Universal molecularly imprinted nanochannel membrane and super-assembly preparation method thereof

Abstract

The application belongs to the technical field of nano ion channel, and particularly relates to a general molecular imprinting nano channel membrane and a super-assembly preparation method thereof. The molecular imprinting polymer is prepared by a precipitation polymerization method, a suitable functional monomer is selected according to the chemical structure of a target object, the molecular imprinting polymer with specific target imprinting cavities is obtained after elution of a template, and finally, the molecular imprinting polymer is loaded on a porous alumina AAO film by using a super-assembly technology to obtain a molecular imprinting nano composite membrane. The molecular imprinting nano composite membrane can realize rapid, high-selectivity and high-sensitivity identification and detection of specific target objects. The application provides a new idea for the nano flow control membrane device in the field of high-selectivity identification of target molecules.

Description

TECHNICAL FIELD

[0001] The application belongs to the technical field of nano ion channels, and particularly relates to a general molecular imprinting nano channel membrane and a super-assembly preparation method thereof. BACKGROUND

[0002] Ion nano channels in biological systems carry cell signal transmission, energy conversion and response to various physiological stimuli, so as to achieve the purpose of controlling ion transmembrane transport and maintaining the balance of physiological environment. Inspired by biological nano channels, artificial ion nano channel biosensing application platforms have developed rapidly in recent years and are widely used in the detection of proteins, nucleic acids, amino acids, tumor cells and ions. Compared with other analytical sensors, artificial ion nano channel sensors have the advantages of high sensitivity, fast detection speed, reversible response and high signal-to-noise ratio. Artificial ion nano channels not only provide ion current as a signal in response to external stimuli or specific recognition, but also provide a nanoscale limited space for the interaction between functional molecules and analytes, which makes the nano channel have higher sensing sensitivity and better recognition selectivity. However, with the rapid development of artificial ion nano channels, the demand for accuracy in sensitivity and selectivity of analysis and detection is gradually increasing, and its disadvantages are gradually emerging. At present, the selective recognition of a certain analyte based on artificial ion nano channels is established on the interaction between molecules (such as covalent interaction, complexation, coordination and redox interaction), which is difficult to achieve high selective recognition of target substances in terms of spatial configuration, shape and size.

[0003] Molecular imprinting technology is a synthesis technology for obtaining a molecular imprinting polymer that is mutually compatible with a target analyte in terms of spatial shape, size and functional groups, and can accurately recognize the chemical bonds and spatial configuration of the target analyte in a complex environment. Its principle is described as the "lock and key principle". Molecular imprinting technology generates a polymer through self-assembly of a target analyte and a functional monomer under the action of a crosslinking agent, and finally removes the target analyte from the polymer by an eluent to obtain a molecular imprinting polymer. At this time, there are a large number of imprinting recognition sites in the molecular imprinting polymer that are chemically bonded to the target analyte, have the same spatial configuration and shape, and only the analyte that completely matches the imprinting site can stimulate the molecular imprinting polymer to produce a response signal, which can effectively achieve high selective recognition of the chemical bonds and spatial configuration of the target analyte. SUMMARY

[0004] The application aims to solve the selective recognition bottleneck faced by current nano channel membrane devices and provides a general molecular imprinting nano channel membrane based on an interface super-assembly strategy and a preparation method thereof. The channel composite membrane prepared by the application can be used to construct a nano-fluidic device with specific recognition, and has a good application prospect in the fields of intelligent ion transport and high selective molecular recognition.

[0005] The application utilizes silica spheres as carriers, generates polymers under the action of cross-linking agent through chemical force (covalent action, non-covalent action and semi-covalent action) between functional monomers and template molecules (target analyte), uses a mixed solution of methanol / acetic acid as eluent, elutes the target from the polymers by solution extraction, obtains molecularly imprinted polymers, and finally prepares a molecularly imprinted nanocomposite membrane on an AAO substrate by vacuum filtration. The composite membrane is obtained by close packing of nanospheres, and has abundant ion transmission channels and imprinted recognition sites. The technical scheme of the application is specifically introduced as follows.

[0006] The application provides a universal molecularly imprinted nanochannel membrane super-assembly preparation method, comprising the following steps:

[0007] (1) dispersing monodisperse silica nanospheres in a solvent to obtain a silica dispersion liquid;

[0008] (2) adding functional monomers and template molecules to the silica dispersion liquid of step (1) and standing at room temperature to perform pre-assembly;

[0009] (3) adding a cross-linking agent and an initiator to the pre-assembled system and performing polymerization under heating, after the polymerization is completed, performing solid-liquid separation, and washing the solid polymerization product;

[0010] (4) dispersing the solid polymerization product in an eluent, leaching the template molecules from the solid polymerization product by using the eluent under heating, after the leaching is completed, sequentially performing solid-liquid separation, washing and drying to obtain molecularly imprinted polymer nanoparticles;

[0011] (5) dispersing the molecularly imprinted polymer nanoparticles in an alcohol-water mixed solvent, loading the molecularly imprinted nanoparticles on a porous alumina AAO by a vacuum filtration method to prepare a molecularly imprinted nanochannel membrane.

[0012] In the application, in step (1), the silica nanospheres are prepared by an alkoxide hydrolysis method in a mixed solvent of ethanol and water, with concentrated ammonia as a catalyst and tetraethyl orthosilicate TEOS as a silicon source; the prepared silica nanoparticles are uniform in size and good in monodispersity.

[0013] In the application, in step (1), the solvent is acetonitrile, and the concentration of the silica dispersion liquid is 1.0-3.0 g / L.

[0014] In the application, in step (2), the mass ratio of the silica, the functional monomer and the template molecule is (7.5-12.5):1:(2-2.5), and the standing time is 1-3 h. In the synthesis process of the molecularly imprinted polymer nanoparticles, the thickness of the imprinting layer is affected by the addition ratio of the functional monomer and the crosslinking agent, and the thickness of the imprinting layer directly affects the performance detection of the molecularly imprinted polymer.

[0015] In the application, in step (2), the functional monomer is acrylamide, and the template molecule is dimethoate. The selected template molecule is different, and the corresponding functional monomer also changes accordingly, so as to obtain a molecularly imprinted polymer which maximizes the matching (covalent interaction, non-covalent interaction or semi-covalent interaction) of the chemical structure of the template molecule (target object).

[0016] In the application, in step (3), the crosslinking agent is ethylene glycol dimethacrylate, the initiator is azobisisobutyronitrile, the addition amount of the crosslinking agent is 0.4%-0.6% of the molar number of the functional monomer, the polymerization temperature is 60-70 DEG C, the polymerization time is 12-36 h, and the solid polymerization product is washed with ethanol.

[0017] In the application, in step (4), the eluent is a mixed solvent of methanol and acetic acid with a volume ratio of 48:2, the feeding ratio of the solid polymerization product and the eluent is 1.0-1.5 g / L, the leaching temperature is 55-65 DEG C, the leaching time is 8-20 h, and the drying is performed in a vacuum drying manner, the vacuum drying temperature is 50-70 DEG C, and the drying time is 8-20 h.

[0018] In the application, in step (5), in the alcohol-water mixed solvent, the volume ratio of ethanol and water is 1:1, and the feeding ratio of the molecularly imprinted polymer nanoparticles and the alcohol-water mixed solvent is 0.4-0.5 g / L. The molecularly imprinted nanofilm made of AAO as the substrate can be supported by the interfacial super-assembly strategy. Different thicknesses of the molecularly imprinted nanocomposite film can be obtained by changing the addition amount of the molecularly imprinted polymer nanoparticles.

[0019] The application also provides a general molecularly imprinted nanochannel film prepared by the above super-assembly preparation method.

[0020] In the above, the application loads the molecularly imprinted polymer nanoparticles in AAO by the molecular imprinting technology and the interfacial super-assembly strategy to obtain a composite nanochannel film. Compared with the prior art, the application has the following beneficial effects:

[0021] The application prepares a molecular imprinting polymer by a precipitation polymerization method, selects a proper functional monomer according to the chemical structure of a target object, obtains a molecular imprinting polymer with specific target imprinting cavities after eluting a template, and finally loads the molecular imprinting polymer on a porous alumina AAO film by using a super assembly technology to obtain a molecular imprinting nano composite film, so that the molecular imprinting nano composite film can realize rapid, high selectivity and high sensitivity identification and detection of specific target objects. BRIEF DESCRIPTION OF DRAWINGS

[0022] Figure 1 is a preparation flowchart of the application based on an interface super assembly strategy.

[0023] Figure 2 is a transmission electron microscope (TEM) image of the molecular imprinting polymer obtained in Example 1 of the application.

[0024] Figure 3 is a scanning electron microscope (SEM) image of the molecular imprinting polymer (a) and the nano composite film cross section (b) prepared in Example 1 of the application.

[0025] Figure 4 is a Fourier transform infrared spectrogram of the molecular imprinting polymer before and after elution of the pesticide dimethoate obtained in Example 1.

[0026] Figure 5 is an ultraviolet absorption graph of the dimethoate imprinting polymer prepared in Example 1 of the application before and after elution of dimethoate. DETAILED DESCRIPTION

[0027] In order to be able to more specifically describe the application, the application is described in detail below in combination with examples and drawings:

[0028] The preparation method of the molecular imprinting nano composite film in Example 1 specifically includes the following steps:

[0029] Step 1: 32.4 mL of ethanol, 49.5 mL of water and 18 mL of ammonia water are mixed in a 500 mL flask, and are uniformly stirred by magnetic stirring at 1100 rpm;

[0030] Step 2: 91 mL of ethanol and 9 mL of tetraethyl silicate are added to the above reaction solution, and are stirred at room temperature at 400 rpm for 2 h;

[0031] Step 3: After the reaction is completed, the obtained product is centrifuged, is washed with water and ethanol alternately, and finally is collected and dried in a 60℃ vacuum drying box overnight;

[0032] Step 4: Take an appropriate amount of the above dried product 100 mg, add 60 mL of acetonitrile, ultrasonic dispersion, then add 10.2 mg of functional monomer acrylamide and 22.9 mg of target analyte dimethoate, stand for 2 h;

[0033] Step 5: After waiting for the above reaction system to complete the pre-assembly, 134.4 μL of crosslinking agent ethylene glycol dimethacrylate is added to the system, nitrogen is blown for 15-20 min to remove air in the system, then 10 mg of reaction initiator azobisisobutyronitrile is added, and the reaction system is placed in a 65°C oil bath for reaction for 24 h;

[0034] Step 6: After the above polymerization reaction is completed, the polymerization product is collected by centrifugation and washed with ethanol for 3 times;

[0035] Step 7: After the product is washed, the solid-liquid ratio is 1.2 g / L, and the whole product is dispersed in methanol / acetic acid (v / v=48:2), ultrasonic dispersion, then the system is placed in a 60°C oil bath for extraction overnight (the synthesis method of the non-imprinted nanochannel is the same as that of the molecularly imprinted nanochannel, except that the target analyte is not added);

[0036] Step 8: The above product is collected by centrifugation and washed with ethanol for 3 times, and then placed in a 60°C vacuum drying box for drying overnight to obtain molecularly imprinted polymer nanoparticles. Through the eluted molecularly imprinted nanochannel, there are imprinting cavities matching the shape, size and functional groups of dimethoate molecules, which can selectively adsorb dimethoate molecules in subsequent tests and cause current changes;

[0037] Step 9: Take an appropriate amount of molecularly imprinted polymer nanoparticles, disperse them in water / ethanol (v / v=1:1) to obtain a solution with a concentration of 0.4 g / L;

[0038] Step 10: Take 1.0 mL of the above solution, load the molecularly imprinted nanoparticles on AAO (pore size 160-200 nm) by vacuum filtration to prepare a molecularly imprinted nanocomposite film.

[0039] Example 2 The preparation method of the molecularly imprinted nanocomposite film, specifically comprising the following steps:

[0040] Step 1: Mix 32.4 mL of ethanol, 49.5 mL of water and 18 mL of ammonia water in a 500 mL flask, and stir uniformly at 1100 rpm under magnetic stirring;

[0041] Step 2: Add 91 mL of ethanol and 9 mL of tetraethyl orthosilicate to the above reaction solution, and stir at room temperature for 2 h at 400 rpm;

[0042] Step 3: After the reaction is completed, the obtained product is centrifuged, washed with water and ethanol alternately, and finally collected and dried in a 60 ℃ vacuum drying oven overnight;

[0043] Step 4: Take an appropriate amount of the dried product 100 mg, add 60 mL of acetonitrile, ultrasonically disperse uniformly, then add 9.24 mg of functional monomer acrylamide and 20.8 mg of target analyte dimethoate, and stand for 2 h;

[0044] Step 5: After the pre-assembly of the above reaction system is completed, 123.2 μL of crosslinking agent ethylene glycol dimethacrylate is added to the system, nitrogen is blown for 15-20 min to remove air in the system, then 10 mg of reaction initiator azobisisobutyronitrile is added, and the reaction system is placed in a 65 ℃ oil bath for reaction for 24 h;

[0045] Step 6: After the above polymerization reaction is completed, the polymerization product is collected by centrifugation and washed with ethanol for 3 times;

[0046] Step 7: After the product is washed, the whole product is dispersed in methanol / acetic acid (v / v=48:2) according to the solid-liquid ratio of 1.0 g / L, ultrasonically dispersed uniformly, and then placed in a 60 ℃ oil bath for extraction overnight (the synthesis method of the non-imprinted nanochannel is the same as that of the molecularly imprinted nanochannel, except that no target analyte is added);

[0047] Step 8: The above product is collected by centrifugation, washed with ethanol for 3 times, and then placed in a 60 ℃ vacuum drying oven for drying overnight to obtain molecularly imprinted polymer nanoparticles. Through the imprinted cavities in the eluted molecularly imprinted nanochannel which are matched with the shape, size and functional groups of dimethoate molecules, dimethoate molecules can be selectively adsorbed in the subsequent test to cause a change in current;

[0048] Step 9: Take an appropriate amount of molecularly imprinted polymer nanoparticles, disperse them uniformly with water / ethanol (v / v=1:1) to obtain a 0.5 g / L solution;

[0049] Step 10: Take 1.0 mL of the above solution, load the molecularly imprinted nanoparticles on AAO by vacuum filtration to prepare a molecularly imprinted nanocomposite film.

[0050] The preparation method of the molecularly imprinted nanocomposite film of Example 3 specifically includes the following steps:

[0051] Step 1: Mix 32.4 mL of ethanol, 49.5 mL of water and 18 mL of ammonia water in a 500 mL flask, and uniformly stir under 1100 rpm magnetic stirring;

[0052] Step 2: 91 mL of ethanol and 9 mL of tetraethyl silicate were added to the above reaction solution, which was stirred at room temperature at 400 rpm for 2 h;

[0053] Step 3: After the reaction was completed, the obtained product was centrifuged, washed with water and ethanol alternately, and finally collected and dried in a 60 ℃ vacuum drying oven overnight;

[0054] Step 4: An appropriate amount of the above dried product 100 mg was taken and added to 60 mL of acetonitrile, and after being uniformly dispersed by ultrasonic, 11.34 mg of functional monomer acrylamide and 25.46 mg of target analyte dimethoate were added, and left to stand for 2 h;

[0055] Step 5: After the pre-assembly of the above reaction system was completed, 151.2 μL of crosslinking agent ethylene glycol dimethacrylate was added to the system, nitrogen was blown for 15-20 min to remove air in the system, then 10 mg of reaction initiator azobisisobutyronitrile was added, and the reaction system was placed in a 65 ℃ oil bath for reaction for 24 h;

[0056] Step 6: After the above polymerization reaction was completed, the polymerization product was collected by centrifugation and washed with ethanol three times;

[0057] Step 7: After the product was washed, the whole product was dispersed in methanol / acetic acid (v / v=48:2) at a solid-liquid ratio of 1.5 g / L, and after being uniformly dispersed by ultrasonic, the system was placed in a 60 ℃ oil bath for extraction overnight (the synthesis method of the non-imprinted nanochannel is the same as that of the molecularly imprinted nanochannel, except that no target analyte is added);

[0058] Step 8: The above product was collected by centrifugation and washed with ethanol three times and then placed in a 60 ℃ vacuum drying oven to dry overnight to obtain molecularly imprinted polymer nanoparticles. Through the elution of the molecularly imprinted nanochannel having imprinting cavities matching the shape, size and functional groups of dimethoate molecules, dimethoate molecules can be selectively adsorbed in the subsequent test to cause a change in current;

[0059] Step 9: An appropriate amount of molecularly imprinted polymer nanoparticles was taken and dispersed uniformly with water / ethanol (v / v=1:1) to obtain a 0.5 g / L solution;

[0060] Step 10: 1.0 mL of the above solution was taken, and the molecularly imprinted nanoparticles were loaded on AAO (pore size 160-200 nm) by vacuum filtration to prepare a molecularly imprinted nanocomposite film.

[0061] Figure 1 The preparation flowchart of the molecularly imprinted nanocomposite film of Example 1 is shown in FIG. 1, Figure 2The TEM image of the molecular imprinting polymer obtained in Example 1 shows that a thin imprinting layer of about 25 nm exists outside the silicon ball from the electron microscope image. Figure 3 a and Figure 3 b are scanning electron microscope images of the molecular imprinting polymer and the molecular imprinting nanocomposite film respectively, Figure 3 The nanoparticles in a are uniform in size, Figure 3 The molecular imprinting nanocomposite film particles in b are closely arranged, and the thickness is about 1.3 microns. Figure 4 The Fourier transform infrared spectrograms of the molecular imprinting polymer obtained by taking the pesticide dimethoate as an example before and after elution of dimethoate show that the characteristic peak of dimethoate disappears after the molecular imprinting polymer elutes the template molecule dimethoate, indicating that dimethoate is successfully eluted from the molecular imprinting polymer. Figure 5 The ultraviolet absorption diagrams of the dimethoate imprinting polymer prepared in the application before and after elution of dimethoate show that the characteristic absorption peak of dimethoate at 230 nm obviously disappears after elution of dimethoate, thereby indicating that the molecular imprinting polymer with specific cavities of dimethoate molecules is successfully synthesized, and the molecular imprinting nanocomposite film is successfully prepared, which can be used in subsequent nanochannel experiment detection.

[0062] In summary, the application successfully prepares a general molecular imprinting nanocomposite film through an interface super-assembly strategy, which can realize rapid, high selectivity and high sensitivity recognition and detection of specific target objects. The application provides a new idea for nanofluidic membrane devices in the field of high selectivity recognition of target molecules.

Claims

1. A method for the superassembly preparation of a general molecularly imprinted nanochannel membrane, characterized in that, The method comprises the following steps: (1) dispersing monodisperse silica nanospheres in a solvent to obtain a silica dispersion; (2) adding a functional monomer and a template molecule to the silica dispersion of step (1) and standing at room temperature to perform pre-assembly; (3) adding a crosslinking agent and an initiator to the pre-assembled system and performing a polymerization reaction under heating, and after the polymerization reaction is completed, solid-liquid separation, washing of the solid polymerization product, and drying are performed; (4) dispersing the solid polymerization product in an eluent, leaching the template molecule from the solid polymerization product by using the eluent under heating, and after the leaching is completed, sequentially performing solid-liquid separation, washing, and drying to obtain a molecularly imprinted polymer nanoparticle; (5) dispersing the molecularly imprinted polymer nanoparticle in an alcohol-water mixed solvent, loading the molecularly imprinted nanoparticle on a porous alumina AAO by vacuum filtration to prepare a molecularly imprinted nanochannel membrane.

2. The hyper-assembly production method according to claim 1, wherein, In step (1), the silica nanospheres are prepared by an alkoxide hydrolysis method in an alcohol-water mixed solvent using concentrated ammonia as a catalyst and tetraethyl orthosilicate TEOS as a silicon source.

3. The hyper-assembly production method according to claim 1, characterized in that, In step (1), the solvent is acetonitrile, and the concentration of the silica dispersion is 1.0-3.0 g / L.

4. The hyper-assembly production method according to claim 1, wherein, In step (2), the mass ratio of silica, functional monomer, and template molecule is (7.5-12.5):1:(2-2.5), and the standing time is 1-3 h.

5. The hyper-assembly production method according to claim 1, wherein, In step (2), the functional monomer is acrylamide, and the template molecule is dimethoate.

6. The hyper-assembly production method according to claim 1, wherein, In step (3), the crosslinking agent is ethylene glycol dimethacrylate, the initiator is azobisisobutyronitrile, the addition amount of the crosslinking agent is 0.4%-0.6% of the number of moles of the functional monomer, the polymerization temperature is 60-70°C, the polymerization time is 12-36 h, and the solid polymerization product is washed with ethanol.

7. The hyper-assembly production method according to claim 1, wherein, In step (4), the eluent is a mixed solvent of methanol and acetic acid in a volume ratio of 48:2, the feeding ratio of the solid polymerization product to the eluent is 1:1-1.5:1 g / L, the leaching temperature is 55-65°C, the leaching time is 8-20 h, the washing is performed with ethanol, the drying is performed by a vacuum drying method, the vacuum drying temperature is 50-70°C, and the drying time is 8-20 h.

8. The hyper-assembly production method of claim 1, wherein, In step (5), in the alcohol-water mixed solvent, the volume ratio of ethanol to water is 1:1, and the feeding ratio of the molecularly imprinted polymer nanoparticle to the alcohol-water mixed solvent is 2:5-1:2 g / L.

9. A general-purpose molecularly imprinted nanochannel membrane prepared by the super-assembly preparation method according to claim 1.

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