Polymer / active gas nanobubble, preparation method and application
By preparing polymer/active gas nanobubbles, the limitations of high-frequency focused ultrasound tumor ablation technology are overcome, uniform ablation of tumor tissue and synergistic treatment of active gas are achieved, and the efficiency and safety of tumor treatment are improved.
Patent Information
- Application Number
- CN202510978367.9
- Authority / Receiving Office
- CN · China
- Patent Type
- Applications(China)
- Current Assignee / Owner
- Filing Date
- 2025-07-16
- Publication Date
- 2025-09-16
AI Technical Summary
Existing high-frequency focused ultrasound tumor ablation technology has problems such as incomplete tumor ablation, damage to normal tissue and single treatment method. Nanobubble systems have problems such as insufficient stability, uneven distribution and uncoordinated active gas delivery in tumor treatment.
Polymer/active gas nanobubbles are used to form stable core-shell structure nanobubbles through the steps of preparing polymer mixed liquid, emulsification, centrifugal freeze-drying, etc., combined with low-frequency ultrasound to trigger gas release, to achieve uniform distribution and synergistic treatment of tumor tissue.
Without damaging normal tissues, it significantly improves the tumor ablation effect, enhances tumor coverage and treatment depth, and achieves uniform distribution of active gases and synergistic treatment functions.
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Figure CN120643692A_ABST
Abstract
Description
Technical Field
[0001] The present invention belongs to the field of biomedicine technology, and in particular relates to polymer / active gas nanobubbles, a preparation method and applications. Background Art
[0002] High-frequency focused ultrasound (HIFU) is a non-invasive tumor ablation technique that achieves ablation of lesions by precisely focusing ultrasound energy on a target area, generating thermal and cavitation effects. This technique has been widely used in the treatment of solid tumors such as uterine fibroids and prostate cancer. Compared with traditional surgery, its significant advantages include avoiding the risk of ionizing radiation and eliminating the need for open surgical incisions. However, existing technologies still have several limitations that restrict their clinical effectiveness. First, the efficiency of thermal ablation is highly dependent on the uniformity of biological tissue. For heterogeneous tumors with abundant blood flow, the focused ultrasound beam is susceptible to energy gradient attenuation due to reflections from tissue interfaces, ultimately resulting in incomplete tumor ablation and the pathological basis for local recurrence. Second, nonspecific thermal diffusion of high-frequency focused ultrasound into tissues outside the focal zone can damage adjacent normal tissues. Third, the ultrasound physical ablation killing mode lacks drug synergy, and single-agent treatments are limited in their biological efficacy. Although microbubble-mediated ultrasound targeted therapy has recently opened up new avenues for synergistic treatment, conventional microbubbles suffer from insufficient stability and are prone to rupture and dissipation. Furthermore, their large particle size makes it difficult to achieve uniform distribution within the high-pressure areas of the tumor interstitium, severely limiting treatment efficiency. Therefore, the design and construction of a nanobubble combined ultrasound treatment system with high stability, uniformly distributed treatment and synergistic treatment capabilities has become an urgent need to break through the bottleneck of ultrasound ablation technology.
[0003] In recent years, nanobubble systems based on active gas loading have demonstrated unique advantages in the field of tumor treatment. Studies have confirmed that active small molecule gases such as nitric oxide (NO), ozone (O3) and hydrogen (H2) can effectively improve the mechanical microenvironment of tumors by regulating the pressure of interstitial fluid in tumor tissues and reshaping the extracellular matrix structure, thereby significantly improving the penetration efficiency of therapeutic carriers in solid tumors. Compared with traditional gas preparations (such as inhaled NO or micron-level ultrasound contrast agents), the new generation of nanobubble systems (<500nm) can achieve targeted enrichment effects in specific areas due to their nanoscale particle size characteristics and acoustic response characteristics, and can use low-frequency ultrasound (1-3MHz) to trigger precise gas release, reducing the risk of exposure to active gases.
[0004] Although studies have attempted to combine nanogas delivery systems with low-frequency ultrasound ablation technology, existing combined treatment systems still have three key issues: First, traditional micron-sized bubbles (>1μm) have low tumor accumulation efficiency due to particle size limitations (EPR effect contribution rate <15%), and rely on high-frequency ultrasound (>5MHz) to trigger gas release, resulting in insufficient tissue penetration depth (<3cm). Conventional lipid / protein-based nanobubble shell materials, while reducing particle size (<500nm), have insufficient Young's modulus (<100MPa) of the shell material, leading to premature rupture under low-frequency ultrasound (1-2MHz) (carrier gas loss rate >40% / h). This makes it difficult to achieve both a long circulation half-life and carrier gas stability, and results in low gas loading efficiency. Second, existing nanobubbles often use inert gases (such as SF6 / C3F8) as cavitation nuclei. While capable of producing a mechanical ablation effect, they are unable to regulate the interstitial pressure (IFP) of the tumor (maintaining it within ±5% of the baseline level), resulting in heterogeneous distribution of the therapeutic carrier within the tumor (with a coefficient of dispersion > 0.5). Third, the spatiotemporal synergy between the delivery of reactive gases (such as NO / CO) and ultrasound energy has not yet been established. High-frequency ultrasound (3MHz) triggers the release of gases, resulting in an uncontrolled gas diffusion range (effective radius < 200μm). This mismatch between the ultrasound energy distribution and the spatial positioning of the bubbles creates a therapeutic blind spot. These deficiencies collectively lead existing technologies into a vicious cycle of low delivery efficiency, weak microenvironmental regulation, and poor energy utilization. Summary of the Invention
[0005] In view of the shortcomings of the prior art, the purpose of the present invention is to provide a polymer / active gas nanobubble, a preparation method and an application, which solve the problems in the prior art.
[0006] The purpose of the present invention can be achieved through the following technical solutions:
[0007] A method for preparing polymer / active gas nanobubbles comprises the following steps:
[0008] S1, mixing polymer powder with an organic solvent to obtain a polymer mixture;
[0009] S2, adding an emulsifier and an aqueous phase to the polymer mixture, and obtaining a primary emulsion by crushing;
[0010] S3, adding a polymer solution and an emulsifier of a certain concentration to the primary emulsion, and emulsifying to obtain a secondary emulsion;
[0011] S4, transferring the double emulsion to a polymer solution of another concentration, stirring and centrifuging, adding a freeze-drying protectant, and packaging and freeze-drying to obtain a freeze-dried powder of the polymer nanomembrane shell;
[0012] S5, taking the freeze-dried powder into a bottle, filling it with active gas, then re-dissolving it with a solvent, and shaking it to obtain polymer / active gas nanobubbles.
[0013] Furthermore, the polymer in S1 is PLGA or PLLA; the polymers in S3 and S4 are PVA or PAM.
[0014] Furthermore, the organic solvent is anhydrous ethanol, dichloromethane or chloroform.
[0015] Furthermore, the emulsifier in S2 is Span 60, Span 65 or Span 80; the emulsifier in S3 is Tween 60, Tween 65 or Tween 80.
[0016] Furthermore, the concentrations of the polymer solutions in S3 and S4 are 5% w / v and 0.5% w / v, respectively.
[0017] Furthermore, the lyoprotectant is mannitol with a concentration of 10% w / v.
[0018] Furthermore, the active gas is: H2, H2S, CO, NO, O2, O3, Xe or SO2.
[0019] A polymer / active gas nanobubble is prepared using the above preparation method.
[0020] A tumor ablation system includes an ultrasound device and an injection device. The injection device includes the above-mentioned polymer / active gas nanobubbles. Under ultrasound stimulation, the polymer / active gas nanobubbles can ablate tumors.
[0021] The application of the above-mentioned polymer / active gas nanobubbles in the preparation of tumor therapeutic drugs and tumor diagnostic kits.
[0022] Beneficial effects of the present invention:
[0023] 1. Compared with other types of nanoscale drug carriers, the polymer nanobubbles of the present invention have good stability, are simple and easy to prepare, are suitable for large-scale production, and have broad application prospects in practical applications;
[0024] 2. Compared with traditional ultrasonic ablation of tumors, the polymer nanobubbles combined with ultrasonic ablation of tumors treatment system of the present invention can have a significant therapeutic effect on tumor tissue without causing thermal damage to normal tissue.
[0025] 3. Compared to polymer nanobubbles loaded with inert gas, the introduction of active gas in this invention promotes uniform distribution of the polymer / active gas nanobubbles within tumor tissue, enhancing overall tumor coverage and ablation. Inert gas bubbles are generally less uniformly distributed within tissue than active gas. Furthermore, upon rupture, active gas bubbles can release therapeutically active molecules, further enhancing the overall therapeutic effect of tumors. In contrast, rupture of inert gas bubbles only produces a physical effect, resulting in a relatively limited therapeutic effect.
[0026] 4. The polymer / active gas nanobubbles of the present invention use polymer as the membrane material to construct nano-microcapsules with good stability and load bioactive gases. By combining with ultrasonic stimulation, on the one hand, the polymer / active gas nanobubbles can be used for tumor ultrasonic imaging, and on the other hand, the polymer / active gas nanobubbles can be induced to produce a cavitation effect, thereby achieving the synergistic therapeutic functions of mechanically destroying tumor cells and extracellular matrix, promoting the diffusion and penetration of active gases in the tumor interstitium, and enhancing the secondary distribution uniformity of nanocarriers. The active gas diffusion and nanocarrier redistribution constitute a dynamic synergistic mechanism, which reduces the interstitial fluid pressure caused by the tumor matrix and increases the subsequent enrichment concentration of nanobubbles in the lesion area, thereby forming a positive feedback loop that enhances the depth and range of cavitation ablation. BRIEF DESCRIPTION OF THE DRAWINGS
[0027] In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the following briefly introduces the drawings required for use in the embodiments or the description of the prior art. Obviously, for ordinary technicians in this field, other drawings can be obtained based on these drawings without any creative work.
[0028] Figure 1 is a transmission electron micrograph of the polymer / active gas nanobubbles of Example 1;
[0029] Figure 2 1 is a particle size distribution diagram and zeta potential diagram of the polymer / active gas nanobubbles of Example 1 and the polymer nanocapsules of Comparative Example 1;
[0030] Figure 3 1 is an in vitro ultrasound imaging effect and quantitative analysis diagram of the polymer / active gas nanobubbles of Example 1;
[0031] Figure 4 This is a graph showing the killing effect of the polymer / active gas nanobubbles of Example 1 on glioma cells GL261 at different concentrations;
[0032] Figure 5 This is a diagram showing the cumulative effect and intratumoral distribution of the polymer / active gas nanobubbles combined with ultrasound in an in vivo subcutaneous tumor model of Example 1;
[0033] Figure 6 This is a histological section analysis diagram of the therapeutic effect of polymer / active gas nanobubbles combined with ultrasound on subcutaneous tumors in vivo in Example 1. DETAILED DESCRIPTION
[0034] The following will clearly and completely describe the technical solutions in the embodiments of the present invention in conjunction with the accompanying drawings. Obviously, the described embodiments are only part of the embodiments of the present invention, not all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without making any creative efforts shall fall within the scope of protection of the present invention.
[0035] A method for preparing polymer / active gas nanobubbles comprises the following steps:
[0036] S1, taking polymer powder, adding organic solvent, and ultrasonically mixing to obtain a polymer mixture;
[0037] S2, adding an emulsifier and an aqueous phase to the polymer mixture, and crushing it by an ultrasonic crusher to obtain a primary emulsion;
[0038] S3, adding a polymer solution and an emulsifier of a certain concentration to the primary emulsion, and performing secondary emulsification under an ultrasonic crusher to obtain a secondary emulsion;
[0039] S4, transferring the double emulsion to a polymer solution of another concentration, stirring the solution by a homogenizer, centrifuging the stirred solution after a certain period of time, adding a freeze-drying protective agent, and performing freeze-drying in aliquots to obtain a freeze-dried powder of the polymer nanomembrane shell;
[0040] S5, taking the freeze-dried powder into a bottle, filling it with active gas, then re-dissolving it with a solvent, shaking it, and finally obtaining polymer / active gas nanobubbles.
[0041] In S1, the polymer is PLGA or PLLA; the organic solvent is anhydrous ethanol, dichloromethane or chloroform; preferably PLGA or dichloromethane;
[0042] In S2, the emulsifier is: Span 60, Span 65 or Span 80; the aqueous phase is: pure water, PBS or normal saline; preferably Span 80 and pure water;
[0043] In S3, the polymer is PVA or PAM, and the emulsifier is Tween 60, Tween 65 or Tween 80; preferably 5% PVA and Tween 80
[0044] In S4, the polymer is PVA or PAM, preferably 0.5% PVA.
[0045] In S3, the concentration of the polymer solution was 5% w / v;
[0046] In S4, the concentration of the polymer solution is 0.5% w / v; the freeze-drying protective agent is mannitol, the concentration of which is 10% w / v; wherein, freeze drying is performed in a freeze dryer, and the freeze drying time is 48-72 hours;
[0047] In S5, the active gas is: H2, H2S, CO, NO, O2, O3, Xe or SO2; the solvent is: pure water, PBS or physiological saline.
[0048] In the following examples, the experimental methods used are conventional methods unless otherwise specified. The materials and reagents used are all commercially available unless otherwise specified. Experimental methods where specific conditions are not specified in the examples are generally performed under conventional conditions or according to the conditions recommended by the manufacturer.
[0049] Example 1
[0050] A method for preparing polymer / active gas nanobubbles (NO-PLGA NBs) comprises the following steps:
[0051] (1) Place 200 mg of PLGA in a vial, add 2 mL of dichloromethane, and fully dissolve in an ultrasonic water bath to obtain a PLGA polymer solution;
[0052] (2) 400 μL of ultrapure water and 100 μL of Tween 80 were added to the PLGA polymer solution, and the mixture was emulsified for 60 s under an ultrasonic cell disruptor with a power of 400 W to obtain a primary emulsion. 5 mL of a 5% (w / v) PVA solution and 100 μL of Tween 80 were added to the primary emulsion, and the mixture was emulsified for 60 s under the same power to obtain a water-in-oil-in-water emulsion.
[0053] (3) The emulsion was transferred to 20 mL of 0.5% (w / v) PVA solution and homogenized at 7200 rpm for 120 min at 25°C using a high-performance homogenizer. The mixture was then centrifuged at 3000 rpm, the supernatant was retained, and the mixture was centrifuged at 7000 rpm, the supernatant was discarded, and the mixture was reconstituted with 5 mL of pure water. The mixture was then packaged and freeze-dried for 48 h to obtain freeze-dried PLGA polymer nanobubble microcapsules.
[0054] (4) 4 mg of the lyophilized sample was placed in a vial and slowly filled with 10 mL of NO. After the filling was complete, the vial was reconstituted with 2 mL of saline and shaken at 50 Hz for 60 s. Thus, polymer / active gas nanobubbles with PLGA as the membrane shell and NO loaded at a concentration of 2 mg / mL were prepared.
[0055] Among them, the transmission electron microscopy analysis of the polymer / active gas nanobubbles obtained in Example 1 showed that Figure 1As shown; it can be seen that the polymer / active gas nanobubbles obtained in Example 1 present a typical core-shell structure, with a high electron density polymer shell (film thickness 18.5±2.3nm) in sharp contrast to a low electron density gas core (diameter 165.4±12.7nm) ( Figure 1 In addition, transmission electron microscopy imaging results show that the polymer / active gas nanobubbles have uniform morphology and good consistency ( Figure 1 b) in the above example.
[0056] Comparative Example 1
[0057] A polymer nanocapsule (PLGA NBs) is not loaded with any type of gas compared to Example 1. The specific preparation method includes the following steps:
[0058] (1) 200 mg of PLGA was placed in a vial, 2 mL of dichloromethane was added, and after ultrasonic dissolution, 400 mL of ultrapure water and 100 mL of Tween 80 were added again. The mixture was emulsified for 60 s in an ultrasonic cell disruptor at 400 W power to obtain a primary emulsion. 5 mL of a 5% (w / v) PVA solution and 100 mL of Tween 80 were added to the primary emulsion and emulsified for 60 s at the same power to obtain a water-in-oil-in-water emulsion.
[0059] (2) The emulsion was transferred to 20 mL of 0.5% (w / v) PVA solution and homogenized at 7200 rpm for 120 min at 25°C using a high-performance homogenizer. The mixture was then centrifuged at 3000 rpm, the supernatant was retained, and the mixture was centrifuged at 7000 rpm, the supernatant was discarded, and the mixture was reconstituted with 5 mL of pure water. The mixture was then packaged and freeze-dried for 48 h to obtain freeze-dried PLGA polymer nanobubble microcapsules.
[0060] (3) 4 mg of the lyophilized sample was placed in a vial, reconstituted with 2 mL of saline, and then shaken at 50 Hz for 60 s. Thus, a 2 mg / mL polymer nanocapsule with a PLGA membrane shell and no gas loaded inside was prepared.
[0061] like Figure 2 a and b in the equation represent particle size distribution and Zeta potential, respectively. Figure 2 As can be seen from a in the figure: Dynamic light scattering characterization shows that the hydrodynamic particle size of the NO-PLGA NBs prepared in Example 1 is 188.38±10 nm (PDI=0.002), which is higher than that of the PLGA NBs prepared in Comparative Example 1 without carrier gas (182.70±10 nm (PDI=0.153), indicating that the PLGA microcapsules were successfully loaded with NO active gas under the steps of Example 1. Figure 2As can be seen in b: Surface charge analysis shows that the Zeta potential of NO-PLGA NBs is (-19.57±2.5) mV, which is close to the Zeta potential of PLGA NBs -(21.43±2) mV. This surface charge characteristic can effectively prevent particle aggregation during storage.
[0062] Example 2
[0063] In this example, we investigated the in vitro ultrasound imaging performance of polymer / active gas nanobubbles. We selected the polymer / active gas nanobubbles prepared in Example 1 and evaluated their in vitro ultrasound imaging enhancement using a standard agarose membrane model. The specific process was as follows:
[0064] A 1.5 mL sample of polymer / active gas nanobubble was added to a preformed well in the agarose membrane. Multimodal ultrasound diagnostics were performed using a 21 MHz high-frequency probe. Dual-mode ultrasound imaging was acquired at 0, 2, 4, 6, and 8 minutes after sample injection using both B-Mode and contrast-enhanced imaging, with simultaneous quantitative signal analysis.
[0065] Among them, the effects of in vitro ultrasound imaging are as follows Figure 3 As shown in a, it can be seen that the polymer / active gas nanobubbles exhibit a characteristic ultrasound enhancement effect in the body membrane model, forming a significant acoustic contrast difference. Figure 3 As shown in Figure (b), the B-Mode peak signal intensity reaches 800,000 a.u., with a half-life of approximately 2 minutes. The Contrast-Mode signal intensity reaches a maximum of approximately 27,500 a.u., with a half-life of approximately 4 minutes. This data confirms that the core-shell structure exhibits excellent ultrasonic response characteristics while maintaining gas stability.
[0066] Example 3
[0067] In this embodiment, the evaluation of the ultrasound-enhanced killing effect of the polymer / active gas nanobubble combined with ultrasound treatment system on glioma cells in vitro includes the following steps:
[0068] (1) GL261 glioma cells were cultured at 1×10 4 Cells were seeded in 96-well plates and cultured for 48 hours to form a monolayer. Then, an untreated control group (Group A) and a nanobubble combined with ultrasound irradiation treatment group (Group BI, nanobubble concentration gradient: 0.4-1.8 mg / mL, Δ=0.2 mg / mL, n=6) were set up.
[0069] (2) Combined treatment: Nanobubble solutions (PBS containing 10% FBS) of different concentrations were incubated with cells for 30 minutes, followed by ultrasound treatment (1 MHz, 1.0 W / cm 2 , duty cycle 50%, duration 5 minutes).
[0070] The cell viability was determined using the CCK-8 assay. Figure 4 The results showed that the nanobubble + ultrasound combined treatment group significantly inhibited the activity of GL261 glioma cells. When the concentration of polymer / active gas nanobubbles reached 1.0 mg / mL, the cell viability decreased by 65.8% compared with the control group. The IC 50 =0.98 mg / mL. Concentration-dependent analysis showed that the killing efficiency was positively correlated with the intensity of the ultrasonic cavitation effect, demonstrating that nanobubbles significantly increased cell membrane permeability through the sonoporation effect, enhancing the cell-killing effect of polymer / active gas nanobubbles.
[0071] Example 4
[0072] In this example, the in vivo near-infrared fluorescence imaging characterization of the polymer / active gas nanobubble therapeutic system was studied, including the following steps:
[0073] (1) Construction of subcutaneous tumor animal model and its grouping treatment: BALB / c mice were selected and injected subcutaneously in the right groin with 5×10 6 GL261 glioma cells were used to construct a subcutaneous tumor model. Seven days after surgery, the tumor volume reached 100-150 mm. 3 The tumor-bearing mice were randomly divided into three groups (n=6): Group A (negative control, tail vein injection of 0.9% NaCl 200mL), Group B (single ultrasound treatment group, 1MHz / 1.0W / cm 2 =The results were published in the Journal of Clinical Oncology. Groups C (single nanobubble treatment group, intratumoral injection of 200 mL of a 1 mg / mL polymer / active gas nanobubble solution) and D (combination treatment group, intratumoral injection of 200 mL of a 1 mg / mL polymer / active gas nanobubble solution, followed 30 minutes later by ultrasound with the same parameters as in Group B) were included. All groups were treated for three consecutive days. Tumor tissues were completely removed and fixed in 4% paraformaldehyde solution after euthanasia on the fourth day.
[0074] (2) A small animal near-infrared fluorescence imaging system was used to perform in vitro fluorescence imaging of mouse tumor tissues. The penetration efficiency of polymer / active gas nanobubble combined with ultrasound therapy was evaluated by analyzing the distribution of near-infrared fluorescence in tumor tissues.
[0075] The results are as follows Figure 5In vivo near-infrared fluorescence imaging of the polymer / active gas nanobubble combined with ultrasound treatment group (Group D) showed that the fluorescence signal intensity in the tumor area increased by about 3 times compared with the saline control group (Group A), and showed good uniform distribution. This significant difference indicates that ultrasound irradiation promotes the directional rupture of nanobubbles through the cavitation effect, effectively degrading tumor extracellular matrix components (such as collagen and laminin), thereby increasing the penetration depth of therapeutic drugs in tumor tissue. This result fully verifies the technical advantage of polymer / active gas nanobubbles in significantly enhancing the penetration efficiency of solid tumor drugs through a dual physical-chemical mechanism of action under the synergistic effect of ultrasound.
[0076] Example 5
[0077] In this embodiment, the polymer / active gas nanobubble therapy system is used to evaluate the efficacy of ultrasound ablation of subcutaneous tumors. The specific steps include
[0078] (1) Construction of subcutaneous tumor animal model and its grouping treatment: BALB / c mice were selected and injected subcutaneously in the right groin with 5×10 6 GL261 glioma cells were used to construct a subcutaneous tumor model. Seven days after surgery, the tumor volume reached 100-150 mm. 3 The tumor-bearing mice were randomly divided into three groups (n=6): Group A (negative control group, tail vein injection of 0.9% NaCl 200mL), Group B (single ultrasound treatment group, 1MHz / 1.0W / cm 2 =The results were published in the Journal of Clinical Oncology. Groups C (single nanobubble treatment group, intratumoral injection of 200 mL of a 1 mg / mL polymer / active gas nanobubble solution) and D (combination treatment group, intratumoral injection of 200 mL of a 1 mg / mL polymer / active gas nanobubble solution, followed 30 minutes later by ultrasound with the same parameters as in Group B) were included. All groups were treated for three consecutive days. Tumor tissues were completely removed and fixed in 4% paraformaldehyde solution after euthanasia on the fourth day.
[0079] (2) Pathological evaluation: After treatment, the mice were killed and the tumor tissues were removed for H&E staining and Picrosirius Red staining to evaluate the potential damage caused by ultrasonic cavitation, including the changes in tumor necrosis area and tumor fibrosis area.
[0080] The results are as follows Figure 6Quantitative analysis of H&E staining showed that the area of tumor core necrosis in Group D reached (82.3±5.7)%, which was 6.8 times, 4.4 times, and 2.4 times the area of tumor core necrosis in Group A (12.1±3.2)%, Group B (18.5±4.1)%, and Group C (34.6±6.8), respectively. Picrosirius Red analysis showed that the area of peritumoral fibrosis in Group D was (7.2±1.3)%, significantly lower than the (19.8±2.1)% in Group B (P<0.01). These results demonstrate that polymer / reactive gas nanobubbles combined with ultrasound can effectively destroy the tumor extracellular matrix and achieve uniform distribution of polymer / reactive gas nanobubbles within the tumor tissue by reducing the degree of fibrosis in the tumor tissue, further enhancing the efficacy of ultrasound-assisted tumor ablation.
[0081] Throughout this specification, references to terms such as "one embodiment," "example," or "specific example" indicate that the specific features, structures, materials, or characteristics described in conjunction with that embodiment or example are included in at least one embodiment or example of the present invention. In this specification, schematic representations of these terms do not necessarily refer to the same embodiment or example. Furthermore, the specific features, structures, materials, or characteristics described may be combined in any suitable manner in any one or more embodiments or examples.
[0082] The basic principles, main features, and advantages of the present invention are shown and described above. Those skilled in the art should understand that the present invention is not limited to the above embodiments. The above embodiments and descriptions are merely illustrative of the principles of the present invention. Various changes and modifications may be made to the present invention without departing from the spirit and scope of the present invention, and such changes and modifications fall within the scope of the invention as claimed.
Claims
1. A method for preparing polymer / active gas nanobubbles, characterized in that: The following steps are involved: S1, mixing polymer powder with an organic solvent to obtain a polymer mixture; S2, adding an emulsifier and an aqueous phase to the polymer mixture, and obtaining a primary emulsion by crushing; S3, adding a polymer solution and an emulsifier of a certain concentration to the primary emulsion, and emulsifying to obtain a secondary emulsion; S4, transferring the double emulsion to a polymer solution of another concentration, stirring and centrifuging, adding a freeze-drying protectant, and packaging and freeze-drying to obtain a freeze-dried powder of the polymer nanomembrane shell; S5, taking the freeze-dried powder into a bottle, filling it with active gas, then re-dissolving it with a solvent, and shaking it to obtain polymer / active gas nanobubbles.
2. The method for preparing polymer / active gas nanobubbles according to claim 1, wherein: The polymer in S1 is PLGA or PLLA; the polymer in S3 and S4 is PVA or PAM.
3. The method for preparing polymer / active gas nanobubbles according to claim 1, wherein: The organic solvent is anhydrous ethanol, dichloromethane or chloroform.
4. The method for preparing polymer / active gas nanobubbles according to claim 1, wherein: The emulsifier in S2 is Span 60, Span 65 or Span 80; the emulsifier in S3 is Tween 60, Tween 65 or Tween 80.
5. The method for preparing polymer / active gas nanobubbles according to claim 1, wherein: The concentrations of the polymer solutions in S3 and S4 were 5% w / v and 0.5% w / v, respectively.
6. The method for preparing polymer / active gas nanobubbles according to claim 1, wherein: The freeze-drying protective agent is mannitol, and the concentration is 10% w / v.
7. The method for preparing polymer / active gas nanobubbles according to claim 1, wherein: The active gas is: H2, H2S, CO, NO, O2, O3, Xe or SO2.
8. A polymer / active gas nanobubble, characterized in that: It is prepared using the preparation method according to any one of claims 1 to 7.
9. A tumor ablation system, comprising an ultrasound device, characterized in that: The invention also includes an injection device, wherein the injection device includes the polymer / active gas nanobubble according to claim 8, and the polymer / active gas nanobubble can ablate tumors under ultrasound stimulation.
10. Use of the polymer / active gas nanobubble according to claim 8 in the preparation of tumor therapeutic drugs and tumor diagnostic kits.