Application of phyllanthus emblica extract in preparation of pharmaceutical composition for improving mitochondrial activity in liver

Abstract

The invention relates to preparation of a pharmaceutical composition for repairing liver injury, in particular to application of a pharmaceutical composition for repairing liver injury by utilizing an emblic leafflower fruit extract. The invention relates to an application of a phyllanthus emblica extract in preparation of a pharmaceutical composition for repairing liver injury. The pharmaceutical composition containing the phyllanthus emblica extract improves the ability of mitochondria in liver cells in contact with free radicals to perform oxidative phosphorylation reaction and adenosine triphosphate (ATP) synthesis. Furthermore, the pharmaceutical composition containing the emblic leafflower fruit extract reduces hydrogen ion leakage of mitochondria, improves the maximum oxygen consumption capacity of the mitochondria, improves the adenosine triphosphate coupling efficiency of the mitochondria, improves the pre-storage oxygen consumption capacity of the mitochondria, and reduces the generation amount of free radicals in liver cells.

Description

[0001] This case is application number: 201810545518.1, application date: May 25, 2018, invention name:

[0002] A divisional application for the use of an extract of Phyllanthus emblica for preparing a pharmaceutical composition for increasing mitochondrial activity in the liver. Technical Field

[0003] The present invention relates to the preparation of a pharmaceutical composition for repairing liver damage, and in particular to the use of an extract of emblica officinalis to prepare a pharmaceutical composition for repairing liver damage. Background Art

[0004] The liver is one of the main metabolic organs in the human body. The liver's work includes metabolizing carbohydrates, proteins and lipids, as well as breaking down, converting and eliminating toxins in the body. However, in the process of the liver's work, especially in breaking down, converting and eliminating toxins in the body, free radicals are also produced as byproducts. Furthermore, for a liver damaged by disease, when its liver function is affected by the disease, the amount of free radicals produced when the liver is working will also increase.

[0005] For liver cells, free radicals have strong oxidative ability, which greatly increases the chances of liver cells and organelles in liver cells being damaged by oxidation. In particular, mitochondria are organelles in liver cells that perform oxidative phosphorylation and synthesize adenosine triphosphate (ATP). When mitochondria are damaged by oxidative damage caused by a large number of free radicals, liver cells cannot obtain sufficient energy supply from the inactive mitochondria. As a result, the inactive mitochondria will have a serious negative impact on the working condition of liver cells, and then have a negative impact on the function of the liver. Summary of the invention

[0006] The present invention provides a use of an extract of Phyllanthus emblica to prepare a pharmaceutical composition for repairing liver damage, thereby enhancing the activity of mitochondria in hepatocytes of the liver, thereby maintaining the normal function of the hepatocytes and repairing liver damage.

[0007] The present invention discloses a use of an extract of Phyllanthus emblica for preparing a pharmaceutical composition for repairing liver damage, wherein the pharmaceutical composition comprising the extract of Phyllanthus emblica enhances the ability of mitochondria in liver cells contacted with free radicals to perform oxidative phosphorylation and synthesize adenosine triphosphate (ATP).

[0008] According to the use of the extract of Phyllanthus emblica for preparing a pharmaceutical composition for repairing liver damage disclosed in the present invention, the pharmaceutical composition containing the extract of Phyllanthus emblica enhances the ability of mitochondria in liver cells that are in contact with free radicals to perform oxidative phosphorylation reactions and synthesize adenosine triphosphate (ATP). In this way, the activity of mitochondria in liver cells is maintained, and the mitochondria in liver cells can produce sufficient adenosine triphosphate for use by liver cells, thereby maintaining normal metabolic work of liver cells. Furthermore, since the efficiency of adenosine triphosphate synthesis required to repair cell damage is improved, damaged liver cells obtain sufficient energy required for repair, thereby accelerating the repair of the liver to a normal state.

[0009] The above description of the disclosed content and the following description of the implementation modes are used to demonstrate and explain the spirit and principle of the present invention, and to provide a further explanation of the scope of the patent application of the present invention. BRIEF DESCRIPTION OF THE DRAWINGS

[0010] Figure 1 This is the infrared absorption spectrum of the Phyllanthus emblica extract used in Examples 1 to 3 of the present invention.

[0011] Figure 2 The HPLC spectrum of the Phyllanthus emblica extract used in Examples 1 to 3 of the present invention is shown.

[0012] Figure 3 Schematic diagram of oxygen consumption for synthesizing adenosine triphosphate in Examples 1 to 3 of the present invention, Comparative Examples and Control Examples.

[0013] Figure 4 Schematic diagram of oxygen consumption for overcoming hydrogen ion leakage in Embodiments 1 to 3 of the present invention, a comparative example and a control example.

[0014] Figure 5 It is a schematic diagram of oxygen consumption of the maximum oxygen consumption capacity of Examples 1 to 3 of the present invention, a comparative example and a control example.

[0015] Figure 6 It is a schematic diagram of coupling efficiency of Embodiments 1 to 3 of the present invention, a comparative example and a control example.

[0016] Figure 7 It is a schematic diagram of the pre-stored oxygen consumption capacity of Examples 1 to 3 of the present invention, a comparative example and a control example.

[0017] Figure 8 Schematic diagram of the amount of free radical generation of hepatocytes induced by oleic acid in Example 2, Example 3, Comparative Example and Control Example of the present invention. DETAILED DESCRIPTION

[0018] The technical solutions provided by the present invention are described in detail below in conjunction with the embodiments, but they should not be construed as limiting the protection scope of the present invention.

[0019] Example 1

[0020] The detailed features and advantages of the present invention are described in detail in the following embodiments, and the contents are sufficient to enable any person skilled in the art to understand the technical content of the present invention and implement it accordingly, and according to the contents disclosed in this specification, the claims and the drawings, any person skilled in the art can easily understand the relevant purposes and advantages of the present invention. The following examples are used to further illustrate the viewpoints of the present invention in detail, but are not intended to limit the scope of the present invention in any way.

[0021] Emblica officinale, also known as pomelo, oil orange, Amalaka, Pokok Melaka, Indian Gooseberry, is a deciduous sub-tree belonging to the genus Emblica in the Euphorbiaceae family. It is distributed from India to Malaysia and southern China, and is generally considered to be native to India.

[0022] The method of obtaining the emblica extract used in the present invention is, for example, to extract the emblica fruit using carbon dioxide as a supercritical fluid, or to extract the emblica fruit using methanol, ethanol, acetone, ethyl acetate, 0.1wt% to 5wt% sodium chloride aqueous solution, potassium chloride aqueous solution, calcium chloride aqueous solution, magnesium chloride aqueous solution, or 0.1wt% to 5wt% sodium chloride ethanol solution, potassium chloride ethanol solution, calcium chloride ethanol solution, magnesium chloride ethanol solution as a solvent to obtain a primary extract; then, the primary extract is filtered and purified to obtain the emblica extract used in the present invention. The emblica extract can be dried using spray drying or vacuum drying to obtain an emblica extract powder that is easy to store.

[0023] The extract of Phyllanthus emblica contains 35wt% to 55wt% of a mixture of Emblicanin-A and Emblicanin-B, 4wt% to 15wt% of Punigluconin, 10wt% to 20wt% of Pedunculagin, 5wt% to 15wt% of Rutin and 10wt% to 30wt% of Gallo-ellagitannoids. The infrared absorption spectrum of the extract of Phyllanthus emblica is 3403.6±5 cm. -1 (cm -1 )、2931.6±5cm -1 (cm -1)、1385.0±5cm -1 (cm -1 )、1318.6±5cm -1 (cm -1 )、1623.5±5cm -1 (cm -1 )、1451.3±5cm -1 (cm -1 )、1352.1±5 cm -1 (cm -1 )、1218.4±5cm -1 (cm -1 )、1148.6±5cm -1 (cm -1 )、1035.7±5cm -1 (cm -1 )、3403.6±5cm -1 (cm -1 ) has characteristic absorption peaks. The HPLC Chromatogram of the extract of Phyllanthus emblica has characteristic peak signals at 1.620±0.5 minutes, 2.148±0.5 minutes, 3.265±0.5 minutes and 4.370±0.5 minutes.

[0024] The structure of Emblicanin-A (2,3-di-O-galloyl-4,6-(S)-hexahydroxydiphenoyl-2-keto-glucono-lactone) is shown in Formula 1.

[0025] Formula 1

[0026]

[0027] The structure of Emblicanin-B (2,3,4,6-bis-(S)-hexahydroxydiphenoyl-2-keto-glucono-lactone) is shown in Formula 2.

[0028] Formula 2

[0029]

[0030] The structure of Punigluconin (2,3-di-O-galloyl-4,6-(S)-hexahydroxydiphenoylgluconicacid) is shown in Formula 3.

[0031] Formula 3

[0032]

[0033] The structure of aristolochicine (2,3,4,6-bis-(S)-hexahydroxydiphenoyl-D-glucose) is shown in Formula 4.

[0034] Formula 4

[0035]

[0036] Rutin (3',4',5,7-tetrahydroxyflavone -1 ,3-O-rhamnoside, 3',4',5,7-tetrahydroxyflavono -1 ,3-O-rhamnoglucoside) has a structure as shown in Formula 5.

[0037] Formula 5

[0038]

[0039] When an aqueous solution of 100 to 500 micrograms per milliliter (μg / ml) of the emblica extract is provided to liver cells in contact with free radicals, the emblica extract entering the liver cells increases the activity of mitochondria in the liver cells. As a result, the amount of adenosine triphosphate synthesized by the oxidative phosphorylation reaction of the mitochondria activated by the emblica extract increases, the maximum oxygen consumption capacity increases, the pre-stored oxygen consumption capacity increases, the adenosine triphosphate coupling efficiency increases, and the hydrogen ion leakage decreases.

[0040] Furthermore, when an aqueous solution of 100 to 500 μg / ml of the extract of Phyllanthus emblica was provided to the hepatocytes in contact with free radicals, the amount of adenosine triphosphate synthesized by the oxidative phosphorylation reaction of the mitochondria activated by the extract of Phyllanthus emblica did not decrease significantly, the coupling efficiency of adenosine triphosphate did not decrease significantly, and the leakage of hydrogen ions did not increase significantly. The maximum oxygen consumption capacity of the mitochondria activated by the extract of Phyllanthus emblica increased slightly, and the pre-stored oxygen consumption capacity of the mitochondria increased slightly. Therefore, the extract of Phyllanthus emblica can be used as an active ingredient of a pharmaceutical composition for repairing liver damage, and the extract of Phyllanthus emblica can maintain the activity of mitochondria in the hepatocytes in contact with free radicals at a level close to that of mitochondria in the hepatocytes not in contact with free radicals. Furthermore, since the efficiency and ability of adenosine triphosphate synthesis required for repairing cell damage are improved, the damaged hepatocytes obtain sufficient energy required for repair, thereby accelerating the repair of the liver to a normal state.

[0041] The method of providing the amla extract or the pharmaceutical composition comprising the amla extract to the liver cells is, for example, to orally ingest the amla extract or the pharmaceutical composition comprising the amla extract. When the amla extract is provided to the cells in an edible manner, the effective dose of the amla extract is 1.081 grams (g) to 5.405 grams (g). The conversion formula is as follows: effective dose for human body (mg) = effective dose for cell experiment (μg / ml) × mouse body weight (g) × conversion coefficient × human body weight (kg). The conversion coefficient is obtained by looking up the conversion coefficient table of dose per kilogram of body weight for animals and humans. When the mouse weight is 20g and the human body weight is 60 kilograms, the conversion coefficient is 9.01.

[0042] In order to facilitate oral ingestion of the emblica extract, the emblica extract can be prepared into a processed emblica extract product in the form of liquid, solid, granular, powder, paste or gel. The processed emblica extract product can be combined with an excipient or flavoring agent as an additive to enhance the flavor and facilitate consumption.

[0043] Excipients include, for example, starches such as wheat starch, rice starch, corn starch, potato starch, dextrin, and cyclodextrin; crystalline celluloses; sugars such as lactose, glucose, granulated sugar, reduced maltose, maltose, fructooligosaccharides, and emulsified oligosaccharides; and sugar alcohols such as sorbitol, erythritol, xylitol, lactitol, and mannitol.

[0044] Flavoring agents include, for example, various fruit juice extracts such as longan extract, lychee extract, and grapefruit extract; various fruit juices such as apple juice, orange juice, and lemon juice; various flavors such as peach flavor, plum flavor, and yogurt flavor; various sweeteners such as acesulfame potassium, sucralose, erythritol, oligosaccharides, mannose, xylitol, and isomerized sugars; various acidulants such as citric acid, malic acid, tartaric acid, and gluconic acid; various tea ingredients such as green tea, oolong tea, Banaba tea, Eucommia tea, Tieguanyin tea, Coix tea, Seven Leaf Gallbladder Tea, Wild Rice Tea, and Kelp tea; various coffee ingredients such as Arabica, Robusta, and Liberica, etc.

[0045] Furthermore, the emblica extract or the pharmaceutical composition comprising the emblica extract can also be coated in a capsule to facilitate oral ingestion of the emblica extract. The emblica extract or the pharmaceutical composition comprising the emblica extract can be coated in a hard capsule in the form of a dry powder. The emblica extract or the pharmaceutical composition comprising the emblica extract can also be coated in a soft capsule in the form of a solution, a suspension, a paste, a powder or a granule.

[0046] The oils and fats used to dissolve or disperse the emblica extract in the soft capsule are, for example, avocado oil, almond oil, linseed oil, fennel oil, perilla oil, olive oil, olive squalene, sweet orange oil, orange roughy oil, sesame oil, garlic oil, cocoa butter, pumpkin seed oil, chamomile oil, carrot oil, cucumber oil, shea fatty acid, macadamia nut oil, cranberry oil, brown rice germ oil, rice oil, wheat germ oil, safflower oil, shea oil, liquid shea oil, perilla oil, soybean oil, evening primrose oil, camellia oil, corn oil, rapeseed oil, saw palmetto extract oil, coix seed oil, peach kernel oil, celery seed oil, castor oil, sunflower seed oil, grape seed oil, borage oil, macadamia nut oil, meadowfoam oil, oil), cottonseed oil, peanut oil, tortoise oil, mink oil, egg yolk oil, fish oil, palm oil, palm kernel oil, wood wax, coconut oil, long-chain / medium-chain / short-chain fatty acid triglycerides, diglycerides, tallow, lard, squalene, squalane, pristane, and hydrogenated products of these oils, etc. Among them, borage oil and evening primrose oil contain a large amount of gamma-linolenic acid (GLA), which is an essential fatty acid for the human body. It has the functions of moisturizing, promoting cell regeneration, and increasing the activity of brown fat to promote fat burning.

[0047] In addition, colorants, preservatives, viscosity enhancers, binders, disintegrants, dispersants, stabilizers, gelling agents, antioxidants, surfactants, preservatives, pH adjusters and other additives that meet the requirements may also be added to the processed products of the emblica extract according to the prescribed dosage standards and processing and production requirements.

[0048] The following examples 1 to 3, control examples and comparative examples of the present invention illustrate the use of the emblica extract disclosed in the present invention for preparing a pharmaceutical composition for repairing liver damage, and experimental tests are performed to illustrate the efficacy of the emblica extract disclosed in the present invention for preparing a pharmaceutical composition for repairing liver damage.

[0049] The extract of the emblica officinalis used in the experiment of the present invention is the emblica officinalis extract powder obtained by soaking the fruit of the emblica officinalis in a 1wt% sodium chloride aqueous solution, heating it in a steam bath at 65°C to 75°C for one hour, filtering it, freezing it for 3 days, and then spray drying it. The emblica officinalis extract after drying is in the form of brown powder. The emblica officinalis extract contains 27wt% of Emblicanin-A, 23wt% of Emblicanin-B, 8wt% of Punigluconin, 14wt% of Long-pedunculate Aristolochicin, 10wt% of Rutin and 10wt% to 30wt% of Gallo-ellagitannoids. The extract of the emblica officinalis of the present invention is not limited to the extract of Emblica Officinalis, and other extracts of the emblica officinalis with different scientific names but similar components also have the same effect.

[0050] Figure 1 This is the infrared absorption spectrum of the Phyllanthus emblica extract used in Examples 1 to 3 of the present invention. Figure 2 The HPLC spectrum used in Examples 1 to 3 of the present invention is shown in FIG. Figure 1 As shown, the infrared absorption spectrum of Phyllanthus emblica extract is at 3403.6 cm -1 (cm -1 ), 2931.6 cm -1 (cm -1 ), 1385.0 cm -1 (cm -1 ), 1318.6 cm -1 (cm -1 ), 1623.5 cm -1 (cm -1 )、1451.3 cm -1 (cm -1 ), 1352.1 cm-1 (cm -1 ), 1218.4 cm -1 (cm -1 )、1148.6 cm -1 (cm -1 )、1035.7 cm -1 (cm -1 )、3403.6 cm -1 (cm -1 ) has characteristic absorption peaks. Figure 2 As shown, the HPLC Chromatogram of the Phyllanthus emblica extract has characteristic peak signals at 1.620 minutes, 2.148 minutes, 3.265 minutes and 4.370 minutes.

[0051] The cells used in the experiment are hepatocytes (Hep-G2 cells). The experimental sample preparation method is to inoculate 20,000 hepatocytes in each well of a 24-well culture plate and culture them for 24 hours. Then, the culture medium in the well is removed, and the hepatocytes in the well are treated according to the conditions of each embodiment, control example and comparative example.

[0052] During the experiment, when preparing the experimental samples of Examples 1 to 3, first, 0.001 ml of an aqueous solution of the emblica extract of a predetermined concentration was added to the well where the hepatocytes were located and incubated for 24 hours; then, the aqueous solution in the well where the hepatocytes were located was removed, and a 1.5 mM H2O2 aqueous solution was added to the well, and the cells were incubated in the 1.5 mM H2O2 aqueous solution for 60 minutes. Then, the H2O2 aqueous solution in the well where the hepatocytes were located was removed, and the hepatocytes were washed with phosphate buffered saline (PBS), completing the preparation of the experimental samples of Examples 1 to 3.

[0053] When preparing the experimental sample of the control example, after removing the culture medium in the well where the hepatocytes are located, the remaining hepatocytes are the experimental samples of the control example. When preparing the experimental sample of the comparative example, firstly, a 1.5mM H2O2 aqueous solution is added to the well, and the hepatocytes are incubated in the 1.5mM H2O2 aqueous solution for 60 minutes. Then, the H2O2 aqueous solution in the well where the hepatocytes are located is removed, and the cells are washed with phosphate buffered saline (PBS), and the experimental sample preparation of the comparative example is completed.

[0054] In Example 1, the concentration of the aqueous solution of the extract of Phyllanthus emblica was 200 micrograms per milliliter (μg / ml). In Example 2, the concentration of the aqueous solution of the extract of Phyllanthus emblica was 250 micrograms per milliliter (μg / ml). In Example 3, the concentration of the aqueous solution of the extract of Phyllanthus emblica was 500 micrograms per milliliter (μg / ml).

[0055] Next, the oxygen consumption of the liver cells in Examples 1 to 3, the control example, and the comparative example in the well was measured using a seahorse bioenergetic meter. According to literature, treating cells with hydrogen peroxide can simulate the condition of free radicals produced by oxidation in cells, thereby studying the mitochondrial activity of cells under oxidative stress. Therefore, after the liver cells in Examples 1 to 3 and the comparative example were treated with hydrogen peroxide, the oxygen consumption of the liver cells was measured using a seahorse bioenergetic meter, and the effect of the extract of Phyllanthus emblica on the activity of mitochondria in liver cells was understood.

[0056] The measurement principle and process of the Hippocampal Bioenergetic Meter are as follows: First, monitor the basal oxygen consumption of the cells in the well. Then, add an ATP synthase inhibitor to inhibit the production of ATP by mitochondria. The reduced oxygen consumption at this time is the oxygen consumption of the mitochondria for oxidative phosphorylation to synthesize ATP, that is, the basal oxygen consumption of mitochondria (Basal Respiration). An ATP synthase inhibitor is, for example, oligomycin. Then, add an anti-coupling agent of appropriate concentration to allow the mitochondria to idle at the limit without destroying the electron transport chain of the mitochondrial inner membrane to evaluate the maximum oxygen consumption capacity of the mitochondria (Maximal Respiration). An anti-coupling agent is, for example, Carbonylcyanide-4-(trifluoromethoxy)phenylhydrazone (FCCP). Finally, add an electron transport chain inhibitor to completely shut down the oxygen consumption of the mitochondria, thereby confirming the background value of the measurement, that is, the non-mitochondrial oxygen consumption (Non-mitochondrial Respiration). An electron transport chain inhibitor is, for example, a combination of rotenone and antimycin A.

[0057] The basal oxygen consumption of mitochondria is equal to the basal oxygen consumption of cells minus the non-mitochondrial oxygen consumption. The basal oxygen consumption of mitochondria minus the oxygen consumed to synthesize ATP is equal to the oxygen consumption to overcome proton leakage. The maximum oxygen consumption capacity of mitochondria minus the basal oxygen consumption of mitochondria is equal to the spare respiratory capacity of mitochondria. The coupling efficiency of mitochondria for ATP is equal to the oxygen consumption for synthesizing ATP divided by the basal oxygen consumption of mitochondria.

[0058] The experimental parameters and experimental measurement results of Examples 1 to 3, Comparative Examples 1 to 2 and the control example are shown in Table 1 and Table 2. Figures 3 to 7 The experimental measurements of picomoles of oxygen consumed per microgram of protein per minute (pmol / min / μg protein) presented in Table 1 are experimental measurements that have been normalized to the amount of cells. Figure 3 Schematic diagram of oxygen consumption for synthesizing adenosine triphosphate in Examples 1 to 3 of the present invention, Comparative Examples and Control Examples. Figure 4 Schematic diagram of oxygen consumption for overcoming hydrogen ion leakage in Embodiments 1 to 3 of the present invention, a comparative example and a control example. Figure 5 It is a schematic diagram of oxygen consumption of the maximum oxygen consumption capacity of Examples 1 to 3 of the present invention, a comparative example and a control example. Figure 6 It is a schematic diagram of coupling efficiency of Embodiments 1 to 3 of the present invention, a comparative example and a control example. Figure 7 It is a schematic diagram of the pre-stored oxygen consumption capacity of Examples 1 to 3 of the present invention, a comparative example and a control example. Figure 8 Schematic diagram of the amount of free radical generation of hepatocytes induced by oleic acid in Example 2, Example 3, Comparative Example and Control Example of the present invention.

[0059] Table 1

[0060]

[0061] like Figure 3 As shown, the oxygen consumption of synthesizing adenosine triphosphate in Examples 1 to 3 is higher than that of synthesizing adenosine triphosphate in the comparative example. Figure 5 As shown, the maximum oxygen consumption capacity of the mitochondria in Examples 1 to 3 is higher than the maximum oxygen consumption capacity of the mitochondria in the comparative example. Figure 6 As shown, the coupling efficiency of the mitochondria in Examples 1 to 3 is higher than that of the mitochondria in the comparative example. Figure 7 As shown in FIG. 1 , the pre-stored oxygen consumption capacity of the mitochondria in Examples 1 to 3 is higher than the pre-stored oxygen consumption capacity of the mitochondria in Comparative Example 1. Figure 3 , Figures 5 to 7 It can be seen that after the hepatocytes of Examples 1 to 3 were treated with the extract of Phyllanthus emblica, the activity of mitochondria in the hepatocytes was enhanced. Furthermore, the increase in the pre-stored oxygen consumption capacity of mitochondria also represents the improvement of the resilience of mitochondria and hepatocytes to various conditions that may cause stress to the cells.

[0062] like Figure 4 As shown in FIG. 1 , the hydrogen ion leakage of the mitochondria of Examples 1 to 3 is lower than that of the mitochondria of the comparative example. Figure 4It can be seen that after the hepatocytes of Examples 1 to 3 were treated with the extract of Phyllanthus emblica, the ability of the mitochondria in the hepatocytes to resist the oxidative damage caused by free radicals was enhanced.

[0063] Furthermore, if Figures 3 to 7 As shown, compared with the mitochondria in the hepatocytes of the control example, the amount of adenosine triphosphate synthesized by the oxidative phosphorylation reaction of the mitochondria activated by the emblica extract in the hepatocytes of Examples 1 to 3 did not decrease significantly, the adenosine triphosphate coupling efficiency did not decrease significantly, and the hydrogen ion leakage did not increase significantly. The maximum oxygen consumption capacity of the mitochondria activated by the emblica extract increased slightly, and the pre-stored oxygen consumption capacity of the mitochondria increased slightly. Therefore, the emblica extract can maintain the activity of the mitochondria in the hepatocytes in contact with free radicals at a level close to the activity of the mitochondria in the hepatocytes not in contact with free radicals.

[0064] Furthermore, if Figure 3 As shown, the oxygen consumption of the synthesis of ATP in Examples 1 to 3 is close to that of the control example and significantly higher than that of the comparative example. Figure 4 As shown in FIG. 1 , the oxygen consumption for overcoming hydrogen ion leakage in Examples 1 to 3 is slightly higher than that in the control example, but significantly lower than that in the comparative example. Figure 3 and Figure 4 It can be seen that the amount of hydrogen ion leakage from the mitochondrial inner membrane of Examples 1 to 3 did not increase as much as that of the mitochondrial inner membrane of the comparative example, which means that there was no serious damage on the mitochondrial inner membrane that caused a large amount of hydrogen ion leakage. Therefore, the amount of oxygen consumed by the mitochondria to transport hydrogen ions to the membrane space again was reduced, so that the oxygen consumption for synthesizing ATP and the ATP coupling efficiency were close to those of the control example.

[0065] like Figure 8 As shown, taking the free radical generation amount in the liver cells of the control example as the benchmark, the free radical generation rate in the liver cells of Examples 2 and 3 is lower than the free radical generation rate in the liver cells of the comparative example, which indicates that the free radical generation amount in the liver cells of Examples 2 and 3 is lower than the free radical generation amount in the liver cells of the comparative example 1. Figure 8 It can be seen that the higher the concentration of the emblica extract provided to the liver cells, the better the effect of inhibiting the generation of free radicals in the liver cells. The liver cells treated with an aqueous solution of the emblica extract at a concentration of 100 to 500 micrograms per milliliter (μg / ml) are protected by the emblica extract, which reduces the damage of the free radicals in the liver cells to the mitochondrial inner membrane in the liver cells. Therefore, the aqueous solution of the emblica extract at a concentration of 100 to 500 micrograms per milliliter (μg / ml) has the effect of protecting the mitochondria in the liver cells. In this way, the time for the collapse of the mitochondria can be delayed, thereby delaying the occurrence of liver cell apoptosis.

[0066] According to the use of the extract of Phyllanthus emblica for preparing a pharmaceutical composition for repairing liver damage disclosed in the present invention, the pharmaceutical composition containing the extract of Phyllanthus emblica enhances the ability of mitochondria in liver cells that are in contact with free radicals to perform oxidative phosphorylation reactions and synthesize adenosine triphosphate (ATP). In this way, the activity of mitochondria in liver cells is maintained, and the mitochondria in liver cells can produce sufficient adenosine triphosphate for liver cells to use, thereby maintaining normal metabolic work of liver cells. Since the efficiency and ability of adenosine triphosphate synthesis required to repair cell damage are improved, damaged liver cells are fully equipped with the energy required for repair, thereby accelerating the repair of the liver to a normal state.

[0067] Furthermore, providing the hepatocytes with the extract of Phyllanthus emblica reduces the damage of the free radicals in the hepatocytes to the inner membrane of the mitochondria in the hepatocytes. Therefore, the aqueous solution of the extract of Phyllanthus emblica also has the effect of protecting the mitochondria in the hepatocytes. In this way, the time for the collapse of the mitochondria can be delayed, thereby delaying the occurrence of apoptosis of the hepatocytes.

[0068]

Explanation of symbols

[0069] none.

[0070] The above is only a preferred embodiment of the present invention. It should be pointed out that for ordinary technicians in this technical field, several improvements and modifications can be made without departing from the principle of the present invention. These improvements and modifications should also be regarded as the scope of protection of the present invention.

Claims

1. Use of an extract of Phyllanthus emblica in preparing any of the following pharmaceutical compositions, a. A pharmaceutical composition for enhancing mitochondrial activity in hepatocytes; b. A pharmaceutical composition for increasing the maximum oxygen consumption capacity of mitochondria; c. A pharmaceutical composition for increasing the pre-stored oxygen consumption capacity of mitochondria; d. A pharmaceutical composition for reducing mitochondrial hydrogen ion leakage and improving mitochondrial adenosine triphosphate coupling efficiency; The concentration of the Phyllanthus emblica extract in the pharmaceutical composition is 200 μg to 500 μg per milliliter; Improving the mitochondrial activity in the liver cells is: improving the ability of multiple mitochondria in multiple liver cells in contact with free radicals to perform oxidative phosphorylation reactions and synthesize adenosine triphosphate; in, The extract of Phyllanthus emblica contains 27 wt % of Emblicanin-A, 23 wt % of Emblicanin-B, 8 wt % of Punigluconin, 14 wt % of Aristolochicine, 10 wt % of Rutin and 10 wt % to 30 wt % of Gallo-ellagitannoids.

2. The use according to claim 1, wherein The infrared absorption spectrum of the emblica extract has a characteristic absorption peak at: 3403.6±5cm -1 、2931.6±5cm -1 、1385.0±5cm -1 、1318.6±5cm -1 、1623.5±5cm -1 、1451.3±5cm -1 、1352.1±5cm -1 、1218.4±5cm -1 、1148.6±5cm -1 、1035.7±5cm -1 、3403.6±5cm -1 .

3. The use according to claim 1, wherein The HPLC spectrum of the emblica extract has characteristic peak signals at 1.620±0.5 minutes, 2.148±0.5 minutes, 3.265±0.5 minutes and 4.370±0.5 minutes.

4. The use according to claim 1, wherein The emblica extract is obtained by extracting emblica fruits using saline as a solvent.

5. The use according to claim 1, wherein The pharmaceutical composition is in oral dosage form.

6. The use according to claim 1, wherein The effective dosage of the emblica extract is 1.081 grams to 5.405 grams.

7. The use according to claim 6, wherein The pharmaceutical composition is encapsulated in a soft capsule or a hard capsule.

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