CN106582578B - High pressure resistant affinity chromatography medium - Google Patents
High pressure resistant affinity chromatography medium Download PDFInfo
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- CN106582578B CN106582578B CN201611201279.5A CN201611201279A CN106582578B CN 106582578 B CN106582578 B CN 106582578B CN 201611201279 A CN201611201279 A CN 201611201279A CN 106582578 B CN106582578 B CN 106582578B
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- high pressure
- pressure resistant
- affinity chromatography
- chromatography medium
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- 239000012501 chromatography medium Substances 0.000 title claims abstract description 25
- 238000001042 affinity chromatography Methods 0.000 title claims abstract description 22
- 239000004005 microsphere Substances 0.000 claims abstract description 14
- 229920000936 Agarose Polymers 0.000 claims abstract description 12
- 229920002125 Sokalan® Polymers 0.000 claims abstract description 12
- 150000002148 esters Chemical class 0.000 claims abstract description 12
- 239000004584 polyacrylic acid Substances 0.000 claims abstract description 12
- 125000000956 methoxy group Chemical group [H]C([H])([H])O* 0.000 claims abstract description 5
- 125000002887 hydroxy group Chemical group [H]O* 0.000 claims abstract description 4
- 238000004519 manufacturing process Methods 0.000 abstract description 4
- 238000000034 method Methods 0.000 abstract description 3
- 238000012986 modification Methods 0.000 abstract description 3
- 230000004048 modification Effects 0.000 abstract description 3
- 238000013461 design Methods 0.000 abstract description 2
- 229920000058 polyacrylate Polymers 0.000 abstract description 2
- HEMHJVSKTPXQMS-UHFFFAOYSA-M Sodium hydroxide Chemical compound [OH-].[Na+] HEMHJVSKTPXQMS-UHFFFAOYSA-M 0.000 description 12
- YXFVVABEGXRONW-UHFFFAOYSA-N Toluene Chemical compound CC1=CC=CC=C1 YXFVVABEGXRONW-UHFFFAOYSA-N 0.000 description 6
- 239000003814 drug Substances 0.000 description 5
- 239000002609 medium Substances 0.000 description 5
- CSCPPACGZOOCGX-UHFFFAOYSA-N Acetone Chemical compound CC(C)=O CSCPPACGZOOCGX-UHFFFAOYSA-N 0.000 description 4
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 description 4
- 229940079593 drug Drugs 0.000 description 4
- 238000001035 drying Methods 0.000 description 4
- 238000001914 filtration Methods 0.000 description 4
- 238000005406 washing Methods 0.000 description 4
- 125000002496 methyl group Chemical group [H]C([H])([H])* 0.000 description 3
- 102000004169 proteins and genes Human genes 0.000 description 3
- 108090000623 proteins and genes Proteins 0.000 description 3
- LRWZZZWJMFNZIK-UHFFFAOYSA-N 2-chloro-3-methyloxirane Chemical compound CC1OC1Cl LRWZZZWJMFNZIK-UHFFFAOYSA-N 0.000 description 2
- 235000019441 ethanol Nutrition 0.000 description 2
- 230000008676 import Effects 0.000 description 2
- 229940070721 polyacrylate Drugs 0.000 description 2
- 238000002360 preparation method Methods 0.000 description 2
- 239000002904 solvent Substances 0.000 description 2
- 229920001817 Agar Polymers 0.000 description 1
- 102000007056 Recombinant Fusion Proteins Human genes 0.000 description 1
- 108010008281 Recombinant Fusion Proteins Proteins 0.000 description 1
- 239000008272 agar Substances 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- 238000004587 chromatography analysis Methods 0.000 description 1
- 230000007812 deficiency Effects 0.000 description 1
- 230000000694 effects Effects 0.000 description 1
- 239000000499 gel Substances 0.000 description 1
- 239000003446 ligand Substances 0.000 description 1
- 125000001997 phenyl group Chemical group [H]C1=C([H])C([H])=C(*)C([H])=C1[H] 0.000 description 1
- 238000004064 recycling Methods 0.000 description 1
- 238000012827 research and development Methods 0.000 description 1
- 239000000126 substance Substances 0.000 description 1
- 238000012360 testing method Methods 0.000 description 1
Classifications
-
- B—PERFORMING OPERATIONS; TRANSPORTING
- B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
- B01J—CHEMICAL OR PHYSICAL PROCESSES, e.g. CATALYSIS OR COLLOID CHEMISTRY; THEIR RELEVANT APPARATUS
- B01J20/00—Solid sorbent compositions or filter aid compositions; Sorbents for chromatography; Processes for preparing, regenerating or reactivating thereof
- B01J20/281—Sorbents specially adapted for preparative, analytical or investigative chromatography
-
- B—PERFORMING OPERATIONS; TRANSPORTING
- B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
- B01J—CHEMICAL OR PHYSICAL PROCESSES, e.g. CATALYSIS OR COLLOID CHEMISTRY; THEIR RELEVANT APPARATUS
- B01J20/00—Solid sorbent compositions or filter aid compositions; Sorbents for chromatography; Processes for preparing, regenerating or reactivating thereof
- B01J20/22—Solid sorbent compositions or filter aid compositions; Sorbents for chromatography; Processes for preparing, regenerating or reactivating thereof comprising organic material
-
- B—PERFORMING OPERATIONS; TRANSPORTING
- B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
- B01J—CHEMICAL OR PHYSICAL PROCESSES, e.g. CATALYSIS OR COLLOID CHEMISTRY; THEIR RELEVANT APPARATUS
- B01J20/00—Solid sorbent compositions or filter aid compositions; Sorbents for chromatography; Processes for preparing, regenerating or reactivating thereof
- B01J20/22—Solid sorbent compositions or filter aid compositions; Sorbents for chromatography; Processes for preparing, regenerating or reactivating thereof comprising organic material
- B01J20/26—Synthetic macromolecular compounds
-
- B—PERFORMING OPERATIONS; TRANSPORTING
- B01—PHYSICAL OR CHEMICAL PROCESSES OR APPARATUS IN GENERAL
- B01J—CHEMICAL OR PHYSICAL PROCESSES, e.g. CATALYSIS OR COLLOID CHEMISTRY; THEIR RELEVANT APPARATUS
- B01J2220/00—Aspects relating to sorbent materials
- B01J2220/50—Aspects relating to the use of sorbent or filter aid materials
- B01J2220/52—Sorbents specially adapted for preparative chromatography
Landscapes
- Chemical & Material Sciences (AREA)
- Analytical Chemistry (AREA)
- Organic Chemistry (AREA)
- Chemical Kinetics & Catalysis (AREA)
- Treatment Of Liquids With Adsorbents In General (AREA)
Abstract
This case is related to high pressure resistant affinity chromatography medium, using polyacrylic acid ester microsphere as kernel, and is bonded with agarose molecules in the polyacrylate microsphere surface；Hydroxyl terminal a part in the agarose molecules connects high pressure resistant group, and remainder connects aglucon；Wherein, the high pressure resistant group is CH2Ph‑p‑CH3Or CH2Ph‑p‑OCH3.This case is improved by the design to affinity chromatography medium surface modification group, introduces the big steric group of rigidity in dielectric surface, so as to improve the separative efficiency of chromatography media at elevated pressures and number is recycled in the mechanical strength of Lai Tigao affinity chromatography medium；Meanwhile this improved method is simple, production cost is low, is conducive to large-scale promotion application.
Description
Technical field
The present invention relates to a kind of chromatography medias, and in particular to a kind of high pressure resistant affinity chromatography medium.
Background technique
It is the biologies such as recombinant protein, antibody currently, how to improve the purity and biological safety of drug in pharmaceutical field
Final steps in medicine research and development and production process.Medicine enterprise in China's relies primarily on import, therefore, state for the acquisition of separating medium
It inside needs to get rid of the dependence to import separating medium, reduces the production cost of drug.
Affinity chromatography medium to protein drug purifies and separates is mainly agar sugar product, but this kind of product mechanical strength
Lower, so as to cause it after loading chromatographic column, proof pressure is lower, limits the room for promotion of its separative efficiency, while
Inhibit the reuse number of agarose.
Summary of the invention
In view of the deficiencies of the prior art, the purpose of the present invention is to provide a kind of high pressure resistant affinity chromatography mediums.
Technical solution of the present invention is summarized as follows:
A kind of high pressure resistant affinity chromatography medium, using polyacrylic acid ester microsphere as kernel, and it is micro- in the polyacrylate
Ball surface is bonded with agarose molecules；Hydroxyl terminal a part in the agarose molecules connects high pressure resistant group, remainder
Connect aglucon；
Wherein, the high pressure resistant group is CH2Ph-p-CH3Or CH2Ph-p-OCH3.(p represents methyl and aligns in phenyl ring)
Preferably, the high pressure resistant affinity chromatography medium, wherein mole of the high pressure resistant group and the aglucon
Than being 5-15: 100.
Preferably, the high pressure resistant affinity chromatography medium, wherein mole of the high pressure resistant group and the aglucon
Than being 8-10: 100.
Preferably, the high pressure resistant affinity chromatography medium, wherein the high pressure resistant group is CH2Ph-p-CH3。
Preferably, the high pressure resistant affinity chromatography medium, wherein the high pressure resistant group is CH2Ph-p-OCH3。
Preferably, the high pressure resistant affinity chromatography medium, wherein the partial size of the polyacrylic acid ester microsphere is 30-
100μm。
The beneficial effects of the present invention are: this case is improved by the design to affinity chromatography medium surface modification group, it is being situated between
Matter surface introduces the big steric group of rigidity, the mechanical strength of Lai Tigao affinity chromatography medium, so as to improve chromatography media in height
Separative efficiency and recycling number under pressure；Meanwhile this improved method is simple, production cost is low, conducive to pushing away on a large scale
Wide application.
Specific embodiment
The present invention will be further described in detail below with reference to the embodiments, to enable those skilled in the art referring to specification
Text can be implemented accordingly.
This case provides the high pressure resistant affinity chromatography medium of an embodiment, using polyacrylic acid ester microsphere as kernel, and poly-
Acrylate microspheres surface bond has agarose molecules；Hydroxyl terminal a part in agarose molecules connects high pressure resistant group, remains
Remaining part point connection aglucon；
Wherein, high pressure resistant group is CH2Ph-p-CH3Or CH2Ph-p-OCH3。
Wherein, the molar ratio of high pressure resistant group and aglucon is 5-15: 100；Preferably 8-10: 100.
Wherein, the partial size of polyacrylic acid ester microsphere is preferably 30-100 μm.
Polyacrylic acid ester microsphere is a kind of conventional commercial commodity, can be bought.Using Ago-Gel as medium carrier
Chromatographic column pressure voltage is generally 0.3-0.5MPa, and has the polyacrylic acid ester microsphere of agarose molecules as medium using surface bond
The chromatographic column pressure voltage of carrier is generally 10-12MPa, and reusing number is 100 times.
The type of aglucon (L:Ligand) is not limited, can be depending on the type of substance to be adsorbed.Such as when needing to make
When standby protein A affinity chromatography medium, aglucon just selects ProA；When needing to prepare protein g affinity chromatography medium, aglucon is just selected
ProG.Whether the selection of aglucon is high pressure resistant to the medium to have no substantial effect.
Embodiment 1
Preparation process is as follows:
1) BrCH is added in the ratio that the molar ratio of high pressure resistant group and aglucon is 5-15: 100 into 1mol carrier2Ph-
p-CH3, 8-10mol toluene and 0.3-0.4mol KOH, be stirred to react 1-1.5h at 70-75 DEG C；Filtering, washing three times, are done
It is dry；Carrier is the polyacrylic acid ester microsphere that surface bond has agarose molecules；
2) 3-4mol acetone, 1.1-1.2mol epoxychloropropane and 1.5-2mol NaOH is added, is stirred at 30-40 DEG C
1-2h, after reaction, drying is washed in filtering；
3) aglucon, the ethyl alcohol of 5-7mol and the NaOH of 2-3mol of 1.2mol is added, is stirred to react 12- at 55-60 DEG C
15h, is filtered to remove solvent after reaction, and washing three times, is drying to obtain chromatography media.
Embodiment 2
Preparation process is as follows:
1) CH is added in the ratio that the molar ratio of high pressure resistant group and aglucon is 5-15: 100 into 1mol carrier2Ph-p-
OCH3, 8-10mol toluene and 0.3-0.4mol KOH, be stirred to react 1-1.5h at 70-75 DEG C；Filtering, washing three times, are done
It is dry；Carrier is the polyacrylic acid ester microsphere that surface bond has agarose molecules；
2) 3-4mol acetone, 1.1-1.2mol epoxychloropropane and 1.5-2mol NaOH is added, is stirred at 30-40 DEG C
1-2h, after reaction, drying is washed in filtering；
3) aglucon, the ethyl alcohol of 5-7mol and the NaOH of 2-3mol of 1.2mol is added, is stirred to react 12- at 55-60 DEG C
15h, is filtered to remove solvent after reaction, and washing three times, is drying to obtain chromatography media.
Following table lists the test result of embodiment 1 and embodiment 2:
| Embodiment 1 | Embodiment 2 | |
| Chromatographic column pressure voltage | 18MPa | 19MPa |
| Reuse number | 200 times | 200 times |
Although the embodiments of the present invention have been disclosed as above, but its is not only in the description and the implementation listed
With it can be fully applied to various fields suitable for the present invention, for those skilled in the art, can be easily
Realize other modification, therefore without departing from the general concept defined in the claims and the equivalent scope, the present invention is simultaneously unlimited
In specific details.
Claims (3)
1. a kind of high pressure resistant affinity chromatography medium, using polyacrylic acid ester microsphere as kernel, and in the polyacrylic acid ester microsphere
Surface bond has agarose molecules；Hydroxyl terminal a part in the agarose molecules connects high pressure resistant group, and remainder connects
Connect aglucon；
Wherein, the high pressure resistant group is CH2Ph-p-CH3Or CH2Ph-p-OCH3；
The molar ratio of the high pressure resistant group and the aglucon is 5-15: 100.
2. high pressure resistant affinity chromatography medium according to claim 1, which is characterized in that the high pressure resistant group is matched with described
The molar ratio of base is 8-10: 100.
3. high pressure resistant affinity chromatography medium according to claim 1, which is characterized in that the grain of the polyacrylic acid ester microsphere
Diameter is 30-100 μm.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201611201279.5A CN106582578B (en) | 2016-12-22 | 2016-12-22 | High pressure resistant affinity chromatography medium |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201611201279.5A CN106582578B (en) | 2016-12-22 | 2016-12-22 | High pressure resistant affinity chromatography medium |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN106582578A CN106582578A (en) | 2017-04-26 |
| CN106582578B true CN106582578B (en) | 2019-06-14 |
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| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201611201279.5A Active CN106582578B (en) | 2016-12-22 | 2016-12-22 | High pressure resistant affinity chromatography medium |
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| Country | Link |
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Families Citing this family (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN107961768A (en) * | 2017-12-06 | 2018-04-27 | 苏州博进生物技术有限公司 | Chromatography media using indoles -3- acethydrazides as functional ligand |
| CN114054006A (en) * | 2021-11-17 | 2022-02-18 | 苏州纳微科技股份有限公司 | Oligo-dT affinity chromatography filler |
Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US4973683A (en) * | 1985-05-23 | 1990-11-27 | Pharmacia Ab | Method of cross-linking a porous polysaccharide gel |
| CN101259406A (en) * | 2007-12-17 | 2008-09-10 | 南京工业大学 | A preparation method of bonding-affinity composite polysaccharide chiral stationary phase |
| CN102617869A (en) * | 2012-03-30 | 2012-08-01 | 中国科学院过程工程研究所 | Polyacrylic ester and polyacrylic ester copolymer material hydrophilic modification products and hydrophilic modification method of polyacrylic ester |
| CN103586008A (en) * | 2013-10-24 | 2014-02-19 | 中国科学院过程工程研究所 | Affinity chromatography medium and preparation method and application thereof |
-
2016
- 2016-12-22 CN CN201611201279.5A patent/CN106582578B/en active Active
Patent Citations (4)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US4973683A (en) * | 1985-05-23 | 1990-11-27 | Pharmacia Ab | Method of cross-linking a porous polysaccharide gel |
| CN101259406A (en) * | 2007-12-17 | 2008-09-10 | 南京工业大学 | A preparation method of bonding-affinity composite polysaccharide chiral stationary phase |
| CN102617869A (en) * | 2012-03-30 | 2012-08-01 | 中国科学院过程工程研究所 | Polyacrylic ester and polyacrylic ester copolymer material hydrophilic modification products and hydrophilic modification method of polyacrylic ester |
| CN103586008A (en) * | 2013-10-24 | 2014-02-19 | 中国科学院过程工程研究所 | Affinity chromatography medium and preparation method and application thereof |
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| CN106582578A (en) | 2017-04-26 |
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