Metal complex with high active oxygen generation efficiency and preparation method and application thereof

Abstract

The application discloses a metal complex with high active oxygen generation efficiency and a preparation method and application thereof, and relates to the technical field of photosensitizers. The application takes terpyridine as an electron acceptor, introduces an electron-donating group to adjust a push-pull electron system of a molecule, further introduces a metal ion to coordinate and adjust electron-withdrawing capacity of the molecule, and finally adjusts a conjugated system, so that a series of metal complexes with high active oxygen generation efficiency are synthesized. The synthesis method is simple, the conditions are mild, the yield is high, the product shows good biocompatibility, shows excellent photodynamic anticancer or antibacterial effects in vitro, and can be used for preparing photodynamic anticancer or antibacterial photosensitizers.

Description

TECHNICAL FIELD

[0001] The present application relates to the technical field of photosensitizers, in particular to a metal complex with high active oxygen generation efficiency and a preparation method and application thereof. BACKGROUND

[0002] At present, the main treatment methods for cancer in clinic include surgery, chemotherapy and radiotherapy, etc. These treatment methods have the problems of invasiveness, large toxic and side effects, difficulty in eliminating latent lesions and high recurrence rate, etc., which seriously affect the quality of life of patients. It is urgent to develop new cancer treatment methods.

[0003] Among the many new cancer treatment methods, photodynamic therapy (PDT) has attracted much attention since its inception. At present, PDT has been approved by the U.S. Food and Drug Administration for the treatment of various cancers such as skin cancer. PDT uses light of appropriate wavelength to irradiate photosensitizers to produce reactive oxygen species (ROS), thereby oxidizing biological macromolecules such as proteins and lipids in tumor cells, causing tumor cells to undergo cell programmed death such as necrosis, apoptosis, autophagy, ferroptosis and pyroptosis. PDT has the advantages of non-invasiveness, small drug resistance, low side effects and ability to eliminate latent lesions, which can effectively overcome the disadvantages of traditional cancer treatment methods such as chemotherapy and radiotherapy.

[0004] Photosensitizers play a crucial role in photodynamic therapy. Among them, metal complex photosensitizers not only have excellent light stability, but also the heavy atom effect of the metal center can enhance the spin-orbit coupling (SOC) of the complex, promote intersystem crossing (ISC), and thus improve the ROS generation capacity. Based on the above advantages, metal complex photosensitizers have attracted widespread attention from researchers. Currently, noble metal complex photosensitizers such as ruthenium and iridium are widely used in photodynamic therapy. However, they have the disadvantages of large toxic and side effects, high metal price which is not conducive to sustainable development, and poor water solubility, etc., which limit their further application.

[0005] It has been reported that essential metal elements for human body, such as zinc and nickel, have excellent biocompatibility for metal center complexes, which can overcome the disadvantage of strong dark toxicity of ruthenium and iridium metal complex photosensitizers. At present, there are some applications of essential metal element complexes in PDT, but there are few related reports. In addition, essential metal elements have very limited improvement on the ISC efficiency of molecules, which leads to the problem of low active oxygen generation efficiency of these complex photosensitizers in the process of photodynamic therapy. SUMMARY

[0006] In order to solve the above problems in the prior art, the present application aims to provide a metal complex with high active oxygen generation efficiency, a preparation method and application thereof, which is constructed by taking metal ions as molecular electron-withdrawing sites to form a push-pull electron system, reduces the molecular singlet-triplet energy system and promotes intersystem crossing, thereby promoting the generation of active oxygen, effectively improving the problem of low active oxygen yield of the existing metal complex photosensitizer; at the same time, the complex has good biocompatibility, and can realize safe and efficient photodynamic therapy of cancer.

[0007] The technical scheme for solving the above technical problems of the present application is as follows: a metal complex with high active oxygen generation efficiency is provided, which has a structure as shown in formula I:

[0008]

[0009] wherein, is

[0010] R1 is

[0011] R is C1-C12 alkyl

[0012] M is a transition metal ion; X is a halogen ion, NO3- or AcO-; a is 1, 2 or 3.

[0013] Further, M is Cr 3+ , Cr 2+ , Mn + , Mn 2+ , Mn 3+ , Fe 2+ , Fe 3+ , Co 2+ , Co 3+ , Ni 2+ , Ni + , Cu + , Cu 2+ or Zn 2+ .

[0014] Still further, the specific structure of the metal complex with high active oxygen generation efficiency is as follows:

[0015]

[0016]

[0017] The preparation method of the metal complex with high active oxygen generation efficiency comprises the following steps:

[0018] (1) dissolving compound A, compound B, palladium catalyst and basic substance in solvent, reacting at 50-60°C for 11-13h under inert atmosphere to obtain compound C;

[0019] (2) dissolving 2-acetylpyridine and sodium hydroxide in organic solvent, adding compound C, reacting at -3-3°C for 1.5-2.5h, adding ammonium acetate, reacting at 55-65°C for 1.5-2.5h to obtain compound D;

[0020] (3) dissolving compound D and MX a in solvent, coordinating for 25-35min to obtain metal complex with high active oxygen generation efficiency as shown in formula I; the synthetic route is as follows:

[0021]

[0022] R2 in compound A is halogen.

[0023] Further, R2 in compound A is Br.

[0024] Further, the solvent in step (1) is mixed solvent of tetrahydrofuran and water, the volume ratio of tetrahydrofuran to water in the solvent is (7-9):1; the organic solvent in step (2) is anhydrous ethanol; the solvent in step (3) is mixed solvent of methanol and dichloromethane, the volume ratio of methanol to dichloromethane in the solvent is 1:(2-3).

[0025] Further, the solvent in step (1) is mixed solvent of tetrahydrofuran and water, the volume ratio of tetrahydrofuran to water in the solvent is 8:1; the solvent in step (3) is mixed solvent of methanol and dichloromethane, the volume ratio of methanol to dichloromethane in the solvent is 1:2.

[0026] Further, step (1) comprises the following steps: dissolving compound A, compound B, palladium catalyst and basic substance in solvent, reacting at 50-60°C for 11-13h under inert atmosphere, cooling to room temperature, removing solvent by reduced pressure distillation, adding water and dichloromethane to extract product, removing dichloromethane by reduced pressure distillation and purifying to obtain compound C.

[0027] Further, silica gel column chromatography is used for purification in step (1), and the eluent is at least one of dichloromethane, ethyl acetate and petroleum ether.

[0028] Further, the reaction temperature in step (1) is 55°C, and the time is 12h.

[0029] Further, the molar ratio of compound A, compound B, palladium catalyst and basic substance is 1:(1.2-1.5):(0.04-0.05):(3.5-4).

[0030] Further, the palladium catalyst is tetrakis(triphenylphosphine)palladium, bis(triphenylphosphine)palladium chloride, [1,1'-bis(diphenylphosphino)ferrocene]dichloropalladium, tris(dibenzylideneacetone)dipalladium or palladium acetate; and the basic substance is sodium tert-butoxide, potassium tert-butoxide, sodium carbonate, potassium carbonate, cesium carbonate, potassium phosphate or barium hydroxide.

[0031] Further, the palladium catalyst is tetrakis(triphenylphosphine)palladium; and the basic substance is potassium carbonate.

[0032] Further, step (2) comprises the following steps: dissolving 2-acetylpyridine and sodium hydroxide in an organic solvent and mixing at -3-3°C for 9-11 minutes, then adding compound C and reacting at -3-3°C for 1.5-2.5 hours, then adding ammonium acetate and reacting at 55-65°C for 1.5-2.5 hours, cooling to room temperature, removing the organic solvent by distillation under reduced pressure, adding water and dichloromethane to extract the product, removing dichloromethane by distillation under reduced pressure and purifying the product to obtain compound D.

[0033] Further, in step (2), the purification is performed by silica gel column chromatography using ethyl acetate and petroleum ether as eluent.

[0034] Further, in step (2), compound C is added and reacted at 0°C for 2 hours, and then ammonium acetate is added and reacted at 60°C for 2 hours.

[0035] Further, in step (2), the molar ratio of compound C, 2-acetylpyridine, sodium hydroxide and ammonium acetate is 1.0:(2.0-2.5):(3.5-4.0):(30.0-40.0).

[0036] Further, step (3) comprises the following steps: dissolving compound D and MX a in a solvent and coordinating for 25-35 minutes, removing the solvent by distillation under reduced pressure, adding water and dichloromethane to extract the product, removing dichloromethane by distillation under reduced pressure and purifying the product to obtain the metal complex having high active oxygen generation efficiency as shown in formula I.

[0037] Further, in step (3), the purification is performed by silica gel column chromatography using at least one of petroleum ether, n-hexane, dichloromethane and methanol as eluent.

[0038] Further, in step (3), the coordination is performed at room temperature for 30 minutes.

[0039] Further, in step (3), the molar ratio of compound D and MX a is 1:(1-1.2).

[0040] The metal complex having high active oxygen generation efficiency is used in the preparation of a photodynamic anticancer or antibacterial photosensitizer.

[0041] The present application has the following beneficial effects:

[0042] (1) The present application introduces electron-donating groups to adjust the push-pull electron system of the molecule, and then introduces metal ion coordination to adjust the electron-withdrawing ability of the molecule, and finally adjusts the conjugated system, to synthesize a series of metal complexes with high active oxygen generation efficiency; by constructing the push-pull electron system and introducing metal ion coordination, the singlet-triplet energy system and the intersystem crossing of the complex are reduced, and the active oxygen generation efficiency of the metal complex is improved.

[0043] (2) The metal complex of the present application has high active oxygen generation efficiency, including singlet oxygen and hydroxyl radicals; the metal complex not only shows good biocompatibility, but also shows excellent photodynamic antitumor effect in vitro and can effectively kill tumor cells in anoxic environment, and also shows excellent in vivo tumor targeting ability and in vivo photodynamic anticancer effect in tumor-bearing mice.

[0044] (3) The metal complex with high active oxygen generation efficiency provided by the present application, as well as the preparation method and application thereof, not only provides a favorable tool for efficient photodynamic tumor therapy, but also provides a more systematic guide for the rational design of the next generation of metal complex photosensitizers. BRIEF DESCRIPTION OF DRAWINGS

[0045] Figure 1 The photophysical properties of the metal complexes and compound D prepared in Examples 1-7 in dichloromethane solvent; wherein A is a UV-vis absorption spectrum, and B is a fluorescence emission spectrum;

[0046] Figure 2 The active oxygen yield test diagram of the metal complexes prepared in Examples 1-7 and the commercially available photosensitizer Ce6; wherein A is total active oxygen, B is singlet oxygen, and C is hydroxyl radical;

[0047] Figure 3 The laser confocal imaging diagram of the total active oxygen / hydroxyl radical produced by the metal complex prepared in Example 1 under dark / light conditions; wherein A is the total active oxygen production confocal imaging diagram, and B is the hydroxyl radical confocal imaging diagram;

[0048] Figure 4 The phototoxicity and dark toxicity results diagram of the metal complex prepared in Example 1 in different cells; wherein A is the phototoxicity and dark toxicity results diagram of the metal complex in mouse breast cancer cells 4T1, B is the phototoxicity and dark toxicity results diagram of the metal complex in human hepatoma cells HepG2, C is the phototoxicity and dark toxicity results diagram of the metal complex in human colon cancer cells HCT116, and D is the phototoxicity and dark toxicity results diagram of the metal complex in human cervical cancer cells HeLa;

[0049] Figure 5 Intracellular reactive oxygen species generation imaging of the metal complex prepared in Example 1 under hypoxia condition in dark / light.

[0050] Figure 6 In vitro photodynamic anticancer performance test chart of the metal complex prepared in Example 1 and the commercially available photosensitizer Ce6 under normoxia / hypoxia condition; wherein, A is the photodynamic anticancer performance test chart of the metal complex prepared in Example 1, and B is the photodynamic anticancer performance test chart of the commercially available photosensitizer Ce6.

[0051] Figure 7 Viable staining chart of mouse breast cancer cells 4T1 under normoxia light of the metal complex prepared in Example 1 and the commercially available photosensitizer Ce6;

[0052] Figure 8 Viable staining chart of mouse breast cancer cells 4T1 under hypoxia condition light of the metal complex prepared in Example 1 and the commercially available photosensitizer Ce6;

[0053] Figure 9 Hemolysis experiment result chart of the metal complex prepared in Example 1.

[0054] Figure 10 Tumor-bearing mouse in vivo imaging and ex vivo organ imaging chart of the metal complex prepared in Example 1; wherein, A is the tumor-bearing mouse in vivo imaging chart, and B is the tumor-bearing mouse ex vivo organ imaging chart.

[0055] Figure 11 In vivo photodynamic therapy of tumor-bearing mice of the metal complex prepared in Example 1 mouse in vivo tumor and ex vivo tumor tissue chart; wherein, A is the mouse in vivo tumor picture, and B is the ex vivo tumor tissue picture.

[0056] Figure 12 In vivo photodynamic therapy of tumor-bearing mice of the metal complex prepared in Example 1 mouse tumor and body weight change chart; wherein, A is the mouse tumor volume change chart over time, B is the ex vivo tumor weight chart, and C is the mouse body weight change chart over time.

[0057] Figure 13 In vivo photodynamic therapy of tumor-bearing mice of the metal complex prepared in Example 1 mouse tumor tissue or organ H&E staining chart; wherein, A is the mouse tumor tissue H&E staining and Ki67 immunohistochemical analysis chart, and B is the mouse organ H&E staining chart. DETAILED DESCRIPTION

[0058] The following examples are intended to illustrate the present application and are not intended to limit the scope of the present application. Unless otherwise indicated, the conditions of the examples are conventional or manufacturer's recommended conditions. Unless otherwise indicated, the reagents or instruments used are conventional products available commercially.

[0059] Example 1

[0060] A metal complex with high active oxygen generation efficiency, having a structure as shown in Formula I-1:

[0061]

[0062] The metal complex with a structure as shown in Formula I-1 in this example is marked as Zn(II)-TPY-TPA-DTZ, which is prepared by the following steps:

[0063] (1) Take 45.0 mL of a mixed solvent of tetrahydrofuran and water (8:1 v / v), then sequentially add 5-bromo-2,2'-bithiophene-5'-carboxaldehyde (819.5 mg, 3.0 mmol), (4-(diphenylamino)phenyl)boronic acid (1012.1 mg, 3.5 mmol), and potassium carbonate (1382.1 mg, 10.0 mmol), and finally add tetrakis(triphenylphosphine)palladium (46.1 mg), and heat the mixture to 55°C under nitrogen protection for 12 h. After cooling the reaction solution, the solvent is removed by distillation, then water and dichloromethane are added to extract the product, after the extraction is completed, the dichloromethane is removed by distillation under reduced pressure, and the obtained product is purified by silica gel column chromatography, with petroleum ether: ethyl acetate (70:1 v / v) as the eluent, and finally 866.4 mg of yellow powder compound C-1 is obtained, with a yield of 83.0%; the synthetic route is as follows:

[0064]

[0065] (2) Dissolve sodium hydroxide (240.0 mg, 6.0 mmol) in 40.0 mL of an EtOH solution, then add 2-acetylpyridine (363.6 mg, 3.0 mmol), and place the mixture in a 0°C environment and stir for 10 min; then add compound C-1 (656.4 mg, 1.5 mmol) and continue stirring for 2 h; after 2 h, add excess ammonium acetate, and then heat the mixture to 60°C and react for 2 h; after cooling the reaction solution, the solvent is removed by distillation, then water and dichloromethane are added to extract the product, after the extraction is completed, the dichloromethane is removed by distillation under reduced pressure, and the obtained product is purified by silica gel column chromatography, with petroleum ether: ethyl acetate (5:1 v / v) as the eluent, and finally 151.4 mg of yellow powder compound D-1 is obtained, marked as TPY-TPA-DTZ, with a yield of 15.0%; the synthetic route is as follows:

[0066]

[0067] Characterization data: 1 H NMR (400 MHz, CDC13) δ 8.75 (d, J = 4.8 Hz, 2H), 8.69 (s, 2H), 8.66 (d, J = 8.0 Hz, 2H), 7.89 (td, J = 7.6, 1.6 Hz, 2H), 7.73 (d, J = 3.8 Hz, 1H), 7.47 (d, J = 8.4 Hz, 2H), 7.39 - 7.35 (m, 2H), 7.28 (t, J = 7.8 Hz, 4H), 7.21 (d, J = 4.0 Hz, 2H), 7.16 - 7.12 (m, 5H), 7.09 - 7.03 (m, 4H). 13 C NMR (100 MHz, CDC13) δ 155.86, 149.00, 147.55, 144.05, 143.29, 139.53, 137.35, 129.50, 127.06, 126.60, 125.39, 124.80, 124.51, 124.15, 123.62, 123.39, 123.13, 121.66, 116.95. HRMS (ESI): m / z [M + H] + calculated for C 41 H 29 N4S2: 641.1834; found: 641.1814.

[0068] (3) TPY-TPA-DTZ (64.1 mg, 0.1 mmol) and ZnCl2(13.6 mg, 0.1 mmol) were added to a mixed solvent of 10.0 mL dichloromethane and 5.0 mL methanol, and the mixture was stirred to coordinate at room temperature for 30 min; after the coordination reaction was completed, the solvent was removed by reduced pressure distillation, followed by adding water and dichloromethane to extract the product, and after the extraction was completed, dichloromethane was removed by reduced pressure distillation, and the obtained product was purified by silica gel column chromatography, eluent was dichloromethane:methanol (65:1 v / v), finally 70.0 mg of red powder Zn(II)-TPY-TPA-DTZ was obtained, the yield was 90.0%; its synthetic route is as follows:

[0069]

[0070] Characterization data: 1H NMR (400 MHz, DMSO-d6) δ 8.88-8.82 (m, 6H), 8.38-8.17 (m, 3H), 7.85-7.83 (m, 2H), 7.62-7.56 (m, 3H), 7.58-7.48 (m, 2H), 7.37-7.34 (m, 4H), 7.13-7.05 (m, 6H), 7.01-6.99 (m, 2H). HRMS (ESI): m / z [M-Cl] + calculated for C 41 H 28 ClN4S2Zn:739.0735;found:739.0734.

[0071] Example 2:

[0072] A metal complex with high active oxygen production efficiency, having a structure as shown in formula I-2:

[0073]

[0074] The metal complex with a structure as shown in formula I-2 in this example is marked as Fe(III)-TPY-TPA-DTZ, and its preparation method steps (1) and (2) are the same as those in Example 1, and step (3) includes the following steps:

[0075] TPY-TPA-DTZ (64.1 mg, 0.1 mmol) and FeCl3(16.2 mg, 0.1 mmol) were added to a mixed solvent of 10.0 mL of dichloromethane and 5.0 mL of methanol, and the mixture was stirred at room temperature for 30 min. After the coordination reaction was completed, the solvent was removed under reduced pressure, followed by adding water and dichloromethane to extract the product. After the extraction was completed, the dichloromethane was removed under reduced pressure, and the obtained product was purified by silica gel column chromatography with dichloromethane:methanol (65:1 v / v) as the eluent. Finally, 70.5 mg of black powder Fe(III)-TPY-TPA-DTZ was obtained, with a yield of 88.8%; its synthesis route is as follows:

[0076]

[0077] Example 3:

[0078] A metal complex with high active oxygen production efficiency, having a structure as shown in formula I-3:

[0079]

[0080] The metal complex having the structure shown in Formula I-3 in this embodiment is labeled as Cr(III)-TPY-TPA-DTZ, and its preparation method steps (1) and (2) are the same as those of Example 1, and step (3) includes the following steps:

[0081] TPY-TPA-DTZ (64.1 mg, 0.1 mmol) and CrCl3 (15.8 mg, 0.1 mmol) were added to a mixed solvent of 10.0 mL of dichloromethane and 5.0 mL of methanol, and the mixture was stirred and coordinated at room temperature for 30 min; after the coordination reaction was completed, the solvent was removed by reduced pressure distillation, then water and dichloromethane were added to extract the product, after the extraction was completed, dichloromethane was removed by reduced pressure distillation, and the obtained product was purified by silica gel column chromatography, with dichloromethane:methanol (65:1 v / v) as the eluent, finally 71.2 mg of black powder Cr(III)-TPY-TPA-DTZ was obtained, with a yield of 89.1%; its synthesis route is as follows:

[0082]

[0083] Example 4:

[0084] A metal complex with high active oxygen generation efficiency, having the structure shown in Formula I-4:

[0085]

[0086] The metal complex having the structure shown in Formula I-4 in this embodiment is labeled as Ni(II)-TPY-TPA-DTZ, and its preparation method steps (1) and (2) are the same as those of Example 1, and step (3) includes the following steps:

[0087] TPY-TPA-DTZ (64.1 mg, 0.1 mmol) and NiCl2 (12.9 mg, 0.1 mmol) were added to a mixed solvent of 10.0 mL of dichloromethane and 5.0 mL of methanol, and the mixture was stirred and coordinated at room temperature for 30 min; after the coordination reaction was completed, the solvent was removed by reduced pressure distillation, then water and dichloromethane were added to extract the product, after the extraction was completed, dichloromethane was removed by reduced pressure distillation, and the obtained product was purified by silica gel column chromatography, with dichloromethane:methanol (65:1 v / v) as the eluent, finally 72.7 mg of black powder Ni(II)-TPY-TPA-DTZ was obtained, with a yield of 94.4%; its synthesis route is as follows:

[0088]

[0089] Example 5:

[0090] A metal complex with high active oxygen generation efficiency, having the structure shown in Formula I-5:

[0091]

[0092] The metal complex having the structure shown as Formula I-5 in this embodiment is labeled as Co(II)-TPY-TPA-DTZ, and its preparation method includes the following steps:

[0093] TPY-TPA-DTZ (64.1 mg, 0.1 mmol) and CoCl2(12.9 mg, 0.1 mmol) were added to a mixed solvent of 10.0 mL of dichloromethane and 5.0 mL of methanol, and the mixture was stirred at room temperature for complexation for 30 min; after the completion of the complexation reaction, the solvent was removed by distillation under reduced pressure, followed by the addition of water and dichloromethane to extract the product, and after the completion of the extraction, dichloromethane was removed by distillation under reduced pressure. The obtained product was purified by silica gel column chromatography, and the eluent was dichloromethane:methanol (65:1 v / v). Finally, 71.6 mg of black powder Co(II)-TPY-TPA-DTZ was obtained, with a yield of 92.9%. The synthetic route is as follows:

[0094]

[0095] Example 6:

[0096] A metal complex with high active oxygen generation efficiency, having the structure shown as Formula I-6:

[0097]

[0098] The metal complex having the structure shown as Formula I-6 in this embodiment is labeled as Mn(II)-TPY-TPA-DTZ, and its preparation method includes the following steps:

[0099] TPY-TPA-DTZ (64.1 mg, 0.1 mmol) and MnCl2(12.6 mg, 0.1 mmol) were added to a mixed solvent of 10.0 mL of dichloromethane and 5.0 mL of methanol, and the mixture was stirred at room temperature for complexation for 30 min; after the completion of the complexation reaction, the solvent was removed by distillation under reduced pressure, followed by the addition of water and dichloromethane to extract the product, and after the completion of the extraction, dichloromethane was removed by distillation under reduced pressure. The obtained product was purified by silica gel column chromatography, and the eluent was dichloromethane:methanol (65:1 v / v). Finally, 73.1 mg of yellow powder Mn(II)-TPY-TPA-DTZ was obtained, with a yield of 95.3%. The synthetic route is as follows:

[0100]

[0101] Example 7:

[0102] A metal complex with high active oxygen production efficiency, having a structure as shown in formula I-7:

[0103]

[0104] The metal complex with a structure as shown in formula I-7 in this example is marked as Cu(II)-TPY-TPA-DTZ, and its preparation method steps (1) and (2) are the same as those of Example 1, and step (3) includes the following steps:

[0105] TPY-TPA-DTZ (64.1 mg, 0.1 mmol) and CuCl2 (13.4 mg, 0.1 mmol) were added to a mixed solvent of 10.0 mL of dichloromethane and 5.0 mL of methanol, and the mixture was stirred at room temperature for 30 min for coordination; after the completion of the coordination reaction, the solvent was removed under reduced pressure, followed by adding water and dichloromethane to extract the product, and after the completion of the extraction, dichloromethane was removed under reduced pressure, and the obtained product was purified by silica gel column chromatography with dichloromethane:methanol (65:1 v / v) as the eluent, and finally 74.8 mg of black powder Cu(II)-TPY-TPA-DTZ was obtained with a yield of 96.5%; and the synthesis route thereof is as follows:

[0106]

[0107] Example 8:

[0108] A metal complex with high active oxygen production efficiency, having a structure as shown in formula I-8:

[0109]

[0110] The metal complex with a structure as shown in formula I-8 in this example is marked as Zn(II)-TPY-TPA-TZ, and is prepared by the following steps:

[0111] (1) Take 45.0 mL of a mixed solvent of tetrahydrofuran and water (8:1 v / v), then sequentially add 5-bromothiophene-2-carboxaldehyde (573.2 mg, 3.0 mmol), (4-(diphenylamino)phenyl)boronic acid (1012.1 mg, 3.5 mmol), and potassium carbonate (1382.1 mg, 10.0 mmol), and finally add tetrakis(triphenylphosphine)palladium (46.1 mg), and heat the mixture to 55°C under nitrogen protection, and reflux for 12 h. After the reaction solution is cooled, the solvent is removed by distillation, then water and dichloromethane are added to extract the product, after the extraction is completed, dichloromethane is removed by distillation under reduced pressure, and the obtained product is purified by silica gel column chromatography, with petroleum ether: ethyl acetate (60:1 v / v) as the eluent, to finally obtain 766.9 mg of yellow powder compound C-2, with a yield of 71.2%; the synthesis route thereof is as follows:

[0112]

[0113] (2) Dissolve sodium hydroxide (240.0 mg, 6.0 mmol) in 40.0 mL of an EtOH solution, then add 2-acetylpyridine (363.6 mg, 3.0 mmol), and place the mixture in a 0°C environment and stir for 10 min; then add compound C-2 (533.2 mg, 1.5 mmol) and continue to stir for 2 h; after 2 h, add excess ammonium acetate, and then heat the mixture to 60°C and react for 2 h. After the reaction solution is cooled, the solvent is removed by distillation, then water and dichloromethane are added to extract the product, after the extraction is completed, dichloromethane is removed by distillation under reduced pressure, and the obtained product is purified by silica gel column chromatography, with petroleum ether: ethyl acetate (5:1 v / v) as the eluent, to finally obtain 120.6 mg of yellow powder compound D-2, labeled as TPY-TPA-TZ, with a yield of 14.4%; the synthesis route thereof is as follows:

[0114]

[0115] Characterization data: 1 H NMR (400 MHz, CDCl3) δ. 8.74 (ddd, J = 4.8, 1.6, 0.8 Hz, 2H), 8.68 (s, 2H), 8.64 (d, J = 8.0 Hz, 2H), 7.86 (td, J = 7.6, 1.8 Hz, 2H), 7.73 (d, J = 3.8 Hz, 1H), 7.56-7.52 (m, 2H), 7.35 (ddd, J = 7.4, 4.8, 1.0 Hz, 2H), 7.31-7.27 (m, 5H), 7.16-7.13 (m, 4H), 7.11-7.04 (m, 4H). 13C NMR (100 MHz, CDC13) δ 156.27, 156.17, 149.27, 147.93, 147.55, 146.17, 143.51, 140.01, 136.96, 129.51, 127.97, 127.01, 126.77, 123.99, 123.54, 123.44, 123.40, 121.47, 116.78. HRMS (ESI): m / z [M+H] + calculated for C 37 H 27 N4S: 559.1956; found: 559.1942.

[0116] (3) TPY-TPA-TZ (55.8 mg, 0.1 mmol) and ZnCl2(13.6 mg, 0.1 mmol) were added to a mixed solvent of 10.0 mL of dichloromethane and 5.0 mL of methanol, and the mixture was stirred and coordinated at room temperature for 30 min; after the coordination reaction was completed, the solvent was removed under reduced pressure, followed by adding water and dichloromethane to extract the product, and after the extraction was completed, dichloromethane was removed under reduced pressure, and the obtained product was purified by silica gel column chromatography, eluent was dichloromethane: methanol (70: 1 v / v), finally 60.4 mg of red powder Zn(II)-TPY-TPA-TZ was obtained, the yield was 86.6%; its synthetic route is as follows:

[0117]

[0118] Characterization data: 1 H NMR (400 MHz, DMSO-d6) δ 8.95-8.66 (m, 6H), 8.40-8.20 (m, 3H), 7.91-7.76 (m, 2H), 7.71-7.67 (m, 3H), 7.39-7.3 (m, 4H), 7.15-7.03 (m, 8H). HRMS (ESI): m / z [M-Cl] + calculated for C 37 H 26 ClN4SZn: 657.0858; found: 657.0862.

[0119] Example 9:

[0120] A metal complex with high active oxygen generation efficiency, having a structure as shown in formula I-9:

[0121]

[0122] The metal complex having the structure as shown in Formula I-9 in this embodiment is labeled as Zn(II)-TPY-TPA-TF, which is prepared by the following steps:

[0123] (1) Take 45.0 mL of a mixed solvent of tetrahydrofuran and water (8:1 v / v), then sequentially add 5-bromofuran-2-carboxaldehyde (525.0 mg, 3.0 mmol), (4-(diphenylamino)phenyl)boronic acid (1012.1 mg, 3.5 mmol), and potassium carbonate (1382.1 mg, 10.0 mmol), and finally add tetrakis(triphenylphosphine)palladium (46.1 mg), and heat the mixture to 55°C under nitrogen protection for 12 h. After cooling, the solvent is removed by distillation, then water and dichloromethane are added to extract the product, after the extraction is completed, dichloromethane is removed by distillation under reduced pressure, and the obtained product is purified by silica gel column chromatography, with petroleum ether: ethyl acetate (65:1 v / v) as the eluent, finally 857.7 mg of compound C-3 in dark yellow powder is obtained, with a yield of 84.0%; the synthesis route is as follows:

[0124]

[0125] (2) Dissolve sodium hydroxide (240.0 mg, 6.0 mmol) in 40.0 mL of EtOH solution, then add 2-acetylpyridine (363.6 mg, 3.0 mmol), and place the mixture in 0°C for stirring for 10 min; then add compound C-3 (510.0 mg, 1.5 mmol) and continue stirring for 2 h; after 2 h, add excess ammonium acetate, and then heat the mixture to 60°C for reaction for 2 h; after cooling, the solvent is removed by distillation, then water and dichloromethane are added to extract the product, after the extraction is completed, dichloromethane is removed by distillation under reduced pressure, and the obtained product is purified by silica gel column chromatography, with petroleum ether: ethyl acetate (5:1 v / v) as the eluent, finally 138.4 mg of compound D-3 in light yellow powder is obtained, labeled as TPY-TPA-TF, with a yield of 17.0%; the synthesis route is as follows:

[0126]

[0127] Characterization data: 1H NMR (400 MHz, DMSO-d6) δ 8.97 - 8.79 (m, 6H), 8.37 - 8.27 (m, 2H), 8.00 - 7.82 (m, 5H), 7.41 - 7.37 (m, 4H), 7.19 - 7.00 (m, 9H). HRMS (ESI): m / z [M - Cl] 13 C NMR (150 MHz, CDC13) δ 156.35, 156.04, 155.39, 149.23, 147.53, 139.57, 136.94, 129.48, 125.42, 124.82, 124.36, 123.93, 123.36, 121.42, 114.91, 111.82, 106.60. HRMS (ESI): m / z [M + H] + calculated for C 37 H 27 N4O: 543.2185; found: 543.2161.

[0128] (3) TPY-TPA-TF (54.3 mg, 0.1 mmol) and ZnCl2(13.6 mg, 0.1 mmol) were added to a mixed solvent of 10.0 mL of dichloromethane and 5.0 mL of methanol, and the mixture was stirred and coordinated at room temperature for 30 min; after the coordination reaction was completed, the solvent was removed under reduced pressure, followed by adding water and dichloromethane to extract the product, and after the extraction was completed, dichloromethane was removed under reduced pressure, and the obtained product was purified by silica gel column chromatography with dichloromethane:methanol (70:1 v / v) as the eluent, and finally 55.0 mg of red powder Zn(II)-TPY-TPA-TF was obtained with a yield of 82.0%; the synthetic route thereof is as follows:

[0129]

[0130] Characterization data: 1 H NMR (400 MHz, DMSO-d6) δ 8.97 - 8.79 (m, 6H), 8.37 - 8.27 (m, 2H), 8.00 - 7.82 (m, 5H), 7.41 - 7.37 (m, 4H), 7.19 - 7.00 (m, 9H). HRMS (ESI): m / z [M - Cl] + calculated forC 37 H 26CIN4OZn: 641.1087; found: 641.1085.

[0131] Example 10:

[0132] A metal complex with high active oxygen generation efficiency, having a structure as shown in formula I-10:

[0133]

[0134] The metal complex with a structure as shown in formula I-10 in this example is marked as Zn(II)-TPY-TPA-Ph, which is prepared by the following steps:

[0135] (1) Take 45.0 mL of a mixed solvent of tetrahydrofuran and water (8:1 v / v), add p-bromobenzaldehyde (555.0 mg, 3.0 mmol), (4-(diphenylamino)phenyl)boronic acid (1012.1 mg, 3.5 mmol), and potassium carbonate (1382.1 mg, 10.0 mmol), and finally add tetrakis(triphenylphosphine)palladium (46.1 mg), and heat the mixture to 55°C under nitrogen protection for 12 h. After the reaction solution is cooled, the solvent is removed by distillation, then water and dichloromethane are added to extract the product, after the extraction is completed, dichloromethane is removed by distillation under reduced pressure, and the obtained product is purified by silica gel column chromatography, with petroleum ether: ethyl acetate (80:1 v / v) as the eluent, finally 911.7 mg of white powder compound C-4 is obtained, with a yield of 87.0%; the synthesis route is as follows:

[0136]

[0137] (2) Dissolve sodium hydroxide (240.0 mg, 6.0 mmol) in 40.0 mL of EtOH solution, then add 2-acetylpyridine (363.6 mg, 3.0 mmol), and place the mixture in 0°C for stirring for 10 min; then add compound C-4 (524.2 mg, 1.5 mmol) and continue stirring for 2 h; after 2 h, add excess ammonium acetate, and then heat the mixture to 60°C for reaction for 2 h; after the reaction solution is cooled, the solvent is removed by distillation, then water and dichloromethane are added to extract the product, after the extraction is completed, dichloromethane is removed by distillation under reduced pressure, and the obtained product is purified by silica gel column chromatography, with petroleum ether: ethyl acetate (5:1 v / v) as the eluent, finally 138.4 mg of white powder compound D-4 is obtained, marked as TPY-TPA-Ph, with a yield of 16.7%; the synthesis route is as follows:

[0138]

[0139] Characterization data: 1H NMR (400 MHz, CDC13) δ 8.80 (s, 2H), 8.74 (ddd, J = 4.8, 1.8, 0.8 Hz, 2H), 8.69 (d, J = 8.0 Hz, 2H), 7.98 (d, J = 8.4 Hz, 2H), 7.88 (td, J = 7.8, 1.8 Hz, 2H), 7.72 (d, J = 8.4 Hz, 2H), 7.56 (d, J = 8.6 Hz, 2H), 7.35 (ddd, J = 7.4, 4.8, 1.2 Hz, 2H), 7.33-7.26 (m, 4H), 7.20-7.14 (m, 6H), 7.09-7.02 (m, 2H). 13 CNMR (100 MHz, CDC13) δ 156.48, 156.11, 149.92, 149.28, 147.77, 147.71, 136.97, 134.28, 129.45, 127.89, 127.83, 127.16, 124.68, 123.93, 123.21, 121.51, 118.76. HRMS (ESI): m / z [M + H] + calculated for C 39 H 29 N4: 553.2392; found: 553.2382.

[0140] (3) TPY-TPA-Ph (55.3 mg, 0.1 mmol) and ZnCl2(13.6 mg, 0.1 mmol) were added to a mixed solvent of 10.0 mL dichloromethane and 5.0 mL methanol, and the mixture was stirred and coordinated at room temperature for 30 min; after the coordination reaction was completed, the solvent was removed by reduced pressure distillation, followed by adding water and dichloromethane to extract the product, and after the extraction was completed, dichloromethane was removed by reduced pressure distillation, and the obtained product was purified by silica gel column chromatography, and the eluent was dichloromethane:methanol (75:1 v / v), and finally 60.2 mg of yellow powder Zn(II)-TPY-TPA-Ph was obtained, with a yield of 89.4%; the synthetic route thereof is as follows:

[0141]

[0142] Characterization data: 1 H NMR (400 MHz, DMSO-d6) δ 9.17 (s, 2H), 9.06-8.99 (m, 2H), 8.90-8.87 (m, 2H), 8.39-8.37 (m, 4H), 7.99-7.88 (m, 4H), 7.81-7.78 (m, 2H), 7.41-7.31 (m, 4H), 7.12-7.10 (m, 8H). HRMS (ESI): m / z [M - Cl]+ Calcd for C 39 H 28 ClN4Zn: 651.1294; found: 651.1272.

[0143] Example 11:

[0144] A metal complex with high active oxygen generation efficiency, having a structure as shown in formula I-11:

[0145]

[0146] The metal complex with a structure as shown in formula I-11 in this example is marked as Zn(II)-TPY-DMA-DTZ, which is prepared by the following steps:

[0147] (1) Take 45.0 mL of a mixed solvent of tetrahydrofuran and water (8:1 v / v), add 5-bromo-2,2'-bithiophene-5'-carboxaldehyde (819.5 mg, 3.0 mmol), 4-dimethylaminobenzenboronic acid (577.5 mg, 3.5 mmol), and potassium carbonate (1382.1 mg, 10.0 mmol), and finally add tetrakis(triphenylphosphine)palladium (46.1 mg), and the mixture is heated to 55°C under nitrogen protection for 12 h. After the reaction solution is cooled, the solvent is removed by distillation, then water and dichloromethane are added to extract the product, after the extraction is completed, dichloromethane is removed by distillation under reduced pressure, and the obtained product is purified by silica gel column chromatography, with petroleum ether: ethyl acetate (100:1 v / v) as the eluent, finally 905.4 mg of white powder compound C-5 is obtained, with a yield of 96.3%; the synthesis route is as follows:

[0148]

[0149] (2) Dissolve sodium hydroxide (240.0 mg, 6.0 mmol) in 40.0 mL of EtOH solution, then add 2-acetylpyridine (363.6 mg, 3.0 mmol), and place the mixture in 0°C for stirring for 10 min; then add compound C-5 (469.5 mg, 1.5 mmol) and continue stirring for 2 h; after 2 h, add excess ammonium acetate, and then heat the mixture to 60°C for reaction for 2 h; after the reaction solution is cooled, the solvent is removed by distillation, then water and dichloromethane are added to extract the product, after the extraction is completed, dichloromethane is removed by distillation under reduced pressure, and the obtained product is purified by silica gel column chromatography, with petroleum ether: ethyl acetate (6:1 v / v) as the eluent, finally 120.4 mg of white powder compound D-5 is obtained, marked as TPY-DMA-DTZ, with a yield of 15.5%; the synthesis route is as follows:

[0150]

[0151] (3) TPY-DMA-DTZ (51.6 mg, 0.1 mmol) and ZnCl2(13.6 mg, 0.1 mmol) were added to a mixed solvent of 10.0 mL of dichloromethane and 5.0 mL of methanol, and the mixture was stirred and coordinated at room temperature for 30 min; after the coordination reaction was completed, the solvent was removed by distillation under reduced pressure, followed by adding water and dichloromethane to extract the product, and after the extraction was completed, dichloromethane was removed by distillation under reduced pressure. The obtained product was purified by silica gel column chromatography, and the eluent was dichloromethane:methanol (65:1 v / v). Finally, 58.6 mg of yellow powder Zn(II)-TPY-DMA-DTZ was obtained, with a yield of 90.1%. The synthetic route is as follows:

[0152]

[0153] Example 12:

[0154] A metal complex with high active oxygen generation efficiency, having a structure as shown in formula I-12:

[0155]

[0156] The metal complex with a structure as shown in formula I-12 in this example is marked as Zn(II)-TPY-TPE-DTZ, which is prepared by the following steps:

[0157] (1) A mixed solvent of tetrahydrofuran and water (8:1 v / v) 45.0 mL was taken, and 5-bromo-2,2'-bithiophene-5'-carboxaldehyde (819.5 mg, 3.0 mmol), (4-(1,2,2-triphenylvinyl)phenyl)boronic acid (1316.5 mg, 3.5 mmol) and potassium carbonate (1382.1 mg, 10.0 mmol) were added. Finally, tetrakis(triphenylphosphine)palladium (46.1 mg) was added, and the mixture was heated to 55°C under nitrogen protection for 12 h. After the reaction solution was cooled, the solvent was removed by distillation, followed by adding water and dichloromethane to extract the product. After the extraction was completed, dichloromethane was removed by distillation under reduced pressure. The obtained product was purified by silica gel column chromatography, and the eluent was petroleum ether:ethyl acetate (90:1 v / v). Finally, 1345.4 mg of white powder compound C-6 was obtained, with a yield of 85.6%. The synthetic route is as follows:

[0158]

[0159] (2) Dissolve sodium hydroxide (240.0 mg, 6.0 mmol) into 40.0 mL of EtOH solution, then add 2-acetylpyridine (363.6 mg, 3.0 mmol), and place the mixture into 0 °C to stir for 10 min; then add compound C-6 (786.5 mg, 1.5 mmol) and continue to stir the reaction for 2 h; after 2 h, add excess ammonium acetate, and then heat the mixture to 60 °C to react for 2 h; after the reaction solution is cooled, remove the solvent by distillation, then add water and dichloromethane to extract the product, after the extraction is completed, remove dichloromethane by distillation under reduced pressure, and purify the obtained product by silica gel column chromatography, using petroleum ether: ethyl acetate (7:1 v / v) as the eluent, to finally obtain 150.9 mg of white powder compound D-6, labeled as TPY-TPE-DTZ, with a yield of 13.8%; the synthesis route thereof is as follows:

[0160]

[0161] (3) Add TPY-TPE-DTZ (72.9 mg, 0.1 mmol) and ZnCl2(13.6 mg, 0.1 mmol) into a mixture of 10.0 mL of dichloromethane and 5.0 mL of methanol, and stir the mixture to coordinate at room temperature for 30 min; after the coordination reaction is completed, remove the solvent by distillation under reduced pressure, then add water and dichloromethane to extract the product, after the extraction is completed, remove dichloromethane by distillation under reduced pressure, and purify the obtained product by silica gel column chromatography, using dichloromethane: methanol (50:1 v / v) as the eluent, to finally obtain 69.6 mg of yellow powder Zn(II)-TPY-TPE-DTZ, with a yield of 80.1%; the synthesis route thereof is as follows:

[0162]

[0163] Example 13:

[0164] A metal complex with high active oxygen generation efficiency, having a structure as shown in formula I-13:

[0165]

[0166] The metal complex with a structure as shown in formula I-13 in this example is labeled as Zn(II)-TPY-TPA-DTZ-NO3, and the preparation method steps (1) and (2) thereof are the same as those of Example 1, and step (3) comprises the following steps:

[0167] TPY-TPA-DTZ (64.1 mg, 0.1 mmol) and Zn(NO3)2(18.9 mg, 0.1 mmol) were added to a mixed solvent of 10.0 mL of dichloromethane and 5.0 mL of methanol, and the mixture was stirred at room temperature for complexation for 30 min; after the completion of the complexation reaction, the solvent was removed by distillation under reduced pressure, followed by adding water and dichloromethane to extract the product, and after the completion of the extraction, dichloromethane was removed by distillation under reduced pressure, and the obtained product was purified by silica gel column chromatography using dichloromethane:methanol (65:1 v / v) as the eluent, and finally 72.0 mg of red powder Zn(II)-TPY-TPA-DTZ-NO3 was obtained, with a yield of 86.7%; the synthetic route thereof is as follows:

[0168]

[0169] Example 14:

[0170] A metal complex with high active oxygen generation efficiency, having a structure as shown in formula I-14:

[0171]

[0172] The metal complex with a structure as shown in formula I-13 in this example is marked as Zn(II)-TPY-TPA-DTZ-ACO, and the preparation method steps (1) and (2) thereof are the same as those of Example 1, and step (3) comprises the following steps:

[0173] TPY-TPA-DTZ (64.1 mg, 0.1 mmol) and Zn(OAc)2(18.9 mg, 0.1 mmol) were added to a mixed solvent of 10.0 mL of dichloromethane and 5.0 mL of methanol, and the mixture was stirred at room temperature for complexation for 30 min; after the completion of the complexation reaction, the solvent was removed by distillation under reduced pressure, followed by adding water and dichloromethane to extract the product, and after the completion of the extraction, dichloromethane was removed by distillation under reduced pressure, and the obtained product was purified by silica gel column chromatography using dichloromethane:methanol (65:1 v / v) as the eluent, and finally 72.0 mg of red powder Zn(II)-TPY-TPA-DTZ-ACO was obtained, with a yield of 86.7%; the synthetic route thereof is as follows:

[0174]

[0175] Experimental Example

[0176] The photophysical properties and the in vitro and in vivo photodynamic anticancer performance of the metal complexes with high active oxygen generation efficiency prepared in Examples 1-7 were investigated. Since the experimental results of the compounds in the application are similar, the reaction conditions are the same, therefore, the photophysical property determination and the in vitro and in vivo photodynamic anticancer performance are mainly taken as examples of the metal complexes Zn(II)-TPY-TPA-DTZ, Fe(III)-TPY-TPA-DTZ, Cr(III)-TPY-TPA-DTZ, Ni(II)-TPY-TPA-DTZ, Co(II)-TPY-TPA-DTZ, Mn(II)-TPY-TPA-DTZ, Cu(II)-TPY-TPA-DTZ prepared in Examples 1-7 of the application, and other examples are not listed one by one because the experimental results and reaction conditions are the same as the examples. The specific results are as follows:

[0177] 1. Photophysical property determination

[0178] 1.1 Absorption and emission determination of metal complexes

[0179] The absorption and emission spectra of the metal complexes Zn(II)-TPY-TPA-DTZ, Fe(III)-TPY-TPA-DTZ, Cr(III)-TPY-TPA-DTZ, Ni(II)-TPY-TPA-DTZ, Co(II)-TPY-TPA-DTZ, Mn(II)-TPY-TPA-DTZ, Cu(II)-TPY-TPA-DTZ and the compound D TPY-TPA-DTZ in the solvent dichloromethane (DCM) were determined, and the results are shown in Figure 1 Compared with the compound D TPY-TPA-DTZ (ligand), the maximum absorption / emission wavelength of the metal complexes Zn(II)-TPY-TPA-DTZ, Fe(III)-TPY-TPA-DTZ, Cr(III)-TPY-TPA-DTZ, Ni(II)-TPY-TPA-DTZ, Co(II)-TPY-TPA-DTZ, Mn(II)-TPY-TPA-DTZ, Cu(II)-TPY-TPA-DTZ is red shifted.

[0180] 1.2 Target molecule active oxygen determination

[0181] The ROS yield of the metal complexes Zn(II)-TPY-TPA-DTZ, Fe(III)-TPY-TPA-DTZ, Cr(III)-TPY-TPA-DTZ, Ni(II)-TPY-TPA-DTZ, Co(II)-TPY-TPA-DTZ, Mn(II)-TPY-TPA-DTZ, Cu(II)-TPY-TPA-DTZ and the commercially available photosensitizer Ce6 in solution was tested with 2,7-dichlorodihydrofluorescein diacetate (H2DCF-DA) detection probe.

[0182] In order to convert H2DCF-DA to 2,7-dichlorodihydrofluorescein (H2DCF), 0.25 mL of H2DCF-DA ethanol solution (1 mM) was added into 1 mL of NaOH (10 mM) aqueous solution, then stirred at room temperature for 30 min, and then adjusted to pH 7.4 with 5 mL of PBS solution, and the obtained solution was stored in a refrigerator for standby. The above-mentioned seven metal complexes and the commercially available photosensitizer Ce6 were added into the above-mentioned solution to make the final concentration of 2.5 μM, and the sample was placed in a fluorescence spectrometer, and the fluorescence intensity of the solution was tested every 5 s (λex: 488 nm), and the results were as follows Figure 2 A, Figure 2 In formula A, I0 is the emission intensity of H2DCF-DA at 525 nm, and I is the emission intensity of H2DCF-DA at 525 nm after the metal complex photosensitizer is added.

[0183] From Figure 2 As can be seen from formula A, the fluorescence intensity of the control group (only H2DCF-DA) at 525 nm does not increase with the extension of light irradiation time; and the fluorescence intensity of the metal complexes Zn(II)-TPY-TPA-DTZ, Fe(III)-TPY-TPA-DTZ, Cr(III)-TPY-TPA-DTZ, Ni(II)-TPY-TPA-DTZ, Co(II)-TPY-TPA-DTZ, Mn(II)-TPY-TPA-DTZ, Cu(II)-TPY-TPA-DTZ and the commercially available photosensitizer Ce6 is increased by 76, 46, 8, 53, 51, 25, 28 and 11 times respectively compared with the initial fluorescence intensity after 60 s of light irradiation; it is confirmed that the seven metal complexes prepared in the application can effectively and quickly generate ROS under light irradiation, and have stronger total active oxygen generation efficiency compared with the commercially available photosensitizer Ce6.

[0184] 1.3 Singlet oxygen yield determination

[0185] ABDA (9,10-anthryl-bis(methylene)dipropionic acid) was used as an indicator to investigate the singlet oxygen generation performance of the metal complexes Zn(II)-TPY-TPA-DTZ, Fe(III)-TPY-TPA-DTZ, Cr(III)-TPY-TPA-DTZ, Ni(II)-TPY-TPA-DTZ, Co(II)-TPY-TPA-DTZ, Mn(II)-TPY-TPA-DTZ, Cu(II)-TPY-TPA-DTZ and the commercially available photosensitizer Ce6 under light irradiation. When ABDA and 1When O2 reaction, ABDA is oxidized into peroxide bridge structure, so that the absorbance value of ABDA at 378 nm decreases, and the speed of decrease can indirectly reflect the O2 yield of photosensitizer under light. 1 First, the absorbance of target molecule is set as blank. Then, ABDA (50 μM) is mixed with target molecule (2.5 μM) solution under dark condition, and the absorbance value of solution is immediately determined, then the solution mixture is irradiated with white light (5 mW / cm 2 ) for a certain time, and the absorbance value of solution is immediately recorded after each irradiation, and the results are shown in Figure 2 B, Figure 2 B, A0 is the initial absorbance of ABDA at 378 nm, and A is the absorbance of ABDA at 378 nm after adding metal complex photosensitizer under different irradiation time.

[0186] As can be seen from Figure 2 B, with the extension of irradiation time, the absorbance value of control group (only ABDA exists) at 378 nm under light does not decrease, while the absorbance of ABDA of the seven metal complexes Zn(II)-TPY-TPA-DTZ, Fe(III)-TPY-TPA-DTZ, Cr(III)-TPY-TPA-DTZ, Ni(II)-TPY-TPA-DTZ, Co(II)-TPY-TPA-DTZ, Mn(II)-TPY-TPA-DTZ, Cu(II)-TPY-TPA-DTZ and commercial photosensitizer Ce6 decreases to 0.1%, 86.2%, 85.4%, 61.5%, 43.6%, 0.7%, 83.7%, 78.9% respectively after 1200 seconds of irradiation; it is confirmed that the metal complexes prepared in examples 1-7 can all produce singlet oxygen under light, and Zn(II)-TPY-TPA-DTZ, Mn(II)-TPY-TPA-DTZ, Ni(II)-TPY-TPA-DTZ and Co(II)-TPY-TPA-DTZ have more excellent singlet oxygen production efficiency than commercial photosensitizer Ce6, and Fe(III)-TPY-TPA-DTZ, Cr(III)-TPY-TPA-DTZ and Cu(II)-TPY-TPA-DTZ have similar singlet oxygen production efficiency to Ce6.

[0187] 1.4 Hydroxyl radical yield determination

[0188] HPF (hydroxyphenyl fluorescein) was used as an indicator to investigate the hydroxyl radical generation performance of the metal complexes prepared in Examples 1 to 7 of the present invention and the commercially available photosensitizer Ce6 under light irradiation. Hydroxyphenyl fluorescein itself has no fluorescence, but when it reacts with hydroxyl radicals, peroxynitrite anions and hypochlorite anions, it will produce strong green fluorescence at 515nm. Under dark conditions, 5mM HPF stock solution was added to 3mL PBS buffer to make the HPF concentration 10μM, and then the target molecule was added to make its concentration 2.5μM. Finally, the 5mW / cm 2 Under the irradiation of incandescent lamp, the changes of sample fluorescence emission at different times were recorded. The results are as follows: Figure 2 C, Figure 2 In C, I0 is the emission intensity of HPF at 515 nm, and I is the emission intensity of HPF at 515 nm under different irradiation times after the addition of metal complex photosensitizer.

[0189] Depend on Figure 3 C shows that with the extension of illumination time, the fluorescence intensity of the control group (only HPF) did not increase significantly, while the fluorescence intensity of HPF of the seven metal complexes Zn(Ⅱ)-TPY-TPA-DTZ, Fe(Ⅲ)-TPY-TPA-DTZ, Cr(Ⅲ)-TPY-TPA-DTZ, Ni(Ⅱ)-TPY-TPA-DTZ, Co(Ⅱ)-TPY-TPA-DTZ, Mn(Ⅱ)-TPY-TPA-DTZ, Cu(Ⅱ)-TPY-TPA-DTZ and commercially available photosensitizer Ce6 after 1200 seconds of illumination was enhanced by 142, 80, 11, 198, 100, 7, 13 and 3 times respectively compared with the initial fluorescence intensity; it is confirmed that these seven metal complexes can quickly and efficiently generate hydroxyl radicals under illumination.

[0190] From the experimental results of 1.1 to 1.4, it can be seen that the metal complexes prepared in Examples 1 to 7 of the present invention have good fluorescence emission performance and excellent reactive oxygen species generation efficiency, and can generate a large amount of singlet oxygen and hydroxyl radicals under light. Among them, Zn(Ⅱ)-TPY-TPA-DTZ has near-infrared emission and the strongest total reactive oxygen species generation efficiency, showing the application potential in efficient photodynamic anti-cancer.

[0191] 2. In vitro photodynamic anti-tumor experiment

[0192] 2.1 Cell culture

[0193] 2.1.1 Cell recovery

[0194] The frozen mouse breast cancer cells (4T1), human cervical cancer cells (HeLa), human colon cancer cells (HCT116) and human liver cancer cells (HepG2) were taken out from liquid nitrogen, and were continuously shaken in a 37°C water bath to promote their melting. The cells were transferred into 5 mL centrifuge tubes, 3 mL of preheated corresponding medium was added, centrifuged at 800 rpm for 3 min, the supernatant was discarded, 2 mL of medium was added, and the cells were gently blown and inoculated into a culture bottle, which was cultured in a cell incubator containing 5% CO2.

[0195] 2.1.2 Cell passage

[0196] The mouse breast cancer cells (4T1), human cervical cancer cells (HeLa), human colon cancer cells (HCT116) and human liver cancer cells (HepG2) cultured for 24 h were taken out, and the cell morphology and density were observed under a microscope. Then, the cell surface was washed with 3 mL of PBS, and the PBS was discarded. The washing was repeated twice. The cells were digested with 1 mL of trypsin for 2 min, and the digestion time of different cells was slightly different. During the digestion, the cells were placed in a 37°C cell incubator for digestion. After complete digestion, 2 mL of corresponding medium was added to stop the digestion, and the cells were blown to be uniform. The cells were transferred into a centrifuge tube, centrifuged at 1000 rpm for 5 min. After centrifugation, the supernatant was discarded, the bottle opening was wiped with alcohol, and 2 mL of corresponding medium was added to blow and mix the cells (blown along the wall for about 20 times). Each 1 mL was inoculated into a culture bottle, a total of two bottles, and cultured in a cell incubator containing 5% CO2. After three generations of cell passage, the cells were plated for cell toxicity test when the cells grew to the logarithmic growth phase.

[0197] 2.2 Intracellular reactive oxygen species imaging

[0198] The human cervical cancer cells (HeLa) were inoculated into a laser confocal dish, and 10 μM of Zn(II)-TPY-TPA-DTZ prepared in Example 1 of the application was added for incubation for 12 h. The cells were irradiated for 1 h under dark conditions or white light (5 mW / cm 2 ), and then H2DCF-DA (50 μM) or HPF (10 μM) was added for incubation for 30 min. After washing with PBS for three times, confocal imaging was performed (H2DCF-DA and HPF: λ ex = 488 nm, λ em = 500-550 nm), and the results are shown in Figure 3 .

[0199] As shown in Figure 3A shows that after incubating Zn(Ⅱ)-TPY-TPA-DTZ with cells for 12 hours and then irradiating with white light for 1 hour, and then adding 50μM H2DCF-DA and incubating for 30 minutes, a green fluorescence signal appeared in the cells. However, under dark conditions, no green fluorescence signal was observed in the cells. The results show that Zn(Ⅱ)-TPY-TPA-DTZ can produce reactive oxygen species in cells under light. Figure 4 As shown in Figure B, after adding Zn(II)-TPY-TPA-DTZ and incubating the cells with light for 1 hour, and then adding 10 μM HPF and incubating for 30 minutes, the cells showed a green fluorescence signal, indicating the generation of hydroxyl radicals in the cells. In contrast, no green fluorescence signal from HPF was observed in the cells under dark conditions.

[0200] In summary, the experimental results show that Zn(Ⅱ)-TPY-TPA-DTZ can effectively produce reactive oxygen species in cells under light irradiation and has the potential for photodynamic therapy.

[0201] 2.3 In vitro photodynamic anticancer activity

[0202] First, mouse breast cancer cells (4T1), human liver cancer cells (HepG2), human colon cancer cells (HCT116), and human cervical cancer cells (HeLa) were digested with trypsin and inoculated into 96-well plates for 12 hours. The culture medium was replaced with 100 μL of DMEM culture medium containing Zn(Ⅱ)-TPY-TPA-DTZ prepared in Example 1. The 96-well plates were then placed in a cell culture incubator at 37°C and 5% CO2 and cultured for 12 hours. The culture medium containing Zn(Ⅱ)-TPY-TPA-DTZ was then discarded, and drug-free complete culture medium was added. Finally, the 96-well plates were illuminated for 60 minutes and then placed in a cell culture incubator for further culturing for 24 hours. After 24 hours, 100 μL of culture medium containing 10% CCK-8 was added to each well of the 96-well plate, and the absorbance OD at 450 nm was measured after incubation in a cell culture incubator for 30 minutes. 450 nm The calculation formula of cell survival rate is as follows:

[0203]

[0204] Where, OD 450nm样品 OD is the absorbance of the wells containing cells, CCK8 solution and drug solution; 450nm空白 OD is the absorbance of wells with culture medium and CCK-8 solution but no cells; 450nm不加药 The absorbance of the wells containing cells and CCK-8 solution but no drug solution; the measured OD value is the average value of 3 independent parallel samples, and the results are expressed as mean ± standard deviation.

[0205] The results are as followsFigure 4 As shown in Figure 5 As shown in Table A-D, the survival rates of the four cancer cells (4T1, HepG2, HCT116 and HeLa) were all above 90% after the dark treatment of 2.5-10 μM Zn(II)-TPY-TPA-DTZ for 12 h, indicating that Zn(II)-TPY-TPA-DTZ had low dark toxicity and good biocompatibility. When irradiated with white light for 1 h, Zn(II)-TPY-TPA-DTZ showed excellent photodynamic therapy effect, and the survival rates of the cancer cells decreased significantly with the increase of the concentration. The survival rates of the cancer cells treated with 10 μM Zn(II)-TPY-TPA-DTZ were all below 20%, showing strong in vitro photodynamic anticancer effect and dose dependence.

[0206] In summary, the experimental results showed that Zn(II)-TPY-TPA-DTZ had no obvious toxicity to mouse breast cancer cells (4T1), human liver cancer cells (HepG2), human colon cancer cells (HCT116) and human cervical cancer cells (HeLa) in the dark, but could efficiently kill various cancer cells under light.

[0207] 2.4 Intracellular reactive oxygen species imaging test under hypoxic (0.1% O2) conditions

[0208] Human cervical cancer cells (HeLa) were inoculated in laser confocal dishes, and the laser confocal dishes were placed in an anaerobic gas bag containing an anaerobic gas bag before Zn(II)-TPY-TPA-DTZ prepared in Example 1 was added. After 3 h of hypoxic culture (0.1% O2), the culture solution was replaced with 1 mL of Zn(II)-TPY-TPA-DTZ at a concentration of 10 μM, and then the culture was continued in the hypoxic environment for another 12 h. The Zn(II)-TPY-TPA-DTZ-containing culture medium was discarded, and drug-free complete culture medium was added. The cells were irradiated with white light (5 mW / cm2) for 1 h in the dark or white light (5 mW / cm2) for 1 h, and then H2DCF-DA (50 μM incubation for 30 min, washed with PBS for three times, and then confocal imaging (H2DCF-DA: λex= 488 nm, λem= 500-550 nm) was performed. The results are shown in Figure 5 .

[0209] As shown in Table A-D, the survival rates of the four cancer cells (4T1, HepG2, HCT116 and HeLa) were all above 90% after the dark treatment of 2.5-10 μM Zn(II)-TPY-TPA-DTZ for 12 h, indicating that Zn(II)-TPY-TPA-DTZ had low dark toxicity and good biocompatibility. When irradiated with white light for 1 h, Zn(II)-TPY-TPA-DTZ showed excellent photodynamic therapy effect, and the survival rates of the cancer cells decreased significantly with the increase of the concentration. The survival rates of the cancer cells treated with 10 μM Zn(II)-TPY-TPA-DTZ were all below 20%, showing strong in vitro photodynamic anticancer effect and dose dependence. Figure 6As can be seen, under hypoxic conditions, after incubation of Zn(II)-TPY-TPA-DTZ with cells for 12 h, white light irradiation for 1 h, and then addition of 50 μM of H2DCF-DA for incubation for 30 min, green fluorescent signals appeared in the cells. However, under dark conditions, no green fluorescence was observed in the cells. The results show that Zn(II)-TPY-TPA-DTZ can produce reactive oxygen species in the cells under light irradiation.

[0210] In summary, the experimental results show that Zn(II)-TPY-TPA-DTZ can effectively produce reactive oxygen species in the cells under hypoxic conditions and light irradiation, indicating that it has the potential to overcome the hypoxic microenvironment of solid tumors for photodynamic therapy.

[0211] 2.5 Photodynamic anticancer activity under hypoxic (0.1% O2) conditions

[0212] Mouse breast cancer cells (4T1) were inoculated in 96-well plates for 12 h. Before addition of Zn(II)-TPY-TPA-DTZ prepared in Example 1 or the commercially available photosensitizer Ce6, the 96-well plates were placed in an anaerobic gas bag equipped with an anaerobic gas bag, and incubated under hypoxic conditions (0.1% O2) for 3 h. Then, the culture solution was replaced with 100 μL of Zn(II)-TPY-TPA-DTZ or the commercially available photosensitizer Ce6 at a concentration of 10 μM, and the incubation was continued under hypoxic conditions for 12 h. The culture medium containing Zn(II)-TPY-TPA-DTZ or the commercially available photosensitizer Ce6 was discarded, and drug-free complete culture medium was added. After light irradiation for 1 h under hypoxic conditions, the incubation was continued under normoxic conditions (21% O2) for 24 h. After 24 h, 100 μL of culture medium containing 10% CCK-8 was added to each well of the 96-well plate, and the absorbance at 450 nm (OD450) was measured after incubation in a cell incubator for 30 min. 450 nm The formula for calculating the cell survival rate was the same as above.

[0213] The results are shown in Table 1. Figures 7-8As shown, under hypoxic (0.1% O2) conditions, after 1 hour of white light irradiation, Zn(Ⅱ)-TPY-TPA-DTZ has excellent photodynamic therapy effect, and the survival rate of cancer cells also decreases significantly with increasing concentration. The survival rate of cancer cells after 10μM Zn(Ⅱ)-TPY-TPA-DTZ light treatment is below 20%, which is similar to the photodynamic therapy effect under normoxic (21% O2) conditions, showing strong in vitro photodynamic anticancer efficacy and dose dependence. The commercially available photosensitizer Ce6 has excellent photodynamic therapy effect under normoxic conditions (21% O2), but its photodynamic therapy effect is significantly reduced under hypoxic (0.1% O2) conditions. The above experimental results show that Zn(Ⅱ)-TPY-TPA-DTZ can effectively overcome the hypoxic microenvironment and achieve efficient photodynamic therapy.

[0214] 2.6 Live-dead staining analysis

[0215] Mouse breast cancer cells (4T1) were seeded in a 96-well plate and cultured for 12 hours. DMEM medium containing 10 μM Zn(Ⅱ)-TPY-TPA-DTZ and commercially available photosensitizer Ce6 was then added and incubated with the 4T1 cells for 12 hours under hypoxia (0.1% O2) / normoxia (21% O2). The medium containing Zn(Ⅱ)-TPY-TPA-DTZ and Ce6 was discarded, and drug-free complete medium was added. The cells were then illuminated for 1 hour and washed 2 to 3 times with PBS. The 4T1 cells were then stained with a Calcein-AM / Propidium Iodide (PI) kit for 40 minutes, washed 2 to 3 times with PBS, and finally photographed and analyzed using a fluorescence microscope. The results are shown in Figure 2. Figure 9 As shown, under normoxic (21% O₂) conditions, neither Zn(Ⅱ)-TPY-TPA-DTZ nor the commercially available photosensitizer Ce6 showed a green fluorescence signal in the Calcein-AM channel after illumination, while a strong red fluorescence signal appeared in the PI channel. This indicates that both Zn(Ⅱ)-TPY-TPA-DTZ and the commercially available photosensitizer Ce6 exhibited excellent in vitro photodynamic antitumor activity under normoxic conditions. However, under hypoxic (0.1% O₂) conditions, neither Zn(Ⅱ)-TPY-TPA-DTZ showed a green fluorescence signal in the Calcein-AM channel after illumination, but a strong red fluorescence signal appeared in the PI channel. Ce6 showed a strong green fluorescence signal in the Calcein-AM channel after illumination, but a small amount of red fluorescence signal appeared in the PI channel. These results indicate that Zn(Ⅱ)-TPY-TPA-DTZ can achieve highly efficient photodynamic therapy in both normoxic and hypoxic environments, outperforming the commercially available photosensitizer Ce6.

[0216] 3 Biocompatibility experiments

[0217] 3.1 Hemolysis assay

[0218] Firstly, 2% red blood cell suspension was prepared: fresh mouse blood 10 mL was taken in a vacuum blood collection tube coated with sodium heparin, gently inverted and mixed, after mixing, centrifuged at 1000 rpm for 10 min, the supernatant was removed, and the red blood cells (RBC) were quantitatively taken by a pipette, washed with about 10 times the volume of RBC sterile phosphate buffer PBS for 3 times, and then diluted with sterile phosphate buffer PBS to prepare a 2% RBC suspension. 20 μL of corresponding volume of Zn(II)-TPY-TPA-DTZ stock solution was added to 180 μL of 2% RBC suspension to obtain a final concentration of 1, 2, 4, 6, 8, 10, 20 μM experimental solution, while the negative control group and the positive control group were added with 20 μL of PBS and Triton X-100 solvent respectively. Each group had 3 parallel holes, and after gentle mixing, it was incubated at 37°C water bath for 1 h, and then centrifuged at 1200 rpm for 5 min. The supernatant was collected, and the ultraviolet absorption at 545 nm was detected by enzyme labeling, and the hemolysis percentage of RBC was calculated according to the following formula:

[0219]

[0220] In the formula, OD 样品 is the absorbance of red blood cell suspension added with Zn(II)-TPY-TPA-DTZ; OD 阳性对照 is the absorbance of red blood cell suspension added with Triton X-100; OD 阴性对照 is the absorbance of red blood cell suspension added with PBS.

[0221] The results are shown in Table 1. Figure 10 After the addition of photosensitizer Zn(II)-TPY-TPA-DTZ to the red blood cell suspension, even if the concentration increased to 20 μM, the hemolysis rate was still less than 5%, indicating that Zn(II)-TPY-TPA-DTZ had good biocompatibility.

[0222] 4 In vivo tumor targeting experiment

[0223] 6-week-old SPF level 20 BALB / c female mice were provided by Chengdu Dashuo Experimental Animal Co., Ltd., and were adapted to the experimental environment for one week before the experiment. The logarithmic growth period of mouse breast cancer cells (4T1) was digested with trypsin, and the 4T1 cells were diluted to 1x10 7 After the right front limb of the mouse was depilated, the right front limb of the BALB / c mouse was first disinfected with 75% alcohol, and 100 μL of the above cells were inoculated subcutaneously in the right front limb of the mouse to model. The right front limb of the mouse was touched every day to observe whether there was a tumor, and the length of the long diameter (A) and the short diameter (B) of the tumor was measured with a vernier caliper, and the volume was calculated according to the following formula: V = (AxB 2) / 2 to calculate the tumor volume, until the tumor grew to 50 mm 3 ~ 80 mm 3 ~ 80 mm 7 The mice were anesthetized with isoflurane, and the in vivo and ex vivo fluorescence imaging of the five organs (heart, liver, spleen, lung, kidney) and tumor were performed at 1, 2, 4, 8, 12, 24, 36, 48, 72 h after the administration of Zn(II)-TPY-TPA-DTZ (1 mM, 100 μL) by tail vein injection, with an excitation wavelength of 465 nm and a fluorescence collection at 700 nm.

[0224] The results are shown in Figure 6. After 8 h of tail vein injection of the metal complex photosensitizer Zn(II)-TPY-TPA-DTZ, Zn(II)-TPY-TPA-DTZ began to accumulate in the tumor site, and the fluorescence intensity at the tumor site was the strongest at 24 h. Until 72 h, Zn(II)-TPY-TPA-DTZ was still accumulated in the tumor cells. The above results show that 24 h after the tail vein injection of Zn(II)-TPY-TPA-DTZ, Zn(II)-TPY-TPA-DTZ presents the highest degree of enrichment in the tumor tissue, indicating that Zn(II)-TPY-TPA-DTZ can be targeted to the tumor site. Figures 11-13

[0225] 5 In vivo photodynamic anticancer performance test

[0226] SPF level 24 BALB / c female mice at 6 weeks of age were provided by Chengdu Dashuo Experimental Animal Co., Ltd. and were adapted to the experimental environment for one week before the experiment. The logarithmic growth period of mouse breast cancer cells (4T1) was digested with trypsin, and the 4T1 cells were diluted to 1 x 10 7 After the right forelimb of the mouse was depilated, the right forelimb of the BALB / c mouse was first disinfected with 75% alcohol, and 100 μL of the above cells were inoculated subcutaneously in the right forelimb of the mouse. The right forelimb of the mouse was touched daily to observe whether there was a tumor, and the length of the long diameter (A) and the short diameter (B) of the tumor was measured with a vernier caliper, and the tumor volume was calculated according to the following formula: V = (A x B 2 ) / 2, until the tumor grew to 50 mm 3 ~ 60 mm 3The model was successfully established. The established mice were divided into four groups for treatment: (1) PBS group; (2) Zn(Ⅱ)-TPY-TPA-DTZ group; (3) Ce6+L group; (4) Zn(Ⅱ)-TPY-TPA-DTZ+L group. Each group consisted of 6 mice, and 100 μL of PBS, Zn(Ⅱ)-TPY-TPA-DTZ (1 mM) or commercially available photosensitizer Ce6 (1 mM) was injected through the tail vein every two days. 24 hours after the injection of drugs in the Ce6+L group and the Zn(Ⅱ)-TPY-TPA-DTZ+L group, the power was 20 mW / cm 2 The incandescent lamp was used to irradiate the tumor site of the mouse for 1 hour. Every two days, the tumor growth and weight of the mouse were recorded, and the long and short diameters of the mouse tumor were measured. The tumor volume was calculated according to the above formula. On the 18th day of treatment, the mice were killed and the tumors and major organs were dissected and separated. The organs and tumors were preserved in 4% paraformaldehyde fixative and sent to Chengdu Lilai Biotechnology Co., Ltd. for hematoxylin-eosin (H&E) staining of the tumors and major organs and tumor Ki67 immunohistochemical analysis. The results are as follows Figures 11-13 As shown, Figure 11 (A) are all PBS groups, (B) are all Zn(Ⅱ)-TPY-TPA-DTZ groups, (C) are all Ce6+L groups, and (D) are all Zn(Ⅱ)-TPY-TPA-DTZ+L groups.

[0227] from Figure 12 As can be seen in the figure, on the 18th day of treatment, there was no significant difference in tumor size between the Zn(Ⅱ)-TPY-TPA-DTZ group and the PBS group, indicating that injection of Zn(Ⅱ)-TPY-TPA-DTZ alone could not inhibit tumor growth. Compared with the PBS group, tumor growth was inhibited in the Ce6+L group and the Zn(Ⅱ)-TPY-TPA-DTZ+L group, especially in the Zn(Ⅱ)-TPY-TPA-DTZ+L group, where the tumor completely disappeared.

[0228] like Figure 12 As shown in A, on the 18th day of treatment, compared with the initial tumor size, the tumor volume of the PBS group increased by nearly 13.9 times; the tumor volume of the Zn(Ⅱ)-TPY-TPA-DTZ group increased by 12.9 times; the tumor volume of the Ce6+L group increased by 6.4 times; and the tumor of the Zn(Ⅱ)-TPY-TPA-DTZ+L group had almost no growth. Figure 12 Figure B shows the weight of the in vitro tumors of mice in each group on treatment day 18. The tumor weights of the PBS group and the Zn(Ⅱ)-TPY-TPA-DTZ group were similar, while the tumor weights of the Ce6+L group and the Zn(Ⅱ)-TPY-TPA-DTZ+L group were significantly reduced compared to the PBS group, and the tumor weight of the Zn(Ⅱ)-TPY-TPA-DTZ+L group was much smaller than that of the Ce6+L group.

[0229] The above results show that Zn(II)-TPY-TPA-DTZ has good photodynamic therapy efficiency in vivo, and the anti-tumor effect is better than that of the commercially available photosensitizer Ce6.

[0230] As shown in Figure 13 C, the weight change of each group of mice during treatment is shown, and it can be seen from the figure that the weight of each group of mice during treatment does not change significantly, indicating that Zn(II)-TPY-TPA-DTZ has good biocompatibility.

[0231] The anti-tumor effect of the four groups of mice was investigated by H&E staining and Ki67 immunohistochemical analysis of tumor sections of each group of mice; the results are shown in Figure 13 A, compared with the PBS group and the Zn(II)-TPY-TPA-DTZ group, the Zn(II)-TPY-TPA-DTZ+L group has very obvious tumor tissue necrosis, and the Ki67 expression level of the Zn(II)-TPY-TPA-DTZ+L group is the lowest. The results show that Zn(II)-TPY-TPA-DTZ has excellent photodynamic therapy efficiency in vivo, and the anti-tumor effect is better than that of the commercially available photosensitizer Ce6. Finally, the H&E staining analysis of the ex vivo organ (heart, liver, spleen, lung and kidney) sections of each group of mice was performed, and the results are shown in ​ B, the H&E staining of the organs of the four groups of mice did not show obvious differences and tissue necrosis; indicating that Zn(II)-TPY-TPA-DTZ has good biocompatibility.

[0232] In summary, the experimental results show that Zn(II)-TPY-TPA-DTZ has good biocompatibility and excellent photodynamic anticancer effect in vivo, and is better than the commercially available photosensitizer Ce6.

[0233] The above only describes the preferred embodiments of the present application, and is not intended to limit the present application. Any modification, equivalent replacement, improvement, etc. made within the spirit and principle of the present application shall be included in the protection scope of the present application.

Claims

1. A metal complex having high active oxygen generation efficiency, characterized in that: The specific structure is as follows: 、 、 、 、 、 、 。 2. The method for preparing a metal complex having high active oxygen generation efficiency according to claim 1, characterized in that: The following steps are involved: (1) Compound A, compound B, a palladium catalyst, and an alkaline substance are dissolved in a solvent, and reacted at 50-60° C. under an inert atmosphere for 11-13 hours to obtain compound C; (2) Dissolve 2-acetylpyridine and sodium hydroxide in an organic solvent, add compound C and react at -3~3°C for 1.5~2.5 hours, then add ammonium acetate and react at 55~65°C for 1.5~2.5 hours to obtain compound D; (3) Compound D and MX a Dissolve in a solvent and react for 25 to 35 minutes to obtain metal complexes with high active oxygen generation efficiency as shown in Formulas I-1 to I-7. The synthesis route is as follows: ； In the compound A, R2 is halogen; in the synthetic route, for ; R1 is ; M is Cr 3 + 、Mn 2+ 、Fe 3+ 、Co 2+ 、Ni 2+ 、Cu 2+ or Zn 2+ ; X is a chloride ion; a is 2 or 3.

3. The method for preparing a metal complex with high active oxygen generation efficiency according to claim 2, characterized in that: The solvent in step (1) is a mixed solvent of tetrahydrofuran and water, and the volume ratio of tetrahydrofuran to water in the solvent is (7-9):1; the organic solvent in step (2) is anhydrous ethanol; and the solvent in step (3) is a mixed solvent of methanol and dichloromethane, and the volume ratio of methanol to dichloromethane in the solvent is 1:(2-3).

4. The method for preparing a metal complex with high active oxygen generation efficiency according to claim 2, wherein: The molar ratio of the compound A, the compound B, the palladium catalyst and the alkaline substance is 1:(1.2-1.5):(0.04-0.05):(3.5-4).

5. The method for preparing a metal complex with high active oxygen generation efficiency according to claim 2 or 4, characterized in that: The palladium catalyst is tetrakis(triphenylphosphine)palladium, bis(triphenylphosphine)palladium chloride, [1,1'-bis(diphenylphosphino)ferrocene]palladium dichloride, tris(dibenzylideneacetone)dipalladium or palladium acetate; the alkaline substance is sodium tert-butoxide, potassium tert-butoxide, sodium carbonate, potassium carbonate, cesium carbonate, potassium phosphate or barium hydroxide.

6. The method for preparing a metal complex with high active oxygen generation efficiency according to claim 2, wherein: In the step (2), the molar ratio of compound C, 2-acetylpyridine, sodium hydroxide and ammonium acetate is 1.0:(2.0-2.5):(3.5-4.0):(30.0-40.0).

7. The method for preparing a metal complex with high active oxygen generation efficiency according to claim 2, characterized in that: In the step (3), compound D and MX a The molar ratio is 1:(1~1.2).

8. Use of the metal complex with high active oxygen generation efficiency according to claim 1 in the preparation of a photodynamic anticancer photosensitizer, wherein the cancer is breast cancer, liver cancer, colon cancer and cervical cancer.

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