CN113876626A

CN113876626A - Composition with protective effect on skin cell damage induced by blue light and application of composition in cosmetics

Info

Publication number: CN113876626A
Application number: CN202111271004.XA
Authority: CN
Inventors: 曹崇江; 尧岚; 程抒劼
Original assignee: China Pharmaceutical University
Current assignee: China Pharmaceutical University
Priority date: 2021-10-29
Filing date: 2021-10-29
Publication date: 2022-01-04
Anticipated expiration: 2041-10-29
Also published as: CN113876626B

Abstract

The invention discloses a composition with a protection effect on skin cell damage induced by blue light, which comprises tremella aurantialba crude polysaccharide and glucosyl hesperidin, wherein the mass ratio of the tremella aurantialba crude polysaccharide to the glucosyl hesperidin is (0.01-1) to (0.01-4), and the mass ratio of the tremella aurantialba crude polysaccharide to the glucosyl hesperidin is more than 1:1. The composition not only has a protective effect on the keratinocyte damaged by blue light induction, but also can promote the proliferation and migration of skin keratinocyte after blue light irradiation, improve the delay of skin barrier repair caused by blue light, and further promote the repair of skin damaged barrier. The invention also discloses application of the composition or the tremella aurantialba crude polysaccharide or the glucosyl hesperidin in preparation of cosmetics, and the cosmetic prepared from the composition or the tremella aurantialba crude polysaccharide or the glucosyl hesperidin, other active ingredients and auxiliary materials is beneficial to inherent self-repair of skin barriers and has the effects of moisturizing, relieving and repairing.

Description

Composition with protective effect on skin cell damage induced by blue light and application of composition in cosmetics

Technical Field

The invention belongs to the technical field of medical beauty skin care products, relates to a composition with a protective effect on blue light induced skin cell damage and application thereof in cosmetics, and particularly relates to a composition containing tremella aurantialba crude polysaccharide and glucosyl hesperidin with a protective effect on blue light induced skin cell damage and application thereof in preparing cosmetics.

Background

Blue light is high-energy visible light with a wavelength of 400-500nm, which is not only present in sunlight, but also can be generated by various artificial light sources such as various displays in LED lamps, fluorescent lamps, incandescent lamps, televisions, tablet computers, and smart phones. People use electronic equipment such as mobile phones, computers and the like more and more frequently, even screen dependence is generated, and blue light emitted by the screens can affect eyes, biological rhythm and skin. To date, studies have shown that blue light causes the skin to produce oxidative stress and inflammatory factors, delays repair of the skin barrier, affects skin barrier homeostasis, and may further cause pigmentation. Although there is also an increasing number of studies showing the therapeutic effect of blue light of wavelength 453nm in chronic inflammatory skin diseases, it has an adverse effect on the normal healthy skin of humans, manifested by a higher TEWL (percutaneous water loss rate) and an increase in pigmentation 24 hours after barrier breakdown. Blue light affects the skin barrier homeostasis of normal healthy skin, and thus normal epidermal barrier function.

The skin consists of an epidermal layer, a dermal layer and a subcutaneous tissue, wherein the epidermal layer is a main barrier of a human body against external injury and consists of a basal layer, a spinous layer, a granular layer and an outermost horny layer. The stratum corneum is composed primarily of keratinocytes, intercellular lipids, and a sebaceous membrane 3, and generally represents the function of the skin barrier by the "brick-mud structure". The corneocytes in the stratum corneum constitute "bricks", the intercellular lipids constitute "mortar", and the intercellular lipids fill the "brick-mud structure" formed by the laminated corneocytes, which maintains the normal epidermal barrier function of the human body. Weather dryness, ultraviolet irradiation, aging, etc. all affect the barrier function of the skin, and maintaining and repairing the barrier function of the epidermis not only helps to prevent and treat skin diseases, but also can relieve the skin diseases to induce further infection and chronic inflammation. Keratinocytes (keratinocytos) are the main cells constituting the epidermis, and the keratinocytes located in the basal layer of the epidermis have the ability to proliferate, migrate and differentiate, are constantly dividing to generate new keratinocytes, migrate from the basal layer upwards, differentiate gradually and mature, and form the stratum spinosum, the stratum granulosum and the stratum corneum in this order. Keratinocytes play a very important role in the complex process of skin barrier repair, and the reduction of proliferation and migration of keratinocytes caused by blue light affects the repair of normal skin barrier.

Plant-derived active ingredients are of great interest for preventing the above-mentioned skin damage caused by blue light, since they are generally less toxic and may act synergistically with a variety of ingredients, such as antioxidant and repair activities. The pathogenesis of sensitive skin is not clear, and the individual difference of the induction factors and clinical manifestations is large. The mechanism is mainly considered to be the reduction of the skin barrier function at present, and aiming at the skin problem, the application of medical skin care products to repair and improve the damaged skin barrier is mainly focused at home and abroad.

At present, the compositions and skin care products claiming to resist blue light on the market are relatively few products for studying the stable state of the blue light to the epidermal barrier from the aspects of absorbing blue light and resisting photoaging. Therefore, it is urgently needed to screen out active ingredients with effective protection effect on blue light induced skin cell damage through cell experiments, and safely apply the active ingredients to skin care products to achieve the effect of repairing the blue light damaged skin barrier.

Disclosure of Invention

The invention aims to provide an active ingredient and a composition thereof with a protective effect on skin cell damage induced by blue light, the composition can obviously reduce ROS induced by blue light, improve weakening of keratinocyte migration capacity caused by blue light, and promote migration of keratinocyte, so that the repair of a damaged barrier is promoted; the composition can be applied to skin care cosmetics to effectively relieve the problems of skin barrier repair delay, skin dryness and the like caused by blue light.

The technical scheme of the invention is as follows:

the composition comprises golden fungus crude polysaccharide and glucosyl hesperidin in a mass ratio of (0.01-1) to (0.01-4), and the mass ratio of the golden fungus crude polysaccharide to the glucosyl hesperidin is more than 1:1.

Preferably, the mass ratio of the tremella aurantialba crude polysaccharide to the glucosyl hesperidin is 1: 1.5-1: 5.

More preferably, the mass ratio of the tremella aurantialba crude polysaccharide to the glucosyl hesperidin is 1: 1.5-1: 4.

Golden fungus (Tremella aurantialba), also known as Tremella aurantialba, is a precious edible and medicinal fungus, and is recorded in Chinese medicinal fungus with warm and cold property, sweet taste, fine colloid, and skin lubricating effect. The tremella aurantialba polysaccharide is one of main active ingredients of tremella aurantialba, and has the effects of immunoregulation, radiation resistance, oxidation resistance, moisture preservation, inflammation resistance and the like, and the tremella aurantialba, which is a rare precious strain in edible fungi, is mainly applied to the field of foods, is less in research and application in cosmetics, and basically has no research related to blue light, so that the application of the tremella aurantialba polysaccharide in future cosmetic raw materials has a wide market prospect. The tremella aurantialba crude polysaccharide is prepared by the following method:

drying and crushing fresh tremella aurantialba sporocarp, adding 95% ethanol according to a material-liquid ratio of 1:15(g: mL or kg: L), stirring at normal temperature for 24 hours, filtering, and drying a filter cake to obtain degreased dry powder;

adding deionized water into the dry degreased powder, stirring until the dry degreased powder swells, adjusting the pH to 5.5-6.5, adding cellulase and pectinase at the temperature of 45-55 ℃, and performing enzymolysis at the temperature of 45-55 ℃ for 30-45 min under heat preservation; and then carrying out water extraction at 75-80 ℃ for 1h, cooling, centrifuging, taking supernatant, carrying out evaporation concentration, adding absolute ethyl alcohol until the concentration reaches 75-80%, carrying out alcohol precipitation at 4 ℃ for 24h, centrifuging, taking precipitate, and drying to obtain the tremella aurantialba crude polysaccharide.

The dosage of the cellulase is 6 wt% of the degreasing dry powder, and the enzyme activity of the cellulase is 22U/mg; the dosage of the pectinase is 4% wt of the degreased dry powder, and the enzyme activity of the pectinase is 31.3U/mg.

Glucosyl Hesperidin (Glucosyl Hesperidin) has good water solubility, is a glucoside formed by hesperetin and rutinose, is a flavanone derivative, is an important component of citrus peel and pulp, and has the content of 1.4% of the weight of fresh citrus fruits.

The inventor finds that the tremella aurantialba crude polysaccharide and glucosyl hesperidin both have a certain protection effect on the skin cell damage induced by blue light, and can improve the skin barrier function by promoting the proliferation and migration capacity of keratinocytes and protecting the skin cells from the influence of artificial blue light from digital equipment. The golden fungus crude polysaccharide and glucosyl hesperidin are combined according to a specific ratio, and the creative discovery shows that the golden fungus crude polysaccharide and glucosyl hesperidin have a synergistic effect. In particular embodiments, the effect of the two combined is significantly better than the effect of the two used alone, in the case of a consistent total amount.

The invention also aims to provide the composition with the protective effect on the skin cell damage induced by blue light or the application of the tremella aurantialba crude polysaccharide or glucosyl hesperidin in the preparation of cosmetics.

A cosmetic, it is with said composition or crude polysaccharide of golden fungus or glucosyl hesperidin with protective action to blue light induced skin cell injury of the invention as the main active ingredient, with supplementary product to make into; wherein, the composition with the protective effect on the skin cell damage induced by blue light or the tremella aurantialba crude polysaccharide or glucosyl hesperidin accounts for 0.1-4% of the total weight of the cosmetic.

Preferably, the composition or the tremella aurantialba crude polysaccharide or the glucosyl hesperidin with the protection effect on the skin cell damage induced by blue light accounts for 0.1-3% of the total mass of the cosmetic according to the skin feel of the cosmetic.

Preferably, the auxiliary materials are as follows: at least one of solvent, thickener, antiseptic, pH regulator, humectant, skin conditioner, emollient, penetration enhancer, chelating agent, emulsifier, essential oil, essence, and pigment.

Specifically, the solvent may be selected from deionized water.

The thickener may be selected from carbomers.

The preservative is at least one of p-hydroxyacetophenone and 1, 2-hexanediol.

The pH modifier may be selected from arginine.

The humectant can be at least one selected from glycerol, sodium hyaluronate, betaine, trehalose, panthenol, polyglutamic acid and ceramide.

The emulsifier may be selected from at least one of cetearyl alcohol (and) cetearyl glucoside, hydrogenated lecithin.

The emollient is at least one selected from jojoba seed oil, isononyl isononanoate, hydrogenated polydecene, caprylic/capric triglyceride, polydimethylsiloxane and cetearyl alcohol.

The chelating agent may be selected from disodium EDTA.

Preferably, the cosmetic contains at least one of the following active ingredients: sodium hyaluronate, allantoin, polyglutamic acid, ceramide, astaxanthin, trehalose, panthenol, betaine, dipotassium glycyrrhizinate, tocopheryl acetate, ascorbyl glucoside, purslane extract, wolfberry fruit extract, ginseng root extract, poria cocos extract, green tea extract and pearl powder.

The cosmetic is selected from the group consisting of smoothing toner, facial mask, essence, eye cream, eye mask, skin care gel, skin care lotion, facial cream, and spray.

The eye cream is prepared from the following components in percentage by mass: 3 to 8% of glycerin, 0.1 to 0.3% of carbomer, 0.01 to 0.1% of sodium hyaluronate, 0.1 to 1% of p-hydroxyacetophenone, 0.01 to 0.1% of disodium EDTA, 1 to 4% of cetearyl alcohol (and) cetearyl glucoside, 0.1 to 3% of jojoba seed oil, 0.2 to 2% of isononyl isononanoate, 0.2 to 2% of hydrogenated polydecene, 0.1 to 2.5% of cetearyl alcohol, 0.1 to 2.5% of polydimethylsiloxane, 0.5 to 3% of hydrogenated lecithin, 0.1 to 2% of caprylic/capric triglyceride, 0.01 to 0.1% of allantoin, 0.4 to 2% of betaine, 0.4 to 2% of trehalose, 0.1 to 2% of tocopheryl acetate, 0.1 to 1% of panthenol, 0.1 to 1% of ceramide, 0.01 to 0.1% of polyglutamic acid, 0.04 to 0.2% of dipotassium glycyrrhizinate, 0.1 to 0.2% of a composition having a protective effect on skin cell damage induced by blue light, or a polysaccharide or a crude polysaccharide or a hesperidin, 0.1 to 1 to 1.1.1 to 1.1% of hesperidin, and a polysaccharide or a composition having a protective effect on skin damage induced by blue light, Adjusting the pH to 5.5-6.5 by using a pH regulator, and adding deionized water to 100%.

The invention also aims to provide a preparation method of the eye cream, which comprises the following steps:

mixing and heating cetearyl alcohol (and) cetearyl glucoside, jojoba seed oil, isononyl isononanoate, hydrogenated polydecene, cetearyl alcohol, polydimethylsiloxane, hydrogenated lecithin and caprylic/capric triglyceride to 75-80 ℃ to form an oil phase;

adding glycerol, carbomer, sodium hyaluronate, p-hydroxyacetophenone, allantoin and EDTA disodium into deionized water, heating to 70-75 ℃, and stirring and dispersing until the materials are completely dissolved to form a water phase;

slowly adding the oil phase into the water phase to mix the oil phase with the water phase, and homogenizing at 65-70 ℃ and 3000rpm for about 4 min;

cooling to 50 ℃, adding betaine, trehalose, panthenol, polyglutamic acid, dipotassium glycyrrhizinate, tocopheryl acetate, ceramide, tremella aurantialba crude polysaccharide and/or glucosyl hesperidin, fully and uniformly stirring, adding 1, 2-hexanediol, and adjusting the pH to 5.5-6.5 by using a pH regulator; standing and cooling to obtain eye cream.

Compared with the prior art, the invention has the beneficial effects that:

(1) according to the composition, the tremella aurantialba crude polysaccharide is compounded with glucosyl hesperidin with a strong antioxidation effect, and the tremella aurantialba crude polysaccharide and the glucosyl hesperidin have synergistic effect, so that the reduction of cell activity and proliferation capacity caused by blue light can be obviously improved, ROS induced by the blue light is reduced, and the migration of keratinocytes is promoted. The composition can protect cells from damage caused by artificial blue light, thereby helping to prevent skin damage caused by artificial blue light irradiation on a screen of an electronic device.

(2) The composition can be matched with other active ingredients and auxiliary materials to prepare related products. The product prepared by the formula can effectively promote the proliferation and regeneration of keratinocytes, thereby promoting the healing of damaged skin barriers and having remarkable moisturizing and repairing effects. In addition, compared with the existing moisturizing repair product which is sold in the market and is added with a plurality of chemical preservatives (such as parabens), the moisturizing repair product has the advantages that the components are simplified through the compounding and the corrosion prevention of the p-hydroxyacetophenone and the 1, 2-hexanediol, the skin is not irritated after long-time use, the moisturizing repair product is safe, mild and comfortable, can be widely applied to the subsequent care of the damaged skin barrier, and is suitable for all skin-type people.

(3) The cosmetic provided by the invention has the advantages of simple preparation process, easiness in reproduction, suitability for large-scale industrial production and wide market prospect.

Drawings

FIG. 1 is a glucose standard curve.

FIG. 2 is a protein standard curve.

FIG. 3 is a graph of the effect of active ingredients on the viability of skin keratinocytes illuminated by blue light; wherein ". x" indicates that the blank of the blue light group (LED-BL group) was significantly different (P < 0.0001) from the blank of the Control group (Control group); in the blue group, "+" indicates that treatments 1,2,3 had significant differences from the blank control (P < 0.01), "+" indicates that treatments 4,5,6,7 had significant differences from the blank control (P < 0.0001), "+" indicates that treatment 6 had significant differences from treatment 2 (P < 0.01), and "+" indicates that treatment 6 had significant differences from treatments 3,4 (P < 0.1).

FIG. 4 shows the effect of active ingredients on ROS production levels in keratinocytes irradiated with blue light; wherein ". x" indicates that blank 1 and blank 2 had significant differences (P < 0.0001); "x" indicates that there was a significant difference between blank 2 and treatments 1,2,3 (P < 0.0001); "x" indicates that treatment 3 and treatment 2 had significant differences (P < 0.01); ". indicates that treatment 3 and treatment 1 had significant differences (P < 0.0001).

FIG. 5 is a graph showing the effect of active ingredients on the ability of a blue light to irradiate keratinocytes to form cells for migration; wherein ". x" indicates that blank 1 and blank 2 had significant differences (P < 0.0001); "x" indicates that there was a significant difference between blank 2 and treatments 1,2,3 (P < 0.0001); ". indicates that treatment 3 was significantly different from treatment 1,2 (P < 0.01).

FIG. 6 shows the results of in vitro (25 ℃ C. + RH 80%) moisturizing tests on eye creams.

FIG. 7 shows the results of in vitro (25 ℃ C. + RH 40%) moisturizing tests on eye creams.

Detailed Description

In order to make the objects, technical solutions and advantages of the present invention more apparent, the technical solutions of the present invention are further described in detail with reference to the embodiments. It will be clear that the description is only illustrative and is not intended to limit the scope of the invention.

Example 1

The preparation method of the tremella aurantialba crude polysaccharide comprises the following steps:

drying and crushing fresh tremella aurantialba sporocarp to obtain dried tremella aurantialba powder, adding 95% ethanol according to a material-liquid ratio of 1:15(g: mL or kg: L), stirring at normal temperature for 24 hours for degreasing, performing suction filtration, and drying a filter cake to obtain degreased dry powder;

adding deionized water into the defatted dry powder obtained in the step (1), stirring until the dry powder is swelled, adding a proper amount of 1mol/L NaOH to adjust the pH value to 5.5, heating to 50 ℃, adding cellulase and pectinase (the enzyme activity of the cellulase is 22U/mg; the enzyme activity of the pectinase is 31.3U/mg; the dosage of the cellulase is 6 wt% of the defatted dry powder; the dosage of the pectinase is 4 wt% of the defatted dry powder), and carrying out heat preservation and enzymolysis for 30min at 50 ℃; extracting with water at 80 deg.C for 1 hr, cooling, centrifuging, collecting supernatant, rotary evaporating for concentration, adding anhydrous ethanol to 75%, precipitating with ethanol at 4 deg.C for 24 hr, centrifuging, collecting precipitate, and drying to obtain Tremella Aurantialba crude polysaccharide.

Determination of total sugar and protein content in tremella aurantialba crude polysaccharide

(1) Determination of the Total sugar content

Respectively placing 0, 0.1, 0.2, 0.4, 0.6, 0.8, and 1.0mL of glucose solution with concentration of 0.1mg/mL in a test tube with a plug, supplementing 1mL with distilled water, and respectively adding 1mL of 5% phenol and 5mL of concentrated H₂SO₄Shaking, standing at room temperature for 30min, measuring absorbance at 490nm, drawing standard curve (FIG. 1), and developing with water as blank control with glucose mass concentration as abscissa and absorbance as ordinate.

Preparing 1mg/mL sample solution with distilled water, sucking 0.5mL 1mg/mL sample solution, placing in a test tube with plug, supplementing 1mL with distilled water, and adding 1mL 5% phenol and 5mL concentrated H₂SO₄Shaking, standing at room temperature for 30min, measuring absorbance at 490nm, and calculating total sugar content according to standard curve.

(2) Determination of protein content

Accurately sucking 0, 0.1, 0.2, 0.4, 0.6, 0.8 and 1.0mL of standard bovine serum albumin solution into a 10mL test tube with a plug, supplementing the solution to 1mL with distilled water, respectively adding 5mL of Coomassie brilliant blue G-250 solution, fully shaking, standing at room temperature for 10min, measuring the absorbance value at the wavelength of 595nm, drawing a standard curve (figure 2), and taking water as a blank control according to the same color development, wherein the protein mass concentration is an abscissa and the absorbance value is an ordinate.

Taking tremella aurantialba crude polysaccharide, preparing a sample solution with the concentration of 1mg/mL by using distilled water, adding distilled water into 0.5mL of the sample solution to complement to 1mL, respectively adding 5mL of Coomassie brilliant blue G-250 solution (10mg of Coomassie brilliant blue, 5mL of 95% ethanol, 10mL of 85% phosphoric acid and distilled water to 100mL), fully shaking, standing at room temperature for 10min, measuring the absorbance value at the position of 595nm, measuring the absorbance value and calculating the protein content.

As can be seen from Table 1, the total sugar content of the crude Tremella aurantialba polysaccharide is 93.66%, and the protein content is 6.60%.

TABLE 1 Total sugar and protein content in Tremella Aurantialba crude polysaccharide

Example 2

A composition with a protection effect on skin cell damage induced by blue light comprises tremella aurantialba crude polysaccharide (example 1) and glucosyl hesperidin in a mass ratio of 1:1.

Example 3

A composition with a protection effect on skin cell damage induced by blue light comprises tremella aurantialba crude polysaccharide (example 1) and glucosyl hesperidin in a mass ratio of 2: 3.

Example 4

A composition with a protection effect on skin cell damage induced by blue light comprises tremella aurantialba crude polysaccharide (example 1) and glucosyl hesperidin in a mass ratio of 3: 7.

Example 5

A composition with a protection effect on skin cell damage induced by blue light comprises tremella aurantialba crude polysaccharide (example 1) and glucosyl hesperidin in a mass ratio of 1: 4.

Example 6

A composition with a protection effect on skin cell damage induced by blue light comprises tremella aurantialba crude polysaccharide (example 1) and glucosyl hesperidin in a mass ratio of 1: 5.

Example 7

Eye creams were prepared according to the formulation of table 2, respectively, by the following method:

mixing and heating cetearyl alcohol (and) cetearyl glucoside, jojoba seed oil, isononyl isononanoate, hydrogenated polydecene, cetearyl alcohol, polydimethylsiloxane, hydrogenated lecithin and caprylic/capric triglyceride to 75-80 ℃ to form an oil phase (phase B) for later use;

adding glycerol, carbomer, sodium hyaluronate, p-hydroxyacetophenone, allantoin and EDTA disodium into deionized water, heating to 70-75 ℃, stirring and dispersing until the mixture is completely dissolved to form a water phase (phase A) for later use;

slowly adding the phase B into the phase A to mix the phase A with the phase B, and homogenizing at the temperature of 70 ℃ and the rotating speed of 3000rpm for about 4 min;

step (4), cooling to 50 ℃, adding phase C components, namely betaine, trehalose, panthenol, polyglutamic acid, dipotassium glycyrrhizinate, tocopheryl acetate, ceramide, tremella aurantialba crude polysaccharide and/or glucosyl hesperidin, fully and uniformly stirring, adding phase D components, namely 1, 2-hexanediol and arginine, and adjusting the pH value to 6.03;

and (5) standing and cooling the cream obtained in the step (4), and bottling.

TABLE 2 formula of eye cream

Note: formulation 1 contained no blue light-resistant composition and was a blank control; formulas 2,3, 4 and 5 contain blue light resisting compositions, and the mass ratio of the tremella aurantialba crude polysaccharide to the glucosyl hesperidin is 2:3, 3:7, 1:4 and 1:5 respectively; formula 6 only contains tremella aurantialba crude polysaccharide; formulation 7 contained only glucosyl hesperidin.

In order to better illustrate the advantages of the present invention, the experimental results of the technical solution provided by the present invention are given below: and (3) examining the influence of the tremella aurantialba crude polysaccharide, glucosyl hesperidin and the combination of the tremella aurantialba crude polysaccharide and the glucosyl hesperidin on the protection effect of blue light damaged cells, the activity of HaCaT cells, the generation of ROS and the cell migration capacity.

First, the Effect of active ingredients on the Activity of skin keratinocytes irradiated with blue light

Selecting logarithmic phase growing keratinocyte (HaCaT cells), digesting, centrifuging, counting, diluting with MEM medium containing 10% fetal calf serum to 5 × 10⁴Concentration of/mL, seeded on 96-well cell culture plates, 100. mu.L per well. At 5% CO₂After 24 hours of incubation at 37 ℃ in an incubator (to ensure that cells were adherent), the medium in the 96-well plate was discarded, and 5 replicates of each group were set by adding MEM complete medium containing 10% fetal bovine serum of different samples (treatment 1: Aurea aurita crude polysaccharide, final concentration 0.04% in medium; treatment 2: glucosyl hesperidin, final concentration 0.04% in medium; treatment 3: example 2 composition, final concentration 0.04% in medium; treatment 4: example 3 composition, final concentration 0.04% in medium; treatment 5: example 4 composition, final concentration 0.04% in medium; treatment 6: example 5 composition, final concentration 0.04% in medium; treatment 7: example 6 composition, final concentration 0.04% in medium; MEM complete medium with 10% fetal bovine serum was blank). That is, HaCaT cells are all pretreated with drugs (after adding drugs, the cells are put into an incubator at 37 ℃ in the incubator with 5% CO)₂Middle incubation) for 24h, the medium was discarded, PBS buffer was added, and an LED blue light lamp (450nm, 100 mw/cm) was used above 15cm from the well plate²) The irradiation was carried out for 1 hour. Immediately after irradiation, the medium was replaced by MEM complete medium containing 10% fetal bovine serum, in a controlled humidified incubator (37 ℃, 5% CO)₂) Incubating for 24h, discarding culture medium in the well, adding 100 μ L MEM complete culture medium containing 10% fetal calf serum and 20 μ LMTT (MTT concentration is 5mg/mL, prepared by PBS buffer) into each well, and culturing for 4 h; terminating the culture, adding 150 mu L of dimethyl sulfoxide into each hole, and standing at room temperature for 1h to fully dissolve crystals; the absorbance (OD value) of each well at a wavelength of 570nm was measured by a microplate reader.

Meanwhile, a Control group (namely a Control group) which is not irradiated by blue light is arranged, the sample is pretreated for 24 hours, the culture medium is discarded, PBS buffer solution is added, and the sample is cultured for 1 hour in a dark place, and other treatments are the same as those of the LED-BL group.

The culture control wells were set to zero, and were free of cells, all other were identical to the assay wells.

The cell activity (%) was calculated according to the following formula, taking the average OD value of 5 parallel wells:

TABLE 3 Effect of active ingredients on the viability of skin keratinocytes irradiated with blue light

As can be seen from fig. 3 and table 3, in the blue light group, the cell activities of treatments 1,2,3, 4,5,6 and 7 were significantly higher than the cell activities of the blank control group, which indicates that the cell activities of tremella aurantialba crude polysaccharide, glucosyl hesperidin and the combination thereof under blue light irradiation can be improved, and the cell activities of treatments 4,5,6 and 7 are significantly higher than the cell activities of treatments 1,2 and 3, so that the ratio of tremella aurantialba crude polysaccharide to glucosyl hesperidin is 1: 1.5-1: 5, and particularly, the combination of the two at a ratio of 2:3, 3:7, 1:4 and 1:5 has a more significant improvement effect on the cell activities.

Second, the Effect of active ingredients on the level of ROS production in keratinocytes formed by irradiation with blue light

Selecting logarithmic phase growing keratinocyte, digesting, centrifuging, counting, diluting with MEM medium containing 10% fetal calf serum to 5 × 10⁴Concentration of/mL, seeded on 6 well cell culture plates, 2mL per well; at 5% CO₂Culturing at 37 deg.C for 24 hr, discarding culture medium in 6-well plate, adding MEM complete culture medium containing 10% fetal calf serum of different samples (treatment 1: Tremella Aurantialba crude polysaccharide, final concentration of 0.04% in culture medium; treatment 2: glucosyl hesperidin, final concentration of 0.0% in culture medium)4 percent; and (3) treatment: composition of example 5, final concentration 0.04% in medium; pretreating cells for 24h with 10% complete MEM culture medium containing fetal calf serum as blank, blank 1 without blue light irradiation, blank 2 with blue light irradiation, and then using LED blue light lamp (450nm, 100 mw/cm)²) The irradiation was carried out for 1 hour. Immediately after irradiation, 1mL of serum-free MEM basal medium containing active oxygen fluorescent probe DCFH-DA (10. mu.M/L) was added to each well, and after incubation for 30min, pictures were taken using a fluorescence microscope to quantitate the fluorescence of intracellular Reactive Oxygen Species (ROS), and the effects of blank controls 1 and 2 and treatments 1,2 and 3 on the amount of active oxygen produced by keratinocytes under blue light were examined, respectively.

TABLE 4 results of the Effect of active ingredients on ROS production levels in keratinocytes formed by irradiation with blue light

Sample (I)	Blank control 1	Blank control 2	Process 1	Treatment 2	Treatment 3
						Amount of active oxygen produced	2.888±0.291	51.999±2.217	38.971±1.365	33.109±2.399	25.256±1.452

As can be seen from fig. 4 and table 4, the active oxygen production amounts of the treatments 1,2 and 3 are significantly less than the percentage of the active oxygen production amount of the blank control 2, and the active oxygen production amount of the treatment 3 is also significantly less than the active oxygen production amounts of the treatments 1 and 2, which indicates that the tremella aurantialba crude polysaccharide and glucosyl hesperidin are synergistic with each other, so that the active oxygen produced under blue light radiation is effectively reduced, and the damage of blue light to the skin is effectively reduced.

Thirdly, the influence of the active ingredients on the migration capability of the blue light irradiation keratin-forming cells

HaCaT cells were cultured at 1.0X 10⁵The cells were inoculated in 6-well cell culture plates (medium: MEM complete medium containing 10% fetal bovine serum) at a concentration of 2mL per well; at 5% CO₂Culturing in 37 deg.C constant temperature incubator until cell fusion rate is 100%, scratching cell with 200 μ L gun head, and scratching with LED blue light lamp (450nm, 100 mw/cm)²) After 1h of irradiation, MEM basal medium containing 1% fetal bovine serum (treatment 1: golden fungus crude polysaccharide, the final concentration in the culture medium is 0.04%; and (3) treatment 2: glucosyl hesperidin with final concentration of 0.04% in culture medium; and (3) treatment: example 5 composition, final concentration 0.04% in culture medium, with MEM basal medium containing 1% fetal bovine serum as blank, blank 1 not irradiated with blue light after scratching, blank 2 irradiated with blue light) were incubated with cells for 24h, observed and photographed under microscope at 0h and 24h (0h for original area, 24h for area after scratching), and cell mobility was calculated using software Image J, formula:

cell mobility ═ (original area-post-scratch area)/original area × 100%

TABLE 5 results of the effect of the active ingredients on the migration ability of the keratinocytes formed by irradiation with blue light

As can be seen from fig. 5 and table 5, the cell migration ability of the treatments 1,2 and 3 is significantly higher than that of the blank controls 1 and 2, and the cell migration ability of the treatment 3 is significantly higher than that of the treatments 1 and 2, further indicating that the tremella aurantialba crude polysaccharide and glucosyl hesperidin can synergize with each other, effectively promote the migration of cells after blue light irradiation, thereby improving the repair ability of the skin barrier.

Fourth, the product has the sense physical and chemical indexes and the in vitro moisture retention test

(1) The sensory physical and chemical indexes of the product

The eye cream prepared in example 7 (formulation 2 for example) was subjected to sensory and stability tests according to conventional cosmetic test methods.

The results are shown in table 6 below, and the eye cream (taking formulation 2 as an example) meets the requirements of sensory and stability tests and has stable quality.

TABLE 6 sensory and stability test results for eye creams

(2) In vitro moisture retention test of the product

The in vitro moisture retention test was carried out at a constant temperature of 25 ℃ with relative humidities of 80% and 40%, respectively, every 2 hours for the first 8 hours, and finally every 24 hours.

Weighing empty beaker to obtain M₁When a proper amount of eye cream is smeared on the bottom of a beaker, the weight is obtained M₂Placing the beaker into a stable constant temperature and humidity box (25 ℃ RH 80%; 25 ℃ RH 40%), taking out the beaker at a certain time interval, and weighing to obtain M₃Calculating the water loss rate according to the following formula:

TABLE 7 in vitro moisture loss rate of eye cream (25 ℃ + RH 80%) for 24h

Sample (I)	Moisture loss ratio (%) for 24h
		Formulation
1	16.579
		Formulation 2	10.430
Formulation 3	11.286
		Formulation 4	12.089
Formulation 5	12.796
		Formulation 6	12.140
Formulation 7	13.095

TABLE 8 in vitro moisturizing of eye creams (25 ℃ + RH 40%) Water loss Rate over 24h

Sample (I)	Moisture loss ratio (%) for 24h
		Formulation
1	66.417
		Formulation 2	52.939
Formulation 3	55.186
		Formulation 4	57.697
Formulation 5	59.266
		Formulation 6	58.186
Formulation 7	59.904

As can be seen from fig. 6, fig. 7, table 7 and table 8, the in vitro moisturizing effect of formulations 2,3, 4,5,6 and 7 is significantly higher than that of formulation 1, indicating that the addition of any one or both of the ingredients of the composition has a better moisturizing effect. Meanwhile, the moisture-keeping effect of the formulas 2,3 and 4 is better than that of the formulas 6 and 7. Therefore, when the mass ratio of the tremella aurantialba crude polysaccharide to the glucosyl hesperidin is 1: 1.5-1: 4, the cosmetic product prepared by compounding the glucosyl hesperidin and the tremella aurantialba crude polysaccharide to form the composition has more remarkable moisturizing capability.

The experimental result shows that under the condition of consistent addition percentage, the two are mixed and used according to the proportion, the effect is better than that of single use, namely the effect of the composition is more obvious.

The above embodiments are only used for illustrating the technical solutions of the present invention, and not for limiting the same, although the present invention is described in detail with reference to the foregoing embodiments, those of ordinary skill in the art should understand that: the technical solutions described in the foregoing embodiments may still be modified, or some or all of the technical features may be equivalently replaced; and the modifications or the substitutions do not make the essence of the corresponding technical solutions depart from the scope of the technical solutions of the embodiments of the present invention.

Claims

1. A composition having a protective effect against blue light induced skin cell damage, comprising: the composition comprises tremella aurantialba crude polysaccharide and glucosyl hesperidin, the mass ratio of the tremella aurantialba crude polysaccharide to the glucosyl hesperidin is (0.01-1) to (0.01-4), and the mass ratio of the tremella aurantialba crude polysaccharide to the glucosyl hesperidin is more than 1:1.

2. The composition for protecting skin cell damage induced by blue light according to claim 1, wherein: the mass ratio of the tremella aurantialba crude polysaccharide to the glucosyl hesperidin is 1: 1.5-1: 5.

3. The composition for protecting skin cell damage induced by blue light according to claim 2, wherein: the mass ratio of the tremella aurantialba crude polysaccharide to the glucosyl hesperidin is 1: 1.5-1: 4.

4. The composition for protecting skin cell damage induced by blue light according to claim 1,2 or 3, wherein: the tremella aurantialba crude polysaccharide is prepared by the following method:

5. Use of the composition according to claim 1 or of a Tremella aurantialba crude polysaccharide or of glucosyl hesperidin for the preparation of a cosmetic.

6. A cosmetic characterized by: the composition is prepared by taking the composition or tremella aurantialba crude polysaccharide or glucosyl hesperidin as a main active ingredient and auxiliary materials, wherein the composition or the tremella aurantialba crude polysaccharide or glucosyl hesperidin is used as the main active ingredient; wherein, the composition with the protective effect on the skin cell damage induced by blue light or the tremella aurantialba crude polysaccharide or glucosyl hesperidin accounts for 0.1-4 percent of the total weight of the cosmetic, and the preferred content is 0.1-3 percent.

7. The cosmetic according to claim 6, characterized in that: the auxiliary materials are as follows: at least one of solvent, thickener, antiseptic, pH regulator, humectant, skin conditioner, emollient, penetration enhancer, chelating agent, emulsifier, essential oil, essence, and pigment.

8. The cosmetic according to claim 6, characterized in that: the cosmetic contains at least one of the following active ingredients: sodium hyaluronate, allantoin, polyglutamic acid, ceramide, astaxanthin, trehalose, panthenol, betaine, dipotassium glycyrrhizinate, tocopheryl acetate, ascorbyl glucoside, purslane extract, wolfberry fruit extract, ginseng root extract, poria cocos extract, green tea extract and pearl powder.

9. The cosmetic according to claim 6, characterized in that: the cosmetic is selected from the group consisting of smoothing toner, facial mask, essence, eye cream, eye mask, skin care gel, skin care lotion, facial cream, and spray.

10. An eye cream characterized by: the paint is prepared from the following components in percentage by mass: 3 to 8% of glycerin, 0.1 to 0.3% of carbomer, 0.01 to 0.1% of sodium hyaluronate, 0.1 to 1% of p-hydroxyacetophenone, 0.01 to 0.1% of disodium EDTA, 1 to 4% of cetearyl alcohol (and) cetearyl glucoside, 0.1 to 3% of jojoba seed oil, 0.2 to 2% of isononyl isononanoate, 0.2 to 2% of hydrogenated polydecene, 0.1 to 2.5% of cetearyl alcohol, 0.1 to 2.5% of polydimethylsiloxane, 0.5 to 3% of hydrogenated lecithin, 0.1 to 2% of caprylic/capric triglyceride, 0.01 to 0.1% of allantoin, 0.4 to 2% of betaine, 0.4 to 2% of trehalose, 0.1 to 2% of tocopheryl acetate, 0.1 to 1% of panthenol, 0.1 to 1% of ceramide, 0.01 to 0.1% of polyglutamic acid, 0.04 to 0.2% of dipotassium glycyrrhizinate, 0.1 to 0.2% of the crude polysaccharide composition having a protective effect on skin cell damage or 0.4 to 1% of hesperidin or the composition according to claim 1 0.1-1% of 1, 2-hexanediol, a pH regulator for regulating the pH to 5.5-6.5, and deionized water for 100%.