Corilagin molecular imprinting electrochemical sensor as well as preparation method and application thereof

Abstract

The invention discloses a corilagin molecular imprinting electrochemical sensor as well as a preparation method and application thereof. The sensor comprises a glassy carbon electrode substrate, a boric acid functionalized Mxene conductive affinity composite material layer covering the glassy carbon electrode substrate and a corilagin molecularly imprinted polymer layer located on the surface of the boric acid functionalized Mxene conductive affinity composite material layer. The preparation method comprises the following steps: preparing a boric acid functionalized Mxene composite material and preparing a modification liquid; dispensing the modified liquid on the pretreated glassy carbon electrode to prepare a modified electrode; immobilizing corilagin template molecules through boric acid affinity reaction; carrying out surface molecular imprinting polymerization; and eluting the template molecules to form an imprinting cavity, thereby obtaining the sensor. The sensor disclosed by the invention has excellent conductivity, high selectivity and high sensitivity by constructing an integrated structure of a conductive nano skeleton and a dual recognition layer, and is wide in corilagin detection linear range, low in detection limit and good in stability.

Description

Technical Field

[0001] This invention relates to the fields of analytical chemistry and sensing technology, specifically to a Corilagin molecularly imprinted electrochemical sensor, its preparation method, and its application. Background Technology

[0002] Corilagin is a key polyphenol quality marker (Q-Marker) for many traditional Chinese medicinal materials, such as Phyllanthus emblica and Phyllanthus urinaria. It possesses various pharmacological activities, including antioxidant, anti-inflammatory, hepatoprotective, and lipid-lowering activities. Its content is closely related to the authenticity and clinical efficacy of the medicinal materials. Currently, high-performance liquid chromatography (HPLC), liquid chromatography-tandem mass spectrometry (LC-MS / MS), capillary electrophoresis (CE), quantitative nuclear magnetic resonance (qNMR), or ultraviolet spectrophotometry (UV) are commonly used for corilagin detection both domestically and internationally. While these methods meet laboratory sensitivity requirements, the high cost of instrument purchase and maintenance makes them difficult to promote in grassroots or production areas; sample pretreatment is complex; and chromatographic systems are prone to matrix effects on tannins, leading to peak tailing and quantitative deviations. Therefore, developing a simple, low-cost, highly sensitive, and highly selective corilagin detection technology is crucial for the rapid evaluation of the quality of traditional Chinese medicinal materials.

[0003] Electrochemical sensors have shown great potential in the field of rapid detection due to their advantages such as low cost, simple operation, fast response, and easy miniaturization. Molecular imprinting technology can construct recognition sites with predetermined selectivity for target molecules and has been widely used in the field of sensing. However, traditional molecular imprinted electrochemical sensors usually face the following challenges: (1) poor conductivity of the imprinted polymer layer, resulting in weak electrochemical signals and low sensitivity; (2) the imprinted cavity is deeply buried inside the polymer, resulting in large mass transfer resistance and slow response speed; (3) in complex sample matrices, selectivity is easily interfered with by structural analogs (such as ellagic acid, gallic acid, etc.).

[0004] In recent years, MXene, a two-dimensional material, has been considered an ideal material for constructing high-performance electrochemical sensing interfaces due to its excellent metallic conductivity, abundant surface functional groups, and good biocompatibility. However, there are currently no reports on its use in the fabrication of Corilagin molecularly imprinted electrochemical sensors. Summary of the Invention

[0005] The purpose of this invention is to overcome the shortcomings of existing detection technologies and provide a novel, easy-to-prepare, low-cost, highly sensitive, selective, and stable corilagin molecularly imprinted electrochemical sensor. The invention also describes its preparation method and its application in the rapid detection of corilagin in complex matrices of traditional Chinese medicinal materials. This invention aims to solve the problems of traditional chromatographic methods, such as strong instrument dependence and complex operation, as well as the poor conductivity and insufficient selectivity of traditional molecularly imprinted sensors.

[0006] To achieve the above objectives, the present invention adopts the following technical solution: In a first aspect, the present invention provides a Corilagin molecularly imprinted electrochemical sensor, comprising: Glassy carbon electrode substrate; A boric acid-functionalized Mxene conductive affinity composite material layer covering the surface of the glassy carbon electrode; And a Corilagin molecularly imprinted polymer layer located on the surface of the conductive affinity composite material layer; wherein the imprinted polymer layer contains imprinted cavities that match the structure of the Corilagin molecules.

[0007] Preferably, the boric acid-functionalized Mxene conductive affinity composite material is formed by modifying Ti3C2T with 4-formylphenylboronic acid via an aminosilane coupling agent. x It is prepared by surface preparation of Mxene.

[0008] Secondly, the present invention provides a method for preparing the above-mentioned Corilagin molecularly imprinted electrochemical sensor, comprising the following steps: (1) Prepare boric acid-functionalized Mxene conductive affinity composite material and disperse it in chitosan acetic acid solution to prepare a modified solution; (2) The modification solution is drop-coated onto the surface of the pretreated glassy carbon electrode and dried to obtain the Mxene-BA modified electrode; (3) Corilagin template molecules are fixed on the surface of the Mxene-BA modified electrode by boric acid affinity reaction; (4) Molecular imprinting polymerization reaction is carried out on the electrode surface on which template molecules are fixed; (5) Elute the template molecules to obtain the Corilagin molecularly imprinted electrochemical sensor.

[0009] Preferably, in step (1), the preparation of the boric acid-functionalized Mxene conductive affinity composite material includes: Multilayer Ti3C2T x Mxene reacts with 3-aminopropyltriethoxysilane in an alkaline ethanol solution to obtain aminated Mxene; wherein the mass-to-volume ratio of Mxene to 3-aminopropyltriethoxysilane is (90-110) mg: (380-420) μL; the pH of the reaction system is 9-11, the reaction temperature is 20-30℃, and the reaction time is 20-28 hours.

[0010] The aminoated Mxene was subjected to a reductive amination reaction with 4-formylphenylboronic acid and sodium cyanoborohydride in methanol to obtain the boric acid-functionalized Mxene; wherein the mass ratio of the aminoated Mxene to 4-formylphenylboronic acid was 1:(1.8-2.2); the reaction temperature was 20-30℃ and the reaction time was 20-28 hours.

[0011] Preferably, in step (1), the concentration of Mxene-BA in the modified solution is 2-6 mg / mL; the mass fraction of chitosan in the chitosan acetic acid solution is 0.8%-1.2%, and the solvent is an aqueous acetic acid solution with a concentration of 0.8%-1.2%.

[0012] Preferably, the pretreatment of the glassy carbon electrode in step (2) includes: mechanically polishing the surface of the glassy carbon electrode to a mirror finish using alumina polishing powder of different particle sizes in sequence; then performing ultrasonic cleaning; and finally performing cyclic voltammetric scanning activation in an electrolyte containing potassium ferrocyanide / potassium ferricyanide until the redox peak potential difference stabilizes within a set threshold, and then drying it with inert gas for later use.

[0013] More preferably, in step (2), the specific conditions for the pretreatment are as follows: polishing is performed sequentially using alumina polishing powder with particle sizes of 1.0 μm, 0.3 μm and 0.05 μm; ultrasonic cleaning is performed using ultrapure water for 2-5 minutes; cyclic voltammetric activation is performed in a solution containing 2-10 mM potassium ferrocyanide / potassium ferrocyanide and 0.05-0.2 M supporting electrolyte, with a scan rate of 20-100 mV / s and a redox peak potential difference threshold set to less than 100 mV.

[0014] Preferably, in step (3), the conditions for immobilizing the Corilagin template molecule include: the concentration of the template molecule solution is 10-50 μg / mL, the solvent is a phosphate buffer solution with pH 8.0-9.0; the incubation temperature is 20-30℃, and the incubation time is 2-4 hours.

[0015] Preferably, in step (4), the molecularly imprinted polymerization reaction uses acrylamide as the monomer, N,N′-methylenebisacrylamide as the crosslinking agent, and ammonium persulfate as the initiator. Specifically, the concentration of acrylamide in the prepolymer solution is 200-300 mmol / L, the molar ratio of N,N′-methylenebisacrylamide to acrylamide is 1:10 to 1:20, the concentration of the ammonium persulfate initiator is 8-12 mmol / L, the polymerization reaction temperature is 20-30℃, and the reaction time is 1.5-3 hours.

[0016] Preferably, in step (5), the eluting template molecules are ultrasonically eluted using an organic solvent mixture, wherein the organic solvent mixture is a mixture of methanol and acetic acid with a volume ratio of (8.5:1) to (9.5:1); and the ultrasonic elution time is 15-25 minutes. In a third aspect, the present invention provides the application of the above-described corilagin molecularly imprinted electrochemical sensor or the sensor prepared by the above method in the detection of corilagin.

[0017] Fourthly, the present invention provides a method for detecting the content of corilagin in a sample, characterized by using the aforementioned corilagin molecularly imprinted electrochemical sensor, comprising the following steps: placing the sensor in a test solution containing corilagin for re-adsorption; detecting the current response signal of the sensor using an electrochemical method; and calculating the content of corilagin in the test sample based on a pre-established quantitative relationship between the sensor response signal and the corilagin concentration. Preferably, the test sample is a traditional Chinese medicine or its extract, wherein the traditional Chinese medicine is Phyllanthus emblica or Phyllanthus urinaria.

[0018] Compared with the prior art, the present invention has the following beneficial effects: This invention utilizes boric acid-functionalized Mxene as the conductive substrate in an electrochemical sensor. Mxene itself possesses high metal-like conductivity, significantly enhancing the electron transfer rate and amplifying the electrochemical response signal, thereby achieving highly sensitive detection of corilagin with a detection limit as low as 0.0049 μg / mL. -1 It can be repeatedly eluted and regenerated ≥5 times.

[0019] This invention combines boric acid affinity with surface molecular imprinting technology. First, the boric acid group can specifically capture corilagin containing a cis-diol structure through reversible covalent bonds, providing the first layer of selectivity. Subsequently, in-situ polymerization and elution occur around the immobilized template molecule, forming an imprinted layer with a precise three-dimensional cavity, providing a second layer of specific recognition based on spatial structure and forces. This dual mechanism of "chemical recognition + spatial recognition" enables the sensor to exhibit extremely high selectivity for corilagin, effectively eliminating the influence of common interfering substances such as phenolic acids and sugars in traditional Chinese medicine extracts.

[0020] By employing a "surface imprinting" strategy, imprinted sites are constructed on the surface of a conductive Mxene-BA material, rather than within the polymer. This structure significantly reduces mass transfer resistance, enabling target molecules to reach the recognition sites more quickly, resulting in a faster response rate.

[0021] The preparation method is clear, the conditions are mild, and it is easy to operate and control. The prepared sensor exhibits good stability under low-temperature sealed storage, can be reused, and reduces the cost per detection.

[0022] This sensor and detection method are simple to operate, fast to detect, and low in cost. They are particularly suitable for rapid screening and quantitative analysis of Corilagin in Chinese medicinal materials producing areas, grassroots drug testing units, or production sites, providing a powerful technical tool for the quality control of Chinese medicine. Attached Figure Description

[0023] Figure 1 The Zeta potential diagram is for the sample prepared in Example 1.

[0024] Figure 2 The XPS full spectrum of the sample prepared in Example 1.

[0025] Figure 3 The XPS high-resolution spectrum of the sample prepared in Example 1 is shown below: where B represents silicon, C represents nitrogen, D represents oxygen, E represents carbon, F represents titanium, and G represents boron.

[0026] Figure 4 The following are scanning electron microscope images of the samples prepared in Example 1, with a scale bar of 1 μm: where A is Ti3C2TX Mxene, B is Mxene-BA, C is before elution of MIP / Mxene-BA, D is after elution of MIP / Mxene-BA, E is before elution of NIP / Mxene-BA, and F is after elution of NIP / Mxene-BA.

[0027] Figure 5 The working curves and fitting results for detecting Corilagin using the MIP / Mxene-BA@GCE sensor are shown: where A represents the fitting results for a concentration range of 0.025-10 μg / mL, and B represents the fitting results for a concentration range of 10-75 μg / mL.

[0028] Figure 6 The results of the specificity and selectivity experiments of MIP / Mxene-BA@GCE are as follows: where A) is the structural formula of COR, EA, GA, phenol, and GLU; B is the difference in current response signal after the sensor re-adsorbs COR, EA, GA, NaCl, GLU, and phenol solutions with a concentration of 50 μg / mL.

[0029] Figure 7 Storage stability assessment for MIP / Mxene-BA@GCE sensors (n=3). Detailed Implementation

[0030] The present invention will now be described in further detail with reference to the accompanying drawings and embodiments, but this does not limit the present invention in any way. Any modifications or improvements made based on the teachings of the present invention shall fall within the protection scope of the present invention.

[0031] The processes, conditions, reagents, and experimental methods used in implementing this invention, except as specifically mentioned below, are all common knowledge and general knowledge in the field, and this invention does not have any particular limitations. Experimental methods in the embodiments that do not specify specific conditions are generally performed under conventional conditions or as recommended by the manufacturer.

[0032] Unless otherwise stated, all technical terms and scientific terms used in this specification have the same meaning as commonly understood by one of ordinary skill in the art to which this invention pertains. However, in the event of any conflict, the specification containing the definitions shall prevail.

[0033] The following is information on the source and specifications of the main raw materials and reagents used in the examples: Corilagin (>98%, 1 g / bottle, Pusi Biotechnology Co., Ltd.); Acrylamide (99.0%, 500 g / bottle, Shanghai Maclean Biotechnology Co., Ltd.); N,N′-Methylenebisacrylamide (analytical grade, 25 g / bottle, Shanghai Maclean Biotechnology Co., Ltd.); Ammonium persulfate (98.5%, 500 g / bottle, Shanghai Maclean Biotechnology Co., Ltd.); Chitosan (biotechnology grade, 25 g / bottle, Shanghai Maclean Biotechnology Co., Ltd.); Acetic acid (>99.5%, 500 mL / bottle, Xilong Scientific Co., Ltd.); Titanium carbide MXene nanosheets Ti3C2T x (54–68 wt%, 1 g / bottle, Pioneer Nano); Ammonia (25–28%, 500 mL / bottle, Shanghai Maclean Biotechnology Co., Ltd.); Anhydrous Ethanol (Analytical grade, 500 mL / bottle, Xilong Scientific Co., Ltd.); 3-Aminopropyltriethoxysilane (99%, 25 mL / bottle, Shanghai Aladdin Biochemical Technology Co., Ltd.); Sodium cyanoborohydride (95%, 25 g / bottle, Shanghai Maclean Biochemical Technology Co., Ltd.); Anhydrous Methanol (≥99.5%, 500 mL / bottle, Guangdong Guanghua Technology Co., Ltd.); Methanol (>99.5%, 500 mL / bottle, Sinopharm Chemical Reagent Co., Ltd.); Potassium Chloride (Superior grade, 500 g / bottle, Shanghai Aladdin Biochemical Technology Co., Ltd.); Potassium Ferricyanide K3[Fe(CN)6] (>99.5%, 500 g / bottle, Xilong Scientific Co., Ltd.); 4-Formylphenylboronic acid (98%) Potassium ferrocyanide trihydrate K4[Fe(CN)6]·3H2O (>99.5%, 500g / bottle, Xilong Scientific Co., Ltd.); 10×PBS (0.22 μm filtered, 500 mL / bottle, FodeBio). Example 1: Preparation of Corilagin Molecularly Imprinted Electrochemical Sensor (MIP / Mxene-BA@GCE) (1) Preparation of boric acid-functionalized Mxene (Mxene-BA) conductive affinity composite material 1) 100 mg of multilayer Ti3C2T X Mxene powder was dispersed in 16 mL of anhydrous ethanol, and 1 mL of ultrapure water and 1.5 mL of 25% ammonia solution were added. The mixture was magnetically stirred at 25 °C for 24 h. Subsequently, 400 μL of 3-aminopropyltriethoxysilane (APTES) was slowly added dropwise, and stirring was continued for another 24 h. After the reaction was completed, the mixture was centrifuged at 4000 rpm for 5 min, washed successively with ultrapure water and methanol until neutral, and dried under vacuum at 45 °C for 12 h to obtain aminated Mxene (Mxene-NH2).

[0034] 2) Take 80 mg Mxene-NH2 and redisperse it in 16 mL of anhydrous methanol. Add 160 mg of 4-formylphenylboronic acid (4-FPBA) and 200 mg of sodium cyanoborohydride (NaBH3CN). Sonicate for 5 min and stir at room temperature for 24 h. After centrifugation, washing with methanol and vacuum drying at 45 °C for 12 h, boronic acid functionalized Mxene (Mxene-BA) is obtained.

[0035] 3) Disperse 4 mg Mxene-BA in 1 mL of ultrapure water and sonicate for 30 min; add 1 mL of 1% (w / w) chitosan acetic acid solution and continue sonicating for 10 min to obtain a homogeneous Mxene-BA / chitosan composite modified solution (4 mg / mL). -1 Store at 4℃ away from light for later use.

[0036] (2) Pretreatment of glassy carbon electrode: A φ3 mm glassy carbon electrode (GCE) was polished to a mirror finish with 1.0 μm, 0.3 μm, and 0.05 μm Al2O3, rinsed with ultrapure water, and then ultrasonically cleaned for 3 min; it was then placed in 5 mM [Fe(CN)6] 3- / 4- (containing 0.1 M KCl) Cyclic voltammetry scan from 0.2 to +0.6 V (vs. Ag / AgCl) until ΔEp < 80 mV, then dry with nitrogen for later use.

[0037] (3) Preparation of Mxene-BA modified electrode (Mxene-BA@GCE) Using a micropipette, 2 μL of the Mxene-BA / chitosan modification solution prepared in step (1) was vertically drop-coated onto the clean glassy carbon electrode surface pretreated in step (2). The electrode was placed under an infrared drying lamp until the droplet was completely dry. Then, another 3 μL of modification solution was added to the same position, and the electrode was dried again. This repeated drop-coating step was intended to form a uniform and stable modification layer. The resulting electrode was designated Mxene-BA@GCE.

[0038] (4) Preparation of Corilagin molecularly imprinted electrochemical sensor (MIP / Mxene-BA@GCE) 1) Template molecule immobilization: The Mxene-BA@GCE prepared in step (3) was immersed in 200 μL of 30 μg / mL corilagin phosphate buffer solution (PBS, 10 mM, pH 8.5) and incubated at 25°C for 3 hours. Under these conditions, the catechol structure (cis-diol) in the corilagin molecule undergoes specific and reversible covalent bonding with the boric acid group on the surface of Mxene-BA, thereby immobilizing it on the electrode surface. After incubation, the electrode surface was gently rinsed several times with ultrapure water to remove the physically adsorbed template molecules, and then dried with nitrogen gas.

[0039] 2) Surface Molecular Imprinted Polymerization: The electrode with the template molecules immobilized was placed in 250 μL of a prepolymerization solution. This prepolymerization solution was prepared by dissolving 250 mmol / L acrylamide (monomer) and 35 mmol / L N,N′-methylenebisacrylamide (crosslinking agent) in 10 mM PBS (pH 7.4). The mixture was allowed to stand at room temperature for 0.5 hours to allow the monomers to fully preassemble around the template molecules. Then, 200 μL of PBS solution containing 10 mmol / L ammonium persulfate (APS, initiator) was added to the system to initiate the polymerization reaction. The reaction was carried out at 25 °C for 2 hours, forming a crosslinked polyacrylamide layer encapsulating the Corylagin template on the electrode surface.

[0040] 3) Elution of template molecules: The electrode after polymerization was immersed in 200 μL of a mixed eluent of methanol and acetic acid (volume ratio 9:1) and sonicated for 20 minutes. This step breaks the borate ester bonds and dissolves the template molecules, eluting them from the polymer network and leaving imprinted cavities in the imprinted polymer layer that are complementary to the shape, size, and functional groups of the corilagin molecules. After elution, the electrode was rinsed with ultrapure water and dried with nitrogen gas to obtain the corilagin molecularly imprinted electrochemical sensor, denoted as MIP / Mxene-BA@GCE.

[0041] 4) Preparation of non-blot sensor (NIP / Mxene-BA@GCE): As a control, except that an equal volume of PBS buffer without corilagin was used instead of the template solution for template molecule fixation, the other steps were exactly the same as the preparation of the blot sensor described above, and a non-blot sensor was obtained.

[0042] Example 2 Characterization of Corilagin Molecularly Imprinted Electrochemical Sensor (MIP / Mxene-BA@GCE) The prepared Corilagin molecularly imprinted electrochemical sensors were characterized by zeta, XPS, potential, and SEM.

[0043] Zeta potential characterization results (see...) Figure 1 This indicates that Ti3C2T X The surface charge properties of its functionalized products change significantly, with the original Ti3C2T... X The negatively charged Mxene-NH2, after amination, is covalently grafted onto 3-aminopropyltriethoxysilane via a silicon-oxygen bond (Si-O-Ti), causing the exposed -NH2 to be protonated to -NH. 3+ The potential reverses to positive. The borate group (-B(OH)2) on Mxene-BA undergoes deprotonation equilibrium (-B(OH)2). -B(OH)O - + H + ), generating negatively charged [B(OH)O - The measured Zeta potential results are consistent with the theoretical results of the modification.

[0044] The chemical composition and bonding state of Ti3C2TXMxene before and after surface modification were investigated using XPS. Results are shown below. Figure 2 The original Ti3C2TX spectrum showed characteristic peaks for Ti 2p, C 1s, O 1s, and F 1s. After amination, N 1s (~399.8 eV, ~402 eV) and Si 2p (~102 eV) peaks were added. The boric acid-modified product (Mxene-BA) retained the above elemental peaks and added a B 1s peak (~192 eV), confirming the introduction of the boric acid group. Further peak fitting of the high-resolution spectra of each element was performed, and the resulting spectra are shown below. Figure 3 As can be seen in the high-resolution spectrum of N 1s: Mxene-NH2 contains -NH 3+ The proportion of -NH2 (402 eV) is greater than that of -NH2 (399.8 eV), indicating that protonation of the amino group is dominant. In Mxene-BA, Schiff base reduction forms a neutral secondary amine bond (-NH-CH2-), and -NH... 3+ The relative proportion decreased. High-resolution C 1s spectrum: Mxene-BA shows a CN bond characteristic peak at 286.2 eV (corresponding to the -CH in the silane chain).2- (NH-), consistent with the amination reaction pathway. High-resolution O1s spectra: The presence of a Si-O-Ti bond peak at 531 eV in Mxene-NH2 confirms the covalent bonding between silane and the hydroxyl groups on the Mxene surface. A new peak at 533.4 eV is added to Mxene-BA, attributed to the BO bond in the borate group. XPS data validate the success of the functionalization modification, corroborating the Zeta potential results.

[0045] Field emission scanning electron microscopy was used to study Ti3C2T X The morphology of Mxene multilayer nanosheets before and after modification, as well as the morphology of MIP / Mxene-BA and NIP / Mxene-BA before and after elution, were characterized (see...). Figure 4 Ti3C2T X Mxene multilayer nanosheets exhibit a typical multilayer stacked structure with a smooth and flat surface. After amination and boric acid modification, the material still maintains its layered structure, but obvious particle adhesion and increased roughness are visible on the surface, indicating that functionalized molecules have been successfully modified onto the Mxene surface. Corilagin is fixed on the surface via boric acid affinity reaction. A clearly visible molecularly imprinted polymer is formed on the surface through crosslinking of functional monomers and crosslinking agents around the template. Eluent elution in an acidic environment simultaneously disrupts the borate ester bond and hydrogen bond network, causing the embedded corilagin molecules to be removed from the polymer, resulting in a corresponding morphological change. As a control, the non-imprinted polymer, lacking corilagin molecules, failed to form a clearly visible polymer on its surface. No significant change in surface morphology was observed after elution. Comprehensive analysis of the morphology of the above materials confirms the successful synthesis of the molecularly imprinted polymer.

[0046] Example 3 Sensor Performance Testing (1) Linear range The MIP / Mxene-BA@GCE sensor prepared in Example 1 was placed in a series of different concentrations (0.1, 0.5, 1, 5, 10, 30, 40, 50, 60, 75 μg / mL) of Corilagin standard PBS solution (pH 7.4) and resorbed at 25°C for 15 minutes. Differential pulse voltammetry (DPV) was used to analyze the 5 mM [Fe(CN)6] solution. 3- / 4- The current response of the sensor was detected in an electrolyte of 0.1 M KCl. A standard curve was plotted with the concentration of Corilagin (C) on the x-axis and the difference in DPV peak current before and after resorption (ΔI) on the y-axis.

[0047] The experimental results are shown in Figure 5The results showed two different linear relationships on both sides of the 10 μg concentration: In the range of 0.025–10 μg / mL, a good linear relationship was observed between the difference in current response and the natural logarithm of the corilagin concentration: ΔI = 3.3662LnC + 19.62, R0 2 =0.9988. In the range of 10⁻⁷⁵ μg / mL, the difference in response current between MIP / Mxene-BA@GCE and the concentration of corilagin satisfies a linear relationship: ΔI = 0.3668 × C + 23.334, R₀ 2 =0.9967. When the concentration to be measured is in the range of 0.025-75 μg / mL, the sensor exhibits a good linear relationship, and its concentration can be quickly and accurately determined based on the electrical signal response value and the linear relationship.

[0048] (2) Limit of detection and limit of quantitation The sensor current response values ​​were measured in parallel 12 times in a blank solution without corilagin. The standard deviation of the sensor was calculated to be 5.825 × 10⁻⁶. -7 Based on the previously obtained method calibration curve and the response value and standard curve slope method, the detection limit and quantitation limit of the method were determined to be: DL = 0.004951 μg / mL and QL = 0.01585 μg / mL, respectively.

[0049] (3) Specificity and selectivity The peak current response signals were measured using MIP / Mxene-BA@GCE and NIP / Mxene-BA@GCE modified electrodes in solutions of 100 mM concentrations of corilagin (COR), ellagic acid (EA), gallic acid (GA), glucose (GLU), phenol, and sodium chloride (NaCl).

[0050] See results Figure 6 The results showed that, at the same concentration, the current response of MIP / Mxene-BA@GCE to the template molecule COR was significantly higher than that of all other substances. This confirms that MIP / Mxene-BA@GCE has good specificity and selectivity for corilagin and can effectively overcome the influence of common interfering substances in complex matrices.

[0051] (4) Repeatability and intermediate precision Repeatability: Six MIP / Mxene-BA@GCE sensors prepared in the same batch were used to detect a 5 μg / mL corilagin solution, and the relative standard deviation (RSD) was calculated. Intermediate precision: Three MIP / Mxene-BA@GCE sensors were prepared and re-adsorbed onto samples of the same concentration of corilagin. Measurements were performed using two different electrochemical workstations (CHI1030C and CHI660E). The average value and relative standard deviation of the six measurements ΔI were calculated.

[0052] The results are shown in Tables 1 and 2. The RSD of the detection results of the same batch of sensors for the same sample was 2.37%, indicating that the sensor preparation process is stable and the method has high measurement consistency under the same conditions. The RSD of the results obtained by testing across two different electrochemical workstations remained at a low level, only 2.50%, indicating that the method has low dependence on the instrument system and good cross-platform applicability.

[0053] Table 1. Results of method repeatability verification (n=6)

[0054] Table 2. Results of intermediate precision testing (n=3)

[0055] (5) Stability The MIP / Mxene-BA@GCE was stored in a sealed container at 2–8 °C and monitored for 10 days. After template molecule elution was completed using the sensor, the sample was processed at 5 mM [Fe(CN)6]. 3- / 4- The DPV peak current value measured in the electrolyte solution (containing 100 mM KCl) was used as the initial response benchmark (defined as 100%), and the subsequent detections at each time point were compared with this initial value.

[0056] See results Figure 7 During the 10-day testing period, the sensor's current response remained at a relatively stable level, which provides important evidence for its further practical application.

[0057] Example 4: Actual Sample Detection Phyllanthus emblica from different origins was pulverized, weighed precisely, extracted with 70% methanol aqueous solution using ultrasound, and the supernatant was taken after centrifugation, diluted appropriately, and the pH was adjusted with PBS.

[0058] The standard addition method was used, and the MIP / Mxene-BA@GCE sensor of this invention was employed for detection. Simultaneously, HPLC was used for comparative verification. The results are shown in Table 3. The results indicate that the average detection result of the two methods was 0.158%, and the RSD was 1.8%. This demonstrates that the sensor established in this invention can be accurately and reliably used for the quantitative analysis of corilagin in complex matrices of actual Chinese medicinal materials.

[0059] Table 3. Results of Corilagin content detection in the same sample by HPLC and electrochemical methods (n=3)

[0060] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention, and not to limit them; although the present invention has been described in detail with reference to the foregoing embodiments, those skilled in the art should understand that modifications can still be made to the technical solutions described in the foregoing embodiments, or equivalent substitutions can be made to some of the technical features; and these modifications or substitutions do not cause the essence of the corresponding technical solutions to deviate from the spirit and scope of the technical solutions of the embodiments of the present invention.

Claims

1. A Corilagin molecularly imprinted electrochemical sensor, characterized in that, include: Glassy carbon electrode substrate; A boric acid-functionalized Mxene conductive affinity composite material layer covering the surface of the glassy carbon electrode; And a Corilagin molecularly imprinted polymer layer located on the surface of the conductive affinity composite material layer; The imprinted polymer layer contains an imprinted cavity that matches the molecular structure of Corilagin.

2. The sensor according to claim 1, characterized in that, The boric acid-functionalized Mxene conductive affinity composite material is formed by modifying 4-formylphenylboronic acid onto Ti3C2T using an aminosilane coupling agent. X It is prepared by surface preparation of Mxene.

3. A method for preparing a Corilagin molecularly imprinted electrochemical sensor as described in any one of claims 1-2, characterized in that, Includes the following steps: (1) Prepare boric acid-functionalized Mxene conductive affinity composite material and disperse it in chitosan acetic acid solution to prepare a modified solution; (2) The modification solution is drop-coated onto the surface of the pretreated glassy carbon electrode and dried to obtain the Mxene-BA modified electrode; (3) Corilagin template molecules are fixed on the surface of the Mxene-BA modified electrode by boric acid affinity reaction; (4) Molecular imprinting polymerization reaction is carried out on the electrode surface on which template molecules are fixed; (5) Elute the template molecules to obtain the Corilagin molecularly imprinted electrochemical sensor.

4. The method according to claim 3, characterized in that, In step (1), the preparation of the boric acid-functionalized Mxene conductive affinity composite material includes: Multilayer Ti3C2T X Mxene reacts with 3-aminopropyltriethoxysilane in an alkaline ethanol solution to give aminated Mxene; The aminoated Mxene was subjected to a reductive amination reaction with 4-formylphenylboronic acid and sodium cyanoborohydride in methanol to obtain the boric acid-functionalized Mxene.

5. The method according to claim 3, characterized in that, In step (3), the conditions for immobilizing the Corilagin template molecule include: template molecule concentration of 10–50 μg / mL, incubation pH of 8.0–9.0, and incubation time of 2–4 hours.

6. The method according to claim 3, characterized in that, In step (4), the molecular imprinting polymerization reaction uses acrylamide as a monomer, N,N′-methylenebisacrylamide as a crosslinking agent, and ammonium persulfate as an initiator.

7. The method according to claim 3, characterized in that, The method for eluting template molecules in step (5) is to use a mixture of methanol and acetic acid for ultrasonic elution.

8. The application of the corilagin molecularly imprinted electrochemical sensor according to any one of claims 1-2 or the corilagin molecularly imprinted electrochemical sensor prepared by the method according to any one of claims 3-7 in the detection of corilagin.

9. A method for detecting the content of corilagin in a sample, characterized in that, Using the Corilagin molecularly imprinted electrochemical sensor as described in any one of claims 1-2 includes the following steps: The sensor was placed in the test solution containing corilagin for re-adsorption; The current response signal of the sensor is detected using an electrochemical method; Based on the pre-established quantitative relationship between the sensor response signal and the concentration of corilagin, the content of corilagin in the sample to be tested is calculated.

10. The method according to claim 9, characterized in that, The sample to be tested is a traditional Chinese medicinal material or its extract.

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