CN106083874B - A kind of preparation and use of isoflavonoid - Google Patents
A kind of preparation and use of isoflavonoid Download PDFInfo
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- CN106083874B CN106083874B CN201610405184.9A CN201610405184A CN106083874B CN 106083874 B CN106083874 B CN 106083874B CN 201610405184 A CN201610405184 A CN 201610405184A CN 106083874 B CN106083874 B CN 106083874B
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- C07—ORGANIC CHEMISTRY
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- C07D493/00—Heterocyclic compounds containing oxygen atoms as the only ring hetero atoms in the condensed system
- C07D493/02—Heterocyclic compounds containing oxygen atoms as the only ring hetero atoms in the condensed system in which the condensed system contains two hetero rings
- C07D493/04—Ortho-condensed systems
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Abstract
The invention belongs to medicine and field of natural product chemistry, and in particular to a kind of new isoflavonoid and preparation method thereof and medical application.Isoflavonoid provided by the present invention has following general structure：In formula, R1、R2=H；(CH2)nCH3；OR’（R’=H；(CH2)nCH3；1 ~ 5 glycosyl）；1 ~ 5 glycosyl, R3、R4=H；(CH2)nCH3；OR’（R’=H；(CH2)nCH3；1 ~ 5 glycosyl）, the glycosyl is glucosyl group or celery glycosyl, n=0 ~ 3.The isoflavonoid of the present invention has significant antioxidation activity.
Description
Technical field
The invention belongs to medicine and field of natural product chemistry, and in particular to a kind of new isoflavonoid and its preparation
Method and medical application.
Background technology
Mung bean is the dry mature seed of legume mung bean (Vigna radiate L.), has clearing heat and detoxicating, eliminating dampness
The effect of.Available for vomit and diarrhoea, the treatment of the disease such as macula, furunculosis, mange.《Collected statements on the herbal foundation》Described in mung bean treat serious case of furuncle,
Ecchymosis, all poison of gold, stone, pellet, fire and cholera are told down.At present, there are not the pharmacological activity of Mung Bean Plant and the correlation of active material
Report.
Isoflavonoid is in plant kingdom's generally existing, but epoxy isoflavonoid is rarely reported.At present, from green
The new isoflavone compounds found in beans plant are 2,3- epoxies -2 '-hydroxyl Dai structure, are the new chemical combination not yet reported
Thing.
In recent years, the filler such as silica gel and ODS Natural Medicine Chemistry composition extracting and developing purifying, preparation process reform,
Quality of the pharmaceutical preparations analysis etc. has wide application study, and is explicitly shown out its unique effect.Utilize silica gel and ODS
Opposite adsorption function and material similar compatibility principle can from Chinese medicine extract separation and purification active ingredient or effective portion
Position, discards the dross and selects the essential, promotes the development of modernization of cmm to greatest extent.
Free radical is in signal transduction and immune etc. has the function that important but excessive active oxygen radical will produce
Raw destruction, by the biomolecule in chain reaction attack cells or in body fluid, such as DNA, lipid, protein, causes
Human normal cell and the damage of tissue, so as to cause a variety of diseases, such as cancer, aging and various angiocardiopathies.It is same with this
When, there is the synthetized oxidation preventive agent of free radical scavenging activity or natural to have turned into the research of domestic and international each ambit
Focus, wherein the research to isoflavonoid is the popular domain in free radical scavenger research.
Isoflavones and its glycoside are a major class native chemical products, widely distributed in plant kingdom, and that is extracted from plant should
Class chemical product has a variety of physiologically actives such as protection cardiovascular and cerebrovascular, anti-inflammatory, anti-oxidant, and it is the chief active of many Chinese herbal medicines
Composition, many of which are developed to novel plant medicine, such as isoflavones, obtain good result, but researcher still exists
Constantly explore, in the hope of obtaining the active ingredient of such more compound, meet the needs of people are increasing to Chinese herbal medicine.
The content of the invention
It is an object of the present invention to provide a kind of new isoflavonoid.
It is another object of the present invention to provide the preparation method of above-mentioned isoflavonoid.
It is yet a further object of the present invention to provide the purposes of the compound.
Isoflavonoid provided by the present invention, in isoflavones C2, C3 positions, double bond is at least partially epoxidized, and generates the different Huang of epoxy
Ketone, i.e. 2,3- epoxy-isoflavonoid, and there is following general structure：
In formula, R1、R2=H；(CH2)nCH3；OR ' (R '=H；(CH2)nCH3；1~5 glycosyl)；1~5 glycosyl R3、R4
=H；(CH2)nCH3；OR ' (R '=H；(CH2)nCH3；1~5 glycosyl)
The glycosyl is glucosyl group or celery glycosyl
N=0~3
Preferably, R1=H；R2、R3、R4=OR ' (R '=H)
The present invention provides the structure and title of one of isoflavonoid, i.e. 2,3- epoxies -2 '-hydroxyl soybean
Element, English entitled 2,3-epoxy-2'-hydroxydaidzein, its concrete structure formula is as follows：
The compound has following spectroscopic properties：As shown in table 1.
- 2 '-hydroxyl Dai of compound 2,3- epoxies of table 1.1H-NMR and13C-NMR and HMBC modal datas
FT-ICR-MS data：For the compound, its high resolution mass spectrum provides molecular ion peak (negative
ion mode):M/z=285.04021 [M-H]-(calcd.for C15H9O6,285.04046,err:0.9ppm), molecular formula：
C15H10O6。
Isoflavonoid provided by the present invention is to separate to prepare from Mung Bean Plant, its preparation method specific steps
It is as follows：
(1) the drying stem branch of Mung Bean Plant is taken, is extracted with ethanol or methanol, filtering, merging filtrate, obtains alcohol extract；By alcohol
Alcohol is recovered under reduced pressure to without alcohol taste in extract, obtains crude extract；
(2) crude extract obtained is extracted with opposed polarity organic solvent successively, discards extract, then extracted with n-butanol
Take, merge butanol extraction liquid, n-butanol is recovered under reduced pressure, obtains Mung Bean Plant extract；
(3) the Mung Bean Plant extract obtained by, is disperseed with proper amount of methanol, is added proper silica gel, is stirred, and is loaded advance
In the silicagel column handled well, the mixed organic solvents (100 that are mixed to get with two kinds of solvents:0~0:100) gradient elution is carried out,
Collect mixed organic solvents (50:50~30:70) eluent, is concentrated into thick, adds proper amount of methanol to disperse；Upper open ODS posts,
With alcohol-water (0:100~100:0), preferably alcohol-water (25:75~40:60) gradient elution is carried out, collects alcohol-water (30:70~
35:65) eluent, it is concentrated to obtain mung bean isoflavonoid；
(4) the mung bean isoflavonoid obtained by, purified with preparing thin-layer chromatography or preparing RPLC,
Obtain 2,3- epoxies -2 '-hydroxyl Dai.
The concentration of ethanol or methanol described in step (1) is 30%~100%.
Extraction described in step (1) is 30%~100% ethanol or methanol homogenate extraction 1~3 with 5~20 times of amounts
It is secondary, and merge extract solution.
Opposed polarity organic solvent described in step (2) includes, but are not limited to：Petroleum ether, hexamethylene, dichloromethane,
Chloroform, ethyl acetate.
The dosage of opposed polarity organic solvent, n-butanol described in step (2) is 0.5~2 times of volume, and extraction times are
1~5 time.
Mixed organic solvents described in step (3) are methylene chloride-methanol, or chloroform-methanol, or dichloromethane-second
Alcohol, or chloroform-ethanol.
Alcohol-water described in step (3) is methanol-water or alcohol-water.
The condition for preparing thin-layer chromatography described in step (4) is：Methylene chloride-methanol (30:1~10:Or dichloromethane 1)
Alkane-ethanol (30:1~10:Or chloroform-methanol (30 1):1~10:Or chloroform-ethanol (30 1):1~10:1).
The condition for preparing RPLC described in step (4) is：Octadecylsilane chemically bonded silica (ODS)
For the chromatographic column of filler, using methanol-water or alcohol-water as mobile phase, gradient elution.
In the present invention, first using positive (silica gel) pillar layer separation, then using anti-phase (open ODS) pillar layer separation,
Prepare the mung bean isoflavones containing the compound；Finally using thin-layer chromatography or preparation is prepared, instead efficiently the progress of phase liquid chromatogram is pure
Change, obtain compound 2,3- epoxies -2 '-hydroxyl Dai.
In the present invention, when being separated using silica gel column chromatography, use a dry method on a sample；During using open ODS pillar layer separations, adopt
With wet method loading.
In the present invention, the pharmacological action of described isoflavonoid is that have significant antioxidation activity.
In the present invention, resulting isoflavonoid has significant Scavenging ability, pre- suitable for preparing
Medicine that is anti-and treating the various diseases caused by interior free yl produces the decline of excessive or Scavenging ability, such as the heart
Popular name for, senile dementia, tumour and aging etc., be particularly suitable for use in prevention and treatment disease of cardiovascular system.And its preparing raw material
Wide material sources, cost is cheap, there is very gratifying application prospect.
Brief description of the drawings
Fig. 1 is the hydrogen spectrogram of 2,3- epoxies -2 '-hydroxyl Dai.
Fig. 2 is the carbon spectrogram of 2,3- epoxies -2 '-hydroxyl Dai.
Fig. 3 is the HMBC spectrograms of 2,3- epoxies -2 '-hydroxyl Dai.
Fig. 4 is the extraction separation process figure of -2 '-hydroxyl Dai of epoxy of the embodiment of the present invention 2,3.
Embodiment
The present invention is further illustrated with specific embodiment below, but is not intended to limit the present invention.
The preparation of embodiment one, 2,3- epoxies -2 '-hydroxyl Dai
1. vegetable material：The drying stem branch of legume mung bean (Vigna radiate L.).
2. method for separating and preparing：Mung Bean Plant dries stem branch 25kg, with the 95% ethanol homogenate extraction 2 times of 10,10 times of amounts,
Filtering, merging filtrate, ethanol is recovered under reduced pressure to without alcohol taste, obtains concentrate about 5L；Respectively with petroleum ether 5L, dichloromethane 5L, just
Butanol 5L respectively extraction 3 times, merge butanol extraction liquid, n-butanol are recovered under reduced pressure, obtains Mung Bean Plant extract；Mung Bean Plant is carried
Thing is taken, is disperseed with proper amount of methanol, and takes proper silica gel to mix sample；The sample mixed is fitted into silicagel column, using dichloromethane-first
Alcohol (100:0~0:100) gradient elution, 6 flow points are obtained, takes (the methylene chloride-methanol (45 of flow point 4:55~35:65)), pass through
Open ODS posts, methanol-water (25:75~40:60) gradient elution obtains 7 flow points, takes (the methanol-water (32.5 of flow point 4:67.5)),
The compound is obtained through preparing thin-layer chromatography, prepares the condition of thin-layer chromatography：Stationary phase is silica gel H；Solvent be dichloromethane-
Methanol (20:1).
The antioxidation activity in vitro test of embodiment two, 2,3- epoxies -2 '-hydroxyl Dai
1. instrument：U-1900 types spectrophotometer (Hitachi), AB135-S types electronic balance (Mettler Toledo),
ELIASA (BIO-RAD), pH-2TC (0.01 grade) precise digital display acidometer (the letter Instrument Ltd. in Shanghai).
2. reagent：Sodium chloride (NaCl), potassium chloride (KCl), potassium dihydrogen phosphate (KH2PO4), dipotassium hydrogen phosphate (K2HPO4)、
Potassium peroxydisulfate (K2S2O8), it is 2,2 '-azine group-bis--(3- ethyl benzo thiazoline quinoline -6- sulfonic acid) di-ammonium salts (ABTS), anti-bad
Hematic acid.
Experiment 2,3- epoxies -2 '-hydroxyl Dai used is made by oneself for inventor, and its chemical constitution is composed through hydrogen, carbon spectrum, matter
Spectrum determination is errorless, and HPLC purity is more than 98%.
3. experimental method：With pH7.4 phosphate buffer solution (PBS) prepare 7mmol/L ABTS solution and
2.45mmol/L potassium persulfate solution, two kinds of solution are mixed in equal volume, and at room temperature, in dark place reaction 12h, derive from
By base cation (ABTS·+·) solution.Then 40 times, avoid light place 30min after dilution are diluted with pH7.4 PBS solution,
It is 0.71 (test that can be used for antioxidation activity) that absorbance is determined at 734nm.It is big that 2,3- -2 '-hydroxyls of epoxy are prepared with methanol
The series concentration solution of legumin and ascorbic acid, concentration be respectively 6mmol/L, 3mmol/L, 1.5mmol/L, 0.3mmol/L and
0.15mmol/L.By each concentration test sample 100 μ L and 3mL ABTS·+·Avoid light place 6min at room temperature, existed with ELIASA
Absorbance A is determined under 734nmS.Negative control is replaced with methanol, and absorbance A is determined under 734nm with ELIASAC。ABTS·+Clearance rate (%) calculation formula is as follows：
ABTS·+Clearance rate (%)=(1-As/Ac) × 100%
With the software calculation of half inhibitory concentration (IC of SPSS 17.050)。
4. test result：It is shown in Table 2.
- 2 '-external ABTS of hydroxyl Dai of compound 2,3- epoxies of table 2.·+Remove result
The invention test result shows that compound 2,3- epoxies -2 '-hydroxyl Dai has ABTS+ inhibitory activity, and
Activity is better than vitamin C.Therefore the compound has significant Scavenging ability, suitable for preparing treatment due to interior free yl
Produce excessive or Scavenging ability decline caused by various diseases medicine, such as heart disease, senile dementia, tumour and
Aging etc., be particularly suitable for use in treatment disease of cardiovascular system, has good application, DEVELOPMENT PROSPECT.
Claims (7)
1. isoflavonoid, it is characterised in that there is following structural formula：
2,3- epoxies -2 '-hydroxyl Dai.
2. a kind of preparation method of compound as claimed in claim 1, it is characterised in that comprise the following steps：
(1) preparation of Mung Bean Plant crude extract：Take Mung Bean Plant appropriate, extracted with ethanol or methanol, filtering, merging filtrate, obtained
Alcohol extract, alcohol is recovered under reduced pressure to without alcohol taste in alcohol extract, obtains crude extract；
(2) preparation of Mung Bean Plant extract：Mung Bean Plant crude extract is taken, is extracted, discarded with opposed polarity organic solvent successively
Extract, then with extracting n-butyl alcohol, merge butanol extraction liquid, n-butanol is recovered under reduced pressure, obtains Mung Bean Plant extract；
(3) preparation of mung bean isoflavonoid：The Mung Bean Plant extract of gained, is disperseed with proper amount of methanol, is added appropriate
Silica gel, stir, be fitted into the silicagel column anticipated, the mixed organic solvents 100 being mixed to get with two kinds of solvents:0
~0:100 carry out gradient elution, collect mixed organic solvents 50:50~30:70 eluents, are concentrated into thick, add appropriate first
Alcohol disperses；Upper open ODS posts, with alcohol-water 0:100~100:0 carries out gradient elution, collects alcohol-water 30:70~35:65 elutions
Liquid, it is concentrated to obtain mung bean isoflavonoid；
(4) preparation of epoxy isoflavone compounds：The mung bean isoflavonoid obtained, through preparing thin-layer chromatography or preparation
RPLC purifies, and obtains 2,3- epoxies -2 '-hydroxyl Dai；
In step (2), after first being extracted 1~5 time with 0.5~2 times of volume petroleum ether or hexamethylene, then with 0.5~2 times of volume dichloro
Methane or chloroform or ethyl acetate extraction 1~5 time, finally with the extracting n-butyl alcohol 1~5 time of 0.5~2 times of volume；
Mixed organic solvents gradient elution described in step (3), its mixed organic solvents are methylene chloride-methanol, or chloroform-
Methanol, or dichloromethane-ethanol, or chloroform-ethanol, in described alcohol-water gradient elution, its alcohol-water is methanol-water or second
Alcohol-water；
In step (4), the condition for preparing thin-layer chromatography is：Lamellae is silica gel H or silica G plate；Solvent is dichloromethane-first
Alcohol (30:1~10:, or dichloromethane-ethanol (30 1):1~10:, or chloroform-methanol (30 1):1~10:, or chloroform-second 1)
Alcohol (30:1~10:1) condition for, preparing RPLC is：Chromatogram column packing is octadecylsilane bonded silica
Glue, using methanol-water or alcohol-water as mobile phase, gradient elution.
3. preparation method as claimed in claim 2, it is characterised in that the 30% of middle 5~20 times of amounts of use of step (1)~
100% ethanol or methanol extract 1~3 time, and merge extract solution.
4. preparation method as claimed in claim 2, it is characterised in that in step (3), upper open ODS posts, with alcohol-water 25:75
~40:60 carry out gradient elution.
5. a kind of pharmaceutical composition, include the isoflavonoid and pharmaceutically acceptable carrier described in claim 1.
6. the pharmaceutical composition described in isoflavonoid or claim 5 described in claim 1 is being prepared for preventing
With treatment because interior free yl produces the application caused by excessive or Scavenging ability declines in the medicine of various diseases.
7. application as claimed in claim 6, it is characterised in that described because interior free yl produces excessive or removes freely
Various diseases caused by the decline of base ability are selected from：Heart disease, senile dementia, tumour or aging disease.
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| CN110108831B (en) * | 2019-06-14 | 2022-04-01 | 山东师范大学 | Thin-layer chromatography developing agent for soybean isoflavone glycoside and detection method |
| CN115429810B (en) * | 2021-06-03 | 2024-03-08 | 苏州禾研生物技术有限公司 | Application of isoflavone compound in preparing medicament for preventing or treating alcoholic liver injury or dispelling alcohol effect and protecting liver |
Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103804443A (en) * | 2014-01-16 | 2014-05-21 | 沈阳药科大学 | Flavonoid glycoside compound and preparation method thereof |
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Patent Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103804443A (en) * | 2014-01-16 | 2014-05-21 | 沈阳药科大学 | Flavonoid glycoside compound and preparation method thereof |
Non-Patent Citations (2)
| Title |
|---|
| Dimethyldioxirane epoxidation of aurones and isoflavones;Waldemar Adam，等;《Synthesis》;19920531(第5期);第436页化合物4b-4c * |
| Enantioselective epoxidation of isoflavones using Jacobsen"s Mn(III)salen catalysts and dimethyldioxirane as the oxygen-atom source;Waldemar Adam，等;《Tetrahedron: Asymmetry》;19981231;第9卷(第7期);第1122页化合物2a-2b，2d-2e * |
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