A FePc modified Ti3C2T x Preparation and application of Rh-loaded nanozymes

Abstract

The present invention introduces a FePc modified Ti3C2T x Preparation and application of Rh-loaded nanozymes, whose main components are Rh‑FePc‑Ti3C2T x , the Rh‑FePc‑Ti3C2T x The raw materials for preparation include FePc, Ti3C2T x and RhCl3·3H2O. The preparation method includes: FePc and Ti3C2T x After mixing, continuous ultrasonication was performed; washing and drying were performed to obtain FePc‑Ti3C2T x ; In FePc‑Ti3C2T x RhCl3·3H2O was added to the mixture and dissolved in polyethylene glycol; ultrasonic dispersion was performed and the mixture was transferred to a reaction kettle; cooling, washing and centrifugation were performed; drying and grinding were performed to obtain Rh‑FePc‑Ti3C2T x Composite nanozyme. The present invention sequentially loads FePc and metal Rh onto two-dimensional Ti3C2T x Nanosheets, synthesized material Rh‑FePc‑Ti3C2T x As a new type of composite nanozyme, it is used to regulate the complex tumor microenvironment of osteosarcoma cells and produce ROS, alleviate the hypoxia problem of tumor cells, and at the same time provide a new research platform for the synergistic treatment of osteosarcoma through photodynamic therapy and sonodynamic therapy.

Description

Technical Field

[0001] The present invention belongs to the field of tumor drug preparation, and specifically relates to a FePc-modified Ti3C2T x Preparation and application of Rh-loaded nanozymes. Background Art

[0002] At present, osteosarcoma is the most common primary bone sarcoma, usually diagnosed in adolescence. It is one of the most threatening malignant tumors with poor prognosis and treatment effects, which seriously affects the normal quality of life of patients. The tumor microenvironment is hypoxic and weakly acidic, and the concentration of hydrogen peroxide is too high. Reactive oxygen species ROS mainly include active superoxide anions, hydroxyl free radicals, and singlet oxygen. Increasing the level of ROS in the tumor environment and enhancing the oxidative stress of cancer cells will lead to tumor cell damage and apoptosis. Therefore, regulating the complex microenvironment of the tumor to generate ROS-mediated treatments is a more feasible method for treating osteosarcoma, such as photodynamic therapy PDT and sonodynamic therapy SDT.

[0003] Clinically, the treatment of osteosarcoma is mainly surgical resection, adjuvant radiotherapy and chemotherapy, etc., but the treatment effect is not good and the prognosis is uncontrollable. In recent years, some nanocatalytic therapies can be used for the treatment of tumors, which is a new treatment method based on nanozymes. Nanozymes are a type of artificial enzymes that have the unique properties of nanomaterials and catalytic functions. Due to their economic, stable and durable advantages, they are widely used in medicine, chemical industry, environment and other fields. Titanium carbide (Ti3C2T x ) is a good nanocarrier load that can be used to load nanoenzymes. Chinese patent CN112903763A discloses a Ti3C2Tx-based ammonia sensor preparation method and its light-assisted application. x The preparation process of ammonia gas sensor includes synthesizing Ti3C2T x , TiO2 / Ti3C2T was synthesized by hydrothermal method x The composite thin film sensor for detecting volatile gas components of meat was prepared by screen printing. The obtained gas sensor can achieve better performance in the application of ammonia detection with the help of ultraviolet light, and is expected to be applied to meat freshness monitoring, realizing the identification and concentration detection of volatile gases in fresh meat, which has important practical significance for ensuring the food safety of meat.

[0004] Chinese patent CN110974978A discloses a nanocatalyst for tumor treatment and its preparation method and application. The nanocatalyst provided includes an erythrocyte membrane and a composite nanozyme and a photosensitizer coated in the erythrocyte membrane; the composite nanozyme includes glucose oxidase and iron nanoparticles wrapped in the inner cavity of the glucose oxidase. The nanocatalyst preferentially accumulates at the target tumor site through targeted biomimetic delivery, and releases the composite nanozyme under near-infrared light irradiation; based on the high glucose uptake and weakly acidic environment at the tumor site, glucose oxidase converts glucose into H2O2, inducing iron nanoparticles to initiate the in situ Fenton reaction, and sequentially catalyzes to produce hydroxyl radicals, inducing oxidative damage to tumor cells, and then killing tumor cells. The nanocatalyst can not only achieve efficient loading of the catalyst, but also effectively prolong the circulation time in the body, and achieve accurate and continuous release at the tumor lesion site, providing a new idea and platform for tumor treatment.

[0005] However, there are many problems in the preparation of some nanozymes at present, such as complex preparation process and large size. There are also many difficulties in the nanozymes themselves, such as being difficult to prepare, having poor water solubility and low biocompatibility. Summary of the invention

[0006] To solve the above problems, we aim to prepare a nanozyme that has good water solubility, small size, uniform dispersion, easy preparation and high in vivo biocompatibility, so as to change the complex tumor microenvironment of osteosarcoma cells, produce ROS, alleviate tumor hypoxia, and achieve the effect of treating osteosarcoma.

[0007] In order to achieve the above effect, the present invention designs a FePc-modified Ti3C2T x Preparation and application of Rh-loaded nanozymes.

[0008] A FePc modified Ti3C2T x Preparation and application of Rh-loaded nanozymes, the main components of which are Rh-FePc-Ti3C2T x , the Rh-FePc-Ti3C2T x The raw materials for preparation include FePc, Ti3C2T x Nanosheets and RhCl3·3H2O; the FePc, Ti3C2T x The mass ratio of the nanosheets is 2-8:1-4, preferably 2:1.

[0009] A FePc modified Ti3C2T x The preparation and application method of the Rh-loaded nanozyme comprises:

[0010] S1, FePc and Ti3C2T x Continuous sonication after mixing;

[0011] S2, washing and drying to obtain FePc-Ti3C2T x ;

[0012] S3, in FePc-Ti3C2T x Add RhCl3·3H2O and dissolve them together in polyethylene glycol;

[0013] S4, ultrasonic dispersion and transfer to a reaction kettle;

[0014] S5, cooling, washing and centrifugation;

[0015] S6, dry and grind to obtain Rh-FePc-Ti3C2T x Composite nanozymes.

[0016] Preferably, in the step S1, FePc and Ti3C2T x The required material ratio is 10-40 mg FePc: 5-20 mg Ti3C2T x : 6-10ml ultrapure water.

[0017] Preferably, in the step S1, the continuous ultrasonication condition after mixing is continuous ultrasonication for 4-8 hours at room temperature.

[0018] Preferably, in the step S2, washing and drying are specifically performed by washing with anhydrous ethanol, centrifuging twice, and then placing in a vacuum drying oven for 12-24 hours to perform FePc-Ti3C2T x collect.

[0019] Preferably, in the step S3, FePc-Ti3C2T x , RhCl3·3H2O, polyethylene glycol in a ratio of 8-12 mg FePc-Ti3C2T x : 3-6 ml RhCl3·3H2O with a concentration of 2 mg / mL: 20-40 ml polyethylene glycol with a number average molecular weight of 200.

[0020] Preferably, in the ultrasonic dispersion in step S4, the ultrasonic time is 1-2 h, until the solution is evenly dispersed.

[0021] Preferably, the reaction time in the reactor of step S4 is 12-16 hours, and the reaction temperature is 160-200°C.

[0022] Preferably, the washing material in step S5 is anhydrous ethanol, and the centrifugation is performed three times.

[0023] Preferably, in step S6, the specific method of drying is placing in a vacuum drying oven for drying, and the drying time is 12-24 hours.

[0024] The advantages and effects of this application are as follows:

[0025] 1. The present invention sequentially loads FePc and metal Rh onto two-dimensional Ti3C2T x Composite nanomaterials Rh-FePc-Ti3C2T synthesized on nanosheets x As a new type of composite nanozyme, it has the advantages of small size, high stability, uniform dispersion and good photothermal performance.

[0026] 2. Rh-FePc-Ti3C2T prepared in this application x The composite nanozyme has catalase-like activity and peroxidase activity, and has good biocompatibility. It regulates and changes the complex tumor microenvironment of osteosarcoma cells, produces ROS and thus alleviates tumor hypoxia.

[0027] 3. Rh-FePc-Ti3C2T prepared in this application x Composite nanozymes can be used as a new type of photo / sonosensitizer, which can catalyze the production of ROS through photodynamic and sonodynamic therapy in the tumor microenvironment, alleviate the hypoxia of tumor cells, and provide a new research platform for composite nanozymes to regulate the tumor microenvironment, produce ROS, and synergistically treat osteosarcoma through photodynamic therapy and sonodynamic therapy.

[0028] The above description is only an overview of the technical solution of the present application. In order to more clearly understand the technical means of the present application so that it can be implemented in accordance with the contents of the specification, and to make the above and other purposes, features and advantages of the present application more obvious and easy to understand, the following is a detailed description of the preferred embodiments of the present application in conjunction with the accompanying drawings as follows.

[0029] Based on the detailed description of the specific embodiments of the present application in combination with the accompanying drawings below, those skilled in the art will become more aware of the above and other objects, advantages and features of the present application. BRIEF DESCRIPTION OF THE DRAWINGS

[0030] In order to more clearly illustrate the technical solutions in the embodiments of the present application or the prior art, the drawings required for use in the embodiments or the prior art descriptions are briefly introduced below. Obviously, the drawings described below are some embodiments of the present application. For ordinary technicians in this field, other drawings can also be obtained based on these drawings without creative work. In all drawings, similar elements or parts are generally identified by similar reference numerals. In the drawings, each element or part is not necessarily drawn according to the actual scale.

[0031] Figure 1 A composite nanozyme Rh-FePc-Ti3C2T provided by the present invention x Flow chart of composite nanozyme preparation;

[0032] Figure 2 The Rh-FePc-Ti3C2T provided by the present invention x High-resolution transmission electron microscopy (HRTEM) images of solids;

[0033] Figure 3 The Rh-FePc-Ti3C2T provided by the present invention x Catalase-like activity detection diagram;

[0034] Figure 4 The Rh-FePc-Ti3C2T provided by the present invention x Peroxidase activity detection diagram;

[0035] Figure 5 The Rh-FePc-Ti3C2T provided by the present invention x Biocompatibility test diagram;

[0036] Figure 6 The Rh-FePc-Ti3C2T provided by the present invention x Effect diagram of killing 143B tumor cells by photodynamic therapy and sonodynamic therapy;

[0037] Figure 7 The Rh-FePc-Ti3C2T provided by the present invention x Effect diagram of the ability to produce ROS in cells. DETAILED DESCRIPTION

[0038] To make the purpose, technical scheme and advantages of the embodiment of the present application clearer, the technical scheme in the embodiment of the present application will be clearly and completely described below in conjunction with the accompanying drawings in the embodiment of the present application. Obviously, the described embodiment is a part of the embodiment of the present application, rather than all of the embodiments. In the following description, specific details such as specific configuration and components are provided only to help fully understand the embodiments of the present application. Therefore, it should be clear to those skilled in the art that various changes and modifications can be made to the embodiments described herein without departing from the scope and spirit of the present application. In addition, for clarity and brevity, the description of known functions and structures is omitted in the embodiment.

[0039] It should be understood that the references to "one embodiment" or "this embodiment" throughout the specification mean that the specific features, structures, or characteristics associated with the embodiment are included in at least one embodiment of the present application. Therefore, the references to "one embodiment" or "this embodiment" appearing throughout the specification do not necessarily refer to the same embodiment. In addition, these specific features, structures, or characteristics may be combined in any suitable manner in one or more embodiments.

[0040] In addition, the present application may repeat reference numerals and / or letters in different examples. This repetition is for the purpose of simplicity and clarity, and does not in itself indicate the relationship between the various embodiments and / or settings discussed.

[0041] The term "and / or" in this article is only a description of the association relationship of associated objects, indicating that there can be three relationships. For example, A and / or B can mean: A exists alone, B exists alone, and A and B exist at the same time. The term " / and" in this article describes another type of association object relationship, indicating that there can be two relationships. For example, A / and B can mean: A exists alone, and A and B exist alone. In addition, the character " / " in this article generally indicates that the previous and next associated objects are in an "or" relationship.

[0042] The term "at least one" in this article is merely a description of the association relationship of associated objects, indicating that there may be three relationships. For example, at least one of A and B can mean: A exists alone, A and B exist at the same time, and B exists alone.

[0043] It should also be noted that, in this article, relational terms such as first and second, etc. are only used to distinguish one entity or operation from another entity or operation, and do not necessarily require or imply any such actual relationship or order between these entities or operations. Moreover, the terms "include", "comprises" or any other variations thereof are intended to cover non-exclusive inclusions.

[0044] Example 1

[0045] This example mainly introduces a FePc-modified Ti3C2T x Preparation and application of Rh-loaded nanozymes.

[0046] A composite nanozyme Rh-FePc-Ti3C2T x , whose main composition is Rh-FePc-Ti3C2T x , the Rh-FePc-Ti3C2T x The raw materials for preparation include FePc, Ti3C2T x and RhCl3·3H2O.

[0047] A composite nanozyme Rh-FePc-Ti3C2T x For the specific preparation process, please refer to Figure 1 , Figure 1 A composite nanozyme Rh-FePc-Ti3C2T provided by the present invention x Preparation flow chart; the preparation method comprises:

[0048] S1, FePc and Ti3C2Tx Continuous sonication after mixing;

[0049] S2, washing and drying to obtain FePc-Ti3C2T x ;

[0050] S3, in FePc-Ti3C2T x Add RhCl3·3H2O and dissolve them together in polyethylene glycol;

[0051] S4, ultrasonic dispersion and transfer to a reaction kettle;

[0052] S5, cooling, washing and centrifugation;

[0053] S6, dry and grind to obtain Rh-FePc-Ti3C2T x Composite nanozymes.

[0054] Furthermore, in the step S1, FePc and Ti3C2T x The materials required for mixing are 10-40 mg of FePc, 5-20 mg of Ti3C2T x And 6-10ml ultrapure water.

[0055] Furthermore, in the step S1, the continuous ultrasonication condition after mixing is continuous ultrasonication at room temperature for 4-8 hours.

[0056] Furthermore, in the step S2, washing and drying are specifically performed by washing with anhydrous ethanol, centrifuging twice, and then placing in a vacuum drying oven for 12-24 hours to perform FePc-Ti3C2T x collect.

[0057] Furthermore, in the step S3, the FePc-Ti3C2T x The mass is 8-12 mg, the RhCl3·3H2O is 3-6 ml, and the polyethylene glycol is 20-40 ml.

[0058] Furthermore, in the ultrasonic dispersion in step S4, the ultrasonic time is 1-2 hours until the solution is evenly dispersed.

[0059] Furthermore, the reaction time in the reactor of step S4 is 12-16 hours, and the reaction temperature is 160-200°C.

[0060] Furthermore, the washing material in step S5 is anhydrous ethanol, and the number of centrifugation is three times.

[0061] Furthermore, in the step S6, the specific method of drying is placing in a vacuum drying oven for drying, and the drying time is 12-24 hours.

[0062] The present invention sequentially loads FePc and metal Rh onto two-dimensional Ti3C2T x As a new type of composite nanozyme, the composite nanomaterial synthesized on the nanosheet has the advantages of small size, uniform dispersion, high stability and good photothermal performance.

[0063] Rh-FePc-Ti3C2T prepared in this application x The composite nanozyme has catalase-like activity and peroxidase activity, and has good biocompatibility. It can regulate the complex tumor microenvironment of osteosarcoma cells, produce ROS, and alleviate tumor hypoxia.

[0064] Rh-FePc-Ti3C2T prepared in this application x Composite nanozymes can be used as a new type of photo / sound sensitizer. They can catalyze the production of ROS in the tumor microenvironment through photodynamic and sonodynamic therapy, which can relieve the hypoxia of tumor cells. This provides a new research platform for composite nanozymes to regulate the tumor microenvironment, produce ROS, and use photodynamic and sonodynamic therapy to synergistically treat osteosarcoma. In addition, they can be used as photoacoustic sensitizers. Under near-infrared light and ultrasound irradiation, they can regulate the complex microenvironment of tumors, decompose hydrogen peroxide to produce O2 and OH-, and relieve the hypoxic environment of tumors.

[0065] Example 2

[0066] Based on the above Example 1, this example mainly introduces Rh-FePc-Ti3C2T x Verification of the effect of composite nanozymes.

[0067] Figure 2 The Rh-FePc-Ti3C2T provided by the present invention x High-resolution transmission electron microscopy (HRTEM) image of a solid; The morphology, size and distribution of the nanomaterial were observed using a high-resolution transmission electron microscope (HRTEM). The results showed that the nanomaterial was uniformly distributed nanoparticles with a size of about 5 nm.

[0068] Figure 3 The Rh-FePc-Ti3C2T provided by the present invention x Catalase-like activity detection diagram; The catalase-like activity detection of nanozyme was detected by dissolved oxygen instrument. It can be seen that when the concentration is 40ug / mL, the nanozyme Rh-FePc-Ti3C2T x The concentration of oxygen produced by decomposing hydrogen peroxide can reach 19 mg / L, which is higher than that of FePc-Ti3C2T x and Ti3C2T x The concentration of oxygen produced by the decomposition of hydrogen peroxide by the nanozyme is dependent on the concentration, indicating that the nanozyme has good catalase-like activity.

[0069] Figure 4 The Rh-FePc-Ti3C2T provided by the present invention x Peroxidase activity detection diagram; TMB was used as a probe to test the peroxidase activity to evaluate the decomposition of hydrogen peroxide by nanozymes to produce OH - The ability of nanozyme Rh-FePc-Ti3C2T x The absorption peak at 652 nm was the lowest at 21 min, indicating the generation of OH - The ability of the nanozyme was the strongest and time-dependent. The results showed that the nanozyme had good peroxidase activity.

[0070] Figure 5 The biocompatibility test diagram of Rh-FePc-Ti3C2Tx provided by the present invention; the biocompatibility of the material was evaluated by CCK-8 experiment after co-culturing the nanozyme Rh-FePc-Ti3C2Tx with HUVECs cells and 143B cells for 24 hours. It can be seen that the survival rate of HUVECs cells is close to 98% in the experimental range of 0-20ug of nanozyme Rh-FePc-Ti3C2Tx, proving that it has no toxicity to normal cells. It has slight toxicity to 143B cells, and the survival rate of 143B cells is about 75% at 20ug.

[0071] Figure 6 The Rh-FePc-Ti3C2T provided by the present invention x The effect diagram of killing 143B tumor cells by photodynamic therapy and sonodynamic therapy; after co-culturing the nanomaterial and 143B cells, applying near-infrared light and ultrasound conditions, the CCK-8 experiment was used to detect the ability of the nanomaterial to kill osteosarcoma cells by photodynamic and sonodynamic methods. The results showed that under the combined action of near-infrared light and ultrasound, the killing power of osteosarcoma cells can reach about 40%, and photodynamic therapy and sonodynamic therapy can effectively kill osteosarcoma cells.

[0072] Rh-FePc-Ti3C2T x Please refer to the diagram showing the ability to generate ROS in cells Figure 7 , Figure 7 The Rh-FePc-Ti3C2T provided by the present invention x The ability to generate ROS in cells is directly reflected by the DCFH-DA fluorescence probe experiment to determine the nanozyme Rh-FePc-Ti3C2T x The ability to produce ROS in cells shows that under the synergistic conditions of photoacoustic dynamics, the nanozyme Rh-FePc-Ti3C2T x The green fluorescence produced is the strongest compared to other groups, proving that it can generate ROS well and enhance oxidative stress in cells.

[0073] The above description is only the preferred embodiment of the present invention, which does not limit the protection scope of the present invention. For those skilled in the art, the present invention may have various modifications and changes. Within the spirit and principle of the present invention, the changes, modifications, replacements, integrations and parameter changes of these embodiments by conventional substitutions or capable of achieving the same functions without departing from the principles and spirit of the present invention all fall within the protection scope of the present invention.

Claims

1. A FePc-modified Ti3C2T x The use of Rh-loaded nanozymes in the preparation of drugs for treating osteosarcoma is characterized in that: The FePc-modified Ti3C2T x The main components of the Rh-loaded nanozyme are Rh-FePc-Ti3C2T x , the Rh-FePc-Ti3C2T x The raw materials for preparation include FePc, Ti3C2T x Nanosheets and RhCl3 . 3H2O; FePc, Ti3C2T x The mass ratio of nanosheets is 2~8:1~4; The FePc-modified Ti3C2T x The preparation method of the Rh-loaded nanozyme comprises the following steps: S1, FePc and Ti3C2T x Continue sonication after mixing; S2, washing and drying to obtain FePc-Ti3C2T x ; S3, in FePc-Ti3C2T x Add RhCl3 . 3H2O and co-dissolved in polyethylene glycol; S4, ultrasonic dispersion and transfer to a reaction kettle; S5, cooling, washing and centrifugation; S6, dry and grind to obtain Rh-FePc-Ti3C2T x Composite nanozymes; In the S1 step, FePc and Ti3C2T x The required material ratio is 10-40 mg FePc: 5-20 mg Ti3C2T x : 6-10ml ultrapure water; In the step S2, washing and drying are specifically performed by washing with anhydrous ethanol, centrifuging twice, and then placing in a vacuum drying oven for 12-24 hours to perform FePc-Ti3C2T x collect; In the S3 step, FePc-Ti3C2T x 、RhCl3 . 3H2O, polyethylene glycol ratio of 8-12mg FePc-Ti3C2T x : 3-6ml RhCl3 with a concentration of 2mg / mL . 3H2O: 20-40ml polyethylene glycol with a number average molecular weight of 200; The reaction time in the reactor of step S4 is 12-16h, and the reaction temperature is 160-200°C; In the step S6, the specific method of drying is to place the mixture in a vacuum drying oven for drying for 12-24 hours.

2. A FePc-modified Ti3C2T3 according to claim 1 x The use of Rh-loaded nanozymes in the preparation of drugs for treating osteosarcoma is characterized in that: In the step S1, the continuous ultrasonication condition after mixing is continuous ultrasonication for 4-8 hours at room temperature.

3. A FePc modified Ti3C2T according to claim 1 x The use of Rh-loaded nanozymes in the preparation of drugs for treating osteosarcoma is characterized in that: In the ultrasonic dispersion in step S4, the ultrasonic time is 1-2 hours until the solution is evenly dispersed.

4. A FePc modified Ti3C2T according to claim 1 x The use of Rh-loaded nanozymes in the preparation of drugs for treating osteosarcoma is characterized in that: The washing material in step S5 is anhydrous ethanol, and the number of centrifugation is three times.

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