An open type fluorescent probe material, a preparation method thereof and a method for detecting phenyl mercaptan

Abstract

The present application relates to material and detection technology, disclose a kind of fluorescent molecular material and opening type fluorescent probe, and it is applied to detect phenylthiol.The fluorescent probe is naphthalimide-triphenylamine as fluorophore, 2,4-dinitrobenzene as recognition group, is a kind of high sensitivity and selective water-soluble fluorescent probe, for the selective detection of phenylthiol.The probe has significant sensitivity and specificity to almost pure aqueous solution of phenylthiol, can realize the qualitative, quantitative detection or visual rapid detection of phenylthiol in actual sample, has good development use prospect, has wide application prospect in the sensing of environment and bioscience.

Description

TECHNICAL FIELD

[0001] The present application relates to the field of detection and materials, and specifically discloses a fluorescent probe and a method for detecting thiophenol. BACKGROUND

[0002] Thiols, including aliphatic thiols and thiophenol, are important molecules in biological systems and chemical industries. Aliphatic thiols with low molecular weight and biological activity, including cysteine (Cys), homocysteine (Hcy), glutathione (GSH), etc., play an important role in a wide range of biological functions. Thiophenol has been widely used as a chemical intermediate for pesticides, drugs and amber dyes, and is a highly toxic and polluting chemical. Long-term exposure to thiophenol can cause serious damage to the central nervous system and other nervous systems, including shortness of breath, muscle weakness, hind limb paralysis, coma, and even death. The presence of thiophenol in water and soil can also cause damage to natural habitats. Thiophenol is toxic to aquatic organisms and animals, with a median lethal concentration (LC50) in fish of 0.01-0.4 mM and a median lethal dose (LD50) in mice of 46.2 mg / kg. Therefore, a detection technique that can selectively distinguish between toxic thiophenol and biologically important aliphatic thiols is of great significance in the fields of chemistry, environment and biology.

[0003] Fluorescence detection using fluorescent probes is considered one of the most attractive methods due to its simplicity, high sensitivity, fast spatial analysis, less biological damage and wide potential for chemical, biological and environmental applications. Since the development of a probe by Wang's research group, which can emit thiophene by a nucleophilic aromatic substitution (SNAr) mechanism, releasing SO2 and fluorophore, resulting in the resurrection of emission, the development of thiophenol fluorescent probes has attracted much attention, mostly with 2,4-dinitrobenzenesulfonyl (DNS) as a quencher, combined with various fluorophores such as naphthalimide, nitrobenzofuran, coumarin, BODIPY, etc., to detect thiophenol through intramolecular charge transfer (ICT) or photoinduced electron transfer (PET) or through bond energy transfer (TBET) mechanism. However, most of the reported probes require a large amount of organic cosolvent (≥30%, v / v) and relatively long time, and cannot well distinguish between thiophenol and aliphatic thiols, so it is necessary to design a "brighter", more sensitive, shorter reaction time, and water-soluble thiophenol sensor from the perspective of practical application. SUMMARY

[0004] The present application aims to provide a fluorescent molecular material, which can be used to prepare an on-off fluorescent probe, and a preparation method and application thereof.

[0005] Another object of the present application is to provide an on-off fluorescent probe for detecting thiophenol, and a preparation method and application thereof.

[0006] The application also provides a method for detecting benzene thiol, to solve the problems of long detection time, low detection sensitivity, narrow detection range, poor water solubility and the like in the detection of benzene thiol in the environment and living cells.

[0007] The application is achieved by the following technical solutions.

[0008] A fluorescent molecular material is a compound C2 (6-(4-diphenylamino) phenyl-2-(4-hydroxyphenyl)-1H-benzoisoquinoline-1,3(2H)-dione) as shown below:

[0009]

[0010] The compound can be excited by ultraviolet, and has ultraviolet absorption intensity at 440 nm. The ultraviolet excitation wavelength is 460 nm, and the emission wavelength is 480 nm. Under the excitation condition at 480 nm, there is strong fluorescence emission intensity at 560 nm.

[0011] The preparation method of the fluorescent molecular material comprises the following steps: refluxing compound C1 (6-bromo-2-(4-hydroxyphenyl)-1H-benzoisoquinoline-1,3(2H)-dione) with 4-boronic acid triphenylamine, a catalyst and a base in an organic solvent.

[0012]

[0013] The molar ratio of C1 to 4-boronic acid triphenylamine, the catalyst and the base is 1:1.1-1.2:0.04-0.1:10-20, preferably 1:1.1-1.4:0.03-0.05:12-18. The reflux reaction condition is that the reaction is carried out at 70-90 DEG C for 6-12 hours, preferably at 75-85 DEG C for 6-7 hours. The base is sodium carbonate, potassium carbonate or triethylamine, preferably potassium carbonate or sodium carbonate. The catalyst is a palladium catalyst, preferably tetra-(triphenylphosphine) palladium.

[0014] The compound C1 is prepared by the following method: 4-bromo-1,8-naphthalene anhydride is dissolved in a solvent, p-aminophenol is added, and the temperature is raised to 100-150 DEG C, and reflux reaction is carried out to obtain an intermediate product C1. Preferably, the reflux reaction temperature is 110-130 DEG C. The solvent is ethylene glycol monobutyl ether or tetrahydrofuran, preferably ethylene glycol monobutyl ether.

[0015]

[0016] The fluorescent molecular material can be used to prepare an open type fluorescent probe.

[0017] A fluorescent probe material for detecting benzene thiol and its derivatives is a compound shown in formula II:

[0018]

[0019] The preparation method of the compound comprises the following steps: heating and refluxing compound C2 and 2,4-dinitrofluorobenzene under a catalyst and a protective atmosphere until the reaction is complete.

[0020]

[0021] Preferably, the catalyst is triethylamine; the reaction solvent is DMF, acetonitrile or chloroform.

[0022] The molar ratio of C2, 2,4-dinitrofluorobenzene and the catalyst is 1:1-1.5:2-4, and in a preferred embodiment of the present application, the molar ratio is 1:1.2:3. The reflux reaction condition is that the reaction is carried out at 70-100°C for 4-12 hours, preferably at 75-90°C for 6-8 hours.

[0023] The compound C1 is prepared by the following method: 4-bromo-1,8-naphthalene anhydride is dissolved in ethylene glycol monobutyl ether, p-aminophenol is added, the temperature is raised to 123°C, and the reaction is stirred under reflux for 5h to obtain an intermediate product C1.

[0024]

[0025] Under the excitation condition of 480nm, the fluorescent probe material has no obvious fluorescence brightness due to 2,4-dinitrophenyl ether as a quencher. After benzene thiol or its derivatives are added, there is a strong green fluorescence emission at 560nm. Meanwhile, the probe alone has two absorption peaks at 308nm and 446nm. However, the addition of benzene thiol compounds to the probe solution causes the absorption peak at 446nm to have a blue shift, and has ultraviolet absorption intensity at 440nm. When other substances are added, the ultraviolet absorption intensity and the fluorescence emission intensity do not change obviously, indicating that the probe has specificity, good fluorescence specific selectivity and anti-interference, and good ultraviolet transfer selectivity. Therefore, the fluorescent probe material can be used for detecting benzene thiol compounds, or for preparing a reagent for detecting benzene thiol compounds.

[0026] The benzene thiol compound is an aromatic thiol, such as benzene thiol and its derivatives.

[0027] Based on the fluorescent probe material for detecting benzene thiol, fluorescence detection can be used for visual detection. The fluorescent probe material can be used for qualitatively and quantitatively detecting benzene thiol compounds.

[0028] The fluorescent probe material can be used for detecting phenylthiol, or used for preparing reagent, test paper, and kit for detecting phenylthiol compound.

[0029] The method for detecting phenylthiol compound comprises the following steps: mixing the fluorescent probe material of formula I with the sample to be detected, and detecting the fluorescence emission intensity or ultraviolet absorption intensity.

[0030] The method for fluorescence detection: if there is obvious fluorescence emission intensity at 560 nm under the excitation of 480 nm, that is, bright green fluorescence is generated, it indicates that the sample contains phenylthiol compound. The fluorescent probe of the present application has no obvious fluorescence absorption, but when phenylthiol compound exists, the probe will have fluorescence opening phenomenon under the ultraviolet light of 365 nm, that is, from no fluorescence to bright yellow-green.

[0031] The method for ultraviolet detection: if the ultraviolet absorption intensity at 306 nm increases, and the absorption peak at 458 nm has left shift phenomenon, it indicates that the sample contains phenylthiol compound.

[0032] Alternatively, the fluorescent probe material is mixed with the sample to be detected, and the ultraviolet absorption intensity (300-500 nm) is detected, that is, the method for ultraviolet detection. If the ultraviolet absorption intensity at 306 nm increases, and the absorption peak at 458 nm has left shift phenomenon, it indicates that the sample contains phenylthiol.

[0033] When the fluorescence method is used for detection, the fluorescence intensity at 560 nm can be used for quantitative detection. The fluorescence emission intensity at 500-800 nm is measured on a fluorescence spectrometer with 480 nm as the excitation wavelength. The working curve of the concentration of phenylthiol is obtained by taking the concentration of phenylthiol as the horizontal coordinate and the fluorescence intensity ratio at 560 nm as the vertical coordinate, so that qualitative and quantitative detection can be performed.

[0034] The method for visually detecting phenylthiol compound comprises the following steps: mixing the fluorescent probe material of formula I with the sample to be detected, and observing the fluorescence and color change under ultraviolet light.

[0035] When the phenylthiol compound is detected, the pH value of the reaction system is 2-12, preferably 6-8. In one preferred embodiment of the present application, the pH value is 7.4. The buffer solution used is preferably HEPES buffer, Tris or PBS buffer. Preferably, the buffer solution contains 0.1-2 mM CTAB.

[0036] The working principle of the probe is that the naphthalimide-triphenylamine part is used as a fluorescent group, the 2,4-dinitrophenyl group is used as a quenching group through a photo-induced electron transfer (PET) effect, and the benzene sulfenyl compound is detected through a fluorescence enhancement (turn-on) probe.

[0037] Since the benzene sulfenyl and derivatives have strong nucleophilicity, the probe molecule is attacked as a nucleophilic attack reagent, a bond between the naphthalimide ring and the 2,4-dinitrophenyl group is broken, a fluorescent signal is released, the product C2 with fluorescence is generated, and a significant fluorescence enhancement phenomenon of the system occurs, so that the benzene sulfenyl can be effectively detected.

[0038] With the above probe and detection method, the rapid response of the benzene sulfenyl can be realized in a short time (15 min), the minimum detection limit of the benzene sulfenyl detected by the fluorescence method is 20 nM, and the probe has high sensitivity, strong anti-interference ability, rapid recognition, and accurate detection results.

[0039] The present application has the following advantages:

[0040] (1) The present application first provides a new type of fluorescent molecule, and on this basis, a high-sensitivity, high-selectivity and water-soluble probe is established for the detection of benzene sulfenyl based on nucleophilic reaction, fluorescence "on-off" effect of the probe and ultraviolet absorption characteristics, and a new type of fluorescent probe is provided, which is an enhanced probe.

[0041] Compared with the fluorescence quenching type probe, the fluorescence enhancement type probe has better specific selection, anti-interference and higher sensitivity, and shows good anti-interference to other related analytes including aliphatic mercapto.

[0042] (2) The present application can quantitatively detect the benzene sulfenyl by fluorescence method, and can realize the rapid response of the benzene sulfenyl in a short time (15 min); at the same time, the probe has a significant linear correlation between the benzene sulfenyl concentration at 0-20 muM and the fluorescence intensity at 460 nm, and the minimum detection limit of the benzene sulfenyl is as low as 20 nM. At the same time, the probe has a good response in water solution, and is more suitable for actual application, such as can be used to detect the benzene sulfenyl in actual water sample and living cells. And the rapid detection can be realized.

[0043] (3) The probe of the present application is easy to prepare, has high sensitivity, strong anti-interference ability, rapid recognition, accurate detection results, simple detection process, can realize the rapid detection of the benzene sulfenyl in actual samples, can realize the application in biological field based on the probe, has good development and use prospect, and has wide application prospect in the sensing aspect of environment and biological science. BRIEF DESCRIPTION OF DRAWINGS

[0044] Figure 1 UV-Vis absorption spectrum of the fluorescent probe 1 of the present application in response to thiophenol,

[0045] Figure 2 Fluorescence spectrum of the fluorescent probe 1 of the present application in response to thiophenol, excitation wavelength 460 nm, emission wavelength 480 nm;

[0046] Figure 3 Fluorescence spectrum of the fluorescent probe 1 of the present application in response to thiophenol, excitation wavelength 460 nm, emission wavelength 480 nm;

[0047] Figure 4 Bar chart of the selectivity of the fluorescent probe 1 of the present application to thiophenol, black column is the probe added with other interfering substances, and gray column is thiophenol and its derivatives.

[0048] Figure 5 Bar chart of the selectivity of the fluorescent probe 1 of the present application to thiophenol, black column is the probe added with other interfering substances, and gray column is thiophenol and its derivatives.

[0049] Figure 6 Response time chart of the fluorescent probe 1 of the present application to recognize thiophenol, excitation wavelength 460 nm; emission wavelength 480 nm;

[0050] Figure 7 pH chart of the fluorescent probe 1 of the present application to recognize thiophenol;

[0051] Figure 8 Concentration titration fluorescence chart of the fluorescent probe 1 of the present application to recognize thiophenol, excitation wavelength 460 nm, emission wavelength 480 nm;

[0052] Figure 9 Chart for drawing the working curve of the fluorescent probe 1 of the present application to recognize thiophenol, excitation wavelength 460 nm; emission wavelength 480 nm;

[0053] Figure 10 Mechanism chart of the fluorescent probe 1 of the present application to recognize thiophenol;

[0054] Figure 11 Mechanism verification high performance liquid chromatogram of the fluorescent probe 1 of the present application to recognize thiophenol. DETAILED DESCRIPTION

[0055] The technical solutions of the present application will be clearly and completely described below in combination with the embodiments of the present application. Obviously, the described embodiments are only some of the embodiments of the present application, rather than all the embodiments. Based on the embodiments in the present application, all other embodiments obtained by those of ordinary skill in the art without creative effort belong to the scope of protection of the present application.

[0056] The chemical reagents, solvents used in the process of preparing the fluorescent probe (hereinafter referred to as probe) in the present application are all purchased from the Exploration Reagent Company, and the metal ions are all purchased from the Aladdin Reagent Company. In the process of confirming and performance testing of the fluorescent probe, a DTX-400 nuclear magnetic resonance spectrometer of Bruker Company is used, deuterated chloro DMSO is used as a solvent, and TMS is used as an internal standard to record nuclear magnetic resonance hydrogen spectrum and carbon spectrum; a Q-Exactive HR-MS mass spectrometer of Thermo Company is used to record high-resolution mass spectrum data. A British Edinburgh FS-5 fluorescence analyzer is used to record fluorescence spectrum.

[0057] Preparation of fluorescent probe 1 in Example 1

[0058]

[0059] 1. Preparation of C1

[0060] 4-bromo-1,8-naphthalic anhydride (2.00 g, 1 mmol) and p-aminophenol (945.26 mg, 1.2 mmol) were dissolved in ethylene glycol monobutyl ether (15 mL) at room temperature, and gradually heated to 123℃ under nitrogen protection to obtain a clear brown solution. Then, the solution became turbid, and refluxed for 5 hours. After the reaction was completed, the reaction solution was cooled to precipitate a solid, which was filtered, washed with water (20 ml), and dried to obtain a brown solid compound (2.49 g, yield 93.6%).

[0061] 2. Preparation of C2

[0062] Compound C1 (500 mg, 0.815 mmol), 4-boronic acid triphenylamine (432.02 mg, 0.8967 mmol) and tetrakis-(triphenylphosphine) palladium (60 mg, 0.0407 mmol) were sequentially dissolved in tetrahydrofuran (15 ml), and then K2CO3 solution (2.24 g / 5 mL water) was added by a syringe. Then, the reaction system was heated to 80℃, and refluxed for 8 h. After the reaction was completed, the reaction solution was cooled to room temperature, and the solvent was removed by rotary evaporation under reduced pressure. The crude product was purified by column chromatography (DCM) to obtain orange-red solid C2 (510.04 mg, yield 57.47%).

[0063] Structure characterization of C2

[0064] Nuclear magnetic resonance determination: 1H NMR (400 MHz, DMSO) δ 9.66 (s, 1H), 8.50 (t, J = 7.1 Hz, 2H), 8.40 (d, J = 8.5 Hz, 1H), 7.86 (t, J = 7.9 Hz, 1H), 7.79 (d, J = 7.6 Hz, 1H), 7.68 - 7.53 (m, 1H), 7.47 (d, J = 8.3 Hz, 2H), 7.37 (t, J = 7.7 Hz, 4H), 7.14 (dd, J = 14.7, 7.0 Hz, 9H), 6.89 (d, J = 8.5 Hz, 2H).13C NMR (101 MHz, DMSO) δ 164.31, 164.08, 157.64, 148.19, 147.25, 146.16, 132.01, 131.91, 131.87, 131.45, 131.17, 130.89, 130.36, 130.20, 129.72, 129.27, 129.16, 128.93, 128.17, 127.72, 127.28, 125.22, 124.25, 123.43, 122.44, 121.77, 115.89.

[0065] 3. Preparation of probe 1

[0066] In a 50 mL two-necked flask, compound 2 (300 mg, 1.0 mmol), 2,4-dinitrofluorobenzene (157.24 mg, 1.5 mmol) and triethylamine (0.303 g, 3.0 mmol) were dissolved in DMF (15 mL). The reaction system was heated to reflux at 80 °C for 6 to 8 hours under the protection of nitrogen, the reaction solution was cooled to room temperature, then extracted with water and dichloromethane, anhydrous sodium sulfate was added, the solvent was removed by rotary evaporation under reduced pressure, and the crude product was purified by column chromatography (PE:EA = 3:1, v / v) to obtain a light yellow solid (96.8 mg, yield 66%).

[0067] Structural characterization of probe 1

[0068] Nuclear magnetic resonance determination: 1 H NMR (400 MHz, DMSO) δ 8.94 (s, 1H), 8.55 (t, J = 6.8 Hz, 3H), 8.45 (d, J = 8.5 Hz, 1H), 7.98 - 7.82 (m, 2H), 7.60 - 7.36 (m, 12H), 7.19-7.13 (m, 7H). 13C NMR (101 MHz, DMSO) δ 164.20, 163.97, 154.97, 154.00, 148.28, 147.28, 146.49, 142.27, 140.25, 134.04, 132.00, 131.90, 131.48, 131.28, 130.98, 130.22, 129.89, 129.10, 128.27, 127.84, 125.25, 124.29, 123.45, 122.49, 122.39, 121.79, 120.81, 120.34.

[0069] HRMS-ESI: calcd for C 42 H 26 N4O7: 698.1801; found: 698.1888.

[0070] The probe obtained above was dissolved in water and could be used for the detection of thiophenol.

[0071] Response of the fluorescent probe to thiophenol compounds in Example 2

[0072] Prepare mixture A of HEPES buffer (10 mM, pH = 7.4, containing 0.5 mM CTAB), and prepare a 1 mM solution of probe 1 in DMSO.

[0073] Add 2 mL of mixture A to a clean cuvette and add 20 μL of dimethyl sulfoxide solution of probe 1 to make the concentration 20 μM. Test on the ultraviolet-visible spectrophotometer and fluorescence spectrum. Two absorption peaks at 308 nm and 446 nm are observed for probe 1 alone. The solution shows very weak fluorescence emission due to 2,4-dinitrophenyl ether as a quencher. However, the addition of p-methoxy mercaptan, i.e. p-methoxy thiophenol (MTP) to the solution of probe 1 causes a blue shift of the absorption peak at 446 nm, and the fluorescence intensity increases significantly at λem= 560 nm with the increase of the concentration of p-methoxy mercaptan. As shown in Figure 1 and Figure 2 .

[0074] The above experimental results show that probe 1 has a good response to thiophenol compounds in mixture A.

[0075] Selectivity of the fluorescent probe in Example 3

[0076] Prepare mixture A of HEPES buffer (10 mM, pH = 7.4, containing 0.5 mM CTAB), and prepare a 1 mM solution of probe 1 in DMSO.

[0077] In a clean fluorescence cuvette, 2 mL of HEPES buffer (10 mM, pH = 7.4, containing 0.5 mM CTAB) was added, and a dimethyl sulfoxide solution of probe 1 was added to make the concentration 10 μM.

[0078] The selectivity of probe 1 to p-methoxythiophenol (MTP) in mixed solution A was investigated by a fluorescence spectrometer. As shown in Figure 1 , under the excitation condition at 480 nm, the probe 1 (20 μM) alone had no obvious fluorescence emission in mixed solution A. When MTP (20 μM) was added, there was a very strong fluorescence emission intensity at 560 nm, but when other substances (100 μM) were added, the fluorescence emission intensity of the solution system had no obvious change compared with that of the probe system alone, i.e. there was no very strong fluorescence emission at 560 nm, as shown in Figure 3 and Figure 4 .

[0079] The above experimental results show that probe 1 has good fluorescence selective selectivity to thiophenol and its derivatives in mixed solution A.

[0080] Example 4 Anti-interference of the fluorescence probe

[0081] Mixed solution A of HEPES buffer (10 mM, pH = 7.4, containing 0.5 mM CTAB) was prepared, and a probe 1 solution with a concentration of 1 mM was prepared with DMSO. In 16 clean fluorescence cuvettes, 2 mL of mixed solution A and 20 μL of dimethyl sulfoxide solution of probe 1 were added, respectively, 10 molar equivalents of other analytes (blank, SO3 2- , SO4 2- , CO3 2- , HCO3 - , HSO3 - , NO2 - , F - , Br - , ClO - , Mg 2+ , Na + , K + , L-cys, GSH, H2S) and 20 μM of p-methoxythiophenol (MTP) were added, and the fluorescence intensity corresponding to different analytes was detected on a fluorescence spectrometer, and the fluorescence emission columnar graph was drawn as shown in Figure 5 .

[0082] Experiments proved that the recognition of the probe to MTP in mixed solution A was not interfered by the above-mentioned other analytes, and the probe had good anti-interference.

[0083] Example 5 Response time of the fluorescence probe to thiophenol compounds

[0084] Mixed solution A was prepared by HEPES buffer (10 mM, pH = 7.4, containing 0.5 mM CTAB) and the solution of probe 1 was prepared by DMSO with the concentration of 1 mM.

[0085] The response time of probe 1 to benzene thiol was investigated by adding 2 mL of mixed solution A and 20 μL of dimethyl sulfoxide solution of probe 1 in a cuvette, and then adding MTP (20 μM) and using a fluorescence spectrometer. The time-dependent fluorescence of the interaction between probe 1 and benzene thiol was studied by monitoring the change of fluorescence intensity at 560 nm over time. As shown in Fig. 2, a strong fluorescence signal was initiated within 5 min and reached saturation within 20 min, while the fluorescence signal of the free probe 1 solution changed negligibly within the same time interval, indicating that the probe is a turn-on probe that reacts rapidly with benzene thiol compounds and has significant fluorescence enhancement. Figure 6

[0086] After adding MTP in mixed solution A and placing it under a 365 nm UV lamp, it can be seen that the mixed solution changed from very weak fluorescence to bright yellow-green after adding MTP, thus achieving rapid response to MTP and enabling visual detection.

[0087] The results show that the probe is a turn-on probe that reacts rapidly with benzene thiol compounds and has significant fluorescence enhancement.

[0088] Example 6 pH determination of probe recognizing benzene thiol

[0089] Mixed solution A was prepared by HEPES buffer (10 mM, pH = 7.4, containing 0.5 mM CTAB) and the solution of probe 1 was prepared by DMSO with the concentration of 1 mM.

[0090] The effect of probe on the response of benzene thiol in mixed solution A at different pH was investigated by adding 2 mL of mixed solution A with different pH (2-12) and 20 μL of dimethyl sulfoxide solution of probe 1 in a cuvette, and then adding benzene thiol (20 μM) and using a fluorescence spectrometer. The emission spectrum of the probe itself remained basically unchanged in a wide pH range of 2.0-12.0, indicating that the probe 1 is very stable, but a significant fluorescence enhancement was observed in the pH range of 6.0-8.0 after adding 14 μM MTP, as shown in Fig. 4. Figure 7

[0091] The results show that probe 1 can detect benzene thiol and its derivatives in environmental and biological research.

[0092] Example 6 Concentration titration of probe recognizing benzene thiol compounds

[0093] ​​Mix solution A was prepared with HEPES buffer (10 mM, pH = 7.4, containing 0.5 mM CTAB), and the probe 1 solution with a concentration of 1 mM was prepared with DMSO. 2 mL of solution A and 20 μL of probe 1 dimethyl sulfoxide solution were added to a clean fluorescence cuvette.

[0094] The fluorescence intensity of probe 1 at different concentrations of MTP was investigated by fluorescence spectroscopy using anhydrous ethanol to prepare a MTP solution with a concentration of 0.005 mol / L. The concentration of MTP was gradually increased to 0, 2 μM, 4 μM, 6 μM, 8 μM, 10 μM, 12 μM, and 14 μM. As the concentration of benzene thiol increased, the fluorescence intensity at 460 nm gradually increased, as shown in FIG. 2. Figure 8

[0095] The results show that the fluorescence intensity changes with the concentration of benzene thiol, and quantitative detection can be performed.

[0096] Example 7 Investigation of the detection limit

[0097] A good detection limit is one of the criteria for testing whether a probe molecule has application value.

[0098] Mix solution A was prepared with HEPES buffer (10 mM, pH = 7.4, containing 0.5 mM CTAB), and the probe 1 solution with a concentration of 1 mM was prepared with DMSO. 2 mL of solution A and 20 μL of probe 1 dimethyl sulfoxide solution were added to a clean fluorescence cuvette. The concentration of benzene thiol was gradually increased to 0, 2 μM, 4 μM, 6 μM, 8 μM, 10 μM, and 12 μM. At the same time, the fluorescence emission intensity was measured on a fluorescence spectrometer with an excitation wavelength of 480 nm. The working curve of the concentration of benzene thiol was obtained with the concentration of benzene thiol as the abscissa and the fluorescence intensity ratio at 560 nm as the ordinate. The linear regression equation was Y = 64433.6394x + 31088.4127, R 2 = 0.99441. The unit of C is μmol / L. The detection limit of the probe molecule for benzene thiol was calculated to be 20 nM, as shown in FIG. 3, and good linearity was obtained. Figure 9

[0099] Example 8 Investigation of the detection mechanism

[0100] ​​Due to the photo-induced electron transfer (PET) behavior from naphthalimide-triphenylamine to 2,4-dinitro-phenyl ether moiety, probe 1 has almost no fluorescence. When phenyl mercaptan was added, strong yellow-green fluorescence appeared. It indicated that probe 1 had a sensing reaction with phenyl mercaptan. Because of the nucleophilicity of phenyl sulfide, we considered that the 2,4-dinitro-chlorobenzene of the probe was attacked by the nucleophilic thiol through the aromatic nucleophilic substitution mechanism, and at the same time, an electronic rearrangement occurred, which was the same as the reported probe. In this process, the phenyl ether bond in probe 1 was cleaved, and the structure of fluorophore C2 and its fluorescence properties were restored Figure 10 ). The reaction process was further studied by high-performance liquid chromatography analysis. Probe 1 had a single chromatographic peak at 11.5 min Figure 11 curve d). After the addition of PhSH (1.4 equivalents), the peak value of probe 1 decreased significantly, and a new peak appeared at 8.6 min Figure 11 curve a), which corresponded to compound C2 Figure 11 curve c). The chromatographic results showed that the PhSH compound could induce the conversion of probe 1 to C2-OH.

Claims

1. An openable fluorescent probe material, characterized in that, Compounds with the following structural formulas: 。 2. The method for preparing the openable fluorescent probe material according to claim 1, characterized in that, Includes the following steps: The C2 compound reacts with 2,4-dinitrofluorobenzene under a catalyst and a protective atmosphere by heating and reflux; the catalyst is triethylamine; the reaction solvent is DMF, acetonitrile, or chloroform. The preparation method of the C2 compound is as follows: 6-bromo-2-(4-hydroxyphenyl)-1H-benzodesisoquinoline-1,3(2H)-dione is reacted with triphenylamine 4-boronic acid, a catalyst, and a base in an organic solvent under reflux; the base is sodium carbonate, potassium carbonate, or triethylamine, and the catalyst is a palladium catalyst.

3. The application of the open-type fluorescent probe material of claim 1 in detecting thiophenol compounds in water samples, or in preparing reagents, test strips, and kits for detecting thiophenol compounds in water samples.

4. A method for detecting thiophene compounds in water samples, characterized in that, The method includes the following steps: mixing the open-type fluorescent probe material of claim 1 with a water sample and detecting the fluorescence emission intensity or ultraviolet absorption intensity; the open-type fluorescent probe material is dissolved in DMSO, and the volume ratio of DMSO to water sample is 20 μL: 2 mL.

5. A method for visually detecting thiophene compounds in water samples, characterized in that, The open-type fluorescent probe material of claim 1 was mixed with a water sample, and its fluorescence and color changes were observed under ultraviolet light; the open-type fluorescent probe material was dissolved in DMSO, and the volume ratio of DMSO to water sample was 20 μL: 2 mL.

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