Plant culture with moisturizing and antioxidant efficacy and application thereof

Abstract

The present application relates to a kind of plant culture with moisturizing antioxidant efficacy and its application, the plant culture with moisturizing antioxidant efficacy is prepared by including the following steps: (1) separating kiwi placenta;(2) callus induction;(3) subculture;(4) suspension culture, harvest cell;(5) broken cell, centrifugal filtration, obtain the plant culture with moisturizing antioxidant efficacy.The present application can obtain stable optimized product by culturing kiwi placenta callus using plant cell biotechnology, and after extraction, it can be used as the raw material component of cosmetic freeze-dried powder, to promote whitening, moisturizing, antioxidant, repair, nourishing, soothing and other effects.

Description

TECHNICAL FIELD

[0001] The present application belongs to the technical field of cosmetics, and particularly relates to a plant culture with moisturizing and antioxidant effects and application thereof. BACKGROUND

[0002] Actinidia chinensis Planch., Chinese name is Chinese kiwi, commonly known as Tengli, Yangtao Teng, Yangtao, kiwi, several fruit, etc., is a plant of Actinidiaceae in Actinidiae. Kiwi is originally from the mountainous and hilly areas in southwest China, and gradually spread to other parts of the world in the middle of the 18th century. At present, the main planting areas include China, New Zealand, Italy, Iran, Chile, Greece and other countries. The main cultivated species of kiwi in the world are A. deliciosa and A. chinensis. The main cultivated varieties in China include Hayward, Qinmei, Cuxiang, Xuxiang, Guichang, Hongyang, Donghong, Jintao, Jinyan, Huayou, etc. The fruit core has green core, red core and yellow core. The main kiwi planting provinces in China include Shaanxi, Sichuan, Zhejiang, Yunnan, Guizhou, Chongqing, etc. The mature period of kiwi is from late August to late October.

[0003] Kiwi is a fruit with main economic and nutritional value, rich in vitamin C, vitamin E, folic acid, potassium, dietary fiber, fructose / glucose, other antioxidants (carotenoids lutein, zeaxanthin and beta-carotene, chlorophyll, quinic acid, caffeic acid glucosyl derivatives, beta-sitosterol, chlorogenic acid, phenolic substances including flavonoids and flavones) and several beneficial protein components. Long-term consumption of kiwi can regulate the intestinal tract, improve constipation, promote digestion, balance metabolic function, reduce blood lipids, protect the cardiovascular system, etc.

[0004] Plant cell culture refers to a kind of plant tissue culture mode in vitro, taking single cell or cell group as unit. This technology is not affected by time and space, external environment and character separation. It can cultivate relatively uniform cells on a large scale in a short time. Compared with animal cells, most plant cells can be cultured in suspension, so they are suitable for large-scale culture. By adjusting the physicochemical conditions of the bioreactor, the best growth, differentiation and metabolism environment of plant cells can be realized, the culture period is shortened, and favorable conditions are created for industrialized culture of plant cells or plants. Through the regulation of growth environment, the generation of various primary and secondary metabolites can also be optimized, so that beneficial ingredients are formed in balance.

[0005] Plant cell in vitro culture firstly needs to induce plant explants to obtain suitable subculture and production of hematic tissue of target product, the hematic tissue is dedifferentiated parenchyma cell generated from the cut part of explant under certain conditions, the cell is changed by human being to original differentiation state, loses original structure and function, and is transformed into undifferentiated cell, then forms a kind of parenchyma cell without specific structure and function which can rapidly increase.

[0006] Plant placenta, also known as plant placenta, is a part of plant fruit, located in edible part of fruit, specifically, the part where seeds are produced in fruit, the part where ovule is born in plant ovary. According to the position of placenta in fruit, the placenta is classified, including marginal placenta, lateral membrane placenta, central axis placenta, special central placenta, basal placenta and apical placenta. Kiwi belongs to central axis placenta type. Compared with other parts of plants, the placenta contains a large number of nutrients with biological activity, which is suitable to be induced into hematic tissue.

[0007] The placental hematic tissue is cultured by plant cell biotechnology, and stable optimized product can be obtained, which can be used as raw material component of cosmetic freeze-dried powder after extraction, and has the effects of promoting whitening, moisturizing, antioxidant, repair, nourishing, soothing and the like. SUMMARY

[0008] In view of the deficiencies of the prior art, the purpose of the present application is to provide a plant culture with moisturizing and antioxidant effects and application thereof.

[0009] To achieve this purpose, the technical scheme adopted by the present application is as follows:

[0010] In a first aspect, the present application provides a plant culture with moisturizing and antioxidant effects, which is prepared by a preparation method comprising the following steps:

[0011] (1) separating kiwi placenta;

[0012] (2) hematic tissue induction;

[0013] (3) subculture;

[0014] (4) suspension culture and harvesting of cells;

[0015] (5) cell disruption, centrifugation and filtration to obtain the plant culture with moisturizing and antioxidant effects.

[0016] The placental hematic tissue of kiwi is cultured by plant cell biotechnology, and stable optimized product can be obtained, which can be used as raw material component of cosmetic freeze-dried powder after extraction, and has the effects of promoting whitening, moisturizing, antioxidant, repair, nourishing, soothing and the like.

[0017] Preferably, the step (1) further comprises sterilizing the separated kiwi placenta.

[0018] Preferably, the sterilizing comprises sequentially washing with sterile water, immersing in an ethanol aqueous solution for 30-90s, washing with sterile water, sterilizing with a sodium hypochlorite solution for 20-40s, washing with sterile water, and drying.

[0019] The immersing time in the ethanol aqueous solution is, for example, 30s, 40s, 60s, 70s, 80s, 90s, etc.

[0020] The sterilizing time is, for example, 20s, 24s, 28s, 32s, 36s, 40s.

[0021] Preferably, the step (2) comprises inoculating the placenta material into a solid induction medium.

[0022] Preferably, the step (3) comprises inoculating the cultured 10-30 day-old callus into a liquid subculture medium, mixing and dispersing, and subculturing for 2-4 generations.

[0023] The 10-30 days are, for example, 10 days, 14 days, 16 days, 21 days, 28 days, 30 days, etc. The subculture generations are, for example, 2 generations, 3 generations, 4 generations, etc. Any specific point value within the above numerical range can be selected.

[0024] Preferably, the step (4) comprises transferring the cells after subculture into a liquid medium and suspending culture for 30-60 days.

[0025] The suspending culture time can be 30 days, 35 days, 40 days, 45 days, 50 days, 55 days, 60 days, etc. Any specific point value within the above numerical range can be selected.

[0026] In a second aspect, the present application provides a use of the plant culture with moisturizing and antioxidant effects according to the first aspect in the preparation of a skin care product.

[0027] The cosmetic prepared from the plant culture with moisturizing and antioxidant effects has good whitening and skin moisturizing effects and low-sensitivity repair effects.

[0028] In a third aspect, the present application provides a cosmetic containing the plant culture with moisturizing and antioxidant effects, which comprises the plant culture with moisturizing and antioxidant effects according to the first aspect, a sea buckthorn extract, and a rose extract.

[0029] Preferably, the cosmetic containing the plant culture with moisturizing and antioxidant effects further comprises a skin conditioning agent, a pH adjusting agent, a moisturizing agent, and a solvent.

[0030] Preferably, the mass ratio of the plant culture with moisturizing antioxidant efficacy, the sea buckthorn extract and the rose extract is (1-3):(2-4):(2-4).

[0031] The present application creatively uses the above three plant active ingredients for whitening and anti-sensitivity repair. The three plant active ingredients complement each other and have synergistic effects. Each component promotes skin whitening, moisturizing, antioxidant and soothing through different pathways or mechanisms, so that the effect of the product is significant and long-lasting.

[0032] The specific point value of 1-3 can be 1, 1.2, 2, 2.3, 2.5, 3, etc.

[0033] The specific point value of 2-4 is independently selected from 2, 2.3, 2.6, 2.8, 3, 3.3, 3.8, 4, etc.

[0034] The specific point values in the above numerical range can be selected, which will not be described one by one here.

[0035] Preferably, the moisturizing agent is selected from any one or a combination of at least two of glycerol, butylene glycol, 1,2-hexanediol, 1,2-pentanediol or 1,3-propanediol.

[0036] Compared with the prior art, the present application has the following beneficial effects:

[0037] The present application can obtain stable and optimized products by culturing kiwi placenta callus using plant cell biotechnology. After extraction, it can be used as a raw material component of cosmetic freeze-dried powder, and has the effects of promoting whitening, moisturizing, antioxidant, repair, nourishment and soothing. DETAILED DESCRIPTION

[0038] To facilitate understanding of the present application, the present application is illustrated as follows. Those skilled in the art should understand that the examples are only to help understand the present application and should not be regarded as specific limitations of the present application.

[0039] Some components in the preparation examples, examples and comparative examples are as follows:

[0040] The rose extract is derived from Hebei Sacred Rose Development Co., Ltd. with model number SDMG1030, the sea buckthorn extract is derived from Shaanxi Snoot Biotechnology Co., Ltd. with model number SNT-095, the gynura acid is derived from Xi'an Ruilin Biotechnology Co., Ltd. with concentration of 90%, and the gynura extract is derived from Shaanxi Jinkangtai Biotechnology Co., Ltd. with concentration of 80%.

[0041] Preparation of solid induction medium for plant tissue culture: (solid induction MS medium: NH4NO3 1650 mg / L, KNO3 1900 mg / L, KH2PO4 440 mg / L, MgSO4·7H2O 370 mg / L, CaCl2·2H2O 440 mg / L, KI 0.83 mg / L, H3BO3 6.2 mg / L, MnSO4·4H2O 22.3 mg / L, ZnSO4·7H2O 8.6 mg / L, Na2MoO4·2H2O 0.25 mg / L, CuSO4·5H2O 0.025 mg / L, CoCl2·6H2O 0.025 mg / L, FeSO4·7H2O 27.8 mg / L, Na2-EDTA·2H2O 37.3 mg / L, myo-inositol 100 mg / L, nicotinic acid 0.5 mg / L, VB6 0.5 mg / L, VB1 0.4 mg / L, glycine 2 mg / L, sucrose 30 g / L, agar 6 g / L, adjust pH to 5.8±0.5, sterilize at 115°C for 20 min, add naphthalene acetic acid (NAA) to 1.0 mg / L, 6 benzyl purine (6-BA) to 1.0 mg / L, pour into plates).

[0042] Preparation Example 1

[0043] Preparation of plant culture with moisturizing and antioxidant effects:

[0044] (1) Select 5 domestic Guichang varieties of Greenheart kiwifruit, cut and separate the placentas with a knife; rinse with water for 20 min, rinse with distilled water 3 times, and dry. Transfer the placenta to a clean bench, rinse with sterile water 3 times, soak in 75% ethanol for 60 s, then rinse with sterile water 3 times. Disinfect with 0.2% sodium hypochlorite solution for 30 s, then rinse with sterile water 4 times, and absorb the excess water on the placenta with sterile filter paper.

[0045] (2) Cut the placenta into small pieces of 3-5 mm and inoculate into solid induction MS medium, and culture at a temperature of 25°C under light with an intensity of 2000 lx for 12 h per day.

[0046] (3) Inoculate the callus cultured for 15 days into liquid subculture medium (MS medium without agar), mix and disperse. Culture at 25°C under light with an intensity of 2000 lx for 12 h per day. Change the medium every 14 days and subculture for 3 generations.

[0047] (4) Select loose and vigorous growth callus after subculture, crush and transfer into liquid medium (MS liquid medium: NH4NO3 1650 mg / L, KNO3 1900 mg / L, KH2PO4 440 mg / L, MgSO4·7H2O 370 mg / L, CaCl2·2H2O 440 mg / L, KI 0.83 mg / L, H3BO3 6.2 mg / L, MnSO4·4H2O 22.3 mg / L, ZnSO4·7H2O 8.6 mg / L, Na2MoO4·2H2O 0.25 mg / L, CuSO4·5H2O 0.025 mg / L, CoCl2·6H2O 0.025 mg / L, FeSO4·7H2O 27.8 mg / L, Na2-EDTA·2H2O 37.3 mg / L, myo-inositol 100 mg / L, nicotinic acid 0.5 mg / L, VB6 0.5 mg / L, VB1 0.4 mg / L, glycine 2 mg / L, sucrose 30 g / L, adjust pH to 5.8±0.5, sterilize at 115°C for 20 min, add NAA to 1.0 mg / L, 6-BA to 0.8 mg / L, culture at 26°C, 16 h light / 8 h dark per day, 100 rpm, suspend culture for 14 days, add 20% fresh medium every 5 days, continue to culture for 20 days. Filter to harvest cell mass, rinse with distilled water, dry. Crush the cells with a crusher, centrifuge the supernatant at 4000 rpm, filter the supernatant to obtain placenta cell culture.

[0048] Preparation Example 2

[0049] Preparation of plant culture with moisturizing and antioxidant effects:

[0050] (1) Select 5 domestic Jintao varieties of red heart kiwifruit, cut and separate the placenta with a knife; rinse with water for 25 min, rinse with distilled water 3 times, and dry. Transfer the placenta to a clean bench, rinse with sterile water 3 times, soak in 75% ethanol for 45 s, then rinse with sterile water 3 times. Disinfect with 0.2% sodium hypochlorite solution for 30 s, then rinse with sterile water 4 times, and dry the excess water on the placenta with sterile filter paper.

[0051] (2) Cut the placenta into small pieces of 3-5 mm and inoculate into solid induction MS medium. Culture at 25°C with light, light intensity 2000 lx, 12 h light per day.

[0052] (3) Inoculate the callus cultured for 14 days into liquid subculture medium (MS medium without agar), mix and disperse. Inoculate into solid induction MS medium, culture at 25°C with light, light intensity 2000 lx, 12 h light per day. Change the medium every 14 days for 3 generations of subculture.

[0053] (4) Select loose and vigorous growth callus after subculture, and then transfer the callus into liquid medium (MS liquid medium, sterilized at 115°C for 20 min, and then add NAA to 1.0 mg / L and 6-BA to 0.8 mg / L) after the callus is broken by glass beads. The culture temperature is 26°C, the light and dark time is 16 h / 8 h, and the rotation speed is 100 rpm. The callus is cultured for 12 days, and then 20% fresh medium is added every 6 days for further culture for 24 days. The cell mass is harvested by filtration, washed with distilled water, and dried. The cell mass is broken by a crusher, and then the supernatant is separated by centrifugation at 4500 rpm. The supernatant is filtered through a 300-mesh sieve to obtain the placenta cell culture.

[0054] Preparation Example 3

[0055] The preparation example provides a preparation method of a plant culture with moisturizing and antioxidant effects. The only difference between the preparation example and Preparation Example 1 is that the subculture generation number is 1 generation, and other conditions remain unchanged. The preparation method is the same as that in Preparation Example 1.

[0056] Preparation Example 4

[0057] The preparation example provides a preparation method of a plant culture with moisturizing and antioxidant effects. The only difference between the preparation example and Preparation Example 1 is that the subculture generation number is 6 generations, and other conditions remain unchanged. The preparation method is the same as that in Preparation Example 1.

[0058] Preparation Example 5

[0059] The preparation example provides a preparation method of a plant culture with moisturizing and antioxidant effects. The only difference between the preparation example and Preparation Example 1 is that “20% fresh medium is added every 5 days for further culture for 20 days” in step (4) is changed to “20% fresh medium is added every 5 days for further culture for 48 days”, and other conditions remain unchanged. The preparation method is the same as that in Preparation Example 1.

[0060] Preparation Example 6

[0061] The preparation example provides a preparation method of a plant culture with moisturizing and antioxidant effects. The only difference between the preparation example and Preparation Example 1 is that “20% fresh medium is added every 5 days for further culture for 20 days” in step (4) is changed to “20% fresh medium is added every 5 days for further culture for 8 days”, and other conditions remain unchanged. The preparation method is the same as that in Preparation Example 1.

[0062] Example 1

[0063] The example provides a lyophilized powder containing a plant culture with moisturizing and antioxidant effects. The lyophilized powder comprises the following components:

[0064] Plant culture with moisturizing and antioxidant effects prepared in Preparation Example 1, sea buckthorn extract, rose extract, total 9 portions (and satisfying the mass ratio of 5:6:6), glycerin 2.5 portions, butylene glycol 1 portion, licorice root extract 0.4 portion, p-hydroxyacetophenone 0.2 portion, 1,2-hexanediol 0.3 portion, sodium hyaluronate 0.25 portion, disodium EDTA 0.05 portion, 1,2-pentanediol 0.3 portion, 1,3-propanediol 1.5 portions, asiatic acid 0.1 portion, oligopeptide-1 0.15 portion, gynostemma extract 0.2 portion, ascorbic acid 0.02 portion, caprylyl glycol 0.02 portion, sodium dihydrogen phosphate 0.002 portion, disodium hydrogen phosphate 0.0024 portion, acetyl hexapeptide-8 0.1 portion, ethylhexylglycerin 0.1 portion, fibronectin 0.15 portion.

[0065] The preparation method for the freeze-dried powder is as follows: the above components are dissolved and uniformly mixed with purified water, divided into 3 ml / bottle, and then vacuum freeze-dried at -70℃ to seal the freeze-dried powder with rubber stoppers.

[0066] Example 2

[0067] The present example provides a freeze-dried powder containing plant culture with moisturizing and antioxidant effects, which comprises the following ingredients:

[0068] Plant culture with moisturizing and antioxidant effects prepared in Preparation Example 1, sea buckthorn extract, rose extract, total 9 portions (and satisfying the mass ratio of 1:2:2), glycerin 2.5 portions, butylene glycol 1 portion, licorice root extract 0.4 portion, p-hydroxyacetophenone 0.2 portion, 1,2-hexanediol 0.3 portion, sodium hyaluronate 0.25 portion, disodium EDTA 0.05 portion, 1,2-pentanediol 0.3 portion, 1,3-propanediol 1.5 portions, asiatic acid 0.1 portion, oligopeptide-1 0.15 portion, gynostemma extract 0.2 portion, ascorbic acid 0.02 portion, caprylyl glycol 0.02 portion, sodium dihydrogen phosphate 0.002 portion, disodium hydrogen phosphate 0.0024 portion, acetyl hexapeptide-8 0.1 portion, ethylhexylglycerin 0.1 portion, fibronectin 0.15 portion.

[0069] The preparation method for the freeze-dried powder is as follows: the above components are dissolved and uniformly mixed with purified water, divided into 3 ml / bottle, and then vacuum freeze-dried at -70℃ to seal the freeze-dried powder with rubber stoppers.

[0070] Example 3

[0071] The present example provides a freeze-dried powder containing plant culture with moisturizing and antioxidant effects, which comprises the following ingredients:

[0072] The plant culture with moisturizing and antioxidant effects prepared in Preparation Example 2, sea buckthorn extract, and rose extract totaling 9 parts (and satisfying the mass ratio of 3:2:2), glycerin 2.5 parts, butylene glycol 1 part, licorice root extract 0.4 parts, p-hydroxyacetophenone 0.2 parts, 1,2-hexanediol 0.3 parts, sodium hyaluronate 0.25 parts, disodium EDTA 0.05 parts, 1,2-pentanediol 0.3 parts, 1,3-propylene glycol 1.5 parts, asiatic acid 0.1 parts, oligopeptide-1 0.15 parts, Centella asiatica extract 0.2 parts, ascorbic acid 0.02 parts, caprylyl glycol 0.02 parts, sodium dihydrogen phosphate 0.002 parts, disodium hydrogen phosphate 0.0024 parts, acetyl hexapeptide-8 0.1 parts, ethylhexylglycerin 0.1 parts, and fibronectin 0.15 parts.

[0073] The preparation method of the lyophilized powder is as follows: the above components are dissolved and mixed evenly with purified water, and the mixture is divided into 3 ml / bottle, and the mixture is freeze-dried in a vacuum freeze-drying state at -70°C and sealed with a rubber stopper to obtain the lyophilized powder.

[0074] Example 4

[0075] This embodiment provides a freeze-dried powder of a plant culture having moisturizing and antioxidant effects. The only difference from Example 1 is that an equal amount of the kiwifruit placenta culture prepared in Preparation Example 1 is replaced by an equal amount of the kiwifruit placenta culture prepared in Preparation Example 3. Other conditions remain unchanged, and the preparation method refers to Example 1.

[0076] Example 5

[0077] This embodiment provides a freeze-dried powder of a plant culture having moisturizing and antioxidant effects. The only difference from Example 1 is that an equal amount of the kiwifruit placenta culture prepared in Preparation Example 1 is replaced by an equal amount of the kiwifruit placenta culture prepared in Preparation Example 4. Other conditions remain unchanged, and the preparation method refers to Example 1.

[0078] Example 6

[0079] This embodiment provides a freeze-dried powder of a plant culture having moisturizing and antioxidant effects. The only difference from Example 1 is that an equal amount of the kiwifruit placenta culture prepared in Preparation Example 1 is replaced by an equal amount of the kiwifruit placenta culture prepared in Preparation Example 5. Other conditions remain unchanged, and the preparation method refers to Example 1.

[0080] Example 7

[0081] This embodiment provides a freeze-dried powder of a plant culture having moisturizing and antioxidant effects. The only difference from Example 1 is that an equal amount of the kiwifruit placenta culture prepared in Preparation Example 1 is replaced by an equal amount of the kiwifruit placenta culture prepared in Preparation Example 6. Other conditions remain unchanged, and the preparation method refers to Example 1.

[0082] Example 8

[0083] The present example provides a cosmetic lyophilized powder containing plant culture with moisturizing and antioxidant effects, which differs from Example 1 only in that the plant culture with moisturizing and antioxidant effects, the sea buckthorn extract, and the rose extract meet a mass ratio of 1:4:6, the total mass of the plant culture with moisturizing and antioxidant effects, the sea buckthorn extract, and the rose extract remains unchanged, and the other conditions remain unchanged. The preparation method is referred to Example 1.

[0084] Example 9

[0085] The present example provides a cosmetic lyophilized powder containing plant culture with moisturizing and antioxidant effects, which differs from Example 1 only in that the plant culture with moisturizing and antioxidant effects, the sea buckthorn extract, and the rose extract meet a mass ratio of 1:6:7, the total mass of the plant culture with moisturizing and antioxidant effects, the sea buckthorn extract, and the rose extract remains unchanged, and the other conditions remain unchanged. The preparation method is referred to Example 1.

[0086] Comparative Example 1

[0087] The present comparative example provides a cosmetic lyophilized powder containing plant culture with moisturizing and antioxidant effects, which differs from Example 1 only in that it does not contain sea buckthorn extract, and the reduced portion of the sea buckthorn extract is proportionally distributed to the plant culture with moisturizing and antioxidant effects and the rose extract, and the other conditions remain unchanged. The preparation method is referred to Example 1.

[0088] Comparative Example 2

[0089] The present comparative example provides a cosmetic lyophilized powder containing plant culture with moisturizing and antioxidant effects, which differs from Example 1 only in that it does not contain rose extract, and the reduced portion of the rose extract is proportionally distributed to the plant culture with moisturizing and antioxidant effects and the sea buckthorn extract, and the other conditions remain unchanged. The preparation method is referred to Example 1.

[0090] Comparative Example 3

[0091] The present comparative example provides a cosmetic lyophilized powder, which differs from Example 1 only in that it does not contain plant culture with moisturizing and antioxidant effects, and the reduced portion of the plant culture with moisturizing and antioxidant effects is proportionally distributed to the rose extract and the sea buckthorn extract, and the other conditions remain unchanged. The preparation method is referred to Example 1.

[0092] Comparative Example 4

[0093] The comparative example provides a plant culture freeze-dried powder containing a plant culture with moisturizing and antioxidant effects, which is different from Example 1 only in that it does not contain rose extract and sea buckthorn extract, and the part of the rose extract and sea buckthorn extract reduced is allocated to the plant culture with moisturizing and antioxidant effects, and the other conditions remain unchanged. The preparation method is referred to Example 1.

[0094] Test Example 1

[0095] Soothing effect - human test

[0096] In addition to meeting the conventional screening conditions, the subjects are further screened by the lactic acid stinging test: 50 μL of 10% lactic acid solution is added to the nasal labial fold on either side at room temperature, and the subjects are asked about their subjective symptoms at 2.5 min and 5 min, and the scores are rated on a scale of 1 to 4 (1 for no stinging, 2 for mild stinging, 3 for moderate stinging, and 4 for severe stinging). The scores at the two inquiry time points are added together, and those with a total score of ≥3 are positive and can be used as subjects.

[0097] One hundred and fifty healthy subjects are screened and randomly divided into a blank group, a test group (Examples 1-9, Comparative Examples 1-4), 10 in each group. Before testing, the subjects are uniformly cleaned on the face with a non-lint absorbent dry paper towel. The blank group is normal saline, and the test group is the reconstituted solution sample of Examples 1-9 and Comparative Examples 1-4 freeze-dried powder. The subjects use the above products once every 2 days, and the lactic acid stinging test and subjective evaluation are carried out after 14 days, 28 days, and 56 days to compare the differences between different time periods and different groups. The spss 26 software is used for statistics. The results of the stinging test are shown in Table 1.

[0098] Table 1

[0099]

[0100] According to the above table, the blank group has a decrease, but the stinging score is not relieved and is still greater than or equal to 3. The stinging scores of the examples and comparative examples decrease significantly, and since the plant culture with moisturizing and antioxidant effects is added, the data of the examples after 56 days have a significant advantage over the comparative examples, indicating that they have a significant soothing effect.

[0101] Test Example 2

[0102] Repairing effect - human test

[0103] The test subjects in this test example are the same as in Test Example 1, and the grouping is the same as in Test Example 1. The subjects are required to undergo a skin transepidermal water loss rate test, and the facial skin is tested using a skin transepidermal water loss rate tester (Germany Courage + Khazaka electronic GmbH, Tewameter TM 300). The subjects are balanced in the test environment for 30 min, and are kept relaxed. The transepidermal water loss rate and hemoglobin content of the test area on the cheeks of the subjects are measured, and are recorded as the base value.

[0104] The transepidermal water loss rate (TEWL) and hemoglobin content are measured after 14 days, 28 days, and 56 days of use of the product, and the blank group uses normal saline. The data are statistically analyzed using spss 26 software, and are shown in Tables 2 and 3.

[0105] Table 2

[0106]

[0107] Table 3

[0108]

[0109] In Tables 2 and 3, the transepidermal water loss rate (TEWL) and hemoglobin content of the examples are significantly lower than those of the comparative examples and the blank group. According to these two data, it can be considered that the examples have a certain repair effect. The reduction effect of Examples 1-3 is more obvious, indicating that the ratio thereof is more reasonable and effective.

[0110] Test Example 3

[0111] Nourishing effect - human test

[0112] Test principle: The nourishing effect of the product is verified by measuring the improvement in skin roughness, gloss, dermal density, or elasticity of the subjects.

[0113] The subjects in this test example are the same as in Test Example 1, and the grouping is the same as in Test Example 1. The above-mentioned subjects are tested for skin roughness, gloss, and elasticity using a skin texture tester (Germany Courage + Khazaka electronic GmbH, VC20 Plus), a skin gloss tester (Germany Courage + Khazaka electronic GmbH, Glossymeter GL200), and a skin elasticity tester (Germany Courage + Khazaka electronic GmbH, Cutometer MPA 580). The subjects are given sample products and comparative sample products, respectively, and the skin roughness, gloss, and elasticity are measured after 14 days, 28 days, and 56 days of use of the product. The data are statistically analyzed using spss 26 software.

[0114] The test results are shown in Tables 4-6. Table 4 is the result of skin roughness, Table 5 is the test result of skin elasticity, and Table 6 is the test result of skin glossiness.

[0115] Table 4

[0116]

[0117] Table 5

[0118]

[0119] Table 6

[0120]

[0121] The three data of skin roughness, skin elasticity and glossiness can directly reflect the function of the test product on skin nourishment and improvement. In Tables 4-6, the skin roughness and wrinkle depth of Examples 1-9 are significantly reduced, and the skin elasticity and glossiness are obviously improved compared with the comparative examples, which indicates that the examples, especially Examples 1-3, have better nourishing effect.

[0122] Test Example 4

[0123] Antioxidant effect - zebrafish model

[0124] After the indicator CM-H2DCFDA is added into the zebrafish larvae culture system, it can produce relatively stable high-intensity fluorescence under the action of intracellular reactive oxygen species. By detecting the fluorescence intensity, the generation of free radicals in zebrafish can be understood, which is suitable for the evaluation of the antioxidant effect of the test sample.

[0125] Randomly selected 3dpf zebrafish larvae with normal development and consistent stages were transferred to 24-well plates, 10 per well. Zebrafish control group, no fish control group, positive group, test group, sample control group were set up, the positive group was 1.5 μmol / L menadione solution; the freeze-dried powder was reconstituted with the corresponding solvent liquid as the test solution, the test group and the sample control group were the dilutions (diluted 2, 4, 8, 16 times, respectively) of Examples 1-9 and Comparative Examples 1-4; the no fish control group only added sample liquid without zebrafish; the zebrafish control group only had zebrafish without adding the test substance. Add 1 ml of sample to each well, add CM-H2DCFDA to 0.5 μg / ml, mix well, transfer the zebrafish to a 96-well plate, 1 fish per well, 100 μL of liquid. Wrap the 96-well plate with aluminum foil to avoid light, incubate in a 28°C constant temperature incubator for 20 h. After incubation, use a multifunctional enzyme marker (Thermo Fisher Varioskan LUX, USA) to detect the fluorescence value S (excitation wavelength 485 nm, emission wavelength 535 nm). The fluorescence value of the test substance treatment group minus the fluorescence value of the no fish group is the relative content of reactive oxygen species ROS in the zebrafish body treated with the test substance. Use spss 26 software for statistics.

[0126] The ROS clearance rate is calculated according to the following formula:

[0127] ROS clearance rate (%) =

[0128] The test results are shown in Table 7.

[0129] Table 7

[0130]

[0131] As can be seen from the above table, the freeze-dried powder sample of Example added with plant culture having moisturizing and antioxidant effects has a higher ROS clearance rate than the comparative example without addition, and still has a certain removal effect at a lower concentration, indicating that the example has better antioxidant effect.

[0132] Test Example 5

[0133] Whitening efficacy-zebrafish model

[0134] The neural crest of zebrafish has three kinds of pigment cells, melanocytes, xanthophores and iridophores, which are differentiated from intermediate progenitor cells, so the surface of zebrafish has pigments, which can be used as an animal model for whitening efficacy evaluation.

[0135] AB wild zebrafish was used for breeding and breeding according to the method of Westerfield. The system water pH value was set to 7.2~7.4, and the conductivity was 450~550 µS / m. The system used filter purified water, and the system water pH value and conductivity were adjusted by automatic suction of sodium chloride and sodium bicarbonate to meet the set parameters. The fish room temperature was 28℃, and the light cycle was 14h light / 10h dark. Light stimulation spawning, collection of embryos, and washing with water. Under a microscope, normal developing embryos were selected and placed in an incubator for further culture. A 24-well plate was used as the test container, and 24hpf zebrafish embryos were randomly placed in it. Each sample was set up an example group, a blank group, and a comparison group, and each group used 10 embryos. The freeze-dried powder reconstituted sample was added to each well at 100μL, and the blank group was added with normal saline. Incubate in an incubator for 48h, and observe the development of the embryo surface melanin under a fluorescence microscope. Take pictures and record, and use ImageJ software (NIH, USA) to calculate the melanin plaque area. Compared with the blank group, the melanin inhibition rate of the test group and the control group was calculated. The test results are shown in Table 8.

[0136] Table 8

[0137]

[0138] According to the analysis of the size of the plaque area, the blank group has no melanin removal effect, the comparison group has a certain effect, the examples have significant differences compared with the comparison group, and the examples 1-3 are significantly higher than the other examples, which shows that the examples of the test have whitening effect.

[0139] The applicant declares that the above-mentioned examples are used to illustrate the detailed process equipment and process flow of the present application, but the present application is not limited to the above-mentioned detailed process equipment and process flow, that is, it does not mean that the present application must rely on the above-mentioned detailed process equipment and process flow to be implemented. Those skilled in the art should understand that any improvement of the present application, equivalent replacement of each raw material of the product of the present application, addition of auxiliary ingredients, selection of specific methods, etc. fall within the protection scope and disclosure scope of the present application.

Claims

1. A method for preparing a plant culture having moisturizing and antioxidant effects, characterized in that: The plant culture with moisturizing and antioxidant effects is prepared by the following steps: (1) Separation of kiwifruit placenta; (2) Callus induction; the callus induction comprises the following steps: inoculating the placenta material into a solid induction medium for culture; the solid induction medium comprises 1650 mg / L NH4NO3, 1900 mg / L KNO3, 440 mg / L KH2PO4, 370 mg / L MgSO4·7H2O, 440 mg / L CaCl2·2H2O, 0.83 mg / L KI, 6.2 mg / L H3BO3, 22.3 mg / L MnSO4·4H2O, 8.6 mg / L ZnSO4·7H2O, 0.25 mg / L Na2MoO4·2H2O, 0.025 mg / L CuSO4·5H2O, 0.025 mg / L CoCl2·6H2O, 27.8 mg / L FeSO4·7H2O, and 27.8 mg / L Na2-EDTA·2H2O. 37.3mg / L, inositol 100mg / L, niacin 0.5mg / L, VB6 0.5mg / L, VB1 0.4mg / L, glycine 2mg / L, sucrose 30g / L, agar 6g / L, NAA 1.0mg / L, 6-BA 1.0mg / L; (3) Subculture; the subculture comprises the following steps: inoculating the callus cultured for 10-30 days into a liquid subculture medium, dispersing it, and subculture it for 2-4 generations; the liquid subculture medium is MS medium without adding agar; (4) suspension culture and cell harvesting; the suspension culture comprises the following steps: transferring the cells after subculture into liquid culture medium and suspending culture for 30-60 days; the liquid culture medium comprises 1650 mg / L NH4NO3, 1900 mg / L KNO3, 440 mg / L KH2PO4, 370 mg / L MgSO4·7H2O, 440 mg / L CaCl2·2H2O, 0.83 mg / L KI, 6.2 mg / L H3BO3, 22.3 mg / L MnSO4·4H2O, 8.6 mg / L ZnSO4·7H2O, 0.25 mg / L Na2MoO4·2H2O, 0.025 mg / L CuSO4·5H2O, 0.025 mg / L CoCl2·6H2O, 0.025 mg / L FeSO4·7H2O 27.8mg / L, Na2-EDTA·2H2O 37.3mg / L, inositol 100mg / L, niacin 0.5mg / L, VB6 0.5mg / L, VB1 0.4mg / L, glycine 2mg / L, sucrose 30g / L, NAA 1.0mg / L, 6-BA 0.8mg / L; (5) Disrupting the cells and centrifuging to obtain the plant culture with moisturizing and antioxidant effects.

2. The method for preparing a plant culture having moisturizing and antioxidant effects according to claim 1, wherein: After the kiwifruit placenta is separated in step (1), disinfection is also performed.

3. The method for preparing a plant culture having moisturizing and antioxidant effects according to claim 2, wherein: The disinfection comprises sequentially washing with sterile water, soaking in an ethanol-water solution for 30-90 seconds, washing with sterile water, disinfecting with a sodium hypochlorite solution for 20-40 seconds, washing with sterile water, and drying.

4. Use of the plant culture with moisturizing and antioxidant effects prepared by the preparation method according to any one of claims 1 to 3 in the preparation of skin care products.

5. A cosmetic containing a plant culture having moisturizing and antioxidant effects, characterized in that: The active ingredients of the cosmetic containing the plant culture with moisturizing and antioxidant effects are the plant culture with moisturizing and antioxidant effects prepared by the preparation method according to any one of claims 1 to 3, sea buckthorn extract and rose extract.

6. The cosmetic containing plant culture having moisturizing and antioxidant effects according to claim 5, characterized in that: The cosmetic containing the plant culture having moisturizing and antioxidant effects further comprises a skin conditioner, a pH adjuster, a moisturizer and a solvent.

7. The cosmetic containing plant culture having moisturizing and antioxidant effects according to claim 5, characterized in that: The mass ratio of the plant culture with moisturizing and antioxidant effects, the sea buckthorn extract and the rose extract is (1-3): (2-4): (2-4).

8. The cosmetic containing plant culture having moisturizing and antioxidant effects according to claim 6, characterized in that: The moisturizing agent is selected from any one of glycerol, butylene glycol, 1,2-hexanediol, 1,2-pentanediol or 1,3-propylene glycol, or a combination of at least two thereof.

Images