KR101976999B1 - The method for preparing a Platycodon grandiflorum extract using a complex enzyme and a composition for improving antioxidant, whitening or wrinkles containing the same - Google Patents

Abstract

The present invention relates to a method for manufacturing balloon flower extracts with improved extraction yield and the content of saponin through a composite enzyme treatment, and to a composition for antioxidation, skin whitening or wrinkle reduction containing the extracts. More specifically, the method for manufacturing balloon flower extracts comprises the steps of: cleaning, drying and pulverizing balloon flowers; treating carbohydrase to the pulverized balloon flowers; extracting the enzyme treated balloon flowers at 40-60°C for 10-15 hours; and depressurizing and concentrating the extracts. According to the present invention, an extraction yield is improved to manufacture balloon flower materials and the content of balloon flower saponin in the extracts can be increased. In addition, the balloon flower extracts manufactured by the manufacturing method of the present invention have antioxidation, skin whitening and wrinkle reduction activities, thereby being used as functional materials for a cosmetic composition.

Description

TECHNICAL FIELD The present invention relates to a method for preparing a bellflower extract using a complex enzyme and a composition for antioxidant, whitening or wrinkle improvement containing the same,

The present invention relates to a method for producing a bellflower extract having enhanced extraction yield and saponin content through complex enzyme treatment, and a composition for antioxidant, whitening or wrinkle improvement containing the extract.

In general, Platycodon grandiflorum is a perennial plant belonging to the lantern family, which is also called kikyou in Oriental medicine. It is native to Korea, China, and Japan. Bellflower is a medicinal herb that has been used in respiratory diseases such as diarrhea, genomes, bronchitis and asthma, and various pharmacological actions are known. For example, triterpenoid saponins among pharmacological agents of phloem phlox are antioxidant, anti-inflammatory, anti-inflammatory, anti-inflammatory, anti-inflammatory, anti-inflammatory, anti- And gastric juice secretion inhibiting action, anticholinergic action of lowering blood pressure by expanding blood vessels, hypoglycemic action, and improvement of cholesterol metabolism. In addition, the extract of the bellflower inhibits the mold aflatoxin, and the inulin fraction is known to have potent antitumor activity against phagocytosis and solid and multiple cancers.

Since such a bellflower contains a physiologically active substance useful in vivo, the physiologically active substance can be advantageously extracted by separate processing. However, bellflower is a medicinal plant that has been used for a long time as a raw material for health functional food, and the development of the usage method is less than known. Particularly, in the extraction of the active ingredient of the bellflower, the hot water extraction method is mainly used, but since the extraction yield is low, there is a problem that the residue remaining after the extraction is discarded with the active ingredient contained therein, making economical bellflower extract difficult to produce.

Patent Publication No. 10-2011-0108687 Patent Publication No. 10-1997-0066516 Patent Publication No. 10-2015-0007810

It is an object of the present invention to provide a method for producing a bellflower extract, which comprises a step of complex enzyme treatment to produce a bellflower extract having enhanced extraction yield and saponin content.

Another object of the present invention is to provide a composition having antioxidant, whitening or wrinkle-reducing effect, which comprises the bellflower extract prepared by the above-mentioned production method.

 In order to accomplish the object of the present invention, the present invention provides a method for cleaning a bellflower, comprising: washing, drying and crushing a bellflower; Treating the crushed bellflower with carbohydrase; Extracting the enzyme treated bellflower at a temperature of 40 to 60 DEG C for 10 to 15 hours; And a step of decompressing and concentrating the extract.

Here, the carbohydrase may be a viscozyme including Arabanase, Cellulase, Beta-glucanase, Hemicellulase, and Xylanase. And termamyl 2X, which contains alpha-amylase.

In addition, the viscose and termaryl 2X are mixed in a ratio of 1: 0.5 to 2.

In addition, the carbohydrase is contained at a concentration of 5% based on the total weight of dried bellflower.

Meanwhile, an embodiment of the present invention provides a cosmetic composition comprising the bellflower extract extracted through the extraction method.

Herein, the cosmetic composition is characterized by containing antioxidant, whitening or wrinkle-improving function.

According to the present invention as described above, it is possible to manufacture a plant material by increasing the extraction yield compared to the existing hot water extraction method, and to improve the content of plant extract of the plant. Also, the bellflower extract prepared according to the production method of the present invention has antioxidative, whitening and wrinkle-reducing activities, and thus can be used as a functional material for a cosmetic composition.

The above and other objects, advantages and novel features of the present invention will become more apparent from the following detailed description when read in conjunction with the accompanying drawings, in which: FIG. It can be understood.

FIG. 1 is a schematic view showing a method for preparing a Plasmid Complex Enzyme-treated extract according to a preferred embodiment of the present invention.
FIG. 2 shows the results of measuring the optimal reaction temperature of a complex enzyme according to a preferred embodiment of the present invention.
FIG. 3 shows the result of measurement of the optimal reaction concentration of a complex enzyme according to a preferred embodiment of the present invention.
FIG. 4 is a graph illustrating the extraction yields of the Plasmid complex enzyme extract and the hot water extract according to a preferred embodiment of the present invention.
FIG. 5 is a graph showing the crude saponin content of Platycodon grandiflorum complex extract and hot water extract according to a preferred embodiment of the present invention.
FIG. 6 is a graph showing the DPPH radical scavenging activity of Doraji complex enzyme extract and hot water extract according to a preferred embodiment of the present invention.
FIG. 7 is a graph showing the ABTS radical scavenging activity of Doraji complex enzyme extract and hot water extract according to a preferred embodiment of the present invention.
FIG. 8 is a graph showing the tyrosinase inhibitory activity of Doraji complex enzyme extract and hot water extract according to a preferred embodiment of the present invention.
FIG. 9 is a graph showing the results of a comparison of elastase inhibitory activities of Plasmodium complexae enzyme extract and hot-water extract according to a preferred embodiment of the present invention.

Hereinafter, the present invention will be described in more detail.

The present invention relates to a method for producing a bellflower extract having enhanced extraction yield and saponin content through complex enzyme treatment, and a composition for antioxidant, whitening or wrinkle improvement containing the extract.

INDUSTRIAL APPLICABILITY According to the present invention, it is possible to produce a broth material economically by improving the extraction yield, and to improve the content of the broth saponin of the extract.

Also, the bellflower extract prepared according to the production method of the present invention has antioxidative, whitening and wrinkle-reducing activities, and thus can be used as a functional material for a cosmetic composition.

Specifically, a method for preparing a Platycodon grandiflorum extract according to an embodiment of the present invention includes washing, drying and pulverizing platycodon; Treating the crushed bellflower with carbohydrase; Extracting the enzyme treated bellflower at a temperature of 40 to 60 DEG C for 10 to 15 hours; And depressurizing and concentrating the extract.

Here, the carbohydrase may be a viscozyme including Arabanase, Cellulase, Beta-glucanase, Hemicellulase, and Xylanase. And termamyl 2X, which contains alpha-amylase, can be used.

The viscose and termaryl 2X may be mixed in a ratio of 1: 0.5 to 2.

In addition, the carbohydrase is contained at a concentration of 5% based on the total weight of the dried bellflower.

Meanwhile, an embodiment of the present invention provides a cosmetic composition comprising the bellflower extract extracted through the extraction method.

The cosmetic composition of the present invention includes antioxidant, whitening or wrinkle-reducing function.

The composition according to the present invention can be manufactured and used in the form of a food composition such as a cosmetic composition, an oral dosage form, a transdermal dosage form, a pharmaceutical form such as an inhaled dosage form, a functional food, a nutritional supplement, a health food or a food additive.

When the composition of the present invention is a cosmetic composition, it is preferable to use a basic cosmetic composition (cleanser, pack, body oil) such as lotion, cream, essence, cleansing foam and cleansing water together with a dermatologically acceptable excipient, , A color cosmetic composition (foundation, lipstick, mascara, makeup base), a hair product composition (shampoo, rinse, hair conditioner, hair gel) and soap.

Such excipients include, but are not limited to, emollients, skin penetration enhancers, colorants, perfumes, emulsifiers, thickeners and solvents. In addition, it may further contain flavors, pigments, bactericides, antioxidants, preservatives, moisturizers and the like, and may include thickeners, inorganic salts and synthetic polymeric substances for the purpose of improving physical properties. For example, when a cleanser and a soap are prepared with the cosmetic composition of the present invention, the above-mentioned Platycodon grandiflorum extract can be easily added to a common cleanser and soap base. In the case of producing a cream, it can be prepared by adding a bellflower extract or a salt thereof to a general underwater type (O / W) cream base. A synthetic or natural material such as a flavor, a chelating agent, a coloring matter, an antioxidant, an antiseptic, and other proteins, minerals, and vitamins for the purpose of improving physical properties may be further added.

The content of the bellflower extract contained in the cosmetic composition of the present invention is not limited thereto, but is preferably 0.001 to 10% by weight, more preferably 0.01 to 5% by weight, based on the total weight of the whole composition. When the content is less than 0.001% by weight, the desired antioxidative, whitening and wrinkle-reducing effects can not be expected. When the content is more than 10% by weight, safety or formability may be difficult.

The bellflower extract of the present invention has antioxidative, whitening and wrinkle-reducing effects and can be used as an effective ingredient of a pharmaceutical composition.

The pharmaceutical composition of the present invention may include a pharmaceutically acceptable salt of a bellflower extract. The term " pharmaceutically acceptable " in the present invention means physiologically acceptable and when administered to humans, it usually means not to carry out an allergic reaction or a similar reaction, Acid addition salts formed by possible free acids are preferred.

The pharmaceutically acceptable salt of the bellflower extract may be an acid addition salt formed using an organic acid or an inorganic acid. The organic acid may be, for example, formic acid, acetic acid, propionic acid, lactic acid, butyric acid, isobutyric acid, trifluoroacetic acid, The present invention also relates to the use of a compound selected from the group consisting of malic acid, maleic acid, malonic acid, fumaric acid, succinic acid, succinic acid monoamide, glutamic acid, tartaric acid, oxalic acid, citric acid, glycolic acid, gluconic acid, ascorbic acid, benzoic acid, phthalic acid, salicylic acid, anthranilic acid, Benzenesulfonic acid, p-toluenesulfonic acid or methanesulfonic acid. The inorganic acid includes, for example, hydrochloric acid, bromic acid, sulfuric acid, phosphoric acid, nitric acid, carbonic acid or boric acid. The acid addition salt may preferably be in the form of a hydrochloride or an acetate, more preferably in the form of a hydrochloride.

The above-mentioned acid addition salts may be prepared by a) directly mixing the bellflowy extract and the acid, or b) dissolving and mixing one of them in a solvent or a water-soluble solvent, or c) placing the bellflower extract in an acid in the solvent or sub- And then mixing them.

Separately, additionally saltable forms include, but are not limited to, the salts of gabapentin, gabapentin, pregabalin, nicotinic acid, adipate, hemimarate, cysteine, acetylcysteine, methionine, arginine, Or aspartate.

In addition, the pharmaceutical composition of the present invention may further comprise a pharmaceutically acceptable carrier.

The pharmaceutically acceptable carrier may further include, for example, a carrier for oral administration or a carrier for parenteral administration. Carriers for oral administration may include lactose, starch, cellulose derivatives, magnesium stearate, stearic acid, and the like. The carrier for parenteral administration may also contain water, suitable oils, saline, aqueous glucose and glycols and the like. In addition, stabilizers and preservatives may be further included. Suitable stabilizers include antioxidants such as sodium hydrogen sulfite, sodium sulfite or ascorbic acid. Suitable preservatives include benzalkonium chloride, methyl- or propyl-paraben and chlorobutanol. Other pharmacologically acceptable carriers may be those listed in the following references (Remington ' s Pharmaceutical Sciences, 19th ed, Mack Publishing Company, Easton, Pa., 1995)

The pharmaceutical composition of the present invention can be administered to mammals including humans by any method. For example, it can be administered orally or parenterally, and parenteral administration methods include, but are not limited to, intravenous, intramuscular, intraarterial, intramedullary, intrathecal, intracardiac, transdermal, subcutaneous, , Intranasal, enteral, topical, sublingual or rectal administration.

The pharmaceutical composition of the present invention can be formulated into oral preparations or parenteral administration preparations according to the administration route as described above. When formulated, one or more buffers (e.g., saline or PBS), antioxidants, bacteriostats, chelating agents (e.g., EDTA or glutathione), fillers, extenders, binders, adjuvants Side), suspending agents, thickening agents, disintegrating agents or surfactants, diluents or excipients.

Solid preparations for oral administration include tablets, pills, powders, granules, solutions, gels, syrups, slurries, suspensions or capsules, etc. These solid preparations can be prepared by incorporating into the pharmaceutical composition of the present invention at least one excipient, , Starch (including corn starch, wheat starch, rice starch and potato starch), calcium carbonate, sucrose, lactose, dextrose, sorbitol, mannitol, xylitol, erythritol maltitol, cellulose , Methyl cellulose, sodium carboxymethyl cellulose and hydroxypropylmethyl-cellulose or gelatin. For example, tablets or tablets may be obtained by combining the active ingredient with a solid excipient, then milling it, adding suitable auxiliaries, and processing the mixture into granules.

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 or syrups. In addition to water or liquid paraffin, which is a simple diluent commonly used, various excipients such as wetting agents, sweeteners, fragrances or preservatives may be included .

In addition, crosslinked polyvinylpyrrolidone, agar, alginic acid, or sodium alginate may optionally be added as a disintegrant, and may further include an anticoagulant, a lubricant, a wetting agent, a flavoring agent, an emulsifying agent and an antiseptic agent .

For parenteral administration, the pharmaceutical compositions of the present invention may be formulated in accordance with methods known in the art in the form of injectable, transdermal and nasal inhalers, together with suitable non-oral carriers. In the case of such injections, they must be sterilized and protected against contamination of microorganisms such as bacteria and fungi. Examples of suitable carriers for injectables include, but are not limited to, solvents or dispersion media containing water, ethanol, polyols (e.g., glycerol, propylene glycol and liquid polyethylene glycol, etc.), mixtures thereof and / or vegetable oils . More preferably, suitable carriers include isotonic solutions such as Hanks' solution, Ringer's solution, phosphate buffered saline (PBS) containing triethanolamine, or sterile water for injection, 10% ethanol, 40% propylene glycol and 5% dextrose Etc. may be used. In order to protect the injection from microbial contamination, various antibacterial and antifungal agents such as parabens, chlorobutanol, phenol, sorbic acid, thimerosal and the like may be further included. In addition, the injections may in most cases additionally include isotonic agents, such as sugars or sodium chloride.

Examples of transdermal dosage forms include ointments, creams, lotions, gels, solutions for external use, pastes, liniments, and air lozenges. In the above, 'transdermal administration' means that the pharmaceutical composition is locally administered to the skin, whereby an effective amount of the active ingredient contained in the pharmaceutical composition is delivered into the skin.

In the case of an inhalation dosage form, the compounds used according to the invention can be formulated into a pressurized pack or a pressurized pack using a suitable propellant, for example dichlorofluoromethane, trichlorofluoromethane, dichlorotetrafluoroethane, carbon dioxide or other suitable gases. It can be conveniently delivered in the form of an aerosol spray from a nebulizer. In the case of a pressurized aerosol, the dosage unit may be determined by providing a valve that delivers a metered amount. For example, gelatin capsules and cartridges for use in an inhaler or insufflator may be formulated to contain a compound, and a powder mixture of a suitable powder base such as lactose or starch. Formulations for parenteral administration are described in Remington's Pharmaceutical Science, 15th Edition, 1975 Mack Publishing Company, Easton, Pennsylvania 18042, Chapter 87: Blaug, Seymour, commonly known in all pharmaceutical chemistries.

The pharmaceutical composition of the present invention can provide an antioxidative effect, a whitening effect and a wrinkle-reducing effect, which are desirable when it contains an effective amount of a bellflower extract. As used herein, the term " effective amount " refers to an amount that exhibits a further reaction than the negative control, and preferably refers to an amount sufficient to improve muscle function. The pharmaceutical composition of the present invention may contain 0.01 to 99.99% of a bellflower extract, and the balance may be a pharmaceutically acceptable carrier. The effective amount of the bellflower extract contained in the pharmaceutical composition of the present invention will vary depending on the form in which the composition is produced.

The total effective amount of the pharmaceutical composition of the present invention may be administered to a patient in a single dose and may be administered by a fractionated treatment protocol administered over a prolonged period of time in multiple doses. The pharmaceutical composition of the present invention may vary in the content of the active ingredient depending on the degree of the disease. In the case of parenteral administration, it is preferably administered in an amount of 0.01 to 50 mg, more preferably 0.1 to 30 mg per 1 kg of body weight per day on the basis of the bellflower extract, and when administered orally, It may be administered one to several times in an amount of preferably 0.01 to 100 mg, more preferably 0.01 to 10 mg per 1 kg. However, the dose of the bellflower extract is determined based on various factors such as age, weight, health, sex, severity of disease, diet and excretion rate, as well as the administration route and the frequency of treatment of the pharmaceutical composition will be. Thus, in view of this, one of ordinary skill in the art will be able to determine the appropriate effective dosage for the particular use of the platycodon extract of the present invention. The pharmaceutical composition according to the present invention is not particularly limited to the formulation, administration route and administration method as long as the effect of the present invention is exhibited.

The pharmaceutical compositions of the present invention may be used alone or in combination with methods using surgery, radiation therapy, hormone therapy, chemotherapy or biological response modifiers.

The pharmaceutical composition of the present invention can also be provided as a formulation of a topical preparation containing the bellflower extract as an active ingredient.

When the pharmaceutical composition of the present invention is used as an external preparation for skin, it may further contain at least one selected from the group consisting of fatty substances, organic solvents, solubilizers, thickening and gelling agents, softening agents, antioxidants, suspending agents, stabilizers, foaming agents, Such as water, an ionic emulsifier, a nonionic emulsifier, a filler, a sequestering agent, a chelating agent, a preservative, a vitamin, a blocker, a wetting agent, a necessary oil, a dye, a pigment, a hydrophilic activator, Such as any other ingredients conventionally used in the field of skin science. The components can also be introduced in amounts commonly used in the field of dermatology.

When the pharmaceutical composition of the present invention is provided as an external preparation for skin, it may be a formulation such as, but not limited to, an ointment, a patch, a gel, a cream or a spray.

The composition of the present invention may also be a food composition. When the platycodon grandiflorum extract of the present invention is a food composition, it can be used for those having antioxidative, whitening and wrinkle-reducing effects.

The food composition of the present invention includes all forms of functional foods, nutritional supplements, health foods, food additives and feeds, It is targeted for eating. Food compositions of this type may be prepared in a variety of forms according to conventional methods known in the art.

Food compositions of this type may be prepared in a variety of forms according to conventional methods known in the art. Common foods include but are not limited to beverages (including alcoholic beverages), fruits and processed foods (eg, canned fruits, jam, maamalade, etc.), fish, meat and processed foods (Eg butter, chewing), edible vegetable oil, margarine (such as corn oil, etc.), breads and noodles (eg udon, buckwheat noodles, ramen noodles, spaghetti, macaroni, etc.), juice, various drinks, cookies, , Vegetable protein, retort food, frozen food, various kinds of seasoning (for example, soybean paste, soy sauce, sauce, etc.). The nutritional supplement may be prepared by adding a plant extract to capsules, tablets, rings, and the like. In addition, the health functional food is not limited to this. For example, the platycodone extract of the present invention itself may be prepared in the form of tea, juice, and drink and liquefied, granulated, encapsulated and powdered It can be ingested. In order to use the bellflower extract as a food additive, it may be prepared in the form of a powder or a concentrated liquid. In addition, it can be prepared in the form of a composition by mixing together a bellflower extract and a known active ingredient known to have antioxidative, whitening and wrinkle-reducing effects.

When the composition of the present invention is used as a health beverage composition, the health beverage composition may contain various flavors or natural carbohydrates as an additional ingredient such as ordinary beverages. The above-mentioned natural carbohydrates include monosaccharides such as glucose and fructose; Disaccharides such as maltose, sucrose; Polysaccharides such as dextrin, cyclodextrin; Xylitol, sorbitol, erythritol, and the like. Sweeteners include natural sweeteners such as tau Martin and stevia extract; Synthetic sweetening agents such as saccharin and aspartame, and the like can be used. The ratio of the natural carbohydrate is generally about 0.01 to 0.04 g, preferably about 0.02 to 0.03 g per 100 mL of the composition of the present invention.

The bellflower extract may be contained as an active ingredient of a food composition, and the amount thereof is not particularly limited to an amount effective to achieve antioxidative, whitening and wrinkle-reducing action, but is preferably 0.01 to 100% by weight based on the total weight of the composition Do. The food composition of the present invention may be prepared by blending with a bellflower extract together with other active ingredients known to have antioxidative, whitening and wrinkle-reducing effects.

In addition to the above, the health food of the present invention may contain various nutrients, vitamins, electrolytes, flavors, colorants, pectic acid, salts of pectic acid, alginic acid, salts of alginic acid, organic acid, protective colloid thickener, pH adjusting agent, Glycerin, an alcohol or a carbonating agent, and the like. In addition, the health food of the present invention may contain flesh for the production of natural fruit juice, fruit juice drink or vegetable drink. These components may be used independently or in combination. The proportion of such additives is not critical, but is generally selected in the range of 0.01 to 0.1 parts by weight per 100 parts by weight of the composition of the present invention.

Hereinafter, embodiments of the present invention will be described in detail with reference to the accompanying drawings. However, the following examples are for the purpose of specifying the contents of the present invention, and thus the present invention is not limited thereto.

Example 1 Experimental Material and Extraction Method

Platycodon grandiflorum (medicinal product market in Yeongju) was washed, dried and pulverized to an appropriate size. Six grams of commercially available enzyme (AMG 300L, Termamyl 2X, BAN 480L, Fungamyl, Viscozyme and Pectinex Ultra, Novozymes) were added to 1 g of the pulverized platycodone powder, and 20 ml of distilled water was added thereto. Lt; / RTI >

As a result, as shown in Table 1 below, it was confirmed that the extraction yield was highest for Plasmodium falciparum extract of Viscozyme and Termamyl 2X among commercially available carbohydrateases.

Enzyme Extraction Yield (%) Control 38.67 ± 0.87 Amg 300L 41.14 + - 0.54 Termamyl 2X 47.27 + 0.14 BAN 480L 39.68 ± 1.07 Fungamyl 43.25 + - 2.43 Viscozyme 48.81 + - 1.02 Pectinex Ultra 41.14 + 1.15

According to the above results, the ratio of the complex enzyme with the highest saponin content and extraction yield and extraction conditions were established using two enzymes with high extraction yield. (Bellows powder 1 g, distilled water 20 ml, enzyme concentration 5%) were extracted under the same conditions as described above, and the mixture was subjected to extraction And the extraction yield of Platycodon grandiflorum extract was compared with that of the enzyme.

As a result, the extraction yield of Platycodon grandiflorum extract according to the enzyme volume ratio was the highest when the volume ratio of viscose: termaryl 2X was 1: 2, as shown in Table 2 below. And the yield was improved by about 43%.

Mixing ratio (volume ratio) Extraction Yield (%) Viscozyme Termamyl 2X One 0.5 55.14 + - 0.32 One One 50.11 ± 0.18 0.5 One 49.17 ± 0.37 Control 38.54 + - 0.42

≪ Example 2 > Establishment of optimum extraction conditions

Experiments were conducted to establish optimal conditions for bellflower extraction using the complex enzyme of Example 1 above. 20 ml of distilled water was added to 1 g of dried bell pepper powder, and a complex enzyme having a ratio of 1: 2 of biscotamate: termaryl 2X was added at a concentration of 5% to prepare a bellflower extract. For optimal extraction temperature setting, each extract was prepared at a temperature of 30 to 70 캜.

As a result, as shown in FIG. 2, it was confirmed that the extraction yield was the highest at around 50 ° C. and that the optimum temperature could be set in the range of 40 to 60 ° C.

In order to confirm the optimum treatment concentration of the complex enzyme, the extraction yields at the respective concentrations were compared by adjusting the enzyme concentration to 2 to 10% under the same experimental conditions. As a result, as shown in FIG. 3, the yield tends to increase in a concentration-dependent manner at a concentration of less than 5%. Therefore, the optimal treatment concentration of the complex enzyme of the present invention was set at 5%.

As a result of extracting the bloom extract at the optimum complex enzyme concentration and temperature set as described above, it was confirmed that the extraction yield was about 43% higher than that of the hot water extract (no enzyme) as the control group (FIG. 4). The content of crude saponin contained in the extract was measured and compared according to a method known by Shibata et al. (Shibata S, Tanaka T, Ando T, Sado M, Tsushima S, and Ohawa T (1996) Chemical studies on oriental plant drugs XIV.Protopanaxadiol, a genuine sapogenin of ginseng saponins.Chem.Pharm.Bull. 600). As a result, it was confirmed that the crude saponin content was increased by about 24% in the extract according to the extraction method using the complex enzyme of the present invention, compared to the general hot water extract (FIG. 5).

Example 3: Antioxidative activity of complex enzyme extracts by radical scavenging ability

3.1 Determination of DPPH radical scavenging ability of extract

In order to confirm the antioxidative activity of the complex enzyme extract of the present invention, the DPPH radical scavenging activity of ascorbic acid, bellflower hot water extract and dorado complex enzyme extract was measured and compared.

The DPPH assay is always one of the most common experiments to measure pharmacokinetics. In this experiment, 1,1-diphenyl-2-picrylhydrazyl, which is a deep purple color, was reduced by antioxidants and changed to light yellow color. Radical activity of 1,1-diphenyl-2-picrylhydrazyl (DPPH, Sigma-Aldrich, St. Louis, Mo., USA) was measured as follows. 190 μl of DPPH dissolved in ethanol was added to 10 μl of each extract of hot water extract, and incubated at 37 ° C for 30 minutes in a dark room. Then, absorbance (VersaMax microplate reader; Molecular Devices, Sunnyvale, CA, USA) was measured at 515 nm. DPPH radical scavenging activity was calculated by the following equation

DPPH radical scavenging ability (%) = [1 - (Aa / Ab)] 100

Aa: Absorbance of the extract-added sphere, Ab: Absorbance of the non-treated sphere

As a result, as shown in FIG. 6, when the concentration of each extract was the same as 5 mg / ml, the DPPH radical scavenging ability of the complex enzyme extract of the present invention was about 1.7 times higher than that of the hot-water extract.

3.2 Measurement of ABTS radical scavenging ability of extract

As shown in FIG. 7, when ABTS radical scavenging ability of ascorbic acid, bellows hydrothermal extract and platycodon complex enzyme extracts were compared, when the concentration of each extract was the same at 500 μg / ml, And high ABTS radical scavenging ability.

Antioxidant activity was measured by ABTS radicals by ABTS + cation decolourisation assay. The ABTS + · radical cation scavenging activity was determined by the addition of 7.4 mM 2,2'-azino-bis-3-ethylbenzothiazoline-6-sulphonic acid (ABTS, Sigma-Aldrich, St. Louis, Mo., USA) and 2.5 mM potassiom persulfate Dissolved in 1: 1 ratio, and reacted for 15 hours at room temperature in the dark room to form radicals with ABTS +. Absorbance of the ABTS + solution was diluted to 0.70 (± 0.01) with distilled water before reacting with the extract at 732 nm (Versa Max microplate reader; Molecular Devices). 10 μl of the hot-water extract was reacted with 190 μl of ABTS + solution at room temperature for 5 minutes in a dark room, and absorbance was measured at 732 nm. The ABTS radical scavenging activity was calculated by the following equation.

ABTS + radical scavenging ability (%) = [1 - (Aa / Ab)] x 100

Aa: Absorbance of the extract-added sphere, Ab: Absorbance of the non-treated sphere

<Example 4> Comparison of tyrosinase inhibitory activity of the complex enzyme extract

In order to confirm the whitening effect of the Plasmodium complex enzyme extract of the present invention, tyrosinase inhibitory activity was measured.

Tyrosinase inhibitory activity was measured by colorimetric assay of DOPA chrome produced by tyrosinase. To 10 μl of the sample in which the extract had been dissolved, 130 μl of 0.1 M phosphate buffer (pH 6.8) was mixed. Twenty microliters (in 0.1 M PBS, pH 6.8) of 250 U / ml tyrosinase (Sigma-Aldrich, St. Louis, Mo., USA) was added and reacted at room temperature for 10 minutes. To the reaction solution was added a substrate solution in which 5 mM of L-3, 4-dihydroxyphenylalanine (L-DDPA, Sigma-Aldrich, St. Louis, Mo., USA) was dissolved in phosphate buffer (0.1 M PBS, pH 6.8) After incubation for 10 min, absorbance was measured at 490 nm (Versa Max microplate reader; Molecular Devices). Tyrosinase inhibition was calculated by the following equation.

Tyrosinase inhibitory effect (%) = [1 - (Aa / Ab)] x 100

Aa: Absorbance of the extract-added sphere, Ab: Absorbance of the non-treated sphere

As a result, as shown in FIG. 8, the tyrosinase inhibitory activity of the complex enzyme extract of the present invention was improved by about 40% as compared with the hot-water extract. From the above results, it can be seen that the whitening activity of the Plasmodix complex enzyme extract of the present invention is improved as compared with the hot water extract.

<Example 5> Elastase inhibitory activity of the complex enzyme extract

In order to confirm the wrinkle-improving effect of the present Plasmid complex enzyme extract, Elastase inhibitory activity was measured.

Elastase activity inhibition was determined by incubation with porcine pancreas elastase (Sigma-Aldrich, St. Louis, Mo., USA) and N-succinyl- (L-Ala) USA). &Lt; / RTI &gt; To 10 μl of the sample in which the extract was dissolved, 130 μl of a phosphate buffer (0.1 M PBS, pH 6.8) and 40 μl of 2.5 mM N-succinyl- (L-Ala) 3-ρ-nitroanilide Were mixed and reacted at 25 DEG C for 10 minutes. 20 μl of 0.25 U / ml (0.1 M PBS, pH 6.8) elastase enzyme was added to the reaction solution, and the reaction was completed at 25 ° C. for 5 minutes. Absorbance was measured at 405 nm (Versa Max microplate reader; Molecular Devices). Elastase activity inhibition was calculated by the following equation as the absorbance reduction ratio of the sample solution in which the extract was dissolved and the non -

Elastase inhibition (%) = [1 - (Aa / Ab)] x 100

Aa: Absorbance of the extract-added sphere, Ab: Absorbance of the non-treated sphere

As a result, as shown in FIG. 9, it was confirmed that the complex enzyme extract of the present invention showed an inhibition of elastase activity by about 40% as compared with the hot water extract of the control group. From the above results, it can be deduced that the Plasmid complex enzyme extract of the present invention exhibits an improved wrinkle-reducing effect.

The present invention has been described with reference to the preferred embodiments. It will be understood by those skilled in the art that various changes in form and details may be made therein without departing from the spirit and scope of the invention as defined by the appended claims. Therefore, the disclosed embodiments should be considered in an illustrative rather than a restrictive sense. The scope of the present invention is defined by the appended claims rather than by the foregoing description, and all differences within the scope of equivalents thereof should be construed as being included in the present invention.

Claims (6)

Washing, drying and crushing the bellflower;
Treating water and at least two carbohydrases in the pulverized dry dandelion;
Preparing an extract of the enzyme treated bellflower at a temperature of 40 to 60 DEG C for 10 to 15 hours; And
And concentrating the extract under reduced pressure,
The carbohydrase
Viscozyme, including Arabanase, Cellulase, Beta-glucanase, Hemicellulase and Xylanase, and the like.
Is termamyl 2X, which contains an alpha-amylase,
Viscox: Termexil 2X in a volume ratio of 2: 1,
The carbohydrase is contained at a concentration of 5% based on the total weight of the dried bellflower,
Wherein the extract has a whitening effect and a wrinkle-improving effect. delete delete delete A cosmetic composition for whitening and wrinkle improvement comprising the bellflower extract extracted through the extraction method of claim 1. delete

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