CN115125153A - Preparation method and application of galactose yeast-like bacteria fermentation product filtrate - Google Patents
Preparation method and application of galactose yeast-like bacteria fermentation product filtrate Download PDFInfo
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Abstract
The invention provides a preparation method and application of a galactose yeast-like bacteria fermentation product filtrate. The invention discloses a yeast of AshbyaTrichosporon asahii) In the preparation of galactose yeast-like bacteria fermentation product filtrateThe use of (a); the galactose yeast-like bacteria fermentation product filtrate prepared by fermenting the aliskirilowii spore yeast is light yellow and transparent, has light fresh bouquet and stable physicochemical indexes, can inhibit the oxidative stress of proinflammatory cytokines by activating an antioxidant system, has excellent antioxidant and anti-inflammatory properties and the like, and can be used for producing antioxidant and anti-inflammatory products.
Description
Technical Field
The invention belongs to the technical field of microbial fermentation. More particularly, relates to a preparation method and application of a galactose yeast-like bacteria fermentation product filtrate.
Background
The skin is the largest organ of the human body and also the first barrier for the human body to directly contact with the outside, and can prevent the human body from dehydrating. The physical barrier established by combining procedures such as differentiation, induced expression and the like of epidermal cells in the skin can protect the body from external physical, chemical and biological damages, such as trauma, UVB radiation and microbial infection, and can also effectively prevent water loss in the skin, and the aging and damage of the skin barrier not only can cause skin problems such as spots, wrinkles, greasiness and the like of the skin, but also can cause potential risks to human health.
The famous material Pitera comes from sake factory, researchers find that the old brewing yeast with full face wrinkles has a young and tender hand, and this strong contrast attracts many researchers to research, and finally, a kind of bacteria is found from yeast contained in the distiller's yeastGeotrichum klebahniThe fermentation product of galactose yeast-like bacteria generated in the fermentation process of special yeast named as Zymomonia is a galactose yeast-like bacteria fermentation product generated in the fermentation process of the special yeast named as Zymomonia, contains free amino acids, mineral substances, organic acids, inorganic acids and the like which are indispensable to skin, and has multiple effects such as whitening, moisturizing, anti-aging and the like. For the production, there is a problem of degeneration of strains, and thus, the search for a wider variety of fermentation products of galacto-yeast-like bacteria is essential for the development of the field of skin care products.
The method has the advantages that a lot of bacteria can metabolize galactose and produce galactose yeast-like bacteria fermentation products in the fermentation process, and fermentation metabolism of different bacteria has obvious difference, so that the research on finding strains which can be fermented to produce safe and skin-care bacteria and have various skin-care effects and the optimization research on the fermentation process of the strains are important for the research and development of producing galactose yeast-like bacteria fermentation product filtrate.
Disclosure of Invention
Aiming at overcoming the defects of the prior art, the invention aims to provide the AshbyaTrichosporon asahii) The application in the preparation of galactose yeast-like bacteria fermentation product filtrate.
Another objective of the invention is to provide a preparation method of the galactose yeast-like bacteria fermentation product filtrate.
The invention also aims to provide the galactose yeast-like bacteria fermentation product filtrate prepared by the method.
Still another object of the present invention is to provide use of the above-mentioned galactose yeast-like bacteria fermentation product filtrate in preparing a composition for medical, cosmetic or cosmetic use.
It is a further object of the present invention to provide a composition for medical, cosmetic or cosmetic use.
The above purpose of the invention is realized by the following technical scheme:
the galactose yeast-like bacteria fermentation product filtrate prepared by fermenting the aliskirilowii sp has excellent oxidation resistance and anti-inflammatory performance by activating an oxidation resistance system to inhibit the oxidative stress of proinflammatory cytokines, so that the application of the aliskirilowii sp in preparing the galactose yeast-like bacteria fermentation product filtrate is within the protection range of the invention.
The invention also provides a leaven for preparing the galactose yeast-like bacteria fermentation product filtrate, which contains the Ashbya isattva.
Preferably, the fermentation medium also contains an Ashbya fermentation medium.
Further preferably, the fermentation medium comprises fermentation medium Y-1 and fermentation medium Y-2;
the fermentation medium Y-1 comprises the following components in percentage by mass: 1.0-2.0% of yeast extract powder, 1.5-2.5% of peptone, 1.0-4.0% of glucose, 0.5-2.0% of lactose and the balance of water;
the preparation method of the fermentation medium Y-2 comprises the following steps: sequentially carrying out enzymolysis, heating and centrifugation on the water extracts of the rice, the millet, the corn grit and the sorghum, and adding supernate obtained by centrifugation into a fermentation culture medium Y-1 to be uniformly mixed.
The composite grain consisting of rice, millet, corn grit and sorghum is subjected to water extraction and enzymolysis, a culture medium used in the fermentation process is creatively optimized, the culture medium is rich in components such as micromolecules, vitamins and various natural oligosaccharides, the fermentation efficiency, the in vivo enzyme activity and the secondary metabolite yield of the ASHISCELLA, and the content of effective active ingredients in the galactose yeast-like fermentation product filtrate are improved, and the product quality is obviously improved. The invention solves the difficult problems of slow breeding and metabolism of the ascharomyces isabellina, obviously accelerates the growth speed of thalli, simultaneously limits the symbiosis of mixed bacteria in the environment and improves the safety of fermentation products.
Further preferably, the fermentation medium Y-1 comprises the following components in percentage by mass: 1.3-1.8% of yeast extract powder, 1.8-2.2% of peptone, 1.5-3.0% of glucose, 0.5-1.5% of lactose and the balance of water.
Most preferably, the fermentation medium Y-1 comprises the following components in percentage by mass: 1.5% of yeast extract powder, 2% of peptone, 2% of glucose, 0.5-1.0% of lactose and the balance of water.
Preferably, the preparation method of the water extract comprises the following steps: adding 60-120 parts by mass of water into 1-5 parts by mass of rice, 1-5 parts by mass of millet, 1-2 parts by mass of corn grit and 1-3 parts by mass of sorghum, and extracting at 75-80 ℃ for 0.5-1.0 h to obtain the water extract.
Further preferably, the rice is 1-3 parts by mass, the millet is 1-3 parts by mass, the corn grit is 1-2 parts by mass, the sorghum is 1-2 parts by mass, and the water is 80-120 parts by mass.
Most preferably, 2 parts by mass of rice, 2 parts by mass of millet, 1 part by mass of corn grit, 2 parts by mass of sorghum, and 100 parts by mass of water.
Preferably, the enzyme used for the enzymatic hydrolysis is an amylase and/or a protease.
Further preferably, the amylase is one or more of high temperature resistant alpha-amylase, beta-amylase or isoamylase.
Further preferably, the protease is one or more of compound protease, subtilisin, papain or neutral protease.
Preferably, the final concentration of the enzyme used for enzymolysis in the water extract is 0.05-0.1 wt%.
Preferably, the enzymolysis is carried out for 2-10 h at 40-50 ℃.
Further preferably, the enzymolysis is carried out at 45 ℃ for 5-6 h.
Preferably, the heating is carried out at 70-80 ℃ for 15-25 min. Most preferably at 75 deg.C for 20 min.
Preferably, the centrifugation is performed for 10-30 min at 600-1000 rpm/min, and most preferably for 20min at 800 rpm/min.
Preferably, the mass volume ratio of the supernatant to the fermentation medium Y-1 is 50-100 g/L. Most preferably 75 g/L.
Preferably, after the blending, the sterilization is also performed.
Further preferably, the sterilization is performed at 115-125 ℃ for 15-25 min. Most preferably, sterilization is carried out at 121 ℃ for 20 min.
In addition, the invention also provides a preparation method of the galactose yeast-like bacteria fermentation product filtrate, which is obtained by fermenting the starter.
Preferably, the preparation method comprises the following steps:
s1, anaerobic fermentation: carrying out anaerobic fermentation on the Ashbya in a fermentation culture medium Y-1 to obtain a seed solution;
s2, micro-aerobic fermentation: adding the seed solution obtained in the step S1 into a fermentation culture medium Y-2, carrying out micro-aerobic fermentation, and centrifuging to obtain a galactose yeast-like bacteria fermentation product filtrate;
wherein the micro-aerobic fermentation of S2 is carried out in an environment with an oxygen content of 1.0-5.0%.
Preferably, the final concentration of the Ashbya in the fermentation medium Y-1 is 1-2 wt%.
Preferably, the temperature of the anaerobic fermentation of S1 is 25-30 ℃, the time is 48-60 h, and the pH is 4.0-6.0.
Preferably, the anaerobic fermentation in S1 is performed on a shaking table, and the rotation speed of the shaking table is 150-200 rpm/min. Most preferably 180 rpm/min.
Preferably, the final concentration of the seed liquid S2 in the fermentation medium Y-2 is 1-3 wt%.
Preferably, the temperature of the micro-aerobic fermentation of S2 is 30-35 ℃, the time is 20-28 h, and the pH is 4.5-6.0. Most preferably, the time is 24 h.
Preferably, the stirring is carried out while the micro-aerobic fermentation is carried out in S2, and the rotating speed of the stirring is 150-200 rpm/min. Most preferably 180 rpm/min.
Preferably, the micro-aerobic fermentation of S2 adopts pasteurization to stop fermentation.
Further preferably, the pasteurization temperature is 60-65 ℃ and the time is 20-30 min.
Preferably, the centrifugation of S2 is centrifugation at 1800-2200 rpm/min for 10-30 min, and most preferably centrifugation at 2000rpm/min for 20 min.
Preferably, after the centrifugation described in S2, an antioxidant is also added to the supernatant obtained by the centrifugation.
Further preferably, the antioxidant is one or more of sodium benzoate, phenoxyethanol and p-hydroxyacetophenone.
More preferably, the mass ratio of the antioxidant to the supernatant is 0.3-1.0: 100.
the galactose yeast-like bacteria fermentation product filtrate is light yellow, pure and transparent, has obvious moistening and lubricating feeling when being smeared on skin, has slight fresh wine aroma, has fresh and soft smell, lower conductivity (300-600 us/cm), and stable physicochemical indexes; the method also obviously improves the fermentation efficiency of the thalli and the content of secondary metabolites, can inhibit the oxidative stress of proinflammatory cytokines by activating an antioxidant system, and has excellent antioxidant and anti-inflammatory properties, so that the galactose yeast-like bacteria fermentation product filtrate prepared by the method and the application of the galactose yeast-like bacteria fermentation product filtrate in preparing the composition for medicine, beauty or cosmetics are all within the protection scope of the invention.
In addition, the invention also provides a medical, cosmetic or cosmetic composition, which comprises the galactose yeast fermentation product filtrate and a pharmaceutically, cosmetically or cosmetically acceptable substrate.
Preferably, the cosmetic is one or more of essence, facial mask liquid, gel, lotion, cream, eye cream and mud film.
Further preferably, the essence comprises the following components in percentage by mass: 3-7% of galactose yeast-like bacteria fermentation product filtrate, 0.2-0.6% of p-hydroxyacetophenone, 0.5-0.7% of 1, 2-hexanediol, 0.05-0.15% of 941 carbomer, 0.08-0.12% of triethanolamine, 0.01-0.03% of EDTA disodium, and the balance of water.
Most preferably, the essence comprises the following components in percentage by mass: 5% of galactose yeast-like bacteria fermentation product filtrate, 0.4% of p-hydroxyacetophenone, 0.6% of 1, 2-hexanediol, 0.1% of 941 carbomer, 0.1% of triethanolamine, 0.02% of EDTA disodium and the balance of water.
The invention has the following beneficial effects:
1. the invention provides a new choice of zymophyte for producing and preparing galactose yeast-like bacteria fermentation product filtrate, namely, the Isaria assamica. The galactose yeast-like bacteria fermentation product filtrate prepared by fermenting the aliskirilowii spore yeast can inhibit the oxidative stress of proinflammatory cytokines by activating an antioxidant system, and has excellent antioxidant and anti-inflammatory properties.
2. The invention adopts a multi-stage culture mode of anaerobic fermentation and micro-aerobic fermentation, combines a specific culture medium, not only improves the appearance and the smell of the galactose yeast-like bacteria fermentation product filtrate, leads the galactose yeast-like bacteria fermentation product filtrate to be in a light yellow transparent state with slight fresh bouquet and more stable physicochemical indexes, but also obviously improves the efficiency of thallus fermentation and the content of secondary metabolites, can inhibit the oxidative stress of proinflammatory cytokines by activating an antioxidant system, and has excellent antioxidant and anti-inflammatory properties.
3. The galactose yeast-like bacteria fermentation product filtrate has mild performance and no skin irritation, and as an active ingredient of the essence, the essence has excellent moisture retention, brightening, oil control and anti-allergy performances, and also has the effects of astringing pores, repairing skin barriers, improving skin injury, redness and swelling and the like.
Detailed Description
The present invention will be further described with reference to the following specific examples, which are not intended to limit the invention in any manner. The reagents, methods and apparatus employed in the present invention are conventional in the art, except as otherwise indicated.
Unless otherwise indicated, reagents and materials used in the following examples are commercially available.
The following examples used Saccharomyces assamissinus commercially available. Specifically, the filamentous yeast assamikayas of examples 1 to 11 and comparative examples 1 to-3 were obtained from Shandong Jiekei Biotech Co., Ltd, the filamentous yeast assamikayas of example 12 was obtained from Beijing Baiohobowenwei biotech Co., Ltd, and the filamentous yeast assamikayas of example 13 was obtained from Shanghai Braseng test biotech Co., Ltd.
EXAMPLE 1 preparation of a galactose yeast-like bacteria fermentation product filtrate
S1, anaerobic fermentation: adding the Ashbya isattva into 1000mL of fermentation medium Y-1 to ensure that the final concentration of the Ashbya isattva in the fermentation medium Y-1 is 2wt%, and then performing anaerobic fermentation shake culture for 48 hours at 30 ℃, pH 5.0 and 180rpm/min to obtain a seed solution;
s2, micro-aerobic fermentation: adding the seed solution obtained in the step S1 into a fermentation culture medium Y-2 to ensure that the final concentration of the seed solution in the fermentation culture medium Y-2 is 2wt%, carrying out micro-aerobic fermentation and stirring culture at 35 ℃, pH 5.0 and 180rpm/min for 24h, carrying out pasteurization at 65 ℃ for 20min to terminate the fermentation, centrifuging at 2000rpm/min for 20min, adding p-hydroxyacetophenone into the supernatant obtained by centrifuging (the mass ratio of the p-hydroxyacetophenone to the supernatant is 0.5: 100), and obtaining the galactose yeast sample fermentation product filtrate;
wherein the micro-aerobic fermentation is carried out in an environment with an oxygen content of 2.0%;
the fermentation medium Y-1 comprises the following components in percentage by mass: 1.5% of yeast extract powder, 2% of peptone, 2% of glucose, 0.5% of lactose and the balance of water;
the preparation method of the fermentation medium Y-2 comprises the following steps: adding 100 parts by mass of purified water into 2 parts by mass of rice, 2 parts by mass of millet, 1 part by mass of corn grit and 2 parts by mass of sorghum, extracting at 75 ℃ for 0.5h to obtain a water extract, and adding the water extract into the mixture according to a mass ratio of 1: 1: 1, the final concentration of the total enzyme dosage in the water extract is 0.1wt%, carrying out enzymolysis for 5h at 45 ℃, then heating for 20min at 75 ℃, centrifuging for 20min at 800rpm/min, finally mixing the supernatant obtained by centrifuging and a fermentation medium Y-1 according to the mass-volume ratio of 75g/L, and sterilizing for 20min at 121 ℃ to obtain the fermentation medium Y-2.
EXAMPLE 2 preparation of a galactose Yeast-like bacteria fermentation product filtrate
The preparation method is the same as that of example 1, except that:
(1) the fermentation medium Y-1 comprises the following components in percentage by mass: 1.8% of yeast extract powder, 1.8% of peptone, 3.0% of glucose, 0.5% of lactose and the balance of water;
(2) in the preparation method of the fermentation medium Y-2, the mass ratio of the rice, the millet, the corn grit, the sorghum and the purified water is 1: 3: 2: 1: 80.
EXAMPLE 3 preparation of a galactose yeast-like bacteria fermentation product filtrate
The preparation method is the same as that of the example 1, except that:
(1) the fermentation medium Y-1 comprises the following components in percentage by mass: 1.3% of yeast extract powder, 2.2% of peptone, 1.5% of glucose, 1.5% of lactose and the balance of water;
(2) in the preparation method of the fermentation medium Y-2, the mass ratio of rice, millet, corn grit, sorghum and purified water is 3: 1: 1: 2: 120.
EXAMPLE 4 preparation of a galactose yeast-like bacteria fermentation product filtrate
S1, anaerobic fermentation: adding the Ashbya into 1000mL of fermentation medium Y-1 to ensure that the final concentration of the Ashbya in the fermentation medium Y-1 is 2wt%, and then performing anaerobic fermentation shake culture for 48h at 25 ℃, pH 4.0 and 150rpm/min to obtain a seed solution;
s2, micro-aerobic fermentation: adding the seed solution obtained in the step S1 into a fermentation culture medium Y-2 to ensure that the final concentration of the seed solution in the fermentation culture medium Y-2 is 3wt%, carrying out micro-aerobic fermentation and stirring culture at 35 ℃, pH 6.0 and 150rpm/min for 28h, carrying out pasteurization at 65 ℃ for 20min to terminate the fermentation, centrifuging at 1800rpm/min for 30min, and adding phenoxyethanol into the supernatant obtained by the centrifugation (the mass-volume ratio of the phenoxyethanol to the supernatant is 1.0: 100) to obtain the galactose yeast sample fermentation product filtrate;
wherein the micro-aerobic fermentation is carried out in an environment with 1.0% of oxygen content;
the fermentation medium Y-1 comprises the following components in percentage by mass: 1.0% of yeast extract powder, 2.5% of peptone, 4.0% of glucose, 0.5% of lactose and the balance of water;
the preparation method of the fermentation medium Y-2 comprises the following steps: adding 60 parts by mass of purified water into 5 parts by mass of rice, 1 part by mass of millet, 2 parts by mass of corn grit and 1 part by mass of sorghum, extracting at 75 ℃ for 0.5h to obtain a water extract, and adding the water extract into the mixture according to a mass ratio of 1: 1, the final concentration of the total enzyme dosage in the water extract is 0.1wt%, carrying out enzymolysis for 2h at 50 ℃, then heating for 25min at 70 ℃, centrifuging for 30min at 600rpm/min, finally mixing the supernatant obtained by centrifuging and the fermentation medium Y-1 according to the mass-volume ratio of 50g/L, and sterilizing for 25min at 115 ℃ to obtain the fermentation medium Y-2.
EXAMPLE 5 preparation of a galactose yeast-like bacteria fermentation product filtrate
S1, anaerobic fermentation: adding the Ashbya into 1000mL of fermentation medium Y-1 to ensure that the final concentration of the Ashbya in the fermentation medium Y-1 is 1wt%, and then performing anaerobic fermentation shake culture for 60h at 30 ℃, pH 6.0 and 200rpm/min to obtain a seed solution;
s2, micro-aerobic fermentation: adding the seed solution obtained in the step S1 into a fermentation culture medium Y-2 to ensure that the final concentration of the seed solution in the fermentation culture medium Y-2 is 1wt%, carrying out micro-aerobic fermentation and stirring culture at 30 ℃, pH 4.5 and 200rpm/min for 20h, carrying out pasteurization at 60 ℃ for 30min to terminate the fermentation, centrifuging at 2200rpm/min for 10min, and adding sodium benzoate (ensuring that the mass ratio of the sodium benzoate to the supernatant is 0.3: 100) into the supernatant obtained by the centrifugation to obtain the fermentation product filtrate of the galacto-yeast;
wherein the micro-aerobic fermentation is carried out in an environment with an oxygen content of 5.0%;
the fermentation medium Y-1 comprises the following components in percentage by mass: 2.0% of yeast extract powder, 1.5% of peptone, 1.0% of glucose, 2.0% of lactose and the balance of water;
the preparation method of the fermentation medium Y-2 comprises the following steps: adding 120 parts by mass of purified water into 1 part by mass of rice, 5 parts by mass of millet, 1 part by mass of corn grit and 3 parts by mass of sorghum, extracting at 80 ℃ for 1.0h to obtain a water extract, and adding the water extract into the mixture according to a mass ratio of 1: 1, performing enzymolysis on the total enzyme in the water extract for 10 hours at 40 ℃ by using the alpha-amylase and the papain with the final concentration of 0.05wt%, heating the mixture for 15 minutes at 80 ℃, centrifuging the mixture for 10 minutes at 1000rpm/min, finally uniformly mixing the supernatant obtained by centrifuging and a fermentation culture medium Y-1 according to the mass-to-volume ratio of 100g/L, and sterilizing the mixture for 15 minutes at 125 ℃ to obtain the fermentation culture medium Y-2.
EXAMPLE 6 preparation of a galactose Yeast-like bacteria fermentation product filtrate
The preparation method is the same as that of example 1, except that in the preparation method of the fermentation medium Y-2, the mass ratio of rice, millet, corn grit, sorghum and purified water is 2: 1: 1: 1: 120.
EXAMPLE 7 preparation of a galactose yeast-like bacteria fermentation product filtrate
The preparation method is the same as that of example 1, except that in the preparation method of the fermentation medium Y-2, the mass ratio of rice, millet, corn grit, sorghum and purified water is 1: 1: 1: 2: 80.
EXAMPLE 8 preparation of a galactose Yeast-like bacteria fermentation product filtrate
The same procedure as in example 1, except that the fermentation medium Y-2 was prepared in such a manner that the mass-to-volume ratio of the supernatant to the fermentation medium Y-1 was 100 g/L.
EXAMPLE 9 preparation of a galactose yeast-like bacteria fermentation product filtrate
The same preparation as in example 1 except that the final concentration of the Saccharomyces assamikowis yeast in the fermentation medium Y-1 was 1wt% in S1.
EXAMPLE 10 preparation of a galactose yeast-like bacteria fermentation product filtrate
The preparation method of example 1 was repeated, except that S1 was carried out in such a manner that the final concentration of the A.assamica yeast in the fermentation medium Y-1 was 1wt%, and the anaerobic shaking culture was carried out for 60 hours.
EXAMPLE 11 preparation of a galactose yeast-like bacteria fermentation product filtrate
The preparation method of example 1 is the same, except that the micro-aerobic fermentation of S2 is performed in an environment with an oxygen content of 5.0%; and the final concentration of the seed liquid in the fermentation medium Y-2 is 3 wt%.
EXAMPLE 12 preparation of a galactose Yeast-like bacteria fermentation product filtrate
The preparation method is the same as that of example 1, except that the used Saccharomyces assamikowii is purchased from Biotech Co., Ltd. of Baiopa Bowei, Beijing.
EXAMPLE 13 preparation of a galactose yeast-like bacteria fermentation product filtrate
The preparation method was the same as that of example 1, except that the used A.assamica yeast was obtained from Shanghai Brasenia test Biotech Co., Ltd.
Comparative example 1
The preparation method was the same as that of example 1, except that the fermentation medium Y-2 used in the microaerophilic fermentation of S2 was replaced with the fermentation medium Y-1.
Comparative example 2
The preparation method is the same as that of example 1, except that the fermentation medium Y-2 is prepared without enzymolysis and heating, that is, the fermentation medium Y-2 is prepared by the following steps: adding 100 parts by mass of purified water into 2 parts by mass of rice, 2 parts by mass of millet, 1 part by mass of corn grit and 2 parts by mass of sorghum, extracting for 0.5h at 75 ℃ to obtain a water extract, cooling to 45 ℃, centrifuging for 20min at 800rpm/min, finally uniformly mixing a supernatant obtained by centrifuging and a fermentation culture medium Y-1 according to the mass-volume ratio of 75g/L, and sterilizing for 20min at 121 ℃ to obtain the fermentation culture medium Y-2.
Comparative example 3
The preparation method is the same as that of example 1, except that the micro-aerobic fermentation in S2 is replaced by anaerobic fermentation, namely S2 specifically comprises the following steps: and (3) adding the seed solution obtained in the step (S1) into a fermentation culture medium Y-2 to ensure that the final concentration of the seed solution in the fermentation culture medium Y-2 is 2wt%, carrying out anaerobic fermentation shake culture at 35 ℃, pH 5.0 and 180rpm/min for 24h, carrying out pasteurization at 65 ℃ for 20min to terminate the fermentation, carrying out centrifugation at 2000rpm/min for 20min, and adding p-hydroxyacetophenone into the supernatant obtained by the centrifugation to obtain the galactose yeast sample bacteria fermentation product filtrate.
Comparative example 4
SK-II essence (90% of Pitera included) was purchased from commercial sources.
Test example 1 physical and chemical index test of galactose yeast-like bacteria fermentation product filtrate
Test method
(1) Appearance and odor:
100g of the galactose yeast-like bacteria fermentation product filtrate obtained in examples 1 to 13 and comparative examples 1 to 3 and the essence dew of comparative example 4 SK-II were placed in a clean beaker, and the appearance was observed and the smell was smelled.
(2) Solid content test:
respectively mixing 2g (m) 0 ) The galactose yeast-like bacteria fermentation product filtrate obtained in the examples 1 to 13 and the comparative examples 1 to 3 and the SK-II essence liquid obtained in the comparative example 4 are dried at 95 ℃ for 120min, taken out and put in a drier, cooled to 25 ℃, and weighed to be m 1 (ii) a Then according to solid content = (m) 1 /m 0 ) The solid content was calculated from the equation 100%.
(3) And (3) irritation test:
the galactose yeast-like bacteria fermentation product filtrates obtained in examples 1 to 13 were subjected to a skin irritation test for a plurality of times by the method in chapter six 4 of the technical standards for cosmetic safety (2015), and an acute eye irritation test was performed by the method in chapter six 5 of the technical standards for cosmetic safety (2015).
(4) And (3) detecting the concentration and the component content of the thallus:
and (3) respectively taking the galactose yeast-like bacteria fermentation product filtrate obtained in the examples 1-13 and the comparative examples 1-3 and the SK-II essence obtained in the comparative example 4, centrifuging for 20min at the rotating speed of 4000r/min, removing supernate, collecting wet thalli, and calculating the concentration of the wet thalli in the filtrate and the contents of micromolecular polypeptide, total sugar, amino acid and organic acid in the filtrate/essence.
Second, test results
The test results are shown in tables 1 and 2.
TABLE 1
In the table, "-" indicates no measurement.
TABLE 2
As can be seen from tables 1 and 2, the galactose yeast-like bacteria fermentation product filtrate obtained in examples 1 to 13 is remarkably superior to comparative examples 1 to 4 in terms of solid content, appearance, odor, wet thallus concentration, small molecular polypeptide content, total sugar content, amino acid content, organic acid content and the like, and it can be seen that the galactose yeast-like bacteria fermentation product filtrate is light yellow and transparent by taking the Ashbya as the fermentation bacteria and adopting a multi-stage culture mode of anaerobic fermentation and micro-aerobic fermentation and combining with a specific culture medium, so that the appearance and the odor of the galactose yeast-like bacteria fermentation product filtrate are improved, the galactose yeast-like bacteria fermentation product filtrate is slightly fresh and fragrant, the physicochemical indexes are more stable, and the thallus fermentation efficiency and the content of secondary metabolites are remarkably improved.
Test example 2 efficacy test of galactose yeast-like bacteria fermentation product filtrate
Test method
(1) DPPH radical scavenging Rate test
Sample group: dissolving 2 mu L of the galactose yeast-like bacteria fermentation product filtrate obtained in the examples 1-13 and the comparative examples 1-3 and the SK-II essence liquid obtained in the comparative example 4 in 98 mu L of distilled water respectively, and adding 100 mu L of 0.1 mM DPPH solution into the system;
control group: mu.L of a 0.1 mM DPPH solution was added to 100. mu.L of distilled water.
The experiments were performed in 96-well plates, each set of three multiple wells, in a 200 μ L system. After the system is constructed, shaking for 10min in a dark place, measuring the absorbance of each group of samples at 520 nm, and determining the clearance rate = [ (% A) 0 -A x ）/A 0 ]The DPPH radical clearance is calculated by the equation x 100%, where A 0 Is a control group; a. the x Are set of samples.
(2) Inflammatory factor inhibition assay
Taking RAW264.7 macrophages as research objects, respectively determining the optimal action concentration of the galacto-yeast-like bacteria fermentation product filtrate obtained in examples 1-13 and comparative examples 1-3 and the optimal action concentration of the SK-II essence 0.5% through MTT cell proliferation experiments, respectively, establishing a cell inflammation model, respectively adding the galacto-yeast-like fermentation product fermentation essence.
Second, test results
The test results are shown in table 3.
TABLE 3
As can be seen from Table 3, the DPPH free radical clearance and the TNF-alpha inhibition rate of the galactose yeast-like bacteria fermentation product filtrate obtained in the examples 1-13 are both significantly higher than those of the comparative examples 1-4, and it can be seen that the galactose yeast-like bacteria fermentation product filtrate prepared by taking the Ashbya as the fermentation bacteria, adopting a multi-stage culture mode of anaerobic fermentation and micro-aerobic fermentation and combining a specific culture medium can inhibit the oxidative stress of proinflammatory cytokines by activating an antioxidant system, and has excellent antioxidant and anti-inflammatory properties.
In addition, taking the galactose yeast-like bacteria fermentation product filtrate obtained in example 1 as an example, the invention entrusts Guangzhou Hua vast detection service company Limited to detect the galactose yeast-like bacteria fermentation product filtrate, and the hyaluronidase inhibition rate (the sample is diluted into a 5% (v/v) aqueous solution) is 38.744 +/-4.929%, which shows that the galactose yeast-like bacteria fermentation product filtrate has excellent anti-allergy effect.
The invention also entrusts Guangdong province product quality detection Limited company to detect the galactose yeast-like bacteria fermentation product filtrate obtained in the example 1, and the 5 alpha-reductase inhibition rate of 2% (v/v) of the sample diluent is 53.21%, and the 5 alpha-reductase inhibition rate of 5% (v/v) of the sample diluent is 59.48%, which shows that the galactose yeast-like bacteria fermentation product filtrate has excellent oil control effect.
Test example 3 short-term persistence test and long-term improvement effect test of essence
The galactose yeast-like bacteria fermentation product filtrate obtained in the examples 1-7, 12 and 13 and the comparative examples 1-3 is prepared into essence according to the following components in percentage by mass: 5% of galactose yeast-like bacteria fermentation product filtrate, 0.4% of p-hydroxyacetophenone, 0.6% of 1, 2-hexanediol, 0.1% of 941 carbopol, 0.1% of triethanolamine, 0.02% of EDTA disodium and the balance of water; comparative example 4 adopts SK-II essence stock solution; the essence without the galactose yeast-like bacteria fermentation product filtrate (prepared from 0.4% of p-hydroxyacetophenone, 0.6% of 1, 2-hexanediol, 0.1% of 941 carbopol, 0.1% of triethanolamine, 0.02% of disodium EDTA, and the balance of water) was used as a blank control.
First, moisture content test
The capacitance method is adopted to measure the moisture content of the human skin horny layer, and the capacitance value of the skin is measured to be different according to the difference of the moisture content of the skin horny layer based on the obvious difference of dielectric constants of water and other substances, so that the capacitance value can represent the moisture content of the skin.
Two calibration areas are randomly selected on the inner sides of two arms of a volunteer (35-45 years old, female) to be selected (the result is averaged), the moisture content of the stratum corneum of the skin is respectively measured by adopting a capacitance method skin moisture measuring instrument, and the volunteer with the moisture content close to that of the stratum corneum before smearing is selected to carry out smearing use experiments. The final selected volunteers were grouped as shown in table 4, 15 per group.
Taking two calibration areas randomly selected from the inner sides of the two arms of the volunteer as essence applying areas, and using a latex finger stall to apply 2.0mg/cm 2 The essence obtained in examples 1 to 7, 12 and 13 and comparative examples 1 to 3 and the essence obtained in comparative example 4 were applied uniformly, and the moisture content of the horny layer of human skin was measured by a capacitance-based skin moisture meter for 0 hour, 2 hours, 4 hours and 8 hours before application and after application of the essence. The results are shown in Table 4.
Table 4 skin stratum corneum moisture content measurement results (%)
As can be seen from table 4, the water retention performance of the essence prepared by the galactose yeast-like bacteria fermentation product filtrates of examples 1 to 7, 12 and 13 is significantly better than that of comparative examples 1 to 4, and it can be seen that the essence has excellent moisture retention performance by using the aliskirilowia alismatifolia yeast as the fermentation bacteria, adopting a multi-stage culture mode of anaerobic fermentation and micro-aerobic fermentation, and combining a specific culture medium to prepare the galactose yeast-like bacteria fermentation product filtrate as the active ingredient of the essence.
Second, lasting oil control effect test
And (4) testing the lasting oil control effect of the essence by adopting a Sebumeter test method. Based on spectrophotometer's theory of operation, adopt the sticky tape to absorb the grease on human skin, the printing opacity volume of extinction sticky tape can change, and absorptive grease is more, and the printing opacity volume will be big more, and then can be through the content of printing opacity survey skin grease.
In a constant temperature and humidity room (the temperature is 21 +/-1 ℃ and the relative humidity is 50 +/-10%), the original value of the facial skin oil content of a to-be-selected volunteer (35-45 years old, female) is measured, and the volunteer with the facial skin oil content close to that before smearing is selected to carry out smearing use experiments. The final selected volunteers were grouped as shown in table 5, 15 per group.
The original value of the skin oil content of the face of the volunteer was taken as "before application" in table 5, the face of the volunteer was cleaned with a standard facial cleanser, and the initial value of the skin oil content was measured after drying (i.e., "after cleaning" in table 5). Then 1.0g of the essence obtained in examples 1-7, 12 and 13 and comparative examples 1-3 and the essence obtained in comparative example 4 were applied to the face of a volunteer, and the amount of skin oil in 2 hours, 4 hours and 8 hours after the application was measured. The results are shown in Table 5.
TABLE 5 measurement of amount of skin oil (mug/cm) 2 ）
As can be seen from table 5, the essential oil control performance of the essence liquid prepared by the galactose yeast-like bacteria fermentation product filtrates of examples 1 to 7, 12 and 13 is significantly better than that of comparative examples 1 to 4, and it can be seen that the essence liquid has excellent oil control performance by using the aliskirilowia as the fermentation bacteria, adopting a multi-stage culture mode of anaerobic fermentation and micro-aerobic fermentation, and combining a specific culture medium to prepare the galactose yeast-like bacteria fermentation product filtrate as the active ingredient of the essence liquid.
Third, long-term improvement effect test
The use effect of the essence obtained in examples 1-7, 12 and 13 and comparative examples 1-3 and the use effect of the essence obtained in comparative example 4 were subjected to a tracking test for 4 weeks.
Firstly, after cleaning the face of volunteers to be selected (35-45 years old, female), respectively, testing skin indexes (oil content, red area, water content, water loss and brightness) by using a VISIA full-face analyzer and a skin tester, selecting volunteers with close skin indexes as initial skin indexes, carrying out a smearing experiment, and finally grouping the selected volunteers according to the table 6 to obtain 15 people in each group.
The volunteers described above were massaged to absorb the essence obtained in examples 1 to 7, 12, and 13 and comparative examples 1 to 3 and the essence obtained in comparative example 4 by using the essence obtained in examples 1 to 7, 12, and 13 and comparative examples 1 to 3, respectively, 1 time each day in the morning and after cleansing at night, about 0.5mL each time, continuously used for 4 weeks (no change or use of other similar products during the test period), and after cleansing the next morning after 4 weeks ended, skin indexes (oil content, red area, water content, water loss amount, and brightness) were tested by using a VISIA full-face analyzer and a skin tester, and the obtained test results were compared with the initial skin indexes, and the corresponding change rates (oil change rate, red area change rate, water content increase rate, water loss change rate, and brightness increase rate) were as shown in table 6.
TABLE 6
As can be seen from table 6, the performance of the essence prepared by the galactose yeast-like bacteria fermentation product filtrates of examples 1 to 7, 12 and 13 in terms of oil change rate, red region change rate, water content increase rate, water loss change rate, brightness increase rate and the like is significantly better than that of comparative examples 1 to 4, and it can be seen that the galactose yeast-like bacteria fermentation product filtrate prepared by using the ashbya as the fermentation bacteria and combining a specific culture medium in an anaerobic fermentation and micro-aerobic fermentation multi-stage culture mode can be used as the active ingredient of the essence, so that the essence has excellent moisture retention and brightness performance, and can effectively repair skin barriers and improve red swelling of skin injury.
The above embodiments are preferred embodiments of the present invention, but the present invention is not limited to the above embodiments, and any other changes, modifications, substitutions, combinations, and simplifications which do not depart from the spirit and principle of the present invention should be construed as equivalents thereof, and all such changes, modifications, substitutions, combinations, and simplifications are intended to be included in the scope of the present invention.
Claims (10)
1. Saccharomyces assamici (S. assamici)Trichosporon asahii) Fermenting in the preparation of galactose yeast-like bacteriaApplication in product filtrate.
2. A starter culture for preparing a galactose yeast-like bacteria fermentation product filtrate is characterized by containing a Saccharomyces assamissimus.
3. The starter culture of claim 2, further comprising an a.
4. The starter culture of claim 3 wherein the fermentation medium comprises fermentation medium Y-1 and fermentation medium Y-2;
the fermentation medium Y-1 comprises the following components in percentage by mass: 1.0-2.0% of yeast extract powder, 1.5-2.5% of peptone, 1.0-4.0% of glucose, 0.5-2.0% of lactose and the balance of water;
the preparation method of the fermentation medium Y-2 comprises the following steps: sequentially carrying out enzymolysis, heating and centrifugation on the water extracts of the rice, the millet, the corn grit and the sorghum, and adding supernate obtained by centrifugation into a fermentation culture medium Y-1 to be uniformly mixed.
5. The starter culture according to claim 4, wherein the water extract is prepared by a method comprising: adding 60-120 parts by mass of water into 1-5 parts by mass of rice, 1-5 parts by mass of millet, 1-2 parts by mass of corn grit and 1-3 parts by mass of sorghum, and extracting at 75-80 ℃ for 0.5-1.0 h to obtain the water extract.
6. A preparation method of a galactose yeast-like bacteria fermentation product filtrate is characterized in that the galactose yeast-like bacteria fermentation product filtrate is obtained by fermentation of the starter culture according to any one of claims 3 to 5.
7. The method of claim 6, comprising the steps of:
s1, anaerobic fermentation: carrying out anaerobic fermentation on the Ashbya in a fermentation culture medium Y-1 to obtain a seed solution;
s2, micro-aerobic fermentation: adding the seed solution obtained in the step S1 into a fermentation culture medium Y-2, carrying out micro-aerobic fermentation, and centrifuging to obtain a galactose yeast-like bacteria fermentation product filtrate;
wherein the micro-aerobic fermentation of S2 is carried out in an environment with an oxygen content of 1.0-5.0%.
8. The galactose yeast-like bacteria fermentation product filtrate prepared by the method of claim 7.
9. Use of the galactoyeast-like fermentation product filtrate of claim 8 for the preparation of a composition for medical, cosmetic or cosmetic use.
10. A medical, cosmetic or cosmetic composition comprising the galactoyeast-like fermentation product filtrate of claim 8 and a pharmaceutically, cosmetically or cosmetically acceptable base.
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| CN117925731A (en) * | 2024-03-21 | 2024-04-26 | 广州华淼生物科技研究院有限公司 | Galactose yeast-like fungus centella asiatica fermentation product and preparation method and application thereof |
| CN117925731B (en) * | 2024-03-21 | 2024-06-04 | 广州华淼生物科技研究院有限公司 | Galactose yeast-like fungus centella asiatica fermentation product and preparation method and application thereof |
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