JP5996706B1 - Tsurmurasaki extract and its use for whitening and sun protection - Google Patents

Abstract

The present invention provides a whitening composition, an anti-ultraviolet composition, or an anti-inflammatory composition. A vineyard extract, which is obtained by adding water or an alcohol to a concentrated liquid obtained by gluing vineyards and heating at 50 to 150 ° C. for about 1 to 6 hours. Extract extract. Use in ointments, creams, lotions, liquid medicines, cosmetic masks, bath essences as a whitening composition, anti-ultraviolet composition or anti-inflammatory composition containing the extract of the vineyard extract. [Selection] Figure 1

Description

The present invention relates to Malabar spinach extract and uses thereof, whitening agents including Malabar spinach extract as an active ingredient, sunscreen agents, skin external preparation containing 及 Bisore, foods, cosmetics, medicines relates like.

  Skin color is affected by melanin in the epidermis. Melanin produced by melanocytes in the basal cell layer is transferred to the surface of the skin through the stratum corneum and pigmented.

  Melanin is produced by a series of chemical reactions and the action of active enzymes. When the skin is irradiated with ultraviolet rays, tyrosinase in melanocytes is activated, tyrosine in melanocytes is oxidized to form dihydroxyphenylalanine (commonly called DOPA), and DOPA is dopachrome by tyrosinase. It is metabolized to (dopachrome), and melanin is synthesized through an oxidation reaction.

Tyrosinase is a polyphenol oxidase containing copper ions. It is synthesized by melanocytes, and since there are three active centers of copper ions at the site where tyrosinase binds to tyrosine, it suppresses tyrosinase. Will use two types of mechanisms:
(1) Inhibits tyrosinase activity by binding to copper ions (2) Promotes competitive inhibition with tyrosinase substrate

  Since the skin is always exposed to ultraviolet rays, various cell hormones and inflammation-causing factors are stimulated to cause inflammation, leading to the development of allergies and spots.

  Among them, kojic acid, arbutin, and the like are applied ingredients for preventing skin darkening phenomenon, spots and freckles and maintaining clean skin, but these are not sufficiently satisfactory. Vitamin C and its derivatives are known as whitening agents that suppress inflammation caused by ultraviolet rays, but they are still insufficient in terms of effects and stability in the preparation.

  The object of the present invention is to (1) add water or alcohol to a concentrated liquid obtained by grated Tsurmuraki extract and heat at 50 to 150 ° C. for about 1 to 6 hours. (2) a liquid rubbed with a paper filter Is obtained by precipitating for 1 to 48 hours in ethanol, and (3) removing the supernatant to obtain a water extract of Tsurmurasaki water or an extract of Tsurmurasaki alcohol to obtain a Tsurmurasaki extract (Basela alba). .

  In order to achieve the object, the alcohols in step (1) are ethanol, glycerol (Glycerin), 1,3-butylene glycol (1,3-Butylene Glycol), and propylene glycol (Propylene Glycol).

  Another object of the present invention is to provide a use of the vineyard extract used for the preparation of the whitening agent. In this case, the extract is the above vineyard extract. Moreover, the pickled vine extract in the present invention has effects such as suppression of skin blackening and suppression of spots.

  Another object of the present invention is to provide a use of the vineyard extract used as an anti-ultraviolet agent. In this case, the extract is the above vineyard extract.

  Another object of the present invention is to provide a use of the vineyard extract used as an anti-inflammatory agent. In this case, the extract is the above vineyard extract.

  Another object of the present invention is to provide a food containing the above pickled vine extract.

Inhibitory amount of melanin production by water extract from Tsurmurasaki and alcohol extract Test for inhibiting tyrosinase activity of water extract from Tsurmurasaki and alcohol extract Tsurumura water extract anti-UV test Tsurmurasaki Alcohol Extract Extract Anti-UV Test IL-6 inhibitory amount of Tsurumura water extract and alcohol extract Tsurmurasaki water extract, alcohol extract extract TNF-α suppression amount

  Unless otherwise defined, technical and scientific terms used in this document are interpreted based on recognition shared by those with ordinary skill in the domain to which the term belongs.

  The composition of the external preparation for skin in the present invention may be appropriately added based on components used for external preparations for skin such as general cosmetics and pharmaceuticals according to the dosage form and necessity.

  Application forms of the vineyard extract of the present invention include, but are not limited to, food, drinking water, health food, animal and human pharmaceutical compositions, food additives, and beverage additives.

  In the above-mentioned Tsurumurasaki extract, one or more kinds of solubilizing agents, buffering agents, preservatives, coloring agents, fragrances, and flavoring agents that are usually used in the formulation area may be used as necessary.

  As another embodiment of the present invention, when an additive is added to the vineyard extract, it is selected from the following group. Vitamin C and its derivatives (Ascorbic Acid and Its Derivatives), Kojic acid (Kojic Acid), Arbutin (Arbutin), Ellagic acid (Ellagic Acid), Camomila ET (Chamomile ET), Tranexamic acid (Trexamic acid) Nicotinamide), potassium methoxysalicylate (Potassium Methoxylate).

  Although the Example of this invention is described below, it is not restricted to this.

<Example 1>
The raw material used in the present invention is Basella alba L .. Tsurmurasaki extract is prepared by the following procedure.
1. Place 300 grams of grated Tsurumura Saki in a beaker, add secondary water or 500 ml of 80% alcohol, and make a hot water bath at 90 ° C. for 3 hours in a constant temperature bath.
2. After cooling, scrape with a 125 mm paper filter, mix the filtrate and 99.5% anhydrous alcohol in a ratio of 1: 2, and precipitate for one day.
3. The preform obtained by centrifuging at a rotational speed of 8000 rpm for 15 to 20 minutes and removing the supernatant is placed in an environment of −80 ° C. for one day to freeze-dry to obtain a Tsurumura powder sample.

<Example 2>
Melanin content measurement (1) 3 × 10 ⁴ cells / well of melanoma B16-F0 cells are cultured for 24 hours.
(2) Tsurmurasaki water extract and alcohol extract were each prepared as a single experiment group of 10 μg / ml and 50 μg / ml (measured with the crude extract concentration of organic matter), and ascorbate (Ethyl) Ascorbic Acid (2.5 mg / ml, 5 mg / ml) is added to the combined experimental group, and ethyl ascorbate (10 mg / ml) is prepared as a positive target and cultured for 24 hours.
(3) After washing twice with PBS, the sample was air-dried, dissolved in 200 μl of 1M sodium hydroxide containing 10% DMSO, heated at 80 ° C. for 1 hour, cooled, and the absorbance was measured at 475 nm. taking measurement.

  In this experiment, as a result of using commercially available ethyl ascorbate as a positive control group, it was found that the water extract of Tsurumurasaki has a good melanin suppressing effect and is superior to ethyl ascorbate (FIG. 1). ).

<Example 3>
Measurement of inhibition rate of tyrosinase (1) Disodium hydrogen phosphate-potassium dihydrogen phosphate (1/15 M, pH 6.8) is prepared as a buffer solution.
(2) Prepare a tyrosinase solution.
(3) Prepare 0.03% L-DOPA.
(4) 50 μL of the sample to be measured and 50 μL of tyrosinase are mixed until they are uniform and left for 15 minutes.
(5) Add 50 μL of 0.03% L-DOPA and leave for 15 minutes.
(6) Measure at OD475nm.

The experimental groups are as follows.
(1) 1% vitamin C,
(2) 0.25% vitamin C,
(3) 10 μg / ml Tsurumura water extract (H10),
(4) 0.25% vitamin C + 10 μg / ml Tsurmurasaki water extract (0.25VitC + H10),
(5) 50 μg / ml Tsurumura water extract (H50),
(6) 0.25% Vitamin C + 50 μg / ml Tsurmurasaki Water Extract (0.25VitC + H50),
(7) 10 μg / ml Tsurumurasaki alcohol extract (E10),
(8) 0.25% vitamin C + 10 μg / ml Tsurumurasaki alcohol extract (0.25VitC + E10),
(9) 50 μg / ml Tsurumurasaki alcohol extract (E50),
(10) 0.25% vitamin C + 50 μg / ml Tsurmurasaki alcohol extract (0.25VitC + E50).

<Experimental result>
In this experiment, it has been demonstrated that Tsurmurasaki extract suppresses the activity of tyrosinase, and the Tsurmurasaki water extract extracts have a better effect of suppressing tyrosinase activity than 1% vitamin C derivative. In the case of the mixing action, the effect is higher when the amount of the vineyard extract is larger (FIG. 2).

<Example 4>
A human fibroblast cell line WI38 (ATCC: CCL-75) is cultured, and UV biological activity is measured and analyzed. In the anti-UV analysis method, cells are irradiated with UV 300 mJ / cm ² for 3, 6, 9, 12, 24 hours. The control group is not irradiated. Analyze cell viability (MTT Assay).

<Survival analysis>
(1) Cells are cultured in a 96-well culture tray (5000 cells / 100 μl / well) and left until the next day.
(2) After irradiation for 6-12 hours according to the experimental group, the culture solution is removed, 100 μl of culture solution containing 500 μg / ml MTT is added, and the mixture is allowed to act at 37 ° C. for 1 hour.
(3) Absorb and remove the culture solution containing MTT, add 100 μl DMSO to dissolve, and read the absorbance at 570 nm using an ELISA reader.

The anti-ultraviolet ability of the WI38 cell line is evaluated at 300 mJ / cm ² UV irradiation for 6 hours. As a result of the experiment, the water extract of Tsurumurasaki water has no effect of protecting the anti-ultraviolet ray ability of the cells (FIG. 3), and the extract of Tsurumurasaki alcohol has a remarkable effect of protecting the anti-ultraviolet ray ability of the cells (FIG. 4).

<Example 5>
WI38 is cultured into cells for analysis of the pro-inflammatory molecule IL-6.
Proinflammatory molecule IL-6 test (1) Add 100 μl primary antibody to each well of a 96-well tray and leave at 4 ° C. for about 16-18 hours.
(2) Wash three times with a wash buffer (300 μl each time, tap to dry).
(3) Add 200 μl 1X assay buffer to each well and leave for 1 hour.
(4) Wash three times using a wash buffer (300 μl each time, tap to dry).
(5) Prepare a standard curve and sample, add 100 μl to each well, and let stand for 2 hours.
(6) Wash three times using a wash buffer (300 μl each time, tap to dry).
(7) Add 100 μl secondary antibody to each well and let stand for 1 hour.
(8) Wash three times with a wash buffer (300 μl each time, tap to dry).
(9) Add 100 μl of HRP to each well and leave it for 30 minutes.
(10) Wash 4 times using a wash buffer (300 μl each time, tap to dry).
(11) Add 100 μl TMB mixture to each well and leave for 15 minutes avoiding light.
(12) Add 100 μl stop solution (2N H ₂ SO ₄ ) to each well and leave it for 30 minutes.
(13) Measure OD450.

  The suppression amount of the pro-inflammatory molecule IL-6 is 60% or more without UV irradiation and 30% or more with UV irradiation. The extract of 5 μg / ml Tsurumurasaki Alcohol had an IL-6 suppression amount of 70% or more in an environment without UV irradiation, and an IL-6 suppression amount of 40% or more was recorded in the UV irradiation environment (FIG. 5).

<Example 6>
WI38 is cultured into cells for analysis of the pro-inflammatory molecule TNF-α. The method for measuring TNF-α is the same as in Example 4 described above, and only the TNF-α antibody is measured.

  As a result of the experiment, 5 μg / ml Tsurmurasaki water extract recorded a TNF-α suppression amount of 70% or more without UV irradiation, and a TNF-α suppression amount of 30% or more even under UV irradiation environment. The extract of 10 μg / ml Tsurmurasaki alcohol had a TNF-α suppression amount of 50% or more without UV irradiation, but could not suppress TNF-α under the UV irradiation environment (FIG. 6).

Claims (2)

A whitening composition or an anti-ultraviolet composition of Basella alba extract,
The vineyard extract is
(1) Water or alcohol is added to a concentrated liquid obtained by grated tsurumurasaki and heated at 50 to 150 ° C. for 1 to 6 hours.
(2) The liquid of (1) rubbed with a paper filter is precipitated in ethanol for 1 to 48 hours.
(3) The supernatant liquid is removed from the liquid of (2), and a Tsurmurasaki water extract or Tsurmurasaki alcohol extract is obtained .
These compositions by Seisuru the skin external or oral administration, or other suitable dosage forms, food, cosmetics, health such chemicals, cosmetic, is applied to pharmaceutical applications,
The external preparation for skin is selected from the group consisting of ointments, creams, lotions, liquid medicines, cosmetic masks, and bath essences ,
A whitening composition or an anti-ultraviolet composition of Tsurmurasaki extract . A whitening compositions or anti-UV composition of claim 1,
Moisturizer, emulsifier, antioxidant, UV absorber, surfactant, plant extract ingredient, various nutrients, vitamin C and its derivatives, kojic acid, arbutin, ellagic acid, chamomile ET, nicotinamide, tranexamic acid, A whitening composition or an anti-ultraviolet composition of Tsurumura extract, to which an additive selected from the group consisting of potassium methoxysalicylate and an oil-soluble or water-soluble component applicable to the composition of claim 1 is added .