CN110628054A - Preparation of interpenetrating network hydrogel chromatographic medium - Google Patents
Preparation of interpenetrating network hydrogel chromatographic medium Download PDFInfo
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- CN110628054A CN110628054A CN201910922362.9A CN201910922362A CN110628054A CN 110628054 A CN110628054 A CN 110628054A CN 201910922362 A CN201910922362 A CN 201910922362A CN 110628054 A CN110628054 A CN 110628054A
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- 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
- B01J20/291—Gel sorbents
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- C—CHEMISTRY; METALLURGY
- C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
- C08J—WORKING-UP; GENERAL PROCESSES OF COMPOUNDING; AFTER-TREATMENT NOT COVERED BY SUBCLASSES C08B, C08C, C08F, C08G or C08H
- C08J3/00—Processes of treating or compounding macromolecular substances
- C08J3/02—Making solutions, dispersions, lattices or gels by other methods than by solution, emulsion or suspension polymerisation techniques
- C08J3/03—Making solutions, dispersions, lattices or gels by other methods than by solution, emulsion or suspension polymerisation techniques in aqueous media
- C08J3/075—Macromolecular gels
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- C—CHEMISTRY; METALLURGY
- C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
- C08J—WORKING-UP; GENERAL PROCESSES OF COMPOUNDING; AFTER-TREATMENT NOT COVERED BY SUBCLASSES C08B, C08C, C08F, C08G or C08H
- C08J3/00—Processes of treating or compounding macromolecular substances
- C08J3/24—Crosslinking, e.g. vulcanising, of macromolecules
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- C—CHEMISTRY; METALLURGY
- C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
- C08J—WORKING-UP; GENERAL PROCESSES OF COMPOUNDING; AFTER-TREATMENT NOT COVERED BY SUBCLASSES C08B, C08C, C08F, C08G or C08H
- C08J2305/00—Characterised by the use of polysaccharides or of their derivatives not provided for in groups C08J2301/00 or C08J2303/00
- C08J2305/08—Chitin; Chondroitin sulfate; Hyaluronic acid; Derivatives thereof
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- C—CHEMISTRY; METALLURGY
- C08—ORGANIC MACROMOLECULAR COMPOUNDS; THEIR PREPARATION OR CHEMICAL WORKING-UP; COMPOSITIONS BASED THEREON
- C08J—WORKING-UP; GENERAL PROCESSES OF COMPOUNDING; AFTER-TREATMENT NOT COVERED BY SUBCLASSES C08B, C08C, C08F, C08G or C08H
- C08J2329/00—Characterised by the use of homopolymers or copolymers of compounds having one or more unsaturated aliphatic radicals, each having only one carbon-to-carbon double bond, and at least one being terminated by an alcohol, ether, aldehydo, ketonic, acetal, or ketal radical; Hydrolysed polymers of esters of unsaturated alcohols with saturated carboxylic acids; Derivatives of such polymer
- C08J2329/02—Homopolymers or copolymers of unsaturated alcohols
- C08J2329/04—Polyvinyl alcohol; Partially hydrolysed homopolymers or copolymers of esters of unsaturated alcohols with saturated carboxylic acids
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- Solid-Sorbent Or Filter-Aiding Compositions (AREA)
- Processes Of Treating Macromolecular Substances (AREA)
- Treatment Of Liquids With Adsorbents In General (AREA)
Abstract
The invention mainly relates to preparation of an interpenetrating network hydrogel chromatographic medium. The spherical chromatographic medium with interpenetrating network hydrogel can be prepared by mixing two different hydrogels according to a certain proportion and utilizing a suspension emulsification method and through composite chemical crosslinking. The spherical chromatographic medium not only has obvious interpenetrating network structure characteristics, but also has the respective characteristics of two hydrogels, stable physicochemical properties and better mechanical strength, and can widen the application field of hydrogel soft matrix chromatographic media.
Description
Technical Field
The invention relates to preparation of an interpenetrating network hydrogel chromatographic medium, which is characterized in that an interpenetrating network hydrogel structure is formed after composite chemical crosslinking, and belongs to the technical field of liquid chromatography.
Background
The liquid chromatographic separation technology is a high-efficiency separation and purification technology widely applied in the biological downstream processing process, and the liquid chromatographic medium is a key factor for determining the performance of liquid chromatographic separation. Due to the large molecular size, complex structure and slow diffusion speed of biological macromolecules (such as proteins, plasmid DNA and the like), mass transfer in a chromatographic medium is always a bottleneck problem in the chromatographic separation process and becomes a main factor for limiting the separation efficiency. The liquid chromatography technology is a main direction for the development of the large-scale separation and purification technology of biomacromolecules. Chromatographic separation media, as an important component of liquid chromatographic separation technology, have been the core and leading edge of the research field of liquid chromatographic technology.
The chitosan is a linear biological polyamino sugar, is obtained by alkaline deacetylation of chitin, has a similar structure to cellulose, contains a large amount of free amino groups in a molecular chain, is an active group, can perform a plurality of functional group reactions, is low in price and easy to obtain as a raw material, is a natural high polymer material with excellent performance, and has good adsorption performance on metal ions, dye molecules and organic matters. The chitosan-chitosan composite material also has good compatibility with biological macromolecules and high hydrophilicity, can allow the biological macromolecules to permeate without deformation, is suitable for separating the biological macromolecules such as protein and the like, but the medium is mostly a soft matrix material originally, has large pressure drop during application, has higher back pressure of a column, is not beneficial to column type operation, has the defects of easy dissolution, poor mechanical property, inconvenience in recycling and the like in an acidic medium, and greatly limits the application and popularization of the chitosan. The polyvinyl alcohol is obtained by hydrolyzing after vinyl acetate polymerization, has rich hydroxyl groups on molecular chains, has high hydrophilicity and biomacromolecule compatibility, is similar to the performance of polysaccharide beads, can be chemically modified to obtain various biochemical separation media by a large amount of hydroxyl groups, and also has excellent chemical stability and antimicrobial corrosivity.
The invention combines the compatibility of the biomacromolecule of the chitosan with the chemical stability of the polyvinyl alcohol, forms the interpenetrating network hydrogel structure after composite chemical crosslinking, has stable physical and chemical properties and higher mechanical strength, is a chromatographic medium with excellent performance, and can be widely applied to biochemical separation. By using the composite material, the respective defects of the material are overcome, the prepared medium has a regular shape, the adhesion phenomenon does not occur, and the mechanical strength of the hydrogel soft matrix medium is improved.
Disclosure of Invention
The invention aims to prepare an interpenetrating network hydrogel chromatographic medium. The method comprises the steps of forming an interpenetrating network hydrogel structure by carrying out composite chemical crosslinking on chitosan and polyvinyl alcohol, preparing a chromatographic medium with uniform particle size distribution by a suspension emulsification method, and analyzing the property of the medium.
The specific steps of the medium are as follows:
(1) preparation of chitosan gel: dissolving 1.25ml acetic acid in 23.75ml deionized water to obtain 5% acetic acid solution, adding 1.5g chitosan, stirring, and mixing.
(2) Preparation of polyvinyl alcohol gel: dissolving 1.5g of polyvinyl alcohol in 25ml of deionized water, soaking for 4h, and then putting into a 90 ℃ water bath kettle to heat and stir for 20 min.
(3) Preparation of composite gel: pouring the prepared polyvinyl alcohol gel into the chitosan gel, and fully stirring for 5 hours by using a magnetic stirrer.
(4) Preparation of the medium: adding 300ml of soybean oil and 8ml of span 80 into a stainless steel reactor, stirring for 20min, adding 50ml of mixed gel, placing the mixture in a water bath at 55 ℃, stirring for 30min at a stirring speed of 1100 rpm, fully dispersing the gel in an oil phase, adding 25% glutaraldehyde and hydrochloric acid, continuing stirring for 1h, adding a certain amount of sodium hydroxide, adjusting the pH to be 8-10, stirring for 30min, adding 3 times of deionized water, standing, collecting microspheres, and sequentially cleaning with acetone, ethanol and deionized water. Then adding epichlorohydrin and dimethyl sulfoxide to activate the microspheres.
(5) Modification of the ligand: adding the cleaned medium into 0.5mol/L DEAE-HCl, simultaneously adding a certain amount of 3.5mol/L sodium hydroxide, placing the mixture in a water bath shaker at 60 ℃ for reaction for 3 hours, cooling, and cleaning with deionized water.
Further, in the step (2), the polyvinyl alcohol is 1788 type, and the alcoholysis degree is 87% -89%.
Further, in the step (4), the 25% glutaraldehyde is used in an amount of 200. mu.l to 1000. mu.l.
Furthermore, in the step (4), the concentration of the hydrochloric acid is 1mol/L, and the addition amount is 1-1.5 ml.
Furthermore, in the step (4), the concentration of the sodium hydroxide is 3.5mol/L, and the addition amount is 5-8 ml.
Drawings
FIG. 1 is a spectrum chart of an interpenetrating network type chromatographic medium having a high degree of crosslinking in example 2.
Detailed Description
According to the invention, an interpenetrating network hydrogel structure is formed by carrying out composite chemical crosslinking on chitosan and polyvinyl alcohol, and a chromatographic medium with uniform particle size distribution is prepared by a suspension emulsification method.
Example 1
The preparation method of the interpenetrating network type chromatographic medium with low crosslinking degree comprises the following steps:
(1) preparation of composite gel: mixing 6% polyvinyl alcohol and 6% chitosan containing 5% acetic acid at a ratio of 1:1, and stirring for 4 hr.
(2) Preparation of the medium: adding 300ml of soybean oil and 8ml of span 80 into a stainless steel reactor, stirring for 20min, adding 50ml of mixed gel, placing the mixture in a water bath at 55 ℃, stirring for 30min at a stirring speed of 1100 rpm, fully dispersing the gel in an oil phase, adding 200 mu l of 25% glutaraldehyde and hydrochloric acid, continuing stirring for 1h, adding a certain amount of sodium hydroxide, adjusting the pH to 8-10, stirring for 30min, adding 3 times of deionized water, standing, collecting microspheres, and sequentially cleaning with acetone, ethanol and deionized water. Then adding epichlorohydrin and dimethyl sulfoxide to activate the microspheres.
(3) Modification of the ligand: adding the cleaned medium into 0.5mol/L DEAE-HCl, simultaneously adding a certain amount of 3.5mol/L sodium hydroxide, placing the mixture in a water bath shaker at 60 ℃ for reaction for 3 hours, cooling, and cleaning with deionized water.
Example 2
The preparation method of the interpenetrating network type chromatographic medium with high crosslinking degree comprises the following specific steps:
(1) preparation of composite gel: mixing 6% polyvinyl alcohol and 6% chitosan containing 5% acetic acid at a ratio of 1:1, and stirring for 4 hr.
(2) Preparation of the medium: adding 300ml of soybean oil and 8ml of span 80 into a stainless steel reactor, stirring for 20min, adding 50ml of mixed gel, placing the mixture in a water bath at 55 ℃, stirring for 30min at a stirring speed of 1100 rpm, fully dispersing the gel in an oil phase, adding 1000 mu l of 25% glutaraldehyde and hydrochloric acid, continuing stirring for 1h, adding a certain amount of sodium hydroxide, adjusting the pH to 8-10, stirring for 30min, adding 3 times of deionized water, standing, collecting microspheres, and sequentially cleaning with acetone, ethanol and deionized water. Then adding epichlorohydrin and dimethyl sulfoxide to activate the microspheres.
(3) Modification of the ligand: adding the cleaned medium into 0.5mol/L DEAE-HCl, simultaneously adding a certain amount of 3.5mol/L sodium hydroxide, placing the mixture in a water bath shaker at 60 ℃ for reaction for 3 hours, cooling, and cleaning with deionized water.
Claims (5)
1. The invention relates to a preparation method of an interpenetrating network hydrogel spherical chromatographic medium, which mainly comprises the following steps:
(1) prefabrication of the composite hydrogel: mixing 3-8% polyvinyl alcohol and 3-8% chitosan containing 2-6% acetic acid at a ratio of 1:1, and stirring for 1-4 hr.
(2) Preparation of the medium: adding 400ml of span 80 and soybean oil mixture of 300-4% by volume into a stainless steel reactor, stirring for 10-30min, adding 50ml of composite gel, placing in a water bath at 50-60 ℃, stirring for 20-40min at the stirring speed of 900-1200rpm, fully dispersing the gel in an oil phase, adding 25% of glutaraldehyde and hydrochloric acid, continuing stirring for 30-60min, adding a certain amount of sodium hydroxide, adjusting the pH to 8-10, stirring for 30min, adding 3 times of deionized water, standing, collecting microspheres, and sequentially cleaning with acetone, ethanol and deionized water. And then activating the microspheres by using epoxy chloropropane and dimethyl sulfoxide, wherein the addition of the dimethyl sulfoxide is used for improving the solubility.
(3) Modification of the ligand: adding the cleaned medium into 0.5mol/L DEAE-HCl, simultaneously adding a certain amount of 3.5mol/L sodium hydroxide, placing the mixture in a water bath shaker at 60 ℃ for reaction for 3h, cooling, and cleaning with deionized water.
2. The method of claim 1, wherein in step (1), the polyvinyl alcohol is of type 1788 with an alcoholysis level of 87% to 89%.
3. The method according to claim 1, wherein in the step (2), the 25% glutaraldehyde is used in an amount of 200 to 1000. mu.l.
4. The method according to claim 1, wherein in the step (2), the hydrochloric acid has a concentration of 1mol/L and is added in an amount of 1 to 1.5 ml.
5. The method according to claim 1, wherein in the step (2), the concentration of the sodium hydroxide is 3.5mol/L, and the addition amount is 5-8 ml.
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| CN201910922362.9A CN110628054A (en) | 2019-09-26 | 2019-09-26 | Preparation of interpenetrating network hydrogel chromatographic medium |
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Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102019213A (en) * | 2010-11-30 | 2011-04-20 | 浙江工业大学 | Method for preparing strongly-acid ion exchange medium |
| CN110026138A (en) * | 2018-01-11 | 2019-07-19 | 中国科学院过程工程研究所 | Polysaccharide microsphere and preparation method thereof |
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- 2019-09-26 CN CN201910922362.9A patent/CN110628054A/en active Pending
Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN102019213A (en) * | 2010-11-30 | 2011-04-20 | 浙江工业大学 | Method for preparing strongly-acid ion exchange medium |
| CN110026138A (en) * | 2018-01-11 | 2019-07-19 | 中国科学院过程工程研究所 | Polysaccharide microsphere and preparation method thereof |
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Application publication date: 20191231 |