CuS-BSA protein complex light-driven motor and preparation method and application thereof

Abstract

The application discloses a CuS-BSA protein compound light-driven motor and a preparation method and application thereof, and belongs to the technical field of organic-inorganic and biomaterial composite. The CuS-BSA compound is prepared by using a bovine serum albumin solution, copper nitrate trihydrate and sodium sulfide, then the CuS-BSA compound is prepared into CuS-BSA hollow vesicles by using n-butanol, and finally the CuS-BSA hollow vesicles are made into the CuS-BSA compound light-driven motor. The CuS-BSA compound light-driven motor prepared by the application has excellent biocompatibility and degradability, does not need additional chemical fuel, and is suitable for physiological environment. The CuS-BSA compound light-driven motor has the advantages of high photo-thermal conversion efficiency, fast and controllable driving speed, simple preparation method, greenness, low cost and the like. The CuS-BSA compound light-driven motor can be applied to multiple fields such as drug targeted delivery, biosensing and detection, minimally invasive surgery and cell micro-operation.

Description

Technical Field

[0001] This invention relates to the field of organic-inorganic and biomaterial composite technology, specifically to a CuS-BSA protein complex optical motor, its preparation method, and its application. Background Technology

[0002] Micro / nanomotors are artificial intelligence devices that can convert external energy into their own mechanical motion, showing broad application prospects in fields such as targeted drug delivery, biosensing, minimally invasive surgery, and environmental governance. Among the many driving methods, light-driven micro / nanomotors have become a hot research topic due to their unique advantages such as non-contact control, high spatiotemporal precision, remote controllability, and gentle energy input. Currently, researchers have developed a variety of light-driven motors, whose driving mechanisms mainly rely on the photothermal effect, photocatalytic reaction, or photoinduced deformation of materials. In the existing technology, the main materials of light-driven motors are mainly divided into two categories: (1) inorganic / metallic materials, such as titanium dioxide (TiO2), zinc oxide (ZnO), or precious metals such as gold and platinum, which generate propulsion through photocatalytic decomposition of fuels (such as hydrogen peroxide) or plasma photothermal effect; (2) synthetic polymer materials, such as azobenzene liquid crystal polymers or thermosensitive hydrogels, which generate mechanical energy by photoinduced molecular conformational changes or volume contraction.

[0003] However, the aforementioned materials have significant shortcomings in practical applications. For inorganic and metallic materials, their poor biodegradability and long-term retention in vivo may trigger toxic or inflammatory responses; furthermore, their driving processes often require the addition of high concentrations of chemical fuels such as hydrogen peroxide, further limiting their application in physiological environments. For synthetic polymers, although they exhibit good flexibility, the biocompatibility of some synthetic monomers remains to be verified, and their complex photoresponsive deformation mechanisms typically require specific molecular design, making it difficult to simultaneously achieve both driving efficiency and stability.

[0004] Therefore, how to develop a light-driven motor with excellent biocompatibility that can be safely driven in physiological environments without additional chemical fuels is a technical problem that urgently needs to be solved in this field. Summary of the Invention

[0005] In view of this, the present invention provides a CuS-BSA protein complex optical motor, its preparation method and application.

[0006] To achieve the above objectives, the present invention adopts the following technical solution: A method for preparing a CuS-BSA protein complex optically driven motor includes the following steps: (1) Add copper nitrate trihydrate (CuNO3·3H2O) solution to bovine serum albumin (BSA) solution and stir until homogeneous.2+ After fully complexing with BSA, the solution turns light blue. Then, sodium hydroxide (NaOH) solution is added dropwise to the mixed solution to adjust the pH, followed by the rapid addition of sodium sulfide (Na2S) solution, at which point the solution turns dark blue-black. The resulting reaction system is heated and stirred, and after stirring, it is kept at a constant temperature and allowed to stand. Finally, it is cooled to room temperature, and the product is purified by dialysis to remove excess metal salt and sodium hydroxide. The purified product is then freeze-dried to obtain CuS-BSA complex solid powder, which is stored for later use. (2) Dissolve the CuS-BSA composite solid powder in water to obtain a composite solution. Add the composite solution to a centrifuge tube containing n-butanol. After vortexing, let it stand at room temperature. Use a liquid gun to remove the upper n-butanol solution to obtain a concentrated solution containing CuS-BSA hollow vesicles. (3) Add concentrated CuS-BSA capsules to a mixture of water and tetrahydrofuran, allow the mixture to stand at room temperature, and then remove the mixture of water and tetrahydrofuran to obtain an asymmetric CuS-BSA structure. Add n-butanol to the collected CuS-BSA asymmetric structure to fix the structure, and you can obtain a CuS-BSA complex photo-driven motor with regular morphology and stable structure.

[0007] Furthermore, the volume ratio of bovine serum albumin solution to copper nitrate trihydrate solution in step (1) is 7.5:1-2; The volume ratio of the bovine serum albumin solution to the sodium sulfide solution is 7.5-8:2.

[0008] Furthermore, the concentration of the bovine serum albumin solution is 33.3 mg / mL; The concentration of the copper nitrate trihydrate solution is 0.2 M; The concentration of the sodium sulfide solution is 48 mg / mL; The sodium hydroxide solution has a concentration of 5 M, and the pH of the resulting mixed solution is adjusted to 12 using the sodium hydroxide solution.

[0009] Furthermore, in step (1), the bovine serum albumin solution and copper nitrate trihydrate solution are mixed and stirred at 200-300 rpm for 5-10 min; Add sodium sulfide solution to the mixed solution after reaction, and react for 30-35 min at 60℃ and 200-300 rpm stirring, then keep warm and let stand for 30 min. The dialysis time was 24 hours. The storage temperature for future use is 4°C.

[0010] Furthermore, the volume ratio of the CuS-BSA complex solution to n-butanol in step (2) is 0.1:0.7-1; The concentration of the CuS-BSA complex solution is 3-5 mg / mL.

[0011] The vortex oscillation time in step (2) is 1-2 min; Let it stand at room temperature for 20-30 minutes.

[0012] Furthermore, the volume ratio of the mixed solution of water and tetrahydrofuran to the hollow vesicles of CuS-BSA in step (3) is 200:2; The volume ratio of hollow vesicles to n-butanol in the CuS-BSA is 2:200.

[0013] Furthermore, the volume ratio of water to tetrahydrofuran in the mixed solution of water and tetrahydrofuran is 100:30-35; The room temperature static reaction time is 24 h.

[0014] Compared with existing light-driven motors based on inorganic / metallic materials or synthetic polymers (such as azobenzene liquid crystal polymers), the present invention has the following advantages: 1. Excellent biocompatibility and biodegradability This invention uses bovine serum albumin (BSA) as the main protein material of the complex. BSA is a naturally derived biological macromolecule that is non-toxic and non-immunogenic, and can be degraded into amino acids by proteases in vivo, ultimately metabolized into water and carbon dioxide. Copper sulfide (CuS) also exhibits good chemical stability in physiological environments, and as BSA degrades, CuS nanoparticles can be cleared by phagocytes or slowly metabolized, avoiding the toxicity or inflammatory reactions caused by long-term retention of inorganic materials. The motor prepared by this invention can be safely used in the biomedical field, such as in vivo drug delivery and minimally invasive surgery.

[0015] 2. No additional chemical fuels required, suitable for physiological environments. Most existing inorganic photodynamic motors rely on high-concentration hydrogen peroxide as fuel, using photocatalytic decomposition to generate oxygen bubbles for propulsion. However, hydrogen peroxide is cytotoxic and cannot be used in physiological environments. This invention completely eliminates chemical fuels, utilizing only the photothermal effect of CuS to generate thermophoretic or bubble propulsion. It can operate normally in water, buffer solutions, cell culture media, serum, and even living tissue fluid, truly achieving safe propulsion in physiological environments.

[0016] 3. High photothermal conversion efficiency, fast and controllable driving speed. The CuS used in this invention is a p-type semiconductor nanomaterial whose localized surface plasmon resonance effect significantly improves its photothermal conversion efficiency compared to many traditional photothermal materials. In this invention, the BSA-mediated synthesized CuS nanoparticles exhibit uniform particle size and good dispersion, enabling rapid heating under near-infrared light irradiation at relatively low power densities, thus driving a motor to generate rapid motion. By adjusting the light power density, irradiation time, or irradiation position, precise control over the motor's speed, direction, and start / stop can be achieved.

[0017] 4. The preparation method is simple, green, and low in cost. In existing technologies, the fabrication of light-driven motors made of precious metals or complex polymers often requires multi-step organic synthesis, high-temperature treatment, or expensive raw materials. This invention utilizes in-situ growth of CuS nanoparticles in an aqueous phase via BSA, employing mild reaction conditions, requiring no complex equipment, and using widely available raw materials (BSA is an industrial-grade product, and copper salts and sulfur sources are inexpensive), making it suitable for large-scale production and widespread application.

[0018] 5. Wide range of applications The CuS-BSA protein complex photoelectric motor prepared by this invention exhibits excellent performance and can be applied to, but is not limited to, the following fields: Targeted drug delivery: Loaded with chemotherapy drugs, the drugs actively move to the tumor site under near-infrared light drive, and combined with the photothermal therapy function of CuS, it realizes photo-driven-photothermal-chemotherapy synergistic treatment.

[0019] Biosensing and Detection: As motion probes, they detect changes in the concentration of biomarkers or chemicals at the microscale (e.g., using changes in motion speed to reflect environmental viscosity, temperature, or pH).

[0020] Minimally invasive surgery and cellular micromanipulation: movement within microvessels or tissue spaces to clear blood clots, remove foreign objects, or deliver gene-editing tools to single cells. Attached Figure Description

[0021] Figure 1 The hydrodynamic radius of pure BSA and CuS-BSA composite in the embodiments of the present invention is shown in (a); transmission electron microscopy image of CuS-BSA composite is shown in (b). Figure 2 Scanning electron microscope (SEM) images of CuS-BSA vesicles in Example 2 of the present invention (a); SEM images of CuS-BSA asymmetric Janus nanobowls (bc); and SEM image of the surface of CuS-BSA nanobowls (d). Figure 3 The image shows the photothermal properties of the CuS-BSA Janus nanobowl under 808 nm laser irradiation in Experimental Example 1 of this invention. Figure 4This is a schematic diagram of the motion behavior of CuS-BSA Janus nanobowls, BSA Janus nanobowls and CuS-BSA hollow vesicles under 808 nm laser irradiation in this invention; Figure 5 The images shown are scanning electron microscope images of the CuS-BSA assemblies of the present invention at concentrations of 1 mg / mL and 8 mg / mL, respectively. Figure 6 The image shows a scanning electron microscope (SEM) image of the CuS-BSA hollow capsule of the present invention in a mixed solution of 25% and 37% water / tetrahydrofuran by volume. Figure 7 This is a flowchart illustrating the preparation process of the light-driven motor based on the CuS-BSA protein complex of this invention. Detailed Implementation

[0022] 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.

[0023] Example 1: Preparation of CuS-BSA composite solid powder Add 1 mL of 0.2 M copper nitrate trihydrate (CuNO3·3H2O) solution to 7.5 mL of bovine serum albumin (BSA) solution with a concentration of 33.3 mg / mL, and stir at 200 rpm for 10 min to allow Cu to settle. 2+ The mixture was fully complexed with BSA, and the resulting solution was light blue. 5 M sodium hydroxide (NaOH) solution was added dropwise to the mixture until the pH of the reaction mixture reached 12. Then, 2 mL of sodium sulfide (Na2S) solution with a concentration of 48 mg / mL was added, at which point the solution turned dark blue-black. The resulting reaction system was reacted at 60℃ and 300 rpm for 30 min, then kept at this temperature and allowed to stand for 30 min. Finally, it was cooled to room temperature and dialyzed for 24 hours to purify the product, removing excess metal salt and sodium hydroxide. The CuS-BSA complex solid powder was obtained by freeze-drying and stored at 4℃ for later use.

[0024] The hydrodynamic radius and microstructure of the CuS-BSA composite were characterized using a nanoparticle size analyzer and transmission electron microscopy, respectively. The results are as follows: Figure 1 As shown.

[0025] Results Analysis: The hydrodynamic radii of BSA and CuS-BSA nanocomposites were characterized using a nanoparticle size analyzer. Figure 1a) The hydrodynamic size of an aqueous solution of pure BSA protein is approximately 11.7 nm. When BSA protein is used as a biotemplate for preparing CuS-BSA nanocomposites, Cu... 2+ When metal ions are added to a BSA aqueous solution, the functional group of the BSA protein itself is Cu. 2+ Metal ion binding provides active sites, thereby inducing the controllable nucleation and growth of CuS nanoparticles, ultimately yielding CuS-BSA nanocomposites with a hydrodynamic size increased to 28.7 nm.

[0026] The morphology, structure, and crystal form of the CuS-BSA composite were characterized using transmission electron microscopy. Low-magnification transmission electron microscopy results showed that the average diameter of CuS-BSA was approximately 5.2 nm, and it exhibited good monodispersity. Figure 1 b). Further analysis of its crystal structure using high-resolution transmission electron microscopy revealed a lattice fringes of 0.41 nm between crystal planes in a single CuS@BSA composite. These results demonstrate the successful synthesis of the CuS-BSA composite.

[0027] Example 2: Preparation of CuS-BSA composite optical motor (1) Take the CuS-BSA complex prepared in Example 1, prepare an aqueous solution of CuS-BSA complex with a concentration of 3 mg / mL, take 100 μL of complex solution with a liquid gun, carefully add it to a centrifuge tube containing 0.9 mL n-butanol, vortex for 1 min, and let stand at room temperature for 30 min to obtain CuS-BSA hollow vesicles with regular morphology.

[0028] (2) 200 μL of a 30% (v / v) water / tetrahydrofuran mixture was added dropwise to 2 μL of concentrated CuS-BSA vesicles, and the mixture was allowed to stand at room temperature for 24 h. Subsequently, the water / tetrahydrofuran mixture was completely removed, and 200 μL of n-butanol was added to the collected CuS-BSA asymmetric structure to fix the structure, thus obtaining a CuS-BSA composite photodynamic motor with regular morphology and stable structure.

[0029] The morphology and structure of the CuS-BSA optical drive motor were characterized using scanning electron microscopy. The results are as follows: Figure 2 As shown.

[0030] Results Analysis: Using CuS-BSA complex as the building block, a large number of CuS-BSA hollow vesicles can be rapidly and easily prepared via the water-n-butanol soft emulsion restricted template method. Figure 2 a) The obtained CuS-BSA capsules were transferred to a water / tetrahydrofuran mixed solution. Under the induction of selective solvent effect, the capsules underwent asymmetric collapse deformation, eventually transforming from a spherical shape into a Janus nanobowl. Figure 2b and Figure 2 c). Characterization of the surface of a single Janus nanobowl reveals that it has a certain degree of roughness, and CuS nanoparticles are uniformly dispersed on the surface of the nanobowl. Figure 2 d). The mechanism of the above deformation process can be summarized as follows: under the induction of solvent exchange, the thermodynamically driven phase separation and the non-equilibrium process of kinetic trapping work synergistically. Among them, the hydrophobic interaction force inside CuS-BSA is the main driving force, which causes the molecular chain segments to undergo asymmetric contraction and directional alignment, and finally reshape the spherical hollow capsule into Janus nanobowls with internal and external chemical differences.

[0031] Experimental Example 1 To evaluate the photothermal properties of CuS-BSA Janus nanobowls, they were irradiated with an 808 nm laser.

[0032] As a control, unmodified CuS-BSA Janus nanobowls were treated under the same conditions. Under laser irradiation (output laser power 1.2 W), the temperature of the CuS-BSA Janus nanobowl dispersion increased significantly (see...). Figure 3 a), and the temperature rise increases with increasing laser output power (see Figure 3 (b) Within 10 min, the maximum temperature change (ΔT) of the CuS-BSA Janus nanobowl at a concentration of 3 mg / mL reached 31.5 K. This significant temperature increase indicates that the nanobowl possesses a highly efficient photothermal effect. As expected, its plasma heating characteristics are related to the laser output power and irradiation time, indicating that the photothermal effect is easily tunable. These results demonstrate that the CuS-BSA Janus nanobowl can efficiently and rapidly convert absorbed laser energy into heat energy, providing technical support for the subsequent motion of a light-driven motor.

[0033] Experimental Example 2 The autonomous motion capability of the CuS-BSA Janus nanobowls was investigated by irradiating them with an 808 nm laser and tracking their motion behavior using an inverted optical microscope. Based on the motion trajectory, the corresponding mean square displacement (MSD) and velocity were derived using a Golestanian self-diffusion model. Under laser irradiation, the Janus nanobowls exhibited directional autonomous motion away from the laser source. This negative phototaxis may be attributed to the asymmetric structure of the Janus nanobowls and the non-uniform plasma heating effect around them.

[0034] Furthermore, compared with a series of control groups (CuS-BSA hollow vesicles, BSA Janus nanobowls), only CuS-BSA Janus nanobowls exhibited high-speed directional autonomous motion, while CuS-BSA hollow vesicles, BSA hollow vesicles, and BSA Janus nanobowls only exhibited enhanced Brownian motion.

[0035] Comparative Example 1 (1) Using the CuS-BSA complex prepared in Example 1, aqueous solutions of CuS-BSA complex with concentrations of 1 mg / mL and 8 mg / mL were prepared respectively. 100 μL of the above complex solution was taken with a liquid pipette and carefully added to a centrifuge tube containing 0.9 mL n-butanol. After vortexing for 1 min, the solution was left to stand at room temperature for 30 min. A large number of CuS-BSA hollow vesicles were collected at the bottom of the centrifuge tube.

[0036] (2) 200 μL of a 30% (v / v) water / tetrahydrofuran mixture was added dropwise to 2 μL of concentrated CuS-BSA vesicles, and the mixture was allowed to stand at room temperature for 24 h. The water / tetrahydrofuran mixture was then completely removed, and 200 μL of n-butanol was added to the collected CuS-BSA asymmetric structure to fix the structure. The morphology and structure of the CuS-BSA assembly were characterized using scanning electron microscopy.

[0037] Results analysis: When constructing Janus nanobowls using the CuS-BSA composite, their morphology and structure significantly depend on the concentration of the CuS-BSA composite building blocks. For example... Figure 5 As shown, when the CuS-BSA concentration is 1 mg / mL, thin-walled flexible vesicles can be formed using the water-n-butanol soft emulsion confined template method. Under subsequent solvent induction, these flexible vesicles further evolve into porous, irregular morphologies with concave features. However, when the CuS-BSA concentration is increased to 8 mg / mL, rigid vesicles with thicker walls are formed; during water / tetrahydrofuran-induced deformation, the vesicles break down directly, failing to form a Janus structure. These results indicate that the successful construction of CuS-BSA Janus nanobowls is closely related to the flexibility of the hollow vesicles.

[0038] Comparative Example 2 (1) Using the CuS-BSA complex prepared in Example 1, prepare an aqueous solution of CuS-BSA complex with a concentration of 3 mg / mL. Take 100 μL of the above complex solution with a liquid gun and carefully add it to a centrifuge tube containing 0.9 mL n-butanol. After vortexing for 1 min, let it stand at room temperature for 30 min to obtain CuS-BSA hollow vesicles with regular morphology.

[0039] (2) 200 μL of water / tetrahydrofuran mixtures with volume ratios of 25% and 37% were added dropwise to a 2 μL concentrate of CuS-BSA vesicles, and the mixtures were allowed to stand at room temperature for one day. The water / tetrahydrofuran mixtures were removed, and 200 μL of n-butanol was added to the collected CuS-BSA asymmetric structure to fix the structure, thus obtaining a CuS-BSA composite photodynamic motor with a regular morphology and stable structure. The morphology and structure of the CuS-BSA assembly were characterized using scanning electron microscopy.

[0040] Results Analysis: The volume ratio of the water / tetrahydrofuran mixed solution significantly affected the deformation process of CuS-BSA hollow capsules. When the water content in the mixed solution was low (25%), the hollow capsules did not undergo significant deformation and maintained their original symmetrical spherical structure (see...). Figure 6 When the water content increases to 37%, the hollow capsule directly disintegrates into a porous spherical framework. This is because water, as a good solvent for BSA proteins, can significantly promote the hydration and swelling of the hydrophilic parts of BSA, generating greater outward expansion stress, thereby leading to the disintegration of the capsule's morphological structure.

[0041] Although embodiments of the present invention have been shown and described above, it is understood that the above embodiments are exemplary and should not be construed as limiting the present invention. Those skilled in the art can make changes, modifications, substitutions and variations to the above embodiments within the scope of the present invention.

Claims

1. A method for preparing a CuS-BSA protein complex optically driven motor, characterized in that, Includes the following steps: (1) Add copper nitrate trihydrate solution to bovine serum albumin solution and stir until Cu 2+ The mixture is fully complexed with BSA, and then sodium hydroxide solution is added dropwise to the mixed solution after the reaction to adjust the pH, followed by sodium sulfide solution. The resulting reaction system is heated and stirred, and after stirring, it is kept at a constant temperature and allowed to stand. Finally, it is cooled to room temperature, and the product is purified by dialyzing. The purified product is freeze-dried to obtain CuS-BSA complex solid powder, which is stored for later use. (2) Dissolve the CuS-BSA composite solid powder in water to obtain a composite solution. Add the composite solution to a centrifuge tube containing n-butanol. After vortexing, let it stand at room temperature. Use a liquid gun to remove the upper n-butanol solution to obtain a concentrated solution containing CuS-BSA hollow vesicles. (3) Add concentrated CuS-BSA capsules to a mixture of water and tetrahydrofuran, allow the mixture to stand at room temperature, and then remove the mixture of water and tetrahydrofuran to obtain an asymmetric CuS-BSA structure. Add n-butanol to the collected CuS-BSA asymmetric structure to fix the structure, and you can obtain a CuS-BSA complex photo-driven motor with regular morphology and stable structure.

2. The method for preparing a CuS-BSA protein complex photoelectric motor according to claim 1, characterized in that, The volume ratio of bovine serum albumin solution to copper nitrate trihydrate solution in step (1) is 7.5:1-2; The volume ratio of the bovine serum albumin solution to the sodium sulfide solution is 7.5-8:

2.

3. The method for preparing a CuS-BSA protein complex photoelectric motor according to claim 2, characterized in that, The concentration of the bovine serum albumin solution was 33.3 mg / mL; The concentration of the copper nitrate trihydrate solution is 0.2 M; The concentration of the sodium sulfide solution is 48 mg / mL; The sodium hydroxide solution has a concentration of 5 M, and the pH of the resulting mixed solution is adjusted to 12 using the sodium hydroxide solution.

4. The method for preparing a CuS-BSA protein complex photoelectric motor according to claim 1 or 2, characterized in that, In step (1), the bovine serum albumin solution and copper nitrate trihydrate solution are mixed and stirred at 200-300 rpm for 5-10 min. Add sodium sulfide solution to the mixed solution after reaction, and react for 30-35 min at 60℃ and 200-300 rpm stirring, then keep warm and let stand for 30 min. The dialysis time was 24 hours. The storage temperature for future use is 4°C.

5. The method for preparing a CuS-BSA protein complex photoelectric motor according to claim 1, characterized in that, The volume ratio of the CuS-BSA complex solution to n-butanol in step (2) is 0.1:0.7-1; The concentration of the CuS-BSA complex solution is 3-5 mg / mL.

6. The method for preparing a CuS-BSA protein complex photoelectric motor according to claim 1 or 5, characterized in that, The vortex oscillation time in step (2) is 1-2 min; The settling time at room temperature is 20-30 minutes.

7. The method for preparing a CuS-BSA protein complex photoelectric motor according to claim 1, characterized in that, The volume ratio of the mixed solution of water and tetrahydrofuran to the hollow vesicles of CuS-BSA in step (3) is 200:2; The volume ratio of hollow vesicles to n-butanol in the CuS-BSA is 2:

200.

8. The method for preparing a CuS-BSA protein complex photoelectric motor according to claim 7, characterized in that, The volume ratio of water to tetrahydrofuran in the mixed solution of water and tetrahydrofuran is 100:30-35; The room temperature static reaction time is 24 h.

9. A CuS-BSA protein complex photoelectric motor, characterized in that, It is prepared by the method described in any one of claims 1-8.

10. The application of the CuS-BSA protein complex photoelectric motor of claim 9 in drug targeted delivery, biosensing and detection, or minimally invasive surgery and cell micromanipulation instruments.

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