An amphiphilic polymer artificial enzyme and a palladium nano-hybrid catalyst thereof, a preparation method and application thereof

Abstract

The application discloses an amphiphilic polymer artificial enzyme and a palladium nano hybrid catalyst thereof, a preparation method and application, and belongs to the technical field of polymer artificial enzyme, biomimetic catalysis, amphiphilic polymer self-assembly and metal nano hybrid catalyst. An amphiphilic statistical copolymer is prepared by copolymerization of OEGMA and 4-VBC as basic monomers (containing styrene), and a catalytic module is post-modified and grafted to the 4-VBC side chain. The polymer self-assembles into a micelle in water: OEGMA forms a hydrophilic outer layer, and aromatic hydrophobic fragments and the catalytic module are enriched in the interior. The obtained artificial enzyme can realize enzyme-mimetic organic catalysis such as water phase ester hydrolysis and hydroxy aldehyde condensation; and the matrix can also act with Pd 2+ , in-situ reduction forms a palladium nanoparticle hybrid catalyst, is used for water phase Suzuki coupling and has a recycling and reuse ability, and realizes the integration of enzyme-mimetic organic catalysis and metal nano catalysis on the same polymer platform.

Description

Technical Field

[0001] This invention belongs to the technical fields of polymeric artificial enzymes, biomimetic catalysis, amphiphilic polymer self-assembly, micelle nanoreactors, and metal nano-hybrid catalysts, specifically relating to an amphiphilic polymeric artificial enzyme and its palladium nano-hybrid catalyst, preparation method, and application. Background Technology

[0002] Natural enzymes achieve highly efficient and selective catalysis through the precise arrangement of active groups within hydrophobic substrate pockets. However, natural enzymes suffer from drawbacks such as high preparation costs, demanding storage conditions, insufficient stability, and limited applicability to specific reaction conditions. Therefore, reconstructing the synergistic catalytic microenvironment of enzymes through synthetic and tunable polymer systems is an important research direction in the field of artificial enzymes.

[0003] Existing technologies have reported catalytic triplet-like enzyme-mimicking catalysts and amphiphilic polymer micellar artificial enzymes, validating the feasibility of polymeric artificial enzymes. However, existing polymeric enzyme-mimicking platforms have significant shortcomings: catalytic site density, hydrophobic microenvironment, and aqueous self-assembled micelle morphology are mutually coupled, making independent regulation on a single platform impossible. This parameter coupling problem easily leads to polymer aggregation, mutual shielding of catalytic sites, and hindered substrate diffusion when catalytic group density increases, making it difficult to establish a composition-performance structure-activity relationship for the system. Furthermore, most existing polymeric artificial enzymes target only single organic catalytic reactions, lacking a modular construction route for introducing metal nanocatalytic functions onto the same polymer framework. This makes it difficult to form an integrated technology chain for micellar enzyme-mimicking organic catalysis, metal nanoparticle stabilization, aqueous CC coupling, and biocompatibility evaluation on a single material platform, limiting the functional expansion and practical application of artificial enzymes. Summary of the Invention

[0004] To address the shortcomings of existing technologies, the present invention aims to provide an amphiphilic polymer artificial enzyme and its palladium nano-hybrid catalyst, preparation method, and application.

[0005] To achieve the above objectives, the technical solution of the present invention is as follows.

[0006] In a first aspect, the present invention provides an amphiphilic polymeric artificial enzyme, which is a core-shell micelle structure formed by self-assembly in an aqueous solution; comprising a statistical copolymer and a catalytic module; wherein the monomers of the statistical copolymer contain polyethylene glycol methyl ether methacrylate and 4-vinylbenzyl chloride, and the catalytic module is covalently grafted onto the side chain of the 4-vinylbenzyl chloride unit of the statistical copolymer; 4-vinylbenzyl chloride units grafted with catalytic modules aggregate inward to form a micelle core, while polyethylene glycol methyl ether methacrylate segments extend outward to form a hydration layer that encapsulates the micelle core. The structure of the catalytic module is as follows: ; R1 includes any one of hydroxyl and 2-guanidinoethyl; R2 includes either 2-imidazolyl or 4-imidazolyl.

[0007] A second aspect of the present invention provides an amphiphilic polymeric artificial enzyme as described in the first aspect, comprising: By copolymerizing mixed monomers, statistical copolymers are obtained; In the statistical copolymer, the side chain benzyl chloride of the 4-vinylbenzyl chloride unit is converted into benzyl azide to obtain the benzyl azide-modified statistical copolymer; The benzyl azide-modified statistical copolymer undergoes a copper-catalyzed azide-alkyne cycloaddition reaction with a catalytic module. The catalytic module covalently grafts the generated 1,2,3-triazole ring onto the side chain of the statistical copolymer. The resulting polymer self-assembles in an aqueous solution to form a core-shell micelle structure.

[0008] A third aspect of the present invention provides the use of the amphiphilic polymeric artificial enzyme described in the first aspect in catalyzing ester hydrolysis and / or aldol reactions.

[0009] In a fourth aspect, the present invention provides a method for preparing 4-nitrophenol, wherein a p-nitrophenyl ester substrate is used as a reactant, and an ester bond hydrolysis reaction is carried out under the catalysis of a catalyst to generate 4-nitrophenol; wherein the catalyst is the amphiphilic polymeric artificial enzyme described in the first aspect.

[0010] A fifth aspect of the present invention provides the use of the amphiphilic polymeric artificial enzyme described in the first aspect in the preparation of antibacterial materials and / or cell-compatible catalytic materials.

[0011] A sixth aspect of the present invention provides a palladium nanohybrid catalyst comprising the amphiphilic polymeric artificial enzyme described in the first aspect and palladium nanoparticles anchored in a micelle matrix of the amphiphilic polymeric artificial enzyme.

[0012] A seventh aspect of the present invention provides the application of the palladium nano-hybrid catalyst described in the sixth aspect in the Suzuki coupling reaction.

[0013] An eighth aspect of the present invention provides a method for preparing biphenyl, wherein bromobenzene and phenylboronic acid are coupled via Suzuki coupling in the presence of a catalyst to obtain biphenyl. The catalyst is the palladium nano-hybrid catalyst described in the sixth aspect.

[0014] The beneficial effects of this invention are as follows: This invention provides an amphiphilic polymeric artificial enzyme, constructed using polyethylene glycol methyl ether methacrylate (PEG) and 4-vinylbenzyl chloride as basic monomers to build a statistical copolymer backbone, with catalytic modules covalently grafted onto the side chains of the 4-vinylbenzyl chloride units. The resulting polymer can form micellar nanoaggregates in an aqueous phase, wherein PEG segments tend to form a hydrated hydrophilic outer layer, while aromatic hydrophobic segments and their grafted catalytic modules tend to be distributed in the relatively hydrophobic inner region. The nominal content of graftable sites can be adjusted by changing the molar ratio of 4-vinylbenzyl chloride; in a series of samples with a constant 4-vinylbenzyl chloride feed ratio, the hydrophobic composition of the copolymer can be adjusted by changing the feed ratio of styrene to PEG. Within the tested composition range, representative samples maintained aqueous dispersion and micellar assembly capabilities, facilitating the examination of the correlation between the nominal content of the catalytic module, the hydrophobic composition, and the catalyst concentration and catalytic performance. Experimental results show that the system exhibits a non-monotonic composition-performance relationship. When the nominal content of the catalytic module or the concentration of the catalyst used is high, the apparent catalytic efficiency may decrease due to site shielding, reduced substrate accessibility, or limited mass transfer. In some experimental systems, appropriately reducing the concentration of the catalyst used can improve the apparent catalytic efficiency parameter.

[0015] The amphiphilic polymeric artificial enzyme provided by this invention contains functional groups such as hydroxyl or guanidinium, carboxyl, and imidazole groups, and can catalyze model reactions such as ester hydrolysis and aldol condensation, achieving aqueous-phase enzyme-like organic catalysis. Furthermore, using P-10%-1 and P-10%-3 artificial enzymes as polymer matrices, the invention utilizes carboxyl or carboxylate salts and nitrogen-containing groups with Pd... 2+ The interaction between the enzymes was investigated, and the artificial enzyme-PdNPs nano-hybrid catalyst was prepared by in-situ reduction with ascorbic acid. TEM results showed that the Pd nanoparticles were dispersed and stable in the polymer matrix, EDX confirmed the presence of Pd, and FTIR results supported the interaction between the carboxylate and the Pd species. The obtained nano-hybrid catalyst could catalyze the Suzuki CC coupling reaction in aqueous phase and could be continuously recycled for five rounds of reaction, but the yield decreased with increasing cycle number.

[0016] Based on the same type of amphiphilic polymer, this invention realizes functions such as aqueous enzyme-like organic catalysis, palladium nanoparticle stabilization, and aqueous Suzuki coupling catalysis, providing a material basis for the study of the composition-performance relationship of polymer artificial enzymes and the expansion of their metal nanoparticle hybrid catalytic functions. Attached Figure Description

[0017] The accompanying drawings, which form part of this invention, are used to provide a further understanding of the invention. The illustrative embodiments of the invention and their descriptions are used to explain the invention and do not constitute an improper limitation of the invention.

[0018] Figure 1 This invention presents the two-step synthesis route of catalytic module 1 and the structures of catalytic modules 2, 3 and 4.

[0019] Figure 2 The present invention provides a reversible addition-fracture chain transfer (RAFT) copolymerization, benzyl chloride azidation, and copper-catalyzed azido-acetylene cycloaddition (CuAAC) grafting route for polyethylene glycol methyl ether methacrylate / styrene / 4-vinylbenzyl chloride in amphiphilic polymer artificial enzymes.

[0020] Figure 3 This is a schematic diagram of the self-assembly of an amphiphilic polymeric artificial enzyme in water to form a micelle nanoreactor, as provided by the present invention.

[0021] Figure 4 TEM characterization of the amphiphilic polymeric artificial enzymes P-5%-1, P-5%-3, P-20%-1, P-20%-3, P-25%50%-1, and P-25%50%-3. The aqueous solution concentration of the polymers for TEM characterization was 0.5 mg / mL. Among them, A is P-5%-1 (4-VBC feed 5 mol%, catalytic module 1), B is P-5%-3 (4-VBC feed 5 mol%, catalytic module 3), C is P-20%-1 (4-VBC feed 20 mol%, catalytic module 1), D is P-20%-3 (4-VBC feed 20 mol%, catalytic module 3), E is P-25%50%-1 (OEGMA / St / 4-VBC=50 / 25 / 25, catalytic module 1), and F is P-25%50%-3 (OEGMA / St / 4-VBC=50 / 25 / 25, catalytic module 3).

[0022] Figure 5 Transmission electron microscopy (TEM, a) and energy-dispersive X-ray spectroscopy (EDX, b) characterize the P-10%-3-PdNPs palladium nanohybrid catalyst.

[0023] Figure 6 FTIR spectra of P-10%-3 and P-10%-3-PdNPs (A), P-10%-1 and P-10%-1-PdNPs (B). Detailed Implementation

[0024] Given that in existing polymeric artificial enzyme platforms, the density of catalytic sites, hydrophobic microenvironment, and the morphology of aqueous self-assembled micelles are coupled with each other and cannot be independently regulated, resulting in easy aggregation and shielding of catalytic sites, difficulty in establishing the composition-performance structure-activity relationship of the system, and limitation on modular expansion of multiple functions such as metal nanocatalysis on the same framework, this invention proposes an amphiphilic polymeric artificial enzyme and its palladium nano-hybrid catalyst, preparation method, and application.

[0025] A typical embodiment of the present invention provides an amphiphilic polymeric artificial enzyme, which is a core-shell micelle structure formed by self-assembly in aqueous solution; it includes a statistical copolymer and a catalytic module; the monomers of the statistical copolymer contain polyethylene glycol methyl ether methacrylate (OEGMA) and 4-vinylbenzyl chloride (4-VBC), and the catalytic module is covalently grafted onto the side chain of the 4-vinylbenzyl chloride unit of the statistical copolymer; 4-vinylbenzyl chloride units grafted with catalytic modules aggregate inward to form a micelle core, while polyethylene glycol methyl ether methacrylate segments extend outward to form a hydration layer that encapsulates the micelle core. The structure of the catalytic module is as follows: ; R1 includes any one of hydroxyl and 2-guanidinoethyl; R2 includes either 2-imidazolyl or 4-imidazolyl.

[0026] This invention constructs an amphiphilic statistical copolymer that can be directly dispersed in water. The copolymer uses polyethylene glycol methyl ether methacrylate (OEGMA) as the hydrophilic monomer and 4-vinylbenzyl chloride (4-VBC) and optionally added styrene as hydrophobic monomers. Based on the amphiphilic composition of the polymer, its aqueous dispersion behavior, and TEM observations, the tested representative polymers can form micellar nanoaggregates in the aqueous phase without the addition of an additional cosolvent. Due to its amphiphilic molecular structure, OEGMA segments tend to form a hydrated hydrophilic outer layer, while 4-VBC-derived units, styrene fragments, and their grafted catalytic modules tend to be distributed in the relatively hydrophobic inner region, thus providing a suitable microenvironment for the enrichment of hydrophobic substrates and aqueous enzyme-mimicking catalysis.

[0027] This invention utilizes 4-VBC units as post-modification sites for polymers. First, a statistical copolymer backbone is obtained through copolymerization. Then, catalytic modules containing hydroxyl or guanidine, carboxyl, and imidazole groups are covalently grafted onto the side chains of the 4-VBC-derived units. By changing the molar ratio of 4-VBCs during the copolymerization stage, the number of available modification sites and the nominal content of the catalytic modules can be adjusted. In a series of samples with a constant 4-VBC feed ratio, the overall hydrophobic composition of the copolymer can be adjusted by changing the styrene to OEGMA feed ratio, and the effects of changes in hydrophobic composition on micellar aggregates and catalytic performance can be compared. Within the tested composition range, representative samples all maintained aqueous phase dispersion and the formation of micellar aggregates.

[0028] This invention prepared a series of samples with different 4-VBC feed ratios and different styrene / OEGMA feed ratios. The correlation between polymer composition, nominal catalytic module content, catalyst concentration, and aggregate morphology and catalytic kinetic parameters was investigated through TEM observation and model catalytic reactions. Experimental results show that the catalytic performance of this amphiphilic polymeric artificial enzyme exhibits a non-monotonic trend with changes in the nominal catalytic module content, the hydrophobic composition of the copolymer, and the catalyst concentration. When the nominal catalytic module content or catalyst concentration is high, the apparent catalytic efficiency may decrease due to mutual shielding of catalytic sites, reduced substrate accessibility, or limited mass transfer. In some experimental systems, appropriately reducing the catalyst concentration can improve the apparent catalytic efficiency parameters, providing experimental basis for the composition design and optimization of usage conditions of this type of polymeric artificial enzyme.

[0029] In this invention, to balance monomer copolymerization activity, segment hydrophilicity, and micelle assembly behavior, the hydrophilic monomer is preferably OEGMA, and more preferably OEGMA 500. It should be noted that this invention is not limited to a specific molecular weight or this single monomer. In alternative embodiments, polyethylene glycol methyl ether methacrylates of different molecular weights (e.g., OEGMA with a molecular weight Mn of 400-600) can be used, as well as polyethylene glycol (meth)acrylates, or other (meth)acrylate monomers with water-soluble side chains. In this invention, the common function of the hydrophilic monomers is to provide water-soluble side chains for the statistical copolymer. When the polymer self-assembles in the aqueous phase, the segments of the hydrophilic monomers extend outward to form a hydrated hydrophilic crown layer of the micelles, ensuring the dispersion stability of the overall micelle nanoreactor in the aqueous phase. As long as the above-mentioned hydrophilic crown layer construction function can be achieved, it can be used for the construction of the amphiphilic statistical copolymer framework of this invention.

[0030] In this invention, to balance copolymerization activity, ease of post-modification reaction, and micellar assembly performance, the hydrophobic monomer is preferably 4-vinylbenzyl chloride (4-VBC), but it is not limited to this hydrophobic monomer. It should be noted that any hydrophobic monomer that can provide hydrophobic structural units and that can be post-modified monomers that can achieve covalent immobilization of the catalytic module can be used to construct the amphiphilic statistical copolymer backbone described in this invention. In alternative embodiments, other styrene monomers or aromatic (meth)acrylate monomers can also be selected as the hydrophobic monomer. These monomers contain aromatic hydrophobic backbones and, after copolymerization, can provide hydrophobic structural segments for the copolymer, participating in the formation of the micellar hydrophobic catalytic core during self-assembly and creating an enzyme-mimicking hydrophobic substrate pocket microenvironment. The vinyl monomers that can be modified are not limited to 4-vinylbenzyl chloride; vinyl monomers containing halomethyl, azide, alkynyl, epoxy, or active ester groups can also be selected. These monomers have active sites that can undergo covalent chemical reactions. After being introduced into the polymer chain through copolymerization, biomimetic catalytic modules such as hydroxyl, guanidine, and imidazole groups can be covalently linked to the polymer side chain through chemical grafting, thereby realizing the modular introduction of catalytic groups into the micelle hydrophobic water area.

[0031] In some embodiments of this implementation, the monomer of the statistical copolymer also contains styrene, hydrophobic aromatic fragments composed of styrene units and 4-vinylbenzyl chloride units, and catalytic modules that aggregate inward to form a micelle core.

[0032] Styrene belongs to the styrene class of hydrophobic monomers. The styrene units obtained after copolymerization possess an aromatic hydrophobic framework. This framework, together with the benzylbenzene ring framework of the 4-vinylbenzyl chloride unit, constitutes the hydrophobic aromatic segment of the system. Even if all the side chains of the 4-vinylbenzyl chloride unit are grafted with polar catalytic modules, its benzyl ring framework still retains its hydrophobic effect. During aqueous self-assembly, the hydrophobic aromatic segment formed by the styrene unit and the 4-vinylbenzyl chloride unit, along with the catalytic module grafted onto it, aggregates inwards through the hydrophobic interactions of the aromatic structures, jointly constructing the hydrophobic catalytic core of the micelles, mimicking the hydrophobic substrate pocket microenvironment of natural enzymes.

[0033] The introduction of styrene allows for independent regulation of the hydrophobicity of the micelle core. This enables the control of the hydrophobic microenvironment without altering the hydrophilic segments or the content of 4-vinylbenzyl chloride units (i.e., without changing the total number of grafting sites in the catalytic module). This decouples the control of catalytic site density and the hydrophobic microenvironment, which is beneficial for optimizing micelle assembly stability and substrate enrichment within the catalytic core. This invention does not limit the addition of styrene; styrene is an optional hydrophobic monomer. When styrene is not added, the micelle core assembly can still be driven solely by the hydrophobic benzene ring framework of the 4-vinylbenzyl chloride units.

[0034] In some embodiments of this implementation, the molar percentage of polyethylene glycol methyl ether methacrylate in the monomers of the copolymer is 30-95 mol%, specifically 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95 mol%, or any two of these values; the molar percentage of 4-vinylbenzyl chloride is 5-25 mol%, specifically 5, 10, 15, 20, 25 mol%, or any two of these values; and the molar percentage of styrene is 0-45 mol%, specifically 0, 5, 10, 15, 20, 25, 30, 35, 40, 45 mol%, or any two of these values. In this invention, 30-95 mol%, 5-25 mol%, and 0-45 mol% refer to the molar percentage of raw materials added during the copolymerization stage. The above molar ratio range takes into account the hydrophilic-hydrophobic balance of the copolymer, the number of post-modifiable sites, and the micelle assembly behavior, enabling the polymer to self-assemble in the aqueous phase to obtain a core-shell micelle nanoreactor with a hydrophilic canopy-hydrophobic catalytic core.

[0035] It should be understood that styrene is an optional hydrophobic monomer, with a molar ratio of 0-45 mol%. 0 mol% represents a system without styrene, where micelle assembly can be achieved solely through the benzene ring skeleton of the 4-vinylbenzyl chloride unit. When styrene is introduced, the styrene unit, with its aromatic hydrophobic skeleton, together with the benzene ring of the 4-vinylbenzyl chloride unit, forms a hydrophobic aromatic segment. By keeping the amount of 4-vinylbenzyl chloride constant, i.e., the catalytic site density essentially unchanged, the hydrophobic microenvironment of the micelle core can be independently controlled by changing the molar percentage of styrene. This decoupling of the catalytic site density and the hydrophobic microenvironment facilitates the control of micelle assembly stability and the enrichment effect of hydrophobic substrates within the catalytic core.

[0036] In some embodiments of this implementation, the monomers of the statistical copolymer contain 80-95 mol% polyethylene glycol methyl ether methacrylate, 5-20 mol% 4-vinylbenzyl chloride, and no styrene.

[0037] In some embodiments of this implementation, the monomers of the statistical copolymer contain 95 mol% polyethylene glycol methyl ether methacrylate, 5 mol% 4-vinylbenzyl chloride, and no styrene.

[0038] In some embodiments of this implementation, the monomers of the statistical copolymer contain 90 mol% polyethylene glycol methyl ether methacrylate, 10 mol% 4-vinylbenzyl chloride, and no styrene.

[0039] In some embodiments of this implementation, the monomers of the statistical copolymer contain 80 mol% polyethylene glycol methyl ether methacrylate, 20 mol% 4-vinylbenzyl chloride, and no styrene.

[0040] The three-component binary statistical copolymers without styrene can achieve aqueous self-assembly to form a core-shell micelle structure by relying solely on the benzylbenzene ring skeleton of the 4-vinylbenzyl chloride unit to provide hydrophobic interactions. By changing the molar ratio of 4-vinylbenzyl chloride, the number of post-modifiable sites can be directly adjusted without changing the type of hydrophobic component, thereby controlling the grafting density of subsequent catalytic modules.

[0041] In some embodiments of this implementation, the monomers of the copolymer contain polyethylene glycol methyl ether methacrylate in a molar ratio of 30-55 mol%, 4-vinylbenzyl chloride in a molar ratio of 20-25 mol%, and styrene in a molar ratio of 25-45 mol%.

[0042] In some embodiments of this implementation, the monomers of the copolymer contain 50 mol% polyethylene glycol methyl ether methacrylate, 25 mol% 4-vinylbenzyl chloride, and 25 mol% styrene.

[0043] In some embodiments of this implementation, the monomers of the copolymer contain 30 mol% polyethylene glycol methyl ether methacrylate, 25 mol% 4-vinylbenzyl chloride, and 45 mol% styrene.

[0044] Two ternary statistical copolymers of styrene were introduced. While maintaining a constant amount of 4-vinylbenzyl chloride (i.e., a constant maximum number of catalytic sites that can be grafted), the overall hydrophobicity of the system was enhanced by increasing the molar percentage of styrene. This independently modulates the hydrophobic microenvironment of the micelle core, achieving decoupling between catalytic site density and the hydrophobic microenvironment. As the styrene percentage increases, the proportion of hydrophobic aromatic fragments also increases, which is beneficial for enhancing the enrichment of hydrophobic substrates. Simultaneously, by reducing the molar percentage of polyethylene glycol methyl ether methacrylate, the overall hydrophilic-hydrophobic balance of the copolymer is maintained, ensuring stable dispersion of the micelles in the aqueous phase.

[0045] In some embodiments of this implementation, the molecular weight of the statistical copolymer ( M n The apparent GPC values ​​were 8.1-76.0 kDa (before azidation, measured using polystyrene as a standard). M n ).

[0046] Within this molecular weight range, combined with the aforementioned monomer molar ratios, the statistical copolymer exhibits excellent aqueous self-assembly behavior: hydrophilic polyethylene glycol methyl ether methacrylate segments form an outward hydration canopy, while hydrophobic aromatic fragments composed of 4-vinylbenzyl chloride units and optional styrene units aggregate inward to construct a hydrophobic catalytic core. The molecular weight dispersion of samples within this range remains between 1.27 and 2.09. Most samples exhibit a dispersion between 1.27 and 1.49, demonstrating good polymerization controllability and facilitating the acquisition of structurally uniform micelles. It should be noted that azidation is a side-chain functional group transformation reaction, ideally not altering the polymer backbone length. Significant fluctuations in molecular weight measured by gel permeation chromatography (GPC) after azidation in some samples are due to minor polymer chain breakage and degradation during post-modification, not the polymerization result of the original statistical copolymer itself. This molecular weight range balances polymer solubility and dispersion, micellar assembly behavior, and substrate diffusion and mass transfer effects, ensuring the catalytic performance of the artificial enzyme.

[0047] In some embodiments of this implementation, the catalytic module is covalently linked to the side chain of the statistical copolymer 4-vinylbenzyl chloride unit via a 1,2,3-triazole ring.

[0048] The catalytic module is covalently linked to the 4-vinylbenzyl chloride unit side chain of the statistical copolymer via a 1,2,3-triazole ring, forming a benzyl-1,2,3-triazole linker. This grafts an amino acid-type catalytic module containing carboxyl, imidazole, hydroxyl, or guanidine groups onto the polymer backbone. The triazole ring exhibits high chemical stability and is not prone to bond breaking and dissociation in an aqueous catalytic environment, ensuring the catalytic module is firmly anchored to the polymer side chain. In congested micelle cores, if the linker is too short, the catalytic group may be encapsulated and shielded by the polymer chain segments, making it difficult for the substrate to approach the active site. The benzyl-1,2,3-triazole linker achieves a stable covalent connection between the catalytic module and the polymer side chain, and provides a certain spatial interval between the polymer backbone and the catalytic group, potentially facilitating contact between the catalytic group and the substrate.

[0049] In some embodiments of this implementation, the grafting reaction of the catalytic module is qualitatively confirmed by the disappearance of the azide group characteristic peak in FTIR.

[0050] In this invention, 5 mol%-25 mol% represents the molar percentage of 4-vinylbenzyl chloride fed during the copolymerization stage. This percentage represents the theoretical maximum proportion of sites available for grafting reactions in the statistical copolymer side chains. This feeding ratio is not equivalent to the actual grafting rate of the catalytic module. This invention qualitatively proves the copper-catalyzed azido-acetylene cycloaddition grafting reaction between the azido group and the acetylene group by observing the disappearance of the characteristic peak of the azido group using Fourier transform infrared spectroscopy (FTIR). This invention does not limit the actual grafting rate of the catalytic module; those skilled in the art can adjust the grafting conversion degree of the catalytic module by adjusting reaction conditions such as reaction time, reaction temperature, and reactant feeding ratio.

[0051] In some embodiments of this implementation, the catalytic module uses an α-amino acid backbone of L-serine or L-arginine as its core, with both a propargyl group and an imidazole methyl group attached to the α-amino nitrogen, and a carboxyl group retained on the α-carbon. The amino acid side chain R is hydroxymethyl (i.e., serine type) or 3-guanidinopropyl (i.e., arginine type). The imidazole methyl group can be linked to a methylene group at the 2 or 4 position of the imidazole ring. The module ends with a propargyl alkynyl group. This terminal alkynyl group can undergo a copper-catalyzed azide-alkynyl cycloaddition (CuAAC) reaction with benzyl azide on the statistical copolymer, thereby covalently grafting the catalytic module onto the statistical copolymer side chain.

[0052] Corresponding to the above structure, the structure of the catalytic module is as follows: ; R1 is selected from hydroxyl and 2-guanidinoethyl, and R2 is selected from 2-imidazolium and 4-imidazolium, forming four typical catalytic module combinations.

[0053] Specifically, R1 is a hydroxyl group, and R2 is a 2-imidazolium group; Alternatively, R1 is a hydroxyl group and R2 is a 4-imidazolium group; Alternatively, R1 is 2-guanidinoethyl and R2 is 2-imidazolyl; Alternatively, R1 is 2-guanidinoethyl and R2 is 4-imidazolyl.

[0054] This catalytic module integrates multiple acid-base synergistic functional groups, such as carboxyl, hydroxyl, guanidinyl, and imidazole groups, onto the same amino acid backbone. After grafting, these groups are positioned within the hydrophobic catalytic core of the micelles, mimicking the multi-group synergistic microenvironment of the active site of natural enzymes to achieve enzyme-mimicking catalysis under aqueous conditions. It should be understood that this invention is not limited to the four specific catalytic modules mentioned above; the substitution positions of the amino acid side chains and imidazole rings can also be adjusted, or they can be replaced with other functional groups possessing nucleophilic and acid-base synergistic effects. This invention prioritizes the protection of the common parent core structure corresponding to the four types of catalytic modules mentioned above.

[0055] Another typical embodiment of the present invention provides a method for preparing the above-mentioned amphiphilic polymeric artificial enzyme, comprising: By copolymerizing mixed monomers, statistical copolymers are obtained; In the statistical copolymer, the side chain benzyl chloride of the 4-vinylbenzyl chloride unit is converted into benzyl azide to obtain a benzyl azide-modified statistical copolymer; The benzyl azide-modified statistical copolymer undergoes a copper-catalyzed azide-alkyne cycloaddition reaction with a catalytic module. The catalytic module covalently grafts the generated 1,2,3-triazole ring onto the side chain of the statistical copolymer. The resulting polymer self-assembles in an aqueous solution to form a core-shell micelle structure.

[0056] The preparation method provided by this invention mainly includes four processes: copolymerization, azidation modification, copper-catalyzed azid-alkyne cycloaddition grafting, and aqueous self-assembly.

[0057] In some embodiments of this implementation, the mixed monomers include polyethylene glycol methyl ether methacrylate and 4-vinylbenzyl chloride, and styrene or other copolymerizable monomers may also be selectively added; the molar ratio of each monomer in the copolymerization stage has been described above and will not be repeated here.

[0058] In some embodiments of this implementation, the copolymerization of the mixed monomers includes the following steps: OEGMA, styrene, 4-vinylbenzyl chloride, chain transfer agent and initiator were dissolved in an organic solvent and subjected to deoxygenation treatment. The reaction was carried out under a protective atmosphere at 60-70 °C to obtain a statistical copolymer.

[0059] Among them, the chain transfer agent can be RAFT chain transfer agent 2-cyano-2-propyl dithiobenzoate (CPADB), the initiator can be azobisisobutyronitrile (AIBN), and the organic solvent can be tetrahydrofuran (THF).

[0060] After the reaction was completed, the reaction system was cooled, diluted with THF, precipitated and purified three times in diethyl ether, and then thoroughly dialyzed with water to obtain the statistical copolymer.

[0061] It should be understood that this invention is not limited to the RAFT polymerization system described above. In addition to RAFT polymerization, atom transfer radical polymerization (ATRP), other types of reversible deactivating radical polymerization, or ordinary radical copolymerization can also be used to construct the statistical copolymer backbone described in this invention. RAFT polymerization combined with CPADB chain transfer agent and AIBN initiator is a well-validated preferred embodiment of this invention. This approach can better control the molecular weight and molecular weight distribution of the copolymer, resulting in a compositionally controllable amphiphilic statistical copolymer, which is more conducive to subsequent post-modification and aqueous micelle self-assembly.

[0062] In some embodiments of this implementation, the catalytic module is prepared as follows: Under a protective atmosphere, amino acid raw materials, imidazole formaldehyde, 3Å molecular sieve, and base (e.g., K2CO3) are dissolved in anhydrous methanol and a reducing amination reaction is carried out by stirring at room temperature. After cooling, a reducing agent (e.g., NaBH4) is added to continue the reaction. After quenching, concentration, washing, and filtration, an imidazole methyl-substituted catalytic module precursor is obtained. The catalytic module precursor, propargyl bromide, and base (e.g., K2CO3) are added to anhydrous ethanol and stirred at room temperature to undergo an N-propargylation reaction. The resulting N-propargylated catalytic module is obtained by filtration and concentration.

[0063] When R1 is a hydroxyl group and R2 is a 4-imidazolium group, the amino acid raw material in the preparation method is L-serine, and the imidazolium formaldehyde is imidazol-4-formaldehyde.

[0064] Wherein, when R1 is a hydroxyl group and R2 is a 2-imidazolium group, in the preparation method, the amino acid raw material is L-serine and the imidazolium formaldehyde is imidazol-2-formaldehyde.

[0065] Wherein, when R1 is 2-guanidinoethyl and R2 is 4-imidazolyl, in the preparation method, the amino acid raw material is L-arginine and the imidazolium formaldehyde is imidazol-4-formaldehyde.

[0066] Wherein, when R1 is 2-guanidinoethyl and R2 is 2-imidazolyl, in the preparation method, the amino acid raw material is L-arginine and the imidazolium formaldehyde is imidazol-2-formaldehyde.

[0067] The method for preparing amphiphilic polymeric artificial enzymes provided by this invention employs RAFT polymerization to obtain compositionally tunable amphiphilic statistical copolymers, followed by azidation and CuAAC "click chemistry" modularization to introduce a biomimetic amino acid-imidazole catalytic module. The resulting polymer can directly self-assemble in an aqueous phase to form a core-shell micelle nanoreactor without solvent replacement. This preparation process exhibits good reaction compatibility, allowing for flexible control of polymer composition, micelle microenvironment, and active site types to construct a substrate-enriched enzyme-like catalytic microenvironment. The process is simple and reproducible.

[0068] The amphiphilic polymeric artificial enzyme constructed in this invention belongs to a general polymeric catalytic platform. Without altering the core polymer backbone, its application functions can be expanded simply by changing the types of catalytic modules with side branches. Sufficient experimental data support the catalytic efficacy of this artificial enzyme for hydrolysis, model aldol reactions, and Suzuki coupling reactions. Based on the modular post-modification capability and microenvironment regulation characteristics of this polymeric platform, it can be used as a candidate catalytic platform for other organic transformations such as activated ester hydrolysis, Knoevenagel condensation, Michael addition, and Heck and Sonogashira couplings by introducing corresponding organic catalytic groups or metal active centers. This artificial enzyme can operate in an aqueous environment, improving its catalytic performance through the substrate enrichment effect of its micellar hydrophobic core, providing a technical foundation for the development of aqueous biomimetic organic catalysis and functional polymeric biomaterials.

[0069] Therefore, another typical embodiment of the present invention provides the application of the above-mentioned amphiphilic polymeric artificial enzyme in catalyzing ester hydrolysis and / or aldol reactions.

[0070] Another typical embodiment of the present invention provides a method for preparing 4-nitrophenol, which uses p-nitrophenyl ester substrates as reactants and generates 4-nitrophenol through ester bond hydrolysis under the catalysis of a catalyst. The catalyst is the aforementioned amphiphilic polymeric artificial enzyme.

[0071] The hydrolysis reaction is carried out in an aqueous system. An amphiphilic polymeric artificial enzyme self-assembles in water to form a core-shell micelle nanoreactor. The hydrophobic catalytic core of the micelles enriches hydrophobic p-nitrophenyl ester substrates, thereby efficiently catalyzing the cleavage of ester bonds to release the product 4-nitrophenol. The study found that this catalytic system exhibits a significant non-monotonic composition-performance relationship: when the concentration of catalytic sites or the overall polymer concentration is too high, the apparent catalytic efficiency of the reaction decreases; appropriately reducing the polymer concentration of the artificial enzyme (e.g., P-20%-1) can significantly improve the catalytic efficiency constant. k cat / K m This phenomenon is mainly due to the combined effects of multiple factors, such as micelle aggregation under high concentration conditions, changes in the microenvironment of active sites, and limited substrate diffusion and mass transfer. This also indicates that for this amphiphilic polymer artificial enzyme, a suitable polymer concentration needs to be matched to fully realize its catalytic potential.

[0072] Another typical embodiment of the present invention provides the application of the above-mentioned amphiphilic polymeric artificial enzyme in the preparation of antibacterial materials and / or cell-compatible catalytic materials.

[0073] In some embodiments of this implementation, the amphiphilic polymeric artificial enzyme is a core-shell micelle structure formed by self-assembly in aqueous solution; it includes a statistical copolymer and a catalytic module; the monomers of the statistical copolymer contain polyethylene glycol methyl ether methacrylate (OEGMA), 4-vinylbenzyl chloride (4-VBC) and styrene, with OEGMA:styrene:4-VBC = 30:45:25 molar percentage; the catalytic module is covalently grafted onto the side chain of the 4-vinylbenzyl chloride unit of the statistical copolymer. 4-vinylbenzyl chloride units grafted with catalytic modules aggregate inward to form a micelle core, while polyethylene glycol methyl ether methacrylate segments extend outward to form a hydration layer encapsulating the micelle core. The structure of the catalytic module is as follows: R1 is 2-guanidinoethyl, and R2 is 2-imidazolyl.

[0074] Another typical embodiment of the present invention provides a palladium nanohybrid catalyst comprising the above-described amphiphilic polymeric artificial enzyme and palladium nanoparticles anchored in the micelle matrix of the amphiphilic polymeric artificial enzyme.

[0075] This palladium nanohybrid catalyst utilizes core-shell micelles formed by the self-assembly of amphiphilic polymeric artificial enzymes in aqueous phase as a support matrix. The carboxylate groups, imidazole groups, and guanidine functional sites carried on the polymer chains, along with the hydrophobic regions within the micelles, synergistically anchor and stabilize palladium nanoparticles through coordination and adsorption, effectively inhibiting their aggregation and growth. The resulting palladium nanohybrid catalyst can be directly dispersed in an aqueous phase for catalyzing the Suzuki coupling reaction. The confined environment of the micelles not only stabilizes the palladium active centers but also enriches organic substrates through the hydrophobic core, enhancing the catalytic effect of the coupling reaction under aqueous conditions. Furthermore, this micellar-based hybrid catalyst exhibits excellent recyclability; after the reaction, the catalyst and product can be easily separated for repeated recycling, providing a recyclable nanocatalytic material for aqueous palladium-catalyzed coupling reactions.

[0076] Another typical embodiment of the present invention provides a method for preparing the above-mentioned palladium nano-hybrid catalyst, comprising the following steps: Palladium salt (e.g., PdCl2) is added to an aqueous solution of an amphiphilic polymeric artificial enzyme, and the Pd salt is stirred gently at room temperature. 2+ The Pd nanoparticles are bound to carboxyl / carboxylate sites; a reducing agent (e.g., ascorbic acid) is added and stirring is continued; unstable excess Pd particles are removed by centrifugation, and the clarified liquid is dried to obtain the palladium nanoparticle hybrid catalyst.

[0077] In aqueous solution, the amphiphilic polymeric artificial enzyme pre-assembles to form core-shell micelles; after the addition of palladium salt, Pd 2+ PdNPs can interact with carboxyl / carboxylate groups and nitrogen-containing sites in the polymer matrix. After mild in-situ reduction with ascorbic acid, the resulting PdNPs are stabilized by the surrounding polymer chains and functional groups. After centrifugation to remove unstabilized excess palladium particles, TEM showed that the PdNPs were dispersed or embedded in the polymer matrix, EDX confirmed the presence of Pd, and FTIR supported interactions between carboxylates and Pd species. This evidence demonstrates that PdNPs are stable within the polymer micelle matrix, but it does not define their unique position within the radial structure of the micelles.

[0078] This palladium nano-hybrid catalyst can efficiently catalyze the Suzuki coupling reaction in an aqueous phase. Continuous cycling experiments show that both palladium nano-hybrid catalysts can be recovered and reused for five Suzuki coupling reactions, but the yield decreases with increasing cycle number, indicating that the material has a certain degree of recyclability and reuse capability, while its cycling activity retention still has room for further optimization.

[0079] It should be understood that the amphiphilic polymeric artificial enzyme of the present invention can serve as a universal carrier for loading various metal or alloy nanoparticles. Utilizing the coordination, adsorption, and confinement stabilization effects of carboxylates, imidazoles, guanidino functional groups, and the hydrophobic microenvironment within the micelles, Pt, Au, Ag, Cu, Ni, and other nanoparticles, or alloy nanoparticles composed of the aforementioned metals, can be anchored and immobilized to construct a series of micelle-metal nanoparticle hybrid catalytic materials, expanding the scope of different types of catalytic reactions. Pt, Au, Ag, Cu, Ni, and their alloy nanoparticles are objects for further expansion and evaluation using this carrier platform. Those skilled in the art can refer to the preparation approach of palladium nanoparticle hybrid catalysts, adjusting the metal precursor and reduction conditions to achieve loading and performance studies of other metal / alloy nanoparticles.

[0080] Another typical embodiment of the present invention provides the application of the above-described palladium nano-hybrid catalyst in the Suzuki coupling reaction.

[0081] Another typical embodiment of the present invention provides a method for preparing biphenyl, wherein bromobenzene and phenylboronic acid are coupled via Suzuki coupling under the action of a catalyst to obtain biphenyl; The catalyst is the palladium nano-hybrid catalyst described above.

[0082] This invention constructs a modular amphiphilic copolymer artificial enzyme. A polymer backbone is obtained through RAFT polymerization, and a self-synthesized imidazole amino acid catalytic module is covalently grafted onto the polymer side chain via azidation and CuAAC "click chemistry." The resulting polymer can directly self-assemble in an aqueous phase to form a core-shell micelle nanoreactor without solvent replacement. Substrate enrichment is achieved through the hydrophobic core, enabling highly efficient catalysis of biomimetic reactions such as ester hydrolysis and aldol reactions. Furthermore, palladium nanoparticles can be in-situ anchored using the multifunctional group coordination adsorption of the micelle matrix to prepare a recyclable aqueous Suzuki-coupled hybrid catalyst. The polymer catalytic platform of this invention has a tunable structure. Without altering the polymer backbone, the biomimetic catalytic performance can be controlled by replacing the catalytic module. It can also be expanded to support other metal or alloy nanoparticles and has potential applications in antibacterial and biomaterial applications, providing a feasible technical solution for the development of aqueous biomimetic catalysis and functional polymer materials.

[0083] To enable those skilled in the art to better understand the technical solution of the present invention, the technical solution of the present invention will be described in detail below with reference to specific embodiments.

[0084] Example 1 This embodiment provides four catalytic modules, the structures and synthetic routes of which are as follows: Figure 1 As shown.

[0085] The specific preparation method is as follows: (1) Serine-type precursor 5: Under nitrogen atmosphere, 2.10 g (20 mmol) of L-serine, 1.92 g (20 mmol) of imidazole-4-carboxaldehyde, 2.0 g of 3 Å molecular sieve and 5.53 g (40 mmol) of K2CO3 were added to 200 mL of anhydrous methanol and stirred at 25 °C for 20 h. After cooling to 0 °C, 1.51 g (40 mmol) of NaBH4 was added and the reaction was continued for 2 h. The mixture was quenched with 1 M HCl, concentrated, washed / filtered with ethanol to obtain 3.00 g of precursor, yield 81%, white solid with melting point (mp) 201-203 °C.

[0086] 13 C NMR (100 MHz, DO) δ 171.4, 135.9, 124.1, 121.5, 63.7, 60.0, 40.0. IR (KBr) ν max 3201, 2982, 2888, 1618, 1393, 1084, 829, 624 cm -1 . MS (ESI, +) m / z208 [(M+Na)+, 100%]. HRMS (M+Na)+:C7H 11 The calculated value of N3O3Na is 208.0698, and the measured value is 208.0701.

[0087] (2) Serine-type precursor 6: Under nitrogen atmosphere, 2.10 g (20 mmol) of L-serine, 20 mmol of imidazole-2-carboxaldehyde, 2.0 g of 3 Å molecular sieve, and 5.53 g (40 mmol) of K2CO3 were added to 200 mL of anhydrous methanol and stirred at 25 °C for 20 h. After cooling to 0 °C, 1.51 g (40 mmol) of NaBH4 was added, and the reaction was continued for 2 h. The reaction was quenched with 1 M HCl, and after concentration, washing / filtration with ethanol, precursor 6 was obtained as 3.15 g, with a yield of 85%.

[0088] 1 H NMR (400 MHz, D2O) δ 7.08-7.05 (m, 2H), 4.66 (s, 1H), 3.92-3.68 (m,3H), 3.19 (t, J=5.2 Hz, 1H). 13C10 NMR (100 MHz, D2O) δ 179.9, 170.8, 148.0, 147.2, 122.9, 122.6, 65.1, 64.3, 63.7, 57.9, 57.0, 44.6 (two tautomers were detected due to sample pH). IR (KBr) ν max 3157, 3075, 2922, 2833, 1583, 1404, 1093, 1050, 985,739 cm -1 . MS (ESI, +) m / z 208 [(M+Na)+, 100%]. HRMS (M+Na)+:C7H 11 The calculated value of N3O3Na is 208.0698, and the measured value is 208.0703.

[0089] (3) Arginine-type precursor 7: Under nitrogen atmosphere, L-arginine (20 mmol), imidazole-4-carboxaldehyde (1.92 g, 20 mmol), 3 Å molecular sieve (2.0 g), and K₂CO₃ (5.53 g, 40 mmol) were added to 200 mL of anhydrous methanol and stirred at 25 °C for 20 h. After cooling to 0 °C, NaBH₄ (1.51 g, 40 mmol) was added, and the reaction was continued for 2 h. The mixture was quenched with 1 M HCl, and after concentration, washing / filtration with ethanol, 74.58 g of the precursor was obtained, with a yield of 90%.

[0090] 1 H NMR (400 MHz, D2O) δ 7.72 (s, 1H), 7.13 (s, 1H), 4.58 (s, 1H), 3.94-3.79 (m, 1H), 3.33-3.16 (m, 3H), 1.91-1.54 (m, 4H). 13 C NMR (100 MHz, D2O) δ 179.7, 177.1, 157.3(4), 157.3, 137.5, 137.0, 136.9, 133.9, 118.5, 117.9, 62.6, 56.7, 55.3, 43.7, 41.4, 41.3, 29.5, 29.2, 25.0, 24.7 (two tautomers were detected due to sample pH). IR (KBr) ν max 3332, 3155, 2971, 2883, 1733, 1652, 1629, 1467,1379, 1127, 950 cm -1MS (ESI, +) m / z 255 [(M+H) + [100%] HRMS (M+H) + :C 10 H 19 The calculated value of N6O2 is 255.1569, and the measured value is 255.1573.

[0091] (4) Arginine-type precursor 8: Under nitrogen atmosphere, L-arginine (20 mmol), imidazole-2-carboxaldehyde (1.92 g, 20 mmol), 3 Å molecular sieve (2.0 g), and K₂CO₃ (5.53 g, 40 mmol) were added to 200 mL of anhydrous methanol and stirred at 25 °C for 20 h. After cooling to 0 °C, NaBH₄ (1.51 g, 40 mmol) was added, and the reaction was continued for 2 h. The mixture was quenched with 1 M HCl, and after concentration, washing / filtration with ethanol, 84.67 g of the precursor was obtained, with a yield of 92%.

[0092] 1 H NMR (400 MHz, CD3OD) δ 6.95 (s, 2H), 3.88 (d, J=14.0 Hz, 1H), 3.71 (d, J=14.0 Hz, 1H), 3.15 (t, J=8.0 Hz, 2H), 3.05 (t, J=8.0 Hz, 1H), 1.73-1.61(m, 4H). 13 C NMR (100 MHz, CD3OD) δ 181.6, 158.7, 148.2, 122.5, 64.5, 46.0, 42.3, 31.6, 26.7. IR (KBr) ν max 3160, 2981, 1634, 1558, 1451, 1398, 1099, 983,745 cm -1 MS (ESI, +) m / z 255 [(M+H) + [100%] HRMS (M+H) + :C 10 H 19 The calculated value of N6O2 is 255.1569, and the measured value is 255.1575.

[0093] (5) Catalytic module 1: N-((1H-imidazol-4-yl)methyl)-N-(prop-2-yn-1-yl)-L-serine 1.85 g (10 mmol) of serine precursor 5, 2.14 mL of propargyl bromide solution (80 wt.% toluene solution, 24 mmol), and 4.15 g (30 mmol) of K2CO3 were added to 40 mL of anhydrous ethanol. The mixture was stirred at 25 °C for 48 h, filtered, and concentrated to obtain 2.05 g of catalytic module 1, with a yield of 92%, as a pale yellow solid with a mp value of 203-204 °C.

[0094] 1 H NMR (400 MHz, D2O) δ 8.75 (s, 1H), 7.72 (s, 1H), 4.54-4.46 (m, 2H), 4.21-4.11 (m, 2H), 3.95-3.89 (m, 3H), 3.04 (t, J=4.0 Hz, 1H). 13 C NMR (100 MHz, D2O) δ 174.0, 135.4, 126.9, 120.7, 78.6, 75.4, 68.2, 59.8, 46.1, 40.8. IR(KBr) ν max 3197, 2971, 2889, 1662, 1590, 1404, 1297, 1066, 879, 726 cm -1 MS (ESI,-) m / z 222 [(MH) - [100%] HRMS (MH) - :C 10 H 12 The calculated value of N3O3 is 222.0879, and the measured value is 222.0878.

[0095] (6) Catalytic module 2: N-((1H-imidazol-2-yl)methyl)-N-(prop-2-yn-1-yl)-L-serine Serine-type precursor 6 (10 mmol), propargyl bromide solution 2.14 mL (80 wt.% toluene solution, 24 mmol), and K2CO3 4.15 g (30 mmol) were added to 40 mL of anhydrous ethanol. The mixture was stirred at 25 °C for 48 h, filtered, and concentrated to obtain catalytic module 2 1.94 g, yield 87%, as a yellow oil.

[0096] 1H NMR (400 MHz, D2O) δ 7.39 (s, 2H), 4.42-4.33 (m, 2H), 4.03-3.94 (m,2H), 3.81 (t, J=5.7 Hz, 1H), 3.68-3.56 (m, 2H), 2.64 (t, J=4.0 Hz, 1H). 13 C NMR (100 MHz, DO) δ 175.1, 146.1, 119.4, 79.0, 75.3, 66.6, 60.6, 46.8, 42.1. IR(KBr) ν max 3287, 3151, 2894, 2242, 1714, 1662, 1432, 1184, 1132, 835, 798 cm -1 MS(ESI, +) m / z 224 [(M+H) + [100%] HRMS (M+H) + :C 10 H 14 The calculated value of N3O3 is 224.1035, and the measured value is 224.1039.

[0097] (7) Catalytic module 3: N 2 -((1H-imidazol-4-yl)methyl)-N 2 -(prop-2-yn-1-yl)-L-arginine Arginine-type precursor 7 (10 mmol), propargyl bromide solution 2.14 mL (80 wt.% toluene solution, 24 mmol), and K2CO3 4.15 g (30 mmol) were added to 40 mL of anhydrous ethanol. The mixture was stirred at 25 °C for 48 h, filtered, and concentrated to obtain catalytic module 3 2.43 g, yield 83%, white solid, mp 218-219 °C.

[0098] 1 H NMR (400 MHz, CD3OD) δ 7.61 (s, 1H), 6.92 (s, 1H), 3.81-3.70 (m,2H), 3.57 (s, 1H), 3.45-3.41 (m, 1H), 3.27-3.13 (m, 4H), 1.83-1.60 (m, 4H). 13CNMR (100 MHz, CD3OD) δ 180.7, 179.7, 158.6, 136.4, 123.4, 68.2, 67.9, 47.4, 42.3, 40.6, 39.4, 28.6, 26.9. IR (KBr) ν max 3128, 2981, 1637, 1560, 1438, 1400,1137, 1106, 829, 755 cm -1 MS (ESI, +) m / z 293 [(M+H) + [100%] HRMS (M+H) + :C 13 H 21 The calculated value of N6O2 is 293.1726, and the measured value is 293.1725.

[0099] (8) Catalytic module 4: N 2 -((1H-imidazol-2-yl)methyl)-N 2 -(prop-2-yn-1-yl)-L-arginine Arginine-type precursor 8 (10 mmol), propargyl bromide solution 2.14 mL (80 wt.% toluene solution, 24 mmol), and K2CO3 4.15 g (30 mmol) were added to 40 mL of anhydrous ethanol. The mixture was stirred at 25 °C for 48 h, filtered, and concentrated to obtain catalytic module 4 2.25 g, yield 77%, as a yellow oil.

[0100] 1 H NMR (400 MHz, D2O) δ 7.41-7.28 (m, 2H), 4.93 (s, 1H), 4.38-4.14 (m,2H), 3.62-3.33 (m, 3H), 3.21 (t, J=6.7 Hz, 2H), 1.95-1.57 (m, 4H). 13 C10 NMR (100MHz, D2O) δ 180.3, 179.3, 157.3, 146.3, 122.2, 121.3, 119.5, 118.4, 115.5, 112.6, 78.8, 75.1, 68.1, 62.8, 55.0, 46.3, 42.6, 41.69, 41.67, 41.4, 41.2, 41.1, 28.7, 27.7, 25.6, 24.7 (two tautomers were detected due to sample pH). IR (KBr) νmax 3362,3174, 2713, 1660, 1429, 1181, 1128, 835, 798 cm -1 MS (ESI, +) m / z 293 [(M+H) + [100%] HRMS (M+H) + :C 13 H 21 The calculated value of N6O2 is 293.1726, and the measured value is 293.1731.

[0101] Table 1. Raw material consumption of the polymer obtained in Example 2

[0102] Note: St in Table 1 represents styrene.

[0103] The catalytic modules 1-4 obtained in this embodiment are a more accurate collective term. Catalytic modules 1 and 2 contain hydroxyl, carboxyl, and imidazole groups, directly mimicking the Ser / Asp / His functional combination; catalytic modules 3 and 4 use guanidinium groups on the arginine side chain instead of hydroxymethyl groups, belonging to enzyme-mimicking variants that enhance hydrogen bonding / ionic interactions.

[0104] Example 2 This embodiment provides a series of methods for preparing amphiphilic statistical copolymer backbones. The raw material amounts for each group are shown in Table 1, and the preparation methods are as follows: OEGMA 500, styrene, 4-vinylbenzyl chloride, CPADB, and AIBN were dissolved in THF, deoxygenated by purging with nitrogen for 20 min, and reacted at 65 °C for 24 h under nitrogen protection. After cooling, the reaction solution was diluted with THF, precipitated three times in diethyl ether, and then thoroughly dialyzed with water.

[0105] The total monomer dosage for each group was 40 mmol, CPADB was 0.20 mmol, AIBN was 0.08 mmol, and THF was 32 mL.

[0106] Example 3 This embodiment provides a method for obtaining an amphiphilic polymeric artificial enzyme by benzyl chloride azide treatment and CuAAC grafting of an amphiphilic statistical copolymer backbone, specifically including the following steps: (1) Benzyl chloride azide Samples P-1 to P-5 obtained in Example 2 were dissolved in anhydrous DMF with NaN3 and reacted at 120 °C for 24 h. After cooling, they were precipitated three times in diethyl ether and dialyzed to obtain P-6 to P-10, respectively. The molecular weight values ​​determined by gel permeation chromatography (GPC) were the apparent molecular weights obtained using polystyrene standards and N,N-dimethylacetamide (DMAc) as the mobile phase.

[0107] Table 2. Raw material usage and molecular weight before and after azidation in the benzyl chloride azidation process.

[0108] (2) CuAAC grafting and aqueous self-assembly, micellar nanoreactors The azide polymers P-6 to P-10, the catalytic modules 1 to 4 obtained in Example 1, CuSO4, and sodium L-ascorbate were dissolved in water / THF (volume ratio 1:1) and reacted at 40 °C until the characteristic peak of the azide group in the infrared spectrum disappeared. After cooling, the mixture was thoroughly dialyzed with water to obtain a series of amphiphilic polymeric artificial enzymes. Benzyl azide forms a 1,2,3-triazole linker with the alkynyl group, allowing the catalytic module to be stably grafted near the aromatic hydrophobic fragment.

[0109] Figure 3 This diagram illustrates the self-assembly of the amphiphilic polymeric artificial enzyme of this invention in an aqueous phase, forming a hydrophilic canopy-hydrophobic catalytic core micelle structure. The resulting amphiphilic polymeric artificial enzyme can be directly dispersed in water to form a transparent or translucent solution without the need for traditional solvent replacement. The OEGMA side chains extend outwards to construct the hydrophilic canopy of the micelles; the styrene units, benzyl-triazole connecting arms, and catalytic modules rely on the synergistic effect of hydrophobicity and hydrogen bonding to drive the chain segments to aggregate inwards, thereby constructing a micelle nanoreactor that combines substrate enrichment capabilities with enzyme-mimicking catalysis.

[0110] Figure 4 The concentration of the polymer aqueous solution characterized by TEM was 0.5 mg / mL. Figure 4 The AF diagrams are P-5%-1, P-5%-3, P-20%-1, P-20%-3, P-25%50%-1, and P-25%50%-3, respectively; where 5%, 20%, and 25% represent the nominal molar fraction of 4-VBC in the polymerization feed, and numbers 1 and 3 represent the connected catalyst module 1 and catalyst module 3, respectively.

[0111] Figure 4 Transmission electron microscopy (TEM) characterization results showed that changing the polymer composition could yield micelle aggregates of different sizes. Under the condition that the nominal content of the catalytic modules remained consistent, the micelle size formed by the catalytic module 3 with introduced guanidine groups was usually larger than the micelles corresponding to the catalytic module 1 containing hydroxyl groups. This experimental phenomenon suggests that guanidine groups may enhance hydrogen bonding and ionic interactions and promote the formation of larger micelle aggregates.

[0112] Note: P-5%-y, P-10%-y and P-20%-y (y=1, 2, 3 or 4): artificial enzymes obtained by CuAAC reaction of P-6, P-7 and P-8 with catalytic module y (y=1, 2, 3 or 4), respectively; 5%, 10% and 20% are the nominal molar fractions of 4-VBC in the polymerization feed.

[0113] P-25%50%-y (y=1, 2, 3 or 4): Obtained by clicking the P-9 intermediate of OEGMA / styrene / 4-VBC=50 / 25 / 25 with the catalyst module y (y=1, 2, 3 or 4); where "25%" represents the VBC feed ratio and "50%" represents the OEGMA feed ratio.

[0114] P-25%30%-y (y=1, 2, 3 or 4): Obtained by clicking the P-10 intermediate of OEGMA / styrene / 4-VBC=30 / 45 / 25 with module y (y=1, 2, 3 or 4).

[0115] Example 4 This embodiment provides a palladium nanohybrid catalyst, comprising the amphiphilic polymeric artificial enzyme obtained in Example 3 and palladium nanoparticles dispersed and stabilized in the micelle matrix of the amphiphilic polymeric artificial enzyme. The preparation method is as follows.

[0116] PdCl2 was added to an aqueous solution of an amphiphilic polymeric artificial enzyme, and the mixture was stirred at room temperature to allow PdCl2 to react with the enzyme. 2+ The enzyme binds to carboxyl / carboxylate sites; ascorbic acid is then added as a mild reducing agent, and stirring is continued for approximately 24 hours. Unstable excess Pd particles are removed by centrifugation, and the clarified liquid is dried to obtain the artificial enzyme-PdNPs nanohybrid. Figure 5 TEM showed that PdNPs were dispersed in the polymer matrix, and EDX confirmed the presence of Pd elements.

[0117] The amphiphilic polymeric artificial enzyme was either P-10%-1 or P-10%-3 obtained in Example 3. The resulting palladium nano-hybrid catalysts were named P-10%-1-PdNPs or P-10%-3-PdNPs, respectively.

[0118] Fourier transform infrared spectroscopy (FTIR) Figure 6 Further data shows that the carboxylic acid concentrations of 2870 and 1726 cm⁻¹ in P-10%-3 are... -1 The signal is transformed into approximately 1622 cm⁻¹ after hybridization. -1 (COO) - (anti-symmetric stretching) and 1397 cm -1 (COO) - (Symmetrical expansion); P-10%-1 corresponds to 2868 and 1726 cm. -1Transformed into 1627 and 1394 cm -1 This supports the interaction between carboxylates and Pd species.

[0119] Example 5 This embodiment provides a method for preparing 4-nitrophenol by hydrolysis of PNB ester, including the following steps: PNB hydrolysis was performed in 25 mM borax-HCl buffer (pH 9.0). Each assay used 2.5 mL of buffer solution, with PNB added as acetonitrile solution, at final substrate concentrations of 10, 25, 50, 100, 150, 200, and 250 μM. A 2.5 μM catalyst (i.e., the amphiphilic polymeric artificial enzyme obtained in Example 3) was used; experiments labeled "low concentration" used 0.25 μM catalyst. The formation of 4-nitrophenol was monitored at 400 nm at 20 °C, and kinetic parameters were calculated using the Lineweaver-Burk double reciprocal method.

[0120] Table 3 Dynamic parameters

[0121] As shown in Table 3, among the various component series, catalytic module 1 typically provides the highest output. k cat / K m ;P-10%-1 k cat / K m 7.759 M -1 s -1 Reducing the catalyst concentration of P-20%-1 by 10 times, its k cat / K m Increased to 21.098 M -1 s -1 This indicates that excessively high site density or polymer concentration may inhibit apparent catalytic efficiency due to aggregation and reduced substrate accessibility. Adding 25 mol% styrene brought the activities of different modules closer together, while increasing styrene to 45 mol% amplified the differences between catalytic modules again, demonstrating the tunable role of the hydrophobic microenvironment.

[0122] Example 6 This embodiment provides a method for preparing 4 via an aldol addition reaction. (4 Nitrophenyl) 2 The method for hydroxycyclohexanone includes the following steps: The model reaction used 1.0 mmol of 4-nitrobenzaldehyde and 5.0 mmol of cyclohexanone as substrates, and added 10 wt.% of the amphiphilic polymeric artificial enzyme obtained in Example 3 as a catalyst. Three sets of conditions were set: Condition A was 2.0 mL of 25 mM borax-HCl buffer (pH 9.0) and reacted at 70 °C for 5-9 h; Condition B was 2.0 mL of water and reacted at 50 °C for 48 h; Condition C was 2.0 mL of water and reacted at 70 °C for 48 h.

[0123] Conversion rate equals the yield measured by separation / NMR, and the diastereomeric proportion (dr) is given by... 1 The enantiomeric excess value (ee) of the anti-isomer was determined by 1H NMR and by chiral high performance liquid chromatography (HPLC).

[0124] Table 4 Results of Aldol Reaction

[0125] in: dr(syn:anti): diastereomeric ratio (cis:trans); de: diastereomer excess value; ee anti Enantiomer excess value of trans isomer.

[0126] Under buffer conditions (condition A), all amphiphilic polymeric artificial enzymes achieved 100% conversion, and the catalytic reaction was typically completed within 10 h. Conversion was lower in water, but diastereoselectivity was improved in most systems. P-25%50%-4 showed the highest conversion of 79.2% at water / 50 °C, while P-25%50%-1 showed the highest de (de) of 47.1% at water / 70 °C. These results indicate that solvent, temperature, styrene content, and catalytic module structure can synergistically regulate reaction rate and relative stereoselectivity.

[0127] Example 7 This embodiment provides a method for preparing biphenyl via Suzuki coupling reaction, comprising the following steps: This embodiment utilizes a palladium nanoparticle hybrid catalyst to catalyze a Suzuki model coupling reaction to prepare biphenyl. The reaction substrate consisted of 212 μL (2.0 mmol) bromobenzene, 268 mg phenylboronic acid, 120 mg (3.0 mmol) sodium hydroxide, and 15.7 mg (5 wt.% palladium nominal mass fraction) of the palladium nanoparticle hybrid catalyst obtained in Example 4. 4.0 mL of water was used as the reaction medium, and the reaction was carried out at 50 °C with stirring for 24 h. After the reaction was completed, the palladium nanoparticle hybrid catalyst was recovered, and catalyst recycling experiments were conducted. The yield of biphenyl was determined by NMR or separation and weighing.

[0128] Table 5 Results of Suzuki Coupling Reaction

[0129] The yields of P-10%-3-PdNPs in the first round were 67.3%, and those of P-10%-1-PdNPs were 58.6%; in the fifth round, the yields remained at 36.3% and 41.5%, respectively. P-10%-3-PdNPs showed higher activity in the first round, while P-10%-1-PdNPs were more stable in multiple cycles, supporting the stability and reusability of PdNPs in this polymer platform.

[0130] Example 8 This embodiment evaluates the antibacterial and cell compatibility properties of amphiphilic polymeric artificial enzymes.

[0131] P-20% and P-25%30% series amphiphilic polymeric artificial enzyme samples were selected for use in Escherichia coli (E. coli). E. coli Inhibition assays and mammalian cell compatibility evaluations were performed. Antibacterial concentrations were set at 0.5 μg / mL, 5 μg / mL, 25 μg / mL, and 50 μg / mL, with gentamicin as a positive control. The results are shown in Table 7. Specifically, P-25%30%-4 at a concentration of 5 μg / mL showed... E. coli The bacterial growth inhibition rate is approximately 44%.

[0132] Cell compatibility assays were performed using human fibroblast 1BR3 cells and primary human osteoblasts. Cells were seeded in 24-well plates at a density of 5 × 10⁶ cells per well. 3 Each sample was incubated with 500 μL of culture medium at 37°C and 5% CO2 for 24 h. Cells were then treated at concentrations of 5 μg / mL, 25 μg / mL, and 50 μg / mL, and cell viability was assessed by flow cytometry. No significant cytotoxicity was observed within the tested concentration ranges using flow cytometry, demonstrating the good in vitro cell biocompatibility of this series of polymeric artificial enzymes.

[0133] The above description is merely a preferred embodiment of the present invention and is not intended to limit the invention. Various modifications and variations can be made to the present invention by those skilled in the art. Any modifications, equivalent substitutions, improvements, etc., made within the spirit and principles of the present invention should be included within the scope of protection of the present invention.

Claims

1. An amphiphilic polymeric artificial enzyme, characterized in that, It is a core-shell micelle structure formed by self-assembly in aqueous solution; it includes a statistical copolymer and a catalytic module; in the monomers of the statistical copolymer, the molar ratio of polyethylene glycol methyl ether methacrylate is 30-95 mol%, the molar ratio of 4-vinylbenzyl chloride is 5-25 mol%, and the molar ratio of styrene is 0-45 mol%, and the mixed monomers are copolymerized to obtain the statistical copolymer; the catalytic module is covalently grafted onto the side chain of the 4-vinylbenzyl chloride unit of the statistical copolymer; 4-vinylbenzyl chloride units grafted with catalytic modules aggregate inward to form a micelle core, while polyethylene glycol methyl ether methacrylate segments extend outward to form a hydration layer that encapsulates the micelle core. The structure of the catalytic module is as follows: R1 includes any one of hydroxyl and 2-guanidinoethyl; R2 includes any one of 2-imidazolyl and 4-imidazolyl.

2. The amphiphilic polymeric artificial enzyme as described in claim 1, characterized in that, The catalytic module is covalently linked to the side chain of the statistical copolymer 4-vinylbenzyl chloride unit via a 1,2,3-triazole ring.

3. The amphiphilic polymeric artificial enzyme as described in claim 1, characterized in that, The hydrophobic aromatic fragments and catalytic modules composed of styrene units and 4-vinylbenzyl chloride units aggregate inward to form a micelle core.

4. A method for preparing the amphiphilic polymeric artificial enzyme according to claim 1, characterized in that, include: By copolymerizing mixed monomers, statistical copolymers are obtained; In the statistical copolymer, the side chain benzyl chloride of the 4-vinylbenzyl chloride unit is converted into benzyl azide to obtain the benzyl azide-modified statistical copolymer; The benzyl azide-modified statistical copolymer undergoes a copper-catalyzed azide-alkyne cycloaddition reaction with a catalytic module. The catalytic module covalently grafts the generated 1,2,3-triazole ring onto the side chain of the statistical copolymer. The resulting polymer self-assembles in an aqueous solution to form a core-shell micelle structure.

5. The use of the amphiphilic polymeric artificial enzyme according to any one of claims 1-3 in catalyzing ester hydrolysis and / or aldol reactions.

6. A method for preparing 4-nitrophenol, characterized in that, Using p-nitrophenyl ester substrates as reactants, ester bonds are hydrolyzed under the catalysis of a catalyst to generate 4-nitrophenol; The catalyst is the amphiphilic polymeric artificial enzyme as described in any one of claims 1-3.

7. The use of the amphiphilic polymeric artificial enzyme according to any one of claims 1-3 in the preparation of antibacterial materials and / or cell-compatible catalytic materials.

8. A palladium nano-hybrid catalyst, characterized in that, It includes the amphiphilic polymeric artificial enzyme as described in any one of claims 1-3 and palladium nanoparticles in the micelle matrix anchoring the amphiphilic polymeric artificial enzyme.

9. The application of the palladium nano-hybrid catalyst according to claim 8 in the Suzuki coupling reaction.

10. A method for preparing biphenyl, characterized in that, In the presence of a catalyst, bromobenzene undergoes Suzuki coupling with phenylboronic acid to yield biphenyl. The catalyst is the palladium nano-hybrid catalyst as described in claim 8.

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