A dunaliella salina extract, compositions comprising the same and uses thereof

Abstract

The application discloses a dunaliella salina extract, a composition containing the same and use, and mainly relates to use of a combination of the dunaliella salina extract and myoxin C in preparation of cosmetics or skin care products with repair effect, anti-photoaging, anti-aging and antioxidation. Tests prove that the composition of the application can significantly improve the cell antioxidation and anti-photoaging capacity, increase skin elasticity, strengthen skin barrier, play an antioxidation role, and make the skin look brighter.

Description

Technical Field

[0001] This invention belongs to the field of bioengineering, specifically relating to a Dunaliella salina extract, a composition containing the same, and its uses. Background Technology

[0002] Dunaliella salina, also known as Dunaliella salina, is a halophilic green microalga belonging to the order Volvariella in the class Chlorophyta. It is widely distributed in high-salinity waters, such as salt lakes, salt pans, and seawater evaporation ponds. Dunaliella salina is an alga with very strong antioxidant capabilities, and it has attracted widespread attention due to its strong salt tolerance and high content of carotenoids (mainly β-carotene).

[0003] Research over many years has shown that Dunaliella salina has the following characteristics:

[0004] 1. Adaptability to extreme environments: It can survive in harsh conditions such as high salinity (salinity up to 30% NaCl), strong light, and high temperature (above 40℃).

[0005] 2. Rich in β-carotene: When grown in high-salt and high-light environments, it synthesizes large amounts of β-carotene, giving it a reddish-orange color, which is used for natural pigment extraction.

[0006] 3. Rich in glycerol: In order to maintain cell osmotic balance in a high osmotic pressure environment, Dunaliella salina synthesizes and accumulates a large amount of glycerol, which has moisturizing and antioxidant effects.

[0007] Dunaliella salina is a highly promising microalgae resource with wide applications in food, health products, pharmaceuticals, cosmetics, bioenergy, and aquaculture. In particular, its high content of β-carotene and glycerol makes it an important ingredient for antioxidants, skincare, and health maintenance. With advancements in biotechnology and industrialization, the prospects for the development and utilization of Dunaliella salina will become even broader.

[0008] L-Ascorbic acid (L-Ascorbic acid) is a natural, water-soluble antioxidant, also known as Vitamin C, and is widely used as an anti-aging ingredient in cosmetics. However, its sensitivity to light and oxygen and its unstable structure limit its application in the cosmetic industry. International patent application WO2022 / 228475A1 discloses an ascorbic acid polypeptide derivative with a novel structure that solves the stability problems of traditional Vitamin C structures. Therefore, all content related to this novel ascorbic acid polypeptide derivative is included in this invention. The above application uses the ascorbic acid polypeptide derivative as the core ingredient to prepare an active molecule with excellent stability and efficacy—a myoglobin C raw material—and commercializes this raw material (this myoglobin C contains 1000ppm EAC-L-Carnosine).

[0009] While Dunaliella salina and Klenokinase C both hold important positions in the cosmetics industry, their mechanisms of action differ, and each has its own advantages and disadvantages when used alone. Although there are already many antioxidant and anti-photoaging products on the market, they are basically limited to the simple layering of ingredients with the same mechanism of action, offering little advantage compared to single ingredients, or lacking a clear synergistic effect. Furthermore, how to extract the key components from Dunaliella salina and combine them with Klenokinase C to maximize efficacy is a problem that urgently needs to be solved in this field. Summary of the Invention

[0010] To address the shortcomings of existing technologies, this invention provides a Dunaliella salina extract, a composition comprising the same, and its uses. This invention combines Dunaliella salina extract with myophotopeptide C, which can significantly enhance the antioxidant and anti-photoaging capabilities of cells, increase skin elasticity, strengthen the skin barrier, and exert an antioxidant effect, making the skin appear brighter and more radiant.

[0011] In a first aspect, the present invention provides the use of a combination of Dunaliella salina extract and myophotopeptide C in the preparation of cosmetics or skin care products with repairing, anti-photoaging, anti-aging and antioxidant effects.

[0012] Furthermore, the structural formula and preparation method of the myophotopeptide C are described in international patent application WO2022 / 228475A1, the specific content of which is cited below:

[0013] This disclosure provides an ascorbic acid polypeptide derivative, which is a compound or salt thereof represented by the following general formula (1):

[0014]

[0015] Where m is 0 and n is 1, or m is 1 and n is 0;

[0016] X1 and X2 are independently C1-C6 alkyl, C1-C6 alkoxy or halogen groups;

[0017] Q1 and Q2 are independently C1 to C6 alkyl chains;

[0018] a, b, and c are independent integers selected from 1 to 10;

[0019] e is 0 and d is an integer from 1 to 10, or e is 1 and d is 0.

[0020] Furthermore, the ascorbic acid polypeptide derivative is a compound represented by the following general formula (3):

[0021]

[0022] Furthermore, the ascorbic acid polypeptide derivative is a compound represented by the following general formula (4):

[0023]

[0024] Furthermore, the ascorbic acid polypeptide derivative is a compound represented by the following general formula (6):

[0025]

[0026] This disclosure provides a method for preparing the above-mentioned ascorbic acid polypeptide derivative, comprising: first activating the carboxyl group of raw material A with an amino protecting group, then reacting it with raw material B, and obtaining the ascorbic acid polypeptide derivative after deprotection of the protecting group;

[0027] Wherein, raw material A is a compound represented by the following general formula (7):

[0028]

[0029] a and b are independent integers selected from 1 to 10;

[0030] The raw material B is a compound represented by the following general formula (8):

[0031]

[0032] k is selected from an integer from 1 to 10; Q is an alkyl chain from C1 to C6;

[0033] Y is a C1-C6 alkyl group, a C1-C6 alkoxy group, or a halogen group.

[0034] Furthermore, raw material A is carnosine with an amino protecting group; raw material B is 3-O-ethyl ascorbic acid;

[0035] Furthermore, the ascorbic acid polypeptide derivative is a compound represented by general formula (3), formula (4), and formula (6);

[0036] Furthermore, the compounds represented by (3), (4), and (6) above are EAC-L-Carnosine. The applicant of the international patent application WO2022 / 228475A1 used EAC-L-Carnosine (a mixture of compounds represented by formulas (3), (4), and (6) in any proportion) as a raw material for myophotopeptide C and commercialized the myophotopeptide C (this myophotopeptide C contains 1000 ppm EAC-L-Carnosine).

[0037] Furthermore, the Dunaliella salina extract is obtained by the following extraction steps:

[0038] 1) Prepare a 10% (w / w) 1,3-butanediol solution using deionized water as the solvent;

[0039] 2) Dissolve Dunaliella salina powder in 1,3-butanediol solution at a mass-to-volume ratio of 1:20 (W / V) and stir well to obtain Dunaliella salina alcohol solution.

[0040] 3) The Dunaliella salina alcohol solution was subjected to ultrasonic treatment;

[0041] 4) After ultrasonic treatment, let the solution stand at 4°C for 1-2 hours or centrifuge at 4000 rpm for 15 minutes. Then take the supernatant and filter it through a 0.45 μm filter membrane.

[0042] 5) Filter the filtered liquid through a 0.22μm sterile filter, and then add 2% by mass of preservative to the filtrate to obtain Dunaliella salina extract.

[0043] Furthermore, the conditions for ultrasonic treatment are: static setting, ultrasonic temperature of 50°C, ultrasonic time of 60 minutes, and ultrasonic power of 360W.

[0044] Furthermore, the preservative is Cosmecare 583.

[0045] Secondly, the present invention provides a method for extracting Dunaliella salina extract, wherein the extraction steps include:

[0046] 1) Prepare a 10% (w / w) 1,3-butanediol solution using deionized water as the solvent;

[0047] 2) Dissolve Dunaliella salina powder in 1,3-butanediol solution at a mass-to-volume ratio of 1:20 (W / V) and stir well to obtain Dunaliella salina alcohol solution.

[0048] 3) The Dunaliella salina alcohol solution was subjected to ultrasonic treatment;

[0049] 4) After ultrasonic treatment, let the solution stand at 4°C for 1-2 hours or centrifuge at 4000 rpm for 15 minutes. Then take the supernatant and filter it through a 0.45 μm filter membrane.

[0050] 5) Filter the filtered liquid through a 0.22μm sterile filter, and then add 2% by mass of preservative to the filtrate to obtain Dunaliella salina extract.

[0051] Furthermore, the conditions for the ultrasonic treatment are: static setting, ultrasonic temperature of 50°C, ultrasonic time of 60 minutes, and ultrasonic power of 360W.

[0052] Furthermore, the preservative is Cosmecare 583.

[0053] Thirdly, the present invention provides a Dunaliella salina extract, obtained by the extraction method described in the second aspect.

[0054] Fourthly, the present invention provides a composition containing 0.1% ascorbic acid polypeptide derivative and 0.1% Dunaliella salina extract as described in the third aspect, based on the total mass of the composition.

[0055] Fifthly, the present invention provides a composition containing 0.25% ascorbic acid polypeptide derivative and 0.25% Dunaliella salina extract as described in the third aspect, based on the total mass of the composition.

[0056] In a sixth aspect, the present invention provides a composition containing 0.1% myophoton peptide C and 0.1% Dunaliella salina extract as described in the third aspect, based on the total mass of the composition.

[0057] In a seventh aspect, the present invention provides a composition containing 0.25% myophoton peptide C and 0.25% Dunaliella salina extract as described in the third aspect, based on the total mass of the composition.

[0058] Eighthly, the present invention provides a composition comprising myophoton peptide C and the Dunaliella salina extract described in the third aspect, wherein the mass ratio of myophoton peptide C to the Dunaliella salina extract is 1:1.

[0059] Ninthly, the present invention provides the use of the compositions described in the sixth aspect and the seventh aspect in the preparation of cosmetics or skin care products having repairing, anti-photoaging, anti-aging and antioxidant effects.

[0060] Compared with the prior art, the present invention has the following beneficial effects:

[0061] The inventors of this application have unexpectedly discovered that the combined use of L-ascorbic acid derivatives (e.g., myophotopeptide C) and Dunaliella salina extract can significantly enhance cell activity after SLS damage, inhibit UVB-induced ROS generation, enhance UVA-induced Col I inhibition, increase collagen content, and significantly improve the cells' antioxidant and anti-photoaging capabilities. This achieves multi-faceted whitening and a significant whitening effect, which is significantly higher than the effect of using L-ascorbic acid derivatives or Dunaliella salina extract alone, demonstrating a clear and significant synergistic effect. Attached Figure Description

[0062] Figure 1 The results are from an experiment testing keratinocyte activity.

[0063] Figure 2 The results of the experiment on the antioxidant capacity of keratinocytes were obtained, in which the composition sample was composed of 0.25% Dunaliella salina extract and 0.25% myophoton peptide C.

[0064] Figure 3 The results of the experiment on the antioxidant capacity of keratinocytes were obtained, in which the composition sample was composed of 0.1% Dunaliella salina extract and 0.1% myophoton peptide C.

[0065] Figure 4 The results are from an experiment on the Col I content in fibroblasts. Detailed Implementation

[0066] Although the numerical ranges and parameter approximations shown in the broad scope of this invention are intended to be approximated, the values ​​described in the specific embodiments are recorded as accurately as possible. However, any numerical value inherently contains a certain degree of error due to the standard deviation present in their respective measurements. Furthermore, all ranges disclosed herein should be understood to encompass any and all subranges contained therein. For example, the stated range “1 to 10” should be considered to include any and all subranges between the minimum value 1 and the maximum value 10 (inclusive); that is, all subranges beginning with a minimum value of 1 or greater, such as 1 to 6.1, and subranges ending with a maximum value of 10 or less, such as 5.5 to 10. Additionally, any references marked “incorporated herein” should be understood to be incorporated herein in their entirety.

[0067] Example 1: Extraction process of Dunaliella salina extract

[0068] 1) Prepare a 10% (w / w) 1,3-butanediol solution using deionized water as the solvent;

[0069] 2) Dissolve Dunaliella salina powder in 1,3-butanediol solution at a mass-to-volume ratio of 1:20 (W / V) and stir well to obtain Dunaliella salina alcohol solution.

[0070] 3) The Dunaliella salina alcohol solution was subjected to ultrasonic treatment;

[0071] 4) After ultrasonic treatment, let the solution stand at 4°C for 1-2 hours, then take the supernatant and filter it using a 0.45μm filter membrane;

[0072] 5) Filter the filtered liquid through a 0.22μm sterile filter, and then add 2% by mass of preservative to the filtrate to obtain Dunaliella salina extract.

[0073] Example 2: Cell damage repair efficacy experiment

[0074] 2.1 Keratinocyte viability test

[0075] Experimental materials: HaCaT cells (SLS induced: 1%)

[0076] Experimental method: CCK-8 (cells were induced for 24 hours after being plated with complete culture medium containing 1% SLS and the corresponding concentration of the test substance, and cell viability was detected by CCK-8 method).

[0077] Test groups: 0.5% Dunaliella salina extract; 0.5% Vitabeptide C; 0.25% Vitabeptide C + 0.25% Dunaliella salina extract (all components are mass fractions). Vitabeptide C is also known as Vitabeptide. Purchased from Hemei Biotechnology (Zhejiang) Co., Ltd.

[0078] Formula for calculating effect:

[0079] Upward adjustment rate (%) = (T / C - 1) * 100%

[0080] Where: T—mean value of cell viability of the test substance (sample group)

[0081] Mean viability of C-negative cells (Model, induced only)

[0082] Table 1. Experimental results of cell damage repair efficacy.

[0083]

[0084] Experimental results show (see) Figure 1 (See Table 1): Under the experimental conditions, compared with the negative control group, 0.5% Dunaliella salina extract and 0.5% myophoton peptide C both enhanced cell activity after SLS injury, while the combination of 0.25% Dunaliella salina extract and 0.25% myophoton peptide C showed a significant synergistic effect.

[0085] 2.2 Keratinocyte ROS test

[0086] Experimental materials: HaCaT cells (UVB induced: 40 mJ / cm²) 2 ).

[0087] Experimental method: Fluorescence method (cells were induced by UVB 24 h after plating, and ROS content of cells was detected by fluorescent probe method after adding the corresponding concentration of sample for 24 h).

[0088] Test groups: 0.5% Dunaliella salina extract; 0.5% Myo-glucosidase C; 0.25% Myo-glucosidase C + 0.25% Dunaliella salina extract (all components are mass fractions).

[0089] Effect calculation formula:

[0090] Inhibition rate (%) = (1 - T / C) * 100%

[0091] Where: T—average ROS content of the test substance (sample group)

[0092] C—Mean ROS content in the negative group (Model, induced only)

[0093] Table 2. Experimental results of keratinocytes' resistance to peroxide damage.

[0094]

[0095] Experimental results show that ( Figure 2 (See Table 2): Under the experimental conditions, compared with the negative control group, both 0.5% Dunaliella salina extract and 0.5% myophoton peptide C inhibited UV-induced ROS generation, while the combination of 0.25% Dunaliella salina extract and 0.25% myophoton peptide C had a synergistic effect.

[0096] In addition, the applicant conducted related experiments on another test group (0.2% Myoglobin C; 0.2% Dunaliella salina extract; 0.1% Myoglobin C + 0.1% Dunaliella salina extract (all components are mass fractions)), and the results are as follows. Figure 3 As shown in Table 3:

[0097] Under the experimental conditions, compared with the negative control group, both 0.2% Dunaliella salina extract and 0.2% myophoton peptide C inhibited UVB-induced ROS generation, while the combination of 0.1% Dunaliella salina extract and 0.1% myophoton peptide C showed a significant synergistic effect, achieving unexpected technical results.

[0098] Table 3. Experimental results of keratinocytes' resistance to peroxide damage.

[0099]

[0100] Example 3: Results of Col I assay in fibroblasts

[0101] Experimental materials: HSF cells (UVA induced: 40 mJ / cm²) 2 )

[0102] Experimental method: ELISA (cells were plated for 24 hours, induced by UVA, and the corresponding concentration of sample was added. The supernatant was collected after 24 hours and the cell Col I content was detected by ELISA).

[0103] Test groups: 0.2% Myo-Einstein C; 0.2% Dunaliella salina extract; 0.1% Myo-Einstein C + 0.1% Dunaliella salina extract (all components are mass fractions).

[0104] Formula for calculating effect:

[0105] Upward adjustment rate (%) = (T / C - 1) * 100%

[0106] Where: T—average content of type I collagen in the test substance (sample group)

[0107] C—Mean type I collagen content in the negative group (Model, induced only)

[0108] Table 4. Results of Col I assay in fibroblasts

[0109]

[0110] Experimental results show (see) Figure 4 (and Table 4): Under the experimental conditions, compared with the negative control group, 0.2% Dunaliella salina extract enhanced the inhibition of UVA-induced Col I and increased the content of collagen. Furthermore, the combined use of 0.1% myophoton peptide C and 0.1% Dunaliella salina extract showed a significant synergistic effect, achieving unexpected technical results.

[0111] The above description represents only preferred embodiments and is provided as an example only, not as a limitation on the combination of features necessary for carrying out the invention. The provided headings are not intended to limit the various embodiments of the invention. Terms such as “comprising,” “including,” and “including” are not intended to be limiting. Furthermore, unless otherwise stated, the plural form is included when not modified by a numeral, and “or” or “or” means “and / or.” Unless otherwise defined herein, all technical and scientific terms used herein have the same meaning as commonly understood by those skilled in the art.

[0112] The above embodiments are preferred embodiments of the present invention, but the embodiments of the present invention are not limited to the above embodiments. Any changes, modifications, substitutions, combinations, or simplifications made without departing from the spirit and principle of the present invention shall be considered equivalent substitutions and shall be included within the protection scope of the present invention.

Claims

1. The use of the combination of Dunaliella salina extract and myophoton peptide C in the preparation of cosmetics with repairing, anti-aging and antioxidant effects.

2. The use according to claim 1, characterized in that, The anti-aging mentioned refers to anti-photoaging.

3. The use according to claim 1, characterized in that, The Dunaliella salina extract was obtained by the following extraction steps: 1) Prepare a 10% (w / w) 1,3-butanediol solution using deionized water as the solvent; 2) Dissolve Dunaliella salina powder in 1,3-butanediol solution at a mass-to-volume ratio of 1:20 (W / V) and stir well to obtain Dunaliella salina alcohol solution. 3) The Dunaliella salina alcohol solution was subjected to ultrasonic treatment; 4) After ultrasonic treatment, let the solution stand at 4°C for 1-2 hours or centrifuge at 4000 rpm for 15 minutes. Then take the supernatant and filter it through a 0.45μm filter membrane. 5) Filter the filtered liquid through a 0.22μm sterile filter, and then add 2% by mass of preservative to the filtrate to obtain Dunaliella salina extract.

4. The use according to claim 3, characterized in that, The conditions for ultrasonic treatment were: static setting, ultrasonic temperature of 50℃, ultrasonic time of 60 minutes, and ultrasonic power of 360W.

5. The use according to any one of claims 3-4, characterized in that, The preservative is Cosmecare 583.

6. The use according to claim 1, characterized in that, The cosmetic contains 0.1% myo-phosphoryl peptide C and 0.1% Dunaliella salina extract, based on the total mass of the cosmetic.

7. The use according to claim 1, characterized in that, The cosmetic contains 0.25% myophoton peptide C and 0.25% Dunaliella salina extract, based on the total mass of the cosmetic.

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