CN106860063B

CN106860063B - A cosmetic

Info

Publication number: CN106860063B
Application number: CN201710215761.2A
Authority: CN
Inventors: 金鑫
Original assignee: Guangzhou Zicai Cosmetic Factory
Current assignee: GUANGZHOU ZICAI COSMETIC FACTORY
Priority date: 2017-04-04
Filing date: 2017-04-04
Publication date: 2020-05-19
Anticipated expiration: 2037-04-04
Also published as: CN106860063A

Abstract

The invention discloses a cosmetic which comprises the following components in percentage by mass: 0.01% of polypeptide, 7-10% of lanolin, 5-9% of vaseline, 4-8% of squalane, 6% of glycerol monostearate, 1% of castor oil, 0.1-0.25% of whitening agent, 15-20% of glycerol, 0.1-0.5% of sodium chondroitin sulfate, 1% of anthocyanin and the balance of water. The cosmetic disclosed by the invention can thoroughly dispel sunburn, is rich in nutrition, smooth and oily, has remarkable effects of improving skin tension and resisting wrinkles, and also has the effect of whitening skin.

Description

A cosmetic

Technical Field

The invention relates to a cosmetic, in particular to a cosmetic for removing sunburn.

Background

Sunburn is caused by ultraviolet rays, toxin accumulation, and irritation of cells. The shape of sunburn appears as an oval bulge or is smooth, predominantly dark brown or light brown in color. The easy-to-appear positions include the outer side of the front hip, the back of the hand, the front side of the lower leg and the face, and the early sunburn can feel pain, itch, desquamation and finally form color spots. The distinction of facial sunburn is mainly whether the sunburn appears on the nasal bridge and outside the cheeks, and is mainly a spot with a granular, punctate, tan color.

For example, CN103655387A discloses a plant essence mask for preventing and treating sunburn, which has the following formula (components and weight percentage thereof) that the tail of Chinese angelica is 6.0-12.0; 4.0-10.0 parts of erysipelas; 10.0-15.0 of talcum powder; aloe 6.0-15.0; radix trichosanthis

3.0-6.0; 9.0-30.0 parts of coix seeds; 3.0-10.0 of borneol; 6.0 to 12.0 portions of eupatorium; 3.0-10.0 parts of honey. The cosmetic is prepared by selecting Chinese herbal medicine components with reasonable compatibility and preparing through a special process. However, the facial mask is added with a large amount of Chinese herbal medicine components, so that the result is complex, the preparation is not easy, the cost is high, and the economical efficiency is further improved.

CN 104306861A discloses a sunburn medicine, which is prepared from 4-10 parts of honeysuckle, 2-5 parts of fructus forsythiae, 10-25 parts of fructus momordicae, 63-8 parts of vitamin B, 5-11 parts of radix codonopsis, 6-11 parts of motherwort, 5-10 parts of radish peel, 5-10 parts of watermelon peel, 6-8 parts of cactus, 5-6 parts of mint, 5-12 parts of fructus amomi, 8-12 parts of bran, 2-5 parts of calcium tablets, 12-15 parts of red wine and 5-12 parts of white hyacinth bean. The medicine is rich in vitamins, contains a large amount of effective components for repairing skin collagen, and is beneficial to eliminating sunburn and repairing skin. Can effectively eliminate sunburn and repair the collagen layer of the skin. And the compatibility is healthy and reasonable, and the health requirements of skin and body are met. However, the same medicine uses a large amount of Chinese herbal medicines, has extremely complicated components, remains questionable about the uniformity and stability of the drug effect, and has complicated preparation process and high cost.

CN106265414A discloses a sunburn removing mask, which comprises the following active ingredients in parts by weight: 5-8 parts of aloe extract, 0.5-0.8 part of ginkgo leaf extract, 1-2 parts of bee pupa extract and 0.2-0.4 part of safflower extract, wherein each facial mask carrier carries 15-25ml of the active ingredients. The facial mask disclosed by the invention has a remarkable effect on sunburn. The ginkgo leaf extract and the bee pupa extract used in the invention are expensive substances, the extraction process is complex, the quality control is not easy to realize, and no remarkable effect is achieved in the aspect of effect, so that the invention has great improvement space.

Based on the defects of the prior art, cosmetics for treating sunburn have great demand in the prior art, and therefore, based on the defects, the invention provides the following technical scheme.

Disclosure of Invention

The invention provides a cosmetic which can effectively remove sunburn.

In one aspect of the invention, the cosmetic capable of killing human melanocytes is provided, wherein the cosmetic comprises the polypeptides shown in SEQ ID Nos. 1-11, and the polypeptides are respectively named as RGT-1-11 correspondingly. The polypeptide was obtained by previous screening of the library by the applicants.

The invention also provides a cosmetic, which contains anthocyanin in addition to the polypeptide of any one of SEQ ID Nos. 1-11.

The invention also provides a cosmetic which is a cream and comprises the following components in percentage by mass: 1-11, 0.01% of polypeptide, 7-10% of lanolin, 5-9% of vaseline, 4-8% of squalane, 6% of glycerol monostearate, 1% of castor oil, 0.1-0.25% of whitening agent, 15-20% of glycerol, 0.1-0.5% of chondroitin sodium sulfate, 1% of anthocyanin and the balance of water.

The present invention also provides a method for preparing a suspension of melanocytes of an adult, which comprises first obtaining epidermal tissues of an adult by a bubble adsorption method. In this step, after the skin is sterilized with alcohol, a syringe is placed on the skin, the skin is sucked with a suction force of 200-300 mmHg for 60-120 minutes to form blisters on the skin, and epidermal tissue containing about 86-90% of keratinocytes is cut offAnd 5-7% melanocytes. Then, the cut epidermal tissue is cut up, soaked with a trypsin solution, then transferred into a trypsin-ethylene diamine tetraacetic acid (trypsin-EDTA) solution to separate melanocytes from the epidermal tissue, and finally, the epidermal cells are collected in a centrifugal mode and cultured in a cell culture dish. In which keratinocytes are rapidly reduced due to their inability to grow and melanocytes continue to proliferate, resulting in a population of cells that approximates pure melanocytes. Since melanocytes belong to adherent cells (adherent cells), Trypsin (Trypsin) is used to decompose the attachment protein between cells and culture dishes, so that melanocytes are exfoliated, which is a common method for collecting adherent cells. Subsequently, the obtained melanocytes were washed. Thereafter, at about 1.2-1.5X10⁶At a density of ml, melanocytes were resuspended in Han's medium (Ham's F12). Han's culture medium (Ham's F12) is a common commercial culture medium, and contains essential substances for cell growth, such as amino acids, saccharides, fatty acids, and salts. In addition to the Han's culture solution (Ham's F12), DMEM, Minimal Essential Medium (MEM), phosphate physiological saline (PBS), 1640 culture solution, normal physiological saline (normal saline) or other physiologically compatible solutions can be used for preparation. Finally, the melanocyte suspension cells are counted and equally divided into test tubes for subsequent experiments.

The invention also provides an experimental method for killing human melanocytes, which comprises the following steps of: 1-11, wherein the addition amount of the polypeptide is 0.01% (m/v); the cells were cultured at 37 ℃ for 5 hours, 10 hours and 12 hours, and the number of viable cells was measured and the cell killing rate was analyzed.

Technical effects

The invention provides specific melanocyte killing polypeptide, and specifically kills local melanocytes, thereby providing final guarantee for thoroughly removing sunburn. In addition, the anthocyanin is added into the cosmetics, so that the formed sunburn on the surface of the skin is effectively eliminated, the anthocyanin and the skin are organically combined, and the combination of the whitening agent and other substances is added, so that the temporary solution and the permanent solution are treated, and the sunburn is thoroughly eliminated to achieve the effect of no relapse. The cosmetic disclosed by the invention is rich in nutrition, smooth and oily, has remarkable effects of improving skin tension and resisting wrinkles, and particularly has the effect of whitening the skin. Most importantly, the invention has low cost and easily obtained materials, and is particularly suitable for large-area popularization and use.

Drawings

FIG. 1 is a flow chart of culturing human melanocytes;

FIG. 2 is a comparison of treatment of anterior skin;

figure 3 comparison of the skin of the face after treatment.

Detailed Description

The following examples illustrate aspects of the invention. They are only used for illustration and do not limit the scope of the invention, and the conventional changes or additions made by those skilled in the art are also within the scope of the invention and should be limited.

Example 1 bulk procurement of human melanocytes

First, epidermal tissues of adults were obtained by a blister adsorption method. In this step, after the skin is sterilized with alcohol, a syringe is placed on the skin, the skin is sucked with a suction force of 200-300 mmHg for 100 minutes to form blisters on the skin, and epidermal tissue, which generally contains about 86-90% of keratinocytes and 5-7% of melanocytes, is cut off. Then, the cut epidermal tissue is cut up, soaked in a 5% trypsin solution, then transferred into a trypsin-ethylenediaminetetraacetic acid (trypsin-EDTA) solution to separate melanocytes from the epidermal tissue, and finally, the epidermal cells are collected by centrifugation at 5000g and cultured in a cell culture dish. In which keratinocytes are rapidly reduced due to their inability to grow and melanocytes continue to proliferate, resulting in a population of cells that approximates pure melanocytes. Since melanocytes belong to adherent cells (adherentcells), 3% Trypsin (Trypsin) is used to break down the attachment protein between cells and culture dish, so that melanocytes are exfoliated. Subsequently, the obtained melanocytes were washed with PBS. Thereafter, at about 1.5X10⁶Density of/ml, fining melaninThe cells were resuspended in Han's medium (Ham's F12). Finally, counting the cells of the melanocyte suspension, and subpackaging the cells into test tubes with equal quantity, wherein the density of the cells in each test tube is about 100g/mL, so as to carry out subsequent experiments.

EXAMPLE 2 preparation of the polypeptide

The polypeptide is expressed and purified by means of polypeptide synthesis or by means of prokaryotic expression, which is commonly used by those skilled in the art, to obtain the amino acid sequence of SEQ ID NO:1 to 11, respectively named RGT-1 to 11.

Example 3 melanocyte killing assay

The sequence of SEQ ID NO:1-11, wherein the original cell concentration is 200/mL, and the addition amount of the polypeptide is 0.01% (m/v); the cells were cultured at 37 ℃ for 5 hours, 10 hours and 12 hours, and the number of viable cells was measured, respectively, and the cell killing rate was analyzed (3 replicates each). The results are shown in Table 1 below, with each result having a P value of less than 0.05.

Kind of product	5h viable cell remaining	10h residual amount of viable cells	Residual amount of viable cells for 12h
				PBS control	195	183	170
RGT-1 Polypeptides	22	18	1
				RGT-2 Polypeptides	40	23	2
RGT-3 Polypeptides	43	20	1
				RGT-4 Polypeptides	50	19	1
RGT-5 Polypeptides	39	30	4
				RGT-6 Polypeptides	40	31	2
RGT-7 Polypeptides	32	29	5
				RGT-8 Polypeptides	29	30	4
RGT-9 Polypeptides	55	38	8
				RGT-10 Polypeptides	60	43	8
RGT-11 Polypeptides	46	32	5

TABLE 1

As can be seen from the results in table 1, the polypeptide of the present invention has the characteristics of good effect of killing melanocytes, short killing time, and significant killing effect. Furthermore, cell death caused by the first lysis of the melanocyte nuclei was observed by electron microscope examination.

EXAMPLE 4 preparation of cosmetic

0.01 percent of RGT-1 polypeptide described by SEQ ID No. 1, 10 percent of lanolin, 5 percent of vaseline, 4 percent of squalane, 6 percent of glycerol monostearate, 1 percent of castor oil, 0.25 percent of whitening agent, 20 percent of glycerol, 0.5 percent of chondroitin sodium sulfate and 1 percent of anthocyanin are respectively taken, fully stirred to be completely dissolved, uniformly mixed, the pH value of the solution is adjusted to 6.5, and finally purified water is used for complementing the total amount of the formula, and the finished product cosmetic is obtained after sterilization, filtration and split charging.

Example 5 preparation of cosmetic RGT-2 to 11

Following the method of example 4, SEQ ID NO:2-11 polypeptides are respectively substituted for RGT-1 polypeptide, and the rest components and the preparation method are the same as the example 4, so that the cosmetic is prepared.

Example 6 Security verification

Test materials and methods

Animals: common-grade experimental animal white rabbit. The animals were examined for abnormalities.

Environmental conditions: room temperature: 25 ℃, relative humidity: 68 percent;

the test method comprises the following steps:

according to the acute skin irritation test of the cosmetic hygiene code (2007 edition), 24 hours before the test, the hair on both sides of the spine of the test animal is cut off, and the hair removing range is 3 cm X3 cm. 0.5 g of the test immunogenicity was applied directly to the right skin, covered with clean gauze and fixed with a non-irritating adhesive tape for 2 hours. The other side was a negative control. Skin reaction at the contact site of the test animal was observed and recorded at 1 hour, 24 hours, 48 hours and 72 hours after the removal of the test substance.

And (4) conclusion:

under the test condition, the test substance containing the polypeptide of the invention is graded as non-irritant according to skin irritation intensity in the acute skin irritation test of white rabbits. The control cosmetic was also non-irritating.

Example 7 Spot removal Effect verification

1. Patients were selected by selecting sunburn patients (signed with informed consent and approved by the ethical committee), who were all women and were randomly divided into two groups. 220 cases of treatment with 20 cases of each polypeptide, mean age 25 years, course 1 month, using the polypeptide cosmetics of examples 4 and 5; control group 20, mean age 25 years, course 3 months, were treated with the cosmetic of example 4 containing no polypeptide. The two groups have comparability (P >0.05) in terms of age, sex, severity of disease condition, course of disease, etc.

2. The treatment method comprises the following steps: the treatment groups are cosmetics prepared respectively based on examples 4 and 5, and are respectively applied to the sunburn affected part for 1 time in the morning and evening, and the treatment course is 1 month; in the control group, the cosmetic containing no polypeptide in example 4 was given as a control, and the control was also applied to the affected part of sunburn for 1 time, with a treatment course of 1 month. During the treatment period, all patients stop using all other functional cosmetics and pay attention to sun protection. The patients in the control group need to monitor liver function and blood lipid regularly, and can continue taking the medicine without abnormality. 30d is 1 course of treatment.

3. Determination of therapeutic effect

the scoring method comprises scoring according to skin damage area, wherein the skin damage area is 0 point without skin damage<3cm²Is 1 minute, and the area of skin damage is 3-5cm²Is 2 minutes, area of skin damage>5cm²the total score is the sum of area score and color score, and the index is calculated by (total score before treatment-total score after treatment)/total score before treatment.

The therapeutic effect judgment standard is that the area of the visible color spot is faded by more than 90 percent and the color of the color spot is basically disappeared, and the decline index after treatment is more than or equal to 0.8 according to the calculation of a grading method; the effect is displayed; the area of the visible color spot is faded by more than 60 percent, the color of the color spot is obviously lighter than that before treatment, and the reduction index after treatment is more than or equal to 0.5 according to the calculation of a grading method; effectively, the area of the color spot which can be seen by naked eyes is faded by more than 30 percent, the color of the color spot becomes light, and the decline index after treatment is more than or equal to 0.3 according to the calculation of a grading method; the effect is not good, the area of the color spot visible to naked eyes is reduced by less than 30%, the color of the color spot is not obviously changed, and the reduction index after treatment is less than or equal to 0 according to the calculation of a grading method. The total effective rate is (ten effective cases are basically cured) per total case x 100%.

The statistical method comprises analyzing data with statistical software SPSS15.0, and analyzing the counting data and the measurement data with chi-shaped software²Method of testing and t-testing, P<0.05 has statistical significance.

The curative effect comparison shows that the total effective rate of the treatment group is more than 95 percent, the total effective rate of the control group is 0 percent, and the curative effect comparison has significant difference (X)²<2.21,P<0.05), see table 2.

TABLE 2

As can be seen from Table 2, the invention has better treatment effect and better application prospect. By taking photographs of a patient treated cosmetically with RGT-1 polypeptide, it can be seen from the results in FIGS. 2 and 3 that the better treatment of the plaques after treatment is substantially healed. The remaining polypeptides are similar in effect and are not shown here.

Example 8 cosmetic moisturization and moisturization time measurement

The MMV (moisteuremeasurementvalue) method measures the MMV value reflecting the ability of the stratum corneum to bind water. The hydration of the skin was tested using a model CM825 skin oil water acid base tester. The measuring principle is that the stratum corneum contains a large amount of electrolytes, the electrolytes in water have conductivity, the phenomenon is generated on the surface of the skin, and the hydration state of the skin is reflected by the change of the conductivity after the test probe is contacted with the skin.

The subjects were healthy, non-dermatosis volunteers. The test conditions are that the temperature and the humidity of the environment are constant during the test period, the temperature is 25 +/-2 ℃, and the relative humidity is 40-60%.

by ① the by ① testing by ① method by ① comprises by ① the by ① following by ① steps by ① of by ① determining by ① 4 by ① testing by ① areas by ① of by ① a by ① left by ① forearm by ① and by ① a by ① right by ① forearm by ①, by ① wherein by ① the by ① area by ① of by ① each by ① area by ① is by ① 4 by ① cmX by ① 4 by ① cm by ①, by ① and by ① testing by ① blank by ① values by ① of by ① the by ① areas by ① respectively by ①, by ① coating by ① a by ① sample by ① on by ① the by ① testing by ① areas by ①, by ① wherein by ① the by ① dosage by ① of by ① the by ① sample by ① is by ① 2 by ① mg by ①/by ① cm by ① 2 by ①, by ① carrying by ① out by ① final by ① testing by ① after by ① 120 by ① min by ① of by ① use by ①, by ① testing by ① 5 by ① points by ① in by ① each by ① area by ①, by ① and by ① averaging by ① the by ① evaluation by ① formula by ① of by ① the by ① moisturizing by ① effect by ① evaluation by ①, by ① wherein by ① in by ① a by ① formula by ① of by ① relative by ① moisturizing by ① rate by ①/by ①% by ① (by ① P by ① sample by ① t by ① is by ① X by ① -by ① P by ① blank by ①) by ①/by ① (by ① P by ① label by ① t by ① is by ① X by ① -by ① P by ① blank by ①) by ① X by ① 100 by ①% by ①, by ① P by ① sample by ① t by ① is by ① X by ① -by ① conductivity by ① of by ① a by ① product by ① after by ① a by ① certain by ① time by ①, by ① P by ① -by ① conductivity by ① of by ① an by ① uncoated by ① sample by ①, by ① P by ① label by ① t by ① is by ① X by ① -by ① conductivity by ① of by ① a by ① reference by ① after by ① a by ① certain by ① time by ①, by ① and by ① the by ① moisturizing by ① rate by ① condition by ① results by ① of by ① the by ① cosmetics by ① and by ① glycerin by ① comparison by ① are by ① shown by ① in by ① the by ① following by ① table by ①: by ①

Kind of product	Moisture retention rate
		Control	0.48
RGT-1 Polypeptides	0.78
		RGT-2 Polypeptides	0.75
RGT-3 Polypeptides	0.74
		RGT-4 Polypeptides	0.76
RGT-5 Polypeptides	0.74
		RGT-6 Polypeptides	0.70
RGT-7 Polypeptides	0.80
		RGT-8 Polypeptides	0.76
RGT-9 Polypeptides	0.77
		RGT-10 Polypeptides	0.75
RGT-11 Polypeptides	0.76

TABLE 3

From the above table, the moisturizing rate of the cosmetic of the present invention is significantly higher than that of glycerin, and it is also demonstrated that the polypeptide of the present invention has a significant promoting effect on improving the moisturizing effect of the cosmetic.

The foregoing is a more particular description ー of the present invention, taken in conjunction with the accompanying preferred embodiments, and it is not intended that the invention be limited to the specific embodiments described. For those skilled in the art to which the invention pertains, several simple deductions or substitutions can be made without departing from the spirit of the invention, and all shall be considered as belonging to the protection scope of the invention.

Sequence listing

Jinxin of < 110 >

(120) A cosmetic

〈160〉11

〈210〉1

〈211〉27

〈212〉PRT

Artificial sequence of < 213 >

〈400〉RGT-1

RIWPVMDQRDRQQHHPWYQRAPCRNQG

〈210〉2

〈211〉25

〈212〉PRT

Artificial sequence of < 213 >

〈400〉RGT-2

VCCCRSTHKPQFCLQLMINFKQFAP

〈210〉3

〈211〉24

〈212〉PRT

Artificial sequence of < 213 >

〈400〉RGT-3

IMGHREQFPRIPRNRPKWHAHWWW

〈210〉4

〈211〉26

〈212〉PRT

Artificial sequence of < 213 >

〈400〉RGT-4

STRRWPKPSTPQSYLEVEQEEMYQPR

〈210〉5

〈211〉21

〈212〉PRT

Artificial sequence of < 213 >

〈400〉RGT-5

IQIPTKSSSRTFLSKIPLDMP

〈210〉6

〈211〉20

〈212〉PRT

Artificial sequence of < 213 >

〈400〉RGT-6

NAEQLYDHNIQWLYAMYWVE

〈210〉7

〈211〉23

〈212〉PRT

Artificial sequence of < 213 >

〈400〉RGT-7

QHPQRLKRSRPETHSMMQPKVWY

〈210〉8

〈211〉25

〈212〉PRT

Artificial sequence of < 213 >

〈400〉RGT-8

DYGPRVVFQQWMKSNRWNWHMNDVV

〈210〉9

〈211〉27

〈212〉PRT

Artificial sequence of < 213 >

〈400〉RGT-9

GHQWYSQTRFEYDDLKIWGKCTFHELS

〈210〉10

〈211〉26

〈212〉PRT

Artificial sequence of < 213 >

〈400〉RGT-10

WHGTTMWHVQFRSSFWAAWWMTPRLC

〈210〉11

〈211〉17

〈212〉PRT

Artificial sequence of < 213 >

〈400〉RGT-11

WHTSHRFKRHPEWHLLI

Claims

1. A cosmetic comprises the following components in percentage by mass: 0.01% of polypeptide, 7-10% of lanolin, 5-9% of vaseline, 4-8% of squalane, 6% of glycerol monostearate, 1% of castor oil, 0.1-0.25% of whitening agent, 15-20% of glycerol, 0.1-0.5% of sodium chondroitin sulfate, 1% of anthocyanin and the balance of water, wherein the polypeptide sequence is shown as SEQ ID NO:2 to 11.

2. A polypeptide, characterized by: the sequence is shown in any one of SEQ ID No. 2-11.

3. The use of the polypeptide of claim 2 for the preparation of a cosmetic product having sunburn removing effect.

4. Use according to claim 3, characterized in that: the cosmetic is facial mask or cream.

5. The cosmetic of claim 1, wherein: it has effects of eliminating sunburn.