A Chinese medicine composition for treating Alzheimer's disease and its preparation method and application

Abstract

The present invention discloses a Chinese medicine composition for preventing and / or treating Alzheimer's disease, wherein the raw materials of the Chinese medicine composition include Zanthoxylum bungeanum and Rehmannia glutinosa. The preparation method of the Chinese medicine composition for treating Alzheimer's disease provided by the present invention adopts easy-to-obtain raw materials, is simple to operate, has stable technology, can maximize the efficacy of the raw material components, and the prepared pharmaceutical composition has good effect and great commercial value; the Chinese medicine compound composition provided by the present invention can improve the lateral ventricular injection of Aβ 25‑35 As well as the spatial and non-spatial learning and memory abilities of AD dementia mice caused by the intraperitoneal injection model of scopolamine.

Description

Technical Field

[0001] The present invention relates to the field of traditional Chinese medicine, and in particular to a traditional Chinese medicine composition for treating Alzheimer's disease, and a preparation method and application thereof. Background Art

[0002] Alzheimer's Disease (AD) is a neurodegenerative disease with slowly progressive cognitive decline. Its onset is insidious, symptoms develop gradually and progressively worsen, and clinical manifestations include decreased cognitive ability and behavioral disorders. The main pathological features of Alzheimer's disease are the deposition of β-amyloid protein (Aβ) and neurofibrillary tangles in the brain. Modern medicine has conducted many years of research on the pathogenesis of Alzheimer's disease, but due to its complex etiology, the pathogenesis of the disease is still unclear. Traditional Chinese medicine therapy focuses on comprehensive treatment strategies, which can exert the synergistic effects of multiple components through multiple targets to achieve the effect of slowing down AD.

[0003] At present, the first type of drug used in clinical practice of Western medicine to improve symptoms is cholinesterase inhibitor (AChEI), and the other is excitatory amino acid receptor (NMDA) antagonist, which can only improve symptoms in the short term and have no effect on the onset and course of AD.

[0004] Patent document CN111110749A clearly discloses a traditional Chinese medicine composition for treating Alzheimer's disease and its preparation method. The composition is composed of raw rehmannia root, cooked rehmannia root, dodder seed, cornus fruit, ginseng, stir-fried atractylodes, astragalus, and kadzu vine. Through the reasonable proportion of each medicinal material and the synergistic effect, it has a clear therapeutic effect on the treatment of Alzheimer's disease.

[0005] Patent document CN109999155A discloses a Chinese medicine composition for treating Alzheimer's disease and a preparation method thereof, wherein the Chinese medicine composition is prepared from the following raw materials: Radix Rehmanniae Preparata, Semen Alpiniae Oxyphyllae, Radix Astragali, Semen Cuscutae, Rhizoma Chuanxiong, Herba Mori Parasiticae, Rhizoma Cassiae, Eucommia, Pheretima, Leech, Rhizoma Gastrodiae, and Rhizoma Uncariae. The invention provides a Chinese medicine composition for treating Alzheimer's disease with significant efficacy and low cost.

[0006] Traditional Chinese medicine has the advantages of overall regulation, multi-link, multi-system and multi-target. my country's rich Chinese medicine resources have great potential for the treatment of Alzheimer's disease. At present, there is no report in the prior art on the use of Rehmannia glutinosa and Zanthoxylum bungeanum extracts to form a Chinese medicine composition for the treatment of AD. In view of this, the present invention is specially proposed. Summary of the invention

[0007] The present invention overcomes the defects in the prior art and provides a Chinese medicine composition for preventing and / or treating Alzheimer's disease and a preparation method and application thereof.

[0008] A first aspect of the present invention provides a traditional Chinese medicine composition for preventing and / or treating Alzheimer's disease, wherein the raw materials of the traditional Chinese medicine composition include Zanthoxylum bungeanum and Rehmannia glutinosa.

[0009] Furthermore, the raw materials of the traditional Chinese medicine composition are composed of Zanthoxylum bungeanum and Rehmannia glutinosa.

[0010] The Chinese medicine composition of the present invention is prepared by mixing the powder of raw material medicine or the extract of raw material medicine.

[0011] Furthermore, the Rehmannia root is selected from one or a combination of two or more of fresh Rehmannia root, raw Rehmannia root and cooked Rehmannia root.

[0012] In one embodiment of the present invention, the Rehmannia root is fresh Rehmannia root.

[0013] In one embodiment of the present invention, the Rehmannia root is fresh Rehmannia root juice.

[0014] In one embodiment of the present invention, the Rehmannia root is a concentrate of fresh Rehmannia root juice.

[0015] In one embodiment of the present invention, the Rehmannia root is a powder obtained by drying fresh Rehmannia root juice concentrate.

[0016] Furthermore, the drying is selected from the group consisting of freeze drying, normal pressure drying, reduced pressure drying, boiling drying, spray drying, infrared drying, and microwave drying.

[0017] Furthermore, the pepper can be red, selected from the peel part of the pepper.

[0018] Furthermore, the Zanthoxylum bungeanum is a Zanthoxylum bungeanum extract.

[0019] Furthermore, the extraction solvent of the Zanthoxylum bungeanum extract is selected from: one of water, alcohol, ketone, ether and ester.

[0020] Furthermore, the alcohol is selected from: one of ethanol solution, isopropanol solution and tert-butanol solution, the ketone is acetone solution, the ether is ether solution or petroleum ether solution, and the ester is ethyl acetate solution or ethyl formate solution.

[0021] Preferably, the extraction solvent of the Zanthoxylum bungeanum extract is an ethanol solution. In one embodiment of the present invention, the Zanthoxylum bungeanum is a concentrate of the Zanthoxylum bungeanum extract.

[0022] In one embodiment of the present invention, the Zanthoxylum bungeanum is a powder obtained by drying a concentrate of Zanthoxylum bungeanum extract.

[0023] Furthermore, in the traditional Chinese medicine composition, the mass ratio of the raw materials Zanthoxylum bungeanum and Rehmannia glutinosa is: 19-152:300-500 (such as 80:400, 80.5:400, 81:400, 81.5:400, 82:400, 82.5:400, 83:400, 83.5:400, 84:400, 84.5:400, 85:400, 85.5:400, 86:400, 86.5:400, 87:400). 00, 87.5:400, 88:400, 88.5:400, 89:400, 89.5:400, 90:400), preferably, the mass ratio of the raw materials Zanthoxylum bungeanum and Rehmannia glutinosa is 38-114:350-450, further preferably, the mass ratio of the raw materials Zanthoxylum bungeanum and Rehmannia glutinosa is 57-95:375-425, further preferably, the mass ratio of the raw materials Zanthoxylum bungeanum and Rehmannia glutinosa is 57-95:400.

[0024] In one embodiment of the present invention, the mass ratio of the raw materials Zanthoxylum bungeanum and Rehmannia glutinosa is 87.4:400.

[0025] Furthermore, in the traditional Chinese medicine composition, the mass ratio of the raw material Zanthoxylum bungeanum extract to the Rehmannia glutinosa extract is: 5-40:60-100 (such as 5:60-100, 10:60-100, 15:60-100, 16:60-100, 17:60-100, 18:60-100, 19:60-100, 20:60-100, 21:60-100, 22:60-100, 23:60-100, 24:60-100, 25:60-100, 26:60-100, 27:60-100). -100, 28: 60-100, 29: 60-100, 30: 60-100, 31: 60-100, 32: 60-100, 33: 60-100, 34: 60-100, 35: 60-100, 36: 60-100, 37: 60-100, 38: 60-100, 39: 60-100, 40: 60-100; 5-40: 60, 5-40: 65, 5-40: 70, 5-40: 71, 5-40: 72, 5-40: 73, 5-40: 74, 5-40: 7 5, 5-40:76, ​​5-40:77, 5-40:78, 5-40:79, 5-40:80, 5-40:81, 5-40:82, 5-40:83, 5-40:84, 5-40:85, 5-40:86, 5-40:87, 5-40:88, 5-40:89, 5-40:90, 5-40:91, 5-40:92, 5-40:93, 5-40:94, 5-40:95, 5-40:96, 5-40:97, 5-40:98, 5-40:99, 5-40:90 0:100), preferably, the mass ratio of the raw material Zanthoxylum bungeanum extract to the Rehmannia glutinosa extract is 10-30:70-90, more preferably, the mass ratio of the raw material Zanthoxylum bungeanum extract to the Rehmannia glutinosa extract is 15-25:75-85, most preferably, the mass ratio of the raw material Zanthoxylum bungeanum extract to the Rehmannia glutinosa extract is 15-25:80 (such as 15:80, 16:80, 17:80, 18:80, 19:80, 20:80, 21:80, 22:80, 23:80, 24:80, 25:80).

[0026] In one embodiment of the present invention, the mass ratio of the raw material Zanthoxylum bungeanum extract to the Rehmannia glutinosa extract is 23:80.

[0027] Preferably, the Chinese medicine composition also includes pharmaceutically acceptable excipients, which are selected from: preservatives, colorants, pH regulators, buffers, antioxidants, metal ion chelators, antibacterial agents, isotonic regulators, carriers, diluents, adhesives, lubricants, sweeteners, absorbents, disintegrants, wetting agents, coating materials. Preferably, the dosage form of the Chinese medicine composition is selected from: decoctions, powders, oral liquids, tablets, capsules, pills, injections, inhalants, suppositories, emulsions, nanoparticles, gels, powders, suspensions, creams, jellies, sprays, etc.

[0028] Preferably, the Chinese medicine composition can be administered in a manner selected from: oral, enteral, subcutaneous, intramuscular, intravenous, nasal, transdermal, subconjunctival, intraocular, orbital, retro-ocular, retinal, choroidal, intrathecal, etc.

[0029] The second aspect of the present invention provides a method for preparing the Chinese medicine composition as described in the first aspect, comprising the following steps:

[0030] (1) Weigh each API according to the weight ratio;

[0031] (2) Prepare drug powder or obtain drug extract by extraction;

[0032] (3) Prepared by mixing the powder of the raw material drug or the extract of the raw material drug, or prepared by adding one or more pharmaceutically acceptable excipients to the powder of the raw material drug or the extract of the raw material drug.

[0033] The extraction method in step (2) is selected from the group consisting of: solvent extraction, percolation, decoction, reflux (such as continuous reflux), ultrasonic extraction, supercritical fluid extraction, and steam distillation;

[0034] In some embodiments of the present invention, the extraction method is selected from: percolation and decoction.

[0035] In some embodiments of the present invention, the raw material Zanthoxylum bungeanum is extracted by percolation.

[0036] In some embodiments of the present invention, the raw material Rehmannia root is extracted by a decoction method, specifically, Rehmannia root (such as fresh Rehmannia root) is squeezed to make juice, and the Rehmannia root juice is steamed, concentrated, and dried.

[0037] Furthermore, the flow rate of the diafiltration method is 0.5 to 2 mL min -1 (e.g. 0.5 mL min -1 、0.6mL·min -1 、0.7mL·min -1 、0.8mL·min -1、0.9mL·min -1 、1.0mL·min -1 、1.1mL·min -1 、1.2mL·min -1 、1.3mL·min -1 、1.4mL·min -1 、1.5mL·min -1 、1.6mL·min -1 、1.7mL·min -1 、1.8mL·min -1 、1.9mL·min -1 , 2.0mL·min -1 ), preferably 1 to 1.5 mL min -1 .

[0038] In some embodiments of the present invention, the flow rate of the diafiltration method is 1 mL min -1 .

[0039] Furthermore, the step (2) further comprises a step of soaking the raw material drug in an extraction solvent before the extraction step.

[0040] Furthermore, the soaking time is 1-20 hours (such as 1, 1.5, 2, 2.5, 3, 3.5, 4, 4.5, 5, 5.5, 6, 6.5, 7, 7.5, 8, 8.5, 9, 9.5, 10, 10.5, 11, 11.5, 12, 12.5, 13, 13.5, 14, 14.5, 15, 15.5, 16, 16.5, 17, 17.5, 18.5, 19, 19.5, 20 hours).

[0041] In one embodiment of the present invention, the soaking time is 12 hours.

[0042] Furthermore, the step (2) further comprises the steps of concentration and drying after the extraction step.

[0043] Furthermore, the concentration can be: atmospheric pressure concentration, reduced pressure concentration, thin film concentration, and multi-effect concentration.

[0044] In some embodiments of the present invention, the concentration is concentration at normal pressure.

[0045] In some embodiments of the present invention, the concentration is concentrated under reduced pressure.

[0046] Furthermore, the drying can be performed by freeze drying, normal pressure drying, reduced pressure drying, boiling drying, spray drying, infrared drying, or microwave drying.

[0047] In one embodiment of the present invention, the drying is freeze-drying.

[0048] Further, the extraction solvent in step (2) is selected from: one of water, alcohol, ketone, ether and ester, preferably, the alcohol is selected from: one of ethanol solution, isopropanol solution and tert-butanol solution, the ketone is acetone solution, the ether is ether solution or petroleum ether solution, and the ester is ethyl acetate solution or ethyl formate solution;

[0049] Preferably, the extraction solvent in step (2) is selected from one of ethanol solution, isopropanol solution and tert-butanol solution.

[0050] In one embodiment of the present invention, the extraction solvent in step (2) is an ethanol solution.

[0051] Furthermore, the concentration of the ethanol solution is 60%-80% (specifically 60%, 65%, 70%, 75%, 80%).

[0052] In one embodiment of the present invention, the concentration of the ethanol solution is 70%.

[0053] Furthermore, in step (2), the extraction times are 1 or more times, for example, 2, 3, 4, 5, 6, 7, 9, or 10 times.

[0054] Furthermore, in step (2), for a single extraction, the mass volume ratio of the raw material (Zanthoxylum bungeanum, specifically such as Zanthoxylum bungeanum peel) to the extraction solvent is 1:5-30 (specifically such as 1:5, 1:10, 1:15, 1:20, 1:25, 1:30).

[0055] In one embodiment of the present invention, the number of extractions is 1, and the mass volume ratio of the raw material (Zanthoxylum bungeanum, specifically such as Zanthoxylum bungeanum peel) to the extraction solvent is 1:20.

[0056] The third aspect of the present invention provides use of the Chinese medicine composition as described in the first aspect in the preparation of a medicament for preventing and / or treating a disease with amnesia as a main symptom.

[0057] Preferably, the disease is Alzheimer's disease.

[0058] Furthermore, the Alzheimer's disease is selected from: early-onset Alzheimer's disease, late-onset Alzheimer's disease and familial Alzheimer's disease.

[0059] Furthermore, the prevention and / or treatment of diseases with amnesia as the main symptom includes the prevention and / or treatment of: memory loss, inattention, social difficulties, inability to think independently, emotional indifference, time orientation disorder, urinary incontinence, memory disorder, aphasia, apraxia, agnosia, spatial skill impairment, executive dysfunction and personality and behavior changes.

[0060] The present invention uses a three-factor three-level orthogonal experiment to optimize the percolation method for the extraction process of Zanthoxylum bungeanum peel, and uses amide components as indicators to optimize the optimal ethanol concentration, solid-liquid ratio and percolation rate. The Zanthoxylum bungeanum peel is extracted according to the optimal percolation conditions to obtain a Zanthoxylum bungeanum extract, and fresh Rehmannia glutinosa juice is prepared by freeze-drying; the Chinese medicine composition for treating Alzheimer's disease provided by the present invention uses natural Chinese medicine as a raw material component, which is green and safe, has mild adverse reactions, is inexpensive, can be used for a long time without drug resistance, has good patient compliance, and good efficacy; the specific targeting effect and cross-function between the components are used to improve the therapeutic effect of the drug on Alzheimer's disease; the preparation method of the Chinese medicine composition for treating Alzheimer's disease provided by the present invention uses raw materials that are easy to obtain, simple to operate, stable technology, can maximize the efficacy of the raw material components, and the prepared pharmaceutical composition has good effect and great commercial value; the Chinese medicine compound composition provided by the present invention can improve the lateral ventricular injection of Aβ 25-35 The spatial learning and memory ability of AD dementia mice caused by the invention can be improved. The rate of entering the stage in the water maze experiment of the dementia mice can be increased, the latency period can be shortened, the number of target quadrant crossings and the target quadrant swimming distance can be increased; the number of central area explorations, the central area exploration distance and the central area exploration time in the open field experiment can be increased; the Chinese medicine compound composition provided by the invention and its formula components can improve the learning and memory disorders of AD mice induced by intraperitoneal injection of scopolamine. The long-term spatial memory of the dementia mice can be improved, the rate of entering the stage in the water maze experiment can be increased, the latency period can be shortened, the number of target quadrant crossings and the target quadrant swimming distance can be increased, the short-term non-spatial memory of the dementia mice can be improved, the novel object discrimination index in the new object recognition experiment can be increased, the passive avoidance reaction of the dementia mice can be improved, the number of dark avoidance errors and the number of probes in the dark avoidance experiment can be reduced, and the dark avoidance error latency can be increased. In the above experiments, the Chinese herbal compound composition has the most significant effect; the Chinese herbal compound composition provided by the present invention and its formula components can improve the cholinergic disorder and increased oxidative stress level associated with learning and memory disorders in AD mice induced by intraperitoneal injection of scopolamine, increase the acetylcholine content and acetylcholine transferase activity in the cerebral cortex of dementia mice, reduce the acetylcholinesterase activity, and reduce the malondialdehyde content in the serum of dementia mice, and the above improvement trends are positively correlated with the improvement of learning and memory ability, which proves that the Chinese herbal compound composition plays a major role in improving AD learning and memory, and its mechanism is related to regulating the cholinergic system and alleviating oxidative stress. BRIEF DESCRIPTION OF THE DRAWINGS

[0061] Figure 1 This is an experimental flow chart for studying the improving effect of a Chinese medicine compound on the learning and memory abilities of dementia mice induced by intraventricular injection of Aβ25-35.

[0062] Figure 2 This is a behavioral study on mice in a positioning navigation experiment (Figure A shows the effect of different doses of the Chinese herbal compound composition on the rate of mice landing on the platform; Figure B shows the effect of different doses of the Chinese herbal compound composition on the escape latency of mice; Figure C shows the effect of different doses of the Chinese herbal compound composition on the percentage of swimming time of mice; Figure D shows the effect of different doses of the Chinese herbal compound composition on the percentage of swimming distance of mice).

[0063] Figure 3 This is the effect of this Chinese herbal compound on the number of platform crossings (A) and the target quadrant travel distance (B) in the space exploration experiment.

[0064] Figure 4 This is the effect of different doses of Chinese herbal compound composition on the total distance traveled, number of central area explorations, exploration time and exploration distance of mice in the open field test.

[0065] Figure 5 This is an experimental flow chart for studying the improving effect of a Chinese medicine compound and its ingredients on the learning and memory abilities of dementia mice induced by intraperitoneal injection of SCOP.

[0066] Figure 6 To study the effects of the Chinese herbal compound and its components on the open field spontaneous activity of SCOP model mice (Figure A shows the effects of the Chinese herbal compound and its components on the total movement distance of mice; Figure B shows the effects of the Chinese herbal compound and its components on the average speed of mice).

[0067] Figure 7 To study the effects of the Chinese herbal compound and its components on the spatial memory ability of SCOP model mice in the water maze (Figure A shows the effects of the Chinese herbal compound and its components on the stage rate of mice; Figure B shows the effects of the Chinese herbal compound and its components on the escape latency of mice; Figure C shows the effects of the Chinese herbal compound and its components on the percentage of swimming time of mice; Figure D shows the effects of the Chinese herbal compound and its components on the percentage of swimming distance of mice).

[0068] Figure 8 To study the effects of a Chinese medicine compound and its ingredients on the novel object discrimination ability of SCOP model mice.

[0069] Fig. 9To study the effects of the Chinese herbal compound and its components on the passive avoidance memory of electrical stimulation in SCOP model mice (Figure A shows the effects of the Chinese herbal compound and its components on the latency of dark-avoidance errors in mice; Figure B shows the effects of the Chinese herbal compound and its components on the number of dark-avoidance errors in mice; Figure C shows the effects of the Chinese herbal compound and its components on the number of dark-avoidance probes in mice).

[0070] Fig.10 To study the effects of the Chinese herbal compound composition and its components on the cholinergic system of SCOP model mice (Figure A shows the effects of the Chinese herbal compound composition and its components on the ACh content in the cerebral cortex of SCOP model mice; Figure B shows the effects of the Chinese herbal compound composition and its components on the ChAT activity in the cerebral cortex of SCOP model mice; Figure C shows the effects of the Chinese herbal compound composition and its components on the AChE activity in the cerebral cortex of SCOP model mice).

[0071] Fig.11 To study the effects of a traditional Chinese medicine compound and its components on the content of oxidative stress-related malondialdehyde in the serum of SCOP model mice. DETAILED DESCRIPTION

[0072] In order to more clearly understand the technical content of the present invention, the following embodiments are given in detail, the purpose of which is only to better understand the content of the present invention but not to limit the protection scope of the present invention.

[0073] Example 1 Optimization of the preparation process of Chinese medicine compound composition

[0074] 1. Experimental Materials

[0075] 1.1 Main experimental instruments and equipment

[0076] Table 1

[0077]

[0078] 1.2 Experimental reagents and consumables

[0079] Table 2

[0080]

[0081] 1.3 Experimental drugs

[0082] Table 3

[0083]

[0084]

[0085] 2. Preparation process

[0086] 2.1 Determination of the content of active ingredients in Zanthoxylum bungeanum extract by HPLC

[0087] (1) Testing conditions

[0088] The mobile phase gradient was 0.1% formic acid: water (A) and acetonitrile (B), and the chromatographic gradient elution conditions were shown in Table 4. The flow rate was set to 1 mL min-1, the column temperature was 30 °C, the detection wavelength was dual wavelength at 284 nm and 360 nm, the injection chamber temperature was 4 °C, and the injection volume was 20 μl.

[0089] Table 4 Chromatographic gradient elution conditions

[0090]

[0091] (2) Preparation of standard products

[0092] Preparation of mixed standard: Weigh 1 mg of hydroxy-α-sanshool, hydroxy-β-sanshool, hydroxy-γ-isosanshool, rutin, hyperoside, and hesperidin respectively in a 10 mL volumetric flask, add 50% methanol to dissolve, make up to volume, and prepare a mixed standard stock solution of 6 components with a concentration of 100 μg / mL. Accurately pipette the mixed standard stock solution prepared above, make up to volume with methanol, and mix thoroughly to obtain mixed standard solutions with mass concentrations of 0.3125, 0.625, 1.25, 2.5, 5, 10, 20, 40, and 80 μg / mL, respectively.

[0093] (3) Preparation of test solution

[0094] Take 0.1g of the test sample, add 1mL (50% methanol) to dissolve, mix well. Dilute the test sample to 300μg / mL with methanol, centrifuge at 13000rpm / min, centrifuge for 5 minutes, take 200μL of the supernatant and put it in the inner liner tube for injection.

[0095] (4) Orthogonal experimental design

[0096] Preliminary experiments have found that pepper ethanol extracts have a strong anti-inflammatory effect in vitro. Therefore, this experiment uses ethanol as the extraction solvent; selects ethanol concentration (A), solid-liquid ratio (B), and percolation rate (C) as factors, and designs the experimental design L9 (3 4 ) Arrange an orthogonal experiment. The factor levels are shown in Table 5.

[0097] Table 5L9(3 4 )Orthogonal test factor level

[0098]

[0099] (5) Extraction and separation of Zanthoxylum bungeanum

[0100] Take 50g of Zanthoxylum bungeanum peel (take the red peel part, remove the white part and the eye part), divide it into 9 groups, place them in a percolation cylinder, soak them in ethanol for 12 hours and then percolate. Collect the percolation liquid, concentrate it at 55℃ with a rotary evaporator, and transfer it to a constant weight evaporating dish after concentrating it to no ethanol. Then put it in a -80℃ low-temperature refrigerator and freeze it for 10 hours. Quickly transfer it to a pre-cooled vacuum freeze dryer for freeze drying, weigh it, and subtract the weight of the evaporating dish to obtain the weight of the dry paste. Vacuum seal the dry paste and store it for later use.

[0101] (6) Calculation of active ingredients in Zanthoxylum bungeanum extract

[0102] According to the HPLC content determination method established under 2.1, the content of the active ingredients in each group of Zanthoxylum bungeanum extracts was determined.

[0103] Extraction rate calculation formula:

[0104]

[0105] Y = active ingredient extraction rate mg / g, c = active ingredient concentration μg / mL, f = dilution factor, v = constant volume mL, M = mass of Zanthoxylum bungeanum peel g, t = extract extraction rate.

[0106] (7) Preparation of fresh Rehmannia glutinosa juice concentrate

[0107] Take fresh Rehmannia root, clean it, and dry the surface moisture at room temperature. Cut Rehmannia root into pieces, put the Rehmannia root pieces into a juicer to extract juice, and record the weight of the fresh Rehmannia root juice. Concentrate the fresh Rehmannia root juice to one tenth of the original volume, cool it at room temperature, and freeze-dry it in a vacuum freeze dryer for 72 hours. Weigh it, seal it and set it aside.

[0108] 3. Optimization of preparation process

[0109] 3.1 HPLC detection of active ingredients in Zanthoxylum bungeanum

[0110] HPLC was used to detect rutin, hyperoside, hesperidin, hydroxy-α-sanshool, hydroxy-β-sanshool, and hydroxy-γ-isosanshool in the extracts, and the total extract extraction rate of each group and the yield of effective components of Zanthoxylum bungeanum extract were calculated. The results are shown in Table 6.

[0111] Table 6 Extraction yield of each active ingredient in Zanthoxylum bungeanum extract

[0112]

[0113] 3.2 Orthogonal experimental results

[0114] The total extraction yield of the three amide components was used as an evaluation index, and SPSS 17.0 was used to analyze and process the experimental results to determine the optimal extraction process for Zanthoxylum bungeanum. From the intuitive analysis of the results in Table 7 and the variance analysis in Table 8, it can be seen that the influence on the content of amide components decreases in the order of percolation rate, solid-liquid ratio, and ethanol concentration. The percolation rate is an important factor in the percolation conditions. Therefore, the percolation conditions were set to A2B3C1, that is, 70% ethanol was added at a solid-liquid ratio of 1:20, and the percolation rate was determined to be 1mL·min -1 .

[0115] Table 7L9(3 4 )Orthogonal experiment results and analysis table

[0116]

[0117]

[0118] Table 8L9(3 4 ) Orthogonal test variance analysis

[0119]

[0120] 3.3 Determine the extraction method of Zanthoxylum bungeanum

[0121] According to the orthogonal design results, the extraction method of Zanthoxylum bungeanum was determined as follows: 450 g of Zanthoxylum bungeanum peel (red pepper) was placed in a percolation cylinder and soaked in 70% ethanol for 12 h. 20 times the volume of 70% ethanol was used at 1 mL min -1 The percolation was performed at a flow rate of 1.50 ℃. The percolation liquid was collected, concentrated to an organic solvent-free state by a rotary evaporator at 55°C, and then freeze-dried. The weight was 92.23 g, and the yield was 20.49%. The amount of the freeze-dried powder equivalent to 1 g of Zanthoxylum bungeanum extract was approximately 4.88 g of crude drug. The freeze-dried powder was collected and vacuum-sealed for later use.

[0122] 3.4 Preparation of fresh Rehmannia glutinosa juice concentrate

[0123] Take 1672g of fresh Rehmannia root, squeeze out 1022g of fresh Rehmannia root juice, boil and concentrate the fresh Rehmannia root juice to one tenth of the original volume, dry it in a vacuum freeze dryer, and obtain 262.13g of black solid powder. The yield is 15.68%, which is equivalent to 1g of Rehmannia root freeze-dried powder, which is about 6.38g of crude drug. Collect the freeze-dried powder, vacuum seal it, and set aside.

[0124] Example 2 Effect of the Chinese herbal compound composition on the learning and memory ability of dementia mice induced by intraventricular injection of Aβ25-35

[0125] 1. Experimental Materials

[0126] 1.1 Experimental animals

[0127] Male Kunming mice, SPF grade, 17-20g, 98 in total, were provided by Beijing Weitonglihua Experimental Animal Co., Ltd. All experimental procedures were in accordance with the requirements of the Animal Research Ethics Committee. The animals were allowed to eat and drink freely, and the experimental animal breeding environment and all experiments were carried out under the conditions of room temperature 25±2℃, humidity 55±10%, and 12h light and 12h dark circadian rhythm. The animals were acclimatized to the environment for 3 days before the experiment.

[0128] 1.2 Main experimental instruments and equipment

[0129] Table 9

[0130]

[0131] 1.3 Main experimental reagents and solutions

[0132] Table 10

[0133]

[0134] 1.4 Preparation of main experimental reagents and solutions

[0135] (1) Modeling agent Aβ 25-35 Solution: According to the instructions, 1 mg of Aβ 25-35 The powder was dissolved in 1 mL of sterile artificial cerebrospinal fluid to obtain a solution with a concentration of 1 mg / mL. After mixing evenly, it was placed in a 37°C cell culture incubator. After incubation for one week, the solution was divided and stored in a -20°C refrigerator for later use.

[0136] (2) Preparation of the Chinese medicine compound composition

[0137] The equivalent dose of the drug calculated according to the crude drug amount is set as the medium dose of the Chinese medicine compound composition, and the low dose of the Chinese medicine compound composition (9.575 mg·mL Zanthoxylum bungeanum extract) is prepared by dissolving in pure water. -1 , fresh Rehmannia glutinosa juice concentrate 33.304 mg·mL -1 ), medium dose (Zanthoxylum bungeanum extract 19.15 mg mL -1 , Fresh Rehmannia glutinosa juice concentrate 66.608 mg·mL -1 ), high dose (Zanthoxylum bungeanum extract 38.3 mg·mL -1 , fresh Rehmannia glutinosa juice concentrate 133.22 mg·mL -1 ).

[0138] (3) Donepezil Hydrochloride

[0139] Weigh 5 mg of donepezil hydrochloride tablets and grind them into powder using a mortar. Dissolve in pure water to a concentration of 0.3 mg / mL -1 , shake well before administration.

[0140] 2. Experimental methods

[0141] 2.1 Experimental Procedure

[0142] The experimental flow chart is as follows Figure 1 As shown, the animals were adapted to the environment for 3 days before the experiment. The animals were administered drugs three days later. Aβ was injected into the lateral ventricle stereotaxically on the 15th day. The water maze test was performed on the 21st day, the open field test was performed on the 27th day, and the samples were collected on the 29th day.

[0143] 2.2 Animal grouping and drug administration

[0144] 98 mice were randomly divided into 7 groups, 14 mice in each group, including normal group, model group, sham operation group, low-dose group of the Chinese medicine compound composition, medium-dose group of the Chinese medicine compound composition, high-dose group of the Chinese medicine compound composition and positive drug group. The drug groups were respectively 10 mL·Kg -1 The mice were intragastrically administered with a concentration of 100 mg / kg / cm2. The normal group, sham operation group, and model group were given an equal amount of pure water, and the positive drug group was given an equal amount of donepezil hydrochloride solution.

[0145] 2.3 Stereotaxic injection into the lateral ventricle

[0146] After the mice were anesthetized by respiratory anesthesia machine, the head was fixed by stereotaxic instrument. The top of the skull of the mouse was disinfected with iodine, and a 1 cm long incision was cut along the midline to separate the periosteum to expose the skull. Referring to the literature reports and the previous experimental results of the research group, the positioning coordinates were determined as follows: bregma was 0, AP-1.0mm, ML±1.0mm, DV3.0 mm. A 10μL microinjection needle was connected to a micro-continuous injection pump, and the injection was performed at a flow rate of 1μL / min. The injection time was 3min, and the needle was retained for 10min to ensure that the drug diffused in the brain.

[0147] 3. Behavioral testing

[0148] 3.1 Water maze experiment

[0149] The water maze experiment (Morris water maze, MWM) uses a mouse water maze real-time monitoring system to detect the activity of mice. The experiment was carried out according to the Morris classic scheme and appropriate improvements were made. The device consists of a circular pool filled with water, which is blackened by adding food coloring. The pool is evenly divided into four quadrants. A cylindrical platform is placed in one of the quadrants, so that it is 1.5 cm below the water surface. The software system locates the activity trajectory of the mouse in real time and collects data. The mice were tested in the water maze on the 7th day of modeling. Positioning navigation training was carried out for 5 days, 3 times a day, one swimming quadrant each time, and each training was adapted for 10 seconds before and after, and the detection time was 90 seconds. During the spatial exploration phase, the safety platform was removed, and the number of times the mouse crossed the platform within 90 seconds and the swimming distance of the target quadrant were monitored.

[0150] 3.2 Open field test

[0151] The open field test (OFT) was used to observe whether the Chinese herbal compound composition had an excitatory or inhibitory effect on the autonomous activity of mice. The instrument consists of 4 boxes with a camera on the top for monitoring. At the beginning of the experiment, the animals were randomly placed in the center of the bottom of the box, and the camera fixed on the top of the box recorded the image, and the behavioral changes and exploration trajectory of the animals within 5 minutes were observed. After the test of each animal was completed, it was returned to the original breeding cage and disinfected with 75% alcohol. The total distance of the animal, the number of explorations in the central area, the exploration time in the central area, and the exploration distance in the central area were recorded.

[0152] 3.3 Experimental Results

[0153] 3.3.1 Water maze experiment

[0154] (1) Positioning and navigation phase

[0155] In the positioning navigation test, the model group's landing rate was significantly reduced on the fourth and fifth days (P<0.01); the model group's escape latency was prolonged, and significantly prolonged on the fourth day (P<0.05); the percentage of swimming time in the target quadrant and the percentage of swimming distance decreased after the third day, but there was no significant difference. Compared with the model group, the landing rates of the positive drug group and each administration group of the Chinese medicine compound composition increased, and on the second day, the medium dose and high dose increased significantly (P<0.05), on the fourth day, the low dose increased significantly, and on the fifth day, the high dose and positive drug increased significantly; the percentage of swimming time in the target quadrant and the percentage of swimming distance increased, but there was no significant difference. The escape latency was reduced overall, and the high dose of the drug significantly decreased on the second day (P<0.05), the low dose of the drug significantly decreased on the fourth day (P<0.05), and the medium dose of the Chinese medicine compound composition and the positive drug significantly decreased on the fifth day (P<0.05). The results are shown in Figure 2 .

[0156] (2) Space Exploration Phase

[0157] On the sixth day of swimming, the platform was removed for a spatial exploration experiment. In the spatial exploration experiment, the number of platform crossings and the target quadrant swimming distance can reflect the spatial memory ability of mice. If the cognitive memory ability is good, the mice will explore around the platform for a long time, increase the target quadrant swimming distance, and increase the number of platform crossings.

[0158] The results showed that compared with the normal group, the model group had significantly lower crossing times (P<0.05) and significantly lower target quadrant travel distance (P<0.05); compared with the model group, the Chinese medicine compound combination-administered groups had higher crossing times and target quadrant travel distance, among which the medium-dose and positive drug groups had significantly higher crossing times (P<0.05), and the low-, medium-, and high-dose and positive drug groups had significantly higher target quadrant travel distance of mice (P<0.01, P<0.05). Figure 3 .

[0159] 3.3.2 Open field test

[0160] The open field test is often used to test the spontaneous activity and exploratory behavior of mice. The number of central area explorations, central area exploration time and central area exploration distance are used to evaluate the mice's autonomous exploration ability in a new environment.

[0161] The results showed that there was no significant difference in the total distance traveled by each group of mice in the open field test, indicating that the surgical operation and drug intervention had no obvious excitatory or inhibitory effect on the mental state and mobility of mice. Compared with the normal group, the number of central area explorations, central area exploration time and central area exploration distance of the model group mice were reduced, among which the number of central area explorations was significantly reduced (P<0.05). Compared with the model group, the number of spatial explorations in the high-dose group of the Chinese medicine compound composition was significantly increased (P<0.05), and the other drug-administered groups also had an increasing trend, but there was no significant difference. Results are shown in Figure 4 .

[0162] Example 3 Effects of Chinese herbal compound and single prescription on the learning and memory ability of scopolamine-induced dementia mice

[0163] 1. Experimental Materials

[0164] 1.1 Experimental animals

[0165] Male Kunming mice, SPF grade, 18-22g, 80 in total, were provided by Beijing Weitonglihua Experimental Animal Co., Ltd. All experimental procedures were in accordance with the requirements of the Animal Research Ethics Committee. The animals were allowed to eat and drink freely, and the experimental animal breeding environment and all experiments were carried out under the conditions of room temperature of 25±2℃ and humidity of 55±10%, and the circadian rhythm of 12h light and 12h dark was maintained. The animals were adapted to the environment for 3 days before the experiment.

[0166] 1.2 Main experimental instruments and equipment

[0167] Table 11

[0168]

[0169] 1.3 Main experimental reagents and solutions

[0170] Table 12

[0171]

[0172]

[0173] 1.4 Preparation of main experimental reagents and solutions

[0174] (1) Modeling agent scopolamine solution: According to the instructions, 4 mg of scopolamine hydrobromide was dissolved in 40 mL of normal saline to obtain a solution with a scopolamine content of 0.075 mg / mL. Prepare and use immediately.

[0175] (2) Preparation of the Chinese medicine compound composition

[0176] The Chinese medicine composition was prepared according to the previous research method. The pepper was sieved to obtain red pepper, then crushed and sieved, and 1:20 (v:v) 70% ethanol was used at 1 mL*min -1 The percolation extraction was performed at a flow rate, and the liquid was concentrated to an organic solvent-free state at 55°C using a rotary evaporator and then freeze-dried; fresh Rehmannia glutinosa juice was squeezed and stored at -20°C for later use. The solvent for oral administration in this experiment was 1% castor oil + 0.5% CMC-Na aqueous solution. The equivalent dose of the drug was calculated based on the amount of the raw drug and set as the high dose of the Chinese medicine composition (194.467 mg·kg of the red pepper extract) -1 , Fresh Rehmannia glutinosa juice concentrate 676.767 mg kg -1 ), the low dose of the compound was 1 / 2 of the high dose. The high dose group of pepper red was given 194.467 mg·kg -1 The high-dose group of Rehmannia glutinosa was 676.767 mg kg fresh Rehmannia glutinosa juice concentrate -1 .

[0177] (3) Donepezil Hydrochloride

[0178] Weigh 10 mg of donepezil hydrochloride tablets, grind them into powder in a mortar, dissolve them in the above solvent and press 3 mg kg -1 The dose was administered by oral gavage.

[0179] 2. Experimental methods

[0180] 2.1 Experimental Procedure

[0181] The animals were acclimatized for 3 days before the experiment. The animals were dosed 3 days later. The open field test was performed 30 minutes after the intragastric administration on the 17th day. Scopolamine was injected intraperitoneally before the start of learning and memory-related tests, and behavioral experiments were performed 20 minutes later. New object recognition experiments were performed on days 18 to 19, water maze positioning and navigation experiments were performed on days 20 to 24, water maze space exploration experiments were performed on day 25, dark avoidance experiments were performed on days 26 to 27, and samples were collected on day 28. See the experimental procedures for details. Figure 5.

[0182] 2.2 Animal grouping and drug administration

[0183] 80 mice were randomly divided into 8 groups according to their body weight: normal group, model group, solvent + model group, high-dose group of red pepper, high-dose group of Rehmannia glutinosa, high-dose group of compound (Chinese medicine composition), low-dose group of compound (Chinese medicine composition), and positive drug (donepezil) group. Each group of animals was intragastrically administered with a volume of 0.2ml / 10g, and the normal group and model group were intragastrically administered with pure water.

[0184] 2.3 Tissue sampling and biochemical testing

[0185] The experimental animals were sampled the next day after all behavioral tests were completed. Blood was collected from the inner canthus of the eye to separate serum, and the mouse brain was peeled off and the cortex was separated. The tissue was quenched with liquid nitrogen and frozen at -80°C. The unilateral cortex of the mouse was weighed and mixed with 0.9% saline and then ground. The supernatant was extracted by centrifugation for 5 minutes according to the requirements of the kit. The protein concentration was determined by the colorimetric BCA kit, and the acetylcholine content and acetylcholinesterase and acetylcholine transferase activities were determined and calculated by the biochemical kit. The malondialdehyde (MDA) content in mouse serum was detected according to the instructions of the biochemical kit.

[0186] 2.3 Behavioral testing

[0187] 2.3.1 Open field test

[0188] The open field experiment was used to observe whether the Chinese herbal compound and its ingredients had an excitatory or inhibitory effect on the autonomous activities of mice. The instrument consists of 3 boxes (each 50cm*50cm*50cm) with a camera on the top for monitoring. At the beginning of the experiment, the animals were randomly placed in the center of the bottom of the box, and the camera fixed on the top of the box recorded the image, and the behavioral changes and exploration trajectory of the animals within 5 minutes were observed. After the test of each animal was completed, it was returned to the original breeding cage and disinfected with 75% alcohol. The total distance of the animal's movement was recorded.

[0189] 2.3.2 Water maze experiment

[0190] The effect of the Chinese medicine compound composition and its ingredients on the spatial long-term learning and memory of mice was observed by water maze experiment. The mouse water maze real-time monitoring system was used to detect the activity of mice. The experiment was carried out according to the Morris classic scheme and appropriate improvements were made. The device consists of a circular pool filled with water, which is blackened by adding food coloring. The pool is evenly divided into four quadrants. A cylindrical platform is placed in one of the quadrants so that it is 1.5 cm below the water surface. The software system locates the activity trajectory of the mouse in real time and collects data. The mice were given medication 1 hour before modeling, modeled 40 minutes, and water maze tested 20 minutes later. Positioning navigation training was carried out for 5 days, 3 times a day, each time in one swimming quadrant, and each training was adapted for 10 seconds before and after, and the detection time was 90 seconds. The mice were recorded for each training (staying for 5 seconds after climbing the hidden platform). After 1 day of spatial exploration experiment, the safety platform was removed, and the number of mice crossing the platform within 90 seconds and the swimming distance and swimming time of the target quadrant were monitored.

[0191] 2.3.3 New object recognition experiment

[0192] The effect of the Chinese medicine compound composition and its ingredients on the non-spatial short-term memory of mice was observed by the novel object recognition test (NORT). The adaptation period is 24 hours before the familiarization period, and the mice are placed in an empty box for 5 minutes. During the familiarization period, the mice are placed in the test box with their backs to two identical objects (A, A') for 5 minutes. During the test period: one of the objects is replaced with an object B with a different shape and color, and the mouse is placed in the test box with its back to the two objects for 5 minutes. Calculate the discrimination index: (exploration time of the animal on the novel object during the test period - exploration time on the familiar object) / (exploration time of the animal on the novel object + exploration time on the familiar object during the test period).

[0193] 2.3.4 Dark avoidance experiment

[0194] The effect of the Chinese medicine compound composition and its ingredients on the non-spatial passive avoidance learning and memory of mice was observed by the passive avoidance test (PAT). The adaptation period was 24 hours before the familiarization period, and the mice were placed in a non-powered test box for 10 minutes. After the dark room was powered on during the familiarization period, the mice were placed in the test box for 5 minutes. During the test period, the mice were placed in a bright room and powered on. The mice entered the dark room from the bright room, which was a dark avoidance error. The dark avoidance error of the mice (the number of dark avoidance errors, the incubation period of dark avoidance errors, and the number of probes) were recorded.

[0195] 3 Experimental results

[0196] 3.1 Behavioral Experiment

[0197] 3.1.1 Open field test

[0198] The open field test is often used to test the spontaneous activity and exploratory behavior of mice. The results showed that there was no significant difference in the total distance traveled by each group of mice in the open field test, indicating that SCOP modeling and drug intervention had no obvious excitatory or inhibitory effects on the mental state and behavioral ability of mice. Figure 6 .

[0199] 3.1.2 Water maze experiment

[0200] (1) Positioning and navigation phase

[0201] The results of the water maze directional navigation experiment showed that the SCOP modeling of mice resulted in a significantly lower rate of appearance on the stage in the model group and the solvent + model group during the training period than in the normal group (P<0.01, P<0.05). The appearance rate of the drug group was significantly higher than that of the solvent + model group starting from Day 3. On Day 5, the appearance rate of the positive drug, red pepper, and Rehmannia high-dose group was significantly higher than that of the solvent + model group (P<0.05). The appearance rate of the low- and high-dose groups of the Chinese medicine combination was significantly higher than that of the model group (P<0.05) and the solvent + model group (P<0.01). Figure 7 A.

[0202] (2) Space Exploration Phase

[0203] On the 6th day of the water maze experiment, the platform was removed for the spatial exploration experiment. In the spatial exploration experiment, the number of platform crossings, the ratio of swimming distance in the target quadrant, and the ratio of time in the target quadrant can reflect the spatial memory ability of mice. If the cognitive memory ability is good, the mouse will spend more time exploring the quadrant where the hidden platform is located, increase the swimming distance in the target quadrant, and increase the number of platform crossings.

[0204] The results of the water maze space exploration experiment showed that the number of times the mice in the model group and the model + solvent group crossed the hidden platform (P<0.01, P<0.05), the target quadrant travel distance ratio (P<0.01), and the target quadrant time ratio (P<0.01) were much lower than those in the normal group, and there was no significant difference between the model group and the model + solvent group. After drug administration, all data were adjusted back, among which the number of crossing the platform of the positive drug donepezil (P<0.01), high-dose compound (P<0.05), and low-dose compound (P<0.05) was significantly higher than that of the model group; the target quadrant travel distance ratio and time ratio of the low-dose compound (P<0.01) and high-dose compound (P<0.05) groups were significantly higher than those of the model group, and the low-dose compound was close to the normal group. The results of the water maze experiment are shown in Figure 7 BD.

[0205] 3.1.3 New object recognition experiment

[0206] The novel object recognition experiment uses the animals' instinctive curiosity and exploration behavior towards novel things to evaluate the non-spatial short-term memory ability of the tested animals by the length of time they explore familiar objects that they have touched and new objects that they have not touched but are different in shape and color. The experimental results are presented by the discrimination index. A long exploration time for novel objects (a large discrimination index) proves that the tested animals have formed memories of familiar objects and have the ability to explore and be curious about novel things. The experimental results showed that the discrimination index of the model group and the solvent + model group decreased significantly (P<0.01). After drug administration, the recognition index of each group was able to recover, among which the high-dose of red pepper, high-dose of Rehmannia glutinosa and high-dose of the compound had a significant recovery effect (P<0.05). The results of the novel object recognition experiment are shown in Figure 8 .

[0207] 3.1.4 Dark avoidance experiment

[0208] The dark avoidance experiment, also known as the passive avoidance experiment, uses the dark tendency of rodents to train animals with electrical stimulation in a dark room, so that the animals acquire the memory of restraining from entering the dark room. If the animals enter the dark room, it proves that they have not acquired a solid dark avoidance memory. The results showed that the number of times the model group and the solvent + model group entered the dark room, that is, the number of errors, increased significantly (P<0.01), the latency of errors in entering the dark room decreased significantly (P<0.01), and the number of probes into the dark room increased significantly (P<0.01).

[0209] The number of dark avoidance errors in the drug-treated group was significantly reduced (P<0.01), and the high-dose compound group could be adjusted back to the normal group. The high-dose of pepper red and the high-dose compound could significantly prolong the dark avoidance latency (P<0.01) and reduce the number of probes (P<0.01). The degree to which the positive drug donepezil reduced the number of probes (P<0.01) was slightly lower than that of the above two groups. The low-dose compound significantly reduced the number of probes, but the effect was slightly lower than that of the positive drug, pepper red and high-dose compound groups (P<0.05). The results of the dark avoidance experiment are shown in Fig. 9 .

[0210] 3.2 Biochemical tests related to the cholinergic system

[0211] After SCOP modeling, the model group and the solvent + model group showed the same trend and there was no significant difference between the groups, proving that the solvent had no significant effect on the efficacy of the modeling drug. SCOP modeling led to a significant decrease in the activity of acetylcholine synthase (ChAT) in the cerebral cortex of mice, a significant increase in the activity of acetylcholine degrading enzyme (AChE), and a decrease in the level of acetylcholine (ACh). Fig.10 .

[0212] 3.3 Oxidative stress-related biochemical tests

[0213] The level of malondialdehyde in the serum of acute AD mice induced by SCOP increased, and after drug administration, all groups showed a downward trend, among which the level of malondialdehyde decreased more significantly after the positive drug donepezil and the low and high doses of the compound (P<0.01). Fig.11 .

Claims

1. A Chinese medicine composition for preventing and / or treating Alzheimer's disease, characterized in that: The Chinese medicine composition consists of Zanthoxylum bungeanum extract and Rehmannia glutinosa extract, the weight ratio of the Zanthoxylum bungeanum extract to the Rehmannia glutinosa extract is 28-29:60-100; the Rehmannia glutinosa extract is a concentrate of fresh Rehmannia glutinosa juice; and the extraction solvent of the Zanthoxylum bungeanum extract is 70% ethanol solution.

2. The Chinese medicine composition according to claim 1, characterized in that The Chinese medicine composition also includes pharmaceutically acceptable excipients, which are selected from: preservatives, colorants, pH regulators, buffers, antioxidants, metal ion chelators, isotonic regulators, diluents, adhesives, lubricants, sweeteners, absorbents, disintegrants, wetting agents, and coating materials. The dosage form of the Chinese medicine composition is selected from: decoctions, powders, oral liquids, tablets, capsules, pills, injections, inhalants, suppositories, emulsions, nanoparticles, gels, suspoemulsions, creams, and sprays.

3. Use of the Chinese medicine composition according to claim 1 in the preparation of a medicament for preventing and / or treating a disease with amnesia as a main symptom, wherein the disease is Alzheimer's disease.

4. The use according to claim 3, characterized in that The Alzheimer's disease is selected from: early-onset Alzheimer's disease, late-onset Alzheimer's disease and familial Alzheimer's disease.

5. The use according to claim 3, characterized in that The prevention and / or treatment of diseases with amnesia as the main symptom includes the prevention and / or treatment of: memory loss and memory disorders.

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