CN104017102B - Ethanol precipitation prepares the method for Sulodexide raw material from heparin byproduct - Google Patents
Ethanol precipitation prepares the method for Sulodexide raw material from heparin byproduct Download PDFInfo
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- CN104017102B CN104017102B CN201410274835.6A CN201410274835A CN104017102B CN 104017102 B CN104017102 B CN 104017102B CN 201410274835 A CN201410274835 A CN 201410274835A CN 104017102 B CN104017102 B CN 104017102B
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Abstract
The invention discloses a kind of method preparing Sulodexide raw material from heparin byproduct separation, heparin byproduct is dissolved, with sodium hydrate regulator solution to high ph-values, does low-temperature sludge with ethanol in proper amount, after going precipitation, obtain sulfated polysaccharide solution; Adjust this sulfated polysaccharide solution to low ph value with hydrochloric acid, do low-temperature sludge with ethanol in proper amount, after going precipitation, obtain sulfated polysaccharide solution; Regulate this solution neutral to pH with sodium hydroxide again, do precipitation at room temperature with ethanol in proper amount, get precipitation and dry the Sulodexide raw material obtaining obtaining.Technique of the present invention is simple, cost is low, easy amplification, be suitable for suitability for industrialized production.
Description
Technical field
The present invention relates to a kind of method preparing Sulodexide raw material, particularly relate to a kind of method being separated Sulodexide raw material from heparin byproduct.
Background technology
Sulodexide Sulodexide, commodity are called big plain VESSELDUEF, and chemical name is glucuronyl glucosaminoglycan vitriol.By the original production of Italian AlfaWassermann company, within 1974, first go on the market in Italy, within 2003, enter Chinese market.Indication is the vascular disease having thrombosis danger, be a kind of can the anticoagulant of oral absorption.In recent years it is shown effective to the treatment of diabetic nephropathy.
Sulodexide is extracted by pig intestinal mucosa, is highly purified glycosaminoglycan, forms the long-chain macromolecule with negative charge by the disaccharides structural unit repeated, and disaccharide unit comprises two and changes structure sugar, namely
n-acetylgalactosamine or
n-acetylglucosamine and uronic acid, as glucuronic acid or iduronic acid.Agargel electrophoresis records the quick travel heparin (fastmovingheparin containing 80%, FMH) (based on electrophoretic mobility) i.e. Low molecular heparin (lowmolecularweightheparin, dermatan sulfate (the dermatansulfate of LMWH) and 20%, DS) and on a small quantity heparin (slowmovingheparin, SMH) (must not 2% be greater than) is moved at a slow speed.LMWH part is wherein the same as with unfraction heparin disaccharides paradigmatic structure, but degree is lower, and the length of polysaccharide chain is shorter.The polysaccharide that the DS that do not degrade is made up of many different disaccharides, average molecular mass 25000Da.
Heparin production process can produce a large amount of by product, wherein containing chondroitin sulfate (chondroitinsulfate, CS), dermatan sulfate, fast mobile heparin, slow mobile heparin etc., is be separated the good material preparing Sulodexide raw material.
Patent " a kind of method (application number 201210064002.8) being separated Sulodexide raw material from heparin byproduct " gives a kind of method being separated Sulodexide raw material from heparin byproduct, adopt the step such as Potassium ethanoate precipitation, Ban Shi agent precipitate, alcohol settling, anion exchange process, from heparin byproduct, isolate Sulodexide raw material.
This patent provides the another kind of method being separated Sulodexide raw material from heparin byproduct, under adopting high pH, low pH, middle pH condition respectively, different invalid components is separated removing by ethanol precipitation, thus from heparin byproduct, isolates Sulodexide raw material quickly and easily.
Summary of the invention
Technical problem to be solved by this invention is to provide a kind of method being separated Sulodexide raw material from heparin byproduct.
In the process of producing heparin, produce a large amount of by product, wherein main composition is chondroitin sulfate, dermatan sulfate, fast mobile heparin, slow mobile heparin etc.Establish a kind of new separation purifying technique in the present invention, isolate Sulodexide raw material from heparin byproduct, the present invention includes following steps:
(1) heparin byproduct is dissolved as 5-15%(m/m) solution, with sodium hydroxide regulate pH to 10.0-11.5, adding ethanol to mass concentration is 29-33%, 2-8 DEG C precipitation 1-2 hour.On collected by centrifugation, liquid obtains the sulfated polysaccharide solution after removing slow heparin.
(2) add water in the sulfated polysaccharide solution through step (1) purifying, make alcohol concn be 24%, with salt acid for adjusting pH to 1.5-2.5, adding ethanol to mass concentration is 38-42%, at 2-8 DEG C of precipitation 1-2 hour.On collected by centrifugation, liquid obtains the sulfated polysaccharide solution after dermatan sulfate decrement.
(3) add water in the solution obtained to step (2), make alcohol concn be 36%, regulate pH to 5.5-8.5, reinforcing body NaCl to be 2% to mass concentration with sodium hydroxide, then to add ethanol to mass concentration be 40-50%, 8-10 DEG C of precipitates overnight.Move away supernatant, obtain precipitation.
(4) precipitation that obtains of step (3) is with 60-75 DEG C of oven dry, is Sulodexide medicine material.
Heparin byproduct strength of solution described in step (1) is preferably 8%-12%, and more preferably 10%, pH value of solution is preferably 11.0, and during precipitation, alcohol concn is preferably 31%; Step (2) pH value of solution is preferably 2.0, and during precipitation, alcohol concn is preferably 40%, (3) pH value of solution is preferably 7.0, and during precipitation, alcohol concn is preferably 45%.Step (1), (2), (3) use ethanol to do organic solvent deposit, but in like manner also can use methyl alcohol, acetone does organic solvent deposit, only parameter need be finely tuned.
Preparation method of the present invention, has that with low cost, technique is simple, the easy feature such as amplification.Detect by the Sulodexide medicine material agarose electrophoresis obtained by the present invention and find that it consists of DS20-30%, fast heparin 70-80%, meet the requirement of Sulodexide drug component.
Accompanying drawing explanation
Fig. 1 is raw material, first time precipitates after product, second time precipitates after product, precipitate after product (end product) agarose gel electrophoretogram for the third time.
Embodiment
Below in conjunction with example, content of the present invention is further elaborated.
Embodiment one
By heparin byproduct (containing CS5%, DS50%, fast heparin 25%, slow heparin 20%, tire 41uspu/mg, specific rotation+15 °) be dissolved as 10%(m/m) solution, with 6M sodium hydroxide regulate pH to 11.0, adding ethanol to mass concentration is 31%, precipitates 2 hours at 2-8 DEG C.The centrifugal 30min of 4000rpm, in collection, liquid obtains the sulfated polysaccharide solution after removing slow heparin.
Add water in the sulfated polysaccharide solution obtained, make alcohol concn be 24%, with 6M salt acid for adjusting pH to 2.0, adding ethanol to mass concentration is 40%, at 2-8 DEG C of precipitation 1-2 hour.On collected by centrifugation, liquid obtains the sulfated polysaccharide solution after dermatan sulfate decrement.
Add water in the solution obtained, make alcohol concn be 36%, regulate pH to 7.0, reinforcing body NaCl to be 2% to mass concentration with 6M sodium hydroxide, then to add ethanol to mass concentration be 50%, 8-10 DEG C of precipitates overnight.Remove supernatant, obtain precipitation.
Precipitation, in an oven with 70 DEG C of oven dry, is Sulodexide medicine material (tire 69uspu/mg, specific rotation+35 ° for DS20%, fast heparin 80%).
Embodiment two
The raw material of Example one, first time precipitates after product, second time precipitates after product, third time precipitates after product (end product) etc. and do agarose gel electrophoresis analysis, and method is with reference to patent " a kind of glass carries the method 200910040788.8 that electrophoretic method analyzes mucopolysaccharide ".Result as shown in Figure 1, M represents heparin byproduct raw material, containing CS, DS, fast heparin, slow heparin in raw material, precipitate through first time and eliminate slow heparin, through second time precipitation, DS content is reduced, through third time precipitation removal CS, obtains final product: DS20-30%, fast heparin 70-80%.
Claims (6)
1. be separated the method preparing Sulodexide raw material from heparin byproduct, its feature comprises the following steps:
(1) heparin byproduct is dissolved as the solution that mass concentration is 5-15%, pH to 10.0-11.5 is regulated with sodium hydroxide solution, adding ethanol to mass concentration is 29-33%, and at 2-8 DEG C of precipitation 1-2 hour, on collected by centrifugation, liquid obtains the sulfated polysaccharide solution after removing slow heparin;
(2) in the sulfated polysaccharide solution through step (1) purifying, purified water is added, alcohol concn is made to reduce to 24%, with salt acid for adjusting pH to 1.5-2.5, adding ethanol to mass concentration is 38-42%, at 2-8 DEG C of precipitation 1-2 hour, on collected by centrifugation, liquid obtains the sulfated polysaccharide solution after dermatan sulfate decrement;
(3) add purified water in the solution obtained to step (2), make alcohol concn be 36%, regulate pH to 5.5-8.5 with sodium hydroxide solution, reinforcing body NaCl is 2% to mass concentration, then to add ethanol to mass concentration be 40-50%, and precipitation at room temperature spends the night, move away supernatant, obtain precipitation;
(4) precipitation that obtains of step (3) is with 60-75 DEG C of oven dry, is Sulodexide medicine material.
2. the method for claim 1, is characterized in that: described in step (1), heparin byproduct is dissolved as the solution of 8-12% massfraction, and regulate pH to 11.0 with sodium hydroxide, during precipitation, in system, ethanol mass concentration is 31%.
3. the method for claim 1, is characterized in that: step (2) is described with salt acid for adjusting pH to 2.0, and adding ethanol to mass concentration is 40%.
4. the method for claim 1, is characterized in that: step (3) is described regulates pH to 7.0 with sodium hydroxide, and adding ethanol to mass concentration is 45%.
5. the method for claim 1, is characterized in that: step (1), (2), (3) are respectively at alkalescence, acid, neutral environment execution alcohol settling.
6. the method for claim 1, is characterized in that: use methyl alcohol or acetone to replace the ethanol in step (1), (2), (3) to do organic solvent deposit.
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| Publication number | Priority date | Publication date | Assignee | Title |
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| CN107991414A (en) * | 2017-12-28 | 2018-05-04 | 山东大学 | A kind of electrophoresis hydrophilic interaction combined gas chromatography mass spectrometry detection method of Sulodexide |
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| CN104398533B (en) * | 2014-11-26 | 2017-03-08 | 山东辰中生物制药有限公司 | The production method of Sulodexide |
| CN106883321B (en) * | 2017-03-06 | 2019-03-19 | 河北常山生化药业股份有限公司 | A method of extracting Sulodexide bulk pharmaceutical chemicals from heparin sodium by-product |
| CN108484789A (en) * | 2018-05-03 | 2018-09-04 | 东营天东制药有限公司 | A kind of preparation method of Sulodexide bulk pharmaceutical chemicals |
| CN113735994A (en) * | 2020-05-29 | 2021-12-03 | 江苏唯高生物科技有限公司 | Process for preparing sulodexide raw material |
| CN114478832B (en) * | 2022-02-14 | 2023-03-03 | 河北常山生化药业股份有限公司 | Method for purifying four polysaccharide products from heparin sodium by-product |
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| Publication number | Priority date | Publication date | Assignee | Title |
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| US20030008844A1 (en) * | 2001-05-17 | 2003-01-09 | Keryx Biopharmaceuticals, Inc. | Use of sulodexide for the treatment of inflammatory bowel disease |
| JP2005139409A (en) * | 2003-11-10 | 2005-06-02 | Medicaraise Corp | Method for producing dermatan sulfate |
| WO2005058235A2 (en) * | 2003-12-10 | 2005-06-30 | Keryx Biopharmaceuticals, Inc. | Methods using sulodexide for the treatment of bladder disease |
| JP2008540448A (en) * | 2005-05-05 | 2008-11-20 | ケリク ビオファルマセウチカルス インコーポレーテッド | Combination therapy with sulodexide and antihypertensive agent in the treatment of diabetic nephropathy |
| CN100384884C (en) * | 2006-05-29 | 2008-04-30 | 南京健友生物化学制药有限公司 | Method for separating and purify dermatansulfate and low-molecular heparan sulfate from sodium heparan product |
| CN101885787B (en) * | 2009-05-11 | 2014-07-09 | 深圳市海普瑞药业股份有限公司 | Method for purifying heparitin sulfate from heparin byproduct |
| CN101885782B (en) * | 2009-05-11 | 2012-07-04 | 深圳市海普瑞药业股份有限公司 | Method for purifying dermatan sulfate from heparin byproduct |
| CN102603924A (en) * | 2012-03-12 | 2012-07-25 | 南京健友生化制药股份有限公司 | Method for separating sulodexide raw materials from heparin by-products |
| CN103044577B (en) * | 2012-12-07 | 2015-05-06 | 青岛九龙生物医药有限公司 | Method for reducing dermatan sulfate content in heparin sodium product |
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| CN107991414A (en) * | 2017-12-28 | 2018-05-04 | 山东大学 | A kind of electrophoresis hydrophilic interaction combined gas chromatography mass spectrometry detection method of Sulodexide |
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Address after: 518057 Guangdong city of Shenzhen province Nanshan District Song Ping Lang Road No. 21 Patentee after: Shenzhen Hepalink Pharmaceutical Group Limited by Share Ltd Address before: 518057 Guangdong city of Shenzhen province Nanshan District Song Ping Lang Road No. 21 Patentee before: Shenzhen Hepalink Pharmaceutical Co., Ltd. |
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