Polypeptide cosmetic compositions and use thereof

Abstract

Peptide cosmetic ingredients can help address skin conditions such as decreased firmness, fine lines and wrinkles, and other age-related skin changes associated with general degeneration of the dermis, typically characterized by collagen loss and structural disorder. Cosmetic ingredients may contain peptides, a first peptide activator containing guanine nucleotides, and a second peptide activator containing succinic acid. Preferably, the peptide has about five amino acid residues or fewer; more preferably, the peptide contains a tripeptide having the amino acid sequence Ser-Thr-Pro-NH2. Preferred peptides can promote collagen production in skin cells after the cosmetic ingredient has been applied.

Description

[Technical Field]

[0001] This disclosure generally relates to cosmetic ingredients, and more specifically to peptide cosmetic ingredients that address skin conditions (including skin aging and appearance). It is incorporated herein by reference.

[0002] The XML sequence list (file name 2401_084296_000012.xml; created on July 9, 2025; size 2336 tuples) is also submitted hereto and its full text is incorporated into this document by reference. [Previous Technology]

[0003] The skin is an important organ composed of multiple layers (dermis, epidermis, and stratum corneum), covering the entire body surface, and has functions such as protection, sensing, immunity, metabolism, and body temperature regulation. Like other organs, the skin also ages.

[0004] The intrinsic or temporal aging of the skin and other organs is the result of genetically programmed aging and biochemical changes caused by endogenous factors. The aging process on the skin is characterized by a slowdown in the regeneration rate of cells and extracellular matrix, the gradual loss of dermal collagen and elastic fibers, which eventually leads to dermal and epidermal atrophy, dryness, reduced elasticity and firmness, fine lines and wrinkles, pigmentation or hypopigmentation blemishes, age spots, etc.

[0005] On the other hand, external aging of the skin may be caused by environmental factors such as pollution, sun exposure and other sources of ultraviolet radiation, diseases, and lifestyle. External and internal aging influence each other and affect the overall rate of skin aging.

[0006] Many consumers are concerned about the appearance of their skin and signs of aging, such as those mentioned above. Many cosmetic ingredients can be used to slow down or prevent the aging process of the skin.

[0007] Collagen plays a vital role in the extracellular matrix and is the most abundant protein in the human body, especially in the skin. Collagen can exist in fibrous or non-fibrous forms both inside and outside cells. Fibrous collagen is the most common, including collagen subtypes I, II, III, V, and XI, with subtypes I, IV, and V most frequently associated with the skin and dermis. Fibrous collagen serves as a structural support material for the skin and extracellular structures, possessing tensile strength that distinguishes it from many other types of proteins. Collagen gives the skin its shape, firmness, and strength. However, collagen production decreases with age, eventually leading to changes in the skin's appearance, such as the appearance of wrinkles. Therefore, collagen plays a crucial role for those who want to combat signs of skin aging.

[0008] Therefore, cosmetic ingredients that target improving collagen production may help address the effects of age-related collagen deficiency. [Summary of the Invention]

[0009] In some embodiments, this disclosure provides a cosmetic composition comprising: a polypeptide that promotes collagen production and comprises about 5 or fewer amino acid residues; a first polypeptide activator comprising guanine nucleotides; and a second polypeptide activator comprising succinic acid.

[0010] In some or other embodiments, this disclosure provides a cosmetic composition comprising: a polypeptide having an amino acid sequence of Ser-Thr-Pro-NH2, wherein the polypeptide is present in an amount of about 2.5 × 10⁻⁵ wt% to about 1.25 × 10⁻⁴ wt% based on the total mass of the cosmetic composition; a first polypeptide activator comprising guanine nucleotide, wherein the guanine nucleotide is present in an amount of about 1 × 10⁻³ wt% to about 3.5 × 10⁻³ wt% based on the total mass of the cosmetic composition; and a second polypeptide activator comprising succinic acid, wherein the succinic acid is present in an amount of about 5 × 10⁻⁵ wt% to about 1.5 × 10⁻⁴ wt% based on the total mass of the cosmetic composition.

[0011] In yet another embodiment, this disclosure provides a cosmetic composition comprising: a polypeptide having the amino acid sequence Ser-Thr-Pro-NH2, wherein the polypeptide is present in an amount of about 2.5 × 10⁻⁵ wt% to about 1.25 × 10⁻⁴ wt% based on the total mass of the cosmetic composition; at least one brine shrimp extract containing guanine nucleotides, wherein the guanine nucleotides are present in an amount of about 1 × 10⁻³ wt% to about 3.5 × 10⁻³ wt% based on the total mass of the cosmetic composition; and at least one lodobacter ssp extract containing succinic acid, wherein the succinic acid is present in an amount of about 5 × 10⁻⁵ wt% to about 1.5 × 10⁻⁴ wt% based on the total mass of the cosmetic composition.

Implementation Method

[0013] This disclosure generally relates to cosmetic ingredients, and more specifically to peptide cosmetic ingredients that can address skin conditions (including skin aging and appearance).

[0014] As described above, cosmetic ingredients that can improve skin appearance and delay or limit the skin aging process are likely highly desirable to consumers. Stimulating collagen production may also be desirable due to the effects of such biomaterials on the skin aging process.

[0015] This disclosure provides cosmetic compositions that address the aforementioned needs and offer related advantages. Specifically, this disclosure provides cosmetic compositions in which a polypeptide containing about five or fewer amino acid residues can interact with at least a first polypeptide activator and a second polypeptide activator to stimulate collagen production. The first polypeptide activator may contain guanine nucleotides, such as diguanosine tetraphosphate, while the second polypeptide activator may contain succinic acid. Surprisingly, the combination of the first and second polypeptide activators interacts with the polypeptide, resulting in a synergistic increase in collagen production compared to using the polypeptide alone (i.e., when the same or similar amounts of the polypeptide are present alone). The term "synergistic interaction" refers to the synergistic effect of the polypeptide with the first and second polypeptide activators, leading to an increase in collagen production greater than the effect of any one or two components of the cosmetic composition acting alone at the same or similar concentrations. That is, the increase in collagen production exceeds the sum of the individual contributions of the first and second polypeptide activators. The advantage is that the synergistic interactions can be achieved within the known concentration range of the individual components of the cosmetic ingredients disclosed in this paper, as described below.

[0016] The peptides contained in the cosmetic ingredients disclosed herein may contain five or fewer amino acid residues, such as three, four, or five amino acid residues, preferably three or four amino acid residues, and most preferably three amino acid residues (tripeptides). Suitable tripeptides may already possess collagen-promoting activity and, as described herein, exhibit increased collagen production when synergistically interacting with a first peptide activator and a second peptide activator. Preferably, the peptides may be ingredients listed in INCI (International Nomenclature for Cosmetic Ingredients).

[0017] In a specific example, the polypeptide may have the amino acid sequence Ser-Thr-Pro-NH2, wherein the C-terminal carboxyl group is replaced with carboxylamine. A more common name for this polypeptide is its INCI name, tripeptide-32. Tripeptide-32 is known to promote collagen production when applied topically to the skin. Another polypeptide suitable for stimulating collagen production may be a tetrapeptide having SEQ ID. NO. 1 (Ser-Pro-Leu-Gln-NH2), with the INCI name tetrapeptide-26. The C-terminal carboxyl group in tetrapeptide-26 is also replaced with carboxylamine. Tripeptide-32 and tetrapeptide-26 are known to mildly stimulate collagen production when used alone. For example, compositions containing tripeptide-32 and tetrapeptide-26, which stimulate collagen production, are described in U.S. Patent 10,149,811, which is incorporated herein by reference. In addition to promoting collagen production, tripeptide-32 and tetrapeptide-26 can also stimulate the circadian locomotor output cycle (CLOCK) gene and cycle homolog 1 (PER1) in keratinocytes. The preferred peptide used in this study is tripeptide-32.

[0018] Surprisingly, the cosmetic composition disclosed herein exhibits a synergistic enhancement in collagen production compared to using the peptide alone at the same or substantially the same peptide concentration. For example, when tripeptide-32 is combined with either a first peptide activator (a composition containing guanine nucleotides) or a second peptide activator (a composition containing succinic acid) alone, collagen production is hardly enhanced or not enhanced at all compared to using tripeptide-32 alone at the same or substantially the same peptide concentration. Conversely, when tripeptide-32 is combined with the first and second peptide activators in appropriate amounts, a significant increase in collagen production is achieved. The increased collagen production when both the first and second peptide activators are present in combination with the peptide indicates a synergistic interaction between them and the peptide.

[0019] Therefore, the cosmetic composition disclosed herein may include: a polypeptide that promotes collagen production and contains about 5 or fewer amino acid residues; a first polypeptide activator containing guanine nucleotides; and a second polypeptide activator containing succinic acid. The first and second polypeptide activators may interact with the polypeptide to produce a synergistic effect on collagen production compared to using the polypeptide alone.

[0020] The polypeptide may be present in the cosmetic composition in an amount ranging from about a1 to about a2 wt% based on the total mass of the cosmetic composition, wherein a1 < a2, and a1 and a2 are independently selected from 2.5×10-6, 5×10-6, 7.5×10-6, 1×10-5, 1.25×10-5, 1.5×10-5, 1.75×10-5, 2×10-5, 2.25×10-5, 2.5×10-5, 3.75×10-5, 5×10-5, 6.25×10-5, 7.5×10-5, 8.75×10-5, 1×10-4, 1.125×10-4, or 1.25×10-4. In non-limiting examples, the peptide may be present in the cosmetic composition in an amount ranging from about 2.5 × 10⁻⁵ wt% to about 1.25 × 10⁻⁴ wt%, or from about 3.75 × 10⁻⁵ wt% to about 7.5 × 10⁻⁵ wt%, or from about 4.5 × 10⁻⁵ wt% to about 6 × 10⁻⁵ wt%, based on the total mass of the cosmetic composition. In some examples, the peptide may be present in an amount of about 5 × 10⁻⁵ wt%, based on the total mass of the cosmetic composition. Tripeptide-32 may be present in any of the foregoing wt% ranges.

[0021] The first polypeptide activator may comprise a nucleotide, preferably a guanine nucleotide. The term "guanine nucleotide" refers to a compound containing a guanine nucleobase, a sugar (ribose or deoxyribose), and a monophosphate or polyphosphate group. Guanine nucleotides may comprise diguanosine tetraphosphate, wherein the two guanine nucleobases are linked together by a monophosphate or polyphosphate group. In some instances, guanine nucleotides may comprise diguanosine tetraphosphate (DGTP) (CAS No. 4130-19-2, also known as diguanosine tetraphosphate).

[0022] In some instances, guanine nucleotides may be present in at least one artemia extract, such as Artemia salina extract. The artemia extract used may provide the following amounts of guanine nucleotides.

[0023] The second polypeptide activator may comprise succinic acid, folic acid, or any combination thereof. Preferably, the second polypeptide activator comprises succinic acid.

[0024] In some instances, succinic acid may be present in at least one lodobacter SSP extract. The amount of lodobacter SSP extract used may provide the following amounts of succinic acid.

[0025] The first polypeptide activator and the second polypeptide activator may be present in appropriate amounts and / or in appropriate proportions to each other to promote synergistic interactions with the polypeptide and stimulate collagen production to the desired extent.

[0026] The first polypeptide activator may be present in the cosmetic composition in a certain amount based on the total mass of the cosmetic composition, such that guanine nucleotides are present in an amount ranging from about b1 to about b2 wt%, wherein b1 < b2, and b1 and b2 are independently selected from 1×10-3, 1.25×10-3, 1.5×10-3, 1.75×10-3, 2×10-3, 2.25×10-3, 2.5×10-3, 2.75×10-3, 3×10-3, 3.25×10-3, or 3.5×10-3. In non-limiting examples, guanine nucleotides may be present in the cosmetic composition in amounts ranging from about 1 × 10⁻³ wt% to about 3.5 × 10⁻³ wt%, or from about 1.5 × 10⁻³ wt% to about 3 × 10⁻³ wt%, or from about 2 × 10⁻³ wt% to about 3 × 10⁻³ wt%, based on the total mass of the cosmetic composition. In some examples, guanine nucleotides may be present in amounts ranging from about 2.5 × 10⁻³ wt%, based on the total mass of the cosmetic composition. Diguanosine tetraphosphate may be present in any of the aforementioned wt% ranges.

[0027] Based on the total mass of the cosmetic composition, the second polypeptide activator may be present in a certain amount in the cosmetic composition, such that succinic acid is present at about C1 to about C2. The quantity exists within the range of wt%, where c1 < c2, and c1 and c2 are independently selected from 5×10⁻⁵, 5.5×10⁻⁵, 6×10⁻⁵, 6.5×10⁻⁵, 7×10⁻⁵, 7.5×10⁻⁵, 8×10⁻⁵, 8.5×10⁻⁵, 9×10⁻⁵, 9.5×10⁻⁵, 1.0×10⁻⁴, 1.05×10⁻⁴, 1.1×10⁻⁴, 1.15×10⁻⁴, 1.2×10⁻⁴, 1.25×10⁻⁴, 1.3×10⁻⁴, 1.35×10⁻⁴, 1.4×10⁻⁴, 1.45×10⁻⁴, or 1.5×10⁻⁴. In non-limiting examples, succinic acid may be present in the cosmetic composition in amounts ranging from about 5 × 10⁻⁵ wt% to about 1.5 × 10⁻⁴ wt%, or about 7 × 10⁻⁵ wt% to about 1.3 × 10⁻⁴ wt%, or about 9 × 10⁻⁵ wt% to about 1.1 × 10⁻⁴ wt%, based on the total mass of the cosmetic composition. In some examples, succinic acid may be present in amounts ranging from about 1 × 10⁻⁴ wt%, based on the total mass of the cosmetic composition.

[0028] In a more specific example, the cosmetic composition may contain about 2.5 × 10⁻⁵ wt% to about 1.25 × 10⁻⁴ wt% of a polypeptide (e.g., tripeptide-32), about 5 × 10⁻⁵ wt% to about 1.5 × 10⁻⁴ wt% of succinic acid, and about 1 × 10⁻³ wt% to about 3.5 × 10⁻³ wt% of guanine nucleotides (e.g., diguanosine tetraphosphate), each based on the total mass of the cosmetic composition. In yet more specific examples, the cosmetic composition may contain about 5 × 10⁻⁵ wt% of a polypeptide (e.g., tripeptide-32) and / or about 1 × 10⁻⁴ wt% of succinic acid and / or about 2.5 × 10⁻³ wt% of guanine nucleotides (e.g., diguanosine tetraphosphate), each based on the total mass of the cosmetic composition.

[0029] In some or other more specific instances, the cosmetic composition may comprise: a polypeptide (e.g., tripeptide-32 having the amino acid sequence Ser-Thr-Pro-NH2), the polypeptide being present in amounts from about 2.5 × 10⁻⁵ wt% to about 1.25 × 10⁻⁴ wt% of the total mass of the cosmetic composition; at least one brine shrimp extract containing guanine nucleotides, the guanine nucleotides being present in amounts from about 1 × 10⁻³ wt% to about 3.5 × 10⁻³ wt% of the total mass of the cosmetic composition; and at least one lodobacter ssp extract containing succinic acid, the succinic acid being present in amounts from about 5 × 10⁻⁵ wt% to about 1.5 × 10⁻⁴ wt% of the total mass of the cosmetic composition.

[0030] The polypeptide (e.g., tripeptide-32) and guanine nucleotide (e.g., diguanosine tetraphosphate) may be present in various ratios (wt:wt). In non-limiting examples, the wt:wt ratio of the polypeptide to the guanine nucleotide may be in the range of about 1:8 to about 1:1400, or about 1:300 to about 1:10, or about 1:30 to about 1:100, or about 1:30 to about 1:60. In some examples, the wt:wt ratio may be about 1:50.

[0031] The polypeptide (e.g., tripeptide-32) and succinic acid may be present in various ratios (wt:wt). In non-limiting examples, the wt:wt ratio of the polypeptide to succinic acid may be in the range of about 2:1 to about 1:60, or about 1:1 to about 1:5, or about 1:1 to about 1:3. In some examples, the wt:wt ratio may be about 1:2.

[0032] Guanine nucleotides (e.g., diguanosine tetraphosphate) and succinic acid may be present in various proportions (wt:wt). In non-limiting examples, the wt:wt ratio of guanine nucleotides (e.g., diguanosine tetraphosphate) to succinic acid may be in the range of about 6.7:1 to about 70:1, or about 50:1 to about 20:1. In some examples, the wt:wt ratio may be about 25:1.

[0033] The cosmetic composition disclosed herein may be in any liquid, semi-solid, or solid form, wherein a suitable liquid form may be an emulsion, solution, or suspension, such as an aqueous emulsion or solution. A suitable liquid form may contain water of about 5 wt% to about 99.9 wt% by weight of the total mass of the cosmetic composition.

[0034] It should be understood that additional ingredients may also be present in the cosmetic compositions disclosed herein, such additional ingredients not interfering with the ability of the peptide to promote collagen production and / or the synergistic interactions between the peptide and the first and second peptide activators. Non-limiting examples of such additional ingredients may include, for example, preservatives, antioxidants, pH adjusters, solvents, oils, moisturizers, surfactants, vitamins, antioxidants, fragrances, minerals, microparticles, and inert ingredients (such as polysiloxanes). Those skilled in the art of cosmetic formulation are familiar with examples of such additional ingredients and their appropriate amounts.

[0035] Therefore, this disclosure provides a cosmetic composition comprising: a polypeptide that promotes collagen production; and a first polypeptide activator and a second polypeptide activator, which work together with the polypeptide to increase the synergistic effect of collagen production compared to using the polypeptide alone.

[0036] The cosmetic ingredients disclosed herein can be applied topically to the skin as part of a user's daily skincare routine. Optionally, the user may have previously determined that they have insufficient collagen synthesis, or suspect that they have such a deficiency. The cosmetic ingredients can be applied to the desired areas of the skin once or more daily, such as after cleansing the skin in the morning and evening.

[0037] The embodiments disclosed herein include the following:

[0038] Example 1. A cosmetic composition comprising: a polypeptide that promotes collagen production and contains about 5 or fewer amino acid residues; a first polypeptide activator that contains guanine nucleotides; and a second polypeptide activator that contains succinic acid.

[0039] Example 2. A cosmetic composition as in Example 1, wherein the polypeptide is a tripeptide.

[0040] Example 3. A cosmetic composition as in Example 2, wherein the polypeptide has an amino acid sequence of Ser-Thr-Pro-NH2.

[0041] Example 4. A cosmetic composition as described in any one of Examples 1 to 3, wherein the polypeptide is present in an amount of about 2.5 × 10⁻⁵ wt% to about 1.25 × 10⁻⁴ wt% based on the total mass of the cosmetic composition.

[0042] Example 5. A cosmetic composition as described in any one of Examples 1 to 4, wherein the succinic acid is present in an amount of about 5 × 10⁻⁵ wt% to about 1.5 × 10⁻⁴ wt% based on the total mass of the cosmetic composition.

[0043] Example 6. A cosmetic composition as described in any one of Examples 1 to 5, wherein the guanine nucleotide comprises diguanosine nucleotide.

[0044] Example 7. A cosmetic composition as described in any one of Examples 1 to 6, wherein the guanine nucleotide comprises diguanosine tetraphosphate.

[0045] Example 8. A cosmetic composition as described in any one of Examples 1 to 7, wherein the guanine nucleotide is present in an amount of about 1 × 10⁻³ wt% to about 3.5 × 10⁻³ wt% based on the total mass of the cosmetic composition.

[0046] Example 9. A cosmetic composition as described in any one of Examples 1 to 8, wherein, based on the total mass of the cosmetic composition, the polypeptide is present in an amount of about 2.5 × 10⁻⁵ wt% to about 1.25 × 10⁻⁴ wt%, the succinic acid is present in an amount of about 5 × 10⁻⁵ wt% to about 1.5 × 10⁻⁴ wt%, and the guanine nucleotide is present in an amount of about 1 × 10⁻³ wt% to about 3.5 × 10⁻³ wt%.

[0047] Example 10. A cosmetic composition as described in any one of Examples 1 to 8, wherein the polypeptide is present at about 5 × 10⁻⁵ wt% based on the total mass of the cosmetic composition.

[0048] Example 11. A cosmetic composition as described in any one of Examples 1 to 10, wherein the polypeptide is present at about 5 × 10⁻⁵ wt% based on the total mass of the cosmetic composition.

[0049] Example 12. A cosmetic composition as described in any one of Examples 1 to 10, wherein the succinic acid is present at about 1 × 10⁻⁴ wt% based on the total mass of the cosmetic composition.

[0050] Example 13. A cosmetic composition as described in any one of Examples 1 to 10, wherein the guanine nucleotide is present at about 2.5 × 10⁻³ wt% based on the total mass of the cosmetic composition.

[0051] Example 14. A cosmetic composition as described in any one of Examples 1 to 13, wherein the guanine nucleotide is present in at least one brine shrimp extract.

[0052] Example 15. A cosmetic composition as in Example 14, wherein the at least one brine shrimp extract contains at least an Artemia salina extract.

[0053] Example 16. A cosmetic composition as described in any one of Examples 1 to 15, wherein the succinic acid is present in at least one lodobacter ssp extract.

[0054] Example 17. A cosmetic composition comprising: a polypeptide having an amino acid sequence of Ser-Thr-Pro-NH2, wherein the polypeptide is present in an amount of about 2.5 × 10⁻⁵ wt% to about 1.25 × 10⁻⁴ wt% based on the total mass of the cosmetic composition; a first polypeptide activator comprising guanine nucleotide, wherein the guanine nucleotide is present in an amount of about 1 × 10⁻³ wt% to about 3.5 × 10⁻³ wt% based on the total mass of the cosmetic composition; and a second polypeptide activator comprising succinic acid, wherein the succinic acid is present in an amount of about 5 × 10⁻⁵ wt% to about 1.5 × 10⁻⁴ wt% based on the total mass of the cosmetic composition.

[0055] Example 18. The composition as in Example 17, wherein the synergistic effect of the first polypeptide activator and the second polypeptide activator on the interaction of the polypeptide to produce collagen production is increased relative to the polypeptide alone.

[0056] Example 19. A cosmetic composition as in Example 17 or Example 18, wherein the guanine nucleotide comprises diguanosine tetraphosphate.

[0057] Example 20. A cosmetic composition comprising: a polypeptide having an amino acid sequence of Ser-Thr-Pro-NH2, wherein the polypeptide is present in an amount of about 2.5 × 10⁻⁵ wt% to about 1.25 × 10⁻⁴ wt% based on the total mass of the cosmetic composition; at least one brine shrimp extract containing guanine nucleotides, wherein the guanine nucleotides are present in an amount of about 1 × 10⁻³ wt% to about 3.5 × 10⁻³ wt% based on the total mass of the cosmetic composition; and at least one lodobacter ssp extract containing succinic acid, wherein the succinic acid is present in an amount of about 5 × 10⁻⁵ wt% to about 1.5 × 10⁻⁴ wt% based on the total mass of the cosmetic composition.

[0058] Example 21. A method comprising: applying a cosmetic composition as described in any one of Examples 1 to 20 to a skin surface; and stimulating collagen production in the skin surface.

[0059] To facilitate a clearer understanding of this disclosure, the following examples of preferred or representative embodiments are provided. These examples should not be construed as limiting or defining the scope of the invention. Examples

[0060] Therapeutic composition: The therapeutic composition is prepared into an aqueous solution containing approximately 5 × 10⁻⁵ wt% of a tripeptide with the sequence Ser-Thr-Pro-NH₂ (INCI name, tripeptide-32), approximately 1 × 10⁻⁴ wt% of succinic acid, and approximately 2.5 × 10⁻³ wt% of diguanosine tetraphosphate, based on the total weight of the treated composition. Succinic acid was obtained from an extract of lodobacter ssp. sprayed onto maltodextrin and then dissolved in water. Diguanosine tetraphosphate (DGTP) was obtained from an extract of brine shrimp.

[0061] Cell seeding: Normal human dermal fibroblasts (NHDF) were obtained from two donors (61 and 62 years old) and seeded at a density of 50,000 cells / well onto a 48-well plate and cultured overnight under standard conditions (37°C / 5% CO2). NHDF were grown in DMEM medium supplemented with 10% fetal bovine serum.

[0062] Preparation and treatment: After the culture is completed, the culture medium is removed from the wells and replaced with fresh culture medium, a reference compound (positive control), the above-mentioned therapeutic composition, individual components of the therapeutic composition, or a mixture of components of the therapeutic composition. After loading, the 48-well plate is then incubated under standard conditions (37°C and 5% CO2) for 72 hours.

[0063] Cell counting: After culturing for 72 hours and collecting the supernatant, cell counting was performed to determine cell viability. Cell counting was performed using a Cellaca high-throughput cell counter.

[0064] Collagen ELISA: Collagen I production was assessed using the Pro Collagen Type I ELISA (Takara MK101) in accordance with the supplier's specifications (Takara Bio, MK101). Optical density (OD) was measured at 450 nm.

[0065] Statistical Analysis: Statistical analysis was performed on all samples using one-way variance analysis and the Dunnett post-hoc test in GraphPad Prism 9. Error bars were calculated from the raw data using Microsoft Excel.

[0066] Results: Figure 1 is a bar graph showing the amount of collagen production promoted by tripeptide-32 and the therapeutic component compared to the blank culture medium control. The results in Figure 1 are averages for subjects aged 61 and 62 years and normalized to mean cell count. The percentage changes of tripeptide-32 and the therapeutic component in Figure 1 are calculated relative to the blank culture medium control. As shown, tripeptide-32 alone increased collagen production by 74% relative to the control group, while the therapeutic component more than doubled collagen production by 134% relative to the control group. Therefore, tripeptide-32 promotes more collagen production in the presence of succinic acid and DGTP than when used alone in the same amount.

[0067] Figure 2 is a bar graph showing the collagen production promoted by the treatment composition, individual components of the treatment composition, and selected mixtures of the treatment composition compared to the blank culture medium control and L-ascorbic acid reference (18 µg / mL, positive control). The individual components or mixtures of components tested in Figure 2 were prepared at the same concentrations present in the treatment composition. The results in Figure 2 are averages for 61- and 62-year-old subjects and are normalized to mean cell count. The percentage changes in Figure 2 are calculated relative to the blank culture medium control. As shown, compared to the control group, the treatment composition again more than doubled collagen production (an increase of 134%), while the same concentration of tripeptide-32 alone only caused a milder increase of 74%.

[0068] Referring again to Figure 2, compared with tripeptide-32, individual components of the treatment composition promoted less collagen production. The increase in collagen production from succinic acid alone was approximately half that of tripeptide-32, while the increase in collagen production from DGTP was essentially the same as that from the control group. Furthermore, the amount of collagen production promoted by the combination of DGTP and succinic acid was essentially the same as that promoted by succinic acid alone. These results indicate that succinic acid and DGTP can increase collagen production through a synergistic interaction with tripeptide-32.

[0069] The potential synergistic interaction between succinic acid and DGTP was further investigated by testing the effects of additional combinations of the components of the therapeutic composition on collagen production. Figure 3 is a bar graph showing the amount of collagen production promoted by the therapeutic composition, individual components of the therapeutic composition, and mixtures of the components of the therapeutic composition compared to the blank culture medium control and L-ascorbic acid reference (18 µg / mL, positive control). The individual components or mixtures of components in Figure 3 were prepared at the same concentrations present in the therapeutic composition. The results in Figure 3 used only NHDF obtained from 62-year-old subjects and were again normalized to mean cell count. The percentage changes in Figure 3 are calculated relative to the blank culture medium control. In this case, individual components of the therapeutic composition showed minimal increase (or even decrease) in collagen production relative to the control group, and binary combinations of individual components of the therapeutic composition also promoted minimal increase (or even decrease) in collagen production relative to the control group. In contrast, the therapeutic composition containing all components and their combinations promoted a robust increase of 142% relative to the control group. The results in Figure 3 show that the combination of tripeptide-32, succinic acid and DGTP has a synergistic effect on collagen production, because these components alone or in binary combinations cannot produce a similar increase in collagen production.

[0070] All documents described herein are incorporated herein by reference in all jurisdictions where such practice is permitted, including any priority documents and / or test procedures, provided that their contents do not conflict with this document. As will be apparent from the foregoing general description and specific embodiments, while the form of this disclosure has been illustrated and described, various changes may be made without departing from the spirit and scope of this disclosure. Therefore, it is not intended to limit this disclosure. For example, the composition described herein may not contain any component or composition not expressly set forth or disclosed herein. Any method may lack any step not set forth or disclosed herein. Similarly, the term "comprising" is considered synonymous with the term "including". Whenever a constituent, element, or group of elements is preceded by the conjunction "consisting of," it should be understood that we also consider the same constituent or group of elements that are preceded by the transitional conjunctions "consisting essentially of," "consisting of," "selected from the group consisting of," or "is," and vice versa.

[0071] Unless otherwise indicated, all numerical values ​​used in this specification and related claims to indicate the amount, properties (such as molecular weight), reaction conditions, etc., of components should be understood to be modified by the term "about" in all cases. Therefore, unless otherwise indicated to the contrary, the numerical parameters set forth in the following specification and the appended claims are approximate values ​​and may vary depending on the desired properties sought to be obtained in embodiments of the invention. To a minimum, and not in an attempt to limit the application of the equivalence principle to the claims, each numerical parameter should be interpreted at least based on the number of significant figures reported and by applying general rounding techniques.

[0072] Whenever a range of values ​​with a lower and upper limit is disclosed, any value falling within that range and any range included therein are specifically disclosed. Specifically, the range of each value disclosed herein (in the form of "from about a to about b" or equivalently "from about a to b" or equivalently "from about a ab") should be understood to list each value and range covered within a broader range of values. Moreover, unless otherwise explicitly and clearly defined by the patent holder, the terms used in the claims have their general, ordinary meaning. Furthermore, the indefinite article "a / an" used in the claims is limited herein to mean one or more of the elements it introduces.

[0073] This document presents one or more illustrative embodiments. For clarity, not all features of the entity implementations are described or shown in this application. It should be understood that in the development of the entity implementations disclosed herein, many implementation-specific decisions must be made to achieve the developer's objectives, such as complying with system-related, business-related, governmental-related restrictions and other limitations, which may vary depending on the implementation and time. While the developer's efforts may be quite time-consuming, such efforts will be routine for those skilled in the art and who benefit from this disclosure.

[0074] Therefore, this disclosure is fully applicable to achieving the mentioned and inherent purposes and advantages. The specific embodiments disclosed above are merely illustrative, as this disclosure can be modified and practiced in different but equivalent ways, as will be apparent to those skilled in the art and who benefit from the teachings herein. Furthermore, the details of the constructions or designs shown herein are not intended to be limited except as described in the following claims. It will be apparent that the specific illustrative embodiments disclosed above can be changed, combined, or modified, and all such changes are considered to be within the spirit and scope of this disclosure. The illustrative embodiments disclosed herein may be suitably practiced in the absence of any elements not specifically disclosed herein and / or any optional elements disclosed herein.

[0075] The dimensions and values ​​disclosed herein should not be construed as strictly limited to the exact numerical values ​​listed. Rather, unless otherwise specified, each of these dimensions refers to the listed value and the functionally equivalent range around that value. For example, a dimension disclosed as "40 mm" is intended to mean "approximately 40 mm".

[0076] Every document cited herein, including any cross-reference or related patent or application, is incorporated herein by reference in its entirety unless expressly excluded or otherwise limited. No reference to any document is an admission that it is prior art with respect to any invention disclosed or claimed herein, nor is it an admission that it, alone or in any combination of any other reference, teaches, implies, or discloses any such invention. Furthermore, in the event of any conflict between the meaning or definition of any term used in this document and the meaning or definition of the same term in any document incorporated herein by reference, the meaning or definition assigned to the term in this document shall prevail.

[0077] Although specific embodiments of the invention have been illustrated and described, it will be apparent to those skilled in the art that various other changes and modifications can be made without departing from the spirit and scope of the invention. Therefore, the appended claims are intended to cover all such changes and modifications falling within the scope of the invention. [Simplified Explanation of the Diagram]

[0012] The following figures are used to illustrate certain aspects of this disclosure and should not be considered as the only embodiments. Those skilled in the art will understand, upon referring to this disclosure, that the disclosed technical content can be modified, altered, combined, and equivalently substituted in form and function. [Figure 1] is a bar graph showing the amount of collagen production promoted by tripeptide-32 and the therapeutic composition of this example. [Figure 2] is a bar graph showing the amount of collagen production promoted by the therapeutic composition of this example, individual components of the therapeutic composition, and selected mixtures of the components of the therapeutic composition. [Figure 3] is a bar graph showing the amount of collagen production promoted by the therapeutic composition of this example, individual components of the therapeutic composition, and mixtures of the components of the therapeutic composition.

Claims

1. A cosmetic ingredient comprising: a polypeptide that promotes collagen production and contains about 5 or fewer amino acid residues; a first polypeptide activator that contains guanine nucleotides; and a second polypeptide activator that contains succinic acid, wherein the polypeptide is a tripeptide and has the amino acid sequence Ser-Thr-Pro-NH2.

2. The cosmetic composition of claim 1, wherein the polypeptide is present in an amount of 2.5 × 10⁻⁵ wt% to 1.25 × 10⁻⁴ wt% based on the total mass of the cosmetic composition.

3. The cosmetic composition of claim 1, wherein the succinic acid is present at a concentration of 5 × 10⁻⁵ wt% to 1.5 × 10⁻⁴ wt% based on the total mass of the cosmetic composition.

4. The cosmetic composition of claim 1, wherein the guanine nucleotide comprises diguanosine nucleotide.

5. The cosmetic composition of claim 4, wherein the guanine nucleotide comprises diguanosine tetraphosphate.

6. The cosmetic composition of claim 1, wherein the guanine nucleotide is present at a concentration of 1 × 10⁻³ wt% to 3.5 × 10⁻³ wt% based on the total mass of the cosmetic composition.

7. The cosmetic composition of claim 1, wherein, based on the total mass of the cosmetic composition, the polypeptide is present at 2.5 × 10⁻⁵ wt% to 1.25 × 10⁻⁴ wt%, the succinic acid is present at 5 × 10⁻⁵ wt% to 1.5 × 10⁻⁴ wt%, and the guanine nucleotide is present at 1 × 10⁻³ wt% to 3.5 × 10⁻³ wt%.

8. The cosmetic composition of claim 7, wherein the polypeptide is present at about 5 × 10⁻⁵ wt% based on the total mass of the cosmetic composition.

9. The cosmetic composition of claim 7, wherein the succinic acid is present at about 1 × 10⁻⁴ wt% based on the total mass of the cosmetic composition.

10. The cosmetic composition of claim 7, wherein the guanine nucleotide is present at about 2.5 × 10⁻³ wt% based on the total mass of the cosmetic composition.

11. The cosmetic composition of claim 1, wherein the guanine nucleotide is present in at least one brine shrimp extract.

12. The cosmetic composition of claim 11, wherein the at least one brine shrimp extract contains at least one Artemia salina extract.

13. The cosmetic composition of claim 1, wherein the succinic acid is present in at least one lodobacter ssp extract.

14. A cosmetic composition comprising: a polypeptide having the amino acid sequence Ser-Thr-Pro-NH2, wherein the polypeptide is present in an amount of 2.5 × 10⁻⁵ wt% to 1.25 × 10⁻⁴ wt% based on the total mass of the cosmetic composition; a first polypeptide activator comprising guanine nucleotide, wherein the guanine nucleotide is present in an amount of 1 × 10⁻³ wt% to 3.5 × 10⁻³ wt% based on the total mass of the cosmetic composition; and a second polypeptide activator comprising succinic acid, wherein the succinic acid is present in an amount of 5 × 10⁻⁵ wt% to 1.5 × 10⁻⁴ wt% based on the total mass of the cosmetic composition.

15. The cosmetic composition of claim 14, wherein the first peptide activator and the second peptide activator interact with the peptide to produce a synergistic effect on collagen production, relative to the peptide alone.

16. The cosmetic composition of claim 14, wherein the guanine nucleotide comprises diguanosine tetraphosphate.

17. A cosmetic composition comprising: a polypeptide having the amino acid sequence Ser-Thr-Pro-NH2, wherein the polypeptide is present in an amount of 2.5 × 10⁻⁵ wt% to 1.25 × 10⁻⁴ wt% based on the total mass of the cosmetic composition; at least one brine shrimp extract containing guanine nucleotides, wherein the guanine nucleotides are present in an amount of 1 × 10⁻³ wt% to 3.5 × 10⁻³ wt% based on the total mass of the cosmetic composition; and at least one lodobacter ssp extract containing succinic acid, wherein the succinic acid is present in an amount of 5 × 10⁻⁵ wt% to 1.5 × 10⁻⁴ wt% based on the total mass of the cosmetic composition.

18. Use of a cosmetic composition as claimed in claim 1 in the manufacture of a medicine that stimulates collagen production in the skin surface.

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