Artificial mitochondrial nanomotor, preparation method therefor, and application

Abstract

Disclosed are an artificial mitochondrial nanomotor, a preparation method therefor, and an application. The artificial mitochondrial nanomotor comprises a monomer and a crosslinking agent, and is formed by initiating a polymerization reaction by means of an initiator. The monomer is a compound having high-density active guanidinyl functional groups and high-energy phosphate bonds, and the crosslinking agent is a compound containing disulfide bonds or diselenide bonds. The artificial mitochondrial nanomotor not only produces energy in cells, but also chemotactically targets disease microenvironments, in order to achieve an ideal disease treatment effect, and has broad application prospects in the field of biomedicine.

Description

An artificial mitochondrial nanomotor, its preparation method and application Technical Field

[0001] This invention pertains to novel biomedical nanomaterials, specifically relating to an artificial mitochondrial nanomotor, its preparation method, and its applications. Background Technology

[0002] Mitochondria play a vital role in cells, particularly in providing ATP through oxidative phosphorylation and central carbon metabolism pathways. Disruptions in their function can severely impact cellular activity, ion homeostasis, and multiple processes related to cell signaling. Impaired ATP production directly affects cellular energy supply, hindering various life activities and triggering a range of diseases, including cardiovascular diseases, neurodegenerative diseases, and metabolic diseases. Therefore, developing a method to enhance mitochondrial ATP production is crucial for treating diseases characterized primarily by mitochondrial damage.

[0003] Existing systems for enhancing mitochondrial ATP production mainly fall into two categories: direct transplantation of viable mitochondria and the construction of artificial systems capable of generating ATP. The former involves extracting fully functional mitochondria from normal cells using specialized methods and transplanting them to the damaged area. The damaged cells then take up the mitochondria and fuse with them to repair their function. This method theoretically offers the advantage of completely replacing the structure and function of damaged mitochondria and is currently the most widely used direct mitochondrial energy repair system in vivo. However, due to the fragility of mitochondrial isolation, the complexity and difficulty of purification and preservation, immunocompatibility during transplantation, and potential ethical issues, its future clinical translation will face significant challenges. As for the latter, some researchers have attempted to construct artificial systems to generate ATP, partially mimicking mitochondrial function. This strategy is highly efficient in vitro; however, most of these artificial systems require additional energy input, such as visible light irradiation, to drive proton transmembrane transport. Their limited in vivo penetration depth restricts their efficiency in generating ATP in vivo. Therefore, despite the promising prospects of artificial ATP synthesis systems in the field of disease treatment, no artificial ATP synthesis system has yet been reported that is stable, applicable in vivo, does not rely on external energy sources, and can effectively target the disease microenvironment. Summary of the Invention

[0004] Purpose of the invention: To address the problems existing in the prior art, this invention provides an artificial mitochondrial nanomotor with stable performance that can effectively target the disease microenvironment in vivo without relying on an external energy source, effectively solving the problem that the currently artificially synthesized ATP system cannot be applied to the disease microenvironment in vivo.

[0005] The present invention also provides a method for preparing an artificial mitochondrial nanomotor and its application.

[0006] Technical Solution: To achieve the above objectives, this invention discloses an artificial mitochondrial nanomotor. The artificial mitochondrial nanomotor comprises a monomer and a cross-linking agent, formed by a polymerization reaction initiated by an initiator. The monomer is a compound with high-density active guanidine functional groups and high-energy phosphate bonds, and the cross-linking agent is a compound containing disulfide or diselenyl groups. This artificial mitochondrial nanomotor not only generates energy within cells but can also chemotactically target the disease microenvironment, achieving ideal therapeutic effects and possessing broad application prospects in the biomedical field.

[0007] The monomeric compound is one or more of L-arginine creatine phosphate derivatives, L-lysine creatine phosphate derivatives, and L-cysteine ​​creatine phosphate derivatives.

[0008] The crosslinking agent is a disulfide crosslinking agent N,N'-bis(acryloyl)cystamine or a diselenyl ether compound crosslinking agent.

[0009] The initiator includes one or more of the following: ammonium persulfate-tetramethylethylenediamine initiating system, benzoin, benzoin ethyl ether and benzoin butyl ether, benzoin dimethyl ether, benzophenone, thioxanthone, camphor porone, or diimidazole.

[0010] The artificial mitochondrial nanomotor is prepared by the following steps:

[0011] (1) Dissolve creatine phosphate in a buffer solution, add N-hydroxysuccinimide and 1-ethyl-(3-dimethylaminopropyl)carbodiimide, then add L-arginine derivative or L-lysine derivative or L-cysteine ​​derivative, stir the reaction to obtain L-arginine creatine phosphate derivative, L-lysine creatine phosphate derivative or L-cysteine ​​creatine phosphate derivative;

[0012] (2) The L-arginine creatine phosphate derivative, L-lysine creatine phosphate derivative or L-cysteine ​​creatine phosphate derivative obtained in step (1) are dissolved in a buffer solution with a crosslinking agent, an initiator is added to carry out a polymerization reaction, and artificial mitochondrial nanomotors are obtained by centrifugation and washing.

[0013] In step (1), the molar ratio of creatine phosphate to N-hydroxysuccinimide is 2-50:1, the molar ratio of N-hydroxysuccinimide to 1-ethyl-(3-dimethylaminopropyl)carbodiimide is 1-50:1, the molar ratio of creatine phosphate to L-arginine derivative, L-lysine derivative, or L-cysteine ​​derivative is 1-10:1, the stirring reaction temperature is 20-30℃, and the reaction time is 10-48h.

[0014] In step (2), the molar ratio of the L-arginine creatine phosphate derivative, L-lysine creatine phosphate derivative, or L-cysteine ​​creatine phosphate derivative to the crosslinking agent is 16:1-1:1, the molar ratio of the L-arginine creatine phosphate derivative, L-lysine creatine phosphate derivative, or L-cysteine ​​creatine phosphate derivative to the initiator is 1:1-50:1, the reaction temperature is 0-30℃, and the reaction time is 1-48h.

[0015] The centrifugation washing process involves collecting nanoparticles through ultracentrifugation, washing them, and freeze-drying them to obtain artificial mitochondrial nanomotors. The ultracentrifugation speed is 5000-12000 rpm, and the centrifugation time is 1-30 min.

[0016] The application of the artificial mitochondrial nanomotor described in this invention in the preparation of drugs, reagents, or health products for generating ATP or treating energy deficiency diseases.

[0017] The energy deficiency diseases mentioned above refer to diseases caused by impaired ATP production capacity affecting cellular energy supply, including cardiovascular diseases, neurodegenerative diseases, and metabolic diseases. This invention utilizes a covalent bond between arginine derivatives and creatine phosphate to prepare a novel monomer containing both arginine and creatine phosphate, which is then constructed into an artificial mitochondrial nanomotor via free radical polymerization. The mechanism of action is that the creatine phosphate portion of the artificial mitochondrial nanomotor structure provides sufficient high-energy phosphate bonds to damaged mitochondria, reacting with ADP in the cytoplasm under enzymatic catalysis to generate ATP. Simultaneously, the arginine portion of its structure can specifically recognize inducible nitric oxide synthase (iNOS), which is highly expressed in damaged mitochondria, thereby achieving chemotactic targeting of the damaged site. The artificial mitochondrial nanomotor reaching the damaged site can not only rapidly generate large amounts of ATP to reduce the burden on damaged mitochondria and restore cellular activity, but also react with reactive oxygen species in the damaged microenvironment to produce NO, effectively alleviating inflammation; the generated NO can also promote mitochondriogenesis, further repairing mitochondrial function.

[0018] Beneficial effects: Compared with the prior art, the present invention has the following advantages.

[0019] The artificial mitochondrial nanomotor of this invention is primarily composed of polymeric segments rich in active guanidine groups and high-energy phosphate bonds. This covalent bonding ensures a sufficiently high density of active groups, enabling it to sensitively respond to the high iNOS concentration gradient in the disease microenvironment and achieve chemotactic targeting of damaged tissues. This overcomes the limitation of currently reported artificial ATP-generating systems being difficult to apply in vivo, and also provides a solution to the problem of artificial mitochondria being unable to target damaged tissues in vivo. Attached Figure Description

[0020] Figure 1 shows the structural formula of the monomer containing arginine and phosphocreatine in Example 1;

[0021] Figure 2 shows the monomers containing arginine and creatine phosphate in Example 1. 1 HNMR spectrum;

[0022] Figure 3 shows a transmission electron microscope image of the artificial mitochondrial nanomotor in Example 1 (scale bar: 500 nm);

[0023] Figure 4 shows the particle size distribution of the artificial mitochondrial nanomotor in Example 2;

[0024] Figure 5 shows the motion trajectory of the artificial mitochondrial nanomotor in Example 3 under a cell injury-stimulated environment.

[0025] Figure 6 shows the velocity distribution of the artificial mitochondrial nanomotor in Example 3 under a damaged and stimulated cell environment.

[0026] Figure 7 shows the motion trajectory of the artificial mitochondrial nanomotor in a normal cellular environment in Example 4.

[0027] Figure 8 shows the velocity distribution of the artificial mitochondrial nanomotor in a normal cellular environment in Example 4.

[0028] Figure 9 shows a representative image of the chemotactic behavior of the artificial mitochondrial nanomotors towards the damaged cellular environment in Example 5.

[0029] Figure 10 shows the fluorescence quantitative normalization results of representative images of the chemotactic behavior of the artificial mitochondrial nanomotors towards the damaged cellular environment in Example 5.

[0030] Figure 11 shows the cell viability of different concentrations of artificial mitochondrial nanomotors in Example 6 after incubation with normal cells for 24 hours to evaluate the biocompatibility of artificial mitochondrial nanomotors for the treatment of ischemic diseases.

[0031] Figure 12 shows the ATP production in the cellular environment stimulated by damage of the artificial mitochondrial nanomotor in Example 7. Detailed Implementation

[0032] Unless otherwise specified, the experimental methods described in the embodiments are conventional methods; unless otherwise specified, the reagents and materials are commercially available.

[0033] The structure of the L-arginine derivative N-methacryloylarginine is as follows:

[0034] For details of the synthesis method, please refer to the literature: A nitric-oxide driven chemotactic nanomotor for enhanced immunotherapy of glioblastoma. Nature Communications, 2023, 14, 941.

[0035] Example 1

[0036] Preparation of artificial mitochondrial nanomotors:

[0037] (1) Creatine phosphate (1.0 g, 4.0 mmol) was dissolved in 20 mL of PBS solution (pH = 7.4), and N-hydroxysuccinimide (0.8 g, 4.0 mmol) and 1-ethyl-(3-dimethylaminopropyl)carbodiimide (0.5 g, 4.0 mmol) were added. Then, N-methacryloylarginine (0.9 g, 4.0 mmol) was added, and the reaction was stirred at 25 °C for 24 h to obtain a monomer containing arginine and phosphocreatine (Figure 1); as shown in Figure 2. 1 HNMR spectra showed that monomers containing arginine and phosphocreatine were successfully prepared.

[0038] (2) The monomer containing arginine and phosphocreatine, namely L-arginine creatine phosphate derivative (140 mg, 0.3 mmol), obtained in step (1), was dissolved in 10 mL of PBS (pH = 7.4) with the GSH-responsive disulfide compound BAC (N,N'-bis(acryloyl)cysteine, 10 mg, 0.04 mmol). An ammonium persulfate-tetramethylethylenediamine initiator system (molar ratio of L-arginine creatine phosphate derivative to ammonium persulfate was 50:1, and molar ratio of ammonium persulfate to tetramethylethylenediamine was 2:1) was added for polymerization (25 °C, 6 h). After centrifugation and washing at 10,000 rpm for 10 min, the supernatant was discarded to obtain the artificial mitochondrial nanomotor (lower precipitate). As shown in Figure 3, the synthesized artificial mitochondrial nanomotor has a particle size of approximately 110 nm and exhibits uniformly dispersed and irregularly shaped spherical nanoparticles.

[0039] Example 2

[0040] DLS particle size determination of artificial mitochondrial nanomotor solution:

[0041] 4 mg of the artificial mitochondrial nanomotor solution prepared in Example 1 was weighed and dispersed in 4 mL of PBS solution to obtain a concentration of 1 mg / mL. 1 mL of this solution was added to a cuvette and placed in a laser particle size analyzer for testing. As shown in Figure 4, the DLS particle size of the artificial mitochondrial nanomotor was approximately 105 nm.

[0042] Example 3

[0043] Study on the motility of artificial mitochondrial nanomotors in a damaged cellular environment:

[0044] (1) Weigh 4 mg of the artificial mitochondrial nanomotor prepared in Example 1, add 4 mL of deionized water, and sonicate for 30 min to disperse it fully. Then add 4 mg of N-hydroxysuccinimide and 8 mg of 1-ethyl-(3-dimethylaminopropyl)carbodiimide. After reacting at room temperature for 3 h, add 20 μL of 1 mg / mL Cy5-NH2 DMSO dye solution and react in the dark for 12 h to obtain Cy5-labeled artificial mitochondrial nanomotor.

[0045] (2) Cardiac cells (H9c2) were prepared at a concentration of 5*10 5 Cells were seeded at a density of 1 / mL in 14mm cell culture dishes with 1mL of complete culture medium and placed in a 37℃ incubator overnight to allow the cells to adhere.

[0046] (3) Take 1 μL of 1 mg / mL lipopolysaccharide DMSO solution, add it to the above culture dish, and place it in a constant temperature incubator at 37℃ for 24 h to stimulate inflammation;

[0047] (3) Take 10 μL of artificial mitochondrial nanomotors loaded with Cy5-NH2 dye at 200 μg / mL and add them to the above adherent cell culture dish. Immediately use a fluorescence microscope to observe and record the movement of the nanomotors.

[0048] (4) Figure 5 shows the trajectory of the artificial mitochondrial nanomotor in damaged cells, demonstrating that the artificial mitochondrial nanomotor can move in the environment of damaged cells. Its velocity distribution was calculated based on the trajectory, as shown in Figure 6. This proves that the artificial mitochondrial nanomotor exhibits significant movement behavior in the damaged environment. The artificial mitochondrial nanomotor prepared in this invention gains movement ability under iNOS catalysis in damaged cell environments with high iNOS content, exhibiting stronger cell permeability and a greater therapeutic effect.

[0049] Example 4

[0050] Study on the motility of artificial mitochondrial nanomotors in a normal cellular environment:

[0051] (1) Weigh 4 mg of the artificial mitochondrial nanomotor prepared in Example 1, add 4 mL of deionized water, and sonicate for 30 min to disperse it fully. Then add 4 mg of N-hydroxysuccinimide and 8 mg of 1-ethyl-(3-dimethylaminopropyl)carbodiimide. After reacting at room temperature for 3 h, add 20 μL of 1 mg / mL Cy5-NH2 DMSO dye solution and react in the dark for 12 h to obtain Cy5-labeled artificial mitochondrial nanomotor.

[0052] (2) H9c2 cells were fed at a concentration of 5*10 5 Cells were seeded at a density of 1 / mL in 14mm cell culture dishes with 1mL of complete culture medium and placed in a 37℃ incubator overnight to allow the cells to adhere.

[0053] (3) Take 10 μL of artificial mitochondrial nanomotor solution loaded with Cy5-NH2 dye and add it to the above adherent cell culture dish. Immediately use a fluorescence microscope to observe and record the movement of the nanomotor.

[0054] (4) Figure 7 shows the trajectory of the artificial mitochondrial nanomotor in normal cells, demonstrating that the artificial mitochondrial nanomotor has no autonomous movement capability in a normal cellular environment. Its velocity distribution was calculated based on the trajectory, as shown in Figure 8. This proves that the artificial mitochondrial nanomotor moves in a random Brownian motion within a normal cellular environment. The nanorobot prepared in this invention does not exhibit autonomous movement in normal cells with normal iNOS levels, but possesses autonomous movement capability in damaged cells.

[0055] Example 5

[0056] Assessment of the chemotactic behavior of artificial mitochondrial nanomotors toward the damaged cellular environment:

[0057] (1) Weigh 4 mg of the artificial mitochondrial nanomotor prepared in Example 1, add 4 mL of deionized water, and sonicate for 30 min to disperse it fully. Then add 4 mg of N-hydroxysuccinimide and 8 mg of 1-ethyl-(3-dimethylaminopropyl)carbodiimide. After reacting at room temperature for 3 h, add 20 μL of 1 mg / mL Cy5-NH2 DMSO dye solution and react in the dark for 12 h to obtain Cy5-labeled artificial mitochondrial nanomotor.

[0058] (2) H9c2 cells were fed at a concentration of 5*10 5 Cells were seeded at a density of 1 / mL in 14mm cell culture dishes with 1mL of complete culture medium and placed in a 37℃ incubator overnight to allow the cells to adhere.

[0059] (3) Take 1 μL of 1 mg / mL lipopolysaccharide DMSO solution, add it to the above culture dish, and place it in a constant temperature incubator at 37℃ for 24 h to stimulate inflammatory damage. Then, use 200 μL of lysis buffer to lyse the cells to obtain lipopolysaccharide pre-stimulated H9c2 cell lysate.

[0060] (3) To observe the collective chemotactic behavior of artificial mitochondrial nanomotors for target cell identification, Y-shaped glass substrate microchannels were used. The main channel was 1 cm long and 0.4 cm wide, and the branch channels were 0.7 cm long and 0.3 cm wide. The chemotactic concentration gradient was generated by different types of cell lysates placed in reservoirs (ii) or (iii) within the branch channels. Briefly, 5 mg of agarose was completely dissolved in 500 μL of PBS at 90 °C. When the melted agarose cooled to room temperature but did not solidify, 50 μL of lipopolysaccharide-prestimulated H9c2 cell lysate was added, and the mixture was then transferred to reservoir (ii) at 4 °C for gelation, or 50 μL of positive PBS was added, and the mixture was then transferred to reservoir (iii) at 4 °C for gelation. Before evaluating the chemotactic movement of the artificial mitochondrial nanomotors, the Y-shaped channels were pre-filled with PBS, and 50 μL of Cy5-labeled artificial mitochondrial nanomotor solution was gently dropped into the reservoir (i). At specific times, fluorescence images of reservoirs (ii) and (iii) were captured using an inverted fluorescence microscope equipped with a 10× objective, as shown in Figure 9. As shown in Figure 10, the corresponding fluorescence intensities were quantified using ImageJ. It was observed that the fluorescence intensity of the reservoir containing pre-stimulated H9C2 cell lysate was significantly higher than that containing PBS, indicating that the artificial mitochondrial nanomotors exhibit chemotactic behavior towards the iNOS concentration gradient. This demonstrates that the artificial mitochondrial nanomotors can chemotactically attract environments containing iNOS concentration gradients, enabling precise targeting of damaged sites for treatment.

[0061] Example 6

[0062] Biocompatibility assessment of artificial mitochondrial nanomotors:

[0063] H9c2 cells were used at a rate of 5*10 4 The artificial mitochondrial nanomotors were seeded at a concentration of 1 / mL in 96-well plates. After the cells adhered, the artificial mitochondrial nanomotors prepared in Example 1 were dispersed in the culture medium. Different concentrations of artificial mitochondrial nanomotors (20, 50, 100, 200, 400 μg / mL) were prepared and incubated with cardiomyocytes at 37°C for 24 h. Cell viability was then detected using the MTT assay. As shown in Figure 11, the cardiomyocyte viability was hardly affected by the increase in material concentration, thus demonstrating that the artificial mitochondrial nanomotors have good biocompatibility.

[0064] Example 7

[0065] The production of ATP in the cellular environment stimulated by damage to artificial mitochondrial nanomotors:

[0066] (1) H9c2 cells were fed at a concentration of 5*10 5 Cells were seeded at a concentration of 1 / mL in 14 mm cell culture dishes with 1 mL of complete culture medium and incubated overnight at 37°C. The culture medium was then replaced with sugar-free DMEM and the dishes were incubated in a hypoxic incubator for 24 h. Artificial mitochondrial nanomotors prepared in Example 1 were added to the sugar-free DMEM at a concentration of 200 μg / mL.

[0067] (2) The ATP production of H9c2 cells was detected according to the requirements of the ATP detection kit (Beyotime, Cat No. S0026), as shown in Figure 12. The ability of cells to produce ATP after injury stimulation was significantly reduced, while the ability of cells to produce ATP after injury stimulation was enhanced after the addition of artificial mitochondrial nanomotors, recovering to or even exceeding the normal level, indicating that artificial mitochondrial nanomotors have a good ability to produce ATP.

[0068] Example 8

[0069] The artificial mitochondrial nanomotor was prepared using the method described in Example 1, with the following differences:

[0070] In step (1), creatine (1.0 g, 7.6 mmol) was dissolved in 30 mL of PBS solution, and N-hydroxysuccinimide (1.6 g, 7.6 mmol) and 1-ethyl-(3-dimethylaminopropyl)carbodiimide (1.0 g, 7.6 mmol) were added. Then, L-arginine derivative N-methacryloylarginine (1.7 g, 7.6 mmol) was added, and the mixture was stirred. The reaction system was 25 °C and the reaction time was 24 h to obtain L-arginine creatine derivative. Subsequently, the reaction was carried out according to step (2) of Example 1 to obtain artificial mitochondrial nanomotor control sample 1, so as to study the difference between artificial mitochondrial nanomotor loaded with creatine phosphate as substrate and loaded with creatine as substrate.

[0071] Example 9

[0072] The artificial mitochondrial nanomotor was prepared using the method described in Example 1, with the following differences:

[0073] In step (1), creatine phosphate (10 g, 40 mmol) was dissolved in 30 mL of PBS solution, and N-hydroxysuccinimide (8.0 g, 40 mmol) and 1-ethyl-(3-dimethylaminopropyl)carbodiimide (5.0 g, 40 mmol) were added. Then, the L-arginine derivative N-methacryloylarginine (0.9 g, 4.0 mmol) was added, and the mixture was stirred. The reaction system was 25 °C and the reaction time was 24 h to obtain the L-arginine creatine derivative. Subsequently, the reaction was carried out according to step (2) of Example 1 to obtain artificial mitochondrial nanomotor control sample 2, so as to study the effect of the amount of creatine phosphate loaded on the artificial mitochondrial nanomotor on its performance.

[0074] Example 10

[0075] The artificial mitochondrial nanomotor was prepared using the method described in Example 1, with the following differences:

[0076] In step (2), the L-arginine creatine derivative and the GSH-responsive disulfide compound BAC were dissolved in 10 mL of PBS solvent at a molar ratio of 1. An ammonium persulfate-tetramethylethylenediamine initiator system (the molar ratio of L-arginine creatine derivative to ammonium persulfate was 50, and the molar ratio of ammonium persulfate to tetramethylethylenediamine was 2) was added to carry out the polymerization reaction. After centrifugation and washing at 10,000 rpm for 10 min, artificial mitochondrial nanomotor control sample 3 was obtained to study the effect of monomer dosage on artificial mitochondrial nanomotor generated by free radical polymerization.

[0077] Example 11

[0078] The artificial mitochondrial nanomotor was prepared using the method described in Example 1, with the following differences:

[0079] In step (2), the L-arginine creatine derivative and the GSH-responsive disulfide compound BAC were dissolved in 10 mL of PBS solvent at a molar ratio of 8. An initiator system of ammonium persulfate-tetramethylethylenediamine (the molar ratio of L-arginine creatine derivative to ammonium persulfate was 10, and the molar ratio of ammonium persulfate to tetramethylethylenediamine was 2) was added to carry out the polymerization reaction. The mixture was centrifuged and washed at 10,000 rpm for 10 min to obtain control sample 4 of the artificial mitochondrial nanomotor, so as to study the effect of the amount of initiator on the artificial mitochondrial nanomotor generated by free radical polymerization.

Claims

1. An artificial mitochondria nanomotor, characterized in that, The artificial mitochondrial nanomotor comprises a monomer and a crosslinking agent, which are formed by a polymerization reaction initiated by an initiator. The monomer is a compound with high-density active guanidine functional groups and high-energy phosphate bonds, and the crosslinking agent is a compound containing disulfide bonds or diselenide bonds.

2. The artificial mitochondrial nanomotor of claim 1, wherein, The monomeric compound is one or more of L-arginine creatine phosphate derivatives, L-lysine creatine phosphate derivatives, and L-cysteine ​​creatine phosphate derivatives.

3. The artificial mitochondrial nanomotor of claim 1, wherein, The crosslinking agent is preferably a disulfide crosslinking agent N,N'-bis(acryloyl)cystamine or a diselenyl ether compound crosslinking agent.

4. The artificial mitochondrial nanomotor of claim 1, wherein, The initiator includes one or more of the following: ammonium persulfate-tetramethylethylenediamine initiating system, benzoin, benzoin ethyl ether and benzoin butyl ether, benzoin dimethyl ether, benzophenone, thioxanthone, camphor porone, or diimidazole.

5. A method of preparing the artificial mitochondria nanomotor of claim 1, wherein, Includes the following steps: (1) Dissolve creatine phosphate in a buffer solution, add N-hydroxysuccinimide and 1-ethyl-(3-dimethylaminopropyl)carbodiimide, then add L-arginine derivative or L-lysine derivative or L-cysteine ​​derivative, stir the reaction to obtain L-arginine creatine phosphate derivative, L-lysine creatine phosphate derivative or L-cysteine ​​creatine phosphate derivative; (2) The L-arginine creatine phosphate derivative, L-lysine creatine phosphate derivative or L-cysteine ​​creatine phosphate derivative obtained in step (1) are dissolved in a buffer solution with a crosslinking agent, an initiator is added to carry out a polymerization reaction, and artificial mitochondrial nanomotors are obtained by centrifugation and washing.

6. The production method according to claim 5, wherein In step (1), the molar ratio of creatine phosphate to N-hydroxysuccinimide is 2-50:1, the molar ratio of N-hydroxysuccinimide to 1-ethyl-(3-dimethylaminopropyl)carbodiimide is 1-50:1, the molar ratio of creatine phosphate to L-arginine derivative, L-lysine derivative, or L-cysteine ​​derivative is 1-10:1, the stirring reaction temperature is 20-30℃, and the reaction time is 10-48h.

7. The preparation method according to claim 5, characterized in that, In step (2), the molar ratio of the L-arginine creatine phosphate derivative, L-lysine creatine phosphate derivative, or L-cysteine ​​creatine phosphate derivative to the crosslinking agent is 16:1:-1:1, the molar ratio of the L-arginine creatine phosphate derivative, L-lysine creatine phosphate derivative, or L-cysteine ​​creatine phosphate derivative to the initiator is 1:1-50:1, the reaction temperature is 0-30℃, and the reaction time is 1-48h.

8. The preparation method according to claim 5, characterized in that, The centrifugation washing in step (2) involves collecting nanoparticles through ultracentrifugation, washing and freeze-drying them to obtain artificial mitochondrial nanomotors. The ultracentrifugation speed is 5000-12000 rpm and the centrifugation time is 1-30 min.

9. The use of the artificial mitochondrial nanomotor of claim 1 in the preparation of drugs, reagents or health products for generating ATP or treating energy deficiency diseases.

10. Use according to claim 9, characterized in that, The energy deficiency diseases mentioned above are diseases caused by damage to ATP production capacity that affects the energy supply of cells, including cardiovascular diseases, neurodegenerative diseases, and metabolic diseases.

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