Collagen-containing anti-aging composition as well as preparation method and application thereof

Abstract

The invention relates to the technical field of collagen cosmetics, in particular to an anti-aging composition containing collagen as well as a preparation method and application of the anti-aging composition. The anti-aging composition comprises recombinant III-type human collagen and a plant active component, the plant active component at least comprises any one of a fruiting body extract of tremella aurantialba (NAEMATELIA AURANTIALBA) and an extract of gentiana triflora (GENTIANA TRIFLORA), the anti-aging composition also comprises an auxiliary agent, and the auxiliary agent is a mixture of the recombinant III-type human collagen, the recombinant III-type human collagen and the plant active component, and the plant active component is a mixture of the recombinant III-type human collagen, the recombinant III-type human collagen and the plant active component. The auxiliary agent at least comprises any one or more of trehalose, mannitol, carbomer, hydroxyethyl cellulose, glycerol, butanediol and pentanediol. The anti-aging composition not only can achieve a synergistic effect in the aspect of anti-aging effect, but also can solve the problem of insufficient stability of recombinant III-type human collagen to a certain extent through reasonable compounding of the components.

Description

Technical Field

[0001] The present invention relates to the technical field of collagen cosmetics, and in particular to an anti-aging composition containing collagen, a preparation method thereof, and an application thereof. Background Art

[0002] In today's society, with people's increasing focus on beauty and health, anti-aging research has become a hot topic. Collagen, as an important biomaterial, has a significant effect on skin anti-aging. Collagen is the primary structural protein in human skin, accounting for 70%-80% of the total skin protein. It maintains skin's elasticity and firmness, reduces wrinkle formation, and maintains a youthful appearance.

[0003] However, traditional single collagen ingredients have certain limitations in anti-aging applications. First, collagen itself has poor stability and is easily affected by external factors such as temperature, light, enzymes, etc., causing degradation or denaturation, resulting in reduced efficacy. Secondly, the role of single collagen in promoting skin cell regeneration, anti-oxidation, and repairing damaged skin tissue is relatively limited, and it cannot fully meet people's demand for comprehensive anti-aging effects. In addition, it is difficult to form a complete anti-aging system using collagen alone, and it is impossible to carry out multi-faceted intervention on the various mechanisms of skin aging. Skin aging is a complex physiological process involving the combined effects of multiple factors such as free radical damage, slowed cell metabolism, loss of collagen and elastin, and skin microcirculation disorders. It is difficult to fundamentally reverse the state of skin aging by relying solely on the supplementation of collagen. Therefore, compositions containing collagen have become the current research and development focus.

[0004] The sources of collagen currently used on the market mainly include animal-derived extraction and genetically engineered-derived extraction. Among them, animal collagen has high biological activity, is extracted from natural biological tissues, and has a complete and stable triple helix structure, high homology, and high compatibility with the human body, but its disadvantages are high immunogenicity and higher viral risk. The collagen prepared from genetically engineered sources is called recombinant collagen, which is obtained by fermenting the collagen DNA in the human body into bacteria such as Escherichia coli and yeast. It has the characteristics of high water solubility, low allergenicity, controllable and mass production, which reduces the occurrence of immune rejection reactions and has no viral risks. However, the defect of recombinant collagen is that its triple helix structure, which plays a major role, is not stable enough, which restricts its use in certain scenarios.

[0005] Although there are many studies on composite anti-aging / repair compositions containing collagen in the prior art, for example, patent CN116617144B uses plant extracts to promote the penetration of active ingredients and achieve safe and non-irritating moisturizing and repair by compounding collagen, modified sodium hyaluronate and eucalyptus / cardamom extracts; patent CN118787723B is mainly based on fish collagen powder, combined with astaxanthin powder and Amomum villosum / Psambar formosana extracts, focusing on scavenging free radicals and improving skin elasticity; patent CN119367238B further emphasizes synergistic enhancement to promote collagen regeneration by compounding active ingredients such as Carrageenan extract, Pheretima striata extract, deer bone collagen, and tripeptide-1. However, similar to the above technologies, although it has solved the problem of synergistic enhancement of collagen to a certain extent, it still has certain limitations, especially it has not specifically solved the core problem of insufficient stability of collagen. Therefore, it is still necessary to develop a more comprehensive and stable composite anti-aging composition to meet market demand. Summary of the Invention

[0006] In order to solve the above technical problems, the purpose of the present invention is to provide an anti-aging composition containing collagen, a preparation method and application thereof, which can not only have a synergistic effect in anti-aging effect, but also, through the rational compounding of components, can to a certain extent solve the problem of insufficient stability of recombinant type III human collagen.

[0007] In order to achieve the above technical effects, the present invention adopts the following technical solutions:

[0008] In a first aspect, the present invention provides an anti-aging composition containing collagen, comprising:

[0009] Recombinant type III human collagen and plant active ingredients, wherein the plant active ingredients include at least one of a NAEMATELIA AURANTIALBA fruiting body extract and a GENTIANA TRIFLORA extract.

[0010] Preferably, the plant active ingredient is a combination of a Tremella fuciformis (Naematelia aurantialba) fruiting body extract and a Gentiana triflora (Gentiana triflora) extract.

[0011] Preferably, the composition further comprises an auxiliary agent, and the auxiliary agent comprises at least one or more of trehalose, mannitol, carbomer, hydroxyethyl cellulose, glycerol, butylene glycol, and pentylene glycol.

[0012] Preferably, the anti-aging composition containing collagen comprises, in parts by mass:

[0013] 1-10 parts of recombinant type III human collagen, 10-20 parts of Naematelia aurantialba fruiting body extract, 10-20 parts of Gentiana triflora extract and 5-20 parts of auxiliary agent.

[0014] Preferably, the Tremella fuciformis fruiting body extract is prepared by the following method: after crushing the Tremella fuciformis fruiting body, extracting it with 5-10 times the amount of 70% ethanol solution at 55-65°C for 2-3 times, each time for 2 hours, combining the extracts, concentrating under reduced pressure and then freeze-drying to obtain the Tremella fuciformis fruiting body extract.

[0015] Preferably, the preparation method of the Gentiana triflora extract is:

[0016] The dried whole herb of Gentiana triflora was crushed, and a 60% methanol solution was added at a solid-liquid ratio of 1:10-15 (g / mL). The mixture was extracted at an ultrasonic power of 300-500W for 30-45 minutes. The extract was collected by filtration and purified using an HPD-600 macroporous resin with 30% ethanol as the eluent. The target fraction was collected and dried to obtain the Gentiana triflora extract.

[0017] Furthermore, during the preparation of the Gentiana triflora extract, the target fraction is collected by first eluting with 1-2 times the volume of pure water from a macroporous resin column, using 30% ethanol as the eluent. The ethanol eluate is then concentrated under reduced pressure and freeze-dried to obtain the Gentiana triflora extract. This preparation method efficiently enriches active substances such as iridoids and flavonoids, including gentiopicroside, to ensure the extract's anti-aging effects.

[0018] Preferably, in the anti-aging composition containing collagen, the auxiliary agent is composed of the following ingredients in parts by mass: 2-8 parts of trehalose, 1-6 parts of mannitol, 1-4 parts of carbomer and 3-10 parts of glycerol. In the auxiliary agent, trehalose and mannitol, as small molecule sugar alcohols, can protect the molecular structure of collagen and plant active ingredients through hydrogen bonding.

[0019] In a second aspect, the present invention further provides a method for preparing the aforementioned anti-aging composition containing collagen, characterized in that it comprises the following steps:

[0020] S1: Premixed solvent preparation: Add 50-70% deionized water by mass based on the total mass of the composition to a reaction kettle, heat to 35-45°C, and control the speed to 200-300 rpm while stirring; slowly add the auxiliary agent and continue stirring for 20-30 minutes until completely dissolved to obtain a base solution;

[0021] S2: Plant extract dispersion: Add the plant active ingredient to the base solution of step S1, reduce the stirring speed to 150-200 rpm, and continue stirring at 35-40°C for 15-25 minutes to fully disperse the plant extract and make it compatible with the base solution to obtain a mixed system;

[0022] S3: Adding recombinant collagen: Lower the temperature of the mixture to 25-30°C, add recombinant human collagen type III and add deionized water, adjust the stirring speed to 100-150 rpm, and stir at low speed for 10-15 minutes until completely dispersed;

[0023] S4: adjusting the pH to 5.5-6.8 and performing sterile filtration to obtain the anti-aging composition containing collagen.

[0024] In a third aspect, the present invention further provides a use of the anti-aging composition containing collagen provided in the first aspect in cosmetics processing.

[0025] Preferably, in the application, the cosmetic is a leave-on cosmetic, and in the cosmetic, the amount of the anti-aging composition containing collagen is 0.1%-5% wt%.

[0026] Compared with the prior art, the present invention has the following beneficial effects:

[0027] The present invention provides an anti-aging composition containing collagen. In terms of plant active ingredients, the preferred combination is the synergistic combination of a Naematelia aurantialba fruiting body extract and a Gentiana triflora extract. Among them, the Naematelia aurantialba fruiting body adopts a specific concentration of ethanol, temperature and multiple extraction processes to efficiently extract effective ingredients; the Gentiana triflora extract is purified by ultrasound-assisted macroporous resin, and a precise fraction collection method based on the characteristic peak of gentiopicroside ensures the high purity and high activity of the extract, providing a solid guarantee for the anti-aging effect of the composition. In this combination, the Naematelia aurantialba fruiting body extract is rich in polysaccharides and flavonoids, has a strong antioxidant capacity, can scavenge free radicals, and delay cell aging; the gentiopicroside of the Gentiana triflora extract can anti-inflammatory repair and accelerate skin cell metabolism. The combination of the three plays a role in multiple dimensions such as nutritional supplementation, anti-oxidation, and anti-inflammatory repair, achieving efficient anti-aging.

[0028] At the same time, the plant active ingredients and adjuvants provided by the present invention also play a key stabilizing role in the composition. The composition is preferably: the adjuvant is composed of 2-8 parts of trehalose, 1-6 parts of mannitol, 1-4 parts of carbomer and 3-10 parts of glycerol. The components of the adjuvant can effectively enhance the synergistic stability between the ingredients, extend the shelf life of the product, and ensure the quality stability of the product during storage and use.

[0029] Finally, the anti-aging composition containing collagen provided by the present invention has a wide range of application prospects in the field of cosmetics processing, and is particularly suitable for the development of leave-on cosmetics. In leave-on cosmetics, such as emulsions, creams, essences, etc., the recommended addition amount of the anti-aging composition is 0.1%-5% (wt%). In actual application, it can be reasonably adjusted within the range of the addition amount according to the specific formula system and expected efficacy of the cosmetics, so that it can not only effectively play an anti-aging effect, but also be well compatible with a variety of cosmetic systems. It can be used for a variety of products such as creams, essences, facial masks, etc. to meet the needs of different consumers, and has great market application potential. DETAILED DESCRIPTION

[0030] The following examples are only used to more clearly illustrate the technical scheme of the present invention and are therefore only used as examples, and cannot be used to limit the scope of the present invention. For those skilled in the art, any equivalent modifications and substitutions to the embodiments described below are also within the scope of the present invention. Therefore, the equalization conversions and modifications made without departing from the spirit and scope of the present invention should all be encompassed within the scope of the present invention. In the examples, if specific conditions are not indicated, they are carried out according to normal conditions or the conditions recommended by the manufacturer. All reagents or instruments are not indicated by manufacturers and are all conventional products that can be purchased commercially.

[0031] To better illustrate the present invention, numerous specific details are provided in the following detailed description. Those skilled in the art will appreciate that the present invention can be practiced without certain specific details. In other embodiments, methods, means, equipment, and steps well known to those skilled in the art are not described in detail in order to highlight the main purpose of the present invention.

[0032] Unless otherwise defined, all technical and scientific terms used herein have the same meanings as those commonly understood by those skilled in the art. Unless otherwise specified, the units used in this specification are all international standard units, and the numerical values and numerical ranges appearing in the present invention should be understood to include the inevitable errors in industrial production.

[0033] Example 1

[0034] This embodiment provides an anti-aging composition containing collagen, which comprises, in parts by mass:

[0035] 3 parts of recombinant human type III collagen, 10 parts of Tremella fuciformis fruiting body extract, and 10 parts of Gentiana triflora extract;

[0036] At the same time, this embodiment also provides a method for preparing the aforementioned anti-aging composition containing collagen, wherein the anti-aging composition contains 3 parts of recombinant human type III collagen, 10 parts of Tremella fuciformis fruiting body extract, and 10 parts of Gentiana triflora extract as active ingredients, and the total concentration of the active ingredients is 3 mg / mL, comprising the following steps:

[0037] S1: Preparation of premixed solvents:

[0038] Add 50-70% of the total mass of the composition to the reactor, heat to 40°C, and control the speed to 300 rpm while stirring;

[0039] S2: Plant extract dispersion:

[0040] (1) Preparation of Tremella fuciformis fruiting body extract:

[0041] Take dried Tremella fuciformis fruiting bodies, crush them into 40-60 meshes, add 70% ethanol solution at a solid-liquid ratio of 1:10 (g / mL), control the temperature at 60°C, stir and extract twice, each time for 2 hours, collect the extracts from the two extractions; combine the extracts, concentrate under reduced pressure using a rotary evaporator, and then transfer the concentrate to a freeze dryer for freeze drying to obtain a Tremella fuciformis fruiting body extract.

[0042] (2) Preparation of Gentiana triflora extract:

[0043] Take the raw materials: take the dried whole herb of Gentiana triflora, crush it, pass it through an 80-mesh sieve, add 60% methanol solution at a material-liquid ratio of 1:12 (g / mL), extract for 40 minutes at 400W ultrasonic power and ultrasonic frequency of 20kHz, filter and collect the extract, and concentrate it to 1 / 3 of the original extract volume; then use HPD-600 macroporous resin purification for purification, first use pure water twice the volume of the macroporous resin column to elute and remove impurities during the purification process, use 30% ethanol as the eluent, collect the ethanol eluate, concentrate the eluate under reduced pressure and then freeze-dry to obtain the Gentiana triflora extract.

[0044] (3) Weigh the plant active ingredients according to the mass ratio in the formula. The plant active ingredients in this embodiment are composed of Tremella fuciformis fruiting body extract and Gentiana triflora extract in a ratio of 1:1. Mix the two and add them to the deionized water in step S1. Reduce the stirring speed to 200 rpm and continue stirring at 40°C for 20 minutes to fully disperse the plant extract and make it compatible with the deionized water to obtain a mixed system.

[0045] S3: Recombinant collagen added:

[0046] Lower the temperature of the mixed system to 30°C, add recombinant human collagen type III according to the mass fraction, add deionized water, adjust the stirring speed to 100-150 rpm, and stir at low speed for 10-15 minutes until completely dispersed;

[0047] S4: Adjust the pH to 5.5-6.8 and perform sterile filtration to obtain the anti-aging composition containing collagen, which is recorded as Experimental Group 1.

[0048] Test Example 1

[0049] This test example provides a plurality of different anti-aging compositions containing collagen based on the above-mentioned Example 1, and conducts efficacy tests on them, specifically including:

[0050] 1.1 Experimental Grouping

[0051] This test case set up multiple experimental groups, and the active ingredient composition of each experimental group is shown in Table 1:

[0052] Table 1 The active ingredient composition of each experimental group

[0053]

[0054] 1.2 Efficacy test and test results

[0055] The test samples were prepared according to the composition of the above experimental groups. The specific method was as follows: each component was weighed according to the mass ratio of the formula, and dissolved in water to prepare a test sample with a total active ingredient concentration of 3 mg / mL for testing.

[0056] Experimental results: The experimental data were analyzed using SPSS, and all data are expressed as mean ± standard deviation (x ± s). One-way analysis of variance was used for comparison between groups. If the results of the variance analysis showed statistically significant differences (P < 0.05), the Tukey test was used for further comparison between groups.

[0057] (1) In vitro antioxidant test

[0058] DPPH free radical scavenging rate test: prepare 6×10 -5 mol / L DPPH ethanol solution, take 2mL (3mg / mL) sample solution and mix it with 2mL DPPH solution, react in the dark at 25±1℃ for 30min, and measure the absorbance at 517nm using a microplate reader, which is recorded as A1; take 2mL sample solution and mix it with 2mL anhydrous ethanol, measure the absorbance, which is recorded as A2; take 2mL DPPH solution and mix it with 2mL anhydrous ethanol, measure the absorbance, which is recorded as A0. The DPPH free radical scavenging rate is calculated according to the following formula:

[0059] DPPH free radical scavenging rate (%) = [(A0-A1) / A0] x 100%. Each sample was measured in parallel 3 times and the average value was taken.

[0060] SOD activity assay (xanthine oxidase method): Prepare 0.2 mmol / L xanthine substrate solution and 0.1 U / mL xanthine oxidase solution. Take 0.1 mL of the sample solution, add 2.8 mL of Tris-HCl buffer (pH 8.2) and 0.1 mL of the xanthine substrate solution, preheat at 37°C for 5 minutes, then add 0.1 mL of the xanthine oxidase solution to initiate the reaction. After 10 minutes, terminate the reaction and measure the absorbance at 560 nm. The amount of sample required to inhibit xanthine oxidase activity by 50% is defined as one SOD activity unit (U). Calculate the SOD activity (U / mL) of the sample. Perform three replicate assays for each sample and take the average value.

[0061] The experimental results are shown in Table 2:

[0062] Table 2 In vitro antioxidant test results of each composition

[0063]

[0064]

[0065] *: indicates significant difference compared with experimental group 9 (P<0.05); **: indicates extremely significant difference compared with experimental group 9 (P<0.01);

[0066] The above experimental results show that the experimental results of experimental group 7 showed that its antioxidant activity (DPPH 84.2%, SOD78.5U / mL) was significantly higher than that of using only Tremella fuciformis or Gentiana scabra alone, and exceeded the sum of the effects of the two alone, indicating that the two extracts can play a synergistic role in antioxidant effect.

[0067] At the same time, the antioxidant activity of experimental group 1 was significantly higher than that of experimental group 4, experimental group 5, and experimental group 7. The experimental data of this group proved that the combination of collagen, golden ear fruiting body extract, and three-flowered gentian extract can achieve a synergistic effect. In addition, compared with experimental groups 2 and 3, experimental group 1 had the best antioxidant effect, and its DPPH scavenging rate was significantly better than that of experimental groups 2 and 3, indicating that the recombinant type III human collagen, golden ear fruiting body extract, and three-flowered gentian extract are optimally matched in a mass ratio of 3:10:10, which can exhibit the best antioxidant activity.

[0068] (2) Human fibroblast experimental cell culture:

[0069] Based on the aforementioned antioxidant effect test, the following test was conducted only on experimental group 1, without testing experimental groups 2 and 3; at the same time, only on experimental group 7, without testing experimental groups 8 and 9. The specific test methods are as follows:

[0070] Cell culture and treatment: Human fibroblasts were seeded in 96-well plates at a density of 5 × 10 4 cells / well and culture in a 37°C, 5% CO2 incubator. After the cells adhere, add 20 μL of the sample from each experimental group. Set up 6 replicate wells for each group. After 72 hours of culture, discard the culture medium and wash the cells three times with PBS, thoroughly aspirating any remaining liquid each time.

[0071] Collagen production assay: Add 100 μL of RIPA lysis buffer containing protease inhibitors to each well, lyse on ice for 30 minutes, centrifuge at 12,000 rpm at 4°C for 15 minutes, and collect the supernatant. Follow the instructions for the Human Collagen Type III ELISA kit, plot a standard curve, and determine the amount of collagen produced in the sample (μg / mL).

[0072] The experimental results are shown in Table 3:

[0073] Table 3 Results of human fibroblast cell culture experiments on the compositions of each experimental group

[0074]

[0075] *: indicates significant difference compared with experimental group 7 (P<0.05), **: indicates extremely significant difference compared with experimental group 7 (P<0.01);

[0076] The above experimental results show that:

[0077] The collagen production in experimental group 1 reached 39.8±1.5μg / mL, an increase of 213.40% compared with experimental group 7. This fully proves that the combination of recombinant human type III collagen with golden ear fruiting body extract and gentiana triflora extract has a significant synergistic effect in promoting collagen production in human fibroblasts. The collagen production in experimental groups 4 and 5 was also higher than that in experimental groups 7 and 6, indicating that the combination of plant extracts and collagen alone can also promote collagen production, but the effect is not as good as the combined effect of the three ingredients. From the difference in the data, it can be seen that the golden ear fruiting body extract and gentiana triflora extract each play an irreplaceable role in the process of synergistically promoting collagen production with collagen, and that working together can achieve better results.

[0078] (3) Human efficacy evaluation

[0079] (1) Subject screening

[0080] Basic conditions: 100 healthy female subjects aged 35-55 were recruited and divided into 5 groups (n=20). The subjects had no obvious inflammation, damage, or allergic history on their facial skin, and no serious systemic diseases (such as diabetes, abnormal thyroid function, and other diseases that may affect skin metabolism). They had not received medical beauty treatments (such as laser, radio frequency, injection fillers, etc.) in the past 3 months, and had not used cosmetics or skin care products with special effects such as anti-aging, whitening, and acne removal in the past month.

[0081] Test Method: Subjects were required to use a product containing the corresponding experimental group composition once daily, morning and evening (refer to Table 1 for the composition of the active ingredients, control the total active ingredient concentration to be consistent and 3 mg / mL, and add 0.4% wt. of the preservative phenoxyethanol to the system to ensure antiseptic requirements). The method of use was: every morning and evening, take 2 mL of the product and apply it evenly to the face, gently massaging until absorbed. This was continued for 28 days. The product was refrigerated at 2-4°C during use.

[0082] (2) Testing process

[0083] Test preparation: The test is conducted on the 0th and 28th day of the test. The subjects are required to keep their faces clean during the test. The test items include:

[0084] (01) Skin hydration measurement: Using the Corneometer skin hydration tester (probe model CM 825), five measurement points were selected: the center of the forehead, the apple muscle area of both cheeks, and the center of the mandible. During measurement, the probe was gently pressed vertically on the skin surface. The value was read after the instrument data stabilized. Each measurement point was measured three times, and the average value was taken as the skin hydration data of that point. Finally, the average value of the five points was taken as the result of the subject's facial skin hydration.

[0085] (02) Skin elasticity measurement: Using the Cutometer skin elasticity tester (model MPA580), select the R0-R7 parameter measurement mode and select one measurement point on each cheek apple muscle area. During measurement, the probe is placed tightly against the skin to ensure that no air is trapped. The instrument is started for measurement. Each measurement point is measured three times. The elasticity parameters (such as R2, R7, etc.) of each measurement are recorded and saved, and the average value is taken for subsequent analysis.

[0086] (03) Wrinkle Depth Measurement: A VISIA skin detector was used to ensure consistent shooting conditions. After the instrument automatically captured facial images, the images were processed using its accompanying analysis software to calculate parameters such as the average depth and length of wrinkles. The changes in each indicator before and after the test were compared, and the improvement rate of each indicator for each subject was calculated. The experimental results are shown in Table 4.

[0087] Table 4 Comparison of experimental results of each test group (n=20)

[0088]

[0089] *: indicates significant difference compared with experimental group 7 (P<0.05), **: indicates extremely significant difference compared with experimental group 7 (P<0.01);

[0090] The above experimental results show that after using the products of different experimental groups for 28 days, the subjects' skin moisture content, elasticity and wrinkle depth were significantly improved. Among them, experimental group 1 performed best in all three indicators, with skin moisture content increased by 42.3±3.1%, elasticity increased by 28.7±2.5%, and wrinkle depth reduced by 36.5±4.2%, which was significantly better than the other experimental groups. This shows that the combination formula can not only effectively promote collagen production in in vitro experiments, but also effectively improve skin condition in actual human use and achieve anti-aging effects. Experimental groups 4 and 5 had the second best effect on improving skin condition, further verifying the importance of the synergistic effect of the two plant extracts and collagen. However, the improvement effects of experimental group 6 containing only collagen and experimental group 7 containing only plant extracts were relatively weak, which once again confirmed the advantages of the combined use of the three ingredients.

[0091] Test Example 2

[0092] Based on the above-mentioned Experiment 1, this embodiment provides a method for testing the stability of an anti-aging composition system containing collagen, which is used to test the stability of macromolecular collagen in different composition systems. The testing method is as follows:

[0093] S1: Sample preparation

[0094] Sample 1: According to the formula of experimental group 1 in Example 1, that is, the total active ingredients are calculated by mass ratio, and its composition is: recombinant type III human collagen: Tremella fuciformis fruiting body extract: Gentiana triflora extract = 3:10:10. An anti-aging composition containing collagen is prepared according to this total active ingredient, and the total active ingredient concentration of the composition is 3 mg / mL.

[0095] Sample 2: The difference from Sample 1 is that the total active ingredients, calculated by mass ratio, are composed of: recombinant human collagen type III: Gentiana triflora extract = 3:20;

[0096] Sample 3: The difference from Sample 1 is that the total active ingredients, calculated by mass ratio, are composed of: recombinant type III human collagen: Tremella fuciformis fruiting body extract = 3:20;

[0097] Sample 4: Based on Sample 1, an adjuvant was added, and the weight ratio of recombinant human type III collagen: Tremella fuciformis fruiting body extract: Gentiana triflora extract: adjuvant is 3:10:10:20, wherein the adjuvant composition is as follows: 6 parts of trehalose, 2 parts of mannitol, 2 parts of carbomer, and 10 parts of glycerol;

[0098] Sample 5: Based on Sample 1, an adjuvant was added, and the mass ratio of recombinant human collagen type III: Tremella fuciformis fruiting body extract: Gentiana triflora extract: adjuvant is 3:10:10:20, wherein the adjuvant composition is as follows (by mass): 8 parts of trehalose, 2 parts of carbomer, and 10 parts of glycerol;

[0099] Sample 6: Based on Sample 1, an adjuvant was added, and the mass ratio of recombinant human collagen type III: Tremella fuciformis fruiting body extract: Gentiana triflora extract: adjuvant is 3:10:10:20, wherein the adjuvant composition is as follows (by mass): 8 parts of mannitol, 2 parts of carbomer, and 10 parts of glycerol;

[0100] Control group: Refer to sample 1, but without the addition of Tremella fuciformis fruiting body extract, Gentiana triflora extract, and adjuvants; the active ingredient was calculated only as recombinant human type III collagen, at approximately 0.391 mg / mL;

[0101] Throughout the experiment, except for the control group, the total active ingredient concentration of each sample was controlled to be consistent at 3 mg / mL (only recombinant human type III collagen, Tremella fuciformis fruiting body extract, and Gentiana triflora extract were counted as active ingredients). After preparation and aliquoting, each sample was stored at 4°C (refrigerated), 25°C (room temperature), and 40°C (accelerated degradation). Samples were collected 30 days later, with three samples per condition in parallel. SDS-PAGE electrophoresis was performed using a 12% separating gel and a 5% stacking gel, with 10 μL sample loaded per well. After electrophoresis, the gel was stained with Coomassie Brilliant Blue R-250 solution for 2 hours with shaking. The gel was then transferred to a destaining solution (acetic acid:methanol:water = 1:4:5) and destained with shaking. The destaining solution was changed every 30 minutes until the background became transparent and the bands were clearly visible. Gel images were acquired using a gel imaging system (Typhoon FLA 9500) and quantitatively analyzed using ImageQuantTL software to calculate collagen retention:

[0102] Retention rate (%) = IOD value of the main band after storage / IOD value of the main band of the control sample on day 0 × 100%;

[0103] The experimental results are shown in Table 5:

[0104] Table 5 Collagen retention rate of each sample under different storage conditions (%)

[0105]

[0106]

[0107] The above experimental results show that:

[0108] Sample 1 showed significant stability improvements under all temperature conditions. Under accelerated conditions at 40°C, its collagen retention rate reached 82.7%, a 46.6% increase over the control group (56.4%). This result confirms the existence of a clear synergistic protective mechanism between the extracts from the fruiting bodies of Tremella fuciformis and the extracts from Gentiana triflora:

[0109] The experimental results of samples 2 and 3 showed that when only one active ingredient was retained, the stability of collagen deteriorated significantly. Among them, sample 2, in which only the Gentiana triflora extract was added, had a retention rate of only 42.1% at 40°C, which was 25.4% lower than the control group. This indicates that the Gentiana triflora extract, when present alone, may promote collagen hydrolysis; this may be related to the organic acids (such as gentisic acid) or a small amount of free amino acids in it. The retention rate of sample 3 at 40°C was 55.2%, which was slightly higher than the control group, but 27.5% lower than sample 1, proving that the physical protective effect of the golden ear extract depends on the antioxidant synergy of gentian.

[0110] At the same time, after adding the adjuvant, samples 4-6 were further optimized based on sample 1, among which the composite adjuvant (sample 4) had the most outstanding effect, indicating that the combination of trehalose and mannitol produced a synergistic effect:

[0111] The above studies have shown that the formula provided by the present invention can significantly reduce the dependence of recombinant type III human collagen on cold chain transportation and low-temperature storage, and is suitable for heat-sensitive product systems such as skin care essences and freeze-dried preparations.

[0112] The above embodiments are intended only to illustrate the technical solutions of the present invention and are not intended to limit the scope of the present invention. Although the present invention has been described in detail with reference to the preferred embodiments, those skilled in the art will appreciate that the technical solutions of the present invention may be modified or replaced with equivalents without departing from the spirit and scope of the technical solutions of the present invention, and such modifications or equivalents shall be encompassed by the claims of the present invention. Any techniques, shapes, and structures not described in detail herein are well known.

Claims

1. An anti-aging composition containing collagen, characterized in that: include: Recombinant type III human collagen and plant active ingredients, wherein the plant active ingredients include at least one of a NAEMATELIA AURANTIALBA fruiting body extract and a GENTIANA TRIFLORA extract.

2. The anti-aging composition containing collagen according to claim 1, characterized in that: The composition further comprises an auxiliary agent, and the auxiliary agent comprises at least one or more of trehalose, mannitol, carbomer, hydroxyethyl cellulose, glycerol, butylene glycol, and pentylene glycol.

3. The anti-aging composition containing collagen according to claim 2, characterized in that: Calculated by mass, including: 1-10 parts of recombinant type III human collagen, 10-20 parts of Naematelia aurantialba fruiting body extract, 10-20 parts of Gentiana triflora extract and 5-20 parts of auxiliary agent.

4. The anti-aging composition containing collagen according to claim 2, characterized in that: The Tremella fuciformis fruiting body extract is prepared by the following method: after crushing the Tremella fuciformis fruiting body, extracting it 2-3 times with 5-10 times the amount of 70% ethanol solution at 55-65° C. for 2 hours each time, combining the extracts, concentrating under reduced pressure and then freeze-drying to obtain the Tremella fuciformis fruiting body extract.

5. The anti-aging composition containing collagen according to claim 2, characterized in that: The preparation method of the Gentiana triflora extract is as follows: The dried whole herb of Gentiana triflora was crushed, and a 60% methanol solution was added at a solid-liquid ratio of 1:10-15 (g / mL). The mixture was extracted at an ultrasonic power of 300-500W for 30-45 minutes. The extract was collected by filtration and purified using an HPD-600 macroporous resin with 30% ethanol as the eluent. The target fraction was collected and dried to obtain the Gentiana triflora extract.

6. The anti-aging composition containing collagen according to claim 5, characterized in that: The target fraction is collected by first eluting with pure water 1-2 times the volume of the macroporous resin column, then using 30% ethanol as an eluent, collecting the ethanol eluate, concentrating the eluate under reduced pressure, and then freeze-drying to obtain the Gentiana triflora extract.

7. The anti-aging composition containing collagen according to claim 1, characterized in that: The auxiliary agent is composed of the following components in parts by mass: 2-8 parts of trehalose, 1-6 parts of mannitol, 1-4 parts of carbomer and 3-10 parts of glycerol.

8. The method for preparing an anti-aging composition containing collagen according to claim 1, wherein: The following steps are involved: S1: Premixed solvent preparation: Add 50-70% deionized water by mass based on the total mass of the composition to a reaction kettle, heat to 35-45°C, and control the speed to 200-300 rpm while stirring; slowly add the auxiliary agent and continue stirring for 20-30 minutes until completely dissolved to obtain a base solution; S2: Plant extract dispersion: Add the plant active ingredient to the base solution of step S1, reduce the stirring speed to 150-200 rpm, and continue stirring at 35-40°C for 15-25 minutes to fully disperse the plant extract and make it compatible with the base solution to obtain a mixed system; S3: Adding recombinant collagen: Lower the temperature of the mixture to 25-30°C, add recombinant human collagen type III and add deionized water, adjust the stirring speed to 100-150 rpm, and stir at low speed for 10-15 minutes until completely dispersed; S4: adjusting the pH to 5.5-6.8 and performing sterile filtration to obtain the anti-aging composition containing collagen.

9. Use of the anti-aging composition containing collagen according to any one of claims 1 to 8 in cosmetics processing.

10. The use according to claim 9, characterized in that: The cosmetic is a leave-on cosmetic, and the amount of the anti-aging composition containing collagen in the cosmetic is 0.1%-5% by weight.

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