CN106045958A - Method for separating and purifying myricetin and dihydromyricetin from ampelopsis grossedentata - Google Patents
Method for separating and purifying myricetin and dihydromyricetin from ampelopsis grossedentata Download PDFInfo
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- CN106045958A CN106045958A CN201610621955.8A CN201610621955A CN106045958A CN 106045958 A CN106045958 A CN 106045958A CN 201610621955 A CN201610621955 A CN 201610621955A CN 106045958 A CN106045958 A CN 106045958A
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- Prior art keywords
- dihydromyricetin
- supernatant
- ampelopsis grossedentata
- ampelopsin
- high speed
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- KJXSIXMJHKAJOD-LSDHHAIUSA-N (+)-dihydromyricetin Chemical compound C1([C@@H]2[C@H](C(C3=C(O)C=C(O)C=C3O2)=O)O)=CC(O)=C(O)C(O)=C1 KJXSIXMJHKAJOD-LSDHHAIUSA-N 0.000 title claims abstract description 238
- KQILIWXGGKGKNX-UHFFFAOYSA-N dihydromyricetin Natural products OC1C(=C(Oc2cc(O)cc(O)c12)c3cc(O)c(O)c(O)c3)O KQILIWXGGKGKNX-UHFFFAOYSA-N 0.000 title claims abstract description 118
- 238000000034 method Methods 0.000 title claims abstract description 52
- 241001018563 Nekemias grossedentata Species 0.000 title claims abstract description 47
- IKMDFBPHZNJCSN-UHFFFAOYSA-N Myricetin Chemical compound C=1C(O)=CC(O)=C(C(C=2O)=O)C=1OC=2C1=CC(O)=C(O)C(O)=C1 IKMDFBPHZNJCSN-UHFFFAOYSA-N 0.000 title abstract 5
- 229940116852 myricetin Drugs 0.000 title abstract 5
- PCOBUQBNVYZTBU-UHFFFAOYSA-N myricetin Natural products OC1=C(O)C(O)=CC(C=2OC3=CC(O)=C(O)C(O)=C3C(=O)C=2)=C1 PCOBUQBNVYZTBU-UHFFFAOYSA-N 0.000 title abstract 5
- 235000007743 myricetin Nutrition 0.000 title abstract 5
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 claims abstract description 48
- 239000000284 extract Substances 0.000 claims abstract description 42
- 239000000843 powder Substances 0.000 claims abstract description 32
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims abstract description 30
- 239000000706 filtrate Substances 0.000 claims abstract description 17
- 239000007788 liquid Substances 0.000 claims abstract description 15
- 238000003756 stirring Methods 0.000 claims abstract description 12
- 239000013049 sediment Substances 0.000 claims abstract 3
- XEKOWRVHYACXOJ-UHFFFAOYSA-N Ethyl acetate Chemical compound CCOC(C)=O XEKOWRVHYACXOJ-UHFFFAOYSA-N 0.000 claims description 138
- 239000006228 supernatant Substances 0.000 claims description 87
- 238000001556 precipitation Methods 0.000 claims description 69
- 238000000703 high-speed centrifugation Methods 0.000 claims description 59
- 238000000605 extraction Methods 0.000 claims description 39
- UIIMBOGNXHQVGW-UHFFFAOYSA-M Sodium bicarbonate Chemical compound [Na+].OC([O-])=O UIIMBOGNXHQVGW-UHFFFAOYSA-M 0.000 claims description 28
- 239000013078 crystal Substances 0.000 claims description 26
- 238000001291 vacuum drying Methods 0.000 claims description 19
- 238000000926 separation method Methods 0.000 claims description 15
- 229910000030 sodium bicarbonate Inorganic materials 0.000 claims description 14
- 235000017557 sodium bicarbonate Nutrition 0.000 claims description 14
- 239000002244 precipitate Substances 0.000 claims description 13
- 230000009514 concussion Effects 0.000 claims description 11
- 238000000746 purification Methods 0.000 claims description 11
- VEXZGXHMUGYJMC-UHFFFAOYSA-N Hydrochloric acid Chemical compound Cl VEXZGXHMUGYJMC-UHFFFAOYSA-N 0.000 claims description 10
- 238000002425 crystallisation Methods 0.000 claims description 10
- 230000008025 crystallization Effects 0.000 claims description 10
- 238000000151 deposition Methods 0.000 claims description 7
- 230000008021 deposition Effects 0.000 claims description 7
- 229920002678 cellulose Polymers 0.000 claims description 6
- 239000001913 cellulose Substances 0.000 claims description 6
- 230000001376 precipitating effect Effects 0.000 claims description 6
- KGBXLFKZBHKPEV-UHFFFAOYSA-N boric acid Chemical compound OB(O)O KGBXLFKZBHKPEV-UHFFFAOYSA-N 0.000 claims description 5
- 239000004327 boric acid Substances 0.000 claims description 5
- 244000144992 flock Species 0.000 claims description 5
- 239000000203 mixture Substances 0.000 claims description 5
- 238000002360 preparation method Methods 0.000 claims description 5
- 238000001816 cooling Methods 0.000 claims description 2
- 239000000126 substance Substances 0.000 claims description 2
- XYIBRDXRRQCHLP-UHFFFAOYSA-N ethyl acetoacetate Chemical compound CCOC(=O)CC(C)=O XYIBRDXRRQCHLP-UHFFFAOYSA-N 0.000 claims 1
- 239000002904 solvent Substances 0.000 abstract description 4
- 238000010992 reflux Methods 0.000 abstract description 3
- 238000001035 drying Methods 0.000 abstract description 2
- 238000003912 environmental pollution Methods 0.000 abstract description 2
- 238000004064 recycling Methods 0.000 abstract description 2
- 235000019441 ethanol Nutrition 0.000 abstract 3
- 238000001914 filtration Methods 0.000 abstract 1
- 230000005484 gravity Effects 0.000 abstract 1
- 238000009776 industrial production Methods 0.000 abstract 1
- 238000007873 sieving Methods 0.000 abstract 1
- 238000002791 soaking Methods 0.000 abstract 1
- 239000003053 toxin Substances 0.000 abstract 1
- 231100000765 toxin Toxicity 0.000 abstract 1
- 238000005303 weighing Methods 0.000 abstract 1
- 239000000243 solution Substances 0.000 description 21
- OKKJLVBELUTLKV-UHFFFAOYSA-N Methanol Chemical compound OC OKKJLVBELUTLKV-UHFFFAOYSA-N 0.000 description 12
- 230000001476 alcoholic effect Effects 0.000 description 6
- 238000002955 isolation Methods 0.000 description 6
- UIIMBOGNXHQVGW-DEQYMQKBSA-M Sodium bicarbonate-14C Chemical compound [Na+].O[14C]([O-])=O UIIMBOGNXHQVGW-DEQYMQKBSA-M 0.000 description 4
- 239000012535 impurity Substances 0.000 description 4
- 238000004321 preservation Methods 0.000 description 4
- 239000013558 reference substance Substances 0.000 description 4
- 238000012360 testing method Methods 0.000 description 4
- 241000196324 Embryophyta Species 0.000 description 3
- LRHPLDYGYMQRHN-UHFFFAOYSA-N N-Butanol Chemical compound CCCCO LRHPLDYGYMQRHN-UHFFFAOYSA-N 0.000 description 3
- 241000219000 Populus Species 0.000 description 3
- GAMYVSCDDLXAQW-AOIWZFSPSA-N Thermopsosid Natural products O(C)c1c(O)ccc(C=2Oc3c(c(O)cc(O[C@H]4[C@H](O)[C@@H](O)[C@H](O)[C@H](CO)O4)c3)C(=O)C=2)c1 GAMYVSCDDLXAQW-AOIWZFSPSA-N 0.000 description 3
- 238000005119 centrifugation Methods 0.000 description 3
- 238000004090 dissolution Methods 0.000 description 3
- 239000003814 drug Substances 0.000 description 3
- 238000005516 engineering process Methods 0.000 description 3
- 229930003944 flavone Natural products 0.000 description 3
- 235000011949 flavones Nutrition 0.000 description 3
- 235000013305 food Nutrition 0.000 description 3
- 238000002156 mixing Methods 0.000 description 3
- 238000005498 polishing Methods 0.000 description 3
- 239000000047 product Substances 0.000 description 3
- 208000006379 syphilis Diseases 0.000 description 3
- VHBFFQKBGNRLFZ-UHFFFAOYSA-N vitamin p Natural products O1C2=CC=CC=C2C(=O)C=C1C1=CC=CC=C1 VHBFFQKBGNRLFZ-UHFFFAOYSA-N 0.000 description 3
- 244000269722 Thea sinensis Species 0.000 description 2
- 230000008901 benefit Effects 0.000 description 2
- 238000005265 energy consumption Methods 0.000 description 2
- 150000002212 flavone derivatives Chemical class 0.000 description 2
- 238000004128 high performance liquid chromatography Methods 0.000 description 2
- 231100000252 nontoxic Toxicity 0.000 description 2
- 230000003000 nontoxic effect Effects 0.000 description 2
- 239000003960 organic solvent Substances 0.000 description 2
- 231100000614 poison Toxicity 0.000 description 2
- 238000011160 research Methods 0.000 description 2
- 241000563984 Ampelopsis Species 0.000 description 1
- 241000345998 Calamus manan Species 0.000 description 1
- 206010013911 Dysgeusia Diseases 0.000 description 1
- 206010017553 Furuncle Diseases 0.000 description 1
- 235000009388 Parthenocissus quinquefolia Nutrition 0.000 description 1
- 235000011158 Prunus mume Nutrition 0.000 description 1
- 244000018795 Prunus mume Species 0.000 description 1
- 206010037660 Pyrexia Diseases 0.000 description 1
- 241000219094 Vitaceae Species 0.000 description 1
- 230000036592 analgesia Effects 0.000 description 1
- 238000004458 analytical method Methods 0.000 description 1
- 230000003110 anti-inflammatory effect Effects 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- 208000002352 blister Diseases 0.000 description 1
- 238000007664 blowing Methods 0.000 description 1
- 210000000988 bone and bone Anatomy 0.000 description 1
- 238000004364 calculation method Methods 0.000 description 1
- 239000003153 chemical reaction reagent Substances 0.000 description 1
- 239000003795 chemical substances by application Substances 0.000 description 1
- 238000011109 contamination Methods 0.000 description 1
- 230000006378 damage Effects 0.000 description 1
- 238000013461 design Methods 0.000 description 1
- 238000001514 detection method Methods 0.000 description 1
- 238000010790 dilution Methods 0.000 description 1
- 239000012895 dilution Substances 0.000 description 1
- 201000010099 disease Diseases 0.000 description 1
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 description 1
- 229940079593 drug Drugs 0.000 description 1
- 230000009977 dual effect Effects 0.000 description 1
- 230000000694 effects Effects 0.000 description 1
- 238000005485 electric heating Methods 0.000 description 1
- 238000010828 elution Methods 0.000 description 1
- 238000002474 experimental method Methods 0.000 description 1
- -1 flavone compound Chemical class 0.000 description 1
- 210000003026 hypopharynx Anatomy 0.000 description 1
- 230000001965 increasing effect Effects 0.000 description 1
- 239000004615 ingredient Substances 0.000 description 1
- 239000007791 liquid phase Substances 0.000 description 1
- 238000004519 manufacturing process Methods 0.000 description 1
- 230000035800 maturation Effects 0.000 description 1
- 238000000874 microwave-assisted extraction Methods 0.000 description 1
- 210000003205 muscle Anatomy 0.000 description 1
- 230000036407 pain Effects 0.000 description 1
- 239000012071 phase Substances 0.000 description 1
- 230000007096 poisonous effect Effects 0.000 description 1
- NHOXRBDMOTVJBL-UHFFFAOYSA-M potassium;dihydrogen phosphate;methanol Chemical compound [K+].OC.OP(O)([O-])=O NHOXRBDMOTVJBL-UHFFFAOYSA-M 0.000 description 1
- 235000012950 rattan cane Nutrition 0.000 description 1
- 239000002994 raw material Substances 0.000 description 1
- 230000007115 recruitment Effects 0.000 description 1
- 239000011347 resin Substances 0.000 description 1
- 229920005989 resin Polymers 0.000 description 1
- 230000001568 sexual effect Effects 0.000 description 1
- 239000007790 solid phase Substances 0.000 description 1
- 238000000638 solvent extraction Methods 0.000 description 1
- 238000005728 strengthening Methods 0.000 description 1
- 238000000194 supercritical-fluid extraction Methods 0.000 description 1
- 230000008961 swelling Effects 0.000 description 1
- 239000003440 toxic substance Substances 0.000 description 1
- 238000002604 ultrasonography Methods 0.000 description 1
- 238000000825 ultraviolet detection Methods 0.000 description 1
Classifications
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07D—HETEROCYCLIC COMPOUNDS
- C07D311/00—Heterocyclic compounds containing six-membered rings having one oxygen atom as the only hetero atom, condensed with other rings
- C07D311/02—Heterocyclic compounds containing six-membered rings having one oxygen atom as the only hetero atom, condensed with other rings ortho- or peri-condensed with carbocyclic rings or ring systems
- C07D311/04—Benzo[b]pyrans, not hydrogenated in the carbocyclic ring
- C07D311/22—Benzo[b]pyrans, not hydrogenated in the carbocyclic ring with oxygen or sulfur atoms directly attached in position 4
- C07D311/26—Benzo[b]pyrans, not hydrogenated in the carbocyclic ring with oxygen or sulfur atoms directly attached in position 4 with aromatic rings attached in position 2 or 3
- C07D311/28—Benzo[b]pyrans, not hydrogenated in the carbocyclic ring with oxygen or sulfur atoms directly attached in position 4 with aromatic rings attached in position 2 or 3 with aromatic rings attached in position 2 only
- C07D311/32—2,3-Dihydro derivatives, e.g. flavanones
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07D—HETEROCYCLIC COMPOUNDS
- C07D311/00—Heterocyclic compounds containing six-membered rings having one oxygen atom as the only hetero atom, condensed with other rings
- C07D311/02—Heterocyclic compounds containing six-membered rings having one oxygen atom as the only hetero atom, condensed with other rings ortho- or peri-condensed with carbocyclic rings or ring systems
- C07D311/04—Benzo[b]pyrans, not hydrogenated in the carbocyclic ring
- C07D311/22—Benzo[b]pyrans, not hydrogenated in the carbocyclic ring with oxygen or sulfur atoms directly attached in position 4
- C07D311/26—Benzo[b]pyrans, not hydrogenated in the carbocyclic ring with oxygen or sulfur atoms directly attached in position 4 with aromatic rings attached in position 2 or 3
- C07D311/28—Benzo[b]pyrans, not hydrogenated in the carbocyclic ring with oxygen or sulfur atoms directly attached in position 4 with aromatic rings attached in position 2 or 3 with aromatic rings attached in position 2 only
- C07D311/30—Benzo[b]pyrans, not hydrogenated in the carbocyclic ring with oxygen or sulfur atoms directly attached in position 4 with aromatic rings attached in position 2 or 3 with aromatic rings attached in position 2 only not hydrogenated in the hetero ring, e.g. flavones
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07D—HETEROCYCLIC COMPOUNDS
- C07D311/00—Heterocyclic compounds containing six-membered rings having one oxygen atom as the only hetero atom, condensed with other rings
- C07D311/02—Heterocyclic compounds containing six-membered rings having one oxygen atom as the only hetero atom, condensed with other rings ortho- or peri-condensed with carbocyclic rings or ring systems
- C07D311/04—Benzo[b]pyrans, not hydrogenated in the carbocyclic ring
- C07D311/22—Benzo[b]pyrans, not hydrogenated in the carbocyclic ring with oxygen or sulfur atoms directly attached in position 4
- C07D311/26—Benzo[b]pyrans, not hydrogenated in the carbocyclic ring with oxygen or sulfur atoms directly attached in position 4 with aromatic rings attached in position 2 or 3
- C07D311/40—Separation, e.g. from natural material; Purification
Landscapes
- Chemical & Material Sciences (AREA)
- Organic Chemistry (AREA)
- Pharmaceuticals Containing Other Organic And Inorganic Compounds (AREA)
- Compounds Of Unknown Constitution (AREA)
Abstract
The invention discloses a method for separating and purifying myricetin and dihydromyricetin from ampelopsis grossedentata. The method comprises the following steps: weighing, finely selecting and crushing the ampelopsis grossedentata, and sieving ampelopsis grossedentata powder through a 35-45-mesh sieve; adding ethyl alcohol and soaking the sieved ampelopsis grossedentata powder for 6 to 10 hours, and then, performing reflux extracting for 3 to 5 hours; filtering an extracting liquid to obtain a filtrate; recycling the ethyl alcohol from the filtrate until the content of the ethyl alcohol in the filtrate is 15 percent or less, and then, performing vacuum concentrating until an extractum with the specific gravity of 1.0 to 1.5 is obtained; fully stirring concentrated extractum with hot water, performing ultrasonic vibrating and high speed centrifuging, and collecting supernate; refrigerating the obtained supernate overnight, performing high speed centrifuging, collecting sediment, and drying the sediment in vacuum to obtain a yellow-green powder extract; separating and purifying the powder extract to obtain pure myricetin and pure dihydromyricetin with the high purity of 98 percent or above. According to the method, the equipment is simple, the process is simple and feasible, and industrial production is facilitated; the used solvent has no toxin and residue, so that environmental pollution cannot be caused; the myricetin and the dihydromyricetin are separated and purified from the ampelopsis grossedentata by utilizing the method, so that the utilization rate is high, the yield is high, and the purity of the obtained myricetin and dihydromyricetin is high.
Description
Technical field
The present invention relates to technical field of Chinese medicine, be more particularly to a kind of separating-purifying ampelopsin and dihydro poplar from Ampelopsis grossedentata
The method of syphilis.
Background technology
Ampelopsis grossedentata belongs to Vitaceae, Ampelopsis, and formal name used at school is Ampelopsis grossedentata (Ampelopsis grossedentata), taste
Sweet, light, cool in nature, there is the effects such as heat-clearing and toxic substances removing, wind-damp dispelling, bone and muscle strengthening, antiinflammatory, analgesia.Among the people by its children young stem and leaf make guarantor
Strong tea, for treating the history of the existing centuries of the diseases such as cold, fever, laryngopharynx swelling and pain, icterohepatitis, bleb furuncle, is a kind of typical case
Medicine food dual purpose plant.Ampelopsis grossedentata principle active component is flavone compound, and with dihydromyricetin
(Dihydromyricetin is called for short DMY) content is the highest.Research shows, in Ampelopsis grossedentata, general flavone content is 43.4%～44.0%
(mass fraction), dihydromyricetin cellulose content is up to 37.4%～40%, but relevant with factors such as geography, climatic environments.In recent years,
Have some scholars both at home and abroad and carried out the research extracting dihydromyricetin from plant, it is common that with organic solvent for extracting
Agent, such as ethyl acetate, n-butyl alcohol, ethanol etc..This method extraction cost is high, and the energy consumption of recycling design is high, there is also time-consuming, carry
The weak points such as the rate that takes is low, poor reproducibility.And solvent for use is the most poisonous, dangerous, easily environment and operator are caused
Harm.Though supercritical extraction dihydromyricetin has saving reagent, the advantage such as pollution-free, but the response rate is poor, and super in order to obtain
Critical condition, equipment one-time investment is relatively big, and operating cost is high.Additionally, also macroreticular resin absorbing method, microwave extraction method, inverse
Stream method, ethanol auxiliary law and ultrasonic assistant solvent extraction method.
Summary of the invention
(1) to solve the technical problem that
The technical problem to be solved in the present invention is how low cost, low energy consumption, extraction rattan safe and nontoxic, free of contamination
Effective ingredient in tea, and a kind of separating-purifying ampelopsin and method of dihydromyricetin from Ampelopsis grossedentata is provided
(2) technical scheme
In order to solve above-mentioned technical problem, the invention provides a kind of separating-purifying ampelopsin and dihydromyricetin from Ampelopsis grossedentata
The method (raw materials used equal city available from) of element, the method comprises the steps:
Step one: preparation: weigh Ampelopsis grossedentata, selected, pulverize, cross 35-45 mesh sieve；Too thin, too thick extraction efficiency is the highest；
Step 2: extraction: add the ethanol of the 75-85% of Ampelopsis grossedentata weight 18-22 times, with 5% sodium bicarbonate solution regulation
PH value is 7.0-8.0, soaks 6-10 hour, then backflow extraction 3-5 hour at a temperature of 35-45 DEG C；Alkalescence condition is conducive to
Extract；
Step 3: secondary extracts: add the ethanol of the 75-85% of Ampelopsis grossedentata weight 8-12 times, refluxes at a temperature of 35-45 DEG C
Extract 3-5 hour；
Step 4: filter: merge twice extraction liquid, be filtrated to get filtrate；Filter out impurity；
Step 5: recovered alcohol: recovered alcohol is less than 15% to alcohol content from filtrate, reaches alcohol-free taste,
Then the extractum obtaining proportion 1.0-1.5 it is concentrated in vacuo to；
Step 6: water precipitating: the hot water of 90-98 DEG C of 18-22 times of weight of concentrated extract is sufficiently stirred for and uses ultrasound wave
Shake 2.5-3.5 hour, high speed centrifugation at a temperature of 80-90 DEG C, collect supernatant；
Step 7: centrifugal: supernatant refrigerated overnight step 6 obtained, carries out high speed centrifugation for second day, collects precipitation,
Vacuum drying obtains yellow greenish powder primary extract；Cold preservation is conducive to ampelopsin and dihydromyricetin crystalline deposit, overnight ensures target
Thing sufficient crystallising separates out；
Step 8: isolated and purified；The primary extract that obtains step 7 is isolated and purified obtain more than 98% high-purity Fructus Myricae rubrae
Element and dihydromyricetin sterling.
Preferably, in step 8, described purification procedures is as follows:
1) will obtain yellow greenish powder primary extract in step 7, the ethyl acetate of point 9 times of weight of 3 use adds yellowish green toner
In the primary extract of end, uniform stirring with ultrasonic vibration 2.5-3.5 hour, ethyl acetate layer becomes yellow green, merges and obtains for three times
Ethyl acetate supernatant and the white depositions that obtains for three times；
2) to step 1) white depositions 8-12 times 5% sodium bicarbonate solution that obtains dissolves, high speed centrifugation, takes supernatant
Liquid, is acidified supernatant, refrigerated overnight, obtains white crystals precipitation, dries the dihydromyricetin obtaining purity at least 90%；Again
It is sufficiently stirred for dissolving with more than 90 DEG C hot water of 5 times of weight by white crystals precipitation, high speed centrifugation, precipitation separation, collects supernatant
Liquid, by supernatant refrigerated overnight, high speed centrifugation, collect and obtain white, needle-shaped crystals, with pure water 3 times, obtain white needles
Crystallization, dries the white needles dihydromyricetin cellulose crystal obtaining purity more than 98%；
3) from step 1) supernatant that obtains reclaims ethyl acetate, vacuum drying obtains yellow powder, by yellow powder
5% sodium bicarbonate solution is used to dissolve, high speed centrifugation, remove precipitation and collect supernatant, supernatant is acidified, chilled overnight, point
More than 98% yellow green needle-like ampelopsin crystallization can be obtained from precipitation vacuum drying.
Preferably, in step 8, described purification procedures is as follows:
1) step 7 will obtain yellow greenish powder primary extract, add 5% boric acid solution of 10 times of primary extract weight, fully
Stirring also ultrasonic vibration 2.5-3.5 hour, then stands overnight precipitation；High speed centrifugation, removes precipitation, collects supernatant；
2) adding 5% aqua calcis in supernatant, be slowly stirred, constantly produce white flock precipitate, regulation is steady
Fixed to PH 7.5, more slowly stir half an hour more than, high speed centrifugation removes precipitation, collects supernatant；
3) by collect supernatant hydrochloric acid regulation pH value to 4.0, refrigerated overnight, high speed centrifugation precipitate, collection obtains
The only precipitation of the mixture of ampelopsin and dihydromyricetin；
4) in step 3) precipitation that obtains point adds for 3 times the ethyl acetate of 9 times of weight, it is sufficiently stirred for and ultrasonic wave concussion
More than 3 hours, stand overnight；High speed centrifugation, collects precipitation and supernatant respectively；
5) by step 4) the precipitation vacuum drying that obtains, obtain the white needles dihydromyricetin cellulose crystal of purity 98%；
6) from step 4) vacuum reclaims ethyl acetate the supernatant that obtains, and obtain body of paste, add 10 times more than 95 DEG C
Hot water, fully dissolves, and filters, cooling, overnight, high speed centrifugation, collect the yellow needles ampelopsin crystalline substance obtaining purity more than 98%
Body.
Preferably, in step one, described sieve is 40 mesh.
Preferably, in step 2 and step 3, described extraction temperature is 40 DEG C.
Preferably, in step 5, described proportion is 1.15-1.20.
Preferably, in step 6, described water precipitating temperature is 95 DEG C.
Preferably, in step 3) in, vacuum is-0.08--0.09MPa.
Preferably, described method comprises the steps:
Step one: preparation: weigh Ampelopsis grossedentata, selected, pulverize, cross 40 mesh sieves；
Step 2: extraction: add the ethanol of the weight concentration 80% of Ampelopsis grossedentata weight 20 times, adjusts with 5% sodium bicarbonate solution
Joint pH value is 7.5, soaks 8 hours, then backflow extraction 4 hours at a temperature of 40 DEG C；
Step 3: secondary extracts: add the ethanol of the weight concentration 80% of Ampelopsis grossedentata weight 10 times, refluxes at a temperature of 40 DEG C
Extract 4 hours；
Step 4: filter: merge twice extraction liquid, be filtrated to get filtrate；
Step 5: recovered alcohol: recovered alcohol is less than 15% to alcohol concentration from filtrate, is then concentrated in vacuo to
Extractum to proportion 1.2；
Step 6: water precipitating: concentrated extract is sufficiently stirred for the hot water of 95 DEG C of 20 times of weight and uses ultrasonic wave concussion 3 little
Time, high speed centrifugation at a temperature of 85 DEG C, collect supernatant；
Step 7: centrifugal: supernatant refrigerated overnight step 6 obtained, carries out high speed centrifugation for second day, collects precipitation,
Vacuum drying obtains yellow greenish powder primary extract；
Step 8: isolated and purified；The primary extract that obtains step 7 is isolated and purified obtain more than 98% high-purity Fructus Myricae rubrae
Element and dihydromyricetin sterling.
Present invention also offers above-mentioned from Ampelopsis grossedentata obtained by the method for separating-purifying ampelopsin and dihydromyricetin
Ampelopsin and dihydromyricetin.
(3) beneficial effect
The used equipment of the inventive method is simple, technique simple possible, it is simple to industrialized production；Solvent for use is nontoxic without residual
Staying, do not produce environmental pollution, the organic solvent usage amount of final wash crystallization is little；The present invention utilizes Ampelopsis grossedentata extraction separation and purification
Utilization rate high, the gained response rate high, all more than 90%, gained ampelopsin, dihydromyricetin purity are the highest, can directly use
In pharmaceutical industries.
Detailed description of the invention
Below in conjunction with embodiment, embodiments of the present invention are described in further detail.Following example are used for illustrating this
Invention, but can not be used for limiting the scope of the present invention.
The solvent that used below, HCL, sodium bicarbonate, ethyl acetate are food grade or analytical pure, must not make recruitment
Industry level.
Embodiment 1
1) 1000 kilograms, the selected remove impurity of Ampelopsis grossedentata, Ampelopsis grossedentata is pulverized, and crosses 40 mesh sieves, but the most meticulous, polishing 1000 kilograms of being weighed
Total amount；
2) extraction for the first time: 20 ton of 80% alcoholic solution sodium bicarbonate regulates PH to 7.5, adds 10 tons of extraction pot and soaks
8 hours then backflow extraction 4 hours at a temperature of 40 DEG C；
3) second time extraction: backflow extraction 4 hours at a temperature of 10 ton of 80% alcoholic solution 40 DEG C；
4) filter with second time lixiviating solution plate and frame type filter-press for the first time, obtain the filtrate totally 28.6 tons of merging；
5) recovered alcohol and concentration: recovered alcohol after filtrate merging, to alcohol concentration less than 15%, is then concentrated in vacuo to
Obtaining the extractum of proportion 1.2, extractum weight is 1340 kilograms；
6) extractum is sufficiently stirred for the hot water of 27 tons more than 95 DEG C and uses ultrasonic wave concussion 3 hours, high under (85 DEG C) while hot
Speed is centrifugal collects supernatant 25.9 tons；
7) supernatant enters freezer, refrigerated overnight, and within second day, high speed centrifugation collects precipitation, and vacuum drying obtains yellow greenish powder
Primary extract 632 kilograms；
8) ampelopsin and the dihydromyricetin to 632 kilograms of yellow green primary extract isolated and purified extraction high-purities more than 98%
Sterling；
8.1) the first isolation and purification method；
8.1.1) step 7) gained yellow green primary extract 312 kilograms, divides 3 times (often by 2700 kilograms of analytical pure ethyl acetate
Secondary 900 kilograms) amount adds in yellow greenish powder, continuous low rate mixing with ultrasonic vibration 3 hours, and ethyl acetate layer becomes yellowish green
Color, last 3 times of ethyl acetate layers are basically unchanged color；
8.1.2) respectively obtain white depositions (1) 210 kilogram and three ethyl acetate supernatant combined wts 2800 are public
Jin；
8.1.3) precipitation (1) is easily dissolved with 1000 kilogram of 5% sodium bicarbonate, high speed centrifugation, supernatant acidifying cold preservation
Overnight white crystals precipitation dries to obtain the dihydromyricetin of at least 129 kilogram of 90% purity, white, needle-shaped crystals is with 600 kilograms
More than 95 DEG C hot water are sufficiently stirred for dissolving, and high speed centrifugation precipitation separation collects supernatant, and supernatant refrigerated overnight high speed centrifugation is received
Collection white, needle-shaped crystals, after pure water 3 times yellow needles dries the white needles dihydromyricetin that can obtain more than 98%
Cellulose crystal 123.3 kilograms.
8.1.4) ethyl acetate supernatant 2800 kilograms；Supernatant reclaims ethyl acetate, and vacuum drying obtains yellow powder 9.1
Kilogram, yellow powder uses 100 kilogram of 5% sodium bicarbonate easily dissolve high speed centrifugation and removes precipitation collection supernatant 106.4
Kilogram, supernatant acidifying chilled overnight precipitation separation vacuum drying can obtain more than 98% yellow green needle-like ampelopsin crystallization 8.7
Kilogram；
8.2) the second isolation and purification method；
8.2.1) step 7) gained yellow greenish powder primary extract 316 kilograms, 5% boric acid adding 3000 kilogram weights is molten
Liquid, is sufficiently stirred for also ultrasonic vibration 3 hours, then stands overnight precipitation；
8.2.2) overnight rear high speed centrifugation removing precipitates to obtain supernatant 3260 kilograms；
8.2.3) supernatant adds 5% aqua calcis, is slowly stirred, and constantly produces white flock precipitate, and regulation is steady
Fixed to PH7.5, slowly stir half an hour more than, high speed centrifugation removes precipitation, collects supernatant 2980 kilograms；
8.2.4) the supernatant hydrochloric acid regulation PH to 4.0 collected, refrigerated overnight；
8.2.5) overnight rear high speed centrifugation precipitates, and collects the precipitation obtaining the only mixture of ampelopsin and dihydromyricetin
(2) 322 kilograms；
8.2.6) precipitation (2) carries out separation ampelopsin and dihydromyricetin by the analytical pure ethyl acetate of 2700 kilograms, and 900
Kilogram of one time analytical pure ethyl acetate is divided 3 times and is sufficiently stirred for and ultrasonic wave concussion more than 3 hours, and high speed centrifugation precipitation collects supernatant
Liquid, precipitation is again ultrasonic by 900 kilograms of ethyl acetate and stirs high speed centrifugation precipitation separation and supernatant again, for the third time with 900
Precipitation is stripped and centrifugation by kilogram ethyl acetate；
8.2.7) dihydromyricetin 121.7 obtaining white needle-like crystals after (3) 119 kg white crystallizations are dried must be precipitated
Kilogram, three supernatant 2690 kilograms of merging；
8.2.8) vacuum reclaims ethyl acetate and obtains body of paste 24.5 kilograms, and body of paste adds 250 kilograms of more than 95 DEG C hot water
Fully dissolution filter, cold goes overnight that high speed centrifugation collects yellow needles body and vacuum drying obtains 8.5 kilograms of ampelopsin purity
More than 98%.
Embodiment 2
1) 1000 kilograms, the selected remove impurity of Ampelopsis grossedentata, Ampelopsis grossedentata is pulverized, and crosses 40 mesh sieves, but the most meticulous, polishing 1000 kilograms of being weighed
Total amount；
2) extraction for the first time: 20 ton of 80% alcoholic solution sodium bicarbonate regulates PH to 7.5, adds 10 tons of extraction pot and soaks
8 hours then backflow extraction 4 hours at a temperature of 40 DEG C；
3) second time extraction: backflow extraction 4 hours at a temperature of 10 ton of 80% alcoholic solution 40 DEG C；
4) filter with second time lixiviating solution plate and frame type filter-press for the first time, obtain the filtrate totally 28.4 tons of merging；
5) recovered alcohol and concentration: recovered alcohol after filtrate merging, to alcohol concentration less than 15%, is then concentrated in vacuo to
Obtaining the extractum of proportion 1.2, extractum weight is 1290 kilograms；
6) extractum is sufficiently stirred for the hot water of 26 tons more than 95 DEG C and uses ultrasonic wave concussion 3 hours, high under (85 DEG C) while hot
Speed is centrifugal collects supernatant 25.1 tons；
7) supernatant enters freezer, refrigerated overnight, and within second day, high speed centrifugation collects precipitation, and vacuum drying obtains yellow greenish powder
Primary extract 628 kilograms；
8) ampelopsin and the dihydromyricetin to 628 kilograms of yellow green primary extract isolated and purified extraction high-purities more than 98%
Sterling；
8.1) the first isolation and purification method；
8.1.1) step 7) gained yellow green primary extract 314 kilograms, divides 3 times (often by 2700 kilograms of analytical pure ethyl acetate
Secondary 900 kilograms) amount adds in yellow greenish powder, continuous low rate mixing with ultrasonic vibration 3 hours, and ethyl acetate layer becomes yellowish green
Color, last 3 times of ethyl acetate layers are basically unchanged color；
8.1.2) respectively obtain white depositions (1) 201 kilogram and three ethyl acetate supernatant combined wts 2880 are public
Jin；
8.1.3) precipitation (1) is easily dissolved with 1000 kilogram of 5% sodium bicarbonate, high speed centrifugation, supernatant acidifying cold preservation
Overnight white crystals precipitation dries to obtain the dihydromyricetin of at least 132.7 kilogram of 90% purity, white, needle-shaped crystals is by 650 public affairs
More than 95 DEG C hot water of jin are sufficiently stirred for dissolving, and high speed centrifugation precipitation separation collects supernatant, supernatant refrigerated overnight high speed centrifugation
After collecting white, needle-shaped crystals, pure water 3 times yellow needles dries the white needles dihydro poplar that can obtain more than 98%
120.2 kilograms of syphilis crystal.
8.1.4) ethyl acetate supernatant 2880 kilograms；Supernatant reclaims ethyl acetate, and vacuum drying obtains yellow powder 9.2
Kilogram, yellow powder uses 100 kilogram of 5% sodium bicarbonate easily dissolve high speed centrifugation and removes precipitation collection supernatant 110.1
Kilogram, supernatant acidifying chilled overnight precipitation separation vacuum drying can obtain more than 98% yellow green needle-like ampelopsin crystallization 8.6
Kilogram；
8.2) the second isolation and purification method；
8.2.1) step 7) gained yellow greenish powder primary extract 314 kilograms, 5% boric acid adding 3000 kilogram weights is molten
Liquid, is sufficiently stirred for also ultrasonic vibration 3 hours, then stands overnight precipitation；
8.2.2) overnight rear high speed centrifugation removing precipitates to obtain supernatant 3310 kilograms；
8.2.3) supernatant adds 5% aqua calcis, is slowly stirred, and constantly produces white flock precipitate, and regulation is steady
Fixed to PH7.5, slowly stir half an hour more than, high speed centrifugation removes precipitation, collects supernatant 3130 kilograms；
8.2.4) the supernatant hydrochloric acid regulation PH to 4.0 collected, refrigerated overnight；
8.2.5) overnight rear high speed centrifugation precipitates, and collects the precipitation obtaining the only mixture of ampelopsin and dihydromyricetin
(2) 317 kilograms；
8.2.6) precipitation (2) carries out separation ampelopsin and dihydromyricetin by the analytical pure ethyl acetate of 2700 kilograms, and 900
Kilogram of one time analytical pure ethyl acetate is divided 3 times and is sufficiently stirred for and ultrasonic wave concussion more than 3 hours, and high speed centrifugation precipitation collects supernatant
Liquid, precipitation is again ultrasonic by 900 kilograms of ethyl acetate and stirs high speed centrifugation precipitation separation and supernatant again, for the third time with 900
Precipitation is stripped and centrifugation by kilogram ethyl acetate；
8.2.7) dihydromyricetin obtaining white needle-like crystals after (3) 129.6 kg white crystallizations are dried must be precipitated
119.8 kilograms, three supernatant 2700 kilograms of merging；
8.2.8) vacuum reclaims ethyl acetate and obtains body of paste 25.5 kilograms, and body of paste adds 250 kilograms of more than 95 degree hot water
Fully dissolution filter, cold goes overnight that high speed centrifugation collects yellow needles body and vacuum drying obtains 8.3 kilograms of ampelopsin purity
More than 98%.
Embodiment 3
1) 1000 kilograms, the selected remove impurity of Ampelopsis grossedentata, Ampelopsis grossedentata is pulverized, and crosses 40 mesh sieves, but the most meticulous, polishing 1000 kilograms of being weighed
Total amount；
2) extraction for the first time: 20 ton of 80% alcoholic solution sodium bicarbonate regulates PH to 7.5, adds 10 tons of extraction pot and soaks
8 hours then backflow extraction 4 hours at a temperature of 40 DEG C；
3) second time extraction: backflow extraction 4 hours at a temperature of 10 ton of 80% alcoholic solution 40 DEG C；
4) filter with second time lixiviating solution plate and frame type filter-press for the first time, obtain the filtrate totally 29.1 tons of merging；
5) recovered alcohol and concentration: recovered alcohol after filtrate merging, to alcohol concentration less than 15%, is then concentrated in vacuo to
Obtaining the extractum of proportion 1.2, extractum weight is 1295 kilograms；
6) extractum is sufficiently stirred for the hot water of 27 tons more than 95 DEG C and uses ultrasonic wave concussion 3 hours, high under (85 DEG C) while hot
Speed is centrifugal collects supernatant 25.7 tons；
7) supernatant enters freezer, refrigerated overnight, and within second day, high speed centrifugation collects precipitation, and vacuum drying obtains yellow greenish powder
Primary extract 638 kilograms；
8) ampelopsin and the dihydromyricetin to 638 kilograms of yellow green primary extract isolated and purified extraction high-purities more than 98%
Sterling；
8.1) the first isolation and purification method；
8.1.1) step 7) gained yellow green primary extract 319 kilograms, divides 3 times (often by 2700 kilograms of analytical pure ethyl acetate
Secondary 900 kilograms) amount adds in yellow greenish powder, continuous low rate mixing with ultrasonic vibration 3 hours, and ethyl acetate layer becomes yellowish green
Color, last 3 times of ethyl acetate layers are basically unchanged color；
8.1.2) respectively obtain white depositions (1) 218 kilogram and three ethyl acetate supernatant combined wts 2770 are public
Jin；
8.1.3) precipitation (1) is easily dissolved with 1000 kilogram of 5% sodium bicarbonate, high speed centrifugation, supernatant acidifying cold preservation
Overnight white crystals precipitation dries to obtain the dihydromyricetin of at least 128.6 kilogram of 90% purity, white, needle-shaped crystals is by 600 public affairs
More than 95 DEG C hot water of jin are sufficiently stirred for dissolving, and high speed centrifugation precipitation separation collects supernatant, supernatant refrigerated overnight high speed centrifugation
After collecting white, needle-shaped crystals, pure water 3 times yellow needles dries the white needles dihydro poplar that can obtain more than 98%
123.9 kilograms of syphilis crystal.
8.1.4) ethyl acetate supernatant 2770 kilograms；Supernatant reclaims ethyl acetate, and vacuum drying obtains yellow powder 8.8
Kilogram, yellow powder uses 100 kilogram of 5% sodium bicarbonate easily dissolve high speed centrifugation and removes precipitation collection supernatant 112.3
Kilogram, supernatant acidifying chilled overnight precipitation separation vacuum drying can obtain more than 98% yellow green needle-like ampelopsin crystallization 8.4
Kilogram；
8.2) the second isolation and purification method；
8.2.1) step 7) gained yellow greenish powder primary extract 319 kilograms, 5% boric acid adding 3000 kilogram weights is molten
Liquid, is sufficiently stirred for also ultrasonic vibration 3 hours, then stands overnight precipitation；
8.2.2) overnight rear high speed centrifugation removing precipitates to obtain supernatant 3350 kilograms；
8.2.3) supernatant adds 5% aqua calcis, is slowly stirred, and constantly produces white flock precipitate, and regulation is steady
Fixed to PH7.5, slowly stir half an hour more than, high speed centrifugation removes precipitation, collects supernatant 3030 kilograms；
8.2.4) the supernatant hydrochloric acid regulation PH to 4.0 collected, refrigerated overnight；
8.2.5) overnight rear high speed centrifugation precipitates, and collects the precipitation obtaining the only mixture of ampelopsin and dihydromyricetin
(2) 317 kilograms；
8.2.6) precipitation (2) carries out separation ampelopsin and dihydromyricetin by the analytical pure ethyl acetate of 2700 kilograms, and 900
Kilogram of one time analytical pure ethyl acetate is divided 3 times and is sufficiently stirred for and ultrasonic wave concussion more than 3 hours, and high speed centrifugation precipitation collects supernatant
Liquid, precipitation is again ultrasonic by 900 kilograms of ethyl acetate and stirs high speed centrifugation precipitation separation and supernatant again, for the third time with 900
Precipitation is stripped and centrifugation by kilogram ethyl acetate；
8.2.7) dihydromyricetin obtaining white needle-like crystals after (3) 133.3 kg white crystallizations are dried must be precipitated
124.1 kilograms, three supernatant 2720 kilograms of merging；
8.2.8) vacuum reclaims ethyl acetate and obtains body of paste 23.7 kilograms, and body of paste adds 250 kilograms of more than 95 degree hot water
Fully dissolution filter, cold goes overnight that high speed centrifugation collects yellow needles body and vacuum drying obtains 8.4 kilograms of ampelopsin purity
More than 98%.
Contrast experiment
Xiangxi Zhangjiajie Ampelopsis grossedentata is analyzed general flavone content through us and is reached 36.5%, and wherein dihydromyricetin contains
Amount is 27.8%, and ampelopsin content is 2.3%, and the commercial run that be there is presently no a kind of maturation by lookup data is extracted simultaneously
Ampelopsin and dihydromyricetin, in industrial Ampelopsis grossedentata, ampelopsin content is low, typically extracts ampelopsin from Fructus Myricae rubrae, in document the most not
Having industrialization high-purity ampelopsin and the extraction process of dihydromyricetin, the present invention has pioneering and initiative.
Experimental example 1
The content assaying method (Deng Danwen, University Of Nanchang's Food Science and technology National Key Laboratory) of ampelopsin
1, instrument
Aglient1260 high performance liquid chromatograph (quaternary pump, UV-detector), Simon Aldrich-C 18 solid phase extracts
Taking pillar, JT2003 precise electronic balance (the Hebei road limited branch company of instrument highway instrument), the swing high speed of DFY-500 is omnipotent
Pulverizer (Lin great Machinery Co., Ltd. of Wenling city), Rotary Evaporators RE-52 (Shanghai Yarong Biochemical Instrument Plant), DHG-9053
Type electric heating constant-temperature blowing drying box (three, Shanghai scientific instrument company limited), DKS-24 type electric-heated thermostatic water bath (Jiaxing City
Zhong Xin Medical Instruments company limited).
2, chromatographic condition
HPLC testing conditions chromatographic column: Thermo-C 18 liquid-phase chromatographic column (4.6mm × 250mm, 5 μm)；Flowing phase:
0.005mol/L potassium dihydrogen phosphate-methanol (volume ratio 30: 70)；Flow velocity is 0.5mL/min, sample introduction 10 μ L, column temperature 25 DEG C,
Detection wavelength 360nm.
3 Specification Curve of Increasings
Accurately weigh ampelopsin mark product 10mg and put in 100mL volumetric flask, dissolve and constant volume with methanol, molten as reference substance
Liquid.With methanol dilution constant volume, it is made into 10, the ampelopsin mark product solution of 20,30,40,50 μ g/mL series concentration, 10 μ L sample introductions,
Entering HPLC under 1.3.5.2 chromatographic condition to analyze, with peak area as vertical coordinate (Y), with concentration as abscissa, (X) draws standard song
Line.Gained regression equation is Y=54.155X-161.47, R=0.999 6, shows that sample closes in 10～50 μ g/mL internal linear
System is good.
Experimental example 2
Dihydromyricetin content assaying method (Wang Jiasheng, Chinese patent medicine in January, 2012 Guizhou University's Life Science College)
1, experimental apparatus
Test apparatus and equipment mainly have KQ-500DE type ultrasonic cleaner (Kunshan Ultrasonic Instruments Co., Ltd.), point
Analysis balance (100,000/, prunus mume (sieb.) sieb.et zucc. Teller-torr benefit Instrument Ltd.), (river, Shanghai believes that science and technology has to multifunctional crusher LX-02
Limit company)；Agilent1200LC (Anjelen Sci. & Tech. Inc of the U.S.)；Dikma chromatographic column (250mm × 4.60mm, 5 μm,
Di Ma company of China).
2, chromatographic condition
Chromatographic condition sample size 10 μ L；Ultraviolet detection wavelength 292nm；Column temperature 40 DEG C；Flowing be mutually 0.1% phosphoric acid-methanol-
Acetonihile gradient elution (0～10min, 74.4: 8: 17.6；10～15min, 36: 20: 44；15.1～18min, 74.4: 8:
17.6)；Volume flow 1mL/min.
3, the preparation of reference substance solution
Precision weighs dihydromyricetin 100.00mg, is placed in 100mL brown measuring bottle, dissolves with methanol and is diluted to carve
Degree, shakes up, obtains the dihydromyricetin reference substance solution that mass concentration is 1.00mg/mL, measure 1.0 respectively, 2.0,4.0,
5.0,8.0,10.0mL, methanol constant volume, to 10mL, obtains control series product solution.
4, linear relationship
Reference substance solution sample introduction 10 μ L, measures peak area, each mass concentration sample introduction 3 times, averages.With sample introduction quality
Concentration be abscissa (X), peak area be vertical coordinate (Y), draw standard curve, the regression equation obtaining dihydromyricetin is Y=2
832.7X-101.24 (r2=0.996 0), the range of linearity is 0.5～10.0 μ g, shows dihydromyricetin line in this range
Sexual relationship is good.
5, sample determination
Need testing solution sample introduction 10 μ l, it is thus achieved that sample chromatogram figure, by the amount of regression equation calculation dihydromyricetin.
Table: testing result
| Sample | Ampelopsin | Dihydromyricetin |
| Embodiment 1 | Method 1,98.3% method 2,98.5% | Method 1,98.7%, method 2,99.1% |
| Embodiment 2 | Method 1,98.0% method 2,98.1% | Method 1,99.2%, method 2,98.9% |
| Embodiment 3 | Method 1,98.4% method 2,98.5% | Method 1,99.0%, method 2,99.1% |
Embodiment of above is merely to illustrate the present invention, rather than limitation of the present invention.Although with reference to embodiment to this
Bright be described in detail, it will be understood by those within the art that, technical scheme is carried out various combination,
Amendment or equivalent, without departure from the spirit and scope of technical solution of the present invention, all should contain the right in the present invention and want
Ask in the middle of scope.
Claims (10)
1. separating-purifying ampelopsin and the method for dihydromyricetin from an Ampelopsis grossedentata, it is characterised in that the method includes as follows
Step:
Step one: preparation: weigh Ampelopsis grossedentata, selected, pulverize, cross 35-45 mesh sieve；
Step 2: extraction: add the ethanol of the 75-85% of Ampelopsis grossedentata weight 18-22 times, with 5% sodium bicarbonate solution regulation pH value
For 7.0-8.0, soak 6-10 hour, then backflow extraction 3-5 hour at a temperature of 35-45 DEG C；
Step 3: secondary extracts: add the ethanol of the 75-85% of Ampelopsis grossedentata weight 8-12 times, backflow extraction at a temperature of 35-45 DEG C
3-5 hour；
Step 4: filter: merge twice extraction liquid, be filtrated to get filtrate；
Step 5: recovered alcohol: recovered alcohol to alcohol concentration is less than 15% from filtrate, is then concentrated in vacuo to and is compared
The extractum of weight 1.0-1.5；
Step 6: water precipitating: the hot water of 90-98 DEG C of 18-22 times of weight of concentrated extract is sufficiently stirred for and uses ultrasonic wave concussion
2.5-3.5 hour, high speed centrifugation at a temperature of 80-90 DEG C, collect supernatant；
Step 7: centrifugal: supernatant refrigerated overnight step 6 obtained, carries out high speed centrifugation for second day, collects precipitation, vacuum
Dry to obtain yellow greenish powder primary extract；
Step 8: isolated and purified；The primary extract that obtains step 7 is isolated and purified obtain more than 98% high-purity ampelopsin and
Dihydromyricetin sterling.
Separating-purifying ampelopsin and the method for dihydromyricetin from Ampelopsis grossedentata the most according to claim 1, it is characterised in that
In step 8, described purification procedures is as follows:
1) will obtain yellow greenish powder primary extract in step 7, the ethyl acetate of point 9 times of weight of 3 use adds at the beginning of yellow greenish powder
In extract, uniform stirring with ultrasonic vibration 2.5-3.5 hour, ethyl acetate layer becomes yellow green, merges the second obtained for three times
Acetoacetic ester supernatant and the white depositions obtained for three times；
2) to step 1) white depositions 8-12 times 5% sodium bicarbonate solution that obtains dissolves, high speed centrifugation, takes supernatant,
Supernatant is acidified, refrigerated overnight, obtains white crystals precipitation, dry the dihydromyricetin obtaining purity at least 90%；Again will
White crystals precipitation is sufficiently stirred for dissolving with more than 90 DEG C hot water of 5 times of weight, high speed centrifugation, and precipitation separation collects supernatant,
By supernatant refrigerated overnight, high speed centrifugation, collect and obtain white, needle-shaped crystals, with pure water 3 times, obtain white needles knot
Crystalline substance, dries the white needles dihydromyricetin cellulose crystal obtaining purity more than 98%；
3) from step 1) supernatant that obtains reclaims ethyl acetate, vacuum drying obtains yellow powder, is used by yellow powder
5% sodium bicarbonate solution dissolves, high speed centrifugation, removes precipitation and collects supernatant, is acidified by supernatant, chilled overnight, and it is heavy to separate
Form sediment and be vacuum dried and can obtain more than 98% yellow green needle-like ampelopsin crystallization.
Separating-purifying ampelopsin and the method for dihydromyricetin from Ampelopsis grossedentata the most according to claim 1, it is characterised in that
In step 8, described purification procedures is as follows:
1) step 7 will obtain yellow greenish powder primary extract, add 5% boric acid solution of 10 times of primary extract weight, be sufficiently stirred for
And ultrasonic vibration 2.5-3.5 hour, then stand overnight precipitation；High speed centrifugation, removes precipitation, collects supernatant；
2) adding 5% aqua calcis in supernatant, be slowly stirred, constantly produce white flock precipitate, regulation is stable extremely
Till PH 7.5, more slowly stirring half an hour more than, high speed centrifugation removes precipitation, collects supernatant；
3) by collect supernatant hydrochloric acid regulation pH value to 4.0, refrigerated overnight, high speed centrifugation precipitate, collect only obtain
The precipitation of the mixture of ampelopsin and dihydromyricetin；
4) in step 3) precipitation that obtains point adds for 3 times the ethyl acetate of 9 times of weight, it is sufficiently stirred for and ultrasonic wave concussion 3 is little
More than time, stand overnight；High speed centrifugation, collects precipitation and supernatant respectively；
5) by step 4) the precipitation vacuum drying that obtains, obtain the white needles dihydromyricetin cellulose crystal of purity 98%；
6) from step 4) vacuum reclaims ethyl acetate the supernatant that obtains, and obtain body of paste, add 10 times of more than 95 DEG C hot water,
Fully dissolve, filter, cooling, overnight, and high speed centrifugation, collect the yellow needles ampelopsin crystal obtaining purity more than 98%.
Separating-purifying ampelopsin and the method for dihydromyricetin from Ampelopsis grossedentata the most according to claim 1, it is characterised in that
In step one, described sieve is 40 mesh.
Separating-purifying ampelopsin and the method for dihydromyricetin from Ampelopsis grossedentata the most according to claim 1, it is characterised in that
In step 2 and step 3, described extraction temperature is 40 DEG C.
Separating-purifying ampelopsin and the method for dihydromyricetin from Ampelopsis grossedentata the most according to claim 1, it is characterised in that
In step 5, described proportion is 1.15-1.20.
Separating-purifying ampelopsin and the method for dihydromyricetin from Ampelopsis grossedentata the most according to claim 1, it is characterised in that
In step 6, described water precipitating temperature is 95 DEG C.
Separating-purifying ampelopsin and the method for dihydromyricetin from Ampelopsis grossedentata the most according to claim 2, it is characterised in that
In step 3) in, vacuum is-0.08--0.09MPa.
9. according to the separating-purifying ampelopsin from Ampelopsis grossedentata described in any one of claim Isosorbide-5-Nitrae-7 and the method for dihydromyricetin,
It is characterized in that, the method comprises the steps:
Step one: preparation: weigh Ampelopsis grossedentata, selected, pulverize, cross 40 mesh sieves；
Step 2: extraction: the ethanol of the 80% of addition Ampelopsis grossedentata weight 20 times, the sodium bicarbonate solution with 5% regulates pH value and is
7.5, soak 8 hours, then backflow extraction 4 hours at a temperature of 40 DEG C；
Step 3: secondary extracts: the ethanol of the 80% of addition Ampelopsis grossedentata weight 10 times, backflow extraction 4 hours at a temperature of 40 DEG C；
Step 4: filter: merge twice extraction liquid, be filtrated to get filtrate；
Step 5: recovered alcohol: recovered alcohol to alcohol content is less than 15% from filtrate, is then concentrated in vacuo to and is compared
Weigh the extractum of 1.2；
Step 6: water precipitating: concentrated extract is sufficiently stirred for the hot water of 95 DEG C of 20 times of weight and uses ultrasonic wave concussion 3 hours,
High speed centrifugation at a temperature of 85 DEG C, collects supernatant；
Step 7: centrifugal: supernatant refrigerated overnight step 6 obtained, carries out high speed centrifugation for second day, collects precipitation, vacuum
Dry to obtain yellow greenish powder primary extract；
Step 8: isolated and purified；The primary extract that obtains step 7 is isolated and purified obtain more than 98% high-purity ampelopsin and
Dihydromyricetin sterling.
10. described in any one of claim 1-9 from Ampelopsis grossedentata obtained by the method for separating-purifying ampelopsin and dihydromyricetin
Ampelopsin and dihydromyricetin.
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| CN113501802A (en) * | 2021-07-14 | 2021-10-15 | 西安金泰生物技术有限公司 | Preparation method of dihydromyricetin |
| CN114960277A (en) * | 2021-07-04 | 2022-08-30 | 惠州市随和科技有限公司 | Ampelopsis grossedentata extract, preparation method thereof and application thereof in water-proof and oil-proof agents |
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN106518829A (en) * | 2016-11-10 | 2017-03-22 | 张清峰 | Method for separating and purifying dihydromyricetin from ampelopsis grossedentata leaves |
| CN108042661A (en) * | 2017-12-16 | 2018-05-18 | 江西天元药业有限公司 | The purposes of Ramulus et Folium Mussaendae Pubescentis extract and preparation medical and health product rich in dihydromyricetin |
| CN114960277A (en) * | 2021-07-04 | 2022-08-30 | 惠州市随和科技有限公司 | Ampelopsis grossedentata extract, preparation method thereof and application thereof in water-proof and oil-proof agents |
| CN113501802A (en) * | 2021-07-14 | 2021-10-15 | 西安金泰生物技术有限公司 | Preparation method of dihydromyricetin |
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Denomination of invention: Method for separating and purifying myricetin and dihydromyricetin from ampelopsis grossedentata Effective date of registration: 20191125 Granted publication date: 20180928 Pledgee: Bank of Changsha Co., Ltd. Kaifu sub branch Pledgor: HUNAN NUTRAMAX INC. Registration number: Y2019430000047 |
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Denomination of invention: A method for separation and purification of myricetin and Dihydromyricetin from Ampelopsis grossedentata Effective date of registration: 20201218 Granted publication date: 20180928 Pledgee: Bank of Changsha Co.,Ltd. Kaifu sub branch Pledgor: HUNAN NUTRAMAX Inc. Registration number: Y2020980009579 |
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