A fluorescent probe based on triphenylamine and benzothiazole derivatives and its preparation method and application

Abstract

The present invention relates to the technical field of organic small molecule fluorescent probes, and provides a fluorescent probe based on triphenylamine and benzothiazole derivatives, and its preparation method and application. The present invention uses triphenylamine as a fluorescent platform to construct a fluorescent probe having a structure shown in Formula ZR-I. Triphenylamine has good planarity, so that it exhibits high-intensity fluorescence characteristics, can respond to sulfur dioxide derivatives in a short time, and achieve the purpose of rapid detection; moreover, the fluorescent probe constructed by the present invention has high sensitivity to sulfur dioxide derivatives, with a detection limit reaching the micromolar level, and also has the advantages of good selectivity, strong anti-interference ability, and fast response, and can quickly and efficiently identify sulfur dioxide derivatives by colorimetry and fluorescence. In addition, the fluorescent probe preparation method is simple and easy to operate.

Description

Technical Field

[0001] The present invention relates to the technical field of organic small molecule fluorescent probes, and in particular to a fluorescent probe based on triphenylamine and benzothiazole derivatives, and a preparation method and application thereof. Background Art

[0002] With the progress of the times, human beings’ demand for food is no longer just to satisfy their hunger, but they have higher requirements for the color, flavor and other properties of food. With the rapid development of the national economy, the application of food additives in the food industry is becoming more and more extensive. SO2 is an important additive to improve the flavor and color of food. It is now widely used to add a more layered color, and it is also used to extend the shelf life of food. In recent years, with the continuous emergence of new foods, the problem of sulfur dioxide (SO2) residues in related foods has also emerged in an endless stream. Many unscrupulous businesses take advantage of consumers’ love for their products, frequently and excessively use additives, and even illegally add them in the processing and production of food. Excessive addition of SO2 without subsequent removal process will lead to its serious residue. SO2 or its derivatives (HSO3 - and SO3 2- ) Once excessive amounts of it are absorbed by the human body, it will cause asthma, allergies, and neurological diseases such as migraines and strokes, which will cause serious harm to the human body.

[0003] Currently, traditional methods for detecting SO2 in food include iodine titration and chromatography. However, these traditional methods all have drawbacks to varying degrees, such as cumbersome sample pretreatment, the need for large-scale instrumentation, complex processes, and lengthy testing times. In recent years, fluorescent probe-based analytical detection has garnered widespread attention from researchers. These methods offer numerous unique advantages, including excellent selectivity, high sensitivity, low cost, and ease of use, while also enabling non-destructive testing. Therefore, the application of fluorescent probes for SO2 detection in food holds considerable promise.

[0004] At present, fluorescent probes used to detect SO2 in food have achieved real-time dynamic monitoring of SO2 to a certain extent, but there are still some problems and there is still much room for improvement:

[0005] (1) Most probes for SO2 detection rely on a single signal change and have low sensitivity and are susceptible to interference from factors such as probe molecule concentration, instrument effects, and the environment;

[0006] (2) There are relatively few reported fluorescent probes for detecting SO2 in food, and most of them are likely to be interfered by biothiols in the system.

[0007] In summary, there is an urgent need to provide a fluorescent probe with high sensitivity and strong anti-interference ability to promote the application of fluorescent probe method in the detection of SO2 and its derivatives. Summary of the Invention

[0008] In light of this, the present invention provides a fluorescent probe based on triphenylamine and benzothiazole derivatives, as well as its preparation method and application. The fluorescent probe provided by the present invention can rapidly and highly selectively identify sulfur dioxide derivatives through colorimetry and fluorescence, with high sensitivity and strong anti-interference ability, and can be used for real-world sample detection and biological imaging research.

[0009] In order to achieve the above-mentioned object of the invention, the present invention provides the following technical solutions:

[0010] A fluorescent probe based on triphenylamine and benzothiazole derivatives, having a structure shown in formula ZR-I:

[0011]

[0012] The present invention also provides a method for preparing the fluorescent probe based on triphenylamine and benzothiazole derivatives described in the above scheme, comprising the following steps:

[0013] Mixing 4-triphenylamine borate, 4-bromo-2-hydroxybenzaldehyde, a basic compound, a palladium catalyst, and an organic solvent to perform a Suzuki coupling reaction to obtain a compound with a structure shown in Formula 1;

[0014]

[0015] 2-methylbenzothiazole, methyl iodide and an organic solvent are mixed to carry out a quaternization reaction to obtain a compound with a structure shown in Formula 2;

[0016]

[0017] The compound represented by Formula 1, the compound represented by Formula 2 and an organic solvent are mixed and subjected to a condensation reaction to obtain a fluorescent probe based on triphenylamine and benzothiazole derivatives and having a structure represented by Formula ZR-I.

[0018] Preferably, the molar ratio of 4-triphenylamine borate to 4-bromo-2-hydroxybenzaldehyde is 1:1.2 to 1:1.5;

[0019] The molar ratio of the 4-triphenylamine borate to the basic compound is 1:3 to 1:6; the basic compound is potassium carbonate;

[0020] The molar ratio of the 4-triphenylamine borate to the palladium catalyst is 1:0.05 to 1:0.1; and the palladium catalyst is PdCl2(dppf).

[0021] Preferably, the organic solvents used in the Suzuki coupling reaction are toluene and anhydrous ethanol; the volume ratio of toluene to anhydrous ethanol is 2:1; the Suzuki coupling reaction is carried out under heating reflux conditions, and the reaction time is 5 to 6 hours.

[0022] Preferably, after the Suzuki coupling reaction is completed, the method further comprises cooling the obtained reaction solution and concentrating it, dissolving the obtained concentrate and sequentially filtering it with diatomaceous earth and separating and purifying it with a silica gel column to obtain a compound with a structure represented by Formula 1; the elution mode of the silica gel column separation and purification is gradient elution, and the eluent used is a mixed solvent of ethyl acetate and petroleum ether. During the gradient elution, the volume ratio of ethyl acetate and petroleum ether in the mixed solvent changes from 1:200 to 1:20.

[0023] Preferably, the molar ratio of 2-methylbenzothiazole to methyl iodide is 1:1 to 1:1.2; the organic solvent used in the quaternization reaction is toluene; the quaternization reaction is carried out under heating reflux conditions, and the reaction time is 12 to 16 hours.

[0024] Preferably, after the quaternization reaction is completed, the process further comprises cooling the obtained reaction solution and filtering it, washing the obtained filter cake and drying it to obtain the compound with the structure shown in Formula 2.

[0025] Preferably, the molar ratio of the compound represented by Formula 1 to the compound represented by Formula 2 is 1:1 to 1:1.2; the organic solvents used in the condensation reaction are toluene and n-butanol; the volume ratio of toluene and n-butanol is 1:1; the condensation reaction is carried out under heating reflux conditions, and the reaction time is 12 to 16 hours.

[0026] Preferably, after the condensation reaction is completed, the obtained reaction solution is cooled and then concentrated and separated and purified by silica gel column to obtain a fluorescent probe based on triphenylamine and benzothiazole derivatives having a structure shown in Formula ZR-I; the elution process of the silica gel column separation and purification includes a first stage and a second stage, the eluent of the first stage is a mixed solvent of dichloromethane and petroleum ether, and the volume ratio of dichloromethane to petroleum ether is 1:1; the elution method of the second stage is gradient elution, and the eluent used is a mixed solvent of methanol and dichloromethane. During the gradient elution, the volume ratio of methanol and dichloromethane in the mixed solvent changes from 1:200 to 1:10.

[0027] The present invention also provides the use of the fluorescent probe based on triphenylamine and benzothiazole derivatives described in the above scheme in detecting sulfur dioxide derivatives.

[0028] The present invention provides a fluorescent probe based on triphenylamine and benzothiazole derivatives, having the structure shown in Formula ZR-I. The present invention uses triphenylamine as a fluorescent platform to construct a fluorescent probe having the structure shown in Formula ZR-I. Triphenylamine has good planarity, resulting in high-intensity fluorescence characteristics, which can respond to sulfur dioxide derivatives in a short time, achieving the purpose of rapid detection. Furthermore, the fluorescent probe constructed in the present invention has high sensitivity to sulfur dioxide derivatives, with a detection limit reaching the micromolar level. It also has the advantages of good selectivity, strong anti-interference ability, and fast response, and can quickly and efficiently identify sulfur dioxide derivatives through colorimetry and fluorescence.

[0029] The present invention also provides a method for preparing the fluorescent probe based on triphenylamine and benzothiazole derivatives described in the above scheme. The present invention adopts a three-step synthesis method to prepare the fluorescent probe based on triphenylamine and benzothiazole derivatives. The synthesis process is simple, convenient and easy to operate. BRIEF DESCRIPTION OF THE DRAWINGS

[0030] Figure 1 is the H NMR spectrum of compound 1 (solvent is CDCl3);

[0031] Figure 2 is the carbon NMR spectrum of compound 1 (solvent is CDCl3);

[0032] Figure 3 is the H NMR spectrum of compound 2 (the solvent is deuterated DMSO);

[0033] Figure 4 is the carbon NMR spectrum of compound 2 (the solvent is deuterated DMSO);

[0034] Figure 5 This is the H NMR spectrum of probe ZR-I (solvent: CDCl3);

[0035] Figure 6 This is the carbon NMR spectrum of probe ZR-I (solvent: CDCl3);

[0036] Figure 7 The fluorescence emission spectra of probe ZR-I for recognizing different concentrations of sulfur dioxide are as follows: the excitation wavelength is 338 nm, and the emission wavelength is 444 nm;

[0037] Figure 8 This is the fluorescence selectivity curve of probe ZR-I for recognizing sulfur dioxide, with an excitation wavelength of 338 nm and an emission wavelength of 444 nm;

[0038] Figure 9 This is the fluorescence interference resistance diagram of probe ZR-I in recognizing sulfur dioxide, with an excitation wavelength of 338 nm and an emission wavelength of 444 nm;

[0039] Figure 10 This is the minimum detection limit of probe ZR-I for identifying sulfur dioxide, with an excitation wavelength of 338 nm and an emission wavelength of 444 nm;

[0040] Figure 11 This is the fluorescence kinetics of probe ZR-I recognizing sulfur dioxide, with an excitation wavelength of 338 nm and an emission wavelength of 444 nm;

[0041] Figure 12 This is the pH applicable range of probe ZR-I for recognizing sulfur dioxide, with an excitation wavelength of 338 nm and an emission wavelength of 444 nm. DETAILED DESCRIPTION

[0042] The present invention provides a fluorescent probe based on triphenylamine and benzothiazole derivatives, having a structure shown in Formula ZR-I:

[0043]

[0044] The present invention also provides a method for preparing the fluorescent probe based on triphenylamine and benzothiazole derivatives described in the above scheme, comprising the following steps:

[0045] Mixing 4-triphenylamine borate, 4-bromo-2-hydroxybenzaldehyde, a basic compound, a palladium catalyst, and an organic solvent to perform a Suzuki coupling reaction to obtain a compound having a structure shown in Formula 1 (denoted as Compound 1);

[0046]

[0047] 2-methylbenzothiazole, methyl iodide and an organic solvent are mixed to carry out a quaternization reaction to obtain a compound with a structure shown in Formula 2 (denoted as Compound 2);

[0048]

[0049] The compound represented by Formula 1, the compound represented by Formula 2 and an organic solvent are mixed and subjected to a condensation reaction to obtain a fluorescent probe based on triphenylamine and benzothiazole derivatives and having a structure represented by Formula ZR-I.

[0050] The synthetic route provided by the present invention is as follows:

[0051]

[0052] The preparation method of the present invention is described in detail below.

[0053] The invention mixes 4-triphenylamine borate, 4-bromo-2-hydroxybenzaldehyde, a basic compound, a palladium catalyst and an organic solvent to carry out a Suzuki coupling reaction, thereby obtaining a compound with a structure shown in Formula 1. In the present invention, the molar ratio of the 4-boric acid triphenylamine and 4-bromo-2-hydroxybenzaldehyde is preferably 1:1.2; the molar ratio of the 4-boric acid triphenylamine to the basic compound is preferably 1:3 to 1:6, more preferably 1:5; the basic compound is preferably potassium carbonate; the molar ratio of the 4-boric acid triphenylamine to the palladium catalyst is preferably 1:0.05 to 1:0.1, more preferably 1:0.1; the palladium catalyst is preferably PdCl2(dppf); the organic solvent used in the Suzuki coupling reaction is preferably toluene and anhydrous ethanol; the volume ratio of toluene and anhydrous ethanol is preferably 2:1; the Suzuki coupling reaction is preferably carried out under heating reflux conditions, the heating reflux temperature is preferably 60°C, the Suzuki coupling reaction time is preferably 5 to 6 hours, and the Suzuki coupling reaction is preferably carried out under nitrogen protection. In a specific embodiment of the present invention, it is preferred to first dissolve 4-triphenylamine borate and 4-bromo-2-hydroxybenzaldehyde in a mixed solvent of toluene and anhydrous ethanol, and then add the alkaline compound and palladium catalyst, followed by heating to reflux for reaction.

[0054] After the Suzuki coupling reaction is completed, the present invention preferably cools the obtained reaction solution and then concentrates it, dissolves the obtained concentrate and then sequentially filters it through diatomaceous earth and separates and purifies it through a silica gel column to obtain a compound with a structure represented by Formula 1; the elution method used in the silica gel column separation and purification is gradient elution, and the eluent is a mixed solvent of ethyl acetate and petroleum ether, and the volume ratio of ethyl acetate and petroleum ether in the mixed solvent is preferably changed from 1:200 to 1:20. The present invention has no special requirements for the specific gradient of the gradient elution, as long as it can achieve sufficient elution of the product; the present invention removes the solvent in the reaction solution by concentration; the solvent used to dissolve the concentrate is preferably dichloromethane.

[0055] The present invention involves mixing 2-methylbenzothiazole, methyl iodide, and an organic solvent for a quaternization reaction to obtain a compound having the structure shown in Formula 2. In the present invention, the molar ratio of 2-methylbenzothiazole to methyl iodide is preferably 1:1 to 1:1.2, more preferably 1:1. The organic solvent used in the quaternization reaction is preferably toluene. The quaternization reaction is preferably carried out under heating and reflux conditions, the heating and reflux temperature is preferably 110°C, and the quaternization reaction time is preferably 12 to 16 hours. After the quaternization reaction is completed, the resulting reaction solution is preferably cooled and filtered, and the resulting filter cake is washed and dried to obtain the compound having the structure shown in Formula 2. The washing detergent is preferably glacial ethanol.

[0056] After obtaining the compound of formula 1 and the compound of formula 2, the present invention mixes the compound of formula 1, the compound of formula 2, and an organic solvent to perform a condensation reaction to obtain a fluorescent probe based on triphenylamine and benzothiazole derivatives having a structure represented by formula ZR-I. In the present invention, the molar ratio of the compound of formula 1 to the compound of formula 2 is preferably 1:1 to 1:1.2, more preferably 1:1.1; the organic solvent used in the condensation reaction is preferably toluene and n-butanol; the volume ratio of toluene and n-butanol is preferably 1:1; the condensation reaction is preferably carried out under heating reflux conditions, the heating reflux temperature is preferably 110°C, and the condensation reaction time is preferably 12 to 16 hours.

[0057] After the condensation reaction is completed, the present invention preferably cools the obtained reaction solution and then concentrates and separates and purifies it on a silica gel column to obtain a fluorescent probe based on triphenylamine and benzothiazole derivatives having a structure represented by formula ZR-I; the elution process of the silica gel column separation and purification preferably includes a first stage and a second stage, the eluent of the first stage is preferably a mixed solvent of dichloromethane and petroleum ether, and the volume ratio of dichloromethane and petroleum ether is preferably 1:1; the elution mode of the second stage is gradient elution, and the eluent used is preferably a mixed solvent of methanol and dichloromethane, and the volume ratio of methanol and dichloromethane in the mixed solvent during the gradient elution is preferably 1:200 to 1:10. The present invention has no special requirements for the specific gradient of the gradient elution, as long as it can achieve sufficient elution of the product.

[0058] The present invention also provides the use of the fluorescent probe based on triphenylamine and benzothiazole derivatives described in the above scheme in detecting sulfur dioxide derivatives; the sulfur dioxide derivative is specifically SO3 2 -; The fluorescence intensity of the fluorescent probe provided by the present invention increases with the increase of the concentration of sulfur dioxide derivatives; in a specific embodiment of the present invention, the fluorescent probe and the analyte are preferably added to the detection system for fluorescence detection, and the content of the sulfur dioxide derivative is determined according to the fluorescence intensity and the standard curve; the detection system is preferably PBS / DMSO, wherein the concentration of PBS is 10mM, and the volume ratio of PBS and DMSO is 8:2, hereinafter referred to as PBS / DMSO (10mM, v / v=8 / 2); the sulfur dioxide derivative is prepared with PBS buffer; the concentration of the fluorescent probe in the detection system is preferably 10μM; the pH value of the detection system is preferably 7-9; the standard curve is preferably a curve showing the relationship between the concentration of sulfur dioxide derivatives and the fluorescence intensity; in the present invention, the conditions for detecting sulfur dioxide derivatives using the fluorescent probe preferably include: an excitation wavelength of 338nm, a voltage of 400V, a detection temperature of 37°C, a slit of 10nm×10nm, and an emission wavelength of 444nm.

[0059] The following will be combined with the embodiments of the present invention to clearly and completely describe the technical solutions of the present invention. Obviously, the embodiments described are only some embodiments of the present invention, not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without making creative efforts are within the scope of protection of the present invention.

[0060] Example 1

[0061] In a 250 mL round-bottom flask, 4-triphenylamine borate (1.735 g, 6 mmol) and 4-bromo-2-hydroxybenzaldehyde (1.447 g, 7.2 mmol) were dissolved in a mixed solution of 60 mL of toluene and 30 mL of ethanol. K2CO3 (4.14 g, 30 mmol) and PdCl2(dppf) (0.439 g, 0.6 mmol) were added. Under a nitrogen atmosphere, the reaction was refluxed at 60°C for 5 h. After cooling to room temperature, the toluene and ethanol were removed by distillation under reduced pressure. The product was dissolved in dichloromethane and passed through diatomaceous earth to preliminarily remove impurities. The product was then separated and purified on a silica gel column using ethyl acetate:petroleum ether = 1:200 to 1:20 as eluent to obtain compound 1.

[0062] The NMR identification data of compound 1 are as follows: 1 H NMR(400MHz,Chloroform-d)δ11.10(s,1H),9.85(s,1H),7.54(d,J=8.1Hz,1H),7.47(d,J =8.7Hz,2H),7.29-7.23(m,4H),7.23-7.18(m,1H),7.17-7.09(m,6H),7.09-7.01(m,3H); 13 C NMR (101 MHz, Chloroform-d) δ 195.87, 162.16, 149.38, 148.95, 147.37, 134.20, 132.27, 129.57, 128.19, 125.16, 123.75, 122.87, 119.31, 118.32, 114.90. H NMR spectrum is shown in Figure 1 As shown, the carbon NMR spectrum is as follows Figure 2 shown.

[0063] Example 2

[0064] 2-Methylbenzothiazole (2 g, 13.4 mmol) and methyl iodide (1.9 g, 13.39 mmol) were dissolved in 20 mL of toluene and reacted under reflux at 110°C for 16 h. After the reaction was completed, the reaction solution was cooled and filtered, and the filter cake was rinsed with ice-cold EtOH and dried to obtain compound 2.

[0065] The NMR identification data of compound 2 are as follows: 1 H NMR (400MHz, DMSO-d6) δ8.46(dd,J=8.3,1.2Hz,1H),8.30(m,1H),7.90(m,1H),7.81(m,1H),4.21(s,3H),3.19(s,3H); 13 C NMR (101 MHz, DMSO-d6) δ 177.21, 141.56, 129.23, 128.68, 128.04, 124.50, 116.76, 36.28, 17.23. H NMR spectrum is shown in Figure 3 As shown, the carbon NMR spectrum is as follows Figure 4 shown.

[0066] Example 3

[0067] Compound 1 (100 mg, 0.27 mmol) and compound 2 (87.67 mg, 0.30 mmol) were dissolved in a mixed solvent of 10 mL of toluene and 10 mL of n-butanol, and the mixture was reacted at 110°C under reflux for 16 h. After the reaction was completed, the solvent was removed by distillation under reduced pressure, and the eluent was first dichloromethane: petroleum ether = 1:1, and then methanol: dichloromethane = 1:200-1:10. The mixture was separated and purified on a silica gel column to obtain the target product ZR-I.

[0068] The nuclear magnetic resonance identification data of ZR-I are as follows: 1 H NMR(400MHz,Chloroform-d)δ8.38(dd,J=8.3,1.2Hz,1H),8.32-8.21(m,2H),8.10(d,J=8.4Hz,1H),8.01(d ,J=15.8Hz,1H),7.87(m,1H),7.78(m,1H),7.68-7.61(m,2H),7.35(m,6H),7.16-7.02(m,9H),4.32(s,3H); 13 CNMR (101MHz, DMSO-d6)δ172.27,158.59,147.90,146.71,145.28,143.29,142.08,131.77,130.52,129.70,129.28,128.19,127.70,127.46,124.66,124.08,123.76,122.35,119.58,117.88,116.65,113.19,112.06,36.08. H NMR spectrum is shown in Figure 3. Figure 5 As shown, the carbon NMR spectrum is as follows Figure 6 shown.

[0069] Example 4

[0070] Fluorescence Spectroscopic Determination of Sulfur Dioxide Derivatives Using Fluorescent Probe ZR-I

[0071] (1) Preparation of fluorescent probe ZR-I solution: Prepare a stock solution of fluorescent probe ZR-I with dimethyl sulfoxide (DMSO);

[0072] (2) Preparation of Na2SO3 solution: Dissolve Na2SO3 powder in 10 mM PBS buffer to prepare a stock solution.

[0073] The detection system used was PBS / DMSO solution (10 mM, pH=8.1, v / v=8 / 2), and the concentration of the fluorescent probe in the control detection system was fixed at 10 μM.

[0074] In PBS / DMSO solution (10 mM, pH = 8.1, v / v = 8 / 2), 10 μM fluorescent probe ZR-I responded to different final concentrations of Na2SO3 solution (0-1000 μM), and its fluorescence emission spectrum was measured under the conditions of excitation wavelength of 338 nm, voltage of 400 V, temperature of 37°C, and slit size of 10 nm × 10 nm (the following examples all adopted this detection condition and will not be described in detail). The results are as follows: Figure 7 As shown. Figure 7 As can be seen, the fluorescence intensity at 444 nm gradually increases with increasing Na₂SO₃ concentration, indicating that the fluorescent probe ZR-I can be used as a fluorescent probe for the quantitative detection of sulfur dioxide. The fluorescent probe ZR-I of the present invention has good solubility and detection sensitivity and has important application value in fluorescent probes, biological detection, and fluorescence imaging.

[0075] Example 5

[0076] Selectivity experiment of fluorescent probe ZR-I

[0077] The fluorescent probe must be able to achieve single recognition of the detected species, so it is necessary to test the fluorescence selectivity of the fluorescent probe ZR-I to different active small molecules. The test solution for the fluorescence selectivity experiment of probe ZR-I is PBS / DMSO solution (10mM, pH=8.1, v / v=8 / 2), and the fixed probe concentration is 10μM. Figure 8As shown in the figure, under 338 nm excitation, the single probe showed almost no fluorescence emission at 444 nm. When Na2SO3 (40 eq.) was added, the fluorescence intensity at 444 nm was significantly enhanced. However, when other active species were added, including CH₃COONa, Al(NO₃)₃, NaBr, CaCO₃, NaCl, NaClO, K₂CO₃, CuSO₄·5H₂O, NaF, FeCl₃, KHCO₃, NaHS, KI, MgSO₄·7H₂O, NaNO₂, Na₂S₂SO₃·5H₂O, CaSO₄·2H₂O, and ZnCl (all added at 40 eq.), and GSH, Cys, and Hcy (all added at 100 eq.), only the solution with NaHS showed a slight increase in fluorescence intensity at 444 nm, but the increase was much smaller than that with Na₂SO₃. The fluorescence intensity at 444 nm of the solutions with the other active species remained comparable to that of the probe alone, showing no significant enhancement. These selectivity experimental results demonstrate that probe ZR-I exhibits good selectivity for sulfur dioxide under 338 nm excitation.

[0078] Example 6

[0079] Anti-interference experiment of fluorescent probe ZR-I

[0080] In order to investigate the anti-interference ability of probe ZR-I in response to sulfur dioxide in a complex environment, other active small molecules were tested using fluorescence emission spectroscopy. The test solution for the fluorescence interference experiment of probe ZR-I was PBS / DMSO solution (10 mM, pH = 8.1, v / v = 8 / 2). Figure 9 As shown, under 338nm excitation, the probe concentration is fixed at 10μM, and CH3COONa, Al(NO3)3, NaBr, CaCO3, NaCl, NaClO, K2CO3, CuSO4·5H2O, NaF, FeCl3, KHCO3, NaHS, KI, MgSO4·7H2O, NaNO2, Na2S2SO3·5H2O, CaSO4·2H2O, ZnCl (the addition amount of the above active species is 40eq.) and GSH, Cys, Hcy (the addition amount of GSH, Cys, Hcy is 100eq.) are added to the individual probe solutions, and then 40eq. of Na2SO3 is added to each of them, and the fluorescence emission intensity of the solution at 444nm is detected. Figure 9 It can be seen that except Cu 2+ 、Fe 3+ , HS - and Zn 2+In addition, the fluorescence intensity of the solution at 434 nm was basically the same as that of the solution with Na2SO3 added alone at 444 nm, which shows that probe ZR-I has a strong anti-interference ability against other active small molecules when detecting sulfur dioxide.

[0081] Example 7

[0082] Minimum detection limit experiment of fluorescent probe ZR-I

[0083] Fluorescent probes are required to have very high detection sensitivity, so the minimum detection limit is also an important indicator to measure the properties of the probe. Fluorescence emission spectroscopy is used to test the minimum detection limit of probe ZR-I for sulfur dioxide. The experimental test solution for the minimum detection limit of probe ZR-I is PBS / DMSO solution (10mM, pH=8.1, v / v=8 / 2). Under 338nm excitation, the concentration of probe ZR-I is fixed at 10μM, the concentration of Na2SO3 in the solution is adjusted (0μM to 90μM), and the fluorescence intensity at 444nm of the probe solution containing different concentrations of Na2SO3 is measured. The results are as follows: Figure 10 As shown. Figure 10 It can be seen that the fluorescence intensity at 444 nm of the solution has a good linear relationship with the Na2SO3 concentration in the range of 0 to 90 μM (R 2 =0.9954). According to IUPAC rules, the detection limit of probe ZR-I for sulfur dioxide was calculated using the detection limit formula (3σ / k) to be 0.66 μM. These detection limit experiments demonstrate that probe ZR-I has high sensitivity for sulfur dioxide and can quantitatively detect extremely low concentrations of sulfur dioxide.

[0084] Example 8

[0085] Fluorescence kinetics experiment of fluorescent probe ZR-I

[0086] Response time is also an important indicator for judging the quality of fluorescent probes. Fluorescence emission spectroscopy was used to test the fluorescence kinetics of probe ZR-I to sulfur dioxide. The experimental solution for probe ZR-I fluorescence kinetics was PBS / DMSO solution (10mM, pH=8.1, v / v=8 / 2). Under 338nm excitation, the fixed probe ZR-I concentration was 10μM, and the changes in 444nm fluorescence intensity of solutions without and with Na2SO3 (100μM) were measured over time. The results are as follows Figure 11 As shown, according to Figure 11 As can be seen, the fluorescence intensity at 444 nm of the Na2SO3 (100 μM) solution increases rapidly over time and reaches a plateau after 9 minutes. The fluorescence kinetics experimental results show that probe ZR-I responds rapidly to sulfur dioxide and can achieve rapid detection.

[0087] Example 9

[0088] pH stability experiment of fluorescent probe ZR-I

[0089] pH is also an important criterion for measuring the versatility of probes. Fluorescence spectroscopy was used to investigate the ability of probe ZR-I to recognize sulfur dioxide in different pH environments. The pH test solution for probe ZR-I was a PBS / DMSO solution (10mM, v / v=8 / 2) with a pH of 1 to 14. The concentrations of the probe and Na2SO3 were 10μM and 100μM, respectively. Figure 12 As shown, the fluorescence emission intensity of the probe alone at 444 nm remained unchanged within the pH range of 1 to 14. However, in the presence of Na₂SO₃, the fluorescence emission intensity at 444 nm of the test solution significantly increased within the pH range of 7 to 9. The pH test results demonstrate that probe ZR-I can recognize sulfur dioxide under physiological conditions.

[0090] Example 10

[0091] Testing food samples

[0092] The food samples used were rock sugar, dried mango, and rock sugar kumquat; the detection method was as follows:

[0093] The food samples were minced appropriately, and an appropriate amount was weighed and ultrasonically extracted with PBS buffer for 30 minutes. The sample was then filtered to prepare the test solution. The test system used was a PBS / DMSO solution (10 mM, pH = 8.1, v / v = 8 / 2). A control group and a test group were set up for each sample. The experiment was repeated three times for each sample. The fluorescence signal of the sample was recorded at 444 nm, and the accuracy of the method was evaluated by spike recovery.

[0094] The test results are shown in Table 1.

[0095] Table 1 Food sample testing

[0096]

[0097] According to the data in Table 1, it can be seen that the fluorescent probe of the present invention is used to detect sulfur dioxide derivatives in food, and the detection results are highly accurate.

[0098] The above is only a preferred embodiment of the present invention. It should be pointed out that for ordinary technicians in this technical field, several improvements and modifications can be made without departing from the principles of the present invention. These improvements and modifications should also be regarded as within the scope of protection of the present invention.

Claims

1. A fluorescent probe based on triphenylamine and benzothiazole derivatives, characterized in that: It has the structure shown in formula ZR-I:

2. The method for preparing a fluorescent probe based on triphenylamine and benzothiazole derivatives according to claim 1, characterized in that: The following steps are involved: Mixing 4-triphenylamine borate, 4-bromo-2-hydroxybenzaldehyde, a basic compound, a palladium catalyst, and an organic solvent to perform a Suzuki coupling reaction to obtain a compound with a structure shown in Formula 1; 2-methylbenzothiazole, methyl iodide and an organic solvent are mixed to carry out a quaternization reaction to obtain a compound with a structure shown in Formula 2; The compound represented by Formula 1, the compound represented by Formula 2 and an organic solvent are mixed and subjected to a condensation reaction to obtain a fluorescent probe based on triphenylamine and benzothiazole derivatives and having a structure represented by Formula ZR-I.

3. The preparation method according to claim 2, characterized in that The molar ratio of 4-triphenylamine borate to 4-bromo-2-hydroxybenzaldehyde is 1:1.2 to 1:1.5; The molar ratio of the 4-triphenylamine borate to the basic compound is 1:3 to 1:6; the basic compound is potassium carbonate; The molar ratio of the 4-triphenylamine borate to the palladium catalyst is 1:0.05 to 1:0.1; and the palladium catalyst is PdCl2(dppf).

4. The preparation method according to claim 2, characterized in that The organic solvents used in the Suzuki coupling reaction are toluene and anhydrous ethanol; the volume ratio of toluene to anhydrous ethanol is 2:1; the Suzuki coupling reaction is carried out under heating reflux conditions, and the reaction time is 5 to 6 hours.

5. The preparation method according to claim 2, characterized in that After the Suzuki coupling reaction is completed, the method further includes cooling the obtained reaction solution and concentrating it, dissolving the obtained concentrate and then filtering it with diatomaceous earth and separating and purifying it with a silica gel column to obtain a compound with a structure represented by Formula 1; the elution method of the silica gel column separation and purification is gradient elution, and the eluent used is a mixed solvent of ethyl acetate and petroleum ether. During the gradient elution, the volume ratio of ethyl acetate and petroleum ether in the mixed solvent changes from 1:200 to 1:

20.

6. The preparation method according to claim 2, characterized in that The molar ratio of 2-methylbenzothiazole to methyl iodide is 1:1 to 1:1.2; the organic solvent used in the quaternization reaction is toluene; the quaternization reaction is carried out under heating reflux conditions, and the reaction time is 12 to 16 hours.

7. The preparation method according to claim 2, characterized in that After the quaternization reaction is completed, the obtained reaction solution is cooled and then filtered, and the obtained filter cake is washed and then dried to obtain a compound with a structure shown in Formula 2.

8. The preparation method according to claim 2, characterized in that The molar ratio of the compound represented by Formula 1 to the compound represented by Formula 2 is 1:1 to 1:1.2; the organic solvents used in the condensation reaction are toluene and n-butanol; the volume ratio of toluene and n-butanol is 1:1; the condensation reaction is carried out under heating reflux conditions, and the reaction time is 12 to 16 hours.

9. The preparation method according to claim 2, characterized in that After the condensation reaction is completed, the method further includes cooling the obtained reaction liquid and then concentrating and separating and purifying it on a silica gel column to obtain a fluorescent probe based on triphenylamine and benzothiazole derivatives having a structure represented by formula ZR-I; the elution process of the silica gel column separation and purification includes a first stage and a second stage, the eluent of the first stage is a mixed solvent of dichloromethane and petroleum ether, and the volume ratio of dichloromethane to petroleum ether is 1:1; the elution method of the second stage is gradient elution, and the eluent used is a mixed solvent of methanol and dichloromethane. During the gradient elution, the volume ratio of methanol and dichloromethane in the mixed solvent changes from 1:200 to 1:

10.

10. Use of the fluorescent probe based on triphenylamine and benzothiazole derivatives according to claim 1 in detecting sulfur dioxide derivatives.

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