CN111939190A - Application of sparrow mouth tea or extract thereof in preparation of medicine or health product for preventing and treating acute liver injury - Google Patents
Application of sparrow mouth tea or extract thereof in preparation of medicine or health product for preventing and treating acute liver injury Download PDFInfo
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K36/00—Medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicines
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- A23L33/00—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof
- A23L33/10—Modifying nutritive qualities of foods; Dietetic products; Preparation or treatment thereof using additives
- A23L33/105—Plant extracts, their artificial duplicates or their derivatives
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- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
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- A61K2236/00—Isolation or extraction methods of medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicine
- A61K2236/30—Extraction of the material
- A61K2236/33—Extraction of the material involving extraction with hydrophilic solvents, e.g. lower alcohols, esters or ketones
- A61K2236/333—Extraction of the material involving extraction with hydrophilic solvents, e.g. lower alcohols, esters or ketones using mixed solvents, e.g. 70% EtOH
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K2236/00—Isolation or extraction methods of medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicine
- A61K2236/30—Extraction of the material
- A61K2236/39—Complex extraction schemes, e.g. fractionation or repeated extraction steps
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Abstract
The invention discloses a new application of Yuezui tea or an extract thereof, namely the application of the Yuezui tea or the extract thereof in preparing a medicine or a health product for preventing and/or treating acute liver injury, belonging to the technical field of natural medicine development or medicine; the experimental result shows that the tea made of sparrow tea has a remarkable effect of preventing and treating acute liver injury; the tea is a natural product, has high safety and simple extraction process, is suitable for industrial production and market popularization and application, and provides a new application approach for development and utilization of the tea.
Description
Technical Field
The invention relates to application of sparrow mouth tea or an extract thereof in preparing a medicine or a health-care product for preventing and/or treating acute liver injury, and belongs to the technical field of medicines.
Background
The types of liver diseases vary from mild hepatocellular injury and necrosis to cirrhosis and liver cancer. Excessive drinking, liver viruses, various toxins and drugs are closely related to acute or chronic inflammation of the liver. Acute Liver Injury (ALI) is a disease that seriously affects human health, and the causes of the disease mainly include: viral infection, autoimmune hepatitis, drugs, alcoholism, chemical poisons, and the like. In recent years, liver metabolism has been seriously compromised by increasingly common Drug abuse, leading to a rapid increase in the incidence of Drug Induced Liver Injury (DILI). Acute Drug Induced Liver Injury (DILI) refers to liver injury caused by drugs themselves or metabolites thereof after excessive or improper use of drugs, the course of disease is generally within 3 months, and the course of cholestatic liver injury can exceed 1 year. In the technology, APAP is selected to establish an acute liver injury model, and because the main reason for the acetaminophen to induce the acute liver injury is oxidative stress, the antioxidation effect can be a new means for preventing and treating acetaminophen-induced liver injury.
The tea is prepared from Vaccinium camphorata (Vaccinium camphorata) belonging to Vaccinium of EricaceaeVaccinium dunalianum) The flower bud of the plant is also called as sparrow tongue tea. The vaccinium dunalianum is mostly produced in shrubs, India (northeast), Plumbum preparatium, Xijin, Myanmar (northeast), Vietnam, Guizhou, Sichuan, Yunnan, Tibet, Taiwan and the like in China are the main production areas of the vaccinium dunalianum. In China, people in the sparrow tongue tea planting area take the sparrow mouth tea as a national medicament for treating the rheumatoid arthritis, and take dry flower buds as a local national drink.
At present, no literature report is found on the research on the prevention and treatment of acute liver injury by the tea sparrow mouth extract at home and abroad.
Disclosure of Invention
The invention aims to provide a new application of the sparrow tea and the extract thereof in the aspect of medicine, namely, the sparrow tea or the extract thereof is applied to the preparation of medicines or health care products for preventing and/or treating acute liver injury.
The invention prevents or/and treats acute liver injury with the maidenhair tea extract as the active ingredient, used for preparing the medicament for preventing and treating acute liver injury, can also add one or more pharmaceutical preparation acceptable supplementary products, or compound with other active ingredients and play the role of synergistically anti-hepatic fibrosis; can be prepared into pharmaceutically suitable dosage forms.
The preparation of the tea extract of the sparrow tea is a conventional method, and the following methods can be adopted but are not limited:
ultrasonically extracting herba Oxalidis Corniculatae with 60-80 vol% methanol water solution for 20-40min, filtering, repeatedly extracting the residue for 1-2 times, mixing filtrates, concentrating, and lyophilizing.
The invention carries out series of researches on the prevention and treatment of acute liver injury by the sparrow tea extract, and proves that the sparrow tea extract has the function of preventing and treating the acute liver injury, so that the sparrow tea extract can be used for preparing and developing the medicine for preventing and treating the acute liver injury.
Compared with the prior art, the invention has the following advantages:
1. the invention explores new medical application of the sparrow mouth tea extract and develops a new research field;
2. experiments show that the tea finch extract can effectively reduce the activity index of acetaminophen (APAP) -induced acute liver injury model mouse tissues under low dose;
3. the prevention and treatment effect of the tea extract is obviously higher than that of a negative control group, and the tea extract is a natural product, has high safety and simple extraction process and has development and application prospects.
Drawings
FIG. 1 is the liver index (mean. + -. variance) of mice for each experimental group; wherein CON is negative control group, APAP is model group, and FTEL is low concentration teaThe concentration group is FTEH which is a high concentration group of the sparrow mouth tea; marker # of significant difference compared to negative control group (c) ((c))p<0.05); the superscript of the significant difference compared to the model group is: (p< 0.05）；
FIG. 2 is the results of ALT content (mean. + -. variance) in serum of mice of each experimental group, in which: marker # of significant difference compared to negative control group (c) ((c))p<0.05); the superscript of the significant difference compared to the model group is: (p< 0.05）；
FIG. 3 is the result of the AST content in serum of mice of each experimental group (mean. + -. variance), in which: marker # of significant difference compared to negative control group (c) ((c))p<0.05); the superscript of the significant difference compared to the model group is: (p< 0.05）；
FIG. 4 is the expression of IL-1 β inflammatory factor in liver tissue (mean. + -. variance), wherein: marker # of significant difference compared to negative control group (c) ((c))p<0.05); the superscript of the significant difference compared to the model group is: (p< 0.05）；
FIG. 5 is the expression of IL-6 inflammatory factor in liver tissue (mean. + -. variance), wherein: marker # of significant difference compared to negative control group (c) ((c))p<0.05); the superscript of the significant difference compared to the model group is: (p< 0.05）；
FIG. 6 is the expression of TNF- α inflammatory factor in liver tissue (mean. + -. variance), wherein: marker # of significant difference compared to negative control group (c) ((c))p<0.05); the superscript of the significant difference compared to the model group is: (p< 0.05）；
FIG. 7 is a H & E staining pattern (H & E X200) of pathological sections of liver tissue from APAP-induced acute liver injury mice and dried prognosis of tea finch (Roque tea) extract, wherein A is a negative control group; b is a model group; c is low dosage group of tea extract of bird's beak; d is high dosage group of extract of tea of bird's beak.
Detailed Description
The invention is further illustrated below with reference to data from experimental examples and pathological section figures. These experimental examples are only for illustrating the present invention and are not intended to limit the scope of application of the present invention. After reading the description of the present invention, those skilled in the art will be able to make various changes, modifications and alterations equivalent to the embodiments of the present invention without specific indications, which are made from conventional commercial products or reagents prepared by conventional methods, and without specific indications, which are made by conventional methods.
Example 1: preparation of extract of Quezui tea
The sparrow mouth tea is collected from Yunnan Wuding, and is naturally dried and then powdered to obtain a sparrow mouth tea sample; weighing 100g of the maidenhair tea sample, performing ultrasonic assisted extraction with 70% methanol aqueous solution by volume concentration at 25 ℃ for 30min according to the ratio of g: mL =1:20, filtering, repeatedly extracting filter residues for 2 times, combining filtrates, concentrating and freeze-drying to obtain the maidenhair tea extract.
Example 2: an aging model is established by applying an acetaminophen (APAP) induction method, and the effect of the tea sparrow extract on preventing acute liver injury of mice is observed through an animal model experiment
1. Establishment and experimental process of acute liver injury model
Male mice of Kunming species (weight 18-22g) were used as study subjects, and each group had 8 mice; after acute modeling for 7 days, evaluating the prevention or delay effect of the tea extract on APAP-induced acute liver injury mice by drenching the tea extract; the mouse is adapted to seven days in a standard environment after being purchased, and is raised according to a conventional raising method, and the specific experimental procedures and groups are as follows:
negative control group: gavage for 7 days, 5mL/kg of physiological saline (0.9%) per day;
model group (APAP group): carrying out intragastric administration with normal saline for 7 days, carrying out intraabdominal injection with 300mg/kg acetaminophen (60 mg/mL) once a day after 1h of the last intragastric administration (fasting is started from night in the sixth day), and carrying out no water inhibition before and after modeling;
low concentration que zui tea group (FTEL): performing intragastric administration of 200mg/kg of the tea extract of the sparrow beak at a dose of 5mL/kg for 7 days continuously, performing intraabdominal injection of 300mg/kg of acetaminophen once a day after the last intragastric administration for 1h for modeling (fasting is started from night of the sixth day), and performing fasting before and after modeling without water inhibition, wherein the extract is diluted by 0.9% physiological saline;
high concentration que zui tea group (FTEH): according to the dose of 5mL/kg, 600mg/kg of the tea extract is continuously infused into the stomach of the sparrow mouth for 7 days, the acetaminophen is injected into the abdominal cavity once a day after the last intragastric infusion for 1h for modeling (the fasting is started from the evening of the sixth day), the water is not forbidden before and after modeling, and the extract is diluted by 0.9% physiological saline;
injecting APAP for 8h, stimulating, collecting blood sample from eyeball, centrifuging at 3500r/min for 10min, collecting upper layer serum, storing in-80 deg.C ultra-low temperature refrigerator, and measuring serum ALT and AST content (mean value + -variance); killing mice by removing necks after blood collection, then collecting liver tissues, washing the liver tissues by using 0.9% cold physiological saline, weighing the mass of the liver tissues by using an electronic balance, and calculating mouse liver indexes (the results are all expressed by mean values +/-variance), wherein the mouse liver indexes (%) = (the weight of the liver tissues/the weight of the mice) multiplied by 100%; another part of liver tissue is crushed by a tissue crusher, 10 percent of tissue homogenate is prepared by normal saline, and relevant inflammatory factor expression determination is respectively carried out (the results are all expressed by mean value +/-variance); fixing liver with 10% neutral formalin, performing H & E staining, performing histopathological analysis, and storing the residual tissue and separated serum in-80 deg.C ultra-low temperature refrigerator. The activity evaluation is carried out by observing the change of mouse liver index, the activity condition in liver tissue and combining with the histopathological analysis of liver.
The determination of the contents of ALT, AST, IL-1 beta, IL-6 and TNF-alpha is carried out by adopting a kit purchased from Nanjing, and the operation is carried out according to the steps of the instruction.
The paraffin section preparation method comprises the following steps:
(1) material taking: fixing fresh liver tissue with 10% formalin solution for more than 24h, taking out liver tissue from the fixing solution, flattening the tissue of a target part in a fume hood with a scalpel, and placing the trimmed tissue and a corresponding label in a dehydration box;
(2) dehydrating and wax dipping: and (5) putting the dehydration box into a dehydration machine for dehydration by gradient alcohol in sequence. 4 hours of 75% alcohol, 2 hours of 85% alcohol, 2 hours of 90% alcohol, 1 hour of 95% alcohol, 30 minutes of absolute ethanol I, 30 minutes of absolute ethanol II, 5-10 minutes of alcohol benzene, 5-10 minutes of xylene I, 5-10 minutes of xylene II, 1 hour of paraffin I melted at 65 ℃, 1 hour of paraffin II melted at 65 ℃, 1h hour of paraffin II melted at 65 ℃ and 1 hour of paraffin III melted at 65 ℃;
(3) embedding: embedding the wax-soaked tissue in an embedding machine. Firstly, putting melted wax into an embedding frame, taking out the tissue from a dehydration box before wax solidification, putting the tissue into the embedding frame according to the requirements of an embedding surface, attaching a corresponding label, freezing and cooling at the temperature of minus 20 ℃, taking out the wax block from the embedding frame after wax solidification, and finishing the wax block;
(4) slicing: placing the trimmed wax block in a paraffin slicer for slicing, wherein the thickness of the wax block is 4 mu m; the slices float on a spreading machine at 40 ℃ warm water to flatten the tissues, the glass slides pick up the tissues, the slices are baked in a 60 ℃ oven, and the slices are taken out and stored at normal temperature for later use after being baked by water, dried and waxed.
HE staining method was as follows:
(1) paraffin section dewaxing to water: sequentially placing the slices into xylene I20 min-xylene II 20 min-absolute ethyl alcohol I5 min-absolute ethyl alcohol II 5min-75% alcohol 5min, and washing with tap water;
(2) hematoxylin staining: staining the slices in hematoxylin staining solution for 3-5min, washing with tap water, differentiating the differentiation solution, washing with tap water, returning blue to the blue solution, and washing with running water;
(3) eosin staining: the slices are dehydrated for 5min respectively by 85 percent and 95 percent gradient alcohol, and are dyed for 5min in eosin dye solution.
(4) Dewatering and sealing: placing the slices in anhydrous ethanol I5 min-anhydrous ethanol II 5 min-anhydrous ethanol III 5 min-xylene I5 min-xylene II 5min for transparency, and sealing with neutral gum;
(5) microscopic examination and image acquisition and analysis.
2. Results
2.1 Effect of tea extract of Quezui on mouse liver index
The effect of the tea finch extract on the liver index of mice is shown in figure 1; compared with a negative control group (CON), the liver index of APAP mice after being injected with 300mg/kg APAP for molding is obviously increased, which indicates that liver injury is caused by APAP, so that the liver is enlarged; the result of the high-concentration sparrow tea group (FTEH) shows that APAP has no obvious influence on the liver index of the mouse, and shows that the liver index can be obviously reduced by the pretreatment of the high-concentration sparrow tea before APAP modeling; the low concentration of sparrow tea pretreatment also reduced the liver index increase caused by APAP compared to the APAP group; the results show that the sparrow tea extract has obvious protective effect on APAP-induced liver injury.
2.2 Effect of the extract of Fujizu tea on ALT and AST in mouse serum
The influence of the tea extract on ALT and AST in mouse serum is shown in FIGS. 2 and 3; compared with CON, the levels of ALT and AST in serum are remarkably increased after 8h of APAP modeling, so that considerable liver injury is caused, which indicates that a liver injury model caused by APAP is successfully established; compared with the APAP group, the mice in the FTEL group can reduce the levels of ALT and AST by being given 200mg/kg of the sparrow mouth tea extract pretreatment; the FTEH group can obviously inhibit the elevation of ALT and AST levels in the serum of the mouse caused by APAP, and can lead the levels of two biochemical indexes to approach the CON group; the above results indicate that the extract of the tea leaves of the bird's beak can significantly protect liver damage caused by APAP.
2.3 Effect of the extract of tea leaves of the Ottelia sparrow on the expression of IL-1 beta, IL-6, TNF-alpha in liver tissue
The results of the effect of the tea finch on the levels of IL-1 beta, IL-6 and TNF-alpha in the liver tissues are shown in FIGS. 4, 5 and 6, and compared with the CON group, the levels of IL-1 beta, IL-6 and TNF-alpha in the liver tissues of APAP mice are obviously increased; compared with the APAP group, the FTEH group mice can obviously inhibit the increase of TNF-alpha, IL-1 beta and IL-6 levels in liver tissues caused by APAP by the pretreatment of 600mg/kg of the tea sparrow tea extract, so that the levels of three inflammatory factors are approximate to the level of the CON group, and the effect of the high-dose group is better than that of the low-dose group; the experimental results show that the tea sparrow extract can prevent and treat acute liver injury of mice caused by APAP and is related to inhibition of inflammatory reaction in liver tissues.
2.4 Effect of tea extract on APAP-induced acute liver injury mouse histopathology
As shown in FIG. 7, when observed under 20-fold microscope, the liver cells in the blank group (FIG. 7A) were arranged radially around the central vein, and were aligned, and the nuclei were large and circular, and were substantially similar, and no significant lesion shrinkage occurred, and the liver tissue structure was normal. The model group (fig. 7B) has obvious pathological changes, and can be observed by a microscope to find that the structure of the liver cells is damaged, the staining of the cell sap is deepened, the cell membranes are cracked and dissolved, and the phenomena of bleeding, extravasated blood, necrosis and the like are obvious; the acetaminophen causes the mouse liver cells to have vacuole and focal necrosis, nucleus shrinkage, aggregation necrosis and more cytoplasm cavities, and the phenomena show that the liver injury molding is successful due to the APAP of 300 mg/kg. Local regional hepatocyte necrosis, most of the lobular structure of the liver was intact, and cells near hepatic veins began to repair were seen in the low concentration group of the tea finch (fig. 7C). The liver tissue result which is obvious in the high-concentration group (figure 7D) of the extract is not damaged when not observed, which shows that the extract can inhibit the APAP from damaging the liver tissue structure of a mouse, so that the liver tissue structure approaches the condition of a normal group of mice, inhibit the chemotaxis of inflammatory cells to the damaged part, inhibit the inflammatory reaction, improve the permeability of a liver cell membrane and relieve the edema of the liver tissue; the results show that the tea finch extract can effectively prevent acute liver injury of mice caused by APAP and protect liver tissues from hepatic veins in a radial manner.
In summary, the following steps: the tea extract of the sparrow mouth can effectively reduce the liver index, lower the ALT and AST levels in serum and obviously improve the pathological changes of liver tissues, thereby showing that the tea extract of the sparrow mouth has obvious protective effect on the acute liver injury of mice caused by APAP.
Claims (2)
1. Application of the tea leaves or the extract thereof in preparing medicines for preventing and/or treating acute liver injury.
2. Application of the tea or the extract thereof in preparing health care products for preventing and/or treating acute liver injury.
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Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| AU2015101749A4 (en) * | 2015-12-03 | 2016-01-14 | Macau University Of Science And Technology | Method of treating or preventing liver injury induced by acetaminophen |
| CN106722608A (en) * | 2016-12-07 | 2017-05-31 | 贵州大学 | A kind of blue fragrant sparrow tongue tea jelly |
-
2020
- 2020-08-23 CN CN202010853377.7A patent/CN111939190A/en active Pending
Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| AU2015101749A4 (en) * | 2015-12-03 | 2016-01-14 | Macau University Of Science And Technology | Method of treating or preventing liver injury induced by acetaminophen |
| CN106722608A (en) * | 2016-12-07 | 2017-05-31 | 贵州大学 | A kind of blue fragrant sparrow tongue tea jelly |
Non-Patent Citations (1)
| Title |
|---|
| 李会瑞: "乙醇浸提苦荞茶和雀嘴茶中的总黄酮及羟自由基清除活性探究", 《河南师范大学学报》 * |
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