Copper-based nano-enzyme, colorimetric sensing array and application of copper-based nano-enzyme and colorimetric sensing array
By utilizing a copper-based nanozyme colorimetric sensor array and taking advantage of the difference in catalytic activity between sheet-like and ring-shaped copper-based nanozymes, combined with multivariate analysis methods, the problem of efficient differentiation and quantitative detection of various phenolic acids in traditional Chinese medicine samples has been solved, achieving rapid, simple, and efficient phenolic acid detection.
Patent Information
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- Filing Date
- 2025-10-14
- Publication Date
- 2026-03-13
AI Technical Summary
Existing methods for detecting phenolic acids are costly, cumbersome, time-consuming, and have strict environmental requirements. They are difficult to efficiently distinguish and detect multiple phenolic acids, especially when complex components coexist in traditional Chinese medicine samples, where the selectivity is poor.
A copper-based nanozyme colorimetric sensor array was used. By constructing multiple sensing units, the catalytic activity differences between sheet-like and ring-shaped copper-based nanozymes at different wavelengths were utilized. Combined with linear discriminant analysis and hierarchical cluster analysis, efficient differentiation and quantitative detection of various phenolic acid substances were achieved.
It enables efficient and accurate differentiation and identification of various phenolic acids in complex Chinese medicine samples, reduces detection costs, improves selectivity and sensitivity, and provides a rapid and convenient detection solution.
Smart Images
Figure CN121648969A_ABST
Abstract
Description
Technical Field
[0001] This invention relates to the field of nanomaterial preparation technology, and more specifically, to a copper-based nanoenzyme, a colorimetric sensing array, and their applications. Background Technology
[0002] Phenolic acids are important active ingredients in traditional Chinese medicine. They are diverse and possess various medicinal values, including clearing heat and detoxifying, promoting blood circulation and removing blood stasis, and regulating immunity, with broad application prospects. Common medicinal phenolic acids such as chlorogenic acid, ferulic acid, caffeic acid, and protocatechuic acid generally exhibit antioxidant, antitumor, antibacterial, and anti-inflammatory pharmacological activities, and their appropriate intake has a positive impact on human health.
[0003] Currently, the main methods for detecting phenolic acids include high-performance liquid chromatography (HPLC), gas chromatography (GC), electrochemical methods, and fluorescence methods. While these methods offer good detection results, they generally suffer from limitations such as high cost, cumbersome operation, long processing times, and stringent environmental requirements, restricting their widespread application. In contrast, colorimetric methods show great potential due to their low cost, ease of operation, speed, and sensitivity. However, traditional Chinese medicine samples are complex, often containing multiple phenolic acids and other components. Therefore, there is an urgent need to develop a method that can simultaneously and efficiently distinguish and detect multiple phenolic acids.
[0004] Encouragingly, the emergence of colorimetric sensor array technology offers an effective solution to this problem. This technology, by constructing multiple sensing units or detection channels, overcomes the limitations of traditional "lock-and-key" single-identification mechanisms. When interacting with a specific target object, different channels in the array can generate distinct colorimetric responses, thus forming a unique "fingerprint" recognition pattern, providing a new approach for the simultaneous detection and differentiation of multiple components.
[0005] Based on this, the present invention develops a novel colorimetric sensor array based on copper-based metal-organic framework nanozymes, providing a new method for the efficient and rapid detection of phenolic acids in traditional Chinese medicine. Summary of the Invention
[0006] To overcome the shortcomings of the prior art, the present invention provides a copper-based nanozyme, a colorimetric sensing array, and its applications.
[0007] The first aspect of this invention provides a method for preparing copper-based nanozymes, the method specifically comprising the following steps: S1. Dissolve 2-methylimidazole in ultrapure water to obtain solution A, and dissolve Cu(NO3)2·3H2O in ultrapure water to obtain solution B; S2. Add solution B to solution A, and then proceed with stirring, settling, centrifugation, and drying to obtain copper-based nanozymes.
[0008] Compared with existing technologies, this preparation method has the advantages of simplicity, efficiency, mild conditions, and ease of scalability. The reaction can be completed through simple solution mixing and room temperature standing, without the need for complex equipment or high-temperature and high-pressure environments, significantly reducing the production threshold and energy consumption. The raw materials used are inexpensive and readily available, the process is streamlined, involving only routine operations such as stirring, standing, and centrifugation, and has high reproducibility, making it suitable for large-scale preparation. This method provides a stable and reliable synthetic basis for the widespread application of copper-based nanozymes.
[0009] In one possible implementation, in step S2, the volume ratio of solution A to solution B is 7:1; The stirring time is 30-50 minutes, and the settling time is 24-25 hours. The parameters for the centrifugation process are as follows: rotation speed of 10,000 rpm and duration of 5 min. The drying parameters are as follows: temperature 60-70℃, time 12-14h.
[0010] Compared with existing technologies, this invention ensures a high degree of controllability and reproducibility of the synthesis process through precisely optimized process parameters (including solution volume ratio, processing time, and temperature). Mild reaction and post-treatment conditions, while maintaining consistent morphology and performance of the nanozymes, significantly reduce energy consumption and operational complexity, giving the method excellent scalability and laying a solid foundation for stable, large-scale industrial production.
[0011] The second objective of this invention is to provide a sheet-like copper-based nanozyme prepared by the above-described preparation method, wherein in step S1, the molar ratio of Cu(NO3)2·3H2O to 2-methylimidazole is 1:55.3.
[0012] Compared with existing technologies, this invention, by employing an extremely high ligand-to-metal salt molar ratio (1:55.3), greatly promotes two-dimensional crystal growth, thereby advantageously obtaining sheet-like nanozymes with uniform morphology and regular structure. This specific ratio is key to forming a unique sheet-like structure, effectively avoiding the generation of other competing morphologies and ensuring high consistency and excellent performance reproducibility between product batches.
[0013] The third objective of this invention is to provide a circular copper-based nanozyme prepared by the above-described method, wherein in step S1, the molar ratio of Cu(NO3)2·3H2O to 2-methylimidazole is 1:6.176.
[0014] Compared with existing technologies, this invention specifically guides the anisotropic growth of crystals and their eventual self-assembly into a ring structure by precisely controlling the molar ratio of metal salt to organic ligand (1:6.176). This key ratio is the decisive factor in achieving the ring morphology, ensuring the high specificity and reproducibility of the synthesis process, effectively avoiding the generation of impurity phases with other morphologies, and providing a reliable guarantee for obtaining nanozyme products with novel structures and unique properties.
[0015] A fourth objective of this invention is to provide a colorimetric sensing array for distinguishing or detecting phenolic acids, wherein the colorimetric sensing array is constructed based on sheet-like copper-based nanozymes and ring-like copper-based nanozymes; the colorimetric sensing array includes four detection channels, namely: the absorbance value of oxidized TMB at 370 nm when sheet-like copper-based nanozymes are used as catalysts, the absorbance value of oxidized TMB at 652 nm when sheet-like copper-based nanozymes are used as catalysts, the absorbance value of oxidized TMB at 370 nm when ring-like copper-based nanozymes are used as catalysts, and the absorbance value of oxidized TMB at 652 nm when ring-like copper-based nanozymes are used as catalysts.
[0016] This invention creatively utilizes the difference in catalytic activity between two structurally distinct nanozymes (sheet-like and ring-like) to synergistically oxidize TMB at two characteristic wavelengths, constructing a four-dimensional sensing array. This design greatly enriches the dimensions of detection information, generating a unique "fingerprint" response spectrum for each phenolic acid substance, thereby achieving efficient and accurate differentiation and identification of multiple structurally similar phenolic acids in complex mixtures, solving the problem of poor selectivity in traditional single detection methods.
[0017] In one possible implementation, the phenolic acid is selected from at least two of ferulic acid, protocatechuic acid, p-hydroxyphenylacetic acid, gallic acid, vanillic acid, chlorogenic acid, and caffeic acid.
[0018] Compared with existing technologies, this invention can efficiently distinguish and detect multiple structurally similar phenolic acids (such as ferulic acid and chlorogenic acid). By utilizing the differential inhibition or enhancement effects of different phenolic acids on nanozyme catalytic reactions, this array can generate a unique "fingerprint" response spectrum for each substance, thereby effectively solving the technical problem of inaccurate identification and recognition when multiple phenolic acids coexist in complex samples, and significantly improving the selectivity and reliability of detection.
[0019] The fifth objective of this invention is to provide a method for distinguishing phenolic acids using a colorimetric sensor array, comprising the following steps: A1. In the reaction system, the phenolic acid sample to be tested was mixed with the TMB-H2O2 colorimetric system and the sheet-like copper-based nanozyme and then reacted at room temperature for 10 min. The absorbance at 375 nm and 652 nm was then measured by a UV spectrophotometer. The phenolic acid sample to be tested was mixed with the TMB-H2O2 colorimetric system and the circular copper-based nanozyme, and then reacted at room temperature for 10 min. The absorbance at 375 nm and 652 nm was then measured by a UV spectrophotometer. A2. Repeat step A1 5 times for all phenolic acid samples to be tested to obtain a data matrix. Use linear discriminant analysis and / or hierarchical cluster analysis to process the data matrix to distinguish the phenolic acid samples to be tested.
[0020] Compared with existing technologies, this invention combines complex chemical responses with powerful pattern recognition algorithms. Stable and reliable four-dimensional signals are obtained through standardized reaction procedures, and statistical significance is ensured through multiple repetitions. Finally, multivariate analysis methods (LDA / HCA) are used to intelligently analyze the signal patterns, thereby transforming subtle response differences into intuitive and accurate classification results, achieving efficient and reliable differentiation and identification of complex phenolic acid mixtures.
[0021] In one possible implementation, in step A1, the concentration of the phenolic acid sample to be tested in the reaction system is 30 μM, and the concentration of either the sheet-like copper-based nanozyme or the ring-like copper-based nanozyme is 0.5 mg / ml.
[0022] The sixth objective of this invention is to provide a method for quantitative detection of phenolic acids in traditional Chinese medicine using a colorimetric sensor array, comprising the following steps: B1. Plotting the standard working curve: Prepare different types of phenolic acid solutions with concentrations of 5µM, 10µM, 20µM, 30µM, 40µM and 50µM, and divide them into two portions; One portion of each phenolic acid solution was mixed with the TMB-H2O2 colorimetric system and the sheet-like copper-based nanozyme and reacted at room temperature for 10 min. The absorbance at 375 nm and 652 nm was then measured using a UV spectrophotometer. Another portion of each phenolic acid solution was mixed with the TMB-H2O2 colorimetric system and the circular copper-based nanozyme and reacted at room temperature for 10 min. The absorbance at 375 nm and 652 nm was then measured using a UV spectrophotometer. Linear discriminant analysis was performed on the obtained absorbance data. The concentration of each phenolic acid solution was used as the x-axis and the first principal factor score obtained in the linear discriminant analysis was used as the y-axis. Linear fitting was performed to obtain the standard working curve equation for each phenolic acid. B2. Determination of unknown samples: Weigh the Chinese medicine sample, soak it in deionized water and reflux at 100℃ for 2 hours. After filtration, centrifugation and dilution, the sample to be tested is obtained. Mix one part of the sample to be tested with the TMB-H2O2 colorimetric system and the sheet copper-based nanozyme and react at room temperature for 10 minutes. Then, use a UV spectrophotometer to measure the absorbance at 375nm and 652nm respectively. Another portion of the sample to be tested was mixed with the TMB-H2O2 colorimetric system and the circular copper-based nanozyme and reacted at room temperature for 10 min. Then, the absorbance at 375 nm and 652 nm was measured using a UV spectrophotometer. The obtained absorbance data is subjected to linear discriminant analysis to obtain its first principal factor score, which is then substituted into the corresponding standard working curve equation in step B1 to calculate the concentration of the phenolic acid to be tested.
[0023] Compared with existing technologies, this invention utilizes the differential inhibition effect of different phenolic acids on the TMB-H2O2 reaction catalyzed by two nanozymes to generate a unique four-dimensional absorbance response. Linear discriminant analysis (LDA) is then used to reduce the dimensionality of the complex response signal, transforming it into a single principal factor score linearly correlated with concentration, thereby establishing a precise quantitative model. Its core advantage lies in combining the high-throughput and high-selectivity recognition capabilities of a sensor array with the powerful resolving power of chemometrics, achieving specific and highly sensitive quantitative detection of multiple structurally similar phenolic acids in complex traditional Chinese medicine matrices, thus solving the problems of poor selectivity and weak anti-interference ability of traditional methods.
[0024] In step B1, the standard working curve equation, linear range, and detection limit of phenolic acid are obtained as follows: Ferulic acid: y = -1.0654x + 27.5253, R 2 = 0.999, linear range is 5-50µM, detection limit = 1.4µM; Protocatechuic acid: y = -0.76282x + 20.78415, R 2 = 0.992, linear range is 5-50µM, detection limit = 2.2µM; p-Hydroxyphenylacetic acid: y = -0.41197x + 10.57688, R 2 = 0.996, linear range is 5-50µM, detection limit = 4.1µM; Gallic acid: y = -1.24524x + 32.00121, R 2 = 0.990, linear range is 5-50µM, detection limit = 0.82µM; Vanillic acid: y = -0.445x + 11.13949, R 2= 0.990, linear range is 5-50µM, detection limit = 3.7µM; Chlorogenic acid: y = -1.66329x + 43.15933, R 2 = 0.996, linear range is 5-50µM, detection limit = 0.56µM; Caffeic acid: y = -0.78288x + 20.27506, R 2 = 0.990, linear range is 5-50µM, detection limit = 1.9µM.
[0025] Where x is the phenolic acid concentration, y is the first principal factor score obtained in the LDA analysis, and R is the correlation coefficient. Attached Figure Description
[0026] Figure 1 This is a scanning electron microscope image of the sheet-like copper-based nanozyme prepared in Example 1; Figure 2 This is a scanning electron microscope image of the ring-shaped copper-based nanozyme prepared in Example 2; Figure 3 The UV-Vis spectra of the catalytic activity of the sheet-like copper-based nanozyme prepared in Example 1 and the ring-like copper-based nanozyme prepared in Example 2 are shown. Figure 4 Differentiation diagrams for seven phenolic acids at 30 μM (A: LDA diagram, B: HCA diagram, C: heatmap, D: radar diagram). Figure 5 This is a standard curve for the detection of caffeic acid; Figure 6 This is a standard curve for the detection of chlorogenic acid; Figure 7 This is a standard curve for the detection of ferulic acid. Figure 8 A standard curve for the detection of gallic acid; Figure 9 A standard curve for the detection of protocatechuic acid; Figure 10 This is a standard curve for the detection of p-hydroxyphenylacetic acid; Figure 11 This is a standard curve for the detection of vanillic acid; Figure 12 This is an LDA diagram showing the differentiation of actual samples in the application examples; Figure 13 The HCA diagram is used to distinguish actual samples in the application examples. Detailed Implementation
[0027] To make the above-mentioned objects, features, and advantages of the present invention more apparent and understandable, specific embodiments of the present invention are described in detail below. It should be noted that the following embodiments are only used to illustrate the implementation methods and typical parameters of the present invention, and are not intended to limit the parameter range described in the present invention. Reasonable variations derived therefrom are still within the protection scope of the claims of the present invention.
[0028] It should be noted that the endpoints and any values of the ranges disclosed herein are not limited to the precise ranges or values, and these ranges or values should be understood to include values close to these ranges or values. For numerical ranges, the endpoint values of the various ranges, the endpoint values of the various ranges and individual point values, and individual point values can be combined with each other to obtain one or more new numerical ranges, which should be considered as specifically disclosed herein.
[0029] Unless otherwise defined, all terms, symbols, and other scientific terms used herein are intended to have the same meaning as commonly understood by one of ordinary skill in the art to which this invention pertains. In some instances, terms having a conventional meaning are defined herein for clarification or ease of reference, and such definitions should not be construed as indicating a significant difference from conventional understanding in the art. The technical methods described or referenced herein are generally well understood by those skilled in the art and employed by conventional methods. Unless otherwise stated, the use of commercially available kits, reagents, and instruments shall be performed according to the manufacturer's instructions and parameters.
[0030] Example 1 This embodiment provides a sheet-like copper-based nanozyme (Cu-MI NPs), prepared by the following method: 27.65 mmol of 2-methylimidazole was dissolved in 35 mL of ultrapure water to form solution A; 0.5 mmol of Cu(NO3)2·3H2O was dissolved in 5 mL of ultrapure water to form solution B; solution B was then added to solution A, stirred for 30 min, and then allowed to stand for 24 h. Finally, the solution was centrifuged (5 min, 10000 rpm), washed three times each with ultrapure water and ethanol, and dried at 60 °C for 12 h to obtain the sheet-like copper-based nanozyme material. The scanning electron microscope image is shown below. Figure 1 ; 10 mg of sheet-like copper-based nanozyme material was dispersed in 20 mL of deionized water to obtain a 0.5 mg / mL sheet-like copper-based nanozyme aqueous solution for later use.
[0031] Example 2 This embodiment provides a ring-shaped copper-based nanozyme material (Cu-PI NPs), prepared by the following method: 3.088 mmol of 2-propylimidazole was dissolved in 35 mL of ultrapure water to form solution A; 0.5 mmol of Cu(NO3)2•3H2O was dissolved in 5 mL of ultrapure water to form solution B; solution B was then added to solution A, stirred for 30 min, and then allowed to stand for 24 h. Finally, the solution was centrifuged (5 min, 10000 rpm), washed three times each with ultrapure water and ethanol, and dried at 60 °C for 12 h to obtain the ring-shaped copper-based nanozyme material. The scanning electron microscope image is shown below. Figure 2 ; 10 mg of circular copper-based nanozyme material was dispersed in 20 mL of deionized water to obtain an aqueous solution of 0.5 mg / mL circular copper-based nanozyme.
[0032] The catalytic activity of the sheet-like copper-based nanozyme prepared in Example 1 and the ring-like copper-based nanozyme prepared in Example 2 was detected by ultraviolet-visible spectrophotometry. Figure 3 The UV-Vis spectra of the catalytic activity of the sheet-like copper-based nanozyme prepared in Example 1 and the ring-like copper-based nanozyme prepared in Example 2 are shown.
[0033] Example 3 This embodiment provides a colorimetric sensing array for distinguishing or detecting phenolic acid substances. The colorimetric sensing array is constructed based on sheet-like copper-based nanozymes and ring-like copper-based nanozymes. The colorimetric sensing array includes four detection channels, namely: the absorbance value of oxidized TMB at 370 nm when sheet-like copper-based nanozymes are used as catalysts, the absorbance value of oxidized TMB at 652 nm when sheet-like copper-based nanozymes are used as catalysts, the absorbance value of oxidized TMB at 370 nm when ring-like copper-based nanozymes are used as catalysts, and the absorbance value of oxidized TMB at 652 nm when ring-like copper-based nanozymes are used as catalysts.
[0034] It is worth mentioning that the above-mentioned phenolic acids are selected from at least two of ferulic acid, protocatechuic acid, p-hydroxyphenylacetic acid, gallic acid, vanillic acid, chlorogenic acid and caffeic acid.
[0035] Example 4 This embodiment provides a specific application of the colorimetric sensor array in Example 3, applying it to a method for distinguishing phenolic acids, including the following steps: Add 200 µL of 5 mM TMB solution, 200 µL of 50 mM H2O2 solution, 200 µL of 100 mM NaAc-HAc (pH=6) buffer solution, 200 µL of 0.5 mg / mL flaky copper-based nanozyme aqueous solution, 30 µM of different types of phenolic acid solutions, and deionized water to a sample tube to make a reaction system of 2 mL. Mix the solutions thoroughly and react at room temperature for 10 minutes. Detect and record the absorbance at 370 nm and 652 nm using a UV-Vis spectrophotometer. Measure the absorbance data when the cyclic copper-based nanozyme aqueous solution is used as the enzyme catalyst using the same procedure. This yields four colorimetric signals for different types of phenolic acids at the same concentration. Each phenolic acid is repeated 5 times to generate a data matrix (4 channels × 7 types of phenolic acids × 5 repetitions). Finally, perform LDA and HCA analysis on the data matrix, as shown below. Figure 4 As shown.
[0036] Example 5 This embodiment provides a specific application of the colorimetric sensor array in Example 3, applying it to a method for detecting phenolic acids, including the following steps: S1. Plotting the working curve: Add 200 µL of 5 mM TMB solution, 200 µL of 50 mM H2O2 solution, 200 µL of 100 mM NaAc-HAc (pH=6) buffer solution, 200 µL of 0.5 mg / mL flaky copper-based nanozyme aqueous solution, phenolic acid solution (using ferulic acid as an example), and deionized water to the sample tube to make a reaction system of 2 mL (the concentrations of ferulic acid in the reaction system are 5, 10, 20, 30, 40, and 50 µM, respectively). Then mix the above solutions thoroughly and react at room temperature for 10 minutes. Detect and record the absorbance at 370 nm and 652 nm using a UV-Vis spectrophotometer. Measure the absorbance data when the cyclic copper-based nanozyme aqueous solution is used as the enzyme catalyst using the same procedure.
[0037] Plotting the concentration of ferulic acid in the reaction system on the x-axis and the value of Factor 1 (>90%) in the LDA analysis on the y-axis, the working curve equation is obtained as: y = -1.0654x + 27.5253 (R 2 = 0.999), the linear detection range is 5-50 µM, and the detection limit is 1.4 μM. Figure 7 As shown.
[0038] The working curves for the remaining six phenolic acids were determined using the same procedure, and the results were as follows: Protocatechuic acid: Working curve equation y = -0.76282x + 20.78415 (R 2= 0.992), the linear detection range is 5-50 µM, and the detection limit is 2.2 μM. Figure 9 As shown; p-Hydroxyphenylacetic acid: Working curve equation y = -0.41197x + 10.57688 (R 2 = 0.996), the linear detection range is 5-50 µM, and the detection limit is 4.1 μM. Figure 10 As shown; Gallic acid: Working curve equation y = -1.24524x + 32.00121 (R 2 = 0.990), the linear detection range is 5-50 µM, and the detection limit is 0.82 μM. Figure 8 As shown; Vanillic acid: Working curve equation y = -0.445x + 11.13949 (R 2 = 0.990), the linear detection range is 5-50 µM, and the detection limit is 3.7 μM. Figure 11 As shown; Chlorogenic acid: Working curve equation y = -1.66329x + 43.15933 (R 2 = 0.996), the linear detection range is 5-50 µM, and the detection limit is 0.56 μM. Figure 6 As shown; Caffeic acid: Working curve equation y = -0.78288x + 20.27506 (R 2 = 0.990), the linear detection range is 5-50 µM, and the detection limit is 1.9 μM. Figure 5 As shown.
[0039] Application Examples Forsythia and honeysuckle were weighed in mass ratios of 4 g:0 g, 3 g:1 g, 2 g:2 g, 1 g:3 g, and 0 g:4 g, and were soaked in 20 ml of deionized water, respectively, and heated under reflux at 100℃ for 2 h. After heating, the samples were filtered and centrifuged to obtain five supernatants. These supernatants were then diluted 100 times to obtain five dilutions. Finally, LDA and HCA analyses were performed on these five dilution samples according to the above experimental procedure. Figure 12 and Figure 13 As shown.
[0040] While the disclosure is as stated above, its scope of protection is not limited thereto. Those skilled in the art can make various changes and modifications without departing from the spirit and scope of this disclosure, and all such changes and modifications will fall within the protection scope of this invention.
Claims
1. A method for preparing a copper-based nanozyme, characterized in that, The preparation method specifically includes the following steps: S1. Dissolve 2-methylimidazole in ultrapure water to obtain solution A, and dissolve Cu(NO3)2·3H2O in ultrapure water to obtain solution B; S2. Add solution B to solution A, and then proceed with stirring, settling, centrifugation, and drying to obtain copper-based nanozymes.
2. A preparation method as described in claim 1, characterized in that, In step S2, the volume ratio of solution A to solution B is 7:1; The stirring time is 30-50 minutes, and the settling time is 24-25 hours. The parameters for the centrifugation process are as follows: rotation speed of 10,000 rpm and duration of 5 min. The drying parameters are as follows: temperature 60-70℃, time 12-14h.
3. A sheet-like copper-based nanozyme, prepared by the method described in claim 2, characterized in that, In step S1, the molar ratio of Cu(NO3)2·3H2O to 2-methylimidazole is 1:55.
3.
4. A circular copper-based nanozyme, prepared by the method described in claim 2, characterized in that, In step S1, the molar ratio of Cu(NO3)2·3H2O to 2-methylimidazole is 1:6.
176.
5. A colorimetric sensing array, characterized in that, The colorimetric sensing array is used to distinguish or detect phenolic acid substances, and the colorimetric sensing array is constructed based on the sheet-like copper-based nanozyme of claim 3 and the ring-like copper-based nanozyme of claim 4; the colorimetric sensing array includes four detection channels, namely: the absorbance value of oxidized TMB at 370 nm when the sheet-like copper-based nanozyme is used as a catalyst, the absorbance value of oxidized TMB at 652 nm when the sheet-like copper-based nanozyme is used as a catalyst, the absorbance value of oxidized TMB at 370 nm when the ring-like copper-based nanozyme is used as a catalyst, and the absorbance value of oxidized TMB at 652 nm when the ring-like copper-based nanozyme is used as a catalyst.
6. The colorimetric sensor array as described in claim 5, characterized in that, The phenolic acid is selected from at least two of ferulic acid, protocatechuic acid, p-hydroxyphenylacetic acid, gallic acid, vanillic acid, chlorogenic acid, and caffeic acid.
7. A method for distinguishing phenolic acids using the colorimetric sensor array of claim 6, characterized in that, Includes the following steps: A1. In the reaction system, the phenolic acid sample to be tested was mixed with the TMB-H2O2 colorimetric system and the sheet-like copper-based nanozyme and then reacted at room temperature for 10 min. The absorbance at 375 nm and 652 nm was then measured by a UV spectrophotometer. The phenolic acid sample to be tested was mixed with the TMB-H2O2 colorimetric system and the circular copper-based nanozyme, and then reacted at room temperature for 10 min. The absorbance at 375 nm and 652 nm was then measured by a UV spectrophotometer. A2. Repeat step A1 5 times for all phenolic acid samples to be tested to obtain a data matrix. Use linear discriminant analysis and / or hierarchical cluster analysis to process the data matrix to distinguish the phenolic acid samples to be tested.
8. The method as described in claim 7, characterized in that, In step A1, the concentration of the phenolic acid sample to be tested in the reaction system is 30 μM, and the concentration of either the sheet-like copper-based nanozyme or the ring-like copper-based nanozyme is 0.5 mg / ml.
9. A method for quantitative detection of phenolic acids in traditional Chinese medicine using the colorimetric sensor array described in claim 6, characterized in that, Includes the following steps: B1. Plotting the standard working curve: Prepare different types of phenolic acid solutions with concentrations of 5µM, 10µM, 20µM, 30µM, 40µM and 50µM, and divide them into two portions; One portion of each phenolic acid solution was mixed with the TMB-H2O2 colorimetric system and the sheet-like copper-based nanozyme and reacted at room temperature for 10 min. The absorbance at 375 nm and 652 nm was then measured using a UV spectrophotometer. Another portion of each phenolic acid solution was mixed with the TMB-H2O2 colorimetric system and the circular copper-based nanozyme and reacted at room temperature for 10 min. The absorbance at 375 nm and 652 nm was then measured using a UV spectrophotometer. Linear discriminant analysis was performed on the obtained absorbance data. The concentration of each phenolic acid solution was used as the x-axis and the first principal factor score obtained in the linear discriminant analysis was used as the y-axis. Linear fitting was performed to obtain the standard working curve equation for each phenolic acid. B2. Determination of unknown samples: Weigh the Chinese medicine sample, soak it in deionized water and reflux at 100℃ for 2 hours. After filtration, centrifugation and dilution, the sample to be tested is obtained. Mix one part of the sample to be tested with the TMB-H2O2 colorimetric system and the sheet copper-based nanozyme and react at room temperature for 10 minutes. Then, use a UV spectrophotometer to measure the absorbance at 375nm and 652nm respectively. Another portion of the sample to be tested was mixed with the TMB-H2O2 colorimetric system and the circular copper-based nanozyme and reacted at room temperature for 10 min. Then, the absorbance at 375 nm and 652 nm was measured using a UV spectrophotometer. The obtained absorbance data is subjected to linear discriminant analysis to obtain its first principal factor score, which is then substituted into the corresponding standard working curve equation in step B1 to calculate the concentration of the phenolic acid to be tested.
10. The method according to claim 9, characterized in that, In step B1, the standard working curve equation, linear range, and detection limit of phenolic acid are obtained as follows: Ferulic acid: y = -1.0654x + 27.5253, R 2 = 0.999, linear range is 5-50µM, detection limit = 1.4µM; Protocatechuic acid: y = -0.76282x + 20.78415, R 2 = 0.992, linear range is 5-50µM, detection limit = 2.2µM; p-Hydroxyphenylacetic acid: y = -0.41197x + 10.57688, R 2 = 0.996, linear range is 5-50µM, detection limit = 4.1µM; Gallic acid: y = -1.24524x + 32.00121, R 2 = 0.990, linear range is 5-50µM, detection limit = 0.82µM; Vanillic acid: y = -0.445x + 11.13949, R 2 = 0.990, linear range is 5-50µM, detection limit = 3.7µM; Chlorogenic acid: y = -1.66329x + 43.15933, R 2 = 0.996, linear range is 5-50µM, detection limit = 0.56µM; Caffeic acid: y = -0.78288x + 20.27506, R 2 = 0.990, linear range is 5-50µM, detection limit = 1.9µM; Where x is the phenolic acid concentration, y is the first principal factor score obtained in the LDA analysis, and R is the correlation coefficient.