Working electrode for electrochemically detecting palmitic acid, preparation method of working electrode and electrochemical sensor and detection system based on working electrode

Abstract

The application belongs to the technical field of fatty acid detection, and particularly relates to a working electrode for electrochemical detection of palmitic acid, a preparation method of the working electrode, and an electrochemical sensor and a detection system based on the working electrode. x MXene is mixed together to carry out a polymerization reaction to obtain a polymer; template molecules in the polymer are eluted to obtain a molecularly imprinted polymer; 2) a gold nanoparticle, an electrochemical signal probe, graphene oxide and a multi-walled carbon nanotube composite material, and the molecularly imprinted polymer are sequentially modified on the surface of a screen-printed electrode to obtain the working electrode. The working electrode for electrochemical detection of palmitic acid, the electrochemical sensor and the portable detection system are prepared by a molecular imprinting method for the first time, and are low in equipment cost, sensitive, rapid, excellent in selectivity and stability.

Description

TECHNICAL FIELD

[0001] The application belongs to the technical field of fatty acid detection, and particularly relates to a working electrode for electrochemical detection of palmitate acid, a preparation method of the working electrode, and an electrochemical sensor and a detection system based on the working electrode. BACKGROUND

[0002] Fats mainly play a role in maintaining normal physiological activities of the human body and maintaining nutritional balance in the human body, can be used as a solvent for some fat-soluble nutrients, and also have certain immune effects. Most of the fatty acids required in the human body are produced by hydrolysis of fats, among which saturated fatty acids mainly provide energy for the human body, and also have the effects of providing cholesterol and neutral fat, and if the intake is insufficient, diseases such as anemia, blood vessel fragility, and cerebral hemorrhage are easily induced. Unsaturated fatty acids also have high nutritional value, including the effects of regulating blood sugar, antioxidant, and preventing atherosclerosis. Therefore, rapid and sensitive detection of fatty acids is very important.

[0003] At present, the detection methods of fatty acids mainly include high performance liquid chromatography (HPLC), liquid chromatography-mass spectrometry (LC-MS), gas chromatography-mass spectrometry (GC-MS), biochemical method, and gas chromatography, each of which has its own characteristics and applicable conditions. However, these methods require expensive testing instruments, have certain requirements for time and personnel, and the pretreatment process is complex. Compared with other technologies, the electrochemical method has the advantages of simple operation, fast response time, time saving, high sensitivity, and easy integration, and is suitable for constructing a portable detection device for rapid detection of fatty acids. SUMMARY

[0004] Palmitate acid (PA) is the most abundant free fatty acid in plasma lipids, is a saturated higher fatty acid, and widely exists in the form of glyceride in animal and vegetable oils. The purpose of the present application is to provide a working electrode for electrochemical detection of palmitate acid, a preparation method of the working electrode, and an electrochemical sensor and a detection system based on the working electrode, so as to overcome the shortcomings of high detection cost, high requirement for detection personnel, long time consumption, and complex operation in the prior art, and realize rapid and sensitive detection of palmitate acid in different samples.

[0005] To achieve the above-mentioned purpose, the technical scheme adopted by the present application is as follows:

[0006] The present application first provides a preparation method of a working electrode for electrochemical detection of palmitate acid, comprising the following steps:

[0007] 1) mixing palmitate acid (template molecule), dopamine (functional monomer), and Ti3C2T MXene together to perform a polymerization reaction to obtain a polymer; eluting the template molecule in the polymer to obtain a molecularly imprinted polymer (MIP-MXene); x ​

[0008] 2) sequentially modifying a gold nanoparticle layer, an electrochemical signal probe layer, a graphene oxide and multi-walled carbon nanotube composite layer, and the molecularly imprinted polymer layer on the surface of the screen-printed electrode (SPE) to obtain the MIP-MXene / Fc-GO-MWNT / AuNPs / SPE electrode.

[0009] Preferably, the palmitic acid, the dopamine and the Ti3C2T x The mass ratio of the MXene is 1-4:4-1:2.5; more preferably 1:2:2.5.

[0010] Preferably, in step 1), the polymerization reaction is carried out at a pH of 7.5-10.5; more preferably at a pH of 8.5.

[0011] Preferably, in step 1), the polymerization reaction is carried out for 20-48 hours.

[0012] Preferably, in step 1), the eluent for elution is selected from at least one of an acetic acid solution with a concentration of 1-20%, acetonitrile, methanol, and ethanol; more preferably an acetic acid solution with a concentration of 5%.

[0013] Preferably, the elution is carried out for 5-25 minutes, more preferably 15 minutes.

[0014] Preferably, in step 2), the electrochemical signal probe is selected from at least one of ferrocene (Fc), thionine, and methylene blue; more preferably ferrocene.

[0015] Preferably, step 2) specifically comprises:

[0016] Gold nanoparticles are deposited on the screen-printed electrode (SPE) by electrodeposition to obtain an AuNPs / SPE electrode;

[0017] A mixture of an electrochemical signal probe, graphene oxide, multi-walled carbon nanotubes, and Nafion solution is drop-coated on the AuNPs / SPE electrode, and dried to obtain a Fc-GO-MWNT / AuNPs / SPE electrode;

[0018] A molecularly imprinted polymer dispersion is drop-coated on the Fc-GO-MWNT / AuNPs / SPE electrode, and dried to obtain a MIP-MXene / Fc-GO-MWNT / AuNPs / SPE electrode.

[0019] Further preferably, the voltage for electrodeposition is -1 V, and the time is 400-1200 seconds (more preferably 800 seconds).

[0020] Further preferably, the concentration of the electrochemical signal probe in the mixture of the electrochemical signal probe, graphene oxide, multi-walled carbon nanotubes and Nafion solution is 6-14 mg / mL, more preferably 10 mg / mL.

[0021] Further preferably, the concentration ratio of graphene oxide to multi-walled carbon nanotubes in the mixture of the electrochemical signal probe, graphene oxide, multi-walled carbon nanotubes and Nafion solution is 1-3:3-1, more preferably 1:1.

[0022] Further preferably, the concentration of the molecularly imprinted polymer dispersion is 1-5 mg / mL, more preferably 2 mg / mL.

[0023] The application also provides a working electrode for electrochemically detecting palmitic acid, which is prepared by the above method.

[0024] The application also provides an electrochemical sensor for electrochemically detecting palmitic acid, which comprises the above working electrode, a carbon electrode as a counter electrode, and a silver / silver chloride electrode as a reference electrode.

[0025] The application also provides a portable detection system for electrochemically detecting palmitic acid, which comprises the above electrochemical sensor, a DPV detection device and a smart device.

[0026] The electrochemical sensor is connected to the DPV detection device.

[0027] The smart device is used to receive electrochemical experimental data of the DPV detection device and calculate the concentration of palmitic acid.

[0028] The DPV detection device comprises:

[0029] A detection cell is used to place a solution to be detected.

[0030] A power module is used to provide power for the DPV detection device.

[0031] A DPV waveform generation circuit is used to provide a scanning voltage for the electrochemical sensor.

[0032] A micro-current detection module is used to collect electrochemical experimental data of the electrochemical sensor.

[0033] A constant potential circuit is used to apply a voltage transmitted from the DPV waveform generation circuit between the working electrode and the reference electrode of the electrochemical sensor, so as to ensure that the voltage meets the requirements.

[0034] A core processor is used to transmit the electrochemical experimental data to the smart device.

[0035] Preferably, the core processor is STM32F103ZET6.

[0036] Preferably, the intelligent device is a smart phone, and the core processor transmits electrochemical experiment data to the smart phone through wireless communication. The wireless communication can be Wifi or Bluetooth. The smart phone realizes the bidirectional transmission of configuration parameter data and signal data between the upper computer and the lower computer through wireless communication. The configuration parameters can be set according to the actual needs of the electrochemical pulse voltammetry scanning parameters. The data processing module in the smart phone can perform baseline calibration processing and calculate the concentration value of the sample to be measured according to the standard working curve. The data storage module is also included to save the configuration parameters and signal data.

[0037] The working electrode for electrochemical detection of palmitic acid, the preparation method thereof, and the electrochemical sensor and detection system based on the working electrode are provided. Palmitic acid is taken as the research object, a palmitic acid sensor is constructed by using a molecular imprinting method, and the palmitic acid in a sample is detected by differential pulse voltammetry (DPV) through an electrochemical method. The beneficial effects are as follows:

[0038] (1) The base electrode used in the application is a disposable screen-printed electrode. The screen-printed electrode is modified with gold nanoparticles, an electrochemical signal probe, a composite material of graphene oxide and multi-walled carbon nanotubes, and a molecular imprinting polymer in sequence. The obtained sensor has excellent selectivity and stability performance, and the cost is low.

[0039] (2) The application uses palmitic acid as a template molecule, dopamine as a functional monomer, and Ti3C2T x MXene to self-polymerize, and then elutes the template molecule to prepare a molecular imprinting polymer (MIP-MXene). The reaction conditions are mild, green and environmentally friendly.

[0040] (3) The application first realizes the preparation of a palmitic acid electrochemical sensor and a portable detection system by a molecular imprinting method. The device has low cost, high sensitivity and fast detection.

[0041] (4) The application can be applied to detect different samples, such as serum, urine, and tissue samples after stirring and centrifugation. BRIEF DESCRIPTION OF DRAWINGS

[0042] In order to more clearly illustrate the embodiments of the application or the technical solutions in the prior art, the drawings needed in the prior art description in the embodiments will be briefly introduced below. Obviously, the drawings in the following description are some embodiments of the application, and those skilled in the art can also obtain other drawings according to these drawings without creative labor.

[0043] Figure 1 The modification process of the working electrode of the present application is shown in the figure.

[0044] Figure 2 The structure diagram of the portable detection system of the present application is shown in the figure.

[0045] Figure 3 The PA detection comparison diagram of the MIP of Example 1 and the NIP of Comparative Example 1 is shown in the figure. DETAILED DESCRIPTION

[0046] The following examples are used to illustrate the present application, but not to limit the scope of the present application. Modifications or replacements of the methods, steps or conditions of the present application, without departing from the spirit and essence of the present application, all belong to the scope of the present application.

[0047] In the following examples, the information of each reagent and instrument used is shown in Table 1.

[0048] Table 1 Information of reagents used in examples

[0049]

[0050] Example 1

[0051] The preparation method of the working electrode for electrochemical detection of palmitic acid can refer to Figure 1 , comprising the following steps:

[0052] (1) Preparation of molecularly imprinted polymer: first, 20 mg of palmitic acid and 40 mg of dopamine are dissolved in 20 ml of 95% anhydrous ethanol, and the mixture is placed in a clean conical flask and ultrasonically cleaned in ice water for 1 hour to obtain a palmitic acid-dopamine mixed solution. Then 50 mg of Ti3C2Tx MXene is dissolved in 30 ml of 95% anhydrous ethanol in a conical flask, and ultrasonically dispersed at room temperature for 2 hours to obtain a MXene suspension. Then the MXene suspension is added to the palmitic acid-dopamine mixed solution, and Tris-hydrochloric acid buffer is added dropwise to make the pH of the whole solution system 8.5. Put the magnetic stirrer rotor into the conical flask, then wrap the conical flask with tin foil paper for light protection, and place it on the magnetic stirrer for sufficient stirring for 24 hours. After removing the tin foil paper, pour the solution into a centrifuge tube, and place it in a centrifuge for centrifugal treatment, with the centrifugal parameters set at 25℃, 8000r, 10min. After centrifugation, the centrifuge tube is taken out gently, the supernatant is removed, and then the solid is taken out. Then use ultrapure water and ethanol to wash the solid to wash away the dopamine that has not been fully reacted in the solid. Then use the eluent to extract the template molecule, i.e. palmitic acid, in the molecularly imprinted polymer. The eluent is 5% acetic acid solution, and the elution time can be 15 minutes. Finally, the molecularly imprinted polymer wrapped on MXene (MIP-MXene) is prepared.

[0053] (2) Clean the electrode surface with ultrapure water and anhydrous ethanol, then put the screen-printed electrode into a 0.5 g / L chloroauric acid (HAuCl4·4H2O) solution, and use chronoamperometry to electrodeposit gold nanoparticles on the surface of the working electrode, with the voltage parameter set to -1 V and the time parameter set to 800 s, to obtain an AuNPs / SPE electrode.

[0054] (3) Mix Fc, GO (graphene oxide), MWNT, and 5 wt% Nafion, with the concentration of Fc being 10 mg / mL, the concentration of GO being 1 mg / mL, and the concentration of MWNT being 1 mg / mL. Ultrasonic for two hours to obtain a uniform Fc-GO-MWNT modification solution, and drop 4 μL of the modification solution on the working electrode to obtain an Fc-GO-MWNT / AuNPs / SPE electrode.

[0055] (4) Weigh the molecularly imprinted polymer (MIP-MXene) of step (1), dissolve it in ultrapure water, and ultrasonic for 0.5 h to disperse it uniformly to obtain a MIP-MXene dispersion liquid with a concentration of 2 mg / mL. Then, use a pipette to draw 4 μL of the MIP-MXene dispersion liquid and drop it on the surface of the Fc-GO-MWNT / AuNPs / SPE working electrode, and naturally air dry to obtain a MIP-MXene / Fc-GO-MWNT / AuNPs / SPE electrode.

[0056] Example 1 also provides a palmitic acid molecularly imprinted sensor, which includes a MIP-MXene / Fc-GO-MWNT / AuNPs / SPE electrode as a working electrode, a carbon electrode as a counter electrode, and a silver / silver chloride electrode as a reference electrode.

[0057] Example 2 constructs a portable detection system for palmitic acid

[0058] Partly with reference to Figure 2 Example 2 provides a portable detection system for palmitic acid, which includes the palmitic acid molecularly imprinted sensor of Example 1, a portable DPV detection device, and a smartphone.

[0059] The portable DPV detection device includes a power module, a core processor, a DPV waveform generation circuit, a micro-current detection module, and a constant potential circuit. The core processor is an STM32F103ZET6. The micro-current detection module further includes an I / V conversion circuit, a low-pass filter circuit, an amplification circuit, and an analog-to-digital conversion circuit.

[0060] The palmitic acid molecularly imprinted sensor constructed in Example 1 is inserted into the interface of the portable DPV detection device, the sensor is provided with a scanning voltage by the DPV waveform generation circuit, the electrochemical experimental data of the sensor is collected by the micro-current detection module, and the core processor is wirelessly communicated to the smart phone application end.

[0061] The smart phone includes a wireless communication module, a data processing module and a data storage module. The data processing module can perform baseline calibration processing, and calculate the concentration value of the sample to be measured according to the standard working curve; the data storage module can store files, save configuration parameters and signal data.

[0062] The above portable palmitic acid detection system is used to detect the sample to be measured, and the specific process is as follows:

[0063] (1) Prepare palmitic acid solutions with concentrations of 10 -9 moL / L, 10 -7 moL / L, 10 -5 moL / L respectively, insert the prepared palmitic acid sensor into the portable DPV detection device, and drop different concentrations of palmitic acid solution into the detection pool for differential pulse voltammetry detection;

[0064] (2) Set the related detection parameters in the smart phone as follows: the voltage range is set to-0.2-0.6V, the interval time is 0.5s, the modulation time is 0.05s, the amplitude potential is 0.025V, and the detection process is carried out at room temperature (25℃). After receiving the detection instruction, the DPV detection device implements the detection and wirelessly transmits the detection signal to the smart phone, and the data processing module in the smart phone obtains the standard curve of the DPV peak current and the concentration of the palmitic acid solution and stores it, and the calculation formula of the standard curve is ΔI(μA)=1.19775LogC PA (nM)+3.03173(R 2 =0.996);

[0065] (3) The experimental material selects pig back muscle as the test object, weighs 250g of pork, stirs it into a paste, grinds it with liquid nitrogen, centrifugally dilutes it, and takes the supernatant to prepare the sample to be measured. A certain amount of sample solution is dropped into the detection pool, the DPV peak current of the sensor is collected by the portable DPV detection device, and is wirelessly transmitted to the data processing module of the smart phone; the data processing module compares and analyzes the detection data with the standard curve, obtains the concentration of palmitic acid in the sample solution, and immediately displays it on the display screen of the smart phone, and can be saved.

[0066] (4) Comparison of experimental results: The same piece of pig back muscle was selected as the experimental material, and its palmitic acid content was detected by gas chromatography. The results were compared with those of the electrochemical method of this invention, as shown in the table below. The results show that the electrochemical method is reliable.

[0067] Table 2 Comparison of Test Results

[0068]

[0069] Comparative Example 1

[0070] Compared to Example 1, the only difference is that no template molecules are added during the preparation of the polymer.

[0071] (1) Preparation of non-molecularly imprinted polymer: First, weigh 40 mg of dopamine and dissolve it in 20 ml of 95% anhydrous ethanol. Place the solution in a clean conical flask and ultrasonically clean it in ice water for 1 hour. Then, add 50 mg of Ti3C2T x MXene was dissolved in 30 ml of 95% anhydrous ethanol and sonicated at room temperature for 2 hours to fully disperse it, obtaining an MXene suspension. The MXene suspension was then added to a dopamine solution, and Tris-hydrochloric acid buffer was added dropwise to adjust the pH of the entire solution to 8.5. A magnetic stirrer rotor was placed in an Erlenmeyer flask, which was then wrapped in aluminum foil for light protection and stirred thoroughly on a magnetic stirrer for 24 hours. After removing the aluminum foil, the solution was poured into centrifuge tubes and centrifuged at 25°C, 8000 rpm for 10 minutes. After centrifugation, the centrifuge tubes were gently removed, the supernatant was discarded, and the lower solid layer was collected. The solid was then washed alternately with ultrapure water and ethanol to remove unreacted dopamine, followed by elution with 5% acetic acid solution for 15 minutes. The final non-molecularly imprinted polymer (NIP-MXene) coated on MXene is prepared (the procedure is consistent with that in Example 1).

[0072] (2) Clean the electrode surface with ultrapure water and anhydrous ethanol, then put the screen-printed electrode into a 0.5 g / L chloroauric acid (HAuCl4·4H2O) solution, and use the chronoamperometry to electrodeposit gold nanoparticles on the working electrode surface. Set the voltage parameter to -1V and the time parameter to 800s to obtain the AuNPs / SPE electrode.

[0073] (3) Mix Fc, GO, MWNT and 5wt% Nafion, the concentration of Fc is 10mg / mL, the concentration ratio of GO and MWNT is 1:1 (1mg / mL GO, 1mg / mL MWCNT). Ultrasonic for two hours to get uniform Fc-GO-MWNT modification solution, drop 4μL modification solution on the working electrode to get Fc-GO-MWNT / AuNPs / SPE electrode.

[0074] (4) Weigh the non-molecularly imprinted polymer (NIP-MXene) of step (1) and dissolve it in ultrapure water, ultrasonic for 0.5h to make it disperse uniformly, get MIP-MXene dispersion liquid with a concentration of 2mg / mL. Then take 4μL of NIP-MXene dispersion liquid by pipette and drop it on the surface of Fc-GO-MWNT / AuNPs / SPE working electrode, and get NIP-MXene / Fc-GO-MWNT / AuNPs / SPE electrode after natural drying.

[0075] Test results: from Figure 3 It can be seen that NIP-MXene / Fc-GO-MWNT / AuNPs / SPE has no special obvious response to different concentrations of PA, which shows that NIP lacks specific binding sites and cannot effectively bind PA. It further illustrates the success of the preparation of MIP-MXene / Fc-GO-MWNT / AuNPs / SPE described in Example 1, and it can effectively detect PA.

[0076] The above examples only describe the preferred embodiments of the present application, and do not limit the scope of the present application. Without departing from the design spirit of the present application, various modifications and improvements to the technical solutions of the present application made by ordinary engineering and technical personnel in the art shall fall within the protection scope determined by the claims of the present application.

Claims

1. A method for preparing a working electrode for electrochemically detecting palmitic acid, characterized by, The method comprises the following steps: 1) palmitic acid, dopamine and Ti3C2T x MXene are mixed together to carry out a polymerization reaction to obtain a polymer; template molecules in the polymer are eluted to obtain a molecularly imprinted polymer; 2) depositing gold nanoparticles on the screen-printed electrode by electro-deposition to obtain an AuNPs / SPE electrode; dropping a mixed solution of an electrochemical signal probe, graphene oxide, multi-walled carbon nanotubes and Nafion solution on the AuNPs / SPE electrode, and drying to obtain an Fc-GO-MWNT / AuNPs / SPE electrode; dropping a molecularly imprinted polymer dispersion on the Fc-GO-MWNT / AuNPs / SPE electrode, and drying to obtain an MIP-MXene / Fc-GO-MWNT / AuNPs / SPE electrode.

2. The production method according to claim 1, characterized by, the palmitic acid, the dopamine, and the Ti3C2T x The mass ratio of the MXene is 1~4:1~4:2.

5.

3. The preparation method according to claim 2, characterized in that, the palmitic acid, the dopamine, and the Ti3C2T x The mass ratio of the MXene is 1:2:2.

5.

4. The method of claim 1, wherein, In step 1), the polymerization is carried out at a pH of 7.5-10.

5.

5. The preparation method according to claim 4, characterized in that, In step 1), the polymerization is carried out at a pH of 8.

5.

6. The method of claim 1, wherein, In step 1), the polymerization is carried out for 20-48 hours.

7. The preparation method according to claim 1, characterized in that, In step 1), the elution is carried out for 5-25 minutes.

8. The preparation method according to claim 7, characterized in that, In step 1), the elution is carried out for 15 minutes.

9. The method of any one of claims 1-8, wherein, In step 2), the electrochemical signal probe is at least one selected from ferrocene, thionine and methylene blue.

10. The preparation method according to claim 9, characterized in that, In step 2), the electrochemical signal probe is ferrocene.

11. The preparation method according to any one of claims 1-8, characterized in that, the voltage for the electro-deposition is -1 V, and the time is 400-1200 seconds; and / or, in the mixed solution of the electrochemical signal probe, graphene oxide, multi-walled carbon nanotubes and Nafion solution, the concentration of the electrochemical signal probe is 6-14 mg / mL; and / or, in the mixed solution of the electrochemical signal probe, graphene oxide, multi-walled carbon nanotubes and Nafion solution, the concentration ratio of graphene oxide to multi-walled carbon nanotubes is 1-3:3-1; and / or, the concentration of the molecularly imprinted polymer dispersion is 1-5 mg / mL.

12. A working electrode for electrochemically detecting palmitic acid, characterized by, prepared according to the preparation method of any one of claims 1-11.

13. An electrochemical sensor for electrochemically detecting palmitic acid, characterized by, The working electrode according to claim 12, a counter electrode, and a silver / silver chloride electrode as a reference electrode.

14. A detection system for electrochemically detecting palmitic acid, characterized by The method comprises: The electrochemical sensor, the DPV detection device and the smart device according to claim 13; The electrochemical sensor is connected to the DPV detection device; The smart device is used to receive electrochemical experimental data of the DPV detection device and calculate the concentration of palmitic acid; The DPV detection device comprises: a detection cell for placing a solution to be tested; a power supply module for providing power supply for the DPV detection device; a DPV waveform generation circuit for providing a scanning voltage for the electrochemical sensor; a micro-current detection module for collecting electrochemical experimental data of the electrochemical sensor; a constant potential circuit for applying a voltage transmitted from the DPV waveform generation circuit between the working electrode and the reference electrode of the electrochemical sensor to ensure that the voltage meets the requirements; a core processor for transmitting the electrochemical experimental data to the smart device.

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