A ti3c2-based nanocomposite material, a preparation method thereof and application thereof in a biosensor

Abstract

The application discloses a kind of nanocomposite based on Ti3C2 and its preparation method and application.It relates to the field of electrochemistry.The above-mentioned nanocomposite based on Ti3C2 includes the following components: Ti3C2, reduced graphene oxide, beta-cyclodextrin-metal organic framework material, IAA antibody and adhesive.The above-mentioned nanocomposite based on Ti3C2 of the application has excellent specificity for IAA, can realize high sensitivity and high stability detection of IAA, can realize detection of IAA concentration from 0.01-10ng / mL at least, and the detection limit reaches 2.88pg / mL (S / N=3).

Description

Technical Field

[0001] This invention relates to the field of electrochemical technology, and in particular to a Ti3C2-based nanocomposite material, its preparation method, and its application in biosensors. Background Technology

[0002] Indoleacetic acid (IAA), an important plant hormone, influences cell division, elongation, and differentiation, as well as the growth, maturation, and senescence of vegetative and reproductive organs. Studies have found that IAA promotes plant and fruit growth and development, accumulating in rapidly dividing and growing parts of the plant, such as stem and leaf meristems, young leaf tissues, young fruits, and seeds. Therefore, due to its crucial role in plant growth regulation, IAA has been widely used in agricultural production. With technological advancements, electrochemical biosensors are gaining increasing attention in IAA detection methods due to their simplicity, speed, high sensitivity, and good selectivity.

[0003] Currently, it has been reported that low concentrations of IAA can be detected using AuNPs / HRP / IgG (gold nanoparticles / antibody / immunoglobulin G) functionalized gold nanoparticles as signal amplification probes and AuNPs / PG / GCE nanocomposite modified electrodes. However, due to the low content of IAA in plant tissues and its sensitivity to external conditions such as light, temperature, and oxidants, coupled with interference from complex metabolites in plant tissues, existing probes or modified electrodes cannot detect IAA with high sensitivity and selectivity.

[0004] Therefore, there is an urgent need for a novel electrochemical sensing element to construct a highly sensitive and selective electrochemical biosensor for detecting IAA. Summary of the Invention

[0005] The first technical problem to be solved by this invention is:

[0006] A Ti3C2-based nanocomposite material is provided.

[0007] The second technical problem to be solved by this invention is:

[0008] A method for preparing the Ti3C2-based nanocomposite material is provided.

[0009] The third technical problem to be solved by this invention is:

[0010] Applications of the Ti3C2-based nanocomposite materials.

[0011] The present invention also proposes a modified electrode, comprising an electrode and a material modified on the electrode, wherein the material modified on the electrode comprises the aforementioned Ti3C2-based nanocomposite material.

[0012] To solve the first technical problem, the technical solution adopted by the present invention is as follows:

[0013] A Ti3C2-based nanocomposite material comprises the following components:

[0014] Ti3C2, reduced graphene oxide, β-cyclodextrin-metal-organic framework materials, IAA antibodies, and adhesives.

[0015] According to embodiments of the present invention, one of the technical solutions has at least one of the following advantages or beneficial effects:

[0016] 1. Titanium carbide (Ti3C2) is an important component of the family of two-dimensional transition metal carbide and nitride materials (MXenes). Ti3C2 has a unique two-dimensional layered structure, which makes it easy to be densely stacked on the electrode. In this invention, Ti3C2 is used as a template material to assemble Ti3C2-based nanocomposite materials, thereby achieving highly sensitive detection of IAA.

[0017] 2. Ti3C2 and reduced graphene oxide have similar layered structures and surface chemical properties, as well as strong interfacial interactions due to the Ti-OC covalent bonds between them. The three-dimensional heterostructure constructed from Ti3C2 and rGO can effectively improve the aggregation of Ti3C2 and rGO, further enhancing the stability of the Ti3C2-based nanocomposite material, thereby enabling stability testing of IAA.

[0018] 3. β-cyclodextrin (β-CD) has a hydrophobic interior with a ring-shaped cavity and a hydrophilic exterior; metal-organic framework materials (MOFs) are metal ions or metal ion clusters of mixed porous materials; in this invention, β-cyclodextrin and metal-organic framework materials are combined to improve their porous structure and high loading capacity, thereby loading materials such as Ti3C2, reduced graphene oxide and IAA antibodies.

[0019] 4. The addition of adhesives is mainly used to fix IAA antibodies.

[0020] 5. The Ti3C2-based nanocomposite material of the present invention has excellent specificity for IAA, enabling highly sensitive and stable detection of IAA, and can detect IAA concentrations from 0.01 to 10 ng / mL, with a detection limit of 2.88 pg / mL (S / N=3).

[0021] According to one embodiment of the present invention, in the β-cyclodextrin-metal-organic framework material, β-cyclodextrin is used as the organic ligand and a metal ion is used as the central ion, wherein the metal ion includes at least one of potassium ions and sodium ions. For the present invention, the role of the metal-organic framework material is mainly reflected in its porous structure and high loading capacity. Based on this, the selection of the central ion in the metal-organic framework material can be made as needed, selecting ions with specific functions, and endowing the metal-organic framework material with the same properties as the ions themselves.

[0022] According to one embodiment of the present invention, the adhesive comprises pentylene glycol and chitosan. Pentylene glycol is primarily used to immobilize antibodies, while chitosan serves both as an adhesive for nanomaterials and as a fixative for pentylene glycol. In this invention, since the primary function of pentylene glycol and chitosan is as adhesives, in specific implementations, pentylene glycol and chitosan can be substituted with other adhesives suitable for biosensors.

[0023] According to one embodiment of the present invention, the mass ratio of Ti3C2 to reduced graphene oxide is 10-20:2-4.

[0024] According to one embodiment of the present invention, in the Ti3C2-based nanocomposite material, the ratio of the total mass of Ti3C2, reduced graphene oxide and β-cyclodextrin-metal-organic framework material to the mass of IAA antibody is 12-24:6-12.

[0025] According to one embodiment of the present invention, a modified electrode is also provided, comprising an electrode and a material modified on the electrode, wherein the material modified on the electrode comprises a Ti3C2-based nanocomposite material as described above.

[0026] According to one embodiment of the present invention, the material modified on the electrode further includes bovine serum albumin (BSA). BSA blocks excess adsorption sites on the modified electrode surface. For the present invention, BSA can be replaced with other similar materials, such as mercaptohexanol (MCH) and 1,6-hexanethiol (DHT). In practice, any material that blocks electrode sites can be substituted in parallel.

[0027] According to one embodiment of the present invention, the base electrode of the modified electrode can be a glassy carbon electrode, a gold electrode, or a platinum electrode, or any electrode can be selected as the base electrode of the modified electrode as needed.

[0028] According to one embodiment of the present invention, the preparation of the modified electrode includes the following steps: 20-40 µL of 5-10 mg / mL β-CD-MOFs / rGO / Ti3C2 composite nanomaterial and 10-20 µL of 6-12 mg / mL chitosan (CS) are ultrasonically mixed; 6-12 µL of the CS and β-CD-MOFs / rGO / Ti3C2 mixed solution is drop-coated onto the electrode surface; the electrode is dried using infrared spectroscopy; 6-12 µL of 2.5-5% glutaraldehyde (GA) (diluted with 10-20 mM PBS at approximately pH 7.4) is added to the modified electrode and reacted for 2-4 h; the electrode is then washed with ddH2O and dried for later use to obtain the modified electrode.

[0029] According to one embodiment of the present invention, the preparation of the modified electrode further includes the following steps: adding 6-12 µL of 1-2 mg mL... -1 Anti-IAA was applied to the surface of a Ti3C2 modified electrode and incubated overnight at 3-4°C. Then, 0.5-1.5 mg / mL of bovine serum albumin (BSA) was added to block excess adsorption sites on the modified electrode surface. After incubation at 3-4°C for 40-60 min, the electrode was washed and dried to obtain the modified electrode with antibody IAA.

[0030] To solve the second technical problem, the technical solution adopted by the present invention is as follows:

[0031] A method for preparing the Ti3C2-based nanocomposite material includes the following steps:

[0032] S1. Ti3C2-rGO composite material was prepared by mixing Ti3C2 and reduced graphene oxide; β-cyclodextrin and metal materials were mixed to prepare β-cyclodextrin-metal-organic framework material.

[0033] S2 mixed Ti3C2-rGO composite material, β-cyclodextrin-metal-organic framework material and adhesive to obtain composite nanomaterials;

[0034] S3. IAA antibody is added to the surface of the dried composite nanomaterial to obtain the Ti3C2-based nanocomposite material.

[0035] According to one embodiment of the present invention, step S3 further includes the following steps: after adding IAA antibody to the surface of the dried composite nanomaterial, incubating it overnight at 3-4°C, then adding bovine serum albumin and incubating it at 3-4°C.

[0036] According to one embodiment of the present invention, step S1, the preparation of the Ti3C2-rGO composite material includes the following steps: weighing 10-20 mg of Ti3C2 and 2-4 mg of rGO, dissolving them in 10-20 mL of ddH2O, vortexing to mix, and then ultrasonically mixing. The Ti3C2-rGO composite material is obtained by centrifugation, washing, and vacuum drying.

[0037] According to one embodiment of the present invention, step S1, the preparation of β-cyclodextrin-metal-organic framework materials includes the following steps: dissolving 2-4 g of β-CD and 0.1-2 g of KOH in 40-60 mL of ultrapure water to obtain a mixture of β-CD and KOH, and then filtering. Methanol is then evaporated into the mixture. After 3-7 days, white crystals appear at the bottom of the container. The washed wet sample of β-CD-MOFs is then placed in a vacuum oven to dry.

[0038] According to one embodiment of the present invention, step S2 further includes the synthesis of β-CD-MOFs / rGO / Ti3C2 composite nanomaterials, specifically including the following steps: dissolving 3-6 mg rGO / Ti3C2 and 3-6 mg β-CD-MOFs in 1-2 mL ddH2O, vortexing and mixing, and then ultrasonically mixing to obtain β-CD-MOFs / rGO / Ti3C2 composite nanomaterials.

[0039] Another aspect of the present invention relates to the application of the Ti3C2-based nanocomposite material in the fabrication of electrochemical sensors. This includes the Ti3C2-based nanocomposite material as described in the first aspect of the embodiments above. Since this application employs all the technical solutions of the aforementioned Ti3C2-based nanocomposite material, it possesses at least all the beneficial effects brought about by the technical solutions of the above embodiments.

[0040] According to one embodiment of the present invention, in the electrochemical sensor, the working electrode is the recognition electrode modified with the Ti3C2-based nanocomposite material, the reference electrode is an Ag / AgCl electrode, and the auxiliary electrode is a platinum wire. Fe(CN)6 3− / 4- As a probe, IAA antibody immunization directly reduces Fe(CN)6. 3− / 4- DPV peak current enables highly sensitive detection of IAA.

[0041] According to one embodiment of the present invention, in the electrochemical sensor, electrochemical measurements are all performed at 10-20 mM 5-10 mM Fe(CN)6 3− / 4- The experiment was conducted in PBS solution. Fe(CN)6 was utilized using DPV. 3− / 4- The reduction in peak current was used to detect IAA. 10-20 μL of IAA in 10-20 mM PBS solution of different concentrations was dropped onto the electrode surface and incubated at about 37°C for 1-2 hours. The peak current (ΔI) of the electrochemical signal reduction was recorded to reflect the value of IAA, and a calibration curve was constructed.

[0042] Another aspect of the present invention relates to the application of the Ti3C2-based nanocomposite material in the detection of plant growth hormones. This includes the Ti3C2-based nanocomposite material as described in the first aspect of the embodiments above. Since this application employs all the technical solutions of the aforementioned Ti3C2-based nanocomposite material, it possesses at least all the beneficial effects brought about by the technical solutions of the above embodiments.

[0043] According to one embodiment of the present invention, the Ti3C2-based nanocomposite material is applied to detect plant auxins in actual samples. During the detection process, the working electrode is an electrode modified with the Ti3C2-based nanocomposite material, and the reference electrode is a glassy carbon electrode. The entire experiment is conducted in a light-protected environment. More specifically, before detection, sample pretreatment is included: fresh Arabidopsis thaliana and rice leaves are selected, ground into powder using liquid nitrogen, 100-200 mg of powder is accurately weighed, 1-2 mL of 80% methanol solution is added, mixed well, and extracted at 4-8 °C for 12-24 h. The extract is then detected by an electrochemical sensor.

[0044] According to one embodiment of the present invention, the detection process for detecting plant auxin in an actual sample includes the following steps: obtaining an initial current peak value I0 using the DPV method; adding a sample for detection to obtain a current peak value I, and calculating the current change value ΔI = I0 - I; and calculating the concentration of plant auxin according to the detection relationship function.

[0045] According to one embodiment of the present invention, the detection process for detecting plant growth hormone in actual samples has the following relationship function: y = 0.27x + 0.3264; the curve equation is y = 3.1187x + 1.6745, the correlation coefficient is 0.9953, y represents the current in μA, and x represents the concentration of IAA in log(ng / mL).

[0046] The plant auxin antibody used in the biosensor of this invention has a strong specific binding ability to auxin, resulting in a biosensor with significant sensitivity advantages. It has been successfully applied in rice and Arabidopsis leaf samples. This biosensor can be used to detect auxin content in plant samples, offering advantages such as simple operation, low detection limit, high sensitivity, and high accuracy. It provides a simple, rapid, sensitive, and accurate method for plant auxin detection. It is expected to provide strong methodological support for electrochemical biosensor detection.

[0047] Other features and advantages of the invention will be set forth in the description which follows, and will be apparent in part from the description, or may be learned by practicing the invention. Attached Figure Description

[0048] The above and / or additional aspects and advantages of the present invention will become apparent and readily understood from the description of the embodiments taken in conjunction with the following drawings, in which:

[0049] Figure 1 This is a schematic diagram of the planar structure of the modified electrode preparation method in Example 1;

[0050] Figure 2 Characterization diagram of the β-CD-MOFs / rGO / Ti3C2 electrode modified material in Example 1;

[0051] Figure 3 The above are electrochemical test diagrams from the preparation process of the modified electrode in Example 1.

[0052] Figure 4 The test diagram shows the detection of different concentrations of IAA using the modified electrode of Example 1.

[0053] Figure 5 This is a test diagram for a selective study of the modified electrode of Example 1. Detailed Implementation

[0054] The embodiments of the present invention are described in detail below. Throughout the embodiments, the same or similar reference numerals denote the same or similar elements or elements having the same or similar functions. The embodiments described below are exemplary and are only used to explain the present invention, and should not be construed as limiting the present invention.

[0055] In the description of this invention, the use of terms such as "first," "second," etc., is for the purpose of distinguishing technical features only and should not be construed as indicating or implying relative importance, or implicitly indicating the number of technical features indicated, or implicitly indicating the order of the technical features indicated.

[0056] The technical solutions of the present invention will be clearly and completely described below with reference to the embodiments of the present invention. Obviously, the described embodiments are only some embodiments of the present invention, and not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those of ordinary skill in the art without creative effort are within the scope of the present invention.

[0057] Unless otherwise specified, the reagents, methods and equipment used in this invention are all conventional reagents, methods and equipment in this technical field.

[0058] Example 1

[0059] A Ti3C2-based nanocomposite material comprises the following components:

[0060] Ti3C2, reduced graphene oxide (rGO), β-cyclodextrin-metal-organic framework materials, IAA antibodies, and adhesives.

[0061] Adhesives include pentylene glycol and chitosan.

[0062] Specifically, the Ti3C2-based nanocomposite material includes anti-IAA / GA / CS-β-CD-MOFs / rGO / Ti3C2.

[0063] A modified electrode includes a Ti3C2-based nanocomposite material and a glassy carbon electrode, and also includes BSA on the surface of the glassy carbon electrode, wherein the BSA is capable of sealing excess adsorption sites on the surface of the modified electrode.

[0064] A method for preparing a modified electrode, such as Figure 1 The diagram shown is a planar structural schematic of the modified electrode fabrication method, which specifically includes the following steps:

[0065] (1) Synthesis of Ti3C2-rGO: 10 mg Ti3C2 and 2 mg rGO were weighed and dissolved in 10 mL ddH2O, vortexed and mixed, and then ultrasonically mixed. The Ti3C2-rGO composite material was obtained by centrifugation, washing and vacuum drying;

[0066] (2) Synthesis of β-CD-MOFs: 2.26 g of β-CD and 0.9 g of KOH were dissolved in 40 mL of ultrapure water to obtain a mixture of β-CD and KOH, which was then filtered. Methanol was then evaporated into the mixture. After 3 days, white crystals appeared at the bottom of the flask. The washed wet sample of β-CD-MOFs was placed in a vacuum oven and dried at 50 °C for 10 hours.

[0067] (3) Synthesis of β-CD-MOFs / rGO / Ti3C2 composite nanomaterials: 3 mg rGO / Ti3C2 and 3 mg β-CD-MOFs were dissolved in 1 mL ddH2O, vortexed and mixed, and then ultrasonically mixed for 30 min to obtain β-CD-MOFs / rGO / Ti3C2 composite nanomaterials. Figure 2 Characterization diagrams of the β-CD-MOFs / rGO / Ti3C2 electrode modified material are shown. Figure 2 A is the infrared spectrum. Figure 2 B is the X diffraction pattern. Figure 2 C represents the Raman spectrum.

[0068] (4) Take 20 µL of 5 mg / mL β-CD-MOFs / rGO / Ti3C2 composite nanomaterial and 10 µL of 6 mg / mL chitosan (CS) and mix them by sonication. Take 6 µL of the CS and β-CD-MOFs / rGO / Ti3C2 mixed solution and drop it onto the surface of the glassy carbon electrode. Dry it with infrared radiation. Add 6 µL of 2.5% glutaraldehyde (GA) (diluted with pH=7.4, 10 mM PBS) to the modified electrode and react for 2 h. Wash with ddH2O and air dry for later use to obtain the glassy carbon electrode loaded with β-CD-MOFs / rGO / Ti3C2 composite nanomaterial.

[0069] (5) Add 6µL of 1 mg / mL solution dropwise. -1 Anti-IAA was applied to the surface of a glassy carbon electrode loaded with β-CD-MOFs / rGO / Ti3C2 composite nanomaterials and incubated overnight at 4°C. Then, 1.0 mg / mL BSA was added to block excess adsorption sites on the electrode surface, and the electrode was incubated at 4°C for 60 min before being washed and dried to obtain the modified electrode. Figure 3 These are electrochemical test graphs from the preparation process of the modified electrode. Among them, Figure 3 In the figure, A represents the DPV electrochemical response curve. Within this curve, a is the electrode modified with β-CD-MOFs / rGO / Ti3C2 nanomaterials; b is the working electrode modified with glutaraldehyde-chitosan-β-CD-MOFs / rGO / Ti3C2 nanomaterials; c is the working electrode modified with anti-IAA-glutaraldehyde-chitosan-β-CD-MOFs / rGO / Ti3C2 nanomaterials; d is the working electrode modified with BSA / anti-IAA-glutaraldehyde-chitosan-β-CD-MOFs / rGO / Ti3C2 nanomaterials; and e is the electrode in d after combining with IAA. Figure 3In the figure, B represents the EIS electrochemical response curve. In this figure, a is the electrode modified with β-CD-MOFs / rGO / Ti3C2 nanomaterials; b is the working electrode modified with glutaraldehyde-chitosan-β-CD-MOFs / rGO / Ti3C2 nanomaterials; c is the working electrode modified with anti-IAA-glutaraldehyde-chitosan-β-CD-MOFs / rGO / Ti3C2 nanomaterials; d is the working electrode modified with BSA / anti-IAA-glutaraldehyde-chitosan-β-CD-MOFs / rGO / Ti3C2 nanomaterials; and e is the electrode in d after IAA is combined with it.

[0070] The modified electrode described in Example 1 of the present invention has excellent specificity for IAA, enabling high sensitivity and high stability detection of IAA. It can detect IAA concentrations from 0.01 to 10 ng / mL, and the detection limit reaches 2.88 pg / mL (S / N=3).

[0071] Example 2

[0072] The difference between Example 2 and Example 1 is that the electrodes are different. In Example 1, the electrode is a glassy carbon electrode, while in Example 2, the electrode is a gold electrode.

[0073] Specifically:

[0074] A Ti3C2-based nanocomposite material comprises the following components:

[0075] Ti3C2, reduced graphene oxide (rGO), β-cyclodextrin-metal-organic framework materials, IAA antibodies, and adhesives.

[0076] Adhesives include pentylene glycol and chitosan.

[0077] Specifically, the Ti3C2-based nanocomposite material includes anti-IAA / GA / CS-β-CD-MOFs / rGO / Ti3C2.

[0078] A modified electrode includes a Ti3C2-based nanocomposite material and a gold electrode, and also includes BSA on the surface of the gold electrode, wherein the BSA is capable of sealing excess adsorption sites on the surface of the modified electrode.

[0079] A method for preparing a modified electrode includes the following steps:

[0080] (1) Synthesis of Ti3C2-rGO: 10 mg Ti3C2 and 2 mg rGO were weighed and dissolved in 10 mL ddH2O, vortexed and mixed, and then ultrasonically mixed. The Ti3C2-rGO composite material was obtained by centrifugation, washing and vacuum drying;

[0081] (2) Synthesis of β-CD-MOFs: 2.26 g of β-CD and 0.9 g of KOH were dissolved in 40 mL of ultrapure water to obtain a mixture of β-CD and KOH, which was then filtered. Methanol was then evaporated into the mixture. After 7 days, white crystals appeared at the bottom of the flask. The washed wet sample of β-CD-MOFs was placed in a vacuum oven and dried at 50 °C for 10 hours.

[0082] (3) Synthesis of β-CD-MOFs / rGO / Ti3C2 composite nanomaterials: 3 mg rGO / Ti3C2 and 3 mg β-CD-MOFs were dissolved in 1 mL ddH2O, vortexed and mixed, and then ultrasonically mixed for 30 min to obtain β-CD-MOFs / rGO / Ti3C2 composite nanomaterials.

[0083] (4) Take 20 µL of 5 mg / mL β-CD-MOFs / rGO / Ti3C2 composite nanomaterial and 10 µL of 6 mg / mL chitosan (CS) and mix them by sonication. Take 6 µL of the CS and β-CD-MOFs / rGO / Ti3C2 mixed solution and drop it onto the surface of the gold electrode. Dry it with infrared radiation. Add 6 µL of 2.5% glutaraldehyde (GA) (diluted with pH=7.4, 10 mM PBS) to the modified electrode and react for 2 h. Wash with ddH2O and air dry for later use to obtain the gold electrode loaded with β-CD-MOFs / rGO / Ti3C2 composite nanomaterial.

[0084] (5) Add 6 µL of 1 mg / mL solution dropwise. -1 Anti-IAA was applied to the surface of a gold electrode loaded with β-CD-MOFs / rGO / Ti3C2 composite nanomaterials and incubated overnight at 4 °C. Then, 1.0 mg / mL BSA was added to block excess adsorption sites on the electrode surface. After incubation at 4 °C for 60 min, the electrode was washed and dried to obtain the modified electrode.

[0085] Example 3

[0086] The difference between Example 3 and Example 1 is that the modified electrode in Example 3 did not use BSA to block the electrode sites.

[0087] A Ti3C2-based nanocomposite material comprises the following components:

[0088] Ti3C2, reduced graphene oxide (rGO), β-cyclodextrin-metal-organic framework materials, IAA antibodies, and adhesives.

[0089] Adhesives include pentylene glycol and chitosan.

[0090] Specifically, the Ti3C2-based nanocomposite material includes anti-IAA / GA / CS-β-CD-MOFs / rGO / Ti3C2.

[0091] A modified electrode comprising a Ti3C2-based nanocomposite material and a glassy carbon electrode.

[0092] A method for preparing a modified electrode includes the following steps:

[0093] (1) Synthesis of Ti3C2-rGO: 10 mg Ti3C2 and 2 mg rGO were weighed and dissolved in 10 mL ddH2O, vortexed and mixed, and then ultrasonically mixed. The Ti3C2-rGO composite material was obtained by centrifugation, washing and vacuum drying;

[0094] (2) Synthesis of β-CD-MOFs: 2.26 g of β-CD and 0.9 g of KOH were dissolved in 40 mL of ultrapure water to obtain a mixture of β-CD and KOH, which was then filtered. Methanol was then evaporated into the mixture. After 7 days, white crystals appeared at the bottom of the container. The washed wet sample of β-CD-MOFs was placed in a vacuum oven and dried at 50 °C for 10 hours.

[0095] (3) Synthesis of β-CD-MOFs / rGO / Ti3C2 composite nanomaterials: 3 mg rGO / Ti3C2 and 3 mg β-CD-MOFs were dissolved in 1 mL ddH2O, vortexed and mixed, and then ultrasonically mixed for 30 min to obtain β-CD-MOFs / rGO / Ti3C2 composite nanomaterials.

[0096] (4) Take 20 µL of 5 mg / mL β-CD-MOFs / rGO / Ti3C2 composite nanomaterial and 10 µL of 6 mg / mL chitosan (CS) and mix them by sonication. Take 6 µL of the CS and β-CD-MOFs / rGO / Ti3C2 mixed solution and drop it onto the surface of the glassy carbon electrode. Dry it with infrared radiation. Add 6 µL of 2.5% glutaraldehyde (GA) (diluted with pH=7.4, 10 mM PBS) to the modified electrode and react for 2 h. Wash with ddH2O and air dry for later use to obtain the glassy carbon electrode loaded with β-CD-MOFs / rGO / Ti3C2 composite nanomaterial.

[0097] (5) Add 6 µL of 1 mg / mL solution dropwise. -1 Anti-IAA was applied to the surface of a glassy carbon electrode loaded with β-CD-MOFs / rGO / Ti3C2 composite nanomaterials and incubated overnight at 4 °C to obtain the modified electrode.

[0098] Example 4

[0099] A method for determining plant auxins using the modified electrode of Example 1 includes the following steps:

[0100] Pretreatment: Extraction of plant growth regulators: Fresh chili pepper leaves were selected and ground into powder using liquid nitrogen. 100 mg of powder was accurately weighed and added to 1 mL of 80% methanol solution. The mixture was stirred and extracted at 4℃ for 12 h. The mixture was then centrifuged at 12000 rpm for 10 min at 4℃. The supernatant was collected, and the extraction was repeated twice. The supernatants were mixed, and all supernatants were placed in the same tube. The tube opening was covered with a dialysis membrane, and a small hole was left in the cap to facilitate water evaporation. The tube was then dried in a vacuum concentrator. The concentrated sample was reconstituted with 20% methanol for analysis.

[0101] 2. Construction of the standard curve:

[0102] The electrode of this invention was used to detect uric acid solutions with concentrations ranging from 0.01 to 10 ng / mL. Under optimal experimental conditions, differential pulse voltammetry (DPV) curves were recorded within the range of -0.1 V to 0.8 V, and the peak current values ​​were analyzed. As the IAA concentration increases, more IAA binds to anti-IAA through specific recognition, weakening the electrochemical signal. The electrochemical signal decreases with increasing IAA concentration, and the peak current of the DPV shows a linear relationship with the common logarithm of the IAA concentration. The detection relationship function is y = 0.27x + 0.3264; the curve equation is y = 3.1187x + 1.6745, with a correlation coefficient of 0.9953. y represents the current in μA; x represents the IAA concentration in log(ng / mL). The detection limit is 2.88 pg / mL (S / N = 3).

[0103] Performance testing:

[0104] IAA assay

[0105] The modified electrode of Example 1 was used to determine plant auxin (IAA). Specifically, the modified electrode of Example 1 was used as the working electrode, the glassy carbon electrode as the reference electrode, and the platinum wire as the auxiliary electrode, with a 5 mM Fe(CN)6 electrode as the reference electrode. 3- / 4- A 10 mM PBS solution was used as the supporting electrolyte to prepare the biosensor. The differential pulse voltammetry was used for electrical measurement, and the differential pulse voltammetry curve of IAA in the range of -0.1 to 0.8 V was recorded. The standard curve method was used for quantitative analysis.

[0106] All electrochemical measurements were performed at 10 mM and 5 mM Fe(CN)6. 3− / 4- The experiment was conducted in PBS solution. Fe(CN)6 was utilized using DPV. 3− / 4-The peak current reduction was used to detect IAA. 10 μL of IAA in 10 mM PBS solution of different concentrations was dropped onto the electrode surface and incubated at 37°C for 1 h. The peak current (ΔI) of the electrochemical signal reduction was recorded to reflect the value of IAA, and a calibration curve was constructed.

[0107] Among them, the anti-IAA on the Ti3C2 modified electrode surface can specifically adsorb IAA in the test solution, forming an anti-IAA-IAA complex, which leads to a decrease in the electron transfer rate on the electrode surface, a decrease in the peak current on the sensor working electrode surface detected by the electrochemical workstation, and a linear relationship within a certain IAA concentration range.

[0108] Specifically, the aforementioned biosensor was used to detect different concentrations of IAA. The IAA concentrations included 0 pg / mL, 10 pg / mL, 50 pg / mL, 100 pg / mL, 200 pg / mL, 500 pg / mL, 1000 pg / mL, 5000 pg / mL, and 10000 pg / mL. The detection results are as follows: Figure 4 As shown. From Figure 4 As can be seen, the peak current of DPV is linearly related to the common logarithm of IAA concentration. Quantitative analysis of the sample was performed using a standard curve, the equation of which is y=3.1187x+1.6745, the correlation coefficient is 0.9953, and the detection limit is 2.88 pg / mL (S / N=3).

[0109] Feasibility study

[0110] To verify the feasibility of using the Ti3C2-based nanocomposite material designed in this invention to detect actual samples, as shown in Table 1, IAA was extracted from leaves of different plants and applied to detect IAA in extracts of Arabidopsis thaliana and rice leaves. The measured IAA values ​​were then compared with reference values ​​obtained by LC-MS / MS using the sensor designed in this paper.

[0111] The extraction of IAA from plant leaves includes the following steps:

[0112] Fresh plant leaves were selected and ground into powder using liquid nitrogen. 100 mg of the powder was accurately weighed and added to 1 mL of 80% methanol solution. The mixture was stirred and extracted at 4°C for 12 h. The mixture was then centrifuged at 12000 rpm for 10 min at 4°C. The supernatant was collected, and the extraction was repeated twice. The supernatants were mixed, and all supernatants were placed in the same tube. The tube opening was covered with a dialysis membrane, with a small hole left in the cap to facilitate water evaporation. The tube was then dried in a vacuum concentrator. The concentrated sample was reconstituted with 20% methanol for analysis.

[0113] Table 1. Comparison of IAA detection methods in actual samples (n≥5)

[0114]

[0115] As shown in the table, the relative deviation between the measured values ​​of IAA in rice extract obtained by the biosensor based on Ti3C2 nanocomposite material of this invention and the reference values ​​obtained by LC-MS / MS method was 3.50%, and the relative deviation between the measured values ​​of IAA in Arabidopsis thaliana extract obtained by the biosensor based on Ti3C2 nanocomposite material of this invention and the reference values ​​obtained by LC-MS / MS method was 7.77%. This indicates that the method can be used to analyze IAA in actual plant samples.

[0116] Selective research

[0117] Good selectivity is an important performance characteristic of electrochemical biosensors for detecting IAA. Therefore, to evaluate the selectivity of the biosensor of the present invention, which is loaded with a Ti3C2-based nanocomposite material, gibberellin 200 ng mL was used to detect IAA. -1 The mixture was incubated with 200 ng / mL 1-Naphthylacetic acid, 200 ng / mL salicylic acid, 200 ng / mL jasmonic acid, 200 ng / mL indole-3-butyric acid, and 10 ng / mL indoleacetic acid (IAA). Figure 5 As shown, the ΔI values ​​for gibberellin, naphthaleneacetic acid, salicylic acid, jasmonic acid, indolebutyric acid, and IAA were 2, 2, 1, 2, 1.5, and 14, respectively. Clearly, IAA caused a significant change in the electrochemical signal, with a ΔI value reaching 14. This indicates the good selectivity of the biosensor prepared based on the IAA-specific antibody-antigen reaction.

[0118] In summary, this invention uses Ti3C2 as a template material and employs a self-assembly method to prepare a β-cyclodextrin-metal-organic framework / rGO / Ti3C2 composite nanomaterial modified electrode, with Fe(CN)6 as the template material. 3− / 4- As a probe, the IAA antibody specifically binds to IAA, reducing Fe(CN)6. 3− / 4- The DPV peak current was used to achieve highly sensitive detection of IAA. The constructed biosensor exhibits high selectivity and good stability, with a linear range of 0.01 ng / mL–10 ng / mL, a detection limit of 2.88 pg / mL (S / N=3), and a recovery rate of 88.52%–102.51%. It has been successfully applied in rice and Arabidopsis thaliana plant samples, providing strong methodological support for the electrochemical biosensor detection of auxin.

[0119] The above are merely embodiments of the present invention and do not limit the patent scope of the present invention. Any equivalent modifications made based on the content of the present invention specification, or direct or indirect applications in related technical fields, are similarly included within the patent protection scope of the present invention.

Claims

1. A Ti3C2-based nanocomposite, characterized by: Includes the following components: Ti3C2, reduced graphene oxide, β-cyclodextrin-metal-organic framework materials, IAA antibodies and adhesives; The adhesive includes glutaraldehyde and chitosan, wherein the glutaraldehyde is used to immobilize IAA antibodies, and the chitosan is used to adhere nanomaterials and immobilize glutaraldehyde. The mass ratio of Ti3C2 to reduced graphene oxide is 10-20:2-4; The Ti3C2 nanocomposite material was prepared using the following method: S1. Ti3C2-rGO composite material was prepared by mixing Ti3C2 and reduced graphene oxide; β-cyclodextrin and metal materials were mixed to prepare β-cyclodextrin-metal-organic framework material. S2 mixed Ti3C2-rGO composite material, β-cyclodextrin-metal-organic framework material and adhesive to obtain composite nanomaterials; S3. IAA antibody is added to the surface of the dried composite nanomaterial to obtain the Ti3C2-based nanocomposite material.

2. The Ti3C2-based nanocomposite of claim 1, wherein: In the β-cyclodextrin-metal-organic framework material, β-cyclodextrin is used as the organic ligand and metal ions are used as the central ions, wherein the metal ions include at least one of potassium ions and sodium ions.

3. The Ti3C2-based nanocomposite material according to claim 1, characterized in that: In the Ti3C2-based nanocomposite material, the ratio of the total mass of Ti3C2, reduced graphene oxide, and β-cyclodextrin-metal-organic framework material to the mass of IAA antibody is 12-24:6-12.

4. The Ti3C2-based nanocomposite material according to claim 1, characterized in that: Step S3 also includes the following steps: after adding IAA antibody to the surface of the dried composite nanomaterial, it is first incubated at 3-4℃, then bovine serum albumin is added and incubated at 3-4℃.

5. A modified electrode, characterized in that: It includes an electrode and a material modified on the electrode, wherein the material modified on the electrode includes a Ti3C2-based nanocomposite material as described in any one of claims 1 to 4.

6. A modified electrode according to claim 5, characterized in that: The material modified on the electrode also includes bovine serum albumin.

7. The application of a Ti3C2-based nanocomposite material as described in any one of claims 1 to 4 in the preparation of electrochemical sensors or plant growth hormone detection.

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