CN1537867A - FB1 polyose and preparaton method and application - Google Patents
FB1 polyose and preparaton method and application Download PDFInfo
- Publication number
- CN1537867A CN1537867A CNA03116403XA CN03116403A CN1537867A CN 1537867 A CN1537867 A CN 1537867A CN A03116403X A CNA03116403X A CN A03116403XA CN 03116403 A CN03116403 A CN 03116403A CN 1537867 A CN1537867 A CN 1537867A
- Authority
- CN
- China
- Prior art keywords
- polysaccharide
- water
- glossy ganoderma
- precipitation
- centrifugal
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Granted
Links
Landscapes
- Medicines Containing Plant Substances (AREA)
- Polysaccharides And Polysaccharide Derivatives (AREA)
- Pharmaceuticals Containing Other Organic And Inorganic Compounds (AREA)
Abstract
A FB1 polyose extracted from ganoderma spore and its extracting process are disclosed. Said polyose has the immunological enhancement action, so it can be used as the medicine for the auxiliary therapy of hypoimmunity diseases, such as tumor.
Description
Technical field
The present invention relates to chemical process extracting effective components from glossy ganoderma, more particularly from Ganoderma spore, extract polysaccharide, determine that chemical structure reaches the immunologic enhancement to body.
Background technology
Cancerous disease constantly threatens human health, it has become one of human dead principal element, can overcome cancer therapy drug Cancerous disease, that side effect is little though wish to search out, not find yet so far that specificity is strong, good effect, the little cancer therapy drug of side effect through people's big quantity research.
Though the tumour medicine kind of using is a lot of at present, most of chemical synthesis process that adopts makes, and toxicity, side effect is bigger.Have some antineoplastic natural products to come out in recent years, but side effect is also bigger, and majority is a raw product.In the nineties later stage, biotherapy has become the important channel of oncotherapy, and immunopotentiating agent is a class medicine that is used for tumor biotherapy.The invention provides the polysaccharide that a kind of immunomodulator promptly obtains from Ganoderma spore.
Glossy ganoderma is a kind of good invigorant in traditional medicine, and its main active ingredient is polysaccharide and Ganodenic acid.Over past ten years glossy ganoderma particularly Ganoderma spore become the focus of domestic and international research.The present invention extracts from Ganoderma spore and obtains homogeneous polysaccharide, determined chemical structure through chemical spectrum analysis, and prove that by pharmacological testing the polysaccharide of this chemical structure has significant immunologic enhancement, can be used as healthcare products and use, especially as the adjuvant therapy medicaments of tumour patient.
Summary of the invention
The object of the invention provides a kind of definite compound of polysaccharide of its structure with immunologic enhancement that extracts from Ganoderma spore.
Another object of the present invention is the extracting method of this polysaccharide of preparation and its medicinal use.
The present invention implements through the following steps:
The Ganoderma spore powder of broken wall soaks several weeks in methyl alcohol or ethanolic soln, take out air-dry, air-dry residue was used the boiling water refluxing extraction 4-5 hour again, treated coldly, filtered, residue is used the same ratio water extraction once again, merge water extraction filtrate twice, filtrate decompression is concentrated into small volume, adds 15% trichoroacetic acid(TCA) in 4 ℃ of following deproteinated by 1: 1 volume ratio, centrifugal, supernatant liquor is neutralized to neutrality with 1mol/L NaOH.To flowing water dialysis three days.See through liquid and be evaporated to 1/5 volume, adding 95% ethanol alcohol concn to the solution is 45-55%, spends the night centrifugation, dry Crude polysaccharides.
Crude polysaccharides is dissolved in distilled water, the centrifugal insolubles that goes, and supernatant liquor carries out the DEAE-Cellulose column chromatography.Water elution to sugar reaction is negative, dialysis, lyophilize gets solid, solid is water-soluble, adds ethanol alcohol concn to the solution and is 25-35%, and is centrifugal, supernatant liquor adds ethanol again makes that alcohol concn is 45-55% in the solution, centrifugal must the precipitation precipitated water-solublely, carries out Sephacryl S-300 column chromatography.0.2mol/LNaCl wash-out.Differential detects, and polysaccharide is partly dialysed, and is evaporated to small volume, presses similarity condition Sephacryl S-300 column chromatography more once, dialysis, concentrating under reduced pressure, lyophilize.This sample detects through HPLC, and the result is a single symmetrical peak, proves that this polysaccharide FB1 is pure product.Can be used as the usefulness of pharmacological testing and structural analysis.
It is 7.2 * 10 that FB1 detects its molecular weight through HPLC
5Da.Specific optical rotation is [a]
D=-23.37 ° of (C0.860, H
2O).The FB1 warp
13CNMR measures, and the anomeric carbon zone only shows that one is positioned at the carbon signal of δ 104.69, and the glucosyl residue that this polysaccharide is described is that pyrans is ring-like, and glucosides is strong that a kind of β-type mode of connection is only arranged, and this supposition with infrared spectra is consistent.In addition, exist
13The bigger signal that is positioned at δ 86.5 of intensity appears in the CNMR spectrum, contain in the sugar chain of prompting FB1 a large amount of 1, the glucopyranose residue that 3-connects; The existence of signal δ 71.7 (should belong to C-6) for the side chain glucosyl residue, prompting polysaccharide FB1 one has the ramose polymkeric substance, and one of tapping point should be positioned at the C-6 position of main chain glucosyl residue.Other signals in the carbon spectrum, δ 77.7,75.4,70.3 and 62.9 can belong to glucosyl group C-5, C-2, C-4 (main chain glucosyl group and C-6 respectively.The above results is in conjunction with methylation reaction, and periodate oxidation, results such as Smith degraded, and the repeating structure that proves FB1 is one to contain 6 glucosyl residues, has the β-1 of a terminal glucose on 6 of a glucosyl residue, the 3-dextran.Detailed structure is as shown in the figure:
Pharmacological testing:
1, mouse spleen lymphocyte proliferation activity: accurately take by weighing FB1 1-2 gram, be dissolved in physiological saline, concentration is 4mg/ml.Being diluted to concentration during use is 10
-3, 10
-2With 10
-1Mg/ml.Mouse causes death with cervical vertebra dislocation, and aseptic condition takes out spleen and separating Morr. cell down, and with nylon net filter, distilled water destroys red corpuscle, be adjusted to wait to ooze, and after nutrient solution washing three times, counting, the adjusting cell concn is 5 * 10
6Cell/ml.In 96 orifice plates, every hole 100ul establishes a control group with above-mentioned enchylema application of sample, adds testing sample and the ConA (5ug/L) or the LPS (10ug/L) of different concns again, places 37 ℃ of 5%CO
2Incubator in cultivated 44 hours, add 20ulMTT and continue to cultivate 4 hours.Cultivate termination, in every hole, add 100ul three liquid (dimethyl formamide and sodium laurylsulfonate), and spend the night in 37 ℃ of placements.Next day, measure the absorbancy of every hole solution in the 570nm place.Make T with the mouse spleen cell then, the test of bone-marrow-derived lymphocyte proliferation activity.The result shows T, bone-marrow-derived lymphocyte proliferation activity, P value difference＜0.01 and 0.001 when concentration 10,100ug/ml.Reach significantly.
2, immunity test in the mouse body:
Qualified ICR mouse, difference abdominal injection 25ug/ml and 50ug/ml FB1, control group injecting normal saline 0.2ml, administration every day secondary, administration is 9 days altogether.The 3rd day sensitization of mouse administration, every immune parameter is measured in dissection in the 5th day.The result shows that this polysaccharide FB1 its P value under above-mentioned two concentration reaches significantly respectively less than 0.01 and 0.001.
Vivo and vitro test according to above-mentioned pharmacological testing shows that FB1 can be used as immunoregulatory drug use.
Specific implementation method
Get 1Kg broken wall red ganoderma spore powder, add 3L 95% alcohol immersion in three weeks.Change ethanol weekly once, take out centrifugal.Solids places the ventilation airing, uses boiling water extraction then, adds deionized water 4L at every turn, extracts twice altogether, merges filtrate twice, and filtrate decompression is concentrated into 1L, is cooled to 4 ℃, adds 15% trichoroacetic acid(TCA) 1L.Finish, leave standstill 3 hours, centrifugal, get supernatant liquor, supernatant liquor was neutralized to pH6-7 with 1mol/LNaOH, to flowing water dialysis three days.Liquid is concentrated into 0.5L in the dialysis tubing.Adding 95% ethanol to the alcohol concn of solution is 50%, 4 ℃ of following standing over night, centrifugal must the precipitation.Precipitation after dehydrated alcohol and acetone wash successively, in 40 ℃ of vacuum-dryings, solids crude polysaccharide 9.7g.(yield 0.97%).
Get Crude polysaccharides 5g, be dissolved in the distilled water, the centrifugal insolubles that goes, supernatant liquor carry out DEAE-Cellulose column chromatography (post 50 * 10cm), water wash-out.Merge the positive reaction part according to sugared color reaction, to distill water dialysis two days.See through liquid and be evaporated to small volume, lyophilize gets cotton shape solid 2.1g.Solid is water-soluble, adds ethanol alcohol concn to the solution and is 30%, and is centrifugal, get supernatant liquor, add ethanol alcohol concn to the solution in the supernatant liquor again and be 50%, centrifugal, must precipitate 1.2g, precipitate water-solublely, carry out Sephacryl S-300 post (100 * 2.6cm) chromatographies.0.2mol/L NaCl wash-out.Differential detects, and merges the polysaccharide part, and dialysis concentrates, and lyophilize gets solid.And then carry out Sephacryl S-300 column chromatography more once by above-mentioned condition, dialysis concentrates, lyophilize, FB1.Detect through HPLC, the result is a single symmetrical peak, proves that the FB1 polysaccharide is the homogeneous component.Can be used as the usefulness of physico-chemical property, chemical structure and pharmacological testing.
Physical and chemical property determining: press the ordinary method of polysaccharide determination, determining molecular weight is 7.2 * 10
5Specific optical rotation is [a]
D=-23.37 °.
Determination of chemical structure: press the determination of chemical structure method of polysaccharide, comprise spectrographic technique (
13CNMR, 1R) and chemical process (all-hydrolytic, methylation reaction, periodate oxidation, Smith degraded, partial hydrolysis) prove that FB1 one contains 6 glucosyl residues, and be to have a branch on 5 an of glucosyl residue of main chain at five glucosyl residues, branch into the β-1 of terminal glucose, the 3-dextran.
Pharmacological testing: get above-mentioned FB1 and carry out the vivo and vitro immunity test by the pharmacological testing method of mentioning in the test respectively.The result shows T, and in the proliferation test of bone-marrow-derived lymphocyte, concentration is 10, the in vitro tests of 100ug/ml can reach significantly and highly significant.In vivo test can reach respectively significantly and highly significant when 25ug/Kg and 50ug/Kg.
Claims (6)
1, a kind of polysaccharide that obtains following structure that from glossy ganoderma, extracts.
2, obtain polysaccharide according to described extraction of claim 1 from glossy ganoderma, it is characterized in that being β-1, the 3-dextran contains 6 glucosyl residues in the repeating structure, and one of them residue is that 6 ramose have a terminal glucose residue, and molecular weight is 7.2 * 10
5, specific optical rotation is [a]
D=-23.37 ° of (C0.860, H
2O).
3, from glossy ganoderma, extract the method that obtains polysaccharide according to claim 1, it is characterized in that comprising
The following step:
(1), breaking trachytectum of glossy ganoderma alcohol degreasing, filtration, air-dry, boiling water extraction, be evaporated to small volume, add rare trichoroacetic acid(TCA) low temperature deproteinated, clear liquid neutralizes, dialyses with diluted alkaline on the Deproteinization, dialyzate is not evaporated to small volume, add ethanol to solution, precipitation, washing get raw sugar.
(2), Crude polysaccharides is water-soluble, is the carrier column chromatography with DEAE-Cellulose, water is moving phase, elutriant dialysis, concentrating under reduced pressure, lyophilize get solid.
(3), solids is water-soluble, add ethanol, centrifugal, go precipitation, supernatant liquor adds ethanol again, centrifugal must the precipitation.
(4), above-mentioned precipitation is water-soluble, gel filtration chromatography, moving phase is water, effluent liquid is through dialysis, lyophilize gets homogeneous polysaccharide.
4, according to the described method that from glossy ganoderma, extract to obtain polysaccharide of claim 3, it is characterized in that seeing through liquid after the dialysis is concentrated into small volume, add ethanol and make that alcohol concn is 45-55% in the solution.
5, according to the described method that obtains polysaccharide of from glossy ganoderma, extracting of claim 3, it is characterized in that raw sugar is through the DEAE-Cellulose chromatography, elutriant gets solids after concentrating, solids is water-soluble again, add ethanol and make that alcohol concn is 25-35% in the solution, the centrifugal precipitation of going, supernatant liquor adds ethanol again makes that alcohol concn is 45-55% in the solution, centrifugal must the precipitation.
6, according to the described method that from glossy ganoderma, extract to obtain polysaccharide of claim 3, it is characterized in that the water-soluble after secondary Sephacryl S-300 column chromatography of above-mentioned throw out, polysaccharide is partly dialysed, concentrating under reduced pressure, lyophilize gets homogeneous polysaccharide.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CNB03116403XA CN100335504C (en) | 2003-04-14 | 2003-04-14 | FB1 polyose and preparaton method and application |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CNB03116403XA CN100335504C (en) | 2003-04-14 | 2003-04-14 | FB1 polyose and preparaton method and application |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN1537867A true CN1537867A (en) | 2004-10-20 |
| CN100335504C CN100335504C (en) | 2007-09-05 |
Family
ID=34320337
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CNB03116403XA Expired - Fee Related CN100335504C (en) | 2003-04-14 | 2003-04-14 | FB1 polyose and preparaton method and application |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN100335504C (en) |
Cited By (12)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN100335505C (en) * | 2005-03-07 | 2007-09-05 | 敖宗华 | Process for preparing antrodia camphorata polysaccharide and antrodia camphorata triterpene with micro-prorous adsorptive resin and its product |
| CN100424098C (en) * | 2006-10-25 | 2008-10-08 | 南京中科集团股份有限公司 | Process for refining glossy ganoderma spore polysaccharide |
| CN102020721A (en) * | 2011-01-13 | 2011-04-20 | 哈尔滨工业大学 | Preparation method of polysaccharide in pine cone from Pinus koraiensis |
| CN102040748A (en) * | 2009-10-16 | 2011-05-04 | 上海医药工业研究院 | Ganoderma sinensis mycelium anti-tumor polysaccharide component GS-C as well as preparation method and application thereof |
| CN101423855B (en) * | 2008-10-28 | 2011-10-26 | 山东好当家海洋发展股份有限公司 | Method for preparing polysaccharide by using lucidum strain fermented laminaria leftover |
| CN104059162A (en) * | 2014-06-25 | 2014-09-24 | 上海市农业科学院 | Ganoderma lucidum polysaccharide and preparation method thereof |
| CN105175575A (en) * | 2015-10-30 | 2015-12-23 | 上海市农业科学院 | Ganoderma lucidum beta-glucan and preparing method and application thereof |
| CN106832042A (en) * | 2017-02-03 | 2017-06-13 | 中国科学院上海药物研究所 | The glucans of β 1,3, its preparation method and pharmaceutical applications |
| CN108159089A (en) * | 2018-03-14 | 2018-06-15 | 广东省微生物研究所（广东省微生物分析检测中心） | Purposes of the lucidum spore powder dietary fiber extract in the preparation for preparing treatment and/or prevention high fat diet cause hepatic injury relevant disease |
| CN108159090A (en) * | 2018-03-14 | 2018-06-15 | 广东省微生物研究所（广东省微生物分析检测中心） | Purposes of the lucidum spore powder dietary fiber extract in treatment and/or prevention intestinal bacilli illness relevant disease preparation is prepared |
| CN110423284A (en) * | 2019-08-14 | 2019-11-08 | 浙江万寿康药业有限公司 | Lucid ganoderma spore powder polysaccharide has effects that auxiliary inhibits the application in the functional food of colon cancer in preparation |
| CN112094358A (en) * | 2020-09-22 | 2020-12-18 | 广东省微生物研究所（广东省微生物分析检测中心） | Tibetan ganoderma lucidum polysaccharide GLP-1 with antioxidant effect, and preparation method and application thereof |
Family Cites Families (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| KR100311317B1 (en) * | 1999-07-16 | 2001-10-18 | 이신영 | Exo-polysaccharide production from submerged mycelial culture of Ganoderma lucidum |
| CN1101855C (en) * | 2000-03-10 | 2003-02-19 | 华东理工大学 | Liquid fermentation process for preparing both ganoderic polyose and ganoderic acid |
| KR100398088B1 (en) * | 2000-08-28 | 2003-09-19 | 주식회사 엠바이오텍 | Mass production of exo-polysaccharide from submerged cultivation of Ganoderma lucidum by agitation and aeration effect under bi-staged pH controlling system of jar fermenter |
| CN1141392C (en) * | 2001-03-20 | 2004-03-10 | 华东理工大学 | Process for preparing ganoderic polyose and ganoderic acid by fermentation during which raw materials are supplemented |
-
2003
- 2003-04-14 CN CNB03116403XA patent/CN100335504C/en not_active Expired - Fee Related
Cited By (15)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN100335505C (en) * | 2005-03-07 | 2007-09-05 | 敖宗华 | Process for preparing antrodia camphorata polysaccharide and antrodia camphorata triterpene with micro-prorous adsorptive resin and its product |
| CN100424098C (en) * | 2006-10-25 | 2008-10-08 | 南京中科集团股份有限公司 | Process for refining glossy ganoderma spore polysaccharide |
| CN101423855B (en) * | 2008-10-28 | 2011-10-26 | 山东好当家海洋发展股份有限公司 | Method for preparing polysaccharide by using lucidum strain fermented laminaria leftover |
| CN102040748A (en) * | 2009-10-16 | 2011-05-04 | 上海医药工业研究院 | Ganoderma sinensis mycelium anti-tumor polysaccharide component GS-C as well as preparation method and application thereof |
| CN102020721A (en) * | 2011-01-13 | 2011-04-20 | 哈尔滨工业大学 | Preparation method of polysaccharide in pine cone from Pinus koraiensis |
| CN104059162B (en) * | 2014-06-25 | 2016-03-23 | 上海市农业科学院 | A kind of ganoderan and preparation method thereof |
| CN104059162A (en) * | 2014-06-25 | 2014-09-24 | 上海市农业科学院 | Ganoderma lucidum polysaccharide and preparation method thereof |
| CN105175575A (en) * | 2015-10-30 | 2015-12-23 | 上海市农业科学院 | Ganoderma lucidum beta-glucan and preparing method and application thereof |
| CN106832042A (en) * | 2017-02-03 | 2017-06-13 | 中国科学院上海药物研究所 | The glucans of β 1,3, its preparation method and pharmaceutical applications |
| CN106832042B (en) * | 2017-02-03 | 2019-05-17 | 中国科学院上海药物研究所 | Beta-1,3-dextran, preparation method and pharmaceutical applications |
| CN108159089A (en) * | 2018-03-14 | 2018-06-15 | 广东省微生物研究所（广东省微生物分析检测中心） | Purposes of the lucidum spore powder dietary fiber extract in the preparation for preparing treatment and/or prevention high fat diet cause hepatic injury relevant disease |
| CN108159090A (en) * | 2018-03-14 | 2018-06-15 | 广东省微生物研究所（广东省微生物分析检测中心） | Purposes of the lucidum spore powder dietary fiber extract in treatment and/or prevention intestinal bacilli illness relevant disease preparation is prepared |
| CN110423284A (en) * | 2019-08-14 | 2019-11-08 | 浙江万寿康药业有限公司 | Lucid ganoderma spore powder polysaccharide has effects that auxiliary inhibits the application in the functional food of colon cancer in preparation |
| CN112094358A (en) * | 2020-09-22 | 2020-12-18 | 广东省微生物研究所（广东省微生物分析检测中心） | Tibetan ganoderma lucidum polysaccharide GLP-1 with antioxidant effect, and preparation method and application thereof |
| CN112094358B (en) * | 2020-09-22 | 2022-05-27 | 广东省微生物研究所（广东省微生物分析检测中心） | Tibetan ganoderma lucidum polysaccharide GLP-1 with antioxidant effect, and preparation method and application thereof |
Also Published As
| Publication number | Publication date |
|---|---|
| CN100335504C (en) | 2007-09-05 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN100335504C (en) | FB1 polyose and preparaton method and application | |
| Sone et al. | Structures and antitumor activities of the polysaccharides isolated from fruiting body and the growing culture of mycelium of Ganoderma lucidum | |
| CN102336840B (en) | Triple-helical Tremellan, preparation method and application thereof | |
| CA2412600A1 (en) | Potent immunostimulatory polysaccharides extracted from microalgae | |
| CN114591448B (en) | Mulberry and phellinus linteus fruiting body manna galactosan and preparation and application thereof | |
| CN1069650C (en) | Method for separation and purification of lentinan | |
| CN103382229B (en) | A kind of preparation method and Structural Identification with the novel SEP-1 of immunoregulation effect | |
| CN110540603A (en) | Rhizoma anemarrhenae polysaccharide, and preparation method, identification method and application thereof | |
| CN110256593A (en) | A kind of Stropharia rugoso-annulata polysaccharide and its preparation method and application | |
| EP0441278A1 (en) | Polysaccharides with immunomodulating properties from astragalus membranaceus and pharmaceutical compositions containing them | |
| KR100871399B1 (en) | Endo-polymer and exo-polymer obtained from submerged mycelial culture of Cordyceps militaris increasing immuno-modulating activities and optimum culture conditions for producing the same | |
| CN113717296B (en) | Eucommia acidic polysaccharide, extraction method and application of eucommia acidic polysaccharide in preparation of anti-colon cancer drugs | |
| CN110218264A (en) | A kind of Lyophyllum decastes granulose and its preparation method and application | |
| Ingólfsdóttir | Bioactive compounds from Iceland moss | |
| CN1165148A (en) | Method for preparing dictyophora fungus polysaccharide | |
| KR20110014820A (en) | Polysaccharide extracted from capsosiphon fulvescens having anti-coagulation and immunostimulating effect, composition containing polysaccharide and process thereof | |
| CN109232753A (en) | A kind of Boletus speciosus polysaccharide BSF-X and the preparation method and application thereof | |
| Wen et al. | A new acidic polysaccharide and its sulfated derivative from cultured Morchella sextelata fruiting bodies and their antioxidant and immunoregulatory activities | |
| KR100397793B1 (en) | Novel Microorganism Isolated from Chinese elm(Ulmus sp.) and Process for Preparing Immunostimulating Exopolysaccharides with Anti-cancer Activity by Employing the Microorganism | |
| KR100348870B1 (en) | Peptido-glyco Compounds from Chinese elm and Process for Preparing the Same | |
| CN100406473C (en) | Preparation method and application of Asiatic polysaccharide | |
| KR100729213B1 (en) | Exo-biopolymer isolated from submerged mycelial culture of Grifola frondosa increasing immune activity | |
| KR100893533B1 (en) | A novel Phellinus strain and the method for isolating ?-1,3 glucomannan therefrom showing immune enhancing activity and antitumor activity | |
| CN1049221C (en) | Ginseng oligosaccharide element and ginseng monomer oligosaccharide and its preparing technology and use | |
| EP0939082A1 (en) | Protein polysaccharide 0041 |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| C06 | Publication | ||
| PB01 | Publication | ||
| C10 | Entry into substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| C14 | Grant of patent or utility model | ||
| GR01 | Patent grant | ||
| CF01 | Termination of patent right due to non-payment of annual fee | ||
| CF01 | Termination of patent right due to non-payment of annual fee |
Granted publication date: 20070905 Termination date: 20200414 |