Application of bamboo leaf green liquor in preparation of medicine for preventing and treating Parkinson disease

Abstract

The application discloses application of bamboo green wine in preparation of medicines for preventing and treating Parkinson's disease, belongs to the technical field of biological medicine, and the concentration of the bamboo green wine is 2.5-40 % Vol; the application further discloses a medicine for preventing and treating Parkinson's disease, which comprises bamboo green wine and pharmaceutically acceptable adjuvants. It is found that the bamboo green wine can enhance the movement ability and stress resistance of nematodes, reduce the abnormal aggregation of alpha-Syn protein, protect the damage of dopaminergic nerve cells, delay the clinical symptoms of Parkinson's disease, and further treat Parkinson's disease. Compared with other medicines for treating Parkinson's disease, the bamboo green wine is a health wine for eating, and the medicinal value of treating Parkinson's disease can be achieved in the process of eating, so that the traditional health wine is beneficial to further development and utilization of the efficacy.

Description

TECHNICAL FIELD

[0001] The application relates to the technical field of biological medicine, in particular to an application of bamboo-leaf-green wine in preparation of a medicine for preventing and treating Parkinson's disease. BACKGROUND

[0002] Parkinson's disease (PD) is a kind of neurodegenerative disease with a continuously increasing incidence in recent years, and is only second to Alzheimer's disease. The disease mainly occurs in people over 60 years old, and the incidence rate of women is higher than that of men, but the mortality rate of men is higher than that of women. The early clinical features of the disease are mainly motor dysfunction, such as slow behavior, muscle rigidity, tremor, movement disorder, balance dysfunction and the like. The symptoms gradually worsen in the later stage, leading to cognitive dysfunction of the patient's brain, language dysfunction and even loss due to damage of the vocal cords, decreased digestive function leading to constipation, decreased sleep quality leading to insomnia and olfactory dysfunction. These clinical features make the life quality and happiness index of the patient decrease linearly, and the disease is a great burden to the patient's family. Therefore, the treatment of Parkinson's disease is imminent.

[0003] Studies have shown that Parkinson's disease is caused by the abnormal accumulation of alpha-syn in dopaminergic neurons, leading to the deformation and death of dopaminergic neurons and loss of functional activity. However, all current studies have not clearly defined the complete pathogenesis of Parkinson's disease. It is reported that family history, gene mutation and environmental factors can all cause Parkinson's disease. At present, a large amount of evidence shows that the main causes of Parkinson's disease are family history and gene mutation. The common mutation genes are SNCA, VPS35, PINK1 and Parkin. SNCA is a gene encoding alpha-syn, and abnormal expression of the gene will lead to abnormal accumulation of alpha-syn to form Lewy bodies, and then cause Parkinson's disease. It has been found that the VPS35 gene is the cause of late-onset familial Parkinson's disease, and the mutation of the gene leads to lysosomes and autophagy. The mechanism of PINK1 and Parkin is mainly the ubiquitination-dependent pathway, and the key proteins are PINK1 and Parkin, which are called PINK1 / Parkin-mediated mitochondrial autophagy pathway. Ultimately, the expression of autophagy protein LC3 is up-regulated, which triggers the function of mitochondrial autophagy and maintains the homeostasis of protein import and the normal functional state of cells. From the current research, it is a recognized difficult problem to find a drug that can treat Parkinson's disease. At present, the main drug for treating Parkinson's disease in the clinic is levodopa, but it has certain side effects, such as arrhythmia, nausea and vomiting, and mental disorders. Many experts point out that food therapy not only has significant effects and small side effects, but more importantly, it can reduce the pain of patients and the burden of their families. Therefore, it is of great significance to find a health food that can delay Parkinson's disease.

[0004] Zhuyeqing wine originated in the Warring States period and has a history of over a thousand years. It has been widely loved and its popularity has endured. Zhuyeqing wine uses Fenjiu, a specialty baijiu from Shanxi Province, as its base. It is made using a special process with 12 kinds of traditional Chinese medicinal herbs, including bamboo leaves, gardenia, chrysanthemum, dried tangerine peel, angelica, cardamom, cloves, costus root, galangal, sandalwood, agastache rugosa, and fragrant grass. It possesses both the distinctive aroma of Fenjiu and the unique flavor of medicinal herbs, resulting in Zhuyeqing wine's unique taste and efficacy. Research shows that Zhuyeqing wine is rich in volatile components, including 11 phenolic acids, 8 alkenes, 7 esters, 4 alcohols, and 1 other type of compound. It also contains 80 non-volatile compounds, including 25 flavonoids, 24 phenolic acids, 14 aldehydes and esters, 3 saponins, 5 terpenes, and 9 other compounds. The unique ratio of volatile to non-volatile components gives Zhuyeqing wine its unparalleled medicinal and edible functions. Current research on Zhuyeqing wine has revealed its antioxidant, anti-inflammatory, anti-tumor, anti-aging, anti-fatigue, antibacterial, and liver-protective effects. However, there are currently no reports on its efficacy in treating the neurodegenerative disease Parkinson's disease. Summary of the Invention

[0005] The purpose of this invention is to provide the application of Zhuyeqing wine in the preparation of drugs for the prevention and treatment of Parkinson's disease, so as to solve the problems existing in the prior art. Zhuyeqing wine can delay the clinical symptoms of Parkinson's disease and play a role in the treatment of Parkinson's disease. This invention provides a new idea for the treatment of Parkinson's disease.

[0006] To achieve the above objectives, the present invention provides the following solution:

[0007] This invention provides the use of Zhuyeqing wine in the preparation of drugs for the prevention and / or treatment of Parkinson's disease.

[0008] Furthermore, the concentration of the Zhuyeqing wine is 2.5-40% Vol.

[0009] Furthermore, the concentration of the Zhuyeqing wine is 20% Vol.

[0010] Furthermore, the bamboo leaf green wine plays a role in preventing and / or treating Parkinson's disease by improving the heat stress resistance, oxidative stress resistance, motor ability and dopaminergic neuron content of nematodes in the Parkinson's disease model, enhancing the food perception behavior of nematodes in the Parkinson's disease model, and reducing the content of α-syn protein.

[0011] The present invention also provides a drug for the prevention and treatment of Parkinson's disease, comprising bamboo leaf green wine and pharmaceutically acceptable excipients.

[0012] Furthermore, the bamboo leaf green wine is the only effective ingredient, with a concentration of 2.5-40% Vol.

[0013] Further, the adjuvant comprises one or more of diluents, fillers, excipients, binders, humectants, disintegrants, absorption promoters, surfactants, adsorptive carriers, lubricants, and flavoring agents.

[0014] Further, the dosage form of the drug comprises tablets, pills, sprays, granules, capsules, soups, or suspensions.

[0015] Further, the mode of administration of the drug comprises oral administration or parenteral administration.

[0016] The present application discloses the following technical effects:

[0017] The present application uses the model organism Caenorhabditis elegans as a carrier to study the therapeutic effect of Zhuyeqing liquor on neurodegenerative disease Parkinson's disease. On the one hand, the effect of Zhuyeqing liquor on the movement ability, stress resistance, and alpha-Syn protein accumulation of Parkinson's disease Caenorhabditis elegans model NL5901 nematodes is studied. On the other hand, the effect of Zhuyeqing liquor on the expression of DA neurons and food perception behavior of BZ555 nematodes is studied. The results show that Zhuyeqing liquor can enhance the movement ability and stress resistance of nematodes and reduce the abnormal aggregation of alpha-Syn protein, thereby protecting the damage of dopaminergic neurons, delaying the clinical symptoms of Parkinson's disease, and treating Parkinson's disease.

[0018] The present application first discovers that Zhuyeqing liquor can treat Parkinson's disease, and compared with other drugs for treating Parkinson's disease, Zhuyeqing liquor can achieve the medicinal value of treating Parkinson's disease during the eating process. BRIEF DESCRIPTION OF DRAWINGS

[0019] In order to more clearly illustrate the technical solutions in the embodiments of the present application or the prior art, the drawings needed in the embodiments will be briefly introduced below. Obviously, the drawings in the following description are only some embodiments of the present application, and other drawings can also be obtained by those skilled in the art without creative labor.

[0020] Figure 1 Influence of Zhuyeqing liquor on the heat stress resistance of Parkinson's disease model NL5901 nematodes;

[0021] Figure 2 Influence of Zhuyeqing liquor on the movement ability of NL5901 nematodes;

[0022] Figure 3 Influence of Zhuyeqing liquor on the expression of DA neurons of BZ555 nematodes; wherein A is the fluorescence microscope observation result, B is the GFP fluorescence intensity statistical result; the scale in A is 20 μm;

[0023] Figure 4 Effects of Zhuyeqing liquor on food perception behavior of BZ555 nematodes;

[0024] Figure 5 Effects of Zhuyeqing liquor on synuclein alpha-syn of NL5901 nematodes; wherein A is fluorescence microscope observation result, and B is GFP fluorescence intensity statistical result; the scale in A is 20 μm;

[0025] Figure 6 Effects of Zhuyeqing liquor on the antioxidant stress resistance of NL5901 nematodes. DETAILED DESCRIPTION

[0026] Various exemplary embodiments of the present application will now be described in detail, which should be considered to be illustrative of certain aspects, features and embodiments of the present application, but not a limitation thereof.

[0027] It should be understood that the terms used in the present application merely describe particular embodiments, and are not intended to limit the present application. In addition, for the numerical range in the present application, it should be understood that each intermediate value between the upper limit and the lower limit of the range is also specifically disclosed. Each smaller range between any stated value or stated range of values, and any other stated value or stated range of values in the described range is also encompassed within the scope of the present application. The upper limit and the lower limit of these smaller ranges can be included or excluded independently from the range.

[0028] Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this application belongs. Although methods and materials similar or equivalent to those described herein can be used in the practice or testing of the present application, preferred methods and materials are described. All publications mentioned in the specification are incorporated by reference to disclose and describe the methods and / or materials in connection with which the publications are cited. The citation of any reference in the specification is not intended as an admission that the reference is prior art to the application described and claimed herein.

[0029] Many modifications and variations of this application specification can be made in light of its teachings, which would be apparent to those of skill in the art. Other embodiments of the application will be apparent to those of skill in the art from the specification and examples given. The specification and examples are illustrative only.

[0030] As used herein, the terms "comprise", "comprising", "include", "including", "have", "having", and the like are open-ended terms that are intended to mean including, but not limited to.

[0031] Example 1

[0032] The term "Zhu Ye Qing wine" used in the present application contains 12 kinds of Chinese herbal medicines, which are purchased from Shanxi Xinghuacun Distillery Co., Ltd.

[0033] In the following examples, NL5901 nematode refers to Caenorhabditis elegans NL5901, which is purchased from Caenorhabditis Genetics Center (CGC); BZ555 nematode refers to transgenic Caenorhabditis elegans strain BZ555, which is purchased from Caenorhabditis Genetics Center (CGC); E. coli OP50 is a gift from Professor Ma Enbo of Shanxi University Institute of Applied Biology.

[0034] Example 1 Zhu Ye Qing wine improves the anti-heat stress ability of Parkinson's disease model NL5901 nematode

[0035] 1. Dilution of ZYQ wine concentration: The commercially available ZYQ wine with an alcohol degree of 45% is first diluted with PBS to 2.5%, 5%, 10%, 20%, and 40%, and then diluted with E. coli OP50 bacterial solution at a dilution ratio of 1:1. The control group is bacterial solution without Zhu Ye Qing wine, which is also diluted with PBS at a dilution ratio of 1:1. After blowing dry in a clean bench, the NL5901 nematodes are evenly coated on NGM plates. After 3 days of treatment, the nematodes are placed in a 37°C constant temperature incubator, recorded as 0h, and then the survival rate of nematodes is recorded every hour until the nematodes completely die (if the nematodes move their heads, they are determined to be alive, and if the nematodes do not move their heads, they are determined to be dead). The experiment is repeated independently for 3 times.

[0036] 2. The data of nematode lifespan experiment is analyzed using the Kaplan-Meier survival function in SPSS22.0 (demo version, Armonk, NY) software, and the log-rank test is used for significance analysis. The experimental results are plotted using OriginLab plotting software. P<0.05 is considered as statistically significant difference between the two groups (*P<0.05 indicates significant difference between the two groups, **P<0.01 indicates extremely significant difference between the two groups).

[0037] Experimental results:

[0038] The experimental results show that, as shown in Figure 1 compared with the control group, the concentrations of 5%, 10%, 20%, and 40% ZYQ can significantly improve the anti-heat stress ability of NL5901 nematodes, among which the improvement effect of 20% ZYQ is extremely significant compared with the control, which is increased by 24.75%. It is proved that ZYQ can improve the anti-heat stress ability of NL5901 nematodes.

[0039] Table 1 Effect of ZYQ on the anti-heat stress ability of Parkinson's disease model nematodes

[0040]

[0041]

[0042] Example 2 Bamboo green liquor can delay the decrease of NL5901 nematode movement ability

[0043] 1. Dilution of ZYQ liquor concentration, the experimental group first dilutes the commercially available ZYQ with an alcohol degree of 45% to 2.5%, 5%, 10%, 20%, and 40% with PBS, and then dilutes the E. coli OP50 bacterial solution at a dilution ratio of 1:1. The control group is the bacterial solution without bamboo green liquor, which is diluted with PBS at a dilution ratio of 1:1. After blowing dry in a clean bench, they are ready for use. Synchronized NL5901 nematodes in L4 stage are picked up with platinum wire and placed on different NGM plates. After 3 days of treatment, the worms are transferred to M9 buffer for 90 seconds of pressure recovery. The sine movement frequency of 30 nematodes in 30 seconds is measured, and the experiment is repeated three times.

[0044] Experimental results:

[0045] The harmful α-Syn aggregation in NL5901 nematodes can cause DA neuron toxicity, leading to movement disorders in nematodes, and has a certain toxic effect on the movement ability of nematodes. Therefore, the present application studies the effect of bamboo green liquor on the movement ability of NL5901 nematodes. The experimental results show that 2.5%, 5%, 10%, 20%, and 40% ZYQ concentrations can improve the movement ability of NL5901 nematodes by 11%, 17%, 24%, 9%, and 2%, respectively. The results are shown in Table 1, indicating that ZYQ can significantly improve the movement ability of NL5901 nematodes and delay the movement dysfunction caused by the loss of DA neuron function. Figure 2

[0046] Example 3 Bamboo green liquor can improve the content of dopaminergic neurons in BZ555 nematodes

[0047] ​1. Dilution of ZYQ alcohol concentration: In the experimental group, commercially available 45% ZYQ alcohol was first diluted to 10%, 20%, and 40% with PBS, and then diluted with E. coli OP50 bacterial suspension at a 1:1 ratio. In the control group, bacterial suspension without ZYQ alcohol was diluted with PBS at the same 1:1 ratio. The solutions were then evenly spread on NGM plates and dried in a clean bench. Thirty synchronized BZ555 nematodes that had reached the L4 stage were picked up with platinum wire and transferred to corresponding NGM plates. After 3 days of treatment, the dopaminergic neuron content of BZ555 nematodes was observed using delta-vision deconvolution fluorescence microscopy, and statistical analysis was performed.

[0048] Experimental results:

[0049] In the BZ555 nematode, dopaminergic neurons exhibit GFP fluorescent labeling signals, and dopaminergic neuronal degeneration and death are characteristic of the brains of PD patients. Based on the experimental results... Figure 3 As shown in Figures A and B, compared to the control group, 20% Vol ZYQ increased the content of dopaminergic neurons in BZ555 nematodes by 10.44%, and 2mM L-dopa by 11.91%. Zhuyeqing wine significantly increased the fluorescence signal intensity of DA neurons in BZ555 nematodes, achieving effects consistent with levodopa, a clinical drug for Parkinson's disease, indicating that Zhuyeqing wine can enhance the expression of DA neurons in nematodes, thus delaying the progression of Parkinson's disease.

[0050] Example 4: Bamboo Leaf Green Wine Enhances Food Sensing Behavior of BZ555 Nematodes

[0051] To dilute the ZYQ alcohol concentration, the experimental group first diluted commercially available 45% ZYQ alcohol to 10%, 20%, and 40% with PBS, and then diluted it with E. coli OP50 bacterial suspension at a 1:1 ratio. The control group used bacterial suspension without ZYQ alcohol diluted with PBS at the same 1:1 ratio. Both solutions were then evenly spread on NGM plates and dried in a clean bench. Thirty synchronized BZ555 nematodes that had reached the L4 stage were picked up with platinum wire and transferred to different NGM plates. After 3 days of treatment, each group of nematodes was transferred to NGM medium containing food and those without. After 3 minutes at room temperature, the movement of the nematodes on the food-containing / food-free NGM plates was observed and recorded under a stereomicroscope for 30 seconds. The food perception behavior of the nematodes was calculated using the following formula:

[0052] Food perception behavior = (Number of nematode movements under sterile conditions - Number of nematode movements under sterile conditions) / Number of nematode movements under sterile conditions × 100%.

[0053] Experimental results:

[0054] The damage of DA neurons makes the movement ability of the nematodes in an environment with sufficient food to be enhanced, and the movement ability of nematodes without damage of DA neurons in an environment with sufficient food is slowed down due to the integrity of foraging behavior. According to the experimental results, it is shown that the ZYQ can significantly improve the food perception behavior of BZ555 nematodes. Figure 4 It can be seen from the above table that, compared with the control group, 10% Vol ZYQ is increased by 10.24%, 20% Vol ZYQ is increased by 17.38%, 40% Vol ZYQ is increased by 7.41%, and 2mM L-dopa is increased by 21.61%, which is consistent with the effect of L-dopa, a clinical drug for Parkinson's disease, indicating that ZYQ has the effect of delaying Parkinson's disease.

[0055] Example 5: ZYQ reduces the aggregation of α-Syn in NL5901 nematodes

[0056] Dilution of ZYQ concentration: the commercially available ZYQ with an alcohol degree of 45% is first diluted with PBS to 10%, 20%, and 40%, and then diluted with E. coli OP50 bacterial solution at a dilution ratio of 1:1. The control group is the bacterial solution without ZYQ, which is also diluted with PBS at a dilution ratio of 1:1, and then uniformly coated on NGM plates and placed in a clean bench to dry. The synchronized NL5901 nematodes reaching the L4 stage are picked up with platinum wire to the corresponding different NGM plates, and after 3 days of treatment, the expression of α-Syn green fluorescence in NL5901 nematodes is observed by laser confocal microscope, and statistical analysis is performed.

[0057] Experimental results:

[0058] The expression amount of α-syn in the striatum of the brain of Parkinson's disease patients is much higher than that of normal people, and the accumulation of a large amount of α-syn forms Lewy bodies, leading to the degeneration and death of dopaminergic neurons, so the content of α-syn is used as an indicator for the detection of Parkinson's disease. The experimental results show that ZYQ can significantly reduce the content of α-syn in NL5901 nematodes, and compared with the control group, 20% Vol ZYQ can extremely significantly reduce the protein content. The results are shown in Figure 5 A-B of the above table.

[0059] Example 6: ZYQ improves the antioxidant stress resistance of NL5901 nematodes

[0060] 1. Dilution of ZYQ alcohol concentration: In the experimental group, commercially available ZYQ with an alcohol content of 45% was first diluted to 10%, 20%, and 40% with PBS, and then diluted with E. coli OP50 bacterial suspension at a 1:1 ratio. In the control group, bacterial suspension without ZYQ alcohol was diluted with PBS at the same 1:1 ratio. The solutions were then evenly spread on NGM plates and dried in a clean bench. Thirty synchronized NL5901 nematodes that had reached the L4 stage were picked up with a platinum wire and placed on different NGM plates. After 3 days of treatment, they were placed in NGM medium containing 10 mM hydrogen peroxide. The nematodes were subjected to oxidative stress at 20°C for 0 h. The survival rate of the nematodes was recorded every hour until they died completely (the nematodes were gently touched with a platinum wire; if their heads moved, they were considered alive; if their heads did not move, they were considered dead). The experiment was independently repeated 3 times.

[0061] 2. The nematode lifespan experimental data were analyzed using the Kaplan-Meier survival function in SPSS 22.0 (demo version, Armonk, NY) software, and the log-rank test was used for significance analysis. Experimental results were plotted using OriginLab software. P < 0.05 was considered statistically significant between the two groups (*P < 0.05 indicates a significant difference between the two groups, **P < 0.01 indicates an extremely significant difference between the two groups).

[0062] Experimental results:

[0063] like Figure 6 As shown in Table 2, compared with the control group, 10%, 20%, and 40% ZYQ all improved the survival rate of NL5901 nematodes. Table 2 also shows that 10% and 20% ZYQ significantly improved the antioxidant stress capacity of NL5901 nematodes in the Parkinson's disease model compared to the control, increasing it by 14.38% and 39.21%, respectively. The 20% Vol ZYQ showed the most significant improvement, demonstrating that ZYQ can enhance the antioxidant stress capacity of NL5901 nematodes and has a delaying effect on Parkinson's disease.

[0064] Table 2. Effects of ZYQ on the antioxidant stress resistance of nematodes in a Parkinson's disease model.

[0065]

[0066] The embodiments described above are merely preferred embodiments of the present invention and are not intended to limit the scope of the present invention. Various modifications and improvements made by those skilled in the art to the technical solutions of the present invention without departing from the spirit of the present invention should fall within the protection scope defined by the claims of the present invention.

Claims

1. The application of Zhuyeqing wine in the preparation of drugs for the prevention and / or treatment of Parkinson's disease, characterized in that, The bamboo leaf green wine plays a role in preventing and / or treating Parkinson's disease by improving the heat stress resistance, oxidative stress resistance, motor ability, and dopaminergic neuron content of nematodes in a Parkinson's disease model, as well as enhancing the food perception behavior of nematodes and reducing α-syn protein content; the concentration of the bamboo leaf green wine in the drug is 20% Vol. The Zhuyeqing liquor is made from Fenjiu as the base liquor and 12 kinds of Chinese herbal medicines, including bamboo leaves, gardenia, chrysanthemum, dried tangerine peel, angelica, amomum, clove, costus root, galangal, sandalwood, lingling incense, and fragrant grass, through processing.

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