CN102464635A - Separation and purification method of arctigenin - Google Patents
Separation and purification method of arctigenin Download PDFInfo
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- CN102464635A CN102464635A CN2010105404802A CN201010540480A CN102464635A CN 102464635 A CN102464635 A CN 102464635A CN 2010105404802 A CN2010105404802 A CN 2010105404802A CN 201010540480 A CN201010540480 A CN 201010540480A CN 102464635 A CN102464635 A CN 102464635A
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Abstract
Belonging to the technical field of medicines, the invention specifically provides a separation and purification method of arctigenin. The method comprises the steps of: dissolution, filtration, separation and purification through high efficiency preparative liquid chromatography, product recovery, etc. Specifically, the separation and purification adopts reversed-phase high efficiency preparative liquid chromatography, a dynamic axial compression column is employed as a chromatographic column, an acetonitrile water solution is taken as a mobile phase, and an ultraviolet detector is used for on-line detection. The method of the invention has the advantages of simple operation, short production period, high separation efficiency, stable technology, low cost, and can realize high-purity separation and preparation of a lot of arctigenin.
Description
Technical field
The present invention relates to biochemical field, be specifically related to a kind of separation purification method of l-arctigenin.
Background technology
L-arctigenin (arctigenin), and chemical name (3R, 4R)-4-[(3,4-dimethoxyphenyl) methyl) dihydro-3-[(4-hydroxy-3-methoxyphenyl) methyl]-2 (3H)-furanone; CAS number: 7770-78-7; Molecular formula: C
21H
24O
6Molecular weight 372.41; Structural formula is following:
L-arctigenin is the effective constituent that can directly be absorbed by the body; Be used for treatment and prevention chronic renal failure and renal fibrosis, antiviral, antitumor, skin whitening, antianaphylaxis, anti-inflammatory, treatment mellitus and complication, dn-, also be used to kill aquatic animal vermin etc.
Natural l-arctigenin mainly is present in composite family (Compasitate) burdock burdock (Arctiumlappal).The method for preparing at present l-arctigenin mainly contains two big types: the one, from plant materials, directly extract; The 2nd, the arctinin higher by content in the plant materials gets through hydrolysis; The l-arctigenin bullion of this two class methods gained all needs purifying, and purification process mainly adopts chromatography, recrystallization method at present; And the combination of two kinds of methods, wherein chromatography comprises silica gel column chromatography and liquid phase method.
The purification process that adopts l-arctigenin among CN200710135648.X and the CN200710135646. was a silica gel H post decompression post, the chloroform-methanol gradient elution, and the purity of final gained l-arctigenin is 98.2%.Gradient elution is had relatively high expectations, and chloroform toxicity is bigger, and the solvent for use amount is big, pollutes greatly, and the purity of last gained l-arctigenin is lower.
Adopting the purification process of l-arctigenin among the CN200810094377.2 is with 60 order silica gel mixed samples with the l-arctigenin bullion; Thick silica gel volume be the l-arctigenin bullion 1-2 doubly, water-bath oven dry, appearance on the dry method; Through 200-300 order silica gel column chromatography; 98: 1 wash-outs of chloroform-methanol, elutriant concentrate to merge and promptly get l-arctigenin, and it is more than 90% that HPLC records purity.
Use silica gel column chromatography among the CN200410015465; Solvent recrystallization or the sub-aglycon of both bonded method purified edible burdocks; Wherein silica gel column chromatography needs through one or many, with chloroform/methanol (100: 0-95: 5) be elutriant, indicate with TLC (chloroform/methanol (97: 3)); Through recrystallizing methanol, getting l-arctigenin is 99% through HPLC detection purity again.Detect behind elder generation's chromatography, detect hysteresis, efficient is low, complex steps, and quality is wayward, and yield is low.
" chromatogram " 2003,31 (1): 52-55), Liu Shiming etc.; The separating and evaluation of micro-lignan arctinin and l-arctigenin in the Herba Arctii leaf, adopt arctinin and l-arctigenin in the reverse phase liquid chromatography separation and purification burdock leaf trace lignan, the chromatographic column of employing is common analytical column, applied sample amount is little; Gradient elution, higher to equipment requirements; Except containing the first alcohol and water, also contain phosphoric acid in the moving phase, cause in product reclaims to be difficult to remove, the purity of the finished product is not high, more than 95%.
Summary of the invention
To the deficiency that exists in the above-mentioned prior art, the present invention provides a kind of method that adopts anti-phase preparative liquid chromatography separation and purification l-arctigenin, and the inventive method adopts the preparative high-performance liquid chromatographic system that l-arctigenin is carried out separation and purification; Through suitable pre-treating process and chromatographic condition; Reach good separating effect, and the UV-detector on-line monitoring, the process intuitive operation is easy; Specific aim is collected l-arctigenin; With clearly defined objective, avoid conventional column chromatography to separate earlier and afterwards detected the wasting of resources that causes, and be easy to control quality product.Products obtained therefrom purity, yield height.The reference substance that can be used as assay uses, and also can directly be used for doing its preparation.
In the high performance preparative liquid chromatography sepn process, the selection of chromatographic condition is extremely important, and it is to decisive role such as the peak sequence of each material in the sample solution, peak shape, separating effects; Chromatographic condition mainly comprises chromatographic column (comprising filler, column length and internal diameter etc.), moving phase (comprise form and flow velocity etc.), column temperature, detection wavelength, detector etc., the selection of each chromatographic condition and make up most important.
The inventor is through great deal of experimental and comparative analysis; Confirmed aforesaid each chromatographic condition; Make the optimizations such as appearance time, peak shape, separating effect of each material in the sample solution, helping the l-arctigenin monomer is well separated with each impurity, and shortens the production cycle.
In order to realize the foregoing invention purpose, the technical scheme that the present invention adopts is following:
The l-arctigenin bullion is dissolved with acetonitrile-aqueous solution, filter, filtrating further is diluted to the l-arctigenin solution of 8-15mg/ml with purified water; Gained solution is separated with the high performance preparative liquid chromatography wash-out; With the online detection of UV-detector, specific aim is collected the preparation component of l-arctigenin, gets l-arctigenin solution; The l-arctigenin solution that makes is removed acetonitrile and water with underpressure distillation and freeze-drying successively, promptly get the pure article of l-arctigenin.
Specifically may further comprise the steps:
(1) dissolving, filtration:
Ratio according to solids quality: solvent volume=1Kg: 50L; With acetonitrile-water (volume ratio 9: (0-3)) solution; With solid l-arctigenin dissolving crude product; The dissolving back is with organic filtering with microporous membrane of 0.45 μ m, and filtrating further is diluted to the solution of 8-15mg (being equivalent to the bullion quality)/ml with purified water, as the raw material of the sub-aglycon of next step preparation purified edible burdock.
Elder generation is with the acetonitrile solution dissolution filter of high density in this step; Remove outside the conventional insolubles and bacterium; Can also remove the bigger impurity of segment polarity; The more excellent concentration range of effect is acetonitrile-water (volume ratio 9: (0-3)), and then further dilutes, in order to be applicable to chromatographic system better.
(2) separation and purification:
With the solution 50-100ml after the dilution in the step (1), inject the reversed phase high efficiency preparative liquid chromatograph, isocratic elution, the online detection of UV-detector, specific aim is collected the preparation component of l-arctigenin, gets l-arctigenin solution.
Wherein the used chromatographic column of reversed phase high efficiency preparative liquid chromatography separation aretigenin is a dynamic axial compression column, diameter 80mm, packed height 250mm; Filler is the octadecylsilane chemically bonded silica microballoon, particle diameter 10 μ m; Moving phase is made up of acetonitrile-water, and the volume ratio of acetonitrile-water is 1: (1-4) isocratic elution; Flow velocity 260ml/min-330ml/min; Sample size 50-100ml; Sample concentration 8-15mg/ml; The online detection of UV-detector detects wavelength 280nm.
Carrying out before reversed phase high efficiency prepares liquid phase separation, can confirm the appearance time and the peak shape of l-arctigenin in the performance liquid chromatography through LC-MS, reference substance location or other present technique field method commonly used.
With the LC-MS method is example, can to separate liquid phase chromatogram condition identical with above-mentioned preparation, adopt the identical chromatographic column of filler, and form identical moving phase, identical detection wavelength etc., column temperature is room temperature (have no special requirements, room temperature gets final product); Get the filtrating sample introduction of step (1) gained, carry out the performance liquid separation and purification of l-arctigenin,, can confirm l-arctigenin pairing peak in liquid chromatography according to the mass spectrometric detection result.
(3) product reclaims
Step (2) high performance preparative liquid chromatography separated in the l-arctigenin solution obtain acetonitrile and water and remove through underpressure distillation (0.08MPa-0.09MPa, 35 ℃-65 ℃) and lyophilization successively, promptly get the l-arctigenin product.
Compared with prior art, excellent effect of the present invention is following:
The analysis mode reversed-phased high performace liquid chromatographic is adopted in the detection of the purity of finished product l-arctigenin, and analyzing and testing purity is more than 99.8%, and single mixing all less than 0.1%; Yield is more than 98%.(chromatographic condition: chromatographic column: the C18 post (4.6mm * 150mm, 5um); Moving phase: the methanol-water volume ratio is 60: 40; Flow velocity: 1.0ml/min; Sample size: 10 μ l; Detect wavelength: 280nm; Column temperature: 25 ℃.)
The inventive method technology is simple, and used organic solvent is all recyclable to be utilized again, and solvent consumption is few; Online detection, efficient is high, and quality product is controlled easily; System is a closed system, and operating environment is polluted little; Isocratic elution is simple to equipment requirements; Moving phase adopts volatile organic solvent and water, in product reclaims, removes easily.
Description of drawings
The high-efficient liquid phase analysis collection of illustrative plates of Fig. 1 l-arctigenin bullion
Fig. 2 prepares the HPLC collection of illustrative plates of gained l-arctigenin by the condition of embodiment 1
Fig. 3 prepares the HPLC collection of illustrative plates of gained l-arctigenin by the condition of embodiment 2
Fig. 4 prepares the HRLC collection of illustrative plates of gained l-arctigenin by the condition of embodiment 3
Embodiment
Below in conjunction with embodiment the present invention is made further detailed description, but protection domain of the present invention is not limited to following embodiment.
(1) dissolving, filtration:
Get l-arctigenin bullion 2KG; Be dissolved in acetonitrile-aqueous solution 100L (volume ratio of acetonitrile and water is 9: 3); With solid l-arctigenin dissolving crude product; The dissolving back is with organic filtering with microporous membrane of 0.45 μ m, and filtrating further is diluted with water to the solution of 8mg/ml, as the raw material of next step preparation l-arctigenin.
(2) separation and purification:
With the solution 100ml after the dilution in the step (1), inject the reversed phase high efficiency preparative liquid chromatograph, moving phase is made up of acetonitrile-water; The volume ratio of acetonitrile-water is 35: 65, isocratic elution, flow velocity 320ml/min; The online detection of UV-detector; Detect wavelength 280nm, specific aim is collected the preparation component of l-arctigenin, gets l-arctigenin solution.
(3) product reclaims:
Step (2) high performance preparative liquid chromatography separated in the l-arctigenin solution obtain acetonitrile and water and remove through underpressure distillation (0.08-0.09MPa, 40-50 ℃) and freeze-drying successively, promptly get the l-arctigenin product.Total recovery can reach 98.2%, and product is measured through HPLC, and purity is 99.89%.
Embodiment 2
(1) dissolving, filtration:
Get l-arctigenin bullion 2KG; Be dissolved in acetonitrile-aqueous solution 100L (volume ratio of acetonitrile and water is 9: 2); With solid l-arctigenin dissolving crude product; The dissolving back is with organic filtering with microporous membrane of 0.45 μ m, and filtrating further is diluted with water to the solution of 10mg/ml, as the raw material of next step preparation l-arctigenin.
(2) separation and purification:
With the solution 80ml after the dilution in the step (1), inject the reversed phase high efficiency preparative liquid chromatograph, moving phase is made up of acetonitrile-water; The portion rate of acetonitrile-water is 45: 55, isocratic elution, flow velocity 290ml/min; The online detection of UV-detector; Detect wavelength 280nm, specific aim is collected the preparation component of l-arctigenin, gets l-arctigenin solution.
(3) product reclaims:
Step (2) high performance preparative liquid chromatography separated in the l-arctigenin solution obtain acetonitrile and water and remove, promptly get the l-arctigenin product through underpressure distillation (0.08-0.09MPa, 40-50 ℃) and lyophilization.Warp is cycles prepare repeatedly, and yield can reach 98.3%, and product is measured through HPLC, and purity is 99.82%.
Embodiment 3
(1) dissolving, filtration:
Get l-arctigenin bullion 2KG; Be dissolved in acetonitrile-aqueous solution 100L (volume ratio of acetonitrile and water is 9: 1); With solid l-arctigenin dissolving crude product; The dissolving back is with organic filtering with microporous membrane of 0.45 μ m, and filtrating further is diluted with water to the solution of 15mg/ml, as the raw material of next step preparation l-arctigenin.
(2) separation and purification:
With the solution 50ml after the dilution in the step (1), inject the reversed phase high efficiency preparative liquid chromatograph, moving phase is made up of acetonitrile-water; The portion rate of acetonitrile-water is 50: 50; Isocratic elution, flow velocity 290ml/min, the online detection of UV-detector; Detect wavelength 280nm, specific aim is collected the preparation component of l-arctigenin.
(3) product reclaims:
Step (2) high performance preparative liquid chromatography separated in the l-arctigenin solution obtain acetonitrile and water through underpressure distillation (0.08-0.09MPa; 40-50 ℃) and lyophilization remove, promptly get the l-arctigenin product, yield can reach 98.5%; Product is measured through HPLC, and purity is 99.86%.
Claims (9)
1. the separation purification method of a l-arctigenin may further comprise the steps:
(1) dissolution filter: the l-arctigenin bullion is dissolved with acetonitrile-aqueous solution, filter, filtrating is further used the l-arctigenin solution of purified water dilution as 8-15mg/ml;
(2) separation and purification: step (1) gained solution is separated with the high performance preparative liquid chromatography wash-out,, collect the preparation component of l-arctigenin, get l-arctigenin solution with the online detection of UV-detector;
(3) product reclaims: the l-arctigenin solution that step (2) is made removes acetonitrile and water with underpressure distillation and lyophilization successively, promptly gets the pure article of l-arctigenin.
2. the separation purification method of l-arctigenin as claimed in claim 1 is characterized in that the chromatographic column of step (2) high performance preparative liquid chromatography is a dynamic axial compression column, and filler is the octadecylsilane chemically bonded silica microballoon, and moving phase is acetonitrile-aqueous solution.
3. the separation purification method of l-arctigenin as claimed in claim 1, the volume ratio that it is characterized in that step (2) acetonitrile and the aqueous solution is 9: 0-3.
4. the separation purification method of l-arctigenin as claimed in claim 2 is characterized in that the dynamic axial compression column diameter is 80mm, and packed height is 250mm.
5. the separation purification method of l-arctigenin as claimed in claim 2, the particle diameter that it is characterized in that the octadecylsilane chemically bonded silica microballoon is 10 μ m.
6. the separation purification method of l-arctigenin as claimed in claim 2 is characterized in that the volume ratio of acetonitrile and water is 1 in the said moving phase: 1-4.
7. the separation purification method of l-arctigenin as claimed in claim 1, the type of elution that it is characterized in that the high performance preparative liquid chromatography post is an isocratic elution.
8. the separation purification method of l-arctigenin as claimed in claim 7, the elution speed that it is characterized in that the high performance preparative liquid chromatography post is 260ml/min-330ml/min.
9. the separation purification method of l-arctigenin as claimed in claim 1, the UV-detector that it is characterized in that high performance preparative liquid chromatography is online detection, the detection wavelength is 280nm.
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Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN105769964A (en) * | 2016-03-30 | 2016-07-20 | 苏州勤浩药物研究开发有限公司 | Application of fructus arctii extract in resistance of skin photoage |
Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JPH02142723A (en) * | 1988-11-25 | 1990-05-31 | Tsumura & Co | Rental disorder-improving agent |
| CN1560265A (en) * | 2004-02-26 | 2005-01-05 | 广州中医药大学 | Preparation process of burduck protoglucoside of anti virus and antiptmour natural medicine |
| CN101392279A (en) * | 2008-10-21 | 2009-03-25 | 浙江工业大学 | Method for preparing fructus arctii aglycone |
| CN101736050A (en) * | 2009-12-08 | 2010-06-16 | 辽宁中医药大学 | Preparation method of arctigenin |
-
2010
- 2010-10-30 CN CN201010540480.2A patent/CN102464635B/en active Active
Patent Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| JPH02142723A (en) * | 1988-11-25 | 1990-05-31 | Tsumura & Co | Rental disorder-improving agent |
| CN1560265A (en) * | 2004-02-26 | 2005-01-05 | 广州中医药大学 | Preparation process of burduck protoglucoside of anti virus and antiptmour natural medicine |
| CN101392279A (en) * | 2008-10-21 | 2009-03-25 | 浙江工业大学 | Method for preparing fructus arctii aglycone |
| CN101736050A (en) * | 2009-12-08 | 2010-06-16 | 辽宁中医药大学 | Preparation method of arctigenin |
Non-Patent Citations (2)
| Title |
|---|
| 刘世名 等: "牛蒡叶中微量木脂体牛蒡子甙和牛蒡子甙元的分离与鉴定", 《色谱》 * |
| 许润春 等: "HPLC梯度洗脱同时测定牛蒡子液体制剂中牛蒡子苷和牛蒡子苷元的含量", 《现代生物医学进展》 * |
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN105769964A (en) * | 2016-03-30 | 2016-07-20 | 苏州勤浩药物研究开发有限公司 | Application of fructus arctii extract in resistance of skin photoage |
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