HK1237357B - Glycopeptide compositions and uses thereof - Google Patents

Description

Glycosylated peptide composition and use thereof

CROSS-REFERENCE TO RELATED APPLICATIONS

This application claims the benefit of priority to U.S. Provisional Patent Application No. 61/914,309, filed December 2013, the contents of which are incorporated herein by reference.

Technical Field

The present disclosure relates to the glycosylation of oligopeptides consisting of 5 to 6 amino acids for use in cosmetic and dermatological compositions and applications.

Background Art

The concern about skin aging is becoming more and more important in the field of dermatology and cosmetic medicine. Skin aging is partly caused by the reduction of extracellular matrix (ECM) components (including collagen, elastin and fibronectin) (Callaghan, TM and Wilhelm, KP (2008) Int J. Cosmet. Sci. 30: 313-322). Type I collagen is the most abundant type of collagen and plays a role in structural integrity, cell adhesion and migration, tissue remodeling and wound healing (Trojanowska, M et al., (1998) J. Mol. Med. 76: 266-267; Chiquet, M. et al., (1996) Biochem. Cell Biol. 74: 737-744). In 1993, a pentapeptide derived from the carboxyl terminus of type I procollagen (Lys-Thr-Thr-Lys-Ser, also known as KTTKS SEQ ID NO: 1) was shown to promote ECM production (Katayama, K et al., (1993) J. Biol. Chem. 268: 9941-9944). In an in vitro model using human lung fibroblasts, the pentapeptide was found to significantly increase ECM proteins by increasing the synthesis of type I and type III collagen and fibronectin.

Although the in vitro data mentioned above demonstrate the ability of the pentapeptide to stimulate collagen and fibronectin synthesis, its electrostatic charge poses a challenge for transdermal penetration. To enhance the activity and stability of the peptide and to increase its lipophilicity, Lintner et al. (Lintner (2003) US 6,620,419) conjugated the pentapeptide to a 16-carbon fatty acid (palmitic acid), thereby creating palmitoyl KTTKS (Pal-KTTKS, SEQ ID NO: 2). This fatty acid modification allows for more efficient delivery through the skin relative to KTTKS alone (Lintner, K (2002) Ann Dermatol Venereol 129: 1S 105; Mas-Chamberlin, C et al., (2002) Ann Dermatol Venereol 129: 1S 456; Tsai, TC and Hantash, BM (2008) Clin Med 2: 1-20). Palmitoyl-KTTKS was one of the first oligopeptides developed as a cosmetic agent and is currently the active ingredient in the cosmetic agent Matrixyl ^™ (Sederma SAS). Pal-KTTKS is formulated with a skin care active and dermatologically acceptable carrier to form a skin care composition for topical treatment of the skin (The Proctor and Gamble Company). The resulting composition has been found to be particularly beneficial for treating wrinkle formation and other effects of skin aging (Robinson (2002) US Pat. No. 6,492,326). Pal-KTTKS has been found to reduce fibroblast viability under certain conditions.

It would be desirable to provide alternative oligopeptides that are capable of promoting the production and secretion of ECM proteins by fibroblasts.

SUMMARY OF THE INVENTION

Generally, the present invention discloses a glycosylated oligopeptide of 5 or 6 amino acids having the following sequence: palmitoyl-X1-Lys-X2-X3-Lys-X4-OH, wherein X1 is absent, Ser with a glycosylated side chain, or Asn with a glycosylated side chain; X2 is Thr, Thr with a glycosylated side chain, Asn with a glycosylated side chain, or Ser with a glycosylated side chain; X3 is Thr, or Thr with a glycosylated side chain; X4 is S er, or Ser with a glycosylated side chain, wherein: at least one of X1, X2, X3, and X4 is an amino acid with a glycosylated side chain; wherein: each glycosylated side chain is independently glycosylated with a carbohydrate selected from the group consisting of glucose, N-acetyl-glucosamine, galactose, N-acetyl-galactosamine, mannose, maltose, lactose, rhamnose, cellobiose, xylose, and trehalose; and wherein: each hydroxyl group on the carbohydrate is independently OH or acetylated. The sequence of the glycosylated peptide is defined as SEQ ID NO: 3.

The hydroxyl groups on the carbohydrates may all be acetylated.

The hydroxyl groups on the carbohydrates may not be acetylated.

The amino acid at position X1 may be absent.

Preferably, at least one amino acid is glycosylated with maltose, glucose, galactose or mannose.

The amino acid at X2 can be Thr with a glycosylated side chain. The glycosylated side chain can be glycosylated with glucose or maltose. The amino acids at X1, X3, and X4 can all be non-glycosylated.

The amino acid at position X4 may be Ser with a glycosylated side chain. The glycosylated side chain may be glycosylated with glucose or galactose. The amino acids at positions X1, X2, and X3 may be non-glycosylated.

In a specific example, a glycosylated oligopeptide having the following sequence is provided: Palmitoyl-Lys-Thr*-Thr-Lys-Ser-OH, wherein Thr* is a threonine glycosylated with peracetylated β-glucose (SEQ ID NO: 4).

Another exemplary oligopeptide disclosed in accordance with the present invention has the following sequence: Palmitoyl-Lys-Thr*-Thr-Lys-Ser-OH, wherein Thr* is a threonine glycosylated with a fully acetylated α-mannose (SEQ ID NO: 5).

In another specific example, a glycosylated oligopeptide having the following sequence is provided: Palmitoyl-Lys-Thr*-Thr-Lys-Ser-OH, wherein Thr* is threonine glycosylated with peracetylated β-maltose (SEQ ID NO: 6).

In another specific example, a glycosylated oligopeptide having the following sequence is provided: Palmitoyl-Lys-Thr-Thr-Lys-Ser*-OH, wherein Ser* is serine glycosylated with peracetylated β-galactose (SEQ ID NO: 7).

In another specific example, a glycosylated oligopeptide having the following sequence is provided: Palmitoyl-Lys-Thr-Thr-Lys-Ser*-OH, wherein Ser* is serine glycosylated with peracetylated β-glucose (SEQ ID NO: 8).

In another specific example, a glycosylated oligopeptide having the following sequence is provided: Palmitoyl-Lys-Thr-Thr*-Lys-Ser-OH, wherein Thr* is a threonine glycosylated with peracetylated β-maltose (SEQ ID NO: 9).

In another specific example, a glycosylated oligopeptide having the following sequence is provided: Palmitoyl-Lys-Thr-Thr-Lys-Ser*-OH, wherein Ser* is serine glycosylated with peracetylated β-maltose (SEQ ID NO: 10).

In another aspect, the present disclosure provides a cosmetic or dermopharmaceutical composition comprising at least one glycosylated oligopeptide as described above.

In another aspect, the present invention discloses a method for increasing the secretion of extracellular matrix proteins by cells in an individual. The method comprises topically administering the composition described above to the individual. The extracellular matrix protein can be elastin, fibronectin, or collagen.

In another aspect, the present invention discloses the use of the glycosylated oligopeptide described above for increasing the secretion of extracellular matrix proteins by cells. The extracellular matrix proteins may be elastin, fibronectin or collagen.

In another aspect, the present disclosure provides a skin care composition comprising: (a) a safe and effective amount of a glycosylated oligopeptide as described above; (b) a safe and effective amount of at least one other skin care active selected from the group consisting of: desquamation actives, anti-acne actives, vitamin _B3 compounds, retinoids, di-, tri-, tetra-, penta- and hexapeptides and derivatives thereof, hydroxy acids, free radical scavengers, chelating agents, anti-inflammatory agents, local anesthetics, tanning actives, skin whitening agents, slimming agents, flavonoids, antimicrobial actives, skin healing agents, antifungal actives, farnesol, phytantriol, allantoin, glucosamine and mixtures thereof; and (c) a dermatologically acceptable carrier.

The peptides disclosed herein, alone or in combination with various other substances, can be used in the following forms: solutions, suspensions, emulsions; encapsulated in carriers such as macrocapsules, microcapsules or nanocapsules, liposomes, or chylomicrons; contained in macroparticles, microparticles or nanoparticles; contained in microsponges; or adsorbed on a powdered organic polymer, talc, bentonite or another mineral support.

The peptides disclosed herein, alone or in combination with various other substances, can be used in the form of: a galenic formulation; an oil-in-water emulsion; a water-in-oil emulsion; a lotion; or a gelling, thickening, surface active or emulsifying polymer, a pomade, a lotion, a capillary, a shampoo, a soap, a powder, a patch, a pencil, a spray or a body oil.

The peptides disclosed herein, alone or in combination with various other substances, can be used with any other components conventionally used, such as extracted lipids, synthetic lipids, gelling, thickening, surface active or emulsifying polymers, water-soluble or fat-soluble active principles, plant extracts, tissue extracts, marine extracts, solar filters, or antioxidants.

The peptides disclosed herein, alone or in combination with various other substances, can be used in cosmetic applications to promote healing, hydration, or skin care, particularly for wrinkle formation or exacerbation, or for natural or accelerated results of skin aging (e.g., heliodermia or pollution).

The peptides disclosed herein, alone or in combination with various other substances, can be used to prepare cosmetic or dermatological agents. The peptides or their agents can be used to promote healing, hydration, or skin care, particularly for wrinkle formation or exacerbation, or for natural or accelerated skin aging (e.g., heliodermia or pollution).

The peptides disclosed herein, alone or in combination with various other substances, can be used with a variety of other skin care actives. Exemplary skin care activities include wound healing, reducing the appearance of stretch marks, psoriasis, skin cancer, dermatitis or eczema.

Other aspects and features of the present disclosure will become apparent to those of ordinary skill in the art upon reviewing the following description of specific embodiments in conjunction with the accompanying drawings.

BRIEF DESCRIPTION OF THE DRAWINGS

Embodiments of the present disclosure will now be described, by way of example only, with reference to the accompanying drawings.

1 is a graph showing a comparison of the efficacy of exemplary glycated peptides on elastin secretion (expressed as % of control) at 48 hours (A) and 72 hours (B) after treatment of human skin fibroblasts.

2 is a graph showing a comparison of the efficacy of exemplary glycated peptides on fibronectin secretion (expressed as % of control) at 48 hours (A) and 72 hours (B) after treatment of human skin fibroblasts.

FIG3A is a graph showing the HPLC profile of the exemplary compound GP001-1 (SEQ ID NO: 4).

FIG3B is a graph showing the mass spectrum of the exemplary compound GP001-1 (SEQ ID NO: 4).

FIG4A is a graph showing the HPLC profile of exemplary compound GP002-3 (SEQ ID NO: 8).

FIG4B is a graph showing the mass spectrum of exemplary compound GP002-3 (SEQ ID NO: 8).

FIG5A is a graph showing the HPLC profile of exemplary compound GP002-7 (SEQ ID NO: 7).

FIG5B is a graph showing the mass spectrum of exemplary compound GP002-7 (SEQ ID NO: 7).

FIG6A is a graph showing the HPLC profile of exemplary compound GP003-9 (SEQ ID NO: 6).

FIG6B is a graph showing the mass spectrum of the exemplary compound GP003-9 (SEQ ID NO: 6).

FIG7A is a graph showing the HPLC profile of exemplary compound GP004-3 (SEQ ID NO: 9).

FIG7B is a graph showing the mass spectrum of exemplary compound GP004-3 (SEQ ID NO: 9).

FIG8A is a graph showing the HPLC profile of exemplary compound GP004-4 (SEQ ID NO: 10).

FIG8B is a graph showing the mass spectrum of exemplary compound GP004-4 (SEQ ID NO: 10).

FIG9 shows phase contrast images illustrating the effects of six exemplary glycosylated peptides (GP001-1, GP002-3, GP002-7, GP003-9, GP004-3, and GP004-4) and a non-glycosylated Pal-KTTKS reference peptide on cell morphology.

10A is a graph showing a comparison of the effects of six exemplary glycosylated peptides (GP001-1, GP002-3, GP002-7, GP003-9, GP004-3, and GP004-4) and a non-glycosylated Pal-KTTKS reference peptide on cell viability (measured by MTT assay) after 48 hours.

10B is a graph showing a comparison of the effects of six exemplary glycosylated peptides (GP001-1, GP002-3, GP002-7, GP003-9, GP004-3, and GP004-4) and a non-glycosylated Pal-KTTKS reference peptide on cell viability (measured by MTT assay) after 72 hours.

11A is a graph showing a comparison of the potency of six exemplary glycated peptides (GP001-1, GP002-3, GP002-7, GP003-9, GP004-3, and GP004-4) and a non-glycosylated Pal-KTTKS reference peptide on elastin secretion (expressed as % of control) in human skin fibroblasts 72 hours after treatment.

11B is a graph showing a comparison of the potency of six exemplary glycosylated peptides (GP001-1, GP002-3, GP002-7, GP003-9, GP004-3, and GP004-4) and a non-glycosylated Pal-KTTKS reference peptide on fibronectin secretion (expressed as % of control) 72 hours after treatment of human skin fibroblasts.

12 is a graph showing a comparison of the potency of three exemplary glycated peptides (GP003-9, GP004-3, and GP004-4) and a non-glycosylated Pal-KTTKS reference peptide on total collagen secretion (expressed as % of control) 72 hours after treatment of human dermal fibroblasts.

13 is a graph showing the efficacy of an exemplary glycosylated peptide GP003-9 and a non-glycosylated Pal-KTTKS reference peptide on elastin secretion (expressed as % of control) in an in vitro 3D model of human skin (composed of human epidermal keratinocytes and human dermal fibroblasts) at 72 hours post-treatment.

Figure 14 shows micrographs of hematoxylin and eosin-stained sections of an in vitro 3-D model of human skin (composed of human epidermal keratinocytes and human skin fibroblasts) showing the morphological effects of an exemplary glycosylated peptide GP003-9 and a non-glycosylated Pal-KTTKS reference peptide.

Detailed Description of the Invention

Post-translational modification of proteins and peptides by the covalent attachment of carbohydrates to specific side chains, known as glycosylation, occurs among the most important processes in regulating protein and peptide function.

Because this process varies with the type of cell, the site of glycosylation, and the size of the carbohydrate chain (polysaccharide), the process is highly regulated within the cell. In humans, the most common sites for protein glycosylation occur on asparagine (N-glycosylation) and on serine and threonine residues (O-glycosylation) using the following sugars: glucose (Glc), trehalose (Fuc), galactose (Gal), mannose (Man), N-acetylglucosamine (GlcNAc), N-acetylgalactosamine (GalNAc), and sialic acid (Sia). Glycosylation can also include glycosylation using the following sugars: lactose (Lac), rhamnose (Rha), cellobiose (Cel), and xylose (Xyl).

Glycosylation of proteins can increase protein binding and efficacy. For example, glycosylation of collagen allows collagen to bind to the phosphatidylcholine domain of membrane receptors, such as urokinase-type plasminogen activator receptor-associated protein (uPARAP/Endo180). Glycosylation of peptides can also increase the stability of the resulting peptide.

Carbohydrates are one of the main components of the skin and play a role in its homeostasis. Many carbohydrates are found on skin cells, acting as ligands for receptors and transmitters of external signals, interacting with neighboring cells. They are involved in cell proliferation and differentiation, protein synthesis, and can also form tissue structure and architectural architecture.

During the aging process, the presence of reducing sugars increases. Glycation is a natural cellular process that results from the covalent bonding of excess endogenous or exogenous reducing sugars to proteins or lipid molecules. Glycation leads to increased cross-linking of extracellular proteins (e.g., collagen, fibronectin, and elastin), causing them to become biologically dysfunctional. Glycation can form cross-links known as advanced glycation end products (AGEs) in the skin's ECM and disrupt the normal function of these biomolecules. In the absence of enzymatic control, glycation occurs in a random manner and has been shown to be associated with wrinkle formation. In the art, it is believed that reducing sugars contribute to the aging process. Glycation should not be confused with the deleterious process of glycation.

The glycosylated peptides disclosed herein can increase the stability of glycosylated peptides and/or increase their effectiveness in delaying aging. The glycosylated peptides disclosed herein can, for example, reduce the degradation of ECM proteins or induce the biosynthesis of ECM proteins. Exemplary ECM proteins include elastin, fibronectin, and collagen.

Generally, the present disclosure provides glycosylated oligopeptides having the sequence palmitoyl-X1-Lys-X2-X3-Lys-X4-OH, wherein: X1 is absent, Ser with a glycosylated side chain, or Asn with a glycosylated side chain; X2 is Thr, Thr with a glycosylated side chain, Asn with a glycosylated side chain, or Ser with a glycosylated side chain; X3 is Thr, or Thr with a glycosylated side chain; and X4 is Ser, or Ser with a glycosylated side chain. At least one of X1, X2, X3, and X4 is an amino acid with a glycosylated side chain. Each glycosylated side chain is independently glycosylated with a carbohydrate selected from the group consisting of glucose, N-acetyl-glucosamine, galactose, N-acetyl-galactosamine, mannose, maltose, lactose, rhamnose, cellobiose, xylose, and trehalose. Each hydroxyl group on the carbohydrate is independently OH or acetylated. The sequence of the glycosylated peptide is defined as SEQ ID NO:3.

In some instances, all hydroxyl groups on the carbohydrate are acetylated. Oligosaccharides having acetylated hydroxyl groups can have improved permeability through cell membranes. This improved permeability can improve delivery through the skin.

Preferred oligopeptides disclosed herein have an N-terminal palmitoyl group. Oligopeptides having an N-terminal palmitoyl group may have improved permeability through cell membranes. This improved permeability may improve delivery through the skin. Preferably, the oligopeptides disclosed herein have 5 amino acids, wherein X1 is absent.

In some preferred oligopeptides, X2 is Thr with a glycosylated side chain. The glycosylated side chain is glycosylated with glucose, mannose, or maltose. The remaining amino acids are preferably non-glycosylated.

In other preferred oligopeptides, X3 is Thr with a glycosylated side chain. The glycosylated side chain is glycosylated with maltose. The remaining amino acids are preferably non-glycosylated.

In other preferred oligopeptides, X4 is Ser with a glycosylated side chain. The glycosylated side chain is glycosylated with glucose, galactose, or maltose. The remaining amino acids are preferably non-glycosylated.

An exemplary oligopeptide disclosed in accordance with the present invention has the following sequence: Palmitoyl-Lys-Thr*-Thr-Lys-Ser-OH, wherein Thr* is a threonine glycosylated with peracetylated β-glucose (SEQ ID NO: 4).

Another exemplary oligopeptide disclosed in accordance with the present invention has the following sequence: Palmitoyl-Lys-Thr*-Thr-Lys-Ser-OH, wherein Thr* is a threonine glycosylated with a fully acetylated α-mannose (SEQ ID NO: 5).

Another exemplary oligopeptide disclosed in accordance with the present invention has the following sequence: Palmitoyl-Lys-Thr*-Thr-Lys-Ser-OH, wherein Thr* is a threonine glycosylated with peracetylated β-maltose (SEQ ID NO: 6).

Another exemplary oligopeptide disclosed herein has the following sequence: Palmitoyl-Lys-Thr-Thr-Lys-Ser*-OH, wherein Ser* is serine glycosylated with peracetylated β-galactose (SEQ ID NO: 7).

Another exemplary oligopeptide disclosed in accordance with the present invention has the following sequence: Palmitoyl-Lys-Thr-Thr-Lys-Ser*-OH, wherein Ser* is serine glycosylated with peracetylated β-glucose (SEQ ID NO: 8).

In another specific example, a glycosylated oligopeptide having the following sequence is provided: Palmitoyl-Lys-Thr-Thr*-Lys-Ser-OH, wherein Thr* is a threonine glycosylated with peracetylated β-maltose (SEQ ID NO: 9).

In another specific example, a glycosylated oligopeptide having the following sequence is provided: Palmitoyl-Lys-Thr-Thr-Lys-Ser*-OH, wherein Ser* is serine glycosylated with peracetylated β-maltose (SEQ ID NO: 10).

Glycosylated oligopeptides disclosed in accordance with the present invention (eg, the exemplary glycosylated oligopeptides listed above) exhibit enhanced efficacy and/or reduced toxicity compared to non-glycosylated Pal-KTTKS.

In the context of the present disclosure, enhanced efficacy refers to higher induction of elastin, fibronectin and total collagen secretion in in vitro human skin fibroblast cultures compared to non-glycosylated Pal-KTTKS, as measured using the following kits: fibronectin (Takara, MK1 15), elastin (Uscn Life Science Inc., E91337Hu) and total collagen (Chondrex Inc., 9062).

In order to form a formulation that can be used as an effective skin care product, the glycosylated peptide preferably must be stable in the formulation, be largely adsorbed on the skin, and be bioactive at the target.

As used herein, the term "safe and effective amount" refers to an amount of a compound or composition that is sufficient to significantly induce a positive benefit, preferably a positive glial tissue appearance or feel benefit, including the benefits disclosed herein, alone or in combination, and is low enough to avoid adverse side effects, i.e., provides a reasonable benefit-risk ratio within the prudent judgment of the skilled artisan.

As used herein, the term "sagging" refers to loose, sagging, or similar conditions of the skin that occur as a result of loss, damage, alteration, and/or abnormality of skin elastin.

As used herein, the term "smoothing" or "softening" refers to altering the surface of keratinous tissue so that its tactile feel is improved.

"Signs of skin aging" include, but are not limited to, all outwardly noticeable and tactile manifestations of skin aging, as well as any other major or minor effects. Such signs may be induced or caused by internal or external factors, such as aging over time and/or environmental damage. These signs may result from inert processes including, but not limited to, the development of textural discontinuities such as wrinkles and deep and coarse wrinkles, striae, fissures, ridges, macropores (associated with adnexal structures such as sweat ducts, sebaceous glands, or hair follicles), unevenness or roughness, loss of skin elasticity (loss and/or inactivation of functional skin elastin), sagging (including puffiness of the eyes and cheeks), loss of skin firmness, loss of skin firmness, loss of skin resilience from deformation, discoloration (including dark circles), mottled areas, sallowness, areas of hyperpigmented skin (e.g., age spots and freckles), keratinous material, abnormal differentiation, hyperkeratosis, elastosis, collagen breakdown, and other histological changes in the stratum corneum, dermis, skin vasculature (e.g., telangiectasias or spider vessels), and subcutaneous tissue (particularly those proximal to the skin).

The glycosylated peptides disclosed herein can be used to therapeutically modulate visible and/or tactile discontinuities in mammalian skin, for example, a decrease in the apparent diameter of pores, an approximation of the apparent height of tissue immediately adjacent to the pores to that of the interadnexal skin, a more even skin tone, and/or a decrease in the length, depth, and/or other dimensions of fine lines and/or wrinkles.

The glycosylated peptides disclosed herein can also be used to regulate the condition of the skin, particularly keratinous tissue. Since various conditions can be induced or caused by a variety of factors internal and/or external to the body, it is necessary to regulate the condition of the skin (i.e., mammalian, particularly human, skin). Examples include environmental damage, radiation exposure (including ultraviolet radiation), aging, menopausal conditions (e.g., postmenopausal changes in the skin), stress, disease, and the like. For example, "regulating skin condition" includes prophylactically regulating and/or therapeutically regulating skin condition and may involve one or more of the following benefits: thickening of the skin (i.e., building up the epidermis and/or dermis and/or subcutaneous (e.g., subcutaneous fat or muscle) layers of the skin and, where applicable, the stratum corneum of the nail and hair shafts) thereby reducing skin atrophy; increasing the convolution of the dermal-epidermal boundary (also known as the epidermal projection); preventing loss of skin elasticity (loss, damage and/or inactivation of functional skin elastin), such as elastosis, sagging, loss of skin resilience due to deformation; non-melanin skin discoloration, such as dark circles, spots (e.g., uneven redness due to rosacea) (hereinafter referred to as "red spots"), sallowness (gray), discoloration due to capillary dilation or spider blood vessels.

As used herein, prophylactically regulating skin condition includes delaying, reducing, and/or preventing visible and/or tactile discontinuities in the skin (eg, textural irregularities in the skin that are detectable by sight or feel).

As used herein, therapeutically regulating skin condition includes alleviating (eg, reducing, diminishing, and/or erasing) discontinuities in the skin.

The glycosylated peptides disclosed herein can also be used to improve the appearance and/or feel of skin. For example, the compositions disclosed herein can be used to regulate the appearance of skin condition by providing an immediate, visible improvement in skin appearance after application of the composition to the skin. Generally, compositions disclosed herein that further comprise a particulate material are most useful for providing an immediate, visible improvement.

The compositions disclosed herein may provide additional benefits, including stability, lack of significant (consumer unacceptable) skin irritation, and good aesthetics.

The compositions disclosed herein are stable. The components used herein, including the glycosylated peptides disclosed herein, are stable in the compositions and are compatible with each other and with other skin care actives, such as niacinamide, phytantriol, farnesol, bisabolol, and salicylic acid. Thus, the compositions (combining oligopeptides with other skin care actives, such as niacinamide) can provide additive and/or synergistic skin benefits. Furthermore, the resulting skin care compositions have good product stability and a reasonably long shelf life.

The resulting composition (comprising a glycosylated peptide disclosed herein, in combination with at least one other skin care active) has good aesthetics. Examples of good aesthetics include compositions (e.g., luxurious creams and lotions) that are (i) refreshing and non-greasy; (ii) have a smooth, silky feel on the skin; (iii) spread easily; and/or (iv) absorb quickly. Other examples of good aesthetics include compositions that have a consumer-acceptable appearance (i.e., do not have an unpleasant odor or exhibit discoloration) and provide a good skin feel.

In preferred embodiments, the skin care product comprises from about 0.01% to about 50% of the glycosylated peptide, by weight of the composition. In more preferred embodiments, the skin care product comprises from about 0.05% to about 20% of the glycosylated peptide, by weight of the composition. In even more preferred embodiments, the skin care product comprises from about 0.1% to about 10% of the glycosylated peptide, by weight of the composition.

The glycosylated oligopeptides disclosed herein can be formulated with at least one other ingredient. In the case where the composition is to be in contact with human keratinous tissue, the other ingredients should be suitable for application on keratinous tissue, that is, when the other ingredients are introduced into the composition, they are suitable for use in contact with human keratinous tissue without causing excessive toxicity, incompatibility, instability, allergic reaction, etc. within the scope of prudent medical judgment. The CTFA Cosmetic Ingredient Handbook, Second Edition (1992) describes a variety of non-limiting cosmetic and pharmaceutical ingredients commonly used in the skin care industry (which are suitable for use in the compositions disclosed herein). Examples of classes of these components include: abrasives, absorbents, aesthetic ingredients (e.g., fragrances, pigments, coloring matter/colorants, essential oils, skin sensates), astringents, e.g., clove oil, menthol, camphor, eucalyptus oil, eugenol, menthyl lactate, witch hazel extract), anti-acne agents, anti-caking agents, defoaming agents, antimicrobial agents (e.g., iodopropyl butylcarbamate), antioxidants, binders, biological additives, buffers, solubilizers, chelating agents, chemical additives, colorants, cosmetic astringents, cosmetic antiseptics, denaturants, pharmaceutical astringents, external analgesics, film formers or materials (e.g., polymers that contribute to the film-forming properties and substantiveness of the composition (e.g., copolymers of eicosene and vinyl pyrrolidone)), opacifiers, pH adjusters, propellants, reducing agents, chelating agents, skin bleaching and lightening agents (e.g., hydroquinone, kojic acid, ascorbic acid, magnesium ascorbyl phosphate, ascorbyl glucosamine), skin conditioning agents (e.g., humectants, including miscellaneous and occlusives), skin soothing and/or healing agents (e.g., panthenol and derivatives (e.g., ethyl panthenol), aloe vera, pantothenic acid and its derivatives, allantoin, bisabolol, and dipotassium glycyrrhizate), skin therapeutic agents, thickeners, and vitamins and their derivatives.

Farnesol

The topical compositions disclosed herein may include a safe and effective amount of farnesol. Farnesol is a naturally occurring substance that is believed to act as a precursor and/or intermediate in the biosynthesis of squalene and sterols (particularly cholesterol). Farnesol is also involved in the modification and regulation of proteins (e.g., protein farnesylation), and there are nuclear receptors that respond to farnesol.

Chemically, farnesol is [2E,6E]-3,7,11-trimethyl-2,6,10-dodecatrien-1-ol, and as used herein, "farnesol" includes isomers and tautomers of the above alcohol. Farnesol is commercially available, for example, under the names farnesol (a mixture of isomers formed from Dragoco, 10 Gordon Drive, Totowa, N.J.) and trans-trans-farnesol (Sigma Chemical Company, P.O. Box 14508, St. Louis, Mo.).

When farnesol is present in the compositions disclosed herein, the compositions preferably comprise from about 0.001% to about 50%, more preferably from about 0.01% to about 20%, even more preferably from about 0.1% to about 15%, even more preferably from about 0.1% to about 10%, even more preferably from about 0.5% to about 5%, and even more preferably from about 1% to about 5% farnesol by weight of the composition.

Phytantriol

The topical compositions disclosed herein may include a safe and effective amount of phytantriol. Phytantriol is the common name for a chemical called 3,7,11,15-tetramethylhexadecane-1,2,3-triol. Phytantriol is available from BASF (1609 Biddle Avenue, Whyandotte, Mich.). For example, phytantriol can be used as a spider vein/red spot repair agent, a dark circle/swollen eye blister repair agent, a sallow repair agent, a sagging repair agent, an antipruritic, a skin thickening agent, a pore shrinking agent, an oil/shine reducing agent, a post-inflammatory pigmentation repair agent, a wound treatment agent, a slimming agent, and to adjust skin texture (including wrinkles and fine lines).

In the compositions disclosed herein, the phytantriol contained preferably accounts for about 0.001% to about 50% by weight of the composition, more preferably about 0.01% to about 20%, even more preferably about 0.1% to about 15%, even more preferably about 0.2% to about 10%, still more preferably about 0.5% to about 10%, and even more preferably about 1% to about 5%.

Desquamation active substances

A safe and effective amount of a desquamation active substance can be added to the composition disclosed herein, more preferably from about 0.1% to about 10% by weight of the composition, even more preferably from about 0.2% to about 5%, and also preferably from about 0.5% to about 4%. Desquamation active substances enhance the skin appearance benefits disclosed herein. For example, desquamation active substances tend to improve the texture of the skin (e.g., smoothness). A desquamation system suitable for the purposes described herein comprises a sulfhydryl compound and a zwitterionic surfactant and is described in Bissett's U.S. Patent No. 5,681,852, which is incorporated herein by reference. Another desquamation system suitable for the purposes described herein comprises salicylic acid and a zwitterionic surfactant and is described in Bissett's U.S. Patent No. 5,652,228, which is incorporated herein by reference. Zwitterionic surfactants (such as those described in the present disclosure) are also used as desquamation agents as described herein, and hexadecyl betaine is particularly preferred.

Anti-acne active ingredients

The compositions disclosed herein may include a safe and effective amount of one or more anti-acne actives. Examples of useful anti-acne actives include resorcinol, sulfur, salicylic acid, benzoyl peroxide, erythromycin, zinc, and the like. Other examples of suitable anti-acne actives are described in further detail in U.S. Patent No. 5,607,980, issued to McAtee et al. on March 4, 1997.

Anti-wrinkle active substances/anti-aging active substances

The compositions disclosed herein may further comprise a safe and effective amount of one or more anti-wrinkle active substances or anti-aging active substances. Exemplary anti-wrinkle/anti-aging active substances suitable for use in the compositions disclosed herein include sulfur-containing D and L amino acids and their derivatives and salts, particularly N-acetyl derivatives, a preferred example of which is N-acetyl-L-cysteine; thiols, such as ethanethiol; hydroxy acids (e.g., α-hydroxy acids such as lactic acid and glycolic acid; or β-hydroxy acids such as salicylic acid and salicylic acid derivatives such as octanoyl derivatives), phytic acid, lipoic acid; lysophosphatidic acid, skin peeling agents (e.g., phenol, etc.), vitamin _B3 compounds, and retinoids, which enhance the appearance benefits of keratinous tissue disclosed herein, particularly regulating the condition of keratinous tissue, such as skin condition.

a) Vitamin _B3 compounds

The compositions disclosed herein may include a safe and effective amount of a vitamin _B3 compound. Vitamin _B3 compounds are particularly useful for regulating skin conditions, as described in co-pending U.S. application Ser. No. 08/834,010, filed April 11, 1997 (equivalent to International Publication No. WO 97/39733 A1, published October 30, 1997). When a vitamin _B3 compound is present in the compositions disclosed herein, the compositions preferably comprise from about 0.1% to about 10%, even more preferably from about 0.5% to about 10%, still more preferably from about 1% to about 5%, and still more preferably from about 2% to about 5%, by weight of the _composition .

As used herein, "vitamin _B3 compound" refers to a compound having the formula:

wherein R is -CONH ₂ (ie, nicotinamide), -COOH (ie, nicotinic acid), or -CH ₂ OH (ie, nicotinic alcohol); derivatives thereof; and salts of any of the foregoing.

Exemplary derivatives of the foregoing vitamin _B3 compounds include hydrochloride esters, non-vasodilating esters including niacin (eg, tocopheryl nicotinate), nicotinyl amino acids, nicotinyl alcohol esters of carboxylic acids, nicotinic acid N-oxide, and niacinamide N-oxide.

Examples of suitable vitamin _B3 compounds are well known in the art and are commercially available from a variety of sources, such as the Sigma Chemical Company (St. Louis, Mo.); ICN Biomedicals, Inc. (Irvin, Calif.); and Aldrich Chemical Company (Milwaukee, Wis.).

The vitamin compound may be included as the substantially pure material, or as an extract obtained by physical and/or chemical separation from natural (eg, plant) sources.

b) Retinoids

The compositions disclosed herein may also include a retinoid. As used herein, "retinoid" includes all natural and/or synthetic analogs of vitamin A or retinol-like compounds (which have the biological activity of vitamin A in the skin), as well as geometric isomers and stereoisomers of these compounds. The retinoid is preferably retinol, a retinol ester (e.g., a _C2 - _C22 alkyl ester of retinol, including retinyl palmitate, retinyl acetate, retinyl propionate), retinal and/or retinoic acid (including all trans-retinoic acid and/or 13-cis-retinoic acid), with the exception of retinoic acid, more preferably a retinoid. These compounds are well known in the art and can be purchased from a variety of sources, such as Sigma Chemical Company (St. Louis, Mo.), and Boerhinger Mannheim (Indianapolis, Ind.). Other retinoids for use herein are described in U.S. Patent No. 4,677,120, issued to Parish et al. on June 30, 1987, U.S. Patent No. 4,885,311, issued to Parish et al. on December 5, 1989, U.S. Patent No. 5,049,584, issued to Purcell et al. on September 17, 1991, U.S. Patent No. 5,124,356, issued to Purcell et al. on June 23, 1992, and U.S. Patent No. Reissue 34,075, issued to Purcell et al. on September 22, 1992. Other suitable retinoids are tocopheryl-retinoate [tocopheryl ester of retinoic acid (trans- or cis-), adapalene {6-[3-(1-adamantane)-4-methoxyphenyl]-2-naphthoic acid}, and tazarotene (ethyl 6-[2-(4,4-dimethylthiochroman-6-yl)-ethynyl]nicotinate). Preferred retinoids are retinol, retinyl palmitate, retinyl acetate, retinyl propionate, retinal, and combinations thereof.

The vitamin compound may be included as the substantially pure material, or in the form of an extract obtained by physical and/or chemical separation from natural (eg, plant) sources. The retinoid is preferably substantially pure, more preferably essentially pure.

The compositions disclosed herein may comprise a safe and effective amount of a retinoid such that the resulting composition is safe and effective for regulating keratinous tissue condition, preferably for regulating visible and/or tactile discontinuities in the skin, more preferably for regulating signs of skin aging, and even more preferably for regulating visible and/or tactile discontinuities in skin texture associated with skin aging. The composition preferably comprises from or about 0.005% to or about 2%, more preferably 0.01% to or about 2% retinoid. Retinol is preferably used in an amount of from or about 0.01% to or about 0.15%; retinol esters are preferably used in an amount of from or about 0.01% to or about 2% (e.g., about 1%); retinoic acid is preferably used in an amount of from or about 0.01% to or about 0.25%; tocopheryl-retinoate, adapalene, and tazarotene are preferably used in an amount of from or about 0.01% to or about 2%.

When the compositions disclosed herein contain a retinoid and a vitamin _B3 compound, the retinoid is preferably used in the amounts described above, and the vitamin _B3 compound is preferably used in an amount from at or about 0.1% to at or about 10%, more preferably from at or about 2% to at or about 5%.

c) Hydroxy acids

The compositions disclosed herein may include a safe and effective amount of a hydroxy acid. Preferred hydroxy acids for use in the compositions disclosed herein include salicylic acid and salicylic acid derivatives. When salicylic acid is present in the compositions disclosed herein, it is preferably used in an amount of from about 0.01% to about 50%, more preferably from about 0.1% to about 20%, even more preferably from about 0.1% to about 10%, still more preferably from about 0.5% to about 5%, and still more preferably from about 0.5% to about 2%.

peptides

Other peptides (including but not limited to di-, tri- and tetrapeptides and their derivatives) can be included in the compositions disclosed herein in safe and effective amounts. As used herein, "peptide" refers to naturally occurring peptides and synthetic peptides. Naturally occurring and commercially available compositions containing peptides can also be used in the present invention.

Suitable dipeptides for use in the present invention include carnosine (β-ala-his). Suitable tripeptides for use in the present invention include gly-his-lys, arg-lys-arg, and his-gly-gly. Preferred tripeptides and their derivatives include palmitoyl-gly-his-lys, which can be purchased from Biopeptide (100 ppm palmitoyl-gly-his-lys, available from Sederma, France); Peptide CK (arg-lys-arg); Peptide CK+ (ac-arg-lys-arg- _NH2 ); and the copper derivative of his-gly-gly available from Sigma (St. Louis, Mo.) as lamin. A tetrapeptide suitable for use in the present invention includes peptide E, arg-ser-arg-lys (SEQ ID NO: 11).

Preferably, the other peptide is selected from palmitoyl-gly-his-lys, β-ala-his, derivatives and combinations thereof. More preferably, the other peptide is selected from palmitoyl-gly-his-lys, derivatives and combinations thereof.

When other peptides are included in the compositions of the present invention, the composition preferably comprises from about 1 ^{x 10-6} % to about 10%, more preferably from about 1 ^{x 10-6} % to about 0.1%, and even more preferably from 1 ^{x 10-5} % to about 0.01% of the other peptide by weight of the composition. In certain embodiments wherein the peptide is included, the composition preferably comprises from about 0.1% to about 5% of the peptide by weight of the composition. In other embodiments wherein a peptide-containing composition comprising a biopeptide is included, the resulting composition preferably comprises from about 0.1% to about 10% of the biopeptide by weight of the composition.

Antioxidants/free radical scavengers

The compositions disclosed herein may include a safe and effective amount of an antioxidant/free radical scavenger. Antioxidants/free radical scavengers are particularly useful for providing protection against UV radiation (which can cause scaling or texture changes in the stratum corneum) and other environmental agents (which can cause skin damage).

A safe and effective amount of an antioxidant/radical scavenger may be added to the subject disclosed compositions, preferably from about 0.1% to about 10%, more preferably from about 1% to about 5% of the composition.

Antioxidants/free radical scavengers may be used, such as ascorbic acid (vitamin C) and its salts, ascorbic acid esters of fatty acids, ascorbic acid derivatives (e.g., magnesium ascorbyl phosphate, sodium ascorbyl phosphate, ascorbyl sorbate), tocopherol (vitamin E), tocopherol sorbate, tocopherol acetate, other esters of tocopherol, butylated hydroxybenzoic acid and its salts, 6-hydroxy-2,5,7,8-tetramethylchromanyl-2-carboxylic acid (sold under the trade name PEG-10), gallic acid and its alkyl esters, particularly gallic acid. [0014] Examples of the antioxidant/free radical scavenger include propyl betaine, uric acid and its salts and alkyl esters, sorbic acid and its salts, lipoic acid, amines (e.g., N,N-diethylhydroxylamine, aminoguanidine), sulfhydryl compounds (e.g., glutathione), dihydroxyfumaric acid and its salts, lycinepidolate, lycine arginine, nordihydroguaiaretic acid, bioflavonoids, turmeric, lysine, methionine, proline, superoxide dismutase, silymarin, tea extract, grape skin/seed extract, melanin, and citronellol extract. Preferred antioxidants/free radical scavengers are selected from tocopherol sorbate and other esters of tocopherol, more preferably tocopherol sorbate. For example, the use of tocopherol sorbate in topical compositions and uses suitable for the present disclosure is described in U.S. Patent No. 4,847,071, issued on July 11, 1989 to Donald L. Bissett, Rodney D. Bush, and Ranjit Chatterjee.

chelating agents

The compositions disclosed herein may also include a safe and effective amount of a chelating agent or chelating agent. As used herein, "chelating agent" or "chelating agent" refers to an active agent that is capable of removing metal ions from a system by forming a complex so that the metal ions cannot readily participate in or catalyze chemical reactions. The inclusion of a chelating agent is particularly useful for providing protection against UV radiation (which can contribute to excessive acne or texture changes) and other environmental agents (which can cause skin damage).

A safe and effective amount of a chelating agent can be added to the composition disclosed herein, preferably from about 0.1% to about 10% of the composition, more preferably from about 1% to about 5%. Exemplary chelating agents useful in the present invention are described in U.S. Patent No. 5,487,884, issued to Bissett et al. on January 30, 1996, International Publication No. 91/16035, issued to Bush et al. on October 31, 1995, and International Publication No. 91/16034, published on October 31, 1995, to Bush et al. Preferred chelating agents useful in the compositions disclosed herein are furildioxime, furilmonoxime, and their derivatives.

flavonoids

The compositions disclosed herein may optionally include flavonoid compounds.Flavonoids are extensively disclosed in US Patent Nos. 5,686,082 and 5,686,367, which are incorporated herein by reference. The flavonoids suitable for use in the present invention are flavanones selected from unsubstituted flavanones, monosubstituted flavanones and mixtures thereof; chalcone selected from unsubstituted chalcone, monosubstituted chalcone, disubstituted chalcone, trisubstituted chalcone and mixtures thereof; flavonoids selected from unsubstituted brass, monosubstituted brass, disubstituted brass and mixtures thereof; one or more isoflavones; coumarin selected from unsubstituted coumarins, monosubstituted coumarins, disubstituted coumarins and mixtures thereof; chromone selected from unsubstituted chromones, monosubstituted chromones, disubstituted chromones and mixtures thereof; one or more dicoumarols; one or more chroman-4-ones; one or more chromanols; isomers (e.g., cis/trans isomers) and mixtures thereof. As used herein, the term "substituted" refers to a flavonoid in which one or more hydrogen atoms are independently substituted by hydroxyl, C1-C8 alkyl, C1-C4 alkoxy, O-glycoside, etc., or a mixture of these substituents.

Examples of suitable flavonoids include, but are not limited to, unsubstituted flavanones, monohydroxyflavanones (e.g., 2'-hydroxyflavanone, 6-hydroxyflavanone, 7-hydroxyflavanone, etc.), monoalkoxyflavanones (e.g., 5-methoxyflavanone, 6-methoxyflavanone, 7-methoxyflavanone, 4'-methoxyflavanone, etc.), unsubstituted chalcone (particularly unsubstituted trans-chalcone), monohydroxychalcone (e.g., 2'-hydroxychalcone, 4'-hydroxychalcone, etc.), dihydroxychalcone (e.g., 2',4-dihydroxychalcone, 2',4'-dihydroxychalcone, 2,2'-dihydroxychalcone, 2',3-dihydroxychalcone, 2',5'-dihydroxychalcone, etc.), and trihydroxychalcone (e.g., 2',3',4'-trihydroxychalcone). chalcone, 4,2',4'-trihydroxychalcone, 2,2',4'-trihydroxychalcone, etc.), unsubstituted flavonoids, 7,2'-dihydroxyflavone, 3',4'-dihydroxynaphthoflavone, 4'-hydroxyflavone, 5,6-benzoflavone and 7,8-benzoflavone, unsubstituted isoflavones, daidzein (7,4'-dihydroxyisoflavone), 5,7-dihydroxy-4'-methoxyisoflavone, soy isoflavones (a mixture extracted from soybeans), unsubstituted coumarins, 4-hydroxycoumarin, 7-hydroxycoumarin, 6-hydroxy-4-methylcoumarin, unsubstituted chromones, 3-formylchromone, 3-formyl-6-isopropylchromone, unsubstituted dicoumarol, unsubstituted chroman-4-one, unsubstituted chromanols and mixtures thereof.

Unsubstituted flavanones, methoxyflavanones, unsubstituted chalcone, 2',4-dihydroxychalcone and mixtures thereof are preferably used in the present invention. Unsubstituted flavanones, unsubstituted chalcone (especially trans-isomers) and mixtures thereof are more preferred.

They can be synthetic materials or obtained as extracts from natural sources (e.g., plants). Naturally derived materials can also be further derivatized (e.g., ester or ether derivatives prepared from natural sources after extraction). Flavonoid compounds used in the present invention can be purchased from a variety of sources, such as Indofine Chemical Company, Inc. (Somerville, N.J.), Steraloids, Inc. (Wilton, N.H.), and Aldrich Chemical Company, Inc. (Milwaukee, Wis.).

Mixtures of the above-mentioned flavonoid compounds may also be used.

The flavonoids described herein are present in the compositions according to the present disclosure at a concentration of about 0.01% to about 20%, more preferably about 0.1% to about 10%, and even more preferably about 0.5% to about 5%.

anti-inflammatory agents

A safe and effective amount of an anti-inflammatory agent can be added to the compositions disclosed herein, preferably from about 0.1% to about 10%, more preferably from about 0.5% to about 5%, of the composition. The anti-inflammatory agent can enhance the skin appearance benefits disclosed herein, for example, such agents contribute to a more even and acceptable skin tone or color. The amount of the anti-inflammatory agent extract used in the composition will depend on the specific anti-inflammatory agent used, as the potency of such agents varies widely.

Steroidal anti-inflammatory agents that can be used include, but are not limited to, corticosteroids such as hydrocortisone, fludroxyl dehydrocortisone, alpha-methyldexamethasone, dexamethasone phosphate, beclomethasone dipropionate, clofloxacin valerate, hydroxyprednisolone acetonide, fludioxonolone diacetate, diflumethasone valerate, fluadrenolone, flucloroxolone acetonide, fludrocortisone, flumethasone trimethylacetate, fluosinolone acetonide, fluocinolone acetate, fluocinolone butyl ester, fluocortolone, fluprednisolone acetate, fluocortine acetonide, halcinonide, hydrocortisone acetate, hydrobutyrate The steroidal anti-inflammatory agent preferably used is hydroxycortisone.

A second class of anti-inflammatory agents that can be used in the compositions includes non-steroidal anti-inflammatory agents. The types of compounds encompassed by this group are well known to those skilled in the art. For detailed disclosure of the chemical structure, synthesis, side effects, etc. of non-steroidal anti-inflammatory agents, one can refer to standard references, including Anti-inflammatory and Anti-Rheumatic Drugs, K.D. Rainsford, Vol. I-III, CRC Press, Boca Raton, (1985), and Anti-inflammatory Agents, Chemistry and Pharmacology, 1, R.A. Scherrer, et al., Academic Press, New York (1974).

Specific nonsteroidal anti-inflammatory agents that can be used in the compositions disclosed herein include, but are not limited to:

1) Oxicams, such as piroxicam, isoxicam, thienoxicam, sudoxicam and CP-14,304;

2) Salicylates, such as aspirin, salsalate, pyraclostrobin, choline magnesium trisalicylate, safapryn, solprin, diflunisal, and fentosal;

3) acetic acid derivatives, for example, diclofenac, fenclorac, indomethacin, sulindac, toluylpyridine acetic acid, isoxacic acid, ethyldihydrobenzofuranacetic acid, dihydrooxydibenzothiazole, azidoindolic acid, acemetacin, dibenzothioic acid, clofenac, clindanac, oxepinic acid, felbinac and ketorolac;

4) Analgesics, such as mefenamic acid, meclofenamic acid, flufenamic acid, niflumic acid, and tolfenamic acid;

5) Propionic acid derivatives such as ibuprofen, naproxen, benoxaprofen, flurbiprofen, ketoprofen, fenoprofen, fenbufen, indopropfen, pyrrol, carprofen, lumefoxine, pranoprofen, miprofen, thioxaprofen, suprofen, alminoprofen and tiaprofen; and

6) Pyrazoles, for example phenylbutazone, oxyphenylbutazone, feprazone, azapropazone and trimethylenebutazone.

Mixtures of these nonsteroidal anti-inflammatory agents can also be used, as well as dermatologically acceptable salts or esters of these agents. For example, etofenamate (a derivative of flufenamic acid) is particularly useful in topical applications. Among the nonsteroidal anti-inflammatory agents, ibuprofen, naproxen, flufenamic acid, etofenamate, aspirin, mefenamic acid, meclofenamic acid, piroxicam, and felbinac are preferred; ibuprofen, naproxen, ketoprofen, etofenamate, aspirin, and flufenamic acid are more preferred.

Finally, so-called "natural" anti-inflammatory agents can be used in the methods disclosed herein. Such agents can be suitably obtained as extracts from natural sources (e.g., plants, fungi, by-products of microbial organisms) by appropriate physical and/or chemical isolation, or can be synthetically prepared. For example, candelilla wax, bisabolol (e.g., alpha bisabolol), aloe vera, phytosterols (e.g., plant steroids), Manjistha (extracted from plants of the genus Rubia, particularly Rubia Cordifolia), and Guggal (extracted from plants of the genus Commiphora, particularly Commiphora Mukul), kola nut extract, chamomile, red clover extract, and gorgonian extract can be used.

Other anti-inflammatory agents that can be used in the present invention include compounds of the Glycyrrhizae family (genus/species of the plant Glycyrrhiza glabra), including and their derivatives (e.g., salts and esters). Suitable salts of the above compounds include metal and ammonium salts. Suitable esters include _C2 - _C24 saturated or unsaturated esters of acids; preferably C10-C24, more preferably C16-C24. Specific examples of the above compounds include oil-soluble European licorice extract, glycyrrhizic acid and glycyrrhetinic acid itself, glycyrrhizic acid monoammonium salt, glycyrrhizic acid potassium salt, dipotassium glycyrrhizate, 1-β-hydrogen succinate glycyrrhetinic acid, glycyrrhetinic acid stearyl ester and 3-stearoyloxy-glycyrrhetinic acid, and 3-succinyloxy-β-glycyrrhetinic acid disodium. Glycyrrhetinic acid stearyl ester is preferred.

slimming supplements

The composition disclosed herein may also include a safe and effective amount of a slimming agent. Suitable agents may include, but are not limited to, xanthine compounds (eg, caffeine, theophylline, theobromine, and aminophylline).

local anesthetics

The composition disclosed herein may further comprise a safe and effective amount of a local anesthetic. Examples of local anesthetic drugs include benzocaine, lidocaine, bupivacaine, chloroprocaine, cinchocaine, etidocaine, mepivacaine, tetracaine, dyclonine, hexylcaine, procaine, cocaine, ketamine, pramoxine, phenol, and pharmaceutically acceptable salts thereof.

Tanning active substances

The compositions disclosed herein may contain a tanning active. When present, it is preferred that the composition contain from about 0.1% to about 20%, more preferably from about 2% to about 7%, and even more preferably from about 3% to about 6%, of dihydroxyacetone as an artificial tanning active, based on the weight of the composition.

Tanning active substance (also known as DHA or 1,3-dihydroxy-2-propanone) is a white to off-white crystalline powder. This material can be represented by the chemical formula C ₃ H ₆ O ₃ and the chemical structure below.

The compound may exist as a mixture of monomers and dimers, with the dimer predominating in the solid crystalline state. Upon heating or melting, the dimer breaks apart to produce the monomer. Conversion of the dimer to the monomeric form may also occur in aqueous solution. Dihydroxyacetone is also known to be more stable at acidic pH values. See The Merck Index, Tenth Edition, entry 3167, p. 463 (1983) and "Dihydroxyacetone for Cosmetics", E. Merck Technical Bulletin, 03-304 1 10, 319 897, 180 588.

Skin whitening agents

The compositions disclosed herein may include a skin-lightening agent. When used, the composition preferably comprises from about 0.1% to about 10%, more preferably from about 0.2% to about 5%, and even more preferably from about 0.5% to about 2%, of the skin-lightening agent, by weight of the composition. Suitable skin-lightening agents include those known in the art, including kojic acid, arbutin, ascorbic acid and its derivatives (e.g., magnesium ascorbyl phosphate or sodium ascorbyl phosphate), and extracts (e.g., mulberry extract, placental extract). Skin lightening agents suitable for use in the present invention also include those described in PCT Publication No. 95/34280 to Hillebrand (equivalent to PCT Application No. U.S. Ser. No. 95/07432 filed June 12, 1995), and in co-pending U.S. Application Serial No. 08/390,152 to Kvalnes, Mitchell A. DeLong, Barton J. Bradbury, Curtis B. Motley and John D. Carter (equivalent to PCT Publication Serial No. 95/23780 published September 8, 1995).

Skin soothing and skin healing active ingredients

The compositions disclosed herein may include a skin-soothing or skin-healing active. Suitable skin-soothing or skin-healing actives for use herein include pantothenic acid derivatives (including panthenol, panthenol, and ethyl panthenol), aloe vera, allantoin, bisabolol, and dipotassium glycyrrhizate. A safe and effective amount of a skin-soothing or skin-healing active may be added to the compositions of the present invention, preferably from about 0.1% to about 30%, more preferably from about 0.5% to about 20%, and even more preferably from about 0.5% to about 10%, by weight of the resulting composition.

a) Bisabolol

The topical compositions disclosed herein may also include a safe and effective amount of bisabolol. Bisabolol is a naturally occurring unsaturated monocyclic terpene alcohol having the following structure.

Bisabolol is the major active ingredient in chamomile extract/oil. Bisabolol can be synthetic (d,1-α-isomer or (+/-)-α-isomer) or of natural origin ((-)-α-isomer) and can be used as an essentially pure compound or a mixture of compounds (e.g., extracts obtained from natural sources, such as chamomile). The α form of bisabolol (α-bisabolol) is used in a variety of cosmetic products as a skin conditioning or soothing agent. As used herein, "bisabolol" includes chamomile extract or oil, and any isomers and tautomers of such substances. Suitable bisabolol compounds are available from Dragoco (Totowa, N.J.) as a natural material under the product name α-Bisabolol (Natural) and from Fluka (Milwaukee, Wis.) as a synthetic material under the product name α-Bisabolol.

In the compositions disclosed herein, the composition preferably comprises from about 0.001% to about 50%, more preferably from about 0.01% to about 20%, even more preferably from about 0.01% to about 15%, still more preferably from about 0.1% to about 10% bisabolol by weight of the composition.

Antimicrobial and antifungal active substances

The compositions disclosed herein may include antimicrobial or antifungal actives. Such actives can destroy microorganisms, prevent their development, or prevent their pathogenic effects. A safe and effective amount of an antimicrobial or antifungal active can be added to the compositions of the present invention, preferably from about 0.001% to about 10%, more preferably from about 0.01% to about 5%, and even more preferably from about 0.05% to about 2%.

Examples of antimicrobial and antifungal active substances include β-lactam drugs, quinolone drugs, ciprofloxacin, norfloxacin, tetracycline, erythromycin, amikacin, 2,4,4'-trichloro-2'-hydroxydiphenyl ether, 3,4,4'-trichloroaniline, phenoxyethanol, phenoxypropanol, phenoxyisopropanol, doxycycline, capreomycin, chlorhexidine, chlortetracycline, oxytetracycline, lintamycin, ethambutol, hexamidine isethionate, metronidazole, pentamicin, gentamicin, kanamycin, tetracycline, methyl Oxytetracycline, methenamine, minocycline, neomycin, netilmicin, paromomycin, streptomycin, tobramycin, myconazole, tetracycline hydrochloride, erythromycin, erythromycin zinc, erythromycin estolate, erythromycin stearate, amikacin sulfate, doxycycline hydrochloride, capreomycin sulfate, chlorhexidine gluconate, chlorhexidine hydrochloride, chlortetracycline hydrochloride, tetracycline hydrochloride, lindamycin hydrochloride, ethambutol hydrochloride, metronidazole hydrochloride, pentamidine hydrochloride, gentamicin sulfate, kanamycin sulfate, lineomycin hydrochloride, methacycline hydrochloride, methenamine hippurate, mandraxamine, minocycline hydrochloride, neomycin sulfate, netilmicin sulfate, paromomycin sulfate, streptomycin sulfate, tobramycin sulfate, myconazole hydrochloride, ketoconazole, amantadine hydrochloride, amantadine sulfate, octopirox, parachlorometa-xylenol, nystatin, tolnaftate, pyrithione zinc, and clotrimazole.

Preferred examples of active substances that can be used in the present invention include those selected from the group consisting of salicylic acid, benzoyl peroxide, 3-hydroxybenzoic acid, glycolic acid, lactic acid, 4-hydroxybenzoic acid, acetylsalicylic acid, 2-hydroxybutyric acid, 2-hydroxyvaleric acid, 2-hydroxyhexanoic acid, cis-retinoic acid, trans-retinoic acid, retinol, phytic acid, N-acetyl-L-cysteine, lipoic acid, azelaic acid, arachidonic acid, benzoyl peroxide, tetracycline, ibuprofen, naproxen, hydrocortisone, acetaminophen, resorcinol, phenoxyethanol, phenoxypropanol, phenoxyisopropanol, 2,4,4'-trichloro-2'-hydroxydiphenyl ether, 3,4,4-trichlorourea, octopirox, lidocaine hydrochloride, clotrimazole, miconazole, ketoconazole, neomycin sulfate and mixtures thereof.

Sunscreen active ingredients

Exposure to ultraviolet light can lead to excessive acne and changes in the texture of the stratum corneum. Therefore, the compositions disclosed herein may optionally include a sunscreen active. As used herein, "sunscreen active" includes sunscreen agents and physical sunscreens. Suitable sunscreen actives can be organic or inorganic.

Inorganic sunscreens that can be used in the present invention include the following metal oxides: titanium dioxide having an average primary particle size of about 15 nm to about 100 nm; zinc oxide having an average primary particle size of about 15 nm to about 150 nm; zirconium oxide having an average primary particle size of about 15 nm to about 150 nm; iron oxide having an average primary particle size of about 15 nm to about 500 nm; and mixtures thereof. When inorganic sunscreens are used in the present invention, the inorganic sunscreens comprise from about 0.1% to about 20%, preferably from about 0.5% to about 10%, and more preferably from about 1% to about 5%, by weight of the composition.

A variety of conventional organic sunscreen actives are suitable for use in the present invention. Sagarin, et al., at Chapter VIII, pages 189 et seq., of Cosmetics Science and Technology (1972) discloses a variety of suitable actives. Specific suitable sunscreen actives include, for example, p-aminobenzoic acid, its salts and derivatives (ethyl, isobutyl, glyceryl esters; p-dimethylaminobenzoic acid); aminobenzoates (i.e., anthranilates; methyl, menthyl, phenyl, benzyl, phenylethyl, linalool, pinoleyl, and cyclohexenyl esters); salicylates (amyl, phenyl, octyl, benzyl, menthyl, glyceryl, and dipropylene glycol esters); cinnamic acid derivatives (menthyl and benzyl esters, α-phenylcinnamonitrile; butyrylcinnamonitrile); keto acids); dihydroxycinnamic acid derivatives (umbelliferone, methylumbelliferone, methylacetyl-umbelliferone); trihydroxycinnamic acid derivatives (esculetin, methylesculetin, daphnetin, glycosides, esculetin and daphnetin); hydrocarbons (diphenylbutadiene, stilbene); dibenzylideneacetone and benzylideneacetophenone; naphthol sulfonates (sodium salts of 2-naphthol-3,6-disulfonic acid and 2-naphthol-6,8-disulfonic acid); dihydroxynaphthoic acid and its salts; o- and p-diphenyldisulfonic acid esters; coumarin derivatives (7-hydroxy, 7-methyl, 3-phenyl); diazoles (2-acetyl-3-bromoindazole, phenylbenzoxazole, methylnaphthoxazole, various arylbenzothiazoles); quinine salts (bisulfate, sulfate, chloride, oleate and tannate); quinoline derivatives (8-hydroxyquinolinate, 2-phenylquinoline); hydroxy- or methoxy-substituted benzophenones; uric acid and violuric acid; tannic acid and its derivatives (e.g., hexaethylether); (butylcarbitol) (6-propyl piperonyl) ether; hydroquinone; benzophenones (hydroxybenzene, sulfisobenzone, dihydroxybenzone, benzoylresorcinol, 2,2',4,4'-tetrahydroxybenzophenone, 2,2'-dihydroxy-4,4'-dimethoxybenzophenone, octabenzophenone; 4-isopropyldibenzoylmethane; butylmethoxydibenzoylmethane; etocrylene; octocrylene; [3-(4'-methylbenzylideneterpan-2-one), terephthalylidene camphor sulfonic acid and 4-isopropyl-dibenzoylmethane.

Among them, 2-ethylhexyl-p-methoxycinnamic acid (available from PARSOL MCX), 4,4'-tert-butylmethoxydibenzoylmethane (available from PARSOL 1789), 2-hydroxy-4-methoxybenzophenone, octyldimethyl-p-aminobenzoic acid, galloylgallic acid trioleate, 2,2-dihydroxy-4-methoxybenzophenone, ethyl-4-(bis(hydroxy-propyl))aminobenzoate, 2-ethylhexyl-2-cyano-3,3-diphenylacrylate, 2-ethylhexyl-salicylate, glyceryl-p-aminobenzoate, 3,3,5-trimethylcyclohexyl salicylate, methylaminobenzoate, p-dimethyl-aminobenzoic acid or aminobenzoate, 2-ethylhexyl-p-dimethyl-amino-benzoate, 2-phenylbenzimidazole-5-sulfonic acid, 2-(p-dimethylaminophenyl)-5-sulfobenzoxazole acid, octocrylene and mixtures of these compounds are preferred.

More preferred organic sunscreen actives for use in the compositions include 2-ethylhexyl-p-methoxycinnamic acid, butyl methoxydibenzoylmethane, 2-hydroxy-4-methoxybenzophenone, 2-phenylbenzimidazole-5-sulfonic acid, octyldimethyl-p-aminobenzoic acid, octocrylene and mixtures thereof.

Sunscreen actives such as those disclosed in U.S. Pat. No. 4,937,370, issued to Sabatelli on June 26, 1990, and U.S. Pat. No. 4,999,186, issued to Sabatelli & Spirnak on March 12, 1991, are also particularly useful in the compositions. The sunscreen agents disclosed herein have two different chromophore moieties in a single molecule that exhibit different ultraviolet radiation absorption spectra. One chromophore moiety absorbs primarily in the UVB radiation range, while the other chromophore moiety absorbs strongly in the UVA radiation range.

Preferred members of this class of sunscreen agents are 4-N,N-(2-ethylhexyl)methyl-aminobenzoate of 2,4-dihydroxybenzophenone; ester of N,N-di-(2-ethylhexyl)-4-aminobenzoic acid with 4-hydroxydibenzoylmethane; ester of 4-N,N-(2-ethylhexyl)methyl-aminobenzoic acid with 4-hydroxydibenzoylmethane; 4-N,N-(2-ethylhexyl)methyl-aminobenzoic acid with 4-hydroxydibenzoylmethane; 4-N,N-(2-hydroxyethoxy)benzophenone -ethylhexyl) methyl-aminobenzoate; 4-(2-hydroxyethoxy)dibenzoylmethane 4-N,N-(2-ethylhexyl)-methylaminobenzoate; 2-hydroxy-4-(2-hydroxyethoxy)benzophenone N,N-di-(2-ethylhexyl)-4-aminobenzoate; 4-(2-hydroxyethoxy)dibenzoylmethane N,N-di-(2-ethylhexyl)-4-aminobenzoate, and mixtures thereof.

Particularly preferred sunscreen actives include 4,4'-tert-butylmethoxydibenzoylmethane, 2-ethylhexyl-p-methoxycinnamic acid, phenylbenzimidazole sulfonic acid and octocrylene.

A safe and effective amount of organic sunscreen active material is used, typically from about 1% to about 20%, more typically from about 2% to about 10%, by weight of the composition. The amount of extract will vary depending on the sunscreen material or materials selected and the desired UV protection factor (SPF).

Granular materials

The compositions disclosed herein may include particulate materials, preferably metal oxides. These particles may be coated or uncoated, charged or uncharged. Charged particulate materials are disclosed in U.S. Patent No. 5,997,887 to Ha et al., which is incorporated herein by reference. Particulate materials that may be used in the present invention include bismuth oxychloride, iron oxide, mica, mica treated with barium sulfate and _TiO2 , silica, nylon, polyethylene, talc, styrene, polypropylene, ethylene/acrylic acid copolymers, sericite, alumina, silicone resins, barium sulfate, calcium carbonate, cellulose acetate, titanium dioxide, polymethyl methacrylate, and mixtures thereof.

Inorganic particulate materials (e.g., TiO ₂ , ZnO, or ZrO ₂ ) are commercially available from a variety of sources. One example of a suitable particulate material includes material available from American Cosmetics (TRONOX TiO ₂ series, SAT-T CR837, rutile TiO ₂ ). Preferably, the particulate material is present in the composition at a level of about 0.01% to about 2%, more preferably about 0.05% to about 1.5%, and even more preferably about 0.1% to about 1%, by weight of the composition.

Conditioning reagents

The compositions disclosed herein may include a conditioning agent selected from a humectant, moisturizer, or skin conditioner. A variety of such materials may be used, and each material may be present at a level of from about 0.01% to about 20%, more preferably from about 0.1% to about 10%, and even more preferably from about 0.5% to about 7%, by weight of the composition. These materials include, but are not limited to, guanidine; urea; glycolic acid and glycolate salts (e.g., ammonium and alkyl quaternary ammonium salts); salicylic acid; lactic acid and lactate salts (e.g., ammonium and alkyl quaternary ammonium salts); aloe vera in any of its various forms (e.g., aloe vera gel); polyols such as sorbitol, mannitol, xylitol, erythritol, glycerol, hexanetriol, butanetriol, propylene glycol, butylene glycol, ethylene glycol, and the like; polyethylene glycols; sugars (e.g., melibiose) and starches; sugar and starch derivatives (e.g., alkoxylated glucose, fructose, glucosamine); hyaluronic acid; lactamide monoethanolamine; acetamide monoethanolamine; panthenol; allantoin; and mixtures thereof. Also useful in the present invention are the propoxylated glycerols described in US Pat. No. 4,976,953, issued Dec. 11, 1990 to Orr et al.

Various _C1 - _C30 monoesters and polyesters of sugars and related materials can also be used. These esters are derived from sugars or polyols and one or more carboxylic acid moieties. Such ester materials are described in U.S. Patent No. 2,831,854, issued to Jandacek on January 25, 1977; U.S. Patent No. 4,005,196, issued to Jandacek on January 25, 1977; U.S. Patent No. 4,005,195, issued to Jandacek on January 25, 1977; U.S. Patent No. 5,306,516, issued to Letton et al. on April 26, 1994; U.S. Patent No. 5,306,515, issued to Letton et al. on April 26, 1994; and U.S. Patent No. 5,306,516, issued to Letton et al. on April 26, 1994. No. 5,305,514 to Tton et al.; U.S. Patent No. 4,797,300 issued to Jandacek et al. on January 10, 1989; U.S. Patent No. 3,963,699 issued to Rizzi et al. on January 15, 1976; U.S. Patent No. 4,518,772 issued to Volpenhein on May 21, 1985; and further described in U.S. Patent No. 4,517,360 issued to Volpenhein on May 21, 1985.

Preferably, the conditioning agent is selected from the group consisting of urea, guanidine, sucrose polyester, panthenol, panthenol, allantoin, and combinations thereof.

Structural reagents

The compositions described herein, and particularly the emulsions described herein, may contain structuring agents. Structuring agents are particularly preferred in the oil-in-water emulsions disclosed herein. Without being limited by theory, it is believed that structuring agents help provide rheological characteristics to the compositions that contribute to the stability of the compositions. For example, structuring agents often contribute to the formation of a liquid crystal gel network structure. The structuring agents may also function as emulsifiers or surfactants. Preferred compositions contain from about 0.1% to about 20%, more preferably from about 0.1% to about 10%, and even more preferably from about 0.5% to about 9% of one or more structuring agents.

Preferred structuring agents are those having an HLB of about 1 to about 8 and a melting point of at least about 45°C. Suitable structuring agents are selected from saturated _C14 to _C30 fatty alcohols, saturated C16 to C30 fatty alcohols containing from about 1 to about 5 moles of polyethylene oxide, saturated _C16 to _C30 diols _, saturated _C16 to _C30 monoglycerol ethers, saturated _C16 to _C30 hydroxy fatty acids, _C14 to _C30 hydroxylated and non-hydroxylated saturated fatty acids, _C14 to _C30 saturated ethoxylated fatty acids, amines and alcohols containing _from about 1 to about 5 moles of ethylene oxide diol, _C14 to _C30 saturated glyceryl monoesters having a monoglyceride content of at least 40%, _C1 to _C30 saturated polyglycerol esters having from about 1 to about 3 alkyl groups and from about 2 to about 3 saturated glycerol units, _C1 to _C30 glyceryl monoesters, _C1 to _C30 sorbitan mono/diesters, _C1 to C30 saturated C1 to _C30 saturated ethoxylated sorbitan mono/diester, _C1 to _C30 saturated methyl glycoside esters, _C1 to _C30 saturated sucrose mono/diester, _C14 to _C30 saturated ethoxylated methyl glycoside esters having from about 1 to about 5 moles of ethylene oxide, C1 to _C30 saturated polyglycosides having from ₁ to 2 glucose units, and mixtures thereof, having a melting point of at least about 45°C.

Preferred structuring agents disclosed herein are selected from the group consisting of stearic acid, palmitic acid, stearyl alcohol, cetyl alcohol, behenyl alcohol, stearic acid, palmitic acid, polyethylene glycol ethers of stearyl alcohol having an average of about 1 to about 5 ethylene oxide units, polyethylene glycol ethers of cetyl alcohol having an average of about 1 to about 5 ethylene oxide units, and mixtures thereof. More preferred structuring agents disclosed herein are selected from the group consisting of stearyl alcohol, cetyl alcohol, behenyl alcohol, polyethylene glycol ethers of stearyl alcohol having an average of about 2 ethylene oxide units (steareth-2), polyethylene glycol ethers of cetyl alcohol having an average of about 2 ethylene oxide units, and mixtures thereof. Even more preferred structuring agents are selected from the group consisting of stearic acid, palmitic acid, stearyl alcohol, cetyl alcohol, behenyl alcohol, steareth-2, and mixtures thereof.

Thickening agents (including thickeners and gelling agents)

The compositions disclosed herein may contain one or more thickening agents, preferably comprising from about 0.1% to about 5%, more preferably from about 0.1% to about 4%, still more preferably from about 0.25% to about 3% by weight of the composition.

Non-limiting classes of thickening agents include those selected from the group consisting of:

a) Carboxylic acid polymers

These polymers are crosslinked compounds comprising one or more monomers derived from acrylic acid, substituted acrylic acids, and salts and esters of these acrylic acids and substituted acrylic acids, wherein the crosslinking agent comprises two or more carbon-carbon double bonds and is derived from a polyol. Polymers useful in the present invention are described more fully in U.S. Patent No. 5,087,445, issued to Haffey et al. on February 11, 1992; in U.S. Patent No. 4,509,949, issued to Huang et al. on April 5, 1985; in U.S. Patent No. 2,798,053, issued to Brown on July 2, 1957; and in CTFA International Cosmetic Ingredient Dictionary, Fourth Edition, 1991, pp. 12 and 80.

Examples of commercially available carboxylic acid polymers that can be used in the present invention include carbomers, which are homopolymers of acrylic acid crosslinked with allyl ethers of sucrose or pentaerythritol. Carbomers are the 900 series available from BFGoodrich (e.g., 954). In addition, other suitable carboxylic acid polymerization reagents include copolymers of _C10-30 alkyl acrylates with one or more monomers of acrylic acid, methacrylic acid, or a short chain (i.e., _C1-4 alcohol) ester thereof, wherein the crosslinking agent is an allyl ether of sucrose or pentaerythritol. These copolymers are known as acrylates/ _C10 - _C30 alkyl acrylate crosspolymers and are available from Pemulen TR-1 and Pemulen TR-2, 1342 and 1382, available from BFGoodrich. In other words, examples of carboxylic acid polymer thickeners that can be used in the present invention are those selected from carbomers, acrylates/ _C10 - _C30 alkyl acrylate crosspolymers, and mixtures thereof.

b) Cross-linked polyacrylate polymers

The compositions disclosed herein may optionally contain cross-linked polyacrylate polymers, including cationic and nonionic polymers, as thickeners or gelling agents, with cationic polymers generally being preferred. Examples of cross-linked nonionic polyacrylate polymers and cross-linked cationic polyacrylate polymers that can be used are described in U.S. Pat. No. 5,100,660, issued to Hawe et al. on March 31, 1992; U.S. Pat. No. 4,849,484, issued to Heard on July 18, 1989; U.S. Pat. No. 4,835,206, issued to Farrar et al. on May 30, 1989; U.S. Pat. No. 4,628,078, issued to Glover et al. on December 9, 1986; U.S. Pat. No. 4,599,379, issued to Flesher et al. on July 8, 1986; and EP 228,868, published by Farrar et al. on July 15, 1987.

c) Polyacrylamide polymer

The compositions disclosed herein may optionally contain polyacrylamide polymers, particularly nonionic polyacrylamide polymers, including substituted branched or unbranched polymers. Of these polyacrylamide polymers, the nonionic polymers given the CTFA designations polyacrylamide, isoparaffin, and laureth-7 (available from Seppic Corporation (Fairfield, N.J.) under the trade name Sepigel 305) are more preferred.

Other polyacrylamide polymers useful in the present invention include multi-block copolymers of acrylamide and substituted acrylamides with acrylic acid and substituted acrylic acid. Commercially available examples of these multi-block copolymers include Hypan SR150H, SS500V, SS500W, and SSSA100H from Lipo Chemicals, Inc., (Patterson, N.J.).

d) Polysaccharides

A variety of polysaccharides can be used in the present invention. "Polysaccharide" refers to a gelling agent comprising a backbone of repeating sugar (i.e., carbohydrate) units. Non-limiting examples of polysaccharide gelling agents include those selected from the group consisting of cellulose, carboxymethyl hydroxyethyl cellulose, cellulose acetate propionate carboxylate, hydroxyethyl cellulose, hydroxyethyl ethyl cellulose, hydroxypropyl cellulose, hydroxypropyl methyl cellulose, methyl hydroxyethyl cellulose, microcrystalline cellulose, sodium cellulose sulfate, and mixtures thereof. Alkyl-substituted celluloses can also be used in the present invention. In these polymers, the hydroxyl groups of the cellulose polymer are hydroxyalkylated (preferably hydroxyethylated or hydroxypropylated) to form hydroxyalkylated cellulose, which is then further modified with _{a C10} - _C30 straight or branched chain alkyl group via an ether linkage. Typically, these polymers are ethers formed between a C10- _C30 straight or branched chain alcohol and a hydroxyalkyl cellulose. Examples of alkyl groups useful in the present invention include those selected from the group consisting of stearoyl, isostearyl, dodecyl, tetradecyl, cetyl, isohexadecyl, cocoyl (i.e., an alkyl group derived from an alcohol of coconut oil), cetyl, oleyl, linoleyl, linolenyl, ricinyl, behenyl, and mixtures thereof. Among the alkyl hydroxyalkyl cellulose ethers, the material given the CTFA designation cetyl hydroxyethylcellulose is preferred, wherein the material is an ether of cetyl alcohol and hydroxyethylcellulose. This material is sold by Aqualon Corporation (Wilmington, Del.) under the trade name CS Plus.

Other useful polysaccharides include scleroglucans, which are linear chains of (1-3) linked glucose units where every third unit has a (1-6) linked glucose, a commercially available example of which is Clearogel ^™ CS11 from Michel Mercier Products Inc. (Mountainside, NJ).

e) Gum

Other thickening and gelling agents useful in the present invention include materials derived primarily from natural sources. Non-limiting examples of these gelling agent gums include gum arabic, agar, algin, alginic acid, ammonium alginate, pullulan, calcium alginate, calcium carrageenan, carnitine, carrageenan, dextrin, gelatin, gellan gum, guar gum, guar hydroxypropyl ammonium chloride, hectorite, hyaluronic acid, hydrated silica, hydroxypropyl chitosan, hydroxypropyl guar gum, karaya gum, kelp, locust bean gum, natto, potassium alginate, potassium carrageenan, propylene glycol alginate, sclerotium gum, sodium carboxymethyl dextran, sodium carrageenan, tragacanth gum, xanthan gum, and mixtures thereof.

Preferred compositions disclosed herein include a thickening agent selected from the group consisting of carboxylic acid polymers, cross-linked polyacrylate polymers, polyacrylamide polymers, and mixtures thereof, more preferably selected from the group consisting of carboxylic acid polymers, polyacrylamide polymers, and mixtures thereof.

Dermatologically acceptable carrier

The topical compositions disclosed herein also include a dermatologically acceptable carrier. As used herein, the phrase "dermatologically acceptable carrier" means that the carrier is suitable for application to keratinous tissue, has good cosmetic properties, is compatible with the active ingredients disclosed herein and any other ingredients, and does not cause any undesirable safety or toxicity issues. A safe and effective amount of the carrier is from about 50% to about 99.99%, preferably from about 80% to about 99.9%, more preferably from about 90% to about 98%, and even more preferably from about 90% to about 95% of the composition.

The carrier can be in various forms. For example, an emulsion carrier can be used in the present invention, including but not limited to oil-in-water, water-in-oil, water-in-oil-in-water, and oil-in-water-in-water emulsions of silicone oil.

Preferred carriers include emulsions, such as oil-in-water emulsions, water-in-oil emulsions, and water-in-silicone emulsions. As will be appreciated by those skilled in the art, a given ingredient will be primarily distributed in the water or oil/silicone phase, depending on the solubility/dispersibility of the ingredient in the composition. Oil-in-water emulsions are particularly preferred.

Emulsions disclosed in accordance with the present invention typically comprise the solution described above and a lipid or oil. Lipids and oils can be derived from animals, plants, or petroleum, and can be natural or synthetic (i.e., artificial). Preferred emulsions also contain a wetting agent, such as glycerol. The emulsion preferably further comprises an emulsifier in an amount of about 0.01% to about 10%, more preferably about 0.1% to about 5%, based on the weight of the carrier. The emulsifier can be nonionic, anionic, or cationic. Suitable emulsifiers are disclosed in, for example, U.S. Patent No. 3,755,560, issued to Dickert et al. on August 28, 1973; U.S. Patent No. 4,421,769, issued to Dixon et al. on December 20, 1983; and McCutcheon's Detergents and Emulsifiers, North American Edition, pages 317-324 (1986).

The emulsion may also contain an anti-foaming agent to reduce foaming when applied to keratinous tissue. Anti-foaming agents include high molecular weight silicone resins and other materials known in the art for such use.

Suitable emulsions can have a wide range of viscosities, depending on the desired product form. Preferred exemplary low viscosity emulsions have a viscosity of about 50 centistokes or less, more preferably about 10 centistokes or less, and even more preferably about 5 centistokes or less.

Preferred water-in-silicone and oil-in-water emulsions are described in more detail below.

A) Water-in-silicone emulsion

Water-in-silicone emulsions contain a continuous silicone phase and a dispersed aqueous phase.

(1) Continuous silicone resin phase

The preselected water-in-silicone emulsions disclosed herein comprise from about 1% to about 60%, preferably from about 5% to about 40%, and more preferably from about 10% to about 20%, by weight of a continuous silicone phase. The continuous silicone phase is present in an external phase that contains or surrounds a discontinuous aqueous phase described below.

The continuous silicone phase comprises a polyorganosiloxane oil. Preferred oil-in-silicone emulsion systems are formulated to provide an oxidatively stable vehicle for retinoids. The continuous silicone phase of these preferred emulsions comprises from about 50% to about 99.9% by weight of organopolysiloxane oil and less than about 50% by weight of non-silicone oil. In particularly preferred embodiments, the continuous silicone phase comprises at least about 50%, preferably from about 60% to about 99.9%, more preferably from about 70% to about 99.9%, even more preferably from about 80% to about 99.9% polyorganosiloxane oil, by weight of the continuous silicone phase, and up to about 50% non-silicone oil, preferably less than about 40%, more preferably less than about 30%, even more preferably less than about 10%, and even more preferably less than about 2% of the continuous silicone phase. These preferred emulsion systems provide greater oxidative stability to the retinoids over extended periods of time compared to comparable water-in-oil emulsions containing lower concentrations of polyorganosiloxane oils. The concentration of non-silicone oils in the continuous silicone phase is minimized or avoided, further enhancing the oxidative stability of the selected retinoids in the compositions. Water-in-silicone emulsions of this type are described in PCT Application WO 97/21423, published June 19, 1997.

The organopolysiloxane oil used in the composition can be a volatile, non-volatile, or a mixture of volatile and non-volatile silicone resins. As used herein, the term "non-volatile" refers to those silicone resins that are liquid under ambient conditions and have a flash point equal to or greater than about 100°C (at one atmosphere of pressure). As used herein, the term "volatile" refers to all other silicone oils. Suitable organopolysiloxanes can be selected from a variety of silicone resins spanning a wide range of volatilities and viscosities. Examples of suitable organopolysiloxane oils include polyalkylsiloxanes, cyclic polyalkylsiloxanes, and polyalkylarylpolysiloxanes.

Polyalkylsiloxanes that can be used in the compositions of the present invention include those having a viscosity of about 0.5 to about 1,000,000 centistokes at 25°C. Such polyalkylsiloxanes can be represented by the general chemical formula _R3SiO [ _R2SiO ] _xSiR3 , where R is an alkyl group having from 1 to about 30 carbon atoms (preferably R is methyl or ethyl, more preferably methyl; _in addition, mixed alkyl groups can be used in the same molecule), and x is an integer from 0 to about 10,000, selected to achieve the desired molecular weight, which can span about 10,000,000. Commercially available polyalkylsiloxanes include polydimethylsiloxanes, also known as dimethicone, examples of which include the Siloxane series sold by General Electric Company and the Dow Corning 200 series sold by Dow Corning Corporation. Specific examples of suitable polydimethylsiloxanes include Dow Corning® 200 fluid having a viscosity of 0.65 centistokes and a boiling point of 100°C, Dow Corning® 225 fluid having a viscosity of 10 centistokes and a boiling point above 200°C, and Dow Corning® 200 fluid having viscosities of 50, 350, and 12,500 centistokes, respectively, and a boiling point above 200°C. Suitable dimethicone oils include those represented by the formula (CH ₃ ) ₃ SiO[(CH ₃ ) ₂ SiO] _x [CH ₃ RSiO] _y Si(CH ₃ ) ₃ , wherein R is a linear or branched alkyl group having from 2 to about 30 carbon atoms, and x and y are both integers of 1 or greater, selected to give the desired molecular weight, which may range from about 10,000,000. Examples of these alkyl-substituted dimethicone oils include hexadecyl dimethicone and lauryl dimethicone.

Suitable cyclic polyalkylsiloxanes for use in the compositions include those represented by the formula [ _SiR2 -O] _n , wherein R is an alkyl group (preferably R is methyl or ethyl, more preferably methyl), and n is an integer from about 3 to about 8, more preferably n is an integer from about 3 to about 7, and even more preferably n is an integer from about 4 to about 6. When R is methyl, these materials are generally referred to as cyclomethicones. Commercially available cyclomethicones include Dow Corning® 244 fluid, having a viscosity of 2.5 centistokes and a boiling point of 172° C., which primarily comprises cyclomethicone tetramers (i.e., n=4); Dow Corning® 344 fluid, having a viscosity of 2.5 centistokes and a boiling point of 178° C., which primarily comprises cyclomethicone pentamers (i.e., n=5); Dow Corning® 245 fluid, having a viscosity of 4.2 centistokes and a boiling point of 205° C., which primarily comprises a mixture of cyclomethicone tetramers and pentamers (i.e., n=4 and 5); and Dow Corning® 345 fluid, having a viscosity of 4.5 centistokes and a boiling point of 217° C., which primarily comprises a mixture of cyclomethicone tetramers, pentamers, and hexamers (i.e., n=4, 5, and 6).

Materials such as trimethylsiloxysilicate may also be used, which is a polymeric material corresponding to the general formula [( _CH2 ) _3SiO1 /2] _x [ _SiO2 ] _y , where x is an integer from about 1 to about 500 and y is an integer from about 1 to about 500. Commercially available trimethylsiloxysilicate is sold as Dow Corning 593 fluid in a mixture with dimethicone.

Dimethiconols are also suitable for use in the compositions. These compounds can be represented by the formulas _{R3SiO [ R2SiO} ] _xSiR2OH and _HOR2SiO [ _R2SiO ] _xSiR2OH , where R is an alkyl group (preferably methyl or ethyl, more preferably methyl), and x is an integer from 0 to about 500, with R and _x being selected to achieve the desired molecular weight. Commercially available dimethiconols are typically sold as mixtures with dimethicone or cyclomethicone (e.g., Dow Corning 1401, 1402, and 1403 fluids).

Polyalkylaryl siloxanes are also suitable for use in the compositions. Polymethylphenyl siloxanes having a viscosity of about 15 to about 65 centistokes at 25°C are particularly useful.

Organopolysiloxanes selected from the group consisting of polyalkylsiloxanes, alkyl-substituted dimethicones, cyclomethicone, trimethylsiloxysilicate, dimethiconol, polyalkylaryl siloxanes, and mixtures thereof are preferred for use in the present invention. Polyalkylsiloxanes and cyclomethicone are more preferred for use in the present invention. Among the polyalkylsiloxanes, dimethicone is preferred.

As described above, the continuous silicone phase may contain one or more non-silicone oils. The concentration of non-silicone oils in the continuous silicone phase is minimized or avoided, thereby further enhancing the oxidative stability of the selected retinoid in the composition. Suitable non-silicone oils have a melting point of about 25° C. at about one atmosphere of pressure. Examples of non-silicone oils suitable for use in the continuous silicone phase are those well known in the chemical art in the form of water-in-oil emulsions in topical personal care products, such as mineral oils, vegetable oils, synthetic oils, semi-synthetic oils, and the like.

(2) Dispersed water phase

The topical compositions disclosed herein comprise from about 30% to about 90%, more preferably from about 50% to about 85%, and even more preferably from about 70% to about 80%, of a dispersed aqueous phase. In emulsion technology, the term "dispersed phase" is a term well known to those skilled in the art and refers to the phase as small particles or droplets suspended in and surrounded by a continuous phase. The dispersed phase is also referred to as the internal phase or discontinuous phase. The dispersed aqueous phase is a dispersion of small aqueous particles or droplets suspended in and surrounded by the continuous silicone phase described above.

The aqueous phase can be water, or a combination of water and one or more water-soluble or dispersible components. Non-limiting examples of such components include thickeners, acids, bases, salts, chelating agents, gums, water-soluble or dispersible alcohols and polyols, buffers, preservatives, sunscreens, colorants, and the like.

The topical compositions disclosed herein typically comprise from about 25% to about 90%, preferably from about 40% to about 80%, more preferably from about 60% to about 80%, of water in the dispersed phase by weight of the composition.

(3) Emulsifier used to disperse the aqueous phase

The water-in-silicone emulsions disclosed herein preferably contain an emulsifier. In preferred embodiments, the composition comprises from about 0.1% to about 10% emulsifier, more preferably from about 0.5% to about 7.5%, and even more preferably from about 1% to about 5%, by weight of the composition. The emulsifier helps disperse and suspend the aqueous phase within the continuous silicone phase.

A variety of emulsifying agents can be used in the present invention to form the preferred water-in-silicone emulsions. Known or conventional emulsifying agents can be used in the compositions, provided that the selected emulsifying agent is chemically and physically compatible with the ingredients of the compositions disclosed herein and provides the desired dispersion characteristics. Suitable emulsifiers include silicone emulsifiers, non-silicone emulsifiers, and mixtures thereof known to those skilled in the art for use in topical personal care products. Preferably, these emulsifiers have an HLB value of about 14 or less, more preferably from about 2 to about 14, and even more preferably from about 4 to about 14. Emulsifiers having HLB values outside the above ranges can be used in combination with other emulsifiers so that the combination achieves an effective weighted average HLB within these ranges.

Silicone emulsifiers are preferred. A variety of silicone emulsifiers can be used in the present invention. These silicone emulsifiers are typically organically modified organopolysiloxanes, also referred to by those skilled in the art as silicone surfactants. Useful silicone emulsifiers include dimethicone copolyols. These materials are polydimethylsiloxanes that have been modified to contain polyether side chains, such as polyethylene oxide chains, polypropylene oxide chains, mixtures of these chains, and polyether chains containing moieties derived from ethylene oxide and propylene oxide. Other examples include alkyl-modified dimethicone copolyols, i.e., compounds containing C2-C30 pendant side chains. Other dimethicone tool polyols that can be used include those with a variety of cationic, anionic, amphoteric, and zwitterionic pendant moieties.

The dimethicone copolyol emulsifiers useful in the present invention can be described by the following general structure:

wherein R is a C ₁ -C ₃₀ linear, branched or cyclic alkyl group, and R ² is selected from:

-(CH ₂ ) _n -O-(CH ₂ CHR ³ O) _m -H,

and

-(CH ₂ )nO-(CH ₂ CHR ³ O) _m -(CH ₂ CHRO)oH,

wherein n is an integer from 3 to about 10; ^R3 and ^R4 are selected from H and _C1 - _C6 linear or branched alkyl groups, such that ^R3 and ^R4 cannot be the same at the same time; and m, o, x, and y are selected so that the molecule has an overall molecular weight of from about 200 to about 10,000,000, and m, o, x, and y are independently selected from integers of 0 or greater, such that m and o cannot be simultaneously 0, and z is independently selected from an integer of 1 or greater. It is recognized that positional isomers of these copolyols can be obtained. The chemical representations depicted above for the ^R2 portion comprising the ^R3 and ^R4 groups are not intended to be limiting and are shown for convenience.

Although the silicone surfactants depicted in the structure of the above paragraph are not strictly classified as dimethicone copolyols, the silicone surfactants can also be used in the present invention where ^R2 is:

-(CH ₂ ) _n -OR ⁵ ,

wherein R ⁵ is a cationic, anionic, amphoteric or zwitterionic moiety.

Non-limiting examples of dimethicone copolyols and other silicone surfactants that can be used as emulsifiers in the present invention include dimethicone polyether copolymers having pendant polyethylene oxide side chains, dimethicone polyether copolymers having pendant polypropylene oxide side chains, dimethicone polyether copolymers having pendant mixed polyethylene oxide and polypropylene oxide side chains, dimethicone polyether copolymers having pendant mixed polyoxy(ethylene)(propylene) side chains, dimethicone polyether copolymers having pendant organic betaine side chains, dimethicone polyether copolymers having pendant carboxylate side chains, dimethicone polyether copolymers having pendant quaternary ammonium side chains; and further modifications of the foregoing copolymers to contain pendant _C2 - _C30 linear, branched, or cyclic alkyl moieties. Examples of commercially available dimethicone copolyols sold by Dow Corning Corporation that can be used in the present invention are Dow Corning 190, 193, Q2-5220, 2501 Wax, 2-5324 Fluid, and 3225C (this material is sold as a mixture with cyclomethicone). Cetyl dimethicone copolyol is commercially available as a mixture with polyglyceryl-4-isostearate (and) hexyl laurate, and is sold under the tradename WE-09 (available from Goldschmidt). Cetyl dimethicone copolyol is also commercially available as a mixture with hexyl laurate (and) polyglyceryl-3 oleate (and) cetyl dimethicone, and is sold under the tradename WS-08 (also available from Goldschmidt). Other non-limiting examples of dimethicone copolyols include dialkyl dimethicone copolyols, dimethicone copolyol acetates, dimethicone copolyol adipic acid, dimethicone copolyol amines, dimethicone copolyol behenate, dimethicone copolyol butyl ether, dimethicone copolyol hydroxystearate, dimethicone copolyol isostearate, dimethicone copolyol laurate, dimethicone copolyol methyl ether, dimethicone copolyol phosphates, and dimethicone copolyol stearate. See International Cosmetic Ingredient Dictionary, Fifth Edition, 1993.

Dimethicone copolyol emulsifiers that can be used in the present invention are described, for example, in U.S. Patent No. 4,960,764, issued to Figueroa et al. on October 2, 1990; in European Patent No. EP 330,369, published by Sano Gueira on August 30, 1989; G. H. Dahms, et al., "New Formulation Possibilities Offered by Silicone Copolyols," Cosmetics & Toiletries, vol. 110, pp. 91-100, March 1995; M. E. Carlotti et al., Optimization of W/O-S Emulsions And Study Of The Quantitative Relationships Between Ester Structure And Emulsion Properties," J. Dispersion Science And Technology, 13(3), 315-336 (1992); P. Hameyer, "Comparative Technological Investigations of Organic and OrganosiliconeEmulsifiers in Cosmetic Water-in-Oil Emulsion Preparations,"HAPPI 28(4),pp.88-128(1991); J.Smid-Korbar et al.,"Efficiency and usability of siliconesurfactants in emulsions,"Provisional Communication International Journal ofCosmetic Science, 12, 135-139 (1990); and D.G. Krzysik et al., "A New Silicone Emulsifier For Water-in-Oil Systems," Drug and Cosmetic Industry, vol. 146 (4) pp. 28-81 (April 1990).

A variety of nonionic and anionic emulsifying agents are included in the silicone-free emulsifiers used in the present invention, such as sugar esters and polyesters, alkoxylated sugar esters and polyesters, _C1 - _C30 fatty acid esters of _C1 - _C30 fatty alcohols, alkoxylated derivatives of _C1 - _C30 fatty acid esters of _C1 - _C30 fatty alcohols, alkoxylated esters of _C1 - _C30 fatty alcohols, polyglyceryl esters of _C1 - _C30 fatty acids, _C1 - _C30 esters of polyols, _C1 - _C30 ethers of polyols, alkyl phosphates, polyethylene glycol fatty ether phosphates, fatty acid amides, acyl lactylates, soaps, and mixtures thereof. Other suitable emulsifiers are described, for example, in McCutcheon's, Detergents and Emulsifiers, North American Edition (1986), Allured Publishing Corporation; U.S. Pat. No. 5,011,681, issued April 30, 1991 to Ciotti et al.; U.S. Pat. No. 4,421,769, issued December 20, 1983 to Dixon et al.; and U.S. Pat. No. 3,755,560, issued August 28, 1973 to Dickert et al.

Non-limiting examples of these silicone-free emulsifiers include: polyethylene glycol 20 sorbitan monolaurate (polysorbate 20), polyethylene glycol 5 soya sterol, steareth-20, ceteareth-20, PPG-2 methyl glucose ether distearate, ceteth-10, polysorbate 80, cetyl phosphate, potassium cetyl phosphate, diethanolamine cetyl phosphate, polysorbate 60, glyceryl stearate, PEG-1 00 Stearate, Polyoxyethylene 20 Sorbitan Trioleate (Polysorbate 85), Sorbitan Monolaurate, Polyoxyethylene 4 Lauryl Ether Sodium Stearate, Polyglyceryl-4 Isostearate, Hexyl Laurate, Steareth-20, Ceteareth-20, PPG-2 Methyl Glucose Ether Distearate, Ceteth-10, Diethanolamine Cetyl Phosphate, Glyceryl Stearate, PEG-100 Stearate and mixtures thereof.

B) Oil-in-water emulsion

Other preferred topical carriers include oil-in-water emulsions having a continuous aqueous phase and a hydrophobic, water-insoluble phase ("oil phase") dispersed therein. Examples of suitable oil-in-water emulsion carriers are described in U.S. Patent Nos. 1 to Turner, D.J. et al., issued December 17, 1991; and 5,073,372 to Turner, D.J. et al., issued December 17, 1991. Particularly preferred oil-in-water emulsions comprising a structuring agent, a hydrophilic surfactant, and water are described in detail below.

(1) Structural reagents

Preferred oil-in-water emulsions contain a structuring agent that aids in the formation of a liquid crystal gel network structure. Without being bound by theory, it is believed that the structuring agent helps provide rheological characteristics to the composition, which contributes to the stability of the composition. The structuring agent may also function as an emulsifier or surfactant. Preferred compositions contain from about 0.5% to about 20%, more preferably from about 1% to about 10%, and even more preferably from about 1% to about 5%, of the structuring agent, based on the weight of the composition.

Preferred structuring agents disclosed herein include stearic acid, palmitic acid, stearyl alcohol, cetyl alcohol, behenyl alcohol, stearic acid, palmitic acid, polyethylene glycol ethers of stearyl alcohol (having an average of about 1 to about 21 ethylene oxide units), polyethylene glycol ethers of cetyl alcohol (having an average of about 1 to about 5 ethylene oxide units), and mixtures thereof. More preferred structuring agents disclosed herein are selected from stearyl alcohol, cetyl alcohol, behenyl alcohol, polyethylene glycol ethers of stearyl alcohol (having an average of about 2 ethylene oxide units (steareth-2)), polyethylene glycol ethers of stearyl alcohol (having an average of about 21 ethylene oxide units (steareth-21)), polyethylene glycol ethers of cetyl alcohol (having an average of about 2 ethylene oxide units), and mixtures thereof. Even more preferred structuring agents are selected from stearic acid, palmitic acid, stearyl alcohol, cetyl alcohol, behenyl alcohol, steareth-2, steareth-21, and mixtures thereof.

(2) Hydrophilic surfactant

Preferred oil-in-water emulsions contain from about 0.05% to about 10%, preferably from about 1% to about 6%, more preferably from about 1% to about 3% of at least one hydrophilic surfactant that can disperse the hydrophobic material in the aqueous phase (based on the weight of the topical vehicle). The surfactant must be at least sufficiently hydrophilic to be dispersible in water.

Preferred hydrophilic surfactants are selected from nonionic surfactants. Among the nonionic surfactants useful in the present invention are those that can be broadly defined as condensation products of long-chain alcohols (e.g., _C8-30 alcohols) with sugar or starch polymers (i.e., glycosides). These compounds can be represented by the formula (S) _n -OR, where S is a sugar moiety, such as glucose, fructose, mannose, and galactose; n is an integer from about 1 to about 1000; and R is a _C8-30 alkyl group. Examples of long-chain alcohols from which alkyl groups can be derived include decyl alcohol, hexadecanol, stearyl alcohol, dodecyl alcohol, tetradecyl alcohol, oleyl alcohol, and the like. Preferred examples of these surfactants include those in which S is a glucose moiety, R is a _C8-20 alkyl group, and n is an integer from about 1 to about 9. Commercially available examples of these surfactants include decyl polyglycoside (available as APG 325 CS from Henkel) and dodecyl polyglycoside (available as APG 600 CS from Henkel).

Other nonionic surfactants that can be used include condensation products of ethylene oxide with fatty acids (i.e., ethylene oxide esters of fatty acids). These materials have the general formula RCO(X) _nOH , where R is a _C10-30 alkyl group, _X is _-OCH2CH2- (i.e., derived from ethylene glycol or an oxide) or _-OCH2CHCH3- (i.e., derived from propylene glycol or an oxide), and n is an integer from about 6 to about 200. These materials have _the general formula RCO(X) _nOOCR , where R is a _C10-30 alkyl group, X is _-OCH2CH2- (i.e., derived from ethylene glycol or an _oxide ) _{or -OCH2CHCH3-} (i.e., derived from propylene glycol or an oxide), and n is an integer from about 6 to about 100. Other nonionic surfactants are condensation products of ethylene oxide with fatty alcohols (i.e., ethylene oxide ethers of fatty alcohols). These materials have the general formula R(X) _nOR ', where R is a _C10-30 alkyl group, X is _{-OCH2CH2- (} i.e., derived from ethylene glycol or an oxide) or _-OCH2CHCH3- (i.e., derived from propylene glycol or an oxide), and n is an integer from about 6 to about 100, and R' is _H or a _C10-30 alkyl group. Other nonionic surfactants are condensation products of ethylene oxide with fatty acids and fatty alcohols [i.e., where one end of the polyoxyalkylene moiety is branched with a fatty acid and the other end is etherified (i.e., linked by an ether linkage) with a fatty alcohol]. These materials have the general formula RCO(X) _nOR ', where R and R' are _C10-30 alkyl _groups , X is _-OCH2CH2- (i.e., derived from ethylene glycol or an oxide) or _-OCH2CHCH3- (i.e., derived from propylene glycol or an oxide), and n is _an integer from about 6 to about 100. Non-limiting examples of these alkylene oxide nonionic surfactants include ceteth-6, ceteth-10, ceteth-12, ceteareth-6, ceteareth-10, ceteareth-12, steareth-6, steareth-10, steareth-12, steareth-21, PEG-6 stearate, PEG-10 stearate, PEG-100 stearate, PEG-12 stearate, PEG-20 glyceryl stearate, PEG-80 glyceryl tallow, PEG-10 glyceryl stearate, PEG-30 glyceryl cocoate, PEG-80 glyceryl cocoate, PEG-200 glyceryl tallow, PEG-8 dilaurate, PEG-10 distearate, and mixtures thereof.

Other nonionic surfactants that can be used include polyhydroxy fatty acid amide surfactants corresponding to the following structural formula:

wherein: ^R1 is H, _dC4 alkyl, 2-hydroxyethyl, or 2-hydroxypropyl, preferably _dC4 alkyl, more preferably methyl or ethyl, and most preferably methyl; ^R2 is _C5 - _C31 alkyl or alkenyl, preferably _C7 - _C19 alkyl or alkenyl, more preferably C9- _C17 alkyl or alkenyl, and most preferably _C11 - _{C15 alkyl} or alkenyl; and Z is a polyhydroxy hydrocarbyl moiety (having a linear hydrocarbyl chain with at least three hydroxyl groups directly attached to the chain) or an alkoxylated derivative thereof (preferably ethoxylated or propoxylated). Z is preferably a sugar moiety selected from glucose, fructose, maltose, lactose, galactose, mannose, xylose, and mixtures thereof. A particularly preferred surfactant corresponding to the above structure is coconut alkyl N-methyl glycoside amide (i.e., wherein the ^R2CO- moiety is derived from coconut oil fatty acid). Processes for preparing compositions containing polyhydroxy fatty acid amides are disclosed, for example, in GB Patent Specification 809,060 to Thomas Hedley & Co., Ltd., published February 18, 1959; U.S. Patent No. 2,965,576, issued to E.R. Wilson on December 20, 1960; U.S. Patent No. 2,703,798, issued to A.M. Schwartz on March 8, 1955; and U.S. Patent No. 1,985,424, issued to Piggott on December 25, 1934, all of which are incorporated herein by reference in their entirety.

Among the nonionic surfactants, those selected from the group consisting of Steareth-21, Ceteareth-20, Ceteareth-12, Sucrose Cocoate, Steareth-100, PEG-100 Stearate and mixtures thereof are preferred.

Other nonionic surfactants suitable for use in the present invention include sugar esters and polyesters, alkoxylated sugar esters and polyesters, d-C30 fatty acid esters of fatty alcohols, alkoxylated derivatives of C1-C30 fatty acid esters of fatty alcohols, alkoxylated ethers of fatty alcohols, polyglycerol esters of d-C30 fatty acids, C1-C30 esters of polyols, C1-C30 esters of polyols, alkyl phosphates, polyoxyalkylene fatty ether phosphates, fatty acid amides, acyl lactylates and mixtures thereof. Non-limiting examples of these emulsifiers include: polyethylene glycol 20 sorbitan monolaurate (polysorbate 20), polyethylene glycol 5 soya sterol, steareth-20, ceteareth-20, PPG-2 methyl glucose ether distearate, ceteth-10, polysorbate 80, cetyl phosphate, potassium cetyl phosphate, diethanolamine cetyl phosphate, polysorbate 60, glyceryl stearate, polyoxyethylene 20 sorbitan trioleate (polysorbate 85), sorbitan monolaurate, polyoxyethylene 4 lauryl ether sodium stearate, polyglyceryl-4 isostearate, hexyl laurate, PPG-2 methyl glucose ether 5 distearate, PEG-100 stearate, and mixtures thereof.

Another group of nonionic surfactants that can be used in the present invention are fatty acid ester blends based on mixtures of sorbitan or sorbitol fatty acid esters and sucrose fatty acids, in each case preferably having a _C8 - _C24 fatty acid, more preferably a _C10 - _C20 fatty acid. Preferred fatty acid ester emulsifiers are blends of sorbitan or sorbitol _C16 - _C20 fatty acid esters and sucrose _C10 - _C16 fatty acid esters, particularly sorbitan stearate and sucrose cocoate. These are available from ICI under the trade name Arlatone 2121.

Other suitable surfactants that can be used in the present invention include comparative cationic, anionic, zwitterionic and amphoteric surfactants, such as those known in the art and discussed more fully below. See, for example, McCutcheon's, Detergents and Emulsifiers, North American 15th Edition (1986), published by Allured Publishing Corporation; U.S. Patent No. 5,011,681, issued to Ciotti et al. on April 30, 1991; U.S. Patent No. 4,421,769, issued to Dixon et al. on December 20, 1983; and U.S. Patent No. 3,755,560, issued to Dickert et al. on December 20, 1973; all four references are incorporated herein by reference in their entirety. The hydrophilic surfactants that can be used in the present invention can include a single surfactant or any combination of suitable surfactants. The exact 20 surfactants (or surfactants) selected will depend on the pH of the combination and the other ingredients present.

Cationic surfactants can also be used in the present invention, particularly dialkyl quaternary ammonium compounds, examples of which are described in U.S. Patent No. 5,151,209; U.S. Patent No. 5,151,210; U.S. Patent No. 5,120,532; U.S. Patent No. 4,387,090; U.S. Patent No. 253,155,591; U.S. Patent No. 3,929,678; U.S. Patent No. 3,959,461; McCutcheon's Deterrents & Emulsifiers, (North American edition 1979) M.C. Publishing Co.; and Schwartz, et al., Surface Active Agents, Their Chemistry and Technology, New York: Interscience Publishers, 1949, which descriptions are incorporated herein by reference. Cationic surfactants that can be used in the present invention include cationic ammonium salts, such as those having the formula:

wherein R ₁ is an alkyl group having from about 12 to about 30 carbon atoms, or an aromatic, aryl, or alkaryl group having from about 12 to about 30 carbon atoms; R ₂ , R ₃ , and R ₄ are independently selected from hydrogen, an alkyl group having from about 1 to about 22 carbon atoms, or an aromatic, aryl, or alkaryl group having from about 12 to about 22 carbon atoms; and X is any compatible anion, preferably selected from chloride, bromide, iodide, acetate, phosphate, nitrate, sulfate, methylsulfate, ethylsulfate, tosylate, lactate, citrate, glycolate, and mixtures thereof. Furthermore, the alkyl groups of R ₁ , R ₂ , R ₃ , and R ₄ may also contain ester and/or ether linkages, or hydroxyl or amino group substituents (e.g., the alkyl groups may contain polyethylene glycol and polypropylene glycol moieties).

More preferably, _R1 is an alkyl group having from about 12 to about 22 carbon atoms; _R2 is selected from H, or an alkyl group having from about 1 to about 22 carbon atoms; _R3 and _R4 are independently selected from H, or an alkyl group having from about 1 to about 3 carbon atoms; and X is as described above.

Still more preferably, R ₁ is an alkyl group having from about 12 to about 22 carbon atoms; R ₂ , R ₃ and R ₄ are selected from H, or an alkyl group having from about 1 to about 3 carbon atoms; and X is as described above.

Alternatively, other cationic emulsifiers that can be used include amino-amides, wherein in the structure described above, R ₁ is alternatively R ₅ CONH-(CH ₂ ) _n , wherein R ₅ is an alkyl group having from about 12 to about 22 carbon atoms, and n is an integer from about 2 to about 6, more preferably from about 2 to about 4, and even more preferably from about 2 to about 3. Non-limiting examples of these cationic emulsifiers include stearamidopropyl PG-dimonium chloride phosphate, behenamidopropyl PG dimonium chloride, stearamidopropyl ethyldimonium ethosulfate, stearamidopropyl dimethyl (tetradecyl acetate) ammonium chloride, stearamidopropyl dimethyl cetearyl ammonium toluenesulfonate, stearamidopropyl dimethyl ammonium chloride, stearamidopropyl dimethyl ammonium lactate, and mixtures thereof. Behenamidopropyl PG dimonium chloride is particularly preferred.

Non-limiting examples of quaternary ammonium salt cationic surfactants include those selected from the group consisting of hexadecyl ammonium chloride, hexadecyl ammonium bromide, dodecyl ammonium chloride, dodecyl ammonium bromide, stearyl ammonium chloride, stearyl ammonium bromide, hexadecyl dimethyl ammonium chloride, hexadecyl dimethyl ammonium bromide, dodecyl dimethyl ammonium chloride, dodecyl dimethyl ammonium bromide, stearyl dimethyl ammonium chloride, stearyl dimethyl ammonium bromide, hexadecyl trimethyl ammonium chloride, hexadecyl trimethyl ammonium bromide, dodecyl trimethyl ammonium chloride, dodecyl trimethyl ammonium bromide, Ammonium bromide, stearyl trimethyl ammonium chloride, stearyl trimethyl ammonium bromide, dodecyl dimethyl ammonium chloride, stearyl dimethyl hexadecyl ditallow dimethyl ammonium chloride, dihexadecyl ammonium chloride, dihexadecyl ammonium bromide, didodecyl ammonium chloride, didodecyl ammonium bromide, distearyl ammonium chloride, distearyl ammonium bromide, dihexadecyl methyl ammonium chloride, dihexadecyl methyl ammonium bromide, didodecyl methyl ammonium chloride, didodecyl methyl ammonium bromide, distearyl methyl ammonium chloride, distearyl methyl ammonium bromide and mixtures thereof. Other quaternary ammonium salts include those in which the C ₁₂ to C ₃₀ alkyl carbon chain is derived from tallow fatty acid or coconut fatty acid. The term "tallow" refers to an alkyl group derived from tallow fatty acid (typically hydrogenated tallow fatty acid), typically having a mixture of alkyl chains ranging from C ₁₆ to C ₁₈ . The term "coconut" refers to alkyl groups derived from coconut fatty acids, which typically have a mixture of alkyl chains in the _C12 to _C14 range. Examples of quaternary ammonium salts derived from these tallow and coconut sources include ditallowdimethylammonium chloride, ditallowdimethylammonium methylsulfate, di(hydrogenated tallow)dimethylammonium chloride, di(hydrogenated tallow)dimethylammonium acetate, ditallowdipropylammonium phosphate, ditallowdimethylammonium nitrate, di(coconut alkyl)dimethylammonium chloride, di(coconut alkyl)dimethylammonium bromide, tallow ammonium chloride, coconut ammonium chloride, stearamidopropyl PG-dimonium chloride phosphate, stearamidopropyl ethyldimonium ethosulfate, stearamidopropyl dimethyl (tetradecyl acetate) ammonium chloride, stearamidopropyl dimethyl cetearyl ammonium p-toluenesulfonate, stearamidopropyl dimethylammonium chloride, stearamidopropyl dimethylammonium lactate, and mixtures thereof. An example of a quaternary ammonium compound (having an ester bond) having an alkyl group is ditallowyloxyethyldimethylammonium chloride.

More preferred cationic surfactants are those selected from the group consisting of behenamidopropyl PG dimonium chloride, didodecyldimonium chloride, distearyldimonium chloride, ditetradecyldimonium chloride, dihexadecyldimonium chloride, distearyldimonium chloride, stearamidopropyl PG-dimonium chloride phosphate, stearamidopropyl ethyldimonium ethosulfate, stearamidopropyl dimethyl (tetradecyl acetate) ammonium chloride, stearamidopropyl dimethyl cetearyl p-toluenesulfonate, stearamidopropyl dimethyl ammonium chloride, stearamidopropyl dimethyl ammonium lactate, and mixtures thereof.

Also more preferred cationic surfactants are those selected from the group consisting of behenamidopropyl PG dimonium chloride, didodecyldimonium chloride, distearyldimonium chloride, ditetradecyldimonium chloride, dicetyldimonium chloride, and mixtures thereof.

A preferred combination of cationic surfactant and structuring agent is behenamidopropyl PG dimonium chloride and/or behenyl alcohol, wherein the ratios are preferably optimized to maintain and enhance physical and chemical stability, particularly when such combinations contain ionic and/or highly polar solvents. This composition is particularly useful for the delivery of sunscreen agents such as zinc oxide and octyl methoxycinnamic acid.

A variety of anionic surfactants may also be used in the present invention. See, for example, U.S. Patent No. 3,929,678, issued to Laughlin et al. on December 30, 1975, which is incorporated herein by reference in its entirety. Non-limiting examples of anionic surfactants include alkanoyl isethionates, and alkyl and alkyl ether sulfates. Alkanoyl isethionates generally have the formula: RCO—OCH ₂ CH ₂ SO ₃ M, wherein R is an alkyl or alkenyl group having from about 10 to about 30 carbon atoms, and M is a water-soluble cation such as ammonium, sodium, potassium, and triethanolamine. Non-limiting examples of these isethionates include alkanoyl isethionates selected from the group consisting of ammonium cocoyl isethionate, sodium cocoyl isethionate, sodium lauroyl isethionate, sodium stearoyl isethionate, and mixtures thereof.

Alkyl and alkyl ether sulfates generally have the following formats: ROSO ₃ M and RO(C ₂ H ₄ O) _x SO ₃ M, respectively, where R is an alkyl or alkenyl group of about 10 to about 30 carbon atoms, x is from about 1 to about 10, and M is a water-soluble cation such as ammonium, sodium, potassium, and triethanolamine. Another class of suitable anionic surfactants are the water-soluble salts of organic sulfuric acid reaction products of the general formula:

R-SO ₃ -M

wherein _R is selected from a linear or branched saturated aliphatic hydrocarbon radical having from about 8 to about 24, preferably from about 10 to about 16, carbon atoms; and M is a cation. Other anionic synthetic surfactants include the designated succinamates, olefin sulfonates, and β-alkoxyalkyl sulfonates having from about 12 to about 24 carbon atoms. Examples of these materials are sodium lauryl sulfate and ammonium lauryl sulfate.

Other anionic materials useful in the present invention are soaps of fatty acids (i.e., alkali metal salts, such as sodium or potassium salts), which typically have from about 8 to about 24 carbon atoms, preferably from about 10 to about 20 carbon atoms. The fatty acids used to prepare the soaps can be obtained from natural sources, such as plant or animal derived glycerides (e.g., palm oil, coconut oil, soybean oil, castor oil, tallow, lard, etc.). The fatty acids can also be prepared synthetically. Soaps are described in more detail in U.S. Patent No. 4,557,853.

Amphoteric and zwitterionic surfactants can also be used in the present invention. Examples of amphoteric and zwitterionic surfactants that can be used in the compositions disclosed herein are broadly described as secondary and tertiary aliphatic amines, wherein the aliphatic radical can be straight or branched, and one of the aliphatic substituents contains from about 8 to about 22 carbon atoms (preferably _C8 - _C18 ), and one of the aliphatic substituents contains an anionic water-solubilizing group, such as carboxyl, sulfonic acid, sulfate _, phosphoric acid, or phosphonic acid. Examples are alkyl iminoacetates, and iminodialkanoates and aminoalkanoates of the formula RN[ _CH2 ) _mCO2M ] ₂ and RNH( _CH2 ) _mCO2M , wherein m is ₁ to 4, R is a _C8 - _C22 alkyl or alkenyl group, and M is H, alkali metal, alkaline earth metal ammonium, or alkylammonium. Also included are imidazoline and ammonium derivatives. Specific examples of suitable amphoteric surfactants include sodium 3-dodecylaminopropionate, sodium 3-dodecylaminopropanesulfonate, N-alkyltaurines, such as those prepared by reacting dodecylamine with sodium isethionate according to the teachings of U.S. Patent No. 2,658,072, which is incorporated herein by reference in its entirety; N-higher alkyl aspartic acids, such as those produced according to the teachings of U.S. Patent No. 2,438,091, which is incorporated herein by reference in its entirety; and products sold under the trade name "Miranol" and described in U.S. Patent No. 2,528,378, which is incorporated herein by reference in its entirety. Other examples of amphoteric substances that can be used include phosphates, such as cocamidopropyl PG-dimonium chloride phosphate (available as Monaquat PTC from Mona Corp.).

Other amphoteric or zwitterionic surfactants that can be used in the present invention include betaines. Examples of betaines include higher alkyl betaines such as coconut dimethyl carboxymethyl betaine, dodecyl dimethyl carboxymethyl betaine, dodecyl dimethyl α-carboxyethyl betaine, hexadecyl dimethyl carboxymethyl betaine, hexadecyl dimethyl betaine (Lonzaine 16SP from Lonza Corp.), dodecyl di-(2-hydroxyethyl) carboxymethyl betaine, stearyl di-(2-hydroxypropyl) carboxymethyl betaine, oleyl dimethyl γ-carboxypropyl betaine, dodecyl di-(2-hydroxypropyl) α-carboxyethyl betaine, coconut dimethyl sulfopropyl betaine, stearyl dimethyl sulfopropyl betaine, dodecyl dimethyl sulfoethyl betaine, dodecyl di-(2-hydroxyethyl) sulfopropyl betaine, amino betaines and amino sulfobetaines (where RCONH(CH ₂ ) ₃ free radical attached to the nitrogen atom of the betaine), oleyl betaine (available from Henkel's amphoteric Velvetex OLB-50), and cocoamidopropyl betaine (available from Henkel's Velvetex BK-35 and BA-35).

Other amphoteric and zwitterionic surfactants that can be used include the sultaines and hydroxysultaines, such as cocoamidopropyl hydroxysultaine (available as Mirataine CBS from Rhone-Poulenc), and the alkanoyl sarcosinates corresponding to the formula RCON( _CH3 ) _CH2CH2CO2M , wherein R is an alkyl or alkenyl group of about 10 to about 20 _carbon atoms and M is a water-soluble cation such as ammonium, sodium, potassium, and trialkanolamines (e.g., triethanolamine), a preferred example _of which is sodium lauroyl sarcosinate.

(3) Water

Preferred oil-in-water emulsions comprise from about 25% to about 98%, preferably from about 65% to about 95%, more preferably from about 70% to about 90% water by weight of the topical vehicle.

The hydrophobic phase is dispersed in the continuous aqueous phase. The hydrophobic phase may comprise water-insoluble or partially soluble materials, such as those known in the art, including but not limited to the silicone resins described herein for the silicone-in-water emulsions, and other oils and lipids, such as those described above for the emulsions.

Topical compositions disclosed herein (including but not limited to lotions and creams) may include dermatologically acceptable emollients. Such compositions preferably include about 1% to about 50% emollients. As used herein, "emollient" refers to a material for preventing or alleviating dryness, and for protecting the skin. A variety of suitable emollients are known and may be used in the present invention. Sagarin, Cosmetics Science and Technology, 2nd Edition, Vol. 1, pp. 32-43 (1972) (incorporated herein by reference) includes various examples of materials suitable for use as emollients. A preferred emollient is glycerol. Preferably, glycerol is used that is from about 0.001% to about 30%, more preferably from about 0.01% to about 20%, and even more preferably from about 0.1% to about 10% (e.g., 5%).

Lotions and creams according to the present disclosure generally comprise a solution carrier system and one or more emollients. Lotions and creams generally comprise from about 1% to about 50%, preferably from about 1% to about 20%, of an emollient; from about 50% to about 90%, preferably from about 60% to about 80%, of water; and pentapeptides and/or pentapeptide derivatives, and other skin care actives (or actives) in the amounts described above. Creams are generally thicker than lotions due to their higher levels of emollients or higher levels of thickeners.

The ointments disclosed herein may comprise a simple carrier base of an animal or vegetable oil or a semisolid hydrocarbon (oleaginous); an absorption ointment base that absorbs water to form an emulsion; or a water-soluble base, such as a water-soluble solution carrier. The ointment may further comprise a thickening agent (such as those described in Sagarin, Cosmetics, Science and Technology, 2nd Edition, Vol. 1, pp. 72-73 (1972), which is incorporated herein by reference), and/or an emollient. For example, the ointment may comprise from about 2% to about 10% of an emollient; from about 0.1% to about 2% of a thickening agent; and the above-mentioned amounts of pentapeptides and/or pentapeptide derivatives, and other skin care active substances (or multiple active substances).

The compositions disclosed herein for cleaning ("cleansers") are formulated using a suitable carrier, such as those described above, and preferably contain from about 1% to about 90%, more preferably from about 5% to about 10%, of a dermatologically acceptable surfactant, in addition to the above amounts of pentapeptides and/or pentapeptide derivatives, and other skin care actives (or actives). The surfactant is suitably selected from anionic, nonionic, zwitterionic, amphoteric and ampholytic surfactants, and mixtures of these surfactants. These surfactants are well known to those skilled in the art of cleansing. Non-limiting examples of viable surfactants include isoceteth-20, sodium methyl cocoyl taurate, sodium methyl oleoyl taurate, and sodium lauryl sulfate. For exemplary surfactants useful in the present invention, see U.S. Patent No. 4,800,197, issued to Kowcz et al. on January 24, 1989, which is incorporated herein by reference in its entirety. Examples of a wide variety of other surfactants that can be used in the present invention are described in McCutcheon's Detergents and Emulsifiers, North American Edition (1986) published by Allured Publishing Corporation.The cleaning compositions may optionally contain other materials conventionally used in cleaning compositions at their art-established levels.

The physical form of the cleansing composition is unimportant. The composition can be formulated, for example, as toilet bars, liquids, shampoos, body washes, hair conditioners, hair tonics, pastes, or mousses. Rinse-off cleansing compositions (e.g., shampoos) require a delivery system sufficient to deposit sufficient levels of the active agent on the skin and scalp. Preferred delivery systems involve the use of insoluble complexes. For a more complete disclosure of such delivery systems, see U.S. Patent No. 4,835,148, issued to Barford et al. on May 30, 1989.

As used herein, the term "foundation" refers to a liquid, semi-liquid, semi-solid, or solid skin cosmetic product, including but not limited to lotions, creams, gels, pastes, blocks, and the like. Foundations are typically applied to large areas of skin, such as the face, to provide a distinctive appearance. Foundations are typically used to provide a base for application of makeup, such as rouge, blush creams, powders, and the like, and tend to conceal skin imperfections and impart a smooth, even appearance to the skin. The foundations disclosed herein include a dermatologically acceptable carrier and may contain conventional ingredients, such as oils, colorants, pigments, emollients, fragrances, waxes, stabilizers, and the like. Exemplary carriers and other such components suitable for use in the present invention are described, for example, in PCT application WO 96/33689 to Canter et al., published October 31, 1996, and in U.K. patent GB 2274585, issued August 3, 1994.

Preparation of composition

The compositions used in the methods disclosed herein are generally prepared by conventional methods, such as those known in the art for preparing topical compositions. Such methods generally involve mixing the various components to a relatively homogeneous state in one or more steps under conditions of heating, cooling, application of a vacuum, or the like, or not.

Methods for regulating skin condition

Compositions disclosed in the present invention can be used to regulate mammal skin condition. This regulation of keratinous tissue condition can include preventive and therapeutic regulation. For example, such regulation methods are directed to thickening keratinous tissue (i.e., constructing the epidermis and/or dermis of the skin, and if used, also constructing the stratum corneum of nails and hair shafts) and preventing and/or delaying the decline of mammal skin, preventing and/or delaying the appearance of spider veins and/or red erythema on mammal skin, preventing and/or delaying the appearance of dark circles under the eyes of mammals, preventing and/or delaying the sallowness of unsatisfactory mammal skin, preventing and/or delaying the sagging of mammal skin, softening and/or smoothing mammal lips, hair and nails, preventing and/or alleviating scabies on mammal skin, regulating skin texture (e.g., wrinkles and fine lines), and improving skin color (e.g., redness, freckles).

Regulating the condition of keratinous tissue involves topically applying a safe and effective amount of the compositions disclosed herein to the keratinous tissue. The amount of composition that can be applied, the frequency of application, and the duration of use will vary widely depending on the level of pentapeptide and/or pentapeptide derivative, and other skin care actives or actives in a given composition, as well as the level of regulation desired (e.g., depending on the level of keratinous tissue damage that is already present or is expected to occur).

In a preferred embodiment, described composition is applied on the skin for a long time. " long-term local application " refers to the time of the prolongation in the life course of study subject, preferably in the time at least about 1 week, more preferably in the time at least about 1 month, even more preferably in at least about 3 months, even more preferably in at least about 6 months, and more preferably in at least about 1 year, the described composition of local application is continuously applied. Although benefit can be obtained after various longest-time uses (for example 5 years, 10 years or 20 years), preferably long-term application continues the life course of whole study subject. Usually, in the time of described prolongation, apply in the rule of about once a day, but apply frequency and can change between about 1 time to every day about 3 times or more weekly.

The compositions disclosed herein can be used in a wide range of amounts to provide skin appearance and/or feel benefits. Typically, the amount of the compositions disclosed herein applied per application is 1 mg of composition/ ^cm2 of skin, ranging from about 0.1 mg/ ^cm2 to about 10 mg/ ^cm2 . Particularly useful application amounts are from about 1 mg/ ^cm2 to about 2 mg/ ^cm2 .

Regulating the condition of keratinous tissue is preferably accomplished by applying a composition in the form of a skin lotion, cream, gel, foam, ointment, paste, emulsion, spray, conditioner, tonic, makeup, lipstick, foundation, nail polish, lotion, or the like, preferably left on the skin or other keratinous structures to achieve some aesthetic, prophylactic, therapeutic, or other benefit (i.e., a "leave-on" composition). After the composition is applied to the skin, it is preferably left on the skin for at least about 15 minutes, more preferably at least about 30 minutes, even more preferably at least about 1 hour, and even more preferably at least several hours (e.g., up to about 12 hours). Any part of the exterior of the face, hair, and/or nails can be treated, such as the face, lips, area under the eyes, eyelids, scalp, neck, trunk, arms, hands, legs, feet, fingernails, toenails, scalp, eyelashes, eyebrows, and the like. The composition can be applied with the fingers or using an implement or instrument (e.g., a pad, cotton ball, applicator, sprayer, etc.).

Another method of ensuring that the skin is continuously exposed to the lowest level of pentapeptides and/or pentapeptide derivatives, and other skin care active substances (or multiple active substances) is to apply the compound by using a patch applied to, for example, the face. This method is particularly useful for problem skin areas (e.g., facial crow's feet, frown lines, under-eye areas, etc.) that require more intensive treatment. The patch can be closed, semi-closed, or non-closed, and can be adhesive or non-adhesive. The pentapeptides and/or pentapeptide derivatives, and other skin care active substances (or multiple active substances) compositions can be included in the patch, or applied to the skin before applying the patch. The patch also contains other active substances, such as chemical initiators for exothermic reactions, such as those described in U.S. Patent Nos. 5,821,250, 5,981,547, and 5,972,957 to Wu et al. The patch is preferably left on the skin for a period of at least about 5 minutes, more preferably at least about 15 minutes, more preferably at least about 30 minutes, even more preferably at least about 1 hour, and even more preferably overnight as a nighttime treatment.

In the following description, for purposes of explanation, numerous details are set forth to provide a thorough understanding of the embodiments. However, it will be apparent to anyone skilled in the art that these specific details are not required.

Example

Materials and methods

General synthetic procedure for glycosylated Pal-KTTKS derivatives

According to the Pal-KTTKS sequence, four groups of similarly modified pentapeptides were prepared by glycosylation of all feasible sites using a variety of sugars (Glc, Gal, GlcNAc, GalNAc, Man, Mai, Lac, Rha, Cel, Xyl, Fuc) in full acetylated and deacetylated forms. By detecting the secretion of elastin, fibronectin and total collagen on in vitro human skin fibroblast cultures and the secretion and history of elastin in an in vitro 3D model of human skin (composed of human epidermal keratinocytes and human skin fibroblasts), these synthetic compounds were subsequently tested for toxicity and biological activity. For the four groups of similarly modified peptides prepared, chemical properties were confirmed by HPLC and mass spectrometry.

Group 1 modifications involve glycosylation of one or both of the two middle threonines in the Pal-KTTKS sequence using fully acetylated or deacetylated forms of glucose, N-acetylglucosamine, galactose, N-acetylgalactosamine, or combinations thereof.

Group 2 modifications involve glycosylation of the terminal serine in the Pal-KTTKS sequence using glucose, N-acetylglucosamine, galactose, N-acetylgalactosamine in their fully acetylated or deacetylated forms.

Group 3 modifications involve glycosylation of the first threonine in the Pal-KTTKS sequence with mannose or maltose in fully acetylated or deacetylated form. Group 3 modifications also include substitution of the first threonine in the Pal-KTTKS sequence with serine, wherein the serine is glycosylated with glucose in fully acetylated or deacetylated form. Group 3 modifications also include substitution of the first threonine in the Pal-KTTKS sequence with asparagine, wherein the asparagine is glycosylated with glucose in fully acetylated or deacetylated form. Group 3 modifications also include addition of an asparagine amino acid to the N-terminus of the Pal-KTTKS sequence, wherein the asparagine is glycosylated with glucose in fully acetylated or deacetylated form.

Group 4 modifications involve glycosylation of the first or second threonine in the Pal-KTTKS sequence, or glycosylation of the terminal serine in the Pal-KTTKS sequence, using fully acetylated forms of galactose, maltose, lactose, rhamnose, cellobiose, xylose, or trehalose.

All chemical reagents were obtained from commercial suppliers of reagent grade or HPLC grade and used without further purification. Dichloromethane (DCM) was dried by distillation from calcium hydride ( _CaH2 ). Unless otherwise stated, all reactions were performed at room temperature. All non-glycosylated Fmoc-protected amino acids used for glycosylated peptide synthesis were obtained from Novabiochem (EMD Millipore). All glycosylated Fmoc-protected amino acids (glycosylated amino acids or glycosylated amino acids) used for glycosylated peptide synthesis were purchased from Sussex Research Laboratories Inc. (Ottawa, Canada) ( http://www.sussex-research.com/products/glycoamino-acids/all-glycoamino-acids/ ) . Preloaded resin (HL-Ser(t-Bu)-2-CI-trityl resin) was purchased from Matrix Innovation (Quebec, Canada). Standard resin (Novabiochem 2-chlorotrityl chloride resin) was obtained from EMD Millipore. Palmitic acid (purity >99%) was purchased from Sigma Aldrich.

Reverse phase high pressure liquid chromatography (RP-HPLC) was performed on a Gilson chromatograph equipped with an 805 Manometric module (maximum pressure 60 MPa), an 811C dynamic mixer, a 305 pump, and a UV/VIS-155 detector. A Phenomenex Luna C18 column (5 μm particle size, 250 x 4.60 mm analytical column) was used. Fractions were composed of mobile phases A and B (water/TFA (100/0.1) and acetonitrile/TFA (100/0.1), respectively). Glycosylated peptides were eluted from the column using a linear gradient from 2% to 70% mobile phase B over 40 minutes at a flow rate of 1 mL/min using UV detection at 214 nm. Preparative HPLC was performed on a Gilson chromatograph (805 Manometric module, maximum pressure 60 MPa; 811C dynamic mixer, 305 pump, UV/VIS-155 detector) using a Phenomenex Luna C18 column (10 μm particle size, 250 x 21.20 mm semi-preparative column). Mobile phases A and B consisted of water/TFA (100/0.1) and acetonitrile/TFA (100/0.1), respectively. Glycosylated peptides were separated from impurities using a linear gradient from 2% to 70% mobile phase B over 35 minutes at a flow rate of 10 mL/min using UV detection at 214 nm.

Analysis of peptide samples was performed on an electrospray mass spectrometer (ESI-MS) using a Micromass ZQ Single Quadrupole mass spectrometer.

General procedure for the synthesis of fully acetylated threonine-glycosylated oligopeptides

Glycosylated peptides were synthesized on a CS Bio CS136XT automated synthesizer using a standard Fmoc solid phase peptide synthesis (SPPS) method. Preloaded resin (H-L-Ser(t-Bu)-2-CI-trityl resin (0.133 g, 0.1 mmol)) was placed in a reaction vessel. Fmoc-protected amino acids (0.4 mmol), Fmoc-protected glycosylated amino acids (0.4 mmol), or palmitic acid (0.4 mmol) were dissolved in 5 mL of DMF and activated with HBTU (0.4 mmol) and DIPEA (0.6 mmol) in DMF to perform the coupling reaction. Fmoc deprotection was performed using a 20% piperidine solution in DMF. After peptide assembly, the resin was washed with DMF (3 x 5 mL), then with DCM (3 x 5 mL), and then dried under high vacuum for 3 hours. The dried resin was then treated with 10 mL of Reagent K (TFA:water:phenol:thioanisole = 8.5:0.5:0.5:0.5) for each 0.1 mmol of resin at room temperature for 3 hours. The crude glycosylated peptide was precipitated with cold ether, purified by RP-HPLC, and freeze-dried to yield 80-100 mg of glycosylated peptide as a white powder. The molecular weight of the glycosylated peptide was confirmed by MS, and the purity was established by analytical RP-HPLC.

General procedure for the synthesis of deacylated glycosylated oligopeptides

To a solution of Pal-KTT*KS-[OH] or Pal-KT*TKS-[OH] (0.05 mmol, respectively) in anhydrous MeOH (5 mL) was added dropwise sodium methoxide (1 M) in anhydrous methanol to adjust the pH to approximately 10. The reaction mixture was stirred at room temperature for 1 hour and then neutralized with acetic acid (pH ~4.0). The solvent was removed under vacuum. The crude deprotected glycosylated peptide was purified by RP-HPLC and freeze-dried to yield typically 30 to 40 mg of the target glycosylated peptide as a white powder.

General procedure for the synthesis of fully acetylated serine glycosylated oligopeptides

Synthesis of pre-loaded H-L-Ser(sugar)-2-Cl-trityl resin: To a solution of Fmoc-Ser(sugar)-OH (0.3 mmol) in anhydrous DCM (5 mL) was added 2-chlorotrityl resin (0.4 g, 0.3 mmol) in a centrifuge tube. After 15 minutes, DIPEA (87 μL, 0.5 mmol) was added. The mixture was shaken on a shaker for 30 minutes, and then DIPEA (120 μL, 0.75 mmol) was added. The mixture was allowed to remain on the shaker for another 3 hours. Methanol (0.4 mL) was added to the end cap, and the mixture was shaken for another 30 minutes. The resin was transferred to a glass funnel equipped with a porous disk and washed with DMF (2x5 mL), then with DCM (2x5 mL), and finally with MeOH (3x5 mL). The resin was dried under vacuum overnight to yield pre-loaded H-L-Ser(sugar)-2-Cl-trityl resin, which was further used in the Fmoc-SPPS synthesis of glycated peptides.

On CS Bio CS136XT automated synthesizer, the glycosylated peptide of serine glycosylation is synthesized by the Fmoc solid phase peptide synthesis method of standard.Preloaded resin (H-L-Ser (sugar)-2-Cl-trityl resin (0.1 mmol)) is placed in the reaction vessel.Through the activation using HBTU (0.4mmol) and DIPEA (0.6mmol), couple the amino acid (0.4mmol is dissolved among the 5mL DMF) of Fmoc protection.Similarly, couple N-terminated palmitic acid (0.4mmol is dissolved among the 5mL DMF).Use the 20% piperidine solution in DMF to remove the Fmoc protecting group.After peptide is assembled, use DMF (3x5mL) washing resin, use DCM (3x5mL) washing then, then dry 3 hours under high vacuum.In dry resin, add 10mL reagent K (TFA: water: phenol: thiophenylmethane=8.5:0.5:0.5:0.5). The mixture was then shaken at room temperature for 3 hours. The crude glycosylated peptide was precipitated from cold ether, purified by RP-HPLC, and freeze-dried to yield 120-150 mg of the target glycosylated peptide as a white powder. The molecular weight of the product glycosylated peptide was confirmed by MS, and the purity was assessed by analytical HPLC.

GP001-1: R ₁ =Ac ₄ -β-Glc, R ₂ =H, R ₃ =H

GP003-9: R ₁ =Ac ₇ -β-Mal, R ₂ =H, R ₃ =H

GP002-3: R ₁ =H, R ₂ =H, R ₃ =Ac ₄ -β-Glc

GP002-7: R ₁ =H, R ₂ =H, R ₃ =Ac ₄ -β-Gal

GP004-3: R ₁ =H, R ₂ =Ac7-β-Mal, R ₃ =H

GP004-4: R ₁ =H, R ₂ =H, R ₃ =Ac ₇ -β-Mal

Exemplary Synthesis of Pal-KT(Ac ₄ -p-Glc)TKS-[OH](GP001-1)

The glycosylated peptide GP001-1 was synthesized on a CS-Bio CS136XT automated synthesizer using standard Fmoc-SPPS methods and the following general procedure on HL-Ser(t-Bu)-2-Cl-trityl resin (133.3 mg, 0.1 mmol). The first amino acid (Fmoc-Lys(Boc)-OH (187.4 mg, 0.4 mmol)) was coupled to the preloaded resin using HBTU (128.4 mg, 0.4 mmol) and DIPEA (104.5 μL, 0.6 mmol) in DMF (5 mL). Subsequently, the Fmoc-amino acid and palmitic acid were coupled in the same manner. In the third coupling reaction, Fmoc-Thr(Ac ₄ -β-Glc)-OH (0.270 mg, 0.4 mmol) was used. After solid-phase synthesis, the glycosylated peptide was cleaved from the resin using Reagent K (10 mL). The crude glycosylated peptide was purified by preparative RP-HPLC to provide Pal-KT(Ac ₄ -β-Glc)TKS-OH, GP001-1 (70 mg, 62%) as a white powder. ESI-MS: m/z: Calcd for C ₅₃ H ₉₃ N ₇ O ₁₉ ; 1132.34; Found: 1133.1 [M+H] ⁺ . Purity determined by RP-HPLC was 95.6%.

Exemplary Synthesis of Pal-KT(p-Glc)-TKS-[OH](GP001-2)

Glycosylated peptide GP001-1 (65 mg, 0.05 mmol) was dissolved in 5 mL of anhydrous methanol and deprotected by adding sodium methoxide in methanol as described above, yielding 32 mg of the target _deacetylated glycosylated GP001-2 as a white powder. ESI-MS: m/z: Calculated for _{C₄₅H₈₅N₇₁₅O₁₅} ; _964.19 ; Found: 965 [M+H]⁺. Purity was determined to be 99 _% by analytical HPLC.

Exemplary Synthesis of Pal-KT(Ac ₇ -β-Mal)TKS-[OH](GP003-9)

On a CS-Bio CS136XT automated synthesizer, the glycosylated peptide GP003-9 was synthesized on HL-Ser(t-Bu)-2-Cl-trityl resin (133.3 mg, 0.1 mmol) using standard Fmoc-SPPS methods and the following general procedure. The first amino acid (Fmoc-Lys(Boc)-OH (187.4 mg, 0.4 mmol)) was coupled to the preloaded resin using HBTU (128.4 mg, 0.4 mmol) and DIPEA (104.5 μL, 0.6 mmol) in DMF (5 mL). Subsequently, the Fmoc-amino acid was coupled in the same manner. In the third coupling reaction, Fmoc-Thr(Ac ₇ -β-Mal)-OH (0.372 mg, 0.4 mmol) was used. After solid phase synthesis, the glycosylated peptide was removed from the resin using reagent K (10 mL). The crude glycosylated peptide was purified by preparative RP-HPLC to provide Pal-KT( _Ac₁₇ -β-Mal)TKS-OH, GP003-9 (90 mg, 63% yield) as _a white powder. ESI-MS: m/z: Calcd for _{C₆₅H₁₀N₁₀O₂₀ : 1420.59} ; Found: 1420.7 [M+H] ^⁺ . Analytical RP-HPLC determined the purity to be 99.1%.

Exemplary Synthesis of Pal-KTTKS(Ac ₄ -β-Glc)-[OH](GP002-3)

a) Fmoc-Ser(Ac4-β-Glc)-OH (0.3 mmol) was loaded onto 2-chlorotrityl resin (0.4 g, 0.3 mmol) by the general procedure described above. The dried pre-loaded resin was then used in subsequent steps without further treatment.

b) The glycosylated peptide GP002-3 was synthesized on a CS Bio CS136XT automated synthesizer using a standard Fmoc solid-phase peptide synthesis method on a preloaded resin prepared as described above. Following solid-phase synthesis, the glycosylated peptide was cleaved from the resin using Reagent K (10 mL). The resulting crude glycosylated peptide was purified by preparative HPLC to provide Pal-KTTKS(Ac4-β-Glc)-OH, GP002-3 (136 mg, 40%) _{as a} white powder. ESI-MS: m/z: Calcd for _C53H93N7O19 ; _1132.34 ; Found: 1132.5 [M+H]+. Purity was determined to be 99.6% by analytical RP-HPLC.

Exemplary Synthesis of PaKTTKS(Ac ₄ -β-Gal)-[OH](GP002-7)

a) Fmoc-Ser(Ac4-β-Gal)-OH (0.3 mmol) was loaded onto 2-chlorotrityl resin (0.4 g, 0.3 mmol) by the general procedure described above. In subsequent steps, the dried pre-loaded resin was used without further treatment.

b) The glycosylated peptide GP002-7 was synthesized on a CS Bio CS136XT automated synthesizer using a standard Fmoc solid-phase peptide synthesis method on a preloaded resin prepared as described above. Following solid-phase synthesis, the glycosylated peptide was cleaved from the resin using Reagent K (10 mL). The resulting crude glycosylated peptide was purified by preparative RP-HPLC to provide Pal-KTTKS(Ac4-β-Gal)-OH, GP002-7 (140 mg, 41%) as _{a white} powder. ESI-MS: m/z: Calcd for _C53H93N7O19 ; _1132.34 ; Found: 1132.5 [M+H]+. Purity was determined to be 99.6% by analytical HPLC.

Exemplary Synthesis of Pal-KTT(Ac ₇ -β-Mal)KS-[OH](GP004-3)

The glycosylated peptide GP004-3 was synthesized on a CS-Bio CS136XT automated synthesizer using a standard Fmoc-SPPS method and the following general procedure on HL-Ser(t-Bu)-2-Cl-trityl resin (133.3 mg, 0.1 mmol). The first amino acid (Fmoc-Lys(Boc)-OH (179.6 mg, 0.4 mmol)) was coupled to the preloaded resin using HBTU (126.3 mg, 0.4 mmol) and DIPEA (104.5 μL, 0.6 mmol) in DMF (5 mL). Subsequently, the Fmoc-amino acid was coupled in the same manner. In the second coupling reaction, Fmoc-Thr(Ac ₇ -β-Mal)-OH (0.383 mg, 0.4 mmol) was used. After solid phase synthesis, the glycosylated peptide was removed from the resin using reagent K (10 mL). The crude glycosylated peptide was purified by preparative RP-HPLC to provide Pal-KT( _Ac₁₇ -β-Mal)TKS-OH, GP004-3 (26 mg _, 18% yield) as _a white powder. ESI-MS: m/z: calcd for _{C₆₅H₁₆N₁₂O₂₂} ; _1420.59 ; found: 1420.7 [M+H] ^⁺ . Analytical RP-HPLC determined the purity to be 98.5%.

Exemplary Synthesis of Pal-KTTKS(Ac ₇ -βp-Mal)-[OH](GP004-4)

a) Fmoc-Ser(Ac7-β-Mal)-OH (0.3 mmol) was loaded onto 2-chlorotrityl resin (0.4 g, 0.3 mmol) by the general procedure described above. The dried pre-loaded resin was then used in subsequent steps without further treatment.

b) Glycosylated peptide GP004-4 was synthesized on a CS Bio CS136XT automated synthesizer using standard Fmoc solid-phase peptide synthesis on a preloaded resin prepared as described above. Following solid-phase synthesis, the glycosylated peptide was cleaved from the resin using Reagent K (10 mL). The resulting crude glycosylated peptide was purified by preparative HPLC to provide Pal-KTTKS(Ac7-β-Mal)-OH, GP004-4 (102 _mg , 24%) as _a white powder. ESI-MS: m/z: Calcd for _C65H109N7O27 ; _1420.59 ; Found: 1420.7 [M+H] ⁺ . Purity was determined to be 97.8% by analytical RP-HPLC.

abbreviation

Ac acetyl

Ac4-β-Glc 2,3,4,6-tetra-O-acetyl-β-D-glucopyranoside

Ac4-β-Gal 2,3,4,6-tetra-O-acetyl-β-D-galactopyranoside

Ac7-β-Mal 2,2',3,3',4',6,6'-hexa-O-β-D-maltopyranoside

Cel D-cellobiose

Cl

Gal D-Galactose

GalNAc N-acetyl-D-galactosamine

Glc D-glucose

GlcNAc N-acetyl-D-glucosamine

DCM dichloromethane

DIPEA N,N-Diisopropylethylamine

DMF dimethylformamide

Fuc L-Trehalose

HBTU N,N,N',N'-Tetramethyl-O-(1H-benzotriazol-1-yl)uronium hexafluorophosphate

Lac D-lactose

Mal D-Maltose

Man D-Mannose

MeOH methanol

Pal Palmitoyl

Rha L-rhamnose

t-Bu tert-butyl

TFA trifluoroacetic acid

Xyl D-Xylose

Mass spectrometry

By being injected into the mass spectrum of obtaining multiple peptides in LC-MS system (HPLC:WatersAlliance 2795) by the sample (2-5 μ L) of the multiple peptides being dissolved in water.Mobile phase is by water: formic acid (100: 0.1) and acetonitrile: formic acid (100: 0.1) forms, water: formic acid (100: 0.1) and acetonitrile: formic acid (100: 0.1) is respectively mobile phase A and B.Flow rate is set at 0.2mL/min, and consists of 50: 50 (mobile phase A: mobile phase B).Mass spectrum (Mass Spec:Waters Micromass ZQ) is set at within 2.0 minutes running times, under positive mode and negative mode, in 1 second, all by 100amu scan to 1800amu.Mass spectrum is extracted by TIC chromatogram.

ECM ELISA test

KMST-6 cells (Japan Health Sciences Foundation, JCRB0433) were seeded at 7,000 cells/ml in complete medium in 12-well tissue culture-treated plates (Falcon, 353043). Complete culture medium consisted of the following: MEM/EBSS (Hyclone, SH30024.01), 10% heat-inactivated FBS (Gibco, 10082-147), 1% L-glutamine (Hyclone, SH30031.01), 1% nucleosides (800 mg/L adenosine, 850 mg/L guanidine, 730 mg/L cytidine, 730 mg/L uridine, and 240 mg/L thymidine), 1% penicillin/streptomycin (HyClone, SV30010), 1% sodium pyruvate (HyClone, SH30239.01), 1% non-essential amino acids (Hyclone, SH30238.01), and 0.04% gentamicin (Gibco, 15710-064). Cells were grown for 2 days before treatment. Aspirate culture medium and use 1X PBS (Fisher, SH30256.01) to wash cells, then use 5 μM (1mL/ well) glycosylated peptide (reference (Pal-KTTKS, CPCScientific, 822197) and control (DMSO, Fisher, BP231-100)) in serum-free medium to process, wherein said serum-free medium is supplemented with 50 μg/mL sodium ascorbate (Sigma, A4034) and 80 μg/mL β-alanine fumarate (Sigma, A3134). The cells processed are incubated at 37 DEG C and 5% CO ₂ for 48 and 72 hours. At each time point, the culture medium obtained by each treatment is collected, and centrifuged at 1500RPM for 15 minutes at 4 DEG C. Supernatant is collected, and stored at-80 DEG C for the quantification of extracellular matrix protein. The following kits are used for the quantification of ECM proteins: fibronectin (Takara, MK115) and elastin (Uscn Life Science Inc., E91337Hu). Cells were harvested and lysed using 250 μL lysis buffer ((20 mM Tris-HCl, Sigma, T1503), 137 mM NaCl (Sigma, S9888), 2 mM EDTA (Fisher, BP118), 1% IGEPAL (Sigma, I8896), 10% glycerol (Sigma, G5516), 100 μM PMSF (Fisher, 36978) and 10 μg/mL leupeptin (Sigma-Aldrich, L2884)). They were placed in an ice container on a flatbed shaker (Lab-Line Instruments, 3595) for 30 minutes. The lysate was centrifuged at 10000 g for 5 minutes at 4 ° C. The supernatant was collected and used for protein quantification (BioRad, 500-0112).

Cell morphology

KMST-6 cells (Japan Health Sciences Foundation, JCRB0433) were seeded at 7,000 cells/ml in complete medium in 12-well tissue culture-treated plates (Falcon, 353043). Complete culture medium consisted of the following: MEM/EBSS (Hyclone, SH30024.01), 10% heat-inactivated FBS (Gibco, 10082-147), 1% L-glutamine (Hyclone, SH30031.01), 1% nucleosides (800 mg/L adenosine, 850 mg/L guanidine, 730 mg/L cytidine, 730 mg/L uridine, and 240 mg/L thymidine), 1% penicillin/streptomycin (HyClone, SV30010), 1% sodium pyruvate (HyClone, SH30239.01), 1% non-essential amino acids (Hyclone, SH30238.01), and 0.04% gentamicin (Gibco, 15710-064). Cells were grown for 2 days before treatment. The culture medium was aspirated and the cells were washed with 1X PBS (Fisher, SH30256.01) and then treated with 5 μM glycosylated peptides (GP001-1, GP002-3, GP002-7, GP003-9, GP004-3 and GP004-4) in serum-free medium supplemented with 50 μg/mL sodium ascorbate (Sigma, A4034) and 80 μg/mL β-alanine fumarate (Sigma, A3134). The treated cells were incubated at 37°C and 5% _CO2 for 48 and 72 hours. At each time point, images were acquired using an Olympus Opti-Tech Scientific Microscope (Lumenera, Infinity 2 camera, 10x magnification), and changes in cell size and morphology were recorded.

Cell viability MTT test

KMST-6 cells (Japan Health Sciences Foundation, JCRB0433) were seeded at 7,000 cells/ml in complete culture medium. Complete culture medium consisted of the following: MEM/EBSS (Hyclone, SH30024.01), 10% heat-inactivated FBS (Gibco, 10082-147), 1% L-glutamine (Hyclone, SH30031.01), 1% nucleosides (800 mg/L adenosine, 850 mg/L guanidine, 730 mg/L cytidine, 730 mg/L uridine, and 240 mg/L thymidine), 1% penicillin/streptomycin (HyClone, SV30010), 1% sodium pyruvate (HyClone, SH30239.01), 1% non-essential amino acids (Hyclone, SH30238.01), and 0.04% gentamicin (Gibco, 15710-064). Cells were grown for 2 days before treatment. Aspirate culture medium and use 1X PBS (Fisher, SH30256.01) to wash cells, then use 5 μM glycosylated peptides (GP001-1, GP002-3, GP002-7, GP003-9, GP004-3 and GP004-4) in serum-free culture medium (reference (Pal-KTTKS, CPCScientific, 822197) and control (DMSO, Fisher, BP231-100)) to process, wherein said serum-free culture medium is supplemented with 50 μg/mL sodium ascorbate (Sigma, A4034) and 80 μg/mL β-alanine fumarate (Sigma, A3134).By processed cells at 37 DEG C and 5% CO ₂ lower incubation 48 and 72 hours.At each time point, by adding 15 μL dye solution (Promega, G4102) to all wells, implement MTT test. The cells were incubated for 2-4 hours. During this process, the living cells converted the MTT tetrazolium component of the dye solution into a formazan product. A dissolution/stop solution (Promega, G4101) was then added to the culture wells to dissolve the formazan product. The absorbance was measured at 570 nm using a microplate reader (Molecular Devices, SpectraMax M2e). We defined cell viability as less than 50% toxicity compared to the control.

Total collagen test

KMST-6 cells (Japan Health Sciences Foundation, JCRB0433) were seeded at 7,000 cells/ml in complete medium on 60 mm tissue culture treated plates (Falcon, 353002). Complete culture medium consisted of the following: MEM/EBSS (Hyclone, SH30024.01), 10% heat-inactivated FBS (Gibco, 10082-147), 1% L-glutamine (Hyclone, SH30031.01), 1% nucleosides (800 mg/L adenosine, 850 mg/L guanidine, 730 mg/L cytidine, 730 mg/L uridine, and 240 mg/L thymidine), 1% penicillin/streptomycin (HyClone, SV30010), 1% sodium pyruvate (HyClone, SH30239.01), 1% non-essential amino acids (Hyclone, SH30238.01), and 0.04% gentamicin (Gibco, 15710-064). Cells were grown for 2 days before treatment. Cells were washed with 1X PBS (Fisher, SH30256.01) and treated with 5 μM glycosylated peptides (GP003-9, GP004-3, GP004-4) (with reference to (Pal-KTTKS, CPCScientific, 822197) and control (DMSO, Fisher, BP231-100)) in serum-free medium supplemented with 50 μg/mL sodium ascorbate (Sigma, A4034) and 80 μg/mL β-alanine fumarate (Sigma, A3134). The cells were incubated at 37°C and 5% CO ₂ for 48 and 72 hours. The culture medium obtained by each treatment was collected at each time point and centrifuged at 1500RPM for 15 minutes at 4°C. Supernatant was collected and stored at -80°C for quantification of extracellular matrix protein. The following kit was used for the quantification of total collagen: Sirius Red Collagen Detection Kit (Chondrex Inc., 9062). Cells were harvested and lysed using lysis buffer ((20mM Tris-HCl, Sigma, T1503), 137mM NaCl (Sigma, S9888), 2mM EDTA (Fisher, BP118), 1% IGEPAL (Sigma, I8896), 10% glycerol (Sigma, G5516), 100μM PMSF (Fisher, 36978) and 10μg/mL leupeptin (Sigma-Aldrich, L2884)). They were placed in an ice container on a regular shaker (VWR, 89032-088) for 30 minutes. The lysate was centrifuged at 10000g for 5 minutes at 4°C. The supernatant was collected and used for protein quantification (BioRad, 500-0112).

3D human in vitro skin model

MatTek's EpiDermFT ^™ 3D human in vitro skin model is produced by MatTek Corporation (Ashland, MA, USA). The 3D skin model is composed of normal human epidermal keratinocytes (NHEK) and normal human dermal fibroblasts (NHDF), which are derived from neonatal foreskin tissue and adult skin, respectively. The history of the tissue indicates 8-12 cell layers plus the stratum corneum, including the basal layer, the spinous layer, and the granular layer. The tissue is grown at the air-liquid interface in Costar Snapwell ^™ single-well tissue culture plate inserts, wherein the inserts have a 0.4 μm pore size, a 1.2 cm diameter, and a 1.0 ^cm2 surface area. For tissue maintenance, serum-free EFT-400-ASY culture medium is used, which contains Dulbecco's Modified Eagle's Medium (DMEM), epidermal growth factor, insulin, hydrocortisone, 5 μg/mL gentamicin, 0.2 μg/ml amphotericin B, and phenol red.

Elastin assessment

The tissue was rinsed twice with 1% PBS and treated with 5 μM GP003-9 (reference (Pal-KTTKS, CPC Scientific, 822197) and control (DMSO, Fisher, BP231-100)). 72 hours after treatment, the supernatant was isolated and stored at -80°C for elastin ELISA analysis (Cedarlane, SE9133Hu). ELISA was performed using a 1:50 sample dilution according to the supplier's protocol.

For total protein quantification, tissues were rinsed with 1% PBS and cold Tissue Protein Extraction Reagent (Pierce, 78510) was added. Tissues were homogenized using a handheld homogenizer for approximately 1 minute. All samples were centrifuged at 16,000 g for 10 minutes at 4°C. After centrifugation, the supernatant was collected and the protein concentration was determined using a BCA Protein Assay Kit (Pierce, 23227).

Histological processing

The tissue was rinsed twice with 1% PBS and treated with 5 μM GP003-9 (reference (Pal-KTTKS, CPC Scientific, 822197) and control (DMSO, Fisher, BP231-100)). 72 hours after treatment, the tissue was fixed overnight in 10% neutral buffered formalin and transferred to 1% PBS the next day. The tissue was then bisected (providing cross sections), dehydrated in a series of graded ethanol, and embedded in paraffin. 5-micron sections were prepared and stained with hematoxylin and eosin for histological observation.

Statistical analysis

One-way ANOVA statistical analysis was used to compare the mean values of three or more samples. The Bonferroni multiple comparison test allowed comparison of differences between glycosylated peptide treatments and controls or references. Statistical significance was derived by assigning a p value (p < 0.05). (Data analysis was performed using GraphPad Prism 6, a scientific software that enables basic statistical testing, data organization, and scientific graphing).

The following examples describe some exemplary modes of preparing or practicing certain compositions and methods described herein. It should be understood that these examples are for illustrative purposes only and are not intended to limit the scope of the compositions and methods described herein.

Example 1

As described above, four batches of modified peptides were prepared and chemical identity confirmed by HPLC and mass spectrometry. Overall, 44 different glycosylated peptides were prepared and are shown in Tables 1A and 1B.

Table 1A. Synthetic glycosylated peptides

Table 1B. Synthetic glycosylated peptides

In order to measure the effect of 5 μM independent glycosylated peptide on the secretion of extracellular matrix protein, described glycosylated peptide is incubated with human skin fibroblast (KMST-6).Control sample cell is processed using DMSO (Fisher, BP231-100).As mentioned above, supernatant (2 time points, 48 hours and 72 hours processing) is collected by the cell of processing, and is stored at-80 DEG C, so that the quantification of extracellular matrix protein is subsequently carried out using ELISA kit (fibronectin (Takara, MK115), elastin (Uscn Life Science Inc., E91337Hu).In order to carry out the mensuration of total cell protein, the cell of processing is harvested and cracked.The lysate is centrifuged, supernatant is collected, and is then used for the quantification (BioRad, 500-0112) of protein.At 450nm absorbance, ELISA plate is read on plate reader (Molecular Devices), and SoftMax Pro software is used to calculate the μ g/ml value of the protein of being paid close attention to that is derived from the 4 parameter fitting standard curve generated. The μ g/ml of various secreted proteins is normalized to the total protein in every hole. By taking the average value of the normalized values of at least 3 independent experiments, and presenting them as the % of the control treatment set to 100%, thereby analyzing the data (Table 2-5). Single-factor ANOVA (p < 0.05) is used to carry out the analysis of statistical significance, wherein " * " represents the statistical significance when compared with the control, and " # " represents the statistical significance when compared with the reference.

The following table shows the effect of all 44 glycosylated peptides on the secretion of elastin (Table 2 and Table 3 at 48 hours and 72 hours, respectively) and the secretion of fibronectin (Table 4 and Table 5 at 48 hours and 72 hours, respectively) after treatment of human skin fibroblasts. Graphs depicting these results are shown in Figures 1A and 1B (secretion of elastin at 48 hours and 72 hours, respectively), and Figures 2A and 2B (secretion of fibronectin at 48 hours and 72 hours, respectively). From these results, it can be seen that fibroblasts treated with glycosylated peptides exhibit increased ECM secretion (elastin and fibronectin) over control-treated fibroblasts.

Table 2. Elastin secretion at 48 hours

Table 3. Elastin secretion at 72 hours

Table 4. Fibronectin secretion at 48 hours

Table 5. Fibronectin secretion at 72 hours

Through this preliminary screening, six glycosylated peptides were identified for further development: GP001-1 (Pal-KT(Ac ₄ -Glc)TKS-OH), GP002-3 ((Pal-KTTKS(Ac 4 -Glc)-OH), GP002-7 (Pal-KTTKS(Ac ₄ -Gal)-OH), GP003-9 (Pal-KT(Ac _{7 -β} -Mal)TKS-OH), GP004-3 (Pal-KTT(Ac ₇ -β-Mal)KS-OH) and GP004-4 (Pal-KTTKS(Ac ₇ -β-Mal)-OH).

GP001-1 (Pal-KT(Ac ₄ -β-Glc)TKS-OH) - Material: Glycosylated peptide (β-Glc peptide PerAc); Molecular weight: 1132; Appearance: White freeze-dried solid; Purity (HPLC) >95% ( FIG. 3A ); Mass spectrum (ESI POS): consistent with [M+H] ⁺ =1132.5 and [M+2H] ⁺ =567 ( FIG. 3B ).

GP002-3 (Pal-KTTKS(Ac ₄ -β-Glc)-OH) - Material: glycosylated peptide (β-Glc peptide PerAc); molecular weight: 1132; appearance: white freeze-dried solid; purity (HPLC) >99.6% ( FIG4A ); mass spectrum (ESI POS): consistent with [M+H]=1132.5 ( FIG4B ).

GP002-7 (Pal-KTTKS (Ac ₄ -β-Gal) -OH) - Material: Glycosylated peptide (β-Glc peptide PerAc); Molecular weight: 1132; Appearance: White freeze-dried solid; Purity (HPLC) > 100% ( FIG5A ); Mass spectrum (ESI POS): consistent with [M+H] = 1132.4 ( FIG5B ).

GP003-9 (Pal-KT (Ac ₇ -β-Mal) TKS-OH) - Material: glycosylated peptide (β-maltodextrin PerAc); molecular weight: 1420; appearance: white freeze-dried solid; purity (HPLC) > 97% ( FIG. 6A ); mass spectrum (ESI POS): consistent with [M+H] = 1420.7 ( FIG. 6B ).

GP004-3 (Pal-KTT (Ac ₇ -β-Mal) KS-OH) - Material: glycosylated peptide (β-maltodextrin PerAc); molecular weight: 1420; appearance: white freeze-dried solid; purity (HPLC) > 95% ( FIG. 7A ); mass spectrum (ESI POS): consistent with [M+H] = 1420.8 ( FIG. 7B ).

GP004-4 (Pal-KTTKS (Ac ₇ -β-Mal) -OH) - Materials: Glycosylated peptide (β-maltodextrin PerAc); Molecular weight: 1420; Appearance: White freeze-dried solid; Purity (HPLC) > 95% ( FIG8A ); Mass spectrum (ESI POS): consistent with [M+H] = 1420.7 ( FIG8B ).

These six glycosylated pentapeptides were subjected to in vitro toxicity tests (cell viability and morphology) compared with the reference pentapeptide Pal-KKTKS.

To determine the morphology and size of the cells, 5 μM of the six exemplary glycosylated peptides were applied to human skin fibroblasts (KMST-6) as described previously. The reference pentapeptide was Pal-KTTKS (CPC Scientific, 822197), and the control sample cells were treated with DMSO (Fisher, BP231-100). Morphological images showed that the reference visibly interfered with cell viability and demonstrated that cell confluence was reduced compared to cells treated with the control and glycosylated peptides. Although none of the six glycosylated peptides tested showed any form of toxicity by comparison, this could be observed by phase contrast microscopy (Figure 9). Importantly, 90% of all glycosylated peptides tested at a concentration of 5 μM did not show any form of toxicity, which could be measured by MTT assay (data not shown).

To determine the effect on cell viability (as a measure of toxicity), 5 μM of 6 exemplary glycosylated peptides were applied to human skin fibroblasts (KMST-6) as described previously. The reference pentapeptide was Pal-KTTKS (CPC Scientific, 822197), and control sample cells were treated with DMSO (Fisher, BP231-100). As described above, the treated cells were incubated at 37°C and 5% _CO2 for 48 hours (Figure 10A) and 72 hours (Figure 10B). At each time point, the MTT test was performed by adding 15 μL of dye solution (Promega, G4102) to all wells. The cells were incubated for 2-4 hours, during which the living cells converted the MTT tetrazolium component of the dye solution into a formazan product. Dissolution/stop solution (Promega, G4101) was then added to the culture wells to dissolve the formazan product. Absorbance was measured at 570 nm using a microplate reader (Molecular Devices, SpectraMaxM2e). Three or more independent experiments were performed and statistical significance was determined using one-way ANOVA (p<0.05). Toxicity was defined as less than 50% cell viability compared to the control. Of the six glycosylated peptides tested, only one showed an effect on cell viability at any time point (Figures 10A and 10B). In contrast, the non-glycosylated reference pentapeptide showed a decrease in cell viability compared to the control at 48 and 72 hours (Figure 10B). Importantly, 90% of all glycosylated peptides tested at a concentration of 5 μM did not show any form of toxicity, which can be measured by MTT assay (data not shown).

To further standardize and investigate the toxicity profile of glycated peptides, additional experiments were performed to examine glycated peptides GP003-9, GP004-3, and GP004-4 (data not shown). Human skin fibroblasts were treated with 5 μM glycated peptides, a reference (Pal-KTTKS, CPC Scientific, 822197), and a control (DMSO, Fisher, BP231-100) at 48 and 72 hours. The following toxicity assays were performed: 1) cell viability (CellTiterAQueous One Solution (MTS) colorimetric assay, Promega, G3580) for measurement of cell metabolic activity; 2) Viability/Cytotoxicity Kit (Molecular Probes, L-3224) for measurement of viable cells in each total cell population; 3) cell proliferation was measured by measuring the following: a) cell count to obtain the total number of viable cells, and b) DNA content (GenomicDNA Mini Kit, Invitrogen, K182002) for quantification of cells with intact DNA; and finally, 4) apoptosis/necrosis (Dead Cell Apoptosis Kit with Annexin V Alexa488 & Propidium Iodide, Molecular Probes, V13241) for measurement of cells undergoing cell death. In all other experiments performed, toxicity was not observed for the three glycosylated peptides, which were subjected to other in vitro toxicity tests.

To demonstrate the effect on extracellular matrix protein secretion, 5 μM glycosylated peptides [GP001-1 (Pal-KT (Ac ₄ -Glc) TKS-OH), GP002-3 ((Pal-KTTKS (Ac ₄ -Glc) -OH), GP002-7 (Pal-KTTKS (Ac ₄ -Gal) -OH), GP003-9 (Pal-KT (Ac ₇ -β-Mal) TKS-OH), GP004-3 (Pal-KTT (Ac ₇ -β-Mal) KS-OH) and GP004-4 (Pal-KTTKS (Ac ₇ -β-Mal) -OH)] were used as reference (Pal-KTTKS, CPC) Human skin fibroblasts (KMST-6) were treated with DMSO (Fisher Scientific, 822197) or blank control (DMSO, Fisher, BP231-100) for 72 hours. Three or more independent experiments were performed. Statistical significance was analyzed using one-way ANOVA (p<0.05), where "*" indicates statistical significance when compared to the control, and "#" indicates statistical significance when compared to the reference.

Table 6. Summary of ECM proteins

Compound# Elastin Fibronectin Total collagen comparison 100% 100% 100% Reference 133 118 110 GP001-1 188 122 GP002-3 GP002-7 116 GP003-9 GP004-3 GP004-4 186

Compared with the control, GP001-1, GP002-3, GP002-7, GP003-9, GP004-3 and GP004-4 induced higher biological activities in the secretion of elastin (Table 6 and FIG. 11A ) and fibronectin (Table 6 and FIG. 11B ). However, compared with the reference peptide, glycated peptides GP002-3 (189%, p<0.0001), GP003-9 (151%, p<0.0001) and GP004-3 (252%, p<0.0001) significantly increased elastin secretion, while compared with the control, GP002-3 (222%, p<0.0001), GP002-7 (107%, p<0.04), GP003-9 (184%, p<0.0001) and GP004-3 (285%, p<0.0001) significantly increased elastin secretion. In addition, GP004-3 (50%, p<0.02) and GP004-4 (65%, p<0.0007) significantly increased the secretion of fibronectin compared with the control, while GP002-3 (35%, p<0.01), GP003-9 (40%, p<0.0004), GP004-3 (68%, p<0.0003) and GP004-4 (83%, p<0.0001) significantly increased the secretion of fibronectin compared with the control. Finally, GP003-9, GP004-3 and GP004-4 were the only glycated peptides tested for total collagen, and the results showed that they all significantly increased the secretion of total collagen compared with the control, by 25% (p<0.004), 31% (p<0.0008) and 24% (p<0.005), respectively, and GP004-3 (21%, p<0.01) significantly increased the secretion of total collagen compared with the reference peptide (Table 6 and Figure 12).

MatTek's EpiDermFT ^™ , a 3D human skin model composed of normal human epidermal keratinocytes and normal human dermal fibroblasts, was used to further investigate the effects of an exemplary glycosylated peptide (GP003-9) on human skin. As described above, tissues were treated with 5 μM GP003-9, a reference (Pal-KTTKS, CPC Scientific, 822197), or a blank control (DMSO, Fisher, BP231-100). Seventy-two hours after treatment, the supernatant was isolated and analyzed for elastin secretion. Figure 13 shows that GP003-9 increased elastin secretion in the 3D human skin model by approximately 1.5-fold compared to the non-glycosylated reference peptide or control. Alternatively, tissues treated with 5 μM GP003-9, a reference (Pal-KTTKS, CPC Scientific, 822197), or a plate control (DMSO, Fisher, BP231-100) were paraffin-embedded, sectioned, and then stained with hematoxylin and eosin for histological evaluation. Figure 14 shows that GP003-9 had no adverse effects on the histology of the 3D human skin model.

The above embodiments are intended to be exemplary only. Those skilled in the art may effect alterations, modifications, and variations to the particular embodiments without departing from the scope thereof, which is defined solely by the claims appended hereto.

Claims (26)

1. A glycosylated oligopeptide consisting of 5 or 6 amino acids comprising the following sequence: palmitoyl-X1-Lys-X2-X3-Lys-X4-OH, wherein: X1 is: Ser, which does not exist, or Asn, which has a glycosylated side chain; X2 is Thr, Thr with glycosylated side chain, Asn with glycosylated side chain, or Ser with glycosylated side chain; X3 is Thr, or Thr with glycosylated side chains; X4 is Ser, or Ser with glycosylated side chains. Wherein: at least one of X1, X2, X3 and X4 is an amino acid with a glycosylated side chain; In this context, each glycosylated side chain is independently glycosylated from the following carbohydrates: glucose, N-acetyl-glucosamine, galactose, N-acetyl-galactosamine, mannose, maltose, lactose, rhamnose, cellobiose, xylose, and trehalose; and Wherein: each hydroxyl group on the carbohydrate is independently OH or acetylated. 2. The glycosylated oligopeptide according to claim 1, wherein the hydroxyl groups on the carbohydrates are all acetylated. 3. The glycosylated oligopeptide according to claim 1, wherein the hydroxyl groups on the carbohydrates are all non-acetylated. 4. The glycosylated oligopeptide according to any one of claims 1-3, wherein X1 is absent. 5. The glycosylated oligopeptide according to any one of claims 1-3, wherein X2 is Thr having a glycosylated side chain. 6. The glycosylated oligopeptide according to claim 5, wherein the glycosylated side chain is glycosylated with glucose, mannose or maltose. 7. The glycosylated oligopeptide according to claim 5, wherein the X1, X3 and X4 amino acids are non-glycosylated. 8. The glycosylated oligopeptide according to any one of claims 1-3, wherein X4 is Ser having a glycosylated side chain. 9. The glycosylated oligopeptide of claim 8, wherein the glycosylated side chain is glycosylated with glucose, galactose or maltose. 10. The glycosylated oligopeptide according to claim 8, wherein the X1, X2 and X3 amino acids are non-glycosylated. 11. The glycosylated oligopeptide according to any one of claims 1-3, wherein X3 is Thr having a glycosylated side chain. 12. The glycosylated oligopeptide according to claim 11, wherein the X1, X2 and X4 amino acids are non-glycosylated. 13. A glycosylated oligopeptide consisting of the following sequence: palmitoyl-Lys-Thr*-Thr-Lys-Ser-OH, wherein Thr* is a threonine glycosylated by a fully acetylated β-glucose. 14. A glycosylated oligopeptide consisting of the following sequence: palmitoyl-Lys-Thr*-Thr-Lys-Ser-OH, wherein Thr* is a threonine glycosylated by peracetylation of α-mannose. 15. A glycosylated oligopeptide consisting of the following sequence: palmitoyl-Lys-Thr*-Thr-Lys-Ser-OH, wherein Thr* is a threonine glycosylated by peracetylation of β-maltose. 16. A glycosylated oligopeptide consisting of the following sequence: palmitoyl-Lys-Thr-Thr-Lys-Ser*-OH, wherein Ser* is a serine glycosylated by peracetylated β-galactose. 17. A glycosylated oligopeptide consisting of the following sequence: palmitoyl-Lys-Thr-Thr-Lys-Ser*-OH, wherein Ser* is a serine glycosylated by a fully acetylated β-glucose. 18. A glycosylated oligopeptide consisting of the following sequence: palmitoyl-Lys-Thr-Thr*-Lys-Ser-OH, wherein Thr* is a threonine glycosylated by peracetylation of β-maltose. 19. A glycosylated oligopeptide consisting of the following sequence: palmitoyl-Lys-Thr-Thr-Lys-Ser*-OH, wherein Ser* is a serine glycosylated by peracetylation of β-maltose. 20. A glycosylated oligopeptide comprising the sequence of any one of SEQ ID NO:12-48. 21. A cosmetic or dermatological composition comprising at least one glycosylated oligopeptide according to any one of claims 1-20. 22. Use of the composition of claim 21 in the preparation of a pharmaceutical agent for increasing the secretion of extracellular matrix proteins by cells in an individual. 23. The use according to claim 22, wherein the extracellular matrix protein is elastin, fibronectin, or collagen. 24. Use of the glycosylated oligopeptide according to any one of claims 1-20 in the preparation of a pharmaceutical agent for increasing the secretion of extracellular matrix proteins by cells. 25. The use according to claim 24, wherein the extracellular matrix protein is elastin, fibronectin, or collagen. 26. A skin care composition comprising: a) A safe and effective amount of the glycosylated oligopeptide according to any one of claims 1-20; b) A safe and effective amount of at least one other skin care active substance selected from: exfoliating active substances, acne-fighting active substances, acne medications, vitamin A compounds, vitamin _B3 compounds, vitamin C compounds, vitamin E compounds, carotene-like substances, vitamin A-like substances, di-, tri-, tetra-, penta- and hexapeptides and their derivatives, hydroxy acids, free radical scavengers, chelating agents, anti-inflammatory agents, local anesthetics, tanning active substances, skin whitening agents, sunscreens, slimming agents, flavonoids, antimicrobial active substances, skin healing agents, antifungal active substances, farnesol, phytosterols, allantoin, glucosamine, hyaluronic acid and mixtures thereof; and c) Dermatologically acceptable carriers.