CN106243173A - Extract and the method for isolated and purified hypolipidemic activity compound Quercetin 3 O β D galactoside from the fine horse Fructus Jujubae of South Sinkiang - Google Patents
Extract and the method for isolated and purified hypolipidemic activity compound Quercetin 3 O β D galactoside from the fine horse Fructus Jujubae of South Sinkiang Download PDFInfo
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Abstract
The present invention is open to be extracted and the method for isolated and purified hypolipidemic activity compound Quercetin 3 O β D galactoside from the fine horse Fructus Jujubae of South Sinkiang, comprising the steps: to extract in the fine horse Fructus Jujubae of (I) South Sinkiang hypolipidemic activity material, jujube pulp is separated by (1 1) with Semen Jujubae；Jujube pulp is extracted by (1 2) with the aqueous solution of alcohol, obtains alcohol extraction thing solution and solid residue, and alcohol extraction thing solution is concentrated to give dry extract；(II) separation of Quercetin 3 O β D galactoside and purification.The present invention is extracted the material with hypolipidemic activity from the fine horse Fructus Jujubae of South Sinkiang, and mouse test shows that it has certain hypolipidemic activity, and separating-purifying has gone out to have hypolipidemic activity compound Quercetin 3 O β D galactoside.
Description
Technical field
The present invention relates to technical field of deep red date processing.Extract and isolated and purified more particularly, to from the fine horse Fructus Jujubae of South Sinkiang
The method of hypolipidemic activity compound Quercetin-3-O-β-D-galactoside.
Background technology
Hyperlipemia shows as people's HypercholesterolemicRats and occurs abnormal, and in blood plasma, a certain or multiple lipide component concentration is high
The low disease exceeding normal range, is coronary heart disease and one of the Main Etiological Factors of tremulous pulse medicated porridge hardening.Owing to gallbladder total in blood plasma is solid
Alcohol and triglyceride are too high, and HDL-C is too low thus a kind of morbid state of occurring.It is primarily due to lipid
Being insoluble or poorly soluble in water, it must form lipoprotein with protein bound could exist human body.Common hyperlipidemia is mainly led
Index change is wanted to have: the content of serum total cholesterol (TC), low density lipoprotein, LDL (LDL-C) and triglyceride (TG) raises；High
Density lipoprotein-cholesterol (HDL-C) reduction etc..
The harm of hyperlipidemia is self-evident, can cause a variety of disease, as hypertension, diabetes, atherosclerosis,
The diseases such as coronary heart disease.In blood fat, leading indicator content exceeds normal range, easily makes blood " blood is thick " occur, may block blood
Pipe, makes vascular pressure uprise, and velocity of blood flow is slack-off, it is easy to cause hypertension symptom, will cause blood in blood vessel time serious
Stream interrupts, if there is the Different Organs in human body, produces different types of disease.If occurring at brain, it may occur that apoplexy；
If occurring at heart, it is likely that coronary heart disease easily occurs；If occurring at lower limb, limbs will be caused to fester, downright bad etc.；Hyperlipidemia
Also can cause liver dysfunction, if liver dysfunction, then lipid and lipoprotein metabolism disorder can occur, so easily draw for a long time
Play fatty liver, even hepatocarcinoma.
Fructus Jujubae is a kind of medicinal and edible food, as medicine, and existing more than 2000 year history, there is abundant nutritional labeling
And active component, there are " tonic king ", the laudatory title of " first-class tonic ".The traditional Chinese medical science thinks that Fructus Jujubae has a variety of edible and pharmacologic action.
Research shows, containing bioactive ingredients such as flavonoid, saccharide, cyclic adenosine monophosphates in Fructus Jujubae, has good pharmacologic action, has
The multiple biological activitys such as enhancing immunity, antitumor, blood fat reducing.
At present, the research to Fructus Jujubae is mainly Middle Eastern Fructus Jujubae, the most isolated and purified, and for being grown in nature bar
Part is superior, and South Sinkiang, southern foot, the Tianshan Mountains Fructus Jujubae hypolipidemic activity research that illumination degree abundant, saline and alkaline is high has no report.South Sinkiang fine horse Fructus Jujubae kind
Planting area wide, yield is high, and quality better, compared with interior ground Fructus Jujubae chemical analysis content, there were significant differences, therefore to South Sinkiang fine horse Fructus Jujubae
The applied research of hypolipidemic activity composition is extremely urgent.
Summary of the invention
It is an object of the present invention to provide in the fine horse Fructus Jujubae of a kind of South Sinkiang and extract and isolated and purified hypolipidemic activity compound
The method of Quercetin-3-O-β-D-galactoside.
For reaching above-mentioned purpose, the present invention uses following technical proposals:
Extract and the side of isolated and purified hypolipidemic activity compound Quercetin-3-O-β-D-galactoside from the fine horse Fructus Jujubae of South Sinkiang
Method, comprises the steps:
(I) South Sinkiang fine horse Fructus Jujubae extracts hypolipidemic activity material；
(1-1) jujube pulp is separated with Semen Jujubae；
(1-2) with alcoholic solution, jujube pulp is extracted, obtain alcohol extraction thing solution and solid residue, by dense for alcohol extraction thing solution
Contract to obtain dry extract；
(II) separation of Quercetin-3-O-β-D-galactoside and purification.
Above-mentioned extraction and isolated and purified hypolipidemic activity compound Quercetin-3-O-β-D-galactoside from the fine horse Fructus Jujubae of South Sinkiang
Method, in step (I), also comprise the steps:
(1-3) being that 1:4g/mL is soluble in water by dry extract according to solid-liquid ratio obtains dry extract solution, with ether to dry extract solution
Extract, obtain ether phase and ether extracts residue aqueous phase.
Above-mentioned extraction and isolated and purified hypolipidemic activity compound Quercetin-3-O-β-D-galactoside from the fine horse Fructus Jujubae of South Sinkiang
Method, also comprise the steps:
(1-4) with chloroform, ether is extracted residue aqueous phase to extract, obtain chloroform phase and chloroform extraction residue aqueous phase.
Above-mentioned extraction and isolated and purified hypolipidemic activity compound Quercetin-3-O-β-D-galactoside from the fine horse Fructus Jujubae of South Sinkiang
Method, also comprise the steps:
(1-5) with ester, chloroform extraction residue aqueous phase is extracted, obtain ester phase and ester extracts residue aqueous phase.
Above-mentioned extraction and isolated and purified hypolipidemic activity compound Quercetin-3-O-β-D-galactoside from the fine horse Fructus Jujubae of South Sinkiang
Method, in step (1-2): the aqueous solution of alcohol is ethanol water, solid-liquid ratio is 1:(2～4) g/mL, second in ethanol water
The volume fraction of alcohol is 60-80%, reflux, extract,.
Above-mentioned extraction and isolated and purified hypolipidemic activity compound Quercetin-3-O-β-D-galactoside from the fine horse Fructus Jujubae of South Sinkiang
Method, in step (1-3): ether is petroleum ether, the solid-liquid ratio of dry extract and petroleum ether is 4:(0.5～2) g/mL.
Above-mentioned extraction and isolated and purified hypolipidemic activity compound Quercetin-3-O-β-D-galactoside from the fine horse Fructus Jujubae of South Sinkiang
Method, in step (1-4): the solid-liquid ratio of dry extract and chloroform is 4:(0.5～2) g/mL.
Above-mentioned extraction and isolated and purified hypolipidemic activity compound Quercetin-3-O-β-D-galactoside from the fine horse Fructus Jujubae of South Sinkiang
Method, in step (1-5): ester is ethyl acetate, the solid-liquid ratio of dry extract and ethyl acetate is 4:(0.5～2) g/mL；Will
To ethyl acetate concentrate mutually afterwards hypolipidemic activity material.
Above-mentioned extraction and isolated and purified hypolipidemic activity compound Quercetin-3-O-β-D-galactoside from the fine horse Fructus Jujubae of South Sinkiang
Method, in step (II), comprise the steps:
(2-1) cross the first silicagel column by step (I) is extracted the hypolipidemic activity material obtained, eluant be petroleum ether and
The volume ratio of the mixture of acetone, petroleum ether and acetone is 5:1；
(2-2) eluent obtained in step (2-1) being crossed the second silicagel column, eluant is the mixing of chloroform and methanol
The volume ratio of thing, chloroform and methanol is 5:1；
(2-3) eluent obtained in step (2-2) being crossed gel column, eluant is the mixture of chloroform and methanol, chlorine
Imitative and methanol volume ratio is 1:1, and every 10mL collects a pipe, is collected to the 123rd pipe eluent by the 41st pipe and merges；
(2-4) the 41st pipe to the 123rd pipe eluent obtained in step (2-3) being crossed middle compression leg, eluant is methanol-water
Solution, the volume fraction of methanol is 70%, and every 10mL collects a pipe, is collected to the 53rd pipe eluent by the 14th pipe and merges；
(2-5) the 14th pipe to the 53rd pipe eluent obtained in step (2-4) is crossed performance liquid chromatographic column, collect eluting
Liquid, what eluent was evaporated concentration obtains yellow powder, is hypolipidemic activity compound Quercetin-3-O-β-D-galactoside.
Above-mentioned extraction and isolated and purified hypolipidemic activity compound Quercetin-3-O-β-D-galactoside from the fine horse Fructus Jujubae of South Sinkiang
Method,
In step (2-1): described first silicagel column is gross porosity zcx-II type silicagel column, and external diameter is 4.5 centimetres, eluent stream
Speed is 5-10mL/min；
In step (2-2): described first silicagel column is gross porosity zcx-II type silicagel column, and external diameter is 4 centimetres, eluant flow velocity
For 1-2mL/min；
In step (2-3): described gel column is Sephadex LH-20 type gel column, and external diameter is 2.5 centimetres, eluent stream
Speed is 0.5-1mL/min；
In step (2-4): medium pressure post external diameter is 4 centimetres, post pressure is 3-4MPa, and eluant flow velocity is 3-4mL/min；
In step (2-5):
Performance liquid chromatographic column is one of following: a) chromatographic column: Agilent Eclipse Plus C18RRHD post,
2.1mm × 100mm, 1.7 μm；Or b) chromatographic column: ACQUITY UPLC BEH C18 post, 2.1mm × 50mm, 1.7 μm；
Flowing is one of following mutually: a) by A phase and B phase composition, A phase is water, and B phase is the mixed solution of acetonitrile and methanol；
B) by A phase and B phase composition, A phase is the mixed solution of water and acetonitrile, and B phase is methanol；C) by A phase and B phase composition, A phase be water and
The mixed solution of acetonitrile, B phase is the mixed solution of acetonitrile and methanol；The volume fraction of acetonitrile in the mixed solution of acetonitrile and methanol
Being 5%, in the mixed solution of water and acetonitrile, the volume fraction of acetonitrile is 10%；
Gradient elution is for one of following: a) illustrate with the ratio of Mobile phase B: after initial volume 5% keeps 5min,
3min increases to 50% holding 3min, and 3min drops to 5% and keeps 5min；B) illustrate with the ratio of Mobile phase B: first initial body
After long-pending 5% holding 4min, 2min increases to 70% holding 5min, and 3min drops to 5% and keeps 5min；C) with the ratio of Mobile phase B
Example illustrates: after initial volume 5% keeps 4min, and 6min increases to 90% holding 1min, and 3min drops to 5% and keeps
5min；
Collecting the outflow phase of 8-11min, what the outflow obtained was evaporated concentration mutually obtains yellow powder, is blood fat reducing and lives
Property compound Quercetin-3-O-β-D-galactoside.
Beneficial effects of the present invention is as follows: be extracted the material with hypolipidemic activity, mouse test from the fine horse Fructus Jujubae of South Sinkiang
Show that it has certain hypolipidemic activity, and separating-purifying has gone out to have hypolipidemic activity compound Quercetin-3-O-β-D-
Galactoside.
Detailed description of the invention
In order to be illustrated more clearly that the present invention, below in conjunction with preferred embodiment, the present invention is described further.Ability
Field technique personnel should be appreciated that following specifically described content is illustrative and be not restrictive, and should not limit this with this
The protection domain of invention.
Part I: extract hypolipidemic activity material and hypolipidemic activity test from the fine horse Fructus Jujubae of South Sinkiang
Hypolipidemic activity material is extracted from the fine horse Fructus Jujubae of South Sinkiang
Embodiment 1
(1-1) Fructus Jujubae used by is fresh Fructus Jujubae, is collected in Alar City, Xinjiang Tarim University gardening experiment centre jujube tree of 7 years, for
The fruit fine horse Fructus Jujubae of Rhamnaceae (Rhamnaceae) zizyphus (Ziziphus Mill.) plant, separates jujube pulp with Semen Jujubae；
(1-2) with the aqueous solution of alcohol, jujube pulp is extracted, obtain alcohol extraction thing solution and solid residue, by molten for alcohol extraction thing
Liquid is concentrated to give dry extract；The aqueous solution of alcohol is ethanol water, and solid-liquid ratio is 1:3 (g/mL), the body of ethanol in ethanol water
Fraction is 70%, reflux, extract,.
(1-3) by dry extract according to solid-liquid ratio be 1:4 (g/mL) soluble in water dry extract solution, molten to dry extract with ether
Liquid extracts, and obtains ether phase and ether extracts residue aqueous phase.Ether is petroleum ether, and dry extract is 4:1 (g/ with the solid-liquid ratio of petroleum ether
mL)。
(1-4) with chloroform, ether is extracted residue aqueous phase to extract, obtain chloroform phase and chloroform extraction residue aqueous phase.Dry extract
It is 4:1 (g/mL) with the solid-liquid ratio of chloroform.
(1-5) by ethyl acetate, chloroform extraction residue aqueous phase is extracted, obtain ester phase and ester extracts residue aqueous phase；Dry leaching
Cream is 4:1 (g/mL) with the solid-liquid ratio of ethyl acetate；Ester is concentrated to give mutually dry extract, is hypolipidemic activity material.
Embodiment 2
(1-1) Fructus Jujubae used by is fresh Fructus Jujubae, is collected in Alar City, Xinjiang Tarim University gardening experiment centre jujube tree of 7 years, for
The fruit fine horse Fructus Jujubae of Rhamnaceae (Rhamnaceae) zizyphus (Ziziphus Mill.) plant, separates jujube pulp with Semen Jujubae；
(1-2) with the aqueous solution of alcohol, jujube pulp is extracted, obtain alcohol extraction thing solution and solid residue, by molten for alcohol extraction thing
Liquid is concentrated to give dry extract；The aqueous solution of alcohol is ethanol water, and solid-liquid ratio is 1:2 (g/mL), the body of ethanol in ethanol water
Fraction is 80%, reflux, extract,.
(1-3) being that 1:10 (g/mL) is soluble in water by dry extract according to solid-liquid ratio obtains dry extract solution, with ether to dry extract
Solution extracts, and obtains ether phase and ether extracts residue aqueous phase.Ether is petroleum ether, and dry extract is 8:1 (g/ with the solid-liquid ratio of petroleum ether
mL)。
(1-4) with chloroform, ether is extracted residue aqueous phase to extract, obtain chloroform phase and chloroform extraction residue aqueous phase.Dry extract
It is 8:1 (g/mL) with the solid-liquid ratio of chloroform.
(1-5) by ethyl acetate, chloroform extraction residue aqueous phase is extracted, obtain ester phase and ester extracts residue aqueous phase, dry leaching
Cream is 8:1 (g/mL) with the solid-liquid ratio of ethyl acetate；Ester is concentrated to give mutually dry extract, is hypolipidemic activity material.
Embodiment 3
(1-1) Fructus Jujubae used by is fresh Fructus Jujubae, is collected in Alar City, Xinjiang Tarim University gardening experiment centre jujube tree of 7 years, for
The fruit fine horse Fructus Jujubae of Rhamnaceae (Rhamnaceae) zizyphus (Ziziphus Mill.) plant, separates jujube pulp with Semen Jujubae；
(1-2) with the aqueous solution of alcohol, jujube pulp is extracted, obtain alcohol extraction thing solution and solid residue, by molten for alcohol extraction thing
Liquid is concentrated to give dry extract；The aqueous solution of alcohol is ethanol water, and solid-liquid ratio is 1:4 (g/mL), the body of ethanol in ethanol water
Fraction is 60%, reflux, extract,.
(1-3) by dry extract according to solid-liquid ratio be 2:5 (g/mL) soluble in water dry extract solution, molten to dry extract with ether
Liquid extracts, and obtains ether phase and ether extracts residue aqueous phase.Ether is petroleum ether, and dry extract is 2:1 (g/ with the solid-liquid ratio of petroleum ether
mL)。
(1-4) with chloroform, ether is extracted residue aqueous phase to extract, obtain chloroform phase and chloroform extraction residue aqueous phase.Dry extract
It is 2:1 (g/mL) with the solid-liquid ratio of chloroform.
(1-5) by ethyl acetate, chloroform extraction residue aqueous phase is extracted, obtain ester phase and ester extracts residue aqueous phase, dry leaching
Cream is 2:1 (g/mL) with the solid-liquid ratio of ethyl acetate；Ester is concentrated to give mutually dry extract, is hypolipidemic activity material.
Hypolipidemic activity is tested
1, material
Animal: Kunming white mice, average weight (20.0 ± 2.0) g, SPF level, production licence: SCXK (newly) 2011-
0001, Xinjiang Disease Control and Prevention Center animal experimental center all it is purchased from normal feedstuff.Freely drink water and ingest.
2, reagent
Test kit: serum total cholesterol test kit, high density lipoprotein test kit, Triglyceride Reagent box, low density lipoprotein
Protein reagent box, chemical pure cholesterol (Changchun Hui Li Bioisystech Co., Ltd).
Other: simvastatin is commercially available；High lipid food (83.0% normal feedstuff, 10% yolk powder, 1.5% cholesterol, 5%
Adeps Sus domestica, 0.5% NaTDC).The reagent such as chloroform, dehydrated alcohol is analytical reagent, is all purchased from Tianjin and causes remote chemistry examination
Agent company limited.
3, method
3.1, hypolipidemic activity material is configured to the sample solution that concentration is 25mg/Kg, is divided into high and low dose group simultaneously
(0.6mL, 0.2mL).
3.2, mice group and feeding
Choose 50 kunming mices, male and female half and half, be equally divided into 5 groups (often groups 10) by body weight equilibrium:
Blank group, raises with normal feedstuff.
Positive controls, raises with high lipid food.
Hyperlipidemia model matched group, raises with high lipid food.
Low dosage test group, raises with high lipid food.
High dose test group, raises with high lipid food.
Within every 10 days, carry out body weight and blood fat dynamic tracking, until modeling success.Then, hyperlipidemia model matched group stops feeding
High lipid food, changes and raises normal feedstuff；1/12 simvastatin solution of positive controls gavage human body dose；Low dosage is tested
Group and high dose test group start to feed various dose sample solution, low dosage test group feeding every day 0.2mL sample solution, height
Dosetest group feeding every day 0.6mL sample solution.
3.3, mice serum preparation and index determining
The preparation of serum: take blood in every 10 days, takes mice fasting 12h before blood, takes mouse tail vein blood, after standing 30min,
10000r/min is centrifuged 15min, and serum moves to clean blood test tube.
Index determining: mice serum T-CHOL (TC), triglyceride (TG), HDL-C (HDL-C)
And low density lipoprotein, LDL (LDL-C) assay uses method well known in the prior art.
It is calculated as follows atherogenic index: atherogenic index=(TC-HDL-C)/(HDL-C).
4, result
Feeding mice through 30 days, mice serum T-CHOL (TC), triglyceride (TG), high density lipoprotein gallbladder are solid
Alcohol (HDL-C) and low density lipoprotein, LDL (LDL-C) are as shown in the table.
Note: * represents the significance compared with hyperlipidemia model group, *: p < 0.05, * *: p < 0.01；# represents compared with blank group
Significance, #:p < 0.05, ##:p < 0.01.
Part II: separate and purification Quercetin-3-O-β-D-galactoside
Separate and purification Quercetin-3-O-β-D-galactoside
Embodiment 4
(2-1) cross the first silicagel column by embodiment 1 is extracted the hypolipidemic activity material obtained, eluant be petroleum ether and
The volume ratio of the mixture of acetone, petroleum ether and acetone is 5:1；Described first silicagel column is gross porosity zcx-II type silicagel column, outward
Footpath is 4.5 centimetres, and eluant flow velocity is 5mL/min；
(2-2) eluent obtained in step (2-1) being crossed the second silicagel column, eluant is the mixing of chloroform and methanol
The volume ratio of thing, chloroform and methanol is 5:1；Described second silicagel column is gross porosity zcx-II type silicagel column, and external diameter is 4 centimetres, washes
De-agent flow velocity is 1mL/min；
(2-3) eluent obtained in step (2-2) being crossed gel column, eluant is the mixture of chloroform and methanol, chlorine
Imitative and methanol volume ratio is 1:1, and every 10mL collects a pipe, is collected to the 123rd pipe eluent by the 41st pipe and merges；Described gel
Post is Sephadex LH-20 type gel column, and external diameter is 2.5 centimetres, and eluant flow velocity is 0.5mL/min；
(2-4) the 41st pipe to the 123rd pipe eluent obtained in step (2-3) being crossed middle compression leg, eluant is methanol-water
Solution, the volume fraction of methanol is 70%, and every 10mL collects a pipe, is collected to the 53rd pipe eluent by the 14th pipe and merges；Described
Middle compression leg external diameter is 4 centimetres, and post pressure is 3MPa, and eluant flow velocity is 3mL/min；
(2-5) the 14th pipe to the 53rd pipe eluent obtained in step (2-4) is crossed performance liquid chromatographic column, collect eluting
Liquid, what eluent was evaporated concentration obtains yellow powder, is hypolipidemic activity compound Quercetin-3-O-β-D-galactoside.
Performance liquid chromatographic column is: a) chromatographic column: Agilent Eclipse Plus C18RRHD post, 2.1mm ×
100mm, 1.7 μm.
Flowing is mutually: a) by A phase and B phase composition, A phase is water, and B phase is the mixed solution of acetonitrile and methanol, acetonitrile and first
In the mixed solution of alcohol, the volume fraction of acetonitrile is 5%.
Gradient elution: a) illustrate with the ratio of Mobile phase B: after initial volume 5% keeps 5min, 3min increases to 50%
3min, 3min is kept to drop to 5% and keep 5min.
Collecting the outflow phase of 8-11min, what the outflow obtained was evaporated concentration mutually obtains yellow powder, gained yellow powder
The purity of middle hypolipidemic activity compound Quercetin-3-O-β-D-galactoside is 89wt%.
Embodiment 5
The present embodiment is from the difference of embodiment 4: chromatographic column is different, the present embodiment following chromatographic column of employing: b) chromatograph
Post: ACQUITY UPLC BEH C18 post, 2.1mm × 50mm, 1.7 μm.Hypolipidemic activity compound Mongolian oak in gained yellow powder
The purity of Pi Su-3-O-β-D-galactoside is 80wt%.
In embodiment 4 and embodiment 5, it is preferred to use the chromatographic column used in embodiment 4, Quercetin-3-O-β-D-gala
Separate between glucosides with other composition and chromatographic peak peak shape is relatively good.And in embodiment 5, Quercetin-3-O-β-D-gala
Separating effect between glucosides and other composition is poor, and chromatographic peak hangover, and product purity and yield all decline.
Embodiment 6
The present embodiment is from the difference of embodiment 4: flow mutually different, and b) by A phase and B phase composition, A phase is water and acetonitrile
Mixed solution, B phase be methanol, water and acetonitrile mixed solution in the volume fraction of acetonitrile be 10%.In gained yellow powder
The purity of hypolipidemic activity compound Quercetin-3-O-β-D-galactoside is 90wt%.
Embodiment 7
The present embodiment is from the difference of embodiment 4: flow mutually different, and c) by A phase and B phase composition, A phase is water and acetonitrile
Mixed solution, B phase is the mixed solution of acetonitrile and methanol；In the mixed solution of acetonitrile and methanol, the volume fraction of acetonitrile is
5%, in the mixed solution of water and acetonitrile, the volume fraction of acetonitrile is 10%.Hypolipidemic activity compound Mongolian oak in gained yellow powder
The purity of Pi Su-3-O-β-D-galactoside is 98wt%.
In embodiment 4, embodiment 6 and embodiment 7, the flowing phase being preferably used in embodiment 7: Quercetin-3-O-β-D-
The chromatographic peak peak shape of galactoside is good, and the response to Quercetin-3-O-β-D-galactoside is relatively big, with the guarantor of other composition
Staying the time is difference maximum in these three flowing mutually, is conducive to isolating the Quercetin-3-O-β-D-galactose that purity is higher
Glycosides.
Embodiment 8
The present embodiment is from the difference of embodiment 7: gradient is different；B) illustrate with the ratio of Mobile phase B: b)
Illustrating with the ratio of Mobile phase B: after initial volume 5% keeps 4min, 2min increases to 70% holding 5min, and 3min drops to
5% and keep 5min.In gained yellow powder, the purity of hypolipidemic activity compound Quercetin-3-O-β-D-galactoside is
93wt%.
Embodiment 9
The present embodiment is from the difference of embodiment 7: gradient is different, c) illustrates with the ratio of Mobile phase B: just
After initial body amasss 5% holding 4min, 6min increases to 90% holding 1min, and 3min drops to 5% and keeps 5min.Gained yellow powder
The purity of middle hypolipidemic activity compound Quercetin-3-O-β-D-galactoside is 92wt%.
In embodiment 7, embodiment 8 and embodiment 9, it is preferred to use the gradient in embodiment 7: be able to ensure that Cortex querci dentatae
Element-3-O-β-D-galactoside separates well with other composition, and retention time is the most stable.
The chemical structural formula of Quercetin-3-O-β-D-galactoside is as follows:
Select the yellow powder obtained in embodiment 7 to be characterized as below: hydrochloric acid-magnesium powder reacting positive, be dissolved in methanol,
A1C13It is that yellow is dashed forward light at 365nm after colour developing.1H-NMR (500MHz, MeOH-d6) δ (ppm): 7.86 (1H, dd, J=2.1Hz,
H-2 '), 7.61 (1H, dd, J=8.5,2.1Hz, H-6 '), 6.89 (1H, d, J=8.4Hz, H-5 '), 6.43 (1H, d, J=
2.1Hz, H-8 '), 6.23 (1H, d, J=2.1Hz, H-6 '), 5.19 (1H, d, J=7.8Hz, H-1 "), 4.87 (1H, d, J=
3.4Hz, H-3 "), sugar moieties other protons δ (ppm):
3.33～3.84；13C-NMR (125MHz, MeOH-d6) δ (ppm): 177.5 (C-4), 165.3 (C-7), 162.7
(C-5),159.4(C-9),155.9(C-2),48.5(C-4′),145.1(C-3′),133.6(C-3),122.03(C-6′),
121.1(C-1′),116.1(C-5′),115.5(C-2′),103.9(C-10),103.9(C-1″),98.8(C-6),93.3(C-
8),75.9(C-5″),3.3(C-3″),71.6(C-2″),68.2(C-4″),60.3(C-6″).With Quercetin in prior art-
The test result of 3-O-β-D-galactoside is consistent.
Hypolipidemic activity is tested
Blood fat reducing test method is identical with the hypolipidemic activity test in the embodiment of the present invention " Part I ", result table
Bright: mouse feeding Quercetin-3-O-β-D-galactoside is after 2 days, i.e. can obviously reduce T-CHOL in mice serum (TC), sweet
Oil three esters (TG), the level of low-density lipoprotein cholesterol (LDL-C), and improve HDL-C (HDL-C)
Level.
Obviously, the above embodiment of the present invention is only for clearly demonstrating example of the present invention, and is not right
The restriction of embodiments of the present invention, for those of ordinary skill in the field, the most also may be used
To make other changes in different forms, cannot all of embodiment be given exhaustive here, every belong to this
What bright technical scheme was extended out obviously changes or changes the row still in protection scope of the present invention.
Claims (10)
1. extract and the side of isolated and purified hypolipidemic activity compound Quercetin-3-O-β-D-galactoside from the fine horse Fructus Jujubae of South Sinkiang
Method, it is characterised in that comprise the steps:
(I) South Sinkiang fine horse Fructus Jujubae extracts hypolipidemic activity material；
(1-1) jujube pulp is separated with Semen Jujubae；
(1-2) with alcoholic solution, jujube pulp is extracted, obtain alcohol extraction thing solution and solid residue, alcohol extraction thing solution is concentrated to give
Dry extract；
(II) separation of Quercetin-3-O-β-D-galactoside and purification.
Extraction and isolated and purified hypolipidemic activity compound Quercetin-3-from the fine horse Fructus Jujubae of South Sinkiang the most according to claim 1
The method of O-β-D-galactoside, it is characterised in that in step (I), also comprise the steps:
(1-3) being that 1:4g/mL is soluble in water by dry extract according to solid-liquid ratio obtains dry extract solution, carries out dry extract solution with ether
Extraction, obtains ether phase and ether extracts residue aqueous phase.
Extraction and isolated and purified hypolipidemic activity compound Quercetin-3-from the fine horse Fructus Jujubae of South Sinkiang the most according to claim 2
The method of O-β-D-galactoside, it is characterised in that also comprise the steps:
(1-4) with chloroform, ether is extracted residue aqueous phase to extract, obtain chloroform phase and chloroform extraction residue aqueous phase.
Extraction and isolated and purified hypolipidemic activity compound Quercetin-3-from the fine horse Fructus Jujubae of South Sinkiang the most according to claim 3
The method of O-β-D-galactoside, it is characterised in that also comprise the steps:
(1-5) with ester, chloroform extraction residue aqueous phase is extracted, obtain ester phase and ester extracts residue aqueous phase.
Extraction and isolated and purified hypolipidemic activity compound Quercetin-3-from the fine horse Fructus Jujubae of South Sinkiang the most according to claim 4
The method of O-β-D-galactoside, it is characterised in that in step (1-2): the aqueous solution of alcohol is ethanol water, solid-liquid ratio is 1:
(2～4) g/mL, in ethanol water, the volume fraction of ethanol is 60-80%, reflux, extract,.
Extraction and isolated and purified hypolipidemic activity compound Quercetin-3-from the fine horse Fructus Jujubae of South Sinkiang the most according to claim 5
The solid-liquid ratio of the method for O-β-D-galactoside, it is characterised in that in step (1-3): ether is petroleum ether, dry extract and petroleum ether
For 4:(0.5～2) g/mL.
Extraction and isolated and purified hypolipidemic activity compound Quercetin-3-from the fine horse Fructus Jujubae of South Sinkiang the most according to claim 6
The method of O-β-D-galactoside, it is characterised in that in step (1-4): dry extract is 4:(0.5～2 with the solid-liquid ratio of chloroform)
g/mL。
Extraction and isolated and purified hypolipidemic activity compound Quercetin-3-from the fine horse Fructus Jujubae of South Sinkiang the most according to claim 7
The material of the method for O-β-D-galactoside, it is characterised in that in step (1-5): ester is ethyl acetate, dry extract and ethyl acetate
Liquor ratio is 4:(0.5～2) g/mL；The ethyl acetate obtained is concentrated mutually and obtains hypolipidemic activity material afterwards.
9. according to arbitrary described the extracting and isolated and purified hypolipidemic activity compound Cortex querci dentatae from the fine horse Fructus Jujubae of South Sinkiang of claim 1-8
The method of element-3-O-β-D-galactoside, it is characterised in that in step (II), comprise the steps:
(2-1) crossing the first silicagel column by extracting the hypolipidemic activity material obtained in step (I), eluant is petroleum ether and acetone
Mixture, the volume ratio of petroleum ether and acetone is 5:1；
(2-2) eluent obtained in step (2-1) being crossed the second silicagel column, eluant is the mixture of chloroform and methanol, chlorine
Imitative and methanol volume ratio is 5:1；
(2-3) eluent obtained in step (2-2) being crossed gel column, eluant is the mixture of chloroform and methanol, chloroform and
The volume ratio of methanol is 1:1, and every 10mL collects a pipe, is collected to the 123rd pipe eluent by the 41st pipe and merges；
(2-4) the 41st pipe to the 123rd pipe eluent obtained in step (2-3) being crossed middle compression leg, eluant is methanol aqueous solution,
The volume fraction of methanol is 70%, and every 10mL collects a pipe, is collected to the 53rd pipe eluent by the 14th pipe and merges；
(2-5) the 14th pipe to the 53rd pipe eluent obtained in step (2-4) is crossed performance liquid chromatographic column, collects eluent,
What eluent was evaporated concentration obtains yellow powder, is hypolipidemic activity compound Quercetin-3-O-β-D-galactoside.
Extraction and isolated and purified hypolipidemic activity compound Quercetin-3-from the fine horse Fructus Jujubae of South Sinkiang the most according to claim 9
The method of O-β-D-galactoside, it is characterised in that
In step (2-1): described first silicagel column is gross porosity zcx-II type silicagel column, external diameter is 4.5 centimetres, and eluant flow velocity is
5-10mL/min；
In step (2-2): described first silicagel column is gross porosity zcx-II type silicagel column, external diameter is 4 centimetres, and eluant flow velocity is 1-
2mL/min；
In step (2-3): described gel column is Sephadex LH-20 type gel column, and external diameter is 2.5 centimetres, and eluant flow velocity is
0.5-1mL/min；
In step (2-4): medium pressure post external diameter is 4 centimetres, post pressure is 3-4MPa, and eluant flow velocity is 3-4mL/min；
In step (2-5):
Performance liquid chromatographic column is one of following: a) chromatographic column: Agilent Eclipse Plus C18RRHD post, 2.1mm ×
100mm, 1.7 μm；Or b) chromatographic column: ACQUITY UPLC BEH C18 post, 2.1mm × 50mm, 1.7 μm；
Flowing is one of following mutually: a) by A phase and B phase composition, A phase is water, and B phase is the mixed solution of acetonitrile and methanol；B) by A
Mutually with B phase composition, A phase is the mixed solution of water and acetonitrile, and B phase is methanol；C) by A phase and B phase composition, A phase is water and acetonitrile
Mixed solution, B phase is the mixed solution of acetonitrile and methanol；In the mixed solution of acetonitrile and methanol, the volume fraction of acetonitrile is
5%, in the mixed solution of water and acetonitrile, the volume fraction of acetonitrile is 10%；
Gradient elution is for one of following: a) illustrating with the ratio of Mobile phase B: after initial volume 5% keeps 5min, 3min increases
3min, 3min is kept to drop to 5% and keep 5min to 50%；B) illustrate with the ratio of Mobile phase B: initial volume 5%
After keeping 4min, 2min increases to 70% holding 5min, and 3min drops to 5% and keeps 5min；C) carry out with the ratio of Mobile phase B
Illustrating: after initial volume 5% keeps 4min, 6min increases to 90% holding 1min, and 3min drops to 5% and keeps 5min；
Collecting the outflow phase of 8-11min, what the outflow obtained was evaporated concentration mutually obtains yellow powder, is hypolipidemic activity
Compound Quercetin-3-O-β-D-galactoside.
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