CN114344206B - Probiotic fermented green plum flower skin care product with whitening effect and preparation method thereof - Google Patents
Probiotic fermented green plum flower skin care product with whitening effect and preparation method thereof Download PDFInfo
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- 230000002087 whitening effect Effects 0.000 title claims abstract description 12
- 238000000855 fermentation Methods 0.000 claims abstract description 80
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- Medicines Containing Plant Substances (AREA)
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Abstract
The invention belongs to the technical field of biological fermentation, and particularly relates to a probiotic fermented green plum flower skin care product with a whitening effect and a preparation method thereof. The green plum flower skin care product fermented by probiotics with the whitening function is obtained through the following steps: (1) superfine grinding green plum flowers; (2) carrying out enzymolysis and sterilization on the crushed green plum flowers; (3) inoculating probiotics, and fermenting twice; (4) Centrifuging and precipitating the fermentation liquor, leaching again, merging the supernatant, and carrying out antiseptic treatment to obtain the green plum probiotic fermentation liquor skin care product. The invention has the beneficial effects that: (1) The results of the B16 melanocyte evaluation experiment on the fermentation broth of the product green plum blossom probiotics show that the product green plum blossom probiotics has a good melanin inhibiting effect; (2) The results of the in vitro antioxidation evaluation experiment of the plum blossom probiotics fermentation liquor show that the plum blossom probiotics fermentation liquor has good antioxidation function.
Description
Technical Field
The invention belongs to the technical field of biological fermentation, and particularly relates to a probiotic fermented green plum flower skin care product with a whitening effect and a preparation method thereof.
Background
Regarding a preparation method of green plum flower extract or a fermentation extraction process of green plum flower, the following patent documents disclose:
CN101536967A discloses application of green plum flower extract in preparing anti-aging cosmetic, and also discloses preparation method of green plum flower extract, 1) and CO 2 Supercritical extraction of nonpolar components of green plum: putting dried green plum flowers with the water content less than or equal to 1.5% into an extraction kettle, and introducing CO of 32-38 Mpa 2 Circularly and dynamically extracting for 1.5-2.5 h at the temperature of 55-65 ℃; then carrying out reduced pressure separation at the separation temperature of 38-42 ℃ and the separation pressure of 3-5 Mpa to obtain nonpolar components in the green plum blossom; 2) Extraction of polar components: extracting the residue obtained in step 1) with 30% ethanolThe weight ratio of the slag to the ethanol is 1:15, and the hot reflux extraction is carried out for 2 hours at the temperature of 80 ℃; filtering to obtain filtrate, and distilling at 78deg.C to obtain polar component in green plum; 3) Mixing the nonpolar component obtained in step 1) and step 2) with the polar component.
The method adopts CO 2 The supercritical extraction technical scheme has the advantages of expensive equipment, higher cost, high cost and complex operation, and is not suitable for large-scale production and application.
CN109806204a is a fermentation extraction process of green plum blossom, which is characterized by comprising the following steps: (1) Taking dried green plum flower, performing wall breaking treatment, sterilizing, and adding distilled water; wherein the weight ratio of the plum blossom after wall breaking to distilled water is 1 (20-30); (2) Continuously adding lactobacillus, leuconostoc and saccharomycetes into the step (1), and carrying out composite anaerobic fermentation for 12-72 hours at the temperature of 20-40 ℃ to obtain a fermentation product; wherein, the weight ratio of the green plum blossom to the lactobacillus, the leuconostoc mesenteroides and the saccharomycetes after wall breaking is 1: (10-15): (10-15): (10-15); (3) Hydrolyzing the fermentation product in the step (2) by adopting protease, and carrying out enzymolysis for 1-6 h at the temperature of 30-50 ℃; wherein the weight ratio of protease to fermentation product is (1-5): 100; (4) Evaporating the fermented product after enzymolysis under reduced pressure to 3-5 times of the weight of the green plum flower after wall breaking, continuously adding preservative, and uniformly stirring to obtain green plum flower extract; wherein the weight ratio of the preservative to the broken plum flower is (2-5) 1000.
The anaerobic fermentation of the composite microorganism containing the saccharomycetes adopted in the patent document can produce a large amount of ethanol in the fermentation process, and the subsequent product application can cause damage to skin care at first.
Therefore, the shortcomings of the method are overcome, and the invention provides the green plum flower skin care product fermented by probiotics with whitening function, which has relatively low cost and excellent efficacy and is suitable for large-scale production and application, and the preparation method thereof.
Disclosure of Invention
In order to solve the technical problems, the invention provides a probiotic fermented green plum flower skin care product with a whitening function and a preparation method thereof.
The green plum flower skin care product with the whitening function and the probiotic fermentation is prepared by taking green plum flowers as raw materials and fermenting the raw materials after enzymolysis.
Specifically, the enzyme adopted in the enzymolysis is at least two of acid cellulase, acid pectase, acid proteinase, amylase and lipase, and the dosage of the enzyme is 0.5-2% of the weight of the green plum flower.
The probiotics used in fermentation are at least one of lactobacillus rhamnosus, lactobacillus paracasei, streptococcus thermophilus, lactobacillus plantarum, saccharomycetes, bifidobacterium longum, lactobacillus plantarum, lactobacillus lactococcus and lactobacillus reuteri.
The inoculation amount of the probiotics is 2-5%, the fermentation temperature is 35-39 ℃ and the fermentation time is 5-7 days.
The preparation method of the green plum flower skin care product fermented by probiotics with the whitening function comprises the following steps of:
(1) Superfine pulverizing green plum flower;
(2) Enzymolysis and sterilization are carried out on the crushed green plum flowers;
(3) Inoculating probiotics, and fermenting twice;
(4) Centrifuging the fermentation liquor, and collecting supernatant; leaching again, centrifuging, collecting supernatant of leaching solution, mixing the supernatant, and antiseptic treating to obtain skin care product.
In the step (1), the green plum flowers are subjected to superfine grinding for 20-30 min until the fineness reaches more than 1500 meshes.
(2) The enzymolysis conditions are as follows: the enzymolysis temperature is 50-60 ℃, the pH value is 4.8-6.5, and the enzymolysis stirring is carried out for 2-3 hours; the enzyme is at least two of acid cellulase, acid pectase, acid proteinase, amylase and lipase, and the dosage of the enzyme is 0.5-2% of the weight of green plum flower;
pasteurization is adopted during sterilization, the sterilization temperature is 80-90 ℃, and the sterilization time is 25-35 min.
(3) Inoculating at least one of lactobacillus rhamnosus, lactobacillus paracasei, streptococcus thermophilus, lactobacillus plantarum, saccharomycetes, bifidobacterium longum, lactobacillus plantarum, lactobacillus lactococcus and lactobacillus reuteri, wherein the inoculation amount is 2-5%, the fermentation temperature is 35-39 ℃ and the fermentation time is 5-7 days;
inoculating at least one of lactobacillus rhamnosus, lactobacillus paracasei, streptococcus thermophilus, lactobacillus plantarum, saccharomycetes, bifidobacterium longum, lactobacillus plantarum, lactobacillus lactococcus and lactobacillus reuteri for 5-7 days after fermentation is finished, wherein the fermentation temperature is 35-39 ℃.
(4) Wherein the centrifugal speed of the fermentation liquor is 8000r/min-11000r/min, and the centrifugal time is 10min-15min;
leaching again by adopting a reflux mode at the temperature of 80-100 ℃ for 30-60 min;
during the antiseptic treatment, at least one of sodium benzoate, potassium sorbate, salicylic acid, methyl parahydroxybenzoate, ethyl parahydroxybenzoate, propyl parahydroxybenzoate and butyl parahydroxybenzoate is added as antiseptic, and the addition amount of the antiseptic is 0.05-0.2% of the weight of the combined supernatant.
Preferably, the preparation method of the green plum flower skin care product fermented by probiotics with whitening function comprises the following steps:
(1) Superfine pulverizing green plum flower for 20-30 min to fineness of above 1500 mesh;
(2) Adding distilled water into crushed green plum flowers according to the weight ratio of 1:25-35, carrying out enzymolysis, pasteurizing, sterilizing at 80-90 ℃ for 25-35 min; the enzymolysis conditions are as follows: the enzymolysis temperature is 50-60 ℃, the pH value is 4.8-6.5, and the enzymolysis stirring is carried out for 2-3 hours; the enzyme is at least two of acid cellulase, acid pectase, acid proteinase, amylase and lipase, and the dosage of the enzyme is 0.5-2% of the weight of green plum flower;
(3) Inoculating at least one of lactobacillus rhamnosus, lactobacillus paracasei, streptococcus thermophilus, lactobacillus plantarum, saccharomycetes, bifidobacterium longum, lactobacillus plantarum, lactobacillus lactococcus and lactobacillus reuteri, wherein the inoculation amount is 2-5%, the fermentation temperature is 35-39 ℃ and the fermentation time is 5-7 days;
inoculating at least one of lactobacillus rhamnosus, lactobacillus paracasei, streptococcus thermophilus, lactobacillus plantarum, saccharomycetes, bifidobacterium longum, lactobacillus plantarum, lactobacillus lactococcus and lactobacillus reuteri for 5-7 days after fermentation is finished, wherein the fermentation temperature is 35-39 ℃;
(4) Centrifuging the fermentation liquor, precipitating, collecting supernatant for later use, adding distilled water into the precipitate to supplement the original total weight, leaching again, combining the supernatant, and carrying out antiseptic treatment to obtain the skin care product of the plum blossom probiotic fermentation liquor;
the centrifugal speed of the fermentation liquor is 8000-11000r/min, and the centrifugal time is 10-15min;
the re-leaching adopts a reflux mode, the temperature is 80-100 ℃, and the leaching time is 30-60min.
The invention has the beneficial effects that:
(1) The results of evaluation experiments on B16 melanocytes (MTT, tyrosinase, melanin) show that the product of the invention, namely the fermentation broth of the green plum probiotics, has a good melanin inhibition effect;
(2) The experimental result of the in vitro antioxidation (DPPH, superoxide anion, hydroxyl radical) evaluation of the plum blossom probiotics fermentation liquor shows that the plum blossom probiotics fermentation liquor has good antioxidation function.
Drawings
FIG. 1 is a graph showing the inhibition of B16 melanocytes by green plum flower probiotic fermentation broth;
FIG. 2 is a graph showing the evaluation results of the in vitro antioxidation of the green plum flower probiotic fermentation liquid.
Detailed Description
For further explanation of the present invention, the present inventors will further explain by way of the following specific examples, but the present invention is not limited thereto.
Example 1
The preparation method of the green plum flower skin care product fermented by probiotics with the whitening function comprises the following steps of:
(1) Weighing 50g of green plum flower superfine powder, adding 1500ml of distilled water, fully and uniformly stirring, and adjusting the pH to 5.0;
(2) Adding 0.5g of acid cellulase and 0.5g of acid pectase while stirring, and stirring and carrying out enzymolysis for 2 hours at 55 ℃;
(3) Transferring the enzymolysis liquid into a sterilizing device, pasteurizing at 85 ℃ for 30 minutes, transferring into a sterile operation table after sterilization, and inoculating 3% lactobacillus rhamnosus when cooling to 40 ℃;
(4) Transferring into a 37 ℃ incubator, standing for culturing, taking and measuring pH and reducing sugar every day, wherein the reducing sugar does not drop any more until fermentation is finished;
(5) After fermentation, inoculating 3% of lactobacillus paracasei;
(6) Transferring into a 37 ℃ incubator, standing for culturing, taking and measuring pH and reducing sugar every day, wherein the reducing sugar does not drop any more until fermentation is finished;
(7) Centrifuging the fermentation broth at 8000r/min-11000r/min for 10min-15min; collecting supernatant for later use;
(8) Adding water to the precipitate to 1550g, transferring into a reflux bottle, and reflux-extracting at 100 ℃ for 1 hour;
(9) Centrifuging the leaching solution at 10000r/min for 10min, collecting supernatant, and mixing with the supernatant after fermentation in step (7);
(10) And (3) adding 0.1% of methyl parahydroxybenzoate into the combined fermentation liquor for preservative treatment to obtain the green plum flower probiotic fermentation liquor.
Example 2
(1) Weighing 50g of green plum flower superfine powder, adding 1500ml of distilled water, fully and uniformly stirring, and adjusting the pH to 5.5;
(2) Adding 0.5g of acid cellulase and 0.5g of acid pectase while stirring, and stirring and carrying out enzymolysis for 2.5 hours at 55 ℃;
(3) Transferring the enzymolysis liquid into a sterilizing device, pasteurizing at 85 ℃ for 30 minutes, transferring into a sterile operation table after sterilization, and inoculating 5% lactobacillus rhamnosus when cooling to 40 ℃;
(4) Transferring into a 37 ℃ incubator, standing for culturing, taking and measuring pH and reducing sugar every day, wherein the reducing sugar does not drop any more until fermentation is finished;
(5) After fermentation, continuing inoculating 5% of lactobacillus paracasei;
(6) Transferring into a 37 ℃ incubator, standing for culturing, taking and measuring pH and reducing sugar every day, wherein the reducing sugar does not drop any more until fermentation is finished;
(7) Centrifuging the fermentation liquor at 8000r/min-11000r/min for 12min;
(8) Collecting supernatant for later use, adding water to the precipitate until 1550g, transferring into a reflux bottle, and reflux-extracting at 100 ℃ for 1 hour;
(9) Centrifuging the leaching solution at 9000r/min for 12min, collecting supernatant, and mixing with the fermented supernatant;
(10) And (3) adding 0.1% of methyl parahydroxybenzoate into the combined fermentation liquor for preservative treatment to obtain the green plum flower probiotic fermentation liquor.
Example 3
(1) Weighing 50g of green plum flower superfine powder, adding 1500ml of distilled water, fully and uniformly stirring, and adjusting the pH to 6;
(2) Adding 0.5g of acid cellulase and 0.5g of acid pectase while stirring, and stirring and carrying out enzymolysis for 2 hours at 55 ℃;
(3) Transferring the enzymolysis liquid into a sterilizing device, pasteurizing at 85 ℃ for 30 minutes, transferring into a sterile operation table after sterilization, and inoculating 7.5% lactobacillus rhamnosus when cooling to 40 ℃;
(4) Transferring into a 37 ℃ incubator, standing for culturing, taking and measuring pH and reducing sugar every day, wherein the reducing sugar does not drop any more until fermentation is finished;
(5) After fermentation, inoculating 7.5% of lactobacillus paracasei;
(6) Transferring into a 37 ℃ incubator, standing for culturing, taking and measuring pH and reducing sugar every day, wherein the reducing sugar does not drop any more until fermentation is finished;
(7) Centrifuging the fermentation liquor at the rotation speed of 10000r/min for 15min;
(8) Collecting supernatant for later use, adding water to the precipitate until 1550g, transferring into a reflux bottle, and reflux-extracting at 100 ℃ for 1 hour;
(9) Centrifuging the leaching solution at 9000r/min for 12min, collecting supernatant, and mixing with the fermented supernatant;
(10) And (3) adding 0.1% of methyl parahydroxybenzoate into the combined fermentation liquor for preservative treatment to obtain the green plum flower probiotic fermentation liquor.
Example 4
(1) Weighing 50g of green plum flower superfine powder, adding 1500ml of distilled water, fully and uniformly stirring, and adjusting the pH to 6.5;
(2) Adding 0.5g of acid cellulase and 0.5g of acid pectase while stirring, and stirring and carrying out enzymolysis for 2 hours at 55 ℃;
(3) Transferring the enzymolysis liquid into a sterilizing device, pasteurizing at 85 ℃ for 30 minutes, transferring into a sterile operation table after sterilization, and inoculating 10% lactobacillus rhamnosus when cooling to 40 ℃;
(4) Transferring into a 37 ℃ incubator, standing for culturing, taking and measuring pH and reducing sugar every day, wherein the reducing sugar does not drop any more until fermentation is finished;
(5) After fermentation, continuing inoculating 10% of lactobacillus paracasei;
(6) Transferring into a 37 ℃ incubator, standing for culturing, taking and measuring pH and reducing sugar every day, wherein the reducing sugar does not drop any more until fermentation is finished;
(7) Centrifuging the fermentation liquor at the rotation speed of 10000r/min for 12min;
(8) Collecting supernatant for later use, adding water to the precipitate until 1550g, transferring into a reflux bottle, and reflux-extracting at 100 ℃ for 1 hour;
(9) Centrifuging the leaching solution at 8000r/min-11000r/min for 10min-15min, collecting supernatant, and mixing with the fermented supernatant;
(10) Adding 0.1% of methyl parahydroxybenzoate into the combined fermentation liquor for preservative treatment to obtain green plum flower probiotic fermentation liquor;
TABLE 1 results of fermentation broth sampling test in examples 1 to 4
In table 1, the conditions for the boiling water extraction are as follows: (1) Weighing 50g of green plum flower superfine powder, adding 1500ml of distilled water, stirring thoroughly, and boiling for 30min at pH of 6.0. (2) Centrifuging the boiled water extract at 10000r/min for 12min, and collecting supernatant. (3) the precipitate was made up to 1550g and was boiled for 30min. (4) Centrifuging the twice boiled water extract at 10000r/min for 12min, and mixing the two supernatants.
As can be seen from the above table, the fermentation broths obtained by extraction in examples 1 to 4 have a total sugar content far lower than that of the boiling water extraction method, a flavone content far higher than that of the boiling water extraction method, and a saponin content almost 3 times that of the boiling water extraction method. This demonstrates that the fermentation broth obtained by the process of the present invention has a far higher content of flavonoids and saponins as active ingredients than the boiling water extraction process.
Example 5
5.1, examining the effect of the green plum flower probiotics fermentation liquor prepared by the invention on inhibiting B16 cell melanin, adopting a NaOH dissolution method, wherein the specific method is as follows:
taking B16 melanoma cells in logarithmic phase, discarding culture solution, washing with PBS solution for 2 times, adding culture solution, and adjusting cell concentration to 0.5X10 4 -1×10 4 The wells correspond to 5X 10 stem cell suspensions 4 -10×10 4 Each (ml-1) was added to a 96-well plate, and 100ul was added per well. After overnight (12 h), the cells were attached to the wall, and the same concentration and volume of boiling water-containing extract as in MTT method and medium of fermentation broth of Thermopsis prinus of examples 1-4 were added, and a control group (cell+medium) was set, each concentration group was provided with 5 multiple wells, each medium contained 5% fetal bovine serum, and 5% CO at 37deg.C 2 After 56h of culture in a constant temperature incubator, the culture solution was aspirated and digested with 0.25% pancreatin, the supernatant was discarded by centrifugation at 1000r/min for 5min, 200ul of NaOH (1 mol/L) solution containing 10% dimethyl sulfoxide was added to the cells under each treatment factor, melanin particles were completely dissolved in a water bath at 65℃and the relative melanin content was detected by ELISA.
5.2, examining the in vitro antioxidation effect of the green plum flower probiotic fermentation liquid prepared by the invention, wherein the specific method is as follows:
(1) DPPH method: sample tubes, sample background, DPPH tubes and solvent background were set up using 10mL test tubes. 1mL boiling water leaching liquor with the same concentration and the green plum blossom probiotics fermentation liquor in the examples 1-4 are respectively added into a sample tube and a sample background. All tubes were filled with water, 3mL was made up, and mixed well. 1mL of DPPH ethanol solution is added into the sample tube and the DPPH tube, the sample background and the solvent background are replaced by 95% ethanol, the mixture is gently shaken, and the mixture is kept stand for 5 minutes at room temperature. DPPH free radical scavenging was detected and calculated using an ultraviolet spectrophotometer.
(2) Hydroxyl radical method: respectively sucking 1.0mL boiling water extract with the same concentration and the fermentation liquor of the green plum probiotics in examples 1-4 into a 10mL colorimetric tube, adding 1.0mL FeSO 9mmol/L 4 After the solution and 1.0mL of 9mmol/L salicylic acid-ethanol solution were thoroughly mixed, 1.0mL of 8.8mmol/L H was added 2 O 2 The reaction was carried out at 37℃for 30min with distilled water as a blank. And detecting and calculating the free radical hydroxyl radical scavenging rate by using an ultraviolet spectrophotometer.
(3) The super anion and cation method comprises the following steps: taking 4.5mL of 50mmol/L Tris-HCl buffer solution (pH 8.2), uniformly mixing, preserving heat for 20min in a water bath at 25 ℃, taking out, respectively adding 1.0mL of boiling water extract, the green plum probiotic fermentation liquor in examples 1-4 and 0.4mL of 25mmol/L pyrogallic acid solution (pyrogallol), shaking uniformly, reacting for 5min in the water bath at 25 ℃, and adding 1.0mL of 8mmol/L HCl solution to terminate the reaction. And detecting and calculating the super anion and cation clearance rate by using an ultraviolet spectrophotometer.
As shown in fig. 2: the superoxide anion detection result shows that the removal rate of the superoxide anion is only 55% by adopting a boiling water leaching method.
In the methods of examples 1 to 4, the superoxide anion scavenger was 70% or more. DPPH results show that the free radical removal rate by boiling water leaching is only 40%, while the removal rates in examples 1 to 4 reach up to 60% more free radical removal rate; in the examination of the hydroxyl radical detection results, the clearance rate of the hydroxyl radical extracted by boiling water is only 87%, while the clearance rate of the hydroxyl radical in examples 1-4 reaches 93% at most.
Claims (2)
1. The skin care product is characterized by being prepared by a preparation method comprising the following steps of:
(1) Superfine pulverizing green plum flower for 20-30 min to fineness of above 1500 mesh;
(2) Adding distilled water into the crushed green plum flowers according to the weight ratio of 1:25-35, carrying out enzymolysis, pasteurizing, and sterilizing at the temperature of 80-90 ℃ for 25-35 min; the enzyme is acid cellulase and acid pectase, the dosage of the enzyme is 0.5-2% of the weight of green plum flower, and the enzymolysis conditions are as follows: the enzymolysis temperature is 50-60 ℃, the pH value is 4.8-6.5, and the enzymolysis stirring is carried out for 2-3 hours;
(3) Inoculating lactobacillus rhamnosus into the sterilized enzymolysis liquid for fermentation, wherein the inoculation amount is 2-5%, the fermentation temperature is 35-39 ℃, and the fermentation time is 5-7 days; inoculating 2-5% of lactobacillus paracasei for fermentation after fermentation is finished, wherein the fermentation temperature is 35-39 ℃ and the fermentation time is 5-7 days;
(4) Centrifuging the fermentation liquor, precipitating, and collecting supernatant for later use; adding distilled water into the precipitate to supplement the original total weight, leaching again by adopting a reflux method, centrifuging the leaching solution at 8000r/min-11000r/min for 10min-15min, and collecting the supernatant of the leaching solution; combining the two supernatants, and performing antiseptic treatment to obtain the green plum flower skin care product fermented by probiotics; the centrifugal speed of the fermentation liquor is 8000-11000r/min, and the centrifugal time is 10-15min; the secondary leaching adopts a reflux mode, the temperature is 80-100 ℃, and the leaching time is 30-60min.
2. The probiotic fermented green plum flower skin care product with whitening function according to claim 1, wherein in (4), leaching is performed again at 100 ℃ for leaching 1 h; during the antiseptic treatment, at least one of sodium benzoate, potassium sorbate, salicylic acid, methyl parahydroxybenzoate, ethyl parahydroxybenzoate, propyl parahydroxybenzoate and butyl parahydroxybenzoate is added as an antiseptic, and the addition amount of the antiseptic is 0.05-0.2% of the weight of the combined supernatant.
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| CN104739710A (en) * | 2015-02-25 | 2015-07-01 | 烟台新时代健康产业日化有限公司 | Skin regeneration compound and use thereof |
| CN108553391A (en) * | 2018-07-01 | 2018-09-21 | 佛山文森特知识产权服务有限公司 | A kind of preparation method of the plant fermentation composition with anti-wrinkle |
| CN109806204A (en) * | 2019-04-04 | 2019-05-28 | 烟台新时代健康产业日化有限公司 | A kind of fermenting extraction process of green plum-blossom |
| CN111840165A (en) * | 2020-08-14 | 2020-10-30 | 山东花物堂生物科技有限公司 | Preparation method of plant whitening fermentation liquor |
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| CN104739710A (en) * | 2015-02-25 | 2015-07-01 | 烟台新时代健康产业日化有限公司 | Skin regeneration compound and use thereof |
| CN108553391A (en) * | 2018-07-01 | 2018-09-21 | 佛山文森特知识产权服务有限公司 | A kind of preparation method of the plant fermentation composition with anti-wrinkle |
| CN109806204A (en) * | 2019-04-04 | 2019-05-28 | 烟台新时代健康产业日化有限公司 | A kind of fermenting extraction process of green plum-blossom |
| CN111840165A (en) * | 2020-08-14 | 2020-10-30 | 山东花物堂生物科技有限公司 | Preparation method of plant whitening fermentation liquor |
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