CN105555278A - Topical composition for skin containing gincenoside Mc - Google Patents

Abstract

The present invention relates to a composition containing, as an active ingredient, gincenoside Mc, for providing effects such as anti-aging, improvement of wrinkles, whitening, improved moisturization, anti-inflammation, improvement of acne, skin troubles, and atopic dermatitis symptoms, skin clearing, and pore tightening, in addition to providing effects such as skin tone improvement, hair growth promotion, improvement of graying hair, anti-dandruff, and antisepsis.

Description

Dermatologic preparation composition containing ginsenoside Mc

Technical field

The present invention relates to a kind of compositions, described compositions contains ginsenoside Mc (ginsenosideMc), thus aging resistance can not only be provided, improve wrinkle of skin, whitening, moisturizing improve effect, but also antiinflammatory can be provided, improve acne and skin problem, and the effect of allergic symptom, convergence skin and pore refining effect, and skin color can be provided to improve effect, promote hair growth, improve poliosis, anti-dandruff and antiseptic effect.

Background technology

Skin, as the first defensive barrier of human body, has and makes intracorporeal organ protect the function of intracorporeal organ from the stimulation of temperature and humidity change and the external environment condition such as ultraviolet, public hazards material.With advancing age, skin will due to multiple inherence, extrinsic factor and changing.That is, from inherent aspect, owing to regulating metabolic various hormone secretion to reduce, and the function of immunocyte and cytoactive reduce, and therefore the immune protein of needed by human body and the biosynthesis of organism constitutive protein reduce.From external aspect, due to the destruction of ozone layer, the ultraviolet content arriving earth's surface from sunray increases, and along with the in-depth of environmental pollution, free radical and active oxygen increase, not only make that the thickness of skin reduces, wrinkle increases, elastic force reduction, and make skin complexion dimmed, often there is problem (trouble) in skin, also can increase nevus and freckle and senile plaque, and cause the various changes such as complexion is deteriorated, the colour of skin is dimmed.

In order to the skin condition change preventing these from occurring due to skin inherence and extrinsic factor, and maintaining healthy skin condition, people make great efforts to be used for improving skin condition by adding in cosmetics to enforcement of going forward side by side such as the biological active substances obtained from existing various animal, plant, microorganism etc. always.

Ginsenoside Mc is that the ginsenoside (Rb1, Rg1, Rd, Rb2, Rc, Rf) of the natural goods form contained in Radix Ginseng is decomposed by digestive enzyme and intestinal microbial and generated.Have and report that the active anticancer of ginsenoside Mc is strong, and record the method used as anticancer preparation by ginsenoside Mc in No. 164266th, Korean granted patent, but do not report and ginsenoside Mc is improved effect as the such as overall skin condition of the compositions of effective ingredient, promote hair growth and improves poliosis, anti-dandruff and the whole structure as skin preparations for extenal use such as anticorrosion.

Summary of the invention

The technical problem solved

To this, the present inventor finds that the ginsenoside Mc contained in Radix Ginseng can not only provide aging resistance, improves wrinkle of skin, whitening and moisturizing improve effect, but also antiinflammatory can be provided, improve acne, the effect of skin problem and allergic symptom, and skin color can be provided to improve effect, promote hair growth, improve poliosis, anti-dandruff and antiseptic effect, thus complete the present invention.

Therefore, the object of the invention is to, provide a kind of Dermatologic preparation composition, described compositions contains ginsenoside Mc, thus the integrality that can demonstrate skin improve and promote hair growth, improve poliosis, anti-dandruff and antiseptic effect.

Technical scheme

To achieve these goals, the invention provides a kind of containing ginsenoside Mc as effective ingredient for aging-resistant Dermatologic preparation composition.

In addition, the invention provides a kind of containing ginsenoside Mc as the Dermatologic preparation composition for improving wrinkle of effective ingredient.

In addition, the invention provides a kind of containing ginsenoside Mc as the Dermatologic preparation composition for moisturizing of effective ingredient.

In addition, the invention provides a kind of containing ginsenoside Mc as the Dermatologic preparation composition for improving acne of effective ingredient.

In addition, the invention provides a kind of containing ginsenoside Mc as the Dermatologic preparation composition for improving color and the colour of skin of effective ingredient.

In addition, the invention provides a kind of containing ginsenoside Mc as the Dermatologic preparation composition for pore refining of effective ingredient.

In addition, the invention provides a kind of containing ginsenoside Mc as the Dermatologic preparation composition for improving allergic skin of effective ingredient.

In addition, the invention provides a kind of containing ginsenoside Mc as the Dermatologic preparation composition for antiinflammatory of effective ingredient.

In addition, the invention provides a kind of containing ginsenoside Mc as the Dermatologic preparation composition for whitening of effective ingredient.

In addition, the invention provides a kind of containing ginsenoside Mc as effective ingredient for trichogenous Dermatologic preparation composition.

In addition, the invention provides a kind of containing ginsenoside Mc as the Dermatologic preparation composition for preventing poliosis of effective ingredient.

In addition, the invention provides a kind of containing ginsenoside Mc as the Dermatologic preparation composition for anti-dandruff of effective ingredient.

In addition, the invention provides a kind of by the natural antiseptic agent compositions of ginsenoside Mc.

Beneficial effect

Compositions of the present invention contains ginsenoside Mc, thus aging resistance can not only be provided, improve wrinkle of skin, whitening, moisturizing improve effect, but also antiinflammatory can be provided, improve the effect of acne and skin problem and allergic symptom, convergence skin and pore refining effect, and skin color can be provided to improve effect, promote hair growth, improve poliosis, anti-dandruff and antiseptic effect.

Preferred forms

Dermatologic preparation composition according to the present invention contains ginsenoside Mc as effective ingredient.

The ginsenoside Mc used in the present invention has the structure of following chemical formula 1:

[chemical formula 1]

Ginsenoside Mc of the present invention can extract from plant, also can be synthesized by method as known in the art and use, also can use the ginsenoside Mc of mercantile-type sale.In addition, ginsenoside Mc can obtain from Radix Ginseng extract.The kind of the Radix Ginseng at this moment used is not particularly limited, and can use water ginseng, Radix Ginseng Rubra, Radix Ginseng, Tai Ji ginseng and tail ginseng etc.Further, described Radix Ginseng extract not only comprises by Radix Ginseng lixiviate, decocts and carry and the leachate that obtains, also comprises and carries out concentrated and concentrate that is that obtain again to a part or whole part of leachate, or concentrated solution described in after drying and impregnating, decoct, the tablet prepared flowing extracts, and be included in Radix Ginseng the chemical substance playing main efficacy results, but also comprise plant itself, and the extract of the whole part of the Radix Ginsengs such as stem, root, leaf, flower and fruit can be used, be not limited to the extract of a certain specific part.In addition, the method extracting ginsenoside Mc from Radix Ginseng extract can use known method.

Particularly, described ginsenoside Mc by method as known in the art, after using water or organic solvent to prepare Radix Ginseng extract, therefrom can be isolated.The organic solvent used in the present invention can be selected from the mixed solvent of ethanol, methanol, butanols, ether, ethyl acetate, chloroform and these organic solvents and water, preferably uses the ethanol of 80%.At this moment, Extracting temperature is preferably 10 ~ 80 DEG C, and can extract 3 ~ 24 hours.If exceed the scope of described Extracting temperature and extraction time, then extraction efficiency can reduce, or the change of composition can occur.

According to compositions of the present invention, in composition total weight, preferably contain the ginsenoside Mc of 0.001 to 50 % by weight.If this is because the content of described ginsenoside Mc is less than 0.001 % by weight, then, if more than 50 % by weight, then can there is the problem in cutaneous safety or dosage form in the effect brought by described composition and low effort.

Compositions of the present invention can use as aging-resistant Dermatologic preparation composition, its improve skin elasticity and improve in wrinkle remarkably productive.

Compositions of the present invention can use as the Dermatologic preparation composition for moisturizing, and it can strengthen skin barrier function, and can the differentiation of induced skin keratinocyte.Therefore, the Dermatologic preparation composition preventing or improve xeroderma, allergic dermatitis, contact dermatitis or the psoriasis etc. that cause because epidermis does not break up completely can be effectively used as.

Compositions of the present invention can use as the Dermatologic preparation composition for improving acne, and its antibacterial effect is excellent, especially excellent to the antibacterial effect of acne pathogenic bacterium, and provides antiphlogistic effects.

Compositions of the present invention can use as the Dermatologic preparation composition for improving color and the colour of skin, when being used in skin, by expansion blood capillary, and nutritional labeling is fed to skin by blood circulation promoting smoothly, and suppress skin aging, therefore, the effect improving color and the colour of skin is remarkable.

Compositions of the present invention can use as pore refining, the Dermatologic preparation composition that regulates sebum and improve skin problem, when being used in skin, suppress the sebum of excessive secretion, and by promoting that the elimination of active oxygen and the synthesis of collagen protein carry out pore refining, and due to the minimizing of inflammatory Cytokines Expression, therefore, suppress the effect of skin problem remarkable.

Compositions of the present invention can use as the compositions for improving allergic skin, it is by means of only suppressing proteolytic enzyme active acceptor-2 (Proteinase-ActivatedReceptor-2 bringing out pruritus, PAR-2) activity, thus excellent antipruritic effect is provided, but also anti-inflammatory effect can be provided by the secretion reducing interleukin (Interleukin-8, IL8).Therefore, described ginsenoside Mc of the present invention can as stabilized susceptibility, zest or allergic skin and scalp, and for the Dermatologic preparation composition that improves or alleviate hotness and inflammation effective ingredient and suitably use.

Compositions of the present invention can use as the compositions for whitening, and it by hindering the activity of tryrosinase, the generation of check melanin, thus can provide excellent whitening effect.

Compositions of the present invention can use as trichogenous Dermatologic preparation composition, it promotes the conversion from resting stage hair cycle to the anagen hair cycle, thus the growth comprising and promote hair is provided, and promote the generation of new hair, and the effect that existing hair health is grown, also provide and prevent and the phenomenon suppressing hair to come off from scalp or hair is scattered or the effect of state that attenuates.

Compositions of the present invention can as preventing from the Dermatologic preparation composition of poliosis to use, it carrys out activation of melanocyte by the expression improving the transcription factor (MITF) in melanocyte, and promote melanic synthesis, thus provide bringing out of advance preventing poliosis, and promote the effect of bringing out hair color.

Compositions of the present invention can as preventing from the Dermatologic preparation composition of the dandruff to use, it purifies scalp by effectively discharging the toxin be accumulated on hair and scalp, and scalp inflammation can be prevented to react by suppressing the propagation of dandruff bacterium and growth, and, because the generation of inhibit activities oxygen and the anti-oxidation efficacy of effect are given prominence to, calm therefore, it is possible to provide and strengthen scalp, and strengthening the effect of intrinsic phylactic power defensive power.

Compositions of the present invention can use as natural antiseptic agent compositions, because it is natural component, therefore while antiseptic effect brilliance, also harmless.

Described compositions of the present invention can containing acceptable carrier on cosmeceutical or Dermatology or base material dosage form.Its whole dosage forms that can use as applicable local provide, and such as, can be provided as solution, gel, solid, the anhydrous product of pasty state in aqueous phase, disperse oil phase and obtain emulsion, suspension, microemulsion, microcapsule, subparticle ball or ion-type (liposome) and nonionic the form of folliculus dispersant, or frost, astringent, lotion, powder, ointment, spray or hide the form of flaw rod.Further, can with foam (foam) form or comprise further compression propellant aerosol combination form use.These compositionss can be prepared according to the method that this area is conventional.

Especially, Dermatologic preparation composition of the present invention is used for preventing the dandruff, for hair growth or for preventing poliosis time, can as the compositions for scalp and hair dosage form, dosage form is not particularly limited, such as, dosage form can turn to hair oil, trichotrophy astringent, scalp care agent, hair-care agent, shampoo, hair conditioner, hair lotion or hair of scalp dual-purpose nursing agent etc.

In addition, fatty material can be comprised according to compositions of the present invention, organic solvent, lytic agent, concentrating agents, gellant, softening agent, antioxidant, suspending agent, stabilizing agent, foaming agent (foamingagent), aromatic, surfactant, water, ion-type or nonionic emulsifier, filler, chelating agen, chelating agent, preservative agent, vitamin, blocker, wetting agent, quintessence oil, dyestuff, pigment, hydrophilic or lipophile activating agent, the adjuvant that lipid folliculus or other composition any etc. being usually used in cosmetics are commonly used in cosmeceutical or Dermatology field.Described adjuvant imports with amount normally used in cosmeceutical or Dermatology field.

In addition, compositions of the present invention, in order to increase skin improvement effects, can contain skin absorption enhancement material.

Embodiment

Below, structure of the present invention and effect will be further illustrated by test example and dosage form example.But, these test examples and dosage form example only in order to help to understand the present invention and illustratively object provide, category of the present invention and scope are not limited to following example.

The preparation of [reference example 1] ginsenoside Mc

The ginsenoside Mc used to test effect of the present composition is purchased from rich (AMBO) institute of peace.

[test example 1] elastase activity suppresses the mensuration of effect

Elastase activity for ginsenoside Mc hinders ability, compares with epigallocatechin gallate (EGCG) (EGCG) and measures.The elastoser used and substrate are the elastoser of commerciality ground purchased from American Sigma-Aldrich (Cat.No.E0127).

By following test method, elastase activity inhibition is tested.

In 96 orifice plates, be modulated in 10mg/L Tri(Hydroxymethyl) Amino Methane Hydrochloride (Tris-HCL) buffer (pH8.0), mixing the ginsenoside Mc of 200 μ L and the 20 μ g/mL elastoser-type III solution of 50 μ L.The EGCG of 250 μMs is used as positive controls, and employs distilled water as the non-process group of negative control group.Afterwards, add the N-succinyl-alanine-Ala-Ala of the 0.4514mg/mL modulated with described buffer of 100 μ L-to p-nitroanilide (N-SUCCINYL-ALA-ALA-ALA-p-NITROANILIDE), and react 15 minutes at 25 DEG C.After reaction terminates, measure absorbance (cooperative effect 2, the microplate reader (BioTek) (VT, the U.S.) under 415nm wavelength.Implement blank assay by identical method and correct.

The computational methods of elastase activity inhibition, as shown in following mathematical expression 1, the results are shown in following table 1.

Mathematical expression 1

Elastase activity obstruction rate (%)=1-(C-D)/(A-B) × 100

A: do not adding substances, and when adding enzyme, the absorbance under 415nm wavelength

B: when not adding substances, enzyme, the absorbance under 415nm wavelength

C: when adding substances, enzyme, the absorbance under 415nm wavelength

D: in interpolation substances, and when not adding enzyme, the absorbance under 415nm wavelength

Table 1

Compound	Suppression degree (%)
		Non-process group	0
EGCG	65
		Ginsenoside Mc	75

As shown in above-mentioned table 1, demonstrate the suppression degree of ginsenoside Mc to elastase activity, significantly more excellent than the EGCG being known as Elastinase activity inhibitor, therefore can confirm that the elastase activity inhibition of ginsenoside Mc of the present invention is excellent.

[test example 2] collagenase (MMP-1) hinders ability

Collagenase for ginsenoside Mc of the present invention generates obstruction ability, compares and measure with tretinoin.

Be equipped with containing 2.5% fetal bovine serum DMEM (Dulbecco'sModifiedEagle'sMedia) culture medium 96 holes slat chain conveyor device (96-wellmicrotiterplate) in, with 5, the amount of 000 cells/well (well) adds human fibroblasts, and at 5%CO ₂, carry out cultivating till growing to 70 ~ 80% degree in 37 DEG C of incubators.After 24 hours, cell culture fluid is taked by the ginsenoside Mc of 10 μ g/ml concentration or retinoic acid treatments.

Utilize commercial available collagenase sensing equipment (AmershamPhamarcia company of the U.S., Catalog#:RPN2610), the collagenase measuring the cell culture fluid taked generates degree.First, in 96-hole flat board (96-wellplate) of uniform application once collagenase antibody, add the cell culture fluid taked, and in temperature chamber, implement antigen-antibody reaction 3 hours.After 3 hours, the secondary collagen antibody being combined with colour developing group is joined in the flat board of 96-hole, and again reacts 15 minutes.After 15 minutes, add 3,3' as colour developing evocating substance, 5,5'-tetramethyl benzidine (3,3', 5,5'-tetramethylbenzidine, Sigma), and at room temperature bring out colour developing 15 minutes, again add 1M sulphuric acid and stop chromogenic reaction, then the color of reactant liquor is in yellow, carries out degree according to what react, and the degree of the yellow demonstrated is different.

Utilize absorptiometer, under 405nm, measure the absorbance in yellow 96-hole flat board, and calculate the synthesis degree of collagenase according to following mathematical expression 2, and the results are shown in following table 2.At this moment, by never with the reaction absorbance of the cell culture fluid taked in the group of compositions-treated as a control group.

Mathematical expression 2

Absorbance × 100 of the absorbance/matched group of collagenase expression degree (%)=mass treatment groups of cells

Table 2

Compound	Expression degree (%)
		Non-process group	100
Tretinoin	75
		Ginsenoside Mc	73

As shown in above-mentioned table 2, compared with can knowing the collagenase expression degree of ginsenoside Mc and being known as the tretinoin of collagenase expression inhibitor, its collagenase expression hinders effect level similar.

By the above results, can confirm that ginsenoside Mc of the present invention has the effect of obstruction matrix metalloproteinase (MMP-1).

[dosage form example 1 and compare dosage form example 1]

According to the composition of following table 3, prepare nourishing cream (unit: % by weight) by the method for routine.

Table 3

[test example 3] skin elasticity improves effect and confirms

In order to confirm the effect improved the skin elasticity of people, utilizing described dosage form example 1 and the dosage form comparing dosage form example 1, and having made following evaluation.

For the healthy women of 20 30 to 40 years old age brackets, two groups are divided into often to organize 10, by dosage form example 1 and compare dosage form example 1 two group nourishing cream be applied in face after 12 weeks with the frequency of 1 time every day, utilize skin elasticity tester (CutometerSEM575, C+K Electronics Co., Ltd. (C+KElectronicCo.), Germany) measure skin elasticity.Its result represents in following table 4.The Δ R8 value of the end value Cutometer (CutometerSEM575) of table 4 is recorded, and wherein R8 value represents the character of skin viscoelasticity (viscoelasticity).

Table 4

Experimental products	Skin elasticity effect
		Dosage form example 1	0.44
Relatively dosage form example 1	0.10

As shown in above-mentioned table 4, the dosage form example 1 containing ginsenoside Mc of the present invention, with smear compare dosage form example 1 group compared with, its skin elasticity improves more.

Therefore, can confirm that the compositions containing ginsenoside Mc of the present invention improves very effective to skin elasticity.

[test example 4] improves the confirmation of wrinkle of skin effect

In order to confirm the effect improving wrinkles of compositions of the present invention to people, make use of described dosage form example 1 and comparing dosage form example 1.

In order to confirm described dosage form example 1 and the effect improving wrinkles comparing dosage form example 1, make following evaluation.For the healthy women of 20 40 years old age brackets, two groups are divided into often to organize 10, by dosage form example 1 and compare dosage form example 1 two group nourishing cream be applied in face after 12 weeks with the frequency of 1 time every day, silicon is utilized to take out replica (replica), and with skinanalysis apparatus (visiometer, SV600, Courage+KhazakaelectronicGmbH, Germany) measure the state of wrinkle, and carry out graphical analysis.Its result represents in following table 5.The result of following table 5 represents in each parameter (parameter) after smearing 12 weeks the meansigma methods after deducting the parameter value before smearing.

Table 5

As shown in above-mentioned table 5, can know that the preparation composition for external use of dosage form example 1 is very excellent to the effect improving wrinkle of skin.

[test example 5] tryrosinase hinders effect

Tryrosinase extracts from mushroom class (Mushroom), employs the tryrosinase of Sigma.First, tyrosine as substrate is dissolved in distilled water the solution making 0.3mg/ml, and this solution is joined after in test tube with the amount of every pipe 10ml, adds the potassium-phosphate buffered solution (0.1mol concentration, pH6.8) of 1.0ml and the distilled water of 0.7ml wherein.

In alcoholic solution of the present invention, mix ginsenoside Mc with 10ppm and prepare sample liquid, the above-mentioned sample liquid of 0.2ml is joined in reactant liquor, then reacting 10 minutes in 37 DEG C of temperature chambers.At this moment, by only add 0.2ml solvent to replace add each sample liquid group as a control group, and ascorbic acid to be used as positive controls.In this reactant liquor, add the tyrosinase solution of the 2500 unit/ml of 0.1ml respectively, and again react 10 minutes in 37 DEG C of temperature chambers.The test tube that this reactant liquor is housed is put into frozen water makes it cool rapidly, thus stopped reaction, and with photoelectricity spectrum analysis instrument, measure the absorbance (synergy 2 under 475nm wavelength, microplate reader (VT, the U.S.), and the results are shown in following table 6.Calculate each tryrosinase by following mathematical expression 3 and hinder effect.

Mathematical expression 3

Tryrosinase obstruction rate (%)=100-(reaction absorbance × 100 of the reaction absorbance/matched group of substances)

Table 6

Substances	Tryrosinase obstruction rate (%)
		Matched group (not adding)	0
Ascorbic acid	52
		Ginsenoside Mc	63

Can know from the result of above-mentioned table 6, ginsenoside Mc according to the present invention is high more a lot of than the ascorbic acid as known tyrosinase inhibitor to the suppression ratio of tryrosinase, and therefore whitening effect is very excellent.

[test example 6] utilizes the melanin of B16/F10 melanoma cell to generate inhibition

Amount respectively using 0.001 % by weight is contained the test portion of ginsenoside Mc and kojic acid as substances, and add in the culture fluid of B16/F10 melanoma cell (bank of Korea Cell system) with finite concentration, and after cultivating 3 days, remove culture fluid, then use phosphate buffer (PBS) to clean, and with after 1NNaOH dissolved cell, under 405nm, measure absorbance (synergy 2, microplate reader (VT, the U.S.).To the cell of substances do not added as a control group, and compare with the melanin content in matched group, and measure the degree that each substances hinders melanin generation.Calculate melanin generating suppression according to mathematical expression 4, and the results are shown in table 7.

Mathematical expression 4

Melanin generating suppression (%)=100-(absorbance × 100 of the absorbance/matched group of substances)

Table 7

Substances	Melanin generating suppression (%)
		Matched group (not adding)	0
Kojic acid	53
		Ginsenoside Mc	66

Can know from the result of above-mentioned table 7, the suppression ratio that ginsenoside Mc according to the present invention generates melanin is high more a lot of than the kojic acid generating Inhibitors as known melanin, and therefore whitening effect is very excellent.

[test example 7] skin moisture-keeping power increases effect measuring

Skin moisture-keeping power being increased to the effect produced to measure ginsenoside Mc, make use of described dosage form example 1 and comparing dosage form example 1, and having made following evaluation.

For the adult men and women being categorized as dry skin of 20 40 to 50 years old age brackets, be divided into two groups often to organize 10, by dosage form example 1 and compare dosage form example 1 two group nourishing cream be applied in face 4 weeks with the frequency of 2 times every day.Before smearing beginning, to smear after 1 week, after 2 weeks, 4 weeks afterwards time, and smear after 2 weeks (altogether through 6 weeks) in stopping, constant temperature, constant humidity condition (24 DEG C, relative humidity 40%) under, use moisture of skin tester (CorneometerCM825, C+K Electronics Co., Ltd. (C+KElectronicCo.), Germany) measure skin beauty water component.Its result represents in following table 8.The result of table 8 be the value of the moisture of skin tester measured before on-test as benchmark, by the result that the increase part of measured value after process a period of time represents with percentage rate.

Table 8

Can confirm from the result of described table 8, smear when comparing dosage form example 1, until through 4 weeks that smearing, demonstrate the moisture increment rate of about 30%, but stop smearing rear skin beauty water component to reduce, on the contrary, when smearing the dosage form example 1 containing ginsenoside Mc, stop smear after also major part demonstrate the moisture of skin increment rate of more than 30%.The skin moisture-keeping power excellent effect of the compositions of the present invention containing ginsenoside Mc can be known thus.

[test example 8] Keratinocyte differentiation facilitation effect measures

As follows, in order to understand the facilitation effect of ginsenoside Mc Human Keratinocytes differentiation, the hornification coating (CornifiedEnvelope, CE) generated when utilizing absorbance to measure Keratinocyte differentiation is measured.

First, by after being separated from the epidermis of baby and the keratinocyte of the people once cultivated puts into cultivation flask, after making it be attached to bottom, with after the ginsenoside Mc process of the concentration of 5ppm in culture fluid, cultivate 5 until cell grows to 70 ~ 80% of bottom area.Now, using low calcium (0.03mM) processed group and high calcium (1.2mM) processed group as negative control group and positive controls.Then above-mentioned cultured cells is obtained, after phosphate buffered saline (PBS) washing, what add 1ml contains 2% sodium lauryl sulphate (sodiumdodecylsulfate, and the dithiothreitol, DTT (Dithiothreitol of 20mM concentration SDS), Tri(Hydroxymethyl) Amino Methane Hydrochloride buffer (the Tris-HCl of 10mM concentration DTT), pH7.4), and carry out supersound process (sonication), boil (boiling), centrifugalize, again precipitation is suspended in 1mlPBS, and the absorbance (synergy 2 under being determined at 340nm, microplate reader (VT, the U.S.).In addition, take out the solution after a part of described supersound process, measure protein content, it can be used as the benchmark evaluating cell differentiation.And the results are shown in following table 9.

Table 9

Substances	Differentiation capability (%) in keratinocyte
		Low calcium (0.03mM) solution (negative control group)	100
High calcium (1.2mM) solution (positive controls)	210
		Ginsenoside Mc	285

As shown in Table 9 above, when can confirm with ginsenoside's Mc process, the differentiation facilitation effect of keratinocyte is excellent.

[test example 9] skin barrier function recovery effects measures

In order to measure the effect that ginsenoside Mc recovers the skin barrier function impaired due to skin injury to produce, implement following experiment.For the upper arm of 10 adult men and women, by the method for tape stripping (TapeStripping), injured skin barrier, smear the dosage form example 2 prepared according to the composition of following table 10 respectively and compare dosage form example 2 two groups, with Vapometer (dolphin (Delfin), Finland), measure the recovery extent of 1 percutaneous moisture loss amount (TWEL) every day, measure seven days, and compare.Here comparison dosage form example 2 is the carrier (vehicle) of negative control group.Its experimental result is shown in following table 11.The result of table 11 is the results difference before treatment before barrier injury and after barrier injury compared as 100% benchmark.

Table 10

Food ingredient	Dosage form example 2	Relatively dosage form example 2
			Purified Water	69	70
Propylene glycol	30	30
			Ginsenoside Mc	1	-

Table 11

Known from above-mentioned table 11, can confirm when using the comparison dosage form example 2 not containing ginsenoside Mc to process, As time goes on, percutaneous moisture loss amount increases gradually, on the contrary, when using the dosage form example 2 containing ginsenoside Mc to process, percutaneous moisture loss amount recovers normal with very fast speed, and barrier injury is restored.

[test example 10] color improves effect

In order to evaluate cosmetic composition according to the present invention to the effect promoting cutaneous circulation, utilize laser Doppler perfusion imaging instrument (LaserDopplerPerfusionImager, LDPI; PeriscanPIMII, lark prestige (Perimed) (Stockholm, Sweden)), measure blood circulation degree in skin.Known LDPI measures the sanguimotor instrument in skin, and it is now widely used instrument, be a kind of speed and amount that not only can measure the blood in capillary of skin, and the very delicate of the flowing in small artery and venule can be measured.

In thermostatic constant wet chamber, after washing one's face with fancy soap, adapt to 30 minutes, and utilize LDPI to determine initial value.First, with LDPI, the initial stage blood flow below the forehead of ice-cold 30 women of trick is at ordinary times measured.Then, make subjects use 1 week described dosage form example 1 and compare dosage form example 1 to measure blood flow afterwards, the result (dermal blood flow change) then the blood flow of mensuration and described initial stage measured value compared represents in following table 12.

Table 12

Can confirm from the result of above-mentioned table 12, cosmetic composition according to the present invention, compared with the comparison dosage form example 1 not containing ginsenoside Mc, is made dermal blood flow significantly increase, and is promoted by this blood circulation and complexion is improved.This finally shows can to effectively transmitting skin supplement ingredient, and suppress also delay skin aging to contribute according to the cosmetic composition containing ginsenoside Mc of the present invention.

[test example 11] colour of skin improves effect

In order to the colour of skin understood described dosage form example 1 and compare dosage form example 1 improves effect, 30 tested objects are made to use (evening 1 times/day respectively, smear 1 week altogether) after, FacialStageDM-3 (jasmine spy (Moritex), Japan) the instrument evaluation colour of skin is utilized to improve degree.For colour of skin improvement rate, adopt the lightness of skin and color measured value, and the lightness of skin and color change value judge, and the results are shown in following table 13.Lightness and the higher expression colour of skin of color change value improve.

Table 13

Can confirm from the result of table 13, not containing the comparison dosage form example 1 of with good grounds ginsenoside Mc of the present invention, not demonstrate the significant colour of skin and improve effect, on the contrary, use containing the dosage form example 1 of ginsenoside Mc as effective ingredient, the colour of skin after using, compared with before use, is greatly improved.

[test example 12] pore refining effect

1. by promoting the effect of the biosynthetic pore refining of collagen protein

Ginsenoside Mc according to the present invention is compared the biosynthetic facilitation effect of collagen protein and transforming growth factor-β (TGF-β) and measures.First, by fibroblast with every hole 10 ⁵the amount of individual cell is inoculated in 24 holes (well), and carries out cultivating until growing to till about 90%.Used after serum-free cell culture medium (DMEM) cultivates 24 hours, used that 10ng/ml's be dissolved in ginsenoside Mc of the present invention in serum-free medium and TGF-β process respectively, and at CO ₂cultivate 24 hours in incubator.Take out their supernatant, and utilize procollagen type (I) enzyme-linked immunosorbent assay (ELISA) test kit (procollagen type (I) (procollagentype (I); Whether #MK101, TAKARA (Shiga, Japan) observe the increase and decrease of procollagen (procollagen), and the results are shown in following table 14.Non-process group is set to 100 and synthesis capability for collagen protein contrasts.

Table 14

Substances	Collagen protein synthesis ability (%)
		Non-process group	100
TGF-β	183.5
		Ginsenoside Mc	195.7

Can confirm from the result of above-mentioned table 14, ginsenoside Mc according to the present invention, compared with positive controls TGF-β, demonstrates the collagen protein synthesis ability of higher levels of excellence.Therefore, can confirm that ginsenoside Mc according to the present invention shrinks by the collagen protein growing amount increasing pore periphery the pore broadened.

2. pore refining effect

In order to understand dosage form example 1 and compare the pore refining effect of dosage form example 1, evaluate as follows.Selecting men and women's tested object that 20 pore sizes are wide, being divided into two groups by often organizing 10, and according to group every day smearing dosage form example 1 on the face and comparing the nourishing cream of dosage form example 1, smear 4 weeks altogether.Judge the effect of pore refining in such a way.Photo before shooting experiment and after 4 weeks, and allow expert be evaluated by naked eyes.Its result represents (opinion rating: 0-does not shrink completely in following table 15; 5-shrinks a lot).

Table 15

Substances	Opinion rating
		Dosage form example 1	4 11 -->
Relatively dosage form example 1	0

Known from the result of above-mentioned table 15, compare the effect that dosage form example 1 does not have pore refining, but dosage form example 1 illustrates the pore refining effect that can with the naked eye confirm, thus known ginsenoside Mc of the present invention is to the excellent effect of pore refining size.

[test example 13] sebum secretion inhibition

1. by suppressing the effect of the suppression skin supersecretion of 5α-reductase activity

In order to confirm the active inhibition of 5α-reductase, determine in HEK293-5 α R2 cell [ ¹⁴c] testosterone change into [ ¹⁴c] ratio of dihydrotestosterone (DHT:dihydrotestosterone).On HEK293 cell after transfection p3xFLAG-CMV-5 α R2, and by every hole 2.5 × 10 ⁵the amount of individual cell is inoculated in 24 orifice plates, and carries out cultivating (Parketal., 2003, JDS.Vol.31, pp.191-98).Within second day, change the new culture medium being added with zymolyte and inhibitor into.By 0.05 μ Ci [ ¹⁴c] testosterone (test kit (AmershamPharmaciabiotech), Britain) is as culture medium substrate.

In order to confirm the active suppression degree of 5α-reductase, add ginsenoside Mc, and at 37 DEG C, 5%CO ₂cultivate 2 hours in incubator.Now, the group not adding ginsenoside Mc is used as negative control group, the group that will add finasteride (finasteride) is used as positive controls.Reclaim culture medium afterwards, and after extracting steroid by 800 μ l ethyl acetate, be separated the organic solvent layer on top, and after drying, to remaining residue, use 50 μ l acetic acid ethyl dissolutions again, and on silastometer thin film silica gel 60F254 (Silicaplasticsheetkieselgel60F254), use ethylacetate-hexane (1:1) to launch as solvent.

By plastics test portion after being dried in air, shower system is employed in order to measure isotopic amount the sheet plastic of drying and x-ray thin film are together joined bath box in, within 1 week, measure the isotopic mass of testosterone and the dihydrotestosterone stayed on thin film afterwards, then according to following mathematical expression 5 and 6, calculate conversion ratio and obstruction rate respectively, and the results are shown in following table 16.

Mathematical expression 5

Radiant in conversion ratio (%)=DHT region/total radiant × 100

Mathematical expression 6

Conversion ratio × 100 of suppression ratio (%)=[conversion ratio of the conversion ratio-substances of matched group]/matched group

Table 16

Substances	Conversion ratio (%)	Obstruction rate (%)
			Negative control group	48.0	-
Positive controls	27.6	42.5
			Ginsenoside Mc	14.8	67

Can confirm from above-mentioned table 16 result, ginsenoside Mc can suppress the activity of 5α-reductase effectively, thus blocking-up testosterone is converted into dihydrotestosterone, and show inhibition more excellent compared with the finasteride of known suppression 5α-reductase activity.Described 5α-reductase makes testosterone be converted into dihydrotestosterone, thus is combined with intracytoplasmic receptor protein and enters in core, thus activation sebocyte cell promote differentiation, thus make the sebum excessive secretion in sebaceous gland.Therefore, the too much secretion of ginsenoside Mc by effectively suppressing the activity of 5α-reductase to suppress sebum is confirmed.

2. sebum secretion inhibition

In order to understand described dosage form example 1 and compare the sebum secretion inhibition of dosage form example 1, carry out following evaluation.Select 30 and think men and women's tested object that sebum secretion is many, smear dosage form example 1 in the appointed part of skin on the face and compare the nourishing cream of dosage form example 1 every day, smear 4 weeks altogether.Sebum is reduced to the judgement of effect, by using sebum tester (SebumeterSM810, C+K Electronics Co., Ltd. (C+KElectronicCo.), Germany) measure the average sebum slip (%) after 2 weeks and 4 weeks respectively, and the results are shown in following table 17.

Table 17

Known from the result of above-mentioned table 17, of the present invention containing ginsenoside Mc as effective ingredient dosage form example 1 with not containing compared with its comparison dosage form example 1, effectively can suppress the sebum of too much secretion.

[dosage form example 3 and compare dosage form example 3 ~ 4]

Prepare dosage form example 3 according to the composition shown in following table 18 and content (% by weight) and compare dosage form example 3 ~ 4, being described as follows.Dosage form example 3 is the material of mixing ginsenoside Mc, relatively dosage form example 3 is do not wrap the material of the effective ingredient improved containing acne skin completely, comparative example 4 is as the standard substance of the benchmark for antimicrbial power, and this material contains many erythromycin (erythromycin) used as acne therapeutic agent.

Dosage form example 3 and to compare the preparation method of dosage form example 3 ~ 4 as follows.The composition of the A item of following table 18 is dissolved completely, and in other dissolving tank, dissolves the composition of B item completely, afterwards B item is added in A item, make it mix solubilized.And wherein, to add the composition of C item according to the mixed proportion recorded in table 18, and after mix homogeneously, filter, thus prepare this compositions.

Table 18

[test example 14] is tested the antimicrbial power of acne bacterium

Use each cosmetic composition prepared according to described dosage form example 3 and the composition that compares dosage form example 3 ~ 4, the propionibacterium acnes (ATCC6919: culture medium-brain heart infusion broth culture medium (BHIbroth)) to as acne pathogenic strain) carry out antimicrbial power test.

As follows to the antimicrbial power test method of acne bacterium.

(1) preparation of test organisms liquid

Use the culture fluid being inoculated in by propionibacterium acnes and carrying out Anaerobic culturel in brain heart infusion broth culture medium as test organisms liquid.

(2) preparation of dilute solution

In the brain heart infusion broth culture medium (pH6.8) or LB broth bouillon (pH4.5) of 15ml, add the described test organisms liquid of 0.15ml, and the mixed liquor mixed is used as dilute solution.

(3) preparation of test portion

By by dosage form example 3 and the cosmetic composition stock solution comparing preparation in dosage form example 3 ~ 4, directly use as sample.

(4) antimicrbial power test

1) add test portion in No. 1, cell culture tube (96wellplate) row in 96 holes, make to mate with initial concentration, and add dilute solution and make total amount be 200 μ l.

2) by after No. 1 mixed liquor mix homogeneously arranged, take out the mixed liquor of 100 μ l and join in No. 2 rows, and after mix homogeneously, again take out the mixed liquor of 100 μ l and join in No. 3 rows.Carry out secondary dilution (doubledilution) in this way.

3) at 32 DEG C, by suspension degree, quiescent culture, after 24 hours and 48 hours, judges whether bacterium rises in value, and the Cmin of the propagation not having bacterium is decided to be minimal inhibitory concentration (MinimumInhibitoryConcentration, MIC) value.If mixed liquor is opaque and be difficult to judge whether bacterium rises in value, then by confirming with microscopic examination.

The antimicrbial power result of the test of acne bacterium is represented in following table 19.For minimal inhibitory concentration, be converted into the concentration of the effective ingredient contained in dosage form and labelling.

Table 19

Project	pH	Propionibacterium acnes
			Dosage form example 3	5.7	>52ppm
Relatively dosage form example 3	5.7	Maximum concentration (not having antimicrbial power)
			Relatively dosage form example 4	5.7	>100ppm

In the result of table 19, in minimal inhibitory concentration, ppm concentration is less, can think that this material is to the effective material of the antimicrbial power of acne bacterium, when using dosage form example 3, with be used as known acne therapeutic agent erythromycin comparison dosage form example 4 compared with, the ppm concentration demonstrated is significantly low, thus can confirm that the compositions containing ginsenoside Mc has more excellent antimicrbial power to test organisms.

[test example 15] lipid synthesis (Lipogenesis) inhibition test

Using the 3T3-L1 cell of the fibroblast (fibroblastcellline) as mice, with 1 × 10 ⁵cells/well is attached at the fetal bovine serum (fetalBovineSerum filled containing 10%, FBS) DMEM (Dulbecco ' smodifiedeagle ' smedium, GIBCOBRL, life technology company) culture medium 6 well culture plates (cultureplate) in.After 2 days, DMEM (FBS containing the 10%) culture medium again more renewed, and cultivate 2 days.Then, with the DMEM (FBS containing 10%) containing 1 μ g/ml insulin (insulin), 0.5mM isobutyl methylxanthine (IBMX) and 0.25 μm of dexamethasone (dexamethasone), induction is carried out to described cultured cells, and with the ginsenoside Mc of 50 μMs and caffeine process, then, after 2 days, be again replaced with the DMEM comprising insulin, and cultivate 5 days.After 5 days, be again replaced with normal incubation medium (DMEM, the FBS containing 10%), and described cell observed and is cultured to described cell and be changing into adipose cell from form.

In order to evaluate ginsenoside Mc to the effect suppressing Fat Accumulation in adipose cell, described in utilization, completing the 3T3-L1 adipose cell of differentiation, implementing the Sudan three and dyeing (S4136, Sigma-Aldrich).At normal temperatures, in phosphate buffer, after 4% paraformaldehyde (pH7.2) fixed fat cell, wash with phosphate buffer, then, take pictures after dyeing with the Sudan three, and compared by naked eyes.To substances do not added or compare material and only use the group of culture medium to use as a control group, for other comparable group, with the caffeine process of 50 μMs.Fat Accumulation suppresses degree to be by the degree of dyeing being divided into +++, ++ ,+,-, thus give grade, and now, more close +++, represent that dye levels is larger.Its result represents in following table 20.

Table 20

Sample

Suppression ratio %

Matched group	+++
		Comparable group	+
Ginsenoside Mc	-

As shown in above-mentioned table 20, can confirm the ginsenoside Mc used in the present invention, not only in adipose cell, the fat mass of accumulation is few, and compared with the caffeine as known lipid synthesis inhibiting substances, also has excellent lipid synthesis inhibition.Therefore, by suppressing lipid synthesis to reduce sebum, thus the generation of acne can be suppressed.

[test example 16] acne is improved and sebum secretion reduces and test with presence or absence of stimulation

Using 30 people suffering from acne as subjects, be divided into three groups often to organize 10, and use by described dosage form example 3 to the subjects corresponding to each group and compare cosmetic composition 1 month prepared by dosage form example 3 ~ 4.Improve standard for acne, be set as 1 point to 5 points, and labelling 1 is divided into " not having ", 3 are divided into " common ", and 5 are divided into " very good ".For experimental result, be marked in following table 21 with the average mark of 10.

Eliminated for acne, to recognize the natural law of elimination as benchmark, for acne recurrence, with or without recurring as benchmark after 1 month period.For the minimizing of sebum secretion, be set as 1 point to 5 points, and be marked as 1 and be divided into " not having ", 3 are divided into " common ", and 5 are divided into " very good ".For experimental result, be marked at following table 21 with the average mark of 10.With (demonstrating the number of irritant reaction)/(overall test number) judge skin irritant with or without.

Table 21

As shown in above-mentioned table 21, can know dosage form example 3 and compare compared with dosage form example 3, acne does not recur, and improves acne on the whole and have excellent effect.In addition, when using the comparison dosage form example 4 containing antimicrbial power standard substance, although demonstrate acne to improve effect, but it is strong to the stimulation of skin when using described material, therefore life-time service is not suitable for, but compositions according to the present invention does not but stimulate, therefore demonstrate and be also applicable to long-time use.

[test example 17] inflammation is improved effect-interleukin 8 (IL-8) and is generated inhibition

Before one day that carries out testing, by Keratoderma epithelial cell (Normalhumanskinkeratinocyte, NHEK buy place: Lonza), with 5 × 10 ⁴cells/well, is inoculated in 96 orifice plates, then at 37 DEG C, and 5%CO ₂cultivate 24 hours in incubator (incubator).After 24 hours, clean cell 2 times with phosphate buffer, and be replaced with serum-free keratinocyte basal culture medium (serumfreekeratinocytebasementmedia (KBM)).In each hole, respectively with the ginsenoside Mc process of 5ppm, 25ppm and 50ppm concentration, and after reacting 30 minutes, use aureus peptide polysaccharide (PGSA) (10 μ g/ml), aureus peptide polysaccharide (50 μ g/ml) and aureus peptide polysaccharide (50 μ g/ml)+lipopolysaccharide (1 μ g/ml) to process respectively.Wherein, aureus peptide polysaccharide (PGSA:peptidoglycanfromS.aureus) is as the Peptidoglycan extracted from staphylococcus (peptidoglycan), be the main constituent of the cell wall of Gram-positive (+) bacterium, and the cell membrane component of known antibacterial can bring out inflammation.Report, especially staphylococcus, allergic dermatitis patient about 90% because of this bacterium produce superinfection.The main constituent of the cell membrane that known lipopolysaccharide (LPS:lypopolysaccaride) is Gram-negative (-) bacterium, and be the main cause that inflammation is brought out.

At 37 DEG C, 5%CO ₂cultivate after 24 hours in incubator, take out culture fluid and the elisa (ELISA) carried out interleukin 8 (Interleukin-8, IL-8), and the results are shown in following table 22.For ELISA, make use of the experimental technique of preparation company (BD science).

Table 22

Classification	The secretion (pg/ml) of interleukin 8
		Non-processor matched group (contrast)	935.12
PGSA(10μg/ml)	4812.60
		PGSA(50μg/ml)	5895.08
PGSA(50μg/ml)+LPS(1μg/ml)	6814.91
		Ginsenoside Mc (5ppm)	1573.32
Ginsenoside Mc (25ppm)	1203.54
		Ginsenoside Mc (50ppm)	1001.23

Can confirm from above-mentioned table 22, ginsenoside Mc can significantly reduce and suppress the secretion of the interleukin 8 increased because of PGSA and lipopolysaccharide LPS.Therefore, can know that Dermatologic preparation composition of the present invention significantly can reduce the secretion of the interleukin 8 increased because of PGSA and LPS, thus excellent anti-inflammatory effect can be provided.

[test example 18] pruritus alleviates evaluation

Before one day that carries out testing, by keratinocyte (cell line title: HaCaT buys place: American Type Culture Collection (ATCC)), with 4 × 10 ⁴cells/well, is inoculated in 96 orifice plates, then at 37 DEG C, and 5%CO ₂cultivate 24 hours in incubator.After 24 hours, with the balanced salt solution (Hanks ' BalancedSaltsolution of hanks, HBSS), after buffer solution for cleaning (washing) 96 orifice plate 2 times, reaction buffer (2 μMs of Fluo-4-AM, 20% pluronic acid (pluronicacid), 2.5mM probenecid (probenecid)) is put in cell.At 37 DEG C, 5%CO ₂react 30 minutes in incubator, and react at normal temperatures after 30 minutes, by HBSS buffer solution for cleaning 2 times, and with the ginsenoside Mc process cell of 0.05%, 0.1%, 0.5% and 1.0% concentration (% by weight).

React after 10 minutes, the trypsin trypsin with 2U/ml) or the PAR-2 bioactive peptide (SLIGKV) of 5 μMs process, and measure Ca in cell ²⁺concentration change 80 seconds.For Ca in cell ²⁺the mensuration of concentration change, make use of microplate reader 3 (FlexStation3: molecular device (MolecularDevice), the U.S.).Trypsin trypsin with ginsenoside Mc and 2U/ml) or 5 μMs PAR-2 bioactive peptide (SLIGKV) process after, measure bending (flex) of 80 seconds, and after obtaining the minima of the value obtained and the difference of maximum, the difference of this value with minima when processing with the PAR-2 bioactive peptide (SLIGKV) of 2U/ml trypsin or 5 μMs and maximum is compared, intracellular suppression ratio (%) is poured in calcium ion and represents in following table 23.

Table 23

Can know from above-mentioned table 23, the calcium ion caused because of trypsin or PAR-2 bioactive peptide (SLIGKV) pours in intracellular, reduce along with the process of ginsenoside Mc, and can confirm the concentration along with improving ginsenoside Mc, calcium ion pours in remarkable minimizing to intracellular.

Therefore, the Dermatologic preparation composition containing ginsenoside Mc of the present invention, by effectively suppressing the PAR-2 bringing out pruritus active, thus can provide excellent antipruritic effect.

[dosage form example 4 and compare dosage form example 5]

Shampoo is prepared with the composition of following table 24.Particularly, surfactant and glycol distearate are added in Purified Water, and be heated to 80 DEG C and after making its uniform dissolution, under agitation gradually be cooled to 40 DEG C, and, add in described mixture according to effective ingredient of the present invention and antiseptic, viscosity modifier, spice and hair conditioner and after mixing, be under agitation cooled to room temperature, thus prepare shampoo.

Table 24

Composition	Dosage form example 4	Relatively dosage form example 5
			Ammonium lauryl sulfate	10	10
Polyoxyethylene lauryl base ammonium sulfate	5	5
			Cocamido propyl betaine	2	2
Glycol distearate	1.5	1.5
			Cocomonoethanolamide	0.8	0.8
Ginsenoside Mc	5.0	-
			Polyquaternium-10	0.2	0.2
Blue No. 1	0.0002	0.0002
			Yellow No. 4	0.0001	0.0001
Methyl parahydroxybenzoate	0.1	0.1
			Spice	0.8	0.8
Citric acid	0.1	0.1
			Dimethicone	1.0	1.0
Water	To 100	To 100

[test example 19] dandruff reduces effect test

The male of 19 to 35 years old that selected 24 dandruff is more, is divided into two groups often to organize 12, and the shampoo using dosage form example 4 with the following methods respectively and compare dosage form example 5 is after 1 month, measures dandruff slip.

Before on-test, usual conventional shampoo cleaning hair, then gather the accumulation dandruff of two days after hair washing, and by the weight of the dandruff of collection with washed a hair with dosage form example 4 and the shampoo that compares dosage form example 5 every two days respectively and the weight of the dandruff of accumulating two days after off-test compares and evaluates.Now, by the dandruff of accumulation with vacuum suction apparatus directly from scalp collection, and obtain dandruff slip according to following mathematical expression 7, and the results are shown in following table 25.

Mathematical expression 7

Dandruff slip (%)=(dandruff weight (mg) of the dandruff weight (mg) before on-test after-one month) dandruff weight (mg) × 100 before/on-test

Table 25

Can know from above-mentioned table 26, the dosage form example 4 containing ginsenoside Mc demonstrates excellent dandruff preventing effectiveness.

The test of [test example 20] pruritus capitis preventing effectiveness

Selected 24 men and women of 25 years old to 45 years old more seriously feeling scalp itch, two groups are divided into often to organize 12, and with three days each dosage form examples 4 of frequency usage once and the shampoo after two weeks that compares dosage form example 5, by following metewand, pruritus capitis preventing effectiveness is evaluated, and the results are shown in following table 26.

[metewand]

Very excellent-5 points

Excellent-4 points

Generally-3 points

Bad-2 points

Very bad-1 point

Table 26

Classification	Dosage form example 4	Relatively dosage form example 5
			The removal effect of pruritus capitis	4.2	2.3

Can know from above-mentioned table 26, the preventing of the dosage form example 4 pairs of pruritus capitis containing ginsenoside Mc demonstrates more excellent effect.

[test example 21] potassium ion channel activity increases effect assessment

Minoxidil as alopeciaing therapeutic agent is known as potential mitochondrion K ~+Channel Opener (K _aTPchannelopener), be the representative drugs used in the treatment of androgenetic alopecia.In order to evaluate the mechanism of this minoxidil, employ following test method(s).Described test method(s) is use the prevention K in the fibroblast of composition scalp corium _aTPthe tolbutamide (SIGMAAIDRICH, T0891) of passage processes, thus the propagation of T suppression cell, and again open potassium-channel and recover cell proliferation.

In order to evaluate the K as this compositions _aTPthe function of channel opener, present invention uses the mouse embryo fibroblasts system (Mouseembryonicfibroblastcellline, NIH3T3 :) as fibroblast.This cell is with 3T3 scheme (protocol), the cell line obtained carrying out nature immortalization from the middle fibroblast be separated of NIH Swiss mouse embryo (Swissmouseembryo).Described cell line, in the DMEM (GibcoBRL, Gaithersburg, MD, the U.S.) containing 10%FBS, at maintenance 5%CO ₂, cultivate 24 hours in the incubator of 37 DEG C.NIH3T3 is inoculated in 96 orifice plates, and cultivate in the incubator of 37 DEG C after 24 hours, process with the tolbutamide of 2.5mM, and after 10 minutes, be used separately as the ginsenoside Mc process of the minoxidil of 10 μMs for positive controls and 2.5ppm, 5ppm and 10ppm concentration, and after drug treating after 48 hours, use WST-1 test kit (Roche (Roche)) to measure ability of cell proliferation.Result represents in following table 27.

Table 27

Classification	Ability of cell proliferation (%)
		Non-processor matched group (contrast)	100
Minoxidil	132
		Ginsenoside Mc (2.5ppm)	115
Ginsenoside Mc (5ppm)	123
		Ginsenoside Mc (10ppm)	130

Can know from above-mentioned table 27, during with ginsenoside Mc process, fibroblastic propagation is restored, and ability of cell proliferation depends on the concentration of the ginsenoside Mc of process and increases, and when can confirm with 10ppm ginsenoside's Mc process, cell proliferation returns to by level during minoxidil process.

The melanin of [test example 22] ginsenoside Mc generates facilitation effect test

Add in the serum-free cell freezing media (RPMI culture medium) fetal bovine serum of 5%, the benzylpenicillin of 100IU and 0.2 μM p-phthalic acid (TPA) culture medium in, by melanocyte, with 50,000 cells/well, is inoculated in 24 orifice plates.Second day, to the cell of inoculation, with the ginsenoside Mc process as substances of the ultimate density of 10ppm or 50ppm.Using by the group of DMSO process of 0.1% as negative control group, above-mentioned each group, as positive controls, is cultivated three days by the group processed by the isobutyl methylxanthine (IBMX) with 100 μMs at 37 DEG C of temperature.After cultivation, with phosphate buffer (PBS) cleaning orifice, and add the 1NNaOH of 100 μ l in every hole after, the melanin in dissolved cell.Utilize slat chain conveyor analyzer (microplatereader), measure under 405nm by melanic absorbance (cooperative effect 2, the microplate reader (VT, the U.S.) of dissolving.The result that the melanin of ginsenoside Mc generation facilitation effect and matched group compare is represented in following table 28.

Table 28

Test portion	B16 cell amount (%)
		DMSO(0.1％)	100 19 -->
IBMX(100μM)	120
		Ginsenoside Mc (10ppm)	112
Ginsenoside Mc (50ppm)	121

Can know from above-mentioned table 28, ginsenoside Mc promotes melanocytic B16 cell, thus increases melanic generation, therefore, it is possible to demonstrate excellent melanin to generate facilitation effect.

The promotion transcription factor (MITF) of [test example 23] ginsenoside Mc in melanocyte and the effect of tryrosinase expression

Utilize 501mel cell line, with 500,000 cells/well, to be inoculated in 6 orifice plates, and in each hole, the dimethyl sulfoxide (DMSO) with 0.1% process as negative control group, with the IBMX process of 100 μMs as positive controls, and with the life saponin Mc process of 10ppm as experimental group, and to cultivate after 24 hours, 48 hours and 72 hours at 37 DEG C of temperature, obtain protein.For the protein so obtained, MITF and tryrosinase is utilized to carry out the test of Western blot (WesternBlot) method.Protein Extraction and Western blot are implemented by the normally used standard method of those skilled in the art.After implementing Western blot, the negative control group in its result is set to 100, and compares with this value and represent in following table 29.

Table 29

As shown in above-mentioned table 29, can confirm that ginsenoside Mc improves the expression of MITF and tyrosinase protein matter in melanocyte.

The antimicrbial power evaluation of [test example 24] ginsenoside Mc

In order to evaluate the antimicrbial power of ginsenoside Mc, implement antibacterial experiment.Concrete test method is as described below.

The staphylococcus aureus (Staphylococcusaureus) used in experiment, escherichia coli (Escherichiacoli) and Pseudomonas aeruginosa (Pseudomonasaeruginosa) cultivate in trypticase soya broth culture medium (TrypticSoyBroth); Candida albicans (Candidaalbicans) and aspergillus niger (Aspergillusniger) cultivate in Sabouraud dextrose broth bouillon (SabouraudDextroseBroth).The diluent that culture fluid carries out diluting with 1/100 (albicans strain is using 1/10) in each culture medium is used as test organisms liquid.For aspergillus niger, will 2 × 10 be prepared into ⁸the spore suspension of cfu/ml is as test organisms liquid.

The test organisms liquid of 0.15ml will be added and the mixed liquor mixed uses as dilute solution in each culture medium of 15ml.

In No. 1,96 orifice plate row, add the ginsenoside Mc of 10ppm with the amount of every hole 16 μ l respectively, and add the dilute solution of 184 μ l respectively.The dilute solution of 100 μ l is added respectively in other hole.After No. 1 mixed liquor mix homogeneously arranged, the mixed liquor taking out 100 μ l joins in No. 2 rows, and after mix homogeneously, the mixed liquor again taking out 100 μ l joins in No. 3 rows.Carry out doubling dilution in this way.

Staphylococcus aureus, escherichia coli and Pseudomonas aeruginosa cultivate in the temperature chamber of 32 DEG C; Candida albicans and aspergillus niger cultivate in the temperature chamber of 25 DEG C.

After 48 hours, whether confirm the increment of bacterium with suspensibility and microscope, thus determine minimal inhibitory concentration (MIC) value, and the results are shown in following table 30.

Table 30

As shown in above-mentioned table 30, can confirm that ginsenoside Mc demonstrates antimicrbial power to multiple bacterial strain, and can predict that ginsenoside Mc can work as natural antiseptic agent or antibacterial in compositions by these.

Claims (22)

1. a Dermatologic preparation composition, is characterized in that, described Dermatologic preparation composition comprises ginsenoside Mc as effective ingredient.

2. Dermatologic preparation composition as claimed in claim 1, it is characterized in that, the following chemical formula 1 of described ginsenoside Mc represents:

3. Dermatologic preparation composition as claimed in claim 1 or 2, is characterized in that, described Dermatologic preparation composition contains the described ginsenoside Mc counting 0.001 ~ 50 % by weight with composition total weight.

4. Dermatologic preparation composition as claimed in claim 1 or 2, is characterized in that, described compositions is used for aging resistance.

5. Dermatologic preparation composition as claimed in claim 1 or 2, it is characterized in that, described compositions is for improving skin elasticity.

6. Dermatologic preparation composition as claimed in claim 1 or 2, it is characterized in that, described compositions is for improving wrinkle of skin.

7. Dermatologic preparation composition as claimed in claim 1 or 2, is characterized in that, described compositions is used for moisturizing.

8. Dermatologic preparation composition as claimed in claim 1 or 2, it is characterized in that, described compositions is for strengthening skin barrier function.

9. Dermatologic preparation composition as claimed in claim 1 or 2, is characterized in that, described compositions is used for the differentiation of induced skin keratinocyte.

10. Dermatologic preparation composition as claimed in claim 1 or 2, it is characterized in that, described compositions is for improving acne.

11. Dermatologic preparation compositions as claimed in claim 1 or 2, is characterized in that, described compositions is used for antibacterial.

12. Dermatologic preparation compositions as claimed in claim 1 or 2, is characterized in that, described compositions is used for antiinflammatory.

13. Dermatologic preparation compositions as claimed in claim 1 or 2, it is characterized in that, described compositions is for improving allergic skin.

14. Dermatologic preparation compositions as claimed in claim 1 or 2, it is characterized in that, described compositions is for improving color and the colour of skin.

15. Dermatologic preparation compositions as claimed in claim 1 or 2, is characterized in that, described compositions is used for pore refining.

16. Dermatologic preparation compositions as claimed in claim 1 or 2, it is characterized in that, described compositions is for regulating sebum.

17. Dermatologic preparation compositions as claimed in claim 1 or 2, it is characterized in that, described compositions is for improving skin problem.

18. Dermatologic preparation compositions as claimed in claim 1 or 2, is characterized in that, described compositions is used for whitening.

19. Dermatologic preparation compositions as claimed in claim 1 or 2, it is characterized in that, described compositions is for promoting the growth of hair.

20. Dermatologic preparation compositions as claimed in claim 1 or 2, it is characterized in that, described compositions is used for preventing poliosis.

21. Dermatologic preparation compositions as claimed in claim 1 or 2, is characterized in that, described compositions is used for anti-dandruff.

22. Dermatologic preparation compositions as claimed in claim 1 or 2, is characterized in that, described compositions is used for natural antiseptic agent.