CN103059094B - A kind of method extracting Tripterine - Google Patents
A kind of method extracting Tripterine Download PDFInfo
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- CN103059094B CN103059094B CN201310027683.5A CN201310027683A CN103059094B CN 103059094 B CN103059094 B CN 103059094B CN 201310027683 A CN201310027683 A CN 201310027683A CN 103059094 B CN103059094 B CN 103059094B
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Abstract
The invention discloses a kind of method extracting Tripterine, wherein, the method comprises and being contacted with the first lower aliphatic alcohols by the plant material containing Tripterine, makes the content obtaining Tripterine be not less than the alcohol extract of 0.005 % by weight; Again described alcohol extract macroporous adsorptive resins is separated, and the elutriant collected containing Tripterine, the aqueous solution of described second lower aliphatic alcohols that the eluent adopted in sepn process is the second lower aliphatic alcohols, concentration is not less than 90 volume % or concentration are not less than the aqueous acetone solution of 60 volume %; Again by concentrated for the described elutriant containing Tripterine also crystallization.Adopt extracting method of the present invention, purity and the higher Tripterine of yield can not only be obtained, but also simplify abstraction and purification process, have prospects for commercial application.
Description
Technical field
The present invention relates to a kind of method extracting Tripterine.
Background technology
Tripterine, has another name called celastrin, Tripterin, celastrine, and English name is celastrol or tripterine, and be a kind of quinone methyl pentacyclic triterpenoid, its structure is as shown in formula I.Tripterine is extensively present in the root skin zone position of Celastraceae plant Thunder God Calamus and celastraceae plants.Tripterine molecular formula is C
29h
38o
4, red needle-like crystal, it is insoluble in water, and be dissolved in the organic solvents such as methyl alcohol, ethanol, acetone, ethyl acetate, chloroform, fusing point is 185-200 DEG C, and ultraviolet full wavelength scanner finds that it has the strongest absorption at 425nm place.
Domestic and international research finds, Tripterine has multiple drug effect, as anti-oxidant, anti inflammatory immunity, antitumor, antifertility, anti-fibrosis, atherosclerosis, inhibiting angiogenesis, cell death inducing etc.Its distinctive Composition and function attracts people's attention, so that cause the global research boom to Tripterine production technology and application and development, especially the new drug research of Tripterine and derivative thereof achieves breakthrough, indicate that it has certain market outlook, the industrial Production requirement to it also increases day by day.
According to the solvability of Tripterine, utilizing organic solvent to extract Tripterine from the higher plant of trypterygine cellulose content, is produce a kind of method that Tripterine generally takes.So far, the extraction process of bibliographical information mainly organic solvent extraction and supercritical CO
2extraction, and the most frequently used still organic solvent extraction.
CN101311186A discloses a kind of method preparing Tripterine, and the method comprises Celastraceae plant powder chloroform dry weight being less than 6 % by weight and carries out supersound extraction and concentrated, obtains chloroform extract; Sherwood oil is added in above-mentioned chloroform extract, sherwood oil is taken out after reflux, and product is added in silicagel column, then carry out wash-out with chloroform, chloroform containing 2-4 weight acetone %, sherwood oil, the sherwood oil containing 4-6 weight ethyl acetate % and the sherwood oil containing 8-12 weight ethyl acetate % successively, then the trypterygine paste sherwood oil obtained and ethyl acetate are carried out recrystallization.
CN101830960A discloses a kind of preparation method of celastrin, the method comprises the following steps: get Celastraceae plant root of Panicled Bittersweet root skin, pulverize, add the methyl alcohol of its quality 4-5 times amount volume, to insert in ultrasonic extraction device supersound extraction 1-3 time, each 0.2-1 hour, extraction power is 30-60KHz, united extraction liquid, filter, reclaim under reduced pressure methyl alcohol, the alkali aqueous solution that residue adds 0.05-0.15mol/L dissolves, filter, filtrate by dilute hydrochloric acid acid solution adjust ph to 3-5, leave standstill, filter, get filter residue, join in alumina chromatographic column, with the methanol-chloroform mixed solvent wash-out that volume ratio is 3:1, collect the elutriant of 2-4 times amount column volume, decompression and solvent recovery is also concentrated, add ethanol-normal hexane mixed solvent crystallization that volume ratio is 1:1.
CN101638425A discloses a kind of method extracting Tripterine in Root of Oriental Bittersweet skin, the method comprises the following steps: with Root of Oriental Bittersweet skin powder for raw material, comprise extraction, be separated, purifying and recrystallization, described extraction first with rudimentary chloroparaffin for solvent, heating and refluxing extraction, the mass ratio of root skin powder and solvent is 1:3-20, time is no less than 2 hours, medicinal extract shape primary extract is obtained after extracting solution desolventizing, then in primary extract, 50-70 volume % methyl alcohol or ethanolic soln is added, abundant stirring, leave standstill extraction, paste primary extract is obtained after extraction liquid desolventizing, described purifying is again with the column chromatography separating purification that stationary phase is purification on normal-phase silica gel or reverse phase silica gel after primary extract dissolve with methanol, collects elutriant, obtains Powdered extract, finally use ethanol-water mixed solvent recrystallization after desolventizing drying.
CN102349917A discloses a kind of purposes and preparation method of Tripterine, and specifically disclose this preparation method and comprise the following steps: (1) is by after trypterygine on the ground liana part and the process of subterraneous root part co-grinding, through alcohol reflux, concentrating under reduced pressure after filtering; (2) by the ultrasonic suspendible of ethanol extraction ethanol water after concentrated, after leaving standstill, supernatant liquor is poured out; (3) supernatant liquor adopts polymeric adsorbent to adsorb, then carries out wash-out with water, 40 volume % aqueous ethanolic solutions, 70 volume % aqueous ethanolic solutions and 90-95 volume % aqueous ethanolic solution successively, collects elutriant; (4) after elution fraction concentrating under reduced pressure, through silica gel chromatography, dry acquisition red powder extract after recycling design; (5), after this extract is dissolved into saturated solution with hot ethanol, place, dry after separating out red colored crystalline.
Because the chemical structure of Tripterine is complicated, be difficult to realize low cost synthesis and produce, and from content relatively high plant extraction and isolation, its feasibility is higher.But the extraction of Tripterine preparation is also only limitted to laboratory on a small scale at present, the Technology in past is often amplified because yield is low, cost is high, preparation amount is little or be difficult to realize technique because process is complicated, thus causes that plant resources utilization rate is very low, Tripterine price is very expensive in the market.
Summary of the invention
The object of the invention is to extract Tripterine purity and the lower defect of yield to overcome existing method, and a kind of method of extraction Tripterine is newly provided.
The invention provides a kind of method extracting Tripterine, wherein, the method comprises and being contacted with the first lower aliphatic alcohols by the plant material containing Tripterine, makes the content obtaining Tripterine be not less than the alcohol extract of 0.005 % by weight; Again described alcohol extract macroporous adsorptive resins is separated, and the elutriant collected containing Tripterine, the aqueous solution of described second lower aliphatic alcohols that the eluent adopted in sepn process is the second lower aliphatic alcohols, concentration is not less than 90 volume % or concentration are not less than the aqueous acetone solution of 60 volume %; Again by concentrated for the described elutriant containing Tripterine also crystallization.
The present inventor finds, prior art adopts macroporous adsorptive resins to be separated the extracting solution containing Tripterine, and needing to adopt water, 40 volume % ethanolic solns, 70 volume % ethanolic solns and 90-95 volume % ethanolic soln to carry out gradient elution successively could separate Tripterine and all the other impurity.And the aqueous solution of described second lower aliphatic alcohols that the present invention directly adopts the second lower aliphatic alcohols, concentration is not less than 90 volume % or concentration are not less than the aqueous acetone solution of 60 volume % as the eluent in macroporous adsorptive resins sepn process, equally also can reach good separating effect, simplify sepn process.In addition, the present invention do not need macroporous adsorptive resins to be separated obtain carry out silica gel column chromatography separation containing the elutriant of Tripterine, and directly described elutriant to be concentrated and crystallization just can obtain the higher Tripterine of purity.
Other features and advantages of the present invention are described in detail in embodiment part subsequently.
Embodiment
Below the specific embodiment of the present invention is described in detail.Should be understood that, embodiment described herein, only for instruction and explanation of the present invention, is not limited to the present invention.
The method of extraction Tripterine provided by the invention comprises and being contacted with the first lower aliphatic alcohols by the plant material containing Tripterine, makes the content obtaining Tripterine be not less than the alcohol extract of 0.005 % by weight; Again described alcohol extract macroporous adsorptive resins is separated, and the elutriant collected containing Tripterine, the aqueous solution of described second lower aliphatic alcohols that the eluent adopted in sepn process is the second lower aliphatic alcohols, concentration is not less than 90 volume % or concentration are not less than the aqueous acetone solution of 60 volume %; Again by concentrated for the described elutriant containing Tripterine also crystallization.
According to the present invention, the kind of the plant material containing Tripterine is known to the skilled person, and such as, can be root and/or the root skin of Celastraceae celastraceae plants and/or tripterygium plant.Particularly, described Celastraceae celastraceae plants such as can be selected from Celastrus hookeri, bastard indigo Stem of Oriental Bittersweet, powder back of the body Stem of Oriental Bittersweet, celastrus angulatus, Stem of Oriental Bittersweet, nerville show in post Stem of Oriental Bittersweet, Celastrus aceleatus Merr, haw rattan, short stalk Stem of Oriental Bittersweet, wide leaf short stalk Stem of Oriental Bittersweet, Du Ziteng, blue or green river rattan and large bud Stem of Oriental Bittersweet one or more.Described tripterygium plant can be such as trypterygine.
Wherein, high performance liquid chromatography (HPLC method) is adopted to measure the content of Tripterine in the dry root skin of above-mentioned 13 kinds of Celastraceae celastraceae plantses and a kind of tripterygium plant, wherein, test condition is: TSK-GEL ODS-100Z C18 chromatographic column (250mm × 4.6mm × 5 μm), flow velocity is 1.0mL/min, determined wavelength is 425nm, the mixed solvent (volume ratio of methyl alcohol and aqueous acetic acid is 87:13) of moving phase to be methyl alcohol and concentration be 1 volume % aqueous acetic acid, records result as follows: Celastrus hookeri: 3.81 % by weight; Bastard indigo Stem of Oriental Bittersweet: 1.10 % by weight; Powder back of the body Stem of Oriental Bittersweet: 1.02 % by weight; Celastrus angulatus: 0.67 % by weight; Stem of Oriental Bittersweet: 0.62 % by weight; Nerville shows post Stem of Oriental Bittersweet: 0.90 % by weight; Celastrus aceleatus Merr: 0.84 % by weight; Haw rattan: 1.73 % by weight; Short stalk Stem of Oriental Bittersweet: 0.55 % by weight; Wide leaf short stalk Stem of Oriental Bittersweet: 0.49 % by weight; Only son rattan: 0.77 % by weight; Blue or green river rattan: 0.03 % by weight; Large bud Stem of Oriental Bittersweet: 0.39 % by weight; Trypterygine: 0.62 % by weight.
As can be seen here, all containing Tripterine in 13 kinds of above-mentioned analyzed celastraceae plantses and the root skin of a kind of tripterygium plant, except the rattan of blue or green river, all may become the raw material extracting Tripterine.That is, described Celastraceae celastraceae plants is preferably Celastrus hookeri, bastard indigo Stem of Oriental Bittersweet, powder back of the body Stem of Oriental Bittersweet, celastrus angulatus, Stem of Oriental Bittersweet, nerville show in post Stem of Oriental Bittersweet, Celastrus aceleatus Merr, haw rattan, short stalk Stem of Oriental Bittersweet, wide leaf short stalk Stem of Oriental Bittersweet, only son rattan and large bud Stem of Oriental Bittersweet one or more.Described tripterygium plant is preferably trypterygine.
According to the present invention, the described plant material containing Tripterine can exist with the form of bulk, also can exist with pulverous form, but in order to more effectively extract Tripterine wherein, under preferable case, the described plant material containing Tripterine exists with pulverous form.
According to the present invention, from the angle being convenient to describe, the lower aliphatic alcohols contacted is called " the first lower aliphatic alcohols ", the lower aliphatic alcohols as eluent is called " the second lower aliphatic alcohols " with the plant material containing Tripterine.According to the present invention, described first lower aliphatic alcohols and the second lower aliphatic alcohols identical or different, and the fatty alcohol that can be less than 8 for existing various carbonatoms, is preferably C
1-C
5monohydroxy-alcohol.Described C
1-C
5the specific examples of monohydroxy-alcohol can include but not limited in methyl alcohol, ethanol, n-propyl alcohol, Virahol, propyl carbinol, isopropylcarbinol, the trimethyl carbinol, 1-amylalcohol, 2-amylalcohol and 3-amylalcohol one or more, be particularly preferably methyl alcohol and/or ethanol.Described first lower aliphatic alcohols can use with pure state, also can use with the form of its aqueous solution, as long as the plant material containing Tripterine can be extracted, preferably uses with the form of its aqueous solution.When described first lower aliphatic alcohols uses in form of an aqueous solutions, its concentration reasonably can be selected according to practical situation, such as, can be 25-75 volume %.
The consumption of the present invention to described first lower aliphatic alcohols is not particularly limited, reasonably can select according to the consumption of the plant material containing Tripterine, such as, with the plant material containing Tripterine of 100 weight parts for benchmark, the consumption of described first lower aliphatic alcohols can be 2-20 weight part.It should be noted that, when described first lower aliphatic alcohols uses in form of an aqueous solutions, the consumption of the first lower aliphatic alcohols described herein does not comprise the content of water.
According to the present invention, be that Tripterine is dissolved in the first fatty alcohol by the order that the plant material containing Tripterine contacts with the first lower aliphatic alcohols.The mode of the present invention to described contact is not particularly limited, and can adopt and well known to a person skilled in the art that various mode is carried out.Such as, at normal temperatures the plant material containing Tripterine can be immersed in (namely normal temperature leaves standstill extraction) in the first lower aliphatic alcohols, also can be under agitation, plant material containing Tripterine is immersed in (stirring at normal temperature extraction) in the first lower aliphatic alcohols at normal temperatures, also the first lower aliphatic alcohols can be allowed at normal temperatures to continue through plant material (normal temperature percolation method) containing Tripterine, or also under microwave condition, the plant material containing Tripterine can be immersed in (microwave loss mechanisms) in the first lower aliphatic alcohols, or also under Ultrasonic Conditions, the plant material containing Tripterine can be immersed in (ultrasonic extraction) in the first lower aliphatic alcohols, under reflux conditions the plant material containing Tripterine can also be immersed in (water bath reflux method) in the first lower aliphatic alcohols.Can know these those skilled in the art, will repeat no more at this.
In the present invention, described normal temperature all refers to 25 DEG C.
The present invention is not particularly limited the condition contacted with the first lower aliphatic alcohols by the plant material containing Tripterine, as long as at least part of Tripterine can be made to be dissolved in the first lower aliphatic alcohols, the content obtaining Tripterine is made to be not less than the alcohol extract of 0.005 % by weight, such as, the condition of described the contact temperature generally including contact and the time contacted.The temperature of described contact can carry out selecting and changing in wider scope, but in order to more be conducive to the extraction of Tripterine, the temperature of described contact is preferably 25 DEG C of boiling points to the first lower aliphatic alcohols.Duration of contact extends the further extraction being conducive to Tripterine, and in order to obtain the higher alcohol extract of Tripterine, the time of described contact is not less than 1 hour usually.Particularly, the different ways of contact has larger difference at corresponding duration of contact, and such as, the duration of contact that normal temperature leaves standstill extraction is generally 6-12 hour; The time of stirring at normal temperature extraction is generally 2-4 hour; The duration of contact of microwave―assisted extraction or ultrasonic-assisted extraction method is generally 0.5-1 hour; The duration of contact of water bath reflux method is generally 1-2 hour.
According to a kind of specific implementation method of the present invention, be that methyl alcohol and/or the ethanol of 25-75 volume % extracts by the plant material concentration containing Tripterine, filter and obtain alcohol extract.It should be noted that, if adopt water bath reflux method (Extracting temperature is more than 30 DEG C), then, before above-mentioned alcohol extract macroporous adsorptive resins being separated, in order to avoid damaging macroporous adsorbent resin, needing described alcohol extract to be cooled to normal temperature.Those skilled in the art should it is easily understood that, carry out in the process extracted at employing methyl alcohol and/or ethanol, once extract and may can't be enough to fully extract containing the Tripterine in the plant material of Tripterine, therefore, in order to make full use of raw material, preferably carry out 2-5 time according to the method described above and extract Tripterine, and carry out wash-out with eluent again after the product that each time is extracted is carried out loading absorption with macroporous resin successively.
According to the present invention, the macroporous adsorbent resin of filling in described macroporous adsorptive resins is preferably nonpolar macroporous adsorption resin to intermediate-polarity macroporous adsorption resin.
As well known to those skilled in the art, nonpolar macroporous adsorption resin generally refers to that charge distribution is even, and there is not the resin of the polar group of positive and negative charge Relatively centralized on a molecular scale, major part is vinylbenzene, divinylbenzene polymer.And in intermediate-polarity macroporous adsorption resin, there is the polar group of ester group one class, there is certain polarity, commonly polyacrylate type polymer, using the methacrylic ester of more function group as linking agent.Particularly, described macroporous adsorbent resin particularly preferably the trade mark be that SP825L(is nonpolar), SP207(is nonpolar), ADS-5(is nonpolar), ADS-8(Semi-polarity), D101-1(is nonpolar), AB-8(low-pole), HP10(is nonpolar), LSA-10(low-pole), LSA-20(low-pole), LSA-21(Semi-polarity), LSA-30(is nonpolar), LSA-33(Semi-polarity) and XDA-6(Semi-polarity) in one or more macroporous adsorbent resin.
According to the present invention, as mentioned above, described macroporous adsorbent resin is polymkeric substance (polystyrenic polymer and/or polyacrylic), wherein also may remain unconverted monomer and other additive, therefore, in order to avoid these materials sepn process impacted and improve the purity of separated product, before above-mentioned alcohol extract being separated with the new macroporous adsorptive resins used first, also preferred the macroporous adsorbent resin of filling in this macroporous adsorptive resins is carried out pre-treatment.Described pretreated method can adopt and well known to a person skilled in the art that various method is carried out, such as, can comprise the following steps: first at 25 DEG C, use alcohol immersion 12-24 hour, not muddy to water outlet with ethanol rinse again, then wash with water to without ethanol taste, be then the salt acid soak 2-4 hour of 3-5 % by weight by concentration, be washed to neutrality, be the soaking with sodium hydroxide 2-4 hour of 2-3 % by weight again by concentration, be washed to neutrality.
The present invention is not particularly limited the condition be separated by described alcohol extract macroporous adsorptive resins, usually, described separation condition comprises the flow velocity of the consumption of the macroporous adsorbent resin of filling in temperature, macroporous adsorptive resins, the flow velocity of alcohol extract and eluent.Described temperature can carry out selecting and changing in wider scope, such as, can be 10-40 DEG C.The consumption of described macroporous absorption resin can be selected according to the concentration of Tripterine in alcohol extract and volume.Particularly, the weight of Tripterine can be calculated according to the Tripterine concentration in alcohol extract and volume, the consumption of macroporous adsorbent resin is selected, as long as this consumption can reach the Tripterine in alcohol extract and other magazins' layout by the weight of Tripterine.Such as, with the Tripterine in alcohol extract described in 1g for benchmark, the volume of the macroporous adsorbent resin of filling in described macroporous adsorptive resins can be 80-200mL.The flow velocity of described alcohol extract can be 2-4 times of column volume/hour.The flow velocity of described eluent can be 1.5-3.5 times of column volume/hour.
As well known to those skilled in the art, above-mentioned alcohol extract macroporous adsorptive resins is being carried out in the process be separated, tlc can be adopted to monitor, with judge Tripterine from flow out to time of flowing out and terminating and collect outflow product during this period of time.In addition, because Tripterine is red, therefore, in sepn process, the time that the time that also can first start to flow out according to the rough trypterygine redness of color terminates to outflow, and then adopt tlc to check the product of outflow during this period of time whether to be rich in target product.All can know these those skilled in the art, will repeat no more at this.
As well known to those skilled in the art, described alcohol extract macroporous adsorptive resins is carried out being separated comprise alcohol extract being introduced from one end of macroporous adsorptive resins to contact with described macroporous adsorbent resin and adsorb, effluent liquid after absorption is drawn from the other end of macroporous adsorptive resins, and carry out wash-out with eluent, collect the elutriant containing Artemisinin; The method also comprises the Extraction solvent described effluent liquid circulation being used as the plant material containing Tripterine, can realize the recycle of Extraction solvent like this.
According to the present invention, the described elutriant containing Tripterine is concentrated and refers to that the solvent major part by described elutriant is removed, thus make the solid content of the enriched product obtained be not less than 90 % by weight, all can know these those skilled in the art, will repeat no more at this.
According to the present invention, the method of described crystallization is known to the skilled person, but, in order to obtain more pure Tripterine, under preferable case, the method of described crystallization comprises first carries out crystallization by the mixed solvent of the product sherwood oil obtained after the solvent removing in elutriant and ethyl acetate, then uses C
1-C
5fatty alcohol or concentration be not less than the C of 95 volume %
1-C
5the aqueous solution of fatty alcohol carry out recrystallization.Wherein, in described mixed solvent, the volume ratio of sherwood oil and ethyl acetate is as being 0.2-5:1.Described C
1-C
5the specific examples of fatty alcohol can include but not limited to: one or more in methyl alcohol, ethanol, n-propyl alcohol, Virahol, propyl carbinol, isopropylcarbinol, the trimethyl carbinol, 1-amylalcohol, 2-amylalcohol and 3-amylalcohol, are particularly preferably methyl alcohol and/or ethanol.The number of times of described recrystallization reasonably can be selected according to practical situation, will repeat no more at this.
Below will be described the present invention by embodiment.
In following examples and comparative example, before above-mentioned alcohol extract is separated with the new macroporous adsorptive resins first used, first the macroporous adsorbent resin of filling in macroporous adsorptive resins is carried out following pre-treatment: first at 25 DEG C, use alcohol immersion 12 hours, not muddy to water outlet with ethanol rinse again, then wash with water to without ethanol taste, be then the salt acid soak 3 hours of 3 % by weight by concentration, be washed to neutrality, be the soaking with sodium hydroxide 3 hours of 3 % by weight again by concentration, be washed to neutrality.
In following examples and comparative example, the purity (%) of Tripterine adopts the high performance liquid chromatography purchased from Japanese Shimadzu Corporation to measure; Yield (%)=(weight × purity of the trypterygine cellulose crystal obtained) ÷ of Tripterine (weight of the plant material containing Tripterine × content) containing Tripterine in the plant material of Tripterine × 100%.
Embodiment 1
This embodiment is for illustration of the method for extraction Tripterine provided by the invention.
Get the dry root skin of 510 grams of celastraceae plants powder back of the body Stem of Oriental Bittersweet, carry out normal temperature (25 DEG C) with the methanol aqueous solution that 5 liters of concentration are 70 volume % at every turn and stir extraction 4 hours, filter, extract three times altogether.The concentration extracting Tripterine in the alcohol extract obtained for three times is followed successively by 0.0673 % by weight, 0.0271 % by weight and 0.0092 % by weight.At 25 DEG C, three alcohol extracts are carried out loading absorption with being equipped with (purchased from the green hundred careless development in science and technology company limiteds in Beijing) in the resin column of the pretreated SP825L macroporous adsorbent resin of 700mL successively, the loading flow velocity controlling alcohol extract is 2500mL/h, wash-out is carried out with methyl alcohol after loading reaches terminal, the flow velocity controlling methyl alcohol is 2200mL/h, collects red elutriant and verifies as the elutriant being rich in Tripterine through thin-layer chromatography.This is rich in the elutriant of Tripterine concentrated after, first use mixed solvent (v/v=1:1) crystallization of petroleum ether-ethyl acetate once, then with recrystallizing methanol once, filter and dry red crystals, obtain the trypterygine cellulose crystal that 3.36 grams of purity are 98.11%, yield is 63.5%.
Embodiment 2
This embodiment is for illustration of the method for extraction Tripterine provided by the invention.
Get the dry root skin of 960 grams of celastraceae plants celastrus angulatus, each with the methanol aqueous solution that 6 liters of concentration are 45 volume % carry out refluxing extraction 1.5 hours, filter, extraction four times altogether.The concentration extracting Tripterine in the alcohol extract obtained for four times is followed successively by 0.0581 % by weight, 0.0300 % by weight, 0.0151 % by weight and 0.0056 % by weight.At 25 DEG C, four alcohol extracts are carried out loading absorption with being equipped with (purchased from the new photoinitiator chemical company limited in Hebei) in the resin column of the pretreated ADS-5 macroporous adsorbent resin of 450mL successively, loading flow velocity is 900mL/h, wash-out is carried out with the aqueous acetone solution that concentration is 65 volume % after loading reaches terminal, controlling the flow velocity of aqueous acetone solution in resin column is 700mL/h, collects red elutriant and verifies as the elutriant being rich in Tripterine through thin-layer chromatography.This is rich in the elutriant of Tripterine concentrated after, first use mixed solvent (v/v=2:1) crystallization of petroleum ether-ethyl acetate once, then with recrystallizing methanol once, filter and dry red crystals, obtain the trypterygine cellulose crystal that 4.80 grams of purity are 98.59%, yield is 73.6%.
Embodiment 3
This embodiment is for illustration of the method for extraction Tripterine provided by the invention.
Get the dry root skin of 1500 grams of tripterygium plant trypterygines, each with the aqueous ethanolic solution that 8 liters of concentration are 65 volume % carry out refluxing extraction 2 hours, filter, extraction three times altogether.The concentration extracting Tripterine in the alcohol extract obtained for three times is followed successively by 0.0762 % by weight, 0.0314 % by weight and 0.0110 % by weight.At 25 DEG C, three alcohol extracts are carried out loading absorption with being equipped with (purchased from Zhengzhou Qin Shi Science and Technology Ltd.) in the resin column of the pretreated AB-8 macroporous adsorbent resin of 900mL successively, loading flow velocity is 2800mL/h, wash-out is carried out with the acetone soln that concentration is 65 volume % after loading reaches terminal, controlling the flow velocity of acetone soln in resin column is 1500mL/h, collects red elutriant and verifies as the elutriant being rich in Tripterine through thin-layer chromatography.This is rich in the elutriant of Tripterine concentrated after, first use mixed solvent (v/v=5:2) crystallization of petroleum ether-ethyl acetate once, again with concentration be the ethyl alcohol recrystallization of 95 volume % once, filter and dry red crystals, obtain the trypterygine cellulose crystal that 5.91 grams of purity are 98.07%, yield is 62.4%.
Embodiment 4
This embodiment is for illustration of the method for extraction Tripterine provided by the invention.
Get the dry root skin of 10 kilograms of celastraceae plants celastrus angulatus, carry out seepage pressure effects with the aqueous ethanolic solution that concentration is 70 volume %, stop when the content of Tripterine is 0.005 % by weight to extracting solution extracting.At 25 DEG C, alcohol extract is carried out loading absorption with (purchased from the green hundred careless development in science and technology company limiteds in Beijing) in the resin column that 7 liters of pretreated SP825L macroporous adsorbent resins are housed, loading flow velocity is 20L/h, wash-out is carried out with methyl alcohol after loading reaches terminal, controlling the flow velocity of methyl alcohol in resin column is 13L/h, collects red elutriant and verifies as the elutriant being rich in Tripterine through thin-layer chromatography.This is rich in the elutriant of Tripterine concentrated after, first use mixed solvent (v/v=2:1) crystallization of petroleum ether-ethyl acetate once, then with recrystallizing methanol once, filter and dry red crystals, obtain the trypterygine cellulose crystal that 50.12 grams of purity are 97.99%, yield is 73.3%.
Embodiment 5
This embodiment is for illustration of the method for extraction Tripterine provided by the invention.
According to the method for embodiment 4, Tripterine is extracted, unlike, in seepage pressure effects process, described concentration is that the isopropanol water solution of 50 volume % of the aqueous ethanolic solution same volume of 70 volume % is replaced.Finally obtain the trypterygine cellulose crystal that 45.27 grams of purity are 98.03%, yield is 66.2%.
Embodiment 6
This embodiment is for illustration of the method for extraction Tripterine provided by the invention.
According to the method for embodiment 4, Tripterine is extracted, unlike, in the process of carrying out being separated with macroporous adsorptive resins, the aqueous acetone solution of 60 volume % of eluent methyl alcohol same volume used substitutes.Finally obtain the trypterygine cellulose crystal that 48.65 grams of purity are 97.79%, yield is 71%.
Comparative example 1
This comparative example is for illustration of the method for the extraction Tripterine of reference.
According to the method for embodiment 5, Tripterine is extracted, unlike, in the process of being carried out being separated by alcohol extract macroporous adsorptive resins, carry out drip washing with the aqueous ethanolic solution of the aqueous ethanolic solution of water, 40 volume %, the aqueous ethanolic solution of 70 volume % and 95 volume % as eluent successively.Finally obtain the trypterygine cellulose crystal that 40.87 grams of purity are 98.09%, yield is 59.8%.
As can be seen from the above results, adopt extracting method of the present invention, purity and all higher Tripterine of yield can not only be obtained, but also simplify abstraction and purification process, have prospects for commercial application.
More than describe the preferred embodiment of the present invention in detail; but the present invention is not limited to the detail in above-mentioned embodiment, within the scope of technical conceive of the present invention; can carry out multiple simple variant to technical scheme of the present invention, these simple variant all belong to protection scope of the present invention.
It should be noted that in addition, each the concrete technical characteristic described in above-mentioned embodiment, in reconcilable situation, can be combined by any suitable mode.In order to avoid unnecessary repetition, the present invention illustrates no longer separately to various possible array mode.
In addition, also can carry out arbitrary combination between various different embodiment of the present invention, as long as it is without prejudice to thought of the present invention, it should be considered as content disclosed in this invention equally.
Claims (13)
1. extract a method for Tripterine, it is characterized in that, the method comprises and being contacted with the first lower aliphatic alcohols by the plant material containing Tripterine, makes the content obtaining Tripterine be not less than the alcohol extract of 0.005 % by weight; Again described alcohol extract macroporous adsorptive resins is separated, and the elutriant collected containing Tripterine, the aqueous solution of described second lower aliphatic alcohols that the eluent adopted in sepn process is the second lower aliphatic alcohols, concentration is not less than 90 volume % or concentration are not less than the aqueous acetone solution of 60 volume %; Again by concentrated for the described elutriant containing Tripterine also crystallization; Described first lower aliphatic alcohols is that the form of the aqueous solution of 25-75 volume % uses with concentration.
2. method according to claim 1, wherein, described first lower aliphatic alcohols and the second lower aliphatic alcohols identical or different, and be C independently of one another
1-C
5monohydroxy-alcohol.
3. method according to claim 2, wherein, described first lower aliphatic alcohols and the second lower aliphatic alcohols identical or different, and be methyl alcohol and/or ethanol independently of one another.
4. according to the method in claim 1-3 described in any one, wherein, the condition of described contact comprises the temperature of contact is 25 DEG C of boiling points to described first lower aliphatic alcohols.
5. according to the method in claim 1-3 described in any one, wherein, described macroporous adsorbent resin is that nonpolar macroporous adsorption resin is to intermediate-polarity macroporous adsorption resin.
6. method according to claim 5, wherein, described macroporous adsorbent resin is selected from the trade mark is one or more macroporous adsorbent resin in SP825L, SP207, ADS-5, ADS-8, D101-1, AB-8, HP10, LSA-10, LSA-20, LSA-21, LSA-30, LSA-33 and XDA-6.
7. according to the method in claim 1-3 described in any one, wherein, the condition of described separation comprises: temperature is 10-40 DEG C; With the Tripterine in alcohol extract described in 1g for benchmark, the volume of the macroporous adsorbent resin of filling in described macroporous adsorptive resins is 80-200mL; The flow velocity of described alcohol extract be 2-4 times of column volume/hour; The flow velocity of described eluent be 1.5-3.5 times of column volume/hour.
8. according to the method in claim 1-3 described in any one, wherein, the method for described crystallization comprises: first carry out crystallization with the mixed solvent of sherwood oil and ethyl acetate, then use C
1-C
5fatty alcohol or concentration be not less than the C of 95 volume %
1-C
5the aqueous solution of fatty alcohol carry out recrystallization.
9. method according to claim 8, wherein, described C
1-C
5fatty alcohol be methyl alcohol and/or ethanol.
10. method according to claim 8, wherein, described alcohol extract macroporous adsorptive resins is carried out being separated comprise alcohol extract being introduced from one end of macroporous adsorptive resins to contact with described macroporous adsorbent resin and adsorb, effluent liquid after absorption is drawn from the other end of macroporous adsorptive resins, and carry out wash-out with eluent, collect the elutriant containing Artemisinin; The method also comprises the Extraction solvent described effluent liquid circulation being used as the plant material containing Tripterine.
11. methods according to claim 1, wherein, the plant material containing Tripterine is root and/or the root skin of Celastraceae celastraceae plants and/or tripterygium plant.
12. methods according to claim 11, wherein, described Celastraceae celastraceae plants is selected from Celastrus hookeri, bastard indigo Stem of Oriental Bittersweet, powder back of the body Stem of Oriental Bittersweet, celastrus angulatus, Stem of Oriental Bittersweet, nerville show in post Stem of Oriental Bittersweet, Celastrus aceleatus Merr, haw rattan, short stalk Stem of Oriental Bittersweet, wide leaf short stalk Stem of Oriental Bittersweet, only son rattan and large bud Stem of Oriental Bittersweet one or more.
13. methods according to claim 11, wherein, described tripterygium plant is trypterygine.
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| CN103242414B (en) * | 2013-05-17 | 2015-09-30 | 成都彼斯特生物科技有限公司 | A kind of method from Stem of Oriental Bittersweet medicinal material separation and purification Tripterine |
| CN103989686A (en) * | 2014-05-13 | 2014-08-20 | 台州恩泽医疗中心(集团) | Drug used for preventing or treating atherosclerosis |
| CN106243181A (en) * | 2016-07-29 | 2016-12-21 | 合肥中科玛卡生物技术有限公司 | A kind of method extracting tripterine from Fructus Momordicae charantiae |
| CN107118251B (en) * | 2017-06-06 | 2019-06-28 | 中国科学院上海药物研究所 | The A crystal form and B crystal form of Celastrol, and its preparation method and application |
| CN107987118A (en) * | 2018-01-17 | 2018-05-04 | 贵州民族大学 | A kind of isolation and purification method of Celastrol |
| CN115785187A (en) * | 2022-11-23 | 2023-03-14 | 湖北民族大学 | Extraction method of tripterine |
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