Composition comprising niacinamide and resveratrol salicylate

Abstract

A topical skincare composition comprising niacinamide and one or more types of resveratrol salicylate. The composition increases energy levels in epidermal keratinocytes while mitigating oxidative damage associated with increased ATP production due to niacinamide application. It also increases the expression of AQP-3, CLDN-1, and CLDN-4, resulting in improved skin barrier function and increased moisture retention. Preferred compositions are in the form of a cream, lotion, serum, or gel, and can be implemented as a mixture or emulsion.

Description

Technical Field

[0001] The present invention is in the field of topical compositions, more specifically, topical skin care compositions that enhance the energy level in skin cells, regulate the water content, while reducing the effects of oxidation.

Background Art

[0002] Adenosine monophosphate-activated protein kinase (AMPK) is considered a major regulator of metabolism that affects numerous metabolic pathways. AMPK is an enzyme complex that detects the intracellular AMP / ATP ratio. When this ratio increases (indicating energy depletion), the AMPK metabolic switch is activated, which leads to several means for increasing the energy level of cells, such as increased fat burning, ketosis, and increased glucose uptake by cells. If the energy deficit is maintained long enough, AMPK induces the process that results in mitochondrial biogenesis (generation of new mitochondria). Among the known activators of AMPK are niacinamide and resveratrol.

[0003] Niacinamide (also known as nicotinamide) is one of three forms of vitamin B3. It is water-soluble and has the following general formula.

[0004]

Chemical Formula

[0005] Niacinamide is used both orally and topically. Topical application of niacinamide is known to treat acne, psoriasis, rosacea, and other inflammatory skin conditions. Niacinamide has been suggested to exhibit anti-inflammatory, anti-aging, anti-cancer, and skin-moisturizing properties. See, for example, Chen, ACN, & Damian, DL (2014). Nicotinamide and the skin. Australasian Journal of Dermatology, 55(3), 169-175. doi: / 10.1111 / ajd.12163.

[0006] Niacinamide is a precursor to nicotinamide adenine dinucleotide (NAD). Within cells, niacinamide is converted to nicotinamide adenine dinucleotide, which acts as an electron transporter and is an essential component for cellular energy production (ATP) via the electron transport system and oxidative phosphorylation. Increased availability of precursor niacinamide is thought to boost ATP production and enhance energy-dependent cellular processes such as DNA repair. Simultaneously, as cells produce more energy, levels of potentially harmful byproducts, particularly reactive oxygen species, also increase.

[0007] Resveratrol, also known as 3,4',5-trihydroxystilbene, has the general formula

[0008] [ka] It is a polyhydroxysubstituted compound having [a specific characteristic].

[0009] Resveratrol possesses potent antioxidant, anti-inflammatory, and life-extending properties. It helps stimulate collagen production while protecting the skin from the damaging effects of free radicals. Due to its perceived rejuvenating properties, resveratrol is incorporated into various cosmetic formulations, such as skin creams. However, one problem with resveratrol is its general instability in cosmetic formulations. This problem has been addressed by derivatizing resveratrol by reacting one or more hydroxyl groups with substituents that confer stability when resveratrol is incorporated into cosmetics. In particular, esterifying resveratrol with inorganic acids, organic carboxylic acids, or reacting hydroxyl groups to form alkoxy substituents or glycosides is known to provide resveratrol derivatives that are stable in cosmetic emulsions and enable the formulation of cosmetics with the aesthetic and stability required for commercially successful products. See, for example, U.S. Patents 8,084,496 and 9,220,669, which describe salicylic acid esters of resveratrol.

[0010] Purpose of the invention The main objective of the present invention is to provide a topical composition that increases energy levels in skin cells by increasing the rate of ATP production, while also providing protection from oxidative species generated in the process.

[0011] Another object of the present invention is to increase the levels of AQP-3, CLDN-1, and CLDN-4 in epidermal keratinocytes, thereby improving the barrier function and increasing moisture retention in the skin.

[0012] A further object of the present invention is to provide a skincare composition comprising niacinamide, at least one resveratrol salicylate, an aqueous phase, and optionally an oil phase. [Overview of the Initiative]

[0013] The present invention relates to a topical skincare composition comprising niacinamide and one or more types of resveratrol salicylate. Preferred compositions are in the form of a cream, lotion, emulsion, gel, serum, and / or powder. [Brief explanation of the drawing]

[0014] [Figure 1] Figure 1 shows the oxygen consumption rate by NHDF treated with niacinamide. [Figure 2] Figure 2 shows the oxygen consumption rate by NHDF treated with niacinamide and salicylic acid resveratrol. [Figure 3] Figure 3 shows the changes in peroxide species levels in NHDF when comparing niacinamide, resveratrol salicylate, and a combination of niacinamide and resveratrol salicylate. [Figure 4] Figure 4 shows the oxygen consumption rate over 90 minutes using NHDF treated with niacinamide. [Figure 5] Figure 5 shows the oxygen consumption rate over 90 minutes using NHDF treated with niacinamide and salicylate resveratrol. [Figure 6] Figure 6 shows NAD+ / NADH measurements in NHDF cells from 28-year-old and 68-year-old individuals treated with various concentrations of salicylic acid resveratrol, niacinamide, and combinations of niacinamide and salicylic acid resveratrol. [Figure 7] Figure 7 shows NAD+ / NADH measurements in NHDF cells from 28-year-old and 68-year-old individuals treated with various concentrations of salicylic acid resveratrol, niacinamide, and combinations of niacinamide and salicylic acid resveratrol. [Figure 8] Figure 8 shows the change in the ratio of phosphorylated (activated) AMPK to total AMPK in NHDF treated with niacinamide and salicylic acid resveratrol. [Figure 9] Figure 9 shows the results from Figure 8, expressed as a percentage of the control group. [Figure 10] Figure 10 shows the mass amount per 1 mL of phosphorylated (activated) AMPK measured by enzyme-linked immunosorbent assay (ELISA). [Figure 11] Figure 11 shows the results of Figure 10, expressed as a percentage of the control. [Figure 12A] Figure 12A shows the changes in the expression of hydration target genes, aquaporin-3, claudin-1, and claudin-4 in male NHEKs aged 28 and 56 years as a result of treatment with the combination of niacinamide and salicylic acid resveratrol. <管理编号: [Figure 12B] Figure 12B shows the changes in the expression of hydration target genes, aquaporin-3, claudin-1, and claudin-4 in male NHEKs aged 28 and 56 years as a result of treatment with the combination of niacinamide and salicylic acid resveratrol. [Figure 12C] Figure 12C shows the changes in the expression of hydration target genes, aquaporin-3, claudin-1, and claudin-4 in male NHEKs aged 28 and 56 years as a result of treatment with the combination of niacinamide and salicylic acid resveratrol. [Figure 13A] Figure 13A shows the changes in the expression of hydration target genes, aquaporin-3, claudin-1, and claudin-4 in male NHEKs aged 28 and 56 years as a result of treatment with the combination of niacinamide and salicylic acid resveratrol. [Figure 13B] Figure 13B shows the changes in the expression of hydration target genes, aquaporin-3, claudin-1, and claudin-4 in male NHEKs aged 28 and 56 years as a result of treatment with the combination of niacinamide and salicylic acid resveratrol. [Figure 13C] Figure 13C shows the changes in the expression of hydration target genes, aquaporin-3, claudin-1, and claudin-4 in male NHEKs aged 28 and 56 years as a result of treatment with the combination of niacinamide and salicylic acid resveratrol.

Mode for Carrying Out the Invention

[0015] The topical skin care composition contains a cosmetically acceptable base, niacinamide, and one or more salicylic acid esters resveratrol. The base is generally aqueous, and the amount of water can range from about 0.1 to 99% by weight, preferably from about 5 to 85% by weight, more preferably from about 7 to 75% by weight of the total composition. Some preferred embodiments of the present invention may include an oil phase. For example, the composition can be in the form of a water-in-oil or oil-in-water emulsion. The amount of oil in the oil phase is preferably in the range of about 1 to 95% by weight, preferably from about 5 to 85% by weight, more preferably from about 7 to 65% by weight of the total composition.

[0016] Salicylic acid esters resveratrol that may be useful in the present invention include 3-salicylate-5,4'-dihydroxystilbene, 5-salicylate-3,4'-dihydroxystilbene, 4'-salicylate-3,5-dihydroxystilbene, 3,5-disalicylate-4'-hydroxystilbene, 3,4'-disalicylate-5-hydroxystilbene, 4',5-disalicylate-3-hydroxystilbene, 3,5,4'-trisalicylate stilbene, or combinations thereof. <00so0101>

[0017] Effect on ATP production and oxidative damage The inventors have demonstrated that when one or more salicylic acid esters resveratrol are used in combination with niacinamide, oxidative damage caused by an increase in ATP production in cells is reduced. The following experimental results demonstrate the usefulness of salicylic acid resveratrol combined with niacinamide. Various measurements were performed on normal human dermal fibroblasts (NHDF) of 28 and 68 years old.

[0018] Figures 1 and 2 show the rate of oxygen consumption by NHDF and the change in ATP production levels. This trend suggests an increase in available cellular energy.

[0019] Figure 3 shows the changes in peroxide species levels in NHDF. Results are normalized so that the H2O2 level in untreated cells is 1. Niacinamide alone significantly increased the levels of oxidized species in 68-year-old cells, but resveratrol salicylate eliminated them all and did more. In 28-year-old cells, resveratrol salicylate reduced oxidation to levels higher than in untreated cells, despite the application of niacinamide.

[0020] Figures 4 and 5 show the oxygen consumption rate by NHDF over 90 minutes. Figure 4 shows cells treated with niacinamide. Figure 5 shows cells treated with a combination of niacinamide and salicylic acid resveratrol. The data suggest that the combination of niacinamide and salicylic acid resveratrol tends to enhance ATP production more than niacinamide alone.

[0021] Figures 6 and 7 show NAD+ / NADH measurements in NHDF cells from 28-year-old and 68-year-old cells. Test samples were prepared with three different concentrations of resveratrol salicylate, three different concentrations of niacinamide, and three different combinations of concentrations. The results indicate that niacinamide increases NADH production in both cell lines.

[0022] Figure 8 shows the change in the ratio of phosphorylated (activated) AMPK to total AMPK in NHDFs aged 28 and 68 years. Measurements were taken 1 and 3 hours after application. Metformin, a known activator of AMPK, is included for comparison. FBS alone was used as a control. This increase in the ratio indicates greater energy production. In Figure 9, the results are expressed as a percentage of the control. Combined treatment with niacinamide (0.04%) and resveratrol salicylate (0.1%) for 1 and 3 hours in low serum medium (e.g., 0.2% FBS) activates the AMPK pathway and generates more energy.

[0023] The levels of phosphorylated (activated) AMPK were also measured by enzyme-linked immunosorbent assay (ELISA). Figure 10 shows the mass per 1 mL, while Figure 11 shows the results as a percentage relative to the control. Measurements were performed 1 hour after application. 1 hour of niacinamide (4%) treatment in low serum medium (e.g., 0.2% FBS) activated the AMPK pathway, generating more energy in mature NHDF cells. 1 hour of salicylic acid resveratrol (0.1%) treatment specifically activated the AMPK pathway, generating more energy in both young and mature NHDF cells (red arrows). Surprisingly, the combination of niacinamide (4%) and salicylic acid resveratrol (0.1%) did not have an additive effect on AMPK activation. In fact, the level of activation was reduced.

[0024] When applied topically, the compositions of the present invention enhance energy production in skin cells and improve cellular function. Oxidative damage associated with increased ATP production due to niacinamide application is significantly reduced by resveratrol salicylate.

[0025] Effects on the levels of AQP-3, CLDN-1, and CLDN-4 in epidermal keratinocytes The combination of resveratrol salicylate and niacinamide has been further demonstrated to modulate the levels of AQP-3, CLDN-1, and CLDN-4 in epidermal keratinocytes, providing an opportunity to improve the barrier function of dry skin and leading to improved moisture content in dry skin. The following experimental results demonstrate the usefulness of resveratrol salicylate combined with niacinamide. Various measurements were performed on normal human epidermal keratinocytes (NHEKs) from men aged 29 and 56 years.

[0026] Trehalose and betaine are osmotic regulators that help retain moisture and ultimately support the hydration of skin cells. The inventors tested the effects of these osmotic regulators individually and in combination with niacinamide-resveratrol salicylate. The inventors also conducted the following gene expression studies in young and mature male NHEK.

[0027] For each age group, 25 seed plates were prepared as follows: Normal human epidermal keratinocytes (75 k / well in 1.875 mL of culture medium) were incubated at 37°C, 95% humidity, and 5% CO2 for 24 hours. The following treatment solutions were prepared. (1) Culture medium (NHEK medium supplemented with Keratinocyte Growth Media) (2) DMSO (dimethyl sulfoxide) in the culture medium (259 μL of DMSO in 6.741 mL of culture medium) (3) 0.2% trehalose in the culture medium (4) 0.5% betaine in the culture medium (5) 0.04% niacinamide + 0.05% resveratrol salicylate in the culture medium (6) Combinations, 2 mL each: 0.8% trehalose, 2% betaine, 0.16% niacinamide, 0.2% resveratrol salicylate.

[0028] Each well was treated with 1 mL of one of the aforementioned treatment solutions, and then incubated at 37°C, 95% humidity, and 5% CO2 for 24 hours. The cells were then washed twice with 1 mL of PBS. Quantitative real-time PCR (qRT-PCR) was performed as follows.

[0029] RNA isolation and quantification were performed using the miRNeasy Micro Kit (Qiagen, catalog number 217084) and the Quant-it® RiboGreen RNA Assay Kit (Invitrogen, catalog number R11490), respectively. 1 μg of RNA was reverse transcribed into complementary DNA (cDNA) using either the SuperScript® IV VILO® Master Mix (ThemoFisher, catalog number 11756050) or the TaqMan microRNA Reverse Transcription Kit (ThemoFisher, catalog number 4366596).

[0030] The obtained cDNA was amplified using a Quant Studio 7 Flex system (Applied Biosystems) containing a Universal PCR Master Mix (Life Technologies, catalog number 4440040) and recommended PCR conditions for quantitative assessment of gene transcript levels in the sample.

[0031] To evaluate the expression of hydration target genes, aquaporin-3, claudin-1, and claudin-4, in NHEKs aged 29 and 56, Taqman assay probes CLDN-1 (Hs00221623_m1), CLDN-4 (Hs00976831_s1), and AQP-3 (Hs00185020_m1) were used.

[0032] The data were normalized against a glyceraldehyde-3-phosphate dehydrogenase control (GAPDH, Hs99999905_m1). qRT-PCR data were calculated using the 2-ΔΔCT method (Livak and Schmittgen, 2001).

[0033] The data represent at least two independent biological replicated experiments. The data were graphed and analyzed using GraphPad Prism software (Prism 7.0). Statistical significance of differences between means was calculated using one-way ANOVA followed by Dunnett's post-hoc test. The results are shown in Figures 12 and 13.

[0034] Referring to Figure 12A, trehalose slightly increased AQP-3 gene expression levels in young keratinocyte cells, while all other test samples showed no increase.

[0035] Referring to Figure 12B, trehalose and betaine had no effect on CLDN-1 levels in young keratinocytes. The niacinamide / resveratrol salicylate combination increased CLDN-1 levels in young keratinocytes in a statistically significant manner. The trehalose / betaine / niacinamide / resveratrol salicylate combination also increased CLDN-1 levels, suggesting that the niacinamide / resveratrol salicylate combination was responsible for the increase.

[0036] Referring to Figure 12C, all treatments provided only a slight increase in CLDN-4 gene expression levels.

[0037] Referring to Figures 13A to 13C, trehalose and betaine have no significant effect on AQP-3, CLDN-1, or CLDN-4 gene expression. The niacinamide / resveratrol salicylate combination increases the levels of AQP-3, CLDN-1, and CLDN-4 in mature keratinocytes in a statistically significant manner, particularly AQP-3. The trehalose / betaine / niacinamide / resveratrol salicylate combination also increases the levels of AQP-3, CLDN-1, and CLDN-4, particularly CLDN-1 and CLDN-4, in mature keratinocytes, which is unexpected since trehalose and betaine alone have no effect or only a slightly negative effect.

[0038] Overall, the fact that the active substance acted better on NHEK in mature men compared to younger cells suggests that young cells already possess healthy basal levels of NMF and osmoregulators, which mask the beneficial effects of the active ingredient, whereas mature cells, due to aging, have lower basal levels of NMF and osmoregulators, and therefore the effects of the treatment are more pronounced.

[0039] In summary, the inventors have demonstrated that when one or more salicylic acid ester resveratrols are used in combination with niacinamide, they reduce oxidative damage caused by increased ATP production in cells. Furthermore, the niacinamide / resveratrol salicylate combination increases the levels of AQP-3, CLDN-1, and CLDN-4, particularly AQP-3, while the trehalose / betaine / niacinamide / resveratrol salicylate combination also increases the levels of AQP-3, CLDN-1, and CLDN-4, particularly CLDN-1 and CLDN-4. Therefore, the compositions according to the present invention are effective in increasing the levels of hydration targets in mature cells, helping them retain more moisture and supporting healthy, hydrated skin.

[0040] In the compositions according to the present invention, the useful level of niacinamide is in the range of about 0.0001% to about 5% of the total weight of the composition, for example, in the range of about 0.001% to about 5%, for example, in the range of about 0.01% to 3% of the total weight of the composition. Preferably, the concentration of niacinamide is 2% to 4%. One or more salicylic acid resveratrols may be present in the range of about 0.001% to 5%, for example, in the range of about 0.005% to 5%, or about 0.01% to 5% of the total weight of the composition. Preferably, the concentration of all salicylic acid resveratrols is about 0.5% to 2%. Some preferred embodiments of the present invention include 0.1% to 2% trehalose and 0.1% to 2% betaine relative to the total weight of the composition.

Claims

1. A topical composition comprising niacinamide and one or more types of resveratrol salicylate.

2. The aforementioned one or more types of resveratrol salicylate 3-Salicylate-5,4'-Dihydroxystilbene 5-Salicylate-3,4'-Dihydroxystilbene 4'-Salicylate-3,5-Dihydroxystilbene 3,5-disalithylate-4'-hydroxystilbene, 3,4'-disalithylate-5-hydroxystilbene, 4',5-disalithylate-3-hydroxystilbene, and A topical composition according to claim 1, selected from the group consisting of 3,5,4'-trisalicylate stilbene.

3. The topical composition according to claim 1, wherein the concentration of niacinamide is in the range of about 0.0001% to about 5% of the total weight of the composition, and the concentration of resveratrol salicylate is in the range of about 0.001% to about 5% of the total weight of the composition.

4. The topical composition according to claim 3, further comprising 0.1% to 2% trehalose and 0.1% to 2% betaine.

5. The topical composition according to claim 3, further comprising an aqueous base acceptable as a cosmetic, and optionally an oil phase.

6. The topical composition according to claim 5, wherein the composition is in the form of a water-in-oil or oil-in-water emulsion.

Images