A flexible sensor for real-time monitoring of gallic acid content in plants in vivo, its preparation method and application

Abstract

This invention belongs to the field of micro-electromechanical biosensing technology, specifically relating to a flexible sensor for real-time monitoring of gallic acid content in plants, its preparation method, and its application. The flexible sensor includes a flexible substrate and a working electrode, a reference electrode, and a counter electrode disposed on the substrate. The working electrode is a laser-induced graphene electrode (LIG) with its surface sequentially modified with black phosphorus nanosheets (BP), sheet-like titanium carbide (MXene), and manganese phosphorus sulfur (MnPS3). This flexible sensor can achieve online detection and analysis of gallic acid in plant tissues at different stages. Because it is a patch-type adaptive sensor for plant surfaces, it does not cause fundamental damage to the detected material, allowing the material to continue growing until maturity. Furthermore, the layer-by-layer assembly of the working electrode with black phosphorus nanosheets, MXene, and MnPS3 achieves highly sensitive detection of gallic acid, offering advantages such as lightweight portability, excellent electrical performance, and high integration.

Description

Technical Field

[0001] This invention belongs to the field of micro-motor biosensing technology, specifically relating to a flexible sensor for real-time monitoring of gallic acid content in living plants, its preparation method, and its application. Background Technology

[0002] Gallic acid is a polyphenolic organic compound widely distributed in nature. It is one of the main active ingredients in many plants such as olives, mangoes, and grapes, possessing various biological activities including anti-inflammatory, anti-mutagenic, antioxidant, and antiviral properties. Furthermore, gallic acid and other phenolic substances have significant inhibitory effects on common foodborne pathogens and spoilage bacteria. Therefore, detecting gallic acid content to select disease-resistant varieties is a relatively effective method for controlling bacterial leaf streak in rice. Gallic acid content is positively correlated with the quality grade of green tea; therefore, gallic acid can be used as a quality evaluation indicator for green tea. The detection of gallic acid is of great significance for the quality evaluation and control of agricultural products and food.

[0003] Currently, researchers both domestically and internationally have developed numerous methods for detecting gallic acid, including chromatography, spectroscopy, and capillary electrophoresis. These methods each possess unique characteristics in terms of sensitivity and stability. However, the instruments used for these methods are expensive, and the requirements for testing personnel are high. Furthermore, they are all in vitro detection methods, requiring sample collection and pretreatment. The collection process can easily damage the plant, while the pretreatment process is complex and time-consuming. Compared to other technologies, electrochemical methods offer advantages such as simple operation, fast response time, time savings, high sensitivity, and ease of integration, and can meet the needs of in-situ detection of living plants. However, traditional sensors based on rigid electrodes (such as glassy carbon electrodes and gold electrodes) cannot achieve effective adhesion to the plant surface, reducing the reliability and accuracy of the detection results. On the other hand, they are difficult to fix on the plant surface for long-term detection. Outdoor environments such as wind, rain, and lightning can easily cause deformation of plant samples, leading to additional damage to the plant from the rigid electrode. Therefore, traditional rigid sensors are difficult to meet the needs of long-term in-situ, fixed-point, and continuous monitoring of plant physiology. Summary of the Invention

[0004] To address the problems existing in the prior art, the purpose of this invention is to provide a flexible sensor for real-time monitoring of gallic acid content in plants, its preparation method, and its application.

[0005] To achieve the above objectives, the present invention adopts the following technical solution:

[0006] A flexible sensor includes a flexible substrate and a working electrode, a reference electrode, and a counter electrode disposed on the flexible substrate;

[0007] The working electrode is a laser-induced graphene electrode (LIG) whose surface is sequentially modified with black phosphorus nanosheets (BP), sheet-like titanium carbide (MXene), and manganese phosphorus sulfur (MnPS3).

[0008] The reference electrode is a laser-induced graphene electrode (LIG) with Ag / AgCl surface modification;

[0009] The counter electrode is a laser-induced graphene electrode (LIG).

[0010] This invention uses a flexible substrate, and the electrode pattern on the flexible substrate is laser-printed graphene (LIG). The resulting flexible electrode has a soft, flexible appearance and a bio-adaptive ability that better fits the shape of the leaf. It can overcome the shortcomings of existing technologies that damage plant samples and can only perform static detection in vitro, and realize long-term, stable and reliable monitoring of living plants. This invention utilizes a laser-induced graphene (LIG) electrode, sequentially modified with black phosphorus nanosheets (BP), sheet-like titanium carbide (MXene), and manganese phosphorus sulfur (MnPS3), as the working electrode for monitoring gallic acid in plants. Two-dimensional black phosphorus nanosheets (BP) possess excellent properties such as large specific surface area, high carrier transport rate and conductivity, easy surface modification, and high catalytic activity, playing a crucial role in electrochemical sensing. MXene, a novel two-dimensional metal carbide, exhibits excellent conductivity, ease of functionalization and large-scale production, high hydrophilicity, and provides a large active surface for loading electroactive molecules. Its good biocompatibility enhances the stability of the electrochemical sensor, making it an ideal material for constructing high-performance electrochemical sensors. MnPS3 has extremely low conductivity, enabling high response even in sensing environments where conductivity changes can be significant. Modifying the LIG electrode substrate with these three nanomaterials improves electrode conductivity, enhances the electrochemical signal, accelerates the reaction rate, and achieves rapid and accurate detection of gallic acid content in living plants.

[0011] Preferably, the working electrode, reference electrode, and counter electrode are all musical note-shaped. Musical note-shaped flexible electrodes feature a large electrode area, good conductivity, aesthetic appeal, simple structure, and compact, integrated design. Furthermore, this shape allows for greater spacing between electrodes, facilitating modification operations and practical applications.

[0012] Further preferably, the musical note shape has a length of 45-50 mm and a width of 20-25 mm.

[0013] Preferably, in the working electrode, the mass ratio of black phosphorus nanosheets (BP), sheet-like titanium carbide (MXene), and manganese phosphorus sulfur (MnPS3) modified on the surface of the laser-induced graphene electrode (LIG) is 0.2–0.6:8–12:1. To optimize sensor performance, the inventors optimized the sensor fabrication conditions. Among the various proportions of the black phosphorus nanosheets (BP), sheet-like titanium carbide (MXene), and manganese phosphorus sulfur (MnPS3) composite material, the highest current response was obtained when the ratio was 0.2–0.6:8–12:1, indicating that the three materials exhibit optimal synergy, allowing the sensor to perform at its maximum potential.

[0014] Preferably, the flexible substrate is made of polydimethylsiloxane (PDMS).

[0015] The present invention also provides a method for fabricating the above-mentioned flexible sensor, comprising:

[0016] Preparation of counter electrode: Laser-induced graphene electrode (LIG) is prepared on polyimide film (PI film), and then transferred to polydimethylsiloxane flexible substrate by polydimethylsiloxane (PDMS) transfer strategy to form counter electrode;

[0017] Preparation of reference electrode: Laser-induced graphene electrode (LIG) is prepared on polyimide film (PI film). The LIG is transferred to a polydimethylsiloxane flexible substrate using a polydimethylsiloxane (PDMS) transfer strategy. Then, Ag / AgCl silver paste is coated on the LIG and cured to form a reference electrode.

[0018] Preparation of working electrode: Laser-induced graphene electrode (LIG) is prepared on polyimide film (PI film). The LIG is transferred to a polydimethylsiloxane (PDMS) flexible substrate using a polydimethylsiloxane (PDMS) transfer strategy. Then, black phosphorus nanosheets (BP), sheet-like titanium carbide (MXene), and manganese phosphorus sulfur (MnPS3) are sequentially coated on the LIG to form the working electrode.

[0019] Preferably, the step of sequentially coating the laser-induced graphene electrode (LIG) with black phosphorus nanosheets (BP), sheet-like titanium carbide (MXene), and manganese phosphorus sulfur (MnPS3) specifically includes:

[0020] A black phosphorus nanosheet dispersion was drop-coated onto a laser-induced graphene electrode (LIG), and then dried to obtain a BP / LIG electrode.

[0021] A flake-shaped titanium carbide dispersion was drop-coated onto the BP / LIG electrode and dried to obtain an MXene / BP / LIG electrode.

[0022] A manganese-phosphorus-sulfur (MnPS3) colloidal solution was drop-coated onto the MXene / BP / LIG electrode and dried to obtain the MnPS3 / MXene / BP / LIG electrode.

[0023] More preferably, the concentration of the black phosphorus nanosheet dispersion is 0.1–0.3 mg / mL;

[0024] The concentration of the flake-shaped titanium carbide dispersion is 3-5 mg / mL;

[0025] The concentration of the manganese-phosphorus-sulfur (MnPS3) colloidal solution is 0.4–0.6 mg / mL. This preferred concentration ensures the uniformity of the film obtained after drying and further enhances the electrochemical signal.

[0026] More preferably, the solvent for the manganese-phosphorus-sulfur (MnPS3) colloidal solution is an aqueous ethanol solution.

[0027] A further preferred embodiment includes: drop-coating a Nafion solution onto a MnPS3 / MXene / BP / LIG electrode and drying it to obtain a Nafion / MnPS3 / MXene / BP / LIG electrode.

[0028] The present invention also provides the application of the above-mentioned flexible sensor in real-time monitoring of gallic acid content in living plants.

[0029] This invention also provides a method for real-time monitoring of gallic acid content in plants, comprising:

[0030] 1) Prepare a series of gallic acid-phosphate buffer solutions of different concentrations, and use the above-mentioned flexible sensor to perform differential pulse voltammetry to obtain a set of relationship curves between concentration and peak current after deducting background current, and make working curves.

[0031] 2) Attach the above-mentioned flexible sensor to the test site of the plant, connect it to the electrochemical workstation, perform differential pulse voltammetry scanning, and calculate the instantaneous concentration of gallic acid in the test site of the plant by using the obtained current signal through the working curve.

[0032] In the above method, the plant to be tested includes crops, flowers, vegetables, fruits, etc., and the part to be tested can be the root, stem, leaf, fruit, etc. of the plant. It can be applied to different stages of plant growth and different environments.

[0033] The beneficial effects of this invention are as follows:

[0034] The flexible sensor provided by this invention enables real-time monitoring of gallic acid content in plants, allowing for online detection and analysis of gallic acid in plant tissues at different stages. Because it is a patch-type sensor that adapts to the plant surface, it does not cause fundamental damage to the material being tested, allowing the material to continue growing until maturity. Furthermore, by assembling the working electrode layer by layer using black phosphorus nanosheets, MXene, and MnPS3, highly sensitive detection of gallic acid is achieved, offering advantages such as lightweight portability, excellent electrical performance, and high integration. Attached Figure Description

[0035] To more clearly illustrate the technical solutions in the embodiments of the present invention or the prior art, the drawings used in the description of the prior art in the embodiments will be briefly introduced below. Obviously, the drawings described below are some embodiments of the present invention. For those skilled in the art, other drawings can be obtained from these drawings without creative effort.

[0036] Figure 1 This is a schematic diagram of the electrodes of the flexible sensor described in this invention.

[0037] Figure 2 This is a schematic diagram illustrating the fabrication of the working electrode of the flexible sensor described in this invention.

[0038] Figure 3 The flexible electrode described in this invention is in 1mM [Fe(CN)6] 4- Cyclic voltammetry curves in solution. Detailed Implementation

[0039] The following examples are used to illustrate the present invention, but are not intended to limit the scope of the invention. Any modifications or substitutions made to the methods, steps, or conditions of the present invention without departing from the spirit and essence of the invention are within the scope of the invention.

[0040] In the following examples, information on the reagents and instruments used is shown in Table 1.

[0041] Table 1. Information on reagents used in the embodiments.

[0042]

[0043] Example 1: Flexible Sensor and Its Fabrication

[0044] See parts Figure 1 , Figure 2 This embodiment provides a flexible sensor, including a PDMS flexible substrate and a note-shaped working electrode, a note-shaped reference electrode and a note-shaped counter electrode disposed on the PDMS flexible substrate, which are about 45 mm long and about 20 mm wide.

[0045] The musical note-shaped working electrode is a laser-induced graphene electrode (LIG) whose surface is sequentially modified with black phosphorus nanosheets (BP), sheet-like titanium carbide (MXene), and manganese phosphorus sulfur (MnPS3).

[0046] The musical note-shaped reference electrode is a laser-induced graphene electrode (LIG) with Ag / AgCl surface modification.

[0047] The musical note-shaped electrode is a laser-induced graphene electrode (LIG).

[0048] The flexible sensor provided in this embodiment is fabricated as follows:

[0049] (1) Polyimide tape (PI) is attached to the glass and then secured with a polymer methyl methacrylate (PMMA) mold. The PI tape is cleaned with distilled water and ethanol. Then, the PI tape is patterned using a computer-controlled laser (laser-induced graphene technology) to obtain three musical note-shaped LIG electrodes.

[0050] (2) Using a spin coater, liquid polydimethylsiloxane (PDMS) (main agent A and curing agent B mixed in a 10:1 ratio and stirred for 10 min) was uniformly applied to a patterned PI tape at 100 rpm for 6 s. The tape was then heated in a vacuum drying oven at 100°C for 10 h. One side of the PDMS / LIG was then peeled off from the PI film to obtain a flexible and stretchable musical note-shaped LIG electrode. The conductive region in the middle of the musical note-shaped LIG electrode was insulated by a liquid PDMS coating and the same heat treatment.

[0051] (3) The musical note-shaped LIG electrode on the left is the counter electrode. The Ag / AgCl silver paste is coated on the musical note-shaped LIG electrode on the right and heated to solidify to make the Ag / AgCl reference electrode. Then the microelectrode is placed in a 0.5M dilute sulfuric acid solution for cyclic voltammetry scanning (0-1.5V) to obtain a typical cyclic voltammetric spectrum, ensuring that the electrode surface is clean.

[0052] (4) 10 μL of BP nanosheet dispersion with a concentration of 0.2 mg / mL was dropped onto the middle phonograph-shaped LIG electrode to obtain the BP / LIG electrode.

[0053] (5) Then, 10 μL of MXene dispersion with a concentration of 5 mg / mL was drop-coated onto the BP / LIG electrode to obtain the MXene / BP / LIG electrode.

[0054] (6) By sonicating MnPS3 crystals in an ethanol / water (volume ratio 2:3) mixture for 2 hours, the bulk MnPS3 crystals were peeled into several layers. The unpeeled material was removed by centrifugation to obtain an MnPS3 colloidal solution. 10 μL of 0.5 mg / mL MnPS3 colloid was dropped onto an MXene / BP / LIG electrode.

[0055] (7) Finally, 5 μL of 0.5% Nafion solution was dropped onto the MnPS3 / MXene / BP / LIG electrode and encapsulated to obtain the Nafion / MnPS3 / MXene / BP / LIG electrode, which is the working electrode.

[0056] Example 2: Application of Flexible Sensors

[0057] The experimental material was potted grapes, with grape leaves from the flowering to fruiting period as the test object.

[0058] (1) Gallic acid-phosphate buffer solutions with concentrations of 0, 0.1, 0.5, 1, 5, 10, 50, 100, and 1000 μM (pH = 4.5) were prepared. Differential pulse voltammetry was used to detect these solutions using the flexible sensor prepared in Example 1 (potential -0.4 to 0.8 V, potential increase 0.004 V, amplitude 0.05 V, pulse width 0.02 s, pulse period 0.5 s, rest time 20 s). A set of curves showing the relationship between concentration and the spike potential after deducting the background current were obtained. A curve for the flexible sensor was constructed, and the linear equation was I. p =0.572C + 1.398 × 10 -7 (Current unit μA, concentration unit μM), linear range up to 0.1-1000μM.

[0059] (2) The flexible sensor was randomly attached to the grape leaf and connected to the electrochemical workstation. The current signal was recorded by differential pulse voltammetry and the gallic acid concentration of the living grape leaf was calculated by the working curve.

[0060] Comparison of experimental results

[0061] Grape leaves from two different varieties, Kyoho and Summer Black, collected at the same time, were analyzed for gallic acid content using high-performance liquid chromatography (HPLC). The results were compared with those obtained using the electrochemical method of this invention, as shown in the table below. The results indicate that the flexible electrode sensing method provides reliable detection results.

[0062] Table 2 Comparison of Test Results

[0063]

[0064] Comparative Example 1

[0065] The difference from Example 1 is that the drop-coating modification of MnPS3 is omitted, and only BP and MXene are drop-coated.

[0066] The fabrication process of the sensor was characterized using cyclic voltammetry (CV). The CV scans were performed at 1 mM [Fe(CN)6]. 4-The CV scan was performed in a solution (containing 0.1 mol KCl). The current response of the CV scan is as follows: Figure 3 As shown, compared to MXene / BP / LIG, the redox peak current increases and the peak-to-peak potential difference decreases when MnPS3 is modified onto MXene / BP / LIG. This is because the high conductivity and catalytic performance of MnPS3 improve the electron transfer rate and enhance reversibility. This indicates that only a combination of these three materials can form a three-dimensional network structure with a large specific surface area and excellent electrical conductivity.

[0067] The above embodiments are merely descriptions of preferred embodiments of the present invention and are not intended to limit the scope of the present invention. Various modifications and improvements made by those skilled in the art to the technical solutions of the present invention without departing from the spirit of the present invention should fall within the protection scope defined by the claims of the present invention.

Claims

1. A flexible sensor, characterized by, It includes a flexible substrate and a working electrode, a reference electrode, and a counter electrode disposed on the flexible substrate; The working electrode is a laser-induced graphene electrode (LIG) whose surface is sequentially modified with black phosphorus nanosheets (BP), sheet-like titanium carbide (MXene), and manganese phosphorus sulfur (MnPS3). The reference electrode is a laser-induced graphene electrode (LIG) with Ag / AgCl surface modification; The counter electrode is a laser-induced graphene electrode (LIG).

2. The flexible sensor of claim 1, wherein, The working electrode, reference electrode, and counter electrode are all shaped like musical notes.

3. The flexible sensor according to claim 1 or 2, characterized in that, In the working electrode, the mass ratio of black phosphorus nanosheets (BP), sheet-like titanium carbide (MXene), and manganese phosphorus sulfur (MnPS3) modified on the surface of the laser-induced graphene electrode (LIG) is 0.2~0.6:8~12:

1.

4. The flexible sensor according to claim 1 or 2, characterized in that, The flexible substrate is made of polydimethylsiloxane.

5. The method for fabricating the flexible sensor according to any one of claims 1-4, characterized in that, include: Preparation of counter electrode: Laser-induced graphene electrode (LIG) is prepared on polyimide film, and then transferred to polydimethylsiloxane flexible substrate by polydimethylsiloxane transfer strategy to form counter electrode; Preparation of reference electrode: Laser-induced graphene electrode (LIG) is prepared on polyimide film. The LIG is then transferred to a polydimethylsiloxane flexible substrate using a polydimethylsiloxane transfer strategy. Ag / AgCl silver paste is then coated onto the LIG and cured to form the reference electrode. Preparation of working electrode: Laser-induced graphene electrode (LIG) is prepared on polyimide film. The LIG is then transferred onto a polydimethylsiloxane flexible substrate using a polydimethylsiloxane transfer strategy. Black phosphorus nanosheets (BP), sheet-like titanium carbide (MXene), and manganese phosphorus sulfur (MnPS3) are then sequentially coated onto the LIG to form the working electrode.

6. The preparation method according to claim 5, characterized in that, The process of sequentially coating black phosphorus nanosheets (BP), sheet-like titanium carbide (MXene), and manganese phosphorus sulfur (MnPS3) onto a laser-induced graphene electrode (LIG) specifically includes: A black phosphorus nanosheet dispersion was drop-coated onto a laser-induced graphene electrode (LIG), and then dried to obtain a BP / LIG electrode. A flake-shaped titanium carbide dispersion was drop-coated onto the BP / LIG electrode and dried to obtain an MXene / BP / LIG electrode. A manganese-phosphorus-sulfur (MnPS3) colloidal solution was drop-coated onto the MXene / BP / LIG electrode and dried to obtain the MnPS3 / MXene / BP / LIG electrode.

7. The preparation method according to claim 6, characterized in that, The concentration of the black phosphorus nanosheet dispersion is 0.1~0.3 mg / mL; The concentration of the flake-shaped titanium carbide dispersion is 3~5 mg / mL; The concentration of the manganese-phosphorus-sulfur (MnPS3) colloidal solution is 0.4~0.6 mg / mL.

8. The preparation method according to claim 6 or 7, characterized in that, Also includes: A Nafion / MnPS3 / MXene / BP / LIG electrode was prepared by drop-coating Nafion solution onto a MnPS3 / MXene / BP / LIG electrode and then drying it.

9. The application of the flexible sensor according to any one of claims 1-4 or the flexible sensor prepared by the preparation method according to any one of claims 5-8 in real-time monitoring of gallic acid content in plants in vivo.

10. A method for real-time monitoring of gallic acid content in plants, characterized in that, include: 1) Prepare a series of gallic acid-phosphate buffer solutions of different concentrations, and use the flexible sensor according to any one of claims 1-4 or the flexible sensor prepared by the preparation method according to any one of claims 5-8 to perform differential pulse voltammetry detection, and obtain a set of relationship curves between concentration and peak current after deducting background current, and make working curves. 2) Attach the flexible sensor according to any one of claims 1-4 or the flexible sensor prepared by the preparation method according to any one of claims 5-8 to the test site of the plant to be tested, connect it to an electrochemical workstation, perform differential pulse voltammetry scanning, and calculate the instantaneous concentration of gallic acid in the test site of the plant to be tested by the obtained current signal through the working curve.

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