1. INTRODUCTION:

Hair is a component of the integumentary system and extends downward into the dermal layer where it sits in the hair follicle. Hairs are the integral part of human beauty. Dandruff is a commonly observed affliction that affects most of the adults across regions, ethnicities, and societies. It often causes itching. It has also been observed that those afflicted with dandruff are prone to losing hair faster than those without dandruff. It is observed in a two-day study that dandruff afflicted individuals lost at least twice the hair count compared to those without the disorder (Franchimont et al., 2006). Hair-care products may be defined as the preparations which are used for cleansing hair and scalp, modifying the texture and giving the healthy look to the hair. There are various types of hair: normal hair, oily hair, dry hair, varies from one human to other human. At present, various treatments of dandruff include therapeutic use of zinc pyrithione, salicylic acid, imidazole derivatives, glycolic acid, steroids and Sulphur etc. However, these agents show certain limitations, either due to poor clinical efficacy (Gaganpreet Kaur et al., 2015). Herbal formulations have growing demand in the world market. Poduthalai (or) Turkey tangle frog fruit (Phyla nodiflora) has Anti-bacterial, Anti-oxidant, Anti-fungal, Anti-dandruff and hair growth promoting activities (Snafi et al., 2019). Shikakai, or acacia concinna, has rich amount of vitamin C, which is beneficial for your hair. Shikakai naturally lowers the pH value and retains the natural oils of the hair and keeps them lustrous and healthy. It is also effective in strengthening and conditioning your hair. t is loaded with antioxidants which can protect hair cells from any kind of damage and also repairs damaged hair. Healthy hair cells will contribute to better hair growth as it will provide a healthy environment for the hair to grow. Pulankelangu (or) White turmeric has anti-fungal and antibacterial properties that can treat scalp conditions and irritations such as dandruff, fungus, dryness, and itchiness. The spice creates a healthier environment on your scalp and promotes healthier hair as it reduces breakage and hair loss. Amla, known as Indian gooseberry, is an ample source of vitamin C. It is full of many health benefits. Powdered Amla is used as an essential of hair tonics, required to enhance hair growth and improve hair pigmentation (Mielke et al., 1997). It helps in darkening of hairs and acts as Hair growth promoter. It gives power to the roots, takes care of the color, and improves luster. Amla oil application to the roots of hair improves hair growth and color. The very popular use is to decrease baldness and hair loss. This quality is due to the tannin contents as tannic acid, ellagic acid, Gallic acid, iron and antioxidant material that freezes free radical damage to hair follicles, caused by dandruff (Gavazzoni et al., 2015). Curry leaves helps in reducing hair fall, treats damage hair, remove clogged hair follicles, Scalp restoration. Curry leaves are very effective and maintain the health of our hairs. Curry leaves are beneficial to remove the dullness of our hairs and helps to keep our hairs healthy and naturally gorgeous. From an itchy scalp to grey hairs and from hairs loss to dullness, it can fight it all and help increase hair strength. Murraya Koenigii, Curry leaves help to control dandruff. Its antifungal properties help in reducing dandruff and itchiness leaving you with a clean scalp and healthy hair. Curry leaves moisturize your scalp, promote hair growth, and prevents dandruff. Curry leaves are rich in cell reinforcements that saturate the scalp while disposing the dead hair follicle, helps in preventing hair fall, premature hair graying, and dandruff (Sravanthi et al., 2021). Venthayam (or) Fenugreek are considered to be one of the most effective remedies to treat hair fall issue. It is rich in folic acid, vitamin A, vitamin K, vitamin C. Venthayam also helps to moisturizing the hairs and bringing back the luster.

Pasipayiru (or) Mung bean for hair growth provides body with a good amount of copper. This ensures that iron does its work efficiently. Proper supply of oxygen to the brain maintains the health of the scalp, giving shiny, long, healthy and thick hair. It is packed with nutrients that help boost hair growth, improve scalp health and protect hair by acting as a UV radiation filter. Mung bean is rich in nutrients that protect keratin, the building blocks of hair (Revansiddappa et al., 2018). Vettiveru (or) Vetivergrass is a cell-renewing agent. It enhances collagen formation, a major component of the hair and strengthens cell structure that replenishes the hair back to life. Besides, any scalp irritation, dandruff, or inflammation can also be cured with this power herb. Rose has Ant-microbial, Anti-oxidant effect. The natural astringent properties of rose water may help soothe scalp irritation and inflammation (Boskabady et al., 2011). Hibiscus rosa-sinensis, malvaceae, The Hibiscus nourishes the hair follicles, soften the hair and make it more manageable. Flowers and leaves contain mucilage and plant proteins that helps in treatment of anti-dandruff and hair loss. Hibiscus carries amino acids, Vitamin A, C and alpha hydroxyl acids along with other nutrients that are highly beneficial for hair and scalp. They keep scalp healthy and minimize the chances of dandruff from hair (Kamat et al., 2015). Kadukkai (or) Indian hog plum rich in Iron, essential Vitamins, Manganese, Selenium and other natural compounds, frequent use of Kadukkai promotes healthy hair growth. It has incredible cleansing powers, making it a popular ingredient in herbal shampoos. Frequent application of kadukkai prevents product build-ups on your scalp. It can also penetrate deeper and unclog your hair follicles by removing sebum, dirt and dead cell plugs. A clean scalp allows for better absorption of haircare products and enhances hair growth. Arappu Powder is made from the leaves of Arappu tree. Arappu is a wonderful natural shampoo used for ages. Produces a gentle lather due to the natural saponin, completely nontoxic- no artificial colors or added fragrance and is proven to have enumerable benefits making the hair soft, shiny and lustrous. Thulasi (or) Basil helps to look after the blood circulation and keeps the scalp calm, thereby reducing irritation and dandruff, hence promotes hair growth. Basil is great in prevention of hair loss as it reduces itching in the scalp region and enhances blood circulation in the head area. Anti-oxidant present in tulsi is very useful in treating premature graying of hair as well as hair fall Neem helps to clean the scalp. It clears the clogged pores and improves hair growth. The regenerative properties are extremely essential for the treatment of dandruff. Neem contains a significant ingredient, fatty acids like linoleic, oleic, and stearic acid which help the scalp to nourish thereby keeping the hair smooth. Karisalankanni (or) False daisy powder helps to empower blood circulation to the scalp by stimulating and triggering hair growth, which has been lost due to any cause, probably, dandruff. Eggshell membranes are an abundant raw material that are a novel source for naturally occurring bioactive compounds such as glucosamine (Picard et al., 1973) chondroitin sulfate (Baker and Balch, 1962), hyaluronic acid (Long FD et al., 2005), fibrous proteins such as collagen Type I(Wong M et al., 1984) and sulfur-rich proteins (Tsai et al., 2006) and other components including lysozyme, ovotransferrin, ovocalixin and desmosine and Isodesmosin. TRICHONEM^TM hair wash powder containing unique combinations of eighteen natural herbs and soluble egg shell membrane helps to reduce Dry scalp, Hair fall, Dandruff, Scaling, Pruritus and also increase the hair growth rate and gives healthy hair condition. It helps someone to boost their confidence.

2. MATERIALS AND METHODS:

TRICHONEM ^TMis a scientifically derived nutritional compositions that has the unique blends of 18 herbal extracts added with Soluble Egg Shell Membrane in an optimized proportion to work as Anti-Dandruff and Hair growth promotor in young adults.

Table 1: Ingredients list of TRICHONEM^TMhair wash powder

S. No	Scientific name of ingredients	Common name of ingredients	Qty (g)	Purity Criteria adopted
1	Phyla nodiflora	Poduthalai (or) Turkey tangle frog fruit	100	yes
2	Acacia concinna	Shikakai	250	yes
3	Curcuma Zedoaria	Pulankelangu (or) White turmeric	250	yes
4	Phyllanthus emblica	Amla extarct	250	yes
5	Murraya koenigii	Curry leaf powder	200	yes
6	Trigonella foenum-graecum	Venthayam (or) Fenugreek	150	yes
7	Vigna radiata	Pasipayiru (or) Mung bean	250	yes
8	Chrysopogon zizanioides	Vettiveru (or) Vetivergrass	250	yes
9	Rosa indica	Rose powder	200	yes
10	Hibiscus rosa-sinensis	Sembaruti (or) Hibiscus powder	500	yes
11	Senna auriculata	Avaram (or) Tanner's cassia	100	yes
12	Terminalia chebula	Kadukkai (or) Indian hog plum	50	yes
13	Abelmoschus esculentus	Lady’s finger powder	450	yes
14	Brassica oleracea	Redcabsol (or) Red cabbage powder	25	yes
15	Albizia amara	Arappu leaf powder	50	yes
16	Ocimum tenuiflorum	Tulsi extract	25	yes
17	Azadirachta indica	Neam Leaf powder	100	yes
18	Eclipta prostrata	Karisalankanni (or) False daisy powder	50	yes
19	Gallus gallus domesticus Egg Shell Membrane – Soluble	Soluble egg shell Membrane	250	yes
		Batch size - 3.5 Kg

Merits of each and every ingredients:

1. Poduthalai (or) Turkey tangle frog fruit (Phyla nodiflora):

Phyla nodiflora, powder from plant called Poduthalai or frog fruit was used at concentration of 100g in TRICHONEM^TM hair wash powder. Poduthalai helps to get rid- off dandruff due to it’s antimicrobial, anti-inflammatory and antioxidant properties. Poduthalai is anodyne, antibacterial, deobstruent, diuretic, emmenagogue, parasiticide and refrigerant. The leaves of this phyla nodiflora have higher levels of terpenoids and have significant levels of the component – Beta Caryophyllene and its oxide. Both these constituents along with natural acetates contributes to its excellent anti-microbial properties.

2. Shikakai (Acacia concinna):

Acacia concinna, was used at concentration of 250g in TRICHONEM^TM hair wash powder. Shikakai is recognised for naturally softening and smoothing hair by releasing essential oils and vitamins that are important for hair growth. With its antifungal, antibacterial, and nutritious characteristics, shikakai is an excellent remedy for dandruff. It has no negative effects and can be used to treat the scalp and get rid of annoying dandruff problems It encourages hair development and maintains hair silky and lustrous. shikakai, fortunately, also has a hair lice treatment. Its low pH value, together with its antifungal and antibacterial capabilities, can prevent hair lice from growing. Shikakai prevents dry scalp by acting as a natural cleaner that does not remove essential oils.

3. Poolankilangu (Curcuma Zedoaria) (or) White turmeric powder:

Poolankilangu (or) White turmeric powder (Curcuma Zedoaria) was used at concentration of 250 g in the TRICHONEM^TM formulation. White turmeric benefits for hair in the form of essential oil has medicinal properties. It controls dandruff and hair loss, stimulate hair growth and thicken hair, Boost hair growth and Controls Premature greying. Because of its antioxidant and fortifying properties, White turmeric powder stimulates hair growth and strengthens hair follicles.

4. Amla Powder (Emblica officinalis):

Emblica officinalis, powder from Amla fruit used at strength of 250 g in the formulation. The phyto-nutrients, vitamins and minerals present in Amla help in increasing the scalp circulation and stimulate healthy growth. Vitamin C of Amla produces collagen protein. This helps in stimulating hair growth, both length and volume wise. Amla can nourish, strengthen and condition your hair thereby making it shiny and adding volume to your hair. Dandruff is usually the result of dryness. This vitamin C rich juice cures the dryness and prevents the accumulation of dandruff. Amla juice is great for cleansing the scalp. It nourishes the scalp and makes the hair shiny. Amla in them as it helps in fighting pigmentation. It also helps in discoloration making the hair long and luscious.

5. Curry leaf powder (Murraya koenigii):

Curry leaf powder (Murraya koenigii) powder of 200 g is used in the TRICHONEM^TM hair wash powder. Curry leaves are rich in antioxidants, vitamin C, and Iron that help strengthen the hair roots and prevent hair loss. It also helps in controlling frizz in hair. The antioxidants present in the curry leaves help moisturize the scalp and remove the dead hair follicles. It contains natural ingredients that retain the luster and shine of hair. Its antioxidant properties assist with dry hair. Beta-carotene, protein, and alkaloids present in the curry leaves are beneficial in retaining natural hair tone, promoting hair growth, and preventing hair loss and hair thinning. Curry leaves have properties to fight and treat fungal, bacterial, and protozoal infections. Thus curry leaves can be used to clear dandruff.

6. Venthayam (or) Fenugreek (Trigonella foenum-graecum):

Trigonella foenum-graecum (Venthayam (or) Fenugreek) Powder was used at a concentration of 150 g in TRICHONEM^TM hair wash powder. Fenugreek has antioxidant and anti-inflammatory properties that help with dandruff, scalp irritation. It stimulates blood flow to the scalp, it also nourishes the hair follicles to promote faster and healthier new growth. Fenugreek acts as a great alternative to store-bought conditioner. The mucilage content replenishes the hair and provides smoothness allowing to detangle hair easily post every wash. Fenugreek is a great source of vitamins-A, B, C, K, protein, iron, calcium, potassium, phosphates, folic acid, saponin, flavonoids, and many other components that are essential for healthy hair growth.

7. Pasi payiru (or) Mung bean powder (Vigna radiata):

250 grams of fine powder obtained from Pasi payiru or Moong dal (Vigna radiata) was used in TRICHONEM^TM hair wash powder. Pasi payiru is known to add shine to the hair. Traditionally it has been used to cleanse the scalp and hair. It keeps hair shiny and healthy. The antioxidants like vitamin C and compounds like phytonutrients and polyphenols present in Pasi payiru or Moong dal protect your hair follicles and scalp from the oxidative damage caused by the free radicals. When your hair follicles are well-nourished, they become strong and healthy and are less prone to damage and breakage. It helps to prevent premature greying and also restores luster and vitality of the hair.

8. Vettiveru (or) Vetivergrass:

Chrysopogon zizanioides, Vettiveru was used in a strength of 250 g in TRICHONEM^TM hair wash powder. Vetivergrass improves blood circulation in the hair follicles, prevents various hair infections and also takes part in collagen formation. It is extremely useful for rejuvenating dull, lifeless hair. Regular use of the oil improves hair texture, adds shine and stimulates hair growth. It has potent anti-microbial properties, so it moisturises the scalp, relieves dryness and prevents itching. Vetiver is a big boost for aging hair to replenish collagen formation and bring hair back to life. The anti-inflammatory and antiseptic properties help with scalp inflammation and scalp ailments like eczema, psoriasis, and even dandruff, and its soothing and calming properties it calms irritated and inflamed scalps.

9. Rose powder (Rosa indica):

Rose (Rosa indica) powder used at a concentration of 200 g in the formulation cleanses the scalp by getting rid of extra sebum and dead skin cells. It Relieves dandruff, improves blood circulation to the scalp, Keeps the hair and scalp hydrated. Rose powder gives more shine and volume to hair and acts as a hair conditioner. Improves hair growth. It has anti-inflammatory properties, which may make it beneficial for certain scalp conditions, like psoriasis and eczema. Rose is a mild astringent which may help to reduce oiliness and dandruff. Roses are rich in antioxidants and antimicrobial properties. They deeply nourish and cleanse the scalp, thus strengthening hair follicles and promoting the growth of healthy hair. They serve as a natural conditioner for the hair and prevent the occurrence of dandruff and scalp infection.

10. Sembaruti (or) Hibiscus powder (Hibiscus rosa-sinensis):

Sembaruti (or) Hibiscus powder (Hibiscus rosa-sinensis) of 500 g was used in the TRICHONEM^TM hair wash powder has multiple benefits to hair. The blend of hibiscus flowers and leaves is moisture-rich and helps nourish your scalp. The mucilage content in the leaves and flowers gives it a slimy consistency that protects your hair from drying out. Hibiscus is used for hair growth as it is rich in amino acids, a prime building block of keratin. Keratin is the protein that nourishes, strengthens, and binds the hair roots. Hibiscus is known to improve the build-up of keratin and boost the formation of new hair follicles, thereby stimulating hair growth. It also thickens the hair strands to reduce hair fall. The soothing benefits of hibiscus leaves and flowers can provide lasting relief to a dry/itchy scalp. Hibiscus has astringent properties that reduce excess oil secretion, thus curing dandruff and flakiness. Hibiscus is an excellent source of antioxidants and vitamins that are needed to produce melanin, which gives hair their natural color.

11. Avaram (or) Tanner's cassia (Senna auriculata):

Senna auriculata, powder from Avaram poo (or) Tanner's cassia used at strength of 100 g in the formulation. Avarampoo has anti-bacterial and anti-inflammatory properties, it helps treat scalp infections and also helps reduce scalp inflammation really well. It contains tannins, phenols, flavonoids, steroids, quinones and coumarins. It adds shine, thickens and strengthens hair.

12. Kadukkai (or) Indian hog plum (Terminalia chebula):

Kasturi manjal or Wild turmeric (Curcuma aromatica) of 50 g is used in the TRICHONEM ^TMhair wash powder. Kadukkai is good for promoting hair growth, controlling hair loss, treating lice, treating dandruff problems, preventing grey hair (white hair) and maintaining the natural colour of your hair. Kadukkai Powder is an amazing ingredient to use for hair. Since Kadukkai has anti-microbial properties, it treats all scalp infections really well. Kadukkai is one of the wonderful herb that can be used for different hair problems and maintain your health of hair. It is rich in Iron, essential Vitamins, Manganese, Selenium and other natural compounds, frequent use Kadukkai promotes healthy hair growth.

13. Lady’s finger powder (Abelmoschus esculentus):

Lady’s finger powder (Abelmoschus esculentus) of 450 g was used in TRICHONEM ^TM hair wash powder, which is a rich source of nutrients like Vitamin A, C, folate and calcium. Lady finger benefits for hair is that it prevents frizzy hair and controls curly hair. It provides softer hair, strengthens the hair follicles and roots. Provides shiny hair: It has power pack nutrients and protein that nourishes hair. Treat dandruff with the help of this vegetable which conditions our hair, moisturises the scalp and helps prevent itchiness and dryness. This curbs dandruff and keeps the scalp healthy and strong. This makes the hair soft and helps in strengthening hair roots and follicles. It is very good for treating hair loss as well.

14. Redcabsol (Brassica oleracea) (or) Red cabbage powder :

Red cabbage powder (Brassica oleracea) was used at a strength of 25 g in the TRICHONEM ^TMhair wash powder. The wealth of nutrients in red cabbage including phytochemicals, antioxidants like Anthocyanins and indoles, vitamins like C, A, E, K, B1, B2 and folate, minerals like calcium, manganese, magnesium, iron and potassium. The goodness of antioxidants (Anthocyanins) and Vitamin A in red cabbage helps to stimulates hair growth. Cabbage nourishes hair, thus providing with silky, shining hair. It is rich in silicon and sulfur, two vital minerals that stimulate hair growth and prevent hair loss. Red cabbage is also beneficial for dry hair as it makes it soft and shiny.

15. Arappu leaf powder (Albizia amara):

Arappu leaf powder (Albizia amara) was used at a concentration of 50g in TRICHONEM ^TMhair wash powder formulation. The Arappu powder removes grime and oil and also has a conditioning effect. Arappu acts as a natural conditioner. It reduces heat of the body and softens the hair. Arappu powder could soothe the scalp letting the cells to grow around the hair follicles producing a new shoot of hair. The natural Arappu leaf powder helps to reduce dandruff which irritates the scalp. It also removes the build ups in the scalp like dirt and oil thus gives you a cleaner and healthier scalp condition. Chemical free natural hair wash powder which keeps your hair thick, fresh and aromatic. It gives natural soft, shining and a natural fragrance for hair after wash.

16. Thulasi (Ocimum tenuiflorum):

Ocimum tenuiflorum (Thulasi or Holy basil) Powder was used at a concentration of 25g in TRICHONEM ^TMhair wash powder has anti-bacterial and anti-fungal benefits that helps to keep the scalp healthy while the anti-inflammatory properties give relief from dryness and itchiness of the scalp. Also basil helps in improving blood circulation in the scalp that leads to optimal nourishment of the hair follicles, rejuvenating them and induce new hair growth. Tulsi benefits hair by rejuvenating the hair follicles and strengthening the roots, which in turn curbs hair loss. Tulsi benefits hair by controlling four types of fungal strains that may cause dandruff. This effect of tulsi offers other benefits like improved hair smoothness and hair shine, along with ease of combing and reduced frizz.

17. Neam leaf Powder (Azadirachta indica):

Neam leaf Powder (Azadirachta indica) was used at a concentration of 100g in TRICHONEM ^TM formulation. Neem leaves and its extracts are commonly used for their antiseptic, anti-inflammatory, antioxidant, and healing properties. This excellent herb is a great source of fatty acids, vitamins and minerals that are needed for healthy hair. It contains active constituents like nimbidin, nimbolide and azadirachtin that have some amazing medicinal properties that can help to get rid of every hair problem. Neem has vitamin E and fatty acids which tighten the skin and thus help prevent signs of ageing. It helps lighten blemishes or scars left behind by acne or pimples. The anti-bacterial and anti-fungal properties of neem protect the skin from infections. vitamin E and other antioxidants can help promote a healthier scalp, subsequently minimizing dandruff and resulting in healthier hair. Neem extract is used in killing head lice treatment. This may be due to its azadirachtin content. Azadirachtin can make it difficult for insects to grow and lay eggs by interfering with their hormones.

18. Karisalankanni (or) False daisy powder (Eclipta prostrata):

Karisalankanni (or) False daisy powder (Eclipta prostrata) was used at a strength of 50 g in the TRICHONEM^TMhair wash powder. Eclipta prostrata rejuvenates and strengthens the scalp and hair shaft. It has antimicrobial and antifungal properties that can help reduce dandruff. The oil also has anti-inflammatory properties, which can help psoriasis or other skin irritations on the scalp. It is also said to improve circulation to the scalp. Karisalankanni may slow or prevent the graying process. Gray hair is also commonly understood as a loss of pigment (melanin). The darkening properties of False daisy may help hair appear less gray. Karisalankanni leaves are a rich source of Vitamin E. Vitamin E fights against the free radical which inhibits the growth of hair.

19. Gallus gallus domesticus Egg Shell Membrane – Soluble:

Soluble Egg Shell Membrane is excellent source of Collagen (I, III, V, X, Elastin (Desmosin + Isodesmosin) formulated at 250g in the TRICHONEM^TMformulation. In general Collagen holds dermal layer and hair follicles together which in turn keeps hair strong and healthy, when collagen production reduces due to ageing, hair follicles becomes brittle, while the dermal layer gets dried up, resulting in hair fall. Collagen begins to decline from the age of 25 years. Collagen supports strong healthy skin hair. The eggshell membrane contains a number of bioactive components such as GAG (glycosaminoglycan), chondroitin sulfate/dermatan sulfate (CS/DS), Keratan sulfate (KS), and heparan sulfate (HS)/heparin, hyaluronic acid (HA) that may impact the physiology and presentation of the hair.

2.1. Methods of preparations:

In this study, formulations were prepared with varying concentrations of all natural ingredients. The concentration of each ingredient was mentioned in (Table 1). The powdered ingredients were sieved using #20 mesh, weighed accurately by using digital balance and added in to double cone blends and rotated both clockwise and anticlockwise for 120 minutes for uniform mixing. This was then stored in an airtight container, labeled and used for further evaluation studies.

2.2. Evaluation of TRICHONEM^TM hair wash powder:

2.2.1. Organoleptic Evaluation:

It refers to the evaluation of the TRICHONEM ^TMhair wash powder by utilizing sensory organs like eyes or nose, the examination of the formulation is performed under this evaluation, and it includes macroscopic characteristics of the product, such as color, odor, texture and appearance. The external characters of the formulation were examined based on the method described by (Pal et al.., 2020).

2.2.2. Total ash content:

Ash value is calculated to determine the inorganic contents which is characteristic for an herb. About 2 gm of hair wash powder was taken in silicon dish previously ignited and weighed. Temperature was increased by gradually increasing the heat not exceeding to red color. After complete burning, ash is cooled and weighed.

2.2.3. Physiochemical evaluation:

Physicochemical parameters such as pH, Washability, Solubility, Loss on drying, Particle size and flow properties were determined,

2.2.3.1 pH:

The acidic or alkaline nature of the TRICHONEM ^TMhair wash powder of combined form was determined by preparing 1% dispersion of powder form in distilled water at room temperature 25°C. The pH was measured by using digital pH Meter.

2.2.3.2 Washability:

TRICHONEM ^TMhair wash powder formulations were applied on the skin and then ease and extent of washing with water were checked manually.

2.2.3.3 Solubility:

Solubility is defined as the ability of the substance to soluble in a solvent. One gram of the TRICHONEM ^TMhair wash powder is weighed accurately and transferred into a beaker containing 100 ml of water. This was shaken well and warmed to increase the solubility. Then cooled and filtered it, the residue obtained is weighed and noted (Sagar et al., 2005).

2.2.3.4 Loss on drying:

Loss on drying is the loss of mass expressed in percent m/m. Two gram of the powder is weighed accurately and

transferred into a dry Petri dish. The Petri dish is placed in a desiccator for 2 days over calcium chloride crystals. Then the powder was taken and weighed accurately to find out the weight loss during drying.

2.2.3.5 Particle size and Flow parameters:

The particle size was tested by microscopy method. The flow property of the dried powder of combined form was evaluated by performing Angle of Repose by funnel method, bulk density and tapped density by Tapping Method (Londhe et al., 2020).

2.2.4. Irritancy test:

Mark an area (1sq.cm) on the left-hand dorsal surface. Definite quantities of prepared TRICHONEM ^TM hair wash powder were applied to the specified area and time was noted. Volunteers response were analyzed for any eye irritation during the application of hair wash powder. Irritancy, erythematic, edema, was checked on skin if any for regular intervals up to 24 hrs. and reported (Banchhor et al., 2009).

2.2.5. Hair wash properties:

2.2.5.1 Dirt dispersion:

Two drops of 1% each shampoo powders were added in a large test tube contain 10 ml of distilled water. 1 drop of India ink was added; the test tube was stoppered and shaken for 10 times. The amount of ink in the foam of was estimated as None, Light, Moderate, or Heavy (Patil et al., 2015).

2.2.5.2 Foaming Index and foam stability:

One gram of the powder was weighed accurately and transferred into 250 ml conical flask containing 100 ml of boiling water. Then it is warmed gently for 30 minutes, cooled and filtered and make up the volume to 100 ml in standard volumetric flask. This extract is taken in 10 test tubes in a series of successive portion of 1, 2, 3….10 ml and remaining volume is made up with water to 10 ml. Then the test tubes were shaken in longwise motion for 15 seconds at speed of 2 frequencies /second. Then the tubes are allowed to stand for 15 minutes. The height of the foam was measured (Gaganpreet Kaur et al., 2016).

Foaming index =1000/a

2.2.6. Stability studies:

Stability testing of prepared TRICHONEM ^TM hair wash powder formulation was conducted by storing at different temperature conditions for the period of 4, 8, 12 months. The packed glass vials of formulation stored at different temperature conditions viz.., Room temperature and 40ºC and were evaluated for physical parameters like Color, Odor, pH, Consistency and feel (Anusha et.al., 2013).

2.2.7 In vitro Antidandruff Activity of TRICHONEM ^TMhair wash powder:

Organism Used (Dermatophyte): Test organisms used in this study are Malassezia furfur (MTCC 1372), Trichophyton mentagrophytes (MTCC 8276) and Microsporum gypseum (MTCC 2819) were obtained from Microbial Type Culture Collection, India. The freeze dried cultures were inoculated into broth and maintained in Sabouraud’s Dextrose.

2.2.7.1 Antifungal Susceptibility Testing-Disk Diffusion Method:

In-vitro antidandruff activity was performed using the disk diffusion assay described by (Sharma et al., 2012) Suspension containing 5 X10 CFU/ml of dandruff causing organisms was swabbed on the surface of the sterile SDA plates using a sterile cotton swab. Sterile filter paper discs impregnated with TRICHONEM ^TM hair wash powder water extracts ranging from 25 µg/ml, 50 µg/ml and 75 µg/ml per disc was aseptically placed over the seeded SDA plates. Similarly, standard antibiotic disc of fluconazole was used as standard drug for comparison of antifungal activity. The plates were incubated at 27°C for 28 hrs and at the end of incubation the diameter of the inhibition zones was measured and tabulated.

2.2.7.2 Determination of Minimum Inhibitory Concentration (MIC):

Different concentrations of TRICHONEM ^TM hair wash powder (0.5 to 2.0 mL) was prepared and dispensed into test tubes and made up to 5 mL with sterile SD broth. One drop of an overnight broth culture of dandruff causing microorganisms was inoculated and incubated for 22-28 hrs at 25°C to determine the MIC. The culture inoculated into the sterile SD broth and placed at 2°C in a refrigerator overnight was used as standard. The MIC value was taken as the lowest concentration of compound at which there is no visible growth of the test organisms after 22-28 hrs of inoculation at 25°C (Anitha et al., 2015).

2.3. Pilot study:

2.3.1 Selection of participants:

The study was conducted in Tamil Nadu, Southern part of India at Erode district. The study enrolled a group of 50 young aged healthy volunteers. They were randomized in double- blind fashion to apply TRICHONEM ^TM hair wash powder or placebo with broken rice powder. 30% of volunteers were in the age range of 22-25 years; 40% were 25-30 years; and 30% were 30-40 years old (Table 2).

2.3.2 Study design:

TRICHONEM ^TM hair wash powder formulation effect on human were compared to placebo in pilot scale study. An open-label 4-week study was performed involving 50 healthy females between the age of 22 years and 40 years. After inclusion in study protocol each volunteer will receive the respective treatment for once a week for 4 weeks. A container of 500 gm TRICHONEM ^TM hair wash powder formulation and container of 500 gm placebo will be given to them. Instruction regarding application of hair wash powder will be given to them. After completion of the treatment for 4 weeks on next day each participant will be asked to give ratings for the following hair parameters such as Dry scalp, Hair fall, Dandruff, Scaling, Pruritus.

3. RESULT AND DISCUSSION:

3.1 Results of evaluation test performed:

3.1.1. Organoleptic Evaluation:

TRICHONEM^TM hair wash powder was evaluated for Organoleptic parameters showed in the (Table 2). The TRICHONEM^TM was powder in nature and greenish in color. The odor of prepared formulations has aroma of the ingredients which was Characteristic and is desirable to hair wash formulations. Texture of the TRICHONEM ^TM hair wash powder was fine and smoothness was acceptable as per requirement of hair wash formulations.

Table 2: Organoleptic Evaluation of TRICHONEM ^TM hair wash powder

S. No	Parameter	Observation
1	Appearance	Powder
2	Color	Green
3	Odor	Characteristic
4	Texture	Fine
5	Smoothness	Smooth

3.1.2. Physiochemical evaluation:

The study of ash values, pH, bulk density, tapped density of TRICHONEM ^TMhair wash powder provided the important feature of physicochemical evaluation. The pH of face pack was found close to neutral. The ash content and moisture content was within limit. The yielded was found to be 3.2% of total ash. Values of particle size, angle of repose, bulk density and tapped density obtained for powder of TRICHONEM ^TM hair wash powder were found to be 20-26μm, 26 ”8, 0.89g/ml and 1.3 g/ml respectively (Table 3), which denoted the free flowing properties of face pack. The TRICHONEM ^TM hair wash powder powder had passable flow property which is suitable for a hair wash powder. The solubility test revealed that the hair wash powder comprises of mostly soluble ingredients which may be helpful in producing comfortable feeling while being mixed with water.

Table 3: Physiochemical evaluation of TRICHONEM ^TMhair wash powder

S. No	Parameter	Observation
1	Total Ash	3.2 %
2	Particle size	20-26μm
3	Angle of Repose	26 ”8
4	Bulk density	0.89g/ml
5	Tapped density	1.3 g/ml
6	pH	6.6
7	Washability	Easily washable
8	Solubility	Easily soluble with water
9	Loss on drying	1.3 % (w/w)

Table 4: Irritancy Test results of TRICHONEM ^TMhair wash powder

S. No	Parameter	Presence	Observation
1	Irritant	NIL	No Irritation
2	Erythema	NIL	No Irritation
3	Edema	NIL	No Irritation

3.1.3 Irritancy Test:

The results of irritancy test were shown in (Table 4). When these TRICHONEM ^TM hair wash powder formulations applied on the skin of left hand, they did not produce any type of redness, edema, inflammation and irritation.

The formulation showed absence of irritation, redness and swelling on both skin and eye during irritancy studies. This shows that TRICHONEM ^TMhair wash powder formulation are safe to use on skin. Most of the synthetic surfactants produce inflammation of the eyelid and corneal irritation. But in this formulation of herbal shampoo powder, the uses of all ingredients are obtained naturally. So it does not produce any harmful effects on skin and eye.

3.1.4 Hair wash properties:

TRICHONEM ^TM has Moderate dirt dispersion activity. This produced a foaming index of 2.5 ml which is a good foaming ability. This results showed that the powder which is capable to produce high foaming property. This is due to the presence of soap nut in TRICHONEM ^TMhair wash powder formulation.

3.1.5 Stability studies:

The stability studies showed a slight change in pH of formulation after 4, 8, 12 months, which was stored at 40ºC No change in color, odor, texture, pH and smoothness was observed for the formulations after 4, 8, 12 months of storage at mentioned conditions of stability at room temperature (Table 5). The odor of formulation was slightly changed after twelve months of stability studies at 40ºC and there was no change in color and odor at other mentioned conditions of stability.

3.1.6 In vitro Antidandruff Activity of TRICHONEM ^TMhair wash powder:

3.1.6.1 Antifungal Susceptibility Testing-Disk Diffusion Method:

In vitro Antidandruff Activity TRICHONEM^TMhair wash powder was performed using disk diffusion assay. Dandruff causing organisms such as Malassezia furfur (MTCC 1372), Trichophyton mentagrophytes (MTCC 8276) and Microsporum gypseum (MTCC 2819) were used in this test. M.furfur, T. mentagrophytes and M. gypseum was sensitive to TRICHONEM ^TMhair wash powder. TRICHONEM ^TMhair wash powder showed inhibition of 1.6 cm, 1.9 cm, 2.2 cm against M. furfur in 25μg/ml, 50μg/ml, 75μg/ml concentrations. There was also good inhibition ranging from 0.6cm, 0.8cm, 1.5cm against T. mentagrophytes at 25μg/ml, 50μg/ml, 75 μg/ml powder concentrations. Similar zones with values of 0.5cm, 0.7cm, 1.3cm were found against another dandruff causing organism M. gypseum in concentrations of 25, 50, 75μg/ml (Table 6). From this it is evident that the dandruff causing organisms were more sensitive to all the concentrations of TRICHONEM^TMhair wash powder.

3.1.6.1 Minimum Inhibitory Concentration (MIC):

Table 7 shows minimum inhibitory concentrations of TRICHONEM^TMagainst dandruff causing organisms. The highest MIC value of 0.22 was obtained for M. furfur (MTCC 1372) at 2ml. There was effective reduction in growth at concentrations of 2 ml with MIC value of 0.36 for T. metagrophyte (MTCC 8276). Growth of M. gypseum (MTCC 2819) was reduced at 2 ml concentration with OD value of 0.29 at 660nm. The growth of the organisms reduced gradually with increase in concentration of TRICHONEM^TM. From the above it has been observed that TRICHONEM^TM was rapidly inhibiting the multiplication of the dandruff causing microorganism in test tube at highest.

Figure 2: Minimum Inhibitory Concentration (MIC) of TRICHONEM ^TM against dandruff causing organisms

Table 6: Zone of inhibition by TRICHONEM ^TM against dandruff causing micro-organisms

Organism	Concentration in μg/mL
Organism	25 μg/mL	50 μg/mL	75 μg/mL
M. furfur	1.6	1.9	2.2
T. metagrophyte	0.6	0.8	1.5
M. gypseum	0.5	0.7	1.3
Zone of inhibition in, cm

Table 7: Minimum Inhibitory Concentration (MIC) of TRICHONEM ^TM against dandruff causing micro-organisms

Organism	OD@ 660 nm
Organism	0.5 ml	1 ml	1.5 ml	2 ml
M. furfur	0.49	0.46	0.32	0.22
T. metagrophyte	0.57	0.48	0.4	0.36
M. gypseum	0.52	0.44	0.39	0.29

3.2. Pilot study:

TRICHONEM ^TMhair wash powder formulations and placebo formulations were compared to placebo was given in the Table 8. TRICHONEM ^TM hair wash powder formulation visibly treats Dry scalp, Hair fall, Dandruff, Scaling, Pruritus after 4 weeks between the age of 22-40, but placebo treated group does not have visible reduction of Dry scalp, Hair fall, Dandruff, Scaling, Pruritus. It was to conclude that TRICHONEM ^TM hair wash powder formulations is very effective. The molecules present in the formulation help fighting against dandruff causing organisms and provide strength to hair follicles. Collagen in soluble egg shell membrane holds dermal layer and hair follicles together which in turn keeps hair strong and healthy. Ratings were given by the volunteers for the TRICHONEM ^TM hair wash powder formulations and Placebo formulation (Fig.3).

Table 8: Skin ratings of TRICHONEM ^TM hair wash powder and Placebo

Ratings	TRICHONEM ^TM	Placebo
Dry Scalp	4	3
Scaling	5	2
Pruritus	4	2
Dandruff	5	3
Hair fall	5	2

Ratings:

5 - Complete mitigation: 90 – 100% relief

4 -Great improvement: 75-90% relief

3 -Significant improvement: 50-74% relief

2 -Mild improvement: 25 – 49% relief

1 -Unsatisfactory: < 25% improvement from the pre-test condition

Figure 3: Comparison of TRICHONEM ^TM and Placebo

CONCLUSIONS:

TRICHONEM ^TMhair wash powder, a unique blend of eighteen herbal powders and soluble egg shell membrane was formulated to develop a stable and functionally effective anti-dandruff hair wash powder by excluding all types of synthetic additives. Harmful effect of chemical products is forcing everyone to switch over to natural products, this led to growing demand for Herbal formulations in the world market. Every Ingredients used in this TRICHONEM ^TMhair wash powder has unique properties in improving the nature of hair and in treating the problems of hair such as Dry scalp, Hair fall, Dandruff, Scaling, Pruritus. TRICHONEM ^TM hair wash powder has soluble Eggshell Membrane which has Collagen that holds dermal layer and hair follicles together which in turn keeps hair strong and healthy. Thus in the present work, It is a very good attempt to formulate the herbal hair wash powder containing naturally available ingredients like Poduthalai (or) Turkey tangle frog fruit, Shikakai, Pulankelangu (or) White turmeric, Amla extarct, Curry leaf powder, Venthayam (or) Fenugreek, Pasipayiru (or) Mung bean, Vettiveru (or) Vetivergrass, Rose powder, Sembaruti(or) Hibiscus powder, Avaram (or) Tanner's cassia, Kadukkai (or) Indian hog plum, Lady’s finger powder, Redcabsol (or) Red cabbage powder, Arappu leaf powder, Tulsi extract, Neam Leaf powder, Karisalankanni (or) False daisy powder and Soluble egg shell Membrane. TRICHONEM ^TM hair wash powder has good organoleptic and flow properties. High foaming index was produced by hair wash powder due to the usage of soap nut as foaming agent which mainly consists of sapoindoside A and B. The eye and skin irritation tests revealed that the TRICHONEM ^TM hair wash powder shows no harmful effect on skin and eye. This is due to the absence of synthetic surfactants. An open-labeled 4-week study on human with TRICHONEM ^TM hair wash powder formulation visibly treats Dry scalp, Hair fall, Dandruff, Scaling, Pruritus. TRICHONEM ^TM treated Volunteers Ratings were high for all hair parameters considered for study in comparison with placebo. TRICHONEM ^TM applied volunteers felt their hair conditions was improved greatly and was overwhelmed by the results of TRICHONEM ^TM hair wash powder effect on their hair in comparison with placebo treated volunteers.

REFERENCES:

1. Franchimont PC, Uhoda XE, Piérard GE. Revisiting dandruff. Int J Cosmet Sci. 2006; 28(5):311-318.

2. Gaganpreet Kaur et al. Formulation and evaluation of anti-dandruff Polyherbal powder shampoo. Journal of Quality Assurance and Pharma Analysis. 2016; Vol 2 (1), 115-121

3. Snafi AE. Pharmacological and therapeutic effects of Lippia nodiflora (Phyla nodiflora). IOSR Journal of Pharmacy (IOSRPHR). 2019; vol. 9, no. 8, pp. 15-25

4. Mielke H. Lead-based hair products: Too hazardous for household use. J Am Pharm Assoc 1997; 85-9

5. Gavazzoni Dias MF. Hair cosmetics: An overview. Int Journal Trichology. 2015; 7(1): 2.

6. Sravanthi et al. A Review on Formulation and Evaluation of Herbal Anti-Dandruff Shampoo. International Journal of Pharmaceutical Research and Applications. June 2021; Volume 6, Issue 3 May, pp: 1300-1311. DOI: 10.35629/7781-060313001311

7. Revansiddappa M, Sharadha R, Abbulu K. Formulation and evaluation of herbal Anti-dandruff shampoo. Journal of Pharmacognosy and Phytochemistry. 2018; 7(4): 764-767

8. Boskabady MH, Shafei MN, Saberi Z, Amini S. Pharmacological Effects of Rosa Damascena. 2011; Jul-Aug; 14(4): 295–307

9. Kamat N, Pearline D, Padma T. Hibiscus Rosa Sinensis -a versatile Indian origin plant. J Chem Pharma Sci. 2015; 8(4): 567-79

10. Picard J, Gardais AP, Vedel M. Sulfated glycoproteins from egg shell membranes and hen oviduct. Isolation and characterization of sulfated glycopeptides. Biochim. Biophys. Actavol. 1973; 320, pp. 427-441.

11. Baker JR, Balch DA. A study of the organic material of hen’s-egg shell. Biochem. J. 1962; pp. 352-361.

12. Long FD, Adams RG, DeVore DP. Preparation of hyaluronic acid from eggshell membrane. USA Patent. 2005; 6946, 551.

13. Wong M, Hendrix MJC, Von der MK. Collagen in the egg shell membranes of the hen. Dev. Biol. 1984; vol-104 pp. 28-36.

14. Tsai WT et al. Characterization and adsorption properties of eggshells and eggshell membrane. Bioresource Technology. 2006; pp. 488-493.

15. Pal et al. Preparation & Assessment of Poly-Herbal Anti-Dandruff Formulation. The Open Dermatology Journal. 2020; 14, 22-27. DOI: 10.2174/1874372202014010022

16. Sagar R, Dixit VK. Formulation and evaluation herbal anti-dandruff shampoo. Nig J Nat Prod Med. 2005; 09: 55-60

17. Banchhor M, Ashawat MS, Saraf S. Herbal cosmetics: Trends in Skin Care Formulation. Pharmacogn Reviews. 2009; 3(5): 82-89

18. Patil SS, Yuvraj JM, Shrinivas KM. Formulation and evaluation of herbal shampoo powder. International Journal of Advanced Research. 2015; Volume 3, Issue 3, 939-946

19. Anusha et.al. Formulation and evaluation of herbal anti-dandruff shampoo. Indian Journal of Research in Pharmacy and Biotechnology. 2013; ISSN: 2321-5674

20. Sharma R, Sharma G, Sharma M. Comparative antifungal study of essential oil with allopathic drugs against M. furfur in vitro. Ind J of Pharma Biological Arch. 2012; 3(1): 89-93.

21. Anitha A, Sreedevi P, Arunkumar D. In vitro Evaluation of Indigenous Medicinal Plants for their Antidandruff Hair Oil Preparation. Global Journal of Pharmacology. 2013; 7 (4): 429-435

22. George W, Latimer JR. Official method of analysis of AOAC. AOAC international. 2019; 21^st Edition.

Received on 11.06.2022 Accepted on 20.07.2022

Research J. Topical and Cosmetic Sci. 2022; 13(2):76-86.

DOI: 10.52711/2321-5844.2022.00013

Parameters	4 months		8 months		12 months
Temperature	RT	40±0.5ºC	RT	40±0.5ºC	RT	40±0.5°C
Color	No change	No change	No change	No change	No change	No change
Odor	Characteristics	Characteristics	Characteristics	Characteristics	Characteristics	Slight change
Texture	Fine	Fine	Fine	Fine	Fine	Fine
pH	6.6±0.5	6.7±0.4	6.6±0.5	6.9±0.4	6.6±0.5	7.03±0.4