Use of a Chinese medicine composition in preparing a drug for preventing or treating nervous system diseases

Abstract

The present invention belongs to the technical field of traditional Chinese medicine, and specifically discloses a use of a traditional Chinese medicine composition in preparing a drug for preventing or treating neurodegenerative diseases. The traditional Chinese medicine composition is made of leeches, chuanxiong, salvia miltiorrhiza and astragalus, has the effects of promoting blood circulation and removing blood stasis, invigorating qi and unblocking collaterals, and can effectively improve general behavioral performance, enhance balance and coordination ability and autonomous activity ability, reduce bradykinesia symptoms, relieve anxiety, and significantly increase substantia nigra TH + The invention discloses a novel Chinese medicine composition for treating Parkinson's disease, which can reduce or delay the loss of dopaminergic neurons in the midbrain, has a neuroprotective effect, and has no liver or kidney damage. It has a significant effect on the treatment of Parkinson's disease and is a safe and effective Chinese medicine composition for the treatment of Parkinson's disease, and has a good prospect for promotion and application.

Description

Technical Field

[0001] The invention belongs to the technical field of traditional Chinese medicines, and particularly relates to use of a traditional Chinese medicine composition in preparing medicines for preventing or treating nervous system diseases. Background Art

[0002] Neurodegenerative diseases are a type of irreversible neurological disease caused by the loss of neurons in the brain and spinal cord, mainly including Alzheimer's disease, Parkinson's syndrome, amyotrophic lateral sclerosis, Huntington's disease, etc. As society enters an aging period, the incidence of neurodegenerative diseases is increasing, causing huge social medical consumption and family pressure.

[0003] Parkinson's disease is a common neurodegenerative disease in the elderly. The dopamine neurons in the substantia nigra of the patients degenerate and die, causing a significant decrease in the dopamine content in the substantia nigra in the striatum. Its clinical manifestations are mainly symptoms such as resting tremor, increased muscle tension, bradykinesia, postural reflex disorders and autonomic dysfunction, which are usually accompanied by mental, sleep and cognitive disorders, causing the patient to lose the ability to live independently and seriously reducing the patient's quality of life.

[0004] At present, the most effective drug for the treatment of Parkinson's disease is levodopa preparation. However, the clinical application reaction of levodopa preparation, although it can improve the symptoms of Parkinson's disease, but as the medication time is prolonged, it has toxic side effects on dopamine nerves, which will lead to the generation of cytotoxic free radicals, thereby destroying the dopamine neurons that are not dead, causing the condition to worsen and cannot be cured. In addition, surgical treatment (neuronucleotomy and deep brain electrical stimulation, etc.) has a greater risk of brain damage and has no significant improvement on axial symptoms such as abnormal posture and gait, dysphagia, etc., and can only be used as a supplementary treatment for drug treatment. How to effectively and safely treat Parkinson's and prevent it from worsening has become one of the mainstream directions of research on the treatment of Parkinson's disease.

[0005] In recent years, in view of the serious toxicity and side effects of Western medicine, the application of traditional Chinese medicine in the treatment of many diseases has begun to receive widespread attention and attention from researchers. Research on the anti-inflammatory treatment of Parkinson's disease with traditional Chinese medicine has made certain progress. Liu Xuewen et al., Observation on the efficacy of ginkgo leaf extract in the treatment of Parkinson's disease (Chinese Tropical Medicine, 2008, 8(6), 977-978), disclosed that ginkgo leaf extract has a good effect on Parkinson's disease, can relieve Parkinson's disease symptoms and improve the quality of life. Gao Junpeng et al., Triptolide can reduce MPP +Induced dopaminergic neuron damage in rats (Neuroscience Bulletin, 2008, 24 (3), 133-142) disclosed that triptolide can protect dopaminergic neurons by inhibiting microglial activation. Polygonatum sibiricum polysaccharide can significantly improve the behavior of Parkinson's disease rats by upregulating the expression of PPAR-γ, inhibiting inflammatory response and cell apoptosis, promoting the regeneration of dopaminergic neurons. Pan Jing et al., Protective Effect of Curcumin on Damage to Substantial Nigra Dopaminergic Neurons in Parkinson's Disease Mouse Model Zuoying (Chinese Journal of Modern Neurological Diseases, 2007, 7 (5), 421-426) proved that curcumin effectively antagonized the loss of substantia nigra dopaminergic neurons in Parkinson's disease mouse model induced by MPTP by reducing the content of reactive oxygen species in substantia nigra dopaminergic neurons and inhibiting inflammatory response. Celastrin can also prevent or delay the progression of Parkinson's disease model through antioxidant and anti-inflammatory effects.

[0006] Traditional Chinese medicine believes that Parkinson's disease is a syndrome of "basic deficiency and superficial excess". The cause of the disease is related to multiple factors such as wind, dampness, cold, deficiency, phlegm, fire, and blood stasis. The pathogenesis is complex. Initially, it is liver and kidney essence deficiency, yin and blood deficiency, tendon and vein malnutrition, and liver wind movement. Then, there is meridian blood blockage, blood stasis and phlegm, and deficiency and excess. The treatment should be based on the principle of promoting blood circulation and warming qi. Chuanzhi Tongluo preparation is made of leech, Chuanxiong, Danshen and Astragalus, and is developed based on Buyang Huanwu Decoction. Leech is the main drug in the formula, which is good at removing blood stasis, breaking blood and eliminating accumulation; Chuanxiong is the ministerial drug, which has an upward nature and can cooperate with leech to reach the brain meridians, activate blood circulation and unblock the meridians, so that the overall qi and blood flow smoothly and the organs function vigorously; Astragalus and Salvia miltiorrhiza are adjuvants, Astragalus replenishes qi to remove blood stasis, warm and unblock the meridians, and Salvia miltiorrhiza activates blood circulation and removes blood stasis, calms the mind and nourishes the heart; the whole formula has the effects of activating blood circulation and removing blood stasis, invigorating qi and unblocking the meridians. Therefore, the research on the use of Chuanxiong Tongluo preparations in the preparation of drugs for the treatment of Parkinson's disease is worthy of in-depth exploration. Summary of the invention

[0007] The object of the present invention is to provide a use of a Chinese medicine composition in preparing a drug for preventing or treating a nervous system disease. The technical solution of the present invention is as follows:

[0008] A use of a Chinese medicine composition in preparing a medicine for preventing or treating nervous system diseases, wherein the Chinese medicine composition is made of the following components: leech, chuanxiong, salvia miltiorrhiza and astragalus.

[0009] The nervous system disease is a degenerative disease of the nervous system in the elderly.

[0010] The nervous system disease is one or more of Alzheimer's disease, Parkinson's syndrome, amyotrophic lateral sclerosis, epilepsy or ischemic stroke.

[0011] Preferably, the neurological disease is Parkinson's syndrome.

[0012] The Chinese medicine composition of the present invention comprises the following components in parts by weight:

[0013] Leech 550-650 weight parts Chuanxiong 550-650 weight parts

[0014] 350-450 parts by weight of Salvia miltiorrhiza and 350-450 parts by weight of Astragalus.

[0015] Preferably, the Chinese medicine composition comprises the following components in parts by weight:

[0016] Leech 600 weight parts Chuanxiong 600 weight parts

[0017] 400 parts by weight of Salvia miltiorrhiza and 400 parts by weight of Astragalus.

[0018] The traditional Chinese medicine components are added with clinically acceptable auxiliary materials through conventional processes to prepare clinically acceptable oral preparations.

[0019] The clinically acceptable oral preparations include but are not limited to granules, capsules, pills, tablets, oral liquids, and mixtures; preferably, the oral preparation is a capsule; more preferably, the oral preparation is a Chuanzhi Tongluo capsule.

[0020] The use described in the present invention is discovered during the clinical application of a marketed drug (Chuanzhi Tongluo Capsule, National Medicine Standard No. Z20090031), but the present invention is not limited to the above-mentioned preparation. The preparation method of the preparation described in the present invention will be described in detail below.

[0021] The method for preparing the oral pharmaceutical preparation of the present invention comprises the following steps:

[0022] A. Crush the leech into coarse powder, add 2-10 times the amount of water, stir and extract at 37°C, centrifuge, take the supernatant, filter and sterilize it, and then concentrate it by ultrafiltration. Then, freeze-dry the concentrate and crush it to obtain leech freeze-dried powder for later use;

[0023] B. Reflux extraction of 2-10 times the amount of 65-95% ethanol is added to Chuanxiong, Danshen and Huangqi, filtering and concentrating the reflux liquid to obtain the alcohol concentrate, adding 2-6 times the amount of water of the raw medicinal materials to the residue, decoction extraction, filtering, concentrating the filtrate to obtain the water concentrate, combining the alcohol concentrate and the water concentrate, spray drying, and obtaining a mixed powder of Chuanxiong, Danshen and Huangqi;

[0024] C. The leech freeze-dried powder obtained in step A and the mixed powder of chuanxiong, danshen and astragalus obtained in step B are uniformly mixed, and the mixture is directly prepared into an oral pharmaceutical preparation through conventional procedures or by adding a pharmaceutically acceptable excipient.

[0025] Preferably, the method for preparing the oral preparation of the present invention comprises the following steps:

[0026] A. The leeches are crushed into coarse powder, 6 times the amount of water is added, and the mixture is extracted at 37°C with a stirring speed of 100 r / min for 20 hours. After centrifugation, the supernatant is pre-filtered through a 1 μm pore size membrane, sterilized by filtration through a 0.2 μm pore size membrane, and then concentrated through an ultrafiltration membrane with a molecular weight cutoff of 6-10 KDa. The concentrate is freeze-dried and crushed to obtain leech freeze-dried powder for later use;

[0027] B. Add 6 times the amount of 80% ethanol to reflux and extract the three ingredients of Chuanxiong, Danshen and Huangqi twice, filter and concentrate the reflux liquid to a relative water density of 1.1-1.3 to obtain the alcohol concentrate, add 4 times the amount of water of the raw medicinal materials to the residue, decoct and extract once, filter, concentrate the filtrate to a relative water density of 1.1-1.3 to obtain the water concentrate, combine the alcohol concentrate and the water concentrate, spray dry, and obtain the mixed powder of Chuanxiong, Danshen and Huangqi;

[0028] C. The leech freeze-dried powder obtained in step A and the mixed powder of chuanxiong, danshen and astragalus obtained in step B are uniformly mixed, and the mixture is directly prepared into an oral pharmaceutical preparation through conventional procedures or by adding a pharmaceutically acceptable excipient.

[0029] In a preferred embodiment, the Chuanzhi Tongluo preparation is in the form of granules, tablets, or capsules, and its preparation method is as follows:

[0030] A. Crush the leech into coarse powder, add 2-10 times the amount of water, stir and extract at 37°C, centrifuge, take the supernatant, filter and sterilize it, and then concentrate it by ultrafiltration. Then, freeze-dry the concentrate and crush it to obtain leech freeze-dried powder for later use;

[0031] The preferred technical scheme in step A is: the amount of water is 6 times, the stirring time is 20h, the stirring speed is 100r / min, the centrifugation time is 30min, the centrifugation speed is 6000r / min, the 1μm pore size membrane is pre-filtered, the 0.2μm pore size membrane is filtered and sterilized, and the retention specification of the ultrafiltration membrane for concentration is 6-10kDa;

[0032] B. Reflux extraction of 2-10 times the amount of 65-95% ethanol is added to Chuanxiong, Danshen and Huangqi, filtering and concentrating the reflux liquid to obtain the alcohol concentrate, adding 2-6 times the amount of water of the raw medicinal materials to the residue, decoction extraction, filtering, concentrating the filtrate to obtain the water concentrate, combining the alcohol concentrate and the water concentrate, spray drying, and obtaining a mixed powder of Chuanxiong, Danshen and Huangqi;

[0033] The preferred technical scheme in step B is: the amount of the refluxed alcohol solution is 6 times, the concentration of the alcohol solution is 80%, the reflux times are 2 times, and the reflux time is 2 hours; the amount of water used to extract the medicinal residue is 4 times, the extraction times are 1 time, and the extraction time is 1 hour; preferably, the relative density of the alcohol concentrate or the water concentrate at 50° C. is 1.1-1.3, and more preferably the relative density is 1.2;

[0034] C. Evenly mix the leech freeze-dried powder obtained in step A and the mixed powder of Chuanxiong, Danshen and Astragalus obtained in step B, add appropriate amount of dextrin and mix well, dry granulate to obtain granules; compress the granules and coat them with enteric coating to obtain tablets; and put the granules into enteric-coated capsules to obtain capsules.

[0035] In another embodiment, the dosage form of the present invention is a pill, and the preparation method thereof comprises the following steps:

[0036] A. Crush the leech into coarse powder, add 2-10 times the amount of water, stir and extract at 37°C, centrifuge, take the supernatant, filter and sterilize it, and then concentrate it by ultrafiltration. Then, freeze-dry the concentrate and crush it to obtain leech freeze-dried powder for later use;

[0037] The preferred technical scheme in step A is: the amount of water is 6 times, the stirring time is 20h, the stirring speed is 100r / min, the centrifugation time is 30min, the centrifugation speed is 6000r / min, the 1μm pore size membrane is pre-filtered, the 0.2μm pore size membrane is filtered and sterilized, and the retention specification of the ultrafiltration membrane for concentration is 6-10kDa;

[0038] B. Reflux extraction of 2-10 times the amount of 65-95% ethanol is added to Chuanxiong, Danshen and Huangqi, filtering and concentrating the reflux liquid to obtain the alcohol concentrate, adding 2-6 times the amount of water of the raw medicinal materials to the residue, decoction extraction, filtering, concentrating the filtrate to obtain the water concentrate, combining the alcohol concentrate and the water concentrate, spray drying, and obtaining a mixed powder of Chuanxiong, Danshen and Huangqi;

[0039] The preferred technical scheme in step B is: the amount of the refluxed alcohol solution is 6 times, the concentration of the alcohol solution is 80%, the reflux times are 2 times, and the reflux time is 2 hours; the amount of water used to extract the medicinal residue is 4 times, the extraction times are 1 time, and the extraction time is 1 hour; preferably, the relative density of the alcohol concentrate or the water concentrate at 50° C. is 1.1-1.3, and more preferably the relative density is 1.2;

[0040] C. The leech freeze-dried powder obtained in step A and the mixed powder of Chuanxiong, Danshen and Astragalus obtained in step B are uniformly mixed, dextrin is added and mixed, and the mixture is made into pills, dried, enteric-coated and polished to obtain pills.

[0041] During the clinical application of Chuanzhi Tongluo preparation, the inventor found that it has a significant effect on the treatment of Parkinson's disease. Later, relevant pharmacodynamic studies confirmed that Chuanzhi Tongluo preparation is an effective formula for the treatment of Parkinson's disease: it can effectively improve the general behavioral performance of Parkinson's disease mice, enhance the balance and coordination ability and autonomous activity ability of Parkinson's disease mice, reduce the symptoms of bradykinesia in Parkinson's disease mice, relieve the anxiety of Parkinson's disease mice, and significantly increase the substantia nigra TH in Parkinson's disease mice. +It can reduce the number of cells, reduce or delay the loss of dopaminergic neurons in the midbrain, has a neuroprotective effect, and has no liver or kidney damage, and has a definite and significant therapeutic effect on Parkinson's disease.

[0042] Experimental Example 1 Effect of Chuanzhi Tongluo Preparation on Behavioral Performance of Parkinson's Disease Model Mice

[0043] 1 Experimental Materials

[0044] 1.1 Experimental animals

[0045] Male C57BL / 6 mice, 9-10 weeks old, SPF grade, body weight 25±2g, provided by Lunan Pharmaceutical Group Co., Ltd., animal license number: SYXK(Lu)20180008. All animals were adaptively raised for 2 weeks before the experiment, with the temperature controlled at (23-25)℃, relative humidity (40-60)%, natural light, good ventilation, and free food, water and movement. Before the experiment, all healthy animals after adaptive feeding were tested for limb balance to screen out animals with flexible limb movement and coordination without obstacles.

[0046] 1.2 Reagents and instruments

[0047] The test drugs were Chuanzhi Tongluo Capsules (produced by Lunan Houpu Pharmaceutical Co., Ltd.); 1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP, Sigma); MSI-DIG-RT DigiGait animal gait detection and analysis system (USA).

[0048] 2 Experimental methods

[0049] 2.1 Modeling and drug administration

[0050] C57 mice were randomly divided into three groups according to the random number table method: normal group, model group and Chuanzhitongluo preparation treatment group, with 10 mice in each group. MPTP (solvent: sterile pure water, freshly prepared and used) was intraperitoneally injected into the model group and Chuanzhitongluo preparation treatment group once a day at 0.2 mL / time / mouse, and the injection doses were: 15 mg / kg on the first day, 20 mg / kg on the second day, and 30 mg / kg on the third to seventh days. The normal group was intraperitoneally injected with an equal volume of sterile pure water. During the period, the behavioral changes of mice were closely monitored, and necessary warming measures were taken appropriately. While administering MPTP, the normal group and the model group were gavaged with normal saline at 50 mL / kg body weight 3 times a day in the morning, noon and evening (each time with an interval of 4 hours), and the treatment group was gavaged with Chuanzhitongluo preparation at 65 mg / kg body weight for two consecutive weeks. During the experiment, the normal environmental conditions required by the animals were met, all animals were free to move and eat, and they were weighed every other day and the dosage was adjusted in time according to the changes in body weight.

[0051] 2.2 Behavioral testing

[0052] After intraperitoneal injection of the corresponding dose of MPTP every day, the general behavioral performance of the mice was closely observed. On the 15th day of the experimental period, the animals' dynamic behavioral performance was tested, including gait test, pole climbing test and tape removal test, to evaluate the rats' limb movement balance ability, bradykinesia symptom degree and motor coordination ability.

[0053] 2.2.1 Gait experiment

[0054] Gait training was performed once a day on the 12th to 14th day of the experimental period. The animals were placed in the transparent compartment of the horizontal gait meter in the opposite direction of the running belt. The running belt speed was set to 10 cm / s. The dynamic gait behavior indicators of the rats within 10 consecutive seconds were recorded using the DigiGait animal gait detection and analysis system: the total contact area of ​​the paws and the angle formed by the paws and the long axis of the rat's movement direction. Data were obtained for each rat three times (each time with an interval of 15 minutes), and the average value was statistically calculated. After each rat was changed, the running belt was thoroughly cleaned with a cotton cloth soaked in soapy water to avoid odor interference.

[0055] 2.2.2 Pole climbing experiment

[0056] A wooden pole (1 cm in diameter, 60 cm in length) with a small metal ball (about 5 cm in diameter) fixed on the top was wrapped with two layers of gauze (anti-slip) and placed vertically on a horizontal platform as a climbing pole device. All animals were trained to climb poles once a day from the 12th to the 14th day of the experimental period. The mice were placed on the top of the small ball and allowed to move naturally. The following residence time (the time taken by the mouse from the time it was placed on the small ball on the top of the pole to the time when it had a tendency to climb down the pole) and climbing pole time (the time taken by the mouse to climb down from the top of the pole to the time when the limbs completely contacted the platform at the bottom of the pole) were recorded. The test was averaged 3 times, with an interval of more than 10 minutes each time. If it paused or crawled in the opposite direction, it was retested. Each test time did not exceed 3 minutes. After each test, the entire climbing pole device and the horizontal platform surface were wiped with 75% alcohol to eliminate odors and avoid mutual interference.

[0057] 2.2.3 Tape removal experiment

[0058] A 0.5 cm × 0.5 cm medical tape was attached to the nose of the mouse, and the time taken for the mouse to completely remove the tape from the application was recorded. The test was repeated three times with an interval of 10 minutes each time. The longest observation time was 60 seconds. If it exceeded 60 seconds, the test was repeated.

[0059] 2.2.4 First, draw lines in the open field experiment analysis system (Smart3.0 software) to divide the bottom of the open field into 9 squares to avoid blind spots. In a relatively quiet environment, 4 mice were placed in the center of an open box with a size of 50cm×50cm×50cm (length×width×height) at the same time, and the animals moved freely. The autonomous activity behavior of the mice was recorded with a camera on the top of the open field. Each test lasted for 10 minutes, and the average of the three tests was taken. After the recording, a clear movement trajectory was obtained for analyzing the total movement distance, rest time, average movement speed, and number of grid crossings of the mice. Each time the mice were changed, the excrement on the inner wall and bottom of the open box should be thoroughly cleaned, and wiped with an alcohol cotton ball to eliminate odor interference.

[0060] 2.3 Statistical analysis

[0061] All experimental data were statistically processed using GraphPad Prism software version 5 (GraphPad Software, USA), and the results were expressed as mean ± standard deviation. P < 0.05 was considered to be significantly different, and P < 0.01 was considered to be extremely significantly different.

[0062] 3 Results

[0063] 3.1 Effects of Chuanzhi Tongluo preparation on general behavioral manifestations of Parkinson's disease model mice

[0064] Compared with the normal group, mice in the model group and Chuanzhi Tongluo preparation treatment group showed acute reactions such as limb tremor, erect hair, arched back, and erect tail 20 minutes after the first intraperitoneal injection of MPTP, accompanied by mild reduction in activity, slow reaction, and quietness. The above symptoms lasted for about 30 minutes and then gradually alleviated, and returned to normal after 1 hour. In addition, with the increase in the number of MPTP and Chuanzhi Tongluo preparation administration, the acute reactions of the model group and Chuanzhi Tongluo preparation treatment group gradually stabilized, with no significant difference, but the model group had more prominent symptoms such as reduced movement, unstable gait, and slow movement than the Chuanzhi Tongluo preparation treatment group.

[0065] 3.2 Effects of Chuanzhi Tongluo Preparation on Gait Behavior of Parkinson's Disease Model Mice

[0066] The results of gait motion analysis are shown in Table 1 and Figure 1 , 2, 3. Compared with the normal group, the total contact area of ​​the paws of the mice in the model group increased significantly (P<0.01), and the distance between the front and rear paws increased significantly (P<0.01); the angles of the right and left hind paws formed by the paws and the long axis of the mouse's movement direction increased significantly (P<0.01), and the angles of the left and right forepaws decreased significantly (P<0.01), indicating that the model group mice had limb movement disorders and weakened balance and coordination ability. Compared with the model group, the balance and coordination ability of the mice in the Chuanzhi Tongluo preparation treatment group was improved to a certain extent, including a significant decrease in the total contact area (P<0.01), a significant decrease in the distance between the front paws (P<0.01), a significant decrease in the angles of the right and left hind paws (P<0.01), and an increase in the angles of the left and right forepaws.

[0067] Table 1 Comparison of total contact area, spacing and angle of paws of mice in each group (n=10)

[0068]

[0069] Note: “**” indicates P<0.01 compared with the normal group, △△ ” indicates P < 0.01 compared with the model group.

[0070] 3.3 Effect of Chuanzhi Tongluo preparation on the climbing behavior of Parkinson's disease model mice

[0071] The results of the pole climbing test are shown in Table 2. Figure 4 As shown in the figure, compared with the normal group, the time taken by the mice in the model group from the time they were placed on the top of the ball to the time they had a tendency to climb down the pole and the time taken to climb the pole completely were significantly prolonged (P<0.01); compared with the model group, the residence time and total climbing time of the group treated with Chuanzhi Tongluo preparation were significantly shortened (P<0.05). In contrast, the degree of bradykinesia symptoms in the group treated with Chuanzhi Tongluo preparation was alleviated.

[0072] Table 2 Comparison of the time mice in each group spent on the ball and the time they spent climbing the pole (n=10)

[0073]

[0074] Note: “*” indicates P<0.05 compared with the normal group, “**” indicates P<0.01 compared with the normal group; △△ ” indicates P < 0.01 compared with the model group.

[0075] 3.4 Effect of Chuanzhi Tongluo preparation on the behavioral performance of adhesive tape removal in Parkinson's disease model mice

[0076] The results of tape removal time are shown in Table 3. Figure 5As shown, compared with the normal group, the time for removing the adhesive tape in the model group was significantly prolonged (P<0.01); compared with the model group, the time for removing the adhesive tape in the Chuanzhi Tongluo preparation treatment group was significantly shortened (P<0.01), and the forelimbs were more coordinated and flexible.

[0077] Table 3 Comparison of adhesive tape removal time in each group of mice (n=10)

[0078]

[0079]

[0080] Note: “**” indicates P<0.01 compared with the normal group, △ ” indicates P < 0.05 compared with the model group.

[0081] 3.5 Effect of Chuanzhi Tongluo preparation on open field behavior of Parkinson's disease model mice

[0082] The activity trajectory diagram finally generated by the system is as follows: Figure 6 As shown, from Figure 6 It can be seen that in the novel environment, the movement routes of the mice in the normal group were complex and varied, and the amount of activity was high, covering the entire bottom of the box, with large amounts of activity in the central area and obvious exploratory behavior; compared with the mice in the normal group, the movement routes of the animals in the model group were simple and clear, with less activity and mostly moving along the walls around the bottom of the open box. The movement routes in the central area were clear, and the exploratory behavior was also significantly reduced, suggesting obvious anxiety; after treatment with Chuanzhi Tongluo preparation, the activity of the mice in the treatment group increased compared with the model group, the movement trajectories on the bottom of the box were more evenly distributed, the autonomous activity and exploratory behavior in the central area increased significantly, and anxiety was relieved.

[0083] Smart 3.0 software was used to further quantify the autonomous activity behavior of the mice in the above groups. The results are shown in Table 4. Figure 7 As shown, compared with the normal group, the total distance moved by mice in the model group was significantly shortened (P<0.01), the average speed was significantly slowed down (P<0.05), the rest time was significantly prolonged (P<0.01), and the number of grid crossings was significantly reduced (P<0.01); compared with the model group, the total distance moved by the Chuanzhi Tongluo preparation treatment group was significantly increased (P<0.05), the average speed was significantly increased (P<0.05), the rest time was significantly reduced (P<0.05), and the number of grid crossings was significantly increased (P<0.01). This indicates that Chuanzhi Tongluo preparation can enhance the spontaneous activity ability of mice.

[0084] Table 4 Comparison of total movement distance, average speed, rest time, and number of grid crossings of mice in each group (n=10)

[0085]

[0086] Note: “*” indicates P<0.05 compared with the normal group, “**” indicates P<0.01 compared with the normal group;

[0087] “ △ " indicates P < 0.05 compared with the model group," △△ ” indicates P < 0.01 compared with the model group.

[0088] Experimental Example 2: Neuroprotective effect of Chuanzhi Tongluo preparation on Parkinson's disease model mice

[0089] 1 Materials and Instruments

[0090] The experimental animals were mice after the behavioral test in Experimental Example 1. 10% chloral hydrate, 4% paraformaldehyde, PBS buffer, sucrose and goat anti-rabbit IgG were purchased from BBI Life Sciences Co., Ltd., OCT embedding medium was purchased from Shanghai SangonBiotech Engineering Co., Ltd., rabbit anti-mouse tyrosine hydroxylase (TH) antibody (Abcam, UK), and ready-to-use immunohistochemistry kit (Wuhan Boster Biotechnology Co., Ltd.).

[0091] 2 Experimental methods

[0092] 2.1 Preparation of frozen sections of substantia nigra brain tissue

[0093] After the behavioral test, the mice were deeply anesthetized by intraperitoneal injection of 10% chloral hydrate (8 mL / kg) based on the last body weight. The heart was exposed by laparotomy. The heart was first perfused with normal saline until the liver turned white, and then perfused with 4% paraformaldehyde / PBS liquid for in vivo tissue fixation (about 50 mL was injected per mouse). The complete brain tissue was then quickly separated on ice and placed in 4% paraformaldehyde solution again at 4°C for another 2 hours in vitro. The tissue was then removed, rinsed with PBS solution twice, and then immersed in 10%, 20%, and 30% sucrose / PBS solution for dehydration for 24 hours until the tissue sank to the bottom. Subsequently, the tissue was carefully clamped and rinsed with PBS solution twice, 1 minute each time, to wash off the paraformaldehyde on the surface, and the water was wiped dry with filter paper. The tissue was embedded in tin foil with OCT embedding medium, and then quickly frozen in an appropriate amount of liquid nitrogen to make a frozen tissue embedding block. Then, continuous coronal frozen sections with a thickness of 10 μm in the midbrain region were prepared in a constant-temperature cryostat, and then dried (about 24 hours) and stored in a slice box at -80°C for subsequent immunohistochemical staining.

[0094] 2.2 Immunohistochemical staining

[0095] Take the slices of the substantia nigra layer of the midbrain stored at -80℃, soak them in PBS for 3 minutes, and dry them at room temperature; circle the tissue range with a tissue pen, and add a methanol solution containing 3% hydrogen peroxide to cover the tissue after it is completely dry, incubate at room temperature for 5-10 minutes to remove endogenous peroxidase; soak in PBS for 5 minutes, soak twice, add 5% bovine serum albumin diluted in PBS, incubate at room temperature for 30 minutes, block non-specific antigens, and dry; add 1:800 rabbit anti-mouse TH antibody diluted in PBS, and incubate at 4℃ overnight. The next day, soak in PBS for 5 minutes, soak twice, add 1:10000 dilution of goat anti-rabbit IgG secondary antibody, incubate at 37℃ for 20-30 minutes, soak in PBS for 5 minutes, soak twice; add SABC solution, and incubate at 37℃ for 30 minutes. DAB color developing solution (A:B=1:1) is used for color development for 3-15 minutes, and the staining depth is observed at any time under the microscope. The binding time is controlled and the reaction is terminated in time with PBS, and hematoxylin is added for counterstaining. Dehydrated with alcohol gradient (70%, 30s; 70%, 30s; 80%, 30s; 95%, 30s; 100%, 1min; 100%, 2min), transitioned with xylene alcohol solution (1:1) (1min), transparent with xylene (100% xylene 5min×2 times), sealed with neutral gum (30μL / slice), and dried at room temperature (avoid touching the slide). Observed and photographed under an optical microscope, and the number of TH-positive cells in each group was counted with the help of the image analysis and measurement software Image Pro Plus 6.0.

[0096] 2.3 Statistical analysis

[0097] All experimental data were statistically processed using GraphPad Prism software version 5 (GraphPad Software, USA), and the results were expressed as mean ± standard deviation. P < 0.05 was considered to be different, and P < 0.01 was considered to be statistically significant.

[0098] 3 Results

[0099] The most obvious pathological lesion of Parkinson's disease is the loss of dopamine in the substantia nigra of the midbrain, while the substantia nigra of the midbrain is positive for TH ( + The expression of ) neurons can indirectly reflect the loss of dopaminergic neurons. Figure 8 As shown, compared with the normal group, the model group TH + The number of cells decreased significantly (P<0.05), indicating that the neuron loss was serious, indicating that the Parkinson's disease model was successfully established; compared with the model group, the TH + The number of cells increased significantly (P<0.05). This suggests that Chuanzhi Tongluo preparation treatment can significantly reduce or delay the loss of dopamine in the substantia nigra of Parkinson's disease mice and protect dopaminergic neurons to a certain extent.

[0100] Table 5 Number of TH+ cells in the substantia nigra of the midbrain (n=10)

[0101]

[0102] Note: “**” indicates P<0.01 compared with the normal group, △△ ” indicates P < 0.01 compared with the model group.

[0103] Experimental Example 3 Effect of Chuanzhi Tongluo Preparation on Liver and Kidney Function in Parkinson's Disease Model Rats

[0104] 1 Experimental Materials

[0105] 1.1 Experimental animals

[0106] 40 healthy adult SPF SD rats, half male and half female, weighing 250±20g, were provided by Lunan Pharmaceutical Group Co., Ltd., animal license number: SYXK(Lu)2018 0008. All animals were adaptively raised for 1 week before the experiment, with the temperature controlled at (23-25)℃, relative humidity (40-60)%, natural light, good ventilation, and free food, water and movement.

[0107] 1.2 Materials and Instruments

[0108] The tested drugs were Chuanzhi Tongluo Capsules (produced by Lunan Houpu Pharmaceutical Co., Ltd.), 6-0HDA (produced by Shanghai Yuanye Biotechnology Co., Ltd.), apomorphine standard (produced by Shanghai Ruiqi Biotechnology Co., Ltd.), vitamin C tablets (produced by Guangdong South China Pharmaceutical Group Co., Ltd.), Madopar (produced by Shanghai Roche Pharmaceuticals Co., Ltd.), ELISA (serum) kits for alanine aminotransferase (ALT), aspartate aminotransferase (AST), creatinine (CR), blood urea nitrogen (BUN) and total bilirubin (TBIL) (produced by BBI Life Sciences Co., Ltd.); brain stereotaxic instrument (STOELTING, USA), SpectraMax M5 microplate spectrophotometer (produced by Molecular Devices, USA).

[0109] 2 Experimental methods

[0110] 2.1 Preparation of Parkinson's disease model

[0111] Under 10% chloral hydrate (350 mg / kg body weight) anesthesia, 6 μL (concentration 4 μg / μL) of 6-0HDA was injected into the left medial forebrain bundle at a single point, with the coordinates of tooth bar: -2.3 mm, antero-posterior: -4.4 mm, medio-lateral: 1.2 mm, dorso-ventral: -7.8 mm. Ten rats in the normal group (half male and half female) underwent sham surgery and were injected with 6 μL of 0.02% vitamin C saline. One month after modeling, 0.5 mg / kg body weight of apomorphine was injected intraperitoneally into the modeling rats. The rats would circle on the horizontal ground, and the rats that rotated more than 7 circles / min to the healthy side were considered to be successful Parkinson's disease models.

[0112] 2.2 Grouping and Dosing

[0113] The rats with successful modeling were randomly divided into 3 groups, each with 10 rats (half male and half female). The normal group and the model group were gavaged with distilled water at 1 ml / 100 g body weight, the Chuanzhi Tongluo preparation treatment group was gavaged with 135 mg / kg Chuanzhi Tongluo preparation, and the Madopar group was gavaged with 75 mg / kg Madopar every day for 30 days.

[0114] 2.3 Collection of materials

[0115] On the 31st day, the rats in each group were anesthetized with 10% chloral hydrate (350 mg / kg body weight) by intraperitoneal injection, and then the thoracic and abdominal cavities of the rats were opened and blood was collected from the abdominal aorta.

[0116] 2.4 Liver and kidney function tests

[0117] The serum of rats in each group was taken, and ALT, AST, CR, BUN and TBIL were detected strictly according to the operating procedures of the kit instructions.

[0118] 2.5 Statistical analysis

[0119] All experimental data were statistically processed using GraphPad Prism software version 5 (GraphPad Software, USA), and the results were expressed as mean ± standard deviation. P < 0.05 was considered to be significantly different, and P < 0.01 was considered to be extremely significantly different.

[0120] 3 Results

[0121] As shown in Table 6, Fig. 9As shown in the data, compared with the normal group of rats, there were no significant differences in ALT, AST, CR, BUN and TBIL levels between the model group and the Chuanzhi Tongluo preparation treatment group (P>0.05), while the ALT (P<0.05), CR (P<0.01) and BUN (P<0.05) levels of the rats in the Madopar group were significantly increased, indicating that Chuanzhi Tongluo preparation reduced liver and kidney function damage in the 6-OHDA-induced Parkinson's disease model rats compared with Madopar.

[0122] Table 6 Changes in biochemical indicators of liver and kidney function (n=10)

[0123]

[0124] Note: “**” indicates P<0.01 compared with the normal group.

[0125] The above experimental results show that the Chuanzhi Tongluo preparation prepared by the present invention can effectively improve the general behavioral performance of Parkinson's disease mice, enhance the balance and coordination ability and autonomous activity ability of Parkinson's disease mice, alleviate the symptoms of bradykinesia in Parkinson's disease mice, relieve the anxiety of Parkinson's disease mice, and significantly increase the TH level in the substantia nigra of Parkinson's disease mice. + It can reduce the number of cells, reduce / delay the loss of dopaminergic neurons in the midbrain, has a neuroprotective effect, and has no liver or kidney damage, and has a definite and significant therapeutic effect on Parkinson's disease. BRIEF DESCRIPTION OF THE DRAWINGS

[0126] Figure 1 This is a comparison of the total contact area of ​​the paws of mice in each group (n=10);

[0127] Figure 2 This is a comparison of the distance between the paws of mice in each group (n=10);

[0128] Figure 3 is a comparison of the paw angles of mice in each group (n=10);

[0129] Figure 4 This is a comparison of the time that mice in each group spent on the ball and the time that they spent on climbing the pole (n=10);

[0130] Figure 5 This is a graph comparing the adhesive tape removal time of mice in each group (n=10);

[0131] Figure 6 Figure 2 shows the open field movement trajectory of mice in each group;

[0132] Figure 7 This is a comparison of the total movement distance, average speed, rest time, and number of grid crossings of mice in each group (n=10);

[0133] Figure 8 Substantia nigra TH+ Cell number (n=10) graph;

[0134] Fig. 9 The figure shows the changes of biochemical indices of liver and kidney function (n=10). DETAILED DESCRIPTION

[0135] The present invention is further described below by means of specific examples, but those skilled in the art should be aware that the examples do not limit the present invention in any way.

[0136] Example 1 Preparation of Chuanzhi Tongluo Granules

[0137] Leech 550g Chuanxiong 550g

[0138] Salvia miltiorrhiza 400g Astragalus 400g

[0139] A. Crush the leech into coarse powder, add 2 times the amount of water, extract at 37°C at 100 r / min for 24 hours, centrifuge at 6000 r / min for 30 minutes, take the supernatant and filter it through a 1 μm filter membrane, sterilize it through a 0.2 μm filter membrane, and then concentrate it through a 6-10 kDa ultrafiltration. The concentrate is freeze-dried and crushed to obtain leech freeze-dried powder for later use;

[0140] B. Add 10 times the amount of 65% ethanol to reflux extract the three ingredients of Chuanxiong, Danshen and Huangqi twice, each time for 2 hours, filter and concentrate the reflux liquid to obtain an alcohol concentrate with a relative density of 1.1 at 50°C, add 4 times the amount of water of the raw medicinal materials to the residue, decoct and extract for 1 hour, filter, concentrate the filtrate to obtain a water concentrate with a relative density of 1.2 at 50°C, combine the alcohol concentrate and the water concentrate, spray dry, and obtain a mixed powder of Chuanxiong, Danshen and Huangqi;

[0141] C. Evenly mix the leech freeze-dried powder obtained in step A and the mixed powder of chuanxiong, danshen and astragalus obtained in step B, add appropriate amount of dextrin and mix well, and dry granulate to obtain granules.

[0142] Example 2 Preparation of Chuanzhi Tongluo Tablets

[0143] Leech 600g Chuanxiong 600g

[0144] Salvia miltiorrhiza 400g Astragalus 400g

[0145] A. Crush the leech into coarse powder, add 6 times the amount of water, extract at 37°C at 100 r / min for 24 hours, centrifuge at 6000 r / min for 30 minutes, take the supernatant and filter it through a 1μm filter membrane, sterilize it through a 0.2μm filter membrane, and then concentrate it through 6-10kDa ultrafiltration. The concentrate is freeze-dried and crushed to obtain leech freeze-dried powder for later use;

[0146] B. Add 5 times the amount of 70% ethanol to reflux extract the three ingredients of Chuanxiong, Danshen and Huangqi twice, each time for 2 hours, filter and concentrate the reflux liquid to obtain an alcohol concentrate with a relative density of 1.3 at 50°C, add 3 times the amount of water of the raw medicinal materials to the residue, decoct and extract for 1 hour, filter, concentrate the filtrate to obtain a water concentrate with a relative density of 1.2 at 50°C, combine the alcohol concentrate and the water concentrate, spray dry, and obtain a mixed powder of Chuanxiong, Danshen and Huangqi;

[0147] C. Evenly mix the leech freeze-dried powder obtained in step A and the mixed powder of Chuanxiong, Danshen and Astragalus obtained in step B, add appropriate amount of dextrin and mix well, dry granulate, press the granules into tablets, and coat with enteric coating to obtain tablets.

[0148] Example 3 Preparation of Chuanzhi Tongluo Capsules

[0149] Leech 600g Chuanxiong 600g

[0150] Salvia miltiorrhiza 400g Astragalus 400g

[0151] A. Crush the leech into coarse powder, add 8 times the amount of water, extract at 37°C at 100 r / min for 24 hours, centrifuge at 6000 r / min for 30 minutes, take the supernatant and filter it through a 1 μm filter membrane, sterilize it through a 0.2 μm filter membrane, and then concentrate it through 6-10 kDa ultrafiltration to a relative density of 1.15. Freeze-dry the concentrate and crush it to obtain leech freeze-dried powder for later use;

[0152] B. Add 6 times the amount of 80% ethanol to reflux extract the three ingredients of Chuanxiong, Danshen and Huangqi twice, each time for 2 hours, filter and concentrate the reflux liquid to obtain an alcohol concentrate with a relative density of 1.2 at 50°C, add 4 times the amount of water of the raw medicinal materials to the residue, decoct and extract for 1 hour, filter, concentrate the filtrate to obtain a water concentrate with a relative density of 1.2 at 50°C, combine the alcohol concentrate and the water concentrate, spray dry, and obtain a mixed powder of Chuanxiong, Danshen and Huangqi;

[0153] C. Evenly mix the leech freeze-dried powder obtained in step A and the mixed powder of Chuanxiong, Danshen and Astragalus obtained in step B, add appropriate amount of starch, mix well, granulate, and fill capsule shells to obtain capsules.

[0154] Example 4 Preparation of Chuanzhi Tongluo Pills

[0155] Leech 650g Chuanxiong 550g

[0156] Salvia miltiorrhiza 350g Astragalus 450g

[0157] A. Crush the leech into coarse powder, add 6 times the amount of water, extract at 37°C at 100 r / min for 24 h, centrifuge at 6000 r / min for 30 min, take the supernatant and pre-filter it through a 1 μm filter membrane, sterilize it through a 0.2 μm filter membrane, and then concentrate it to 600 mL (relative density 1.15 at 25°C) through 6-10 kDa ultrafiltration. Freeze-dry the concentrate and crush it to obtain leech freeze-dried powder for later use;

[0158] B. Add 6 times the amount of 80% ethanol to reflux extract the three ingredients of Chuanxiong, Danshen and Huangqi twice, each time for 2 hours, filter and concentrate the reflux liquid to obtain an alcohol concentrate with a relative density of 1.2 at 50°C, add 4 times the amount of water of the raw medicinal materials to the residue, decoct and extract for 1 hour, filter, concentrate the filtrate to obtain a water concentrate with a relative density of 1.3 at 50°C, combine the alcohol concentrate and the water concentrate, spray dry, and obtain a mixed powder of Chuanxiong, Danshen and Huangqi;

[0159] C. The leech freeze-dried powder obtained in step A and the mixed powder of Chuanxiong, Danshen and Astragalus obtained in step B are uniformly mixed, dextrin is added and mixed, and the mixture is made into pills, dried, enteric-coated and polished to obtain pills.

[0160] Example 5 Preparation of Chuanzhi Tongluo Oral Liquid

[0161] Leech 600g Chuanxiong 600g

[0162] Salvia miltiorrhiza 400g Astragalus 400g

[0163] A. Crush the leeches into coarse powder, add 8 times the amount of water, stir and extract at 37℃, 100r / min for 24h, centrifuge at 6000r / min for 30min, take the supernatant and filter it through a 1μm filter membrane, sterilize it through a 0.2μm filter membrane, and then concentrate it through a 6-10kDa ultrafiltration to a relative density of 1.15 for use;

[0164] B. Add 8 times the amount of 80% ethanol to reflux extract the three herbs of Chuanxiong, Danshen and Huangqi twice, each time for 2 hours, filter and concentrate the reflux liquid to obtain an alcohol concentrate with a relative density of 1.1 at 50°C, add 4 times the amount of water of the raw medicinal materials to the residue, decoct and extract for 1 hour, filter, concentrate the filtrate to obtain a water concentrate with a relative density of 1.2 at 50°C, and combine the alcohol concentrate and the water concentrate;

[0165] C. Evenly mix the leech concentrate obtained in step A and the concentrated solutions of rhizoma chuanxiong, salvia miltiorrhiza and astragalus obtained in step B, add water for injection and appropriate amounts of sodium benzoate and sucrose, adjust the pH to 7.0, concentrate, let stand, filter, fill and sterilize to obtain an oral solution.

[0166] Example 6 Preparation of Chuanzhitongluo Mixture

[0167] Leech 600g Chuanxiong 600g

[0168] Salvia miltiorrhiza 400g Astragalus 400g

[0169] A. Crush the leeches into coarse powder, add 6 times the amount of water, stir and extract at 37℃, 100r / min for 24h, centrifuge at 6000r / min for 30min, take the supernatant and filter it through a 1μm filter membrane, sterilize it through a 0.2μm filter membrane, and then concentrate it through a 6-10kDa ultrafiltration to a relative density of 1.15;

[0170] B. Add 6 times the amount of 80% ethanol to reflux extract 2 times for 2 hours each time, filter and concentrate the reflux liquid to obtain an alcohol concentrate with a relative density of 1.2 at 50°C, add 4 times the amount of water of the raw medicinal materials to the residue, decoct and extract for 1 hour, filter, concentrate the filtrate to obtain a water concentrate with a relative density of 1.2 at 50°C, and combine the alcohol concentrate and the water concentrate;

[0171] C. Evenly mix the leech concentrate obtained in step A and the concentrated solutions of rhizoma chuanxiong, salvia miltiorrhiza and astragalus obtained in step B, add appropriate amount of sodium benzoate, stir well, and add water to prepare a mixture.

[0172] Example 7 Preparation of Chuanzhi Tongluo Capsules

[0173] Leech 550g Chuanxiong 650g

[0174] Salvia miltiorrhiza 450g Astragalus 350g

[0175] A. Crush the leech into coarse powder, add 10 times the amount of water, extract at 37°C at 100 r / min for 24 hours, centrifuge at 6000 r / min for 30 minutes, take the supernatant and filter it through a 1 μm filter membrane, sterilize it through a 0.2 μm filter membrane, and then concentrate it through 6-10 kDa ultrafiltration to a relative density of 1.15. Freeze-dry the concentrate and crush it to obtain leech freeze-dried powder for later use;

[0176] B. Add 7 times the amount of 95% ethanol to reflux extract the three ingredients of Chuanxiong, Danshen and Huangqi twice, each time for 2 hours, filter and concentrate the reflux liquid to obtain an alcohol concentrate with a relative density of 1.15 at 50°C, add 4 times the amount of water of the raw medicinal materials to the residue, decoct and extract for 1 hour, filter, concentrate the filtrate to obtain a water concentrate with a relative density of 1.2 at 50°C, combine the alcohol concentrate and the water concentrate, spray dry, and obtain a mixed powder of Chuanxiong, Danshen and Huangqi;

[0177] C. Evenly mix the leech freeze-dried powder obtained in step A and the mixed powder of Chuanxiong, Danshen and Astragalus obtained in step B, add appropriate amount of starch, mix well, granulate, and fill capsule shells to obtain capsules.

[0178] Example 8 Preparation of Chuanzhi Tongluo Capsules

[0179] Leech 650g Chuanxiong 650g

[0180] Salvia miltiorrhiza 350g Astragalus 350g

[0181] A. Crush the leech into coarse powder, add 6 times the amount of water, extract at 37°C at 100 r / min for 24 h, centrifuge at 6000 r / min for 30 min, take the supernatant and pre-filter it through a 1 μm filter membrane, sterilize it through a 0.2 μm filter membrane, and then concentrate it to 600 mL (relative density 1.15 at 25°C) through 6-10 kDa ultrafiltration. Freeze-dry the concentrate and crush it to obtain leech freeze-dried powder for later use;

[0182] B. Add 6 times the amount of 80% ethanol to reflux extract the three ingredients of Chuanxiong, Danshen and Huangqi twice, each time for 2 hours, filter and concentrate the reflux liquid to obtain an alcohol concentrate with a relative density of 1.2 at 50°C, add 4 times the amount of water of the raw medicinal materials to the residue, decoct and extract for 1 hour, filter, concentrate the filtrate to obtain a water concentrate with a relative density of 1.2 at 50°C, combine the alcohol concentrate and the water concentrate, spray dry, and obtain a mixed powder of Chuanxiong, Danshen and Huangqi;

[0183] C. Evenly mix the leech freeze-dried powder obtained in step A and the mixed powder of Chuanxiong, Danshen and Astragalus obtained in step B, add appropriate amount of starch, mix well, granulate, and fill capsule shells to obtain capsules.

Claims

1. Use of a Chinese medicine composition in the preparation of a drug for preventing or treating Parkinson's syndrome, wherein the Chinese medicine composition is made of the following components in parts by weight: Leech 550-650 weight parts Chuanxiong 550-650 weight parts 350-450 parts by weight of Salvia miltiorrhiza and 350-450 parts by weight of Astragalus.

2. The use according to claim 1, characterized in that The Chinese medicine composition is made of the following components in parts by weight: Leech 600 weight parts Chuanxiong 600 weight parts 400 parts by weight of Salvia miltiorrhiza and 400 parts by weight of Astragalus.

3. The use according to claim 1 or 2, characterized in that The traditional Chinese medicine components are added with clinically acceptable auxiliary materials through conventional processes to prepare clinically acceptable oral preparations.

4. The use according to claim 3, characterized in that: The clinically acceptable oral preparations are granules, capsules, pills, tablets, oral liquids or mixtures.

5. The use according to claim 4, characterized in that The oral preparation is a capsule.

6. The use according to claim 5, characterized in that The capsule is Chuanzhi Tongluo Capsule.

Images