WO2022218173A1 - 一种面膜液、其制备方法及应用 - Google Patents

一种面膜液、其制备方法及应用 Download PDF

Info

Publication number
WO2022218173A1
WO2022218173A1 PCT/CN2022/084840 CN2022084840W WO2022218173A1 WO 2022218173 A1 WO2022218173 A1 WO 2022218173A1 CN 2022084840 W CN2022084840 W CN 2022084840W WO 2022218173 A1 WO2022218173 A1 WO 2022218173A1
Authority
WO
WIPO (PCT)
Prior art keywords
facial mask
mask liquid
hours
barley
pineapple
Prior art date
Application number
PCT/CN2022/084840
Other languages
English (en)
French (fr)
Inventor
吴迪
刘继涛
安全
霍彤
刘平平
王昌涛
石秀芹
Original Assignee
云南白药集团健康产品有限公司
云南白药集团股份有限公司
北轻家化(黄山)科技有限公司
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by 云南白药集团健康产品有限公司, 云南白药集团股份有限公司, 北轻家化(黄山)科技有限公司 filed Critical 云南白药集团健康产品有限公司
Publication of WO2022218173A1 publication Critical patent/WO2022218173A1/zh

Links

Images

Classifications

    • AHUMAN NECESSITIES
    • A61MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61KPREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K8/00Cosmetics or similar toiletry preparations
    • A61K8/18Cosmetics or similar toiletry preparations characterised by the composition
    • A61K8/96Cosmetics or similar toiletry preparations characterised by the composition containing materials, or derivatives thereof of undetermined constitution
    • A61K8/97Cosmetics or similar toiletry preparations characterised by the composition containing materials, or derivatives thereof of undetermined constitution from algae, fungi, lichens or plants; from derivatives thereof
    • A61K8/9783Angiosperms [Magnoliophyta]
    • A61K8/9789Magnoliopsida [dicotyledons]
    • AHUMAN NECESSITIES
    • A61MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61KPREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K8/00Cosmetics or similar toiletry preparations
    • A61K8/18Cosmetics or similar toiletry preparations characterised by the composition
    • A61K8/92Oils, fats or waxes; Derivatives thereof, e.g. hydrogenation products thereof
    • A61K8/922Oils, fats or waxes; Derivatives thereof, e.g. hydrogenation products thereof of vegetable origin
    • AHUMAN NECESSITIES
    • A61MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61KPREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K8/00Cosmetics or similar toiletry preparations
    • A61K8/18Cosmetics or similar toiletry preparations characterised by the composition
    • A61K8/96Cosmetics or similar toiletry preparations characterised by the composition containing materials, or derivatives thereof of undetermined constitution
    • A61K8/97Cosmetics or similar toiletry preparations characterised by the composition containing materials, or derivatives thereof of undetermined constitution from algae, fungi, lichens or plants; from derivatives thereof
    • A61K8/9783Angiosperms [Magnoliophyta]
    • A61K8/9794Liliopsida [monocotyledons]
    • AHUMAN NECESSITIES
    • A61MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61QSPECIFIC USE OF COSMETICS OR SIMILAR TOILETRY PREPARATIONS
    • A61Q19/00Preparations for care of the skin
    • AHUMAN NECESSITIES
    • A61MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61QSPECIFIC USE OF COSMETICS OR SIMILAR TOILETRY PREPARATIONS
    • A61Q19/00Preparations for care of the skin
    • A61Q19/08Anti-ageing preparations
    • AHUMAN NECESSITIES
    • A61MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61KPREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K2800/00Properties of cosmetic compositions or active ingredients thereof or formulation aids used therein and process related aspects
    • A61K2800/20Chemical, physico-chemical or functional or structural properties of the composition as a whole
    • A61K2800/30Characterized by the absence of a particular group of ingredients
    • AHUMAN NECESSITIES
    • A61MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61KPREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K2800/00Properties of cosmetic compositions or active ingredients thereof or formulation aids used therein and process related aspects
    • A61K2800/40Chemical, physico-chemical or functional or structural properties of particular ingredients
    • A61K2800/59Mixtures
    • A61K2800/592Mixtures of compounds complementing their respective functions
    • A61K2800/5922At least two compounds being classified in the same subclass of A61K8/18

Definitions

  • the invention belongs to the field of cosmetics, and in particular relates to a facial mask liquid, a preparation method and application thereof.
  • Natural cosmetics are more and more popular nowadays. Natural cosmetics, in addition to natural ingredients, do not add flavors, preservatives and other potentially irritating ingredients.
  • enzymolysis can transform macromolecular components into small molecules, which are accompanied by the production of other active substances such as polysaccharides and polypeptides.
  • Plants contain a variety of enzymes, including protease, cellulase, amylase, esterase, etc.
  • the enzymes contained in plants can be used to hydrolyze macromolecular proteins into oligopeptides, and starch and cellulose into oligosaccharides. , lipolysis into glycerol, fatty acid.
  • the application of these products in cosmetics is more conducive to the skin's absorption of functional ingredients, and finally achieves a certain effect.
  • the purpose of the present invention is to overcome the defects in the prior art, and to provide a facial mask liquid, its preparation method and application.
  • MDA malondialdehyde
  • PBS refers to: Phosphate Buffered Saline.
  • t-test refers to: T-test.
  • MTT Cytotoxicity Test
  • VE Vitamin E
  • VC Vitamin C
  • a first aspect of the present invention provides a facial mask liquid, the facial mask liquid does not contain any chemical additives, and is made from pure plant raw materials and water comprising the following components: pineapple, oat, highland barley, barley and vegetable oil;
  • the vegetable oil is selected from one or more of the following: olive oil, green prickly ash oil, peony seed oil, macadamia nut oil, babassu oil, toona sinensis oil; preferably olive oil.
  • the proportion by weight of each raw material component in the facial mask liquid is:
  • the second aspect of the present invention provides the preparation method of the facial mask liquid described in the first aspect, and the preparation method may comprise the following steps:
  • step (2) after the oats, highland barley, and barley are physically pulverized and sieved, they are added to the extract obtained in step (1), heated and stirred, cooled to room temperature, and centrifuged to obtain the supernatant to obtain an enzymatic hydrolysis treatment solution;
  • step (3) adding vegetable oil to the supernatant liquid obtained in step (2) and homogenizing to obtain the facial mask liquid.
  • the ratio of pineapple to water is 1:1 ⁇ 500g/mL, preferably 1:5 ⁇ 500g/mL, more preferably 1:10 ⁇ 400g /mL, more preferably 1:50 ⁇ 200g/mL; and/or
  • the standing time is 1 to 5 hours, preferably 2 to 4 hours, and most preferably 3 hours.
  • step (2) the oats, highland barley, and barley are pulverized and passed through an 80-mesh sieve.
  • the mass ratio of the oatmeal and the extract obtained in the step (1) is 1-50:100, preferably 1-20:100, more It is preferably 1 to 10:100, more preferably 1 to 5:100;
  • the mass ratio of the highland barley to the extract obtained in step (1) is 1-10:100, preferably 1-5:100, more preferably 1-3:100, further preferably 1.25-2:100; and/or
  • the mass ratio of the coix seed and the extract obtained in step (1) is 1-10:100, preferably 1-5:100, more preferably 1-3:100, further preferably 1.25-2:100.
  • the heating temperature is 20-150°C, preferably 25-100°C, more preferably 30-100°C, most preferably 40°C;
  • the heating time is 1-10 hours, preferably 1-8 hours, more preferably 2-6 hours, most preferably 4 hours;
  • the stirring rate is 10-1000r/min, preferably 10-800r/min, more preferably 10-600r/min, most preferably 200r/min; and/or
  • the centrifugal rotation speed is 10-10000r/min, preferably 100-8000r/min, more preferably 200-6000r/min, most preferably 1200r/min.
  • the added amount of the vegetable oil is 1-50%, preferably 1-40%; more preferably 10-30%, further preferably 10-40% 20%;
  • the homogenization speed is 3000 ⁇ 8000r/min, preferably 3000 ⁇ 7000r/min, more preferably 4000 ⁇ 6000r/min; further preferably 4000 ⁇ 5000r/min; and/or
  • the homogenization time is 1 to 60 minutes, preferably 1 to 50 minutes, more preferably 10 to 40 minutes, still more preferably 20 to 30 minutes.
  • the method further comprises the following steps:
  • step (3) gained facial mask liquid is sterilized
  • the sterilization temperature is 80-100°C, preferably 80°C; and/or the sterilization time is 1-5 hours, preferably 2 hours.
  • a third aspect of the present invention provides a facial mask comprising the facial mask liquid of the first aspect or the facial mask liquid prepared according to the method of the second aspect.
  • a fourth aspect of the present invention provides a face sticker, the face sticker comprising the facial mask liquid of the first aspect or the facial mask liquid prepared according to the method of the second aspect.
  • the fifth aspect of the present invention provides the application of the facial mask solution described in the first aspect or the facial mask solution prepared by the method described in the second aspect in the preparation of an anti-aging product.
  • the anti-aging is to significantly increase the collagen content of fibroblasts.
  • the lipid peroxide is malondialdehyde.
  • a sixth aspect of the present invention provides a method for anti-aging, the method comprising: administering to a subject in need:
  • the preparation method of facial mask liquid of the present invention comprises the following steps:
  • the ratio of the pineapple to water is 1:1 ⁇ 500g/mL, preferably 1:5 ⁇ 500g/mL, more preferably 1:10 ⁇ 400g/mL, most preferably 1:100g/mL mL;
  • the ratio of the oatmeal to the extract (1) is 1:1 ⁇ 300g/mL, preferably 1:20 ⁇ 200g/mL, more preferably 1:20 ⁇ 150g/mL, most preferably 1:100g/mL;
  • the ratio of the highland barley to the extract (1) is 1:1 to 200 g/mL, preferably 1:10 to 150 g/mL, more preferably 1:30 to 100 g/mL, and most preferably 1:80 g/mL;
  • the ratio of the coix seed and the extract (1) is 1:1 ⁇ 200g/mL, preferably 1:10 ⁇ 150g/mL, more preferably 1:50 ⁇ 100g/mL, most preferably 1:80g/mL;
  • the heating temperature is 1-150°C, preferably 20-100°C, more preferably 30-100°C, most preferably 40°C;
  • the heating time is 1-10 hours, preferably 1-8 hours, more preferably 2-6 hours, most preferably 4 hours
  • the stirring rate is 10-1000 r/min, preferably 10-800 r/min, more preferably 10-600 r/min, most preferably 200 r/min.
  • the centrifugal rotation speed is 10-10000r/min, preferably 100-8000r/min, more preferably 200-6000r/min, most preferably 1200r/min.
  • the added amount of the vegetable oil is 1-50%, preferably 1-40%; more preferably 10-30%, most preferably 20%;
  • the homogenization speed is 3000-8000r/min, preferably 3000-7000r/min, more preferably 4000-6000r/min; the most preferably 5000r/min;
  • the homogenization time is 1 to 60 minutes, preferably 1 to 50 minutes, more preferably 10 to 40 minutes, and most preferably 30 minutes.
  • the facial mask liquid of the present invention does not add any potentially irritating ingredients such as essence, preservatives, etc., in addition to the natural ingredients in the formula. Tests have proved that the facial mask liquid of the present invention has a promoting effect on the proliferation of fibroblasts, and has a certain transdermal absorption effect.
  • Figure 1 shows the HPLC profile of sample 1 at 220 nm in Example 1.
  • FIG. 2 shows the HPLC profile of sample 2 at 220 nm in Example 1.
  • FIG. 3 shows the HPLC profile of the blank group at 220 nm in Example 1.
  • FIG. 4 shows the toxicity results of different concentrations of facial mask liquid of the present invention on fibroblasts in Test Example 1.
  • FIG. 5 shows the transdermal absorption results of the mask liquid of the present invention in Test Example 2.
  • FIG. 6 shows the test results of the effect of the facial mask liquid of the present invention on the content of collagen in fibroblasts in Test Example 3.
  • FIG. 6 shows the test results of the effect of the facial mask liquid of the present invention on the content of collagen in fibroblasts in Test Example 3.
  • Oats (fried and cooked) were purchased from Zhangjiakou Jianjun Oat Food Co., Ltd.;
  • Barley was purchased from Beijing Tongrentang (Group) Co., Ltd., China;
  • Olive oil was purchased from Yunnan Yuchenglong Economic and Trade Co., Ltd.;
  • Fibroblasts were purchased from Saibaikang (Shanghai) Biotechnology Co., Ltd.
  • PBS purchased from Beijing Soleibo Technology Co., Ltd.
  • NaOH was purchased from Sinopharm Chemical Reagent Co., Ltd.
  • Homogenizer purchased from IKA Co., Ltd., model: T25;
  • the medical centrifuge was purchased from Wuxi Ruijiang Analytical Instrument Co., Ltd., model RJ-TGL-10B.
  • This example is used to illustrate the selection of the pineapple enzyme solution of the present invention.
  • the experimental method is as follows:
  • pineapple enzyme solution 1 is Barry pineapple
  • pineapple enzyme solution 2 is golden diamond pineapple.
  • Sample preparation use the mobile phase as the solvent to prepare a sample with a concentration of 5 mg/ml, and then filter it with a microporous membrane (0.45 ⁇ m) for injection.
  • Standard sample bovine serum albumin (Mr 67000), B12 (Mr 1335), and oxidized glutathione (Mr 614) were prepared into mixed standards, and the content of each substance was 5mg/ml.
  • Figures 1 to 3 show the HPLC chromatograms of sample 1, sample 2 and blank group at 220 nm in Example 1, respectively.
  • the peak time, area and molecular weight of sample 1, sample 2 and blank group are shown in Table 1 below.
  • This example is used to illustrate the preparation method of the facial mask liquid of the present invention.
  • step (3) heating the mixed solution obtained in step (2) at 70° C. for 4 hours, stirring at 100 r/min; cooling to room temperature, centrifuging at 500 r/min, and taking the supernatant.
  • the mask liquid is filled into the mask bag (including the mask cloth).
  • This example is used to illustrate the preparation method of the facial mask liquid of the present invention.
  • step (3) The mixed solution obtained in step (2) was heated at 40° C. for 4 hours, stirred at 500 r/min; cooled to room temperature, centrifuged at a speed of 2000 r/min, and the supernatant was taken.
  • the mask liquid is filled into the mask bag (including the mask cloth).
  • Oats, highland barley, and barley crushed and passed through 80 mesh for use.
  • step (3) The mixed solution obtained in step (2) was heated at 40° C. for 4 hours, stirred at 200 r/min; cooled to room temperature, centrifuged at a speed of 1200 r/min, and the supernatant was taken.
  • the homogenizer rotating speed is 5000r/min and homogenizing for 25 minutes.
  • the mask liquid is filled into the mask bag (including the mask cloth).
  • Oats, highland barley, and barley crushed and passed through 80 mesh for use.
  • step (3) The mixed solution obtained in step (2) was heated at 40° C. for 4 hours, stirred at 200 r/min; cooled to room temperature, centrifuged at 1200 r/min, and the supernatant was taken.
  • step (3) (4) adding peony seed oil with a mass of 10% of the supernatant to the supernatant obtained in step (3), and the homogenizer rotates at 4000 r/min for 30 minutes.
  • the mask liquid is filled into the mask bag (including the mask cloth).
  • Oats, highland barley, and barley crushed and passed through 80 mesh for use.
  • step (3) The mixed solution obtained in step (2) was heated at 80° C. for 6 hours, and stirred at 600 r/min; cooled to room temperature, centrifuged at a speed of 6000 r/min, and the supernatant was taken.
  • step (3) (4) adding macadamia nut oil with a mass of 30% of the supernatant to the supernatant obtained in step (3), and the homogenizer rotating speed is 6000r/min for 40 minutes.
  • the mask liquid is filled into the mask bag (including the mask cloth).
  • Oats, highland barley, and barley crushed and passed through 80 mesh for use.
  • pineapple and water are mixed and beaten according to a ratio of 1:500 g/mL, and allowed to stand at room temperature for 5 hours to obtain an extract (1).
  • step (3) The mixed solution obtained in step (2) was heated at 100° C. for 1 hour, stirred at 800 r/min; cooled to room temperature, centrifuged at a rotating speed of 8000 r/min, and the supernatant was taken.
  • the mask liquid is filled into the mask bag (including the mask cloth).
  • This test example is used to illustrate the effect of the facial mask liquid prepared in Example 2 on the proliferation of fibroblasts.
  • Collect logarithmic phase cells adjust the concentration of cell suspension, add 100 ⁇ L to each well (96-well plate), and plate the cells to adjust the density to 5000 cells/well (the edge wells are filled with sterile PBS), 5% CO 2 , Incubate at 37°C until the cells grow to a certain density, change the medium and add the samples to be tested with different concentration gradients, and use the culture medium without the drug to be tested as a control. 5% CO 2 , incubated at 37°C for 24 hours, 20 ⁇ L of MTT solution (5 mg/mL, 0.5% MTT tetrazolium salt) was added to each well, and the culture was continued for 4 hours.
  • MTT solution 5 mg/mL, 0.5% MTT tetrazolium salt
  • the culture medium can be discarded after centrifugation, and the culture medium containing MTT (FM-Fibroblast Medium purchased from ScienCell) can be added after carefully washing with PBS for 2-3 times. The culture was terminated, and the medium in the well was carefully aspirated. 150 ⁇ L of dimethyl sulfoxide was added to each well, and it was shaken at low speed on a shaker for 10 min to fully dissolve the crystals. The absorbance value of each well was measured at the OD value of 490 nm in an enzyme-linked immunosorbent assay.
  • MTT FM-Fibroblast Medium purchased from ScienCell
  • Cell viability (OD value of assay well-OD value of blank control)/(OD value of cell control group-OD value of blank control)*100%.
  • Results The test results of different concentration gradients of the samples were compared. To investigate the toxicity of different concentrations of facial mask liquid to fibroblasts. The results are shown in Figure 4. The experimental results show that the mask liquid has no obvious cytotoxicity, and the cell survival rate increases significantly in the range of the sample concentration of 5% to 40%, about 140% to 180%. It shows that the mask liquid has a promoting effect on the proliferation of fibroblasts.
  • mice were sacrificed by decapsulation of their cervical vertebrae, their back hair was quickly shaved with a razor, the back skin was peeled off, and subcutaneous fat and blood vessels were removed, rinsed repeatedly with distilled water until clean, and then rinsed with normal saline several times, and placed in a -80°C refrigerator. Refrigerate for later use (use within 5 days).
  • Q is the cumulative permeation amount
  • S is the transdermal diffusion area
  • V is the volume of the receiving chamber of the improved Franz diffusion cell
  • Cn is the concentration of the receiving liquid during the nth sampling
  • Ci is the concentration of the receiving liquid during the i-th sampling
  • 0.5 is the sample size.
  • This test example is used to illustrate the effect of the facial mask liquid prepared in Example 3 on the collagen content of fibroblasts.
  • Dilution of the standard dilute the standard lyophilized powder to 150 ⁇ l (160ng/ml) with the standard diluent, and then mix for 30s. Take five clean EP tubes, add 150 ⁇ l of standard dilution to each tube, and mark them with 80ng/ml, 40ng/ml, 20ng/ml, 10ng/ml and 5ng/ml respectively. Take 150 ⁇ l of the standard solution and add it to the tube marked 80ng/ml. After mixing, take out 150 ⁇ l of the standard solution and add it to the next tube, and so on to the last tube. (Zero wells directly add standard dilution)
  • b. Dilution of biotin antigen Centrifuge the concentrated biotin antigen at 6000-10000rpm for 30s, then take 1ml of biotin antigen dilution into the concentrated biotin antigen, mix for 15s until completely dissolved, then pour all the liquid into In the diluent bottle, mix well to obtain the biotin antigen working solution.
  • Dilution of avidin-HRP centrifuge the concentrated biotin antigen at 6000-10000rpm for 30s, transfer all the concentrated avidin-HRP to the diluent bottle, and mix well to obtain the avidin-HRP working solution .
  • Dilution of washing solution Pour the 25-fold concentrated solution in the kit into a 500ml volumetric flask, and dilute to 500ml with distilled water, which is the working solution.
  • Washing method first suck off the liquid in the enzyme-labeled wells, pat dry on absorbent paper, add 300 ⁇ l of washing solution to each well, shake gently for 30s, then shake off, pat dry on absorbent paper, repeat 4-5 times.
  • Standard wells add 50 ⁇ l of diluted standard to each well, add 50 ⁇ l of standard/sample diluent to zero well, and then add 50 ⁇ l of biotin antigen working solution.
  • Sample well add 50 ⁇ l of sample, and then add 50 ⁇ l of biotin antigen working solution.
  • the first washing Carefully peel off the sealing film, discard the liquid, spin dry, add 200 ⁇ l of washing solution to each well, let it stand for 30 s, and then discard, repeat 5 times, and drain.
  • the second washing Carefully peel off the sealing film, discard the liquid, spin dry, fill each well with washing liquid, let it stand for 30 s, then discard, repeat this for 5 times, and pat dry.
  • Color development First add 50 ⁇ l of color developer A to each well, then add 50 ⁇ l of color developer B, gently shake and mix, and develop color at 37°C for 10 minutes in the dark.
  • results The test results of the effect of facial mask liquid on the content of collagen in fibroblasts are shown in Figure 6.
  • the model group was the cell group that was only irradiated with UVA and no sample was added, the blank group was the cell group that was not irradiated and no sample was added, and the positive control group was the positive control sample VC added with a mass concentration of 0.1 g/L after UVA irradiation. cell group.
  • the test found that UVA irradiation will seriously damage the collagen of the cells.
  • the sample group was compared with the model group, blank group and positive control group. As can be seen from the figure, the mask liquid can significantly increase the content of collagen and achieve anti-aging effect.
  • This test example is used to illustrate the effect of the facial mask liquid prepared in Examples 2 to 7 on the content of malondialdehyde (MDA) in fibroblasts.
  • MDA malondialdehyde
  • Seeding Seeding fibroblasts in a 6-well plate at a seeding density of 2.2 ⁇ 10 5 cells/well, and incubating in an incubator (37° C., 5% CO 2 ) overnight.
  • UVA irradiation According to the test grouping, 30J/cm 2 of UVA irradiation was performed on the groups with UVA irradiation.
  • the cells were further cultured in an incubator (37°C, 5% CO 2 ) for 24 hours, and the cells were washed 3 times with PBS;
  • liquid phase detection collect cells, add 100 ⁇ L of NaOH (1N) to each tube to crack for 30min, measure the protein concentration of each group of samples, and finally adopt high performance liquid chromatography to detect the MDA content of the sample.
  • results of the effect of the mask liquid prepared in Examples 2 to 7 on the content of malondialdehyde (MDA) in fibroblasts are shown in Table 10. It was found that UVA irradiation would increase the content of MDA in fibroblasts. It can be seen that the facial masks prepared in all the examples can significantly reduce the content of MDA, and the facial masks prepared in Examples 2, 3, 4, 5, and 6 have a stronger effect on reducing the MDA content than the positive control VE. MDA content is an important parameter reflecting the body's anti-oxidative potential, which can reflect the lipid peroxidation rate and intensity of the body. In anti-aging research, MDA content is also a commonly used indicator. Therefore, the facial mask liquid prepared according to the present invention is obtained from the experimental results. Has anti-oxidant and anti-aging properties.

Landscapes

  • Health & Medical Sciences (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Public Health (AREA)
  • Veterinary Medicine (AREA)
  • Animal Behavior & Ethology (AREA)
  • General Health & Medical Sciences (AREA)
  • Birds (AREA)
  • Epidemiology (AREA)
  • Biotechnology (AREA)
  • Engineering & Computer Science (AREA)
  • Botany (AREA)
  • Microbiology (AREA)
  • Mycology (AREA)
  • Dermatology (AREA)
  • Chemical & Material Sciences (AREA)
  • Oil, Petroleum & Natural Gas (AREA)
  • Gerontology & Geriatric Medicine (AREA)
  • Medicines Containing Plant Substances (AREA)
  • Cosmetics (AREA)

Abstract

本发明提供了一种面膜液,所述面膜液不含任何化学添加成分,由包含以下成分的纯植物原料及水制成:菠萝、燕麦、青稞、薏仁和植物油;还提供了其制备方法和应用。本发明的面膜液除了配方成分天然以外,同时不添加香精、防腐剂等具有潜在刺激性成分。试验证明,本发明面膜液对成纤维细胞的增殖具有促进作用,具有一定的透皮吸收效果,本面膜液可显著提高胶原蛋白的含量,同时还可以显著增加成纤细胞内脂质过氧化物含量,达到抗衰老的功效。

Description

一种面膜液、其制备方法及应用
相关申请的交叉引用
本申请要求2021年04月16日提交的第CN202110413450.3号中国发明专利申请的优先权,所述申请以引用的方式整体并入本文。
技术领域
本发明属于化妆品领域,具体涉及一种面膜液,及其制备方法和应用。
背景技术
天然化妆品如今越来越受到人们的青睐。天然化妆品,除了配方成分天然以外,同时不添加香精、防腐剂等具有潜在刺激性成分。
酶解作为一种广泛应用于食品、药品及化妆品领域的技术,可将大分子成分改造成小分子,并伴随多糖、多肽等其他活性物质产生。植物中含有的酶多种多样,有蛋白酶、纤维素酶、淀粉酶、酯酶等,利用植物中含有的酶可以将大分子蛋白质酶解成寡肽,将淀粉、纤维素酶解成寡糖,将脂肪酶解成甘油、脂肪酸。将这些产物应用在化妆品中,更有利于皮肤吸收功效成分,最终达到一定的功效。
发明内容
因此,本发明的目的在于克服现有技术中的缺陷,提供一种面膜液,及其制备方法和应用。
在阐述本发明内容之前,定义本文中所使用的术语如下:
术语“MDA”是指:丙二醛。
术语“PBS”是指:磷酸缓冲盐溶液。
术语“t-test”是指:T检验。
术语“MTT”是指:细胞毒性实验。
术语“VE”是指:维生素E。
术语“VC”是指:维生素C。
为实现上述目的,本发明的第一方面提供了一种面膜液,所述面膜液不含任何化学添加成分,由包含以下成分的纯植物原料及水制成:菠萝、燕麦、青稞、薏仁和植物油;
优选地,所述植物油选自以下一种或多种:橄榄油、青刺果油、牡丹籽油、澳洲坚 果油、巴巴苏子油、椿花油;优选为橄榄油。
根据本发明第一方面的面膜液,其中,所述面膜液中各原料成分的重量份数配比为:
菠萝1~10份,燕麦0.1~10份,青稞0.5~5份,薏仁0.5~5份;
优选地,菠萝1~5份,燕麦0.2~6份,青稞0.5~3份,薏仁0.5~3份;
更优选地,菠萝1~2份,燕麦0.5~5份,青稞0.5~2份,薏仁0.5~2份。
本发明的第二方面提供了第一方面所述的面膜液的制备方法,该制备方法可以包括以下步骤:
(1)将菠萝清洗去皮切块后与水混合打浆,静置,得到提取液,即为酶液;
(2)将燕麦、青稞、薏仁进行物理粉碎过筛后,加入步骤(1)所得提取液中,加热搅拌,冷却至室温,离心取上清,得酶解处理液;
(3)向步骤(2)所得上清液中加入植物油均质,得到所述面膜液。
根据本发明第二方面的制备方法,其中,步骤(1)中,菠萝与水的比例为1:1~500g/mL,优选为1:5~500g/mL,更优选为1:10~400g/mL,进一步优选为1:50~200g/mL;和/或
所述静置时间为1~5小时,优选为2~4小时,最优选为3小时。
根据本发明第二方面的制备方法,其中,步骤(2)中,所述燕麦、青稞、薏仁粉碎后过80目筛。
根据本发明第二方面的制备方法,其中,所述步骤(2)中,所述燕麦和步骤(1)所得提取液的质量比为1~50:100,优选为1~20:100,更优选为1~10:100,进一步优选为1~5:100;
所述青稞和步骤(1)所得提取液的质量比为1~10:100,优选为1~5:100,更优选为1~3:100,进一步优选为1.25~2:100;和/或
所述薏仁和步骤(1)所得提取液的质量比为1~10:100,优选为1~5:100,更优选为1~3:100,进一步优选为1.25~2:100。
根据本发明第二方面的制备方法,其中,步骤(2)中,所述加热温度为20~150℃,优选为25~100℃,更优选为30~100℃,最优选为40℃;
所述加热时间为1~10小时,优选为1-8小时,更优选为2~6小时,最优选为4小时;
所述搅拌速率为10~1000r/min,优选为10~800r/min,更优选为10~600r/min,最优选为200r/min;和/或
所述离心转速为10~10000r/min,优选为100~8000r/min,更优选为200~6000r/min,最优选为1200r/min。
根据本发明第二方面的制备方法,其中,步骤(3)中,所述植物油的加入量为1~50%,优选为1~40%;更优选为10~30%,进一步优选为10~20%;
所述均质速度为3000~8000r/min,优选为3000~7000r/min,更优选为4000~6000r/min;进一步优优为4000~5000r/min;和/或
均质时间为1~60分钟,优选为1~50分钟,更优选为10~40分钟,进一步优选为20~30分钟。
根据本发明第二方面的制备方法,其中,所述方法进一步包括以下步骤:
(4)对步骤(3)所得面膜液进行灭菌;
优选地,所述灭菌温度为80~100℃,优选为80℃;和/或所述灭菌时间为1~5小时,优选为2小时。
本发明的第三方面提供了一种面膜,所述面膜包括第一方面所述的面膜液或根据第二方面所述方法制备的面膜液。
本发明的第四方面提供了一种面贴,所述面贴包括第一方面所述的面膜液或根据第二方面所述方法制备的面膜液。
本发明的第五方面提供了第一方面所述的面膜液或按照第二方面所述的方法制备的面膜液在制备用于抗衰老产品中的应用。
根据本发明第五方面的应用,其中,所述抗衰老为显著提高成纤细胞胶原蛋白含量。
根据本发明第五方面的应用,其中,所述抗衰老为显著增加成纤细胞内脂质过氧化物含量;
优选地,所述脂质过氧化物为丙二醛。
本发明的第六方面提供了一种用于抗衰老的方法,所述方法包括:对有需要的受试者给予:
第一方面所述的面膜液;
按照第二方面所述的方法制备的面膜液;
第三方面所述的面膜;或
第四方面所述的面贴。
根据本发明的一个具体实施方案,本发明面膜液的制备方法包括以下步骤:
(1)将菠萝与水混合打浆。室温静置3小时。
(2)将燕麦、青稞、薏仁进行物理粉碎后,加入提取液(1)中,加热搅拌,离心,取上清。
(3)(2)中加入植物油均质,得到所述面膜液。
(4)100℃ 2h灭菌,无菌条件下灌装。
步骤(1)中,所述菠萝和水的比例为1:1~500g/mL,优选为1:5~500g/mL,更优选为1:10~400g/mL,最优选为1:100g/mL;
步骤(2)中,所述燕麦和提取液(1)的比例为1:1~300g/mL,优选为1:20~200g/mL,更优选为1:20~150g/mL,最优选为1:100g/mL;
所述青稞和提取液(1)的比例为1:1~200g/mL,优选为1:10~150g/mL,更优选为1:30~100g/mL,最优选为1:80g/mL;
所述薏仁和提取液(1)的比例为1:1~200g/mL,优选为1:10~150g/mL,更优选为1:50~100g/mL,最优选为1:80g/mL;
所述加热温度为1~150℃,优选为20~100℃,更优选为30~100℃,最优选为40℃;
所述加热时间为1~10小时,优选为1-8小时,更优选为2~6小时,最优选为4小时
所述搅拌速率为10~1000r/min,优选为10~800r/min,更优选为10~600r/min,最优选为200r/min。
所述离心转速为10~10000r/min,优选为100~8000r/min,更优选为200~6000r/min,最优选为1200r/min。
步骤(3)中,所述植物油的加入量为1~50%,优选为1~40%;更优选为10~30%,最优选为20%;
所述均质速度为3000~8000r/min,优选为3000~7000r/min,更优选为4000~6000r/min;最优为5000r/min;
均质时间为1~60分钟,优选为1~50分钟,更优选为10~40分钟,最优选为30分钟。
本发明的面膜液可以具有但不限于以下有益效果:
本发明的面膜液除了配方成分天然以外,同时不添加香精、防腐剂等具有潜在刺激性成分。试验证明,本发明面膜液对成纤维细胞的增殖具有促进作用,具有一定的透皮吸收效果,本面膜液可显著提高胶原蛋白的含量,达到抗衰老的功效。
附图的简要说明
以下,结合附图来详细说明本发明的实施方案,其中:
图1示出了实施例1中220nm样品1的HPLC图谱。
图2示出了实施例1中220nm样品2的HPLC图谱。
图3示出了实施例1中220nm空白组的HPLC图谱。
图4示出了试验例1中本发明不同浓度的面膜液对成纤维细胞的毒性结果。
图5示出了试验例2中本发明面膜液的透皮吸收结果。
图6示出了试验例3中本发明面膜液对成纤维细胞内胶原蛋白含量的影响测试结果。
实施发明的最佳方式
下面通过具体的实施例进一步说明本发明,但是,应当理解为,这些实施例仅仅是用于更详细具体地说明之用,而不应理解为用于以任何形式限制本发明。实施例中未注明具体技术或条件者,按照本领域内的文献所描述的技术或条件,或者按照产品说明书进行。所用试剂或仪器未注明生产厂商者,均为可通过正规渠道商购获得的常规产品。
本部分对本发明试验中所使用到的材料以及试验方法进行一般性的描述。虽然为实现本发明目的所使用的许多材料和操作方法是本领域公知的,但是本发明仍然在此作尽可能详细描述。本领域技术人员清楚,在上下文中,如果未特别说明,本发明所用材料和操作方法是本领域公知的。
以下实施例中使用的试剂和仪器如下:
材料:
菠萝购自购自广东省湛江市红星农场;
燕麦(炒制熟)购自张家口建军燕麦食品有限公司;
青稞购自中国北京同仁堂(集团)有限责任公司;
薏仁购自中国北京同仁堂(集团)有限责任公司;
橄榄油购自云南誉成隆经贸有限公司;
VE,购自Sigma公司;
VC,I型胶原蛋白(Col I)测试盒,均购自碧云天生物技术有限公司;
成纤维细胞,购自赛百慷(上海)生物技术股份有限公司。
试剂:
MTT试剂盒,购自ScienCell有限公司;
PBS,购自北京索莱宝科技有限公司;
NaOH,购自国药集团化学试剂有限公司。
仪器:
均质机,购自IKA有限公司,型号:T25;
酶联免疫检测仪,购自Thermo Fisher Scientific OY有限公司,型号1510-00662C;
医用离心机,购自无锡市瑞江分析仪器有限公司,型号RJ-TGL-10B。
实施例1
本实施例用于说明本发明对菠萝酶液的选择。
本实验探究了菠萝作为酶液,燕麦、青稞与薏仁作为底物时,不同品种菠萝所得酶液对产物多肽分子量的影响。
实验方法为以下步骤:
(1)将不同品种的菠萝清洗去皮切块后与水按照1:1的质量比混合打浆,制得的菠萝酶液为菠萝酶液1、菠萝酶液2。其中样品1为巴里菠萝、样品2为金钻凤梨。
(2)将燕麦进行物理粉碎与水按照质量比1:25混合,得到预处理液(2)。
(3)按照酶液:底物=1:50的质量比向预处理液(2)中加入(1)。所得的混合液编号分别为样品1、样品2,其中空白组为不加入菠萝酶液的预处理液(2)。
(4)将(3)中的混合液于35℃保温1小时,后置于95℃水浴0.5小时使酶灭活。
(5)将(4)中的混合液于5000r/min离心20分钟得到上清液。
(6)利用高效液相色谱法对上清液的多肽分子量进行测定。具体测定方法如下:
(a)色谱条件
色谱柱:TsK gel 2000 SWXL 300mm×7.8mm
流动相:0.05mol/L磷酸盐缓冲液(pH 7)+0.3mol/LNaCl
检测波长:UV 220nm
流速:1ml/min
柱温:25℃
样品制备:以流动相为溶剂配制浓度为5mg/ml的样品,再在用微孔膜(0.45μm)过滤后供进样。
标准样品:将牛血清蛋白(Mr 67000)、B12(Mr 1335)、氧化型谷胱甘肽(Mr 614)配成混标,每中物质含量均为5mg/ml。
(b)标准曲线的测定
将三种标准样品按5mg/ml配制,按照上述色谱条件制作标准曲线,采用最小二乘法,求出直线的回归方程为:y=-0.3913x+7.4563,其回归系数R 2=0.9982。
由方程的回归系数R 2=0.9982可知,混合标样的标准曲线线性关系较好,可以提高计算的精确程度。关系式中X代表洗脱时间,Y代表分子量对数。这样当面膜样品进行HPLC分析时,即可通过洗脱图谱中每种物质洗脱峰的出现时间计算其对应的分子量。
(c)分子量测定结果
图1至图3分别示出了实施例1中220nm样品1、样品2和空白组的HPLC图谱。样品1、样品2和空白组的出峰时间、面积和分子量如下表1所示。
表1 样品1、样品2和空白组的出峰时间、面积和分子量
Figure PCTCN2022084840-appb-000001
Figure PCTCN2022084840-appb-000002
实验结果:空白组样品中,分子量为262946.30Da的多肽含量为6.14%,分子量为69342.58Da多肽含量为71.11%,分子量为2266.84Da的多肽含量为6.95%,分子量为1661.12Da的多肽含量为81.29%,分子量为142.01Da的多肽含量为11.76%;样品1中,分子量为1394.14Da的多肽含量为43.35%,分子量为955.95Da的多肽含量为41.72%,分子量为146.21Da的多肽含量为14.93%;样品2中,分子量为1413.49Da的多肽含量为87.19%,分子量为146.83Da的多肽含量为12.81%。结果表明,与空白组相比,加入菠萝酶液所得的上清液中,分子量为1600Da以下的多肽占比增加,尤其在样品1中,在菠萝中蛋白酶的作用下,分子量在1000以下的多肽含量为56.65%。由此说明,菠萝蛋白酶能够将燕麦中的高分子量多肽降解为较低分子量多肽,并且对比样品1、样品2的多肽分子量分布结果,最终选定后续实验中选用制备样品1所用的菠萝制备酶液。
实施例2
本实施例用于说明本发明面膜液的制备方法。
1、原料处理
菠萝:清洗、去皮、切块。
燕麦、青稞、薏仁:粉碎后过80目备用。
2、面膜液制备
(1)菠萝与水按比例为1:50g/mL混合打浆,室温静置3小时,得提取液(1)。
(2)燕麦、青稞、薏仁与提取液(1)添加比例如表2所示,形成混合液。
表2 预处理液的添加比例
原料 质量份数
提取液(1) 100
燕麦 5
青稞 2
薏仁 2
(3)将步骤(2)所得混合液70℃加热4小时,100r/min进行搅拌;冷却至室温,以转速500r/min离心,取上清。
(4)在步骤(3)所得上清液中加入上清液质量10%的橄榄油,均质机转速为4000r/min均质30分钟。
3、面膜液灭菌处理
面膜液灌装入面膜袋(含面膜布)。
80℃灭菌2小时。
实施例3
本实施例用于说明本发明面膜液的制备方法。
1、原料处理
菠萝:清洗、去皮、切块。
燕麦、青稞、薏仁:粉碎后过80目备用。
2、面膜液制备
(1)菠萝与水按照比例为1:200g/mL混合,室温静置3小时,得到提取液(1)。
(2)燕麦、青稞、薏仁与提取液(1)添加比例如表3所示,形成混合液。
表3 预处理液的添加比例
原料 质量份数
提取液(1) 100
燕麦 1
青稞 1.25
薏仁 1.25
(3)将步骤(2)所得混合液40℃加热4小时,500r/min进行搅拌;冷却至室温,以转速2000r/min离心,取上清。
(4)在步骤(3)所得上清液中加入上清液质量20%的橄榄油,均质机转速为5000r/min均质20分钟。
3、面膜液灭菌处理
面膜液灌装入面膜袋(含面膜布)。
80℃灭菌2小时。
实施例4
本实施例用于说明本发明面膜液的制备方法
1、原料处理
菠萝:清洗、去皮、切块。
燕麦、青稞、薏仁:粉碎后过80目备用。
2、面膜液制备
(1)菠萝与水按照比例为1:100g/mL混合打浆,室温静置3小时,得到提取液(1)。
(2)燕麦、青稞、薏仁与提取液(1)添加比例如表4所示,形成混合液。
表4 预处理液的添加比例
原料 质量份数
提取液(1) 100
燕麦 4
青稞 1
薏仁 1
(3)将步骤(2)所得混合液40℃加热4小时,200r/min进行搅拌;冷却至室温,以转速1200r/min离心,取上清。
(4)在步骤(3)所得上清液中加入上清液质量20%的青刺果油,均质机转速为5000r/min均质25分钟。
3、面膜液灭菌处理
面膜液灌装入面膜袋(含面膜布)。
80℃灭菌2小时。
实施例5
本实施例用于说明本发明面膜液的制备方法
1、原料处理
菠萝:清洗、去皮、切块。
燕麦、青稞、薏仁:粉碎后过80目备用。
2、面膜液制备
(1)菠萝与水按照比例为1:150g/mL混合打浆,室温静置3小时,得到提取液(1)。
(2)燕麦、青稞、薏仁与提取液(1)添加比例如表5所示,形成混合液。
表5 预处理液的添加比例
原料 质量份数
提取液(1) 100
燕麦 2
青稞 0.5
薏仁 0.5
(3)将步骤(2)所得混合液40℃加热4小时,200r/min进行搅拌;冷却至室温, 以转速1200r/min离心,取上清。
(4)在步骤(3)所得上清液中加入上清液质量10%的牡丹籽油,均质机转速为4000r/min均质30分钟。
3、面膜液灭菌处理
面膜液灌装入面膜袋(含面膜布)。
80℃灭菌2小时。
实施例6
本实施例用于说明本发明面膜液的制备方法
1、原料处理
菠萝:清洗、去皮、切块。
燕麦、青稞、薏仁:粉碎后过80目备用。
2、面膜液制备
(1)菠萝与水按照比例为1:400g/mL混合打浆,室温静置4小时,得到提取液(1)。
(2)燕麦、青稞、薏仁与提取液(1)添加比例如表6所示,形成混合液。
表6 预处理液的添加比例
原料 质量份数
提取液(1) 100
燕麦 6
青稞 3
薏仁 3
(3)将步骤(2)所得混合液80℃加热6小时,600r/min进行搅拌;冷却至室温,以转速6000r/min离心,取上清。
(4)在步骤(3)所得上清液中加入上清液质量30%的澳洲坚果油,均质机转速为6000r/min均质40分钟。
3、面膜液灭菌处理
面膜液灌装入面膜袋(含面膜布)。
90℃灭菌3小时。
实施例7
本实施例用于说明本发明面膜液的制备方法
1、原料处理
菠萝:清洗、去皮、切块。
燕麦、青稞、薏仁:粉碎后过80目备用。
2、面膜液制备
(1)菠萝与水按照比例为1:500g/mL混合打浆,室温静置5小时,得到提取液(1)。
(2)燕麦、青稞、薏仁与提取液(1)添加比例如表7所示,形成混合液。
表7 预处理液的添加比例
原料 质量份数
提取液(1) 100
燕麦 10
青稞 5
薏仁 5
(3)将步骤(2)所得混合液100℃加热1小时,800r/min进行搅拌;冷却至室温,以转速8000r/min离心,取上清。
(4)在步骤(3)所得上清液中加入上清液质量40%的椿花油,均质机转速为7000r/min均质50分钟。
3、面膜液灭菌处理
面膜液灌装入面膜袋(含面膜布)。
100℃灭菌2小时。
试验例1
本试验例用于说明实施例2制备的面膜液的对成纤维细胞的增殖的影响。
成纤维细胞毒性实验(MTT)
收集对数期细胞,调整细胞悬液浓度,每孔加入100μL(96孔板),铺板使待测细胞调密度至5000个/孔,(边缘孔用无菌PBS填充),5%CO 2,37℃孵育,至细胞生长至一定密度,换液加入不同浓度梯度的待测样品,以不含待测药品的培养液为对照。5%CO 2,37℃孵育24小时,每孔加入20μL MTT溶液(5mg/mL,即0.5%MTT四唑盐),继续培养4h。使药物与MTT充分反应,可先离心后弃去培养液,小心用PBS冲2-3遍后,再加入含MTT的培养液(FM-Fibroblast Medium购自ScienCell)。终止培养,小心吸去孔内培养液。每孔加入150μL二甲基亚砜,置摇床上低速振荡10min,使结晶物充分溶解。在酶联免疫检测仪OD值490nm处测量各孔的吸光值。
细胞活率=(测定孔OD值-空白对照OD值)/(细胞对照组OD值-空白对照OD值)*100%。
结果:对比样品不同浓度梯度的试验结果。考察不同浓度的面膜液对成纤维细胞的毒性。结果如图4所示。实验结果表明该面膜液未检出明显的细胞毒性,且在样品浓度为5%~40%范围内,细胞存活率明显增加,约为140%~180%。说明本面膜液对成纤维细胞的增殖具有促进作用。
试验例2
本试验例用于说明实施例2制备面膜液的透皮吸收结果
(1)鼠皮的制备
将裸鼠脱颈椎处死,迅速将其背部毛用剃须刀剃净,剥离背部皮肤并除去皮下脂肪,血管,用蒸馏水反复冲洗至净,再用生理盐水冲洗数遍,置-80℃冰箱中冷藏备用(在5天内用完)。
(2)Franz扩散池进行体外透皮吸收实验
实验过程:向体外渗透扩散装置恒温槽中加入适量水。开启电源和恒温槽磁力搅拌,设定恒温槽中水温为37±0.1℃。将制备好的鼠皮用铁夹子固定在两室扩散池之间,向垂直扩散池的接受池内加入5mL接受液,放入体外渗透扩散装置恒温槽中预热,设定接受池搅拌速度为400r/min。向供给池内分别加入供给液,以保鲜膜封住上口。开始记时,当样品渗透0、2、4、6、8、24小时(根据实际样品确定具体时间间隔),分别取样500μl置具塞离心管中,每次取样的同时向接受池中补充等量的接受液并排除池中的气泡。将测定样品透明质酸钠含量。再根据一下公式计算出累积透过量Q(mg/cm 2)。其中,扩散池底部直径为1.50cm,样品接触面积为1.77cm 2
Figure PCTCN2022084840-appb-000003
注:Q为累积透过量,S为透皮扩散面积,V为改进Franz扩散池接受室的体积,Cn为第n次取样时接受液的浓度,Ci为第i次取样时接受液的浓度,0.5为取样量。通过以上公式计算后,做出累积量与时间的曲线图。接受液为浓度为0.9%的NaCl溶液(生理盐水);供给液分别为待检测样品。
结果:以多糖为考察指标,测试结果如图5。在保证待测液总糖含量一致的情况下,在0-24h面膜液作用时间内,多糖透过率随作用时间延长而不断增加,在8h时,多糖透皮过程趋于稳定。说明本面膜液具有一定的透皮吸收效果。
试验例3
本试验例用于说明实施例3制备的面膜液对成纤维细胞胶原蛋白含量的影响。
本试验例中采用南京建成生物工程研究所!!HYPERLINK"http://www.njjcbio.com/products.asp?id=701"\t"/Users/lpp/Documents\\x/_blank"
Figure PCTCN2022084840-appb-000004
I型胶原蛋白(Col I)测试盒 检测成纤维细胞中胶原蛋白的含量。
(1)实验准备
a.标准品的稀释:用标准品稀释液将标准品冻干粉定容至150μl(160ng/ml),然后混匀30s。取五只干净的EP管,每管加150μl标准品稀释液,分别标记上80ng/ml,40ng/ml,20ng/ml,10ng/ml,5ng/ml。取150μl标准品原液加入标记为80ng/ml的管中, 混匀后同样取出150μl,加入下一管,以此类推至最后一管。(零孔直接加标准品稀释液)
b.生物素抗原的稀释:将浓缩型生物素抗原于6000-10000rpm离心30s,然后取生物素抗原稀释液1ml到浓缩型生物素抗原中,混匀15s至完全溶解,然后将所有液体倒入稀释液瓶中,混匀即为生物素抗原工作液。
c.亲和素-HRP的稀释:将浓缩型生物素抗原于6000-10000rpm离心30s,将浓缩亲和素-HRP全部转移到稀释液瓶中,混匀后即为亲和素-HRP工作液。
样品处理:
d.洗涤液稀释:将试剂盒中25倍浓缩液倒入500ml容量瓶中,用蒸馏水定容至500ml即为工作液。
洗板方法:先吸去酶标孔中的液体,在吸水纸上拍干,每孔加入300μl洗涤液,轻轻摇晃30s,然后甩掉,在吸水纸上拍干,重复4-5次。
(2)详细实验操作
a.使用前将试剂盒在室温下平衡半个小时
b.空白孔不加样,只加显色剂A,B和终止液用于调零。
c.标准品孔:每孔加入稀释好的标准品50μl,零孔加入标准品/样品稀释液50μl,然后加入生物素抗原工作液50μl。
d.样品孔:加入样品50μl,然后加入生物素抗原工作液50μl。
轻轻摇晃,盖上封板膜,37℃培养箱中孵育30min。
e.将25倍浓缩洗涤液用蒸馏水25倍稀释后备用。
f.第一次洗涤:小心揭掉封板膜,弃去液体,甩干,每孔加入200μl洗涤液,静置30s后弃去,如此重复5次,排干。
g.加入50μl亲和素-HRP到标准品孔和样品孔中,轻轻摇晃,盖上封板膜,37℃培养箱中孵育30min。
h.第二次洗涤:小心揭掉封板膜,弃去液体,甩干,每孔加满洗涤液,静置30s后弃去,如此重复5次,拍干。
i.显色:每孔先加入显色剂A 50μl,再加入显色剂B 50μl,轻轻震荡混匀,37℃避光显色10分钟。
j.终止:每孔加终止液50μl终止反应(此时蓝色立转黄色)
k.测定:以空白孔调零,450nm波长依次测量各孔的吸光度(OD值)。测定应在终止液后10min内进行。
l.计算:根据浓度和OD值算出标准曲线和回归方程,用专用软件进行计算,拟合模型选用logistic曲线(四参数)。标曲示意图如下表8所示:
表8 标曲示意图
浓度 Blank S5 S4 S3 S2 S1 S0
OD值 VB V5 V4 V3 V3 V1 V0
校准值 VB-VB V5-VB V4-VB V3-VB V2-VB V1-VB V0-VB
结果:面膜液对成纤维细胞内胶原蛋白含量的影响测试结果如图6。模型组是只通过UVA照射、未添加样品的细胞组,空白组为未照射且未添加样品的细胞组,阳性对照组为UVA照射后,添加质量浓度为0.1g/L的阳性对照样品VC的细胞组。
经试验发现UVA照射会严重损伤细胞的胶原蛋白,样品组与模型组、空白组和阳性对照组对比,由图可知,本面膜液可显著提高胶原蛋白的含量,达到抗衰老的功效。
试验例4
本试验例用于说明实施例2~7制备的面膜液对成纤维细胞内丙二醛(MDA)含量的影响。
测试方案如表9
表9 测试方案
Figure PCTCN2022084840-appb-000005
详细操作步骤:
(1)接种:按2.2×10 5个/孔的接种密度接种成纤维细胞至6孔板,培养箱(37℃、5%CO 2)中孵育过夜。
(2)给药:根据表9测试方案,待6孔板中细胞铺板率达到40%~60%时,进行分组给药,每孔加样2mL,每组设3个复孔。给药完成后将6孔板放置在培养箱(37℃、5%CO 2)中培养24h。
(3)UVA辐照:根据试验分组,对有UVA照射的组别进行30J/cm 2的UVA辐照。
(4)辐照结束后,培养箱(37℃、5%CO 2)中继续培养24h后,PBS清洗细胞3次;
(5)液相检测:收集细胞,每管加入100μL的NaOH(1N)裂解30min,测定每组样 品的蛋白浓度,最后采用高效液相色谱进行样品MDA含量检测。
(6)结果分析:汇总各组MDA含量,用t-test方法对进行统计分析。
结果:实施例2~7制备的面膜液对成纤维细胞内丙二醛(MDA)含量的影响结果如表10所示,经实验发现UVA的照射会增加成纤维细胞内MDA的含量,从表中可以看出,所有的实施例制备的面膜液能够显著降低MDA的含量,其中实施例2、3、4、5、6制备的面膜降低MDA含量作用强于阳性对照VE。MDA含量是反映机体抗氧化潜在能力的重要参数,可以反映机体脂质过氧化速率和强度,在抗衰老研究中,MDA含量也是一个常用指标,因此从实验结果得出根据本发明制备的面膜液具有抗氧化抗衰老的功效。
表10 成纤维细胞内丙二醛(MDA)含量变化
Figure PCTCN2022084840-appb-000006
备注:用t-test方法进行统计分析时,NC组与BC组相比,显著性以#表示,P-value<0.05表示为#,P-value<0.01表示为##;PC组、样品组与NC组相比,显著性以*表示,P-value<0.05表示为*,P-value<0.01表示为**。
尽管本发明已进行了一定程度的描述,明显地,在不脱离本发明的精神和范围的条件下,可进行各个条件的适当变化。可以理解,本发明不限于所述实施方案,而归于权利要求的范围,其包括所述每个因素的等同替换。

Claims (15)

  1. 一种面膜液,其特征在于,所述面膜液不含任何化学添加成分,由包含以下成分的纯植物原料及水制成:菠萝、燕麦、青稞、薏仁和植物油;
    优选地,所述植物油选自以下一种或多种:橄榄油、青刺果油、牡丹籽油、澳洲坚果油、巴巴苏子油、椿花油;优选为橄榄油。
  2. 根据权利要求1所述的面膜液,其特征在于,所述面膜液中各原料成分的重量份数配比为:
    菠萝1~10份,燕麦0.1~10份,青稞0.5~5份,薏仁0.5~5份;
    优选地,菠萝1~5份,燕麦0.2~6份,青稞0.5~3份,薏仁0.5~3份;
    更优选地,菠萝1~2份,燕麦0.5~5份,青稞0.5~2份,薏仁0.5~2份。
  3. 根据权利要求1或2所述的面膜液的制备方法,其特征在于,所述方法包括以下步骤:
    (1)将菠萝清洗去皮切块后与水混合打浆,静置,得到提取液;
    (2)将燕麦、青稞、薏仁进行物理粉碎过筛后,加入步骤(1)所得提取液中,加热搅拌,冷却至室温,离心取上清液;
    (3)向步骤(2)所得上清液中加入植物油均质,得到所述面膜液。
  4. 根据权利要求3所述的方法,其特征在于,步骤(1)中,菠萝与水的比例为1:1~500g/mL,优选为1:5~500g/mL,更优选为1:10~400g/mL,进一步优选为1:50~200g/mL;和/或
    所述静置时间为1~5小时,优选为2~4小时,最优选为3小时。
  5. 根据权利要求3或4所述的方法,其特征在于,步骤(2)中,所述燕麦、青稞、薏仁粉碎后过80目筛。
  6. 根据权利要求3至5中任一项所述的方法,其特征在于,所述步骤(2)中,所述燕麦和步骤(1)所得提取液的质量比为1~50:100,优选为1~20:100,更优选为1~10:100,进一步优选为1~5:100;
    所述青稞和步骤(1)所得提取液的质量比为1~10:100,优选为1~5:100,更优选为1~3:100,进一步优选为1.25~2:100;和/或
    所述薏仁和步骤(1)所得提取液的质量比为1~10:100,优选为1~5:100,更优选为1~3:100,进一步优选为1.25~2:100。
  7. 根据权利要求3至6中任一项所述的方法,其特征在于,步骤(2)中,所述加热温度为20~150℃,优选为25~100℃,更优选为30~100℃,最优选为40℃;
    所述加热时间为1~10小时,优选为1-8小时,更优选为2~6小时,最优选为4小时;
    所述搅拌速率为10~1000r/min,优选为10~800r/min,更优选为10~600r/min,最优选为200r/min;和/或
    所述离心转速为10~10000r/min,优选为100~8000r/min,更优选为200~6000r/min,最优选为1200r/min。
  8. 根据权利要求3至7中任一项所述的方法,其特征在于,步骤(3)中,所述植物油的加入量为1~50%,优选为1~40%;更优选为10~30%,进一步优选为10~20%;
    所述均质速度为3000~8000r/min,优选为3000~7000r/min,更优选为4000~6000r/min;进一步优优为4000~5000r/min;和/或
    均质时间为1~60分钟,优选为1~50分钟,更优选为10~40分钟,进一步优选为20~30分钟。
  9. 根据权利要求3至8中任一项所述的方法,其特征在于,所述方法进一步包括以下步骤:
    (4)对步骤(3)所得面膜液进行灭菌;
    优选地,所述灭菌温度为80~100℃,优选为80℃;和/或所述灭菌时间为1~5小时,优选为2小时。
  10. 一种面膜,其特征在于,所述面膜包括权利要求1或2所述的面膜液或按照权利要求4至9中任一项所述的方法制备的面膜液。
  11. 一种面贴,其特征在于,所述面贴包括权利要求1或2所述的面膜液或按照权利要求3至9中任一项所述的方法制备的面膜液。
  12. 根据权利要求1或2所述的面膜液或按照权利要求3至9中任一项所述的方法制备的面膜液在制备用于抗衰老产品中的应用。
  13. 根据权利要求12所述的应用,其特征在于,所述抗衰老为显著提高成纤细胞胶原蛋白含量。
  14. 根据权利要求12所述的应用,其特征在于,所述抗衰老为显著增加成纤细胞内脂质过氧化物含量;
    优选地,所述脂质过氧化物为丙二醛。
  15. 一种用于抗衰老的方法,其特征在于,所述方法包括:对有需要的受试者给予:
    权利要求1或2所述的面膜液;
    按照权利要求3至9中任一项所述的方法制备的面膜液;
    根据权利要求10所述的面膜;或
    根据权利要求11所述的面贴。
PCT/CN2022/084840 2021-04-16 2022-04-01 一种面膜液、其制备方法及应用 WO2022218173A1 (zh)

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
CN202110413450.3 2021-04-16
CN202110413450.3A CN113230166A (zh) 2021-04-16 2021-04-16 一种面膜液、其制备方法及应用

Publications (1)

Publication Number Publication Date
WO2022218173A1 true WO2022218173A1 (zh) 2022-10-20

Family

ID=77128420

Family Applications (1)

Application Number Title Priority Date Filing Date
PCT/CN2022/084840 WO2022218173A1 (zh) 2021-04-16 2022-04-01 一种面膜液、其制备方法及应用

Country Status (2)

Country Link
CN (1) CN113230166A (zh)
WO (1) WO2022218173A1 (zh)

Families Citing this family (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN113230166A (zh) * 2021-04-16 2021-08-10 云南白药集团股份有限公司 一种面膜液、其制备方法及应用
CN113274334B (zh) * 2021-04-16 2022-07-22 云南白药集团股份有限公司 具有抗衰老作用的菠萝提取液组合物、其制备方法和应用
CN113197818A (zh) * 2021-04-16 2021-08-03 云南白药集团股份有限公司 一种面膜液的制备方法及面膜产品
CN113274333B (zh) * 2021-04-16 2022-07-26 云南白药集团股份有限公司 具有抗氧化作用的菠萝提取液组合物、其制备方法和应用

Citations (11)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
JP2002275081A (ja) * 2001-03-21 2002-09-25 Kose Corp 皮膚外用剤
CN101953758A (zh) * 2009-07-21 2011-01-26 吉林修正药业新药开发有限公司 燕麦蛋白肽为基本成分的系列护肤品及制备方法
WO2013136174A1 (fr) * 2012-03-12 2013-09-19 Isp Investments Inc. Utilisation cosmétique d'un extrait d'avoine en tant qu'agent activateur de la caspase-14
CN104069045A (zh) * 2014-06-24 2014-10-01 樊世菊 一种祛斑防皱面膜
CN104434647A (zh) * 2014-11-17 2015-03-25 上海应用技术学院 一种含有青稞β-葡聚糖的滋养调理润肤液及其制备方法
CN106420531A (zh) * 2016-11-24 2017-02-22 青岛科技大学 一种含藏荆芥具有美白保湿功效的中药组合提取物及应用
CN108498447A (zh) * 2018-03-15 2018-09-07 广州佳静健康产业有限公司 一种植物酵素清洁泡泡液体制剂
CN108635236A (zh) * 2018-06-23 2018-10-12 无锡伟通商标代理服务有限公司 一种木瓜美肤面膜
CN113197817A (zh) * 2021-04-16 2021-08-03 云南白药集团股份有限公司 一种菠萝提取液组合物的美白应用
CN113197818A (zh) * 2021-04-16 2021-08-03 云南白药集团股份有限公司 一种面膜液的制备方法及面膜产品
CN113230166A (zh) * 2021-04-16 2021-08-10 云南白药集团股份有限公司 一种面膜液、其制备方法及应用

Family Cites Families (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
FR2834718B1 (fr) * 2002-01-15 2004-12-24 Cognis France Sa Substances actives cosmetiques et/ou pharmaceutiques
CN111110615A (zh) * 2020-01-17 2020-05-08 云南白药集团股份有限公司 一种松茸面膜液、其制备方法及应用
CN111110616A (zh) * 2020-01-17 2020-05-08 云南白药集团股份有限公司 一种松茸美白面膜液的制备方法

Patent Citations (11)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
JP2002275081A (ja) * 2001-03-21 2002-09-25 Kose Corp 皮膚外用剤
CN101953758A (zh) * 2009-07-21 2011-01-26 吉林修正药业新药开发有限公司 燕麦蛋白肽为基本成分的系列护肤品及制备方法
WO2013136174A1 (fr) * 2012-03-12 2013-09-19 Isp Investments Inc. Utilisation cosmétique d'un extrait d'avoine en tant qu'agent activateur de la caspase-14
CN104069045A (zh) * 2014-06-24 2014-10-01 樊世菊 一种祛斑防皱面膜
CN104434647A (zh) * 2014-11-17 2015-03-25 上海应用技术学院 一种含有青稞β-葡聚糖的滋养调理润肤液及其制备方法
CN106420531A (zh) * 2016-11-24 2017-02-22 青岛科技大学 一种含藏荆芥具有美白保湿功效的中药组合提取物及应用
CN108498447A (zh) * 2018-03-15 2018-09-07 广州佳静健康产业有限公司 一种植物酵素清洁泡泡液体制剂
CN108635236A (zh) * 2018-06-23 2018-10-12 无锡伟通商标代理服务有限公司 一种木瓜美肤面膜
CN113197817A (zh) * 2021-04-16 2021-08-03 云南白药集团股份有限公司 一种菠萝提取液组合物的美白应用
CN113197818A (zh) * 2021-04-16 2021-08-03 云南白药集团股份有限公司 一种面膜液的制备方法及面膜产品
CN113230166A (zh) * 2021-04-16 2021-08-10 云南白药集团股份有限公司 一种面膜液、其制备方法及应用

Non-Patent Citations (1)

* Cited by examiner, † Cited by third party
Title
CHEN MIN: "Preliminary Study on Antioxidant Activity of Oat Peptide and Its Regulatory Pathway", CHINESE MASTER'S THESES FULL-TEXT DATABASE, ENGINEERING SCIENCE I, no. 2, 15 February 2021 (2021-02-15), XP055976979 *

Also Published As

Publication number Publication date
CN113230166A (zh) 2021-08-10

Similar Documents

Publication Publication Date Title
WO2022218173A1 (zh) 一种面膜液、其制备方法及应用
WO2022218174A1 (zh) 一种面膜液的制备方法及面膜产品和应用
WO2022218172A1 (zh) 一种面膜液的制备方法及其产品
WO2022218171A1 (zh) 一种菠萝提取液组合物的美白应用
Montano et al. Vitamin content and amino acid composition of pickled garlic processed with and without fermentation
Lagreze et al. The peptides ADNF-9 and NAP increase survival and neurite outgrowth of rat retinal ganglion cells in vitro
CN113274334B (zh) 具有抗衰老作用的菠萝提取液组合物、其制备方法和应用
WO2022218169A1 (zh) 具有抗氧化作用的菠萝提取液组合物、其制备方法和应用
Li et al. A study of the physicochemical properties of rabbit glycated myofibrillary protein with high solubility in low ionic strength medium
CN110950925A (zh) 具有修复营养激活细胞功能的ace抑制肽及制备方法
CN111793140A (zh) 一种保护肝细胞氧化损伤的黑木耳寡糖及制备方法和用途
CN113397175B (zh) 一种刺参低聚肽交联微胶囊及其制备方法和应用
CN112891559B (zh) 促进牙髓干细胞增殖和成骨分化的多肽共修饰柚皮苷纳米脂质体及其制备方法和用途
Garner et al. A preliminary study of the dynamic aspects of age dependent changes in the abundances of human lens polypeptides
Li et al. Preparation and antithrombotic activity identification of Perinereis aibuhitensis extract: A high temperature and wide pH range stable biological agent
CN105400761A (zh) 一种低分子量纤溶酶及其制备方法和应用
CN109369781B (zh) 一种麒麟菜抗氧化四肽及其应用
CN106473027A (zh) 一种罗非鱼鱼皮液态发酵活性产物的制备方法
Greville et al. Observations on the fragmentation of isolated flight-muscle mitochondria from Calliphora erythrocephala (Diptera)
US20220248737A1 (en) Oyster peptide with effect of improving sexual function and preparation method thereof
CN110025499A (zh) 牡丹肽-富勒烯的制备方法
US10941432B1 (en) Method for the preparation of low molecular weight porcine lympho-reticular polypeptides
CN116370362A (zh) 白参菌-三七发酵物、其美容产品和应用
CN107779490B (zh) 一种胃蛋白酶制备苦味肽的方法及苦味肽的应用
CN105085618B (zh) 一种缢蛏活性八肽及其制备方法和应用

Legal Events

Date Code Title Description
121 Ep: the epo has been informed by wipo that ep was designated in this application

Ref document number: 22787400

Country of ref document: EP

Kind code of ref document: A1

NENP Non-entry into the national phase

Ref country code: DE

122 Ep: pct application non-entry in european phase

Ref document number: 22787400

Country of ref document: EP

Kind code of ref document: A1