CN105085453B - A kind of utilization high-speed countercurrent chromatography method that separation prepares oligomeric stilbene compound from Chinese small iris - Google Patents
A kind of utilization high-speed countercurrent chromatography method that separation prepares oligomeric stilbene compound from Chinese small iris Download PDFInfo
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- CN105085453B CN105085453B CN201510458256.1A CN201510458256A CN105085453B CN 105085453 B CN105085453 B CN 105085453B CN 201510458256 A CN201510458256 A CN 201510458256A CN 105085453 B CN105085453 B CN 105085453B
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- C—CHEMISTRY; METALLURGY
- 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
- C07D307/93—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 condensed with a ring other than six-membered
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- C—CHEMISTRY; METALLURGY
- 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
- C07D307/80—Radicals substituted by oxygen atoms
Abstract
The invention provides a kind of utilization high-speed countercurrent chromatography method that separation prepares oligomeric stilbene compound from Chinese small iris.Compared with traditional column chromatography separation technology, the present invention uses high speed adverse current chromatogram (HSCCC) isolation technics, is not only effectively shortened disengaging time, and reduce the consumption of the organic solvent in technical process and energy consumption.The present invention is separated by HSCCC twice and can obtain 4 kinds of oligomeric stilbene compounds of high-purity (can reach more than 95%), i.e. vitisin A, ε viniferin, vitisin B and vitisin C.
Description
Technical field
The present invention relates to a kind of utilization high-speed countercurrent chromatography the side for preparing oligomeric stilbene compound is separated from Chinese small iris
Method.
Background technology
Chinese small iris (Iris lactea Pall.var.chinensis (Fisch.) Koidz.), also known as Ma Lian, kalimeris, royal purple
Grass, Orchid etc., are Iridaceae (Iridaceae) Jris (Iris L.) perennial herb perennial plants.Chinese small iris NATURAL DISTRIBUTION
In China northeast, North China, northwest and other places, it is distributed with grassland region more universal.Chinese small iris is sweet, mild-natured, cooling blood and hemostasis, heat-clearing profit
It is wet.Modern age is among the people to be had as the disease such as oral contraceptive and treatment functional uterine bleeding, icteric hepatitis and cervical carcinoma.Mesh
Before, the isolated multiple compounds from Jris mainly include flavone compound and its glucosides, isoflavonoid
And its glucosides, benzoquinone compound, triterpene compound and its saponin(e, stilbene compound.
Oligomeric stilbene compound has extensive pharmacological action, such as antitumor, antibacterial, antiviral, anti-inflammatory, anti-oxidant, god
Through the effect such as protection.For effective exploitation utilizes Chinese small iris child resource, and the pharmacological activity for further studying oligomeric stilbene compound
And mechanism of action, quickly the method for the preparation oligomeric stilbene compound of high-purity has great importance from Chinese small iris for exploitation.And
To using traditional pillar layer separation, because traditional column chromatography has, separation cycle is long, consumption more than the separation of oligomeric stilbene compound
The deficiency such as a large amount of organic solvents, serious to the dead absorption of sample.Therefore, it is necessary to it is high-purity to prepare to explore some efficient methods
The oligomeric stilbene compound of degree.
High speed adverse current chromatogram (High-speed counter-current chromatography, abbreviation HSCCC) is separated
Technology is a kind of liquid luquid partition chromatography isolation technics of continuous high-efficient, it without any solid-state supporter or carrier, using two
Phase solvent system sets up a kind of special one-way fluid dynamic equilibrium in the helix tube of high speed rotation, wherein one mutually makees
It is fixing phase, it is another as mobile phase, can retain substantial amounts of fixing phase, the separation foundation of material during continuous wash-out
Its in two-phase distribution coefficient difference and realize, be particularly suitable for the separation of natural product active ingredient.
The content of the invention
Extracted from Chinese small iris using HSCCC technologies it is an object of the invention to provide one kind and separate four kinds of oligomeric stilbene classes
The method of compound.
Specifically, separated from Chinese small iris using high-speed countercurrent chromatography the invention provides one kind and prepare oligomeric stilbene class
The method of compound, it includes following operating procedure：
(1) preparation of crude extract：
After Chinese small iris is crushed, extracted with 50-90%v/v ethanol, ethanol is obtained after extract solution recycling design concentrated under reduced pressure
Crude extract, i.e., sample to be separated；
(2) vitisin A, flow point II and ε-viniferin (ε-grape element) are separated using high-speed countercurrent chromatography：
It is 2~6: 4~7: 2~6: 3~7 to take n-hexane, ethyl acetate, the volume ratio of first alcohol and water, is placed in separatory funnel
Middle preparation two-phase solvent system, takes phase for fixing phase, and lower phase is mobile phase；By chromatographic column of the fixing phase full of adverse current chromatogram,
At 25~35 DEG C, main frame is rotated forward, and rotating speed is 800~900r/min, then enters flowing with the flow pump of 1.5~10ml/min
Phase, poised state is reached to two-phase solvent in post；Crude extract prepared by step (1), dissolves by solvent of mobile phase, prepares sample
Product solution；By sample solution sample introduction, each target component is received respectively under UV-detector, concentrate, dry, you can respectively obtain
Vitisin A, flow point II and ε-viniferin；
(3) vitisin B and vitisin C are separated using high-speed countercurrent chromatography：
It is 2~7: 2~7: 2~7: 2~7 to take petroleum ether, ethyl acetate, the volume ratio of first alcohol and water, is placed in separatory funnel
Middle preparation two-phase solvent system, it be mutually fixing phase to remove, and upper phase is mobile phase；By chromatographic column of the fixing phase full of adverse current chromatogram,
At 25-35 DEG C, main frame is rotated forward, and rotating speed is 800~900r/min, then enters mobile phase with the flow pump of 1.5~10ml/min,
Poised state is reached in post to two-phase solvent；Flow point II is taken, is dissolved by solvent of mobile phase, prepare sample solution；By sample
Solution sample introduction, receives each target component respectively under UV-detector, concentration, dries, you can respectively obtain vitisin B and
vitisin C；
Wherein, the ultraviolet detection wavelength in step (2), (3) is 325nm.
Further, in step (1), concentration of alcohol be 75~90%v/v, preferably 80~90%v/v, more preferably
85%v/v.
Further, in step (1), ethanol extraction time is 1~5 time, preferably 3 times.
Further, in step (2), petroleum ether, ethyl acetate, the volume ratio of first alcohol and water are 2.5~3: 6~6.5: 4
~4.5: 5~5.5.
Further, in step (2), vitisin A, the appearance time of flow point II and ε-viniferin be followed successively by 40~
130min, 75~175min, 90~205min.
In a specific embodiment of the invention, n-hexane, ethyl acetate, the volume ratio of first alcohol and water are 3: 6.5: 4.2:
5.5, vitisin A, the appearance time of each compositions of flow point II and ε-viniferin be followed successively by 60~80min, 115~135min,
140~155min.
In another embodiment of the present invention, n-hexane, ethyl acetate, the volume ratio of first alcohol and water are 2.5: 6:
4.5: 5, vitisin A, the appearance time of each compositions of flow point II and ε-viniferin be followed successively by 90~130min, 145~
175min, 180~205min.
In another embodiment of the present invention, n-hexane, ethyl acetate, the volume ratio of first alcohol and water are 3: 6.4: 5,
Vitisin A, the appearance time of each compositions of flow point II and ε-viniferin be followed successively by 40~60min, 75~85min, 90~
105min。
Further, in step (2), at 25 DEG C, main frame is rotated forward, and rotating speed is 900r/min, then with 1.5~2.5ml/
The flow pump of min enters mobile phase.
Further, in step (3), petroleum ether, ethyl acetate, the volume ratio of first alcohol and water are 5: 5: 3: 6~7.
Further, in step (3), the appearance time of vitisin C and vitisin B be followed successively by 135~182min,
145~200min.
In a specific embodiment of the invention, petroleum ether, ethyl acetate, the volume ratio of first alcohol and water are 5: 5: 3: 6,
The appearance time of vitisin C and vitisin B is followed successively by 135~142min and 145~155min.
In another embodiment of the present invention, petroleum ether, ethyl acetate, the volume ratio of first alcohol and water are 5: 5: 3: 7,
The appearance time of vitisin C and vitisin B is followed successively by 155~182min and 185~200min.
In another embodiment of the present invention, petroleum ether, ethyl acetate, the volume ratio of first alcohol and water are 5: 5: 3: 6,
The appearance time of vitisin C and vitisin B is followed successively by 135~142min and 145~155min.
Further, in step (3), at 35 DEG C, main frame is rotated forward, and rotating speed is 800r/min, then with 3.0~5.0ml/
The flow pump of min enters mobile phase.
Advantages of the present invention：
1st, compared with traditional column chromatography separation technology, the present invention uses high speed adverse current chromatogram (HSCCC) isolation technics, not only
Disengaging time is effectively shortened, and reduces the consumption of the organic solvent in technical process and energy consumption.
2nd, the present invention is separated by HSCCC twice and can obtain 4 kinds of oligomeric stilbene classes of high-purity (can reach more than 95%)
Compound, i.e. vitisin A, ε-viniferin, vitisin B and vitisin C.
3rd, the present invention prepare 4 kinds of oligomeric stilbene compounds of gained respectively through UV, 1D (1H-NMR and13C-NMR) and 2D (1H-1H
COSY, HMBC and HSQC) nuclear-magnetism spectrum carry out structural confirmation identification.
With reference to specific embodiment, the present invention is further elaborated, but the present invention is not limited to following examples.Institute
State method and be conventional method unless otherwise instructed.The raw material can be obtained from open commercial sources unless otherwise instructed.
Brief description of the drawings
Fig. 1 be the sub- ethanol crude extract of Chinese small iris, flow point II, vitisin A, ε-viniferin, vitisin B and
The high-efficient liquid phase chromatogram of vitisin C
Fig. 2 is vitisin A, ε-viniferin, the full wavelength scanner figure of vitisin B and vitisin C
Fig. 3 is that present invention application high-speed countercurrent chromatography separation prepares vitisin A, ε-viniferin and flow point II
Separate and prepare figure
Fig. 4 is that present invention application high-speed countercurrent chromatography separates the separation preparation figure for preparing vitisin B and vitisin C
When Fig. 5 solvent systems are " n-hexane, ethyl acetate, the volume ratio of first alcohol and water are 4: 6: 5: 5 ", flow point 1, flow point
The separation of II and flow point 2 prepares collection of illustrative plates
Fig. 6 solvent systems for " petroleum ether, ethyl acetate, first alcohol and water volume ratio be 5: 5: 3: 6 " when, flow point 3 and stream
4 separation is divided to prepare collection of illustrative plates
Specific embodiment
Following embodiment instruments are as follows with reagent：
TBE-300C high-speed counter-current chromatographs (Shanghai Tongtian Biotechnology Co., Ltd.)；It is furnished with TBP5002 constant currents simultaneously
Pump (Shanghai Tongtian Biotechnology Co., Ltd.), DC-0506 low temperature thermostat baths (Shanghai Sunny Hengping Scientific Instrument Co., Ltd.),
(Shanghai is same for UV-2000D detectors (Shanghai Tongtian Biotechnology Co., Ltd.) and EasyChrom-1000 Data Processing in Chromatography Workstation
Field Bioisystech Co., Ltd).The high performance liquid chromatographs of Agilent 1260 (Agilent company), equipped with G1311C four-stage pumps,
G1329B automatic samplers, G1316A column ovens and G1315D PDADs.
Purity detecting (peak area normalization method), its chromatographic condition are carried out to each flow point using HPLC methods in following embodiments
For：Eclipse XDB C18Post (250mm × 4.6mm, 5 μm)；Mobile phase：Methanol-acetonitrile-the aqueous solution；Gradient elution program has
Body is as follows：0-20min, mobile phase for 30-50% methyl alcohol, 5%-5% acetonitriles；21-35min, mobile phase is 50-65%'s
Methyl alcohol, 5%-5% acetonitriles；Flow velocity：1.0ml/min；Column oven temperature is 30 DEG C；Detection wavelength is 325nm.
Embodiment 1
A kind of utilization high-speed countercurrent chromatography separates the method for preparing oligomeric stilbene compound including following from Chinese small iris
Step：
(1) after Chinese small iris is crushed, extracted 3 times with 85% ethanol, filtering, it is thick to obtain ethanol after recycling design concentrated under reduced pressure
Extract, i.e., sample to be separated
(2) by the well mixed rear stratification of the organic solvent system containing n-hexane, ethyl acetate, first alcohol and water, point
Do not take upper strata solvent and lower floor's solvent, ultrasound degassing 20min, using upper strata solvent as fixing phase, lower floor's solvent as mobile phase,
To the fixing phase is pumped into the separating pipe of high-speed counter-current chromatograph, 25 DEG C of separating pipe temperature is kept, with the rotating speed of 900rpm just
After turning 20min, mobile phase is pumped into, after after system balance, from sample introduction valve injection, and it is inverse to carry out high speed with the flow velocity of 1.5ml/min
Flow chromatography is separated, and Detection wavelength is 325nm, and flow point 1, flow point II and flow point 2 are received successively, is concentrated under reduced pressure and is dried to constant weight, is obtained
To the vitisin A (flow point 1) and ε-viniferin (flow point 2) of high-purity, wherein flow point II is one group of mixture；
Wherein n-hexane, ethyl acetate, the volume ratio of first alcohol and water are 3: 6.5: 4.2: 5.5；
The appearance time of flow point 1, flow point II and flow point 2 is followed successively by 60~80min, 115~135min, 140~155min；
(3) flow point II is taken, is separated using high speed adverse current chromatogram, separation method is as follows：
By the well mixed rear stratification of the organic solvent system containing petroleum ether, ethyl acetate, first alcohol and water, take respectively
Upper strata solvent and lower floor's solvent, ultrasound degassing 20min, using lower floor's solvent as fixing phase, upper strata solvent is used as mobile phase, Xiang Gao
The fixing phase is pumped into the separating pipe of fast counter-current chromatograph, 35 DEG C of separating pipe temperature is kept, is rotated forward with the rotating speed of 800rpm
After 20min, mobile phase is pumped into, after after system balance, from sample introduction valve injection, and high-speed counter-current is carried out with the flow velocity of 3.5ml/min
Chromatographic isolation, Detection wavelength is 325nm, and flow point 4 and flow point 3 are received successively, is concentrated under reduced pressure and dries to constant weight, obtains high-purity
Vitisin C (flow point 4) and vitisin B (flow point 3)；
Wherein petroleum ether, ethyl acetate, the volume ratio of first alcohol and water are 5: 5: 3: 6；
The appearance time of flow point 4 and flow point 3 is followed successively by 135~142min and 145~155min；
The structural confirmation result of the embodiment products therefrom is as follows：
14 kinds of compounds of table13C NMR datas
24 kinds of compounds of table1H NMR datas
It can be seen from above-mentioned testing result, 4 kinds of isolated compositions of the present invention are respectively：
Embodiment 2
A kind of utilization high-speed countercurrent chromatography separates the method for preparing oligomeric stilbene compound including following from Chinese small iris
Step：
(1) after Chinese small iris is crushed, extracted 3 times with 85% ethanol, filtering, it is thick to obtain ethanol after recycling design concentrated under reduced pressure
Extract, i.e., sample to be separated；
(2) by the well mixed rear stratification of the organic solvent system containing n-hexane, ethyl acetate, first alcohol and water, point
Do not take upper strata solvent and lower floor's solvent, ultrasound degassing 20min, using upper strata solvent as fixing phase, lower floor's solvent as mobile phase,
To the fixing phase is pumped into the separating pipe of high-speed counter-current chromatograph, 25 DEG C of separating pipe temperature is kept, with the rotating speed of 900rpm just
After turning 20min, mobile phase is pumped into, after after system balance, from sample introduction valve injection, and it is inverse to carry out high speed with the flow velocity of 2.0ml/min
Flow chromatography is separated, and Detection wavelength is 325nm, and flow point 1, flow point II and flow point 2 are received successively, is concentrated under reduced pressure and is dried to constant weight, is obtained
To vitisin A and the ε-viniferin of high-purity, wherein flow point II is one group of mixture；
Wherein n-hexane, ethyl acetate, the volume ratio of first alcohol and water are 2.5: 6: 4.5: 5；
The appearance time of flow point 1, flow point II and flow point 2 be followed successively by 90~130min, 145~175min, 180~
205min；
(3) flow point II is taken, is separated using high speed adverse current chromatogram, separation method is as follows：
By the well mixed rear stratification of the organic solvent system containing petroleum ether, ethyl acetate, first alcohol and water, take respectively
Upper strata solvent and lower floor's solvent, ultrasound degassing 20min, using lower floor's solvent as fixing phase, upper strata solvent is used as mobile phase, Xiang Gao
The fixing phase is pumped into the separating pipe of fast counter-current chromatograph, 35 DEG C of separating pipe temperature is kept, is rotated forward with the rotating speed of 800rpm
After 20min, mobile phase is pumped into, after after system balance, pump into sample solution, and high-speed counter-current is carried out with the flow velocity of 5.0ml/min
Chromatographic isolation, Detection wavelength is 325nm, and flow point 4 and flow point 3 are received successively, is concentrated under reduced pressure and dries to constant weight, obtains high-purity
Vitisin C and vitisin B；
Wherein petroleum ether, ethyl acetate, the volume ratio of first alcohol and water are 5: 5: 3: 7；
The appearance time of flow point 4 and flow point 3 is followed successively by 155~182min and 185~200min；
Embodiment 3
A kind of utilization high-speed countercurrent chromatography separates the method for preparing oligomeric stilbene compound including following from Chinese small iris
Step：
(1) after Chinese small iris is crushed, extracted 3 times with 85% ethanol, filtering, it is thick to obtain ethanol after recycling design concentrated under reduced pressure
Extract, i.e., sample to be separated；
(2) by the well mixed rear stratification of the organic solvent system containing n-hexane, ethyl acetate, first alcohol and water, point
Do not take upper strata solvent and lower floor's solvent, ultrasound degassing 20min, using upper strata solvent as fixing phase, lower floor's solvent as mobile phase,
To the fixing phase is pumped into the separating pipe of high-speed counter-current chromatograph, 25 DEG C of separating pipe temperature is kept, with the rotating speed of 900rpm just
After turning 20min, mobile phase is pumped into, after after system balance, from sample introduction valve injection, and it is inverse to carry out high speed with the flow velocity of 2.5ml/min
Flow chromatography is separated, and Detection wavelength is 325nm, and flow point 1, flow point II and flow point 2 are received successively, is concentrated under reduced pressure and is dried to constant weight, is obtained
To vitisin A and the ε-viniferin of high-purity, wherein flow point II is one group of mixture；
Wherein n-hexane, ethyl acetate, the volume ratio of first alcohol and water are 3: 6.4: 5；
The appearance time of flow point 1, flow point II and flow point 2 is followed successively by 40~60min, 75~85min, 90~105min；
(3) flow point II is taken, is separated using high speed adverse current chromatogram, separation method is as follows：
By the well mixed rear stratification of the organic solvent system containing petroleum ether, ethyl acetate, first alcohol and water, take respectively
Upper strata solvent and lower floor's solvent, ultrasound degassing 20min, using lower floor's solvent as fixing phase, upper strata solvent is used as mobile phase, Xiang Gao
The fixing phase is pumped into the separating pipe of fast counter-current chromatograph, 35 DEG C of separating pipe temperature is kept, is rotated forward with the rotating speed of 800rpm
After 20min, mobile phase is pumped into, after after system balance, pump into sample solution, and high-speed counter-current is carried out with the flow velocity of 3.0ml/min
Chromatographic isolation, Detection wavelength is 325nm, and flow point 4 and flow point 3 are received successively, is concentrated under reduced pressure and dries to constant weight, obtains high-purity
Vitisin C and vitisin B.
Wherein petroleum ether, ethyl acetate, the volume ratio of first alcohol and water are 5: 5: 3: 6；
The appearance time of flow point 4 and flow point 3 is followed successively by 135~142min and 145~155min；
Discuss：
Early stage of the present invention additionally uses other solvent systems and carries out the separation of each compound, but fails to reach preferable point
It is specific as follows from purification effect：
Flow point 1, flow point II and the separation condition 1 of flow point 2：N-hexane, ethyl acetate, the volume ratio of first alcohol and water are 4: 6: 5:
5, other conditions are consistent with embodiment.Result finds, under the system condition, flow point 1, the separating degree between flow point II and flow point 2
It is bad, and flow point 1 fails to be separated (Fig. 5) with impurity.The purity of HPLC purity checks, flow point 1, flow point II and flow point 2 does not have
Embodiment is high.
Flow point 4 and the separation condition of flow point 3：Petroleum ether, ethyl acetate, the volume ratio of first alcohol and water are 5: 5: 3: 6, with upper strata
, used as fixing phase, lower floor's solvent is used as mobile phase, and other conditions are consistent with embodiment for solvent.Flow point 3 and flow point 4 fail to separate
(see Fig. 6).
Claims (12)
1. a kind of utilization high-speed countercurrent chromatography separates the method for preparing oligomeric stilbene compound from Chinese small iris, and its feature exists
In：It includes following operating procedure：
(1) preparation of crude extract：
After Chinese small iris is crushed, extracted with 50-90%v/v ethanol, obtaining ethanol after extract solution recycling design concentrated under reduced pressure slightly carries
Thing, i.e., sample to be separated；
(2) vitisin A, flow point II and ε-viniferin are separated using high-speed countercurrent chromatography：
It is 2~6: 4~7: 2~6: 3~7 to take n-hexane, ethyl acetate, the volume ratio of first alcohol and water, is placed in separatory funnel and matches somebody with somebody
Two-phase solvent system processed, takes phase for fixing phase, and lower phase is mobile phase；By chromatographic column of the fixing phase full of adverse current chromatogram, 25
At~35 DEG C, main frame is rotated forward, and rotating speed is 800~900r/min, then enters mobile phase with the flow pump of 1.5~10ml/min, extremely
Two-phase solvent reaches poised state in post；Crude extract prepared by step (1), dissolves by solvent of mobile phase, prepares sample molten
Liquid；By sample solution sample introduction, each target component is received respectively under UV-detector, concentrate, dry, you can respectively obtain
Vitisin A, flow point II and ε-viniferin；
(3) vitisin B and vitisin C are separated using high-speed countercurrent chromatography：
It is 2~7: 2~7: 2~7: 2~7 to take petroleum ether, ethyl acetate, the volume ratio of first alcohol and water, is placed in separatory funnel and matches somebody with somebody
Two-phase solvent system processed, it be mutually fixing phase to remove, and upper phase is mobile phase；By chromatographic column of the fixing phase full of adverse current chromatogram, in 25-
At 35 DEG C, main frame is rotated forward, and rotating speed is 800~900r/min, then enters mobile phase with the flow pump of 1.5~10ml/min, to two
Phase solvent reaches poised state in post；Flow point II is taken, is dissolved by solvent of mobile phase, prepare sample solution；By sample solution
Sample introduction, receives each target component respectively under UV-detector, concentration, dries, you can respectively obtain vitisin B and
vitisin C；
Wherein, the ultraviolet detection wavelength in step (2), (3) is 325nm.
2. method according to claim 1, it is characterised in that：In step (1), concentration of alcohol is 75~90%v/v.
3. method according to claim 2, it is characterised in that：In step (1), concentration of alcohol is 80~90%v/v.
4. method according to claim 3, it is characterised in that：In step (1), concentration of alcohol is 85%v/v.
5. method according to claim 1, it is characterised in that：In step (1), ethanol extraction time is 1~5 time.
6. method according to claim 5, it is characterised in that：In step (1), ethanol extraction time is 3 times.
7. method according to claim 1, it is characterised in that：In step (2), n-hexane, ethyl acetate, first alcohol and water
Volume ratio is 2.5~3: 6~6.5: 4~4.5: 5~5.5.
8. method according to claim 1, it is characterised in that：In step (2), vitisin A, flow point II and ε-
The appearance time of viniferin is followed successively by 40~130min, 75~175min, 90~205min.
9. method according to claim 1, it is characterised in that：In step (2), at 25 DEG C, main frame is rotated forward, and rotating speed is
900r/min, then enters mobile phase with the flow pump of 1.5~2.5ml/min.
10. method according to claim 1, it is characterised in that：In step (3), petroleum ether, ethyl acetate, first alcohol and water
Volume ratio be 5: 5: 3: 6~7.
11. methods according to claim 1, it is characterised in that：In step (3), vitisin C and vitisin B's goes out
Peak time is followed successively by 135~182min and 145~200min.
12. methods according to claim 1, it is characterised in that：In step (3), at 35 DEG C, main frame is rotated forward, and rotating speed is
800r/min, then enters mobile phase with the flow pump of 3.0~5.0ml/min.
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| CN110590881B (en) * | 2019-09-23 | 2023-05-16 | 中国科学院西北高原生物研究所 | Oligomeric stilbene compounds in Iris seed kernel, and extraction method and application thereof |
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