CN106668088A - Preparation method of hyperforin perforatum extract - Google Patents
Preparation method of hyperforin perforatum extract Download PDFInfo
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- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K36/00—Medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicines
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- A61K2236/10—Preparation or pretreatment of starting material
- A61K2236/19—Preparation or pretreatment of starting material involving fermentation using yeast, bacteria or both; enzymatic treatment
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- A—HUMAN NECESSITIES
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- A61K2236/30—Extraction of the material
- A61K2236/33—Extraction of the material involving extraction with hydrophilic solvents, e.g. lower alcohols, esters or ketones
- A61K2236/333—Extraction of the material involving extraction with hydrophilic solvents, e.g. lower alcohols, esters or ketones using mixed solvents, e.g. 70% EtOH
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
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- A61K2236/00—Isolation or extraction methods of medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicine
- A61K2236/50—Methods involving additional extraction steps
- A61K2236/51—Concentration or drying of the extract, e.g. Lyophilisation, freeze-drying or spray-drying
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K2236/00—Isolation or extraction methods of medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicine
- A61K2236/50—Methods involving additional extraction steps
- A61K2236/55—Liquid-liquid separation; Phase separation
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Abstract
The invention relates to a preparation method of a hyperforin perforatum extract and the prepared hyperforin perforatum extract. The method is simple in step and high in yield, industrial mass production is very convenient, and the hyperforin perforatum extract which comprises three active ingredients of over 6.0wt% of hypericin, over 2.0wt% of hyperforin and over 75wt% of flavonoid constituent is obtained.
Description
Technical field
The present invention relates to the preparation method of Herba Hyperici perforati extract.
Background technology
Herba Hyperici perforati Hypericum perforatum L., are Guttiferae hypericum, also known as Herba Hyperici Monogyni,
It is distributed widely in the ground such as Europe, Asia, Australia, is one of medical herbs for more early recognizing for the mankind and utilizing.Mainly it is distributed in China
In Hebei, Shaanxi, Gansu, Shandong and other places, its dry aerial parts has been widely used in clinic as Chinese traditional herbs.According to
Record, Herba Hyperici perforati nature and flavor are pungent, tremble with fear, return liver warp, the effects such as with dispersing the stagnated live-QI to relieve the stagnation of QI, clearing away heat-damp and promoting diuresis, detumescence lactogenesis, for hepatic depression
Knot, feelings will is not smooth, and ambition is gloomy, arthralgia, acute mastitis, and breast is few；Also can herb fresh goods mash or dry product grinds deposited affected part, control wound
Hinder bleeding, pain furuncle and phyma poison and burn and scald etc..Germany is called Herba Hyperici perforati, makees that antidepressant is existing more than 100 years to be gone through with it
History；Also there are a commerial growing in the U.S., Canada, and by countries such as Germany, the U.S. have been embodied in pharmacopeia, are widely used in clinic.
With the development of Hydrolysis kinetics technology and going deep into for pharmacological research, Herba Hyperici perforati is in antiviral, antidepressant, antitumor
Etc. the effect of aspect increasingly paid close attention to by the mankind.Herba Hyperici perforati extract ingredient is extremely complex, is broadly divided into three classes：Naphthalene
Parallel dianthrone compound (naphthodianthrones), phloroglucinol derivatives compound (phiorogluCinol S) and flavone
Class compound (flavonoids).Wherein, naphthalene a pair of horses going side by side dianthrone compound for Herba Hyperici perforati characteristic component, including hypericin
Hypericin, pseudohypericin (pseudohypericin), former hypericin (ProtohyperiCin), former pseudohypericin
(protopseudohypericin) etc.；Phloroglucinol derivatives compound mainly includes hyperforine (hypefonn) and adds to pass through
Leaf hypericin (adhyperforin) etc., flavone compound includes Quercetin (quercetin), Quercitroside
(quercitrin), isoquercitrin (iso-quercitrin), rutin (rutin), luteolin (1uteolin), hyperin
(hyperin/hyperoside) etc..
Early stage research thinks that hypericin is the main component of Herba Hyperici perforati, therefore develops and various carry from Herba Hyperici perforati
The method for taking hypericin, such as CN10508224A disclose a kind of method that hypericin is extracted from Herba Hyperici perforati, including
Immersion, decocting in water, digest, deoil, the step of alkali carries, column chromatography and drying, finally give Determination of Hypericin from Extraction for 5-6% whole product
Thing.But with the further investigation to antidepressant and mechanism, it is believed that including hypericin, pass through leaf spun gold
Fructus Persicae element, flavones ingredient together constitute its antidepressant active component in interior Multiple components.Therefore it provides having abundant work
The Herba Hyperici perforati extract of property composition becomes current study hotspot, rather than is confined to single component.
The preparation method of existing Herba Hyperici perforati extract has the heavy crystallization process of alcohol extraction acid, macroreticular resin absorbing method, alcohol extraction extraction
Take chromatography, CTAB inverse micelle abstraction methods, supersound extraction macroreticular resin absorbing method etc..Leaf is passed through as CN1181855A discloses one kind
The method of Fructus Forsythiae extract, including ethanol extraction, adsorption column separation, solvent-extracted method, obtain containing more than 0.6% spun gold
The Herba Hyperici perforati extract of Fructus Persicae element and 50-70% flavone；CN1880328A is disclosed and is separated and carry with inverse micelle abstraction method, resin
Take the preparation method of Herba Hyperici perforati hyperin and hypericin.Extract in order to be able to obtain the Herba Hyperici perforati containing more effective ingredient
Thing, the present inventor continues to grind on the basis of achievement in research " a kind of method that hypericin is extracted from Herba Hyperici perforati " before
Study carefully, the present invention is obtained then.
The content of the invention
It is an object of the invention to provide a kind of preparation method of Herba Hyperici perforati extract and its obtained Herba Hyperici perforati are extracted
Thing.
The preparation method of the Herba Hyperici perforati extract of the present invention specifically includes following steps：
（1）Take Herba Hyperici perforati and be dried herb（Including flower and fruit）, crush, cross 40 mesh sieves；Add raw material weight 20-40%'s
Water, adjusts pH to 4.0-6.0, and adds enzyme preparation, closed stir process 1-3h of lucifuge, knot in the ratio of raw material weight 0.5-3%
60-80 DEG C is heated to before beam, continues 5-15min, be down to room temperature, separation discards liquid, the raw material after must processing；
（2）The raw material after processing is taken, is extracted in extraction pot by Extraction solvent of alkali liquor, raw material weight is pressed in extraction pot
0.1-1% adds stabilizer, obtains extracting solution A；
（3）The pH value of extracting solution A is adjusted to into acidity, centrifugation obtains supernatant C；
（4）Supernatant C is isolated and purified through simulated moving bed chromatography, eluent D is obtained；
（5）By eluent D concentrations, then it is freeze-dried obtain Herba Hyperici perforati extract, extract contains Determination of Hypericin from Extraction.
Wherein, step（1）In enzyme preparation can be xylanase, pectase or its mixture, preferred xylanase with
The mixture of pectase, particularly xylanase and pectase 3:1 mixed enzyme；Step（1）The addition of middle enzyme preparation is preferred
1-2 weight %, more preferably 1.5 weight %；The intensification for terminating before processing preferably reaches 80 DEG C, continues 5min.
Wherein, step（2）Alkali liquor be 1% sodium hydroxide solution, every kilogram of raw material uses 20-25L Extraction solvents, 40-
60 DEG C of extraction 2-3h；Stabilizer can be Butylated hydroxyanisole or dibenzylatiooluene.
Wherein, step（3）In, adjust pH to 3-4 using 10% hydrochloric acid solution；Centrifuge speed is 4000 turns/min, from
The heart time is 10min.
Wherein, step（4）In, the adsorbent of simulated moving bed chromatography filling is nonpolar macroporous adsorption resin, preferably
D101 non-polar resins or HPD100 non-polar resins, water wash zone is pure water, and strippant is 60% ethanol, and resin absorption regeneration is molten
Agent is 95% ethanol or 2-4% sodium hydrate aqueous solutions, and adsorption zone flow velocity 1-2BV/h, water wash zone flow velocity 1-3 BV/h, desorption zone is flowed
Fast 1-3 BV/h, renewing zone flow velocity 2-3 BV/h, switching time 600-800s, at 40-60 DEG C, Stress control exists temperature control
0.1-0.5Mpa。
Wherein, step（5）In, eluent D is concentrated into into solid quality percentage composition for 30-40%, then it is freeze-dried
Obtain Herba Hyperici perforati extract.
Present invention also offers have Herba Hyperici perforati extract obtained in said method, comprising more than 6.0 weights in the extract
The amount hypericin of %, the hyperforine more than 2.0 weight %, more than the flavones ingredient of 75 weight %.
The present inventor be based on before achievement in research, i.e., using moving-bed adsorption chromatographic technique, but not only with gold
Silk Fructus Persicae element is target component, and hyperforine, flavones ingredient etc. are classified as after consideration target, has extensively consulted existing text
Part, therefrom have selected possible means, and by a large amount of work, the means that have selected optimum Herba Hyperici perforati are combined, and
This method for being surprised to find that the present invention obtains splendid effect, essentially consists in：
（1）As well known to those skilled in the art using processing with enzyme preparation raw material, the specific processing with enzyme preparation of Jing can be removed well
Vegetable material has grease-like composition, the inventors discovered that, although the mesh of remove impurity can be realized using conventional enzymatic isolation method
, but because Herba Hyperici perforati ingredient is more unstable composition, the process of enzymolysis will also cause various active composition
Loss, the oxidation of such as flavones ingredient, conversion of hyperforine etc., are unfavorable for the goal of the invention of the present invention.Therefore, send out
A person of good sense has attempted the technological means such as addition stabilizer, antioxidant, lucifuge be closed, and effect not substantially, Jing after many kinds are attempted, is sent out
A person of good sense has found that enzymolysis is carried out under conditions of lucifuge is closed can be realized and add stabilizer and result as antioxidant, and
And the introducing of impurity is also reduced, more suitably；Subsequently, inventor is surprised to find that the enzymolysis under lucifuge airtight condition very much
End stage carries out the intensification of a short time, not only obtains impurity-eliminating effect well, after last whole method terminates,
The extract comprising more target components can also be obtained.This seems to be disagreed with the thermal instability of Herba Hyperici perforati composition, invention
People's conjecture may be between the composition of inside plants mutual conversion it is relevant, need further research and confirm.
（2）The present inventor further have adjusted the relevant parameter of whole method so that except the short time high temperature of enzymolysis step
Outward, remaining all step controls to be carried out in the environment of not higher than 60 DEG C, further reduces the loss of effective ingredient.This
A person of good sense once attempted the step of instantaneously being heated up in each stage on the basis of above-mentioned having been surprisingly found that, also, did not as a result make us
Satisfied, either extracting part or the intensification of separate section will all clearly result in the reduction of active constituent content.
（3）The each step of the method for the present invention is mutually assisted, the method for constituting an entirety, is finally obtained comprising being more than
The hypericin of 6.0 weight %, the hyperforine more than 2.0 weight %, more than 75 weight % flavones ingredient three major types
The Herba Hyperici perforati extract of active component, active component total content >=80%.It is non-and method of the present invention step is simple, high income
Often it is easy to industrialized great production.
Present inventor has carried out lot of experiments, experiment provided below to obtain technical scheme
As a result it is only used for showing the superiority of the preparation method relative to additive method of the present invention, raw material is chosen 100g Herba Hyperici perforatis and entered
Row test.
1st, extract active component contrast test
The inventive method：According to the method for embodiment 1.
Contrast method 1：Method of the present inventor in front inventive embodiment 1.
Contrast method 2：The method of CN1181855A embodiments 1.
Contrast method 3：The method of CN11198097A embodiments 2.
Contrast method 4：The method of CN105085224A embodiments 1.
Contrast method 5：The method of CN1392129A embodiments 1.
Experimental result see the table below, and numerical value represents that such composition accounts for extract obtained percentage by weight.
2nd, the extract obtained antidepressant activity test of the present invention
Test method：Using the experimental model of rat antidepressant effect, the antidepressant activity of Herba Hyperici perforati extract is verified.Test
The extract obtained by mentioned component contrast test with medicine, dosage is 50mg/kg, is configured with 0.3%CMCNa before use.System
Count and represented with x ± SD.As a result see the table below.
Can see, the inventive method is extract obtained can substantially to shorten the rats'swimming dead time.
Specific embodiment
The present invention is illustrated in more detail with reference to detailed description below.
Embodiment 1
Take Herba Hyperici perforati and be dried herb（Including flower and fruit）1kg, crushes, and crosses 40 mesh sieves；Raw material weight 0.4L water is added, is adjusted
Section pH adds xylanase and pectase 3 to 4.5 in the ratio of raw material weight 1.5%:1 mixed enzyme, the closed stirring of lucifuge
2h is processed, 80 DEG C are heated to before terminating, continue 5min, be down to room temperature, separation discards liquid, the raw material after must processing；Plus
1% sodium hydrate aqueous solution for entering 20L is extracted in extraction pot, raw material weight 0.5% is pressed in extraction pot and adds stabilizer
BHA, obtains extracting solution A；The pH value of extracting solution A is adjusted to into 3.5 with 10% hydrochloric acid solution, centrifugation obtains supernatant C；Supernatant C
Separate into moving-bed adsorption, adsorbent is D101 nonpolar macroporous adsorption resins, and water wash zone is pure water, and strippant is
60% ethanol, resin absorption regenerated solvent be 3% sodium hydrate aqueous solution, adsorption zone flow velocity 1.5BV/h, the BV/ of water wash zone flow velocity 2
H, the BV/h of desorption zone flow velocity 3, the BV/h of renewing zone flow velocity 3, switching time 800s, at 45 DEG C, Stress control exists temperature control
0.3Mpa, obtains eluent D；Concentrated juice solid quality percentage composition is 30%, then the freeze-dried Herba Hyperici perforati that obtains is extracted
Thing.Jing determine, in extract containing 6.3 weight % hypericins, 3.1 weight % hyperforines, 81.2 weight % flavonoids into
Point.
Embodiment 2
Take Herba Hyperici perforati and be dried herb（Including flower and fruit）1kg, crushes, and crosses 40 mesh sieves；Raw material weight 0.4L water is added, is adjusted
Section pH adds xylanase and pectase 3 to 4.0 in the ratio of raw material weight 1.5%:1 mixed enzyme, the closed stirring of lucifuge
2h is processed, 80 DEG C are heated to before terminating, continue 5min, be down to room temperature, separation discards liquid, the raw material after must processing；Plus
1% sodium hydrate aqueous solution for entering 20L is extracted in extraction pot, raw material weight 0.5% is pressed in extraction pot and adds stabilizer
BHA, obtains extracting solution A；The pH value of extracting solution A is adjusted to into 3.5 with 10% hydrochloric acid solution, centrifugation obtains supernatant C；Supernatant C
Separate into moving-bed adsorption, adsorbent is D101 nonpolar macroporous adsorption resins, and water wash zone is pure water, and strippant is
60% ethanol, resin absorption regenerated solvent be 3% sodium hydrate aqueous solution, adsorption zone flow velocity 1.5BV/h, the BV/ of water wash zone flow velocity 2
H, the BV/h of desorption zone flow velocity 3, the BV/h of renewing zone flow velocity 3, switching time 800s, at 45 DEG C, Stress control exists temperature control
0.3Mpa, obtains eluent D；Concentrated juice solid quality percentage composition is 30%, then the freeze-dried Herba Hyperici perforati that obtains is extracted
Thing.Jing determine, in extract containing 6.0 weight % hypericins, 2.8 weight % hyperforines, 79.5 weight % flavonoids into
Point.
Claims (7)
1. a kind of preparation method of Herba Hyperici perforati extract, comprises the steps：
（1）Take Herba Hyperici perforati and be dried herb（Including flower and fruit）, crush, cross 40 mesh sieves；Add raw material weight 20-40%'s
Water, adjusts pH to 4.0-6.0, and adds enzyme preparation, closed stir process 1-3h of lucifuge, knot in the ratio of raw material weight 0.5-3%
60-80 DEG C is heated to before beam, continues 5-15min, be down to room temperature, separation discards liquid, the raw material after must processing；
（2）The raw material after processing is taken, is extracted in extraction pot by Extraction solvent of alkali liquor, raw material weight is pressed in extraction pot
0.1-1% adds stabilizer, obtains extracting solution A；
（3）The pH value of extracting solution A is adjusted to into acidity, centrifugation obtains supernatant C；
（4）Supernatant C is isolated and purified through simulated moving bed chromatography, eluent D is obtained；
（5）By eluent D concentrations, then it is freeze-dried obtain Herba Hyperici perforati extract, extract contains Determination of Hypericin from Extraction.
2. preparation method as claimed in claim 1, step（1）In enzyme preparation can be xylanase, pectase or it is mixed
The mixture of compound, preferred xylanase and pectase, particularly xylanase and pectase 3:1 mixed enzyme；Enzyme preparation
Preferred 1-2 weight % of addition, more preferably 1.5 weight %；The intensification for terminating before processing preferably reaches 80 DEG C, continues 5min.
3. preparation method as claimed in claim 1 or 2, step（2）Alkali liquor be 1% sodium hydroxide solution, every kilogram of raw material
Using 20-25L Extraction solvents, 40-60 DEG C of extraction 2-3h；Stabilizer can be Butylated hydroxyanisole or butylated hydroxy-a
Benzene.
4. preparation method as claimed in claim 1 or 2, step（3）In, adjust pH to 3-4 using 10% hydrochloric acid solution；From
Scheming rotating speed is 4000 turns/min, and centrifugation time is 10min.
5. preparation method as claimed in claim 1 or 2, step（4）In, the adsorbent of simulated moving bed chromatography filling is non-pole
Property macroporous adsorbent resin, preferred D101 non-polar resins or HPD100 non-polar resins, water wash zone is pure water, and strippant is 60%
Ethanol, resin absorption regenerated solvent is 95% ethanol or 2-4% sodium hydrate aqueous solutions, and adsorption zone flow velocity 1-2BV/h, water wash zone is flowed
Fast 1-3 BV/h, desorption zone flow velocity 1-3 BV/h, renewing zone flow velocity 2-3 BV/h, switching time 600-800s, temperature control exists
40-60 DEG C, Stress control is in 0.1-0.5Mpa.
6. preparation method as claimed in claim 1 or 2, step（5）In, eluent D is concentrated into into solid quality percentage and is contained
Measure as 30-40%, then freeze-dried obtain Herba Hyperici perforati extract.
7. Herba Hyperici perforati extract obtained in the preparation method as described in claim 1-4, includes in the extract and is more than 6.0 weights
The amount hypericin of %, the hyperforine more than 2.0 weight %, more than the flavones ingredient of 75 weight %.
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Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN109394843A (en) * | 2019-01-03 | 2019-03-01 | 重庆工商大学 | A method of preparing the Hypericum Perforatum P.E rich in hypericin |
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| CN1392128A (en) * | 2002-07-18 | 2003-01-22 | 中山大学 | Process for preparing extract containing hypericin and flavon compounds |
| CN1392129A (en) * | 2002-07-18 | 2003-01-22 | 中山大学 | Process for preparing hypericum perforatum extract |
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| CN1198097A (en) * | 1995-09-29 | 1998-11-04 | 威廉施瓦布博士有限公司 | Stable extract of hypericum perforatum L., process for preparing the same and pharmaceutical compositions |
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| CN103450000A (en) * | 2013-10-08 | 2013-12-18 | 白心亮 | Method for extracting hypericin from hyperforin perforatum |
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Cited By (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN109394843A (en) * | 2019-01-03 | 2019-03-01 | 重庆工商大学 | A method of preparing the Hypericum Perforatum P.E rich in hypericin |
| CN109394843B (en) * | 2019-01-03 | 2021-03-26 | 重庆工商大学 | Method for preparing hypericum perforatum extract rich in hypericin |
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Application publication date: 20170517 |