Preparation method of engine-detachable braking nano motor

Abstract

The invention belongs to the technical field of functional nano materials, and particularly relates to a preparation method of a detachable engine type braking nano motor, a nano drug carrier is jointly composed of HMSN and CAT modified PNA, tetraethyl orthosilicate serves as a silicon source, and solid silicon dioxide balls are synthesized; on the basis, tetraethyl orthosilicate is used as a silicon source, CTAB is used as a pore-forming agent to synthesize mesoporous silica coated solid silica spheres; etching the core silicon dioxide ball through sodium carbonate to obtain a hollow mesoporous silicon dioxide ball; then, amino modification is carried out on the CAT modified PNA2 through 3-aminopropyltrimethoxysilane, PNA1 is connected with HMSN through an amido bond, and a double-chain structure is formed by the CAT modified PNA2 and PNA1 through complementary base pairing. According to the nano motor provided by the invention, the engine CAT of the nano motor can be detached by adding the unwinding chain PNA3, so that in-vivo deceleration and braking are realized.

Description

Technical Field

[0001] This invention belongs to the field of functional nanomaterials technology, specifically relating to a method for preparing a detachable, brakeable nanomotor. Background Technology

[0002] The development of nanotechnology has brought new opportunities to the biomedical field. Nanomaterials, due to their size effect, surface effect, and designability, show broad application prospects in drug delivery, molecular imaging, and integrated disease diagnosis and treatment. In recent years, nanomotors, as functional nanostructures capable of autonomous or directional movement at the micro- and nanoscale, have attracted widespread attention. By achieving active movement and penetration in complex biological environments, they can effectively overcome the insufficient penetration problem faced by traditional nanomedicine carriers that rely on passive diffusion, and are therefore considered an important direction for promoting precision medicine and novel intelligent healthcare.

[0003] Existing nanomotors primarily rely on various motion mechanisms for propulsion, such as chemical fuel-driven, light-driven, ultrasound-driven, and magnetic field-driven mechanisms. The controllability of motion speed is a crucial factor in nanomotor applications; excessively fast or slow motion can affect delivery efficiency and therapeutic efficacy within the target environment. Therefore, nanomotors capable of "braking" or stopping are of significant importance. Currently, most nanomotors capable of braking rely on the removal of the external energy field, exhaustion of fuel, or the application of a counterforce. However, these strategies still have significant limitations. For example, in the complex environment of the body, the penetration depth of the external energy field is limited, making effective control of deep tissues difficult; the fuel supply cannot be precisely adjusted, easily leading to uncontrollable motion; and the magnitude and range of the counterforce are also difficult to control. These problems severely restrict the clinical translational application prospects of nanomotors.

[0004] Therefore, there is an urgent need to develop a novel nanomotor design that can achieve braking through internal field control. Summary of the Invention

[0005] The purpose of this invention is to provide a method for preparing a detachable, brakeable nanomotor (HMSN-PNA1-PNA2-CAT) to address the problem of rapid metabolism of nanomotors in vivo due to their rapid motility. A double-helix structure is formed by the self-assembly of complementary PNA1 and CAT-modified PNA2, creating a CAT-driven nanomotor. CAT catalyzes the decomposition of H2O2 to generate O2, providing the driving force for the nanomotor. Once the nanomotor reaches a designated location in the body, the CAT-PNA2 on the nanomotor is replaced by injecting unhelicated PNA3, thus achieving engine detachment. This prolongs the residence time at the specific site and improves the efficiency of the nanomotor.

[0006] To achieve the above-mentioned technical objectives and effects, the present invention is implemented through the following technical solution:

[0007] This invention provides a method for preparing a detachable, brakeable nanomotor, comprising the following steps:

[0008] 1) Preparation of SN: Solid silica was prepared by sol-gel method. TEOS was rapidly added to a mixture of deionized water, anhydrous ethanol and ammonia, and the mixture was stirred at room temperature to form SN suspension. Finally, the product was centrifuged, washed and the SN particles were dispersed in deionized water.

[0009] 2) Preparation of HMSN: CTAB was added to a mixture of deionized water, anhydrous ethanol and ammonia and stirred until homogeneous; then, SN particles dispersed in deionized water were sonicated and added to the mixture, and stirring was continued; TEOS was then slowly added dropwise and stirred at a constant speed; after the reaction was completed, the mixture was centrifuged and washed with water to obtain the intermediate product mesoporous silica-CTAB, denoted as MSN-CTAB.

[0010] The core SN was removed by etching: MSN-CTAB prepared by reaction was ultrasonically dispersed in deionized water, and then anhydrous sodium carbonate was added to the dispersion. The mixture was reacted at 40-60℃. After the reaction was completed, the white product was collected by centrifugation and washed several times with ethanol to obtain hollow mesoporous silica-CTAB, denoted as HMSN-CTAB.

[0011] The obtained product was extracted with a mixture of ethanol and hydrochloric acid at 75–85 °C to remove the template CTAB; the product was collected by centrifugation and washed with alcohol to obtain the product HMSN.

[0012] 3) Preparation of HMSN-NH2: HMSN was dispersed in ethanol, and 3-aminopropyltriethoxysilane was slowly added dropwise to the HMSN solution while stirring at room temperature; the product HMSN-NH2 was collected by centrifugation and washed with deionized water, and then dispersed in deionized water;

[0013] 4) Preparation of HMSN-PNA1-PNA2-CAT: Deionized water was added to a tube of PNA1 and dispersed by sonication. 1-Ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride and N-hydroxysuccinimide were added and stirred to activate the mixture. HMSN-NH2 was taken, sonicated, and added to the above solution. The mixture was stirred overnight at room temperature. After centrifugation and washing with water, the supernatant was retained, and the precipitate was dispersed with deionized water to obtain HMSN-RNA1. Tris-HCl was added to a tube of PNA2 and dispersed by sonication. Add HMSN-RNA1 and stir overnight; centrifuge and wash with water, retain the supernatant, and disperse the precipitate with deionized water to obtain HMSN-PNA1-PNA2-NH2; dissolve catalase in a mixed solution containing deionized water, EDC·HCl and NHS to activate it, stir at room temperature, and then mix the above solution with the HMSN-PNA1-PNA2-NH2 solution and stir overnight; finally centrifuge and wash with water several times, disperse in deionized water to obtain HMSN-PNA1-PNA2-CAT.

[0014] Further, in step 1), the mass ratio of ethanol, deionized water and ammonia solution in the mixture is 25-50:5:1-4, and the concentration of ammonia solution is 25-28%; the mass ratio of the mixture and TEOS is 31-59:3.

[0015] Further, in step 2), the mass ratio of deionized water, anhydrous ethanol and ammonia in the mixture is 600:600:11, and the concentration of the ammonia solution is 25-28%; the mass ratio of the mixture, SN particles and TEOS is 3633:6000:10-30.

[0016] Further, in step 2), the mass ratio of MSN-CTAB to anhydrous sodium carbonate is 1000:2-5, and the mass ratio of ethanol to hydrochloric acid in the mixture is 100:9.

[0017] Further, in step 3), the mass ratio of HMSN to 3-aminopropyltriethoxysilane is 1:12.

[0018] Further, in step 4), the mass ratio of 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride to N-hydroxysuccinimide is 40 to 100:69.

[0019] Furthermore, in step 4), the molar ratio of PNA1 to PNA2 is 1:1.

[0020] The sequence of PNA1 is 5'-HOOC-O-linker-TGTTGACGAGCG-3';

[0021] The sequence of PNA2 is 5'-CAT-O-linker-CGTCAACA-3';

[0022] The sequence of PNA3 is 5'-O-linker-CGCTCGTCDDCD-3'.

[0023] Further, in step 4), the mass ratio of deionized water, EDC·HCl and NHS in the mixture is 100:379:345.

[0024] The present invention also provides an engine-detachable brakeable nanomotor, which is prepared by the preparation method described in any one of claims 1-9.

[0025] The beneficial effects of this invention are:

[0026] 1. The nanomotor synthesized in this invention is powered by complementary PNA coupling enzymes.

[0027] 2. The nanomotor synthesized in this invention can achieve motion braking by internal field regulation in biological bodies, thereby prolonging the dwell time at specific locations.

[0028] Of course, any product implementing this invention does not necessarily need to achieve all of the above advantages at the same time. Attached Figure Description

[0029] To more clearly illustrate the technical solutions of the embodiments of the present invention, the accompanying drawings used in the description of the embodiments will be briefly introduced below. Obviously, the drawings described below are only some embodiments of the present invention. For those skilled in the art, other drawings can be obtained based on these drawings without creative effort.

[0030] Figure 1 TEM image of HMSN-PNA1-PNA2-CAT prepared in Example 1;

[0031] Figure 2 The flowchart shows the preparation process of HMSN-PNA1-PNA2-CAT.

[0032] Figure 3 Particle size distribution diagrams of HMSN, HMSN-PNA1-PNA2-CAT and HMSN-PNA1-PNA3;

[0033] Figure 4 Optical microscopic images of HMSN-PNA1-PNA2-CAT catalyzing the production of O2 from H2O2 and schematic diagram of the successful CAT modification evaluation by BCA method;

[0034] Figure 5 Trajectory diagrams of Brownian motion and self-propulsion motion for HMSN-PNA1-PNA2 and HMSN-PNA1-PNA2-CAT;

[0035] Figure 6 Cell viability of 4T1 and MCF-7 cells after 48 hours of incubation with HMSN-PNA1-PNA2-CAT.

[0036] Figure 7 Cell viability of 4T1 cells after incubation with HMSN-PNA1-PNA2-CAT and 0.01% H2O2 for 48 hours. Detailed Implementation

[0037] The technical solutions of the embodiments of the present invention will be clearly and completely described below with reference to the accompanying drawings. Obviously, the described embodiments are only some embodiments of the present invention, and not all embodiments. Based on the embodiments of the present invention, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of the present invention.

[0038] This invention provides a method for preparing a detachable, brakeable nanomotor (HMSN-PNA1-PNA2-CAT), the preparation process of which is as follows: Figure 2 As shown, the specific experimental steps are as follows:

[0039] 1) Preparation of SN: Solid silica was prepared using the sol-gel method. 6 mL of TEOS was rapidly added to a mixture of ethanol (50-100 mL), deionized water (10 mL), and ammonia solution (2-8 mL). The mixture was then stirred at room temperature for 0.5 h to form a solid silica sphere (SN) suspension. Finally, the product was centrifuged at 15000 r / min for 10 min, washed three times with ethanol and deionized water, and dispersed in 50 mL of water.

[0040] 2) Preparation of HMSN: Add 300-500 mg of CTAB to a mixture of 90 mL deionized water, 90 mL anhydrous ethanol, and 1.65 mL ammonia, and stir at 30°C for 2 h. Then, add 300 mg of SN particles dispersed in 20 mL deionized water, sonicated for 30 min, to the mixture, and continue stirring for 30 min. Then, slowly add 0.5-1.5 mL of TEOS, and stir uniformly at 30°C for 6 h. After the reaction, centrifuge and wash three times with water to obtain the intermediate product mesoporous silica-CTAB (MSN-CTAB). Remove the core SN by etching: Take 400 mg of MSN-CTAB prepared in the previous step, sonicate it into 60 mL of deionized water, and then add 0.8-2.0 g (0.2 M) of anhydrous sodium carbonate (Na₂CO₃) to the dispersion. React the mixture at 50°C for 6 h. After the reaction, collect the white product by centrifugation. Hollow mesoporous silica-CTAB (HMSN-CTAB) was obtained by washing three times with ethanol. The resulting product was extracted in a mixture of 100 mL ethanol and 9 mL hydrochloric acid at 80 °C for 24 h to remove the template CTAB. The product was collected by centrifugation and washed three times with ethanol to obtain the product HMSN.

[0041] 3) Preparation of HMSN-NH2: HMSN was dispersed in ethanol to a concentration of 1 mg·mL. -1 120 μL of 3-aminopropyltriethoxysilane (APTES) was slowly added dropwise to 10 mL of HMSN solution with stirring at room temperature. After 12 h, the product HMSN-NH2 was collected by centrifugation and washed three times with deionized water, and then dispersed in 10 mL of water.

[0042] 4) Preparation of HMSN-PNA1-PNA2-CAT: Add 300 μL of deionized water to a tube of PNA1 (10-50 nmol), and sonicate to disperse. Take 100 μL of this solution, add 2-5 mg of 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride (EDC·HCl) and 3.45 mg of N-hydroxysuccinimide (NHS), and stir to activate for 30 min. Take 300 μL of HMSN-NH2 (0.5 mg / mL), sonicate for 30 min, and add it to the above solution. Stir overnight at room temperature. Centrifuge and wash once with water, retaining the supernatant. Disperse the precipitate with 150 μL of deionized water to obtain HMSN-RNA1 (1 mg / mL). Add 300 μL of Tris-HCl (pH=8, 50 mM, 50 mM NaCl) to a tube of PNA2 (25 nmol), disperse by sonication, and take 100 μL to add HMSN-RNA1 (1 mg / mL). Stir overnight. Centrifuge and wash once with water, retain the supernatant, and disperse the precipitate with deionized water to obtain HMSN-PNA1-PNA2-NH2 (2 mg / mL). Activate 5 mg of catalase (CAT) in a mixed solution containing 1 mL of water, 3.79 mg of EDC·HCl, and 3.45 mg of NHS. Stir at room temperature for 30 min, then mix the above solution with the HMSN-PNA1-PNA2-NH2 solution and stir overnight. Finally, centrifuge and wash three times with water, disperse in 150 μL of deionized water to obtain HMSN-PNA1-PNA2-CAT (1 mg / mL).

[0043] Specific embodiments of the present invention are as follows:

[0044] Example 1

[0045] A method for fabricating a detachable, brakeable nanomotor includes the following steps:

[0046] 1) Preparation of SN: Solid silica was prepared using the sol-gel method. 6 mL of TEOS was rapidly added to a mixture of ethanol (74 mL), deionized water (10 mL), and ammonia solution (3.2 mL). The mixture was then stirred at room temperature for 0.5 h to form a solid silica sphere (SN) suspension. Finally, the product was centrifuged at 15000 r / min for 10 min, washed three times with ethanol and deionized water, and dispersed in 50 mL of water.

[0047] 2) Preparation of HMSN: 450 mg of CTAB was added to a mixture of 90 mL deionized water, 90 mL anhydrous ethanol, and 1.65 mL ammonia, and stirred at 30 °C for 2 h. Then, 300 mg of SN particles dispersed in 20 mL deionized water were sonicated for 30 min and added to the mixture, and stirring was continued for another 30 min. Next, 0.75 mL of TEOS was slowly added dropwise, and the mixture was stirred uniformly at 30 °C for 6 h. After the reaction, the mixture was centrifuged and washed three times with water to obtain the intermediate product mesoporous silica-CTAB (MSN-CTAB). The core SN was removed by etching: 400 mg of MSN-CTAB prepared in the previous step was sonicated and dispersed in 60 mL deionized water. Then, 1.44 g (0.2 M) of anhydrous sodium carbonate (Na₂CO₃) was added to the dispersion, and the mixture was reacted at 50 °C for 6 h. After the reaction, the white product was collected by centrifugation. After washing three times with ethanol, hollow mesoporous silica-CTAB (HMSN-CTAB) was obtained. The obtained product was extracted in a mixture of 100 mL ethanol and 9 mL hydrochloric acid at 80 °C for 24 h, and the template CTAB was removed. The product was collected by centrifugation and washed three times with alcohol to obtain the product HMSN.

[0048] 3) Preparation of HMSN-NH2: HMSN was dispersed in ethanol to a concentration of 1 mg·mL. -1 120 μL of 3-aminopropyltriethoxysilane (APTES) was slowly added dropwise to 10 mL of HMSN solution with stirring at room temperature. After 12 h, the product HMSN-NH2 was collected by centrifugation and washed three times with deionized water, and then dispersed in 10 mL of water.

[0049] 4) Preparation of HMSN-PNA1-PNA2-CAT: Add 300 μL of deionized water to a tube of PNA1 (25 nmol), and sonicate to disperse. Take 100 μL of this solution, add 3.79 mg of 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride (EDC·HCl) and 3.45 mg of N-hydroxysuccinimide (NHS), and stir to activate for 30 min. Take 300 μL of HMSN-NH2 (0.5 mg / mL), sonicate for 30 min, and add it to the above solution. Stir overnight at room temperature. Centrifuge and wash once with water, retaining the supernatant. Disperse the precipitate with 150 μL of deionized water to obtain HMSN-RNA1 (1 mg / mL). Add 300 μL Tris-HCl (pH=8, 50 mM, 50 mM NaCl) to a tube of PNA2 (25 nmol), disperse by sonication, and take 100 μL. Add HMSN-RNA1 (1 mg / mL). Stir overnight. Centrifuge and wash once with water, retain the supernatant, and disperse the precipitate with deionized water to obtain HMSN-PNA1-PNA2-NH2 (2 mg / mL). Activate 5 mg catalase (CAT) in a mixed solution containing 1 mL water, 3.79 mg EDC·HCl, and 3.45 mg NHS. Stir at room temperature for 30 min, then mix the above solution with the HMSN-PNA1-PNA2-NH2 solution and stir overnight. Finally, centrifuge and wash three times with water, disperse in 150 μL of deionized water to obtain HMSN-PNA1-PNA2-CAT (1 mg / mL).

[0050] 5) Engine enzyme disassembly of HMSN-PNA1-PNA2-CAT: Add 300 μL Tris-HCl (pH=8, 50 mM, 50 mM NaCl) to a tube of PNA3 (25 nmol), and disperse by sonication. Take 100 μL, add 150 μg HMSN-PNA1-PNA2-CAT, disperse by sonication, and shake overnight at 37℃. After the reaction, wash once with deionized water, retain the supernatant, and disperse the precipitate by sonication with deionized water to obtain HMSN-PNA1-PNA3 (1 mg / mL). Transmission electron microscopy (TEM) images of the prepared material are shown below. Figure 1 .

[0051] 6) For example Figure 3 The figure shows the particle size distribution of hollow mesoporous silica (HMSN) and HMSN-PNA1-PNA2-CAT. It can be seen that the size of the nanomotor increased after modification with APTES and PNA1, PNA2, and catalase. However, the particle size decreased after the addition of PNA3.

[0052] like Figure 4 As shown, when HMSN-PNA1-PNA2-CAT was placed in a 0.5% H2O2 solution, a large number of bubbles were generated. The activity of CAT was evaluated using a BCA kit, and the figure shows that CAT was successfully modified onto HMSN.

[0053] like Figure 5 As shown, the motion of HMSN-PNA1 in H2O2 solution is typical Brownian motion, while HMSN-PNA1-PNA2-CAT exhibits directional self-propulsion motion.

[0054] like Figure 6 As shown in the image, the cell viability graphs of 4T1 and MCF-7 cells after 48 hours of incubation with HMSN-PNA1-PNA2-CAT demonstrate that HMSN-PNA1-PNA2-CAT exhibits good biocompatibility. Figure 7 As shown in the figure, the cell viability of 4T1 cells after incubation with HMSN-PNA1-PNA2-CAT and 0.01% H2O2 for 48 hours shows that HMSN-PNA1-PNA2-CAT and 0.01% H2O2 do not exhibit cytotoxicity.

[0055] Example 2

[0056] A method for fabricating a detachable, brakeable nanomotor includes the following steps:

[0057] 1) Preparation of SN: Solid silica was prepared using the sol-gel method. 6 mL of TEOS was rapidly added to a mixture of ethanol (74 mL), deionized water (10 mL), and ammonia solution (3.2 mL). The mixture was then stirred at room temperature for 0.5 h to form a solid silica sphere (SN) suspension. Finally, the product was centrifuged at 15000 r / min for 10 min, washed three times with ethanol and deionized water, and dispersed in 50 mL of water.

[0058] 2) Preparation of HMSN: 450 mg of CTAB was added to a mixture of 60 mL deionized water, 90 mL anhydrous ethanol, and 1.65 mL ammonia, and stirred at 30 °C for 2 h. Then, 300 mg of SN particles dispersed in 20 mL deionized water were sonicated for 30 min and added to the mixture, and stirring was continued for another 30 min. Then, 0.75 mL of TEOS was slowly added dropwise, and the mixture was stirred uniformly at 30 °C for 6 h. After the reaction, the mixture was centrifuged and washed three times with water to obtain the intermediate product mesoporous silica-CTAB (MSN-CTAB). The core SN was removed by etching: 400 mg of MSN-CTAB prepared in the previous step was sonicated and dispersed in 60 mL deionized water. Then, 2.0 g (0.2 M) of anhydrous sodium carbonate (Na₂CO₃) was added to the dispersion, and the mixture was reacted at 50 °C for 6 h. After the reaction, the white product was collected by centrifugation. After washing three times with ethanol, hollow mesoporous silica-CTAB (HMSN-CTAB) was obtained. The obtained product was extracted in a mixture of 100 mL ethanol and 9 mL hydrochloric acid at 80 °C for 24 h, and the template CTAB was removed. The product was collected by centrifugation and washed three times with alcohol to obtain the product HMSN.

[0059] 3) Preparation of HMSN-NH2: HMSN was dispersed in ethanol to a concentration of 1 mg·mL. -1 120 μL of 3-aminopropyltriethoxysilane (APTES) was slowly added dropwise to 10 mL of HMSN solution with stirring at room temperature. After 12 h, the product HMSN-NH2 was collected by centrifugation and washed three times with deionized water, and then dispersed in 10 mL of water.

[0060] 4) Preparation of HMSN-PNA1-PNA2-CAT: Add 300 μL of deionized water to a tube of PNA1 (25 nmol), and sonicate to disperse. Take 100 μL of this solution, add 3.79 mg of 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride (EDC·HCl) and 3.45 mg of N-hydroxysuccinimide (NHS), and stir to activate for 30 min. Take 300 μL of HMSN-NH2 (0.5 mg / mL), sonicate for 30 min, and add it to the above solution. Stir overnight at room temperature. Centrifuge and wash once with water, retaining the supernatant. Disperse the precipitate with 150 μL of deionized water to obtain HMSN-RNA1 (1 mg / mL). Add 300 μL Tris-HCl (pH=8, 50 mM, 50 mM NaCl) to a tube of PNA2 (25 nmol), disperse by sonication, and take 100 μL. Add HMSN-RNA1 (1 mg / mL). Stir overnight. Centrifuge and wash once with water, retain the supernatant, and disperse the precipitate with deionized water to obtain HMSN-PNA1-PNA2-NH2 (2 mg / mL). Activate 5 mg catalase (CAT) in a mixed solution containing 1 mL water, 3.79 mg EDC·HCl, and 3.45 mg NHS. Stir at room temperature for 30 min, then mix the above solution with the HMSN-PNA1-PNA2-NH2 solution and stir overnight. Finally, centrifuge and wash three times with water, disperse in 150 μL of deionized water to obtain HMSN-PNA1-PNA2-CAT (1 mg / mL).

[0061] Example 3

[0062] A method for fabricating a detachable, brakeable nanomotor includes the following steps:

[0063] 1) Preparation of SN: Solid silica was prepared by the sol-gel method. 8 mL of TEOS was rapidly added to a mixture of ethanol (50 mL), deionized water (10 mL), and ammonia solution (5 mL). The mixture was then stirred at room temperature for 0.5 h to form a solid silica sphere (SN) suspension. Finally, the product was centrifuged at 15000 r / min for 10 min, washed three times with ethanol and deionized water, and then dispersed in 50 mL of water.

[0064] 2) Preparation of HMSN: 300 mg of CTAB was added to a mixture of 90 mL deionized water, 90 mL anhydrous ethanol, and 1.65 mL ammonia, and stirred at 30 °C for 2 h. Then, 300 mg of SN particles dispersed in 20 mL deionized water were sonicated for 30 min and added to the mixture, and stirring was continued for another 30 min. Then, 0.75 mL of TEOS was slowly added dropwise, and the mixture was stirred uniformly at 30 °C for 6 h. After the reaction, the mixture was centrifuged and washed three times with water to obtain the intermediate product mesoporous silica-CTAB (MSN-CTAB). The core SN was removed by etching: 400 mg of MSN-CTAB prepared in the previous step was sonicated and dispersed in 60 mL deionized water. Then, 1.44 g (0.2 M) of anhydrous sodium carbonate (Na₂CO₃) was added to the dispersion, and the mixture was reacted at 50 °C for 6 h. After the reaction, the white product was collected by centrifugation. After washing three times with ethanol, hollow mesoporous silica-CTAB (HMSN-CTAB) was obtained. The obtained product was extracted in a mixture of 100 mL ethanol and 9 mL hydrochloric acid at 80 °C for 24 h, and the template CTAB was removed. The product was collected by centrifugation and washed three times with alcohol to obtain the product HMSN.

[0065] 3) Preparation of HMSN-NH2: HMSN was dispersed in ethanol to a concentration of 1 mg·mL. -1 100 μL of 3-aminopropyltriethoxysilane (APTES) was slowly added dropwise to 10 mL of HMSN solution with stirring at room temperature. After 12 h, the product HMSN-NH2 was collected by centrifugation and washed three times with deionized water, then dispersed in 10 mL of water.

[0066] 4) Preparation of HMSN-PNA1-PNA2-CAT: Add 300 μL of deionized water to a tube of PNA1 (25 nmol), and sonicate to disperse. Take 100 μL of this solution, add 3.79 mg of 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride (EDC·HCl) and 3.45 mg of N-hydroxysuccinimide (NHS), and stir to activate for 30 min. Take 300 μL of HMSN-NH2 (0.5 mg / mL), sonicate for 30 min, and add it to the above solution. Stir overnight at room temperature. Centrifuge and wash once with water, retaining the supernatant. Disperse the precipitate with 150 μL of deionized water to obtain HMSN-RNA1 (1 mg / mL). Add 300 μL Tris-HCl (pH=8, 50 mM, 50 mM NaCl) to a tube of PNA2 (25 nmol), disperse by sonication, and take 100 μL. Add HMSN-RNA1 (1 mg / mL). Stir overnight. Centrifuge and wash once with water, retain the supernatant, and disperse the precipitate with deionized water to obtain HMSN-PNA1-PNA2-NH2 (2 mg / mL). Activate 5 mg catalase (CAT) in a mixed solution containing 1 mL water, 3.79 mg EDC·HCl, and 3.45 mg NHS. Stir at room temperature for 30 min, then mix the above solution with the HMSN-PNA1-PNA2-NH2 solution and stir overnight. Finally, centrifuge and wash three times with water, disperse in 150 μL of deionized water to obtain HMSN-PNA1-PNA2-CAT (1 mg / mL).

[0067] Example 4

[0068] A method for fabricating a detachable, brakeable nanomotor includes the following steps:

[0069] 1) Preparation of SN: Solid silica was prepared using the sol-gel method. 6 mL of TEOS was rapidly added to a mixture of ethanol (74 mL), deionized water (10 mL), and ammonia solution (3.2 mL). The mixture was then stirred at room temperature for 0.5 h to form a solid silica sphere (SN) suspension. Finally, the product was centrifuged at 15000 r / min for 10 min, washed three times with ethanol and deionized water, and dispersed in 50 mL of water.

[0070] 2) Preparation of HMSN: 450 mg of CTAB was added to a mixture of 90 mL deionized water, 90 mL anhydrous ethanol, and 1.65 mL ammonia, and stirred at 30 °C for 2 h. Then, 300 mg of SN particles dispersed in 20 mL deionized water were sonicated for 30 min and added to the mixture, and stirring was continued for another 30 min. Then, 1.0 mL of TEOS was slowly added dropwise, and the mixture was stirred uniformly at 30 °C for 6 h. After the reaction, the mixture was centrifuged and washed three times with water to obtain the intermediate product mesoporous silica-CTAB (MSN-CTAB). The core SN was removed by etching: 400 mg of MSN-CTAB prepared in the previous step was sonicated and dispersed in 60 mL deionized water. Then, 1.0 g (0.2 M) of anhydrous sodium carbonate (Na₂CO₃) was added to the dispersion, and the mixture was reacted at 50 °C for 6 h. After the reaction, the white product was collected by centrifugation. After washing three times with ethanol, hollow mesoporous silica-CTAB (HMSN-CTAB) was obtained. The obtained product was extracted in a mixture of 100 mL ethanol and 9 mL hydrochloric acid at 80 °C for 24 h, and the template CTAB was removed. The product was collected by centrifugation and washed three times with alcohol to obtain the product HMSN.

[0071] 3) Preparation of HMSN-NH2: HMSN was dispersed in ethanol to a concentration of 1 mg·mL. -1 120 μL of 3-aminopropyltriethoxysilane (APTES) was slowly added dropwise to 10 mL of HMSN solution with stirring at room temperature. After 12 h, the product HMSN-NH2 was collected by centrifugation and washed three times with deionized water, and then dispersed in 10 mL of water.

[0072] 4) Preparation of HMSN-PNA1-PNA2-CAT: Add 300 μL of deionized water to a tube of PNA1 (50 nmol), and sonicate to disperse. Take 100 μL of this solution, add 3.79 mg of 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide hydrochloride (EDC·HCl) and 3.45 mg of N-hydroxysuccinimide (NHS), and stir to activate for 30 min. Take 300 μL of HMSN-NH2 (0.5 mg / mL), sonicate for 30 min, and add it to the above solution. Stir overnight at room temperature. Centrifuge and wash once with water, retaining the supernatant. Disperse the precipitate with 150 μL of deionized water to obtain HMSN-RNA1 (1 mg / mL). Add 300 μL Tris-HCl (pH=8, 50 mM, 50 mM NaCl) to a tube of PNA2 (25 nmol), disperse by sonication, and take 100 μL. Add HMSN-RNA1 (1 mg / mL). Stir overnight. Centrifuge and wash once with water, retain the supernatant, and disperse the precipitate with deionized water to obtain HMSN-PNA1-PNA2-NH2 (2 mg / mL). Activate 5 mg catalase (CAT) in a mixed solution containing 1 mL water, 3.79 mg EDC·HCl, and 3.45 mg NHS. Stir at room temperature for 30 min, then mix the above solution with the HMSN-PNA1-PNA2-NH2 solution and stir overnight. Finally, centrifuge and wash three times with water, disperse in 150 μL of deionized water to obtain HMSN-PNA1-PNA2-CAT (1 mg / mL).

[0073] The nanomedicine carrier of this invention is composed of hollow mesoporous silica spheres (HMSN) and catalase (CAT)-modified peptide nucleic acid (PNA). The material uses tetraethyl orthosilicate as the silicon source to synthesize 130 nm solid silica spheres; based on this, 220 nm mesoporous silica is synthesized to coat the solid silica spheres using tetraethyl orthosilicate as the silicon source and hexadecyltrimethylammonium bromide (CTAB) as the pore-forming agent. Hollow mesoporous silica spheres (150 nm) are obtained by etching the core silica spheres with sodium carbonate. Subsequently, 3-aminopropyltrimethoxysilane is used to modify the spheres with amino groups, linking PNA1 (5'-HOOC-O-linker-TGTTGACGAGCG-3') to HMSN via amide bonds, and forming a double-stranded structure with catalase (CAT)-modified PNA2 (5'-CAT-O-linker-CGTCAACA-3') through base complementarity pairing. The nanomotor provided by this invention can achieve deceleration and braking within the body by adding a decoy chain PNA3 (5'-O-linker-CGCTCGTCDDCD-3') to disassemble the nanomotor's engine CAT.

[0074] The preferred embodiments of the present invention disclosed above are merely illustrative of the invention. These preferred embodiments do not exhaustively describe all details, nor do they limit the invention to specific implementations. Clearly, many modifications and variations can be made based on the content of this specification. This specification selects and specifically describes these embodiments to better explain the principles and practical applications of the invention, thereby enabling those skilled in the art to better understand and utilize the invention. The invention is limited only by the claims and their full scope and equivalents.

Claims

1. A method for preparing an engine-disassembled brakeable nanomotor, characterized in that, The method comprises the following steps: 1) Preparation of SN: solid silica is prepared by sol-gel method, TEOS is quickly added into the mixture of deionized water, anhydrous ethanol and ammonia water, and then the mixture is stirred at room temperature to form a SN suspension; finally, the product is centrifuged and washed, and the SN particles are dispersed in deionized water; 2) Preparation of HMSN: CTAB is added into the mixture of deionized water, anhydrous ethanol and ammonia water, and stirred uniformly; then the SN particles dispersed in deionized water are ultrasonically treated and added into the mixture, and stirring is continued; TEOS is slowly added dropwise, and stirring is uniformly carried out; after the reaction is completed, centrifugation and water washing are carried out to obtain the intermediate product mesoporous silica-CTAB, which is recorded as MSN-CTAB; The core SN is removed by etching method: the prepared MSN-CTAB is ultrasonically dispersed in deionized water, and then anhydrous sodium carbonate is added into the dispersion, and the mixture is reacted at 40-60℃; after the reaction is completed, the white product is collected by centrifugation, and is washed with ethanol for multiple times to obtain hollow mesoporous silica-CTAB, which is recorded as HMSN-CTAB; The obtained product is extracted in a mixture of ethanol and hydrochloric acid at 75-85℃ to remove the template CTAB; the product is collected by centrifugation and is alcohol-washed to obtain the product HMSN; 3) Preparation of HMSN-NH2: HMSN is dispersed into ethanol, and 3-aminopropyltriethoxysilane is slowly added dropwise into the HMSN solution under stirring at room temperature; the product HMSN-NH2 is collected by centrifugation and is washed with deionized water, and then is dispersed in deionized water; 4) Preparation of HMSN-PNA1-PNA2-CAT: deionized water is added into a tube of PNA1, and is ultrasonically dispersed, and 1-ethyl-3-(3-dimethylaminopropyl) carbonyldiimidazole hydrochloride and N-hydroxysuccinimide are added and stirred to activate; HMSN-NH2 is taken, is ultrasonically treated, and is added into the above solution, and is stirred at room temperature overnight; centrifugation and water washing are carried out, the supernatant is reserved, and the precipitate is dispersed in deionized water to obtain HMSN-PNA1; Tris-HCl is added into a tube of PNA2, is ultrasonically dispersed, and is added into HMSN-PNA1, and is stirred overnight; centrifugation and water washing are carried out, the supernatant is reserved, and the precipitate is dispersed in deionized water to obtain HMSN-PNA1-PNA2-NH2; catalase is dissolved in a mixed solution containing deionized water, EDC·HCl and NHS to activate, and after stirring at room temperature, the above solution is mixed with the HMSN-PNA1-PNA2-NH2 solution and is stirred overnight; finally, centrifugation and water washing are carried out for multiple times, and the product is dispersed in deionized water to obtain HMSN-PNA1-PNA2-CAT.

2. The production method according to claim 1, wherein In step 1), the mass ratio of ethanol, deionized water and ammonia solution in the mixture is 25-50:5:1-4, and the concentration of the ammonia solution is 25-28%; the mass ratio of the mixture and TEOS is 31-59:

3.

3. The production method according to claim 1, wherein In step 2), the mass ratio of deionized water, anhydrous ethanol and ammonia water in the mixed solution is 600:600:11, the concentration of the ammonia water solution is 25-28%, and the mass ratio of the mixed solution, SN particles and TEOS is 3633:6000:10-30.

4. The production method according to claim 1, wherein In step 2), the mass ratio of MSN-CTAB and anhydrous sodium carbonate in the mixture is 1000:2-5, and the mass ratio of ethanol and hydrochloric acid in the mixture is 100:

9.

5. The production method according to claim 1, wherein In step 3), the mass ratio of HMSN and 3-aminopropyltriethoxysilane is 1:

12.

6. The production method according to claim 1, wherein In step 4), the mass ratio of 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide hydrochloride and N-hydroxysuccinimide is 40-100:

69.

7. The production method according to claim 1, wherein In step 4), the molar ratio of PNA1 and PNA2 is 1:

1.

8. The production method according to claim 1, wherein In step 4), the sequence of PNA1 is 5'-HOOC-O-linker-TGTTGACGAGCG-3'; The sequence of PNA2 is 5'-CAT-O-linker-CGTCAACA-3'; The sequence of PNA3 is 5'-O-linker-CGCTCGTCDDCD-3'.

9. The production method according to claim 1, wherein In step 4), the mass ratio of deionized water, EDC·HCl and NHS in the mixed solution is 100:379:

345.

10. An engine disassembled brakeable nanomotor, characterized in that, Prepared by the preparation method of any one of claims 1-9.

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