Fermented peony seed oil as well as preparation method and application thereof

Abstract

The invention discloses a preparation method of fermented peony seed oil. The fermentation process is optimized, different strains such as lactic acid bacteria, saccharomycetes and bifidobacteria are innovatively combined for multiple times of fermentation, and the lactic acid bacteria and saccharomycetes mixed strains are used for primary fermentation, so that components are preliminarily decomposed, and the viscosity is reduced; a bifidobacterium strain is used for secondary fermentation, so that the components are further optimized. After fermentation, supercritical CO2 extraction and freezing purification are performed to obtain the high-quality peony seed oil. According to the process, the content of monounsaturated fatty acid oleic acid in the peony seed oil is increased to 80.06%, the content of polyunsaturated fatty acid linoleic acid and linolenic acid is reduced, the product stability is improved, the skin feeling is fresh and cool when the peony seed oil is used, the permeation rate is increased, the DPPH free radical scavenging rate is increased, and the peony seed oil has an anti-eczema effect.

Description

Technical Field

[0001] This invention belongs to the field of natural product technology, specifically relating to fermented peony seed oil prepared through a double microbial fermentation process and its use in skin care products such as anti-aging, antioxidant, and anti-eczema products. By optimizing fermentation process parameters and bacterial strain combinations, this invention significantly improves the permeability and sensory experience of peony seed oil, protects against cellular oxidative stress damage, and enhances its in vitro antioxidant activity, making it suitable for a variety of skin care product forms, including essence oils, creams, lotions, and masks. Background Art

[0002] Peony seed oil, an emerging plant oil resource, has recently attracted significant attention for its unique chemical composition and remarkable biological activity. Peony seeds contain 24.13-37.83% oil. Its chemical composition reveals that peony seed oil is rich in unsaturated fatty acids, comprising 82-93% unsaturated fatty acids. Linoleic, linolenic, and oleic acids are predominant among these unsaturated fatty acids, with α-linolenic acid comprising a whopping 42%. This essential fatty acid has been shown to lower cholesterol and blood lipids, prevent cardiovascular disease, enhance immunity, combat allergies, combat aging, promote fat metabolism, and promote liver cell regeneration. Linoleic acid inhibits cholesterol synthesis, has anti-cancer and antioxidant properties, and can prevent diabetes. Peony seed oil also contains a variety of active ingredients, including polyphenols, anthocyanins, vitamins A / E, phytosterols, and minerals. These ingredients possess potent antioxidant, anti-inflammatory, and skin barrier repair properties, demonstrating significant potential for use in skincare products.

[0003] However, despite its numerous advantages, peony seed oil still faces some pressing challenges in its practical application. First, the peony seed oil extracted using traditional methods has a relatively long molecular chain, resulting in high viscosity and poor permeability. This makes it difficult for peony seed oil to be quickly absorbed by the skin when applied to the surface, hindering its full skincare benefits. For example, in some anti-aging skincare products, if peony seed oil cannot effectively penetrate deep into the skin, it will not provide sufficient nutrients to skin cells, making it difficult to achieve the desired anti-aging effect.

[0004] Secondly, the release efficiency of active ingredients in traditional peony seed oil is low. Active ingredients such as polyphenols and anthocyanins are often encapsulated within oil molecules or bound to other ingredients, making it difficult for them to fully release and interact with skin cells to exert their effects. This not only reduces the skincare benefits of peony seed oil but can also cause some active ingredients to gradually inactivate during storage or use, affecting product quality and stability.

[0005] Furthermore, from a sensory perspective, traditional peony seed oil has a greasy texture, forming a thick, oily film on the skin after application, causing discomfort. This greasy feeling is particularly noticeable in summer or for people with oily skin, and can easily lead to clogged pores, acne, and other skin problems, thus limiting its widespread use in skincare products.

[0006] Furthermore, traditional peony seed oil also has shortcomings in its emulsification capacity. Due to limitations in its molecular structure and surface properties, peony seed oil has poor emulsification with other skincare ingredients (such as aqueous phase components), making it difficult to form a stable emulsion or cream system. This not only affects the product's appearance and texture, but can also lead to delamination and deterioration during use, reducing its effectiveness and safety.

[0007] In summary, developing a preparation method that can effectively solve the above-mentioned problems of traditional peony seed oil is of great significance for giving full play to the efficacy of peony seed oil in the field of skin care products and improving product quality and market competitiveness. Summary of the Invention

[0008] To address the above-mentioned problems in the prior art, the present invention provides a fermented peony seed oil. Compared with traditional plant oils and fats, the fermented peony seed oil of the present invention has a lighter texture and a pleasant odor. It also has strong antioxidant, anti-aging, and anti-eczema properties. Compared with unfermented peony seed oil, the content of monounsaturated fatty acids such as oleic acid is increased, making it suitable as a good cosmetic base oil.

[0009] The present invention also provides a preparation method and application of the fermented peony seed oil. This method, through a double fermentation process, further addresses the challenges of traditional technologies, such as incomplete release of active ingredients, a heavy feel on the skin, and low bioavailability, while also improving product safety and long-lasting efficacy.

[0010] In order to achieve the above-mentioned object of the invention, the present invention adopts the following technical solutions: A method for preparing fermented peony seed oil comprises the following steps: 1) Primary Fermentation: For the first fermentation, lactic acid bacteria and yeast are selected as strains (lactic acid bacteria can produce organic acids to lower the pH value and inhibit the growth of miscellaneous bacteria; yeast can secrete various enzymes to initially decompose long-chain fatty acids). These strains are added to YPD medium to prepare lactic acid bacteria seed liquid and yeast seed liquid, respectively. Peony seed oil, the lactic acid bacteria seed liquid, and the yeast seed liquid are added to the YPD medium and mixed under a sterile environment for the first fermentation to obtain a primary fermentation product. 2) Secondary fermentation: The second fermentation uses bifidobacteria (bifidobacteria can secrete proteases, lipases, etc. to further decompose residual macromolecules). After filtering the primary fermentation product, bifidobacteria are inoculated for a second fermentation to obtain the secondary fermentation product. 3) Post-processing: Supercritical CO2 extraction technology and cryo-purification technology are used to extract and purify the secondary fermentation product to obtain clear, transparent and fragrant fermented peony seed oil.

[0011] Specifically, in step 1), the YPD medium is prepared by the following steps: taking 9 to 11 g of glucose, 9 to 11 g of peptone, and 4 to 6 g of yeast extract, adding 0.9 to 1.1 L of deionized water, and then sterilizing at 99 to 108°C for 26 to 34 min.

[0012] Specifically, in step 1), the lactic acid bacteria seed liquid or yeast seed liquid is prepared by the following steps: placing the frozen tubes of lactic acid bacteria or yeast in an incubator at a temperature of 25-27°C and a humidity of 75-85% for activation for 10-14 hours, and then inoculating the lactic acid bacteria or yeast into a pre-sterilized YPD culture medium at a mass ratio of 1.8-2.4:1, and culturing at 25-27°C in a shaking incubator for 60-84 hours.

[0013] Furthermore, in step 1), the conditions for the first fermentation culture are as follows: peony seed oil, YPD medium, lactic acid bacteria seed liquid, and yeast seed liquid are mixed in a sterile environment at a volume ratio of 26-31:26-31:1:1, and then fermented in a constant temperature shaking incubator at 31-38°C for 36-52 hours at a rotation speed of 140-160 rpm; the pH value is regularly tested during the fermentation process to maintain the pH between 4.0 and 4.5.

[0014] Specifically, in step 2), the second fermentation culture conditions are as follows: after filtering the first fermentation product, inoculating Bifidobacterium at a 2.5-3.6% inoculum, and then fermenting at a temperature of 26-31°C and a light intensity of 1800-2200 lux for 60-84 hours. During the fermentation process, sterile water is regularly added to maintain a stable volume of the fermentation broth.

[0015] Specifically, in step 3), the supercritical CO2 extraction conditions are as follows: the secondary fermentation product is subjected to supercritical CO2 extraction at a pressure of 22-29 MPa and a temperature of 35-45°C for 1-3 hours to separate the fermented peony seed oil and remove moisture, impurities and residual bacteria.

[0016] Furthermore, in step 3), the freeze purification conditions are as follows: freezing the fermented peony seed oil after CO2 extraction at -4~4°C for 18~24h, and filtering out the precipitated crystals at 10~15°C to obtain clear, transparent and fragrant fermented peony seed oil.

[0017] The present invention provides fermented peony seed oil prepared by the above preparation method.

[0018] The present invention also provides the use of the fermented peony seed oil in the preparation of anti-aging and / or anti-oxidation foods, skin care products or medicines.

[0019] The present invention also provides the use of the fermented peony seed oil in preparing anti-eczema skin care products or medicines.

[0020] The present invention discloses a preparation method of fermented peony seed oil. The present invention optimizes the fermentation process and innovatively adopts a combination of different strains such as lactic acid bacteria, yeast and bifidobacterium to carry out multiple fermentations. A mixed strain of lactic acid bacteria and yeast is used for the first fermentation to preliminarily decompose the components and reduce the viscosity; a bifidobacterium strain is used for the second fermentation to further optimize the components. After fermentation, the oil is extracted by supercritical CO2 and frozen for purification to obtain high-quality peony seed oil. This process increases the content of monounsaturated fatty acid oleic acid in peony seed oil to 80.06%, reduces the content of polyunsaturated fatty acids linoleic acid and linolenic acid, improves product stability, and makes the skin feel refreshed, the penetration rate is increased, the DPPH free radical scavenging rate is increased, and the oil has an anti-eczema effect. The present invention provides high-efficiency and high-quality raw materials for the field of skin care products, and has broad prospects.

[0021] Compared with the prior art, the present invention has the following advantages and beneficial effects: 1) Ingredient optimization: The double fermentation process of the present invention increases the content of monounsaturated fatty acid oleic acid from 22.64% in traditional peony seed oil to 80.06%.

[0022] 2) Sensory enhancement: The fermented peony seed oil obtained after fermentation of the present invention has a refreshing and non-greasy skin feel, a fresh and elegant smell, is clear and transparent, and has an improved penetration rate.

[0023] 3) Anti-aging effect: This invention strengthens the skin barrier function, reduces water loss, improves collagen synthesis ability, delays the skin aging process, and has a strong effect on the common DPPH, ABTS, O 2- The ability to scavenge free radicals is enhanced.

[0024] 4) Anti-eczema effect: The present invention reduces the release of the specific inflammatory factor thymic stromal lymphopoietin (TSLP), thereby reducing the intensity of itching and the area of ​​skin lesions, thereby achieving the purpose of anti-eczema. BRIEF DESCRIPTION OF THE DRAWINGS

[0025] Figure 1 This is a comparison chart of the HPLC test results of the peony seed oil of Comparative Example 1 (Figure A) and the fermented peony seed oil of Example 1 (Figure B). DETAILED DESCRIPTION

[0026] The following will clearly and completely describe the technical solutions in the embodiments of the present invention in conjunction with the present invention. Obviously, the described embodiments are only part of the embodiments of the present invention, not all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without making creative efforts are within the scope of protection of the present invention.

[0027] In the following examples, the lactic acid bacteria (Lactobacillus fermentum) CGMCC No. 1.15608 and the yeast (Saccharomyces cerevisiae) CGMCC No. 2.3973 used were purchased from the China General Microorganism Culture Collection Center, and the Bifidobacterium (Bifidobacterium bifidum) CGMCC No. 1.5091 was purchased from the China General Microorganism Culture Collection Center.

[0028] Peony seed oil was purchased from Heze Zhonghe Jianyuan Biotechnology Co., Ltd.

[0029] Example 1 A method for preparing a peony seed oil fermented product comprises the following steps: 1. Primary fermentation (1) Preparation of strain seed solution YPD medium: Take 10 g of glucose, 10 g of peptone, and 5 g of yeast extract, add 1 L of deionized water, and then sterilize at 105°C for 30 min.

[0030] Seed solution preparation: Place the frozen tubes of lactic acid bacteria or yeast in a mold incubator at a temperature of 26°C and a humidity of 80% for activation overnight (12 hours). Then, inoculate the lactic acid bacteria or yeast into pre-sterilized YPD medium at a mass ratio of 2:1 and culture in a shaking incubator at 26°C for 72 hours to obtain the seed solution of lactic acid bacteria or yeast.

[0031] (2) Fermentation: 280 mL of peony seed oil was mixed under sterile conditions with a volume ratio of 28:28:1:1 for peony seed oil, YPD medium, lactic acid bacteria seed solution, and yeast seed solution. The mixture was then fermented in a constant temperature shaking incubator at 35°C for 48 hours at 150 rpm to obtain a primary fermentation product. The pH value was regularly monitored during the fermentation process and maintained between 4.0 and 4.5.

[0032] 2. Secondary fermentation Fermentation conditions: After filtering the primary fermentation product, inoculate with Bifidobacterium (3%) and ferment at 28°C under 2000 lux for 72 hours to obtain the secondary fermentation product. Sterile water was regularly added during the fermentation process to maintain a stable volume.

[0033] 4. Post-processing Supercritical CO2 extraction: The secondary fermentation product was subjected to supercritical CO2 extraction at a pressure of 25 MPa and a temperature of 40°C for 2 h to separate the fermented peony seed oil and remove moisture, impurities and residual bacteria.

[0034] Freeze purification technology: The fermented peony seed oil after CO2 extraction is frozen at 2°C for 18 hours, and the precipitated crystals are filtered out at 15°C to obtain clear, transparent and fragrant fermented peony seed oil.

[0035] Comparative Example 1 The sample of this comparative example is peony seed oil that has not been fermented.

[0036] Comparative Example 2 The difference between the sample of this comparative example and the example is that the fermented product is obtained by the first fermentation using only lactic acid bacteria and yeast.

[0037] Comparative Example 3 The preparation of peony seed oil fermented with Bifidobacterium involves the following steps: culturing Bifidobacterium in a high-pressure environment to prepare a seed solution; adding the peony seed oil and Bifidobacterium seed solution to YPD medium (inoculation rate: 3%); and fermenting for 72 hours at 28°C and 2000 lux of light intensity. Sterile water is regularly added during the fermentation process to maintain a stable volume of the fermentation solution.

[0038] Preparation of bifidobacterium seed liquid: Place the frozen tubes of bifidobacteria in a mold incubator at a temperature of 26°C and a humidity of 80% for activation overnight (12 hours), and then inoculate bifidobacteria into pre-sterilized YPD culture medium at a 5% inoculum rate, and culture in a shaking incubator at 26°C for 48 hours to obtain the bifidobacterium seed liquid.

[0039] Comparative Example 4 The preparation of peony seed oil through co-fermentation with lactic acid bacteria, yeast, and bifidobacteria involves the following steps: co-culturing lactic acid bacteria, yeast, and bifidobacteria in a high-pressure environment to prepare a mixed seed liquid; adding the peony seed oil and mixed seed liquid to YPD medium (inoculation rate: 3%); and fermenting at 28°C under a light intensity of 2000 lux for 72 hours. Sterile water is regularly added during the fermentation process to maintain a stable volume of the fermentation liquid.

[0040] Preparation of mixed seed liquid of lactic acid bacteria, yeast and bifidobacterium: Place the frozen tubes of lactic acid bacteria, yeast and bifidobacterium in a mold incubator at a temperature of 26°C and a humidity of 80% for activation overnight (12 hours), and then inoculate lactic acid bacteria, yeast and bifidobacteria into pre-sterilized YPD culture medium at an inoculum rate of 5%, and culture in a shaking incubator at 26°C for 48 hours to obtain a mixed seed liquid of lactic acid bacteria, yeast and bifidobacteria.

[0041] Test Example 1 The fermented peony seed oil obtained in Example 1 and the peony seed oil in Comparative Example 1 were subjected to high performance liquid chromatography (HPLC) and compared with a reference substance. The specific test steps are as follows: (1) Chromatographic conditions: Agilent Poroshell 120 EC-C18 (3.0×50 nm, 2.7 μm) column, DAD detector; detection wavelength 214 nm; column temperature 30°C; flow rate 0.6 mL / min; injection volume 10 μL; mobile phase: acetonitrile (A)-0.1% phosphoric acid (B); gradient elution order: 0 to 8 min, A 8% to 16%, B 92% to 84%; 8 to 19 min, A 16% to 60%, B 84% to 40%; 19 to 30 min, A 60% to 90%, B 40% to 10%.

[0042] (2) Preparation of reference solution: Accurately weigh appropriate amounts of myristic acid, palmitic acid, palmitoleic acid, heptadecanoic acid, cis-10-heptadecenoic acid, stearic acid, oleic acid, linoleic acid, α-linolenic acid and cis-11-eicosenoic acid reference substances, place them in a 50 mL brown volumetric flask, add 80% methanol to dissolve and dilute to the mark to obtain mixed reference solutions with concentrations of 25.84 μg / mL, 29.21 μg / mL, 38.54 μg / mL, 26.59 μg / mL, 24.51 μg / mL, 51.84 μg / mL, 48.48 μg / mL, 42.21 μg / mL, 27.25 μg / mL and 26.87 μg / mL, respectively.

[0043] (3) Preparation of test solution: 125 μL of the peony seed oil obtained in Comparative Example 1 and the fermented peony seed oil obtained in Example 1 were accurately pipetted into a 25 mL volumetric flask, 23 mL of methanol was added, and ultrasonication was performed for 30 min to fully dissolve. After cooling, the solution was diluted to the mark with methanol and shaken to obtain the solution.

[0044] (4) Chromatographic conditions: Agilent C18 high-performance chromatographic column (3.0 mm × 50 mm, 2.7 μm), mobile phase: acetonitrile-0.1% phosphoric acid aqueous solution (volume ratio 85:15), flow rate 1 mL / min, column temperature 35°C, detection wavelength 203 nm, injection volume 20 μL.

[0045] (5) Results, see Figure 1 and Table 1.

[0046] Table 1 Changes in relative content of fatty acids before and after fermentation The HPLC fingerprints of 10 fatty acids in peony seed oil before fermentation (Comparative Example 1) and after fermentation (Example 1) are as follows: Figure 1 The relative peak area results are shown in Table 1. After fermentation, the oleic acid content increased from 22.63% to 80.06%, the linoleic acid content decreased from 28.73% to 8.59%, and the linolenic acid content decreased from 40.58% to 0.67%.

[0047] Test Example 2 Antioxidant Ability The test objects were the peony seed oil products prepared in Example 1 and Comparative Examples 1 to 4.

[0048] (1) DPPH free radical scavenging rate test: Preparation of DPPH solution (0.2 mmol / L): Weigh 0.0197 g of DPPH, dissolve it in anhydrous ethanol, and dilute to 250 mL. Place 2 mL of sample in a centrifuge tube, add 2 mL of DPPH solution, shake well, and incubate in the dark for 15 minutes. Measure the absorbance (A1) at 515 nm. Also, add 2 mL of sample and 2 mL of anhydrous ethanol, shake well, incubate in the dark for 15 minutes, and measure the absorbance (A2) at 515 nm. Also, add 2 mL of anhydrous ethanol and 2 mL of DPPH solution, shake well, incubate in the dark for 15 minutes, and measure the absorbance (A0) at 515 nm. The DPPH radical scavenging rate is calculated using the following formula.

[0049] DPPH clearance rate%=[1-(A1-A2) / A0]×100%.

[0050] (2) ABTS free radical scavenging rate Preparation of ABTS working solution: Mix 0.2 mL of ABTS solution (7.4 mmol / L) and 0.2 mL of K2S2O8 solution (2.6 mmol / L). Incubate in the dark at room temperature for 12 h. Dilute the solution 40-50 times with anhydrous ethanol to an absorbance of 0.7 ± 0.2 at 734 nm. Store the diluted solution in a refrigerator at 4°C in the dark until ready for use.

[0051] Place 0.8 mL of ABTS working solution in a centrifuge tube, add 100 μL of sample solution (diluted 100-fold), mix thoroughly, oscillate for 20 seconds, let stand for 6 minutes, and measure the absorbance (A) at 734 nm. Using distilled water as a control, measure the absorbance (A0) under the same conditions. Calculate the ABTS free radical scavenging rate using the following formula.

[0052] ABTS free radical scavenging rate%=(A0-A) / A ×100%.

[0053] (3) O2 - Free radical scavenging rate Take three 10 mL test tubes, add 2.5 mL of Tris-HCl solution (50 mmol / L) to test tube 1, then add 1 mL of sample and 0.6 mL of pyrogallol solution (20 mmol / L), mix well, place the test tube in a 25°C water bath for 20 minutes, and finally add 0.5 mL of HCl to terminate the reaction. Measure the absorbance of the solution at 320 nm. j ; In test tube 2, water is used instead of pyrogallol solution, and the subsequent treatment is the same as before, and the absorbance value of the solution is measured. i ; In test tube 3, water is used to replace the sample solution, and the subsequent treatment is the same as before, and the absorbance value of the solution A0 is measured. - The free radical scavenging rate was calculated using the following formula.

[0054] O2 - Clearance % = (A j -A i ) / A0 ×100%.

[0055] (4) Results: Three sets of parallel samples were tested for each sample to be tested. The test results are shown in Table 2.

[0056] Table 2 DPPH, ABTS and O2 - Free radical scavenging rate Different letters in the table represent significant differences. P <0.05.

[0057] In Table 2, the scavenging rates of Example 1 and Comparative Examples 1 to 4 on DPPH free radicals varied between 66.21% and 91.21%. Example 1 had the strongest antioxidant capacity (91.21%), followed by Comparative Example 4 (76.35%). As for the ABTS free radical scavenging capacity of fermented peony seed oil, Example 1 had the strongest free radical scavenging capacity (87.89%), followed by Comparative Example 4 (80.54%). - During the experiment, O2- The free radical scavenging rate values ​​ranged from 70.68% to 88.97%. Example 1 had the strongest antioxidant capacity (88.97%), followed by Comparative Example 4 (77.79%). The in vitro antioxidant test results showed that Example 1 had the best in vitro antioxidant capacity compared to the other comparative examples.

[0058] Test Example 3 Anti-eczema The normal 3D epidermal model EpiKutis (EpiKutis ® RHE, Guangdong Boxi Biotechnology Co., Ltd.) were transferred to a 6-well plate to which 0.9 mL EpiLife™ culture medium (MEPI500CA, ThermoFisherScientific) had been added in advance and divided into a blank control group (BC), a negative control group (NC), a positive control group (PC), and sample test groups (Example 1 group, Comparative Examples 1, 2, 3, and 4 groups). An eczema-like model was constructed by adding 25 μL of 24 μg / mL Poly I∶C (CSGC14710, Wuhan Costan Biotechnology Co., Ltd.) and 50 μL of 20 μg / mL LPS solution to the surface of the NC group model. In the PC group, 12.5 μL of 48 μg / mL Poly I∶C, 50 μL of 40 μg / mL LPS solution, and 12.5 μL of 0.01% dexamethasone were added to the surface of the model. In the sample test group, 12.5 μL of 48 μg / mL Poly I∶C, 50 μL of 40 μg / mL LPS solution, and 12.5 μL of the test sample were added to the surface of the model. The models were then incubated in a CO2 incubator (37°C, 5% CO2, relative humidity 96%) for 24 h. After incubation, the test substances remaining on the surface of the model were washed with sterile PBS, and the residual liquid inside and outside the model was wiped off with a sterile cotton swab. The TSLP content was detected by ELISA kit.

[0059] Data processing: Each test was repeated three times, and the results were expressed as "mean ± SD". Statistical differences between groups were analyzed using t-test or ANOVA with SPSS 26. P > 0.05 was considered to be no significant difference, and p < 0.05 and p < 0.01 were considered to be significant and extremely significant differences.

[0060] TSLP can induce itch by directly stimulating cutaneous sensory nerve fibers. TSLP expression levels are positively correlated with itch intensity and lesion area, and can be used as an indicator of disease activity and severity in patients with eczema. Table 3 shows the TSLP content in 3D epidermal models from the five groups.

[0061] Table 3 TSLP content of 3D epidermal model Different letters in the table represent significant differences. P <0.05.

[0062] Table 3 shows the TSLP content (pg / mL) of different groups (BC, NC, PC, Example 1, and Comparative Examples 1-4). The mean and standard deviation for each data set are provided, and significant differences between groups are indicated with letters (a, b, c, d). The results for the PC group validated the effectiveness of the experimental system, demonstrating that the positive control treatment significantly reduced TSLP expression. The TSLP content in Example 1 was similar to that in the PC group and significantly lower than that in the NC group, indicating that the sample in Example 1 was significantly effective in reducing TSLP expression. The TSLP content in Comparative Example 1 was similar to that in the NC group, indicating that its sample had no significant effect. The TSLP content in Comparative Examples 2 and 3 (and Comparative Example 4 as a supplementary reference), while lower than that in the NC group, was higher than that in the PC group and Example 1, indicating that these samples or treatments had limited effectiveness in reducing TSLP expression.

[0063] Test Example 4 Sensory Evaluation 1. Scoring criteria.

[0064] Sixty trained volunteers (30 men and 30 women) were asked to evaluate the sensory experience of the peony seed oils prepared in Example 1 and Comparative Examples 1-4, and their average scores were calculated. Transparency, odor, and flavor were measured according to GB / T 5525-2008, "Test Methods for Transparency, Odor, and Flavor of Vegetable Oils and Fats." Detailed scoring criteria for the sensory evaluation of fermented oils are shown in Table 4.

[0065] Table 4 Scoring criteria for sensory evaluation of fermented oil 2. Rating results: The sensory rating results are shown in Table 5.

[0066] Table 5 Sensory evaluation results As can be seen in Table 5, the sensory properties of peony seed oil are better after fermentation, and the effect is better after two fermentations compared to one. Peony seed oil has a distinct, heavy, oily smell before fermentation, but after fermentation, the smell becomes lighter and the aroma becomes clearer. The color is slightly lighter after fermentation, making it more suitable for subsequent processing. As for transparency, the transparency is good before and after fermentation, proving that the fermentation process does not make the oil turbid. The skin feel before and after fermentation differs significantly. Before fermentation, the skin feel is heavy and sticky, but after fermentation, the skin feel is refreshing and has strong ductility. Absorption is significantly improved after fermentation, and it penetrates the skin more quickly after application. This may be because the bacterial strain metabolites during the fermentation process break down the larger molecular weight fatty acid esters into smaller molecular weight fatty acids, such as oleic acid, making them more easily absorbed by the body and more refreshing, without the heavy feeling before processing.

Claims

1. A method for preparing fermented peony seed oil, characterized in that: The following steps are involved: 1) Primary Fermentation: For the first fermentation, lactic acid bacteria and yeast strains are selected and added to YPD medium to prepare lactic acid bacteria seed liquid and yeast seed liquid, respectively. Peony seed oil, the lactic acid bacteria seed liquid, and the yeast seed liquid are added to the YPD medium and mixed under a sterile environment for the first fermentation to obtain a primary fermentation product. 2) Secondary fermentation: The second fermentation uses bifidobacteria. After filtering the primary fermentation product, bifidobacteria are inoculated for a second fermentation culture to obtain the secondary fermentation product. 3) Post-processing: Apply supercritical CO2 extraction technology and freezing purification technology to extract and purify the secondary fermentation product.

2. The method for preparing fermented peony seed oil according to claim 1, wherein In step 1), the YPD medium is prepared by the following steps: 9 to 11 g of glucose, 9 to 11 g of peptone, and 4 to 6 g of yeast extract are added with 0.9 to 1.1 L of deionized water, and then sterilized at 99 to 108°C for 26 to 34 min.

3. The method for preparing fermented peony seed oil according to claim 1, wherein In step 1), the lactic acid bacteria seed liquid or yeast seed liquid is prepared by the following steps: placing the frozen tubes of lactic acid bacteria or yeast in an incubator at a temperature of 25-27°C and a humidity of 75-85% for activation for 10-14 hours, and then inoculating the lactic acid bacteria or yeast into a pre-sterilized YPD culture medium at a mass ratio of 1.8-2.4:1, and culturing at 25-27°C in a shaking incubator for 60-84 hours to obtain the product.

4. The method for preparing fermented peony seed oil according to claim 1, wherein In step 1), the conditions for the first fermentation culture are as follows: peony seed oil, YPD medium, lactic acid bacteria seed liquid, and yeast seed liquid are mixed in a sterile environment at a volume ratio of 26-31:26-31:1:1, and then fermented in a constant temperature shaking incubator at 31-38° C. for 36-52 hours; the pH is maintained between 4.0 and 4.5 during the fermentation process.

5. The method for preparing fermented peony seed oil according to claim 1, wherein In step 2), the conditions for the second fermentation culture are as follows: after filtering the first fermentation product, inoculating bifidobacteria at an inoculum rate of 2.5-3.6%, and then fermenting at a temperature of 26-31°C and a light intensity of 1800-2200 lux for 60-84 hours.

6. The method for preparing fermented peony seed oil according to claim 1, wherein In step 3), the supercritical CO2 extraction conditions are as follows: the secondary fermentation product is subjected to supercritical CO2 extraction at a pressure of 22-29 MPa and a temperature of 35-45° C. for 1-3 h.

7. The method for preparing fermented peony seed oil according to claim 1, wherein In step 3), the freeze purification conditions are as follows: the fermented peony seed oil after CO2 extraction is frozen at -4~4°C for 18~24 hours, and the precipitated crystals are filtered out at a temperature of 10~15°C to obtain the product.

8. Fermented peony seed oil prepared by the preparation method according to any one of claims 1 to 7.

9. Use of the fermented peony seed oil according to claim 8 in the preparation of anti-aging and / or antioxidant foods, skin care products or medicines.

10. Use of the fermented peony seed oil according to claim 8 in the preparation of anti-eczema skin care products or medicines.

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