Method for improving N-(4-aminobutyl)-N-ethyl isoluminol cathode electrogenerated chemiluminescence signal intensity and application

Abstract

The invention discloses a method for improving the intensity of an N-(4-aminobutyl)-N-ethyl isoluminol cathode electrogenerated chemiluminescence signal and application of the method. A novel green and efficient signal enhancement method is established, and the problem of biotoxicity of a traditional co-reactant is solved. According to the method, an innovative solution is provided for N-(4-aminobutyl)-N-ethyl isoluminol cathode electrogenerated chemiluminescence signal enhancement, and a new technical path is provided for application expansion of the field of trace analysis and detection.

Description

Technical Field

[0001] This invention belongs to the field of electrochemiluminescence technology, and relates to a method and application for improving the intensity of N-(4-aminobutyl)-N-ethylisoluminol cathode electrochemiluminescence signal. Background Technology

[0002] Electrochemiluminescence (ECL) is an analytical technique that excites luminescent materials through electrochemical reactions to generate light signals. It boasts advantages such as high sensitivity, low background signal, and strong controllability, and has broad application prospects in biosensing, environmental monitoring, and clinical diagnostics. Luminol, a classic ECL luminescent material, has attracted considerable attention in cathode ECL systems due to its excellent chemiluminescent properties and good water solubility. It was one of the earliest artificially synthesized luminescent materials and is often used as a luminescent label, but it often suffers from decreased luminescence efficiency. Subsequent development of luminol derivatives has successfully overcome this deficiency. Therefore, this invention selects a luminol derivative—N-(4-aminobutyl)-N-ethylisoluminol (95%)—as the ECL luminescent material. However, the cathode ECL system of this derivative still faces problems such as weak signal, low efficiency, and poor stability, limiting its application in trace high-sensitivity analysis. The main reason for this is that the efficiency of the luminol derivative cathode ECL reaction is limited by the catalytic conversion ability of the co-reactants (such as hydrogen peroxide and dissolved oxygen). Currently, commonly used co-reactant systems have drawbacks such as low catalytic activity and numerous side reactions, resulting in weak ECL signals at the luminol derivative cathode, which makes it difficult to meet the requirements for high-sensitivity detection.

[0003] While current research focuses on developing efficient co-reactants and catalysts to enhance electrochemiluminescence signals, the biotoxicity of traditional co-reactants severely limits their application in the field of life analysis. Summary of the Invention

[0004] Objective of the Invention: The technical problem to be solved by the present invention is to provide a method for improving the signal intensity of N-(4-aminobutyl)-N-ethyl isoluminol cathode ECL. This method achieves efficient generation of co-reactants under light-assisted conditions by precisely optimizing the selection and structural design of co-reactant catalytic materials, thereby significantly improving the luminescence intensity of N-(4-aminobutyl)-N-ethyl isoluminol cathode ECL and significantly reducing the concentration of the luminescent material used.

[0005] Another technical problem to be solved by the present invention is to provide a method for determining the concentration of N-(4-aminobutyl)-N-ethyl isoluminol.

[0006] Technical Solution: To solve the above-mentioned technical problems, the present invention provides a method for improving the intensity of N-(4-aminobutyl)-N-ethylisoluminol cathodic electrochemiluminescence signal, comprising the following steps: (1) PLL-black phosphorus nanosheets (PLL / BP-NPs) were prepared by modifying thin-layer black phosphorus nanosheets (BP-NPs) with L-polylysine (PLL). (2) A working electrode was prepared by modifying PLL-black phosphorus nanosheets (PLL / BP-NPs) and carbon black composite material (CB) onto the surface of a glassy carbon electrode, and a photoelectric conversion system containing a working electrode, a counter electrode and a reference electrode was constructed. (3) Add N-(4-aminobutyl)-N-ethyl isoluminol solution to the reaction tank, and use a laser to illuminate the working electrode. Under the action of an applied voltage, detect the intensity of the electrochemiluminescence signal of N-(4-aminobutyl)-N-ethyl isoluminol molecule.

[0007] In step (1), thin-layer black phosphorus nanosheets (BP-NPs) are prepared by ultrasonic-assisted liquid phase exfoliation. The preparation method of PLL-black phosphorus nanosheets is as follows: weigh L-polylysine and dissolve it in the deoxygenated thin-layer black phosphorus nanosheet solution, and stir overnight.

[0008] In step (2), the modification is carried out by drip coating and natural drying in a glove box, and the mass ratio of PLL-black phosphorus nanosheets and carbon black composite material is 1:1.

[0009] In step (3), the laser used has a wavelength of 638 nm and an illumination time of 3 to 20 minutes.

[0010] In step (3), the applied voltage refers to the cathode electrochemiluminescence signal test performed under the action of a potential window of 0 ~ -0.9 V.

[0011] The present invention also includes the application of the method in determining the concentration of N-(4-aminobutyl)-N-ethyl isoluminol.

[0012] The present invention also includes a method for determining the concentration of N-(4-aminobutyl)-N-ethyl isoluminol, comprising the following steps: (S1) PLL-black phosphorus nanosheets were prepared by modifying thin-layer black phosphorus nanosheets with L-polylysine; (S2) A working electrode was prepared by modifying the surface of a glassy carbon electrode with PLL-black phosphorus nanosheets and carbon black composite material, and a photoelectric conversion system containing a working electrode, a counter electrode and a reference electrode was constructed. (S3) Add N-(4-aminobutyl)-N-ethyl isoluminol solutions of different concentrations to the reaction vessel, and use a laser to illuminate the working electrode. Under the action of an applied voltage, detect the electrochemiluminescence signal intensity of N-(4-aminobutyl)-N-ethyl isoluminol molecules. Based on the established relationship between the concentration of N-(4-aminobutyl)-N-ethyl isoluminol solution and the signal intensity, establish a standard curve. (S4) The N-(4-aminobutyl)-N-ethyl isoluminol solution to be tested is added to the reaction tank, and the working electrode is illuminated by a laser. Under the action of an applied voltage, the electrochemiluminescence signal intensity of N-(4-aminobutyl)-N-ethyl isoluminol molecules in the test solution is detected, and the concentration of N-(4-aminobutyl)-N-ethyl isoluminol molecules is calculated according to the standard curve.

[0013] The preparation method of PLL-black phosphorus nanosheets in step (S1) is as follows: L-polylysine is added to thin-layer black phosphorus nanosheets, and in step (S2), the modification is carried out by drop coating and natural drying in a glove box. The mass ratio of PLL-black phosphorus nanosheets to carbon black composite material is 1:1.

[0014] In step (S3), the laser used has a wavelength of 638 nm and an illumination time of 3 to 20 minutes.

[0015] In steps (S3) and (S4), the applied voltage refers to the cathode electrochemiluminescence signal test performed under the action of a potential window of 0 ~ -0.9 V.

[0016] This invention significantly enhances the intensity of the cathodic electrochemiluminescence signal of luminol derivative (N-(4-aminobutyl)-N-ethylisoluminol) through two key approaches: an in-situ self-generation of co-reactants strategy: utilizing a 638 nm laser to excite thin-layer black phosphorus nanosheets to generate singlet oxygen (¹O2), serving as a highly efficient co-reactant for the cathodic electrochemiluminescence of N-(4-aminobutyl)-N-ethylisoluminol; and a catalytic substrate material optimization strategy: selecting nanomaterials with both catalytic function and co-reactant self-generation capability as the substrate. The technical principle of this invention lies in the electro-optical conversion effect generated under an applied voltage. By applying a cathode potential, a redox reaction is induced in N-(4-aminobutyl)-N-ethylisoluminol and its co-reactants, converting the electrochemical signal into an optical signal, which is then acquired by a photomultiplier tube (PMT). This invention uses an ultrasound-assisted liquid-phase exfoliation method to prepare thin-layer black phosphorus nanosheets and significantly improves the material stability through L-polylysine (PLL); it innovatively combines laser excitation with electrochemical regulation to achieve in-situ generation and efficient utilization of ¹O2; and it enhances the electrochemiluminescence signal by applying an external voltage.

[0017] Beneficial Effects: Compared with existing technologies, this invention has the following advantages: This invention establishes a green and efficient new method for signal enhancement, avoiding the biotoxicity problems of traditional co-reactants. This method not only provides an innovative solution for enhancing the cathodic electrochemiluminescence signal of N-(4-aminobutyl)-N-ethylisoluminol, but also provides a new technical path for expanding applications in the field of trace analysis and detection. Attached Figure Description

[0018] Figure 1 Characterization diagram of thin-layer black phosphorus nanosheets: Figure 1 a), b), c), d), and e) are, respectively, TEM, HR-TEM, SAED, AFM, and Raman spectra of thin-layer black phosphorus nanosheets.

[0019] Figure 2 The figures show the electrochemical activity characterization of different modified electrodes. Figure 2 a) Cyclic voltammetry plots of bare glassy carbon electrode (GCE, black), carbon black-modified glassy carbon electrode (CB / GCE, green), and glassy carbon electrode modified with a mixture of carbon black and PLL-black phosphorus nanosheets (CB-PLL-BP / GCE, red) in a 5 mM potassium ferricyanide and 0.1 M potassium chloride solution, with a scan rate of 10 mV s. -1 The scanning potential window is -0.1~0.55 V. Figure 2 b) shows the cyclic voltammetry of a bare glassy carbon electrode (GCE, a, green), a glassy carbon electrode modified with PLL-black phosphorus nanosheets (PLL-BP / GCE, b, blue), a glassy carbon electrode modified with carbon black (CB / GCE, c, black), and a glassy carbon electrode modified with a mixture of carbon black and PLL-black phosphorus nanosheets (CB-PLL-BP / GCE, d, red) in saturated oxygen (O2), with a scan rate of 10 mV s. -1 The scanning potential window is -0.9~0.1 V. Figure 2 c) Cyclic voltammetry of a glassy carbon electrode (CB-PLL-BP / GCE) modified with a mixture of carbon black and PLL-black phosphorus nanosheets in saturated nitrogen (N2, a, black), saturated air (b, green), and saturated oxygen (O2), at a scan rate of 10 mV s. -1 The scanning potential window is -0.9~0.1 V.

[0020] Figure 3 The linear range of N-(4-aminobutyl)-N-ethyl isoluminol luminescent material was measured in saturated air using a glassy carbon electrode (CB-PLL-BP / GCE) modified with a mixture of carbon black and PLL-black phosphorus nanosheets. The concentrations of the N-(4-aminobutyl)-N-ethyl isoluminol luminescent material were 30 nM, 50 nM, 200 nM, 300 nM, and 500 nM.

[0021] Figure 4 To characterize the enhanced intensity of N-(4-aminobutyl)-N-ethylisoluminol cathodic electrochemiluminescence after laser irradiation, the cathodic electrochemiluminescence signal of PLL-thin black phosphorus nanosheets was irradiated with a 638 nm laser for different irradiation times (0 min, 1 min, 3 min, 5 min, 8 min, and 10 min).

[0022] Figure 5 The characterization results show that PLL-thin black phosphorus nanosheets can generate more singlet oxygen (¹O2) under light irradiation compared to bulk black phosphorus. The ¹O2 is generated by irradiation with a wavelength of 638 nm laser. Figure 5 a) and Figure 5 b) shows the UV-Vis absorption spectra of ¹O₂ scavenger 1,3-diphenylisobenzofuran (DPBF) after irradiation of bulk black phosphorus and thin-layer black phosphorus nanosheets with a 638 nm laser for a certain period of time. The irradiation times were 0 min, 5 min, 10 min, 15 min, and 20 min, respectively. Figure 5 c) UV-Vis absorption spectra of thin-layer black phosphorus nanosheets and DPBF in the presence of sodium azide (NaN3) as an O2 sacrificial agent. Detailed Implementation

[0023] The embodiments of the present invention will be described in detail below with reference to the examples. The following examples are only for illustrating the present invention. Unless otherwise specified, specific conditions in the examples are performed under conventional conditions or conditions recommended by the manufacturer. Reagents or instruments used, unless otherwise specified, are all commercially available conventional products.

[0024] The technical solution of the present invention will be further described and illustrated below with reference to specific embodiments and accompanying drawings. It should be noted that those skilled in the art can make various modifications and improvements without departing from the principles of the present invention, and these should also be considered within the scope of protection of the present invention.

[0025] The reagents and instruments used in this experiment are as follows: Reagents used in the experiment: Black phosphorus crystalline solids were purchased from Jiangsu Xianfeng Nanomaterials Technology Co., Ltd. (catalog number: 105572). N-methylpyrrolidone (NMP) was purchased from Aladdin Industrial Co., Ltd. (Shanghai, China). L-polylysine (PLL, catalog number: ST509) and N-(4-aminobutyl)-N-ethyl isoluminol (95%, catalog number: Y245805) were both purchased from Beyotime Biotechnology Co., Ltd. (Shanghai, China). Potassium ferricyanide, potassium ferrocyanide, sodium hydroxide, and hydrogen peroxide (H2O2, 35%) were all purchased from Sigma-Aldrich (Shanghai, China). Unless otherwise stated, all chemicals used in this invention are analytical grade and have not been further purified. Distilled water (≥18.2 MΩ cm) was obtained from an ultrapure water system (Direct-Q 3UV, Millipore, USA).

[0026] The instruments used in the experiment included: transmission electron microscopy (TEM), high-magnification transmission electron microscopy (HR-TEM), and selected area electron diffraction (SAED) analysis, all performed using a JEOL JEM-2100 transmission electron microscope (Japan). The thickness of the thin-layer black phosphorus nanosheets was characterized using a JEOL Dimenson ICON atomic force microscope (AFM). For electrochemical performance testing, the catalytic activity assessment of the thin-layer black phosphorus nanosheets and the determination of the applied voltage required to drive the N-(4-aminobutyl)-N-ethylisoluminol cathodic electrochemiluminescence signal were performed using a CHI660E electrochemical workstation manufactured by Shanghai Chenhua Instrument Co., Ltd., with cyclic voltammetry (CV) as the primary testing method. The glove box used for preparing the thin-layer black phosphorus nanosheets was a Delis RH model, which provided an inert gas protective environment. The ultrasonic tissue homogenizer used was a Lawson DH-250 model. Centrifugation was performed using a Thermo Fisher Scientific MULTIF low-temperature high-speed centrifuge, and ultrasonic dispersion was carried out using a Kunshan Ultrasonic Instrument Co., Ltd. KQ3200DE CNC ultrasonic cleaner. All instruments and equipment were used and maintained in accordance with standard operating procedures to ensure the accuracy and reliability of experimental data.

[0027] Example 1: Preparation of PLL-black phosphorus nanosheets and working electrode Thin-layer black phosphorus nanosheets were prepared by ultrasound-assisted liquid-phase exfoliation. The specific steps included: First, 20.00±0.1 mg of black phosphorus lumps were accurately weighed in a glove box under inert gas protection and ground into a gray state using an agate mortar (10.00±1 cm in diameter). Then, the mixture was transferred to a 50.00±1 mL screw-top double-walled glass bottle, and 20.00±0.1 mL of nitrogen-treated N-methylpyrrolidone (NMP) solvent was added. After sealing, the mixture was sonicated for 6 h using a cell disruptor at an ultrasonic power of 900 W, with an ultrasonic cycle of 2 s and a 4 s pause, under circulating cooling water (0℃). Finally, the desired thin-layer nanosheets were obtained by centrifugation using a gradient centrifugation method. Specifically: First, centrifuge at 6000 rpm for 20 min, and collect the supernatant in a glove box; then, centrifuge the supernatant again at 10000 rpm for 20 min, washing and centrifuging with acetone and distilled water respectively, retaining the precipitate, which is the prepared thin-layer black phosphorus nanosheets (<10 nm). Note: All weighing, grinding, and other operations involved in the entire process must be completed in an oxygen-free atmosphere in a glove box; the N-methylpyrrolidone, acetone, and distilled water used must all be purged with nitrogen to remove oxygen.

[0028] Preparation of PLL-black phosphorus nanosheets: Accurately weigh 2±0.1 mg of L-polylysine (PLL) and dissolve it in 1 mL of deoxygenated thin-layer black phosphorus nanosheet solution, and stir overnight for 12 h to obtain the final product.

[0029] Preparation of carbon black / PLL-protected thin-layer black phosphorus nanosheet electrode: PLL-thin-layer black phosphorus nanosheets and conductive carbon black (manufacturer: CABOT, USA, model: VXC-72, CAS: 1333-86-4) were mixed in 1 mL of N,N-dimethylformamide (DMF) solution at a mass ratio of 1:1. The mixture was stirred at room temperature in a glove box for 8 h. 5 μL of the mixture was accurately pipetted onto the clean glassy carbon electrode surface and allowed to air dry in a glove box to obtain the modified electrode.

[0030] Preparation of carbon black-modified electrodes: 1 ± 0.1 mg of conductive carbon black powder was accurately weighed and dissolved in 1 mL of DMF. The mixture was stirred at room temperature for 8 h. 5 μL of the carbon black composite solution was accurately transferred and applied to the clean surface of the glassy carbon electrode. The electrode was then air-dried in a glove box to obtain the modified electrode.

[0031] Example 2: Construction of an enhanced N-(4-aminobutyl)-N-ethylisoluminol cathodic electrochemiluminescence signaling system To enhance its ECL signal, this embodiment constructs an efficient and stable enhancement system, mainly including the following aspects: (1) Optimization of the electrochemical interface: The N-(4-aminobutyl)-N-ethylisoluminol cathodic electrochemiluminescence system was constructed using a three-electrode system. A bare glassy carbon electrode, an electrode with carbon black / PLL-protected thin-layer black phosphorus nanosheets prepared in Example 1, or an electrode modified only with carbon black were used as the working electrode. An Ag / AgCl electrode was used as the reference electrode, and a Pt sheet electrode was used as the counter electrode. A quartz trough with a diameter of approximately 15.00 ± 1 cm and a height of approximately 10 ± 1 cm was used as the reaction apparatus. Cyclic voltammetry scans were performed in a substrate containing 5 mM potassium ferricyanide and 0.1 M potassium chloride to characterize the electrochemical active area of ​​different electrodes. The scanning potential window was -0.1 to 0.55 V, and the scan rate was 10 mV / s. Figure 2 As shown in the green curve (a) in Figure 1), the carbon black-modified electrode exhibits a larger peak current in a 5 mM potassium ferricyanide and 0.1 M potassium chloride substrate compared to the bare glassy carbon electrode (black curve). Randles-Sevcik According to the formula, the active area of ​​the bare glassy carbon electrode is approximately 0.0701 cm². 2 The active area of ​​the carbon black-modified electrode is approximately 0.1533 cm². 2 This indicates that carbon black-modified electrodes are beneficial for increasing the electrochemical activity of the electrodes. For example... Figure 2 As shown by the red curve in a), the electrochemically active area of ​​the black phosphorus nanosheets modified with carbon black and protected by PLL is approximately 0.2311 cm². 2 The electrochemical active area was significantly increased. These results indicate that modifying carbon black and PLL-black phosphorus nanosheets is beneficial for promoting the electrochemical reaction.

[0032] (2) Self-generation of co-reactants: Cyclic voltammetry scans were performed on a bare glassy carbon electrode, an electrode with carbon black / PLL-protected thin-layer black phosphorus nanosheets prepared in Example 1, and a carbon black-modified glassy carbon electrode, respectively, in PBS (0.1 M, pH=7.0) containing 30 nM N-(4-aminobutyl)-N-ethyl isoluminol substrate with oxygen (O2) bubbled through it. The applied voltage was -0.9 to 0.1 V, and the scan rate was 10 mV / s. Figure 2 b) Cyclic voltammetry of different electrodes (bare glassy carbon electrode, electrode with carbon black / PLL protected thin-layer black phosphorus nanosheets prepared in Example 1, and carbon black modified glassy carbon electrode) in saturated O2, with a scanning potential window of -0.9 to 0.1 V and a scan rate of 10 mV s. -1The main purpose was to demonstrate that the carbon black and PLL-black phosphorus nanosheet composite material exhibits higher electrocatalytic activity for O2. Specifically, cyclic voltammetry diagrams of bare glassy carbon electrode (GCE, a, green), PLL-black phosphorus nanosheet-modified glassy carbon electrode (PLL-BP / GCE, b, blue), carbon black-modified glassy carbon electrode (CB / GCE, c, black), and a glassy carbon electrode modified with a mixture of carbon black and PLL-black phosphorus nanosheets (CB-PLL-BP / GCE, d, red) in saturated oxygen (O2) were presented. These results further demonstrate that the constructed sensor CB-PLL-BP / GCE exhibits better electrocatalytic activity for oxygen. Figure 2 c) shows the cyclic voltammetry of a glassy carbon electrode (CB-PLL-BP / GCE) modified with carbon black and PLL-black phosphorus nanosheets in saturated nitrogen (N2, a, black), saturated air (b, green), and saturated oxygen (O2). This further demonstrates that the introduction of carbon black and thin-layer black phosphorus nanosheets has a higher catalytic effect on oxygen reduction and lays the foundation for enhancing the cathodic electrochemiluminescence of N-(4-aminobutyl)-N-ethylisoluminol.

[0033] Example 3: Optimization of low-concentration electrochemiluminescent agent N-(4-aminobutyl)-N-ethylisoluminol Based on Example 2, an electrochemiluminescence signal response value of N-(4-aminobutyl)-N-ethyl isoluminol was tested in a saturated air atmosphere using a three-electrode system with a glassy carbon electrode (CB-PLL-BP / GCE) modified with carbon black and PLL-black phosphorus nanosheets as the working electrode, an Ag / AgCl electrode as the reference electrode, and a Pt sheet electrode as the auxiliary electrode. The concentrations of the N-(4-aminobutyl)-N-ethyl isoluminol luminescent material were 30 nM, 50 nM, 200 nM, and 300 nM, respectively. The test potential window was -0.9 V to 0.1 V, the scan rate was 10 mV / s, and PMT = 300. Linear curves were plotted based on the concentration of the N-(4-aminobutyl)-N-ethyl isoluminol luminescent material and its response signal value. Figure 3 The results showed that the glassy carbon electrode (CB-PLL-BP / GCE) modified with carbon black and PLL-black phosphorus nanosheets detected N-(4-aminobutyl)-N-ethyl isoluminol luminescent material. The ECL signal intensity gradually increased with increasing luminescent material concentration, and the lowest detectable concentration reached 30 nM. The linear equation was: Y = 10179.63 X +2222.76, R=0.9919, where Y refers to the ECL signal response value and X refers to the concentration of the luminescent material.

[0034] Example 4: Performance of PLL-black phosphorus nanosheets in generating singlet oxygen (¹O2) after light irradiation PLL-black phosphorus nanosheets exhibit excellent photosensitivity in photodynamic therapy and photocatalysis due to their unique layered structure and tunable band gap. As a novel two-dimensional material, PLL-black phosphorus nanosheets show significant light absorption under 638 nm near-infrared laser irradiation, enabling efficient electron transitions and the generation of reactive oxygen species, particularly singlet oxygen (¹O2) with strong oxidizing capabilities. This property makes them ideal for constructing self-powered electrochemiluminescence (ECL) systems, where the generated ¹O2 can directly serve as a co-reactant in the N-(4-aminobutyl)-N-ethylisoluminol cathode ECL reaction, achieving significant signal enhancement.

[0035] The surface of the glassy carbon electrode (CB-PLL-BP / GCE) modified with carbon black and PLL-black phosphorus nanosheet composite prepared in Example 1 was irradiated with a 638 nm laser for 0 min, 1 min, 3 min, 5 min, 8 min and 10 min. Then, ECL signals were acquired in a substrate containing 30 nM N-(4-aminobutyl)-N-ethylisoluminol under the conditions of potential window of -0.9 V to 0.1 V, scan rate of 10 mV / s and PMT=300. Figure 4 The results showed that under illumination, PLL-black phosphorus nanosheets could generate singlet oxygen (¹O₂) as a co-reactant for ECL luminescence, further enhancing the intensity of N-(4-aminobutyl)-N-ethylisoluminol cathodic electrochemiluminescence. Furthermore, with prolonged irradiation time, the intensity of N-(4-aminobutyl)-N-ethylisoluminol cathodic electrochemiluminescence gradually increased. Figure 4 As can be seen, under 638 nm laser irradiation, the intensity of the cathode ECL shows a significant increasing trend with the extension of illumination time. This phenomenon is attributed to the ¹O₂ generated by photoexcitation of thin-layer black phosphorus nanosheets, which provides sufficient co-reactant for the ECL reaction, thereby achieving cascade amplification of the signal. It is noteworthy that this photo-controlled ECL strategy not only improves detection sensitivity but also enables dynamic control of the ECL process.

[0036] To further investigate the ability of PLL-black phosphorus nanosheets to form, this invention used 1,3-diphenylisobenzofuran (DPBF, CAS: 5471-63-6) as an O2 scavenger for systematic verification experiments. DPBF molecules themselves exhibit fluorescence at 412 nm, but the fluorescence intensity decreases upon reaction with O2. A DPBF solution was prepared by accurately weighing 0.0027 g of DPBF solid powder and dissolving it in 1 mL of dimethyl sulfoxide (DMSO). 1 mL of catalyst solution (1 mg / mL of a bulk crystalline black phosphorus aqueous solution and 1 mg / mL of an aqueous solution of PLL-black phosphorus nanosheets prepared in Example 1) was added to 20 μL of a 2.7 mg / mL DPBF solution, and the absorbance was measured at 412 nm under light-protected conditions in the wavelength range of 300–600 nm. Figure 5 In the figure, 'a' represents the fluorescence signal value of DPBF after the bulk black phosphorus crystal solution generates ¹O2 under illumination. The results show that the fluorescence signal of DPBF at 412 nm decreases more slowly with increasing illumination time. Figure 5 In the figure, b represents the fluorescence signal value of DPBF after PLL-black phosphorus nanosheets generate ¹O2 under illumination. The results show that the fluorescence signal of DPBF at 412 nm decreases rapidly with increasing illumination time. This indicates that under illumination, PLL-black phosphorus nanosheets are more conducive to ¹O2 generation than bulk black phosphorus. This is attributed to the fact that ultrathin black phosphorus nanosheets (<10 nm) have a larger specific surface area and more active edge sites, which can improve the yield of ¹O2. However, in the presence of the ¹O2 sacrificial agent sodium azide (NaN3), (… Figure 5 c), the absorbance values ​​of PLL-black phosphorus nanosheets and DPBF UV-Vis absorption spectra did not change even under illumination, further indicating that the PLL-black phosphorus nanosheets can generate ¹O2 as an ECL luminescent co-reactant for the luminescent body N-(4-aminobutyl)-N-ethyl isoluminol, further enhancing the cathode ECL intensity.

Claims

1. A method for increasing the intensity of the cathodic electrochemiluminescence signal of N-(4-aminobutyl)-N-ethylisoluminol, characterized in that, Includes the following steps: (1) PLL-black phosphorus nanosheets were prepared by modifying thin-layer black phosphorus nanosheets with L-polylysine; (2) A working electrode was prepared by modifying the surface of a glassy carbon electrode with PLL-black phosphorus nanosheets and carbon black composite material, and a photoelectric conversion system containing a working electrode, a counter electrode and a reference electrode was constructed. (3) Add N-(4-aminobutyl)-N-ethyl isoluminol solution to the reaction tank, and use a laser to illuminate the working electrode. Under the action of an applied voltage, detect the intensity of the electrochemiluminescence signal of N-(4-aminobutyl)-N-ethyl isoluminol molecule.

2. The method for increasing the intensity of N-(4-aminobutyl)-N-ethylisoluminol cathode electrochemiluminescence signal according to claim 1, characterized in that, The preparation method of PLL-black phosphorus nanosheets in step (1) is as follows: weigh L-polylysine and dissolve it in the deoxygenated thin-layer black phosphorus nanosheet solution, and stir overnight.

3. The method for increasing the intensity of the N-(4-aminobutyl)-N-ethylisoluminol cathode electrochemiluminescence signal according to claim 1, characterized in that, In step (2), the modification is carried out by drip coating and natural drying in a glove box, and the mass ratio of PLL-black phosphorus nanosheets and carbon black composite material is 1:

1.

4. The method for increasing the intensity of the N-(4-aminobutyl)-N-ethylisoluminol cathode electrochemiluminescence signal according to claim 1, characterized in that, The laser used in step (3) has a wavelength of 638 nm and an illumination time of 3 to 20 minutes.

5. The method for increasing the intensity of the N-(4-aminobutyl)-N-ethylisoluminol cathode electrochemiluminescence signal according to claim 1, characterized in that, The applied voltage mentioned in step (3) refers to the cathode electrochemiluminescence signal test performed under the action of a potential window of 0 ~ -0.9 V.

6. The application of the method according to any one of claims 1 to 5 in the determination of N-(4-aminobutyl)-N-ethyl isoluminol concentration.

7. A method for determining the concentration of N-(4-aminobutyl)-N-ethyl isoluminol, characterized in that, Includes the following steps: (S1) PLL-black phosphorus nanosheets were prepared by modifying thin-layer black phosphorus nanosheets with L-polylysine; (S2) A working electrode was prepared by modifying the surface of a glassy carbon electrode with PLL-black phosphorus nanosheets and carbon black composite material, and a photoelectric conversion system containing a working electrode, a counter electrode and a reference electrode was constructed. (S3) Add N-(4-aminobutyl)-N-ethyl isoluminol solutions of different concentrations to the reaction vessel, and use a laser to illuminate the working electrode. Under the action of an applied voltage, detect the electrochemiluminescence signal intensity of N-(4-aminobutyl)-N-ethyl isoluminol molecules. Based on the established relationship between the concentration of N-(4-aminobutyl)-N-ethyl isoluminol solution and the signal intensity, establish a standard curve. (S4) The N-(4-aminobutyl)-N-ethyl isoluminol solution to be tested is added to the reaction tank, and the working electrode is illuminated by a laser. Under the action of an applied voltage, the electrochemiluminescence signal intensity of N-(4-aminobutyl)-N-ethyl isoluminol molecules in the test solution is detected, and the concentration of N-(4-aminobutyl)-N-ethyl isoluminol molecules is calculated according to the standard curve.

8. The method for determining the concentration of N-(4-aminobutyl)-N-ethyl isoluminol according to claim 7, characterized in that, The preparation method of PLL-black phosphorus nanosheets in step (S1) is as follows: L-polylysine is added to thin-layer black phosphorus nanosheets, and in step (S2), the modification is carried out by drop coating and natural drying in a glove box. The mass ratio of PLL-black phosphorus nanosheets to carbon black composite material is 1:

1.

9. The method for determining the concentration of N-(4-aminobutyl)-N-ethyl isoluminol according to claim 7, characterized in that, The laser used in step (S3) has a wavelength of 638 nm and an illumination time of 3 to 20 min.

10. The method for determining the concentration of N-(4-aminobutyl)-N-ethyl isoluminol according to claim 7, characterized in that, The applied voltage mentioned in steps (S3) and (S4) refers to the cathode electrochemiluminescence signal test performed under the action of a potential window of 0 ~ -0.9 V.

Images