An anti-aging and anti-glycation Eucommia ulmoides leaf extract composition, its preparation method, and its application in oral beauty products.

Abstract

This invention provides an anti-aging and anti-glycation Eucommia ulmoides leaf extract composition, its preparation method, and its application in oral cosmetic products, belonging to the field of food technology. The composition of this invention consists of Eucommia ulmoides leaf powder, Cistanche deserticola powder, Cornus officinalis powder, and Gastrodia elata powder, prepared using deep eutectic solvent extraction, microbial fermentation, subcritical water extraction, and ethanol reflux extraction processes, respectively, significantly improving the extraction efficiency and bioavailability of the active ingredients. This composition, combined with type III collagen peptides, vitamin C, and other excipients, is formulated into a cosmetic oral liquid, possessing multiple cosmetic effects such as anti-glycation, anti-oxidation, promoting collagen synthesis, and strengthening the skin barrier. Through in vitro, animal, and clinical trials, the cosmetic oral liquid of this invention has been verified to have anti-glycation, anti-aging, epidermal layer thickness-increasing, skin barrier health-improving, and skin firmness-enhancing effects.

Description

Technical Field

[0001] This invention relates to the field of food technology, specifically to an anti-aging and anti-glycation Eucommia ulmoides leaf extract composition, its preparation method, and its application in oral beauty products. Background Technology

[0002] With the development of society and the economy and the improvement of people's living standards, people's pursuit of beauty is growing. Beauty and skincare have shifted from simple external care to a comprehensive beauty concept that considers both internal and external aspects. Against this backdrop, oral beauty products, as an effective way to improve skin condition and delay aging from the inside out, have gained widespread market favor.

[0003] Currently, oral beauty products on the market can be mainly divided into the following categories: 1) Single-ingredient supplements: such as supplements for collagen, vitamin C, vitamin E, hyaluronic acid, etc. These products have relatively simple functions and are difficult to address the complex mechanisms of skin aging. Furthermore, the bioavailability of some synthetically derived ingredients is limited. 2) Compound products containing chemically synthesized substances: These are products that combine multiple chemically synthesized nutrients. Although the ingredients are clearly defined, long-term use may lead to potential side effects. 3) Plant extracts (phytoextracts) have become a research hotspot in the field of oral beauty due to their naturalness, safety, and rich content of various bioactive components (such as polyphenols, flavonoids, polysaccharides, saponins, etc.). However, traditional extraction methods (such as water extraction and alcohol extraction) are relatively simple, with low extraction rates and selectivity for specific active ingredients. Moreover, during the extraction process, high temperatures and prolonged processing can easily deactivate heat-sensitive components, affecting the efficacy of the final product. Furthermore, when applying plant extract compositions to oral liquids, multiple technical bottlenecks are faced, including issues with taste (bitterness, grassy taste), stability (precipitation, stratification, degradation of active ingredients), and preservation. Poor taste and appearance can seriously affect consumer compliance.

[0004] Therefore, there is an urgent need in this field to develop a new and efficient method for preparing plant extract compositions that can retain and enrich active ingredients with synergistic cosmetic effects to the maximum extent, and at the same time successfully apply the composition to oral liquid formulations with good taste, good stability and high bioavailability. Summary of the Invention

[0005] The purpose of this invention is to provide an anti-aging and anti-glycation Eucommia ulmoides leaf extract composition, its preparation method, and its application in oral beauty products. The Eucommia ulmoides leaf extract composition provided by this invention has excellent anti-glycation, antioxidant, and whitening effects.

[0006] To achieve the above-mentioned objectives, the present invention provides the following technical solution:

[0007] This invention provides a Eucommia ulmoides leaf extract composition comprising the following raw materials in parts by weight: 40-60 parts Eucommia ulmoides leaf powder, 25-35 parts Cistanche deserticola powder, 15-25 parts Cornus officinalis powder, and 8-12 parts Gastrodia elata powder.

[0008] Preferably, the preparation method of the Eucommia ulmoides leaf powder includes: pulverizing Eucommia ulmoides leaves, adding deep eutectic solvent extract, ultrasonic extraction, filtration, purification, concentration, and drying to obtain Eucommia ulmoides leaf powder.

[0009] More preferably, the molar ratio of choline chloride to glycerol in the deep eutectic solvent extract is 1:1.5-2.5.

[0010] Preferably, the preparation method of the Cistanche deserticola powder includes: pulverizing dried Cistanche deserticola slices, mixing with water, inoculating with compound microorganisms for anaerobic fermentation, filtering, and drying to obtain Cistanche deserticola powder.

[0011] More preferably, the composite microorganism consists of Bifidobacterium lactis BL-99 and Lactobacillus delbrueckii subsp. bulgaricus Ouya-D-L5.

[0012] Preferably, the preparation method of the Cornus officinalis powder includes: pressing fresh Cornus officinalis to obtain Cornus officinalis juice and Cornus officinalis residue; mixing the Cornus officinalis residue with deionized water and extracting it with subcritical water to obtain a fruit residue extract; mixing the Cornus officinalis juice and the fruit residue extract, concentrating and drying them to obtain Cornus officinalis powder.

[0013] Preferably, the preparation method of the gastrodia powder includes: crushing gastrodia slices and mixing them with an ethanol solution, reflux extraction, filtering, concentrating and drying the filtrate to obtain gastrodia powder.

[0014] The present invention also provides a method for preparing the above-mentioned Eucommia ulmoides leaf extract composition, comprising: mixing Eucommia ulmoides leaf powder, Cistanche deserticola powder, Cornus officinalis powder and Gastrodia elata powder to obtain the Eucommia ulmoides leaf extract composition.

[0015] The present invention also provides an application of the above-described Eucommia ulmoides leaf extract composition in the preparation of a cosmetic oral liquid.

[0016] The present invention also provides a cosmetic oral liquid containing the above-mentioned Eucommia ulmoides leaf extract composition, comprising: 40-60 parts of the Eucommia ulmoides leaf extract composition, 40-60 parts of type III collagen peptide powder, 25-35 parts of fructooligosaccharides, 20-30 parts of hydroxypropyl-β-cyclodextrin, 70-80 parts of maltodextrin, 0.5-2 parts of stevia, 1-5 parts of citric acid, 0.01-0.5 parts of vitamin C, and 1-3 parts of xanthan gum.

[0017] Compared with the prior art, the present invention has the following beneficial effects:

[0018] This invention discloses a Eucommia ulmoides leaf extract composition composed of Eucommia ulmoides leaf powder, Cistanche deserticola powder, Cornus officinalis powder, and Gastrodia elata powder. These components are prepared using deep eutectic solvent extraction, microbial fermentation, subcritical water extraction, and ethanol reflux extraction processes, respectively, significantly improving the extraction efficiency and bioavailability of the active ingredients. In vitro experiments verified the synergistic effect between the plant extracts of this invention (the whole Cistanche deserticola powder, Cornus officinalis powder, and Gastrodia elata powder and Eucommia ulmoides leaf powder), demonstrating excellent effects in anti-glycation, anti-oxidation, and whitening.

[0019] This invention relates to a cosmetic oral liquid formulated with Eucommia ulmoides leaf extract and excipients such as type III collagen peptides and vitamin C. This oral liquid possesses multiple cosmetic benefits, including anti-glycation, anti-oxidation, promotion of collagen synthesis, and enhancement of the skin barrier. Animal and clinical trials have verified that this cosmetic oral liquid exhibits anti-glycation, anti-aging, increases epidermal thickness, improves skin barrier health, and enhances skin firmness, thus solving problems such as low extraction rates and poor taste in traditional plant extract products. Detailed Implementation

[0020] This invention provides a Eucommia ulmoides leaf extract composition comprising the following raw materials in parts by weight: 40-60 parts Eucommia ulmoides leaf powder, 25-35 parts Cistanche deserticola powder, 15-25 parts Cornus officinalis powder, and 8-12 parts Gastrodia elata powder.

[0021] The preferred method for preparing Eucommia ulmoides leaf powder according to the present invention includes: pulverizing Eucommia ulmoides leaves, adding a deep eutectic solvent extract, ultrasonically extracting, filtering, purifying, concentrating, and drying to obtain Eucommia ulmoides leaf powder. More preferably, it includes: pulverizing Eucommia ulmoides leaves through a 50-100 mesh sieve, mixing with a deep eutectic solvent extract at a material-to-liquid ratio of 1:15-1:20 (g / mL), extracting 2-4 times at 400-500W, 20-40kHz, and 45-55℃, each time for 25-35 minutes, filtering, combining the filtrates, concentrating the filtrate to 1 / 2-1 / 4 of its volume, adsorbing with HPD-300 macroporous resin, and then eluting sequentially with 3-5 BV of deionized water and 4-6 BV of 50%-60% ethanol solution, collecting the ethanol eluent, concentrating the eluent to a relative density of 1.10-1.15, and freeze-drying under vacuum to a water content of 1wt%-4wt% to obtain Eucommia ulmoides leaf powder. The preferred method for preparing the deep eutectic solvent extract includes: mixing choline chloride and glycerol at a molar ratio of 1:1.5-2.5, stirring at 70-80℃ and 350-450rpm for 80-100min to obtain a deep eutectic solvent, mixing it with deionized water at a volume ratio of 2-4:5-8, and stirring at 250-350rpm for 10-20min to obtain the deep eutectic solvent extract.

[0022] This invention utilizes a green deep eutectic solvent combined with ultrasonic extraction and macroporous resin purification technology to produce refined Eucommia ulmoides leaf powder rich in highly active ingredients (genipin, flavonoids, polyphenols, etc.). In a cosmetic oral liquid, it exerts its cosmetic effects through a multi-target mechanism: In anti-glycation, high-purity genipin, as a highly efficient inhibitor of advanced glycation end products (AGEs), specifically captures active dicarbonyl compounds, blocks protein cross-linking reactions, and significantly reduces glycation levels in skin tissue; in anti-aging and improving skin firmness, genipin and flavonoids synergistically activate the collagen synthesis pathway in fibroblasts, significantly promoting the gene expression of type I and type III collagen and elastin, while effectively inhibiting matrix metalloproteinase activity, bidirectionally regulating the synthesis and degradation metabolism of the extracellular matrix, thereby enhancing skin elasticity, improving wrinkle depth, and achieving anti-aging effects. In terms of increasing epidermal thickness and barrier function, the active ingredients of Eucommia ulmoides leaves can promote the synthesis of filaggrin and inner lining protein, markers of keratinocyte differentiation, accelerate the complete construction of the lipid barrier of the stratum corneum, effectively enhance the epidermis's water retention capacity, and repair the physical barrier function. The polyphenolic components retained by this preparation process provide a basic guarantee for the above-mentioned skin improvement effects by scavenging free radicals and inhibiting oxidative stress.

[0023] The preferred method for preparing Cistanche deserticola powder according to the present invention includes: pulverizing dried Cistanche deserticola slices, mixing with water, inoculating with a compound microorganism for anaerobic fermentation, filtering, and drying to obtain Cistanche deserticola powder. More preferably, it includes: pulverizing dried Cistanche deserticola slices through a 30-80 mesh sieve to obtain Cistanche deserticola powder, mixing with a liquid fermentation medium at a ratio of 1:12 (g / mL), sterilizing, inoculating with a compound microorganism at 5‰-8‰ of the weight of the Cistanche deserticola powder, anaerobic fermenting at 35-38℃ and pH 5-6 for 36-72 hours, filtering, sterilizing, concentrating the filtrate to a relative density of 1.17-1.22, and freeze-drying under vacuum to a water content of 2wt%-5wt% to obtain Cistanche deserticola powder. The compound microorganisms preferably consist of Bifidobacterium lactis BL-99 and Lactobacillus delbrueckii subsp. bulgaricus Ouya-D-L5, and the preferred viability of Bifidobacterium lactis BL-99 in the compound microorganisms is (2-5)×10⁻⁶. 9 pcs / g, more preferably 3×10 9 The optimal viability of *Lactobacillus delbrueckii* subsp. bulgaricus Ouya-D-L5 is (3-8) × 10⁻⁶ cells / g. 9 5 × 10⁻⁶ cells / g, more preferably 5 × 10⁻⁶ cells / g. 9 per g.

[0024] This invention employs a specific composition to ferment *Cistanche deserticola*, which efficiently converts glycosides to increase the content of active ingredients (such as phenylethanoid glycosides, polysaccharides, and amino acids), produces beneficial metabolites, and significantly improves the inherent odor of *Cistanche deserticola*. The *Cistanche deserticola* powder of this invention possesses antioxidant activity, which can help scavenge free radicals in the body. Simultaneously, it regulates metabolism with the complex microbial fermentation products, helps inhibit non-enzymatic glycation reactions, and exerts anti-glycation and anti-aging effects. Amino acids provide raw materials for skin cell renewal, promote epidermal cell proliferation, and help increase the thickness of the epidermis. Polysaccharide components and metabolites can enhance the integrity of the stratum corneum, reduce moisture loss, and improve skin barrier health. As precursors for collagen synthesis, amino acids can help increase the collagen content in the skin, thereby helping to improve skin firmness and providing skin health support for the beauty oral liquid.

[0025] The preferred method for preparing Cornus officinalis powder according to the present invention includes: pressing fresh Cornus officinalis to obtain Cornus officinalis juice and Cornus officinalis residue; mixing the Cornus officinalis residue with deionized water and extracting it with subcritical water to obtain a fruit residue extract; mixing the Cornus officinalis juice and the fruit residue extract, concentrating and drying them to obtain Cornus officinalis powder. A more preferred method includes: pressing the pulp of Cornus officinalis to obtain juice and pomace; pumping the juice into a 200-300 Da nanofiltration membrane system and performing nanofiltration at 1.5-2.5 MPa and 35-45 °C, collecting the retentate liquid to obtain concentrated juice; crushing the pomace through a 10-20 mesh sieve, mixing it with deionized water at a ratio of 1:13-18 (g / mL) and adding it into a subcritical water reactor, purging with nitrogen to replace the air, extracting at 130-140 °C and 2.5-3.5 MPa for 20-30 min, filtering, mixing the filtrate with the concentrated juice, concentrating to a relative density of 1.15-1.20, and freeze-drying under vacuum to a water content of 2wt%-5wt% to obtain Cornus officinalis powder.

[0026] This invention utilizes a synergistic process of subcritical water extraction and nanofiltration membranes to significantly improve the bioavailability of active ingredients (iridosides, polysaccharides, etc.), exhibiting anti-glycation, anti-aging, and tight junction protein expression-promoting effects. Its anti-glycation effect stems from the potent inhibition of AGEs formation by iridoids, reducing the damage of advanced glycation end products (AGEs) to collagen; the anti-aging effect is achieved through free radical scavenging and MMP-1 expression inhibition.

[0027] The preferred method for preparing Gastrodia elata powder according to the present invention includes: pulverizing Gastrodia elata slices and mixing them with an ethanol solution, reflux extraction, filtering, concentrating and drying the filtrate to obtain Gastrodia elata powder. More preferably, the method includes: pulverizing Gastrodia elata slices through a 30-80 mesh sieve, mixing them with a 65%-75% ethanol solution at a ratio of 1:8-12 (g / mL), reflux extraction at 83-88℃ 1-3 times, 1-2 hours each time, filtering, combining the filtrates, concentrating to a relative density of 1.08-1.12, and freeze-drying under vacuum to a water content of 2wt%-5wt% to obtain Gastrodia elata powder.

[0028] This invention employs ethanol reflux extraction to extract Gastrodia elata, thereby improving the extraction efficiency of its active ingredients. The Gastrodia elata powder of this invention can effectively scavenge free radicals, inhibit lipid peroxidation, protect fibroblasts and keratinocytes, delay cell aging, and slow down the degradation of collagen and elastin. It also promotes the synthesis of barrier-related proteins such as filaggrin and inner lining protein in keratinocytes, and stimulates the production of ceramides, improving dryness and roughness, making the skin more hydrated and plump. Furthermore, it has a calming and anti-anxiety neuroregulatory effect, helping to alleviate skin problems exacerbated by mental stress.

[0029] The Eucommia ulmoides leaf extract composition provided by this invention, in which Eucommia ulmoides leaf powder, Cistanche deserticola powder, Cornus officinalis powder, and Gastrodia elata powder work together through a multi-target synergistic mechanism to exert a systemic anti-aging and beautifying effect. Regarding anti-glycation, geniposide in Eucommia ulmoides leaf powder and iridoid glycosides in Cornus officinalis powder jointly block non-enzymatic glycation reactions of proteins; fermentation products of Cistanche deserticola powder assist in inhibiting AGEs receptor pathways; and phenolic components in Gastrodia elata powder effectively remove active carbonyl groups that promote glycation. These multiple mechanisms work together to reduce collagen glycation damage. Regarding anti-aging and barrier repair, Eucommia ulmoides leaf powder and Cistanche deserticola powder synergistically promote the synthesis of type I and III collagen and inhibit matrix metalloproteinase activity, significantly improving dermal firmness; polysaccharides from Cornus officinalis powder and active ingredients from Gastrodia elata powder jointly promote keratinocyte differentiation, enhance filaggrin and ceramide synthesis, and synergistically construct a complete skin lipid barrier. Furthermore, the antioxidant activities of all four components form a combined defense system, effectively reducing the multiple damages to the skin caused by oxidative stress. This synergistic effect based on different targets enables the Eucommia ulmoides leaf extract composition to intervene in the skin aging process from multiple dimensions, including anti-glycation, collagen synthesis promotion, barrier function enhancement and anti-oxidation, achieving a more comprehensive and significant cosmetic improvement effect than a single ingredient.

[0030] The present invention also provides a method for preparing the above-mentioned Eucommia ulmoides leaf extract composition, comprising: mixing Eucommia ulmoides leaf powder, Cistanche deserticola powder, Cornus officinalis powder and Gastrodia elata powder to obtain the Eucommia ulmoides leaf extract composition.

[0031] The present invention also provides an application of the above-mentioned Eucommia ulmoides leaf extract composition in the preparation of a cosmetic oral liquid, wherein the cosmetic benefits preferably include anti-glycation, anti-aging, increasing epidermal thickness, improving skin barrier health, and enhancing skin firmness.

[0032] The present invention also provides a cosmetic oral liquid containing the above-mentioned Eucommia ulmoides leaf extract composition, comprising: 40-60 parts of the Eucommia ulmoides leaf extract composition, 40-60 parts of type III collagen peptide powder, 25-35 parts of fructooligosaccharides, 20-30 parts of hydroxypropyl-β-cyclodextrin, 70-80 parts of maltodextrin, 0.5-2 parts of stevia, 1-5 parts of citric acid, 0.01-0.5 parts of vitamin C, and 1-3 parts of xanthan gum.

[0033] The oral beauty liquid provided by this invention achieves comprehensive skin improvement through the combination of Eucommia ulmoides leaf extract and a scientific formula. The Eucommia ulmoides leaf extract, as the core active ingredient, exerts a systemic anti-aging effect through the multi-target synergistic action of Eucommia ulmoides leaf powder, Cistanche deserticola powder, Cornus officinalis powder, and Gastrodia elata powder, including anti-glycation, promotion of collagen synthesis, enhancement of barrier function, and antioxidant properties. Type III collagen peptide powder, as a direct raw material for collagen synthesis, synergizes with the collagen-promoting active ingredients in the Eucommia ulmoides leaf extract to jointly improve skin firmness and elasticity. Vitamin C is an essential cofactor for collagen synthesis and, as a powerful antioxidant, works synergistically with vitamin E to scavenge free radicals and enhance the overall antioxidant network. Fructooligosaccharides function as both a prebiotic and a natural sweetener, indirectly supporting skin health by regulating gut microbiota. Hydroxypropyl-β-cyclodextrin significantly improves its solubility and bioavailability through inclusion complexation, while effectively masking unpleasant flavors. Maltodextrin serves as a carrier and filler to ensure product stability. Xanthan gum acts as a stabilizer to prevent component precipitation and ensure system homogeneity. Stevia and citric acid work together to adjust the taste, creating a pleasantly sweet and sour flavor. The scientifically formulated ingredients combine to create a beauty oral liquid with clear efficacy, excellent stability, and a pleasant taste.

[0034] In this invention, unless otherwise specified, all raw material components are commercially available products well known to those skilled in the art.

[0035] The technical solutions of this invention will be clearly and completely described below with reference to the embodiments thereof. Obviously, the described embodiments are only a part of the embodiments of this invention, and not all of them. All other embodiments obtained by those skilled in the art based on the embodiments of this invention without creative effort are within the scope of protection of this invention.

[0036] Unless otherwise specified, the following embodiments are all conventional methods.

[0037] Unless otherwise specified, all materials and reagents used in the following examples are commercially available.

[0038] The liquid fermentation medium for the examples and comparative examples was prepared by adding 10g beef extract powder, 5g peptone, 3g yeast extract powder, 5g glucose and 1g soluble starch to 1000mL of distilled water.

[0039] Bifidobacterium lactis BL-99 was purchased from the China General Microbiological Culture Collection Center (CGMCC No. 15650), and is disclosed in patent CN110964657A; Lactobacillus delbrueckii subsp. bulgaricus Ouya-D-L5 was purchased from the China General Microbiological Culture Collection Center (CGMCC No. 17959), and is disclosed in patent CN110484477A; Bifidobacterium longum strain LTBL16 was purchased from the China Center for Type Culture Collection (CCTCC No. M 2019028), and is disclosed in patent CN110452829A; Lactobacillus plantarum ZF603 was purchased from the Guangdong Provincial Microbiological Culture Collection Center (GDMCC No. 61734), and is disclosed in patent CN113717879A.

[0040] Type III collagen peptide powder is a solid beverage made from type III collagen peptide powder (composed of type III collagen peptide, collagen peptide, and elastin peptide), purchased from Lianyi International Biotechnology (Zhejiang) Co., Ltd. Fructooligosaccharides were purchased from Shanxi Xize Biotechnology Co., Ltd.

[0041] DPPH free radical scavenging rate assay kit, catalog number ml095224; total antioxidant capacity (ABTS method) assay kit, catalog number ml092653; mouse superoxide dismutase (SOD) ELISA assay kit, catalog number ml001998; glutathione peroxidase (GSH-PX) assay kit / colorimetric method, catalog number ml092941; mouse hydroxyproline (Hyp) assay kit, catalog number ml631122; malondialdehyde (MDA) assay kit (TBA colorimetric method), catalog number ml094963, were purchased from Shanghai Enzyme-Link Biotechnology Co., Ltd.

[0042] Example 1

[0043] Preparation of Eucommia ulmoides leaf extract composition

[0044] (1) Preparation of Eucommia ulmoides leaf powder

[0045] Choline chloride and glycerol were mixed at a molar ratio of 1:2 and stirred at 75°C and 400 rpm for 90 min to obtain a deep eutectic solvent. This solvent was then mixed with deionized water at a volume ratio of 3:7 and stirred at 300 rpm for 15 min to obtain a deep eutectic solvent extract.

[0046] Eucommia ulmoides leaves were pulverized and passed through an 80-mesh sieve. They were mixed with deep eutectic solvent extract at a material-to-liquid ratio of 1:18 (g / mL) and extracted three times at 450W, 30kHz, and 50℃ for 30 minutes each time. The extracts were filtered, and the filtrates were combined. The filtrates were concentrated to 1 / 3 of their volume and then adsorbed onto HPD-300 macroporous resin. The extracts were then eluted sequentially with 4 BV of deionized water and 5 BV of 55% ethanol solution. The ethanol eluent was collected and concentrated to a relative density of 1.13. The eluent was then freeze-dried under vacuum to a water content of 2 wt% to obtain Eucommia ulmoides leaf powder.

[0047] (2) Preparation of Cistanche deserticola powder

[0048] The compound microbial community consists of Bifidobacterium lactis BL-99 and Lactobacillus delbrueckii subsp. bulgaricus Ouya-D-L5. The viability of Bifidobacterium lactis BL-99 in the compound microbial community is 3 × 10⁻⁶. 9 The bacterial count of *Lactobacillus delbrueckii* subsp. bulgaricus Ouya-D-L5 was 5 × 10⁻⁶ cells / g. 9 bacteria / g;

[0049] The dried desert Cistanche deserticola was sliced, pulverized, and passed through a 50-mesh sieve to obtain Cistanche deserticola powder. The powder was mixed with liquid fermentation medium at a ratio of 1:12 (g / mL), sterilized, and inoculated with 7‰ of the weight of the Cistanche deserticola powder as a compound microorganism. The mixture was then anaerobic fermented at 37℃ and pH 5.5 for 54 hours. After filtration and sterilization, the filtrate was concentrated to a relative density of 1.2 and then freeze-dried under vacuum to a water content of 3wt% to obtain Cistanche deserticola powder.

[0050] (3) Preparation of Cornus officinalis powder

[0051] The pulp of Cornus officinalis fruit was pressed to obtain juice and pomace. The juice was pumped into a 250 Da nanofiltration membrane system and nanofiltration was performed at 2 MPa and 40 °C. The retentate was concentrated to 1 / 4 of its volume to obtain concentrated juice. The pomace was crushed and passed through a 15-mesh sieve. It was mixed with deionized water at a ratio of 1:15 (g / mL) and added to a subcritical water reactor. After purging with nitrogen to replace the air, it was extracted at 135 °C and 2 MPa for 25 min. After filtration, the filtrate was mixed with the concentrated juice and concentrated to a relative density of 1.18. It was then freeze-dried under vacuum to a water content of 3 wt% to obtain Cornus officinalis powder.

[0052] (4) Preparation of Gastrodia elata powder

[0053] Gastrodia elata slices were pulverized and passed through a 50-mesh sieve. They were mixed with a 70% ethanol solution at a ratio of 1:10 (g / mL) and extracted twice by reflux at 80℃ for 1.5 hours each time. The mixtures were filtered, and the filtrates were combined and concentrated to a relative density of 1.10. The mixtures were then freeze-dried under vacuum to a water content of 4 wt% to obtain Gastrodia elata powder.

[0054] (5) Preparation of Eucommia ulmoides leaf extract composition

[0055] Eucommia ulmoides leaf powder, Cistanche deserticola powder, Cornus officinalis powder and Gastrodia elata powder were mixed in a weight ratio of 50:30:20:10 to obtain an Eucommia ulmoides leaf extract composition.

[0056] Example 2

[0057] Preparation of Eucommia ulmoides leaf extract composition

[0058] (1) Preparation of Eucommia ulmoides leaf powder

[0059] Choline chloride and glycerol were mixed at a molar ratio of 1:1.5 and stirred at 70°C and 350 rpm for 100 min to obtain a deep eutectic solvent. This solvent was then mixed with deionized water at a volume ratio of 2:5 and stirred at 250 rpm for 20 min to obtain a deep eutectic solvent extract.

[0060] Eucommia ulmoides leaves were pulverized and passed through a 50-mesh sieve. They were mixed with deep eutectic solvent extract at a material-to-liquid ratio of 1:15 (g / mL) and extracted four times at 400W, 20kHz, and 45℃ for 25 minutes each time. The extracts were filtered, and the filtrates were combined. The filtrates were concentrated to half their volume and then adsorbed onto HPD-300 macroporous resin. The extracts were then eluted sequentially with 3 BV of deionized water and 4 BV of 50% ethanol solution. The ethanol eluent was collected and concentrated to a relative density of 1.10. The eluent was then freeze-dried under vacuum to a water content of 2 wt% to obtain Eucommia ulmoides leaf powder.

[0061] (2) Preparation of Cistanche deserticola powder

[0062] The compound microbial community consists of Bifidobacterium lactis BL-99 and Lactobacillus delbrueckii subsp. bulgaricus Ouya-D-L5. The viability of Bifidobacterium lactis BL-99 in the compound microbial community is 2 × 10⁻⁶. 9 The bacterial count of *Lactobacillus delbrueckii* subsp. bulgaricus Ouya-D-L5 was 3 × 10⁻⁶ cells / g. 9 pcs / g;

[0063] The dried desert Cistanche deserticola was sliced, pulverized, and passed through a 30-mesh sieve to obtain Cistanche deserticola powder. The powder was mixed with liquid fermentation medium at a ratio of 1:10 (g / mL), sterilized, and inoculated with 8‰ of the weight of the Cistanche deserticola powder as a compound microorganism. The mixture was then anaerobic fermented at 35℃ and pH 5 for 72 hours. After filtration and sterilization, the filtrate was concentrated to a relative density of 1.17 and then freeze-dried under vacuum to a water content of 2wt% to obtain Cistanche deserticola powder.

[0064] (3) Preparation of Cornus officinalis powder

[0065] The pulp of Cornus officinalis fruit was pressed to obtain juice and pomace. The juice was pumped into a 200 Da nanofiltration membrane system and nanofiltration was performed at 1.5 MPa and 35 °C. The retentate was concentrated to 1 / 3 of its volume and collected to obtain concentrated juice. The pomace was crushed and passed through a 10-mesh sieve and mixed with deionized water at a ratio of 1:13 (g / mL) in a subcritical water reactor. After purging with nitrogen to replace the air, extraction was carried out at 130 °C and 2.5 MPa for 30 min. The mixture was filtered, and the filtrate was mixed with the concentrated juice and concentrated to a relative density of 1.15. The mixture was then freeze-dried under vacuum to a water content of 2wt%-5wt% to obtain Cornus officinalis powder.

[0066] (4) Preparation of Gastrodia elata powder

[0067] Gastrodia elata slices were pulverized and passed through a 30-mesh sieve. They were mixed with a 65% ethanol solution at a ratio of 1:8 (g / mL) and extracted by reflux at 88℃ for 1 hour each time. The mixtures were filtered, and the filtrates were combined and concentrated to a relative density of 1.08. The mixtures were then freeze-dried under vacuum to a water content of 2 wt% to obtain Gastrodia elata powder.

[0068] (5) Preparation of Eucommia ulmoides leaf extract composition

[0069] Eucommia ulmoides leaf powder, Cistanche deserticola powder, Cornus officinalis powder and Gastrodia elata powder were mixed in a weight ratio of 40:35:25:12 to obtain an Eucommia ulmoides leaf extract composition.

[0070] Example 3

[0071] Preparation of Eucommia ulmoides leaf extract composition

[0072] (1) Preparation of Eucommia ulmoides leaf powder

[0073] Choline chloride and glycerol were mixed at a molar ratio of 1:2.5 and stirred at 80°C and 450 rpm for 80 min to obtain a deep eutectic solvent. This solvent was then mixed with deionized water at a volume ratio of 4:8 and stirred at 350 rpm for 10 min to obtain a deep eutectic solvent extract.

[0074] Eucommia ulmoides leaves were pulverized and passed through a 100-mesh sieve. They were mixed with deep eutectic solvent extract at a material-to-liquid ratio of 1:20 (g / mL) and extracted twice at 500W, 40kHz, and 55℃ for 35 min each time. The extracts were filtered, and the filtrates were combined. The filtrates were concentrated to 1 / 4 of their volume and then adsorbed onto HPD-300 macroporous resin. The extracts were then eluted sequentially with 5 BV of deionized water and 6 BV of 60% ethanol solution. The ethanol eluent was collected and concentrated to a relative density of 1.15. The eluent was then freeze-dried under vacuum to a water content of 4 wt% to obtain Eucommia ulmoides leaf powder.

[0075] (2) Preparation of Cistanche deserticola powder

[0076] The compound microbial system consists of Bifidobacterium lactis BL-99 and Lactobacillus delbrueckii subsp. bulgaricus Ouya-D-L5. The viability of Bifidobacterium lactis BL-99 in the compound microbial system is 5 × 10⁻⁶. 9 The bacterial count of *Lactobacillus delbrueckii* subsp. bulgaricus Ouya-D-L5 was 5 × 10⁻⁶ cells / g. 9 pcs / g;

[0077] The dried desert Cistanche deserticola was sliced, pulverized, and passed through a 30-80 mesh sieve to obtain Cistanche deserticola powder. The powder was mixed with liquid fermentation medium at a ratio of 1:15 (g / mL), sterilized, and inoculated with 5‰ of the weight of the Cistanche deserticola powder as a compound microorganism. The mixture was then anaerobic fermented at 38℃ and pH 6 for 36 hours. After filtration and sterilization, the filtrate was concentrated to a relative density of 1.22 and then freeze-dried under vacuum to a water content of 5wt% to obtain Cistanche deserticola powder.

[0078] (3) Preparation of Cornus officinalis powder

[0079] The pulp of Cornus officinalis fruit was pressed to obtain juice and pomace. The juice was pumped into a 300 Da nanofiltration membrane system and nanofiltration was performed at 2.5 MPa and 45 °C. The retentate was concentrated to 1 / 5 of its volume and collected to obtain concentrated juice. The pomace was crushed and passed through a 20-mesh sieve. It was mixed with deionized water at a ratio of 1:18 (g / mL) and added to a subcritical water reactor. After purging with nitrogen to replace the air, it was extracted at 140 °C and 3.5 MPa for 20 min. After filtration, the filtrate was mixed with the concentrated juice and concentrated to a relative density of 1.20. It was then freeze-dried under vacuum to a water content of 5 wt% to obtain Cornus officinalis powder.

[0080] (4) Preparation of Gastrodia elata powder

[0081] Gastrodia elata slices were pulverized and passed through an 80-mesh sieve. They were mixed with a 75% ethanol solution at a ratio of 1:12 (g / mL) and extracted by reflux at 88℃ for 2 hours. The mixture was then filtered, and the filtrates were combined and concentrated to a relative density of 1.12. The mixture was then freeze-dried under vacuum to a water content of 5 wt% to obtain Gastrodia elata powder.

[0082] (5) Preparation of Eucommia ulmoides leaf extract composition

[0083] Eucommia ulmoides leaf powder, Cistanche deserticola powder, Cornus officinalis powder and Gastrodia elata powder were mixed in a weight ratio of 60:25:15:8 to obtain an Eucommia ulmoides leaf extract composition.

[0084] Example 4

[0085] Preparation of oral beauty liquid

[0086] (1) Weigh the following ingredients in 1000 parts by weight: 50 parts of Eucommia ulmoides leaf extract composition, 50 parts of type III collagen peptide powder, 30 parts of fructooligosaccharide, 25 parts of hydroxypropyl-β-cyclodextrin, 75 parts of maltodextrin, 1 part of stevia, 3 parts of citric acid, 0.2 parts of vitamin C, 2 parts of xanthan gum, and the remainder is deionized water.

[0087] (2) Preparation of oral beauty liquid

[0088] 100 parts by weight of deionized water were mixed with hydroxypropyl-β-cyclodextrin and stirred at 55°C and 350 rpm for 35 min to obtain a cyclodextrin solution. Eucommia ulmoides leaf extract was added and stirred at 55°C and 550 rpm for 40 min to obtain a premix. 100 parts by weight of deionized water, fructooligosaccharides and maltodextrin were stirred at 250 rpm for 18 min to obtain a sugar solution. This sugar solution was mixed with the premix, xanthan gum and vitamin C, stirred at 700 rpm for 20 min, and homogenized at 30 MPa for 15 min to obtain a colloidal solution. Type III collagen peptide powder, stevia, citric acid and the remaining deionized water were added at 250 rpm, stirred for 15 min, and sterilized to obtain a cosmetic oral liquid.

[0089] Example 5

[0090] Preparation of oral beauty liquid

[0091] (1) Weigh the following ingredients in 1000 parts by weight: 40 parts of Eucommia ulmoides leaf extract composition, 60 parts of type III collagen peptide powder, 25 parts of fructooligosaccharide, 30 parts of hydroxypropyl-β-cyclodextrin, 70 parts of maltodextrin, 2 parts of stevia, 1 part of citric acid, 0.5 parts of vitamin C, 1 part of xanthan gum and the remainder of deionized water.

[0092] (2) Preparation of oral beauty liquid

[0093] 100 parts by weight of deionized water were mixed with hydroxypropyl-β-cyclodextrin and stirred at 50°C and 300 rpm for 40 min to obtain a cyclodextrin solution. Eucommia ulmoides leaf extract was added and stirred at 50°C and 500 rpm for 45 min to obtain a premix. 100 parts by weight of deionized water, fructooligosaccharides and maltodextrin were stirred at 200 rpm for 20 min to obtain a sugar solution. This sugar solution was mixed with the premix, xanthan gum and vitamin C, stirred at 700 rpm for 20 min, and homogenized at 25 MPa for 20 min to obtain a colloidal solution. Type III collagen peptide powder, stevia, citric acid and the remaining deionized water were added at 200 rpm, stirred for 18 min, and sterilized to obtain a cosmetic oral liquid.

[0094] Example 6

[0095] Preparation of oral beauty liquid

[0096] (1) Weigh the following ingredients in 1000 parts by weight: 60 parts of Eucommia ulmoides leaf extract composition, 40 parts of type III collagen peptide powder, 35 parts of fructooligosaccharide, 20 parts of hydroxypropyl-β-cyclodextrin, 80 parts of maltodextrin, 0.5 parts of stevia, 5 parts of citric acid, 0.2 parts of vitamin C, 3 parts of xanthan gum, and the remainder is deionized water.

[0097] (2) Preparation of oral beauty liquid

[0098] 100 parts by weight of deionized water were mixed with hydroxypropyl-β-cyclodextrin and stirred at 60°C and 400 rpm for 30 min to obtain a cyclodextrin solution. Eucommia ulmoides leaf extract was added and stirred at 60°C and 600 rpm for 30 min to obtain a premix. 100 parts by weight of deionized water, fructooligosaccharides and maltodextrin were stirred at 300 rpm for 15 min to obtain a sugar solution. This sugar solution was mixed with the premix, xanthan gum and vitamin C, stirred at 800 rpm for 18 min, and homogenized at 35 MPa for 10 min to obtain a colloidal solution. Type III collagen peptide powder, stevia, citric acid and the remaining deionized water were added at 300 rpm, stirred for 12 min, and sterilized to obtain a cosmetic oral liquid.

[0099] Comparative Example 1 The specific implementation method is the same as that in Example 1, except that the "deep eutectic solvent extract" in step (1) is replaced with a 75% volume fraction ethanol solution.

[0100] Comparative Example 2 The specific implementation method is the same as in Example 1, except that the composite microorganism in step (2) consists of Bifidobacterium longum strain LTBL16 and Lactobacillus plantarum ZF603, and the viability of Bifidobacterium longum strain LTBL16 in the composite microorganism is 3×10⁻⁶. 9 The bacterial activity of *Lactobacillus plantarum* ZF603 was 5 × 10⁶ cells / g. 9 pcs / g, other conditions remain unchanged.

[0101] Comparative Example 3 The specific implementation method is the same as in Example 1, except that the preparation method of Cornus officinalis powder in step (3) is as follows: the pulp of Cornus officinalis is pressed to obtain juice and pomace; the juice is pumped into a 250Da nanofiltration membrane system and nanofiltration is performed at 2MPa and 40℃. The retentate is concentrated to 1 / 4 of the retentate volume to obtain concentrated juice; the pomace is crushed and passed through a 15-mesh sieve, mixed with deionized water at a ratio of 1:15 (g / mL), and refluxed twice at 102℃ for 30 min each time. The mixture is filtered, the filtrates are combined, the filtrate is mixed with the concentrated juice, concentrated to a relative density of 1.18, and freeze-dried under vacuum to a water content of 3wt% to obtain Cornus officinalis powder.

[0102] Comparative Example 4 The specific implementation method is the same as that in Example 1, except that step (4) "70% ethanol solution" is replaced with deionized water.

[0103] Comparative Example 5 The specific implementation method is the same as in Example 1, except that Eucommia ulmoides leaf powder is omitted. The Eucommia ulmoides leaf extract composition is composed of Cistanche deserticola powder, Cornus officinalis powder and Gastrodia elata powder in a weight ratio of 55:37:18.

[0104] Comparative Example 6 The specific implementation method is the same as in Example 1, except that the powders of Cistanche deserticola, Cornus officinalis, and Gastrodia elata are omitted, and the Eucommia ulmoides leaf extract composition is only Eucommia ulmoides leaf powder.

[0105] Comparative Example 7 The specific implementation method is the same as that in Example 4, except that "Example 1 Eucommia ulmoides leaf extract composition" is replaced with "Comparative Example 1 Eucommia ulmoides leaf extract composition".

[0106] Comparative Example 8 The specific implementation method is the same as that in Example 4, except that "Example 1 Eucommia ulmoides leaf extract composition" is replaced with "Comparative Example 2 Eucommia ulmoides leaf extract composition".

[0107] Comparative Example 9 The specific implementation method is the same as that in Example 4, except that "Example 1 Eucommia ulmoides leaf extract composition" is replaced with "Comparative Example 3 Eucommia ulmoides leaf extract composition".

[0108] Comparative Example 10 The specific implementation method is the same as that in Example 4, except that "Example 1 Eucommia ulmoides leaf extract composition" is replaced with "Comparative Example 4 Eucommia ulmoides leaf extract composition".

[0109] Comparative Example 11 The specific implementation method is the same as that in Example 4, except that "Example 1 Eucommia ulmoides leaf extract composition" is replaced with "Comparative Example 5 Eucommia ulmoides leaf extract composition".

[0110] Comparative Example 12 The specific implementation method is the same as that in Example 4, except that "Example 1 Eucommia ulmoides leaf extract composition" is replaced with "Comparative Example 6 Eucommia ulmoides leaf extract composition".

[0111] Experimental Example 1

[0112] In vitro testing

[0113] (1) Anti-glycation

[0114] The inhibition rates of Eucommia ulmoides leaf extract compositions (samples) in Examples 1-3 and Comparative Examples 1-6 on advanced glycation end products (AGEs) were tested respectively.

[0115] The Eucommia ulmoides leaf extract was prepared into a sample solution with a concentration of 20 mg / mL using PBS buffer (0.1 mol / L, pH 7.4). A 10 mg / mL bovine serum albumin (BSA) solution was prepared using PBS buffer (0.1 mol / L, pH 7.4); a 0.5 mol / L glucose solution was prepared using PBS buffer (0.1 mol / L, pH 7.4); and a 20 mg / mL aminoguanidine solution (positive control) was prepared using PBS buffer (0.1 mol / L, pH 7.4). The sample solution and the aminoguanidine solution were used as the test solutions.

[0116] Add the reagents sequentially to the centrifuge tubes according to Table 1 under light-protected conditions.

[0117] Table 1. Reagent dosage for each group

[0118] Three replicates (n=3) were set up for each sample group and positive group.

[0119] Seal all centrifuge tubes and incubate them in a 37°C water bath with shaking for 7 days in the dark. After incubation, remove the reaction system and place it on ice to terminate the reaction.

[0120] Accurately pipette 200 μL of reaction solution from each reaction tube and transfer it to a black 96-well plate. Using a fluorescence spectrophotometer, set the excitation wavelength (Ex) to 370 nm and the emission wavelength (Em) to 440 nm, measure the fluorescence intensity (FI) of each well, and calculate the AGEs inhibition rate. The specific results are shown in Table 2.

[0121] AGEs inhibition rate (%) = [1 - (FI test group / FI blank group)] × 100%.

[0122] Table 2. AGEs inhibition rate (%) of each group of sample solutions

[0123] Table 2 shows that although the AGEs inhibition rate of the sample solutions in Examples 1-3 was slightly lower than that of the positive group (93.03%), it still reached an excellent level of anti-glycation. This indicates that the Eucommia ulmoides leaf extract composition, prepared by a specific process and mixed in a specific ratio from Eucommia ulmoides leaf powder, Cistanche deserticola powder, Cornus officinalis powder, and Gastrodia elata powder, is significantly effective in blocking non-enzymatic glycosylation of proteins and reducing AGEs formation. This is consistent with the mechanism by which the four raw materials exert anti-glycation effects through multi-target synergistic action. The AGEs inhibition rate of Comparative Example 1 was much lower than that of Example 1, indicating that the deep eutectic solvent-assisted ultrasonic extraction technology can more efficiently retain the anti-glycation active ingredients in Eucommia ulmoides leaves, which is a key process to ensure the anti-glycation effect. The AGEs inhibition rate of Comparative Example 2 was lower than that of Example 1, indicating that the complex microorganism composed of Bifidobacterium lactis BL-99 and Lactobacillus delbrueckii subsp. bulgaricus Ouya-D-L5 has a greater advantage in transforming Cistanche deserticola glycosides and generating auxiliary anti-glycation metabolites. The AGEs inhibition rate of Comparative Example 3 was lower than that of Example 1, indicating that subcritical water extraction and nanofiltration membrane technology are more conducive to the extraction of anti-glycation components such as iridoid glycosides from Cornus officinalis. The AGEs inhibition rates of Comparative Examples 5 and 6 were much lower than those of Example 1, fully demonstrating that the four raw materials need to work synergistically to achieve efficient anti-glycation. Single raw materials or the lack of key raw materials cannot achieve the desired effect, further verifying the importance of the multi-target synergistic anti-glycation mechanism.

[0124] (2) Antioxidant

[0125] The antioxidant capacity of the Eucommia ulmoides leaf extract compositions (samples) in Examples 1-3 and Comparative Examples 1-6 were tested respectively.

[0126] The Eucommia ulmoides leaf extract composition was prepared into a sample solution with a concentration of 5 mg / mL using PBS buffer (0.1 mol / L, pH 7.4).

[0127] The DPPH radical scavenging rate and ABTS radical scavenging rate of the sample solution were tested according to the instructions of the DPPH radical scavenging rate test kit and the total antioxidant capacity (ABTS method) test kit, respectively. The specific results are shown in Table 3.

[0128] Table 3. DPPH radical scavenging rate and ABTS radical scavenging rate of each group of sample solutions

[0129] Table 3 shows that the free radical scavenging rates of the sample solutions in Examples 1-3 were all higher than those of the Eucommia ulmoides leaf extract compositions in Comparative Examples 1-6, indicating that the Eucommia ulmoides leaf extract compositions can effectively scavenge free radicals and reduce oxidative stress, providing a solid guarantee for skin anti-aging. This is attributed to the fact that all four raw materials contain antioxidant active ingredients (such as Eucommia ulmoides leaf polyphenols and Cistanche deserticola polysaccharides), and form a synergistic defense system. The antioxidant properties of Comparative Examples 1-4 were all lower than those in Example 1, indicating that the optimized extraction process (deep eutectic solvent, subcritical water extraction, specific microorganisms, and specific solvents) can better retain the antioxidant components in the raw materials and improve the overall antioxidant effect. The antioxidant effects of Comparative Examples 5-6 were significantly lower than those in Example 1, indicating that specific complex microorganisms can promote the generation of more antioxidant metabolites from Cistanche deserticola, and that the synergistic effect of the four raw materials is necessary to maximize the antioxidant capacity; single or partial combinations of raw materials cannot achieve the same effect.

[0130] (3) Whitening

[0131] The inhibition rate of tyrosinase by the Eucommia ulmoides leaf extract compositions (samples) of Examples 1-3 and Comparative Examples 1-6 was tested respectively.

[0132] The Eucommia ulmoides leaf extract composition was prepared into a sample solution with a concentration of 5 mg / mL using PBS buffer (0.1 mol / L, pH 6.8). A 100 U / mL tyrosinase solution was prepared using PBS buffer (0.1 mol / L, pH 6.8); a 2.5 mL DOPA substrate solution was prepared using PBS buffer (0.1 mol / L, pH 6.8).

[0133] The reaction system according to Table 4 was carried out in a cuvette or a 96-well plate.

[0134] Table 4. Reagent dosage for each group

[0135] Each group has three duplicate holes (n=3).

[0136] Following the dosages in Table 4, PBS, sample solution, and tyrosinase solution were added sequentially to the 96-well plate. After mixing, the plate was incubated at 37°C for 10 min to ensure sufficient contact between the inhibitor and enzyme. L-DOPA solution at 37°C was then added to initiate the reaction. Under constant temperature (37°C), absorbance values ​​were recorded every 30 s at a wavelength of 475 nm for 3 min to obtain the reaction kinetic curve. A linear fit was performed on the data for each well, with time on the x-axis and absorbance values ​​on the y-axis, to obtain the slope, i.e., the reaction rate V.

[0137] The tyrosinase inhibition rate of the samples was calculated, and the specific results are shown in Table 5.

[0138] Tyrosinase inhibition rate (%) = [1 - (V sample group / V blank group)] × 100%.

[0139] Table 5 Tyrosinase inhibition rate (%) of each group of sample solutions

[0140] Table 5 shows that the tyrosinase inhibition rate of the sample solutions in Examples 1-3 was all above 77%, indicating that the Eucommia ulmoides leaf extract composition can effectively inhibit tyrosinase activity, reduce melanin production, and has a good whitening effect. This is related to the synergistic inhibition of the melanin synthesis pathway by the active ingredients in the raw materials (such as gastrodin and cornus iridoid glycosides). The tyrosinase inhibition rate of Comparative Examples 1-4 was lower than that of Example 1, indicating that the optimized extraction process (deep eutectic solvent, subcritical water extraction, specific microorganisms, and specific solvents) can increase the content of tyrosinase-inhibiting active ingredients in the raw materials and enhance the whitening effect. The tyrosinase inhibition rate of Comparative Examples 5-6 was much lower than that of Example 1, indicating that the synergistic effect of the four raw materials and specific microbial fermentation can form a more efficient tyrosinase inhibition system. Single raw materials or non-specific microorganisms cannot achieve the ideal whitening effect.

[0141] Experimental Example 2

[0142] animal experiments

[0143] SPF-grade 8-week-old female ICR mice (18-22g) were selected, acclimatized for 7 days, and randomly divided into 9 groups (blank group, model group 9, example group 4, comparative group 7, comparative group 8, comparative group 9, comparative group 10, comparative group 11 and comparative group 12), with 10 mice in each group.

[0144] Except for the control group, mice in other groups were subcutaneously injected with D-galactose (200 mg / kg) in the back of the neck daily for 7 consecutive weeks to establish the model. The control group was injected with an equal volume of physiological saline. Two hours after the daily injection, mice were administered the cosmetic oral solution of Example 4 / Comparative Examples 7-12 by gavage at a dose of 200 mL / kg. The control group and the model group were administered an equal weight of physiological saline by gavage for 7 consecutive weeks.

[0145] Twenty-four hours after the last gavage, mice were euthanized by cervical dislocation. Hair was removed from the backs of the mice, and skin tissue was harvested. Subcutaneous fat and connective tissue were removed, and the tissue was rinsed with pre-cooled physiological saline and dried with filter paper. 0.5g of skin tissue was taken and mixed with nine times its weight of pre-cooled physiological saline. The mixture was homogenized in an ice bath, and the resulting homogenate was repeatedly frozen and thawed three times to ensure complete cell disruption and release of contents. The homogenate was then centrifuged at 8000 rpm for 20 minutes, and the supernatant was collected.

[0146] The activities of superoxide dismutase (SOD) and glutathione peroxidase (GSH-Px) in skin tissue, as well as the contents of hydroxyproline (Hyp) and malondialdehyde (MDA) in skin tissue, were measured according to the kit instructions. The specific results are shown in Table 6.

[0147] Table 6. Activities of SOD and GSH-Px enzymes, and contents of MDA and hydroxyproline in skin tissue of mice in each group.

[0148] Table 6 shows that, in terms of SOD and GSHPx enzyme activities, although the SOD and GSHPx enzyme activities in the skin tissue of mice in Example 4 were slightly lower than those in the blank group, they were much higher than those in the model group and each comparative group. This indicates that the oral beauty liquid of the present invention can effectively enhance the activity of antioxidant enzymes in mouse skin tissue, strengthen the skin's antioxidant capacity, and reduce oxidative damage. The comparative groups showed low enzyme activity: the SOD and GSHPx enzyme activities of the oral beauty liquids in Comparative Examples 7-12 were all lower than those in Example 4, further demonstrating the importance of the synergistic effect of the four raw materials in the Eucommia ulmoides leaf extract composition and the optimized process. The absence of raw materials or improper processing can lead to a decrease in the antioxidant capacity of the oral liquid, failing to effectively restore the activity of skin antioxidant enzymes.

[0149] Regarding the hydroxyproline (Hyp) content, the Hyp content in Example 4 was close to that of the blank group and significantly higher than that of each comparative group, indicating that the oral beauty liquid of the present invention can effectively promote collagen synthesis in mouse skin and improve skin firmness and elasticity. This is attributed to the Eucommia ulmoides leaf extract composition promoting fibroblast collagen synthesis (such as Eucommia ulmoides leaf genipin and Cistanche deserticola amino acids), and its combination with type III collagen peptide powder, which provides raw materials and pathway activation support for collagen synthesis. The Hyp content of comparative groups 7-12 was lower than that of Example 4, indicating that the synergistic effect of raw materials in the Eucommia ulmoides leaf extract composition, the optimized extraction process, and the combination with type III collagen peptide powder are the key to promoting collagen synthesis and increasing Hyp content. The absence of any one of these steps will lead to a significant decrease in the effect.

[0150] Regarding the malondialdehyde (MDA) content, the MDA content in Example 4 was significantly lower than that in the model group and also significantly lower than that in each comparative group. This indicates that the oral cosmetic liquid of the present invention can effectively reduce the generation of lipid peroxidation products (MDA) in mouse skin tissue, alleviate oxidative damage, and protect skin cells and tissues. This is closely related to the strong antioxidant capacity of the Eucommia ulmoides leaf extract composition and the free radical scavenging effect of vitamin C. The MDA content in comparative groups 7-12 was higher than that in Example 4, indicating that the lack of synergistic effect of raw materials or improper processing can lead to a decrease in the antioxidant capacity of the oral liquid, making it unable to effectively inhibit lipid peroxidation, and thus unable to reduce the MDA content, resulting in ineffective repair of skin oxidative damage.

[0151] Experimental Example 3

[0152] Clinical trial validation

[0153] This test case was conducted by Shanghai Micro-Spectrum Testing Technology Group Co., Ltd., report number SHA01-25041858-JC-01.

[0154] To fully evaluate the anti-glycation and antioxidant effects of the beauty oral liquid, 40 volunteers were selected and enrolled, with 38 ultimately yielding valid samples. Before starting use of the beauty oral liquid, and after 14 and 28 days of continuous use, volunteers conducted self-assessments and non-invasive skin testing to evaluate its anti-wrinkle and antioxidant effects.

[0155] I. Volunteer Entry Criteria

[0156] Inclusion criteria: 1) Age between 30 and 60 years old; 2) Fine lines or wrinkles at the corners of both eyes that meet the criteria of SKIN AGING ATLAS 1 to 5; 3) Good health, no obvious brain, heart, liver, lung, kidney, or blood diseases, no history of long-term medication use, and a commitment to actively cooperate with the experiment; 4) Ability to read Chinese, accurately understand the experimental process, voluntarily participate, and sign an informed consent form; 5) No use of drugs or health products that may affect the results during the experiment; 6) Ability to attend follow-up visits on time, promptly report any changes in health status and medication use or adverse reactions, and comply with research requirements and schedules.

[0157] Exclusion criteria: 1) Pregnant or lactating women; 2) Women with severe systemic diseases, immunodeficiency, or autoimmune diseases; 3) Women who have undergone skin treatments, cosmetic procedures, or other interventions that may affect test results at the test site; 4) Women with active allergic diseases; 5) Women with highly sensitive constitutions; 6) Women who have used hormonal drugs or immunosuppressants within the past month; 7) Women who have participated in other clinical trials at the test site in the current or past three months.

[0158] The characteristics of the volunteers are shown in Table 7. A total of 40 volunteers were recruited, and 2 were excluded.

[0159] Table 7. Volunteer Population Characteristics RD0001 female 34 no no 2 experimental group RD0002 male 41 yes yes 3 experimental group RD0003 female 54 yes yes 3 experimental group RD0004 female 51 yes yes 3 experimental group RD0005 female 46 yes yes 2 experimental group RD0006 female 57 yes yes 4 experimental group RD0007 female 45 yes yes 3 experimental group RD0008 male 35 yes yes 2 experimental group RD0009 female 49 yes yes 2 experimental group RD0010 female 42 yes yes 3 experimental group RD0011 female 42 yes yes 4 experimental group RD0012 female 39 yes yes 2 experimental group RD0013 female 44 yes yes 2 experimental group RD0014 female 46 yes yes 2 experimental group RD0015 male 34 yes yes 1 experimental group RD0016 female 43 yes yes 2 experimental group RD0017 female 37 yes yes 1 experimental group RD0018 male 35 yes yes 1 experimental group RD0019 female 34 yes yes 1 experimental group RD0020 female 42 yes yes 2 experimental group RD0021 female 44 yes yes 2 control group RD0022 female 49 yes yes 2 control group RD0023 female 36 yes yes 1 control group RD0024 female 45 yes yes 3 control group RD0025 female 45 yes yes 2 control group RD0026 female 48 yes yes 4 control group RD0027 female 42 yes yes 2 control group RD0028 female 49 yes yes 3 control group RD0029 female 33 yes yes 1 control group RD0030 female 41 yes yes 2 control group RD0031 male 40 yes yes 1 control group RD0032 female 37 yes yes 2 control group RD0033 female 45 no no 3 control group RD0034 female 45 yes yes 3 control group RD0035 female 43 yes yes 2 control group RD0036 female 50 yes yes 4 control group RD0037 male 50 yes yes 4 control group RD0038 female 46 yes yes 2 control group RD0039 male 44 yes yes 3 control group RD0040 female 47 yes yes 3 control group

[0160] II. Test Plan

[0161] Thirty-eight volunteers were randomly divided into two groups of 19 each, ensuring no significant differences in age and gender between the two groups. The experiment employed a single-center, single-blind, pre- and post-control design. Volunteers in the experimental group took one vial (10 mL) of the beauty oral liquid twice daily, morning and evening, for a total of two vials. Volunteers in the control group took one vial (10 mL) of a non-functional control drink (physiological saline) twice daily, morning and evening, for a total of two vials.

[0162] Baseline value D0 was collected before administering the oral liquid; experimental values ​​D14 and D28 were collected at 14 and 28 days, respectively. Before data collection, subjects cleansed their faces with a facial cleanser and sat in a constant temperature and humidity environment for 30 minutes. Images were then taken to record the baseline value D0. Similarly, during the 14-day and 28-day follow-up, subjects underwent facial cleansing and 30-minute sitting in a constant temperature and humidity environment, and experimental values ​​D14 and D28 were recorded. Subjects were also asked to complete a self-assessment questionnaire.

[0163] 1. Evaluation Methods (1) Image capture and analysis (a) Facial imaging:

[0164] The Visia7 (Canfield, USA) utilizes multi-source imaging technology and image analysis software for multi-dimensional and quantitative skin analysis. Standard light (SD / Standard) provides universal white light, offering excellent assessment of most skin features through balanced cross-illumination. Cross-polarized light (CP / Cross-polarized) uses a polarized light source and a vertical polarizer on the lens to filter out surface reflections, enabling clearer visualization of subsurface details such as vascular lesions and pigment deposits.

[0165] (b) Image analysis:

[0166] Image ProPlus7 (Media Cybernetics, USA) extracts useful quantitative information by processing and analyzing digital images. Skin yellowness (b* value) has no specific unit; a significantly lower b* value indicates a reduction in skin yellowing, suggesting the sample effectively improves skin yellowing. The percentage of crow's feet area also has no specific unit; the lower the value, the smaller the area of ​​crow's feet.

[0167] (c) Skin ultrasound:

[0168] The UC22 Ultrascan (Courage+Khazaka, Germany) utilizes ultrasound imaging technology. Ultrasound waves are reflected when they encounter tissue structures of varying densities; these reflected waves are received and processed by the sensor to form a two-dimensional ultrasound image. By analyzing the ultrasound image, parameters such as skin thickness and density can be obtained. Epidermal thickness is measured in micrometers (μm). A significant increase in epidermal thickness indicates that the sample contributes to improving skin epidermal thickness.

[0169] (2) Subjective assessment Volunteers completed a self-assessment questionnaire on days 14 and 28.

[0170] (3) Safety assessment Record any adverse reactions on the skin, such as spots, edema, papules, and blisters.

[0171] 2. Test Results (1) Image analysis results (a) Epidermal thickness: Volunteers used the test product for 14 and 28 consecutive days, and the change in UC22 epidermal thickness in the test product area was significantly better than that in the blank control area (Table 8).

[0172] Table 8. UC22 Skin Thickness Test Results

[0173] Note: vs. base value change rate (%) = (average test value in the nth hour / day - average base value test value) / average base value test value * 100%.

[0174] (b) VISIA skin color b* value:

[0175] Volunteers used the test product continuously for 28 days, and the change in VISIA skin color b* value in the test product area was significantly better than that in the blank control area (Table 9).

[0176] Table 9. Results of VISIA Skin Color b* Value Test

[0177] (c) VISIA data on the percentage of area covered by crow's feet wrinkles:

[0178] Volunteers used the experimental product continuously for 28 days, and the change in the proportion of VISIA crow's feet area in the experimental product area was significantly better than that in the blank control area (Table 10).

[0179] Table 10 Results of VISIA Eye Wrinkle Area Percentage Test

[0180] (2) Instrument measurement results

[0181] (a) Moisture content of the stratum corneum:

[0182] Volunteers used the test product for 14 and 28 consecutive days. The change in skin stratum corneum moisture content in the test product area was significantly better than that in the blank control area (Table 11).

[0183] Table 11 Results of Skin Stratum Corneum Moisture Content Test

[0184] (b) Skin elasticity R2:

[0185] Volunteers used the test product continuously for 14 and 28 days, and the change in skin elasticity R2 in the test product area was significantly better than that in the blank control area (Table 12).

[0186] Table 12 Skin elasticity R2 test results (unit: %)

[0187] (c) Skin firmness F4:

[0188] Volunteers used the test product for 14 and 28 consecutive days. The skin firmness F4 change value in the test product area was significantly better than that in the blank control area (Table 13).

[0189] Table 13 Skin Tightness F4 Test Results (Unit: mm·s)

[0190] (d) Skin glycation levels (AGEs):

[0191] Volunteers used the test product for 14 and 28 consecutive days. The changes in AGEs (glycation end products) in the skin of the test product area were significantly better than those in the blank control area (Table 14).

[0192] Table 14 Results of Skin Glycation Level (AGEs) Test

[0193] (3) Volunteer self-evaluation results

[0194] After 14 days of continuous use of the test product, the experimental group showed higher evaluations of the reduction in nasolabial folds and the product's moisturizing effect compared to the control group. After 28 days of continuous use of the test product, the experimental group showed higher evaluations of the product's improvement in crow's feet and reduction of fine lines compared to the 14-day period. Furthermore, the experimental group also showed higher evaluations of the product's improvement in crow's feet, reduction of fine lines, moisturizing effect, skin brightening effect, and improvement in dull skin tone compared to the control group (Table 15).

[0195] Table 15 Results of Volunteers' Self-Assessment Questionnaire on Product D14

[0196] (3) Adverse reactions or adverse events

[0197] Adverse reactions in volunteers were followed up during the trial. No adverse reactions were reported in volunteers during this efficacy trial.

[0198] Table 16 Results of Adverse Reaction Follow-up Survey

[0199] In summary, the clinical trial results show that after 28 days of product use, the changes in UC22 epidermal thickness, VISIA skin color b* value, and VISIA crow's feet area in the product area were significantly better than those in the control area, demonstrating the product's anti-wrinkle efficacy. The change in stratum corneum moisture content in the product area was significantly better than that in the control area, indicating excellent moisturizing effects. Changes in skin elasticity (R2) and firmness (F4) were also significantly better in the product area than in the control area, indicating that the product improves skin firmness. Furthermore, the change in AGEs (advanced glycation end products) in the product area was significantly better than that in the control area, indicating that the product can effectively improve advanced glycation end products. No adverse reactions were observed during the trial.

[0200] The above description is only a preferred embodiment of the present invention. It should be noted that for those skilled in the art, several improvements and modifications can be made without departing from the principle of the present invention, and these improvements and modifications should also be considered within the scope of protection of the present invention.

Claims

1. The use of Eucommia ulmoides leaf extract composition in the preparation of oral products for anti-aging, anti-glycation, increasing epidermal thickness, improving skin barrier health, and enhancing skin firmness, characterized in that, The Eucommia ulmoides leaf extract composition consists of the following raw materials in parts by weight: 40-60 parts Eucommia ulmoides leaf powder, 25-35 parts Cistanche deserticola powder, 15-25 parts Cornus officinalis powder, and 8-12 parts Gastrodia elata powder; Eucommia ulmoides leaves were pulverized, added to a deep eutectic solvent extract, and then subjected to ultrasonic extraction, filtration, purification, concentration, and drying to obtain Eucommia ulmoides leaf powder. The molar ratio of choline chloride to glycerol in the deep eutectic solvent extract is 1:1.5-2.5; The preparation method of the Cistanche deserticola powder includes: pulverizing dried Cistanche deserticola slices, mixing with water, inoculating with compound microorganisms for anaerobic fermentation, filtering, and drying to obtain Cistanche deserticola powder; The composite microorganism consists of Bifidobacterium lactis BL-99 and Lactobacillus delbrueckii subsp. bulgaricus Ouya-D-L5; The preparation method of the Cornus officinalis powder includes: pressing the pulp of Cornus officinalis fruit to obtain juice and pomace; pumping the juice into a 200-300 Da nanofiltration membrane system and performing nanofiltration at 1.5-2.5 MPa and 35-45℃, collecting the liquid on the retentate side to obtain concentrated juice; crushing the pomace and passing it through a 10-20 mesh sieve, mixing it with deionized water at a ratio of 1:13-18 g / mL and adding it into a subcritical water reactor, purging the air with nitrogen, extracting at 130-140℃ and 2.5-3.5 MPa for 20-30 min, filtering, mixing the filtrate with the concentrated juice, concentrating to a relative density of 1.15-1.20, and freeze-drying under vacuum to a water content of 2wt%-5wt% to obtain Cornus officinalis powder; The preparation method of the gastrodia powder includes: crushing gastrodia slices and mixing them with an ethanol solution, reflux extraction, filtering, concentrating and drying the filtrate to obtain gastrodia powder.

2. The use according to claim 1, characterized in that, The preparation method of the Eucommia ulmoides leaf extract composition includes: mixing Eucommia ulmoides leaf powder, Cistanche deserticola powder, Cornus officinalis powder and Gastrodia elata powder to obtain the Eucommia ulmoides leaf extract composition.

3. A cosmetic oral liquid containing the Eucommia ulmoides leaf extract composition as described in claim 1, characterized in that, It comprises the following components: 40-60 parts of the Eucommia ulmoides leaf extract composition, 40-60 parts of type III collagen peptide powder, 25-35 parts of fructooligosaccharides, 20-30 parts of hydroxypropyl-β-cyclodextrin, 70-80 parts of maltodextrin, 0.5-2 parts of stevia, 1-5 parts of citric acid, 0.01-0.5 parts of vitamin C, and 1-3 parts of xanthan gum.

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