KR101855207B1 - Cosmetic composition containing fermentative extract of terminalia ferdinandiana with increased amount of vitamin c fermented by aureobasidium pullulans - Google Patents

Abstract

The present invention relates to a cosmetic composition comprising a kakaduplum black yeast fermented extract. The kakaduplum black yeast fermented extract of the present invention can be effectively used as a cosmetic composition for antioxidant, anti-inflammation, promotion of collagen synthesis, improvement of skin wrinkles, moisturizing, whitening, skin barrier improvement.

Description

Technical Field [0001] The present invention relates to a cosmetic composition containing a kakadu plum extract having increased vitamin C content through black yeast fermentation. [0002] The present invention relates to a cosmetic composition containing a kakadu plum extract,

The present invention relates to a cosmetic composition comprising a kakaduplum extract as an active ingredient, and more particularly, to a cosmetic composition containing a fermented extract having an increased vitamin C content by fermenting kakaduplum extract with a black yeast mycelium.

Skin aging can be caused by active oxygen species activated by ultraviolet rays, stress, disease states, environmental factors, wounds, and age. When such conditions are intensified, the antioxidant defense network in the living body is destroyed, and cells and tissues are damaged, thereby promoting skin aging. Specifically, lipids, proteins, polysaccharides and nucleic acids, which are major constituents of the skin, are oxidized to destroy skin cells and tissues, resulting in skin aging. The oxidation of protein breaks down collagen, hyaluronic acid, elastin, proteoglycan, and fibronectin, which are skin connective tissues, causing severe inflammation reaction and affect skin elasticity. When it gets worse, mutation of DNA leads to mutation, cancer, and immune function.

Therefore, it is necessary to protect the cell membrane by eradicating reactive oxygen species that are mediated by the body's metabolic process, ultraviolet radiation, inflammatory reaction, and regenerate the already damaged cells by active metabolism to proliferate the cells. It can restore and maintain healthy skin.

On the other hand, the skin is an organ that functions as a protective film for protecting the living body from the external environment, and serves to prevent loss of biocomponents in the human body, that is, water and electrolytes, and to prevent harmful substances from entering the human body from the outside. The skin is divided into the skin layer, the dermal layer and the subcutaneous fat layer. The epidermal layer consists of keratinocytes and melanocytes. Since the keratinocytes in the outermost layer of the skin are in direct contact with the external environment, they are required to have strong resistance against physical, chemical, or material permeation and prevent moisture from being lost to the outside At the same time as zooming, it should maintain its flexibility by holding proper water on its own.

Generally, the moisture content of the skin is about 70% in the dermal layer, but decreases to the skin layer, which is about 10 to 30% in the keratin layer. The water supplied from the dermis layer mainly migrates to the upper part of the stratum corneum by passive diffusion and eventually to the outside. This is called Trans Epidermal Water Loss (TEWL). It is known that the lipid component of the keratinocyte layer and epidermal lipid, which maintains the transdermal water loss at an appropriate level, is the lipid component.

It is known that the keratinocyte layer has a hydrophilic water retention ability called "Natural Moisturizing Factor (NMF)" and plays an important role in moisturizing the skin. Normal keratinocyte layer maintains skin smoothness and softness and maintains normal body protection function when moisture is maintained at about 10 to 30%. However, when the moisture content of the keratinocyte layer is less than 10%, the skin becomes rough and the protective function of the body is lost and the aging phenomenon occurs. For example, in the case of dry skin, the keratinocyte aggregation is weakened, and a scaling phenomenon appears in which the keratinocytes are peeled off like scales from the skin surface. The reason why the skin is dried in this way is that the moisture content of keratinocyte layer is smaller than that of normal skin. In addition to this, even healthy skin may be in a harsh environment, such as wind, cold weather, sunshine, cleansing, shaving, etc., which causes water shortage, which can lead to poor skin condition.

Therefore, it is very important to properly maintain the moisture content of the keratinocyte layer. To this end, cosmetics were added with ingredients similar to sebum, moisturizers such as NMF or polyol, and the like. For example, glycerin or sorbitol having three or more hydroxyl groups as a water-soluble polyol exhibits excellent moisture resistance, but is highly sticky and makes it feel uncomfortable when used. Propylene glycol 1,3-butylene glycol having two hydroxyl groups, . In addition, other natural moisturizing factors such as sodium pyrrolidonecarboxylate (PCA-Na), sodium lactate, urea, etc., are highly electrolytic, which deteriorates emulsion stability of the cosmetic. Amino acids, collagen, elastin, etc. are also capable of moisturizing, but also had a limited ability to moisturize.

The ability of the horny layer to retain moisture can be controlled by natural moisturizing factor (NMF), which consists of sebum components, amino acids, lactic acid, urea, and inorganic salts. And the development of a substance with high moisturizing effect is one of the important research subjects in cosmetics.

Meanwhile, in recent years, many cosmetic products using natural products have been developed to reduce skin irritation caused by various chemical substances and the like. In addition to low adverse effects on skin, natural materials have recently become increasingly appreciated as a cosmetic raw material as consumers' response to cosmetics using natural materials has increased.

The present inventors have studied the applicability of various natural products to cosmetics. As a result, they have found that the extract of kakaduplum fermented with the mycelia of black yeast produced antioxidative, anti-inflammatory, promoting collagen synthesis, Wrinkle improvement, moisturizing, whitening, skin barrier improvement, and completed the present invention.

US Patent Publication No. 2016/0008270 relates to a cosmetic composition having a whitening effect or the like including Terminalia ferdinandiana fruit extract and the like.

The present inventors intend to provide a natural extract which is applicable to skin cosmetics and which is harmless to human body and is highly safe, and a cosmetic composition containing it as an active ingredient.

The present invention also provides a cosmetic composition having a natural extract obtained by fermenting Kakadu plum extract with black yeast mycelium and having an effect as an active ingredient and antioxidant, anti-inflammation, promotion of collagen synthesis, skin wrinkle improvement, moisturizing and skin barrier improvement The purpose.

In order to achieve the above object, the present invention provides a cosmetic composition containing a kakaduplum black yeast fermentation extract.

Preferably, the cosmetic composition may be used for antioxidation, anti-inflammation, skin wrinkle improvement, skin whitening, skin moisturizing or skin barrier improvement.

Preferably, the kakaduplum black yeast fermentation extract can be obtained by fermenting kakadu plum extract with black yeast for 1 to 7 days while maintaining a pH of 5 to 7 under a temperature condition of 20 to 40 ° C.

In addition, the cosmetic composition may contain 0.0001 to 30.0% by weight of kakaduplum black yeast fermentation extract based on the total weight of the cosmetic composition.

Preferably the cosmetic composition is in the form of a solution, suspension, emulsion, paste, gel, cream, lotion, powder, soap, surfactant-containing cleansing, oil, powder foundation, emulsion foundation, wax foundation and spray And can have any one of the formulations selected.

INDUSTRIAL APPLICABILITY The present invention can provide a natural extract which is applicable to skin cosmetics and which is harmless to the human body and has excellent safety, and a cosmetic composition containing it as an active ingredient.

The present invention also provides a cosmetic composition having a natural extract obtained by fermenting Kakadu plum extract with black yeast mycelium and an effective ingredient thereof and having antioxidant, anti-inflammatory, promoting collagen synthesis, improving skin wrinkles, moisturizing, whitening, and skin barrier can do.

Also, it is possible to provide a cosmetic composition in which the content of vitamin C is increased by fermenting the kakaduplum extract of the present invention with black yeast.

Fig. 1 shows HPLC graphs of the kakadu plum extract before (a) and after (b) fermentation of black yeast.

The present inventors have conducted studies to prepare extracts from various natural materials as a raw material by various extraction methods and to select substances having excellent effects such as antioxidation, anti-inflammation, promotion of collagen synthesis, improvement of skin wrinkles, moisturizing and skin barrier improvement . As a result, it was found that the extract prepared by fermenting the black yeast mycelium into the extract of kakaduplum is most suitable for the purpose of the present invention described above.

Kakadu plum (Terminalia Ferdinandiana ) is the most abundant vitamin C in the world's existing plants and has recently become an antioxidant. In addition, Kakaduplum has a vitamin C content of 3200 mg to 5000 mg per 100 g, which is comparable to that of orange 50 mg / 100 g. Vitamin C promotes collagen synthesis of proline and amino acids. It also improves skin elasticity, reducing wrinkles and protecting skin from active oxygen and ultraviolet rays. It is resistant to pests, has excellent self-sufficiency, can be cultivated without pesticides, and fruits, leaves, roots and trees suitable for the well-being era can be used as raw materials for food tea, medicines or cosmetics.

Aureobasidium pullulans ) is a microorganism used to make beer, wine, etc. It is a collective of single celled organisms that are fungi or mushroom flocks but do not have hyphae and no photosynthesis or motility. It is a microorganism that humans have used for food since 5000 years ago, and yeast itself is used as a cheap fat or protein source. It is rich in vitamin B group and contains vitamin D. In addition, black yeast is a microorganism that produces beta-glucan, and it is known that the black yeast fermented product alone exhibits a moisturizing effect, an antioxidant, and soothing effect. Black yeast beta-glucan contains a large amount of components known to have physiological activities such as anticancer and insulin secretion inhibition.

Until now, extracts of kakaduplum have been widely used for other purposes. However, as in the present invention, the effect of the fermentation form, particularly the black yeast fermentation extract of kakaduplum, is not specifically known.

The cosmetic compositions of the present invention Kakadu plum fermentation extract the black two plum extract Kaka yeast (Aureobasidium pullulans ), and exhibits excellent antioxidant, anti-inflammation, promotion of collagen synthesis, improvement of skin wrinkles, moisturizing, whitening, skin barrier improvement.

In the present invention, kakadooplum extract can be produced according to a conventional method known in the art. Specifically, seeds, leaves or stems of kakaduplum can be dried and pulverized, and then, using conventional solvents, under the conditions of ordinary temperature and pressure. Specifically, the kakadu plum extract is extracted using a solvent selected from the group consisting of water, an anhydrous or a lower alcohol having 1 to 4 carbon atoms, acetone, ethyl acetate, butyl acetate and 1,3-butylene glycol, The seeds, leaves or stems of the two plums can be extracted using the above-mentioned solvent, followed by mixing them.

The solvent is preferably an alcohol, more preferably ethanol. At this time, the ethanol is preferably 70% ethanol. The suitable amount of the extraction solvent is 1 to 15 times, more preferably 5 to 10 times, and most preferably 7 times the weight of the dried kakadu plume.

On the other hand, the kakaduplum extract of the present invention includes not only the extract obtained by the solvent extraction method described above, but also the extract obtained through a conventional purification process. For example, by separation using an ultrafiltration membrane having a constant molecular weight cut-off value, separation by various chromatographies (made for separation by size, charge, hydrophobicity or affinity), and the like, Fraction is also considered to be included in the kakadooplum extract of the present invention.

In addition, the kakaduplum extract of the present invention may be prepared in powder form by an additional process such as vacuum distillation and freeze-drying or spray drying.

The kakaduplum black yeast fermentation extract can be prepared by fermenting the above-described kakaduplum extract with black yeast. Specifically, the kakaduplum extract can be prepared by the following method. First, the saccharide can be sterilized by adding 1 to 5% by weight of glucose as a carbon source to kakaduplum extract, and adding 0.1 to 1%, preferably 0.2 to 0.5% by weight, of peptone. It can be cultured by inoculating a culture medium of Aureobasidium pullulans strain so as to have a concentration of 25 g / L. The culture can be carried out using a 5 L fermenter for 1 to 7 days at 20 to 40 ° C while maintaining the pH at 5-7. After culturing, the culture broth was centrifuged and the culture was firstly removed and sterilized (121 ° C, 15 minutes, 1.5 atm) to prevent further culture.

After the fermentation, the kakaduprum fermented product, from which the cultures were firstly removed, was refluxed with ethanol to make a final 70% (v / v) ethanol solution, cooled, and filtered. When the filtration is completed, the extract obtained in the extraction step is transferred to a concentration tank and concentrated under reduced pressure or freeze-dried at 50 ° C or lower to prepare the kakaduplum black yeast fermentation extract of the present invention.

According to a preferred embodiment of the present invention, the kakaduplum black yeast fermented extract added to the cosmetic composition may be contained in an amount of 0.0001 to 30.0% by weight based on the total weight of the cosmetic composition. When the content of the kakaduplum black yeast fermentation extract is less than 0.0001% by weight, the effect of antioxidation, anti-inflammation, promotion of collagen synthesis, improvement of skin wrinkles, moisturizing, whitening and skin barrier improvement is insignificant.

Meanwhile, the cosmetic composition of the present invention may contain, as an active ingredient, components commonly used in cosmetic compositions in addition to the kakaduplum black yeast fermentation extract described above. Examples thereof include antioxidants, stabilizers, solubilizers, vitamins, Such as conventional adjuvants, and carriers.

Meanwhile, the cosmetic composition of the present invention may be prepared in any form conventionally produced in the art, and examples thereof include a solution, a suspension, an emulsion, a paste, a gel, a cream, a lotion, a powder, a soap, Containing cleansing, oils, powder foundations, emulsion foundations, wax foundations and sprays. However, the present invention is not limited thereto. More specifically, it can be manufactured in the form of a soft lotion, a convergent lotion, a nutritional lotion, a nutritional cream, a massage cream, an essence, an eye cream, a cleansing cream, a cleansing foam, a cleansing water, a pack, a spray or a powder.

When the formulation of the cosmetic composition of the present invention is a paste, a cream or a gel, an animal oil, vegetable oil, wax, paraffin, starch, tracant, cellulose derivative, polyethylene glycol, silicone, bentonite, silica, talc or zinc oxide Can be used.

When the formulation of the cosmetic composition of the present invention is a powder or a spray, lactose, talc, silica, aluminum hydroxide, calcium silicate or polyamide powder may be used as a carrier component, Propane / butane, or dimethyl ether. ≪ RTI ID = 0.0 >

When the formulation of the cosmetic composition of the present invention is a solution or an emulsion, a solvent, a dissolving agent or an emulsifying agent is used as a carrier component, and examples thereof include water, ethanol, isopropanol, ethyl carbonate, ethyl acetate, benzyl alcohol, benzyl benzoate , Propylene glycol, 1,3-butyl glycol oil, glycerol aliphatic ester, polyethylene glycol or sorbitan fatty acid esters.

When the formulation of the cosmetic composition of the present invention is in the form of a suspension, the carrier component may include water, a liquid diluent such as ethanol or propylene glycol, ethoxylated isostearyl alcohol, polyoxyethylene sorbitol ester and polyoxyethylene sorbitan ester Suspending agent, microcrystalline cellulose, aluminum metahydroxide, bentonite, agar or tracant, etc. may be used.

When the formulation of the cosmetic composition of the present invention is a cleansing agent containing an interfacial active agent, the carrier component may include aliphatic alcohol sulfate, aliphatic alcohol ether sulfate, sulfosuccinic acid monoester, isethionate, imidazolinium derivative, methyltaurate, Fatty acid amide ether sulfate, alkylamido betaine, aliphatic alcohol, fatty acid glyceride, fatty acid diethanolamide, vegetable oil, lanolin derivative, or ethoxylated glycerol fatty acid ester.

On the other hand, the kakaduplum black yeast fermentation extract of the present invention exhibits an antioxidant effect (Experimental Example 2), anti-inflammatory effect (Experimental Example 4), collagen synthesis effect (Experimental Example 5) and skin whitening effect (Experimental Example 6). In addition, the cosmetic composition prepared by containing the kakaduplum black yeast fermented extract as an active ingredient having such an effect showed excellent skin moisturizing effect (Experimental Example 7) and skin barrier improving effect (Experimental Example 8).

Hereinafter, the present invention will be described in more detail with reference to the following Production Examples. However, the following Production Examples are intended to further illustrate the present invention, and the scope of the present invention is not limited to the following Production Examples.

Comparative Manufacturing Example 1: Kakadu Plum  Preparation of extract

The outposts of Kakadu plum were washed with purified water, dried, and then pulverized and finely ground using a sieve of 100 mesh. 70% ethanol was added to 100 g / L of Kakadu plum pulverized product, and the mixture was refluxed three times for 5 hours. After cooling, the mixture was filtered through Whatman # 3 filter paper, concentrated under reduced pressure at 50 캜 or less and freeze- To prepare kakadu plum extract.

Manufacturing example  1: Kakadu Plum Black yeast  Preparation of fermented extract

To 5 g of the Kakadooplum extract prepared in Comparative Preparation Example 1, 2% of glucose was added as a carbon source, and 0.5% of peptone was further added to sterilize. Then, the culture was inoculated with a culture solution of Aureobasidium pullulans strain so as to have a concentration of 25 g / L. Cultures were cultured in a 5 L fermenter for 7 days at 37 ° C and pH 5-7. After culturing, the culture broth was centrifuged and the culture was firstly removed and sterilized (121 ° C, 15 minutes, 1.5 atm) to prevent further culture.

After the fermentation, the kakaduplum fermented product, from which the cultures were firstly removed, was refluxed three times for 5 hours by adding ethanol to make a final 70% (v / v) ethanol aqueous solution. After cooling, Lt; / RTI > When the filtration is completed, the extract obtained in the extraction step is transferred to a concentration tank and concentrated under reduced pressure or freeze-dried at 50 ° C or lower.

Comparative Manufacturing Example  2 : Black yeast  Preparation of fermentation broth

It was added a 2% Four clique as a carbon source, and sterilized by adding an additional 0.5% peptone. Then, the culture was inoculated with a culture solution of Aureobasidium pullulans strain so as to have a concentration of 25 g / L. Cultures were cultured in a 5 L fermenter for 7 days at 37 ° C and pH 5-7. After culturing, the culture broth was centrifuged and the culture was firstly removed and sterilized (121 ° C, 15 minutes, 1.5 atm) to prevent further culture.

After the fermentation, the cultures were firstly removed, ethanol was added to make a final 70% (v / v) ethanol aqueous solution, refluxed three times for 5 hours, cooled, and filtered through Whatman # 3 filter paper. When the filtration was completed, the filtrate was transferred to a concentration tank and concentrated under reduced pressure or freeze-dried at 50 ° C or lower to prepare a black yeast fermentation broth free from kakaduplum extract.

Comparative Manufacturing Example  3 to 6: Preparation of Kakadu Plum Fermentation Extract Using Various Bacteria

A kakaduplum fermentation extract was prepared in the same manner as in Production Example 1, except that the mycelium of Table 1 below was used instead of black yeast.

mycelium Comparative Production Example 3 Saccharomyces cerevisiae Comparative Production Example 4 Lactobacillus casei Comparative Preparation Example 5 Asperillus oryzae Comparative Preparation Example 6 Bacillus subtilis

Experimental Example  1: Vitamin C analysis

The vitamin C content in the kakaduplum extract of Comparative Preparation Example 1 and the kakaduplum black yeast fermentation extract of Preparation Example 1 was analyzed by HPLC (mobile phase 25 mM potassium phosphate (KH2PO4), pH 2.5). The analytical column was analyzed with OptiPack C18, column temperature 30 ° C and ultraviolet detector absorbance wavelength 254 nm.

The results of analysis of the vitamin C content of the kakadu plum extract before (a) and after (b) of black yeast fermentation are shown in Fig.

The vitamin C contents of the kakaduplum black yeast fermentation extract of Preparation Example 2 and the kakaduprum fermentation extracts of Production Examples 3 to 7 were measured and shown in Table 2.

The content of vitamin C before and after fermentation Before fermentation (ppm) After fermentation (ppm) Production Example 1 (Black yeast fermentation) 2,913.4 19,452.5 Comparative Preparation Example 3 (yeast fermentation) 2,913.4 2,915.43 Comparative Preparation Example 4 (lactic acid fermentation) 2,913.4 2,913.21 Comparative Preparation Example 5 (fermentation of M. tuberculosis) 2,913.4 2,915.41 Comparative Preparation Example 6 (Bacillus subtilis fermentation) 2,913.4 2,913.25

The content of vitamin C before and after the black yeast fermentation of Kakadu plum extract was 2913.4 ppm before fermentation and 18,292.5 ppm after fermentation (Production Example 1). However, in the case of other mycelial fermented products, no significant increase in vitamin C content was observed.

Experimental Example  2: Kakadu Plum  Extract and Kakadu Plum Black yeast  Fermented extract DPPH method  Evaluation of antioxidant activity

Free radical scavenging activity was evaluated according to the concentrations (31.5, 62.5, 125, 250, 500, 1000 占 퐂 / ml) of Preparation Example 1 and Comparative Preparation Examples 1 to 6.

The DPPH assay measures absorbance at 540 nm to the extent that the inhibitor decolorizes the stable radical DPPH (2,2-Diphenyl-1-picrylhydrazyl radical). The used samples are shown in Table 3 below, and the free radical scavenging activity was measured using the following equation (1). The control group was Epigallocatechin gallate (EGCG).

material Control group Blank of C Exp. Group Blank of E A DPPH (MeOH) 180 (180) 180 (180) B Sample (solvent) (20) (20) 20 20

A: Abs of experimental group

B: Abs blank of experimental group

C: Abs of control group

D: Abs of blank of control group

Free radical scavenging activity Production Example 1 Comparative Preparation Example 1 Comparative Production Example 2 Comparative Production Example 3 Comparative Production Example 4 Comparative Preparation Example 5 Comparative Preparation Example 6 EGCG 25uM - - - - - - - 55.34 0.001 wt% 43.15 13.25 5.95 24.15 16.57 14.16 23.19 - 0.002 wt% 67.24 23.17 6.10 32.10 23.20 25.14 27.45 - 0.005 wt% 82.41 24.57 7.14 39.45 33.45 35.25 37.66 - 0.01 wt% 97.58 36.26 9.09 57.20 45.30 46.47 47.27 -

As shown in Table 4, the kakaduplum fermentation extract (Preparation Example 1 and Comparative Preparation Examples 3 to 6) was superior to the kakaduplum extract (Preparation Example 1) in free radical scavenging activity. Among fermented extracts, black yeast fermented extract (Preparation Example 1) showed the highest free radical scavenging activity. In addition, superior antioxidative effect was also confirmed in comparison with Comparative Production Example 2 which contained only black yeast fermentation broth. Therefore, it was confirmed that the kakaduplum black yeast fermentation extract of the present invention has the highest antioxidative effect.

Experimental Example  3: Evaluation of cell survival rate of kakadu plum extract and kakaduplum fermented extract

The toxicity of Preparation Example 1 and Comparative Preparation Examples 1 to 6 on cells was measured. Cytotoxicity was performed by modifying Mosmann's method of measuring cell viability using 3- (4,5-dimethylthiazol-2-yl) -2-5-diphenyltetrazolium bromide (MTT) reagent.

HDF was dispensed into 96-well plates at a concentration of 1 x 10 ⁴ cells / well and cultured at 37 ° C and 5% CO ₂ for 24 hours. The culture medium was removed and the sample was cultured for 24 hours in a concentration-treated medium. The medium was removed and washed twice with PBS (phosphate buffered saline). MTT was dissolved in PBS at 5 mg / mL, and 50 μL was added and cultured at 37 ° C and 5% CO ₂ for 2 hours. DMSO was added to each well (100 μL), stirred for 10 minutes, and then absorbance was measured at 540 nm.

The measurement results are shown in Table 5 below. As shown in Table 5, no cytotoxicity was observed at all concentrations of each sample. From this, it was confirmed that the kakaduplum black yeast fermented extract of the present invention is harmless to human body and has excellent safety.

Cell survival rate (%) Name of sample Treatment concentration (PPM) Cell survival rate (%) Production Example 1 50 104.5 100 103.1 Comparative Preparation Example 1 50 102.3 100 104.7 Comparative Production Example 2 50 98.7 100 101.9 Comparative Production Example 3 50 97.9 100 101.5 Comparative Production Example 4 50 99.5 100 102.5 Comparative Preparation Example 5 50 98.5 100 98.7 Comparative Preparation Example 6 50 97.1 100 101.9

Experimental Example  4: Evaluation of anti-inflammatory properties of kakaduplum extract and kakaduprum fermented extract (5- Lipoxygenase  Inhibition activity evaluation)

The 5-Lipoxygenase inhibitory activity of Preparation Example 1 and Comparative Preparation Examples 1 to 6 was evaluated to evaluate the anti-inflammatory effect.

Lipoxygenase produces a variety of substances from arachidonic acid that act as various chemical mediators involved in inflammatory and allergic reactions in vivo. Therefore, a substance that inhibits lipoxygenase may result in inhibiting the production of various chemical mediators, so it can be assumed that it is effective for allergy. The activity level of lipoxygenase is an experiment to measure the degree of peroxide formation using substrate and enzyme.

The used samples were tested as shown in Table 6 below, and the 5-lipoxygenase inhibitory activity was measured using the equation (2). The control group was nordihydroguaiaretic acid (NDGA).

material Control group Blank of C Exp. Group Blank of E A Buffer solution 2ml 2ml 2ml 2ml B Sample (solvent) (20 [mu] l) (20 [mu] l) 20ul 20ul C Enzyme 40ul - 40ul - D Substrate 70ul 70ul 70ul 70ul

Production Example 1 Comparative Preparation Example 1 Comparative Production Example 2 Comparative Production Example 3 Comparative Production Example 4 Comparative Preparation Example 5 Comparative Preparation Example 6 NDGA 250 nM - - - - - - - 52.55 0.001 wt% 34.20 20.25 3.27 26.45 22.57 22.42 36.07 - 0.002 wt% 42.94 27.50 5.08 33.06 36.15 35.14 32.78 - 0.005 wt% 63.40 34.98 5.48 34.23 35.99 38.77 32.93 - 0.01 wt% 75.73 33.90 5.39 49.79 42.96 43.78 42.95 -

As shown in the results of Table 7, the kakaduprum fermentation extract (Preparation Example 1 and Comparative Preparation Examples 3 to 6) was superior to kakadu plum extract (Comparative Preparation Example 1) and black yeast fermentation broth (Comparative Preparation Example 2) 5-lipoxygenase inhibition was excellent. Among the fermented extracts, the kakaduplum black yeast fermented extract (Preparation Example 1) showed the highest 5-lipoxygenase inhibitory ability. Generally, the higher the inhibitory effect of 5-lipoxygenase, the more anti-inflammatory effect. Therefore, the above results confirm that the kakaduplum black yeast fermented extract has a higher anti-inflammatory effect than the ordinary kakadooplum extract.

Experimental Example  5: Evaluation of collagen synthesis amount of kakaduplum extract and kakaduprum fermented extract

The increase rate of collagen biosynthesis in Production Example 1 and Comparative Production Examples 1 to 6 was evaluated.

HDF was added to a 24-well plate at a concentration of 5 x 10 ⁴ cells / well and cultured at 37 ° C and 5% CO ₂ for 24 hours. Serum-free DMEM medium supplemented with Preparation Example 1 and Comparative Preparations 1 to 6 and a control group were further cultured in serum-free DMEM medium containing no kakaduplum fermentation extract or kakadooplum extract for 24 hours. After the incubation, the supernatant of each well was collected and the amount of procollagen type I C-peptide (PICP) was measured using a collagen kit (Takara, Japan) and converted into ng / ml, Respectively. Collagen biosynthesis increase rate (%) was calculated according to the following formula (3), and the control group was L-ascorbic acid (L-AA).

Production Example 1 Comparative Preparation Example 1 Comparative Production Example 2 Comparative Production Example 3 Comparative Production Example 4 Comparative Preparation Example 5 Comparative Preparation Example 6 L-AA 0.3mM - - - - - - - 133.41 0.001 wt% 127.55 102.09 102.43 106.14 115.35 107.76 114.74 - 0.002 wt% 164.78 107.30 103.76 116.22 116.46 115.57 127.47 - 0.005 wt% 175.64 116.77 104.88 121.67 123.72 125.34 127.51 - 0.01 wt% 183.47 117.32 104.97 128.22 135.24 126.72 132.32 -

As can be seen from the results of Table 8, the kakaduplum black yeast fermented extract (Preparation Example 1) of the present invention contains the kakaduplum extract (Comparative Preparation Example 1), the black yeast fermentation broth (Comparative Preparation Example 2) The amount of collagen synthesis was superior to that of fermented Kakaduprum fermented extract (Comparative Preparation Examples 3 to 6) fermented as a bacterium.

Experimental Example  6: B16F1  Measurement of inhibition of melanin synthesis using melanocytes

In Experimental Example 6, arbutin known as a whitening agent was used as a comparative sample and B16F1 melanocyte was used to confirm the melanin synthesis inhibitory effect of the samples obtained in Production Examples 1 to 6 and Comparative Production Example 1.

The B16F1 melanocyte used in Experimental Example 6 is a cell line derived from a mouse and is a cell that secretes a melanin pigment called melanin. The inhibitory effect of B16F1 melanin on melanin synthesis was measured as follows.

B16F1 melanocytes were plated at a concentration of 2 x 10 ⁶ per well in a 6-well plate, and after attaching the cells, the samples were treated at a concentration not causing toxicity and cultured for 72 hours. After culturing for 72 hours, cells were detached with trypsin-EDTA, the number of cells was measured, and the cells were recovered by centrifugation. Quantification of intracellular melanin was carried out with a slight modification of the method of Rotan (R Lotan and D Lotan, Cancer Res, 40: 3345-3350, 1980). Cell pellets were washed once with PBS and then 1 ml of homogenization buffer (50 mM sodium phosphate, pH 6.8, 1% Triton X-100, 2 mM PMSF) was added and vortexed for 5 minutes to disrupt the cells. After centrifugation (3,000 rpm, 10 min), 1N NaOH (10% DMSO) was added to the cell filtrate to dissolve the extracted melanin, and the absorbance of melanin was measured at 405 nm with a microplate reader. Melanin was quantified The percent inhibition of melanin formation in the sample was measured. The inhibition rate (%) of melanin formation of B16F1 melanocytes was calculated by the following equation (4).

A: Amount of melanin in wells to which no sample was added

B: Amount of melanin in the well to which the sample was added

Production Example 1 Comparative Preparation Example 1 Comparative Preparation Example 1 Comparative Production Example 3 Comparative Production Example 4 Comparative Preparation Example 5 Comparative Preparation Example 6 Arbutin 0.2 wt% - - - - - - - 52.33 0.001 wt% 47.35 6.07 3.29 7.16 5.36 5.96 14.64 - 0.002 wt% 63.58 11.18 11.87 5.35 15.37 15.76 26.77 - 0.005 wt% 75.24 16.67 20.75 16.54 21.32 23.35 29.35 - 0.01 wt% 83.65 25.27 23.47 18.33 22.36 25.36 33.75 -

The inhibitory effect of melanin on B16F1 melanocytes was measured and the results are shown in Table 9. < tb > < TABLE > The kakaduplum black yeast fermentation extract (Preparation Example 1) of the present invention was prepared by mixing the kakaduplum extract (Comparative Preparation Example 1), the black yeast fermentation broth (Comparative Preparation Example 2) and the kakaduplum fermentation extract fermented with various bacteria (Examples 3 to 6). From the above results, it was confirmed that the melanin synthesis inhibition rate of the kakaduplum black yeast fermentation extract was excellent, and the excellent whitening effect of the composition for external application for skin of the present invention was confirmed based on this.

Prescription  Manufacturing: Kakadu Plum Black yeast  Fermented extract Cosmetic  Produce

A cosmetic composition containing 20.0% by weight of the kakaduplum black yeast fermentation extract of Preparation Example 1 was prepared according to the composition shown in Table 10 below, referred to as Prescription Example 1, and a cosmetic composition containing Kakadu Plum Extract (Comparative Preparation Example 1) The composition was compared to Comparative Example 1, and a cosmetic composition containing a black yeast fermentation broth (Comparative Preparation Example 2) was prepared and referred to as Comparative Prescription Example 2.

ingredient Prescription Example 1
(weight%) Comparative Prescription Example 1
(weight%) Comparative Prescription Example 2
(weight%) Production Example 1 20.0 - - Comparative Preparation Example 1 - 20.0 - Comparative Production Example 2 - - 20.0 1,3-BG 10.0 10.0 10.0 glycerin 5.1 5.1 5.1 Propylene glycol 4.2 4.2 4.2 Tocopheryl acetate 3.0 3.0 3.0 Liquid paraffin 4.6 4.6 4.6 Triethanolamine 1.0 1.0 1.0 Squalane 3.1 3.1 3.1 Macadamia nut oil 2.5 2.5 2.5 Polyahyuvait 60 1.6 1.6 1.6 Azithromycin 1.6 1.6 1.6 Propylparaben 0.6 0.6 0.6 Carboxyl vinyl polymer 1.5 1.5 1.5 incense a very small amount a very small amount a very small amount antiseptic a very small amount a very small amount a very small amount Purified water Balance Balance Balance system 100 100 100

Experimental Example  7: Kakadu Plum Black yeast  Skin of fermented extract Moisture power  Improvement effect

In order to examine the effect of the cosmetic composition containing the fermented extract of kakaduplum black yeast fermentation extract of the present invention on the skin moisturizing ability, the following experiment was conducted. Twenty patients in the 20s to 40s without skin diseases were divided into two groups each consisting of 20 patients per group. The nutritional creams of the cream of Prescription Example 1 and Comparative Prescription Example 1 prepared in Table 10 were divided into two groups, And applied to the face and forehead for 1 month. The skin conductivity was measured by using a corneometer (CM820 courage Khazaka electronic GmbH, Germany) under the constant temperature and humidity conditions (24 ° C, 40% humidity) before the start of application, and the skin conductivity was measured at 1 week, 2 weeks, The rate of increase in skin conductivity (%) was measured and the rate of increase was evaluated. The results are shown in Table 11 below.

sample After 1 week after 2 weeks After 4 weeks Prescription Example 1 54 59 65 Comparative Prescription Example 1 29 35 40 Comparative Prescription Example 2 17 18 24

As shown in the results of Table 11, in the case of Formulation Example 1, which is a cosmetic containing the fermented extract of kakaduplum black yeast of the present invention, the rate of skin conductivity increase was much higher than that of Comparative Formulation Examples 1 and 2. Since the skin conductivity is generally proportional to the skin moisture content, the above results confirm that the cosmetic containing the germinated black yeast fermented extract keeps the skin moisture content higher than that of the cosmetic containing no germinated black yeast fermented extract.

Experimental Example  8: Kakadu Plum Black yeast  Effect of fermented extract on skin barrier

In order to examine the skin barrier improvement effect of the cosmetic composition containing the kakaduplum black yeast fermented extract of the present invention, the following experiment was conducted. In the present experimental example, the cosmetic materials of Table 10 were evaluated by conducting a comparative experiment using a Tewameterr (TM300, Courage and Khazaka Electronic Co., Germany) for transepidermal water loss (TEWL) in humans. Each sample was applied to the right and left faces of 40 women in their 20s and 40s. Percutaneous water loss before, 1 hour, 2 hours, 4 hours, and 6 hours after applying 1cm downward to the right side of 3cm from the eyes The results are shown in Table 1. The transdermal water loss was measured using a Tewameterr (TM300, Courage and Khazaka Electronic Co., Germany) three times and the average value was evaluated.

division 1 hours 2 hours 4 hours 6 hours Prescription Example 1 12.9 13.8 13.3 9.7 Comparative Prescription Example 1 17.8 18.0 19.2 12.5 Comparative Prescription Example 2 17.9 18.6 19.9 15.6

As shown in Table 12, the amount of transdermal water loss was measured. As a result, the amount of transdermal water loss was reduced in the facial skin of an experimenter who applied cream containing Kakaduplum black yeast fermented extract, .

While the present invention has been particularly shown and described with reference to exemplary embodiments thereof, it is to be understood that the same is by way of illustration and example only and is not to be construed as limiting the scope of the present invention. It is therefore intended that the scope of the invention be defined by the claims appended hereto and their equivalents.

Claims (10)

A terminalia ferdinandiana extract is treated with Aureobasidium pullulans at a temperature of 20 to 40 ° C and maintained at a pH of 5 to 7 for 1 to 7 days to obtain a cosmetic composition containing a fermented Kakaduplum black yeast extract As a composition,
Wherein the cosmetic composition has at least one effect selected from the group consisting of an antioxidative effect, an anti-inflammatory effect, a skin wrinkle improving effect, a skin whitening effect, a skin moisturizing effect and a skin barrier improving effect. The method according to claim 1,
Wherein the kakaduplum black yeast fermented extract is added in an amount of 0.0001 to 30.0% by weight based on the total weight of the cosmetic composition. The method according to claim 1,
The cosmetic composition may be in the form of a solution, suspension, emulsion, paste, gel, cream, lotion, powder, soap, surfactant-containing cleansing, oil, powder foundation, emulsion foundation, wax foundation and spray Wherein the cosmetic composition has one formulation. delete delete delete delete delete delete delete

Images