Panax notoginseng-derived exosome-like nanovesicles, preparation method and application thereof

Abstract

This invention relates to a high-purity Panax notoginseng-derived exosome nanovesicle, its preparation method, and its application in improving Parkinson's disease depression. The nanovesicles possess a typical bilayer lipid membrane structure, exhibiting a "teacup" or "biconcave saucer" appearance; their particle size distribution is concentrated, with an average particle size of 55-70 nm, a Zeta potential of -30 mV to -45 mV, and a particle concentration as high as 1.0 × 10⁻⁶. 12 The particle count is above 1000 mg / mL; the chemical composition is rich in triterpenoids (approximately 16.2%) and fatty acids and their compounds. Experiments have demonstrated that the Panax notoginseng exosomes provided by this invention can significantly improve motor dysfunction and depressive-like behavior in a Parkinson's disease-associated depression model mouse induced by MPTP combined with LPS. The nanovesicles provided by this invention have high purity, good stability, and strong biological activity, and have broad application prospects in the treatment of nervous system diseases and drug delivery.

Description

Technical Field

[0001] This invention belongs to the field of biomedical technology, specifically relating to a high-purity plant-derived exosome-like nanoparticle (PELNs) extracted from Panax notoginseng, and its application in the preparation of drugs for treating neurological diseases, especially Parkinson's disease and its comorbid depressive symptoms. Background Technology

[0002] Parkinson's disease (PD) is the second most common neurodegenerative disease after Alzheimer's disease, characterized by the progressive loss of dopaminergic neurons in the substantia nigra of the midbrain. In addition to typical motor symptoms (such as tremor, rigidity, and bradykinesia), approximately 50% of Parkinson's patients experience non-motor symptoms such as depression and anxiety, significantly impacting their quality of life. Current clinical treatment primarily relies on dopamine replacement therapy, such as levodopa; however, long-term use can easily lead to side effects such as dyskinesia, and its effectiveness in improving symptoms like depression is limited.

[0003] Exosomes are nanoscale vesicles secreted by cells, ranging from 30 to 150 nm in diameter. They possess a bilayer lipid membrane structure, protecting their internal load from enzymatic degradation and exhibiting low immunogenicity and good tissue penetration (including the blood-brain barrier). In recent years, plant-derived exosomes have become a research hotspot due to their wide availability, high yield, low cost, and good safety profile. Panax notoginseng, a traditional and valuable Chinese medicinal herb, has significant effects in promoting blood circulation, removing blood stasis, and neuroprotection. However, current research on Panax notoginseng exosomes is limited. Existing techniques often suffer from low extraction purity, wide particle size distribution, and significant loss of active ingredients, and there is a lack of pharmacodynamic evidence for the complex pathological model of Parkinson's disease combined with depression. Therefore, developing a high-purity, highly active Panax notoginseng exosome preparation that can simultaneously improve motor and psychiatric symptoms is of significant clinical importance. Summary of the Invention

[0004] The purpose of this invention is to solve the problems existing in the prior art, thereby providing a high-purity, high-concentration Panax notoginseng-derived exosome nanovesicle rich in specific active metabolites, and demonstrating its excellent stability at the level and significant efficacy in treating Parkinson's disease with depressive symptoms.

[0005] To solve the above-mentioned technical problems, the present invention is achieved through the following technical solution.

[0006] The first aspect of this invention provides the application of Panax notoginseng-derived exosome nanovesicles in the preparation of drugs for treating nervous system diseases.

[0007] Preferably, the neurological disease is selected from one or more of Parkinson's disease, depression, and Parkinson's disease with depression.

[0008] Preferably, the exosome-like nanovesicles derived from Panax notoginseng are prepared by the following method:

[0009] (1) Take fresh Panax notoginseng rhizomes, wash and chop them, add buffer solution and homogenize lightly;

[0010] (2) After homogenization, filter and centrifuge to remove plant fibers, then pre-filter through a 0.45μm filter membrane and take the supernatant;

[0011] (3) Centrifuge the supernatant obtained in step (2) for the first time to remove large vesicles, then take the supernatant and place it in a centrifuge tube for the second centrifugation; take the precipitate and resuspend it in the buffer solution for the third centrifugation; take the precipitate and resuspend it in the buffer solution to obtain the product.

[0012] Preferably, the cleaning in step (1) specifically involves cleaning with clean water; more preferably, cleaning with clean water 1-5 times.

[0013] Preferably, in step (1), the Panax notoginseng rhizomes are chopped into small pieces 0.3-0.8 cm wide.

[0014] Preferably, the buffer solution in step (1) is selected from PBS buffer; more preferably, the buffer solution is selected from PBS buffer with a concentration of 0.1 mol / L.

[0015] Preferably, the mass-to-volume ratio (g:mL) of Panax notoginseng rhizome to buffer solution in step (1) is 1:2-8.

[0016] Preferably, the centrifugation temperature in step (2) is 1-4℃, the rotation speed is 1000-5000g, and the time is 10-20min.

[0017] Preferably, the temperature of the first centrifugation in step (3) is 1-4℃, the rotation speed is 5000-15000g, and the time is 20-60min.

[0018] Preferably, the temperature of the second centrifugation in step (3) is 1-4℃, the rotation speed is 80000-110000g, and the time is 60-100min.

[0019] Preferably, the temperature of the third centrifugation in step (3) is 1-4℃, the rotation speed is 115000-150000g, and the time is 60-100min.

[0020] Preferably, in step (3), the precipitate is resuspended in PBS buffer and centrifuged a third time; more preferably, the concentration of the PBS buffer is 0.1 mol / L; most preferably, the PBS buffer is a pre-cooled PBS buffer with a concentration of 0.1 mol / L.

[0021] Preferably, in step (3), the precipitate is resuspended in PBS buffer; more preferably, the concentration of the PBS buffer is 0.1 mol / L; most preferably, the PBS buffer is a pre-cooled PBS buffer with a concentration of 0.1 mol / L.

[0022] A second aspect of the present invention provides a pharmaceutical composition for treating nervous system diseases, comprising exosome-like nanovesicles derived from Panax notoginseng and a pharmaceutically acceptable carrier.

[0023] Preferably, the neurological disease is selected from one or more of Parkinson's disease, depression, and Parkinson's disease with depression.

[0024] Preferably, the pharmaceutically acceptable carrier is selected from one or more of fillers, binders, disintegrants, lubricants, preservatives, antioxidants, chelating agents, colorants, flavoring agents, solvents, matrices, gelling agents, adhesives, humectants, softeners, plasticizers, crosslinking agents, pH adjusters, and fragrances.

[0025] Preferably, the exosome-like nanovesicles derived from Panax notoginseng are prepared by the following method:

[0026] (1) Take fresh Panax notoginseng rhizomes, wash and chop them, add buffer solution and homogenize lightly;

[0027] (2) After homogenization, filter and centrifuge to remove plant fibers, then pre-filter through a 0.45μm filter membrane and take the supernatant;

[0028] (3) Centrifuge the supernatant obtained in step (2) for the first time to remove large vesicles, then take the supernatant and place it in a centrifuge tube for the second centrifugation; take the precipitate and resuspend it in the buffer solution for the third centrifugation; take the precipitate and resuspend it in the buffer solution to obtain the product.

[0029] Preferably, the cleaning in step (1) specifically involves cleaning with clean water; more preferably, cleaning with clean water 1-5 times.

[0030] Preferably, in step (1), the Panax notoginseng rhizomes are chopped into small pieces 0.3-0.8 cm wide.

[0031] Preferably, the buffer solution in step (1) is selected from PBS buffer; more preferably, the buffer solution is selected from PBS buffer with a concentration of 0.1 mol / L.

[0032] Preferably, the mass-to-volume ratio (g:mL) of Panax notoginseng rhizome to buffer solution in step (1) is 1:2-8.

[0033] Preferably, the centrifugation temperature in step (2) is 1-4℃, the rotation speed is 1000-5000g, and the time is 10-20min.

[0034] Preferably, the temperature of the first centrifugation in step (3) is 1-4℃, the rotation speed is 5000-15000g, and the time is 20-60min.

[0035] Preferably, the temperature of the second centrifugation in step (3) is 1-4℃, the rotation speed is 80000-110000g, and the time is 60-100min.

[0036] Preferably, the temperature of the third centrifugation in step (3) is 1-4℃, the rotation speed is 115000-150000g, and the time is 60-100min.

[0037] Preferably, in step (3), the precipitate is resuspended in PBS buffer and centrifuged a third time; more preferably, the concentration of the PBS buffer is 0.1 mol / L; most preferably, the PBS buffer is a pre-cooled PBS buffer with a concentration of 0.1 mol / L.

[0038] Preferably, in step (3), the precipitate is resuspended in PBS buffer; more preferably, the concentration of the PBS buffer is 0.1 mol / L; most preferably, the PBS buffer is a pre-cooled PBS buffer with a concentration of 0.1 mol / L.

[0039] A third aspect of this invention provides a method for preparing exosome-like nanovesicles derived from Panax notoginseng, comprising the following steps:

[0040] (1) Take fresh Panax notoginseng rhizomes, wash and chop them, add buffer solution and homogenize lightly;

[0041] (2) After homogenization, filter and centrifuge to remove plant fibers, then pre-filter through a 0.45μm filter membrane and take the supernatant;

[0042] (3) Centrifuge the supernatant obtained in step (2) for the first time to remove large vesicles, then take the supernatant and place it in a centrifuge tube for the second centrifugation; take the precipitate and resuspend it in the buffer solution for the third centrifugation; take the precipitate and resuspend it in the buffer solution to obtain the product.

[0043] Preferably, the cleaning in step (1) specifically involves cleaning with clean water; more preferably, cleaning with clean water 1-5 times.

[0044] Preferably, in step (1), the Panax notoginseng rhizomes are chopped into small pieces 0.3-0.8 cm wide.

[0045] Preferably, the buffer solution in step (1) is selected from PBS buffer; more preferably, the buffer solution is selected from PBS buffer with a concentration of 0.1 mol / L.

[0046] Preferably, the mass-to-volume ratio (g:mL) of Panax notoginseng rhizome to buffer solution in step (1) is 1:2-8.

[0047] Preferably, the centrifugation temperature in step (2) is 1-4℃, the rotation speed is 1000-5000g, and the time is 10-20min.

[0048] Preferably, the temperature of the first centrifugation in step (3) is 1-4℃, the rotation speed is 5000-15000g, and the time is 20-60min.

[0049] Preferably, the temperature of the second centrifugation in step (3) is 1-4℃, the rotation speed is 80000-110000g, and the time is 60-100min.

[0050] Preferably, the temperature of the third centrifugation in step (3) is 1-4℃, the rotation speed is 115000-150000g, and the time is 60-100min.

[0051] Preferably, in step (3), the precipitate is resuspended in PBS buffer and centrifuged a third time; more preferably, the concentration of the PBS buffer is 0.1 mol / L; most preferably, the PBS buffer is a pre-cooled PBS buffer with a concentration of 0.1 mol / L.

[0052] Preferably, in step (3), the precipitate is resuspended in PBS buffer; more preferably, the concentration of the PBS buffer is 0.1 mol / L; most preferably, the PBS buffer is a pre-cooled PBS buffer with a concentration of 0.1 mol / L.

[0053] The fourth aspect of the present invention provides exosome-like nanovesicles derived from Panax notoginseng prepared according to the above preparation method.

[0054] Preferably, the median diameter of the exosome-like nanovesicles derived from Panax notoginseng is 50-70 nm, and the average particle size is 55-70 nm.

[0055] Preferably, the zeta potential range of the exosome-like nanovesicles derived from Panax notoginseng is -30mV to -45mV.

[0056] Preferably, the original concentration of the exosome-like nanovesicles derived from Panax notoginseng is ≥1.0×10⁻⁶. 12 Particles / mL.

[0057] Preferably, the relative content of triterpenoids in the exosome-like nanovesicles derived from Panax notoginseng is ≥15%.

[0058] Compared with the prior art, the present invention has the following advantages:

[0059] This invention, through an optimized extraction process, yields nanovesicles with uniform particle size and extremely high concentration, overcoming the low recovery rate problem of traditional methods. This invention is the first to demonstrate the effectiveness of Panax notoginseng exosomes in a comorbid model of Parkinson's disease and depression, providing a new strategy for clinical treatment. The active ingredients of this invention are derived from medicinal and edible plants, endogenously rich in triterpenoid active substances, exhibiting good biocompatibility and low risk of side effects. Suitable for long-term use, it has great potential in the prevention and improvement of depression, and possesses significant clinical significance and application value. Attached Figure Description

[0060] Figure 1 This is a schematic diagram of the TEM morphology characterization results of the Panax notoginseng-derived exosome nanovesicles prepared in Example 1.

[0061] Figure 2 This is a schematic diagram of the NTA analysis results of the Panax notoginseng-derived exosome nanovesicles prepared in Example 1.

[0062] Figure 3 This is a schematic diagram of the NanoFCM analysis results of the Panax notoginseng-derived exosome nanovesicles prepared in Example 1.

[0063] Figure 4 This is a schematic diagram showing the Zeta potential analysis results of the Panax notoginseng-derived exosome nanovesicles prepared in Example 1.

[0064] Figure 5 This is a schematic diagram (positive ion mode) showing the non-targeted metabolomics detection and analysis results of the Panax notoginseng-derived exosome nanovesicles prepared in Example 1.

[0065] Figure 6 This is a schematic diagram of the non-targeted metabolomics detection and analysis results of the Panax notoginseng-derived exosome nanovesicles prepared in Example 1 (negative ion mode).

[0066] Figure 7 This is a schematic diagram showing the composition and classification of core metabolites in the Panax notoginseng-derived exosome nanovesicles prepared in Example 1.

[0067] Figure 8This is a schematic diagram showing the pole turning time results in the pole climbing experiment of Example 2.

[0068] Figure 9 This is a schematic diagram showing the total climbing time results of the pole climbing experiment in Example 2.

[0069] Figure 10 This is a schematic diagram of the total motion distance results in the open field experiment of Example 2.

[0070] Figure 11 This is a schematic diagram showing the number of times the shuttle passes through the central region in the open field experiment of Example 2.

[0071] Figure 12 This is a schematic diagram showing the dwell time results in the central region of the open field experiment in Example 2.

[0072] Figure 13 This is a schematic diagram of the motion trajectory results in the open field experiment of Example 2.

[0073] Figure 14 This is a schematic diagram of the stationary time results of the tail suspension test in Example 2.

[0074] Figure 15 This is a schematic diagram of the immobility time results in the forced swimming experiment of Example 2. Detailed Implementation

[0075] To make the objectives, technical solutions, and effects of this invention clearer and more explicit, the invention will be further described in detail below with reference to embodiments. It should be understood that the specific embodiments described herein are merely illustrative of the invention and are not intended to limit the invention.

[0076] Unless otherwise specified, the experimental animals used in this invention are adult male C56BL6 / J mice, 2 months old and weighing 26g, purchased from Vital River Pharmaceuticals. All experimental animals were housed in an SPF-grade animal facility at a temperature maintained at 23±2℃ and humidity controlled at 50±15%. Normal acclimatization was performed one week prior to the start of the experiments, with all animals having free access to food and water. All experiments were conducted with the approval of the animal ethics committee, and all behavioral experiments were conducted daily from 8:00 to 18:00. All experimental methods used in this invention, including animal experiments, are conventional methods and techniques in the art.

[0077] Example 1

[0078] First, exosome-like nanovesicles derived from Panax notoginseng were prepared. The specific steps are as follows:

[0079] (1) Take 1 kg of fresh Panax notoginseng rhizomes, wash them three times with clean water to remove withered, shriveled or other undesirable tissues, cut the Panax notoginseng into small pieces 0.5 cm wide, add 0.1 mol / L PBS buffer at a ratio of 1:3 (g:mL), and homogenize lightly using a homogenizer.

[0080] (2) Filter the supernatant using a fine-mesh filter and retain the supernatant. Centrifuge the supernatant initially at 3000×g for 15 minutes at 4℃ to remove plant fibers (this can be repeated until no visible precipitate is visible). Pre-filter the supernatant through a 0.45μm filter membrane and collect the supernatant.

[0081] (3) Centrifuge the supernatant at 10000×g for 30 minutes at 4℃ to remove large vesicles. Resuspend the supernatant in an ultrafiltration tube and centrifuge at 100000×g for 70 minutes at 4℃. Discard the supernatant and collect the precipitate. Add an appropriate amount of pre-cooled 0.1mol / L PBS buffer to resuspend the precipitate and centrifuge a second time at 120000×g for 70 minutes at 4℃. Discard the supernatant and collect the precipitate. Add 200μL of pre-cooled 0.1mol / L PBS buffer to resuspend the precipitate. This is the collected exosome.

[0082] The exosome-like nanovesicles derived from Panax notoginseng prepared above were observed using transmission electron microscopy (TEM), and the results are shown in Figure 1. The results showed that the extract exhibited a typical "teacup-shaped" or "biconcave saucer-shaped" appearance, with a clear and complete outline of the double-layer lipid membrane. The membranes were uniform in size, free from impurities and ruptures, and the background of the electron microscope field was clean, with no obvious aggregation of extraneous proteins or cell debris, confirming the high efficiency of the extraction process and the extremely high purity of the sample.

[0083] The particle size was determined using NTA technology, and the results are as follows: Figure 2 As shown in the figure. The results show that the main peak is located at 130.3 nm, with a concentration of 3.5 × 10⁻⁶. 11 Particles / mL. NanoFCM technology was used for higher precision detection, and the results are as follows: Figure 3 As shown in the figure. The results revealed a large number of small-diameter vesicles, with a median diameter of 57.8 nm and an average diameter of 62.3 nm, resulting in a total concentration as high as 1.21 × 10⁻⁶. 12 Particles / mL.

[0084] Subsequently, the Zeta potential of the Panax notoginseng-derived exosome-like nanovesicles prepared above was detected, and the results are as follows: Figure 4 As shown in the figure. The results show that its Zeta potential is -38.24±2.04mV, indicating that the particle surface carries a strong negative charge and the system is stable.

[0085] Furthermore, the exosome-like nanovesicles derived from Panax notoginseng prepared above were subjected to non-targeted metabolomics detection (LC-MS), and the results are as follows. Figure 5-7 As shown in the figure. The results show that the chromatographic baseline is stable and the peaks are sharp, with a total of 308 metabolites identified (of which 167 were identified in positive ion mode). Figure 5 ), 141 ( ) were identified in the negative ion mode. Figure 6 The main components are triterpenoids (16.2%), fatty acids and their compounds (14.3%), amino acids and their derivatives, etc. Figure 7 The high abundance retention of triterpenoids proves that this extraction method can effectively enrich the core medicinal components of Panax notoginseng.

[0086] Example 2

[0087] First, a combined model of Parkinson's disease and depression was constructed. Specifically, adult male C56BL6 / J mice, aged 2 months and weighing 26g, were selected. They were injected intraperitoneally with MPTP at a concentration of 30mg / kg for 7 days to induce neurotoxicity, and then injected with LPS at a concentration of 0.5mg / kg for 1 day to induce neuroinflammation.

[0088] After modeling, mice were randomly divided into three groups of eight each: the model group, the Panax notoginseng exosome group, and the blank control group. Treatment was administered via nasal drops. The Panax notoginseng exosome group received exosome-like nanovesicles derived from Panax notoginseng prepared in Example 1 at a concentration of 1 mg / mL, administered in single doses of 20 μL. The model group and the blank control group received an equal volume of physiological saline. Treatment continued for 7-14 days.

[0089] The motor function of each group of mice was then assessed using the pole test.

[0090] The pole-climbing test is a standard technique used in animal models of movement disorders such as Parkinson's disease to evaluate motor function deficits. It's an experimental method for assessing bradykinesia and coordination. The animal is placed head-up at the top of a vertical pole. In this environment, the animal's instinctive behavior is to climb back to safety. Another common behavior is an attempt to turn around, often with slow or stiff movements, inevitably exhibiting bradykinesia. Observing and recording a series of parameters during the animal's turning and downward climbing can be used to evaluate the efficacy of anti-Parkinson's drugs or neuroprotective agents. The specific test method is as follows: The mouse is removed from its cage and placed head-up at the top of a vertical, fixed, rough-surfaced round pole (50 cm high, 1 cm in diameter). The mouse is allowed to turn freely and climb downwards. The time it takes for the mouse to turn completely (head-down) from head-up is recorded; the total time is recorded when the mouse continues climbing until all four limbs fully contact the base. At the end of the experiment (or when the mouse has not completed the test within the set maximum cutoff time of 60 seconds), the mouse is removed from the testing device and returned to its cage. The mouse's turning time and total time from being placed on the pole to the bottom are recorded and analyzed using a manual timing or video tracking system.

[0091] The results are as follows Figure 8-9 As shown in the figure. The results showed that, compared with the model group, the time for climbing and turning on the pole and the total time were significantly shortened in the Panax notoginseng exosome group, indicating that the motor impairment was improved.

[0092] Furthermore, the constructed Parkinson's disease-associated depression mouse model was subjected to the Open Field Test (OFT), Forced Swimming Test (FST), and Tail Suspension Test (TST) to evaluate the antidepressant effect of Panax notoginseng-derived exosome nanovesicles.

[0093] The open field test is a standard technique used in animal models of anxiety or depression to evaluate neuropsychiatric states. It is an experimental method for assessing general spontaneous activity and anxiety levels in animals. Animals are placed in an open and unfamiliar environment. In this environment, rodents, due to their natural instinct to avoid predators, often move along the edges (tactile tropism). Another common behavior is exploring the central open area out of curiosity about the new environment. The conflict between the animal's desire to explore and its fear of the unknown is thus revealed. A series of parameters are observed and recorded during the animal's free exploration, which can be used to assess the efficacy of anti-anxiety drugs or the animal's motor function. The specific testing method is as follows: Mice are removed from their cages and gently placed in the central area of ​​a square open field test chamber (e.g., 50cm × 50cm × 50cm, with solid-colored inner walls and bottom) without a lid. The mice are allowed to explore freely for 5 minutes (or 10 minutes) in a quiet, evenly lit environment. At the end of the experiment, the mice were removed from the reaction chamber and returned to their cages. The bottom and inner walls of the reaction chamber were thoroughly wiped with 75% alcohol to eliminate any interference from feces or odors for the next mouse. Key parameters such as the total distance the mice traveled, the time spent in the central area, and the number of times they traversed the central area during the test were recorded and analyzed using behavioral video tracking software such as ANY-maze.

[0094] Forced swimming is a technique used in animal model studies of depression to assess the criteria for depression. It is an experimental method for assessing despair. Animals are placed in water at a suitable temperature. In this environment, struggling is a common behavior, as is attempting to escape but often failing or being unable to escape. This creates an unavoidable oppressive environment. After a period of time, the animals exhibit a typical "immobile state." A series of parameters are observed and recorded during the animals' movement and floating, which can be used to evaluate the efficacy of antidepressants. The specific test method is as follows: Mice are removed from their cages and placed in a transparent glass jar (50 cm high, 30 cm in diameter) filled with water to a depth of approximately 30 cm (23±2℃). The mice are allowed to swim freely for 6 minutes. When a mouse floats on the surface without struggling, it is considered immobile. At the end of the test, the mice are removed from the water, dried, and returned to their cages. The total immobile time in the last 4 minutes of the 6-minute test is recorded and analyzed using ANY-maze software.

[0095] Tail suspension behavior testing is widely used in basic research on antidepressant drug testing. It is a classic method for rapidly assessing the efficacy of antidepressants, stimulants, and sedatives. The principle involves mice suspended by their tails, attempting to escape but failing, eventually giving up and entering a specific state of depressive immobility. The duration of immobility is recorded to reflect the depressive state; antidepressants and stimulants can significantly shorten or alter this state. The experimental animal's tail is fixed, leaving its head hanging downwards. The animal struggles in this environment, attempting to escape. After failing to escape, it exhibits intermittent immobility, displaying a state of "behavioral despair." The animal's desperate struggle to escape provides an inescapable, oppressive environment. After a period of experimentation, a series of parameters are recorded during the process of the animal developing this state of despair and immobility. This typical "immobility" reflects a state called "behavioral despair," a model similar to depression. This model is sensitive to most antidepressants, and its efficacy is significantly correlated with clinical efficacy, thus it is widely used in the initial screening of antidepressants. The specific testing method is as follows: The mouse was placed in a sealed soundproof box that isolated both hearing and vision. It was attached to a hook about 50cm above the ground with tape about 1cm from the tip of the mouse's tail. The animal's activity was recorded at the same time. The total immobility time in the last 4 minutes of the 6-minute test time was recorded and analyzed using ANY-maze software (Stoeling Co. Ltd.; USA).

[0096] Experimental results are as follows Figure 10-15 As shown in the figure. The results showed that the total movement distance of mice in the Panax notoginseng exosome group increased, the number of times they shuttled through the central area increased, the time spent in the central area increased, the immobile time decreased significantly, and the struggling time increased, indicating that the depressive-like behavior was significantly improved.

[0097] The above detailed embodiments provide a specific description of the analytical methods involved in this invention. It should be noted that the above description is only intended to help those skilled in the art better understand the methods and ideas of this invention, and is not intended to limit the scope of the invention. Without departing from the principles of this invention, those skilled in the art can make appropriate adjustments or modifications to this invention, and such adjustments and modifications should also fall within the protection scope of this invention.

Claims

1. Application of Panax notoginseng-derived exosome nanovesicles in the preparation of drugs for treating nervous system diseases.

2. The application according to claim 1, characterized in that, The neurological disease is selected from one or more of Parkinson's disease, depression, and Parkinson's disease with depression.

3. The application according to claim 1, characterized in that, The exosome-like nanovesicles derived from Panax notoginseng were prepared by the following method: (1) Take fresh Panax notoginseng rhizomes, wash and chop them, add buffer solution and homogenize lightly; (2) After homogenization, filter and centrifuge to remove plant fibers, then pre-filter through a 0.45μm filter membrane and take the supernatant; (3) Centrifuge the supernatant obtained in step (2) for the first time to remove large vesicles, then take the supernatant and place it in a centrifuge tube for the second centrifugation; take the precipitate and resuspend it in the buffer solution for the third centrifugation; take the precipitate and resuspend it in the buffer solution to obtain the product.

4. A pharmaceutical composition for treating nervous system diseases, characterized in that, It contains exosome-like nanovesicles derived from Panax notoginseng, and pharmaceutically acceptable carriers.

5. The pharmaceutical composition according to claim 4, characterized in that, The neurological disease is selected from one or more of Parkinson's disease, depression, and Parkinson's disease with depression.

6. The pharmaceutical composition according to claim 4, characterized in that, The exosome-like nanovesicles derived from Panax notoginseng were prepared by the following method: (1) Take fresh Panax notoginseng rhizomes, wash and chop them, add buffer solution and homogenize lightly; (2) After homogenization, filter and centrifuge to remove plant fibers, then pre-filter through a 0.45μm filter membrane and take the supernatant; (3) Centrifuge the supernatant obtained in step (2) for the first time to remove large vesicles, then take the supernatant and place it in a centrifuge tube for the second centrifugation; take the precipitate and resuspend it in the buffer solution for the third centrifugation; take the precipitate and resuspend it in the buffer solution to obtain the product.

7. A method for preparing exosome-like nanovesicles derived from Panax notoginseng, characterized in that, Includes the following steps: (1) Take fresh Panax notoginseng rhizomes, wash and chop them, add buffer solution and homogenize lightly; (2) After homogenization, filter and centrifuge to remove plant fibers, then pre-filter through a 0.45μm filter membrane and take the supernatant; (3) Centrifuge the supernatant obtained in step (2) for the first time to remove large vesicles, then take the supernatant and place it in a centrifuge tube for the second centrifugation; take the precipitate and resuspend it in the buffer solution for the third centrifugation; take the precipitate and resuspend it in the buffer solution to obtain the product.

8. The Panax notoginseng-derived exosome nanovesicles prepared by the preparation method according to claim 7.

9. A pharmaceutical preparation, characterized in that, The median diameter of the exosome-like nanovesicles derived from Panax notoginseng is 50-70 nm, and the average particle size is 55-70 nm.

10. The pharmaceutical preparation according to claim 9, characterized in that, The zeta potential range of the exosome-like nanovesicles derived from Panax notoginseng is -30mV to -45mV.

Images