FIELD OF THE INVENTION

The present invention relates to a wound healing agent possessing cell migration characteristics comprising an effective amount of at least one of the actives of Formula 1, and particularly, relates to a cosmetic or pharmaceutical composition, especially dermopharmaceutical composition comprising the said actives in association with a cosmetically/ dermopharmceutically acceptable vehicle with or without other skin benefiting agents.

H-R-R'-H

Wherein R-R'= Saturated hydrocarbons having 14-20 carbon atoms

Formula 1

More particularly, the invention relates to a simple and cost effective dermopharmaceutical/ cosmetic compositions for external application facilitating cell migration and therefore wound healing comprising at least anyone or more of eicosane of Formula 2 and hexadecane of Formula 3 as wound healing agent preferably sourced from a safe and renewable plant source, Syzygium cuminii Skeels (syn. Eugenia jambolana Lam)., which can be used singly or in combination with cosmetically/ dermopharmaceutically acceptable vehicle/ carriers.

CH3-(CH2)i8-CH3
Formula 2 (Eicosane)

Cn3_(CH2ji4 "CH3
Formula 3 (Hexadecane)

BACKGROUND OF THE INVENTION

Skin is the major structural and functional organ protecting the body from external environment, injuries or insults. Any damage to the skin (wound or injury) should be rapidly and efficiently repaired by the healing process.

Cell migration is a fundamental activity intrinsic to development and maintenance of homeostasis in processes such as wound healing, neo-vascularization and the workings of the immune system. When the cells fail to migrate or there is
substantial movement of invasive cells into inappropriate locations, which is the core reason for many disease processes such as dysregulated wound healing and cancer. In addition to the above, understanding cell migration is critical to emerging technologies such as tissue generation and metastasis of transformed cells.

Wound healing is a well orchestrated process comprising of three major overlapping phases: inflammatory phase, proliferative phase and maturational or remodeling phase (Diegelmann et al., Front Biosci., 2004, 9, 283-89). Usually the skin gets repaired and heals uneventfully. But many of the wounds like: ulcers, burns, traumatic wounds, non-healing ulcers, diabetic ulcers, pressure ulcers, ischemic ulcers and cancer extirpations fail to heal naturally. Some times even the chronic ulcers are also associated with mortality. More over the prevalence of chronic and non healing wounds increases with age and other patho-physiological conditions of the patients like diabetes, AIDS, etc.

During the first phase of the healing, clotting cascades both the intrinsic and extrinsic pathways that initiates the inflammatory phase. The platelets release an array of cytokines like Etablissement Francais des Greffes (EFG), firbonectin, fibrinogen, platelet derived growth factor (PDGF) etc. (Broughton et.al., Plast Reconstr Surg. 2006, 117(Suppl), le-32e). Neutrophils and macrophages gets attracted to the wound site and debride the wound area by releasing various proteloytic enzymes (Sugihara et. al., Proc Soc Exp Biol Med. 2000, 225, 58-64) that activate the resident skin cells like fibroblasts and keratinocytes.

The second phase of the wound healing, the proliferative phase is crucial, during which only various cellular activities responsible for the closure of the wound takes place. Cell migration, cell proliferation, matrix deposition and re-epithelialization are the key events occurring in the phase.

Cell migration is an important event in various physiological and pathological processes. During the healing process the cells from the periphery of the wound migrate into the wound area to fill the gap, followed by proliferation and matrix production. Hence this event plays a pivotal role in accelerating the wound healing of the skin.

Millions of dollars have been spent in developing various recombinant growth factors, skin grafts, organotypic skin replacements for wound over the past decades, which are far from the reach of the common man.

Number of compositions facilitating wound repair like collagen implants (Ksander et. al., USP: 4,950,483, 1990), collagen sheets (Zimmerman et. al., USP: 4,453,939, 1984), collagen with fibrinogen component, with thrombin component and with fibronectin (Silver et al., USP: 4,970,298, 1990), hyaluronic acid (Broughton et.al., Plast Reconstr Surg. 2006, 117(Suppl), le-32e; Nakamura et. al., Exp Eye Res., 1997, 64, 355-359), chondroitin sulphate proteoglycan mediated cell migration of fibroblasts (Svee et al., Clin Invest., 1996, 98, 1713-27) are known in the art.
Topical gel and cream formulations comprising human plasma fibronectin for healing of cutaneous wounds, a composition of EFG, fibronectin, a synthetic collagenase inhibitor, aprotinin etc, gelatin binding domains of fibronectins stimulating fibroblast migration (Ellis et. al., Exp Cell Res., 1996, 228, 326-33) are also known. In 1995, Steed et al., reported on the RFDS-containing cell binding domain of fibronectin stimulating fibroblast migration in accelerated wound healing Argidene Gel or Telio-Derm Gel with RGD peptide matrix are available commercially to ease and accelerate wound healing (Steed et al., Diabetes Care, 1995, 18, 39-46).

Topical applications of honey has been used in the treatment of burns and wounds for many centuries, with documents describing this use dating back to 1700 BC. A number of properties inherent to honey might contribute to its ability to fight infection and promote healing. Its high sugar content allows it to draw infection and fluid from wounds by a process called "osmosis." Honey prevents bacterial growth through its acidic pH and through the work of an enzyme that produces small amounts of hydrogen peroxide. Its ability to keep the area around a wound moist and protected promotes fast healing and prevents scarring (http://bastyrcenter.org/content/view/810/).

Several people uses petroleum jelly to accelerate wound healing process. The petroleum jelly acting as sealing on cuts and burns, which inhibits germs from getting into the wound and keeps the injured area supple by preventing the skin's moisture from evaporating. (http://en.wikipedia.org/wiki/Petroleum_jelly).

Inspite of the above mentioned medications being available, the severity of the problem especially with respect to non healing wounds or ulcers irrespective of the responsible pathophysiological conditions draws continuous attention in bringing a more effective ways of easing the wound healing process by addressing different aspects of the wound healing process.

Lack of selective herbal extracts is the bane of the herbal personal or health care industry. Herbal extracts are generally mixtures of several compounds from the plant/s. Even when such extracts are obtained from single plant, there remains scanty information on the nature of the active ingredient (s) and on the effective amount of the active ingredient required in the extract for the product to be effective. Plant ingredients are known to vary depending on the strain of the plant used, the nature of the soil in which the plant grows, the age the plant, the time of the harvest and related factors. Several compounds/ mixtures have been reported in literature for wound healing property.

Leon et al., studied bioassay guided investigation of the ethanol extract of Peperomia galioides for wound healing property and identified (+)-epi-a-bisbolol as an active compound by in-vivo method wherein both oc-bisbolol and a-terpineol were reported to have wound healing properties (Leon et. al., J. Nat. Prod., 2001, 64, 1357-59). Novel hybrid compounds derived from solanesol have been reported in possessing wound healing property (Srivastava et al., Indian J. Chem., 2009, 48B, 237-247). Morganti taught that chitin nanofibrils are able to activate fibroblast proliferation and cytokine production, favouring gaint cell migration, macrophage activation and neovascularization. A component of Aloe vera, acemanan, has been reported to accelerate wound healing and reduce pain in aphthous stomatitis and prevent stress induced gastric ulceration in rats. The mannan derivatives stimulate collagen synthesis and fibroblast activity (Ghosh et. al., Clinical Science, 2004, 104, 547-556). Ebba et. al., disclosed, resveratrol, a natural polyphenols compound available in many plants to inhibit endothelial cell growth and act as an oral angiogenesis inhibitor (The FASEB Journal., 2001, 15, 1798-1800). The anti-angiogenic and wound healing compound, trans-4', 5'-dihydroxy-3-methoxystilbene-5-0-[a-L-rhamnopyronosyl-(1^2)-[a-L-rhamonopyranosyl-(1^6)-(3-D-glucopyranoside was isolated from medicinal plant, Boswellia papyriferai (Hussain et. al., BMC Cell

Biology, 2009, 10, 30). Okoli et.al., teaches that the aerial parts of Phyllanthus niruri showed wound healing and anti-ulcer properties (Okoli et. al., Am. J. Pham. & Toxicol., 2009, 4, 118-126).

In another US Patent 6,174,541 by Jin et. al, plant derived cell migration agent, Indole-3-acetic acid (Auxin) and its derivatives were taught to facilitate the healing process of skin tissue. The said cell migration agent was also useful in treating the wrinkled skin to minimize the effect of photo aging.

Therefore the above existing state of the art clearly reveals that although several efficacious wound healing and cell migration inducing compounds are already known, there has been a continuing need in the art to provide for newer alternatives/ superior skin care agents which would promote wound healing and would be safe to apply on the skin and specially harnessed from renewable resource materials such as plants.

OBJECTS OF THE INVENTION

It is thus the principal object of the present invention to provide for a wound healing agent having cell migration characteristics that would be safe and compatible as a skin care active agent selectively extracted from safe natural and renewable sources such as plants.

Yet another object of the present invention is directed to a skin care formulation including cosmetic or dermatological composition involving the said extract in an effective amount along with a cosmetically acceptable vehicle with or without other skin benefiting agents that would be effective in promoting wound healing to thereby cater to the needs of various people.

Yet another object of the present invention is directed to a wound healing agent comprising a selective fraction of the natural material extract/ isolate and /or formulations thereof that can be sourced simply and cost-effectively from natural renewable materials for variety of skin care applications/ formulations and the like.
A still further object of the present invention is directed to skin care formulations which besides possessing cell migration or wound healing characteristics involving

selective extracts/ actives isolated from plant sources would also have good skin compatibility.

SUMMARY OF THE INVENTION

Thus according to the basic aspect of the invention there is provided a wound healing agent comprising an effective amount of cell migrating active comprising anyone or more of hydrocarbon compounds of the general

Formula 1
H-R-R'-H Formula 1
Wherein R-R'= Saturated hydrocarbons having 14-20 carbon atoms
In a preferred aspect of the invention the said wound healing agent is a linear hydrocarbon comprising an effective amount of the active ingredient selected from at least anyone or more of eicosane of Formula 2 and hexadecane of Formula 3
CH3~(CH2)i8~CH3
Formula 2 (Eicosane)
CH3-(Cri2)i4 ~Crl3 Formula 3 (Hexadecane)

Also, in keeping with the requirement for a wound healing/ skin care agent possessing cell migration characteristics, it has surprisingly been found that the above said compounds meet the much desired criteria of a superior wound healing agent with superior cell migrating characteristics by way of the enhanced distance traveled by the cells which is found to be simple in affecting wound healing of the skin under low effective concentrations, which is also cost-effective and safe in having less or no side effects in association to said wound healing effect.

Most importantly, efforts were further directed to provide said selective compounds of Formula 1 and specifically of Formula 2 and Formula 3 revealing cell migration characteristics resulting in wound healing of the skin sourced from plant species and according to yet another aspect of the invention there is provided the said wound healing/ skin care agent wherein the said actives of Formula 1 preferably selected

from at least anyone or more of eicosane of Formula 2 and hexadecane of Formula 3 are obtained either via., from plant extracts/ concentrates more preferably from appropriate plant such as Ex. Syzygium cuminii Skeels.

It is important to clarify herein that the selective finding of the present invention resides in the identification of the above said selective fraction of the extract from the plant Syzygium cuminii in a particular solvent media sourcing the said actives of Formula 1 revealing cell migration characteristics. The said selective plant sourcing the said selective actives of Formula 1 is indeed surprising and in significant deviation from its prior known characteristics/ biological properties as discussed hereunder:
An unidentified compound isolated from ethanolic extract of the seeds of S. cuminii showed hypoglycemic and anti-diabetic activity (Chanez et al., C. R. Acad. Sci., Paris, Ser. D. 1967, 268, 1119-1123). Further two compounds, betulinic and platanic acids were isolated from the leaves of S. clavaiflorum wherein such compounds were found to be inhibitors of HIV replication in H9 lymphocyte cells (Fujioka et. al., J. Nat. Prod., 1994, 57, 243-47). An anti-fertility activity compound, oleanolic acid was isolated from the flowers of Eugenia jambolana, which was evaluated in male albino rats that showed positive results (Rajasekaran et. al., J. ethnopharmacol., 1988, 24, 115-121). Two anti-diabteic compounds, peptidyglycan and an oligosaccharide of 6.0 and 1.2kD were also isolated by a simple two step purification from fruit pulp of E. jambolana (Kelkar et. al, Phytomedicine, 1997, 3, 353-59). Also the leaf essential oil of 5. cuminii was found to possess substantial antibacterial property (Shafi et. al., Fitoterapia, 2002, 73, 414-16).The ethanolic extract of the bark and seed extract of S. cuminii were shown to hold anti¬inflammatory activity (Muruganandan et.al., Fitoterapia, 2001, 72, 369-75 and Chaudhuri et. al., Phytotherapy Research, 1990,4, 5-10). Again, the leaf extract of S. cuminii was found to be effective in reducing the radiation induced DNA damage in the cultured human peripheral blood lymphocytes (Jagetia et. al., Toxicology Letters, 2002, 132, 19-25).

In the backdrop of the abovesaid, it is thus a surprising finding by way of the present invention that the actives of Formula 1, preferably of Formula 2 and 3 can be indeed sourced from Ex. Syzygium cuminii Skeels by the identification of a selective fraction
in the extract of the said plant wherein the said actives in the said selective fraction were surprisingly and selectively efficacious with regard to wound healing activity.
In another aspect of the invention there is provided a wound healing agent wherein the said actives of Formula 1 and preferably eicosane of Formula 2 or hexadecane of Formula 3 are obtained via., Synthesis or petrochemical process or from selective plant extract extracted either as a single entity of anyone or more of the actives of Formula 1 preferably eicosane of Formula 2 or hexadecane of Formula 3 or as a semi pure mixture of the compounds of Formula 1 preferably a semi-pure mixture of eicosane of Formula 2 and hexadecane of Formula 3.

In yet another aspect of the invention there is provided a wound healing agent wherein the said agent comprises at least one or more of said hydrocarbon compounds of Formula 1 and preferably eicosane of Formula 2 or preferably hexadecane of Formula 3 as extracts/ concentrates from plant Ex. Syzygium cuminii Skeels., preferably as cell migrating active in methanol or more preferably in ethyl acetate or most preferably in hexane.

Preferably, the said wound healing agent comprises the selective fraction of Ex. Syzygium cuminii Skeels comprising extracts identified for purification and showing a spot at Rf 0.75 in thin layer chromatography using pre-coated silica gel plates with hexane as the mobile phase comprising of at least three ingredients; one of which has a mass spectrometric M+value of m/z 198 mass units and is present to an extent of not less than 10 % of the three identified peak areas of GC eluting at Rt of 7.8 minutes, second peak M+ value of m/z 226 mass units and not less than 15%, eluting in GC at 10.59 min, the third peak M+ value of m/z 282 mass units and not more than 15% eluting in GC at 28.22 minutes.

More preferably, the said selective fraction preferably comprises a selective ingredient eicosane of Formula 2 comprising a mass spectrometric m/z value of 282 mass units and retention time (Rt) at 28.22 minutes in GC or hexadecane of Formula 3 comprising a mass spectrometric m/z value of 226 mass units and retention time (Rt) at 10.59 minutes in GC.

In another aspect of the invention there is provided a dermopharmaceutical / cosmetic composition for topical use comprising an effective amount of said wound healing agent

and

a cosmetically/ dermopharmaceutically acceptable vehicle with or without other skin care benefiting agents.

In a preferred aspect of the present invention there is provided a dermopharmaceutical/ cosmetic composition for topical use comprising an effective amount of wound healing agent from extracts/ concentrates from the said plants Ex. Syzygium cuminii Skeels and preferably as a methanol extract/ concentrate or more preferably as a ethyl acetate extract/ concentrate or most preferably as a hexane extract/ concentrate

and

a cosmetically/ dermopharmaceutically acceptable vehicle with or without other skin care benefiting agents.

In yet another preferred aspect of the present invention there is provided a dermopharmaceutical/ cosmetic composition for topical use comprising
a) 0.0001 wt. % to 20 wt. %, preferably 0.001 wt. % to 10 wt. % and more
preferably 0.01 wt. % to 5 wt. % of the said wound healing agent
and
b) a cosmetically/ dermopharmaceutically acceptable vehicle with or without other

skin care benefiting agents.

Advantageously, the said dermopharmaceutical/ cosmetic composition for topical/ external use in the form of products such as creams, emulsions, gels, lotions, oils, sticks, sprays, packs, wraps, woven or non-woven wipes, films or patches and the like as a vehicle of the said wound healing composition.

In yet another aspect of the present invention there is provided the said dermopharmaceutical /cosmetic composition comprising said wound healing agent as the cell migrating active comprising selectively any one or more of (a) pure compound of Formula 1 preferably selected from eicosane of Formula 2 or hexadecane of Formula 3; (b) an extract/ concentrate of an appropriate natural source material comprising the anyone or more of the compounds of Formula 1 preferably eicosane of Formula 2 or hexadecane of Formula 3; (c) a selective fraction of an extract/ concentrate of an appropriate natural source material comprising the said compounds of Formula 1 as a semi pure mixture effective for wound healing preferably eicosane of Formula 2 or hexadecane of Formula 3 and (d) any natural source material comprising anyone or more of the said active linear hydrocarbon compounds of Formula 1 or preferably eicosane of Formula 2 or hexadecane of Formula 3.

In still another aspect of the present invention there is provided a process for producing the said wound healing agent comprising selectively extracting the wound healing agent comprising anyone or more of the actives of Formula 1 and preferably of Formula 2 or Formula 3 from natural renewable plant source (Ex. Syzygium cuminii Skeel)
comprising the steps of:

a) subjecting a selective source from said plants preferably leaves, in powder form,
to solvent extraction technique to provide for an extract;

b) removing the solvent from the extract under vacuum to provide for a concentrate;

c) subjecting the concentrate to vacuum liquid chromatography for obtaining selective fractions;

d) subjecting the said selective fractions to biological studies;

e) subjecting the active identified fraction to column chromatography;

f) pooling the identified active fractions and concentrating the resulting solution to dryness in vacuo to provide for a concentrate and obtaining therefrom at least one active wound healing agent of Formula 1.

The details of the invention, its objects and advantages are explained hereunder in greater detail in relation to non-limiting exemplary illustrations as per the following exemplary illustrations:

DETAILED DESCRIPTION OF THE INVENTION

As discussed herein before, the present invention provides a single compound or mixture of linear hydrocarbon compounds as extracts or pure compounds acting as wound healing/ skin care agent and, in particular, cell migrating/ promoting agent in a cosmetic/ dermopharmaceutical composition for topical use. The present invention also includes a process for the isolation of the said selective skin care actives or pure compounds/ agents and manufacturing the said compositions with or without other carriers/ skin care additives after its selective isolation from plants (Ex. Syzygium cumini Skeels., Syn. Eugenia jambolana) and its illustration in achieving cell migrating effect.

Syzygium cuminii (Syn. Eugenia jambolana), is a large evergreen tree attaining 30 m in height and 3.6 m in girth. The ripe fruit is widely eaten in India. It is generally sub-acidic and astringent to taste, but is palatable. A wine is prepared from ripe fruit in Goa. Fruits are used for making preserves, squashes and jellies. A good jelly can be made from the purple-fleshed fruits in combination with the white fleshed fruits or other fruits having high content of pectin. A jam can also be prepared from pitted fruits. Steam distillation of leaves gave an essential oil with a pleasant odor. The essential oil contains terpenes, 1-limonene and dipentene 20%, sesquiterpens of cadalane type 40% and sesquiterpene of azulene type 10% or less. The stem bark contains betulenic acid, p-sitosterol, freidelin, epi-friedelanol with a fatty acid, tannins (10-12%), gallic acid, ellagic acid and myricetin (The wealth of India).
Accordingly the present invention provides for a wound healing agent comprising an effective amount of cell migrating active comprising anyone or more of linear hydrocarbon compounds of the general Formula 1

H-R-R'-H Formula 1
Wherein R-R'= Saturated hydrocarbons having 14-20 carbon atoms

The said wound healing agent comprising of linear hydrocarbons comprises an effective amount of the active ingredient eicosane of Formula 2 or hexadecane of Formula 3.
CH3-(CH2)l8"CH3
Formula 2 (Eicosane)
CH3-(CH2)14-CH3 Formula 3 (Hexadecane)
A further aspect of the invention is based upon the selective and surprising finding wherein the abovesaid actives selected from anyone or more of linear hydrocarbons of Formula 1 or specifically eicosane of Formula 2 or hexadecane of Formula 3 are derived from the selectively identified fraction of the extract of appropriate plant variety such as Ex. Syzygium cumini Skeels., effectively providing for dermopharmaceutical/ cosmetic composition with excellent wound healing thereby possessing cell migration characteristics.
Accordingly, the said cosmetic composition for topical use possessing wound healing activity comprises selectively pure compound of Formula 1 as a concentrate of an appropriate naturaJ isolate or an extract/ concentrate of the natural source material or even the natural source material itself.
According to a further aspect of the invention, the said cosmetic composition for topical use includes skin care/ cell migration actives comprising extract/ concentrate from selective plant (Ex. leaves of Syzygium cuminii Skeels.). The said extract or concentrate is preferably in methanol or more preferably in ethyl acetate and most preferably in hexane containing eicosane of Formula 1 or specifically of Formula 2 or hexadecane of Formula 3 or atleast one compound of Formula 1.
The preferable solvents are less polar solvents like: hexane, benzene, toluene etc. and chlorinated solvents like: dichloromethane, chloroform, carbon tetrachloride etc. Also, solvents such as esters like: methyl acetate, ethyl acetate; ethers like: diethyl ether, methyl ethyl ether etc.; ketones like: acetone, ethyl methyl ketone etc.; and alcohols like: methanol, ethanol, propanol, butanol etc.; and aqueous alcoholic

solutions like: aq. methanol, aq. ethanol etc. and or combination thereof can be used.
In another aspect, the present invention relates to a process for providing a compound isolated from plants to be used as active ingredient in the cosmetic compositions of the invention. The processes may be devised through isolation procedures from renewable natural resources such as plants. The process for isolating the compound from S. cuminii Skeels of the invention comprises their extraction from leaves, in air-dried form, using appropriate solvents, followed by purifying the extract and separating the compound/s of the invention according to procedures generally known to those skilled in the art. The isolation process may result in a pure compound or in a mixture of the compounds of the invention in different and defined proportions, or may yield an extract enriched in the compound/s of the invention.
In accordance with another aspect of the present invention there is provided a process for the extraction and isolation of the said skin care/ cell migrating agent for wound healing comprising anyone or more of the active compounds of Formula 1 and preferably of eicosane of Formula 2 or hexadecane of Formula 3 harnessed preferably from a plant source (Ex. Syzygium cuminii Skeel)
comprising the steps of:

a) Subjecting a selective source from plants, in powder form, to solvent extraction technique to provide for an extract;

b) Removing the solvent from the extract under vacuum to provide for a concentrate;

c) Subjecting the concentrate to Vacuum liquid chromatography;

d) subjecting the fractions to biological studies;

e) Subjecting the active concentrate to column chromatography;

f) Pooling the active fractions and concentrating the resulting solution to dryness in vacuo to provide for a concentrate and obtaining there from atleast one active agent of the above said Formula 2 or Formula 3.

Accordingly, the invention provides for a selective compound as wound healing agent, a cosmetic/ dermopharmaceutical composition for topical use comprising of the said selective wound healing agent, a process for manufacturing the said composition either by isolation from natural sources, as a single entity of Formula 2 or Formula 3 or any one of the actives of Formula 1 or in a semi pure mixture of compounds of Formula 1 to achieve wound healing effect.

Preferably, the wound healing agent involving the selective fraction of the extract comprise extracts identified for purification showing a spot at Rf 0.75 in thin layer chromatography using pre-coated silica gel plates with hexane as the mobile phase that comprises of at least four ingredients; one of which has a mass spectrometric M+ value of m/z 198 mass units and is present to an extent of not less than 10 % of the three identified peak areas of GC eluting at Rt of 7.80 minutes, second peak M+ value of m/z 226 mass units and not less than 15%, eluting in GC at 10.59 min, the third peak M+ value of m/z 282 mass units and not more than 15% eluting in GC at 28.22 minutes

The said cosmetic/ dermopharmaceutical composition comprising of the said wound healing agent is a selective fraction of the selective herbal extract comprising purified or processed extract on the basis of its cell migrating/ promoting characteristics as measured by the increase in cell viability.

Preferably, the said wound healing selective fraction comprises a selective ingredient eicosane of Formula 2 comprising a mass spectrometric m/z value of 282 mass units and retention time (Rt) at 28.22 minutes in GC or hexadecane of Formula 3 comprising a mass spectrometric m/z value of 226 mass units and retention time (Rt) at 10.59 minutes in GC.

More preferably, a dermopharmaceutical/ cosmetic composition is provided wherein the said wound healing agent comprises selectively any one or more of (a) selective fraction of the selective herbal extract/ concentrate in an effective amount in appropriate solvent; (b) a purified and processed selective fraction of the selective extract/ concentrate in an effective amount in appropriate solvent; and (c) extract/ concentrate of the appropriate natural source material itself in an effective amount

extracted by an appropriate solvent in the presence of an adsorption material such as silica gel.

It is thus possible by way of the present invention to provide for a selective wound healing agents sourced from plant (Ex. S. cuminii Skeels.) that would be safe and compatible as a skin care active agent.

Importantly, the inventive compounds and compositions are directed to meet the much required simple, cost-effective and variety of skin care applications/ formulations and the like which would be safe for topical use.

In accordance with another aspect of invention a dermopharmaceutical/ cosmetic composition for topical use is provided comprising an effective amount of wound healing agent and a cosmetically acceptable vehicle with or without other skin care benefiting agents.

Specifically various components are included in the cosmetic hair care compositions of the present invention that are usually used for cosmetic compositions such as, an aqueous component, an oily component, a powder component, alcohols, esters, a surfactant, a moisturizer, a whitening agent, an antioxidant, an ultraviolet ray absorbent, a thickener, a colouring material, perfume, an anti-oxidant, pH regulator, a chelating agent, and antiseptics, that can be blended with these cosmetics.

The ingredients essentially employed in such a composition of this invention are surfactants, oil bodying agents, emulsifiers, fats and waxes, stabilizers, polymers, silicone compounds, deodorants and antimicrobial agents, film formers, swelling agents preservatives, thickeners / herbal extracts / concentrates / active ingredients, pH adjusting agents, perfumes oils, and water to qs.

The composition according to the invention can be used for the production of cosmetic and/or dermopharmaceutical preparations for topical use in the form of products, e.g. lotions, creams, gels, oils, sticks, sprays, packs, wraps, woven or non-woven wipes, films, patches, emulsions, wax/fat compositions, as a vehicle of the said composition.

These preparations can also comprise further auxiliaries and additives, such as, surfactants, oily bodies, emulsifiers, pearlescent waxes, bodying agents, thickeners, superfatting agents, stabilizers, polymers, silicone compounds, fats, waxes, lecithins, phospholipids, biogenic active ingredients, deodorants, antiperspirants, film formers, swelling agents, hydrotropes, solubilizers, preservatives, perfume oils, dyes, pH regulating agents and the like.
Thus the hair care composition of the invention can also incorporate conventional cosmetic and hair/ skin care ingredients as discussed hereunder:
Surfactants

Surface-active substances which may be present are anionic, nonionic, cationic and/or amphoteric surfactants, the content of which in the compositions is usually about 1 to 70% by weight, preferably 5 to 50% by weight and in particular 10 to 30% by weight.
Oily Bodies

Suitable oily bodies can also be incorporated which include for example, Guerbet alcohols based on fatty alcohols having 6 to 18, preferably 8 to 10, carbon atoms, esters of linear C6 -C22-fatty acids with linear or branched C6-C22-fatty alcohols or esters of branched C6-C13-carboxylic acids with linear or branched C6-C22-fatty alcohols. Such oily bodies can be used in amounts of 0.1 to 50.0 % by wt.
Emulsifiers

Suitable emulsifiers are, for example, nonionic surfactants.
The addition products of ethylene oxide and/ or of propylene oxide onto fatty alcohols, fatty acids, alkylphenols or onto castor oil are also known, commercially available products which can be included. Alkyl and/or alkenyl oligoglycosides, their preparation and their use are known from the prior art.
Furthermore, zwitterionic surfactants can be used as emulsifiers. The term "zwitterionic surfactants" refers to those surface-active compounds which carry at least one quaternary ammonium group and at least one carboxylate and one sulfonate group in the molecule. Likewise suitable emulsifiers are ampholytic

surfactants. The term "ampholytic surfactants" means those surface-active compounds which, apart from a C.sub.8/18- alkyl or -acyl group in the molecule, contain at least one free amino group and at least one —COOH or — S03H group and are capable of forming internal salts. Finally, cationic surfactants are also suitable emulsifiers, those of the quaternary ester type, preferably methyl quaternized difatty acid triethanolamine ester salts, being particularly preferred. Such emulsifiers can be used in amounts of 0.1 to 50.0 % by wt.
Fats, hydrocarbons and Waxes

Typical examples of fats are glycerides, i.e. solid or liquid vegetable or animal products which consist essentially of mixed glycerol esters of higher fatty acids, hydrocarbons are obtained from petroleum products, which contains more than 20 carbon atoms, suitable waxes are inter alia natural waxes. In addition to the fats, suitable additives are also fat-like substances, such as lecithins and phospholipids. The term lecithin is understood by the person skilled in the art as meaning those glycerophospholipids which form from fatty acids, glycerol, phosphoric acid and choline by esterification. Lecithins are thus frequently also known as phosphatidylcholines (PC). Examples of natural lecithins which may be mentioned are the cephalins, which are also referred to as phosphatidic acids and represent derivatives of 1, 2-diacyl-syn-glycerol-3-phosphoric acids. By contrast, phospholipids are usually understood as meaning mono- and, preferably, diesters of phosphoric acid with glycerol (glycerophosphates), which are generally considered to be fats. In addition, sphingosines and sphingolipids are also suitable. Such fats, hydrocarbons and waxes can be used in amounts of 0.1 to 99.5 % by wt.
Moreover, known Pearlescent Waxes, Bodying Agents and Thickeners as well as superfatting agents can also be used.

Superfatting agents which can be used are substances such as, for example, lanolin and lecithin, and polyethoxylated or acylated lanolin and lecithin derivatives, polyol fatty acid esters, monoglycerides and fatty acid alkanolamides, the latter also serving as foam stabilizers. Such super fatting agents can be used in amounts of 0.1 to 15 % by wt.

Stabilizers
Stabilizers which can be used are metal salts of fatty acids, such as, for example, magnesium, aluminum and/or zinc stearate or ricinoleate. Such stabilizers can be used in amounts of 0.1 to 10% by wt.
Polymers

Suitable polymers include anionic, zwitterionic, amphoteric, nonionic and cationic polymers. Suitable polymers are, for example, suitable polymers and thickeners are listed in Cosm. Toil. 108, 95 (1993). Such polymers can be used in amounts of 0.001 to 5 % by wt.

Silicone Compounds
Suitable silicone compounds which are volatile or non-volatile, for example, dimethylpolysiloxanes, methylphenylpolysiloxanes, linear and cyclic silicones, and amino-, fatty acid-, alcohol-, polyether-, epoxy-, fluorine-, glycoside- and/or alkyl-modified silicone compounds, which can either be liquid or in resin form at room temperature. Also suitable are simethicones, which are mixtures of dimethicones having an average chain length of from 200 to 300 dimethylsiloxane units and hydrogenated silicates. Such silicone compounds can be used in amounts of 0.1 to 10% by wt.
Biogenic Active Ingredients

Biogenic active ingredients can also be included meaning those which do not arise from the plant S. cuminii Skeels, such as, for example, tocopherol acetate, tocopherol palmitate, ascorbic acid, (deoxy)ribonucleic acid and fragmentation products thereof, retinol, bisabolol, allantoin, phytantriol, panthenol, menthol, anethol, AHA acids, amino acids, ceramides, pseudoceramides, essential oils, further plant extracts and additional vitamin complexes. Such biogenic active ingredients can be used in amounts of 0.001 to 5.0% by wt.

Deodorants and Antimicrobial Agents
Cosmetic deodorants are also possible ingredients which can be added to counteract,
mask or remove body odors. Body odors arise as a result of the effect of skin bacteria on apocrine perspiration, with the formation of degradation products which

have an unpleasant odor. Accordingly, deodorants comprise active ingredients which act as antimicrobial agents, enzyme inhibitors, odor absorbers or odor masking agents.
Antiperspirants can also be added in the cosmetic preparation to reduce the formation of perspiration by influencing the activity of the eccrine sweat glands, thus counteracting underarm wetness and body odor.
Suitable astringent antiperspirant active ingredients are primarily salts of aluminum, zirconium or of zinc.

Customary water-soluble additives are, for example, preservatives, water-soluble fragrances, pH regulators, e.g. buffer mixtures, water-soluble thickeners, e.g. water-soluble natural or synthetic polymers, such as, for example, xanthane gum, hydroxyethylcellulose, polyvinylpyrrolidone or high molecular weight polyethylene oxides.

Such deodorant and antimicrobial agents can be used in amounts of 0.001 to 5.0% by wt.
Film Formers

Conventionally the film formers are selected from, for example, chitosan,
microcrystalline chitosan, quaternized chitosan, polyvinylpyrrolidone, vinylpyrrolidone - vinyl acetate copolymers, polymers of the acrylic acid series, quaternary cellulose derivatives, collagen, hyaluronic acid and salts thereof, and similar compounds. Suitable film formers can be used in amounts of 0.001 to 5.0% by wt.
Thickeners

Thickeners/ rheology modifiers may also be utilized as part of the cosmetically acceptable carrier of compositions according to the present invention. Amounts of the thickener may range from 0.0001 to 10%, usually from 0.001 to 5%, by weight.

Swelling Agents
The swelling agents for aqueous phases may be montmorillonites, clay mineral substances, Pemulen, and a I kyl-modified Carbopol grades (Goodrich). Such additives can be incorporated in amounts of 0.001 to 2.0% by wt.
Hydrotropes

To improve the flow behavior, hydrotropes can be added, such as, for example, ethanol, isopropyl alcohol, or polyols, can also be used. Such additives can be incorporated in amounts of 0.1 to 15% by wt.
Preservatives

Suitable preservatives are, for example, phenoxyethanol, formaldehyde solution, parabenes, pentanediol or sorbic acid, and the other classes of substance listed in Annex 6, Part A and B of the Cosmetics Directive. Such preservatives can be incorporated in amounts of 0.001 to 5 % by wt.
Other adjunct minor components may also be incorporated into the cosmetic compositions. These ingredients may include coloring agents, opacifiers, and perfumes. Amounts of these other adjunct minor components may range anywhere from 0.001 % up to 20 % by wt. of the composition.
The details of the invention, its objects and advantages are explained hereunder in greater detail in relation to non-limiting exemplary illustrations as per the following examples:

EXAMPLES:
Example 1 - Extraction of leaves of Syzygium cuminii Skeels
The air-dried leaves of S. cuminii Skeels., (190 g) was powdered, extracted with methanol through soxhlet apparatus for about 8 hrs. The dilute extract was concentrated under reduced pressure to get 52.0 g of crude methanol concentrate. The crude methanol concentrate showed cell migration property.

Example 2 - Isolation of elcosane and hexadecane from methanolic extract of S. cuminii Skeels

The methanol concentrate of S. cuminii Skeels (52g) was dissolved in methanol and adsorbed on silica gel (100-200 mesh, 150g). 350g of silica gel (100-200 mesh) was packed in glass G3 sintered funnel followed by loading the adsorbed silica gel which was then washed with chloroform, ethyl acetate and finally with methanol to get fractions weighing 6g, 3g and 26.5g correspondingly. The chloroform fraction was found to show cell migration property. The said chloroform fraction was purified by repeated silver nitrate impregnated silica gel columns by using hexane as mobile phase and finally isolated 78 mg of the compounds with Rf 0.75 in Hexane. The compound was identified as mixture of three major compounds, hexadecane, tetradecane and, eicosane based on GC MS data. All the said compounds were investigated for cell migration property as the chloroform fraction containing and then showed good cell migrating activity.

Example 3: GC Instrumentation and Chromatography Conditions i) The Shimadzu GC-17A capillary GC unit consists of FID detector. The chromatographic system consists of capillary glass column of DB-1 column (30 m length X 0.53 mm ID, 5^ film thickness) where nitrogen was used as the carrier gas. The total flow rate of nitrogen is 21 ml/ minute. The oven and Injection port temperatures are 200°C and 225°C and detector temperature is 250°C. The active fraction was dissolved in heptane and directly injected for the GC analysis. The retention time of the three major peaks are 7.8, 10.59, and 28.21 minutes.

Example: 4 Fibroblast Migration assay:

Materials and Methods

Primary cultures of human skin dermal fibroblasts were established from human split skin grafts. The process of handling the skin biopsy and isolation of the cells from the biopsy is approved by the institutional ethical committee.

The collected skin biopsy is transferred into a 50 ml tube with phosphate buffer solution (PBS). It was washed with betadine or antibiotics for 10 minutes. It was further incubated in 10X antibiotics for 10 minutes. It was rinsed with sterile PBS or water 3 times. The skin biopsy is then transferred to a new petri dish with epidermis side up. Trypsin/dispase solution is transferred in to a 60 mm Petri dish. The skin was lifted with two forceps and was floated on trypsin/ dispase solution without leaving any folds on the skin. It was incubated overnight at 4°C. Next day, the skin was transferred to a new Petri dish with forceps. The epidermis was kept faced down and the dermis was separated from the epidermis. The dermis was transferred to a separate Petri dish for preparation of fibroblasts. The Dermis was chopped in to small pieces with scissors and incubated in DMEM with 0.03% collagenase for 24-48 hours.
The released fibroblasts were cultured in Dulbecco's modified Eagle's medium, supplemented with 100 ng/ml penicillin, 100 (Kj/ml streptomycin, and 10% FCS, at 37 C and 5% C02 in a humidified atmosphere. The cells used were between passage 3 and 6.

Methodolgy:

IX 105 cells were seeded and allowed to adhere for 24 hours. Cells were grown to 80% confluency and serum starved for 24 hours. In vitro scratched wound is created by removing the monolayer cells using a sterile rubber policemen or pipette tip, rinsed with phosphate buffer saline and supplemented with fresh medium without serum. The cells were treated with different concentrations of the compound of the invention both in the presence and absence of the said compound. Cells treated with 10% serum or 10 nM EFG served as positive control and with vehicle alone as controls. 24 hours after the treatment the cells were photographed and the distance traveled by the cells is measured under inverted microscope. Distance traveled in the presence of the compound is compared to that of the distance traveled in the absence of the compound and expressed as percentage of cell migration.

It is thus concluded from the above table 1 that indicates that active fraction is superior than the crude in methanol which promotes the wound healing process. It is also noteworthy that the active eicosane or hexadecane when administered at 25 lKj/ml solution induces wound healing at a faster rate with % of cell migration at 70.

Thus the present invention provides for the hitherto desired dermopharmaceutical/cosmetic composition for external application in wound healing and methods for making the same. More specifically, the present invention is directed to serve the much desired need for skin care actives/ formulations which would serve as more efficacious, safe to apply and natural healing agent with better consumer acceptance. Importantly, the present invention thus favours in obtaining a wound healing/ skincare agent where the actives of the above said Formula 1 and eicosane of Formula 2 or hexadecane of Formula 3 are either preferably obtained from selective fraction of the extract of the selective safe and renewable sources, or obtained as pure compound via., Synthesis or isolated as an extract from appropriate plant variety such as Syzygium cuminii Skeels that can be used either singly or in combination with a cosmetically/ dermopharmaceutically acceptable vehicle.

WE CLAIM:

1. A wound healing agent comprising an effective amount of cell migrating active comprising anyone or more of hydrocarbon compounds of the general Formula 1

H-R-R'-H
Formula 1
Wherein R-R'= Saturated hydrocarbons having 14-20 carbon atoms

2. A wound healing agent as claimed in claim 1, wherein said hydrocarbon comprises an effective amount of the active ingredient selected from at least anyone or more of eicosane of Formula 2 and hexadecane of Formula 3.

CH3-(CH2.)l8"CH3

Formula 2 (Eicosane)

CH3-(CH2)14-CH3 Formula 3 (Hexadecane)

3. A wound healing agent as claimed in claims 1 and 2 wherein the said actives of Formula 1 and preferably eicosane of Formula 2 or hexadecane of Formula 3 are obtained via., Synthesis or petrochemicals or from selective plant extract either as a single entity of anyone or more of the actives of Formula 1 preferably eicosane of Formula 2 or hexadecane of Formula 3 or as a semi pure mixture of the compounds of Formula 1 preferably a semi-pure mixture of eicosane of Formula 2 and hexadecane of Formula 3.

4. A wound healing agent as claimed in anyone of the preceding claims wherein the said agent comprises at least any one or more of said hydrocarbon compounds of Formula 1 preferably eicosane of Formula 2 or preferably hexadecane of Formula 3 as extracts/ concentrates from plant Ex. Syzygium cuminii Skeels., preferably as cell migrating active in methanol or more preferably in ethyl acetate or most preferably in hexane.

5. A wound healing agent as claimed in anyone of the preceding claims wherein the said agent comprises the selective fraction of Ex. Syzygium cuminii Skeels comprising extracts identified for purification and showing a spot at Rf 0.75 in thin layer chromatography using pre-coated silica gel plates with hexane as the mobile phase comprising of at least three ingredients; one of which has a mass spectrometric M+ value of m/z 198 mass units and is present to an extent of not less than 10 % of the three identified peak areas of GC eluting at Rt of 7.80 minutes, second peak M+ value of m/z 226 mass units and not less than 15%, eluting in GC at 10.59 min, the third peak M+ value of m/z 282 mass units and not more than 15% eluting in GC at 28.22 minutes

6. A wound healing agent as claimed in claim 5 wherein the said selective fraction preferably comprises a selective ingredient eicosane of Formula 2 comprising a mass spectrometric m/z value of 282 mass units and retention time (Rt) at 28.22 minutes in GC or hexadecane of Formula 3 comprising a spectrometric m/z value of 226 mass units and retention (Rt) at 10.59 minutes in GC.

7. A dermopharmaceutical/ cosmetic composition for topical use comprising an effective amount of wound healing agent as claimed in claims 1 to 6

and

a cosmetically/ dermopharmaceutically acceptable vehicle with or without other skin care benefiting agents.

8. A dermopharmaceutical/ cosmetic composition for topical use as claimed in claim 7 comprising an effective amount of wound healing agent from extracts/ concentrates from the said plants Ex. Syzygium cuminii Skeels and preferably as a methanol extract/ concentrate or more preferably as a ethyl acetate extract/ concentrate or most preferably as a hexane extract/ concentrate

and

a cosmetically/ dermopharmaceutically acceptable vehicle with or without other skin care benefiting agents.

9. A dermopharmaceutical/ cosmetic composition for topical use as claimed in Claims

7 and 8 comprising

a) 0.0001 wt. % to 20 wt. %, preferably 0.001 wt. % to 10 wt. % and more
preferably 0.01 wt. % to 5 wt. % of the said wound healing agent

and

b) a cosmetically/ dermopharmaceutically acceptable vehicle with or without other skin care benefiting agents.

10. A dermopharmaceutical/ cosmetic composition as claimed in anyone of claims 7 to 9 for topical/ external use in the form of products such as creams, emulsions, gels, lotions, oils, sticks, sprays, packs, wraps, woven or non-woven wipes, films or patches and the like as a vehicle of the said wound healing composition.

11. A dermopharmaceutical /cosmetic composition as claimed in anyone of claims 7 to 10 comprises said wound healing agent as the cell migrating active comprising selectively any one or more of (a) pure compound of Formula 1 preferably selected from eicosane of Formula 2 or hexadecane of Formula 3; (b) an extract/ concentrate of an appropriate source material comprising anyone or more of the compounds of Formula 1 preferably eicosane of Formula 2 or hexadecane of Formula 3 ; (c) a selective fraction of an extract/ concentrate of an appropriate natural source material comprising the said compounds of Formula 1 as a semi pure mixture effective for wound healing preferably eicosane of Formula 2 or hexadecane of Formula 3 and (d) any natural source material comprising anyone or more of the said active linear hydrocarbon compounds of Formula 1 or preferably eicosane of Formula 2 or hexadecane of Formula 3.

12. A process for producing a wound healing agent as claimed in anyone of claims 1 to 6 comprising selectively extracting the wound healing agent comprising anyone or more of the actives of Formula 1 and preferably of Formula 2 or Formula 3 from natural renewable plant source (Ex. Syzygium cuminii Skeel)

comprising the steps of:

a) subjecting a selective source from said plants preferably leaves, in powder
form, to solvent extraction technique to provide for an extract;

b) removing the solvent from the extract under vacuum to provide for a
concentrate;

c) subjecting the concentrate to vacuum liquid chromatography for obtaining
selective fractions;

d) subjecting the said selective fractions to biological studies;

e) subjecting the active identified fraction to column chromatography;

f) pooling the identified active fractions and concentrating the resulting solution
to dryness in vacuo to provide for a concentrate and obtaining therefrom at least one active wound healing agent of Formula 1.

13. A wound healing agent comprising one or more of hydrocarbons compounds of Formula 1 as an active ingredient in effective amounts for wound healing and a dermopharmaceutical/ cosmetic composition obtained thereof substantially as herein described and illustrated with reference to the accompanying examples.