Formulations for treating hair loss and for stimulating pigmentation in hair follicles

Abstract

The invention provides formulations for treating hair loss, methods of treatment for hair loss, and cosmetic methods for reducing hair loss or stimulating hair growth or regrowth. An exemplary formulation for treating hair loss includes: one or more of wedelolactone, demethylwedelolactone, and demethylwedelolactone glucoside; ß-sitosterol; and one or more of thymoquinone, thymohydroquinone, dithymoquinone and thymol.

Description

[0001] Formulation for treating hair loss

[0002] Technical Field

[0003] The present invention relates to a formulation for treating hair loss or stimulating hair growth or regrowth. Methods of treatment using the formulation are also provided, as are cosmetic methods for reducing hair loss or stimulating hair growth or regrowth.

[0004] Background

[0005] Normally, all humans shed around 50 to 100 of the more than 100,000 hairs on the scalp each day. But problems begin when too many follicles enter rest phase permanently or are damaged or killed. When this happens, the rate of replacement may not keep up, eventually causing loss of hair density, known as alopecia.

[0006] Alopecia can be categorised as either hormonal or non-hormonal. The loss of hair caused by testosterone and its active metabolite dihydrotestosterone (DHT) is known as androgenic alopecia (AGA) and has been investigated extensively for developing therapies for treating hair loss. Non-hormonal causes of hair loss include aging, scalp inflammatory disorders, auto-immune disorder, and mechanical stress. Certain types of alopecia present as "focal" or "patchy" hair loss and are caused by an underlying medical disorder. This includes non-scarring focal alopecias such as tinea capitis and alopecia areata and rare scarring cicatricial alopecia. On the other hand, diffuse alopecias are characterised by hair thinning and include telogen effluvium (TE), anagen effluvium (AE), and AGA.

[0007] A popular treatment where hair loss is caused by AGA or AE is topical application of Minoxidil, a potent vasodilator. The mechanism by which Minoxidil promotes hair growth is not fully understood. However, Minoxidil has been reported to promote the production of growth factors thought to stimulate hair regrowth in dermal papilla cells including vascular endothelial growth factor (VEGF), hepatocyte growth factor (HGF), and insulinlike growth factor-1 (IGF-1). Minoxidil also increases the flow of nutrients and oxygen to the hair follicle by dilating hair follicle arteries.

[0008] Another group of treatments for AGA include 5o-reductase inhibitors such as Finasteride and Dutasteride. 5o-reductase is an enzyme which converts testosterone to DHT. DHT causes hair loss by triggering hair follicle miniaturisation, reducing the anagen (active growing) phase and prolonging the telogen (rest) phase of the hair cycle. Other treatments for hair loss include triamcinolone acetonide for alopecia areata, and antifungal agents (e.g., itraconazole, griseofulvin, or fluconazole) for tinea capitis. i Unfortunately, these drugs have limited therapeutic efficacy and possess serious side effects. For example, male patients using Finasteride may experience loss of libido, erectile dysfunction, and gynecomastia. On the other hand, antifungal agents can affect liver function. While Minoxidil represents a safer treatment for hair loss, reportedly only 50% of users achieve hair growth after daily use for four months, and hair regrowth tends to be vellus (non-adult, baby) hair. Further, termination of treatment results in progressive hair loss within 12 to 24 weeks.

[0009] Due to the significance of hair, both physiologically and socially, as well as the emotional impact of hair loss, efforts have been made to try to understand the mechanisms which control hair follicle development and cycling in the hope of finding new therapies. Unlike other organs, each hair follicle normally regenerates itself cyclically in a manner that recapitulates embryonic hair follicle development. There are three main phases of the hair follicle cycle: anagen, catagen, and telogen. Anagen phase is the active hair growth phase and lasts between 2 and 8 years in human scalp hair. Catagen, which lasts between 2 and 3 weeks, is the regression phase in which growth of the hair shaft comes to an end and the lower portion of the follicle involutes. Telogen is the resting phase and ends with the original hair falling out followed by the appearance of new hair.

[0010] Both hair follicle development and cycling are controlled by a complex signalling network involving both inhibitory and stimulatory signalling cascades. These include Sonic Hedgehog (shh), Bone Morphogenetic Protein (BMP), Transforming Growth Factor-p (TGF- P), and Wingless-Integrated p-catenin (Wnt / p-catenin). Thus, attempts have been made to target one or more of these one or more of these signalling cascades to develop new therapies for alopecia. However, the development of a reliable and effective therapy exploiting these signalling pathways remains elusive.

[0011] Consequently, there is a need for alternative therapies that are effective in treating hair loss in both men and women.

[0012] Summary of Invention

[0013] Accordingly, a first aspect of the invention provides a formulation for treating hair loss, the formulation comprising: B-sitosterol; one or more of wedelolactone, demethylwedelolactone and demethylwedelolactone glucoside; and one or more of thymoquinone, thymohydroquinone and thymol.

[0014] A second aspect of the invention provides a method of treatment for hair loss, the method comprising applying a formulation according to any one of the preceding claims to a treatment area to stimulate hair growth in the treatment area and / or reduce hair loss in the treatment area.

[0015] A third aspect of the invention provides a cosmetic method for reducing hair loss or stimulating hair growth or regrowth, the method comprising applying a formulation according to any one of the preceding claims to a treatment area to stimulate hair growth in the treatment area.

[0016] A fourth aspect of the invention provides a cosmetic method for stimulating pigmentation in hair follicles.

[0017] A fifth aspect of the invention provides a formulation for treating hair loss, the formulation comprising: E. alba extract, wherein the E. alba extract comprises one or more of wedelolactone; demethylwedelolactone; and demethylwedelolactone glucoside; Saw palmetto extract, wherein the Saw palmetto extract comprises B-sitosterol; N. sativa extract, wherein the N. sativa extract comprises one or more of thymoquinone, thymohydroquinone, dithymoquinone and thymol; and P. niruri extract.

[0018] A sixth aspect of the invention provides a formulation for treating hair loss, the formulation comprising: E. alba extract, wherein the E. alba extract comprises one or more of wedelolactone; demethylwedelolactone; and demethylwedelolactone glucoside; Saw palmetto extract, wherein the Saw palmetto extract comprises B-sitosterol; N. sativa extract, wherein the N. sativa extract comprises one or more of thymoquinone, thymohydroquinone, dithymoquinone and thymol; P. niruri extract; and C. tinctorius extract.

[0019] Formulations according to the invention have been found to provide noticeable hair regrowth of non-vellus hair within as little as 5 to 6 weeks. Crucially, the invention has been found to maintain the effects of hair growth long after the completion of the treatment regimen. This is surprising because prior art treatments for hair growth typically require sustained use, with effects often dwindling shortly after discontinuation. Notably, this is the case for popular treatments such as Minoxidil and Finasteride, where discontinuation often leads to a cessation of the benefits and a return of hair loss.

[0020] Particularly surprising is the finding by the inventor that hair growth has been stimulated in scalp scars when the formulation is applied to the scalp scar. This finding is remarkable because it is widely thought that established scar tissue does not contain active hair follicles. This suggests that formulations according to the invention can activate hair follicles which are dormant or even generate new hair follicles, indicating an immunomodulatory effect of the formulation according to the invention. This is a significant improvement over formulations of the prior art which do not stimulate hair growth in scalp scars and have no evidence of inducing an immunomodulatory effect.

[0021] Description

[0022] The present invention provides a formulation for treating hair loss and / or reduced hair growth or regrowth. The formulation may be used to treat hair loss or reduced hair growth due to various underlying causes including any one or more of androgenic alopecia (AGA), telogen effluvium (TE), anagen effluvium (AE), alopecia senescence, traction alopecia or alopecias caused by mechanical stress, fungal or bacterial infections. In particular embodiments, the formulation is for treating hair loss and / or reduced hair growth or regrowth due to traction alopecia. The subject is a mammal, optionally a human, and may be male or female. The subject may be an adult human. Alternatively, the subject may be a different variety of mammal, for example an equine subject, such as a horse, a dog, a cat, or a livestock animal.

[0023] The formulation comprises B-sitosterol, one or more of thymoquinone, thymohydroquinone, dithymoquinone and thymol; and wedelolactone and / or a derivative thereof, for instance demethylwedelolactone or demethylwedelolactone glucoside.

[0024] Without being bound by theory, the inventor believes that formulations according to the invention act to slow hair loss and promote hair growth by simultaneously targeting multiple mechanisms involved in regulating hair growth. In particular, the inventor believes that the claimed combination may act to promote the proliferation and differentiation of hair follicle cells, to regulate the hair growth cycle, and to maintain a healthy hair follicle microenvironment. In this way, the invention provides a comprehensive approach which addresses the multifactorial nature of hair loss.

[0025] B-sitosterol may be able to activate the WNT / B-catenin signalling pathway in hair follicles by upregulating the expression of genes involved in the pathway, including B-catenin, and LEF-1 [1]. The WNT / B-catenin pathway plays an important role in regulating the growth and differentiation of hair follicle stem cells, and its dysregulation has been implicated in various types of hair loss, including AGA. The inclusion of B-sitosterol in the formulation may promote the growth and differentiation of hair follicle stem cells and thereby promote hair growth and / or regrowth. Wedelolactone and B-sitosterol may have anti-androgenic activity, thus making them effective to target the underlying cause of AGA. For instance, wedelolactone may inhibit the action of the enzyme 5o-reductase [2]. The mechanism of anti-androgenic activity in B-sitosterol is not known, however, one study reported that B-sitosterol inhibited the 5o- reductase type 1 isoform in hamster prostate [3].

[0026] The inventor has surprisingly recognised that the combination of both wedelolactone (and / or one of its derivatives demethylwedelolactone or demethylwedelolactone glucoside) with B-sitosterol provides an overall more powerful effect than a single agent with anti-androgenic activity alone. In particular, the combination of wedelolactone and B- sitosterol may target different steps in the androgen pathway, or different 5o-reductase isoforms, thereby providing a more comprehensive approach to androgen-dependent hair loss.

[0027] B-sitosterol, wedelolactone, thymoquinone, thymohydroquinone and thymol also have anti-inflammatory and antioxidant activity. The application of these compounds to the treatment area therefore acts to reduce oxidative and inflammatory damage to the hair follicle and maintain a normal hair growth cycle. Importantly, the inventor believes that the claimed combination provides an overall more powerful anti-inflammatory effect by targeting a variety of different pathways or mediators involved in inflammation. Each of the claimed constituents exerts its anti-inflammatory effect via a number of molecular mechanisms and signalling pathways, however the primary mechanisms differ for the different constituents. For instance, thymoquinone is thought to exert its effect predominantly via modulation of pro-inflammatory enzymes, such as cyclooxygenase-2 (COX-2)

[0030] . On the other hand, the primary mechanism of anti-inflammatory action of wedelolactone is thought to be the inhibition of the nuclear factor-kappa B (NF-KB) pathway [31, 32]. In contrast, B-sitosterol is thought to achieve its anti-inflammatory effect by reducing inflammation markers, such as TNF-o, IL-ip, IL-6, IL-8, while promoting the anti-inflammatory IL-10

[0033] . B-sitosterol is also thought to inhibit nuclear translocation of NF-KB

[0034] . Therefore, by combining B-sitosterol, wedelolactone, thymoquinone, thymohydroquinone and thymol, an enhanced anti-inflammatory action can be achieved.

[0028] B-sitosterol is also purported to have angiogenic effects, and thus may improve blood flow to the hair follicle, thus helping to improve the flow of nutrients an oxygen to the hair follicle [5]. Thymoquinone, thymohydroquinone, dithymoquinone and thymol also all have potent anti-fungal activity [6], [7]. As such, these compounds may be effective in treating types of alopecia which are associated with fungal infections, such as tinea capitis. The inventor has found that these compounds may have beneficial effects on patients with no fungal infection by mimicking the action of Ketoconazole in treating alopecia. Ketoconazole is an antifungal agent which has been found to be effective in treating alopecias having various underlying causes, including those which are not associated with fungal infection. The mechanism by which Ketoconazole treats alopecia is not fully understood. However, some types of alopecia, despite not being traditionally associated with fungal infections, may still involve fungal colonisation or subclinical fungal overgrowth that is not easily detectable. Antifungal agents could help address these hidden fungal factors and potentially improve hair loss conditions.

[0029] Although certain effects described hereinabove are described with reference to wedelolactone, in formulations according to the invention, wedelolactone may be substituted by one or more of demethylwedelolactone or demethylwedelolactone glucoside. Demethylwedelolactone and demethylwedelolactone glucoside are derivatives of wedelolactone which retain many of the core characteristics of wedelolactone.

[0030] Optionally, the formulation comprises at least two different 5o-reductase inhibitors. Combining two different 5o-reductase inhibitors may provide benefits over using a single 5o-reductase inhibitor alone. In particular, different 5o-reductase inhibitors are likely to involve different mechanisms of action to block the conversion of testosterone to DHT. The inventor has also recognised different 5o-reductase inhibitors are likely to have different durations of action. By combining two different 5o-reductase inhibitors with different pharmacokinetic profiles, more sustained inhibition of DHT production may be achieved.

[0031] Optionally, the formulation includes at least three different 5o-reductase inhibitors. Further optionally, the formulation includes at least four different 5o-reductase inhibitors. Further optionally, the formulation includes at least five different 5o-reductase inhibitors. Further optionally, the formulation includes at least six different 5o-reductase inhibitors. By increasing the number of different 5o-reductase inhibitors, an even greater spread of inhibitory action can be achieved, thus providing more sustained inhibition of DHT production.

[0032] The 5o-reductase inhibitors in the formulation may be natural or synthetic. Examples of 5o-reductase inhibitors which may be included in the formulation include wedelolactone, B-sitosterol, Carthamus tinctorius extract, Saw palmetto extract, pumpkin seed extract, stinging nettle extract, Pygeum bark extract, Phyllanthus niruri extract, Phyllanthus embilica extract, lion's mane mushroom extract, azelaic acid, curcumin, gamma-Linolenic acid, o-linolenic acid, linoleic acid, palmitoleic acid, oleic acid, myristoleic acid, green tea catechins including (-)-epigallo-catechin-3-gallate (EGCG), riboflavin (vitamin B2), or any other suitable agent. For instance, in some embodiments, the formulation may comprise a combination of wedelolactone and / or B-sitosterol and saw palmetto and / or C. tinctorius. In yet other embodiments, the formulation comprises any combination selected from the group E. alba extract, C. tinctorius extract, Saw Palmetto extract, P. embilica extract and P. Niruri extract.

[0033] As used herein, the term "extract" refers to any preparation or substance obtained from a plant, herb, or other organic material, which concentrates the active or beneficial constituents of the source material. This includes, but is not limited to, oils, powders, tinctures, infusions, distillates, concentrates, suspensions, or any other form containing the active or beneficial properties of the original material. The extraction method may include any suitable technique known to those skilled in the art, such as cold pressing, solvent extraction, supercritical fluid extraction, steam distillation, maceration, or any other suitable method or combination of methods. The "extract" can be in pure form or it may be combined with other substances, carrier agents or additives, in a manner that does not materially alter the essential characteristics of the extract. In some cases, the extract may also be synthetically produced or obtained through biotechnological methods. Hence, the term "extract" as used in this document should be interpreted broadly to encompass any such preparations or substances derived or synthesised from the source material, whether in isolated, concentrated, diluted, solid, liquid, or gaseous forms.

[0034] In particular embodiments, the formulation comprises E. alba extract, Saw palmetto extract and N. sativa extract. The inventor has shown that this combination provides a surprisingly effective and synergistic hair growth formulation. In particular embodiments, the formulation comprises E. alba extract, Saw palmetto extract, N. sativa extract and P. niruri extract. In particular embodiments, the formulation comprises E. alba extract, Saw palmetto extract, N. sativa extract, P. niruri extract and C. tinctorius extract. These formulations provide a blend of phytochemicals with a spread of 5o-reductase activity action. In some embodiments, the formulation comprises a combination of all 5 of E. alba, C. tinctorius, Saw palmetto, P. embilica and P. niruri. The inventor believes that the combination of E. alba, C. tinctorius, Saw palmetto, P. embilica and P. niruri provides a blend of phytochemicals with a spread of 5o-reductase activity action over short, medium and long term. This provides a cumulative effect which significantly reduces hair follicle miniaturisation and extends the duration of anagen phase relative to the telogen phase. In some embodiments, the formulation comprises a combination of all 6 of E. alba, C. tinctorius, Saw palmetto, N. sativa, P. embilica and P. niruri. The inventor believes that the combination of E. alba, C. tinctorius, Saw palmetto, N. sativa, P. embilica and P. niruri provides a blend of phytochemicals with a spread of 5o-reductase activity action over short, medium and long term. This provides a cumulative effect which significantly reduces hair follicle miniaturisation and extends the duration of anagen phase relative to the telogen phase.

[0035] In some embodiments, the formulation comprises E. alba powder, Saw palmetto powder and N. sativa oil. In some embodiments, the formulation comprises E. alba powder, Saw palmetto powder, N. sativa oil and P. niruri powder. In some embodiments, the formulation comprises E. alba powder, Saw palmetto powder, N. sativa oil, P. niruri powder, and C. tinctorius oil.

[0036] Optionally, the formulation comprises vitamin C or a derivative thereof. Vitamin C and its derivatives are potent antioxidants and therefore serve to reduce oxidative damage to the hair follicle. In particular, the inventor believes that the combination of vitamin C and the one or more of thymoquinone, thymohydroquinone, dithymoquinone and thymol in the formulation may provide particular benefits by acting synergistically. Whilst each of vitamin C, thymoquinone, thymohydroquinone, dithymoquinone and thymol are potent antioxidants, each compound exerts its effect via different mechanisms of action. The addition of vitamin C in the formulation therefore provides protection against different types of oxidative stress. Compared to thymoquinone, thymohydroquinone, dithymoquinone and thymol, vitamin C has a much greater capacity to directly scavenge and neutralise reactive oxygen species (ROS), including superoxide anion radicals, hydroxyl radicals, and singlet oxygen. Further, whilst vitamin C is water-soluble and primarily acts to neutralise reactive oxygen species in the aqueous compartment of the cell, each of thymoquinone, thymohydroquinone, dithymoquinone and thymol are lipid soluble and may therefore be more active within the lipid compartments of the cell, such as the cell membrane or within organelles surrounded by lipid membranes. Combining vitamin C with the one or more of thymoquinone, thymohydroquinone, dithymoquinone and thymol, may therefore provide more comprehensive protection against oxidative damage.

[0037] Further, although thymoquinone and thymohydroquinone primarily behave as antioxidants, it has been observed that, under certain conditions, both compounds can exhibit pro-oxidant activity [7], [8]. Vitamin C is a potent antioxidant, and exhibits a property known as "redox cycling", which allows it to act as an antioxidant multiple times. The inventor believes that this property of vitamin C could potentially help to neutralise any pro-oxidant effects of the other compounds.

[0038] Optionally, the formulation comprises at least 1% (w / v) vitamin C, for example, in the range 1 to 5 % (w / v) vitamin C.

[0039] Optionally, the formulation comprises a phenolic compound selected from the group gallic acid, methyl gallate, chlorogenic acid, ellagic acid, caffeic acid, and ferulic acid.

[0040] These phenolic compounds are known for their potent antioxidant and anti-inflammatory properties. Further, certain phenolic compounds, including caffeic acid and ferulic acid, have been found to be pro-angiogenic. The inventor therefore recognises that the inclusion of a phenolic compound selected from the group gallic acid, methyl gallate, chlorogenic acid, caffeic acid, and ferulic acid in the formulation may stimulate blood circulation in the scalp and enhance the delivery of nutrients and oxygen to hair follicles [9, 10].

[0041] Optionally, the formulation comprises a tannin selected from the group embilicanin A, embilicanin B, punigluconin, pedunculagin, chebulagic acid, chebulinic acid, corilagin, trigallayl glucose. These tannins have anti-oxidant, anti-inflammatory and antimicrobial properties and can therefore contribute to maintaining a healthy microenvironment and reducing damage to the hair follicle. Further, tannins are known to have the ability to bind to and precipitate proteins. The inventor has recognised that this property of tannins could potentially strengthen the hair shaft, providing more resilience and less breakage.

[0042] Optionally, the formulation comprises one or more lignan compounds selected from the group phyllanthin, hypophyllanthin and niranthin. Lignans have antioxidant and antiinflammatory activity. In addition, lignans have been associated with antimicrobial effects against certain bacteria and fungi, therefore potentially helping to treat bacterial and fungal infections such as dandruff or seborrheic dermatitis which can contribute to hair loss by causing inflammation and damage to hair follicles.

[0043] Optionally, the formulation comprises one or more o-amino acids selected from the group glutamic acid, aspartic acid, alanine, lysine, proline and cysteine. Including these o-amino acids in the formulation helps to ensure an adequate supply for protein synthesis and hair growth.

[0044] Optionally, the formulation comprises one or more flavonoids. The flavonoids may comprise one or more anthoxanthins, optionally one or more flavones selected from the group apigenin, chrysin and luteolin; and / or one or more flavonols selected from the group astragalin, azalein, isoquercetin, kaempferol, isorhamnetin, quercetin, quericitin, myricetin, fisetin and rutin. Additionally, or alternatively, the flavonoids may comprise one or more flavan-3-ols selected from the group catechin (C), catechin 3-gallate (CG), epicatechins (EC), epigallocatechin (EGC) epicatechin 3-gallate (ECG), epigallcatechin 3- gallate (EGCG), gallocatechin (GC) and gallcatechin 3-gallate (GCG). Additionally, or alternatively, the flavonoids may comprise one or more coumestans selected from the group coumestrol, plicadin, wedelolactone, demethylwedelolactone strynolactone, isodemethylwedelolactone and demethylwedelolactone glucoside.

[0045] Flavonoids are known for their potent antioxidant and anti-inflammatory properties, which help to combat oxidative stress and damage to the hair follicle. In addition, some flavonoids have been found to regulate signalling cascades and influence growth factors which are important for hair follicle function and growth. For example, one study found that the flavonol quercetin promoted hair growth in mice by upregulating the HIF-lo signalling cascade in endothelial cells, in turn promoting angiogenesis around hair follicles and facilitating the transition from the resting phase to the active growth phase of the hair cycle

[0013] . Certain flavonoids are also believed to have 5o-reductase inhibitory activity. For instance, it has been suggested that the structure of myricetin, quercetin, and fisetin may allow them to inhibit 5o-reductase isoenzyme 1, while kaempferol has been shown to inhibit isoenzyme 2 in transfected Rat 1A cells

[0014] .

[0046] Optionally, the formulation comprises one or more of o-amyrin and B-amyrin. Certain studies have suggested that a- and B- amyrin have an inhibitory effect on the activity of 5-lipoxygenase, an enzyme that catalyses the conversion of arachidonic acid into leukotrienes

[0015] . Leukotrienes are lipid signalling molecules which have pro-inflammatory effects that can damage hair follicles. In addition, leukotrienes have been shown to inhibit the proliferation of human dermal papilla cells. By inhibiting the proliferation of these cells, leukotrienes can disrupt the hair growth cycle and lead to hair loss. The inventor has therefore recognised that the inclusion of a- and B- amyrin in the formulation may reduce the levels of leukotrienes in the scalp, thereby reducing damage to the hair follicles and acting to maintain a normal hair growth cycle.

[0047] Optionally, the formulation comprises one or more fatty acids selected from the group stearic acid, oleic acid, palmitic acid, myristic acid, linolenic acid, linoleic acid, arachidonic acid, eicosadienoic acid, almitoleic acid and lauric acid. Certain fatty acids, such as stearic acid, palmitic acid and linoleic acid have emollient properties that help moisturize and hydrate the hair and scalp. This can improve the hair's moisture retention, preventing dryness and promoting healthier hair growth. Further, certain fatty acids possess antiinflammatory properties that can alleviate scalp inflammation and support overall scalp health. It has also been suggested that certain fatty acids including lauric acid and myristic acid may inhibit the action of 5o-reductase, thus helping to reduce levels of DHT

[0016] .

[0048] Optionally, the formulation comprises non-trace amounts of minerals selected from the group magnesium, zinc, and iron. Zinc is involved in the regulation of various enzymes and hormones that are important for hair growth and maintenance and zinc deficiency has been associated with hair loss. It is also a potent antioxidant that helps protect the hair follicle from oxidative stress.

[0049] Like zinc, iron also plays a critical role in hair growth and is involved in the regulation of various enzymes and hormones that are important for hair growth and maintenance. Hair is extremely sensitive to iron deficiency and iron deficiency has been associated with a higher risk of hair loss and reduced hair density.

[0050] Magnesium is also involved in the regulation of various enzymes and hormones that are important for hair growth and maintenance. It is also a potent antioxidant that helps protect the hair follicle from oxidative stress. Further, magnesium plays an important role in the formation of keratin, the protein that makes up the hair shaft, and may help improve hair strength and resilience.

[0051] The aforementioned minerals may be present in the formulation as salts, chelates, or complexes, which have been shown to be absorbed and utilised by the body. These forms may include, but are not limited to, any one or more of ferrous sulfate, ferrous gluconate, ferric ammonium citrate, magnesium citrate, magnesium glycinate, zinc gluconate and zinc citrate.

[0052] Optionally, the formulation comprises o-tocopherol. o-tocopherol is a potent lipophilic antioxidant which can integrate into the lipid bilayer of cells, including those of the hair follicles and the cuticle layer of the hair shaft, protecting these structures from lipid peroxidation caused by reactive oxygen species (ROS). By neutralizing ROS, o-tocopherol mitigates oxidative stress, helping to repair cellular damage and maintain the integrity and function of the hair follicles and shaft.

[0053] Optionally, the formulation comprises an extract of Eclipta alba. Major constituents found in E. alba extract include wedelolactone, the flavonoids luteolin and apigenin, the phytosterols B-sitosterol and stigmasterol. As a result, the extract of E. alba provides potent anti-oxidant and anti-inflammatory action mediated via a variety of mechanisms. Further, it has been suggested that E. alba may inhibit the TGF-B 1 signalling pathway in hair follicles, thus reducing TGF-B 1-induced apoptosis in human dermal papilla cells

[0017] . In addition, some studies suggest that E. alba may have 5o-reductase inhibitory activity

[0028] .

[0054] Optionally, the formulation comprises an extract of Phyllanthus embilica. Phytochemicals found in P. embilica extract include the flavonoids quercetin and kaempferol; phenolic compounds including gallic acid, ellagic acid methyl gallate; tannins including emblicanin A and B, phyllaemblicin B, punigluconin, pedunculagin, chebulinic acid, corilagin, geraniin; amino acids including glutamic acid, aspartic acid, alanine, lysine, proline and cysteine; fatty acids including stearic acid, oleic acid, palmitic acid, myristic acid, linolenic acid; and linoleic acid, alkaloids including phyllantine, phyllembein, phyllantidine, the lignan compounds phyllanthin, hypophyllanthin and niranthin. In addition, P. embilica has one of the highest vitamin C contents of any natural source, and also has a high content of minerals including calcium, iron, phosphorus, potassium, zinc, copper, manganese and magnesium.

[0055] Accordingly, P. embilica extract is rich in various nutrients and bioactive compounds that may contribute to hair growth and maintenance. In particular, the inventor has recognised that the high vitamin C content of P. embilica can provide powerful protection against oxidative damage to the hair follicle. The high zinc and iron content of P. embilica may also contribute to the regulation of hormones and enzymes involved in hair growth and maintenance, as described hereinabove. P. embilica has also been reported to act as a 5o- reductase inhibitor and thus may reduce DHT levels when included in the formulation

[0018] .

[0056] Optionally, the formulation comprises a Nigella sativa seed extract, for example, N. sativa oil. Key constituents of N. sativa seed are: fatty acids including linoleic acid, linolenic acid, arachidonic acid, eicosadienoic acid, oleic acid, palmitic acid, stearic acid and myristic acid; sterols including B-sitosterol, campesterol, and stigmasterol; alkaloids, including nigellidine and nigellicine; volatile oils including thymoquinone, thymohydroquinone, dithymoquinone, nigellone, p-cymene; and minerals including calcium, phosphorous, potassium and iron.

[0057] Optionally, the formulation comprises an extract of Saw palmetto. Saw palmetto is primarily comprised of fatty acids including lauric acid and myristic acid, phytosterols including B-sitosterol, B-carotene, vitamin E derivatives, and polysaccharides

[0019] . Saw palmetto has also been reported to be an inhibitor of both 5o-reductase isoforms [20, 21]. Further, studies have shown that stimulates hair regrowth in murine models via TGF-B and mitochondrial signalling pathways

[0023] . Additionally, in vitro models have demonstrated SP's ability to inhibit inflammatory gene expression in human keratinocytes

[0024] .

[0058] Optionally, the formulation comprises an extract of Carthamus tinctorius. C. tinctorius extract has been found to inhibit 5o-reductase from rat livers

[0018] . Therefore, the inventor has recognised that the inclusion of C. tinctorius in the formulation may stimulate hair growth by reducing androgen activity in the scalp. C. tinctorius also contains a high concentration of fatty acids such as linoleic acid and oleic acid which may have moisturising and nourishing effects on the scalp and hair. C. tinctorius extract also contains a number of phenolic acids chlorogenic acid, ferulic acid and caffeic acid have been suggested to have antioxidant and anti-inflammatory properties that may help to protect the hair follicle from oxidative stress and inflammation.

[0059] Optionally, the formulation comprises linalool. Linalool has potent anti-inflammatory and anti-oxidant properties, which the inventor has recognised can reduce damage to the hair follicle. In addition, linalool has vasodilatory properties. Therefore, the inventor has recognised that linalool may improve circulation to the scalp to help promote hair growth by delivering more oxygen and nutrients to the hair follicles.

[0060] Optionally, the linalool is provided by an extract of a plant selected from the group coriander, rose and lavender.

[0061] Optionally, the formulation comprises linalyl acetate. Linalyl acetate has anti-inflammatory and antioxidant effects and may also increase blood circulation in the scalp.

[0062] Optionally, the formulation comprises Lavandula extract, for example lavender oil. Lavender oil contains linalool and linalyl. In addition, lavender oil may act to increase the expression of growth factors including VEGF and IGF-1 in hair follicles

[0025] .

[0063] Optionally, the formulation comprises Phyllanthus niruri extract. Major constituents of P. niruri are lignans including phyllanthin, hypophyllanthin and niranthin; phenolic compounds including gallic acid and ellagic acid; flavonoids including quercetin, luteolin, and apigenin; tannins including corilagin; and alkaloids including phyllanthine, hypophyllanthine, and nirurine. Further, one study has reported that petroleum ether extract of P. niruri inhibited the activity of type-2 5o-reductase in vitro

[0026] . Therefore, the inventor believes the inclusion of P. niruri in the formulation to be effective in treating alopecia, for example traction alopecia, in particular, by reducing DHT levels in the scalp. Optionally, the formulation comprises a penetrant enhancer. The term "penetrant enhancer" herein refers to an ingredient that increases the absorption of the active ingredients through the skin, in particular through the stratum corneum into the deeper layers of the epidermis. In particular, the penetrant enhancer may increase the amount of active ingredients which reach the hair follicles. The inventor recognises that the use of a penetrant enhancer may improve the efficacy of the hair loss tonic and enhance its ability to stimulate hair growth. Any suitable penetrant enhancer may be used, including surfactants, solvents, and fatty acids. Examples of penetrant enhancers which may be used in formulations according to the invention include propylene glycol, ethanol, dimethyl sulfoxide (DMSO) and menthol.

[0064] Optionally, the formulation comprises one or more of an extract of Gingko biloba, an extract of Panax ginseng and Peppermint oil.

[0065] Gingko biloba is known for its vasodilating properties. The inventor has therefore recognised that the inclusion of Gingko biloba in the formulation may increase blood flow in the scalp and thereby increase the flow of oxygen and nutrients to the hair follicles. Gingko biloba also has potent antioxidant and anti-inflammatory effects. In addition, some evidence suggests that Gingko biloba may increase VEGF and HGF levels and decrease inflammatory factors such as TNF-o, thereby potentially stimulating hair growth and / or regrowth

[0027] .

[0066] Panax ginseng also has antioxidant and anti-inflammatory properties. Further, some research suggests that Panax ginseng can have an anti-apoptotic activity

[0029] . Apoptosis plays a critical role in the transition from the anagen phase to the catagen phase. In the catagen phase, a significant portion of the hair follicle undergoes apoptosis, leading to a regression of the follicle. Therefore, the inventor believes that including Panax ginseng in the formulation may extend the anagen phase and thereby reduce hair loss and / or increase hair growth.

[0067] Peppermint oil may provide the formulation with menthol which may act as a penetrant enhancer to increase the amount of active ingredients in the formulation which reach the hair follicles. Peppermint oil may also provide the formulation with antibacterial and / or antifungal compounds including any one or more of menthone, menthyl acetate and limonene. Optionally, the formulation comprises a thickening agent. Suitable thickening agents include any dermatologically-acceptable thickening agent, such as any one or more of cetyl alcohol, stearyl alcohol, carnauba wax, stearic acid or castor oil. The term "dermatologically acceptable" herein means having no undue toxicity, incompatibility, instability, and the like.

[0068] Optionally, the formulation has a selected viscosity to make it suitable for topical application. Specifically, the formulation may have a selected viscosity to avoid significant dripping or pooling after application to skin. The term "topical application" herein refers to applying or spreading the formulation onto the surface of keratinous tissue.

[0069] Optionally, the formulation comprises carriers prepared according to conventional techniques using, conventional ingredients and agents which are well known in the industry. Adjuvants and other agents which may also be dispersed in the carrier include the following: moisturizers, humectants, anti-dusting agents, emulsifiers, and selected amino acids. Further, the carrier is not particularly limited. Specific examples include, but are not limited to, ointments such as hydrophilic vaseline or polyethylene glycol ointment; pastes such as gum (e.g., xanthan gum); solutions such as alcoholic, aqueous or buffer solution; gels such as aluminum hydroxide or sodium alginate gel; albumins such as human or animal albumin; collagens such as human or animal collagen; celluloses such as alkyl cellulose, hydroxyalkyl cellulose or alkyl hydroxyalkyl cellulose; methyl cellulose, hydroxyethyl cellulose, carboxymethyl cellulose, hydroxypropylmethyl cellulose and hydroxypropyl cellulose; polymers such as Pluronic™ polyol as exemplified by Pluronic F- 127; tetronics such as Tetronic 1508; and alginates such as sodium alginate.

[0070] The method of treatment for hair loss comprises applying a formulation according to any one of the preceding claims to a treatment area to stimulate hair growth in the treatment area and / or reduce hair loss in the treatment area. The term treatment area as it used herein, refers to an area of mammalian skin affected by hair loss and / or reduced hair growth or regrowth. References to the formulation being applied to a treatment area herein include both topical application of the formulation to the undisrupted skin surface as well as delivery of the formula trans- dermally / epidermally. For example, in some embodiments, the formulation may be administered to part or all of the epidermis, or deeper into the papillary dermis, reticular dermis, and / or hypodermis, during or after mechanical disruption of the epidermal barrier, using electroporation, RF ablation, laser- poration, laser ablation (fractional or non-fractional), skin abrasion, exfolitation (eg. a prepeel), micro-needling or any other suitable method. In some embodiments the formulations disclosed herein may be prepared with carriers that deliver the active ingredients to a target site, for example, the hair follicle. The carriers may be liposomes, micelles, or other suitable vesicles.

[0071] The treatment area may refer to any area of mammalian skin. For instance, in some embodiments of the method, the treatment area may be located on the scalp. In other embodiments of the method the treatment area may be located on the eyebrows.

[0072] Surprisingly, the inventor has found that hair growth has been stimulated in scalp scars when the formulation is applied to the scalp scar. This finding is remarkable because scar tissue does not contain active hair follicles because the hair follicles may be damaged or destroyed. As such, the fact that this hair loss tonic is able to stimulate hair growth in scalp scars suggests that formulations according to the invention can activate dormant hair follicles or even generate new ones.

[0073] Optionally the method comprises applying the formulation to the treatment area at least twice weekly, optionally at least 3 times weekly, optionally at least 4 times weekly, optionally at least 5 times weekly, optionally at least 6 times weekly, optionally at least once daily. In some embodiments, the method comprises applying the formulation regularly for a period of at least 5 weeks, optionally at least 6 weeks, optionally at least 7 weeks, optionally at least 8 weeks, optionally at least 10 weeks, optionally at least 11 weeks, optionally at least 12 weeks.

[0074] Other objects, features and advantages of the present invention will become apparent from the following detailed description. It should be understood, however, that the detailed description and the examples, while indicating specific embodiments of the invention, are given by way of illustration only. Additionally, it is contemplated that changes and modifications within the spirit and scope of the invention will become apparent to those skilled in the art from this detailed description.

[0075] Brief Description of the Figures

[0076] The invention will now be described in detail, by way of example only, with reference to the accompanying figures, in which:

[0077] Figure 1 shows the effects after 20 weeks of the 3 times a week times weekly application of the formulation according to the invention to the scalp of a 40 to 50 year old man with TE and AGA (Figure 1A shows hair density before treatment and Figure IB shows hair density after 20 weeks of treatment); Figure 2 shows the effects after 12 weeks of the 3 times weekly application of the formulation according to the invention to the scalp of a 45 to 55 year old man with AGA (Figure 2A shows hair density before treatment and Figure 2B shows hair density after 12 weeks of treatment);

[0078] Figure 3 shows the effects after 16 weeks of the twice weekly application of the formulation according to the invention to the scalp of a 30 to 40 year old woman with traction alopecia (Figure 3A shows hair density before treatment and Figure 3B shows hair density after 16 weeks of treatment);

[0079] Figure 4 shows the effects after 20 weeks of the 3 times weekly application of the formulation according to the invention to the scalp of a 45 to 55 year old man with AGA (Figure 4A shows hair density before treatment and Figure 4B shows hair density after 20 weeks of treatment);

[0080] Figure 5 shows the effects after 16 weeks of the 2 times weekly application of the formulation according to the invention to the scalp of a 55 to 65 year old woman with traction alopecia (Figure 5A shows hair density before treatment and Figure 5B shows hair density after 16 weeks of treatment); and

[0081] Figure 6 shows the effects after 12 weeks and after 24 weeks of the 3 times weekly application of the formulation according to the invention to the scalp of a 40 to 50 year old man with AGA (Figure 6A shows hair density before treatment, Figure 6B shows hair density after 12 weeks of treatment and Figure 6C shows hair density after 24 weeks of treatment);

[0082] Figure 7 shows the results for the subject of figures 6A to 6C 14 months after ceasing application; and

[0083] Figure 8 shows the effects after 16 weeks and after 56 weeks of the twice weekly application of the formulation according to the invention to the scalp of a 45 to 55 man with a scalp scar (Figure 8A shows hair density before treatment, Figure 8B shows hair density after 16 weeks of treatment and Figure 8C shows hair density after 56 weeks of treatment).

[0084] Figure 9 shows a 47 year old female with traction alopecia and diffuse androgenic thinning treated with a formulation of Bhringraj and castor oil as a carrier. Figure 9A shows hair density before treatment, Figure 9B shows hair density after 40 applications of treatment over a 4 month period. The red dotted lines denote the border of the widows peak where hair growth is observed.

[0085] Figure 10 shows a 46 year old man with androgenic and traction hair loss treated with a formulation of Saw Palmetto and castor oil as a carrier. Figure 10A shows hair density before treatment, Figure 10B shows hair density after 9 weeks of treatment.

[0086] Figure 11 shows a 68 year old man with androgenic hair loss treated with a formulation of black seed oil and castor oil as a carrier. Figure 11A shows hair density before treatment, Figure 11B shows hair density after 9 months of treatment. The red dot in each image denotes the site of a mole on the hairline, above which sparse new hair growth is observed.

[0087] Figure 12 shows a man with atypical hair loss treated with a formulation of Bhringraj, Saw Palmetto, black seed oil and castor oil as a carrier. Figure 12A shows hair density before treatment, Figure 12B shows hair density after treatment after less than one month of treatment.

[0088] Example 1

[0089] A. Ingredients

[0090] A formulation was prepared according to the below quantities.

[0091] B. Methodology

[0092] The formulation according to Example 1 was prepared by mixing the castor oil, the black seed oil, the safflower oil and the amla oil. The powdered ingredients (Bhringraj powder, Phyllanthus niruri powder and Amla powder) and the lavender oil were then added and allowed to infuse for 2 weeks with daily shaking to homogenise the mixture.

[0093] While the format (e.g., oil, powder) of each ingredient is indicated for the exemplified formulation, the skilled person will appreciate that other formats of the ingredients are encompassed by the present invention (for example, Saw palmetto extract may be provided as an oil rather than a powder).

[0094] A 120 mL bottle of the formulation was administered to 20 trial subjects who applied the formulation topically to the scalp 2 to 3 times weekly.

[0095] C. Results

[0096] Of the 20 trial subjects, 18 reported a noticeable increase in hair density. The remaining two subjects did not participate in the follow-up process. A number of the subjects reported a mild shedding phase occurring after around 4 to 5 weeks of application, followed by growth of non-vellus hair. Critically, trial subjects that ceased use of the formulation did not induce hair loss suggesting that the hair stimulated by the formulation possessed greater resistance to DHT and Telogen Effluvium. Only 1 subject experienced side effects: this subject had pale skin type 1 skin experienced pigmentary melanocyte stimulation. Figures 1 to 8 show the effects of the formulation according to Example 1 in a sample of the trial subjects after various periods. It can be seen that there is a marked increase in hair density for all of the subjects depicted.

[0097] Figure 6A to 6C show the effects after 12 weeks and after 24 weeks of the 3 times weekly application of the formulation according to Example 1 to the scalp of a 40 to 50 year old man with AGA. Figure 7 shows the results for the same subject 14 months after ceasing application. It can be seen that the increased hair density achieved after 24 weeks of treatment is sustained over the long term, even in the absence of continued treatment.

[0098] Referring in particular to Figures 8A to 8C, it was found that the application of the formulation according to the above methodology stimulated hair growth in scar tissue on the subject's scalp. Figure 8A shows the subject's scalp prior to the treatment. It can be seen that the subject has a scar extending transversely across the occipital region of the scalp. The scar was caused by follicular unit transplant (FUT) performed 10 years prior to the beginning of treatment with the formulation according to Example 1. Therefore, in this case the scar tissue was established and can be assumed to contain no active hair follicles.

[0099] Figure 8B shows the subject's scalp after 16 weeks of treatment. It can be seen that there is a noticeable increase in hair density over all regions of the scalp. In particular, it can be seen that new hair growth has been stimulated in the scar tissue. Therefore, it can be inferred that treatment with the formulation according to Example 1 has either reactivated hair follicles which were present in the scar tissue, but which were dormant due to damage, or has stimulated generation of new hair follicles.

[0100] Figure 8C shows the subject's scalp after 56 weeks of treatment. It can be seen that there is a further noticeable increase in hair density over all regions of the scalp compared to Figure 8B such that hair now appears uniformly dense.

[0101] Surprisingly, it was found that application of the formulation according to Example 1 stimulated pigmentation in hair follicles in some subjects presenting with grey hair. These subjects exhibited regrowth of black hairs after undergoing treatment. This points to the formulation's potential ability to influence melanin production, a factor contributing to hair colour.

[0102] Further surprisingly, the formulation according to Example 1 also showed positive effects when tested on a horse. This demonstrates that the formulation has broader applicability across various mammalian species. Example 2

[0103] To test the synergy of the invention, three patients were treated with a formulation containing a different active ingredient (Bhringraj, Saw Palmetto, or black seed oil) in a carrier of castor oil, as follows:

[0104] A 47 year old female patient with traction alopecia and diffuse androgenic thinning was treated with a formulation of Bhringraj and castor oil as a carrier for four months (see Figure 9).

[0105] A 46 year old man with androgenic and traction hair loss was treated with a formulation of Saw Palmetto and castor oil as a carrier for 9 weeks (see Figure 10).

[0106] A 68 year old man with androgenic hair loss was treated with a formulation of black seed oil and castor oil as a carrier for 9 months (see Figure 11).

[0107] A further patient was treated with a formulation containing Bhringraj, Saw Palmetto, and black seed oil in a carrier of castor oil (see Figure 12). While the three patients treated with a single ingredient only displayed sparse hair growth after treatment (Figures 9-11), the patient treated with the combination of ingredients displayed significant hair growth after treatment, showing a clear synergistic effect of the ingredients. The inventor believes that the combination of ingredients provides an immunomodulatory effect, working at stem cell level in the treatment area to stimulate hair growth.

[0108] In view of the many possible embodiments to which the principles of the disclosed invention may be applied, it should be recognised that the illustrated embodiments are only preferred examples of the invention and should not be taken as limiting the scope of the invention. Rather, the scope of the invention is defined by the following claims.

[0109] References:

[0110] 1. Truong, V.L., Bak, MJ., Lee, C., Jun, M., Jeong, W.S. (2017). Hair Regenerative Mechanisms of Red Ginseng Oil and Its Major Components in the Testosterone-Induced Delay of Anagen Entry in C57BL / 6 Mice. Molecules, 22, 1505. 2. Pandit, S., Chauhan, N. S., & Dixit, V. K. (2009). Effect of Wedelolactone on In Vitro 5o-Reductase Inhibition and Hair Growth Promotion in Male Wistar Rats. Journal of Ethnopharmacology, 123(3), 423-427.

[0111] 3. Cabeza M, Bratoef E, Heuze I, Ramirez E, Sanchez M, Flores E. Effect of betasitosterol as inhibitor of 5alpha-reductase in hamster prostate. Proceedings- Western Pharmacology Society; 2003. p. 153-5.

[0112] 5. Moon, EJ., Lee, Y.M., Lee, OH. et al. (1999). A novel angiogenic factor derived from Aloe vera gel: 0-sitosterol, a plant sterol. Angiogenesis, 3, 117-123.

[0113] 6. Taha, M., A. Z. Azeiz, & W. Saudi. (2010). Antifungal Effect Of Thymol, Thymoquinone And Thymohydroquinone Against Yeasts, Dermatophytes And NonDermatophyte Molds Isolated From Skin And Nails Fungal Infections. Egyptian Journal of Biochemistry & Molecular Biology, 28(2).

[0114] 7. Elsharkawy, E. R., Abdallah, E. M., & Markb, A. A. (2021). Potential Cytotoxic, Antifungal, and Antioxidant Activity of Dithymoquinone and Thymoquinone. Journal of Hunan University Natural Sciences, 48(9).

[0115] 8. Mizuno, M., & Fukuhara, K. (2022). Antioxidant and Prooxidant Effects of Thymoquinone and Its Hydroquinone Metabolite. Biological and Pharmaceutical Bulletin, 45(9), 1389-1393.

[0116] 9. Lin C.M., Chiu J.H., Wu I.H., Wang B.W., Pan C.M., Chen Y.H. (2010). Ferulic acid augments angiogenesis via VEGF, PDGF and HIF-1 A. Journal of Nutritional Biochemistry, 21, 627-633.

[0117] 10. Vuthijumnonk J., Heyes J., & Molan A. (2016). Total anthocyanins, chlorogenic acid concentration, antioxidant and in ovo anti-angiogenic activities of rabbiteye blueberries. International Food Research Journal, 23, 515.

[0118] 13. Zhao, Q., Zheng, Y., Zhao, D., Zhao, L., Geng, L., Ma, S., Cai, Y., Liu, C., Yan, Y., Belmonte, J. C. I., Wang, S., Zhang, W., Liu, G.-H., Qu, J., (2002). Single-cell profiling reveals a potent role of quercetin in promoting hair regeneration. Protein 8c Cell. 14. Hiipakka RA, Zhang HZ, Dai W, et al. (2002). Structure-activity relationships for inhibition of human 5o-reductases by polyphenols. Biochemical Pharmacology, 63(6), 1165-1176.

[0119] 15. Kweifio-Okai, G., & Macrides, T. A. (1992). Antilipoxygenase activity of amyrin triterpenes. Research Communications in Chemical Pathology and Pharmacology, 78(3), 367-372.

[0120] 16. Veeresh B, Patil AA, & Warke YB. (2016). Combination of lauric acid and myristic acid prevents benign prostatic hyperplasia symptoms in animal model. African Journal of Pharmacy and Pharmacology, 10(8), 101-106.

[0121] 17. Begum S, Lee MR, Gu LJ, Hossain J, Sung CK. (2015). Exogenous stimulation with Eclipta alba promotes hair matrix keratinocyte proliferation and downregulates TGF-pi expression in nude mice. International Journal of Molecular Medicine, 35(2), 496-502.

[0122] 18. Kumar, N., Rungseevijitprapa, W., Narkkhong, N., Suttajit, M., & Chaiyasut, C. (2012). 5o-reductase inhibition and hair growth promotion of some Thai plants traditionally used for hair treatment. Journal of Ethnopharmacology, 139(3), 765-771.

[0123] 19. Evron E, Juhasz M, Babadjouni A, & Mesinkovska NA. (2020). Natural Hair Supplement: Friend or Foe? Saw Palmetto, a Systematic Review in Alopecia. Skin Appendage Disorders, 6(6), 329-337.

[0124] 20. lehle C, Delos S, Guirou O, Tate R, Raynaud JP, & Martin PM. (1995). Human prostatic steroid 5o-reductase isoforms - a comparative study of selective inhibitors. Journal of Steroid Biochemistry and Molecular Biology, 54(5-6), 273-9.

[0125] 21. Sultan C, Terraza A, Devillier C, Carilla E, Briley M, Loire C, et al. (1984). Inhibition of androgen metabolism and binding by a liposterolic extract of "Serenoa repens B" in human foreskin fibroblasts. Journal of Steroid Biochemistry, 20(1), 515-9.

[0126] 23. Zhu HL, Gao YH, Yang JQ, Li JB, & Gao J. (2018). Serenoa repens extracts promote hair regeneration and repair of hair loss mouse models by activating TGF-p and mitochondrial signalling pathway. European Review for Medical and Pharmacological Sciences, 22(12), 4000-8. 24. Chittur S, Parr B, & Marcovici G. (2011). Inhibition of inflammatory gene expression in keratinocytes using a composition containing carnitine, thioctic acid and saw palmetto extract. Evidence-Based Complementary and Alternative Medicine, 2011, 985345.

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[0129] 28. Datta, K.; Singh, A.T.; Mukherjee, A.; Bhat, B.; Ramesh, B.; Burman, A.C. Eclipta alba extract with potential for hair growth promoting activity. J. Ethnopharmacol. 2009, 124, 450-456.

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Claims

Claims1. A formulation for treating hair loss, the formulation comprising: one or more of wedelolactone; demethylwedelolactone; and demethylwedelolactone glucoside;B-sitosterol; and one or more of thymoquinone, thymohydroquinone, dithymoquinone and thymol.

2. A formulation according to claim 1, wherein the formulation includes at least two different 5a-reductase inhibitors.

3. A formulation according to any preceding claim, wherein the formulation comprises vitamin C or a derivative thereof.

4. A formulation according to any preceding claim, wherein the formulation comprises one or more flavonoids selected from the group quercetin, kaempferol apigenin and luteolin.

5. A formulation according to any preceding claim, wherein the formulation comprises one or more triterpenes selected from the group: one or more of o-amyrin and B-amyrin.

6. A formulation according to any preceding claim, wherein the formulation comprises one or more fatty acids selected from the group stearic acid, oleic acid, palmitic acid, myristic acid, linolenic acid, linoleic acid, arachidonic acid, eicosadienoic acid, almitoleic acid, lauric acid.

7. A formulation according to any preceding claim, wherein the formulation comprises non-trace amounts of one or more of magnesium, zinc, and iron.

8. A formulation according to any preceding claim, wherein the formulation comprises Eclipta alba extract.

9. A formulation according to any preceding claim, wherein the formulation comprises Phyllanthus embilica extract.

10. A formulation according to any preceding claim, wherein the formulation comprises Nigella sativa seed extract.26SUBSTITUTE SHEET (RULE 26)11. A formulation according to any preceding claim, wherein the formulation comprises Saw palmetto extract.

12. A formulation according to any preceding claim, wherein the formulation comprises an extract of Cartham us tinctorius extract.

13. A formulation according to any preceding claim, wherein the formulation comprises lavender oil.

14. A formulation according to any preceding claim, wherein the formulation comprises Phyllanthus niruri extract.

15. A formulation according to any preceding claim, wherein the formulation comprises one or more of an extract of Gingko biloba extract, Panax ginseng extract and Peppermint oil.

16. A method of treatment for hair loss, the method comprising applying a formulation according to any one of the preceding claims to a treatment area to stimulate hair growth in the treatment area and / or reduce hair loss in the treatment area.

17. A method of treatment for hair loss according to claim 16, wherein the method is a method for regenerating hair in skin scars, further wherein the method comprises applying the formulation to skin scar tissue to stimulate hair growth in the skin scar tissue.

18. A method of treatment for hair loss according to claim 16, wherein the method comprises applying the formulation to the treatment area at least twice weekly.

19. A cosmetic method for reducing hair loss or stimulating hair growth or regrowth, the method comprising applying a formulation according to any one of claims 1 to 15 to a treatment area to stimulate hair growth in the treatment area.

20. A cosmetic method for stimulating pigmentation in hair follicles, the method comprising applying a formulation according to any one of claims 1 to 15 to stimulate pigmentation in hair follicles in the treatment area.27SUBSTITUTE SHEET (RULE 26)

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