Preparation method of acoustic / photosensitizer-manganese dioxide nanoparticles and product thereof

Abstract

The invention provides a preparation method of an acoustic / photosensitizer-manganese dioxide nanoparticle and a product thereof. According to the method, firstly, a potassium permanganate reduction method is adopted to precisely synthesize the high-purity MnO2 nano-substrate, and according to the method, the size uniformity and dispersion stability of nano-particles can be precisely controlled by regulating and controlling the pH value and temperature of a reaction system and the concentration of a reducing agent. On the basis, amphiphilic high-molecular polyethylene glycol-polycaprolactone (PEG-PCL) is modified on the surface of the MnO2 nanometre. The whole preparation process is green and mild, and complex equipment is not needed; the reaction conditions are mild, nanoparticles are uniform, and the acoustic / photosensitizer can be combined and applied to an imaging technology to realize real-time monitoring.

Description

TECHNICAL FIELD

[0001] The present application relates to the field of nanomedicine, and particularly relates to a preparation method of a sound / light sensitizer manganese dioxide nanoparticle and a product thereof. BACKGROUND

[0002] MnO2 has good biocompatibility, low toxicity in a physiological environment, and reduces the biological safety risk of medical materials. It has pH responsiveness and redox activity, and can be degraded in a tumor microenvironment and a high active oxygen atmosphere, to realize intelligent control of drugs and diagnosis and treatment functions. MnO2 in a nanometer scale has a high specific surface area, can efficiently load drugs, and has certain light-heat conversion efficiency and magnetic resonance imaging contrast capability. Therefore, MnO2 has become a functional material with great potential in the field of nanomedicine due to its unique physical and chemical and biological properties.

[0003] An invention with a publication number CN110104689A discloses a hollow manganese dioxide nanoparticle. The hollow nanostructure is obtained by reducing potassium permanganate in a hydrochloric acid aqueous solution environment through one-time feeding or multiple feeding of a reducing organic acid citric acid. The hollow manganese dioxide nanoparticle has the advantages of simple preparation, stable properties, and controllable size and structure, is rich in carboxyl groups on the surface, and can be applied in the fields of targeted modification, drug carriers, and magnetic resonance imaging.

[0004] The rapid development of nanomedicine promotes the innovation of non-invasive and precise diagnosis and treatment technology. As the core functional molecule of a sound / light diagnosis and treatment system, a sound / light sensitizer has the response characteristics to specific physical signals, carries out sound and light dynamic therapy, becomes a key material for solving the problems of poor targeting and strong side effects of traditional treatment, and has a continuously highlighted application potential in the field of nanomedicine. In the treatment of traditional tumors, bacterial infections and other diseases, chemotherapy and antibiotic means are prone to drug resistance and have significant side effects, and thus there is an urgent need for new minimally invasive and precise treatment methods. The integration of "diagnosis-treatment-efficacy evaluation" is increasingly urgent, and a sound sensitizer and a light sensitizer can be combined with imaging technology to realize real-time monitoring of the treatment process. SUMMARY

[0005] The present application aims to provide a sound / light sensitizer to realize real-time monitoring.

[0006] Still another object of the present application is to provide a preparation method of the sound / light sensitizer.

[0007] Still another object of the present application is to provide a sound / light sensitizer manganese dioxide nanoparticle product prepared by the above method.

[0008] The object of the present application is achieved by the following scheme: In a first aspect, the present application provides a preparation method of a sound / light sensitizer manganese dioxide nanoparticle.

[0009] In the present application, the sound / light sensitizer refers to a sound sensitizer or a light sensitizer. The sound sensitizer (Acoustic Sensitizer) has a sensitive performance for sound wave excitation or detection, and is a kind of compound capable of responding to sound waves or sound. They will produce a series of physical or chemical changes under the action of sound waves or sound, so as to realize the functions of sound detection, sensing or conversion. The photoinitiator (photoinitiator) is also called photosensitizer (photosensitizer) or photocuring agent (photocuring agent), which is a kind of compound that can absorb energy of a certain wavelength in the ultraviolet light region (250-420 nm) or visible light region (400-800 nm), and generate free radicals, cations and the like, so as to initiate monomer polymerization, crosslinking and curing.

[0010] The present application adopts the potassium permanganate reduction method to accurately synthesize high-purity MnO2nanometer substrate, and the method can realize accurate control of the uniformity and dispersion stability of the nanoparticle size by adjusting the pH value, temperature and reducing agent concentration of the reaction system. On this basis, the amphiphilic polymer polyethylene glycol-poly (ε-caprolactone) (PEG-PCL) is modified on the surface of the MnO2nanoparticles.

[0011] The preparation method of the present application comprises the following steps: Step (1): slowly drop 1 part of potassium permanganate aqueous solution into 1.2-3 parts of reducing agent solution, and ultrasonic dispersion. The pH value is adjusted to 2-7, the reaction temperature is controlled at 25-60 ℃ range, the reaction is carried out for 4-24 hours, the solution changes from purple red to brown black, and the solution is neutralized to neutral with sodium hydroxide; Step (2): add 1-5 parts of polyethylene glycol-poly (ε-caprolactone) (PEG-PCL) solution, stir at room temperature, and react for 6-12 hours; Step (3): according to 10%-20% of the mass of PEG-PCL, the sound sensitizer or light sensitizer is dissolved in dimethyl sulfoxide, and then added to the solution of step (2) and stirred; Step (4): the mixed solution is dialyzed through water overnight, centrifuged, and the unreacted substances are removed, and the MnO2nanoparticles loaded with sound sensitizer or light sensitizer are collected.

[0012] Preferably, in step (1), the reducing agent is selected from one or more of common reducing agents such as oxalic acid, sodium sulfite, polyallylamine hydrochloride (preferably with an average molecular weight of 15000). The weight of the reducing agent used is 1.2-2 parts, for example 1.5-1.7 parts, etc., based on 1 part of the weight of potassium permanganate.

[0013] Preferably, after ultrasonic dispersion, the pH value is adjusted to 3-6, or 4-5.

[0014] Preferably, the reaction temperature of step (1) is controlled in the range of 35-45 ℃, for example 38, 40, 42, 44 ℃, etc.

[0015] Preferably, the reaction of step (1) is carried out for 6-12 hours; or for 8-10 hours.

[0016] Preferably, the product of step (1) is neutral in pH, and a base, including a strong base or a weak base, can be used to adjust the pH to neutral, depending on the pH of the solution to be adjusted. Generally, the pH of a neutral solution is 7. In one preferred embodiment of the present application, sodium hydroxide is used to adjust the solution to neutral.

[0017] Preferably, in step (2), the average molecular weight of PEG-PCL is preferably 7000.

[0018] Preferably, in step (2), the amount of polyethylene glycol-polycaprolactone used is 2-5 parts, for example 3-4 parts.

[0019] The reaction temperature of step (2) is room temperature, which is understood to be 15-30 ℃ or 18-28 ℃, preferably 20-25 ℃, including 22, 23, 24 ℃, etc.

[0020] Preferably, in step (3), the amount of sonophoto sensitizer used is 15-20% of the mass of polyethylene glycol-polycaprolactone, including 16%, 17%, 18%, 19%, etc.

[0021] Preferably, in step (3), the sonophoto sensitizer can be selected from common sonophoto sensitizers such as protoporphyrin, chlorin E6, indocyanine green, etc., including various organic sonophoto sensitizers. In one preferred embodiment of the present application, chlorin E6 is used.

[0022] In a second aspect, the present application provides a sonophoto sensitizer@manganese dioxide nanoparticle, which can be prepared according to any of the above-mentioned methods. Among them, polyethylene glycol (PEG) endows the nanoparticle with excellent water solubility and biocompatibility, and poly-caprolactone (PCL) forms a hydrophobic core to efficiently load the liposoluble organic sonophoto sensitizer through hydrophobic interaction.

[0023] The sonophoto sensitizer@manganese dioxide nanoparticle of the present application contains manganese dioxide nanoparticles, polyethylene glycol, poly-caprolactone, and a sonophoto sensitizer, with manganese dioxide nanoparticles as the substrate, poly-caprolactone forming a hydrophobic core to load the sonophoto sensitizer.

[0024] In a third aspect, the present application provides the use of the sonophoto sensitizer@manganese dioxide nanoparticle, for example, for preparing a contrast agent.

[0025] The prepared sound / light sensitizer MnO2 nanoparticles have broad application prospects in the field of disease diagnosis and treatment. In the treatment of Helicobacter pylori and other bacterial infections, the sound / light sensitizer can be delivered to the target through targeting, and the active oxygen (ROS) generated by the synergistic activation of ultrasound and light can kill bacteria efficiently. The test results of the present application show that the prepared sound / light sensitizer MnO2 nanoparticles have excellent antibacterial activity.

[0026] Meanwhile, the manganese ions generated by the degradation of MnO2 can be used as a magnetic resonance imaging (MRI) contrast agent to realize accurate diagnosis of the infection site and real-time evaluation of the treatment effect.

[0027] In tumor treatment, the nanoparticles not only have synergistic anti-tumor effect through sonodynamic and photodynamic therapy, but also can improve the hypoxic microenvironment of tumors through MnO2 to enhance the sensitivity of chemotherapy and radiotherapy, and realize real-time monitoring during treatment by combining with the MRI contrast function.

[0028] The sound / light sensitizer MnO2 nanoparticles of the present application are also used for real-time monitoring of the to-be-measured index, including the whole process of tumor treatment.

[0029] The present application has the following outstanding advantages: Polyethylene glycol (PEG) endows the nanoparticles with excellent water solubility and biocompatibility, and polycaprolactone (PCL) forms a hydrophobic core. The sound / light sensitizer MnO2 nanoparticles of the present application cooperate with each other to efficiently load the liposoluble organic sound / light sensitizer through hydrophobic interaction, realize stable encapsulation and targeted delivery of drugs, and avoid the drawbacks of traditional drug preparations, such as easy aggregation in vivo and low bioavailability. The present application can be used to construct a "diagnosis-treatment-treatment effect evaluation" integrated diagnosis and treatment platform, provide a new technical solution for clinical precision medicine, and effectively improve the precision and effectiveness of disease treatment. BRIEF DESCRIPTION OF DRAWINGS

[0030] In order to more clearly illustrate the technical solutions of the present application, the following will briefly introduce the drawings needed in the embodiments. Obviously, each drawing in the following description is directed to some embodiments of the present application, and those skilled in the art can also obtain other drawings without creative labor on the basis of these drawings.

[0031] Figure 1 The ultraviolet-visible absorption spectrum of the Ce6@MnO2 nanoparticles prepared in Example 1 of the present application.

[0032] Figure 2 The anti-Helicobacter pylori activity of the nanoparticles prepared in the present application under ultrasound mediation. DETAILED DESCRIPTION

[0033] The technical solution will be clearly and completely described below through embodiments of this application. Obviously, the described embodiments are only some preferred embodiments of this application, and not all embodiments. Based on the embodiments of this application, all other embodiments obtained by those of ordinary skill in the art without creative effort are within the protection scope of this application.

[0034] Example 1 A type of manganese dioxide nanoparticle loaded with Ce6 photosensitizer was prepared by precisely synthesizing a high-purity MnO2 nano-substrate using a potassium permanganate reduction method. The amphiphilic polymer polyethylene glycol-polycaprolactone (PEG-PCL) was modified onto the MnO2 nano-surface, comprising: 1 part potassium permanganate, 1.2 parts reducing agent polyallylamine hydrochloride, and 2 parts PEG-PCL, prepared according to the following steps: 1) Slowly add 1 part of potassium permanganate aqueous solution to 1.2 parts of polyallylamine hydrochloride solution, disperse by ultrasonication, adjust the pH value to 7, react at 25℃ for 4 hours, the solution changes from purple-red to brown-black, and MnO2 nano-substrate is obtained. 2): Add 2 parts of polyethylene glycol-polycaprolactone, stir at room temperature, and react for 12 hours to obtain a solution of polyethylene glycol-polycaprolactone (PEG-PCL) modified on the MnO2 nano surface; 3): Dissolve 10% (by mass) of the photosensitizer dihydroporphyrin E6 (Ce6) in dimethyl sulfoxide, add it to the solution from step 2), and stir to obtain a mixed solution; Step (4): Dialyze the mixture obtained in step 3) overnight with water, centrifuge to remove unreacted material, and collect the MnO2 nanoparticles loaded with photosensitizer, Ce6@MnO2.

[0035] The UV-Vis absorption spectrum of the prepared Ce6@MnO2 nanoparticles is shown in the figure below. Figure 1 As shown.

[0036] Example 2 A manganese dioxide nanoparticle loaded with Ce6 photosensitizer was prepared similarly to that in Example 1, except that the reducing agent in step 1) was oxalic acid, the pH was adjusted to 3, the reaction temperature was 45 °C, and the reaction time was 6 hours. The rest of the reaction was the same as in Example 1, yielding sheet-like nanoparticles with a specific surface area of ​​150-200 m². 2 / g, the prepared nanoparticles were loaded with 20% Ce6 by mass of PEG-PCL, dialyzed with water overnight, centrifuged to obtain Ce6@MnO2, and then proceeded with the following steps: 1) Slowly add 1 part of potassium permanganate aqueous solution to 1.2 parts of oxalic acid solution, disperse by ultrasonication, adjust the pH to 3, and react at 45℃ for 6 hours. The solution changes from purplish-red to brownish-black, yielding sheet-like nanoparticles with a specific surface area of ​​150-200 m².2 / g; 2): Add 2 parts of polyethylene glycol-polycaprolactone, stir at room temperature, and react for 12 hours to obtain a solution of polyethylene glycol-polycaprolactone (PEG-PCL) modified on the MnO2 nano surface; 3): Dissolve 20% (by mass) of photosensitizer Ce6 in dimethyl sulfoxide, add it to the solution from step 2), and stir to obtain a mixed solution; Step (4): Dialyze the mixture obtained in step 3) overnight with water, centrifuge to remove unreacted material, and collect the MnO2 nanoparticles loaded with photosensitizer, Ce6@MnO2.

[0037] Example 3 A manganese dioxide nanoparticle loaded with indocyanine green (ICG) was prepared similarly to that in Example 1, except that in step 2), 5 parts of PEG-PCL solution were added, and the mixture was stirred at room temperature for 6 h. The resulting nanoparticles were loaded with 20% (by mass) of indocyanine green (ICG) from PEG-PCL. ICG@MnO2 was then prepared according to the following steps: 1) Slowly add 1 part of potassium permanganate aqueous solution to 1.2 parts of polyallylamine hydrochloride solution, disperse by ultrasonication, adjust the pH value to 7, react at 25℃ for 4 hours, the solution changes from purple-red to brown-black, and MnO2 nano-substrate is obtained. 2): Add 5 parts of polyethylene glycol-polycaprolactone, stir at room temperature, and react for 6 hours to obtain a solution of polyethylene glycol-polycaprolactone (PEG-PCL) modified on the MnO2 nano surface; 3): Dissolve 20% (by mass) of indocyanine green (ICG) in dimethyl sulfoxide, add it to the solution from step 2), and stir to obtain a mixed solution; Step (4): Dialyze the mixture obtained in step 3) overnight with water, centrifuge to remove unreacted substances, and collect ICG@MnO2.

[0038] Example 4 Manganese dioxide nanoparticles loaded with protoporphyrin (PPIX) were prepared similarly to those in Example 1, except that the reducing agent in step 1) was sodium sulfite, the pH was adjusted to 6, the reaction temperature was 35 °C, and the reaction time was 8 hours. The rest was the same as in Example 1. Rod-shaped nanoparticles were obtained. The prepared nanoparticles were loaded with 15% by mass of PEG-PCL protoporphyrin (PPIX), dialyzed with water overnight, and centrifuged to obtain PPIX@MnO2.

[0039] 1) Slowly add 1 part of potassium permanganate aqueous solution to 1.2 parts of sodium sulfite solution, disperse by ultrasonication, adjust the pH value to 6, react at 35℃ for 8 hours, the solution changes from purple-red to brown-black, and rod-shaped MnO2 nanoparticles are obtained. 2) Add 2 parts of polyethylene glycol-polycaprolactone, stir at room temperature, and react for 12 hours to obtain a solution of polyethylene glycol-polycaprolactone (PEG-PCL) modified on the MnO2 nano surface; 3) Dissolve 15% of protoporphyrin (PPIX) by mass of PEG-PCL in dimethyl sulfoxide, add it to the solution in step 2), and stir to obtain a mixed solution; Step (4): Dialyze the mixture obtained in step 3) overnight with water, centrifuge to remove unreacted material, and collect the MnO2 nanoparticles loaded with PPIX, PPIX@MnO2.

[0040] Example 5 Anti-Helicobacter pylori test: with a Helicobacter pylori growth concentration of 10 6 Each milliliter of bacterial colony constitutes a sample. 30 μL of this sample was co-incubated with the nanoparticles prepared in Examples 1-4, with a sonication agent concentration of 5 g / mL. The ultrasound-mediated conditions were 1 MHz and 1.5 W / cm². -2 The antibacterial rate was calculated using the plate method.

[0041] The results are as follows Figure 2 As shown, the nanoparticles prepared in Examples 1-4 all exhibit antibacterial properties, with values ​​of approximately 86%, 100%, 70%, and 81%, respectively. The overall antibacterial rate exceeds 60%, with Example 2 showing the highest and most stable antibacterial rate.

[0042] In the description of this application, the terms "comprising," "including," or any other variations thereof are intended to cover a non-exclusive inclusion, such that a process, method, article, or apparatus that comprises a list of elements includes not only those elements but also other elements not expressly listed, or elements inherent to such a process, method, article, or apparatus. Without further limitation, an element defined by the phrase "comprising a reference structure" does not exclude the presence of other identical elements in the process, method, article, or apparatus that includes said element.

[0043] It should be noted that in this article, relational terms such as "first" and "second" are used only to distinguish one entity or operation from another, and do not necessarily require or imply any such actual relationship or order between these entities or operations.

[0044] The embodiments described above are merely specific implementations of this application, but the scope of protection of this application is not limited thereto. Any variations or substitutions that can be conceived by those skilled in the art within the scope of the technology disclosed in this application without creative effort should be included within the scope of protection of this application. Therefore, the scope of protection of this application should be determined by the scope of protection of the claims in this application.

Claims

1. A method for preparing acoustic / photosensitizer@manganese dioxide nanoparticles, characterized in that, High-purity MnO2 nano-substrate was precisely synthesized using the potassium permanganate reduction method. The amphiphilic polymer polyethylene glycol-polycaprolactone (PEG-PCL) was then used to modify the MnO2 nano-surface, including the following steps: Step (1): Slowly add 1 part by weight of potassium permanganate aqueous solution into 1 to 3 parts of reducing agent solution. After ultrasonic dispersion, adjust the pH value to 2 to 7 and react in the range of 25 to 60 °C for 4 to 24 hours. The solution changes from purple-red to brown-black. Neutralize the solution to neutral to obtain MnO2 nano-substrate. Step (2): Add 1 to 5 parts of polyethylene glycol-polycaprolactone, stir at room temperature, and react for 6 to 12 hours to obtain a solution of polyethylene glycol-polycaprolactone (PEG-PCL) modified on the MnO2 nano surface; Step (3): Dissolve 10% to 20% of the PEG-PCL mass of the sound sensitizer or photosensitizer in dimethyl sulfoxide, add it to the solution in step (2) and stir to obtain a mixed solution; Step (4): Dialyze the mixture obtained in step (3) with water overnight, centrifuge to remove unreacted substances, and collect MnO2 nanoparticles loaded with acoustic or photosensitizers, i.e., acoustic / photosensitizer@manganese dioxide nanoparticles.

2. The method for preparing acoustic / photosensitizer@manganese dioxide nanoparticles according to claim 1, characterized in that, In step (1), the reducing agent is selected from oxalic acid, sodium sulfite, and polyallylamine hydrochloride; the solution is neutralized to neutral using an alkaline solution.

3. The method for preparing acoustic / photosensitizer@manganese dioxide nanoparticles according to claim 1, characterized in that, In step (3), the acoustic / photosensitizer is selected from protoporphyrin, dihydroporphyrin E6, or indocyanine green.

4. The method for preparing acoustic / photosensitizer@manganese dioxide nanoparticles according to claim 1, characterized in that, In step (2), the average molecular weight of PEG-PCL is 7000.

5. The method for preparing acoustic / photosensitizer@manganese dioxide nanoparticles according to claim 1, characterized in that, In step (1), the amount of reducing agent used is 1.2 to 2 parts, the pH value is adjusted to 3 to 6 after ultrasonic dispersion, the reaction temperature is controlled in the range of 35 to 45 ℃, and the reaction time is 6 to 12 hours; In step (2), the amount of polyethylene glycol-polycaprolactone used is 2 to 5 parts, and the reaction temperature is 15℃ to 30℃; In step (3), the amount of acoustic / photosensitizer used is 15% to 20% of the mass of polyethylene glycol-polycaprolactone.

6. The method for preparing acoustic / photosensitizer@manganese dioxide nanoparticles according to claim 1, characterized in that, In step (1), the reaction is carried out for 8 to 10 hours; the solution is neutralized with sodium hydroxide until it is neutral.

7. A sound / photosensitizer@manganese dioxide nanoparticles, characterized in that, The method for preparing the acoustic / photosensitizer@manganese dioxide nanoparticles contains manganese dioxide nanoparticles, polyethylene glycol, polycaprolactone, and an acoustic or photosensitizer. The manganese dioxide nanoparticles serve as the substrate, with polycaprolactone forming a hydrophobic core and the acoustic or photosensitizer loaded onto it.

8. The acoustic / photosensitizer@manganese dioxide nanoparticles according to claim 7, characterized in that, The aforementioned acoustic / photosensitizer@manganese dioxide nanoparticles are obtained using the preparation method described in any one of claims 1-6, wherein polyethylene glycol, as an amphiphilic polymer, endows the nanoparticles with excellent water solubility and biocompatibility.

9. The application of the acoustic / photosensitizer@manganese dioxide nanoparticles according to any one of claims 7-8, characterized in that, The aforementioned acoustic / photosensitizer@manganese dioxide nanoparticles are used to prepare contrast agents.

10. The application of the acoustic / photosensitizer@manganese dioxide nanoparticles according to any one of claims 7-8, characterized in that, The aforementioned acoustic / photosensitizer@manganese dioxide nanoparticles are used for real-time monitoring of the target indicator.

Images