CN102381962A - Extraction method of effective components of Chinese angelica - Google Patents
Extraction method of effective components of Chinese angelica Download PDFInfo
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- CN102381962A CN102381962A CN2011103181043A CN201110318104A CN102381962A CN 102381962 A CN102381962 A CN 102381962A CN 2011103181043 A CN2011103181043 A CN 2011103181043A CN 201110318104 A CN201110318104 A CN 201110318104A CN 102381962 A CN102381962 A CN 102381962A
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Abstract
The invention relates to an extraction method of effective components of Chinese angelica. The extraction method comprises the following steps: 1, Chinese angelica is crushed; 2, crushed Chinese angelica is added to a solvent A solution with the pH value of 5.5; 3, cellulase with the concentration of 0.3mg.mL<-1> is added to the solution obtained in step 2; 4, the solution obtained in step 3 is subjected to enzymatic hydrolysis for 6h at a constant temperature of 55DEG C; 5, isoamylase with the concentration of 0.3mg.ml<-1> is added to the solution obtained in step 4; 6, a solution obtained in step 5 is subjected to enzymatic hydrolysis for 6h at a constant temperature of 55DEG C; 7, the solution obtained in step 6 is subjected to reduced pressure filtration after completing the enzymatic hydrolysis, and the obtained filtrate is preserved in a dark place; 8, the filter residue obtained by filtering in step 7 is used for extracting ligustilide, an eight time solvent B is added each time, reflux extraction is twice carried out for 2h, the obtained first filtrate and the obtained second filtrate are mixed to obtain a mixed filtrate which is a ligustilide solution, and the ligustilide solution is preserved in a low temperature dark place; 9, the filter residue obtained in step 8 is used for extracting angelica polysaccharides, an fifteen time solvent C is added to the filter residue to extract; 10, the filtrate obtained in step 9 is concentrated to obtain a concentrate; and 11, a solvent D is added to the concentrate obtained in step 10 until that the concentration of the solvent D reaches 75% to obtain a deposition, and the deposition is centrifuged by a centrifuge to obtain the angelica polysaccharides.
Description
Technical field
The invention belongs to the Chinese medicine technology for extracting effective component.
Background technology
Medicinal material is famous Chinese medicine simply when the root that is classified as the umbelliferae per nnial herb.
At present from Radix Angelicae Sinensis, extract related technology and mainly comprise above-mentioned topmost three kinds of activeconstituentss, that is: be the fat-soluble component of representative with the Z-ligustilide and be the water soluble component of representative with FLA and Radix Angelicae Sinensis polysaccharide.Relevant research confirms with the Z-ligustilide to be the Radix Angelicae Sinensis fat-soluble component of representative and to be that the Radix Angelicae Sinensis water soluble component of representative is all unstable with the FLA; Isomerization reactions such as dehydrogenation, oxidation, hydrolysis, degraded take place easily, therefore extraction, separation and purification and preservation condition are required relatively harsher.
Process for extracting based on Chinese traditional medicine angelica effective constituent FLA mainly is traditional wet distillation and organic solvent extraction.But FLA is a determination system of thermal unstable material, is heated for a long time, can cause the decomposition of FLA, so adopt the yield of FLA in this type of heating extraction Radix Angelicae Sinensis not high.
The process for extracting of Radix Angelicae Sinensis volatile oil mainly includes machine solvent leaching method, steam distillation, supercritical CO
2Extraction process.The organic solvent soaking extraction method is extracted Radix Angelicae Sinensis volatile oil and has that treatment capacity is big, energy consumption is lower, is easy to characteristics such as operate continuously, but this method solvent load is big, and in treating processes, causes the loss of water soluble component easily.The yield that steam distillation extracts Radix Angelicae Sinensis volatile oil is lower, simultaneously because the leaching process temperature is high, is prone to cause thermally labile is reached the destruction that is prone to oxydised component.
Radix Angelicae Sinensis polysaccharide is made up of D-semi-lactosi, L pectinose, D-wood sugar, glucuronic acid, galacturonic acid etc., and Radix Angelicae Sinensis polysaccharide adopts the traditional method for extracting of poach alcohol precipitation usually at present.
The Chinese medicine ingredient is very complicated, and existing effective constituent has invalid components and toxic ingredient again.In order to improve the inner quality of Chinese medicine preparation, strengthen its result of treatment, reduce toxic side effect, select for use rational extraction process extremely important.The process for extracting that the Radix Angelicae Sinensis active part is commonly used at present has traditional water steam distillation extraction and organic solvent extraction.These traditional methods have been passed by the very long historical years, because its scientific and technical kernel that rationally is suitable for, being continuous is still Radix Angelicae Sinensis extraction method commonly used so far; But these methods exist tangible deficiency and problem; Embody a concentrated expression of, modern pharmacological research shows that low polarity compounds (comprising volatile oil) is the important substance basis of its some drug effect of Radix Angelicae Sinensis performance, yet tradition is taked water vapour distillation; So high temperature is unfavorable for the extraction of unsettled low polarity compounds; The drug effect of Chinese medicine is not from a certain single active chemical components in addition, but from the synergy between the various active composition, even relevant with some non-active ingredient; But traditional extraction technique is mostly to a certain (or a certain type) component; And be that index comes Different Extraction Method is estimated with the extracted amount of a certain component (or a certain type) component, so the utilization ratio of medicinal material is not high, the yield of activeconstituents is low.Methods such as supercritical extraction, ultrasonic and microwave-assisted extraction belong to the application of new technology in medicinal material extract; It realizes that the big production of pharmacy also needs the regular hour; Although can avoid the destruction of HTHP to effective constituent; But higher to equipment requirements, and the preparation apparatus early investment is bigger, and maintenance cost is higher.In addition, the composition that pharmaceutical use arranged in the botanical herbs is some secondary metabolites of medicinal plant itself often, and the structural material that the overwhelming majority of medicinal plant is it relies and grow; Like Mierocrystalline cellulose, semicellulose, xylogen, pectin; Do not have pharmaceutical use, no matter be China's traditional Chinese medicines concocting method, or the extraction process of modern large enterprises; All be to adopt poach or the structural material of the composition of managing with a large amount of organic solvents Chinese medicine is had a pharmaceutical use and no pharmaceutical use separates; Its limitation is that extracts active ingredients is insufficient or only to the extraction of certain or some compositions, and curative effect is reduced, and limited natural resources of Chinese medicinal materials is wasted in a large number.
Although existing processes can satisfy the separation of Radix Angelicae Sinensis activeconstituents in existing enterprise's Chinese medicinal materials processing; But; Existing processes all is certain effective component that is directed against wherein, for example: extract FLA, adopt to add thermal distillation or organic solvent extraction no matter be; The capital causes the loss to Z-ligustilide, promptly adopts existing the report after technology is extracted FLA can not extract Z-ligustilide again; Equally, no matter be to adopt distillation method or adopt organic solvent method to extract Z-ligustilide, residual residue all can not be used for the extraction of FLA.
Summary of the invention
The object of the invention provides a kind of Radix Angelicae Sinensis extraction of effective components.
The present invention is a kind of Radix Angelicae Sinensis extraction of effective components, the steps include:
(1) exsiccant Radix Angelicae Sinensis piece root is crushed to 100 order particles in kibbler;
(2) take by weighing Radix Angelicae Sinensis after the pulverizing, with 1: 8 different feed liquid than or mass ratio add the solvent orange 2 A solution of pH5.5;
(3) in the solution of (2), add 0.3mgmL again
-1Cellulase, fully stir;
(4) with (3) gained solution enzymolysis 6 hours under 55 ℃ of constant temperatures;
(5) in the solution of (4), add 0.3mgmL again
-1Isoamylase, fully stir;
(6) (5) gained solution is continued to carry out the constant temperature enzymolysis at 55 ℃, time remaining 6 hours;
(7) filtration under diminished pressure after enzymolysis is accomplished, gained filtrating is FLA solution, and this solution keeps in Dark Place;
(8) the gained filter residue is filtered in (7) and be used to extract Z-ligustilide, add the solvent B of 8 times of amounts, refluxing extraction 2 times, each 2 hours at every turn; To filtrate for the first time and filtrating merging for the second time, the filtrating after the merging is Z-ligustilide solution, this solution lucifuge cryopreservation;
(9) the gained filter residue is filtered in (8) and be used to extract Radix Angelicae Sinensis polysaccharide, add the solvent C of 15 times of amounts in the filter residue, extracted 3 hours, extracting temperature is 80 ± 5 ℃;
(10) with 8 layers of filtered through gauze, discard filter residue after extraction is accomplished, filtrate volume is concentrated 5 times;
(11) in (10) liquid concentrator, add solvent D, fully stir, reach 75% to solvent strength, produce a large amount of flockss, with whizzer 3000rpm centrifugal 10 minutes, separation obtaining Radix Angelicae Sinensis polysaccharide.
Described solvent orange 2 A is a Glacial acetic acid min. 99.5; Solvent B is 70% ethanol; Solvent C is a zero(ppm) water; Solvent D is 95% ethanol.
Compare with traditional method, traditional method for extracting FLA yield is 0.52mg/g, and the Z-ligustilide yield is 120.6mg/g, and polysaccharide yield is 76.9mg/g, and FLA and Z-ligustilide can't carry out common extraction; FLA yield of the present invention is 0.55mg/g, and the Z-ligustilide yield is 119.1mg/g, and polysaccharide yield is 93.6mg/g, and the present invention can realize the common extraction of main active ingredient FLA, Z-ligustilide in the Radix Angelicae Sinensis and the extraction of later stage Radix Angelicae Sinensis polysaccharide.
Process for extracting of the present invention has following advantage: because the high efficiency and the specificity of enzyme, action condition is gentle, and extraction conditions of the present invention is simpler, equipment is more simplified, and not only cuts down the consumption of energy but also practice thrift cost; Relative prior art, this method realize to the common extraction of Z-ligustilide and the extraction of later stage Radix Angelicae Sinensis polysaccharide having the drug quality of guaranteeing, reduce extraction cost, reduce the natural resources of Chinese medicinal materials waste when effectively putting forward the FLA yield.
Description of drawings
Fig. 1 is a FLA standard substance HPLC color atlas, and Fig. 2 is a FLA color atlas in the sample, and Fig. 3 is a Z-ligustilide standard substance HPLC color atlas, and Fig. 4 is a Z-ligustilide color atlas in the sample.
Embodiment
The present invention is a kind of Radix Angelicae Sinensis extraction of effective components, the steps include:
(1) exsiccant Radix Angelicae Sinensis piece root is crushed to 100 order particles in kibbler;
(2) take by weighing Radix Angelicae Sinensis after the pulverizing, with different feed liquid than or mass ratio add the solvent orange 2 A solution of pH5.5;
(3) in the solution of (2), add 0.3mgmL again
-1Cellulase, fully stir;
(4) with (3) gained solution enzymolysis 6 hours under 55 ℃ of constant temperatures;
(5) in the solution of (4), add 0.3mgmL again
-1Isoamylase, fully stir;
(6) (5) gained solution is continued to carry out the constant temperature enzymolysis at 55 ℃, time remaining 6 hours;
(7) filtration under diminished pressure after enzymolysis is accomplished, gained filtrating is FLA solution, and this solution keeps in Dark Place;
(8) the gained filter residue is filtered in (7) and be used to extract Z-ligustilide, add the solvent B of 8 times of amounts, refluxing extraction 2 times, each 2 hours at every turn; To filtrate for the first time and filtrating merging for the second time, the filtrating after the merging is Z-ligustilide solution, this solution lucifuge cryopreservation;
(9) the gained filter residue is filtered in (8) and be used to extract Radix Angelicae Sinensis polysaccharide, add the solvent C of 15 times of amounts in the filter residue, extracted 3 hours, extracting temperature is 80 ± 5 ℃;
(10) extraction is used filtered through gauze after accomplishing, and discards filter residue, and filtrate volume is concentrated 5 times;
(11) in (10) liquid concentrator, add solvent D, fully stir, reach 75% to solvent strength, produce a large amount of flockss, with whizzer 3000rpm centrifugal 10 minutes, separation obtaining Radix Angelicae Sinensis polysaccharide.
Described solvent orange 2 A is a Glacial acetic acid min. 99.5; Solvent B is 70% ethanol; Solvent C is a zero(ppm) water; Solvent D is 95% ethanol.
Above-described prozyme is a cellulase, perhaps polygalacturonase, perhaps isoamylase, perhaps zytase, perhaps above-described two or more combination.
Described prozyme is cellulase and isoamylase preferably, in 1: 1 ratio combination.
Adopt the HPLC method to detect main active ingredient in the Radix Angelicae Sinensis to method of the present invention
1, detect with the extraction yield variation of HPLC method to the Enzymatic Extraction FLA, chromatographic condition is chromatographic column: the reverse post of ODS (4.6 μ m * 250mm, 5 μ m); Moving phase: methyl alcohol-0.085% phosphoric acid (55: 45, v/v); Flow velocity: 1ml/min; Sample size 20 μ l; Detect wavelength: 322nm; Column temperature: room temperature.Under this chromatographic condition, impurity peaks is better separated in target peak and the sample.The FLA color atlas is like Fig. 1, shown in 2 in FLA standard substance color atlas, the sample.
The production standard curve
Accurately take by weighing FA standard substance 1mg, be dissolved in the 1ml moving phase, be mixed with 1000 μ g/ml standard reserving solutions.Get an amount of standard reserving solution and be diluted to the standardized solution that concentration is respectively 0.5,1,5,10,25,50,100 μ g/ml, respectively sample detection with moving phase.With standard substance concentration is X-coordinate, and the HPLC peak area is that ordinate zou carries out linear regression, makes the FLA typical curve.
This method gained regression equation is y=117053x+32234, dependency R
2=0.9999, the result shows accurately ferulaic acid content in the test sample of HPLC method.
2, detect with the extraction yield variation of HPLC method to Z-ligustilide in the Radix Angelicae Sinensis, chromatographic condition is chromatographic column: the reverse post of ODS (4.6 μ m * 250mm, 5 μ m); Moving phase: methanol-water (70: 30, v/v); Flow velocity: 1ml/min, sample size 20 μ l; Detect wavelength: 327nm
[1]Under this chromatographic condition, impurity peaks is better separated in target peak and the sample.The Z-ligustilide color atlas is like Fig. 3, shown in 4 in Z-ligustilide standard substance color atlas, the sample.
The production standard curve
Accurately take by weighing Z-ligustilide standard substance 1mg, add the dissolving of 500 μ l moving phases, and gradient dilution gets the standardized solution that concentration is 5,10,25,50,100,250,500,1000,2000 μ g/ml, sample detection.With ester concentration in the manuscript is X-coordinate, and the HPLC peak area is that ordinate zou carries out linear regression, makes the Z-ligustilide typical curve.
This method gained regression equation is y=3453.3x-4070.2, dependency R
2=0.9997, the result shows accurately Z-ligustilide content in the test sample of HPLC method.
Further launch the present invention with more concrete embodiment below.
Embodiment 1
Main active ingredient in the Enzymatic Extraction Radix Angelicae Sinensis
Taking by weighing the Radix Angelicae Sinensis powder 10g after the pulverizing, is the glacial acetic acid solution that 1: 8 ratio (mass ratio) adds pH5.5 with solid-liquid ratio, adds 0.3mgmL again
-1Cellulase, fully stir enzymolysis 6h under 55 ℃ of constant temperature; Add 0.3mgmL again
-1Isoamylase, fully stir, continue 55 ℃ of constant temperature enzymolysis 6h.Filtration under diminished pressure after enzymolysis is accomplished, gained filtrating is FLA solution, and this solution keeps in Dark Place, and detects the FLA yield with the HPLC method.To filter the gained filter residue and be used to extract Z-ligustilide; (promptly 70% alcohol reflux is 2 times with traditional alcohol extracting method; 2h/ time) extract, will filtrate for the first time and filtrating merging for the second time, the filtrating after the merging is Z-ligustilide solution; This solution lucifuge cryopreservation, and with HPLC method detection Z-ligustilide yield.This step gained filter residue extracts Radix Angelicae Sinensis polysaccharide with traditional water extraction and alcohol precipitation method then, detects polysaccharide yield with sulfuric acid-phynol method.
The FLA yield is 0.55mg/g among the present invention, and the Z-ligustilide yield is 119.1mg/g, and polysaccharide yield is 93.6mg/g, and the yield of main active ingredient is all suitable with traditional method in the Radix Angelicae Sinensis, but to the utilization ratio of crude drug far above traditional method.
Combined-enzyme method extracts main active ingredient in the Radix Angelicae Sinensis
Taking by weighing the Radix Angelicae Sinensis powder 10g after the pulverizing, is the glacial acetic acid solution that 1: 8 ratio (mass ratio) adds pH5.5 with solid-liquid ratio, adds 0.2mgmL again
-1Cellulase, fully stir, enzymolysis 5h under 55 ℃ of constant temperature adds 0.2mgmL again
-1Polygalacturonase, fully stir, continue 55 ℃ of constant temperature enzymolysis 5h.Filtration under diminished pressure after enzymolysis is accomplished, gained filtrating is FLA solution, and this solution keeps in Dark Place, and detects the FLA yield with the HPLC method.To filter the gained filter residue and be used to extract Z-ligustilide; (promptly 70% alcohol reflux is 2 times with traditional alcohol extracting method; 2h/ time) extract, will filtrate for the first time and filtrating merging for the second time, the filtrating after the merging is Z-ligustilide solution; This solution lucifuge cryopreservation, and with HPLC method detection Z-ligustilide yield.This step gained filter residue extracts Radix Angelicae Sinensis polysaccharide with traditional water extraction and alcohol precipitation method then, detects polysaccharide yield with sulfuric acid-phynol method.
The FLA yield is 0.52mg/g among the present invention, and the Z-ligustilide yield is 121.3mg/g, and polysaccharide yield is 90.1mg/g,
Embodiment 3
Combined-enzyme method extracts main active ingredient in the Radix Angelicae Sinensis
Taking by weighing the Radix Angelicae Sinensis powder 10g after the pulverizing, is the glacial acetic acid solution that 1: 8 ratio (mass ratio) adds pH5.5 with solid-liquid ratio, adds 0.3mgmL again
-1Cellulase, fully stir, enzymolysis 5h under 55 ℃ of constant temperature adds 0.3mgmL again
-1Zytase, fully stir, continue 55 ℃ of constant temperature enzymolysis 5h.Filtration under diminished pressure after enzymolysis is accomplished, gained filtrating is FLA solution, and this solution keeps in Dark Place, and detects the FLA yield with the HPLC method.To filter the gained filter residue and be used to extract Z-ligustilide; (promptly 70% alcohol reflux is 2 times with traditional alcohol extracting method; 2h/ time) extract, will filtrate for the first time and filtrating merging for the second time, the filtrating after the merging is Z-ligustilide solution; This solution lucifuge cryopreservation, and with HPLC method detection Z-ligustilide yield.This step gained filter residue extracts Radix Angelicae Sinensis polysaccharide with traditional water extraction and alcohol precipitation method then, detects polysaccharide yield with sulfuric acid-phynol method.
The FLA yield is 0.51mg/g among the present invention, and the Z-ligustilide yield is 118.2mg/g, and polysaccharide yield is 87.3mg/g.
Claims (3)
1. a Radix Angelicae Sinensis extraction of effective components the steps include:
(1) exsiccant Radix Angelicae Sinensis piece root is crushed to 100 order particles in kibbler;
(2) take by weighing Radix Angelicae Sinensis after the pulverizing, add the solvent orange 2 A solution of pH5.5 with 1: 8 solid-liquid ratio or mass ratio;
(3) in the solution of (2), add 0.3mgmL again
-1Cellulase, fully stir;
(4) with (3) gained solution enzymolysis 6 hours under 55 ℃ of constant temperatures;
(5) in the solution of (4), add 0.3mgmL again
-1Isoamylase, fully stir;
(6) (5) gained solution is continued to carry out the constant temperature enzymolysis at 55 ℃, time remaining 6 hours;
(7) filtration under diminished pressure after enzymolysis is accomplished, gained filtrating is FLA solution, and this solution keeps in Dark Place;
(8) the gained filter residue is filtered in (7) and be used to extract Z-ligustilide, add the solvent B of 8 times of amounts, refluxing extraction 2 times, each 2 hours at every turn; To filtrate for the first time and filtrating merging for the second time, the filtrating after the merging is Z-ligustilide solution, this solution lucifuge cryopreservation;
(9) the gained filter residue is filtered in (8) and be used to extract Radix Angelicae Sinensis polysaccharide, add the solvent C of 15 times of amounts in the filter residue, extracted 3 hours, extracting temperature is 80 ± 5 ℃;
(10) extraction is used filtered through gauze after accomplishing, and discards filter residue, and filtrate volume is concentrated 5 times;
(11) in (10) liquid concentrator, add solvent D, fully stir, reach 75% to solvent strength, produce a large amount of flockss, with whizzer 3000rpm centrifugal 10 minutes, separation obtaining Radix Angelicae Sinensis polysaccharide;
Described solvent orange 2 A is a Glacial acetic acid min. 99.5; Solvent B is 70% ethanol; Solvent C is a zero(ppm) water; Solvent D is 95% ethanol.
2. Radix Angelicae Sinensis extraction of effective components according to claim 1 is characterized in that described prozyme is a cellulase, perhaps polygalacturonase, perhaps isoamylase, perhaps zytase, perhaps above-described two or more combination.
3. Radix Angelicae Sinensis extraction of effective components according to claim 1 is characterized in that described prozyme preferably cellulase and isoamylase, in 1: 1 ratio combination.
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| CN103709029A (en) * | 2013-12-13 | 2014-04-09 | 大兴安岭嘉迪欧营养原料有限公司 | Process for extracting and purifying ferulic acid from angelica cartilaginomarginata Nakai |
| CN105246868A (en) * | 2013-05-21 | 2016-01-13 | 罗地亚经营管理公司 | Optimized process for extraction of ferulic acid with pretreatment |
| CN106692234A (en) * | 2017-03-03 | 2017-05-24 | 四川德成动物保健品有限公司 | Method for extracting active ingredients of angelica sinensis in Yimu biochemical mixture |
| CN109867598A (en) * | 2017-12-01 | 2019-06-11 | 北京国康本草物种生物科学技术研究院有限公司 | Radix Angelicae Sinensis extraction process of effective component and its extract |
| CN112273709A (en) * | 2020-10-23 | 2021-01-29 | 湖北中烟工业有限责任公司 | Preparation method of Shiyao angelica extract for perfuming tobacco |
| CN115364132A (en) * | 2022-09-30 | 2022-11-22 | 兰州海关技术中心 | Preparation method of angelica sinensis extract |
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Cited By (8)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN105246868A (en) * | 2013-05-21 | 2016-01-13 | 罗地亚经营管理公司 | Optimized process for extraction of ferulic acid with pretreatment |
| US10202328B2 (en) | 2013-05-21 | 2019-02-12 | Rhodia Operations | Optimized process for extraction of ferulic acid with pretreatment |
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| CN103709029B (en) * | 2013-12-13 | 2016-06-29 | 大兴安岭嘉迪欧营养原料有限公司 | A kind of technique of extraction purification ferulic acid from Guyuling Capsule |
| CN106692234A (en) * | 2017-03-03 | 2017-05-24 | 四川德成动物保健品有限公司 | Method for extracting active ingredients of angelica sinensis in Yimu biochemical mixture |
| CN109867598A (en) * | 2017-12-01 | 2019-06-11 | 北京国康本草物种生物科学技术研究院有限公司 | Radix Angelicae Sinensis extraction process of effective component and its extract |
| CN112273709A (en) * | 2020-10-23 | 2021-01-29 | 湖北中烟工业有限责任公司 | Preparation method of Shiyao angelica extract for perfuming tobacco |
| CN115364132A (en) * | 2022-09-30 | 2022-11-22 | 兰州海关技术中心 | Preparation method of angelica sinensis extract |
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| CN102381962B (en) | 2013-12-11 |
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