KR101386727B1 - A Phamaceutical Composition Comprising a Fermented Purple Sweet Potato for Treating or Preventing Liver Disease - Google Patents

Abstract

The present invention relates to a pharmaceutical composition for preventing or treating liver disease containing a fermentation broth obtained by fermenting a purple sweet potato extract, wherein the fermentation broth obtained by fermenting the purple sweet potato extract of the present invention is effective for inhibiting liver damage to ethanol using HepG2 / 2E1. It can be usefully used as a pharmaceutical composition for preventing or treating liver disease or as a health functional food for preventing or improving liver disease.

Description

Pharmaceutical composition for preventing or treating liver disease containing purple sweet potato extract fermentation broth as an active ingredient {A Phamaceutical Composition Comprising a Fermented Purple Sweet Potato for Treating or Preventing Liver Disease}

The present invention relates to a pharmaceutical composition for preventing or treating liver disease, in particular, a pharmaceutical composition applied to alcoholic liver disease, comprising a fermentation broth obtained by fermenting purple sweet potato extract, and to a health functional food having a prophylactic or improvement effect of liver disease. It is about.

Liver disease is a series of processes that usually progress from hepatitis to liver fibrosis, cirrhosis, and liver cancer. Hepatitis-causing factors in the early stage include hepatotoxic substances, hepatitis virus, drugs, alcohol, and autoimmune reactions. Repeated inflammatory responses resulting from these lead to the next step.

Acute alcohol consumption is also known to cause toxic effects directly or indirectly in humans, and products such as acetaldehyde and acetate cause damage to liver cells (Shiraishi K et al, Ethanol Ethanol, 28: 59-64). , 1993). More than 80-90% of the ingested ethanol is metabolized in the liver and ethanol is oxidized to acetaldehyde. The toxic effects of many ethanol in the liver are explained by the oxidative stress caused by ethanol metabolism (Enomoto N, et al, J. Gastroenterol. Hepatol, 15: D20-5, 2000; Senhilkumar R et al Asia Pac. J Clin.Nutr, 11: 157-63, 2002). Ethanol or its metabolites cause autooxidation in hepatocytes and cause hepatitis by acting as a pro-oxidative factor or by reducing antioxidant levels. The association of lipid oxidation with membrane damage is also an important feature in ethanol liver damage (Senthilkumar R, et al, J. Nutr. Biochem, 14: 452-8, 2003; Senthilkumar R et al. Pol. J. Pharmacol, 55: 603-11, 2003).

Therefore, there is a need for development of a hepatoprotective active substance or a hepatitis therapeutic agent that can inhibit their initial reaction. Silymarin is a representative liver treatment currently used clinically. It has been isolated from the fruit of Silybum marianum, an asteraceae plant, and the recently developed dimethyl dimethoxybiphenylate is a component of Schizandra chinensis. Hepatotoxic effects have been reported with synthetic compounds similar to schizandrin. However, in view of the severity and frequency of these diseases, it is urgently needed to develop a preventive or therapeutic agent for liver disease in natural products.

On the other hand, purple sweet potato (Purple-Fleshed Sweet Potato) was artificially crossed in 1993 using a variety of dried varieties of processed varieties of dry brown rice and purple pigment containing Yamakawamurasaki in the production test and local adaptation test. Multiplicity was confirmed, and breeding was carried out in three new varieties: Jami in 1998, Borami in 2000, and Shinjami in 2001. These new varieties have all of the native ingredients of wild sweet potatoes, similar in size to regular sweet potatoes, but harvest more than twice as much as the wild ones, and have the advantage of having sweet potatoes' own saccharides and fibrin. The purple sweet potato contains a large amount of pigment extract, so that not only the epidermal layer but also the entire flesh has a deep purple color.The functional pigment content is 10 to 60 times higher than a cream berry and 5 to 7 times higher than a grape berry, as well as a functional food source. Excellent value as a material.

The main active ingredient of these purple sweet potatoes is anthocyanin, a flavonoid pigment (J Agric. Food Chem. 51, pp 628-633, 2003; J Nat. Prod. 62, p 802, 1999; J Med. Food. 4 pp 211-218, 2001), anti-diabetic (J Agric Food Chem. 49, pp 1948-1951, 2001), and anticancer effects (Cancer Lett. In press pp 1-12, 2005). .

In addition, Korean Patent No. 0792029 discloses that purple sweet potato pigment extract inhibits the activities of cytochrome P450 enzymes (P450 1A1, P450 2E1), which are liver metabolism enzymes involved in metabolism of chemicals that cause liver damage such as acetaminophen. In addition, it is disclosed that by increasing the detoxifying enzyme that promotes the release of the body through the incorporation of a metabolite of a chemical that causes liver damage, it can be usefully used for the prevention and treatment of liver disease.

An object of the present invention to provide a pharmaceutical composition for the prevention or treatment of liver disease using a fermentation broth obtained by fermenting the purple sweet potato extract.

In addition, another object of the present invention to provide a health functional food for the prevention or improvement of liver disease using a fermentation broth obtained by fermenting the purple sweet potato extract.

Another object of the present invention is to ferment the purple sweet potato extract to produce an active ingredient for the prevention or treatment of alcoholic liver disease ( Isatchenkia orientalis) orientalis ) to provide PSP-HAN [Accession No .: KFCC11519P].

The present invention provides a pharmaceutical composition for preventing or treating liver disease, which contains a fermentation broth obtained by fermenting a purple sweet potato extract as an active ingredient.

In the pharmaceutical composition for preventing or treating liver disease of the present invention, the purple sweet potato extract is characterized in that the hot water extract or an extract of a lower alcohol solution of 2 to 4 carbon atoms.

In the pharmaceutical composition for preventing or treating liver disease of the present invention, the hot water extract is characterized in that extracted 0.5 ~ 24 hours at 80 ~ 105 ℃.

The hot water extract in the liver disease treatment and prevention composition of the present invention is characterized in that extracted for 0.5 to 24 hours at 80 ~ 105 ℃.

The fermentation broth in the liver disease treatment and prophylactic composition of the present invention is the Issachenchia orientalis of purple sweet potato extract ( Issatchenkia orientalis ) fermentation broth.

In the pharmaceutical composition for preventing or treating liver disease of the present invention, the issatchenkia orientalis is characterized in that the Issatchenkia orientalis PSP-HAN [Accession No .: KFCC11519P].

In the pharmaceutical composition for preventing or treating liver disease of the present invention, after inoculating yeast to the purple sweet potato extract, it is characterized by fermentation at 25 to 40 ° C. for 20 to 100 hours.

In the pharmaceutical composition for preventing or treating liver disease of the present invention, the purple sweet potato is extracted with hot water at 80 to 105 ° C. for 0.5 to 24 hours, and the hot water extract is inoculated with Issatchenkia orientalis and then 25 to 40 ° C. It characterized in that it comprises a fermentation broth fermented in 18 to 78 hours as an active ingredient.

In the pharmaceutical composition for preventing or treating liver disease of the present invention, the purple sweet potato is characterized in that the sweet potato of any one or more varieties selected from Yamakawamurasaki, Yeonmi, Borami and Shinjami.

In the pharmaceutical composition for preventing or treating liver disease of the present invention, the liver disease is characterized by being an alcoholic liver disease.

In another aspect, the present invention provides a health functional food having a prevention or improvement effect of liver disease containing a fermentation broth obtained by fermenting the purple sweet potato extract as an active ingredient.

In addition, the present invention is fermented purple sweet potato extract to produce an active ingredient for the prevention or treatment of alcoholic liver disease ( Isatchenkia orientalis) orientalis ) PSP-HAN [Accession No .: KFCC11519P].

The fermentation broth obtained by fermenting the purple sweet potato extract of the present invention shows the effect of inhibiting liver damage to ethanol using HepG2 / 2E1, and is useful as a pharmaceutical composition for preventing or treating liver disease or as a health functional food for preventing or improving liver disease. Can be. In particular, the fermentation broth obtained by fermenting the purple sweet potato extract of the present invention inhibits liver damage as the anthocyanin pigment is enhanced with pH reduction during fermentation, even though an anthocyanin pigment is used as a crude extract that is not separated or purified using column chromatography. Efficacy is significantly enhanced.

1 is a 18s rDNA partial sequencing phylogenetic table of YPD Large bacteria isolated in the present invention.
Figure 2 is a 18s rDNA partial sequencing phylogenetic table of the YPD Small bacteria isolated in the present invention.
Figure 3 is a graph showing the pH change of the fermentation broth with the fermentation time in Example 1.
Figure 4 is a graph showing the change in absorbance at 530 nm with the fermentation time in Example 1.
FIG. 5 is a graph showing cytotoxicity against HepG2 / 2E1 cell lines according to the sample concentrations of Comparative Example 1 and Experimental Example 1 in Experimental Example 2. FIG.
Figure 6 is a graph showing the hepatocyte protective effect according to the addition of the sample of Comparative Example 1 and Experimental Example 1 HepG2 / 2E1 cell line damaged by ethanol in Experimental Example 3.
7 is a graph showing the antioxidant effect according to the addition of the sample of Comparative Example 1 and Experimental Example 1 in Experimental Example 4.

The pharmaceutical composition or health functional food for preventing and treating liver disease of the present invention contains a fermentation broth obtained by fermenting the purple sweet potato extract as an active ingredient.

In the present invention, purple sweet potatoes are higher in anthocyanin content than general sweet potatoes, and are not particularly limited, but Yamakawamura, zucchini, Borami or Shinzami are preferable, and more preferable are Yamakawamura or Shinzami , And most preferably believer.

The purple sweet potato extract of the present invention is preferably an extract having a high content of anthocyanin together with sugar as a substrate used by the yeast.

Purple sweet potato extract of the present invention is preferably hot water extract. For example, it can be extracted in water of 80 to 105 ℃, preferably 90 to 100 ℃ 0.5 to 24 hours, preferably 1 to 6 hours. If the extraction temperature is below the lower limit of the extraction temperature, the extraction efficiency of the anthocyanin is lowered and the germicidal effect against the germs can not be sufficiently attained. Therefore, there is a problem that the germs can be proliferated by increasing the extraction time. The sterilization efficiency of the germ is increased, but the anthocyanin is destroyed and the color of the extract becomes turbid. If the extraction time is less than the lower limit of the extraction time, sufficient anthocyanin is difficult to be extracted, resulting in loss of raw materials and lower yield of the final product. If the extraction time is increased, the extraction efficiency is not increased and the anthocyanin is destroyed . In order to increase the anthocyanin extraction efficiency of the purple sweet potato, it is preferable to acidify the pH of the water to 2 to 6, preferably pH 3 to 5 by adding an acid. To lower the pH of the water, organic acids such as citric acid, lactic acid, succinic acid, malic acid, malic acid, and acetic acid may be used although not particularly limited.

As a purple sweet potato extract of the present invention, an extract by a lower alcohol aqueous solution having 2 to 4 carbon atoms may be used. For example, it is extracted with an aqueous alcohol solution such as ethanol, methanol, isopropanol, preferably 20 to 80% by weight of an aqueous alcohol solution, more preferably 50 to 70% by weight of an aqueous alcohol solution. In order to increase the anthocyanin extraction efficiency of the aqueous alcohol solution, an acid may be added to the alcohol in the same manner as the hot water extraction. If alcohol extract is used, concentrate the alcohol or alcohol concentrate or alcohol extract to lower the alcohol content by evaporating alcohol from the alcohol extract before yeast is inoculated, and then dissolve in water before use. In addition, in order to suppress the propagation of various bacteria, the alcohol extract is used after sterilization for 0.5 to 24 hours and preferably 1 to 6 hours at 80 to 105 ° C, preferably 90 to 100 ° C.

The strain inoculated for the fermentation of the purple sweet potato extract of the present invention is yeast, preferably Issatchenkia orientalis ), more preferably Issatchenkia orientalis PSP-HAN (Accession No .: KFCC11519P). The inoculation amount of the yeast is used in an amount of 0.001 to 10 parts by weight, preferably 0.01 to 5 parts by weight, and more preferably 0.1 to 3 parts by weight based on 100 parts by weight of the purple sweet potato extract based on 1 × 10 ⁷ cells / ml.

After inoculating the purple sweet potato extract in the pharmaceutical composition or health functional food for liver disease prevention or treatment of the present invention issakenchia orientalis 25 to 40 ℃, preferably 30 to 35 ℃ 20 to 100 hours, preferably Fermentation is 50 to 95 hours, more preferably 70 to 90 hours.

The fermentation broth obtained by fermenting the purple sweet potato extract in the pharmaceutical composition or health functional food for preventing or treating liver disease of the present invention is characterized in that the fermentation broth is contained in an amount of 0.01 to 50% by weight (based on solids).

Fermentation broth fermented purple sweet potato extract of the present invention can be used in various ways as a health functional food for preventing or improving liver disease. Health functional food containing the purple sweet potato extract fermentation broth of the present invention as an active ingredient can be used in the form of various foods, for example, beverages, gum, tea, vitamin complex, powder, granules, tablets, capsules.

When the fermentation broth obtained by fermenting the purple sweet potato extract of the present invention is included in the health functional food, the amount thereof may be added as 0.01 to 15% by weight of the total food weight when dried or solid, and 0.02 to 10 g based on 100 ml for beverages. Preferably, it can be added in the ratio of 0.3-1 g.

When the health functional food of the present invention is prepared in the form of a beverage in addition to containing the purple sweet potato extract fermentation broth as an essential ingredient in the indicated ratio, there are no particular limitations on the liquid components and various flavors or natural carbohydrates, such as ordinary drinks. May be contained as an additional component. Examples of the above-mentioned natural carbohydrates include monosaccharides such as disaccharides such as glucose and fructose such as maltose, sucrose and the like and polysaccharides such as dextrin, cyclodextrin and the like Sugar, and sugar alcohols such as xylitol, sorbitol, and erythritol. As natural flavors other than those described above, natural flavors (such as tau martin, stevia extract (e.g., rebaudioside A, glycyrrhizin)) and synthetic flavors (saccharin, aspartame, etc.) have. The ratio of the natural carbohydrate is generally about 1 to 20 g, preferably about 5 to 12 g per 100 ml of the beverage of the present invention.

In addition, the health functional food of the present invention, in addition to the fermentation broth fermented purple sweet potato extract used as an active ingredient, various nutrients, vitamins, minerals (electrolytes), synthetic flavors and natural flavors such as flavoring agents, coloring agents and neutralizing agents (cheese) , Chocolate and the like), pectic acid and salts thereof, alginic acid and salts thereof, organic acids, protective colloid thickeners, pH adjusting agents, stabilizers, preservatives, glycerin, alcohols, carbonation agents used in carbonated drinks and the like.

In addition to the fermentation broth obtained by fermenting the purple sweet potato extract, the health functional food of the present invention may contain a flesh for preparing natural fruit juice and fruit juice beverage and vegetable beverage. These components may be used independently or in combination. The proportion of such additives is not so critical, but is generally selected in the range of 0 to about 20 parts by weight per 100 parts by weight of the composition of the present invention.

The health functional food of the present invention is prepared by fermenting the extract of purple sweet potato for use as food, and can be used with confidence even when taken for a long time for the purpose of prevention or treatment with little toxicity and side effects.

The present invention provides a pharmaceutical composition for preventing or treating liver disease, which comprises a fermentation broth fermented with purple sweet potato extract as an active ingredient.

The pharmaceutical composition of the present invention comprises 0.1 to 50% by weight (based on solids) of the fermentation broth obtained by fermenting the purple sweet potato extract based on the total weight of the pharmaceutical composition. However, the composition is not limited thereto, and may vary depending on the condition of the patient, the type of disease, and the progress of the disease.

The pharmaceutical compositions of the present invention may further comprise suitable carriers, excipients and diluents commonly used in the manufacture of pharmaceutical compositions.

The pharmaceutical compositions according to the invention can be used in the form of oral dosage forms, external preparations, suppositories, and sterile injectable solutions, such as powders, granules, tablets, capsules, suspensions, emulsions, syrups, aerosols, etc., respectively, according to conventional methods. Carriers, excipients and diluents that may be included in the pharmaceutical composition include lactose, dextrose, sucrose, sorbitol, mannitol, xylitol, erythritol, maltitol, starch, acacia rubber, alginate, gelatin, calcium phosphate, calcium silicate , Cellulose, methyl cellulose, microcrystalline cellulose, polyvinyl pyrrolidone, water, methylhydroxybenzoate, propylhydroxybenzoate, talc, magnesium stearate and mineral oil. In the case of formulation, a diluent or excipient such as a filler, an extender, a binder, a wetting agent, a disintegrant, or a surfactant is usually used. Solid preparations for oral administration include tablets, pills, powders, granules, capsules, and the like, and the solid preparations may include at least one excipient such as starch, calcium carbonate, sucrose in the extract. ) Or lactose, gelatin and the like are mixed. In addition to simple excipients, lubricants such as magnesium stearate talc are also used. Examples of the liquid preparation for oral use include suspensions, solutions, emulsions, and syrups. In addition to water and liquid paraffin, simple diluents commonly used, various excipients such as wetting agents, sweeteners, fragrances, preservatives and the like may be included . Formulations for parenteral administration include sterilized aqueous solutions, non-aqueous solutions, suspensions, emulsions, freeze-dried preparations, and suppositories. Examples of the suspending agent include propylene glycol, polyethylene glycol, vegetable oil such as olive oil, injectable ester such as ethyl oleate, and the like. As the base of the suppository, witepsol, macrogol, tween 61, cacao butter, laurin butter, glycerogelatin and the like can be used.

The preferred dosage of the pharmaceutical composition of the present invention varies depending on the condition and the weight of the patient, the degree of disease, the drug form, the administration route and the period, but can be appropriately selected by those skilled in the art. However, for the desired effect, it is preferable to administer from 0.01 mg / kg to 10 g / kg, preferably 1 mg / kg to 1 g / kg per day based on the fermentation broth solid fermentation of the purple sweet potato extract of the present invention . The administration may be carried out once a day or divided into several doses. Thus, the dosage amounts are not intended to limit the scope of the invention in any manner.

The pharmaceutical composition of the present invention can be administered to mammals such as rats, mice, livestock, humans, and the like in various routes. All modes of administration may be expected, for example, by oral, rectal or intravenous, intramuscular, subcutaneous, intra-uterine or intracerebroventricular injections.

Hereinafter, the present invention will be described in more detail through Examples, Comparative Examples, Experimental Examples, and Formulation Examples. The following examples are illustrative of the present invention and are not intended to limit the scope of the present invention.

Comparative Example  1: purple sweet potato extract

After cutting the purple sweet potato of Shinjami varieties within 1 month of harvesting to 1 cm thickness, extracted with sweet potato 3 times the weight of water at 90 ℃ for 60 minutes, the extract was cooled to 25 ℃ and filtered to filter the filtrate and residue The separated filtrate obtained by separation was concentrated under reduced pressure at 35 ° C. using a vacuum condenser equipped with a cold inhaler to obtain a concentrate, and the concentrate was prepared using a freeze vacuum dryer to prepare a purple sweet potato extract powder.

Example  1: Purple Sweet Potato Extract Fermentation Solution

1) Isolation of Fermented Strains from Purple Sweet Potato Fermented Beverage

The main causes of fermentation were isolated from purple sweet potato fermented beverage-producing mixed seedlings. Two colonies (YPD Large and YPD Small) were isolated from YPD agar medium at 25 ° C. for 24 hours.

This was commissioned by the Korea Center for Microbiological Conservation (KCCM) and examined homology after blast search of NCBI through 18S rDNA sequence. The 18S rDNA sequence of YPD Large was shown in SEQ ID NO: 1, and the 18S rDNA sequence of YPD Small was shown in SEQ ID NO: 2. Both YPD Large and Small are 99% homologous, and Issatchenkia orientalis ) (FIGS. 1 and 2).

Although the two species of Isakenchia orientalis have almost similar growth patterns, the YPD Large bacteria having excellent activity of the bacteria is Isatchenkia Orientalis ( Issatchenkia). orientalis ) was named PSP-HAN, which was deposited with the Korea Microorganism Conservation Center on November 16, 2011 and was assigned the accession number KFCC11519P.

2) Preparation of Purple Sweet Potato Extract Fermentation Solution

After cutting the purple sweet potato of Shinjami varieties within 1 month after harvesting to 1 cm thickness, extracted with sweet potato weight of 3 times the weight 90 ℃, 60 minutes, and cooled the extract to 25 ℃ 1 × 10 ⁷ cell / After inoculating 1 wt% of ml Isakenchia orientalis PSP-HAN, it was fermented at 30 ° C. for 24 to 96 hours. The fermentation broth was filtered to separate the filtrate and the residue. The fermentation filtrate was concentrated under reduced pressure at 35 ° C. using a vacuum condenser equipped with a cold inhaler, and the concentrate was obtained by using a freeze vacuum dryer. Extract fermentation broth powder was prepared.

The pH change of the fermentation broth according to the fermentation time is shown in FIG. 3, and the change in absorbance at 530 nm for detecting ansocyanine pigment is shown in FIG. 4.

When the fermentation time was 0 hours (the same as Comparative Example 1), the pH was rapidly decreased and the absorbance was increased more than 24 hours.Therefore, there was no significant change in the 24-hour to 72-hour fermentation. As it became lower, the absorbance increased further. The preliminary test results showed that the fermentation broth fermented for 24 hours to 72 hours was excellent in inhibiting the hepatocyte damage.

Experimental Example  1: General composition analysis

In Comparative Example 1 and Example 1, the extraction filtrate and the fermentation filtrate before concentration and drying were measured as a sample according to the Food Code, and the total nitrogen, crude fat, moisture, and ash content were measured and carbohydrate content was calculated and shown in Table 1.

division Total nitrogen
(weight%) Crude fat
(weight%) moisture
(weight%) Ash
(weight%) carbohydrate
(weight%) Comparative Example 1 0.03 0.09 95.1 0.23 4.39 Example 1 0.03 0.05 97.2 0.18 2.39

The fermentation filtrate of Example 1 was confirmed to reduce the content of ash, fat, and carbohydrates compared to the extraction filtrate of Comparative Example 1, which means that the yeast used in the fermentation of minerals, fats, sugars, etc. contained in the purple sweet potato water extract It appears to have broken down these components using it as a nutrient source.

Experimental Example 2: Cytotoxicity Test

In order to confirm the cytotoxicity of the purple sweet potato extract and purple sweet potato extract fermentation broth of the present invention obtained in Comparative Example 1 and Example 1 was measured as follows by modifying the method of Rochem and the like. (Documents: Dai, Y. and AI . Cederbaum 1995. cytotoxicity of acetaminophen in human cytochrome P4502E1-transfected HepG2 cells J. Pharmacol Exp Ther 273:.... 1497 - 1505)

In order to confirm whether the purple sweet potato extract and the purple sweet potato extract fermentation broth obtained in Comparative Example 1 and Example 1 cause toxicity to cells, HepG2 / 2E1 cell line [HepG2 (ATCC HB8065) was used as a 293GPG cell line (Clontech) for that cell; modified using a 24 wells plate 2 × 10 ⁵ for 24 hours after the frequency division to the cells / well of serum (serum) containing 10% medium {[Minimum Essential medium (MEM) , Hyclone], [fetal bovine serum (FBS), Hyclone]}.

Remove the medium after the primary culture and the medium containing 3% of the serum (serum) in which the sample was dissolved {[Minimum Essential Medium (MEM), Hyclone], [fetal bovine serum (FBS), Hyclone]} 1 The mL was dispensed and the reculture was repeated for 5 days. On day 7 after incubation, 0.25 ml of XTT-PMS solution [1 mg XTT (XTT Sodium Cell Culture Test, Sigma) and 0.0375 mg PMS (phenazine methosulfate, Sigma) / mL of Minimum Essential Medium in each well (Hyclone) mixed solution] was added and incubated again for 2 hours. The cytotoxicity of the cells was measured for absorbance at 450 nm using a microplate reader (BIO-TEK, 1010-1) to determine the degree of formazan formation.

As a result of the experiment, it was confirmed that the purple sweet potato extract and the purple sweet potato extract fermentation broth of the present invention did not exhibit cytotoxicity up to a concentration of 1,000 ㎍ / mL.

Experimental Example 3: Hepatic Injury Inhibitory Activity

Hepatic damage inhibition effect of the purple sweet potato extract of Comparative Example 1 and the purple sweet potato extract fermentation solution of the present invention obtained in Example 1 was measured by applying the method described in the literature as follows (Documents: Jamenez-Lpez, JM and AI Cederbaum. 2003. Green tea polyphenol epigallocatechin-3-gallate protects HepG2 cells against CYP2E1-dependent toxicity.Free radic. Biol. & Med. 36: 359-370).

Hepatoprotective activity of the purple sweet potato extract and purple sweet potato extract fermentation broth samples obtained in Comparative Examples 1 and 1 was 24 well for HepG2 / 2E1 cell line [cell modified with HepG2 (ATCC HB8065) using 293GPG cell line (Clontech)]. After dispensing the plate at 5 × 10 ⁴ cells / well, the medium containing 10% serum ([Minimum Essential Medium (MEM), Hyclone], [fetal bovine serum (FBS), Hyclone] for 16 hours. Incubated.

After 16 hours of incubation, the medium [Minimum Essential Medium (MEM)] was removed and each well contained a sample and 200 mM EtOH. Medium containing 3% serum [Minimum Essential Medium (MEM), Hyclone] was added and reacted in a 37 ° C. incubator for 24 hours. This process was repeated for 5 days, and on the 7th day, the medium containing the sample and ethanol was removed, and the XTT-PMS solution [1 mg XTT (XTT Sodium Cell Culture Test, Sigma) and 0.0375 mg PMS (phenazine methosulfate, Sigma) / mL of Minimum Essential Medium (Hyclone) solution] was added and incubated again for 2 hours. The damage inhibition effect of liver cells was confirmed by measuring the absorbance at 450 nm using a microplate reader (BIO-TEK, 1010-1) to form the resultant formazan. Negative control (Control) was used only the culture medium instead of the sample, the positive control was treated only 200 mM ethanol.

As shown in FIG. 6, the survival rate of the HepG2 / 2E1 cell line was significantly reduced to 72.8 ± 8.2 in the positive control group treated with ethanol as compared to the negative control group, and the purple sweet potato extract of Comparative Example 1 was added together with ethanol. Survival increased up to 83.8 ± 4.3, but it was still different from the negative control, and when the purple sweet potato extract fermentation broth was added with ethanol, the survival rate increased to 97.0 ± 2.3, equivalent to the negative control. It was confirmed that the purple sweet potato extract fermentation broth of Example 1 was significantly superior in inhibiting liver damage by ethanol compared to the purple sweet potato extract of Comparative Example 1.

Experimental Example  4: Antioxidant ability  Experiment

The antioxidant capacity of the purple sweet potato extract of Comparative Example 1 and the purple sweet potato extract fermentation broth of the present invention obtained in Example 1 was measured by applying the method described in the literature as follows (Documents: Yun-Mi J. and Kyoung-Chan P. 2005. Cytoprotective effect of green tea extract and quercetin against hydrogen peroxide-induced oxidative stress.Arch Pharm Res. 28 (11): 1251-1256).

Antioxidant activity of the purple sweet potato extract and purple sweet potato extract fermentation broth samples obtained in Comparative Examples 1 and 1 was obtained by dividing HepG2 cell line [HepG2 (Korea Cell Line Bank 88065) cell line] into 5 × 10 ⁴ cells / well in a 24 well plate. Subsequently, the cultures were incubated with medium containing 10% serum ([Minimum Essential Medium (MEM), Hyclone], [fetal bovine serum (FBS), Hyclone]} for 24 hours.

After 24 hours of incubation, the medium [Minimum Essential Medium (MEM)] is removed and each well contains a sample. 3% serum-containing medium [Minimum Essential Medium (MEM), Hyclone] was added, and reacted in a 37 ° C. incubator for 24 hours, and then a medium containing 0.3 mM hydrogen peroxide was added and reacted in a 37 ° C. incubator for 24 hours. At the end of the reaction, remove the medium containing hydrogen peroxide and mix with XTT-PMS solution [1 mg XTT (XTT Sodium Cell Culture Test, Sigma) and 0.0375 mg PMS (phenazine methosulfate, Sigma) / mL of Minimum Essential Medium (Hyclone) Solution] was added and incubated again for 2 hours. The damage inhibition effect of liver cells was confirmed by measuring the absorbance at 450 nm using a microplate reader (BIO-TEK, 1010-1) to form the resultant formazan. Negative control (Control) was used only the culture medium instead of the sample, the positive control was treated with only hydrogen peroxide.

As shown in FIG. 7, the positive control group treated with hydrogen peroxide only significantly reduced the survival rate of the HepG2 cell line by 48% compared to the negative control group, and 58% of the purple sweet potato extract of Comparative Example 1 was added together with hydrogen peroxide. Although the survival rate was increased to some extent, when the purple sweet potato extract fermentation broth of Example was added together with ethanol, the survival rate increased to 70% compared to the positive control, and the purple sweet potato extract fermentation broth of Example 1 was purple in Comparative Example 1. It was confirmed that the antioxidant efficacy by hydrogen peroxide was superior to the sweet potato extract.

Hereinafter, an example of the preparation containing the purple sweet potato extract fermentation broth of the present invention will be described, but the present invention is not intended to be limited thereto, but is intended to be described in detail.

Preparation of Formulation Example 1 Powder (unit: weight%)

Purple Sweet Potato Extract Fermentation Solution of Example 1 66.0

Lactose 34.0

The above components are mixed and packed in airtight bags to prepare powders.

Formulation Example 2. Preparation of tablets (unit:% by weight)

Purple Sweet Potato Extract Fermentation Solution of Example 1 33.0

Lactose 33.0

Corn Starch 33.3

Magnesium stearate 0.7

After mixing the above components, tablets are prepared by tableting according to a conventional tablet preparation method.

Formulation Example 3 Preparation of Wire (Unit: Weight%)

40% of brown rice, 30% of barley, 20% of yulmu and 10% of alpha derivative are mixed and crushed to make cereal powder having a grain size of 110 mesh. 40% of black beans, 30% of black sesame seeds and 30% of perilla seeds are distributed, and then a seed powder having a particle size of 110 mesh is prepared by a grinder. 75% of the grains, 20% of the seeds, and 5% of the purple sweet potato extract fermentation broth powder of Example 1 were granulated and then granulated.

Preparation Example 4. Preparation of chewing gum (unit:% by weight)

Chewing gum is prepared by a conventional method by combining gum base 20%, sugar 76%, fragrance 1.5% and water 2% and purple sweet potato extract fermentation broth powder 0.5%.

Formulation Example 5. Preparation of beverage (unit: wt%)

5% by weight of honey, 3% fructose, 0.0001% of sodium riboflavin hydrochloride, 0.0001% of pyridoxine hydrochloride, and 91.9998% of water and 1% of the purple sweet potato extract fermentation broth of Example 1 were combined to prepare a healthy beverage.

Korea Microorganism Conservation Center (Domestic) KFCC11519P 20111116

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Claims (17)

delete delete delete delete delete delete delete delete delete Issatchenkia fermented purple sweet potato extract to produce an active ingredient for preventing or treating alcoholic liver disease orientalis ) PSP-HAN [Accession No .: KFCC11519P]. A method for producing a purple sweet potato fermentation broth showing an activity for preventing or treating alcoholic liver disease, comprising fermenting the purple sweet potato extract with Issatchenkia orientalis PSP-HAN [Accession No .: KFCC11519P] of claim 10.
The method of claim 11, wherein the purple sweet potato extract is a method of producing a purple sweet potato fermentation broth showing the activity of preventing or treating alcoholic liver disease, characterized in that the extract of hot water extract or a lower alcohol solution of 2 to 4 carbon atoms.
The method of claim 12, wherein the hot water extract is a method for producing a purple sweet potato fermentation broth showing an alcoholic liver disease prevention or treatment activity, characterized in that extracted for 0.5 to 24 hours at 80 ~ 105 ℃.
The purple sweet potato extract according to claim 11, wherein the purple sweet potato extract is inoculated with Issatchenkia orientalis and then fermented at 25 to 40 ° C. for 20 to 100 hours. Method of producing sweet potato fermentation broth.
The method of claim 11, wherein the purple sweet potato is a sweet potato of any one or more varieties selected from Yamakawamurasaki, yeonjami, Borami and Shinjami production of purple sweet potato fermentation broth showing the activity of preventing or treating alcoholic liver disease Way.
A pharmaceutical composition for preventing or treating alcoholic liver disease, comprising a fermentation broth obtained by fermenting a purple sweet potato extract with Issatchenkia orientalis PSP-HAN [Accession No .: KFCC11519P] of claim 10 as an active ingredient.
A health functional food for preventing or improving alcoholic liver disease, which comprises a fermentation broth obtained by fermenting purple sweet potato extract with Issatchenkia orientalis PSP-HAN [Accession No .: KFCC11519P] of claim 10 as an active ingredient.

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