CN111320732A - Amphiphilic block copolymer with near-infrared thermal responsiveness and preparation and application thereof - Google Patents

Abstract

The invention relates to an amphiphilic block copolymer with near-infrared photothermal responsiveness and its preparation and application. The present invention prepares the amphiphilic block copolymer PBnMA-b-P (BAPMA-co-PEGMA) through RAFT polymerization, and then reacts with the ketoacid cyanine dye NIR 800 with near-infrared photothermal response to obtain PBnMA-b- P(APMA‑co‑PEGMA)@NIR800. The self-assembly of amphiphilic block copolymers was carried out in the organic solvent/water mixed solvent system, respectively. The self-assembly results show that the present invention successfully obtains an amphiphilic block copolymer micelle with near-infrared photothermal responsiveness.

Description

Amphiphilic block copolymers with near-infrared photothermal responsiveness and their preparation and applications

technical field

The invention relates to the field of polymer synthesis, in particular to an amphiphilic block copolymer with near-infrared photothermal responsiveness and its preparation and application.

Background technique

Photothermal responsive materials refer to a class of materials that absorb light and convert energy into heat to heat up when illuminated by a light source of a certain wavelength. Since near-infrared light can penetrate deep biological tissues and has little effect on normal cells, nanomaterials with near-infrared photothermal response are often used in photothermal therapy of some human tissues. Inorganic near-infrared photothermal responsive materials have high photothermal conversion efficiency, but they are not an ideal class of photothermal reagents because they are difficult to exclude in vivo and will remain for a long time.

At present, more and more people are devoted to the development of organic NIR photothermal responsive materials. For example, CN103002921A discloses a functional cross-linked nanostructure for combined optical imaging and therapy, wherein the optical substance includes a cross-linked block copolymer, a linker and a therapeutic agent, and each block copolymer contains hydrophilic and hydrophobic blocks. The linker contains an optically active moiety, and the optical substance forms a supramolecular structure in an aqueous solution. CN103254371A discloses a method for synthesizing an amphiphilic block polymer with near-infrared fluorescence properties. The RAFT synthesis method is used to synthesize PNIPAM-b-PVDHBI, the side chain of which contains 2-benzimidazolyl-β-naphthalene chromophore group. It is necessary to develop more types of organic NIR photothermal responsive materials.

SUMMARY OF THE INVENTION

In order to solve the above technical problems, the purpose of the present invention is to provide an amphiphilic block copolymer with near-infrared photothermal responsiveness and its preparation and application.

An amphiphilic block copolymer (PBnMA-b-P(APMA-co-PEGMA)@NIR800) with near-infrared photothermal responsiveness of the present invention is characterized in that it comprises the structure shown in formula (1):

Wherein, x=5-30; y=5-30; z=5-30; m=5-21.

The present invention also provides a preparation method of the amphiphilic block copolymer represented by formula (1), comprising the following steps:

(1) In a protective atmosphere, benzyl methacrylate BnMA was reacted in an organic solvent at 65-85 °C under the action of RAFT regulator α-dithionaphthoic acid isobutyronitrile ester CPDN and thermal initiator , the polymer PBnMA is obtained after the reaction is complete; its structural formula is as follows:

(2) In a protective atmosphere, using the polymer PBnMA as a macromolecular regulator, the tert-butyl carbamate group-protected methacrylamide BAPMA and methacrylate polyethylene glycol monomethyl ether ester PEGMA were thermally induced Under the action of the agent, the reaction is carried out in an organic solvent at 65-85 ° C, and the block polymer PBnMA-b-P (BAPMA-co-PEGMA) is obtained after the reaction is complete; its structural formula is as follows:

(3) removing the protective group tert-butyl carbamate (BOC) on BAPMA in the block polymer PBnMA-b-P (BAPMA-co-PEGMA) to expose the amino group in the methacrylamide to obtain PBnMA-b-P (APMA-co-PEGMA); its structural formula is as follows:

(4) The PBnMA-b-P (APMA-co-PEGMA) and the ketone acid cyanine dye NIR 800 were reacted in an organic solvent at a reaction temperature of 20-25 °C, and the amphiphilic block copolymer PBnMA-b-P was obtained after the reaction was completed. (APMA-co-PEGMA)@NIR800; wherein, the structural formula of the ketoacid cyanine dye NIR 800 is as follows:

Further, in step (1), the molar ratio of BnMA, CPDN and thermal initiator is 5-30:1:0.3.

In step (1), the molecular weight of the polymer PBnMA is 2400-3600 g/mol. The molecular weight distribution index M _w / _{Mn is} less than 1.13. Polymers with different molecular weights can be designed and synthesized by adjusting the ratio of monomer and RAFT reagent CPDN, and the molecular weight distribution of polymers is very narrow.

Further, in step (1), the reaction time is 20h.

Further, in step (2), the molar ratio of BAPMA, PEGMA, PBnMA and thermal initiator is 10:5-10:1:0.3.

In step (2), a block polymer PBnMA-b-P (BAPMA-co-PEGMA) includes the lipophilic segment PBnMA, and the P in the polymer (BAPMA-co-PEGMA) is random of PEGMA and BAPMA. The copolymer, in which PEGMA is a hydrophilic segment, and BAPMA is a functional segment, when the BOC protecting group is removed, the amino group is exposed, and the amino group is used to react with the carboxyl group in the ketone acid cyanine dye NIR 800. Block polymers covalently link groups that are responsive to near-infrared light.

Further, in step (2), the reaction time is 24h. The molecular weight of the block polymer PBnMA-b-P (BAPMA-co-PEGMA) is 5600-9900 g/mol. Block copolymers of different molecular weights can be synthesized by adjusting the ratio of comonomers BAPMA and PEGMA, and using PBnMA of different molecular weights, and the molecular weight distribution of the polymers is mostly narrow. The macromolecular RAFT synthesized in step (1) described The reagent PBnMA has a high degree of terminal functionalization.

Further, in steps (1) and (2), the thermal initiator is azobisisobutyronitrile (AIBN).

Further, in step (3), the method for removing tert-butyl carbamate group comprises the following steps:

The block polymer PBnMA-b-P (BAPMA-co-PEGMA) is subjected to an acid hydrolysis reaction under the action of trifluoroacetic acid (TFA) in an organic solvent, and the reaction temperature is 20-25°C.

Further, in step (3), the reaction time is 24h.

Further, in step (4), the molar ratio of PBnMA-b-P (APMA-co-PEGMA) and NIR 800 is 0.1-0.5:1.

Further, in step (4), the reaction time is 24h.

In step (4), the preparation route of ketoacid cyanine dye NIR 800 is as follows:

In step (4), since both ends of the ketocyanine dye NIR 800 contain carboxyl groups, when it reacts with PBnMA-b-P (APMA-co-PEGMA), one of the carboxyl groups will interact with one of the carboxyl groups on APMA. In the case of amino reaction, in addition, two carboxyl groups react with amino groups on two APMAs, respectively. Since in the same polymer chain, the two closest APMAs are separated by a longer chain PEGMA, so this The APMAs of the two reactions are more likely to be located on different polymer chains, resulting in cross-linking (ie intermolecular cross-linking) between some APMAs on different polymer chains relying on NIR 800.

The present invention also provides an amphiphilic block copolymer micelle with near-infrared photothermal responsiveness, comprising the amphiphilic block copolymer represented by formula (1), the amphiphilic block copolymer micelle It includes a lipophilic inner core and a hydrophilic outer shell wrapped around the inner core. The inner core includes PBnMA, the outer shell includes P(APMA-co-PEGMA), and the outer shell is covalently linked with a near-infrared photothermal responsive ketone acid. Cyanine dyes.

Further, the particle size of the amphiphilic block copolymer micelles is 200-600 nm. Under the condition that the length of the lipophilic segment PBnMA in the amphiphilic block copolymer is kept unchanged, the chain length of the hydrophilic segment is extended, and the obtained micelle size will also increase accordingly. At the same time, changing the ratio of lipophilic segment and hydrophilic segment in the amphiphilic block copolymer also changes the micelle size.

Further, the amphiphilic block copolymer micelles are obtained by self-assembly of the amphiphilic block copolymer represented by formula (1) in a co-solvent of an organic solvent and water.

Specifically, when preparing micelles, the amphiphilic block copolymer represented by formula (1) is dissolved in an organic solvent, and the obtained organic solution is slowly added dropwise to water, or water is slowly added dropwise to the obtained organic solution. , the volume ratio of water and organic solvent is greater than 10:1. Under the condition of excessive volume of water, the amphiphilic block copolymer represented by formula (1) relies on the hydrophilic and hydrophobic interaction in the co-solvent of organic solvent and water. self-assembly by force to obtain amphiphilic block copolymer micelles with a hydrophilic shell and a hydrophobic core.

Further, when preparing the amphiphilic block copolymer micelles, the steps of dialysis and centrifugation to remove excess NIR800 are also included after the dropwise addition.

The invention also discloses the application of the above-mentioned amphiphilic block copolymer micelles with near-infrared photothermal responsiveness in preparing near-infrared light-responsive materials.

Further, the near-infrared light-responsive material is a near-infrared light-responsive photothermal therapy preparation.

Since the amphiphilic block copolymer micelles of the present invention contain ketone acid cyanine dyes, they can absorb near-infrared light and convert light energy into heat, so they are suitable for preparing near-infrared light-responsive materials for non-medical purposes, and even It can be used to prepare photothermal treatment preparations for medical purposes, and utilize the photothermal effect of the preparations to exert its therapeutic function.

By means of the above scheme, the present invention has at least the following advantages:

The present invention prepares the amphiphilic block copolymer PBnMA-b-P (BAPMA-co-PEGMA) with PEGMA as the hydrophilic monomer by RAFT polymerization, and uses the ketone acid cyanine dye NIR 800 to post-modify the block copolymer, The amphiphilic block copolymer PBnMA-b-P(APMA-co-PEGMA)@NIR800 with near-infrared photothermal responsiveness was obtained.

The PBnMA-b-P(APMA-co-PEGMA)@NIR800 of the present invention can be self-assembled in a mixed solvent system of organic solvent/water, thereby obtaining an amphiphilic block copolymer micelle with near-infrared photothermal responsiveness , the micelles can be used to prepare near-infrared light-responsive materials.

The above description is only an overview of the technical solution of the present invention. In order to understand the technical means of the present invention more clearly, and implement it according to the content of the description, the preferred embodiments of the present invention are described in detail below with the accompanying drawings.

Description of drawings

Fig. 1 is the hydrogen nuclear magnetic spectrum (DMSO-d ₆ ) of the intermediate product PT-1 synthesized in Example 1;

Fig. 2 is the hydrogen nuclear magnetic spectrum (DMSO-d ₆ ) of the intermediate product PT-2 synthesized in Example 1;

Fig. 3 is the nuclear magnetic spectrum of the near-infrared dye NIR 800 synthesized in Example 1 (DMSO-d ₆ );

Fig. 4 is the UV-Vis absorption spectrogram of the near-infrared dye NIR800 synthesized in Example 1 in different solutions and the relationship diagram of absorption intensity and concentration;

Figure 5 is the hydrogen nuclear magnetic spectrum (CDCl ₃ ) of PBnMA synthesized in Example 2, PBnMA-bP (BAPMA-co-PEGMA) synthesized in Example 3, and PBnMA-bP (BAPMA-co-PEGMA) synthesized in Example 4 ;

Fig. 6 is the hydraulic diameter of the modified P7 sample after NIR, the TEM image and the TEM image of the P7 self-assembly;

Figure 7 shows the UV-Vis absorption curve and the time-dependent curve of photothermal conversion temperature of the modified P7 sample after NIR.

Detailed ways

The specific embodiments of the present invention will be described in further detail below with reference to the accompanying drawings and embodiments. The following examples are intended to illustrate the present invention, but not to limit the scope of the present invention.

Chemical reagents used in the following examples of the present invention: benzyl methacrylate (BnMA), polyethylene glycol monomethyl ether methacrylate (PEGMA, M _n =500g/mol), azobisisobutyronitrile (AIBN ), acetic acid (CH ₃ COOH), 2-mercaptothiophene, 2-mercaptothiophene, tert-butyl N-(3-hydroxypropyl)carbamate, anhydrous sodium sulfate (Na ₂ SO ₄ ), tetrahydrofuran (THF), N,N-dimethylformamide (DMF), toluene, methacryloyl chloride and trifluoroacetic acid (TFA) were mainly purchased from Jiangsu Qiangsheng Chemical Co., Ltd. and used directly.

Test apparatus used in the following examples of the present invention: The near-infrared absorption intensity of compound NIR 800 and post-modified micelles PBnMA-bP (APMA-co-PEGMA)@NIR 800 was determined by an ultraviolet spectrophotometer (UV-2600 spectrophotometer) measured. The number-average molecular weight (M _{n, GPC} ) and molecular weight distribution (M _w /M _n ) of homopolymer PBnMA and amphiphilic block copolymer PBnMA-bP (APMA-co-PEGMA) obtained by RAFT polymerization Detector (TOSOH) gel permeation chromatography (GPC, TOSOH, HLC-8320). The instrument is equipped with a guard column (TSKgel SuperMP-N type, 4.6 x 20 mm), two separation columns (TSKgel SupermultiporeHZ-N type, 4.6 x 150 mm), a test column (molecular weight determination range from 5 x 10 ² to 1.9 x 10 ^{5 )} g/mol). The test temperature of the instrument was 40 °C, DMF was selected as the mobile phase, and the flow rate was 0.35 mL/min. GPC samples were injected using a TOSOH plus autosampler, and the molecular weights of the samples were calculated from PS standards purchased from TOSOH. The real-time temperature of NIR800 and post-modified micelles under near-infrared light irradiation was measured by a thermal imager. The ¹ H NMR spectra of the synthesized compounds NIR 800, BAPMA, homopolymers obtained by polymerization, block copolymers and acid hydrolysis products were all determined by nuclear magnetic resonance (Bruker 300MHz) using DMSO-d ₆ or CDCl ₃ as solvent , TMS is the internal standard. The particle size and particle size polydispersity index of the self-assembled micelles were measured by dynamic light scattering (DLS, NanoBrook 90Plus). The micelles were dispersed in water, and the test temperature was 25°C. The morphology and size of self-assembled micelles were measured by transmission electron microscopy (TEM, HITACHI HT7700) at an accelerating voltage of 120 kV. Use a micropipette to pipette 20uL of the dialyzed micellar aqueous solution and carefully drop it on the center of the carbon-coated copper mesh. After standing for one minute, remove the excess liquid with filter paper. Pipette 20 uL of phosphotungstic acid aqueous solution (1.0 wt. %) again and add it dropwise to the copper mesh, wait for half a minute, and absorb the excess liquid. Phosphotungstic acid aqueous solution has a dyeing effect and can better observe the morphology. The prepared copper mesh can be baked under infrared light for a while to make the water evaporate completely.

Example 1: Synthesis of ketoacid cyanine NIR 800

(1) Synthesis of PT-1

Under the protection of argon atmosphere, 2-mercaptothiophene (4.87 g, 41.9 mmol) and methyl 4-piperidinecarboxylate (9.02 g, 63.0 mmol) were dissolved in 50 mL of toluene, and heated under reflux at 110 °C for 2 hours. After the reaction, the reaction solvent, toluene, was removed by rotary evaporation. The product PT-1 was isolated by intelligent fast liquid preparative chromatography system (eluent V _{petroleum ether} : V _{ethyl acetate} =9:1), and the yield was 73.1%; ¹ H NMR (300 MHz, CDCl ₃ ): δ6.76 (d, 1H), δ6.62 (d, 1H), δ6.14 (d, 1H), δ3.71 (s, 3H), δ3.55～3.46 (m, 2H), δ3.06～2.79 ( m, 2H), δ2.49-2.39 (m, 1H), δ2.05-2.00 (m, 2H), δ1.96-1.83 (m, 2H) (Fig. 1).

(2) Synthesis of PT-2

First, PT-1 (5.02 g, 22.3 mmol) was dissolved in a certain amount of sodium hydroxide aqueous solution (0.50 mol/L, 2.93 g NaOH dissolved in 146 mL H ₂ O), and refluxed at 90° C. for 4 hours. When the reaction solution was cooled to room temperature, an aqueous acetic acid solution (1.7 mmol/L, 7.00 g CH ₃ COOH dissolved in 70 mL H ₂ O) was added dropwise to it until a white precipitate appeared. During the dropping process, use a wide range of pH test paper to detect the pH value of the mixed solution. If the pH value of the solution drops to about 2, stop the dropwise addition of the aqueous acetic acid solution. Excess acetic acid and other water-soluble impurities were removed by suction filtration and multiple water washings, and the product was dried in a vacuum oven overnight to obtain a light blue solid product PT-2 with a yield of 59.9%; ¹ H NMR (300 MHz, DMSO- d ₆ ): δ6.73(d, 1H), δ6.70(d, 1H), δ6.13(d, 1H), δ3.45～3.38(m, 2H), δ2.82～2.73(m, 2H), δ2.42-2.33 (m, 1H), δ1.93-1.88 (m, 2H), δ1.73-1.59 (m, 2H) (Fig. 2).

(3) Synthesis of NIR 800

Under the protection of argon atmosphere, PT-2 (1.95g, 9.2mmol) and ketone acid (0.65g, 4.6mmol) were successively added to the mixed solvent of toluene and n-butanol (V _toluene : V _{keto acid} =50mL: 50 mL), refluxed at 90°C for 2 hours. The reaction solvent was removed by suction filtration and washed with anhydrous methanol for several times to remove soluble impurities. Drying in a vacuum oven overnight gave a solid powder with metallic luster in a yield of 84.3%. ¹ H NMR (300MHz, DMSO-d ₆ ): δ12.44(d, 2H), δ8.54(d, 2H), δ7.05(d, 2H), δ4.03～3.99(m, 4H), δ3.51-3.53 (m, 2H), δ3.33-3.31 (m, 4H), δ2.27-2.03 (m, 4H), δ1.75-1.72 (m, 4H) (Fig. 3).

Figure 4 is the UV-Vis absorption spectrum of near-infrared dye NIR800 in (a) buffer solution (pH=8.0) and (b) DMF and (c) the relationship between absorption intensity and concentration (λ=780nm, pH=8.0 buffer solution). The results show that it can absorb near-infrared light, and the absorption capacity is proportional to the concentration.

Example 2: Synthesis of PBnMA

PBnMA was synthesized according to the following method, and the molar ratio of [BnMA] ₀ : [CPDN] ₀ : [AIBN] ₀ was changed to synthesize products P1, P2 and P3 respectively, and the molar ratio of each component was [BnMA] ₀ : [CPDN] ₀ :[AIBN] ₀ = 5:1:0.3 as an example, the method is as follows:

Monomer BnMA (0.5 mL, 3.0 mmol), RAFT regulator CPDN (0.1623 g, 0.6 mmol), initiator AIBN (29.6 mg, 0.18 mmol), solvent toluene (1.0 mL) and a crystalline and dry magnetic stir bar were combined Sequentially add to 5mL ampoules. The oxygen in the ampoules was removed by three cycles of freeze-pump-thaw on a double-row tube device, and the tubes were sealed by flame fusion. After the oil bath has warmed to 80°C, the ampoule is placed in it and placed in stirring mode. After 20 hours of reaction, the ampoule was taken out and cooled under running water to quench the reaction. After the tube is broken, add an appropriate amount of THF to the ampoule to dissolve the polymer, and the diluent is added dropwise to a large amount of petroleum ether for precipitation. After sufficient precipitation time, the polymer was obtained by suction filtration. The polymer after suction filtration was dried in a vacuum oven at 35°C to a constant weight, and the monomer conversion was calculated by the weighing method. The performance test results of the synthesized PBnMA are shown in Table 1. Polymers with different molecular weights can be designed and synthesized by adjusting the ratio of the monomer and the RAFT reagent CPDN, and the molecular weight distribution of the polymers is very narrow (M _w /M _n is less than 1.13). .

Table 1: Performance test results of PBnMA

In Table 1, R=[BnMA] ₀ /[CPDN] ₀ /[AIBN] ₀ molar ratio. ^a indicates that the molecular weight and molecular weight distribution are measured by GPC, the standard of GPC is PS in DMF (0.1 wt.% LiBr) solution, ^b indicates that the molecular weight is calculated by hydrogen NMR spectroscopy.

Example 3: Synthesis of PBnMA-b-P (BAPMA-co-PEGMA)

Different PBnMAs (P1, P2, P3) synthesized in Example 2 were used as macromolecular regulators, and BAPMA and PEGMA were used as monomers to synthesize different PBnMA-bP (BAPMA-co-PEGMA) (P4-P9), wherein, The molecular weight of PEGMA) is 500g/mol. Taking [BAPMA] ₀ :[PEGMA] ₀ :[PBnMA] ₀ :[AIBN] ₀ =10:10:1:0.3 as an example, the synthesis method is as follows:

BAPMA (0.1013g, 0.42mmol), PEGMA (0.2083g, 0.42mmol), PBnMA (0.10g, 0.042mmol), AIBN (2.1mg, 0.013mmol), toluene (3mL) and a clean and dry small magnet were sequentially mixed Add to 5mL ampoules. After three freeze-pump-thaw cycles, the tube was sealed with a flame, and the ampoule was placed in an oil bath at 80°C for 24 hours of reaction. After the reaction was completed, the ampoule was taken out and cooled to room temperature. Break the tube, add an appropriate amount of THF to the ampoule to dissolve the polymer, and add the diluent dropwise to a large amount of petroleum ether for precipitation. After sufficient precipitation time, the polymer was obtained by suction filtration. The polymer after suction filtration was dried in a vacuum oven at 35°C to a constant weight, and the monomer conversion was calculated by the weighing method. The synthesized PBnMA-b-P (BAPMA-co-PEGMA) is shown in Table 2. Blocks with different molecular weights can be synthesized by adjusting the ratio of comonomer BAPMA and PEGMA, and using PBnMA with different molecular weights (P1, P2, P3). Copolymers and the molecular weight distribution of the polymers are mostly narrow, indicating that the macromolecular RAFT reagent PBnMA has a high degree of terminal functionalization.

Table 2 Synthesis of PBnMA-b-P (BAPMA-co-PEGMA)

Example 4: Synthesis of PBnMA-b-P (BAPMA-co-PEGMA)

The tert-butyl carbamate group in the monomer BAPMA in the different products synthesized in Example 3 was removed to expose the amino group to react with the carboxyl group in NIR 800 for post-modification. That is, PBnMA-b-P(APMA-co-PEGMA) can be obtained by acid hydrolysis of PBnMA-b-P(BAPMA-co-PEGMA). Specific steps are as follows:

The polymer PBnMA-b-P (BAPMA-co-PEGMA) (100 mg, 0.042 mmol) synthesized by the above was dissolved in 2.0 mL of dichloromethane, and 0.5 mL of trifluoroacetic acid (TFA) was quickly added to it, and the reaction was carried out under sealing conditions. The solution was stirred at room temperature overnight. After the reaction, the reaction mixture was precipitated with petroleum ether for 2 to 3 times, and the precipitate was dried in a constant temperature vacuum oven at 35°C. From the H NMR spectrum (Fig. 5(c), the complete disappearance of the peak at 1.40 ppm to 1.50 ppm was observed, which proved that the tert-butoxycarbonyl group (BOC) was successfully removed, and PBnMA-b-P(APMA- co-PEGMA).

Example 5: Synthesis of PBnMA-b-P(APMA-co-PEGMA)@NIR800

PBnMA-bP(APMA-co-PEGMA)@NIR800 was prepared by post-modification reaction of PBnMA-bP(APMA-co-PEGMA) and NIR800. The details are as follows: different acid hydrolysis products PBnMA-bP(APMA-co-PEGMA) (2mg) prepared in Example 4 and an appropriate amount of ketoacid cyanine dye NIR 800 (0.6mg, 1.14×10 ^-3 mmol) were simultaneously dissolved in 1.0 In mL DMF, the reaction was carried out overnight to complete the reaction to achieve the purpose of post-modification to obtain PBnMA-bP(APMA-co-PEGMA)@NIR800.

Example 6: Preparation of micelles

The PBnMA-b-P(APMA-co-PEGMA)@NIR800 synthesized from the product P7 synthesized in Example 3 was used as the raw material to prepare micelles, and the micelles were obtained by assembling with a selective solvent, and the method was as follows:

First, the polymer PBnMA-b-P(APMA-co-PEGMA)@NIR800 (2 mg) was dissolved in 1.0 mL of DMF, and the solution was fully shaken for half an hour in a sonicator to obtain a completely dissolved polymer solution. 10 mL of deionized water was added dropwise to the polymer solution via a syringe pump over 2 hours at a constant temperature of 25°C with gentle stirring. After the dropwise addition, the obtained solution was placed in a dialysis bag with a molecular weight cut-off of 3500 g/mol, dialyzed against deionized water for 24 hours and continuously changed the water to remove DMF in the reaction solution by dialysis. Since the mixed solution gradually changed from alkaline to neutral, excess NIR 800 that did not participate in the reaction was precipitated from the solution, so it was centrifuged at a speed of 10,000 r/min for 10 minutes in a high-speed centrifuge to remove the precipitate. The supernatant after centrifugation, that is, the assembly solution, was dialyzed again in a buffer solution with pH=8.0 for 12 hours to remove the NIR800 embedded in the micelles. Finally, it was dialyzed in deionized water for 24 hours. At this time, all the NIR 800 structures in the micellar aqueous solution were chemically bound to the micellar core-shell structure, and there was no physical entrapment form.

As a control, PBnMA-b-P (BAPMA-co-PEGMA) synthesized in Example 3 and PBnMA-b-P (BAPMA-co-PEGMA) synthesized in Example 4 were prepared into micelles according to the same method, and the polymers were dissolved. In 1.0 mL of DMF, fully shaken for half an hour in a sonicator to obtain a completely dissolved polymer solution. 10 mL of deionized water was added dropwise to the polymer solution via a syringe pump over 2 hours at a constant temperature of 25°C with gentle stirring. After the dropwise addition, the obtained solution was placed in a dialysis bag with a molecular weight cut-off of 3500 g/mol, dialyzed against deionized water for 24 hours and continuously changed the water to remove DMF in the reaction solution by dialysis.

The particle size and particle size distribution of each micelle are shown in Table 3. In Table 3, the particle size and particle size distribution are obtained by DLS test, wherein the polymers 1-3 are respectively P4-P9 synthesized in Example 3. Raw materials are obtained, polymer 1 represents the micelle formed by the product PBnMA-b-P (BAPMA-co-PEGMA) synthesized in Example 3; Polymer 2 represents the product PBnMA-b-P (APMA-co-PEGMA) synthesized by Example 4 ); polymer 3 represents the micelle formed by the product PBnMA-b-P(APMA-co-PEGMA)@NIR800 synthesized in Example 5.

Table 3 Self-assembly of different block polymers

The self-assembly results showed that (Table 3), under the same conditions of the lipophilic chain length and the degree of polymerization of BAPMA, the longer the chain length of the hydrophilic chain PPEGMA, the better the amphiphilic block copolymer PBnMA-b-P (BAPMA-co-PEGMA) The larger the size of the micelles obtained by self-assembly, the size of the assembled micelles of the block copolymer after acidolysis showed the same trend. This is caused by the increase of the repulsive force between the hydrophilic shells as the hydrophilic chain length increases. The block copolymer P7, which forms micelles of smaller size, was selected for post-modification of NIR800. Figure 6(c) is the TEM image of the micelle formed by the block copolymer P7, and Figure 6b is the TEM image of the micelle formed by the NIR 800 modified P7 polymer. The results show that the post-modified block copolymer undergoes the same assembly. After that, the size of the obtained micelles decreased further. The reason is speculated as follows: in the process of amidation, one carboxyl group of NIR 800 is not only chemically bonded to the copolymer chain, but also the other hydroxyl group reacts with the amino group, and the amino group is more likely to be located on a different copolymer chain, so that Different copolymer chains were slightly cross-linked, which made the micellar structure obtained by subsequent assembly more dense, and the particle size of the assembly decreased to 114 nm. It can be seen from the TEM image (Fig. 6(b)) that the assemblies are spherical particles with a corresponding diameter of about 38 nm, which is much lower than the 114 nm measured by DLS (Fig. 6(a)). This is mainly due to the fact that the diameter of the self-assembly in the dry state is measured by TEM.

Figure 7(a) is the UV absorption curve of NIR800 post-modified polymer PBnMA-bP(APMA-co-PEGMA)@NIR800 micelles after 5-fold dilution, where C _{NIR 800} =0.07 mg/mL, from which we calculated NIR The modification efficiency of 800 pairs of deprotected block copolymer PBnMA-bP (APMA-co-PEGMA) was 39.3%. Under the irradiation of 810 nm near-infrared light (0.028 W/cm ² ), the temperature of the post-modified micelle aqueous solution increased from the initial ambient temperature of 20 °C to about 55 °C within one hour (Fig. 7(b)). This indicates that the micelles formed by PBnMA-bP(APMA-co-PEGMA)@NIR800 also have good near-infrared photothermal responsiveness.

The present invention adopts the RAFT polymerization method to synthesize the amphiphilic block copolymer PBnMA-bP (BAPMA-co-PEGMA), wherein, benzyl methacrylate (BnMA) is used as the lipophilic monomer, and methyl methacrylates with a molecular weight of 500 are respectively used. Polyethylene glycol monomethyl ether acrylate (PEGMA ₅₀₀ ) is a hydrophilic monomer, BAPMA is a functional monomer, α-dithionaphthoic acid isobutyronitrile (CPDN) is a chain transfer agent, azodiiso Nitrile (AIBN) is the initiator. The self-assembly of the obtained amphiphilic block copolymers was studied by solution self-assembly method. With the growth of the chain segment, the particle size of the micelle increases. The growth of the hydrophilic chain leads to the increase of the repulsive force between the micelle shells, which leads to the size change of the micelles. Subsequently, the carboxyl group on the ketoacid cyanine dye NIR 800, which has a strong absorption of 800 nm near-infrared light and has a good photothermal conversion ability, was used to directly interact with the deprotected amino group-containing block in DMF solution. The copolymer reaction, which played a post-modification role, successfully reacted NIR 800 to the copolymer chain with a modification efficiency of 39.3%. The post-modified block copolymer was also assembled by solution self-assembly to obtain spherical nanomicelles with a size of 114 nm and a polydispersity index (PDI) of 0.059. At the same time, the post-modified micelles can also be irradiated by near-infrared light. When the temperature was raised to above 50 °C, nanomicelles with good near-infrared photothermal response properties were prepared, which opened up a route for modifying amphiphilic block copolymers with ketone acid cyanine dyes.

The above are only the preferred embodiments of the present invention and are not intended to limit the present invention. It should be pointed out that for those skilled in the art, some improvements and modifications can be made without departing from the technical principles of the present invention. , these improvements and modifications should also be regarded as the protection scope of the present invention.

Claims (10)

1. An amphiphilic block copolymer having near-infrared thermal responsiveness, characterized in that it comprises a structure represented by formula (1):

wherein x is 5-30; y is 5-30; z is 5-30; and m is 5-21.

2. A method for preparing the amphiphilic block copolymer according to claim 1, comprising the steps of:

(1) in a protective atmosphere, reacting benzyl methacrylate BnMA in an organic solvent at 65-85 ℃ under the action of a RAFT regulator α -isobutylnaphthoate CPDN and a thermal initiator, and obtaining a polymer PBnMA after complete reaction;

(2) in a protective atmosphere, taking the polymer PBnMA as a macromolecular regulator, reacting methacrylamide BAPMA protected by tert-butyl carbamate and polyethylene glycol monomethyl ether methacrylate PEGMA in an organic solvent at 65-85 ℃ under the action of a thermal initiator, and obtaining a block polymer PBnMA-b-P (BAPMA-co-PEGMA) after the reaction is completed;

(3) removing a protecting group tert-butyl carbamate group on BAPMA in the block polymer PBnMA-b-P (BAPMA-co-PEGMA) to expose amino in methacrylamide to obtain PBnMA-b-P (APMA-co-PEGMA);

(4) reacting the PBnMA-b-P (APMA-co-PEGMA) and croconium cyanine dye NIR800 in an organic solvent at the temperature of 20-25 ℃ to obtain the amphiphilic block copolymer after the reaction is completed; wherein the structural formula of the croconium cyanine dye NIR800 is as follows:

3. the method of claim 2, wherein: in step (1), the molar ratio of BnMA, CPDN and thermal initiator is 5-30:1: 0.3.

4. The method of claim 2, wherein: in step (2), the molar ratio of BAPMA, PEGMA, PBnMA and thermal initiator is 10:5 to 10:1: 0.3.

5. The production method according to claim 2, wherein in the step (3), the method for removing a tert-butylcarbamate group comprises the steps of:

the block polymer PBnMA-b-P (BAPMA-co-PEGMA) is subjected to acidolysis reaction in an organic solvent under the action of trifluoroacetic acid, and the reaction temperature is 20-25 ℃.

6. The method of claim 2, wherein: in step (4), the molar ratio of PBnMA-b-P (APMA-co-PEGMA) to NIR800 is 0.1-0.5: 1.

7. An amphiphilic block copolymer micelle with near-infrared thermal responsiveness, which is characterized in that: the amphiphilic block copolymer of claim 1, wherein the amphiphilic block copolymer micelle comprises a lipophilic inner core and a hydrophilic outer shell wrapping the outer part of the inner core, wherein the inner core comprises PBnMA, the outer shell comprises P (APMA-co-PEGMA), and the outer shell is connected with croconium cyanine dye with near infrared photo-responsiveness through covalent bonds.

8. The amphiphilic block copolymer micelle having near-infrared thermal responsiveness according to claim 7, wherein: the particle size of the amphiphilic block copolymer micelle is 100-600 nm.

9. The amphiphilic block copolymer micelle having near-infrared thermal responsiveness according to claim 7, wherein: the amphiphilic block copolymer micelle is obtained by self-assembling the amphiphilic block copolymer according to claim 1 in a co-solvent of an organic solvent and water.

10. Use of the amphiphilic block copolymer micelle having a near-infrared thermo-responsiveness according to claim 7 in preparation of a near-infrared photo-responsive material.

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