Near-infrared second near-infrared region luminescent compounds, aggregates thereof, preparation methods and applications thereof

Abstract

The present invention belongs to the field of biomedical materials, and discloses a class of organic near-infrared second near-infrared region (NIR-II) luminescent materials based on pyrrolo[2,3-b:4,5-b']diindole derivatives, a preparation method thereof, and an application thereof in kidney transplantation surgery. In the present invention, a nine-ring pyrrole derivative is used as an electron-donating group, and 2-(5,6-difluoro-3-oxo-2,3-dihydro-1H-inden-1-ylidene) malononitrile with strong electron-withdrawing ability is combined to design an organic NIR-II molecule with an A-D-A structure. Then, through simple aggregation, an AIE aggregate DIPT-ICF Aggs with both NIR-II absorption and emission is obtained. The present invention exhibits excellent transplanted kidney angiography and transplanted kidney reperfusion imaging capabilities in kidney transplantation surgery, has a wide application prospect in the field of biomedicine, and is expected to be widely applied in the field of fluorescence imaging for surgical operations.

Description

Technical Field

[0001] The present invention belongs to the field of biomedical materials, and particularly relates to a preparation method of a near-infrared second-region luminescent aggregate contrast agent and its application in kidney transplantation surgery. Background Art

[0002] The incidence of end-stage renal disease (ESRD) is increasing year by year globally, and it has poor prognosis and high cost, which has become a serious public health problem endangering human health and increasing the economic burden globally. Kidney transplantation is the preferred treatment method for ESRD patients. Compared with hemodialysis and peritoneal dialysis, kidney transplantation patients have higher survival rates and better quality of life. The success of kidney transplantation surgery mainly depends on the patency after vascular anastomosis and the quality of renal reperfusion. Currently, methods for evaluating intraoperative vascular anastomosis and renal reperfusion quality include angiography, color Doppler ultrasound, and renal tissue oxygenation measurement, etc. However, these techniques all have certain defects, such as the nephrotoxicity of angiography contrast agents, X-ray exposure, the limited area of ultrasound detection, and they cannot display renal vascular anastomosis and renal perfusion in real time and accurately. Therefore, safer and more sensitive techniques are needed to achieve real-time monitoring of kidney transplantation surgery.

[0003] Due to the characteristics of real-time and high resolution of near-infrared second-region (NIR-II) fluorescence imaging technology, it has been increasingly favored by surgeons. However, currently approved NIR fluorescence contrast agents by the FDA, indocyanine green (ICG) and methylene blue (MB), not only have the phenomenon of aggregation-caused fluorescence quenching (ACQ), but also have disadvantages such as poor stability, low brightness, and short circulation time (short best imaging window period), and cannot display anatomical structures with high quality throughout the surgery, so they are limited in clinical use.

[0004] Contrary to aggregation-caused fluorescence quenching (ACQ), NIR-II fluorescence contrast agents with aggregation-induced emission (AIE) characteristics have weakened brightness in the dispersed state, but increased brightness and stability in the aggregated state. Although there are many reports on contrast agents with AIE characteristics and NIR-II emission currently, there are few reports on AIE aggregate contrast agents with both NIR-II absorption and emission. Therefore, in the present invention, an AIE molecule with NIR-II emission was designed and synthesized, and an aggregate contrast agent with both NIR-II absorption and emission was prepared by further aggregation. Summary of the Invention

[0005] The primary object of the present invention is to provide a novel AIE aggregate contrast agent with both NIR-II absorption and emission.

[0006] Another object of the present invention is to provide a preparation method of the above novel AIE aggregate contrast agent and its application in kidney transplantation surgery.

[0007] The object of the present invention is achieved based on the following technical solutions: When designing, it is particularly important to select a suitable central donor (D) unit. The stronger the electron-donating ability of the central D unit, the wider the absorption range of the constructed small molecule, and thus the more likely it is to obtain high-performance. Therefore, pyrrolo[2,3-b:4,5-b']diindole (DIP) is selected as the D unit, and fluorinated and non-fluorinated 2-(3-oxo-2,3-dihydro-1H-inden-1-ylidene)malononitrile (IC and ICF) are used as the A unit to construct a series of A-D-A type small molecule acceptors or donor materials based on N,S-heterocycles. With the aid of surfactants, further improvement of photophysical properties and good dispersibility in aqueous solutions are achieved through aggregation.

[0008] The present invention has at least the following beneficial effects:

[0009] 1. The NIR-II fluorescence contrast agent described in the present invention has a long retention time in blood circulation, high imaging resolution, large tissue penetration depth, long optimal imaging window period, good stability, and good biocompatibility. Currently, the fluorescence contrast agents approved by the FDA for clinical use have a fast excretion rate, low imaging resolution, and poor stability, which are not conducive to long-term surgical navigation;

[0010] 2. The NIR-II aggregate fluorescence contrast agent described in the present invention is simple to prepare. Description of the Drawings

[0011] Figure 1 Shown are the synthetic routes of DIPT-IC and DIPT-ICF.

[0012] Figure 2 Shown is the synthetic route of DIPT-ICF Aggs.

[0013] Figure 3 (a) and (b) are the absorption spectra and emission spectra of DIPT-IC and DIPT-ICF in THF, respectively; (c) and (d) are the emission spectra of DIPT-IC and DIPT-ICF in DMF / H2O mixtures with different f w values; (e) is the curve of the emission intensity versus the water content in the DMF / H2O mixture.

[0014] Figure 4 (a) is the molar absorbance of DIPT-IC and DIPT-ICF; (b) is the fluorescence spectra of DIPT-IC, DIPT-ICF, and IR-26 at five different concentrations; (C) is the quantification curve of the molar absorbance, quantum yield, and brightness of DIPT-IC and DIPT-ICF.

[0015] Figure 5Characterization of DIPT-IC and DIPT-ICF Aggs: (a) Absorption and emission spectra of DIPT-ICF Aggs in deionized water; (b) Absorption spectra of DIPT-IC, DIPT-ICF and DIPT-ICF Aggs.

[0016] Figure 6 (a) Dynamic light scattering (DLS) data and transmission electron microscopy (TEM) images (scale bar in the inset is 50 nm); (b) Particle size stability of DIPT-ICF Aggs in deionized water at different pH values and different times.

[0017] Figure 7 Toxicity analysis of DIPT-ICF Aggs on HeLa and 3T3 cells at different concentrations.

[0018] Figure 8 DIPT-ICF Aggs were intravenously injected into Balb / c nude mice (a) and New Zealand white rabbits (b). After 2 weeks, hematological examinations (blood cell and blood biochemical tests) were performed, and there were no significant differences in various indicators between the experimental group and the control group.

[0019] Figure 9 DIPT-ICF Aggs were intravenously injected into Balb / c nude mice (a) and New Zealand white rabbits (b). After 2 weeks, important organs (heart, liver, spleen, lung, kidney, intestine, skin) of the nude mice and rabbits were removed for HE staining; according to the experimental results, no organic damage to the important organs of mice and rabbits was found caused by DIPT-ICF Aggs.

[0020] Figure 10 (a) NIR-I / II images of capillary glass tubes containing DIPT-ICF Aggs saline solution. The arrows correspond to the positions and directions of the cross-sectional fluorescence intensity distribution maps in the NIR-I and NIR-II regions. Chicken breast tissue with different thicknesses (1.0 mm, 2.0 mm, 3.0 mm, 4.0 mm and 5.0 mm) was covered on the capillary glass tubes; (b) and (c) respectively show that the NIR-I / II fluorescence signal intensity decreases with the increase in the thickness of the chicken breast tissue covering the capillary glass tubes; (d) shows that there is a statistically significant difference in the SBR between NIR-II and NIR-I imaging at depths within 3 mm of each chicken breast (P < 0.05).

[0021] Figure 11(a) NIR-II angiography of the subcutaneous blood vessels in the abdominal wall of mice was performed under the conditions of an 808 nm laser - 900 nm long-pass filter (LP900 nm), an 808 nm laser - 1319 nm long-pass filter (LP1319 nm), and a 980 nm laser - 1319 nm long-pass filter, respectively; (b) NIR-II angiography of the subcutaneous blood vessels in the abdomen of mice was performed under the conditions of a 980 nm laser combined with LP1020 nm, LP1100 nm, and LP1319 nm filters; (c, d) Under different laser and filter conditions, there were significant differences in the SBR of NIR-II angiography; paired t-tests were used to calculate the significance, **p < 0.01, *p < 0.05.

[0022] Figure 12 Shows the retention time of DIPT-ICF Aggs in the blood circulation of mice: (a) The NIR-II fluorescent contrast agent DIPT-ICF Aggs was injected into mice via the tail vein. Serum was collected at different time points after administration (0 h, 0.5 h, 1 h, 2 h, 4 h, 8 h, 24 h, 48 h, 72 h, 96 h, 120 h, and 144 h) and placed in a 96-well plate for NIR-II imaging; (b) The near-infrared signal intensity distribution curve of DIPT-ICF Aggs in (a); (c) The quantitative analysis curve of the near-infrared-average signal intensity of DIPT-ICF Aggs in (a); (d) Angiography of the subcutaneous blood vessels in the abdomen of mice using a 980 nm laser - long-pass 1319 nm filter; (e) The NIR-II angiography signal intensity curve (dotted line marked) of the subcutaneous blood vessels in the abdomen of mice in (d); (f) The signal-to-background ratio (SBR) of the subcutaneous blood vessel angiography in the abdomen of mice in (d).

[0023] Figure 13 Shows the retention time of DIPT-ICF Aggs in the blood circulation of rabbits: (a) DIPT-ICF Aggs was injected via the ear vein of rabbits, and then serum was collected at different time points after injection (0 h, 0.25 h, 0.5 h, 1 h, 2 h, 4 h, 8 h, 20 h, 48 h, 72 h, 96 h, and 120 h) and placed in a 96-well plate for NIR-II imaging (980 nm laser, 1319 nm long-pass filter, and exposure time 200 ms); (b) The NIR-II signal intensity distribution curve of DIPT-ICF Aggs in (a); (c) The quantitative analysis curve of the NIR-II average signal intensity of DIPT-ICF Aggs in (a); (d) NIR-II angiography of the ear blood vessels of rabbits at different time points (980 nm laser, 1319 nm long-pass filter, and exposure time 200 ms); (e) The NIR-II angiography signal intensity curve of the ear blood vessels of rabbits in (d); (f) The NIR-II signal-to-background ratio (SBR) of the ear blood vessels of rabbits marked with a yellow dotted line in (d).

[0024] Figure 14 NIR-II Imaging after Renal Transplantation Vascular Anastomosis with DIPT-ICF Aggs as the Contrast Agent: (a)-(d) show the bright-field images (upper) and NIR-II images (lower) of successful vascular anastomosis, respectively; (a) image of successful anastomosis of the transplanted kidney vessels; (b) partial stenosis of the renal artery anastomosis; (c) partial and complete stenosis of the renal vein anastomosis; (d) complete stenosis of the renal artery; (e)-(h) are the quantitative curves of NIR-II fluorescence signal intensity in (a), (b), (c), and (d), respectively; (j) quantitative analysis of the average NIR-II fluorescence intensity after reperfusion of the transplanted kidney in a-d. Detailed implementation manners

[0025] The present invention will be further described in detail below in conjunction with the embodiments and the drawings, but the implementation manners of the present invention are not limited thereto.

[0026] The reagents used in the following examples are all commercially available.

[0027] Example 1: Synthesis of NIR-II Luminescent Materials (DIPT-IC and DIPT-ICF) Based on High-Performance Fused-Ring Electron Acceptors Pyrrolo[2,3-b:4,5-b']diindole (DIP)

[0028]

[0029] The synthesis route is as Figure 1 shown:

[0030] (1) Under an argon atmosphere, compound 1 (3.962 g, 4.75 mmol), ethyl 2-bromothiophene-3-carboxylate (3.35 g, 14.25 mmol), Pd(PPh3)4 (145 mg, 0.12 mmol), K2CO3 (1.93 g, 13.96 mmol), distilled water (6 mL), toluene (30 mL), and a drop of trioctylmethylammonium chloride were successively added to a 100 mL round-bottom flask. The mixture was slowly heated to 115 °C and stirred for 22 h. After the reaction device was cooled to room temperature, the reaction solution was extracted with dichloromethane and then column chromatographed using petroleum ether / dichloromethane as the eluent to obtain intermediate 2 (DIP-COOEt) with a yield of 65%.

[0031] (2) Under the protection of argon gas, in a low-temperature environment of -78 °C, add THF (16 mL), 1-bromo-4-hexylbenzene (2.85 g, 11.8 mmol) to a 100 mL round-bottom flask respectively. Next, slowly add dropwise a solution of 2.4 M n-butyllithium in hexane (4.50 mL, 10.82 mmol). After keeping warm and stirring at -78 °C for 2 h, slowly add dropwise compound DIP-COOEt (1.751 g, 1.97 mmol) dissolved in THF (15 mL), then move to room temperature and stir overnight. After the reaction is completed, quench with water, extract with ethyl acetate and rotary evaporate to dryness, transfer the crude product to a 250 mL three-necked flask, then add acetic acid (40 mL) and octane (40 mL) and reflux for 5 h. After cooling to room temperature, quench the reaction with water, extract with ethyl acetate, and purify by column chromatography using petroleum ether as the eluent to obtain a blue-yellow intermediate 3 (DIPT) with a yield of 60%.

[0032] (3) In an argon atmosphere, first add 10.19 ml of anhydrous DMF to a 100 mL round-bottom flask, place the reaction device in an ice-water bath and cool to 0 °C, then add dropwise 2.03 ml of phosphorus oxychloride. After stirring at 0 °C for 2 h, slowly add dropwise intermediate 3 (0.721 g, 0.51 mmol) dissolved in 1,2-dichloroethane (10 ml). After the addition is complete, move the reaction device to an 85 °C oil bath and stir for 10 h. After the reaction is completed, pour the mixture into ice water, stir for 1 h, extract with dichloromethane, and purify by column chromatography using petroleum ether / dichloromethane (1:1) as the eluent to obtain an orange-red intermediate 4 (DIPT-CHO) with a yield of 60%.

[0033] (4) Under an argon atmosphere, add compound DIPT-CHO (0.42 g, 0.28 mmol), 2-(3-oxo-2,3-dihydro-1H-inden-1-ylidene) malononitrile (IC) (0.22 g, 1.12 mmol), pyridine (1 mL) and chloroform (40 mL) to a 100 mL round-bottom flask in sequence, then reflux for 8 h. After the reaction is completed, drop the mixture into methanol (200 mL) for precipitation, filter by suction and collect the solid. Purify by column chromatography using petroleum ether / dichloromethane as the eluent to obtain DIPT-IC, a bright black solid DIPT-IC, with a yield of 70%.

[0034] 11H NMR (400 MHz, CDCl3, δ / ppm) δ 8.87 (s, 2H), 8.65 (s, 2H), 7.89 (s, 2H), 7.68 (d, J = 12 Hz, 8H), 7.60 (s, 2H), 7.19 (d, J = 4 Hz, 8H), 7.07 (d, J = 8 Hz, 8H), 4.40 - 4.28 (m, 4H), 4.3 (d, J = 8 Hz, 2H), 2.56 (t, J = 8 Hz, 8H), 2.22 - 2.10 (m, 2H), 1.97 - 1.86 (m, 2H), 1.42 - 1.15 (m, 59H), 0.92 - 0.82 (m, 20H), 0.71 (t, J = 8 Hz, 4H), 0.60 (t, J = 8 Hz, 4H); 13 13C NMR (100 MHz, CDCl3, δ / ppm): 13 13C NMR (101 MHz, CDCl3) δ 188.85, 163.47, 160.76, 157.37, 147.30, 142.13, 141.92, 141.46, 140.18, 140.10, 138.35, 136.99, 134.87, 134.14, 129.87, 129.57, 128.63, 127.83, 125.23, 123.47, 121.10, 120.23, 117.10, 115.33, 115.05, 104.05, 77.48, 77.16, 76.84, 67.39, 62.33, 51.28, 40.04, 35.68, 31.84, 31.51, 30.87, 30.71, 29.25, 29.22, 28.78, 28.61, 24.30, 23.91, 23.21, 22.98, 22.72, 14.21, 14.07, 13.81, 11.08, 11.02, 10.74, 1.16; MS (MALDI-TOF, m / z) C 124 H 135 N7O2S2: calculated: 1819.611; found: 1818.794.

[0035] (5) Based on DIPT-IC, a blue-black solid DIPT-ICF was synthesized with a yield of 63%.

[0036] 11H NMR(400MHz,CDCl3,δ / ppm)δ8.87(s,2H),8.54 - 8.50(m,2H),7.71 - 7.63(m,6H),7.16(t,J=4Hz 8H),7.07(d,4Hz,8H),4.40 - 4.27(m,4H),4.24(d,J=4Hz,2H),2.20 - 2.09(m,2H),1.95 - 1.86(m,2H),1.28 - 1.15(m,59H),0.92 - 0.88(m,20H),0.85(t,J=8Hz 4H),0.60(t,J=8Hz 4H); 13 13C NMR(100MHz,CDCl3,δ / ppm):186.46,164.63,157.78,147.54,142.03,141.57,140.05,138.36,130.17,129.46,128.66,127.78,121.29,119.32,117.32,115.10,114.90,104.27,100.13,77.48,77.16,76.84,62.31,35.67,31.84,31.52,29.25,29.22,23.23,22.97,22.73,14.22,14.08,13.81,1.17,0.14; MS(MALDI - TOF,m / z)C 124 H 131 F4N7O2S2: calculated: 1891.571; found: 1890.181.sd.

[0037] Example 2: Characterization of the AIE properties of molecules DIPT - IC and DIPT - ICF

[0038] Figure 3 It is the fluorescence spectrogram of the materials obtained in Example 1 under different water content conditions. It can be seen from the figure that the two example materials emit weakly in the good solvent dimethylformamide. With the addition of the poor solvent water, the fluorescence gradually starts to increase. When the water content increases to 90%, the fluorescence intensity reaches the strongest state, indicating that both DIPT - IC and DIPT - ICF photosensitizers have AIE properties. In addition, the peak absorption wavelengths of the materials obtained in Example 1 are 746nm and 818 nanometers, and the peak fluorescence emission wavelengths are 979nm and 1014nm respectively, indicating that after introducing 4 fluorine atoms into molecule DIPT - IC, the absorption and emission are significantly red - shifted, and the photophysical properties of DIPT - ICF are better than those of DIPT - IC.

[0039] Example 3: Brightness characterization of molecules DIPT - IC and DIPT - ICF

[0040] Figure 4 The molar absorbance and quantum yield curves of DIPT-IC and DIPT-ICF. As can be seen from Figure 4 Figure a, the molar absorbance of DIPT-ICF at 808 nm is 3.13 times that of DIPT-IC. The quantum yields of DIPT-IC and DIPT-ICF are 2% and 0.7% respectively. Even though the quantum yield of DIPT-ICF is lower than that of DIPT-IC, the brightness of DIPT-ICF is higher than that of DIPT-IC.

[0041] Example 4: Preparation of DIPT-ICF Aggs (DIPT-ICF Aggregates)

[0042] From the characterization parameters of Examples 2 and 3, it can be seen that DIPT-ICF has better photophysical properties than DIPT-IC. Therefore, DIPT-ICF is selected as the molecular material of the aggregate contrast agent, and further aggregated to prepare DIPT-ICF Aggs.

[0043] The preparation route is as Figure 2 shown. Preparation of DIPT-ICF Aggs by solvent replacement method: Weigh 1 mg of DIPT-ICF and 6 mg of F-127 and dissolve them in 2 mL of THF solution respectively to obtain THF solution A and solution B. Then mix solution A and solution B to obtain mixed solution C; dropwise add solution C into 9 mL of ultrapure water, stir magnetically overnight to remove the solvent THF, and obtain a dark blue clear DIPT-ICF Aggs contrast agent solution.

[0044] Example 5: Characterization of the Photophysical Properties of DIPT-ICF Aggs

[0045] Figure 5 The spectral curves of DIPT-ICF Aggs in aqueous solution and the changes in the absorption spectra from the molecule DIPT-IC to DIPT-ICF Aggs. The absorption and emission peaks of DIPT-ICF Aggs are 974 nm and 1074 nm respectively. The absorption spectrum redshifts by 72 nm from the small molecule DIPT-IC to DIPT-ICF, and the absorption peak redshifts to 818 nm; from DIPT-ICF to DIPT-ICF Aggs, the absorption redshifts by 156 nm again, and the absorption peak redshifts to 974 nm, and the absorption spectrum tails to 1100 nm. Through the molecular engineering and aggregation process, the absorption spectrum of DIPT-ICF Aggs redshifts by a total of 228 nm, achieving simultaneous absorption and emission in the second near-infrared region, improving the penetration depth of living tissues and the signal-to-noise ratio of imaging.

[0046] Example 6: Characterization of the particle size and stability of DIPT-ICF Aggs

[0047] Figure 6 It is about the dynamic light scattering particle size distribution of DIPT-ICF Aggs, the aggregate morphology by transmission electron microscopy (TEM), and the stability of the aggregates in different pH environments. The particle size of DIPT-ICF Aggs is about 80 nm, and good morphology and luminescence stability are exhibited in acidic, neutral, and alkaline environments.

[0048] Example 7: Biocompatibility evaluation of DIPT-ICF Aggs at the cellular and animal levels

[0049] HeLa cells and 3T3 cells were used to evaluate the cytotoxicity of DIPT-ICF Aggs. HeLa cells and 3T3 cells were seeded in 96-well plates and cultured for 24 h, and then co-cultured with different concentrations of DIPT-ICF Aggs for 24 h. MTT was used to detect the toxicity of the two types of cells. The test results are shown in Figure 7 , and DIPT-ICF Aggs has no obvious cytotoxicity.

[0050] Balb / c nude mice and New Zealand white rabbits were used to evaluate the toxicity of DIPT-ICF Aggs at the animal level. 200 μL of 0.5 mg / mL DIPT-ICF Aggs was injected into Balb / c nude mice via the tail vein. After 2 weeks, blood samples and important organs (heart, liver, spleen, lung, kidney, intestine, and skin) of the nude mice were collected for blood biochemistry, blood cell count, and pathological analysis of important organs. 10 mL of 0.5 mg / mL DIPT-ICF Aggs was injected into New Zealand white rabbits via the marginal ear vein. After 2 weeks, blood and important organ (heart, liver, spleen, lung, kidney, intestine, and skin) specimens were collected for blood biochemistry, blood cell count, and pathological analysis of important organs. The results are shown in Figure 8 , Figure 9 .

[0051] Combined with Figure 7 , Figure 8 and Figure 9 it can be seen that DIPT-ICF Aggs has excellent biocompatibility.

[0052] Example 8: Comparison of NIR-I and NIR-II resolutions of DIPT-ICF Aggs as a contrast agent in in vitro vascular simulation angiography

[0053] 0.33 mg / mL DIPT-ICF Aggs contrast agent was aspirated into a capillary glass tube, and the capillary glass tube was covered with chicken breast tissue of different thicknesses (1 mm, 2 mm, 3 mm, 4 mm, and 5 mm in sequence). NIR-II imaging was performed using a small animal imager (the capillary glass tube was used to simulate blood vessels passing through animal tissues, and the chicken breast tissue was used to simulate animal living tissues). As Figure 9 shown, the fluorescence signal intensity and signal-background ratio (SBR) decreased with the increase in the depth of the chicken breast. When the chicken breast tissue with a depth less than 3 mm was covered, the SBR of NIR-II was significantly higher than that of NIR-I, indicating that DIPT-ICF Aggs is an excellent NIR-II fluorescence contrast agent.

[0054] Example 9: Resolution evaluation of NIR-II angiography of animal body surface blood vessels using DIPT-ICF Aggs as a contrast agent

[0055] 200 μL of 0.5 mg / mL DIPT-ICF Aggs was injected into Balb / c nude mice via the tail vein and imaged using a small animal imager. Multiple groups of angiography images of the body surface blood vessels were taken using different lasers (808 nm and 980 nm) combined with different filters (long-pass 900 nm, long-pass 1020 nm, long-pass 1100 nm, and long-pass 1319 nm). As Figure 11 shown, in the group of 980 nm laser + long-pass 1319 nm filter, the abdominal blood vessel course was the clearest and its SBR was the highest.

[0056] Example 10: DIPT-ICF Aggs has a long retention time in the blood circulation of Balb / c nude mice and New Zealand white rabbits

[0057] 200 μL of 0.5 mg / mL DIPT-ICF Aggs was injected into Balb / c nude mice via the tail vein. Serum was collected at different time points (0 h, 0.5 h, 1 h, 2 h, 4 h, 8 h, 24 h, 48 h, 72 h, 96 h, 120 h, and 144 h) and placed in a 96-well plate for NIR-II imaging. As Figure 12 shown, 8 h after tail vein injection, the DIPT-ICF Aggs retained in the serum still had a high fluorescence signal; the angiography image of the abdominal subcutaneous blood vessels in mice showed that the abdominal blood vessel texture was still clearly visible 10 h after the fluorescence contrast agent was given.

[0058] Meanwhile, 10 mL of DIPT-ICF Aggs at a concentration of 0.5 mg / mL was injected into New Zealand white rabbits via the marginal ear vein, and sera were collected at different time points after injection (0 h, 0.5 h, 1 h, 2 h, 4 h, 8 h, 24 h, 48 h, 72 h, 96 h, and 120 h). Then the collected sera were placed in 96-well plates for NIR-II imaging. After a single administration, the high NIR-II signal intensity of DIPT-ICF Aggs in rabbit sera could be maintained for 20 h. As Figure 13 shown, angiography of the rabbit ear 24 h after injection showed clearly visible blood vessels in the ear and a relatively high SBR.

[0059] From Figure 12 and Figure 13 it can be seen that the aggregated contrast agent DIPT-ICF Aggs has a long circulation retention time in Balb / c nude mice and New Zealand white rabbits, a high SBR in angiography, and excellent long-term angiography ability.

[0060] Example 11: As an NIR-II contrast agent, DIPT-ICF Aggs was used to perform real-time high-resolution imaging of the patency of vascular anastomoses during renal transplantation and to monitor renal allograft reperfusion

[0061] Four New Zealand white rabbits (weighing 2.5 - 3 kg) were anesthetized successfully by inhaling isoflurane. An incision about 15 cm long was made along the midline of the abdomen from the pubic symphysis to the xiphoid process. The intestines were pushed to the right side to expose the left retroperitoneal renal tissue and the left renal artery and vein. The left renal artery was quickly dissected and ligated, and a 24G intravenous indwelling needle was implanted at the distal end of the left renal artery. Pre-cooled kidney preservation solution was quickly injected until the color of the kidney turned yellowish-white, and then the left renal artery was transected. The left renal vein was ligated and transected, and the perfused left kidney was removed and placed in kidney preservation solution at 4°C. The left kidneys of the other three successfully anesthetized New Zealand white rabbits were removed in the same way and stored in pre-cooled kidney preservation solution. Allogeneic orthotopic renal transplantation was performed on New Zealand white rabbits respectively (model classification: good renal vascular anastomosis, renal vein anastomotic stenosis, partial stenosis of the renal artery anastomosis, and complete stenosis of the renal artery anastomosis). The experimental rabbits were placed in a small animal imaging instrument in the second near-infrared region, and the focal length was adjusted to clearly see the kidney and renal blood vessels in the bright field state. An 8 mL physiological saline solution of DIPT-ICF Aggs (concentration 0.33 mg / mL) was quickly injected into the experimental rabbits via the marginal ear vein, and images were collected in real time. From Figure 13 it can be seen that the patency of the renal transplantation vascular anastomoses and the renal allograft reperfusion in the four experimental models were clearly shown. The stenosis and reperfusion status of the renal transplantation vascular anastomoses could be clearly seen through NIR-II imaging, which helped urologists to timely detect the deficiencies of the surgery and improve the success rate of renal transplantation surgery.

[0062] The above embodiments are preferred embodiments of the present invention, but the embodiments of the present invention are not limited to the above embodiments. Any other changes, modifications, substitutions, combinations, and simplifications made without departing from the spirit and principle of the present invention shall be equivalent replacement methods and are all included in the protection scope of the present invention.

Claims

1. A near-infrared second near-infrared region luminescent compound, characterized in that, The near-infrared second-region luminescent compound has a structure represented by Formula I or II as follows:

2. The near-infrared second near-infrared region luminescent compound according to claim 1, characterized in that, The near-infrared second-region luminescent compound has AIE properties.

3. A preparation method of the near-infrared second near-infrared region luminescent compound according to claim 1 or 2, characterized in that, Its synthetic route is as follows:

4. The preparation method according to claim 3, characterized in that, It includes the following steps: Under an argon atmosphere, mix Compound 1, ethyl 2-bromothiophene-3-carboxylate, Pd(PPh3)4, K2CO3, distilled water, toluene, and trioctylmethylammonium chloride, slowly heat and stir. After the reaction device cools to room temperature, the reaction solution is extracted with dichloromethane and then column chromatographed using petroleum ether / dichloromethane as the eluent to obtain Intermediate 2 (DIP-COOEt); Under the protection of argon, in an environment at -78 °C, add THF, 1-bromo-4-hexylbenzene, and hexane solution of 2.4 M n-butyllithium, stir and then dropwise add Compound DIP-COOEt dissolved in THF. After the reaction is complete, quench with water, extract with ethyl acetate and rotary evaporate, transfer the crude product, then add acetic acid and octane and reflux. After cooling to room temperature, quench the reaction with water, extract with ethyl acetate and then column chromatograph using petroleum ether as the eluent to separate, obtaining the blue-yellow Intermediate 3 (DIPT); In an argon atmosphere, first add anhydrous DMF, cool the reaction device to 0 °C in an ice-water bath, then dropwise add phosphorus oxychloride. After stirring at 0 °C, dropwise add Intermediate 3 dissolved in 1,2-dichloroethane. After the addition is complete, transfer the reaction device to an 85 °C oil bath and stir. After the reaction is complete, pour the mixture into ice water and stir, extract with dichloromethane and then purify by column chromatography using petroleum ether / dichloromethane as the eluent to obtain the orange-red Intermediate 4 (DIPT-CHO); Under an argon atmosphere, sequentially add Compound DIPT-CHO, 2-(3-oxo-2,3-dihydro-1H-inden-1-ylidene) malononitrile (IC), pyridine, and chloroform, then reflux. After the reaction is complete, drop the mixture into methanol for precipitation, filter by suction and collect the solid, and separate by column chromatography using petroleum ether / dichloromethane as the eluent to obtain DIPT-IC.

5. A near-infrared second near-infrared region luminescent compound aggregate, characterized in that, The near-infrared second-region luminescent compound aggregate is aggregated by small molecules having the following structure, 6. The near-infrared second near-infrared region luminescent compound aggregate according to claim 5, characterized in that, The absorption spectrum and emission spectrum of the near-infrared second-region luminescent compound aggregate are both in the near-infrared second region.

7. Use of the near-infrared second near-infrared region luminescent compound aggregate according to any one of claims 5 or 6 in the preparation of a fluorescence contrast agent material.

8. Non-diagnostic or therapeutic use of the near-infrared second near-infrared region luminescent compound aggregate according to any one of claims 5 or 6 in renal transplantation surgery.

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