Application of tetramethylcurcumin in whitening

Abstract

The application belongs to the technical field of cosmetics, and discloses application of tetramethylcurcumin in whitening. The application first discovers that tetramethylcurcumin has the effect of whitening, specifically, can inhibit melanin production and tyrosinase activity, and the inhibiting effect is strong, and tetramethylcurcumin has no irritation to red blood cells, indicating that tetramethylcurcumin has certain safety as a potential raw material component of cosmetics.

Description

Technical Field

[0001] This invention belongs to the field of cosmetic technology, and specifically relates to the application of tetramethylcurcumin in whitening. Background Technology

[0002] Melanin, a biological pigment, is widely found in both plants and animals. Its primary source is melanocytes, where it is formed from tyrosine or 3,4-dihydroxyphenylalanine through a series of oxidation reactions, ultimately producing polymerized melanin granules. The presence of melanin has significant physiological importance. On one hand, melanin absorbs ultraviolet radiation, playing a crucial defensive role against radiation-induced DNA damage and oxidative stress. On the other hand, its content directly affects skin and hair color. Melanin granules produced by melanocytes located in the basal layer of the skin are continuously transported to surrounding keratinocytes via cell dendrites. The melanin granules taken in by keratinocytes ascend to the stratum corneum as keratinocytes proliferate and differentiate, thus influencing skin color. Abnormal synthesis, metabolism, or uneven distribution of melanin in the body can easily lead to localized pigmentation, causing age spots and other problems that not only affect appearance but may also cause lesions such as melanoma.

[0003] With the expanding demand in the skin whitening consumer market, more researchers are focusing on active ingredients that regulate melanin production, transport, and metabolism. A growing number of skin whitening ingredients have emerged on the market, including natural and synthetic monomers, extracts, and other forms. Currently, common skin whitening ingredients on the market can be broadly categorized into three types based on their mechanisms of action: 1) Inhibiting melanin production, most of which are tyrosinase inhibitors, such as kojic acid, arbutin, resveratrol, glycyrrhizin, phloretin, phenylethyl resorcinol, and rhododendronol; 2) Inhibiting melanin transport, such as niacinamide; and 3) Promoting melanin reduction, primarily composed of strong antioxidants, such as vitamin C and its derivatives. Many single chemical ingredients, while possessing good melanin-inhibiting effects, also exhibit strong irritant properties. For example, rhododendronol, if used improperly, can easily lead to skin allergies and even adverse reactions such as vitiligo, and has therefore been banned in some countries and regions. Therefore, how to achieve scientific and correct skin whitening and develop more efficient, gentle, and safe skin whitening ingredients is a technical challenge that urgently needs to be addressed by those skilled in the art. Summary of the Invention

[0004] The present invention aims to at least solve one of the technical problems existing in the prior art. To this end, the present invention provides the application of tetramethylcurcumin in whitening. Tetramethylcurcumin can effectively inhibit melanin production and tyrosinase activity, and has a low effective concentration and low irritation, making it suitable as a whitening ingredient for use in cosmetics.

[0005] In a first aspect, the present invention provides the use of tetramethylcurcumin in the preparation of products with whitening effects.

[0006] In some embodiments of the present invention, the amount of tetramethylcurcumin added to the product with whitening effect is ≤10wt%, preferably ≤6wt%, more preferably ≤3wt%, and even more preferably ≤1wt%.

[0007] In some embodiments of the present invention, the amount of tetramethylcurcumin added to the product with whitening effect is ≤0.5wt%, preferably 0.0004ppm-0.5wt%, more preferably 0.04ppm-0.4wt%, and even more preferably 0.2ppm-0.3wt%.

[0008] In some embodiments of the present invention, the amount of tetramethylcurcumin added to the product with whitening effect is 0.0001wt%-0.2wt%, preferably 0.001wt%-0.2wt%, and more preferably 0.01wt%-0.2wt%.

[0009] In a second aspect, the present invention provides a composition having whitening effects, the composition comprising one or more of tetramethylcurcumin or pharmaceutically acceptable acids, bases, salts, esters, solvates, stereoisomers, and tautomers thereof.

[0010] In some embodiments of the present invention, the content of the tetramethylcurcumin in the composition having whitening effect is ≤10wt%, preferably ≤6wt%, more preferably ≤3wt%, and even more preferably ≤1wt%.

[0011] In some embodiments of the present invention, the content of the tetramethylcurcumin in the composition having whitening effect is ≤0.5wt%, preferably 0.0004ppm-0.5wt%, more preferably 0.04ppm-0.4wt%, and even more preferably 0.2ppm-0.3wt%.

[0012] In some embodiments of the present invention, the content of the tetramethylcurcumin in the composition having whitening effect is 0.0001wt%-0.2wt%, preferably 0.001wt%-0.2wt%, and more preferably 0.01wt%-0.2wt%.

[0013] In a third aspect, the present invention provides a cosmetic with whitening effects, the cosmetic comprising one or more of tetramethylcurcumin or pharmaceutically acceptable acids, bases, salts, esters, solvates, stereoisomers, and tautomers thereof, or comprising the composition described in the second aspect of the present invention.

[0014] In some embodiments of the present invention, the content of the tetramethylcurcumin in the cosmetic with whitening effect is ≤10wt%, preferably ≤6wt%, more preferably ≤3wt%, and even more preferably ≤1wt%.

[0015] In some embodiments of the present invention, the content of the tetramethylcurcumin in the cosmetic with whitening effect is ≤0.5wt%, preferably 0.0004ppm-0.5wt%, more preferably 0.04ppm-0.4wt%, and even more preferably 0.2ppm-0.3wt%.

[0016] In some embodiments of the present invention, the content of the tetramethylcurcumin in the cosmetic with whitening effect is 0.0001wt%-0.2wt%, preferably 0.001wt%-0.2wt%, and more preferably 0.01wt%-0.2wt%.

[0017] In some embodiments of the present invention, the whitening cosmetic further includes excipients, which include one or more of emulsifiers, moisturizers, thickeners, chelating agents, and preservatives.

[0018] In some embodiments of the present invention, the dosage form of the cosmetic includes any one of aqueous solution, emulsion, spray, cream, serum or mask.

[0019] Compared with the prior art, the beneficial effects of the present invention are as follows:

[0020] This invention is the first to discover that tetramethylcurcumin has a whitening effect, specifically by inhibiting melanin production and tyrosinase activity with a strong inhibitory effect. Furthermore, tetramethylcurcumin is non-irritating to red blood cells, indicating that tetramethylcurcumin has a certain degree of safety as a potential raw material component for cosmetics. Detailed Implementation

[0021] To enable those skilled in the art to more clearly understand the technical solutions described in this invention, the following embodiments are provided for illustration. It should be noted that the following embodiments do not constitute a limitation on the scope of protection claimed by this invention.

[0022] Unless otherwise specified, the raw materials, reagents, and apparatus used in the following examples can be obtained from conventional commercial sources or by existing known methods.

[0023] Example 1

[0024] This embodiment provides a phenolic compound, tetramethylcurcumin (CAS: 52328-97-9).

[0025] Comparative Example 1

[0026] This comparative example provides a phenolic compound, resveratrol (CAS: 501-36-0).

[0027] Comparative Example 2

[0028] This comparative example provides a phenolic compound, phenylethyl resorcinol (CAS: 85-27-8).

[0029] Comparative Example 3

[0030] This comparative example provides a flavonoid compound, glycyrrhizin (CAS: 59870-68-7).

[0031] Comparative Example 4

[0032] This comparative example provides a flavonoid compound, phlorizin (CAS: 60-82-2).

[0033] Comparative Example 5

[0034] This comparative example provides a phenolic compound, glucosyl syringic acid (CAS: 33228-65-8).

[0035] Comparative Example 6

[0036] This comparative example provides a phenolic compound, protohematoxylin A (CAS: 102036-28-2).

[0037] Comparative Example 7

[0038] This comparative example provides a phenolic curcumin derivative, tetrahydrocurcumin (CAS: 36062-04-1).

[0039] Comparative Example 8

[0040] This comparative example provides a phenolic curcumin derivative, hexahydrocurcumin (CAS: 36062-05-2).

[0041] Comparative Example 9

[0042] This comparative example provides a phenolic curcumin derivative, demethoxycurcumin (CAS: 22608-11-3).

[0043] Experiment 1: Inhibition of Melanin Synthesis and Tyrosinase Activity

[0044] 1.1 Preparation of sample solution

[0045] The substances provided in Example 1 and Comparative Examples 1-9 were dissolved in dimethyl sulfoxide (DMSO) to prepare sample solutions with concentrations of 0.01 μM, 0.1 μM, 1 μM, and 5 μM, respectively, for testing.

[0046] 1.2 Experimental Principle

[0047] Melanin is a biological pigment synthesized and secreted by melanocytes. Within melanocytes, tyrosine is catalyzed by tyrosinase and undergoes a series of reactions to ultimately produce melanin.

[0048] B16-F10 mouse melanoma cells are widely used as test cells for assessing the efficacy of skin-whitening chemicals. α-MSH, an α-melanocyte-stimulating agent, can promote melanin secretion from B16-F10 cells and increase intracellular tyrosinase activity. By inducing B16-F10 cells with α-MSH, the inhibitory effects of test substances on melanin synthesis and tyrosinase activity can be measured.

[0049] 1.3 Experimental Methods

[0050] (1) Cell Culture and Processing

[0051] B16-F10 cells were cultured in 1640 complete medium (1640 medium + 10% FBS + antibiotics) in a carbon dioxide incubator at 37°C and 5% CO2, and passaged every 2-3 days.

[0052] (2) Measurement of intracellular melanin content

[0053] B16-F10 cells in logarithmic growth phase were selected, digested with 0.25% trypsin, and seeded into 6-well plates with 2 mL of culture medium per well. The plates were incubated at 37°C in a 5% CO2 incubator. The day after seeding, culture medium containing different concentrations of the test substance and 1 μM α-MSH was added, and a control group (containing only 1 μM α-MSH) was set up with 2 mL per well. Each concentration was repeated in triplicate. After incubation at 37°C in a 5% CO2 incubator for 48 h, the supernatant was discarded, and 0.5 mL of 0.25% trypsin digestion solution was added to each well. Digestion was carried out at room temperature for 1 min. Digestion was stopped by adding 1 mL of culture medium, and the cells were pipetted into a single-cell suspension. Take 20 μL for cell counting, take 0.7 mL of cell suspension, centrifuge at 2000 r / min for 5 min, discard the supernatant, add 1 mL of 1 mol / L NaOH solution (containing 10% DMSO by mass), and incubate in an 80℃ water bath for 30 min. Then measure the absorbance at 490 nm using an ELISA reader.

[0054] (3) Measurement of intracellular tyrosinase activity

[0055] After the cells were treated with the above drugs, they were digested and counted with trypsin (specific steps are the same as 1.3-(2)). 0.7 mL of the cell suspension was centrifuged at 2000 r / min for 5 min, the supernatant was discarded, and 1 mL of 0.5% sodium deoxycholate solution was added. The cells were incubated on ice for 15 min to lyse them and prepare an extract containing tyrosinase. After preheating at 37℃, 0.5 mL of 0.3% dopa solution was added, and the reaction was carried out at 37℃ for 10 min. The absorbance was then measured at 475 nm using an ELISA reader.

[0056] (4) Results Statistics

[0057] The OD value is calibrated by the number of cells.

[0058] Melanin synthesis inhibition rate (%) = [1 - (As / Ds) / (Ac / Dc)] × 100;

[0059] Tyrosinase activity inhibition rate (%) = [1 - (As / Ds) / (Ac / Dc)] × 100;

[0060] In the formula: As is the absorbance of the sample group, Ac is the absorbance of the control group, Ds is the cell concentration of the sample group, and Dc is the cell concentration of the control group.

[0061] 1.4 Experimental Results

[0062] The results are shown in Table 1-3.

[0063] Table 1

[0064]

[0065] Table 2

[0066]

[0067]

[0068] Table 3

[0069]

[0070] It is understandable that skin tone, dullness, and problems such as pigmentation and uneven pigmentation are all closely related to melanin. Melanin is mainly produced by melanocytes, and tyrosinase is the key enzyme that catalyzes melanin synthesis during its production. Therefore, by using melanocytes, the whitening effect of the test product can be judged by assessing the melanin content and tyrosinase activity.

[0071] Currently, in the skin whitening market, resveratrol and phenylethyl resorcinol with phenolic structures, and glycyrrhizin and phloretin with flavonoid structures are all well-known cosmetic raw materials with good whitening effects. As shown in Tables 1 and 2, comparative examples 1 (resveratrol), 2 (phenylethyl resorcinol), 3 (glycyrrhizin), and 4 (phloretin) all have certain effects in inhibiting melanin production and tyrosinase activity. However, comparative examples 5 (glucosyl eugenol) and 6 (proto-hematoxylin A), which also belong to the phenolic structure, are not currently listed in the catalog of cosmetic raw materials and did not show obvious inhibitory effects at concentrations of 0.01-1 μM. Example 1 (tetramethylcurcumin) of this invention exhibited similar inhibitory activity to Comparative Examples 1-4, and at a concentration of 1 μM, its inhibitory effect was significantly better than that of common whitening ingredients (Comparative Examples 1-4). Furthermore, at a low concentration of 0.01 μM, while the inhibitory effects of Comparative Examples 1-4 were significantly reduced, the effect of Example 1 (tetramethylcurcumin) remained at approximately 10%. Meanwhile, as shown in Table 3, Example 1 (tetramethylcurcumin) demonstrated superior tyrosinase inhibitory effects compared to other curcumin derivatives (Comparative Examples 7-9).

[0072] The results in Tables 1-3 above indicate that Example 1 (tetramethylcurcumin) exhibits strong inhibitory effects on melanin production and tyrosinase activity, and has a wider effective concentration range compared to Comparative Examples 1-4. This suggests that Example 1 (tetramethylcurcumin) may achieve a whitening effect with only a small amount added in practical applications. Furthermore, Example 1 (tetramethylcurcumin) demonstrates stronger whitening activity compared to other curcumin derivatives.

[0073] Experimental Example 2: Red Blood Cell Hemolysis Test

[0074] 1.1 Experimental Principle

[0075] Eye irritants can denature proteins or dissolve cell membranes. By applying the test substance to red blood cells, the occurrence of hemolysis and the denaturation of hemoglobin released from the cells are detected. The extent of damage to the cell membrane by the test substance is evaluated by measuring the amount of leaked hemoglobin and the degree of denaturation, thus inferring its potential eye irritation. The red blood cell hemolysis assay simulates damage similar to that to the cornea.

[0076] 1.2 Experimental Methods (Refer to EURL ECVAM DB-ALM Protocol No. 99 and EURL ECVAM DB-ALM Protocol No. 37)

[0077] (1) The test substance—Example 1—and Comparative Examples 1-4 were prepared into suspensions with mass fractions of 0.001%, 0.01%, 0.1%, 1%, 10%, 20%, 30%, and 40% using physiological saline as a solvent. These suspensions were then mixed with an equal volume ratio of red blood cell suspension, incubated at 37°C for 3 hours, and the supernatant was collected by centrifugation. The absorbance was measured at 410 nm, 540 nm, and 575 nm using an ELISA reader, and the results were calculated and statistically analyzed. Physiological saline was used as a negative control (i.e., no hemolysis), and 0.1% SDS solution was used as a positive control (i.e., complete hemolysis).

[0078] (2) Results Statistics

[0079] Hemolysis rate (%) = (A 410nm -C 410nm ) / (B 410nm -C 410nm )×100;

[0080] Protein denaturation index DI (%) = (A 575nm / A 540nm ) / (B 575nm / B 540nm )×100;

[0081] In the formula, A is the test sample, B is the positive control, and C is the average absorbance of the negative control. In the DI% calculation, it refers to the average absorbance of the A / B samples at a concentration of 1%.

[0082] Plot a curve with sample concentration on the x-axis and hemolysis rate on the y-axis, and determine the concentration of the test substance HC50 (mg / L) that causes 50% hemolysis of red blood cells based on the regression equation.

[0083] (3) Result determination

[0084] Calculate the L / D value of the sample based on the HC50 and DI values: L / D = HC50 / DI;

[0085] According to the European Centre for Verification of Alternative Methods (ECVAM) RBC test grading standard, the irritation level of cosmetics is graded, and the grading standard is shown in Table 4.

[0086] Table 4

[0087] L / D Classification ﹥100 Non-irritating 10 < L / D ≤ ​​100 Mild irritation 1 < L / D ≤ ​​10 Mild irritation 0.1 < L / D ≤ ​​1 moderate irritation L / D≤0.1 Severe irritation

[0088] 1.3 Experimental Results

[0089] The results are shown in Table 5.

[0090] Table 5

[0091]

[0092] Comparative Examples 1-4 are all popular and effective cosmetic ingredients currently available in the whitening field. As shown in Table 5, comparing the irritation of different samples to red blood cells reveals that Example 1 (tetramethylcurcumin) is non-irritating and relatively mild, similar to Comparative Examples 1-4. Therefore, from a safety perspective, Example 1 (tetramethylcurcumin) of this invention has the potential for application in the cosmetic field.

[0093] Although tetramethylcurcumin is not currently listed in the cosmetic raw material catalog, it has low irritation and good whitening effects. These characteristics provide good data support for its application in the cosmetic field, especially in whitening products.

[0094] Application Example 1

[0095] A composition with whitening effect, comprising 10 wt% tetramethylcurcumin and the remainder being physiological saline solvent.

[0096] Application Example 2

[0097] A composition with whitening effect, comprising 0.5 wt% tetramethylcurcumin and the remainder being physiological saline solvent.

[0098] Application Example 3

[0099] A composition with whitening effect, comprising 0.2 wt% tetramethylcurcumin and the remainder being physiological saline solvent.

[0100] Application Example 4

[0101] A composition with whitening effect, comprising 0.0001 wt% tetramethylcurcumin and the balance being physiological saline solvent.

[0102] Application Example 5

[0103] A whitening toner, composed of the following components by weight percentage:

[0104] The ingredients are: 6% glycerol, 0.08% sodium hyaluronate, 0.05% xanthan gum, 0.4% p-hydroxyacetophenone, 1% 1,2-hexanediol, 10% tetramethylcurcumin, and the balance being water.

[0105] Application Example 6

[0106] A facial mask with whitening effects is composed of the following ingredients by weight percentage:

[0107] The ingredients are: 6% glycerol, 0.02% disodium EDTA, 0.08% sodium hyaluronate, 0.05% xanthan gum, 0.15% carbomer, 0.4% p-hydroxyacetophenone, 1% 1,2-hexanediol, 0.5% tetramethylcurcumin, and water as balance.

[0108] Application Example 7

[0109] A whitening lotion is composed of the following components by weight percentage:

[0110] 10% glycerol, 0.01% disodium EDTA, 0.1% ammonium acryloyldimethyl taurate / VP copolymer, 0.05% sodium hyaluronate, 0.4% p-hydroxyacetophenone, 1% 1,2-hexanediol, 1.5% squalane, 0.01% tetramethylcurcumin, and water balance.

[0111] It is readily apparent that the compositions of Application Examples 1-4 and the cosmetics of Application Examples 5-7 of the present invention, due to containing the tetramethylcurcumin of the present invention, possess excellent whitening effects.

[0112] The preferred embodiments of the present invention have been described in detail above, but the present invention is not limited to the embodiments described. Those skilled in the art can make various equivalent modifications or substitutions without departing from the spirit of the present invention, and these equivalent modifications or substitutions are all included within the scope defined by the claims of this application.

Claims

1. Application of tetramethylcurcumin in the preparation of products with whitening effects.

2. The application according to claim 1, characterized in that, The amount of tetramethylcurcumin added to the product is ≤10wt%.

3. The application according to claim 2, characterized in that, The amount of tetramethylcurcumin added to the product is ≤0.5wt%.

4. The application according to claim 3, characterized in that, The amount of tetramethylcurcumin added to the product is 0.0001wt%-0.2wt%.

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