A pharmafood product for improving memory and application thereof

Abstract

The application discloses a kind of improving memory of food-herbal product and application, food-herbal product contains Chinese medicinal material, the Chinese medicinal material is composed of Fructus Alpiniae Oxophyllae, lily, lotus leaf, or, the Chinese medicinal material is composed of Fructus Alpiniae Oxophyllae extract, lily extract, lotus leaf extract.The product of the application intervenes memory decline related pathway by regulating JUN, FOS, SRC, TP53, ESR1 and other key target points, effectively improves the spatial memory and recognition memory ability of rat, reduces hippocampal CA1 region neuron morphological structure damage, increases the number of neuron and Nissl body and up-regulates the expression of neurotrophic factor, human taste test test proves that the product can improve the memory of memory decline population Memory quotient, Montreal cognitive assessment scale and psychological resilience scale score, reduce Hamilton anxiety scale and fatigue rating scale score, relieve the core symptoms of TCM such as turning around forgetting, forgetting easily, spirit fatigue and so on, can be used to improve memory.

Description

Technical Field

[0001] This invention relates to the field of health and wellness products, specifically to a food-medicine homology product that improves memory. Background Technology

[0002] Memory, a fundamental function in daily life, is closely related to aging, chronic diseases, lifestyle, education level, and apolipoprotein E4 (APOE) genotype. Memory decline is not a single, isolated symptom, but rather a continuous spectrum encompassing everything from normal aging to severe neurodegenerative diseases, including age-related memory impairment, subjective cognitive decline, mild cognitive impairment, and dementia. Memory loss is prevalent in the population, affecting quality of life and work efficiency, and increasing the risk of dementia.

[0003] Modern research has found that the memory decline process can be stabilized or even reversed by taking appropriate measures, preventing progression to a disease state (Zahodne LB, Wall MM, Schupf N, et al. Late-life memorytrajectories in relation to incident dementia and regional brain atrophy[J].J Neurol. 2015, 262:2484-90). Some compounds have been found to improve cognitive function. These nootropic drugs can enhance cognition through various neurotransmitter systems, such as inhibiting acetylcholinesterase activity (e.g., donepezil, galantamine), regulating the dopaminergic system (e.g., amphetamine, modafinil, armodafinil), or antagonizing NMDA receptors (e.g., memantine).

[0004] However, most of the aforementioned medications are prescription drugs, primarily targeting clearly defined pathological cognitive impairments (such as Alzheimer's disease and vascular dementia). Long-term use of these prescription drugs may produce varying degrees of side effects. For example, donepezil and galantamine commonly cause nausea and vomiting. Donepezil may also cause arrhythmias, mental disturbances, and extrapyramidal syndromes. Amphetamine-type drugs have adverse reactions including headache, anxiety, tremors, anorexia, sweating, and stomach upset, and carry significant risks of addiction and drug dependence; they are controlled substances. Furthermore, prescription drugs are not suitable for daily health maintenance in the general population for physiological memory decline caused by aging, stress, or fatigue. DHA supplements, phosphatidylserine, and ginkgo biloba extract, as supplements for improving memory, have single ingredients, limited targets, and lack high-quality clinical evidence. Summary of the Invention

[0005] The purpose of this invention is to provide a food-medicine product that can improve memory, offering a safe, effective, and suitable daily health care product for people with memory decline who have not yet reached a disease state.

[0006] In a first aspect, the present invention provides a food-medicine product for improving memory, containing traditional Chinese medicine ingredients, wherein the traditional Chinese medicine ingredients are composed of Alpinia oxyphylla, lily bulb, and lotus leaf, or wherein the traditional Chinese medicine ingredients are composed of Alpinia oxyphylla extract, lily bulb extract, and lotus leaf extract.

[0007] Optional or preferred, the dosage of Alpinia oxyphylla, lily bulb, and lotus leaf is calculated by weight as follows: 5 parts Alpinia oxyphylla, 5 parts lily bulb, and 3 parts lotus leaf.

[0008] Secondly, the present invention provides an application of a food-medicine homology product in the preparation of a memory-improving product, wherein the food-medicine homology product contains traditional Chinese medicine raw materials, wherein the traditional Chinese medicine raw materials are composed of Alpinia oxyphylla, lily bulb, and lotus leaf, or wherein the traditional Chinese medicine raw materials are composed of Alpinia oxyphylla extract, lily bulb extract, and lotus leaf extract.

[0009] Optional or preferred, the dosage of Alpinia oxyphylla, lily bulb, and lotus leaf is calculated by weight as follows: 5 parts Alpinia oxyphylla, 5 parts lily bulb, and 3 parts lotus leaf.

[0010] Optionally or preferably, the memory-improving product is a plant-based beverage, which is prepared by the following method: (1) Powdered and mixed Alpinia oxyphylla, Lilium brownii and Nelumbo nucifera leaves were extracted in two stages. In the first stage of extraction, 7 times the volume of 50% ethanol aqueous solution was added and extracted at room temperature for 1 hour. The filtrate was collected. The residue was subjected to the second stage of extraction. 6 times the volume of 50% ethanol aqueous solution was added and extracted at room temperature for 1 hour. The filtrate was collected from the two stages of extraction. The filtrates were combined and concentrated at 60-70℃ to a density of 1.10-1.15 g / mL to obtain concentrated extract. (2) Based on the calculation per gram of concentrated extract, add 0.14g of water-soluble mono- and diglycerides of fatty acids, 0.02g of soybean lecithin, and 0.200g of γ-cyclodextrin, respectively; mix evenly and finally pass through a 400-mesh sieve, add purified water, heat to 80℃ and stir to dissolve for 30 minutes, cool to room temperature, pass the liquid through a 400-mesh sieve again before filling, fill into 30 mL bags, sterilize and fill to obtain the plant beverage. The present invention provides the following beneficial effects.

[0011] Compared with the prior art, the present invention has the following beneficial effects: This application describes a formula composed of the traditional Chinese medicine ingredients Alpinia oxyphylla, lily bulb, and lotus leaf. Based on the clinical experience of Professor Zhang Yunling, a renowned traditional Chinese medicine practitioner and a recipient of the first Qihuang Scholar award, and combined with previous literature and clinical research, this formula was developed using subjects with memory impairment as the research subjects. The basic treatment principle is "warming the kidneys and spleen, calming the mind and soothing the nerves, and promoting the upward movement of clear yang." This formula is a functional traditional Chinese medicine ingredient used in a food-medicine homology product called Yihe Yin to improve memory. In over 10 years of clinical practice, it has achieved good therapeutic effects, improving memory and learning abilities, alleviating anxiety, and improving sleep.

[0012] In this formula, the principal herb is Alpinia oxyphylla, which enters the kidney and spleen meridians. Its main components are volatile oils, terpenes, flavonoids, and diarylheptane compounds, which possess various pharmacological activities such as neuroprotection, cognitive improvement, antidiarrheal, sedation, anti-inflammation, and antioxidant effects. The volatile oil, chloroform extract, and n-butanol extract of Alpinia oxyphylla have been shown to improve learning and memory deficits by regulating hippocampal neuronal apoptosis, regulating the activity of central ACh synthase and degradative enzymes, inhibiting β-secretase activity and reducing Aβ (1–42) levels, alleviating oxidative stress, and regulating eosinophilic degeneration of neurons.

[0013] Lily bulb, a key ingredient in this formula, enters the lung and heart meridians, and its functions include nourishing yin, moistening dryness, relieving cough, clearing the heart, and calming the mind. Water extracts of lily bulb have been found to enhance memory retention and spatial memory in memory-deficient mice by increasing the activation of brain-derived neurotrophic factor (BDNF) / cAMP response element-binding protein (CREB) and mitogen-activated protein kinase / extracellular signal-regulated kinase (MEK / ERK) in the hippocampus, thereby improving synaptic plasticity and learning and memory abilities.

[0014] Lotus leaf, used as an adjuvant herb, enters the liver, spleen, and stomach meridians. Its main functions are clearing summer heat and dampness, promoting the upward movement of clear yang, cooling the blood, and stopping bleeding. The lotus leaf alkaloid derivative (2-hydroxy-1-methoxyaporphine) in lotus leaf has high acetylcholinesterase inhibitory activity, which can correct the decline in the synthesis, release, and uptake of acetylcholine, promote the release of neurotransmitters, and correct the decline in memory and cognitive function. Lotus leaf flavonoid extract has been found to have sedative and hypnotic effects, shortening the sleep latency and prolonging sleep time.

[0015] The combination of Alpinia oxyphylla, lily bulb, and lotus leaf can enhance intelligence, calm the mind, and relieve anxiety.

[0016] All components of this product are derived from substances that are both food and medicine according to the "Announcement of Substances That Are Traditionally Both Food and Chinese Medicine" issued by the National Health Commission and the State Administration for Market Regulation. No non-food ingredients or non-food and medicine ingredients are added. No obvious adverse reactions have been found. It is suitable for long-term daily health care consumption by people with various types of memory decline.

[0017] Network pharmacology and molecular docking revealed that the product of this invention may intervene in memory decline through targets such as JUN, FOS, SRC, TP53, and ESR1. Animal experiments showed that the product of this invention can effectively improve spatial memory and recognition memory in rats. It exerts a neuroprotective effect by repairing the morphology, structure, and arrangement of neurons in the CA1 region of the hippocampus, increasing the number of neurons and Nissl bodies, and upregulating the expression of various neurotrophic factors, thereby improving memory ability.

[0018] Human trials have shown that the product of this invention can improve memory quotient, MoCA score, and CD-RISC score in people with memory loss, reduce HAMA score and FAS score, and improve symptoms in traditional Chinese medicine such as forgetfulness, absent-mindedness, dull expression, apathy, depression, fatigue, lethargy, drowsiness, slow reaction, and slow thinking.

[0019] In summary, the product of this invention can intervene in memory-related pathways by regulating key targets such as JUN, FOS, SRC, TP53, and ESR1, effectively improving spatial and recognition memory abilities in rats, reducing morphological and structural damage to neurons in the CA1 region of the hippocampus, increasing the number of neurons and Nissl bodies, and upregulating the expression of neurotrophic factors. Human trials have demonstrated that Yiheyin can also improve memory quotient, Montreal Cognitive Assessment Scale and Psychological Resilience Scale scores in people with memory loss, reduce Hamilton Anxiety Scale and Fatigue Rating Scale scores, and alleviate core symptoms in traditional Chinese medicine such as forgetfulness, absent-mindedness, and fatigue, thus it can be used to improve memory. Attached Figure Description

[0020] Figure 1 This is a production flow chart of the plant-based beverage Yiheyin in Example 1.

[0021] Figure 2 This is the network diagram of "Traditional Chinese Medicine - Main Active Ingredients - Potential Targets" in Example 2.

[0022] Figure 3 This is a Venn diagram of the memory loss target in Example 2.

[0023] Figure 4 This is a Venn diagram of the drug-disease common target in Example 2.

[0024] Figure 5 This is a visualization of the PPI protein interaction network of the overlapping therapeutic targets in Example 2.

[0025] Figure 6 This is a graph showing the GO analysis results of the intersection targets of Yiheyin in treating memory loss in Example 2.

[0026] Figure 7 This is a KEGG analysis result of the intersection target of Yihe Decoction in treating memory loss in Example 2.

[0027] Figure 8 The statistical results of neurological deficit scores in each group of rat models in Example 3 are as follows ( n = 10), NS indicates that there is no significant difference between groups.

[0028] Figure 9 The average swimming speed of rats in each group during the Morris water maze experiment in Example 3 is statistically analyzed. n =10), NS indicates that there is no significant difference between groups.

[0029] Figure 10 The statistical results of the escape latency of each group of rats in the localization and navigation experiment in Example 3 are as follows ( n = 10), Compared with the sham surgery group, P <0.05, Compared with the sham surgery group, P <0.01, # Compared with the model group, P <0.05, ## Compared with the model group, P <0.01.

[0030] Figure 11 The statistical results of the number of times the rats traversed the space exploration test platform in each group in Example 3 are as follows ( n = 10), Compared with the sham surgery group, P <0.01, ## Compared with the model group, P <0.01.

[0031] Figure 12 The statistical results of the dwell time in the target quadrant of rats in each group in Example 3 are as follows ( n = 10), Compared with the sham surgery group, P <0.05, # Compared with the model group, P <0.05.

[0032] Figure 13 The statistical results of the discrimination index of rats in the training phase of the new object recognition test in each group of rats in Example 3 are as follows: n = 10), NS indicates that there is no significant difference between groups.

[0033] Figure 14 The statistical results of the discrimination index of rats in the new object recognition test of each group of rats in Example 3 are as follows: n = 10), Compared with the sham surgery group, P <0.05, # Compared with the model group, P <0.05.

[0034] Figure 15 The results of HE staining of the hippocampus of rats in each group in Example 3 are shown.

[0035] Figure 16 The statistical results of the number of neurons in the CA1 region of the hippocampus of rats in each group in Example 3 ( n = 5), Compared with the sham surgery group, P <0.01, # Compared with the model group, P <0.05.

[0036] Figure 17 The results of Nielsen staining of the hippocampus in each group of rats in Example 3 are shown.

[0037] Figure 18 The statistical results of the number of Nissl bodies in the CA1 region of the hippocampus of each group of rats in Example 3 ( n = 5), Compared with the sham surgery group, P <0.05, # Compared with the model group, P <0.05.

[0038] Figure 19 The statistical results of BDNF, NGF, NT-3, and NT-4 levels in the hippocampus of rats in each group in Example 2 are as follows: n = 5), Compared with the sham surgery group, P <0.05, Compared with the sham surgery group, P <0.01, # Compared with the model group, P <0.05, ## Compared with the model group, P <0.01. Detailed Implementation

[0039] To enable those skilled in the art to better understand the technical solutions of the present invention, the technical solutions of the present invention will be clearly and completely described below in conjunction with the embodiments and accompanying drawings. Based on the embodiments in this application, other similar embodiments obtained by those skilled in the art without creative effort should all fall within the scope of protection of this application.

[0040] Example 1: Optimization of Concentration and Extraction Process of Traditional Chinese Medicine Composition and Preparation of Plant Beverage The ingredients and proportions of each Chinese herbal raw material are as follows: 5g of Alpinia oxyphylla, 5g of lily bulb, and 3g of lotus leaf.

[0041] Optimization of the concentration and extraction process of traditional Chinese medicine compositions: 1. Determination of Product Quality Evaluation Indicators and Testing Methods Based on preliminary literature review, the types of components contained in the three medicinal herbs, Alpinia oxyphylla, lotus leaf, and lily bulb, were summarized and compiled. This included the main active ingredients and the functional components that play a role in nourishing yin and strengthening the spleen, calming the mind and improving intelligence; all of which can be used as quality evaluation indicators for the product. Given the complex composition of the extract and the significant differences in the physical and chemical properties of various components, it is impossible to extract them all using a single extraction method. Therefore, based on their proportion in the formula, their content, their correlation with the target efficacy, and considering the influence of pharmacopoeia standards and the simplicity and accuracy of detection in practical operations, the quality markers for this formula product were determined to be lotus leaf alkaloids, total flavonoids, and terpenes.

[0042] The product quality evaluation and analysis methods are shown in Table 1 below.

[0043] Table 1. Detection and Analysis Methods for Product Quality Evaluation Indicators 2. Extraction process development Based on the traditional process (decoction), some process parameters have been optimized and improved. The optimized parameters are the type and concentration of extraction solvent, the dosage, extraction temperature, extraction time and extraction grade.

[0044] 2.1 Investigation of extraction solvent and concentration Accurately weigh equal amounts of Chinese herbal raw materials, pulverize them through a 60-mesh sieve, and investigate the extraction solvents of water, 50%, 60%, and 70% ethanol (all solvents are water, all are volume percentages). The extraction volume was increased by 15 times, the extraction time was 2 hours, and the temperature was room temperature with stirring (water was extracted by reflux). After filtration, the supernatant was collected. The transfer rates of lotus leaf alkaloids, total flavonoids, and terpenoids in the final extract were used as evaluation indicators, as shown in Table 2 below. Finally, 50% ethanol was determined to be the optimal solvent.

[0045] Table 2 Extraction rates of product quality evaluation indicators in the extract when water and ethanol of different concentrations are used as extraction solvents. 2.2 Investigation of extraction temperature A precise weight of the same amount of Chinese herbal raw material was taken, pulverized through a 60-mesh sieve, and extracted with 50% ethanol (50% ethanol aqueous solution by volume) as the solvent. The extraction volume was 15 times the original volume, and the extraction time was 2 hours. The temperature conditions were investigated at 25℃, 50℃, and 70℃. After filtration, the supernatant was collected. The transfer rates of lotus leaf alkaloids, total flavonoids, and terpenoids in the final extract were used as evaluation indicators, as shown in Table 3 below. Considering actual production energy consumption and subsequent formulation issues, 25℃ was finally determined as the optimal extraction temperature.

[0046] Table 3 Extraction rates of product quality evaluation indicators in the extract at different extraction temperatures 2.3 Investigation of Solvent Multiplier Accurately weigh equal amounts of the herbal raw materials, pulverize them through a 60-mesh sieve, and extract them using 50% ethanol (50% ethanol aqueous solution by volume) as the solvent. Extraction time is 2 hours, and the temperature is room temperature with stirring. The extraction ratios are 10, 15, and 20 times. After filtration, the supernatant is collected. The transfer rates of lotus leaf alkaloids, total flavonoids, and terpenoids in the final extract are used as evaluation indicators, as shown in Table 4 below. The final extraction ratio for ethanol is determined to be 15 times.

[0047] Table 4 Extraction rates of product quality evaluation indicators in the extract at different solvent concentrations 2.4 Examination of extraction time Accurately weigh equal amounts of Chinese herbal raw materials, pulverize them through a 60-mesh sieve, use 50% ethanol (50% ethanol aqueous solution by volume) as the extraction solvent, apply 15 times the volume ratio, and extract for 0.5, 1.0, 1.5, and 2.0 hours at room temperature with stirring. After filtration, collect the supernatant. The transfer rates of lotus leaf alkaloids, total flavonoids, and terpenoids in the final extract were used as evaluation indicators, as shown in Table 5 below. The final extraction time for each stage was determined to be 1.5 hours.

[0048] Table 5 Extraction rates of product quality evaluation indicators in extracts at different extraction times 2.5 Examination of Extraction Levels Accurately weigh equal amounts of Chinese herbal raw materials, pulverize them through a 60-mesh sieve, and use 50% ethanol (50% ethanol aqueous solution by volume) as the extraction solvent. Maintain room temperature with stirring. Optimize the extraction stages by reducing the amount of raw material, increasing the number of stages, and correspondingly reducing the extraction time. Collect the supernatant after filtration. The transfer rates of lotus leaf alkaloids, total flavonoids, and terpenoids in the final extract are used as evaluation indicators, as shown in Table 6 below. The final extraction stages are determined to be two.

[0049] Table 6 Extraction rates of product quality evaluation indicators in the extract at different extraction stages 2.6 Determination of Extraction Process Parameters Table 7 Extraction rates of product quality evaluation indicators in the extract under different process parameters As can be seen from the table above, after two-stage extraction, the extraction rate of the target components is basically above 80%.

[0050] Based on the above experimental results, the final extraction process was determined as follows: 7 / 6 times the amount (7 times the amount for the first stage and 6 times the amount for the second stage) of 50% ethanol (50% ethanol aqueous solution by volume) at room temperature (25℃) for two stages of extraction, each stage for 1 hour, and the filtrates were combined after passing through a 400-mesh sieve.

[0051] 3. Development of plant-based beverage formulation technology Once the formulation and extraction process are determined, the key issues to be addressed in producing palatable plant-based beverages are the solubility of the extract, the taste of the formulation, and the formulation preparation process.

[0052] 3.1 Concentration Process After the target components are extracted using the optimal process, the extract needs to be concentrated to remove the solvent. During concentration, issues such as material precipitation and insolubility may occur. Therefore, through small-scale laboratory trials, it was determined that, depending on the amount of components to be dissolved, a co-solvent can be added during concentration to achieve an encapsulation effect. Thus, the process parameters for concentration were determined as follows: concentration temperature 60-70℃, concentration to a density of 1.10-1.15 g / mL to obtain a concentrated extract. The excipients added are: 0.14 g of water-soluble mono- and diglycerides of fatty acids, 0.02 g of soybean lecithin, and 0.200 g of γ-cyclodextrin per gram of concentrated extract; and the final product must be passed through a 400-mesh sieve.

[0053] 3.2 Preparation process After concentration, the extract requires the addition of flavor-correcting excipients to improve taste. The proportion of excipients added was determined based on the ratio of components per gram of concentrated extract. Therefore, during preparation, the concentrated extract, various excipients (including solubilizers and flavoring agents), and purified water were added in the specified proportions. The mixture was then heated to 80°C, stirred for 30 minutes to dissolve, and then cooled to room temperature. The solution was filtered through a 400-mesh sieve before bottling.

[0054] 3.3 Filling process The prepared liquid beverage is bottled in 30 mL sachets and named Yiheyin. During packaging, 10 sachets are randomly sampled every 20 minutes to check for volume variation, appearance, and sealing. Volume variation must not exceed ±0.5 mL.

[0055] 3.4 Sterilization process The packaged material (plant beverage) is placed into the sterilization device, the device is pressurized and heated, and the timer is set when the temperature reaches 115℃. After 20 minutes, the sterilization is completed, the device is depressurized and cooled, and the material is taken out and drained.

[0056] 3.5 Production of Sample Products In April 2025, Hebei Baixiaodan Pharmaceutical Co., Ltd. produced the products required for human trials of the plant-based beverage, in accordance with the product's process specifications. Product information is as follows: Table 8 Product Information of Yihe Drink, a Plant Beverage Example 2 Network Pharmacology Analysis of Yihe Drink 1 Method 1.1 Screening of Active Ingredients and Target Prediction The traditional Chinese medicine raw materials of Yihe Drink consist of lily, semen alpiniae oxyphyllae, and lotus leaf, which are the functional ingredients that enable this plant beverage to improve memory. First, the pinyin of each component drug was entered into the TCMSP database (https: / / tcmspw.com / tcmsp.php) and the HERB2.0 database (http: / / 47.92.70.12) in sequence to obtain their ingredient information. In the TCMSP database, the screening criteria were set as oral bioavailability (OB) not less than 30% and drug-likeness (DL) ≥ 0.18. According to this screening standard, potential active ingredients were screened out. By integrating the results obtained from the two databases, the active ingredients of Yihe Drink and their corresponding target proteins were determined. Further, with the help of the Uniprot database (http: / / www.uniprot.org), the above target proteins were verified and uniformly converted into standard gene names, restricting the species to "Homosapiens", and finally, the target genes corresponding to the active ingredients in Yihe Drink were obtained.

[0057] 1.2 Screening of Targets for Memory Decline Using "MemoryDisorders" as the keyword, relevant therapeutic targets for memory decline were searched in the GeneCards database (https: / / www.genecards.org / ), DrugBank database (http / / www.drugbank.ca / ), OMIM database (http: / / www.omim.org / ), and TTD database (http: / / database.idrb.cqu.edu.cn / TTD / ). And the jvenn (https: / / jvenn.toulouse.inra.fr / ) online tool was used to draw the Venn diagram of the memory decline targets. The jvenn (https: / / jvenn.toulouse.inra.fr / ) was used for visual analysis of the intersection targets of drugs and diseases and to draw the Venn diagram.

[0058] 1.3 Construction of the "Traditional Chinese Medicine - Main Active Ingredients - Potential Targets" Interaction Network The main active ingredients of lily, Alpinia oxyphylla, and lotus leaf, along with common target genes related to memory decline, were imported into Cytoscape 3.10.0 software to establish a "traditional Chinese medicine-ingredient-target" interaction network and generate a visualization map. Finally, the core active ingredients that play a key role in the treatment of memory decline by Yiheyin were screened.

[0059] 1.4 Constructing a protein-protein interaction (PPI) network of potential targets Based on the intersection targets obtained through screening, a PPI network of potential therapeutic targets was obtained using the STRING11.0 database (https: / / string-db.org / ) with the biological species set as "Homo Sapiens" and the minimum interaction threshold set as "Highest Confidence" (>0.9). The obtained graph data was imported into the built-in plugin of the visualization software CytoScape to perform topological analysis on the PPI network. By calculating the degree values ​​of the nodes in the network, core targets were screened out.

[0060] 1.5 Gene Ontology (GO) enrichment, Kyoto enrichment, and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis The potential targets of Yiheyin in treating memory loss were imported into RStudio, and enrichment analyses were performed on the common targets of the drug components and the disease, including biological processes (BP), cellular components (CC), molecular functions (MF), and KEGG. Based on the enrichment analysis results, the top 10 significant items in the GO analysis and the top 30 in the KEGG pathway analysis were selected and visualized.

[0061] 1.6 Molecular docking verification Based on the PPI results, the top 5 compounds were selected for molecular docking verification with the core target. Molecular structure files of the main active ingredients were obtained from the PubChem database, and the receptor protein structure was retrieved and downloaded from the PDB database. The receptor protein was pretreated using PyMOL software, including the removal of water molecules and proligands. Pretreatment operations such as hydrogenation and charge balancing were performed using AutoDock Tools software. Molecular docking was completed using AutoDock Vina, and the results were finally visualized using PyMOL.

[0062] 2 Results 2.1 Screening of active ingredients in Yiheyin By searching the TCMSP and HERB2.0 databases and supplementing with relevant published literature, after screening and removing duplicates, 55 effective active ingredients were found in the three traditional Chinese medicines of Yihe Yin. 620 potential target proteins were identified through screening. The interaction relationships between each traditional Chinese medicine, active ingredient, and potential target protein are detailed in (see...). Figure 2 Green represents the main active ingredient, and orange represents the main target for combating memory loss.

[0063] 2.2 Targets related to memory loss and therapeutic targets of Yiheyin Using the Genecard, DrugBank, OMIM, and TTD databases, a search was conducted using "MemoryDisorders" as the keyword. After merging and deduplication, 14,529 disease targets were obtained. (See [link to database]). Figure 3 Through intersection analysis of drug and disease targets, 559 potential targets for Yiheyin in treating memory loss were identified and visualized using a Venn diagram. (See attached image.) Figure 4 .

[0064] 2.3 Construction of the interaction network between traditional Chinese medicine, main active ingredients, and potential targets The active ingredients in Yiheyin and 559 intersection targets of memory decline were input into Cytoscape 3.10.1 software to construct a visual network relating to the traditional Chinese medicine components. Through network topology analysis, the degree values ​​of the compounds were calculated, and the top 10 compounds were selected (see Table 9).

[0065] Table 9. Information on the top 10 compounds in Yiheyin with moderate alcohol content. The results showed that quercetin, yakuchinone a, palmitoleic acid, and tectochrysin were ranked highly, indicating that these components may be the core active ingredients in Yiheyin for treating memory loss.

[0066] 2.4 Construction of PPI Network in Yiheyin for Treating Memory Loss A total of 559 potential therapeutic targets corresponding to Yihe Yin and memory impairment were imported into the STRING 11.0 database to construct a PPI network, and the results were imported into the Cytoscape 3.10.1 software for visualization. The color depth of nodes and interaction edges represents the target association strength. The closer the node color is to red, the larger the degree value, and the closer the node is to yellow, the smaller the degree value. The darker the color of the edge, the larger the degree value. Further, with the aid of the CytoNCA plug-in, core targets were screened by combining parameters such as degree, betweenness, and closeness. The results showed that targets such as STAT3, SRC, ESR1, TP53, JUN, and EGFR had relatively high degree values, as shown in Figure 5 . It is suggested that these proteins may play a key role in the process of Yihe Yin treating memory impairment.

[0067] 2.5 GO Functional and KEGG Pathway Enrichment Analysis GO enrichment analysis was performed on 559 potential targets for Yihe Yin to treat memory impairment. This analysis covered three categories: biological process (BP), molecular function (MF), and cellular component (CC). Set P <0.05, and after sorting by P value, the top 10 entries were selected to draw bar charts and bubble charts, as shown in Figure 6 . KEGG pathway analysis showed that these targets were mainly enriched in signal pathways such as calcium signaling pathway, chemical carcinogenesis - reactive oxygen species, HIF-1 signaling pathway, PI3K-Akt signaling pathway, glioma, and cellular senescence, suggesting that Yihe Yin may exert a therapeutic effect on memory impairment through the above signal pathways. According to the P value and Count value sorting, the top 30 enriched entries were selected to draw bar charts and bubble charts, as shown in Figure 7 .

[0068] 2.6 Molecular Docking Verification For molecular docking verification, according to the PPI results, the top 5 compounds were taken for molecular docking with the core targets. That is, the binding energies of quercetin, palmitoleic acid, penduletin, tectochrysin, and yakuchinone a with the core targets JUN, FOS, SRC, TP53, and ESR1 were all less than -5.0 kcal / mol, as shown in Table 10.

[0069] Table 10 Docking Binding Energy (Affinity) of Core Targets and Active Ingredients The results showed that there was a strong binding activity between the core targets and the compounds. Therefore, Yihe Drink may intervene in the related pathways of memory decline through the above core targets, achieving the effect of treating memory decline.

[0070] Example 3 Animal Experiment on the Improvement of Memory by the Plant Beverage Yihe Drink 1 Experimental Materials 1.1 Experimental Animals 60 SPF-grade male SD rats, weighing 200 - 220 g, were purchased from Beijing Spepharm Biotechnology Co., Ltd. (Production License No.: SCXK(Beijing)2024 - 0001). They were housed in the SPF animal room of Xiyuan Hospital, China Academy of Chinese Medical Sciences (Animal Breeding License No.: SYXK(Beijing)2023 - 0053). The breeding environment was suitable, with the room temperature maintained at 23 ± 2°C, humidity at 50 ± 100%,%, and the light / dark time at 12 h / 12 h. The rats had free access to food and water.

[0071] 1.2 Experimental Reagents 1.2.1 Administration to Animals Yihe Drink: Composed of Alpinia oxyphylla, Lily, and Lotus Leaf, provided by Hebei Baixiaodan Pharmaceutical Co., Ltd. (prepared by the process method of Example 1).

[0072] Donepezil Hydrochloride Tablets (also known as Aricept): Purchased from Eisai (China) Pharmaceutical Co., Ltd., National Drug Approval No. H20070181.

[0073] 1.2.2 Model Establishment Pentobarbital Sodium: Shanghai Tongshan Biotechnology Co., Ltd., Tc - P8411; Microspheres (106 - 125μm, 180 - 212μm): Cospheric Company, USA, UVPMS - BY2 - 1.00; Low - molecular - weight Dextran Amino Acid Injection: Guangdong Leiyunshang Pharmaceutical Co., Ltd., National Drug Approval No. H44025079; Heparin Sodium Injection: Changzhou Qianhong Biochemical Pharmaceutical Co., Ltd., 86901379000083.

[0074] 1.2.3 Pathological Morphology Table 11 Reagents Used for Detecting Pathological Morphology 1.2.4 Enzyme - Linked Immunosorbent Assay (ELISA) Detection Table 12 Reagents Used for ELISA Detection 1.3 Experimental Equipment Table 13 Equipment used in animal experiments Consumables required for model making and material collection: Gillette blades, straight scissors (large / small), hemostatic forceps, dental forceps, ophthalmic curved forceps, arterial clamps, needle holders, disposable sterile suture needles with sutures (4-0), disposable sterile syringes (1ml, 5ml, 10ml), 75% alcohol, povidone-iodine, physiological saline, sterile gauze, cotton balls, cotton swabs, medical gauze, masks, disposable sterile gloves, liquid nitrogen, etc.

[0075] 1.4 Experimental Methods 1.4.1 Rat Model Preparation A rat model of vascular cognitive impairment was established using microsphere embolization. Rats were fasted for 12 hours preoperatively, allowed free access to water, and anesthetized with intraperitoneal injection of sodium pentobarbital (40 mg / kg). The rats were fixed in a supine position, and a midline incision was made in the neck. The right common carotid artery (CCA), internal carotid artery (ICA), and external carotid artery (ECA) were separated. Small arteries branching from the ECA were ligated, and the distal end of the ECA was ligated. Blood flow to the CCA was temporarily blocked with an arterial clamp. A small incision was made in the ECA, and 0.2 mL of fluorescent microsphere suspension (microsphere size: 106-125 μm and 180-212 μm, dissolved in 5% low molecular weight dextran amino acid injection solution) was injected into the right ICA via the right ECA using a blunted injection needle (5 mL syringe needle). The arterial clamp was then released, and the microspheres dispersed from the right ICA into various cerebral arteries via the blood flow. Finally, the wound was sutured layer by layer with surgical sutures and disinfected with iodine. In the sham surgery group, only the neck skin was cut open and blood vessels were separated; no microspheres were injected.

[0076] 1.4.2 Neurological deficit score Twenty-four hours after the modeling was completed, the Zea-Longa method was used to assess neurological deficits. The Zea-Longa method has a 5-point scale: 0 points indicates no behavioral deficits and suggests no neurological damage; 1 point indicates adduction and flexion of the contralateral forelimb when the tail is lifted, which can be considered mild neurological damage; 2 points indicates rotation to the contralateral side when crawling, which can be considered moderate neurological damage; 3 points indicates falling to the contralateral side when standing or crawling, which can be considered severe neurological damage; and 4 points indicates no voluntary activity but impaired consciousness. All scores were taken independently by two researchers, and the average score was used.

[0077] A neurological deficit score of 1-3 indicates successful model establishment; rats with a neurological deficit score of 0 or 4 are excluded.

[0078] 1.4.3 Grouping of experimental animals and administration of drugs via gavage Fifty rats that successfully established the model were selected and randomly divided into the following five groups using a random number table: ①Sham surgery group; ② Model group; ③ Donepezil hydrochloride group, dose was 1.04 mg / kg·d; ④ The medium-dose group of Yiheyin (YHY medium) had a dose of 1.35g crude drug / kg·d; ⑤YHY high-dose group, with a dose of 2.70g crude drug / kg·d.

[0079] Each rat model group was administered the above dosage by gavage. The sham-operated group and the model group were given an equal volume of 0.9% sodium chloride solution. The administration was repeated once every 24 hours for 28 days.

[0080] 1.4.4 Behavioral Experiments 1.4.4.1 Morris Water Maze Experiment The Morris water maze experiment setup consists of a circular water tank, a moving platform, and a camera mounted on top. The circular water tank is filled with tap water, maintained at a temperature of 25 ± 1℃. The tank is divided into four equal quadrants, with the moving platform located 3 cm below the water surface in the center of one quadrant. Initially, rats are allowed to swim freely for 2 minutes to familiarize themselves with the maze environment. The orientation and navigation experiment is conducted for 5 days. The platform is placed in quadrant 1, and the rats are placed facing a curtain. The camera on top records the time it takes for the rats to find the platform (escape latency), average swimming speed, and swimming trajectory. The rats should find the platform within 90 seconds. If they do not find it within 90 seconds, the experimenter guides them to the platform, where they remain for 10 seconds, and this time is recorded as 90 seconds. For the four training sessions, rats are placed from fixed locations in different quadrants. The average escape latency of the four orientation and navigation experiments each day is used as the final result. The orientation and navigation experiment records the rats' average swimming speed, escape latency, and swimming trajectory.

[0081] On day 6 of the Morris water maze experiment, a spatial exploration test was conducted. The moving platform was removed, and the rats were placed in the water from the same position in the same quadrant and allowed to swim freely for 90 seconds. The number of times the rats crossed the platform quadrant and the time spent in the target quadrant were recorded.

[0082] 1.4.4.2 New Object Recognition Experiment The novel object recognition experimental setup consisted of a gray open box and a camera mounted on top. Tracking and data analysis were automated using an animal movement tracking system and an animal behavior video analysis system. The experiment was divided into three phases: adaptation, training, and testing. In the adaptation phase, each rat was placed in the open box empty for 5 minutes to familiarize itself with the experimental setup. In the training phase, two identical objects (A1 and A2) were placed in the instrument, and each rat had 5 minutes of free exploration time; the movement trajectory and time of exploration were recorded. Twenty-four hours after the training phase, the testing phase was conducted. Object A2 was replaced with an object B of a different shape. All rats had 5 minutes of exploration time after each object replacement, and the movement trajectory and time of exploration were recorded. We defined a discrimination index (IR = novel object exploration time / total exploration time × 100%) to assess each rat's learning and memory abilities. After each trial, the open box and objects were wiped clean with 70% ethanol to remove any odors, feces, etc., left by the previous rat.

[0083] 1.4.4.3 Tissue paraffin embedding and section preparation (1) Fixation and sampling: Under fasting conditions, rat brain tissue was anesthetized by intraperitoneal injection of sodium pentobarbital (40 mg·kg-1), and fresh rat brain tissue was dissected and fixed in tissue fixation solution for more than 24 hours.

[0084] (2) Dehydration and paraffin embedding: Rat brain tissue was placed in different reagents in sequence through the following steps: 75% alcohol for 4 hours, 85% alcohol for 2 hours, 90% alcohol for 2 hours, 95% alcohol for 1 hour, anhydrous ethanol I for 30 minutes, and anhydrous ethanol II for 30 minutes. Clearing was then performed: benzene for 5-10 minutes, xylene I for 5-10 minutes, and xylene II for 5-10 minutes. Infiltration was then performed: molten paraffin I, II, and III at 65°C for 1 hour each. The paraffin-impregnated tissue was then embedded.

[0085] (3) Paraffin sections: Coronal sections were prepared using a microtome. Each section was 4 μm thick. The sections were then transferred and laid flat on a glass slide and baked in an oven at 60°C for 2 hours.

[0086] 1.4.4.4 HE staining (1) Prepare paraffin sections of rat brain tissue, following the same steps as before.

[0087] (2) Stain with hematoxylin for 5 minutes, then rinse the slides with tap water to remove excess staining solution. This process takes about 10 minutes. Afterward, rinse the slides with distilled water for 10 seconds.

[0088] (3) Place the sections in the differentiation solution for 30 seconds. Then, rinse the sections with tap water for about 10 minutes to remove the differentiation solution. Place the sections in the blueing solution and then rinse with distilled water continuously for 10 seconds.

[0089] (4) The sections were placed in 85% and 95% graded alcohol for 5 min each for dehydration, and then stained in eosin staining solution for 5 min.

[0090] (5) Gradient dehydration and clearing: 70% alcohol, 80% alcohol, 90% alcohol, xylene I, xylene II for 5 min each.

[0091] (6) Sealing: After applying neutral resin, seal the film.

[0092] (7) Observation under light microscope: Observe the morphology of hippocampal neurons under light microscope and take pictures at the same time.

[0093] 1.4.4.5 Nissl staining (1) Dewaxing paraffin sections to water: Place the sections in xylene I and xylene II for 20 min each, 100% ethanol for 10 min, 95%, 90%, 80% and 70% alcohol for 5 min each, and rinse the sections with distilled water several times.

[0094] (2) Staining: Stain with 1% toluidine blue solution for 10 min.

[0095] (3) Differentiation: Use 0.1% glacial acetic acid for slight differentiation, and wash with tap water to terminate the reaction.

[0096] (4) Sealing: After drying in the oven, add xylene for 10 minutes, and then seal with neutral resin.

[0097] (5) Observation under light microscope: The staining results show that Nissl bodies in neurons are dark blue granules and cell nuclei are light blue. Observe the morphology and number of hippocampal neurons and take pictures at the same time.

[0098] 1.4.4.6 Enzyme-linked immunosorbent assay (ELISA) (1) Move all reagents to room temperature (18-25℃) and equilibrate for at least 30 minutes.

[0099] (2) Solution preparation: The standard solution is serially diluted, and the 30-fold concentrated washing solution is diluted 30 times with distilled water before use. (3) Sample addition: Set up blank wells (blank control wells without sample and enzyme labeling reagent, the rest of the steps are the same), standard wells, and sample wells. Add 50 μl of different concentrations of standard to the standard wells of the enzyme-labeled plate, and add 50 μl of the sample to be tested diluted 5 times to the sample wells.

[0100] (4) Incubation: After sealing the plate with sealing film, incubate at 37℃ for 30 minutes.

[0101] (5) Washing: Carefully peel off the sealing film, discard the liquid, spin dry, add 350 μl of washing solution to each well, let stand for 30 seconds, discard and spin dry. After each washing, pat dry on filter paper and repeat the washing process 5 times.

[0102] (6) In addition to the blank wells, add 50 μl of enzyme labeling reagent. Add the sample to the bottom of the well of the enzyme labeling plate, trying not to touch the well wall, and gently shake to mix.

[0103] (7) Incubation: After sealing the plate with sealing film, incubate at 37℃ for 30 minutes.

[0104] (8) Washing: Carefully peel off the sealing film, discard the liquid, spin dry, add 350 μl of washing solution to each well, let stand for 30 seconds, discard and spin dry. After each washing, pat dry on filter paper and repeat the washing process 5 times.

[0105] (9) Color development: Add 50 μl of color developer A to each well, then add 50 μl of color developer B, gently shake to mix, and develop color at 37°C in the dark for 10 min.

[0106] (10) Termination: Add 50 μl of stop solution to each well to terminate the reaction (at this time, the blue color will immediately turn yellow).

[0107] (11) Measurement: Zero the instrument with the blank well and measure the absorbance (OD value) of each well in sequence at a wavelength of 450 nm. The measurement should be performed within 15 minutes after adding the stop solution.

[0108] 1.5 Experimental Results 1.5.1 Neurological deficit score results Statistical analysis was performed on the neurological deficits in the five groups of rats 24 hours after modeling, such as... Figure 8 As shown in Table 14.

[0109] Table 14 Statistical results of neurological deficits in rats 24 hours after modeling. Note: The sham-operated group showed no neurological deficits, and all animals scored 0. The model group, donepezil hydrochloride group, and the medium and high dose groups of Yiheyin were compared. P >0.05.

[0110] The results showed that there were no statistically significant differences in neurological deficit scores among the model group, donepezil hydrochloride group, and the medium and high dose groups of Yiheyin (a traditional Chinese medicine formula). P >0.05), indicating that the degree of nerve deficit is basically the same in each group.

[0111] 1.5.2 Results of the Morris water maze experiment Statistical analysis was performed on the average swimming speed of rats in each group, such as... Figure 9 As shown in Table 15.

[0112] Table 15 Statistical results of average swimming speed of rats in each group The results showed no statistically significant difference in swimming speed among the groups of rats. P (>0.05), all rats in each group had normal motor ability and survival motivation.

[0113] Analysis of the escape latency time in 1-5 day positioning navigation experiments revealed that as the training time for positioning navigation experiments increased, such as... Figure 10 As shown in Table 16.

[0114] Table 16 Statistical results of escape latency in the rat positioning and navigation experiment Note: a This indicates that compared to the sham surgery group, P <0.05; b This indicates that compared to the sham surgery group, P <0.01; c This indicates a comparison with the model group. P <0.05; d This indicates a comparison with the model group. P <0.01.

[0115] The escape latency time of all five groups of rats decreased, and the overall trend was downward.

[0116] On days 1 and 2 of the orientation navigation experiment, compared with the sham-operated group, the escape latency of rats in the model group was significantly prolonged. P <0.05, P <0.01); Compared with the model group, there was no significant difference in the escape latency time between the donepezil hydrochloride group and the medium and high dose groups of Yiheyin (<0.01); P >0.05). During days 3-5 of the orientation navigation experiment, compared to the sham-operated group, the escape latency of rats in the model group was significantly prolonged ( P <0.01); During the positioning and navigation experiment on days 3-5, compared with the model group, the donepezil hydrochloride group ( P <0.01, P <0.05, P <0.05), Yiheyin medium-dose group ( P <0.01, P <0.05, P <0.01), Yiheyin high-dose group ( P <0.01, P <0.05, P <0.01) The duration of the escape incubation period was significantly reduced.

[0117] In the space exploration experiment on day 6, such as Figure 11 and Figure 12 As shown in the figure, and in Table 17.

[0118] Table 17 Statistical results of the number of times the rat navigation platform was traversed and the time spent in the target quadrant. Note: a This indicates that compared to the sham surgery group, P <0.05; b This indicates that compared to the sham surgery group, P <0.01; c This indicates a comparison with the model group. P <0.05; d This indicates a comparison with the model group. P <0.01.

[0119] The results showed that, compared with the sham-operated group, the number of platform crossings and the dwell time in the target quadrant were significantly reduced in the model group rats. P <0.01, P <0.05; compared with the model group, the number of platform crossings in the medium-dose group of Yiheyin was significantly increased ( P <0.01, the target quadrant dwell time in both the medium-dose and high-dose groups of Yiheyin was significantly increased ( P <0.05).

[0120] The results of this experiment show that the spatial memory ability of rats was severely impaired after modeling. Both donepezil hydrochloride and Yiheyin (medium and high doses) can significantly improve the spatial memory ability of rats. Among them, Yiheyin showed better performance in improving the escape latency, platform crossing number and dwell time in the target quadrant of rats.

[0121] 1.5.3 Experimental Results of New Object Recognition like Figure 13 As shown, during the training phase, the discrimination index of the five groups of rats for two identical objects was approximately 50%, and there was no statistically significant difference between the groups. P >0.05), indicating that the rats in each group showed no selective preference for the location or environment of the two identical objects. Figure 14 As shown in Table 18.

[0122] Table 18 Statistical results of discrimination index in the training and testing phases of the rat novel object recognition experiment. Note: a This indicates that compared to the sham surgery group, P <0.05; c This indicates a comparison with the model group. P <0.05.

[0123] The results showed that, during the novel object recognition test phase, compared with the sham-operated group rats, the model group rats exhibited a decreased ability to distinguish novel objects. P <0.05. Compared with the model group, the recognition index of new objects in rats in the medium-dose and high-dose groups of Yiheyin was increased after administration ( P <0.05).

[0124] The results of this experiment show that the recognition and memory abilities of rats were severely reduced after modeling, and Yiheyin (medium and high doses) could improve the recognition and memory abilities of rats to a certain extent.

[0125] 1.5.4 HE staining results HE staining results are as follows Figure 15 and Figure 16 And as shown in Table 19.

[0126] Table 19. Statistical results of neuronal number in the CA1 region of the rat hippocampus. Note: b This indicates that compared to the sham surgery group, P <0.01; c This indicates a comparison with the model group. P <0.05.

[0127] The results showed that in the sham-operated group, hippocampal neurons in the rats had normal morphology and intact structure, with few neurons showing obvious necrosis or degeneration; the neurons were regularly arranged, densely structured, and without widening of intercellular spaces. Compared with the sham-operated group, the number of neurons in the CA1 region of the hippocampus in the model group was significantly reduced ( P <0.01), abnormal neuronal cell morphology, condensed nuclei, and darkened staining; disordered neuronal arrangement, loose structure, and widened intercellular spaces; compared with the model group, the donepezil hydrochloride group showed an increase in the number of neurons in the CA1 region ( P <0.05%, a small number of condensed nuclei were observed with darker staining; neurons were arranged relatively neatly, with a compact structure and reduced intercellular spaces. Compared with the model group, there was no statistically significant difference in the number of neurons in the CA1 region in the medium-dose group of Yiheyin (<0.05). P >0.05), some nuclei showed condensation and darkened staining; neuronal arrangement remained relatively disordered. The high-dose group of Yiheyin showed an increase in the number of neurons in the CA1 region ( P <0.05), a small number of condensed nuclei were observed, with darker staining; neurons were arranged relatively neatly, with a compact structure and reduced intercellular spaces.

[0128] The results of this experiment show that the number of neurons in the CA1 region of the hippocampus of cognitively impaired rats is significantly reduced, with abnormal cell morphology, pyknosis of the nuclei, and darker staining; neurons are disordered in arrangement, loose in structure, and widened in intercellular spaces. However, intervention with donepezil hydrochloride and Yiheyin (especially high doses) can significantly increase the number of neurons in the CA1 region and repair the morphology, structure, and arrangement of neurons.

[0129] 1.5.5 Nissl staining results Nissl staining results as follows Figure 16 and Figure 17 As shown in the figure and Table 20.

[0130] Table 20 Statistical results of Nissl bodies in the CA1 region of the rat hippocampus Note: a This indicates that compared to the sham surgery group, P <0.05; c This indicates a comparison with the model group. P <0.05.

[0131] The results showed that in the sham-operated group, the neurons in the CA1 region of the hippocampus were arranged more neatly and densely, with deeper cytoplasmic staining and a greater number of Nissl bodies, appearing dark blue. Compared with the sham-operated group, the hippocampal neurons in the model group showed necrosis or deformation, and a reduced number of Nissl bodies. P <0.05). Compared with the model group, the donepezil hydrochloride group showed more orderly neuronal arrangement and an increased number of Nissl bodies ( P <0.05. In the medium-dose group of Yiheyin, neuronal necrosis or degeneration was observed, with disordered neuronal arrangement and loose structure; the number of Nissl bodies showed no statistically significant difference. P >0.05). Although the high-dose group of Yiheyin showed disordered neuronal arrangement, loose structure, and increased intercellular spaces, the number of Nissl bodies increased. P <0.05).

[0132] The results of this experiment indicate that the number of Nissl bodies in the CA1 region of the hippocampus of rats decreased after modeling. However, intervention with donepezil hydrochloride and Yiheyin (especially at high doses) significantly increased the number of Nissl bodies in the CA1 region.

[0133] 1.5.6 ELISA staining results The expression levels of neurotrophic factor (BDNF), nerve growth factor (NGF), neurotrophic factor-3 (NT-3), and neurotrophic factor-4 (NT-4) in brain tissue were detected using an ELISA kit. The results are as follows: Figure 18 And as shown in Table 21.

[0134] Table 21. Expression levels of different proteins in rat hippocampus (ng / g protein) Note: a This indicates that compared to the sham surgery group, P <0.05; b This indicates that compared to the sham surgery group, P <0.01; c This indicates a comparison with the model group. P <0.05; d This indicates a comparison with the model group. P <0.01.

[0135] Compared with the sham-operated group, the levels of BDNF, NGF, and NT-4 in the hippocampus of rats in the model group were significantly reduced. P <0.01, P <0.05, P <0.05. Compared with the model group, the levels of BDNF and NGF in the hippocampus of rats in the donepezil hydrochloride group were significantly increased ( P <0.05, P <0.01, the levels of BDNF, NGF, NT-3, and NT-4 in the hippocampus of rats in the medium-dose group of Yiheyin were significantly increased ( P <0.01, P <0.01, P <0.05, P <0.05. Compared with the model group, there was no statistically significant difference in the levels of BDNF, NGF, NT-3, and NT-4 in the hippocampus of rats in the high-dose Yiheyin group ( P >0.05).

[0136] The results of this experiment show that medium doses of Yiheyin can significantly increase the levels of BDNF, NGF, NT-3 and NT-4 in the hippocampus of rats, indicating that it may exert a neuroprotective effect by upregulating the expression of multiple neurotrophic factors.

[0137] Example 4: Human Trial Experiment 1. Overall Design This was a clinical exploratory study, a randomized, double-blind, placebo-controlled trial. There were 60 patients in the intervention group (n=30) and the control group (n=30).

[0138] This study employed a blinding method (researchers and subjects). ① Blinding: Randomization was conducted using a randomized coding table generated by SPSS 27.0 software with a given seed number. Assignments were performed by designated personnel not directly involved in the trial, and a blinding record was created and maintained. ② Emergency Letters: Paper emergency letters were used, one for each subject. Each letter contained a sealed oral solution corresponding to the random number, and was sent to the research center along with the corresponding oral solution. ③ Emergency Unblinding: In emergency situations, if researchers believed that knowing the oral solution taken by the subject would be beneficial for managing adverse events, they could open the emergency letter for emergency unblinding. The researcher opening the emergency letter should also note the reason for the emergency unblinding, the date, and sign it. ④ Unblinding Rules: Unblinding in this study was conducted after the statistical plan, data review report (or blinding review report) were finalized and the database was locked. The oral solution corresponding to the random number was revealed to facilitate statistical analysis of all data after grouping. The unblinding document was jointly signed by the principal investigator, the responsible party, and the statistician.

[0139] 2. Selection of subjects From December 2025 to April 2026, 60 patients with memory impairment were recruited at Xiyuan Hospital of China Academy of Chinese Medical Sciences. This study was approved by the Medical Ethics Committee of Xiyuan Hospital of China Academy of Chinese Medical Sciences (Approval No.: 2025XLA152-2) and has been registered on the International Traditional Medicine Clinical Trial Registry Platform (Registration No.: ITMCTR2025002587).

[0140] 2.1 Inclusion Criteria (1) Age between 18 and 60 years old; (2) The chief complaint is subjective memory loss, and the memory loss symptoms have lasted for more than 6 months; (3) The cutoff values ​​for the Montreal Cognitive Assessment-Basic (MoCA-B) are: less than 6 years of education, ≤19 points; 7-12 years of education, ≤22 points; more than 12 years of education, ≤24 points; (4) No regular use of traditional Chinese and Western medicine for memory loss in the past 30 days, and no use of anti-anxiety, antidepressant, sedative and hypnotic Chinese and Western medicines; (5) The subject was informed and signed an informed consent form.

[0141] 2.2 Exclusion Criteria (1) Imaging examinations showed obvious organic pathological damage to the brain; (2) Other neurological diseases that may affect cognitive function (such as Alzheimer's disease, frontotemporal dementia, severe Parkinson's disease, etc.) or other diseases that may affect cognitive function (such as hypothyroidism, severe anemia, syphilis, etc.); those with severe neurological deficits who cannot complete the examination (such as hemiplegia, various aphasias, visual and auditory impairments, etc.); severe mental illnesses (such as depression, schizophrenia, severe anxiety) and epilepsy; those whose cognitive function is affected by drugs or poisoning; (3) Alcohol and drug abusers; (4) Subjects with allergic constitution or allergic to Yiheyin oral liquid; (5) Abnormal laboratory parameters: Liver and kidney function indicators exceed 1.5 times the upper limit of normal; (6) Individuals currently participating in other clinical trials that may affect the evaluation of the results of this study; (7) Pregnant women (a pregnancy test is required, and those with a positive pregnancy result are excluded).

[0142] 2.3 Termination / Withdrawal of the Test The study is terminated if a participant, already enrolled, develops conditions during the trial that make them unfit to continue, preventing them from completing the trial according to the protocol. Termination measures should generally be taken in the following situations: (1) The subject experienced severe discomfort during the blood draw process, such as severe fainting of blood or fainting of needles.

[0143] (2) If a subject’s condition worsens or deteriorates rapidly during the trial and he is unable to cooperate with the scale assessment, the clinical trial should be stopped according to the doctor’s judgment.

[0144] (3) During the trial, the subject developed certain comorbidities, complications or special physiological changes, making it unsuitable to continue the trial.

[0145] (4) If a subject experiences a serious adverse reaction during the study, the study should be terminated; (5) If, during the study, the subjects fail to follow the clinical trial protocol in terms of the use of Yiheyin oral liquid, receiving visits, and follow-ups, and the researchers determine that this may affect the authenticity of the study results, the study should be terminated.

[0146] (6) External information (such as other high-quality studies or evidence) proves that the protocol is ineffective or meaningless, and there is no need to continue the current clinical trial.

[0147] 3. Intervention measures 3.1 Test drug and dosage Yiheyin Group (Experimental Group): Take Yiheyin oral liquid, 2 times a day, 1 sachet each time.

[0148] Placebo group (control group): Administered placebo, 1 sachet twice daily.

[0149] The treatment course lasts for 30 days, with a follow-up period of 1 month.

[0150] Yiheyin Oral Solution (Composition: Alpinia oxyphylla 5g, Lilium brownii 5g, Nelumbo nucifera leaf 3g; Specification: 30ml × 30 sachets). The placebo is made by diluting the original Yiheyin Oral Solution with purified water to a concentration of 5% (i.e., containing 1 / 20 of the original solution). Both are basically identical in terms of outer packaging, appearance, and color, and have the same usage and dosage. Both are produced by Hebei Baixiaodan Pharmaceutical Co., Ltd.

[0151] 3.2 Regulations for Combined Medication Use When researchers become aware that a participant has comorbidities requiring treatment, they inform the participant and pay attention to any concomitant medications that may interfere with clinical trial results or participant safety. Concomitant medications include long-term oral medications for chronic diseases such as hypertension, diabetes, coronary heart disease, and hyperlipidemia.

[0152] During the study, the use of other traditional Chinese medicines with cognitive-improving effects is prohibited, including oral preparations (such as Ginkgo biloba tablets, Compound Cistanche deserticola capsules), decoctions (such as Angelica sinensis and Paeonia lactiflora powder, Gui Pi Tang, etc.), injections (such as Shu Xue Ning injection), and traditional Chinese medicine teas (such as Polygala tenuifolia tea). The use of other Western medicines with cognitive-improving effects is also prohibited, including cholinesterase inhibitors (such as donepezil, rivastigmine, galantamine), glutamate receptor blockers (such as memantine), antihyperxia drugs (such as lamotrigine), brain metabolism activators (such as piracetam, piracetam, piracetam), microcirculation-improving drugs (such as ergot alkaloid preparations), and calcium channel blockers (such as nimodipine). The use of unmarketed drugs or other drugs used in clinical trials is also prohibited during the study.

[0153] 4. Observation Indicators and Evaluation Criteria 4.1 General Information The subjects' personal information includes name, gender, date of birth, ethnicity, marital status, occupation, and education level.

[0154] 4.2 Observation Indicators Changes in subjects' scores on the Clinical Memory Scale (compiled by the Institute of Psychology, Chinese Academy of Sciences) on day 1 and day 30 after enrollment; changes in memory quotient, Montreal Cognitive Assessment-B (MoCA-B), Psychological Resilience Scale (CD-RISC), Fatigue Rating Scale (FAS), Hamilton Anxiety Scale (HAMA), and Traditional Chinese Medicine Core Symptom Observation Scale.

[0155] 4.3 Security (1) Vital signs: body temperature, blood pressure, respiration, heart rate, etc. on day 1 and day 30 after enrollment; (2) Laboratory tests such as routine blood tests, liver and kidney function tests, routine stool tests, and routine urine tests on day 1 and day 30 after enrollment, as well as examinations such as electrocardiogram and abdominal ultrasound. (3) Adverse reactions and adverse events during treatment and follow-up.

[0156] 4.4 Statistical Analysis Statistical analysis was performed using SPSS 27.0 software. Normally distributed continuous data were expressed as mean ± standard deviation. Paired-samples t-tests were used for within-group comparisons, and independent-samples t-tests were used for between-group comparisons. Data not conforming to a normal distribution were expressed as median (upper and lower quartiles) [M( P 25, P 75) indicates that within-group comparisons were performed using the signed-rank test, and between-group comparisons were performed using the Wilcoxon rank-sum test. Categorical data were described as frequencies or percentages and analyzed using the χ² test. P <0.05 indicates a statistically significant difference.

[0157] Categorical variables are expressed as number of cases (component ratio / percentage), and Fisher's exact test is used for comparisons between groups. When the contingency table has excessively small expected frequencies (e.g., n < 40 or expected frequency T < 5), the chi-square test is not applicable; therefore, Fisher's exact test is used for correction analysis. P <0.05 indicates a statistically significant difference.

[0158] 4.5 Results 4.5.1 Baseline Comparability This study included 66 patients. Four patients dropped out of the Yiheyin group (lost to follow-up), and two dropped out of the placebo group (one withdrew voluntarily, one lost to follow-up). Ultimately, 60 patients were enrolled: 30 in the Yiheyin group and 30 in the placebo group.

[0159] There were no statistically significant differences in gender, age, and years of education between the two groups of patients. P (>0.05), indicating comparability. See Table 22.

[0160] Table 22 Baseline data of patients [M ( P 25, P 75) 4.5.2 Comparison of memory quotient scores before and after treatment between the two groups of patients There was no statistically significant difference in memory quotient scores between the two groups of patients before treatment. t = -0.407, P = 0.685>0.05, indicating comparability. Compared with pre-treatment levels, the memory quotient score of patients in the Yiheyin group increased after treatment ( t = -7.482,P <0.001); Compared with pre-treatment levels, the placebo group showed an increase in memory quotient scores after treatment ( t = -5.492, P <0.001); Compared with the placebo group, the memory quotient score of patients in the Yiheyin group increased after treatment ( t = -2.066, P = 0.043 < 0.05). See Table 23.

[0161] Table 23 Comparison of memory quotient scores before and after treatment in the two groups of subjects with memory decline (scores, x̄ ± s) 4.5.3 Comparison of MoCA scores before and after treatment in the two groups of patients There was no statistically significant difference in MoCA scores between the two groups before treatment (z = -0.515). P = 0.606>0.05), indicating comparability. Compared with pre-treatment levels in this group, the MoCA score of patients in the Yiheyin group increased after treatment (z = -4.639, P <0.001); Compared with pre-treatment levels, the MoCA score of patients in the placebo group increased after treatment (z = -4.173, P <0.001); Compared with the placebo group, the Yiheyin group showed a more significant improvement in MoCA scores after treatment (z = -3.344, P <0.001). See Table 24.

[0162] Table 24 Comparison of MoCA scores before and after treatment in two groups of subjects with memory impairment [points, M ( P 25, P 75) 4.5.4 Comparison of HAMA scores before and after treatment in the two groups of patients There was no statistically significant difference in HAMA scores between the two groups before treatment. t = 0.336, P = 0.715>0.05), indicating comparability. Compared with pre-treatment levels, the HAMA score of the Yiheyin group decreased after treatment (z = -3.994, P <0.001); Compared with pre-treatment levels, the placebo group showed a decrease in HAMA score after treatment (z = -1.847, P = 0.065>0.05); compared with the placebo group, the Yiheyin group showed more significant improvement in HAMA scores after treatment ( t = 3.523, P<0.001). See Table 25.

[0163] Table 25 Comparison of HAMA scores before and after treatment in two groups of subjects with memory impairment (scores, x̄ ± s) 4.5.5 Comparison of CD-RISC scores before and after treatment in the two groups of patients There was no statistically significant difference in CD-RISC scores between the two groups before treatment. t = -1.604, P = 0.114>0.05), indicating comparability. Compared with pre-treatment levels, the CD-RISC score of patients in the Yiheyin group increased after treatment (z = -3.324, P <0.001); There was no statistically significant difference in CD-RISC scores before and after treatment in the placebo group (z = -1.675, P =0.094>0.05); compared with the placebo group, the Yiheyin group showed a more significant improvement in CD-RISC scores after treatment (z = -3.425, P <0.001). See Table 26.

[0164] Table 26 Comparison of CD-RISC scores before and after treatment in the two groups of subjects with memory impairment [scores, x̄ ± s, M ( ) ] P 25, P 75) 4.5.6 Comparison of FAS scores before and after treatment between the two groups of patients There was no statistically significant difference in FAS scores between the two groups before treatment (z = -1.431). P = 0.153>0.05, indicating comparability. Compared with pre-treatment levels, the FAS score of patients in the Yiheyin group decreased after treatment ( t = 2.947, P = 0.006<0.05; There was no statistically significant difference in FAS scores before and after treatment in the placebo group ( t = 1.665, P = 0.107>0.05); compared with the placebo group, the Yiheyin group showed a more significant improvement in FAS scores after treatment (z = -2.003, P = 0.045 < 0.05). See Table 27.

[0165] Table 27 Comparison of FAS scores before and after treatment in two groups of subjects with memory impairment [points, M ( P 25 , P 75)] 4.5.7 Comparison of TCM core symptom observation scale scores before and after treatment between the two groups of patients Compared with the placebo group, the Yiheyin group showed a higher rate of symptom disappearance after treatment, including forgetfulness, absent-mindedness, listlessness, apathy, depression, fatigue, lethargy, drowsiness, slow reaction, and slow thinking. P <0.05). See Table 28.

[0166] Table 28 Comparison of the disappearance rate of TCM core symptoms before and after treatment in two groups of subjects with memory loss [% (n / N)] 4.5.8 Security Safety checks were performed on both groups of subjects before and after treatment, including general vital signs, routine blood, urine, and stool tests, liver and kidney function tests, electrocardiograms, and abdominal ultrasounds. All laboratory indicators showed no significant abnormalities. No adverse reactions occurred in any of the patients during the treatment period.

[0167] This article uses specific examples to illustrate the inventive concept in detail. The description of the above embodiments is only for the purpose of helping to understand the core idea of ​​the present invention. It should be noted that any obvious modifications, equivalent substitutions or other improvements made by those skilled in the art without departing from the inventive concept should be included within the protection scope of the present invention.

Claims

1. A food-medicine homology product for improving memory, characterized in that, It contains Chinese medicinal materials, which are composed of Alpinia oxyphylla, lily bulb, and lotus leaf, or the Chinese medicinal materials are composed of Alpinia oxyphylla extract, lily bulb extract, and lotus leaf extract.

2. The food-medicine homology product according to claim 1, characterized in that, The dosage of Alpinia oxyphylla, lily bulb, and lotus leaf is calculated by weight as follows: 5 parts Alpinia oxyphylla, 5 parts lily bulb, and 3 parts lotus leaf.

3. The application of a food-medicine homology product in the preparation of memory-improving products, characterized in that, The food-medicine homology product contains Chinese medicinal materials, which are composed of Alpinia oxyphylla, lily bulb, and lotus leaf, or the Chinese medicinal materials are composed of Alpinia oxyphylla extract, lily bulb extract, and lotus leaf extract.

4. The application according to claim 3, characterized in that, The dosage of Alpinia oxyphylla, lily bulb, and lotus leaf is calculated by weight as follows: 5 parts Alpinia oxyphylla, 5 parts lily bulb, and 3 parts lotus leaf.

5. The application according to claim 3, characterized in that, The memory-improving product is a plant-based beverage, which is prepared using the following method: (1) Powdered and mixed Alpinia oxyphylla, Lilium brownii and Nelumbo nucifera leaves were extracted in two stages. In the first stage of extraction, 7 times the volume of 50% ethanol aqueous solution was added and extracted at room temperature for 1 hour. The filtrate was collected. The residue was subjected to the second stage of extraction. 6 times the volume of 50% ethanol aqueous solution was added and extracted at room temperature for 1 hour. The filtrate was collected from the two stages of extraction. The filtrates were combined and concentrated at 60-70℃ to a density of 1.10-1.15 g / mL to obtain concentrated extract. (2) Calculated per gram of concentrated extract, add 0.14g of water-soluble mono- and diglycerides of fatty acids, 0.02g of soybean lecithin and 0.200g of γ-cyclodextrin respectively; after mixing evenly, finally pass through a 400-mesh sieve, add purified water, heat to 80℃ and stir to dissolve for 30 minutes, cool to room temperature, pass the liquid through a 400-mesh sieve again before filling, fill at 30 mL per bag, sterilize and fill to obtain the plant beverage.

Images