CN110204590A - The extracting method of Celastrol - Google Patents
The extracting method of Celastrol Download PDFInfo
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- CN110204590A CN110204590A CN201910585654.8A CN201910585654A CN110204590A CN 110204590 A CN110204590 A CN 110204590A CN 201910585654 A CN201910585654 A CN 201910585654A CN 110204590 A CN110204590 A CN 110204590A
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Abstract
The invention discloses a kind of extracting methods of Celastrol, using Fagopyrum plant as raw material, liquid nitrogen grinding, enzymolysis processing are passed sequentially through, ultrasonic extraction, extraction, silica gel post separation and recrystallization process, the Celastrol high income finally extracted, purity is high, and a kind of new raw material and extracting method are provided for the extraction process of Celastrol, low raw-material cost is applicable to industrialized production.
Description
Technical field
The invention belongs to active components of plants extractive technique fields, and in particular to a kind of extracting method of Celastrol.
Background technique
Celastrol is eponymous with qinghaosu, is classified as by " cell " magazine and most possibly is developed to be the 5 of modern medicines
One of natural medicinal compound of kind tradition.Celastrol is one of traditional Chinese medicine active ingredients of thunder god vine.Harvard is big within 2015
The institute Umut Ozcan that studies medicine professor research team reports the weight-reducing of Celastrol and leptin sensitization for the first time more makes this
The one fashionable world of traditional Chinese medicine ingredient.In the same year, the Elisabetta Mueller from U.S. NIH is it has also been found that Celastrol swashs
Heat shock factor HSF1 living can increase energy consumption and mouse is helped to resist the obesity that high fat diet induces.This shows to pass through Thunder God
Rattan red pigment activation HSF1 can be used as fat and associated metabolic syndrome potential treatment means and further be developed, this for
Fat and associated metabolic syndrome treatment has certain prompt meaning.Cinta Blade is also indicated that from food compounds
Leptin can be regulated and controled and achieve the purpose that weight-reducing.Umut Ozcan team further investigations have shown that, in central nervous system
IL1R1 can be used as the major target class of Celastrol, provide potential therapeutic strategy for treatment obesity and related disease.
Umut professor Ozcan also found that Celastrol reduces cholesterol levels, improve liver function and glucose metabolism, these
Collective effect can reduce the risk of heart disease, fatty liver and diabetes B.However Celastrol commercial synthesis step is more,
It is at high cost, it is difficult to industrialization；Celastrol is present on a small quantity in tripterygium plant root, but the root of the platymiscium and flower are also
Many other toxic compound compositions, to lose weight, there may be danger by taking some extracts of tripterygium wilfordii by people；And
And balsam pear Celastrol (Charantin) content is extremely low, Celastrol function is dose dependent effect, therefore is hindered
Celastrol is developed into the process of Xenical.
Celastrol is a kind of natural products with multiple biological activities, initially extracts from the plant of Celastraceae Thunder God Calamus
The root skin of object tripterygium wilfordii then plants tripterygium hypoglaucum hutcs, Celastraceae Celastrus plant celastrus orbiculatus, Celastraceae in Celastraceae Thunder God Calamus
Valve Teng belongs to monotype platymiscium valve Teng and the extraction separation acquisition of Curcurbitaceae balsam pear platymiscium balsam pear forever forever.Also there are other reports i.e. with plant
Object cell culture processes prepare triptolide, Celastrol isoreactivity ingredient.But there is presently no from Fagopyrum plant
The middle report for extracting Celastrol.
Summary of the invention
For above-mentioned deficiency in the prior art, the present invention provides a kind of extracting method of Celastrol, the present invention
Celastrol directly is extracted from cymose buckwheat rhizome, bitter buckwheat and sweet tea buckwheat seed, leaf, stalk and root, is the extraction work of Celastrol
Skill provides a kind of new approaches.
To achieve the above object, the technical solution adopted by the present invention to solve the technical problems is:
A kind of extracting method of Celastrol, unlike the prior art, this method be from Fagopyrum plant into
Row extracts.
Further, Fagopyrum plant is cymose buckwheat rhizome, bitter buckwheat or sweet tea buckwheat.
Further, said extracted method the following steps are included:
(1) Fagopyrum plant seed, leaf, stalk and/or root is dry, liquid nitrogen is added and is ground, powder is obtained；
(2) powder is mixed with deionized water, and adjusting solution ph is 5.0-6.0, and 0.5-0.8% powder is then added
The mixed enzyme of weight is digested, and hydrolysis temperature is 25-50 DEG C, enzymolysis time 30-40min, is filtered, must be filtered after the completion of enzymatic hydrolysis
Slag；Wherein, mixed enzyme is the mixture that cellulase and pectase are 2-5:1 mixing by weight；
It (3) is that 1g:20-80mL is mixed by solid-to-liquid ratio with extracting solution by filter residue obtained by step (2), it is then close in ultrasonic power
Degree is 100-150W/cm2, supersonic frequency carry out ultrasonic extraction under the conditions of being 30-50kHz, Extracting temperature is 45-70 DEG C, is extracted
Time is 20-40min, is separated by solid-liquid separation to obtain supernatant；
(4) by volume it is that 1:3-8 is mixed with organic solvent by supernatant obtained by step (3), extracts 3-4 times, separation has
Then organic phase is concentrated under reduced pressure under the conditions of 40-50 DEG C, obtains medicinal extract by machine phase；
(5) by weight it is that 1:30-60 is mixed with the silica gel of 80-100 mesh by medicinal extract obtained by step (4), is then charged into layer
It analyses in column, is first eluted with petroleum ether, then carried out with the mix reagent that petroleum ether and ethyl acetate are 6-10:1 mixing by volume
Elution, the flow velocity for controlling eluent is 2-5mL/min, collects eluent, then does eluent under the conditions of 45-60 DEG C
It is dry；
(6) by step (5) gains with 30-40 DEG C, it is molten that volume fraction is that the ethanol solution of 70-80% is dissolved into saturation
Then cold water is added until crystal is no longer precipitated in liquid, crystal is dry, and Celastrol is made.
Further, detailed process in step (2) are as follows: powder is mixed with deionized water, and adjusts solution ph and is
5.5, the mixed enzyme that 0.6% powder weight is then added is digested, and hydrolysis temperature is 45 DEG C, enzymolysis time 30min, enzymatic hydrolysis
It filters after the completion, obtains filter residue.
Further, it is the mixture that 4:1 is mixed that mixed enzyme is cellulase and pectase in step (2) by weight.
Further, detailed process in step (3) are as follows: it is 1g that filter residue obtained by step (2) and extracting solution, which are pressed solid-to-liquid ratio:
Then 50mL mixing is 120W/cm in ultrasonic power density2, supersonic frequency be 35kHz under the conditions of carry out ultrasonic extraction, extract
Temperature is 60 DEG C, and extraction time 30min is separated by solid-liquid separation to obtain supernatant.
Further, in step (3) extracting solution be ethyl alcohol that volume fraction is 70%-85% and ionic liquid by volume
For 3-6:1-2 mixing mixture, preferably volume fraction be 75% ethyl alcohol and ionic liquid be by volume what 5:2 was mixed
Mixture.
Further, ionic liquid is at least one in [BMIM] Br, [BMIM] Cl, [BMIM] BF4 and [BMIM] PF6
Kind.
Further, in step (4) organic solvent be chloroform, methylene chloride, 1,2- dichloroethanes, carbon tetrachloride,
At least one of ethyl acetate and acetone.
It further, with the silica gel of 100 mesh is by weight that 1:35 is mixed by medicinal extract obtained by step (4) in step (5)；It washes
De- Shi Xianyong petroleum ether elution, then eluted with the mix reagent that petroleum ether and ethyl acetate are 8:1 mixing by volume, it controls
The flow velocity of eluent processed is 5mL/min.
The extracting method of Celastrol provided by the invention, has the advantages that
The stalk or leaf of Fagopyrum plant used in the present invention, conventional processing mode are to burn, this will cause money
Source waste and environmental pollution, Fagopyrum stalk etc. are not reasonably utilized.Fagopyrum stalk, the leaf that the present invention will discard
Son, root are recycled, and therefrom extract Celastrol, this provides a kind of new original for the extraction process of Celastrol
Material, and low raw-material cost, are applicable to industrialized production.
In extraction process of the present invention, Fagopyrum plant used is mainly cymose buckwheat rhizome, the seed of bitter buckwheat or sweet tea buckwheat, leaf, stem
Stalk and root are first dried seed, leaf, stalk and root, and liquid nitrogen is then added and is ground, liquid nitrogen temperature is very low, straw
Cells of these tissues of stalk, leaf, root, seed in liquid nitrogen can adfreezing, when grinding, can reach brokenly born of the same parents' effect, so that inside cell
Substance release come out.
But all cell ruptures can not be made during the grinding process, therefore, enzyme is carried out to gained powder after having ground
Solution processing.Because seed, leaf, stalk and root tissue have cell wall, contain a large amount of cellulose and pectin, the two phase in cell wall
Mutually interweave so that cell wall forms fine and close structure, therefore to sufficiently extract intracellular active material, need to cell wall into
Row processing.The selected enzyme of the application is the mixture that cellulase and pectase are 2-5:1 mixing by weight, then in spy
Determine under pH environment, the mixed enzyme that 0.5-0.8% powder weight is added is digested, and hydrolysis temperature is 25-50 DEG C, and enzymolysis time is
30-40min.On the basis of liquid nitrogen grinding, the case where for not broken born of the same parents, after being digested with this condition, can effectively it degrade
Cellulase and pectase in cell wall, release intracellular reactive ingredient.It is ground simultaneously by liquid nitrogen
Resulting powder powder is thinner after mill, and specific surface area is relatively large, and with enzyme contact area with regard to big, powder can sufficiently be dropped by enzyme
Solution shortens degradation time, saves production cost.
It, need to be from Multiple components to mesh after intracellular active constituent sufficiently releases when cell wall is sufficiently damaged
Ingredient extract, handled as the compounding substances that ultrasound forms filter residue obtained by after enzymatic hydrolysis and extracting solution.The step
According to " cavitation " of ultrasonic wave, on the one hand filter residue is dispersed, increase the active area of filter residue and extracting solution, improves reaction
Efficiency；Another aspect liquid enables the impurity for being attached to filter residue surface to fall off rapidly, so that extracting solution faster mentions
Purpose ingredient is got, and improves the purity of purpose ingredient.The present invention is by ultrasonic power density domination in 100-150W/cm2, ultrasound
Frequency control carries out ultrasonic extraction in 30-50kHz, and at 45-70 DEG C, extraction time is controlled in 20-40min model for Extracting temperature control
In enclosing, extraction process can either be made more abundant, improve extraction rate, shorten extraction time, and Extraction solvent can be saved
Usage amount.Ethyl alcohol and ionic liquid are mixed during the extraction process and are used as extracting solution, volume fraction of ethanol 70%-85%,
In contain moisture, water when extracting under ultrasound condition to filter residue, can be substantially improved as the diluent for reducing system viscosity
Extraction efficiency, and extraction process is environmentally protective, not can cause environmental pollution, and harm will not be brought to operator.
After gained filter residue effectively extract after using extracting solution to enzymatic hydrolysis, then extracted with organic solvent, by purpose at
Divide and be extracted into organic solvent, organic solvent is preferably chloroform, methylene chloride, 1,2- dichloroethanes, carbon tetrachloride, acetic acid
At least one of ethyl ester and acetone, can further remove the impurity being dissolved in extracting solution in this way, so improve purpose at
Point purity, while can also enough silicagel columns be effectively separated.It is pressed when by the silica gel of medicinal extract and 80-100 mesh obtained by step (4)
Weight ratio is that after 1:30-60 is sufficiently mixed, medicinal extract can be dispersed in silica gel, first eluted with petroleum ether, then use petroleum
The mixed solvent of ether and ethyl acetate is eluted, and in the case where controlling certain elution rate, can be effectively to purpose
Ingredient is eluted, and can obtain that purity is higher and the higher purpose ingredient of yield.
After the ethanol solution of substance heat after to elution recrystallizes, purpose ingredient can be further improved
Purity, while agents useful for same is at low cost, and recrystallization effect is good.
Specific embodiment
Embodiment 1
A kind of extracting method of Celastrol, this method are extracted using cymose buckwheat rhizome, bitter buckwheat or sweet tea buckwheat as raw material,
Specific extraction step the following steps are included:
(1) cymose buckwheat rhizome, the seed of bitter buckwheat or sweet tea buckwheat, leaf, stalk and root is dry, liquid nitrogen is added and is ground, powder is obtained；
(2) powder is mixed with deionized water, and adjusting solution ph is 5.0, and the mixed of 0.5% powder weight is then added
Synthase is digested, and hydrolysis temperature is 25 DEG C, enzymolysis time 40min, is filtered after the completion of enzymatic hydrolysis, is obtained filter residue；Wherein, mixed enzyme
It is by weight the mixture of 2:1 mixing for cellulase and pectase；
It (3) is that 1g:20mL is mixed by solid-to-liquid ratio with extracting solution by filter residue obtained by step (2), then in ultrasonic power density
For 100W/cm2, supersonic frequency carry out ultrasonic extraction under the conditions of being 30kHz, Extracting temperature is 45 DEG C, extraction time 40min,
It is separated by solid-liquid separation to obtain supernatant；Wherein, extracting solution is ethyl alcohol that volume fraction is 70% and ionic liquid is 3:1 mixing by volume
Mixture, ionic liquid be [BMIM] Br；
(4) by volume it is that 1:3 is mixed with organic solvent by supernatant obtained by step (3), extracts 3 times, separate organic phase,
Then organic phase is concentrated under reduced pressure under the conditions of 40 DEG C, obtains medicinal extract；Wherein, organic solvent is chloroform；
(5) by weight it is that 1:30 is mixed with the silica gel of 100 mesh by medicinal extract obtained by step (4), is then charged into chromatographic column,
It is first eluted with petroleum ether, then is eluted with the mix reagent that petroleum ether and ethyl acetate are 6:1 mixing by volume, control is washed
The flow velocity of de- liquid is 2mL/min, collects eluent, then eluent is dried under the conditions of 45 DEG C；
(6) by step (5) gains with 30 DEG C, the ethanol solution that volume fraction is 70% is dissolved into saturated solution, then
Cold water is added until crystal is no longer precipitated, crystal is dry, Celastrol is made.
It is extracted by raw material of the seed of cymose buckwheat rhizome, leaf, stalk and root, yield 3.58%, purity 99.3%；
It is extracted by raw material of the seed of bitter buckwheat, leaf, stalk and root, yield 3.31%, purity 99.0%；
It is extracted by raw material of the seed of sweet tea buckwheat, leaf, stalk and root, yield 2.85%, purity 98.8%.
Above-mentioned yield=(obtained Celastrol crystal quality/plant material quality) × 100%.
Embodiment 2
A kind of extracting method of Celastrol, this method are extracted using cymose buckwheat rhizome, bitter buckwheat or sweet tea buckwheat as raw material,
Specific extraction step the following steps are included:
(1) cymose buckwheat rhizome, the seed of bitter buckwheat or sweet tea buckwheat, leaf, stalk and root is dry, liquid nitrogen is added and is ground, powder is obtained；
(2) powder is mixed with deionized water, and adjusting solution ph is 6.0, and the mixed of 0.8% powder weight is then added
Synthase is digested, and hydrolysis temperature is 50 DEG C, enzymolysis time 30min, is filtered after the completion of enzymatic hydrolysis, is obtained filter residue；Wherein, mixed enzyme
It is by weight the mixture of 5:1 mixing for cellulase and pectase；
It (3) is that 1g:80mL is mixed by solid-to-liquid ratio with extracting solution by filter residue obtained by step (2), then in ultrasonic power density
For 150W/cm2, supersonic frequency carry out ultrasonic extraction under the conditions of being 50kHz, Extracting temperature is 70 DEG C, extraction time 20min,
It is separated by solid-liquid separation to obtain supernatant；Wherein, extracting solution is ethyl alcohol that volume fraction is 85% and ionic liquid is 3:2 mixing by volume
Mixture, ionic liquid be [BMIM] Cl；
(4) by volume it is that 1:8 is mixed with organic solvent by supernatant obtained by step (3), extracts 4 times, separate organic phase,
Then organic phase is concentrated under reduced pressure under the conditions of 50 DEG C, obtains medicinal extract；Wherein, organic solvent is methylene chloride；
(5) by weight it is that 1:60 is mixed with the silica gel of 100 mesh by medicinal extract obtained by step (4), is then charged into chromatographic column,
It is first eluted with petroleum ether, then is eluted with the mix reagent that petroleum ether and ethyl acetate are 10:1 mixing by volume, controlled
The flow velocity of eluent is 5mL/min, collects eluent, then eluent is dried under the conditions of 60 DEG C；
(6) by step (5) gains with 40 DEG C, the ethanol solution that volume fraction is 70% is dissolved into saturated solution, then
Cold water is added until crystal is no longer precipitated, crystal is dry, Celastrol is made.
It is extracted by raw material of the seed of cymose buckwheat rhizome, leaf, stalk and root, yield 3.67%, purity 99.4%；
It is extracted by raw material of the seed of bitter buckwheat, leaf, stalk and root, yield 3.39%, purity 99.0%；
It is extracted by raw material of the seed of sweet tea buckwheat, leaf, stalk and root, yield 3.02%, purity 99.1%.
Above-mentioned yield=(obtained Celastrol crystal quality/plant material quality) × 100%.
Embodiment 3
A kind of extracting method of Celastrol, this method are extracted using cymose buckwheat rhizome, bitter buckwheat or sweet tea buckwheat as raw material,
Specific extraction step the following steps are included:
(1) cymose buckwheat rhizome, the seed of bitter buckwheat or sweet tea buckwheat, leaf, stalk and root is dry, liquid nitrogen is added and is ground, powder is obtained；
(2) powder is mixed with deionized water, and adjusting solution ph is 6.0, and the mixed of 0.6% powder weight is then added
Synthase is digested, and hydrolysis temperature is 35 DEG C, enzymolysis time 40min, is filtered after the completion of enzymatic hydrolysis, is obtained filter residue；Wherein, mixed enzyme
It is by weight the mixture of 3:1 mixing for cellulase and pectase；
It (3) is that 1g:30mL is mixed by solid-to-liquid ratio with extracting solution by filter residue obtained by step (2), then in ultrasonic power density
For 120W/cm2, supersonic frequency carry out ultrasonic extraction under the conditions of being 35kHz, Extracting temperature is 55 DEG C, extraction time 30min,
It is separated by solid-liquid separation to obtain supernatant；Wherein, extracting solution is ethyl alcohol that volume fraction is 75% and ionic liquid is 6:1 mixing by volume
Mixture, ionic liquid be [BMIM] BF4；
(4) by volume it is that 1:4 is mixed with organic solvent by supernatant obtained by step (3), extracts 4 times, separate organic phase,
Then organic phase is concentrated under reduced pressure under the conditions of 50 DEG C, obtains medicinal extract；Wherein, organic solvent 1,2- dichloroethanes；
(5) by weight it is that 1:40 is mixed with the silica gel of 100 mesh by medicinal extract obtained by step (4), is then charged into chromatographic column,
It is first eluted with petroleum ether, then is eluted with the mix reagent that petroleum ether and ethyl acetate are 7:1 mixing by volume, control is washed
The flow velocity of de- liquid is 3mL/min, collects eluent, then eluent is dried under the conditions of 50 DEG C；
(6) by step (5) gains with 35 DEG C, the ethanol solution that volume fraction is 75% is dissolved into saturated solution, then
Cold water is added until crystal is no longer precipitated, crystal is dry, Celastrol is made.
It is extracted by raw material of the seed of cymose buckwheat rhizome, leaf, stalk and root, yield 3.82%, purity 99.3%；
It is extracted by raw material of the seed of bitter buckwheat, leaf, stalk and root, yield 3.45%, purity 98.9%；
It is extracted by raw material of the seed of sweet tea buckwheat, leaf, stalk and root, yield 3.12%, purity 99.1%.
Above-mentioned yield=(obtained Celastrol crystal quality/plant material quality) × 100%.
Embodiment 4
A kind of extracting method of Celastrol, this method are extracted using cymose buckwheat rhizome, bitter buckwheat or sweet tea buckwheat as raw material,
Specific extraction step the following steps are included:
(1) cymose buckwheat rhizome, the seed of bitter buckwheat or sweet tea buckwheat, leaf, stalk and root is dry, liquid nitrogen is added and is ground, powder is obtained；
(2) powder is mixed with deionized water, and adjusting solution ph is 6.0, and the mixed of 0.8% powder weight is then added
Synthase is digested, and hydrolysis temperature is 45 DEG C, enzymolysis time 35min, is filtered after the completion of enzymatic hydrolysis, is obtained filter residue；Wherein, mixed enzyme
It is by weight the mixture of 4:1 mixing for cellulase and pectase；
It (3) is that 1g:60mL is mixed by solid-to-liquid ratio with extracting solution by filter residue obtained by step (2), then in ultrasonic power density
For 140W/cm2, supersonic frequency carry out ultrasonic extraction under the conditions of being 45kHz, Extracting temperature is 65 DEG C, extraction time 30min,
It is separated by solid-liquid separation to obtain supernatant；Wherein, extracting solution is ethyl alcohol that volume fraction is 80% and ionic liquid is 5:1 mixing by volume
Mixture, ionic liquid be [BMIM] PF6；
(4) by volume it is that 1:6 is mixed with organic solvent by supernatant obtained by step (3), extracts 4 times, separate organic phase,
Then organic phase is concentrated under reduced pressure under the conditions of 50 DEG C, obtains medicinal extract；Wherein, organic solvent is acetone；
(5) by weight it is that 1:50 is mixed with the silica gel of 100 mesh by medicinal extract obtained by step (4), is then charged into chromatographic column,
It is first eluted with petroleum ether, then is eluted with the mix reagent that petroleum ether and ethyl acetate are 9:1 mixing by volume, control is washed
The flow velocity of de- liquid is 4mL/min, collects eluent, then eluent is dried under the conditions of 50 DEG C；
(6) by step (5) gains with 30 DEG C, the ethanol solution that volume fraction is 80% is dissolved into saturated solution, then
Cold water is added until crystal is no longer precipitated, crystal is dry, Celastrol is made.
It is extracted by raw material of the seed of cymose buckwheat rhizome, leaf, stalk and root, yield 3.90%, purity 99.6%；
It is extracted by raw material of the seed of bitter buckwheat, leaf, stalk and root, yield 3.50%, purity 99.2%；
It is extracted by raw material of the seed of sweet tea buckwheat, leaf, stalk and root, yield 3.31%, purity 99.5%.
Above-mentioned yield=(obtained Celastrol crystal quality/plant material quality) × 100%.
Embodiment 5
A kind of extracting method of Celastrol, this method are extracted using cymose buckwheat rhizome, bitter buckwheat or sweet tea buckwheat as raw material,
Specific extraction step the following steps are included:
(1) cymose buckwheat rhizome, the seed of bitter buckwheat or sweet tea buckwheat, leaf, stalk and root is dry, liquid nitrogen is added and is ground, powder is obtained；
(2) powder is mixed with deionized water, and adjusting solution ph is 5.5, and the mixed of 0.6% powder weight is then added
Synthase is digested, and hydrolysis temperature is 45 DEG C, enzymolysis time 30min, is filtered after the completion of enzymatic hydrolysis, is obtained filter residue；Wherein, mixed enzyme
It is by weight the mixture of 4:1 mixing for cellulase and pectase；
It (3) is that 1g:50mL is mixed by solid-to-liquid ratio with extracting solution by filter residue obtained by step (2), then in ultrasonic power density
For 120W/cm2, supersonic frequency carry out ultrasonic extraction under the conditions of being 35kHz, Extracting temperature is 60 DEG C, extraction time 30min,
It is separated by solid-liquid separation to obtain supernatant；Wherein, extracting solution is ethyl alcohol that volume fraction is 75% and ionic liquid is 5:2 mixing by volume
Mixture, ionic liquid be [BMIM] Br；
(4) by volume it is that 1:6 is mixed with organic solvent by supernatant obtained by step (3), extracts 4 times, separate organic phase,
Then organic phase is concentrated under reduced pressure under the conditions of 50 DEG C, obtains medicinal extract；Wherein, organic solvent is methylene chloride；
(5) by weight it is that 1:35 is mixed with the silica gel of 100 mesh by medicinal extract obtained by step (4), is then charged into chromatographic column,
It is first eluted with petroleum ether, then is eluted with the mix reagent that petroleum ether and ethyl acetate are 8:1 mixing by volume, control is washed
The flow velocity of de- liquid is 5mL/min, collects eluent, then eluent is dried under the conditions of 50 DEG C；
(6) by step (5) gains with 35 DEG C, the ethanol solution that volume fraction is 75% is dissolved into saturated solution, then
Cold water is added until crystal is no longer precipitated, crystal is dry, Celastrol is made.
It is extracted by raw material of the seed of cymose buckwheat rhizome, leaf, stalk and root, yield 3.98%, purity 99.8%；
It is extracted by raw material of the seed of bitter buckwheat, leaf, stalk and root, yield 3.61%, purity 99.4%；
It is extracted by raw material of the seed of sweet tea buckwheat, leaf, stalk and root, yield 3.39%, purity 99.1%.
Above-mentioned yield=(obtained Celastrol crystal quality/plant material quality) × 100%.
Comparative example 1
Documents 1 using the seed of cymose buckwheat rhizome, leaf, stalk and root as raw material extract, extracting method compared with Example 5,
Liquid nitrogen grinding is not had in step (1), but conventional crushing, remaining step are same as Example 5.
Gained yield is 3.49%, purity 99.2%；
Yield=(obtained Celastrol crystal quality/plant material quality) × 100%.
Comparative example 2
Documents 2 using the seed of cymose buckwheat rhizome, leaf, stalk and root as raw material extract, extracting method compared with Example 5,
Lack step (2), directly reacts powder obtained by step (1) with extracting solution, remaining step is same as Example 5.
Gained yield is 3.39%, purity 99.3%；
Yield=(obtained Celastrol crystal quality/plant material quality) × 100%.
Comparative example 3
Documents 3 using the seed of cymose buckwheat rhizome, leaf, stalk and root as raw material extract, extracting method compared with Example 5,
Step (3) is only to mix gained filter residue with extracting solution to extract, and Extracting temperature is 60 DEG C, extraction time 30min,
Remaining step is same as Example 5.
Gained yield is 3.45%, purity 98.5%；
Yield=(obtained Celastrol crystal quality/plant material quality) × 100%.
Comparative example 4
Documents 4 using the seed of cymose buckwheat rhizome, leaf, stalk and root as raw material extract, extracting method compared with Example 5,
The extracting solution of step (3) is only the ethyl alcohol that volume fraction is 75%, remaining step is same as Example 5.
Gained yield is 3.49%, purity 99.2%；
Yield=(obtained Celastrol crystal quality/plant material quality) × 100%.
Comparative example 5
Documents 5 using the seed of cymose buckwheat rhizome, leaf, stalk and root as raw material extract, extracting method compared with Example 5,
Gained filter residue is mixed with extracting solution in step (3) and is extracted, Extracting temperature is 60 DEG C, extraction time 30min, and is extracted
Liquid is only the ethyl alcohol that volume fraction is 75%, remaining step is same as Example 5.
Gained yield is 3.33%, purity 98.4%；
Yield=(obtained Celastrol crystal quality/plant material quality) × 100%.
Comparative example 6
Documents 6 using the seed of cymose buckwheat rhizome, leaf, stalk and root as raw material extract, extracting method compared with Example 5,
Step (1) does not have to liquid nitrogen grinding, but conventional crushing；Lack step (2), directly by powder obtained by step (1) and extracting solution
It being reacted, and extracting solution is only the ethyl alcohol that volume fraction is 75%, Extracting temperature is 60 DEG C, extraction time 30min, remaining
Step is same as Example 5.
Gained yield is 3.20%, purity 97.6%；
Yield=(obtained Celastrol crystal quality/plant material quality) × 100%.
Comparative example 7
Documents 7 using the seed of cymose buckwheat rhizome, leaf, stalk and root as raw material extract, extracting method compared with Example 5,
Step (1)-step (4) is same as Example 5, and later step is different, specifically:
Medicinal extract obtained by step (4) is added in petroleum ether step (5), is heated to reflux 30min, and petroleum is removed in filtering while hot
Ether is repeated twice, and obtains residue, and the dry petroleum ether that volatilizees；Wherein, the ratio of medicinal extract and petroleum ether obtained by step (4) is 1g:
15mL；
(6) step (5) gains are added in 75% ethyl alcohol, are heated to 60 DEG C of dissolutions, are slow added into same volume
Then water steams most of ethyl alcohol, be placed in cooling in ice-water bath, obtain Celastrol crude extract；
(7) a small amount of ethyl alcohol of Celastrol crude extract is dissolved, is filled by pretreated AB-8 macroporous absorbent resin 20g
Column simultaneously carries out post separation, with 70% ethanol elution, collects eluent, dry, then presses volume with hexamethylene and dioxane again
Mixture than mixing for 9:1 is recrystallized, and Celastrol is made.
Gained yield is 3.33%, purity 99.2%；
Yield=(obtained Celastrol crystal quality/plant material quality) × 100%.
Claims (10)
1. a kind of extracting method of Celastrol, which is characterized in that extracted from Fagopyrum plant.
2. the extracting method of Celastrol according to claim 1, which is characterized in that Fagopyrum plant be cymose buckwheat rhizome,
Bitter buckwheat or sweet tea buckwheat.
3. the extracting method of Celastrol according to claim 1 or 2, which comprises the following steps:
(1) Fagopyrum plant seed, leaf, stalk and/or root is dry, liquid nitrogen is added and is ground, powder is obtained；
(2) powder is mixed with deionized water, and adjusting solution ph is 5.0-6.0, and 0.5-0.8% powder weight is then added
Mixed enzyme digested, hydrolysis temperature be 25-50 DEG C, enzymolysis time 30-40min, enzymatic hydrolysis after the completion of filter, obtain filter residue；
Wherein, mixed enzyme is the mixture that cellulase and pectase are 2-5:1 mixing by weight；
(3) it is that 1g:20-80mL is mixed by solid-to-liquid ratio with extracting solution by filter residue obtained by step (2), is then in ultrasonic power density
100-150W/cm2, supersonic frequency carry out ultrasonic extraction under the conditions of being 30-50kHz, Extracting temperature is 45-70 DEG C, extraction time
For 20-40min, it is separated by solid-liquid separation to obtain supernatant；
(4) by volume it is that 1:3-8 is mixed with organic solvent by supernatant obtained by step (3), extracts 3-4 times, separate organic phase,
Then organic phase is concentrated under reduced pressure under the conditions of 40-50 DEG C, obtains medicinal extract；
(5) by weight it is that 1:30-60 is mixed with the silica gel of 80-100 mesh by medicinal extract obtained by step (4), is then charged into chromatographic column
In, it is first eluted with petroleum ether, then eluted with the mix reagent that petroleum ether and ethyl acetate are 6-10:1 mixing by volume,
The flow velocity for controlling eluent is 2-5mL/min, collects eluent, then eluent is dried under the conditions of 45-60 DEG C；
(6) by step (5) gains with 30-40 DEG C, volume fraction is that the ethanol solution of 70-80% is dissolved into saturated solution, so
Cold water is added afterwards until crystal is no longer precipitated, crystal is dry, Celastrol is made.
4. the extracting method of Celastrol as claimed in claim 3, which is characterized in that detailed process in step (2) are as follows: by powder
End is mixed with deionized water, and adjusting solution ph is 5.5, and the mixed enzyme that 0.6% powder weight is then added is digested, enzyme
Solving temperature is 45 DEG C, enzymolysis time 30min, filters after the completion of enzymatic hydrolysis, obtains filter residue.
5. the extracting method of Celastrol as claimed in claim 4, which is characterized in that mixed enzyme is cellulose in step (2)
Enzyme and pectase are the mixture of 4:1 mixing by weight.
6. the extracting method of Celastrol as claimed in claim 3, which is characterized in that detailed process in step (3) are as follows: will walk
Suddenly filter residue obtained by (2) is that 1g:50mL is mixed by solid-to-liquid ratio with extracting solution, is then 120W/cm in ultrasonic power density2, ultrasound
Frequency carries out ultrasonic extraction under the conditions of being 35kHz, Extracting temperature is 60 DEG C, and extraction time 30min is separated by solid-liquid separation to obtain supernatant
Liquid.
7. the extracting method of Celastrol as claimed in claim 6, which is characterized in that extracting solution is volume point in step (3)
The ethyl alcohol and ionic liquid that number is 70%-85% are the mixture of 3-6:1-2 mixing by volume.
8. the extracting method of Celastrol as claimed in claim 7, which is characterized in that ionic liquid be [BMIM] Br,
At least one of [BMIM] Cl, [BMIM] BF4 and [BMIM] PF6.
9. the extracting method of Celastrol as claimed in claim 3, which is characterized in that organic solvent is trichlorine in step (4)
At least one of methane, methylene chloride, 1,2- dichloroethanes, carbon tetrachloride, ethyl acetate and acetone.
10. the extracting method of Celastrol as claimed in claim 3, which is characterized in that will be obtained by step (4) in step (5)
Medicinal extract is that 1:35 is mixed with the silica gel of 100 mesh by weight；The elution of Shi Xianyong petroleum ether is eluted, then with petroleum ether and ethyl acetate
The mix reagent for 8:1 mixing is eluted by volume, and the flow velocity for controlling eluent is 5mL/min.
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