An aggregation-induced luminescence fluorescent probe for detecting cuprous ions Cu(I) and its preparation method and application

Abstract

The present invention discloses an aggregation-induced luminescence fluorescent probe for detecting cuprous ions Cu(I), as well as its preparation method and application. The structure of the aggregation-induced luminescence fluorescent probe is shown in Formula I below; wherein R is selected from one of H, an alkyl group with carbon atoms ranging from C1 to C4, and COOH. The fluorescent probe provided by the present invention uses a chromophore quinoline nitrile derivative as an AIE structural unit and a sulfide-rich bis(2-((2-(ethylsulfide)ethyl)-thio)ethyl)amine as a Cu(I) recognition unit. It has excellent aggregation-induced luminescence properties, exhibits excellent selectivity and high sensitivity response to Cu(I), and can in situ detect the process of Cu(II) being reduced to Cu(I), providing technical support for revealing the role of Cu ions in cell physiology and pathology in the future.

Description

Technical Field

[0001] The present invention relates to the field of fluorescent probe preparation, and more specifically to an aggregation-induced luminescence fluorescent probe for detecting cuprous ions Cu(I), and a preparation method and application thereof. Background Art

[0002] Copper ions, as an indispensable coenzyme factor for a variety of enzymes and proteins, play a vital role in biochemistry, mediating multiple activities such as redox reactions in the cellular respiratory chain, defense mechanisms against oxidative stress, and neuronal function. When copper ion homeostasis is imbalanced, it may be closely related to a variety of pathological conditions, including but not limited to Wilson's disease, neurodegenerative diseases (such as Parkinson's disease and Alzheimer's disease), and atherosclerosis. In the reducing environment of cells, copper ions usually exist in the form of monovalent cations Cu(I). Free Cu(I) is considered to be more biotoxic than divalent cations Cu(II) in vivo. Therefore, real-time monitoring of the dynamic changes of intracellular Cu(I) is crucial for revealing its role in cellular physiological and pathological processes.

[0003] In biological imaging and molecular probe technology, fluorescent probes have become ideal tools for studying the dynamic changes and subcellular distribution of Cu(I) in vivo due to their high sensitivity, high selectivity and real-time monitoring capabilities. However, traditional fluorescent probes often suffer from photobleaching and are prone to severe aggregation quenching (ACQ), resulting in poor photostability and "false positive" reactions (Yanting L, Xiaoyan C, Qi W, et al. Advanced Functional Materials, 2021, 32(6)). The new AIE (Aggregation-Induced Emission) type fluorescent probes, with their "aggregation-enhanced emission" characteristics, overcome the limitations of traditional organic fluorescent probes in concentration quenching, and AIE type fluorescent probes are more resistant to photobleaching, so they have superior photostability and higher signal reliability. In addition, AIE type fluorescent probes have excellent biocompatibility and low cytotoxicity, which makes them show great application potential and significant technical advantages in the field of biological imaging and detection. However, there are relatively few aggregation-induced emission fluorescent probes for detecting Cu(I). Therefore, constructing Cu(I) fluorescent probes with AIE properties is of great significance for the detection of Cu(I) in organisms and cells. Summary of the Invention

[0004] The first objective of the present invention is to provide an aggregation-induced emission fluorescent probe for detecting cuprous ions (Cu(I)). The fluorescent probe prepared in this invention exhibits aggregation-induced emission (AIE) properties, exhibits a good response and excellent selectivity for Cu(I), and can sensitively detect the reduction of Cu(II) to Cu(I) in situ, providing technical support for future research revealing the role of Cu ions in cellular physiological and pathological processes.

[0005] The second object of the present invention is to provide a method for preparing the aggregation-induced emission fluorescent probe as described above.

[0006] The third object of the present invention is to provide a use of the aggregation-induced emission fluorescent probe described above in the preparation of a product for detecting cuprous ions Cu(I).

[0007] The fourth object of the present invention is to provide a method for detecting cuprous ions Cu(I) based on the aggregation-induced emission fluorescent probe described above.

[0008] To achieve the first objective, the present invention discloses an aggregation-induced luminescence fluorescent probe for detecting cuprous ions Cu(I). The structure of the aggregation-induced luminescence fluorescent probe is shown in Formula I below:

[0009]

[0010] Wherein, R is selected from one of H, an alkyl group with carbon atoms ranging from C1 to C4, and COOH.

[0011] By screening a large number of Cu(I) recognition units and AIE structural units, the present invention constructs a fluorescent probe with AIE effect for detecting Cu(I), which uses sulfide-rich bis(2-((2-(ethylsulfide)ethyl)-thio)ethyl)amine (NS4) as the Cu(I) recognition unit and a chromophore quinolinecarbonitrile derivative as the AIE structural unit. In the fluorescent probe, Cu(I) can combine with the fluorescent probe through coordination with NS4 to change the fluorescent signal. Therefore, the fluorescent probe provided by the present invention has good response and excellent selectivity to Cu(I), and the fluorescent probe can sensitively detect the process of Cu(II) being reduced to Cu(I) in situ.

[0012] Furthermore, the R is selected from methyl (named QM-NS4) or COOH (named QMCOOH-NS4).

[0013] To achieve the second objective, the present invention discloses a method for preparing the aggregation-induced emission fluorescent probe as described above, comprising the following steps:

[0014]

[0015] (1) Sodium ethoxide, 4-[bis-(2-chloroethyl)amino]benzaldehyde, and 2-(ethylthio)ethanethiol were dissolved in anhydrous ethanol and refluxed under an inert atmosphere. After the reaction, deionized water was added to the system, and the mixture was extracted with dichloromethane. The organic phase was washed with deionized water and saturated brine in sequence, dried over anhydrous sodium sulfate, concentrated, and purified by column chromatography to obtain compound 1;

[0016] (2) Compound 1, compound 2 and a catalyst are added to a solvent, and the mixture is refluxed under a nitrogen atmosphere. After the reaction is completed, the mixture is concentrated and purified by column chromatography to obtain the product.

[0017] Furthermore, the molar ratio of sodium ethoxide, 2-(ethylthio)ethanethiol, and 4-[bis-(2-chloroethyl)amino]benzaldehyde in step (1) is 3:2:0.5-1.

[0018] Furthermore, the molar volume ratio of sodium ethoxide to ethanol in step (1) is 1 mmol:(1.5-2) mL.

[0019] Furthermore, the reflux reaction time in step (1) is 3-8 hours.

[0020] Furthermore, the eluent used in the silica gel column chromatography in step (1) is a mixture of petroleum ether and ethyl acetate, with a volume ratio of 100-10:1.

[0021] Furthermore, the catalyst in step (2) is piperidine and the solvent is ultra-dry acetonitrile.

[0022] Furthermore, the molar ratio of compound 1 to compound 2 in step (2) is 1:0.5-1.

[0023] Furthermore, the reflux reaction time in step (2) is 8-14 hours.

[0024] Furthermore, the eluent used in the silica gel column chromatography in step (2) is a mixture of petroleum ether and ethyl acetate, with a volume ratio of 100-1:1.

[0025] To achieve the third objective, the present invention discloses an application of the aggregation-induced emission fluorescent probe described above in the preparation of a product for detecting cuprous ions Cu(I).

[0026] To achieve the fourth objective, the present invention discloses a method for detecting cuprous ions Cu(I) based on the aggregation-induced emission fluorescent probe described above, comprising the following steps:

[0027] S1. preparing a probe stock solution using the aggregation-induced emission fluorescent probe;

[0028] S2. adding the test solution to the probe mother solution to obtain a test solution, and diluting the probe mother solution with a solvent to obtain a probe reference solution, so that the final concentration of the aggregation-induced emission fluorescent probe in the test solution and the probe reference solution is the same;

[0029] S3. Based on the comparison of the emission peak fluorescence intensities of the probe reference solution and the test solution, determine whether the test solution contains cuprous ions Cu(I).

[0030] Further, step S3 is specifically as follows:

[0031] The emission peak fluorescence intensity of the probe reference solution and the test solution at 615 nm is tested respectively. If the emission peak fluorescence intensity of the test solution decreases compared with the probe reference solution, the test solution contains cuprous ions Cu(I). If the emission peak fluorescence intensity of the test solution does not change compared with the probe reference solution, the test solution does not contain cuprous ions Cu(I).

[0032] Furthermore, during the detection process, the final concentration of the aggregation-induced emission fluorescent probe was guaranteed to be above 10 μM.

[0033] Based on the above application, the present invention also explored the fluorescence response test of Cu(II) with different concentration gradients after reduction, and determined that as the concentration of generated Cu(I) increased, the emission peak intensity at 615nm gradually decreased, with obvious fluorescence changes.

[0034] Based on the above applications, the present invention also explores the changes in the fluorescence response of aggregation-induced luminescence fluorescent probes to common metal ions and reactive oxygen species in cells. Metal ions and reactive oxygen species are added to the probe mother solution and their fluorescence response changes are tested. The metal ions include Na + , K + , Mg 2+ , Ca 2+ , Mn 2+ ,Co 2+ , Ni 2+ , Cu 2+ , Zn 2+ and Cu + , reactive oxygen species include H2O2, ONOO - and ClO - Due to the specific coordination of the NS4 structural unit with Cu(Ⅰ), the fluorescence intensity changes significantly only when Cu(Ⅰ) is added. The addition of other metal ions or reactive oxygen species has no obvious effect on the fluorescence of the aggregation-induced emission fluorescent probe.

[0035] Based on the above application, the present invention further explored the in situ detection of the reduction of Cu(II) to Cu(I) by aggregation-induced emission fluorescent probe. By real-time monitoring of the fluorescence signal of the system, a trend of gradually decreasing emission peak intensity at 615 nm of the system was shown.

[0036] The beneficial effects of the present invention are as follows:

[0037] The present invention discloses an aggregation-induced emission fluorescent probe for detecting cuprous ions (Cu(I). The fluorescent probe uses sulfide-rich bis(2-((2-(ethylsulfide)ethyl)-thio)ethyl)amine (NS4) as the Cu(I) recognition unit and a chromophore quinolinenitrile derivative as the AIE structural unit. The fluorescent probe has the following characteristics:

[0038] 1. The fluorescence intensity of the fluorescent probe provided by the present invention increases with the increase of water content, and it has a significant AIE effect.

[0039] 2. The fluorescent probe provided by the present invention has a gradually decreasing emission peak intensity at 615 nm as the concentration of Cu(I) increases, showing a significant fluorescence change, which can achieve high-sensitivity detection of Cu(I) with a detection limit (LOD) of 2.03×10 -7 M.

[0040] 3. Since the fluorescent probe of the present invention has a high basic fluorescence, it can resist photobleaching, so that the fluorescent probe can achieve long-term monitoring of Cu(I).

[0041] 4. The fluorescent probe provided by the present invention is not interfered by various metal ions and reactive oxygen species, wherein the metal ions include Na + , K + , Mg 2+ , Ca 2+ , Mn 2+ ,Co 2+ , Ni 2+ , Cu 2+ , Zn 2+ and Cu + , reactive oxygen species include H2O2, ONOO - and ClO - Only after the addition of Cu(Ⅰ), the fluorescence intensity changes significantly, so the highly selective detection of Cu(Ⅰ) can be achieved, and the process of Cu(Ⅱ) reduction to Cu(Ⅰ) can be detected in situ.

[0042] 5. The fluorescent probe provided by the present invention emits weak light in a dispersed state, so it is expected to avoid interference from background signals when used for cell Cu(I) imaging, have a high signal-to-noise ratio, and do not require additional washing steps to remove non-aggregated fluorescent probes during cell imaging. BRIEF DESCRIPTION OF THE DRAWINGS

[0043] The specific embodiments of the present invention will be further described in detail below with reference to the accompanying drawings.

[0044] Figure 1 Compound 1 of the present invention 1 HNMR spectrum.

[0045] Figure 2 The fluorescent probe QM-NS4 of the present invention 1 HNMR spectrum.

[0046] Figure 3 This is the AIE property test of the fluorescent probe QM-NS4 of the present invention; the inset shows the change in relative fluorescence intensity at the emission peak of 615nm as the water content changes.

[0047] Figure 4 This is a response test of the AIE type fluorescent probe QM-NS4 of the present invention to Cu(Ⅰ).

[0048] Figure 5 This is the response curve of the AIE type fluorescent probe QM-NS4 of the present invention to different concentrations of Cu(I).

[0049] Figure 6 This is the selectivity test result of the AIE type fluorescent probe QM-NS4 of the present invention.

[0050] Figure 7 The stability of the AIE-type fluorescent probe QM-NS4 of the present invention and the test results of in-situ detection of Cu(II) reduction to Cu(I); the inset shows the color change of the system before and after the response to Cu(I). DETAILED DESCRIPTION

[0051] In order to more clearly illustrate the present invention, the present invention is further described below in conjunction with preferred embodiments and accompanying drawings. Similar components in the accompanying drawings are represented by the same reference numerals. It should be understood by those skilled in the art that the following detailed description is illustrative rather than restrictive and should not be used to limit the scope of protection of the present invention.

[0052] Unless otherwise specified, the raw materials used in the present invention can be obtained commercially. Any range described in the present invention includes the end value and any numerical value between the end values ​​and any sub-range formed by the end value or any numerical value between the end values.

[0053] Example 1

[0054] Synthesis of compound 1

[0055]

[0056] Sodium ethoxide (0.408 g, 6 mmol) was dissolved in anhydrous ethanol (12 mL), and 2-(ethylthio)ethanethiol (0.5 mL, 4 mmol) and 4-[bis-(2-chloroethyl)amino]benzaldehyde (0.5 g, 2 mmol) were added thereto. The mixture was stirred at reflux at 80 ° C for 4 h under nitrogen atmosphere. After the reaction, deionized water (20 mL) was added to the system, and then extracted with dichloromethane (20 mL × 3). The organic phase was washed with deionized water (20 mL × 1) and saturated brine (20 mL × 1), dried with a small amount of anhydrous sodium sulfate, filtered, concentrated, and separated and purified by silica gel column chromatography. The eluent was petroleum ether and ethyl acetate (10 / 1, v / v) to obtain compound 1 as an orange oil. Among them, compound 1 1 HNMR spectrum Figure 1 shown.

[0057] Synthesis of AIE-type fluorescent probe QM-NS4

[0058] To compound 1 (150 mg, 0.359 mmol) and 2-(1-ethyl-2-methylquinolin-4(1H)-ylidene)malononitrile (85 mg, 0.359 mmol) were added catalyst piperidine (0.2 mL) and solvent ultra-dry acetonitrile (12 mL), and the mixture was refluxed at 90°C under nitrogen atmosphere for 10 h. After the reaction, the solvent was concentrated and the mixture was separated and purified by silica gel column chromatography with petroleum ether and ethyl acetate (1 / 1, v / v) as the eluent to obtain an orange-red solid compound, QM-NS4. 1 HNMR spectrum Figure 2 shown.

[0059] Preparation of probe stock solution

[0060] Preparation of 1 mM QM-NS4 probe stock solution: Accurately weigh 6.3 mg of QM-NS4 probe molecules on an analytical balance, pipette 10 mL of dimethyl sulfoxide (DMSO) to dissolve the sample, and prepare a 1 mM probe stock solution. Wrap it in tin foil and store it in a refrigerator at 4°C away from light until use. The probe stock solution in the following examples is prepared using the same method.

[0061] Example 2

[0062] Synthesis of compound 1

[0063] Sodium ethoxide (0.408 g, 6 mmol) was dissolved in anhydrous ethanol (12 mL), and 2-(ethylthio)ethanethiol (0.5 mL, 4 mmol) and 4-[bis-(2-chloroethyl)amino]benzaldehyde (0.5 g, 2 mmol) were added thereto. The mixture was refluxed and stirred at 80 ° C for 6 h under nitrogen atmosphere. After the reaction, deionized water (20 mL) was added to the system, and then extracted with dichloromethane (20 mL×3). The organic phase was washed with deionized water (20 mL×1) and saturated brine (20 mL×1), respectively, and dried over a small amount of anhydrous sodium sulfate, filtered, and concentrated. The product was separated and purified by silica gel column chromatography with petroleum ether and ethyl acetate (10 / 1, v / v) as the eluent to obtain compound 1 as an orange oil.

[0064] Synthesis of AIE-type fluorescent probe QM-NS4

[0065] To compound 1 (150 mg, 0.359 mmol) and 2-(1-ethyl-2-methylquinolin-4(1H)-ylidene)malononitrile (85 mg, 0.359 mmol) were added catalyst piperidine (0.2 mL) and solvent ultra-dry acetonitrile (12 mL). The mixture was refluxed at 90°C under a nitrogen atmosphere for 12 h. After the reaction, the solvent was concentrated and the product was separated and purified by silica gel column chromatography with petroleum ether and ethyl acetate (1 / 1, v / v) as the eluent to obtain an orange-red solid compound, QM-NS4.

[0066] Determination of probe AIE properties

[0067] Preparation of blank solution: In a 50mL conical flask, add 50mL, 45mL, 40mL, 35mL, 30mL, 25mL, 20mL, 15mL, 10mL, 5mL, and 0mL of anhydrous ethanol respectively, and then add ultrapure water to make the total volume 50mL to prepare a series of C2H5OH / H2O mixed solvents with a water volume fraction of f w They are 0 (pure C2H5OH), 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, and 100%, respectively, and are prepared for use.

[0068] The AIE property of the probe was tested by adding 1.98 mL of the blank solution prepared in advance to a 10 mm cuvette, and adding 20 μL of the 1 mM QM-NS4 probe stock solution to obtain a C2H5OH / H2O mixed solution of QM-NS4 with a water content ranging from 0% to 99%. The fluorescence spectra of the fluorescent probe in solutions with different water contents were measured using a fluorescence spectrometer. The changes in the fluorescence emission spectra are shown in Figure 2. Figure 3 The inset shows the relative fluorescence intensity change at the emission peak of 615 nm as the water content changes. Figure 3As can be seen from the figure and its illustration, the AIE-type fluorescent probe QM-NS4 has obvious aggregation fluorescence enhancement performance with the increase of water content.

[0069] In order to ensure the dispersion of the fluorescent probe in the solution, the volume fraction of water f was selected in subsequent experiments. w =70% was tested as the test solution.

[0070] Example 3

[0071] Synthesis of compound 1

[0072] Sodium ethoxide (0.408 g, 6 mmol) was dissolved in anhydrous ethanol (12 mL), and 2-(ethylthio)ethanethiol (0.5 mL, 4 mmol) and 4-[bis-(2-chloroethyl)amino]benzaldehyde (0.5 g, 2 mmol) were added thereto. The mixture was refluxed and stirred at 80°C under nitrogen atmosphere for 8 h. After the reaction, deionized water (20 mL) was added to the system, and then extracted with dichloromethane (20 mL×3). The organic phase was washed with deionized water (20 mL×1) and saturated brine (20 mL×1), respectively, and dried over a small amount of anhydrous sodium sulfate, filtered, and concentrated. The product was separated and purified by silica gel column chromatography with petroleum ether and ethyl acetate (10 / 1, v / v) as the eluent to obtain compound 3 as an orange oil.

[0073] Synthesis of AIE-type fluorescent probe QM-NS4

[0074] To compound 1 (150 mg, 0.359 mmol) and 2-(1-ethyl-2-methylquinolin-4(1H)-ylidene)malononitrile (85 mg, 0.359 mmol) were added catalyst piperidine (0.2 mL) and solvent ultra-dry acetonitrile (12 mL). The mixture was refluxed at 90°C under a nitrogen atmosphere for 14 h. After the reaction, the solvent was concentrated and the product was separated and purified by silica gel column chromatography with petroleum ether and ethyl acetate (1 / 1, v / v) as the eluent to obtain an orange-red solid compound, QM-NS4.

[0075] Determination of the fluorescence emission spectrum changes of the probe in response to Cu(Ⅰ)

[0076] Preparation of stock buffer: Accurately pipette 3.75 mL of 4-hydroxyethylpiperazineethanesulfonic acid (HEPES, 1 M), 45 mL of anhydrous ethanol, and 101.25 mL of ultrapure water into a 250 mL conical flask to prepare a stock buffer containing 25 mM HEPES, with a water volume fraction of f w =70%.

[0077] Prepare NaCl solution with a concentration of 100 mM using the stock buffer solution. + , K + , Mg 2+ , Ca2+ , AANA, H2O2 aqueous solution, Mn with a concentration of 1mM 2+ ,Co 2+ , Ni 2+ , Cu 2+ , Zn 2+ aqueous solution and 10 mM ClO - 、ONOO - of aqueous solution.

[0078] Probe response test to Cu(I): 20 μL QM-NS4 probe stock solution was added to a 10 mm cuvette, a certain amount of reserve buffer solution was added thereto, and 20 μL Cu(II) solution, 20 μL AANa solution, 20 μL Cu(II) and 20 μL sodium ascorbate (AANa) solution were added thereto to make the final volume 2 mL. The fluorescence spectrum of the probe in response to Cu(II), AANa and Cu(I) was tested using a fluorescence spectrometer. The changes in the fluorescence emission spectrum are shown in Figure 2. Figure 4 .

[0079] Probe response test to different concentrations of Cu(I): Add 20μL QM-NS4 probe stock solution to a 10mm cuvette, add a certain amount of reserve buffer solution and 20μL AANa solution, and add 0-20μL Cu(II) solution to make the final volume 2mL. Use fluorescence spectrometer to test the fluorescence spectrum of the probe in response to different concentrations of Cu(I) solution. The changes in fluorescence emission spectrum are shown in Figure 5 .

[0080] from Figure 4 and Figure 5 It can be seen that the fluorescent probe has a good response to Cu(Ⅰ), and with the increase of Cu(Ⅰ) concentration, the fluorescence intensity of the fluorescent probe at 615 nm gradually decreases, and its detection limit LOD is 2.03×10 -7 M.

[0081] Selectivity test of the probe for Cu(I): 20 μL of QM-NS4 probe stock solution was added to a 10 mm cuvette, and a certain amount of reserve buffer solution and 20 μL of the interfering substance to be tested were added thereto to make the final volume 2 mL. The interfering substance to be tested was selected from Na + , K + , Mg 2+ , Ca 2+ 、Mn 2+ ,Co 2+ , Ni 2+ , Cu 2+ , Zn 2+ ,AANa,H2O2,ClO - , ONOO -Or Cu(Ⅰ), the fluorescence spectrum of the fluorescent probe to each interfering substance was tested by fluorescence spectrometer, and the fluorescence intensity at the emission peak of 615nm was shown in Figure 2. Figure 6 .from Figure 6 It can be seen that only after Cu(Ⅰ) was added, the fluorescence intensity changed significantly, indicating that the fluorescent probe has very good selectivity for Cu(Ⅰ).

[0082] The stability of the probe and the process of in-situ detection of Cu(Ⅱ) reduction to Cu(Ⅰ): 20 μL QM-NS4 probe stock solution was added to a 10 mm cuvette, a certain amount of reserve buffer solution and 20 μL AANa solution were added thereto, and 0 and 20 μL Cu(Ⅱ) solution were added thereto respectively to make the final volume 2 mL. The fluorescence spectra of the probe and the response of the probe to Cu(Ⅰ) solution over time were tested using a fluorescence spectrometer. The changes in the fluorescence emission spectrum are shown in FIG. Figure 7 The illustration shows the color change of the system before and after the probe responds to Cu(Ⅰ). Figure 7 As can be seen from the illustration, the fluorescent probe has good stability and can in situ test the process of Cu(Ⅱ) reduction to Cu(Ⅰ). During this process, the fluorescence intensity of the fluorescent probe at 615nm gradually weakens, and the color of the system changes from orange to yellow.

[0083] Obviously, the above embodiments of the present invention are merely examples for clearly illustrating the present invention, and are not limitations on the implementation methods of the present invention. For ordinary technicians in the relevant field, other different forms of changes or modifications can be made based on the above description. It is impossible to list all the implementation methods here. All obvious changes or modifications derived from the technical solution of the present invention are still within the scope of protection of the present invention.

Claims

1. An aggregation-induced luminescence fluorescent probe for detecting cuprous ions Cu(I), characterized in that: The structure of the aggregation-induced emission fluorescent probe is shown in Formula I below: I； Wherein, R is selected from one of H and an alkyl group with carbon atoms ranging from C1 to C4.

2. The aggregation-induced emission fluorescent probe according to claim 1, characterized in that The R is selected from methyl.

3. The method for preparing the aggregation-induced emission fluorescent probe according to claim 1 or 2, wherein: The steps include: (1) Sodium ethoxide, 4-[bis-(2-chloroethyl)amino]benzaldehyde, and 2-(ethylthio)ethanethiol were dissolved in anhydrous ethanol and refluxed under an inert atmosphere. After the reaction, deionized water was added to the system, and the mixture was extracted with dichloromethane. The organic phase was washed with deionized water and saturated brine in sequence, dried over anhydrous sodium sulfate, concentrated, and purified by column chromatography to obtain compound 1. (2) Compound 1, Compound 2 and a catalyst are added to a solvent, and the mixture is refluxed under a nitrogen atmosphere. After the reaction is completed, the mixture is concentrated and purified by column chromatography to obtain the product; Wherein, the catalyst in step (2) is piperidine.

4. The preparation method according to claim 3, characterized in that The molar ratio of sodium ethoxide, 2-(ethylthio)ethanethiol, and 4-[bis-(2-chloroethyl)amino]benzaldehyde in step (1) is 3:2:0.5-1; The molar volume ratio of the sodium ethoxide to the ethanol is 1 mmol:(1.5-2) mL.

5. The preparation method according to claim 3, characterized in that The reflux reaction time in step (1) is 3-8 h; The reflux reaction time in step (2) is 8-14 h.

6. The preparation method according to claim 3, characterized in that The solvent in step (2) is ultra-dry acetonitrile.

7. The preparation method according to claim 3, characterized in that The molar ratio of compound 1 to compound 2 in step (2) is 1:0.5-1.

8. Use of the aggregation-induced emission fluorescent probe according to claim 1 or 2 in the preparation of a product for detecting cuprous ions Cu(I).

9. A method for detecting cuprous ions Cu(I) based on aggregation-induced emission fluorescent probe, characterized in that: The detection method is not intended for the diagnosis and treatment of diseases and includes the following steps: S1. preparing a probe mother solution using the aggregation-induced emission fluorescent probe according to claim 1 or 2; S2. adding the test solution to the probe mother solution to obtain a test solution, and diluting the probe mother solution with a solvent to obtain a probe reference solution, so that the final concentration of the aggregation-induced emission fluorescent probe in the test solution and the probe reference solution is the same; S3. Based on the comparison of the emission peak fluorescence intensities of the probe reference solution and the test solution, determine whether the test solution contains cuprous ions Cu(I).

10. The detection method according to claim 9, characterized in that: Step S3 is specifically as follows: The emission peak fluorescence intensity of the probe reference solution and the test solution at 615 nm is tested respectively. If the emission peak fluorescence intensity of the test solution decreases compared with that of the probe reference solution, the test solution contains cuprous ions Cu(I). If the emission peak fluorescence intensity of the test solution does not change compared with that of the probe reference solution, the test solution does not contain cuprous ions Cu(I).

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