CN115475258A - Purification process of tenecteplase for injection - Google Patents
Purification process of tenecteplase for injection Download PDFInfo
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- CN115475258A CN115475258A CN202211084472.0A CN202211084472A CN115475258A CN 115475258 A CN115475258 A CN 115475258A CN 202211084472 A CN202211084472 A CN 202211084472A CN 115475258 A CN115475258 A CN 115475258A
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
- A61L2/00—Methods or apparatus for disinfecting or sterilising materials or objects other than foodstuffs or contact lenses; Accessories therefor
- A61L2/0005—Methods or apparatus for disinfecting or sterilising materials or objects other than foodstuffs or contact lenses; Accessories therefor for pharmaceuticals, biologicals or living parts
- A61L2/0011—Methods or apparatus for disinfecting or sterilising materials or objects other than foodstuffs or contact lenses; Accessories therefor for pharmaceuticals, biologicals or living parts using physical methods
- A61L2/0017—Filtration
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
- A61L2/00—Methods or apparatus for disinfecting or sterilising materials or objects other than foodstuffs or contact lenses; Accessories therefor
- A61L2/0005—Methods or apparatus for disinfecting or sterilising materials or objects other than foodstuffs or contact lenses; Accessories therefor for pharmaceuticals, biologicals or living parts
- A61L2/0082—Methods or apparatus for disinfecting or sterilising materials or objects other than foodstuffs or contact lenses; Accessories therefor for pharmaceuticals, biologicals or living parts using chemical substances
- A61L2/0088—Liquid substances
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- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N9/00—Enzymes; Proenzymes; Compositions thereof; Processes for preparing, activating, inhibiting, separating or purifying enzymes
- C12N9/14—Hydrolases (3)
- C12N9/48—Hydrolases (3) acting on peptide bonds (3.4)
- C12N9/50—Proteinases, e.g. Endopeptidases (3.4.21-3.4.25)
- C12N9/64—Proteinases, e.g. Endopeptidases (3.4.21-3.4.25) derived from animal tissue
- C12N9/6421—Proteinases, e.g. Endopeptidases (3.4.21-3.4.25) derived from animal tissue from mammals
- C12N9/6424—Serine endopeptidases (3.4.21)
- C12N9/6456—Plasminogen activators
- C12N9/6459—Plasminogen activators t-plasminogen activator (3.4.21.68), i.e. tPA
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- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12Y—ENZYMES
- C12Y304/00—Hydrolases acting on peptide bonds, i.e. peptidases (3.4)
- C12Y304/21—Serine endopeptidases (3.4.21)
- C12Y304/21068—Tissue plasminogen activator (3.4.21.68), i.e. tPA
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61L—METHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
- A61L2202/00—Aspects relating to methods or apparatus for disinfecting or sterilising materials or objects
- A61L2202/20—Targets to be treated
- A61L2202/21—Pharmaceuticals, e.g. medicaments, artificial body parts
Abstract
The invention relates to a purification process of injection tenecteplase, which comprises the following steps: taking a pretreatment solution containing tenecteplase protein, subjecting the pretreatment solution to Blue Sepharose 6Fastflow affinity chromatography, and eluting with an eluent A to obtain a Blue chromatography target protein solution; performing Lysine HyperD affinity chromatography on the Blue chromatography target protein solution, and then eluting by using an eluent B to obtain a Lysine chromatography target protein solution; diluting the Lysine chromatography target protein solution, adjusting the pH value of the Lysine chromatography target protein solution to acidity, then carrying out virus inactivation treatment, adjusting the pH value of the Lysine chromatography target protein solution to neutrality, and filtering by using a 0.1um filter and a 20nm virus removal filter to obtain a tenecteplase protein solution; the process can achieve 1 × 10 removal/inactivation effect on viruses in the tenecteplase 10 Dose/granule, much more than 1X 10 6 The technical requirements of the preparation/granule industry effectively improve the product quality of the injection tenecteplase.
Description
Technical Field
The invention relates to the technical field of biological product production, in particular to a purification process of injection tenecteplase.
Background
The tenecteplase for injection is prepared by using recombinant Chinese Hamster Ovary (CHO) cells containing high-expression human histotypic plasminogen activator modified protein gene, performing amplification culture by using an inoculation amplification culture medium, then performing amplification by using an inoculation tank, performing cell culture and separation and purification on a tank-harvested culture medium to obtain a human histotypic plasminogen activator modified protein stock solution, and then adding a proper amount of auxiliary materials to prepare a freeze-dried powder injection which is mainly used for thrombolytic therapy.
And the possibility of exogenous virus pollution exists in the CHO cell culture process, so that virus removal and inactivation are needed in the process of separation and purification of tenecteplase, so as to ensure the safety of clinical medication. However, in the existing tenecteplase purification process, two-step chromatography processes, namely Blue Sepharose 6FastFlow affinity chromatography and lysine HyperD affinity chromatography, are usually adopted, and although the purification process can effectively remove impurities in tenecteplase, the virus removal effect is not ideal, so that a new technical scheme is needed to overcome the defects.
Disclosure of Invention
The invention aims to provide a purification process of injection tenecteplase, which has good virus removal and inactivation effects and can effectively improve the product quality of injection tenecteplase.
In order to achieve the purpose of the invention, the following technical scheme is adopted:
a purification process of injection tenecteplase comprises the following steps:
step 1: taking a pretreatment solution containing tenecteplase protein, subjecting the pretreatment solution to Blue Sepharose 6Fastflow affinity chromatography, and eluting with an eluent A to obtain a Blue chromatography target protein solution;
step 2: performing Lysine HyperD affinity chromatography on the Blue chromatography target protein solution, and eluting with an eluent B to obtain a Lysine chromatography target protein solution;
and, step 3: diluting the Lysine chromatography target protein solution, adjusting the pH value of the Lysine chromatography target protein solution to acidity, then performing virus inactivation treatment, adjusting the pH value of the Lysine chromatography target protein solution to neutrality after virus inactivation is completed, and filtering by using a 0.1um filter and a 20nm virus removal filter in sequence to obtain the tenecteplase protein solution.
Preferably, in the step 1, the binding capacity of the Blue Sepharose 6Fastflow affinity chromatography to the tenecteplase protein is 10-20mg/ml.
Preferably, in the step 1, the eluent a has the following components: 10-50mM phosphate buffer, 0.043% polysorbate 20, 0.5-1M sodium chloride, 2-4M urea, and 5-10mg/L aprotinin.
Preferably, the eluent A has a pH value of 7.2-8.0, and is used in an amount of 3-5 times the column volume.
Preferably, in the step 2, the binding capacity of the lysine HyperD affinity chromatography to the tenecteplase protein is 5-10mg/ml.
Preferably, the eluent B comprises the following components: 0.2-1.0M arginine, 0.1-0.5M aminocaproic acid, 0.043% polysorbate 20, 6-10mg/L aprotinin, and 5-18mg/mL phosphoric acid.
Preferably, the eluent B has a pH value of 7.4-7.8, and is used in an amount of 3-5 times the column volume.
Preferably, in the step 3, the Lysine chromatography target protein solution is diluted by water for injection until the content of the target protein is 0.19-0.21 mg/ml.
Preferably, in the step 3, when the virus inactivation treatment is performed, the pH value of the Lysine chromatography target protein solution is 3.3 to 3.7.
Compared with the prior art, the invention has the following beneficial effects:
compared with the existing purification process, the method adopted by the invention has the advantages that the removal/inactivation effect of the process on the virus in the tenecteplase can reach 1 x 10 10 Dose/granule, much more than 1X 10 6 The technical requirements of the agent/particle industry meet the requirement that every 100 ten thousand doses of virus particles are less than 1; effectively improves the product quality of the injection tenecteplase.
Detailed Description
In order to make the objects, technical solutions and advantages of the embodiments of the present invention clearer, the following will clearly and completely describe the technical solutions in the embodiments of the present invention, and it is obvious that the described embodiments are some embodiments of the present invention, but not all embodiments.
The invention provides a purification process of injection tenecteplase, which specifically comprises the following steps:
step 1: taking a pretreatment solution containing tenecteplase protein, subjecting the pretreatment solution to Blue Sepharose 6Fastflow affinity chromatography, and eluting with an eluent A to obtain a Blue chromatography target protein solution;
wherein the binding capacity of the Blue Sepharose 6Fastflow affinity chromatography to the tenecteplase protein is 10-20mg/ml;
specifically, the steps of the Blue Sepharose 6Fastflow affinity chromatography are as follows:
step 1a: blue Sepharose 6Fastflow affinity chromatography before and after loading equilibration: before and after the pretreatment solution is loaded, balancing the chromatographic column by using a balance buffer solution A respectively; the components of the equilibration buffer A are as follows: 10-50mM phosphate buffer solution, 0.043% polysorbate 20, 0.1-0.5M sodium chloride and 1-3mg/L aprotinin, wherein the pH value is 7.2-8.0, and the dosage is 3-5 times of the column volume;
step 1b: blue Sepharose 6FastFlow affinity chromatography impurity elution: after the sample loading is finished, impurity elution is carried out on the chromatographic column by using an impurity eluent A, and the impurity eluent A comprises the following components: 10-50mM phosphate buffer solution, 0.043% polysorbate 20, 1-3M sodium chloride and 1-3mg/L aprotinin, wherein the pH value is 7.2-8.0, and the dosage is 5-10 times of the column volume;
step 1c: blue Sepharose 6FastFlow affinity chromatography activity peak elution: after the impurity elution is finished, eluting the chromatographic column by using an eluent A, and collecting the target protein activity peak to obtain a Blue chromatographic target protein solution; the eluent A comprises the following components: 10-50mM phosphate buffer solution, 0.043% polysorbate 20, 0.5-1M sodium chloride, 2-4M urea and 5-10mg/L aprotinin, wherein the pH value is 7.2-8.0, and the dosage is 3-5 times of the column volume.
Step 2: performing Lysine HyperD affinity chromatography on the Blue chromatography target protein solution, and then eluting by using an eluent B to obtain a Lysine chromatography target protein solution; wherein the binding capacity of lysine HyperD affinity chromatography to tenecteplase protein is 5-10mg/ml;
specifically, the steps of the lysine HyperD affinity chromatography are as follows:
step 2a: blue target protein solution was diluted: before the Blue target protein solution is loaded, diluting the Blue target protein solution by using a dilution buffer solution for 5-6 times and then loading the Blue target protein solution; the composition of the dilution buffer was: 10-50mM phosphate buffer solution, 0.043% polysorbate 20 and 1-3mg/L aprotinin, wherein the pH value is 7.4-8.0;
and step 2b: equilibration before and after loading of lysine HyperD affinity chromatography: before and after the Blue target protein solution is loaded, balancing the chromatographic column by using a balance buffer solution B respectively; the components of the balance buffer solution B are as follows: 10-50mM phosphate buffer solution, 0.043% polysorbate 20 and 2mg/L aprotinin, wherein the pH value is 7.2-8.0, and the dosage is 3-5 times of the column volume;
and step 2c: lysine HyperD affinity chromatography impurity elution: after Blue target protein solution is loaded, impurity elution treatment is carried out on the chromatographic column by using impurity eluent B, and the impurity eluent B comprises the following components: 10-50mM phosphate buffer, 1M sodium chloride, 0.043% polysorbate 20 and 2mg/L aprotinin, wherein the pH value is 7.2-8.0, and the dosage is 5-15 times of the column volume.
Step 2d: lysine HyperD affinity chromatography active peak elution: after the impurity elution is finished, eluting the chromatographic column by using an eluent B, and collecting the active peak of the target protein to obtain a Lysine chromatography target protein solution; the eluent B comprises the following components: 0.2-1.0M arginine, 0.1-0.5M aminocaproic acid, 0.043% polysorbate 20, 6-10mg/L aprotinin, and 5-18mg/mL phosphoric acid; the pH value is 7.4-7.8, and the dosage is 3-5 times of the column volume.
And 3, step 3: diluting the Lysine chromatography target protein solution by using water for injection until the content of target protein is 0.19-0.21 mg/ml, then adjusting the pH value of the Lysine chromatography target protein solution to 3.3-3.7, then performing virus inactivation treatment, standing for 90 minutes at 20-24 ℃ to complete virus inactivation, then adjusting the pH value of the Lysine chromatography target protein solution to 7.2-8.0, and sequentially filtering by using a 0.1um filter and a 20nm virus removal filter to obtain the tenecteplase protein solution.
Example 1
A purification process of injection tenecteplase comprises the following steps:
step 1: taking a pretreatment solution containing tenecteplase protein, subjecting the pretreatment solution to Blue Sepharose 6Fastflow affinity chromatography, and eluting with an eluent A to obtain a Blue chromatography target protein solution;
specifically, the steps of the Blue Sepharose 6Fastflow affinity chromatography are as follows:
step 1a: blue Sepharose 6Fastflow affinity chromatography before and after loading equilibration: before and after the pretreatment solution is loaded, balancing the chromatographic column by using a balance buffer solution A respectively; the components of the equilibration buffer A are as follows: 20mM phosphate buffer solution, 0.043% polysorbate 20, 0.15M sodium chloride and 2mg/L aprotinin, wherein the pH value is 7.6, and the dosage is 3 times of the column volume;
step 1b: blue Sepharose 6FastFlow affinity chromatography impurity elution: after the sample loading is finished, impurity elution is carried out on the chromatographic column by using an impurity eluent A, and the impurity eluent A comprises the following components: 20mM phosphate buffer, 0.043% polysorbate 20, 2M sodium chloride and 2mg/L aprotinin, wherein the pH value is 7.6, and the dosage is 5 times of the column volume;
step 1c: blue Sepharose 6FastFlow affinity chromatography activity peak elution: after the impurity elution is finished, eluting the chromatographic column by using an eluent A, and collecting a target protein active peak to obtain a Blue chromatographic target protein solution; the eluent A comprises the following components: 20mM phosphate buffer, 0.043% polysorbate 20, 1M sodium chloride, 3M urea and 6mg/L aprotinin, at pH 7.6, in an amount of 3 times the column volume.
And 2, step: performing Lysine HyperD affinity chromatography on the Blue chromatography target protein solution, and then eluting by using an eluent B to obtain a Lysine chromatography target protein solution;
specifically, the steps of the lysine hyperD affinity chromatography are as follows:
step 2a: blue target protein solution was diluted: before Blue target protein solution is loaded, diluting the Blue target protein solution by 6 times by using a dilution buffer solution and then loading the Blue target protein solution; the composition of the dilution buffer was: 20mM phosphate buffer, 0.043% polysorbate 20 and 2mg/L aprotinin, at pH 7.6;
and step 2b: equilibration before and after loading of lysine HyperD affinity chromatography: before and after the Blue target protein solution is loaded, balancing the chromatographic column by using a balance buffer solution B respectively; the components of the balance buffer solution B are as follows: 20mM phosphate buffer, 0.043% polysorbate 20 and 2mg/L aprotinin, wherein the pH value is 7.6, and the dosage is 3 times of the column volume;
and step 2c: lysine HyperD affinity chromatography impurity elution: after Blue target protein solution is loaded, impurity elution treatment is carried out on the chromatographic column by using impurity eluent B, and the impurity eluent B comprises the following components: 20mM phosphate buffer, 1M sodium chloride, 0.043% polysorbate 20 and 2mg/L aprotinin, at pH 7.6, in an amount 10 times the column volume.
Step 2d: lysine HyperD affinity chromatography active peak elution: after the impurity elution is finished, eluting the chromatographic column by using an eluent B, and collecting the active peak of the target protein to obtain a Lysine chromatography target protein solution; the eluent B comprises the following components: 0.5M arginine, 0.2M aminocaproic acid, 0.043% polysorbate 20, 10mg/L aprotinin, and 8mg/mL phosphoric acid; the pH was 7.4 and the amount used was 3 times the column volume.
And step 3: diluting the Lysine chromatography target protein solution by using water for injection until the content of the target protein is 0.20mg/ml, then adjusting the pH value of the Lysine chromatography target protein solution to 3.7, then performing virus inactivation treatment, standing at 20 ℃ for 90 minutes to complete virus inactivation, then adjusting the pH value of the Lysine chromatography target protein solution to 7.6, and sequentially filtering by using a 0.1um filter and a 20nm virus removal filter to obtain the tenecteplase protein solution.
Example 2
A purification process of injection tenecteplase comprises the following steps:
step 1: taking a pretreatment solution containing tenecteplase protein, subjecting the pretreatment solution to Blue Sepharose 6Fastflow affinity chromatography, and eluting with an eluent A to obtain a Blue chromatography target protein solution;
specifically, the steps of the Blue Sepharose 6Fastflow affinity chromatography are as follows:
step 1a: blue Sepharose 6Fastflow affinity chromatography pre-and post-loading equilibration: before and after the pretreatment solution is loaded, balancing the chromatographic column by using a balance buffer solution A respectively; the components of the equilibration buffer A are as follows: 35mM phosphate buffer solution, 0.043% polysorbate 20, 0.3M sodium chloride and 1mg/L aprotinin, wherein the pH value is 7.4, and the dosage is 4 times of the column volume;
step 1b: blue Sepharose 6FastFlow affinity chromatography impurity elution: after the sample loading is finished, impurity elution is carried out on the chromatographic column by using an impurity eluent A, and the impurity eluent A comprises the following components: 35mM phosphate buffer, 0.043% polysorbate 20, 1M sodium chloride and 1mg/L aprotinin, wherein the pH value is 7.4, and the dosage is 7 times of the column volume;
step 1c: blue Sepharose 6FastFlow affinity chromatography activity peak elution: after the impurity elution is finished, eluting the chromatographic column by using an eluent A, and collecting a target protein active peak to obtain a Blue chromatographic target protein solution; the eluent A comprises the following components: 35mM phosphate buffer, 0.043% polysorbate 20, 0.7M sodium chloride, 2M urea and 8mg/L aprotinin, at pH 7.4, in an amount of 4 column volumes.
Step 2: performing Lysine HyperD affinity chromatography on the Blue chromatography target protein solution, and then eluting by using an eluent B to obtain a Lysine chromatography target protein solution;
specifically, the steps of the lysine HyperD affinity chromatography are as follows:
step 2a: diluting the Blue target protein solution: before the Blue target protein solution is loaded, diluting the Blue target protein solution by 6 times by using a dilution buffer solution and then loading the Blue target protein solution; the composition of the dilution buffer was: 35mM phosphate buffer, 0.043% polysorbate 20 and 1mg/L aprotinin, at pH 7.4;
and step 2b: equilibration of lysine HyperD affinity chromatography before and after loading: before and after the Blue target protein solution is loaded, balancing the chromatographic column by using a balance buffer solution B respectively; the components of the balance buffer solution B are as follows: 35mM phosphate buffer, 0.043% polysorbate 20 and 2mg/L aprotinin, wherein the pH value is 7.4, and the dosage is 4 times of the column volume;
and step 2c: lysine HyperD affinity chromatography impurity elution: after Blue target protein solution is loaded, impurity elution treatment is carried out on the chromatographic column by using impurity eluent B, and the impurity eluent B comprises the following components: 35mM phosphate buffer, 1M sodium chloride, 0.043% polysorbate 20 and 2mg/L aprotinin, at pH 7.4, in an amount 5 times the column volume.
And step 2d: lysine HyperD affinity chromatography active peak elution: after the impurity elution is finished, eluting the chromatographic column by using an eluent B, and collecting the active peak of the target protein to obtain a Lysine chromatography target protein solution; the eluent B comprises the following components: 0.8M arginine, 0.3M aminocaproic acid, 0.043% polysorbate 20, 6mg/L aprotinin, and 12mg/mL phosphoric acid; the pH was 7.6 and the amount used was 5 times the column volume.
And step 3: diluting the Lysine chromatography target protein solution by using water for injection until the content of the target protein is 0.18mg/ml, then adjusting the pH value of the Lysine chromatography target protein solution to 3.5, then performing virus inactivation treatment, standing at 22 ℃ for 90 minutes to complete virus inactivation, then adjusting the pH value of the Lysine chromatography target protein solution to 7.8, and sequentially filtering by using a 0.1um filter and a 20nm virus removal filter to obtain the tenecteplase protein solution.
The detection results of the tenecteplase protein solutions obtained in the embodiments 1 and 2 respectively show that after the tenecteplase is purified by adopting the process, the virus removal/inactivation index is 18.151og10, and the retrovirus safety index is 101og10 at the moment; the virus removal effect reaches 1 x 10 10 Dosage/granule of 1 × 10 6 Technical requirements of the agent/granule industry.
Various modifications to these embodiments will be readily apparent to those skilled in the art, and the generic principles defined herein may be applied to other embodiments without departing from the spirit or scope of the invention.
Claims (9)
1. A purification process of injection tenecteplase is characterized in that: the method comprises the following steps:
step 1: taking a pretreatment solution containing tenecteplase protein, subjecting the pretreatment solution to Blue Sepharose 6Fastflow affinity chromatography, and eluting with an eluent A to obtain a Blue chromatography target protein solution;
step 2: performing Lysine HyperD affinity chromatography on the Blue chromatography target protein solution, and then eluting by using an eluent B to obtain a Lysine chromatography target protein solution;
and, step 3: diluting the Lysine chromatography target protein solution, adjusting the pH value of the Lysine chromatography target protein solution to acidity, then performing virus inactivation treatment, adjusting the pH value of the Lysine chromatography target protein solution to neutrality after virus inactivation is completed, and filtering by using a 0.1um filter and a 20nm virus removal filter in sequence to obtain the tenecteplase protein solution.
2. The purification process of tenecteplase for injection according to claim 1, wherein: in the step 1, the binding capacity of the Blue Sepharose 6Fastflow affinity chromatography to the tenecteplase protein is 10-20mg/ml.
3. The purification process of tenecteplase for injection according to claim 1, wherein: in the step 1, the eluent A comprises the following components: 10-50mM phosphate buffer, 0.043% polysorbate 20, 0.5-1M sodium chloride, 2-4M urea, and 5-10mg/L aprotinin.
4. The purification process of tenecteplase for injection according to claim 3, wherein: the pH value of the eluent A is 7.2-8.0, and the dosage of the eluent A is 3-5 times of the column volume.
5. The purification process of tenecteplase for injection as claimed in claim 1, wherein: in the step 2, the binding capacity of the lysine HyperD affinity chromatography to the tenecteplase protein is 5-10mg/ml.
6. The purification process of tenecteplase for injection as claimed in claim 1, wherein: the eluent B comprises the following components: 0.2-1.0M arginine, 0.1-0.5M aminocaproic acid, 0.043% polysorbate 20, 6-10mg/L aprotinin, and 5-18mg/mL phosphoric acid.
7. The purification process of tenecteplase for injection according to claim 6, wherein: the pH value of the eluent B is 7.4-7.8, and the dosage of the eluent B is 3-5 times of the column volume.
8. The purification process of tenecteplase for injection as claimed in claim 1, wherein: in the step 3, the Lysine chromatography target protein solution is diluted by water for injection until the content of the target protein is 0.19-0.21 mg/ml.
9. The purification process of tenecteplase for injection as claimed in claim 8, wherein: in the step 3, when the virus inactivation treatment is carried out, the pH value of the Lysine chromatography target protein solution is 3.3-3.7.
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Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101942429A (en) * | 2010-08-25 | 2011-01-12 | 广州铭康生物工程有限公司 | Purification method for recombined human tissue type plasminogen exciter TNK mutant rhTNK-tPA |
| CN106103464A (en) * | 2014-01-17 | 2016-11-09 | 建新公司 | Aseptic chromatography and manufacturing process |
| CN108424897A (en) * | 2018-03-13 | 2018-08-21 | 广州铭康生物工程有限公司 | A kind of purification process of rhTNK-tPA cells harvest liquid |
| CN109843904A (en) * | 2016-10-17 | 2019-06-04 | 恩泽生物科学有限公司 | For reducing the heterogeneous continuation method of human cytokines |
| CN110257358A (en) * | 2019-06-10 | 2019-09-20 | 广东双林生物制药有限公司 | A kind of production method of high-purity Complex |
-
2022
- 2022-09-06 CN CN202211084472.0A patent/CN115475258A/en active Pending
Patent Citations (5)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101942429A (en) * | 2010-08-25 | 2011-01-12 | 广州铭康生物工程有限公司 | Purification method for recombined human tissue type plasminogen exciter TNK mutant rhTNK-tPA |
| CN106103464A (en) * | 2014-01-17 | 2016-11-09 | 建新公司 | Aseptic chromatography and manufacturing process |
| CN109843904A (en) * | 2016-10-17 | 2019-06-04 | 恩泽生物科学有限公司 | For reducing the heterogeneous continuation method of human cytokines |
| CN108424897A (en) * | 2018-03-13 | 2018-08-21 | 广州铭康生物工程有限公司 | A kind of purification process of rhTNK-tPA cells harvest liquid |
| CN110257358A (en) * | 2019-06-10 | 2019-09-20 | 广东双林生物制药有限公司 | A kind of production method of high-purity Complex |
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