Carbon dot assembly and preparation method and application thereof

Abstract

The invention belongs to the technical field of carbon nanomaterials, and particularly relates to a carbon dot assembly and a preparation method and application thereof. Compared with a traditional carbon dot, the donor-acceptor structure promotes the charge separation process in the carbon dot assembly, so that electrons can be transferred from an inner core serving as an electron donor to an interface serving as an electron acceptor, and then the electrons act with surrounding oxygen to promote generation of superoxide free radicals and photogenerated holes. The carbon dot assembly provided by the invention has excellent properties, and can specifically generate I-type active oxygen and oxidize important biological substrates through photogenerated holes, so that efficient anti-tumor photodynamic therapy is realized. In addition, the particle size of the carbon dot assembly is larger, so that the carbon dot assembly can be targeted to tumor tissues through enhanced permeability and retention effect of tumor tissue characteristics, and further efficient tumor killing is realized.

Description

Technical Field

[0001] The present invention belongs to the technical field of carbon nanomaterials, and in particular relates to a carbon dot assembly and a preparation method and application thereof. Background Art

[0002] Photodynamic therapy is an effective tumor treatment method with the advantages of low side effects, controllable treatment time and space, good tissue selectivity and low trauma. Photodynamic therapy relies on the reactive oxygen species produced by the reaction of photosensitizers with surrounding substrates or oxygen under light radiation to work. According to the type of reactive oxygen species produced, photodynamic therapy can be divided into type I photodynamic therapy and type II photodynamic therapy. Type I photodynamic therapy mainly produces type I reactive oxygen species, such as hydroxyl free radicals and superoxide free radicals, through electron or proton transfer; type II photodynamic therapy mainly produces type II reactive oxygen species, such as singlet oxygen, through charge transfer. Compared with type II photodynamic therapy, type I photodynamic therapy is less dependent on oxygen. Considering the hypoxic characteristics of the tumor tissue microenvironment, type I photodynamic therapy is more beneficial to the treatment of tumors. In addition, the reactive oxygen species produced by photodynamic therapy can also effectively kill bacteria. Compared with traditional antibacterial therapy, photodynamic sterilization does not produce drug resistance.

[0003] Carbon dots (CDs), a zero-dimensional photoluminescent carbon nanomaterial with a size of less than 10 nm, were first discovered in 2004. Due to their simple preparation, readily available raw materials, low cost, good water solubility and biocompatibility, they are widely used in various fields such as drug delivery, photothermal therapy of tumors and photodynamic therapy. However, in the field of tumor photodynamic therapy, most of the carbon dots reported so far produce singlet oxygen and exert type II photodynamic therapy effects, but have poor therapeutic effects on tumors with hypoxic microenvironments. Effective charge transfer is the key to the generation of reactive oxygen species. By enhancing charge transfer, the ability of molecules or particles to produce type I reactive oxygen species can be significantly improved. However, there is currently a lack of effective strategies to enhance charge transfer within carbon dots.

[0004] Therefore, there is an urgent need to develop a strategy to promote charge transfer within carbon dots so that they can specifically generate type I reactive oxygen species and further apply them to photodynamic therapy of tumors and microbial infections with hypoxic microenvironments.

[0005] In view of this, the present invention is proposed. Summary of the invention

[0006] The present invention aims to solve at least one of the above-mentioned technical problems existing in the prior art.

[0007] For this reason, the principle of the present invention is:

[0008] The present invention provides a donor-acceptor type carbon dot assembly, the structure of which has carbon dots aggregated in the inner core as electron donors and interfaces formed by melted molecules containing electron-withdrawing groups as electron acceptors, and the interfaces between different carbon dots and the outer layers of the carbon dots are both composed of melted molecules containing electron-withdrawing groups. Charge separation can occur efficiently inside the carbon dot assembly of this structure, promoting light-induced electron transfer. Under illumination conditions, excited electrons are transferred to the interface of the carbon dot assembly and further transferred to the surrounding oxygen, generating a large number of superoxide free radicals. At the same time, the photogenerated holes are stabilized in the core of the carbon dot assembly, have a certain oxidation capacity, and can oxidize important biological substrates. By obtaining the above carbon dot assembly through this donor-acceptor strategy, type I reactive oxygen - superoxide free radicals can be specifically generated, promoting the application of carbon dots in tumor treatment.

[0009] It can be seen that the core of the present invention is the electron transfer between the carbon dot core and the electron acceptor material interface, rather than the material composition of the carbon dot, or the particle size, or the specific type of electron acceptor material. Those skilled in the art should know that the carbon dots prepared by other methods can also be prepared into carbon dot assemblies by the method of the present invention. Other types of solvents can also be used to prepare the carbon dots. For example, formic acid carbon nanodots, acetic acid carbon nanodots, and acetone carbon nanodots prepared by different solvents in CN116173209A can also be used to prepare the carbon dot assembly of the present invention. Further, carbon dots prepared by other raw materials except citric acid and urea can also be used to prepare the carbon dot assembly of the present invention. Similarly, electron acceptor materials with other electron-withdrawing groups, through the preparation method of the present invention, form carbon dot assemblies with similar electron acceptor material interfaces, and also have similar technical effects of the present invention.

[0010] The first aspect of the present invention aims to provide a carbon dot assembly.

[0011] The second aspect of the present invention is to provide a method for preparing a carbon dot assembly.

[0012] The third aspect of the present invention is to provide an application of a carbon dot assembly.

[0013] The fourth aspect of the present invention aims to provide a fungicide.

[0014] The fifth aspect of the present invention aims to provide a medicine.

[0015] In order to achieve the above-mentioned purpose of the present invention, the technical solution adopted by the present invention is:

[0016] In a first aspect of the present invention, a carbon dot assembly is provided, the carbon dot assembly comprising an electron donor core and an electron acceptor interface between and on the surface of the electron donor core; the electron donor core comprises carbon dots.

[0017] Preferably, the electron acceptor material is bonded to the carbon dots via chemical bonds.

[0018] Preferably, the chemical bond includes at least one of a covalent bond, an ionic bond, and a metallic bond.

[0019] In the first aspect of the present invention, the so-called carbon dot assembly refers to an electron acceptor material that is combined with multiple carbon dots through covalent bonds or non-covalent bonds to form an aggregate of carbon dots. In the carbon dot assembly, the electron acceptor material wraps multiple carbon dots, and the electron acceptor material's adsorbing sub-groups form an acceptor interface that can accept electrons with different carbon dot surfaces. In the carbon dot assembly, the carbon dots and the electron acceptor material are tightly combined, not simply physically mixed or dispersed.

[0020] Preferably, the electron acceptor material comprises a material containing an electron withdrawing group.

[0021] Preferably, the electron-withdrawing group includes one or more of a nitro group, a cyano group, a halogen group, an alkynyl group, a carboxyl group, a sulfonic acid group, a formyl group, and an acyl group.

[0022] Preferably, the electron withdrawing group comprises a cyano group.

[0023] In one embodiment of the present invention, the carbon dot assembly is formed by combining carboxyl groups on the aggregated carbon dots and hydroxyl groups on the material containing electron-withdrawing groups through ester bonds.

[0024] In one embodiment of the present invention, the electron acceptor material includes 2,3-dicyanohydroquinone.

[0025] Preferably, the particle size of the carbon dots is 1 to 10 nm.

[0026] Preferably, the particle size of the carbon dots is 1.5 to 3.5 nm.

[0027] The carbon dots in the present invention are those having a large number of sp 2 Conjugated domains and carbon dots with electron donor properties.

[0028] In one embodiment of the present invention, the carbon dots are prepared by solvothermal reaction of citric acid and urea.

[0029] Preferably, the mass ratio of the citric acid to the urea is 1:(1-5); more preferably, it is 1:(2-4).

[0030] Preferably, the solvent is at least one of N,N-dimethylformamide, dimethyl sulfoxide, formic acid, acetic acid, acetone and N-methylpyrrolidone; more preferably, the solvent is N,N-dimethylformamide.

[0031] Preferably, the reaction temperature is 120-200° C., and the reaction time is 0.5-24 h.

[0032] Preferably, the particle size of the carbon dot assembly is 15 to 30 nm; more preferably, it is 15 to 20 nm.

[0033] The second aspect of the present invention provides a method for preparing the carbon dot assembly of the first aspect of the present invention, comprising the following steps:

[0034] The carbon dots, the electron acceptor material and the solution are mixed and reacted to obtain.

[0035] Preferably, the reaction comprises at least one of a solvothermal reaction, a microwave reaction, a deposition reaction, and a calcination reaction.

[0036] Preferably, the reaction is a solvothermal reaction.

[0037] Preferably, the temperature of the solvent thermal reaction is 40 to 200° C.; the time of the solvent thermal reaction is 0.5 to 24 hours.

[0038] Preferably, the solvent includes at least one of N,N-dimethylformamide, dimethyl sulfoxide, formic acid, acetic acid, acetone and N-methylpyrrolidone; a mixture of two or more thereof may also be selected, and the mixing ratio is not limited.

[0039] In one embodiment of the present invention, the carbon dot assembly is prepared from a carbon dot solution and an electron acceptor material under solvothermal conditions.

[0040] Preferably, the carbon dots are prepared using citric acid and urea as raw materials, and the mass ratio of the citric acid to the urea is 1:(1-5); more preferably, it is 1:(2-4).

[0041] Preferably, the mass ratio of the carbon dots to the electron acceptor material is 1:(0.1-3).

[0042] Preferably, the reaction temperature is 80-200° C., and the reaction time is 0.5-24 h.

[0043] In some embodiments of the present invention, the reaction temperature of the solvothermal reaction is 80-200°C, for example, 80°C, 90°C, 100°C, 110°C, 120°C, 130°C, 140°C, 145°C, 150°C, 155°C, 160°C, 165°C, 170°C, 175°C, 180°C, 185°C, 190°C, 195°C or 200°C, etc.; the reaction time is 0.5-24h, for example, 0.5h, 1h, 2h, 3h, 4h, 5h, 6h, 7h, 8h, 9h, 10h, 11h, 12h, 13h, 14h, 15h, 16h, 17h, 18h, 19h, 20h, 21h, 22h, 23h or 24h, etc.

[0044] Furthermore, in some embodiments, the product obtained from the solvothermal reaction is purified by centrifugation, the speed of the centrifugation is 8000 rpm, each centrifugation lasts 10 min, and the centrifugation is performed three times in total.

[0045] The third aspect of the present invention provides the use of the carbon dot assembly of the first aspect of the present invention in 1) to 3):

[0046] 1) Producing active oxygen;

[0047] 2) preparing a fungicide;

[0048] 3) Preparation of anti-tumor drugs.

[0049] Preferably, the bactericide is a photocatalytic bactericide.

[0050] Preferably, the anti-tumor drug is a photocatalytic drug.

[0051] Preferably, the tumor comprises at least one of a solid tumor and a hematological tumor; and further comprises a solid tumor.

[0052] Preferably, the solid tumor comprises liver cancer, colorectal cancer, bladder cancer, breast cancer, cervical cancer, prostate cancer, glioma, melanoma, pancreatic cancer, nasopharyngeal cancer, lung cancer, gastric cancer, adrenocortical carcinoma, pararenal cortical carcinoma, anal cancer, appendix cancer, astrocytoma, atypical teratoma, rhabdoid tumor, basal cell carcinoma, bile duct cancer, bladder cancer, bone cancer, brain tumor, bronchial tumor, Burkitt's lymphoma, carcinoid tumor, heart tumor, bile duct epithelial carcinoma, chordoma, colorectal cancer, craniopharyngioma, ductal carcinoma in situ, embryonic tumor, endometrial cancer, ependymoma, esophageal cancer, olfactory neuroblastoma, intracranial germ cell tumor, extragonadal germ cell tumor, eye cancer, fallopian tube cancer, gallbladder cancer, head and neck cancer, hypopharyngeal cancer, Kaposi's sarcoma, renal cancer, Langerhans cell tissue cell At least one of: leukocyte proliferation, laryngeal cancer, lip cancer, oral cancer, Merkel cell carcinoma, malignant mesothelioma, multiple endocrine neoplasia syndrome, mycosis fungoides, sinonasal cancer, neuroblastoma, non-small cell lung cancer, ovarian cancer, pancreatic neuroendocrine tumors, islet cell tumors, papillomatosis, paraganglioma, sinonasal cancer, parathyroid cancer, penile cancer, pharyngeal cancer, pituitary tumor, pleuropulmonary blastoma, primary peritoneal cancer, retinoblastoma, salivary gland tumor, sarcoma, Sezary syndrome, skin cancer, small cell lung cancer, small intestine cancer, soft tissue sarcoma, squamous cell carcinoma, testicular cancer, thymoma and thymic cancer, thyroid cancer, urethral cancer, uterine cancer, endometrial and uterine sarcoma, vaginal cancer, vascular tumor, vulvar cancer and single myeloma.

[0053] Preferably, the blood tumor is selected from at least one of B-cell acute lymphoid leukemia (BALL), T-cell acute lymphoid leukemia (TALL), acute lymphoid leukemia (ALL), chronic myeloid leukemia (CML), chronic lymphocytic leukemia (CLL), B-cell prolymphocytic leukemia, blastic plasmacytoid dendritic cell tumor, Burkitt's lymphoma, diffuse large B-cell lymphoma, follicular lymphoma, hairy cell leukemia, small cell or large cell-follicular lymphoma, MALT lymphoma, mantle cell lymphoma, marginal zone lymphoma, multiple myeloma, non-Hodgkin's lymphoma, plasmablastic lymphoma, plasmacytoid dendritic cell tumor, Waldenstrom macroglobulinemia and preleukemia.

[0054] In one embodiment of the present invention, the tumor is breast cancer.

[0055] Those skilled in the art can anticipate that, since the oxidative stress mechanism is a common mechanism for treating cancer, reactive oxygen species can exert the effect of tumor treatment by promoting tumor apoptosis, directly causing tumor cell necrosis, promoting autophagic cell death in tumor cells, and so on. Therefore, in the absence of prior art disclosures of tumor types that are specifically resistant to reactive oxygen species, the specific type of tumor will not affect the application of the present invention.

[0056] The fourth aspect of the present invention provides a fungicide comprising the carbon dot assembly of the first aspect of the present invention.

[0057] Preferably, the bactericide comprises other bactericidal components.

[0058] Preferably, the other bactericidal components include but are not limited to at least one of antibiotics and antimicrobial peptides.

[0059] Preferably, the bactericide further comprises a pharmaceutically acceptable carrier or excipient.

[0060] The fifth aspect of the present invention provides an anti-tumor drug, comprising the carbon dot assembly of the first aspect of the present invention.

[0061] Preferably, the drug also includes other anti-tumor drugs.

[0062] Preferably, the drug further comprises a pharmaceutically acceptable excipient.

[0063] Preferably, the pharmaceutically acceptable excipients include but are not limited to at least one of a desiccant, an antioxidant, a stabilizer, a binder, a dispersant, a filler, a buffer, and a coating material.

[0064] Preferably, the dosage form of the drug includes at least one of capsules, tablets, microcapsules, injections, suppositories, sprays, powders, soft capsules, pellets, honey pills, pills, granules, honey pastes, sustained-release preparations, oral liquid preparations, chewable tablets, buccal tablets, transdermal patches, and effervescent tablets.

[0065] The beneficial effects of the present invention are:

[0066] The present invention provides a carbon dot assembly, which is composed of a core formed by aggregated carbon dots with electron-donating properties and an interface formed by molecules containing electron-withdrawing groups. Compared with traditional carbon dots, the carbon dot assembly has a larger particle size, which is more conducive to its targeting to tumor tissues through the enhanced permeability and retention effect of tumor tissue characteristics, thereby achieving efficient tumor killing. The carbon dot assembly provided by the present invention has excellent properties, and can oxidize important biological substrates through photogenerated holes while specifically generating type I reactive oxygen species, thereby achieving efficient anti-tumor photodynamic therapy. Compared with traditional small molecule photosensitizers, the carbon dot assembly has better water solubility and is more conducive to its application in organisms. The carbon dot assembly provided by the present invention generates reactive oxygen species through a photodynamic process to kill pathogenic microorganisms, and its killing effect will not be affected by the current drug resistance state of the microorganisms, and will not cause the microorganisms to develop drug resistance. The donor-acceptor type carbon dot assembly provided by the present invention is simple to prepare, and is directly prepared by reacting carbon dots with electron-donating properties and electron acceptor materials containing electron-withdrawing groups under solvent thermal conditions, without the need for complex reaction processes and operations. BRIEF DESCRIPTION OF THE DRAWINGS

[0067] In order to more clearly illustrate the technical solutions of the embodiments of the present invention, the drawings required for use in the embodiments are briefly introduced below. It should be understood that the following drawings only show certain embodiments of the present invention and therefore should not be regarded as limiting the scope. For ordinary technicians in this field, other related drawings can be obtained based on these drawings without creative work.

[0068] Figure 1 A schematic diagram of the principle of the preparation process of a donor-acceptor type carbon dot assembly provided in one embodiment of the present invention.

[0069] Figure 2 TEM images (a, b) of supra-CDs of Example 1 of the present invention, TEM image (c) of CDs of Comparative Example 1 and their particle size distribution graph (d).

[0070] Figure 3 The ultraviolet-visible (UV-Vis) absorption spectra of the supra-CDs of Example 1 of the present invention, the CDs of Comparative Example 1, the electron-withdrawing group-containing molecule DCHQ, and a simple mixture of CDs and DCHQ.

[0071] Figure 4 Fluorescence spectra of the active oxygen probe DCFH solution (a), the DCFH solution containing CDs of Comparative Example 1 (b), and the DCFH solution containing supra-CDs of Example 1 (c) after different illumination times, and their fluorescence intensity quantification graph at 525 nm (d).

[0072] Figure 5The fluorescence spectra of the superoxide radical probe DHR123 solution (a), the DHR123 solution containing CDs of Comparative Example 1 (b), and the DHR123 solution containing supra-CDs of Example 1 (c) after different illumination times, and their fluorescence intensity quantification graph at 528 nm (d).

[0073] Figure 6 Electron paramagnetic resonance spectra of the DMSO solution containing CDs of comparative example 1 and free radical scavenger DMPO (a) and the DMSO solution containing supra-CDs of example 1 and free radical scavenger DMPO (b) after different illumination times, and their signal intensity quantification graph at 335.6 mT (c).

[0074] Figure 7 The electron paramagnetic resonance spectra of the DMSO solution of CDs of Comparative Example 1 (a) and the DMSO solution of supra-CDs of Example 1 (b) after different illumination times, and a quantitative graph of their signal intensities at 337.5 mT (c); the electron paramagnetic resonance spectra of the DMSO solution of supra-CDs of Example 1 at different times after illumination for 5 min and then stopping the illumination (d) and a quantitative graph of their signal intensities at 337.5 mT (e).

[0075] Figure 8 The UV-Vis absorption spectra of BH4 solution (a) and NADH solution (b) added with supra-CDs of Example 1 after different illumination times.

[0076] Fig. 9 The figures are cytotoxicity tests of different concentrations of CDs of Comparative Example 1 (a) and supra-CDs of Example 1 (b) on mouse breast cancer cells (4T1) under dark and light conditions, and the ability of CDs of Comparative Example 1 and supra-CDs of Example 1 to generate reactive oxygen species (c) and superoxide radicals (d) in 4T1 cells.

[0077] Fig.10 The growth curves of mouse tumors in different treatment groups (a), the tumor weight of mouse tumors 16 days after treatment (b), the photos of mouse tumors 16 days after treatment (c) and the typical comparison pictures of mouse tumors before and after treatment (d).

[0078] Fig.11 The antibacterial rate test of supra-CDs of Example 1 at different concentrations against Staphylococcus aureus under light conditions. DETAILED DESCRIPTION

[0079] The following will be combined with the embodiments to clearly and completely describe the concept of the present invention and the technical effects produced, so as to fully understand the purpose, characteristics and effects of the present invention. Obviously, the described embodiments are only part of the embodiments of the present invention, not all of them. Based on the embodiments of the present invention, other embodiments obtained by those skilled in the art without creative work are all within the scope of protection of the present invention.

[0080] If the specific conditions are not specified in the examples, the experiments were carried out under conventional conditions or conditions recommended by the manufacturer. If the manufacturers of the reagents or instruments are not specified, they are all conventional products that can be purchased commercially.

[0081] Example 1 A method for preparing a donor-acceptor type carbon dot assembly

[0082] This embodiment provides a method for preparing a donor-acceptor type carbon dot assembly, comprising the following steps:

[0083] Disperse 1g of citric acid and 2g of urea in 10mL of N,N-dimethylformamide, sonicate for 30min to assist dissolution, place the mixed solution in a high-pressure reactor equipped with a polytetrafluoroethylene liner, and then place it in a high-temperature oven for solvothermal reaction at a temperature of 180°C for 4h. After the reaction is cooled to room temperature, add 1g of 2,3-dicyanohydroquinone (DCHQ) to the above solution, place it in a high-temperature oven to continue the solvothermal reaction at a temperature of 180°C for 1h. After the reaction is cooled to room temperature, take out the reaction solution, add twice the volume of anhydrous ethanol to the reaction solution, centrifuge at 8000 rpm for 10min, discard the supernatant and collect the precipitate. Continue to add anhydrous ethanol and repeat washing for 3 times, then collect the precipitate. Freeze-drying to obtain a black solid is the donor-acceptor carbon dot assembly (supra-CDs).

[0084] Example 2

[0085] This embodiment provides a method for preparing a donor-acceptor type carbon dot assembly, which is different from Embodiment 1 only in that the second solvothermal reaction time is 0.5 h.

[0086] Example 3

[0087] This embodiment provides a method for preparing a donor-acceptor type carbon dot assembly, which is different from Embodiment 1 only in that the second solvothermal reaction time is 2 h.

[0088] Example 4

[0089] This embodiment provides a method for preparing a donor-acceptor type carbon dot assembly, which is different from Embodiment 1 only in that the second solvothermal reaction temperature is 160°C.

[0090] Example 5

[0091] This embodiment provides a method for preparing a donor-acceptor type carbon dot assembly, which is different from Embodiment 1 only in that the second solvothermal reaction temperature is 140°C.

[0092] Example 6

[0093] This embodiment provides a method for preparing a donor-acceptor type carbon dot assembly, which is different from Embodiment 1 only in that the second solvothermal reaction temperature is 120°C.

[0094] Example 7

[0095] This embodiment provides a method for preparing a donor-acceptor type carbon dot assembly, which is different from Embodiment 1 only in that the second solvothermal reaction temperature is 100°C.

[0096] Example 8

[0097] This embodiment provides a method for preparing a donor-acceptor type carbon dot assembly, which is different from Embodiment 1 only in that the second solvothermal reaction temperature is 80°C.

[0098] Example 9

[0099] This embodiment provides a method for preparing a donor-acceptor type carbon dot assembly, which is different from Example 1 only in that the mass of the added DCHQ is 500 mg.

[0100] Example 10

[0101] This embodiment provides a method for preparing a donor-acceptor carbon dot assembly, which is different from Example 1 only in that the mass of the added DCHQ is 200 mg.

[0102] Comparative Example 1

[0103] This comparative example provides a method for preparing carbon dots, comprising the following steps:

[0104] Disperse 1g of citric acid and 2g of urea in 10mL of N,N-dimethylformamide, sonicate for 30min to assist dissolution, place the mixed solution in a high-pressure reactor equipped with a polytetrafluoroethylene liner, and then place it in a high-temperature oven for solvent thermal reaction at 180°C for 4h. After the reaction is cooled to room temperature, take out the reaction solution, add twice the volume of anhydrous ethanol to the reaction solution, centrifuge at 8000 rpm for 10min, discard the supernatant and collect the precipitate. Continue to add anhydrous ethanol and repeat washing for 3 times, then collect the precipitate. Freeze-dry to obtain a black solid, which is carbon dots (CDs).

[0105] Effect Example 1

[0106] The morphologies of supra-CDs obtained in Example 1 and CDs obtained in Comparative Example 1 were tested by transmission electron microscopy (TEM) and dynamic light scattering (DLS). The results are as follows: Figure 2 shown. Figure 2 TEM images and DLS images of supra-CDs in Example 1 of the present invention and CDs in Comparative Example 1.

[0107] from Figure 2 It can be seen that CDs and supra-CDs are evenly distributed in aqueous solution. High-resolution transmission electron microscopy (HRTEM) images show that CDs have 0.21nm lattice fringes, which are attributed to the (100) crystal plane of graphene. Supra-CDs are composed of multiple aggregated CDs and also have 0.21nm lattice fringes. DLS results show that the particle size of CDs is about 2.5nm, and the particle size of supra-CDs is about 20nm.

[0108] Effect Example 2

[0109] Ultraviolet absorption tests were performed on the supra-CDs obtained in Example 1, the CDs obtained in Comparative Example 1, the raw material DCHQ, and the mixture of CDs and DCHQ.

[0110] Figure 3 The results in show that CDs have two characteristic absorption peaks at 416nm and 550nm, the characteristic absorption peak of DCHQ is at 348nm, the mixture of CDs and DCHQ has the characteristic absorption peaks of both CDs and DCHQ, and supra-CDs shows a new characteristic absorption peak at 389nm, which proves the successful synthesis of supra-CDs and that supra-CDs is not a simple physical mixture of CDs and DCHQ.

[0111] Effect Example 3

[0112] The supra-CDs obtained in Example 1 and the CDs obtained in Comparative Example 1 were tested for their ability to produce active oxygen. 2,7-Dichlorofluorescein (DCFH) is an active oxygen probe that has no fluorescence itself. It emits green fluorescence after being oxidized by active oxygen. Therefore, the ability to produce active oxygen can be judged by detecting its fluorescence signal. The DCFH used was purchased from Shanghai Aladdin Biochemical Technology Co., Ltd. with the item number H131224. The fluorescence spectra of DCFH (4 μM), DCFH (4 μM) + CDs (5 ppm), and DCFH (4 μM) + supra-CDs (5 ppm) in the range of 490 nm to 650 nm were detected by fluorescence spectrometer after white light irradiation for different times, and the fluorescence intensity at 525 nm was selected for quantitative analysis.

[0113] Figure 4 The results show that the fluorescence of the DCFH solution itself does not increase significantly under illumination; however, in the presence of CDs, the fluorescence of the DCFH solution gradually increases after illumination, indicating that CDs can produce a small amount of reactive oxygen species; compared with CDs, in the presence of supra-CD, the fluorescence enhancement of the DCFH solution after illumination is more obvious, indicating that supra-CDs can produce more reactive oxygen species than CDs.

[0114] Effect Example 4

[0115] The superoxide free radical generating ability of supra-CDs obtained in Example 1 and CDs obtained in Comparative Example 1 was tested. Dihydrorhodamine 123 (DHR23) is a superoxide free radical probe, which has no fluorescence itself, but emits green fluorescence after being oxidized by superoxide free radicals. Therefore, the superoxide free radical generating ability can be judged by detecting its fluorescence signal. The DHR123 used was purchased from Shanghai Maokang Biotechnology Co., Ltd., and the item number is MX4455. The fluorescence spectra of DHR123 (4 μM), DHR23 (4 μM) + CDs (5 ppm), and DHR123 (4 μM) + supra-CDs (5 ppm) in the range of 510 nm to 650 nm after white light irradiation for different times were detected by fluorescence spectrometer, and the fluorescence intensity at 528 nm was selected for quantitative analysis.

[0116] Figure 5 The results show that the fluorescence of DHR123 solution itself does not increase significantly under illumination; however, in the presence of CDs, the fluorescence of DHR123 solution gradually increases after illumination, indicating that CDs can produce a small amount of superoxide radicals; compared with CDs, the fluorescence of DHR123 solution in the presence of supra-CD increases more significantly after illumination, indicating that supra-CDs can produce more superoxide radicals than CDs.

[0117] Effect Example 5

[0118] The superoxide free radical generating ability of supra-CDs obtained in Example 1 and CDs obtained in Comparative Example 1 was tested by electron paramagnetic resonance (EPR) technology. 5,5-Dimethyl-1-pyrroline-N-oxide (DMPO) is a free radical scavenger that can capture superoxide free radicals in a dimethyl sulfoxide (DMSO) solution and show four characteristic signals with an intensity ratio of 1:1:1:1. The DMPO used was purchased from Sigma-Aldrich (Shanghai) Trading Co., Ltd. with the item number 222-011-1. The EPR spectra of DMPO (1%) + CDs (100 ppm) and DMPO (1%) + supra-CDs (100 ppm) after white light irradiation for different times were detected by electron paramagnetic resonance (EPR) spectroscopy.

[0119] Figure 6 The results showed that in the presence of CDs, the DMPO solution showed four characteristic signals of 1:1:1:1 after illumination, indicating that CDs can produce a small amount of superoxide free radicals; compared with CDs, in the presence of supra-CD, the DMPO solution also showed four characteristic signals of 1:1:1:1 after illumination, but the signal intensity was significantly stronger, indicating that supra-CDs can produce more superoxide free radicals than CDs.

[0120] Effect Example 6

[0121] The supra-CDs obtained in Example 1 and the CDs obtained in Comparative Example 1 were tested for their photogenerated hole generation capabilities using electron paramagnetic resonance (EPR) technology.

[0122] Figure 7 The results show that the DMSO solution of CDs has almost no EPR signal under illumination, while the DMSO solution of supra-CDs shows a gradually enhanced EPR signal under illumination, indicating that supra-CDs can generate photogenerated holes under illumination. After 5 minutes of illumination, the EPR signal of the DMSO solution of supra-CD was monitored at different intervals. The results showed that after the illumination was stopped, the EPR signal of the DMSO solution of supra-CD could still exist. After 40 minutes, the EPR signal still remained above 80% of the highest value. These results show that photogenerated holes can exist for a long time, proving that there is effective long-term charge separation in supra-CDs.

[0123] Effect Example 7

[0124] Taking tetrahydrobiopterin (BH4) and reduced coenzyme I (NADH) as representatives, the oxidation ability of the photogenerated holes generated by supra-CDs obtained in Example 1 was tested. The BH4 used was purchased from MCE (MedChemExpress) with the item number HY-107383; the NADH used was purchased from Beijing Solebow Technology Co., Ltd. with the item number IN0020. The visible-ultraviolet absorption spectra of supra-CDs (50ppm) + BH4 (100μM) and supra-CDs (50ppm) + NADH (100μM) after white light irradiation for different times were detected by UV-visible spectrometer.

[0125] Figure 8 The results in show that with the extension of illumination time, the absorption peak of BH4 at 300nm decreases, while the absorption peak of its oxidation product dihydrobiopterin (BH2) at 280nm increases, indicating that the photogenerated holes generated by supra-CDs under illumination can oxidize BH4. The oxidation test results of NADH show that with the extension of illumination time, the absorption peak of NADH at 340nm decreases, while the absorption peak of its oxidation product at 260nm increases, indicating that the photogenerated holes generated by supra-CDs under illumination can oxidize NADH. These results prove that the photogenerated holes generated by supra-CDs under illumination can oxidize some important biological substrates, thereby affecting cell growth.

[0126] Effect Example 8

[0127] Mouse breast cancer cells (4T1) were used as tumor cell models to test the ability of supra-CDs obtained in Example 1 and CDs obtained in Comparative Example 1 to kill tumor cells and produce reactive oxygen species in cells. 4T1 cells were plated in a 96-well plate at a density of 8,000 cells per well, incubated overnight in a cell culture incubator, and then the culture medium in the well plate was replaced with 100 μL of cell culture medium containing different concentrations (500, 250, 100, 50, 25, 10, 5, 0 ppm) of CDs or supra-CDs. After 12 hours of culture, the illumination group was illuminated with a white light flashlight for 10 minutes, and after continuing to culture for 12 hours, CCK-8 reagent was added, and the OD value of each well at 450 nm was measured with an ELISA instrument, and the cell viability was calculated.

[0128] Fig. 9The results showed that under dark conditions, both CDs and supra-CDs had only weak cytotoxicity; under light conditions, CDs and supra-CDs showed cell killing ability that increased with concentration. At a concentration of 500ppm, the cell survival rate after CDs treatment was about 36%, while the cell survival rate after supra-CDs treatment was only 4%, indicating that supra-CDs has a stronger tumor cell killing ability under light conditions. Subsequently, the ability of supra-CDs and CDs to produce reactive oxygen species in tumor cells was measured. The groups for reactive oxygen species determination were: supra-CDs, L (white light irradiation for 10min), CDs+L, supra-CDs+L. 4T1 cells were cultured at 3×10 4 The cells were plated in an 8-well chamber at a density of 100 cells. After incubation in a cell culture incubator overnight, the medium in the chamber was replaced with 200 μL of cell culture medium containing 500 ppm of CDs or supra-CDs, and the culture was continued for 12 hours. After replacing the medium in the chamber with the medium containing DCFH or DHR123, the illumination group was illuminated with a white light flashlight for 10 minutes, and after incubation in the cell culture medium for 30 minutes, the green fluorescence of each group was observed using a confocal microscope. The results showed that under illumination conditions, CDs could only produce a small amount of reactive oxygen in tumor cells, while supra-CDs could produce a large amount of reactive oxygen in tumor cells, and the reactive oxygen produced was mainly superoxide free radicals.

[0129] Effect Example 9

[0130] BALB / c mice loaded with breast cancer cells (4T1) were used as animal models to test the inhibitory ability of supra-CDs obtained in Example 1 and CDs obtained in Comparative Example 1 on tumor growth at the animal level. 6PBS with 4T1 cells was added to the mice. One week later, the mice were randomly divided into 5 groups: PBS, L (white light irradiation for 10 min), supra-CDs, CDs+L and supra-CDs+L, with 5 mice in each group. Each group was injected with 100 μL of PBS, CDs (10 mg / kg) or supra-CDs (10 mg / kg) through the tail vein. 12 hours later, the mice in the L, CDs+L and supra-CDs groups were irradiated with white light for 10 min. The tumor volume of the mice was measured in the following two days. On the 8th day of treatment, 100 μL of PBS, CDs (10 mg / kg) or supra-CDs (10 mg / kg) was injected into the tail vein again. 12 hours later, the mice in the L, CDs+L and supra-CDs groups were irradiated with white light for 10 min, and the tumor volume of the mice was continuously monitored. On the 16th day of treatment, the mice were killed and the tumors of the mice were dissected. The weight of the tumors of the mice was weighed and the tumors of the mice were photographed. The animal experiments involved in this example have been approved by the Ethics Committee of the Experimental Animal Research Center of Southern University of Science and Technology, with approval number SUSTC-JY2019092.

[0131] Fig.10 The results showed that light alone or supra-CDs without light had no inhibitory effect on tumor growth. CDs could slow down the growth of tumors under light conditions, while supra-CDs could completely inhibit tumor growth under light conditions.

[0132] In summary, the present invention mainly provides a donor-acceptor carbon dot assembly and its preparation method and application. The donor-acceptor carbon dot assembly supra-CDs is obtained by eutectic melting CDs and a molecule DCHQ containing an electron-withdrawing group. The supra-CDs are composed of a core formed by the aggregation of CDs and an interface formed by DCHQ. The introduction of the electron-withdrawing molecule DCHQ promotes electron transfer in supra-CDs. Under light conditions, excited electrons are transferred from the core of supra-CD to the interface, and further react with oxygen in the surrounding environment to generate superoxide radicals; while the photogenerated holes with long life are stabilized in the core of supra-CD, and these photogenerated holes can further oxidize some important biological substrates. Under the combined action of ultrasonic free radicals and photogenerated holes, supra-CD can effectively kill tumors under light. Using mouse breast cancer cells (4T1) as a tumor cell model and BALB / c mice with subcutaneous 4T1 as an animal model, the experimental results prove the excellent anti-tumor therapeutic effect of supra-CDs.

[0133] Effect Example 10

[0134] This example tests the antibacterial rate of supra-CDs of different concentrations against Staphylococcus aureus under illumination conditions. The specific operation is as follows: Staphylococcus aureus is cultured in a 37°C incubator using fresh Luria-Bertani (LB) medium until it reaches the logarithmic growth phase. Subsequently, it is washed three times with sterile PBS, centrifuged at 10,000 rpm for 2 minutes, the absorbance value at 600 nm is measured and adjusted to 1, and then 400 μL of bacterial solution containing different concentrations of supra-CDs is added to a 48-well plate and irradiated with white light for 10 minutes. Supra-CDs with a concentration of 0 is used as a control group and continued to be cultured in a 37°C incubator for 120 minutes. Subsequently, 100 μL of bacterial solution is spread on an LB agar plate, cultured overnight at 37°C, and the number of colonies is observed and counted.

[0135] The calculation process of the antibacterial rate is as follows: Antibacterial rate = (number of colonies in the control group - number of colonies in the experimental group) / number of colonies in the control group × 100%.

[0136] The results are as follows Fig.11 As shown in the figure, as the concentration of supra-CDs increases, the antibacterial rate after white light irradiation gradually increases, showing a dose-dependent effect. When supra-CDs reaches 500ppm, its antibacterial rate reaches 90%.

Claims

1. A carbon dot assembly, characterized in that: The carbon dot assembly includes an electron donor core and an electron acceptor interface between and on the surface of the electron donor core; The electron donor core includes carbon dots.

2. The carbon dot assembly according to claim 1, characterized in that: The electron acceptor interface is connected to the electron donor core via chemical bonds.

3. The carbon dot assembly according to claim 2, characterized in that: The electron acceptor is melted between the electron donor core and the surface to form an interface.

4. The carbon dot assembly according to any one of claims 1 to 3, characterized in that: The electron acceptor includes a material containing an electron-withdrawing group; Preferably, the electron-withdrawing group includes one or more of a nitro group, a cyano group, a halogen group, an alkynyl group, a carboxyl group, a sulfonic acid group, a formyl group, and an acyl group.

5. The method for preparing the carbon dot assembly according to claim 4, characterized in that: reacting the carbon dots with the electron acceptor material to obtain the carbon dot assembly; The electron acceptor material includes a material containing an electron withdrawing group.

6. The preparation method according to claim 5, characterized in that: The reaction comprises at least one of a solvent thermal reaction, a microwave reaction, a deposition reaction, and a calcination reaction; Preferably, the reaction is a solvothermal reaction; Preferably, the temperature of the solvent thermal reaction is 40 to 200° C.; the time of the solvent thermal reaction is 0.5 to 24 hours.

7. The preparation method according to claim 6, characterized in that: The mass ratio of the carbon dots to the electron acceptor material is 1:(0.1-3).

8. Use of the carbon dot assembly according to claim 4 in 1) to 3): 1) Producing active oxygen; 2) preparing a fungicide; 3) Preparation of anti-tumor drugs; Preferably, the bactericide is a photocatalytic bactericide; Preferably, the anti-tumor drug is a photocatalytic drug.

9. A bactericide comprising the carbon dot assembly according to claim 4; Preferably, the bactericide further comprises other bactericidal components; Preferably, the bactericide further comprises a pharmaceutically acceptable carrier or excipient.

10. An anti-tumor drug comprising the carbon dot assembly according to claim 4; Preferably, the drug also includes other anti-tumor drugs; Preferably, the drug further comprises a pharmaceutically acceptable excipient; Preferably, the tumor comprises breast cancer.

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