Compound, preparation method of compound, photo-thermal agent and application of photo-thermal agent in obesity treatment

Abstract

The invention relates to the technical field of biological medicine, in particular to a compound and a preparation method thereof, a photo-thermal agent and application of the photo-thermal agent in obesity treatment. The compound has a structure as shown in a formula I in the specification, wherein R is selected from at least one of anthracene-containing groups, dibenzothiophene-containing groups or carbazole-containing groups, and X is selected from at least one of halogen atoms, PF6 <-> and BF4 <->. The compound provided by the invention has mitochondrial targeting property and good photothermal conversion capability, and can realize obesity treatment by promoting the browning process of white adipose tissues.

Description

TECHNICAL FIELD

[0001] The present application relates to the technical field of biological medicine, and particularly relates to a compound, a preparation method thereof, a photothermal agent and application of the photothermal agent in obesity treatment. BACKGROUND

[0002] Obesity is the main cause of type 2 diabetes, hypertension, coronary heart disease, stroke, fatty liver and osteoarthritis and other diseases. In addition, obesity is also closely related to a variety of cancers, including breast cancer, colon cancer and endometrial cancer. More attention is paid to the fact that the obesity problem is increasingly attacking the adolescent population. Overweight in childhood and adolescence directly affects health and is associated with increased and early onset of diseases such as diabetes and atherosclerosis. In addition, obesity in children and adolescents also has adverse social and psychological consequences.

[0003] The healthiest way to control weight usually includes dietary intervention and regular exercise, which helps maintain overall health but has limited effect on some diseases caused by obesity. At present, in addition to liposuction surgery with certain risks in the field of medical cosmetology, there are other medical means for treating obesity. White adipose tissue (WAT) is the main fat tissue in the human body for storing excess energy, and its excessive accumulation can lead to obesity. In contrast, brown adipose tissue (BAT) is rich in a large number of mitochondria and can convert the energy in food into heat. The mitochondria in BAT contain uncoupling protein 1 (UCP1), which can prevent the energy generated by glucose and fatty acid metabolism from being used to synthesize adenosine triphosphate (ATP), thereby promoting the release of energy in the form of heat. Therefore, the browning of white adipose tissue (i.e. the transformation of WAT to BAT) as a potential obesity treatment strategy is attracting more and more attention from researchers. Studies have shown that activating transient receptor potential vanilloid 1 (TRPV1) is one of the key factors to promote the remodeling of WAT and achieve its transformation to BAT. TRPV1 can be activated by heating (>42°C).

[0004] Photothermal agents (PTA) can convert high-energy light into strong heat and are widely studied in photothermal therapy (PTT) and have been applied in the fields of anti-tumor and antibacterial. PTA can meet the conditions for activating TRPV1, so it has played an important role in the treatment of obesity in recent years.

[0005] In view of the above, the present application is proposed. SUMMARY

[0006] The present application aims to provide a compound, a preparation method thereof, a photothermal agent and an application in obesity treatment. The compound has mitochondrial targeting and good photothermal conversion ability, and can realize the treatment of obesity by promoting the browning process of white adipose tissue.

[0007] To achieve the above-mentioned purpose of the present application, the first aspect of the present application provides a compound having the structure as shown in formula I: wherein R is selected from at least one of anthracene, dibenzothiophene or carbazole groups, and X is selected from at least one of halogen atoms, PF6 - and BF4 - .

[0008] In some embodiments, R is selected from any one of , and .

[0009] In some embodiments, X is selected from at least one of I - , PF6 - and BF4 - .

[0010] In some embodiments, the compound has the following structural formula: , , , , .

[0011] The second aspect of the present application provides a preparation method of the compound of the first aspect, comprising the following steps: (a) performing Suzuki coupling reaction of compound A and compound B in a solvent to obtain compound C; (b) performing reaction of compound C and 1,4-dimethylpyridine-1-iodide in a solvent to obtain a compound as shown in formula I1; wherein the structural formulae of compound A, compound B, compound C and compound I1 are as follows: , , , .

[0012] In some embodiments, when X is PF6 - or BF4 - ​The preparation method further includes reacting compound I1 with KPF6 or NaBF4 in a solvent to obtain a compound as shown in Formula I.

[0013] A third aspect of the present invention provides a photothermal agent comprising the compound provided in the first aspect of the present invention.

[0014] In a specific embodiment of the present invention, the photothermal agent comprises a compound and an amphiphilic copolymer. Further, the amphiphilic copolymer comprises amphiphilic copolymer F127.

[0015] In a specific embodiment of the present invention, the mass ratio of the compound to the amphiphilic copolymer is 1:(5-15).

[0016] The fourth aspect of the present invention provides the use of the compounds of the first aspect or the photothermal agents provided in the third aspect of the present invention in the preparation of medical imaging agents, optical imaging agents or staining agents.

[0017] The fifth aspect of the present invention provides the use of the compounds of the first aspect or the photothermal agents provided in the third aspect of the present invention in the preparation of a medicament for the treatment of obesity.

[0018] In some embodiments, the drug is used to promote the transformation of white adipocytes into brown adipocytes.

[0019] Compared with the prior art, the advantages of the present invention are as follows: The compounds of this invention have mitochondrial targeting properties and good photothermal conversion capabilities, enabling them to treat obesity by promoting the browning process. Attached Figure Description

[0020] To more clearly illustrate the specific embodiments of the present invention or the technical solutions in the prior art, the drawings used in the description of the specific embodiments or the prior art will be briefly introduced below. Obviously, the drawings described below are some embodiments of the present invention. For those skilled in the art, other drawings can be obtained from these drawings without creative effort.

[0021] Figure 1 The UV-Vis absorption spectra (a) and normalized fluorescence emission spectra (b) of TPBNaI, TPBThI, and TPBCaI in DMSO; the UV-Vis absorption spectra (c) and normalized fluorescence emission spectra (d) of TPBCaI, TPBCaPF6, and TPBCaBF4 in DMSO. Figure 2 Normalized fluorescence emission spectra of TPBNaI, TPBThI, and TPBCaI in the solid state; Figure 3The graph shows the change in fluorescence intensity of TPBNaI, TPBThI, and TPBCaI as a function of water content, representing the ratio of their initial fluorescence intensity to the fluorescence intensity. Figure 4 The temperature changes of TPBNaI, TPBThI, TPBCaI, TPBCaPF6, and TPBCaBF4 under different irradiation times; Figure 5 This is a size distribution diagram of TPBCaI NPs; Figure 6 The UV-Vis absorption spectra (a) and normalized fluorescence emission spectra (b) of TPBaI NPs, TPBThI NPs, TPBcaI NPs, TPBcaPF6 NPs and TPBcaBF4 NPs in water are shown. Figure 7 The temperature changes of TPBNaI NPs, TPBThI NPs, TPBCaI NPs, TPBCaPF6 NPs, and TPBCaBF4 NPs under different irradiation times are shown. Figure 8 Infrared thermal images of TPBNaI NPs, TPBThI NPs, TPBCaI NPs, TPBCaPF6 NPs, and TPBCaBF4 NPs under different irradiation times; Figure 9 Temperature variation of TPBCaI NPs under 4 on-off cycles (a), and photothermal conversion efficiency (PCE, η) of TPBCaI NPs (b); Figure 10 The cytotoxicity of different concentrations of TPBCaI NPs on 3T3 cells (a) and the phototoxicity of 10 μM TPBCaI NPs on 3T3 cells (b). Figure 11 Images of Mito-Tracker Green, TPBCaI NPs, and combined confocal microscopy. Figure 12 Confocal laser scanning microscope fluorescence image of 3T3 stained with calcein-AM / PI; Figure 13 A schematic diagram of adipogenesis and differentiation in 3T3 cells (a), Oil Red O staining results of intracellular lipid droplets after treatment under different conditions (b), fluorescence images of intracellular calcium ion concentration after treatment under different conditions (c), immunofluorescence images of UCP1-labeled (green) cells after treatment under different conditions (d), and schematic diagrams of intracellular ATP levels (e) and increased intracellular calcium concentration and UCP1 generation after treatment under different conditions (f). Figure 14(a) Relative body weight change, Lee's index in normal and obese mouse model groups (b) Photographs of iWAT and eWAT in normal group (c) Photographs of iWAT and eWAT in obese mouse model group (d) In vivo biodistribution of TPBCaI NPs in major organs and adipose tissue of mice 1 h and 24 h after subcutaneous injection (e). Figure 15 Schematic diagram of the obese mouse model and photothermal treatment of obese mice (a), photos of mice after treatment under different conditions on day 14 (b), relative weight changes during treatment (c), photos of iWAT and eWAT of mice after treatment under different conditions after euthanasia (d) and relative fat content (e). Figure 16 Serum levels of TG (a), TC (b), HDL-C (c), LDL-C (d), FFA (e), and insulin (f), and H&E staining images of iWAT, scale bar = 100 μm (g). Data represent mean ± standard deviation (n = 3 mice per group); one-way ANOVA: ns = not significant; *P<0.05, **P<0.01, ***P<0.001 compared with the control group; ##P<0.01, ###P<0.001 between specified groups. Detailed Implementation

[0022] The technical solution of the present invention will be clearly and completely described below with reference to the accompanying drawings and specific embodiments. However, those skilled in the art will understand that the embodiments described below are some embodiments of the present invention, but not all embodiments, and are only used to illustrate the present invention, and should not be regarded as limiting the scope of the present invention. Based on the embodiments of the present invention, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of the present invention. Where specific conditions are not specified in the embodiments, conventional conditions or conditions recommended by the manufacturer shall be followed. Where the manufacturers of reagents or instruments are not specified, they are all conventional products that can be purchased commercially.

[0023] The first aspect of this invention provides a compound having the structure shown in Formula I: ; Wherein, R is selected from at least one of anthracene, dibenzothiophene, or carbazole groups, and X is selected from halogen atoms, PF6 - and BF4 - At least one of them.

[0024] The compounds of this invention include a highly distorted structure, strong electron-donating and accepting groups, and cations, resulting in PTT dominance. The compounds of this invention possess mitochondrial targeting and high photothermal conversion efficiency (PCE), enabling the treatment of obesity by promoting the browning process of white adipose tissue.

[0025] In some implementations, R is selected from... , and Any of the above, preferably * represents the linking site of the corresponding compound structure of R.

[0026] The inventors of this invention discovered in their research that compounds with carbazole as a donor have superior photothermal properties compared to compounds with anthracene or dibenzothiophene as donors.

[0027] In some implementations, X is selected from I - PF6 - and BF4 - At least one of them.

[0028] In some embodiments, the compound has the following structural formula: (Named TPBAnI) (Named TPBThI) (Named TPBCaI) (Named TPBCaPF6) (Named TPBCaBF4).

[0029] The second aspect of the present invention provides a method for preparing the compound of the first aspect, comprising the following steps: (a) Compound A and compound B undergo a Suzuki coupling reaction in a solvent to give compound C; (b) Compound C reacts with 1,4-dimethylpyridine-1-iodide in a solvent to give the compound shown in Formula I1; The structural formulas of compounds A, B, C, and I1 are as follows: , , , .

[0030] In some embodiments, the Suzuki coupling reaction of compound A and compound B in step (a) can be carried out under conventional conditions. In an alternative embodiment, the catalyst comprises potassium phosphate and tetrakis(triphenylphosphine)palladium; the molar amount of potassium phosphate can be 1 to 1.5 times, such as 1.2 times, the molar amount of tetrakis(triphenylphosphine)palladium can be 5% to 15%, such as 10%, the molar amount of compound A.

[0031] In some embodiments, in step (a), the molar ratio of compound A to compound B is 1:(1 to 1.5), preferably 1:1.2.

[0032] In some embodiments, compound B includes any one of anthracene-2-ylboronic acid, dibenzo[b,d]thiophene-2-boronic acid, and (9-ethyl-9H-carbazole-3-yl)boronic acid.

[0033] In some embodiments, in step (a), the solvent includes, but is not limited to, 1,4-dioxane.

[0034] In some embodiments, in step (a), the Suzuki coupling reaction is carried out under a protective atmosphere, including but not limited to argon. Further, in step (a), the temperature of the Suzuki coupling reaction is 90–110°C, and the reaction time is 6–12 h.

[0035] In some embodiments, in step (a), after the Suzuki coupling reaction is completed, the reacted material is cooled to room temperature; the reacted material is extracted with ethyl acetate and the crude product is obtained by rotary evaporation under reduced pressure; then, the product is purified by column chromatography to obtain compound C.

[0036] The synthetic route for obtaining compound C from compound A and compound B in this invention can be found below:

[0037] In some embodiments, the preparation of compound A includes reacting 4-iodobenzaldehyde, diphenylyne, and 4-bromostyrene in a solvent in the presence of a catalyst. Further, the catalyst includes lithium chloride, triphenylphosphine, palladium acetate, and sodium bicarbonate. The molar amount of lithium chloride can be 60%–80% (e.g., 70%) of the molar amount of 4-iodobenzaldehyde; the molar amount of triphenylphosphine can be 5%–15% (e.g., 10%) of the molar amount of 4-iodobenzaldehyde; the molar amount of palladium acetate can be 2%–8% (e.g., 5%) of the molar amount of 4-iodobenzaldehyde; and the molar amount of sodium bicarbonate can be 1.5–2.5 times (e.g., 2 times) the molar amount of 4-iodobenzaldehyde.

[0038] In some embodiments, the molar ratio of 4-iodobenzaldehyde, diphenylyne and 4-bromostyrene in the preparation of compound A is 1:(3-5):1, preferably 1:4:1.

[0039] In some embodiments, the solvent used in the preparation of compound A includes, but is not limited to, N,N-dimethylformamide (DMF).

[0040] In some embodiments, the preparation of compound A is carried out under a protective atmosphere, including but not limited to argon. Further, in the preparation of compound A, the reaction temperature is 110–130°C, and the reaction time is 4–8 h.

[0041] In some embodiments, in the preparation of compound A, after the reaction is completed, the reactants are cooled to room temperature; the reactants are extracted with dichloromethane and the crude product is obtained by rotary evaporation under reduced pressure; then, compound A is purified by column chromatography.

[0042] The synthetic route for compound A of this invention can be found as follows:

[0043] In some embodiments, in step (b), the solvent includes, but is not limited to, tetrahydrofuran and methanol. Further, the volume ratio of tetrahydrofuran to methanol in the solvent is 1:(2-4), such as 1:3.

[0044] In some embodiments, in step (b), the reaction temperature is 70–80°C and the reaction time is 6–10 h.

[0045] In some embodiments, the reaction of compound C with 1,4-dimethylpyridine-1-iodide is carried out under the catalysis of piperidine. The amount of piperidine added is the conventional catalytic dosage, and there is no specific limitation.

[0046] In some embodiments, the molar ratio of compound C to 1,4-dimethylpyridine-1-iodide is 1:(1 to 1.5), preferably 1:1.2.

[0047] In some embodiments, in step (b), after the reaction is complete, the mixture is cooled to room temperature and rotary evaporated under reduced pressure to obtain a crude product; then, column chromatography is performed to purify the product to obtain the compound shown in Formula I1.

[0048] The synthetic route for the compound of the present invention as shown in Formula I1 can be found as follows:

[0049] In some implementations, when X is PF6 - Or BF4 -The preparation method also includes reacting compound I1 with KPF6 or NaBF4 in a solvent to obtain the compound shown in Formula I.

[0050] In some embodiments, the solvent in the reaction of compound I1 with KPF6 or NaBF4 includes, but is not limited to, dichloromethane.

[0051] In some embodiments, the reaction of compound I1 with KPF6 or NaBF4 specifically includes: dissolving compound I1 in a solvent, adding KPF6 or NaBF4 pre-dissolved in the solvent to the solvent at room temperature, and stirring the reaction at room temperature. Further, the stirring reaction time is 1–3 hours.

[0052] In some embodiments, the molar ratio of compound I1 to KPF6 or NaBF4 is 1:(1 to 1.5), preferably 1:1.

[0053] In some embodiments, after the reaction is completed by stirring at room temperature, the reactants are filtered, the filtrate is collected, and the compound is obtained by rotary evaporation under reduced pressure as shown in Formula I.

[0054] A third aspect of the present invention provides a photothermal agent comprising the compound provided in the first aspect of the present invention.

[0055] In some embodiments, the photothermal agent comprises a compound and an amphiphilic copolymer. Further, the amphiphilic copolymer comprises amphiphilic copolymer F127.

[0056] In some embodiments, the mass ratio of the compound to the amphiphilic copolymer is 1:(5 to 15), specifically a range of 1:5, 1:8, 1:10, 1:12, 1:15, or any two of these.

[0057] In some embodiments, the photothermal agent is nanoparticles. Further, the average particle size of the nanoparticles is 40–55 nm, specifically within the range of 40 nm, 42 nm, 45 nm, 48 nm, 50 nm, 52 nm, 55 nm, or any combination thereof.

[0058] The fourth aspect of the present invention provides the use of the compounds of the first aspect or the photothermal agents provided in the third aspect of the present invention in the preparation of medical imaging agents, optical imaging agents or staining agents.

[0059] The fifth aspect of the present invention provides the use of the compounds of the first aspect or the photothermal agents provided in the third aspect of the present invention in the preparation of a medicament for the treatment of obesity.

[0060] In some embodiments, the drug is used to promote the conversion of white adipocytes into brown adipocytes. Further, the heat generated by the drug under irradiation conditions activates the TRPV1 channel, promoting the conversion of WAT to BAT.

[0061] Example 1 This embodiment provides a method for preparing compound TPBAnI, and the synthetic route is as follows:

[0062] Specifically, the preparation methods of compound TPBAnI include: (1) Preparation of compound A: 4-Iodobenzaldehyde (2.32 g, 10 mmol), diphenylyne (7.12 g, 40 mmol), 4-bromostyrene (1.30 mL, 10 mmol), lithium chloride (0.30 g, 7 mmol), triphenylphosphine (0.26 g, 1 mmol), palladium acetate (0.11 g, 0.5 mmol), and sodium bicarbonate (1.7 g, 20 mmol) were added to a 100 mL round-bottom flask, followed by 20 mL of DMF. The mixture was purged with argon gas three times, and then reacted in an oil bath at 120 °C for 6 h. After the reaction was completed by TLC, the mixture was cooled to room temperature, and the reactants were extracted three times with dichloromethane and water. The dichloromethane phase was collected, dried over MgSO4, and purified by rotary evaporation under reduced pressure. The purified product, compound A, was obtained by silica gel column chromatography (mobile phase: ethyl acetate and petroleum ether in a volume ratio of 1:10) in 57% yield.

[0063] The structural characterization data of compound A are as follows: 1 H NMR (400 MHz, Chloroform- d ) δ 9.96 (d, J = 86.8 Hz, 1H), 7.91 (d, J = 7.9 Hz, 1H), 7.54 (d, J = 8.0 Hz, 1H), 7.48 (d, J = 7.9 Hz, 1H), 7.41 – 7.34(m, 3H), 7.28 (d, J = 5.5 Hz, 1H), 7.24 (d, J = 2.5 Hz, 2H), 7.18 (qd, J = 6.7,6.1, 4.0 Hz, 3H), 7.06 (dd, J = 8.3, 4.7 Hz, 5H), 6.89 – 6.82 (m, 2H), 6.25(dd, J = 16.0, 5.2 Hz, 1H).13 C NMR (101 MHz, CDCl3) δ (ppm): 192.06, 142.10,141.91, 141.60, 140.68, 140.38, 139.71, 132.97, 132.86, 131.87, 131.79,131.36, 131.02, 129.60, 128.99, 128.41, 128.06, 127.76, 126.90. HRMS (APCl, m / z ) Calcd for C 29 H 21 BrO [M+H] + : 465.0854, found: 468.0851. (2) Preparation of compound C: Compound A (0.93 g, 2 mmol), anthracene-2-ylboronic acid (0.53 g, 2.4 mmol), potassium phosphate (0.82 g, 2.4 mmol), and tetrakis(triphenylphosphine)palladium (0.23 g, 0.2 mmol) were dissolved in 10 mL of 1,4-dioxane. The mixture was refluxed at 100 °C for 10 h, cooled to room temperature, and extracted with ethyl acetate. The mixture was washed with water, and the organic layer was collected and dried over anhydrous MgSO4. The solvent was evaporated under reduced pressure, and the mixture was purified by silica gel column chromatography (mobile phase: ethyl acetate and petroleum ether, v / v) to give compound C, an orange solid, in 67.4% yield.

[0064] The structural characterization data of compound C are as follows: 1 H NMR (400 MHz, Chloroform- d ) δ 9.86 (s, 1H), 8.44 (d, J = 11.7 Hz, 3H), 8.18 (s, 1H), 8.06 (d, J = 8.8 Hz, 2H), 7.93 (d, J = 8.0 Hz, 3H), 7.70(dd, J = 15.3, 8.5 Hz, 5H), 7.55 (d, J = 8.1 Hz, 2H), 7.48 – 7.43 (m, 4H), 7.36 – 7.30 (m, 5H), 6.90 (s, 2H), 6.48 – 6.28 (m, 2H). 13C NMR (101 MHz, CDCl3) δ191.93, 139.83, 134.02, 133.64, 132.09, 132.03, 131.84, 131.76, 131.48,131.44, 131.36, 131.14, 130.88, 130.86, 129.49, 128.88, 128.84, 128.29,128.24, 128.23, 128.17, 128.07, 127.88, 127.64, 127.48, 127.44, 127.33,127.24, 127.19, 126.61, 126.05, 125.57, 125.47, 125.39, 125.12. HRMS (APCl, m / z ) Calcd for C 43 H 30 O [M+Na] + : 585.2194, found: 585.2197. (3) Preparation of compound TPBAnI: Compound C (1.13 g, 2 mmol) and 1,4-dimethylpyridine-1-iodide (0.56 g, 2.4 mmol) were dissolved in 28 mL of a tetrahydrofuran / methanol mixture (v / v ratio 1:3). A few drops (e.g., 5 drops) of piperidine were added to the mixture, and the mixture was refluxed at 75 °C for 8 h. The reactants were cooled to room temperature, and the solvent was removed by rotary evaporation under reduced pressure. The product was purified by alumina column chromatography (mobile phase: methanol and dichloromethane, v / v ratio 1:14) to give the red solid product compound TPBAnI in 45.7% yield.

[0065] The structural characterization data of compound TPBAnI are as follows: 1 H NMR (400 MHz, CDCl3) δ 8.78 (s, 1H), 8.30 (d, J = 13.0 Hz, 2H), 8.06(s, 1H), 7.92 (d, J = 8.9 Hz, 2H), 7.87 (d, J = 5.9 Hz, 2H), 7.60 (s, 2H), 7.58– 7.50 (m, 3H), 7.42 – 7.31 (m, 3H), 7.27 (dd, J= 16.8, 8.1 Hz, 4H), 7.17(dt, J = 15.4, 6.3 Hz, 8H), 7.10 – 7.01 (m, 3H), 6.98 (d, J = 2.9 Hz, 2H), 6.87– 6.80 (m, 2H), 6.35 – 6.26 (m, 1H), 4.34 (s, 1H). 13 C NMR (101 MHz, CDCl3) δ143.67, 139.19, 138.79, 131.16, 130.99, 130.72, 130.44, 130.18, 129.91,128.26, 127.82, 127.16, HRMS (APCl, m / z ) Calcd for C 50 H 38 N + [M+Na] + : 652.3004, found: 652.3008. Example 2 This embodiment provides a method for preparing compound TPBThI. The synthetic route is the same as that in Example 1, except that compound B anthracene-2-ylboronic acid in Example 1 is replaced with an equimolar amount of dibenzo[b,d]thiophene-2-boronic acid. The resulting compounds C and TPBThI have different structures.

[0066] Specifically, the preparation methods of compound TPBThI include: (1) Preparation of compound A: Same as in Example 1.

[0067] (2) Structural formula and preparation method of compound C:

[0068] Compound A (0.93 g, 2 mmol), dibenzo[b,d]thiophene-2-boronic acid (0.54 g, 2.4 mmol), potassium phosphate (0.82 g, 2.4 mmol), and tetrakis(triphenylphosphine)palladium (0.23 g, 0.2 mmol) were dissolved in 10 mL of 1,4-dioxane. The mixture was refluxed at 100 °C for 10 h, cooled to room temperature, and extracted with ethyl acetate. The mixture was washed with water, and the organic layer was collected and dried over anhydrous MgSO4. The solvent was evaporated under reduced pressure, and the product was purified by silica gel column chromatography (mobile phase: ethyl acetate and petroleum ether, v / v) to give compound C, an orange solid, in 70.1% yield.

[0069] The structural characterization data of compound C are as follows: 1 H NMR (400 MHz, Chloroform- d ) δ 10.08 (s, 1H), 8.32 (s, 1H), 8.20(d, J = 5.9 Hz, 1H), 7.94 (s, 2H), 7.92 – 7.88 (m, 3H), 7.67 (d, J = 8.3 Hz, 3H), 7.62 (d, J = 8.1 Hz, 3H), 7.53 (d, J = 8.2 Hz, 4H), 7.47 (d, J = 9.0 Hz, 3H), 7.33 (d, J = 8.0 Hz, 4H), 6.90 (s, 1H), 6.40 (dd, J = 16.0, 5.0 Hz, 2H). 13CNMR (101 MHz, CDCl3) δ 191.97, 149.13, 142.13, 140.61, 139.93, 136.45,136.15, 135.49, 135.20, 133.66, 132.02, 131.75, 131.47, 131.43, 131.35,131.13, 130.45, 129.48, 128.87, 128.28, 128.22, 128.07, 127.64, 127.51,127.47, 127.18, 127.13, 126.93, 126.71, 125.83, 124.49, 123.10, 122.96,121.64, 119.69. HRMS (APCl, m / z ) Calcd for C 41 H 28 OS [M+Na] + : 591.1759, found:591.1760. (3) Structural formula and preparation method of compound TPBThI:

[0070] Compound C (1.14 g, 2 mmol) and 1,4-dimethylpyridine-1-iodide (0.56 g, 2.4 mmol) were dissolved in 28 mL of a tetrahydrofuran / methanol mixture (v / v ratio 1:3). A few drops (e.g., 5 drops) of piperidine were added to the mixture, and the mixture was refluxed at 75 °C for 8 h. The reactants were cooled to room temperature, and the solvent was removed by rotary evaporation under reduced pressure. The product was purified by alumina column chromatography (mobile phase: methanol and dichloromethane, v / v ratio 1:14) to give the red solid compound TPBThI in 43.5% yield.

[0071] The structural characterization data of compound TPBThI are as follows: 1 H NMR (400 MHz, CDCl3) δ 8.90 (s, 1H), 8.83 (s, 1H), 8.23 ​​(s, 1H), 8.18 – 7.84 (m, 2H), 7.80 (s, 2H), 7.57 – 7.47 (m, 3H), 7.40 – 7.32 (m, 4H),7.24 (d, J = 7.5 Hz, 14H), 6.98 – 6.77 (m, 4H), 6.31 (t, J= 14.2 Hz, 1H), 4.40 (s, 1H). 13 C NMR (101 MHz, CDCl3) δ 153.47, 144.79, 144.79, 144.67, 144.66,132.19, 131.95, 131.46, 131.34, 131.17, 128.23, 128.00, 127.66, 127.58,127.45, 127.16, 125.80, 124.52, 124.46, 124.09, 123.84, 123.06, 122.91,121.64, 121.50, 119.63, 44.96. HRMS (APCl, m / z ) Calcd for C 48 H 36 S + [M+Na] + :658.2568, found: 658.2573. Example 3 This embodiment provides a method for preparing compound TPBCaI. The synthetic route is the same as that in Example 1, except that compound B anthracene-2-ylboronic acid in Example 1 is replaced with an equimolar amount of (9-ethyl-9H-carbazole-3-yl)boronic acid. The resulting compounds C and TPBCaI have different structures.

[0072] Specifically, the preparation methods of compound TPBCaI include: (1) Preparation of compound A: Same as in Example 1.

[0073] (2) Structural formula and preparation method of compound C:

[0074] Compound A (0.93 g, 2 mmol), 9-ethyl-3-carbazoleboronic acid (0.57 g, 2.4 mmol), potassium phosphate (0.82 g, 2.4 mmol), and tetrakis(triphenylphosphine)palladium (0.23 g, 0.2 mmol) were dissolved in 10 mL of 1,4-dioxane. The mixture was refluxed at 100 °C for 10 h, cooled to room temperature, and extracted with ethyl acetate. The product was washed with water, and the organic layer was collected and dried over anhydrous MgSO4. The solvent was evaporated under reduced pressure, and the product was purified by silica gel column chromatography (mobile phase: ethyl acetate and petroleum ether, v / v) to give compound C, an orange solid, in 65.5% yield.

[0075] The structural characterization data of compound C are as follows: 1 H NMR (400 MHz, Chloroform- d ) δ 10.09 (s, 1H), 8.30 (s, 2H), 8.12(s, 1H), 7.93 (d, J = 8.1 Hz, 2H), 7.70 (d, J = 9.7 Hz, 2H), 7.62 (s, 3H), 7.55(d, J = 7.7 Hz, 3H), 7.45 (d, J = 8.9 Hz, 5H), 7.34 (s, 5H), 7.07 (s, 2H), 6.91(d, J = 7.0 Hz, 2H), 4.39 (d, J = 7.2 Hz, 2H), 1.45 (s, 3H). 13 C NMR (101 MHz, CDCl3) δ 192.02, 191.96, 141.74, 141.52, 141.06, 140.42, 139.53, 135.58,135.16, 133.99, 133.93, 132.05, 131.78, 131.57, 131.51, 131.47, 131.39,131.17, 130.31, 129.90, 129.50, 128.88, 128.29, 128.21, 128.05, 127.82,127.64, 127.36, 127.32, 127.30, 127.17, 127.11, 126.65, 125.91, 124.91,123.50, 123.05, 120.51, 119.01, 118.69, 108.74, 108.66, 37.69, 13.87. HRMS(APCl, m / z ) Calcd for C 43 H 33 NO [M+Na] + : 602.2460, found: 602.2458. (3) Structural formula and preparation method of compound TPBCaI:

[0076] Compound C (1.16 g, 2 mmol) and 1,4-dimethylpyridine-1-iodide (0.56 g, 2.4 mmol) were dissolved in 28 mL of a tetrahydrofuran / methanol mixture (v / v ratio 1:3). A few drops (e.g., 5 drops) of piperidine were added to the mixture, and the reaction was carried out under reflux at 75 °C for 8 h. The reactants were cooled to room temperature, and the solvent was removed by rotary evaporation under reduced pressure. The product was purified by alumina column chromatography (mobile phase: methanol and dichloromethane, v / v ratio 1:14) to give the red solid product compound TPBCaI in 52.8% yield.

[0077] The structural characterization data of compound TPBCaI are as follows: 1 H NMR (400 MHz, Chloroform-d) δ 8.88 (s, 1H), 8.86 (s, 1H), 8.26 (d, J = 14.5 Hz, 1H), 8.08 (d, J = 7.7 Hz, 1H), 7.95 (d, J = 6.3 Hz, 1H), 7.82 (s,1H), 7.69 (d, J = 21.8 Hz, 2H), 7.60 (d, J = 7.7 Hz, 3H), 7.43 (d, J = 6.5 Hz, 2H), 7.37 (d, J = 8.1 Hz, 4H), 7.30 (d, J = 8.3 Hz, 3H), 7.23 (d, J = 4.4 Hz, 6H), 7.15 (s, 2H), 7.05 (d, J = 3.2 Hz, 3H), 6.97 – 6.84 (m, 3H), 6.35 (d, J =13.4 Hz, 1H), 4.40 (s, 2H), 4.31 (s, 1H), 1.39 (s, 3H). 13C NMR (101 MHz, CDCl3) δ 152.18, 143.76, 141.38, 140.57, 139.28, 139.18, 138.40, 132.42,131.16, 130.93, 130.46, 130.37, 130.19, 129.35, 127.14, 126.96, 126.54,126.21, 126.07, 125.56, 124.91, 123.81, 122.93, 122.68, 122.36, 121.05,119.45, 118.03, 117.47, 107.82, 107.65, 47.12, 36.64, 12.83. HRMS (APCl, m / z )Calcd for C 50 H 41 N2 + [M+Na] + : 669.3270, found: 669.3272. Example 4 This embodiment provides a method for preparing compound TPBCaPF6. The structural formula and preparation method of compound TPBCaPF6 are as follows.

[0078]

[0079] Compound TPBCaI (0.796 g, 1 mmol) was dissolved in 5 mL of dichloromethane to obtain a TPBCaI solution; KPF6 (0.184 g, 1 mmol) was dissolved in the minimum volume of dichloromethane (1 mL) to obtain a KPF6 solution. At room temperature, the KPF6 solution was slowly added to the TPBCaI solution (at a dropping rate of 0.5 mL / min), and the reaction was stirred for 2 h after the addition was complete. After the reaction was complete, the filtrate was collected by filtration; then, the solvent was removed by rotary evaporation under reduced pressure at 40 °C to obtain the red solid product compound TPBCaPF6, with a yield of 92%.

[0080] The structural characterization data of compound TPBCaPF6 are as follows: 1 H NMR (400 MHz, CDCl3) δ 8.88 (s, 1H), 8.85 (s, 1H), 8.27 (s, 1H), 8.10 (s, 2H), 7.96 (s, 2H), 7.72 – 7.67 (m, 2H), 7.64 (dd, J= 7.7, 4.5 Hz, 6H), 7.46 (dd, J = 14.5, 7.5 Hz, 5H), 7.40 (s, 2H), 7.38 (s, 1H), 7.34 (s,1H), 7.31 (s, 3H), 7.11 – 7.06 (m, 4H), 6.99 – 6.94 (m, 3H), 6.44 (s, 1H), 4.41 (s, 2H), 4.35 (s, 1H), 1.42 (s, 3H). 13 C NMR (101 MHz, CDCl3) δ 152.15,143.74, 143.66, 141.40, 140.49, 140.44, 139.32, 139.28, 139.21, 138.43,134.72, 132.50, 131.12, 130.89, 130.46, 130.38, 130.33, 130.18, 127.22,127.18, 126.97, 126.62, 126.56, 126.21, 126.09, 125.96, 125.57, 124.94,123.83, HRMS (APCl, m / z ) Calcd forC 50 H 41 F6N2P [M+H] + : 815.2990, found: 815.2993. Example 5 This embodiment provides a method for preparing compound TPBCaBF4. The structural formula and preparation method of compound TPBCaBF4 are as follows.

[0081]

[0082] Compound TPBCaI (0.796 g, 1 mmol) was dissolved in 5 mL of dichloromethane to obtain a TPBCaI solution; NaBF4 (0.110 g, 1 mmol) was dissolved in the minimum volume of dichloromethane (1 mL) to obtain a NaBF4 solution. At room temperature, the NaBF4 solution was slowly added to the TPBCaI solution (at a dropping rate of 0.5 mL / min), and the reaction was stirred for 2 h after the addition was complete. After the reaction was complete, the filtrate was collected by filtration; then, the solvent was removed by rotary evaporation under reduced pressure at 40 °C to obtain the red solid product compound TPBCaBF4, with a yield of 92%.

[0083] The structural characterization data of compound TPBCaBF4 are as follows: 1 H NMR (400 MHz, CDCl3) δ 8.88 (s, 2H), 8.27 (s, 1H), 8.10 (s, 2H), 7.96 (s, 2H), 7.72 – 7.67 (m, 2H), 7.64 (dd, J = 7.7, 4.5 Hz, 6H), 7.46 (dd, J = 14.5, 7.5 Hz, 5H), 7.40 (s, 2H), 7.38 (s, 1H), 7.34 (s, 2H), 7.31 (s, 2H),7.11 – 7.06 (m, 4H), 6.99 – 6.94 (m, 3H), 6.44 (s, 1H), 4.41 (s, 2H), 4.35(s, 1H), 1.42 (s, 3H). 13 C NMR (101 MHz, CDCl3) δ 152.18, 143.76, 141.38,140.57, 139.28, 139.18, 138.40, 132.42, 131.16, 130.93, 130.46, 130.37,130.19, 129.35, 127.14, 126.96, 126.54, 126.21, 126.07, 125.56, 124.91,123.81, 122.93, 122.68, 122.36, 121.05, 119.45, 118.03, 117.47, 107.82,107.65, 47.12, 36.64, 12.83. HRMS (APCl, m / z ) Calcd for C 50 H 41BF4N2[M+H] + :757.3377, found: 757.3385. Example 6 group This embodiment group provides a method for preparing nanoparticles, including the following steps: Weigh 3 mg of the compound and 30 mg of the amphiphilic copolymer F127 and dissolve them thoroughly in 3 mL of DMSO solution. Then, quickly add the solution to 27 mL of ultrapure water under ultrasonic treatment. Concentrate the solution by centrifugation using an ultrafiltration centrifuge tube to obtain nanoparticles dispersed in ultrapure water.

[0084] Example 6a: The compound was TPBAnI obtained in Example 1, and the nanoparticles obtained were named TPBAnINPs; Example 6b: The compound is TPBThI obtained in Example 2, and the obtained nanoparticles are named TPBThINPs; Example 6c: The compound is TPBCaI obtained in Example 3, and the obtained nanoparticles are named TPBCaINPs; Example 6d: The compound was TPBCaPF6 obtained in Example 4, and the nanoparticles obtained were named TPBCaPF6NPs. Example 6e: The compound is TPBCaBF4, which was prepared in Example 5, and the nanoparticles obtained are named TPBCaBF4NPs.

[0085] Experimental Example 1 Optical measurement of matter (1) Photophysical property testing of compounds: UV-Vis absorption and fluorescence emission spectroscopy tests: First, weigh out a certain mass of TPBNaI, TPBThI, TPBCaI, TPBCaPF6, and TPBCaBF4 solid powders and dissolve them in an appropriate amount of DMSO to prepare a concentration of 10. -3 For each mol / L stock solution, take 30 μL of the stock solution and add 2970 μL of DMSO, mix well to prepare a 10 mol / L solution. -5 The working solutions were prepared at mol / L, and the UV absorption and fluorescence emission spectra of each working solution were measured. The UV-Vis absorption spectra and normalized fluorescence emission spectra of TPBNaI, TPBThI, TPBCaI, TPBCaPF6, and TPBCaBF4 in DMSO were measured as follows: Figure 1As shown in the figure, the maximum absorption wavelengths of these compound molecules are in the range of 400 nm to 500 nm. In DMSO, the normalized maximum emission wavelengths are 570 nm (TPBAnI), 605 nm (TPBCaI), 679 nm (TPBThI), 607 nm (TPBCaPF6), and 587 nm (TPBCaBF4).

[0086] Solid-state fluorescence spectroscopy: TPBNaI, TPBThI, and TPBCaI solid powders were evenly spread in a solid test cell to measure solid-state fluorescence spectra. Figure 2 As shown. Due to the typical D-π-A structure, the emission peak in the solid state is located at 813 nm, with the wavelength tail extending to 1000 nm, covering the near-infrared II region.

[0087] AIE property test: Take the above concentration as 10 -3 The stock solution of mol / L was diluted 10 times with DMSO to prepare a solution with a concentration of 1×10⁻⁶. -4 A solution of mol / L; then a concentration of 1×10 -4 A solution of mol / L, DMSO, and water were mixed in different proportions to prepare solutions with water contents of 0%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, and 90%, respectively, with a total volume of 3 mL and a concentration of 1×10⁻⁶. - 5 Fluorescence spectroscopy was performed on the mixed solution of mol / L, and the test results are as follows: Figure 3 As shown in the figure, I / I0 refers to the ratio of the fluorescence intensity of the mixed solution with the corresponding water content to the initial fluorescence intensity (the fluorescence intensity of the mixed solution with 0% water content). The figure shows that when the water content is below 50%, these three compounds show almost no fluorescence signal. With increasing water content, the fluorescence intensity rises sharply and then decreases slowly, exhibiting typical aggregation-induced emission enhancement (AIEE) characteristics.

[0088] Photothermal performance testing: First, a certain amount of TPBNaI, TPBThI, TPBCaI, TPBCaPF6, and TPBCaBF4 solid powders were weighed and dissolved in DMSO to prepare 1 mM stock solutions. Then, the stock solutions were diluted with DMSO to prepare 200 μM working solutions. Next, 0.5 mL of each of the above working solutions (0.5 mL of DMSO as a control) was placed in a 1.5 mL conical centrifuge tube and centrifuged at 470 nm (350 mW / cm²). 2 The sample was irradiated with a laser for 8 minutes continuously. Temperature was precisely recorded every 30 seconds using an infrared thermal imager during the irradiation period. The test results are as follows: Figure 4As shown in the figure, these compounds all exhibit excellent photothermal conversion capabilities due to the TPB unit and methylpyridinium. After 8 minutes of laser irradiation, the temperatures of the three compounds increased to 49.0℃ (TPBAnI), 53.3℃ (TPBThI), and 58.4℃ (TPBCaI), respectively; TPBcaPF6 and TPBcaBF4 exhibited similar photophysical properties and photothermal therapeutic performance to TPBcaI.

[0089] (2) Photophysical property testing of nanoparticles: Size distribution testing: The particle size distribution of the five nanoparticles prepared in Example 6 was measured by dynamic light scattering (DLS), and the results are shown in Table 1. Taking TPBCaI NPs as an example, the size distribution of TPBCaI NPs measured by DLS is shown in the figure below. Figure 5 As shown.

[0090] UV-Vis absorption and fluorescence emission spectroscopy tests: First, the concentrations of the corresponding compounds (TPBAnI, TPBThI, TPBCaI, TPBCaPF6, and TPBCaBF4) in the nanoparticles were obtained using a UV standard curve; then, appropriate amounts of water were added to prepare concentrations of 10... -3 30 μL of the stock solution (based on the compounds contained therein; the concentration of nanoparticles mentioned thereafter is based on the content of those compounds) was added to 2970 μL of water to prepare a 10 mol / L solution. -5 The working solutions were prepared at mol / L, and the UV absorption and fluorescence emission spectra of each working solution were measured. The UV-Vis absorption spectra and normalized fluorescence emission spectra of the obtained TPBaI NPs, TPBThI NPs, TPBcaI NPs, TPBcaPF6 NPs, and TPBcaBF4 NPs in water are shown below. Figure 6 As shown.

[0091] Photothermal performance testing: Referring to the photothermal performance testing methods of the above compounds, the temperature changes of TPBaI NPs, TPBThI NPs, TPBCaI NPs, TPBCaPF6 NPs, and TPBCaBF4 NPs under different irradiation times are as follows: Figure 7 As shown; infrared thermal images at different irradiation times, as follows Figure 8 As shown in the figure, the temperature of the solution containing each nanoparticle rapidly increased within 10 minutes, with the plateau temperature ranging from 47.5℃ to 57.3℃ (the specific temperatures after the increase are shown in Table 1); and the excellent photothermal conversion performance can be visually observed from the infrared thermal images. Among them, TPBCaI NPs exhibited the most significant photothermal therapeutic potential.

[0092] Subsequently, the photothermal stability and photothermal conversion efficiency (η) of each nanoparticle were tested, with TPBCaI NPs as an example for specific description. The test results are as follows: Figure 9 As shown in (a) and (b), TPBCaI NPs exhibited a stable temperature change during the four heating and cooling cycles, with minimal temperature variation at the highest increase. The η of TPBCaI NPs was further calculated to be approximately 53.07%. Detailed data for the other four nanoparticles are shown in Table 1.

[0093] The photothermal stability test included: diluting the nanoparticle mother liquor with water to prepare a 200 μM working solution (based on the compounds contained therein); taking 0.5 mL of the above working solution into a 1.5 mL conical centrifuge tube and placing it at 470 nm (350 mW / cm²). 2 Irradiate the working fluid with a laser for 5 minutes continuously; record the temperature rise of the photothermal conversion agent under illumination to obtain a temperature rise curve; turn off the light source and allow the working fluid to cool to room temperature under natural conditions, recording the temperature drop process to obtain a cooling curve. Repeat the above irradiation and cooling steps a total of 4 times, recording the complete temperature change process each time, and plot the temperature change curve.

[0094] Calculation of photothermal conversion efficiency:

[0095] In the formula, I is the laser power, A is the absorbance of the 200 μM working fluid at 470 nm, and ΔT and ΔT water Temperature changes were represented for the 200 μM working solution and the blank sample water, respectively. The heat transfer coefficient is denoted by h, and s represents the surface area of ​​the container. These parameters were determined using the following formula:

[0096] m is the mass of the solution (approximately 1 g), and c is the specific heat capacity of the solvent (4.2 J·g for water). -1 ·℃ -1 ), τ s The time constant can be determined over the cooling cycle by the following formula:

[0097] θ is a dimensionless parameter that varies with time, called the driving force temperature, and is defined as follows:

[0098] T max and T surr The maximum temperature and room temperature of the 200 μM working solution are respectively.

[0099] Table 1. Test results of different nanoparticles

[0100] Experimental Example 2 Therapeutic effects of TPBCaI NPs at the cellular level Cell culture: 3T3 cells (mouse embryonic fibroblast cell line) were obtained from the National Cell Bank (NICR) and cultured in Dubick modified Eagle medium (DMEM) containing 10% fetal bovine serum and 2% penicillin-streptomycin. The culture conditions were carried out in a constant temperature incubator at 37°C and 5% CO2.

[0101] (1) Cytotoxicity test: 3T3 cells were subjected to 5×10 3 Cells were seeded at a density of 100 cells / well in 96-well plates and incubated for 24 h. Cells were then incubated with different concentrations of TPBCaI NPs in fresh medium. After a further 24 h of incubation, the medium was removed and the cells were washed three times with PBS. Cells were then incubated in the dark with fresh serum-free medium containing 10% MTT for 4 h. All medium was then removed and 150 µL of DMSO was added. Finally, the absorbance of the products was measured at 570 nm using a microplate reader. The results are expressed as the percentage of cell survival after different treatments relative to untreated control cells.

[0102] Cell phototoxicity assay: Culture cells as described above. After cell adhesion, discard the original culture medium and select the maximum effective concentration of TPBCaI NPs (e.g., 50 μM) under conditions of high cell viability for phototoxicity testing. Randomly divide the wells of the 96-well plate into four groups: 1) Control group (containing only culture medium); 2) Light control group (culture medium + white light irradiation); 3) TPBCaI NPs group (containing TPBCaI NPs, no light irradiation); 4) TPBCaI NPs + light group (containing TPBCaI NPs + white light irradiation). The no-light groups (control group and TPBCaI NPs group) were completely protected from light by wrapping them in aluminum foil; the light groups (light control group and TPBCaI NPs + light group) were exposed to white light (5 mW·cm²). -2 Irradiate for 10 min. After irradiation, all groups were incubated in a 37℃, 5% CO2 incubator for 24 h. After the culture was completed, the relative cell viability was calculated using the same procedure as above.

[0103] The cytotoxicity of different concentrations (final concentrations) of TPBCaI NPs on 3T3 cells and the phototoxicity of 50 μM TPBCaINPs on 3T3 cells are as follows: Figure 10As shown in (a) and (b) in the figure, TPBCaI NPs exhibit good biocompatibility in 3T3 cells. At a concentration as high as 50 μM, the cell viability is as high as 83%, indicating that the cytotoxic effect is negligible.

[0104] (2) Cell colocalization imaging: 3T3 cells were seeded in Ф 20 mm glass-bottomed cell culture dishes (1.0 ± 0.05 × 10⁶ cells per dish). 6 After culturing overnight in a humidified incubator at 37°C and 5% CO2, the culture medium was removed, and cells were stained with TPB CaI NPs (50 µM) for 15 min. After washing three times with PBS, 3T3 cells were fixed with 4% fixative solution for 10 min. Before imaging, each dish was washed three times with PBS. To co-localize with Mito-Tracker Green, the fixed cells were stained with Mito-Tracker Green (50 nM) at 37°C for 30 min. Mito-Tracker Green (mitochondria), TPB CaI NPs, and combined confocal microscopy images are shown below. Figure 11 As shown (where green channel 500–550 nm, red channel 550–600 nm). Cellular colocalization imaging revealed that TPBCaI NPs exhibit mitochondrial targeting. In vitro images elucidated that, due to the highly negative mitochondrial membrane potential, TPBCaI NPs were extensively internalized into the mitochondria, with a Pearson correlation coefficient of 0.87. Mitochondrial targeting is crucial for maximizing the efficacy of PTT in treating obesity, as mitochondria are essential for ATP production and fat storage. Simultaneously, photothermal agents can kill cells under radiation conditions.

[0105] (3) Live / dead cell (Calcein-AM / PI) staining: First, 3T3 cells were seeded and cultured in glass-bottomed culture dishes for 24 h, and then subjected to the following different treatments: blank control group (culture medium only), light-only group (culture medium + white light irradiation 5 mW·cm). -2 ), TPBCaI NPs only (50 μM), TPBCaI NPs + light irradiation group (50 μM TPBCaI NPs + white light irradiation 5 mW·cm), -2The cells were irradiated with white light for 10 min. Then, they were stained sequentially with calcein-AM in PBS for 20 min and PI in PBS for 5 min. Afterward, the cells were gently washed and imaged using a confocal laser scanning microscope. Conditions: Excitation wavelength: calcein-AM 488 nm, PI 543 nm; Emission filter: calcein-AM 500–550 nm, PI 550–650 nm. The confocal laser scanning microscope fluorescence image of 3T3 cells stained with calcein-AM / PI is shown below. Figure 12 As shown in the figure, approximately 71% of 3T3 cells were at 5 mW·cm⁻¹. -2 The cells were killed after treatment with TPBCaI NPs for 10 min under white light irradiation. In the calcein-AM / PI staining experiment, the strong red fluorescence signal indicated excellent photothermal conversion efficiency and cytotoxicity.

[0106] (4) Adipogenic differentiation of 3T3 cells: 3T3-L1 adipogenic differentiation induction medium solutions A and B were prepared according to the instructions for 3T3-L1 (mouse embryonic fibroblast) adipogenic differentiation induction medium from Pronosei for subsequent use. When the confluence of 3T3 cells reached 80%–90%, they were digested with 0.25% trypsin. The digested 3T3 cells were counted, and the results were used to determine the cell count (2–3 × 10⁶ cells / year). 4 3T3 cells were seeded in six-well plates, with 2 mL of complete culture medium added to each well. The uniformly seeded 3T3 cells were then incubated at 37°C with 5% CO2. When cell confluence reached 100%, the complete culture medium was carefully aspirated from the wells, and 2 mL of adipogenic differentiation medium A solution was added to each well. After induction with solution A for 2-3 days, the complete induction medium was aspirated from the six-well plates, and 2 mL of complete adipogenic differentiation medium B solution was added to each well to maintain induction for 1 day. After alternating induction with medium A and medium B 3-5 times, when obvious and sufficient lipid droplets were observed in the stem cells, the cells could be cultured in solution B for 3-6 days (changing the medium every 2-3 days) until the lipid droplets became sufficiently large and abundant. Induction could then be terminated, and the cells could be processed according to experimental requirements. A schematic diagram of 3T3 cell adipogenic differentiation is shown below. Figure 13 As shown in (a), successfully differentiated adipocytes are observed using a microscope.

[0107] Oil Red O staining: Remove the induction differentiation medium from the culture dish and rinse 1-2 times with PBS. Add 4% neutral formaldehyde solution (just enough to cover the cell surface) and fix the cells for 15 min. During cell fixation, prepare the Oil Red O working solution (saturated Oil Red O solution: distilled water = 3:2, mix thoroughly and filter through neutral filter paper to remove impurities). Remove the 4% neutral formaldehyde solution and rinse twice with PBS to remove background impurities. Using a Φ20 mm glass-bottomed cell culture dish as an example: Add 1 mL of Oil Red O working solution to each well and stain at room temperature for 30 min. Aspirate the Oil Red O working solution, rinse 1-2 times with PBS to remove background impurities, and then observe the induction and staining effects under a microscope. The experiment was set up with the following 4 groups: blank control group (medium only), light-only group (medium + white light irradiation 5 mW·cm²), etc. -2 ), TPBCaI NPs only (50 μM), TPBCaI NPs + light irradiation group (50 μM TPBCaI NPs + white light irradiation 5 mW·cm), -2 Oil Red O staining results of intracellular lipid droplets after treatment under different conditions are shown below. Figure 13 As shown in (b).

[0108] Intracellular calcium ion concentration test: The experiment was set up with the following 4 groups: 1) Blank control group (culture medium only), 2) Light-only group (culture medium + white light irradiation 5 mW·cm). -2 ), 3) TPBCaI NPs only (50 μM), 4) TPBCaI NPs + light irradiation group (50 μM TPBCaI NPs + white light irradiation 5 mW·cm -2 The specific experimental steps are as follows: The culture medium was aspirated, and the cells were fixed with 4% paraformaldehyde for 15 min at room temperature, then washed three times with PBS. TPBCaI NPs were diluted to 10 μM with DMEM medium. Groups 3) and 4) were incubated with medium containing 50 μM TPBCaI NPs for 30 min. Then they were washed three times with PBS. Groups requiring light were exposed to white light (5 mW·cm²). -2 The cells were incubated for 10 min in one group, while the other groups were kept in the dark. Fluo-3 AM was diluted to 3 μM, and 1 mL of diluted Fluo-3 AM was added to each of the four groups of cells. The cells and dye were incubated in an incubator for 30 min, and then washed twice with PBS. After washing, the cells were incubated for another 20 min to ensure that Fluo-3 AM was completely converted to Fluo-3 in the cells. 1 mL of PBS was added to each culture dish, and the fluorescence signal was immediately detected using a Zeiss LSM 980. Fluorescence images of intracellular calcium ion concentration after treatment under different conditions are shown below. Figure 13 As shown in (c).

[0109] UCP1 staining: As described above, four groups were set up for the experiment. The specific experimental steps are as follows: The culture medium in the successfully induced adipocytes was aspirated, and the cells were fixed with 4% paraformaldehyde at room temperature for 15 min, then washed three times with PBS. TPBCaINPs were diluted to 50 μM with DMEM medium. Groups 3) and 4) were incubated with medium containing 50 μM TPBCaINPs for 30 min, and then washed three times with PBS. The groups requiring illumination were irradiated with white light (5 mW·cm²). - ², 10 min), the remaining groups were treated in the dark. Add 1 mL of blocking buffer and block for 30 min, then add 500 μL of UCP1 antibody and incubate at room temperature in the dark for 1 h, followed by washing three times with PBS. Add 1 mL of secondary antibody and incubate at room temperature in the dark for 1 h, followed by washing three times with PBS. Finally, stain with DAPI for 10 min, wash three times with PBS, add 1 mL of PBS buffer to each dish, and immediately detect the fluorescence signal using a confocal laser scanning microscope. Immunofluorescence images of UCP1-labeled (green) after treatment under different conditions are shown below. Figure 13 As shown in (d), the cell nucleus is stained blue.

[0110] ATP concentration test: (1) Preparation of standard curve determination: Thaw the reagents to be used in an ice bath, and dilute the ATP standard solution with ATP detection lysis buffer to a concentration gradient of 0.01, 0.025, 0.05, 0.1, 0.25, 0.5, 1, 2.5, 5 and 10 μM. (2) Sample preparation: The experiment was set up with the following 4 groups: 1) Blank control group (culture medium only), 2) Light only group (culture medium + white light irradiation 5 mW·cm). -2 ), 3) TPBCaI NPs only (50 μM), 4) TPBCaI NPs + light irradiation group (50 μM TPBCaI NPs + white light irradiation 5mW·cm -2 Groups 3) and 4) were incubated with medium containing 50 μM TPBCaI NPs for 30 min. They were then washed three times with PBS. Groups requiring light were exposed to white light (5 mW·cm²). -2, 10 min), while the other groups were kept in the dark. After treatment, the culture medium was removed, and 200 μL of lysis buffer was added to each group to lyse the cells. After lysis, centrifuge at 4℃ and 12000 g for 5 min, and take the supernatant for subsequent determination. (3) Preparation of ATP detection working solution: Prepare an appropriate amount of ATP detection working solution according to the ratio of 100 μL of ATP detection working solution required for each sample or standard. Thaw the reagent to be used on an ice bath. Take an appropriate amount of ATP detection reagent and dilute the ATP detection reagent with ATP detection reagent diluent at a ratio of 1:4. The diluted ATP detection reagent is the ATP detection working solution for subsequent experiments. The ATP detection working solution can be temporarily stored on an ice bath. (4) Determination of ATP concentration: Add 100 μL of ATP detection working solution to the detection well or detection tube. Place at room temperature for 3-5 min to allow all the background ATP to be consumed, thereby reducing the background. Add 20 μL of sample or standard to the detection well or tube, mix quickly with a pipette, and measure the RLU value using a microplate reader after at least 2 seconds. Intracellular ATP levels after treatment under different conditions are as follows: Figure 13 As shown in (e) in the diagram.

[0111] The test results above show that under the influence of TPBCaI NPs+Light, the cell morphology of white adipocytes changed significantly and gradually transformed into brown adipocytes. According to... Figure 13 As shown in (c), the intracellular calcium concentration in the TPBCaI NPs + Light group was increased, demonstrating TRPV1 channel activation. According to... Figure 13 As shown in (d), the brown adipocyte-specific mitochondrial protein UCP1, responsible for thermogenic respiration, was highly expressed after TPBCaI NPs + Light, indicating that white adipocytes transform into brown-like adipocytes via the TRPV1-mediated pathway. In summary, the PTT of TPBCaI NPs can induce browning in white adipocytes, which can be confirmed by intracellular calcium and UCP1 concentrations. As previously mentioned, UCP1 can release ATP produced by the mitochondrial respiratory chain as heat energy, thereby preventing its storage and conversion into fat, thus achieving a fat-reducing effect. In UCP1-rich brown adipocytes, intracellular ATP levels decrease sharply, such as... Figure 13 As shown in (e).

[0112] Experimental Example 3 The therapeutic effect of TPBCaI NPs on obesity in live mice I. Experimental Methods Animal culture: BALB / c (male, 5 weeks old) mice were purchased from Chengdu Dashuo Experimental Animal Co., Ltd. Housing conditions: All animals were kept under controlled temperature (22–26 °C), humidity (50%–60%), and light (12 h light / 12 h dark, 15–20 LX).

[0113] Establishment of a mouse obesity model: BALB / c mice were used as experimental subjects. A high-fat diet was used to induce obesity by feeding the mice a high-fat diet, causing their energy intake to far exceed their energy expenditure, resulting in fat accumulation and weight gain. A suitable high-fat diet was selected, generally with a fat content of around 40% to 60%, and the carbohydrate and protein content were adjusted accordingly. The mice were fed this diet continuously for more than two weeks. Their weight was recorded daily and compared with that of mice in a normal-fed group to calculate their Lee's index.

[0114] After successfully establishing the obesity model, mice were randomly divided into 4 groups (n=5 / group): blank control group (PBS): no treatment; light-only group (PBS+Light): 100 μL PBS was injected into the inguinal fat area, and the inguinal fat area was irradiated with white light (200 mW·cm). - ², 10 min); Drug-only group (TPBCaI NPs): 100 μL of TPBCaI NPs diluted in PBS (50 μM concentration) was injected into the inguinal fat region, and the solution was protected from light (wrapped in aluminum foil); Drug-only + Light group (TPBCaI NPs + Light): 100 μL of TPBCaI NPs solution diluted in PBS (50 μM concentration) was injected into the inguinal fat region, and the solution was irradiated under the same conditions as the light-only group. Mouse weight was recorded daily for 14 days.

[0115] II. Results Analysis The results are as follows Figures 14 to 16 As shown: To further verify the therapeutic effect of PTT on obesity in vivo, an obesity model was constructed using a high-fat diet (HFD) for 2 weeks. According to... Figure 14 As shown in (a) and (b), the weight and Lee's index of mice fed HFD were significantly higher than those in the normal group, indicating the development of obesity-related metabolic syndrome. Figure 14 The photographs of iWAT and eWAT in (c) and (d) illustrate the development of obesity-related metabolic syndrome. To avoid burning other tissues, especially the skin, TPBCaI NPs were injected into the iWAT using an insulin needle. Figure 14As shown in (e), a large number of TPBCaI NPs were observed to remain on iWAT 1 h after injection, providing sufficient time for PTT-based obesity treatment. After 24 h, both major organ and adipose tissue signals were negative, indicating that TPBCaI NPs have good biocompatibility and are easily metabolized.

[0116] Subsequently, different treatments were applied to the obese mouse model. Schematic diagrams of the obese mouse model and photothermal treatment are shown below. Figure 15 As shown in (a). After treatment with PBS, light alone, TPBCaI NPs, and TPBCaI NPs + Light, respectively, the photographs and weight data of the mice demonstrated that TPBCaI NPs + Light completely inhibited the development of obesity, as shown in (a). Figure 15 As shown in (b) and (c). During the treatment period, mice treated with PBS gained 10.2% body weight, while mice treated with TPBCaI NPs + Light lost 7.9% body weight. Furthermore, fat sampling analysis and histological analysis were performed. Both iWAT and eWAT showed significant shrinkage after treatment, which can be directly observed in… Figure 14 As observed in (d), the percentage of adipose tissue to body weight is an important indicator. Compared to a single body weight indicator, the ratio of fat mass to body weight in mice provides a more intuitive reflection of obesity and health status. Figure 14 As shown in (e), after PTT treatment, both "iWAT / weight" and "eWAT / weight" showed significant improvement, decreasing by 4.6% and 37.6%, respectively.

[0117] The anti-obesity effect was also assessed through blood lipid tests. Relatively high levels of serum triglycerides (TG), total cholesterol (TC), and low-density lipoprotein cholesterol (LDL-C) can lead to obesity, arteriosclerosis, and other diseases. Conversely, high-density lipoprotein cholesterol (HDL-C) transports cholesterol to the liver for metabolism and excretion. Its plasma levels are negatively correlated with the risk of cardiovascular disease. Figure 16 As shown in (a) to (c), during the treatment, serum TG, TC, and LDL-C levels were effectively improved, cured, and reduced by 36.2%, 10.4%, and 15.6%, respectively. Meanwhile, the TPBCaI NPs + Light group was the most effective in increasing serum HDL-C levels, with significantly higher effects than other groups. Figure 16 (d) After browning, BAT adipocytes consume large amounts of free fatty acids (FFA) to generate heat, leading to a decrease in FFA levels, such as... Figure 16 (e) The reduction in WAT quality after PTT treatment can also improve systemic insulin sensitivity, allowing for better glucose absorption and utilization, thereby providing a feedback reduction in serum insulin secretion, such as... Figure 16(f) in the middle. Finally, Figure 16 The H&E staining images of iWAT in (g) depict mice receiving TPBCaI NPs + Light that experienced a higher degree of adipocyte browning compared to the other three groups.

[0118] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention, and not to limit them; although the present invention has been described in detail with reference to the foregoing embodiments, those skilled in the art should understand that modifications can still be made to the technical solutions described in the foregoing embodiments, or equivalent substitutions can be made to some or all of the technical features; and these modifications or substitutions do not cause the essence of the corresponding technical solutions to deviate from the scope of the technical solutions of the embodiments of the present invention.

Claims

1. A compound, characterized in that, It has the structure shown in Equation I: ； Wherein, R is selected from at least one of anthracene, dibenzothiophene, or carbazole groups, and X is selected from halogen atoms, PF6 - and BF4 - At least one of them.

2. The compound according to claim 1, characterized in that, R is selected from , and Any one of them.

3. The compound according to claim 1, characterized in that, X is selected from I - PF6 - and BF4 - At least one of them.

4. The compound according to claim 1, characterized in that, The compound has the following structural formula: 、 、 、 、 。 5. A method for preparing the compound according to any one of claims 1 to 4, characterized in that, Includes the following steps: (a) Compound A and compound B undergo a Suzuki coupling reaction in a solvent to give compound C; (b) Compound C reacts with 1,4-dimethylpyridine-1-iodide in a solvent to give the compound shown in Formula I1; The structural formulas of compounds A, B, C, and I1 are as follows: 、 、 、 。 6. The preparation method according to claim 5, characterized in that, When X is PF6 - Or BF4 - The preparation method further includes reacting compound I1 with KPF6 or NaBF4 in a solvent to obtain a compound as shown in Formula I.

7. A photothermal agent, characterized in that, Includes the compounds described in any one of claims 1 to 4.

8. The photothermal agent according to claim 7, characterized in that, The photothermal agent includes compounds and amphiphilic copolymers; Preferably, the amphiphilic copolymer includes amphiphilic copolymer F127; Preferably, the mass ratio of the compound to the amphiphilic copolymer is 1:(5-15).

9. The use of the compound according to any one of claims 1 to 4 or the photothermal agent according to any one of claims 7 to 8 in the preparation of medical imaging agents, optical imaging agents or staining agents.

10. The use of the compound according to any one of claims 1 to 4 or the photothermal agent according to any one of claims 7 to 8 in the preparation of a medicament for the treatment of obesity.

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