An organic fluorescent dye compound with a DAD structure, its preparation method and application

Abstract

This invention belongs to the field of multi-stimulus responsive luminescent materials technology. It provides an organic fluorescent dye compound with a D-A-D structure, its preparation method, and its applications. The method includes the following steps: S1, in a protective gas atmosphere, a tricyclic compound, a thiophene compound, a palladium catalyst, an alkaline compound solution, and a solvent are mixed and subjected to a Suzuki coupling reaction to obtain an intermediate product; S2, in a protective gas atmosphere, the intermediate product, 2-amino-4,6-dimethylpyrimidine, tetrabutylammonium hydrogen sulfate, an alkaline compound solution, and a solvent are mixed and subjected to a Knauvengel condensation reaction to obtain the organic fluorescent dye compound with a D-A-D structure. The organic fluorescent dye compound provided by this invention allows for the regulation of its luminescent properties by changing the steric hindrance of the power supply system, showing promising application prospects in organic optoelectronic materials, biochemical detection, and cell imaging.

Description

Technical Field

[0001] This invention relates to the field of multi-stimulus responsive luminescent materials technology, and in particular to an organic fluorescent dye compound with a DAD structure, its preparation method, and its application. Background Technology

[0002] In recent years, stimulus-responsive materials (such as those responding to light, pH, ions, gases, and mechanical forces) have shown potential applications in sensors, optoelectronic devices, security systems, and displays, becoming a hot topic in science and technology. To date, various compounds responding to single stimuli from the external environment have been reported. However, compounds exhibiting high-contrast multicolor reactions to multiple external stimuli and specifically recognizing biothiols (Hcy) and reactive oxygen species (·OH) remain rare. Currently, the most studied probes for reactive sulfur compounds both domestically and internationally focus on the simultaneous detection of three reactive sulfur species; developing probes that specifically distinguish one reactive sulfur species still presents challenges. To construct multi-stimuli-responsive systems, we have developed numerous methods, including constructing donor-acceptor system models, combining stimulus structural motifs, or introducing weak intermolecular interactions, enabling rapid responses to external environmental stimuli. Among these, stimulus-responsive molecules with donor-acceptor structures possess intramolecular charge transfer (ICT) properties, with electron-donating (D) and electron-withdrawing (A) groups via π-conjugated linkers attracting increasing attention because they can be used as photoactive materials in biochemical fluorescence technology and photochemical sensing technology.

[0003] Pyrimidines possess good coplanarity and high electron affinity, making them suitable building blocks for constructing chromophores in chemical sensor materials. Combining freely rotatable triphenylamine and rigid planar anthracene as the power supply system to construct donor-acceptor-donor (DAD) configurations is highly attractive for developing novel stimulus-responsive materials. However, to date, the synthesis and properties of novel DAD-structured organic fluorescent dye compounds have not been reported in any literature or patents. Therefore, how to provide an organic fluorescent dye compound with a DAD structure has become a problem urgently needing to be solved by those skilled in the art. Summary of the Invention

[0004] In view of this, the present invention provides an organic fluorescent dye compound with a DAD structure, its preparation method, and its application. Its purpose is to solve the technical problem that existing technologies cannot prepare stimulus-responsive materials with a DAD structure.

[0005] To achieve the above objectives, the present invention adopts the following technical solution:

[0006] This invention provides an organic fluorescent dye compound having a DAD structure, which has the following characteristics:

[0007] The structure of Equation 1 or Equation 2:

[0008]

[0009] This invention provides a method for preparing the above-mentioned organic fluorescent dye compound having a DAD structure, characterized by comprising the following steps:

[0010] S1. Under a protective gas atmosphere, a tricyclic compound, a thiophene compound, a palladium catalyst, an alkaline compound solution, and a solvent are mixed and subjected to a Suzuki coupling reaction to obtain an intermediate product.

[0011] S2. Under a protective gas atmosphere, the intermediate product, 2-amino-4,6-dimethylpyrimidine, tetrabutylammonium hydrogen sulfate, alkaline compound solution and solvent are mixed and subjected to Knauvengel condensation reaction to obtain an organic fluorescent dye compound with a DAD structure.

[0012] Furthermore, the tricyclic compound is (4-(diphenylamine)phenyl)boronic acid or 9-bromoanthracene; the thiophene compound is 5-bromothiophene-2-carboxaldehyde or 2-boronic acid-thiophene.

[0013] Furthermore, the palladium catalyst is one or more of Pd(PPh3)4, PdCl2(dppf)2, Pd(dppf)Cl2, and Pd(OAc)2.

[0014] Furthermore, the concentration of the alkaline compound solution in steps S1 and S2 is independently 1–10 mol / L; the alkaline compound is independently one or more of Na2CO3, Ba(OH)2, K3PO4, Cs2CO3, K2CO3, TiOH, KF, CsF, TBAF, NaOH, and diethylisopropylamine; the protective gas in steps S1 and S2 is independently nitrogen, argon, or neon; and the solvent in steps S1 and S2 is independently one or more of THF, CH2Cl2, DMF, NaOH, and CH3CN.

[0015] Furthermore, in step S1, the molar ratio of the tricyclic compound and the thiophene compound is 1:1 to 2; the palladium catalyst accounts for 5 to 15 mol% of the total reactants; and the molar volume of the thiophene compound, the alkaline compound solution, and the solvent is 1 to 100 mmol: 60 to 120 mL: 80 to 250 mL.

[0016] Furthermore, in step S1, the temperature of the Suzuki coupling reaction is 80–90°C, and the time of the Suzuki coupling reaction is 11–13 h.

[0017] Further, in step S2, the molar ratio of the intermediate product to 2-amino-4,6-dimethylpyrimidine is 1:1 to 2; the molar volume ratio of the intermediate product, the alkaline compound solution, and the solvent is 1 to 3000 mmol: 30 to 120 mL: 60 to 150 mL; and the tetrabutylammonium hydrogen sulfate is 20 to 40 mol% of the total reactants.

[0018] Furthermore, in step S2, the temperature of the Knauvengay condensation reaction is 100–150°C, and the time of the Knauvengay condensation reaction is 8–12 h.

[0019] This invention also provides the application of the above-mentioned organic fluorescent dye compounds with DAD structures in organic optoelectronic materials, biochemical detection, or cell imaging.

[0020] As can be seen from the above technical solution, compared with the prior art, the beneficial effects of the present invention are as follows:

[0021] The structures of Formula 1 and Formula 2 provided by this invention can be adjusted by the polarity of acid, metal ions and solvent, and have high contrast multicolor display; the structure of Formula 2 can also specifically recognize reactive oxygen species ·OH, while Formula 1 can not only specifically recognize reactive oxygen species ·OH, but also specifically recognize biothiol Hcy.

[0022] The organic fluorescent dye compound with a DAD structure prepared in this invention exhibits high thermal stability, chemical stability, and strong fluorescence intensity. It can be used in acid-base and metal ion-responsive luminescent materials, security inks, anti-counterfeiting materials, and materials for detecting bioactive sulfur and oxygen. This compound can also enter HeLa cells and exhibit excellent cell staining function, making it a promising candidate as a cellular fluorescent marker in the field of cell dyes. The raw materials used to prepare this compound are relatively inexpensive, and the synthesis method is simple, facilitating industrial production. Attached Figure Description

[0023] Figure 1 The hydrogen spectrum of BTPAT-PA prepared in Example 1;

[0024] Figure 2 The hydrogen spectrum of BAT-PA prepared in Example 2;

[0025] Figure 3 In the diagram, (A) is the fluorescence emission spectrum of compound BTPAT-PA, (B) is the fluorescence emission spectrum of compound BAT-PA, (C) is the ultraviolet absorption spectrum of compound BTPAT-PA, and (D) is the ultraviolet absorption spectrum of compound BAT-PA.

[0026] Figure 4 (A) in the figure is the absorption spectrum variation curve of BTPAT-PA (inset: H)+ (A) Effect of concentration on absorption intensity at 530 nm, (B) is the emission intensity variation curve of BTPAT-PA (inset: H) + The effect of concentration on emission intensity at 574 nm, λ ex =390nm). (C) shows the absorption spectrum change of BAT-PA (inset: H) + (D) shows the effect of concentration on absorption intensity at 463 nm, and (H) shows the change in emission intensity of BAT-PA (inset: H). + (Effect of concentration on emission intensity at 545 nm);

[0027] Figure 5 (A) is the absorption spectrum curve of BTPAT-PA in solutions with different pH values, (B) is the fluorescence spectrum curve of BTPAT-PA in solutions with different pH values, (C) is the absorption spectrum curve of BAT-PA in solutions with different pH values, and (D) is the fluorescence spectrum curve of BAT-PA in solutions with different pH values.

[0028] Figure 6 Photographs showing the color changes of BTPAT-PA and BAT-PA on the test strips in solutions with different pH values;

[0029] Figure 7 In the image, (A) is the absorption spectrum of BTPAT-PA after adding equal amounts of various metal ions, and (B) is the fluorescence spectrum of BTPAT-PA after adding equal amounts of various metal ions.

[0030] Figure 8 For BTPAT-PA in THF for Al 3+ Cr 3+ and Fe 3+ The titration absorption spectrum change curve is shown, where (A) represents Al. 3+ / EDTA-induced absorption spectrum change curve, (B) is Cr 3+ / EDTA-induced absorption spectrum change curve, (C) is Fe 3+ / EDTA-induced absorption spectrum change curve;

[0031] Figure 9 For BTPAT-PA in THF for Al 3+ Cr 3+ and Fe 3+ The titration fluorescence spectrum change curve, where (A) represents Al. 3+ / EDTA-induced fluorescence spectrum change curve, (B) is Cr 3+ / EDTA-induced fluorescence spectrum change curve, (C) is Fe 3+ / EDTA-induced fluorescence spectrum change curve;

[0032] Figure 10 In the diagram, (A) is the absorption spectrum of BAT-PA after adding equal amounts of various metal ions, and (B) is the fluorescence spectrum of BAT-PA after adding equal amounts of various metal ions.

[0033] Figure 11 For BAT-PA in THF for Al 3+ Cr 3+ and Fe 3+ The titration absorption spectrum change curve is shown, where (A) represents Al. 3+ / EDTA-induced absorption spectrum change curve, (B) is Cr 3+ / EDTA-induced absorption spectrum change curve, (C) is Fe 3+ / EDTA-induced absorption spectrum change curve;

[0034] Figure 12 For BAT-PA in THF for Al 3+ Cr 3+ and Fe 3+ The titration fluorescence spectrum change curve, where (A) represents Al. 3+ / EDTA-induced fluorescence spectrum change curve, (B) is Cr 3+ / EDTA-induced fluorescence spectrum change curve, (C) is Fe 3+ / EDTA-induced fluorescence spectrum change curve;

[0035] Figure 13 In the image, (A) shows the change of BTPAT-PA with different metal ions, and (B) shows the change of BAT-PA with different metal ions.

[0036] Figure 14 In the image, (A) is the absorption spectrum of BTPAT-PA after adding equal amounts of various active oxides, and (B) is the fluorescence spectrum of BTPAT-PA after adding equal amounts of active oxides.

[0037] Figure 15 The graphs show the changes in absorption and fluorescence spectra of BTPAT-PA in THF after titration with ·OH. In the graphs, (A) shows the change in absorption spectrum induced by ·OH, and (B) shows the change in fluorescence spectrum induced by ·OH.

[0038] Figure 16 The graphs show the changes in absorption and fluorescence spectra of BTPAT-PA in THF during titration with Hcy. (A) shows the Hcy-induced absorption spectrum change, and (B) shows the Hcy-induced fluorescence spectrum change.

[0039] Figure 17 In the image, (A) is the absorption spectrum of BAT-PA after adding equal amounts of various active oxides, and (B) is the fluorescence spectrum of BAT-PA after adding equal amounts of active oxides.

[0040] Figure 18 The graphs show the changes in absorption and fluorescence spectra of BAT-PA in THF after titration with ·OH. In the graphs, (A) shows the changes in absorption spectra induced by ·OH, and (B) shows the changes in fluorescence spectra induced by ·OH.

[0041] Figure 19 In the image, (A) shows the change of BTPAT-PA with different active oxides, and (B) shows the change of BAT-PA with different active oxides.

[0042] Figure 20 Photographs showing the color changes of BTPAT-PA and BAT-PA test strips in aqueous solutions of different metal ions;

[0043] Figure 21 In the image, (A) shows the image changes of BTPAT-PA filter paper treated with TFA and TEA steam, and (B) shows the image changes of BAT-PA filter paper treated with TFA and TEA steam.

[0044] Figure 22 The images show the cellular fluorescence imaging of compounds BTPAT-PA and BAT-PA, where A is the dark-field fluorescence imaging of the blank, B is the bright-field imaging of the blank, and C is the superimposed field imaging of the blank. Detailed Implementation

[0045] This invention provides an organic fluorescent dye compound having a DAD structure, which has the following characteristics:

[0046] The structure of Equation 1 or Equation 2:

[0047]

[0048] This invention provides a method for preparing the above-mentioned organic fluorescent dye compound having a DAD structure, characterized by comprising the following steps:

[0049] S1. Under a protective gas atmosphere, a tricyclic compound, a thiophene compound, a palladium catalyst, an alkaline compound solution, and a solvent are mixed and subjected to a Suzuki coupling reaction to obtain an intermediate product.

[0050] S2. Under a protective gas atmosphere, the intermediate product, 2-amino-4,6-dimethylpyrimidine, tetrabutylammonium hydrogen sulfate, alkaline compound solution and solvent are mixed and subjected to Knauvengel condensation reaction to obtain an organic fluorescent dye compound with a DAD structure.

[0051] In this invention, the reaction process for preparing Formula 1 is as follows:

[0052]

[0053] In this invention, the reaction process for preparing Formula 2 is as follows:

[0054]

[0055] In this invention, the tricyclic compound is (4-(diphenylamine)phenyl)boronic acid or 9-bromoanthracene, preferably (4-(diphenylamine)phenyl)boronic acid; the thiophene compound is 5-bromothiophene-2-carboxaldehyde or 2-boronic acid-thiophene, preferably 5-bromothiophene-2-carboxaldehyde.

[0056] In this invention, the palladium catalyst is one or more of Pd(PPh3)4, PdCl2(dppf)2, Pd(dppf)Cl2 and Pd(OAc)2, preferably Pd(PPh3)4, Pd(dppf)Cl2 and Pd(OAc)2, and more preferably Pd(PPh3)4 and / or Pd(OAc)2.

[0057] In this invention, the concentration of the alkaline compound solution in steps S1 and S2 is independently 1–10 mol / L, preferably 2–8 mol / L, and more preferably 4–6 mol / L; the alkaline compound is independently one or more of Na2CO3, Ba(OH)2, K3PO4, Cs2CO3, K2CO3, TiOH, KF, CsF, TBAF, NaOH, and diethylisopropylamine, preferably one or more of Na2CO3, Ba(OH)2, K3PO4, Cs2CO3, K2CO3, TiOH, KF, NaOH, and diethylisopropylamine. The solvent is preferably one or more of Na2CO3, Ba(OH)2, K3PO4, K2CO3, KF, NaOH, and diethylisopropylamine; the protective gas in steps S1 and S2 is independently nitrogen, argon, or neon, preferably nitrogen or argon, and more preferably nitrogen; the solvent in steps S1 and S2 is independently one or more of THF, CH2Cl2, DMF, NaOH, and CH3CN, preferably one or more of THF, DMF, NaOH, and CH3CN, and more preferably one or more of THF, DMF, and CH3CN.

[0058] In this invention, in step S1, the molar ratio of the tricyclic compound and the thiophene compound is 1:1 to 2, preferably 1:1.2 to 1.8, and more preferably 1:1.4 to 1.6; the palladium catalyst accounts for 5 to 15 mol% of the total reactants, preferably 8 to 12 mol%, and more preferably 10 mol%; the molar volume of the thiophene compound, the alkaline compound solution, and the solvent is 1 to 100 mmol: 60 to 120 mL: 80 to 250 mL, preferably 3 to 80 mmol: 70 to 110 mL: 100 to 220 mL, and more preferably 10 to 50 mmol: 80 to 100 mL: 150 to 200 mL.

[0059] In this invention, in step S1, the temperature of the Suzuki coupling reaction is 80-90°C, preferably 82-86°C, and more preferably 85°C; the time of the Suzuki coupling reaction is 11-13 hours, preferably 11.5-12.5 hours, and more preferably 12 hours.

[0060] In this invention, after the Suzuki coupling reaction is completed, the reaction product is cooled to room temperature and the solvent is removed by rotary evaporation to obtain the rotary evaporation residue; the rotary evaporation residue is extracted, and the obtained organic phase is dried and filtered in sequence to obtain the filtrate; the filtrate is separated by column chromatography to obtain the intermediate product.

[0061] In this invention, the extractant used for extraction is preferably dichloromethane; the drying agent used for drying is preferably anhydrous magnesium sulfate; and the eluent used for column chromatography separation is preferably petroleum ether.

[0062] In this invention, in step S2, the molar ratio of the intermediate product to 2-amino-4,6-dimethylpyrimidine is 1:1 to 2, preferably 1:1.2 to 1.8, and more preferably 1:1.4 to 1.6; the molar volume ratio of the intermediate product, the alkaline compound solution, and the solvent is 1 to 3000 mmol: 30 to 120 mL: 60 to 150 mL, preferably 10 to 1000 mmol: 50 to 100 mL: 80 to 140 mL, and more preferably 20 to 800 mmol: 60 to 80 mL: 100 to 120 mL; the tetrabutylammonium bisulfate is 20 to 40 mol% of the total reactants, preferably 25 to 35 mol%, and more preferably 30 mol%.

[0063] In this invention, in step S2, the temperature of the Knauvengay condensation reaction is 100-150°C, preferably 110-140°C, and more preferably 120-130°C; the time of the Knauvengay condensation reaction is 8-12 hours, preferably 9-11 hours, and more preferably 10 hours.

[0064] In this invention, after the Knauvengel condensation reaction is completed, the reaction product is cooled to room temperature and then sequentially filtered, washed and dried to obtain the residue; the residue is then separated by column chromatography to obtain the organic fluorescent dye compound.

[0065] In this invention, the drying agent used for drying is preferably anhydrous magnesium sulfate; the eluent used for column chromatography separation is preferably a mixture of petroleum ether and ethyl acetate, and the volume ratio of petroleum ether to ethyl acetate is preferably 5 to 8:1, more preferably 6 to 7:1.

[0066] This invention also provides the application of the above-mentioned organic fluorescent dye compounds with DAD structures in organic optoelectronic materials, biochemical detection, or cell imaging.

[0067] The technical solutions provided by the present invention will be described in detail below with reference to the embodiments, but they should not be construed as limiting the scope of protection of the present invention.

[0068] Example 1

[0069] Under nitrogen protection, 5-bromothiophene-2-carboxaldehyde (0.38 g, 2.00 mmol) and Pd(PPh3)4 (10 mol%) were dissolved in THF (80.0 mL). After stirring for 30 min, (4-(diphenylamino)phenyl)boric acid (0.43 g, 1.50 mmol) and 60 mL of 5.0 mol / L Na2CO3 solution were added. The mixture was heated under reflux at 85 °C for 12 h, and the reaction was stopped and cooled to room temperature. The solvent was removed by rotary evaporation, and the mixture was extracted with dichloromethane. The organic phases were combined and dried over anhydrous magnesium sulfate. The mixture was filtered, and the solvent was removed by rotary evaporation of the filtrate. Column chromatography was performed using petroleum ether as the eluent to obtain 0.30 g of green solid 5-(4-(diphenylamino)phenyl)thiophene-2-carboxaldehyde, with a yield of 75%.

[0070] Under nitrogen protection, 2-amino-4,6-dimethylpyrimidine (0.37 g, 3.00 mmol) was dissolved in NaOH solution (5.0 mol / L, 30 mL), stirred for 10 min, and then 5-(4-(diphenylamine)phenyl)thiophene-2-carboxaldehyde (2.13 g, 3.00 mmol), TBAHS (0.099 g, 3.0 mmol) and THF (80.0 mL) were added. The mixture was heated under reflux at 120 °C for 12 h, the reaction was stopped, and the mixture was cooled to room temperature. The reaction mixture was then filtered, washed with water, and purified by neutral alumina column chromatography using petroleum ether / ethyl acetate (v / v = 5:1) as the eluent to give a yellow-orange solid 4,4-((2-aminopyrimidine-4,6-diyl)bis(ethylene-2,1-diyl))bis(thiophene-5,2-diyl)bis(N,N-diphenylaniline) (1.27 g, 1.60 mmol), named BTPAT-PA, with a yield of 53.3%.

[0071] Figure 1 The proton NMR spectrum of the BTPAT-PA prepared in this embodiment is as follows: 1 H NMR (500MHz, DMSO-d6): δ6.48(s,2H),6.72(d,J=15.0Hz,2H),6.85(s,1H),6.98(d,J=10.0Hz,4H),7.15– 7.04(m,12H),7.34(t,J=7.5Hz,10H),7.42(d,J=5.0Hz,2H),7.61(d,J=10Hz,4H),7.87(d,J=20.0Hz,2H).

[0072] Example 2

[0073] Under nitrogen protection, 9-bromoanthracene (0.38 g, 1.5 mmol) and Pd(PPh3)4 (10 mol%) were dissolved in 80.0 mL of THF. After stirring for 30 min, 2-thiopheneboronic acid (0.31 g, 2.0 mmol) and 60 mL of 5.0 mol / L Na2CO3 solution were added. The mixture was heated under reflux at 85 °C for 12 h, and the reaction was stopped and cooled to room temperature. The solvent was removed by rotary evaporation, and the mixture was extracted with dichloromethane. The organic phases were combined and dried over anhydrous magnesium sulfate. The mixture was filtered, and the solvent was removed by rotary evaporation of the filtrate. Column chromatography was performed using petroleum ether as the eluent to obtain 0.35 g of the green solid 9-(5-thiophenealdehyde)anthracene, with a yield of 66%.

[0074] Under nitrogen protection, 2-amino-4,6-dimethylpyrimidine (0.37 g, 3.00 mmol) was dissolved in NaOH solution (5.0 mol / L, 30 mL). After stirring for 10 min, 9-(5-thienyl)anthracene (1.73 g, 6.00 mmol), TBAHS (0.099 g, 3.0 mmol), and 80.0 mL of THF were added. The mixture was heated under reflux at 120 °C for 12 h, and the reaction was stopped and cooled to room temperature. The reaction mixture was then filtered, washed with water, and purified by neutral alumina column chromatography using petroleum ether / ethyl acetate (v / v = 5:1) as the eluent to give a yellow-green solid, 4,6-bis(2-(9-(5-thienyl)anthracene-2-vinyl))pyrimidine-2-amino (0.79 g, 1.20 mmol), named BAT-PA, in 40% yield.

[0075] Figure 2 The proton NMR spectrum of BAT-PA prepared in this embodiment is as follows: 1H NMR (500MHz, DMSO-d6): δ6.53(s,2H),6.84(d,J=15.0Hz,2H),6.96(s,1H),7.28(s,2H),7.56(m,8H ), 7.63 (s, 2H), 7.85 (d, J = 10.0Hz, 4H), 8.06 (d, J = 15.0Hz, 2H), 8.19 (d, J = 10.0Hz, 4H), 8.78 (s, 2H).

[0076] Performance Characterization

[0077] Figure 3 In the diagram, (A) is the fluorescence emission spectrum of compound BTPAT-PA, (B) is the fluorescence emission spectrum of compound BAT-PA, (C) is the ultraviolet absorption spectrum of compound BTPAT-PA, and (D) is the ultraviolet absorption spectrum of compound BAT-PA. Figure 3 It can be seen that with the increase of solvent polarity, the fluorescence emission wavelengths of compounds BTPAT-PA and BAT-PA in different solvents show a significant Stokes shift. The absorption intensities of both BTPAT-PA and BAT-PA are slightly enhanced, indicating that both BTPAT-PA and BAT-PA have small dipole moments and are not easily affected by the increase in ground-state solvent polarity.

[0078] Figure 4 (A) in the figure is the absorption spectrum variation curve of BTPAT-PA (inset: H) + (A) Effect of concentration on absorption intensity at 530 nm, (B) is the emission intensity variation curve of BTPAT-PA (inset: H) + The effect of concentration on emission intensity at 574 nm, λ ex =390nm). (C) shows the absorption spectrum change of BAT-PA (inset: H) + (D) shows the effect of concentration on absorption intensity at 463 nm, and (H) shows the change in emission intensity of BAT-PA (inset: H). + The effect of concentration on emission intensity at 545 nm), where the concentrations of BTPAT-PA and BAT-PA are both 2.0 × 10⁻⁶. -5 mol / L. From Figure 4It was found that in acid-base titrations with an acid concentration of 0.01 mol / L, the maximum absorption values ​​of both BTPAT-PA and BAT-PA gradually decreased, and new peaks and obvious isoabsorption points appeared. Furthermore, the fluorescence spectra of both exhibited a certain degree of quenching accompanied by a significant redshift, proving that both BTPAT-PA and BAT-PA underwent protonation, with the protonation process primarily acting on the N-1 of pyrimidines. This reveals that they both possess high-contrast acidic staining and sensitive reaction characteristics.

[0079] Figure 5 In the diagram, (A) shows the absorption spectrum of BTPAT-PA in solutions with different pH values, (B) shows the fluorescence spectrum of BTPAT-PA in solutions with different pH values, (C) shows the absorption spectrum of BAT-PA in solutions with different pH values, and (D) shows the fluorescence spectrum of BAT-PA in solutions with different pH values. Figure 5 It can be seen that at pH > 7, there is no significant change in the absorption and fluorescence of either, indicating that the alkali has little effect on BTPAT-PA and BAT-PA. However, at pH 3–6, the spectral changes are similar to those observed in other samples. Figure 2 The basic agreement demonstrates that the protonation process of BTPAT-PA and BAT-PA acts on the N-1 of pyrimidine. As the pH is further reduced to 1 and 2, new peaks are clearly observed in the absorption peaks of BTPAT-PA and BAT-PA, indicating that BTPAT-PA and BAT-PA achieve multi-stage protonation of the pyrimidine amino group under extremely acidic conditions.

[0080] Figure 6 Photographs showing the color changes of BTPAT-PA and BAT-PA on the test strips in solutions with different pH values. Figure 6 It can be seen that both BTPAT-PA and BAT-PA can be visually identified as having high-contrast color changes on test strips with solutions of different pH values.

[0081] Figure 7 In the image, (A) is the absorption spectrum of BTPAT-PA after adding equal amounts of various metal ions, and (B) is the fluorescence spectrum of BTPAT-PA after adding equal amounts of various metal ions. Figure 7 It can be concluded that, compared with other metal ions, BTPAT-PA has a greater effect on Al. 3+ Cr 3+ , and Fe 3+ It is highly selective.

[0082] Figure 8 For BTPAT-PA in THF for Al 3+ Cr 3+ and Fe 3+The titration absorption spectrum change curve is shown, where (A) represents Al. 3+ / EDTA-induced absorption spectrum change curve, (B) is Cr 3+ / EDTA-induced absorption spectrum change curve, (C) is Fe 3+ The graph shows the EDTA-induced absorption spectrum changes, where the concentration of BTPAT-PA is 2.0 × 10⁻⁶. -5 mol / L.

[0083] Figure 9 For BTPAT-PA in THF for Al 3+ Cr 3+ and Fe 3+ The titration fluorescence spectrum change curve, where (A) represents Al. 3+ / EDTA-induced fluorescence spectrum change curve, (B) is Cr 3+ / EDTA-induced fluorescence spectrum change curve, (C) is Fe 3+ / EDTA-induced fluorescence spectrum change curve, where the concentration of BTPAT-PA is 2.0 × 10 -5 mol / L.

[0084] Figure 10 In the diagram, (A) is the absorption spectrum of BAT-PA after adding equal amounts of various metal ions, and (B) is the fluorescence spectrum of BAT-PA after adding equal amounts of various metal ions. Figure 10 It can be concluded that, compared with other metal ions, BAT-PA has a greater effect on Al. 3+ Cr 3+ , and Fe 3+ It is highly selective.

[0085] Figure 11 For BAT-PA in THF for Al 3+ Cr 3+ and Fe 3+ The titration absorption spectrum change curve is shown, where (A) represents Al. 3+ / EDTA-induced absorption spectrum change curve, (B) is Cr 3+ / EDTA-induced absorption spectrum change curve, (C) is Fe 3+ / EDTA-induced absorption spectrum change curve, where the concentration of BAT-PA is 2.0×10 -5 mol / L.

[0086] Figure 12 For BAT-PA in THF for Al 3+ Cr 3+ and Fe 3+The titration fluorescence spectrum change curve, where (A) represents Al. 3+ / EDTA-induced fluorescence spectrum change curve, (B) is Cr 3+ / EDTA-induced fluorescence spectrum change curve, (C) is Fe 3+ / EDTA-induced fluorescence spectrum change curve, where the concentration of BAT-PA is 2.0×10 -5 mol / L.

[0087] Figure 13 In the image, (A) shows the change of BTPAT-PA with different metal ions, and (B) shows the change of BAT-PA with different metal ions.

[0088] Figure 14 In the image, (A) shows the absorption spectrum of BTPAT-PA after adding equal amounts of various active oxides, and (B) shows the fluorescence spectrum of BTPAT-PA after adding equal amounts of active oxides, where the concentration of BTPAT-PA is 2.0 × 10⁻⁶. -5 mol / L. From Figure 14 It can be seen that BTPAT-PA exhibits high selectivity for Hcy and ·OH compared to other active oxides.

[0089] Figure 15 The figures show the titration absorption and fluorescence spectra of BTPAT-PA in THF induced by ·OH. (A) shows the ·OH-induced absorption spectrum change, and (B) shows the ·OH-induced fluorescence spectrum change. The concentration of BTPAT-PA is 2.0 × 10⁻⁶. -5 mol / L.

[0090] Figure 16 The figures show the titration absorption and fluorescence spectra of BTPAT-PA in THF in response to Hcy, with (A) showing the Hcy-induced absorption spectrum and (B) showing the Hcy-induced fluorescence spectrum. The concentration of BTPAT-PA is 2.0 × 10⁻⁶. -5 mol / L.

[0091] Figure 17 In the diagram, (A) shows the absorption spectrum of BAT-PA after adding equal amounts of various active oxides, and (B) shows the fluorescence spectrum of BAT-PA after adding equal amounts of active oxides. Figure 17 It can be seen that BAT-PA exhibits high selectivity for Hcy and ·OH compared to other active oxides.

[0092] Figure 18The figures show the titration absorption and fluorescence spectra of BAT-PA in THF induced by ·OH. Figure (A) shows the ·OH-induced absorption spectrum change, and figure (B) shows the ·OH-induced fluorescence spectrum change. The concentration of BAT-PA is 2.0 × 10⁻⁶. -5 mol / L.

[0093] Figure 19 In the diagram, (A) shows the change of BTPAT-PA with different active oxides, and (B) shows the change of BAT-PA with different active oxides.

[0094] Figure 20 Photographs showing the color changes of BTPAT-PA and BAT-PA test strips in aqueous solutions of different metal ions.

[0095] Figure 21 In the image, (A) shows the image changes of BTPAT-PA filter paper treated with TFA and TEA steam, and (B) shows the image changes of BAT-PA filter paper treated with TFA and TEA steam.

[0096] Application examples

[0097] HeLa cells were cultured in bovine serum supplemented with 10% (w / v) DMEM at 37°C under a 5% CO2 and 95% air atmosphere. The HeLa cells were then stored overnight in 20 mm cell culture dishes. After washing the HeLa cells with phosphate-buffered saline (PBS), the cell staining material (i.e., BTPAT-PA or BAT-PA, both at a concentration of 10 μmol / L) was added to the culture medium and incubated for 30 minutes. After washing the HeLa cells three times with PBS, the cells were imaged using an OLYMPUS FV1000 confocal laser scanning microscope, using fluorescence emission at 500–650 nm as the collection channel. The results are shown below. Figure 22 As shown, Figure 22 These are cellular fluorescence imaging images of compounds BTPAT-PA and BAT-PA. A1 is the dark-field fluorescence image of the blank sample, B1 is the bright-field fluorescence image of the blank sample, and C1 is the superimposed field image of the blank sample. A2 is the dark-field fluorescence image of compound BTPAT-PA, B2 is the bright-field fluorescence image of compound BTPAT-PA, and C2 is the superimposed field image of compound BTPAT-PA. A3 is the dark-field fluorescence image of compound BAT-PA, B3 is the bright-field fluorescence image of compound BAT-PA, and C3 is the superimposed field image of compound BAT-PA. Figure 22As can be seen, both compounds provided by this invention can effectively enter HeLa cells, and both compounds exhibit good cell staining function. Therefore, the compounds provided by this invention can be used as cell fluorescent markers and have potential application value in the field of cell dyes.

[0098] The above description is only a preferred embodiment of the present invention. It should be noted that for those skilled in the art, several improvements and modifications can be made without departing from the principle of the present invention, and these improvements and modifications should also be considered within the scope of protection of the present invention.

Claims

1. An organic fluorescent dye compound having a DAD structure, characterized in that, It has the structure of either Equation 1 or Equation 2:

2. The method for preparing the organic fluorescent dye compound with a DAD structure according to claim 1, characterized in that, Includes the following steps: S1. Under a protective gas atmosphere, a tricyclic compound, a thiophene compound, a palladium catalyst, an alkaline compound solution, and a solvent are mixed and subjected to a Suzuki coupling reaction to obtain an intermediate product. S2. In a protective gas atmosphere, the intermediate product, 2-amino-4,6-dimethylpyrimidine, tetrabutylammonium hydrogen sulfate, alkaline compound solution and solvent are mixed and subjected to Knauvengel condensation reaction to obtain an organic fluorescent dye compound with a DAD structure. The tricyclic compound is (4-(diphenylamine)phenyl)boronic acid or 9-bromoanthracene; the thiophene compound is 5-bromothiophene-2-carboxaldehyde or 2-boronic acid-thiophene. The structural formula of the intermediate product is selected from one of the following chemical formulas:

3. The preparation method according to claim 2, characterized in that, The palladium catalyst is one or more of Pd(PPh3)4, PdCl2(dppf)2, Pd(dppf)Cl2 and Pd(OAc)2.

4. The preparation method according to claim 2 or 3, characterized in that, The concentration of the alkaline compound solution in steps S1 and S2 is independently 1–10 mol / L; the alkaline compound is independently one or more of Na2CO3, Ba(OH)2, K3PO4, Cs2CO3, K2CO3, KF, CsF, TBAF, NaOH, and diethylisopropylamine; the protective gas in steps S1 and S2 is independently nitrogen, argon, or neon; the solvent in steps S1 and S2 is independently one or more of THF, CH2Cl2, DMF, NaOH, and CH3CN.

5. The preparation method according to claim 4, characterized in that, In step S1, the molar ratio of the tricyclic compound and the thiophene compound is 1:1 to 2; the palladium catalyst accounts for 5 to 15 mol% of the total reactants; and the molar volume of the thiophene compound, the alkaline compound solution, and the solvent is 1 to 100 mmol: 60 to 120 mL: 80 to 250 mL.

6. The preparation method according to claim 2 or 5, characterized in that, In step S1, the temperature of the Suzuki coupling reaction is 80–90°C, and the time of the Suzuki coupling reaction is 11–13 h.

7. The preparation method according to claim 2, 3 or 5, characterized in that, In step S2, the molar ratio of the intermediate product to 2-amino-4,6-dimethylpyrimidine is 1:1 to 2; the molar volume ratio of the intermediate product, the alkaline compound solution, and the solvent is 1 to 3000 mmol: 30 to 120 mL: 60 to 150 mL; and the tetrabutylammonium hydrogen sulfate is 20 to 40 mol% of the total reactants.

8. The preparation method according to claim 7, characterized in that, In step S2, the temperature of the Knauvengay condensation reaction is 100–150°C, and the time of the Knauvengay condensation reaction is 8–12 h.

9. The use of the organic fluorescent dye compound with the DAD structure as described in claim 1 in the preparation of organic optoelectronic materials, biochemical detection agents, or cell imaging agents.

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