TW201914589A - Use of a composition in manufacturing a medicament for protecting and repairing mitochondria - Google Patents

Abstract

A use of a composition in manufacturing a medicament for protecting and repairing mitochondria. The composition comprises Emblicanin-A and Emblicanin-B.

Description

Use of a composition for preparing a preparation for protecting and repairing mitochondria

The present invention relates to the use of a composition for the protection and repair of mitochondria, and in particular to a preparation comprising a composition of Emblicanin-A and Emblicanin-B for the preparation of a preparation for protecting and repairing mitochondria.

Mitochondria is the main site for intracellular oxidative phosphorylation and synthesis of adenosine triphosphate (ATP). Since adenosine triphosphate is the energy source of cellular activity, the mitochondria is also known as the "cell energy factory." In addition to providing energy to cells, mitochondria are involved in cell differentiation, cell signaling, and apoptosis, and have the ability to regulate cell growth cycles.

However, some of the by-products produced by the mitochondria during the oxidative phosphorylation reaction are detrimental to the inner membrane of the mitochondria. Long-term accumulation, severely damaged mitochondrial inner membrane will trigger mitochondrial disintegration, which in turn triggers apoptosis. Therefore, how to protect and repair mitochondria to slow the apoptosis triggered by mitochondrial disintegration has become an important issue.

The present invention provides a use of a composition comprising Emblicanin-A and Emblicanin-B for the preparation of a protective mitochondria, which can delay the rate of apoptosis triggered by mitochondrial disintegration by administering the preparation.

The present invention discloses the use of a composition for the preparation of a formulation for protecting and repairing mitochondria, the composition comprising Emblicanin-A and Emblicanin-B.

According to the above-mentioned invention, the composition comprising Emblicanin-A and Emblicanin-B is used for preparing a preparation for protecting and repairing mitochondria, and when the preparation is supplied to cells, Emblicanin-A and Emblicanin-B can be protected. Repair the inner membrane of the mitochondria to delay the disintegration of the mitochondria. In this way, the rate at which mitochondrial disintegration triggers apoptosis can be slowed down.

The above description of the disclosure and the following description of the embodiments of the present invention are intended to illustrate and explain the spirit and principles of the invention, and to provide further explanation of the scope of the invention.

The detailed features and advantages of the present invention are set forth in the Detailed Description of the Detailed Description of the <RTIgt; </ RTI> <RTIgt; </ RTI> </ RTI> </ RTI> <RTIgt; The objects and advantages associated with the present invention can be readily understood by those skilled in the art. The following examples are intended to describe the present invention in further detail, but are not intended to limit the scope of the invention.

The Emblicanin-A and Emblicanin-B used in the present invention are obtained by extracting Phyllanthus Emblica or Emblica Officinale. Yu Ganzi, also known as Yuyouzi, citrus, Amalaka, Pokok Melaka, Indian Gooseberry, belongs to the deciduous sub-arbor of Emblica. From India to Malaysia and southern China, India is generally considered to be the country of origin.

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

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

The method for obtaining Emblicanin-A and Emblicanin-B used in the present invention is, for example, extracting the fruit of Phyllanthus emblica with carbon dioxide as a supercritical fluid, or using methanol, ethanol, acetone, ethyl acetate, aqueous sodium chloride solution, potassium chloride aqueous solution, chlorine The calcium extract solution, the magnesium chloride aqueous solution, the sodium chloride ethanol solution, the potassium chloride ethanol solution, the calcium chloride ethanol solution or the magnesium chloride ethanol solution is used as a solvent to extract the melon fruit to obtain a preliminary extract. The weight percentage concentration of the aqueous sodium chloride solution, the aqueous potassium chloride solution, the aqueous calcium chloride solution or the aqueous magnesium chloride solution is, for example, 0.1 to 5%. The weight percent concentration of the sodium chloride ethanol solution, the potassium chloride ethanol solution, the calcium chloride ethanol solution or the magnesium chloride ethanol solution is, for example, 0.1 to 5%. Next, the primary extract was filtered and purified to obtain Emblicanin-A and Emblicanin-B used in the present invention.

In the preparation of the examples of the present invention, the weight ratio of Emblicanin-A to Emblicanin-B (Emblicanin-A/Emblicanin-B) is not particularly limited. In a formulation of a portion of the present invention, the weight ratio of Emblicanin-A to Emblicanin-B is from 3/2 to 2/3. In a formulation of another embodiment of the invention, the weight ratio of Emblicanin-A to Emblicanin-B is from 6/5 to 5/4. In a formulation of still another embodiment of the invention, the weight ratio of Emblicanin-A to Emblicanin-B is 27/23.

When the total weight concentration of Emblicanin-A and Emblicanin-B is 14 to 22 micrograms (μg/ml) per ml of the preparation, Emblicanin-A and Emblicanin-B entering the cell can protect the inside of the mitochondria. membrane. As a result, the oxidative phosphorylation reaction in the inner membrane of the mitochondria improves the efficiency of synthesizing the triphosphate. In detail, the amount of triphosphonoside synthesized by oxidative phosphorylation of mitochondria protected and repaired by Emblicanin-A and Emblicanin-B is increased, and the basic oxygen consumption (Basal Respiration) and maximum oxygen consumption of the mitochondria Maximal Respiration, ATP Production, Spare Respiratory Capacity, and Coupling Efficiency were significantly improved. In addition, the mitochondria of Emblicanin-A and Emblicanin-B were obtained, and their mitochondrial activity was improved. The energy supply to the cells in which the mitochondria activated by Emblicanin-A and Emblicanin-B are located is increased, so that the ability of the cells to resist stress is also improved.

A method of administering Emblicanin-A and Emblicanin-B to a cell is, for example, oral administration of a composition comprising Emblicanin-A and Emblicanin-B or a preparation thereof. When Emblicanin-A and Emblicanin-B are administered orally in an oral manner, the effective dose of Emblicanin-A and Emblicanin-B is 151 mg (mg) to 237 mg. The effective dose here is obtained by converting the effective dose of the cell experiment to the human body kilogram. The conversion formula is as follows: effective dose of human body = effective dose of cell experiment × mouse body weight × conversion coefficient × human body kilogram. Among them, it is assumed that the mouse has a body weight of 20 grams, the human body has a kilogram of 60 kilograms, and the conversion coefficient is 9.01. The conversion coefficient is obtained from a table of conversion factors per kilogram of body weight of the animal and the human body.

In order to facilitate the oral administration of Emblicanin-A and Emblicanin-B, the composition comprising Emblicanin-A and Emblicanin-B can be made, for example, in the form of a liquid, a solid, a granule, a powder, a paste or a gel. preparation.

In some embodiments of the invention, only Emblicanin-A and Emblicanin-B may be included in the formulation. In another embodiment of the present invention, the preparation may also contain other ingredients or additives, such as carriers, diluents, adjuvants, and additives, without affecting the effects and achievable effects of the present invention. Forming agent or flavoring agent. Excipients make the formulation convenient and practical, while flavoring agents can enhance the flavor of the formulation.

The formulation may be a hard capsule whereby the composition comprising Emblicanin-A and Emblicanin-B is coated in a dry powder form. The preparation may also be a soft capsule by coating a composition comprising Emblicanin-A and Emblicanin-B in the form of a solution, a suspension, a paste, a powder or a granule.

The oil or fat used in the soft capsule for dissolving or dispersing the composition comprising Emblicanin-A and Emblicanin-B is, for example, avocado oil, almond oil, linseed oil, cumin oil, white succulent oil, olive oil, olive squalene. , orange oil, orange roughy oil, sesame oil, garlic oil, cocoa butter, pumpkin seed oil, chamomile oil, carrot oil, cucurbit oil, tallow fatty acid, Hawaiian stone fruit oil, orange oil, brown rice germ Oil, rice oil, wheat germ oil, safflower oil, shea oil, liquid avocado oil, perilla oil, soybean oil, evening primrose oil, camellia oil, corn oil, rapeseed oil, saw palmetto extract oil (saw palmetto Extract oil), eucalyptus oil, peach oil, parsley oil, castor oil, sunflower oil, grape seed oil, borage oil, Australian walnut oil, meadowfoam oil, cottonseed oil, peanut oil, turtle oil , oyster sauce, egg butter, fish oil, palm oil, palm kernel oil, wood wax, coconut oil, long chain/medium chain/short chain fatty acid triglyceride, diglyceride, butter, lard, squalene , squalane, decane, Such fats and oils of hydride. Among them, borage oil and evening primrose oil contain a large amount of Gamma-Linolenic Acid (GLA), which is an essential fatty acid of the human body, which has moisturizing, promotes cell regeneration and promotes brown fat activity. Degree to promote the function of fat burning.

Excipients are, for example, wheat starch, rice starch, corn starch, potato starch, dextrin, cyclodextrin and the like; crystalline cellulose; lactose, glucose, sugar, reduced maltose, sucrose, fructooligosaccharide, emulsified oligosaccharide Such as sugars; sugar alcohols such as sorbitol, erythritol, xylitol, lactitol, and mannitol.

The flavoring agents are various juice extracts such as longan extract, lychee extract, and grapefruit extract; various juices such as apple juice, orange juice, and lemon juice; various flavors such as peach flavor, plum flavor, and yoghurt flavor; Various sweeteners such as potassium acid, sucralose, erythritol, oligosaccharides, mannose, xylitol, isomerized sugars; various acidulants such as citric acid, malic acid, tartaric acid, gluconic acid; green tea, oolong tea, Various tea ingredients such as Banaba tea, Eucommia tea, Tieguanyin tea, scented tea, safflower tea, scented white tea, and kelp tea.

In addition, colorants, preservatives, tackifiers, binders, disintegrants, dispersants, stabilizers, gelling agents, antioxidants, surfactants, preservatives, pH adjusters, etc. are added in accordance with government regulations. The substance may also be added to the preparation containing Emblicanin-A and Emblicanin-B according to the dosage standard prescribed by the government unit and the demand for processing and production.

Further, the preparation of the present invention can be used for protecting and repairing mitochondria of various cells, such as mitochondria of cells such as fibroblasts, skeletal muscle cells, retinal cells, liver cells, and the like. Further, the preparation of the present invention activates the mitochondria, thereby providing more energy and cells in which the mitochondria are located, so that the cell activity of the cells in which the mitochondria are located is enhanced.

Hereinafter, the method for protecting the mitochondria disclosed in the present invention will be described by way of examples and comparative examples of the present invention, and experimental tests are carried out to demonstrate the efficacy of the method for protecting mitochondria disclosed in the present invention.

The cells used in this experiment were human skin fibroblasts CCD-966SK, purchased from the Food Industry Research Institute, ATCC No. CRL-1881. The experimental samples were prepared by implanting about 20,000 human fibroblasts in each well of a 24-well plate and culturing for 24 hours.

During the experiment, a total of 18 μg/ml of Emblicanin-A and Emblicanin-B was first added to the wells and soaked for 24 hours. Finally, the oxygen consumption of the cells in the wells was measured with a hippocampus bioenergy meter.

The measurement principle and flow of the hippocampus bioenergy analyzer are as follows. First, the medium in the well was changed to the culture medium for analysis. Next, the basal oxygen consumption of the cells in the well is detected. Next, a triphosphate synthase inhibitor is added to inhibit the production of triphosphates by the mitochondria, and the reduced oxygen consumption is the oxygen consumption of the synthetic triphosphate. The inhibitor of the triphosphate synthase is, for example, Oligomycin. Next, an appropriate concentration of the anti-coupling agent is added, and the mitochondria are idling at the extreme conditions without damaging the electron transport chain of the inner membrane of the mitochondria to evaluate the maximum oxygen consumption capacity of the mitochondria. The anti-coupling agent is, for example, Carbonyl cyanide-4-(trifluoromethoxy)phenylhydrazone (FCCP). Finally, the addition of an electron transport chain inhibitor completely shuts off the oxygen consumption of the mitochondria, thereby confirming the measured background value, ie, non-mitochondrial Respiration. The electron transport chain inhibitor is, for example, a combination of rotenone and antimycin A.

The basic oxygen consumption of the mitochondria is equal to the basic oxygen consumption of the cells minus the non-mitochondrial oxygen consumption. The basal oxygen consumption of the mitochondria minus the amount of oxygen consumed by the synthetic triphosphate is equal to the oxygen consumption of the Proton Leakage. The maximum oxygen consumption capacity of the mitochondria minus the basal oxygen consumption of the mitochondria is equal to the pre-existing oxygen consumption capacity of the mitochondria. The mitochondrial trinucleotide mediation efficiency is equal to the oxygen consumption of the synthetic triphosphate glycosides divided by the basal oxygen consumption of the mitochondria.

The concentrations and experimental measurements of the compositions comprising Emblicanin-A and Emblicanin-B of the examples and comparative examples are shown in Table 1.

Table I

Please refer to FIG. 1 to FIG. 6 and Table 1. BRIEF DESCRIPTION OF THE DRAWINGS Fig. 1 is a schematic view showing the pre-existing oxygen consumption capacity of mitochondria in normal human fibroblasts of Examples and Comparative Examples. Fig. 2 is a graph showing the basic oxygen consumption of mitochondria in normal human fibroblasts of the examples and comparative examples. Fig. 3 is a graph showing the maximum oxygen consumption capacity of the mitochondria in normal human fibroblasts of the examples and the comparative examples. Fig. 4 is a graph showing the oxygen consumption of the synthesis of the triphosphates of the mitochondria in the normal human fibroblasts of the examples and the comparative examples. Fig. 5 is a graph showing the efficiency of trinucleotide-series mediation of mitochondria in normal human fibroblasts of the examples and the comparative examples. Fig. 6 is a graph showing the oxygen consumption of the mitochondria in the normal human fibroblasts of the examples and the comparative examples against hydrogen ion leakage.

As shown in Figure 1, the pre-existing oxygen consumption capacity of the examples was significantly higher than that of the comparative examples (p < 0.001). As shown in Fig. 2, the basic oxygen consumption of the examples was significantly higher than that of the comparative examples (p < 0.05). As shown in Figure 3, the maximum oxygen consumption capacity of the examples was significantly higher than that of the comparative examples (p < 0.001). As shown in Fig. 4, the oxygen consumption of the synthetic triphosphate glycosides of the examples was significantly higher than that of the comparative examples (p < 0.001). As shown in Fig. 5, the triethylglycoside mediation efficiency of the examples was significantly higher than that of the comparative examples (p < 0.05). As shown in FIG. 6, the oxygen consumption of the embodiment against hydrogen ion leakage is higher than that of the comparative example.

According to the above experimental test results, the mitochondria treated with the total concentration of Emblicanin-A and Emblicanin-B at a concentration of 18 μg/ml were protected by Emblicanin-A and Emblicanin-B to reduce the oxidant in the inner membrane of the granule. damage. At the same time, the pre-existing oxygen consumption capacity of the mitochondria is increased, so that the elasticity of the mitochondria which can be adjusted or withstood when subjected to changes or pressures is increased.

The above values were brought into the following formula to calculate BHI (Bioenergetic Healthy Index) values of the examples and comparative examples.

BHI=[Oxygen consumption of synthetic triphosphates* Pre-existing oxygen consumption capacity]/[Oxygen consumption over hydrogen ion leakage × Non-mitochondrial oxygen consumption]

It can be seen from the above formula that the BHI value of the example is about 0.21, and the BHI value of the comparative example is about 0.05, that is, the BHI value of the example is about 4 times of the BHI value of the comparative example, so that Emblicanin-A and Emblicanin can be inferred. The mitochondria of the Example after the treatment of -B were indeed protected and repaired.

In addition, it can be seen from the above experimental test results that Emblicanin-A and Emblicanin-B can protect and repair the mitochondria of human fibroblasts and enhance the activity of mitochondria in human fibroblasts. Therefore, human fibroblasts containing these mitochondria are activated, thereby improving the strain capacity and differentiation ability of human fibroblasts, such as continuous secretion of collagen, elastin, intercellular matrix and growth factors, and stimulation of angiogenesis. With the tissue repair function, etc., to achieve the effect of tissue renewal and regeneration. In this way, activated human fibroblasts can be further used in various applications, such as filling wrinkles, repairing mucous membranes and muscles, treating wounds such as diabetic ulcers, burns, stress urinary incontinence, Achilles tendinitis. Gingival atrophy and so on.

According to the method for protecting mitochondria disclosed in the above invention, Emblicanin-A and Emblicanin-B are supplied to cells to protect the inner membrane of the mitochondria and delay the disintegration of the mitochondria. In this way, the rate at which mitochondrial disintegration triggers apoptosis can be slowed down.

Furthermore, the mitochondria of Emblicanin-A and Emblicanin-B were obtained, and their mitochondrial activity was improved. The energy supply to the cells in which the mitochondria activated by Emblicanin-A and Emblicanin-B are located is increased, so that the ability of the cells to resist stress is also improved.

Furthermore, Emblicanin-A and Emblicanin-B can protect and repair the mitochondria of human fibroblasts, thereby improving the strain capacity and differentiation ability of human fibroblasts, thereby achieving the effects of tissue renewal and regeneration, and then applied to Various uses.

Although the present invention has been disclosed above in the foregoing embodiments, it is not intended to limit the invention. It is within the scope of the invention to be modified and modified without departing from the spirit and scope of the invention. Please refer to the attached patent application for the scope of protection defined by the present invention.

no.

Fig. 1 is a schematic view showing the pre-existing oxygen consumption capacity of the mitochondria in the human fibroblasts of the examples and the comparative examples. Fig. 2 is a graph showing the basic oxygen consumption of the mitochondria in the human fibroblasts of the examples and the comparative examples. Fig. 3 is a graph showing the maximum oxygen consumption capacity of the mitochondria in the human fibroblasts of the examples and the comparative examples. Fig. 4 is a graph showing the oxygen consumption of the synthesis of triphosphates of the mitochondria in the human fibroblasts of the examples and the comparative examples. Fig. 5 is a graph showing the efficiency of trinucleotide mediation of mitochondria in human fibroblasts of the examples and the comparative examples. Fig. 6 is a graph showing the oxygen consumption of the mitochondria in the normal human fibroblasts of the examples and the comparative examples against hydrogen ion leakage.

Claims (14)

A use of a composition for the preparation of a formulation for protecting and repairing mitochondria, wherein the composition comprises an Emblicanin-A and an Emblicanin-B. The use of claim 1, wherein the weight ratio of the Emblicanin-A to the Emblicanin-B is from 3/2 to 2/3. The use of claim 1, wherein the total concentration of the Emblicanin-A and the Emblicanin-B is 14 to 22 micrograms per milliliter (μg/ml). The use according to claim 1, wherein the total effective dose of the human body of the Emblicanin-A and the Emblicanin-B is 151 mg to 237 mg. The use of claim 1, wherein the mitochondria are mitochondria in human fibroblasts. The use of claim 1, wherein the cellular activity of a cell in which the mitochondria are protected and repaired is elevated. The use of claim 1, wherein the formulation increases the maximum oxygen consumption of the mitochondria (Maximal Respiration). The use of claim 1, wherein the formulation increases the oxygen consumption rate of the synthetic mitochondrial (ATP Production) of the mitochondria. The use of claim 1, wherein the formulation increases the Spare Respiratory Capacity of the mitochondria. The use of claim 1, wherein the formulation increases the basic oxygen consumption of the mitochondria (Basal Respiration). The use of claim 1, wherein the formulation increases the Coupling Efficiency of the mitochondrial. The use of claim 1, wherein the formulation further comprises a pharmaceutically or pharmaceutically acceptable carrier, excipient, diluent or adjuvant. The use according to claim 1, wherein the preparation is an oral preparation. The use according to claim 1, wherein the preparation is in the form of a hard capsule or a soft capsule.