CN104725559B - Thiophilic chromatography material and its preparation method and application - Google Patents

Abstract

The invention discloses a thiolophilic chromatographic material and its preparation method and application. The preparation method comprises: a. In the presence of an initiator, adding glycidyl methacrylate and methylenebisacrylamide to a porogen Carry out copolymerization reaction in and make copolymer poly(GMA-co-MBAA); b, in the presence of the first organic solvent, carry out ring-opening reaction with copolymer poly(GMA-co-MBAA) and sulfonamide to make copolymer poly (SFA-(GMA-co-MBAA)). The thiophile chromatographic material prepared by the method has high mechanical strength, good permeability and stable modified sulfone-sulfide group, and the thiophilic chromatographic material can selectively enrich and separate antibodies.

Description

Thiophilic chromatography material and its preparation method and application

technical field

The invention relates to a chromatographic material, in particular to a thiophilic chromatographic material and its preparation method and application.

Background technique

With the rapid development of molecular biology, immune engineering and genetic engineering, antibodies have played an increasingly extensive role in the fields of disease treatment, biopharmaceuticals and biotechnology. In the domestic and foreign markets, the demand for antibodies is increasing year by year. According to the latest survey by Business Communication, the demand for antibodies has continued to grow at a growth rate of 4.4% since 1993. Among them, the market share of monoclonal antibodies alone has reached tens of billions of dollars [see Nature Biotech., 2010, 28, 917-924]. However, the use of antibodies, especially in vivo applications, has strict requirements on purity. The development of low-cost and high-efficiency antibody purification technologies and methods can not only promote the progress of scientific research, but also contain huge business opportunities.

Thiophilic chromatography, also known as thiophilic chromatography [see Analytica Chimica Acta.2010, 658, 18–31], is a method for antibody enrichment and separation. So far, scholars at home and abroad have been working on the preparation of various thiolophilic chromatography materials and applying them to the enrichment and purification of antibodies. As early as the 1980s, Porath et al [see FEBS Lett, 1985, 185, 306-310] used divinyl sulfone to activate agarose to prepare T2gel gel, which was the earliest thiophile chromatography separation material. Thereafter, Yannick et al. [see J.Chromatogr.B, 2004, 808, 51–56] modified the thophilic reagent on polyethylene divinyl alcohol on the hollow fiber membrane to achieve the separation of IgG in complex systems. In 2010, Lakhiar et al [see Chromatogr., 2010,72,205-213] bonded ethylenediamine to the surface of dextran to neutralize the electronegativity of dextran, and then introduced sulfone-sulfide groups , Successfully enriched and separated antibodies in mouse ascites. Fu Rong et al. from Fuzhou University in my country [see Natural Science, 2000, 28(6), 110-114] used commercial T2gel to separate and purify yolk immunoglobulin (IgY), and the activity recovery rate can be as high as 70%. Qian Hao et al. from Huaqiao University [see Biotech.Progr., 2009, 25, 376-383] are engaged in the related research work of thiophilic magnetic microspheres. They fully hydrolyze superparamagnetic polymer microspheres, and then pass Michael addition reaction , a thiophilic group is coupled on the surface of the magnetic microspheres, and the thiophilic magnetic microspheres can be used for the purification of antibodies in human serum with excellent results.

However, agarose and gel are soft matrices and cannot withstand high pressure; and post-modification on magnetic microspheres is prone to loss of functional groups. This type of material has low mechanical strength, high cost, difficult storage, and cannot be reused many times, which cannot meet high-throughput antibody purification.

Contents of the invention

The purpose of the present invention is to provide a thiolophilic chromatographic material and its preparation method and application. The thiolophilic chromatographic material prepared by the method has high mechanical strength, good permeability and stable modified sulfone-sulfide groups. Thiophilic chromatographic materials enable selective enrichment and separation of antibodies.

In order to achieve the above object, the invention provides a preparation method of thiophilic chromatographic material, the preparation method comprising:

a, in the presence of an initiator, glycidyl methacrylate and methylenebisacrylamide are copolymerized in a porogen to obtain a copolymer poly(GMA-co-MBAA);

b. In the presence of the first organic solvent, the copolymer poly(GMA-co-MBAA) is subjected to ring-opening reaction with sulfonamide to prepare the copolymer poly(SFA-(GMA-co-MBAA)).

The present invention also provides a thiolophilic chromatographic material, which is prepared by the above method.

The present invention also provides an application of the above-mentioned thiolophilic chromatographic material in the enrichment and separation of antibodies.

Through the above-mentioned technical scheme, the present invention firstly carries out copolymerization reaction with glycidyl methacrylate and methylenebisacrylamide in the presence of an initiator to obtain a copolymer poly(GMA-co-MBAA); then sulfonamide and copolymerization The compound poly(GMA-co-MBAA) undergoes a ring-opening reaction, thereby modifying the sulfone-sulfide group to the copolymer poly(GMA-co-MBAA) to form a copolymer poly(SFA-(GMA-co-MBAA) ), the thiolophilic chromatographic material. The thiolophilic chromatographic material has a cross-linked network structure, high mechanical strength and good permeability. In the presence of high-concentration salt, the thiophilic chromatographic material can specifically adsorb to the appropriate site on the surface of the antibody, and in the presence of low-concentration salt, the antibody can be eluted from the thiophilic chromatographic material.

Other features and advantages of the present invention will be described in detail in the following detailed description.

Description of drawings

The accompanying drawings are used to provide a further understanding of the present invention, and constitute a part of the description, together with the following specific embodiments, are used to explain the present invention, but do not constitute a limitation to the present invention. In the attached picture:

Fig. 1 is the schematic diagram of the reaction principle of preparing copolymer poly(SFA-(GMA-co-MBAA)) in embodiment 1;

Fig. 2 is the Fourier transform-infrared absorption spectrogram of the thiolophilic chromatographic material A1 in detection example 1;

Fig. 3 is the scanning electron micrograph of thiolophilic chromatographic material A2 in detection example 2 under magnification 1300 times;

Fig. 4 is the scanning electron micrograph of thiophilic chromatographic material A2 in detection example 2 under magnification 5000 times;

Fig. 5 is the MALDI-TOF MS figure of thiol-phile chromatographic material A2 to IgY eluent in application example 3;

Fig. 6 is the electrophoresis diagram of thiophile chromatographic material A2 to IgY eluate in application example 4.

detailed description

Specific embodiments of the present invention will be described in detail below. It should be understood that the specific embodiments described here are only used to illustrate and explain the present invention, not to limit the present invention.

The present invention provides a kind of preparation method of sulfophilic chromatographic material, and described preparation method comprises:

a, in the presence of an initiator, glycidyl methacrylate (GMA) and methylenebisacrylamide (MBAA) are copolymerized in a porogen to obtain a copolymer poly(GMA-co-MBAA);

b. In the presence of the first organic solvent, the copolymer poly(GMA-co-MBAA) is subjected to ring-opening reaction with sulfonamide (SFA) to prepare the copolymer poly(SFA-(GMA-co-MBAA)).

In the present invention, initiator is any conventional initiator in the art, can be organic peroxide initiator, inorganic peroxide initiator, azo initiator or redox initiator, considering reaction temperature Mildness, preferably, the initiator is an azo initiator, in order to further improve the initiation efficiency, more preferably, the initiator is selected from azobisisobutyronitrile (AIBN), azobisisovaleronitrile One or more of (AMBN), dimethyl azobisisobutyrate (AIBME).

In the present invention, the specific type of porogen can be selected in a wide range, but in order to improve the yield of the copolymerization reaction, preferably, the porogen is selected from dimethyl sulfoxide, dodecanol, acetone, PEG One of -200 (polyethylene glycol with brand name PEG-200), PEG-300 (polyethylene glycol with brand name PEG-300) and PEG-10000 (polyethylene glycol with brand name PEG-1000) or more.

In step a of the present invention, the specific amount of each raw material can be selected within a wide range, but in order to improve the rate and productivity of the copolymerization reaction, preferably, relative to 1 weight part of glycidyl methacrylate, the The amount of methylenebisacrylamide is 1-5 parts by weight, the amount of the initiator is 0.1-5% by weight, and the amount of the porogen is 1-10 parts by weight.

In step a of the present invention, the specific reaction conditions of the copolymerization reaction can be selected in a wide range, but in order to improve the rate and the productivity of the copolymerization reaction, preferably, the copolymerization reaction satisfies the following conditions: the reaction temperature is 40-90 °C, the reaction time is 2-24h.

In step b of the present invention, the first organic solvent can be any organic solvent used in the ring-opening reaction in the art, but in order to improve the rate and yield of the ring-opening reaction, preferably, the first organic solvent One or more selected from acetonitrile, acetone, methanol and ethanol.

In step b of the present invention, the specific amount of each substance can be selected within a wide range, but in order to improve the rate and yield of the ring-opening reaction, preferably, relative to 1 weight part of the copolymer poly(GMA-co- MBAA), the amount of the sulfonamide is 0.5-2 parts by weight, and the amount of the first organic solvent is 10-50 parts by weight.

In step b of the present invention, the specific reaction conditions of the ring-opening reaction can be selected in a wide range, but in order to improve the rate and yield of the ring-opening reaction, preferably, the ring-opening reaction meets the following conditions: the reaction temperature is 65-75°C, the reaction time is 10-14h.

In the present invention, in order to remove the unreacted small molecular substance attached on the copolymer poly(GMA-co-MBAA), thereby improve the copolymer poly(SFA-(GMA-co-MBAA)) that the ring-opening reaction makes Purity, preferably, before step b, the method further includes: extracting or rinsing the copolymer poly(GMA-co-MBAA) with a second organic solvent. Wherein, the extraction can be performed by a Soxhlet extractor, and the washing can be performed by column chromatography.

In the present invention, in order to remove the unreacted small molecular substance attached on the copolymer poly(SFA-(GMA-co-MBAA)), further improve the purity of the copolymer poly(SFA-(GMA-co-MBAA)), Preferably, after step b, the method further includes: extracting or rinsing the copolymer poly(SFA-(GMA-co-MBAA)) with a third organic solvent. Wherein, the extraction can also be performed by a Soxhlet extractor, and the washing can also be performed by column chromatography.

In the above step of extraction or rinsing, the second organic solvent and the third organic solvent are conventional organic solvents in the art, and can be selected in a wide range, but in order to improve the removal effect of small molecular substances, preferably, the second The second organic solvent and the third organic solvent are each independently selected from one or more of methanol, acetonitrile and ethanol.

In the present invention, the reaction vessels of the copolymerization reaction and the ring-opening reaction can be conventional glass containers in the art, such as flasks, test tubes and chromatographic columns, but in order to obtain specific shapes of thiophilic chromatographic materials, preferably, the copolymerization reaction and ring-opening reactions were carried out in open glass containers or sealed capillary tubes. In this way, the thiolophilic chromatographic material in the open glass container is in the form of a block, and then it is ground into a powdered thiolophilic chromatographic material, which can be poured into the chromatographic column, thereby enriching and separate. Of course, in addition to block thiolophilic chromatographic materials, by carrying out copolymerization and ring-opening reactions in sealed capillary tubes, the thiolophilic chromatographic materials prepared in this way are fixed in capillaries, and such thiolophilic chromatographic materials can be carried out in capillary tubes. reuse.

In the above embodiment, in order to make the prepared thiolophilic chromatographic material more stably fixed in the capillary, preferably, before step a, the method further includes: cleaning the capillary, and then putting the cleaned capillary in The contact reaction is performed with 3-(methacryloyloxy)propyltrimethoxysilane (γ-MAPS) in the presence of the fourth organic solvent. In this way, as shown in Figure 1, the silane groups of γ-MAPS can react with the hydroxyl groups on the silicon oxide on the inner surface of the capillary, and then the carbon-carbon double bonds of γ-MAPS can react with the copolymer poly(GMA- co-MBAA) to react with the double bond on the copolymer poly(GMA-co-MBAA) in the capillary, and then the SFA and the copolymer poly(GMA-co-MBAA) undergo a ring-opening reaction, so that the preparation The obtained copolymer poly(SFA-(GMA-co-MBAA)) was tightly fixed in the capillary.

In the present invention, the fourth organic solution can be any one in the art, but in order to make the copolymer poly(SFA-(GMA-co-MBAA)) more stably located in the capillary, preferably, the fourth The organic solvent is selected from one or more of methanol, acetonitrile and ethanol.

When the capillary is processed, the amount of each substance can be selected in a wide range, but in order to improve the stability of the copolymer poly(SFA-(GMA-co-MBAA)), preferably, relative to 1 part by weight of 3 -(Methacryloyloxy)propyltrimethoxysilane, the amount of the fourth organic solvent is 0.5-10 parts by weight.

In the present invention, the specific conditions of the contact reaction can be selected in a wide range, but in order to make SFA react as much as possible with the hydroxyl groups on the silicon oxide on the inner surface of the capillary, preferably, the contact reaction satisfies the following conditions : The reaction temperature is 40-60°C, and the reaction time is 8-12h.

In addition, the cleaning of the capillary can be a conventional cleaning method in the art, such as pickling, alkali cleaning, distilled water and organic solvent cleaning, but in order to make the capillary clean more cleanly, preferably, the capillary is sequentially subjected to alkali washing, water washing, acid washing, etc. Washing, washing with water and cleaning with organic solvents.

The present invention provides a thiolophilic chromatographic material, which is prepared by the above method.

The present invention also provides an application of the above-mentioned thiolophilic chromatographic material in the enrichment and separation of antibodies.

The present invention will be described in detail below by way of examples. In the following examples, the infrared spectrum parameters are measured by a Fourier-infrared spectrometer (IR-21, Shimadzu Corporation, Japan), and the scanning electron microscope spectrum parameters and X-ray energy spectrum parameters are all measured by a field emission scanning electron microscope (S-4800, Japan Hitachi, Hitachi), mass spectrometry parameters were measured by matrix-assisted laser desorption ionization time-of-flight mass spectrometry (4800TOF/TOF, AB Sciex, USA), and electrophoresis parameters were measured by capillary electrophoresis (P/ACE ^tm MDQ, Beckman Corporation, USA) Measured.

Glycidyl methacrylate, methylenebisacrylamide, 3-(methacryloyloxy)propyltrimethoxysilane, and azobisisobutyronitrile were purchased from Aladdin and were purchased from Shanghai Jingchun Biochemical Technology Co., Ltd. The company's commercial products.

Aiming at the enrichment and separation ability of the thiolophilic chromatographic material, the thiolophilic chromatographic material is detected with egg yolk immunoglobulin (IgY) in the following test examples.

Example 1

1) Put 0.3g of GMA, 0.6g of MBA, 2g of DMSO and 1.74g of dodecanol in a centrifuge tube, vortex, ultrasonically degas, mix well, then add 10mg of initiator AIBN, vortex again Spinning and ultrasonication to obtain a homogeneous and clear solution, then co-polymerize at a constant temperature in a water bath at 75°C for 12 hours to obtain block material M1;

2) Take 3 g of the block material M1 and cut it into small pieces, put it into a Soxhlet extractor, add 50 mL of methanol, and extract at 110° C. for 24 hours to obtain the block material M2.

3) Place the block material M2 in a vacuum drying oven and dry at 100° C. for 12 hours to obtain the copolymer poly(GMA-co-MBAA).

4) 5 g of sulfonamide was dissolved in 10 ml of acetonitrile, and then reacted with 3 g of copolymer poly(GMA-co-MBAA) at a temperature of 70° C. for 12 hours to obtain block material M3.

5) Then put 3g of the bulk material M3 into the extractor, add 50mL of methanol, and extract at 110°C for 24 hours to obtain the copolymer poly(SFA-(GMA-co-MBAA)), that is, the thiolophilic chromatographic material A1 .

Example 2

1) Wash the capillary with NaOH (0.1M), water, HCl (0.1M), water and methanol for 30min in sequence. Then, the inner wall of the washed capillary was blown with nitrogen flow for 10 h to dry. At 50°C, place the dried capillary in a mixed solution of γ-MAPS and methanol (the volume ratio of γ-MAPS and methanol is 1:1, and the mass ratio is 1.3:1) for contact reaction for 10 h. After the reaction, use Rinse the capillary with methanol and dry it with nitrogen to obtain a clean capillary.

2) Put GMA (30mg), MBAA (60mg), DMSO (200mg), dodecanol (174mg) and AIBN (1mg) in a 1.5mL centrifuge tube, vortex and sonicate to obtain a uniform and clear solution. Then the solution was quickly injected into the above-mentioned clean capillary, sealed with silicone rubber at both ends, and subjected to a constant temperature copolymerization reaction in a water bath at 75° C. for 12 hours.

3) Take out the above-mentioned capillary tube, connect it to a high-pressure liquid chromatography pump, wash the capillary tube with methanol as the mobile phase to obtain the copolymer poly(GMA-co-MBAA).

4) Dissolve 25.0 mg of sulfonamide in 1 ml of acetonitrile, and continuously inject it into the above-mentioned capillary containing the copolymer poly(GMA-co-MBAA) at a temperature of 70°C with a high-pressure pump, and perform a ring-opening reaction for 12 hours. After the reaction, the capillary was taken out, connected to the high-pressure liquid chromatography pump, and the monolithic column was washed with acetonitrile and methanol in sequence to obtain the copolymer poly(SFA-(GMA-co-MBAA)), that is, the thiolophilic chromatography material A2.

Test example 1

The ^thiolophilic chromatographic material A1 was detected by a Fourier-infrared spectrometer (IR-21, Shimadzu Corporation, Japan). 657.7, 952.9cm ^-1 ) and other characteristic group absorption peaks, and then concluded that the sulfone-sulfide group has been successfully modified on the thiolophilic chromatography material A1.

Test example 2

Carry out X-ray elemental analysis and detection to thiophile chromatographic material A2 and copolymer poly(GMA-co-MBAA) in Example 2 by field emission scanning electron microscope (S-4800, Hitachi, Japan, Hitachi), and the detection results are shown in Table 1 , It can be seen from the table that, through step 4) of Example 2, the sulfonamide group has been successfully modified on the copolymer poly(GMA-co-MBAA).

Table 1

Test example 3

The thiolophilic chromatographic material A2 was detected by a field emission scanning electron microscope (S-4800, Hitachi, Japan), and the test results are shown in Figure 3 and Figure 4. It can be seen from the figure that the thiolophilic chromatographic material A2 is a highly cross-linked porous material , the material is a 3D skeleton structure material with a diameter of 30-70nm.

Preparation Example 1

Preparation of 1 mg/mL IgY standard solution: Dissolve 2 mg IgY in 2 ml of 0.5 M sodium sulfate aqueous solution at 25°C to prepare 1 mg/mL IgY standard solution.

Preparation example 2

Preparation of 1M citric acid solution: Dissolve 2.1014g of citric acid in an appropriate amount of distilled water at 25°C, and dilute to 10mL to prepare 1M citric acid aqueous solution.

Preparation example 3

Preparation of sample solution (CPB): Dissolve 0.7163g disodium hydrogen phosphate and 3.5510g sodium sulfate in 45mL of distilled water at 25°C, then adjust the pH to 6.2 with 1M citric acid aqueous solution, and then dilute to 50mL to obtain the sample solution.

Preparation Example 4

Preparation of the eluent (PB): at 25°C, 0.7163 g of disodium hydrogen phosphate and 45 mL of distilled water were adjusted to pH 6.2 with 1 M aqueous citric acid solution, and then the volume was adjusted to 50 mL to obtain the eluent.

Preparation Example 5

Preparation of MALDI-TOF MS matrix solution: prepared by mixing 100 mg of gallic acid (SA), 3 mL of acetonitrile and 7 mL of 0.1% trifluoroacetic acid at 25°C.

Preparation example 6

Preparation of electrophoretic separation buffer solution: Dissolve 1.9501g of disodium hydrogen phosphate, 0.7305g of sodium chloride, and 7.2095g of sodium lauryl sulfate in 240mL of distilled water, adjust the solution to pH 6.5 with 1M aqueous sodium hydroxide solution, and set Make up to 250mL to prepare electrophoresis separation buffer solution.

Application example 1

1) Inject 1 mL of the sample solution into the thiol-phile chromatography material A2 to equilibrate for 200 min, and then inject 50 μL of 1 mg/ml IgY standard solution for immobilization for 60 min;

2) After IgY is immobilized, inject 20 μL of the sample solution into the thiophile chromatography material A2 for washing, and obtain the residual solution that is not eluted after IgY enrichment;

3) Inject 6 μL of the eluent into the thiophile chromatographic material A2 to elute IgY to obtain the IgY eluent;

4) Mix 0.5 μL of IgY eluate with 0.5 μL of MALDI-TOF MS matrix solution, then drop onto the MALDI-TOF MS target plate, then dry, wash with ice water to remove saline, and finally use matrix-assisted laser Analytical ionization time-of-flight mass spectrometry (4800TOF/TOF, AB Sciex, USA) was used for mass spectrometry characterization. The characterization results are shown in Figure 5, in which curve a is the mass spectrum of the blank control group (0.5M _Na2SO4 solution ₎ , curve b is the mass spectrum of the 1mg/mL IgY standard solution, and curve c is the mass spectrum of the IgY enrichment The mass spectrogram of the residual liquid that has not been eluted, the d curve is the mass spectrogram of the IgY eluent, it can be seen from this figure that the thiophile chromatography material A2 can effectively enrich IgY in the presence of high concentration of salt, and at low Efficient elution of IgY in the presence of concentrated salt.

It can be seen from Figure 5 that using the MALDI-TOF MS method, it can be known that the thiol-philic chromatographic material A1 can effectively enrich IgY in the presence of high-concentration salt, and can wash IgY in the presence of low-concentration salt. come out.

Application example 2

1) Inject 1 mL of the sample solution into the thiol-phile chromatography material A2 to equilibrate for 200 min, and then inject 50 μL of 1 mg/ml IgY standard solution for immobilization for 60 min;

2) After IgY is immobilized, inject 20 μL of the sample solution into the thiophile chromatography material A2 for washing, and obtain the residual solution that is not eluted after IgY enrichment;

3) Inject 6 μL of the eluent into the thiophile chromatographic material A2 to elute IgY to obtain the IgY eluent;

4) The IgY eluate was detected by electrophoresis using a capillary electrophoresis instrument (P/ACE ^tm MDQ, Beckman Company, USA), and the specific detection conditions were: a fused silica capillary with an inner diameter of 75 μm (total length 56.5 cm, effective length 50cm); the infrared detection wavelength is 214nm, and the injection volume is 5s×0.5psi. The results of the electrophoresis test are shown in Figure 6, in which curve a is the electrophoresis of the blank control group (0.5M _Na2SO4 solution ₎ , curve b is the electrophoresis of the 1mg/mL IgY standard solution, and curve c is the enrichment of IgY The electropherogram of the residual liquid that has not been eluted after that, the d curve is the electropherogram of the IgY eluent. It can be seen from this figure that the thiophile chromatography material A2 can effectively enrich IgY in the presence of high-concentration salt. Efficient elution of IgY in the presence of low concentrations of salt.

Using this method, it can also be known that the thiol-philic chromatographic material A1 can effectively enrich IgY in the presence of high-concentration salt, and can elute IgY in the presence of low-concentration salt.

The preferred embodiments of the present invention have been described in detail above, but the present invention is not limited to the specific details in the above embodiments. Within the scope of the technical concept of the present invention, various simple modifications can be made to the technical solutions of the present invention. These simple modifications All belong to the protection scope of the present invention.

In addition, it should be noted that the various specific technical features described in the above specific embodiments can be combined in any suitable way if there is no contradiction. The combination method will not be described separately.

In addition, various combinations of different embodiments of the present invention can also be combined arbitrarily, as long as they do not violate the idea of the present invention, they should also be regarded as the disclosed content of the present invention.

Claims (16)

1. a kind of preparation method of thiophilic chromatography material, it is characterised in that the preparation method includes：

A, in the presence of initiator, GMA and methylene-bisacrylamide are carried out in pore-foaming agent Copolyreaction is obtained copolymer poly (GMA-co-MBAA)；

B, in the presence of the first organic solvent, copolymer poly (GMA-co-MBAA) is opened with P-aminobenzene-sulfonamide Ring reaction is obtained copolymer poly (SFA- (GMA-co-MBAA))；

Wherein, relative to the GMA of 1 weight portion, the consumption of the methylene-bisacrylamide is 1-5 Weight portion, the consumption of the initiator is 0.1-5% weight portions, and the consumption of the pore-foaming agent is 1-10 weight portions；Relative to 1 weight The copolymer poly (GMA-co-MBAA) of part is measured, the consumption of the P-aminobenzene-sulfonamide is 0.5-2 weight portions, described first The consumption of organic solvent is 10-50 weight portions.

2. preparation method according to claim 1, wherein, the initiator is selected from azodiisobutyronitrile, azo diisoamyl One or more in nitrile, azo-bis-iso-dimethyl.

3. preparation method according to claim 1, wherein, the pore-foaming agent is selected from dimethyl sulfoxide, lauryl alcohol, acetone, board One kind or many in the polyethylene glycol of the polyethylene glycol, the polyethylene glycol of trade mark PEG-300 and trade mark PEG-1000 of number PEG-200 Kind.

4. the preparation method according to any one in claim 1-3, wherein, the copolyreaction meets following condition： Reaction temperature is 40-90 DEG C, and the reaction time is 2-24h.

5. preparation method according to claim 4, wherein, first organic solvent is selected from acetonitrile, methyl alcohol and ethanol One or more.

6. the preparation method according to any one in claim 1-3, wherein, the ring-opening reaction meets following condition： Reaction temperature is 65-75 DEG C, and the reaction time is 10-14h.

7. the preparation method according to any one in claim 1-3, wherein, before step b, methods described is also wrapped Include：Second organic solvent is extracted or drip washing to copolymer poly (GMA-co-MBAA).

8. the preparation method according to any one in claim 1-3, wherein, after step b, methods described is also wrapped Include：3rd organic solvent is extracted or drip washing to copolymer poly (SFA- (GMA-co-MBAA)).

9. the preparation method according to any one in claim 1-3, wherein, second organic solvent and the described 3rd Organic solvent is each independently selected from one or more in methyl alcohol, acetonitrile and ethanol.

10. the preparation method according to any one in claim 1-3, wherein, the copolyreaction and ring-opening reaction exist Carried out in open glass container or the capillary of sealing.

11. preparation methods according to claim 10, wherein, before step a, methods described also includes：By capillary Cleaned, then by the capillary after cleaning in the presence of the 4th organic solvent with 3- (methacryloxypropyl) propyl group front three TMOS carries out haptoreaction.

12. preparation methods according to claim 11, wherein, the 4th organic solvent be selected from acetonitrile, acetone, methyl alcohol and One or more in ethanol.

13. preparation method according to claim 11 or 12, wherein, relative to the 3- (methacryloxypropyl) of 1 weight portion Propyl trimethoxy silicane, the consumption of the 4th organic solvent is 0.5-10 weight portions.

14. preparation methods according to claim 13, wherein, the haptoreaction meets following condition：Reaction temperature is 40-60 DEG C, the reaction time is 8-12h.

15. a kind of thiophilic chromatography materials, it is characterised in that the thiophilic chromatography material is by any one in claim 1-14 Method described in is prepared.

A kind of 16. thiophilic chromatography materials according to claim 15 antibody enrichment with separate in application.