CN106928170A - A kind of dihydrofuran biphenyl compound and its preparation method and application - Google Patents
A kind of dihydrofuran biphenyl compound and its preparation method and application Download PDFInfo
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- C07—ORGANIC CHEMISTRY
- C07D—HETEROCYCLIC COMPOUNDS
- C07D307/00—Heterocyclic compounds containing five-membered rings having one oxygen atom as the only ring hetero atom
- C07D307/77—Heterocyclic compounds containing five-membered rings having one oxygen atom as the only ring hetero atom ortho- or peri-condensed with carbocyclic rings or ring systems
- C07D307/78—Benzo [b] furans; Hydrogenated benzo [b] furans
- C07D307/79—Benzo [b] furans; Hydrogenated benzo [b] furans with only hydrogen atoms, hydrocarbon or substituted hydrocarbon radicals, directly attached to carbon atoms of the hetero ring
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Abstract
The invention discloses a kind of dihydrofuran biphenyl compound and its preparation method and application, described biphenyl compound molecular formula is C18H18O4, the Compound nomenclature is 2 isopropenyl, 6 methoxy 7 hydroxy (4 hydroxyphenyl) dihydrobenzofuran, with following structural formula：The preparation method is with dry Guttiferae arbor branch, leaf and/or fruit as raw material, through medicinal extract extraction, organic solvent extraction, silica gel column chromatography, high performance liquid chromatography separation step.Described application is application of the biphenyl compound in anti-rotavirus medicaments are prepared.Through anti-rotavirus activity experiment, from virazole as control, CCs of the hydroxy of 2 isopropenyl, 6 methoxy 7 (4 hydroxyphenyl) dihydrobenzofuran to rotavirus50And EC50Value is respectively 164.2 and 12.9μMol/L, it has preferable anti-rotavirus activity.The compounds of this invention simple structure activity is good, can have good application prospect as the guiding compound of anti-rotavirus medicaments.
Description
Technical field
The invention belongs to effective ingredients in plant extractive technique field, and in particular to a kind of dihydrofuran biphenyl compound and
Its preparation method and application.
Background technology
Guttiferae (Garcinia L.) about 450 kinds of the plant whole world, Asia, Africa south and Polynesia western part are produced, I
State-owned 21 kinds, it is distributed in the south such as Guangdong, Guangxi, Yunnan provinces and regions.Garcinia maingayii or natural xanthone（xanthones）
One of Main Resources of constituents, the xanthone rich in isopentene group substitution（xanthones）, this kind of constituent structure is novel more
Sample, and with extensive pharmacological activity, it is especially most representative with gambogicacid (gambogic acid), it is anti-with broad spectrum high-effect
Tumor promotion, is one of study hotspot of Antitumor Natural Products in recent years, and Chinese scholar is developing its parenteral solution for anti-swollen
The kind new medicine of knurl one.Except xanthone（xanthone）Outside class, Benzophenone class (benzophenones), bisflavones
(bioflavonoids) and the compound such as biphenyl class (biphenyls) is also the characteristic chemical constituent of graminaceous plant, it may have various
Bioactivity.In order to more effectively utilize China's Garcinia maingayii resource, the active component with DEVELOPMENT PROSPECT is therefrom found, I
Select to Garcinia maingayii carry out compared with system study of active components work.
The content of the invention
The first object of the present invention is to provide a kind of biphenyl compound；Second purpose is to provide the biphenyl class chemical combination
The preparation method of thing；3rd purpose is the application for providing the biphenyl compound in anti-rotavirus medicaments are prepared.
The first object of the present invention is achieved in that described biphenyl compound is with dry Guttiferae arbor branch
Bar, leaf or fruit are raw material, are obtained through medicinal extract extraction, organic solvent extraction, silica gel column chromatography, high performance liquid chromatography separation,
The compound molecule formula is C18H18O4, it is named as 2-isopropenyl-6-methoxy-7-hydroxy- (4-
Hydroxyphenyl)-dihydrobenzofuran, with following structural formula：
。
The second object of the present invention is achieved in that the preparation method of described biphenyl compound, is with dry
Guttiferae arbor branch, leaf and/or fruit are raw material, through medicinal extract extraction, organic solvent extraction, silica gel column chromatography, efficient liquid phase
Chromatographic isolation is obtained, specially：
A, medicinal extract are extracted：By Guttiferae arbor branch, leaf or fruit coarse crushing to 20 ~ 40 mesh, with organic solvent ultrasonic extraction 2 ~ 4
Secondary, 40 ~ 60min every time, extract solution merges；Extract solution is filtered, and during extract solution to 1/4 ~ 1/2 volume that is concentrated under reduced pressure, is stood,
Sediment is filtered, medicinal extract a is condensed into；
B, organic solvent extraction：Add weight than 1 ~ 2 times of water of amount in medicinal extract a, 3 are extracted with the organic solvent isometric with water
~ 5 times, merge organic solvent extraction phase, be concentrated under reduced pressure into medicinal extract b；
C, silica gel column chromatography：Medicinal extract b weight is dissolved than 1.5 ~ 3 times of organic solvents of amount, then 0.8 ~ 1.2 times is weighed with medicinal extract
100 ~ 200 mesh silica gel mixed samples, then go up silica gel column chromatography, dress post silica gel is 100 ~ 200 mesh, and consumption is 6 ~ 8 times of medicinal extract b weight
Amount；It is 1 with volume ratio:0~0:1 mixed organic solvents gradient elution, collects gradient eluent, concentration, is monitored through TLC, merges
Identical part；
D, reversed phase column chromatography：Will be with 7:Reversed phase column chromatography on the eluent that the organic solvent of 3 proportionings is afforded, it is anti-phase
Post is to fill post with reversed material C-8, C-18, ODS or MCI；Gradient is carried out with the methanol aqueous solution that volume content is 20 ~ 100% to wash
It is de-, collect each several part eluent and concentrate, monitored through TLC, merge identical part；
E, high performance liquid chromatography separation：The eluent that will be afforded with the methanol aqueous solution of volume content 55 ~ 80% is through efficient liquid phase
Chromatographic separation and purification, obtains final product described biphenyl compound 2-isopropenyl-6-methoxy-7-hydroxy- (4-
hydroxyphenyl)-dihydrobenzofuran；
It with 50 ~ 70% methyl alcohol is mobile phase, 10 ~ 14mL/ of flow velocity that high performance liquid chromatography separation purifying described in F, E step is
The Zorbax PrepHT GF reverse phase preparative columns of min, 21.2 × 250 mm, 5um are fixing phase, UV-detector Detection wavelength
It is 254 nm, each 45 ~ 60uL of sample introduction collects the chromatographic peak of 16 ~ 32min, is evaporated after repeatedly adding up.Obtain final product described biphenyl class
Compound 2-isopropenyl-6-methoxy-7-hydroxy- (4-hydroxyphenyl)-dihydrobenzofuran.
Biphenyl compound of the present invention is separated first, by nuclear magnetic resonance and other spectroscopic technique measure sides
Method is defined as biphenyl compound, and characterizes its concrete structure and be：
Compound 2-isopropenyl-6-methoxy-7-hydroxy- (4-hydroxyphenyl)-
Dihydrobenzofuran, is orange-yellow jelly；Ultraviolet spectra（Solvent is methyl alcohol）,λ max（logε）：570 (1.79),
266 (3.69), 226 (3.82), 204 (3.96) nm；Infrared spectrum（Pressing potassium bromide troche）V max 3432, 2924,
2852, 1630, 1503, 1450, 1374, 1243, 1173, 1096, 1054, 1027, 905, 855, 824,
591 cm–1；HRESIMS shows the compounds of this invention quasi-molecular ion peakm/z 298.1202 [M]+（Calculated value is
298.1205）, with reference to13C and1H H NMR spectroscopies（Fig. 1 and Fig. 2, carbon spectrum hydrogen modal data ownership is shown in Fig. 4）Providing its molecular formula is
C18H18O4。1H NMR（C5D5N, 500 MHz）With13C NMR（C5D5N, 125 MHz）Data, are shown in Fig. 4.
HRESIMS shows that its quasi-molecular ion peak is 298.1202 [M]+(calculated value 298.1205), with reference to13C
H NMR spectroscopy determines that molecular formula is C18H18O4, degree of unsaturation is 10.Infrared absorption spectroscopy is in 3432 cm-1 Show the presence of hydroxyl,
In 1630,1503 cm-1Show the presence of phenyl ring, ultraviolet spectra has absorption maximum to also demonstrate that phenyl ring in 204,226,266 nm
Presence.13C NMR and1H NMR modal datas show the characteristic signal of biphenyl structural.1H NMR data displays have a virtue
Fragrant protonδ H6.81 (1H, s), show the compound be have five substituted biphenyl structurals.Aromaticδ H 6.81 (1H,
S) be H-5 because shown in HMBC it and C-4 (δ C 128.8)、C-9 (δ C 118.7)、C-7 (δ C 132.1)、C-
6 (δ C150.0) and C-1 ' ' (δ C132.5) there is correlation.1One phenyl ring of contraposition substitution of H NMR data displays [δ H
7.58 (2H, d, J=8.5 Hz, H-2 ' ', H-6 ' ') and 7.33 (2H, d,J = 8.5 Hz, H-3′′,
5 ' ')], part signal for isopropenyl coumarone [δ H 3.44 (1H, dd, J = 15.2, 9.0 Hz, H-3),
3.23 (H, dd, J = 15.2, 8.6 Hz, H-3), 5.30 (1H, t, J = 8.6 Hz, H-2), 4.88 (1H,
S, H-2 '), 5.22 (1H, s, H-2 ') and 1.72 (3H, s, H-3 ')], also one methoxyl group (δ H 3.84,
3H, s, 6-OMe).In HMBC, H-3 and C-4, C-9, C-8 (δ C149.1) related, H-2 is related to C-8, C-9,
It is known that isopropenyl coumarone is connected with C-8, C-9.Methoxyl group (δ H3.84) be substituted in C-6, can be by it and C-
The related confirmation of 6 HMBC.Understand that two hydroxyls are located at C-7, C-4 ' ' position respectively according to its molecular formula.So far, the compound
Structure determined, be named as 2-isopropenyl-6-methoxy-7-hydroxy- (4-hydroxyphenyl)-
dihydrobenzofuran。
The third object of the present invention is achieved in that described biphenyl compound in anti-rotavirus medicaments are prepared
Application.Through anti-rotavirus activity experiment, from virazole as control, 2-isopropenyl-6-methoxy-7-
CCs of hydroxy- (the 4-hydroxyphenyl)-dihydrobenzofuran to rotavirus50And EC50Value is respectively
164.2 and 12.9μMol/L, it has preferable anti-rotavirus activity.The compounds of this invention simple structure activity is good, can
As the guiding compound of anti-rotavirus medicaments, there is good application prospect.
Brief description of the drawings
Fig. 1 be compound 2-isopropenyl-6-methoxy-7-hydroxy- (4-hydroxyphenyl)-
The carbon-13 nmr spectra of dihydrobenzofuran（13C NMR）.
Fig. 2 be compound 2-isopropenyl-6-methoxy-7-hydroxy- (4-hydroxyphenyl)-
The proton nmr spectra of dihydrobenzofuran（1H NMR）.
Fig. 3 be compound 2-isopropenyl-6-methoxy-7-hydroxy- (4-hydroxyphenyl)-
The main HMBC of dihydrobenzofuran（→）It is related.
Fig. 4 is the carbon spectrum hydrogen modal data ownership of compound.
Fig. 5 is the anti-rotavirus activity of compound.
Specific embodiment
The present invention is further illustrated below in conjunction with the accompanying drawings, but the present invention is any limitation as never in any form, base
In present invention teach that any conversion or improvement made, each fall within protection scope of the present invention.
Biphenyl compound of the present invention is the warp with the branch of dry Guttiferae arbor, leaf or fruit as raw material
Medicinal extract extraction, organic solvent extraction, silica gel column chromatography, high performance liquid chromatography separation are obtained, and the compound molecule formula is
C18H18O4, be named as 2-isopropenyl-6-methoxy-7-hydroxy- (4-hydroxyphenyl)-
Dihydrobenzofuran, with following structural formula：
。
The preparation method of biphenyl compound of the present invention, it is characterised in that be with dry Guttiferae arbor branch
Bar, leaf and/or fruit are raw material, are obtained through medicinal extract extraction, organic solvent extraction, silica gel column chromatography, high performance liquid chromatography separation,
Specially：
A, medicinal extract are extracted：By Guttiferae arbor branch, leaf or fruit coarse crushing to 20 ~ 40 mesh, with organic solvent ultrasonic extraction 2 ~ 4
Secondary, 30 ~ 60min every time, extract solution merges；Extract solution is filtered, and during extract solution to 1/4 ~ 1/2 volume that is concentrated under reduced pressure, is stood,
Sediment is filtered, medicinal extract a is condensed into；
B, organic solvent extraction：Add weight than 1 ~ 2 times of water of amount in medicinal extract a, 3 ~ 5 are extracted with the organic solvent isometric with water
It is secondary, merge organic solvent extraction phase, it is concentrated under reduced pressure into medicinal extract b；
C, silica gel column chromatography：Then medicinal extract b is weighed the 80 of 0.8 ~ 1.2 times than 1.5 ~ 3 times of acetone solutions of amount with weight with medicinal extract
~ 100 mesh silica gel mixed samples, then go up silica gel column chromatography, and dress post silica gel is 100 ~ 200 mesh, and consumption is 6 ~ 8 times of amounts of medicinal extract b weight；With
Volume ratio is 1:0~0:1 mixed organic solvents gradient elution, collects gradient eluent, concentration, is monitored through TLC, merges identical
Part；
D, reversed phase column chromatography：Will be with 7:Reversed phase column chromatography, reversed-phase column on the eluent that the organic solvent of 3 proportionings is afforded
It is to fill post with reversed material C-8, C-18, ODS or MCI；Gradient is carried out with the methanol aqueous solution that volume content is 20 ~ 100% to wash
It is de-, collect each several part eluent and concentrate, monitored through TLC, merge identical part；
E, high performance liquid chromatography separation：The eluent that will be afforded with the methanol aqueous solution of volume content 55 ~ 80% is through efficient liquid phase
Chromatographic separation and purification, obtains final product described biphenyl compound 2-isopropenyl-6-methoxy-7-hydroxy- (4-
hydroxyphenyl)-dihydrobenzofuran；
It with 50 ~ 70% methyl alcohol is mobile phase, 10 ~ 14mL/ of flow velocity that high performance liquid chromatography separation purifying described in F, E step is
The reverse phase preparative column of min, 21.2 × 250 mm, 5um is fixing phase, and UV-detector Detection wavelength is 254 nm, each sample introduction
45 ~ 60uL, collects the chromatographic peak of 16 ~ 32min, is evaporated after repeatedly adding up.Obtain final product described biphenyl compound 2-
isopropenyl-6-methoxy-7-hydroxy-(4-hydroxyphenyl)-dihydrobenzofuran：
Organic solvent described in step A is 70 ~ 100% acetone, ethanol or methyl alcohol；
Organic solvent described in step B is ethyl acetate, chloroform, ether, petroleum ether or benzene；
Mixed organic solvents described in step C are n-hexane-acetone, chloroform-acetone, chloroform-methanol, petroleum ether-acetone or stone
Oily ether-ethyl acetate；
The volume proportion of the mixed organic solvents described in step C is 1:0、20:1、9:1、8:2、7:3、3:2、1:1、1:2、0:1.
Application of the biphenyl compound of the present invention in anti-rotavirus medicaments are prepared.
Garcinia maingayii of the present invention is not limited by area and kind, can realize the present invention.
Embodiment 1
Dry Guttiferae arbor branch, leaf and/or the kg of fruit 5.7 are taken, meal is broken to 40 mesh, with 70% acetone ultrasound
Extract 5 times, each 60min, extract solution merges；Extract solution is filtered, and is concentrated under reduced pressure into the 1/4 of volume；Stand, filter sediment,
It is condensed into 368g medicinal extract a；600g water is added in medicinal extract a, is extracted 4 times with the chloroform isometric with water, merge extraction phase, decompression
It is condensed into 247g medicinal extract b；Post is filled with 200 mesh silica gel 1600g, the acetone solution of 600g is added in medicinal extract b, be subsequently adding 100
Mesh silica gel 250g mixes sample, mixes upper prop after sample；1 is respectively with volume ratio:0、20:1、9:1、8:2、7:3、3:2、1:1、1:2、0:1
Chloroform-methanol mixed organic solvents gradient elution, collect gradient eluent, concentration, monitored through TLC, merge identical part,
Obtain 9 parts, volume ratio 7:The eluent c of 3 chloroform-methanol mixed organic solvents is 67g；Filled with reversed material C-18
Post, reversed-phase column on eluent c carries out gradient elution by 20 ~ 100% methanol aqueous solution of volume content, collects each several part wash-out
Liquid is simultaneously concentrated, and is monitored through TLC, merges identical part；Take the wash-out afforded with the methanol aqueous solution of volume content 50 ~ 70%
Liquid, then with 60% methyl alcohol be mobile phase, the Zorbax PrepHT anti-phase systems of GF of flow velocity 8mL/min, 21.2 × 250mm, 5um
Standby post is fixing phase, and UV-detector Detection wavelength is 254 nm, each sample introduction 50uL, collects the chromatographic peak of 15min, repeatedly tired
Plus after be evaporated, obtain final product described biphenyl compound 2-isopropenyl-6-methoxy-7-hydroxy- (4-
hydroxyphenyl)-dihydrobenzofuran。
Embodiment 2
Dry Guttiferae arbor branch, leaf and/or fruit 3kg are taken, meal is broken to 20 mesh, extracted with 100% EtOH Sonicate
3 times, each 60min, extract solution merges；Extract solution is filtered, and is concentrated under reduced pressure into the 1/3 of volume；Stand, filter sediment, concentrate
Into 189g medicinal extract a；The water of 360g is added in medicinal extract a, is extracted 4 times with the chloroform isometric with water, merge extraction phase, subtracted
Pressure is condensed into 158g medicinal extract b；Post is filled with 100 mesh silica gel 900g, the acetone solution of 260g is added in medicinal extract b, be subsequently adding
100 mesh silica gel 260gSample is mixed, upper prop after sample is mixed；1 is respectively with volume ratio:0、20:1、9:1、8:2、3:2、1:1、1:2、0:1
N-hexane-acetone mixed organic solvents gradient elution, collects gradient eluent, concentration, is monitored through TLC, merges identical part；
Volume ratio 9:The eluent c of 1 n-hexane-acetone mixed organic solvents is 76g；Post, eluent c are filled with reversed material C-8
Upper reversed-phase column, gradient elution is carried out by 20 ~ 100% methanol aqueous solution of volume content, is collected each several part eluent and is concentrated,
Monitored through TLC, merge identical part；Take the eluent afforded with the methanol aqueous solution of volume content 50 ~ 70%, then with
68% methyl alcohol is mobile phase, and the Zorbax PrepHT GF reverse phase preparative columns of flow velocity 14mL/min, 21.2 × 250mm, 5um are
Fixing phase, UV-detector Detection wavelength is 254 nm, each sample introduction 50uL, collects the chromatographic peak of 21min, is steamed after repeatedly adding up
It is dry, obtain final product described biphenyl compound 2-isopropenyl-6-methoxy-7-hydroxy- (4-hydroxyphenyl)-
dihydrobenzofuran。
Embodiment 3
Dry Guttiferae arbor branch, leaf and/or fruit 6kg are taken, meal is broken to 30 mesh, with 80% methyl alcohol ultrasonic extraction 4
Secondary, each 30min, extract solution merges；Extract solution is filtered, and is concentrated under reduced pressure into the 1/2 of volume；Stand, filter sediment, be condensed into
392g medicinal extract a；The water of 700g is added in medicinal extract a, is extracted 4 times with the ether isometric with water, merge extraction phase, depressurized dense
Shorten 242g medicinal extract b into；Post is filled with 100 mesh silica gel 1600g, the acetone solution of 320g, Ran Houjia are added in medicinal extract b
Enter 100 mesh silica gel 260g and mix sample, mix upper prop after sample；1 is respectively with volume ratio:0、20:1、9:1、8:2、7:3、3:2、1:1、
1:2、0:1 chloroform-acetone mixed organic solvents gradient elution, collects gradient eluent, concentration, is monitored through TLC, merges identical
Part；Volume ratio 9:The eluent c of 1 chloroform-acetone mixed organic solvents is 45g；Post is filled with reversed material ODS, is washed
Reversed-phase column on de- liquid c, gradient elution is carried out by 20 ~ 100% methanol aqueous solution of volume content, collects each several part eluent
And concentrate, monitored through TLC, merge identical part；Take the wash-out afforded with the methanol aqueous solution of volume content 50 ~ 70%
Liquid, then be mobile phase with 55% methyl alcohol, the Zorbax PrepHT GF of flow velocity 12mL/min, 21.2 × 250mm, 5um are anti-
Post is mutually prepared for fixing phase, UV-detector Detection wavelength is 254nm, each sample introduction 50uL, collects the chromatographic peak of 19min,
It is evaporated after repeatedly adding up, obtains final product described biphenyl compound 2-isopropenyl-6-methoxy-7-hydroxy- (4-
hydroxyphenyl)-dihydrobenzofuran。
Embodiment 4
Dry Guttiferae arbor branch, leaf and/or fruit 5.3kg are taken, meal is broken to 40 mesh, is carried with 90% EtOH Sonicate
Take 3 times, each 45min, extract solution merges；Extract solution is filtered, and is concentrated under reduced pressure into the 1/4 of volume；Stand, filter sediment,
It is condensed into 321g medicinal extract a；The water of 680g is added in medicinal extract a, with the petroleum ether extraction isometric with water 4 times, is closed
And extraction phase, it is concentrated under reduced pressure into 235g medicinal extract b；Post is filled with 100 mesh silica gel 1450g, the acetone of 290g is added in medicinal extract b
Dissolving, is subsequently adding 100 mesh silica gel 265gSample is mixed, upper prop after sample is mixed；1 is respectively with volume ratio:0、20:1、9:1、8:2、7:3、
3:2、1:1、1:2、0:1 petroleum ether-acetone mixed organic solvents gradient elution, collects gradient eluent, concentration, through TLC
Monitoring, merges identical part；Volume ratio 9:The eluent c of 1 petroleum ether-acetone mixed organic solvents is 52g；With anti-
Phase material MCI fills post, and reversed-phase column on eluent c carries out gradient elution by 20 ~ 100% methanol aqueous solution of volume content, receives
Collection each several part eluent is simultaneously concentrated, and is monitored through TLC, merges identical part；Take water-soluble with the methyl alcohol of volume content 50 ~ 70%
The eluent that liquid is afforded, then with 70% methyl alcohol be mobile phase, flow velocity 10mL/min, 21.2 × 250mm, 5um's
Zorbax PrepHT GF reverse phase preparative columns are fixing phase, and UV-detector Detection wavelength is 254nm, collects the chromatogram of 14min
Peak, is evaporated after repeatedly adding up, and obtains final product described biphenyl compound 2-isopropenyl-6-methoxy-7-hydroxy- (4-
hydroxyphenyl)-dihydrobenzofuran。
Embodiment 5
Dry Guttiferae arbor branch, leaf and/or the kg of fruit 10 are taken, meal is broken to 20 mesh, with 70% methyl alcohol ultrasonic extraction 4
Secondary, 35 min every time, extract solution merges；Extract solution is filtered, and is concentrated under reduced pressure into the 1/2 of volume；Stand, filter sediment, concentrate
Into 879g medicinal extract a；The water of 1700g is added in medicinal extract a, is extracted 5 times with the benzene isometric with water, merge extraction phase, depressurized dense
Shorten 445g medicinal extract b into；Post is filled with the g of 200 mesh silica gel 3330, the acetone solution of 900 g is added in medicinal extract b, be subsequently adding 100
Mesh silica gel 580g mixes sample, mixes upper prop after sample；1 is respectively with volume ratio:0、20:1、9:1、8:2、3:2、1:1、1:2、0:1 stone
Oily ether-ethyl acetate mixed organic solvents gradient elution, collects gradient eluent, concentration, is monitored through TLC, merges identical portion
Point；Volume ratio 9:The eluent c of 1 petroleum ether-ethyl acetate mixed organic solvents is 105 g；Post is filled with reversed material C-18,
Reversed-phase column on eluent c, gradient elution is carried out by 20 ~ 100% methanol aqueous solution of volume content, collects each several part eluent
And concentrate, monitored through TLC, merge identical part；Take the wash-out afforded with the methanol aqueous solution of volume content 50 ~ 70%
Liquid, then be mobile phase with 50% methyl alcohol, the Zorbax PrepHT GF of flow velocity 12 mL/min, 21.2 × 250 mm, 5um are anti-
Post is mutually prepared for fixing phase, UV-detector Detection wavelength is 254 nm, collect the chromatographic peak of 31 min, steamed after repeatedly adding up
It is dry, obtain final product described biphenyl compound 2-isopropenyl-6-methoxy-7-hydroxy- (4-hydroxyphenyl)-
dihydrobenzofuran。
Embodiment 6
Dry Guttiferae arbor branch, leaf and/or the kg of fruit 8.1 are taken, meal is broken to 20 mesh, is carried with 100% acetone ultrasound
Take 4 times, every time 30 min, extract solution merges；Extract solution is filtered, and is concentrated under reduced pressure into the 1/2 of volume；Stand, filter sediment, it is dense
Shorten 638g medicinal extract a into；The water of 1200g is added in medicinal extract a, is extracted 5 times with the benzene isometric with water, merge extraction phase, decompression
It is condensed into 362g medicinal extract b；Post is filled with the g of 200 mesh silica gel 2400, the acetone solution of 500 g is added in medicinal extract b, be subsequently adding
100 mesh silica gel 400g mix sample, mix upper prop after sample；1 is respectively with volume ratio:0、20:1、9:1、8:2、3:2、1:1、1:2、0:1
Petroleum ether-acetone mixed organic solvents gradient elution, collects gradient eluent, concentration, is monitored through TLC, merges identical part；
Volume ratio 9:The eluent c of 1 petroleum ether-ethyl acetate mixed organic solvents is 87 g；Post, wash-out are filled with reversed material ODS
Reversed-phase column on liquid c, gradient elution is carried out by 20 ~ 100% methanol aqueous solution of volume content, and collection each several part eluent is simultaneously dense
Contracting, monitors through TLC, merges identical part；The eluent afforded with the methanol aqueous solution of volume content 50 ~ 70% is taken, then
It is mobile phase, the Zorbax PrepHT anti-phase systems of GF of flow velocity 12 mL/min, 21.2 × 250 mm, 5um with 62% methyl alcohol
It is fixing phase for post, UV-detector Detection wavelength is 254 nm, collects the chromatographic peak of 22 min, is evaporated after repeatedly adding up, i.e.,
Described biphenyl compound 2-isopropenyl-6-methoxy-7-hydroxy- (4-hydroxyphenyl)-
dihydrobenzofuran。
Embodiment 7
Compound 2-isopropenyl-6-methoxy-7-hydroxy- (4-hydroxyphenyl) prepared by Example 1-
Dihydrobenzofuran, is orange-yellow jelly；Assay method is：With nuclear magnetic resonance, identified with reference to other spectroscopic techniques and tied
Structure：
（1）Ultraviolet spectra（Solvent is methyl alcohol）,λ max（logε）：570 (1.79), 266 (3.69), 226 (3.82),
204 (3.96) nm；
（2）Infrared spectrum（Pressing potassium bromide troche）V max 3 432, 2 924, 2 852, 1 630, 1 503, 1 450, 1
374, 1 243, 1 173, 1 096, 1 054, 1 027, 905, 855, 824, 591 cm–1；
（3）HRESIMS shows the compounds of this invention quasi-molecular ion peakm/z 298.1202 [M]+（Calculated value is
298.1205）, with reference to13C and1H H NMR spectroscopies（Fig. 1 and Fig. 2, carbon spectrum hydrogen modal data ownership is shown in Fig. 4）Providing its molecular formula is
C18H18O4。1H NMR（C5D5N, 500 MHz）With13C NMR（C5D5N, 125 MHz）Data, are shown in Fig. 4；
HRESIMS shows that its quasi-molecular ion peak is 298.1202 [M]+(calculated value 298.1205), with reference to13C H NMR spectroscopies
Determine that molecular formula is C18H18O4, degree of unsaturation is 10.Infrared absorption spectroscopy is in 3432 cm-1 Show the presence of hydroxyl,
1630、1503 cm-1Show the presence of phenyl ring, ultraviolet spectra has absorption maximum to also demonstrate that phenyl ring in 204,226,266 nm
In the presence of.13C NMR and1H NMR modal datas show the characteristic signal of biphenyl structural.1H NMR data displays have a fragrance
Protonδ H6.81 (1H, s), show the compound be have five substituted biphenyl structurals.Aromaticδ H 6.81 (1H, s)
Be H-5 because shown in HMBC it and C-4 (δ C 128.8)、C-9 (δ C 118.7)、C-7 (δ C 132.1)、C-6
(δ C150.0) and C-1 ' ' (δ C132.5) there is correlation.1One phenyl ring of contraposition substitution of H NMR data displays [δ H
7.58 (2H, d, J=8.5 Hz, H-2 ' ', H-6 ' ') and 7.33 (2H, d,J = 8.5 Hz, H-3′′,
5 ' ')], part signal for isopropenyl coumarone [δ H 3.44 (1H, dd, J = 15.2, 9.0 Hz, H-3),
3.23 (H, dd, J = 15.2, 8.6 Hz, H-3), 5.30 (1H, t, J = 8.6 Hz, H-2), 4.88 (1H,
S, H-2 '), 5.22 (1H, s, H-2 ') and 1.72 (3H, s, H-3 ')], also one methoxyl group (δ H 3.84,
3H, s, 6-OMe).In HMBC, H-3 and C-4, C-9, C-8 (δ C149.1) related, H-2 is related to C-8, C-9,
It is known that isopropenyl coumarone is connected with C-8, C-9.Methoxyl group (δ H3.84) be substituted in C-6, can be by it and C-
The related confirmation of 6 HMBC.Understand that two hydroxyls are located at C-7, C-4 ' ' position respectively according to its molecular formula.So far, the compound
Structure determined, be named as 2-isopropenyl-6-methoxy-7-hydroxy- (4-hydroxyphenyl)-
dihydrobenzofuran。
Embodiment 8
Compound prepared by Example 2, is orange-yellow jelly；Structure determination is carried out by the method in embodiment 6, as a result
For：With embodiment 6, molecular formula is C to its structure18H18O4.Confirm that compound prepared by embodiment 2 is described biphenyl compound
2-isopropenyl-6-methoxy-7-hydroxy-(4-hydroxyphenyl)-dihydrobenzofuran。
Embodiment 9
Compound prepared by Example 3, is orange-yellow jelly；Structure determination is carried out by the method in embodiment 6, as a result
For：With embodiment 6, molecular formula is C to its structure18H18O4.Confirm that compound prepared by embodiment 3 is described biphenyl compound
2-isopropenyl-6-methoxy-7-hydroxy-(4-hydroxyphenyl)-dihydrobenzofuran；
Fig. 4 compounds1H and13C NMR datas（Solvent is C5D5N）(125 and 500 MHz).
Embodiment 10
Compound prepared by Example 4, is orange-yellow jelly；Structure determination is carried out by the method in embodiment 6, as a result
For：With embodiment 6, molecular formula is C to its structure18H18O4.Confirm that compound prepared by embodiment 4 is described biphenyl compound
2-isopropenyl-6-methoxy-7-hydroxy-(4-hydroxyphenyl)-dihydrobenzofuran。
Embodiment 11
Compound prepared by Example 5, is orange-yellow jelly；Structure determination is carried out by the method in embodiment 6, as a result
For：With embodiment 6, molecular formula is C to its structure18H18O4.Confirm that compound prepared by embodiment 5 is described biphenyl compound
2-isopropenyl-6-methoxy-7-hydroxy-(4-hydroxyphenyl)-dihydrobenzofuran。
Embodiment 12
Any biphenyl compound prepared by Example 1 ~ 5 carries out cytotoxicity assay experiment, and test situation is as follows：
Cell line:Rhesus monkey kidney cell line (MA-104) is provided by Shanghai Wei Meng bio tech ltd.
Experimental design:MA-104 cells and various concentrations compound incubation 72 hours, every plant of experiment counterpoise of cell
It is multiple once to carry out data processing with the result tested twice, the suppression that compound on intracellular is bred is evaluated using improvement MTT methods
Processing procedure degree, calculates inhibiting rate, and IC is calculated using Logit methods according to inhibiting rate50, the extracorporeal anti-tumor work of comparative compound
Property.
EC50Refer to medium effective concentration, refer to cause 50% experimental animal to produce a certain specific reaction, or certain reaction
Index is suppressed concentration during half.
CC50Refer to half cytotoxic concentration, refer to that concentration needed for toxic action is produced to half cell.In this experiment
In, refer to the drug concentration needed for causing 50% cell death.
Compound Cytotoxicity is determined
Compound dimethyl sulfoxide (DMSO) (dimethylsulfoxide, DMSO) dissolves, and in the min of microwave sterilization 10, uses MEM
The mother liquor for being made into l mg/mL is standby, and MEM solution is diluted to required concentration.96 porocyte culture plates, plus l × 105 /
The MA-104 cell suspensions of mL concentration, 100uL/ holes, 37 DEG C, 5%CO2 The h of incubator culture 24, in well-grown individual layer
Concentration respectively l mg/mL, 0.2 mg/mL, 40 are separately added on cellug/mL、8 ug/mL、1.25 uThe chemical combination of g/mL
Thing;100uL/ holes, each concentration sets 3 multiple holes, while setting normal cell controls.It is placed in 37 DEG C, 5%CO2 Incubator continues to train
After supporting 24h, MTT methods detection cell survival rate.
Compounds on viral infection mitigation is acted on
It is 10 with concentration4/ mL, per hole l00uL inoculating cells are cultivated 24 hours in 96 orifice plates, see that cell grows up to list
Simultaneously upgrowth situation is good for layer, and 100 are respectively with concentrationug/mL、75 ug/mL、50 ug/mL、25 ug/mL、l uG/mL's
Compound is every with the rotavirus of 100 TCID 50/mL after the 37 DEG C of incubator advance h of function cells 1.5, PBS washings
Hole 100uL is discarded after adsorbing 1 h, plus MEM culture mediums 100uL/ holes maintain, and put 37 DEG C, 5% CO2Incubator, often
Day observation of cell lesion situation.Viral suppression is detected with MTT methods after 48h.
Cell survival rate is determined
Using MTT methods, 5 mg/mL methyl thiazolyl tetrazoliums (methyl are added in the cell of culture 48h
Thiazolyl tetrazolium, MTT) 20uL, continues to cultivate 3-4 h, abandons supernatant, adds DMSO per hole 100uL,
Vibration make in the hole in crystallize be completely dissolved after immediately 490nmMensuration absorbance A values under wavelength：
The average A values of cell survival rate=medicine group/cell controls group A value × 100%
Viral suppression=[the average virus control group average A-value of A values one of experimental group]/[the average virus of A values one of cell controls group
The average A values of control group] × 100%
Therapeutic index (TI)=half toxic concentration (CC50)/half-inhibition concentration (IC50)。
Experimental result
Test result indicate that：Through anti-rotavirus activity experiment, from virazole as control, 2-isopropenyl-6-
CCs of methoxy-7-hydroxy- (the 4-hydroxyphenyl)-dihydrobenzofuran to rotavirus50And EC50
Value is respectively 164.2 and 12.9μMol/L, it has preferable anti-rotavirus activity（As shown in Figure 5）.
Claims (7)
1. a kind of dihydrofuran biphenyl compound, it is characterized in that：Described biphenyl compound is tall with dry Guttiferae
Wooden branch bar, leaf or fruit are raw material, are obtained through medicinal extract extraction, organic solvent extraction, silica gel column chromatography, high performance liquid chromatography separation
, the compound molecule formula is C18H18O4, it is named as 2-isopropenyl-6-methoxy-7-hydroxy- (4-
Hydroxyphenyl)-dihydrobenzofuran, with following structural formula：
。
2. the preparation method of the biphenyl compound described in a kind of claim 1, it is characterised in that be tall with dry Guttiferae
Wooden branch bar, leaf and/or fruit are raw material, through medicinal extract extraction, organic solvent extraction, silica gel column chromatography, high performance liquid chromatography separation
Obtain, specially：
A, medicinal extract are extracted：By Guttiferae arbor branch, leaf or fruit coarse crushing to 20 ~ 40 mesh, with organic solvent ultrasonic extraction 2 ~ 4
Secondary, 30 ~ 60 min every time, extract solution merges；Extract solution is filtered, quiet during extract solution to 1/4 ~ 1/2 volume that is concentrated under reduced pressure
Put, filter sediment, be condensed into medicinal extract a；
B, organic solvent extraction：Add weight than 1 ~ 2 times of water of amount in medicinal extract a, extracted with the organic solvent isometric with water
3 ~ 5 times, merge organic solvent extraction phase, be concentrated under reduced pressure into medicinal extract b；
C, silica gel column chromatography：Medicinal extract b weight is dissolved than 1.5 ~ 3 times of organic solvents of amount, then 0.8 is weighed with medicinal extract ~
1.2 times of 100 ~ 200 mesh silica gel mixed samples, then go up silica gel column chromatography, and dress post silica gel is 100 ~ 200 mesh, and consumption is medicinal extract
6 ~ 8 times of amounts of b weight；It is 1 with volume ratio:0~0:1 mixed organic solvents gradient elution, collects gradient eluent, concentration,
Monitored through TLC, merge identical part；
D, reversed phase column chromatography：Will be with 4:Reversed phase column chromatography on the eluent that the organic solvent of 1 proportioning is afforded, it is anti-phase
Post is to fill post with reversed material C-8, C-18, ODS or MCI；With volume content for 40 ~ 100% methanol aqueous solution carries out ladder
Degree wash-out, collects each several part eluent and concentrates, and is monitored through TLC, merges identical part；
E, high performance liquid chromatography separation：The eluent that will be afforded with the methanol aqueous solution of volume content 55 ~ 80% is through efficient liquid
Phase chromatographic separation and purification, obtains final product described biphenyl compound 2-isopropenyl-6-methoxy-7-hydroxy- (4-
hydroxyphenyl)-dihydrobenzofuran；
It with 50 ~ 70% methyl alcohol is mobile phase, the mL/ of flow velocity 10 ~ 14 that high performance liquid chromatography separation purifying described in F, E step is
The reverse phase preparative column of min, 21.2 × 250 mm, 5 um is fixing phase, and UV-detector Detection wavelength is 254 nm, is entered every time
The uL of sample 45 ~ 60, collects the chromatographic peak of 16 ~ 34 min, is evaporated after repeatedly adding up；Obtain final product described biphenyl compound 2-
isopropenyl-6-methoxy-7-hydroxy-(4-hydroxyphenyl)-dihydrobenzofuran。
3. the preparation method of the biphenyl compound according to claim 2, it is characterized in that：It is organic molten described in step A
Agent is 80 ~ 100% acetone, ethanol or methyl alcohol.
4. the preparation method of the biphenyl compound according to claim 2, it is characterized in that：It is organic molten described in step B
Agent is ethyl acetate, chloroform, ether, petroleum ether or benzene.
5. the preparation method of the biphenyl compound according to claim 2, it is characterized in that：Being mixed with described in step C
Machine solvent is n-hexane-acetone, chloroform-acetone, chloroform-methanol, petroleum ether-acetone or petroleum ether-ethyl acetate.
6. the preparation method of biphenyl compound according to claim 2, it is characterized in that：Mixing described in step C is organic
The volume proportion of solvent is 1:0、20:1、9:1、8:2、7:3、3:2、1:1、1:2、0:1.
7. application of the biphenyl compound described in a kind of claim 1 in anti-rotavirus medicaments are prepared.
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| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN109053447A (en) * | 2018-08-06 | 2018-12-21 | 云南民族大学 | A kind of isoamyl alkyl-substituted biphenyls class compound and its preparation method and application |
| CN109485569A (en) * | 2018-08-06 | 2019-03-19 | 云南民族大学 | A kind of acetoxyl group isopentene group substituted biphenyl class compound and its preparation method and application |
| CN109485570A (en) * | 2018-08-06 | 2019-03-19 | 云南民族大学 | A kind of isopentene group substituted biphenyl class compound and its preparation method and application |
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Cited By (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN109053447A (en) * | 2018-08-06 | 2018-12-21 | 云南民族大学 | A kind of isoamyl alkyl-substituted biphenyls class compound and its preparation method and application |
| CN109485569A (en) * | 2018-08-06 | 2019-03-19 | 云南民族大学 | A kind of acetoxyl group isopentene group substituted biphenyl class compound and its preparation method and application |
| CN109485570A (en) * | 2018-08-06 | 2019-03-19 | 云南民族大学 | A kind of isopentene group substituted biphenyl class compound and its preparation method and application |
| CN109053447B (en) * | 2018-08-06 | 2021-04-06 | 云南民族大学 | Isopentyl substituted biphenyl compound and preparation method and application thereof |
| CN109485570B (en) * | 2018-08-06 | 2021-04-09 | 云南民族大学 | Isopentene group substituted biphenyl compound and preparation method and application thereof |
| CN109485569B (en) * | 2018-08-06 | 2021-05-18 | 云南民族大学 | Acetoxy isopentene group substituted biphenyl compound and preparation method and application thereof |
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