A water-in-oil nanoemulsion and a preparation method thereof

Abstract

The present invention provides a water-in-oil nanoemulsion, comprising an oil phase, an aqueous phase, and an emulsifier, wherein the aqueous phase comprises a hyaluronic acid composition. The present invention adds the hyaluronic acid composition to the water-in-oil nanoemulsion formula. As the internal phase of the nanoemulsion, the hyaluronic acid composition can act as a reservoir in the dermis, enhancing the retention of active ingredients in the skin. Combined with the small particle size and easy permeability of the nanoemulsion, the active ingredients can be delivered to the deeper layers of the skin, which are more hydrophilic, synergistically enhancing transdermal absorption. At the same time, the water-in-oil nanoemulsion can encapsulate water-soluble active ingredients with poor stability, enhancing stability and resolving irritation issues.

Description

Technical Field

[0001] The present invention belongs to the field of biomedicine, and in particular relates to a water-in-oil nanoemulsion and a preparation method thereof. Background Art

[0002] Nanoemulsions are transparent or translucent systems composed of an aqueous phase, an oil phase, a surfactant, and a co-surfactant. Compared to traditional emulsions, most nanoemulsions effectively prevent delamination, sedimentation, and aggregation, leading to their widespread application in petroleum, food, and cosmetics.

[0003] In the practical application of skin care products, active ingredients must penetrate specific skin sites to exert their efficacy. The transdermal ability of active ingredients is related to their molecular weight. Generally, ingredients with a molecular weight greater than 500 are less likely to penetrate the skin. Nanoformulations such as nanoemulsions, microemulsions, and liposomes can be used to promote the transdermal delivery of active ingredients. Furthermore, the human stratum corneum is lipophilic, making oil-soluble ingredients more easily accessible to the skin than water-soluble active ingredients. Water-in-oil nanoemulsions, with their oily external phase, have a greater affinity for the skin, making them an effective means of promoting the absorption of active ingredients.

[0004] However, conventional water-in-oil formulations only enhance the absorption of oil-soluble ingredients in the external phase, without significantly promoting the absorption of water-soluble active ingredients in the internal aqueous phase. The present invention incorporates a sodium hyaluronate composition with a permeation-promoting effect into the internal phase. This composition itself is transdermal and can carry water-soluble active ingredients into the skin, promoting absorption. Therefore, the water-in-oil nanoemulsion obtained by the present invention has good stability and excellent transdermal absorption, which is of great significance for enhancing skin care efficacy. Summary of the Invention

[0005] In view of the above problems existing in the prior art, the present invention provides a water-in-oil nanoemulsion and a preparation method thereof.

[0006] Specifically, the present invention relates to the following aspects:

[0007] 1. A water-in-oil nanoemulsion, characterized in that the nanoemulsion comprises an oil phase, an aqueous phase, and an emulsifier, wherein the aqueous phase comprises a hyaluronic acid composition.

[0008] 2. The nanoemulsion according to item 1, characterized in that the mass percentage of the oil phase in the nanoemulsion is 5-80%, preferably 10-60%, the mass percentage of the hyaluronic acid composition is 0.01%-10%, preferably 0.01%-1%, further preferably 0.01%-0.5%, and the mass percentage of the emulsifier is 0.01-20%, preferably 0.5-10%.

[0009] 3. The nanoemulsion according to item 1, characterized in that the hyaluronic acid composition comprises medium molecular weight hyaluronic acid or a salt thereof, acetylated hyaluronic acid or a salt thereof and hydrolyzed hyaluronic acid or a salt thereof, preferably, the molecular weight of the medium molecular weight hyaluronic acid or a salt thereof is 200-700 kDa, preferably 200-400 kDa, further preferably, the molecular weight of the acetylated hyaluronic acid or a salt thereof is 20-40 kDa, and further preferably, the molecular weight of the hydrolyzed hyaluronic acid or a salt thereof is 3-18 kDa.

[0010] 4. The nanoemulsion according to claim 3, characterized in that, in terms of mass percentage in the hyaluronic acid composition, the content of the medium molecular weight hyaluronic acid or its salt is 20-60%, preferably 25-40%, the content of the acetylated hyaluronic acid or its salt is 10-50%, preferably 20-35%, and the content of the hydrolyzed hyaluronic acid or its salt is 30-70%, preferably 40-55%.

[0011] 5. The nanoemulsion according to item 1, characterized in that the aqueous phase further comprises a water-soluble active ingredient.

[0012] 6. The nanoemulsion according to item 1, characterized in that the water-soluble active ingredient is one or more of a whitening ingredient, a moisturizing ingredient, and an anti-aging ingredient, wherein the whitening ingredient is selected from one or more of nonapeptide-1, tranexamic acid, niacinamide, thiotaurine, potassium 4-methoxysalicylate, arbutin, glutathione, and ethylbisiminomethylguaiacol manganese chloride (EUK-134); the moisturizing ingredient is selected from one or more of ectoine, carnosine, and β-glucan; and the anti-aging ingredient is selected from one or more of polydeoxyribonucleotides (PDRN), γ-aminobutyric acid, and collagen.

[0013] 7. The nanoemulsion according to claim 1, wherein the oil phase is selected from one or more of squalane, rice bran oil, caprylic / capric triglyceride, coconut oil, and isononyl isononanoate (ISIS).

[0014] 8. The nanoemulsion according to claim 1, wherein the emulsifier is selected from one or more of polyglyceryl-2 dipolyhydroxystearate, polyglyceryl-2 oleate, polyglyceryl-3 diisostearate, cetyl PEG / PPG-10 / 1 polydimethylsiloxane, PEG40 hydrogenated castor oil, sorbitan sesquioleate, and PEG-20 glyceryl tristearate, preferably a mixture of polyglyceryl-3 diisostearate and PEG 40 hydrogenated castor oil, and further preferably, the mass ratio of polyglyceryl-3 diisostearate to PEG 40 hydrogenated castor oil is 1:4-2:3.

[0015] 9. The method for preparing the nanoemulsion according to any one of items 1 to 8, characterized in that the method comprises the following steps:

[0016] Dissolve the emulsifier in liquid oil to form an oil phase.

[0017] dissolving the hyaluronic acid composition in water to form an aqueous phase,

[0018] Mixing and homogenizing the oil phase and the water phase at the same temperature to obtain a nanoemulsion;

[0019] Preferably, the step of dissolving the hyaluronic acid composition in water to form an aqueous phase comprises:

[0020] The hyaluronic acid composition and the water-soluble active ingredient are dissolved in water to form an aqueous phase.

[0021] 10. A cosmetic comprising the nanoemulsion according to any one of items 1 to 8.

[0022] 11. Use of a hyaluronic acid composition for increasing the stability and / or permeability of a nanoemulsion, wherein the hyaluronic acid composition comprises medium molecular weight hyaluronic acid or a salt thereof, acetylated hyaluronic acid or a salt thereof, and hydrolyzed hyaluronic acid or a salt thereof;

[0023] Preferably, the molecular weight of the medium molecular weight hyaluronic acid or its salt is 200-700 kDa, preferably 200-400 kDa, the molecular weight of the acetylated hyaluronic acid or its salt is 20-40 kDa, and the molecular weight of the hydrolyzed hyaluronic acid or its salt is 3-18 kDa;

[0024] Further preferably, in terms of mass percentage in the hyaluronic acid composition, the content of the medium molecular weight hyaluronic acid or its salt is 20-60%, preferably 25-40%, the content of the acetylated hyaluronic acid or its salt is 10-50%, preferably 20-35%, and the content of the hydrolyzed hyaluronic acid or its salt is 30-70%, preferably 40-55%.

[0025] The present invention adds a hyaluronic acid composition to the water-in-oil nanoemulsion formula. As the internal phase of the nanoemulsion, it can act as a reservoir in the dermis and enhance the retention of active ingredients in the skin. Combined with the characteristics of the nanoemulsion's small particle size and easy penetration, the active ingredients can be delivered to the deeper layers of the skin with better hydrophilicity, synergistically enhancing the transdermal absorption effect. At the same time, the water-in-oil nanoemulsion can encapsulate water-soluble active ingredients with poor stability, thereby enhancing stability and solving the problem of irritation. DETAILED DESCRIPTION

[0026] The present invention is further described below with reference to the 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.

[0027] Unless otherwise defined, technical and scientific terms used in this specification have the same meaning as those commonly understood by those skilled in the art. Although methods and materials similar or identical to those described herein can be used in experiments or practical applications, the materials and methods are described herein below. In the event of a conflict, the present specification, including definitions, will prevail. In addition, the materials, methods, and examples are illustrative only and not restrictive. The present invention is further described below with reference to specific examples, but is not intended to limit the scope of the invention.

[0028] The present invention provides a water-in-oil nanoemulsion, wherein the nanoemulsion comprises an oil phase, an aqueous phase encapsulating the oil phase, and an emulsifier, wherein the aqueous phase comprises a hyaluronic acid composition.

[0029] Furthermore, the mass percentage of the oil phase in the nanoemulsion is 5-80%, for example, 5%, 6%, 7%, 8%, 9%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, preferably 10-60%. The mass percentage of the hyaluronic acid composition is 0.01%-10%, for example, 0.01%, 0.1%, 0.5%, 1%, 2%, 3%, 4%, 5%, 6%, 7%, 8%, 9%, 10%, preferably 0.01%-1%, more preferably 0.01%-0.5%. The mass percentage of the emulsifier is 0.01-20%, 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%, 18%, 19%, 20%, preferably 0.5-10%.

[0030] The hyaluronic acid composition of the present invention may refer to any composition comprising various forms of hyaluronic acid or its salts.

[0031] In a specific embodiment, the hyaluronic acid composition includes medium molecular weight hyaluronic acid or its salt, acetylated hyaluronic acid or its salt, and hydrolyzed hyaluronic acid or its salt.

[0032] In a specific embodiment, the molecular weight of the medium molecular weight hyaluronic acid or its salt is 200-700 kDa, preferably 200-400 kDa, for example, 200 kDa, 300 kDa, 400 kDa, 500 kDa, 600 kDa, or 700 kDa.

[0033] Acetylated Hyaluronate (AcHA) is obtained by acetylation of sodium hyaluronate. The introduction of acetyl groups makes hyaluronic acid lipophilic.

[0034] In a specific embodiment, the molecular weight of the acetylated hyaluronic acid or its salt is 20-40 kDa, for example, 20 kDa, 25 kDa, 30 kDa, 35 kDa, or 40 kDa.

[0035] Hydrolyzed hyaluronic acid or its salts are low-molecular-weight hyaluronic acids obtained by enzymatic hydrolysis of hyaluronic acid or its salts. By controlling the enzymatic hydrolysis conditions, hydrolyzed hyaluronic acid of varying molecular weights can be obtained. Research has found that the physiological effects of hyaluronic acid are closely related to its molecular weight, with hyaluronic acid of varying molecular weights exhibiting distinct biological activities. Hyaluronic acid with a lower molecular weight exhibits immune activation, promotes endothelial cell proliferation, and reverses multidrug resistance in tumor cells. Furthermore, due to its lower molecular weight, hydrolyzed hyaluronic acid can penetrate deep into the epidermis.

[0036] In a specific embodiment, the molecular weight of the hydrolyzed hyaluronic acid or its salt is 3-18 kDa, for example, 3 kDa, 4 kDa, 5 kDa, 6 kDa, 7 kDa, 8 kDa, 9 kDa, 10 kDa, 11 kDa, 12 kDa, 13 kDa, 14 kDa, 15 kDa, 16 kDa, 17 kDa, or 18 kDa.

[0037] Furthermore, in the hyaluronic acid composition, the content of the medium molecular weight hyaluronic acid or its salt is 20-60% by mass percentage in the hyaluronic acid composition, for example, it can be 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, preferably 25-40%. The content of the acetylated hyaluronic acid or its salt is 10-50%, for example, it can be 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%. Preferably, it is 20-35%. The content of the hydrolyzed hyaluronic acid or its salt is 30-70%, for example, it can be 30%, 35%, 40%, 45%, 50%, 50%, 55%, 60%, 65%, 70%, preferably 40-55%.

[0038] In a specific embodiment, the hyaluronic acid composition consists of medium molecular weight hyaluronic acid or its salt, acetylated hyaluronic acid or its salt, and hydrolyzed hyaluronic acid or its salt.

[0039] In a specific embodiment, in the hyaluronic acid composition, the content of the medium molecular weight hyaluronic acid or its salt is 25-40%, the content of the acetylated hyaluronic acid or its salt is 20-35%, and the content of the hydrolyzed hyaluronic acid or its salt is 40-55%, calculated as a percentage by mass in the hyaluronic acid composition.

[0040] In a specific embodiment, in the hyaluronic acid composition, the content of the medium molecular weight hyaluronic acid or its salt is 30%, the content of the acetylated hyaluronic acid or its salt is 25%, and the content of the hydrolyzed hyaluronic acid or its salt is 45%, calculated as a percentage by mass in the hyaluronic acid composition.

[0041] In a specific embodiment, in the hyaluronic acid composition, the content of the medium molecular weight hyaluronic acid or its salt is 20%, the content of the acetylated hyaluronic acid or its salt is 10%, and the content of the hydrolyzed hyaluronic acid or its salt is 70%, calculated as a percentage by mass in the hyaluronic acid composition.

[0042] In a specific embodiment, in the hyaluronic acid composition, the content of the medium molecular weight hyaluronic acid or its salt is 20%, the content of the acetylated hyaluronic acid or its salt is 50%, and the content of the hydrolyzed hyaluronic acid or its salt is 30%, calculated as a percentage by mass in the hyaluronic acid composition.

[0043] In a specific embodiment, in the hyaluronic acid composition, the content of the medium molecular weight hyaluronic acid or its salt is 60%, the content of the acetylated hyaluronic acid or its salt is 10%, and the content of the hydrolyzed hyaluronic acid or its salt is 30%, calculated by mass percentage in the hyaluronic acid composition.

[0044] In a specific embodiment, the oil phase is selected from one or more of squalane, rice bran oil, caprylic / capric triglyceride, coconut oil, and isononyl isononanoate (ISIS).

[0045] In a specific embodiment, the emulsifier is selected from one or more of polyglyceryl-2 dipolyhydroxystearate, polyglyceryl-2 oleate, polyglyceryl-3 diisostearate, cetyl PEG / PPG-10 / 1 polydimethicone, PEG 40 hydrogenated castor oil, sorbitan sesquioleate, and PEG-20 glyceryl tristearate, preferably a mixture of polyglyceryl-3 diisostearate and PEG 40 hydrogenated castor oil. Further preferably, the mass ratio of polyglyceryl-3 diisostearate to PEG 40 hydrogenated castor oil is 1:4-2:3, for example, 1:4, 1:3, 1:2, 1:1, 2:3, etc.

[0046] The water-in-oil nanoemulsion of the present invention may further contain a water-soluble active ingredient in the aqueous phase, wherein the water-soluble active ingredient is one or more of a whitening ingredient, a moisturizing ingredient, and an anti-aging ingredient.

[0047] The whitening functional ingredient is selected from one or more of nonapeptide-1, tranexamic acid, niacinamide, thiotaurine, potassium 4-methoxysalicylate, arbutin, glutathione, and ethylbisiminomethylguaiacol manganese chloride (EUK-134); the moisturizing functional ingredient is selected from one or more of ectoine, carnosine, and β-glucan; and the anti-aging functional ingredient is selected from one or more of polydeoxyribonucleotide (PDRN), γ-aminobutyric acid, and collagen.

[0048] The present invention also provides a method for preparing the nanoemulsion, which comprises the following steps:

[0049] Dissolve the emulsifier in liquid oil to form an oil phase.

[0050] dissolving the hyaluronic acid composition in water to form an aqueous phase,

[0051] The oil phase and the water phase are mixed and homogenized at the same temperature to obtain a nanoemulsion.

[0052] Wherein, the step of dissolving the hyaluronic acid composition in water to form an aqueous phase may further include:

[0053] The hyaluronic acid composition and the water-soluble active ingredient are dissolved in water to form an aqueous phase.

[0054] Wherein, the composition and content of the hyaluronic acid composition, water-soluble active ingredients, etc. are as described above.

[0055] The present invention also provides a cosmetic comprising the nanoemulsion.

[0056] The present invention also provides use of a hyaluronic acid composition in increasing the stability and / or permeability of a nanoemulsion, wherein the hyaluronic acid composition comprises medium molecular weight hyaluronic acid or a salt thereof, acetylated hyaluronic acid or a salt thereof, and hydrolyzed hyaluronic acid or a salt thereof.

[0057] In a specific embodiment, the molecular weight of the medium molecular weight hyaluronic acid or its salt is 200-700 kDa, preferably 200-400 kDa, the molecular weight of the acetylated hyaluronic acid or its salt is 20-40 kDa, and the molecular weight of the hydrolyzed hyaluronic acid or its salt is 3-18 kDa.

[0058] In a specific embodiment, the content of the medium molecular weight hyaluronic acid or its salt is 20-60%, preferably 25-40%, the content of the acetylated hyaluronic acid or its salt is 10-50%, preferably 20-35%, and the content of the hydrolyzed hyaluronic acid or its salt is 30-70%, preferably 40-55%.

[0059] The present invention uses a hyaluronic acid composition, particularly a composition comprising medium molecular weight hyaluronic acid, acetylated hyaluronic acid, and hydrolyzed hyaluronic acid, and a specific emulsifier to prepare a nanoemulsion, so that the nanoemulsion has high stability at both low and high temperatures. Furthermore, the hyaluronic acid composition, as the internal phase of the nanoemulsion, can act as a reservoir in the dermis, enhancing the retention of active ingredients in the skin. Combined with the nanoemulsion's small particle size and easy permeability, the active ingredients can be delivered to the deeper layers of the skin, which are more hydrophilic, synergistically enhancing transdermal absorption. At the same time, the water-in-oil nanoemulsion can encapsulate water-soluble active ingredients with poor stability, enhancing stability and addressing irritation issues.

[0060] Example

[0061] Unless otherwise specified, the experimental methods used in the following examples are all conventional methods.

[0062] Unless otherwise specified, the materials and reagents used in the following examples can be obtained from commercial sources, and the sodium hyaluronate used was purchased from Bloomage Biotech Co., Ltd.

[0063] Example 1

[0064] (1) 2 g of polyglyceryl-3 diisostearate and 8 g of PEG40 hydrogenated castor oil were added to 55 g of squalane and heated to 80° C. to obtain an oil phase. 0.2 g of a hyaluronic acid composition (wherein, in the hyaluronic acid composition, the content of medium molecular weight sodium hyaluronate is 30%, and the molecular weight is 200-400 kDa, the content of acetylated sodium hyaluronate is 25%, and the molecular weight is 20-30 kDa, and the content of hydrolyzed sodium hyaluronate is 45%, and the molecular weight is 3-10 kDa) and 3 g of γ-aminobutyric acid were dissolved in 31.8 g of ultrapure water and heated to 80° C. to dissolve and obtain an aqueous phase.

[0065] (2) heating the two phases to 80°C, mixing and stirring them uniformly, and homogenizing them with a homogenizer to obtain a crude emulsion;

[0066] (3) The obtained crude emulsion was placed in a high-pressure homogenizer and homogenized twice, each time for 5 minutes to obtain a nanoemulsion.

[0067] Examples 2-5

[0068] Examples 2-5 differ from Example 1 only in the amounts of the hyaluronic acid composition and ultrapure water added. Specifically, the amount of the hyaluronic acid composition added in Example 2 was 0.1 g, the amount of the hyaluronic acid composition added in Example 3 was 0.5 g, the amount of the hyaluronic acid composition added in Example 4 was 1 g, and the amount of the hyaluronic acid composition added in Example 5 was 0.005 g. The specific amounts of each component are shown in Table 1.

[0069] Examples 6-12

[0070] Examples 6-12 differ from Example 1 only in the composition of the hyaluronic acid composition. Example 6 differs from Example 1 in the content of medium molecular weight sodium hyaluronate, acetylated sodium hyaluronate, and hydrolyzed sodium hyaluronate: the content of medium molecular weight sodium hyaluronate is 20%, with a molecular weight of 200-400 kDa, the content of acetylated sodium hyaluronate is 10%, with a molecular weight of 20-30 kDa, and the content of hydrolyzed sodium hyaluronate is 70%. Example 7 differs from Example 1 in the content of medium molecular weight sodium hyaluronate, acetylated sodium hyaluronate, and hydrolyzed sodium hyaluronate: the content of medium molecular weight sodium hyaluronate is 20%, with a molecular weight of 200-400 kDa, the content of acetylated sodium hyaluronate is 50%, with a molecular weight of 20-30 kDa, and the content of hydrolyzed sodium hyaluronate is 30%. Example 8 differs from Example 1 in the content of medium molecular weight sodium hyaluronate, acetylated sodium hyaluronate, and hydrolyzed sodium hyaluronate. The content of medium molecular weight sodium hyaluronate is 60% with a molecular weight of 200-400 kDa, the content of acetylated sodium hyaluronate is 10% with a molecular weight of 20-30 kDa, and the content of hydrolyzed sodium hyaluronate is 30%. The hyaluronic acid composition in Example 9 contains only medium molecular weight sodium hyaluronate with a molecular weight of 200-400 kDa. The hyaluronic acid composition in Example 10 contains only acetylated sodium hyaluronate with a molecular weight of 20-30 kDa. The hyaluronic acid composition in Example 11 contains only hydrolyzed sodium hyaluronate with a molecular weight of 3-10 kDa. Example 12 differs from Example 1 in the content of medium molecular weight sodium hyaluronate, acetylated sodium hyaluronate, and hydrolyzed sodium hyaluronate. The content of medium molecular weight sodium hyaluronate is 60%, with a molecular weight of 200-400 kDa; the content of acetylated sodium hyaluronate is 15%, with a molecular weight of 20-30 kDa; and the content of hydrolyzed sodium hyaluronate is 25%. The specific content of each component is shown in Table 1.

[0071] Examples 13-16

[0072] Examples 13-16 differ from Example 1 only in the composition of the emulsifier. In Example 13, the emulsifiers are 4g of polyglyceryl-3 diisostearate and 6g of PEG40 hydrogenated castor oil; in Example 14, the emulsifier is 10g of polyglyceryl-3 diisostearate; in Example 15, the emulsifier is 10g of PEG40 hydrogenated castor oil; and in Example 16, the emulsifier is 6g of polyglyceryl-3 diisostearate and 4g of PEG40 hydrogenated castor oil. The specific component contents are shown in Table 1.

[0073] Comparative Example 1

[0074] The only difference between Comparative Example 1 and Example 1 is that the aqueous phase does not contain the hyaluronic acid composition. That is, when preparing the aqueous phase in step 1, no hyaluronic acid composition is added, but only 3 g of γ-aminobutyric acid is added.

[0075] Comparative Example 2

[0076] Comparative Example 2 is a solution obtained by directly dissolving 3 g of γ-aminobutyric acid in 97 g of ultrapure water.

[0077] Table 1

[0078]

[0079]

[0080]

[0081]

[0082] Test example

[0083] Test Example 1 Stability Test

[0084] High temperature test: The nanoemulsions prepared in the examples and comparative examples were placed in a suitable clean container as test samples and placed at 55°C for 10 days. Samples were taken on the 5th and 10th days to observe the stratification of the samples.

[0085] Low temperature test: Place the sample to be tested in a suitable clean container and place it at -20℃ for 5 days. Take samples on the 5th day and observe the stratification of the samples.

[0086] Centrifugal stability: Fill a centrifuge tube to approximately 2 / 3 of its height with the sample to be tested and secure it with a stopper. Place the tube in an incubator pre-adjusted to 38°C for 1 hour. Immediately transfer the tube to a centrifuge at a speed of 2000 rpm. Spin for 30 minutes and remove the tube for observation.

[0087] The test results are shown in Table 2 below. From the data in Table 2, it can be seen that the addition of the hyaluronic acid composition has an important influence on the stability of the emulsion. The high-temperature stability of the oil-in-water nanoemulsion with the addition of the hyaluronic acid composition is significantly improved; the amount of different emulsifiers in the system has an important influence on the stability of the system.

[0088] Table 2

[0089]

[0090]

[0091] Test Example 2: Penetration Test

[0092] The samples of Examples 1-4, 6-8, 13 and the comparative example without stability issues were subjected to Franz diffusion cell permeation experiments to explore the penetration-enhancing effect of the hyaluronic acid composition on the active substance.

[0093] Excised pig skin was fixed between the supply and receiving reservoirs of a Phoenix DB-6 transdermal testing system. Approximately 15 ml of PBS buffer solution was added to the sampling tube, depending on the liquid level. Air was expelled to ensure close contact between the dermis and the receiving solution. 49 μL of the test sample was then applied to the skin surface. Using a stainless steel stirring rod, the sample was evenly spread radially from the center of the skin toward the periphery. Three replicates of each sample were prepared, maintained in a constant temperature water bath at (32 ± 1)°C and stirred at 300 rpm / min. After 20 hours of diffusion, samples from the skin surface, the skin interior, and the receiving solution were collected and analyzed by high-performance liquid chromatography to determine in vitro transdermal efficiency.

[0094] in,

[0095] Relative retention (%) = the amount of test substance detected in the skin / the amount of sample loaded * 100%

[0096] Skin retention ratio = the amount of the test substance detected in the skin / the amount of the comparative example 2 detected in the skin

[0097] Table 3 Skin retention ratio test results

[0098]

[0099] As can be seen from Table 3, at 20 hours, the retention of the active substance in the skin was increased in all the Example groups compared to the Comparative Example group. Example 1 showed the most significant improvement, with retention increasing by 18%. This demonstrates that the hyaluronic acid composition can promote the retention of the active substance, thereby further promoting transdermal absorption of the active substance by the skin.

[0100] Test Example 3 Patch test

[0101] Open the packaging of the patch tester and add 0.025 mL or 0.025 g of the prepared test samples from Examples 1, 2, 5, Comparative Examples 1, and 2 to the chamber. Apply the patch tester to the curved side of the subject's forearm, gently pressing with the palm of your hand to evenly adhere the patch to the skin for 24 hours.

[0102] 30 minutes, 24 hours, and 48 hours after removing the spot tester, the reaction results were observed and recorded according to Table 4. The results are shown in Table 5. As can be seen from the results in the table, compared with the group without the hyaluronic acid composition, the irritation was reduced after adding the hyaluronic acid composition. Compared with the pure GABA solution, the addition of this active ingredient to the water-in-oil emulsion also significantly reduced its irritation.

[0103] Table 4 Skin reaction grading standards for skin occlusive patch test

[0104]

[0105] Table 5 Patch test results

[0106] Group Experimental results Example 1 All 20 people were negative Example 2 All 20 people were negative Example 5 One person had a grade 1 reaction, and the rest were negative Comparative Example 1 Two people had grade 1 reactions, and the rest were negative Comparative Example 2 6 people had grade 1 reactions, and the rest were negative

Claims

1. A water-in-oil nanoemulsion, characterized in that The nanoemulsion comprises an oil phase, an aqueous phase, and an emulsifier, wherein the aqueous phase comprises a hyaluronic acid composition; The hyaluronic acid composition comprises medium molecular weight hyaluronic acid or a salt thereof, acetylated hyaluronic acid or a salt thereof, and hydrolyzed hyaluronic acid or a salt thereof, wherein the molecular weight of the medium molecular weight hyaluronic acid or a salt thereof is 200-700 kDa, the molecular weight of the acetylated hyaluronic acid or a salt thereof is 20-40 kDa, and the molecular weight of the hydrolyzed hyaluronic acid or a salt thereof is 3-18 kDa; Calculated by mass percentage in the hyaluronic acid composition, the content of the medium molecular weight hyaluronic acid or its salt is 20-60%, the content of the acetylated hyaluronic acid or its salt is 10-50%, and the content of the hydrolyzed hyaluronic acid or its salt is 30-70%; The emulsifier is selected from a mixture of polyglyceryl-3 diisostearate and PEG 40 hydrogenated castor oil, and the mass ratio of polyglyceryl-3 diisostearate to PEG 40 hydrogenated castor oil is 1:4-2:3; The mass percentage of the oil phase in the nanoemulsion is 5-80%, the mass percentage of the hyaluronic acid composition is 0.01%-10%, and the mass percentage of the emulsifier is 0.01-20%.

2. The nanoemulsion according to claim 1, wherein The mass percentage of the oil phase in the nanoemulsion is 10-60%.

3. The nanoemulsion according to claim 1, wherein The mass percentage of the hyaluronic acid composition is 0.01%-1%.

4. The nanoemulsion according to claim 1, wherein The mass percentage of the hyaluronic acid composition in the nanoemulsion is 0.01%-0.5%.

5. The nanoemulsion according to claim 1, wherein The mass percentage of the emulsifier is 0.5-10%.

6. The nanoemulsion according to claim 1, characterized in that The molecular weight of the medium molecular weight hyaluronic acid or its salt is 200-400 kDa.

7. The nanoemulsion according to claim 1, wherein Calculated by mass percentage in the hyaluronic acid composition, the content of the medium molecular weight hyaluronic acid or its salt is 25-40%.

8. The nanoemulsion according to claim 1, wherein Calculated by mass percentage in the hyaluronic acid composition, the content of the acetylated hyaluronic acid or its salt is 20-35%.

9. The nanoemulsion according to claim 1, characterized in that Calculated by mass percentage in the hyaluronic acid composition, the content of the hydrolyzed hyaluronic acid or its salt is 40-55%.

10. The nanoemulsion according to claim 1, characterized in that The aqueous phase also includes water-soluble active ingredients.

11. The nanoemulsion according to claim 10, characterized in that The water-soluble active ingredient is one or more of a whitening ingredient, a moisturizing ingredient, and an anti-aging ingredient, wherein the whitening ingredient is selected from one or more of nonapeptide-1, tranexamic acid, niacinamide, thiotaurine, potassium 4-methoxysalicylate, arbutin, glutathione, and ethylbisiminomethylguaiacol manganese chloride (EUK-134); the moisturizing ingredient is selected from one or more of ectoine, carnosine, and β-glucan; and the anti-aging ingredient is selected from one or more of polydeoxyribonucleotides (PDRN), γ-aminobutyric acid, and collagen.

12. The nanoemulsion according to claim 1, characterized in that The oil phase is selected from one or more of squalane, rice bran oil, caprylic / capric triglyceride, coconut oil, and isononyl isononanoate (ISIS).

13. The method for preparing the nanoemulsion according to any one of claims 1 to 12, characterized in that: The preparation method comprises the following steps: Dissolve the emulsifier in liquid oil to form an oil phase. dissolving the hyaluronic acid composition in water to form an aqueous phase, The oil phase and the water phase are mixed and homogenized at the same temperature to obtain a nanoemulsion.

14. The method for preparing the nanoemulsion according to claim 13, wherein The step of dissolving the hyaluronic acid composition in water to form an aqueous phase comprises: The hyaluronic acid composition and the water-soluble active ingredient are dissolved in water to form an aqueous phase.

15. A cosmetic comprising the nanoemulsion according to any one of claims 1 to 12.

16. Use of a hyaluronic acid composition for increasing the stability of a water-in-oil nanoemulsion, wherein the hyaluronic acid composition comprises medium molecular weight hyaluronic acid or a salt thereof, acetylated hyaluronic acid or a salt thereof, and hydrolyzed hyaluronic acid or a salt thereof; The molecular weight of the medium molecular weight hyaluronic acid or its salt is 200-700 kDa, the molecular weight of the acetylated hyaluronic acid or its salt is 20-40 kDa, and the molecular weight of the hydrolyzed hyaluronic acid or its salt is 3-18 kDa; Calculated by mass percentage in the hyaluronic acid composition, the content of the medium molecular weight hyaluronic acid or its salt is 20-60%, the content of the acetylated hyaluronic acid or its salt is 10-50%, and the content of the hydrolyzed hyaluronic acid or its salt is 30-70%.

17. The use according to claim 16, characterized in that The molecular weight of the medium molecular weight hyaluronic acid or its salt is 200-400 kDa.

18. The use according to claim 16, characterized in that Calculated by mass percentage in the hyaluronic acid composition, the content of the medium molecular weight hyaluronic acid or its salt is 25-40%.

19. The use according to claim 16, characterized in that Calculated by mass percentage in the hyaluronic acid composition, the content of the acetylated hyaluronic acid or its salt is 20-35%.

20. The use according to claim 16, characterized in that Calculated by mass percentage in the hyaluronic acid composition, the content of the hydrolyzed hyaluronic acid or its salt is 40-55%.

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