CN1329406C - Method of preparing cerebroside compound from flax root - Google Patents
Method of preparing cerebroside compound from flax root Download PDFInfo
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- CN1329406C CN1329406C CNB200510068282XA CN200510068282A CN1329406C CN 1329406 C CN1329406 C CN 1329406C CN B200510068282X A CNB200510068282X A CN B200510068282XA CN 200510068282 A CN200510068282 A CN 200510068282A CN 1329406 C CN1329406 C CN 1329406C
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Abstract
The present invention discloses a new method for preparing cerebroside compounds, particularly a method for preparing the cerebroside compound beta of daucosterol, a compound 1 and a compound 2 from linen through extraction, separation and recrystallization.
Description
Technical field
Invention relates to a kind of new method for preparing cerebroside compound, promptly extracts cerebroside compound from flax.
Background technology
Flax (Linum usitatissimum L.) is the flax family (Linaceae) linum, blazons all parts of the country.This plant is an annual herb, and stem is bright and clean level and smooth, plant height 0.75～1.20m.There is branch on the top, is umbrella shape; The cane cross section is rounded, is greyish-green; Flax leaf greyish-green, alternate, the edge is the wire lanceolar, and leaf is intensive, have an even surface, long 2～3.5cm, wide 2～4cm, leaf base are gradually narrow, are blue-greenish colour; Spend that scattered bennet is upright on the bennet top, long 2～7mm, raceme or open cyme; 5 pieces of sepals, long and narrow or long avette, long 5～7mm, full edge, no nectary, the edge indentation, total sepal length is 1/2 of petal; Petal blueness, white or incarnadine are about more than the 1cm, are obovate; It is linear that column cap is, and extends downward the inboard of style; Fruit is the capsule with Room 5, ovoid, and 7～10mm is long, is surrounded by Su Cunyue, and pericarp does not divide; Seed is no more than 10 in each fruit, and is yellow or light brown to Vandyke brown, glossy, differs in size, and dry granular weighs 3～12g.Seed is flat, and is avette, an end slyness, and the other end is sharp-pointed.
Flax is a kind of good weaving and oil crops, has multiple medicinal function again.In textile industry, though compare with cotton, wool, flax fiber flexibility and elasticity deficiency, powerful high, brute force is only second to ramie in crudefiber crop.Flax fiber can be spun into high grade cotton yarn and be made into exquisite fabric with special textile technology.Flex seed contains the protein of 20%-30%, with its oleum lini of can squeezing out.80% oleum lini is used for producing paint.Behind the linseed oil expressed oil, residue can be pressed into linseed meal, and linseed meal still contains the oleum lini of 3%-6%.Because protein content height, linseed meal are considered to a kind of fabulous animal feed.Aspect medicinal, mainly use its seed, have relax bowel, the effect of nourishing blood to expel wind, be used for the treatment of the dry constipation of intestines, allergic dermatitis, skin pruritus etc.
Although document record flax root is also pharmaceutically acceptable, aspect relevant scientific research and flax root chemical constitution study, do not see any formal research report so far.
Cerebroside is the lipoid that contains sugar, lipid acid and sphingosine, and it is constructed as follows:
Cerebroside mainly is present in animal kingdom, and distribution is also arranged in the plant.Cerebronic monomer component is white amorphous pellets; Studies show that, most cerebroside compounds have the important physical activity, as antitumor, immunomodulatory, anti-HIV-1 virus, anti-liver poison, antiulcer agent, and effects such as skin care, hair care, because cerebroside membrane structure composition, therefore its active function is generally comparatively gentle, is expected to be developed as medicine [Sang Yishu, the Min Zhida of some chronic diseases of control, the cerebroside compound progress, China's biochemical drug magazine, 2000,21 (4): 211-213].
Summary of the invention
The object of the present invention is to provide the cerebronic method of a kind of new preparation.
For realizing purpose of the present invention, the present invention adopts following technical scheme:
Comprise the steps:
(a) flax (Linum usitatissimum L.) solubilizing agent extracts, and filters, and concentrates to obtain crude extract;
(b) crude extract water dissolution with low polar organic solvent extraction, keeps water layer;
(c) go up the water layer that the step obtains, use organic solvent extraction, keep organic solvent layer, carry thing in concentrated;
(d) total extract separates with silicagel column, increases progressively by polarity with elutriant and carries out wash-out, obtains containing the flow point of cerebroside compound;
(e) merge the flow point that contains cerebroside compound, concentrate, get cerebroside compound with pure crystallization.
Particularly,
Raw material: flax (Linum usitatissimum L.) plant, described flax plant comprises from flex seed, seed germination is divided the whole process of flax growths such as seedling stage, fir shape phase, fast growing period, existing thunder flowering period, ripening stage, preferably flax plant in ripening stage.Ripening stage is divided into green ripe stage, yellow maturity initial stage, the stage of yellow ripeness, full ripe stage four-stage, more preferably flax plant in yellow maturity stage again.Described flax plant comprises the various pieces such as root, stem, leaf, flower, fruit and seed of flax, also comprises the former stem of flax, and the stem bits behind the flax straw extraction fiber; Preferred flax plant is flax underground part, i.e. the root of flax.
Raw material can the appropriateness drying.
For improving the efficient of extracting; Flax raw material can suitably be pulverized, and is beneficial to increase the contact area with solvent, raises the efficiency.
Extract:
The extraction solvent of flax raw material is selected from the mixture of water, alcohols, other organic solvents commonly used or water and alcohols.Preferred solvent is the mixture of water and alcohols, for example contains the water of pure 1-95% (volume ratio).Preferred alcohols comprises methyl alcohol, ethanol, Virahol, butanols etc.Organic solvent commonly used comprises vinyl acetic monomer, chloroform, sherwood oil etc.
The effect of extracting for guaranteeing, use during extraction quantity of solvent be former medicine weight 3-14 doubly; Each amount of extracting solvent that adds will there be medicinal raw material.Extraction can be carried out under static state or dynamic condition, preferably carries out under dynamic condition.In order to improve the efficient of extraction, can use ultrasonic wave etc.The temperature of extracting be from room temperature (for example 20 ℃) to the scope of solvent refluxing temperature in, preferably under the temperature of backflow.Extraction can be carried out continuously or intermittently, can repeat 1-4 time when intermittently extracting.
After extracting end, filter, use the ordinary skill in the art to reclaim solvent and get crude extract.
The crude extract water dissolution.Then with the organic solvent extraction of rudimentary property to remove the rudimentary property composition in the crude extract, preferred rudimentary property organic solvent comprises chloroform, benzene, hexanaphthene, sherwood oil, solvent oil, alkyl alcohol, vinyl acetic monomer; Vinyl acetic monomer more preferably.The 1-6 that used rudimentary property organic solvent amount can be the crude extract alcoholic solution doubly.The number of times of extraction is 2-5 time, preferably 3-4 time.Discard rudimentary property organic solvent part after extraction is finished for the first time, keep water layer.
Obtain the aqueous solution and use organic solvent extraction again, keep organic solvent layer, discard water layer to remove water soluble component.Organic solvent preferably uses propyl carbinol.Can extract 1-6 time, preferably 3-5 time, be more preferably 3 times.The amount of the organic solvent that each extraction is used is 0.1-5 a times of volume of water, and preferably 0.2-1.5 doubly.Merge organic solvent extraction liquid and select NaHCO for use
3Aqueous solution extraction, use H again
2The O washing is to neutral.Concentrate evaporate to dryness at last and get total extract.
The a small amount of organic solvent dissolution of total extract, methyl alcohol preferably, silica gel mixed sample with 2-13 times of weight, separate with silica gel column chromatography, separate weight with silica gel and be applied sample amount 8-80 times, preferably 10-30 times, the specification of silica gel can be column chromatography or thin-layer chromatography silica gel, preferably 200-300 order column chromatography silica gel.The elutriant of silicagel column can use polarity solvent elution from small to large, preferred elutriant is chloroform, vinyl acetic monomer or dichloromethane-ethanol or methanol mixed solvent, its volume ratio can progressively increase the content of ethanol or methyl alcohol since 50: 1, to improve polarity.When chloroform: when methyl alcohol is adjusted to 25: 1 by volume, be the wash-out flow point that wash-out obtains containing The compounds of this invention soon.
Reclaim the eluting solvent of each wash-out stream part respectively, and with behind the dissolve with methanol, place, stream part of containing The compounds of this invention promptly has crystallization to separate out, and recrystallizations such as available organic solvent such as methyl alcohol, gets cerebroside compound, is the unformed powder of white.Can certainly use the outflow of additive method detection The compounds of this invention, for example: nuclear magnetic resonance technique etc.
Also can merge wash-out stream part of containing cerebroside compound, concentrate.Resistates is placed with small amount of methanol or dissolve with ethanol, and crystallization is filtered, and gets cerebroside compound.
Advantage of the present invention:
The present invention with in the Taro Aso's product process of the prior art Central Asia as being to be used as raw material as flax root of waste, flax stem bits, extract cerebroside compound with multiple physiologically active, realized the comprehensive utilization of resource.
Embodiment
The experiment of the following examples and pharmaceutical activity is used for further specifying the present invention, but this and do not mean that any limitation of the invention.
Embodiment
26kg flax root is 1 * 1cm through being ground into granularity
2Behind the order, with 6 times of amount 95% alcohol reflux 3 times (2h, 1h * 2), the leaching extracting solution merges, and is concentrated into paste, use the 4000mL dissolved in distilled water, use ethyl acetate extraction successively three times (2000mL, 1500mL * 2), n-butanol extraction three times (2000mL, 1500mL * 2); The butanol extraction liquid that obtains is concentrated into about 3000mL, then with 3 times (1500mL, 1000mL * 2) of 5% sodium bicarbonate aqueous solution extraction, water is extracted to neutrality (1000mL * 2) again, the reclaim under reduced pressure propyl carbinol gets brown n-butanol extract, is thick cerebroside extract (31g).
Get 30g and carry out the column chromatography separation with 10 times of amount 200～300 order silica gel, carry out gradient elution with trichloromethane/methyl alcohol (100: 1～5: 1) solvent system, every 250ml collects 1 part,
1H NMR detects, and merges identical flow point, gets β-daucosterol (980mg) 61-69 part; The compound warp that obtains 100-110 and 115-130 part
1HNMR detects and is respectively cerebroside compound 1 (110mg) and 2 (70mg).
Structure is identified:
Compound 1, white unformed powder, its IR spectrum signify hydroxy (3300cm
-1) and amino (1630cm
-1) absorption peak.Quasi-molecular ions 866[M+Na among the ESIMS]
+, 848[M-H
2O+Na]
+, 704[M-162+Na]
+, 686[M-180+Na]
+Show that its molecular weight is 843, with document [Wu Tong, Kong Deyun, Li Huiting. the structure of two new plant brain glycosides is identified in the longan of dark green mountain, Acta Pharmaceutica Sinica, 2004,39 (7): 525-527] known compound 3 unanimities of report.Compound 1
1H NMR spectrum and 3 closely similar (tables 1), main difference is 1
1In the H NMR spectrum, ethylene linkage proton signal δ 5.57 (1H, dt, J=15.5,6.0Hz, H-8), 5.46 (1H, dt, J=15.5,6.5Hz, coupling constant H-9) is trans coupling.964cm in the IR spectrum
-1Absorption also show the existence of trans double bond.1
13C is also very similar to 3 with DEPT NMR spectrum, except that a plurality of methylene signals in δ 22.9-33.5 scope, also shows 1 stronger methyl resonance signal, the methylene signals of 2 company's oxygen, 11 methyne resonance signals and 1 quaternary carbon signal.1 methanolic acid hydrolysis obtains the longer chain fatty acid methyl esters, the quasi-molecular ions 398[M among its EIMS]
+, 339[M-COOCH
3]
+Be indicated as tetracosa carbon acid methyl esters.In conjunction with
1H-
1H COSY, TOCSY, HSQC, the analysis of HMBC spectrum, the structure of conclusive evidence compound 1 is as follows.
Table 1
1HNMR and
13C NMR data of compound 1(500MHz，C
5D
5N)
| Carbon | δ H | δ C | 1H- 1H COSY | HMBC |
| 1 2 3 4 5 6 7 8 9 10 11～15 16 17 18 1′ 2′ 3′ 4′ 5′～21′ 22′ 23′ 24 1″ 2″ 3″ 4″ 5″ 6″ NH | 4.66dd(11.0，6.5) 4.48dd(11.6，4.5) 5.23m 4.24.dd(6.5.4.5) ～4.15m 2.23，m 1.85m 1.75m 1.99m 5.57dt(15.5，6.0) 5.46dt(15.5，6.5) 2.05m 1.23～1.33m 1.23-1.33m 1.23-1.33 0.87t(6.5) 4.54dt(6.5，2.5) 1.95m 2.15m 1.69m 1.23～1.33m 1.23～133m 1.23～133m 0.87t(6.5) 4.91d(8.0) 3.96t(8.0) 4.15m 4.15m 3.82m 4.45，dd(11.5，2.0) 4.30dd(11.5，5.0) 8.15d(9.5) | 70.4 51.7 75.8 72.4 34.0 26.6 33.0a 130.6b 130.8b 33.3a 29.5～30.0 32.1 23.0 14.3 175.6 72.4 35.5 25.8 29.5～30.0 32.1 23.0 14.3 105.5 75.1 78.4 71.4 78.5 62.6 | 2-H 2-H 1-H，3-H 2-H，4-H 3-H，5-H 4-H 8-H 7-H，9-H 8-H，10-H 3′-H 2′-H 2″-H 1″-H，3″-H 2″-H，4″-H 3″-H，5″-H 4″-H，6″-H 5″-H 5″-H 2-H | 1-C 3-C，1-C，1′-C 1-C，2-C，4-C，5-C 3-C，6-C 7-C，9-C，10-C 7-C，8-C，10-C 16-C，17-C 3′-C，1′-C 22′-C，23′-C 1-C，3″-C，5″-C 3″-C，1″-C 5″-C 6″-C 1″-C，6″-C，4″-C 5″-C，4″-C |
1：8E
3：8Z
The structure of compound 1,3
Compound 2, white unformed powder, ESIMS m/z:736[M+Na]
+,
1H NMR (400MHz, C
5D
5N) δ: 0.87 (6H, t, J=5.6Hz, CH
3* 2), 3.90 (1H, m, H-5 "), 4.02 (1H, t, J=8.0Hz; H-2 "), 4.21 (2H, m, H-1a, 3 ", 4 "), 4.35 (1H, dd, J=12.0,5.6Hz, H-6 " a), 4.51 (1H, dd; J=12.0,2.4Hz, H-6 " b), 4.57 (1H, dd, J=7.6,3.6Hz, H-2 '), 4.71 (1H, dd, J=10.4,6.0Hz, H-1b), 4.75 (1H, t, J=6.4Hz, H-3), 4.81 (1H, m, H-2), 4.92 (1H, d, J=7.6Hz, H-1 "); 5.48 (2H, m, H-8,9), 5.89 (1H, dt; J=15.2,6.0Hz, H-5), 5.99 (1H, dd, J=15.2; 5.6Hz, H-4), 8.35 (1H, d, J=8.0Hz, NH);
13C NMR (100MHz, C
5D
5N) δ: 69.9 (C-1), 54.3 (C-2), 72.4 (C-3), 132.1 (C-4), 131.8 (C-5), 32.9 (C-6), 32.8 (C-7), 131.0 (C-8), 129.8 (C-9), 32.6 (C-10), 175.7 (C-1 '), 72.1 (C-2 '), (35.5 C-3 '), 14.2 (C-18,16 '), 105.4 (C-1 "), 75.0 (C-2 "), 78.4 (C-3 "); 71.4 (C-4 "), 78.3 (C-5 "), 62.5 (C-6 ").Its methanolic acid hydrolysis obtains longer chain fatty acid methyl esters, EIMSm/z:286[M]
+, 227[M-COOCH
3]
+, be indicated as 16 methyl carbonates.Above data and document [.Shibuya H, Kawashima K, Sakagami M, et al.Sphingolipids andglycerolipids.I.Chemical structures and ionophoretic activities ofsoyacerebrosides I and II from Soybean, Chem Pharm Bull, the structure of 1990,38 (11): 2933-2938.] unanimity, so authenticating compound 2 is as follows.
The structure of compound 2
Claims (3)
1, a kind of preparation method of cerebroside compound is characterized in that, comprises the steps:
(a) flax root (Linum usitatissimum L.) solubilizing agent extracts, and filters, and concentrates to obtain crude extract;
(b) crude extract water dissolution with low polar organic solvent extraction, keeps water layer;
(c) go up the water layer that the step obtains, use organic solvent extraction, keep organic solvent layer, merge organic solvent extraction liquid and select NaHCO for use
3Aqueous solution extraction, use H again
2O washing is to neutral, concentrate total extract;
(d) total extract separates with silicagel column, increases progressively by polarity with elutriant and carries out wash-out, obtains containing the flow point of cerebroside compound;
(e) merge the flow point that contains cerebroside compound, concentrate, get cerebroside compound with pure crystallization;
And the solvent described in the step (a) is selected from water, alcohol or both mixtures; Organic solvent described in the step (b) comprises chloroform, benzene, hexanaphthene, sherwood oil, solvent oil, alkyl alcohol, vinyl acetic monomer;
Organic solvent described in the step (c) is a propyl carbinol;
Described silica gel weight in the step (d) be applied sample amount 8-80 doubly, the specification of silica gel is the 200-300 order; Elutriant described in the step (d) is chloroform, vinyl acetic monomer or dichloromethane-ethanol or methanol mixed solvent, and its volume ratio can progressively increase methyl alcohol or alcoholic acid content since 50: 1;
Alcohol described in the step (e) is selected from methyl alcohol or ethanol.
According to the preparation method of claim 1, it is characterized in that 2, described cerebroside compound is selected from β-daucosterol, compound 1, compound 2
Compound 1
Compound 2
According to the preparation method of claim 1, it is characterized in that 3, described alcohol comprises methyl alcohol, ethanol, Virahol, butanols.
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|---|---|---|---|
| CNB200510068282XA CN1329406C (en) | 2005-05-08 | 2005-05-08 | Method of preparing cerebroside compound from flax root |
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|---|---|---|---|
| CNB200510068282XA CN1329406C (en) | 2005-05-08 | 2005-05-08 | Method of preparing cerebroside compound from flax root |
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| CN1329406C true CN1329406C (en) | 2007-08-01 |
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Families Citing this family (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101703515B (en) * | 2009-11-23 | 2011-12-21 | 南京医科大学 | Application of cerebroside compound |
| CN101735885B (en) * | 2009-12-14 | 2012-07-04 | 山东省中医药研究院 | Method for extracting shelled cedar seed oil and daucosterol from shelled cedar seed |
| CN101885747B (en) * | 2010-06-12 | 2013-03-13 | 暨南大学 | Separation and purification method for cerebroside and ceramide type compound |
| CN103087119B (en) * | 2013-01-22 | 2015-08-05 | 李桂凤 | One grows flax Cerebrosides compound and extraction and separation method thereof |
| CN105218604B (en) * | 2015-10-19 | 2018-06-29 | 浙江大学 | The method that subcritical abstraction obtains natural glycolipid |
| CN114437150A (en) * | 2022-01-20 | 2022-05-06 | 辽宁省海洋水产科学研究院 | Method for separating and extracting stichopus japonicus cerebroside from stichopus japonicus |
Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1317477A (en) * | 2000-04-13 | 2001-10-17 | 胡德甫 | Process for preparing high-purity alpha-linolenic acid |
| CN1394865A (en) * | 2001-07-11 | 2003-02-05 | 北京万博力科技发展有限公司 | Method for extracting, separating and purifying linolenic lignan by using linseed cake |
-
2005
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Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1317477A (en) * | 2000-04-13 | 2001-10-17 | 胡德甫 | Process for preparing high-purity alpha-linolenic acid |
| CN1394865A (en) * | 2001-07-11 | 2003-02-05 | 北京万博力科技发展有限公司 | Method for extracting, separating and purifying linolenic lignan by using linseed cake |
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