Whitening, freckle-removing and face-beautifying liquid and preparation method thereof

Abstract

The invention discloses a whitening, freckle-removing and face-beautifying liquid. The whitening, freckle-removing and face-beautifying liquid is prepared from the following components: nicotinamide, glutathione, ascorbyl tetraisopalmitate, 3-o-ethyl ascorbic acid, papain, 4-hydroxyethyl piperazine ethanesulfonic acid, a citrus peel extract, tetrahydrocurcumin, ascorbyl tetraisopalmitate and the like. The invention also provides a preparation method of the whitening, freckle-removing and face-beautifying liquid. The invention has the following beneficial effects: with the assistance of the specific process, the components have a synergistic effect, and by inhibiting tyrosinase, lightening melanin, promoting metabolism and renewal of epidermis, blocking melanin upward movement, improving intercellular space, improving the white brightness and luster of skin and lightening melanin, the whitening, spot-fading and skin-brightening effects are excellent, and the whitening, spot-fading and skin-brightening effects are good. The moisturizing cream is high in safety, remarkable in moisturizing effect, low in cost and suitable for large-scale production and use of vast user groups.

Description

Technical Field

[0001] This invention relates to the field of cosmetic technology, and in particular to a whitening and freckle-removing lotion and its preparation method. Background Technology

[0002] With the improvement of living standards and the diversification of aesthetic concepts, consumers' demand for skin whitening and even skin tone is becoming increasingly prominent. Pigmentation problems such as age spots and dullness are mainly caused by abnormal melanin metabolism, and their formation is influenced by a variety of internal and external factors, including endocrine disorders, ultraviolet radiation, mental stress, skin inflammation, and environmental pollution. Melanin is produced in melanocytes through a series of oxidation and polymerization reactions catalyzed by tyrosinase. Its metabolic process involves the branching conversion of dopaquinone into eumelanin and pheomelanin, a process regulated by various enzymes and genes.

[0003] When skin is exposed to ultraviolet radiation and external pollution for extended periods, it is prone to excessive accumulation of reactive oxygen species, leading to oxidative stress, lipid peroxidation, protein denaturation, and DNA damage. This accelerates skin aging and triggers various skin problems. While existing antioxidant cosmetics possess some ability to scavenge free radicals, they generally suffer from high concentrations, high formulation costs, low transdermal efficiency, and poor stability. These limitations not only restrict their antioxidant effects but may also cause skin irritation or allergic reactions. Furthermore, their high cost restricts their target audience.

[0004] Whitening methods mainly include inhibiting tyrosinase activity, blocking melanin synthesis signals, accelerating keratinocyte metabolism, and providing antioxidant, nourishing, and anti-allergic effects. However, most existing whitening and brightening products rely on chemically synthesized ingredients, such as retinol and resorcinol. While these have some whitening and brightening effects, they are highly irritating, and long-term use may lead to skin sensitivity, dryness, or even allergies. Most existing cosmetic whitening combinations have relatively singular effects, with ingredients having the same function piled up in the formula without a complete and systematic synergy to achieve optimal results. Furthermore, these formulas contain large amounts of preservatives and additives that are harmful to the skin, which may have negative effects on skin health with long-term use. Although some formulas use natural ingredients, the whitening and brightening effects are not significant, and the cost is high. Besides ingredient selection, the poor efficacy of existing cosmetics is largely due to the neglect of penetration in most formulas. Without good penetrating ingredients, the added active ingredients remain on the skin's surface and cannot be absorbed, resulting in waste and limited effectiveness. Existing cosmetic formulas lack soothing ingredients, which may cause users to experience redness, peeling, and rashes.

[0005] Chinese invention patent application CN 111956557 A discloses a whitening and freckle-removing lotion, cosmetic, and its preparation method, all possessing whitening and freckle-removing effects. The whitening and freckle-removing lotion and cosmetic can prevent melanin aggregation and transfer, reduce and depigment melanin; inhibit tyrosinase activity, and scavenge oxygen free radicals to alter the melanin production pathway; thereby preventing melanin production and further achieving a whitening and freckle-removing effect. However, this technology is ineffective in promoting skin metabolism, the formula has a relatively simple function, its application is limited, it is costly, and it cannot meet people's comprehensive functional needs.

[0006] In conclusion, the market urgently needs to develop natural compound active ingredients that are both highly effective and safe, and have multiple functions such as whitening, fading spots, anti-oxidation, and skin repair and brightening, in order to meet consumers' multiple needs for product efficacy and gentleness. Summary of the Invention

[0007] The purpose of this invention is to overcome the shortcomings of the prior art and provide a whitening and freckle-removing essence with advantages such as high safety, whitening and freckle removal, brightening the skin, moisturizing and anti-allergy, strong penetration, significant efficacy and low cost, so as to meet the needs of most consumers.

[0008] Another objective of this invention is to provide a method for preparing the above-mentioned whitening and freckle-removing essence. The process is simple, can maximize the synergistic effects of various functional components, and the system is stable with an excellent skin feel.

[0009] The objective of this invention is achieved through the following technical solution: a whitening and freckle-removing serum, comprising the following weight proportions:

[0010]

[0011]

[0012] Preferably, the whitening and freckle-removing serum is prepared from the following components by weight:

[0013]

[0014]

[0015] The citrus (CITRUS RETICULATA) peel extract can be a citrus extract produced by Shaanxi Snowt Biotechnology Co., Ltd. under the brand name Snowt Bio.

[0016] The Sclerotium RolfssII gel can be the Sclerotium gel from Hubei Xinghengye Technology Co., Ltd.

[0017] The Bacillus / soybean fermentation product extract may be a Bacillus / soybean fermentation product extract produced by Shanghai Yuyu New Material Technology Co., Ltd.

[0018] The preferred Phellodendron amurense bark extract is Phellodendron amurense bark powder produced by Ningxia Xiangcao Biotechnology Co., Ltd.

[0019] The ROSA HYBRID flower extract may be the ROSA HYBRID flower extract sold by the supplier, Shanghai Yuyu New Material Technology Co., Ltd.

[0020] The gardenia fruit extract mentioned may be the gardenia fruit extract sold by Baoji Liupanyun Biotechnology Co., Ltd. under the brand name Liupanyun.

[0021] A method for preparing a whitening and freckle-removing serum, comprising the following steps:

[0022] (1) Heat the A phase mixture to 75-80℃, stir at 20-60r / min until all components are completely dissolved, and then homogenize at 2000-3000r / min for 1-2min to obtain the A phase mixture;

[0023] (2) Heat the B phase mixture to 75-80℃ and stir at 20-60r / min until completely dissolved to obtain the B phase mixture;

[0024] (3) Slowly add the B phase mixture to the A phase mixture through a filter screen, stir for 1-2 minutes, and then homogenize at 2000-3000 r / min for 2-5 minutes;

[0025] (4) After cooling to 60-70℃, add the C phase component, stir at 20-60 r / min for 1-2 min, and then homogenize at 2000-3000 r / min for 2-5 min;

[0026] (5) After cooling to 35-45℃, add the D phase component, stir at 20-60r / min for 1-2min, and then homogenize at 2000-3000r / min for 2-5min;

[0027] (6) After passing the inspection, the product is canned and packaged.

[0028] In step (5), after stirring, homogenization is performed at 2500 r / min while simultaneously applying ultrasonic treatment at 200 W and 25 kHz for 2 minutes, followed by homogenization at 3000 r / min for 2 minutes. This combination of ultrasonic treatment at a specific frequency and mechanical homogenization removes air from the formulation, prevents aggregation, improves suspension stability, and enhances the activity of active components. The subsequent homogenization after ultrasonic treatment further refines the microemulsion particle size and removes any gases that may be generated under ultrasonic action, thus improving product efficacy and formulation stability.

[0029] Preferably, the stirring speed in the above steps is 30 r / min and the homogenization speed is 2500 r / min.

[0030] The preferred temperature in steps (1) and (2) is 78°C, the preferred temperature in step (3) is 65°C, and the preferred temperature in step (4) is 40°C.

[0031] The sieve used for filtration in step (3) has a mesh size of 100-150.

[0032] When cooling from 75℃ to 65℃ in step (3), a gradient cooling rate of 5℃ / min is used.

[0033] The order of adding phase D in step (4) is as follows: first add nicotinamide, inositol, taurine, tranexamic acid and glutathione, then add trehalose and panthenol, followed by 1,2-hexanediol, 1,2-pentanediol, caprylyl glycol and ethylhexylglycerin, then add ascorbic acid tetraisopalmitate, then add 3-o-ethyl ascorbic acid and tetrahydrocurcumin, then add citrus peel extract, phellodendron bark extract, hybrid rose extract and Bacillus / soybean fermentation product extract, then slowly add sclerotium gum and hydroxyethyl deacetylated chitosan, briefly increase the stirring speed to 50 r / min and stir for 1 min, then return to 30 r / min, then add papain, stir quickly for 1 min after adding, then add folic acid, and finally add phenoxyethanol and fragrance. After all the ingredients are added, continue stirring at 30 r / min for 2-3 min to mix well.

[0034] Compared with the prior art, the present invention has the following beneficial effects:

[0035] (1) High safety: Most of the ingredients are natural and contain almost no chemical additives that are harmful to the skin. It is suitable for long-term use, especially for sensitive skin.

[0036] (2) Significant whitening and brightening effects: By adding specific amounts of niacinamide, glutathione, ascorbate tetraisopalmitate, 3-o-ethyl ascorbic acid, papain, 4-hydroxyethylpiperazine ethanesulfonic acid, and citrus peel extract, in synergy with other components of this formula, the whitening and radiance of the skin are significantly improved, and melanin deposition and dull skin problems are reduced.

[0037] (3) Good moisturizing effect: The formula system contains moisturizing components such as sodium hyaluronate, which can deeply moisturize and hydrate, and improve the skin pigmentation problem caused by dry skin;

[0038] (4) Strong antioxidant capacity: The formula system contains tetrahydrocurcumin and ascorbate tetraisopalmitate, which effectively enhance the damage of free radicals to the skin, protect cells, and effectively inhibit the production of tyrosinase.

[0039] (5) Lower cost: Compared with other high-end whitening, spot-fading and brightening products, the formula cost is lower, making it suitable for large-scale production and use by a wide range of users. Detailed Implementation

[0040] To make the objectives, technical solutions, and advantages of the present invention clearer, the present invention will be further described in detail below with reference to specific embodiments, but the embodiments of the present invention are not limited thereto.

[0041] In this formulation, the desmomeric arginine / lysine carboxyl termini produced by papain hydrolysis can sever the connections between keratinocytes, exfoliating dead skin cells. Within the granular layer of epidermal cells, caspase 14 degrades filaggrin, generating abundant non-toxic moisturizing proteins (NMFs), keeping the stratum corneum hydrated and soft, promoting skin renewal. NMFs are crucial molecules affecting skin hydration; their reduction or absence weakens the skin barrier function, leading to various skin diseases. 4-Hydroxyethylpiperazine ethanesulfonic acid gently promotes the exfoliation of old keratinocytes in the epidermis, enhances penetration and absorption of active ingredients, and softens the stratum corneum. This formulation combines papain and 4-hydroxyethylpiperazine ethanesulfonic acid to address surface skin problems, exfoliate old keratinocytes, and promote epidermal renewal.

[0042] Niacinamide is used to inhibit the process of melanin transfer in the skin, prevent photosensitivity and skin cancer caused by ultraviolet radiation, and inhibit melanin production, while stimulating collagen production and DNA synthesis. Niacinamide acts on the dermis to block and inhibit the upward migration of melanin.

[0043] Glutathione has extremely strong reducing properties, which can reduce melanin in cells into lighter pigmentation, inhibit the formation of age spots, increase the moisture content of the stratum corneum, and promote metabolism. In this formula, glutathione works synergistically with other components to lighten melanin and engulf melanocytes.

[0044] Ascorbate tetraisopalmitate can inhibit intracellular tyrosinase activity and melanin production, preventing lipid peroxidation and skin aging—acting as an antioxidant—and has excellent transdermal absorption, effectively reducing UV-induced cellular DNA damage. 3-o-Ethyl ascorbic acid, as a vitamin C derivative, can penetrate deep into the skin, promoting absorption and effectively inhibiting melanin production. This invention incorporates these two vitamin C derivatives, working synergistically with other components of the formulation to effectively inhibit tyrosinase and provide antioxidant effects.

[0045] Hydroxyethyl deacetylated chitosan (CAS No.: 123938-86-3) comprises self-assembled hydroxyethyl deacetylated chitosan nanoparticles. As a transdermal delivery carrier, it effectively delivers functional components to the skin surface and inner layers through intercellular spaces, stratum corneum cells, hair follicles, sebaceous glands, and sweat pores via similar compatibility, hydration, diffusion, and structural changes. In this formulation, hydroxyethyl deacetylated chitosan effectively promotes synergistic penetration of the active ingredients, further enhancing their efficacy and promoting skin absorption.

[0046] The combination of Bacillus / soy ferment extract and folic acid effectively inhibits the release of histamine from leukocytes, exhibiting superior anti-allergic effects compared to sodium cromoglycate, while being safer. It also effectively enhances the survival ability of fibroblasts under UV radiation. This formula, with the addition of Bacillus / soy ferment extract and folic acid, provides excellent soothing effects and is suitable for most skin types.

[0047] Citrus peel extract flattens and firms epidermal keratinocytes, reduces intercellular spaces, and improves keratinocyte structure; it also reduces the area of ​​scattered light, resulting in smoother and more translucent skin. Tetrahydrocurcumin has excellent antioxidant properties, protecting cells and making them healthier and more delicate; it inhibits hyaluronidase, protecting hyaluronic acid from decomposition and preserving its water-locking function; and it effectively scavenge free radicals. In this formula, the added citrus peel extract and tetrahydrocurcumin work synergistically with other components to effectively improve skin radiance.

[0048] This invention primarily uses the aforementioned components as its efficacy base, supplemented by other components described in the formula. Through a specific preparation method, the components synergistically enhance their effects. This formula system inhibits tyrosinase, lightens melanin, promotes epidermal metabolism and renewal, blocks melanin migration, improves intercellular spaces, and enhances skin brightness, radiance, and melanin reduction, resulting in excellent skin whitening, spot-fading, and brightening effects. Most of the components are natural raw materials, safe and non-irritating, suitable for people of all ages and skin types, and have a relatively high efficacy, thus appealing to a wide range of users.

[0049] A whitening and blemish-removing serum is made from the following components in parts by weight:

[0050]

[0051]

[0052] The preparation method of the above-mentioned whitening and freckle-removing serum includes the following steps:

[0053] (1) Deionized water, glycerol, butanediol, p-hydroxyacetophenone, PPG-26-butanol polyether-26 (polyethylene oxide polypropylene oxide monobutyl ether), PEG-40 hydrogenated castor oil (polyoxyethylene 40 hydrogenated castor oil) and sodium hyaluronate from phase A were added sequentially to an aqueous phase pot and mixed. The mixture was heated to 76°C and stirred at 30 r / min for 2 min until all components were completely dissolved. Then, the mixture was homogenized at 2500 r / min for 2 min to obtain phase A mixture.

[0054] (2) Isododecane, polydimethylsiloxane, stearyl alcohol polyether-21, cetearyl alcohol, stearyl alcohol polyether-2 and cetearyl glucoside in phase B are added to the oil phase pot in sequence and mixed. The mixture is heated to 76°C and stirred at 30r / min for 3min until all components are completely dissolved to obtain phase B mixture.

[0055] (3) The B phase mixture is slowly added to the A phase mixture through a filter screen, stirred at 30 r / min for 2 min, and then homogenized at 2800 r / min for 2 min;

[0056] (4) After cooling to 65°C, polysorbate-80, tocopheryl acetate, sodium acrylate / sodium acryloyl dimethyl taurate copolymer and isohexadecane in phase C are added in sequence, while stirring at 30 r / min for 2 min, and then homogenizing at 2500 r / min for 3 min.

[0057] (5) Cool to 38℃ and remove the following ingredients from phase D: nicotinamide, 3-o-ethyl ascorbic acid, phenoxyethanol, trehalose, ascorbate tetraisopalmitate, panthenol, taurine, ethylhexylglycerin, tetrahydrocurcumin, inositol, 4-hydroxyethylpiperazine ethanesulfonic acid, 1,2-hexanediol, hydroxyethyl deacetylated chitosan, papain, folic acid, caprylyl glycol, tranexamic acid, 1,2-pentanediol, Citrus reticulata peel extract, Phellodendron amurense bark extract, Rosa hybrid flower extract, Sclerotium rolfsii gum, Bacillus / soybean fermentation product extract, and Gardenia. FLORIDA fruit extract, glutathione and fragrance were added in sequence. First, the mixture was stirred at 30 r / min for 2 min. Then, it was homogenized at 2500 r / min and ultrasonically treated at 200 W and 25 kHz for 2 min. Finally, it was homogenized at 3000 r / min for 2 min.

[0058] (6) After passing the routine inspection, the product is canned and packaged.

[0059] Example 4

[0060] Compared to the above embodiment, the difference is that in step (5), no ultrasonic treatment is applied. Instead, the mixture is stirred at 30 r / min for 2 min and then homogenized at 2500 r / min for 3 min.

[0061] Effect test

[0062] I. Whitening Efficacy Test

[0063] 1. Experimental materials: Whitening and freckle-removing serums prepared in Examples 3, 4 and Comparative Examples 1-5.

[0064] 2. Subjects: Seventy volunteers aged 20-45 with sensitive skin and dull complexion were selected, including 35 men and 35 women. The volunteers were divided into 7 groups with 5 men and 5 women in each group.

[0065] 3. Test instruments: Skin red and melanin content tester (Mexameter MX18, CK GmbH, Germany), skin color tester (Skin-Colorimeter CL 400, CK GmbH, Germany).

[0066] Before volunteers tried the sample, a skin red and melanin tester and a special test probe were used to collect melanin MI value data on a pre-set test area on the volunteers' faces. The test area was a 5cm×5cm area of ​​skin on the cheeks, and the data obtained was used as the skin background value.

[0067] 4. Test methods:

[0068] Volunteers first cleansed their faces according to a standardized procedure. After their skin dried, they applied 1 mL of the whitening and spot-removing serum prepared according to the examples and comparative proportions evenly to the cheek test area, once in the morning and once in the evening, for four consecutive weeks. Each group consisted of 10 participants. The skin melanin content (MI value) and skin color (ITA° value) were measured using a skin red and melanin content analyzer and a skin color meter after 1, 2, and 4 weeks of use, respectively. Five measurements were taken for each test area, and the average value was calculated. The average values ​​of all volunteers were then used. The results are shown in the table below.

[0069] Table 1 Melanin content (MI value)

[0070] Melanin content Before use Use for 1 week Use for 2 weeks Use for 4 weeks Example 3 159.87 154.12 143.56 138.92 Example 4 160.15 158.38 157.21 150.57 Comparative Example 1 160.23 158.24 153.65 152.12 Comparative Example 2 160.23 159.54 158.87 157.12 Comparative Example 3 159.45 159.22 158.15 156.89 Comparative Example 4 160.08 157.26 149.34 145.78 Comparative Example 5 159.51 156.38 147.21 142.57

[0071] A lower MI value indicates a lower melanin content in the skin, resulting in a more significant whitening effect. Example 3 of this invention exhibits the most significant whitening effect. Although Example 5 contains all the necessary ingredients, its preparation lacks ultrasonic synergistic treatment, leading to a reduced whitening effect. This demonstrates that the ultrasonic step optimizes the preparation process of the whitening serum, helping each component to function more effectively. Comparative Example 2, which did not contain any whitening components, has the highest melanin content, proving that the unadded components in Comparative Example 2 have a whitening effect. Comparative Example 3, which did not contain hydroxyethyl deacetylated chitosan, also has a higher melanin content than Example 3, demonstrating that the hydroxyethyl deacetylated chitosan of this invention plays a role in promoting the penetration of other ingredients. Comparative Example 1's effect is slightly worse than Example 3, indicating that the two exfoliating components work synergistically, and the component promoting skin epidermal renewal has an auxiliary effect on whitening.

[0072] Table 2 ITA° values

[0073] ITA° value Before use Use for 1 week Use for 2 weeks Use for 4 weeks Example 3 25.51 33.48 40.24 49.96 Example 4 24.15 29.91 30.67 32.22 Comparative Example 1 25.21 32.53 33.81 34.25 Comparative Example 2 24.53 24.82 25.33 26.13 Comparative Example 3 24.36 26.82 27.33 28.13 Comparative Example 4 25.19 31.75 36.56 37.04 Comparative Example 5 24.97 27.35 29.12 30.87

[0074] A higher ITA° value indicates brighter skin; a lower value indicates duller skin. The data shows that Example 3 of this invention has the most significant effect. Example 4, although containing all the necessary ingredients, lacks ultrasonic synergistic treatment, resulting in a reduced whitening effect. Comparative Example 2, without any whitening components, has the lowest ITA° value. Comparative Example 3, without hydroxyethyl deacetylated chitosan, also has a lower ITA° value compared to Example 3, demonstrating that the hydroxyethyl deacetylated chitosan of this invention plays a role in promoting the penetration of other ingredients. Comparative Example 1 is less effective than Example 3, indicating that exfoliating and skin renewal-promoting ingredients have an auxiliary effect on whitening. Comparative Example 4 is slightly less effective than Example 3, indicating the absence of two ingredients reduces the brightening effect, significantly lower than the complete formula, while also demonstrating the synergistic effect of niacinamide, glutathione, ascorbate tetraisopalmitate, and 3-o-ethylascorbic acid.

[0075] II. Tyrosinase Activity Inhibition Test

[0076] The tyrosinase inhibition rate was determined according to the test method in standard T / GDCA 006-2021, "Test Method for Inhibition of Tyrosinase Activity by Cosmetic Raw Materials (In Vitro Method)". The results are shown in the table below.

[0077] Table 3 Tyrosinase inhibition rate

[0078] Tyrosinase inhibition rate (%) Example 3 56.2％ Example 4 20.3％ Comparative Example 1 26.2％ Comparative Example 2 10.3％ Comparative Example 3 22.4％ Comparative Example 4 38.7％ Comparative Example 5 38.2％

[0079] The data above show that the whitening and spot-removing serum prepared in Example 3 of this application has a higher tyrosinase inhibition rate. The inhibition rate of Comparative Example 2 is significantly lower than that of Example 3, indicating that the use of niacinamide, glutathione, tetraisopalmitate ascorbate, and 3-o-ethyl ascorbic acid can achieve a better tyrosinase inhibition effect in the serum. Comparative Example 1, lacking exfoliating and skin-renewing components, cannot effectively remove melanin-containing aging keratinocytes, indirectly affecting the tyrosinase inhibition effect. The inhibition rate of Comparative Example 5 is lower than that of Example 1 but higher than that of Comparative Example 2, indicating that although the brightening ingredients do not directly inhibit tyrosinase, they also have an auxiliary whitening effect, further demonstrating the importance of the synergistic effect of the various functional components.

[0080] III. Antioxidant Test

[0081] A blank matrix was prepared containing all components except the active ingredients papain, 4-hydroxyethylpiperazine ethanesulfonic acid, nicotinamide, glutathione, ascorbate tetraisopalmitate, 3-o-ethylascorbic acid, hydroxyethyl deacetylated chitosan, Bacillus / soybean fermentation product extract, folic acid, citrus peel extract, and tetrahydrocurcumin. The phosphate buffer solutions of Examples 3, 4, and Comparative Examples 1-5 were diluted to a concentration of 2% and filtered sterilized for later use. Trolox standard solutions at concentrations of 50, 100, and 200 μM were prepared using phosphate buffer.

[0082] Add 25 μL of the above sample, 25 μL of Trolox standard solution, and 25 μL of blank matrix to each well of a 96-well plate. Then add 25 μL of phosphate buffer to each well, setting up three replicate wells to ensure data accuracy. Subsequently, add 150 μL of sodium fluorescein solution to each well, mix well, and incubate at 37°C for 30 min. Immediately add 25 μL of free radical initiator (AAPH) solution to each well, mix well, and immediately measure fluorescence values ​​using a microplate reader at an excitation wavelength of 485 nm and an emission wavelength of 538 nm. Measure at 37°C for 60 min, reading data every 1 min.

[0083] The fluorescence decay curves (AUC values) for each well were calculated using Graphpad Prism software.

[0084] Net AUC sample = AUC sample - AUC blank matrix;

[0085] Net AUC standard = AUC standard - AUC blank matrix;

[0086] Reduce fluorescence interference that may be caused by blank matrix.

[0087] A linear regression curve was plotted with the concentration of the Trolox standard on the x-axis (X) and its corresponding net AUC value on the y-axis (Y). The net AUC value of each sample was substituted into the standard curve equation to calculate its corresponding Trolox equivalent concentration. The ORAC value of the sample was calculated based on the sample dilution factor, expressed in μmol Trolox / g.

[0088] 2. Calculation of predicted ORAC values ​​and assessment of synergistic antioxidant activity

[0089] Assuming the effects of each component in the color-enhancing serum are additive, the formula for predicting the ORAC value is as follows:

[0090] ORAC=(C1×ORAC1)+(C2×ORAC2)+…(C n ×ORAC m )

[0091] Where C1, C2…C n The concentrations of each component in the whitening and spot-removing serum are ORAC1, ORAC2…ORAC. n To obtain the ORAC values ​​of each component measured in a blank matrix, all measurements were performed in triplicate to ensure data accuracy.

[0092] Data were analyzed using Graphpad Prism software to compare the actual measured values ​​with the calculated predicted values ​​to determine if there were significant differences. When the actual ORAC measured value > the ORAC predicted value, a synergistic antioxidant effect was identified; when the actual ORAC measured value is approximately equal to the ORAC predicted value, the antioxidant effect was considered simple additive; and when the actual ORAC measured value < the ORAC predicted value, an antagonistic effect was identified. The antioxidant activity results for each component are shown in the table below:

[0093] Table 4 ORAC values

[0094]

[0095] The data above show that the actual ORAC value of the best example 3 is significantly greater than the predicted value, indicating that example 3 has a significant synergistic antioxidant effect. The difference between the actual and predicted ORAC values ​​in example 4 is small, indicating the key role of ultrasound in synergistic preparation. The difference between the actual and predicted ORAC values ​​in comparative example 2 is small, but the values ​​are all low, indicating that nicotinamide, glutathione, tetraisopalmitate ascorbate, and 3-o-ethyl ascorbic acid whitening and spot-removing essence are key components for achieving antioxidant effects. The difference between the actual and predicted ORAC values ​​in comparative example 3 is small, indicating that the lack of hydroxyethyl deacetylated chitosan prevents the antioxidant components from effectively penetrating and exerting their effects. The significantly decreased ORAC value in comparative example 5 indicates that citrus peel extract and tetrahydrocurcumin also play key roles in the antioxidant effect.

[0096] IV. Metabolic Test

[0097] ATP, or adenosine triphosphate, directly reflects the energy metabolism status of various organs. Therefore, intracellular ATP concentration is a key indicator for assessing cellular health and metabolic function. This experiment uses an ATP assay kit to detect intracellular ATP levels to assess cellular energy levels. The experiment includes the following steps:

[0098] (1) HaCaT cells (human epidermal keratinocytes) were cultured to the logarithmic growth phase, and after trypsin digestion, the cell suspension was diluted with DMEM complete medium to adjust the cell density to 5 × 10⁶ cells / year. 4 Cells / mL, 100 μL per well, were seeded into black 96-well cell culture plates. The cell suspension served as the experimental group, and the complete culture medium without cells served as the control group. Three control groups were set up to ensure the accuracy of the data. The cell plates were incubated at 37°C in a 5% CO2 cell culture incubator for 24 h.

[0099] (2) The whitening and freckle-removing serums of Examples 3, 4 and Comparative Examples 1-5 were diluted to a concentration of 2% with serum-free DMEM medium and filtered through a 0.22 μm filter membrane for sterilization. The original culture medium in the cell plate was removed. 100 μL of serum-free culture medium containing the corresponding sample was added to the experimental group, and 100 μL of serum-free culture medium was added to the control group. The cell plates were placed in a 37°C, 5% CO2 cell culture incubator for further culture. At culture times of 0 h, 1 h, 3 h, 6 h, 12 h, and 24 h, a group of wells was selected, ATP detection lysis buffer was added, and the relative optical units (RLU) value of the well was read.

[0100] (3) The intracellular ATP level was detected using an ATP assay kit (Promega, model: G7571), following the instructions. The ATP assay kit and the cultured cell plate were brought to room temperature for approximately 15 minutes. 100 μL of ATP assay working solution was added to each well, and the mixture was shaken for 2 minutes in the dark to fully lyse the cells. After standing at room temperature for 10 minutes, the ATP fluorescence signal was detected using a microplate reader, and the RLU value of each well was read. The results are shown in the table below:

[0101] Table 5 Cell Energy Levels

[0102]

[0103]

[0104] The data above show that Example 3 exhibits a highly significant effect in enhancing cell energy, proving that the whitening and spot-removing essence can effectively enhance cell energy. Comparative Example 5 shows a significant decrease, demonstrating that tetrahydrocurcumin and citrus peel extract are the main components affecting cell energy growth. Example 4 shows an extremely low cell energy growth rate, indicating a lack of ultrasound synergistic treatment; the synergistic activity of each component was not activated, further demonstrating the importance of ultrasound synergistic treatment. Comparative Example 3 shows very low cell energy, indicating that the nanoparticle hydroxyethyl deacetylated chitosan plays an important role in promoting the function of other components. Comparative Example 2 shows an extremely low cell energy growth rate, indicating that this comparative essence has almost no function in promoting cell energy; the results are not significantly different from the blank control group, suggesting that nicotinamide, glutathione, ascorbate tetraisopalmitate, and 3-o-ethyl ascorbic acid are the core components promoting cell energy growth.

[0105] V. Moisturizing Efficacy Test

[0106] 1. Experimental Materials: Whitening and freckle-removing serums prepared in Examples 3 and 4 and Comparative Examples 1-5. 2. Experimental Subjects: 75 volunteers aged 20-45 years were selected, including 35 males and 35 females, and were divided into 7 groups.

[0107] 3. Testing Instrument: Skin testing instrument (Corneometer CM825)

[0108] 4. Test environment conditions: temperature 20-22℃, humidity 40-60%, and real-time dynamic detection.

[0109] 5. Test methods:

[0110] Moisturizing performance was tested according to the method in "QB / T4256-2011 Guidelines for Evaluation of Moisturizing Efficacy of Cosmetics". Ten participants were tested in each sample group. Each subject applied 1 mL of sample to the marked area. Data were collected before use, 2 hours after use, and 4 hours after use. The average results were used to calculate the growth rate of skin stratum corneum moisture content. The growth rate results are shown in the table below:

[0111] Table 6. Growth Rate of Moisture Content

[0112] Use 2h Use 4h Example 3 53.6％ 43.9％ Example 4 41.3％ 33.2％ Comparative Example 1 37.3％ 28.6％ Comparative Example 2 23.3％ 12.6％ Comparative Example 3 28.9％ 18.8％ Comparative Example 4 49.3％ 38.2％ Comparative Example 5 45.8％ 36.8％

[0113] The data above shows that the whitening and spot-removing serum of Optimal Example 3 has a good moisturizing effect. The significant decrease in Comparative Example 1 indicates that papain and 4-hydroxyethylpiperazine ethanesulfonic acid, by removing dead skin cells, make subsequent moisturizing ingredients easier to penetrate and absorb. Their absence reduces the penetration efficiency of moisturizing ingredients and significantly decreases the moisture growth rate. The sharp decrease in Comparative Example 3 indicates that hydroxyethyl deacetylated chitosan is an important component for delivering moisturizing ingredients deep into the stratum corneum. The lower concentration in Example 4 indicates that ultrasonic treatment ensures thorough mixing of the active ingredients, enhancing the synergistic moisturizing effect of each component. The lowest concentration in Comparative Example 2 indicates that the whitening ingredient is the core ingredient, also possessing moisturizing properties; its absence weakens the overall basic function of the formula, resulting in the worst moisturizing effect.

[0114] VI. Spot-fading efficacy test

[0115] 1. Experimental materials: Whitening and freckle-removing serums prepared in Examples 3, 4 and Comparative Examples 1-5.

[0116] 2. Subjects: Seventy volunteers with melasma, aged 20-45, were selected, including 35 men and 35 women. The volunteers were divided into 7 groups.

[0117] 3. Test methods:

[0118] Volunteers first cleansed their faces according to a standardized procedure, with each person spending the same amount of time. After their skin dried, they applied 5 mL of the above sample evenly to their cheeks, using it twice a day, morning and evening, for one month. Each group consisted of 10 participants. Subjects evaluated the skincare product based on their skin improvement, assigning a score (X). The evaluation criteria were: significant fading of melasma (1 ≤ X < 3), slight fading (3 ≤ X < 4), and no change or worsening (4 ≤ X < 5). The results are shown in the table below.

[0119] Table 7 Skin Evaluation

[0120] evaluate Example 3 1.69 Example 4 4.01 Comparative Example 1 3.53 Comparative Example 2 4.07 Comparative Example 3 3.83 Comparative Example 4 3.12 Comparative Example 5 4.03

[0121] The data above shows that the whitening and freckle-removing essence of this invention has a significant freckle-fading effect. Comparative Example 2 indicates that niacinamide, glutathione, tetraisopalmitate ascorbate, and 3-o-ethyl ascorbic acid are the main active ingredients in the whitening and freckle-removing essence for freckle fading. Comparative Example 5 shows a significant decrease in effect, indicating the absence of antioxidant and melanin transfer inhibition pathways, resulting in reduced improvement in melasma. Comparative Example 1 shows poor effect, indicating the lack of components promoting epidermal renewal, which fails to effectively promote pigment metabolism; even if melanin production is inhibited, existing pigmentation is difficult to remove quickly. Comparative Example 3 shows poor effect, indicating that components assisting penetration play an important role in the essence. Examples 4 and Comparative Example 2 show almost no effect, indicating that the ultrasound-modified process and whitening components play a core role in improving melasma.

[0122] VII. Skin Safety Test

[0123] 1. Test materials: patch test tape: Finn chambers on Scanpor-8mm (Smart Practice, USA); hospital dermatology pen: used for positioning and skin marking.

[0124] 2. Subjects: Seventy volunteers aged 20-45 years were selected, including 35 men and 35 women, and were divided into 7 groups.

[0125] 3. Test Methods: A closed patch test was conducted on human skin according to the 2015 edition of the "Cosmetic Safety Technical Specifications". The results are shown in the table below:

[0126] Table 8 Test Evaluation Standards

[0127] grade Use 2h 0 Negative reaction: no irritation, no erythema 1 Suspicious reaction: faint erythema 2 Weak positive reaction: erythema, infiltration, papules 3 Strong positive reaction: erythema, edema, papules, vesicles 4 Extremely positive reaction: erythema, edema, bullae

[0128] Table 9. Results of the patch test

[0129] 1 week 4 weeks total Example 3 0 0 0 Example 4 0 0 0 Comparative Example 1 0 0 0 Comparative Example 2 0 0 0 Comparative Example 3 0 0 0 Comparative Example 4 0 0 0 Comparative Example 5 0 0 0

[0130] The formula in this application is mostly composed of natural ingredients. No skin reactions were observed in the 1-week and 4-week tests of each embodiment and comparative example. Within the permissible range of the judgment criteria, no adverse skin reactions were produced on the human body, indicating that the whitening and freckle-removing lotion described in this application is safe and non-irritating. Furthermore, the formula in this application contains a compound of Bacillus / soybean fermentation product extract and folic acid, which has excellent soothing and anti-allergic effects, helping the skin build long-term tolerance, making it suitable for long-term use by all types of people.

[0131] VIII. Stability Testing

[0132] The testing method involves thoroughly mixing the sample to ensure no significant stratification or precipitation. 20 mL of sample is placed in a transparent glass container, which is then sealed to prevent leakage. The sealed container is placed in a constant temperature incubator at 50°C for one month. After one month, the sample is removed, cooled to room temperature, and observed for oil and water phase separation, precipitation, clumping, discoloration, or off-odors. The stability evaluation criteria are: A: Consistent with the initial sample, no change; B: Slight change, still applicable; C: Moderate change, significant stratification or precipitation, unacceptable; D: Severe change, severe precipitation. The results are shown below:

[0133] Table 10 Stability Evaluation

[0134] evaluate Example 3 A Example 4 B Comparative Example 1 A Comparative Example 2 B Comparative Example 3 A Comparative Example 4 A Comparative Example 5 B

[0135] The data above shows that the whitening and freckle-removing lotion of the best embodiment 3 of this invention has excellent stability. Slight discoloration in Comparative Example 5 indicates that the lack of a major antioxidant component may lead to some oils or active ingredients in the formula being more easily oxidized at high temperatures, resulting in slight discoloration or decreased oil phase stability. Slight stratification in Comparative Example 2 indicates that the lack of whitening substances may have altered the compatibility ratio of the entire formula, leading to a slight decrease in system stability. Slight discoloration and stratification in Example 4 indicate that the lack of ultrasonic treatment prevented complete removal of oxygen from the formula system, and the inability to achieve deep emulsification resulted in slightly poorer stability at high temperatures, demonstrating the importance of the ultrasonic-modified process.

[0136] Based on the formulation components of this application, glycerin and butylene glycol reduce water activity (Aw), which can both moisturize and reduce the microbial survival environment, thus helping to prevent spoilage; PEG-40 hydrogenated castor oil solubilizes fragrances, essential oils, or oil-soluble active ingredients, preventing them from precipitating and forming turbidity; phenoxyethanol is a traditional preservative, while p-hydroxyacetophenone and ethylhexylglycerin are novel mild preservatives, which achieve a broad-spectrum preservative effect through compound use; sodium acrylate / sodium acryloyldimethyl taurate copolymer, taurine, and sclerotium gum increase the viscosity of the external phase and reduce the droplet movement speed, thereby delaying stratification.

[0137] This invention is mainly prepared by mixing the various phases in a specific ratio and through key processes such as temperature extraction and temperature-controlled emulsification (the process conditions necessary for the efficacy described in this application). This process fully extracts and retains the active ingredients that can synergistically enhance each component. It is rich in various active ingredients that inhibit tyrosinase activity, lighten melanin, promote epidermal metabolism and renewal, block melanin migration, improve intercellular spaces, accelerate keratin metabolism, and provide antioxidant effects. These ingredients effectively reduce pigmentation and lighten existing dark spots, resulting in significant whitening, spot-fading, and skin-brightening effects. The addition of trace amounts of highly active vitamin C derivatives and niacinamide further enhances its effectiveness in preventing pigmentation and fading spots. This formula also contains soothing and repairing ingredients, which can strengthen the skin barrier function and reduce potential irritation while whitening.

[0138] Although the above embodiments have provided a detailed description of the present invention, they are only some embodiments of the present invention, and not all embodiments. People can obtain other embodiments based on these embodiments without creative effort.

Claims

1. A whitening and freckle-removing serum, characterized in that, It is made from an array of the following weight parts:

2. The whitening and freckle-removing serum according to claim 1, characterized in that, The citrus (CITRUSRETICULATA) peel extract mentioned above is a citrus extract produced by Shaanxi Snowt Biotechnology Co., Ltd. under the brand Snowt Biotechnology; the sclerotium rolfsii gum mentioned above is sclerotium gum produced by Hubei Xinghengye Technology Co., Ltd.; the Bacillus / soybean fermentation product extract mentioned above is a Bacillus / soybean fermentation product extract produced by Shanghai Yuyu New Material Technology Co., Ltd.; the Phellodendron amurense bark extract mentioned above is Phellodendron amurense bark powder produced by Ningxia Xiangcao Biotechnology Co., Ltd.; the Rosa hybrid flower extract mentioned above is a Rosa hybrid flower extract sold by Shanghai Yuyu New Material Technology Co., Ltd.; and the Gardenia flower extract mentioned above is a Gardenia flower extract sold by Baoji Liupanyun Biotechnology Co., Ltd. under the brand Liupanyun.

3. The preparation method of the whitening and freckle-removing serum according to claim 1, characterized in that, The steps include the following: (1) Heat the A phase mixture to 75-80℃, stir at 20-60r / min until all components are completely dissolved, and then homogenize at 2000-3000r / min for 1-2min to obtain the A phase mixture; (2) Heat the B phase mixture to 75-80℃ and stir at 20-60r / min until completely dissolved to obtain the B phase mixture; (3) Add the B phase mixture to the A phase mixture, stir for 1-2 min, and then homogenize at 2000-3000 r / min for 2-5 min; (4) After cooling to 60-70℃, add the C phase component, stir at 20-60 r / min for 1-2 min, and then homogenize at 2000-3000 r / min for 2-5 min; (5) After cooling to 35-45℃, add the D phase component, stir at 20-60r / min for 1-2min, and then homogenize at 2000-3000r / min for 2-5min.

4. The preparation method of the whitening and freckle-removing serum according to claim 3, characterized in that, In step (5), after stirring, the mixture is homogenized at a speed of 2500 r / min while being subjected to ultrasonic treatment with a power of 200 W and a frequency of 25 KHz for 2 min, and then homogenized at a speed of 3000 r / min for 2 min.

5. The preparation method of the whitening and freckle-removing serum according to claim 3, characterized in that, The stirring speed was 30 r / min, and the homogenization speed was 2500 r / min.

6. The preparation method of the whitening and freckle-removing serum according to claim 3, characterized in that, The preferred temperature in steps (1) and (2) is 78°C, the preferred temperature in step (3) is 65°C, and the preferred temperature in step (4) is 40°C.

7. The preparation method of the whitening and freckle-removing serum according to claim 3, characterized in that, When cooling from 75℃ to 65℃ in step (3), a gradient cooling rate of 5℃ / min is used.

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