Composition for promoting long-term skin moisturizing, use thereof and moisturizing product

Abstract

The present invention provides a composition for promoting long-term moisturizing of the skin, the composition comprising: acetylated hyaluronic acid or its salt, wherein the mass ratio of the acetylated hyaluronic acid or its salt in the composition is greater than or equal to 30%, preferably greater than or equal to 40%. The composition of the present invention contains hyaluronic acid and its derivatives of different molecular weights, and through the reasonable compounding of several hyaluronic acids and their derivatives, it can not only keep the skin moisture to play a physical moisturizing effect, but also penetrate into the deep layer of the skin, and promote epidermal cells to up-regulate the expression of HAS2, HABP2 and AQP3 so that epidermal cells continuously secrete hyaluronic acid, improve the ability and water content of epidermal binding water, play a biological moisturizing effect, thereby achieving the effect of long-term moisturizing and barrier repair.

Description

Technical Field

[0001] The invention belongs to the field of material science, and in particular relates to a composition for promoting long-term moisturizing of the skin, a use thereof and a moisturizing product. Background Art

[0002] The skin is a natural barrier between the body and the environment. It protects the body from environmental influences by maintaining an effective epidermal barrier, which not only resists external factors but also prevents the body from losing water. The hydration level not only affects the visible skin condition, such as skin softness, but also affects cell metabolism, enzyme activity, and signal transduction within the epidermis. Therefore, adequate skin hydration is essential to maintaining skin health, and moisturizing is an important part of basic skin care. The water content of the normal stratum corneum is 20%-35%. When the water content of the stratum corneum drops below 10%, the skin barrier function will be impaired. Atopic dermatitis, psoriasis, acne, and dry skin are all related to damaged skin barriers.

[0003] Existing moisturizing compositions usually achieve the purpose of maintaining or increasing the moisture content of the skin through physical moisturizing. They mainly add some substances with water absorption capacity to increase the moisture content of the skin, or form a closed film on the skin surface through high molecular weight polymer components to slow down the water loss on the skin surface. They cannot fundamentally change the overall hydration and state of the skin, so the effect lasts for a short time. Summary of the invention

[0004] In view of the problems existing in the prior art, the present invention provides a composition for promoting long-term moisturizing of the skin and a preparation method thereof.

[0005] Specifically, the present invention relates to the following aspects:

[0006] 1. A composition for promoting long-term moisturizing of the skin, characterized in that the composition comprises:

[0007] Acetylated hyaluronic acid or a salt thereof,

[0008] The mass ratio of the acetylated hyaluronic acid or its salt in the composition is greater than or equal to 30%, preferably greater than or equal to 40%, and more preferably 40%-60%.

[0009] 2. The composition according to item 1, characterized in that the composition further comprises:

[0010] Hydrolyzed hyaluronic acid or its salt,

[0011] One or both of low molecular weight hyaluronic acid or its salts.

[0012] 3. The composition according to item 2, characterized in that the hydrolyzed hyaluronic acid or its salt is selected from one or more of a first hydrolyzed hyaluronic acid or its salt having a molecular weight of less than 1 kDa, a second hydrolyzed hyaluronic acid or its salt having a molecular weight of 1.1k-5kDa, and a third hydrolyzed hyaluronic acid or its salt having a molecular weight of 5.1k-10kDa.

[0013] 4. The composition according to item 3, characterized in that the hydrolyzed hyaluronic acid or its salt consists of the first hydrolyzed hyaluronic acid or its salt, the second hydrolyzed hyaluronic acid or its salt, and the third hydrolyzed hyaluronic acid or its salt.

[0014] 5. The composition according to item 4 is characterized in that, in the composition, the mass ratio of the first hydrolyzed hyaluronic acid or its salt is 0.01%-17.5%, preferably 10%-15%, the mass ratio of the second hydrolyzed hyaluronic acid or its salt is 0.01%-17.5%, preferably 10%-15%, and the mass ratio of the third hydrolyzed hyaluronic acid or its salt is 0.01%-17.5%, preferably 10%-15%.

[0015] 6. The composition according to item 2 is characterized in that the composition consists of the acetylated hyaluronic acid or its salt, the hydrolyzed hyaluronic acid or its salt, and the low molecular weight hyaluronic acid or its salt. Preferably, in the composition, the mass ratio of the hydrolyzed hyaluronic acid or its salt is 0.03%-52.5%, preferably 30%-45%, and the mass ratio of the low molecular weight hyaluronic acid or its salt is 0.01%-17.5%, preferably 10%-15%.

[0016] 7. The composition according to item 2, characterized in that the molecular weight of the low molecular weight hyaluronic acid or its salt is 200k-400kDa.

[0017] 8. The composition according to item 1, characterized in that the molecular weight of the acetylated hyaluronic acid or its salt is 30k-100kDa.

[0018] 9. The composition according to any one of items 1 to 7, characterized in that the composition further comprises cross-linked hyaluronic acid or a salt thereof.

[0019] 10. Use of the composition according to any one of items 1 to 9 for preparing a composition for epidermal cells expressing HAS2, HABP2 and / or AQP3.

[0020] 11. Use of the composition according to any one of Items 1 to 9 for preparing a moisturizing composition.

[0021] 12. A moisturizing product comprising the composition according to any one of items 1 to 9.

[0022] The composition of the present invention contains hyaluronic acid and its derivatives of different molecular weights. Through the reasonable compounding of several hyaluronic acids and their derivatives, it can not only maintain skin moisture and play a physical moisturizing effect, but also penetrate deep into the skin, promote epidermal cells to upregulate the expression of HAS2, HABP2 and AQP3, so that epidermal cells continuously secrete hyaluronic acid, improve the ability of the epidermis to bind water and the water content, play a biological moisturizing role, and thus achieve the effect of long-lasting moisturizing and barrier repair. BRIEF DESCRIPTION OF THE DRAWINGS

[0023] Figure 1 Shown is the structure of the 3D full-thickness skin model after using Examples 1-2 and Comparative Examples 1-4.

[0024] Figure 2 Shown is the HAS2 expression of the 3D full-thickness skin model after using Examples 1-2 and Comparative Example 1.

[0025] Figure 3 Shown is the expression of HABP2 in the 3D full-thickness skin model after using Examples 1-2 and Comparative Example 1.

[0026] Figure 4 The figure shows the AQP3 expression of the 3D full-thickness skin model after using Examples 1-2 and Comparative Example 1. DETAILED DESCRIPTION

[0027] The present invention is further described below in conjunction with examples. It should be understood that the examples are only used to further illustrate and explain the present invention, and are not intended to limit the present invention.

[0028] Unless otherwise defined, the technical and scientific terms in this specification are the same as those commonly understood by those skilled in the art. Although similar or identical methods and materials as described herein can be used in experiments or practical applications, materials and methods are described herein below. In the event of a conflict, the present specification including the definitions therein shall prevail. In addition, materials, methods and examples are for illustration only and are not restrictive. The present invention is further described below in conjunction with specific embodiments, but is not intended to limit the scope of the present invention.

[0029] Biological moisturizing is reflected in the following three points: 1. The increase in the thickness of the stratum corneum and the integrity of the granular layer structure. A healthy, complete, and thick stratum corneum and a well-differentiated granular layer are the embodiment of good barrier function, and are also the first line of defense for the epidermis to prevent water loss. 2. The content of glycosaminoglycans in the extracellular matrix of the epidermis. Endogenous hyaluronic acid is the main glycosaminoglycan in the epidermis, and it is the main substance that binds water in the epidermis. The increase in the content of endogenous hyaluronic acid can indirectly reflect the increase in the hydration capacity of the epidermis. Hyaluronan synthase 2 (HAS2) is a secretory protein on the cell membrane surface of keratinocytes and fibroblasts, which can secrete endogenously synthesized hyaluronic acid, while hyaluronan binding protein 2 (HABP2) reflects the content of endogenous hyaluronic acid in the skin. 3. The ability of epidermal cells to transport water. Since there are no capillaries in the epidermis, the transportation of water and nutrients mainly depends on some receptors on the surface of the cell membrane. Aquaporin 3 (AQP3) is a water channel protein and a cell membrane surface receptor for epidermal cells to transport water. Increased expression of AQP3 can increase the ability of epidermal cells to transport and absorb water.

[0030] Through the above three different dimensions, not only can the loss of moisture in the epidermis be prevented, but the skin can also continuously "drink up" moisture and store the moisture it "drank" to achieve long-lasting moisturizing effect.

[0031] Hyaluronic acid is a non-sulfated glycosaminoglycan and is the main component of the extracellular matrix. 50% of the hyaluronic acid in the human body is present in the skin. Together with collagen and chondroitin sulfate, it forms the extracellular matrix of the skin, which can play a role in retaining moisture, maintaining extracellular space and regulating osmotic pressure. Since hyaluronic acid itself contains a large number of carboxyl and hydroxyl groups, it can bind a large amount of water to stabilize water retention.

[0032] Hyaluronic acid in the skin is mainly synthesized by hyaluronan synthase (HAS). Three HAS subtypes have been identified in mammals. HAS is involved in the synthesis of hyaluronic acid chains of different molecular weights on the inner side of the plasma membrane, and secretes the synthesized hyaluronic acid into the extracellular matrix. It is reported that HAS1 and HAS2 synthesize 20-2000kDa hyaluronic acid, and HAS3 synthesizes 100-1000kDa hyaluronic acid. Fibroblasts and keratinocytes mainly use HAS2. Human hyaluronan binding protein 2 (HABP2) can reflect the content of hyaluronic acid in the epidermis.

[0033] Aquaporins (AQP) are proteins located on the cell membrane. They form channels on the cell membrane and can control the flow of water through the cell membrane like a "cell pump". So far, 11 types of AQP have been found in the human body. Some references have confirmed that AQP3 plays an important role in the restoration of skin barrier function. By regulating the expression of AQP3, the skin's moisture content can be increased and skin dryness can be improved.

[0034] By regulating the above three targets, epidermal cells can continuously secrete hyaluronic acid, thereby improving the epidermis' ability to bind water and its water content, thus achieving a long-lasting moisturizing effect.

[0035] It is reported in the prior art that hyaluronic acid has the function of physical moisturizing, and it is also reported that hyaluronic acid can bind to four receptors in the skin: CD44, TLR2 / TLR4, RHAMM, and ICAM-1 receptors to regulate the physiological activities and functions of keratinocytes and fibroblasts. However, there are few studies on what kind of hyaluronic acid or its combination to promote the expression of hyaluronic acid-promoting cell moisturizing-related proteins, thereby essentially improving the moisturizing ability of the epidermis and achieving the effect of biological long-term moisturizing.

[0036] In order to achieve the effect of long-term moisturizing of the skin, the present invention provides a composition for promoting long-term moisturizing of the skin, wherein the composition comprises acetylated hyaluronic acid or a salt thereof, wherein the mass ratio of the acetylated hyaluronic acid or a salt thereof in the composition is greater than or equal to 30%, for example, it can be greater than or equal to 35%, for example, it can be 30%, 32%, 35%, 38%, 40%, 42%, 45%, 48%, 50%, 52%, 55%, 58%, 60%, 62%, 65%, 68%, 70%, 75%, 80%, preferably greater than or equal to 40%, and further preferably 40%-60%.

[0037] The composition may include only acetylated hyaluronic acid or its salt, or may further include one or more of hydrolyzed hyaluronic acid or its salt, low molecular weight hyaluronic acid or its salt, and cross-linked hyaluronic acid or its salt.

[0038] In a specific embodiment, the composition consists of acetylated hyaluronic acid or a salt thereof.

[0039] In a specific embodiment, the composition consists of acetylated hyaluronic acid or its salt, and hydrolyzed hyaluronic acid or its salt.

[0040] In a specific embodiment, the composition consists of acetylated hyaluronic acid or a salt thereof, and low molecular weight hyaluronic acid or a salt thereof.

[0041] In a specific embodiment, the composition consists of acetylated hyaluronic acid or its salt, and cross-linked hyaluronic acid or its salt.

[0042] In a preferred embodiment, the composition consists of acetylated hyaluronic acid or its salt, hydrolyzed hyaluronic acid or its salt, and low molecular weight hyaluronic acid or its salt.

[0043] Furthermore, in the composition, the mass ratio of the hydrolyzed hyaluronic acid or its salt is 0.03%-52.5%, for example, it can be 0.03%, 0.1%, 0.5%, 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 11%, 12%, 13%, 14%, 15%, 16%, 17%, 18%, 19%, 20%, 21%, 22%. %, 23%, 24%, 25%, 26%, 27%, 28%, 29%, 30%, 31%, 32%, 33%, 34%, 35%, 36%, 37%, 38%, 39%, 40%, 41%, 42%, 43%, 44%, 45%, 46%, 47%, 48%, 49%, 50%, 51%, 52%, 52.2%, preferably 30%-45%. The mass ratio of the low molecular weight hyaluronic acid or its salt is 0.01%-17.5%, for example, it can be 0.01%, 0.1%, 0.5%, 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 11%, 12%, 13%, 14%, 15%, 16%, 17%, 17.5%, preferably 10%-15%.

[0044] In a specific embodiment, the composition includes acetylated hyaluronic acid or its salt, hydrolyzed hyaluronic acid or its salt, low molecular weight hyaluronic acid or its salt, cross-linked hyaluronic acid or its salt.

[0045] Among them, acetylated hyaluronate (AcHA) is obtained by acetylation of sodium hyaluronate. The introduction of acetyl groups brings lipophilicity to hyaluronic acid, enhances the affinity and adsorption of hyaluronic acid to the skin, and improves the moisturizing performance of hyaluronic acid.

[0046] In a specific embodiment, the molecular weight of the acetylated hyaluronic acid or its salt is 30k-100kDa, for example, 30kDa, 40kDa, 50kDa, 60kDa, 70kDa, 80kDa, 30kDa, 90kDa, or 100kDa.

[0047] Hydrolyzed hyaluronic acid or its salt is a low molecular weight hyaluronic acid obtained by hydrolyzing hyaluronic acid or its salt. By controlling the hydrolysis conditions, hydrolyzed hyaluronic acid of different molecular weights can be obtained. Studies have found that the physiological effects of hyaluronic acid are closely related to the size of its molecular weight, and hyaluronic acid of different molecular weights has different biological activities. Hyaluronic acid with a smaller molecular weight has the effects of immune activity, promoting the proliferation of vascular endothelial cells and reversing the multidrug resistance of tumor cells. At the same time, due to its small molecular weight, hydrolyzed hyaluronic acid can penetrate into the epidermis.

[0048] Further, the hydrolyzed hyaluronic acid or its salt is selected from one or more of a first hydrolyzed hyaluronic acid or its salt having a molecular weight of less than 1 kDa, a second hydrolyzed hyaluronic acid or its salt having a molecular weight of 1.1 k-5 kDa, and a third hydrolyzed hyaluronic acid or its salt having a molecular weight of 5.1 k-10 kDa. That is, the hydrolyzed hyaluronic acid or its salt can be any one, any two, or three of the first hydrolyzed hyaluronic acid or its salt having a molecular weight of less than 1 kDa, the second hydrolyzed hyaluronic acid or its salt having a molecular weight of 1.1 k-5 kDa, and the third hydrolyzed hyaluronic acid or its salt having a molecular weight of 5.1 k-10 kDa. Wherein the molecular weight of the first hydrolyzed hyaluronic acid or its salt is below 1kDa, for example, it can be 1kDa, 990Da, 980Da, 970Da, 960Da, 950Da, 940Da, 930Da, 920Da, 910Da, 900Da, 890Da, 880Da, 870Da, 860Da, 850Da, 840Da, 830Da, 820Da, 810Da, 800Da, 790Da, 780Da, 770Da, 760Da, 750Da, 740Da, 730Da, 720Da, 710Da, 700Da. The molecular weight of the second hydrolyzed hyaluronic acid or its salt is 1.1k-5kDa, for example, it can be 1.1kDa, 1.5kDa, 2kDa, 2.5kDa, 3kDa, 3.5kDa, 4kDa, 4.5kDa, 5kDa. The molecular weight of the third hydrolyzed hyaluronic acid or its salt is 5.1k-10kDa, for example, 5.1kDa, 5.5kDa, 6kDa, 6.5kDa, 7kDa, 7.5kDa, 8kDa, 8.5kDa, 9kDa, 9.5kDa, or 10kDa.

[0049] In a specific embodiment, the hydrolyzed hyaluronic acid or its salt consists of the first hydrolyzed hyaluronic acid or its salt.

[0050] In a specific embodiment, the hydrolyzed hyaluronic acid or a salt thereof consists of the second hydrolyzed hyaluronic acid or a salt thereof.

[0051] In a specific embodiment, the hydrolyzed hyaluronic acid or a salt thereof consists of the third hydrolyzed hyaluronic acid or a salt thereof.

[0052] In a specific embodiment, the hydrolyzed hyaluronic acid or a salt thereof consists of the first hydrolyzed hyaluronic acid or a salt thereof, and the second hydrolyzed hyaluronic acid or a salt thereof.

[0053] In a specific embodiment, the hydrolyzed hyaluronic acid or a salt thereof consists of the first hydrolyzed hyaluronic acid or a salt thereof, and the third hydrolyzed hyaluronic acid or a salt thereof.

[0054] In a specific embodiment, the hydrolyzed hyaluronic acid or a salt thereof consists of the second hydrolyzed hyaluronic acid or a salt thereof, and the third hydrolyzed hyaluronic acid or a salt thereof.

[0055] In a preferred embodiment, the hydrolyzed hyaluronic acid or a salt thereof consists of the first hydrolyzed hyaluronic acid or a salt thereof, the second hydrolyzed hyaluronic acid or a salt thereof, and the third hydrolyzed hyaluronic acid or a salt thereof.

[0056] In a specific embodiment, in the composition, the mass ratio of the first hydrolyzed hyaluronic acid or its salt is 0.01%-17.5%, for example, it can be 0.01%, 0.1%, 0.5%, 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 11%, 12%, 13%, 14%, 15%, 16%, 17%, 17.5%, preferably 10%-15%; the mass ratio of the second hydrolyzed hyaluronic acid or its salt is 0.01%-17.5%, for example, it can be 0.01%, 0.1%, 0.5%, 1% , 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 11%, 12%, 13%, 14%, 15%, 16%, 17%, 17.5%, preferably 10%-15%; the mass ratio of the third hydrolyzed hyaluronic acid or its salt is 0.01%-17.5%, for example, it can be 0.01%, 0.1%, 0.5%, 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, 11%, 12%, 13%, 14%, 15%, 16%, 17%, 17.5%, preferably 10%-15%.

[0057] In a specific embodiment, the molecular weight of the low molecular weight hyaluronic acid or its salt is 200k-400kDa. For example, it can be 200kDa, 210kDa, 220kDa, 230kDa, 240kDa, 250kDa, 260kDa, 270kDa, 280kDa, 290kDa, 300kDa, 310kDa, 320kDa, 330kDa, 340kDa, 350kDa, 360kDa, 370kDa, 380kDa, 390kDa, 400kDa.

[0058] In a specific embodiment, the composition is composed of 12.5% ​​of the first hydrolyzed hyaluronic acid or its salt, 12.5% ​​of the second hydrolyzed hyaluronic acid or its salt, 12.5% ​​of the third hydrolyzed hyaluronic acid or its salt, 50% of the acetylated hyaluronic acid or its salt, and 12.5% ​​of the low molecular weight hyaluronic acid or its salt, according to the mass ratio of the composition. Among them, the molecular weight of the first hydrolyzed hyaluronic acid or its salt is 1kDa, the molecular weight of the first hydrolyzed hyaluronic acid or its salt is 2.6kDa, the molecular weight of the third hydrolyzed hyaluronic acid or its salt is 7kDa, the molecular weight of the acetylated hyaluronic acid or its salt is 30kDa, and the molecular weight of the low molecular weight hyaluronic acid or its salt is 373kDa.

[0059] In a specific embodiment, the composition is composed of 15% of the first hydrolyzed hyaluronic acid or its salt, 15% of the second hydrolyzed hyaluronic acid or its salt, 15% of the third hydrolyzed hyaluronic acid or its salt, 40% of the acetylated hyaluronic acid or its salt, and 15% of the low molecular weight hyaluronic acid or its salt, according to the mass ratio of the composition. Among them, the molecular weight of the first hydrolyzed hyaluronic acid or its salt is 1kDa, the molecular weight of the first hydrolyzed hyaluronic acid or its salt is 2.6kDa, the molecular weight of the third hydrolyzed hyaluronic acid or its salt is 7kDa, the molecular weight of the acetylated hyaluronic acid or its salt is 30kDa, and the molecular weight of the low molecular weight hyaluronic acid or its salt is 373kDa.

[0060] In a specific embodiment, the composition is composed of 10% of the first hydrolyzed hyaluronic acid or its salt, 10% of the second hydrolyzed hyaluronic acid or its salt, 10% of the third hydrolyzed hyaluronic acid or its salt, 60% of the acetylated hyaluronic acid or its salt, and 10% of the low molecular weight hyaluronic acid or its salt, according to the mass ratio of the composition. Among them, the molecular weight of the first hydrolyzed hyaluronic acid or its salt is 1kDa, the molecular weight of the first hydrolyzed hyaluronic acid or its salt is 2kDa, the molecular weight of the third hydrolyzed hyaluronic acid or its salt is 7kDa, the molecular weight of the acetylated hyaluronic acid or its salt is 30kDa, and the molecular weight of the low molecular weight hyaluronic acid or its salt is 373kDa.

[0061] In a specific embodiment, according to the mass ratio of the composition, the composition is composed of 12.5% ​​of the first hydrolyzed hyaluronic acid or its salt, 12.5% ​​of the second hydrolyzed hyaluronic acid or its salt, 12.5% ​​of the third hydrolyzed hyaluronic acid or its salt, 50% of the acetylated hyaluronic acid or its salt, and 12.5% ​​of the low molecular weight hyaluronic acid or its salt. Among them, the molecular weight of the first hydrolyzed hyaluronic acid or its salt is 900Da, the molecular weight of the first hydrolyzed hyaluronic acid or its salt is 3kDa, the molecular weight of the third hydrolyzed hyaluronic acid or its salt is 9kDa, the molecular weight of the acetylated hyaluronic acid or its salt is 50kDa, and the molecular weight of the low molecular weight hyaluronic acid or its salt is 250kDa.

[0062] The cross-linked hyaluronic acid in the present invention refers to ultra-high molecular weight hyaluronic acid formed by cross-linking reaction of hyaluronic acid, which is a dense three-dimensional network structure, evenly covers the surface of the skin to form a breathable film, which can enhance the skin barrier function, reduce the evaporation of water inside the skin, and the protective film formed on the skin surface is more stable and has a longer-lasting effect.

[0063] The present invention also provides a moisturizing product comprising the above composition. The above different types of hyaluronic acid in the composition of the moisturizing product can be added simultaneously or separately to the moisturizing product. Further, other ingredients can be added to the moisturizing product according to different needs and applications.

[0064] The present invention also provides use of the composition in preparing a moisturizing composition.

[0065] The present invention also provides use of the above composition in preparing a composition for promoting epidermal cells to express HAS2, HABP2 and / or AQP3.

[0066] The results of the effect of the composition on the tissue structure of the 3D full-thickness skin model showed that the composition of the present invention can significantly increase the thickness of the stratum corneum, reduce water loss, improve epidermal hydration, and keep the skin in a healthy state; it can promote epidermal cells to express HAS2, HABP2, and AQP3, thereby promoting epidermal cells to generate and secrete hyaluronic acid, improve the ability of epidermal hydration, and achieve a long-lasting moisturizing effect.

[0067] Example

[0068] The experimental methods used in the following examples are all conventional methods unless otherwise specified.

[0069] Unless otherwise specified, the materials, reagents, etc. used in the following examples can be obtained from commercial sources, and the hyaluronic acid used was purchased from Bloomage Biotech Co., Ltd.

[0070] Example 1

[0071] Weigh the hydrolyzed sodium hyaluronate with a molecular weight of 1 kDa (trade name: Hybloom 1000) TM ) 12.5g, hydrolyzed sodium hyaluronate with a molecular weight of 2.6kDa (trade name: Ximinxiu TM Super active hyaluronic acid) 12.5g, hydrolyzed sodium hyaluronate with a molecular weight of 7kDa (trade name: Nano ) 12.5 g, acetylated sodium hyaluronate with a molecular weight of 30 kDa (trade name: Hymagic TM -AcHA) 50g, low molecular weight sodium hyaluronate with a molecular weight of 373kDa (trade name: Hybloom TM Low molecular weight sodium hyaluronate) 12.5g.

[0072] The weighed components are stirred evenly to obtain a hyaluronic acid composition.

[0073] Example 2

[0074] Weigh 30 kDa acetylated sodium hyaluronate (trade name: Hymagic TM -AcHA) 100g.

[0075] That is, a hyaluronic acid composition is obtained.

[0076] Comparative Example 1

[0077] Weigh a 1 kDa molecular weight (trade name: Hybloom 1000) TM ) 50g of hydrolyzed sodium hyaluronate, 2.6kDa of hydrolyzed sodium hyaluronate (trade name: Ximinxiu TM Super active hyaluronic acid) 12.5g, hydrolyzed sodium hyaluronate with a molecular weight of 7kDa (trade name: Nano ) 12.5 g, acetylated sodium hyaluronate with a molecular weight of 30 kDa (trade name: Hymagic TM -AcHA) 12.5g, low molecular weight sodium hyaluronate with a molecular weight of 373kDa (trade name: Hybloom TM Low molecular weight sodium hyaluronate) 12.5g.

[0078] The weighed components are stirred evenly to obtain a hyaluronic acid composition.

[0079] Comparative Example 2

[0080] Weigh a 1 kDa molecular weight (trade name: Hybloom 1000) TM ) 12.5g of hydrolyzed sodium hyaluronate, 2.6kDa of hydrolyzed sodium hyaluronate (trade name: Ximinxiu TMSuper active hyaluronic acid) 50g, hydrolyzed sodium hyaluronate with a molecular weight of 7kDa (trade name: Nano ) 12.5 g, acetylated sodium hyaluronate with a molecular weight of 30 kDa (trade name: Hymagic TM -AcHA) 12.5g, low molecular weight sodium hyaluronate with a molecular weight of 373kDa (trade name: Hybloom TM Low molecular weight sodium hyaluronate) 12.5g.

[0081] The weighed components are stirred evenly to obtain a hyaluronic acid composition.

[0082] Comparative Example 3

[0083] Weigh a 1 kDa molecular weight (trade name: Hybloom 1000) TM ) 12.5g of hydrolyzed sodium hyaluronate, 2.6kDa of hydrolyzed sodium hyaluronate (trade name: Ximinxiu TM Super active hyaluronic acid) 12.5g, hydrolyzed sodium hyaluronate with a molecular weight of 7kDa (trade name: Nano ) 50g, acetylated sodium hyaluronate with a molecular weight of 30kDa (trade name: Hymagic TM -AcHA) 12.5g, low molecular weight sodium hyaluronate with a molecular weight of 373kDa (trade name: Hybloom TM Low molecular weight sodium hyaluronate) 12.5g.

[0084] The weighed components are stirred evenly to obtain a hyaluronic acid composition.

[0085] Comparative Example 4

[0086] Weigh a 1 kDa molecular weight (trade name: Hybloom 1000) TM ) 12.5g of hydrolyzed sodium hyaluronate, 2.6kDa of hydrolyzed sodium hyaluronate (trade name: Ximinxiu TM Super active hyaluronic acid) 12.5g, hydrolyzed sodium hyaluronate with a molecular weight of 7kDa (trade name: Nano ) 12.5g, molecular weight 30kDa acetylated sodium hyaluronate (trade name: Hymagic TM -AcHA) 12.5g, low molecular weight sodium hyaluronate with a molecular weight of 373kDa (trade name: Hybloom TM Low molecular weight sodium hyaluronate) 50g.

[0087] The weighed components are stirred evenly to obtain a hyaluronic acid composition.

[0088] The conditions of the above embodiments and comparative examples are shown in Table 1.

[0089] Table 1 Composition and content of different embodiments and comparative examples (g)

[0090]

[0091]

[0092] Test example

[0093] The 3D full-thickness skin model is an active tissue similar to the structure of human skin that is reconstructed in vitro using normal human skin cells using tissue engineering technology and appropriate culture medium. It can be used to screen active substances. Active substances that are beneficial to the skin can be preliminarily screened out by exploring the effects of active substances on the tissue structure of the skin model.

[0094] Test Example 1

[0095] The efficacy of the compositions was evaluated by assessing their effects on the histological structure of a 3D full-thickness skin model.

[0096] 1. Sample preparation

[0097] The compositions prepared in Examples 1-2 and Comparative Examples 1-4 were dissolved in a certain amount of DMEM culture medium to obtain hyaluronic acid composition-culture medium solutions (Samples 1-6) with a final concentration of 0.2%.

[0098] 2. Construction of 3D full-thickness skin model

[0099] The dermis layer of the 3D full-thickness skin model was constructed by combining collagen with human dermal fibroblasts, and then human keratinocytes were inoculated on the dermis layer to form the epidermis layer, thereby obtaining a complete skin model with a dermis-epidermis structure. Samples 1-6 were then added to treat them, and the control group was not treated at all.

[0100] 3. 3D full-thickness skin model paraffin tissue sections

[0101] The tissue was placed in an embedding box and dehydrated in 4% paraformaldehyde, 70% ethanol-water solution, 80% ethanol-water solution, 90% ethanol-water solution, 95% ethanol-water solution, anhydrous ethanol I, anhydrous ethanol II, methylcyclohexane I, and methylcyclohexane II, respectively, for 1 hour in each tank. After dehydration, the tissue was embedded in paraffin, trimmed, and cut into paraffin sections with a thickness of 5 μm.

[0102] 4. Hematoxylin-eosin staining of 3D full-thickness skin model

[0103] Soak the paraffin sections in methylcyclohexane I, methylcyclohexane II, anhydrous ethanol, 95% ethanol-water solution, 80% ethanol-water solution, 70% ethanol-water solution, and distilled water for 10 minutes for dewaxing and rehydration. After staining in hematoxylin staining solution for 8 minutes, place them in hydrochloric acid-ethanol solution for differentiation for 2 seconds and reblue in ammonia solution for 1 minute. Then stain with eosin solution for 1 minute, and finally dehydrate in 70% ethanol-water solution, 95% ethanol-water solution, anhydrous ethanol, and methylcyclohexane solution, and seal with neutral gum. Take pictures under a microscope. See the results. Figure 1 .

[0104] 5. Results

[0105] When the water content of the skin increases, the skin barrier is strengthened and the skin is in a healthy and active state. The various structures in the skin will also tend to be well-differentiated. Therefore, by evaluating the thickness of the epidermis and stratum corneum of the 3D full-thickness skin model after adding the sample, the effect of the sample on the hydration state of the 3D full-thickness skin model can be judged.

[0106] By adding samples to the structure of the 3D full-thickness skin model ( Figure 1 ) We can see that the embodiment, especially embodiment 1, makes the 3D full-thickness skin model well differentiated compared with the control, the stratum corneum, stratum granulosum, and stratum spinosum are well differentiated, the structure is clear, the epidermis thickness is increased, and the stratum corneum thickness is increased. At the same time, the basal layer structure is complete and the cells are closely arranged. The embodiment, especially embodiment 1, can significantly increase the thickness of the stratum corneum, reduce water loss, improve epidermal hydration, and keep the skin in a healthy state.

[0107] Test Example 2

[0108] The efficacy of the composition was evaluated by assessing the expression levels of HAS2, HABP2, and AQP3 in a 3D full-thickness skin model.

[0109] 1. Sample preparation

[0110] The compositions prepared in Examples 1 and 2 and Comparative Example 1 were dissolved in a certain amount of DMEM culture medium to obtain hyaluronic acid composition-culture medium solutions (Samples 1, 2, and 3) with a final concentration of 0.2%.

[0111] 2. Construction of 3D full-thickness skin model

[0112] The dermis layer of the 3D full-thickness skin model was constructed by combining collagen with human dermal fibroblasts, and then human keratinocytes were inoculated on the dermis layer to form the epidermis layer, thus obtaining a skin model with a complete dermis-epidermis structure. Samples 1, 2, and 3 were then added to treat them, and the control group was not treated at all.

[0113] 3. 3D full-thickness skin model frozen tissue sections

[0114] The 3D full-thickness skin model was placed in an OCT tissue embedding medium and rapidly cooled by adding liquid nitrogen. The embedded tissue block was placed in a freezing microtome and cut into slices with a thickness of 5 μm.

[0115] 4. Immunofluorescence staining of 3D full-thickness skin model ice-cut tissue sections

[0116] The sections were fixed with ice methanol at -20°C, washed three times with PBS, and incubated with blocking solution at room temperature for 1 hour. After incubation, the corresponding primary antibody was added and incubated at 4°C overnight. The next day, the sections were washed three times with PBS, and the corresponding secondary antibody was added and incubated at room temperature in the dark for 1.5 hours. Finally, the sections were stained with DAPI and sealed, and then observed and photographed under a fluorescence microscope. The results are shown in Figure 2 , Figure 3 , Figure 4 .

[0117] 5. Results

[0118] Next, by comparing the expressions of HAS2, HABP2 and AQP3 in the epidermis in Example 1, Example 2 and Comparative Example 1, the effect of the long-term moisturizing on the skin was studied. Figure 2 , 3 In 4, HAS2, HABP2, and AQP3 are staining of target proteins. DAPI is staining of cell nuclei, indicating the cell localization of the 3D full-thickness skin model structure. Merge is the superposition of target protein staining and cell nucleus staining. Figure 2 , 3 , 4 show that HAS2 and AQP3 are expressed on the epidermal cell membrane, and HABP2 is expressed on the epidermal extracellular matrix, which is consistent with the localization reported in the literature.

[0119] Figure 2 It shows that the HAS2 fluorescence expression in the epidermis of the embodiments, especially embodiment 1, is brighter than that in comparative example 1, and the fluorescence expression area in a single cell is larger and more uniform, that is, the HAS2 expression is significant, indicating that embodiment 1 can significantly promote the synthesis and secretion of hyaluronic acid by human epidermal cells.

[0120] Figure 3 The expression of HABP2 in the stratum corneum of Example 1 is also consistent with this result. Compared with Comparative Example 1 and Example 2, the stratum corneum of Example 1 has strong expression of HABP2.

[0121] Figure 4 It shows that the AQP3 fluorescence expression in the epidermis of the examples, especially Example 1, is brighter than that in Comparative Example 1, and the cells located in the basal layer express AQP3 strongly, indicating that Example 1 can significantly promote the expression of AQP3 in human epidermal cells and regulate the hydration balance of the epidermis.

[0122] The above results show that Example 1 can promote epidermal cells to express HAS2, HABP2, and AQP3, thereby promoting epidermal cells to generate and secrete hyaluronic acid, improving the ability of epidermal hydration, and achieving a long-lasting moisturizing effect. By comparing Example 1 and Example 2, it can be seen that the long-lasting moisturizing ability of acetylated sodium hyaluronate by compounding the other four different sodium hyaluronates (Example 1) is significantly greater than that of acetylated sodium hyaluronate (Example 2) with the same total concentration.

Claims

1. A composition for promoting long-term moisturizing of the skin, characterized in that: The composition consists of the following ingredients: Acetylated hyaluronic acid or its salt, hydrolyzed hyaluronic acid or its salt, and low molecular weight hyaluronic acid or its salt, The molecular weight of the acetylated hyaluronic acid or its salt is 30k-100kDa, The molecular weight of the low molecular weight hyaluronic acid or its salt is 200k-400kDa, The hydrolyzed hyaluronic acid or its salt is composed of a first hydrolyzed hyaluronic acid or its salt having a molecular weight of less than 1 kDa, a second hydrolyzed hyaluronic acid or its salt having a molecular weight of 1.1 kDa to 5 kDa, and a third hydrolyzed hyaluronic acid or its salt having a molecular weight of 5.1 kDa to 10 kDa. Wherein, in the composition, the mass ratio of the acetylated hyaluronic acid or its salt is 40%-60%, The mass ratio of the low molecular weight hyaluronic acid or its salt is 10%-15%, The mass ratio of the first hydrolyzed hyaluronic acid or its salt is 10%-15%, the mass ratio of the second hydrolyzed hyaluronic acid or its salt is 10%-15%, and the mass ratio of the third hydrolyzed hyaluronic acid or its salt is 10%-15%.

2. The composition according to claim 1, characterized in that In the composition, the mass ratio of the hydrolyzed hyaluronic acid or its salt is 30%-45%.

3. Use of the composition according to claim 1 or 2 in the preparation of a composition for promoting epidermal cells to express HAS2, HABP2 and / or AQP3.

4. A moisturizing product comprising the composition according to claim 1 or 2.

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