CN114480152A - Preparation method and application of vitronectin-containing yeast fermentation product filtrate composition - Google Patents
Preparation method and application of vitronectin-containing yeast fermentation product filtrate composition Download PDFInfo
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- 238000000855 fermentation Methods 0.000 title claims abstract description 81
- 230000004151 fermentation Effects 0.000 title claims abstract description 81
- 240000004808 Saccharomyces cerevisiae Species 0.000 title claims abstract description 53
- 239000000706 filtrate Substances 0.000 title claims abstract description 36
- 239000000203 mixture Substances 0.000 title claims abstract description 34
- 239000000047 product Substances 0.000 title claims abstract description 22
- 108010031318 Vitronectin Proteins 0.000 title claims abstract description 17
- 102100035140 Vitronectin Human genes 0.000 title claims abstract description 17
- 238000002360 preparation method Methods 0.000 title claims abstract description 13
- 230000000694 effects Effects 0.000 claims abstract description 13
- 235000014680 Saccharomyces cerevisiae Nutrition 0.000 claims description 51
- 239000007788 liquid Substances 0.000 claims description 32
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 claims description 30
- 230000001580 bacterial effect Effects 0.000 claims description 30
- VEXZGXHMUGYJMC-UHFFFAOYSA-N Hydrochloric acid Chemical compound Cl VEXZGXHMUGYJMC-UHFFFAOYSA-N 0.000 claims description 26
- 238000003756 stirring Methods 0.000 claims description 24
- XEKOWRVHYACXOJ-UHFFFAOYSA-N Ethyl acetate Chemical compound CCOC(C)=O XEKOWRVHYACXOJ-UHFFFAOYSA-N 0.000 claims description 21
- 238000000034 method Methods 0.000 claims description 17
- 239000001963 growth medium Substances 0.000 claims description 16
- 239000003755 preservative agent Substances 0.000 claims description 15
- TXFPEBPIARQUIG-UHFFFAOYSA-N 4'-hydroxyacetophenone Chemical compound CC(=O)C1=CC=C(O)C=C1 TXFPEBPIARQUIG-UHFFFAOYSA-N 0.000 claims description 14
- SRBFZHDQGSBBOR-IOVATXLUSA-N D-xylopyranose Chemical compound O[C@@H]1COC(O)[C@H](O)[C@H]1O SRBFZHDQGSBBOR-IOVATXLUSA-N 0.000 claims description 14
- YRKCREAYFQTBPV-UHFFFAOYSA-N acetylacetone Chemical compound CC(=O)CC(C)=O YRKCREAYFQTBPV-UHFFFAOYSA-N 0.000 claims description 12
- 230000004913 activation Effects 0.000 claims description 12
- 238000001914 filtration Methods 0.000 claims description 12
- 239000011259 mixed solution Substances 0.000 claims description 11
- 238000010438 heat treatment Methods 0.000 claims description 10
- 239000005740 Boscalid Substances 0.000 claims description 9
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- 229940118790 boscalid Drugs 0.000 claims description 9
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- 125000002915 carbonyl group Chemical group [*:2]C([*:1])=O 0.000 claims description 8
- 238000001816 cooling Methods 0.000 claims description 8
- 230000002335 preservative effect Effects 0.000 claims description 8
- 239000002994 raw material Substances 0.000 claims description 8
- 229940015975 1,2-hexanediol Drugs 0.000 claims description 7
- PYMYPHUHKUWMLA-UHFFFAOYSA-N arabinose Natural products OCC(O)C(O)C(O)C=O PYMYPHUHKUWMLA-UHFFFAOYSA-N 0.000 claims description 7
- SRBFZHDQGSBBOR-UHFFFAOYSA-N beta-D-Pyranose-Lyxose Natural products OC1COC(O)C(O)C1O SRBFZHDQGSBBOR-UHFFFAOYSA-N 0.000 claims description 7
- FHKSXSQHXQEMOK-UHFFFAOYSA-N hexane-1,2-diol Chemical compound CCCCC(O)CO FHKSXSQHXQEMOK-UHFFFAOYSA-N 0.000 claims description 7
- -1 sodium triacetoxyborohydride Chemical compound 0.000 claims description 7
- 239000012321 sodium triacetoxyborohydride Substances 0.000 claims description 7
- WQZGKKKJIJFFOK-GASJEMHNSA-N Glucose Natural products OC[C@H]1OC(O)[C@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-GASJEMHNSA-N 0.000 claims description 6
- 239000001888 Peptone Substances 0.000 claims description 6
- 108010080698 Peptones Proteins 0.000 claims description 6
- KWYUFKZDYYNOTN-UHFFFAOYSA-M Potassium hydroxide Chemical compound [OH-].[K+] KWYUFKZDYYNOTN-UHFFFAOYSA-M 0.000 claims description 6
- HEMHJVSKTPXQMS-UHFFFAOYSA-M Sodium hydroxide Chemical compound [OH-].[Na+] HEMHJVSKTPXQMS-UHFFFAOYSA-M 0.000 claims description 6
- 230000003321 amplification Effects 0.000 claims description 6
- 238000006482 condensation reaction Methods 0.000 claims description 6
- 238000007865 diluting Methods 0.000 claims description 6
- 239000008103 glucose Substances 0.000 claims description 6
- 238000009630 liquid culture Methods 0.000 claims description 6
- WRUGWIBCXHJTDG-UHFFFAOYSA-L magnesium sulfate heptahydrate Chemical compound O.O.O.O.O.O.O.[Mg+2].[O-]S([O-])(=O)=O WRUGWIBCXHJTDG-UHFFFAOYSA-L 0.000 claims description 6
- 229940061634 magnesium sulfate heptahydrate Drugs 0.000 claims description 6
- 229910000402 monopotassium phosphate Inorganic materials 0.000 claims description 6
- 235000019796 monopotassium phosphate Nutrition 0.000 claims description 6
- 238000003199 nucleic acid amplification method Methods 0.000 claims description 6
- 238000005457 optimization Methods 0.000 claims description 6
- 235000019319 peptone Nutrition 0.000 claims description 6
- PJNZPQUBCPKICU-UHFFFAOYSA-N phosphoric acid;potassium Chemical compound [K].OP(O)(O)=O PJNZPQUBCPKICU-UHFFFAOYSA-N 0.000 claims description 6
- 239000000843 powder Substances 0.000 claims description 6
- 230000001954 sterilising effect Effects 0.000 claims description 6
- 239000006228 supernatant Substances 0.000 claims description 6
- 239000006071 cream Substances 0.000 claims description 4
- 238000004659 sterilization and disinfection Methods 0.000 claims description 4
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- 229920001817 Agar Polymers 0.000 claims description 2
- 239000008272 agar Substances 0.000 claims description 2
- 238000007599 discharging Methods 0.000 claims description 2
- 239000012528 membrane Substances 0.000 claims description 2
- SESFRYSPDFLNCH-UHFFFAOYSA-N benzyl benzoate Chemical group C=1C=CC=CC=1C(=O)OCC1=CC=CC=C1 SESFRYSPDFLNCH-UHFFFAOYSA-N 0.000 claims 1
- 230000020477 pH reduction Effects 0.000 claims 1
- 210000003491 skin Anatomy 0.000 abstract description 24
- 230000003020 moisturizing effect Effects 0.000 abstract description 8
- 210000004207 dermis Anatomy 0.000 abstract description 4
- 229920002683 Glycosaminoglycan Polymers 0.000 abstract description 3
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- 230000002500 effect on skin Effects 0.000 abstract 1
- 238000004519 manufacturing process Methods 0.000 abstract 1
- HHEAADYXPMHMCT-UHFFFAOYSA-N dpph Chemical compound [O-][N+](=O)C1=CC([N+](=O)[O-])=CC([N+]([O-])=O)=C1[N]N(C=1C=CC=CC=1)C1=CC=CC=C1 HHEAADYXPMHMCT-UHFFFAOYSA-N 0.000 description 8
- 238000012545 processing Methods 0.000 description 5
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- GLEVLJDDWXEYCO-UHFFFAOYSA-N Trolox Chemical compound O1C(C)(C(O)=O)CCC2=C1C(C)=C(C)C(O)=C2C GLEVLJDDWXEYCO-UHFFFAOYSA-N 0.000 description 2
- 239000004480 active ingredient Substances 0.000 description 2
- 150000001413 amino acids Chemical class 0.000 description 2
- MGJZITXUQXWAKY-UHFFFAOYSA-N diphenyl-(2,4,6-trinitrophenyl)iminoazanium Chemical compound [O-][N+](=O)C1=CC([N+](=O)[O-])=CC([N+]([O-])=O)=C1N=[N+](C=1C=CC=CC=1)C1=CC=CC=C1 MGJZITXUQXWAKY-UHFFFAOYSA-N 0.000 description 2
- 230000001804 emulsifying effect Effects 0.000 description 2
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- 229910052500 inorganic mineral Inorganic materials 0.000 description 2
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- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Chemical compound O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 2
- DDADCBXAKYGDEH-UHFFFAOYSA-N 2-(3-hydroxypropyl)oxane-2,3,4-triol Chemical compound OCCCC1(O)OCCC(O)C1O DDADCBXAKYGDEH-UHFFFAOYSA-N 0.000 description 1
- 241000235342 Saccharomycetes Species 0.000 description 1
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- 210000001519 tissue Anatomy 0.000 description 1
- 150000003741 xylose derivatives Chemical class 0.000 description 1
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- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N1/00—Microorganisms, e.g. protozoa; Compositions thereof; Processes of propagating, maintaining or preserving microorganisms or compositions thereof; Processes of preparing or isolating a composition containing a microorganism; Culture media therefor
- C12N1/14—Fungi; Culture media therefor
- C12N1/16—Yeasts; Culture media therefor
- C12N1/18—Baker's yeast; Brewer's yeast
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K8/00—Cosmetics or similar toiletry preparations
- A61K8/18—Cosmetics or similar toiletry preparations characterised by the composition
- A61K8/30—Cosmetics or similar toiletry preparations characterised by the composition containing organic compounds
- A61K8/49—Cosmetics or similar toiletry preparations characterised by the composition containing organic compounds containing heterocyclic compounds
- A61K8/4973—Cosmetics or similar toiletry preparations characterised by the composition containing organic compounds containing heterocyclic compounds with oxygen as the only hetero atom
- A61K8/498—Cosmetics or similar toiletry preparations characterised by the composition containing organic compounds containing heterocyclic compounds with oxygen as the only hetero atom having 6-membered rings or their condensed derivatives, e.g. coumarin
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K8/00—Cosmetics or similar toiletry preparations
- A61K8/18—Cosmetics or similar toiletry preparations characterised by the composition
- A61K8/96—Cosmetics or similar toiletry preparations characterised by the composition containing materials, or derivatives thereof of undetermined constitution
- A61K8/97—Cosmetics 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
- A61K8/9728—Fungi, e.g. yeasts
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61Q—SPECIFIC USE OF COSMETICS OR SIMILAR TOILETRY PREPARATIONS
- A61Q19/00—Preparations for care of the skin
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61Q—SPECIFIC USE OF COSMETICS OR SIMILAR TOILETRY PREPARATIONS
- A61Q19/00—Preparations for care of the skin
- A61Q19/08—Anti-ageing preparations
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- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12P—FERMENTATION OR ENZYME-USING PROCESSES TO SYNTHESISE A DESIRED CHEMICAL COMPOUND OR COMPOSITION OR TO SEPARATE OPTICAL ISOMERS FROM A RACEMIC MIXTURE
- C12P1/00—Preparation of compounds or compositions, not provided for in groups C12P3/00 - C12P39/00, by using microorganisms or enzymes
- C12P1/04—Preparation of compounds or compositions, not provided for in groups C12P3/00 - C12P39/00, by using microorganisms or enzymes by using bacteria
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K2800/00—Properties of cosmetic compositions or active ingredients thereof or formulation aids used therein and process related aspects
- A61K2800/80—Process related aspects concerning the preparation of the cosmetic composition or the storage or application thereof
- A61K2800/85—Products or compounds obtained by fermentation, e.g. yoghurt, beer, wine
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- Gerontology & Geriatric Medicine (AREA)
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Abstract
The invention provides a preparation method and application of a vitronectin-containing yeast fermentation product filtrate composition. The vitreous color can indirectly increase the production of dermal collagen due to the effect of promoting the synthesis of glycosaminoglycan, achieve the effects of promoting the repair of dermis, improving the elasticity of skin and increasing the firmness of skin, can be used for improving skin, and has the effects of moisturizing and resisting aging.
Description
Technical Field
The invention relates to the technical field of microbial fermentation, chemical industry and cosmetics, in particular to a preparation method and application of a vitronectin-containing yeast fermentation product filtrate composition.
Background
The fermentation product filtrate composition obtained by yeast fermentation post-treatment contains various active ingredients such as rich amino acids, polypeptides, minerals and various vitamins, which are important components of human body and play an important role in improving skin conditions.
Vitriol, which is chemically named as hydroxypropyl tetrahydropyrane triol, is a xylose derivative with anti-aging activity and is commonly used in cosmetics. Researches show that the vitronectin can affect extracellular matrixes in three-layer structures of the skin, affect the secretion of glycosaminoglycan, promote the synthesis of glycosaminoglycan in the skin, improve the bonding degree between dermis and epidermis, promote the regeneration of damaged tissues, help to maintain the elasticity of the dermis, effectively keep the skin tight and fine and delay the aging of the skin.
The vitronectin is combined with the saccharomycete fermentation product filtrate, so that the cosmetic is rich in nutrition, replenishes various nutrients required by skin, makes the skin tender, smooth and elastic, and is a very good cosmetic raw material.
Disclosure of Invention
The technical scheme of the invention is as follows: a preparation method and application of a vitronectin-containing yeast fermentation product filtrate composition comprise the following steps:
s1, strain activation: unfreezing the frozen yeast strain, putting the unfrozen yeast strain into a culture medium, and performing activation culture under the fermentation conditions of pH value range of 5.2-6.2, fermentation temperature of 27-32 ℃, shaking table speed of 100-200 rpm and fermentation time of 12-24 hours;
S2, strain optimization: diluting the activated bacterial liquid in the step S1, and selecting a high-quality bacterial colony with strong bacterial strain growth activity from the diluted bacterial liquid in a flat-plate scribing mode;
s3, strain propagation: inoculating the high-quality bacterial colony screened in the step S2 into a liquid culture medium for amplification culture to obtain a fermented yeast liquid;
s4, in the main fermentation stage: preparing a mixed solution with the concentration of 25g/L glucose, 8.5g/L peptone, 7.5g/L yeast powder, 0.16g/L potassium dihydrogen phosphate and 0.50g/L magnesium sulfate heptahydrate, and inoculating 3-10% of the yeast liquid obtained in the step S3; fermenting for 12-24 hours under the conditions that the pH value ranges from 5.2 to 6.2, the fermentation temperature ranges from 27 to 32 ℃, and the shaking table speed ranges from 100 to 200 rpm;
s5, collecting the filtrate: after fermentation is finished, sterilizing at 90-100 ℃, discharging, cooling to room temperature, and filtering the obtained fermentation product to obtain supernatant filtrate;
s6, preparation of vitronectin: taking xylose as a raw material, carrying out condensation reaction with acetylacetone for 1h under a strong alkaline condition, acidifying by hydrochloric acid, and processing by ethanol to obtain an intermediate; reducing carbonyl by adopting milder sodium triacetoxyborohydride for 5 hours, treating by using 2mol/L hydrochloric acid, and extracting by using ethyl acetate to obtain a vitreous chromogen;
S7, adding a vitreous factor: heating the filtrate to about 45 ℃, adding 20-50% mass percent of boscalid, and stirring for dissolving;
s8, adding a preservative: and adding a preservative into the finally obtained mixed solution according to the proportion of 0.2-0.8% of the total mass of the mixed solution.
Further, the yeast in the step S1 is saccharomyces cerevisiae, and the culture medium is a bangladesh red agar culture medium from beijing land bridge.
Further, the sterilization temperature in the step S5 is 90 ℃, and the sterilization time is 1 hour.
Further, the filtrate collected in the step S5 has a polysaccharide content of 0.03-0.30% by taking the polysaccharide content as a reference standard.
Further, in step S5, the method further includes performing an activated carbon adsorption step after cooling to room temperature: adding 0.6% of activated carbon into the obtained mixed solution, and continuously stirring for 30 minutes at the rotating speed of 70 rpm; and then subsequently filtered.
Further, in the step S5, the pore size of the filter membrane or filter paper for filtration is 3 μm, 1 μm, 0.45 μm in this order after three times of filtration.
Further, as a preferred embodiment, the mixture in the step S4 is fermented in a fermentation tank with a stirring device, the stirring speed is 120-150 r/min, and the stirring time is: stirring was carried out for 30 minutes every 2 hours. And introducing sterile air into the fermentation tank, and keeping the pressure in the fermentation tank at 0.03-0.05 MPa.
Further, the preservatives added in step S8 are 1, 2-hexanediol and p-hydroxyacetophenone.
The added preservative is 1, 2-hexanediol and p-hydroxyacetophenone.
Further, the yeast fermentation product filtrate composition containing the boscalid prepared by the invention is applied to the field of preparing cosmetics; the cosmetic preparation comprises cream, lotion and aqua.
Further, in the step S6, the strong alkaline condition is controlled by sodium hydroxide or potassium hydroxide, the concentration of the hydrochloric acid is 5-10%, the ethanol is 95% medical grade ethanol, the sodium triacetoxyborohydride is from the far infrared chemical industry, the ethyl acetate is analytically pure, and the xylose is a conventional market raw material.
The invention applies the microbial fermentation technology, and the fermented product filtrate is rich in various fermentation active ingredients such as amino acid, polypeptide, mineral substances, various vitamins and the like, and is added with the vitronectin so as to help maintain the elasticity of dermis, effectively keep the skin tight and delicate and delay the aging of the skin.
Drawings
FIG. 1 is a method for preparing a vitronectin-containing yeast fermentation product filtrate composition and applications thereof.
FIG. 2 is an instantaneous moisture test chart of skin after example 3 is applied to the skin.
FIG. 3 is a graph of the DPPH radical scavenging test results for compositions prepared in example 3.
Detailed Description
The present invention will be described in further detail with reference to the drawings and examples. It should be understood that the specific embodiments described herein are merely illustrative of the invention and do not limit the invention.
Example 1
As shown in fig. 1, a method for preparing a vitronectin-containing yeast fermentation product filtrate composition comprises the following steps:
step S1, strain activation: unfreezing the frozen yeast strain, putting the unfrozen yeast strain into a culture medium, and performing activation culture under the fermentation conditions of pH value range of 5.6, fermentation temperature of 30 ℃, shaking table speed of 130rpm and fermentation time of 12 hours.
Step S2, strain optimization: diluting the activated bacterial liquid in the step S1, and selecting a high-quality bacterial colony with strong bacterial strain growth activity from the bacterial liquid in a plate scribing mode.
Step S3, expanding culture of strains: and (4) inoculating the high-quality bacterial colony screened in the step (S2) into a liquid culture medium for amplification culture to obtain the yeast liquid after fermentation.
Step S4, the main fermentation stage: preparing a mixed solution with the concentration of 25g/L glucose, 8.5g/L peptone, 7.5g/L yeast powder, 0.16g/L potassium dihydrogen phosphate and 0.5g/L magnesium sulfate heptahydrate, and inoculating the yeast liquid which is obtained in the step S3 and accounts for 5 percent of the total weight of the composition; fermenting for 24 hours under the fermentation conditions that the pH value range is 5.2-6.2, the fermentation temperature is 30 ℃ and the shaking table speed is 130 rpm.
Step S5, collecting the filtrate: after fermentation, the fermentation liquid is sterilized at 90 ℃ and discharged, the fermentation liquid is cooled to room temperature, and the supernatant is obtained by filtering for three times by using filter paper with the aperture of 3um, 1um and 0.45um in sequence.
Step S6, preparing a vitreous silica: taking xylose as a raw material, carrying out condensation reaction with acetylacetone for 1h under a strong alkaline condition, acidifying by hydrochloric acid, and processing by ethanol to obtain an intermediate; reducing carbonyl for 5h by adopting milder sodium triacetoxyborohydride, treating by using 2mol/L hydrochloric acid, and extracting by using ethyl acetate to obtain vitreous chromogen;
step S7, adding a vitreous color factor: heating the filtrate to about 45 ℃, adding 30 mass percent of boscalid, stirring and dissolving.
Step S8, adding a preservative: 0.2% of 1, 2-hexanediol and 0.7% of p-hydroxyacetophenone were added to the finally obtained mixture in terms of mass fraction as preservatives.
Example 2
Referring to fig. 1, a method for preparing a vitronectin-containing yeast fermentation product filtrate composition includes the steps of:
step S1, strain activation: unfreezing the frozen yeast strain, putting the unfrozen yeast strain into a culture medium, and performing activation culture under the fermentation conditions of pH value range of 5.8, fermentation temperature of 31 ℃, shaking table speed of 120rpm and fermentation time of 14 hours.
Step S2, strain optimization: diluting the activated bacterial liquid in the step S1, and selecting a high-quality bacterial colony with strong bacterial strain growth activity from the bacterial colony in a flat-plate streaking mode.
Step S3, strain propagation: and (4) inoculating the high-quality bacterial colony screened in the step S2 into a liquid culture medium for amplification culture to obtain the fermented yeast liquid.
Step S4, fermentation stage: preparing a mixed solution with the concentration of 25g/L glucose, 8.5g/L peptone, 7.5g/L yeast powder, 0.16g/L potassium dihydrogen phosphate and 0.5g/L magnesium sulfate heptahydrate, and inoculating the yeast liquid which is obtained in the step S3 and accounts for 4 percent of the total weight of the composition; fermenting for 18 hours under the fermentation conditions that the pH value range is 5.5-6.0, the fermentation temperature is 31 ℃ and the shaking table speed is 120 rpm.
Step S5, collecting the filtrate: after fermentation, the fermentation liquid is sterilized at 90 ℃ and discharged, the fermentation liquid is cooled to room temperature, and the supernatant is obtained by filtering for three times by using filter paper with the aperture of 3um, 1um and 0.45um in sequence.
Step S6, preparing a vitreous silica: taking xylose as a raw material, carrying out condensation reaction with acetylacetone for 1.5h under a strong alkaline condition, acidifying by hydrochloric acid, and processing by ethanol to obtain an intermediate; then reducing carbonyl by adopting milder sodium triacetoxyborohydride for 6 hours, treating by 1.5mol/L hydrochloric acid, and extracting by ethyl acetate to obtain the vitreous chromogen.
Step S7, adding a vitreous color factor: heating the filtrate to about 45 ℃, adding 40% of vitreous color factor by mass, stirring and dissolving;
step S8, adding a preservative: 0.3% of 1, 2-hexanediol and 0.5% of p-hydroxyacetophenone were added to the finally obtained mixture in terms of mass fraction as preservatives.
Example 3
The starting materials were prepared as in example 1. The preparation method comprises the following steps:
step S1, strain activation: unfreezing the frozen yeast strain, putting the unfrozen yeast strain into a culture medium, and performing activation culture under the fermentation conditions of pH value range of 6.0, fermentation temperature of 31 ℃, shaking table speed of 120rpm and fermentation time of 12 hours.
Step S2, strain optimization: diluting the activated bacterial liquid in the step S1, and selecting a high-quality bacterial colony with strong bacterial strain growth activity from the bacterial liquid in a plate scribing mode.
Step S3, expanding culture of strains: and (4) inoculating the high-quality bacterial colony screened in the step (S2) into a liquid culture medium for amplification culture to obtain the yeast liquid after fermentation.
Step S4, the main fermentation stage: preparing a mixed solution with the concentration of 20g/L glucose, 6.5g/L peptone, 6.5g/L yeast powder, 0.15g/L potassium dihydrogen phosphate and 0.45g/L magnesium sulfate heptahydrate, and inoculating the yeast liquid which is obtained in the step S3 and accounts for 8 percent of the total weight of the composition; fermenting for 24 hours under the fermentation conditions that the pH value range is 5.2-6.0, the fermentation temperature is 29 ℃ and the shaking table speed is 120 rpm.
Step S5, collecting the filtrate: after fermentation, the fermentation liquid is sterilized at 90 ℃ and discharged, the fermentation liquid is cooled to room temperature, and the supernatant is obtained by filtering for three times by using filter paper with the aperture of 3um, 1um and 0.45um in sequence.
Step S6, preparing a vitreous silica: taking xylose as a raw material, carrying out condensation reaction with acetylacetone for 2h under a strong alkaline condition, acidifying by hydrochloric acid, and processing by ethanol to obtain an intermediate; then reducing the carbonyl group by adopting milder sodium triacetoxyborohydride for 4h, treating by 3mol/L hydrochloric acid, and extracting by ethyl acetate to obtain the vitreous chromogen.
Step S7, adding a vitreous color factor: heating the filtrate to about 45 ℃, adding 35% mass fraction of vitreous color factor, and stirring for dissolving;
step S8, adding a preservative: 0.5% of 1, 2-hexanediol and 0.4% of p-hydroxyacetophenone were added to the finally obtained mixture in terms of mass fraction as preservatives.
Example 4
The starting materials were prepared as in example 1. The preparation method comprises the following steps:
step S1, strain activation: unfreezing the frozen yeast strain, putting the unfrozen yeast strain into a culture medium, and performing activation culture under the fermentation conditions of pH value range of 5.8, fermentation temperature of 28 ℃, shaking table speed of 140rpm and fermentation time of 16 hours.
Step S2, strain optimization: diluting the activated bacterial liquid in the step S1, and selecting a high-quality bacterial colony with strong bacterial strain growth activity from the bacterial liquid in a plate scribing mode.
Step S3, strain propagation: and (4) inoculating the high-quality bacterial colony screened in the step S2 into a liquid culture medium for amplification culture to obtain the fermented yeast liquid.
Step S4, fermentation stage: preparing a mixed solution with the concentration of 20g/L glucose, 7.5g/L peptone, 7.5g/L yeast powder, 0.13g/L potassium dihydrogen phosphate and 0.6g/L magnesium sulfate heptahydrate, and inoculating the yeast liquid which is obtained in the step S3 and accounts for 5 percent of the total weight of the composition; fermenting for 24 hours under the fermentation conditions that the pH value range is 5.5-6.0, the fermentation temperature is 28 ℃ and the shaking table speed is 140 rpm.
Step S5, collecting the filtrate: after fermentation, the fermentation liquid is sterilized at 90 ℃ and discharged, the fermentation liquid is cooled to room temperature, and the supernatant is obtained by filtering for three times by using filter paper with the aperture of 3um, 1um and 0.45um in sequence.
Step S6, preparing a vitreous silica: taking xylose as a raw material, carrying out condensation reaction with acetylacetone for 1h under a strong alkaline condition, acidifying by hydrochloric acid, and processing by ethanol to obtain an intermediate; then reducing carbonyl by adopting milder sodium triacetoxyborohydride for 5h, treating by using 2mol/L hydrochloric acid, and extracting by using ethyl acetate to obtain the vitreous chromogen.
Step S7, adding: heating the filtrate to about 45 ℃, adding 30% mass fraction of vitronectin, and stirring to dissolve.
Step S8, adding a preservative: 0.8% of 1, 2-hexanediol and 0.3% of p-hydroxyacetophenone were added to the finally obtained mixture in terms of mass fraction as preservatives.
And (3) displaying the effects:
evaluation of skin moisturizing and improving efficacy:
in order to measure the immediate moisturizing and improving effect of the skin, the composition of example 3 was randomly applied to the skin, and then the moisture content of the skin was observed by Epsilon.
Specifically, the composition of example 3 was applied to the arms of 5 subjects, and the skin moisture content was measured, using pure distilled water as a control.
FIG. 2 shows the results of measurement of the skin moisture improving effect of example 3 applied to the skin. As shown in fig. 2, the moisture content of the skin immediately after the composition of example 3 was applied to the skin increased by 1.5 times. The skin moisture content was significantly increased compared to the control group.
The experimental result shows that the cosmetic composition has an instant moisturizing effect.
DPPH radical scavenging test
First, reagent preparation
(1)3.943mgDPPH was made up to 50mL with 95% ethanol
(2)10mgTrolox is fixed to 100mL by 95% ethanol
(3) The composition of the present invention (hereinafter, referred to as example 3)
Second, the experimental procedure
(1) UV zero calibration, 95% ethanol as blank
(2) Preliminary experiments (DPPH concentration selection)
Shaking 500 μ L DPPH +1000 μ L95% ethanol for 10s, standing for 10min, and measuring absorbance (absorbance at 520nm of 0.45-0.55, can be used as DPPH solution for the following experiment)
(3) Determination of Trolox
| Sample | Trolox(μL) | 95% ethanol (μ L) | DPPH(μL) |
| |
100 | 900 | 500 |
| |
100 | 900+500 | 0 |
| |
0 | 100+900 | 500 |
| |
0 | 100+900+500 | 0 |
Adding DPPH or 95% ethanol, shaking for 10s, standing for 10min, and measuring absorbance
(4) Experimental procedure
Adding DPPH or 95% ethanol, shaking for 10s, and standing for 10min
(5) The absorbance values were measured at 520nm, tabulated and plotted to obtain IC50 values.
Calculating the formula: inhibition rate (%) [1- (S-SB)/(C-CB) ]. 100
Third, experimental results
| Inner(μL) | Con(μL/mL) | DPPH(％) |
| 150 | 100 | 77.841 |
| 50 | 33.33 | 54.187 |
| 20 | 13.3 | 32.778 |
The experimental results are as follows: example 3 has the effect of scavenging DPPH free radicals and has an IC50 value of about 31. mu.L/mL.
Application examples
(1) Moisturizing and anti-aging essence lotion
The anti-inflammatory moisturizing essence water process comprises the following steps:
heating and dissolving A phase at 80 ℃, stirring for 40 minutes, and then cooling
2. Adding phase B into phase A, stirring and mixing uniformly
3. Cooling to 60 deg.C, sequentially adding C, D, and stirring
4. Stirring, cooling to 40 deg.C, adding phase D, stirring for 5min, sampling, testing, filtering with 250 mesh gauze, and packaging
(2) Moisturizing and anti-aging essence emulsion
The anti-inflammatory repair emulsion process comprises the following steps:
dissolving phase A at 80 deg.C for 20min
Heating and stirring the B phase at 80 ℃ for dissolving for 30min
3, after the A phase and the B phase are completely dissolved, adding the A phase into the B phase, emulsifying for 3min after 5500 revolutions
4. And after sampling and detecting, stirring, cooling to 40 ℃, adding the phase D, filtering with 250-mesh gauze, and packaging.
(3) Moisturizing and anti-aging essence cream
The anti-inflammatory repair cream process comprises the following steps:
dissolving phase A by heating at 80 deg.C
Dissolving B phase in 80 deg.C under heating and stirring
Adding phase B into phase A, emulsifying for 3min at 5000r/min
4. Cooling to about 50 deg.C, sequentially adding C, D phases, stirring for dissolving, and mixing
The foregoing description describes preferred embodiments of the invention, and, as noted above, it is to be understood that the invention is not limited to the forms disclosed herein, but is not intended to be exhaustive or to be construed as other embodiments, and is capable of use in various other combinations, modifications, and environments and is capable of changes within the scope of the inventive concept as described herein, commensurate with the above teachings, or the skill or knowledge of the relevant art. And that modifications and variations may be effected by those skilled in the art without departing from the spirit and scope of the invention as defined by the appended claims.
Claims (9)
1.A method for preparing a vitronectin-containing yeast fermentation product filtrate composition is characterized by comprising the following steps of: the method comprises the following steps:
s1, strain activation: unfreezing the frozen yeast strain, putting the unfrozen yeast strain into a culture medium, and performing activation culture under the fermentation conditions of pH value range of 5.2-6.2, fermentation temperature of 27-32 ℃, shaking table speed of 100-200 rpm and fermentation time of 12-24 hours;
S2, strain optimization: diluting the activated bacterial liquid in the step S1, and selecting a high-quality bacterial colony with strong bacterial strain growth activity from the diluted bacterial liquid in a flat-plate scribing mode;
s3, strain propagation: inoculating the high-quality bacterial colony screened in the step S2 into a liquid culture medium for amplification culture to obtain a fermented yeast liquid;
s4, a formal fermentation stage: preparing a mixed solution with the concentration of 25g/L glucose, 8.5g/L peptone, 7.5g/L yeast powder, 0.16g/L potassium dihydrogen phosphate and 0.50g/L magnesium sulfate heptahydrate, and inoculating 3-10% of the yeast liquid obtained in the step S3; fermenting for 12-24 hours under the conditions that the pH value ranges from 5.2 to 6.2, the fermentation temperature ranges from 27 to 32 ℃, and the shaking table speed ranges from 100 to 200 rpm;
s5, collecting the filtrate: after fermentation is finished, sterilizing at 90-100 ℃, discharging, cooling to room temperature, and filtering the obtained fermentation product to obtain supernatant filtrate;
s6, preparation of vitronectin: taking xylose as a raw material, carrying out condensation reaction with acetylacetone for 1h under a strong alkaline condition, and carrying out acidification with hydrochloric acid and ethanol treatment to obtain an intermediate; reducing the carbonyl group by adopting milder sodium triacetoxyborohydride for 5 hours, treating the reduced carbonyl group by using 2mol/L hydrochloric acid, and extracting the treated carbonyl group by using ethyl acetate to obtain a vitreous chromogen;
S7, adding boscalid: heating the filtrate to about 45 ℃, adding 20-50% of the filtrate by mass, and stirring for dissolving;
s8, adding preservatives: and adding a preservative into the finally obtained mixed solution according to the proportion of 0.2-0.8% of the total mass of the mixed solution.
2. The method of claim 1 for preparing a boscalid-containing yeast fermentation product filtrate composition, wherein: the yeast in the step S1 is Saccharomyces cerevisiae, and the culture medium is Bengal agar culture medium from Beijing Luqiao.
3. The method of claim 1 for preparing a boscalid-containing yeast fermentation product filtrate composition, wherein: the sterilization temperature in step S5 was 90 ℃, and the sterilization time was 1 hour.
4. The method of claim 1 for preparing a boscalid-containing yeast fermentation product filtrate composition, wherein: in the step S4, the added yeast liquid is 3%, the fermentation temperature is 28 ℃, and the fermentation time is 24 hours.
5. The method of claim 1 for preparing a boscalid-containing yeast fermentation product filtrate composition, wherein: and in the step S5, filtering is carried out for three times, and the aperture of a filter membrane or filter paper for filtering is 3 mu m, 1 mu m and 0.45 mu m in sequence.
6. The method of claim 1, wherein the method comprises the steps of: and in the step S4, fermenting the mixture in a fermentation tank with a stirring device, wherein the stirring speed is 120-150 r/min, and the stirring time is as follows: stirring was carried out for 30 minutes every 2 hours.
7. The method of claim 6, wherein the method comprises the steps of: the strong alkaline condition is controlled by sodium hydroxide or potassium hydroxide.
8. The method of claim 1 for preparing a boscalid-containing yeast fermentation product filtrate composition, wherein: the preservatives added in the step S8 are 1, 2-hexanediol and p-hydroxyacetophenone.
9. The application of a vitronectin-containing yeast fermentation product filtrate composition in cosmetics is characterized in that: the vitronectin-containing yeast fermentation product filtrate composition is prepared by the preparation method of any one of claims 1 to 8; the cosmetic preparation comprises cream, lotion and aqua.
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