Boron affinity chromatography medium and preparation method thereof

Abstract

The present invention provides a boron affinity chromatography medium and a preparation method thereof. The boron affinity chromatography medium comprises a silica gel coated with poly(styrene-divinylbenzene) as a matrix, 3-acrylamidophenylboronic acid is grafted onto the matrix, and then modified with 6-aminopyridine boronic acid to obtain a boron affinity chromatography medium. The present invention can increase the surface binding sites of the matrix and improve the adsorption capacity of the chromatography medium by coating the silica gel surface with poly(styrene-divinylbenzene). Meanwhile, 6-aminopyridine boronic acid is used for modification to increase the adsorption sites of the chromatography medium, thereby further improving the adsorption capacity.

Description

Technical Field

[0001] The present invention relates to chromatography separation technology, in particular to a boron affinity chromatography medium and a preparation method thereof. Background Art

[0002] Boron affinity materials are a new type of affinity material developed based on the principle that phenylboronic acid undergoes structural changes in different pH environments, allowing it to specifically bind to cis-dihydroxy groups. They have the ability to enrich, extract, and separate various biomolecules containing cis-dihydroxy structures, such as carbohydrates, glycoproteins, and nucleotides. In physiological and slightly alkaline environments, they can reversibly react with cis-ortho-dihydroxy groups to form five- or six-membered cyclic esters. However, because most current boron affinity materials are prepared using traditional bonding reactions of bifunctional compounds, they have limited adsorption sites and low adsorption capacity.

[0003] In addition, under the action of hydrophobic forces caused by high concentrations of salt ions, the binding performance of boron affinity materials with substances containing cis-vicinal dihydroxy groups is significantly reduced, and they are only suitable for chromatographic separation of low concentration salt ion solutions. Therefore, their scope of application is limited. Summary of the Invention

[0004] The object of the present invention is to provide a boron affinity chromatography medium and a preparation method thereof, so as to solve the problems of low adsorption capacity and narrow application range of the boron affinity chromatography medium in the prior art.

[0005] In order to solve the above technical problems, the technical solutions of the present invention are as follows:

[0006] A boron affinity chromatography medium is prepared by using silica gel coated with poly(styrene-divinylbenzene) as a matrix, grafting 3-acrylamidophenylboronic acid onto the matrix, and then modifying the matrix with 6-aminopyridineboronic acid to obtain the boron affinity chromatography medium.

[0007] Preferably, the silica gel with the surface coated with poly(styrene-divinylbenzene) is obtained by mixing activated silica gel with poly(styrene-divinylbenzene) and then curing.

[0008] Preferably, the mass ratio of the activated silica gel to poly(styrene-divinylbenzene) is 10:(3-5).

[0009] Preferably, the activated silica gel is acid-activated silica gel.

[0010] Preferably, the activated silica gel is obtained by the following method: taking silica gel, adding 5.5-6.0 mol / L hydrochloric acid thereto, heating and refluxing for 5-6 hours, filtering and drying to obtain activated silica gel.

[0011] Preferably, the mass ratio of the matrix to 3-acrylamidophenylboronic acid is 1:(1.8-2.8).

[0012] Preferably, the mass ratio of the matrix to the 6-aminopyridine boronic acid is 1:(0.1-0.5).

[0013] The present invention also provides a method for preparing a boron affinity chromatography medium, comprising the following steps:

[0014] (1) 3-acrylamidophenylboronic acid is added to a silica gel coated with poly(styrene-divinylbenzene) to obtain a grafted silica gel after reaction;

[0015] (2) adding 6-aminopyridine boronic acid to the grafted silica gel obtained in step (1), and reacting to obtain a boron affinity chromatography medium.

[0016] Preferably, the silica gel with a surface covered with poly(styrene-divinylbenzene) is obtained by the following method: taking silica gel, adding 5.5-6.0 mol / L hydrochloric acid thereto, heating and refluxing for 5-6 hours, filtering and drying to obtain activated silica gel; adding the activated silica gel to a poly(styrene-divinylbenzene) solution, stirring at 60-80°C until the volatile components are volatilized, to obtain silica gel with a surface covered with poly(styrene-divinylbenzene).

[0017] Preferably, step (1) specifically comprises: dispersing silica gel coated with poly(styrene-divinylbenzene) in toluene, adding 4-chloromethylphenyltrimethoxysilane thereto, reacting at 100-120° C. for 8-16 hours, filtering to obtain silica gel with a surface-fixed initiator; dispersing the silica gel with the surface-fixed initiator in N,N-dimethylformamide, adding 3-acrylamidophenylboronic acid, 2,2'-bipyridine and cuprous bromide thereto, stirring and reacting at 90-100° C. for 5-10 hours to obtain the grafted silica gel.

[0018] Preferably, the amount of the poly(styrene-divinylbenzene)-coated silica gel to toluene is 0.8-1.5 g per 10 ml of toluene. The mass ratio of the poly(styrene-divinylbenzene)-coated silica gel to 4-chloromethylphenyltrimethoxysilane is 1:(1.2-1.5). The amount of the silica gel as the surface-immobilized initiator to the N,N-dimethylformamide is 1.2-1.8 g per 10 ml of N,N-dimethylformamide. The mass ratio of the silica gel, 2,2'-bipyridine, and cuprous bromide as the surface-immobilized initiator is 1:0.5:0.3.

[0019] Preferably, step (2) specifically comprises: adding 6-aminopyridine boronic acid to the grafted silica gel obtained in step (1), stirring and reacting at 110-130° C. for 2-8 hours to obtain a boron affinity chromatography medium.

[0020] The above solution of the present invention includes at least the following beneficial effects:

[0021] (1) The boron affinity chromatography medium of the present invention is based on a silica gel coated with poly(styrene-divinylbenzene) as a matrix, 3-acrylamidophenylboronic acid is grafted onto the matrix, and then modified with 6-aminopyridine boronic acid to obtain a boron affinity chromatography medium. The present invention can increase the surface binding sites of the matrix and improve the adsorption capacity of the chromatography medium by coating the silica gel surface with poly(styrene-divinylbenzene). At the same time, for the silica gel matrix, its residual silanol groups are easy to adsorb amino substances, which leads to poor selectivity. By coating the silica gel with poly(styrene-divinylbenzene), the influence of its silanol groups can be reduced to improve the selectivity of the chromatography medium. In addition, the present invention uses 6-aminopyridine boronic acid for modification, which also increases the adsorption sites of the chromatography medium and can further improve the adsorption capacity of the chromatography medium.

[0022] (2) The boron affinity chromatography medium of the present invention can maintain a good chromatographic separation effect in a high-concentration salt ion solution and has a wider range of applications. DETAILED DESCRIPTION

[0023] In the examples of the present invention, if specific conditions are not specified, the experiments were carried out under conventional conditions or the conditions recommended by the manufacturer. Reagents or instruments used without specifying the manufacturer are all commercially available conventional products. Raw materials of different manufacturers and types do not affect the implementation of the technical solutions of the present invention and the achievement of the technical effects.

[0024] Example 1

[0025] The boron affinity chromatography medium in this example is based on a silica gel coated with poly(styrene-divinylbenzene) as a matrix, 3-acrylamidophenylboronic acid is grafted onto the matrix, and then modified with 6-aminopyridineboronic acid to obtain a boron affinity chromatography medium.

[0026] The silica gel coated with poly(styrene-divinylbenzene) is obtained by mixing activated silica gel with poly(styrene-divinylbenzene) and curing the mixture. The activated silica gel is acid-activated silica gel. The mass ratio of the activated silica gel to poly(styrene-divinylbenzene) is 10:3. The matrix, i.e., the silica gel coated with poly(styrene-divinylbenzene), has a mass ratio of 1:2.8 to 3-acrylamidophenylboronic acid. The mass ratio of the matrix to 6-aminopyridineboronic acid is 1:0.3.

[0027] The method for preparing the boron affinity chromatography medium described in this embodiment comprises the following steps:

[0028] (1) Silica gel coated with poly(styrene-divinylbenzene) is dispersed in toluene, 4-chloromethylphenyltrimethoxysilane is added thereto, the mixture is reacted at 120°C for 8 hours, and filtered to obtain silica gel with a surface-fixed initiator; the silica gel with the surface-fixed initiator is dispersed in N,N-dimethylformamide, 3-acrylamidophenylboronic acid, 2,2'-bipyridine, and cuprous bromide are added thereto, the mixture is stirred and reacted at 90°C for 10 hours, and filtered to obtain a grafted silica gel.

[0029] The amount of the poly(styrene-divinylbenzene)-coated silica gel to toluene is 0.8 g per 10 ml of toluene. The mass ratio of the poly(styrene-divinylbenzene)-coated silica gel to 4-chloromethylphenyltrimethoxysilane is 1:1.2. The amount of the surface-immobilized initiator silica gel to N,N-dimethylformamide is 1.2 g per 10 ml of N,N-dimethylformamide. The mass ratio of the surface-immobilized initiator silica gel, 2,2'-bipyridine, and cuprous bromide is 1:0.5:0.3.

[0030] In this embodiment, the silica gel with a surface covered with poly(styrene-divinylbenzene) is obtained by the following method: taking silica gel, adding 5.5 mol / L hydrochloric acid thereto, heating reflux for 6 hours, filtering and drying to obtain activated silica gel; adding the activated silica gel to a poly(styrene-divinylbenzene) solution, stirring at 60°C until the volatile components are evaporated, so that the poly(styrene-divinylbenzene) is solidified on the surface of the silica gel, and obtaining silica gel with a surface covered with poly(styrene-divinylbenzene).

[0031] The volume ratio of the silica gel to the hydrochloric acid is at least 1:5. The poly(styrene-divinylbenzene) solution is an aqueous solution with a concentration of 0.5 mg / mL.

[0032] (2) Add 6-aminopyridine boronic acid to the grafted silica gel obtained in step (1), and stir the reaction at 130° C. for 2 h to obtain a boron affinity chromatography medium.

[0033] Example 2

[0034] The boron affinity chromatography medium in this example is based on a silica gel coated with poly(styrene-divinylbenzene) as a matrix, 3-acrylamidophenylboronic acid is grafted onto the matrix, and then modified with 6-aminopyridineboronic acid to obtain a boron affinity chromatography medium.

[0035] The poly(styrene-divinylbenzene)-coated silica gel is obtained by mixing activated silica gel with poly(styrene-divinylbenzene) and curing the mixture. The activated silica gel is acid-activated silica gel. The mass ratio of the activated silica gel to poly(styrene-divinylbenzene) is 10:4. The mass ratio of the matrix to 3-acrylamidophenylboronic acid is 1:1.8. The mass ratio of the matrix to 6-aminopyridineboronic acid is 1:0.5.

[0036] The method for preparing the boron affinity chromatography medium described in this embodiment comprises the following steps:

[0037] (1) Silica gel coated with poly(styrene-divinylbenzene) is dispersed in toluene, 4-chloromethylphenyltrimethoxysilane is added thereto, the mixture is reacted at 100°C for 12 hours, and filtered to obtain silica gel with a surface-fixed initiator; the silica gel with the surface-fixed initiator is dispersed in N,N-dimethylformamide, 3-acrylamidophenylboronic acid, 2,2'-bipyridine, and cuprous bromide are added thereto, the mixture is stirred and reacted at 100°C for 7 hours to obtain a grafted silica gel.

[0038] The amount of the poly(styrene-divinylbenzene)-coated silica gel to toluene is 1.5 g per 10 ml of toluene. The mass ratio of the poly(styrene-divinylbenzene)-coated silica gel to 4-chloromethylphenyltrimethoxysilane is 1:1.3. The amount of the silica gel as the surface-immobilized initiator to N,N-dimethylformamide is 1.8 g per 10 ml of N,N-dimethylformamide. The mass ratio of the silica gel, 2,2'-bipyridine, and cuprous bromide as the surface-immobilized initiator is 1:0.5:0.3.

[0039] In this embodiment, the silica gel with a surface covered with poly(styrene-divinylbenzene) is obtained by the following method: taking silica gel, adding 6.0 mol / L hydrochloric acid thereto, heating reflux for 5 hours, filtering and drying to obtain activated silica gel; adding the activated silica gel to a poly(styrene-divinylbenzene) solution, stirring at 70°C until the volatile components are evaporated, so that the poly(styrene-divinylbenzene) is solidified on the surface of the silica gel, and obtaining silica gel with a surface covered with poly(styrene-divinylbenzene).

[0040] The volume ratio of the silica gel to the hydrochloric acid is at least 1:5. The poly(styrene-divinylbenzene) solution is an aqueous solution with a concentration of 0.5 mg / mL.

[0041] (2) Add 6-aminopyridine boronic acid to the grafted silica gel obtained in step (1), and stir the reaction at 120° C. for 5 h to obtain a boron affinity chromatography medium.

[0042] Example 3

[0043] The boron affinity chromatography medium in this example is based on a silica gel coated with poly(styrene-divinylbenzene) as a matrix, 3-acrylamidophenylboronic acid is grafted onto the matrix, and then modified with 6-aminopyridineboronic acid to obtain a boron affinity chromatography medium.

[0044] The poly(styrene-divinylbenzene)-coated silica gel is obtained by mixing activated silica gel with poly(styrene-divinylbenzene) and curing the mixture. The activated silica gel is acid-activated silica gel. The mass ratio of the activated silica gel to poly(styrene-divinylbenzene) is 10:5. The mass ratio of the matrix to 3-acrylamidophenylboronic acid is 1:2. The mass ratio of the matrix to 6-aminopyridineboronic acid is 1:0.1.

[0045] The method for preparing the boron affinity chromatography medium described in this embodiment comprises the following steps:

[0046] (1) Silica gel coated with poly(styrene-divinylbenzene) is dispersed in toluene, 4-chloromethylphenyltrimethoxysilane is added thereto, the mixture is reacted at 110°C for 16 hours, and filtered to obtain silica gel with a surface-fixed initiator; the silica gel with the surface-fixed initiator is dispersed in N,N-dimethylformamide, 3-acrylamidophenylboronic acid, 2,2'-bipyridine, and cuprous bromide are added thereto, the mixture is stirred and reacted at 95°C for 5 hours to obtain a grafted silica gel.

[0047] The amount of the poly(styrene-divinylbenzene)-coated silica gel to toluene is 1.2 g per 10 ml of toluene. The mass ratio of the poly(styrene-divinylbenzene)-coated silica gel to 4-chloromethylphenyltrimethoxysilane is 1:1.5. The amount of the silica gel as the surface-immobilized initiator to N,N-dimethylformamide is 1.5 g per 10 ml of N,N-dimethylformamide. The amount of the silica gel, 2,2'-bipyridine, and cuprous bromide as the surface-immobilized initiator is 1:0.5:0.3.

[0048] In this embodiment, the silica gel with a surface covered with poly(styrene-divinylbenzene) is obtained by the following method: taking silica gel, adding 5.8 mol / L hydrochloric acid thereto, heating reflux for 6 hours, filtering and drying to obtain activated silica gel; adding the activated silica gel to a poly(styrene-divinylbenzene) solution, stirring at 80°C until the volatile components are volatilized, so that the poly(styrene-divinylbenzene) is solidified on the surface of the silica gel, and obtaining silica gel with a surface covered with poly(styrene-divinylbenzene).

[0049] The volume ratio of the silica gel to the hydrochloric acid is at least 1:5. The poly(styrene-divinylbenzene) solution is an aqueous solution with a concentration of 0.5 mg / mL.

[0050] (2) Add 6-aminopyridine boronic acid to the grafted silica gel obtained in step (1), and stir the reaction at 110° C. for 8 h to obtain a boron affinity chromatography medium.

[0051] Example 4

[0052] The boron affinity chromatography medium in this example is based on a silica gel coated with poly(styrene-divinylbenzene) as a matrix, 3-acrylamidophenylboronic acid is grafted onto the matrix, and then modified with 6-aminopyridineboronic acid to obtain a boron affinity chromatography medium.

[0053] The poly(styrene-divinylbenzene)-coated silica gel is obtained by mixing activated silica gel with poly(styrene-divinylbenzene) and curing the mixture. The activated silica gel is acid-activated silica gel. The mass ratio of the activated silica gel to poly(styrene-divinylbenzene) is 10:3.8. The mass ratio of the matrix to 3-acrylamidophenylboronic acid is 1:2.4. The mass ratio of the matrix to 6-aminopyridineboronic acid is 1:0.3.

[0054] The method for preparing the boron affinity chromatography medium described in this embodiment comprises the following steps:

[0055] (1) Silica gel coated with poly(styrene-divinylbenzene) is dispersed in toluene, 4-chloromethylphenyltrimethoxysilane is added thereto, the mixture is reacted at 110°C for 16 hours, and filtered to obtain silica gel with a surface-fixed initiator; the silica gel with the surface-fixed initiator is dispersed in N,N-dimethylformamide, 3-acrylamidophenylboronic acid, 2,2'-bipyridine, and cuprous bromide are added thereto, the mixture is stirred and reacted at 95°C for 8 hours to obtain a grafted silica gel.

[0056] The amount of the poly(styrene-divinylbenzene)-coated silica gel to toluene is 1.2 g per 10 ml of toluene. The mass ratio of the poly(styrene-divinylbenzene)-coated silica gel to 4-chloromethylphenyltrimethoxysilane is 1:1.4. The amount of the silica gel as the surface-immobilized initiator to N,N-dimethylformamide is 1.2 g per 10 ml of N,N-dimethylformamide. The mass ratio of the silica gel, 2,2'-bipyridine, and cuprous bromide to the surface-immobilized initiator is 1:0.5:0.3.

[0057] In this embodiment, the silica gel with a surface covered with poly(styrene-divinylbenzene) is obtained by the following method: taking silica gel with a particle size of 5 μm, adding 5.8 mol / L hydrochloric acid thereto, heating and refluxing for 6 hours, filtering and drying to obtain activated silica gel; adding the activated silica gel to a poly(styrene-divinylbenzene) solution, stirring at 70°C until the volatile components are volatilized, so that the poly(styrene-divinylbenzene) is solidified on the surface of the silica gel, and obtaining silica gel with a surface covered with poly(styrene-divinylbenzene).

[0058] The volume ratio of the silica gel to the hydrochloric acid is at least 1:5. The poly(styrene-divinylbenzene) solution is an aqueous solution with a concentration of 0.5 mg / mL.

[0059] (2) Add 6-aminopyridine boronic acid to the grafted silica gel obtained in step (1), and stir the reaction at 110° C. for 6 h to obtain a boron affinity chromatography medium.

[0060] Comparative Example 1

[0061] In this comparative example, the boron affinity chromatography medium was prepared by the same method as in Example 4, except that silica gel was directly used as the matrix, that is, the silica gel coated with poly(styrene-divinylbenzene) was replaced by silica gel.

[0062] Comparative Example 2

[0063] In this comparative example, the boron affinity chromatography medium was prepared by the same method as in Example 4, except that 6-aminopyridine boronic acid was not used for modification, that is, step (2) was omitted, and the grafted silica gel obtained in step (1) was used as the boron affinity chromatography medium.

[0064] Comparative Example 3

[0065] In this comparative example, a boron affinity chromatography medium was prepared by the same method as in Example 4, except that the mass ratio of the activated silica gel to the poly(styrene-divinylbenzene) in the silica gel coated with the poly(styrene-divinylbenzene) was 10:2.

[0066] Comparative Example 4

[0067] In this comparative example, a boron affinity chromatography medium was prepared by the same method as in Example 4, with the only difference being that in the silica gel coated with poly(styrene-divinylbenzene), the mass ratio of the activated silica gel to the poly(styrene-divinylbenzene) was 10:6.

[0068] Comparative Example 5

[0069] In this comparative example, the boron affinity chromatography medium was prepared by the same method as in Example 4, except that the mass ratio of the matrix to the 6-aminopyridine boronic acid was 1:0.6.

[0070] Effect Experiment Example

[0071] To verify the technical effect of the boron affinity chromatography medium of the present invention, the following experiments were performed:

[0072] The boron affinity chromatography media prepared in Examples 1-4 and Comparative Examples 1-5 were tested for adsorption capacity: catechol, a substance containing cis-o-dihydroxyl groups, was used as the target adsorbent, and the adsorption capacity of each boron affinity chromatography medium for the target adsorbent was determined by ultraviolet high-performance liquid chromatography.

[0073] The boron affinity chromatography media prepared in Examples 1-4 and Comparative Examples 1-5 were used to test adsorption selectivity: a mixed solution of catechol, aniline, hydroquinone, and 5-hydroxytryptamine was prepared in a 50 mmol / L NH4Cl-NH3 buffer solution with a pH of 8.5; the concentrations of catechol, aniline, hydroquinone, and 5-hydroxytryptamine in the mixed solution were all 5 μg / mL. The mixed solution was subjected to adsorption extraction using the boron affinity chromatography media prepared in Examples 1-4 and Comparative Examples 1-5, followed by elution. The eluate was analyzed by ultraviolet high-performance liquid chromatography, and the purity of the catechol in the eluate was calculated.

[0074] The boron affinity chromatography media prepared in Examples 1-4 and Comparative Examples 1-5 were tested for adsorption capacity at different ion concentrations: fructose, a substance containing cis-ortho-dihydroxy groups, was used as the target adsorbent. Fructose was added to a 0.01 mol / L phosphate buffered saline solution to obtain a low ion concentration loading solution. Fructose was added to a 0.1 mol / L phosphate buffered saline solution to obtain a high ion concentration loading solution. The fructose concentration in each loading solution was 5 μg / mL. The adsorption capacity of each boron affinity chromatography media for catechol in the above loading solutions was determined using ultraviolet high-performance liquid chromatography.

[0075] Effect experiment results:

[0076] After experimentation, the results are as follows:

[0077]

[0078]

[0079] According to the contrast of embodiment 1-4 and comparative example 1, boron affinity chromatography medium of the present invention adopts surface covered with poly (styrene-divinylbenzene) as matrix, not only significantly improves the adsorption capacity of chromatography medium, and has higher adsorption selectivity, in the solution of high concentration salt ion, still can efficiently adsorb target adsorbent. According to the contrast of embodiment 4 and comparative example 3-4, described surface is covered with in the silica gel of poly (styrene-divinylbenzene), the consumption of poly (styrene-divinylbenzene) is too little, can't give full play to its effect. And the consumption of poly (styrene-divinylbenzene) is too much, then the adsorption selectivity of the chromatography medium obtained only slightly raises, but adsorption capacity declines instead, especially in the solution of high concentration salt ion, adsorption capacity declines obviously. According to the contrast of embodiment 1-4 and comparative example 2, adopt 6-aminopyridine boric acid to carry out modification, the impact on adsorption selectivity is very little, but can significantly increase the adsorption site of chromatography medium, show higher adsorption capacity, in the solution of high concentration salt ion, also can avoid the decline of adsorption capacity to a certain extent. According to the comparison between Example 4 and Comparative Example 5, it can be seen that excessive use of 6-aminopyridine boronic acid has no significant effect on increasing the adsorption capacity, but instead leads to a decrease in its adsorption capacity in a solution with a high concentration of salt ions.

[0080] It is understood from common technical knowledge that the present invention may be implemented by other embodiments that do not depart from its spirit or essential features. Therefore, the embodiments disclosed above are, in all respects, merely illustrative and not exclusive. All modifications within the scope of the present invention or equivalent to the scope of the present invention are intended to be encompassed by the present invention.

Claims

1. A boron affinity chromatography medium, characterized in that Silica gel coated with poly(styrene-divinylbenzene) is used as a matrix, 3-acrylamidophenylboronic acid is grafted onto the matrix, and then modified with 6-aminopyridineboronic acid to obtain a boron affinity chromatography medium.

2. The boron affinity chromatography medium according to claim 1, characterized in that The silica gel with the poly(styrene-divinylbenzene) coated on the surface is obtained by mixing activated silica gel with poly(styrene-divinylbenzene) and then solidifying.

3. The boron affinity chromatography medium according to claim 2, characterized in that The mass ratio of the activated silica gel to poly(styrene-divinylbenzene) is 10:(3-5).

4. The boron affinity chromatography medium according to claim 3, characterized in that The activated silica gel is acid-activated silica gel.

5. The boron affinity chromatography medium according to claim 1, characterized in that The mass ratio of the matrix to 3-acrylamidophenylboronic acid is 1:(1.8-2.8).

6. The boron affinity chromatography medium according to claim 1, characterized in that The mass ratio of the matrix to the 6-aminopyridine boronic acid is 1:(0.1-0.5).

7. A method for preparing a boron affinity chromatography medium, characterized in that: The steps include: (1) 3-acrylamidophenylboronic acid is added to a silica gel coated with poly(styrene-divinylbenzene) to obtain a grafted silica gel after reaction; (2) adding 6-aminopyridine boronic acid to the grafted silica gel obtained in step (1), and reacting to obtain a boron affinity chromatography medium.

8. The method for preparing a boron affinity chromatography medium according to claim 7, wherein: The silica gel with a surface covered with poly(styrene-divinylbenzene) is obtained by the following method: taking silica gel, adding 5.5-6.0 mol / L hydrochloric acid thereto, heating and refluxing for 5-6 hours, filtering and drying to obtain activated silica gel; adding the activated silica gel to a poly(styrene-divinylbenzene) solution, stirring at 60-80° C. until the volatile components are volatilized, to obtain silica gel with a surface covered with poly(styrene-divinylbenzene).

9. The method for preparing a boron affinity chromatography medium according to claim 7, wherein: Step (1) specifically comprises: dispersing silica gel coated with poly(styrene-divinylbenzene) in toluene, adding 4-chloromethylphenyltrimethoxysilane thereto, reacting at 100-120° C. for 8-16 hours, filtering to obtain silica gel with a surface-fixed initiator; dispersing the silica gel with the surface-fixed initiator in N,N-dimethylformamide, adding 3-acrylamidophenylboronic acid, 2,2'-bipyridine and cuprous bromide thereto, stirring and reacting at 90-100° C. for 5-10 hours, and obtaining grafted silica gel.

10. The method for preparing a boron affinity chromatography medium according to claim 7, wherein: Step (2) specifically comprises: adding 6-aminopyridine boronic acid to the grafted silica gel obtained in step (1), stirring and reacting at 110-130° C. for 2-8 hours to obtain a boron affinity chromatography medium.

Images