CN110343250B

CN110343250B - Preparation method of molecularly imprinted high molecular polymer material formed based on two or more times of polymerization

Info

Publication number: CN110343250B
Application number: CN201910568010.8A
Authority: CN
Inventors: 李房有; 林煊豪
Original assignee: National University of Singapore
Current assignee: National University of Singapore
Priority date: 2018-06-28
Filing date: 2019-06-27
Publication date: 2021-11-30
Anticipated expiration: 2039-06-27
Also published as: CN110343250A

Abstract

The invention relates to the technical field of high molecular materials, in particular to a preparation method of a molecularly imprinted high molecular polymer material formed on the basis of two or more times of polymerization.

Description

Preparation method of molecularly imprinted high molecular polymer material formed based on two or more times of polymerization

Technical Field

Background

Molecularly imprinted polymeric materials have been widely used in solid phase extraction, catalysts, and sensing of chemical and biochemical molecules. The traditional preparation method of the molecularly imprinted polymer material is to mix template molecules and monomers and then polymerize and crosslink the mixture, and a polymer matrix wraps the template molecules. After the template molecules are removed from the polymer matrix, the polymer matrix leaves a gap with the imprinted template molecules, and the polymer is the molecularly imprinted high molecular polymer material. The target analyte (template) molecule can bind to the complementary molecular vacancies in the molecularly imprinted polymeric material to embed vacancies, whereas competing molecules are difficult to embed vacancies due to insufficient matching of molecular size, shape, and interaction forces with the molecular vacancies (fig. 1). Traditional molecular imprinting methods use only one polymerization to form the molecularly imprinted vacancies. The molecularly imprinted polymer material has characteristic template cognition vacancy and is prepared by imprinting a templateThe monomer of the molecule is polymerized and then the template molecule is removed. The monomer interacts with the template molecule through covalent bonds or non-covalent bonds, mainly non-covalent bonds. After the monomer is polymerized, the template molecule is wrapped in the high polymer material matrix. Next, the template molecules are removed from the matrix of the high molecular weight polymeric material leaving imprinted vacancies. Molecular recognition is achieved by molecular binding vacancies of the same size, shape and function as the template molecule. There are many patents describing conventional molecularly imprinted polymeric materials. In U.S. Pat. No. 20150342869, silane, tetraalkyl (C)₁-C₄) Orthosilicate, pore-forming solvent and C₁₄-C₂₀The fatty acid is polymerized to form the molecular engram high molecular polymer material. If the cosmetic formulation incorporates such a molecularly imprinted polymeric material, it is said to have an effect of preventing and caring for dandruff. There are some patents that describe dual polymerization, but are not related to molecular imprinting.

Disclosure of Invention

In order to solve the defects and problems of the molecularly imprinted high molecular polymer in the prior art, a preparation method of a molecularly imprinted high molecular polymer material formed based on two or more times of polymerization is provided.

The technical scheme adopted by the invention for solving the technical problem is as follows: the invention relates to a preparation method of a molecularly imprinted polymer material formed on the basis of two or more times of polymerization, which comprises the following steps:

(1) dissolving monomer, initiator, cross-linking agent and template molecule with water or organic solvent, and mixing;

(2) carrying out primary polymerization under a first reaction condition, and firstly imprinting a certain part of a template molecule; (3) then carrying out second polymerization under a second reaction condition and then imprinting other parts of the template molecules;

(4) if necessary, polymerizing the rest parts of the imprinted template molecule for more times;

(5) and finally, removing the template, the unpolymerized or uncrosslinked monomer, the low molecular weight oligomer and other impurities to obtain the molecularly imprinted polymer material formed on the basis of two or more times of polymerization.

The polymerization conditions of the two or more times are different from each other.

The polymerization includes, but is not limited to, condensation, free radical, thermal, electrical, emulsion, solution, suspension, precipitation, light, plasma, reversible addition-fragmentation chain transfer, atom transfer free radical, nitroxide assisted, ring opening polymerization.

Monomers used in the polymerization may include, but are not limited to, silanes, acrylics, olefins, epoxies, bisphenol-a, amides, imines, fluorinated monomers, ionic liquids, glycerol, fatty acids, amino acids, nucleotides, monosaccharides.

The monomers used in the polymerization may have one, two or more functional moieties that can be polymerized, and may also have one, two or more functional groups that can interact with the template molecule.

The template molecule is imprinted by a first polymerization at a certain site, and is imprinted by a second polymerization at another site, and so on if there is a third or more polymerization.

In order to facilitate the entrance and exit of large template molecules, an imprinting opening can be manufactured, one end molecule of the imprinting molecule is protected through polymerization of low crosslinking or no crosslinking at a certain time, and finally, the low crosslinking or no crosslinking polymer at the time is removed to leave a large imprinting opening so as to facilitate the entrance and exit of target molecules.

When it is desired to recognize a class of molecules rather than just one, the imprinting cavities may selectively interact with only features of the class of target molecules.

The same template molecule is imprinted by the functional monomer for two or more times, the shape of the molecular imprinting vacancy left after the template molecule is removed is precise, the monomer local functional group type and the space position of the acting force between local molecules are caused to be immobilized, the template molecule has characteristic selective adsorption, the molecular imprinting vacancy can be provided with an opening for the target molecule to enter, and the imprinting vacancy can interact with all parts of the target molecule or selectively interacts with only the characteristic parts.

The invention has the beneficial effects that: compared with the prior art, the preparation method of the molecularly imprinted polymer material based on two or more times of polymerization has the advantages that the same template molecule is imprinted by the functional monomer for two or more times in sequence, the shape of the molecular imprinting vacancy left after the template molecule is removed can be precise, the type and the space position of the monomer local functional group causing the local intermolecular force are fixed, and the purposes of higher selective adsorption, better sensitivity and better anti-interference performance on the template molecule are achieved. The traditional imprinting method only uses one-time polymerization to form a molecular imprinting vacancy, a large biomolecule generally has a plurality of functional groups, a plurality of subunits, different sizes and shapes, different spatial configurations when the pH quality changes, and molecular chirality or local chirality, the simple and rough one-step polymerization imprinting technology used in the preparation of the traditional molecular imprinting high molecular material is difficult to imprint such complex biomacromolecules, and the invention can effectively solve the problem by two-step or multi-step one-step polymerization.

Drawings

The following will further describe a preparation method of a molecularly imprinted polymer material formed based on two or more polymerizations according to the present invention with reference to the accompanying drawings and the following detailed description.

FIG. 1 is a schematic flow chart of the formation of a molecularly imprinted polymeric material;

FIG. 2 is a graph demonstrating two and more polymerizations of silane monomers and acrylic acid monomers and template concanavalia protein molecules;

FIG. 3 is a schematic representation of the design of two and more polymerizations to produce a molecularly imprinted polymeric material-based chemical sensor that can achieve precisely defined and controlled molecularly imprinted vacancies by step-wise imprinted polymerization of template molecules;

fig. 4 is a graph illustrating small and large particles in a mixed solution of silane monomers and template concanavalin a protein molecules.

The specific implementation mode is as follows:

as shown in fig. 1 to 4, the preparation method of the molecularly imprinted polymer material formed by two or more polymerizations according to the present invention is demonstrated by the molecularly imprinted polymer material prepared by two polymerizations of silane monomers and acrylic acid monomers using sword bean a protein molecules as model template molecules. The monomer used in the present invention may be formed by mixing a plurality of monomers, and is not limited to the two-stage polymerization of silane and acrylic acid. The silane monomers and acrylic acid monomers and template sword bean A protein molecules for demonstration are shown in figure 2 and comprise the following components:

aminopropyltrimethoxysilane or aminopropyltriethoxysilane (APTMS or APTES): silane monomers having only one amino functional group.

N- (2-aminoethyl) -3-aminopropyltrimethoxysilane (AEAPTMS): silane monomers having two amino functional groups.

Methacrylate oxy-propyltrimethoxysilane (MAPTMS): the bifunctional silane and acrylic monomers are both silane monomers and acrylic monomers.

Tetraethoxysilane (TEOS): used as a crosslinking agent for the polymerization of silane monomers, does not contribute to the polymerization of acrylic acid.

Azobisisobutyronitrile (AIBN): used as a crosslinking agent for the polymerization of acrylic monomers, does not contribute to the polymerization of silanes.

Jack bean a protein molecule (Con a): a model template molecule, a lectin, having a molecular weight of 10.4-11.2 kilodaltons.

FIG. 3 illustrates two and more polymerization blots, but the monomers are not limited to the example illustrated in FIG. 2 and the polymerization method is not limited to the example illustrated in FIG. 3. Aminopropyltriethoxysilane (APTES) in FIG. 2 is a silane monomer with amino functional groups, whose amino groups readily interact with the template molecule by hydrogen bonding. Methacrylate oxy-propyltrimethoxysilane (MAPTMS) is a bifunctional silane and acrylic monomer, both silane and acrylic. Its ester group can perform hydrogen bond interaction with the template molecule. For Tetraethoxysilane (TEOS)The crosslinking agent for silane monomer polymerization is prepared. First, aminopropyltriethoxysilane, methacryloxy-propyltrimethoxysilane, and tetraethoxysilane were adsorbed on template concanavalin A protein lectin molecules in a buffer solution of pH 8.3. The silane is then gradually hydrolyzed to yield a silanol group, which is further partially polymerized (condensation reaction) during the aging of the solution. The solution is then coated on a substrate, such as a gold-plated quartz wafer. During the drying and aging process at room temperature, the silanol groups are further polymerized and crosslinked to form- (O-Si-O-) with a certain crosslinking degree on the surface of the gold_nA polysilane. The film was very strongly bonded to the gold surface. The tetraethoxysilane crosslinking agent can effectively control the primary silane polymerization. If the tetraethoxysilane is in excess, the degree of crosslinking is too high, resulting in too tight a binding of the template molecules by the highly crosslinked matrix, and subsequent template removal processes tend to result in the collapse, deformation and collapse of the molecular imprinting vacancies, which reduces sensitivity and selectivity for the target template analyte. If the amount of tetraethoxysilane is insufficient and the degree of crosslinking is insufficient to fix the position of the template molecule to be imprinted, the template molecule may move in a subsequent step, resulting in incomplete imprinting of the size shape and spatial configuration of the imprinted vacancy to the template molecule, and the sensitivity and selectivity for the target template analyte may be reduced. Through the initial silane monomer polymerization, partial sites of the sword bean A protein molecule have hydrogen bonding action with amino groups and ester groups in a polymer matrix, the spatial positions of the sites are fixed, and the whole protein molecule is substantially fixed. After drying at room temperature and aging, the dry film is heated to initiate a second polymerization, i.e., polymerization of the methacrylate. This is a radical polymerization of methacrylate monomers initiated by azo or peroxide initiators under the action of ultraviolet light or heat. More kinds or amounts of acrylic monomers and crosslinking agents may be used for the acrylic polymerization, facilitating a second polymerization with a suitable degree of polymerization and crosslinking. In this exemplary reaction, heating to 90 degrees Celsius for 2 hours is sufficient to polymerize the acrylic monomer. The second polymerization is carried out with a suitable degree of polymerization and crosslinking such that the polymeric matrix further encapsulates the template molecule. Second polymerizationThe ester group position in the polymeric matrix is further fixed, so that the template molecule which has interaction with the ester group is further fixed, and the whole template molecule is more tightly wrapped by the matrix. If necessary, more monomers and different initiators can be added into the demonstration system to realize multiple times of polymerization so as to achieve more precise control of the size and the geometry of the molecular imprinting vacancy, the types of the peripheral matrix functional groups and the density and the spatial configuration. Finally, the template molecules are removed to create precisely defined and controlled molecularly imprinted vacancies. The method disclosed by the invention has the advantages that rough one-step polymerization and fine multistep polymerization are carried out, and the molecular imprinting vacancies can be better regulated and controlled to be more matched with template molecules, so that high selectivity and high sensitivity on target template analytes are achieved.

How to regulate the molecularly imprinted polymer material by the preparation method.

After each polymerization reaction, the imprinted vacancies can be tested to see if the template molecule binds to the generated imprinted vacancies to determine if each step of molecularly imprinted polymerization is just, insufficient, or over-headed. Target template analyte molecules interact with corresponding functional groups on the imprinted vacancies and are adsorbed on the vacancies, and the size, geometry, types, density and spatial configuration of the molecularly imprinted vacancies affect the adsorption firmness and the adsorbed molecular species. The molecular imprinting vacancies then influence the number of adsorbed molecules. For example, a quartz microbalance can be used for the evaluation. A sufficiently large change in the resonance frequency of the quartz indicates a sufficiently high sensitivity. The quartz resonance frequency changes only for target template analyte molecules and does not change for interfering molecules, indicating good selectivity. Whether the molecular imprinting polymerization is insufficient or too late is difficult to judge, and the judgment needs to be carried out by combining the adsorption condition of interfering molecules. When the target template analyte molecule adsorption amount is low, if the interfering molecule adsorption amount is large, the imprinting polymerization should be insufficient, otherwise, the imprinting polymerization should be over-terminated. If the imprinted polymerization is insufficient, polymerization may be carried out in additional steps until imprinted polymerization is sufficient. If the imprinted polymerization is too late, the previous polymerization, including the type and amount of the cross-linking agent and the corresponding monomer, needs to be adjusted. In this way, molecular imprinting can be well controlled to achieve the desired sensitivity and selectivity.

A large biomolecule typically has many functional groups, several subunits, different sizes and shapes, different spatial configurations when pH properties change, and molecular chirality or local chirality. The simple and rough one-step polymerization imprinting technology used in the preparation of the traditional molecular imprinting high molecular polymer material is difficult to imprint such complex biological macromolecules. The present invention can effectively solve this problem by polymerizing one-step polymerization in two or more steps. Complex features of biological macromolecules are precisely imprinted one by one in a two-step or multi-step polymeric blot. The molecularly imprinted polymer material formed based on two or more times of polymerization has the following important characteristics through the special preparation method:

1) the molecularly imprinted void can have an opening

A large template biomolecule, usually several nanometers in size, can be difficult to remove if it is completely encapsulated by the polymeric matrix. The large template molecule can only be degraded into small fragments by acid, alkali, oxidant and biological enzyme, and then the small fragments are removed by various methods such as solvent extraction, surfactant emulsification or electrophoresis, etc., leaving imprinting vacancies. However, even if the large template biomolecules are degraded to clear the remaining molecularly imprinted vacancies, it is difficult for the same large biomolecules as the target template analyte to reach the closed imprinted vacancies through layer-by-layer obstacles. Surface imprinting is a solution, but sensitivity is affected because the two-dimensional surface volume is much smaller than three-dimensional space. The invention can selectively print only one end with relative characteristics of macromolecules, and the other end is firstly protected by low cross-linked or uncross-linked polymer, and then the low cross-linked or uncross-linked polymer is dissolved, and the protected end can be made into a print opening channel.

2) The imprinting empty spaces can have dimension space surplus

A biomacromolecule is generally not strictly spherical. If the imprinting gap is the same size as the biological macromolecule, the target analyte biological macromolecule can enter the gap smoothly only by adjusting the angle to a certain degree, and the secondary angle is very small. If the imprinting void energy is slightly larger than the molecule, the angle of entry will be larger, facilitating the capture of the target analyte molecule by the imprinting void. The invention can control the surplus of some dimension space left in the imprinting vacancy by the amount of the imprinting functional groups and the number of times of polymerization imprinting, increases the angle of target analyte molecules entering the imprinting vacancy, and improves the sensitivity.

3) The imprinted vacancies can selectively interact with target analyte molecular features

A macromolecule typically has many functional groups thereon. The macromolecules of the same class are generally characterized by some functional groups or fragments. It is sometimes desirable to imprint a class of molecules rather than just one molecule. The benefit of imprinting a class of molecules is that the imprinted polymeric material can interact with both of these classes of molecules. When used as a chemical sensor, a class of molecules can be detected. The invention can selectively polymerize and engrave the characteristic functional groups of a molecule, and is realized by two or more times of polymerization.

Polymerization as used herein includes, but is not limited to, condensation (e.g., silane), free radical, thermal, electrical, emulsion, solution, suspension, precipitation, light, plasma, reversible addition-fragmentation chain transfer, atom transfer free radical, nitroxide assisted, ring opening polymerization, and the like.

The two-time or multiple-time polymerization imprinted high molecular polymer material can also be imprinted with two or more template molecules simultaneously. The first template molecule may be imprinted by primary polymerization, the second template molecule may be imprinted by secondary polymerization, and so on. Alternatively, a first polymerization may imprint certain functional groups of multiple template molecules, then a second polymerization may imprint other functional groups of multiple template molecules, and so on.

The two-time or multiple-time polymerization imprinted high molecular polymer material can be prepared by surface imprinting. The template molecule is firstly adsorbed on the surface of the particle or the substrate, then two or more times of polymerization imprinting are carried out, and finally the polymer is stripped from the surface of the particle or the substrate. The stripping process may be used to elute or dissolve the adsorption surface.

The preparation process of the two-time or multi-time polymerization imprinting high molecular polymer material is more complex and time-consuming than that of the traditional single-step imprinting method, but the obtained technical advantages are very obvious, and the problems of low selectivity and low sensitivity of the large biological molecular imprinting polymer material are solved.

Examples of the embodiments

The following illustrates the preparation process of the two or more times of the polymeric imprinted polymer material of the present invention, using the non-imprinted polymer material as a control.

1) Premix A was 200. mu.l of aminopropyltrimethoxysilane or aminopropyltrimethoxysilane aminopropyltriethoxysilane, 100. mu.l of methacrylate oxy-propyl trimethoxysilane, 100. mu.l of tetraethoxysilane, 8 mg of azobisisobutyronitrile.

2) Premix B10.0 mg of jack bean A protein molecules were dissolved in 4.0 ml of 1-fold concentrated phosphate buffered saline or deionized water. (if a non-imprinted polymer material is prepared, the sword bean A protein molecule is not added)

3) Preparing AB solution by primary polymerization-hydrolytic condensation reaction: transfer 40. mu.l of A into B, mix well, and age for 25 minutes.

4) Premix C20. mu.l AB (supernatant) was transferred to 140. mu.l phosphate buffered saline or deionized water and mixed.

5) Continuing primary polymerization and coating and fixing the high polymer material on the gold surface: transfer 40 μ l of C to a 12 mm diameter gold plated circular quartz wafer.

6) Naturally drying at room temperature.

7) And (3) second polymerization: heat to 90 degrees celsius for 2 hours.

8) Removing the template molecules: the solution was washed 3 times with 50 ml of deionized water, 10 times the concentration of phosphate buffered saline, ethanol, and deionized water.

As shown in Table 1, the selectivity of the imprinted polymer material with a film thickness of 300 nm is slightly inferior to that of the imprinted polymer material with a film thickness of 150 nm. When the silane monomer is added into the sword bean A protein molecular water solution and stirred evenly, the siliconThe methoxy or ethoxy groups of the alkane monomer begin to hydrolyze to produce Si-OH silanol groups, which in turn react with each other to partially condense to form Si-O-Si siloxane bonds. As the reaction time becomes longer, the condensation reaction produces more Si-O-Si siloxy bonds, which in turn forms polysilane particles. As the reaction time increased, the particles grew progressively larger and began to sink due to gravity without stirring. Small particles sink more slowly and large particles sink more quickly. The lower layer of the slow reaction solution showed more polysilane macro-particles, while the upper layer of the slow reaction solution mostly retained polysilane micro-particles (FIG. 4). The small particles have a relatively large specific surface, which is beneficial to improving the sensitivity. Therefore, it is preferable to use the upper end solution. As shown in Table 1, sample #270918A was prepared from the upper solution and had the highest target molecule-to-interfering molecule frequency variation ratio Df₁/Df₂(1/9.8) ratio of change in target molecule interfering molecule frequency Df to that of sample #270918B prepared from the intermediate layer solution₁/Df₂(1-Canavalia A protein molecule, 2-bovine serum protein molecule) 1/10 was slightly higher. While sample #270918C, prepared from the bottom solution, did not adsorb the target molecule, the sword bean A protein molecule at all, but adsorbed only the interfering molecule, bovine serum albumin. The frequency change was measured by a quartz microbalance.

TABLE 1

As shown in table 2, the low concentration of the mixed solution of silane and concanavalin a protein molecules (# 170817a and # 170817D) facilitates the formation of small polysilane particles, and thus can improve selectivity. The results of the experiment in Table 2 demonstrate that the second polymerization, methacrylic acid polymerization, further improves selectivity. When the mixed solution of silane and the concanavalin A protein molecules is diluted by 20 times (# 170817A), the selectivity of the imprinted polymer material to the target molecule concanavalin A protein molecule is improved by 10.03 times, and the frequency change ratio Df of the target molecule interfering molecules is₁/Df₂Increasing from 1/5.9 (# 131216A) to 1.7/1 (# 170817A). Samples #170817A and #131216A were all polymerized only once (silane)Polymerized) imprinted polymeric materials. Sample #170817D is a two-shot polymer imprinted polymeric material, the first shot was a silane polymerization and the second shot was an acrylic polymerization. Target molecule interfering molecule frequency variation ratio Df of sample #170817D₁/Df₂Further increased to 4.1/1, which is 2.41 times higher than sample #170817A from a single polymerization run. That is, the second polymerization, acrylic polymerization, further increases the selectivity to the target molecule.

TABLE 2

Alternatively, the first polymerization-silane polymerization can be performed first in solution. As shown in table 3, sample #021017E silane polymerization was completed in solution under ammonia catalysis, and the particles then adhered to the gold surface by van der waals forces. The target molecule interferes with the frequency change ratio Df of the molecule₁/Df₂Is 1/2.4 (repeat data points 1/2.7) to Df for sample #131216A, which was only partially polymerized in solution₁/Df₂The 1/5.9 is better by a factor of 2.46. The aging time of sample #021017A3 was much longer than the aging time of sample #131216A, 20 minutes, resulting in a Df₁/Df₂1/7.5 (repeat data points 1/6.9) is greater than the Df of sample #131216A₁/Df₂1/5.9 is smaller and the selectivity is worse. If the aging time is too long (e.g., 90 minutes), the upper solution may contain a large portion of small particles which sink or grow larger, and may contain too few small particles. This also demonstrates the necessity to select a supernatant containing a high number of small particles. The aging time is well controlled.

TABLE 3

As shown in Table 4, it was also successful to prepare imprinted polymer materials of two or more times polymeric imprinted polymer materials using non-aqueous solution systems. The aging time of the non-aqueous solution system is also a key parameter and needs to be strictly controlledNot to be too long. 25 minutes is sufficient. Samples #091017A and #091017B aged 5 and 24 hours, respectively, and their Df₁/Df₂Df for sample #131216A aged 1/10.6 and 1/15.1, respectively, much less than 25 minutes₁/Df₂ 1/5.9。

TABLE 4

As shown in Table 4, sample #091017C uses a 1% Sodium Dodecyl Sulfate (SDS) reagent to remove template protein molecules, which is used in many documents and patents. It is undeniably very effective for this agent to eliminate protein molecules. However, the experiments of the present invention demonstrate that this reagent is very likely to leave sequelae by removing template protein molecules, possibly disrupting some of the imprinted vacancies, resulting in a more than half-reduced sensitivity to the target template analyte molecules.

As shown in Table 5, the aging time of the aqueous solutions for preparing sample #170817D and sample #180917A was 25 minutes and 24 hours, respectively, and the target molecule interfering molecule frequency change ratio Df thereof₁/Df₂The selectivity of the adsorption on the target template molecules is greatly reduced by 23.78 times when the adsorption is reduced from 4.1/1 to 1/5.8. Moreover, the sensitivity is much reduced. Neither sample used ammonia as a catalyst for silane polymerization and the particles would grow or aggregate into larger particles with reaction time. Therefore, the polymerization aging time of the aqueous system is also an important parameter and is strictly controlled.

TABLE 5

The present invention has been described in accordance with embodiments thereof with the understanding that the present invention is not limited thereto but rather it is to be understood that variations and/or modifications may be made by those skilled in the art without departing from the scope of the invention as defined by the appended claims, and any such modifications, equivalents and the like as fall within the true spirit and scope of the invention.

Claims

1. A method for preparing a molecularly imprinted polymeric material based on two or more polymerizations, the method comprising the steps of:

(1) dissolving a monomer, an initiator, a cross-linking agent and a template molecule together or step by using water or an organic solvent, and then uniformly mixing, wherein the monomer is one or more of the following monomers: silane, acrylic, olefin, epoxy resin, bisphenol A, amide, imine, fluorinated monomer, ionic liquid, glycerol, fatty acid, amino acid, nucleotide and monosaccharide;

(2) carrying out primary polymerization under a first reaction condition, and firstly imprinting a certain part of a template molecule;

(3) then carrying out second polymerization under a second reaction condition and then imprinting other parts of the template molecules;

2. The method for preparing a molecularly imprinted polymeric material based on two or more polymerizations according to claim 1, wherein: the polymerization conditions of the two or more times are different from each other.

3. The method for preparing a molecularly imprinted polymeric material based on two or more polymerizations according to claim 1, wherein: the polymerization comprises condensation, free radical, heat, electricity, emulsion, solution, suspension, precipitation, light, plasma, reversible addition-fragmentation chain transfer, atom transfer free radical, nitrogen oxide assisted and ring opening polymerization.

4. The method for preparing a molecularly imprinted polymeric material based on two or more polymerizations according to claim 3, wherein: the monomers used in the polymerization have one, two or more functional moieties that can be polymerized and also have one, two or more functional groups that interact with the template molecule.

5. The method for preparing a molecularly imprinted polymeric material based on two or more polymerizations according to claim 1, wherein: the first part of the template molecule is polymerized and imprinted for the first time, the second part is polymerized and imprinted for the second time, and if a third part or more parts exist, the third time or more polymerization is carried out.

6. The method for preparing a molecularly imprinted polymeric material based on two or more polymerizations according to claim 1, wherein: in order to facilitate the entrance and exit of large template molecules, an imprinting opening needs to be manufactured, one end molecule of the imprinting molecule is protected through polymerization of low crosslinking or no crosslinking at a certain time, and finally, the low crosslinking or no crosslinking polymer at the time is removed to leave a large imprinting opening so as to facilitate the entrance and exit of target molecules.

7. The method for preparing a molecularly imprinted polymeric material based on two or more polymerizations according to claim 1, wherein: where it is desired to recognize a class of molecules rather than just one, the imprinting vacancies may selectively interact only with features of the class of target molecules.

8. The method for preparing a molecularly imprinted polymeric material based on two or more polymerizations according to claim 1, wherein: the same template molecule is imprinted by the functional monomer for two or more times, the shape of the molecular imprinting vacancy left after the template molecule is removed is precise, the monomer local functional group type and the space position of the acting force between local molecules are caused to be immobilized, the template molecule has characteristic selective adsorption, the molecular imprinting vacancy is provided with an opening for the target molecule to enter, and the imprinting vacancy interacts with all parts of the target molecule or selectively interacts with only the characteristic part.