Amino-functionalized Nile blue dye as well as synthesis method and application thereof

Abstract

The invention discloses amino-functionalized Nile blue dye as well as a synthesis method and application thereof, and belongs to the field of organic synthesis. According to the method, by optimizing the catalyst type, the reaction temperature and the solvent system, the generation of isomer byproducts is effectively inhibited, and the reaction selectivity is improved. Compared with the prior art, the synthesis process disclosed by the invention has the advantages of simple reaction steps, mild and controllable conditions, high product yield and high purity; meanwhile, large-scale use of strong acid, strong alkali and expensive noble metal catalysts is avoided, and the production cost is remarkably reduced. The obtained amino-functionalized Nile blue dye has long wavelength absorption, excellent cell membrane permeability and wide pH tolerance range (pH 2-11), and can generate superoxide anion free radicals under red light excitation. Based on the characteristics, the compound can be widely applied to the fields of biological fluorescence labeling and functional material preparation.

Description

Technical Field

[0001] This invention belongs to the field of organic synthesis, specifically relating to an amino-functionalized Nile Blue dye, its synthesis method, and its application. Background Technology

[0002] Nile Blue, also known as Nile Blue sulfate, is a benzophenoxazine compound. Due to its excellent photophysical and photochemical properties, it shows broad application prospects in fields such as biological fluorescent staining, laser dyes, optoelectronic devices, and chemical sensing. The conjugated system in its parent structure endows it with strong absorption and emission properties in the visible light region, making it a commonly used labeled probe in analytical chemistry and biological imaging research.

[0003] Currently, the synthesis of amino-functionalized Nile blue dyes mainly relies on post-modification of existing Nile blue backbones or multi-step construction starting from simple precursors. However, existing synthetic routes have revealed the following technical shortcomings in practical applications:

[0004] (1) Cumbersome reaction steps: Traditional methods often require multiple steps such as nitration and reduction, which are lengthy and not only prolong the production cycle but also increase the risk of product loss.

[0005] (2) Regioselectivity is difficult to control: Since the Nile Blue molecule itself has multiple reaction sites, it is easy to generate multiple isomers when directly aminated, which makes it difficult to separate and purify the target product and the yield is low.

[0006] (3) Harsh reaction conditions: Some synthetic routes involve strong acid, strong base or high temperature conditions, which require high equipment and are easy to destroy the original conjugated structure of the dye, affecting its fluorescence performance;

[0007] (4) Complex post-processing: The reaction system often contains a large number of byproducts and unreacted raw materials, requiring multiple column chromatography separations. This not only increases solvent consumption and production costs but also hinders the greening of the process. Therefore, existing methods for synthesizing amino-functionalized Nile Blue dyes still have significant shortcomings in terms of reaction efficiency, product purity, and process economy. There is an urgent need to develop a synthetic process with simple reaction steps, mild conditions, high selectivity, and easy purification to meet the application needs of this functional dye in the fields of life sciences and materials science. Summary of the Invention

[0008] The purpose of this invention is to provide an amino-functionalized Nile Blue dye and its preparation method. The method has simple reaction steps, mild and controllable conditions, is easy to scale up, and the target product has high purity and excellent yield.

[0009] The technical solution of the present invention: an amino-functionalized Nile Blue dye having a structure as shown in general formula I:

[0010]

[0011] In general formula I, R1 and R2 are each independently selected from C1-C5 straight-chain or branched alkyl groups. More preferably, in the embodiments of the present invention, R1 is selected as ethane and R2 is selected as ethane.

[0012] X is selected from oxygen, sulfur, selenium, and tellurium. More preferably, in the embodiments of the present invention, X is selected as oxygen or sulfur.

[0013] Y - Selected from one of halide ions, perchlorate ions, tetrafluoroborate ions, and hexafluorophosphate ions, and more preferably, in the embodiments of the present invention, Y - Chloride ions were selected.

[0014] n is selected from positive integers from 1 to 6, and more preferably, in the embodiments of the present invention, n is 1.

[0015] This invention also discloses a method for preparing the above-mentioned amino-functionalized Nile blue dye, including the preparation of aniline derivatives with targeting groups using aniline or naphthylamine as raw materials, the preparation of oxygen-, sulfur-, or selenium-containing elemental condensation intermediates, and the synthesis steps for preparing amino-functionalized Nile blue dye using the above two intermediates. Specifically, it includes the following steps:

[0016]

[0017] (1) 1-naphthylamine, inorganic base, catalyst and pyridine-2-carboxylic acid were added to an organic solvent at 80℃-100℃ and reacted. After the reaction, the mixture was concentrated and purified under reduced pressure to obtain compound 2.

[0018] (2) Compound 2, dihaloalkane, inorganic base and catalyst were added to an organic solvent at 80℃-100℃ to carry out the reaction. After depressurization, concentration and purification, compound 3 was obtained.

[0019] (3) Compound 3 and an inorganic base were added to an organic solvent at 70℃-90℃ to carry out the reaction. After depressurization, concentration and purification, compound 4 was obtained.

[0020] (4) Synthesis of target dye: Compound 1a or compound 1b is oxidized with compound 4 in an organic solvent at 0-120℃ using an oxidant or catalyzed by a base to form a ring. After the reaction is completed, the mixture is treated with acid, concentrated and purified to obtain the amino-functionalized Nile blue dye NBX-NI as shown in general formula I.

[0021] The preparation of compound 1a is as follows:

[0022]

[0023] Aniline containing R1 and R2 substituents was dissolved in an aqueous solution of aluminum sulfate. NaS2O3 and ZnCl2 were added sequentially under stirring at 0℃-3℃, and an oxidant was added dropwise. After the reaction was complete, the precipitate was obtained by filtration and then purified by recrystallization to obtain compound 1a.

[0024] The preparation of compound 1b is as follows:

[0025]

[0026] Phenol containing R1 and R2 substituents was added to an acid with a mass percentage of 5-50%; sodium nitrite was added at 0℃-3℃, the reaction was allowed to proceed, the mixture was allowed to stand, and the precipitate was obtained by filtration and washing to obtain compound 1b.

[0027] Furthermore, the organic solvent is selected from one or more of dichloromethane, dichloroethane, butanediol, ethanol, acetonitrile, ethyl acetate, acetic acid, and dimethyl sulfoxide.

[0028] Furthermore, the inorganic base is selected from at least one of sodium hydroxide, potassium hydroxide, lithium hydroxide, potassium carbonate, cesium carbonate, calcium carbonate, sodium carbonate, potassium phosphate, and sodium ethoxide.

[0029] Furthermore, the catalyst is selected from at least one of cuprous iodide, cuprous bromide, cuprous chloride, cuprous oxide, copper sulfate, copper sulfate pentahydrate, copper oxide, copper chloride, and silver carbonate.

[0030] Furthermore, the oxidant is selected from at least one of potassium dichromate, potassium permanganate, potassium perchlorate, and potassium chlorate.

[0031] Furthermore, in step (4), the acid used for acid treatment is selected from hydrochloric acid, hydrobromic acid, hydrofluoric acid, hydroiodic acid, perchloric acid, fluoroboric acid, and hexafluorophosphate.

[0032] The acid used in the preparation of compound 1b is selected from hydrochloric acid, sulfuric acid, hydrobromic acid, and acetic acid.

[0033] Furthermore, in step (1), the molar ratio of 1-naphthylamine, inorganic base, catalyst and pyridine-2-carboxylic acid is 1:1-1.5:0.03-0.07:1.5-2.5.

[0034] Furthermore, in step (2), the molar ratio of compound 2, dihaloalkane, inorganic base, and catalyst is 1:2-3:1-1.5:0.03-0.07;

[0035] In step (3), the molar ratio of compound 3 to the inorganic base is 1:1-2;

[0036] In step (4), the molar ratio of compound 1a or compound 1b to compound 4 is 1:1-1.5.

[0037] The application of an amino-functionalized Nile blue dye, specifically its use in the preparation of photodynamic therapy drugs. Specifically, it is used in the preparation of photodynamic therapy drugs for lung cancer.

[0038] The beneficial effects of this invention are:

[0039] 1. This invention adopts a relatively simple amino-functionalized modification strategy, avoiding the cumbersome multi-step covalent modification process. It directly obtains a series of amino-functionalized Nile Blue dyes using a simple and efficient synthetic route. This is a simple and reliable preparation method, providing a brand-new strategy for the large-scale production of Nile Blue dyes.

[0040] 2. The series of amino-functionalized Nile blue dyes synthesized in this invention are cationic molecules that retain the near-infrared absorption characteristics of the Nile blue parent compound, exhibiting deep tissue penetration and low phototoxicity. They also demonstrate good reactive oxygen species generation capacity in aqueous solutions, making them suitable for preparing reagents for tumor cell diagnosis and treatment. Attached Figure Description

[0041] Figure 1 High-resolution mass spectrum of amino-functionalized Nile blue dye NBO-NI.

[0042] Figure 2 This is a high-resolution mass spectrum of the amino-functionalized Nile Blue dye NBS-NI.

[0043] Figure 3 The UV absorption spectra of amino-functionalized Nile Blue dye NBS-NI in different solvents.

[0044] Figure 4 The spectrum is a performance verification spectrum of amino-functionalized Nile Blue dye NBS-NI in generating superoxide radicals under light irradiation; (a) is the spectrum of the control group and (b) is the spectrum of the experimental group.

[0045] Figure 5 The UV absorption spectra of amino-functionalized Nile Blue dye NBS-NI under different pH conditions.

[0046] Figure 6 This is a graph illustrating the in vitro photodynamic therapy capability of the amino-functionalized Nile Blue dye NBS-NI in live cells. Detailed Implementation

[0047] The specific embodiments of the present invention will be further described below with reference to the accompanying drawings and technical solutions.

[0048] An amino-functionalized Nile Blue dye has a structure as shown in general formula I:

[0049]

[0050] In general formula I, R1 and R2 are each independently selected from C1-C5 straight-chain or branched alkyl groups. More preferably, in the embodiments of the present invention, R1 is selected as ethane and R2 is selected as ethane.

[0051] X is selected from oxygen, sulfur, selenium, and tellurium. More preferably, in the embodiments of the present invention, X is selected as oxygen or sulfur.

[0052] Y - Selected from one of halide ions, perchlorate ions, tetrafluoroborate ions, and hexafluorophosphate ions, and more preferably, in the embodiments of the present invention, Y - Chloride ions were selected.

[0053] n is selected from positive integers from 1 to 6, and more preferably, in the embodiments of the present invention, n is 1.

[0054] This invention also discloses a method for preparing the above-mentioned amino-functionalized Nile blue dye, including the preparation of aniline derivatives with targeting groups using aniline or naphthylamine as raw materials, the preparation of oxygen-, sulfur-, or selenium-containing elemental condensation intermediates, and the synthesis steps for preparing amino-functionalized Nile blue dye using the above two intermediates. Specifically, it includes the following steps:

[0055]

[0056] (1) Synthesis of oxygen-containing / sulfur intermediates: Aniline containing R1 and R2 substituents is dissolved in an aqueous solution of aluminum sulfate. NaS2O3 and ZnCl2 are added sequentially under stirring at 0℃-3℃. Oxidizing agent is added dropwise. After the reaction is complete, the precipitate is obtained by filtration and then purified by recrystallization to obtain compound 1a; or, phenol containing R1 and R2 substituents is added to an acid with a mass percentage of 5-50%; sodium nitrite is slowly added under stirring at 0℃-3℃. After the reaction is complete, the precipitate is obtained by filtration and washed to obtain compound 1b;

[0057] (2) At 80-100℃, 1-naphthylamine, inorganic base, catalyst and pyridine-2-carboxylic acid were added to an organic solvent and reacted for 24 h. Compound 2 was obtained by depressurization, concentration and purification.

[0058] (3) At 80-100℃, in an organic solvent, the above compound 2, dihaloalkane, inorganic base and catalyst were added and reacted for 12h. After depressurization, concentration and purification, compound 3 was obtained.

[0059] (4) At 70-90℃, in an organic solvent, add the above compound 3 and an inorganic base, react for 24 h, and then obtain compound 4 by depressurization, concentration and purification;

[0060] (5) Synthesis of target dye: Compound 1a or compound 1b is oxidized with compound 4 in an organic solvent at 0-120℃ using an oxidant or catalyzed by a base to form a ring. After the reaction is completed, the mixture is treated with an acid containing Y, concentrated and purified to obtain the amino-functionalized Nile blue dye NBX-NI of general formula I.

[0061] More preferably, in step (1) above, the oxidant is selected from at least one of potassium dichromate, potassium permanganate, potassium perchlorate, and potassium chlorate; and the acid is selected from at least one of hydrochloric acid, sulfuric acid, hydrobromic acid, and acetic acid.

[0062] More preferably, the organic solvent is selected from one or a mixture of several of the following: dichloromethane, dichloroethane, butanediol, ethanol, acetonitrile, ethyl acetate, acetic acid, and dimethyl sulfoxide; the inorganic base is selected from at least one of the following: sodium hydroxide, potassium hydroxide, lithium hydroxide, potassium carbonate, cesium carbonate, calcium carbonate, sodium carbonate, potassium phosphate, and sodium ethoxide; and the catalyst is selected from at least one of the following: cuprous iodide, cuprous bromide, cuprous chloride, cuprous oxide, copper sulfate, copper sulfate pentahydrate, copper oxide, copper chloride, and silver carbonate.

[0063] More preferably, in step (5) above, the acid containing Y is selected from at least one of hydrochloric acid, hydrobromic acid, hydrofluoric acid, hydroiodic acid, perchloric acid, fluoroboric acid, and hexafluorophosphate.

[0064] Example 1: Preparation of amino-functionalized Nile Blue dye:

[0065] The synthesis method of dyes includes the following steps:

[0066]

[0067] Weigh 0.83 g (5.0 mmol) of 3-diethylaminophenol and dissolve it in 10 mL of dilute hydrochloric acid. Place the solution in an ice-water bath, stir, and cool to 0°C. At 0°C, add sodium nitrite aqueous solution (prepared by dissolving 2.5 g of sodium nitrite in 8 mL of deionized water) dropwise over 30 min using a constant-pressure dropping funnel. A significant exothermic reaction was observed during the addition. After the addition was complete, maintain the temperature at 0°C and continue stirring for 4 h. After the reaction was complete, filter the solution. The filter cake was dried to obtain compound 1b (a grayish-white solid). This product did not require further purification and was used directly in the next step. Yield: 0.60 g, 41%.

[0068]

[0069] 1-Naphthylamine (1.43 g, 10.0 mmol), pyridine-2-carboxylic acid (2.46 g, 20.0 mmol), cuprous iodide (0.095 g, 0.5 mmol), and cesium carbonate (3.26 g, 10.0 mmol) were weighed and added to a reaction flask containing 20 mL of dichloromethane. The reaction was carried out under nitrogen protection and stirred at room temperature for 12 h. After the reaction was complete, the mixture was cooled to room temperature, and the solvent was removed by vacuum distillation. The crude product obtained did not require further purification and was used directly in subsequent reactions. The yield was 1.52 g, with a yield of 51%.

[0070]

[0071] Compound 2 (2.48 g, 10.0 mmol), cuprous iodide (0.095 g, 0.5 mmol), and cesium carbonate (3.26 g, 10 mmol) were weighed and added to 20 mL of dichloroethane. The mixture was reacted at 90 °C for 24 h under nitrogen protection. After the reaction was complete, the solvent was removed by vacuum distillation. No further purification was required, and the mixture was used directly in the next step. The yield was 2.53 g, with a yield of 70%.

[0072]

[0073] Compound 3 (3.10 g, 10.0 mmol) and sodium hydroxide (0.6 g, 15 mmol) were weighed and added to 20 mL of ethanol. The mixture was reacted at 90 °C for 24 h under nitrogen protection. After the reaction was complete, the solvent was removed by vacuum distillation. No further purification was required, and the mixture was used directly in the next step. The yield was 0.97 g, with a yield of 57%.

[0074]

[0075] Compound 4 (34 mg, 0.2 mmol, 1 equivalent) and compound 1b (23 mg, 0.1 mmol, 1 equivalent) were stirred and heated to 115 °C in HOAc (2 mL) for 4 h, then cooled to room temperature. The mixture was concentrated and purified by silica gel column chromatography (V... 二氯甲烷 V 甲醇 =10:1), yielding a deep blue solid, which is the amino-functionalized Nile Blue dye NBO-NI, MS(ESI): m / z for C 22 H 22 N3O + ([M]+): Calculated value 344.1757; Measured value 344.1762.

[0076] Example 2: Preparation of amino-functionalized Nile Blue dye:

[0077] The synthesis method of dyes includes the following steps:

[0078]

[0079] Weigh 3.66 g (5.5 mmol) of aluminum sulfate octadechydrate and dissolve it thoroughly in 10 mL of deionized water. Add N,N-diethyl-p-phenylenediamine (0.83 g, 5.0 mmol) to the aluminum sulfate solution and stir until homogeneous. Then, add sodium thiosulfate pentahydrate (2.85 g, 11.5 mmol) and zinc chloride (0.72 g, 5.25 mmol) sequentially. Cool to 0°C in an ice-water bath. Separately, dissolve 0.44 g (1.5 mmol) of potassium dichromate in 4 mL of deionized water. Under ice-bath cooling and vigorous stirring, slowly add this potassium dichromate solution to the above reaction system using a constant-pressure dropping funnel. After the addition is complete, continue stirring at a constant temperature in an ice bath for about 2 hours. After the reaction is complete, collect the precipitated solid by filtration. Transfer the obtained solid to 12 mL of methanol, heat under reflux for 10 min to remove impurities, then cool to room temperature and filter again. After vacuum drying, the filter cake yielded compound 1a (a grayish-white solid). This product required no further purification and was used directly in the next coupling reaction.

[0080] The synthesis of intermediate compound 2-4 in NBS-NI is the same as that of intermediate compound 2-4 in NBO-NI.

[0081]

[0082] Compound 4 (34 mg, 0.2 mmol, 1 equivalent) and compound 1a (23 mg, 0.1 mmol, 1 equivalent) were weighed and added to 3 mL of dimethyl sulfoxide. Potassium dichromate (0.162 g, 0.55 mmol) was slowly added, and the mixture was stirred at room temperature for 10 min. Then, 50 mL of methanol and 6 mL of dilute hydrochloric acid solution (1.0 mol / L) were added, and the reaction was continued at room temperature for another 40 min. After the reaction was complete, the solvent was removed by rotary evaporation under reduced pressure, and the mixture was purified by column chromatography (V...). 二氯甲烷 V 甲醇 =10:1) yielded a deep blue solid, which is the amino-functionalized Nile Blue dye NBS-NI. MS(ESI): m / z for C 22 H 22 N3S + ([M]+): Calculated value 360.1529; Measured value 360.1535.

[0083] Example 3:

[0084] The equipment and feed ratio were the same as in Example 1, but the catalyst in the synthesis of intermediate compound 2 was replaced with 1.0 mmol of cuprous bromide, and the solvent was replaced with dimethyl sulfoxide. The reaction was carried out at room temperature with stirring for 12 h. After the reaction, post-processing was performed in the same manner as in Example 1. The yield was 1.07 g, with a yield of 36%.

[0085] The reaction continued smoothly even after replacing it with cuprous bromide, demonstrating the universality of copper-based catalytic systems.

[0086] Example 4:

[0087] The equipment and raw material feed ratio were the same as in Example 1, but the catalyst in the synthesis of intermediate compound 3 was replaced with 1.0 mmol of cuprous bromide, and the reaction temperature was set at 90°C. The reaction was carried out under constant temperature and stirring for 24 hours. After the reaction, post-processing was performed in the same manner as in Example 1. The yield was 1.89 g, with a yield of 52%.

[0088] The reaction continued smoothly after being replaced with cuprous bromide, demonstrating the universality of the copper-based catalytic system; however, the reaction rate slowed down and the yield decreased, further establishing cuprous iodide as the optimal catalyst.

[0089] Example 5: Spectral performance testing of NBS-NI:

[0090] To evaluate the photophysical properties of NBS-NI in Example 2, the UV-Vis absorption spectra of this dye in various solvents of different polarities were tested. The test results are as follows: Figure 3 As shown, the dye was dissolved in dimethyl sulfoxide (DMSO), water (H2O), ethanol (EtOH), methanol (MeOH), acetonitrile (ACN), N,N-dimethylformamide (DMF), acetone (Acetone), tetrahydrofuran (THF), and phosphate-buffered saline (PBS) to prepare test solutions. The dye exhibited strong absorption in the visible to near-infrared region. In most test solvents (such as water, alcohols, THF, PBS, etc.), the maximum absorption peak (λmax) of this compound was mainly concentrated between approximately 630 nm and 650 nm, with a shoulder peak near 590 nm. The test results indicate that NBS-NI has excellent long-wavelength absorption, a characteristic that facilitates deep tissue penetration and effectively avoids interference from biological autofluorescence. Simultaneously, the dye's high sensitivity to solvent polarity / microenvironment makes it a promising candidate for applications in complex biological microenvironments and specific solvent identification.

[0091] Example 6: Superoxide radical generation capacity test of NBS-NI:

[0092] To determine whether NBS-NI in Example 2 could release superoxide anion radicals under red light excitation, dihydrorhodamine 123 (DHR123) was used as a metric for superoxide anion radicals (·O2). − The fluorescence spectrum of DHR123 was determined using an indicator. Because DHR123 is in O2... − In the presence of DHR123, it will be oxidized to DHR123, which has strong green fluorescence. Therefore, the generation of O2 by the photosensitizer can be evaluated by observing the increase in fluorescence emission intensity at 540 nm in the system in DMSO solvent. − The ability. For example... Figure 4 As shown, the control group is shown in the left figure (a), and the experimental group is shown in the right figure (b). In the blank control group without NBS-NI, after irradiation with an LED light source with a wavelength of 640-660 nm for 0-4 min, the fluorescence emission peak intensity of the system near 520 nm to 530 nm remained almost unchanged, consistently maintaining a very low baseline level, effectively eliminating background interference from spontaneous oxidation of the probe. In the experimental group, after irradiation with a mixed solution of NBS-NI dissolved in DMSO and DHR123 for only 60 s, the fluorescence emission of DHR123 at 540 nm showed a significant enhancement. This comparative result fully demonstrates that the NBS-NI described in this invention can efficiently undergo photochemical reactions and sensitize the generation of a large number of reactive oxygen species such as superoxide anion radicals under red light excitation.

[0093] Example 7 pH tolerance test of NBS-NI:

[0094] To evaluate the pH tolerance of NBS-NI in Example 2, the changes in the UV absorption spectrum of NBS-NI in Example 2 between pH = 2 and 11 were tested. Figure 5 As shown, the absorption spectrum of Example NBS-NI does not change significantly in acidic and neutral solution environments, indicating the dye's acid tolerance. However, in alkaline environments, even at pH = 11, the absorption spectrum of Example NBS-NI is minimally affected, demonstrating significant alkali tolerance. Therefore, the amino-functionalized Nile Blue dye molecules constructed in this invention possess excellent acid and alkali tolerance, overcoming the traditional limitation that Nile Blue-like structures are unusable in alkaline environments.

[0095] Example 8: Evaluation of Cellular Photodynamic Therapy Capability:

[0096] Cells were stained using calcein-AM and propidium iodide (PI) to detect the cytotoxic effect of NBS-NI on tumor cells in Example 2. Calcein-AM only stains live cells, producing green fluorescence, while PI, as a nuclear staining dye, only stains dead cells, producing red fluorescence. Figure 6As shown, after irradiation with a 640-660nm LED lamp, NBS-NI-treated A549 cells showed almost no signal in the green channel but a significant signal in the red channel, indicating that the A549 cells were almost completely dead. Conversely, NBS-NI-treated cells showed strong fluorescence only in the green channel in the absence of light, indicating that cell death was negligible. This suggests that the amino-functionalized Nile Blue dye exhibits good tumor cell activity (low dark toxicity) after endocytosis under dark conditions, while under light conditions, a large number of tumor cells are killed and cell activity is very low (high phototoxicity).

[0097] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention, and not to limit them; although the present invention has been described in detail with reference to the foregoing embodiments, those skilled in the art should understand that modifications can still be made to the technical solutions described in the foregoing embodiments, or equivalent substitutions can be made to some or all of the technical features; and these modifications or substitutions do not cause the essence of the corresponding technical solutions to deviate from the scope of the technical solutions of the embodiments of the present invention.

Claims

1. An amino-functionalized Nile blue dye, characterized in that, It has a structure as shown in general formula I: ； in: R1 and R2 are each independently selected from C1-C5 straight-chain or branched alkyl groups; X is selected from one of oxygen, sulfur, selenium, and tellurium; Y - one selected from the group consisting of halide ions, perchlorate ions, tetrafluoroborate ions, hexafluorophosphate ions; n is a positive integer selected from 1 to 6.

2. The process for the preparation of an amino-functionalized Nile Blue dye according to claim 1, characterized in that, Includes the following steps: ； (1) 1-naphthylamine, inorganic base, catalyst and pyridine-2-carboxylic acid were added to an organic solvent at 80℃-100℃ and reacted. After the reaction, the mixture was concentrated and purified under reduced pressure to obtain compound 2. (2) Compound 2, dihaloalkane, inorganic base and catalyst were added to an organic solvent at 80℃-100℃ to carry out the reaction. After depressurization, concentration and purification, compound 3 was obtained. (3) Compound 3 and an inorganic base were added to an organic solvent at 70℃-90℃ to carry out the reaction. After depressurization, concentration and purification, compound 4 was obtained. (4) Synthesis of target dye: Compound 1a or compound 1b is oxidized with compound 4 in an organic solvent at 0-120℃ using an oxidant or catalyzed by a base to form a ring. After the reaction is completed, the mixture is treated with acid, concentrated and purified to obtain the amino-functionalized Nile blue dye NBX-NI as shown in general formula I. The preparation of compound 1a is as follows: ； Aniline containing R1 and R2 substituents was dissolved in an aqueous solution of aluminum sulfate. NaS2O3 and ZnCl2 were added sequentially under stirring at 0℃-3℃, and an oxidant was added dropwise. After the reaction was complete, the precipitate was obtained by filtration and then purified by recrystallization to obtain compound 1a. The preparation of compound 1b is as follows: ； Phenol containing R1 and R2 substituents was added to an acid with a mass percentage of 5-50%; sodium nitrite was added at 0℃-3℃, the reaction was allowed to proceed, the mixture was allowed to stand, and the precipitate was obtained by filtration and washing to obtain compound 1b.

3. The method for preparing amino-functionalized Nile blue dye according to claim 2, characterized in that: The organic solvent is selected from one or more of dichloromethane, dichloroethane, butanediol, ethanol, acetonitrile, ethyl acetate, acetic acid, and dimethyl sulfoxide.

4. The method for preparing amino-functionalized Nile blue dye according to claim 2, characterized in that: The inorganic base is selected from at least one of sodium hydroxide, potassium hydroxide, lithium hydroxide, potassium carbonate, cesium carbonate, calcium carbonate, sodium carbonate, potassium phosphate, and sodium ethoxide.

5. The method for preparing amino-functionalized Nile blue dye according to claim 2, characterized in that: The catalyst is selected from at least one of cuprous iodide, cuprous bromide, cuprous chloride, cuprous oxide, copper sulfate, copper sulfate pentahydrate, copper oxide, copper chloride, and silver carbonate.

6. The method for preparing amino-functionalized Nile blue dye according to claim 2, characterized in that: The oxidant is selected from at least one of potassium dichromate, potassium permanganate, potassium perchlorate, and potassium chlorate.

7. The method for preparing amino-functionalized Nile blue dye according to claim 2, characterized in that: In step (4), the acid used for acid treatment is selected from hydrochloric acid, hydrobromic acid, hydrofluoric acid, hydroiodic acid, perchloric acid, fluoroboric acid, and hexafluorophosphate. The acid used in the preparation of compound 1b is selected from hydrochloric acid, sulfuric acid, hydrobromic acid, and acetic acid.

8. The method for preparing amino-functionalized Nile blue dye according to claim 2, characterized in that: In step (1), the molar ratio of 1-naphthylamine, inorganic base, catalyst and pyridine-2-carboxylic acid is 1:1-1.5:0.03-0.07:1.5-2.5; In step (2), the molar ratio of compound 2, dihaloalkane, inorganic base, and catalyst is 1:2-3:1-1.5:0.03-0.07; In step (3), the molar ratio of compound 3 to the inorganic base is 1:1-2; In step (4), the molar ratio of compound 1a or compound 1b to compound 4 is 1:1-1.

5.

9. The application of the amino-functionalized Nile Blue dye according to claim 1, characterized in that: The application of the amino-functionalized Nile blue dye in the preparation of photodynamic therapy drugs.

10. The application of an amino-functionalized Nile Blue dye according to claim 9, characterized in that: The application of the amino-functionalized Nile blue dye in the preparation of photodynamic therapy drugs for lung cancer.

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