CN107693558A - The separation method of general ginsenoside - Google Patents
The separation method of general ginsenoside Download PDFInfo
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- CN107693558A CN107693558A CN201711097772.1A CN201711097772A CN107693558A CN 107693558 A CN107693558 A CN 107693558A CN 201711097772 A CN201711097772 A CN 201711097772A CN 107693558 A CN107693558 A CN 107693558A
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- A61K2236/50—Methods involving additional extraction steps
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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
- A61K2236/55—Liquid-liquid separation; Phase separation
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Abstract
The invention provides a kind of separation method of general ginsenoside, including：Ginseng pulverate is added in Extraction solvent, at room temperature normal pressure ultrasonic extraction, then centrifuged, collect liquid, and diluted so as to obtain Ginseng extract with Extraction solvent；Multi-walled carbon nanotube is added in Ginseng extract total saposins are adsorbed, then centrifuged, collect solid；Solid is added in eluting solvent and eluted, is then centrifuged, eluent is collected, that is, obtains general ginsenoside solution.The method separative efficiency of the present invention is high, it is time-consuming it is short, solvent-oil ratio is small, simple to operate and device is simple.
Description
Technical field
The invention belongs to natural medicinal ingredients preparing technical field, in particular it relates to a kind of general ginsenoside
Separation method, more particularly it relates to a kind of method of extraction and quick separating general ginsenoside from ginseng.
Background technology
Ginseng (Panax ginseng C.A.Mayer) is perennial araliaceae ginseng plant, main product in East Asia Region,
In the pharmaceutical ground of the existing more than one thousand years of China.Qin-Han times《Sheng Nong's herbal classic》It is classified as top grade.Ginsenoside is by soap
Aglycon be connected with sugared substituent form a kind of glycoside compound, be ginseng play anti-aging, it is antitumor, hypoglycemic, improve in
The main active of the pharmacological action such as secretion and regulation immune system.Ginsenoside in ginseng be divided into protopanaxadiol-type,
Protopanaxatriol type and oleanolic acid type.Protopanaxadiol-type's saponin(e mainly includes Rb1, Rb2, Rc and Rd；Protopanaxatriol type
Saponin(e mainly includes Rg1, Re and Rf；Oleanolic acid type saponin mainly includes Ro.This seven kinds of saponin(es are the main saponin(es in ginseng,
Content accounts for more than the 70% of total saposins.At present, the method for extracting general ginsenoside mainly has ultrasound assisted extraction, Microwave-assisted Extraction
Take, SPE, pressure fluid extraction, supercritical fluid extraction etc., be mainly water suitable for industrial Extraction solvent.Carry
Take the impurity such as the sugar in thing generally to utilize macroreticular resin, separated using organic solvents such as ethanol as eluant, eluent and purify the total soap of ginseng
Glycosides.Water-saturated n-butanol extraction is the method for another conventional separation general ginsenoside.Both approaches are selectively good, behaviour
Make simply, but be required for consuming substantial amounts of organic solvent and time, it is impossible to general ginsenoside and sugar are separated in a short time, it is difficult
To realize that high flux obtains the medicinal ingredients such as ginsenoside.
CNT is that have nano level one-dimensional tunnel structure carbon material, can be divided into single wall and multi-walled carbon nanotube.
Single-walled carbon nanotube is formed by mono-layer graphite curling, aperture 1-2nm.Multi-walled carbon nanotube is made up of Multi-layer graphite, and aperture can
To reach 2-50nm.CNT has very high specific surface area, good chemistry, machinery and heat endurance, can be formed
Stable π-π interactions.These design features make it show many unique performances.In Analysis of Chinese Traditional Medicine field, carbon nanometer
Pipe is mainly used in the residues of pesticides being enriched with Chinese medicine, and the application in terms of traditional Chinese medicine extyaction separation is still less, and uses
Report is then had no in general ginsenoside separation.
The content of the invention
The goal of the invention of the present invention is the defects of being directed to prior art, there is provided a kind of separation method of general ginsenoside.
The separation method of the general ginsenoside of the present invention, comprises the following steps：
(1) ginseng pulverate is added in Extraction solvent, at room temperature normal pressure ultrasonic extraction, then centrifuged, collector
Join extract solution, and the Ginseng extract so as to be diluted is diluted with Extraction solvent；
(2) multi-walled carbon nanotube is added in the Ginseng extract of dilution and total saposins is adsorbed, then centrifugation point
From collection solid；
(3) solid is added in eluting solvent and eluted, then centrifuged, collect eluent.
Further, it is 1 according to the w/v of ginseng pulverate and Extraction solvent in step (1):Particle diameter is by (2-5)
The ginseng pulverate of 20 mesh to 60 mesh is added in Extraction solvent, at room temperature normal pressure ultrasonic extraction 10 minutes to 20 minutes, then with
4000rpm to 8000rpm is centrifuged 3 minutes to 5 minutes, collects Ginseng extract, and diluted 10- with Extraction solvent
100 times so as to the Ginseng extract diluted.
Further, the Extraction solvent is any one in following solvent：Methanol, ethanol, water, methanol or ethanol with
The binary mixed solvent that water is formed.
Further, in step (2), the w/v of the multi-walled carbon nanotube and the Ginseng extract of the dilution
It is 1:(100-200), preferably 1:160.
Further, in step (2), the internal diameter of the multi-walled carbon nanotube is 2nm to 16nm, and joint length is 0.5 μm
To 10 μm.
Further, in step (2), centrifuged 1 minute to 5 minutes with 6000rpm to 8000rpm revolution；It is preferred that
Ground, centrifuged 2 minutes with 8000rpm revolution.
Further, it is characterised in that in step (3), the eluting solvent is ethanol, acetonitrile, acetone, normal propyl alcohol, just
Butanol or acetone；
Or the eluting solvent is that the binary that ethanol, acetonitrile, acetone, normal propyl alcohol, n-butanol or acetone are formed with water is mixed
Bonding solvent, wherein, the volume ratio of the eluting solvent reclaimed water is no more than 30%.
Further, in step (3), centrifuged 1 minute to 5 minutes with 6000rpm to 8000rpm revolution；It is preferred that
Ground, centrifuged 2 minutes with 8000rpm revolution.
Further, in step (3), solid is added in eluting solvent and carries out recycling elution, preferred cycle elution two
It is secondary.
Further, method of the invention also includes：The solid being centrifugally separating to obtain in step (3) is precipitated into calcining to live
Change.
Technical scheme has the advantages that：
The method of the present invention is adsorbed using multi-walled carbon nanotube to the ginsenoside in ginseng extract, passes through more walls
π-the π of CNT tube wall and ginsenoside aglycon interact, and form stable adhesion, rapidly adsorb Ginseng extract
In ginsenoside.The inertia tube wall of the stronger component of sugared isopolarity and multi-walled carbon nanotube does not interact in extract solution,
It can not enter in the one-dimensional channels of multi-walled carbon nanotube, not adsorbed, be retained in Ginseng extract by multi-walled carbon nanotube.Gu
The multi-walled carbon nanotube of phase can utilize the method quick separating of centrifugation with liquid phase Ginseng extract.
Compared to traditional organic solvent deposit except sugar and water-saturated n-butanol extract and separate general ginsenoside method, this hair
Bright method saves plenty of time cost, and reduces consumption quantity of solvent.Further, there is method of the invention operation to walk
Rapid simple, separative efficiency is high, and general ginsenoside loss amount is few, for different Panax sample universals it is good the characteristics of, and fill
Put simply, be easy to be fabricated to large automatic device.
Brief description of the drawings
Fig. 1 is that the High Performance Liquid Chromatography/Mass Spectrometry of Ginseng extract in embodiment 1 scans total ion current figure.
Fig. 2 is the High Performance Liquid Chromatography/Mass Spectrometry full scan of the Ginseng extract after multi-walled carbon nanotube absorption in embodiment 1
Total ion current figure.
Fig. 3 be embodiment 1 in from CNT desorption general ginsenoside High Performance Liquid Chromatography/Mass Spectrometry full scan always from
Subflow figure.
Fig. 4 is the superposition total ion current figure that MRM modal cutoffs detect 8 kinds of ginsenosides in embodiment 1.
Embodiment
In order to be fully understood by the purpose of the present invention, feature and effect, by following embodiments, the present invention is made detailed
Describe in detail bright.For the process of the present invention in addition to the description below, remaining uses the conventional method or device of this area.Following nouns
Term is unless otherwise stated, be respectively provided with the implication that those skilled in the art are generally understood that.
Currently, it is also high without a kind of simple to operate, time-consuming short and separative efficiency in terms of general ginsenoside separation and Extraction
Method.For the defect, the invention provides a kind of method of quick separating general ginsenoside, including：(1) ginseng pulverate is added
Into Extraction solvent, normal pressure ultrasonic extraction, is then centrifuged at room temperature, collects Ginseng extract, and will with Extraction solvent
It is diluted so as to the Ginseng extract diluted；(2) multi-walled carbon nanotube is added in the Ginseng extract of dilution to total
Saponin(e is adsorbed, and is then centrifuged, and collects solid；(3) solid is added in eluting solvent and eluted, then from
The heart separates, and collects eluent.This method can be realized in the short time by using the selective absorption performance of multi-walled carbon nanotube
General ginsenoside is efficiently separated from ginseng extract, reaches the effect for rapidly purifying general ginsenoside in complicated ginseng extract
Fruit.
Specifically, the method for quick separating general ginsenoside of the invention comprises the following steps：
The first step, extract Ginseng extract：Ginseng pulverate is added in Extraction solvent, at room temperature normal pressure ultrasonic extraction,
Then centrifuge, collect Ginseng extract, and the Ginseng extract so as to be diluted is diluted with Extraction solvent.
In the present invention, " ginseng " refers to araliaceae ginseng plant, includes but is not limited to, ginseng and its processed product are (red
Ginseng), American Ginseng, pseudo-ginseng and Vietnam's ginseng etc..The method of the present invention does not have particular/special requirement to the source of ginseng, to any source
Ginseng is applicable, the ginseng mainly planted in the following embodiments using Northeast Area of China, but this is exemplary
's.
In a kind of preferred embodiment, Ginseng extract is extracted with the following method：
First, fresh ginseng is cleaned, dried by the way of 20 DEG C of -40 DEG C of low-temperature air-dryings, then by dry people
Ginseng is crushed to 20 mesh to 60 mesh (national standard sieve).
Then, Extraction solvent is added in appropriate ginseng pulverate, the quality of ginseng pulverate and the volume ratio of Extraction solvent are 1:2
To 1:5.Normal pressure ultrasonic extraction ginseng pulverate 10 minutes to 20 minutes at room temperature, then centrifuge, collect Ginseng extract.It is excellent
Selection of land, repeat the step twice, and merge the Ginseng extract of collection.
Finally, according to 1:10-1:100 ratio (v:V) Ginseng extract collected is diluted with Extraction solvent, is diluted
Ginseng extract.
Preferably, Extraction solvent can be methanol, ethanol or water, or, Extraction solvent can also be methanol or ethanol with
The binary mixed solvent that water is formed, wherein, the volume ratio that Extraction solvent reclaimed water accounts for is no more than 20%.
Preferably, in above-mentioned centrifugal separation processes, centrifugation revolution is 4000rpm to 8000rpm, and the time is 3 points
Clock was to 5 minutes, and preferably 5000rpm, 4 minutes.
Second step, adsorption step：Multi-walled carbon nanotube is added in the Ginseng extract of dilution and adsorbs total soap therein
Glycosides, then centrifuge, collect solid, that is, adsorbed the multi-walled carbon nanotube of general ginsenoside.It is centrifugally separating to obtain
Liquid is the Ginseng extract after absorption.
Preferably, it is 1 according to the w/v of multi-walled carbon nanotube and the Ginseng extract of dilution:(100-200) (more
It is preferred that 1:160) multi-walled carbon nanotube is added in the Ginseng extract of dilution, and the two is fully contacted, such as pass through whirlpool
Rotation (such as 10 seconds) makes multi-walled carbon nanotube fully be contacted with the Ginseng extract diluted, total in Ginseng extract to adsorb
Saponin(e.
Preferably, the internal diameter of multi-walled carbon nanotube is 2nm to 16nm, and joint length is 0.5 μm to 10 μm.The present invention is to more
The external diameter of wall carbon nano tube does not require that conventional outside dimension is applicable (such as external diameter is 18nm to 37nm).Conventional commercial
Multi-walled carbon nanotube be used equally in the present invention, for example, commercially available from Nanometer Port Co., Ltd., Shenzhen.
Preferably, centrifuge in this step and centrifuged 1 minute to 5 minutes with 6000rpm to 8000rpm revolution；
It is highly preferred that centrifuged 2 minutes with 8000rpm revolution.
Measured respectively using phend-sulphuric acid dilution Ginseng extract and absorption after Ginseng extract in sugar contain
Amount, and calculate multi-walled carbon nanotube removes sugared rate.
Wherein, the specific steps of phend-sulphuric acid include：Two are added in the glucose control product of same volume various concentrations
The concentrated sulfuric acid of 6% phenol solution of/mono- volume and 2 times of volumes, vibration shake up, and boiling water bath is cooled to room temperature after 10 minutes,
Reference substance absorbance is determined at 490nm wavelength with ultraviolet-visible spectrophotometer, external standard method calculates sugar in Ginseng extract and contained
Amount.
Wherein, multi-walled carbon nanotube refers to except sugared rate：The ginseng of sugared content/dilution in Ginseng extract after absorption
Sugared content × 100% in extract solution.Except sugared rate is closer to 100%, illustrate multi-walled carbon nanotube to sugared in Ginseng extract
Suction-operated is lower, is more advantageous to the separation of sugar and ginsenoside.
By measurement and calculate, in the method for the invention, multi-walled carbon nanotube except sugared rate is 95%-100%, explanation
Multi-walled carbon nanotube is extremely low to suction-operated sugared in Ginseng extract.
3rd step, elution step：The multi-walled carbon nanotube for having adsorbed general ginsenoside is added in eluting solvent and carried out
Elution, then centrifuge, collect eluent, that is, the general ginsenoside solution being desorbed.The solid being centrifugally separating to obtain is
Multi-walled carbon nanotube.Wherein, general ginsenoside includes Rb1, Rb2, Rc, Rd, Re, Rg1, Rf, Ro.
Preferably, the w/v of multi-walled carbon nanotube and eluting solvent is 1:10-1:100.
Preferably, eluting solvent is ethanol, acetonitrile, acetone, normal propyl alcohol, n-butanol or acetone, or, eluting solvent is second
The binary mixed solvent that alcohol, acetonitrile, acetone, normal propyl alcohol, n-butanol or acetone are formed with water, wherein, the body of eluting solvent reclaimed water
Product is than being no more than 30%.
Preferably, centrifuge in this step and centrifuged 1 minute to 5 minutes with 6000rpm to 8000rpm revolution；
It is highly preferred that centrifuged 2 minutes with 8000rpm revolution.
Preferably, repeat this step (for example, repeating this step once), and the eluent collected every time merged,
The general ginsenoside solution being desorbed.
The general ginsenoside solution of desorption is determined using high performance liquid chromatography-triple quadrupole bar mass spectrography respectively and ginseng carries
Content of ginsenoside in liquid is taken, and calculates adsorption efficiency of the multi-walled carbon nanotube to ginsenoside.Wherein, first will before test
General ginsenoside solution is through membrane filtration, and filter sizes are preferably 0.22 μm, 0.45 μm, 0.6 μm, more preferably 0.22 μm.
Wherein, high performance liquid chromatography-triple quadrupole bar mass spectrography is as follows：
In high performance liquid chromatography program, mobile phase is respectively the aqueous formic acid of A phases 0.1%, B phase acetonitriles, chromatographically pure；Gradient
Elution requirement (volume fraction)：0-5min (25%B~30%B), 5-8min (30%-36%B), 8-15min (36%-48%
B), 15-20min (48%-70%B), 20-25min (70%-90%B), 25-28min (90%B), 28-34min (25%B)；
Flow velocity：0.2mL/min, using conventional C18 chromatographic columns (2.1mm × 100mm, 1.7 μm), column temperature is preferably 25-40 DEG C.Triple four
Pole bar mass ion source condition：ESI sources anion MRM patterns, sheath gas are preferably：30-42；Auxiliary gas be preferably：6-15；Purging
Gas is preferably：0-1；Ion transfer tube temperature is preferably 300-3600 DEG C, and spray voltage is preferably -2000--3000V.8 kinds of people
The quota ion pair information for joining saponin(e is as shown in table 1：
The target substance MRM mode detection parameters of table 1
8 kinds of ginsenoside hybrid standard product solution are configured, standard concentration is made with the peak area of hybrid standard product solution
Figure draws the standard curve of 8 kinds of ginsenosides, and each component peak area in testing sample is substituted into standard curve and calculates each component
Concentration.
Multi-walled carbon nanotube refers to the separative efficiency of ginsenoside：The ginseng of content of ginsenoside/dilution of desorption carries
Take content of ginsenoside × 100% in liquid.Separative efficiency represents multi-walled carbon nanotube from Ginseng extract closer to 100%
The content of ginsenoside of separation is more, and the loss of ginsenoside is fewer in separation process.
By measurement and calculate, in the method for the invention, multi-walled carbon nanotube is to the adsorption efficiency of ginsenoside
80%-100%, illustrate that the content of ginsenoside that multi-walled carbon nanotube separates from Ginseng extract is high, ginseng in separation process
The loss of saponin(e is lacked.
In the method for the invention, the separation of the general ginsenoside in ginseng extract and sugar is (i.e. from the extract solution of dilution
With multi-walled carbon nanotube contact start, untill eluting solvent separates with multi-walled carbon nanotube) only need to complete for 7 minutes,
So that the method fast and easy of the present invention.
Preferably, after separation terminates, calcining and activating is carried out to multi-walled carbon nanotube.Specifically, tube furnace is placed in nitrogen
Under gas atmosphere protection, 200-250 DEG C of simultaneously constant temperature 1 hour is warming up to 2-10 DEG C/min programming rate since room temperature, then
Room temperature is naturally cooled to, so as to reuse multi-walled carbon nanotube.
In the present invention, inventor is that 2nm to 16nm, external diameter are 18nm to 37nm, single length by studying selection internal diameter
Degree is that 0.5 μm to 10 μm of multi-walled carbon nanotube is used for the separation of general ginsenoside in Ginseng extract.
For other carbon nanomaterials, such as graphene, graphene oxide, amination graphene etc., due to these materials
Two-dimension plane structure, them is not had the type of selecting of multi-walled carbon nanotube, i.e., can not according to size, molecular weight to determinand select
Select absorption.And in the present invention, it is 2nm to 16nm multi-walled carbon nanotube by using internal diameter, enters ginsenoside molecule
In the duct of multi-walled carbon nanotube, and the polysaccharide constituents in Ginseng extract are unable in access aperture road because molecular weight is big, because
This is advantageous to the separation of ginsenoside and carbohydrate content.The ginsenoside realized using multi-walled carbon nanotube and carbohydrate content
Excellent separating effect is other carbon nanomaterials, such as graphene, graphene oxide, amination graphene etc., can not be realized
's.
The joint length for the multi-walled carbon nanotube that the present invention uses is 0.5 μm to 10 μm, the ginsenoside that can be adsorbed
Content is high, is advantageous to improve adsorption capacity (i.e. the adsorbable sample size of unit mass multi-walled carbon nanotube).Due to dimensions length
Advantage so that the multi-walled carbon nanotube that uses of the present invention can have adsorption capacity more more preferable than other carbon nanomaterials.
The methods of multi-walled carbon nanotube surface free used in the present invention aoxidizes is modified, and shows as chemical inertness.Compared to
The multi-walled carbon nanotube or graphene oxide of surface oxidation or the carbon nanomaterial of other surface modifications, more walls that the present invention uses
CNT has more preferable hydrophobicity, and good oleophobic property is shown in Extraction solvent and eluting solvent, therefore pole
Easily separated with dicyandiamide solution, greatly shorten disengaging time.
Other carbon nanomaterials of multi-walled carbon nanotube and such as graphene are all by sp2The six of the carbon atom composition of hydridization
Yuan of rings periodic arrangement is formed, but unlike other carbon nanomaterials, multi-walled carbon nanotube is rolled up by Multi-layer graphite piece
It is curved into one-dimensional channels (and graphene is the two-dimension single layer graphite flake that is formed by hexatomic ring).This periodicity six-membered ring structure
Conjugatedπbond be present, metastable π-π interactions can be formed with the compound of similar structure.Aglycon in ginsenoside
It with similar structure, therefore can be adsorbed by it, with reference to relatively stable, but agent can be eluted and eluted easily.And graphene
Two-dimension plane structure compared with the one-dimentional structure of the bending of multi-walled carbon nanotube, can produce stronger active force to absorbate and
The problem of causing to be not easy to be desorbed.In addition, for other materials, such as Flavonoid substances, because these materials contain phenyl ring, double
The big conjugatedπbond such as key, phenolic hydroxyl group and strong electron-withdrawing group group, it will also result in difficult the problem of being desorbed after absorption.Therefore, inventor passes through
Research finds, separation of the multi-walled carbon nanotube particularly suitable for general ginsenoside in Ginseng extract.
In addition, multi-walled carbon nanotube technology of preparing in the market, characterizing method are all highly developed, and conduct
Commodity are commercially available on a large scale.And the material such as graphene oxide, graphene, ammonification graphene can not be accomplished to mass produce at present,
It is and costly.Therefore, method material of the invention easily obtains and cost is low.
Embodiment
The present invention is further illustrated below by the mode of embodiment, but does not therefore limit the present invention to described reality
Apply among a scope.The experimental method of unreceipted actual conditions in the following example, conventionally and condition, or according to business
Product specification selects.
The phend-sulphuric acid used in following embodiments is specifically：
Glucose control product 10mg accurately is weighed, 0.1g/L solution is prepared into water.Respectively draw 0.2,0.4,0.6,
0.8th, 1.0,1.2,1.4,1.6,1.8,2.0mL is placed in tool plug test tube, is supplied less than 2mL with water, is then added the examination of 6% phenol
Agent 1mL, then concentrated sulfuric acid 5mL is slowly added to pipette, shaken well, boiling water bath 10min, it is cooled to room temperature.Use ultraviolet-visible
Spectrophotometer determines reference substance solution absorbance A at 490nm wavelength.It is abscissa by ordinate, mass concentration of A, paints
Glucose standard curve equation processed.The A of measure Ginseng extract and the Ginseng extract after absorption substitutes into standard curve side respectively
Cheng Zhong, the sugared content in Ginseng extract before and after carbon nanotube adsorption is calculated.
The high performance liquid chromatography used in following embodiments-triple quadrupole bar mass spectrum quantitative analysis ginseng extract solution and ginseng
8 kinds of ginsenoside Rb1s in total saposins solution, Rb2, Rc, Rd, Re, Rg1, Rf, the detecting step of Ro contents are：
8 kinds of ginsenoside hybrid standard product solution of configuration, concentration 0.5,1.0,2.0,4.0,8.0,16.0,32.0,
64.0,100.0 μ g/mL.Ginseng extract and general ginsenoside solution be configured to the solution of same concentrations, all preparations it is molten
Liquid crosses 0.22 μm of filter membrane, is detected into High Performance Liquid Chromatography/Mass Spectrometry using foregoing routine, wherein 35 DEG C of column temperature, sheath gas：38,
Aid in gas：11, purge gass：1, ion transfer tube temperature：329 DEG C, spray voltage：-2500V.With the peak of hybrid standard product solution
Area is mapped to ginsenoside concentration draws the standard curve of 8 kinds of ginsenosides, and each component peak area in testing sample is substituted into
Each component concentration is calculated in standard curve.
The multi-walled carbon nanotube used in following embodiments is bought from Nanometer Port Co., Ltd., Shenzhen, wherein, embodiment 1
The specification of the multi-walled carbon nanotube of use is that internal diameter is 3nm to 12nm, and external diameter is 28nm to 34nm, and joint length is 1 μm to 5 μ
M, the specification for the multi-walled carbon nanotube that embodiment 2 to 4 uses is that internal diameter is 2nm to 6nm, and external diameter is 15nm to 20nm, single length
Degree is 3 μm to 8 μm, and the specification for the multi-walled carbon nanotube that embodiment 5 to 7 uses is that internal diameter is 6nm to 15nm, external diameter be 21nm extremely
30nm, joint length are 0.8 μm to 6 μm.
Embodiment 1
(1) Ginseng extract is extracted
The root of fresh ginseng is taken, is cleaned, is positioned in the air dry oven that temperature control is 35 DEG C and dries to constant weight.It is dry
Dry ginseng is crushed with pulverizer, is crossed 40 mesh sieves and is obtained ginseng pulverate.
5g ginseng pulverates are taken to be added in the ethanol of 15 milliliter 80% (volume fraction), room temperature normal pressure ultrasonic extraction 15 minutes.
4000rpm is centrifuged 5 minutes, separates ginseng pulverate.The ginseng pulverate of separation repeats extraction 2 times using aforementioned condition, and it is molten to merge extraction
Agent, produce the Ginseng extract of dilution again with 80% ethanol dilution extract solution 10.
(2) adsorb
0.3 gram of multi-walled carbon nanotube is taken to be placed in the Ginseng extract of 50 milliliters of dilutions, being vortexed 10 seconds makes multi-wall carbon nano-tube
Manage and fully contacted with the Ginseng extract diluted.Then centrifuged 2 minutes with 8000rpm, after separation multi-walled carbon nanotube and absorption
Ginseng extract.Using phend-sulphuric acid quantitative analysis ginseng extract solution with absorption after Ginseng extract in sugared content,
Except sugared rate is 99.6% in the present embodiment.
(3) elute
The CNT of separation is immersed in 20 milliliters of water-saturated n-butanol solution, and being vortexed 10 seconds makes CNT and positive fourth
Alcoholic solution fully contacts, and then 8000rpm centrifuges 2 minutes separating carbon nano-tubes and butanol solution.After repeating 1 time, close
And water-saturated n-butanol solution, obtain general ginsenoside solution.With high performance liquid chromatography-triple quadrupole bar mass spectrum quantitative analysis person
Join 8 kinds of ginsenoside Rb1s in extract solution and general ginsenoside solution, Rb2, Rc, Rd, Re, Rg1, Rf, Ro content.This implementation
The desorption efficiency of 8 kinds of ginsenosides is 87.5-96.3% in example.
CNT after separation is placed in 25 DEG C of air dry ovens and dried, and is subsequently placed in tube furnace and is continuously flowing
Calcining and activating is carried out in the atmosphere of nitrogen protection, calcination procedure is：200 DEG C and constant temperature 1 are warming up to 4 DEG C/min since room temperature
Hour, then naturally cool to room temperature.
Fig. 1 is the full scan total ion current figure of the High Performance Liquid Chromatography/Mass Spectrometry of the gained Ginseng extract of embodiment 1, from this
It can be seen from the figure that, the saponin(e being largely extracted are efficiently separated, and component of the retention time before 1 minute is carbohydrate, because
It is very high for polarity, so the reservation in C18 chromatographic columns is very weak, flow out at first.Fig. 2 is that multi-walled carbon nanotube is inhaled in embodiment 1
The full scan total ion current figure of the High Performance Liquid Chromatography/Mass Spectrometry of attached Ginseng extract, it will be apparent from this figure that more wall carbon
Ginsenoside has been can't detect in Ginseng extract after nanotube absorption, and carbohydrate then remains in Ginseng extract
In.Fig. 3 is the full scan of the High Performance Liquid Chromatography/Mass Spectrometry for the general ginsenoside being desorbed in embodiment 1 from multi-walled carbon nanotube
Total ion current figure, by the it can be seen from the figure that, the key component being desorbed in solution is identical with Ginseng extract, illustrates by carbon
The ginsenoside of nanotube absorption can be eluted solvent desorbing.By Fig. 1 and Fig. 3 it can also be seen that between the ginsenoside of part
There is no baseline separation, such as Re and Rg1, this will reduce the accuracy of ginsenoside quantitative analysis results.By using triple four
The mass spectrographic MRM scan patterns of pole bar mass spectrum can solve this problem, as shown in figure 4, under MRM scan patterns, 8 kinds of ginseng soaps
Glycosides independently detects respective signal response, improves the accuracy of content of ginsenoside analysis.
Embodiment 2
For experimental method with embodiment 1, the Extraction solvent when difference with embodiment 1 is to extract Ginseng extract is first
Alcohol, 4000rpm are centrifuged 10 minutes.The desorption efficiency of ginsenoside is 88.2-94.8%.
Embodiment 3
For experimental method with embodiment 1, the Extraction solvent when difference with embodiment 1 is to extract Ginseng extract is 95%
Ethanol, 8000rpm are centrifuged 4 minutes.The desorption efficiency of ginsenoside is 85.1-95.4%.
Embodiment 4
Experimental method is with embodiment 1, the multi-walled carbon nanotube when difference with embodiment 1 is to extract Ginseng extract
Quality is 0.5 gram, is vortexed 25 seconds, and 6000rpm is centrifuged 6 minutes.The desorption efficiency of ginsenoside is 89.3-97.6%.
Embodiment 5
Experimental method with embodiment 1, when the difference with embodiment 1 is to elute the desorption solvent that uses for acetone,
6000rpm is centrifuged 6 minutes.The desorption efficiency of ginsenoside is 84.7-92.5%.
Embodiment 6
Experimental method is with embodiment 1, and the desorption solvent that the difference with embodiment 1 is to use when eluting is 70% ethanol.
Ginsenoside desorption efficiency is 86.2-97.6%.
Embodiment 7
Experimental method is with embodiment 1, and the desorption solvent that the difference with embodiment 1 is to use when eluting is methanol.Ginseng
Saponin(e desorption efficiency is 83.2-91.8%.
The present invention is hereinbefore disclosed with preferred embodiment, but it should be understood by those skilled in the art that, these
Embodiment is only used for describing the present invention, and should not be construed as limiting the scope of the present invention.It should be noted that every implement with these
Example equivalent change and displacement, all should be set to be covered by scope of the presently claimed invention.Therefore, protection scope of the present invention
It should be defined by the scope defined in claims.
Claims (10)
1. a kind of separation method of general ginsenoside, it is characterised in that comprise the following steps：
(1) ginseng pulverate is added in Extraction solvent, at room temperature normal pressure ultrasonic extraction, then centrifuged, collected ginseng and carry
Liquid is taken, and the Ginseng extract so as to be diluted is diluted with Extraction solvent；
(2) multi-walled carbon nanotube is added in the Ginseng extract of dilution and total saposins is adsorbed, then centrifuged, received
Collect solid；
(3) solid is added in eluting solvent and eluted, then centrifuged, collect eluent.
2. separation method according to claim 1, it is characterised in that in step (1), according to ginseng pulverate and Extraction solvent
W/v is 1:Particle diameter is added in Extraction solvent by (2-5) for the ginseng pulverate of 20 mesh to 60 mesh, and normal pressure surpasses at room temperature
Sound extracts 10 minutes to 20 minutes, is then centrifuged 3 minutes to 5 minutes with 4000rpm to 8000rpm revolution, collector
Join extract solution, and 10 to 100 times are diluted so as to the Ginseng extract diluted with Extraction solvent.
3. separation method according to claim 1, it is characterised in that the Extraction solvent is any one in following solvent
Kind：The binary mixed solvent that methanol, ethanol, water or methanol or ethanol are formed with water.
4. separation method according to claim 1, it is characterised in that in step (2), the multi-walled carbon nanotube with it is described
The w/v of the Ginseng extract of dilution is 1:(100-200), preferably 1:160.
5. separation method according to claim 1, it is characterised in that in step (2), the internal diameter of the multi-walled carbon nanotube
It is 2nm to 16nm, joint length is 0.5 μm to 10 μm.
6. separation method according to claim 1, it is characterised in that in step (2), with turning for 6000rpm to 8000rpm
Number centrifuges 1 minute to 5 minutes；Preferably, centrifuged 2 minutes with 8000rpm revolution.
7. separation method according to claim 1, it is characterised in that in step (3), the eluting solvent is ethanol, second
Nitrile, acetone, normal propyl alcohol, n-butanol or acetone；
Or the two end number mixing that the eluting solvent is ethanol, acetonitrile, acetone, normal propyl alcohol, n-butanol or acetone with water is formed is molten
Agent, wherein, the volume ratio of the eluting solvent reclaimed water is no more than 30%.
8. separation method according to claim 1, it is characterised in that in step (3), with turning for 6000rpm to 8000rpm
Number centrifuges 1 minute to 5 minutes；Preferably, centrifuged 2 minutes with 8000rpm revolution.
9. separation method according to claim 1, it is characterised in that in step (3), solid is added in eluting solvent
Recycling elution is carried out, preferred cycle elutes twice.
10. separation method according to claim 1, it is characterised in that further comprise：It will be centrifuged in step (3)
Obtained solid precipitation calcining and activating.
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| CN108553494A (en) * | 2018-06-11 | 2018-09-21 | 通化鑫业生物科技研发有限公司 | Fresh ginseng activity extract and its application in preparing mitigation surgical patient pain medication |
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| CN101781351A (en) * | 2009-01-21 | 2010-07-21 | 重庆华森制药有限公司 | Method for extracting ginsenoside Rb1 from American ginseng and powder-injection thereof |
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN101781351A (en) * | 2009-01-21 | 2010-07-21 | 重庆华森制药有限公司 | Method for extracting ginsenoside Rb1 from American ginseng and powder-injection thereof |
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| 张崇禧等: "不同方法提取人参总皂苷工艺的优化研究", 《人参研究》 * |
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| CN108553494A (en) * | 2018-06-11 | 2018-09-21 | 通化鑫业生物科技研发有限公司 | Fresh ginseng activity extract and its application in preparing mitigation surgical patient pain medication |
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