阅读说明：本技术 一种七甲川吲哚菁染料及其制备方法和应用 (Heptamethine indocyanine dye and preparation method and application thereof ) 是由 杨靓月 郭英杰 王兴宗 李响 晋腾达 唐煜文 于 2021-06-30 设计创作，主要内容包括：本发明提供了一种七甲川吲哚菁染料及其制备方法和应用,属于精细化工技术领域。本发明提供的七甲川吲哚菁染料,具有式1所示结构,其在近红外区有强的近红外吸收,且具有良好的光稳定性和光热转换效率,其光热转换率、光热稳定性相较于吲哚菁绿明显提高。同时,本发明提供的七甲川吲哚菁染料具有良好的荧光性能。本发明中的七甲川吲哚菁染料属于有机小分子,细胞毒性低且化学位点多,易于进一步设计合成功能性分子；其生物相容性好,在生物体内易于清除,具有明确的生物降解途径,能够应用于生物标记、光学成像、光热成像、光声成像、光动力疗法和光热治疗领域。(The invention provides a heptamethine indocyanine dye and a preparation method and application thereof, belonging to the technical field of fine chemical engineering. The heptamethine indocyanine dye provided by the invention has a structure shown in formula 1, has strong near-infrared absorption in a near-infrared region, has good light stability and light-heat conversion efficiency, and has a light-heat conversion rate and light-heat stability which are obviously improved compared with indocyanine green. Meanwhile, the heptamethine indocyanine dye provided by the invention has good fluorescence property. The heptamethine indocyanine dye belongs to small organic molecules, has low cytotoxicity and multiple chemical sites, and is easy to enterDesigning and synthesizing functional molecules; the nano-particles have good biocompatibility, are easy to remove in organisms, have a definite biodegradation way, and can be applied to the fields of biomarkers, optical imaging, photothermal imaging, photoacoustic imaging, photodynamic therapy and photothermal therapy.)

1. A heptamethine indocyanine dye has a structure shown in formula 1:

in formula 1, n is 0 or 1;

R₁is of the formula R_1-1、R_1-2、R_1-3And R_1-4Any one of:

R₁in the formula, m is 0, 1 or 2;

R₂is of the formula R_2-1Or formula R_2-2：

R₃Is of the formula R_3-1Or formula R_3-2：

When R is₂Is R_2-1When R is₃Is R_3-1(ii) a When R is₂Is R_2-2When R is₃Is R_3-2。

2. A process for the preparation of a heptamethine indocyanine dye as claimed in claim 1, comprising the steps of:

when R is₁Is composed ofThe preparation method of the heptamethine indocyanine dye comprises the following steps:

carrying out condensation reaction on a compound with a structure shown in a formula a and a compound with a structure shown in a formula b to obtain a compound with a structure shown in a formula c;

carrying out nucleophilic substitution reaction on a compound with a structure shown in a formula c and a nucleophilic reagent to obtain a heptamethine indocyanine dye;

the nucleophilic reagent has a structure shown in a formula d-1, a formula d-2 or a formula d-3;

when R is₁Is composed ofWhen the dye is a heptamethine indocyanine dye, the heptamethine indocyanine dye has a structure shown in a formula 1-1:

the preparation method of the heptamethine indocyanine dye with the structure shown in the formula 1-1 comprises the following steps:

the heptamethine indocyanine dye prepared by the method has a structure shown in a formula 1-2;

carrying out BOC protecting group removing reaction on the heptamethine indocyanine dye with the structure shown in the formula 1-2 to obtain the heptamethine indocyanine dye with the structure shown in the formula 1-1.

3. The preparation method according to claim 2, wherein the molar ratio of the compound having the structure shown in the formula a to the compound having the structure shown in the formula b is 1: 2-2.1;

the condensation reaction is carried out in the presence of sodium acetate, the temperature of the condensation reaction is reflux temperature, and the time is 4-6 h.

4. The preparation method according to claim 2, wherein when the nucleophilic reagent has a structure represented by formula d-1 or formula d-3, the temperature of the nucleophilic substitution reaction is 80-90 ℃ and the time is 4-6 h;

and when the nucleophilic reagent has a structure shown as a formula d-2, the temperature of the nucleophilic substitution reaction is room temperature, and the time is 17-24 h.

5. The preparation method according to claim 2, wherein the deprotection reagent used in the BOC protecting group removing reaction is an aqueous solution of trifluoroacetic acid; the reaction time for removing the BOC protecting group is 2-6 h.

6. The method according to claim 2, wherein the method for preparing the compound having the structure represented by the formula a comprises the following steps:

mixing phosphorus oxychloride with N', N-dimethylformamide to obtain Vilsmeier-Haack weak nucleophilic reagent;

mixing the Vilsmeier-Haack weak nucleophilic reagent with cycloalkanone and aniline to perform Vilsmeier-Haack reaction to obtain a compound with a structure shown in a formula a; the cycloalkanone is cyclohexanone or cyclopentanone.

7. The method according to claim 2, wherein the method for preparing the compound having the structure represented by formula b comprises the following steps:

mixing 1,1, 2-trimethyl-1H-benzo [ e ] indole with an electrophilic reagent, and carrying out electrophilic reaction to obtain a compound with a structure shown in a formula b;

the electrophilic reagent is 1, 3-propane sultone or 1, 4-butane sultone.

8. The preparation method according to claim 7, wherein the molar ratio of the 1,1, 2-trimethyl-1H-benzo [ e ] indole to the electrophile is 1 (1.2-1.7);

the temperature of the electrophilic reaction is reflux temperature, and the time is 18-24 h.

9. Use of the heptamethine indocyanine dye of claim 1 or the heptamethine indocyanine dye prepared by the method of any one of claims 2 to 8 in the preparation of reagents or drugs for biological labeling, fluorescence imaging, photothermal imaging and photoacoustic imaging.

10. Use of the heptamethine indocyanine dye of claim 1 or the heptamethine indocyanine dye prepared by the method of any one of claims 2 to 8 in the preparation of a photothermal therapy drug or a photodynamic therapy drug.

Technical Field

The invention relates to the technical field of fine chemical engineering, and particularly relates to a heptamethine indocyanine dye as well as a preparation method and application thereof.

Background

The photothermal material is a material which converts absorbed light energy into kinetic energy of electron or hole resonance through a surface local plasma resonance effect or generates energy through electron transition, and transmits the energy to the surrounding environment through vibration energy scattered by crystal lattices so as to increase the environmental temperature, and is widely researched in the field of photothermal therapy.

The ideal near-infrared photothermal material should have strong near-infrared absorption and good stability, and can effectively convert the absorbed near-infrared light energy into heat energy, i.e. have high photothermal conversion efficiency. The near-infrared photothermal materials are divided into inorganic near-infrared photothermal materials and organic near-infrared photothermal materials, wherein the inorganic near-infrared photothermal materials mainly comprise nano-gold materials, nano-carbon materials, nano-palladium materials, copper sulfide nano-materials and the like, although the inorganic materials show good photothermal conversion characteristics, most of the inorganic materials have poor biocompatibility, are not biodegradable, can remain in the body for a long time and have potential toxicity, and thus the wide application of the inorganic near-infrared photothermal materials is limited. Therefore, the organic near-infrared photothermal material is gradually favored by researchers, and compared with the inorganic near-infrared photothermal material, the organic near-infrared photothermal material not only has good photothermal conversion efficiency, but also has the advantages of good biocompatibility, low toxic and side effects and the like.

The cyanine dye is one of the currently commonly used organic near-infrared photothermal materials, taking indocyanine green (ICG) as an example, it has the advantages of high molar absorption coefficient, high fluorescence quantum yield, capability of photothermal conversion, low cytotoxicity and the like, and is the only near-infrared fluorescent dye approved by FDA and applicable to clinical diagnosis. However, indocyanine green (ICG) has poor light stability and low photothermal conversion efficiency, which limits its wide application.

Disclosure of Invention

In view of the above, the present invention aims to provide a heptamethine indocyanine dye, a preparation method and an application thereof, wherein the heptamethine indocyanine dye provided by the present invention has strong absorption in a near infrared region, and has good light stability and photo-thermal conversion efficiency.

In order to achieve the above object, the present invention provides the following technical solutions:

the invention provides a heptamethine indocyanine dye which has a structure shown in a formula 1:

in formula 1, n is 0 or 1;

R₁is of the formula R_1-1、R_1-2、R_1-3And R_1-4Any one of:

R₁in the formula, m is 0, 1 or 2;

R₂is of the formula R_2-1Or formula R_2-2：

R₃Is of the formula R_3-1Or formula R_3-2：

When R is₂Is R_2-1When R is₃Is R_3-1(ii) a When R is₂Is R_2-2When R is₃Is R_3-2。

The invention provides a preparation method of the heptamethine indocyanine dye, which comprises the following steps:

when R is₁Is composed ofThe preparation method of the heptamethine indocyanine dye comprises the following steps:

carrying out condensation reaction on a compound with a structure shown in a formula a and a compound with a structure shown in a formula b to obtain a compound with a structure shown in a formula c;

carrying out nucleophilic substitution reaction on a compound with a structure shown in a formula c and a nucleophilic reagent to obtain a heptamethine indocyanine dye;

the nucleophilic reagent has a structure shown in a formula d-1, a formula d-2 or a formula d-3;

when R is₁Is composed ofWhen the dye is a heptamethine indocyanine dye, the heptamethine indocyanine dye has a structure shown in a formula 1-1:

the preparation method of the heptamethine indocyanine dye with the structure shown in the formula 1-1 comprises the following steps:

the heptamethine indocyanine dye prepared by the method has a structure shown in a formula 1-2;

carrying out BOC protecting group removing reaction on the heptamethine indocyanine dye with the structure shown in the formula 1-2 to obtain the heptamethine indocyanine dye with the structure shown in the formula 1-1.

Preferably, the molar ratio of the compound with the structure shown in the formula a to the compound with the structure shown in the formula b is 1: 2-2.1;

the condensation reaction is carried out in the presence of sodium acetate, the temperature of the condensation reaction is reflux temperature, and the time is 4-6 h.

Preferably, when the nucleophilic reagent has a structure shown as a formula d-1 or a formula d-3, the temperature of the nucleophilic substitution reaction is 80-90 ℃, and the time is 4-6 h;

and when the nucleophilic reagent has a structure shown as a formula d-2, the temperature of the nucleophilic substitution reaction is room temperature, and the time is 17-24 h.

Preferably, the deprotection reagent used in the reaction for removing the BOC protecting group in the second step is aqueous solution of trifluoroacetic acid; the reaction time for removing the BOC protecting group is 2-6 h.

Preferably, the preparation method of the compound with the structure shown in the formula a comprises the following steps:

mixing phosphorus oxychloride with N', N-dimethylformamide to obtain Vilsmeier-Haack weak nucleophilic reagent;

mixing the Vilsmeier-Haack weak nucleophilic reagent with cycloalkanone and aniline to perform Vilsmeier-Haack reaction to obtain a compound with a structure shown in a formula a; the cycloalkanone is cyclohexanone or cyclopentanone.

Preferably, the preparation method of the compound with the structure shown in the formula b comprises the following steps:

mixing 1,1, 2-trimethyl-1H-benzo [ e ] indole with an electrophilic reagent, and carrying out electrophilic reaction to obtain a compound with a structure shown in a formula b;

the electrophilic reagent is 1, 3-propane sultone or 1, 4-butane sultone.

Preferably, the molar ratio of the 1,1, 2-trimethyl-1H-benzo [ e ] indole to the electrophilic reagent is 1 (1.2-1.7);

the temperature of the electrophilic reaction is reflux temperature, and the time is 18-24 h.

The invention provides application of the heptamethine indocyanine dye in preparation of reagents or medicines for biological marking, fluorescence imaging, photothermal imaging and photoacoustic imaging.

The invention provides application of the heptamethine indocyanine dye in preparing photo-thermal treatment medicines or photodynamic treatment medicines.

The invention provides a heptamethine indocyanine dye which has a structure shown in a formula 1. The heptamethine indocyanine dye provided by the invention has strong near-infrared absorption in a near-infrared region, good light stability and photo-thermal conversion efficiency, and the photo-thermal conversion rate and the photo-thermal stability of the heptamethine indocyanine dye are obviously improved compared with indocyanine green. The invention introduces a ring rigid structure (such as a six-membered ring or a five-membered ring) containing unsaturated bonds into the conjugated chain of the indocyanine dye, so that the polymethine chain is partially rigidized and the steric hindrance of the polymethine chain is increased; meanwhile, the substitution of different heteroatoms for the polymethine chain can influence the electron distribution state of a conjugated system, and further influence the optical stability and the photo-thermal conversion efficiency of molecules. Meanwhile, the heptamethine indocyanine dye provided by the invention has good fluorescence property. The heptamethine indocyanine dye belongs to small organic molecules, has low cytotoxicity and many chemical sites, and is easy to further design and synthesize functional molecules; the nano-particles have good biocompatibility, are easy to remove in organisms, have a definite biodegradation way, and can be applied to the fields of biomarkers, optical imaging, photothermal imaging, photoacoustic imaging, photodynamic therapy and photothermal therapy.

The invention provides a preparation method of the heptamethine indocyanine dye, which is simple to operate and easy to realize industrial mass production.

Drawings

FIG. 1 is a nuclear magnetic resonance hydrogen spectrum of a heptamethine indocyanine dye Cy4 according to the present invention;

FIG. 2 is a UV spectrum of Cy2, Cy3, and Cy 4;

FIG. 3 is a fluorescence spectrum of Cy2, Cy3 and Cy 4;

FIG. 4 shows Cy2, Cy3, Cy4 passing through 808nm 2.5W/cm²A temperature rise curve graph of laser irradiation for 10 minutes;

fig. 5 shows the results of the photothermal conversion test of Cy 2;

fig. 6 shows the results of the photothermal conversion test of Cy 3;

fig. 7 shows the results of the photothermal conversion test of Cy 4;

FIG. 8 shows UV absorption spectra before and after exposure to Cy 2;

FIG. 9 shows UV absorption spectra before and after exposure to Cy 3;

fig. 10 shows ultraviolet absorption spectra before and after Cy4 irradiation.

Detailed Description

The invention provides a heptamethine indocyanine dye which has a structure shown in a formula 1:

in formula 1, n is 0 or 1;

R₁is of the formula R_1-1、R_1-2、R_1-3And R_1-4Any one of:

R₁in the formula, m is 0, 1 or 2;

R₂is of the formula R_2-1Or formula R_2-2：

R₃Is of the formula R_3-1Or formula R_3-2：

When R is₂Is R_2-1When R is₃Is R_3-1(ii) a When R is₂Is R_2-2When R is₃Is R_3-2。

In the present invention, in the case of the present invention,indicates the attachment site.

In the invention, the structure of the heptamethine indocyanine dye is preferably as shown in a formula 2-9, and is specifically shown in a table 1.

TABLE 1 specific structural formula of heptamethine indocyanine dyes

The heptamethine indocyanine dye provided by the invention has strong near-infrared absorption in a near-infrared region, and the wavelength of the near-infrared region is preferably 620-850 nm. The heptamethine indocyanine dye provided by the invention has good light stability and photo-thermal conversion efficiency, and the photo-thermal conversion rate and the photo-thermal stability of the heptamethine indocyanine dye are obviously improved compared with indocyanine green. Meanwhile, the heptamethine indocyanine dye provided by the invention has good fluorescence property. The heptamethine indocyanine dye belongs to small organic molecules, has low cytotoxicity and many chemical sites, and is easy to further design and synthesize functional molecules; the nano-particles have good biocompatibility, are easy to remove in organisms, have a definite biodegradation way, and can be applied to the fields of biomarkers, optical imaging, photothermal imaging, photoacoustic imaging, photodynamic therapy and photothermal therapy.

The invention provides a preparation method of the heptamethine indocyanine dye, which comprises the following steps:

when R is₁Is composed ofThe preparation method of the heptamethine indocyanine dye comprises the following steps:

reacting a compound with a structure shown in a formula a with a compound with a structure shown in a formula b to obtain a compound with a structure shown in a formula c;

carrying out nucleophilic substitution reaction on a compound with a structure shown in a formula c and a nucleophilic reagent to obtain a heptamethine indocyanine dye;

the nucleophilic reagent has a structure shown in a formula d-1, a formula d-2 or a formula d-3;

in the present invention, the preparation method of the compound having the structure represented by formula a preferably comprises the following steps:

mixing phosphorus oxychloride with N', N-dimethylformamide to obtain Vilsmeier-Haack weak nucleophilic reagent;

mixing the Vilsmeier-Haack weak nucleophilic reagent with cycloalkanone and aniline, and carrying out Vilsmeier-Haack reaction to obtain a compound with a structure shown in a formula a; the cycloalkanone is cyclohexanone or cyclopentanone.

Phosphorus oxychloride and N' N-dimethylformamide are mixed to obtain the Vilsmeier-Haack weak nucleophilic reagent. In the present invention, the mixing is preferably performed under nitrogen protection; the molar ratio of the phosphorus oxychloride to the N' N-dimethylformamide is preferably 2.3: 3.2. In the invention, the mixing temperature is preferably-2-5 ℃, and more preferably 0-2 ℃; the mixing time is preferably 0.5-2 h, and more preferably 1-1.5 h. In the present invention, the phosphorus oxychloride and N' N-dimethylformamide are capable of forming a Vilsmeier-Haack weak nucleophile.

In the invention, the Vilsmeier-Haack weak nucleophilic reagent is mixed with cycloalkanone and aniline to carry out Vilsmeier-Haack reaction, thus obtaining a compound with a structure shown in a formula a; the cycloalkanone is cyclohexanone or cyclopentanone. In the invention, the molar ratio of the cycloalkanone, the phosphorus oxychloride and the aniline is preferably 1: 2.3: 2-2.2. In the present invention, the aniline is preferably added in the form of an ethanol solution of aniline, and the molar ratio of aniline to ethanol is preferably 1: 1.

In the present invention, the mixing is preferably performed in the following manner: mixing the Vilsmeier-Haack weak nucleophilic reagent with the cyclic alkanone, heating and refluxing, cooling, and adding aniline. In the invention, the temperature of the heating reflux is preferably 153-165 ℃, and more preferably 155-160 ℃; the time is preferably 1 h.

In the invention, the temperature of the Vilsmeier-Haack reaction is preferably room temperature, and the time is preferably 0.5-2 h, and more preferably 1-1.5 h.

After the Vilsmeier-Haack reaction, the present invention preferably performs a post-treatment of the obtained Vilsmeier-Haack reaction product, which preferably comprises the steps of:

and (3) recrystallizing, washing, filtering and drying the obtained Vilsmeier-Haack reaction product in sequence to obtain a compound solid with the structure shown in the formula a. In the present invention, the recrystallization is preferably performed under a cold water bath condition, and the solvent used for the recrystallization is preferably concentrated hydrochloric acid. In the invention, the temperature of the cold water is preferably 0-4 ℃, and the volume ratio of the water in the concentrated hydrochloric acid to the hydrochloric acid is preferably 8-10: 1.

The present invention does not require any particular manner of washing, filtering and drying, and the above-described operations, which are well known to those skilled in the art, may be used.

In the present invention, the preparation method of the compound having the structure represented by formula b preferably comprises the following steps:

mixing the 1,1, 2-trimethyl-1H-benzo [ e ] indole with an electrophilic reagent, and carrying out electrophilic reaction to obtain the compound with the structure shown in the formula b.

In the present invention, the electrophile is preferably 1, 3-propane sultone or 1, 4-butane sultone.

In the present invention, the electrophilic reaction is carried out in an organic solvent, which is preferably toluene.

In the invention, the molar ratio of the 1,1, 2-trimethyl-1H-benzo [ e ] indole to the electrophilic reagent is preferably 1:1 (1.2-1.7), and more preferably 1: 1.5. In the invention, the temperature of the electrophilic reaction is preferably the reflux temperature, and the time is preferably 18-24 h, and more preferably 20-22 h.

After the electrophilic reaction is completed, the present invention preferably performs a post-treatment on the obtained electrophilic reaction solution, and the post-treatment preferably comprises the following steps:

and (3) sequentially carrying out solid-liquid separation on the electrophilic reaction liquid, and recrystallizing the obtained solid to obtain the compound solid with the structure shown in the formula b.

The present invention has no special requirement on the solid-liquid separation mode, and the solid-liquid separation mode known to those skilled in the art can be used, such as filtration. In the invention, the solvent used for recrystallization is preferably a mixed solution of methanol and diethyl ether, and the volume ratio of the methanol to the diethyl ether in the mixed solution is preferably 1 to (1-2).

In the present invention, a compound having a structure represented by formula a is reacted with a compound having a structure represented by formula b to obtain a compound having a structure represented by formula c.

In the invention, the molar ratio of the compound with the structure shown in the formula a to the compound with the structure shown in the formula b is preferably 1: 2-2.1, and more preferably 1: 2.

In the present invention, the reaction is preferably carried out in the presence of anhydrous sodium acetate, and the molar ratio of the compound having the structure represented by formula a to the anhydrous sodium acetate is preferably 1: 2.3. In the present invention, the anhydrous sodium acetate provides an alkaline condition for the reaction.

In the present invention, the organic solvent used for the reaction is preferably absolute ethanol.

In the present invention, after the reaction is completed, the present invention preferably subjects the obtained reaction product to a post-treatment, which preferably comprises the steps of:

and (3) sequentially removing the solvent and recrystallizing the obtained reaction product to obtain a compound solid with the structure shown in the formula c.

The invention does not require any particular solvent removal means, and can be achieved by solvent removal means known to those skilled in the art. In the invention, the solvent used for recrystallization is preferably a mixed solution of methanol and diethyl ether, and the volume ratio of the methanol to the diethyl ether in the mixed solution is preferably 1 to (1-2).

In the invention, the preparation process of the compound having the structure shown in formula c is shown in formula A:

after the compound with the structure of the formula c is obtained, the compound with the structure of the formula c and a nucleophilic reagent are subjected to nucleophilic substitution reaction to obtain a heptamethine indocyanine dye;

the nucleophilic reagent has a structure shown in a formula d-1, a formula d-2 or a formula d-3;

in the invention, the nucleophilic reagent attacks the conjugated chain meso-position carbon atom of the compound with the structure shown in the formula c, and the compound with the structure shown in the formula c meso-position chlorine is substituted through nucleophilic substitution reaction to prepare the heptamethine indocyanine dye.

In the present invention, the solvent used in the nucleophilic substitution reaction is preferably one or more of methanol, DMSO, DMF, and acetone. In the present invention, the nucleophilic substitution reaction is preferably at N₂Under protection.

In the invention, when the nucleophilic reagent has a structure shown as a formula d-1 or a formula d-3, the temperature of the nucleophilic substitution reaction is preferably 80-90 ℃, and the time is preferably 4-6 h, and more preferably 5 h.

In the present invention, when the nucleophile has a structure represented by formula d-2, the nucleophilic substitution reaction is preferably performed under basic conditions, and in the present invention, the base used to provide basic conditions is preferably sodium hydride. In the invention, when the nucleophilic reagent has a structure shown as a formula d-2, the temperature of the nucleophilic substitution reaction is preferably room temperature, and the time is preferably 17-24 h, and more preferably 20-22 h.

In the present invention, the reaction process of the nucleophilic substitution reaction is shown as formula B:

in the invention, after the nucleophilic substitution reaction is finished, the invention preferably performs C18 reverse column chromatography separation on the obtained nucleophilic substitution reaction liquid to obtain the pure product of the heptamethine indocyanine dye. In the invention, the packing material for the C18 reverse phase column chromatography is preferably silica gel, and the mobile phase is preferably petroleum ether and ethyl acetate.

When R is₁Is composed ofWhen the dye is a heptamethine indocyanine dye, the heptamethine indocyanine dye has a structure shown in a formula 1-1:

the preparation method of the heptamethine indocyanine dye with the structure shown in the formula 1-1 comprises the following steps:

the heptamethine indocyanine dye prepared by the method has a structure shown in a formula 1-2;

carrying out BOC protecting group removing reaction on the heptamethine indocyanine dye with the structure shown in the formula 1-2 to obtain the heptamethine indocyanine dye with the structure shown in the formula 1-1.

In the invention, the deprotection reagent used in the BOC protecting group removing reaction is preferably trifluoroacetic acid aqueous solution, and the volume ratio of trifluoroacetic acid to water in the trifluoroacetic acid aqueous solution is preferably 1-2: 1. In the invention, the temperature of the BOC protecting group removing reaction is preferably room temperature, and the time is preferably 2-6 h.

In the invention, the reaction process of the BOC protecting group removing reaction is shown as a formula C.

The invention provides application of the heptamethine indocyanine dye in preparation of reagents or medicines for biological marking, fluorescence imaging, photothermal imaging and photoacoustic imaging. .

The invention provides application of the heptamethine indocyanine dye in preparing photo-thermal treatment medicines or photodynamic treatment medicines.

The heptamethine indocyanine dyes provided by the present invention, the preparation methods and applications thereof will be described in detail with reference to the following examples, but they should not be construed as limiting the scope of the present invention.

Example 1

(1) Preparation of Compound (Compound 1) having formula a

1mL POCl was added to 1.2mL anhydrous DMF at 2 deg.C₃Reacting for 30min, adding 0.5mL of cyclohexanone, and heating and refluxing for 1 h; then cooling to 20 ℃, and dropwise adding 2.9mL of aniline/ethanol mixture, wherein the molar ratio of aniline to ethanol in the mixture is 1: 1; after stirring for 1.5 hours, pour into concentrated HCl, recrystallize in an ice-water bath, filter, wash, and dry in vacuo to give 1.044g of Compound 1, 61% yield.

LCMS (ESI +) calculation of [ M + H ]]⁺323.1, 323.1 was detected.

The synthetic route is shown as formula D:

(2) preparation of Compound (Compound 2) having formula b

Toluene, 1.3g of 1,1, 2-trimethyl-1H-benzo [ e ] indole and 1.1mL of 1, 3-propanesultone were heated under reflux for 18 hours; cooling to room temperature, the resulting blue crystals were filtered, washed with ether, and the product recrystallized from methanol and ether and dried in vacuo to give 1.81g of a grey solid as compound 2 in 88% yield.

LCMS (ESI +) calculation of [ M + H ]]⁺332.1, 332.1 was measured.

The synthetic route is shown as formula E:

(3) synthesis of Compound (Compound 3) having the Structure represented by formula c

Under a nitrogen atmosphere, the prepared compound 1(360mg), compound 2(662mg) and anhydrous sodium acetate were heated under reflux in an anhydrous ethanol solution for 4 hours, and the progress of the reaction was monitored by Thin Layer Chromatography (TLC); the solvent ethanol was removed under reduced pressure and the resulting solid was washed with diethyl ether and separated by column chromatography to give the crude product which was purified by recrystallisation from methanol and diethyl ether and finally filtered and dried in vacuo to give compound 3(759mg) in 95% yield.

The synthetic route is shown as formula F:

(4) synthesis of heptamethine indocyanine dye Cy1

Weighing 800mg of 4-hydroxybenzylamine and 1.56g of di-tert-butyl dicarbonate, adding sodium bicarbonate to dissolve in methanol, heating and refluxing overnight, and monitoring the reaction process by TLC; after the reaction, the mixture was cooled to room temperature, the solvent methanol was removed under reduced pressure, water and ethyl acetate were added for extraction, and compound 4(1.14g) was isolated by post-treatment and column chromatography techniques in 79% yield.

LCMS (ESI-) [ M-H ] is calculated]^-222.1, 222.1 was measured.

In N₂836mg of Compound 4 are weighed out in anhydrous DMF under atmosphere and directed upwardsTo the solution was added 150mg of sodium hydride, and the mixture was stirred at room temperature for 1 hour. Adding 200mg of compound 3 to the solution, and reacting at room temperature for 20 hours; the solvent was evaporated to dryness and ethyl acetate was added, stirred, filtered and dried under vacuum to give the crude product, which was then purified by column chromatography to give the heptamethine indocyanine dye Cy 1.

The synthetic route is shown as formula G:

example 2

200mg of heptamethine indocyanine dye Cy1 was weighed, 10mL of a mixture of trifluoroacetic acid (TFA) and water was added, stirred at room temperature for 4 hours, the solvent was evaporated to dryness, and ethyl acetate was added thereto, stirred, filtered, and dried under vacuum to give a crude product, which was then separated and purified by column chromatography to give heptamethine indocyanine dye Cy 2.

LCMS (ESI +) calculation of [ M + H ]]⁺886.3, respectively; 886.3 was measured.

The synthetic route is shown as formula H:

example 3

Weighing a compound, namely the heptamethine indocyanine dye compound 3(200mg) and p-phenylenediamine (270mg), dissolving the mixture in anhydrous DMF, heating the mixture for reaction for 4-6 hours, monitoring the reaction progress by TLC, cooling to room temperature, adding diethyl ether, stirring, filtering, drying in vacuum to obtain a crude product, and separating and purifying by C18 reverse column chromatography to obtain the heptamethine indocyanine dye Cy 3.

LCMS (ESI +): calculating C₅₀H₅₄N₄O₆S₂，m/z 870.3，[M+2H]2+ 435.5; 435.2 was measured.

The synthetic route is shown as formula J:

example 4

200mg of the compound heptamethine indocyanine dye compound 3, 215.5mg of anhydrous piperazine were weighed and dissolved in anhydrous DMF, the mixture was heated to react for 5 hours, TLC monitored the progress of the reaction, then cooled to room temperature and added with ether, stirred, filtered, vacuum dried to obtain a crude product, which was recrystallized with methanol and ether to purify the crude product, and finally filtered and vacuum dried to obtain heptamethine indocyanine dye Cy4(160mg) with a yield of 75%.

LCMS (ESI +): calculate [ M]⁺849.1, respectively; 849.1 was measured.

The synthetic route is shown as formula K:

the hydrogen nuclear magnetic resonance spectrum of the heptamethine indocyanine dye Cy4 is shown in FIG. 1.

The hydrogen spectrum data are:

¹HNMR(400MHz，DMSO-d₆)δ8.18(d，J＝8.8Hz，2H)，8.02(apps，2H)，8.00(apps，2H)，7.80(d，J＝13.2Hz，2H)，7.73(d，J＝8.8Hz，2H)，7.81-7.58(m，2H)，7.45-7.41(m，2H)，6.17(d，J＝13.2Hz，2H)，4.36-4.32(m，4H)，3.73(m，4H)，3.17-3.16(m，4H)，2.64-2.56(comp，8H)，2.09-2.02(m，4H)，1.94(s，12H)，1.79-1.73(m，2H).

performance testing

Ultraviolet absorption spectrum

The DMSO solvents were used to prepare Cy2, Cy3, and Cy4 at 10 concentrations^-2mol·L^-1Diluting the solution to be detected to 10 deg.C in methanol^-5mol·L^-1The ultraviolet absorption spectrum was measured, and the results are shown in FIG. 2. As can be seen from fig. 2, the maximum absorption wavelength of Cy2 in methanol is 808nm, the maximum absorption wavelength of Cy3 in methanol is 725nm, and the maximum absorption wavelength of Cy4 in methanol is 808nm, which have strong absorption in the near infrared region.

(II) fluorescent Property

The DMSO solvents were used to prepare Cy2, Cy3, and Cy4 at 10 concentrations^-2mol·L^-1Diluting the solution to be detected to 10 deg.C in methanol^-5mol·L^-1The fluorescence properties were measured, and the results are shown in FIG. 3. As can be seen from fig. 3, the maximum emission wavelength of Cy2 in methanol is 835nm, the maximum emission wavelength of Cy3 in methanol is 834nm, the maximum emission wavelength of Cy4 in methanol is 830nm, the fluorescence intensity of Cy2 and Cy4 is higher, and excellent fluorescence performance is shown.

(III) photothermal Effect

The compounds Cy2, Cy3 and Cy4 were mixed in an aqueous solution of 0.2mmol/L at 880nm and 2.5W/cm²The laser was irradiated for 10 minutes and the change in temperature of the solution system during irradiation was recorded. The results are shown in FIG. 4. As can be seen from FIG. 4, at 880nm, 2.5W/em²Under the laser irradiation, after Cy4 is irradiated by near infrared light for 10 minutes, the temperature is increased by 27.8 ℃; the temperature of Cy2 increased by 21.7 ℃; the temperature of Cy3 increased by 12.7 ℃; the heptamethine indocyanine dyes Cy2, Cy3 and Cy4 provided by the invention have good photo-thermal conversion effect, wherein the Cy4 has higher photo-thermal conversion efficiency and the most obvious temperature rise effect.

(IV) photothermal conversion stability

Firstly, 808nm, 1W/cm are used²After continuously irradiating the compounds Cy2, Cy3 and Cy410 with the laser light, the compound was naturally cooled, and then the temperature was raised by the same laser irradiation for 5 minutes to naturally cool for 5 minutes as one cycle period, and 5 cycle periods were tested. The results of the Cy2 test are shown in FIG. 5, the Cy3 test is shown in FIG. 6, and the Cy4 test is shown in FIG. 7. As can be seen from fig. 5 to 7, the temperature rise degree of the solution system was nearly uniform for each repetition of Cy3 and Cy4, indicating that Cy3 and Cy4 have excellent photothermal conversion stability.

(V) light stability

The compounds Cy2, Cy3 and Cy4 were mixed at 880nm, 1W/cm²The laser irradiation was continued for 10min, and absorption spectra of compounds Cy2, Cy3, and Cy4 before and after the irradiation with light were measured. The results of the Cy2 test are shown in FIG. 8, the Cy3 test is shown in FIG. 9, and the Cy4 test is shown in FIG. 10.

As can be seen from FIGS. 8 to 10, after 10 minutes of irradiation, the absorption intensities of the compounds Cy2, Cy3 and Cy4 are all reduced, but the maximum absorption wavelength and the peak shape are not changed basically, which shows that the compounds have better light stability.

The test shows that the heptamethine indocyanine dye provided by the invention has strong absorption in a near infrared region, and has good light stability and photo-thermal conversion efficiency.

The foregoing is only a preferred embodiment of the present invention, and it should be noted that, for those skilled in the art, various modifications and decorations can be made without departing from the principle of the present invention, and these modifications and decorations should also be regarded as the protection scope of the present invention.