Near-infrared fluorescent silicon dots, preparation method thereof and application thereof in antibacterial and tumor treatment

Abstract

This invention discloses a near-infrared fluorescent silicon dot, its preparation method, and its application in antibacterial and tumor treatment. The silicon dot is generated via a one-step hydrothermal method through a high-temperature condensation reaction between the carboxyl group of a 4-carboxyporphyrin dye and the amino group of a silane coupling agent. The silicon dot synthesized in this invention exhibits excellent water dispersibility, high near-infrared fluorescence intensity and pH stability, uniform morphology, good photostability, good biocompatibility, and high reactive oxygen species (ROS) yield. Furthermore, the preparation method is simple and convenient, uses widely available raw materials, and is low-cost, making it suitable for industrial production. Based on these excellent properties, it can be used to prepare fluorescent probes as fluorescent markers for cell and biological imaging. Its excellent singlet oxygen generation performance can be used for photodynamic therapy against bacteria and tumors. In addition, the silicon dot also possesses peroxidase properties, catalyzing the generation of hydroxyl radicals from hydrogen peroxide, thus enabling its use in chemokinetic therapy.

Description

Technical Field

[0001] This invention belongs to the field of chemical nanomaterials, specifically relating to a near-infrared fluorescent silicon dot, its preparation method, and its applications in fluorescence imaging, wound antibacterial treatment, tumor photodynamic therapy, and chemokinetic therapy. Background Technology

[0002] In recent years, the fields of nanomaterials and nanomaterial processing technology have developed rapidly, greatly promoting the development of nanobiotechnology. In particular, the application prospects of nanomaterials in bioimaging and biomedicine are extremely attractive. Among them, quantum dots stand out due to their excellent optical properties, including high fluorescence, single excitation, tunable emission spectrum, and resistance to photobleaching. They can be used as fluorescent probes to image or detect analytes in living organisms, providing reliable data for bioanalysis. Traditional quantum dots usually refer to cadmium-based semiconductor crystals, most of which contain heavy metals such as lead and cadmium. While causing harm to humans and the environment, this also limits the development of quantum dots in the biological and medical fields. Therefore, finding other novel quantum dot materials with excellent biocompatibility is particularly important.

[0003] Research has revealed that when silicon materials are shrunk to the nanoscale, their properties undergo a dramatic transformation, resulting in superior and novel properties in multiple aspects. Such silicon materials are called silicon quantum dots. They are zero-dimensional semiconductor fluorescent nanomaterials. Besides their excellent optical properties, silicon quantum dots possess many advantages that other quantum dots lack, such as excellent biocompatibility, stable resistance to photofluorescence bleaching, simple synthesis processes, and tunable emission spectra. In particular, water-soluble organosilicon quantum dots have shown enormous development potential in electrochemical performance, photochemical performance, and biocompatibility.

[0004] In addition, the reported organosilicon quantum dots also have some drawbacks, such as short emission wavelength, high cytotoxicity, low quantum yield, weak tissue penetration, and cumbersome and complex synthesis steps. Overcoming these problems would lead to further development of silicon quantum dots in biological applications. Summary of the Invention

[0005] Purpose of the invention: To address the problems existing in the prior art, this invention prepares a near-infrared fluorescent silicon dot using a one-step hydrothermal method. This silicon dot has a high fluorescence quantum yield, good fluorescence stability, is non-toxic, has low preparation cost, and is simple to prepare. Furthermore, this silicon quantum dot can be used for fluorescence imaging of organisms, antibacterial treatment of wounds, photodynamic therapy (PDT) of tumors, and chemokinetic therapy (CDT).

[0006] The present invention also provides a method for preparing the near-infrared fluorescent silicon dots and their applications.

[0007] Technical solution: In order to achieve the above objectives, the present invention provides a near-infrared fluorescent silicon dot, which is generated by a one-step hydrothermal method through a high-temperature condensation reaction between the carboxyl group of 4-carboxyphenylporphyrin (TCPP) and the amino group of a silane coupling agent.

[0008] The near-infrared fluorescent silicon dots have a diameter in the range of 1-10 nm and mainly contain four elements: C, N, O, and Si.

[0009] The near-infrared fluorescent silicon dots exhibit fluorescence properties, with a strong fluorescence emission peak in the near-infrared band of 650 nm-700 nm.

[0010] The silane coupling reagent includes any one or more of diethylenetriaminepropyltrimethoxysilane (AEEA), 3-aminopropyltriethoxysilane (AMEO), 3-aminopropyltrimethoxysilane (APTMS), and N-(beta-aminoethyl)-gama-aminopropyltrimethoxysilane (DAMO).

[0011] The method for preparing near-infrared fluorescent organosilicon quantum dots according to the present invention includes the following steps:

[0012] (1) The reaction is carried out by a one-step hydrothermal method using fluorescent dye TCPP, silane coupling reagent, and deionized water;

[0013] (2) The solid powder obtained by centrifugation and dialysis of the reaction solution in step (1) followed by freeze drying is near-infrared fluorescent organosilicon quantum dots.

[0014] In step (1), TCPP and deionized water are first mixed at a mass ratio of 1-3:400, and then silanizing reagent is added to the above mixture. The mixture is then sonicated to make it uniform. The molar ratio of TCPP to silane coupling reagent is 1:80-120.

[0015] The hydrothermal reaction in step (1) is carried out at a temperature of 140-200℃ for 3-8 hours.

[0016] Preferably, the method for preparing fluorescent organosilicon quantum dots according to the present invention includes the following steps:

[0017] The fluorescent dye TCPP solid was dissolved in deionized water, and the silane coupling reagent AEEA was added by pipette. After hydrothermal reaction, heating was stopped, the solution was cooled, the reaction solution was removed, unreacted reagents were removed by dialyzing, and the supernatant was retained by centrifugation.

[0018] The solid powder obtained by freeze-drying the supernatant in step (1) is a near-infrared fluorescent organosilicon quantum dot.

[0019] In step (1), TCPP is mixed with an appropriate amount of deionized water at a mass ratio of 3:400, and then AEEA is added; the preferred molar ratio of TCPP to AEEA is 1:100. The hydrothermal reaction in step (1) is carried out at a temperature of 160 ℃ for 4 hours.

[0020] The present invention describes the application of near-infrared fluorescent silicon dots in the preparation of antibacterial agents for biological wounds and in fluorescent imaging reagents.

[0021] The application of the near-infrared fluorescent silicon dots described in this invention in the preparation of photodynamic agents for tumor treatment.

[0022] The application of the near-infrared fluorescent silicon dots described in this invention in the preparation of chemokinetics for tumor treatment.

[0023] Furthermore, the near-infrared fluorescent silicon dots can be used as biofluorescent probes or as PDT / CDT combination therapy formulations for antibacterial and antitumor applications.

[0024] This invention utilizes a mixture of TCPP aqueous solution and silane coupling reagent to carry out a high-temperature reaction in a hydrothermal reactor. The synthesized fluorescent organosilicon quantum dots exhibit excellent biocompatibility, superior near-infrared luminescence properties, and outstanding photochemical stability, making them applicable to the biomedical field.

[0025] The near-infrared fluorescent silicon dot described in this invention is synthesized in one step via a hydrothermal method using an aqueous solution of the fluorescent probe TCPP and a silane coupling reagent. Four silane coupling reagents—divinyltriaminepropyltrimethoxysilane (AEEA), 3-aminopropyltriethoxysilane (AMEO), 3-aminopropyltrimethoxysilane (APTMS), and N-(beta-aminoethyl)-gama-aminopropyltrimethoxysilane (DAMO)—showed essentially the same experimental results. AEEA is used as a representative example; the near-infrared emitting fluorescent silicon dot is formed by a high-temperature condensation reaction between the carboxyl group of TCPP and the amino group of AEEA.

[0026] This invention prepares a spherical, zero-dimensional near-infrared fluorescent silicon dot, synthesized in one step via a hydrothermal method from an aqueous TCPP solution and the silane coupling reagent AEEA. Due to its low preparation cost, simple method, and advantages such as good photochemical stability, excellent biocompatibility, and high fluorescence quantum yield, it shows great potential in the biomedical field. The fluorescent silicon quantum dot material of this invention exhibits good singlet oxygen generation performance, making it suitable as a PDT (proton pump dehydrogenase) agent; furthermore, it possesses peroxidase activity, catalyzing the conversion of hydrogen peroxide into harmful hydroxyl radicals, making it suitable as a CDT (co-dioxanone iontophoresis) agent. Therefore, this near-infrared organosilicon quantum dot can be used as a PDT / CDT combined therapeutic agent for biological wound antibacterial and tumor diagnosis and treatment. In a mouse model, this near-infrared fluorescent silicon dot achieved good therapeutic effects through PDT / CDT combined therapy.

[0027] This invention is the first to propose a zero-dimensional near-infrared fluorescent silicon dot for wound antibacterial and tumor treatment applications. It utilizes simple raw materials, a straightforward preparation method, and is non-toxic, making it advantageous for biomedical applications. Compared to the two previous cumbersome and complex "top-down" synthesis methods, this one-step hydrothermal method can control the size of the silicon dots, resulting in uniform morphology and high purity, without requiring expensive equipment or stringent conditions. Furthermore, the raw materials are inexpensive, allowing for rapid mass production.

[0028] This invention utilizes specific raw materials and a specific preparation method to prepare near-infrared fluorescent silicon dots. The phototherapy porphyrin compounds possess the ability to generate high levels of reactive oxygen species and low dark toxicity, making them powerful photosensitizers. However, TCPP itself has poor water solubility and weak luminescence. After the raw material TCPP reacts with a silane reagent to form silicon dots, the increased water solubility due to the combination of carboxyl and amino groups results in a higher photodynamic therapy (PDT) effect. Simultaneously, it can catalyze the decomposition of hydrogen peroxide to generate hydroxyl radicals, which can be used for chemikinetic therapy. Furthermore, the nanoscale size improves biodistribution, and the prepared near-infrared fluorescent silicon dots exhibit photodynamic therapy (PDT) / chemikinetic synergistic therapeutic capabilities. In the field of silicon quantum dots, this material, with its simple and stable production process, along with its multiple effects such as fluorescence imaging and synergistic therapy, makes it unique. Meanwhile, quantum dots are generally formed due to their good fluorescence properties (mostly blue, green or yellow light), water dispersibility, and the ability to be controlled by the fluorescent optical modulation of silicon nanoparticles for fluorescent labeling. In addition to these, this invention synthesizes near-infrared luminescent silicon dots due to the properties of the raw materials themselves and the combination between the raw materials, and produces unique PDT / CDT synergistic therapeutic and antibacterial effects. It can be used for fluorescence imaging and tumor treatment at the same time.

[0029] The presence of the carboxyl group in this invention endows TCPP with hydrogen bonding and strong intramolecular reactivity. Furthermore, TCPP exhibits high thermal stability, and the carboxyl group can rapidly combine with the amino group of the silane coupling reagent via high-temperature condensation, resulting in more efficient production and increased yield. TCPP itself is also an important photosensitive material, possessing photophysical properties similar to porphyrin compounds. When combined with a silane reagent, it exhibits enhanced stability and higher water solubility, and upon photoexcitation, it produces strong near-infrared fluorescence and a stronger photosensitizing effect.

[0030] This invention utilizes TCPP's high thermal stability and strong intramolecular reactivity due to its carboxyl groups, resulting in a shorter reaction time with silane reagents compared to existing near-infrared silicon dot synthesis methods. The synthesis can be achieved in as little as 3 hours via a one-step hydrothermal method, making it more efficient. The prepared silicon dots exhibit high stability and good water solubility. Furthermore, while most existing silicon dots only possess visible light emission properties and are primarily used in biosensing and bioimaging, the near-infrared fluorescent silicon dots prepared in this invention not only have high singlet oxygen generation capabilities but can also catalyze the decomposition of hydrogen peroxide to generate hydroxyl radicals, enabling simultaneous photodynamic and chemodynamic therapy – a crucial function of this invention.

[0031] In this invention, the silane coupling agent itself does not have antibacterial or antitumor effects. TCPP itself can be used as a photosensitive material, generating singlet oxygen and exhibiting photodynamic therapy properties. However, organic dye molecules have poor water solubility and photostability. The specific preparation method of this invention, after generating silicon dots, enhances water solubility, photostability, and luminescence efficiency, increases singlet oxygen yield, and improves photodynamic therapy efficiency. Furthermore, it can catalyze the generation of hydroxyl radicals from hydrogen peroxide, thereby enhancing chemodynamic therapy efficiency. The combined photodynamic / chemodynamic therapy significantly enhances antibacterial and antitumor effects.

[0032] The silicon dots synthesized in this invention not only possess the synergistic therapeutic effect of PDT / CDT and can be used as anti-tumor nanomedicines, but also, compared with silicon dots that emit blue, yellow, and green light as general fluorescent imaging reagents, the silicon dots produced by combining TCPP with silane coupling reagents have near-infrared luminescence properties, making them more advantageous in the field of bioimaging. Near-infrared fluorescence can better penetrate biological tissues, achieving higher imaging resolution and depth. Furthermore, due to the passive targeting performance of nanoparticles to tumor sites, they are superior as bioimaging probes.

[0033] The key raw material of this invention is 4-carboxyphenylporphyrin (TCPP), and the key step is a one-step hydrothermal method at 160°C, which enables a high-temperature condensation reaction between the carboxyl group of TCPP and the amino group of the silane coupling agent. This high-temperature and high-pressure carboxyl-amino condensation reaction enhances the stability and water solubility of the product, and also makes it superior to the TCPP raw material itself in various aspects. A comparison of the reactive oxygen yield of the TCPP raw material itself and the silicon dots of the synthesized product in this invention is presented. Figure 9 The reactive oxygen species yield of the silicon dot synthesized in this invention is much greater than that generated by TCPP itself. A comparison of the fluorescence stability of the TCPP raw material and the synthesized silicon dot at different pH values ​​is presented. Figure 10 The silicon dots of this invention exhibit photostability that TCPP does not possess at pH values ​​in the range of 2-12.

[0034] The present invention relates to the silicon dot Lineweaver-Burk ( Figure 11Vmax and Km are obtained as 8.98125x10. -5 M / min and 3.30245x10 -3 M, stronger than known horseradish peroxidases, proves that Si NDs possess peroxidase properties, capable of efficiently catalyzing the conversion of H2O2 to •OH. Si NDs can serve as high-performance nanozymes for chemokinetic (CDT) therapy. A comparative experiment was conducted on the MTT cell viability before and after laser treatment of the TCPP raw material and the synthesized silicon dots. Figure 12 By comparing the survival rates at the same concentration, it was demonstrated that the silicon dots of this invention have a more efficient tumor cell killing effect than the raw material TCPP under the premise of low toxicity. A comparative experiment of cell viability and mortality staining before and after laser lasing was conducted on the TCPP raw material itself and the synthesized silicon dots. Figure 13 The PI / AM staining pattern also demonstrated that the silicon dots of this invention, at the same concentration, exhibit a more efficient PDT / CDT synergistic effect in killing tumor cells than the raw material TCPP. Intracellular ROS levels of both the TCPP raw material and the synthesized silicon dots were measured using a DCFH staining comparison experiment. Figure 14 This also demonstrates higher levels of reactive oxygen species (ROS) than the raw material TCPP. A comparative experiment was conducted on the bacterial viability and mortality staining of the TCPP raw material and the synthesized silicon dots before and after laser lasing. Figure 15 Comparison of bacterial survival rates in in vitro antibacterial test groups ( Figure 16 ), the antibacterial treatment effect of subcutaneous wounds in mice infected with Staphylococcus aureus (S-aureus) Figure 17 Both of these studies demonstrate that the silicon dot effect of this invention is significantly stronger than that of TCPP.

[0035] Beneficial effects: Compared with the prior art, the present invention has the following advantages:

[0036] 1. This invention directly utilizes TCPP aqueous solution and silane coupling reagent to prepare zero-dimensional near-infrared fluorescent organosilicon quantum dots via a specific one-step hydrothermal method. The quantum dots are formed by the reaction between the carboxyl group of TCPP and the amino group of AEEA. Since TCPP itself has a certain fluorescence intensity and the fluorescence of the synthesized quantum dots is significantly improved, the quantum dots exhibit high fluorescence intensity, high quantum yield, uniform morphology, and good fluorescence stability, which is beneficial for biomedical applications. Furthermore, the preparation method is simple and convenient, has good biocompatibility and safety, and is low in cost.

[0037] 2. The fluorescent organosilicon quantum dots prepared in this invention exhibit strong near-infrared fluorescence and good water solubility, with a strong fluorescence emission peak in the 650 nm-700 nm wavelength range, demonstrating excellent fluorescence spectral performance. Therefore, silicon quantum dots have relatively good cell and tissue penetration. Due to their unique optical properties and relatively good cell and tissue penetration, they can be effectively used for in vivo fluorescence imaging.

[0038] 3. In addition to excellent near-infrared optical properties and good biocompatibility, the fluorescent organosilicon quantum dots prepared by this invention also have excellent singlet oxygen generation performance and peroxidase performance. They can be used as photodynamic / chemodynamic synergistic therapeutic agents for antibacterial and antitumor applications, and have no toxic side effects on organisms themselves. Attached Figure Description

[0039] Figure 1 The images show the near-infrared fluorescent organosilicon quantum dots prepared according to this invention before and after synthesis, with the fluorescent organosilicon quantum dots before synthesis (left) and after synthesis (right).

[0040] Figure 2 The images show a comparison of the fluorescence of the near-infrared fluorescent organosilicon quantum dots prepared in this invention under sunlight and a 660 nm laser. From left to right, these are the fluorescence comparison images obtained under sunlight and a 660 nm laser, respectively.

[0041] Figure 3 The ultraviolet absorption spectra of the dye molecule TCPP and the fluorescent organosilicon quantum dots prepared in this invention;

[0042] Figure 4 Fluorescence absorption spectra of the dye molecule TCPP and the fluorescent organosilicon quantum dots prepared in this invention;

[0043] Figure 5 The XRD pattern of the fluorescent organosilicon quantum dots prepared in this invention;

[0044] Figure 6 TEM image of fluorescent organosilicon quantum dots prepared in this invention;

[0045] Figure 7 Infrared spectrum of fluorescent organosilicon quantum dots prepared in this invention;

[0046] Figure 8 The XPS spectrum and peak fitting diagram of the near-infrared fluorescent organosilicon quantum dots prepared in this invention are shown, wherein A is the XPS spectrum of the fluorescent organosilicon quantum dots, B is the XPS spectrum of the fluorescent organosilicon quantum dots, and C is the peak fitting diagram of the fluorescent organosilicon quantum dots. 1s Fitted graph, C represents fluorescent organosilicon quantum dots N 1s Fitted graph, D represents fluorescent organosilicon quantum dots O 1s Fitted graph, E represents fluorescent organosilicon quantum dots Si 2pFitted plot;

[0047] Figure 9 The image shows the photocatalytic performance of the near-infrared fluorescent organosilicon quantum dots prepared in this invention. In the image, A represents the UV absorption changes of DPHA and MB at different times; B represents the UV absorption changes of DPHA and TCPP at different times; C represents the UV absorption changes of DPHA and SiNDs at different times; and D represents the singlet oxygen yields of TCPP and fluorescent organosilicon quantum dots with methylene blue (MB) as a control, with relative yields of 21.51% and 49.87%, respectively.

[0048] Figure 10 In Figure A, the fluorescence spectrum of the dye molecule TCPP at different pH values ​​is shown. In Figure B, the fluorescence spectrum of the fluorescent organosilicon quantum dots prepared in this invention at different pH values ​​is shown. In Figure C, the fluorescence intensity of TCPP and Si NDs corresponding to A and B is shown as a function of pH.

[0049] Figure 11 The following is a chemical kinetics test diagram of the near-infrared fluorescent organosilicon quantum dot CDT prepared in this invention. In this diagram, A is the UV-vis absorption spectrum of TMB after adding different concentrations of H2O2 in the presence of Si NDs, B is the 652 nm time-path absorbance of PBS solution containing Si NDs and TMB at different H2O2 concentrations, C is the Michaelis-Menten kinetic diagram of Si NDs, and D is the Lineweaver-Burk diagram of Si NDs.

[0050] Figure 12 For the in vitro cytotoxicity characterization of MTT, A shows the relative survival rate of 4T1 cells before and after 660 nm laser irradiation with PBS; B shows the relative survival rate of 4T1 cells before and after 660 nm laser irradiation with TCPP; and C shows the relative survival rate of 4T1 cells before and after 660 nm laser irradiation with the fluorescent organosilicon quantum dots prepared in this invention. );

[0051] Figure 13 The images show the fluorescence of PBS, TCPP, and Si NDs in 4T1 cells before and after 660 nm laser irradiation, co-stained with calcein AM / PI. The scale bars represent 100 µm.

[0052] Figure 14 CLSM images of DCF staining of PBS, TCPP and Si NDs in 4T1 cells before and after 660 nm laser irradiation, scale bar 10µm.

[0053] Figure 15CLSM fluorescence imaging of TCPP and Si NDs in vitro antibacterial activity, with PBS as the control group, and fluorescence images of calcein AM / PI co-staining in Staphylococcus aureus before and after 660 nm laser irradiation. Live bacteria are marked with AM in green and dead bacteria are marked with PI in red. The scale bars represent 100 µm.

[0054] Figure 16 This invention demonstrates the in vitro antibacterial efficacy of the Si NDs prepared in this invention. In this image, A shows colony images of *Staphylococcus aureus* and *Escherichia coli* exposed to PBS, TCPP, Si NDs, PBS + Light, TCPP + Light, and Si NDs + Light; B shows the bacterial survival rate corresponding to A. );

[0055] Figure 17 This invention demonstrates the in vivo antibacterial efficacy of the fluorescent organosilicon quantum dots prepared in this invention in live experiments. A shows photographs and corresponding histological sections of *Staphylococcus aureus* abscesses treated with PBS, TCPP (100 μg / mL), Si NDs, TCPP + Laser, and Si NDs + Laser, with a scale bar of 50 µm. B shows TUNEL staining measurements of wound tissue sections from different groups of mice, with a scale bar of 50 µm. C shows histological observation of normal tissue after Si NDs treatment, with a scale bar of 50 µm.

[0056] Figure 18 Fluorescence imaging of tumors in live animals after tail vein injection of near-infrared fluorescent organosilicon quantum dots prepared in this invention at different time points.

[0057] Figure 19 In vitro fluorescence imaging of major organs and tumors in mice 120 hours after injection of the near-infrared fluorescent organosilicon quantum dots (SiNDs) prepared in this invention.

[0058] Figure 20 These are representative images of tumor-bearing mice in the control group and different treatment groups after treatment on day 12 in in vivo experiments.

[0059] Figure 21 This is a graph showing the changes in body weight of tumor-bearing mice during in vivo experimental treatment.

[0060] Figure 22 Tumor growth curves of tumor-bearing mice after administration of different formulations during in vivo experiments ( );

[0061] Figure 23Photographs of xenografted tumors dissected from mice on day 12 after treatment with different agents in an in vivo experiment. Scale bar: 2cm.

[0062] Figure 24 The tumor weight of xenografted tumors dissected from mice on day 12 after treatment with different formulations in in vivo experiments. ), G1: PBS, G2: TCPP, G3: Si NDs, G4: TCPP+Laser, G5: SiNDs +Laser;

[0063] Figure 25 TUNEL staining (a) and H&E staining (b) of tumor tissue sections from mice treated with different formulations in in vivo on day 12. From left to right, they are PBS group, TCPP group, Si NDs, TCPP+Laser and Si NDs+Laser group. The scale bar is 50 µm.

[0064] Figure 26 The images show tissue sections of near-infrared fluorescent organosilicon quantum dots (SiNDs) prepared in this invention, from left to right: heart, liver, spleen, lung, and kidney sections of a mouse. Detailed Implementation

[0065] The present invention will be further described below with reference to the accompanying drawings and embodiments.

[0066] Unless otherwise specified, all materials and reagents used in the following examples are commercially available. Experimental methods not specifically described in the examples are generally performed under standard conditions or as recommended by the manufacturer.

[0067] Among them, TCPP: 14609-54-2, SKU: P115336-100mg, manufacturer: Aladdin; AEEA: cas: 35141-30-1, SKU: R008710-25ml, manufacturer: RHAWN.

[0068] Example 1

[0069] (1) Weigh 30 mg TCPP using an electronic balance and dissolve it in 4 mL of deionized water. Add the silanizing coupling reagent AEEA to the above mixture so that the molar ratio of TCPP to AEEA is 1:100. Use an ultrasonic cleaner to mix it evenly.

[0070] (2) After mixing thoroughly, the above solution was transferred to a 15 ml polytetrafluoroethylene-lined autoclave and tightened. The mixture was then kept in a constant-temperature drying oven at 160 °C for 4 h for hydrothermal reaction. After cooling to room temperature, the reacted solution was centrifuged (8000 r, 5 min) to remove unreacted reagents. After centrifugation, the supernatant was collected. The supernatant was placed in a dialysis bag (500 Da) and dialyzed in deionized water for 24 h to remove excess silanizing reagents.

[0071] (3) The solution in the dialysis bag in step (2) is freeze-dried to obtain solid powder near-infrared fluorescent organosilicon quantum dots SiNDs.

[0072] Example 2

[0073] The near-infrared fluorescent organosilicon quantum dots (Si NDs) prepared in Example 1 of this invention were tested using methods including ultraviolet absorption, fluorescence, XRD, TEM, infrared, XPS, etc.

[0074] Figure 1 The images show near-infrared fluorescent organosilicon quantum dots (Si NDs) before and after the reaction. Using the method of Example 1, the near-infrared fluorescent organosilicon quantum dot solution after the hydrothermal reaction became significantly lighter in color, turning a light red, indicating that a reaction occurred after heating. The left side shows the solution before the reaction (i.e., the mixed solution from step 1 of Example 1), and the right side shows the solution after the reaction. Figure 2 The image shows a comparison of the fluorescence of the fluorescent organosilicon quantum dot aqueous solution prepared in Example 1 under sunlight and 660 nm red light excitation. It can be seen that it has strong fluorescence under 660 nm red light irradiation. Figure 3 The ultraviolet absorption spectrum of fluorescent organosilicon quantum dots, from Figure 3 As can be seen from the above, the fluorescent organosilicon quantum dot aqueous solution prepared in Example 1 has a strong absorption peak at 410 nm. Figure 4 Fluorescence spectra of the fluorescent molecule TCPP and fluorescent organosilicon quantum dots, from Figure 4 As can be seen from the above, the fluorescent organosilicon quantum dot solution prepared in Example 1 has a strong fluorescence emission peak at a wavelength of 655 nm. Figure 5 The XRD powder diffraction pattern of fluorescent organosilicon quantum dots (SiNDs) is shown below. Figure 5 It can be seen that the fluorescent organosilicon quantum dots prepared in Example 1 did not have obvious diffraction characteristic peaks. The diffraction peaks at around 20° to 30° were generated by the glass substrate, and the rest were some inconspicuous small and broad impurity peaks. This may be because the synthesized silicon quantum dots were amorphous or the quantum dots were small in size. Figure 6 The image shows the TEM spectrum of fluorescent organosilicon quantum dots (Si NDs). The quantum dots prepared in Example 1 have a size within 1-10 nm. Figure 7The infrared spectrum of fluorescent organosilicon quantum dots (SiNDs) is shown. Figure 7 The characteristic peaks of each element are indicated, demonstrating the functional groups present on the fluorescent organosilicon quantum dots. AEEA silane coupling reagent possesses amino groups, and Si NDs show a peak at 1640 cm⁻¹. -1 The presence of a distinct absorption peak at the position of the amide group indicates that the carboxyl group of the raw material TCPP reacts with the amino group of AEEA under high temperature conditions to generate an amide group. Figure 8 The XPS spectrum and peak fitting spectrum of the fluorescent organosilicon quantum dots are shown. The spectrum also shows that the fluorescent organosilicon quantum dots prepared in Example 1 mainly contain C and O elements, and are also doped with N and Si elements.

[0075] Figure 9 In step A, methylene blue (MB) reacts with a DPHA solution used for detecting reactive oxygen species in an aerobic environment under a fixed power of 0.5 W / cm². 2 The changes in ultraviolet absorption of the solution at different times after irradiation with 660 nm excitation light were recorded, providing a basis for calculating the singlet oxygen yield of the fluorescent organosilicon quantum dots prepared in Example 1. Figure 9 In the diagram, B and C represent TCPP and fluorescent organosilicon quantum dots prepared in Example 1, respectively, in a DPHA solution for detecting reactive oxygen species under an aerobic environment and a fixed power of 0.5 W / cm². 2 The UV absorption of the solution was recorded at different times after irradiation with 660 nm excitation light. The UV absorption peak of DPHA at 355 nm decreased with the increase of reactive oxygen species. Figure 9 Figure D shows the changes in the UV absorption peak of DPHA at 355 nm with time corresponding to Figures A, B, and C. The ROS yield of TCPP was 21.51%, and the ROS yield of near-infrared fluorescent silicon quantum dots was 49.87%, indicating that near-infrared fluorescent organosilicon quantum dots have a strong ability to generate singlet oxygen.

[0076] Figure 10 AC represents the relationship between the fluorescence intensity of TCPP and the near-infrared fluorescent organosilicon quantum dots prepared in Example 1 and pH value. Figure 10 It can be seen that the fluorescence intensity of the fluorescent molecule TCPP is greatly affected by the pH value. The prepared fluorescent organosilicon quantum dots are relatively stable in fluorescence within the pH range of 2-12, and the fluorescence intensity decay is not significant, indicating that the prepared fluorescent organosilicon quantum dots have good fluorescence stability.

[0077] Figure 11 The Michaelis-Menten steady-state dynamics of near-infrared fluorescent organosilicon quantum dots (Si NDs) were further investigated by the Chinese Academy of Sciences (CAS). Figure 11In samples A and B, TMB was used as a colorimetric reagent for determination, exhibiting characteristic absorbance at 652 nm. Specifically, different concentrations of H2O2 (1.25 × 10⁻⁶) were used. -5 1.875×10 -5 2.5×10 -5 and 3.75×10 -5 M) was added to PBS containing 1 mg / mL Si NDs and 0.8 mg / mL TMB, and the time-history absorbance of the reaction solution at 652 nm was plotted. Figure 11 In C, the relationship between the initial oxidation velocity ν0 of TMB and the H2O2 concentration was plotted to obtain the Michaelis-Menten curve. Figure 11 D is the Lineweaver-Burk fitting plot obtained through linear double reciprocal transformation, yielding Vmax and Km, which are 8.98125x10⁻¹⁰ respectively. -5 M / min and 3.30245x10 -3 M. This catalytic activity is stronger than that of horseradish peroxidase, demonstrating that Si NDs possess peroxidase properties and can efficiently catalyze the conversion of H2O2 to •OH. Si NDs can serve as high-performance nanozymes for chemokinetic therapy (CDT).

[0078] Example 3

[0079] The near-infrared fluorescent organosilicon quantum dots (Si NDs) prepared in Example 1 of this invention were subjected to an MTT cytotoxicity test. 4T1 cells were cultured overnight in 96-well plates, with a cell density of approximately 1 × 10⁻⁶ cells per well. 4 PBS, TCPP, and fluorescent organosilicon quantum dots prepared in Example 1 (TCPP and fluorescent organosilicon quantum dots were dispersed in PBS at different concentrations, and then diluted with cell culture medium at a 1:9 ratio) were added to 4T1 cells at final concentrations of 0, 6.25, 12.5, 25, 50, and 100 μg / mL (the amount of free TCPP added was kept the same as the amount of TCPP contained in the silicon dots). The cells were incubated with these solutions at 37°C for 24 hours. Then, MTT solution (MTT powder, prepared as a 5 mg / mL solution with sterile pH 7.4 PBS, and then diluted with cell culture medium at a 1:9 ratio) was added and incubated for 4 hours. The supernatant was discarded, the cells were washed twice with PBS, dissolved in 100 μL of DMSO for 5–10 min, transferred to a clean culture plate, and measured using a microplate reader to read the OD value. Cytotoxicity was detected using the MTT assay. Figure 12As shown in A and B, PBS and TCPP exhibit cell viability greater than or equal to 80% without corresponding 660 nm laser irradiation, demonstrating extremely low cytotoxicity. In contrast, fluorescent organosilicon quantum dots still exhibit strong CDT therapeutic effects without laser irradiation, with a cell killing rate reaching 50.51% at 100 μg / mL.

[0080] PBS, TCPP at a final concentration of 0-100 μg / mL, and near-infrared fluorescent organosilicon quantum dots prepared in Example 1 were mixed with 4T1 cells (4T1 cells were cultured overnight in 96-well plates, with a cell density of approximately 1 × 10⁶ cells per well). 4 After incubating together for 24 hours (keeping the amount of free TCPP added the same as that contained in the silicon spot), the solution was changed using 0.5 W / cm². 2 Irradiation with a 660 nm laser for 5 minutes corresponds to Figure 12 Error bar data in B and C represent the therapeutic effects of TCPP and fluorescent organosilicon quantum dots on PDT / CDT in 4T1 cells, derived from... Figure 12 As can be seen from C, the therapeutic effect of the fluorescent organosilicon quantum dots prepared in Example 1 on cells increases with increasing concentration, and at a concentration of 100 μg / mL, the cell survival rate is extremely low, and the cell death rate reaches 78.67% at 100 μg / mL, while the corresponding cell death rate of TCPP is 48.54%. The synergistic therapeutic effect of TCPP is significant, while the silane reagent alone does not have a therapeutic effect.

[0081] Example 4

[0082] The near-infrared fluorescent organosilicon quantum dots (Si NDs) prepared in Example 1 of this invention were subjected to live / dead cell staining tests. Four T1 cells (approximately 4 × 10⁻⁶ cells / mL) were pre-seeded in six confocal dishes. 4 (10 ... 2 Irradiate the cells under a 660 nm laser for 5 minutes (5 seconds of irradiation followed by a 5-second pause). Add calcein AM (1 μM) and PI (2 μM) to all dishes and incubate for 30 minutes. Aspirate the supernatant, wash the cells three times with PBS, and then add 1 mL of PBS for fluorescence confocal imaging. Figure 13The first two rows show AM / PI fluorescence imaging of live and dead cells in 4T1 cells before and after 660 nm laser irradiation with PBS and the fluorescent molecule TCPP. Figure 13 The third row shows AM / PI fluorescence imaging of live and dead cells of fluorescent organosilicon quantum dots in 4T1 cells before and after 660 nm laser irradiation. Before laser irradiation, PBS and TCPP had little effect on the cells, with only a few cells dying. However, due to its own chemical kinetics, the Si NDs group still showed some cell apoptosis even without laser irradiation, appearing red. After laser irradiation, the fluorescent organosilicon quantum dots prepared in Example 1 showed a large proportion of cell apoptosis, with a significantly stronger killing effect than the TCPP group after laser irradiation. These results indicate that the synthesized fluorescent organosilicon quantum dots have a high killing power against tumor cells through PDT / CDT synergistic therapy.

[0083] Example 5

[0084] Cellular ROS detection was performed on the near-infrared fluorescent organosilicon quantum dots (SiNDs) prepared in Example 1 of this invention. Four T1 cells (approximately 4 × 10⁻⁶ cells / mL) were pre-seeded in three cell culture dishes. 4 (One sample was taken). TCPP and the lyophilized fluorescent organosilicon quantum dot powder from Example 1 were dissolved in deionized water to form an aqueous solution. This solution was then diluted with cell culture medium at a ratio of 1:9 to achieve a final concentration of 100 μg / mL of fluorescent organosilicon quantum dots in the medium (the amount of free TCPP added was kept the same as that contained in the silicon dots). After incubation for 24 hours, the solution was added at 0.5 W / cm². 2 Irradiate the cells with a 660 nm laser for 5 min (5 s irradiation followed by a 5 s pause), then change the medium and add 2,7-dichlorodiacetic acid dichlorofluorescein (DCFH-DA). Detect the generation of ROS in 4T1 cells after incubation with fluorescent organosilicon quantum dots (SiNDs). Specifically, dilute DCFH-DA 1:1000 with DMEM cell culture medium, add 1 ml of DCFH diluent to each dish, incubate at 37°C for 20 min, and then wash the cells three times with serum-free culture medium to thoroughly remove any DCFH-DA that has not entered the cells. Figure 14 The images show DCFH staining patterns of 4T1 cells co-incubated with PBS, TCPP, and fluorescent organosilicon quantum dots prepared in Example 1, respectively. The results show that the green fluorescence of cells incubated with Si NDs was significantly enhanced compared to the control group, corresponding to the results of Examples 3 and 4.

[0085] Example 6

[0086] The near-infrared fluorescent organosilicon quantum dots (Si NDs) prepared in Example 1 of this invention were subjected to a live / dead bacterial staining test. *Staphylococcus aureus* (OD=0.5) was pre-inoculated into four confocal dishes and incubated at 37°C for 6 hours. Then, TCPP and the lyophilized fluorescent organosilicon quantum dot powder from Example 1 were dissolved in deionized water to form an aqueous solution. This solution was then mixed with a nutrient broth solution (1.8 g of solid dissolved in 100 ml of deionized water, dispensed into Erlenmeyer flasks, and autoclaved at 121°C for 15 minutes) at a ratio of 1:9 to dilute the Si NDs in the culture medium to a final concentration of 100 μg / mL (the amount of free TCPP added was kept the same as the amount of TCPP contained in the silicon dots). Two dishes were added to each material. After co-incubating with bacteria at 37°C for 6 hours, one dish corresponding to each material was incubated at 0.5 W / cm². 2 Irradiate the cells under a 660 nm laser for 5 minutes (5 seconds of irradiation followed by a 5-second pause). PBS serves as a control group. All dishes are incubated for 30 minutes with calcein AM (1 μM) and PI (2 μM). Calcein AM fluoresces live bacteria with intact cell membranes green, while PI fluoresces dead bacteria with damaged cell membranes red. Figure 15 As shown, the PBS group, PBS+light group, and TCPP group emitted green fluorescence, while the red fluorescence was weak. In the group treated with fluorescent organosilicon quantum dots (Si NDs), some areas were marked green, and some were marked green, indicating that Si NDs still exhibited a certain CDT therapeutic effect even without light irradiation. When bacteria were treated with Si NDs+light, the number of red spots increased significantly, indicating that the large amount of singlet oxygen generated by the fluorescent organosilicon quantum dots under the action of oxygen and light significantly enhanced the bactericidal effect. After the introduction of light, almost no green spots were observed, and most bacteria showed red fluorescence, with a significantly better effect than TCPP+light irradiation. This result proves that fluorescent organosilicon quantum dots (Si NDs) have a superior PDT / CDT sterilization effect compared to pure TCPP, while silane reagents alone do not possess antibacterial effects.

[0087] Example 7

[0088] In vitro antibacterial tests were performed on the near-infrared fluorescent organosilicon quantum dots (SiNDs) prepared in Example 1 of this invention. 800 μL of *Staphylococcus aureus* and *Escherichia coli* (OD value 1) bacterial suspension were incubated with 200 μL of PBS and solutions of TCPP and SiNDs at 37°C for 6 h. The final concentration of SiNDs was 100 μg / mL. The amount of free TCPP added was kept the same as that contained in the silicon dots. Irradiation and non-irradiation treatments were performed, with irradiation involving changing the solution after incubation and then applying 0.5 W / cm³ of PBS. 2Irradiate the bacteria under a 660 nm laser for 5 minutes (5 seconds of irradiation followed by a 5-second pause). Spread 10 μL of the final-treated bacterial suspension onto solid agar medium (3.3 g of solid was dissolved in 100 ml of deionized water, dispensed into Erlenmeyer flasks, and autoclaved at 121 °C for 15 minutes). Incubate at 37 °C for 18 h and count the colonies. Si NDs are a potential antibacterial agent for PDT treatment of bacterial infections. Therefore, the in vitro antibacterial activity of fluorescent organosilicon quantum dots against Gram-positive Staphylococcus aureus and Gram-negative Escherichia coli was evaluated. Figure 16 As shown in Figure A, the PBS group and the PBS+light irradiation group formed a large number of live bacterial colonies, indicating that surface light irradiation alone did not affect the bacteria. However, the Si NDs group, even without laser irradiation, still showed some antibacterial effect after CDT treatment. Figure 16 According to B Figure 16 The survival rate of the corresponding colonies was obtained by counting using A and ImageJ software. The TCPP group and the TCPP+ light-illuminated group showed antibacterial activity, but the effect was weak. After light treatment, the inhibition rate of the TCPP+ light-illuminated group for *Staphylococcus aureus* was 43.45%, and the inhibition rate of the TCPP+ light-illuminated group for *Escherichia coli* was 38.63%. Clearly, due to the chemical kinetics (CDT) effect, the Si NDs material itself can kill bacteria. The Si NDs+ light group showed better antibacterial effect than other groups, achieving an inhibition rate of 92.7% in *Staphylococcus aureus*. In terms of killing *Escherichia coli*, the corresponding treatment groups showed the same antibacterial trend as *Staphylococcus aureus*. The inhibition rate of the Si NDs+ light group for *Escherichia coli* reached 84.66%, confirming that the fluorescent organosilicon quantum dots prepared in Example 1 of this invention can be applied to antibacterial experiments and have a highly efficient antibacterial effect.

[0089] Example 8

[0090] Inspired by the in vitro bactericidal effect and biocompatibility of Si NDs, this study further investigated the in vivo antibacterial effect of near-infrared fluorescent organosilicon quantum dots (NIVs) using a mouse subcutaneous abscess model infected with Staphylococcus aureus. BALB / c mice (6 weeks old, weighing approximately 20 grams) were used. To establish an abscess xenograft model, 100 μL (103 g2) was subcutaneously injected into the upper right leg region of BALB / c mice. 7CFU / mL) S-aureus. After 48 hours, subcutaneous abscesses formed in mice. Mice were grouped according to abscess size (4 mice per group): PBS group, TCPP group, Si NDs group, TCPP + Laser group, and Si NDs + Laser group. 30 μL of material (100 μg / mL) was injected for 15 minutes, followed by 0.5 W / cm². 2 The abscesses were irradiated with 660 nm light for 5 min (5 s irradiation followed by a 5 s pause), with the amount of free TCPP added maintained at the same level as that contained in the silicon dots. Abscesses were photographed and weighed daily. After 10 days of treatment, skin tissue, heart, liver, spleen, lung, and kidney tissues were collected, fixed with 4% paraformaldehyde, and analyzed by H&E staining. Figure 17A shows the in vivo antibacterial efficacy of fluorescent organosilicon quantum dots, along with photographs and corresponding histological sections of *Staphylococcus aureus* abscesses treated with PBS, TCPP, Si NDs, TCPP + Laser, and Si NDs + Laser. The scale bar is 50 μm. The results showed that the scar rate in the Si NDs + Laser group was significantly better than other groups. After 10 days of treatment, the scars in the Si NDs + Laser group disappeared, and the wound was basically healed, indicating that the material played an important role in wound healing due to the synergistic effect of CDT and PDT. Furthermore, TUNEL analysis of the wound on the tenth day (Figure 17B) showed a significant improvement in the repair treatment effect. H&E staining further evaluated the healing of infected wounds. Figure 17C shows the histological observation of normal tissues (heart, liver, spleen, lung, and kidney) after Si NDs treatment, with a scale bar of 50 μm. The Si NDs + Laser group showed an intact epidermal layer and fewer inflammatory cells in the wound tissue, while the other groups showed disordered epidermal layers and significant inflammatory cell infiltration. Therefore, the antibacterial effect of fluorescent organosilicon quantum dots under aerobic conditions is further demonstrated.

[0091] Example 9

[0092] Further evaluation of the antitumor efficacy of Si NDs was conducted. In vivo antitumor experiments were performed using 4T1 tumor-bearing mice. First, all mice were BALB / c mice (6 weeks old, weighing approximately 20 grams). The mice were divided into five groups of four mice each: Control group (PBS group) (G1), TCPP group (G2), Si NDs (near-infrared fluorescent organosilicon quantum dots prepared in Example 1) group (G3), TCPP + Laser group (G4), and Si NDs + Laser (fluorescent organosilicon quantum dots prepared in Example 1 + laser) group (G5). Approximately 1×10⁻⁶ ppm of Si NDs were suspended in 20 µL of PBS. 6 4T1 cells were injected subcutaneously into the right leg of mice. When the tumor volume reached approximately 50 mm... 3The Si NDs group and the Si NDs + Laser group were injected via tail vein with 100 μL of a fluorescent organosilicon quantum dot aqueous solution at a concentration of 100 μg / mL. The amount of free TCPP added was kept the same as that contained in the silicon dots. The control group (PBS group) was injected with an equal volume of PBS. For the Si NDs + Laser group, after the material was injected, a 0.5 W / cm² solution was used. 2 The tumor site was irradiated with a 660 nm laser for 5 minutes (5 seconds of irradiation followed by a 5-second pause, for a total of 5 minutes). For all groups, tumor volume and mouse weight were recorded every two days, and the tail vein was injected every three days for a total of 12 days. Afterward, the mice were dissected, solid tumors were removed for size comparison, and the heart, liver, spleen, lung, and kidney were removed for section analysis. Tunel and H&E analyses were performed on the tumors.

[0093] Figure 18 Fluorescence imaging of live mice was performed using tail vein injection of fluorescent organosilicon quantum dots (TCPP). Due to the fluorescence properties of TCPP, fluorescence imaging of mouse tumor sites was achieved using the FOBI live animal imaging system (NeoScience, South Korea). Due to the EPR effect of the nanoparticles, Si NDs accumulated in the tumor. Mice were sacrificed 72 hours after administration, and in vitro fluorescence imaging (of the resected major organs and tumors) was performed. Figure 19 As shown, after Si NDs were administered, bright fluorescence was visible on the tumor, while the fluorescence on the heart, spleen, kidney, and tumors treated with DMNRs was relatively weak.

[0094] Figure 20 Compared with live mice in the PBS group, TCPP group, Si NDs group, TCPP + Laser group and Si NDs + Laser group, the tumor size in the treatment group (Si NDs + Laser group) was significantly smaller than that in the other four groups, indicating that near-infrared fluorescent organosilicon quantum dots have a significant therapeutic effect on mouse tumors under laser irradiation, and the synergistic effect of PDT / CDT is obvious.

[0095] Figure 21 The graph shows the change in mouse weight. The weight change of all mice was not significant over 12 days, indicating that fluorescent organosilicon quantum dots have no toxic side effects on mice.

[0096] Figure 22 The tumor volume changes in mice over 12 days showed that the tumor size in the Si NDs + Laser group was very small and almost unchanged, while the tumor size in the other four groups increased significantly, indicating that fluorescent organosilicon quantum dots have a significant synergistic effect of PDT / CDT on the tumor region of mice.

[0097] Figure 23The tumor size of the five groups was compared after dissecting the mice. The tumor size of the Si NDs + Laser group was significantly smaller than that of the Control group (PBS group) and TCPP group, indicating that fluorescent organosilicon quantum dots have a significant therapeutic effect on mouse tumors under laser irradiation.

[0098] Example 10

[0099] After dissecting the mice in Example 9, tumor masses were removed, and the weight of the tumor in each mouse was recorded. The TCPP group, Si NDs group, TCPP + Laser group, and Si NDs + Laser group were compared with the Control group (PBS group). The tumor inhibition rates of the TCPP group, Si NDs group, and TCPP + Laser group were calculated to be 20.01%, 55.05%, and 65.75%, respectively. The Si NDs + Laser group had the highest tumor inhibition rate of 84.76%. Figure 24 The comparison of tumor inhibition rates by photothermal therapy demonstrates that fluorescent organosilicon quantum dots exhibit outstanding synergistic PDT / CDT effects on tumor regions in mice.

[0100] Example 11

[0101] H&E staining: After dissecting the mice treated in Example 9, tissue blocks from the heart, liver, spleen, lung, kidney, and tumor were removed and placed in a pre-prepared fixative (10% formalin). After successful fixation, the blocks were trimmed to 25px × 25px × 5px, placed in an embedding cassette, and rinsed with running water (to remove the fixative) for 30 min. The tissue blocks were gradually dehydrated using alcohol, gradually increasing the concentration. The tissue blocks were then cleared in xylene, a clearing agent soluble in both alcohol and paraffin, replacing the alcohol in the tissue blocks with xylene. The cleared tissue blocks were then placed in melted paraffin and kept warm in a paraffin bath. After the paraffin had completely penetrated the tissue blocks, they were embedded and allowed to cool and solidify. Once the embedded tissue blocks had hardened, they were sliced ​​into very thin sections (5-8 μm) using a microtome. The slices were then flattened in hot water, mounted on glass slides, and dried in a 45°C oven. Before staining, the paraffin in the sections was removed with xylene. The sections were then passed through high-concentration to low-concentration alcohol, followed by staining in distilled water and then in a hematoxylin solution for several minutes. They were then separated in acidic and ammonia solutions for a few seconds each, rinsed with running water for 1 hour, briefly immersed in distilled water, and then dehydrated in 70% and 90% alcohol for 10 minutes each. Finally, eosin ethanol staining solution was added for 2-3 minutes. After staining, the sections were dehydrated with pure alcohol and then cleared with xylene. Resin was dripped onto the cleared sections, and they were sealed with coverslips. After the resin had slightly dried, labels were attached, and the specimens could then be observed under a microscope.

[0102] TUNEL staining: Add 100 μL of TUNEL Equilibration Buffer to each sample and incubate at room temperature for 5 min. Prepare the TUNEL reaction mixture in advance: Add 1 μL of TdT enzyme to every 50 μL of TUNEL Reaction Buffer. Discard the TUNEL Equilibration Buffer and add 50 μL of TUNEL reaction mixture to each sample. Use a coverslip to evenly cover the sample with the buffer. Place the sample flat in a humidified chamber and incubate at 37°C for 2 h (place a slightly damp paper towel at the bottom of the chamber to maintain humidity). Then remove the reaction solution and rinse twice in a staining container of 1×PBS for 5 min each time. Next, wash the sample three times for 5 min each time with an appropriate amount of 0.1% Triton X-100 buffer containing 5 mg / mL BSA in PBS to reduce background. Observe using a fluorescence microscope or flow cytometry.

[0103] Figure 25 The images show the tumors in mice: TUNEL (a) and H&E (b). The tumors in the G3 group showed mild necrosis, while the tumors in the G5 treatment group showed extensive necrosis. Unstructured eosinophilic material and a large number of necrotic cell fragments were visible around the necrotic areas, confirming the effective therapeutic effect of Si NDs + Laser.

[0104] Figure 26 The analysis of mouse tissue sections shows tissue sections of the heart, liver, spleen, lung, and kidney (from left to right). The top and bottom sections are a comparison between the Control group (PBS group) and the Treatment group (Si NDs + Laser group). The results show that the pathological damage caused by the material to the major organs is negligible, indicating that the fluorescent organosilicon quantum dots prepared in Example 1 of this invention have no effect on the heart, liver, spleen, lung, and kidney of mice.

Claims

1. A near-infrared fluorescent silicon dot, characterized in that, The near-infrared fluorescent silicon dots are generated by a one-step hydrothermal method through a high-temperature condensation reaction between the carboxyl group of 4-carboxyphenylporphyrin (TCPP) and the amino group of a silane coupling agent; the silane coupling agent includes any one or more of diethylenetriaminepropyltrimethoxysilane (AEEA), 3-aminopropyltriethoxysilane (AMEO), 3-aminopropyltrimethoxysilane (APTMS), and N-(β-aminoethyl)-γ-aminopropyltrimethoxysilane (DAMO).

2. The near-infrared fluorescent silicon dots according to claim 1, characterized in that, The diameter of the near-infrared fluorescent silicon dots is in the range of 1-10 nm.

3. The near-infrared fluorescent silicon dots according to claim 1, wherein, The near-infrared fluorescent silicon dots exhibit fluorescence properties, with a strong fluorescence emission peak in the near-infrared band of 650 nm-700 nm.

4. A method of preparing near-infrared fluorescent silicon dots according to claim 1, characterized in that, Includes the following steps: (1) The reaction is carried out by a one-step hydrothermal method using 4-carboxyphenylporphyrin (TCPP), silane coupling reagent, and deionized water; (2) The solid powder obtained by centrifugation and dialysis of the reaction solution in step (1) followed by freeze drying is a near-infrared fluorescent silicon dot.

5. The preparation method according to claim 4, characterized in that, In step (1), TCPP and deionized water are first mixed at a mass ratio of 1-3:400, and then silane coupling reagent is added to the mixture and ultrasonicated to make it uniform; the molar ratio of TCPP to silane coupling reagent is 1:80-120.

6. The preparation method according to claim 4, characterized in that, The reaction temperature of the one-step hydrothermal method in step (1) is 140-200℃, and the time is 3-8 hours.

7. The application of the near-infrared fluorescent silicon dot according to claim 1 in the preparation of antibacterial and fluorescent imaging reagents for biological wounds.

8. The use of the near-infrared fluorescent silicon dot according to claim 1 in the preparation of photodynamic agents for tumor treatment.

9. The use of the near-infrared fluorescent silicon dot according to claim 1 in the preparation of a chemokinetic agent for tumor treatment.

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