WO2008114988A1

WO2008114988A1 - Whitening cosmetic composition comprising an extract of halocynthia species shell

Info

Publication number: WO2008114988A1
Application number: PCT/KR2008/001510
Authority: WO
Inventors: Ji-Ho Park; Sun-Yeou Kim; Min-Jae Lee; Tae-Jin Yoon
Original assignee: Handock Cosmetics Co. Ltd.
Priority date: 2007-03-20
Filing date: 2008-03-18
Publication date: 2008-09-25
Also published as: KR20080085432A

Abstract

The present invention provides a whitening cosmetic composition comprising an extract of Halocynthia sp. shell. The extract of Halocynthia sp. shell effectively reduces the melanin pigment formation level without causing cytotoxicity. Therefore, the extract is useful for whitening cosmetics for e.g., face or skin.

Description

WHITENING COSMETIC COMPOSITION COMPRISING AN EXTRACT OF

HALOCYNTHIA SPECIES SHELL

TECHNICAL FIELD

The present invention relates to a whitening cosmetic composition comprising an extract of Halocynthia sp. shell.

BACKGROUND ART

The skin color of human beings depends on the content of melanin, carotene, and hemoglobin therein. Among them, melanin is the most determining element. Melanin is synthesized in melanocytes, i.e., pigment cells existing in the stratum basale within epidermis, and the synthesized melanin is transferred to neighboring keratin cells, thereby determining the skin color. When melanin is abnormally generated in excess, freckles are formed. On the other hand, when melanin is generated abnormally in a small amount, skin lesions such as leucoderma occur. Pigmentation is caused by various external stimuli, such as exposure to ultraviolet light or inflammations, and a level of pigmentation is proportional to the reaction level of melanocytes existing in the stratum basale within epidermis. The mechanism of pigmentation caused by UV light is based on tyrosinase. Various external stimuli induce pigmentation by facilitating activation of melanocyte-activating factors promoting tyrosinase expression and materials involved in the chain reaction by arachidonic acid, one of chemical mediators in inflammatory system. In addition, it is known that oxygen free radicals whose generation is facilitated by UV light irradiation are involved in the pigment formation. It is generally known that the mechanism of melanin excretion includes melanosome transfer from the melanocytes to the neighboring keratinocytes. Recently, it is reported that pigmentation occurs when melanosome is transferred to keratinocytes. Melanocyte-activating factors activate melanocytes by mediating α-MSH

(melanocyte-stimulating hormone) and ACTH (adrenocorticotrophin) in keratinocytes (Suzuki I. et al., J. Endocrinol., 137, p1627, 1996). ET-1 (Endothelin-1), which is formed in vascular endothelial cells and has an activity of blood vessel constriction, activates PKC (protein kinase C) and adenylcyclase systems to facilitate melanocyte proliferation and tyrosinase activation, thereby resulting in pigmentation (Imokawa G. et al., J. Invest. Dermatol., 105, p32, 1995). Recently, it has been reported that ET-1 is generated in keratinocytes by UVB irradiation (Yada Y. et al., J. Biol. Chem., 266, p18352, 1991). In addition, it is reported that, as another factor, SCF (stem cell factor) and its receptor, i.e., c-kit play an important role in the migration and proliferation of primary melanocytes (Grabbe J. et al., Arch. Dermatol. Res., 287, p78, 1994). Meanwhile, various whitening cosmetic compositions containing an herbal extract or a material isolated therefrom have been disclosed. For example, KR Patent Registration No. 654,959 discloses a whitening cosmetic composition including extracts of Corni Fructus, Lycii Fructus, Rubi Fructus, Pini Semen, and pearl powder. KR Patent Registration No. 614,467 discloses a whitening cosmetic composition including an extract of Euphorbiae Helioscopiae Herba, Moutan Cortex, and Aloe as an active ingredient. KR Patent Registration No. 628,651 discloses a whitening cosmetic composition including Mori Ramulus extracts and Neem tree extracts as an active ingredient. KR Patent Registration No. 560,408 discloses a whitening cosmetic composition including methyl trans-cinnamate extracted from Tricholoma Matsutake Mushroom. In addition, KR Patent Registration No. 602,684 discloses a whitening cosmetic composition including an extract of Erigeron breviscapus as an active ingredient.

DETAILED DESCRIPTION OF THE INVENTION

TECHNICAL PROBLEM

The present Inventors have performed extensive studies in order to develop various natural product-derived sources which are useful for whitening cosmetics; and found that an extract of Halocynthia sp. shell effectively reduces the melanin pigment formation level. Therefore, it is an object of the present invention to provide a whitening cosmetic composition including an extract of Halocynthia sp. shell.

TECHNICAL SOLUTION

According to an aspect of the present invention, there is provided a whitening cosmetic composition comprising an extract of Halocynthia sp. shell.

ADVANTAGEOUS EFFECTS

The extract of Halocynthia sp. shell effectively reduces the melanin pigment formation level without causing cytotoxicity. Therefore, the extract is useful for whitening cosmetics for e.g., face or skin.

BRIEF DESCRIPTION OF THE DRAWINGS

FIG. 1 is a graph showing the effects of the extract of Halocynthia sp. shell obtained in Example 1 on melanin formation by melanocytes and cell viability;

FIG. 2 is a graph showing the effects of the extract of Halocynthia sp. shell obtained in Example 2 (aqueous fraction) on melanin formation by melanocytes and cell viability;

FIG. 3 is a graph showing the effects of the extract of Halocynthia sp. shell obtained in Example 2 (butanol fraction) on melanin formation by melanocytes and cell viability; and FIG. 4 is a graph showing the effects of PTU on melanin formation by melanocytes and cell viability.

BEST MODE FOR CARRYING OUT THE INVENTION

There are two kinds of Halocynthia sp. in Korea: Halocynthia roretzi living or being cultured on the southern and western seashores and Halocynthia aurantium living on the eastern seashore. It is known that Halocynthia roretzi has halocyamine, an antimicrobial tetrapeptide-like substance (Azumi, K. et al., Experientia, 1990, 46, 1066-1068; K. Azumi et al., Biochemistry, 1990, 29, 159-165). Halocynthia aurantium has an antimicrobial peptide, referred to dicynthaurin (Lee, I. H. et al., Biochem. Biophys. Acta, 2001 ; and KR Patent Registration No. 491 ,423). A whitening cosmetic composition according to the present invention may include an extract obtained from shells of any kind of Halocynthia species, including Halocynthia roretzi and Halocynthia aurantium.

The extract of Halocynthia sp. shell included in a whitening cosmetic composition according to the present invention may be obtained by extracting Halocynthia sp. shell with water, CrC₄ alcohol, or a mixed solvent of water and C1-C₄ alcohol, preferably water, as an extraction solvent.

In preparing the extract of Halocynthia sp. shell, the extraction temperature may be in the range of about 0 to 100 "C, preferably room temperature, although the temperature depends on the extracting solvent used. The amount of the extracting solvent used may be in the range of about 1 to about 20 times greater than that of Halocynthia sp. shell. For example, the extracting solvent may be used in an amount of about 500 to 2000 ml, preferably about 1000 ml, per 30 g of Halocynthia sp. shell. The extraction time may be in the range of about 1 to 3 hours and the extraction may be performed once or more than once, although not being limited to these.

The extractaction for preparing an extract of Halocynthia sp. shell may be performed by cold extraction, hot extraction, superfluid extraction, centrifugal extraction, sonication-extraction, reflux- extraction, or the like. If necessary, the extraction may be performed after drying Halocynthia sp. shell using conventional drying methods, such as a freeze-drying or drying under reduced pressure. Preferably, the extract of Halocynthia sp. shell may be obtained in a liquid form by sonicating Halocynthia sp. shell (specifically, dried Halocynthia sp. shell) in water and then filtering the resultant extract solution using conventional methods to remove impurities. More preferably, the extract of Halocynthia sp. shell may be obtained in a powder form by drying the resultant extract solution obtained from sonicating Halocynthia sp. shell, using conventional drying methods, such as drying under reduced pressure, freeze-drying, drying with Speed Savant, or the like.

The extract of Halocynthia sp. shell included in a whitening cosmetic composition according to the present invention may be obtained by performing an extraction process including additional purification steps, that is, an active-fraction extraction process. An extract solution obtained from the sonication-extraction process may be directly applied to the active-fraction extraction process; or a powder form, which is made from an extract solution obtained from the sonication-extraction process, may be applied to the active-fraction extraction process. That is, the extract of Halocynthia sp. shell may be obtained by an extraction process comprising: (a) sonicating Halocynthia sp. shell in water; (b) adding ethyl acetate to the extract solution obtained in Step (a) to obtain an aqueous fraction; and (c) adding butanol to the aqueous fraction obtained in Step (b) to obtain an aqueous fraction. Alternatively, the extract of Halocynthia sp. shell may be obtained by an extraction process comprising: (a') sonicating Halocynthia sp. shell in water; (b^J) drying the extract solution obtained in Step (a¹) to obtain a powder; (c¹) adding water and ethyl acetate to the powder obtained in Step (b¹) to obtain an aqueous fraction; and (d^J) adding butanol to the aqueous fraction obtained in Step (c¹) to obtain an aqueous fraction. The extraction process including additional purification steps (i.e., an active-fraction extraction process) may further comprise a drying step of the aqueous fraction, e.g., freeze-drying, drying with Speed Savant, after performing the extraction process.

The whitening cosmetic composition according to the present invention includes the extract of Halocynthia sp. shell as an effective ingredient, in an effective amount of achieving a whitening function, e.g., in the range of 0.0001 to 20.0 % by weight, preferably in about 2 % by weight, based on the total weight of the composition. When the amount of the extract of Halocynthia sp. shell is less than 0.0001 % by weight, it may be difficult to obtain desired whitening effects. On the other hand, when the content of extract of Halocynthia sp. shell is greater than 20.0 % by weight, skin irritation may occur and stability of the formulation obtained therefrom may be affected. The whitening cosmetic composition according to the present invention may include, in addition to the extract, any conventional components used in the filed of a cosmetic composition, such as, additives, e.g., an antioxidant, a stabilizer, a solubilizer, vitamins, dyes, or perfumes; and a carrier. The whitening cosmetic composition according to the present invention may be formulated into any conventional types of formulations in the art. For example, the whitening cosmetic composition may be formulated into, but not limited to, solution, suspension, emulsion, paste, gel, cream, lotion, powder, soap, surfactant-containing cleansing, oil, powder foundation, emulsion foundation, wax foundation, or spray. Specifically, the whitening cosmetic composition can be formulated into softening skin lotion, nourishing milk lotion, nourishing cream, massage cream, essence, eye cream, cleansing cream, cleansing foam, cleansing water, packs, spray, or powder.

When the whitening cosmetic composition is formulated into paste, cream, or gel, an available carrier may be animal oil, plant oil, wax, paraffin, starch, tragacanth, cellulose derivatives, polyethylene glycol, silicon, bentonite, silica, talc, zinc oxide, or the like.

When the whitening cosmetic composition is formulated into powder or spray, an available carrier may be lactose, talc, silica, aluminum hydroxide, calcium silicate, or polyamide powder. Specifically, when the whitening cosmetic composition is formulated into spray, a propellant such as chlorofluorohydroncarbon, propan/butane, or dimethyl ether may be included.

When the whitening cosmetic composition is formulated into solution or emulsion, an available carrier may be a solvent, a solubilizer, or an emulsifier, and specifically water, ethanol, isopropanol, ethyl carbonate, ethyl acetate, benzyl alcohol, benzyl benzoate, propylene glycol, 1 ,3-butylglycol oil, glycerol aliphatic esters, polyethylene glycol, or sorbitan fatty acid esters.

When the whitening cosmetic composition is formulated into suspension, an available carrier may be a liquid diluent, such as water, ethanol, or propylene glycol; a suspending agent, such as ethoxylated isostearyl alcohol, polyoxyethylene sorbitol esters, or polyoxyethylene sorbitan esters; microcrystalline cellulose; aluminum metahydroxide; bentonite; agar; or tragacanth. When the whitening cosmetic composition is formulated into surfactant-containing cleansing, an available carrier may be aliphatic alcohol sulfate, aliphatic alcohol ether sulfate, sulfosuccinc acid monoester, isethionate, imidazolinium derivatves, methylta urate, sarcosinate, fatty acid amide ester sulfate, alkylamidobetain, aliphatic alcohol, fatty acid glycerides, fatty acid diethanolamides, plant oil, lanolin derivatives, or ethoxylated glycerol fatty acid esters.

In addition, the whitening cosmetic composition according to the present invention may further include, in addition to the extract of Halocynthia sp. shell, other conventional materials having a whitening activity. In this regard, it would be obvious to one of ordinary skill in the art to determine the kind and content of conventional materials.

Hereinafter, the present invention will be described more specifically with reference to the following examples and experimental examples. The following examples are for illustrative purposes only and are not intended to limit the scope of the invention.

Example 1. Preparation of extract of Halocynthia sp. shell

The shell of Halocynthia roretzi was freeze-dried. 30 g of the obtained dried sample was sonicated in 1 L of water for one hour at room temperature. The sonication process was performed three times. Each extracts were filtered with a filtering sheet and the filtrate was collected. The combined filtrate was concentrated under reduced pressure using a rotary vacuum evaporator to obtain 1.7 g of an extract of Halocynthia sp. shell in a powder form.

Example 2. Preparation of extract of Halocynthia sp. shell

500 ml of water was added to 1.7 g of the powdered extract prepared in Example

1 to obtain a suspension. 500 ml of the suspension and 500 ml of ethyl acetate were added to a separatory funnel and then separated into an ethyl acetate fraction and an aqueous fraction. 500 ml of water-saturated butanol was added to the obtained aqueous fraction and then separated into a butanol fraction and an aqueous fraction. Each ethyl acetate fraction, butanol fraction, and aqueous fraction obtained in the above were concentrated under reduced pressure using a rotary vacuum evaporator and then freeze-dried to obtain extracts of Halocynthia sp. shell.

Experimental Example

1. CeII Culture

Melan-A cell line (Skin Research Center of Pacific Co. Ltd) was incubated in a RPMI 1640 (Gibco, Invitrogen, USA) supplemented with 10 % Fetal Bovine Serum (Gibco, Invitrogen, USA) and 200 nm of phorbol 12-myristate 13-acetate (Sigma). About 5 x 10⁵ of cells were inoculated on 10 mi of the culture medium in a 100π cell culture plate and then incubated for three to four days at 37 ^°C , under 5% CO₂. When the cells were fused and grew, 24 well plates were inoculated with 1 X 10⁵ cells per well and incubation was performed for 24 hours. While exchanging with 990 ul of the culture medium everyday, each wells were treated with the extract of Halocynthia sp. shell prepared in Example 1 , the extracts of Halocynthia sp. shell prepared in Example 2 (butanol fraction and aqueous fraction), or PTU (phenylthiourea; Tokyo Kasei Kogyo Co Ltd, Japan) consecutively for three days, each being in 10 ul of volume at the concentration of 1 , 10, or 100 ug/ml (in a mixture of 50% propylene glycol, 30% ethanol, and 20% distilled water), and then incubation was performed for one day. Herein, PTU, which is a melanin formation inhibitor, was used as a positive control.

2. Measurement of the melanin pigment formation level

In order to identify inhibitory effects of the extract of Halocynthia sp. shell on melanin pigmentation, the melanin pigment formation level was measured. After the culture medium was removed from each well, the cells were washed with PBS and then

1 ml of 1 N sodium hydroxide was added thereto to dissolve melanin. Then, the absorbance was measured at a wavelength of 405 nm in ELISA. This experiment was performed three times. An average of three test results and a standard deviation are shown in Table 1 and FIGS. 1-4. Table 1

Concentration Melanin Formation Level (%)

Extract of Halocynthia sp. 1 ug/ml 83.7 + 4.2 shell of Example 1 10 ug/ml 72.0 ± 5.7

100 ug/ml 52.0 + 7.9

Extract of Halocynthia sp. 1 ug/ml 102.8 + 4.1 shell of Example 2 10 ug/ml 80.8 ± 7.9 (Aqueous Faction)

100 ug/ml 60.9 + 4.9

Extract of Halocynthia sp. 1 ug/ml 69.9 1.2 shell of Example 2 10 ug/ml 56.1 4.8 (Butanol Faction)

PTU 1 ug/ml 87.6 ± 1.1

10 ug/ml 48.0 ± 1.4

100 ug/ml 25.1 ± 8.8

3. Measurement of Cell Viability

In order to identify effects of the extract of Halocynthia sp. shell on melan-A cell viability, the living cells were dyed with a crystal violet solution (0.1% crystal violet, 10% EtOH, and 89.9% PBS) and then cell viabilities were measured. The culture medium was removed from each well and then the cells were washed with PBS. 200 μi of the crystal violet solution was added to the cells in each well, which were incubated for 5 minutes at room temperature and then washed with water twice. 1 ml of ethanol was added to each well and then shaken at room temperature for 10 minutes. Then, the absorbance was measured at a wavelength of 570 nm. This experiment was performed three times. An average of three test results and a standard deviation are shown in Table 2 and FIGS. 1-4.

Table 2

4. Test Results

As shown in Tables 1 and 2 and FIGS. 1-4, the extract of Halocynthia sp. shell effectively reduces the melanin pigment formation level without causing cytotoxicity. In addition, the extract of Halocynthia sp. shell showed higher cell viability than PTU used as a positive control in all concentration used. Therefore, the extract of Halocynthia sp. shell can be used, instead of conventional whitening materials having cytotoxicity, such as phenylthiourea.

Formulation 1. Preparation of softening skin lotion

3 parts by weight of glycerin, 2 parts by weight of butylene glycol, 2 parts by weight of propylene glycol, and a preservative were dissolved in 80.9 parts by weight of purified water under stirring. 1 part by weight of polyoxyethylene hardened castor oil 60 was dissolved under heating at 60 ^°C and then a perfume was added thereto. To the mixture, 1 part by weight of the powdered extract of Halocynthia sp. shell prepared in Example 1 , 10 parts by weight of ethanol, 0.1 parts by weight of triethanolamine, and a pigment were added and sufficiently stirred to obtain a softening skin lotion. Formulation 2. Preparation of nourishing lotion

The mixture of 1.7 parts by weight of sitosterol, 1.5 parts by weight of polyglyceryl-2 oleate, 0.7 parts by weight of ceramide, 1.2 parts by weight of ceteareth-4, and 1.5 parts by weight of cholesterol was homogenized at a constant temperature.

To the homogenized mixture, 1 part by weight of the powdered extract of Halocynthia sp. shell prepared in Example 1 , 0.4 parts by weight of dicetylphosphate, 5.0 parts by weight of concentrated glycerin, 10 parts by weight of sunflower oil, 0.2 parts by weight of carboxyvinylpolymer, 0.3 parts by weight of xanthan gum, and a preservative were added and dissolved under heating. 76.5 parts by weight of purified water was added to the mixture, which was then emulsified using a homogenizer. Then, a perfume was added to the mixture, which was mixed by stirring to obtain a nourishing lotion.

Formulation 3. Preparation of nourishing cream The mixture of 4 parts by weight of sitosterol, 3 parts by weight of polyglyceryl-2 oleate, 0.7 parts by weight of ceramide, 2 parts by weight of ceteareth-4, and 3 parts by weight of cholesterol was homogenized at a constant temperature. To the homogenized mixture, 3 parts by weight of the powdered extract of Halocynthia sp. shell prepared in Example 1 , 0.4 parts by weight of dicetylphosphate, 5.0 parts by weight of concentrated glycerin, 22 parts by weight of sunflower oil, 0.5 parts by weight of carboxyvinylpolymer, and a preservative were added and dissolved under heating. 56.4 parts by weight of purified water was added to the mixture, which was then emulsified using a homogenizer. Then, a perfume was added to the mixture, which was mixed by stirring to obtain a nourishing cream.

Claims

1. A whitening cosmetic composition comprising an extract of Halocynthia sp. shell.

2. The composition of claim 1 , wherein the extract of Halocynthia sp. shell is obtained by extracting with an extraction solvent selected from the group consisting of water, CrC₄ alcohol, and a mixed solvent of water and C₁-C₄ alcohol.

3. The composition of claim 2, wherein the extraction solvent is water.

4. The composition of claim 1 , wherein the extract of Halocynthia sp. shell is obtained by sonicating Halocynthia sp. shell in water.

5. The composition of claim 4, wherein the extract of Halocynthia sp. shell is obtained by sonicating Halocynthia sp. shell in water and then drying the resultant extract solution.

6. The composition of claim 1 , wherein the extract of Halocynthia sp. shell is obtained by an extraction process comprising: (a) sonicating Halocynthia sp. shell in water;

(b) adding ethyl acetate to the extract solution obtained in Step (a) to obtain an aqueous fraction; and

(c) adding butanol to the aqueous fraction obtained in Step (b) to obtain an aqueous fraction.

7. The composition of claim 1 , wherein the extract of Halocynthia sp. shell is obtained by an extraction process comprising:

(a¹) sonicating Halocynthia sp. shell in water;

(b¹) drying the extract solution obtained in Step (a¹) to obtain a powder; (c¹) adding water and ethyl acetate to the powder obtained in Step (b¹) to obtain an aqueous fraction; and (cT) adding butanol to the aqueous fraction obtained in Step (c') to obtain an aqueous fraction.

8. The composition of claims 6 or 7, further comprising drying the aqueous fraction after performing the extraction process.

9. The composition of claim 8, wherein the drying is performed by freeze-drying.