Hair care compositions containing extracellular metabolite preparation from bacillus coagulans

Abstract

Disclosed are hair care compositions containing partially purified extracellular metabolite preparation from strains of Bacillus coagulans. Specifically, the uses of compositions containing extracellular metabolite preparation from a strain of Bacillus coagulans for increasing hair growth, inhibition of 5α-reductase and proliferation of follicle dermal papilla cells and in the management of androgenic alopecia, are disclosed.

Description

HAIR CARE COMPOSITIONS CONTAINING EXTRACELLULAR METABOLITE PREPARATION FROM BACILLUS COAGULANS CROSS-REFERENCE TO RELATED APPLICATIONS This is a PCT application claiming priority from US provisional application no. 62516077 filed on 6<com:Sup>th< / com:Sup> June 2017 BACKGROUND OF THE INVENTION Field of the invention [PARA 001] The invention in general relates to hair care compositions. More specifically the invention relates to hair care composition containing partially purified extracellular metabolite preparation from Bacillus coagulans. Description of prior art [PARA 002] Hair is one of the fastest growing tissues of the human body. Hair follicles undergo repetitive regenerative cycles consisting of three stages: anagen (rapid growth, active stage), catagen (apoptosis-driven regression, physiological involution stage), and telogen (resting stage). Reduction in anagen stage can result in miniaturization of hair follicles and hair loss. Thus extending the anagen phase is an effective treatment for the prevention of hair loss. [PARA 003] Growth and proliferation of follicle papilla cells are important for increasing the number of hair follicles and thereby to promote hair growth. Dermal papilla cells accumulate below undifferentiated epidermis which is then stimulated to form the hair "peg". Interaction between the hair peg and the dermal papilla cells promotes differentiation into a mature hair follicle. During the anagen phase the dermal papilla cells secrete several growth factors, like insulin like growth factor (IGF-1), fibroblast growth factor (FGF-7) which stimulates the proliferation and elongation of hair shaft. Hence, the dermal papilla has to be present in good numbers to ensure sufficient hair growth. Certain clinical conditions like androgenic alopecia, seborrhoeic dermatitis (dandruff) etc reduce the number of dermal papilla and increase hair fall. TGF-β2 is specifically expressed in human dermal papilla cells. TGF-β has been shown to exert unique multidirectional effect by inducing premature hair follicle regression in adult hair growth SUBSTITUTE SHEET (RULE 26) cycle and is required for hair folliculogenesis. The role of TGF- β is well documented in the following references: 1. Foitzik K, Paus R, Doetschman T, et al. The TGF-β2 isoform is both a required and sufficient inducer of murine hair follicle morphogenesis. Dev Biol. 1999; 212:278–89 2. Hibino T, Nishiyama T. Role of TGF-β2 in the human hair cycle. J Dermatol Sci.2004; 35: 9-18. 3. Jamora C, Lee P, Kocieniewski P, et al. A signaling pathway involving TGF-\(\beta\)2 and snail in hair follicle morphogenesis. PLoS Biol.2004; 3: e11. 4. Keita Inoue, Noriyuki Aoi, Yuji Yamauchi, Takahiro Sato, Hirotaka Suga, Hitomi Eto, Harunosuke Kato, Yasuhiko Tabata, Kotaro Yoshimura. TGF-β2 is specifically expressed in human dermal papilla cells and modulates hair folliculogenesis J. Cell. Mol. Med. Vol 13, No 11-12, 2009 pp. 4643-4656 [PARA 004] Vascular endothelial growth factor (VEGF) is essential for angiogenesis and vascular permeability and is responsible for maintaining proper vasculature around the hair follicle during the anagen growth phase. VEGF mRNA is strongly expressed in dermal papilla cells (DPC) in the anagen phase, but during the catagen and telogen phases. VEGF mRNA is less strongly expressed (Kozlowska et al., Expression of vascular endothelial growth factor (VEGF) in various compartments of the human hair follicle Arch Dermatol Res (1998) 290 :661–668; Yano et al., Control of hair growth and follicle size by VEGF-mediated angiogenesis J Clin Invest. 2001;107(4):409-417). Both VEGF and TGF-β2 promote the proliferation of dermal papilla cells and thereby increase hair growth. [PARA 005] The dermal papilla also produces an enzyme called <com:Math><semantics>5α<annotation encoding="application / x-tex">5\alpha< / annotation>< / semantics>< / com:Math>-reductase (3-oxo-<com:Math><semantics>5α<annotation encoding="application / x-tex">5\alpha< / annotation>< / semantics>< / com:Math>- steroid-delta 4-dehydrogenase;), a NADPH-dependent enzyme, which converts Testosterone to dihydrotestosterone (DHT). Overproduction of <com:Math><semantics>5α<annotation encoding="application / x-tex">5\alpha< / annotation>< / semantics>< / com:Math>-reductase increases the conversion of testosterone to DHT in hair follicles, thereby disrupting the natural hair cycle leading to hair fall and development of androgenic alopecia. [PARA 006] Natural 5α-reductase inhibitors like mushrooms, green tea etc. slow down the production of DHT and prevent hair loss. The use of probiotic in hair care in now garnering attention owing to the fact that probiotics can alter the microbiome of the skin responsible for hair loss. US7374750, discloses a composition containing a probiotic organism for hair care. The beneficial effects of probiotics for hair growth and increasing follicle dermal papilla is also well documented by Levkovich et al. (2013); Probiotic Bacteria Induce a 'Glow of Health', PLoS One. 2013; 8(1): e53867. Probiotics and their extracellular products are also reported to exhibit excellent anti-fungal activity, making them a valuable addition individually or as an adjuvant in anti-dandruff compositions. [PARA 007] It is well known in the scientific art that biological effects of probiotics or products thereof are strain specific and cannot be generalised among genera, species and strains (Probiotics: In Depth / NCCIH, U.S. Department of Health and Human Services, National Institutes of Health). Hence, there exists a need to find a superior probiotic strain and its extracellular product which promotes the growth of dermal papilla cells and prevents hair loss by inhibiting the <com:Math><semantics>5α<annotation encoding="application / x-tex">5\alpha< / annotation>< / semantics>< / com:Math>-reductase. Also, the beneficial effects of extracellular product obtained from probiotics on hair growth remain to be validated. The present invention solves the above problem by disclosing the beneficial effect of partially purified extracellular metabolite preparation of Bacillus coagulans on dermal papilla proliferation. [PARA 008] It is the principle objective of the invention to disclose a composition containing the partially purified extracellular metabolite preparation of Bacillus coagulans to increase hair growth. [PARA 009] It is another objective of the invention to disclose a composition containing the partially purified extracellular metabolite preparation of Bacillus coagulans to inhibit the activity of 5α-reductase. [PARA 0010] It is yet another objective of the invention to disclose a composition containing the partially purified extracellular metabolite preparation of Bacillus coagulans for use in the therapeutic management of androgenic alopecia. DEPOSIT OF BIOLOGICAL MATERIAL [PARA 0011] The deposit of biological material Bacillus coagulans bearing accession number MTCC 5856, mentioned in the instant application has been made on 19<com:Sup>th< / com:Sup> September 2013 at Microbial Type SUBSTITUTE SHEET (RULE 26) Culture Collection & Gene Bank (MTCC), CSIR-Institute of Microbial Technology, Sector 39-A, Chandigarh – 160036, India. SUMMARY OF THE INVENTION [PARA 0012] The present invention discloses a hair care composition containing the partially purified extracellular metabolite preparation isolated from a strain of Bacillus coagulans. More specifically, the invention discloses the use of a composition containing the extracellular metabolite isolated from a strain of Bacillus coagulans for the enhancing hair growth by increasing the proliferation follicle dermal papilla cells and inhibition of <com:Math><semantics>5α<annotation encoding="application / x-tex">5\alpha< / annotation>< / semantics>< / com:Math>-reductase. BRIEF DESCRIPTION OF THE DRAWINGS [PARA 0013] Fig.1a is the graphical representation of the percentage increase in dermal papilla cell proliferation compared to control by partially purified extracellular metabolite preparation from Bacillus coagulans MTCC 5856. [PARA 0014] Fig.1b and 1c depict the dose dependent increase in the proliferation of dermal papilla cells in the presence of partially purified extracellular metabolite preparation from Bacillus coagulans MTCC 5856. [PARA 0015] Fig.2 is the graphical representation of the percentage increase in the expression of TGF-β2 in the dermal papilla cells in the presence of partially purified extracellular metabolite preparation from Bacillus coagulans MTCC 5856. [PARA 0016] Fig.3 is the graphical representation of the percentage increase in the expression of VEGF in the dermal papilla cells in the presence of partially purified extracellular metabolite preparation from Bacillus coagulans MTCC 5856. [PARA 0017] Fig.4a depicts the percentage inhibition of 5α-reductase by partially purified extracellular metabolite preparation from Bacillus coagulans MTCC 5856. [PARA 0018] Fig.4b is the graphical representation showing decrease in concentration of dihydrotestosterone due to the inhibition of <com:Math><semantics>5α<annotation encoding="application / x-tex">5\alpha< / annotation>< / semantics>< / com:Math>-reductase activity by partially purified extracellular metabolite preparation from Bacillus coagulans MTCC 5856. SUBSTITUTE SHEET (RULE 26) DESCRIPTION OF THE MOST PREFERRED EMBODIMENTS [PARA 0019] In a most preferred embodiment, the invention discloses a hair care composition containing the partially purified extracellular metabolite preparation from a strain of Bacillus coagulans formulated with pharmaceutically / cosmeceutically acceptable excipients, adjuvants, bases, diluents, carriers, conditioning agents and preservatives. In a related embodiment, the composition containing the partially purified extracellular metabolite preparation from a strain of Bacillus coagulans can be combined with and / or incorporated into formulations containing hair care ingredients. In another related embodiment, the hair care composition is administered topically in the form of creams, gels, lotions, shampoo, serum, oil, suspensions, emulsions, and compacts. In another embodiment, the strain of Bacillus coagulans is selected from the group consisting of Bacillus coagulans MTCC 5856, Bacillus coagulans ATCC 31284 and Bacillus coagulans ATCC 7050 [PARA 0020] In another related embodiment, the invention relates to a method of increasing hair growth in mammals, said method comprising steps of administering effective concentration of partially purified extracellular metabolite preparation from a strain of Bacillus coagulans to said mammals, to bring about increase in hair growth. In a related embodiment, the increase in hair growth is brought about by a) increasing the expression of secretory factors and b) increasing the proliferation of follicle dermal papilla cells. In another related embodiment, the secretory factors are selected from the group consisting of vascular endothelial growth factor (VEGF) and transforming growth factors – <com:Math><semantics>β<annotation encoding="application / x-tex">\beta< / annotation>< / semantics>< / com:Math> (TGF-<com:Math><semantics>β<annotation encoding="application / x-tex">\beta< / annotation>< / semantics>< / com:Math>). In another related embodiment, the effective concentration of the partially purified extracellular metabolite preparation is 0.01% v / v to 2.0% v / v of the total composition. In another embodiment, the strain of Bacillus coagulans is selected from the group consisting of Bacillus coagulans MTCC 5856, Bacillus coagulans ATCC 31284 and Bacillus coagulans ATCC 7050 [PARA 0021] In another preferred embodiment, the invention relates to a method of inhibiting <com:Math><semantics>5α<annotation encoding="application / x-tex">5\alpha< / annotation>< / semantics>< / com:Math>-reductase activity, said method comprising step of bringing into contact said follicle dermal papilla cells with effective concentration of partially purified extracellular metabolite preparation from a strain of Bacillus coagulans to bring about 5α-reductase inhibition in follicle dermal papilla cells. In a related embodiment, the effective concentration of the partially purified extracellular metabolite preparation in the composition is 0.01% v / v to 2.0% v / v of the total SUBSTITUTE SHEET (RULE 26) composition. In another embodiment, the strain of Bacillus coagulans is selected from the group consisting of Bacillus coagulans MTCC 5856, Bacillus coagulans ATCC 31284 and Bacillus coagulans ATCC 7050. [PARA 0022] In another preferred embodiment, the invention discloses a method for therapeutic management of androgenic alopecia in mammals, said method comprising steps administering an effective concentration of a composition containing partially purified extracellular metabolite preparation from a strain of Bacillus coagulans to mammals in need of such therapy. In a related embodiment, the composition is formulated with pharmaceutically / cosmeceutically acceptable excipients, adjuvants, bases, diluents, carriers, conditioning agents and preservatives and / or incorporated into formulations containing hair care ingredients and administered topically in the form of creams, gels, lotions, shampoo, serum, oil, suspensions, emulsions, and compacts. In a related embodiment, the effective concentration of the partially purified extracellular metabolite preparation in the composition is 0.01% v / v to 2.0% v / v of the total composition. In another embodiment, the strain of Bacillus coagulans is selected from the group consisting of Bacillus coagulans MTCC 5856, Bacillus coagulans ATCC 31284 and Bacillus coagulans ATCC 7050. In another related embodiment, the mammal is preferably human. [PARA 0023] In another preferred embodiment, the invention discloses a method for therapeutic management of seborrhoeic dermatitis in mammals, said method comprising steps administering an effective concentration of a composition containing partially purified extracellular metabolite preparation from a strain of Bacillus coagulans, along with standard anti-dandruff ingredients to mammals in need of such therapy. In a related embodiment, the composition is formulated with pharmaceutically / cosmeceutically acceptable excipients, adjuvants, bases, diluents, carriers, conditioning agents and preservatives and administered topically in the form of creams, gels, lotions, shampoo, serum, oil, suspensions, emulsions, and compacts. In another embodiment, the anti-dandruff ingredients are selected from the group consisting of, but not limited to, coleus oil, tea tree oil, clove oil, basil oil and extract, rosemary oil, neem extract, cedarwood oil, selenium sulphide, Zinc Pyrithione, Salicylic acid, Ketoconazole, Climbazole and Ciclopirox Olamine. In a related embodiment, the effective concentration of the partially purified extracellular metabolite preparation in the composition is 0.01% v / v to 2.0% v / v of the total composition. In another embodiment, the strain of Bacillus coagulans is selected from the group consisting of SUBSTITUTE SHEET (RULE 26) Bacillus coagulans MTCC 5856, Bacillus coagulans ATCC 31284 and Bacillus coagulans ATCC 7050. In another related embodiment, the mammal is preferably human. [PARA 0024] Specific illustrative examples enunciating the most preferred embodiments are included herein below [PARA 0025] EXAMPLE 1: Proliferation of dermal papilla [PARA 0026] Isolation of extracellular metabolite [PARA 0027] The extracellular metabolite from Bacillus coagulans MTCC 5856 was isolated as per the steps outlined in US9596861. The product is commercially available under the tradename LACTOSPORIN® (INCI: Bacillus ferment filtrate extract) from Sabinsa Corporation, USA. [PARA 0028] Materials and Method [PARA 0029] Cells: Dermal Papilla Cells (human HFDPC) were purchased from Promocell (Germany), and maintained as a monolayer culture in Fibroblast growth media ( Promocell, Germany) at 37 °C in a humidified 5% CO<com:Sub>2< / com:Sub> incubator. Dermal Papilla Cells were cultured in DMEM supplemented with 10% FBS. The confluent cultures were harvested by trypsinization and expanded during two more passages before they were used for the experiments. Medium and other culture components were renewed after 48–72 h. All cell cultures were maintained at 37°C in 95% air and 5% CO<com:Sub>2< / com:Sub> in a CO<com:Sub>2< / com:Sub> incubator. [PARA 0030] The Dermal Papilla Cells were used for the assay at a seeding density of 2000 cells per well of 24 -well clear bottom microplate in DMEM and incubated at 37°C in 95% air and 5% CO<com:Sub>2< / com:Sub> in a CO<com:Sub>2< / com:Sub> incubator for 24 hours. The cell monolayers were fixed with 50% (w / v) trichloroacetic acid for 1hour at 4°C and stained with 0.4% SRB for 30 min. The excess dye was removed by washing repeatedly with 1% (v / v) acetic acid. The protein-bound dye is dissolved in 10 mMTris base solution and the optical density (OD) was read at 492 nm using a microplate reader. Cell proliferation in the presence of the samples was calculated based on the increased optical density due to the viable cells, as percentage of proliferation compared to the controls in the absence of test samples. SUBSTITUTE SHEET (RULE 26) [PARA 0031] Results [PARA 0032] The results indicated that the extracellular metabolite preparation increased the proliferation of Dermal papilla cells (Fig. 1a, 1b and 1c). It also stimulated the anagen phase of follicular cell growth indicating its potential as a hair care cosmetic. [PARA 0033] Expression of secretory factors [PARA 0034] Materials and Method [PARA 0035] Cells: Dermal Papilla Cells (human HFDPC) were purchased from Promocell (Germany), and maintained as a monolayer culture in Fibroblast growth media ( Promocell, Germany) at 37 °C in a humidified 5% CO<com:Sub>2< / com:Sub> incubator. Dermal Papilla Cells were cultured in DMEM supplemented with 10% FBS. The confluent cultures were harvested by trypsinization and expanded during two more passages before they were used for the experiments. Medium and other culture components were renewed after 48–72 h. All cell cultures were maintained at 37°C in 95% air and 5% <com:Math><semantics>CO2<annotation encoding="application / x-tex">CO_2< / annotation>< / semantics>< / com:Math> in a <com:Math><semantics>CO2<annotation encoding="application / x-tex">CO_2< / annotation>< / semantics>< / com:Math> incubator. [PARA 0036] Dermal Papilla Cells were used for the assay at a seeding density of 5000 cells per well of 96 -well clear bottom microplate in DMEM and incubated at 37°C in 95% air and 5% CO<com:Sub>2< / com:Sub> in a CO<com:Sub>2< / com:Sub> incubator for 24 hours. Culture supernatants were collected and stored at -80°C until use. TGF-\beta and VEGF -1 were detected using ELISA kits (Quantikine® ELISA kit for human TGF-β2 and human VEGF kit ,Krishgen biosystems) as per manufacturer's instructions [PARA 0037] Result [PARA 0038] The extracellular metabolite increased the expression of TGF-β2 (Fig. 2) by 70.3% at a concentration of 0.5% v / v while a 30% increase in VEGF expression (Fig. 3) was observed, indicating that the extracellular metabolite from Bacillus coagulans MTCC 5856 increases the proliferation of dermal papilla and thereby promoting hair growth. [PARA 0039] EXAMPLE 2: 5α-reductase Inhibition [PARA 0040] Isolation of extracellular metabolite SUBSTITUTE SHEET (RULE 26) [PARA 0041] The extracellular metabolite from Bacillus coagulans MTCC 5856 was isolated as per the steps outlined in US9596861. The product is commercially available under the tradename LACTOSPORIN® (INCI: Bacillus ferment filtrate extract) from Sabinsa Corporation, USA. [PARA 0042] Materials and Method [PARA 0043] Cells: Dermal Papilla Cells (human HFDPC) were purchased from Promocell (Germany), and maintained as a monolayer culture in Fibroblast growth media ( Promocell, Germany) at 37 °C in a humidified 5% CO<com:Sub>2< / com:Sub> incubator. Dermal papilla cells (DPC) were cultured in DMEM medium with 10% FBS and seeded in 96 well tissue culture plate. 24 hrs post seeding, DPC were treated with 50nM testosterone with & without the test samples and incubated for 48 hrs in a 5% CO<com:Sub>2< / com:Sub> incubator at 37°C. After 48hrs the supernatant was collected from each well. The amount of <com:Math><semantics>5α<annotation encoding="application / x-tex">5\alpha< / annotation>< / semantics>< / com:Math>-DHT produced by the various treated DPCs was quantified using ELISA and absorbance was measured at 450 nm using a microplate reader [PARA 0044] Conclusion [PARA 0045] The results indicated that the extracellular metabolite preparation brought about 16.2% inhibition in <com:Math><semantics>5α<annotation encoding="application / x-tex">5\alpha< / annotation>< / semantics>< / com:Math>-reductase activity (Fig. 4a), and dihydrotestosterone levels (Fig. 4b) in vitro at <com:Math><semantics>0.5%<annotation encoding="application / x-tex">0.5\%< / annotation>< / semantics>< / com:Math> indicating its potential as a potent <com:Math><semantics>5α<annotation encoding="application / x-tex">5\alpha< / annotation>< / semantics>< / com:Math>-reductase inhibitor for treating clinical conditions like androgenic alopecia. [PARA 0046] EXAMPLE 3: Hair care formulations containing extracellular metabolite preparation from Bacillus coagulans [PARA 0047] Tables 1- provide illustrative examples of hair care formulations containing partially purified extracellular metabolite preparation from Bacillus coagulans MTCC 5856 (Bacillus ferment filtrate extract) [PARA 0048] Table 1: Hair serum [Image disponible dans le document PDF, Image available in the PDF document] Cationic polymers (Galsilk 700), Disodium EDTA, glycerin, Preservatives, non-ionic surfactant (Tween 20), non-ionic solubilizers and emulsifying agents (Cremophor RH 40), Bioavailability enhancers (Piperine extract or Tetrahydropiperine (Cosmoperine®)), Fragrance, Thickeners (Cellulose derivatives or Acrylates Cross Polymer) [PARA 0049] Table 2: Hair serum Active Ingredients Bacillus ferment filtrate extract 0.01% - 2% Amla extract, Cocus nucifera extract, Cosmetic peptides, Selenium sulphide, Vitamin A, C Excipients Cationic polymers (Galsilk 700), Disodium EDTA, glycerin, Preservatives, non-ionic surfactant (Tween 20), non-ionic solubilizers and emulsifying agents (Cremophor RH 40), Bioavailability enhancers (Piperine extract or Tetrahydropiperine (Cosmoperine®)), Fragrance, Thickeners (Cellulose derivatives or Acrylates Cross Polymer) [PARA 0050] Table 3: Hair oil Active Ingredients Bacillus ferment filtrate extract 0.01% - 2% Tea tree oil / coleus oil, Cedarwood oil Excipients Flavouring agent, Preservatives, Carrier oils, Bioavailability enhancers (Piperine extract), Selenium containing amino acids, Selenium sulfide, Antioxidants (Vitamins, rosmarinic acid) [PARA 0051] Table 4: Hair oil Active Ingredients Bacillus ferment filtrate extract 0.01% - 2% Almond oil, Amino acids (Methionine, Cysteine), Selenium Sulfide, Vitamins [Image disponible dans le document PDF, Image available in the PDF document] [PARA 0052] Table 5: Anti-dandruff shampoo [Image disponible dans le document PDF, Image available in the PDF document] [PARA 0053] The above formulations are merely illustrative examples; any formulation containing the above active ingredient intended for the said purpose will be considered equivalent. [PARA 0054] Other modifications and variations to the invention will be apparent to those skilled in the art from the foregoing disclosure and teachings. Thus, while only certain embodiments of the invention have been specifically described herein, it will be apparent that numerous modifications may be made thereto without departing from the spirit and scope of the invention. The scope of the invention is to be interpreted only in conjunction with the appended claims. सुरमजीव प्रस्ता संवर्षन सम्रह एवं जीन बैंक [Image disponible dans le document PDF, Image available in the PDF document] • MICROBIAL TYPE CULTURE COLLECTION & GENE BANK [Image disponible dans le document PDF, Image available in the PDF document] - कुरत १० ८ मानुन्न १०००म गान<com:Br / >सिम्मानीय भूगानुनुक्ता संस्त्रात CSIR-INSTITUTE OF MICROBIAL TECHNOLOGY (A CONSTITUENT ESTABLISHMENT OF CSW) Sector 39-A, Chandigarh-160036 (INDIA) Dr. G.S. Prasad By Courier 17.12.2013 Senior Principal Scientist 30 Dr. Muhammed Majerel Sami Labs. Limited 19 / 1 & 19 / 2, I Mai, 11 Phase Prenya Industrial Area Bangalore - 560 058 Kamataka, India Dear Dr. Muhammed Majood, Your microbial culture has been accepted for deposit in MTCC-IDA under Budapest. Treaty. It was .... assigned MTCC number and preserved in the patent collection of MTCC. Mill details and as follows, . . . . . . . . . . . . . . . . . . . . . . Taxonomic designation Manification Relevence MTCC Number Assigned Date of Accession Bacillus coaquians SBC-37-01 MTCC 5858 19.09.2013 Enclosed here with are the relevant documents of the above strain, Form 8PI4 - Receipt and acceptance of the culture in MTCC Form 8Pi9 - Viability statement of the culture Please acknowledge the receipt of the forms. Sincerely yours, a might fight the same of the same of the same of the same of the same of the same of the same of the same of the same of the same of the same of the same of the same of the same of the same of the same of the same of the . . . . ) G. S. Prasal . . . . . . . . . . . . . . . . Exilia (+31-135-8682121 (m.c.c. quice) 833 -(491-133-2699319 Bethell +91-172-6665152 (Information Centra) ANN - +61-172-2800032 si sar. hadmilijostm dini : naitamellai Phone: +91-172-2690362 (Nesd MTCC) Head, WYCC: curator@imtech.res.in Patent Deposits Sciamfocijismiech restin 1. vi, eva davtvii ootivii, qtid : deW . . . . ٠. [Image disponible dans le document PDF, Image available in the PDF document] . BUDAPEST TREATY ON THE INTERNATIONAL RECOGNITION OF THE DEPOSIT OF MICROORGANISMS FOR THE PURPOSES OF PATENT PROCEDURE . INTERNATIONAL FORM 1.0 [Image disponible dans le document PDF, Image available in the PDF document] . .* erelika ere ereken. . . #6.88 persol<com:Br / > ************************************ 1,15 . ... • ... d. Same and the second . ************************************* . Angar MITTER BETT (miss) many . . . • 2000 . BUDAPEST TREATY ON THE INTERNATIONAL RECOGNITION OF THE DEPOSIT OF MICROORGANISMS FOR THE PURPOSES OF PATENT PROCEDURE INTERNATIONAL FORM 30 Dr. Muhammed Majeed Sami Labs. Limited 19 / 1 & 19 / 2, I Mai, II Phase Bangalore - 560 058 Peenya Industrial Area issued paramet in Rate (0.2 by the INTERNATIONAL DEPOSITIONAL ADTRIONALY VISBBLITY STATEMENT Karnataka, India NAME AND ADDRESS OF THE PARTY ages griveded out on baltiteth TO WIFOM THE YLABILITY STATEMENT IS INFERENT Stage State of X011264436.1 II. IDENTIFICATION OF THE MICRODRGANISM 30 4-5 Dr. Muhammed Majeed V525. . . . . . . . . . . . . . . . . . . . . Accession burnler given by its SYTERNATES ALLENSYSBARY STERMERY ه کر کر کر کر کر کر کر کر کر کر کر کر کر :: : Nikhbis. Sami Labs. Limited MTCC 5856 Space (All Control of Control of Control of Control of Control of Control of Control of Control of Control of Control of Control of Control of Control of Control of Control of Control of Control of Control of Control of Co 19 / 1 & 19 / 2, I Mai, II Phase<com:Br / >Peenya Industrial Area<com:Br / >Bangalore – 560 058 Date of the deposit of of the transfer 19-09-2013 SATTIBEATHE, TONIG Karnataka, India 30 33 20 20 *************************************** HE YEARREST STATEMENT *************************************** The risbility of the microsegment identified under II above was bened m 17-09-2013 <com:Sup>2< / com:Sup> on that date, the sold microorganism was (XX) (1388) eldein regard on '{____} Indicate the date of the original deposit or, where the new deposit or a two site has been reads, the most recent relevant date (dute of the new deposit or dute of the inserior). test etablish meng tena alit ne relet f (ii) fon (ii) (ii). Whit di di binara nema nema kati binara (ii) (ii) sod pickering out escess o lieu kodle. ... <com:Math><semantics>ℒi(x,y)<annotation encoding="application / x-tex">\mathcal{L}_{i}(x,y)< / annotation>< / semantics>< / com:Math> <com:Math><semantics>ΔAiai=Φ0<annotation encoding="application / x-tex">\Delta_{A_{i}a_{i}} \stackrel{\Phi}{=} 0< / annotation>< / semantics>< / com:Math> . 11 8 6 . . 150 ٠. Telephone reconstruction and the contract of the contract of the contract of the contract of the contract of the contract of the contract of the contract of the contract of the contract of the contract of the contract of t (1990), SAN TOO HOW WORK . . - ;• [Image disponible dans le document PDF, Image available in the PDF document] *Form MINX 1999 (second and last page) 4 e e

Claims

1. Use of an effective concentration of extracellular metabolite preparation from Bacillus coagulans MTCC 5856 for increasing hair growth in mammals.

2. The use of claim 1, wherein the increase in hair growth is brought about by a) increasing the expression of TGF-β2 and VEGF in dermal papilla, b) increasing the proliferation of follicle dermal papilla cells and c) inhibiting 5 α 5\alpha -reductase activity in follicle dermal papilla cells.

3. The use of claim 1 or 2, wherein the effective concentration of the extracellular metabolite preparation is 0.01 % 0.01\% v / v to 2.0 % 2.0\% v / v of the total composition.

4. Use of a composition containing extracellular metabolite preparation from Bacillus coagulans MTCC 5856 and a carrier for therapeutic management of androgenic alopecia in mammals.

5. The use of any one claims 1-4, wherein the composition is formulated with pharmaceutically / cosmeceutically acceptable excipients, adjuvants, bases, diluents, carriers, conditioning agents and preservatives and / or incorporated into formulations containing hair care ingredients and for an administration topically in the form of creams, gels, lotions, shampoo, serum, oil, suspensions, emulsions, and compacts.

6. The use of any one of claims 1-5, wherein the mammal is a human.