KR20220079352A - Anti-oxidant composition comprising the extract of soybean hull as an effective component - Google Patents

Abstract

The present invention relates to an antioxidant composition containing an extract of soybean ( Glycine max ) hull (hereinafter also referred to as 'soybean hull') extract as an active ingredient, and more specifically, hot water or It relates to a composition having a strong antioxidant ability containing an ethanol extract as an active ingredient. The soybean husk extract as an active ingredient of the antioxidant composition of the present invention has a strong scavenging ability against active cation radicals and nitrite, as demonstrated through the examples of the present specification. The active ingredient is processed into various forms such as extracts, powders, pills, and tablets, and is a very useful invention to be applied to pharmaceutical compositions, health functional foods, cosmetic compositions, etc. having antioxidant activity.

Description

Antioxidant composition containing soybean hull extract as an active ingredient {ANTI-OXIDANT COMPOSITION COMPRISING THE EXTRACT OF SOYBEAN HULL AS AN EFFECTIVE COMPONENT}

The present invention relates to an antioxidant composition containing an extract of soybean ( Glycine max ) hull (hereinafter also referred to as 'soybean hull') extract as an active ingredient, and more specifically, hot water or It relates to a composition having a strong antioxidant ability containing an ethanol extract as an active ingredient.

The human body that breathes oxygen cannot avoid oxidative stress, and this oxidative stress is manifested by the accumulation of reactive oxygen species. In other words, when the final electron acceptor and oxygen react during the metabolic process of organic matter and are converted to water (H ₂ O), cytotoxicity does not appear, but hydrogen peroxide (H ₂ O ₂ ), superoxide (O ₂ ·) When , hydroxy radicals (OH·), etc. are generated, the activated oxygen species maintain a high concentration of radicals in the cell, causing damage to proteins or DNA, and various oxidative stresses, which eventually damage the cell structure and damage the cells. Loss of function or deterioration (Choi et al. 2008. J. Korean Soc. Food Sci. Nutr. 37: 276-281). In addition, recent research results show that oxidative stress is directly related to the formation of blood clots related to cardiovascular disease (Bijak M et al., 2012, Thrombosis Res. 130, 123-128), and the structural structure of intravascular fibrinogen caused by oxidative stress. It is known that changes can promote thrombus formation and inhibit blood flow, leading to thrombosis (Martinez M et al., 2013. Free Radical Biol. Med. 65, 411-418). According to a recent report, it has been confirmed that oxidative stress in vivo is associated with carcinogenesis, aging, dementia, inflammation, and various cardiovascular diseases (Zhang et al., 2013. Chines J. Cancer biotheraphy, 20. 153). -158; Cho Min-hyun, 2014. J. Korean Soc. Pediatr. Nephrol. 18: 7-12: Shin Jeong-hee et al., 2012. Journal of the Korean Society of Industry-Academic Technology 13: 3561-3569). Therefore, it is determined that, while ensuring stability, natural ingredients capable of removing various active radicals can be expected to significantly help health.

To date, various substances capable of scavenging various reactive oxygen species and radical substances have been developed as antioxidants, and vitamin C, vitamin E, BHT (butylated hydroxy toluene) and BHA (butylated hydroxy anisole) are representative food additives, It is used as a health functional food and medicine. However, side effects of BHT and BHA, which are chemically synthesized antioxidants showing strong antioxidant power, have been reported (Ha, Hyeon-Ju et al., 2014. Korean J. Culinary Res. 20: 16-26; Choe and Yang, 1982. Korean J. Food Sci. Technol. 14: 283-288), it was necessary to develop a strong natural antioxidant that can replace chemically synthesized antioxidants. (Ahn et al., 2009. Korean J. Microbiol. Biotechnol. 37: 266-272; Kim et al., 2009. Korean J. Microbiol. Biotechnol. 37: 133-139).

Meanwhile, soybeans ( Glycine max ) are known to be native to the entire Korean Peninsula, including Manchuria, and are widely cultivated worldwide as a source of vegetable protein and fat. Currently, domestic consumption of soybeans in Korea is more than 1.4 million tons per year, of which 900 to 1 million tons are used for milking and feed, and about 450,000 tons are eaten, while domestic soybean production is 150,000 tons a year, so the self-sufficiency rate of edible beans is 33%. are dependent on imports for more than 90%.

Soybeans contain about 40% protein, 30% carbohydrates, and 20% fat, as well as various physiologically active substances including isoflavones, anthocyanins, and saponins, so the demand for health functional foods is gradually increasing. are doing On the other hand, although mature soybean seeds contain a large amount of various functional ingredients, many non-nutritive elements are also included. Major non-nutritive factors include lipoxygenase enzyme protein that causes fishy smell by participating in the oxidation process of unsaturated fatty acids, and Kunitz trypsin inhibitor, which causes indigestion by inhibiting the action of trypsin There is a protein, and in the case of a human, raffinose and stachyose, which cannot be decomposed and absorbed in the body, are representative. In addition, raw soybeans contain hemagglutinin and soyin, so infants with weak digestive enzymes, people with weak gastric juice and pancreatic function, and people who consume a lot of fats and oils consume raw soybeans, causing diarrhea and It causes digestive problems and headaches. Therefore, because soybeans cannot be eaten raw due to their unique fishy taste, they must be heated or subjected to microbial fermentation to destroy or remove non-nutritive factors and toxic components. However, fermented soybean products such as soybean paste, cheonggukjang, and natto are difficult for the general public to easily manufacture and use, and there is a problem that the taste is not high due to the peculiar taste and odor during fermentation. Therefore, non-fermented heated products of soybeans are more easily manufactured, distributed, and used than fermented products, and there are representative soymilk, tofu, and isolated soybean protein products.

Typically, soybean hulls are removed from the blood after heating soybeans, and the removed soybean hulls account for about 12% of the total soybeans. Soybean hulls have been partially used for livestock feed, but most of them are being discarded without specific uses.

For research on soybean hull, development of an oxidative biodegradable bioplastic film using agricultural by-products such as jade hull, soybean hull, rice hull and wheat hull (Youngseon Yoo et al., 2014, Clean technology 20: 205-211), functional ingredients using by-products of soybean processing Characteristics and applications of soybean hull fraction (protein, fat, carbohydrate, ash, etc.) using separation and physiological activity (Jumseon Park, 2007. Chungnam National University doctoral thesis), ultra-fine grinding/air classification Journal of Science 28: 506-513), research on the development of a fermented feed using soybean hull that improved nutrition by fermenting soybean hull with Aspergillus niger (Yanghee Lee et al., 1972, Journal of the Korean Nutrition Society 5:33-42), using microwave The effect of soybean hull addition on the quality of instant beaksulgi production (Seunghee Kim et al., 2010, Journal of the Korean Society for Food Storage and Distribution 17: 334-342), and the effect of dietary fiber isolated from soybean hull on the improvement of diarrhea (Lim Ji-hyeok et al., 2007. Journal of Food Science and Technology 39: 588-592), evaluation of the effectiveness of special nutritional food using soybean dietary fiber (Han Jae-heum et al., 2008. CNU Journal of Agricultural Science 35: 41-51) and In Vitro ruminant fermentation characteristics and The effect of replacing corn and soybean meal through an appropriate mixing level (Chung Park et al., 2011, Journal of Animal Resources and Science 53: 441-450) is known. However, to date, studies on useful components and useful physiological activities of soybean hull are very limited, and the strong active cation scavenging activity and nitrite scavenging ability of soybean hull extract are not known.

Patents related to soybean hull include “soybean hull peeling device” in Korean Patent No. 10-0507711, and “fried flour containing flour coated with soybean hull fine powder and manufacturing method thereof” in Korean Patent No. 10-0708229 , “Fermentation method of soybean hull or black scalp using microorganisms of the genus Bacillus and composition containing fermented product of soybean hull or black scalp” in Korean Patent Registration No. 10-0710275, and “food” in Korean Patent Registration No. 10-1525658 A composition for a biomass film using wheat or soybean hull as a by-product and a biomass film using the same”, Republic of Korea Patent No. 10-1028119 discloses “a rumen-enhanced antibacterial fermented feed composition and manufacturing method”, disclosed in the Republic of Korea Patent No. 10-2007-0019265 discloses “High-fiber soybean food containing soybean hull and non-fat ingredients and manufacturing method thereof” and Korean Patent Publication No. 10-2001-0047621 discloses “Soybean extract with physiological activity” have. However, there are no patents or research results on soybean hull extract showing strong cation scavenging activity and nitrite scavenging activity.

KR 10-2001-0047621 A

Separation and physiological activity of functional ingredients using by-products of soybean processing, Park Jeom-seon, 2007. Chungnam National University Ph.D. thesis

The present invention has been devised to solve the problems of the prior art as described above, and an object to be solved in the present invention is to provide an antioxidant composition containing soybean husk extract as an active ingredient.

In order to solve the above problems, the present invention provides an antioxidant composition containing soybean ( Glycine max ) bark extract as an active ingredient.

The soybean hull extract is preferably a soybean hull ethanol extract or a soybean hull hot water extract.

The composition is preferably selected from the group consisting of pharmaceutical compositions, health functional foods and cosmetic compositions.

The soybean hull extract as an active ingredient of the antioxidant composition of the present invention has a strong scavenging ability against active cation radicals and nitrite, as demonstrated through the Examples of the present specification. The active ingredient is processed into various forms such as extracts, powders, pills, tablets, etc., and is a very useful invention to be applied to pharmaceutical compositions having antioxidant activity, health functional foods, cosmetic compositions, and the like.

1 is a photograph showing soybeans, soybean hulls and powder after drying soybean hulls used in the present invention.
2 is an evaluation diagram of antioxidant activity at various concentrations of hot water extract and ethanol extract of soybean hull and soybean (●: ethanol extract, ○: hot water extract, ▼: vitamin C).

Hereinafter, the present invention will be described in detail.

The inventors of the present invention recovered soybean hulls by cleaning, drying, cracking, and dehulling soybeans in order to test the antioxidant efficacy of soybean hulls, which are mostly discarded as agricultural waste. After heating and drying, hot water and ethanol extracts of soybean hull were prepared by a certain method. It was attempted to utilize the soybean hull extract as an antioxidant composition by confirming that it had excellent thermal stability and acid stability while exhibiting no hemolytic activity.

Specifically, the present inventors prepared an extract of dried and pulverized soybean hull to develop an antioxidant composition using soybean hull, a by-product removed in the manufacturing step of soybean processed food, and evaluated their scavenging ability against various active radicals. , The strong active cation scavenging activity and nitrite scavenging activity were confirmed in the hot water and ethanol extracts of soybean hull compared to the hot water and ethanol extracts of soybean. In addition, it was confirmed that the soybean hull extract did not show haemolytic activity against human red blood cells, so it could actually be used as a pharmaceutical composition for antioxidant use, a health functional food, and a cosmetic composition.

More specifically, in order to develop an antioxidant composition using soybean hulls, the present inventors selected the most grown and organically grown soybeans in Korea (cleaning), drying (tempering), crushing (cracking), and dehulling (dehulling). The soybean hull was recovered through a process such as a soybean hull, and in the dehulling process, the soybean hull produced from the dried and crushed soybean was separated using a pneumatic aspirator. Silver was removed using a vibrating screen. Thereafter, the soybean hulls were dried at 80° C. for 3-4 hours and pulverized to 50 mesh. A hot water extract and an ethanol extract were prepared from the pulverized soybean hulls, respectively, and the scavenging ability of these extracts against various active radicals was evaluated. did On the other hand, as a control, soybeans without soybean hulls were dried at 80°C for 3-4 hours and pulverized into 50 mesh. The scavenging ability was evaluated.

As a result, strong ABTS cation scavenging activity and nitrite scavenging activity were confirmed in hot water and ethanol extracts of soybean hull. In particular, after evaluating the antioxidant activity of hot water extract and ethanol extract of soybean hull extract at various concentrations, RC ₅₀ indicating 50% scavenging activity was calculated. As a result, the RC ₅₀ of ABTS cation scavenging activity and nitrite scavenging activity of the ethanol extract was 113.1 and It was confirmed to be 27.7 μg/ml, and the RC ₅₀ of the ABTS cation scavenging ability and nitrite scavenging ability of the hot water extract were 103.8 and 45.4 μg/ml, respectively. On the other hand, the RC ₅₀ of ABTS cation scavenging activity and nitrite scavenging activity of whole soybean hot water and ethanol extract were 1637.3 and 1673.7 μg/ml, respectively, and the RC ₅₀ of ABTS cation scavenging activity and nitrite scavenging activity of hot water extract were 1833.3 and 339.3 μg/ml, respectively. was confirmed as

In addition, it was confirmed that the hot water extract among the soybean hull extracts did not show hemolytic activity against human red blood cells and thus did not induce acute toxicity. % hemolysis. However, considering that amphotericin B, which is used as an anticancer and antifungal agent with hemolytic activity, exhibits a hemolytic activity of 28.1% at a concentration of 0.003 mg/ml, it is judged that soybean hull extract will not cause a separate problem of red blood cell hemolysis.

Accordingly, the present invention provides an antioxidant composition containing soybean ( Glycine max ) husk extract as an active ingredient.

The soybean hull extract is preferably a soybean hull ethanol extract or a soybean hull hot water extract.

The composition is preferably selected from the group consisting of pharmaceutical compositions, health functional foods and cosmetic compositions.

The soybean hull can be recovered from soybean hulls of various varieties, but preferably soybean hulls of the 'Seonpung' variety.

Hereinafter, the method for preparing the soybean hull extract of the present invention and the efficacy test will be described in more detail.

The present invention comprises the steps of harvesting organic soybeans, removing foreign substances; tempering and cracking the soybeans; recovering soybean hulls from crushed soybeans using a pneumatic aspirator and a vibrating screen; drying soybean hulls and pulverizing them to 50 mesh; preparing an extract of ground soybean hull; Antioxidant activity evaluation step of the extract; and a step of examining the stability of the active substance of the soybean hull extract exhibiting antioxidant activity.

The organic solvent used for soybean hull extraction of the present invention is water (cold water, hot water), alcohol, anhydrous or hydrous lower alcohol having 1 to 4 carbon atoms (methanol, ethanol, alcohol, propanol, ethyl acetate, etc.), the lower alcohol and water A mixed solvent and the like can be used, and hot water or 95% ethanol extraction is preferable.

The soybean hull extract of the present invention may be prepared as a powder through a conventional powdering process such as drying under reduced pressure, freeze drying, or spray drying. They are not degraded by various degrading enzymes in plasma, and they maintain activity even at 100°C heat treatment and at pH 2 in the human stomach.

The soybean hull extract of the present invention exhibits strong antioxidant activity and can be used as a material for pharmaceutical compositions, health functional foods, and cosmetic compositions for the prevention or treatment of various diseases related to oxidative stress.

The various diseases related to the oxidative stress include, for example, various cancers, myocardial infarction, stroke, hypertension, dementia, Parkinson's disease, Alzheimer's disease, Huntington's disease, diabetes, hyperlipidemia, angina, inflammatory bowel disease, cardiovascular disease, skin wrinkles, erythema, Freckles, skin pigmentation, atopic dermatitis, acne, etc. may be mentioned, but are not limited thereto.

The pharmaceutical composition of the present invention is formulated in various forms such as oral formulations such as powders, granules, tablets, capsules, suspensions, emulsions, syrups and aerosols, and injections of sterile injection solutions according to conventional methods for each purpose of use. and can be administered orally, or administered through various routes including intravenous, intraperitoneal, subcutaneous, rectal, topical administration, and the like. The pharmaceutical composition may further include a carrier, excipient or diluent, and examples of suitable carriers, excipients or diluents that may be included include lactose, dextrose, sucrose, sorbitol, mannitol, xylitol, erythritol, maltitol, starch. , acacia gum, alginate, gelatin, calcium phosphate, calcium silicate, cellulose, methyl cellulose, amorphous cellulose, polyvinyl pyrrolidone, water, methylhydroxybenzoate, propylhydroxybenzoate, talc, magnesium stearate and mineral oil, etc. can be heard In addition, the pharmaceutical composition of the present invention may further include a filler, an anti-aggregant, a lubricant, a wetting agent, a fragrance, an emulsifier, a preservative, and the like.

The pharmaceutical composition of the present invention is administered in a pharmaceutically effective amount. In the present invention, "pharmaceutically effective amount" means an amount sufficient to treat a disease with a reasonable benefit/risk ratio applicable to medical treatment, and the effective dose level is determined by the type, severity, activity of the drug, and the type of disease in the patient; Sensitivity to the drug, time of administration, route of administration and excretion rate, duration of treatment, factors including concomitant drugs and other factors well known in the medical field. The pharmaceutical composition of the present invention may be administered as an individual therapeutic agent or in combination with other therapeutic agents, may be administered sequentially or simultaneously with conventional therapeutic agents, and may be administered single or multiple. In consideration of all of the above factors, it is important to administer an amount that can obtain the maximum effect with a minimum amount without side effects, which can be easily determined by those skilled in the art.

In a preferred embodiment, the effective amount of the soybean hull extract having excellent antioxidant activity in the pharmaceutical composition of the present invention may vary depending on the age, sex, and weight of the patient, and is generally 1 to 5,000 mg per body weight, preferably 100 to 5,000 mg per body weight. 3,000 mg may be administered daily or every other day, or divided into 1 to 3 times a day. However, since it may increase or decrease depending on the route of administration, disease severity, sex, weight, age, etc., the dosage is not intended to limit the scope of the present invention in any way. The pharmaceutical composition of the present invention may be administered to a subject through various routes. Any mode of administration can be envisaged, for example, by oral, rectal or intravenous, intramuscular, subcutaneous, intrauterine dural or intracerebroventricular injection. In the present invention, "administration" means providing a predetermined substance to a patient by any suitable method, and the administration route of the pharmaceutical composition of the present invention is oral or parenteral through all general routes as long as it can reach the target tissue. It can be administered orally. In addition, the composition of the present invention may be administered using any device capable of delivering an active ingredient to a target cell. In the present invention, the "subject" is not particularly limited, but includes, for example, humans, monkeys, cattle, horses, sheep, pigs, chickens, turkeys, quails, cats, dogs, mice, rats, rabbits or guinea pigs. and preferably a mammal, more preferably a human.

In addition, the health functional food of the present invention can be used in a variety of ways, such as foods and beverages effective for the improvement of various diseases related to oxidative stress. Foods containing the soybean hull extract having excellent antioxidant activity of the present invention include, for example, various foods, beverages, gum, tea, vitamin complexes, health supplements, and the like, and include powders, granules, tablets, capsules, or beverages. available in the form. In general, the active extract of the present invention may be added in an amount of 0.01 to 15% by weight of the total food weight, and the health beverage composition may be added in a proportion of 0.02 to 10g, preferably 0.3 to 1g, based on 100ml. The health functional food of the present invention may contain, as an additional ingredient, in addition to containing the above compound as an essential ingredient in the indicated ratio, a food pharmaceutically acceptable food supplement additive, such as natural carbohydrate and various flavoring agents. Examples of the natural carbohydrate include monosaccharides such as glucose and fructose, disaccharides such as maltose and sucrose, and common sugars such as polysaccharides such as dextrin and cyclodextrin, and sugar alcohols such as xylitol, sorbitol, and erythritol. As the flavoring agent, thaumatin, rebaudioside A, glycyrrhizin, saccharin, aspartame, and the like may be used. The ratio of the flavoring agent is generally about 1 to 20 g, preferably about 5 to 12 g per 100 ml of the health functional food of the present invention. In addition to the above, the health functional food of the present invention includes various nutrients, vitamins, minerals, flavoring agents such as synthetic and natural flavoring agents, colorants and thickeners, pectic acid and its salts, alginic acid and its salts, organic acids, protective colloids It may contain thickeners, pH adjusters, stabilizers, preservatives, glycerin, alcohol, carbonation agents used in carbonated beverages, and the like. In addition, the health functional food of the present invention may contain natural fruit juice, fruit juice for the production of beverages, vegetable beverages, and the like. These components may be used independently or in combination. The proportion of these additives is generally selected in the range of 0.01 to about 20 parts by weight per 100 parts by weight of the active extract of the present invention.

The cosmetic composition of the present invention may be prepared by using the soybean hull extract having excellent antioxidant activity of the present invention in the form of, for example, essence, nourishing cream, body lotion, conditioner, shampoo, body cleanser, lotion, skin, mask pack, etc. , but not limited thereto. Preparation in the above form can be easily carried out according to various manufacturing processes known to those skilled in the art.

Hereinafter, a specific method of the present invention will be described in more detail through examples. The following examples are only one preferred embodiment of the present invention, and the scope of the present invention is not limited to the scope of the following examples.

[Example]

Example 1: Preparation of soybean hull extract and analysis of components of the extract

In 2019, organically grown soybeans were harvested in Bonghwa-gun, Gyeongbuk, and soybean hulls were recovered through cleaning, tempering, cracking, and dehulling processes. In the dehulling process, dried and crushed soybeans The soybean hull produced in the above was separated and recovered using a pneumatic aspirator, and at this time, the soybean kernel fragments separated along with the soybean hull were removed using a vibrating screen. Thereafter, the soybean hulls were dried at 80° C. for 3-4 hours and pulverized into 50 mesh, and a hot water extract and an ethanol extract were prepared from the pulverized soybean hulls, respectively.

Meanwhile, the moisture content and component analysis results of the soybeans and soybean hulls used are shown in Table 1.

[Table 1] Moisture content and component characteristics of soybeans and soybean hulls used in the experiment

As shown in Table 1, soybean hulls showed 1/3 of the protein content and very low lipid content compared to soybeans, and the carbohydrate content was more than twice that of soybeans. At this time, the carbohydrate of soybean hull contained more than 60% of non-digestible fiber including ligrin.

On the other hand, the hot water extract of soybean hull was prepared by adding distilled water 20 times the weight of the sample, extracting it once at 100° C. for 1 hour, collecting the extract, filtering, and concentrating under reduced pressure to prepare a powder, and the ethanol extract 20 times the amount of ethanol (95%, Deoksan, Korea) was added to the silver sample weight, extracted twice at room temperature for 8 hours, the extract was collected, filtered, and concentrated under reduced pressure to prepare a powder. The results of each extraction efficiency and component analysis of the extract are shown in Table 2. The content of total polyphenols, total flavonoids, total sugars and reducing sugars was determined by component analysis. For the total polyphenol content, 50 μl of Folin-ciocalteau and 100 μl of Na ₂ CO ₃ saturated solution were added to 400 μl of the extraction sample solution, and the absorbance was measured at 725 nm after standing at room temperature for 1 hour. As a standard reagent, tannic acid was used. For the total flavonoid content, each sample was extracted with methanol for 18 hours, and 4 ml of 90% diethylene glycol was added to 400 μl of the filtered extraction sample solution, 40 μl of 1 N NaOH was added again, and the absorbance was measured at 420 nm after reaction at 37° C. for 1 hour. As a standard reagent, rutin was used. Reducing sugar was quantified by DNS method and total sugar was quantified by phenol-sulfuric acid method.

[Table 2] Extraction efficiency and useful component analysis of soybean hull extract

As shown in Table 2, the hot water extraction efficiency of soybeans was very high at 27.1%, but the ethanol extraction efficiency was low at 2%. appeared low. However, as a result of analysis of the total polyphenol and total flavonoid content, the soybean hull extract showed significantly higher polyphenol content than the soybean extract. In particular, the hot water extract of soybean hull showed a polyphenol content of 0.1 mg/g, whereas the hot water extract of soybean hull had a polyphenol content of 24.6 mg/g. showed a significantly higher content. In addition, high content was shown in both hot water and ethanol extracts of soybean hull, indicating that soybean hull contains various polyphenols and flavonoids, water-soluble and fat-soluble.

On the other hand, the total sugar content was high in the ethanol extract from soybean hull and the ethanol extract from soybean, indicating that the components of soybean and soybean hull were significantly different. Due to these results, it was expected that soybean hull would exhibit various useful physiological activities compared to soybean, and it was confirmed that soybean hull has a high potential for development as a functional material.

Example 2: Antioxidant activity of soybean hull and soybean extract

The antioxidant activity of soybean hull and soybean extracts of Example 1 was evaluated, and the extract was dissolved in dimethylsulfoxide (DMSO) and diluted to an appropriate concentration to obtain DPPH (1,1-diphenyl-2-picryl hydrazyl) anion radical scavenging ability, ABTS[2] ,2-azobis (3-ethylbenzo thiazoline-6-sulfonate)] was used to measure cation radical scavenging ability, nitrite scavenging ability and reducing power. First, in the case of DPPH scavenging ability, 380 μl of 2x10 ^-4 M DPPH solution dissolved in 99.5% ethanol was added to 20 μl of sample diluted to various concentrations, mixed, and reacted at 37° C. for 30 minutes, followed by a microplate reader (Asys Hitech, Expert 96) at 516 nm. , Asys Co., Austria) was used to measure the absorbance. The DPPH radical scavenging ability was expressed as a percentage of the sample-added group and non-additive group. In the case of ABTS scavenging activity, 5ml of 7mM ABTS (Sigma Co., USA) and 88ml of 140mM potassium persulfate were mixed, and light was blocked at room temperature for 16 hours to form ABTS cations. Then, this solution was mixed with ethanol so that the absorbance value at 414nm was 1.5. diluted with After mixing 190 ml of the prepared diluted solution and 10 ml of samples prepared at various concentrations, reacting at room temperature for 6 minutes, absorbance was measured at 734 nm, and the ABTS radical scavenging ability was calculated by the following equation.

ABTS radical scavenging ability (%) = [(C-S)/C] x 100,

C: absorbance upon addition of solvent control DMSO,

S: Absorbance upon sample addition.

On the other hand, in the case of measuring nitrite scavenging ability, the sample solution was added to a nitrite solution (1 mM), and 0.1N HCl was added thereto to adjust the pH to 1.2, and after reacting at 37°C for 1 hour, Griess reagent (Sigma Co., USA) was added and mixed. After leaving at room temperature for 15 minutes, absorbance was measured at 520 nm to measure the amount of remaining nitrite. The nitrite scavenging ability (%) was calculated by the following formula.

NSA (%) = [1-(A-C)/B] x 100,

A: Absorbance after adding the sample to 1 mM nitrite solution and reacting for 1 hour,

B: absorbance of 1 mM nitrite solution,

C: Absorbance of the extract sample.

In the above antioxidant experiment, vitamin C (Sigma Co., USA) was used as a control, and DMSO was used as a solvent control. Each activity evaluation was expressed as the average and deviation of the experiment repeated three times, and finally, each antioxidant activity was expressed as the concentration of the extract required for 50% radical scavenging activity (RC ₅₀ ).

On the other hand, in the case of reducing power evaluation, it was measured by modifying the method of Oyaizu et al. (Sunmi Ahn et al., 2011. J. Life Sci. 21: 576-583). 2.5ml of 0.2M sodium phosphate buffer (pH 6.6) and 2.5ml of 10% potassium ferricyanide were added to 2.5ml of sample dissolved in ethanol, and after reacting at 50℃ for 20 minutes, 2.5ml of 10% trichloroacetic acid was added to terminate the reaction and centrifuged at 4,000 rpm for 10 minutes to recover the supernatant. The recovered supernatant was diluted two-fold with distilled water, mixed with a freshly prepared 0.1% ferric chloride solution at a ratio of 5:1 (v/v), and absorbance was measured at 700 nm to evaluate. The measurement results of each antioxidant power are shown in Table 3.

[Table 3] Antioxidant activity of soybean hull and soybean extract

As shown in Table 3, when the antioxidant activity was evaluated using the soybean hull extract at a concentration of 0.5 mg/ml, the ABTS cation scavenging ability and nitrite scavenging ability were significantly superior to those of the soybean extract. This strong scavenging ability was shown in both hot water and ethanol extracts, confirming that soybean hull contains a large amount of (-)-charged antioxidants and nitrite-removing substances capable of scavenging active cations.

Therefore, the concentration of soybean hull and soybean extract was varied, the antioxidant activity was evaluated, and RC ₅₀ indicating 50% scavenging ability was calculated, and the results are shown in Table 4 and FIG. 2 .

[Table 4] RC of soybean hull and soybean extract ₅₀ (Concentration evaluation showing 50% active radical scavenging ability)

As shown in Table 4 and Figure 2, the RC50 of the ABTS cation scavenging activity and the nitrite scavenging activity of the soybean hull ethanol extract were 113.1 and 27.7 μg/ml, respectively, and the RC50 of the ABTS cation scavenging activity and the nitrite scavenging activity of the hot water extract were 103.8 and 103.8 and nitrite scavenging activity, respectively. It was found to be 45.4 μg/ml. On the other hand, the RC50 of ABTS cation scavenging activity and nitrite scavenging ability of soybean hot water and ethanol extract were 1637.3 and 1673.7 μg/ml, respectively, and the RC ₅₀ of ABTS cation scavenging ability and nitrite scavenging ability of hot water extract were 1833.3 and 339.3 μg/ml, respectively. became Therefore, soybean hull extract exhibits the same effect at 1/14 concentration (ethanol extract) or 1/18 concentration (hot water extract) than soybean extract for scavenging active cations. It was confirmed that the same effect was exhibited at the concentration of 62 (ethanol extract) or 1/7 concentration (hot water extract). The above results suggest that it is possible to use soybean hull extract as a commercial antioxidant composition.

Example 3: Human hemolytic activity of soybean hull and soybean extract

Although soybean hull cannot be eaten raw due to its unique fishy taste, it has been used as livestock feed since the past. In particular, it can be used as an excellent food material when it increases the digestibility and removes harmful microorganisms through heat treatment such as steaming. Therefore, it is known that soybean hull extract does not show any toxicity. In the present invention, human red blood cell hemolytic activity was evaluated to evaluate the acute toxicity of soybean hull extract, and the results are shown in Table 5. At this time, the hemolytic activity was evaluated according to the previous report (Miseon Kim et al., 2014. J. Life Sci. 24: 515-521), and simply, 100 μl of human red blood cells washed three times with PBS was added to a 96-well microplate, and various concentrations were added. 100 μl of the sample solution was added, followed by reaction at 37° C. for 30 minutes. Then, the reaction solution was centrifuged (1,500 rpm) for 10 minutes, and 100 μl of the supernatant was transferred to a new microtiter plate. . DMSO (2%) was used as a solvent control of the sample, and Triton X-100 (1mg/ml) was used as an experimental control for hemolysis of red blood cells. Hemolytic activity was calculated using the following formula.

[Table 5] Human hemolytic activity of soybean hull and soybean extract

As a result, as shown in Table 5, first, DMSO and water used as controls had no hemolytic activity, and it was confirmed that triton X-100 hemolyzed red blood cells 100% at a concentration of 1 mg/ml. Soybean ethanol and hot water extracts did not show hemolytic activity up to a concentration of 1 mg/ml. Meanwhile, among soybean hull extracts, the hot water extract showed no hemolytic activity at a concentration of 1 mg/ml, and the ethanol extract did not show hemolytic activity at a concentration of 0.5 mg/ml, and showed a hemolysis of 27.5% at a concentration of 1 mg/ml. However, considering that amphotericin B, which is used as an anticancer and antifungal agent with hemolytic activity, shows 28.1% of hemolysis at a concentration of 0.003 mg/ml, soybean hull extract does not appear to cause a separate hemolysis problem.

Example 4: Evaluation of Plasma, Acid and Thermal Stability of Soybean Husk Extract

Plasma stability, thermal stability, and acid stability with respect to antioxidant activity of the soybean hull extract obtained in Example 1 were confirmed. The samples showed high stability because no significant decrease in antioxidant activity of ABTS cation scavenging activity and nitrite scavenging activity was observed even after 1 hour heat treatment at 100° C., 1 hour treatment at pH 2 (0.01M HCl), and 1 hour treatment in plasma. it was Therefore, the soybean hull extract is expected to maintain excellent antioxidant activity during the digestion and absorption process and food manufacturing process.

[R&D project]

Department name: Ministry of Education, Science and Technology

Research and management institution: Andong University LINC+ project group

Research project name: Action Learning

Research project name: Development of high-quality livestock/insect feed through chemical and biological treatment of soybean hull, a by-product of agriculture

Contribution rate: 1/1

Organized by: Andong University Industry-University Cooperation Foundation

Research period: 2020.09.01~2020.12.28

Claims (3)

An antioxidant composition containing soybean ( Glycine max ) husk extract as an active ingredient. The antioxidant composition according to claim 1, wherein the soybean hull extract is a soybean hull ethanol extract or a soybean hull hot water extract. The antioxidant composition according to claim 1 or 2, wherein the composition is selected from the group consisting of a pharmaceutical composition, a health functional food and a cosmetic composition.

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