Application of bamboo-leaf-green wine in preparation of anti-aging product

Abstract

The present application relates to the technical field of biological medicine, and particularly relates to application of bamboo-leaf-green wine in preparation of anti-aging products. The anti-aging products are prepared from bamboo-leaf-green wine as raw material through dealcoholization and drying, and can better retain active ingredients. The extract has the effects of antioxidation and nerve protection, can efficiently scavenge free radicals, improve the survival rate of oxidatively damaged neurons, reduce the accumulation of intracellular reactive oxygen species, up-regulate the expression of antioxidant enzymes, inhibit the secretion of pro-inflammatory factors, and improve the cell cycle arrest of neurons caused by oxidative stress. Animal experiments show that the extract can improve the physiological state of aging mice, improve the spatial learning and memory ability, protect brain neurons, and inhibit abnormal activation of glial cells; meanwhile, the extract can improve the antioxidant capacity of the body and inhibit central nervous inflammation. The extract breaks the vicious cycle of oxidative stress and neural inflammation through the synergistic effect of the antioxidant pathway and the anti-inflammatory pathway, and plays an anti-aging role.

Description

Technical Field

[0001] This invention relates to the field of biomedicine, and in particular to the application of Zhuyeqing wine in the preparation of anti-aging products. Background Technology

[0002] As the global population ages, age-related neurodegenerative diseases, such as Alzheimer's, Parkinson's, and frontotemporal dementia, have become prominent issues affecting the physical and mental health of middle-aged and elderly people. Aging is a complex biological process driven by multiple factors, including genetics and environment, with its core characteristic being the progressive decline in the physiological functions of various organ systems. In the central nervous system, oxidative stress and neuroinflammation are the two core intrinsic mechanisms driving age-related pathological changes and inducing neurodegenerative diseases. Excessive accumulation of reactive oxygen species (ROS) is the core manifestation of oxidative stress. Excessive ROS can induce lipid peroxidation, protein denaturation, and DNA oxidative damage, directly disrupting the integrity of neuronal structure and function. Simultaneously, ROS can damage the mitochondrial respiratory chain, causing energy metabolism disorders and further triggering the neuronal apoptosis cascade, which is an important pathological basis for cognitive decline and reduced learning and memory abilities. In addition, with the aging process, the central nervous system spontaneously produces chronic, low-grade neuroinflammation, mainly manifested as excessive activation of astrocytes and microglia. Activated glial cells can release large amounts of pro-inflammatory factors, which not only directly cause toxic damage to neurons, but also further promote ROS generation by activating inflammatory signaling pathways, forming a vicious cycle with oxidative stress and jointly accelerating the occurrence and development of neurodegenerative diseases.

[0003] Currently, drugs used clinically to delay aging and improve neurodegenerative diseases are mainly chemically synthesized. However, these drugs generally suffer from problems such as single-target effects, significant toxic side effects, and poor long-term safety. They cannot systematically intervene in complex pathological regulatory networks, greatly limiting their application in the long-term prevention and control of aging and neurodegenerative diseases. Therefore, developing natural medicinal and edible active ingredients that combine antioxidant, anti-neuroinflammatory, and multi-target synergistic regulatory advantages, while being safe, low in toxicity, and suitable for long-term use, has become a research hotspot and urgent need in this field.

[0004] Zhuyeqing wine is a traditional Chinese health-preserving beverage with a long history. It uses high-quality Fenjiu liquor as its base, combined with 12 authentic Chinese medicinal herbs including bamboo leaves, gardenia, angelica, dried tangerine peel, chrysanthemum, and cardamom. It is refined using a traditional intangible cultural heritage soaking process and was approved as a health food in 1998. Currently, systematic and in-depth research on the anti-aging and neuroprotective effects of Zhuyeqing wine is still relatively scarce. In particular, there are significant research gaps in core issues such as the molecular mechanisms of oxidative stress and neuroinflammation in regulating the aging process and improving learning and memory functions. This severely restricts the in-depth development and industrial application of Zhuyeqing wine in the fields of anti-aging functional foods and the broader health industry. Summary of the Invention

[0005] This invention aims to at least solve one of the technical problems existing in related technologies. Therefore, the object of this invention is to provide the application of Zhuyeqing wine in the preparation of anti-aging products.

[0006] To achieve the above objectives, the technical solution adopted by the present invention is as follows: The application of Zhuyeqing wine in the preparation of anti-aging products, wherein the anti-aging products include an extract of Zhuyeqing wine, the extract being obtained by using Zhuyeqing wine as raw material after de-alcoholization and drying.

[0007] Furthermore, the anti-aging product is used to prevent or treat at least one of the following age-related diseases: oxidative stress damage, neuroinflammation, cognitive impairment, and neurodegenerative diseases.

[0008] Furthermore, the anti-aging product has a neuroprotective effect.

[0009] Furthermore, the anti-aging product exerts a neuroprotective effect through anti-oxidation and / or anti-inflammation.

[0010] Furthermore, the anti-aging product enhances the body's antioxidant capacity by scavenging free radicals, including at least one of ABTS free radicals, DPPH free radicals, and superoxide anion free radicals; ABTS is 2,2'-bis(3-ethylbenzothiazoline-6-sulfonic acid) diammonium salt, and DPPH is 1,1-diphenyl-2-trinitrophenylhydrazine.

[0011] Furthermore, the anti-aging product can reduce abnormally elevated malondialdehyde levels by at least one of the following antioxidant mechanisms: restoring the activity of superoxide dismutase, glutathione, and glutathione peroxidase in the body.

[0012] Furthermore, the anti-aging product combats inflammation by inhibiting the mRNA expression of at least one of the following pro-inflammatory cytokines: tumor necrosis factor-α, interleukin-1β, and interleukin-6.

[0013] Furthermore, the anti-aging product can improve the body's spatial learning and memory abilities.

[0014] Furthermore, the anti-inflammatory effect includes anti-central nervous system inflammation; The anti-aging product inhibits the excessive activation of astrocytes and microglia in brain tissue, downregulates the mRNA expression of NF-κB, TNF-α, and IL-1β in brain tissue, and blocks the central nervous system inflammatory cascade, thereby achieving anti-central nervous system inflammation.

[0015] Furthermore, the anti-aging product includes at least one of anti-aging drugs and anti-aging foods.

[0016] The above-described one or more technical solutions in the embodiments of the present invention have at least one of the following technical effects: The present invention provides the application of bamboo leaf green wine in the preparation of anti-aging products. The anti-aging products include extracts of bamboo leaf green wine, which are obtained by de-alcoholizing and drying bamboo leaf green wine as raw material. The preparation process is simple and green, and the active ingredients are well retained.

[0017] The extract of Zhuyeqing wine exhibits excellent in vitro antioxidant capacity and neuroprotective effects. At a concentration of 5 mg / mL, its total antioxidant capacity is 3.84 μmol / mL, and its scavenging rates of ABTS, DPPH, and superoxide anion free radicals are 81.8%, 42.4%, and 14.3%, respectively, demonstrating a significant free radical scavenging effect. Cell experiments showed that this extract can significantly improve the cell survival rate of HT-22 neurons in a hydrogen peroxide (H2O2) injury model and reduce the accumulation of intracellular reactive oxygen species (ROS). Simultaneously, it can upregulate the gene expression of antioxidant enzymes CAT and SOD, inhibit the secretion of pro-inflammatory factors IL-1β and IL-6, and improve neuronal cell cycle arrest induced by oxidative stress. This product achieves effective neuronal protection through its dual antioxidant and anti-inflammatory effects.

[0018] Extracts from Zhuyeqing wine exhibit significant in vivo anti-aging and cognitive function improvement effects. They effectively alleviated the slow weight gain and reduced food intake observed in D-galactose-induced aging mice, improved age-related declines in liver and spleen indices, and restored the body to its normal physiological state. Regarding cognitive function, Zhuyeqing wine extract significantly enhanced spatial learning and memory abilities in aging mice: in the Morris water maze test, it significantly shortened escape latency, prolonged time spent in the target quadrant, and increased the number of platform crossings. Furthermore, Zhuyeqing wine extract maintained the morphological integrity of neurons in the CA1 and DG regions of the hippocampus in mice, reduced neuronal shrinkage and degeneration, and inhibited the excessive activation of astrocytes and microglia in the brain tissue of D-galactose-induced aging mice.

[0019] The extract of Zhuyeqing wine possesses clear in vivo antioxidant and anti-neuroinflammatory effects. This extract can significantly restore the activity of antioxidant enzymes SOD, GSH, and GSH-PX in the serum of aging mice, reduce the content of lipid peroxidation product MDA, and restore the balance of the body's antioxidant defense system. Simultaneously, it can significantly inhibit the excessive activation of GFAP-positive astrocytes and Iba1-positive microglia in the hippocampus of aging mice, downregulate the mRNA expression of NF-κB, TNF-α, and IL-1β in brain tissue, and effectively block the central nervous system inflammatory cascade.

[0020] The extract of Zhuyeqing wine exerts anti-aging and neuroprotective effects through a dual-pathway synergy: on the one hand, it activates the SIRT1-Nrf2-HO-1 antioxidant signaling pathway, upregulates the expression of key molecules in the pathway, and enhances the body's endogenous antioxidant capacity; on the other hand, it inhibits the NF-κB-mediated inflammatory signaling pathway, reduces the transcription and release of pro-inflammatory factors, and fundamentally blocks the vicious cycle of oxidative stress and neuroinflammation, thereby achieving systemic aging intervention.

[0021] Additional aspects and advantages of the invention will be set forth in part in the description which follows, and in part will be obvious from the description, or may be learned by practice of the invention. Attached Figure Description

[0022] Figure 1 This is a bar chart comparing the in vitro antioxidant activity of the extract of Zhuyeqing wine provided in Example 2 of the present invention.

[0023] Figure 2 This describes the effect of the extract of bamboo leaf green wine provided in Example 2 of the present invention on the activity of HT-22 cells.

[0024] Figure 3 The figures shown are the H2O2 cytotoxicity experiment results provided in Example 3 of this invention; where Figure A shows the cell survival rate at different H2O2 concentrations, and Figure B shows the dose-response curve fitting graph.

[0025] Figure 4 This describes the effects of different groups provided in Example 3 of the present invention on the H2O2-induced viability of HT-22 cells.

[0026] Figure 5 These are the results of the detection of intracellular reactive oxygen species (ROS) levels in different groups provided in Example 3 of the present invention.

[0027] Figure 6 This is a stacked bar chart of different groups of cell cycle distribution provided in Embodiment 3 of the present invention.

[0028] Figure 7 These are the RT-qPCR gene expression detection results of different groups provided in Example 3 of the present invention.

[0029] Figure 8 The results of physiological index detection of different groups of mice provided in Example 4 of the present invention are shown in Figure A, which is a curve of body weight change over time; Figure B is a bar chart of the reduction changes of food intake in different groups of mice; Figure C is a bar chart of spleen index in different groups of mice; and Figure D is a bar chart of liver index in different groups of mice.

[0030] Figure 9 The figures shown are the results of the Morris water maze experiment provided in Embodiment 4 of the present invention; wherein, Figure A is the result of the escape latency of the mouse positioning and navigation experiment, Figure B is a bar chart of the time the mouse stayed in the target quadrant, Figure C is a bar chart of the number of times the mouse crossed the platform, and Figure D is a representative swimming trajectory of the mouse.

[0031] Figure 10 The results are the serum antioxidant index detection results provided in Embodiment 4 of the present invention; wherein, Figure A is the serum SOD detection result, Figure B is the serum MDA detection result, Figure C is the serum GSH detection result, and Figure D is the serum GSH-PX detection result.

[0032] Figure 11 The images show pathological sections and immunofluorescence detection results of mouse hippocampal tissue provided in Example 4 of this invention; where Figure A is an H&E staining image and Figure B is an immunofluorescence staining image.

[0033] Figure 12 This describes the mRNA expression of SIRT1, Nrf2, HO-1, NF-κB, TNF-α, and IL-1β in mouse brain tissue provided in Example 4 of this invention. Detailed Implementation

[0034] To make the objectives, technical solutions, and advantages of this invention clearer, the technical solutions of this invention will be clearly and completely described below in conjunction with specific embodiments. Obviously, the described embodiments are only some embodiments of this invention, not all embodiments. Based on the embodiments of this invention, all other embodiments obtained by those skilled in the art without creative effort are within the scope of protection of this invention. The following embodiments are used to illustrate this invention, but cannot be used to limit the scope of this invention.

[0035] In the following embodiments, unless otherwise specified, the experimental methods used are conventional methods, and the materials and reagents used are commercially available, unless otherwise specified, and are carried out in accordance with the techniques or conditions described in the literature in this field or in accordance with the product instructions.

[0036] Example 1 I. Preparation of Zhuyeqing Wine.

[0037] Zhuyeqing liquor uses light-aroma baijiu as its base and is made by extracting 12 kinds of herbal ingredients in a specific ratio, as follows: Weigh the 12 herbal ingredients according to the following proportions and mix them evenly: Lophatherum gracile (10-50 parts), Gardenia jasminoides (10-50 parts), Citrus reticulata peel (10-50 parts), Chrysanthemum morifolium (10-50 parts), Kaempferia galanga (2-10 parts), Amomum villosum (2-10 parts), Angelica sinensis (2-10 parts), Ligusticum striatum (2-10 parts), Aristolochia debilis (1-10 parts), Sandalwood (1-10 parts), Aucklandia lappa (1-10 parts), Clove (1-10 parts); Place the evenly mixed herbal raw materials in a ceramic jar, add the herbal raw materials and base liquor (20-70% vol light aroma type baijiu) in a ratio of 1:5 to 1:30 (kg:L), soak for 10-30 days, then filter to obtain Zhuyeqing wine.

[0038] II. Preparation of extracts from bamboo leaf green wine.

[0039] Take 1000 mL of Zhuyeqing liquor and place it in a rotary evaporator. Set the water bath temperature to 50℃ and the vacuum degree to 0.09MPa. Rotary evaporate until anhydrous ethanol is fully distilled out to obtain an aqueous concentrate.

[0040] The above concentrate was transferred to a vacuum freeze dryer, and the cold trap temperature was set to -55℃ and the vacuum degree to 5 Pa. The freeze dryer was set for 48 h to constant weight, and 19.7 g of powdered Zhuyeqing wine extract was obtained, with a yield of 1.97 g / 100 mL. The extract was sealed and stored at 4℃. The Zhuyeqing wine extract was designated as ZLE in subsequent experiments.

[0041] Example 2 Determination of the in vitro antioxidant activity of ZLE.

[0042] ZLE prepared in Example 1 was used to prepare 1 mg / mL and 5 mg / mL ZLE solutions with distilled water. Using commercially available kits and following the instructions, the total antioxidant capacity, superoxide anion radical scavenging rate, DPPH radical scavenging capacity, and ABTS radical scavenging capacity of the samples were determined. The results are shown below. Figure 1As shown in the figure, the total antioxidant capacity of ZLE ranges from 1.06 to 3.84 µmol / mL when the concentration is 1–5 mg / mL. At a concentration of 5 mg / mL, the scavenging rates of superoxide anion radicals, DPPH radicals, and ABTS radicals were 14.3%, 42.4%, and 81.8%, respectively; while at a concentration of 1 mg / mL, the scavenging rates of these three radicals were 3.1%, 7.6%, and 9.4%, respectively. These results indicate that ZLE possesses significant concentration-dependent in vitro antioxidant activity, and exhibits excellent total antioxidant capacity and free radical scavenging activity at a concentration of 5 mg / mL.

[0043] Example 3 The protective effect of ZLE on H2O2-induced HT-22 neurons.

[0044] I. Cell Culture.

[0045] HT-22 cells were cultured in DMEM complete medium containing 10% fetal bovine serum and 1% penicillin-streptomycin antibiotics. The cells were then placed in a 37°C, 5% CO2 incubator and the medium was replaced with fresh medium every 24–48 h. When the cell confluence reached 80%–90%, the cells were passaged, and cells in the logarithmic growth phase were selected for subsequent experiments.

[0046] II. Effects of ZLE on HT-22 cell viability.

[0047] HT-22 cells were loaded at 5 × 10 3 Cells were seeded at a density of 10 cells / well in 96-well plates and cultured for 24 h. Fresh medium containing 10, 25, 50, 100, and 500 μg / mL ZLE was added for intervention. The blank control group was replaced with an equal volume of fresh medium without ZLE. After culturing for another 24 h, CCK-8 reagent was added to each well, and the cells were incubated at 37°C. The absorbance (OD) at 450 nm was measured using a microplate reader. Data were recorded, and cell viability was calculated for each group. The experimental results are shown below. Figure 2 As shown in the figure, it can be seen that the concentration range of bamboo leaf green wine extract in the range of 25~500 μg / mL has no significant toxic effect on the viability of HT-22 cells.

[0048] III. Half-maximal inhibitory concentration (IC50) of H2O2-induced oxidative damage in HT-22 cells 50 ) Measurement.

[0049] HT-22 cells were seeded in 96-well plates and cultured for 24 h. Then, culture media containing 200 μM, 400 μM, 600 μM, 800 μM, and 1000 μM H2O2 were added, respectively. After 4 h of incubation, cell viability was assessed using the CCK-8 assay. The results are shown below. Figure 3 As shown; Figure A shows the cell survival rate at different H2O2 concentrations, and Figure B shows the dose-response curve fitting graph. The figure shows that the IC50 value for H2O2-induced apoptosis in HT-22 cells is... 50 The concentration was 650 μM, so this concentration was selected as the modeling concentration for the subsequent cellular oxidative damage model.

[0050] IV. Effects of Bamboo Leaf Green Wine Extract on H2O2-Induced Viability of HT-22 Cells.

[0051] Cells were divided into a control group (Ctrl), a model group (H2O2), and ZLE treatment groups (10, 50, 100, and 500 μg / mL). The treatment groups were pretreated with culture medium containing the corresponding concentration of extract for 24 h. The Ctrl and H2O2 groups were added to complete culture medium. Except for the control group, all other groups were added to culture medium containing 650 μM H2O2. After culturing for another 4 h, cell viability was measured using the CCK-8 assay. The results are as follows: Figure 4 As shown in the figure, it can be seen that the extract at a concentration of 500 μg / mL reduces the survival rate of HT-22 cells treated with H2O2; while pretreatment with extracts at concentrations of 50 μg / mL and 100 μg / mL can significantly improve the survival rate of HT-22 cells damaged by H2O2.

[0052] V. Detection of intracellular ROS levels.

[0053] Cells were divided into a control group (Ctrl), a model group (H2O2), a low-dose ZLE group (10 μg / mL), and a high-dose ZLE group (100 μg / mL). After drug administration and model establishment as described above, cells were collected, incubated with the DCFH-DA probe, and intracellular fluorescence intensity was detected by flow cytometry to characterize ROS levels. The results are as follows: Figure 5 As shown in the figure, it can be seen that the intracellular ROS level in the H2O2 group was significantly increased, while ZLE at 10 μg / mL and 100 μg / mL could significantly reduce intracellular ROS accumulation and alleviate oxidative stress damage.

[0054] VI. Cell cycle detection.

[0055] HT-22 cells were treated with ZLE at concentrations of 10 μg / mL and 100 μg / mL, respectively, to investigate the regulatory effect of ZLE on neuronal cell cycle progression induced by oxidative stress. The results are as follows: Figure 6 As shown in the figure, H2O2 treatment significantly increased the proportion of HT-22 cells in the G1 phase and significantly decreased the proportion in the G2 phase, indicating that oxidative stress can cause cell cycle arrest in HT-22 cells; while 100 μg / mL ZLE can significantly reverse the abnormal cell cycle distribution and alleviate the cell cycle arrest caused by oxidative stress.

[0056] VII. Detection of intracellular antioxidant and inflammation-related gene expression.

[0057] The effects of ZLE on the regulation of key antioxidant enzymes (CAT, SOD) and pro-inflammatory cytokines (TNF-α, IL-1β, IL-6) in H2O2-induced damaged HT-22 cells were investigated using real-time quantitative polymerase chain reaction (RT-qPCR). The results are as follows: Figure 7 As shown in the figure, ZLE can upregulate the mRNA expression of catalase (CAT) and superoxide dismutase (SOD) in HT-22 cells, while downregulating the mRNA expression of pro-inflammatory cytokines tumor necrosis factor-α (TNF-α), interleukin-1β (IL-1β), and interleukin-6 (IL-6). This result indicates that ZLE has both antioxidant and anti-inflammatory activities.

[0058] Example 4 The protective effect of bamboo leaf green wine extract on D-galactose-induced aging mice.

[0059] I. Laboratory Animals and Grouping

[0060] Eight-week-old male SPF-grade ICR mice were selected and, after one week of acclimatization, were randomly divided into five groups (n=8): normal control group (CON), model group (MOD), vitamin C positive group (VC, 100 mg / kg / d), low-dose ZLE group (ZL, 100 mg / kg / d), and high-dose ZLE group (ZH, 200 mg / kg / d).

[0061] II. Modeling and Drug Administration.

[0062] Except for the CON group, mice in the other groups were subcutaneously injected with 500 mg / kg D-galactose daily, while the CON group was injected with an equal volume of physiological saline. Simultaneous drug intervention was implemented during model establishment: mice in the VC, ZL, and ZH groups were administered the corresponding dose of the test substance via gavage daily, while the normal control group and model group were administered an equal volume of distilled water via gavage. This intervention lasted for 8 weeks. During the experiment, mice had free access to food and water, and changes in mouse weight and food intake were recorded weekly.

[0063] III. Sample Collection and Processing.

[0064] After 8 weeks of intervention, all mice were fasted for 12 hours and blood was collected from the orbital cavity under anesthesia. The blood samples were centrifuged at 3000 g for 15 min at 4°C, and the supernatant serum was separated and stored at -80°C. The mice were dissected, and the spleen and liver were separated and weighed. The organ indices of each group of mice were calculated. The brain tissue was dissected and divided into two parts: one part was fixed with 4% paraformaldehyde for pathological and immunofluorescence detection; the other part was frozen at -80°C for molecular biological index detection.

[0065] IV. Results of physiological index testing in mice.

[0066] The results of physiological index tests on different groups of mice, such as Figure 8 As shown in the figure; Figure A is a curve of body weight change over time, Figure B is a bar chart of the reduction changes of food intake in different groups of mice, Figure C is a bar chart of spleen index in different groups of mice, and Figure D is a bar chart of liver index in different groups of mice. from Figure 8 It can be seen that, compared with the normal control group, the model group mice showed a significantly slower rate of weight gain, a significant decrease in food intake, and a significant decrease in spleen and liver indices starting from week 4 of intervention. This suggests that D-galactose-induced aging model mice exhibit growth retardation, abnormal feeding, and atrophy and damage to immune and metabolic organs. Compared with the model group, both low- and high-dose ZLE intervention can effectively improve the slow weight gain in aging mice, correct the abnormal decrease in food intake, significantly reverse the decline in spleen and liver indices, and effectively restore the normal physiological growth and organ function of mice.

[0067] V. Morris water maze test to assess spatial learning and memory abilities in mice.

[0068] The Morris water maze (MWM) test was used to assess the spatial learning and memory abilities of mice. The experiment included 5 days of orientation and navigation training and 1 day of spatial exploration. The results are as follows: Figure 9 As shown; Figure A shows the escape latency results of the mouse positioning and navigation experiment; Figure B shows the bar chart of the time the mouse spent in the target quadrant; Figure C shows the bar chart of the number of times the mouse crossed the platform; and Figure D shows the representative swimming trajectory of the mouse. from Figure 9It can be seen that during the 4th to 5th day of the navigation training, the escape latency of mice in the ZL and ZH groups was significantly shorter than that in the model group, suggesting that ZLE can improve the spatial learning ability of aging mice. In the spatial exploration experiment, the time spent in the target quadrant of mice in the ZL and ZH groups was significantly longer than that in the model group, the number of platform crossings was significantly increased, and the swimming trajectory of the mice was more concentrated in the target quadrant, indicating that ZLE can effectively improve the spatial learning and memory function impairment in aging mice.

[0069] VI. Serum antioxidant index detection.

[0070] Antioxidant markers in mouse serum, including the levels of superoxide dismutase (SOD), malondialdehyde (MDA), glutathione (GSH), and glutathione peroxidase (GSH-PX), were determined using appropriate detection kits. The results are as follows: Figure 10 As shown; Figure A shows the serum SOD test results, Figure B shows the serum MDA test results, Figure C shows the serum GSH test results, and Figure D shows the serum GSH-PX test results. from Figure 10 It can be seen that compared with the normal control group, the serum SOD, GSH, and GSH-PX levels of the model group mice were significantly reduced, while the MDA content was significantly increased. After ZLE intervention, the serum SOD, GSH, and GSH-PX activities of aging mice were significantly restored, while the MDA content was significantly reduced. This indicates that ZLE can effectively alleviate D-galactose-induced oxidative stress damage and enhance the antioxidant capacity of aging mice.

[0071] VII. Hippocampal histopathology and immunofluorescence detection.

[0072] Paraffin sections of mouse brain tissue were prepared. H&E staining was used to observe the pathological morphology of the hippocampus, and GFAP and Iba1 immunofluorescence staining was used to detect the activation of inflammatory cells in the brain tissue. The results are as follows: Figure 11 As shown; Figure A shows the H&E staining pattern. From this figure, we can see that the neurons in the CA1 and DG regions of the hippocampus of the normal control group mice are neatly arranged and have intact morphology; the neurons in the CA1 and DG regions of the hippocampus of the model group mice are disordered, with obvious nuclear condensation, neuronal degeneration and other aging pathological damage; after ZLE intervention, the pathological morphology of the hippocampal neurons of mice was significantly improved and the degree of neuronal damage was significantly reduced. Figure B shows the immunofluorescence staining pattern. From this figure, it can be seen that the area of ​​GFAP and Iba1 positive cells in the hippocampus of the model group mice was significantly increased, indicating that astrocytes and microglia in the brain tissue of aging mice were abnormally activated in large numbers, inducing central nervous system inflammation. ZLE intervention can significantly reduce the area of ​​GFAP and Iba1 positive signal expression, effectively inhibit the excessive activation of astrocytes and microglia induced by D-galactose, and reduce the central nervous system inflammatory response in mice.

[0073] 8. Detection of molecular pathways in brain tissue.

[0074] Total RNA was extracted from mouse brain tissue using the TRIzol method, and the mRNA expression levels of SIRT1, Nrf2, HO-1, NF-κB, TNF-α, and IL-1β were detected by RT-qPCR. The results are as follows: Figure 12 As shown in the figure, ZLE significantly downregulates the mRNA expression levels of NF-κB, TNF-α, and IL-1β in mouse brain tissue, effectively inhibiting the activation of NF-κB-mediated inflammatory pathways. Simultaneously, it significantly upregulates the mRNA expression levels of SIRT1, Nrf2, and HO-1 in brain tissue, activating the SIRT1-Nrf2-HO-1 antioxidant signaling pathway. These results further confirm at the molecular level that ZLE can exert its effects of antioxidative damage, inhibiting neuroinflammation, and alleviating aging in mice by regulating antioxidant and inflammation-related pathways.

[0075] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention, and not to limit them; although the present invention has been described in detail with reference to the foregoing embodiments, those skilled in the art should understand that modifications can still be made to the technical solutions described in the foregoing embodiments, or equivalent substitutions can be made to some of the technical features; and these modifications or substitutions do not cause the essence of the corresponding technical solutions to deviate from the spirit and scope of the technical solutions of the embodiments of the present invention.

Claims

1. Use of a bamboo-leaf-green liquor in the preparation of an anti-aging product, characterized in that, The anti-aging product includes an extract of Zhuyeqing wine, which is obtained by removing alcohol from Zhuyeqing wine and drying it.

2. The application of the bamboo leaf green wine as described in claim 1 in the preparation of anti-aging products, characterized in that, The anti-aging product is used to prevent or treat at least one of the following age-related diseases: oxidative stress damage, neuroinflammation, cognitive impairment, and neurodegenerative diseases.

3. The application of the bamboo leaf green wine as described in claim 1 in the preparation of anti-aging products, characterized in that, The anti-aging product has a neuroprotective effect.

4. The application of the bamboo leaf green wine as described in claim 3 in the preparation of anti-aging products, characterized in that, The anti-aging product exerts a neuroprotective effect through anti-oxidation and / or anti-inflammation.

5. The application of the bamboo leaf green wine as described in claim 4 in the preparation of anti-aging products, characterized in that, The anti-aging product enhances the body's antioxidant capacity by scavenging free radicals, including at least one of ABTS free radicals, DPPH free radicals, and superoxide anion free radicals. ABTS is 2,2'-bis(3-ethylbenzothiazoline-6-sulfonic acid) diammonium salt, and DPPH is 1,1-diphenyl-2-trinitrophenylhydrazine.

6. The application of the bamboo leaf green wine as described in claim 4 in the preparation of anti-aging products, characterized in that, The anti-aging product can achieve antioxidant effects through at least one of the following: restoring the activity of superoxide dismutase, glutathione, and glutathione peroxidase in the body, and reducing abnormally elevated malondialdehyde levels.

7. The application of the bamboo leaf green wine as described in claim 4 in the preparation of anti-aging products, characterized in that, The anti-aging product combats inflammation by inhibiting the mRNA expression of at least one of the following pro-inflammatory cytokines: tumor necrosis factor-α, interleukin-1β, and interleukin-6.

8. The application of the bamboo leaf green wine as described in claim 1 in the preparation of anti-aging products, characterized in that, The anti-aging products can improve the body's spatial learning and memory abilities.

9. The application of the bamboo leaf green wine as described in claim 4 in the preparation of anti-aging products, characterized in that, The anti-inflammatory effects include those against central nervous system inflammation; The anti-aging product inhibits the excessive activation of astrocytes and microglia in brain tissue, downregulates the mRNA expression of NF-κB, TNF-α, and IL-1β in brain tissue, and blocks the central nervous system inflammatory cascade, thereby achieving anti-central nervous system inflammation.

10. The application of the bamboo leaf green wine as described in claim 1 in the preparation of anti-aging products, characterized in that, The anti-aging products include at least one of anti-aging drugs and anti-aging foods.

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