Traditional Chinese medicine composition for resisting Alzheimer disease as well as preparation method and application of traditional Chinese medicine composition

Abstract

The invention provides a traditional Chinese medicine composition for resisting Alzheimer's disease as well as a preparation method and application of the traditional Chinese medicine composition. The traditional Chinese medicine composition is prepared from the following active ingredients in parts by weight: 6-50 parts of astragalus membranaceus, 6-50 parts of salvia miltiorrhiza, 6-50 parts of uncaria, 3-30 parts of ginkgo leaves, 3-30 parts of gardenia, 3-30 parts of cistanche, 3-30 parts of rhodiola rosea, 3-30 parts of corydalis tuber, 3-30 parts of ligusticum wallichii, 3-30 parts of rhizoma anemarrhenae, 3-30 parts of medlar, 3-30 parts of semen raphani, 3-30 parts of teasel root, 3-30 parts of golden cypress, 3-30 parts of acorus tatarinowii, 3-30 parts of pseudo-ginseng and 2-30 parts of coptis chinensis. The raw materials of the medicine are common, the preparation method is simple and easy to operate, the medicine has wide application value, after oral administration, cognitive impairment of a rat model with the Alzheimer disease can be remarkably improved, the content of inflammatory factors IL-6 and IL-1beta in brain tissue is reduced, meanwhile, the number of activated microglial cells and astrocytes in the brain tissue is reduced, and the curative effect of the medicine is improved. The compound can inhibit excessive phosphorylation of tau protein in brain tissues, and has a new application of an anti-Alzheimer disease drug.

Description

Technical Field

[0001] This invention belongs to the field of biomedical technology, specifically relating to a traditional Chinese medicine composition for treating Alzheimer's disease, its preparation method, and its application in the preparation of drugs for treating or preventing Alzheimer's disease. Background Technology

[0002] Alzheimer's disease (AD) is a neurodegenerative disease characterized by progressive cognitive impairment and memory loss. It seriously harms the physical and mental health of the elderly and imposes a heavy economic burden on families and society. According to the AD Association, there are currently about 50 million people with AD worldwide, and the annual medical costs for treating AD have reached $1 trillion.

[0003] In Traditional Chinese Medicine (TCM), Alzheimer's disease (AD) falls under the categories of "dullness syndrome," "forgetfulness," "literary idiocy," "healing syndrome," and "depression syndrome." Zhang Jingyue of the Ming Dynasty first proposed dementia as an independent disease in his book *Jingyue Quanshu*, elaborating on its causes as primarily "liver stagnation, phlegm obstruction, and stomach deficiency." Physicians throughout history have believed this disease is closely related to kidney deficiency and marrow depletion, liver qi stagnation, and phlegm and blood stasis obstructing the orifices. Therefore, tonifying the kidneys and replenishing marrow, soothing the liver and relieving stagnation, calming the mind and opening the orifices, strengthening the spleen and replenishing qi, and resolving phlegm and removing blood stasis are the basic methods for treating this disease.

[0004] Currently, there is no specific treatment for Alzheimer's disease (AD) in modern medicine. Clinically available treatments for AD include cholinesterase inhibitors (donepezil, galantamine, rivastigmine), N-methyl-D-aspartate (NMDA) receptor antagonists (memantine), and Aβ monoclonal antibodies (aducanumab). However, these drugs can only slow the progression of dementia symptoms and alleviate some symptoms within a limited timeframe; they cannot delay or stop the progression of AD. Furthermore, they have limitations such as easy drug resistance and significant side effects (e.g., headaches, diarrhea, insomnia). Traditional Chinese medicine and natural products, due to their multi-target effects and fewer side effects, are increasingly favored by clinical researchers.

[0005] Therefore, developing safer and more effective traditional Chinese medicine formulas for treating Alzheimer's disease (AD) is an urgent problem to be solved. Summary of the Invention

[0006] To overcome the shortcomings of existing technologies, this invention provides a traditional Chinese medicine composition for treating Alzheimer's disease, its preparation method, and its application.

[0007] In a first aspect, the present invention provides a traditional Chinese medicine composition for treating Alzheimer's disease, wherein the effective components of the traditional Chinese medicine composition are made from raw materials comprising the following parts by weight: Astragalus membranaceus 6-50, Salvia miltiorrhiza 6-50, Uncaria rhynchophylla 6-50, Ginkgo biloba leaf 3-30, Gardenia jasminoides 3-30, Cistanche deserticola 3-30, Rhodiola rosea 3-30, Corydalis yanhusuo 3-30, Ligusticum chuanxiong 3-30, Anemarrhena asphodeloides 3-30, Lycium barbarum 3-30, Raphanus sativus seed 3-30, Dipsacus asper 3-30, Phellodendron chinense 3-30, Acorus tatarinowii 3-30, Panax notoginseng 3-30, and Coptis chinensis 2-30. In a second aspect, the present invention provides a method for preparing the traditional Chinese medicine composition described in the first aspect, comprising the following steps:

[0008] (1) Weigh each raw material according to the ratio, soak it in distilled water at room temperature for 12-24 hours in 10-20 times the weight of all the medicinal materials, heat and extract for 0.5-2 hours, filter, and repeat the above extraction steps twice;

[0009] (2) Combine the three filtrates, remove all water under reduced pressure, and freeze dry to obtain a fine powder of the active ingredients of the traditional Chinese medicine composition.

[0010] Thirdly, the present invention provides the application of the traditional Chinese medicine composition described in the first aspect in the preparation of a drug for treating Alzheimer's disease.

[0011] The beneficial effects of this invention include: the traditional Chinese medicine composition of this invention has been experimentally proven to significantly improve symptoms and slow disease progression. The raw materials used in this invention are common, and the preparation method is simple and easy to operate, making it widely applicable. Oral administration of the drug can significantly improve cognitive impairment in a rat model of Alzheimer's disease, reduce the levels of inflammatory factors IL-6 and IL-1β in brain tissue, while simultaneously reducing the number of activated microglia and astrocytes in brain tissue and inhibiting excessive phosphorylation of tau protein in brain tissue, thus possessing novel medicinal uses in the treatment of Alzheimer's disease. Attached Figure Description

[0012] Figure 1A , 1B 1A and 1B represent the effects of CFI on the movement path (1A), the time the rat spent in the center of the open field (1B), and the movement speed (1C) of the rat in the open field experiment, respectively. * p<0.05, ** p<0.01.

[0013] Figure 2A , 2B 2A and 2C represent the effects of CFI on STZ-induced recognition memory in rats on the following aspects: (2A) the movement path of the new object recognition test in rats, (2B) the time of exploring new objects in rats, and (2C) the NORT index in rats (n=10). * p<0.05, **p<0.01.

[0014] Figure 3A , 3B 3A and 3C represent the effects of CFI on STZ-induced spatial memory in rats: (3A) swimming path during the test period, (3B) escape latency during the training period, and (3C) time spent in the target quadrant during the test period (n=10). * p<0.05, ** p<0.01, *** p<0.001, **** p<0.0001.

[0015] Figure 4A and 4B The effects of CFI on the levels of IL-6 (4A) and IL-1β (4B) in STZ-induced rat brain tissue were respectively (n=6). * p<0.05, ** p<0.01, *** p<0.001, **** p<0.0001.

[0016] Figure 5 This diagram shows the effects of CFI on STZ-induced astrocytes (Figures A and B) and microglia (Figures C and D) in the cerebral cortex of rats (n=3). GFAP: glial fibrillary acidic protein, astrocyte-specific marker; DAPI: 4',6-diamidin-2-phenylindole, nuclear-specific marker; Iba-1: ion-calcium binding linker molecule 1, microglia-specific marker; Merge: integration diagram. * p<0.05, ** p<0.01, *** p<0.001, **** p<0.0001.

[0017] Figure 6 These are protein blot bands representing the expression of tau protein phosphorylation sites Thr205, Ser396, and Ser404 in rat brain tissue induced by STZ.

[0018] Figure 7A , 7B 7C represents the effect of CFI on the expression levels of tau protein phosphorylation sites Thr205, Ser396, and Ser404 in the brain tissue of STZ-induced rats (n=6). * p<0.05, ** p<0.01, *** p<0.001. Detailed Implementation

[0019] In this article, "room temperature" and "normal temperature" both refer to a temperature between 23-25℃.

[0020] This invention provides a traditional Chinese medicine composition for treating Alzheimer's disease. The effective components of the composition are made from the following raw materials in parts by weight: Astragalus membranaceus 6-50, Salvia miltiorrhiza 6-50, Uncaria rhynchophylla 6-50, Ginkgo biloba 3-30, Gardenia jasminoides 3-30, Cistanche deserticola 3-30, Rhodiola rosea 3-30, Corydalis yanhusuo 3-30, Ligusticum chuanxiong 3-30, Anemarrhena asphodeloides 3-30, Lycium barbarum 3-30, Raphanus sativus 3-30, Dipsacus asper 3-30, Phellodendron chinense 3-30, Acorus tatarinowii 3-30, Panax notoginseng 3-30, and Coptis chinensis 2-30.

[0021] Among the above-mentioned active ingredients, all traditional Chinese medicines possess anti-Alzheimer's disease and neuroprotective effects: Bioactive compounds such as astragaloside A, quercetin, isoflavones, tanshinone IIA, and salvianolic acid A, isolated from Astragalus membranaceus and Salvia miltiorrhiza, can significantly treat AD by inhibiting the formation and deposition of β-amyloid protein (Aβ), tau protein hyperphosphorylation, neuroinflammation, and synaptic defects; Uncaria rhynchophylla extract and its active ingredients, isorhein and rhynchophylline, can exert neuroprotective effects on different AD models through multiple pathways. Ginkgo biloba extract has been shown to treat age-related cognitive impairment. Bioactive compounds isolated from gardenia, cistanche, and coptis, respectively, as well as from rhodiola rosea extract, can improve learning and memory function in AD models by inhibiting apoptosis, oxidative stress, and inflammation. Major compounds of corydalis, such as corydaline, berberine, and berberine, as well as the water extract of dipsacus, have been found to have acetylcholinesterase inhibitory effects. Z-ligustilide, a bioactive marker of chuanxiong, can regulate tumor necrosis factor. TNF-α activates the nuclear factor κB (NF-κB) signaling pathway to protect against Aβ25-35-induced neurotoxicity; Anemarrhena asphodeloides extract and its active compounds, such as anemarrhena saponins B-II and A-III, have shown effects in improving memory impairment in AD animal models by inhibiting β-secretase 1 (BACE1); Lycium barbarum extract can improve memory deficits in AD mice by reducing Aβ deposition, increasing NeuN-positive cells, and upregulating the expression of neurotrophic factors; sulforaphane, an active compound of radish seed, can alleviate cognitive dysfunction by inhibiting Aβ deposition, tau protein phosphorylation, and neuroinflammation, while improving synaptic plasticity in different animal models; Phellodendron chinense cortex has a neuroprotective effect against Aβ-induced neurotoxicity in PC12 cells through an anti-apoptotic mechanism; the active components of Acorus tatarinowii can alleviate AD pathological syndromes by improving myelin damage in mice and inhibiting tau protein phosphorylation; Panax notoginseng saponins have also been shown to inhibit Aβ formation and deposition, prevent excessive tau protein phosphorylation, and maintain acetylcholine homeostasis in AD models. This invention, through the synergistic effect of the above-mentioned active ingredients, has the advantages of significantly improving symptoms and slowing down the disease progression. After oral administration, it can significantly improve cognitive impairment in a rat model of Alzheimer's disease, reduce the content of inflammatory factors IL-6 and IL-1β in brain tissue, reduce the number of activated microglia and astrocytes in brain tissue, and inhibit the excessive phosphorylation of tau protein in brain tissue, thus having a new drug use in the fight against Alzheimer's disease.

[0022] In a preferred embodiment, the weight proportions of each raw material in the active ingredient are as follows: Astragalus membranaceus 15%, Salvia miltiorrhiza 15%, Uncaria rhynchophylla 14%, Ginkgo biloba 10%, Gardenia jasminoides 10%, Cistanche deserticola 10%, Rhodiola rosea 10%, Corydalis yanhusuo 10%, Ligusticum chuanxiong 10%, Anemarrhena asphodeloides 10%, Lycium barbarum 10%, Raphanus sativus 10%, Dipsacus asper 10%, Phellodendron chinense 10%, Acorus tatarinowii 9%, Panax notoginseng 6%, and Coptis chinensis 4%. This invention establishes an Alzheimer's disease rat model to observe the therapeutic effect of the active ingredient (CFI) in the preferred embodiment on streptozotocin (STZ)-induced cognitive impairment in a rat model of Alzheimer's disease, and analyzes its possible mechanism of action. Evaluation of various behavioral indicators in rats revealed that CFI significantly improves cognitive impairment in rats, and its mechanism of action may be related to inhibiting inflammatory responses and inhibiting excessive phosphorylation of tau protein. Currently, there are no literature reports on the efficacy of CFI in alleviating cognitive impairment or its application in the treatment of Alzheimer's disease.

[0023] This invention demonstrates through in vivo experiments that CFI can improve cognitive impairment in rats. Biochemical reagent kits were used to measure the levels of interleukin-6 (IL-6) and interleukin-1β (IL-1β) in brain tissue; histopathological methods were used to observe the activation of microglia and astrocytes in the cerebral cortex; and Western blotting was used to detect the phosphorylation level of the key protein tau in brain tissue. These findings further demonstrate that CFI has a good therapeutic effect on STZ-induced cognitive impairment and shows promising potential for development into an anti-Alzheimer's disease drug or functional food. Details are as follows: 1. The therapeutic effect of the traditional Chinese medicine composition of this invention on Alzheimer's disease.

[0024] 1.1 Experimental Materials

[0025] 1.1.1 Laboratory animals and drugs

[0026] SPF-grade male SD rats, 220-240g, were purchased from the Laboratory Animal Centre of the Chinese University of Hong Kong. The animals were housed separately, with bedding changed twice weekly. An acclimatization period of 7 days was provided, during which time they had free access to water and food. The ambient temperature was 22±2℃, relative humidity was 60%, and the environment featured a 12-hour light-dark cycle.

[0027] 1.1.2 Preparation of test reagents

[0028] Source: Extracted from the laboratory.

[0029] Preparation of the active ingredient (CFI) in the traditional Chinese medicine composition: All medicinal materials (one dose containing 15g Astragalus membranaceus, 15g Salvia miltiorrhiza, 14g Uncaria rhynchophylla, 10g Ginkgo biloba, 10g Gardenia jasminoides, 10g Cistanche deserticola, 10g Rhodiola rosea, 10g Corydalis yanhusuo, 10g Ligusticum chuanxiong, 10g Anemarrhena asphodeloides, 10g Lycium barbarum, 10g Raphanus sativus, 10g Dipsacus asper, 10g Phellodendron chinense, 9g Acorus tatarinowii, 6g Panax notoginseng, and 4g Coptis chinensis) were soaked in 10 times the amount of water for 24 hours, heated under reflux for 1 hour, filtered, and the above extraction steps were repeated twice. The filtrates from the three extractions were combined, recovered under reduced pressure, and freeze-dried to obtain the extract (i.e., the active ingredient, CFI), with an extraction rate of 37% (w / w). The above extract was prepared into test solution samples with concentrations of 0.3g, 0.6g, and 0.9g raw material / mL using distilled water.

[0030] Positive control group test solution (PC): Accurately weigh 20 mg donepezil hydrochloride and dissolve it in distilled water to prepare a test solution sample with a concentration of 0.5 mg / mL. After preparation, store the above-mentioned drugs at 4°C for later use.

[0031] 1.1.3 Experimental Reagents

[0032] Modeling and drug administration reagents: streptozotocin (STZ), Santa Cruz Biosciences, USA; donepezil hydrochloride, Sigma-Aldrich, USA.

[0033] Rat IL-6 and IL-1β kit, Abclonal (China).

[0034] Reagents used for frozen section preparation and staining: sucrose, Sigma-Aldrich, USA; OCT cryoembedding medium, Sakura Corporation, USA.

[0035] Reagents used in Western blot assays: RIPA lysis buffer, protease inhibitor cocktail, PMSF (100mM), phosphorylated protease inhibitor, protein loading buffer, SDS-PAGE gel preparation reagent, TRIS buffer, glycine, SDS, Tween-20 (Sigma-Aldrich, USA); PVDF membrane (0.22μm) (Bio-Rad); BSA (Thermo Scientific); GAPDH (Cell Signal Technology); tau protein, tau protein phosphorylated Thr205, Ser396, and Ser404 antibodies (Abcam); protein markers, HRP-labeled goat anti-rabbit antibody, and HRP-labeled goat anti-mouse antibody (Thermo Scientific).

[0036] 1.1.4 Experimental Apparatus

[0037] SHIMADZU analytical balance, Guangzhou Xiangyi Electromechanical Equipment Co., Ltd.; PL-203 electronic balance, Mettler Toledo Instruments (Shanghai) Co., Ltd.; IKA10 tissue homogenizer, IKA GmbH, Germany; FLUO star Optima microplate reader, BMG Labtec GmbH, Germany; Azure Biosystems c300 developing system, Azure Biosystems, USA; Zeiss Axio Scope A1 microscope, Zeiss GmbH, Germany; benchtop high-speed refrigerated centrifuge, Heal Force; SuperMaze V2.0 behavioral analysis system, Shanghai Xinruan, China.

[0038] 1.2 Animal grouping and administration

[0039] Rats were anesthetized with xylazine (10 mg / kg) and ketamine (75 mg / kg) and fixed on a rat brain localization device. The location of the lateral ventricles was determined by the fontanelle position according to coordinates (0.8 mm posterior to the anterior fontanelle, ±1.4 mm lateral to the sagittal plane, and 4.0 mm ventral to the skull surface). STZ (2 mg / kg, 5 μL) was injected into both lateral ventricles at a rate of 1 μL / min using a micro-injector. After injection, the needle was left in place at the injection site for 5 minutes before being withdrawn, and the incision was sutured. Rats in the sham-operated group received 5 μL of 0.9% saline in the lateral ventricles. After surgery, the rats were returned to their cages to recover. The day after surgery, the rats were randomly divided into: a model group (STZ group), a low-dose CFI treatment group (3 g / kg), a medium-dose CFI treatment group (6 g / kg), a high-dose CFI treatment group (9 g / kg), a positive control group (PC, 5 mg / kg), and a sham-operated group (n=10 per group). The sham-operated group and the model group were given the same volume of distilled water. The remaining groups of rats were administered CFI and PC at the above concentrations by gavage once daily for 6 weeks.

[0040] 1.3 Rat Open Field Test (OFT)

[0041] One hour prior to the experiment, rats were transferred to the behavioral laboratory to acclimatize. The rats were placed in a square open field facility (80cm × 80cm × 80cm), and each rat was allowed 10 minutes to explore. Video recording and analysis were performed using SuperMaze V2.0 (Shanghai Xinruan, China). Between each experiment, the open field facility was wiped clean with 70% ethanol.

[0042] 1.4. Rat Novel Object Recognition Test

[0043] The Novel Object Recognition Test (NORT) is a rapid and effective method for testing rats' learning and memory at different stages. This experiment consists of three phases: an acclimatization period, a training period, and a testing period. During the acclimatization period, the rat is placed in the center of an open area (test box) and allowed to explore freely for 10 minutes. The training period involves visual exploration of two identical objects. After 24 hours of acclimatization, the rat is removed from its cage and placed in the center of the test box, equidistant from the two identical objects, allowing it to explore freely for 5 minutes. During the testing period, one of the previously explored objects is replaced with a new object. Because rodents have a natural preference for novelty, they will spend more time exploring new objects if they remember familiar ones. During the testing period, one of the objects used in the training period (the familiar object) and the new object are placed in the test box, allowing the rat to explore for 5 minutes. Video recording and analysis are performed using SuperMaze V2.0 (Shanghai Xinruan, China). The test box is wiped clean with 70% ethanol between each test.

[0044] 1.5. Rat Water Maze Test

[0045] The Water Maze Test (MWMT) is an experiment to evaluate spatial learning and memory functions. A circular pool (120 cm in diameter and 30 cm in height) is placed in a behavioral laboratory, and the water temperature is maintained at 25 degrees Celsius. The water maze is divided into four equal quadrants, and four different shapes are placed on the pool walls to mark the different quadrants. A hidden platform (8 cm in diameter) is placed 0.5 cm below the water surface at the midpoint of one of the quadrants. The MWMT lasts for five days. The first four days are the spatial training period, training rats to find the hidden platform in the water maze. On the first day of spatial training, each rat swims three times from different directions, each time for a maximum of 60 seconds, and then each rat is guided to the platform and stays on the platform for 30 seconds. On days 2 to 4, each rat is trained to swim for a maximum of 60 seconds each time. Escape latency is recorded during the training period. On day 5, the hidden platform is removed, and the rats are allowed to swim freely in the water for 60 seconds without the hidden platform. The swimming parameters are recorded and analyzed using the video tracking software SuperMaze V2.0 (Shanghai Xinruan, China).

[0046] 1.6. Serum and brain tissue sampling

[0047] After the experiment, blood was collected from rats via cardiac puncture. The blood samples were allowed to stand at room temperature for 2 hours, then centrifuged at 3000 rpm for 15 minutes, and the supernatant was separated to obtain serum samples. After blood collection, the rats were euthanized by cervical dislocation, and brain tissue was immediately removed. The cortex and hippocampus were extracted and frozen at -80°C for subsequent experiments. Four rats were randomly selected from each group for cardiac perfusion, and the brains were then fixed in 4% (v / v) paraformaldehyde for 24 hours for preparing pathological tissue sections.

[0048] 1.7 Western Blot detection of related protein expression

[0049] 1.7.1 Total Protein Extraction

[0050] Wash tissue blocks 2-3 times with cold PBS to remove blood, cut into small pieces and place in a homogenizer. Add 10 times the tissue volume of lysis buffer (add protease inhibitor a few minutes before use) and homogenize thoroughly on ice. Transfer the homogenate to a 1.5 mL centrifuge tube and vortex. Incubate on ice for 30 min, repeatedly pipetting during this time to ensure complete cell lysis. Then centrifuge at 12000g for 10 min and collect the supernatant, which is the total protein solution.

[0051] 1.7.2 Protein concentration determination: Protein concentration was determined by the Coomassie brilliant blue method.

[0052] 1.7.3 SDS-PAGE electrophoresis

[0053] Prepare the SDS-PAGE gel according to the instructions of the SDS-PAGE gel preparation kit. After adding sufficient electrophoresis buffer, load the sample into the electrophoresis wells for electrophoresis (stacking gel voltage 70V, 1h; separating gel voltage 120V, 30min). Stop electrophoresis when bromophenol blue just runs out, and then proceed with the membrane transfer.

[0054] 1.7.4 Transfer of film

[0055] Prepare six 7×9cm filter papers and one appropriately sized PVDF membrane. Activate the PVDF membrane with methanol for 15 seconds before use. Place the transfer clamp, two sponge pads, filter paper, and the activated PVDF membrane in a bowl containing transfer buffer. Open the clamp so the black side is horizontal. Place the sponge pads and three layers of filter paper on the mats. Carefully peel off the separating gel and place it on the filter paper. Place the membrane on the gel, removing any air bubbles. Then cover the membrane with three more layers of filter paper, removing any air bubbles. Finally, cover with the remaining sponge pad. Transfer conditions (wet transfer): 250mA constant current for 45 minutes.

[0056] 1.7.5 Immune Response

[0057] The transferred membrane was blocked for 2 hours on a decolorizing shaker at room temperature with 5% (w / v) skim milk (prepared with 0.5% (w / v) TBST). The primary antibody was diluted (5% (w / v) skim milk dissolved in TBST; phosphorylated proteins were treated with 5% (w / v) BSA dissolved in TBST), and incubated overnight at 4°C. The membrane was then washed three times with TBST on a decolorizing shaker at room temperature for 5 minutes each time. The secondary antibody was diluted 2000-fold with TBST and incubated for 2 hours at room temperature. The membrane was then washed three times with TBST on a decolorizing shaker at room temperature for 5 minutes each time.

[0058] 1.7.6 Chemiluminescence

[0059] Mix equal volumes of ECLA and ECLB reagents in a centrifuge tube. Place the PVDF membrane, protein side up, into the mixture for 1-2 minutes. Remove any remaining liquid, wrap the membrane, and place it in a dark box for exposure. Finally, develop and fix the membrane using developing and fixing reagents, adjusting the exposure conditions according to different light intensities.

[0060] 1.8 Pathological and Histological Observation

[0061] Brain tissue was fixed in 4% (v / v) paraformaldehyde for 24 h, then transferred to 30% (w / v) sucrose solution for dehydration, followed by OCT embedding and then frozen sectioning. Sections were blocked in 5% (w / v) BSA at room temperature for 20 min. Sections were incubated overnight at 4°C with primary antibodies (including anti-GFAP polyclonal antibody (Catalog No.: C106874, Sigma) and anti-Iba-1 antibody (Catalog No.: 019-19741, Wako)). The next day, sections were incubated with donkey anti-rabbit IgG H&L (Alexa). 488 (Abcam) secondary antibody was incubated at room temperature in the dark for 1 hour. Finally, it was incubated with ProLong (containing DAPI, Invitrogen) TM Diamond Antifade Mountant mounting allows for simultaneous staining of cell nuclei. Images were taken using a Zeiss Axio Scope A1 microscope.

[0062] 1.9 Determination of serum and brain tissue inflammatory factor levels

[0063] The procedure was performed according to the ELISA kit instructions. The brain tissue was accurately weighed, and after adding cell lysis buffer, a tissue homogenate was prepared using a high-speed homogenizer. The homogenate was centrifuged at 12,000×g and 4℃ for 15 min, and the supernatant was separated to determine the content of IL-1β and IL-6 in the brain tissue.

[0064] 1.10 Statistical Analysis

[0065] Data are expressed as mean ± standard error (x ± SEM). GraphPad Prism 9 software was used for graphing and statistical analysis. One-way ANOVA was used to compare differences between groups, and Dunnett's method was used for pairwise multiple comparisons between groups. A p-value < 0.05 was considered statistically significant. Furthermore, compared with the normal group, #p < 0.05 and ##p < 0.01; compared with the model group, *p < 0.05 and **p < 0.01.

[0066] 2. Experimental results of the therapeutic effect of the traditional Chinese medicine composition of the present invention on Alzheimer's disease.

[0067] 2.1 Effects of CFI on STZ-induced cognitive impairment in rats

[0068] To assess whether CFI treatment affects STZ-induced cognitive impairment in rats, this invention used several behavioral tests, including OFT, NORT, and MWMT. The results are as follows:

[0069] 2.1.1 Effects of CFI on STZ-induced anxiety and activity levels in rats

[0070] To measure anxiety, the time spent in the center of the open field was determined. For example... Figure 1A , 1B As shown in Figure 1C, the study of this invention found that after 6 weeks of continuous administration, rats in the STZ control group spent less time in the center of the open field than sham-operated rats (F(5,37)=4, p<0.05), while CFI treatment (3, 6, and 9 g / kg) significantly increased the time rats spent in the center of the open field (p<0.01, p<0.01, and p<0.05, respectively). There was no significant difference in mean speed between the groups, indicating that the rats' spontaneous activity capacity was not affected by STZ or CFI treatment. The results suggest that CFI can reduce STZ-induced anxiety behavior in rats.

[0071] 2.1.2 Effects of CFI on STZ-induced recognition memory in rats

[0072] like Figure 2A , 2B As shown in Figure 2C, the activity pathways of the rats indicate that rats in the sham-operated group, CFI treatment group, and donepezil group may prefer exploring new objects, while STZ-induced rats prefer to spend more time exploring familiar objects. By comparing the NORT index, STZ-induced rats showed significantly greater recognition memory impairment than the sham-operated group (F(5,37)=7.25, p<0.01), while CFI (9g / kg) significantly improved recognition memory impairment and STZ-induced cognitive impairment in rats (p<0.01).

[0073] 2.1.3 Effects of CFI on STZ-induced spatial recognition memory in rats

[0074] In the same MWMT test of spatial memory and learning function in rats, all rats exhibited shortened platform escape latency during training. By day four of training, there were significant differences in mean latency between the CFI (3, 6, and 9 g / kg) treatment groups and the STZ group (p<0.05). During the test period, STZ group rats spent significantly less time in the target quadrant compared to the sham-operated group (p<0.0001), while CFI (3 and 6 g / kg) treatment effectively improved the time spent in the target quadrant in STZ-treated rats (both p<0.05). These findings suggest that CFI can improve STZ-induced learning and memory impairments in rats, such as… Figure 3A , 3B As shown in 3C.

[0075] 2.2 Effects of CFI on STZ-induced neuroinflammation in rats

[0076] 2.2.1 Effects of CFI on the levels of inflammatory factors in STZ-induced rat brain tissue

[0077] like Figure 4A and 4B As shown, the study of this invention found that the levels of IL-6 and IL-1β in the brain tissue of STZ-induced rats were significantly reduced in the CFI treatment group.

[0078] 2.2.2 Effects of CFI on STZ-induced microglia and astrocytes in the rat cerebral cortex

[0079] like Figure 5 As shown, the study of this invention found that the activation of microglia and astrocytes in the cerebral cortex of rats induced by STZ was significantly reduced after CFI treatment (p<0.01 compared with the model group), indicating that CFI has a significant inhibitory effect on the neuroinflammation induced by STZ in rats.

[0080] 2.3 Effects of CFI on the expression of STZ-induced phosphorylated tau protein in rat brain tissue

[0081] like Figure 6 The image shows the protein blot bands representing the expression of tau protein at phosphorylation sites Thr205, Ser396, and Ser404 in rat brain tissue induced by STZ. Figure 7A , 7BFigures 7 and 7C represent the effects of CFI on the expression levels of tau protein phosphorylation sites Thr205, Ser396, and Ser404 in the brain tissue of STZ-induced rats. The study found that STZ-induced hyperphosphorylation of tau protein at these phosphorylation sites was elevated in rat brain tissue. CFI treatment significantly reduced tau protein phosphorylation (p<0.01 compared to the model group), indicating that CFI inhibits STZ-induced hyperphosphorylation of tau protein in rats.

[0082] 3. Discussion

[0083] Alzheimer's disease (AD) is a progressive neurodegenerative disease, primarily characterized by cognitive impairment. AD symptoms worsen over time. Generally, AD progression is defined as three stages: mild, moderate, and severe. The mild stage is the early stage of AD, typically characterized by cognitive impairment symptoms that do not significantly impact daily life, such as forgetting familiar words or the location of everyday objects, and short-term memory loss. In addition, patients in the mild stage may experience neuropsychiatric symptoms such as anxiety, apathy, irritability, and depression. Alleviating cognitive impairment symptoms in the mild stage of AD and slowing disease progression to reduce its impact on the patient's quality of life are crucial aspects of AD treatment.

[0084] The traditional Chinese medicine composition of the present invention contains Astragalus membranaceus, Salvia miltiorrhiza, Uncaria rhynchophylla, Ginkgo biloba leaf, Gardenia jasminoides, Cistanche deserticola, Rhodiola rosea, Corydalis yanhusuo, Ligusticum chuanxiong, Anemarrhena asphodeloides, Lycium barbarum, Raphanus sativus seed, Dipsacus asper, Phellodendron chinense, Acorus tatarinowii, Panax notoginseng, and Coptis chinensis, which have the effects of tonifying the kidney and marrow, soothing the liver and relieving depression, calming the mind and opening the orifices.

[0085] This invention employs an in vivo model of STZ-induced cognitive impairment in rats as a validation model simulating human Alzheimer's disease. In the study, rats injected with STZ intracerebrally exhibited significant cognitive impairment symptoms in behavioral experiments, including decreased spatial memory, cognitive decline, and anxious behavior. Rats treated with CFI showed significant improvement in spatial memory and cognitive abilities, suggesting that CFI has a therapeutic effect on STZ-induced cognitive impairment in rats.

[0086] The pathogenesis of Alzheimer's disease is complex, with currently accepted mechanisms including neuroinflammation and ganglion tangles caused by tau protein hyperphosphorylation. Neuroinflammation is an innate immune system response responsible for protecting the central nervous system from mechanical damage and tissue damage caused by bacterial and viral invasion. While inflammation helps clear the original stimulus, excessive inflammatory responses can also cause tissue damage. Under inflammatory conditions, microglia are activated and produce pro-inflammatory cytokines such as interleukin-6 (IL-6), tumor necrosis factor-α (TNF-α), and IL-1β. Activated microglia and neuroinflammation may lead to oxidative stress and neuronal damage, resulting in the formation of amyloid plaques and tangles in the brains of AD patients. This study suggests that the anti-AD effect of CFI may be related to its anti-neuroinflammation properties. In the CFI treatment group, the levels of pro-inflammatory cytokines IL-6 and IL-1β in the rat brain were significantly reduced, and the number of activated microglia and astrocytes in the cerebral cortex was also significantly reduced. These results suggest that inhibiting neuroinflammation may be the molecular biological mechanism of action of CFI in combating AD.

[0087] Intraneuronal neurofibrillary tangles (NFTs) are a pathological marker of Alzheimer's disease (AD), primarily composed of tau protein. Tau is a microtubule-associated protein mainly expressed in the axonal compartments of neurons, contributing to the regulation of microtubule assembly, nucleation, fasciculation, and axonal transport. Studies have shown that tau protein dysfunction is associated with tangle formation in the brain. Hyperphosphorylation of tau protein affects its affinity for tubulin and microtubule assembly, promotes tau protein relocalization to somatic dendritic compartments, and increases cytoplasmic tau protein levels, thereby enhancing the potential for tau protein interaction and aggregation, leading to the formation of tau protein multimers that cause cytotoxicity throughout the cell, particularly at synapses. Subsequently, intracellular tau protein aggregation impairs axonal transport, mitochondrial function, cytoskeleton dynamics, and increases neuronal oxidative stress, resulting in neuronal damage and cognitive deficits. This invention suggests that the anti-AD effect of CFI may be related to the inhibition of tau protein phosphorylation. In the CFI treatment group, the protein expression levels of tau protein phosphorylation sites Thr205, Ser396, and Ser404 in brain tissue were significantly reduced. This result suggests that inhibiting tau protein hyperphosphorylation may be the molecular biological mechanism of action of CFI against Alzheimer's disease (AD).

[0088] In summary, CFI has good activity in alleviating cognitive impairment and fighting AD, and its effect may be related to inhibiting neuroinflammation and inhibiting the hyperphosphorylation of tau protein.

[0089] The traditional Chinese medicine composition of the present invention also includes medically acceptable excipients, which can be formulated with the active ingredients into any pharmaceutically acceptable oral dosage form, such as granules, capsules, concentrated pills, oral liquids, or tablets. These oral dosage forms are illustrated below through some examples.

[0090] Example 1 (Tablet)

[0091] Take 1000g of this CFI (hereinafter also referred to as "the extract"), add 480g of lactose and 500g of starch, mix well, use 300g of 7% (w / v) starch slurry as a binder, wet granulate, dry, add 20g of magnesium stearate, mix well, and compress into 10,000 tablets, each containing 100mg of the extract, with a net weight of 0.2g per tablet. For oral administration, take 3 tablets twice daily for the treatment of Alzheimer's disease and cognitive impairment. Symptoms include: forgetfulness and irritability.

[0092] Example 2 (tablet)

[0093] Take 800g of this CFI, add 480g of lactose and 700g of starch, mix well, use 300g of 7% (w / v) starch slurry as a binder, wet granulate, dry, add 20g of magnesium stearate, mix well, and compress into 10,000 tablets, each containing 80mg of the extract, with a net weight of 0.2g per tablet. For oral administration, take 3 tablets twice daily for the treatment of Alzheimer's disease and cognitive impairment. Symptoms include: forgetfulness and irritability.

[0094] Example 3 (tablet)

[0095] Take 500g of this CFI, add 480g of lactose and 1000g of starch, mix well, use 300g of 7% (w / v) starch slurry as a binder, wet granulate, dry, add 20g of magnesium stearate, mix well, and compress into 10,000 tablets, each containing 50mg of the extract, with a net weight of 0.2g per tablet. For oral administration, take 3 tablets twice daily for the treatment of Alzheimer's disease and cognitive impairment. Symptoms include: forgetfulness and irritability.

[0096] Example 4 (tablet)

[0097] Take 1000g of this CFI, add 2984g of starch and mix well. Use 350g of 7% (w / v) starch slurry as a binder, wet granulate, dry, add 16g of magnesium stearate and mix well, compress into 10,000 tablets, each containing 100mg of the extract, with a net weight of 0.4g per tablet. For oral administration, take 3 tablets twice daily for the treatment of Alzheimer's disease and cognitive impairment. Symptoms include: forgetfulness and irritability.

[0098] Example 5 (tablet)

[0099] Take 800g of this CFI, add 3184g of starch and mix well. Use 350g of 7% (w / v) starch slurry as a binder, wet granulate, dry, add 16g of magnesium stearate and mix well, compress into 10,000 tablets, each containing 80mg of the extract, with a net weight of 0.4g per tablet. For oral administration, take 3 tablets twice daily for the treatment of Alzheimer's disease and cognitive impairment. Symptoms include: forgetfulness and irritability.

[0100] Example 6 (tablet)

[0101] Take 500g of this CFI, add 3484g of starch and mix well. Use 350g of 7% (w / v) starch slurry as a binder, wet granulate, dry, add 16g of magnesium stearate and mix well, compress into 10,000 tablets, each containing 50mg of the extract, with a net weight of 0.4g per tablet. For oral administration, take 3 tablets twice daily for the treatment of Alzheimer's disease and cognitive impairment. Symptoms include: forgetfulness and irritability.

[0102] Example 7 (Tablet)

[0103] Take 1000g of this CFI, add 594g of lactose and 390g of starch, mix well, use 350g of 7% (w / v) starch slurry as a binder, wet granulate, dry, add 16g of magnesium stearate, mix well, and compress into 10,000 tablets, each containing 100mg of the extract, with a net weight of 0.2g per tablet. For oral administration, take 3 tablets twice daily for the treatment of Alzheimer's disease and cognitive impairment. Symptoms include: forgetfulness and irritability.

[0104] Example 8 (tablet)

[0105] Take 800g of this CFI, add 614g of lactose and mix well, add 570g of starch and mix well, use 350g of 7% (w / v) starch slurry as a binder, wet granulate, dry, add 16g of magnesium stearate and mix well, compress into 10,000 tablets, each containing 80mg of extract, with a net weight of 0.2g per tablet. For oral administration, take 3 tablets twice daily for the treatment of Alzheimer's disease and cognitive impairment. Symptoms include: forgetfulness and irritability.

[0106] Example 9 (Tablet)

[0107] Take 500g of this CFI, add 574g of lactose and mix well. Use 350g of 7% (w / v) starch slurry as a binder, wet granulate, dry, add 16g of magnesium stearate and mix well, compress into 10,000 tablets, each containing 50mg of the extract, with a net weight of 0.2g per tablet. For oral administration, take 3 tablets twice daily for the treatment of Alzheimer's disease and cognitive impairment. Symptoms include: forgetfulness and irritability.

[0108] Example 10 (Tablet)

[0109] Take 1000 g of this CFI, add 480 g of dextrin and 1114 g of starch, mix well, use 350 g of 7% (w / v) starch slurry as a binder, wet granulate, dry, add 16 g of magnesium stearate, mix well, and compress into 10,000 tablets, each containing 100 mg of the extract, with a net weight of 0.1 g per tablet. For oral administration, take 3 tablets twice daily for the treatment of Alzheimer's disease and cognitive impairment. Symptoms include: forgetfulness and irritability.

[0110] Example 11 (Tablet)

[0111] Take 800g of this CFI, add 480g of dextrin and 1134g of starch, mix well, use 350g of 7% (w / v) starch slurry as a binder, wet granulate, dry, add 16g of magnesium stearate, mix well, and compress into 10,000 tablets, each containing 80mg of the extract, with a net weight of 0.2g per tablet. For oral administration, take 3 tablets twice daily for the treatment of Alzheimer's disease and cognitive impairment. Symptoms include: forgetfulness and irritability.

[0112] Example 12 (Tablet)

[0113] Take 500 g of this CFI, add 480 g of dextrin and 1094 g of starch, mix well, use 350 g of 7% (w / v) starch slurry as a binder, wet granulate, dry, add 16 g of magnesium stearate, mix well, and compress into 10,000 tablets, each containing 50 mg of the extract, with a net weight of 0.2 g per tablet. For oral administration, take 3 tablets twice daily for the treatment of Alzheimer's disease and cognitive impairment. Symptoms include: forgetfulness and irritability.

[0114] Example 13 (Tablet)

[0115] Take 1000g of this CFI, add 480g of lactose and 1114g of starch, mix well, use 350g of 7% (w / v) starch slurry as a binder, wet granulate, dry, add 16g of talc powder, mix well, and compress into 10,000 tablets, each containing 100mg of the extract, with a net weight of 0.2g per tablet. For oral administration, take 3 tablets twice daily for the treatment of Alzheimer's disease and cognitive impairment. Symptoms include: forgetfulness and irritability.

[0116] Example 14 (Tablet)

[0117] Take 800g of this CFI, add 480g of lactose and 1134g of starch, mix well, use 350g of 7% (w / v) starch slurry as a binder, wet granulate, dry, add 16g of talc powder, mix well, and compress into 10,000 tablets, each containing 80mg of the extract, with a net weight of 0.2g per tablet. For oral administration, take 3 tablets twice daily for the treatment of Alzheimer's disease and cognitive impairment. Symptoms include: forgetfulness and irritability.

[0118] Example 15 (Tablet)

[0119] Take 500g of this CFI, add 480g of lactose and 1094g of starch, mix well, use 350g of 7% (w / v) starch slurry as a binder, wet granulate, dry, add 16g of talc powder, mix well, and compress into 10,000 tablets, each containing 50mg of the extract, with a net weight of 0.2g per tablet. For oral administration, take 3 tablets twice daily for the treatment of Alzheimer's disease and cognitive impairment. Symptoms include: forgetfulness and irritability.

[0120] Example 16 (Tablet)

[0121] Take 1000g of this CFI, add 480g of dextrin and 504g of starch, mix well, use 350g of 7% (w / v) starch slurry as a binder, wet granulate, dry, add 16g of talc powder, mix well, and compress into 10,000 tablets, each containing 100mg of the extract, with a net weight of 0.2g per tablet. For oral administration, take 3 tablets twice daily for the treatment of Alzheimer's disease and cognitive impairment. Symptoms include: forgetfulness and irritability.

[0122] Example 17 (Tablet)

[0123] Take 800g of this CFI, add 480g of dextrin and 704g of starch, mix well, use 350g of 7% (w / v) starch slurry as a binder, wet granulate, dry, add 16g of talc powder, mix well, and compress into 10,000 tablets, each containing 80mg of the extract, with a net weight of 0.2g per tablet. For oral administration, take 3 tablets twice daily for the treatment of Alzheimer's disease and cognitive impairment. Symptoms include: forgetfulness and irritability.

[0124] Example 18 (Tablet)

[0125] Take 500g of this CFI, add 480g of dextrin and 1004g of starch, mix well, use 350g of 7% (w / v) starch slurry as a binder, wet granulate, dry, add 16g of talc powder, mix well, and compress into 10,000 tablets, each containing 50mg of the extract, with a net weight of 0.2g per tablet. For oral administration, take 3 tablets twice daily for the treatment of Alzheimer's disease and cognitive impairment. Symptoms include: forgetfulness and irritability.

[0126] Example 19 (Capsule)

[0127] Take 1000g of this CFI, add 980g of lactose and 1004g of starch, mix well, use 350g of 7% (w / v) starch slurry as a binder, wet granulate, dry, add 16g of magnesium stearate, mix well, fill into No. 1 capsules to make 10,000 capsules, each capsule contains 100mg of extract, and each capsule has a net weight of 0.3g. For oral administration, take 3 capsules twice daily for the treatment of Alzheimer's disease and cognitive impairment. Symptoms include: forgetfulness and irritability.

[0128] Example 20 (Capsule)

[0129] Take 800g of this CFI, add 980g of lactose and 1204g of starch, mix well, use 350g of 7% (w / v) starch slurry as a binder, wet granulate, dry, add 16g of magnesium stearate, mix well, fill into No. 1 capsules to make 10,000 capsules, each capsule contains 80mg of extract, each capsule has a net weight of 0.3g. For oral administration, take 3 capsules twice daily for the treatment of Alzheimer's disease and cognitive impairment. Symptoms include: forgetfulness and irritability.

[0130] Example 21 (Capsule)

[0131] Take 500g of this CFI, add 980g of lactose and 1504g of starch, mix well, use 350g of 7% (w / v) starch slurry as a binder, wet granulate, dry, add 16g of magnesium stearate, mix well, fill into No. 1 capsules to make 10,000 capsules, each capsule contains 50mg of extract, and each tablet has a net weight of 0.3g. For oral administration, take 3 capsules twice daily for the treatment of Alzheimer's disease and cognitive impairment. Symptoms include: forgetfulness and irritability.

[0132] Example 22 (Capsule)

[0133] Take 1000g of this CFI, add 2984g of starch and mix well. Use 350g of 7% (w / v) starch slurry as a binder, wet granulate, dry, add 16g of magnesium stearate and mix well, fill into No. 1 capsules to make 10,000 capsules, each with a net weight of 0.4g and containing 100mg of extract. For oral administration, take 3 capsules twice daily for the treatment of Alzheimer's disease and cognitive impairment. Symptoms include forgetfulness and irritability.

[0134] Example 23 (Capsule Formulation)

[0135] Take 800g of this CFI, add 3184g of starch and mix well. Use 350g of 7% (w / v) starch slurry as a binder, wet granulate, dry, add 16g of magnesium stearate and mix well, fill into No. 1 capsules to make 10,000 capsules, each with a net weight of 0.4g and containing 80mg of extract. For oral administration, take 3 capsules twice daily for the treatment of Alzheimer's disease and cognitive impairment. Symptoms include: forgetfulness and irritability.

[0136] Example 24 (Capsule)

[0137] Take 500g of this CFI, add 3484g of starch and mix well. Use 350g of 7% (w / v) starch slurry as a binder, wet granulate, dry, add 16g of magnesium stearate and mix well, fill into No. 1 capsules to make 100,000 capsules, each with a net weight of 0.4g and containing 50mg of extract. For oral administration, take 3 capsules twice daily for the treatment of Alzheimer's disease and cognitive impairment. Symptoms include forgetfulness and irritability.

[0138] Example 25 (Capsule)

[0139] Take 1000g of this CFI, add 1480g of lactose and 2504g of starch, mix well, use 350g of 7% (w / v) starch slurry as a binder, wet granulate, dry, add 16g of magnesium stearate, mix well, fill into size 0 capsules to make 10,000 capsules, each capsule contains 100mg of extract, each capsule has a net weight of 0.5g. For oral administration, take 3 capsules twice daily for the treatment of Alzheimer's disease and cognitive impairment. Symptoms include: forgetfulness and irritability.

[0140] Example 26 (Capsule)

[0141] Take 800g of this CFI, add 1480g of lactose and 2704g of starch, mix well, use 350g of 7% (w / v) starch slurry as a binder, wet granulate, dry, add 16g of magnesium stearate, mix well, fill into size 0 capsules to make 10,000 capsules, each capsule contains 80mg of extract, and each capsule has a net weight of 0.5g. For oral administration, take 3 capsules twice daily for the treatment of Alzheimer's disease and cognitive impairment. Symptoms include: forgetfulness and irritability.

[0142] Example 27 (Capsule)

[0143] Take 600g of this CFI, add 1480g of lactose and 2904g of starch, mix well, use 350g of 7% (w / v) starch slurry as a binder, wet granulate, dry, add 16g of magnesium stearate, mix well, fill into size 0 capsules to make 10,000 capsules, each capsule contains 50mg of extract, and each capsule has a net weight of 0.5g. For oral administration, take 3 capsules twice daily for the treatment of Alzheimer's disease and cognitive impairment. Symptoms include: forgetfulness and irritability.

[0144] Example 28 (Capsule)

[0145] Take 500g of this CFI, add 1480g of lactose and 3004g of starch, mix well, use 350g of 7% (w / v) starch slurry as a binder, wet granulate, dry, add 16g of magnesium stearate, mix well, fill into No. 2 capsules to make 10,000 capsules, each capsule contains 50mg of extract, and each capsule has a net weight of 0.2g. For oral administration, take 3 capsules twice daily for the treatment of Alzheimer's disease and cognitive impairment. Symptoms include: forgetfulness and irritability.

[0146] Example 29 (Capsule)

[0147] Take 400g of this CFI, add 480g of lactose and 1104g of starch, mix well, use 350g of 7% (w / v) starch slurry as a binder, wet granulate, dry, add 16g of magnesium stearate, mix well, fill into No. 2 capsules to make 10,000 capsules, each capsule contains 40mg of extract, and each capsule has a net weight of 0.2g. For oral administration, take 3 capsules twice daily for the treatment of Alzheimer's disease and cognitive impairment. Symptoms include: forgetfulness and irritability.

[0148] Example 30 (Capsule)

[0149] Take 250g of this CFI, add 580g of lactose and 1154g of starch, mix well, use 350g of 7% (w / v) starch slurry as a binder, wet granulate, dry, add 16g of magnesium stearate, mix well, fill into No. 2 capsules to make 10,000 capsules, each capsule contains 25mg of extract, and each capsule has a net weight of 0.2g. For oral administration, take 3 capsules twice daily for the treatment of Alzheimer's disease and cognitive impairment. Symptoms include: forgetfulness and irritability.

[0150] Example 31 (Oral Liquid)

[0151] Take 50g of this CFI, add 15% (v / v) ethanol, and stir thoroughly. Then add appropriate excipients (such as 0.1% (w / v) sorbic acid as an antibacterial agent, 0.1% (w / v) citric acid as an antioxidant, and 0.1% (w / v) mannitol as a flavoring agent), dissolve evenly, filter to clarify, and fill the contents into ampoules or easy-open bottles, then sterilize. The specification is an oral solution containing 166mg of extract per bottle, with a net content of 10mL per bottle. For oral administration, once daily, one bottle each time, for the treatment of Alzheimer's disease and cognitive impairment. Symptoms include: forgetfulness and irritability.

[0152] Example 32 (Oral Liquid)

[0153] Take 100g of this CFI, add 15% (v / v) ethanol, and stir thoroughly. Then add appropriate excipients (such as 0.1% (w / v) sorbic acid as an antibacterial agent, 0.1% (w / v) citric acid as an antioxidant, and 0.1% (w / v) mannitol as a flavoring agent), dissolve evenly, filter to clarify, and fill the contents into ampoules or easy-open bottles, then sterilize. The specification is an oral solution containing 300mg of extract per bottle, with a net content of 10mL per bottle. For oral administration, once daily, one bottle each time, for the treatment of Alzheimer's disease and cognitive impairment. Symptoms include: forgetfulness and irritability.

[0154] Example 33 (Oral Liquid)

[0155] Take 200g of this CFI, add 15% (v / v) ethanol, and stir thoroughly. Then add appropriate excipients (such as 0.1% (w / v) sorbic acid as an antibacterial agent, 0.1% (w / v) citric acid as an antioxidant, and 0.1% (w / v) mannitol as a flavoring agent), dissolve evenly, filter to clarify, and fill the contents into ampoules or easy-open bottles, then sterilize. The specification is an oral solution containing 600mg of extract per bottle, with a net content of 10mL per bottle. For oral administration, once daily, one bottle each time, for the treatment of Alzheimer's disease and cognitive impairment. Symptoms include: forgetfulness and irritability.

[0156] Example 34 (Oral Liquid)

[0157] Take 300g of this CFI, add 15% (v / v) ethanol, and stir thoroughly. Then add appropriate excipients (such as 0.1% (w / v) sorbic acid as an antibacterial agent, 0.1% (w / v) citric acid as an antioxidant, and 0.1% (w / v) mannitol as a flavoring agent), dissolve evenly, filter to clarify, and fill the contents into ampoules or easy-open bottles, then sterilize. The specification is an oral solution containing 900mg of extract per bottle, with a net content of 10mL per bottle. For oral administration, once daily, one bottle each time, for the treatment of Alzheimer's disease and cognitive impairment. Symptoms include: forgetfulness and irritability.

[0158] Example 35 (Oral Liquid)

[0159] Take 400g of this CFI, add 15% (v / v) ethanol, and stir thoroughly. Then add appropriate excipients (such as 0.1% (w / v) sorbic acid as an antibacterial agent, 0.1% (w / v) citric acid as an antioxidant, and 0.1% (w / v) mannitol as a flavoring agent), dissolve evenly, filter to clarify, and fill the contents into ampoules or easy-open bottles, then sterilize. The specification is an oral solution containing 1333mg of extract per bottle, with a net content of 10mL per bottle. For oral administration, once daily, one bottle each time, for the treatment of Alzheimer's disease and cognitive impairment. Symptoms include: forgetfulness and irritability.

[0160] Example 36 (Oral Liquid)

[0161] Take 500g of this CFI, add 15% (v / v) ethanol, and stir thoroughly. Then add appropriate excipients (such as 0.1% (w / v) sorbic acid as an antibacterial agent, 0.1% (w / v) citric acid as an antioxidant, and 0.1% (w / v) mannitol as a flavoring agent), dissolve evenly, filter to clarify, and fill the contents into ampoules or easy-open bottles, then sterilize. The specification is an oral solution containing 1666mg of extract per bottle, with a net content of 10mL per bottle. For oral administration, once daily, one bottle each time, for the treatment of Alzheimer's disease and cognitive impairment. Symptoms include: forgetfulness and irritability.

[0162] In summary, the inventors have demonstrated through in vivo experiments that CFI can significantly improve cognitive impairment. This invention reveals that CFI has a good therapeutic effect on STZ-induced cognitive impairment and can be used to prepare drugs and functional foods that improve cognitive impairment, providing a new option for the treatment of Alzheimer's disease. Currently, the pathogenesis of AD is related to ganglion intractability caused by excessive phosphorylation of tau protein in brain tissue and neuroinflammation. The experiments of this invention have confirmed that CFI can significantly reduce the content of inflammatory factors IL-6 and IL-1β in brain tissue and inhibit the activation of microglia and astrocytes in the cerebral cortex, elucidating the anti-AD mechanism of CFI from the perspective of neuroinflammatory response. In addition, CFI inhibits the protein expression level of excessive phosphorylation of tau protein, elucidating the mechanism of CFI in treating AD from the perspective of inhibiting the production of ganglion intractability.

[0163] The background section of this invention may include background information about the problems or environment in which the invention is being developed, and is not necessarily a description of prior art. Therefore, the content included in the background section does not constitute an admission of prior art by the applicant.

[0164] The above description provides a further detailed explanation of the present invention in conjunction with specific / preferred embodiments, and it should not be construed that the specific implementation of the present invention is limited to these descriptions. For those skilled in the art, various substitutions or modifications can be made to these described embodiments without departing from the concept of the present invention, and all such substitutions or modifications should be considered within the scope of protection of the present invention. In the description of this specification, the reference to terms such as "an embodiment," "some embodiments," "preferred embodiment," "example," "specific example," or "some examples," etc., indicates that the specific features, structures, materials, or characteristics described in connection with that embodiment or example are included in at least one embodiment or example of the present invention. In this specification, the illustrative expressions of the above terms do not necessarily refer to the same embodiment or example. Moreover, the specific features, structures, materials, or characteristics described can be combined in any suitable manner in one or more embodiments or examples. Without contradiction, those skilled in the art can combine and integrate the different embodiments or examples described in this specification and the features of different embodiments or examples. Although the embodiments of the present invention and their advantages have been described in detail, it should be understood that various changes, substitutions, and modifications can be made herein without departing from the scope of protection of the patent application.

Claims

1. A traditional Chinese medicine composition for treating Alzheimer's disease, characterized in that, The active ingredients in the traditional Chinese medicine composition are made from the following raw materials in parts by weight: Astragalus membranaceus 6-50g, Salvia miltiorrhiza 6-50g, Uncaria rhynchophylla 6-50g, Ginkgo biloba 3-30g, Gardenia jasminoides 3-30g, Cistanche deserticola 3-30g, Rhodiola rosea 3-30g, Corydalis yanhusuo 3-30g, Ligusticum chuanxiong 3-30g, Anemarrhena asphodeloides 3-30g, Lycium barbarum 3-30g, Raphanus sativus 3-30g, Dipsacus asper 3-30g, Phellodendron chinense 3-30g, Acorus tatarinowii 3-30g, Panax notoginseng 3-30g, Coptis chinensis 2-30g.

2. The traditional Chinese medicine composition for treating Alzheimer's disease according to claim 1, characterized in that, The weight proportions of each raw material in the effective ingredients are as follows: Astragalus membranaceus 15, Salvia miltiorrhiza 15, Uncaria rhynchophylla 14, Ginkgo biloba 10, Gardenia jasminoides 10, Cistanche deserticola 10, Rhodiola rosea 10, Corydalis yanhusuo 10, Ligusticum chuanxiong 10, Anemarrhena asphodeloides 10, Lycium barbarum 10, Raphanus sativus 10, Dipsacus asper 10, Phellodendron chinense 10, Acorus tatarinowii 9, Panax notoginseng 6, and Coptis chinensis 4.

3. The traditional Chinese medicine composition for treating Alzheimer's disease according to claim 1, characterized in that, The traditional Chinese medicine composition also includes medically acceptable excipients.

4. The traditional Chinese medicine composition for treating Alzheimer's disease according to any one of claims 1-3, characterized in that, The traditional Chinese medicine composition is any pharmaceutically acceptable oral dosage form.

5. The traditional Chinese medicine composition for treating Alzheimer's disease according to claim 4, characterized in that, The traditional Chinese medicine composition is in the form of granules, capsules, concentrated pills, oral liquids, or tablets.

6. The traditional Chinese medicine composition for treating Alzheimer's disease as described in any one of claims 1-3, characterized in that: The active ingredients in the traditional Chinese medicine composition are prepared by extracting the raw materials by heating after soaking them in a solvent.

7. A method for preparing a traditional Chinese medicine composition for treating Alzheimer's disease according to any one of claims 1-6, characterized in that, Includes the following steps: (1) Weigh each raw material according to the ratio, soak it in distilled water at room temperature for 12-24 hours in 10-20 times the weight of all the medicinal materials, heat and extract for 0.5-2 hours, filter, and repeat the above extraction steps twice; (2) Combine the three filtrates, remove all water under reduced pressure, and freeze dry to obtain a fine powder of the active ingredient of the Chinese herbal composition.

8. The preparation method according to claim 7, characterized in that, It also includes the following steps: (4) The fine powder and a predetermined amount of medically acceptable excipients are prepared into any pharmaceutically acceptable oral dosage form.

9. The use of the traditional Chinese medicine composition according to any one of claims 1-6 in the preparation of a medicament for treating Alzheimer's disease.

10. The application as described in claim 9, characterized in that, The Alzheimer's disease mentioned refers to streptozotocin (STZ)-induced Alzheimer's disease.

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