Oil-soluble plant compound and preparation method thereof

Abstract

The invention provides an oil-soluble plant compound and a preparation method thereof, and belongs to the field of medical, dental or cosmetic preparations. The composition consists of chrysanthellum indicum, ligusticum wallichii, frankincense, exocarpium citri grandis essential oil and a soybean-radix puerariae co-fermentation product. The technical core is as follows: soybean and radix puerariae are subjected to synergistic liquid fermentation by using aspergillus oryzae, lysophosphatide, soybean polypeptide and puerarin aglycone are generated in situ through biotransformation, and an endogenous self-emulsifying carrier is constructed. According to the process, the transdermal penetration promoting capacity of active components is remarkably enhanced by utilizing the regulating effect of ligusticum wallichii oil on cuticle lipid arrangement and combining a micro-micelle structure formed by a co-fermentation system. Nanoscale self-solubilization of four essential oil matrixes in a fermentation substrate is achieved, the skin barrier repair and inflammation inhibition effects are enhanced through multi-target cooperation, and the bioavailability, penetration depth and safety of active ingredients are greatly improved.

Description

Technical Field

[0001] This invention belongs to the field of formulations for medical, dental or cosmetic use, and particularly relates to an oil-soluble plant complex and its preparation method. Background Technology

[0002] In the fields of modern cosmetics and biotechnology, the research and development of soothing and repairing products has always held a central position. With the increasing density of people with sensitive skin and the widespread prevalence of skin barrier damage, developing highly bioactive repair compositions has become a key area of ​​technological innovation in the industry. Traditional repairing products often rely on simple blends of plant extracts or the addition of synthetic chemical ingredients. However, under current technological conditions, achieving efficient delivery, stable release, and improved bioavailability of natural active ingredients still faces multiple technical bottlenecks.

[0003] Plant essential oils, as a class of natural components with significant bioactivity, have shown great potential in the field of skin repair due to their small molecular weight, strong penetrability, and multi-dimensional biological functions. However, in practical applications, essential oils exhibit strong hydrophobicity and instability. Current technologies typically require the addition of large amounts of synthetic surfactants, chemical cosolvents, or high concentrations of polyols to introduce these non-polar oily components into aqueous or emulsion systems. While these exogenous additives solve the solubility problem at the physical level, they also bring significant negative technical effects: on the one hand, high doses of synthetic emulsifiers may disrupt the arrangement of the skin's own lipid barrier, creating a risk of secondary irritation, which contradicts the original intention of repair product development; on the other hand, complex chemical cosolvent systems often interfere with the structural stability of plant active molecules, reducing their bioactive expression on the skin surface.

[0004] Meanwhile, bio-fermentation technology, as an important means to enhance plant efficacy, has been widely used in raw material processing. Traditional fermentation processes mostly focus on the transformation of single plant substrates, and the functional dimensions of their products are relatively limited. Moreover, the metabolites produced during fermentation often lack sufficient physicochemical characteristics to support the carrying capacity of other active components.

[0005] In the study of synergistic effects of plant-based raw materials, existing technologies mostly focus on the physical superposition of active ingredients, lacking a systematic approach from the perspective of microbial metabolic induction and intermolecular interactions. For example, whether metabolic coupling exists between different types of plant substrates during co-fermentation, and whether endogenous metabolites produced during fermentation can form spontaneous physical encapsulation systems with essential oil components of specific polarities, still lack in-depth theoretical support and mature practical solutions in current published literature. Especially under the industry trend of pursuing "green chemistry" and "simplified formulations," how to utilize biotransformation methods to generate interfacially active biomolecules in situ, thereby replacing synthetic chemicals to achieve self-solubilization of essential oils, is a key issue that urgently needs to be addressed in the field of cosmetic raw materials.

[0006] In conclusion, existing soothing and repairing compositions still struggle to achieve a perfect balance between naturalness and stability, high activity and low irritation, and the physical compatibility between complex components. Summary of the Invention

[0007] The first objective of this invention is to provide a plant composition comprising, by weight, the following components: Golden chamomile extract 5-12 Ligusticum chuanxiong oil 2-5 Frankincense oil 3-8 1-2g of Citrus reticulata essential oil Soybean-kudzu co-fermentation product: 70-90; The soybean-kudzu co-fermented product is obtained by mixing soybeans and kudzu at a mass ratio of 3-5:1, inoculating with Aspergillus oryzae for fermentation, and then extracting with a medium-to-low polar solvent.

[0008] Preferably, the soybean-kudzu co-fermentation product is extracted with a medium-low polarity solvent, then purified by ultrafiltration membrane with a molecular weight cutoff of 10,000 Da, and the permeate is collected and concentrated under reduced pressure to a solid content of ≥30%.

[0009] Preferably, the golden chamomile extract, chuanxiong oil, frankincense oil, and tangerine peel essential oil are alcohol-soluble oily components obtained from their respective plants through physical separation or solvent extraction.

[0010] Preferably, the preparation method of the soybean-kudzu co-fermentation product includes the following steps: S1. Take defatted soybeans and kudzu root, dry them, grind them into powder and mix them; add water at a material-to-liquid ratio of 1:8-12; then add at least 0.2% (w / w) potassium dihydrogen phosphate and at least 0.1% (w / w) magnesium sulfate heptahydrate based on the total mass of liquid; adjust the initial pH to between 5.5 and 6.0, sterilize and obtain the fermentation substrate; S2, Aspergillus oryzae was inoculated into an induction medium containing at least 1% (w / w) soybean lecithin for induction culture and seed culture was prepared; S3, inoculate the fermentation substrate with Aspergillus oryzae seed liquid; ferment according to the following parameters; Temperature 30±1℃, aeration rate 1.0-1.5VVM; fermentation time at least 60h; S4, after fermentation, is inactivated and centrifuged to obtain the supernatant; ethanol aqueous solution is added to the supernatant for extraction, and the alcohol extract is obtained; S5, the alcohol extract is passed through an ultrafiltration membrane with a molecular weight cutoff of 10000 Da, and the permeate is collected; the permeate is concentrated under reduced pressure until the solid content is ≥30%.

[0011] In step S3, the inoculation amount of Aspergillus oryzae seed liquid is at least 5% of the substrate volume.

[0012] In step S4, the condition for determining the completion of fermentation is: the pH value of the fermentation broth drops to its lowest point and then rises again, with the rise being at least 0.2 units, and the final pH value is ≥4.3.

[0013] In step S4, when adding an ethanol-water solution to the supernatant for extraction, the amount of ethanol-water solution added is 2-3 times the volume of the supernatant; the concentration of the ethanol-water solution is 75-95%.

[0014] A second object of the present invention is to provide a method for preparing a plant composition, comprising the following steps: S1. Preheat the soybean-kudzu co-fermentation product to 40-55℃ and keep stirring. S2, mix the remaining components and add them to the soybean-kudzu co-fermentation product while stirring to obtain a coarse mixture; S3, homogenize the crude mixture to obtain the plant composition.

[0015] A third objective of this invention is to provide the application of the aforementioned plant composition in the preparation of cosmetics. This includes, but is not limited to, dosage forms such as serums, creams, lotions, and lyophilized powders. The plant composition is present in a cosmetic at a concentration of ≥1%.

[0016] The plant composition provided by this invention is based on the use of microbial fermentation engineering to transform the natural macromolecules of soybean and kudzu into a functional matrix with high bioactivity and self-solubilizing properties, and uses this matrix as a carrier to achieve efficient and stable loading of plant essential oils.

[0017] 1. Microbial-mediated in situ phospholipid transformation and autoemulsification mechanisms Soybeans are rich in phosphatidylcholine (PC). This invention utilizes Aspergillus oryzae cultured with soybean lecithin-induced culture and its highly active extracellular phospholipases (especially Phospholipase A) during liquid fermentation to directionally hydrolyze the fatty acid chain at the sn-2 position of soybean phospholipid molecules, thereby generating lysophospholipids in situ.

[0018] Lysophospholipids possess a high hydrophilic-lipophilic balance (HLB) value, spontaneously forming extremely fine micelles in aqueous solutions. This endogenously generated biosurfactant, combined with fermented soybean peptides and plant glycolipids, constructs a natural self-emulsifying system. This system can encapsulate alcohol-soluble oily components such as golden chamomile, chuanxiong, and frankincense within nanoscale micelles, solving the technical challenge of easy precipitation and poor dispersion of non-polar active ingredients in aqueous cosmetic systems.

[0019] 2. Biotransformation enhances component polarity and absorption rate. During fermentation, β-glucosidase secreted by Aspergillus oryzae acts on puerarin compounds in kudzu root, converting them into daidzein, a smaller molecular weight, higher bioactivity, and stronger polarity. Simultaneously, soybean protein is degraded into small molecule peptides below 10,000 Da. The blend of these small molecule metabolites with essential oil components exhibits superior transdermal penetration and skin biocompatibility.

[0020] 3. Metabolic Coupling Between Minerals and Dynamic pH Regulation The addition of potassium dihydrogen phosphate and magnesium sulfate heptahydrate to the fermentation medium not only provides essential elements for microbial growth, but the magnesium ions, acting as a metal cofactor for phospholipases, significantly enhance the rate of enzymatic reactions. By monitoring specific metabolic inflection points where pH changes from a decrease to a rebound (0.2-0.5 units), the equilibrium point between the highest lysophospholipid production and the lowest excessive protein degradation can be identified, ensuring the stability of the product's physical properties.

[0021] 4. Synergistic metabolic logic of soybean-kudzu co-fermentation This invention specifically employs a co-fermentation process involving soybeans and kudzu root, rather than separate fermentations followed by physical mixing. The core of this approach lies in the induced metabolic response of microorganisms: the phospholipid components in soybeans and the flavonoid glycosides in kudzu root, within the same system, can collectively induce Aspergillus oryzae to secrete a more targeted enzyme spectrum. Experiments have shown that Aspergillus oryzae can produce more lysophospholipids in the presence of kudzu root polysaccharides.

[0022] During co-fermentation, small peptides produced from soybean protein degradation and aglycones converted from kudzu root achieve in-situ self-assembly under microscopic conditions. This spontaneously formed "peptide-phospholipid-aglycone" complex in the bioreactor exhibits higher stability and significantly enhanced encapsulation capacity (drug loading) of essential oils compared to later artificial physical mixing.

[0023] This invention utilizes defatted soybeans as a fermentation substrate (although defatted soybeans remove most of the triglycerides, they still contain approximately 0.5% - 1.5% residual phospholipids, mainly cell membrane components), effectively removing neutral fats that interfere with system stability. Simultaneously, by utilizing the residual cell membrane phospholipids and the initial phospholipids introduced during the induction culture stage, highly active lysophospholipids are accumulated in situ within the fermentation broth through the efficient biotransformation of Aspergillus oryzae. This 'enrichment and transformation' process based on a defatted matrix ensures that the final co-fermented product, with a solid content ≥30%, exhibits higher emulsification efficiency and superior skin feel compared to a full-fat matrix.

[0024] 5. Synergistic Mechanism of the Four Essential Oil Combinations Golden chamomile, rich in bisabolol and azulene, primarily acts on the skin's endogenous anti-inflammatory pathway (inhibiting cyclooxygenase-2). Frankincense, rich in boswellic acid and volatile sesquiterpenes, inhibits leukotriene production and guides repair against chronic, deep tissue damage. Ligusticum striatum, rich in ligustilide and volatile lactones, not only possesses significant sedative and anti-inflammatory effects, but its hydrophobic components also act as natural penetration enhancers, temporarily altering the lipid arrangement of the stratum corneum to facilitate the penetration of active ingredients from chamomile, frankincense, and Citrus reticulata into the deeper layers of the skin. Citrus reticulata essential oil, rich in limonene and flavonoids, effectively scavenge free radicals and inhibit the expression of pro-inflammatory factors. Its unique sesquiterpenes synergistically work with frankincense and chamomile to construct a stepwise anti-inflammatory network from the superficial barrier to deep tissues. This combination, based on "multi-target inflammation inhibition, deep repair, and efficient penetration guidance," encapsulated in a co-fermentation carrier, achieves sustained release and targeted delivery of active ingredients.

[0025] Compared with the prior art, the present invention has the following significant advantages: (1) It achieves the self-solubilizing function of all plant-derived products. This invention utilizes endogenous surfactants generated through soybean-kudzu co-fermentation technology to achieve self-solubilization of essential oils from golden chamomile, chuanxiong, frankincense, and tangerine peel. In cosmetic formulations, ideal dispersion can be achieved without the need for additional PEG-based synthetic solubilizers, significantly enhancing the product's natural properties and safety.

[0026] (2) Significantly improved the bioavailability of active ingredients The fermentation broth was purified using 10000Da ultrafiltration to remove large molecular proteins and bacterial residues, while enriching small molecular active peptides, isoflavone aglycones, and lysophospholipids. Experiments showed that the core active ingredients in the composition processed using this method had significantly better skin penetration depth and absorption rate than those obtained through physical compound extracts.

[0027] (3) Stepwise penetration enhancement and synergistic soothing effect This invention utilizes the instantaneous alteration of lipid arrangement in the stratum corneum by Ligusticum chuanxiong oil to construct a highly efficient physical permeation-enhancing channel, assisting in the targeted delivery of components such as Citrus reticulata peel and chamomile. Combined with fermented peptides and phospholipids, it forms a complex action chain of "physical permeation guidance, lipid barrier reconstruction, and deep inflammation inhibition," exhibiting a significant synergistic effect in increasing skin hydration.

[0028] (4) Micromicelle carriers and self-solubilizing and permeation-enhancing technology By utilizing a co-fermentation carrier, essential oils are contained in the form of tiny lipid micelles. This structure not only solves the common oily feeling of essential oils but also enhances transdermal absorption by mimicking the properties of biological membranes. While achieving self-solubilization of essential oils, this micromicelle carrier significantly shortens the time for active ingredients to penetrate the barrier, completely resolving the conflict between highly active components and the poor permeability of traditional matrices. Attached Figure Description

[0029] Appendix Figure 1 This is a laser irradiation image of sample number 1 in Embodiment 6 of the present invention. Detailed Implementation

[0030] To better understand the present invention, the present invention will be further described below with reference to specific serial numbers. The terminology used in the serial numbers is for describing specific embodiments and does not constitute a limitation on the scope of protection of the present invention.

[0031] In the specific implementation methods, unless otherwise specified, the experimental methods used are all conventional methods, and the materials and reagents used are all commercially available unless otherwise specified.

[0032] Unless otherwise specified, percentages, % and so on in the specific implementation method are assumed to be mass percentages.

[0033] The raw materials used in this invention are as follows: Aspergillus oryzae.

[0034] Bacillus subtilis.

[0035] Golden Chamomile Extract, INCI name: Golden Chamomile (CHRYSANTHELLUM INDICUM) Extract.

[0036] Ligusticum chuanxiong oil, INCI name: Ligusticum chuanxiong oil.

[0037] Frankincense oil, INCI name: Boswellia Carterii oil.

[0038] Citrus grandis essential oil is an essential oil component extracted from the peel of Citrus grandis.

[0039] Soybeans refer to mature, peeled seeds of GLYCINE MAX, and the soybeans used in this invention have been pre-degreased.

[0040] Kudzu root refers to the root of Pueraria Lobata.

[0041] Example 1: Preparation of a high-proportion soybean-kudzu co-fermentation product, comprising the following steps: S1, take defatted soybeans and kudzu root in a mass ratio of 3:1; dry them, pulverize them through a 40-mesh sieve, and then mix them; add deionized water in a material-to-liquid ratio of 1:8; 0.2% (w / w) of potassium dihydrogen phosphate and 0.1% (w / w) of magnesium sulfate heptahydrate were added based on the total liquid mass; the initial pH was adjusted to 5.5, and the fermentation substrate was obtained after high temperature and high pressure sterilization. S2, take Aspergillus oryzae strain, inoculate it on PDA slant, and culture it at 30℃ for 3 days until the slant is covered with a large number of bluish-brown or yellowish-green conidia.

[0042] The induction medium should be prepared according to the following composition: 1% glucose, 1% peptone, 2% soybean lecithin, 0.3% potassium dihydrogen phosphate, 0.15% magnesium sulfate heptahydrate.

[0043] Aspergillus oryzae conidia were inoculated into an induction medium and cultured until the logarithmic growth phase, and then Aspergillus oryzae seed culture was prepared.

[0044] S3, inoculate the fermentation substrate with 5% (v / v) Aspergillus oryzae seed culture; ferment according to the following parameters; Temperature 30±1℃, aeration rate 1.0-1.2 VVM; fermentation time at least 60h; The pH value was measured every 6 hours. The pH value of the fermentation broth dropped to its lowest point before 60 hours and then rebounded. Fermentation is considered complete when the pH value is ≥4.3 after 60 hours.

[0045] S4, after fermentation, heat at 85℃ for 10 min to inactivate; centrifuge the fermentation broth at 4000 rpm for 15 min; collect the supernatant; Add 3 times the volume of ethanol-water (75% concentration) to the supernatant and reflux at 45℃ twice, 1.5 h each time; filter again and collect the filtrate to obtain the ethanol extract; The alcohol extract was passed through an ultrafiltration membrane with a molecular weight cutoff of 10,000 Da, and the permeate was collected. The permeate was concentrated under reduced pressure at 50°C until the solid content was 30%, yielding a high-proportion soybean-kudzu co-fermentation product.

[0046] Example 2: Preparation of a low-ratio soybean-kudzu co-fermentation product, comprising the following steps: S1, take defatted soybeans and kudzu root in a mass ratio of 5:1; dry them, pulverize them through a 40-mesh sieve, and then mix them; add deionized water in a material-to-liquid ratio of 1:12; 0.2% (w / w) of potassium dihydrogen phosphate and 0.1% (w / w) of magnesium sulfate heptahydrate were added based on the total liquid mass; the initial pH was adjusted to 6.0, and the fermentation substrate was obtained after high temperature and high pressure sterilization. S2, take Aspergillus oryzae strain, inoculate it on PDA slant, and culture it at 30℃ for 3 days until the slant is covered with a large number of bluish-brown or yellowish-green conidia.

[0047] The induction medium should be prepared according to the following composition: 2% glucose, 1% peptone, 1% soy lecithin, 0.3% potassium dihydrogen phosphate, 0.15% magnesium sulfate heptahydrate.

[0048] Aspergillus oryzae conidia were inoculated into an induction medium and cultured until the logarithmic growth phase, and then Aspergillus oryzae seed culture was prepared.

[0049] S3, inoculate the fermentation substrate with 5% (v / v) Aspergillus oryzae seed culture; ferment according to the following parameters; Temperature 30±1℃, aeration rate 1.2-1.5 VVM; fermentation time at least 60 h; The pH value was measured every 6 hours. The pH value of the fermentation broth dropped to its lowest point before 60 hours and then rebounded. Fermentation is considered complete when the pH value is ≥4.3 after 60 hours.

[0050] S4, after fermentation, heat at 85℃ for 10 min to inactivate; centrifuge the fermentation broth at 4000 rpm for 15 min; collect the supernatant; Add 2 times the volume of ethanol-water (95% concentration) to the supernatant and reflux at 45℃ twice, 1.5 h each time; filter again and collect the filtrate to obtain the ethanol extract; The alcohol extract was passed through an ultrafiltration membrane with a molecular weight cutoff of 10,000 Da, and the permeate was collected. The permeate was concentrated under reduced pressure at 50°C until the solid content was 30%, yielding a low-proportion soybean-kudzu co-fermentation product.

[0051] Example 3: Preparation of non-induced soybean-kudzu co-fermentation product The difference from Example 1 is that the induction culture medium ratio in step S2 is as follows: 2% glucose, 1% peptone, 1% glycerol, 0.3% potassium dihydrogen phosphate, 0.15% magnesium sulfate heptahydrate.

[0052] Example 4: Preparation of a mixed soybean and kudzu root ferment, comprising the following steps: The difference from Example 1 is that in step S1, defatted soybeans and kudzu root are dried, pulverized, and then not mixed; defatted soybeans and kudzu root are fermented and refined separately according to the process and technical parameters of Example 1; after obtaining separate soybean fermentation products and kudzu root fermentation products with a solid content of 30%, they are mixed together.

[0053] Example 5: Preparation of Bacillus soy-kudzu co-fermentation product, including the following steps: S1, take defatted soybeans and kudzu root in a mass ratio of 3:1; dry them, pulverize them through a 40-mesh sieve, and then mix them; add deionized water in a material-to-liquid ratio of 1:8; 0.2% (w / w) of potassium dihydrogen phosphate and 0.1% (w / w) of magnesium sulfate heptahydrate were added based on the total liquid mass; the initial pH was adjusted to 7.0, and the fermentation substrate was obtained after high temperature and high pressure sterilization. S2, take Bacillus subtilis strain, inoculate it on LB solid slant medium, and incubate at 37℃ for 1 day.

[0054] The induction medium should be prepared according to the following composition: 2% soybean lecithin, 1% peptone, 0.5% yeast powder, 1% sodium chloride, 0.3% potassium dihydrogen phosphate, 0.15% magnesium sulfate heptahydrate.

[0055] Bacillus subtilis was inoculated from slant culture medium into induction medium and cultured with shaking at 37°C and 200 rpm until the logarithmic growth phase to prepare Bacillus subtilis seed culture.

[0056] S3, inoculate the fermentation substrate with 5% (v / v) Bacillus subtilis seed culture; ferment according to the following parameters; Temperature 37±1℃, aeration rate 1.2-1.5VVM; stirring speed 200rpm; fermentation time at least 36h; The pH value was measured every 6 hours. The pH value of the fermentation broth dropped to its lowest point before 60 hours and then rebounded. Fermentation is considered complete when the pH value is in the range of 6.5-7.0 after 60 hours.

[0057] S4, after fermentation, heat at 100℃ for 15 min to inactivate; centrifuge the fermentation broth at 4000 rpm for 15 min; take the supernatant; filter the supernatant through a 0.22 μM polyethersulfone PES microfiltration membrane and collect the microfiltration permeate; Add 3 times the volume of ethanol-water (75% concentration) to the microfiltration permeate and reflux at 45°C twice, 1.5 h each time; filter again and collect the filtrate to obtain the ethanol extract; The alcohol extract was passed through an ultrafiltration membrane with a molecular weight cutoff of 10,000 Da, and the permeate was collected. The permeate was concentrated under reduced pressure at 50°C until the solid content was 30%, yielding Bacillus soy-kudzu co-fermentation product.

[0058] Test Example 1: Determination of Lysophospholipid (LPC / LPE) Content The lysophospholipid (LPC / LPE) content was determined for the above examples to illustrate the differences between various fermentation methods. The specific methods are as follows: (1) Instruments and reagents Instruments: High performance liquid chromatograph (equipped with ELSD detector), ultrasonic cleaner, analytical balance (0.1 mg).

[0059] Reagents: n-hexane, isopropanol, methanol, glacial acetic acid (all chromatographic grade); ultrapure water.

[0060] Standards: Lysophosphatidylcholine (LPC) standard, lysophosphatidylethanolamine (LPE) standard.

[0061] (2) Preparation of standard solutions 1. Accurately weigh 10 mg each of LPC and LPE standards and place them in a 10 mL volumetric flask.

[0062] 2. Dissolve and dilute with isopropanol-n-hexane (1:1, v / v) to prepare a mixed standard stock solution of 1.0 mg / mL.

[0063] 3. Dilute the stock solution with the mobile phase to prepare a series of standard working solutions.

[0064] (3) Sample pretreatment Since the sample is an oily paste with a solid content ≥30%, lipid extraction is required. 1. Sampling: Accurately weigh 1.0g of fermentation product.

[0065] 2. Extraction: Add 10 mL of chloroform-methanol (2:1, v / v) mixed solvent and sonicate for 20 min to allow the lipids to fully enter the solvent layer.

[0066] 3. Remove impurities: Centrifuge (8000 rpm, 10 min) and collect the supernatant.

[0067] 4. Resolution: Dry the supernatant under nitrogen at 40°C, and redissolve the residue with 2 mL of mobile phase A.

[0068] 5. Filtration: Filtered through a 0.22μm organic filter membrane, ready for testing.

[0069] (4) Chromatographic conditions Chromatographic column: Silica column (250mm×4.6mm, 5μm).

[0070] Mobile phase: Gradient elution is used to separate different types of phospholipids.

[0071] Solution A: n-hexane-isopropanol.

[0072] Solution B: Isopropanol-Water-Glacial Acetic Acid-Triethylamine.

[0073] Flow rate: 1.0 mL / min.

[0074] Column temperature: 35℃.

[0075] ELSD parameters: drift tube temperature 60-80℃, carrier gas flow rate 2.0-2.5L / min.

[0076] (5) Content Calculation 1. Establishing the curve: Since the response of the ELSD detector is usually exponential, it is necessary to take the logarithm of the peak area (A) and concentration (C) and perform linear regression: logA=a·logC+b.

[0077] 2. Result conversion: Substitute the sample peak area into the regression equation to calculate the concentrations of LPC and LPE, and then convert them into the mass percentages in the original sample.

[0078] The results are shown in Table 1 below: Table 1 The test results showed that the total content of lysophospholipids (LPC+LPE) in Examples 1 and 2 was significantly higher than that in other control groups. This demonstrates that the soybean lecithin introduced in step S2, combined with the substrate ratio in step S1, can effectively activate the expression of Aspergillus oryzae phospholipase A, achieving efficient in-situ conversion of endogenous biosurfactants.

[0079] Comparing Examples 1 / 2 with Example 3, it was found that the lack of specific substrate induction led to a significant decrease in lysophospholipids, confirming that "pre-induction" is a key switch for initiating metabolic pathways. Comparing Examples 1 and 4 (physical compound group), it was observed that even with the same substrate, the lysophospholipids produced by the co-fermentation process were nearly 85% higher than those produced by separate fermentation, indicating a significant biochemical synergistic effect between soybeans and kudzu in the same fermentation system.

[0080] Furthermore, the extremely low detection level in Example 5 demonstrates that Bacillus subtilis is not suitable for the co-fermentation system of this invention.

[0081] Example 6: Preparation of oil-soluble plant complex, comprising the following steps: S1, Weigh each component according to Table 2, preheat component A to 40-55℃ and keep stirring at 200 rpm; S2, mix component B and add it to component A while stirring to obtain a coarse mixture; S3, homogenize the crude mixture at 4000 rpm for 8 min; then cool to room temperature to obtain the oil-soluble plant complex.

[0082] Table 2 Test Example 2: Testing of Oil-Soluble Plant Complexes (1) Performance of nano-dispersed systems Dilute sample number 1 in Table 2 20 times with water and place it in a beaker. In a dark room, illuminate the sample horizontally with a red laser pointer, as shown... Figure 1 As shown, the red laser forms a continuous, clear, and bright optical path in the liquid, with a stable and uniform colloidal morphology in the background; the system marked with serial number 1 contains a large number of nanoscale micelles, and a self-emulsifying system has been successfully established.

[0083] (2) Hyaluronidase inhibition experiment This experiment, a biochemical titration method, was used to demonstrate the protective ability of the complex on the dermal matrix.

[0084] 1. Preparation of sample stock solution (1% active concentration) Sampling: Accurately weigh 1.00g of each sample number in Example 6.

[0085] Solubilization: Add 5 mL of phosphate buffer (PBS, pH 5.35) containing 0.1% Tween-80 and sonicate for 10 min to fully disperse the essential oil nanomicelles.

[0086] Volume adjustment: Add PBS to a final volume of 100 mL, shake well, and obtain a 1% (w / v) experimental working solution.

[0087] 2. Experimental Procedure Reaction system: Add 0.25 mL of hyaluronidase solution (0.1%) to a test tube, add 0.5 mL of 1% sample working solution, and incubate at 37°C for 20 min.

[0088] Activation: Add 0.1 mL of calcium chloride activator (2.5 mmol / L) and continue incubation for 20 min.

[0089] Substrate reaction: Add 0.5 mL of sodium hyaluronate solution (0.5 mg / mL) and react at 37°C for 30 min.

[0090] Color development: The reaction was terminated by adding 0.1 mL of sodium hydroxide solution (0.4 mol / L), followed by the addition of the colorimetric reagent (p-dimethylaminobenzaldehyde), and the absorbance A was measured at 585 nm. S .

[0091] Control group: A0 was measured using PBS instead of the sample; blank A was measured using PBS instead of the enzyme solution. b .

[0092] 3. Calculation formula The results are shown in Table 3.

[0093] (3): IL-6 inflammatory factor inhibition assay (ELISA method) This experiment, based on a cell model, was used to demonstrate the inhibitory effect of the complex on deep inflammation.

[0094] 1. Cell-level sample preparation Dissolution: Weigh 0.1g of each sample number in Example 6 and add 10mL of serum-free culture medium (containing 0.5% DMSO) and sonicate to aid dissolution.

[0095] Sterilization: Filter through a 0.22μm sterile filter membrane to obtain a 1% sterile stock solution.

[0096] 2. Experimental Procedure Cell seeding: RAW264.7 (macrophages) were seeded at a rate of 1 × 10⁻⁶. 5 Inoculate one well per well into a 96-well plate and incubate for 24 hours.

[0097] Induction and drug administration: Model group: Added culture medium containing LPS (lipopolysaccharide, 1 μg / mL).

[0098] Experimental group: Add LPS (1 μg / mL) + 1% sample diluent.

[0099] Control group: only normal culture medium was added.

[0100] Incubation: Continue incubation for 24 hours.

[0101] Supernatant collection: Collect the culture supernatant from each well and centrifuge at 4°C and 3000 rpm for 5 min to remove cell debris.

[0102] ELISA test: Add the coating antibody, standard / sample, biotinylated antibody, HRP marker, and substrate TMB in sequence according to the kit instructions.

[0103] After adding the stop solution, the OD value was read at 450 nm using an ELISA reader.

[0104] 3. Calculation formula The results are shown in Table 3.

[0105] Table 3 This test case aims to evaluate the synergistic effect of different fermentation carriers (component A) on the bioactivity of the essential oil matrix (component B). Experimental data show that the plant complexes (numbers 1, 2, and 9) prepared using Examples 1 and 2 of this invention exhibit excellent barrier repair and anti-inflammatory properties at a 1% experimental concentration. Specifically, number 1 showed a hyaluronidase inhibition rate of 88.4% and an IL-6 inhibition rate of 82.6%, significantly superior to the groups using no-inducing carrier (number 3), physically compounded carrier (number 4), and Bacillus carrier (number 5).

[0106] Compared to No. 4, under the premise of completely identical essential oil components, No. 1 showed an increase of approximately 43% in hyaluronidase inhibition rate and approximately 55% in IL-6 inhibition rate. This quantitatively confirms that the endogenous lysophospholipid matrix produced by co-fermentation not only has excellent self-emulsifying carrying capacity but also serves as a highly efficient penetration enhancer, significantly improving the bioavailability of active ingredients such as Ligusticum chuanxiong oil and chamomile extract; demonstrating the synergistic penetration-enhancing effect of the carrier.

[0107] All indicators of No. 1 (essential oil blend) were higher than those of the single essential oil groups (No. 6, 7, 8), demonstrating the multi-target synergistic effect of Ligusticum chuanxiong oil in improving microcirculation, chamomile in inhibiting inflammatory factors, and frankincense in repairing the matrix. It can build a complete skin chemical defense barrier at a concentration of 1%, showing the synergistic effect of the essential oil matrix.

[0108] The extremely low activity of item 5 (inhibition rate below 35%) further confirms the incompatibility between Bacillus metabolites and the essential oil system of this invention.

[0109] The above detailed description is a specific description of one of the feasible embodiments of the present invention. This embodiment is not intended to limit the patent scope of the present invention. All equivalent implementations or modifications that do not depart from the present invention should be included within the scope of the technical solution of the present invention.

Claims

1. A plant composition, characterized in that, The following components are included in parts by mass: Golden Chamomile Extract 5-12 Ligusticum chuanxiong oil 2-5 Frankincense oil 3-8 1-2g of Citrus reticulata essential oil Soybean-kudzu co-fermentation product: 70-90; The soybean-kudzu co-fermented product is obtained by mixing soybeans and kudzu at a mass ratio of 3-5:1, inoculating with Aspergillus oryzae for fermentation, and then extracting with a medium-to-low polar solvent.

2. The plant composition according to claim 1, characterized in that, The soybean-kudzu co-fermentation product was extracted with a medium-low polarity solvent, then purified by ultrafiltration with a molecular weight cutoff of 10,000 Da. The permeate was collected and concentrated under reduced pressure until the solid content was ≥30%.

3. The plant composition according to claim 1, characterized in that, The golden chamomile extract, chuanxiong oil, frankincense oil, and tangerine peel essential oil are alcohol-soluble oily components obtained from their respective plants through physical separation or solvent extraction.

4. The plant composition according to claim 1, characterized in that, The preparation method of the soybean-kudzu co-fermentation product includes the following steps: S1. Take defatted soybeans and kudzu root, dry them, grind them into powder and mix them; add water at a material-to-liquid ratio of 1:8-12; then add at least 0.2% (w / w) potassium dihydrogen phosphate and at least 0.1% (w / w) magnesium sulfate heptahydrate based on the total mass of liquid; adjust the initial pH to between 5.5 and 6.0, sterilize and obtain the fermentation substrate; S2, Aspergillus oryzae is inoculated into an induction medium containing at least 1% (w / w) soybean lecithin for induction culture and seed culture is prepared; S3, inoculate the fermentation substrate with Aspergillus oryzae seed liquid; Ferment according to the following parameters; Temperature 30±1℃, aeration rate 1.0-1.5VVM; fermentation time at least 60h; S4, after fermentation, is inactivated and centrifuged to obtain the supernatant; ethanol aqueous solution is added to the supernatant for extraction, and the alcohol extract is obtained; S5, the alcohol extract is passed through an ultrafiltration membrane with a molecular weight cutoff of 10000 Da, and the permeate is collected; the permeate is concentrated under reduced pressure until the solid content is ≥30%.

5. The plant composition according to claim 4, characterized in that, In step S3, the inoculation amount of Aspergillus oryzae seed liquid is at least 5% of the substrate volume.

6. The plant composition according to claim 4, characterized in that, In step S4, the condition for determining the completion of fermentation is: the pH value of the fermentation broth drops to its lowest point and then rises again, with the rise being at least 0.2 units, and the final pH value is ≥4.

3.

7. The plant composition according to claim 4, characterized in that, In step S4, when adding an ethanol-water solution to the supernatant for extraction, the amount of ethanol-water solution added is 2-3 times the volume of the supernatant; the concentration of the ethanol-water solution is 75-95%.

8. A method for preparing the plant composition according to any one of claims 1-7, characterized in that, Includes the following steps, S1. Preheat the soybean-kudzu co-fermentation product to 40-55℃ and keep stirring. S2, mix the remaining components and add them to the soybean-kudzu co-fermentation product while stirring to obtain a coarse mixture; S3, homogenize the crude mixture to obtain the plant composition.

9. The use of the plant composition according to any one of claims 1-8 in the preparation of cosmetics.

10. The application according to claim 9, characterized in that, The plant composition is present in ≥1% of the cosmetic.

Images