CN114591632A

CN114591632A - Azaindole-hemicyanine dye, and synthesis method and application thereof

Info

Publication number: CN114591632A
Application number: CN202210083686.XA
Authority: CN
Inventors: 杜健军; 韩富平; 张寒; 潘静巍; 樊江莉; 彭孝军
Original assignee: Dalian University of Technology
Current assignee: Dalian University of Technology
Priority date: 2022-01-23
Filing date: 2022-01-23
Publication date: 2022-06-07
Anticipated expiration: 2042-01-23
Also published as: CN114591632B

Abstract

The invention discloses azaindole-hemicyanine dyes, a synthesis method and application thereof. This type of dye has longer absorption and emission wavelengths than conventional hemicyanine dyes. And the change of the photophysical properties of the novel dye molecules compared with the traditional hemicyanine dye is verified through related experiments, and the dye has good biocompatibility, and can be applied to the aspects of biological recognition imaging, cell imaging, protein labeling, specific recognition of antibodies, nucleic acid labeling, fluorescent probes, DNA sequencing, tumor photodynamic therapy and the like.

Description

Azaindole-hemicyanine dye, and synthesis method and application thereof

Technical Field

The invention relates to the technical field of organic dyes, in particular to azaindole-hemicyanine dyes, a synthetic method thereof and application thereof.

Background

The hemicyanine dye has the advantages of large Stokes shift, large molar extinction coefficient and the like, so that the hemicyanine dye is continuously used as a dye matrix to be developed into a fluorescent probe or a photosensitizer, and particularly, researchers continuously develop a plurality of fluorescent probes with excellent performance by utilizing the unique optical property of the hemicyanine dye in recent years, and the hemicyanine dye is used for protein labeling, gene sequencing and in-vivo fluorescence imaging; or the hemicyanine dye is used as a matrix to design a photosensitizer which generates active oxygen under the irradiation of light and is used for the photodynamic therapy of tumors in organisms.

Hemicyanine dyes themselves have limitations, particularly in that their parent structures are relatively fixed, which limits the maximum absorption wavelength of the hemicyanine dye molecules to between 650nm and 720nm, and the emission wavelength to between 680nm and 750 nm. For the aspects of in vivo deep imaging, photodynamic and the like, due to the problems of autofluorescence in organisms and the thickness of tissues, the absorption wavelength and the emission wavelength of the needed probe and photosensitizer are longer, and dye molecules which absorb over 720nm and emit over 750nm are more suitable for the application of the scenes. However, there is no relevant literature in the prior art, and how to properly modify the parent of the hemicyanine dye to synthesize a hemicyanine dye structure with longer absorption and emission wavelengths, so that the novel hemicyanine dye has a better application prospect, which is also a technical problem to be solved by those skilled in the art.

Disclosure of Invention

In order to solve the problems, the invention provides a novel azaindole-hemicyanine dye, a synthetic method thereof and application of the dye in the fields of biology and medicine.

A first aspect of the present application is directed to a class of azaindole-hemicyanine dyes having a structural formula as shown in formula I:

In the general formula I, the compound has the following structure,

R₁and R₃Each independently selected from one of hydrogen, halogen, alkyl with 1-18 carbons, carboxyalkyl with 1-18 carbons, aryl, arylcarboxylic acid, alkylsulfonate, arylsulfonate, alkylsulfonate, or arylsulfonate; most preferably one selected from hydrogen, halogen, carboxyalkyl having 1 to 8 carbons, alkylsulfonate;

R₂、R₄and R₅Each independently selected from one of hydrogen, alkyl having 1-18 carbons, carboxyalkyl having 1-18 carbons, hydroxyalkyl having 1-18 carbons, alkylsulfonate having 1-18 carbons, aryl, arylcarboxylic acid, alkylsulfonate, arylsulfonate, alkylsulfonate, or arylsulfonate; more preferably one of hydrogen, alkyl group having 1-8 carbons, carboxyalkyl group having 1-8 carbons, aryl group, alkylsulfonate group having 1-8 carbons, arylcarboxylic acid group; most preferably one of hydrogen, aryl, alkyl having 1 to 8 carbons, alkyl sulfonate having 1 to 8 carbons;

x is selected from one of oxygen, sulfur, selenium and tellurium; most preferably one of oxygen and sulfur;

Y^-selected from halogen ion, ClO₄ ^-、BF₄ ^-、CH₃COO^-、CF₃COO^-Or OTs^-One of (1); most preferably halogen ion, ClO ₄ ^-To (3) is provided.

A second aspect of the present application is directed to a method of synthesizing a class of azaindole-hemicyanine dyes, comprising the steps of:

(1) in an organic solvent at 60-120 deg.C, containing R₁Substituted J-1 and R₂Substituted halogenated alkane (N alkylating reagent) reacts for 3 to 24 hours, and N-R is obtained through recrystallization₂Quaternary ammonium salts of substituted side chains J-2; wherein, contains R₁Substituted compound J-1 and N-alkylating agent R₂The molar ratio of the substituted halogenated alkanes is 1:1 to 10, most preferably 1:3 to 8;

(2) at 20-80 deg.C, adding S-1 and R₃、R₄、R₅Dissolving substituted S-2 in polar solvent, reacting for 10-24h under catalysis of inorganic base, extracting, concentrating, and purifying to obtain the product containing R₃、R₄、R₅A substituted intermediate S-3;

(3) in organic solvent at 50-120 deg.C₁And R₂Substituted J-2 and R₃、R₄、R₅And dissolving the substituted S-3 in an organic solvent, carrying out condensation reaction under the catalysis of organic base, and carrying out recrystallization purification to obtain the aza-hemicyanine near-infrared fluorescent dye I.

With respect to the above-mentioned technical means, preferably, in the step (1), the organic solvent is selected from any one of benzene, toluene, o-dichlorobenzene, and DMF; the recrystallization solvent is selected from one or a mixture of more of methanol, ethanol, acetonitrile, ethyl acetate, diethyl ether, acetone and propanol;

For the technical solution described above, preferably, in the step (2), the polar solvent is one or a mixture of several selected from DMSO, DMF, methanol, and acetonitrile; the inorganic base is selected from any one of sodium hydroxide, potassium carbonate, cesium carbonate, sodium acetate and sodium ethoxide;

with respect to the above technical solution, preferably, in the step (3), the organic solvent is selected from any one or a mixture of several of ethanol, acetic acid, acetic anhydride and DMF;

with respect to the above technical solution, preferably, in the step (3), the solvent used for recrystallization is selected from any one or a mixture of several of methanol, ethanol, acetonitrile, water, ethyl acetate, diethyl ether, acetone and propanol; the organic base is selected from any one of triethylamine, pyridine and DIPEA.

A third aspect of the present application is to protect the use of said azaindole-hemicyanine dyes in the biological and medical field.

For the above-described solutions, more preferred application areas include: the field of deep biological in vivo imaging and tumor treatment of the azaindole-hemicyanine dye; for example: the application in cell imaging, protein labeling, specific recognition of antibodies, nucleic acid labeling, fluorescent probes, DNA sequencing and tumor photodynamic therapy.

For the above technical solution, preferably, the azaindole-hemicyanine dye of the present invention has an excitation wavelength of 600-950nm and a fluorescence detection wavelength of 650-1000nm when applied.

For the above-described solutions, it is preferred that the working concentration of the azaindole-hemicyanine dyes according to the invention is below 12. mu. mol/L.

Compared with the prior art, the beneficial effect of this application:

1. the azaindole-hemicyanine dye provided by the invention has the advantages that the position of quaternary ammonium nitrogen and the type of electron donating groups are changed, so that the molecular charge separation degree and the electron mobility are increased, and the azaindole-hemicyanine dye has longer absorption and emission wavelengths.

Compared with the maximum absorption wavelength of the traditional hemicyanine dye, the azaindole-hemicyanine dye has a large red shift, and the embodiment of the application shows that the maximum absorption wavelength of the compound 1 is 728nm, the maximum absorption wavelength of the compound 2 is 754nm, and the maximum absorption wavelengths are respectively red-shifted by 57nm and 83nm compared with the maximum absorption wavelength 671nm of the traditional hemicyanine dye compound 3.

Compared with the maximum emission wavelength of the traditional hemicyanine dye, the azaindole-hemicyanine dye has a large red shift, and the embodiment of the application shows that the maximum emission wavelengths 719nm of the

compounds

1 and 2 are respectively red-shifted by 45nm and 58nm compared with the maximum emission wavelength 719nm of the compound 3 (traditional hemicyanine dye).

Therefore, the dye is more suitable for being applied to scenes of long-wavelength absorption and emission, and can be well applied to the fields of deep biological in-vivo imaging and tumor treatment.

2. Compared with the traditional hemicyanine dye, the dye provided by the invention has changed photophysical properties and good biocompatibility, and after MCF-7 cells are cultured for 48 hours by using the

compounds

1 and 2 with different concentrations, the cells still show good survival rate, and even if the concentration is increased to 12 mu mol/L, the survival rate of the cells is still high, which indicates that the azaindole-hemicyanine dye has very good biocompatibility and can not generate toxic and side effects on the cells within a working concentration range; can be applied to the aspects of biological recognition imaging, cell imaging, protein labeling, antibody specificity recognition, nucleic acid labeling, fluorescent probes, DNA sequencing, tumor photodynamic therapy and the like.

Drawings

FIG. 1 is a high resolution mass spectrum of Compound 1;

FIG. 2 is a high resolution mass spectrum of Compound 2;

FIG. 3 is a graph of normalized absorption spectra of Compound 1, Compound 2, and Compound 3 in methanol;

FIG. 4 is a normalized fluorescence spectra of compound 1, compound 2 and compound 3 in methanol;

FIG. 5 is a graph of MTT assay for compound 1 and compound 2.

Detailed Description

The present invention will be described in further detail below.

Unless otherwise indicated, the terms used herein have the following meanings.

The term "halogen" as used herein includes fluorine, chlorine, bromine and iodine.

The term "alkyl" as used herein includes straight chain and branched chain alkyl groups.

The term "MTT" as used herein refers to a method for detecting cell survival and growth.

Y is used herein^-Denotes an anion, which may be any suitable anion, including inorganic anions and organic anions, such as but not limited to halogen ions, ClO₄ ^-、PF₆ ^-、BF₄ ^-、CH₃COO^-、CF₃COO^-Or OTs^-。

The instruments and equipment used in the examples:

in the column chromatography process, 200-plus 300-mesh and 100-plus 200-mesh column chromatography silica gel purchased from Qingdao Meigaoji Limited and 20-40-mesh analytical pure quartz sand purchased from Tianda chemical reagent factory are adopted.

In the process of detecting the compound, the mass spectrometer uses a Synapt G2-Si HDMS high-resolution mass spectrometer of waters company in the United states, and adopts a double-spray-needle electrospray ion source to detect the compound in a positive and negative mode.

Dye absorption and emission spectra were measured using the Cary 60 uv-vis spectrophotometer and Cary Eclipse fluorescence spectrophotometer from Agilent corporation.

Cytotoxicity assays were performed using a Varioskan LUX Multimode Microplate Reader instrument from Thermofisher, USA.

EXAMPLE 1 preparation of Compound 1

The structural formula of compound 1:

example 1.1

To 2-hydrazinopyridine (2.182g,20mmol) dissolved in 60mL toluene was added 3-methyl-2-butanone (3.445g,40mmol) at room temperature. Stirring and heating under the protection of nitrogen, refluxing, and stopping reaction after 12h of reaction. And cooling to room temperature. Removing most of toluene, adding 12mL of polyphosphoric acid into the residue, heating and stirring at 140 deg.C for 45min, and reactingThe mixture was poured into 200mL of ice water, ammonia was added dropwise, the pH was adjusted to weak alkalinity, extracted with ethyl acetate and Na anhydrous₂SO₄Drying, evaporation of the solvent and purification on silica gel column afforded compound 1.1 as a nitrogen yellow solid (1.280g,8mmol, Y ═ 40%).

Example 1.2

Compound 1.1(0.640g,4mmol, 1.0eq) was added to 15mL of o-dichlorobenzene, 1, 3-propanesultone (0.977g,8mmol,2.0eq) was added, the reaction was stopped at 65 ℃ for 12h, the reaction was stopped, cooled to room temperature, 100mL of ethyl acetate was added, and the precipitate was collected to give compound 1.2(0.960g,3.4mmol, Y ═ 85%).

Example 1.3

9mL of phosphorus tribromide was slowly added dropwise to a mixed solution of 12mL of DMF and 25mL of chloroform at 0 ℃ and stirred for 1 hour, then the mixture was warmed to room temperature, 4mL of cyclohexanone was added and stirred for 12 hours, and then the reaction was stopped. The solution was introduced into 100mL of ice water and NaHCO was added ₃Adjusting the solution to neutral with CH₂Cl₂Extracting, washing with saturated brine, and removing anhydrous Na₂SO₄Drying and evaporation of the solvent gave the title compound 1.3(6.0g,31.74mmol, Y ═ 73.8%).

Example 1.4

At room temperature, compound 1.1(2.93g,15.5mmol,1.0eq) and 4-diethylamino salicylaldehyde (3.0g,15.5mmol,1.0eq) were added to 15mL of DMF, cesium carbonate (5.05g,15.5mmol,1.0eq) was added, the reaction was stopped after stirring for 16h, the solution was poured into 100mL of water, and CH was used₂Cl₂Extracting, washing with saturated brine, and removing anhydrous Na₂SO₄The organic phase was dried, concentrated and purified by silica gel column to give yellow brown compound 1.4(1.4g,4.94mmol, Y ═ 31.9%).

Production of Compound 1

Compound 1.2(0.100g,0.35mmol,1eq) and compound 1.4(0.149g,0.53mmol,1.5eq) were dissolved in 10mL of acetic anhydride at 100 ℃, 0.2mL of triethylamine was added to catalyze the reaction, the reaction was stopped after stirring for 2 hours, the reaction mixture was cooled to room temperature and then added dropwise to 150mL of ethyl acetate to recrystallize, and the resulting crude product was purified by a silica gel column to give compound 1(0.068g,0.12mmol, Y ═ 35.2%) and a high-resolution mass spectrum chart of fig. 1.

Example 2 preparation of Compound 2

The structural formula of compound 2:

example 2.1

To 2-hydrazino-4-bromopyridine (1.00g,5.32mmol,1.0eq) dissolved in 20mL toluene was added 3-methyl-2-butanone (0.92g,10.64mmol,2.0eq) at room temperature. Stirring and heating under the protection of nitrogen, refluxing, and stopping reaction after 12h of reaction. And cooling to room temperature. Removing most of toluene, adding 12mL of polyphosphoric acid to the residue, heating and stirring at 140 deg.C for 45min, pouring the mixture into 200mL of ice water, adding dropwise ammonia water, adjusting pH to weak alkalinity, extracting with ethyl acetate, and extracting with Na ₂SO₄After drying and evaporation of the solvent, purification was performed by silica gel column chromatography to obtain 2.1(0.760g,3.18mmol, Y59.8%) as a nitrogen yellow solid.

Example 2.2

To compound 2.1(0.500g,2.09mmol, 1.0eq) was added to 15mL of acetonitrile, benzyl bromide (0.715g,4.18mmol,2.0eq) was added, the reaction was stopped at 70 ℃ for 12h, the reaction was cooled to room temperature, added to 100mL of ethyl acetate, and the precipitate was collected to give compound 2.2(0.610g,1.49mmol, Y ═ 71.1%).

Preparation of Compound 2

Compound 1.2(0.100g,0.35mmol,1eq) and compound 2.2(0.217g,0.53mmol,1.5eq) were dissolved in 10mL of acetic anhydride at 100 ℃, 0.2mL of triethylamine was added to catalyze the reaction, the reaction was stopped after stirring for 2 hours, the reaction mixture was cooled to room temperature and then added dropwise to 150mL of ethyl acetate to recrystallize, and the resulting crude product was purified by a silica gel column to give compound 2(0.095g,0.14mmol, Y39.8%), and a high-resolution mass spectrum is shown in fig. 2.

Example 3 production of Compound 3

The structural formula of compound 3 is as follows: used as a comparative example

Example 3.1

Dissolving 4-iodophenylhydrazine (2.00g,8.55mmol,1eq) and 3-methyl-2-butanone (1.47g,17.09mmol,2eq) in 20mL of acetic acid at room temperature, heating and refluxing for 12h under the protection of nitrogen, cooling to room temperature after stopping reaction, removing most of solvent under reduced pressure, pouring the rest solution into saturated NaHCO ₃In aqueous solution, extracted with ethyl acetate and Na₂SO₄Drying, evaporation of the solvent and purification on silica gel column gave yellow-brown compound 3.1(1.85g,6.49mmol, Y75.9%).

Example 3.2

To compound 3.1(0.500g,1.75mmol, 1.0eq) was added to 15mL of acetonitrile, iodomethane (1.37g,8.77mmol,5.0eq) was added, the reaction was stopped at 70 ℃ for 12h, the reaction was cooled to room temperature, and the precipitate was collected by adding to 100mL of diethyl ether to obtain compound 3.2(0.58g,1.31mmol, Y75.0%).

Example 3.3

At room temperature, compound 1.1(2g,10.58mmol,1.0eq) and 4-methoxysalicylaldehyde (1.61g,10.58mmol,1.0eq) were added to 15mL of DMF, cesium carbonate (2g,10.58mmol,1.0eq) was added as stirring for 16h and the reaction was stopped, the solution was poured into 100mL of water, and CH was used₂Cl₂Extracting, washing with saturated brine, and removing anhydrous Na₂SO₄Drying, concentrating the organic phase, dissolving the crude product in 20mL dichloromethane, adding 5mL boron tribromide dropwise in ice bath, stirring at room temperature for 12h, pouring the solution into ice water, adding NaHCO₃Adjusting the aqueous solution to neutral, extracting with ethyl acetate and adding Na₂SO₄Drying, evaporation of the solvent and purification on silica gel column gave yellow-brown compound 3.3(0.75g,3.10mmol, Y ═ 29.3%).

Preparation of Compound 3

Compound 3.3(0.100g,0.438mmol,1eq) and compound 3.2(0.900g,0.6570mmol,1.5eq) were dissolved in 10mL of acetic anhydride at 100 ℃, 0.2mL of triethylamine was added to catalyze the reaction, the reaction was stopped after stirring for 2 hours, the reaction mixture was cooled to room temperature and then added dropwise to 150mL of ethyl acetate to recrystallize, and the resulting crude product was purified by a silica gel column to give compound 3(0.088g,0.135mmol, Y ═ 30.8%).

Example 4 determination of the UV-Vis absorption Spectroscopy and fluorescence Spectroscopy of

Compounds

1, 2 and 3

The dye after vacuum drying is accurately weighed by a ten-thousandth balance, 5mmol/L DMSO dye mother liquor is prepared in a brown sample bottle and stored in a refrigerator at 4 ℃ for standby.

When testing the ultraviolet visible absorption spectrum and the fluorescence spectrum, a micro liquid-transferring gun is used for measuring 1.2 mu L of dye mother liquor, the dye mother liquor is dissolved in a quartz cuvette containing 3mL of a solvent to be tested, the mixture is uniformly mixed, the concentration of the obtained dye is 2.0 mu mol/L, and the dye is used for testing the absorption spectrum and the fluorescence emission spectrum. All tests were done at 25 ℃.

As shown in fig. 3, in the methanol solution, the maximum absorption wavelength of the conventional hemicyanine dye compound 3 is 671nm, while the maximum absorption wavelength of the compound 1 is 728nm and the maximum absorption wavelength of the compound 2 is 754nm, which are red-shifted by 57nm and 83nm, respectively, compared to the compound 3. The maximum absorption wavelength of the novel azaindole-hemicyanine dye is greatly red-shifted compared with that of the traditional hemicyanine dye.

As shown in fig. 4, the maximum emission wavelengths of compound 1 and compound 2 in methanol are 764nm and 777nm, and compared with the maximum emission wavelength of compound 3, 719nm, the maximum emission wavelengths are respectively red-shifted by 45nm and 58nm, which indicates that the maximum emission wavelength of the novel azaindole-hemicyanine dye is more red-shifted than that of the conventional hemicyanine dye, so that the dye is more suitable for being applied to scenes of long-wavelength absorption and emission, and can be well used in the fields of deep-layer in vivo imaging and tumor therapy.

EXAMPLE 5 cytotoxicity assays of Compound 1 and Compound 2

Toxicity of the dye molecules to the cells was assessed by MTT assay. The principle is as follows: succinate dehydrogenase in mitochondria of living cells can reduce exogenous MTT into water-insoluble blue-violet crystalline Formazan (Formazan) and deposit the Formazan in the cells, but dead cells do not have the function. Dimethyl sulfoxide (DMSO) can dissolve formazan in cells, and the light absorption value of the formazan is measured at the wavelength of 570nm by an enzyme-labeling instrument, so that the quantity of living cells can be indirectly reflected.

MCF-7 cells are inoculated in a 96-well plate, after a period of culture, a certain concentration of compound 1 or compound 2 is added into different wells respectively to ensure that the concentration of the compound is 0-12 mu mol/L respectively, and the cells are continuously incubated for 48h and then the cell activity is detected through an MTT experiment. The experimental data are shown in figure 5, after MCF-7 cells are cultured for 48 hours by using different concentrations of compound 1 and compound 2, the cells still show good survival rate, even if the concentration is increased to 12 mu mol/L, the survival rate of the cells is still high, and the azaindole-hemicyanine dye has very good biocompatibility, does not generate toxic or side effect on the cells within a working concentration range, and can be applied to the fields of biology and medicine.

The foregoing is a further description of the invention in connection with specific preferred embodiments thereof and is not intended to limit the invention to the particular forms disclosed. It will be apparent to those skilled in the art that various changes and modifications can be made therein without departing from the scope of the invention as defined in the appended claims.

Claims

1. An azaindole-hemicyanine dye, the dye having a structural formula as shown in formula I:

in the general formula I, the compound has the following structure,

R₁and R₃Each independently is one selected from hydrogen, halogen, alkyl with 1-18 carbons, carboxyalkyl with 1-18 carbons, aryl, arylcarboxylic acid, alkylsulfonate, arylsulfonate, alkylsulfonate, or arylsulfonate;

R₂、R₄and R₅Each independently selected from one of hydrogen, alkyl having 1-18 carbons, carboxyalkyl having 1-18 carbons, hydroxyalkyl having 1-18 carbons, alkylsulfonate having 1-18 carbons, aryl, arylcarboxylic acid, alkylsulfonate, arylsulfonate, alkylsulfonate, or arylsulfonate;

x is selected from one of oxygen, sulfur, selenium and tellurium;

Y^-selected from halogen ion, ClO₄ ^-、BF₄ ^-、CH₃COO^-、CF₃COO^-Or OTs^-One kind of (1).

2. Dye according to claim 1, characterized in thatIs characterized in that: r is₁And R₃Can be independently selected from one of hydrogen, halogen, carboxyalkyl with 1-8 carbons and alkyl sulfonate.

3. Dye according to claim 1, characterized in that R₂、R₄、R₅Can be one selected from hydrogen, aryl, alkyl with 1-8 carbons and alkyl sulfonate with 1-8 carbons independently.

4. Dye according to claim 1, characterized in that X is chosen from one of oxygen, sulphur.

5. Dye according to claim 1, characterized in that Y^-Selected from halogen ions, ClO₄ ^-One kind of (1).

6. A process for the synthesis of azaindole-hemicyanine dyes as claimed in claim 1, characterized in that: the method comprises the following synthetic steps:

(1) in an organic solvent at 60-120 deg.C, containing R₁Substituted J-1 and R₂Reacting the substituted halogenated alkane for 3 to 24 hours, and recrystallizing to obtain the product containing N-R₂Quaternary ammonium salts of substituted side chains J-2; wherein, contains R₁Substituted J-1 and R₂The molar ratio of the substituted halogenated alkanes is 1:1 to 10, most preferably 1:3 to 8;

(2) at 20-80 deg.C, adding S-1 and R₃、R₄、R₅Dissolving substituted S-2 in polar solvent, reacting for 10-24h under catalysis of inorganic base, extracting, concentrating, and purifying to obtain the product containing R ₃、R₄、R₅A substituted intermediate S-3;

7. The method of synthesis according to claim 6, characterized in that: in the step (1), the organic solvent is selected from any one of benzene, toluene, o-dichlorobenzene and DMF; the recrystallization solvent is selected from one or a mixture of more of methanol, ethanol, acetonitrile, ethyl acetate, diethyl ether, acetone and propanol;

in the step (2), the polar solvent is selected from one or a mixture of several of DMSO, DMF, methanol and acetonitrile; the inorganic base is selected from any one of sodium hydroxide, potassium carbonate, cesium carbonate, sodium acetate and sodium ethoxide;

in the step (3), the organic solvent is selected from one or a mixture of several of ethanol, acetic acid, acetic anhydride and DMF; the recrystallization solvent is selected from one or a mixture of more of methanol, ethanol, acetonitrile, water, ethyl acetate, ether, acetone and propanol; the organic base is any one of triethylamine, pyridine and DIPEA.

8. Use of azaindole-hemicyanine dyes as claimed in claim 1 in the biological and medical field.

9. Use according to claim 8, characterized in that: the application comprises the application of azaindole-hemicyanine dye in the aspects of cell imaging, protein labeling, specific recognition of antibody, nucleic acid labeling, DNA sequencing and tumor photodynamic therapy.

10. Use according to claim 8 or 9, characterized in that: when the azaindole-hemicyanine is applied, the excitation wavelength is 600-950nm, and the fluorescence detection wavelength is 650-1000 nm.