Magnetic polymer microsphere containing p-toluenesulfonyl as well as preparation method and application of magnetic polymer microsphere

Abstract

The invention provides a paratoluensulfonyl-containing magnetic polymer microsphere as well as a preparation method and application thereof. The preparation method comprises the following steps: firstly, coating the surface of the magnetic polymer microsphere with a benzene ring-containing polymer; then sequentially carrying out chloromethyl modification, hydroxyl modification and p-toluenesulfonyl chloride modification on the magnetic polymer microspheres coated with the benzene ring polymer, so as to obtain magnetic polymer microspheres containing p-toluenesulfonyl; the preparation method has the advantage of high group utilization rate, and the prepared paratoluensulfonyl-containing magnetic polymer microspheres have the advantages of high detection accuracy, low physical adsorption of non-target objects and the like, and can be applied to multiple aspects of targeted capture, separation and purification and the like.

Description

Technical Field

[0001] The present invention belongs to the technical field of magnetic microspheres, and particularly relates to a magnetic polymer microsphere containing tosyl group, and a preparation method and application thereof. Background Art

[0002] Magnetic microspheres are composite materials composed of polymer-coated magnetic nanoparticles, which have superparamagnetism and are prone to magnetic response under the action of an external magnetic field; in addition, magnetic particles with superparamagnetism are usually composed of oxides of iron, cobalt, nickel or added with various other elements such as copper, strontium, barium, etc. However, iron, cobalt, and nickel particles are not friendly to the biological environment, so iron oxide particles in the form of magnetite or maghemite are often used in the biomedical field. Due to the high surface energy caused by the existence of van der Waals forces and dipole-dipole attraction trends of magnetic particles and their biological sensitivity to oxygen, acidity and alkalinity, salt components, humidity, etc. in the environment, it is necessary to modify their surfaces. Magnetic microspheres show versatility, high resolution and good reactivity in the introduction of various functional groups such as hydroxyl, carboxyl, amino, mercapto, etc., providing possibilities for diverse bioactive couplings, and are therefore widely used in fields such as in vitro diagnosis, immunoprecipitation, cell separation, drug purification, chemiluminescence, etc.

[0003] CN116643042A discloses a preparation method of carboxyl magnetic beads. Sodium carboxymethyl cellulose, ferric salt and ferrous salt are dissolved in a mixed solution of ethylene glycol and ethylenediamine to obtain Fe3O4 nanoparticles. Then, tetraethyl orthosilicate is added under alkaline conditions, and a silica coating layer is obtained after a polymerization reaction. Subsequently, vinyltriethoxysilane and cyclohexane carboxylic anhydride are used to prepare carboxylated silane-modified Fe3O4 nanoparticles to obtain carboxyl superparamagnetic particles. The reagents used in this method are easily available, but it is difficult to adjust the size of the magnetic particles, and the carboxyl activation sites are not easy to control.

[0004] CN111013504A provides a method for modifying amino groups on the surface of magnetic microspheres coated with glycidyl methacrylate. First, oleic acid-coated Fe3O4 magnetic fluid is prepared, and then an initiator, Fe3O4 magnetic fluid and a monomer are mixed to form an organic phase, and an aqueous solution of polyvinyl alcohol and sodium chloride is used as the aqueous phase. The two are mixed evenly and a polymerization reaction occurs at a certain temperature to obtain magnetic polymer microspheres containing epoxy groups. The microspheres are suspended in a mixed solvent of tetrahydrofuran and ethanol containing lithium hydroxide, and ethylenediamine is slowly added under an ice-water bath, and the reaction is carried out overnight. After washing, magnetic polymer microspheres with amino groups on the surface are obtained. However, this method has high requirements for temperature changes and it is not easy to control the content of amino groups.

[0005] Therefore, developing a magnetic polymer microsphere containing tosyl group with high group utilization rate and high detection accuracy is still a technical problem urgently to be solved in this field. Summary of the Invention

[0006] Aiming at the deficiencies of the prior art, the purpose of the present invention is to provide a magnetic polymer microsphere containing a p-toluenesulfonyl group, a preparation method thereof and an application. The preparation method coats a polymer containing a benzene ring on the surface of the magnetic polymer microsphere, and then carries out p-toluenesulfonyl chloride modification in sequence, and finally obtains a magnetic polymer microsphere containing a p-toluenesulfonyl group with a group content and group distribution meeting the usage requirements, which has good application prospects in application directions such as separation and purification.

[0007] To achieve this purpose, the present invention adopts the following technical solutions:

[0008] In the first aspect, the present invention provides a preparation method of a magnetic polymer microsphere containing a p-toluenesulfonyl group. The preparation method includes: coating the magnetic polymer microsphere with a polymer containing a benzene ring, and then carrying out chloromethyl modification, hydroxyl modification and p-toluenesulfonyl chloride modification in sequence to obtain the magnetic polymer microsphere containing a p-toluenesulfonyl group.

[0009] The preparation method provided by the present invention first coats the surface of the magnetic polymer microsphere with a polymer containing a benzene ring, which can not only reduce the leakage of magnetic particles, but also graft a polymer chain segment with a certain molecular weight on the surface of the magnetic polymer microsphere. The number and molecular weight of the polymer chain segments can be tuned, and polymer chain segments of different lengths can be introduced for combination. Then, the magnetic polymer microsphere coated with the benzene ring polymer is subjected to chloromethyl modification, hydroxyl modification and p-toluenesulfonyl chloride modification in sequence to obtain the magnetic polymer microsphere containing a p-toluenesulfonyl group. This preparation method has the advantage of high group utilization rate, and the obtained magnetic polymer microsphere containing a p-toluenesulfonyl group has the advantages of high detection accuracy and low physical adsorption of non-target substances, and can be applied in many aspects such as targeted capture and separation and purification.

[0010] Preferably, the magnetic polymer microsphere is prepared by the following method, and the method includes:

[0011] (1A) Reacting a polymer microsphere, an epoxy monomer, a functional monomer, a cross-linking monomer and an initiator in an aqueous solution of an emulsifier to obtain a polymer microsphere with epoxy groups;

[0012] (2A) Reacting the polymer microsphere with epoxy groups obtained in step (1A) with an amino compound to obtain a polymer microsphere with amino groups;

[0013] (3A) Depositing magnetic nanoparticles on the surface of the polymer microsphere with amino groups obtained in step (2A) to obtain the magnetic polymer microsphere.

[0014] Preferably, the particle size of the polymer microsphere in step (1A) is 0.2 - 3 μm, such as 0.5 μm, 1 μm, 1.5 μm, 2 μm or 2.5 μm, etc.

[0015] Preferably, the polymer microspheres in step (1A) include polystyrene microspheres or polyacrylate microspheres.

[0016] Preferably, the polymer microspheres in step (1A) are prepared by any one of emulsion polymerization, microemulsion polymerization, soap-free emulsion polymerization or dispersion polymerization.

[0017] Preferably, the polymer microspheres in step (1A) are obtained by reacting a monomer and an initiator in a solvent.

[0018] Preferably, the monomer includes any one or a combination of at least two of styrene, methyl acrylate, methyl methacrylate, glycidyl methacrylate, 2-hydroxyethyl methacrylate or vinyl acetate.

[0019] Preferably, based on 1 g of the mass of the monomer, the mass of the initiator is 0.01 - 0.5 g, such as 0.05 g, 0.1 g, 0.2 g, 0.3 g or 0.4 g, etc.

[0020] Preferably, the initiator includes any one or a combination of at least two of potassium persulfate, ammonium persulfate, sodium persulfate, azobisisobutyronitrile, azodiisovaleronitrile, dimethyl 2,2'-azobis(2-methylpropionate), cumene hydroperoxide, dicumyl peroxide, benzoyl peroxide or tert-butyl peroxybenzoate.

[0021] Preferably, based on 1 g of the mass of the monomer, the mass of the solvent is 1 - 50 g (excluding the solvent used to dissolve the emulsifier and stabilizer), such as 5 g, 10 g, 20 g, 30 g or 40 g, etc.

[0022] Preferably, the solvent includes any one or a combination of at least two of water, methanol, ethanol, ethyl acetate, isopropanol or tetrahydrofuran.

[0023] Preferably, an emulsifier and / or a stabilizer are added during the reaction process, and both are previously dissolved in a small amount of solvent.

[0024] Preferably, based on 100% of the total mass of the solvent, the mass of the emulsifier is not higher than 5%, and the mass of the stabilizer is also not higher than 5%.

[0025] Preferably, the emulsifier includes any one or a combination of at least two of sodium naphthenate, sodium oleate, sodium dodecyl sulfate, sodium benzenesulfonate, sodium isopropylsulfonate, potassium methylene bis(isopropylnaphthalenesulfonate), diacetyl tartaric acid esters of mono- and diglycerides, polyoxyethylene sorbitan fatty acid esters, octylphenol polyoxyethylene ether, dodecylphenol ethers or dinonylphenol ethers.

[0026] Preferably, the stabilizer includes any one or a combination of at least two of polyvinylpyrrolidone, polyethylene glycol, polyvinyl alcohol, methylcellulose, ethylcellulose, or polyacrylic acid.

[0027] Preferably, the temperature of the reaction is 50 - 80 °C, such as 55 °C, 60 °C, 65 °C, 70 °C, or 75 °C, etc.

[0028] Preferably, the reaction time is 16 - 30 h, such as 20 h, 23 h, 26 h, or 29 h, etc.

[0029] Preferably, after the reaction, it further includes the step of washing multiple times with one or more solvents such as water, methanol, ethanol, or ethyl acetate.

[0030] Preferably, based on the mass of the polymer microspheres in step (1A) being 1 g, the total mass of the epoxy monomer, functional monomer, and crosslinking monomer is 5 - 500 g, such as 10 g, 50 g, 100 g, 200 g, 300 g, or 400 g, etc.

[0031] Preferably, the mass ratio of the epoxy monomer, functional monomer, and crosslinking monomer in step (1A) is (10 - 30):(50 - 85):(5 - 20), such as 10:85:5, 20:70:10, or 30:50:20, etc.

[0032] Preferably, the epoxy monomer in step (1A) includes any one or a combination of at least two of glycidyl methacrylate, n-butyl glycidyl ether, resorcinol diglycidyl ether, ethylene glycol diglycidyl ether, tert-butyl glycidyl ether, or allyl glycidyl ether.

[0033] Preferably, the functional monomer in step (1A) includes any one or a combination of at least two of methyl acrylate, methyl methacrylate, ethyl acrylate, ethyl methacrylate, n-butyl acrylate, n-butyl methacrylate, 2-hydroxyethyl methacrylate, isobutyl acrylate, or methacrylic acid.

[0034] Preferably, the crosslinking monomer in step (1A) includes any one or a combination of at least two of butanediol dimethacrylate, ethylene glycol dimethacrylate, glycerol dimethacrylate, trimethylolpropane trimethacrylate, divinylbenzene, butadiene, or N,N'-methylenebisacrylamide.

[0035] Preferably, based on the total mass of the epoxy monomer, functional monomer, and crosslinking monomer in step (1A) being 100 g, the mass of the initiator is 1 - 10 g, such as 2 g, 4 g, 6 g, or 8 g, etc.

[0036] Preferably, the initiator in step (1A) includes any one or a combination of at least two of potassium persulfate, ammonium persulfate, sodium persulfate, azobisisobutyronitrile, azodiisopentanenitrile, dimethyl azodicarboxylate, cumene hydroperoxide, dicumyl peroxide, benzoyl peroxide, or tert-butyl perbenzoate.

[0037] Preferably, the mass percentage content of the emulsifier in the aqueous solution of the emulsifier in step (1A) is 0.01 - 5%, such as 0.1%, 0.5%, 1%, 1.5%, 3%, or 3.5%, etc.

[0038] Preferably, the emulsifier in step (1A) includes any one or a combination of at least two of sodium naphthenate, sodium oleate, sodium dodecyl sulfate, sodium benzenesulfonate, sodium isopropylsulfonate, potassium methylene bis(isopropylnaphthalenesulfonate), diacetyl tartaric acid esters of mono- and diglycerides, polyoxyethylene sorbitan fatty acid esters, octylphenol polyoxyethylene ether, dodecylphenol ether, or dinonylphenol ether.

[0039] Preferably, a stabilizer is further added during the reaction in step (1A). The stabilizer can be dissolved in a small amount of solvent in advance, and then dispersed by machines such as a high-speed shear emulsifier, a high-pressure homogenizing disperser, an ultrasonic cell disruptor, etc. to produce droplets and then added to carry out the reaction.

[0040] Preferably, based on the mass of the aqueous solution of the emulsifier in (1A) being 100%, the mass of the stabilizer is 0.5 - 5%.

[0041] Preferably, the stabilizer includes any one or a combination of at least two of polyvinylpyrrolidone, polyethylene glycol, polyvinyl alcohol, methylcellulose, ethylcellulose, or polyacrylic acid.

[0042] Preferably, the reaction temperature in step (1A) is 30 - 80 °C, such as 35 °C, 40 °C, 50 °C, 60 °C, or 70 °C, etc.

[0043] Preferably, the reaction time in step (1A) is 8 - 40 h, such as 10 h, 15 h, 20 h, 25 h, 30 h, or 35 h, etc.

[0044] Preferably, step (1A) specifically includes: mixing polymer microspheres, epoxy monomers, functional monomers, crosslinking monomers, and an initiator in an aqueous solution of an emulsifier at 30 - 50 °C for 8 - 20 h, then adding a stabilizer solution, dispersing using machines such as a high-speed shear emulsifier, a high-pressure homogenizing disperser, an ultrasonic cell disruptor, etc., adding to a reactor, and reacting at 40 - 80 °C for 8 - 20 h to obtain the polymer microspheres with epoxy groups.

[0045] Preferably, after the reaction in step (1A), it further includes a step of washing with one or several of anhydrous ethanol, methanol, isopropanol, tetrahydrofuran, ethyl acetate or water for multiple times.

[0046] Preferably, based on 1 g of the polymer microspheres with epoxy groups in step (2A), the dosage of the amino compound is 10 - 100 mol, such as 20 mol, 40 mol, 60 mol or 80 mol, etc.

[0047] Preferably, the amino compound in step (2A) includes any one or a combination of at least two of diisopropylamine, ethanolamine, triethanolamine, ethylenediamine, ammonia water, tetraethylenepentamine or polyethyleneimine;

[0048] Preferably, the temperature of the reaction in step (2A) is 30 - 80 °C, such as 40 °C, 50 °C, 60 °C or 70 °C, etc.

[0049] Preferably, the reaction time in step (2A) is 8 - 20 h, such as 10 h, 12 h, 14 h, 16 h or 18 h, etc.

[0050] Preferably, after the reaction in step (2A), it first undergoes multiple washings with one or several of methanol, ethanol, n - hexane, isopropanol, tetrahydrofuran, etc., and then multiple washings with water.

[0051] Preferably, the method for surface - depositing magnetic nanoparticles in step (3A) specifically includes: reacting the polymer microspheres with amino groups obtained in step (2A), ferric salts, ferrous salts and surfactants in a solvent to complete the deposition of magnetic nanoparticles.

[0052] Preferably, based on 1 g of the polymer microspheres with amino groups, the dosage of the ferric salt is 1 - 20 mmol, such as 2 mmol, 4 mmol, 6 mmol, 8 mmol, 10 mmol, 12 mmol, 14 mmol, 16 mmol or 18 mmol, etc.

[0053] Preferably, based on 1 g of the polymer microspheres with amino groups, the dosage of the ferrous salt is 1 - 15 mmol, such as 2 mmol, 4 mmol, 6 mmol, 8 mmol, 10 mmol, 12 mmol or 14 mmol, etc.

[0054] Preferably, the ferric salt includes any one or a combination of at least two of ferric chloride, ferric sulfate or ferric nitrate.

[0055] Preferably, the ferrous salt includes any one or a combination of at least two of ferrous chloride, ferrous sulfate, ferrous ammonium sulfate or ferrous nitrate.

[0056] In the present invention, both the ferric salt and the ferrous salt can be dissolved in a small amount of solvent in advance.

[0057] Preferably, the surfactant includes any one or a combination of at least two of undecylenic acid, methyl undecylenate, butyl undecylenate, oleic acid, methyl palmitoleate, sodium oleate, dodecylamine, octadecylamine, sodium laurate, coconut amine, sodium stearate, sodium dodecyl sulfate or sodium dodecylbenzenesulfonate.

[0058] Preferably, based on the mass of the solvent used in this step being 100%, the mass of the surfactant is 0.1 - 5%, such as 0.5%, 1%, 2%, 3% or 4%, etc.

[0059] Preferably, the reaction is carried out under alkaline conditions, and the pH value of the reaction can be adjusted to be alkaline by adding alkaline substances such as ammonia water, sodium hydroxide, potassium hydroxide, sodium bicarbonate, sodium carbonate or potassium carbonate.

[0060] Preferably, the temperature of the reaction is 50 - 90 °C, such as 60 °C, 70 °C or 80 °C, etc.

[0061] Preferably, the reaction time is 0.5 - 5 h, such as 1 h, 2 h, 3 h or 4 h, etc.

[0062] Preferably, after the reaction, it further includes the step of washing several times with one or several of methanol, ethanol, isopropanol, tetrahydrofuran, etc., and then washing several times with water.

[0063] Preferably, the preparation method specifically includes the following steps:

[0064] (1) Reacting magnetic polymer microspheres, monomers containing benzene rings, crosslinking monomers and initiators in an aqueous solution of an emulsifier to obtain magnetic polymer microspheres with benzene rings;

[0065] (2) Reacting the magnetic polymer microspheres with benzene rings obtained in step (1) and chloromethyl compounds in a solvent to obtain magnetic polymer microspheres with chloromethyl groups;

[0066] (3) Reacting the magnetic polymer microspheres with chloromethyl groups obtained in step (2) and hydroxyl compounds to obtain magnetic polymer microspheres with hydroxyl groups;

[0067] (4) Reacting the magnetic polymer microspheres with hydroxyl groups obtained in step (3) and p-toluenesulfonyl chloride to obtain the magnetic polymer microspheres containing p-toluenesulfonyl groups.

[0068] Preferably, based on 1 g of the mass of the magnetic polymer microspheres described in step (1), the total mass of the benzene ring-containing monomer and the crosslinking monomer is 0.2 to 75 g, such as 0.5 g, 1 g, 10 g, 20 g, 30 g, 40 g, 50 g, or 60 g, etc.

[0069] Preferably, the mass ratio of the benzene ring-containing monomer to the crosslinking monomer described in step (1) is (50 to 80):(20 to 50), such as 60:40, 70:30, or 75:25, etc.

[0070] Preferably, the benzene ring-containing monomer described in step (1) includes any one or a combination of at least two of styrene, methylstyrene, or 4-tert-butylstyrene.

[0071] Preferably, the crosslinking monomer described in step (1) includes any one or a combination of at least two of butanediol dimethacrylate, ethylene glycol dimethacrylate, glycerol dimethacrylate, trimethylolpropane trimethacrylate, divinylbenzene, butadiene, or N,N'-methylenebisacrylamide.

[0072] Preferably, based on 100% of the total mass of the benzene ring-containing monomer and the crosslinking monomer described in step (1), the mass of the initiator is 1 to 10%, such as 2%, 4%, 6%, or 8%, etc.

[0073] Preferably, the initiator described in step (1) includes any one or a combination of at least two of potassium persulfate, ammonium persulfate, sodium persulfate, azobisisobutyronitrile, azobisisoheptonitrile, dimethyl azobisisobutyrate, cumene hydroperoxide, dicumyl peroxide, benzoyl peroxide, or tert-butyl peroxybenzoate.

[0074] Preferably, the mass percentage content of the emulsifier in the aqueous solution of the emulsifier described in step (1) is 0.01 to 5%, such as 0.05%, 0.1%, 0.5%, 1%, 2%, 3%, or 4%, etc.

[0075] Preferably, the emulsifier includes any one or a combination of at least two of sodium alkanoate, sodium oleate, sodium dodecyl sulfate, sodium benzenesulfonate, potassium methylene diisopropylnaphthalene sulfonate, diacetyl tartaric acid esters of mono- and diglycerides, polyoxyethylene sorbitan fatty acid esters, octylphenol polyoxyethylene ether, dodecylphenol ether, or dinonylphenol ether.

[0076] Preferably, the temperature of the reaction described in step (1) is 50 to 80 °C, such as 55 °C, 60 °C, 65 °C, 70 °C, or 75 °C, etc.

[0077] Preferably, the reaction time described in step (1) is 3 to 20 h, such as 5 h, 7 h, 9 h, 11 h, 13 h, 15 h, 17 h, or 19 h, etc.

[0078] Preferably, after the reaction in step (1), there is also a step of washing by centrifugation or magnetic separation one or more times with one or more of methanol, ethanol, isopropanol, tetrahydrofuran, ethyl acetate or water.

[0079] Preferably, based on 1 g of the amount of the magnetic polymer microspheres with a benzene ring in step (2), the amount of the chloromethyl compound is 10 - 100 mmol, such as 20 mmol, 40 mmol, 60 mmol or 80 mmol, etc.

[0080] Preferably, the chloromethyl compound in step (2) includes any one or a combination of at least two of dichloromethyl methyl ether, 2 - chloroethyl methyl ether, dichloroisopropyl ether, paraformaldehyde, dimethoxymethane or chloromethyl ethyl ether.

[0081] Preferably, the solvent in step (2) includes any one or a combination of at least two of 1,4 - dioxane, dichloromethane, chloroform or dimethyl sulfoxide.

[0082] Preferably, a catalyst is also added in the reaction in step (2).

[0083] Preferably, based on 1 g of the amount of the magnetic polymer microspheres with a benzene ring in step (2), the amount of the catalyst is 5 - 50 mmol, such as 10 mmol, 20 mmol, 30 mmol or 40 mmol, etc.

[0084] Preferably, the catalyst includes any one or a combination of at least two of tin tetrachloride, aluminum trichloride, boron trifluoride or zinc chloride;

[0085] Preferably, the temperature of the reaction in step (2) is 20 - 40 °C, such as 25 °C, 30 °C or 35 °C, etc.

[0086] Preferably, the reaction time in step (2) is 16 - 24 h, such as 18 h, 20 h or 22 h, etc.

[0087] Preferably, after the reaction in step (2), there is also a step of washing the magnetic polymer microspheres one or more times with one or more of 1,4 - dioxane, methanol, ethanol, tetrahydrofuran, water, etc.

[0088] Preferably, based on 1 g of the mass of the magnetic polymer microspheres with chloromethyl in step (3), the mass of the hydroxy compound is 0.1 - 100 g, such as 0.5 g, 1 g, 5 g, 10 g or 50 g, etc.

[0089] Preferably, the hydroxy compound in step (3) includes any one or a combination of at least two of n-butanol, isobutanol, n-propanol, ethylene glycol, polyethylene glycol 200, polyethylene glycol 400, polyethylene glycol 600, or polyethylene glycol 800.

[0090] Preferably, the solvent in step (3) includes any one or a combination of at least two of ethanol, n-hexane, dichloromethane, chloroform, acetonitrile, or tetrahydrofuran.

[0091] Preferably, the reaction in step (3) is carried out under alkaline conditions, and the alkaline conditions can be achieved by adding an inorganic base and / or an organic base.

[0092] Preferably, the inorganic base includes but is not limited to: sodium bicarbonate, ammonium bicarbonate, sodium carbonate, potassium carbonate, cesium carbonate, sodium hydroxide, or potassium hydroxide, and the organic base includes but is not limited to: ethanolamine, triethanolamine, triethylamine, ethylenediamine, pyridine, diisopropylamine, sodium methoxide, or tert-butyllithium.

[0093] Preferably, the temperature of the reaction in step (3) is 50 - 80 °C, such as 55 °C, 60 °C, 65 °C, 70 °C, or 75 °C, etc.

[0094] Preferably, the reaction time in step (3) is 8 - 20 h, such as 10 h, 12 h, 14 h, 16 h, or 18 h, etc.

[0095] Preferably, after the reaction in step (3), it further includes a step of magnetically attracting and washing several times with one or several of anhydrous ethanol, methanol, isopropanol, tetrahydrofuran, ethyl acetate, water, etc.

[0096] Preferably, based on the mass of 1 g of the magnetic polymer microspheres with hydroxyl groups in step (4), the mass of p-toluenesulfonyl chloride is 0.1 - 10 g, such as 2 g, 4 g, 6 g, or 8 g, etc.

[0097] Preferably, the solvent in step (4) includes any one or a combination of at least two of tetrahydrofuran, 1,4-dioxane, dimethyl sulfoxide, N,N-dimethylformamide, ether, or acetonitrile.

[0098] Preferably, an organic base is further added to the reaction in step (4), and the organic base includes but is not limited to: triethylamine, ethylenediamine, 2-phenylpyridine, pyridine, triethylenediamine, N-methylmorpholine, or tetramethylethylenediamine.

[0099] Preferably, based on the mass of 1 g of the magnetic polymer microspheres with hydroxyl groups in step (4), the mass of the organic base is 0.1 - 10 g, such as 2 g, 4 g, 6 g, or 8 g, etc.

[0100] Preferably, the temperature of the reaction in step (4) is 5 to 50 °C, such as 10 °C, 15 °C, 20 °C, 25 °C, 30 °C, 35 °C, 40 °C or 45 °C, etc.

[0101] Preferably, the reaction time in step (4) is 2 to 30 h, such as 5 h, 10 h, 15 h, 20 h or 25 h, etc.

[0102] Preferably, after the reaction in step (4), it further includes magnetic suction washing with one or several of anhydrous ethanol, methanol, isopropanol, tetrahydrofuran, ethyl acetate, water, etc. multiple times, and then adding an appropriate amount of water for dispersion and storing in a refrigerator at 2 - 8 °C.

[0103] Preferably, the reactions in steps (1) - (4) are all carried out under the protection of a protective gas, such as under the protection of nitrogen.

[0104] In a second aspect, the present invention provides a magnetic polymer microsphere containing a p - toluenesulfonyl group, and the magnetic polymer microsphere is prepared by the preparation method as described in the first aspect.

[0105] Preferably, the particle size of the magnetic polymer microsphere is 3 - 10 μm, such as 4 μm, 5 μm, 6 μm, 7 μm, 8 μm or 9 μm, etc.

[0106] In a third aspect, the present invention provides an application of the magnetic polymer microsphere as described in the second aspect in targeted capture or separation and purification.

[0107] Compared with the prior art, the present invention has the following beneficial effects:

[0108] The preparation method of the magnetic polymer microsphere containing a p - toluenesulfonyl group provided by the present invention first coats the surface of the magnetic polymer microsphere with a benzene - ring - containing polymer, and then sequentially carries out chloromethyl modification, hydroxyl modification and p - toluenesulfonyl chloride modification on the magnetic polymer microsphere coated with the benzene - ring polymer. This preparation method has the advantage of high group utilization rate, and the obtained magnetic polymer microsphere containing a p - toluenesulfonyl group has the advantages of high detection accuracy and low physical adsorption of non - target substances, and can be applied in many aspects such as targeted capture, separation and purification. Description of the Drawings

[0109] Figure 1 It is a scanning electron microscope image of the magnetic polymer microsphere containing a p - toluenesulfonyl group provided in Example 1. Detailed Embodiments

[0110] The technical solutions of the present invention will be further described below through specific embodiments. Those skilled in the art should understand that the embodiments are only for helping to understand the present invention and should not be regarded as specific limitations to the present invention.

[0111] Preparation Example 1

[0112] A magnetic polymer microsphere, and its preparation method includes the following steps:

[0113] (1) Take 1.8 g of styrene and add it to a 100 mL three-necked flask, then add 10 mL of purified water; weigh 0.03 g of sodium persulfate, dissolve it in 10 mL of purified water under ultrasonic conditions and add it to the three-necked flask. Under a nitrogen environment, react at 60 °C for 24 h. After the reaction is completed, the obtained polymer microspheres have a particle size of about 0.75 μm. After centrifugally washing 3 times with purified water, add an appropriate amount of purified water for dispersion and store in a refrigerator at 5 °C;

[0114] (2) Take 1 g of the polymer microspheres obtained in step (1) and transfer them to a 500 mL four-necked flask; add 15 g of glycidyl methacrylate, 42 g of methyl acrylate, 3 g of divinylbenzene and 1.5 g of azobisisobutyronitrile to a 500 mL beaker. After ultrasonic dissolution of the solids, add 200 mL of an aqueous solution of sodium oleate with a mass percentage of 0.15%. Disperse it using an ultrasonic cell disruptor, then add it to the four-necked flask and react at 40 °C for 16 h; afterwards, weigh 2 g of methyl cellulose into a 250 mL beaker, add 150 mL of an aqueous solution of sodium oleate with a mass percentage of 0.15%, dissolve it ultrasonically and add it to the above 500 mL four-necked flask. React at 60 °C for 20 h under a nitrogen environment. After the reaction is completed, centrifugally wash 3 times with absolute ethanol and then 3 times with purified water, add an appropriate amount of purified water for dispersion to obtain polymer microspheres with epoxy groups;

[0115] (3) Take 2 g of the polymer microspheres with epoxy groups obtained in step (2), transfer them to a 100 mL three-necked flask with 50 mL of purified water. Add 1 g of ethylenediamine to the three-necked flask and react at 60 °C for 10 h under a nitrogen environment. After the reaction is completed, centrifugally wash 5 times with purified water to obtain polymer microspheres with amino groups;

[0116] (4) Take 2 g of the polymer microspheres with amino groups obtained in step (3) and transfer them to a 500 mL three-necked flask. Weigh 1.42 g of ferric chloride hexahydrate into a 100 mL beaker, add 40 mL of purified water, dissolve it ultrasonically and add it to the three-necked flask. Then weigh 0.84 g of ferrous sulfate heptahydrate into a 100 mL beaker, add 40 mL of purified water, dissolve it ultrasonically and add it to the three-necked flask; pass nitrogen into the system and set the reaction temperature to 70 °C. After reaching the preset temperature and stabilizing for 10 min, add 0.2 g of sodium oleate dissolved in 15 mL of purified water to the three-necked flask. Add 10 mL of ammonia water with a mass concentration of 25% to the three-necked flask in a dropwise manner, and the dropping time is 1 h. After the dropping is completed, continue to react at 80 °C for 1 h. After the reaction is completed, centrifugally wash 5 times with purified water to obtain the magnetic polymer microspheres.

[0117] Preparation Example 2

[0118] A magnetic polymer microsphere, the preparation method thereof comprising the following steps:

[0119] (1) Take 2 g of methyl methacrylate and add it to a 100 mL three-necked flask, then add 20 mL of ethanol; weigh 0.1 g of azobisisobutyronitrile and add it to the three-necked flask, then weigh 0.45 g of methylcellulose, dissolve it in 20 mL of ethanol and add it to the three-necked flask. Under a nitrogen atmosphere, react at 50 °C for 16 h. After the reaction is completed, the obtained polymer microspheres have a particle size of about 1.1 μm, and are centrifugally washed 3 times with anhydrous ethanol and 3 times with purified water, and then dispersed in an appropriate amount of purified water and stored in a refrigerator at 5 °C;

[0120] (2) Take 0.5 g of the polymer microspheres obtained in step (1) and transfer them to a 500 mL four-necked flask; add 2.5 g of butyl glycidyl ether, 7 g of methacrylic acid, 0.5 g of glycerol dimethacrylate and 0.1 g of azobisisobutyronitrile to a 250 mL beaker. After ultrasonic dissolution of the solid, add 200 mL of an aqueous solution of polyoxyethylene sorbitan fatty acid ester with a mass percentage of 0.5%, disperse it using an ultrasonic cell disruptor, and then add it to the four-necked flask and react at 50 °C for 16 h; then, weigh 2.5 g of polyacrylic acid into a 250 mL beaker, add 100 mL of an aqueous solution of polyoxyethylene sorbitan fatty acid ester with a mass percentage of 0.5%, ultrasonically dissolve it and add it to the above 500 mL four-necked flask, and react at 50 °C for 20 h under a nitrogen atmosphere. After the reaction is completed, centrifuge and wash 3 times with anhydrous ethanol and then 3 times with purified water, and disperse in an appropriate amount of purified water to obtain polymer microspheres with epoxy groups;

[0121] (3) Take 2 g of the polymer microspheres with epoxy groups obtained in step (2), transfer them to a 100 mL three-necked flask with 10 mL of purified water, add 1.5 g of ethanolamine to the three-necked flask, and react at 50 °C for 20 h under a nitrogen atmosphere. After the reaction is completed, centrifuge and wash 5 times with purified water to obtain polymer microspheres with amino groups;

[0122] (4) Take 2 g of the polymer microspheres with amino groups obtained in step (3), transfer them to a 500 mL three-necked flask. Weigh 1.6 g of ferric chloride hexahydrate into a 100 mL beaker, add 40 mL of purified water, ultrasonically dissolve it and then add it to the three-necked flask. Then weigh 1.32 g of ferrous sulfate heptahydrate into a 100 mL beaker, add 40 mL of purified water, ultrasonically dissolve it and then add it to the three-necked flask. Weigh 0.25 g of sodium stearate into a 50 mL beaker, add 15 mL of purified water, ultrasonically disperse it and then add it to the three-necked flask. Then add 100 mL of a sodium bicarbonate solution with a concentration of 30 mmol / L to the three-necked flask. The system is purged with nitrogen and reacted at 80 °C for 1 hour. After the reaction is completed, centrifuge and wash 5 times with purified water to obtain the magnetic polymer microspheres.

[0123] Preparation Example 3

[0124] A magnetic polymer microsphere, and its preparation method includes the following steps:

[0125] (1) Take 2 g of 2-hydroxyethyl methacrylate and add it to a 100 mL three-necked flask, and then add 10 mL of ethyl acetate. Weigh 0.03 g of azobisisobutyronitrile, ultrasonically dissolve it in 10 mL of ethyl acetate and add it to the three-necked flask. Under a nitrogen atmosphere, react at 80 °C for 20 h. After the reaction is completed, the obtained polymer microspheres, with a particle size of about 0.8 μm, are centrifuged and washed 3 times with ethyl acetate, 3 times with absolute ethanol, and 3 times with purified water by centrifugation, and then dispersed in an appropriate amount of water and stored in a refrigerator at 5 °C;

[0126] (2) Take 0.5 g of the polymer microspheres obtained in step (1) and transfer them to a 500 mL four-necked flask. Add 12.5 g of allyl glycidyl ether, 35 g of isobutyl acrylate, 2.5 g of divinylbenzene and 0.1 g of azobisisobutyronitrile to a 250 mL beaker, ultrasonically dissolve the solids and then add 100 mL of an aqueous solution of sodium oleate with a mass percentage of 0.1%. After pulverizing for 15 min using an ultrasonic cell disruptor, add it to the four-necked flask and react at 50 °C for 8 h. Then, weigh 2 g of ethyl cellulose into a 250 mL beaker, add 100 mL of purified water, ultrasonically dissolve it and add it to the above-mentioned 500 mL four-necked flask. React at 70 °C for 16 h under a nitrogen atmosphere. After the reaction is completed, centrifuge and wash 3 times with absolute ethanol and then 3 times with purified water by centrifugation, and disperse in an appropriate amount of purified water to obtain polymer microspheres with epoxy groups;

[0127] (3) Take 2 g of the polymer microspheres with epoxy groups obtained in step (2), transfer them to a 100 mL three-necked flask with 20 mL of purified water. Add 4.2 g of triethanolamine to the three-necked flask and react at 40 °C for 20 h under a nitrogen atmosphere. After the reaction is completed, centrifuge and wash 5 times with purified water to obtain polymer microspheres with amino groups;

[0128] (4) Take 2 g of the polymer microspheres with amino groups obtained in step (3), transfer them to a 500 mL three-necked flask. Weigh 2 g of ferric chloride hexahydrate into a 100 mL beaker, add 80 mL of deoxygenated water, ultrasonically dissolve it and then add it to the three-necked flask. Then weigh 1.07 g of ferrous chloride tetrahydrate into a 100 mL beaker, add 80 mL of deoxygenated water, ultrasonically dissolve it and add it to the three-necked flask. Weigh 0.6 g of sodium dodecyl sulfate, ultrasonically dissolve it in 20 mL of deoxygenated water and add it to the three-necked flask. Then add 20 mL of ammonia water with a mass concentration of 25% to the three-necked flask. The system is purged with nitrogen and the reaction temperature is set at 60 °C. React for 1 hour. After the reaction is completed, centrifuge and wash 5 times with purified water to obtain the magnetic polymer microspheres.

[0129] Preparation Example 4

[0130] A magnetic polymer microsphere, and its preparation method comprises the following steps:

[0131] (1) Take 2 g of glycidyl methacrylate and add it to a 100 mL three-necked flask, and then add 10 mL of purified water. Weigh 0.1 g of sodium persulfate, ultrasonically dissolve it in 10 mL of purified water and add it to the three-necked flask. Then weigh 0.06 g of sodium oleate, dissolve it in 20 mL of purified water and add it to the three-necked flask. Under a nitrogen atmosphere, react at 50 °C for 20 h. After the reaction is completed, the obtained polymer microspheres, with a particle size of about 1.5 μm, are centrifuged and washed 3 times with purified water, then dispersed in an appropriate amount of water and stored in a refrigerator at 5 °C;

[0132] (2) Take 1 g of the polymer microspheres obtained in step (1) and transfer them to a 500 mL four-necked flask. Add 25 g of butyl glycidyl ether, 70 g of methyl methacrylate, 5 g of divinylbenzene and 2 g of azobisisobutyronitrile to a 250 mL beaker. Ultrasonically dissolve the solids and then add 100 mL of an aqueous solution of diacetyl tartaric acid ester of mono- and diglycerides with a mass percentage content of 0.25%. After pulverizing with an ultrasonic cell disruptor for 15 min, add it to the four-necked flask and react at 40 °C for 16 h. Then, weigh 4 g of methyl cellulose into a 250 mL beaker, add 100 mL of purified water, ultrasonically dissolve it and add it to the above 500 mL four-necked flask. React at 70 °C for 16 h under a nitrogen atmosphere. After the reaction is completed, centrifuge and wash 3 times with absolute ethanol and then 3 times with purified water, and disperse in an appropriate amount of purified water to obtain polymer microspheres with epoxy groups;

[0133] (3) Take 2 g of the polymer microspheres with epoxy groups obtained in step (2), transfer them to a 100 mL three-necked flask with 20 mL of purified water. Add 2.5 g of diisopropylamine to the three-necked flask and react at 60 °C for 10 h under a nitrogen atmosphere. After the reaction is completed, centrifuge and wash 5 times with purified water to obtain polymer microspheres with amino groups;

[0134] (4) Take 2 g of the amino-group-containing polymer microspheres obtained in step (3), transfer them to a 1000 mL three-necked flask. Weigh 7.6 g of ferric sulfate into a 250 mL beaker, add 100 mL of deoxygenated water, ultrasonically dissolve it and then add it to the three-necked flask. Then weigh 2.6 g of ferrous chloride tetrahydrate into a 250 mL beaker, add 100 mL of deoxygenated water, ultrasonically dissolve it and add it to the three-necked flask. Weigh 0.5 g of sodium dodecylbenzenesulfonate, ultrasonically dissolve it in 20 mL of deoxygenated water and add it to the three-necked flask. Then add 100 mL of a 100 mmol / L sodium hydroxide solution to the three-necked flask. The system is purged with nitrogen and the reaction temperature is set at 70 °C. React for 3 h. After the reaction is completed, centrifuge and wash 5 times with purified water to obtain the magnetic polymer microspheres.

[0135] Example 1

[0136] A preparation method of a magnetic polymer microsphere containing a p-toluenesulfonyl group specifically comprises the following steps:

[0137] (1) Take 2 g of the magnetic polymer microspheres provided in Preparation Example 1, transfer them to a 250 mL three-necked flask with 40 mL of a sodium naphthenate aqueous solution with a mass percentage of 0.5%. Take 4.8 g of styrene and 3.2 g of divinylbenzene into a 100 mL beaker, add 0.8 g of azobisisobutyronitrile, and then add 40 mL of a sodium naphthenate aqueous solution with a mass percentage of 0.5% thereto. Disperse it with an ultrasonic cell disruptor. Under a nitrogen atmosphere, set the reaction temperature at 60 °C. After reaching the preset temperature, stabilize for 10 minutes and add the above solvent mixed with the initiator and monomer dropwise to the three-necked flask, controlling the dropping time to be 2 h. After the dropping is completed, continue to react for 16 h. After the reaction is completed, magnetically wash 3 times with purified water to obtain the magnetic polymer microspheres with benzene rings.

[0138] (2) Take 2 g of the magnetic polymer microspheres with benzene rings obtained in step (1), displace them 3 times with 40 mL of absolute ethanol and 40 mL of dichloromethane respectively. After magnetically removing the supernatant, transfer them to a 100 mL three-necked flask with 60 mL of dichloromethane. Add 3.4 g of chloromethyl ethyl ether, and slowly add 4.6 g of tin tetrachloride within 40 min during stirring in an ice-water bath at 5 °C. After the addition of tin tetrachloride is completed, continue to react at 25 °C for 16 h. After the reaction is completed, wash 3 times with tetrahydrofuran, 3 times with absolute ethanol, and 3 times with purified water to obtain the magnetic polymer microspheres with chloromethyl groups.

[0139] (3) Take 2 g of the magnetic polymer microspheres with chloromethyl groups obtained in step (2), displace them with 40 mL of anhydrous ethanol three times and 40 mL of tetrahydrofuran three times in sequence. After magnetic separation, transfer the microspheres to a reaction vessel with 40 mL of tetrahydrofuran. Add 0.05 g of ethylenediamine to the reaction vessel. Measure 4 mL of ethylene glycol, dissolve it in 20 mL of tetrahydrofuran, and add it to the reaction vessel drop by drop. Control the dropping time to be 2 h. Under a nitrogen atmosphere, react at 60 °C for 16 h. After the reaction is completed, wash with anhydrous ethanol three times and purified water three times to obtain magnetic polymer microspheres with hydroxyl groups.

[0140] (4) Take 2 g of the magnetic polymer microspheres with hydroxyl groups obtained in step (3), displace them with 40 mL of anhydrous ethanol and 40 mL of tetrahydrofuran three times. After magnetic separation, transfer them to a reaction vessel with 40 mL of tetrahydrofuran. Add 2 g of p-toluenesulfonyl chloride and 4 mL of tetramethylethylenediamine to the reaction vessel, and react at 40 °C for 5 h. After the reaction is completed, wash with anhydrous ethanol three times, and wash with purified water multiple times until the pH value of the supernatant is close to neutral. After magnetic separation to remove the supernatant, add an appropriate amount of purified water to disperse to obtain magnetic polymer microspheres containing p-toluenesulfonyl groups, with a particle size of about 3.2 μm, and store them in a refrigerator at 2 - 8 °C.

[0141] Example 2

[0142] A preparation method of magnetic polymer microspheres containing p-toluenesulfonyl groups specifically includes the following steps:

[0143] (1) Take 2 g of the magnetic polymer microspheres provided in Preparation Example 2, transfer them to a 100 mL three-necked flask with 20 mL of an aqueous solution of polyoxyethylene sorbitan fatty acid ester with a mass percentage of 0.5%. Add 0.2 g of azobisisobutyronitrile, and then add 40 mL of an aqueous solution of polyoxyethylene sorbitan fatty acid ester with a mass percentage of 0.5%. Disperse them using an ultrasonic cell disruptor. Under a nitrogen atmosphere, set the reaction temperature to 65 °C. After reaching the preset temperature, stabilize for 10 min and add the above solvent mixed with the initiator and monomer to the three-necked flask drop by drop. Control the dropping time to be 2 h. After the dropping is completed, react at 50 °C for 10 h. After the reaction is completed, wash with purified water by magnetic separation three times to obtain magnetic polymer microspheres with benzene rings.

[0144] (2) Take 2 g of the magnetic polymer microspheres with a benzene ring obtained in step (1), and replace them 3 times with 40 mL of anhydrous ethanol, 40 mL of tetrahydrofuran, and 40 mL of 1,4-dioxane respectively. After removing the supernatant by magnetic attraction, transfer them to a 250 mL three-necked flask with 60 mL of 1,4-dioxane. Add 4.48 g of dichloroethyl ether, and slowly add 4.6 g of tin tetrachloride within 40 min during stirring in an ice-water bath at 5°C. After adding the tin tetrachloride, continue the reaction at 25°C for 16 h. After the reaction is completed, wash it 3 times with tetrahydrofuran, 3 times with anhydrous ethanol, and 3 times with purified water to obtain magnetic polymer microspheres with chloromethyl groups;

[0145] (3) Take 2 g of the magnetic polymer microspheres with chloromethyl groups obtained in step (2), and replace them 3 times with 40 mL of anhydrous ethanol and 3 times with 40 mL of n-hexane respectively. After magnetic attraction, transfer the magnetic polymer microspheres to a reaction vessel with 40 mL of n-hexane. Take 0.1 g of ethanolamine and add it to the reaction vessel; measure 4 mL of ethylene glycol, dissolve it in 20 mL of n-hexane, and add it to the reaction vessel dropwise. Control the dropping time within 2 h, and react at 60°C for 20 h in a nitrogen environment. After the reaction is completed, wash it 3 times with anhydrous ethanol and 3 times with purified water to obtain magnetic polymer microspheres with hydroxyl groups;

[0146] (4) Take 2 g of the magnetic polymer microspheres with hydroxyl groups obtained in step (3), and replace them 3 times with 40 mL of anhydrous ethanol and 3 times with 40 mL of acetonitrile respectively. After magnetic attraction, transfer them to a reaction vessel with 40 mL of acetonitrile; add 2 g of p-toluenesulfonyl chloride and 2 mL of triethylenediamine to the reaction vessel, and react at 50°C for 2 h. After the reaction is completed, wash it 3 times with anhydrous ethanol, wash it multiple times with purified water until the pH value of the supernatant is close to neutral, remove the supernatant by magnetic attraction, and then add an appropriate amount of purified water to disperse it to obtain magnetic polymer microspheres containing p-toluenesulfonyl groups, with a particle size of about 2.9 μm, and store them in a refrigerator at 2 - 8°C.

[0147] Example 3

[0148] A preparation method of magnetic polymer microspheres containing p-toluenesulfonyl groups specifically includes the following steps:

[0149] (1) Take 2 g of the magnetic polymer microspheres provided in Preparation Example 3, transfer them to a 250 mL three-necked flask with 40 mL of an aqueous sodium oleate solution with a mass percentage of 0.25%. Take 14 g of methylstyrene and 6 g of ethylene glycol dimethacrylate into a 100 mL beaker, add 0.25 g of azobisisobutyronitrile, and then add 40 mL of an aqueous sodium oleate solution with a mass percentage of 0.25% and transfer. Disperse it with an ultrasonic cell disruptor. Under a nitrogen environment, set the reaction temperature to 60 °C. After reaching the preset temperature, stabilize for 10 min and add the above solvent mixed with the initiator and monomer dropwise to the three-necked flask, control the dropping time to 2 h. After the dropping is completed, react at 60 °C for 20 h. After the reaction is completed, wash it 3 times with purified water by magnetic absorption to obtain magnetic polymer microspheres with a benzene ring;

[0150] (2) Take 2 g of the magnetic polymer microspheres with a benzene ring obtained in step (1), replace them 3 times with 40 mL of absolute ethanol, 40 mL of tetrahydrofuran, and 40 mL of 1,4-dioxane respectively. After magnetic absorption to remove the supernatant, transfer them to a 250 mL three-necked flask with 60 mL of 1,4-dioxane. Add 3.2 g of dichloromethyl methyl ether, and slowly add 3.6 g of tin tetrachloride within 40 min during stirring in an ice-water bath at 5 °C. After the addition of tin tetrachloride is completed, continue to react at 25 °C for 20 h. After the reaction is completed, wash it 3 times with tetrahydrofuran, 3 times with absolute ethanol, and 3 times with purified water to obtain magnetic polymer microspheres with a chloromethyl group;

[0151] (3) Take 2 g of the magnetic polymer microspheres with a chloromethyl group obtained in step (2), replace them 3 times with 40 mL of isopropanol and 40 mL of tetrahydrofuran respectively. After magnetic absorption, transfer the magnetic polymer microspheres to a reaction vessel with 40 mL of tetrahydrofuran. Take 0.1 g of pyridine and add it to the reaction vessel; measure 4 mL of polyethylene glycol 800, dissolve it in 20 mL of tetrahydrofuran, and add it dropwise to the reaction vessel, control the dropping time to 2 h. Under a nitrogen environment, react at 60 °C for 16 h. After the reaction is completed, wash it 3 times with absolute ethanol and 3 times with purified water to obtain magnetic polymer microspheres with a hydroxyl group;

[0152] (4) Take 2 g of the magnetic polymer microspheres with a hydroxyl group obtained in step (3), replace them 3 times with 40 mL of absolute ethanol and 40 mL of tetrahydrofuran respectively. After magnetic absorption, transfer them to a reaction vessel with 40 mL of tetrahydrofuran; add 2 g of p-toluenesulfonyl chloride and 2 g of 2-phenylpyridine to the reaction vessel, react at 30 °C for 2 h. After the reaction is completed, wash it 3 times with absolute ethanol, wash it multiple times with purified water until the pH value of the supernatant is close to neutral. After magnetic absorption to remove the supernatant, add an appropriate amount of purified water to disperse to obtain magnetic polymer microspheres containing p-toluenesulfonyl groups, with a particle size of about 4.0 μm, and store them in a refrigerator at 2 - 8 °C.

[0153] Example 4

[0154] A preparation method of magnetic polymer microspheres containing tosyl groups, specifically including the following steps:

[0155] (1) Take 2 g of the magnetic polymer microspheres provided in Preparation Example 4, transfer them to a 250 mL three-necked flask with 30 mL of a 0.5% (by mass) aqueous Tween solution. Take 11.2 g of styrene and 4.8 g of glycerol dimethacrylate into a 100 mL beaker, add 0.16 g of azobisisobutyronitrile, then add 40 mL of a 0.5% (by mass) aqueous Tween solution thereto, disperse with an ultrasonic cell disruptor. Under a nitrogen atmosphere, set the reaction temperature to 55 °C. After reaching the preset temperature and stabilizing for 10 min, add the above solvent mixed with the initiator and monomer dropwise to the three-necked flask, control the dropping time to 2 h. After the dropping is completed, react at 65 °C for 20 h. After the reaction is completed, wash with purified water by magnetic attraction 3 times to obtain magnetic polymer microspheres with benzene rings;

[0156] (2) Take 2 g of the magnetic polymer microspheres with benzene rings obtained in step (1), replace them 3 times with 40 mL of absolute ethanol and 40 mL of dichloromethane respectively. After magnetic attraction to remove the supernatant, transfer them to a 250 mL three-necked flask with 60 mL of dichloromethane. Add 1.5 g of 2-chloroethyl methyl ether, and slowly add 2.4 g of stannic chloride within 30 min during stirring in an ice-water bath at 5 °C. After the addition of stannic chloride is completed, continue to react at 30 °C for 16 h. After the reaction is completed, wash with absolute ethanol 3 times and with purified water 3 times to obtain magnetic polymer microspheres with chloromethyl groups;

[0157] (3) Take 2 g of the magnetic polymer microspheres with chloromethyl groups obtained in step (2), replace them 3 times with 40 mL of isopropanol and 40 mL of tetrahydrofuran respectively. After magnetic attraction, transfer the magnetic polymer microspheres to a reaction vessel with 40 mL of tetrahydrofuran. Take 0.2 g of sodium carbonate and add it to the reaction vessel; measure 4 mL of n-butanol dissolved in 20 mL of tetrahydrofuran and add it dropwise to the reaction vessel, control the dropping time to 2 h. Under a nitrogen atmosphere, react at 50 °C for 20 h. After the reaction is completed, wash with absolute ethanol 3 times and with purified water 3 times to obtain magnetic polymer microspheres with hydroxyl groups;

[0158] (4) Take 2 g of the magnetic polymer microspheres with hydroxyl groups obtained in step (3), replace them 3 times with 40 mL of absolute ethanol and 40 mL of dimethyl sulfoxide respectively. After magnetic attraction, transfer them to a reaction vessel with 40 mL of dimethyl sulfoxide; add 2 g of p-toluenesulfonyl chloride and 2 g of N-methylmorpholine to the reaction vessel, and react at 30 °C for 24 h. After the reaction is completed, wash with absolute ethanol 3 times, wash with purified water multiple times until the pH value of the supernatant is close to neutral. After magnetic attraction to remove the supernatant, add an appropriate amount of purified water to disperse to obtain magnetic polymer microspheres containing tosyl groups, with a particle size of about 7.2 μm, and store them in a refrigerator at 2 - 8 °C.

[0159] Example 5

[0160] A method for preparing magnetic polymer microspheres containing p-toluenesulfonyl group, which is different from Example 1 in that the addition amount of styrene is 3.2 g, the addition amount of divinylbenzene is 4.8 g, and other substances, conditions and steps are the same as those in Example 1;

[0161] The particle size of the magnetic polymer microspheres containing p-toluenesulfonyl group finally obtained in this example is about 4.1 μm.

[0162] Example 6

[0163] A method for preparing magnetic polymer microspheres containing p-toluenesulfonyl group, which is different from Example 1 in that the addition amount of styrene is 6.8 g, the addition amount of divinylbenzene is 1.2 g, and other substances, conditions and steps are the same as those in Example 1;

[0164] The particle size of the magnetic polymer microspheres containing p-toluenesulfonyl group finally obtained in this example is about 3.3 μm.

[0165] Comparative Example 1

[0166] A method for preparing magnetic polymer microspheres containing p-toluenesulfonyl group,

[0167] (1) Take 2 g of the magnetic polymer microspheres provided in Preparation Example 1 and transfer them to a 250 mL three-necked flask with 40 mL of an aqueous solution of sodium naphthenate with a mass percentage of 0.5%. Take 6.5 g of 4-tert-butylstyrene and 3.2 g of divinylbenzene into a 100 mL beaker, add 0.9 g of azobisisobutyronitrile, and then add 40 mL of an aqueous solution of sodium naphthenate with a mass percentage of 0.5% thereto. Disperse it with an ultrasonic cell disruptor. Under a nitrogen atmosphere, set the reaction temperature to 75 °C. After reaching the preset temperature and stabilizing for 10 minutes, add the above solvent mixed with the initiator and monomer dropwise to the three-necked flask, control the dropping time to be 2 h. After the dropping is completed, continue the reaction for 16 h. After the reaction is completed, wash it with purified water by magnetic attraction 3 times to obtain magnetic polymer microspheres with a benzene ring;

[0168] (2) Take 2 g of the magnetic polymer microspheres with a benzene ring obtained in step (1), replace them 3 times with 40 mL of absolute ethanol and 40 mL of tetrahydrofuran. After magnetically attracting and removing the supernatant, transfer them to a reaction vessel with 40 mL of tetrahydrofuran. Add 2 g of p-toluenesulfonyl chloride and 4 mL of tetramethylethylenediamine to the reaction vessel and react at 37 °C for 5 h. After the reaction is completed, wash it 3 times with absolute ethanol and wash it with purified water multiple times until the pH value of the supernatant is close to neutral. After magnetically attracting and removing the supernatant, add an appropriate amount of purified water to disperse it to obtain magnetic polymer microspheres containing p-toluenesulfonyl group with a particle size of about 3.4 μm, and store them in a refrigerator at 2-8 °C.

[0169] Performance test:

[0170] (1) Particle size: The magnetic polymer microspheres containing p-toluenesulfonyl group were tested by using a scanning electron microscope (Hitachi Regulus 8100).

[0171] The scanning electron microscope image of the magnetic polymer microspheres containing p-toluenesulfonyl group provided in Example 1 tested by using the above test method is as shown in Figure 1 shown. It can be seen from Figure 1 that the magnetic polymer microspheres containing p-toluenesulfonyl group provided in Example 1 have uniform size, the particle size is 3.2 ± 0.1 μm, the surface structure has small irregular protrusions and depressions to varying degrees, and the specific surface area is increased compared with the magnetic polymer microspheres without surface modification, which helps to improve the specific adsorption of antibodies in applications.

[0172] (2) Specific adsorption performance: The magnetic polymer microspheres containing p-toluenesulfonyl group were tested on a fully automatic chemiluminescence immunoassay analyzer; specifically, it includes: 10 mg of the magnetic polymer microspheres containing p-toluenesulfonyl group were coated with PCT monoclonal antibody and then diluted to 0.5 mg / mL, and a double-antibody sandwich reagent was composed with alkaline phosphatase-labeled PCT antibody to test the standard PCT antigen, and the signal values at different PCT antigen concentrations were recorded.

[0173] The magnetic polymer microspheres containing p-toluenesulfonyl group provided in Examples 1 to 6 and Comparative Example 1 were tested according to the above test method, and the test results are shown in Table 1:

[0174] Table 1

[0175]

[0176] It can be seen from the data in Table 1 that

[0177] when the antigen concentration is 0 μIU / mL, the signal values of Examples 1 to 4 are relatively low, indicating that the magnetic polymer microspheres containing p-toluenesulfonyl group provided in Examples 1 to 4 have low non-specific adsorption to non-target antibodies, and after the antigen concentration gradually increases, the signal values increase very significantly;

[0178] Compared with Example 1, the background value of Comparative Example 1 is significantly higher.

[0179] Compared with Example 1, the magnetic polymer microspheres containing p-toluenesulfonyl group provided in Examples 5 to 6 have higher background values and higher non-specific adsorption to non-target antibodies, and as the antigen concentration increases, the signal values are significantly smaller than those of Example 1.

[0180] The applicant declares that the present invention illustrates a magnetic polymer microsphere containing p-toluenesulfonyl group, its preparation method and application through the above-mentioned embodiments. However, the present invention is not limited to the above-mentioned embodiments, that is, it does not mean that the present invention must rely on the above-mentioned embodiments to be implemented. Those skilled in the art should understand that any improvement to the present invention, the equivalent substitution of each raw material of the products of the present invention, the addition of auxiliary components, the selection of specific methods, etc., all fall within the protection scope and the disclosure scope of the present invention.

Claims

1. A preparation method of magnetic polymer microspheres containing tosyl groups, characterized in that, The preparation method includes: coating the magnetic polymer microspheres with a benzene ring-containing polymer, followed by chloromethyl modification, hydroxyl modification, and p-toluenesulfonyl chloride modification in sequence to obtain the magnetic polymer microspheres containing p-toluenesulfonyl groups.

2. The preparation method according to claim 1, wherein The magnetic polymer microspheres are prepared by the following method, which includes the following steps: (1A) Reacting polymer microspheres, epoxy monomers, functional monomers, crosslinking monomers, and an initiator in an aqueous solution of an emulsifier to obtain polymer microspheres with epoxy groups; (2A) Reacting the polymer microspheres with epoxy groups obtained in step (1A) with an amino compound to obtain polymer microspheres with amino groups; (3A) Depositing magnetic nanoparticles on the surface of the polymer microspheres with amino groups obtained in step (2A) to obtain the magnetic polymer microspheres; Preferably, the particle size of the polymer microspheres in step (1A) is 0.2 - 3 μm; Preferably, the polymer microspheres in step (1A) are prepared by any one of emulsion polymerization, microemulsion polymerization, soap-free emulsion polymerization, or dispersion polymerization; Preferably, the polymer microspheres in step (1A) include polystyrene microspheres or polyacrylate microspheres; Preferably, based on the mass of the polymer microspheres in step (1A) being 1 g, the total mass of the epoxy monomers, functional monomers, and crosslinking monomers is 5 - 500 g; Preferably, the mass ratio of the epoxy monomers, functional monomers, and crosslinking monomers in step (1A) is (10 - 30):(50 - 85):(5 - 20); Preferably, the epoxy monomers in step (1A) include any one or a combination of at least two of glycidyl methacrylate, n-butyl glycidyl ether, resorcinol diglycidyl ether, ethylene glycol diglycidyl ether, tert-butyl glycidyl ether, or allyl glycidyl ether; Preferably, the functional monomers in step (1A) include any one or a combination of at least two of methyl acrylate, methyl methacrylate, ethyl acrylate, ethyl methacrylate, n-butyl acrylate, n-butyl methacrylate, 2-hydroxyethyl methacrylate, isobutyl acrylate, or methacrylic acid; Preferably, the crosslinking monomers in step (1A) include any one or a combination of at least two of butanediol dimethacrylate, ethylene glycol dimethacrylate, glycerol dimethacrylate, trimethylolpropane trimethacrylate, divinylbenzene, butadiene, or N,N'-methylenebisacrylamide; Preferably, based on the total mass of the epoxy monomers, functional monomers, and crosslinking monomers in step (1A) being 100 g, the mass of the initiator is 1 - 10 g; Preferably, the initiator in step (1A) includes any one or a combination of at least two of potassium persulfate, ammonium persulfate, sodium persulfate, azobisisobutyronitrile, azodiisooctanenitrile, dimethyl azodicarboxylate, cumene hydroperoxide, dicumyl peroxide, benzoyl peroxide, or tert-butyl peroxybenzoate; Preferably, the mass percentage content of the emulsifier in the aqueous solution of the emulsifier in step (1A) is 0.01 - 5%; Preferably, the emulsifier in step (1A) includes any one or a combination of at least two of sodium naphthenate, sodium oleate, sodium dodecyl sulfate, sodium benzenesulfonate, sodium isopropylsulfonate, potassium methylene bis(isopropylnaphthalenesulfonate), diacetyl tartaric acid esters of mono- and diglycerides, polyoxyethylene sorbitan fatty acid esters, octylphenol polyoxyethylene ether, dodecylphenol ether or dinonylphenol ether; Preferably, a stabilizer is further added during the reaction in step (1A); Preferably, the stabilizer includes any one or a combination of at least two of polyvinylpyrrolidone, polyethylene glycol, polyvinyl alcohol, methylcellulose, ethylcellulose or polyacrylic acid; Preferably, the temperature of the reaction in step (1A) is 30-80 °C; Preferably, the reaction time in step (1A) is 8-40 h; Preferably, based on 1 g of the polymer microspheres with epoxy groups in step (2A), the amount of the amino compound is 10-100 mol; Preferably, the amino compound in step (2A) includes any one or a combination of at least two of diisopropylamine, ethanolamine, triethanolamine, ethylenediamine, ammonia water, tetraethylenepentamine or polyethyleneimine; Preferably, the temperature of the reaction in step (2A) is 30-80 °C; Preferably, the reaction time in step (2A) is 8-20 h; Preferably, the method for depositing magnetic nanoparticles in step (3A) specifically includes: reacting the polymer microspheres with amino groups obtained in step (2A), ferric salt, ferrous salt and surfactant in a solvent to complete the deposition of magnetic nanoparticles; Preferably, based on 1 g of the polymer microspheres with amino groups, the amount of the ferric salt is 1-20 mmol, and the amount of the ferrous salt is 1-15 mmol; Preferably, the ferric salt includes any one or a combination of at least two of ferric chloride, ferric sulfate or ferric nitrate; Preferably, the ferrous salt includes any one or a combination of at least two of ferrous chloride, ferrous sulfate, ferrous ammonium sulfate or ferrous nitrate; Preferably, the surfactant includes any one or a combination of at least two of undecylenic acid, methyl undecylenate, butyl undecylenate, oleic acid, methyl palmitoleate, sodium oleate, dodecylamine, octadecylamine, sodium laurate, coconut amine, sodium stearate, sodium dodecyl sulfate or sodium dodecylbenzenesulfonate; Preferably, the reaction is carried out under alkaline conditions; Preferably, the temperature of the reaction is 50-90 °C; Preferably, the reaction time is 0.5-5 h.

3. The preparation method according to claim 1 or 2, characterized in that, The preparation method specifically includes the following steps: (1) Reacting magnetic polymer microspheres, benzene ring-containing monomers, crosslinking monomers and initiators in an aqueous solution of an emulsifier to obtain magnetic polymer microspheres with benzene rings; (2) Reacting the magnetic polymer microspheres with benzene rings obtained in step (1) and chloromethyl compounds in a solvent to obtain magnetic polymer microspheres with chloromethyl groups; (3) Reacting the magnetic polymer microspheres with chloromethyl groups obtained in step (2) and hydroxyl compounds in a solvent to obtain magnetic polymer microspheres with hydroxyl groups; (4) React the magnetic polymer microspheres with hydroxyl groups obtained in step (3) and p-toluenesulfonyl chloride in a solvent to obtain the magnetic polymer microspheres containing p-toluenesulfonyl groups.

4. The preparation method according to claim 3, wherein Based on the mass of the magnetic polymer microspheres described in step (1) being 1 g, the total mass of the benzene ring-containing monomer and the crosslinking monomer is 0.2 - 75 g; Preferably, the mass ratio of the benzene ring-containing monomer to the crosslinking monomer described in step (1) is (50 - 80):(20 - 50); Preferably, the benzene ring-containing monomer described in step (1) includes any one or a combination of at least two of styrene, methylstyrene, or 4-tert-butylstyrene; Preferably, the crosslinking monomer described in step (1) includes any one or a combination of at least two of butanediol dimethacrylate, ethylene glycol dimethacrylate, glycerol dimethacrylate, trimethylolpropane trimethacrylate, divinylbenzene, butadiene, or N,N-methylenebisacrylamide; Preferably, based on the total mass of the benzene ring-containing monomer and the crosslinking monomer described in step (1) being 100%, the mass of the initiator is 1 - 10%; Preferably, the initiator described in step (1) includes any one or a combination of at least two of potassium persulfate, ammonium persulfate, sodium persulfate, azobisisobutyronitrile, azodiisooctanenitrile, dimethyl azodicarboxylate, cumene hydroperoxide, dicumyl peroxide, benzoyl peroxide, or tert-butyl peroxybenzoate; Preferably, the temperature of the reaction described in step (1) is 50 - 80 °C; Preferably, the time of the reaction described in step (1) is 3 - 20 h.

5. The preparation method according to claim 3 or 4, characterized in that, Based on the amount of the magnetic polymer microspheres with a benzene ring described in step (2) being 1 g, the amount of the chloromethyl compound is 10 - 100 mmol; Preferably, the chloromethyl compound described in step (2) includes any one or a combination of at least two of dichloromethyl methyl ether, 2-chloroethyl methyl ether, dichloroethyl ether, paraformaldehyde, dimethoxymethane, or chloromethyl ethyl ether; Preferably, the solvent described in step (2) includes any one or a combination of at least two of 1,4-dioxane, dichloromethane, chloroform, or dimethyl sulfoxide; Preferably, a catalyst is further added in the reaction described in step (2); Preferably, based on the amount of the magnetic polymer microspheres with a benzene ring described in step (2) being 1 g, the amount of the catalyst is 5 - 50 mmol; Preferably, the catalyst includes any one or a combination of at least two of tin tetrachloride, aluminum trichloride, boron trifluoride, or zinc chloride; Preferably, the temperature of the reaction described in step (2) is 20 - 40 °C; Preferably, the time of the reaction described in step (2) is 16 - 24 h.

6. The preparation method according to any one of claims 3 to 5, characterized in that, Based on the mass of the magnetic polymer microspheres with chloromethyl groups described in step (3) being 1 g, the mass of the hydroxyl compound is 0.1 - 100 g; Preferably, the hydroxyl compound described in step (3) includes any one or a combination of at least two of n-butanol, isobutanol, n-propanol, ethylene glycol, polyethylene glycol 200, polyethylene glycol 400, polyethylene glycol 600, or polyethylene glycol 800; Preferably, the solvent in step (3) includes any one or a combination of at least two of ethanol, n-hexane, dichloromethane, chloroform, acetonitrile, or tetrahydrofuran. Preferably, the reaction in step (3) is carried out under alkaline conditions; Preferably, the temperature of the reaction in step (3) is 50 - 80 °C; Preferably, the reaction time in step (3) is 8 - 20 h.

7. The preparation method according to any one of claims 3 to 6, characterized in that, Based on the mass of 1 g of the magnetic polymer microspheres with hydroxyl groups in step (4), the mass of p-toluenesulfonyl chloride is 0.1 - 10 g; Preferably, the solvent in step (4) includes any one or a combination of at least two of tetrahydrofuran, 1,4-dioxane, dimethyl sulfoxide, N,N-dimethylformamide, ether, or acetonitrile; Preferably, the temperature of the reaction in step (4) is 5 - 50 °C; Preferably, the reaction time in step (4) is 2 - 30 h.

8. A magnetic polymer microsphere containing p-toluenesulfonyl group, characterized in that, The magnetic polymer microspheres are prepared by the preparation method described in any one of claims 1 - 7.

9. The magnetic polymer microspheres according to claim 8, wherein The particle size of the magnetic polymer microspheres is 3 - 10 μm.

10. Use of the magnetic polymer microspheres according to claim 8 or 9 in targeted capture or separation and purification.

Images