CN117257681A - Rhodiola rosea extract extraction method, rhodiola rosea extract and application of rhodiola rosea extract - Google Patents
Rhodiola rosea extract extraction method, rhodiola rosea extract and application of rhodiola rosea extract Download PDFInfo
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K8/00—Cosmetics or similar toiletry preparations
- A61K8/18—Cosmetics or similar toiletry preparations characterised by the composition
- A61K8/96—Cosmetics or similar toiletry preparations characterised by the composition containing materials, or derivatives thereof of undetermined constitution
- A61K8/97—Cosmetics or similar toiletry preparations characterised by the composition containing materials, or derivatives thereof of undetermined constitution from algae, fungi, lichens or plants; from derivatives thereof
- A61K8/9783—Angiosperms [Magnoliophyta]
- A61K8/9789—Magnoliopsida [dicotyledons]
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- A61Q17/04—Topical preparations for affording protection against sunlight or other radiation; Topical sun tanning preparations
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- A—HUMAN NECESSITIES
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- A61Q19/02—Preparations for care of the skin for chemically bleaching or whitening the skin
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- A—HUMAN NECESSITIES
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Abstract
The invention provides an extraction method of rhodiola rosea extract, the extract and application thereof. The method for extracting the rhodiola rosea extract provided by the invention aims at the characteristics of rhodiola crenulata, and can effectively extract the effective components of rhodiola crenulata; the rhodiola rosea extract prepared and extracted by the invention has the effects of moisturizing, whitening, anti-wrinkle, antioxidation, sun protection and the like; the invention adopts the compound microbial inoculum to ferment the rhodiola rosea extracted cells, can obviously improve the moisturizing, antioxidation and repairing performances of the rhodiola rosea, and has good application prospect in the repairing field. The extract prepared by the method provided by the invention can realize long-acting moisturizing when applied to the fields of facial masks, moisturizing creams and the like.
Description
Technical Field
The invention relates to the technical field of daily chemicals, in particular to an extraction method of rhodiola rosea extract, the extract and application thereof.
Background
Rhodiola rosea is a plant of rhodiola genus of Crassulaceae family, is an important herb and food homologous plant commonly used in mountain or cold areas, and is called "mountain ginseng" and "Gao Shanxian grass". The rhodiola root herb can be used as medicine, and has sweet and bitter taste, mild nature, and lung and heart meridian entered. Has effects of invigorating qi, promoting blood circulation, dredging collaterals, and relieving asthma. In high altitude areas, rhodiola rosea is often used to alleviate altitude stress and to relieve fatigue.
The field growth environment of rhodiola rosea is extremely severe and variable, such as hypoxia, low-temperature drying, strong ultraviolet radiation, large day-night temperature difference and the like. The rhodiola rosea is genetically adapted to severe cold and variable dangerous environments, and is also provided with special adaptive substances which are rare for other plants. Rhodiola crenulata is the only rhodiola crenulata variety recorded in Chinese pharmacopoeia 2005.
Up to now, 180 kinds of compounds are identified from rhodiola crenulata, including a large amount of glycosides, flavonoids, volatile oils, tannins, cyanogenic glycosides, etc. The current method for extracting plant components is to dissolve the components in the plant by a solvent so as to extract the effective components in the plant. However, the components obtained by the method are complex and uncontrollable, the specificity is poor, the content of the effective components is low, and the skin absorbability is poor.
Aiming at the phenomenon, the invention adopts a novel rhodiola root extraction method.
Disclosure of Invention
The invention provides an extraction method of rhodiola rosea extract, which comprises the following steps:
s01, pretreatment: washing the cultivated rhodiola rosea young roots with flowing sterile water for 1h, and then slicing the rhodiola rosea young roots on a sterile super clean bench by using a sterile blade; then cleaning the sliced rhodiola rosea young root slices with sterile water for 2min;
s02, inducing callus: inoculating the S01 cleaned rhodiola rosea young root slices into a tissue culture bottle containing hormone for inducing callus;
induction conditions: dark culturing at 25+ -1deg.C for 24 hr, and adding light for 8 hr every day; after 2 weeks, the edges of the materials grow massive callus; cutting off the callus after 3 weeks of culture;
s03, subculturing the callus: transferring the callus with loose upper part, good state and no browning into a secondary culture medium, and performing dark culture at 25+/-1 ℃ for 24 hours, and then illuminating for 8 hours per day with illumination intensity of 1500Lx;
s04, shake flask culture: inoculating the callus subjected to the secondary culture for three times into a liquid culture medium, performing shake culture at 100rpm with the inoculum size of one third, performing dark culture for 24 hours, and performing illumination culture for 8 hours every day at the culture temperature of 25+/-1 ℃;
s05, culturing in a bioreactor: inoculating liquid suspension cells cultured in a shake flask as seed cells, wherein the inoculation volume is 15% of the volume of a culture medium for culture;
the culture conditions are as follows: the initial ventilation is 50L/h, the stirring speed is 100rpm, the culture temperature is 26+/-1 ℃, and the dark culture is carried out for 24 hours after inoculation; then illumination is carried out for 8 hours every day, and the illumination intensity is 1500Lx; and simultaneously changing the stirring speed to 140rpm, and continuously culturing for 2 weeks;
s06, fermenting and culturing: filtering the culture solution in the step S05, and then inoculating the culture solution into a fermentation medium for fermentation.
Preferably, the medium in the step S02 comprises the following components:
MS fixed media (7% agar), 0.5 mg/L6-benzyladenine, 6.5mg/L a-naphthylacetic acid, 40g/L sucrose, 0.2mg/L indoleacetic acid, natural pH.
Preferably, the medium in the step S03 comprises the following components:
MS fixed media (7% agar), 0.2mg/L vitamin A,6.0mg/L a-Naacetic acid, 35g/L glucose, 6mg/L PVP, natural pH.
Preferably, the medium composition in step S04 is:
MS liquid medium (7% agar), 6.0mg/L a-Naacetic acid, 50g/L sucrose, 0.2mg/L glutamine, 5mg/LPVP, natural pH.
Preferably, the strain used in step S06 is: pentose milk plant stems and lactobacillus terreus.
In another aspect, the invention provides a rhodiola rosea extract, which is prepared by the rhodiola rosea extraction method.
In another aspect, the invention provides the rhodiola rosea extract for protecting or enhancing skin barrier function.
In another aspect, the invention provides the rhodiola rosea extract, which is used for protecting skin cells from apoptosis caused by ultraviolet irradiation.
In another aspect, the invention provides the rhodiola rosea extract for whitening skin.
In another aspect, the invention provides the rhodiola rosea extract which is used for resisting aging or oxidization.
By adopting the technical scheme, the invention has the following beneficial effects:
1. the method for extracting the rhodiola rosea extract provided by the invention aims at the characteristics of rhodiola crenulata, and can effectively extract the effective components of rhodiola crenulata;
2. the rhodiola rosea extract prepared and extracted by the invention has the effects of moisturizing, whitening, anti-wrinkle, antioxidation, sun protection and the like;
3. the invention adopts the compound microbial inoculum to ferment the rhodiola rosea extracted cells, can obviously improve the moisturizing, antioxidation and repairing performances of the rhodiola rosea, and has good application prospect in the repairing field. The extract prepared by the method provided by the invention can realize long-acting moisturizing when applied to the fields of facial masks, moisturizing creams and the like.
Drawings
In order to more clearly illustrate the embodiments of the invention or the technical solutions in the prior art, the drawings which are required in the description of the embodiments or the prior art will be briefly described, it being obvious that the drawings in the description below are some embodiments of the invention and that other drawings may be obtained from these drawings without inventive effort for a person skilled in the art.
FIG. 1 is a graph showing the experimental effect of human body efficacy;
fig. 2 is a graph showing the experimental effect of human body efficacy.
Detailed Description
The following description of the present invention will be made clearly and fully, and it is apparent that the embodiments described are some, but not all, of the embodiments of the present invention. All other embodiments, which can be made by those skilled in the art based on the embodiments of the invention without making any inventive effort, are intended to be within the scope of the invention.
The invention provides an extraction method of rhodiola rosea extract, which comprises the following steps:
s01, pretreatment: washing the cultivated rhodiola rosea young roots with flowing sterile water for 1h, and then slicing the rhodiola rosea young roots on a sterile super clean bench by using a sterile blade; then cleaning the sliced rhodiola rosea young root slices with sterile water for 2min;
s02, inducing callus: inoculating the S01 cleaned rhodiola rosea young root slices into a tissue culture bottle containing hormone for inducing callus;
induction conditions: dark culturing at 25+ -1deg.C for 24 hr, and adding light for 8 hr every day; after 2 weeks, the edges of the materials grow massive callus; cutting off the callus after 3 weeks of culture;
s03, subculturing the callus: transferring the callus with loose upper part, good state and no browning into a secondary culture medium, and performing dark culture at 25+/-1 ℃ for 24 hours, and then illuminating for 8 hours per day with illumination intensity of 1500Lx;
s04, shake flask culture: inoculating the callus subjected to the secondary culture for three times into a liquid culture medium, performing shake culture at 100rpm with the inoculum size of one third, performing dark culture for 24 hours, and performing illumination culture for 8 hours every day at the culture temperature of 25+/-1 ℃;
s05, culturing in a bioreactor: inoculating liquid suspension cells cultured in a shake flask as seed cells, wherein the inoculation volume is 15% of the volume of a culture medium for culture;
the culture conditions are as follows: the initial ventilation is 50L/h, the stirring speed is 100rpm, the culture temperature is 26+/-1 ℃, and the dark culture is carried out for 24 hours after inoculation; then illumination is carried out for 8 hours every day, and the illumination intensity is 1500Lx; and simultaneously changing the stirring speed to 140rpm, and continuously culturing for 2 weeks;
s06, fermenting and culturing: and (3) inoculating the culture solution obtained in the step S05 into a fermentation medium for fermentation.
Preferably, the medium in the step S02 comprises the following components:
MS fixed media (7% agar), 0.5 mg/L6-benzyladenine, 6.5mg/L a-naphthylacetic acid, 40g/L sucrose, 0.2mg/L indoleacetic acid, natural pH.
Preferably, the medium in the step S03 comprises the following components:
MS fixed media (7% agar), 0.2mg/L vitamin A,6.0mg/L a-Naacetic acid, 35g/L glucose, 6mg/L PVP, natural pH.
Preferably, the medium composition in step S04 is:
MS liquid medium (7% agar), 6.0mg/L a-Naacetic acid, 50g/L sucrose, 0.2mg/L glutamine, 5mg/LPVP, natural pH.
Preferably, the medium in the step S05 comprises the following components: MS liquid culture medium (7% agar), 0.6 mg/L6-benzyladenine, 6.0mg/L a-naphthylacetic acid, 30g/L sucrose, 0.2mg/L glutamine, 5mg/L PVP, natural pH.
Preferably, the strain used in step S06 is: pentose milk plant stems and lactobacillus terreus.
The specific step S06 is as follows:
1mL of Lactobacillus pentosus solution (10) 6 CFU/mL), 2mL of Lactobacillus terreus bacterial liquid (10) 6 CFU/mL) was inoculated into the culture broth of 300mLS step 05 (inoculum size 1%), fermented in a fermenter at 29 ℃ for 3 days at 110rpm, followed by filtration to obtain rhodiola crenulata extract.
In another aspect, the invention provides a rhodiola rosea extract, which is prepared by the rhodiola rosea extraction method.
In another aspect, the invention provides the rhodiola rosea extract for protecting or enhancing skin barrier function.
In another aspect, the invention provides the rhodiola rosea extract, which is used for protecting skin cells from apoptosis caused by ultraviolet irradiation.
In another aspect, the invention provides the rhodiola rosea extract for whitening skin.
In another aspect, the invention provides the rhodiola rosea extract which is used for resisting aging or oxidization.
In the invention, the rhodiola rosea extract prepared by the method is a rhodiola rosea root extract, namely RHODIOLA CRENULATA.
The technical solutions provided by the present invention are described in detail below with reference to examples, but they should not be construed as limiting the scope of the present invention.
The lactobacillus pentosus Lactiplantibacillus pentosus CICC percent 24333, the lactobacillus terreus Sporolactobacillus terrae CICC percent 10726 and the saccharomyces cerevisiae Saccharomyces cerevisiae CICC percent 32883 are purchased from the China industry microbiological culture collection center.
Example 1:
the embodiment provides an extraction method of rhodiola rosea extract, which comprises the following steps:
s01, pretreatment: washing the cultivated rhodiola rosea young roots with flowing sterile water for 1h, and then slicing the rhodiola rosea young roots on a sterile super clean bench by using a sterile blade; then cleaning the sliced rhodiola rosea young root slices with sterile water for 2min;
s02, inducing callus: inoculating the S01 cleaned rhodiola rosea young root slices into a tissue culture bottle containing hormone for inducing callus;
induction conditions: dark culturing at 25+ -1deg.C for 24 hr, and adding light for 8 hr every day; after 2 weeks, the edges of the materials grow massive callus; cutting off the callus after 3 weeks of culture;
the culture medium comprises the following components:
MS fixed media (7% agar), 0.5 mg/L6-benzyladenine, 6.5mg/L a-naphthylacetic acid, 40g/L sucrose, 0.2mg/L indoleacetic acid, natural pH.
S03, subculturing the callus: transferring the callus with loose upper part, good state and no browning into a secondary culture medium, and performing dark culture at 25+/-1 ℃ for 24 hours, and then illuminating for 8 hours per day with illumination intensity of 1500Lx;
the culture medium comprises the following components:
MS fixed media (7% agar), 0.2mg/L vitamin A,6.0mg/L a-Naacetic acid, 35g/L glucose, 6mg/L PVP, natural pH.
S04, shake flask culture: inoculating the callus subjected to the secondary culture for three times into a liquid culture medium, performing shake culture at 100rpm with the inoculum size of one third, performing dark culture for 24 hours, and performing illumination culture for 8 hours every day at the culture temperature of 25+/-1 ℃;
the culture medium comprises the following components:
MS liquid medium (7% agar), 6.0mg/L a-Naacetic acid, 50g/L sucrose, 0.2mg/L glutamine, 5mg/LPVP, natural pH.
S05, culturing in a bioreactor: inoculating liquid suspension cells cultured in a shake flask as seed cells, wherein the inoculation volume is 15% of the volume of a culture medium for culture;
the culture medium comprises the following components: MS liquid culture medium (7% agar), 0.6 mg/L6-benzyladenine, 6.0mg/L a-naphthylacetic acid, 30g/L sucrose, 0.2mg/L glutamine, 5mg/L PVP, natural pH.
The culture conditions are as follows: the initial ventilation is 50L/h, the stirring speed is 100rpm, the culture temperature is 26+/-1 ℃, and the dark culture is carried out for 24 hours after inoculation; then illumination is carried out for 8 hours every day, and the illumination intensity is 1500Lx; and simultaneously changing the stirring speed to 140rpm, and continuously culturing for 2 weeks;
s06, fermenting and culturing: 1mL of Lactobacillus pentosus solution (10) 6 CFU/mL), 2mL of Lactobacillus terreus bacterial liquid (10) 6 CFU/mL) was inoculated into the culture broth of 300mLS step 05 (inoculum size 1%), fermented in a fermenter at 29 ℃ for 3 days at 110rpm, followed by filtration to obtain rhodiola crenulata extract.
Example 2:
the embodiment provides an extraction method of rhodiola rosea extract, which comprises the following steps:
s01, pretreatment: washing the cultivated rhodiola rosea young roots with flowing sterile water for 1h, and then slicing the rhodiola rosea young roots on a sterile super clean bench by using a sterile blade; then cleaning the sliced rhodiola rosea young root slices with sterile water for 2min;
s02, inducing callus: inoculating the S01 cleaned rhodiola rosea young root slices into a tissue culture bottle containing hormone for inducing callus;
induction conditions: dark culturing at 25+ -1deg.C for 24 hr, and adding light for 8 hr every day; after 2 weeks, the edges of the materials grow massive callus; cutting off the callus after 3 weeks of culture;
the culture medium comprises the following components:
MS fixed media (7% agar), 0.5 mg/L6-benzyladenine, 6.5mg/L a-naphthylacetic acid, 40g/L sucrose, 0.2mg/L indoleacetic acid, natural pH.
S03, subculturing the callus: transferring the callus with loose upper part, good state and no browning into a secondary culture medium, and performing dark culture at 25+/-1 ℃ for 24 hours, and then illuminating for 8 hours per day with illumination intensity of 1500Lx;
the culture medium comprises the following components:
MS fixed media (7% agar), 0.2mg/L vitamin A,6.0mg/L a-Naacetic acid, 35g/L glucose, 6mg/L PVP, natural pH.
S04, shake flask culture: inoculating the callus subjected to the secondary culture for three times into a liquid culture medium, performing shake culture at 100rpm with the inoculum size of one third, performing dark culture for 24 hours, and performing illumination culture for 8 hours every day at the culture temperature of 25+/-1 ℃;
the culture medium comprises the following components:
MS liquid medium (7% agar), 6.0mg/L a-Naacetic acid, 50g/L sucrose, 0.2mg/L glutamine, 5mg/LPVP, natural pH.
S05, culturing in a bioreactor: inoculating liquid suspension cells cultured in a shake flask as seed cells, wherein the inoculation volume is 15% of the volume of a culture medium for culture;
the culture medium comprises the following components: MS liquid culture medium (7% agar), 0.6 mg/L6-benzyladenine, 6.0mg/L a-naphthylacetic acid, 30g/L sucrose, 0.2mg/L glutamine, 5mg/L PVP, natural pH.
The culture conditions are as follows: the initial ventilation is 50L/h, the stirring speed is 100rpm, the culture temperature is 26+/-1 ℃, and the dark culture is carried out for 24 hours after inoculation; then illumination is carried out for 8 hours every day, and the illumination intensity is 1500Lx; and simultaneously changing the stirring speed to 140rpm, continuously culturing for 2 weeks, and filtering to obtain radix Rhodiolae extract.
Example 3:
the embodiment provides a method for extracting rhodiola rosea extract,
1mL of Lactobacillus pentosus solution (10) 6 CFU/mL), 2mL of Lactobacillus terreus bacterial liquid (10) 6 CFU/mL) was inoculated into 300mL of rhodiola rosea extract (inoculation amount 1%), fermented in a fermenter at 29℃for 3 days at a rotation speed of 110rpm, followed by filtration to obtain rhodiola crenulata extract.
The rhodiola rosea extract is purchased from the biological technology company of the American style of Guangzhou.
Example 4: the difference from example 1 is that step S06 is: 1mL of Lactobacillus pentosus solution (10) 6 CFU/mL) was inoculated into the culture broth of 300mLS step 05 (inoculum size 1%), fermented in a fermenter at 29 ℃ for 3 days at 110rpm, followed by filtration to obtain rhodiola crenulata extract.
Example 5: the difference from example 1 is that step S06 is: 2mL of the Lactobacillus terreus bacterial liquid (10) 6 CFU/mL) was inoculated into the culture broth of 300mLS step 05 (inoculum size 1%), fermented in a fermenter at 29 ℃ for 3 days at 110rpm, followed by filtration to obtain rhodiola crenulata extract.
Example 6: the difference from example 1 is that step S06 is: 2mL of Saccharomyces cerevisiae liquid (10) 6 CFU/mL) was inoculated into the culture broth of 300mLS step 05 (inoculum size 1%), fermented in a fermenter at 29 ℃ for 3 days at 110rpm, followed by filtration to obtain rhodiola crenulata extract.
Performance testing
The rhodiola crenulata extracts prepared in examples 1-6 were subjected to performance testing. Before a specific test, 10% of rhodiola crenulata extract prepared in examples 1-6, 0.1% of EDTA disodium, 0.2% of carbomer and the balance of water are prepared into 1-6 to be tested.
Performance test 1: elastase inhibition test: the anti-wrinkle efficacy of examples 1-6 was tested according to the laboratory method (HMC-W1-028 elastase inhibition), and the results of the elastase inhibition test performed on the test samples and the negative control were compared, and examples 1-6 were considered to have some anti-wrinkle efficacy if the inhibition of the test samples was higher than that of the negative control and there was a significant difference (p < 0.05).
Treatment of control and test samples
Examples 1 to 6: diluting with pure water to a sample concentration of 1%;
positive control (epigallocatechin gallate): diluting with water to a positive control concentration of 0.1%;
negative control: pure water.
Elastase inhibition ratio (%) = (1- (C-D)/(a-B)) ×100
Wherein: a-is the absorbance of the reaction solution without the sample;
b-is the absorbance of the reaction solution without sample and enzyme;
c-is the absorbance of the reaction solution containing the sample and the enzyme;
d-is the absorbance of the reaction solution containing the sample and containing no enzyme.
The statistical analysis software is SPSS, and the comparison among the elastase inhibition rates of the test sample, the positive control and the negative control adopts independent sample t test. The above statistical analysis was a two-tailed test with a significance level of a=0.05. P >0.05, indicating no significant difference between the two groups; p <0.05, indicating a significant difference between the two groups.
TABLE 1 test results of Performance test 1
| Examples | Results of the test,% | P value |
| Example 1 | 51.081±7.157 | <0.05 |
| Example 2 | 32.563±2.581 | <0.05 |
| Example 3 | 37.213±5.143 | <0.05 |
| Example 4 | 40.671±1.346 | <0.05 |
| Example 5 | 43.281±6.351 | <0.05 |
| Example 6 | 30.197±5.397 | <0.05 |
| Negative control | -8.121±5.135 | / |
| Positive control | 78.031±3.235 | <0.05 |
Performance test 2: moisture retention performance test
And carrying out a moisturizing performance experiment on the to-be-detected products 1-6.
30 healthy skin volunteers with ages of 25-35 are selected, no skin care product is used in 24 hours before the experiment, the experiment is carried out after the skin volunteers enter a test environment for resting for 4 hours, 1-6 g of the product to be tested is respectively used after the faces of the volunteers are cleaned, the skin is coated until the skin absorption is completed, the moisture content of the forehead and the cheek of the face mask after 0 hour, 1 hour, 2 hours and 4 hours after the skin moisture tester (Corneometer CM825 in Germany) is used for measuring, 4 times are selected in the same area, and the average value is recorded as the MVV value of the moisture content of the skin. Each test sample was tested by 5 volunteers, and the measurement results at each time point were averaged from 5 parallel experiments to avoid the influence of individual skin level.
General criteria for MVV values:
MVV <35 is noted as dry skin;
MVV value is 35-50, and the skin moisture is medium;
MVV >50 skin moisture was adequate.
The measurement results are shown in the following table 2:
TABLE 2 moisturizing Performance test results
Performance test 3: repair Performance test
30 volunteers in performance test 2 were cleaned with clean water, and then applied with adhesive tape 3 times to break skin barrier after peeling, and 1-6 samples were used 1mg/cm per day 2 The application is carried out until the skin absorption is completed. After 3D, the skin transepidermal water loss measurement (skin moisture loss tester), the skin redness degree a (multifunctional 3D skin imaging analyzer) and the skin redness index EI (multifunctional 3D skin imaging analyzer) were tested respectively.
(1) The skin transepidermal water loss rate was calculated to be:
improvement rate of skin TEWL after application of product t (D)
Wherein: d0— skin parameter base values before product application in the test area
Dt- -skin parameter value after product application in the test area
N- -number of subjects
(2) The value of a is calculated to obtain:
improvement in skin a% after application of product t (D)
Wherein: d0— skin parameter base values before product application in the test area
Dt- -skin parameter value after product application in the test area
N- -number of subjects
(2) And (3) calculating an EI value to obtain:
improvement rate of skin EI value after use of product t (D)%
Wherein: d0— skin parameter base values before product application in the test area
Dt- -skin parameter value after product application in the test area
N- -number of subjects
TABLE 3 test results of Performance test 3
| Examples | Improvement rate of TEWL% | Improvement rate of a% | Improvement rate of EI value% |
| Example 1 | 6.15 | 4.21 | 4.35 |
| Example 2 | 3.61 | 3.01 | 3.09 |
| Example 3 | 4.35 | 3.12 | 3.22 |
| Example 4 | 5.75 | 3.64 | 3.74 |
| Example 5 | 5.99 | 3.81 | 3.95 |
| Example 6 | 2.15 | 1.75 | 1.82 |
Wherein, the experimental effect of part of human body efficacy is shown in figures 1 and 2.
Finally, it should be noted that: the above embodiments are only for illustrating the technical solution of the present invention, and not for limiting the same; although the invention has been described in detail with reference to the foregoing embodiments, it will be understood by those of ordinary skill in the art that: the technical scheme described in the foregoing embodiments can be modified or some or all of the technical features thereof can be replaced by equivalents; such modifications and substitutions do not depart from the spirit of the invention.
Claims (10)
1. An extraction method of rhodiola rosea extract is characterized by comprising the following steps:
s01, pretreatment: washing the cultivated rhodiola rosea young roots with flowing sterile water for 1h, and then slicing the rhodiola rosea young roots on a sterile super clean bench by using a sterile blade; then cleaning the sliced rhodiola rosea young root slices with sterile water for 2min;
s02, inducing callus: inoculating the S01 cleaned rhodiola rosea young root slices into a tissue culture bottle containing hormone for inducing callus;
induction conditions: dark culturing at 25+ -1deg.C for 24 hr, and adding light for 8 hr every day; after 2 weeks, the edges of the materials grow massive callus; cutting off the callus after 3 weeks of culture;
s03, subculturing the callus: transferring the callus with loose upper part, good state and no browning into a secondary culture medium, and performing dark culture at 25+/-1 ℃ for 24 hours, and then illuminating for 8 hours per day with illumination intensity of 1500Lx;
s04, shake flask culture: inoculating the callus subjected to the secondary culture for three times into a liquid culture medium, performing shake culture at 100rpm with the inoculum size of one third, performing dark culture for 24 hours, and performing illumination culture for 8 hours every day at the culture temperature of 25+/-1 ℃;
s05, culturing in a bioreactor: inoculating liquid suspension cells cultured in a shake flask as seed cells, wherein the inoculation volume is 15% of the volume of a culture medium for culture;
the culture conditions are as follows: the initial ventilation is 50L/h, the stirring speed is 100rpm, the culture temperature is 26+/-1 ℃, and the dark culture is carried out for 24 hours after inoculation; then illumination is carried out for 8 hours every day, and the illumination intensity is 1500Lx; and simultaneously changing the stirring speed to 140rpm, and continuously culturing for 2 weeks;
s06, fermenting and culturing: filtering the culture solution in the step S05, and then inoculating the culture solution into a fermentation medium for fermentation.
2. The method according to claim 1, wherein the medium in step S02 comprises the following components:
MS fixed culture medium, 0.5 mg/L6-benzyladenine, 6.5 mg/La-naphthylacetic acid, 40g/L sucrose, 0.2mg/L indoleacetic acid, natural pH value.
3. The method according to claim 1, wherein the medium in step S03 comprises the following components:
MS fixed culture medium, 0.2mg/L vitamin A,6.0 mg/La-naphthylacetic acid, 35g/L glucose, 6mg/L PVP, natural pH value.
4. The method for extracting rhodiola rosea extract according to claim 1, wherein the medium in step S04 comprises the following components:
MS liquid culture medium, 6.0mg/L a-naphthylacetic acid, 50g/L sucrose, 0.2mg/L glutamine, 5mg/L PVP and natural pH value.
5. The method for extracting rhodiola rosea extract according to claim 1, wherein the bacterial species used in step S06 are: pentose milk plant stems and lactobacillus terreus.
6. A rhodiola rosea extract prepared by the extraction method of any one of claims 1-5.
7. The rhodiola rosea extract according to claim 6, for protecting or enhancing skin barrier function.
8. Rhodiola rosea extract according to claim 6, for protecting the skin cells from apoptosis caused by uv irradiation.
9. Rhodiola rosea extract according to claim 6, for whitening the skin.
10. Rhodiola rosea extract according to claim 6, for anti-ageing or anti-oxidation.
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