Imidazolyl zinc complex crystal and application thereof as ester hydrolysis mimic enzyme
The imidazole zinc complex crystal formed by L-carnosine and zinc ions solves the problem of low catalytic efficiency of existing imidazole zinc complexes, achieving efficient and stable ester hydrolysis catalysis, especially showing excellent catalytic activity for aromatic esters, and reducing synthesis costs.
Patent Information
- Application Number
- CN202511332552.7
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- Filing Date
- 2025-09-18
- Publication Date
- 2025-11-14
AI Technical Summary
Existing imidazole zinc complexes have lower catalytic efficiency than natural enzymes, especially for esters with large steric hindrance, where their catalytic activity is significantly reduced. Furthermore, traditional synthesis methods are complex, energy-intensive, and costly.
L-carnosine was used as a ligand to form an imidazole zinc complex crystal with zinc ions, which was synthesized in one step by a solvothermal method. Zinc ions act as Lewis acid nucleophiles to attack ester bonds, while L-carnosine provides hydrogen bonding sites, which enhances the stability of the substrate transition state. The resulting product has high catalytic activity and good stability.
It achieves catalytic activity higher than that of natural enzymes, especially exhibiting excellent catalytic performance for esters with large steric hindrance. The synthesis process is simple, low-cost, highly stable, and applicable to a wide range of conditions.
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Figure CN120943784A_ABST
Abstract
Description
Technical Field
[0001] This invention belongs to the field of organometallic complex technology, specifically relating to an imidazole zinc complex crystal and its application as an ester hydrolysis mimic enzyme. Background Technology
[0002] Ester hydrolysis is a key step in biological metabolic processes (such as lipolysis and neurotransmitter regulation) and industrial production (such as biodiesel production and chiral drug resolution). Natural hydrolases (such as esterases and lipases) can achieve efficient ester bond cleavage under mild conditions due to their high catalytic efficiency and stereoselectivity. However, the industrial application of natural enzymes has significant limitations: poor stability (susceptible to temperature, pH, and organic solvents), high separation and purification costs, low reusability, and difficulty in adapting to non-physiological industrial environments. Therefore, developing artificial enzyme systems with similar catalytic properties has become a research hotspot in the fields of chemistry and materials science.
[0003] To overcome the aforementioned defects of natural enzymes, some researchers have proposed using metal complexes to mimic the active site structure of enzymes, utilizing transition metal ions (such as ZnO2) 2+ Cu 2+ Fe 3+ Zinc ions, with their Lewis acidity and coordination ability, catalyze ester hydrolysis. Due to their high biocompatibility, redox inertness, and coordination flexibility, zinc ions have become the preferred metal center for constructing ester hydrolysis mimics. Early research mainly focused on complexes of zinc with multidentate macrocyclic ligands (such as cyclodextrins and porphyrins), which, while exhibiting some catalytic activity, suffered from complex synthesis steps, limited substrate selectivity, and low catalytic efficiency (only 1%–10% of that of natural enzymes). Therefore, subsequent research shifted towards small molecule ligand systems (such as amino acid derivatives and imidazoles) to improve catalytic performance by optimizing the coordination microenvironment. Among these, the imidazole group, due to its similarity to the histidine residues in the active site of natural enzymes, is considered an ideal module for constructing biomimetic catalytic sites.
[0004] In recent years, imidazole zinc complexes have made some progress in the field of ester hydrolysis mimics. Existing imidazole complexes exhibit some catalytic activity towards nitrobenzene ester substrates under alkaline conditions, but their catalytic efficiency remains lower than that of natural enzymes, especially for sterically hindered esters (such as aromatic esters), where their catalytic activity significantly decreases. Furthermore, the synthesis methods of traditional imidazole complexes typically require high-temperature reflux, inert gas protection, or multi-step purification, resulting in high energy consumption, and residual byproducts may inhibit catalytic activity. Therefore, developing ester hydrolysis mimics with high catalytic efficiency, high stability, and simple preparation processes is a pressing issue that needs to be addressed in this field. Summary of the Invention
[0005] The purpose of this invention is to provide an imidazole zinc complex crystal, which forms a new imidazole zinc complex crystal by coordinating zinc ions with L-carnosine. This crystal can be used as an ester hydrolysis mimic enzyme and has higher catalytic activity than traditional natural enzymes. It can effectively solve the technical problems of relatively low catalytic efficiency and stability of existing ester hydrolysis mimic enzymes. This invention also provides a method for preparing the above-mentioned imidazole zinc complex crystals. The imidazole zinc complex crystals can be synthesized in one step by using a solvothermal method. The overall synthesis process is simple and the synthesis cost is low, thereby solving the problems of relatively complex operation and high energy consumption in traditional imidazole complex crystal synthesis methods.
[0006] The present invention also provides an application of the above-mentioned imidazole zinc complex crystals as an ester hydrolysis mimic enzyme.
[0007] To achieve the above objectives, the first aspect of the present invention provides an imidazole zinc complex crystal with the chemical formula [Zn(L)](CH3CH2OH), wherein L represents L-carnosine, and its chemical structural formula is shown below:
[0008] One asymmetric structural unit of the imidazole zinc complex crystal contains a Zn(II), an L-carnosine ligand, and an ethanol molecule. The crystal structure of the imidazole zinc complex is monoclinic, space group P21; wherein, the coordination number of Zn(II) is 4, and each Zn(II) is coordinated with N or O atoms at four different positions on four L-carnosine ligands, wherein the N atoms are N1 and N4 on the imidazole ring and N3 on the amino group, and the O atoms are hydroxyl O1 on the carboxyl group.
[0009] The nitrogen and oxygen atoms on L-carnosine are labeled as follows:
[0010] Furthermore, the unit cell parameters of the imidazole zinc complex crystal are a=9.4155(2) Å, b=9.25933(13) Å, c=9.4236(2) Å; α=90°, β=118.740(3)°, γ=90°.
[0011] The imidazole zinc complex crystal of the present invention uses zinc ions as the metal center and L-carnosine as the organic ligand. The complex has a single crystal structure, exhibits the properties of an ester hydrolysis mimic enzyme, and has high catalytic activity. It can effectively solve the technical problem that the catalytic efficiency of existing ester hydrolysis mimic enzymes is significantly lower than that of natural enzymes. At the same time, the imidazole zinc complex also has good catalytic stability, and its catalytic activity is relatively less affected by factors such as temperature, pH value, and salt concentration.
[0012] Specifically, during catalytic ester hydrolysis, zinc ions, acting as Lewis acid nucleophiles, attack the carbonyl carbon in the ester bond, accelerating ester bond cleavage. L-carnosine has numerous metal coordination sites, enabling it to form complexes with metals. Simultaneously, L-carnosine contains flexible carboxylic acid side chains, which can utilize the carboxylic acid groups to provide additional hydrogen bonding sites, mimicking the electrostatic stabilizing function of "oxygen anion holes" in natural enzymes, thereby effectively enhancing the stabilization ability of the substrate transition state.
[0013] It is particularly important to note that the selection of metal ions and ligands is crucial in this application. Although some existing studies have attempted to use zinc complexes as ester hydrolases, the catalytic effect has been unsatisfactory due to inappropriate ligand selection, especially with a significant decrease in catalytic activity for sterically hindered esters (such as aromatic esters), as detailed in Table 1. In addition, the inventors of this application have also tried using carnosine complexes of other metal ions (such as Cu), or complexes formed by zinc ions with other ligands containing imidazole groups for ester hydrolysis, but the results have also been unsatisfactory.
[0014] According to any of the technical solutions described in the first aspect of the present invention, the imidazole zinc complex crystal has a diffraction peak at 10.9±0.2° in the XRD diffraction pattern.
[0015] The second aspect of the present invention also provides a method for preparing imidazole zinc complex crystals as described in the first aspect of the present invention, comprising: causing L-carnosine to undergo a coordination reaction with a soluble zinc salt in a reaction system containing water and anhydrous ethanol; and then cooling and filtering the reaction product to obtain the imidazole zinc complex crystals.
[0016] This invention enables the one-step synthesis of the imidazolyl zinc complex crystals using a solvothermal method. The overall process is simple and effectively ensures the high stability and catalytic activity of the obtained complex crystals. It should also be noted that the choice of solvent is crucial in this invention. Using a water-ethanol mixed solvent instead of toxic organic media not only allows for simultaneous crystallization and purification of the complex without complex post-processing, resulting in lower production costs, but also affects the microstructure and catalytic activity of the obtained complex. By using the mixed solvent of this invention, complexes with stable single-crystal structures can be obtained over a wide range of conditions, thus further ensuring the catalytic activity of the complex crystals. The inventors attempted to use other solvents mixed with water, but only the ethanol-water mixture as a solvent yielded imidazolyl zinc complex crystals with large size, high quality, good catalytic performance, and ease of detection and application.
[0017] According to any of the technical solutions described in the second aspect of the present invention, based on 1 mmol of L-carnosine, the total volume of the mixed solvent composed of water and anhydrous ethanol is 10-20 mL; the volume ratio of water to anhydrous ethanol in the mixed solvent is 4:1 to 1:4. The amount of reaction solvent added and the volume ratio of water to anhydrous ethanol affect the solubility of L-carnosine in the reaction system, thereby affecting the yield of the complex. The present invention optimizes and controls the amount of reaction solvent added and the volume ratio of water to anhydrous ethanol, thereby ensuring good solubility of L-carnosine in the reaction system, enabling the coordination reaction to proceed smoothly, and obtaining a complex with an ideal single-crystal structure.
[0018] More preferably, in the mixed solvent composed of water and anhydrous ethanol, the volume ratio of water to anhydrous ethanol is 1:(1~2).
[0019] According to any of the technical solutions described in the second aspect of the present invention, the entire reaction process does not require the addition of sodium methoxide or sodium ethoxide. In the prior art, sodium methoxide or sodium ethoxide is used to synthesize L-carnosine zinc complexes for ligands with active hydrogen. These, as strong bases, can deprotonate the nitrogen atom on the imidazole ring and neutralize the carboxylic acid, allowing carnosine to dissolve in organic solvents, activating the carnosine molecule, and enabling it to efficiently bind with zinc ions. The present invention eliminates the need for sodium ethoxide activation, forming a different coordination structure than the method using sodium ethoxide, resulting in high-quality complex crystals.
[0020] According to any of the technical solutions described in the second aspect of the present invention, the molar ratio of zinc ions to L-carnosine in the soluble zinc salt is 8:1 to 1:8. More specifically, the soluble zinc salt is preferably zinc nitrate hexahydrate, zinc chloride, or zinc acetate.
[0021] According to any of the technical solutions described in the second aspect of the present invention, the reaction temperature of the coordination reaction is 60~100℃ and the reaction time is 24~72 hours.
[0022] Furthermore, the reaction temperature of the coordination reaction is 75~85℃; Furthermore, the reaction temperature of the coordination reaction is 78~82℃; Furthermore, the coordination reaction was carried out at a temperature of 80°C. Synthesis at 80°C yielded high-quality bulk single crystals, with the crystal structure ultimately refined and converged to R1=0.024 and wR2=0.0637, indicating a highly reliable structural model. However, at lower temperatures, such as 20–50°C, it is difficult to obtain ideal crystals or high-quality single crystals.
[0023] Further optimization and control of reaction temperature and reaction time can help ensure the smooth progress of coordination reaction and the yield of the obtained complex crystals. However, if the reaction time is too long, not only will more energy be consumed, but the reaction yield will also be difficult to improve further due to the constraints of reaction kinetics.
[0024] According to any of the technical solutions described in the second aspect of the present invention, after the coordination reaction is completed, the reaction mixture is cooled to 20~30°C and filtered to obtain imidazole zinc complex crystals.
[0025] The third aspect of the present invention also provides the application of the imidazole zinc complex crystals described in the first aspect of the present invention, or the imidazole zinc complex crystals prepared by the preparation method described in the second aspect of the present invention, as ester hydrolysis mimics.
[0026] According to the application described in the third aspect of the present invention, the imidazole zinc complex crystals are applied to the hydrolysis reaction of p-nitrobenzene acetate. Based on 1 mmol of p-nitrobenzene acetate, the amount of imidazole zinc complex crystals used is preferably 500-3000 mg, more preferably 1500-3000 mg; the hydrolysis reaction temperature is 30-90°C, and the pH of the reaction solution is 3-11. The imidazole zinc complex crystals of the present invention exhibit high catalytic activity for the hydrolysis of p-nitrobenzene acetate, and also high catalytic stability. When parameters such as the temperature, pH, and salt concentration of the hydrolysis reaction change, the catalytic activity and catalytic efficiency of the complex crystals fluctuate relatively little.
[0027] Furthermore, the temperature of the hydrolysis reaction is preferably 50-90°C, and the pH value of the reaction solution is preferably 5-11, and even more preferably 9-11.
[0028] In summary, by adopting the technical solution provided by this invention, the following beneficial effects can be achieved compared with the prior art: (1) The present invention provides an imidazole zinc complex crystal formed with L-carnosine as a ligand. The imidazole zinc complex crystal has a crystal structure different from that of imidazole zinc complexes in the prior art. It has high crystal stability and has ester hydrolysis catalytic activity that is significantly higher than that of imidazole zinc complexes in the prior art and significantly higher than that of complexes formed by other metals and carnosine. At the same time, the imidazole zinc complex crystal of the present invention has catalytic activity that is significantly higher than that of traditional natural enzymes. Compared with the catalytic activity of traditional natural enzymes, which is affected by temperature, pH, salt concentration, etc., the imidazole zinc complex crystal of the present invention is almost unaffected by these factors and has high and stable catalytic activity under a wider range of applicable conditions. This is because the flexible carboxylic acid side chain contained in L-carnosine can provide additional hydrogen bond interaction sites, which can simulate the electrostatic stabilization function of "oxygen anion hole" in natural enzymes and enhance the substrate transition state stabilization ability. As a result, the complex crystal has high ester hydrolysis catalytic activity, especially able to meet the catalytic activity requirements of ester hydrolysis with large steric hindrance.
[0029] (2) The present invention can directly synthesize the imidazole zinc complex crystals in one step by using a solvothermal method. The overall operation is simple. In particular, by using a water + ethanol mixed solvent, not only can the formation of single crystal structure complexes be guaranteed, which is beneficial to improving the catalytic activity of the obtained complexes, but the preparation conditions are not demanding. The crystallization and purification of the complexes can be completed simultaneously without complicated post-processing, which can further reduce production costs. At the same time, the size of the obtained imidazole zinc complex crystals is relatively large and the crystal quality is stable, which is conducive to detection and application.
[0030] (3) The present invention further optimizes the specific reaction process parameters of the coordination reaction, such as the amount of mixed solvent used, the volume ratio of water to ethanol, and the reaction temperature and time, to improve the solubility of L-carnosine, so that the coordination reaction can proceed smoothly and the yield of the obtained complex crystals can be guaranteed.
[0031] (4) The imidazole zinc complex of the present invention can be effectively used to catalyze ester hydrolysis reactions. When applied to the hydrolysis of p-nitrobenzene acetic acid, it can catalyze the hydrolysis of p-nitrobenzene acetic acid quickly and efficiently. Its catalytic activity is better than that of lipase and has high catalytic stability. It can effectively resist the influence of changes in parameters such as temperature and pH on catalytic activity. Attached Figure Description
[0032] Figure 1 This is a two-dimensional structural diagram of the imidazole zinc complex crystal prepared in Example 3 of the present invention along the b-axis (hydrogen atoms and solvent molecules have been omitted). Figure 2 These are the complexes A1-A7 prepared in the embodiments of the present invention and their simulated PXRD spectra; Figure 3This is a comparison chart of the conversion rates of the imidazole zinc complex crystals A1-A7 prepared in Examples 1-7 to the substrate p-nitrophenyl acetate; Figure 4 This is a schematic diagram of the catalytic hydrolysis reaction mechanism of the imidazole zinc complex crystals of the present invention; color codes: Zn, turquoise; C, gray; O, red; N, blue; H, white; Figure 5 This is a comparison chart of the conversion rates of the substrate p-nitrophenyl acetate by different amounts of imidazole zinc complex crystals mimicking enzymes. Figure 6 This is a comparison chart of the conversion rates of imidazole zinc complex crystals and lipase to the substrate p-nitrophenyl acetate under different temperature conditions. Figure 7 This is a comparison of the conversion rates of imidazole zinc complex crystals and lipase to the substrate p-nitrobenzene acetate under different pH conditions. Figure 8 This is a comparison chart of the conversion rates of imidazole zinc complex crystals and lipase to the substrate p-nitrobenzene acetate under different salt concentrations. Figure 9 This is a comparison chart of the conversion rates of different metal carnosine complexes and lipases to the substrate p-nitrobenzene acetate. Figure 10 This is a comparison chart of the conversion rates of each complex in Comparative Examples 7-9 to the substrate p-nitrophenyl acetate. Figure 11 This is a comparison chart showing the conversion rates of complexes prepared using different reaction solvents to the substrate p-nitrophenyl acetate. Figure 12 These are the PXRD spectra of different imidazole zinc complexes synthesized with different solvents; Figure 13 This is a two-dimensional structural diagram of the Cu-LC crystal along the b-axis in Comparative Example 5 (hydrogen atoms and solvent molecules have been omitted). Figure 14 The PXRD spectrum of Cu-LC in Comparative Example 5 is shown. Figure 15 These are the PXRD spectra of the complexes A8-A13 prepared in the embodiments of the present invention; Figure 16 This is a comparison chart of the conversion rates of the complexes A8-A13 prepared in the embodiments of the present invention to the substrate p-nitrophenyl acetate. Detailed Implementation
[0033] This invention provides an imidazole zinc complex crystal, its preparation method, and its application as an ester hydrolysis mimic enzyme. The imidazole zinc complex has a single-crystal structure (see the crystal structure in Example 3 below), and its chemical formula is [Zn(L)](CH3CH2OH), where L represents L-carnosine. The chemical structural formula is shown below: ; An asymmetric structural unit in an imidazole zinc complex crystal contains a Zn(II), an L-carnosine ligand, and an ethanol molecule. The crystal structure of the imidazole zinc complex is a monoclinic crystal system with space group P21; the coordination number of Zn(II) is 4, and each Zn(II) is coordinated with N or O atoms from four different positions on four L-carnosine ligands, where the N atoms are N1 and N4 on the imidazole ring and N3 on the amino group, and the O atom is the hydroxyl group O1 on the carboxyl group.
[0034] The imidazole zinc complex crystals of the present invention possess the properties of an ester hydrolysis enzyme, exhibiting high catalytic activity and stability, and low cost. The present invention will be further described below with reference to specific embodiments; however, it should be understood that all the following embodiments are merely illustrative and not limiting, and should not be construed as limiting the scope of protection of the present invention.
[0035] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention pertains. Furthermore, the ranges of concentrations, amounts, and other numerical data defined herein should be flexibly interpreted to include not only the values explicitly stated as the limits of the range, but also all individual values or subranges encompassed within said range, as if each value and subrange were explicitly stated. For example, a numerical range of 1 to 4.5 should be interpreted to include not only the explicitly stated limits of 1 to 4.5, but also individual numbers (such as 2, 3, 4) and subranges (such as 1 to 3, 2 to 4, etc.). The same principle applies to ranges that describe only a single value, such as "less than 4.5," which should be interpreted to include all the values and ranges described above. Moreover, this interpretation should apply regardless of the breadth of the range or feature described.
[0036] The following descriptions of the sources of pharmaceuticals and solvents used in the examples, comparative examples, and test cases are as follows: p-nitrobenzene acetate (also known as "p-nitrobenzene acetate", CAS No.: 830-03-5), p-nitrophenol, lipase (CAS No.: 9001-62-1, 20,000 U / g, product number L874984), and L-carnosine were all purchased from Shanghai Maclean Biochemical Technology Co., Ltd. Anhydrous ethanol, acetonitrile, anhydrous methanol, and N,N'-dimethylformamide (DMF) were purchased from Lianyungang Bohua (Tianjin) Pharmaceutical Chemical Co., Ltd. Zinc nitrate hexahydrate and sodium chloride were purchased from Chengdu Kelong Chemical Co., Ltd., and the water was double-distilled water.
[0037] The test methods used in the following examples, comparative examples, and test cases are as follows: elemental analysis was performed using the Elementar UNICUBE elemental analyzer (Germany); infrared spectroscopy was performed using the Nicolet is5 Fourier transform infrared spectrometer (Thermo Fisher Scientific, USA); PXRD was performed using the PAN alytical X-ray powder diffractometer X'PertPro (Netherlands); single-crystal diffraction was performed using the Rigaku XtaLAB Synergy-DW single-crystal diffractometer (Japan); and ultraviolet-visible spectroscopy was performed using the Shimadzu UV-2700 ultraviolet-visible spectrophotometer (Japan).
[0038] Example 1 Take 0.226 g (1.0 mmol) L-carnosine, 0.297 g (1.0 mmol) Zn(NO3)2•6H2O, and 20 mL of a mixed solvent of water and anhydrous ethanol (volume ratio of water to anhydrous ethanol is 1:1), sonicate for 5 minutes, and react in a constant temperature oven at 80 °C for 48 hours to obtain colorless crystals A1 with a yield of 60%.
[0039] Elemental analysis of A1 yielded a result of C. 11 H 18 N4O4Zn: C, 39.11%; N, 16.56%; H, 5.39%; theoretically calculated values are: C, 39.36%; N, 16.69%; H, 5.41%, indicating that the elemental analysis results are consistent with the theoretical calculations. IR (KBrdisc, cm⁻¹) -1 ): 3440, 3250, 3155, 3097, 2968, 1643, 1619, 1576, 1482, 1385, 1343, 1272, 1237, 1188, 1127, 1056, 1034, 1007, 981, 875, 742, 704, 660, 470, 431; of which 3440cm -1 This is the stretching vibration peak of NH on the amino group; 1619 cm⁻¹ -1 These are the stretching vibration peaks of C=N and C=C on the imidazole ring; 1576 cm⁻¹ -1 and 1385cm -1 COO - Asymmetric stretching vibration peaks and symmetric stretching vibration peaks; 470 cm -1 The peak represents the stretching vibration of Zn-N; 431 cm⁻¹ -1 The peak represents the Zn-O stretching vibration. PXRD analysis of A1 is as follows... Figure 2 As shown, in the XRD diffraction pattern, 2θ has a diffraction peak at 10.9±0.2°.
[0040] Example 2 This embodiment uses the method of Example 1 to prepare imidazole zinc complex crystals. The difference from Example 1 is that the Zn(NO3)2•6H2O content in this embodiment is 0.125 mmol. This embodiment yields colorless crystals A2 with a yield of 62%.
[0041] Elemental analysis of A2 yielded a result of C. 11 H 18 N4O4Zn: C, 39.40%; N, 16.81%; H, 5.33%, while the theoretically calculated values are: C, 39.36%; N, 16.69%; H, 5.41%, indicating that the elemental analysis results are consistent with the theoretical calculations. PXRD detection is as follows... Figure 2 As shown.
[0042] Example 3 This embodiment uses the method of Example 1 to prepare imidazole zinc complex crystals. The difference from Example 1 is that the Zn(NO3)2•6H2O content in this embodiment is 0.25 mmol. This embodiment yields colorless crystals A3 with a yield of 62%.
[0043] Elemental analysis of A3 yielded a result of C. 11 H 18 N4O4Zn: C, 39.45%; N, 16.83%; H, 5.21%, while the theoretically calculated values are: C, 39.36%; N, 16.69%; H, 5.41%, indicating that the elemental analysis results are consistent with the theoretical calculations. PXRD detection is as follows... Figure 2 As shown.
[0044] Example 4 This embodiment uses the method of Example 1 to prepare the imidazole zinc complex. The difference from Example 1 is that the Zn(NO3)2•6H2O content in this embodiment is 0.5 mmol. This embodiment yields colorless crystals A4 with a yield of 62%.
[0045] The elemental analysis of A4 yielded a result of C. 11 H 18 N4O4Zn: C, 39.38%; N, 16.73%; H, 5.32%, while the theoretically calculated values are: C, 39.36%; N, 16.69%; H, 5.41%, indicating that the elemental analysis results are consistent with the theoretical calculations. PXRD detection is as follows... Figure 2 As shown.
[0046] Example 5 This embodiment uses the method of Example 1 to prepare imidazole zinc complexes. The difference from Example 1 is that the Zn(NO3)2•6H2O content in this embodiment is 2 mmol. This embodiment yields colorless crystals A5 with a yield of 62%.
[0047] Elemental analysis of A5 yielded a result of C. 11 H 18 N4O4Zn: C, 39.57%; N, 16.77%; H, 5.22%, while the theoretically calculated values are: C, 39.36%; N, 16.69%; H, 5.41%, indicating that the elemental analysis results are consistent with the theoretical calculations. PXRD detection is as follows... Figure 2 As shown.
[0048] Example 6 This embodiment uses the method of Example 1 to prepare imidazole zinc complexes. The difference from Example 1 is that the Zn(NO3)2•6H2O content in this embodiment is 4 mmol. This embodiment yields colorless crystals A6 with a yield of 62%.
[0049] Elemental analysis of A6 yielded a result of C. 11 H 18 N4O4Zn: C, 39.44%; N, 16.65%; H, 5.35%, while the theoretically calculated values are: C, 39.36%; N, 16.69%; H, 5.41%, indicating that the elemental analysis results are consistent with the theoretical calculations. PXRD detection is as follows... Figure 2 As shown.
[0050] Example 7 This embodiment uses the method of Example 1 to prepare imidazole zinc complexes. The difference from Example 1 is that the Zn(NO3)2•6H2O content in this embodiment is 8 mmol. This embodiment yields colorless crystals A7 with a yield of 62%.
[0051] Elemental analysis of A7 yielded a result of C. 11 H 18 N4O4Zn: C, 39.47%; N, 16.72%; H, 5.36%. The theoretical calculated values are: C, 39.36%; N, 16.69%; H, 5.41%, indicating that the elemental analysis results are consistent with the theoretical calculations. PXRD detection is as follows... Figure 2 As shown.
[0052] The PXRD results of A1 to A7 show that, within the range of 1:8 to 8:1 in the molar ratio of zinc to carnosine, the imidazole zinc complex single crystals prepared by the method of this invention all have the same structure, that is, the same imidazole zinc complex crystal structure as in Example 3. According to the analysis results of the imidazole zinc complex crystal, the cell parameters of the crystal are a=9.4155(2) Å, b=9.25933(13) Å, c=9.4236(2) Å; α=90°, β=118.740(3)°, γ=90°.
[0053] Application test in the hydrolysis reaction of p-nitrobenzene acetate: The imidazole zinc complex crystals (Zn-LC) prepared in Examples 1-7 were used in the hydrolysis reaction of p-nitrobenzene acetate, and the specific operations are as follows: Add 4.9 mL of PBS buffer (pH 7.4) to a 20 mL glass vial, then add 5.0 mg of the imidazole zinc complex crystals prepared in Examples 1-7 respectively. Stir magnetically in a 40 °C water bath for 10 minutes, then add 100 μL of 0.5 mol•L... -1 An acetonitrile solution of p-nitrobenzene acetate was prepared. After 4 hours, 100 μL of the reaction solution was taken and diluted with 2.9 mL of PBS buffer (pH 7.4). The absorbance was then measured at 400 nm. The catalytic results of the hydrolysis of p-nitrobenzene acetate by the imidazole zinc complex crystals prepared in different examples are shown below. Figure 3 As shown. Combined with Figure 3 It can be seen that when the molar ratio of zinc ions to L-carnosine in the soluble zinc salt during the preparation process is 8:1 to 1:8, the resulting zinc complex crystals all have high catalytic activity for the hydrolysis of p-nitrobenzene acetate, and the hydrolysis conversion rate of p-nitrobenzene acetate is higher than 73%, with a conversion rate range of 73.80% to 78.38%. This indicates that the conversion rate of p-nitrobenzene acetate hydrolyzed by the imidazole zinc complex crystals of the present invention is not only high but also very stable.
[0054] Catalytic mechanism: Combination Figure 4As shown, the potential reaction pathway for Zn-LC ester hydrolysis mimic enzymes as catalysts to promote the hydrolysis of p-nitrophenylacetic acid (P-NPA) involves several key steps. First, the p-nitrophenylacetic acid molecule is activated through interaction with the catalyst, which provides a Lewis acid center, thereby enhancing the electrophilicity of the carbonyl carbon in the acetate group. Nucleophiles coordinated to the catalyst (such as water molecules or hydroxide ions) attack the carbonyl carbon, forming a tetrahedral intermediate. The catalyst stabilizes this intermediate through coordination or hydrogen bonding, and the electron-withdrawing nitro group on the substrate benzene ring further disperses the negative charge. Finally, the reaction ends with the release of the acetate product and the regeneration of the catalytic site, allowing the Zn-LC ester hydrolysis mimic enzyme catalyst to participate in subsequent reaction cycles. The Zn-LC mimic enzyme catalyst effectively accelerates the hydrolysis reaction by enhancing the electrophilicity of the carbonyl group, stabilizing the transition state, promoting nucleophilic attack, and stabilizing the leaving group.
[0055] To further demonstrate the stability of the imidazole zinc complex crystals of the present invention, the effects of hydrolysis reaction temperature, pH value, salt concentration, and amount of complex on the hydrolysis catalytic effect of acetic acid on the complex crystals prepared in Example 3 are studied below, and compared with the catalytic effect of lipase.
[0056] Test Example 1 The imidazole zinc complex crystals prepared in Example 3 were applied to the hydrolysis reaction of p-nitrobenzene acetate, and the effect of the amount of complex on the activity of the ester hydrolysis mimic enzyme was studied. The specific operation is as follows: Add 4.9 mL of pH 7.4 PBS buffer to a 20 mL glass vial, then add 2.5, 5.0, 7.5, 10.0, and 15.0 mg of imidazole zinc complex crystals. Stir magnetically in a 40°C water bath for 10 minutes. Then add 100 μL of 0.5 mol / L... -1 An acetonitrile solution of p-nitrobenzene acetate was prepared. After 4 hours, 100 μL of the reaction solution was taken and diluted with 2.9 mL of PBS buffer (pH 7.4). The absorbance was then measured at 400 nm. The results are as follows. Figure 5 As shown. From Figure 5 It can be seen that with the increase of the amount of imidazole zinc complex crystals, the hydrolysis conversion rate of p-nitrobenzene acetate under the same conditions gradually increases, until the amount of added imidazole zinc complex crystals reaches 10 mg, at which point the hydrolysis conversion rate of p-nitrobenzene acetate reaches over 90%. Further increases in the amount of imidazole zinc complex crystals have a smaller effect on the conversion rate. Therefore, based on 1 mmol of p-nitrobenzene acetate, the preferred amount of imidazole zinc complex is 500-3000 mg, more preferably 1500-3000 mg, and even more preferably 2000-3000 mg.
[0057] Test Example 2 The imidazole zinc complex crystals prepared in Example 3 were applied to the hydrolysis reaction of p-nitrobenzene acetate, and the effect of temperature on the activity of the ester hydrolysis mimic enzyme was studied. The specific operation is as follows: Add 4.9 mL of pH 7.4 PBS buffer to a 20 mL glass vial, then add 10.0 mg of imidazole zinc complex crystals. Stir magnetically in a water bath at 30, 40, 50, 60, 70, 80, and 90 °C for 30 minutes each, then stir magnetically in a 40 °C water bath for 10 minutes. Finally, add 100 μL of 0.5 mol•L... -1 An acetonitrile solution of p-nitrobenzene acetate was prepared. After 4 hours, 100 μL of the reaction solution was taken and diluted with 2.9 mL of PBS buffer (pH 7.4), and the absorbance was measured at 400 nm. The results are as follows. Figure 6 As shown.
[0058] Comparative Example 1 All other conditions were the same as in Test Example 2, except that a natural enzyme (Lipase, also added at 10 mg) was used instead of the imidazole zinc complex crystals. The catalytic reaction results are as follows. Figure 6 As shown.
[0059] Figure 6 The results showed that, under the same conditions, the conversion rate of the imidazole zinc complex crystals to the hydrolysis of p-nitrobenzene acetate remained above 87% in the range of 30–90 °C. In contrast, the catalytic activity of the natural enzyme (Lipase, also added at 10 mg) was greatly affected by temperature, with a conversion rate of about 80% in the range of 30–60 °C, but only about 30% in the range of 70–90 °C.
[0060] Test Example 3 The imidazole zinc complex crystals prepared in Example 3 were applied to the hydrolysis reaction of p-nitrobenzene acetate, and the effect of pH on the activity of the ester hydrolysis mimic enzyme was studied. The specific operation is as follows: Add 4.9 mL of PBS buffer solutions with pH values of 3.0, 5.0, 7.0, 9.0, and 11.0 to a 20 mL glass vial, then add 10.0 mg of imidazole zinc complex crystals. Stir magnetically at room temperature for 8 hours, then stir magnetically in a 40 °C water bath for 10 minutes. Finally, add 100 μL of 0.5 mol / L... -1 Acetonitrile solution of p-nitrobenzene acetate. After 4 hours, 100 μL of the reaction solution was taken and diluted with 2.9 mL of PBS buffer at pH 3.0, 5.0, 7.0, 9.0, and 11.0, respectively, and the absorbance was measured at 400 nm. The results are as follows. Figure 7 As shown.
[0061] Comparative Example 2 All other conditions were the same as in Test Example 3, except that a natural enzyme (Lipase, also added at 10 mg) was used instead of the imidazole zinc complex crystals. The catalytic reaction results are as follows. Figure 7 As shown.
[0062] Figure 7 The results showed that, under the same conditions, within a pH range of 3 to 11, the conversion rate of imidazole zinc complex crystals for the hydrolysis of p-nitrobenzene acetate remained above 87%, while the catalytic activity of the natural enzyme (Lipase, also added at 10 mg) was greatly affected by pH, with a conversion rate of 78 to 84% at around pH 5 to 7, while the conversion rate was only about 60% at lower or higher pH.
[0063] Test Example 4 The imidazole zinc complex crystals prepared in Example 3 were applied to the hydrolysis reaction of p-nitrobenzene acetate, and the effect of salt concentration on the activity of the simulated enzyme was studied. The specific operation is as follows: Add 4.9 mL of PBS buffer (pH 7.4) to a 20 mL glass vial, then add 10.0 mg of imidazole zinc complex crystals, followed by the addition of NaCl to achieve NaCl concentrations of 0, 150, 300, and 500 mmol / L. -1 Then, stir magnetically in a 40°C water bath for 10 minutes, followed by adding 100 μL of 0.5 mol•L⁻¹. -1 An acetonitrile solution of p-nitrobenzene acetate was prepared. After 4 hours, 100 μL of the reaction solution was taken and diluted with 2.9 mL of PBS buffer (pH 7.4), and the absorbance was measured at 400 nm. The results are as follows. Figure 8 As shown.
[0064] Comparative Example 3 All other conditions were the same as in Test Example 4, except that a natural enzyme (Lipase, also added at 10 mg) was used instead of the imidazole zinc complex crystals. The catalytic reaction results are as follows. Figure 8 As shown.
[0065] Figure 8 The results showed that, under the same conditions, within the salt concentration range of 0–500 mM, the conversion rate of the imidazole zinc complex crystals for the hydrolysis of p-nitrobenzene acetate remained above 82%, while the catalytic activity of the natural enzyme (Lipase, also added at 10 mg) fluctuated significantly due to the influence of salt concentration.
[0066] Therefore, in summary Figures 6-8The test results show that, compared with lipase, the imidazole zinc complex of the present invention not only has higher catalytic activity for the hydrolysis of p-nitrobenzene acetate, but also has better stability. The catalytic activity of lipase fluctuates greatly due to the influence of temperature, pH and salt concentration of the hydrolysis reaction, while the catalytic activity of the imidazole zinc complex of the present invention is less affected by the above factors.
[0067] It should be noted that, due to space limitations, this section only uses the complex prepared in Example 3 as an example to illustrate some performance test examples under specific application conditions. When the complex crystals used in each test example are the same but other hydrolysis reaction process parameters change (such as adjusting the pH value or the amount of complex in the test example within a certain range), the comparison results of their performance tests can also lead to the above conclusions.
[0068] Comparative Example 4 In this comparative example, lipase was used to catalyze the hydrolysis of p-nitrobenzene acetate. The specific process parameters were basically the same as in Test Example 1, and the amount of lipase used was 5 mg.
[0069] Comparative Example 5 In this comparative example, Cu was used instead of zinc to coordinate with L-carnosine to form Cu carnosine complex crystals. The specific preparation method was as follows: 0.226 g (1.0 mmol) of L-carnosine, 0.200 g (1.0 mmol) of Cu (CH3COO)2•H2O, and 20 mL of a mixed solvent of water, anhydrous methanol and DMF (volume ratio of water, anhydrous methanol and DMF was 1:3:1) were taken, sonicated for 5 minutes, and reacted in a constant temperature oven at 80℃ for 48 hours to obtain blue crystals with a yield of 59%.
[0070] The elemental analysis of the sample yielded the result of C9H. 16 CuN5O4: C, 33.43%; N, 17.32%; H, 4.91%, while the theoretically calculated values are: C, 33.38%; N, 17.30%; H, 4.98%, indicating that the elemental analysis results are in good agreement with the theoretical calculations. IR (KBrdisc, cm⁻¹) -1 (): 3425, 3260, 3150, 3064, 2950, 1618, 1561, 1501, 1468, 1435, 1403, 1366, 1320, 1287, 1274, 1235, 1180, 1141, 1109, 1039, 1003, 967, 898, 878, 840, 687, 655, 536, 473; obtained by single-crystal diffraction testing as follows Figure 13 The structure shown is PXRD detected as follows. Figure 14 As shown.
[0071] Figure 13In the copper complex crystal, an asymmetric structural unit comprises a Cu(II), an L-carnosine ligand, a water molecule coordinated with copper, and a free water molecule; the crystal structure is a monoclinic crystal system with space group P3121; the coordination number of Cu(II) is 5, and each Cu(II) is coordinated with N003 on the peptide bond of the L-carnosine ligand, N00E on the amino group, O002 on the carboxyl group, O009 on the water molecule, and N008 on the imidazole ring of another L-carnosine ligand; the unit cell parameters are a = 8.548 Å, b = 8.548 Å, c = 30.625 Å; α = 90°, β = 90°, γ = 120°.
[0072] The carnosine complex crystals of Cu in this comparative example were used in the hydrolysis reaction of p-nitrobenzene acetate. The specific application process parameters were basically the same as those in Test Example 1, and the amount of carnosine complex of Cu was 5 mg.
[0073] The conversion rates of Comparative Examples 4 and 5, and 5 mg Zn-LC under the same conditions, for the hydrolysis of p-nitrobenzene acetate are compared as follows: Figure 9 As shown in the figure, the catalytic activity of the imidazole zinc complex crystal (Zn-LC) for p-nitrobenzene acetate in this application is much higher than that of the Cu carnosine complex crystal; the catalytic effect of the Cu carnosine complex crystal on p-nitrobenzene acetate is lower than that of lipase, while the catalytic activity of the imidazole zinc complex crystal (Zn-LC) for p-nitrobenzene acetate in this application is higher than that of lipase.
[0074] Comparative Examples 6-8 In Comparative Examples 6-8, 5-carboxybenzimidazole, 2-hydroxymethyl-5-carboxybenzimidazole, and 2-methylimidazolium-4,5-dicarboxylic acid were used as organic ligands, respectively, and coordinated with zinc ions and other metal ions to form corresponding imidazole complexes. Each imidazole complex was then used for the hydrolysis catalytic reaction of p-nitrobenzene acetate under essentially the same reaction conditions as in Test Example 1, with each complex used at a concentration of 10 mg. The catalytic conversion rates of each complex for p-nitrobenzene acetate are shown below. Figure 10 As shown. According to Figure 10 It can be seen that the ester hydrolysis catalytic activity of the complexes formed by coordinating 5-carboxybenzimidazole, 2-hydroxymethyl-5-carboxybenzimidazole, and 2-methylimidazolium-4,5-dicarboxylic acid with zinc ions or other metal ions using 5-carboxybenzimidazole, 2-hydroxymethyl-5-carboxybenzimidazole, and 2-methylimidazolium-4,5-dicarboxylic acid as organic ligands is much lower than that of the imidazolium zinc complex crystal in this application (Test Example 1). Figure 5 (Conversion rate was 90.58% at 10mg).
[0075]
[0076] Table 1 shows comparative data on zinc complexes used for ester hydrolysis catalysis in other existing studies. The data in Table 1 further demonstrates that the imidazole zinc complex crystals in this application have a significantly better ester hydrolysis catalytic effect than existing technologies.
[0077] Note: The literature [1] is Mimic Carbonic Anhydrase Using Metal–Organic Frameworks forCO2Capture and Conversion[J / OL]. Inorganic Chemistry, 2018, 57(4): 2169-2174. The literature [2] is A Zinc Coordination Complex Mimicking Carbonic Anhydrase for CO2Hydrolysis and Sequestration [J / OL]. Inorganic Chemistry, 2019, 58(15):9916-9921. Literature [3] is Influence of ZIF-8 polymorphism catalysts and nanosizes inp-nitrophenyl ester hydrolysis[J / OL]. Applied Organometallic Chemistry, 2023,37(7): e7136. Literature [4] is Bio-inspired nanozyme: a hydratase mimic in a zeoliticimidazolate framework [J / OL]. Nanoscale, 2019, 11(13): 5960-5966. Comparative Examples 9-11 Compared to Example 3, the main difference between Comparative Examples 9-11 is that methanol (MeOH):water volume ratio 1:1, acetonitrile (MeCN):water volume ratio 1:1, and N,N-dimethylformamide (DMF):water volume ratio 1:1 were used instead of the ethanol-water composite solvent in Example 3 to prepare the imidazole zinc complex (PXRD spectra shown in Figure 1). Figure 12 (As shown in the figure), and the resulting complexes were used for the hydrolysis catalytic reaction of p-nitrobenzene acetate, with reaction conditions basically the same as in Test Example 1, and the amount of complex used was 5 mg. The test comparison results are as follows. Figure 11As shown in the figure, compared to using toxic organic media (acetonitrile aqueous solution, DMF aqueous solution), using ethanol (EtOH:water volume ratio 1:1) as the reaction solvent not only achieves simultaneous crystallization and purification of the complex without complex post-treatment, but also further improves the catalytic effect of the obtained complex on the hydrolysis of p-nitrobenzene acetate. However, when methanol aqueous solution is used as the reaction solvent, the catalytic effect of the obtained complex on the hydrolysis of p-nitrobenzene acetate is significantly reduced compared to that in ethanol aqueous solution.
[0078] Example 8 This embodiment uses the method of Example 3 to prepare imidazole zinc complex crystals. The difference from Example 3 is that the volume ratio of water to anhydrous ethanol in this embodiment is 3:1. This embodiment yields colorless crystals A8 with a yield of 61%.
[0079] Elemental analysis of A8 yielded a result of C. 11 H 18 N4O4Zn: C, 39.41%; N, 16.82%; H, 5.21%, while the theoretically calculated values are: C, 39.36%; N, 16.69%; H, 5.41%, indicating that the elemental analysis results are consistent with the theoretical calculations. IR (KBrdisc, cm⁻¹) -1 PXRD detection, for example Figure 15 As shown.
[0080] Example 9 This embodiment uses the method of Example 3 to prepare imidazole zinc complex crystals. The difference from Example 3 is that the volume ratio of water to anhydrous ethanol in this embodiment is 1:3. This embodiment yields colorless crystals A9 with a yield of 62%.
[0081] Elemental analysis of A9 yielded a result of C. 11 H 18 N4O4Zn: C, 39.35%; N, 16.73%; H, 5.31%, while the theoretically calculated values are: C, 39.36%; N, 16.69%; H, 5.41%, indicating that the elemental analysis results are consistent with the theoretical calculations. PXRD detection is as follows... Figure 15 As shown.
[0082] Example 10 This embodiment uses the method of Example 3 to prepare imidazole zinc complex crystals. The difference from Example 3 is that the reaction temperature in this embodiment is 60°C. This embodiment yields colorless crystals A10 with a yield of 61%.
[0083] A10 was tested, and elemental analysis revealed that it contained C. 11 H 18N4O4Zn: C, 39.39%; N, 16.75%; H, 5.48%, while the theoretically calculated values are: C, 39.36%; N, 16.69%; H, 5.41%, indicating that the elemental analysis results are consistent with the theoretical calculations. PXRD detection is as follows... Figure 15 As shown.
[0084] Example 11 This embodiment uses the method of Example 3 to prepare imidazole zinc complex crystals. The difference from Example 3 is that the reaction temperature in this embodiment is 100°C. This embodiment yields colorless crystals A11 with a yield of 61%.
[0085] Elemental analysis of A11 yielded a result of C. 11 H 18 N4O4Zn: C, 39.34%; N, 16.71%; H, 5.45%, while the theoretically calculated values are: C, 39.36%; N, 16.69%; H, 5.41%, indicating that the elemental analysis results are consistent with the theoretical calculations. PXRD detection is as follows... Figure 15 As shown.
[0086] Example 12 This embodiment uses the method of Example 3 to prepare imidazole zinc complex crystals. The difference from Example 3 is that the reaction time in this embodiment is 72 hours. This embodiment yields colorless crystals A12 with a yield of 60%.
[0087] A12 was tested, and elemental analysis revealed that it was C. 11 H 18 N4O4Zn: C, 39.31%; N, 16.74%; H, 5.48%, while the theoretically calculated values are: C, 39.36%; N, 16.69%; H, 5.41%, indicating that the elemental analysis results are consistent with the theoretical calculations. PXRD detection is as follows... Figure 15 As shown.
[0088] Example 13 This embodiment uses the method of Example 3 to prepare imidazole zinc complex crystals. The difference from Example 3 is that the amount of water and anhydrous ethanol mixture used in this embodiment is 10 ml. This embodiment yields colorless crystals A13 with a yield of 60%.
[0089] Elemental analysis of A13 yielded a result of C. 11 H 18 N4O4Zn: C, 39.32%; N, 16.71%; H, 5.44%, while the theoretically calculated values are: C, 39.36%; N, 16.69%; H, 5.41%, indicating that the elemental analysis results are consistent with the theoretical calculations. PXRD detection is as follows... Figure 15 As shown.
[0090] The imidazole zinc complex crystals (Zn-LC) prepared in Examples 8-13 were used in the hydrolysis reaction of p-nitrobenzene acetate, and the specific operations are as follows: Add 4.9 mL of pH 7.4 PBS buffer to a 20 mL glass vial, then add 5.0 mg of imidazole zinc complex crystals to each vial. Stir magnetically in a 40 °C water bath for 10 minutes. Then add 100 μL of 0.5 mol / L... -1 An acetonitrile solution of p-nitrobenzene acetate was prepared. After 4 hours, 100 μL of the reaction solution was taken and diluted with 2.9 mL of PBS buffer (pH 7.4). The absorbance was then measured at 400 nm. The catalytic results of the hydrolysis of p-nitrobenzene acetate by the imidazole zinc complex crystals obtained in different examples are shown below. Figure 16 As shown.
Claims
1. A crystal of an imidazolium zinc complex, characterized in that, Its chemical formula is [Zn(L)](CH3CH2OH), where L represents L-carnosine. The chemical structure of the L-carnosine is shown below: One asymmetric structural unit of the imidazole zinc complex crystal contains a Zn(II), an L-carnosine ligand, and an ethanol molecule. The crystal structure of the imidazole zinc complex is monoclinic, space group P21; wherein, the coordination number of Zn(II) is 4, and each Zn(II) is coordinated with N or O atoms at four different positions on four L-carnosine ligands, wherein the N atoms are N1 and N4 on the imidazole ring and N3 on the amino group, and the O atoms are hydroxyl O1 on the carboxyl group.
2. The imidazole zinc complex crystal according to claim 1, characterized in that, The unit cell parameters of the imidazole zinc complex crystal are a=9.4155(2) Å, b=9.25933(13) Å, c=9.4236(2) Å; α=90°, β=118.740(3)°, γ=90°.
3. The imidazole zinc complex crystal according to claim 1, characterized in that, The XRD diffraction pattern of the imidazole zinc complex crystal shows a diffraction peak at 10.9 ± 0.2° for 2θ.
4. A method for preparing imidazole zinc complex crystals as described in any one of claims 1 to 3, characterized in that, include: In a reaction system containing water and anhydrous ethanol, L-carnosine is subjected to a coordination reaction with a soluble zinc salt; the reaction product is then cooled and filtered to obtain the imidazole zinc complex crystals.
5. The preparation method according to claim 4, characterized in that, Based on 1 mmol of L-carnosine, the total volume of the mixed solvent consisting of water and anhydrous ethanol is 10-20 mL; the volume ratio of water to anhydrous ethanol in the mixed solvent is 4:1 to 1:
4.
6. The preparation method according to claim 5, characterized in that, The molar ratio of zinc ions to L-carnosine in the soluble zinc salt is 8:1 to 1:
8.
7. The preparation method according to claim 6, characterized in that, The coordination reaction is carried out at a temperature of 60-100℃ for 24-72 hours.
8. The preparation method according to claim 7, characterized in that, After the coordination reaction is completed, the reaction mixture is cooled to 20~30℃ and filtered to obtain the imidazole zinc complex crystals.
9. The use of an imidazole zinc complex crystal as described in any one of claims 1 to 3 as an ester hydrolysis mimic enzyme.
10. The application according to claim 9, characterized in that, The imidazole zinc complex crystals were applied to the hydrolysis reaction of p-nitrobenzene acetate. Based on 1 mmol of p-nitrobenzene acetate, the amount of imidazole zinc complex crystals was 500-3000 mg. The hydrolysis reaction temperature was 30-90℃, and the pH of the reaction solution was 3-11.