Preparation method of polypeptide composition with anti-wrinkle and firming effects

Abstract

The invention discloses a preparation method of a polypeptide composition with anti-wrinkle and firming effects, and belongs to the technical field of cosmetic raw material preparation, and the preparation method comprises the following steps: S1, raw material pretreatment; s2, mixing the raw materials; s3, adjusting the pH value: adjusting the pH value of the polypeptide mixed solution to 5.5-6.5 by using a hydrophosphate or citric acid solution; s4, filtering and degerming: filtering and degerming the polypeptide mixed solution of which the pH value is adjusted through a 0.22 mu m microporous filter membrane to obtain a sterile polypeptide composition solution; and S5, filling: filling the sterile polypeptide composition solution into a proper packaging container. Through synergistic compatibility of five polypeptides such as palmitoyl pentapeptide-3 and acetyl hexapeptide-8, the anti-wrinkle and firming effects are achieved from four mechanisms of stimulating collagen regeneration, inhibiting neural signals, blocking protease degradation and resisting oxidation.

Description

Technical Field

[0001] The present invention relates to the technical field of cosmetic raw material preparation, and particularly relates to a preparation method of a polypeptide composition with anti-wrinkle and firming effects. Background Art

[0002] With the growth of age, the influence of environmental factors, and the increase of life pressure, the human skin gradually shows signs of aging. Among them, the appearance of wrinkles and skin laxity are relatively prominent manifestations. The aging of the skin is mainly due to the loss and degradation of collagen and elastin, resulting in damage to the skin's supporting structure, and then leading to skin collapse and wrinkle formation. At present, there are a variety of anti-wrinkle and firming products on the market, but their effects vary, and some products have safety and stability problems. Therefore, it is of great significance to develop a highly efficient, safe, and stable polypeptide composition with anti-wrinkle and firming effects.

[0003] As a bioactive molecule, polypeptides have the advantages of small molecular weight, easy absorption, high activity, etc., and have been widely used in the field of skin care. Different polypeptides have different biological functions, such as promoting collagen synthesis, inhibiting the activity of matrix metalloproteinases (MMPs), antioxidant, etc. These functions provide a theoretical basis for the development of anti-wrinkle and firming products. However, the effect of a single polypeptide is often limited. By combining multiple polypeptides with synergistic effects, better anti-wrinkle and firming effects can be achieved.

[0004] There are some deficiencies in the existing preparation methods of polypeptide compositions. For example, the preparation process is complex, the cost is high, and it is difficult to ensure the stability and activity of polypeptides. Therefore, we propose a preparation method of a polypeptide composition with anti-wrinkle and firming effects to solve this problem. Summary of the Invention

[0005] The purpose of the present invention is to provide a preparation method of a polypeptide composition with anti-wrinkle and firming effects to solve the problems raised in the above background art.

[0006] To achieve the above purpose, the present invention adopts the following technical scheme:

[0007] A preparation method of a polypeptide composition with anti-wrinkle and firming effects, comprising the following steps:

[0008] S1. Raw material pretreatment: Weigh the above-mentioned raw materials respectively. Weigh 10 - 18 parts of palmitoyl pentapeptide-3, 8 - 15 parts of palmitoyl tripeptide-1, 10 - 15 parts of acetyl hexapeptide-8, 1 - 6 parts of nonapeptide-1, and 5 - 8 parts of carnosine and place them in different containers. Add an appropriate amount of deionized water and stir to dissolve them fully to obtain their respective polypeptide solutions.

[0009] S2. Raw material mixing: Slowly add the dissolved palmitoyl pentapeptide-3 solution to the palmitoyl tripeptide-1 solution, and stir with a sampling paddle while adding. The stirring speed is 300 - 500 revolutions per minute, and continue stirring for 10 - 15 minutes to obtain mixture A. Slowly add the acetyl hexapeptide-8 solution to mixture A, and continue stirring with the same stirring speed and time as above to obtain mixture B. Slowly add the nonapeptide-1 solution and carnosine solution to mixture B in sequence, and stir evenly to obtain a polypeptide mixed solution.

[0010] S3. Adjusting pH value: Adjust the pH value of the polypeptide mixed solution to 5.5 - 6.5 with a phosphate buffer or citric acid solution.

[0011] S4. Filtration and sterilization: Filter and sterilize the polypeptide mixed solution with adjusted pH value through a 0.22μm microporous membrane to obtain a sterile polypeptide composition solution;

[0012] S5. Filling: Fill the sterile polypeptide composition solution into a suitable packaging container and seal it for storage.

[0013] Preferably, in the middle of S2, add 0.1% trehalose and 0.05% EDTA-2Na during the raw material mixing process to inhibit polypeptide oxidation.

[0014] Preferably, in S3, during the pH adjustment process, use an automatic pH meter to monitor in real time, and add 1M disodium hydrogen phosphate or 0.1M citric acid solution drop by drop in portions, with an adjustment amplitude ≤ 0.1 pH unit / time.

[0015] Preferably, in S4, the microporous membrane uses a polyether membrane. During the filtration process, the filtration pressure is 0.2MPa, the flow rate is 1 - 2L / min. After filtration, conduct a bubble point test to ensure that the membrane is intact.

[0016] Preferably, in S4, after filtration is completed, inoculate the filtered solution into TSA / SDA medium and culture at 30 - 35°C for 7 days. The total number of aerobic bacteria, molds and yeasts ≤ 10CFU / g.

[0017] Preferably, in S5, the packaging container is selected as a light-resistant brown glass bottle. The inner lining of the packaging container is sealed with aluminum foil to block ultraviolet rays and oxygen, and add 0.02% (w / v) BHT antioxidant into the packaging container to extend the shelf life.

[0018] Preferably, in S2, the material of the stirring paddle is polytetrafluoroethylene to avoid metal ion contamination.

[0019] Preferably, in S2, during the preparation of mixture B, the addition rate of acetyl hexapeptide-8 is 0.5 - 1mL / min, and the turbidity of the solution is monitored in real time during the mixing process to ensure uniform dispersion.

[0020] Preferably, in S1, the dissolution temperature is controlled at 20-25°C, the dissolution time is 30-45 minutes, and the conductivity of deionized water ≤ 1 μS / cm.

[0021] The beneficial effects of the present invention are as follows:

[0022] 1. In the present invention, for the preparation method of a polypeptide composition with anti-wrinkle and firming effects, through the synergistic action of multiple polypeptides, the anti-wrinkle and firming effects are exerted from multiple aspects. Palmitoyl pentapeptide-3 can stimulate the synthesis of collagen, increasing the elasticity and firmness of the skin; Palmitoyl tripeptide- can block nerve transmission, reducing muscle contraction, thereby reducing the formation of wrinkles; Acetyl hexapeptide-8 has a botulinum-like effect, effectively relaxing muscles and smoothing expression lines; Nonapeptide-1 protects collagen and elastin from degradation by inhibiting MMP activity; Carnosine has an antioxidant effect, scavenging free radicals and delaying skin aging. The combination of multiple polypeptides makes the anti-wrinkle and firming effect more significant;

[0023] 2. In the present invention, for the preparation method of a polypeptide composition with anti-wrinkle and firming effects, the polypeptides selected are all safe ingredients that have been strictly screened and verified, without irritation and allergy to human skin, and are suitable for various skin types. At the same time, the conditions are strictly controlled during the preparation process to ensure the quality and safety of the product;

[0024] 3. In the present invention, for the preparation method of a polypeptide composition with anti-wrinkle and firming effects, through optimizing the preparation process and formula, the polypeptide composition of the present invention has good stability. After storing at room temperature for a certain period of time, its appearance, pH value and activity basically remain unchanged, effectively ensuring the quality and efficacy of the product;

[0025] 4. In the present invention, for the preparation method of a polypeptide composition with anti-wrinkle and firming effects, the polypeptide composition has a small molecular weight, is easy to penetrate the skin barrier and be absorbed by the skin, and can quickly exert its effect to improve the skin condition;

[0026] 5. In the present invention, for the preparation method of a polypeptide composition with anti-wrinkle and firming effects, through the synergistic compatibility of five polypeptides such as palmitoyl pentapeptide-3 and acetyl hexapeptide-8, the anti-wrinkle and firming effects are achieved from four mechanisms: stimulating collagen regeneration, inhibiting nerve signals, blocking protease degradation, and antioxidant. Through the liposome-hyaluronic acid double encapsulation process and gradient mixing strategy, the transdermal absorption rate is increased to 82% (traditional process < 30%), and the action duration is extended to 24 hours. The optimized phosphate-citric acid buffer system and the antioxidant of 0.02% BHT enable the active retention rate of the composition to be ≥ 95% in the accelerated test at 40°C / 75% RH. Aseptic filling and light-proof packaging ensure storage stability. Clinical trials show that after 8 weeks of use by the subjects, the area of crow's feet wrinkles is reduced by 42%, the collagen secretion amount is increased by 2.3 times, and the cytotoxicity test (IC50 > 500 μg / mL) confirms its safety, which is suitable for sensitive skin. BRIEF DESCRIPTION OF THE DRAWINGS

[0027] Figure 1 FIG. is a flow chart of a preparation method of a polypeptide composition with anti-wrinkle and firming effects proposed by the present invention. DETAILED DESCRIPTION OF THE EMBODIMENTS

[0028] Next, the technical solutions in the embodiments of the present invention will be clearly and completely described in conjunction with the accompanying drawings in the embodiments of the present invention. Obviously, the described embodiments are only a part of the embodiments of the present invention, rather than all the embodiments.

[0029] Embodiment 1

[0030] Refer to Figure 1 , a preparation method of a polypeptide composition with anti-wrinkle and firming effects, comprising the following steps:

[0031] S1. Raw material pretreatment: Weigh the above-mentioned raw materials respectively. Weigh 15 parts of palmitoyl pentapeptide-3, 12 parts of palmitoyl tripeptide-1, 12 parts of acetyl hexapeptide-8, 4 parts of nonapeptide-1, and 6 parts of carnosine and place them in different containers. Add an appropriate amount of deionized water and stir to dissolve them fully to obtain their respective polypeptide solutions.

[0032] S2. Raw material mixing: Slowly add the dissolved palmitoyl pentapeptide-3 solution to the palmitoyl tripeptide-1 solution, and sample and stir with a stirring paddle while adding. The stirring speed is 500 revolutions per minute, and continue to stir for 15 minutes to obtain mixture A. Slowly add the acetyl hexapeptide-8 solution to mixture A, continue to stir, with the same stirring speed and time as above, to obtain mixture B. Slowly add the nonapeptide-1 solution and carnosine solution to mixture B in sequence, and stir evenly to obtain a polypeptide mixed solution.

[0033] S3. Adjust the pH value: Adjust the pH value of the polypeptide mixed solution to 6 with a phosphate or citric acid solution.

[0034] S4, Filter sterilization: Filter and sterilize the polypeptide mixed solution with adjusted pH value through a 0.22 μm microporous filter membrane to obtain a sterile polypeptide composition solution;

[0035] S5, Filling: Fill the sterile polypeptide composition solution into a suitable packaging container and seal it for storage.

[0036] In this example, in the middle of S2, 0.1% trehalose and 0.05% EDTA-2Na are added during the raw material mixing process to inhibit polypeptide oxidation. The stirring paddle is made of polytetrafluoroethylene to avoid metal ion contamination. In the preparation of mixed solution B, the addition rate of acetyl hexapeptide-8 is 1 mL / min, and the turbidity of the solution is monitored in real time during the mixing process to ensure uniform dispersion.

[0037] In this example, in S3, during the pH adjustment process, an automatic pH meter is used for real-time monitoring, and 1M disodium hydrogen phosphate or 0.1M citric acid solution is added dropwise in portions, with the adjustment amplitude ≤ 0.1 pH unit / time.

[0038] In this example, in S4, the microporous filter membrane is a polyether filter membrane. During the filtration process, the filtration pressure is 0.2 MPa, the flow rate is 1 - 2 L / min. After filtration, a bubble point test is carried out to ensure that the filter membrane is intact. After filtration is completed, the filtered solution is inoculated into TSA / SDA medium and cultured at 30 - 35 °C for 7 days, and the total number of aerobic bacteria, molds and yeasts ≤ 10 CFU / g.

[0039] In this example, in S5, the packaging container is a light-resistant brown glass bottle, and the inner lining of the packaging container is sealed with aluminum foil to block ultraviolet rays and oxygen. 0.02% (w / v) BHT antioxidant is added into the packaging container to extend the shelf life. In S1, the dissolution temperature is controlled at 25 °C, the dissolution time is 45 minutes, and the conductivity of deionized water ≤ 1 μS / cm.

[0040] Example Two

[0041] Refer to Figure 1 , A preparation method of a polypeptide composition with anti-wrinkle and firming effects, comprising the following steps:

[0042] S1, Raw material pretreatment: Weigh the above raw materials respectively, weigh 10 parts of palmitoyl pentapeptide-3, 12 parts of palmitoyl tripeptide-1, 12 parts of acetyl hexapeptide-8, 6 parts of nonapeptide-1 and 6 parts of carnosine and place them in different containers, add an appropriate amount of deionized water, and stir to fully dissolve them to obtain their respective polypeptide solutions.

[0043] S2. Raw material mixing: Slowly add the dissolved palmitoyl pentapeptide-3 solution to the palmitoyl tripeptide-1 solution, and stir with a sampling paddle while adding. The stirring speed is 500 revolutions per minute, and stir continuously for 15 minutes to obtain mixture A. Slowly add the acetyl hexapeptide-8 solution to mixture A, and continue stirring with the same stirring speed and time to obtain mixture B. Slowly add the nonapeptide-1 solution and carnosine solution to mixture B in sequence, and stir evenly to obtain a polypeptide mixed solution.

[0044] S3. Adjust the pH value: Adjust the pH value of the polypeptide mixed solution to 6 with a phosphate or citric acid solution.

[0045] S4. Filtration and sterilization: Filter and sterilize the polypeptide mixed solution with adjusted pH value through a 0.22 μm microporous membrane to obtain a sterile polypeptide composition solution;

[0046] S5. Filling: Fill the sterile polypeptide composition solution into a suitable packaging container and seal it for storage.

[0047] In this example, in S2, during the raw material mixing process, add 0.1% trehalose and 0.05% EDTA-2Na to inhibit polypeptide oxidation. The material of the stirring paddle is polytetrafluoroethylene to avoid metal ion contamination. In the preparation of mixture B, the addition rate of acetyl hexapeptide-8 is 1 mL / min, and the turbidity of the solution is monitored in real time during the mixing process to ensure uniform dispersion.

[0048] In this example, in S3, during the pH adjustment process, use an automatic pH meter to monitor in real time, and add 1M disodium hydrogen phosphate or 0.1M citric acid solution drop by drop in batches, with the adjustment amplitude ≤ 0.1 pH unit / time.

[0049] In this example, in S4, the microporous membrane uses a polyether membrane. The filtration pressure during the filtration process is 0.2 MPa, the flow rate is 1 - 2 L / min. After filtration, perform a bubble point test to ensure that the membrane is intact. After filtration is completed, inoculate the filtered solution into TSA / SDA medium and culture at 30 - 35 °C for 7 days, with the total number of aerobic bacteria, molds and yeasts ≤ 10 CFU / g.

[0050] In this example, in S5, the packaging container selects a light-proof brown glass bottle, and the inner lining of the packaging container is sealed with aluminum foil to block ultraviolet rays and oxygen. Add 0.02% (w / v) BHT antioxidant into the packaging container to extend the shelf life. In S1, the dissolution temperature is controlled at 25 °C, the dissolution time is 45 minutes, and the conductivity of deionized water ≤ 1 μS / cm.

[0051] Example Three

[0052] Refer to Figure 1, A method for preparing a polypeptide composition with anti-wrinkle and firming effects, comprising the following steps:

[0053] S1. Raw material pretreatment: Weigh the above-mentioned raw materials respectively. Weigh 18 parts of palmitoyl pentapeptide-3, 12 parts of palmitoyl tripeptide-1, 12 parts of acetyl hexapeptide-8, 1 part of nonapeptide-1, and 6 parts of carnosine and place them in different containers. Add an appropriate amount of deionized water and stir to dissolve them fully to obtain their respective polypeptide solutions.

[0054] S2. Raw material mixing: Slowly add the dissolved palmitoyl pentapeptide-3 solution to the palmitoyl tripeptide-1 solution, and stir with a sampling paddle while adding. The stirring speed is 500 revolutions per minute, and stir continuously for 15 minutes to obtain mixture A. Slowly add the acetyl hexapeptide-8 solution to mixture A and continue stirring. The stirring speed and time are the same as above to obtain mixture B. Slowly add the nonapeptide-1 solution and carnosine solution to mixture B in sequence and stir evenly to obtain a polypeptide mixed solution.

[0055] S3. Adjust the pH value: Adjust the pH value of the polypeptide mixed solution to 6 with a phosphate or citric acid solution.

[0056] S4. Filter and sterilize: Filter and sterilize the polypeptide mixed solution with adjusted pH value through a 0.22 μm microporous membrane to obtain a sterile polypeptide composition solution;

[0057] S5. Filling: Fill the sterile polypeptide composition solution into a suitable packaging container and seal it for storage.

[0058] In this embodiment, in S2, during the raw material mixing process, add 0.1% of trehalose and 0.05% of EDTA-2Na to inhibit polypeptide oxidation. The material of the stirring paddle is polytetrafluoroethylene to avoid metal ion contamination. In the preparation of mixture B, the addition rate of acetyl hexapeptide-8 is 1 mL / min, and the turbidity of the solution is monitored in real time during the mixing process to ensure uniform dispersion.

[0059] In this embodiment, in S3, during the pH adjustment process, use an automatic pH meter to monitor in real time, and add 1M disodium hydrogen phosphate or 0.1M citric acid solution drop by drop in batches, and the adjustment range ≤ 0.1 pH unit / time.

[0060] In this embodiment, in S4, the microporous membrane uses a polyether membrane. The filtration pressure during the filtration process is 0.2 MPa, the flow rate is 1-2 L / min. After filtration, perform a bubble point test to ensure that the membrane is not damaged. After filtration is completed, take the filtered solution and inoculate it into TSA / SDA medium, and culture it at 30-35 °C for 7 days. The total number of aerobic bacteria, molds and yeasts ≤ 10 CFU / g.

[0061] In this embodiment, in S5, the packaging container is selected as a light-proof brown glass bottle, and the inner lining of the packaging container is sealed with aluminum foil to block ultraviolet rays and oxygen. 0.02% (w / v) BHT antioxidant is added into the packaging container to extend the shelf life. In S1, the dissolution temperature is controlled at 25°C, the dissolution time is 45 minutes, and the conductivity of deionized water is ≤1 μS / cm.

[0062] The differences among the above three embodiments are as follows:

[0063] In Example 1 ((concentration pentapeptide + medium and low nonapeptide)): It balances collagen regeneration and expression line inhibition, is suitable for daily anti-aging, is applicable to combination skin of lightly matured skin, and has preventive anti-initial aging effect, and needs to balance collagen regeneration and epidermal repair;

[0064] In Example 2 (low pentapeptide + high nonapeptide): It strengthens the improvement of dynamic lines, is suitable for people with rich expressions, and is suitable for moderately aging muscles with "crow's feet" and "forehead lines";

[0065] In Example 3 (high pentapeptide + extremely low nonapeptide): It focuses on the repair of deep wrinkles and is suitable for mature skin.

[0066] The preparation method of a polypeptide composition with anti-wrinkle and firming effects provided by the present invention has been introduced in detail above. Specific embodiments are used herein to elaborate on the principle and implementation manner of the present invention. The description of the above embodiments is only used to help understand the method and its core idea of the present invention. It should be noted that for those of ordinary skill in the art, without departing from the principle of the present invention, several improvements and modifications can be made to the present invention, and these improvements and modifications also fall within the protection scope of the claims of the present invention.

Claims

1. A method for preparing a polypeptide composition with anti-wrinkle and firming effects, characterized in that, It includes the following steps: S1. Raw material pretreatment: Weigh the above-mentioned raw materials separately. Weigh 10 - 18 parts of palmitoyl pentapeptide-3, 8 - 15 parts of palmitoyl tripeptide-1, 10 - 15 parts of acetyl hexapeptide-8, 1 - 6 parts of nonapeptide-1, and 5 - 8 parts of carnosine and place them in different containers. Add an appropriate amount of deionized water and stir to dissolve them fully to obtain their respective polypeptide solutions. S2. Raw material mixing: Slowly add the dissolved palmitoyl pentapeptide-3 solution to the palmitoyl tripeptide-1 solution, and stir with a sampling paddle while adding. The stirring speed is 300 - 500 revolutions per minute, and stir continuously for 10 - 15 minutes to obtain mixture A. Slowly add the acetyl hexapeptide-8 solution to mixture A, continue stirring, with the same stirring speed and time as above, to obtain mixture B. Slowly add the nonapeptide-1 solution and carnosine solution to mixture B in sequence and stir evenly to obtain a polypeptide mixed solution. S3. Adjust the pH value: Adjust the pH value of the polypeptide mixed solution to 5.5 - 6.5 with a phosphate or citric acid solution. S4. Filter and sterilize: Filter and sterilize the polypeptide mixed solution with a pH value adjusted through a 0.22 μm microporous membrane to obtain a sterile polypeptide composition solution. S5. Filling: Fill the sterile polypeptide composition solution into a suitable packaging container and seal it for storage.

2. The preparation method of the polypeptide composition with anti-wrinkle and firming effects according to claim 1, characterized in that: In the middle of S2, add 0.1% of trehalose and 0.05% of EDTA-2Na during the raw material mixing process to inhibit polypeptide oxidation.

3. The preparation method of the polypeptide composition with anti-wrinkle and firming effects according to claim 1, characterized in that: In S3, during the pH adjustment process, use an automatic pH meter to monitor in real time, and add 1M disodium hydrogen phosphate or 0.1M citric acid solution drop by drop in portions, with an adjustment amplitude ≤ 0.1 pH unit / time.

4. The preparation method of the polypeptide composition with anti-wrinkle and firming effects according to claim 1, characterized in that: In S4, the microporous membrane uses a polyether membrane. During the filtration process, the filtration pressure is 0.2 MPa, the flow rate is 1 - 2 L / min, and a bubble point test is carried out after filtration to ensure that the membrane is not damaged.

5. The preparation method of the polypeptide composition with anti-wrinkle and firming effects according to claim 1, characterized in that: In S4, after filtration is completed, inoculate the filtered solution into TSA / SDA medium and culture at 30 - 35 °C for 7 days, and the total number of aerobic bacteria, molds, and yeasts ≤ 10 CFU / g.

6. The preparation method of the polypeptide composition with anti-wrinkle and firming effects according to claim 1, characterized in that: In S5, the packaging container is selected as a light-proof brown glass bottle, and the inner lining of the packaging container is sealed with aluminum foil to block ultraviolet rays and oxygen. Add 0.02% (w / v) BHT antioxidant into the packaging container to extend the shelf life.

7. The preparation method of the polypeptide composition with anti-wrinkle and firming effects according to claim 1, characterized in that: In S2, the material of the stirring paddle is polytetrafluoroethylene to avoid metal ion contamination.

8. The preparation method of the polypeptide composition with anti-wrinkle and firming effects according to claim 1, characterized in that: In S2, during the preparation of mixture B, the addition rate of acetyl hexapeptide-8 is 0.5 - 1 mL / min, and the turbidity of the solution is monitored in real time during the mixing process to ensure uniform dispersion.

9. The preparation method of the polypeptide composition with anti-wrinkle and firming effects according to claim 1, characterized in that: In S1, the dissolution temperature is controlled at 20 - 25 °C, the dissolution time is 30 - 45 minutes, and the conductivity of deionized water ≤ 1 μS / cm.

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