CN108358878A - Isoflavones noval chemical compound and its preparation method and application in big fruit banyan - Google Patents
Isoflavones noval chemical compound and its preparation method and application in big fruit banyan Download PDFInfo
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- CN108358878A CN108358878A CN201810022626.0A CN201810022626A CN108358878A CN 108358878 A CN108358878 A CN 108358878A CN 201810022626 A CN201810022626 A CN 201810022626A CN 108358878 A CN108358878 A CN 108358878A
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- C07—ORGANIC CHEMISTRY
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- 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
- C07D311/26—Benzo[b]pyrans, not hydrogenated in the carbocyclic ring with oxygen or sulfur atoms directly attached in position 4 with aromatic rings attached in position 2 or 3
- C07D311/34—Benzo[b]pyrans, not hydrogenated in the carbocyclic ring with oxygen or sulfur atoms directly attached in position 4 with aromatic rings attached in position 2 or 3 with aromatic rings attached in position 3 only
- C07D311/36—Benzo[b]pyrans, not hydrogenated in the carbocyclic ring with oxygen or sulfur atoms directly attached in position 4 with aromatic rings attached in position 2 or 3 with aromatic rings attached in position 3 only not hydrogenated in the hetero ring, e.g. isoflavones
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- A61P35/00—Antineoplastic agents
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- 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
- C07D311/26—Benzo[b]pyrans, not hydrogenated in the carbocyclic ring with oxygen or sulfur atoms directly attached in position 4 with aromatic rings attached in position 2 or 3
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Abstract
The invention discloses a kind of preparation method and applications of isoflavones noval chemical compound in big fruit banyan, preparing for the isoflavones noval chemical compound is as follows：It by big fruit banyan root grinding and sieving, is extracted with alcoholic solution cold soaking, medicinal extract is obtained after reduced pressure, distillation aqueous suspension is added in medicinal extract, extracts to obtain each position medicinal extract with petroleum ether, ethyl acetate successively；Ethyl acetate extract obtains isoflavones noval chemical compound through column chromatography, efficient liquid phase.The isoflavones noval chemical compound that the present invention is extracted from big fruit banyan, which has, preferably inhibits three kinds of tumor cell proliferation effects.
Description
Technical field
The present invention relates to a kind of natural plant extracts, and in particular in a kind of big fruit banyan isoflavones noval chemical compound and its
Preparation method and application.
Background technology
Big fruit banyan（Ficus auriculatalour）For Moraceae Ficus（Ficus）Plant, alias steamed bun fruit, fruit taste
Sweet tea is edible, is distributed mainly on the torrid zone and subtropical zone, Hainan, Guangxi, Yunnan, Guizhou, Sichuan etc. are mainly distributed in China
Ground,《Southern herbal garden plant list》Record big fruit banyan fruit with wind-dispelling facilitaing lung, tonifying kidney and benefiting sperm effect, cure mainly cough with lung heat, seminal emission,
It spits blood.Big fruit banyan chemical composition and bioactivity research are shown in big fruit banyan containing the ingredients such as isoflavones in recent years and with compared with
Good bioactivity.Qi Cuicui etc.（A new isoflavone from the roots ofFicus auriculata[J].
Natural Product Research, 2018, 32(1):43）Isolated 4 have antibacterial activity from big fruit banyan root
Isoflavone compound.Tan Lin etc.（Big fruit banyan fruit ethanol extract antioxidant activity and rightαGlucuroide and acetyl
Cholinesterase inhibition [J] Food Sciences, 2016. 37 (13): 77）To polyphenol content in big fruit banyan fruit and its resist
Oxidation,αGlucuroide and inhibiting activity of acetylcholinesterase are studied, the results showed that big fruit banyan fruit polyphenol has good
Oxidation activity well,αGlucuroide and certain inhibiting activity of acetylcholinesterase.At present still lack big fruit banyan isoflavones at
The system research divided, antitumor activity need further to study.
Invention content
Present invention aims at solving the above problems, isoflavones with anti-tumor activity new in a kind of big fruit banyan is provided
Noval chemical compound and its preparation method and application.
Realizing the technical solution of the object of the invention is：A kind of isoflavones noval chemical compound, structure such as formula（Ⅰ）It is shown：
（Ⅰ）.
The preparation method of above-mentioned isoflavones noval chemical compound includes the following steps：
1. with medicinal extract is obtained after alcoholic solution extraction concentration after big fruit banyan tissue nature is dried in the shade, is crushed, medicinal extract adds distillation aqueous suspension
Petroleum ether, ethyl acetate extract is obtained by extraction with petroleum ether, ethyl acetate afterwards；
2. 1. ethyl acetate extract that step obtains is obtained 7 component A-G through silica gel column chromatography, component C is again through petroleum ether-second
Acetoacetic ester system gradient elution merges similar fraction and obtains 5 component Fr1-5；
3. 2. Fr3 that step is obtained is through the isolated isoflavones noval chemical compound of efficient liquid phase.
In above-mentioned steps 1. described in big fruit banyan be organized as big fruit banyan root, stem, leaf or fruit, alcoholic solution is that volume fraction is
The ethanol solution of 75%-95%, the distilled water are 1 with medicinal extract volume ratio:1.
It is 100 that 2. the agent of center pillar chromatographic elution, which is petroleum ether-ethyl acetate volume ratio, in above-mentioned steps:0-0:100.
Above-mentioned steps 3. described in efficient liquid phase dicyandiamide solution be -55% water of 45% methanol.
Application of the above-mentioned isoflavones noval chemical compound on inhibiting tumour cell, the tumour cell are that cervical cancer Hela cells are thin
Born of the same parents, MCF-7 Breast Cancer Cell or Lung Adenocarcinoma A 549 Cell.
The good effect that the present invention has：Osajin chemical combination with anti-tumor activity new in a kind of big fruit banyan is provided
Object and preparation method thereof.
Description of the drawings
Fig. 1 is the isoflavone compounds of the present invention1H-NMR is composed（Nuclear magnetic resonance spectroscopy）Figure.
Fig. 2 is the isoflavone compounds of the present invention13C-NMR is composed（Carbon-13 nmr spectra）Figure.
Fig. 3 is the hsqc spectrum figure of the isoflavone compounds element of the present invention.
Fig. 4 is the HMBC spectrograms of the isoflavone compounds of the present invention.
Fig. 5 is the isoflavone compounds of the present invention1H-1H COSY spectrums（Hydrogen hydrogen Correlated Spectroscopy）Figure.
Fig. 6 is the HRESIMS of the isoflavone compounds of the present invention（High-resolution electrospray ionization mass spectrometry）Spectrogram.
Specific implementation mode
Preparation method of the present invention is further illustrated with reference to specific embodiment, but and the scope of the present invention
It is not limited to this.
（Embodiment）
The structural formula of the isoflavone compound of the present embodiment such as formula（Ⅰ）It is shown：
（Ⅰ）.
The preparation method of the isoflavone compound includes the following steps：
1. the big fruit banyan root grinding and sieving that 20kg is dried in the shade naturally, ethyl alcohol soak extraction 3 at room temperature for being 95% with volume fraction
It is secondary, 6 days every time, recycling ethyl alcohol is concentrated under reduced pressure and obtains total medicinal extract 460g, isometric distillation aqueous suspension medicinal extract is added, first uses 3L stones
Oily ether extracts 3 times, obtains petroleum ether part medicinal extract 175g, is repeatedly extracted with 3L ethyl acetate until upper solution color is apparent afterwards
It is thin out to obtain ethyl acetate extract 70g.
2. by step 1. in ethyl acetate extract 70g petroleum ether-ethyl acetates（100:0-0:100, V/V）Gradient
Elution, flow point is collected per 500mL, and the detection of TCL contact plates merges similar compositions and obtains 7 component A-G；Component C is again through petroleum ether-
Ethyl acetate (3:1, V/V) column chromatography, the detection of TCL contact plates merge similar compositions and obtain 5 component Fr1-5；
3. by step 2. in Fr3 through efficient liquid phase (methanol/water, 45:55, 3 mL/min, tR=17.6 min) it detaches
To 5.6 mg of isoflavones noval chemical compound.
（Test case）
Product analysis is carried out to the isoflavone compound that embodiment obtains：
（1）The isoflavone compounds1H-NMR spectrums are shown in Fig. 1（Solvent acetone-d 6, 400Hz）.
（2）The isoflavone compounds13C-NMR spectrums are shown in Fig. 2（Solvent acetone-d 6, 100MHz）.
Specific data are shown in Table 1.
Nuclear magnetic resonance data (400/100 MHz, acetone- of 1 Ficaurisoflavone A of tabled 6)
（3）The hsqc spectrum of the isoflavone compounds is shown in Fig. 3（Solvent acetone-d 6）.
（4）The HMBC spectrums of the isoflavone compounds are shown in Fig. 4（Solvent acetone-d 6）.
（5）The isoflavone compounds1H-1H COSY spectrums（Hydrogen hydrogen Correlated Spectroscopy）See Fig. 5（Solvent acetone-d 6）.
（6）The high-resolution electrospray ionization mass spectrometry figure of the isoflavone compounds is shown in Fig. 6.
Isoflavones noval chemical compound：Pale yellow powder is soluble in acetone, methanol; m.p. 157-158 ℃; [α]25 D +
26.5 (c 0.1, CH3OH); UV (CH3OH) λ max (logε): 260 (2.52) nm, 201 (3.56) nm。
It is C that HRESI-MS, which provides the compound molecule formula,20H18O6（m/z 355.11771 [M + H]+, calculated value is
355.11761）, degree of unsaturation 12.In UV spectrum, there is absorption at 260 nm, illustrate that the compound contains phenyl ring, prompts
The compound may be flavone compound.1Low field area provides 1 typical position flavones C-5 and intramolecular carbonyl in H H NMR spectroscopies
Base forms the hydroxyl signal of hydrogen bondδ H12.99 (1H, s), 1 positions typical isoflavones C-2 alkene hydrogen signalδ H 8.20
(1H, s), 1 positions C-6 alkene hydrogenδ H6.28 (1H, d, J=2.2 Hz), 1 positions C-8 alkene hydrogenδ H6.42 (1H, d,J=2.2 Hz), 3 group 1, the phenyl ring hydrogen signal of 3,4 substitutionsδ H 7.36 (1H, dd, J = 8.3, 2.2 Hz)、7.30
(1H, d, J=2.2 Hz) and 6.91 (1H, d,J=8.3 Hz), further prove that the compound is osajin
Object is closed, in addition,12 cis- alkene hydrogen signals are given in H H NMR spectroscopiesδ H 6.44 (1H, J=9.8 Hz) and 5.77 (1H,J
=9.8 Hz), 2 identical methyl signalsδ H1.43 (6H, s)；13Provided in C H NMR spectroscopies 1 four substituted benzene ring (δ C
165.5,163.9,159.1,106.1,100.0,94.6), 11,3,4- substituted benzene ring (δ C 154.0, 130.6,
128.0,123.8,122.0,116.8), 1α, βBeta-unsaturated ketone (δ C 181.5,154.5,124.5), information above is being just
Meet the skeleton of 15 carbon of isoflavones well, in addition,13Given in C H NMR spectroscopies 2 olefinic carbon signals (δ C 132.0,122.8), 2
A identical methyl carbon signal (δ C 28.3).It can speculate that the compound is the substituted different Huang in the positions C-3' in conjunction with information above
Ketone compounds.The connection type of cis-double bonds is determined by HMBC, in HMBC spectrumsδ H 6.44 (1H, J = 9.8
Hz) and 5.77 (1H,J=9.8 Hz) all withδ C 122.0 (C-3') are related, illustrate that double bond is connected to C-3', 2 methylδ H1.43 (6H, s) all withδ C 132.0 (C-2'') are related to 77.2 (C-3''), illustrate that 2 methyl are connected on C-3''.
In conclusion the compound is new isoflavone compounds, it is named as：Ficaurisoflavone A.
The structural formula of Ficaurisoflavone A
The HMBC of Ficaurisoflavone A is related
（Test example, anti tumor activity in vitro evaluation）
Test method：The isolated isoflavone compounds of Example carry out antitumor activity.
Tumor cell line：Cervical cancer Hela cells, MCF-7 Breast Cancer Cell, Lung Adenocarcinoma A 549 Cell（University Of Hebei's pharmacy
Institute provides）.
Positive control：Adriamycin.
Instrument：Refrigerator, autoclave, microplate reader（BioTek ELx800）, microscope, freeze drier, CO2Constant temperature trains case,
96 orifice plates, pipettor, electric heating constant-temperature blowing drying box, microstrainer and 0.22 μm of filter membrane, supercentrifuge.
Reagent：RPMI1640 culture solutions, pancreatin, MTT, DMSO, top grade newborn bovine serum.
Specific experiment method：
(1) inoculating cell：With the culture solution containing 10% fetal calf serum（DMEM）It is made into individual cells suspension, with every hole 5 × 104、5
×105A/mL is inoculated into 96 orifice plates, and per 100 μ L of pore volume, attached cell shifts to an earlier date 12 hours inoculated and cultureds.
(2) testing compound solution is added（40 μ g/mL primary dcreening operations of fixed concentration inhibit growth of tumour cell in the concentration
Compound near 50% sets 5 concentration and enters gradient secondary screening, i.e. detectable concentration is respectively 5,1,0.2,0.04,0.008 μ g/
ML is diluted sample to be tested with culture solution）100 μ L, per 100 μ L of hole final volume, each processing is all provided with 3 multiple holes.
(3) it develops the color：After 37 degrees Celsius are cultivated 48 hours, add 20 μ L of MTT solution per hole.Continue to be incubated 4 hours, terminates training
It supports, culture supernatant in hole is abandoned in suction, and the DMSO solution of 150 μ L is added per hole, crystal is made fully to melt.
(4) colorimetric：Select 490nm wavelength, microplate reader（BioTek ELx800）Each hole absorbance value is read, result is recorded.
Calculate the IC of compound50Value.
Data analysis：
Make ordinate with cell inhibitory rate, the logarithm of concentration is abscissa, makees regression curve, calculates IC50Value.Wherein, inhibit
Rate=（Add the OD values of OD values-test group of PBS control group）/ plus PBS control group OD value × 100%.
Test result is as follows for antitumor activity of compound：
2 isoflavone compounds of table inhibit tumor cell proliferation result
AdriamycinaFor positive control.
The result shows that isoflavone compounds all have three kinds of tumor cell proliferations effects of preferable inhibition.
Claims (6)
1. a kind of isoflavones noval chemical compound, structure such as formula（Ⅰ）It is shown：
（Ⅰ）.
2. the preparation method of isoflavones noval chemical compound described in claim 1, it is characterised in that include the following steps：
1. with medicinal extract is obtained after alcoholic solution extraction concentration after big fruit banyan tissue nature is dried in the shade, is crushed, medicinal extract adds distillation aqueous suspension
Petroleum ether, ethyl acetate extract is obtained by extraction with petroleum ether, ethyl acetate afterwards；
2. 1. ethyl acetate extract that step obtains is obtained 7 component A-G through silica gel column chromatography, component C is again through petroleum ether-second
Acetoacetic ester system gradient elution merges similar fraction and obtains 5 component Fr1-5；
3. 2. Fr3 that step is obtained is through the isolated isoflavones noval chemical compound of efficient liquid phase.
3. the preparation method of isoflavones noval chemical compound according to claim 2, it is characterised in that：Above-mentioned steps 1. described in
Big fruit banyan be organized as big fruit banyan root, stem, leaf or fruit, alcoholic solution is the ethanol solution that volume fraction is 75%-95%, the steaming
Distilled water is 1 with medicinal extract volume ratio:1.
4. the preparation method of isoflavones noval chemical compound according to claim 2, it is characterised in that：Above-mentioned steps 2. center pillar layer
It is 100 that analysis eluant, eluent, which is petroleum ether-ethyl acetate volume ratio,:0-0:100.
5. the preparation method of isoflavones noval chemical compound according to claim 2, it is characterised in that：Above-mentioned steps 3. described in
Efficient liquid phase dicyandiamide solution is -55% water of 45% methanol.
6. application of the isoflavones noval chemical compound described in claim 1 on inhibiting tumour cell, the tumour cell is uterine neck
Cancer Hela cells, MCF-7 Breast Cancer Cell or Lung Adenocarcinoma A 549 Cell.
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Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2014207069A1 (en) * | 2013-06-25 | 2014-12-31 | Universita' Degli Studi Di Siena | Multitarget hedgehog pathway inhibitors and uses thereof |
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| Publication number | Priority date | Publication date | Assignee | Title |
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| WO2014207069A1 (en) * | 2013-06-25 | 2014-12-31 | Universita' Degli Studi Di Siena | Multitarget hedgehog pathway inhibitors and uses thereof |
Non-Patent Citations (2)
| Title |
|---|
| NICOLE DARBOUR ET AL.: ""Isoflavones from Ficus nymphaefolia"", 《NATURAL PRODUCT RESEARCH》 * |
| STEPHEN S. NYANDORO ET AL.: ""Flavonoids from Erythrina schliebenii"", 《J. NAT. PROD.》 * |
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