CN111057165A - Preparation method of high-purity heparin sodium - Google Patents
Preparation method of high-purity heparin sodium Download PDFInfo
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- CN111057165A CN111057165A CN202010030423.3A CN202010030423A CN111057165A CN 111057165 A CN111057165 A CN 111057165A CN 202010030423 A CN202010030423 A CN 202010030423A CN 111057165 A CN111057165 A CN 111057165A
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- C—CHEMISTRY; METALLURGY
- C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
- C08B—POLYSACCHARIDES; DERIVATIVES THEREOF
- C08B37/00—Preparation of polysaccharides not provided for in groups C08B1/00 - C08B35/00; Derivatives thereof
- C08B37/006—Heteroglycans, i.e. polysaccharides having more than one sugar residue in the main chain in either alternating or less regular sequence; Gellans; Succinoglycans; Arabinogalactans; Tragacanth or gum tragacanth or traganth from Astragalus; Gum Karaya from Sterculia urens; Gum Ghatti from Anogeissus latifolia; Derivatives thereof
- C08B37/0063—Glycosaminoglycans or mucopolysaccharides, e.g. keratan sulfate; Derivatives thereof, e.g. fucoidan
- C08B37/0075—Heparin; Heparan sulfate; Derivatives thereof, e.g. heparosan; Purification or extraction methods thereof
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Abstract
The invention discloses a preparation method of high-purity heparin sodium, which comprises the following steps: preparing a crude heparin sodium solution; stirring and standing the crude heparin sodium solution for 30-60min, filtering after standing, and precipitating out impurity precipitates to obtain pre-configured solution; placing the pre-configured solution into a reaction container, adding protease into the reaction container, carrying out enzymolysis on the pre-configured solution in the reaction container, and obtaining an enzymolysis configured solution after enzymolysis; heating the enzymolysis preparation solution, cooling the enzymolysis preparation solution to 35-45 ℃ after heating, adding a sodium hydroxide solution to adjust the pH, and obtaining a regulating solution after adjusting the pH; adding an adsorbing material into the regulating solution, stirring, placing the regulating solution into a centrifugal device for high-speed centrifugation after stirring, and collecting clear liquid after centrifugation; oxidizing the clear liquid to obtain an oxidized liquid; and (3) carrying out precipitation treatment on the oxidation liquid, collecting precipitates after the precipitation treatment, and dehydrating the precipitates to obtain the high-purity heparin sodium. The preparation method has high efficiency, and the obtained heparin sodium has high purity.
Description
Technical Field
The invention relates to the technical field of preparation of related heparin sodium, in particular to a preparation method of high-purity heparin sodium.
Background
Heparin sodium is the most widely used anticoagulant in clinical medicine, and is a mucopolysaccharide sulfate consisting of D-glucosamine, N-acetylglucosamine, D-glucuronic acid and L-iduronic acid, which is mainly present in mast cell granules in animal viscera, and is mostly present in combination with proteins as protein-heparin complexes, and is mostly present in small intestinal mucosa and lung. Heparin sodium is mainly used for anticoagulation and is one of the most important anticoagulation and antithrombotic biochemical drugs at present, but a medicament prepared by taking heparin sodium as a raw material is usually administrated by an injection mode, so the heparin sodium has high purity to ensure medical safety.
The existing heparin sodium preparation method generally uses a heparin sodium crude product as a raw material to prepare high-purity heparin sodium, but the preparation process is easy to leave a certain amount of impurities such as small molecular proteins in the heparin sodium finished product, so that the protein content of the final heparin sodium is relatively high, and the quality of the heparin sodium is influenced.
Disclosure of Invention
The invention aims to provide a preparation method of high-purity heparin sodium, which aims to solve the problems in the background technology.
In order to achieve the purpose, the invention provides the following technical scheme:
a preparation method of high-purity heparin sodium comprises the following steps:
the method comprises the following steps: preparing a crude heparin sodium solution;
step two: stirring and standing the crude heparin sodium solution for 30-60min, filtering after standing, and precipitating out impurity precipitates to obtain pre-configured solution;
step three: placing the pre-configured solution into a reaction container, adding protease into the reaction container, carrying out enzymolysis on the pre-configured solution in the reaction container for 4-8h at the reaction temperature of 30-40 ℃, and obtaining an enzymolysis configured solution after enzymolysis;
step four: heating the enzymolysis preparation solution to 80-90 ℃, keeping for 20-40min, cooling the enzymolysis preparation solution to 35-45 ℃ after heating, adding a sodium hydroxide solution to adjust the pH, adjusting the pH to 7-10, and obtaining an adjusting solution after adjusting the pH;
step five: adding an adsorbing material into the regulating solution, stirring, placing the regulating solution into a centrifugal device for high-speed centrifugation after stirring, removing insoluble impurities, and collecting clear liquid after centrifugation;
step six: oxidizing the clear liquid collected in the fifth step to obtain an oxidized liquid;
step seven: and (3) carrying out precipitation treatment on the oxidation liquid, collecting precipitates after the precipitation treatment, dehydrating the precipitates, and carrying out vacuum drying and sterilization to obtain the high-purity heparin sodium.
As a preferred technical scheme of the invention, the preparation of the crude heparin sodium solution in the first step specifically comprises the following steps: step a: preparing a sodium chloride solution, wherein the sodium chloride solution is prepared by mixing and dissolving edible sodium chloride and water according to the proportion of 3: 5-13; step b: taking crude heparin sodium, mixing and dissolving the crude heparin sodium and a sodium chloride solution to prepare a crude heparin sodium solution, wherein the ratio of the crude heparin sodium to the sodium chloride solution is 1: 5-15.
As a preferable technical solution of the present invention, the oxidation treatment in the sixth step includes: adding hydrogen peroxide solution accounting for 2-5% of the total mass of the clear liquid into the clear liquid, stirring, adjusting the pH value to 8.5-9.5, and then centrifuging at high speed and low temperature to obtain the oxidation liquid.
As a preferred technical solution of the present invention, the precipitation treatment in the seventh step includes: adding absolute ethyl alcohol which accounts for 40-70% of the total mass of the oxidizing solution into the oxidizing solution, adjusting the temperature to 0-6 ℃, adding a sodium chloride solution after adjustment, stirring, filtering by using a filter membrane, and filtering to obtain a precipitate.
As a preferred technical scheme of the invention, the adsorption material in the fourth step is nano-alumina, nano-kaolinite or nano-montmorillonite.
Compared with the prior art, the invention has the beneficial effects that:
the invention adopts crude heparin sodium as raw material to prepare high-purity heparin sodium, has low preparation cost and high preparation method efficiency, and the obtained heparin sodium has high purity and less impurities, adopts the adsorption material to adsorb protein in the preparation process, is beneficial to protein precipitation, and can better remove protein components in liquid by matching with centrifugal treatment.
Detailed Description
The technical solutions in the embodiments of the present invention are clearly and completely described below, and it is obvious that the described embodiments are only a part of the embodiments of the present invention, and not all embodiments. All other embodiments, which can be derived by a person skilled in the art from the embodiments given herein without making any creative effort, shall fall within the protection scope of the present invention.
Example 1: the invention provides a technical scheme that: a preparation method of high-purity heparin sodium comprises the following steps:
the method comprises the following steps: preparing a crude heparin sodium solution;
step two: stirring and standing the crude heparin sodium solution for 30min, filtering after standing, and precipitating out impurity precipitates to obtain pre-configured solution;
step three: placing the pre-configured solution into a reaction container, adding protease into the reaction container, carrying out enzymolysis on the pre-configured solution in the reaction container for 4 hours at the reaction temperature of 35 ℃, and obtaining an enzymolysis configured solution after enzymolysis;
step four: heating the enzymolysis preparation solution to 80 ℃, keeping the temperature for 30min, cooling the enzymolysis preparation solution to 35 ℃ after heating, adding a sodium hydroxide solution to adjust the pH, adjusting the pH to 8, and obtaining a regulating solution after adjusting the pH;
step five: adding an adsorbing material into the regulating solution, stirring, placing the regulating solution into a centrifugal device for high-speed centrifugation after stirring, removing insoluble impurities, and collecting clear liquid after centrifugation;
step six: oxidizing the clear liquid collected in the fifth step to obtain an oxidized liquid;
step seven: and (3) carrying out precipitation treatment on the oxidation liquid, collecting precipitates after the precipitation treatment, dehydrating the precipitates, and carrying out vacuum drying and sterilization to obtain the high-purity heparin sodium.
In this embodiment, the preparation of the crude heparin sodium solution in the first step specifically includes the following steps: step a: preparing a sodium chloride solution, wherein the sodium chloride solution is prepared by mixing and dissolving edible sodium chloride and water according to the proportion of 3: 8; step b: taking crude heparin sodium, mixing and dissolving the crude heparin sodium and a sodium chloride solution to prepare a crude heparin sodium solution, wherein the ratio of the crude heparin sodium to the sodium chloride solution is 1: 7.
In this embodiment, the oxidation treatment in the sixth step includes: adding hydrogen peroxide solution accounting for 3% of the total mass of the clear liquid into the clear liquid, stirring, adjusting the pH value to 8.5, and then centrifuging at high speed and low temperature to obtain the oxidation liquid.
In this embodiment, the precipitation treatment in step seven includes: adding anhydrous ethanol accounting for 40% of the total mass of the oxidizing solution into the oxidizing solution, adjusting the temperature to 0-6 ℃, adding a sodium chloride solution after adjustment, stirring, filtering by using a filter membrane, and obtaining a precipitate after filtering.
In this example, the adsorbent in step four is nano-montmorillonite.
Example 2:
the method comprises the following steps: preparing a crude heparin sodium solution;
step two: stirring and standing the crude heparin sodium solution for 40min, filtering after standing, and precipitating out impurity precipitates to obtain pre-configured solution;
step three: placing the pre-configured solution into a reaction container, adding protease into the reaction container, carrying out enzymolysis on the pre-configured solution in the reaction container for 6 hours at the reaction temperature of 30 ℃, and obtaining an enzymolysis configured solution after enzymolysis;
step four: heating the enzymolysis preparation solution to 90 ℃, keeping the temperature for 20min, cooling the enzymolysis preparation solution to 45 ℃ after heating, adding a sodium hydroxide solution to adjust the pH, adjusting the pH to 10, and obtaining an adjusting solution after adjusting the pH;
step five: adding an adsorbing material into the regulating solution, stirring, placing the regulating solution into a centrifugal device for high-speed centrifugation after stirring, removing insoluble impurities, and collecting clear liquid after centrifugation;
step six: oxidizing the clear liquid collected in the fifth step to obtain an oxidized liquid;
step seven: and (3) carrying out precipitation treatment on the oxidation liquid, collecting precipitates after the precipitation treatment, dehydrating the precipitates, and carrying out vacuum drying and sterilization to obtain the high-purity heparin sodium.
In this embodiment, the preparation of the crude heparin sodium solution in the first step specifically includes the following steps: step a: preparing a sodium chloride solution, wherein the sodium chloride solution is prepared by mixing and dissolving edible sodium chloride and water according to the proportion of 3: 7; step b: taking crude heparin sodium, mixing and dissolving the crude heparin sodium and a sodium chloride solution to prepare a crude heparin sodium solution, wherein the ratio of the crude heparin sodium to the sodium chloride solution is 1: 9.
In this embodiment, the oxidation treatment in the sixth step includes: adding hydrogen peroxide solution accounting for 2-5% of the total mass of the clear liquid into the clear liquid, stirring, adjusting the pH value to 9, and then centrifuging at high speed and low temperature to obtain the oxidation liquid.
In this embodiment, the precipitation treatment in step seven includes: adding anhydrous ethanol accounting for 55% of the total mass of the oxidizing solution into the oxidizing solution, adjusting the temperature to 0-6 ℃, adding a sodium chloride solution after adjustment, stirring, filtering by using a filter membrane, and obtaining a precipitate after filtering.
In this embodiment, the adsorbing material in the fourth step is nano kaolinite.
Example 3:
the method comprises the following steps: preparing a crude heparin sodium solution;
step two: stirring and standing the crude heparin sodium solution for 30min, filtering after standing, and precipitating out impurity precipitates to obtain pre-configured solution;
step three: placing the pre-configured solution into a reaction container, adding protease into the reaction container, carrying out enzymolysis on the pre-configured solution in the reaction container for 4 hours at the reaction temperature of 40 ℃, and obtaining an enzymolysis configured solution after enzymolysis;
step four: heating the enzymolysis preparation solution to 80 ℃, keeping the temperature for 40min, cooling the enzymolysis preparation solution to 35 ℃ after heating, adding a sodium hydroxide solution to adjust the pH, adjusting the pH to 10, and obtaining an adjusting solution after adjusting the pH;
step five: adding an adsorbing material into the regulating solution, stirring, placing the regulating solution into a centrifugal device for high-speed centrifugation after stirring, removing insoluble impurities, and collecting clear liquid after centrifugation;
step six: oxidizing the clear liquid collected in the fifth step to obtain an oxidized liquid;
step seven: and (3) carrying out precipitation treatment on the oxidation liquid, collecting precipitates after the precipitation treatment, dehydrating the precipitates, and carrying out vacuum drying and sterilization to obtain the high-purity heparin sodium.
In this embodiment, the preparation of the crude heparin sodium solution in the first step specifically includes the following steps: step a: preparing a sodium chloride solution, wherein the sodium chloride solution is prepared by mixing and dissolving edible sodium chloride and water according to the proportion of 3: 8; step b: taking crude heparin sodium, mixing and dissolving the crude heparin sodium and a sodium chloride solution to prepare a crude heparin sodium solution, wherein the ratio of the crude heparin sodium to the sodium chloride solution is 1: 10.
In this embodiment, the oxidation treatment in the sixth step includes: adding a hydrogen peroxide solution accounting for 4% of the total mass of the clear liquid into the clear liquid, stirring, adjusting the pH value to 8.5, and then centrifuging at high speed and low temperature to obtain an oxidation liquid.
In this embodiment, the precipitation treatment in step seven includes: adding anhydrous ethanol accounting for 45% of the total mass of the oxidizing solution into the oxidizing solution, adjusting the temperature to 0-6 ℃, adding a sodium chloride solution after adjustment, stirring, filtering by using a filter membrane, and obtaining a precipitate after filtration.
In this example, the adsorbent in step four is nano-montmorillonite.
The invention adopts crude heparin sodium as raw material to prepare high-purity heparin sodium, has low preparation cost and high preparation method efficiency, and the obtained heparin sodium has high purity and less impurities, adopts the adsorption material to adsorb protein in the preparation process, is beneficial to protein precipitation, and can better remove protein components in liquid by matching with centrifugal treatment.
Although embodiments of the present invention have been shown and described, it will be appreciated by those skilled in the art that changes, modifications, substitutions and alterations can be made in these embodiments without departing from the principles and spirit of the invention, the scope of which is defined in the appended claims and their equivalents.
Claims (5)
1. The preparation method of the high-purity heparin sodium is characterized by comprising the following steps of:
the method comprises the following steps: preparing a crude heparin sodium solution;
step two: stirring and standing the crude heparin sodium solution for 30-60min, filtering after standing, and precipitating out impurity precipitates to obtain pre-configured solution;
step three: placing the pre-configured solution into a reaction container, adding protease into the reaction container, carrying out enzymolysis on the pre-configured solution in the reaction container for 4-8h at the reaction temperature of 30-40 ℃, and obtaining an enzymolysis configured solution after enzymolysis;
step four: heating the enzymolysis preparation solution to 80-90 ℃, keeping for 20-40min, cooling the enzymolysis preparation solution to 35-45 ℃ after heating, adding a sodium hydroxide solution to adjust the pH, adjusting the pH to 7-10, and obtaining an adjusting solution after adjusting the pH;
step five: adding an adsorbing material into the regulating solution, stirring, placing the regulating solution into a centrifugal device for high-speed centrifugation after stirring, removing insoluble impurities, and collecting clear liquid after centrifugation;
step six: oxidizing the clear liquid collected in the fifth step to obtain an oxidized liquid;
step seven: and (3) carrying out precipitation treatment on the oxidation liquid, collecting precipitates after the precipitation treatment, dehydrating the precipitates, and carrying out vacuum drying and sterilization to obtain the high-purity heparin sodium.
2. The method for preparing high-purity heparin sodium according to claim 1, wherein the preparation of the crude heparin sodium solution in the first step specifically comprises the following steps: step a: preparing a sodium chloride solution, wherein the sodium chloride solution is prepared by mixing and dissolving edible sodium chloride and water according to the proportion of 3: 5-13; step b: taking crude heparin sodium, mixing and dissolving the crude heparin sodium and a sodium chloride solution to prepare a crude heparin sodium solution, wherein the ratio of the crude heparin sodium to the sodium chloride solution is 1: 5-15.
3. The method for preparing high-purity heparin sodium according to claim 1, wherein the oxidation treatment in the sixth step comprises: adding hydrogen peroxide solution accounting for 2-5% of the total mass of the clear liquid into the clear liquid, stirring, adjusting the pH value to 8.5-9.5, and then centrifuging at high speed and low temperature to obtain the oxidation liquid.
4. The method for preparing high-purity heparin sodium according to claim 1, wherein the precipitation treatment in the seventh step comprises: adding absolute ethyl alcohol which accounts for 40-70% of the total mass of the oxidizing solution into the oxidizing solution, adjusting the temperature to 0-6 ℃, adding a sodium chloride solution after adjustment, stirring, filtering by using a filter membrane, and filtering to obtain a precipitate.
5. The method for preparing high-purity heparin sodium according to claim 1, wherein the adsorbing material in the fourth step is nano-alumina, nano-kaolinite or nano-montmorillonite.
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Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1566162A (en) * | 2003-07-07 | 2005-01-19 | 张国良 | Heparin sodium and its preparing process |
| US20080269134A1 (en) * | 1999-04-05 | 2008-10-30 | Emisphere Technologies, Inc. | Disodium salts, monohydrates, and ethanol solvates for delivering active agents |
| CN102993336A (en) * | 2011-09-14 | 2013-03-27 | 浦江亚太肠衣有限公司 | Crude heparin sodium purification technology |
| CN108456262A (en) * | 2018-03-13 | 2018-08-28 | 广元市海天实业有限责任公司 | A kind of preparation process of high purity heparin sodium |
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2020
- 2020-01-13 CN CN202010030423.3A patent/CN111057165A/en active Pending
Patent Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US20080269134A1 (en) * | 1999-04-05 | 2008-10-30 | Emisphere Technologies, Inc. | Disodium salts, monohydrates, and ethanol solvates for delivering active agents |
| CN1566162A (en) * | 2003-07-07 | 2005-01-19 | 张国良 | Heparin sodium and its preparing process |
| CN102993336A (en) * | 2011-09-14 | 2013-03-27 | 浦江亚太肠衣有限公司 | Crude heparin sodium purification technology |
| CN108456262A (en) * | 2018-03-13 | 2018-08-28 | 广元市海天实业有限责任公司 | A kind of preparation process of high purity heparin sodium |
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