CN111759855A - Traditional Chinese medicine composition for preventing and treating Alzheimer's disease and application thereof - Google Patents

Traditional Chinese medicine composition for preventing and treating Alzheimer's disease and application thereof Download PDF

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CN111759855A
CN111759855A CN202010782377.2A CN202010782377A CN111759855A CN 111759855 A CN111759855 A CN 111759855A CN 202010782377 A CN202010782377 A CN 202010782377A CN 111759855 A CN111759855 A CN 111759855A
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chinese medicine
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medicine composition
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张宁
徐艳明
孙慧峰
杨波
王发善
王喜军
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Heilongjiang University of Chinese Medicine
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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/395—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins
    • A61K31/435—Heterocyclic compounds having nitrogen as a ring hetero atom, e.g. guanethidine or rifamycins having six-membered rings with one nitrogen as the only ring hetero atom
    • A61K31/47—Quinolines; Isoquinolines
    • A61K31/4738—Quinolines; Isoquinolines ortho- or peri-condensed with heterocyclic ring systems
    • A61K31/4745—Quinolines; Isoquinolines ortho- or peri-condensed with heterocyclic ring systems condensed with ring systems having nitrogen as a ring hetero atom, e.g. phenantrolines
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/075—Ethers or acetals
    • A61K31/085—Ethers or acetals having an ether linkage to aromatic ring nuclear carbon
    • A61K31/09—Ethers or acetals having an ether linkage to aromatic ring nuclear carbon having two or more such linkages
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/33—Heterocyclic compounds
    • A61K31/335—Heterocyclic compounds having oxygen as the only ring hetero atom, e.g. fungichromin
    • A61K31/35—Heterocyclic compounds having oxygen as the only ring hetero atom, e.g. fungichromin having six-membered rings with one oxygen as the only ring hetero atom
    • A61K31/352—Heterocyclic compounds having oxygen as the only ring hetero atom, e.g. fungichromin having six-membered rings with one oxygen as the only ring hetero atom condensed with carbocyclic rings, e.g. methantheline 
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/70—Carbohydrates; Sugars; Derivatives thereof
    • A61K31/7028—Compounds having saccharide radicals attached to non-saccharide compounds by glycosidic linkages
    • A61K31/7034—Compounds having saccharide radicals attached to non-saccharide compounds by glycosidic linkages attached to a carbocyclic compound, e.g. phloridzin
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/70—Carbohydrates; Sugars; Derivatives thereof
    • A61K31/7042—Compounds having saccharide radicals and heterocyclic rings
    • A61K31/7048—Compounds having saccharide radicals and heterocyclic rings having oxygen as a ring hetero atom, e.g. leucoglucosan, hesperidin, erythromycin, nystatin, digitoxin or digoxin
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P25/00—Drugs for disorders of the nervous system
    • A61P25/28—Drugs for disorders of the nervous system for treating neurodegenerative disorders of the central nervous system, e.g. nootropic agents, cognition enhancers, drugs for treating Alzheimer's disease or other forms of dementia

Abstract

The invention discloses a traditional Chinese medicine composition for preventing and treating Alzheimer's disease, which comprises 10-30 parts by weight of neferine, 5-10 parts by weight of farrerol, 5-10 parts by weight of alpha-asarone, 10-20 parts by weight of verbascoside and 10-30 parts by weight of naringin, wherein the weight ratio of the naringin to the farrerol is 1: (0.5-0.6). The invention also discloses application of the traditional Chinese medicine composition for preventing and treating the Alzheimer's disease in learning memory protection, learning memory improvement and PC12 cell protection. The traditional Chinese medicine composition for preventing and treating the Alzheimer disease and the application thereof provided by the invention can obviously increase the object recognition index of a scopolamine model mouse; the escape latency of a scopolamine model mouse is obviously shortened, and the platform crossing times and the target quadrant residence time are obviously increased; the incubation period and the error times of the scopolamine model mouse are obviously increased.

Description

Traditional Chinese medicine composition for preventing and treating Alzheimer's disease and application thereof
Technical Field
The invention relates to a traditional Chinese medicine composition for preventing and treating Alzheimer's disease and application thereof, belonging to the technical field of pharmacy.
Background
Alzheimer's Disease (AD) is a central nervous system disease, and its main clinical manifestations include gradual memory loss, cognitive dysfunction, language expression disorder, disability, mood and behavior changes. According to the international conference on the Alzheimer's Association in 2018, 1 dementia patient was produced every 3 seconds in the world. With the increasing aging degree of the population, more people will suffer from AD, and the health and the life quality of the old people are seriously affected. However, the pathogenesis of AD is not yet elucidated, and the existing drugs for treating AD only slightly improve the clinical symptoms of patients, and cannot achieve the effects of prevention and cure. Therefore, the definition of the pathogenesis of AD and the search for effective therapeutic agents are reluctant.
Disclosure of Invention
In order to solve the technical problems mentioned in the background technology, the invention provides a traditional Chinese medicine composition for preventing and treating Alzheimer's disease and application thereof.
In order to solve the technical problems, the technical scheme adopted by the invention is as follows:
the traditional Chinese medicine composition for preventing and treating Alzheimer's disease comprises 10-30 parts by weight of neferine, 5-10 parts by weight of azalea, 5-10 parts by weight of alpha-asarone, 10-20 parts by weight of verbascoside and 10-30 parts by weight of naringin, wherein the weight ratio of the naringin to the azalea is 1: (0.5-0.6).
The traditional Chinese medicine composition can be prepared into one or more of oral liquid, powder, tablets, capsules and pills.
The traditional Chinese medicine composition comprises auxiliary agents within a pharmaceutically allowable range, wherein the auxiliary agents are one or more of wetting agents, bacteriostats, emulsifying agents, antioxidants, solubilizing agents, thickening agents, suspending agents, excipients, sweetening agents or stabilizing agents.
The application of the traditional Chinese medicine composition for preventing and treating Alzheimer's disease in learning and memory protection.
The application of the traditional Chinese medicine composition for preventing and treating Alzheimer's disease in learning and memory improvement.
Application of a traditional Chinese medicine composition for preventing and treating Alzheimer's disease in PC12 cell protection.
The traditional Chinese medicine composition is applied to the regulation of an ER-P38/MAPK signal pathway.
The traditional Chinese medicine composition can be used for being combined with an anti-AD medicine for delaying the development of AD diseases.
Neferine (Trolline) with molecular formula of C12H13NO3Molecular weight is 219, and it is colorless flaky crystal. Is derived from flos Trollii of Ranunculaceae; the Chinese globeflower has the alias: eclipta alba, callicarpa pedunculata and keloid; the name learning: trollius chinensis Bunge is a plant of the genus globeflower of the family Ranunculaceae. Annual or perennial herbs, trollius chinensis is cold resistant, exists at 2-15 ℃ throughout the year, and widely grows in the north of China. The Chinese globeflower has the medicinal functions, can clear away heat and toxic materials, is mainly used for treating tonsillitis and otitis media, is used for treating acute and chronic tonsillitis, acute otitis media, acute myringitis, acute conjunctivitis, acute lymphangitis and the like, and also has antiviral activity.
Farrerol (Farrerol), alternative name: 5, 7-dihydroxy-2- (4-hydroxyphenyl) -6, 8-dimethyl-2, 3-dihydrobenzopyran-4-one or farrerol, with the molecular formula C17H16O5The molecular weight is 300, the farrerol is a medicine extracted from rhododendron dauricum and other rhododendron plants and has the effect of eliminating phlegm, the medicine is synthesized artificially at present, the medicine directly acts on respiratory mucosa, promotes cilia movement, enhances the function of mechanically clearing foreign matters from trachea and bronchus, can break acid glycoprotein fibers in phlegm, reduce sialic acid content, reduce phlegm viscosity, thin phlegm and easy expectoration, and the medicine is also used for treating phlegm deficiency, cough, asthma, coughThe amount of phlegm gradually decreases. Has definite effect of eliminating phlegm for patients with chronic tracheitis, and has lasting and stable curative effect, thereby obviously reducing the amount of sputum, reducing the viscosity and reducing the symptoms. The traditional Chinese medicine composition is mainly used for treating excessive and viscous phlegm and the like caused by chronic bronchitis in clinic.
α -Asarone (named as alpha-Asarone) with molecular formula of C12H16O3α -asarone is the main effective component of traditional Chinese medicine grassleaf sweelflag rhizome, and has the functions of relieving asthma, relieving cough, eliminating phlegm, tranquilizing, relieving spasm, resisting convulsion, etc. clinically, it is used for adult and children bacterial pneumonia, intrapulmonary infection, acute and chronic bronchitis, bronchial asthma, obstructive emphysema, pulmonary heart disease, bronchiectasis and bronchopulmonary carcinoma, and cough, expectoration and wheeze caused by cold.
Verbascoside (Verbascoside), alternative name: acteoside and acteoside with molecular formula C29H36O15The extract is white needle crystal powder, is easily soluble in ethanol, methanol, and ethyl acetate, and is extracted from dry fleshy stem with scale leaf of Cistanchis herba Deserticola Y.C.Ma of Orobanchaceae. Acteoside is ATP competitive PKC inhibitor, IC50Has a value of 25 μ M, and has antitumor, antiinflammatory, and neuropathic pain relieving effects.
The naringin is totally called naringenin-7-O-neohesperidoside, is a dihydroflavonoid compound, is one of the main effective components of the traditional Chinese medicine rhizoma drynariae for tonifying the kidney, and researches show that the naringin has stronger biological activity in the aspects of reducing blood fat, calming, resisting oxidation, tumors, fungi, atherosclerosis, spasmolysis, easing pain, regulating blood sugar and the like.
The rhododendron and the alpha-asarone have the effect of eliminating phlegm, the nekaline and the verbascoside have the anti-inflammatory effect, the naringin has the effect of tonifying the kidney, and the alpha-asarone has the effect of tonifying the brain. The five traditional Chinese medicines are combined together to play the roles of tonifying the kidney and benefiting the brain, and removing blood stasis and removing phlegm, thereby providing a new treatment idea for AD.
Estrogen is a broadly active steroid hormone secreted by the ovary, in a variety of forms, such as estrone, estradiol (E2) and estriol (E3). The effects of estrogens are mainly mediated through Estrogen Receptors (ER). ER includes two subtypes, ER alpha and ER beta. ER alpha is mainly present in hypothalamus and amygdala, ER beta is mainly distributed in hippocampus, cerebral cortex and olfactory lobe, and the brain areas are all related to cognitive functions and the like, which suggests that estrogen is closely related to memory, emotion and cognitive functions. Many epidemiological studies have shown that AD prevalence is significantly different between sexes, with postmenopausal women having a prevalence significantly higher than that of age-matched men by 1.5-3 times that of men. These results reveal that estrogen is closely associated with the onset of AD. It is also postulated that decreased levels of estrogen and progestogen in women lead to increased incidence of AD. Meanwhile, the artificial synthesis of estrogen has many side effects in treating AD, such as the increase of the incidence rate of cardiovascular events and breast cancer. It is therefore important to find safe and effective therapeutic drugs.
Phytoestrogens (PE) are plant-derived compounds similar in structure to the human endogenous estrogen 17- β -estradiol, PE is similar to estrogen and can also bind to the in vivo estrogen receptor ER, PE natural drugs are used as substitutes for estrogen, have a wide and important role in the central nervous system, and have small toxic and side effects, such as reducing a β production, improving cholinergic nerve function, resisting oxidative stress and scavenging free radicals, reducing hyperphosphorylation of Tau protein, inhibiting apoptosis, improving cerebral blood circulation and cerebral metabolism, and the like.
The neferine, farrerol, alpha-asarone, verbascoside and naringin of the present invention are all commercially available.
The application of the combination of neferine, farrerol, alpha-asarone, verbascoside and naringin in the aspect of treating AD is not reported at present.
Detailed Description
In order to make the objects, technical solutions and advantages of the present invention more clear, the present invention is further described in detail with reference to the following embodiments. The specific embodiments described herein are merely illustrative of the invention and are not intended to be limiting.
Example 1
A traditional Chinese medicine composition for preventing and treating Alzheimer's disease comprises 10 parts by weight of neferine, 5 parts by weight of rhododendron, 5 parts by weight of alpha-asarone, 1 part by weight of verbascoside and 10 parts by weight of naringin, wherein the weight ratio of the naringin to the rhododendron is 1: 0.5.
the traditional Chinese medicine composition can be prepared into one or more of oral liquid, powder, tablets, capsules and pills.
The application of the traditional Chinese medicine composition for preventing and treating Alzheimer's disease in learning and memory protection.
The application of the traditional Chinese medicine composition for preventing and treating Alzheimer's disease in learning and memory improvement.
Application of a traditional Chinese medicine composition for preventing and treating Alzheimer's disease in PC12 cell protection.
The application of the traditional Chinese medicine composition for preventing and treating Alzheimer's disease and the application of the traditional Chinese medicine composition in regulating ER-P38/MAPK signal pathways.
The research on the learning and memory protection effects of the traditional Chinese medicine composition on the scopolamine model mouse comprises the following steps: taking 3-month-old male Kunming mice, and randomly dividing the mice into a blank group, a scopolamine group and a traditional Chinese medicine composition group according to body weight. The mice in the blank groups of the experiment days 1-21 and the scopolamine group are subjected to intragastric administration of distilled water, the mice in the traditional Chinese medicine composition group are subjected to intragastric administration of a traditional Chinese medicine composition solution (100 mg/kg. d), the blank group of the experiment day 22 is subjected to intraperitoneal injection of normal saline, the other groups are subjected to intraperitoneal injection of the same amount of scopolamine (100 mg/kg. d), the modeling is finished on the 28 th day, the behaviourological experiment is started on the 29 th day, and the effect of the traditional Chinese medicine composition on the learning and memory capacity of the mice is observed by adopting new object recognition and water maze experiments.
New object identification experiment: the experiment was carried out in three stages: a familiarity period, a training period, and a testing period. In the familiarity phase, the mice were free to move for 5min in the box. The mouse training method comprises a training period and a testing period, wherein in the training period, two objects with the same shape, size, color and texture are placed at one end of a box body, a mouse is placed at the midpoint of the wall of the opposite side, back to the objects, and the mouse is familiar with the training period for 5 min. After 1h, a test period was followed by replacing one of the objects with another completely different object in shape, size, color and texture. Mice were placed from the same location and the contact time to the new object (Tn) and to the old object (Tf) within 5min of the mice was recorded. The recognition index (Tn-Tf)/(Tn + Tf) is used to express the recognition ability of the mouse for the object. The high or low recognition index represents the strength of the memory of the mouse.
Water maze experiment: the Morris water maze experiment comprises a positioning navigation experiment and a space exploration experiment. Positioning navigation experiment: and (3) placing the circular platform at the position 2cm under the water in the fourth quadrant, and keeping the position of the platform unchanged. Mice were placed in the water facing the pool wall at the water entry points specified in quadrants 1, 2, and 3. Within 90s, the mouse with the target platform is found and stays on the platform for 30s, and the mouse without the platform is manually placed on the platform for 30s to enhance the memory. The time for the mouse to find the platform is the escape latency. The experimental period was four days and the escape latency was recorded. Space exploration experiment: the platform was removed and the mice were placed in the water maze at quadrant 1, 2, and 3 entry points for 90s and their residence time in the target quadrant (quadrant 4) was recorded. The stage position, the number of passes of the active area (2 stage diameter area) was recorded.
The results of the novel object recognition experiments show that the traditional Chinese medicine composition can obviously increase the novel object recognition index of a scopolamine model mouse.
The water maze experiment result shows that the traditional Chinese medicine composition can obviously shorten the escape latency of a scopolamine model mouse and obviously increase the platform crossing times and the target quadrant residence time.
Secondly, the research on the learning and memory improving effects of the traditional Chinese medicine composition on scopolamine model mice: 75 Kunming mice of 3 months of age were randomly divided into 5 groups according to body weight, a blank group, a scopolamine model group (SC), an anti-encephalasthenia capsule control group (KNSC), a donepezil hydrochloride control group (DPH), a traditional Chinese medicine composition group (ZHW), a traditional Chinese medicine composition group and an ER blocking agent group (ZHW + ICI 182780). Distilled water is given to the blank group and the scopolamine model group on the 1 st to 21 th days of the experiment through intragastric administration; performing intragastric gavage on an anti-encephalasthenia capsule control group, a donepezil hydrochloride control group and a Chinese medicinal composition group respectively to obtain an anti-encephalasthenia capsule (585 mg/kg. d), a donepezil hydrochloride (0.65 mg/kg. d) and a Chinese medicinal composition (100 mg/kg. d), performing intraperitoneal injection on the Chinese medicinal composition and an ER blocking agent group to obtain an ICI182780(0.072 mg/kg. d), and performing intragastric administration on the Chinese medicinal composition and the ER blocking agent group after 15min to obtain the Chinese medicinal composition (100 mg/kg. d). The molding is started on the 22 th day, the normal saline is injected into the abdominal cavity of the blank group, the scopolamine (100 mg/kg. d) with the same amount is injected into the abdominal cavity of the other groups, the molding is finished on the 28 th day, the ethological experiment is started on the 29 th day, and the molding agent is given 30min before the ethological experiment. And respectively adopting new object identification, water maze and diving platform experiments to observe the learning and memory abilities of the mouse.
New object identification experiment: the experiment was carried out in three stages: a familiarity period, a training period, and a testing period. In the familiarity phase, the mice were free to move for 5min in the box. The mouse training method comprises a training period and a testing period, wherein in the training period, two objects with the same shape, size, color and texture are placed at one end of a box body, a mouse is placed at the midpoint of the wall of the opposite side, back to the objects, and the mouse is familiar with the training period for 5 min. After 1h, a test period was followed by replacing one of the objects with another completely different object in shape, size, color and texture. Mice were placed from the same location and the contact time to the new object (Tn) and to the old object (Tf) within 5min of the mice was recorded. The recognition index (Tn-Tf)/(Tn + Tf) is used to express the recognition ability of the mouse for the object. The high or low recognition index represents the strength of the memory of the mouse.
Water maze experiment: the Morris water maze experiment comprises a positioning navigation experiment and a space exploration experiment. Positioning navigation experiment: and (3) placing the circular platform at the position 2cm under the water in the fourth quadrant, and keeping the position of the platform unchanged. Mice were placed in the water facing the pool wall at the water entry points specified in quadrants 1, 2, and 3. Within 90s, the mouse with the target platform is found and stays on the platform for 30s, and the mouse without the platform is manually placed on the platform for 30s to enhance the memory. The time for the mouse to find the platform is the escape latency. The experimental period was four days and the escape latency was recorded. Space exploration experiment: the platform was removed and the mice were placed in the water maze at quadrant 1, 2, and 3 entry points for 90s and their residence time in the target quadrant (quadrant 4) was recorded. The stage position, the number of passes of the active area (2 stage diameter area) was recorded.
A jump bench experiment: the mouse is placed into a reaction box to adapt for 3min, then is electrified, and after the mouse is shocked, the mouse jumps to the platform. The mouse jumps to the copper grid for multiple times and quickly jumps back to the platform after being shocked, the training is carried out for 5min, after 24 hours, the mouse is directly and lightly placed on the platform under the condition that the copper grid at the bottom is electrified, the time from the mouse being placed on the platform to the mouse jumping down for the first time is recorded, the latency period (LT) is obtained, the jumping down times (n) of the mouse within 5min are recorded, and the learning and memory abilities of the mouse are judged through the LT and the n.
The results of the novel object recognition experiments show that the traditional Chinese medicine composition can obviously increase the novel object recognition index of a scopolamine model mouse.
The water maze experiment result shows that the traditional Chinese medicine composition can obviously shorten the escape latency of a scopolamine model mouse and obviously increase the platform crossing times and the target quadrant residence time.
The results of the diving platform experiments show that the traditional Chinese medicine composition can obviously increase the latency and the error times of the scopolamine model mouse.
Thirdly, the Chinese medicinal composition pair A β25-35Study of protective effects of induced injury PC12 cells:
1 materials of the experiment
1.1 cell source: PC12 cells (high differentiation, cat # ZQ0150) were purchased from Geohio-Xinzhou Biotech, Inc. in Shanghai.
1.2 preparation of reagents and medicines:
preparing a high-glucose DMEM culture medium: 89% DMEM medium, 10% fetal bovine serum and 1% penicillin-streptomycin are mixed uniformly and stored at 4 ℃.
E2The solution is prepared by weighing 10.90mg of estradiol, adding 10 μ L DMSO for assisting dissolution, adding DMEM complete culture medium for dissolving to constant volume of 20mL, and preparing into 2 × 10-3Diluting the mother liquor of mol/L to 2 × 10 according to the requirement-9Filtering the working solution with mol/L concentration by a 0.22 mu m filter membrane, and storing at-20 ℃ for later use.
Aβ25-35The solution was prepared by mixing 5mg of A β25-35The powder was dissolved in 15. mu.L DMSO and then 31.438mLPBS was added to prepare a 150. mu. mol/L stock solution at 37 ℃ in 5% CO2The mixture was allowed to stand in an incubator for one week, aged and coagulated, and stored at-20 ℃. When used, the solution is diluted to a final concentration of 20. mu. mol/L.
The Chinese medicinal composition solution is prepared by precisely weighing 23.06mg powder, adding 20 μ L DMSO for dissolving, and preparing into 2 × 10 with 20mL DMEM culture solution-3mol/L mother liquor, diluted 1 × 10 before experiment-6The mol/L concentration is stored at-20 ℃ for later use.
Preparation of ER blocker (ICI182780) solution: 3.00mg was weighed precisely, and ICI182780 stock solution with 20. mu. mol. L-1 final concentration was prepared with DMEM culture solution, sterilized by filtration with 0.22 μm filter membrane, and stored at-20 ℃ for further use. Diluting to the required use concentration of 1 mu mol/L before use, and preheating at 37 ℃.
Preparation of P38 blocker (SB203580) solution: 3.80mg of SB203580 solid powder was weighed, prepared into SB203580 stock solution with a final concentration of 200. mu. mol/L using DMEM complete culture solution, sterilized by filtration using a 0.22 μm filter membrane, and stored at-20 ℃ for further use. Diluting to the required 10 mu mol/L use concentration before use, and preheating at 37 ℃.
2 method of experiment
2.1 revival, culture, passage and cryopreservation of PC12 cells
Placing the cells taken out from the liquid nitrogen in a 37 ℃ water bath kettle, rapidly shaking for 1min to melt the cells to avoid the ice crystals from puncturing the cells, transferring the cells into a 15ml centrifuge tube after melting, adding 3-5ml of DMEM complete culture medium, gently mixing the cells uniformly, centrifuging at 1000rpm for 5min, adding 3-7ml of DMEM complete culture medium for resuspension, sucking 1ml of resuspension solution, placing the resuspension solution in a 25cm container2Adding 3-6ml DMEM complete medium into a culture flask, placing at 37 ℃ and 5% CO2In an incubator, each timeThe cells were passaged at a rate of 1: 4 or more after 24h replacement until the cells grew to 80% or more, using 0.25% trypsinized cells. After digesting and centrifuging cells growing to 80% state, the cells were resuspended in complete culture medium containing 8% DMSO, 10% fetal bovine serum and 82% DMEM, and the suspension was transferred to a freezing tube at 1ml per tube. Gradually cooling in a programmed cooling box, standing overnight at-80 deg.C, and storing in liquid nitrogen tank (-196 deg.C) for a long period.
2.2 Experimental groups and dosing
Blank group: logarithmic phase cells seeded at 25cm2And (3) incubating in a culture bottle or a pore plate for 24 hours, then discarding waste liquid, and continuously adding fresh culture solution for culturing for 24 hours.
Model group (A β)25-35): logarithmic phase cells seeded at 25cm2Incubating in culture flask or well plate for 24 hr, removing waste liquid, and adding fresh DMEM to obtain complete culture medium with concentration of 1.5 × 10-4mol/L of A β25-35Mixing to obtain final concentration of 2 × 10-5mol/L of A β25-35The culture was continued for 24 h.
Estradiol + A β25-35Group (E)2+Aβ25-35): logarithmic phase cells seeded at 25cm2Incubating in culture flask or well plate for 24 hr, removing waste liquid, and adding fresh DMEM complete culture medium at concentration of 2 × 10-9mol/L of E2The solution is mixed evenly to prepare the final concentration of 1 × 10-9mol/L of E2Culturing in solution for 2h, and adding 1.5 × 10-4mol/L of A β25-35At this time, 1.5 × 10-4mol/L of A β25-35、2×10-9mol/L of E2The solution is mixed with the rest culture solution in the pore plate to prepare the final concentration of 2 × 10-5mol/L of A β25-35And 1 × 10-9mol/L of E2The solution was incubated for a further 22 h.
Chinese medicinal composition + A β25-35Group (ZHW + A β)25-35): logarithmic phase cells seeded at 25cm2Incubating for 24 hr in a culture flask, discarding waste liquid, and transferring into 2 × 10-7Culturing in solution of Chinese medicinal composition at mol/L concentration for 2 hr, and adding 1.5 × 10-4mol/L of A β25-35Mixing with the rest culture solution to obtain a final concentration of 20 μmol/LAβ25-35The cultivation was continued for 22 h.
Chinese medicinal composition + A β25-35+ ER blocker group (ZHW + A β)25-35+ ICI 182780): logarithmic phase cells seeded at 25cm2Culturing in a culture flask for 24h, changing to ICI182780 culture solution with final concentration of 1 μmol/L, culturing for 1h, and changing to 2 × 10- 7Culturing in solution of Chinese medicinal composition at mol/L concentration for 2 hr, and adding 1.5 × 10-4mol/L of A β25-35Mixing with the rest culture solution to obtain A β with final concentration of 20 μmol/L25-35The cultivation was continued for 22 h.
Chinese medicinal composition + A β25-35+ P38 blocker group (ZHW + A β)25-35+ SB 203580): logarithmic phase cells seeded at 25cm2Culturing in culture flask for 24h, changing to SB203580 culture solution with final concentration of 10 μmol/L, culturing for 1h, and changing to 2 × 10- 7Culturing in solution of Chinese medicinal composition at mol/L concentration for 2 hr, and adding 1.5 × 10-4mol/L of A β25-35Mixing with the rest culture solution to obtain A β with final concentration of 20 μmol/L25-35The cultivation was continued for 22 h.
3 results of the experiment
3.1 Chinese medicinal composition and blocker pair A β25-35Effect of damaged PC12 cell proliferation Rate
The results are shown in Table 1, A β compared to the blank25-35The proliferation rate of the cells in the group is very obviously reduced (P is less than 0.01), and A β25-35Group comparison, ZHW groups, E2The proliferation rate of the cells in the group is extremely obviously increased (P is less than 0.01) and the cells show protective effect, and compared with the ZHW group, the ICI182780+ ZHW + A β25-35The proliferation rate of the cells in the group is very obviously reduced (P is less than 0.01), which indicates that the traditional Chinese medicine composition plays a role by activating ER, and compared with the ZHW group, the SB203580+ ZHW + A β25-35The proliferation rate of the group cells is extremely obviously reduced (P is less than 0.01), which indicates that the traditional Chinese medicine composition is A β25-35Function by activating the P38/MAPK signaling pathway.
TABLE 1, ZHW and blocker Pair A β25-35Effect of impaired proliferation Rate of PC12 cells (
Figure BDA0002620707250000091
n＝6)
Figure BDA0002620707250000092
Note: p compared to blank group<0.05 as P<0.01, and A β25-35Group comparison, # # is P<0.01; in comparison with the ZHW group, the test piece was,&&is P<0.01。
Example 2
The traditional Chinese medicine composition for preventing and treating the Alzheimer's disease comprises 30 parts by weight of neferine, 9 parts by weight of rhododendron, 10 parts by weight of alpha-asarone, 20 parts by weight of verbascoside and 15 parts by weight of naringin, wherein the weight ratio of the naringin to the rhododendron is 1: 0.6.
example 3
A traditional Chinese medicine composition for preventing and treating Alzheimer's disease comprises 20 parts by weight of neferine, 10 parts by weight of azalea, 8 parts by weight of alpha-asarone, 15 parts by weight of verbascoside and 30 parts by weight of naringin, wherein the weight ratio of the naringin to the azalea is 3: 1.
example 4
A traditional Chinese medicine composition for preventing and treating Alzheimer's disease comprises 15 parts by weight of neferine, 10 parts by weight of rhododendron, 7 parts by weight of alpha-asarone, 12 parts by weight of verbascoside and 20 parts by weight of naringin, wherein the weight ratio of the naringin to the rhododendron is 1: 0.5.
example 5
A traditional Chinese medicine composition for preventing and treating Alzheimer's disease comprises 15 parts by weight of neferine, 10 parts by weight of azalea, 9 parts by weight of alpha-asarone, 18 parts by weight of verbascoside and 16.7 parts by weight of naringin, wherein the weight ratio of the naringin to the azalea is 1: 0.6.
the foregoing shows and describes the general principles, essential features, and advantages of the invention. It will be understood by those skilled in the art that the present invention is not limited to the embodiments described above, which are described in the specification and illustrated only to illustrate the principle of the present invention, but that various changes and modifications may be made therein without departing from the spirit and scope of the present invention, which fall within the scope of the invention as claimed. The scope of the invention is defined by the appended claims and equivalents thereof.

Claims (6)

1. The traditional Chinese medicine composition for preventing and treating Alzheimer's disease is characterized by comprising the following components in parts by weight: the composition comprises 10-30 parts by weight of neferine, 5-10 parts by weight of farrerol, 5-10 parts by weight of alpha-asarone, 10-20 parts by weight of verbascoside and 10-30 parts by weight of naringin, wherein the weight ratio of the naringin to the farrerol is 1: (0.5-0.6).
2. The traditional Chinese medicine composition for preventing and treating alzheimer's disease according to claim 1, which is characterized in that: the traditional Chinese medicine composition can be prepared into one or more of oral liquid, powder, tablets, capsules and pills.
3. The use of the Chinese medicinal composition for preventing and treating Alzheimer's disease according to any one of claims 1-2 in learning and memory protection.
4. The use of the Chinese medicinal composition for preventing and treating Alzheimer's disease according to any one of claims 1-2 in learning and memory improvement.
5. The use of the Chinese medicinal composition for preventing and treating Alzheimer's disease according to any one of claims 1-2 in PC12 cytoprotection.
6. The use of the Chinese medicinal composition for preventing and treating Alzheimer's disease according to claim 5, wherein: the traditional Chinese medicine composition is applied to the regulation of an ER-P38/MAPK signal pathway.
CN202010782377.2A 2020-08-06 2020-08-06 Traditional Chinese medicine composition for preventing and treating Alzheimer's disease and application thereof Pending CN111759855A (en)

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Cited By (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN115501223A (en) * 2022-10-08 2022-12-23 青岛大学 Application of farrerol in preparation of medicine for resisting cerebral ischemia reperfusion injury

Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
KR20160004688A (en) * 2014-07-04 2016-01-13 이범천 A cosmetic composition containing extracts of Trollius hondoensis
CN107693530A (en) * 2017-09-03 2018-02-16 江西青峰药业有限公司 Aurantiamarin and/or aurantiin are preparing the application in improving cognition dysfunction medicine
CN110840866A (en) * 2018-08-20 2020-02-28 成都新睿泰康科技有限公司 Asarin pharmaceutical composition and application thereof in preventing and treating neurodegenerative diseases

Patent Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
KR20160004688A (en) * 2014-07-04 2016-01-13 이범천 A cosmetic composition containing extracts of Trollius hondoensis
CN107693530A (en) * 2017-09-03 2018-02-16 江西青峰药业有限公司 Aurantiamarin and/or aurantiin are preparing the application in improving cognition dysfunction medicine
CN110840866A (en) * 2018-08-20 2020-02-28 成都新睿泰康科技有限公司 Asarin pharmaceutical composition and application thereof in preventing and treating neurodegenerative diseases

Non-Patent Citations (5)

* Cited by examiner, † Cited by third party
Title
CUI B. ET AL: "Farrerol attenuates β-amyloid-induced oxidative stress and inflammation through Nrf2/Keap1 pathway in a microglia cell line", 《BIOMEDICINE & PHARMACOTHERAPY》 *
石效荣 等: "HPLC法同时测定天麻醒脑胶囊中9种成分", 《中成药》 *
董芝欣 等主编: "《老年医疗保健手册》", 31 August 2003, 中国海洋大学出版社 *
谢文英: "《养生茶饮新知全书》", 31 May 2013, 陕西科学技术出版社 *
邢海燕 等: "BDNF乙酰化修饰参与毛蕊花糖苷对Aβ25-35致PC12细胞损伤的保护作用", 《中国新药杂志》 *

Cited By (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN115501223A (en) * 2022-10-08 2022-12-23 青岛大学 Application of farrerol in preparation of medicine for resisting cerebral ischemia reperfusion injury
CN115501223B (en) * 2022-10-08 2024-01-16 青岛大学 Application of azalea essence in preparing medicine for resisting cerebral ischemia reperfusion injury

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Application publication date: 20201013