CN107951765B - Purple bamboo shoot tea extract and application thereof - Google Patents

Abstract

The invention relates to the field of cosmetics, in particular to application of a purple bamboo shoot tea extract in cosmetics and a cosmetic composition containing the purple bamboo shoot tea extract. The first purpose of the invention is to provide a purple bamboo shoot tea extract, the second purpose of the invention is to provide the application of the purple bamboo shoot tea extract in preparing cosmetics, and the third purpose of the invention is to provide a cosmetic composition. The extract of the present invention has excellent skin beautifying effect, especially in skin whitening, and can inhibit the activity of tyrosinase in vitro and reduce or lighten melanin contained in melanocyte. Meanwhile, the composition can resist the invasion of free radicals to skin cells; promoting the proliferation of human fibroblasts and delaying the skin aging.

Description

Purple bamboo shoot tea extract and application thereof

Technical Field

The invention relates to the field of cosmetics, in particular to application of a purple bamboo shoot tea extract in cosmetics and a cosmetic composition containing the purple bamboo shoot tea extract.

Background

The main influencing factor of skin color is melanin in the skin, and the shade of skin color is proportional to the content of melanin and the content of melanin transferred to keratinocytes. Changes in melanin content are caused by ultraviolet radiation, changes in female hormones and progestagens, fine particle pollutants in the atmosphere, inflammatory factors, mental stress, decline in metabolic capacity due to aging, and the like. An increase in melanin content causes a darkening of the overall skin color and the appearance of color spots.

In the development of whitening cosmetics, various whitening active raw materials are often added for whitening skin color or improving abnormal pigmentation of skin, and the principle is as follows:

1. anti-oxidation (also called reduction whitening) is a key step for whitening because oxidation radicals participate in partial reactions during the production of melanin.

2. Inhibit "three enzymes one". Four key substances playing a key role in the process of melanin formation, and inhibiting the activity of the four substances blocks the melanin formation.

3. The exfoliating (metabolic whitening) action is to allow melanin already produced to the skin surface and then metabolize them away.

With the growing concern of consumers on their own health and safety in recent years, raw materials derived from natural sources and having excellent effects are preferred over many biochemically synthesized cosmetic efficacy raw materials.

The Gushu purple bamboo tea is produced in Gushu mountain in Changxing county of Zhejiang province, also named as 'Huzhou purple bamboo tea', and is a famous top-quality tribute tea in China. From Tang dynasty, Gushushan has been the emperor 'tea garden'. Slaughter Li Jifu, written in the legend of Yuan and county: after the year of' Zhenyuan, the purple bamboo shoot tea is served for the mountain every year, the work is thirty thousand, and the month-old method is finished. The full condition of Tang Dynasty tribute tea is explained. The tea made of the tea leaves has the name because the fresh tea buds are purplish and the tender leaves are coiled like bamboo shells. The Gushu purple bamboo shoots are produced in Gushu mountain areas in Changxing county in lake of Zhejiang province. The tea is exquisite in tea making process, tender in tea buds, purple in color and like bamboo shoots, so that the purple bamboo shoot tea is obtained. More than 80% of tea containing purple bamboo shoots in the tea retaining mountain areas grows in rotten stone spaces or grave soil on two sides of mountain stream over hundreds of meters above the sea level. The tea garden in flat land cultivated artificially in Jie is small in proportion. Wild ancient tea mountains preserved for more than one thousand years are managed by a traditional method: after the tea is picked in spring, shrubs (only a small part of the shrubs are high and are used for shading the sun) and weeds around the tea trees are removed in late autumn, and the shrubs and the weeds are piled at the roots of the tea trees and are used as fertilizers.

The tea made from the tea leaves of Gushu purple bamboo is a famous tea, and is rich in polyphenols and various minerals. The substances can effectively play a role in health care, such as fat burning, weight losing, body building and beauty treatment. And has effects of refreshing brain, improving thinking, relieving fatigue, resisting bacteria, reducing blood lipid and blood pressure, quenching thirst, removing summer heat, removing toxic substance, promoting urination, tonifying heart, relieving spasm, and preventing dental caries.

Disclosure of Invention

The first purpose of the invention is to provide a purple bamboo shoot tea extract, the second purpose of the invention is to provide the application of the purple bamboo shoot tea extract in preparing cosmetics, and the third purpose of the invention is to provide a cosmetic composition. The extract of the present invention has excellent skin beautifying effect, especially in skin whitening, and can inhibit the activity of tyrosinase in vitro and reduce or lighten melanin contained in melanocyte. Meanwhile, the composition can resist the invasion of free radicals to skin cells; promoting the proliferation of human fibroblasts and delaying the skin aging.

In order to achieve the first object, the invention adopts the following technical scheme:

the Zisun tea extract is a liquid or solid component of Zisun tea obtained by water extraction, organic solvent extraction or supercritical CO2 extraction method.

Preferably, the organic solvent is one or more of ethanol, butanediol and acetone.

In order to achieve the second object, the invention adopts the following technical scheme:

application of the extract of the purple bamboo shoot tea in preparing cosmetics with one, two or three effects of resisting aging and oxidation and inhibiting melanin synthesis and transfer.

The application of the extract of the tea of the purple bamboo shoot in preparing cosmetics for promoting the proliferation of HaCaT cells and HSF cells, resisting oxidation activity, inhibiting key receptors in synthesis and transport pathways of melanin or promoting the expression of hyaluronic acid synthetase 1(HAS 1).

Preferably, the key receptors in the synthesis and transport pathways of melanin include Tyrosinase (TYP), tyrosine-related protein 2(TRP-2), melanocyte melanocortin receptor 1(MC1R), and protease-activated receptor 2 (PAR-2).

In order to achieve the third object, the present invention adopts the following technical solutions:

a cosmetic composition comprises a bamboo shoot tea extract and auxiliary materials acceptable in the field of cosmetics.

Preferably, the cosmetically acceptable adjuvant is selected from one or more of solvents, solubilizers, preservatives, antioxidants, pH regulators, penetration enhancers, liposomes, moisturizers, thickeners, chelating agents, skin feel modifiers, surfactants, emulsifiers, fragrances, and pigments, and other efficacy additives.

Preferably, the composition is in the form of a cream, lotion, solution, film, aerosol or spray.

Preferably, the composition is in the form of a facial cleanser, body wash, lotion, gel, lotion, cream, essence, eye cream, mask, aerosol or spray.

Preferably, the extract of the purple bamboo shoot tea accounts for 0.001-100%, preferably 0.01-50%, more preferably 0.5-20% of the total weight of the composition.

The cosmetic composition may be formulated, for example, as a solution, a suspension, an emulsion, a paste, a gel, a cream, a lotion, a powder, a soap, a surfactant-containing cleanser, an oil, a powder foundation, an emulsion foundation, a wax foundation, a spray, etc., but is not limited thereto. More preferably, it can be made into basic cosmetics such as skin softening lotion, nourishing lotion, emulsion, body lotion, nourishing cream, massage cream, moisturizing cream, hand cream, essence, eye cream, cleansing foam, cleansing lotion, pack, gel, patch, oil-in-water (O/W) emulsion, water-in-oil (O/W) emulsion, etc., color cosmetics such as lipstick, pre-makeup emulsion, foundation, etc., cleansing agent such as hair lotion, hair conditioner, bath lotion, toothpaste, mouth wash, etc., or hair care cosmetics such as hair tonic, hair setting agent such as gel or mousse, hair growth promoter, hair dye, etc.

The cosmetic composition may comprise a cosmetically acceptable medium or base, and may provide any form of topical administration, such as a solution, gel, anhydrous solid or paste, oil-in-water emulsion, suspension, microemulsion, microcapsule, microparticle or ionic (liposome) type and/or nonionic vesicle dispersion, cream, lotion, emulsion, powder, ointment, spray or concealer stick. These compositions can be prepared according to methods commonly used in the art.

When the dosage form of the present invention is a solution or emulsion, a solvent, a dissolving agent or an emulsifier may be used as a carrier. For example, water, ethanol, isopropanol, ethyl carbonate, ethyl acetate, benzyl alcohol, benzyl benzoate, propylene glycol, 1, 3-butylene glycol, glycerol fatty acid esters, polyethylene glycol, or fatty acid esters of sorbitan may be used.

When the dosage form of the present invention is a suspension, liquid diluents such as water, ethanol or propylene glycol, suspending agents such as ethoxylated isostearyl alcohols, polyoxyethylene sorbitol esters and polyoxyethylene sorbitol esters, microcrystalline cellulose, meta-aluminate, bentonite, agar or tragacanth may be used as carriers.

When the formulation of the present invention is a paste, cream or gel, animal oil, vegetable oil, wax, paraffin, starch, tragacanth, cellulose derivative, polyethylene glycol, silicone, bentonite, silica, talc, zinc oxide, or the like can be used as a carrier.

When the formulation of the present invention is a powder or a spray, lactose, talc, silicon dioxide, aluminum hydroxide, calcium silicate or polyamide powder may be used as a carrier. In particular, when the dosage form is a spray, it may further comprise a propellant, such as chlorofluorocarbon (chlorofluorohydrocarbon), propane/butane or dimethyl ether.

When the formulation of the present invention is surfactant-containing, fatty alcohol sulfate, fatty alcohol ether sulfate, sulfosuccinic acid monoester, isethionate, imidazoline derivative, methyl taurine, sarcosinate, fatty acid amide ether sulfate, alkylamide betaine, fatty alcohol, fatty acid glyceride, fatty acid diethanolamide, vegetable oil, lanolin derivative, ethoxylated glycerin fatty acid ester, or the like can be used as a carrier.

The cosmetic composition of the present invention may further comprise a thickener. The thickener contained in the cosmetic composition of the present invention may be methylcellulose, carboxymethylcellulose, carboxymethyl-hydroxy-guanine, hydroxymethyl cellulose, hydroxyethyl cellulose, carbopol, polyquaternary ammonium salt, cetostearyl alcohol, stearic acid, carrageenan (carrageenan), or the like. Preferably, one or more thickeners selected from the group consisting of carboxymethyl cellulose, carbopol, and polyquaternary ammonium salts may be used. More preferably, carbopol may be used.

In an exemplary embodiment of the present invention, the cosmetic composition may include various bases and additives as appropriate, and the kind and content thereof may be easily determined by those skilled in the art. The cosmetic composition may contain acceptable additives such as preservatives, colorants, additives and the like commonly used in the art. Preferably, phenoxyethanol, 1, 2-hexanediol, etc. can be used as the preservative, and a synthetic perfume can be used.

Further, the cosmetic composition of the present invention may comprise a substance selected from the group consisting of water-soluble vitamins, oil-soluble vitamins, polypeptides, polysaccharides, sphingolipids and seaweed extracts. In addition, it may further comprise oils, fats, moisturizers, emollients, surfactants, organic or inorganic pigments, organic powders, ultraviolet absorbers, preservatives, bactericides, antioxidants, plant extracts, pH adjusters, alcohols, colorants, fragrances, blood circulation stimulants, coolants, antiperspirants, purified water, and the like.

However, the ingredients contained in the cosmetic composition may not be limited thereto. Moreover, the content of the components may be determined within a range not affecting the object and effect of the present invention.

The invention also discloses a skin care cream containing the purple bamboo shoot tea, which comprises the following components in percentage by weight:

the invention also discloses a skin care emulsion containing the purple bamboo shoot tea, which comprises the following components in percentage by weight:

the invention also discloses a purple bamboo shoot tea toning lotion which comprises the following components in percentage by weight:

the invention also discloses a purple bamboo shoot tea facial cleanser which comprises the following components in percentage by weight:

the extract of the present invention has excellent skin beautifying effect, especially in skin whitening, and can inhibit the activity of tyrosinase in vitro and reduce or lighten melanin contained in melanocyte. Meanwhile, the composition can resist the invasion of free radicals to skin cells; promoting the proliferation of human fibroblasts and delaying the skin aging.

The invention takes human keratinocyte (HaCaT cell), human fibroblast (HSF cell) and human melanoma cell (A375 cell) cultured in vitro as research objects, and researches the corresponding in vitro efficacy of the tea of the purple bamboo shoot by the experimental methods of MTT, L-DOPA oxidation method, DPPH free radical scavenging experiment, cell-based E L ISA, western blot, cell immunohistochemistry and the like, and the result shows that the extract of the tea of the purple bamboo shoot is in the proper concentration range (0.2, 0.4, 0.6, 0.8 and 1.0g L)^-1) The tea HAS the effects of remarkably promoting the proliferation of HaCaT cells and HSF cells and simultaneously HAS extremely strong antioxidant activity, and the tea of the purple bamboo shoots remarkably inhibits key receptors in the synthesis and transport pathways of melanin, including Tyrosinase (TYP), tyrosine-related protein 2(TRP-2), melanocyte melanocortin receptor 1(MC1R) and protease activation receptor 2(PAR-2), and the tea of the purple bamboo shoots can promote the expression of hyaluronic acid synthetase 1(HAS 1). The combination of the above results shows that the purple bamboo shoot tea has good effects of resisting aging and oxidation and inhibiting melanin synthesis and transfer in vitro level.

Drawings

FIG. 1 shows that the purple bamboo shoot tea promotes HaCaT cell proliferation experiment. Results are expressed as mean ± SD; p <0.01vsblank, p <0.05vs blank, n-6.

FIG. 2 shows HSF cell proliferation promotion experiments with Zisun bamboo tea. Results are expressed as mean ± SD; p <0.01vs blank, p <0.05vs blank, N ═ 6.

FIG. 3DPPH radical clearance. Results are expressed as mean ± SD; n is 2.

FIG. 4 rate of change of tyrosinase activity. N is 1.

FIG. 5Cell-based elisa tests the effect of tea shoots on the expression of the A375 Cell receptor MC 1R. Results are expressed as mean ± SD; p <0.05vs blank, p <0.01vs blank, N-5.

FIG. 6A: and detecting the influence of the tea of the bamboo shoot on the expression of the receptor protein MC1R of the A375 cells by Western blot. B: and (5) carrying out grey level analysis on protein bands. Results are expressed as mean ± SD; p <0.05vs blank, p <0.01vs blank, N-3.

FIG. 7Cell-based elisa assay of the effect of tea of Zisun bamboo on the expression of TRP-2, a375 cellular receptor protein. Results are expressed as mean ± SD; p <0.05vs blank, p <0.01vs blank, N-5.

FIG. 8Cell-based elisa assay of the effect of tea shoots on the expression of the keratinocyte receptor PAR-2. Results are expressed as mean ± SD; p <0.05vs blank, p <0.01vs blank, N-5.

FIG. 9Cell Immunohistochemical assay of the effect of Phyllostachys nigra on PAR-2 expression in HaCaT cells (A) blank set, (B) Phyllostachys nigra 0.2g L^-10.4g of purple bamboo shoot tea L^-1(D) 0.6g of purple bamboo shoot tea L^-1(E) 0.8g of purple bamboo shoot tea L^-1(F) purple bamboo shoot tea 1.0g L^-1. According to the scoring principle, (A) is strongly positive (+++), (B) is strongly positive (+++), (C) and (D) are positive (++), and (E) and (F) are weakly positive (+).

FIG. 10cell-based elisa tests the effect of tea of Zisun bamboo on the expression of HSF cell receptor HAS 1. Results are expressed as mean ± SD; p <0.05vs blank, N ═ 5.

Detailed Description

The extract of Zisun tea described in examples 1-4 below was produced from Changxing Zhejiang.

Example 1 preparation of skin cream

The skin-care cream comprises the following components in percentage by weight:

the preparation method of the skin cream comprises the following steps of adding glycerol, 1, 3-propylene glycol, sodium polyacrylate and water into a water phase pot, heating and stirring to 75-80 ℃, uniformly dispersing to obtain a phase A, adding methyl glucose sesquistearate, PEG-20 methyl glucose sesquistearate, cetearyl alcohol, isohexadecane, shea butter, polydimethylsiloxane, octyldodecanol myristate, cetearyl ethyl hexanoate and tocopherol into the oil pot, heating and stirring to 75-80 ℃, stirring to completely dissolve to obtain a phase B, pumping the phase A into a homogenization reaction pot, pumping the phase B into the homogenization reaction pot, homogenizing for 5 minutes, adding SIMU L GE 35305, homogenizing at a high speed for 3 minutes, starting circulating cooling water, stirring and cooling, adding a bamboo shoot purple tea extract, an essence and a preservative when the temperature is 40-45 ℃, uniformly stirring, stopping stirring after the temperature is not higher than 38 ℃, and finishing stirring.

Example 2 preparation of skin care emulsions

The skin care emulsion comprises the following components in percentage by weight:

the preparation method of the skin care emulsion comprises the following steps: adding water, 1, 3-propylene glycol, butanediol, an acryloyl dimethyl ammonium taurate/VP copolymer and potassium cetyl phosphate into a water phase pot, heating and stirring to 75-80 ℃, and uniformly stirring and dispersing to obtain a phase A; adding behenyl alcohol, squalane, shea butter, polydimethylsiloxane, caprylic/capric triglyceride and mineral oil into an oil pan, heating and stirring to 75-80 ℃, and stirring until the mixture is completely dissolved to serve as a phase B; pumping the phase A into a homogenizing reaction kettle, starting stirring, pumping the phase B, homogenizing for 5 minutes, starting cooling circulating water, and stirring for cooling; adding the purple bamboo shoot tea extract, essence and preservative when the temperature is 40-45 ℃, and stirring and dissolving completely; and (4) continuing stirring until the material temperature is not higher than 38 ℃, stopping stirring, and ending.

Example 3 preparation of astringent

The toning lotion comprises the following components in percentage by weight:

the preparation method of the toning lotion comprises the following steps: adding water, 1, 3-propylene glycol, dipropylene glycol, xanthan gum and sodium polyacrylate into a reaction pot, starting stirring, heating to 75-80 ℃, and uniformly dispersing; cooling circulating water is started, stirring is carried out, the temperature is reduced, when the temperature is reduced to 40-45 ℃, the purple bamboo shoot tea extract and the preservative are added into a reaction pot, and stirring and dissolving are carried out completely; and (5) continuously stirring and cooling until the material temperature is lower than 38 ℃, stopping stirring, and ending.

EXAMPLE 4 preparation of facial cleanser

The facial cleanser comprises the following components in percentage by weight:

the preparation method of the facial cleanser comprises the following steps: adding water, potassium laureth phosphate, glycerol and PEG-120 methyl glucose dioleate into a reaction kettle, starting stirring, heating to 75-80 ℃, and uniformly stirring and dispersing; cooling circulating water is started, the temperature is reduced to 65-70 ℃, acrylic acid (ester) copolymer, sodium cocoyl amphoacetate, citric acid, lauryl hydroxy sulfobetaine and PEG-7 glycerol cocoate are added, and the mixture is uniformly dispersed; when the temperature is reduced to 40-45 ℃, adding the purple bamboo shoot tea extract, essence and preservative, and stirring and dispersing uniformly; and (5) continuing stirring and cooling to below 38 ℃, stopping stirring and ending.

Test examples

Because the purple bamboo shoot tea has unique geographical conditions and is not found in relevant efficacy research documents, the research carries out relevant in-vitro efficacy research on the purple bamboo shoot tea and observes the efficacy effect of the purple bamboo shoot tea on skin at an in-vitro level as an active ingredient.

1 experimental part

1.1 Primary reagents and instruments

Purple bamboo shoot tea extract, oshi romance biology ltd; DMEM medium, Thermo scientific; fetal bovine serum, Thermo scientific; a375 cells, shanghai kanlang biotechnology limited; HSF cells, shanghai kanlang biotechnology limited; HaCaT cells, shanghai medaka cell bank; thiazole blue (MTT), Sigma company; PAR-2 rabbit anti-human antibody, BIOSS Biotech Ltd; TRP-2 Rabbit anti-human antibody, BIOSS Biotech Co., Ltd; MCIR rabbit anti-human antibodies, BIOSS biotechnology limited; goat anti-rabbit IgG antibody, bioscience, ltd; SABC immunohistochemical kit, BOSTER biotechnology ltd; multiskan GO full-wavelength microplate reader, Thermo scientific; CKX53 inverted imaging microscope, olympus (china) ltd; 5200Multi full-automatic chemiluminescence/fluorescence image analysis System, Shanghai Nature technologies, Inc.

1.2 Experimental methods

1.2.1 in vitro cell culture

The culture of the HaCaT cells, the A375 cells and the HSF cells uses a DMEM medium containing 10% fetal calf serum, the frozen cells are recovered, and the cells are cultured to 3-4 generations to a logarithmic growth phase for carrying out experiments.

1.2.2 cell proliferation assay

Using cells grown in DMEM culture containing 10% fetal calf serum for passage 3-4, HaCaT cells or HSF cells grown nearly confluent were cultured at 1 × 10⁵The cells/well density was uniform and seeded in 96-well culture plates. After culturing for 24h, dividing the cells into a blank group and a sample group for culturing, adding the appropriate concentration of the bamboo shoot tea extract into the sample group, adding MTT after culturing for 24h, culturing for 4h, dissolving with DMSO, and determining the OD value.

1.2.3DPPH free radical scavenging experiment

Formulation restriction 1mmol L^-1Mixing DPPH anhydrous ethanol solution with appropriate concentration of the extract of the purple bamboo shoot tea, standing in dark and closed condition for 30min, and measuring OD value.

1.2.4 tyrosinase inhibition experiments

The cells grown in DMEM medium containing 10% fetal calf serum for 3-4 passages were used to grow nearly confluent A375 cells at 1 × 10⁶Uniformly inoculating the cells/holes in a 24-hole culture plate, culturing for 24h, then dividing the cells into a blank group and a sample group for culturing after the cells adhere to the wall, adding a proper concentration of the purple bamboo shoot tea extract into the sample group, culturing for 24h, and then performing a tyrosinase inhibition experiment by using an L-DOPA oxidation method.

1.2.5Cell-based E L ISA experiment

The cells growing near to fusion are cultured at 1 × 10 by using the cells growing in DMEM culture solution containing 10% fetal bovine serum for passage 3-4⁵Uniformly inoculating the cells/holes in a 96-hole culture plate, culturing for 24h, then dividing the cells into a blank group and a sample group for culturing, adding a proper concentration of the purple bamboo shoot tea extract into the sample group, and performing E L ISA detection after culturing for 24 h.

1.2.6Western blot experiment

The cells grown in DMEM medium containing 10% fetal calf serum for 3-4 passages were used to grow nearly confluent A375 cells at 1 × 10⁶The cells/well density was uniform and seeded in 96-well culture plates. After culturing for 24h, dividing the cells into a blank group and a sample group for culturing, and adding the appropriate concentration of the bamboo shoot tea extract into the sample group. And culturing for 24h to extract total cell protein, and performing western blot detection.

1.2.7 cell immunohistochemical experiments

The cells grown in DMEM medium containing 10% fetal calf serum for 3-4 passages were used to grow nearly confluent A375 cells at 1 × 10⁶Uniformly inoculating the cells/well in 96-well culture plate, culturing for 24 hr, separating the cells into blank group and sample group, and adding appropriate concentration (0.2, 0.4, 0.6, 0.8, 1.0g L)^-1) And (4) extracting the purple bamboo shoot tea. After 24h of culture, a celllimmonohistochemical experiment was performed.

1.2.7 statistical analysis

And (3) performing statistical analysis by adopting corresponding statistical software, wherein data is expressed by x +/-s, and performing t test or one-way variance analysis, wherein the difference is statistically significant when P <0.05 is used.

2 results and discussion

2.1 influence of Zisun tea on HaCaT cell growth

The sample groups were selected to appropriate concentrations (0.2, 0.4, 0.6, 0.8, 1.0g L)^-1) The experimental results are shown in figure 1, and the results are expressed as mean value +/-SD., and when the concentration of the purple bamboo shoot tea is 0.2-1.0g x L^-1The keratinocyte growth is remarkably promoted in the range of 0.2-0.6g L^-1Has obvious proliferation effect at concentration and has statistical significance.

2.2 influence of Zisun tea on HSF cell growth

Selecting appropriate concentration (0.2, 0.4, 0.6, 0.8, 1.0g L)^-1) The purple bamboo shoot tea extract directly acts on the HSF cell level, the cell growth condition is observed, the experimental result is shown in figure 2, the concentration of the purple bamboo shoot tea extract is found to be 0.2-1.0 g/L^-1Within the range, HSF cell growth is significantly promoted, especially at concentrations of 0.6-0.8g L^-1Has obvious proliferation effect at concentration and has statistical significance. Therefore, the purple bamboo shoot tea can obviously promote the proliferation of fibroblasts under a certain range of concentration and has a certain anti-aging effect.

2.3DPPH radical scavenging experiment

The skin aging and the body aging have important relation with free radicals, so that the detection of the scavenging effect of the purple bamboo shoot tea on the free radicals is important for the anti-aging effect of the purple bamboo shoot tea, and the appropriate concentrations (0.2, 0.4, 0.6, 0.8 and 1.0g L) are used by combining the experimental results^-1) The DPPH free radical scavenging experiment was carried out on the extract of Zisun bamboo shoot tea the results (FIG. 3) show that the extract of Zisun bamboo shoot tea was at a concentration of 0.2-1.0 g/L^-1The radical clearance rate is increased with increasing concentration, especially at concentrations of 0.4-1.0g L^-1The clearance rate of free radicals reaches 80 percentThe above.

2.4 Effect of Zisun tea on tyrosinase Activity inhibition

Melanocytes in the skin are affected by various intrinsic and extrinsic factors caused by the environment and neighboring cells, including UV, MSH, ASP, ET1, DKK1, and a large number of growth factors and cytokines, the hyperpigmentation of epidermal pigments is mainly due to excessive melanin synthesis and/or the transfer of excessive melanin to keratinocytes, and in order to lighten pigmented skin, the main objective is to keep the melanin synthesis and cut off its transfer pathway^[3]。

Melanocyte biosynthesis melanin takes tyrosine as a substrate, melanin is synthesized under the action of tyrosinase, and the melanin production amount is related to the activity of tyrosinase, so that the melanin production amount can be controlled by controlling the activity of tyrosinase [4]Using appropriate concentrations (0.2, 0.4, 0.6, 0.8, 1.0g L)^-1) The tyrosinase inhibition experiment was performed by applying the extract solution to a375 cells, and the results of the experiment (fig. 4, table 1) show that, compared to the initial values, the rate of change of the tyrosinase activity in a375 cells after the application of the extract solution was decreased with the increase of the concentration, and the concentration was 1.0 g/L^-1When compared to the initial value, the tyrosinase activity decreased by 23.3% (Table 1), i.e.the tyrosinase was most strongly inhibited.

TABLE 1 rate of change of tyrosinase activity

2.5 Effect of Zisun tea on MC1R protein expression

α -MSH as a naturally occurring endogenous peptide of melanocortin family, which stimulates melanin synthesis, α -MSH acts in the pigmentation region by binding to the MC1R receptor on the surface of the melanin cell membrane^[5,6]Meanwhile, the melanocortin receptor 1(MC1R) of the melanocyte is expressed on the surface of the melanocyte of a human body, is a G protein coupled receptor, is a receptor of α -MSH in the melanocyte, and is a key protein for regulating the proliferation and the function of the melanocyteDeposition, and activation of DNA damage responses in human melanocytes, including DNA repair pathways^[7,8,9]。

The result of the experiment through the cell-based E L ISA experiment is shown in figure 5, the A375 cell expression MSH receptor MC1R protein is reduced with the concentration of the purple bamboo shoot tea rising, the concentration is dependent, and the concentration is 0.4-1.0g L^-1When the protein expression of MC1R is significantly inhibited, especially at a concentration of 0.6-1.0g L^-1When the protein expression level is extremely obviously inhibited, the protein expression level has statistical significance.

Further western blot experiments were performed to verify that (FIG. 6), at appropriate concentrations (0.2, 0.4, 0.6, 0.8, 1.0g L)^-1) The A375 cell expressed MC1R protein of the extract liquid of the zizhu bamboo shoot tea is reduced along with the increase of the concentration, and has concentration dependence, and the protein strip is subjected to gray scale analysis by Tanon self-contained analysis software, and the result is represented by MC1R/β -actin, when the concentration of the zizhu bamboo shoot tea is 0.6-1.0 g/L^-1When the protein is expressed, the expression of the MC1R protein is extremely obviously inhibited, and the protein has statistical significance, particularly 0.8-1.0g L^-1At the concentration, the protein inhibition rate reaches more than 50 percent, and the aim of mutual verification is achieved.

2.6 Effect of Zisun tea on TRP-2 protein expression

As mentioned above, tyrosinase-related protein 2(TRP-2) is capable of catalyzing the subsequent steps of the melanin synthesis process, accelerating melanogenesis^[10]The results of the experiment (fig. 7) show that the concentration of the purple bamboo shoot tea is 0.2-1.0g L^-1Along with the increase of the concentration of the purple bamboo shoot tea, the expression of the TRP-2 protein of the A375 cells is reduced and has concentration dependence, and when the concentration is 0.2-1.0 g/L^-1When the expression of TRP-2 protein is significantly inhibited statistically, especially at a concentration of 0.6-1.0g L^-1When the protein is expressed, the expression level of the protein is remarkably inhibited.

2.7 Effect of Zisun tea on PAR-2 protein expression

The protein activator receptor 2(PAR-2) is expressed only in keratinocytes and not in melanocytes. The inventors have found that the increase of melanin in keratinocytes and the transport of melanin are observed by a method of co-culturing keratinocytes and melanocytes in three dimensionsIs related to, but is not related to the synthesis of melanin, and further finds that PAR-2 is involved in the transport process of melanin, is a main key receptor protein for melanin transport, and researches prove that the PAR-2 can mediate keratinocytes to actively phagocytose melanosomes and participate in the transport of melanin, thereby being capable of regulating skin pigmentation^[11,12,13]。

Through cell-based E L ISA experiments (figure 8), the expression of PAR-2 protein of HaCaT cells is reduced with the increase of the concentration of the purple bamboo shoot tea, and the expression is dependent on the concentration, particularly the concentration is 0.6-1.0 g/L^-1When the expression of PAR-2 protein is significantly inhibited. Subsequently, a cell immunohistochemical verification experiment (fig. 9) is carried out, a data graph is shot under a 10 × 20 microscope, 6 visual fields are randomly selected, and it is found that as the concentration of the purple bamboo shoot tea rises, the brown-yellow color of the HaCaT cells becomes lighter, which has concentration dependence and accords with the result shown in fig. 8.

2.8 Effect of Zisun tea on expression of HAS1 protein

Hyaluronic acid is a high molecular weight glycosaminoglycan, is a main component of an extracellular matrix, is a polymer with a water retention function naturally existing in human skin, is a main component of the skin extracellular matrix, has the effects of retaining water, lubricating, regulating osmotic pressure and the like, protects normal cells from being invaded by toxic cells, free radicals and the like, can promote cell proliferation and differentiation, can regulate and control the synthesis of collagen, and is widely considered as a natural skin moisturizing factor to have stronger moisturizing and anti-aging effects^[14,15,16]. Hyaluronic acid synthase exists on the plasma membrane and is capable of specifically acting on the hyaluronic acid synthesis process.

Through the cell-based E L ISA experiment, it is found that the HSF cell HAS1 protein expression is increased along with the increase of the concentration of the purple bamboo shoot tea (figure 9), and when the concentration is 0.8-1.0g L^-1When the HAS1 protein expression is remarkably promoted.

3 conclusion of the experiment

The Zisun tea is a tea which has no literature to verify the function in the field of skin care, and the research is used for in vitro level research and verification of the function of the Zisun tea as an active ingredient on the effects of resisting aging, whitening, resisting oxidation and the like of skin cells. The research finds that the purple bamboo shoot tea remarkably promotes the proliferation of keratinocytes and fibroblasts in a proper concentration range (figure 1, figure 2) and has a very strong anti-oxidation effect (figure 3). And the purple bamboo shoot tea has extremely strong effects of inhibiting TYR activity and TRP-2 protein expression (figure 4 and figure 7), and has extremely obvious inhibition effect on a receptor MC1R (figure 5 and figure 6), which indicates that the purple bamboo shoot tea can inhibit a melanin biosynthesis pathway. Experiments show that the purple bamboo shoot tea has a strong effect on melanin transport process, especially on the process that keratinocytes phagocytose melanosomes. And aiming at the synthesis of hyaluronic acid which is a natural moisturizing factor, the purple bamboo shoot tea HAS a good effect of promoting the expression of HAS1 (figure 10). According to the experimental results, the purple bamboo shoot tea has the cosmetic effects of resisting oxidation, whitening and resisting aging to a certain extent, and can be applied to skin care products as an additive.

Claims (10)

1. Application of Zisun bamboo shoot tea extract in preparing cosmetics with one, two or three of anti-aging, anti-oxidation and melanin synthesis and transfer inhibiting effects is prepared by extracting Zisun bamboo shoot tea with water, organic solvent or supercritical CO₂Liquid or solid components of the extraction process; the organic solvent is one or more of ethanol, butanediol and acetone.

2. Use of the extract of Phyllostachys Pubescens tea as claimed in claim 1 for the preparation of cosmetics having the effects of promoting proliferation of HaCaT cells and HSF cells, antioxidant activity, inhibiting key receptors in the synthesis and transport pathway of melanin or promoting expression of hyaluronic acid synthase 1(HAS 1).

3. The use according to claim 2, wherein the key receptors in the pathway for the synthesis and transport of melanin comprise Tyrosinase (TYP), tyrosine-related protein 2(TRP-2), melanocyte melanocortin receptor 1(MC1R), protease activated receptor 2 (PAR-2).

4. A cosmetic composition characterized by comprisingThe extract is prepared by carrying out water extraction, organic solvent extraction or supercritical CO extraction on the purple bamboo shoot tea₂Liquid or solid components of the extraction process; the organic solvent is one or more of ethanol, butanediol and acetone.

5. The purple bamboo shoot tea cosmetic composition of claim 4, wherein: the auxiliary materials acceptable in the field of cosmetics are selected from one or more of solvents, solubilizers, preservatives, antioxidants, pH regulators, penetration enhancers, liposomes, humectants, thickeners, chelating agents, skin feel regulators, surfactants, emulsifiers, essences, pigments and other efficacy additives.

6. The purple bamboo shoot tea cosmetic composition of claim 5, wherein: the composition is in the form of cream, lotion, solution, film, aerosol or spray.

7. The purple bamboo shoot tea cosmetic composition of claim 6, wherein: the composition is in the form of facial cleanser, bath lotion, cosmetic water, skin care gel, skin care lotion, skin care cream, essence, eye cream, facial mask, aerosol or spray.

8. The purple bamboo shoot tea cosmetic composition of claim 7, wherein: the purple bamboo shoot tea extract accounts for 0.001-100% of the total weight of the composition.

9. The purple bamboo shoot tea cosmetic composition of claim 8, wherein: the purple bamboo shoot tea extract accounts for 0.01-50% of the total weight of the composition.

10. The purple bamboo shoot tea cosmetic composition of claim 8, wherein: the purple bamboo shoot tea extract accounts for 0.5-20% of the total weight of the composition.

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