KR101103283B1 - The Manufacturing Method of Fermented Aloe for Whitening Effect in skin, and the Functional Whitening Cosmetics Containing Fermented Aloe - Google Patents

Abstract

The present invention relates to aloe fermented product for a whitening cosmetic material of a functional material obtained by inoculating mushroom mycelium on a natural medium based on aloe and a whitening cosmetic using the same.

The aloe fermented product for whitening cosmetic material of the present invention is a substrate containing aloe as a main ingredient, and inoculated with the mushroom mycelium 5-15% (w / w) in the aloe natural medium, and the mushroom mycelium is The inoculated aloe natural medium is cultured and fermented for 3 to 10 days at a temperature of 24 ~ 34 ℃, aeration 0.4 ~ 0.8vvm to obtain aloe fermented product for whitening cosmetic material.

Aloe, Fermented, Functional Food, Ganoderma lucidum

Description

Aloe fermented product for whitening cosmetics and whitening cosmetics using the same {The Manufacturing Method of Fermented Aloe for Whitening Effect in skin, and the Functional Whitening Cosmetics Containing Fermented Aloe}

The present invention relates to aloe fermented product for a whitening cosmetic material of a functional material obtained by inoculating mushroom mycelium on a natural medium based on aloe and a whitening cosmetic using the same.

Melanin is a biopolymer of phenols that is widely distributed in nature. It is a complex of black pigment and protein, and melanin synthesis is the browning or animal that occurs when the cut surface of apples, potatoes, and bananas is exposed to the air. It is found on the outer skin, feathers, skin, hair and eyes. The function of melanin increases the viability of microorganisms against ultraviolet radiation, propagation, drying, and extreme temperatures, and in animals, melanin in the skin or head functions as a means of self-protection. In addition, melanin in invertebrates and fungi acts as an immune system against microorganisms and contributes to the quality of coffee, tea and tobacco (see J. Amm. Rev. Phytophathol. 240, 411. 1986). However, pigmentation in the skin, such as human blemishes and freckles, is due to the ideal increase of melanin in the epidermis, and melanin is biosynthesized in melanosomes in pigment cells called melanocytes in the basement layer of the epidermis. The main production process of melanin is oxidizing tyrosine, a kind of amino acid, to dopa and dopaquinone by tyrosinase, and they are metabolized into dopachrome, indole carboxylic acid and indole quinones by the action of enzyme and automatic oxidation reaction. Synthesize melanin (see J. Mutation research 571, 121. 2005).

By inhibiting tyrosinase, an important enzyme in the biosynthetic action of melanin, it is possible to reduce the production of melanin, which has a skin whitening effect, which is known from the literature (J. Soc. Cosmet Chem. 42, 361, 1991).

Currently, melanin biosynthesis inhibitors have been applied to the treatment of skin diseases, skin whitening agents, food browning inhibitors, etc. in pharmaceuticals, cosmetics, foods, etc., the prevention of pigmentation in the skin is mainly studied from four perspectives. The first is to develop tyrosinase inhibitors or antagonists of tyrosinase substrates to modulate tyrosinase activity, the main enzyme of melanin synthesis. The second is to develop substances that are toxic to melanocytes in order to reduce the function of melanocytes, the site of melanin biosynthesis in animals. The third is to develop the waveguide reducing material to prevent the waveguide oxidation. Finally, there are methods to reduce the activity of the first enzyme tyrosinase, a melanin production mechanism, and an enzyme that catalyzes the conversion of dopachrome to DHICA.

Representative examples of melanin inhibitors have been developed so far arbutin, koji acid, ascorbic acid diphosphate magnesium, vitamin C, hydroquinone, plant extracts. However, kojic acid acts to inhibit enzymatic activity by chelating copper ions present in the active site of tyrosinase. Magnesium ascorbic acid phosphate ester is involved in the reaction for producing melanin from tyrosine to reduce the intermediate chain dopaquinone and to inhibit melanin production. The other is to act as a brown reducing type melanin by reducing the dark oxidized melanin by directly acting on melanin, but there is a problem in stability by rapid decomposition and coloration in solution. Vitamin C and its derivatives also have a whitening effect, but due to the instability that is well oxidized, it is difficult to use as a raw material for cosmetics. Due to its irreversible whitening effect and its high skin irritation, HITRO quinone has been limited in its use as a safety issue. Most plant extracts show relatively high inhibitory effect of tyrosinase activity only at high concentrations, and almost no inhibitory effects at low concentrations.

Recently, cosmetic raw material for skin whitening of silkworm Cordyceps sinensis mycelium culture medium using mulberry leaf extract of Korean Patent Application Publication No. 10-2008-0078371 (Publication No.) using mulberry leaf extract and silkworm Cordyceps mycelium mycelium. Obtained by culturing mushroom mycelium on a natural medium such as a whitening cosmetic containing the mushroom mycelium culture of Korean Patent Publication No. 10-0848515 (registration number), which is inoculated and cultured by inoculating the mushroom mycelium on a medium. Raw materials for various whitening cosmetics using cultures have been developed.

Aloe is a perennial herb belonging to the genus Aloe of the family Liaceae, and is said to be the best plant that has been used as a folk or herbal medicine for about 4,000 years, and among the various aloe varieties, aloe is used directly for food. , Aloe aboresense, saponaria, etc. There are three to four species. The main ingredients of aloe are polymer polysaccharide, alomichin, aloin, aloetic acid, aloe ulcin, etc., which shows the effects and effects of hypertension lowering, antitumor, psoriasis, and cell regeneration.

Aloe is developed as a health functional food only by the above effects and effects, and functional whitening cosmetic raw materials (materials) using aloe have not been developed yet.

Therefore, due to the limited development of aloe, the demand for aloe does not increase, which damages aloe growers. Therefore, there is an urgent need to develop high value-added technologies using aloe.

In order to solve the above problems, the present invention was invented, and to provide aloe fermented product for whitening cosmetic material having less skin irritation because it has excellent skin whitening effect and no cytotoxicity by culturing and fermenting mushroom mycelium with aloe as a natural medium. And also to provide a whitening cosmetics using the fermented product.

Aloe fermented product for whitening cosmetic material of the present invention to achieve the above object,

Inoculate the mushroom mycelium pre-cultivated in aloe-based natural medium at 5-15% (w / w), and the aloe natural medium in which the mushroom mycelium is inoculated has a temperature of 24-34 ° C. and aeration rate of 0.4-0.8vvm. Obtained by incubation and fermentation for 3 to 10 days under conditions.

The aloe natural medium, the aloe leaf is processed by harvesting-washing-coating-grinding-grinding step to obtain a viscous gel, the obtained viscous gel alone or viscous gel 25 to 99% by weight of purified water 1 to 75 weight The mixture is added in a fermentation culture tank and heated to a temperature of 113 ~ 125 ℃ for 2-30 minutes to sterilize to prepare aloe natural medium,

Alternatively, 0.5 ~ 7.0% by weight of dry aloe and 93.0 ~ 99.5% by weight of purified water are mixed and mixed in a fermentation culture tank, followed by sterilization by heating at 113-125 ° C. for 2-30 minutes to prepare aloe natural medium.

Mushroom mycelium is inoculated (cultured) as a spawn on the natural medium using a mushroom mycelium obtained by incubating for 4-8 days under the conditions of temperature 24 ~ 34 ℃, aeration 0.3 ~ 0.8vvm,

The mushroom mycelium used is Ganoderma lucidum lucidum ), Phellinus linteus ), Red Cordyceps militaris ), silkworm cordyceps ( Paecilomyces japonica ), Roe Deer Mushroom ( Hericium) erinaceum ), Agaricus blazei ( Agaricus blazei ) is selected from the group consisting of one is used alone.

Using the aloe of the present invention made as described above as a natural medium to ferment the mushroom mycelium using aloe fermented product for a whitening cosmetic material can be provided with a whitening cosmetic having no skin irritation, skin irritation effect.

In particular, the aloe fermentation product for the whitening cosmetic material is not added to the additives other than aloe and mushroom cells, it is possible to eliminate irritation to the skin by using a fermentation broth liquid cultured with mushroom mycelium in aloe natural medium.

The present invention is to inoculate the mushroom mycelium in a natural medium based on aloe, and to cultivate the mushroom mycelium under optimum conditions to obtain aloe fermented product for whitening cosmetic material.

The natural medium based on the aloe is usually used aloe leaves, and dry aloe may be used.

The aloe natural medium based on the aloe green leaf as a substrate, the aloe green leaf is obtained by collecting the viscous-washing-deriving-crushing step to obtain a gel of viscous phase in the fermentation culture tank in the viscous gel alone or viscous gel in 25 ~ 99% by weight of purified water It is prepared by adding 1 to 75% by weight, heat sterilizing at a temperature of 113 to 125 ° C. for 2 to 30 minutes, and quenching to room temperature (20 to 25 ° C.). Unlike other aloe fermentation methods that add other additives such as sugar, it uses pure 100% aloe as a natural medium.

In general, dried aloe is processed by harvesting aloe leaves, washing, peeling, and grinding to obtain a viscous gel, and the obtained viscous gel is lyophilized or dried by hot air drying.

The aloe natural medium based on the dried aloe is mixed with 0.5 to 7.0% by weight of dry aloe and 93.0 to 99.5% by weight of purified water, added to a fermentation broth, mixed and then heated to a temperature of 113 to 125 ° C. for 2 to 30 minutes to sterilize. It is prepared by treatment and quenching at a temperature of 20 to 25 ° C.

Mushroom mycelium inoculated in aloe natural medium is Ganoderma lucidum ), Phellinus linteus ), Red Cordyceps militaris ), Silkworm Cordyceps sinensis ( Paecilomyces) japonica ), Roe Deer Mushroom ( Hericium) erinaceum ), Agaricus blazei ( Agricus blazei ) is selected from the group consisting of 1 used alone.

The mushroom mycelium is used incubated for 4 to 8 days at a temperature of 24 ~ 34 ℃, aeration amount 0.3 ~ 0.8vvm conditions.

Inoculate mushroom mycelium pre-cultivated in natural aloe medium at 5-15% (w / w), and incubate and ferment for 3 to 10 days at the optimum temperature of 24 ~ 32 ℃, aeration 0.4 ~ 0.8vvm.

Through the following Examples to prepare aloe fermentation for whitening cosmetic material of the present invention.

Example  One

Ganoderma lucidum mushroom lucidum ) Aloe fermented product (GLAL) for whitening cosmetics using mycelium.

① Preparing the aloe natural medium (S1): After collecting the aloe leaves, washing, peeling, and grinding to obtain a viscous gel, add 10L to 20L bioreactor and sterilize at 120 ℃ for 20 minutes. Prepare a 100% aloe natural medium by quenching to a temperature of 25 ℃.

② Preparation of Ganoderma lucidum mycelium (seedling) (S2): In order to use Ganoderma lucidum mycelium as spawn (preculture), 5 days of primary liquid culture, 4 days of secondary liquid culture, Prepare the spawn (preculture) by incubating at the optimum condition (temperature 30 ℃, Shaking Rate 120 RPM) for 4 days in the tertiary liquid culture.

③ step of inoculating Ganoderma lucidum mycelium (S3): 8% (w) of the preculture (seed) prepared in the step (S2) preparing Ganoderma lucidum spawn in the bioreactor (Aloe-natural medium) prepared in the above step (S1) / w) inoculate.

④ Cultivating and fermenting aloe natural medium with aloe inoculated Ganoderma lucidum mycelium (S4): The optimum conditions (Temperature: 30 ℃, aeration amount 0.4 ~ 0.5vvm inoculated with Ganoderma lucidum mycelium in the step (S3) Fermentation for 4 days.

⑤ finally obtaining the aloe fermented product (S5): in the step (S4) to obtain the finished aloe fermented in low temperature hot water extraction, and then filtered (0.45㎛ ~ 0.26㎜) and concentrated to 10 ~ 30brix To obtain aloe fermented product for functional whitening cosmetic material, or to concentrate the filtered fermented product to 7 ~ 13 brix and then powdered by using a freeze dryer to obtain aloe fermented product powder for functional whitening cosmetic material.

Example 2

Phellinus linteus ) Aloe fermented product (PLAL) for whitening cosmetics using mycelium.

① Preparing the aloe natural medium (S1): After collecting the aloe leaves, washing, peeling, and grinding to obtain a viscous gel, add 10L to 20L bioreactor and sterilize at 120 ℃ for 20 minutes. Prepare 100% natural aloe broth by quenching to 25 ℃. (Add 2-3 drops of soybean oil which acts as a defoaming agent when natural aloe is added to bioreactor.)

② Preparation of the situation mushroom mycelium (seedling) (S2): 6 days of primary liquid culture, 6 days of secondary liquid culture after 6 days of plate culture to use the situation mushroom mycelium as a seed culture (preculture) Prepare the spawn (preculture) by incubating at the optimum condition (temperature 29 ℃, Shaking Rate 120 RPM) for 4 days in the tertiary liquid culture.

③ Inoculating the situation mushroom mycelium (S3): 7% (w) of the pre-culture (seed) prepared in the step (S2) to prepare the situation mushroom mycelium in the bioreactor (Aloe-natural medium) prepared in the step (S1) / w) inoculate.

④ Cultivation and fermentation of natural medium in aloe inoculated with mycelium mushroom mycelium (S4): The optimal condition (Temperature: 30 ℃, aeration amount 0.4 ~ 0.5vvm inoculated with Ganoderma lucidum mushroom in the step (S3) Fermentation for 4 days.

⑤ finally obtaining the aloe fermented product (S5): to obtain the aloe fermented culture in the step (S4) to extract the low temperature hot water, and then filtered (0.45㎛ ~ 0.26㎜) and concentrated to 10 ~ 30brix To obtain the aloe fermented product for functional whitening cosmetic material, or to concentrate the filtered fermented product to 7 ~ 13 brix and then powdered using a freeze dryer to obtain aloe fermented product powder for functional whitening cosmetic material.

Example  3

Roe Deer Mushroom ( Hericeum erinaceum ) Production of aloe fermented products (HEAL) for whitening cosmetic materials using mycelia.

① Preparing the aloe natural medium (S1): After collecting the aloe leaves, washing, peeling, and grinding to obtain a viscous gel, add 10L to the 20L bioreactor and sterilize at 120 ℃ for 20 minutes. Prepare 100% aloe natural medium by quenching to a temperature of 25 ° C.

② Preparing the scab mycelium mycelium (seedling) (S2): In order to use the scab mycelium as the spawn (preculture), the first liquid culture 6 days, the second liquid culture 4 after 7 days of plate culture Prepare the spawn (preculture) by culturing at the optimum condition (temperature 25 ℃, Shaking Rate 125 RPM) for 4 days in the first and third liquid cultures.

③ step of inoculating the roe deer mushroom seedlings (S3): 8% (w / w) of the roe deer mushroom preculture (seed) prepared in step (S2) in the bioreactor (aloe-natural medium) prepared in the step (S1). ) Inoculate.

④ Cultivation and fermentation of natural aloe vera medium inoculated with roe deer mushroom spawn (S4): in a step (S3) aloe natural medium inoculated with roe deer mycelium optimum conditions (Temperature: 25 ℃, aeration 0.5 ~ Culture fermentation for 5 days at 0.6vvm).

⑤ finally obtaining the aloe fermented product (S5): to obtain the aloe fermented culture in the step (S4) to extract the low temperature hot water, and then filtered (0.45㎛ ~ 0.26㎜) and concentrated to 10 ~ 30brix To obtain aloe fermented product for functional whitening cosmetic material, or concentrated to 7 ~ 13 brix filtered fermented product and then powdered using a freeze dryer to obtain aloe fermented product powder for functional whitening cosmetic material.

The aloe fermented powder for functional whitening cosmetic material obtained as described above was subjected to a melanogenesis inhibition assay, and the test method is a guideline for the evaluation of the effectiveness of functional cosmetics [1] (Food and Drug Administration, 2003). Followed.

Test Methods

This test method is to compare the amount of melanin produced in the cell culture with the blank sample.

In this experiment, murine melanoma (B-16 F1) or similar cells were inoculated with DMEM containing an appropriate amount of FBS (fetal bovine serum) or 1 × 105 cells per well in a 6-well plate after growth. Incubate at 5% CO ₂ , 37 ° C until the cells adhere to at least 80% of the bottom of the well.

After incubation, the medium is removed, the sample is replaced with a medium diluted to an appropriate concentration, and then incubated at a constant time at 5% CO ₂ , 37 ° C. The concentration range of the sample is determined by toxicity test using crystal violet assay. The cells from which the medium has been removed are washed with PBS (phosphated buffer saline) and treated with trypsin to recover the cells. The recovered cells are measured by using a hematocytometer and centrifuged at 5,000 to 10,000 rpm for 10 minutes, and then the supernatant is removed to obtain pellets.

The cell pellet was dried at a temperature of 60 ° C., and then 100 μl of 1M sodium hydroxide solution containing 10% DMSO (dimethyl sulfoxide) or an appropriate amount of cell lysis buffer was used to obtain intracellular melanin in a 60 ° C. thermostat.

Using this solution, measure the absorbance at 490 nm with a microplate reader to determine the amount of melanin per fixed cell number or melanin per constant protein. Perform a blank test and calibrate.

The B16 F10 cells used in the experiment were purchased from the Bank of Korea Cell Line, Histopathology was melanoma, Media was Minimum essential medium, 25mM HEPES (4- (2-hydroxyethyl) -1-piperazineethanesulfonic acid) and 25mM NaHCO3, 90%; heat inactivated fetal bovine serum (FBS), 10%.

1) Determination of sample concentration through the toxicity test results

No significant cytotoxicity was seen at all concentrations treated (high: 300 ug / ml, low: 30 ug / ml) for sample determination including safety. In the melanin production inhibition test, it was decided to treat at a concentration of 300 ug / ml, 30 ug / ml (final concentration), respectively.

TABLE 1 Cytotoxicity test in B16F10 cells. High: 300 ug / ml, low: 30 ug / ml (final concentration).

I) Intracellular  Melanin production inhibition test result

Comparison of melanin production in B16F10 cells. High: 300 ug / ml, low: 30 ug / ml (final concentration) (see FIG. 5).

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The experiments were conducted at two concentrations of 300 ug / ml and 30 ug / ml (final concentration), respectively. When the blank was converted to 100%, melanin production was 100 ± 3.4%.

In the Arbutin-treated group, melanin production was 96.5 ± 3.2% and 97.3 ± 1.2% at high and low concentrations, respectively.

In the kojic acid-treated group, melanin production was 82.1 ± 1.7% and 82.0 ± 5.0% at high and low concentrations, respectively.

The NRAL treatment group showed melanin production of 95.0 ± 4.9% at high concentrations and 92.1 ± 4.9% at low concentrations, and no significant effects were observed.

The GLAL treatment group had no significant effect at high concentrations (96.6 ± 1.3%) but had a significant inhibitory effect on melanogenesis (87.3 ± 0.4%).

The HEAL treatment group showed no significant changes at high concentrations (93.4 ± 4.1%) and suppressed melanogenesis at low concentrations (76.0 ± 3.9%).

• The group treated with PLAL samples showed no effect at high concentrations (111.4 ± 3.8%) but showed a significant effect at low concentrations (74.8 ± 0.4%).

C) summary of experimental results

As a result of observing the melanin production inhibitory effect of NRAL, GLAL, HEAL, PLAL samples, it was confirmed that NRAL did not inhibit melanin production, but GLAL, HEAL, PLAL had a melanin production inhibitory effect.

1 is a fermentation product of fermented ganoderma lucidum in aloe natural medium prepared with aloe leaves in accordance with an embodiment of the present invention.

Figure 2 is a fermentation product of fermenting a situation mushroom in aloe natural medium prepared with aloe leaves in accordance with an embodiment of the present invention.

Figure 3 is a fermentation product of fermented Militaris cordyceps in aloe natural medium prepared with aloe leaves in accordance with an embodiment of the present invention.

4 is a fermentation product of fermenting Ganoderma lucidum mushroom in aloe natural medium prepared with dry aloe according to an embodiment of the present invention.
5 is a comparison table of melanin production in B16F10 cells

Claims (6)

In the whitening cosmetic material which uses mushroom mycelium culture inoculated by inoculating mushroom mycelium on natural medium as an active ingredient, Aloe is a natural medium, Ganoderma lucidum , Ganoderma lucidum , Phellinus linteus , Red Cordyceps ( Cordyceps militaris ), Silkworm Cordyceps ( Paecilomyces japonica ), Inoculated with 5-15% (w / w) of mushroom mycelium which is used alone by selecting one species from the group consisting of roe deer mushroom ( Hericium erinaceum ) and Agaricus blazei , Aloe fermented for whitening cosmetic material, characterized in that the cultured aloe inoculated with the mushroom mycelium cultured and fermented for 3 to 10 days at a temperature of 24 ~ 34 ℃, aeration 0.4 ~ 0.8vvm. The method of claim 1, The aloe natural medium, the aloe leaf is processed by harvesting-washing-coating-grinding-grinding step to obtain a viscous gel, the obtained viscous gel alone or viscous gel 25 to 99% by weight of purified water 1 to 75 weight Aloe fermented product for whitening cosmetics, characterized in that the mixture is added in a fermentation culture tank and heated to a temperature of 113-125 ° C. for 2 to 30 minutes to sterilize to prepare aloe natural medium. The method of claim 1, The aloe natural medium is mixed with 0.5 ~ 7.0% by weight of dry aloe and 93.0 ~ 99.5% by weight of purified water, added to the fermentation culture and mixed, and then heated to a temperature of 113 ~ 125 ℃ for 2 to 30 minutes sterilized by aloe natural medium Aloe fermented product for whitening cosmetic material, characterized in that the production. The method of claim 1, The mushroom mycelium is aloe fermented for whitening cosmetic material, characterized in that using the mushroom mycelium obtained by incubating for 4-8 days at a temperature of 24 ~ 34 ℃, aeration amount 0.3 ~ 0.8vvm. A whitening cosmetic using the aloe fermented product for whitening cosmetic material of any one of Claims 1-4. delete

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