to obtain the chromone compound 7-hydroxy-5- (3-hydroxy-2-oxypropyl) -2-methyl-4H-chromen-4-one.
Example 3
The preparation method of the chromone compound in the embodiment comprises the following steps:
a3, taking 6.5kg of dried branches, leaves and/or fruits of the cassia plant, coarsely crushing the branches, leaves and/or fruits to 30 meshes, ultrasonically extracting the branches, leaves and/or fruits with 80% methanol for 4 times, each time for 30min, and combining the extracting solutions; filtering the extract, and concentrating under reduced pressure to 1/2 of the volume; standing, filtering out precipitate, and concentrating into 416g of extract a;
adding 460g of water into the extract a, extracting for 4 times by using diethyl ether with the same volume as the water, combining the extracted phases, and concentrating under reduced pressure to 243g of extract B;
c3, filling 1400g of 180-mesh silica gel into a column, adding 340g of acetone into the extract b for dissolution, then adding 250g of 90-mesh silica gel for sample mixing, and filling the column after sample mixing; gradient eluting with dichloromethane-acetone mixed organic solvent with volume ratio of 1:0, 20:1, 9:1, 8:2, 7:3, 6:4, 1:2, and 0:1 respectively, collecting gradient eluent, concentrating, monitoring by TLC, and mixing the same parts to obtain 8 components, wherein the 5 th component, i.e. eluent of dichloromethane-acetone mixed organic solvent with volume ratio of 7:3 is 50g;
loading the column with an anti-phase material ODS, loading the eluent of the 3 rd component in the step C3 on the anti-phase column, carrying out gradient elution by using a methanol water solution with the volume content of 20-100%, adopting solutions with three concentration ratios with sequentially increased methanol concentration in the gradient elution process, collecting and concentrating each part of eluent, monitoring by TLC, and merging the same parts to obtain 3 components;
e3, carrying out gradient elution on the eluent of the 1 st component in the step D3 by using a normal phase silica gel column, wherein the eluent is petroleum ether-ethyl acetate solution, and the volume ratio of the petroleum ether-ethyl acetate solution in the gradient elution process is 1:0, 20:1, 9:1, 8:2, 7:3, 6:4, 1:2 and 0:1 to obtain 8 components;
f3, eluting with an organic solvent with a volume ratio of 7:3 in the step E3 to obtain an eluent, taking a methanol aqueous solution with a volume content of 40-60% as a mobile phase, taking a Zorbax PrepHT GF reversed phase preparation column with a flow rate of 10ml/min and a detection wavelength of 254nm and 5 mu m as a stationary phase, feeding 50 mu L each time, collecting chromatographic peaks for 24min, accumulating for multiple times, and evaporating to dryness to obtain the chromone compound 7-hydroxy-5- (3-hydroxy-2-oxypropyl) -2-methyl-4H-chromen-4-one.
Example 4
The preparation method of the chromone compound in the embodiment comprises the following steps:
a4, taking 5.5kg of dried branches, leaves and/or fruits of the cassia plant, coarsely crushing the branches, leaves and/or fruits to 40 meshes, extracting the branches, leaves and/or fruits with 90% ethanol for 3 times, and combining the extracting solutions; filtering the extract, and concentrating under reduced pressure to 1/4 of the volume; standing, filtering out precipitate, and concentrating into 420g of extract a;
adding 490g of water into the extract a, extracting with petroleum ether with the same volume as the water for 4 times, combining the extracted phases, and concentrating under reduced pressure to 260g of extract B;
c4, filling 1400g of 160-mesh silica gel into a column, adding 300g of acetone into the extract b for dissolution, then adding 260g of 80-mesh silica gel for sample mixing, and filling the column after sample mixing; gradient eluting with chloroform-acetone mixed organic solvent with volume ratio of 1:0, 20:1, 9:1, 8:2, 7:3, 6:4, 1:2, and 0:1 respectively, collecting gradient eluent, concentrating, monitoring by TLC, and mixing the same parts to obtain 8 components, wherein the eluent obtained by eluting the 5 th component with chloroform-acetone mixed organic solvent with volume ratio of 7:3 is 55g;
loading the column with a reversed phase material C-8, loading the 5 th component in the step C3 onto the reversed phase column, carrying out gradient elution by using a methanol aqueous solution with the volume content of 20-100%, adopting solutions with three concentration ratios with sequentially increased methanol concentration in the gradient elution process, collecting eluent of each part, concentrating, monitoring by TLC, and merging the same parts to obtain 3 components;
e4, carrying out gradient elution on the eluent of the 1 st component in the step D4 by using a normal phase silica gel column, wherein the eluent is petroleum ether-ethyl acetate solution, and the volume ratio of the petroleum ether-ethyl acetate solution in the gradient elution process is 1:0, 20:1, 9:1, 8:2, 7:3, 6:4, 1:2 and 0:1 to obtain 8 components;
f4, eluting with an organic solvent with a volume ratio of 7:3 in the step E3 to obtain an eluent, taking a methanol aqueous solution with a volume content of 40-60% as a mobile phase, taking a Zorbax PrepHT GF reversed phase preparation column with a flow rate of 10ml/min and a wavelength of 21.2mm multiplied by 25cm and 5 mu m as a stationary phase, collecting chromatographic peaks with a detection wavelength of 254nm for 20min, accumulating for multiple times, and evaporating to obtain the chromone compound 7-hydroxy-5- (3-hydroxy-2-oxypropyl) -2-methyl-4H-chromen-4-one.
Example 5
The preparation method of the chromone compound in the embodiment comprises the following steps:
a5, taking 5.7kg of dried branches, leaves and/or fruits of the cassia plant, coarsely crushing the branches, leaves and/or fruits to 20 meshes, ultrasonically extracting the branches, leaves and/or fruits with 70% methanol for 4 times, each time for 35min, and combining the extracting solutions; filtering the extract, and concentrating under reduced pressure to 1/2 of the volume; standing, filtering out precipitate, and concentrating into 310g of extract a;
adding 500g of water into the extract a, extracting with benzene with the same volume as the water for 5 times, combining the extraction phases, and concentrating under reduced pressure to 120g of extract B;
loading 850g of 200-mesh silica gel into a column, adding 130g of acetone into the extract b for dissolution, adding 120g of 100-mesh silica gel into the extract b, stirring, and loading the mixture into the column after stirring; gradient eluting with petroleum ether-chloroform mixed organic solvent with volume ratio of 1:0, 20:1, 9:1, 8:2, 7:3, 6:4, 1:2 and 0:1 respectively, collecting gradient eluent, concentrating, monitoring by TLC, and combining the same parts to obtain 8 components, wherein the eluent obtained by the 5 th component, namely petroleum ether-chloroform mixed organic solvent system with volume ratio of 7:3 is 58g;
loading the column with an anti-phase material ODS, loading the eluent of the 5 th component obtained by eluting in the step C1 on the anti-phase column, carrying out gradient elution by using a methanol aqueous solution with the volume content of 20-100%, adopting solutions with three concentration ratios with sequentially increased methanol concentration in the gradient elution process, collecting and concentrating each part of eluent, monitoring by TLC, and merging the same parts to obtain 3 components;
e5, carrying out gradient elution on the eluent of the 1 st component in the step D1 by using a normal phase silica gel column, wherein the eluent is petroleum ether-ethyl acetate solution, and the volume ratio of the petroleum ether-ethyl acetate solution in the gradient elution process is 1:0, 20:1, 9:1, 8:2, 7:3, 6:4, 1:2 and 0:1 to obtain 8 components;
f5, eluting with an organic solvent with a volume ratio of 7:3 in the step E5 to obtain an eluent, taking a methanol aqueous solution with a volume content of 40-60% as a mobile phase, taking a Zorbax PrepHT GF reversed phase preparation column with a flow rate of 12ml/min and a wavelength of 21.2mm multiplied by 25cm and 5 mu m as a stationary phase, collecting chromatographic peaks with a detection wavelength of 254nm by an ultraviolet detector for 21min, accumulating for multiple times, and evaporating to obtain the chromone compound 7-hydroxy-5- (3-hydroxy-2-oxypropyl) -2-methyl-4H-chromen-4-one.
Example 6
The preparation method of the chromone compound in the embodiment comprises the following steps:
a6, taking 11kg of dried branches, leaves and/or fruits of the cassia plant, coarsely crushing the branches, leaves and/or fruits to 20 meshes, ultrasonically extracting the branches, leaves and/or fruits with 100% ethanol for 2 times, each time for 30min, and combining the extracting solutions; filtering the extract, and concentrating under reduced pressure to 1/3 of the volume; standing, filtering out precipitate, and concentrating to 1300g of extract a;
b6, adding 1300g of water into the extract a, extracting 3 times by chloroform with the same volume as the water, combining the extracted phases, and concentrating under reduced pressure to 550g of extract B;
loading 3500g of 160-mesh silica gel into a column, adding 540g of acetone into the extract b for dissolution, adding 550g of 80-mesh silica gel into the extract b, stirring, and loading the mixture into the column after stirring; gradient eluting with n-hexane-acetone mixed organic solvents with volume ratios of 1:0, 20:1, 9:1, 8:2, 7:3, 6:4, 1:2 and 0:1 respectively, collecting gradient eluates, concentrating, monitoring by TLC, and combining the same parts to obtain 8 components, wherein the 5 th component, i.e. the eluent of the n-hexane-acetone mixed organic solvent with volume ratio of 7:3 is 120g;
loading the column with a reversed phase material C-18, loading the eluent of the 5 th component obtained by eluting in the step C6 on the reversed phase column, carrying out gradient elution by using a methanol aqueous solution with the volume content of 20-100%, adopting solutions with three concentration ratios with sequentially increased methanol concentration in the gradient elution process, collecting and concentrating eluent of each part, monitoring by TLC, and merging the same parts to obtain 3 components;
e6, carrying out gradient elution on the eluent of the 1 st component in the step D6 by using a normal phase silica gel column, wherein the eluent is petroleum ether-ethyl acetate solution, and the volume ratio of the petroleum ether-ethyl acetate solution in the gradient elution process is 1:0, 20:1, 9:1, 8:2, 7:3, 6:4, 1:2 and 0:1 to obtain 8 components;
f6, eluting with an organic solvent with a volume ratio of 7:3 in the step E6 to obtain an eluent, taking a methanol aqueous solution with a volume content of 40-60% as a mobile phase, taking a Zorbax PrepHT GF reversed phase preparation column with a flow rate of 13ml/min and a detection wavelength of 254nm and 5 mu m as a stationary phase, feeding 50 mu L each time, collecting chromatographic peaks for 16min, accumulating for multiple times, and evaporating to dryness to obtain the chromone compound 7-hydroxy-5- (3-hydroxy-2-oxypropyl) -2-methyl-4H-chromen-4-one.
Example 7
The preparation method of the chromone compound in the embodiment comprises the following steps:
a7, taking 1.2kg of dried branches, leaves and/or fruits of the cassia plant, coarsely crushing the branches, leaves and/or fruits to 30 meshes, ultrasonically extracting the branches, leaves and/or fruits with 80% methanol for 4 times, each time for 30min, and combining the extracting solutions; filtering the extract, and concentrating under reduced pressure to 1/2 of the volume; standing, filtering out precipitate, and concentrating to 130g of extract a;
b7, adding 130g of water into the extract a, extracting for 4 times by using diethyl ether with the same volume as the water, combining the extracted phases, and concentrating under reduced pressure to 45g of extract B;
c7, filling a column with 450g of 180-mesh silica gel, adding 110g of acetone into the extract b for dissolution, then adding 45g of 90-mesh silica gel for sample mixing, and filling the column after sample mixing; gradient eluting with dichloromethane-acetone mixed organic solvent with volume ratio of 1:0, 20:1, 9:1, 8:2, 7:3, 6:4, 1:2, and 0:1 respectively, collecting gradient eluent, concentrating, monitoring by TLC, and mixing the same parts to obtain 8 components, wherein the 5 th component, i.e. eluent of dichloromethane-acetone mixed organic solvent with volume ratio of 7:3 is 7g;
loading the column with a reversed phase material C-18, loading the eluent of the 5 th component in the step C7 on the reversed phase column, carrying out gradient elution by using a methanol aqueous solution with the volume content of 20-100%, adopting solutions with three concentration ratios with sequentially increased methanol concentration in the gradient elution process, collecting and concentrating eluent of each part, monitoring by TLC, and merging the same parts to obtain 3 components;
e7, carrying out gradient elution on the eluent of the 1 st component in the step D1 by using a normal phase silica gel column, wherein the eluent is petroleum ether-ethyl acetate solution, and the volume ratio of the petroleum ether-ethyl acetate solution in the gradient elution process is 1:0, 20:1, 9:1, 8:2, 7:3, 6:4, 1:2 and 0:1 to obtain 8 components;
f7, eluting with an organic solvent with a volume ratio of 7:3 in the step E7 to obtain an eluent, eluting with 40-60% methanol aqueous solution to obtain a mobile phase, wherein the flow rate is 12ml/min, the flow rate is 21.2mm multiplied by 25cm, a Zorbax PrepHT GF reversed phase preparation column with a wavelength of 5 μm is used as a stationary phase, the detection wavelength of an ultraviolet detector is 254nm, 50 mu L of sample is injected each time, chromatographic peaks for 20min are collected, accumulated for multiple times, and evaporated to dryness to obtain the chromone compound 7-hydroxy-5- (3-hydroxy-2-oxypropyl) -2-methyl-4H-chromen-4-one.
Example 8
The compound 7-hydroxy-5- (3-hydroxy-2-oxoopyyl) -2-methyl-4H-chromen-4-one prepared in example 1 was taken as a pale brown powder;
the measuring method comprises the following steps: nuclear magnetic resonance, in combination with other spectroscopic techniques, is used to identify structures.
(1) Ultraviolet spectrum (methanol as solvent), lambda max (logε)：296(3.29),252(3.98)nm；
(2) Infrared spectrum (Potassium bromide tablet) v max 3419，2971，2928，1648，1622，1577，1396，1273，1160，944，840，601cm –1 ；
(3) HRESIMS shows that the molecular ion peak of the compound of the invention is m/z 249.0765[ M+H ]] + (calculated as 249.0757), combined with 13 C and C 1 The H NMR spectrum (FIGS. 1 and 2, hydrocarbon spectrum data is summarized in Table 1) shows that the molecular formula is C 13 H 12 O 5 。 1 H NMR(400MHz，CD 3 OD) and 13 CNMR(100MHz,CD 3 OD) data are shown in table 1.
HRESIMS shows that its excimer ion peak is an excimer ion peak m/z 249.0765[ M+H ]] + (calculated as 249.0757), combined with 13 C NMR spectrum to determine the molecular formula as C 13 H 12 O 5 The unsaturation was 8. Infrared spectra showed hydroxyl groups (3419 cm -1 ) Conjugated carbonyl (1648 cm) -1 ) Is not shown in the figure). At the position of 13 In C-NMR, C-4 (delta) C 179.3 The presence of conjugated carbonyl groups was also confirmed. The compound is 1 H-NMR data showed that it contained 1 methyl group (. Delta.) H 2.02 2 methylene groups (delta) H 4.54,4.94) 1 allyl hydrocarbon (. Delta H 6.03 And 1 meta-substituted phenyl [ delta ] H 7.09(1H,d,J＝2.3Hz),7.01(1H,d,J＝2.3Hz)]. In addition, the compounds 13 C-NMR and DEPT spectra (Table 1) show 13 carbon signals, including 2 carbonyl carbons (delta) C 209.15,179.28), 6 aromatic carbons (delta) C 162.57,160.67,139.27,120.63,114.87,103.42), 2 olefin carbons (delta) C 164.72,110.80), 2 methylene groups (delta) C 45.47,69.54 in which 1 is oxymethylene group) and 1 methyl group (delta) C 19.47). The above spectral data illustrate that the compound is a chromone compound, similar to the spectral data of the known compound 5-acetyl-7-hydroxy-2-methyl chromone. Except that an acetone group was attached to the C-5 position in 5-acetyl-7-hydroxy-2-methyl chromone, but a hydroxyl group was attached to the acetone group in this compound. In the HMBC spectra of the compounds, H-3 (delta H 6.03 And C-2 (delta) C 164.72)，C-10(δ C 114.87)，C-14(δ C 19.47 With a clear correlation signal, H-6 (delta) H 7.01 And C-8 (delta) C 103.42)，C-10(δ C 114.87)，C-11(δ C 45.47 Related to H-8 (delta) H 7.09 And C-6 (delta) C 120.63)，C-9(δ C 160.67)，C-10(δ C 114.87 Related to H-14 (delta) H 2.02 And C-2 (delta) C 164.72)，C-3(δ C 110.80 With obvious correlation signals to determine the connection position of methyl group as C-2, H-11 (delta) H 4.54 And C-5 (delta) C 139.27)，C-12(δ C 209.15 With a clear correlation signal, the 3-hydroxy-acetonyl group was identified as C-5,H-13 (. Delta.) H 4.94 And C-12 (delta) C 209.15 With a clear correlation signal confirming that methylene was attached to C-12. The structure of this compound was thus determined and named 7-hydroxy-5- (3-hydroxy-2-oxoopyyl) -2-methyl-4H-chromen-4-one.
Example 9
The compound prepared in example 2 was taken as a pale brown powder; the structure was determined as in example 8, with the following results: the structure is the same as that of example 8, and the molecular formula is C 13 H 12 O 5 . The compound prepared in example 2 was identified as the chromone compound 7-hydroxy-5- (3-hydroxy-2-oxoopyl) -2-methyl-4H-chromen-4-one.
Example 10
The compound prepared in example 3 was taken as a pale brown powder; the structure was determined as in example 8, with the following results: the structure is the same as that of example 8, and the molecular formula is C 13 H 12 O 5 . The compound prepared in example 3 was identified as the chromone compound 7-hydroxy-5- (3-hydroxy-2-oxoopyl) -2-methyl-4H-chromen-4-one.
The compounds of Table 1 1 H and 13 c NMR data (400/100 MHz, C) 5 D 5 N)
Example 11
The compound prepared in example 4 was taken as a pale brown powder; the structure was determined as in example 8, with the following results: the structure is the same as that of example 8, and the molecular formula is C 13 H 12 O 5 . The compound prepared in example 4 was identified as the chromone compound 7-hydroxy-5- (3-hydroxy-2-oxoopyl) -2-methyl-4H-chromen-4-one.
Example 12
The compound prepared in example 5 was taken as a pale brown powder; structure measurement was performed as in example 8, and the result was thatThe method comprises the following steps: the structure is the same as that of example 8, and the molecular formula is C 13 H 12 O 5 . The compound prepared in example 4 was identified as the chromone compound 7-hydroxy-5- (3-hydroxy-2-oxoopyl) -2-methyl-4H-chromen-4-one.
Example 13
The compound prepared in example 6 was taken as a pale brown powder; the structure was determined as in example 8, with the following results: the structure is the same as that of example 8, and the molecular formula is C 13 H 12 O 5 . The compound prepared in example 4 was identified as the chromone compound 7-hydroxy-5- (3-hydroxy-2-oxoopyl) -2-methyl-4H-chromen-4-one.
Example 14
The compound prepared in example 7 was taken as a pale brown powder; the structure was determined as in example 8, with the following results: the structure is the same as that of example 8, and the molecular formula is C 13 H 12 O 5 . The compound prepared in example 4 was identified as the chromone compound 7-hydroxy-5- (3-hydroxy-2-oxoopyl) -2-methyl-4H-chromen-4-one.
Example 15
The tobacco mosaic virus resistance activity test was performed on any one of the chromone compounds prepared in examples 1 to 7, and the test conditions were as follows:
the test host, TMV dead spot host, heart leaf smoke Nicotiana glutinosa L, TMV system infects host common smoke Nicotiana tabacum L.K326 and insect prevention greenhouse cultivation.
Test toxin source: tobacco mosaic virus (TMV, U1 strain) was stored on common smoke K326 by tobacco chemistry key laboratory of Yunnan tobacco institute.
Virus purification: with reference to the method of good et al, a slight modification is made. Typical symptomatic disease She Jingcha, PEG precipitation and discontinuous density gradient centrifugation of 10-40% sucrose followed by virus purification. The mass concentration of the purified virus is determined to be 20mg/mL by ultraviolet scanning
Purified virus was stored at-20℃and diluted to 32. Mu.g/mL with 0.01M PBS prior to use.
Inhibition of infection assay: the local spot-drying method is adopted. Test compounds were dissolved in DMSO and diluted to the desired concentration with distilled water, and ningnanmycin was used as a positive control. Healthy and vigorous 5-6 leaf stage heart leaf tobacco is selected, the left half leaf is inoculated with a mixed solution of a compound and viruses in equal volume, and the right half leaf is inoculated with distilled water (containing a small amount of DMSO) and the mixed solution of the viruses in equal volume as a negative control. After inoculation, the cells were rinsed with water. 4-5 leaves are inoculated in each treatment, the treatment is repeated for 3 times, the number of dead spots is counted after 3-4 days, and the inhibition rate is calculated. Specifically, the inhibition ratio= (number of control spots-number of treated spots)/number of control spots×100%
Test results
The chromone compound is separated for the first time, is determined to be the chromone compound by a nuclear magnetic resonance and mass spectrometry method, and characterizes the specific structure of the chromone compound. Through experiments of resisting tobacco mosaic virus, the inhibition rate reaches 28.7+/-4.1%, and the inhibition rate has good activity of resisting tobacco mosaic virus, and is similar to that of a positive control, namely naningmycin (32.2+/-3.2%). The results show that the compound has good application prospect in preparing the tobacco mosaic virus resistant medicines. The compound has simple structure and good activity, and can be used as a lead compound of a tobacco mosaic virus resistant drug.
In the description of the present specification, a description referring to terms "one embodiment," "some embodiments," "examples," "specific examples," or "some examples," etc., means that a particular feature, structure, material, or characteristic described in connection with the embodiment or example is included in at least one embodiment or example of the present invention. In this specification, schematic representations of the above terms do not necessarily refer to the same embodiments or examples. Furthermore, the particular features, structures, materials, or characteristics described may be combined in any suitable manner in any one or more embodiments or examples.
The foregoing examples illustrate only a few embodiments of the invention and are described in detail herein without thereby limiting the scope of the invention. It should be noted that it will be apparent to those skilled in the art that several variations and modifications can be made without departing from the spirit of the invention, which are all within the scope of the invention. Accordingly, the scope of protection of the present invention is to be determined by the appended claims.
Claims (6)
1. A chromone compound characterized by the following chemical structural formula:
2. a process for the preparation of the chromone compound of claim 1, comprising the steps of:
A. extracting extract: pulverizing branches, leaves or fruits of cassia plants to 20-40 meshes, ultrasonically extracting for 2-4 times by using an organic solvent for 30-60 min each time, combining the extracting solutions, filtering the extracting solutions, concentrating the extracting solutions under reduced pressure to 1/4-1/2 of the volume, standing, filtering out precipitate, and concentrating into an extract a;
B. extracting with an organic solvent: adding water into the extract a, wherein the weight of the added water is 1-2 times of that of the extract a, extracting for 3-5 times by using an organic solvent with the same volume as that of the water, combining the organic solvent extraction phases, and concentrating under reduced pressure to obtain an extract b;
C. silica gel column chromatography: dissolving the extract b by using an organic solvent with the weight of 1.5-3 times, mixing the extract b with 200-300 meshes of silica gel with the weight of 0.8-1.2 times, loading the extract b on a silica gel column for chromatography, wherein the loading silica gel is 200-300 meshes, and the loading silica gel is 6-8 times of the weight of the extract b; gradient eluting with two mixed organic solvents at a volume ratio of 1:0-0:1, collecting gradient eluent, concentrating, monitoring by TLC, and mixing the same parts;
D. reversed phase column chromatography: c, loading the eluent after the same parts are combined in the step C on a reversed phase column for chromatography, wherein the reversed phase column is filled with reversed phase materials; gradient elution is carried out by using a methanol aqueous solution with the volume content of 20-100%, three solutions with sequentially increased methanol concentration are adopted in the gradient elution process, eluent of each part is collected and concentrated, TLC monitoring is carried out, and the same parts are combined to obtain 3 components;
E. gradient eluting the 1 st component of the eluent in the step D by using a normal phase silica gel column, wherein the eluent is petroleum ether-ethyl acetate solution, and 8 petroleum ether-ethyl acetate solutions with sequentially increased ethyl acetate concentration are adopted in the gradient elution process to obtain 8 components;
F. high performance liquid chromatography separation: separating and purifying the concentrated solution of the 5 th component of the eluent in the step E through high performance liquid chromatography to obtain the chromone compound;
wherein, the organic solvent adopted in the extract extraction in the step A is 80-100% of acetone, ethanol or methanol, the organic solvent adopted in the extract extraction in the step B is chloroform, diethyl ether, petroleum ether or benzene, and the mixed organic solvent in the step C is n-hexane-acetone, chloroform-acetone or petroleum ether-acetone.
3. The method for preparing a chromone compound according to claim 2, wherein the step of separating and purifying the concentrated solution of the 5 th component of the eluent of the E step by high performance liquid chromatography comprises:
loading the concentrated solution of the 5 th component of the eluent in the step E on a high performance liquid chromatographic column, taking 45-65% methanol as a mobile phase, taking a reversed phase preparation column with the flow rate of 8-14 ml/min,21.2mm multiplied by 25cm and 5 mu m as a stationary phase, detecting the wavelength of 254nm by an ultraviolet detector, injecting 45-60 mu L each time, collecting chromatographic peaks for 15-33 min, and evaporating after accumulating for a plurality of times.
4. The method for preparing a chromone compound according to claim 2, wherein the volume ratio of the mixed organic solvent in the step C is 1:0, 20:1, 9:1, 8:2, 7:3, 6:4, 1:2, 0:1.
5. Use of a chromone compound according to claim 1 in the preparation of a medicament against tobacco mosaic virus.
6. The use of a chromone compound according to claim 5 for the preparation of a tobacco mosaic virus resistant drug comprising said chromone compound as the sole active ingredient.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202110385027.7A CN113185484B (en) | 2021-04-09 | 2021-04-09 | Chromone compound and preparation method and application thereof |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202110385027.7A CN113185484B (en) | 2021-04-09 | 2021-04-09 | Chromone compound and preparation method and application thereof |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN113185484A CN113185484A (en) | 2021-07-30 |
| CN113185484B true CN113185484B (en) | 2023-05-02 |
Family
ID=76975388
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN202110385027.7A Active CN113185484B (en) | 2021-04-09 | 2021-04-09 | Chromone compound and preparation method and application thereof |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN113185484B (en) |
Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN109265423A (en) * | 2018-11-20 | 2019-01-25 | 云南民族大学 | A kind of chromone compounds and its preparation method and application |
-
2021
- 2021-04-09 CN CN202110385027.7A patent/CN113185484B/en active Active
Patent Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN109265423A (en) * | 2018-11-20 | 2019-01-25 | 云南民族大学 | A kind of chromone compounds and its preparation method and application |
Also Published As
| Publication number | Publication date |
|---|---|
| CN113185484A (en) | 2021-07-30 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| ITO et al. | Constituents of Murraya exotica L. Structure elucidation of new coumarins | |
| CN104945360A (en) | Preparation method and application of phenylpropanoid compound in tobacco | |
| CN104387402A (en) | Isocoumarin compound and preparation method and use thereof | |
| CN105175239A (en) | Sesquiterpenoid compound capable of inhibiting activity of tobacco mosaic virus in tobacco, preparation method and applications thereof | |
| CN106831365B (en) | Hydroxyl methoxy substituted biphenyl compound and preparation method and application thereof | |
| Glombitza et al. | Phlorotannins from the brown algae Cystophora torulosa and Sargassum spinuligerum | |
| CN104292202B (en) | A kind of flavonoid compound and its preparation method and application | |
| Peres et al. | Naturally occurring pentaoxygenated, hexaoxygenated and dimeric xanthones: a literature survey | |
| CN113185484B (en) | Chromone compound and preparation method and application thereof | |
| CN103232427B (en) | Xanthone compound as well as preparation method and application thereof | |
| CN110357894B (en) | Tricyclic alkaloid compound and preparation method and application thereof | |
| CN111205302B (en) | Litsea pungens fruit extract, extraction method, preparation method and application | |
| CN104974122A (en) | Coumarin compound originated from tobacco, and preparation method and application thereof | |
| CN111560016A (en) | Method for extracting and separating coptisine from celandine | |
| CN104387361B (en) | A kind of Isocoumarin compounds and its production and use | |
| CN103626812B (en) | Gloomy glycosides compound of a kind of new Bali and uses thereof in rhizoma Gastrodiae | |
| CN113717184B (en) | Quinoline alkaloid with tobacco mosaic virus resisting activity in cigar and preparation method and application thereof | |
| CN113105471B (en) | Phenolic acid compound with antioxidant activity and application thereof | |
| CN113200911B (en) | Quinoline alkaloid compound and preparation method and application thereof | |
| CN114436802A (en) | Cadinane sesquiterpene compound and preparation method and application thereof | |
| CN103012425B (en) | Benzofuran compound, and preparation method and application thereof | |
| CN113149950A (en) | Dalbergia odorifera lactone compound and preparation method and application thereof | |
| CN109456163B (en) | Cycloalkenone compound with symmetrical structure and preparation method and application thereof | |
| CN110372707B (en) | Benzazepine alkaloid compound and preparation method and application thereof | |
| CN104650053A (en) | Flavonoids compound with novel structure, as well as preparation method and applications thereof |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| PB01 | Publication | ||
| PB01 | Publication | ||
| SE01 | Entry into force of request for substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| GR01 | Patent grant | ||
| GR01 | Patent grant |