CN114540426B - Peony fermentation filtrate and fermentation process thereof - Google Patents
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Abstract
The invention provides a peony fermentation filtrate and a fermentation process thereof. The fermentation process comprises the following steps: (1) pulverizing and sieving radix moutan to obtain radix moutan fine powder; (2) mixing fine powder of peony root, carbon source, vitamins, inorganic salt and water uniformly, sterilizing under high pressure to obtain a sterilized culture medium, and cooling for later use; (3) carrying out enlarged culture on the lactobacillus to obtain a secondary seed solution; (4) transferring the secondary seed liquid to a sterilization culture medium, and performing constant-temperature culture and fermentation at 35 +/-1 ℃ to obtain a fermentation liquid; (5) and (4) after the fermentation liquor is treated at high temperature, adding a preservative, and uniformly mixing to obtain a peony fermentation filtrate. The peony fermentation filtrate obtained by the invention shows good anti-inflammatory effect and scar-resistant effect, can inhibit melanin synthesis, has antioxidant/cell protection effect, and has simple preparation raw materials and process, safety, environmental protection and controllability.
Description
Technical Field
The invention relates to the field of plant fermentation liquor, and particularly relates to peony fermentation filtrate and a fermentation process thereof.
Background
In China, the peony cultivation history is long and is a beautiful name of the national flower. With the development of economy and the progress of science and technology, the peony is not only taken as an ornamental plant, but also has wide development on medicinal value and economic value, and hundreds of products such as peony seed oil, peony tea, peony cosmetics, peony daily chemicals, peony functional food, peony biological medicine products and the like are also developed, so that the peony goes on a way of deep, multi-field and comprehensive utilization.
Paeonol is one of the effective components in peony root, the content can reach about 1.5-3.0%, and the Paeonol has the effects of resisting aging, resisting oxidation, scavenging free radicals, resisting bacteria, diminishing inflammation, preventing cardiovascular system diseases, resisting tumors and the like, has the advantages of short half life, few adverse reactions and the like, and is gradually applied to the field of cosmetics. However, paeonol is easily dissolved in alcohol organic solvent and has poor water solubility, so the existing paeonol extraction process mainly adopts CO 2 Supercritical extraction method, organic solvent extraction method, steam distillation method, direct distillation method, macroporous resin method, etc., but these processes are not environment-friendly enough in the process of extracting paeonol and the extraction rate is not high. Chinese patent CN106699535A discloses a process for producing low-temperature subcritical extraction of paeonol, which uses dimethyl ether as an extractant to carry out low-temperature subcritical extraction of paeonol from peony root residue powder. The dimethyl ether used in the method is not environment-friendly enough, and although the extracted paeonol has high content, the paeonol has difficult water solubility and obvious smell, and finally the use is limited.
Based on the method, the invention provides the peony root fermented filtrate and the fermentation process thereof, and the fermentation of the lactobacillus to the peony root is adopted to convert paeonol into glycosylated paeonol derivatives, so that the water solubility, the biological activity and the cell tolerance of the glycosylated paeonol derivatives are improved, and the peony root fermented filtrate which is safer to use, more stable to store and better in effect is obtained.
Disclosure of Invention
The invention provides a peony fermentation filtrate, which is prepared from raw materials including peony root fine powder, lactic acid bacteria, a carbon source, vitamins, inorganic salt, a preservative and water; the inoculation amount of the lactobacillus is 0.5-5 times of the weight of the fine powder of the peony root.
Preferably, the inoculation amount of the lactic acid bacteria is 1.4 times of the weight of the fine powder of the peony root.
In a preferred embodiment, the lactic acid bacteria are selected from at least one of lactobacillus, bifidobacterium, streptococcus thermophilus.
In a preferred embodiment, the lactobacillus is selected from at least one of lactobacillus acidophilus, lactobacillus casei, lactobacillus delbrueckii, lactobacillus fermentum, lactobacillus plantarum, lactobacillus rhamnosus. More preferably, the Lactobacillus is Lactobacillus plantarum.
In a preferred embodiment, the lactic acid bacterium is Lactobacillus plantarum (Lactobacillus plantarum).
In a preferred embodiment, the mesh size of the fine powder of peony root is 40 to 200 mesh.
In a preferred embodiment, the mesh number of the fine powder of peony root is 60 to 80 mesh, and more preferably, the mesh number of the fine powder of peony root is 60 mesh.
In a preferred embodiment, the weight ratio of the fine powder of peony root, the carbon source, the vitamins, the inorganic salts and the water is (2-8): (2-6): (0.2-1): (0.5-1): (80-90).
In a preferred embodiment, the vitamin is selected from at least one of the vitamin B group, vitamin H, vitamin E.
In a preferred embodiment, the vitamin is a vitamin B group.
In a preferred embodiment, the carbon source is selected from at least one of a monosaccharide source, a disaccharide source, a polysaccharide source, glycerol, a small molecule alcohol, an inorganic carbon source.
In a preferred embodiment, the monosaccharide source is selected from at least one of glucose, fructose, galactose, mannose, arabinose,
in a preferred embodiment, the disaccharide source is selected from at least one of maltose, sucrose, lactose, trehalose.
In a preferred embodiment, the carbon source is glucose.
In a preferred embodiment, the inorganic salt is sodium chloride.
In a preferred embodiment, the weight ratio of the fine powder of peony root, the carbon source, the vitamin, the inorganic salt and the water is 6:4:0.4:0.6: 89.
The second aspect of the invention provides a fermentation process of peony fermentation filtrate, which comprises the following steps:
(1) pulverizing and sieving radix moutan to obtain radix moutan fine powder;
(2) mixing fine powder of peony root, carbon source, vitamins, inorganic salt and water uniformly, sterilizing under high pressure to obtain a sterilized culture medium, and cooling for later use;
(3) carrying out enlarged culture on the lactobacillus to obtain a secondary seed solution;
(4) transferring the secondary seed liquid to a sterilization culture medium, and performing constant-temperature culture and fermentation at 35 +/-1 ℃ to obtain a fermentation liquid;
(5) and (4) after the fermentation liquor is treated at high temperature, adding a preservative, and uniformly mixing to obtain a peony fermentation filtrate.
In a preferred embodiment, the specific process of step (3) is as follows: transferring lactobacillus into MRS liquid culture medium, performing static culture at 35 + -1 deg.C to obtain first-stage seed solution, performing amplification culture in MRS liquid culture medium, and performing static culture at 35 + -1 deg.C for 12-18h to obtain second-stage seed solution.
In a preferred embodiment, the specific process of step (4) is as follows: transferring the secondary seed liquid into a sterilization culture medium, and culturing at constant temperature of 35 + -1 deg.C for 24-96h with stirring speed of constant temperature oscillator of 60-100r/min to obtain fermentation liquid.
More preferably, the incubation time is 72h, and the stirring speed of the constant temperature oscillator is 85 r/min.
In a preferred embodiment, the specific process of step (5) is as follows: carrying out first high-temperature treatment on the fermentation liquor, cooling, then carrying out ultrasonic treatment to obtain treated fermentation liquor, then filtering and removing residues to obtain clear liquid, carrying out second high-temperature treatment on the clear liquid, cooling, and then adding p-hydroxyacetophenone and 1, 2-hexanediol to obtain peony fermentation filtrate.
Wherein the weight ratio of the p-hydroxyacetophenone to the 1, 2-hexanediol is 1: 5.
In a preferred embodiment, the treatment temperature of the first high-temperature treatment is 70 to 80 ℃ and the treatment temperature of the second high-temperature treatment is 80 to 90 ℃.
More preferably, the treatment temperature of the first high-temperature treatment is 75 ℃, and the treatment temperature of the second high-temperature treatment is 85 ℃.
In a preferred embodiment, the specific process of step (5) is as follows: carrying out primary high-temperature treatment on the fermentation liquor at 75 ℃ for 15min, cooling to room temperature, carrying out ultrasonic treatment with the ultrasonic power of 200-500W and the temperature of 40-50 ℃ for 45-60min to obtain treated fermentation liquor, filtering to remove residues to obtain clarified liquid, carrying out secondary high-temperature treatment on the clarified liquid at 85 ℃ for 25min, cooling to room temperature, and adding 0.2wt% of p-hydroxyacetophenone and 1.0wt% of 1, 2-hexanediol to obtain peony fermentation filtrate.
In the application, the fermentation liquor is subjected to two-step high-temperature treatment, so that the fermentation time of the lactobacillus plantarum can be accurately controlled, the types and the contents of active ingredients such as paeonol derivatives and the like in the final fermentation filtrate product are ensured, and the effects of oxidation resistance, inflammation resistance and the like of the peony fermentation liquor are ensured. The applicant believes that the reason is that in the two-step high-temperature treatment process, the first step of high-temperature treatment at 70-80 ℃ is combined with ultrasonic treatment, the cell structure of the lactobacillus plantarum is destroyed, the lactobacillus plantarum in the fermentation liquor is inactivated, the fermentation effect of the lactobacillus plantarum is stopped, the fermentation time of the lactobacillus plantarum is accurately controlled, the types and the content of active ingredients are ensured, the second step of high-temperature treatment at 80-90 ℃ is performed on the filtered fermentation liquor, the fermentation liquor is further purified, bacteria possibly bred and brought in the treatment process are killed, and the sterile environment of the fermentation liquor is ensured. In the experimental process, the applicant finds that if two high-temperature treatment processes are combined into one treatment, the condition that the activity of the lactobacillus plantarum can not be controlled in time is easy to occur, and the using effect of the fermentation liquor is finally influenced.
Compared with the prior art, the invention has the following beneficial effects:
1. the peony root fermented filtrate provided by the invention is obtained by fermenting peony roots through lactobacillus plantarum, and the preparation raw materials are simple, safe and environment-friendly, and the preparation process is safe and controllable.
2. According to the peony root fermentation process provided by the invention, the fermentation time of lactobacillus plantarum is accurately controlled through two-step high-temperature treatment on the fermentation liquor, the types and contents of active ingredients such as paeonol derivatives and the like in the final fermentation filtrate product are ensured, and the antioxidant and anti-inflammatory effects and the like of the peony fermentation liquor are ensured.
3. According to the peony root fermentation filtrate and the fermentation process, the glycosylated paeonol derivative is obtained by performing enzymatic reaction on a cell level through the fermentation process of lactobacillus plantarum, the water dissolution difficulty of the paeonol component is improved, and a fermentation broth product containing paeonol active groups with better water solubility is obtained.
4. According to the peony root fermentation filtrate and the fermentation process, the fermentation filtrate contains paeonol active substances, and also contains various active ingredients such as saccharides and organic acid, so that the peony root fermentation filtrate has good anti-inflammatory and scar-resisting effects, can inhibit melanin synthesis, and has the effects of resisting oxidation and protecting cells.
5. The peony root fermentation filtrate prepared by the method is convenient to store, simple to use, good in solubility in hot water and wide in application, and is particularly suitable to be used as an additive component of cosmetics and skin care products.
Drawings
FIG. 1 is a graph showing the effect of peony fermentation filtrate on SOD and ROS concentration in RAW264.7 macrophages in example 1;
FIG. 2 is a diagram of a peony fermentation filtrate test for human skin prick and discomfort;
FIG. 3 is a graph showing the effect of peony fermentation filtrate on the soothing sensitivity of human skin on the red areas of the face tested in example 1;
FIG. 4 is a graph showing the effect of the peony fermentation filtrate on the sensitivity of the peony fermentation filtrate to skin soothing of the human body in the test of facial redness.
Detailed Description
Example 1
In the first aspect of this embodiment, a peony root fermented filtrate is provided, and the raw materials for preparation include, by weight, 6 parts of peony root fine powder, 8.4 parts of Lactobacillus plantarum (Lactobacillus plantarum), 4 parts of glucose, 0.4 part of vitamin B group, 0.6 part of sodium chloride, 1.2 parts of preservative, and 89 parts of water. Wherein the lactobacillus plantarum is a lactobacillus plantarum secondary seed solution.
In a second aspect, this embodiment provides a fermentation process of peony root fermentation filtrate, which comprises the following steps:
(1) pulverizing and sieving radix moutan to obtain 60 mesh fine powder;
(2) mixing fine powder of peony root, glucose, vitamin B group, sodium chloride, and water, sterilizing at 121 deg.C under 0.105MPa for 30min to obtain sterilized culture medium, and cooling;
(3) transferring lactobacillus plantarum into 100mL of MRS liquid culture medium, performing static culture at 35 +/-1 ℃ to obtain first-stage seed liquid, performing amplification culture on the first-stage seed liquid in the MRS liquid culture medium, and performing static culture at 35 +/-1 ℃ for 18 hours to obtain second-stage seed liquid;
(4) transferring the secondary seed liquid into a sterilization culture medium, and culturing at constant temperature of 35 +/-1 ℃ for 72h, wherein the stirring speed of a constant temperature oscillator is 80r/min, so as to obtain a fermentation liquid;
(5) carrying out primary high-temperature treatment on the fermentation liquor at the temperature of 75 ℃ for 15min, carrying out ultrasonic treatment at the ultrasonic power of 400W and the temperature of 45 ℃ for 50min after cooling to room temperature to obtain treated fermentation liquor, then carrying out filtration and deslagging to obtain clarified liquid, carrying out secondary high-temperature treatment on the clarified liquid at the temperature of 85 ℃ for 25min, cooling to room temperature, and adding 0.2wt% of p-hydroxyacetophenone and 1.0wt% of 1, 2-hexanediol to obtain a peony fermentation filtrate.
The resulting peony fermentation filtrate was subjected to routine data testing as shown in table 1 below:
TABLE 1
Example 2
The embodiment provides a peony fermentation filtrate and a fermentation process thereof, and the specific implementation manner is the same as that in embodiment 1, the difference is that the granularity of the peony root fine powder is 80 meshes, and the weight ratio of the peony root fine powder, glucose, vitamin B group, sodium chloride and water is 5: 3: 0.3: 0.5: 85.
example 3
The embodiment provides a peony fermentation filtrate and a fermentation process thereof, and the specific implementation manner is the same as that of embodiment 1, except that the fermentation process is as follows:
(1) pulverizing and sieving radix moutan to obtain 60 mesh fine powder;
(2) mixing fine powder of peony root, glucose, vitamin B group, sodium chloride, and water, sterilizing at 121 deg.C under 0.105MPa for 30min to obtain sterilized culture medium, and cooling;
(3) transferring lactobacillus plantarum into 100mL of MRS liquid culture medium, performing static culture at 35 +/-1 ℃ to obtain first-stage seed liquid, performing amplification culture on the first-stage seed liquid in the MRS liquid culture medium, and performing static culture at 35 +/-1 ℃ for 18h to obtain second-stage seed liquid;
(4) transferring the secondary seed liquid into a sterilization culture medium, and culturing at constant temperature of 35 +/-1 ℃ for 72h, wherein the stirring speed of a constant temperature oscillator is 80r/min, so as to obtain a fermentation liquid;
(5) filtering the fermentation liquor, keeping the temperature at 85 ℃ for 30min, cooling to room temperature, and adding 0.2wt% of p-hydroxyacetophenone and 1.0wt% of 1, 2-hexanediol to obtain a peony fermentation filtrate.
The resulting peony fermentation filtrate was subjected to routine data testing as shown in table 2 below:
TABLE 2
Performance testing
1. The peony root fermentation filtrate prepared in example was subjected to an anti-inflammatory soothing test:
in vitro cultured RAW264.7 macrophage as model, 0.15%, 0.3%, 0.6% W/W peony fermentation filtrate was added, 1. mu.g/ml LPS (lipopolysaccharide) was added as a stimulator after 2h of culture, and the concentrations of inflammatory factors TNF-alpha, IL-1, IL-8, MCP-1, PGE2 were measured after 24h, and the results are shown in Table 3 and the intracellular SOD and ROS concentrations were measured as shown in Table 4.
TABLE 3
TABLE 4
The results show that: LPS enables the concentration of inflammatory factors generated by macrophages to be remarkably improved, the concentration of various inflammatory factors can be remarkably reduced by pre-culturing peony fermentation filtrates with different concentrations for 2h before contacting with LPS, meanwhile, the oxidation resistance in cells is improved, the SOD enzyme activity is remarkably improved, and the ROS is remarkably reduced. Analysis of signal path related protein shows that the protein can reduce PPAR gamma expression and JNK, p38 and IKK beta phosphorylation, block signal transmission, and has the same anti-inflammatory action mechanism as paeonol.
2. The peony root fermentation filtrate prepared in example was subjected to an antioxidant test:
in vitro test of 0.10%, 0.30%, 0.50% W/W concentration of peony fermentation filtrate on hydroxyl radical clearance (Fe-salicylic acid method), superoxide anion radical clearance (pyrogallol method), total antioxidant capacity (ammonium molybdate method), Fe 3+ Reduction (FeCl) 3 Method) lipid peroxidation inhibition (thiobarbituric acid method), DPPH radical clearance (DPPH method), and the data are reported in table 5.
TABLE 5
3. The peony root fermentation filtrate prepared in example was subjected to melanin suppressor cell test:
the human melanoma HM3KO cells were used as the experimental subjects, and the cells were expanded and cultured in MEM medium at 3 × 10 5 The density of each bottle is inoculated in a T75 bottle and kept stand overnight until the density of each bottle is reachedThe cells were attached to the wall of the flask, and the medium was replaced with MEM medium containing 1000ppm of the fermentation filtrate of the different examples, and MEM medium containing no fermentation filtrate was used as a control. Fresh medium was replaced at 3 day intervals until the flask was full of cultured cells. The cells were collected, washed with PBS, dissolved in a 1mol/L aqueous solution of sodium hydroxide, and the absorbance at 500nm was measured. The melanin production inhibition was calculated.
Melanin production inhibition% = (control absorbance-experimental absorbance)/control absorbance 100%.
The data are recorded in table 6.
TABLE 6
4. The fermentation broths prepared in examples 1-3 and the blanks (without fermentation broths) were tested for human skin stinging and discomfort by the following method:
test objects: selecting 20 female volunteers, and 30-50 years old; skin care products should not be used during the test.
The test method comprises the following steps: after cleaning the face, wait 30 minutes. The cheek and the nasolabial sulcus of a volunteer are selected as test areas, 10% lactic acid is respectively used for a patch test, after 5 minutes, the skin generates stabbing pain and discomfort, the use example obtains a product with the concentration of 1.0% W/W and no product (blank), the stabbing pain and discomfort of the test part are measured after 0 minute, 5 minutes, 15 minutes and 30 minutes, the score is carried out according to a 4-point method (0 is no stabbing pain, 1 is mild stabbing pain, 2 is moderate stabbing pain and 3 is severe stabbing pain), and the average score is calculated and used as an index for evaluating the skin sensitivity. The specific detection results are shown in fig. 2.
5. The fermentation broth prepared in example 1 was used for the test of skin soothing sensitivity of human:
testing an instrument: skin tester VISIA 7
Test objects: selecting 20 persons with slight skin sensitivity, and 30-50 years old;
the using method comprises the following steps: after cleaning the face, the emulsion of 1.0% W/W peony root fermented filtrate was applied to the face, and after 15 minutes, it was removed and massaged until absorption. 1 time daily, 1 test per week, and 4 weeks of connected tests. No other products must be used during the test.
As a result: the red areas of the face were tested by the skin tester VISIA 7 and the overall reduction in the red areas of the volunteer's face was in the range of 28-45.5%. The 1.0% W/W peony root fermented filtrate has better effects on relieving sensitivity and repairing red blood streak, wherein the effect of the red area of the face is shown in figure 3, and the effect of the red blood streak of the face is shown in figure 4.
Claims (1)
1. A peony fermentation filtrate is characterized in that preparation raw materials comprise, by weight, 6 parts of peony root fine powder, 8.4 parts of Lactobacillus plantarum (Lactobacillus plantarum), 4 parts of glucose, 0.4 part of vitamin B group, 0.6 part of sodium chloride, 1.2 parts of preservative and 89 parts of water; wherein the lactobacillus plantarum is lactobacillus plantarum secondary seed liquid;
the fermentation process of the peony fermentation filtrate comprises the following steps:
(1) pulverizing and sieving radix moutan to obtain 60 mesh fine powder;
(2) mixing fine powder of peony root, glucose, vitamin B group, sodium chloride, and water, sterilizing at 121 deg.C under 0.105MPa for 30min to obtain sterilized culture medium, and cooling;
(3) transferring lactobacillus plantarum into 100mL of MRS liquid culture medium, performing static culture at 35 +/-1 ℃ to obtain first-stage seed liquid, performing amplification culture on the first-stage seed liquid in the MRS liquid culture medium, and performing static culture at 35 +/-1 ℃ for 18h to obtain second-stage seed liquid;
(4) transferring the secondary seed liquid into a sterilization culture medium, and culturing at constant temperature of 35 +/-1 ℃ for 72h, wherein the stirring speed of a constant temperature oscillator is 80r/min, so as to obtain a fermentation liquid;
(5) carrying out primary high-temperature treatment on the fermentation liquor at the temperature of 75 ℃ for 15min, carrying out ultrasonic treatment at the ultrasonic power of 400W and the temperature of 45 ℃ for 50min after cooling to room temperature to obtain treated fermentation liquor, then carrying out filtration and deslagging to obtain clarified liquid, carrying out secondary high-temperature treatment on the clarified liquid at the temperature of 85 ℃ for 25min, cooling to room temperature, and adding 0.2wt% of p-hydroxyacetophenone and 1.0wt% of 1, 2-hexanediol to obtain a peony fermentation filtrate.
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| CN113197784A (en) * | 2021-04-13 | 2021-08-03 | 北京源天彩生物科技有限公司 | Preparation method and application of hair dyeing product containing melanin precursor prepared by biological fermentation |
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| CN113197784A (en) * | 2021-04-13 | 2021-08-03 | 北京源天彩生物科技有限公司 | Preparation method and application of hair dyeing product containing melanin precursor prepared by biological fermentation |
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