CN113750018A - Plant composition and preparation method and application thereof - Google Patents

Plant composition and preparation method and application thereof Download PDF

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Publication number
CN113750018A
CN113750018A CN202110433276.9A CN202110433276A CN113750018A CN 113750018 A CN113750018 A CN 113750018A CN 202110433276 A CN202110433276 A CN 202110433276A CN 113750018 A CN113750018 A CN 113750018A
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culture medium
ganoderma lucidum
fermentation
plant composition
bidirectional
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CN113750018B (en
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邱显荣
刘有停
吕永博
苏牧楠
杨素珍
李燕
韩婷婷
王晓娜
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Shandong Furida Biological Co ltd
Nutri Woods Bio Tech Beijing Co ltd
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Shandong Furida Biological Co ltd
Nutri Woods Bio Tech Beijing Co ltd
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    • 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
    • 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
    • A61K8/9728Fungi, e.g. yeasts
    • 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
    • A61QSPECIFIC USE OF COSMETICS OR SIMILAR TOILETRY PREPARATIONS
    • A61Q19/00Preparations for care of the skin
    • A61Q19/02Preparations for care of the skin for chemically bleaching or whitening 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
    • A61K2800/40Chemical, 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
    • 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
    • A61K2800/80Process related aspects concerning the preparation of the cosmetic composition or the storage or application thereof
    • A61K2800/85Products or compounds obtained by fermentation, e.g. yoghurt, beer, wine

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  • Cosmetics (AREA)

Abstract

The invention discloses a plant composition and a preparation method and application thereof, and belongs to the field of plant extraction research. The plant composition comprises dendrobium officinale, rhizoma polygonati, tuberose and lucid ganoderma and is prepared by a bidirectional fermentation process. Compared with the traditional water extraction method, the plant composition prepared by the bidirectional fermentation process has high content of active ingredient polysaccharide. The plant composition achieves a good anti-aging effect by inhibiting non-enzymatic glycosylation and inhibiting clock gene expression, achieves a good whitening effect by inhibiting tyrosinase activity, and can achieve double effects of anti-aging and whitening by adding the plant composition into cosmetics and/or skin care products.

Description

Plant composition and preparation method and application thereof
Technical Field
The invention relates to the field of plant extraction research, and particularly relates to a plant composition and a preparation method and application thereof.
Background
The bidirectional fermentation of Chinese medicine is a new high-tech Chinese medicine preparing technology combining modern microbial engineering with the research result of microecology. The 'bidirectional fermentation' is characterized in that traditional Chinese medicinal materials or herb residues with certain active ingredients are used as a medicinal substrate to replace a traditional nutrient substrate, and optimized strains are added into the medicinal substrate for microbial transformation, so that a fermented composition formed by the medicinal materials or herb residues is called medicinal mycoplasm. The bidirectional property is realized in that the medicinal matrix provides nutrition required by fungi, and simultaneously is influenced by enzymes in the fungi to change self tissues and components to generate new flavor functions. Therefore, the microbial fermentation of the traditional Chinese medicine is environment-friendly, high in extraction efficiency and good in effect, can be applied to multiple industries such as food and medicines, but is less in application in cosmetics, can provide a way for expanding new components and new effects in the field of cosmetic raw materials, and has a very wide application prospect.
The patent (CN 107898931A) discloses a regulator of clock gene expression and a preparation method thereof, wherein the regulator is a dendrobium extract. The dendrobium extract is found to promote the expression of Clock gene for the first time. However, studies (Sun, BMAL1/CLOCK regulation of keratinocyte apoptosis, DNA damage and immunity under UVB irradiation [ D ]. Tianjin university, 2018.) show that (i) under normal culture conditions without UV irradiation, small RNA-mediated silencing of CLOCK gene can promote keratinocyte proliferation and induce differentiation of primary keratinocytes; secondly, the silencing of the Clock gene can prevent the apoptosis of primary keratinocytes and HaCat cells caused by UVB; thirdly, in keratinocytes, silencing Clock can effectively inhibit the high expression of the cell factor caused by UVB; in primary keratinocytes, Clock gene silencing strongly inhibited the increase in total p53 protein and phosphorylation caused by UVB. In conclusion, the promotion of Clock can cause adverse reaction on skin, inhibit the expression of Clock gene, and effectively reduce keratinocyte apoptosis and resist the adverse reaction of UVB on organisms.
Aiming at the fact that the single dendrobium extract promotes the expression of Clock gene and the adverse reaction on skin, it is necessary to develop a plant composition which has high active ingredient content, can inhibit non-enzymatic glycosylation reaction, inhibit the expression of Clock gene and inhibit the activity of tyrosinase, and has the effects of resisting aging and whitening skin.
Disclosure of Invention
Based on the defects of the prior art, the invention provides the plant composition which has high active matter content, can inhibit non-enzymatic glycosylation reaction, inhibit clock gene expression and inhibit tyrosinase activity, and has the effects of resisting aging and whitening.
The technical problem to be solved by the invention is realized by the following technical scheme:
in a first aspect, the present invention provides a plant composition comprising the following components: dendrobium officinale, rhizoma polygonati, tuberose and lucid ganoderma.
According to the invention, the plant composition is prepared by a bidirectional fermentation process.
According to the invention, the addition amount of dendrobium officinale in the plant composition is 1-10% by weight of the bidirectional fermentation medium.
According to the invention, the addition amount of rhizoma polygonati in the plant composition is 0.01-1% by weight of the bidirectional fermentation medium.
According to the invention, the tuberose is added into the plant composition in an amount of 0.01-1% by weight of the bidirectional fermentation medium.
According to the invention, the plant composition contains 5.11-5.98mg/mL of polysaccharide and 0.11-0.23mg/mL of flavone.
According to some embodiments of the invention, the plant composition has a polysaccharide content of 5.23 to 5.98mg/mL and a flavone content of 0.13 to 0.23 mg/mL.
According to the present invention, the plant composition is prepared by a preparation method comprising:
pretreating raw materials: pulverizing herba Dendrobii, rhizoma Polygonati, and tuberose respectively, and sieving;
② activating ganoderma lucidum: preparing a ganoderma lucidum culture medium, inoculating ganoderma lucidum, and culturing to obtain ganoderma lucidum seed liquid;
preparing a bidirectional fermentation culture medium;
inoculating and fermenting: inoculating the ganoderma seed liquid into a bidirectional fermentation culture medium for culture;
fifthly, post-treatment of fermentation liquor: sterilizing and filtering.
According to the invention, the culture medium for activating the ganoderma lucidum in the step II of the plant combination preparation method is 0.5-5% of starch, 0.1-5% of yeast powder, 0.1-2% of sodium chloride, 0.01-0.1% of calcium carbonate and water added to 100%.
According to the invention, the culture temperature of the ganoderma lucidum is 25-35 ℃.
According to the invention, the culture speed of the ganoderma lucidum is 50-300 rpm.
According to the invention, the culture time of the ganoderma lucidum is 50-80 h.
According to the invention, in the third step of the plant combination preparation method, the bidirectional fermentation medium is 0.5-5% of starch, 0.1-5% of yeast powder, 0.1-2% of sodium chloride, 0.01-0.1% of calcium carbonate, 1-10% of dendrobium officinale, 0.01-1% of rhizoma polygonati, 0.01-1% of tuberose and water to 100%.
According to some embodiments of the present invention, in the third step of the preparation method of the plant composition, the bidirectional fermentation medium is 0.5 to 5% of starch, 0.1 to 5% of yeast powder, 0.1 to 2% of sodium chloride, 0.01 to 0.1% of calcium carbonate, 1 to 2% of dendrobium officinale, 0.01 to 0.2% of rhizoma polygonati, 0.01 to 0.2% of tuberose, and water to 100%.
According to the invention, the temperature of the bidirectional fermentation culture in the third step is 25-35 ℃.
According to the invention, the rotation speed of the bidirectional fermentation culture in the third step is 50-300 rpm.
According to the invention, the bidirectional fermentation culture time in the third step is 1-7 d.
According to the invention, the pH value of the bidirectional fermentation culture in the third step is 3.5-7.5.
According to some embodiments of the present invention, the temperature of the bidirectional fermentation culture in the third step is 30 ℃.
According to some embodiments of the present invention, the rotation speed of the bidirectional fermentation culture in the third step is 150 rpm.
According to some embodiments of the present invention, the time for the bidirectional fermentation culture in step (iii) is 3 d.
According to some embodiments of the present invention, the pH value of the bidirectional fermentation culture in the third step is 5.5.
According to the invention, the inoculation amount of the ganoderma lucidum seed liquid in the step (iv) in the plant combination preparation method is 1-15%.
According to some embodiments of the present invention, the inoculation amount of the ganoderma lucidum seed solution in the fourth step of the plant combination preparation method is 10%.
According to the invention, the sterilization mode in the step (v) of the plant combination preparation method is a conventional sterilization mode in the field.
According to the invention, the sterilization mode in the fifth step of the plant combination preparation method is high-temperature sterilization.
According to the invention, the high temperature sterilization condition in the step (c) of the plant combination preparation method is that the sterilization temperature is 115-121 ℃.
According to the invention, the high temperature sterilization condition in the fifth step of the plant combination preparation method is that the sterilization time is 10-60 min.
According to the invention, the preparation of the ganoderma lucidum culture medium in the step II of the plant combination preparation method also comprises subpackaging and sterilization.
According to the invention, in the step II of the plant combination preparation method, the ganoderma lucidum culture medium is subpackaged in a 250mL triangular flask, and the liquid filling amount in each flask is 50-100 mL.
According to the invention, the sterilization mode of the ganoderma lucidum culture medium in the step II of the plant combination preparation method is a conventional sterilization mode in the field.
According to the invention, the ganoderma lucidum culture medium is sterilized at high temperature in the step II of the plant combination preparation method.
According to the invention, the high-temperature sterilization temperature of the ganoderma lucidum culture medium in the step II of the plant combination preparation method is 115-121 ℃.
According to the invention, the high-temperature sterilization time of the ganoderma lucidum culture medium in the step II of the plant combination preparation method is 10-60 min.
According to the invention, the preparation of the bidirectional fermentation medium in the third step of the plant combination preparation method also comprises subpackaging and sterilizing.
According to the invention, the bidirectional fermentation medium in the third step of the plant combination preparation method is distributed into 500mL triangular flasks, and the liquid filling amount of each flask is 50-200 mL.
According to the invention, the sterilization mode of the bidirectional fermentation culture medium in the third step of the plant combination preparation method is a conventional sterilization mode in the field.
According to the invention, the plant combination preparation method comprises the step III of sterilizing the bidirectional fermentation culture medium at high temperature.
According to the invention, the high temperature sterilization temperature of the bidirectional fermentation medium in the step III in the plant combination preparation method is 115-121 ℃.
According to the invention, the high-temperature sterilization time of the bidirectional fermentation medium in the third step of the plant combination preparation method is 10-60 min.
According to the invention, the post-treatment of the fermentation liquor in the fifth step of the plant combination preparation method also comprises the addition of auxiliary materials after sterilization.
According to the invention, in the step (v) of the plant combination preparation method, the fermentation liquor is post-treated and auxiliary materials are added, wherein the auxiliary materials comprise glycerol, pentanediol and hexanediol.
According to the invention, the auxiliary materials added in the post-treatment of the fermentation liquor in the step (v) in the preparation method of the plant combination comprise 10-50% of glycerol, 0.1-5% of pentanediol and 0.1-5% of hexanediol by weight of the bidirectional fermentation liquor.
According to the invention, the post-treatment of the fermentation liquor in the fifth step of the plant combination preparation method also comprises the sterilization after the addition of auxiliary materials.
According to the invention, in the step (v) of the plant combination preparation method, the fermentation liquor is subjected to post-treatment, auxiliary materials are added, and the sterilization temperature is 70-100 ℃.
According to the invention, in the step (v) of the preparation method of the plant combination, the fermentation liquor is subjected to post-treatment, auxiliary materials are added, and then the sterilization time is 20-60 min.
According to the invention, in the step of the plant combination preparation method, the dendrobium officinale kimura et migo is crushed and sieved by a sieve with 80-120 meshes in the pretreatment of the raw materials, and the parts below the sieve with 80 meshes and above the sieve with 120 meshes are taken for standby.
According to the invention, in the step of raw material pretreatment, the rhizoma polygonati is crushed and sieved by a 10-25-mesh sieve, and the parts below the 10-mesh sieve and above the 25-mesh sieve are taken for standby.
According to the invention, in the preparation method of the plant combination, the tuberose in the pretreatment of the raw materials is crushed and sieved by a sieve with 10-25 meshes, and the parts below the sieve with 10 meshes and above the sieve with 25 meshes are taken for standby.
In a second aspect, the present invention provides a method for preparing a plant composition according to the first aspect of the present invention.
According to the invention, the preparation method comprises the following steps:
pretreating raw materials: pulverizing herba Dendrobii, rhizoma Polygonati, and tuberose respectively, and sieving;
② activating ganoderma lucidum: adding 0.5-5% of starch, 0.1-5% of yeast powder, 0.1-2% of sodium chloride and 0.01-0.1% of calcium carbonate into 100% of water, mixing, subpackaging and sterilizing to prepare a ganoderma lucidum culture medium; selecting Ganoderma mycelia, inoculating to culture medium, and culturing at 25-35 deg.C and 50-300rpm for 50-80 hr to obtain Ganoderma seed solution;
preparing a bidirectional fermentation culture medium: mixing starch 0.5-5% and yeast powder 0.1-5%; 0.1-2% of sodium chloride, 0.01-0.1% of calcium carbonate, 1-10% of dendrobium officinale, 0.01-1% of rhizoma polygonati, 0.01-1% of tuberose, and water which is added to 100% are mixed, subpackaged and sterilized to prepare a bidirectional fermentation culture medium;
inoculating and fermenting: inoculating the ganoderma lucidum seed liquid obtained in the step (II) to a fermentation culture medium according to the inoculation amount of 1-15%, and then culturing for 1-7d at the temperature of 25-35 ℃ and at the speed of 50-300 rpm;
fifthly, post-treatment of fermentation liquor: sterilizing and filtering.
According to some embodiments of the present invention, in the third step of the preparation method of the plant composition, the bidirectional fermentation medium is 0.5 to 5% of starch, 0.1 to 5% of yeast powder, 0.1 to 2% of sodium chloride, 0.01 to 0.1% of calcium carbonate, 1 to 2% of dendrobium officinale, 0.01 to 0.2% of rhizoma polygonati, 0.01 to 0.2% of tuberose, and water to 100%.
According to the invention, the pH value of the bidirectional fermentation culture in the third step is 3.5-7.5.
According to some embodiments of the present invention, the pH value of the bidirectional fermentation culture in the third step is 5.5.
According to some embodiments of the present invention, the temperature of the bidirectional fermentation culture in the third step is 30 ℃.
According to some embodiments of the present invention, the rotation speed of the bidirectional fermentation culture in the third step is 150 rpm.
According to some embodiments of the present invention, the time for the bidirectional fermentation culture in step (iii) is 3 d.
According to the invention, the inoculation amount of the ganoderma lucidum seed liquid in the step (iv) in the plant combination preparation method is 1-15%.
According to some embodiments of the present invention, the inoculation amount of the ganoderma lucidum seed solution in the fourth step of the plant combination preparation method is 10%.
According to the invention, the sterilization mode in the step (v) of the plant combination preparation method is a conventional sterilization mode in the field.
According to the invention, the sterilization mode in the fifth step of the plant combination preparation method is high-temperature sterilization.
According to the invention, the high temperature sterilization condition in the step (c) of the plant combination preparation method is that the sterilization temperature is 115-121 ℃.
According to the invention, the high temperature sterilization condition in the fifth step of the plant combination preparation method is that the sterilization time is 10-60 min.
According to the invention, in the step II of the plant combination preparation method, the ganoderma lucidum culture medium is subpackaged in a 250mL triangular flask, and the liquid filling amount in each flask is 50-100 mL.
According to the invention, the sterilization mode of the ganoderma lucidum culture medium in the step II of the plant combination preparation method is a conventional sterilization mode in the field.
According to the invention, the ganoderma lucidum culture medium is sterilized at high temperature in the step II of the plant combination preparation method.
According to the invention, the high-temperature sterilization temperature of the ganoderma lucidum culture medium in the step II of the plant combination preparation method is 115-121 ℃.
According to the invention, the high-temperature sterilization time of the ganoderma lucidum culture medium in the step II of the plant combination preparation method is 10-60 min.
According to the invention, the bidirectional fermentation medium in the third step of the plant combination preparation method is distributed into 500mL triangular flasks, and the liquid filling amount of each flask is 50-200 mL.
According to the invention, the sterilization mode of the bidirectional fermentation culture medium in the third step of the plant combination preparation method is a conventional sterilization mode in the field.
According to the invention, the plant combination preparation method comprises the step III of sterilizing the bidirectional fermentation culture medium at high temperature.
According to the invention, the high temperature sterilization temperature of the bidirectional fermentation medium in the step III in the plant combination preparation method is 115-121 ℃.
According to the invention, the high-temperature sterilization time of the bidirectional fermentation medium in the third step of the plant combination preparation method is 10-60 min.
According to the invention, the post-treatment of the fermentation liquor in the fifth step of the plant combination preparation method also comprises the addition of auxiliary materials after sterilization.
According to the invention, in the step (v) of the plant combination preparation method, the fermentation liquor is post-treated and auxiliary materials are added, wherein the auxiliary materials comprise glycerol, pentanediol and hexanediol.
According to the invention, the auxiliary materials added in the post-treatment of the fermentation liquor in the step (v) in the preparation method of the plant combination comprise 10-50% of glycerol, 0.1-5% of pentanediol and 0.1-5% of hexanediol by weight of the bidirectional fermentation liquor.
According to the invention, the post-treatment of the fermentation liquor in the fifth step of the plant combination preparation method also comprises the sterilization after the addition of auxiliary materials.
According to the invention, in the step (v) of the plant combination preparation method, the fermentation liquor is subjected to post-treatment, auxiliary materials are added, and the sterilization temperature is 70-100 ℃.
According to the invention, in the step (v) of the preparation method of the plant combination, the fermentation liquor is subjected to post-treatment, auxiliary materials are added, and then the sterilization time is 20-60 min.
According to the invention, in the step of the plant combination preparation method, the dendrobium officinale kimura et migo is crushed and sieved by a sieve with 80-120 meshes in the pretreatment of the raw materials, and the parts below the sieve with 80 meshes and above the sieve with 120 meshes are taken for standby.
According to the invention, in the step of raw material pretreatment, the rhizoma polygonati is crushed and sieved by a 10-25-mesh sieve, and the parts below the 10-mesh sieve and above the 25-mesh sieve are taken for standby.
According to the invention, in the preparation method of the plant combination, the tuberose in the pretreatment of the raw materials is crushed and sieved by a sieve with 10-25 meshes, and the parts below the sieve with 10 meshes and above the sieve with 25 meshes are taken for standby.
In a third aspect, the present invention provides the use of a plant composition according to the first aspect of the present invention or a plant composition prepared by a method for preparing a plant composition according to the second aspect of the present invention in cosmetics and/or toiletries.
According to the invention, the plant composition is applied to the preparation of skin care products and/or cosmetics with anti-aging and/or whitening effects.
According to the present invention, the cosmetic is not particularly limited in kind or formulation, and may be, for example, a mask, a cream, a face lotion, a essence, a milky lotion, or the like.
The invention has the beneficial effects that:
1. the plant composition is prepared by a bidirectional fermentation process, the dendrobium officinale, the polygonatum sibiricum and the tuberose are subjected to low-temperature fermentation by adopting the fungus lucid ganoderma which is homologous in medicine and food, the dendrobium officinale, the polygonatum sibiricum and the tuberose are subjected to deep fermentation decomposition into a micromolecule carbon source by utilizing a strong enzyme system which is capable of decomposing substances such as cellulose and the like of the fungus, the utilization rate of active ingredients of plant raw materials is increased, and the polysaccharide content of the active ingredients of the plant composition is improved.
2. The plant composition achieves a good anti-aging effect by inhibiting non-enzymatic glycosylation and inhibiting Clock gene expression, achieves a good whitening effect by inhibiting tyrosinase activity, and can achieve double effects of anti-aging and whitening by adding the plant composition into cosmetics and/or skin care products.
Drawings
In order to more clearly illustrate the embodiments of the present invention or the technical solutions in the prior art, the drawings used in the following will be briefly described.
FIG. 1 is a graph showing the results of Clock expressing genes.
Detailed Description
The invention is further illustrated below with reference to specific examples, to which, however, the invention is not restricted.
The experimental methods described in the following examples are all conventional methods unless otherwise specified; the experimental materials and reagents are commercially available, unless otherwise specified.
The names of the main raw materials, suppliers/manufacturers and production places of the invention are shown in table 1:
TABLE 1 raw materials and sources
Name of raw materials Supplier/producer Producing area
DendrobiumDendrobium officinale Kimura et Migo Kunming plant institute of Chinese academy of sciences Yunnan province
Polygonatum sibiricum Beijing Qiancao Chinese medicinal drink Co Ltd Hebei river
Tuberose Yunnan Rui flower teaLimited company Yunnan province
Ganoderma lucidum China center for preservation and management of industrial microbial strains China
Hexanediol De Zhi Xin (Shanghai) Co., Ltd Germany
Pentanediol De Zhi Xin (Shanghai) Co., Ltd Germany
Example 1
Pretreating raw materials: crushing Dendrobium officinale, sieving with a 80-mesh sieve, respectively crushing rhizoma Polygonati and tuberose, and sieving with a 10-mesh sieve;
② activating ganoderma lucidum: 2% of starch, 1.5% of yeast powder, 0.3% of sodium chloride, 0.02% of calcium carbonate, and 100% of pure water, preparing a ganoderma lucidum culture medium, and subpackaging in 250mL conical flasks, wherein the liquid filling amount of each flask is 100 mL. Respectively placing the culture medium in a sterilization pot, sterilizing at 121 deg.C for 20min, and cooling in a super clean bench after sterilization; selecting mycelium from the ganoderma lucidum slant, and inoculating the mycelium into a ganoderma lucidum culture medium; culturing the inoculated culture medium in a constant temperature shaking table at the culture temperature of 30 ℃ and the rotation speed of the shaking table of 150rpm for 3d to obtain ganoderma lucidum seed liquid;
preparing a bidirectional fermentation culture medium: according to the mixture ratio of the raw materials in the table 2, 2% of starch, 1.5% of yeast powder, 0.3% of sodium chloride and 0.02% of calcium carbonate are added, water is added to 100%, a bidirectional fermentation culture medium is prepared, the initial pH value is 5.5, the bidirectional fermentation culture medium is subpackaged in 250mL conical flasks, the liquid filling amount of each flask is 100mL, and each culture medium is 3 flasks. Respectively placing the two-way fermentation culture medium in a sterilization pot, sterilizing at 121 deg.C for 20min, and cooling in a super clean bench after sterilization;
inoculating and fermenting: inoculating lucid Ganoderma seed liquid according to the inoculation amount of 10%, then placing the culture medium in a constant-temperature shaking table for culturing at the culture temperature of 30 ℃ and the rotation speed of the shaking table of 150rpm for 3d, wherein each group of samples comprises 3 bottles;
fifthly, post-treatment of fermentation liquor: and (4) after the fermentation is finished, sterilizing the fermentation liquor for 20min at 121 ℃. And (4) filtering the fermentation liquor after sterilization, and detecting the content of polysaccharide and flavone in the filtrate.
TABLE 2 optimization of the amount of Chinese medicinal materials added
Sample numbering Dendrobium officinale (%) Sealwort (%) Tuberose (%)
Sample 1 1 0.1 0.05
Sample 2 1 0.2 0.1
Sample 3 1 0.4 0.2
Sample No. 4 2 0.1 0.1
Sample No. 5 2 0.2 0.2
Sample No. 6 2 0.4 0.05
Sample 7 4 0.1 0.2
Sample 8 4 0.2 0.05
Sample 9 4 0.4 0.1
Example 2
Pretreating raw materials: crushing Dendrobium officinale, sieving with a 80-mesh sieve, respectively crushing rhizoma Polygonati and tuberose, and sieving with a 10-mesh sieve;
② activating ganoderma lucidum: 2% of starch, 1.5% of yeast powder, 0.3% of sodium chloride, 0.02% of calcium carbonate, and 100% of pure water, preparing a ganoderma lucidum culture medium, and subpackaging in 250mL conical flasks, wherein the liquid filling amount of each flask is 100 mL. Respectively placing the culture medium in a sterilization pot, sterilizing at 121 deg.C for 20min, and cooling in a super clean bench after sterilization; selecting mycelium from the ganoderma lucidum slant, and inoculating the mycelium into a ganoderma lucidum culture medium; culturing the inoculated culture medium in a constant temperature shaking table at the culture temperature of 30 ℃ and the rotation speed of the shaking table of 150rpm for 3d to obtain ganoderma lucidum seed liquid;
preparing a bidirectional fermentation culture medium: 2% of dendrobium officinale, 0.2% of rhizoma polygonati, 0.2% of tuberose, 2% of starch, 1.5% of yeast powder, 0.3% of sodium chloride, 0.02% of calcium carbonate, and 100% of water is added to prepare a bidirectional fermentation culture medium, the initial pH value is 5.5, the bidirectional fermentation culture medium is subpackaged in 250mL conical flasks, the liquid filling amount of each flask is 100mL, and each culture medium is 3 flasks. Respectively placing the two-way fermentation culture medium in a sterilization pot, sterilizing at 121 deg.C for 20min, and cooling in a super clean bench after sterilization;
inoculating and fermenting: inoculating glossy ganoderma seed liquid according to the inoculation amount (2%, 5%, 10%, 15%) in the table 3, then placing the culture medium in a constant temperature shaking table for culturing for 3d at the culture temperature of 30 ℃ and the rotation speed of the shaking table of 150rpm, and culturing each group of samples for 3 bottles;
fifthly, post-treatment of fermentation liquor: and (4) after the fermentation is finished, sterilizing the fermentation liquor for 20min at 121 ℃. And (4) filtering the fermentation liquor after sterilization, and detecting the content of polysaccharide and flavone in the filtrate.
TABLE 3 fermentation inoculum size optimization
Sample numbering Inoculum size (%)
Sample 10 2
Sample 11 5
Sample 12 10
Sample 13 15
Example 3
Pretreating raw materials: crushing Dendrobium officinale, sieving with a 80-mesh sieve, respectively crushing rhizoma Polygonati and tuberose, and sieving with a 10-mesh sieve;
② activating ganoderma lucidum: 2% of starch, 1.5% of yeast powder, 0.3% of sodium chloride, 0.02% of calcium carbonate, and 100% of pure water, preparing a ganoderma lucidum culture medium, and subpackaging in 250mL conical flasks, wherein the liquid filling amount of each flask is 100 mL. Respectively placing the culture medium in a sterilization pot, sterilizing at 121 deg.C for 20min, and cooling in a super clean bench after sterilization; selecting mycelium from the ganoderma lucidum slant, and inoculating the mycelium into a ganoderma lucidum culture medium; culturing the inoculated culture medium in a constant temperature shaking table at the culture temperature of 30 ℃ and the rotation speed of the shaking table of 150rpm for 3d to obtain ganoderma lucidum seed liquid;
preparing a bidirectional fermentation culture medium: 2% of dendrobium officinale, 0.2% of rhizoma polygonati, 0.2% of tuberose, 2% of starch, 1.5% of yeast powder, 0.3% of sodium chloride, 0.02% of calcium carbonate, and 100% of water is added to prepare a bidirectional fermentation culture medium, the initial pH value is 5.5, the bidirectional fermentation culture medium is subpackaged in 250mL conical flasks, the liquid filling amount of each flask is 100mL, and each culture medium is 3 flasks. Respectively placing the two-way fermentation culture medium in a sterilization pot, sterilizing at 121 deg.C for 20min, and cooling in a super clean bench after sterilization;
inoculating and fermenting: inoculating the ganoderma lucidum seed liquid according to the inoculation amount of 10%, and then placing the culture medium in a constant-temperature shaking table for culture at the culture temperature of 30 ℃ and the rotation speed of the shaking table of 150rpm for 1d, 3d, 5d and 7d, wherein each group of samples comprises 3 bottles;
fifthly, post-treatment of fermentation liquor: and (4) after the fermentation is finished, sterilizing the fermentation liquor for 20min at 121 ℃. And (4) filtering the fermentation liquor after sterilization, and detecting the content of polysaccharide and flavone in the filtrate.
TABLE 4 fermentation time optimization
Sample numbering Fermentation time (d)
Sample 14 1
Sample 15 3
Sample 16 5
Sample 17 7
Example 4
Pretreating raw materials: crushing Dendrobium officinale, sieving with a 80-mesh sieve, respectively crushing rhizoma Polygonati and tuberose, and sieving with a 10-mesh sieve;
② activating ganoderma lucidum: 2% of starch, 1.5% of yeast powder, 0.3% of sodium chloride, 0.02% of calcium carbonate, and 100% of pure water, preparing a ganoderma lucidum culture medium, and subpackaging in 250mL conical flasks, wherein the liquid filling amount of each flask is 100 mL. Respectively placing the culture medium in a sterilization pot, sterilizing at 121 deg.C for 20min, and cooling in a super clean bench after sterilization; selecting mycelium from the ganoderma lucidum slant, and inoculating the mycelium into a ganoderma lucidum culture medium; culturing the inoculated culture medium in a constant temperature shaking table at the culture temperature of 30 ℃ and the rotation speed of the shaking table of 150rpm for 3d to obtain ganoderma lucidum seed liquid;
preparing a bidirectional fermentation culture medium: 2% of dendrobium officinale, 0.2% of rhizoma polygonati, 0.2% of tuberose, 2% of starch, 1.5% of yeast powder, 0.3% of sodium chloride, 0.02% of calcium carbonate, and 100% of water is added to prepare a bidirectional fermentation culture medium, the initial pH value is 5.5, the bidirectional fermentation culture medium is subpackaged in 250mL conical flasks, the liquid filling amount of each flask is 100mL, and each culture medium is 3 flasks. Respectively placing the two-way fermentation culture medium in a sterilization pot, sterilizing at 121 deg.C for 20min, and cooling in a super clean bench after sterilization;
inoculating and fermenting: inoculating the ganoderma lucidum seed liquid according to the inoculation amount of 10%, then placing the culture medium in a constant-temperature shaking table for culturing at the culture temperature of 28 ℃, 30 ℃ and 35 ℃ at the shaking table rotating speed of 150rpm for 3d, wherein each group of samples comprises 3 bottles;
fifthly, post-treatment of fermentation liquor: and (4) after the fermentation is finished, sterilizing the fermentation liquor for 20min at 121 ℃. And (4) filtering the fermentation liquor after sterilization, and detecting the content of polysaccharide and flavone in the filtrate.
TABLE 5 fermentation temperature optimization
Sample numbering Fermentation temperature (. degree.C.)
Sample 18 28
Sample 19 30
Sample 20 35
Example 5
Pretreating raw materials: crushing Dendrobium officinale, sieving with a 80-mesh sieve, respectively crushing rhizoma Polygonati and tuberose, and sieving with a 10-mesh sieve;
② activating ganoderma lucidum: 2% of starch, 1.5% of yeast powder, 0.3% of sodium chloride, 0.02% of calcium carbonate, and 100% of pure water, preparing a ganoderma lucidum culture medium, and subpackaging in 250mL conical flasks, wherein the liquid filling amount of each flask is 100 mL. Respectively placing the culture medium in a sterilization pot, sterilizing at 121 deg.C for 20min, and cooling in a super clean bench after sterilization; selecting mycelium from the ganoderma lucidum slant, and inoculating the mycelium into a ganoderma lucidum culture medium; culturing the inoculated culture medium in a constant temperature shaking table at the culture temperature of 30 ℃ and the rotation speed of the shaking table of 150rpm for 3d to obtain ganoderma lucidum seed liquid;
preparing a bidirectional fermentation culture medium: 2% of dendrobium officinale, 0.2% of rhizoma polygonati, 0.2% of tuberose, 2% of starch, 1.5% of yeast powder, 0.3% of sodium chloride and 0.02% of calcium carbonate, adding water to 100%, preparing a bidirectional fermentation culture medium, adjusting the pH to 3.5 by using citric acid, adjusting the pH to 7.5 by using sodium hydroxide, and adjusting the pH of the culture medium to 5.5 in a natural state. The medium was dispensed into 250mL conical flasks, each containing 100mL of the medium, 3 flasks for each medium. Respectively placing the two-way fermentation culture medium in a sterilization pot, sterilizing at 121 deg.C for 20min, and cooling in a super clean bench after sterilization;
inoculating and fermenting: inoculating lucid Ganoderma seed liquid according to the inoculation amount of 10%, then placing the culture medium in a constant-temperature shaking table for culturing at the culture temperature of 30 ℃ and the rotation speed of the shaking table of 150rpm for 3d, wherein each group of samples comprises 3 bottles;
fifthly, post-treatment of fermentation liquor: and (4) after the fermentation is finished, sterilizing the fermentation liquor for 20min at 121 ℃. And (4) filtering the fermentation liquor after sterilization, and detecting the content of polysaccharide and flavone in the filtrate.
TABLE 6 fermentation initial pH optimization
Sample numbering pH value of fermentation
Sample 21 3.5
Sample 22 5.5
Sample 23 7.5
Example 6
Pretreating raw materials: crushing Dendrobium officinale, sieving with a 80-mesh sieve, respectively crushing rhizoma Polygonati and tuberose, and sieving with a 10-mesh sieve;
② activating ganoderma lucidum: 2% of starch, 1.5% of yeast powder, 0.3% of sodium chloride, 0.02% of calcium carbonate, and 100% of pure water, preparing a ganoderma lucidum culture medium, and subpackaging in 250mL conical flasks, wherein the liquid filling amount of each flask is 100 mL. Respectively placing the culture medium in a sterilization pot, sterilizing at 121 deg.C for 20min, and cooling in a super clean bench after sterilization; selecting mycelium from the ganoderma lucidum slant, and inoculating the mycelium into a ganoderma lucidum culture medium; culturing the inoculated culture medium in a constant temperature shaking table at the culture temperature of 30 ℃ and the rotation speed of the shaking table of 150rpm for 3d to obtain ganoderma lucidum seed liquid;
preparing a bidirectional fermentation culture medium: 2% of dendrobium officinale, 0.2% of rhizoma polygonati, 0.2% of tuberose, 2% of starch, 1.5% of yeast powder, 0.3% of sodium chloride, 0.02% of calcium carbonate, and 100% of water is added to prepare a bidirectional fermentation culture medium, the initial pH value is 5.5, the bidirectional fermentation culture medium is subpackaged in 250mL conical flasks, the liquid filling amount of each flask is 100mL, and each culture medium is 3 flasks. Respectively placing the two-way fermentation culture medium in a sterilization pot, sterilizing at 121 deg.C for 20min, and cooling in a super clean bench after sterilization;
inoculating and fermenting: inoculating lucid Ganoderma seed liquid according to the inoculation amount of 10%, then placing the culture medium in a constant-temperature shaking table for culturing at the culture temperature of 30 ℃ and the rotation speed of the shaking table of 150rpm for 3d, wherein each group of samples comprises 3 bottles;
fifthly, post-treatment of fermentation liquor: after the fermentation is finished, sterilizing the fermentation liquor at 121 ℃ for 20min, cooling to normal temperature, finely filtering, adding 30% of glycerol, 1% of hexanediol and 2% of pentanediol, sterilizing at 95 ℃ for 40min, and cooling to obtain the product of example 6.
Example 7
Pretreating raw materials: pulverizing herba Dendrobii, sieving with 80 mesh sieve, pulverizing rhizoma Polygonati and tuberose respectively, and sieving with 10 mesh sieve;
② activating ganoderma lucidum: 2% of starch, 1.5% of yeast powder, 0.3% of sodium chloride, 0.02% of calcium carbonate, and 100% of pure water, preparing a ganoderma lucidum culture medium, and subpackaging in 250mL conical flasks, wherein the liquid filling amount of each flask is 100 mL. Respectively placing the culture medium in a sterilization pot, sterilizing at 121 deg.C for 20min, and cooling in a super clean bench after sterilization; selecting mycelium from the ganoderma lucidum slant, and inoculating the mycelium into a ganoderma lucidum culture medium; culturing the inoculated culture medium in a constant temperature shaking table at the culture temperature of 30 ℃ and the rotation speed of the shaking table of 150rpm for 3d to obtain ganoderma lucidum seed liquid;
preparing a traditional Chinese medicine bidirectional fermentation culture medium: 1% of dendrobium officinale, 0.01% of rhizoma polygonati, 0.01% of tuberose, 2% of starch, 1.5% of yeast powder, 0.3% of sodium chloride, 0.02% of calcium carbonate, and 100% of water are added to prepare a traditional Chinese medicine bidirectional fermentation culture medium, the initial pH value is 5.5, the traditional Chinese medicine bidirectional fermentation culture medium is subpackaged in 250mL conical flasks, the liquid filling amount of each flask is 100mL, and each culture medium is 3 flasks. Respectively placing the two-way fermentation culture medium of the traditional Chinese medicine in a sterilization pot, sterilizing at 121 ℃ for 20min, and after the sterilization is finished, placing the two-way fermentation culture medium in a super clean bench for cooling for later use;
inoculating and fermenting: inoculating lucid Ganoderma seed liquid according to the inoculation amount of 10%, then placing the culture medium in a constant-temperature shaking table for culturing at the culture temperature of 30 ℃ and the rotation speed of the shaking table of 150rpm for 3d, wherein each group of samples comprises 3 bottles;
fifthly, post-treatment of fermentation liquor: after the fermentation is finished, sterilizing the fermentation liquor at 121 ℃ for 20min, cooling to normal temperature, finely filtering, adding 30% of glycerol, 1% of hexanediol and 2% of pentanediol, sterilizing at 95 ℃ for 40min, and cooling to obtain the product of example 7.
Example 8
Pretreating raw materials: pulverizing herba Dendrobii, sieving with 80 mesh sieve, pulverizing rhizoma Polygonati and tuberose respectively, and sieving with 10 mesh sieve;
② activating ganoderma lucidum: 2% of starch, 1.5% of yeast powder, 0.3% of sodium chloride, 0.02% of calcium carbonate, and 100% of pure water, preparing a ganoderma lucidum culture medium, and subpackaging in 250mL conical flasks, wherein the liquid filling amount of each flask is 100 mL. Respectively placing the culture medium in a sterilization pot, sterilizing at 121 deg.C for 20min, and cooling in a super clean bench after sterilization; selecting mycelium from the ganoderma lucidum slant, and inoculating the mycelium into a ganoderma lucidum culture medium; culturing the inoculated culture medium in a constant temperature shaking table at the culture temperature of 30 ℃ and the rotation speed of the shaking table of 150rpm for 3d to obtain ganoderma lucidum seed liquid;
preparing a traditional Chinese medicine bidirectional fermentation culture medium: 10% of dendrobium officinale, 1% of rhizoma polygonati, 1% of tuberose, 2% of starch, 1.5% of yeast powder, 0.3% of sodium chloride, 0.02% of calcium carbonate, and adding water to 100% to prepare a traditional Chinese medicine bidirectional fermentation culture medium, wherein the initial pH value is 5.5, the traditional Chinese medicine bidirectional fermentation culture medium is subpackaged in 250mL conical flasks, the liquid filling amount of each flask is 100mL, and each culture medium is 3 flasks. Respectively placing the two-way fermentation culture medium of the traditional Chinese medicine in a sterilization pot, sterilizing at 121 ℃ for 20min, and after the sterilization is finished, placing the two-way fermentation culture medium in a super clean bench for cooling for later use;
inoculating and fermenting: inoculating lucid Ganoderma seed liquid according to the inoculation amount of 10%, then placing the culture medium in a constant-temperature shaking table for culturing at the culture temperature of 30 ℃ and the rotation speed of the shaking table of 150rpm for 3d, wherein each group of samples comprises 3 bottles;
fifthly, post-treatment of fermentation liquor: after the fermentation is finished, sterilizing the fermentation liquor at 121 ℃ for 20min, cooling to normal temperature, finely filtering, adding 30% of glycerol, 1% of hexanediol and 2% of pentanediol, sterilizing at 95 ℃ for 40min, and cooling to obtain the product of example 8.
Example 9
Pretreating raw materials: pulverizing herba Dendrobii, sieving with 80 mesh sieve, pulverizing rhizoma Polygonati and tuberose respectively, and sieving with 10 mesh sieve;
② activating ganoderma lucidum: 2% of starch, 1.5% of yeast powder, 0.3% of sodium chloride, 0.02% of calcium carbonate, and 100% of pure water, preparing a ganoderma lucidum culture medium, and subpackaging in 250mL conical flasks, wherein the liquid filling amount of each flask is 100 mL. Respectively placing the culture medium in a sterilization pot, sterilizing at 121 deg.C for 20min, and cooling in a super clean bench after sterilization; selecting mycelium from the ganoderma lucidum slant, and inoculating the mycelium into a ganoderma lucidum culture medium; culturing the inoculated culture medium in a constant temperature shaking table at the culture temperature of 30 ℃ and the rotation speed of the shaking table of 150rpm for 3d to obtain ganoderma lucidum seed liquid;
preparing a traditional Chinese medicine bidirectional fermentation culture medium: 10% of dendrobium officinale, 1% of rhizoma polygonati, 0.01% of tuberose, 2% of starch, 1.5% of yeast powder, 0.3% of sodium chloride, 0.02% of calcium carbonate, and adding water to 100% to prepare a traditional Chinese medicine bidirectional fermentation culture medium, wherein the initial pH value is 5.5, the traditional Chinese medicine bidirectional fermentation culture medium is subpackaged in 250mL conical flasks, the liquid filling amount of each flask is 100mL, and each culture medium is 3 flasks. Respectively placing the two-way fermentation culture medium of the traditional Chinese medicine in a sterilization pot, sterilizing at 121 ℃ for 20min, and after the sterilization is finished, placing the two-way fermentation culture medium in a super clean bench for cooling for later use;
inoculating and fermenting: inoculating lucid Ganoderma seed liquid according to the inoculation amount of 10%, then placing the culture medium in a constant-temperature shaking table for culturing at the culture temperature of 30 ℃ and the rotation speed of the shaking table of 150rpm for 3d, wherein each group of samples comprises 3 bottles;
fifthly, post-treatment of fermentation liquor: after the fermentation is finished, sterilizing the fermentation liquor at 121 ℃ for 20min, cooling to normal temperature, finely filtering, adding 30% of glycerol, 1% of hexanediol and 2% of pentanediol, sterilizing at 95 ℃ for 40min, and cooling to obtain the product of example 9.
Comparative example 1
2 percent of dendrobium officinale, 0.2 percent of rhizoma polygonati and 0.2 percent of tuberose, adding water to 100 percent, extracting for 2 hours at 80 ℃, filtering, and detecting the content of polysaccharide and flavone in the filtrate.
Comparative example 2
Pretreating raw materials: respectively crushing rhizoma Polygonati and tuberose, and sieving with 10 mesh sieve;
② activating ganoderma lucidum: 2% of starch, 1.5% of yeast powder, 0.3% of sodium chloride, 0.02% of calcium carbonate, and 100% of pure water, preparing a ganoderma lucidum culture medium, and subpackaging in 250mL conical flasks, wherein the liquid filling amount of each flask is 100 mL. Respectively placing the culture medium in a sterilization pot, sterilizing at 121 deg.C for 20min, and cooling in a super clean bench after sterilization; selecting mycelium from the ganoderma lucidum slant, and inoculating the mycelium into a ganoderma lucidum culture medium; culturing the inoculated culture medium in a constant temperature shaking table at the culture temperature of 30 ℃ and the rotation speed of the shaking table of 150rpm for 3d to obtain ganoderma lucidum seed liquid;
preparing a bidirectional fermentation culture medium: 0.2% of rhizoma polygonati, 0.2% of tuberose, 2% of starch, 1.5% of yeast powder, 0.3% of sodium chloride, 0.02% of calcium carbonate, and adding water to 100% to prepare a bidirectional fermentation culture medium, wherein the initial pH value is 5.5, the bidirectional fermentation culture medium is subpackaged in 250mL conical flasks, the liquid filling amount of each flask is 100mL, and each culture medium is 3 flasks. Respectively placing the two-way fermentation culture medium in a sterilization pot, sterilizing at 121 deg.C for 20min, and cooling in a super clean bench after sterilization;
inoculating and fermenting: inoculating lucid Ganoderma seed liquid according to the inoculation amount of 10%, then placing the culture medium in a constant-temperature shaking table for culturing at the culture temperature of 30 ℃ and the rotation speed of the shaking table of 150rpm for 3d, wherein each group of samples comprises 3 bottles;
fifthly, post-treatment of fermentation liquor: and (4) after the fermentation is finished, sterilizing the fermentation liquor for 20min at 121 ℃. And after sterilization, filtering the fermentation liquor, and detecting the contents of polysaccharide and flavone in the filtrate.
Comparative example 3
Pretreating raw materials: crushing Dendrobium officinale, sieving with a 80-mesh sieve, respectively crushing rhizoma Polygonati and tuberose, and sieving with a 10-mesh sieve;
② activating ganoderma lucidum: 2% of starch, 1.5% of yeast powder, 0.3% of sodium chloride, 0.02% of calcium carbonate, and 100% of pure water, preparing a ganoderma lucidum culture medium, and subpackaging in 250mL conical flasks, wherein the liquid filling amount of each flask is 100 mL. Respectively placing the culture medium in a sterilization pot, sterilizing at 121 deg.C for 20min, and cooling in a super clean bench after sterilization; selecting mycelium from the ganoderma lucidum slant, and inoculating the mycelium into a ganoderma lucidum culture medium; culturing the inoculated culture medium in a constant temperature shaking table at the culture temperature of 30 ℃ and the rotation speed of the shaking table of 150rpm for 3d to obtain ganoderma lucidum seed liquid;
preparing a bidirectional fermentation culture medium: 0.5% of dendrobium officinale, 0.2% of rhizoma polygonati, 0.2% of tuberose, 2% of starch, 1.5% of yeast powder, 0.3% of sodium chloride and 0.02% of calcium carbonate, adding water to 100% to prepare a bidirectional fermentation culture medium, wherein the initial pH value is 5.5, the culture medium is subpackaged in 250mL conical flasks, the liquid filling amount of each flask is 100mL, and each culture medium is 3 flasks. Respectively placing the two-way fermentation culture medium in a sterilization pot, sterilizing at 121 deg.C for 20min, and cooling in a super clean bench after sterilization;
inoculating and fermenting: inoculating lucid Ganoderma seed liquid according to the inoculation amount of 10%, then placing the culture medium in a constant-temperature shaking table for culturing at the culture temperature of 30 ℃ and the rotation speed of the shaking table of 150rpm for 3d, wherein each group of samples comprises 3 bottles;
fifthly, post-treatment of fermentation liquor: and (4) after the fermentation is finished, sterilizing the fermentation liquor for 20min at 121 ℃. And after sterilization, filtering the fermentation liquor, and detecting the contents of polysaccharide and flavone in the filtrate.
Comparative example 4
Pretreating raw materials: respectively crushing rhizoma Polygonati and tuberose, and sieving with 10 mesh sieve;
② activating ganoderma lucidum: 2% of starch, 1.5% of yeast powder, 0.3% of sodium chloride, 0.02% of calcium carbonate, and 100% of pure water, preparing a ganoderma lucidum culture medium, and subpackaging in 250mL conical flasks, wherein the liquid filling amount of each flask is 100 mL. Respectively placing the culture medium in a sterilization pot, sterilizing at 121 deg.C for 20min, and cooling in a super clean bench after sterilization; selecting mycelium from the ganoderma lucidum slant, and inoculating the mycelium into a ganoderma lucidum culture medium; culturing the inoculated culture medium in a constant temperature shaking table at the culture temperature of 30 ℃ and the rotation speed of the shaking table of 150rpm for 3d to obtain ganoderma lucidum seed liquid;
preparing a bidirectional fermentation culture medium: 0.2% of rhizoma polygonati, 0.2% of tuberose, 2% of starch, 1.5% of yeast powder, 0.3% of sodium chloride, 0.02% of calcium carbonate, and adding water to 100% to prepare a bidirectional fermentation culture medium, wherein the initial pH value is 5.5, the bidirectional fermentation culture medium is subpackaged in 250mL conical flasks, the liquid filling amount of each flask is 100mL, and each culture medium is 3 flasks. Respectively placing the two-way fermentation culture medium in a sterilization pot, sterilizing at 121 deg.C for 20min, and cooling in a super clean bench after sterilization;
inoculating and fermenting: inoculating lucid Ganoderma seed liquid according to the inoculation amount of 10%, then placing the culture medium in a constant-temperature shaking table for culturing at the culture temperature of 30 ℃ and the rotation speed of the shaking table of 150rpm for 3d, wherein each group of samples comprises 3 bottles;
fifthly, post-treatment of fermentation liquor: and (4) after the fermentation is finished, sterilizing the fermentation liquor for 20min at 121 ℃. After sterilization, the fermentation broth was filtered, 30% glycerol, 1% hexanediol, 2% pentanediol were added, sterilized at 95 ℃ for 40min, and cooled to obtain comparative example 4.
Comparative example 5
Pretreating raw materials: crushing Dendrobium officinale, sieving with a 80-mesh sieve, respectively crushing rhizoma Polygonati and tuberose, and sieving with a 10-mesh sieve;
② activating ganoderma lucidum: 2% of starch, 1.5% of yeast powder, 0.3% of sodium chloride, 0.02% of calcium carbonate, and 100% of pure water, preparing a ganoderma lucidum culture medium, and subpackaging in 250mL conical flasks, wherein the liquid filling amount of each flask is 100 mL. Respectively placing the culture medium in a sterilization pot, sterilizing at 121 deg.C for 20min, and cooling in a super clean bench after sterilization; selecting mycelium from the ganoderma lucidum slant, and inoculating the mycelium into a ganoderma lucidum culture medium; culturing the inoculated culture medium in a constant temperature shaking table at the culture temperature of 30 ℃ and the rotation speed of the shaking table of 150rpm for 3d to obtain ganoderma lucidum seed liquid;
preparing a bidirectional fermentation culture medium: 0.5% of dendrobium officinale, 0.2% of rhizoma polygonati, 0.2% of tuberose, 2% of starch, 1.5% of yeast powder, 0.3% of sodium chloride and 0.02% of calcium carbonate, adding water to 100% to prepare a bidirectional fermentation culture medium, wherein the initial pH value is 5.5, the culture medium is subpackaged in 250mL conical flasks, the liquid filling amount of each flask is 100mL, and each culture medium is 3 flasks. Respectively placing the two-way fermentation culture medium in a sterilization pot, sterilizing at 121 deg.C for 20min, and cooling in a super clean bench after sterilization;
inoculating and fermenting: inoculating lucid Ganoderma seed liquid according to the inoculation amount of 10%, then placing the culture medium in a constant-temperature shaking table for culturing at the culture temperature of 30 ℃ and the rotation speed of the shaking table of 150rpm for 3d, wherein each group of samples comprises 3 bottles;
fifthly, post-treatment of fermentation liquor: and (4) after the fermentation is finished, sterilizing the fermentation liquor for 20min at 121 ℃. After sterilization, the fermentation broth was filtered, 30% glycerol, 1% hexylene glycol, 2% pentanediol were added, and the mixture was quenched at 95 ℃ for 40min, and cooled to obtain comparative example 5.
Comparative example 6
Pretreating raw materials: crushing the dendrobium officinale and sieving the crushed dendrobium officinale with a 80-mesh sieve;
② activating ganoderma lucidum: 2% of starch, 1.5% of yeast powder, 0.3% of sodium chloride, 0.02% of calcium carbonate, and 100% of pure water, preparing a ganoderma lucidum culture medium, and subpackaging in 250mL conical flasks, wherein the liquid filling amount of each flask is 100 mL. Respectively placing the culture medium in a sterilization pot, sterilizing at 121 deg.C for 20min, and cooling in a super clean bench after sterilization; selecting mycelium from the ganoderma lucidum slant, and inoculating the mycelium into a ganoderma lucidum culture medium; culturing the inoculated culture medium in a constant temperature shaking table at the culture temperature of 30 ℃ and the rotation speed of the shaking table of 150rpm for 3d to obtain ganoderma lucidum seed liquid;
preparing a bidirectional fermentation culture medium: 2% of dendrobium officinale, 2% of starch, 1.5% of yeast powder, 0.3% of sodium chloride, 0.02% of calcium carbonate, and adding water to 100% to prepare a bidirectional fermentation culture medium, wherein the initial pH value is 5.5, the bidirectional fermentation culture medium is subpackaged in 250mL conical flasks, the liquid filling amount of each flask is 100mL, and each culture medium is 3 flasks. Respectively placing the two-way fermentation culture medium in a sterilization pot, sterilizing at 121 deg.C for 20min, and cooling in a super clean bench after sterilization;
inoculating and fermenting: inoculating lucid Ganoderma seed liquid according to the inoculation amount of 10%, then placing the culture medium in a constant-temperature shaking table for culturing at the culture temperature of 30 ℃ and the rotation speed of the shaking table of 150rpm for 3d, wherein each group of samples comprises 3 bottles;
fifthly, post-treatment of fermentation liquor: and (4) after the fermentation is finished, sterilizing the fermentation liquor for 20min at 121 ℃. After sterilization, the fermentation broth was filtered, 30% glycerol, 1% hexylene glycol, 2% pentanediol were added, and the mixture was quenched at 95 ℃ for 40min, and cooled to obtain comparative example 6.
Comparative example 7
Pretreating raw materials: crushing the dendrobium officinale, sieving the crushed dendrobium officinale with a 80-mesh sieve, crushing the polygonatum sibiricum, and sieving the crushed polygonatum sibiricum with a 10-mesh sieve;
② activating ganoderma lucidum: 2% of starch, 1.5% of yeast powder, 0.3% of sodium chloride, 0.02% of calcium carbonate, and 100% of pure water, preparing a ganoderma lucidum culture medium, and subpackaging in 250mL conical flasks, wherein the liquid filling amount of each flask is 100 mL. Respectively placing the culture medium in a sterilization pot, sterilizing at 121 deg.C for 20min, and cooling in a super clean bench after sterilization; selecting mycelium from the ganoderma lucidum slant, and inoculating the mycelium into a ganoderma lucidum culture medium; culturing the inoculated culture medium in a constant temperature shaking table at the culture temperature of 30 ℃ and the rotation speed of the shaking table of 150rpm for 3d to obtain ganoderma lucidum seed liquid;
preparing a bidirectional fermentation culture medium: 2% of dendrobium officinale, 0.2% of rhizoma polygonati, 2% of starch, 1.5% of yeast powder, 0.3% of sodium chloride, 0.02% of calcium carbonate, and adding water to 100% to prepare a bidirectional fermentation culture medium, wherein the initial pH value is 5.5, the bidirectional fermentation culture medium is subpackaged in 250mL conical flasks, the liquid filling amount of each flask is 100mL, and each culture medium is 3 flasks. Respectively placing the two-way fermentation culture medium in a sterilization pot, sterilizing at 121 deg.C for 20min, and cooling in a super clean bench after sterilization;
inoculating and fermenting: inoculating lucid Ganoderma seed liquid according to the inoculation amount of 10%, then placing the culture medium in a constant-temperature shaking table for culturing at the culture temperature of 30 ℃ and the rotation speed of the shaking table of 150rpm for 3d, wherein each group of samples comprises 3 bottles;
fifthly, post-treatment of fermentation liquor: and (4) after the fermentation is finished, sterilizing the fermentation liquor for 20min at 121 ℃. After sterilization, the fermentation broth was filtered, 30% glycerol, 1% hexylene glycol, 2% pentanediol were added, and the mixture was quenched at 95 ℃ for 40min, and cooled to obtain comparative example 7.
Comparative example 8
Pretreating raw materials: crushing the dendrobium officinale, sieving with a 80-mesh sieve, respectively crushing the tuberose, and sieving with a 10-mesh sieve;
② activating ganoderma lucidum: 2% of starch, 1.5% of yeast powder, 0.3% of sodium chloride, 0.02% of calcium carbonate, and 100% of pure water, preparing a ganoderma lucidum culture medium, and subpackaging in 250mL conical flasks, wherein the liquid filling amount of each flask is 100 mL. Respectively placing the culture medium in a sterilization pot, sterilizing at 121 deg.C for 20min, and cooling in a super clean bench after sterilization; selecting mycelium from the ganoderma lucidum slant, and inoculating the mycelium into a ganoderma lucidum culture medium; culturing the inoculated culture medium in a constant temperature shaking table at the culture temperature of 30 ℃ and the rotation speed of the shaking table of 150rpm for 3d to obtain ganoderma lucidum seed liquid;
preparing a bidirectional fermentation culture medium: 2% of dendrobium officinale, 0.2% of tuberose, 2% of starch, 1.5% of yeast powder, 0.3% of sodium chloride, 0.02% of calcium carbonate, and adding water to 100% to prepare a bidirectional fermentation culture medium, wherein the initial pH value is 5.5, the bidirectional fermentation culture medium is subpackaged in 250mL conical flasks, the liquid filling amount of each flask is 100mL, and each culture medium is 3 flasks. Respectively placing the two-way fermentation culture medium in a sterilization pot, sterilizing at 121 deg.C for 20min, and cooling in a super clean bench after sterilization;
inoculating and fermenting: inoculating lucid Ganoderma seed liquid according to the inoculation amount of 10%, then placing the culture medium in a constant-temperature shaking table for culturing at the culture temperature of 30 ℃ and the rotation speed of the shaking table of 150rpm for 3d, wherein each group of samples comprises 3 bottles;
fifthly, post-treatment of fermentation liquor: and (4) after the fermentation is finished, sterilizing the fermentation liquor for 20min at 121 ℃. After sterilization, the fermentation broth was filtered, 30% glycerol, 1% hexylene glycol, 2% pentanediol were added, and the mixture was quenched at 95 ℃ for 40min, and cooled to obtain comparative example 8.
Comparative example 9
Pretreating raw materials: pulverizing rhizoma Polygonati, and sieving with 10 mesh sieve;
② activating ganoderma lucidum: 2% of starch, 1.5% of yeast powder, 0.3% of sodium chloride, 0.02% of calcium carbonate, and 100% of pure water, preparing a ganoderma lucidum culture medium, and subpackaging in 250mL conical flasks, wherein the liquid filling amount of each flask is 100 mL. Respectively placing the culture medium in a sterilization pot, sterilizing at 121 deg.C for 20min, and cooling in a super clean bench after sterilization; selecting mycelium from the ganoderma lucidum slant, and inoculating the mycelium into a ganoderma lucidum culture medium; culturing the inoculated culture medium in a constant temperature shaking table at the culture temperature of 30 ℃ and the rotation speed of the shaking table of 150rpm for 3d to obtain ganoderma lucidum seed liquid;
preparing a bidirectional fermentation culture medium: 0.2% of rhizoma polygonati, 2% of starch, 1.5% of yeast powder, 0.3% of sodium chloride, 0.02% of calcium carbonate, and 100% of water are added to prepare a bidirectional fermentation culture medium, the initial pH value is 5.5, the bidirectional fermentation culture medium is subpackaged in 250mL conical flasks, the liquid filling amount of each flask is 100mL, and each culture medium is 3 flasks. Respectively placing the two-way fermentation culture medium in a sterilization pot, sterilizing at 121 deg.C for 20min, and cooling in a super clean bench after sterilization;
inoculating and fermenting: inoculating lucid Ganoderma seed liquid according to the inoculation amount of 10%, then placing the culture medium in a constant-temperature shaking table for culturing at the culture temperature of 30 ℃ and the rotation speed of the shaking table of 150rpm for 3d, wherein each group of samples comprises 3 bottles;
fifthly, post-treatment of fermentation liquor: and (4) after the fermentation is finished, sterilizing the fermentation liquor for 20min at 121 ℃. After sterilization, the fermentation broth was filtered, 30% glycerol, 1% hexylene glycol, 2% pentanediol were added, and the mixture was quenched at 95 ℃ for 40min, and cooled to obtain comparative example 9.
Comparative example 10
Pretreating raw materials: grinding tuberose, and sieving with a 10-mesh sieve;
② activating ganoderma lucidum: 2% of starch, 1.5% of yeast powder, 0.3% of sodium chloride, 0.02% of calcium carbonate, and 100% of pure water, preparing a ganoderma lucidum culture medium, and subpackaging in 250mL conical flasks, wherein the liquid filling amount of each flask is 100 mL. Respectively placing the culture medium in a sterilization pot, sterilizing at 121 deg.C for 20min, and cooling in a super clean bench after sterilization; selecting mycelium from the ganoderma lucidum slant, and inoculating the mycelium into a ganoderma lucidum culture medium; culturing the inoculated culture medium in a constant temperature shaking table at the culture temperature of 30 ℃ and the rotation speed of the shaking table of 150rpm for 3d to obtain ganoderma lucidum seed liquid;
preparing a bidirectional fermentation culture medium: 0.2% of tuberose, 2% of starch, 1.5% of yeast powder, 0.3% of sodium chloride, 0.02% of calcium carbonate, and adding water to 100% to prepare a bidirectional fermentation culture medium, wherein the initial pH value is 5.5, the bidirectional fermentation culture medium is subpackaged in 250mL conical flasks, the liquid filling amount of each flask is 100mL, and each culture medium is 3 flasks. Respectively placing the two-way fermentation culture medium in a sterilization pot, sterilizing at 121 deg.C for 20min, and cooling in a super clean bench after sterilization;
inoculating and fermenting: inoculating lucid Ganoderma seed liquid according to the inoculation amount of 10%, then placing the culture medium in a constant-temperature shaking table for culturing at the culture temperature of 30 ℃ and the rotation speed of the shaking table of 150rpm for 3d, wherein each group of samples comprises 3 bottles;
fifthly, post-treatment of fermentation liquor: and (4) after the fermentation is finished, sterilizing the fermentation liquor for 20min at 121 ℃. After sterilization, the fermentation broth was filtered, 30% glycerol, 1% hexylene glycol, 2% pentanediol were added, and the mixture was quenched at 95 ℃ for 40min, and cooled to obtain comparative example 10.
1. Active substance content detection
Detection method of total flavone
Using NaNO2-Al(NO3)3In colorimetric test samplesThe content of flavone refers to a spectrophotometric method for determining the content of total flavone in propolis in GB/T20574-2006.
② polysaccharide detection method
95 percent ethanol is adopted for precipitation, and after the precipitate is dissolved, the content of polysaccharide is detected by adopting a phenol-sulfuric acid method, which refers to aloe juice and powder for cosmetics QB/T2488-.
2. Non-enzymatic glycosylation assay
One of the main features of skin aging is skin yellowing and the appearance of age spots, mainly due to lipid peroxidation and non-enzymatic glycosylation reactions. The non-enzymatic glycosylation reaction in organisms refers to that under the condition of no enzyme catalysis, the aldehyde group or ketone group of reducing sugar and the amino group of macromolecules such as protein and the like undergo Maillard reaction to generate yellow brown glycosylation end products (AGEs).
By inhibiting the nonenzymatic glycosylation of proteins, the inhibitory effect of substances on the deposition of brown pigment can be reflected. The specific test method is as follows:
0.5mol/L bovine serum albumin solution (BCA) and 20mg/mL fructose solution are mixed in equal volume to obtain BCA-fructose reaction solution. The reagents were added sequentially in the order of Table 7 and mixed by vortexing after addition. After incubation for 5 days at 37 ℃ in the dark, the fluorescence intensity was measured at an excitation wavelength of 370nm and an emission wavelength of 440 nm. The inhibition degree of the sample to be tested on the non-enzymatic glycosylation reaction is calculated according to the Fluorescence intensity (RFU, Relative Fluorescence Unit), and is shown in the formula (1).
Figure DEST_PATH_IMAGE002
TABLE 7 grouping and addition of non-enzymatic glycosylation test experiments
Group of Sample to be tested (μ L) PBS buffer (μ L) BCA-fructose reaction (μ L)
Test group/Positive control + - +
Negative control - + +
Blank group + + -
3. Inhibition of tyrosinase assay
Skin pigmentation is caused by the accumulation of melanin in the epidermis, and excessive melanin synthesis causes skin pigmentation, formation of spots, and dullness of the skin. Generally, the higher the tyrosinase activity, the more melanin synthesis. By inhibiting tyrosinase activity, the rate of melanin synthesis can be controlled. The whitening effect of the substances applied to cosmetics was evaluated by the tyrosinase activity inhibition rate, and the experimental reaction system is shown in table 8. After the sample addition, incubation was carried out at 25 ℃ for 30min, and the absorbance was measured at 475 nm.
TABLE 8 tyrosinase reaction System
Test tube numbering PBS buffer (μ L) L-tyrosine (mu L) sample/Positive control (μ L) Tyrosinase (μ L)
A + + - +
B + + - -
C + + + -
D + + + -
The tyrosinase inhibition rate is calculated according to the formula (2):
Figure DEST_PATH_IMAGE004
in formula (2): A. b, C, D is the absorbance of negative group, the blank absorbance of negative control group, the absorbance of sample group/positive group, and the blank absorbance of sample group/positive group.
4. Clock gene clock expression assay
In nature, all life activities from single cells to higher animals and plants and human beings have a periodic life activity phenomenon which runs according to a certain rule, and is called biological rhythm. It is now believed that a time control mechanism system, called a biological clock system, widely exists in natural organisms to control and regulate various rhythmic activities and maintain periodic oscillations in the internal environment of the organism. The molecular elements of biological clocks are composed of a series of core Clock components, including genes such as Clock, Bmal1, Per, Cry, Tim, etc., and their related protein products. UVB-induced DNA damage repair responses are also reported to exhibit clock-dependent rhythmic changes. In addition, circadian clocks are involved in regulating multiple aspects of skin homeostasis such as: hair growth, cell proliferation, stem cell function, carcinogenesis, aging, immunity, and the like.
Clock gene Clock expression experiment, the concrete operation is as follows:
based on the detection of cytotoxicity
The test sample is set with 8 concentrations, each concentration is set with 3 repeated holes, and meanwhile, the test is set with a negative control (culture medium), a zero setting hole (deionized water) and a positive control (culture medium containing 5% DMSO (dimethyl sulfoxide)). The assay employs a MTT activity assay to screen the maximum safe concentration for cell administration. The specific operation steps are as follows:
a. cell inoculation: by 1 × 104Cell/well inoculation Density cells were plated onto 96 well plates, incubators (37 ℃, 5% CO)295% Relative Humidity (RH)) overnight.
b. Preparing liquid: test substances were prepared at different concentrations according to the concentration settings in table 9.
TABLE 9 test concentration settings
Sample name Sample concentration (v/v)
Bidirectional fermentation liquor 0.08%、0.16%、0.31%、0.63%、1.25%、2.5%、5%、10%
c. Administration: the administration was carried out when the plating rate of the cells in the 96-well plate reached 40-60%. Adding 200 mu L of culture solution into each well of the solvent control group; adding 200 mu L of culture solution containing 10% DMSO into each well of the positive control group; adding 200 mu L of culture solution containing samples with corresponding concentrations into each well of the sample group; the zero-adjusted group was seeded without cells, and only 200. mu.L of cell culture medium was added. After completion of the administration, the 96-well plate was placed in an incubator (37 ℃ C., 5% CO)295% RH).
d. And (3) detection: after 24h of cell incubation culture, discarding the supernatant, adding MTT working solution (0.5 mg/mL), incubating for 4h at 37 ℃ in a dark place, discarding the supernatant after the incubation is finished, adding 150 muL DMSO into each well, and reading the OD value at 490 nm.
e. Relative cell viability: according to the calculation formula:
Figure DEST_PATH_IMAGE006
② morphological examination
a. Cell inoculation: and (3) setting a sample group and a solvent control group, wherein each group is provided with two multiple holes. The cells were seeded at the corresponding seeding density in 24-well plates, incubators (37 ℃ C., 5% CO)295% RH) overnight.
b. Preparing liquid: according to the MTT detection result, determining the morphological observation concentration of the detection sample, and preparing the working solution of the test object with different concentrations.
c. Administration: when the cell plating rate of the 24-pore plate reaches 40%, the drug is administered, the samples are added with the test substances with different concentrations, the solvent control group is added with the cell culture solution, and the incubator (37 ℃, 5% CO)295% RH) for 24 h.
d. And (3) cell observation: after incubation, cell morphology was observed under a microscope and photographed.
③ Gene detection
a. Cell inoculation: by 2.5X 105Cell/well inoculation Density cells were plated onto 6 well plates, incubators (37 ℃, 5% CO)295% RH) overnight.
b. Administration: grouping according to the experiment of table 10, when the plating rate of the cells in the 6-well plate reaches 40-60%, the administration is carried out in groups, each well is loaded with 2mL, and each group is provided with 3 multiple wells. After completion of the administration, the 6-well plate was placed in an incubator (37 ℃ C., 5% CO)295% RH) for 24 h.
c. Collecting cells: after 24h incubation, cell supernatants were discarded, washed twice with 1 mL/well PBS, 1mL RNAioso Plus was added to each well, lysed cells were aspirated, and samples were collected.
d. And (3) gene expression detection: extracting RNA, reverse transcribing to cDNA, and fluorescent quantitative PCR detection with 2-△△CTThe method performs result calculation
e. And (4) analyzing results: the plots were generated using GraphPad Prism Program software and statistically analyzed between groups using t-test.
TABLE 10 test grouping
Figure DEST_PATH_IMAGE008
5. Analysis of test results
5.1 active ingredient content
TABLE 11 comparison of active ingredients in samples
Sample numbering Polysaccharide content (mg/mL) Flavone content (mg/mL)
Sample 1 5.11 0.15
Sample 2 5.45 0.16
Sample 3 5.23 0.13
Sample No. 4 5.83 0.18
Sample No. 5 5.98 0.21
Sample No. 6 5.79 0.15
Sample 7 5.38 0.12
Sample 8 5.55 0.15
Sample 9 5.21 0.11
Sample 10 5.17 0.12
Sample 11 5.32 0.16
Sample 12 5.89 0.19
Sample 13 5.78 0.18
Sample 14 5.68 0.14
Sample 15 5.85 0.18
Sample 16 5.74 0.21
Sample 17 5.33 0.23
Sample 18 5.68 0.14
Sample 19 5.82 0.15
Sample 20 5.74 0.17
Sample 21 5.32 0.15
Sample 22 5.79 0.14
Sample 23 5.13 0.11
Comparative example 1 3.11 0.11
Comparative example 2 2.64 0.17
Comparative example 3 2.73 0.13
The polysaccharide content and the flavone content are taken as research targets, the plant composition is prepared by adopting a bidirectional fermentation process, the influence of fermentation raw materials, fermentation inoculum size (%), fermentation time (d), fermentation temperature (DEG C) and fermentation pH value on the polysaccharide content and the flavone content in the plant composition is considered, and the optimized preparation process comprises the following steps: 1-10% of dendrobium officinale, 0.01-1% of rhizoma polygonati and 0.01-1% of tuberose by weight of the bidirectional fermentation medium; 1-15% of fermentation inoculation amount; fermenting for 1-7 days at 25-35 deg.C and pH3.5-7.5, wherein the plant composition contains polysaccharide 5.11-5.98mg/mL and flavone 0.11-0.23 mg/mL.
The plant composition is prepared by a bidirectional fermentation process, the dendrobium officinale, the polygonatum and the tuberose are combined according to a specific proportion and then subjected to bidirectional fermentation by adopting the fungus lucid ganoderma which is homologous in medicine and food, and the dendrobium officinale, the polygonatum and the tuberose are subjected to deep fermentation decomposition into a small molecular carbon source by utilizing a strong enzyme system of the fungus for decomposing substances such as cellulose and the like, so that the utilization rate of active ingredients of plant raw materials is increased, and the active ingredient polysaccharide of the plant composition is improved.
5.2 non-enzymatic glycosylation assay
TABLE 12 inhibition of nonenzymatic glycosylation of the samples
Sample numbering Non-enzymatic glycosylation inhibition ratio (%)
2.5% example 6 73
2.5% example 7 67
2.5% example 8 75
2.5% example 9 74
2.5% comparative example 4 36
2.5% comparative example 5 32
2.5% comparative example 6 24
2.5% comparative example 7 31
2.5% comparative example 8 22
2.5% comparative example 9 24
2.5% comparative example 10 21
0.5% Vc ethyl ether 58
The results of the non-enzymatic glycosylation test are shown in table 12, and the plant compositions prepared in examples 6 to 9 of the present invention have higher inhibition rate for non-enzymatic glycosylation than the plant compositions obtained in comparative examples 4 to 10 and 0.5% of Vc ethyl ether (positive control) at the same addition amount (2.5%), which indicates that the plant compositions of the present invention can inhibit the non-enzymatic glycosylation better than the fermentation products obtained by any two-way fermentation of the ganoderma lucidum from a single raw material and the fermentation products obtained by any two-way fermentation of the ganoderma lucidum from any two combined raw materials. The test results prove that the dendrobium officinale, the polygonatum sibiricum and the tuberose have obvious synergistic effect after being subjected to bidirectional fermentation by the ganoderma lucidum according to a specific proportion, and compared with the combination or dosage range provided in a comparative example, the combination and proportion provided by the application can obviously increase the inhibition effect on non-enzymatic glycosylation reaction after being treated by a bidirectional fermentation process of the ganoderma lucidum.
5.3 tyrosinase assay
TABLE 13 tyrosinase inhibition rate of the samples
Sample numbering Tyrosinase inhibition (%)
15% of example 6 63
15% example 7 59
15% of example 8 64
15% of example 9 65
15% comparative example 4 23
15% comparative example 5 19
15% comparative example 6 11
15% comparative example 7 18
15% comparative example 8 14
15% comparative example 9 13
15% comparative example 10 12
0.075% alpha-arbutin 87
The tyrosinase test results are shown in table 13, and compared with comparative examples 4-10, the plant compositions prepared in examples 6-9 of the present invention have a good tyrosinase activity inhibiting effect at the same addition amount (15%), and the tyrosinase activity inhibiting effect is lower than that of α -arbutin (positive control). The plant composition can better inhibit the activity of tyrosinase and has better whitening effect than a fermentation product obtained by any ganoderma lucidum bidirectional fermentation single raw material and a fermentation product obtained by any two combined raw materials of the ganoderma lucidum bidirectional fermentation. The test results prove that the dendrobium officinale, the polygonatum sibiricum and the tuberose have obvious synergistic effect after being subjected to bidirectional fermentation by the ganoderma lucidum according to a specific proportion, and compared with the combination or dosage range provided in a comparative example, the combination and proportion provided by the application can obviously increase the inhibition effect on the tyrosinase activity after being treated by a bidirectional fermentation process of the ganoderma lucidum.
5.4 clock Gene expression assay
TABLE 14 summary of results for Clock genes
Sample name Mean value (Mean) Standard Deviation (SD) Significance of difference: (p-value)
Blank control 1.00 0.07 /
0.025% example 6 0.88 0.02 0.043*
0.05% of example 6 0.84 0.03 0.023*
0.075% of example 6 0.77 0.04 0.007**
0.025% comparative example 6 1.04 0.04 0.032*
0.05% comparative example 6 1.11 0.05 0.028*
0.075% of comparative example 6 1.17 0.03 0.025*
As shown in table 14 and fig. 1, compared with the blank control, the example 6 has a good effect of inhibiting the expression of the Clock gene in the range of 0.025% to 0.075%, and the effect is significantly better than that of the comparative example 6, which shows that the plant composition of the present invention has a better effect of inhibiting the expression of the Clock gene than the fermentation product obtained by ganoderma lucidum bi-directionally fermenting dendrobium officinale. The test results prove that the dendrobium officinale, the polygonatum and the tuberose have obvious synergistic effect after being subjected to bidirectional fermentation by the ganoderma lucidum according to a specific proportion, and compared with the fermentation product of the dendrobium officinale provided in the comparative example 6, the combination and proportion of the application can obviously increase the inhibition effect on clock gene expression after being treated by the bidirectional fermentation process of the ganoderma lucidum.
According to the invention, dendrobium officinale, rhizoma polygonati and tuberose are combined in a specific ratio and then are inoculated with ganoderma lucidum to be subjected to bidirectional fermentation to obtain a fermentation product with the effects of inhibiting non-enzymatic glycosylation reaction and inhibiting Clock gene expression, and the fermentation product can also inhibit tyrosinase activity and has double effects of resisting aging and whitening.

Claims (10)

1. A plant composition is characterized in that the addition amount of dendrobium officinale in the plant composition is 1-10%, the addition amount of rhizoma polygonati is 0.01-1%, and the addition amount of tuberose is 0.01-1% by weight of a bidirectional fermentation medium;
the plant composition is prepared by a bidirectional fermentation process.
2. The plant composition of claim 1, wherein the plant composition has a polysaccharide content of 5.11-5.98mg/mL and a flavone content of 0.11-0.23 mg/mL.
3. The plant composition according to claim 1 or 2, wherein the plant composition is prepared by a preparation method comprising the steps of:
pretreating raw materials: pulverizing herba Dendrobii, rhizoma Polygonati, and tuberose respectively, and sieving;
② activating ganoderma lucidum: preparing a ganoderma lucidum culture medium, inoculating ganoderma lucidum, and culturing to obtain ganoderma lucidum seed liquid;
preparing a bidirectional fermentation culture medium;
inoculating and fermenting: inoculating the ganoderma lucidum seed liquid into a bidirectional fermentation culture medium for culture;
fifthly, post-treatment of fermentation liquor: sterilizing and filtering.
4. The plant composition according to claim 3, wherein the culture medium for the activated ganoderma lucidum in step (II) of the preparation method of the plant composition comprises 0.5-5% of starch, 0.1-5% of yeast powder, 0.1-2% of sodium chloride, 0.01-0.1% of calcium carbonate, and water to 100%; the culture conditions of the ganoderma lucidum are 25-35 ℃, 50-300rpm, and the ganoderma lucidum is cultured for 50-80 h.
5. The plant composition according to claim 3, wherein the bidirectional fermentation medium in step (c) of the preparation method of the plant composition is 0.5-5% of starch, 0.1-5% of yeast powder, 0.1-2% of sodium chloride, 0.01-0.1% of calcium carbonate, 1-10% of dendrobium officinale, 0.01-1% of rhizoma polygonati, 0.01-1% of tuberose and water to 100%.
6. The plant composition according to claim 3, wherein the inoculation amount of the ganoderma lucidum seed solution in the step (iv) in the preparation method of the plant composition is 1-15%.
7. The plant composition according to claim 3, wherein the bidirectional fermentation culture conditions in the step (iv) of the preparation method of the plant composition are 25-35 ℃, 50-300rpm, and the culture time is 1-7 days.
8. A method of preparing a botanical composition, said method comprising the steps of:
pretreating raw materials: pulverizing herba Dendrobii, rhizoma Polygonati, and tuberose respectively, and sieving;
② activating ganoderma lucidum: adding 0.5-5% of starch, 0.1-5% of yeast powder, 0.1-2% of sodium chloride and 0.01-0.1% of calcium carbonate into 100% of water, mixing, subpackaging and sterilizing to prepare a ganoderma lucidum culture medium; selecting Ganoderma mycelia, inoculating to culture medium, and culturing at 25-35 deg.C and 50-300rpm for 50-80 hr to obtain Ganoderma seed solution;
preparing a bidirectional fermentation culture medium: mixing starch 0.5-5% and yeast powder 0.1-5%; 0.1-2% of sodium chloride, 0.01-0.1% of calcium carbonate, 1-10% of dendrobium officinale, 0.01-1% of rhizoma polygonati, 0.01-1% of tuberose, and water which is added to 100% are mixed, subpackaged and sterilized to prepare a bidirectional fermentation culture medium;
inoculating and fermenting: inoculating the ganoderma lucidum seed liquid obtained in the step (II) to a culture medium according to the inoculation amount of 1-15%, and then culturing for 1-7d at the temperature of 25-35 ℃ and under the condition of 50-300 rpm;
fifthly, post-treatment of fermentation liquor: sterilizing and fine filtering.
9. Use of the plant composition according to any one of claims 1 to 7 or of the plant composition prepared by the preparation process according to claim 8 in skin care and/or cosmetics.
10. Use of the plant composition according to any one of claims 1 to 7 or the plant composition prepared by the preparation method according to claim 8 in the preparation of skin care products and/or cosmetics with anti-aging and/or whitening effects.
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Citations (7)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US20060018867A1 (en) * 2004-05-12 2006-01-26 Ichimaru Pharcos Co., Ltd Cosmetic composition and production thereof
CN104593425A (en) * 2015-02-03 2015-05-06 周礼红 Dendrobium monascus preparation method
CN105395458A (en) * 2015-12-15 2016-03-16 上海相宜本草化妆品股份有限公司 Herbal composition and application thereof
CN108567912A (en) * 2018-07-17 2018-09-25 上海家化联合股份有限公司 A kind of Chinese medical extract and its enzymolysis and tunning
CN109528568A (en) * 2018-12-18 2019-03-29 北京工商大学 A kind of composition and preparation method thereof with anti-aging and white-skinned face function
CN110755344A (en) * 2019-10-25 2020-02-07 北京京瑜科技有限公司 Ganoderma lucidum-rhizoma polygonati bidirectional fermentation process and composition
CN112245538A (en) * 2020-10-28 2021-01-22 广州环亚化妆品科技有限公司 Traditional Chinese medicine composition, traditional Chinese medicine fermentation product containing traditional Chinese medicine composition, and preparation method and application of traditional Chinese medicine fermentation product

Patent Citations (7)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US20060018867A1 (en) * 2004-05-12 2006-01-26 Ichimaru Pharcos Co., Ltd Cosmetic composition and production thereof
CN104593425A (en) * 2015-02-03 2015-05-06 周礼红 Dendrobium monascus preparation method
CN105395458A (en) * 2015-12-15 2016-03-16 上海相宜本草化妆品股份有限公司 Herbal composition and application thereof
CN108567912A (en) * 2018-07-17 2018-09-25 上海家化联合股份有限公司 A kind of Chinese medical extract and its enzymolysis and tunning
CN109528568A (en) * 2018-12-18 2019-03-29 北京工商大学 A kind of composition and preparation method thereof with anti-aging and white-skinned face function
CN110755344A (en) * 2019-10-25 2020-02-07 北京京瑜科技有限公司 Ganoderma lucidum-rhizoma polygonati bidirectional fermentation process and composition
CN112245538A (en) * 2020-10-28 2021-01-22 广州环亚化妆品科技有限公司 Traditional Chinese medicine composition, traditional Chinese medicine fermentation product containing traditional Chinese medicine composition, and preparation method and application of traditional Chinese medicine fermentation product

Non-Patent Citations (1)

* Cited by examiner, † Cited by third party
Title
左锦辉等: "中药双向发酵技术在化妆品中的应用", 《日用化学工业》 *

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