WO2010097695A2

WO2010097695A2 - Herbal cleansing composition and methods thereof

Info

Publication number: WO2010097695A2
Application number: PCT/IB2010/000404
Authority: WO
Inventors: Ramesh Surianarayanan; Anindita Das; Ekta Saxena
Original assignee: Hymalaya Global Holdings Ltd.
Priority date: 2009-02-27
Filing date: 2010-02-27
Publication date: 2010-09-02
Also published as: WO2010097695A3; WO2010097695A8

Abstract

Disclosed herein is a herbal cleansing composition comprising extracts of herbs an surfactant selected from non-ionic surfactant and non-ethoxylated anionic surfactant an pharmaceutically acceptable excipients wherein the extract is prepared employing a super critical fluid extraction (SCFE), percolation or hot- soxhalation method or enzymatic extraction method.

Description

HERBAL CLEANSING COMPOSITION AND METHODS THEREOF

Field of the Invention

This invention, in general relates to a herbal cleansing composition and methods thereof. In particular, the present invention provides herbal cleanser composition comprising non-ionic surfactant and non-ethoxylated anionic surfactant along with herbal actives and method for preparing the same.

Background of the Invention In general the cosmetic cleansing compositions such as for skin and hair comprises of water, surfactants, viscosity controller for thickening the formulation, solubility enhancer for water-insoluble substances, perfume oils, preservatives and other active ingredients for the care of skin and hair.

A cleansing composition particularly for hair commonly comprises ethoxylated anionic surfactant, like, sodium lauryl ether sulfate, etc., these formulations are further containing an additional amphoteric surfactant like, cocoamidopropyl betaine. The formulations also contain additional silicones for hair conditioning benefits. Usage of large amounts of conditioning agents that work to control flyaway by coating and weighing down the hair commonly results in a poor impression of hair cleanliness and leaving the hair and hands with a tacky and dirty feeling.

It is therefore necessary to develop a herbal cleanser composition that is devoid of large amounts of conditioning agents to provide better conditioning and cleansing properties and is safe to use.

Accordingly, the present invention provides a composition for various types of cleansers, which avoids the limitation associated with the composition known in the art.

Related Art

U.S. Patent No. 20070286838A1 to Axelrod et al. discloses a dermatological composition which may be used for the topical treatment of the skin, hair, or nails of mammals, comprising tamanu oil, an unsaturated fatty acid, and arnica oil, optionally including Vitamins E and A and herbal extract of Aloe Barbadensis Leaf Extract, Chamomilla Recutita (Matricaria) Flower Extract, Triticum Vulgare (Wheat) Germ Oil, Triticum Vulgare (Wheat) Gluten, Avena Sativa (Oat) Kernel Extract, Althaea Officinalis (Marshmallow) Root Extract, Glycyrrhiza Glabra (Licorice) Extract, Epilobium Angustifolium Extract or Squalane Extract and an anionic surfactant. The composition may be in the form of a shampoo, conditioner, lotion, cream, spray, or salve. Patent No. WO2007068390A1 to Gargano et al. discloses aqueous pre-mixes for making personal or home care compositions, consisting of surfactants, plant extracts, and optionally, preservatives, and the use of said pre-mixtures for making various types of compositions. The pre-mixes consisting of anionic, non-ionic and/or amphoteric surfactants, botanical extracts, and preservatives.

JP2004292359A to Asada discloses a shampoo for curly hair comprises a surfactant including an anionic surfactant, a nonionic surfactant and an amphoteric surfactant and vegetable essence. The shampoo for curly hair includes the vegetable essence which is selected from licorice essence, essence of Arnica Montana, essence of Hedera helix L., garlic essence, essence of Rome chamomile, essence of Rosmarinus officiealis, essence of Hamamelis, essence of Hypericum erectum, essence of Aesculus hippocastanum, pinetree essence, grape leaf essence, burdock essence, essence of Netherlands mustard and essence of Lamium album.

US patent application no. 20060165636 Al to Hasebe et al. discloses a composition for hair treatment containing γ-polyglutamic acid or a salt thereof, a hair cosmetic for damaged hair containing such a composition, and their uses. The composition also contains Glycyrrhiza glabra, Cucumis sativus, Carica papaya and other herbs extract.

Summary of the Invention It is a principal object of the present invention to provide a cleansing composition having excellent cleansing effect with the use of non-ionic surfactant, anionic surfactant, benefited with the goodness of herbal actives.

It is one other object of the present invention to provide a cleansing composition having conditioning benefits such as smoothness and softness, without the use of silicones.

The above and the other objects of the present invention are attended according to following preferred embodiments of the present invention, however the scope of the invention is not restricted to any particular embodiment.

In accordance with one embodiment of the present invention, there is provided a herbal cleanser composition comprising extracts of herbs and surfactant selected from non- ionic surfactant and non-ethoxylated anionic surfactant and pharceutically acceptable excipients.

In accordance with further embodiment of the present invention, there is provided a herbal cleanser composition comprising extracts obtained from a blend of herbs selected from Eclipta prostate, Glycyrrhiza glabra, Phyllanthus emblica, Terminalia bellarcia, Terminalia chebula, Cicer arietinum, Lawsonia inermis, Carica papaya, Sapindus mukurrosi, Cucumis sativus, Glycine max, Rosmarinus officinalis and in various combination thereof.

In accordance with further embodiment of the present invention, there is provided a herbal cleanser composition comprising extracts of is obtained from a blend of herbs selected from Eclipta prostate, Glycyrrhiza glabra, Phyllanthus emblica, Terminalia bellarcia, Terminalia chebula, Cicer arietinum and Lawsonia inermis.

In accordance with further embodiment of the present invention, there is provided a herbal cleanser composition comprising extracts of is obtained from a blend of herbs selected from Carica papaya, Sapindus mukurrosi, Cucumis sativus, Glycine max, Glycyrrhiza glabra and Rosmarinus officinalis.

In accordance with another embodiment of the present invention, there is provided a herbal cleanser composition, wherein the blend is used in the range of 0.0001% to 50%.

In accordance with yet another embodiment of the present invention, there is provided a method of preparing the herbal cleanser composition, wherein said composition is prepared by the method comprising of blending the herbs and extracting the resultant herbal blend employing super critical fluid extraction (SCFE), mixing the resultant extract of blend with non-ionic surfactant and non-ethoxylated anionic surfactant and pharmaceutically acceptable excipients.

In accordance with yet another embodiment of the present invention, there is provided a method of preparing the herbal cleanser composition, wherein said composition is prepared by the method comprising of blending the herbs and extracting the resultant herbs employing percolation or soxhalation using water, mixing the resultant extract of blend with non-ionic surfactant and non-ethoxylated anionic surfactant and pharmaceutically acceptable excipients.

In accordance with yet another embodiment of the present invention, there is provided a method of preparing the herbal cleanser composition, wherein said composition is prepared by the method comprising of blending the herbs and extracting the resultant herbs employing enzymes, mixing the resultant extract of blend with non-ionic surfactant and non-ethoxylated anionic surfactant and pharmaceutically acceptable excipients.

Description of the Invention

While this specification concludes with claims particularly pointing out and distinctly claiming that, which is regarded as the, invention, it is anticipated that the invention can be more readily understood through reading the following detailed description of the invention and study of the included examples. This invention provides a synergistic combination of non-ionic surfactant, like, alkylpolyglucoside and non-ethoxylated anionic surfactant, like, ammonium lauryl sulfate, with herbal actives for cleansing preparations including quaternary compounds for conditioning benefits.

The composition according to the present invention can be formulated in various kinds of cleansers, preferably in a shampoo formulation.

The cleansing composition according to the present invention, wherein said extract of herbs can be prepared by using any part of herbs, preferably whole plant of Eclipta prostate, roots of Glycyrrhiza glabra, leaves of Lawsonia inermis, fruits of Phyllanthus emblica, Terminalia bellarica, and Terminalia chebula and seeds of Cicer arietinum.

The cleansing composition according to the present invention, wherein said extract of herbs can be prepared by using any part of herbs, preferably roots of Glycyrrhiza glabra, leaves of Rosmarinus officinalis, fruits of Carica papaya, Sapindus mukurrosi and Cucumis sativus, and seeds of Glycine max.

The following non-limiting examples illustrate in details about the invention. However, they are, not intended to be limiting the scope of present invention in any way.

Example- 1

Experimental Procedure:

Five different shampoo formulations were made with different combinations of Alkylpolyglucoside and Ammonium Lauryl Sulfate for evaluation on hair swatches and the constituent of each formulation is mentioned in the below table.

These formulations were also compared with the conventional formulation containing Sodium Laureth Sulfate and Coco Amidopropylbetaine, the formula for which is also mentioned in the table.

Experimental Formulation:

Manufacturing Method:

1 . Decyl Glucoside and Lauryl Glucoside is weighed in the main vessel and mixed well for 10 minutes. The mixture pH is adjusted by 25% citric acid solution around 5.50.

2. Heat the main vessel to 5O⁰C. Cool to 45⁰C under slow stirring and add Polyquaternium 7 and continue mixing for 20minutes.

3. A portion of DM water is weighed in a side vessel and kept in a propeller stirrer. Guar Hydroxypropyltrimonumchloride is transferred into the DM water slowly. This is mixed for 20 minutes to ensure complete mixing. This is transferred into main vessel and mixed for 20 minutes.

4. Potassium sorbate is dissolved in a small portion of water and added to the main vessel and mixed well for 10 minutes.

5. Sodium Benzoate is dissolved in a small portion of water and added to the main vessel and mixed well for 10 minutes.

6. Methyl chloroisothiazolinone & Methyl isothiazolinone is added into the main vessel and mixed well for 10 minutes.

7. Perfume is added into the main vessel and mixed well for 10 minutes.

8. Paraben-free Herbal active is mixed with alkylpolyglucoside and added to the main vessel and mixed well for 10 minutes.

9. DM water is added finally to adjust the batch quantity to 100%. Conventional Formulation:

Manufacturing Method:

1. Sodium laureth sulfate and Cocamidopropylbetaine are weighed in the main vessel and mixed well for 10 minutes.

2. Heat the main vessel to 50⁰C. Cool to 45°C under slow stirring and add

Polyquaternium 7 and continue mixing for 20minutes.

6. The mixture pH is adjusted by 25% citric acid solution to around 5.50.

7. Methyl chloroisothiazolinone & Methyl isothiazolinone is added into the main vessel and mixed well for 10 minutes.

8. Perfume is added into the main vessel and mixed well for 10 minutes.

9. Paraben-free Herbal active is mixed with alkylpolyglucoside and added to the main vessel and mixed well for 10 minutes.

10. DM water is added finally to adjust the batch quantity to 100%. Evaluation Details:

Conditioning Evaluation - Experiment

1. Hair tress/Hair Swatch In-vitro evaluation

2. Chemically untreated human hairs were selected for evaluation that has no permanent waves, no hair color or even beaten up from styling with curling irons.

3. Approximately 20 cm long and 15g weight hair swatches were selected.

4. 1.5gram of shampoo was taken for the wash and hair swatches were washed manually and rinsed in the running water.

5. Cold air blowers dried up the hair swatches.

6. The hair swatches were kept in a stand.

7. Trained panel members evaluated the hair swatches.

8. They were asked to rank the product in 10 scales/

9. The ranking scale was given below 1 2 3 4 5 6 7. .8 9 10

Not acceptable Acceptable Excellent

Results of the Evaluation: The 'Formulas A, B, C, D and E' are compared with each other in hair swatches

Result : The 'Formula E', which is with 0.01% Paraben-free Herbal Active and 15.00% level of Ammonium Lauryl Sulfate and 30% level of Alkylpolyglucoside , was found to excel out in comparison to Formula A, B, C and D which have Paraben-free Herbal Active at 0.0001%, 0.005%, 0.002% and 0.001% level, respectively, and (ALS:APG) combination at a level of (15: 15), (20:20), (20:20) and (15: 15), respectively.

Further confirming the synergistic effect of Alkylpolyglucosides and Ammonium Lauryl Sulfate with Paraben-free Herbal Active and quaternary compounds, 'Formula E' is evaluated against 'Formula F', which consists of conventional surfactants like Sodium laureth sulfate, Cocamidopropyl betaine with Paraben-free Herbal Active and quaternary compounds. The hair tress evaluation results are as below

Formula Formula

Parameters E F

In use feel 8 4.5

Wet combing 8.5 3.5

Dry combing 9 3

Softness of hair 9 5

The ratings clearly show 'Formula E' is significantly superior to 'Formula F' in all aspects.

There is clear synergism witnessed between non-ionic surfactant, like, alkylpolyglucoside and non-ethoxylated anionic surfactant, like, ammonium lauryl sulfate, with herbal actives. Example 2

Preparation of extract from Eclipta prostata by percolation method: The shade dried material of whole plant of Eclipta prostata was pulverized to coarse powder and about 10 Kg each of powdered material placed in different percolators and extracted with n-hexane, acetone, ethyl alcohol, methanol, ethyl alcohol and water (1 : 1), methanol and water (1 : 1 ) and water at room temperature for 24 h to 48 h., then plant extracts were filtered and concentrated to dryness on rotatory evaporator or on steam bath at optimum temperature and under reduced pressure.

Example 3

Preparation of extract from Eclipta prostata by hot-soxlation method:

The coarse powdered material of whole plant of Eclipta prostate was subjected to hot- soxlation by placing 10 Kg of material in each soxlator using solvents n-hexane, acetone, ethyl alcohol, methanol, ethyl alcohol and water (1 : 1 ), methanol and water (1 : 1 ) and water at refluxing temperature of each solvent and recycled the process until extraction is completed, then plant extracts were filtered and concentrated to dryness on rotatory evaporator or on steam bath at optimum temperature.

All extracts such as n-hexane extract (EP-I), acetone extract (EP-2), ethyl alcohol extract (EP-3), methanol extract (EP-4), ethyl alcohol and water (1 : 1) extract (EP-5), methanol and water (1 : 1) extract (EP-6) and water extract (EP-7) prepared from the whole plant of Eclipta prostata by percolation method or hot-soxlation method were subjected to HPTLC (High Performance Thin Layer Chromatography) and HPLC (High performance Liquid chromatography) in various mobile phases on precoated TLC plates (Merck) and ODS column for qualitative and quantitative estimation of marker compounds and active principles.

Example 4

Preparation of extract from Eclipta prostata by Super Critical Fluid Extraction The shade dried material of whole plant of Eclipta prostata was pulverized to coarse powder and about 100 Kg of powdered material placed in a SCF extractor at the temperature of 40-50⁰C at high pressure of 300-350 bar using carbon dioxide as super critical fluid for extraction upto 4 to 6 hours and then the extract was collected in the collection vessel and evaporated at room temperature to remove any further residues of carbon dioxide. The extract thus obtained was free from any solvent residues and in highest pure form.

Example 5 Preparation of extract from Ecliota prostata by Enzyme extraction The shade dried material of whole plant of Eclipta prostata was pulverized to powder and about 10 Kg each of powdered material placed in Stainless Steel container and 0.5% to 5% of cellulase and pectinase enzyme combination was added in 4 volumes of distilled water. The enzyme extraction was processed at the temperature of 55⁰C to 60⁰C under occasional stirring upto 4-6 hours, and then plant extracts were filtered and concentrated to dryness on rotatory evaporator or on steam bath at optimum temperature and under reduced pressure.

Example 6

Preparation of extract from Glycyrrhiza glabra by percolation method: The shade dried material of glycyrrhiza glabra roots was pulverized to coarse powder and about 10 Kg each of powdered material placed in different percolators and extracted with n-hexane, acetone, ethyl alcohol, methanol, ethyl alcohol and water (1 : 1 ), methanol and water (1 : 1 ) and water at room temperature for 24 h to 48 h., then plant extracts were filtered and concentrated to dryness on rotatory evaporator or on steam bath at optimum temperature and under reduced pressure. ' ^!

Example 7

Preparation of extract from glycyrrhiza glabra by hot-soxlation method: The coarse powdered material of roots of glycyrrhiza glabra was subjected to hot- soxlation by placing 10 Kg of material in each soxlator using solvents n-hexane, acetone, ethyl alcohol, methanol, ethyl alcohol and water (1 : 1), methanol and water (1 : 1 ) and water at refluxing temperature of each solvent and recycled the process until extraction is completed, then plant extracts were filtered and concentrated to dryness on rotatory evaporator or on steam bath at optimum temperature.

All extracts such as n-hexane extract (GG-I), acetone extract (GG-2), ethyl alcohol extract (GG-3), methanol extract (GG-4), ethyl alcohol and water (1 : 1) extract (GG-5), methanol and water (1 : 1 ) extract (GG-6) and water extract (GG-7) prepared from the roots of glycyrrhiza glabra by percolation method or hot-soxlation method were subjected to HPTLC (High Performance Thin Layer Chromatography) and HPLC (High performance Liquid chromatography) in various mobile phases on precoated TLC plates (Merck) and ODS column for qualitative and quantitative estimation of marker compounds and active principles.

Example 8

Preparation of extract from glycyrrhiza glabra by Super Critical Fluid Extraction The shade dried material of roots of glycyrrhiza glabra was pulverized to coarse powder and about 100 Kg of powdered material placed in a SCF extractor at the temperature of 40-50⁰C at high pressure of 300-350 bar using carbon dioxide as super critical fluid for extraction upto 4 to 6 hours and then the extract was collected in the collection vessel and evaporated at room temperature to remove any further residues of carbon dioxide. The extract thus obtained was free from any solvent residues and in highest pure form.

Example 9

Preparation of extract from glycyrrhiza glabra by Enzyme extraction The shade dried material of roots of glycyrrhiza glabra was pulverized to powder and about 10 Kg each of powdered material placed in Stainless Steel container and 0.5% to 5% of cellulase and pectinase enzyme combination was added in 4 volumes of distilled water. The enzyme extraction was processed at the temperature of 55⁰C to 60⁰C under occasional stirring upto 4-6 hours, and then plant extracts were filtered and concentrated to dryness on rotatory evaporator or on steam bath at optimum temperature and under reduced pressure.

Example 10

Preparation of extract from Phyllanthus emblica by percolation method: The shade dried material of fruits of Phyllanthus emblica was pulverized to coarse powder and about 10 Kg each of powdered material placed in different percolators and extracted with n-hexane, acetone, ethyl alcohol, methanol, ethyl alcohol and water (1 : 1 ), methanol and water (1 : 1) and water at room temperature for 24 h to 48 h., then plant extracts were filtered and concentrated to dryness on rotatory evaporator or on steam bath at optimum temperature and under reduced pressure.

Example 1 1

Preparation of extract from Phyllanthus emblica by hot-soxlation method: The coarse powdered material of fruits of Phyllanthus emblica was subjected to hot-soxlation by placing 10 Kg of material in each soxlator using solvents n-hexane, acetone, ethyl alcohol, methanol, ethyl alcohol and water (1 : 1), methanol and water (1 : 1) and water at refluxing temperature of each solvent and recycled the process until extraction is completed, then plant extracts were filtered and concentrated to dryness on rotatory evaporator or on steam bath at optimum temperature.

All extracts such as n-hexane extract (PE-I), acetone extract (PE-2), ethyl alcohol extract (PE-3), methanol extract (PE-4), ethyl alcohol and water (1 : 1) extract (PE-5), methanol and water ( 1 : 1 ) extract (PE-6) and water extract (PE-7) prepared from the fruits of Phyllanthus emblica by percolation method or hot-soxlation method were subjected to HPTLC (High Performance Thin Layer Chromatography) and HPLC (High performance Liquid chromatography) in various mobile phases on precoated TLC plates (Merck) and ODS column for qualitative and quantitative estimation of marker compounds and active principles. ,

Example 12

Preparation of extract from Phyllanthus emblica by Super Critical Fluid Extraction The shade dried material of fruits of Phyllanthus emblica was pulverized to coarse powder and about 100 Kg of powdered material placed in a SCF extractor at the temperature of 40-50⁰C at high pressure of 300-350 bar using carbon dioxide as super critical fluid for extraction upto 4 to 6 hours and then the extract was collected in the collection vessel and evaporated at room temperature to remove any further residues of carbon dioxide. The extract thus obtained was free from any solvent residues and in highest pure form.

Example 13

Preparation of extract from Phyllanthus emblica by Enzyme extraction The shade dried material of fruits of Phyllanthus emblica was pulverized to powder and about 10 Kg each of powdered material placed in Stainless Steel container and 0.5% to 5% of cellulase and pectinase enzyme combination was added in 4 volumes of distilled water. The enzyme extraction was processed at the temperature of 55⁰C to 60⁰C under occasional stirring upto 4-6 hours, and then plant extracts were filtered and concentrated to dryness on rotatory evaporator or on steam bath at optimum temperature and under reduced pressure.

Example 14

Preparation of extract from Terminalia bellerica by percolation method: The shade dried material of fruits of Terminalia bellerica was pulverized to coarse powder and about LO Kg each of powdered material placed in different percolators and extracted with n-hexane, acetone, ethyl alcohol, methanol, ethyl alcohol and water (1 : 1), methanol and water (1 : 1 ) and water at room temperature for 24 h to 48 h., then plant extracts were filtered and concentrated to dryness on rotatory evaporator or on steam bath at optimum temperature and under reduced pressure.

Example 15

Preparation of extract from Terminalia bellerica by hot-soxlation method: The coarse powdered material of fruits of Terminalia bellerica was subjected to hot- soxlation by placing 10 Kg of material in each soxlator using solvents n-hexane, acetone, ethyl alcohol, methanol, ethyl alcohol and water (1 : 1), methanol and water (1 : 1) and water at refluxing temperature of each solvent and recycled the process until extraction is completed, then plant extracts were filtered and concentrated to dryness on rotatory evaporator or on steam bath at optimum temperature. All extracts such as n-hexane extract (TB-I), acetone extract (TB-2), ethyl alcohol extract (TB-3), methanol extract (TB-4), ethyl alcohol and water (1 :1) extract (TB-5), methanol and water (1 : 1) extract (TB-6) and water extract (TB-7) prepared from the fruits of Terminalia bβllerica by percolation method or hot-soxlation method were subjected to HPTLC (High Performance Thin Layer Chromatography) and HPLC (High performance Liquid chromatography) in various mobile phases on precoated TLC plates (Merck) and ODS column for qualitative and quantitative estimation of marker compounds and active principles.

Example 16

Preparation of extract from Terminalia bellerica by Super Critical Fluid Extraction The shade dried material of fruits of Terminalia bellerica was pulverized to coarse powder and about 100 Kg of powdered material placed in a SCF extractor at the temperature of 40-50⁰C at high pressure of 300-350 bar using carbon dioxide as super critical fluid for extraction upto 4 to 6 hours and then the extract was collected in the collection vessel and evaporated at room temperature to remove any further residues of carbon dioxide. The extract thus obtained was free from any solvent residues and in highest pure form.

Example 17

Preparation of extract from Terminalia bellerica by Enzyme extraction The shade dried material of Terminalia bellerica fruits was pulverized to powder and about 10 Kg each of powdered material placed in Stainless Steel container and 0.5% to 5% of cellulase and pectinase enzyme combination was added in 4 volumes of distilled water. The enzyme extraction was processed at the temperature of 55⁰C to 60⁰C under occasional stirring upto 4-6 hours, and then plant extracts were filtered and concentrated to dryness on rotatory evaporator or on steam bath at optimum temperature and under reduced pressure.

Example 18

Preparation of extract from Terminalia chebula by percolation method: The shade dried material of Terminalia chebula fruits was pulverized to coarse powder and about 10 Kg each of powdered material placed in different percolators and extracted with n-hexane, acetone, ethyl alcohol,¹ methanol, ethyl alcohol and water (1 : 1), methanol and water ( 1 : 1 ) and water at room temperature for 24 h to 48 h., then plant extracts were filtered and concentrated to dryness on rotatory evaporator or on steam bath at optimum temperature and under reduced pressure.

Example 19 Preparation of extract from Terminalia chebula by hot-soxlation method: The coarse powdered material Terminalia chebula fruits was subjected to hot- soxlation by placing 10 Kg of material in each soxlator using solvents n-hexane, acetone, ethyl alcohol, methanol, ethyl alcohol and water (1 :1), methanol and water (1 : 1) and water at refluxing temperature of each solvent and recycled the process until extraction is completed, then plant extracts were filtered and concentrated to dryness on rotatory evaporator or on steam bath at optimum temperature.

All extracts such as n-hexane extract (TC-I), acetone extract (TC-2), ethyl alcohol extract (TC-3), methanol extract (TC-4), ethyl alcohol and water (1 : 1) extract (TC-5), methanol and water (1 : 1 ) extract (TC-6) and water extract (TC-7) prepared from the fruits of Terminalia chebula by percolation method or hot-soxlation method were subjected to HPTLC (High Performance Thin Layer Chromatography) and HPLC (High performance Liquid chromatography) in various mobile phases on precoated TLC plates (Merck) and ODS column for qualitative and quantitative estimation of marker compounds and active principles.

Example 20

Preparation of extract from Terminalia chebula by Super Critical Fluid Extraction The shade dried material of fruits of Terminalia chebula was pulverized to coarse powder and about 100 Kg of powdered material placed in a SCF extractor at the temperature of 40-50⁰C at high pressure of 300-350 bar using carbon dioxide as super critical fluid for extraction upto 4 to 6 hours and then the extract was collected in the collection vessel and evaporated at room temperature to remove any further residues of carbon dioxide. The extract thus obtained was free from any solvent residues and in highest pure form.

Example 21

Preparation of extract from Terminalia chebula by Enzyme extraction The shade dried material of fruits of Terminalia chebula was pulverized to powder and about 10 Kg each of powdered material placed in Stainless Steel container and 0.5% to 5% of cellulase and pectinase enzyme combination was added in 4 volumes of distilled water. The enzyme extraction was processed at the temperature of 55⁰C to 60⁰C under occasional stirring upto 4-6 hours, and then plant extracts were filtered and concentrated to dryness on rotatory evaporator or on steam bath at optimum temperature and under reduced pressure.

Example'22

Preparation of extract from Cicer arietinum by percolation method: The shade dried material of seeds of Cicer arietinum was pulverized to coarse powder and about 10 Kg each of powdered material placed in different percolators and extracted with n-hexane, acetone, ethyl alcohol, methanol, ethyl alcohol and water (1 : 1 ), methanol and water ( 1 : 1 ) and water at room temperature for 24 h to 48 h., then plant extracts were filtered and concentrated to dryness on rotatory evaporator or on steam bath at optimum temperature and under reduced pressure.

Example 23

Preparation of extract from by hot-soxlation method:

The coarse powdered material of seeds of Cicer arietinum was subjected to hot- soxlation by placing 10 Kg of material in each soxlator using solvents n-hexane, acetone, ethyl alcohol, methanol, ethyl alcohol and water (1 : 1), methanol and water (1 : 1) and water at refluxing temperature of each solvent and recycled the process until extraction is completed, then plant extracts were filtered and concentrated to dryness on rotatory evaporator or on steam bath at optimum temperature.

All extracts such as n-hexane extract (CA-I), acetone extract (CA-2), ethyl alcohol extract (CA-3), methanol extract (CA-4), ethyl alcohol and water (1 : 1) extract (CA-5), methanol and water (1 : 1) extract (CA-6) and water extract (CA-7) prepared from the seeds of Cicer arietinum by percolation method or hot-soxlation method were subjected to HPTLC (High Performance Thin Layer Chromatography) and HPLC (High performance Liquid chromatography) in various mobile phases on precoated TLC plates (Merck) and ODS column for qualitative and quantitative estimation of marker compounds and active principles.

Example 24

Preparation of extract from seeds of Cicer arietinum by Super Critical Fluid Extraction

The shade dried material of seeds of Cicer arietinum was pulverized to coarse powder and about 100 Kg of powdered material placed in a SCF extractor at the temperature of 40- 50⁰C at high pressure of 300-350 bar using carbon dioxide as super critical fluid for extraction upto 4 to 6 hours and then the extract was collected in the collection vessel and evaporated at room temperature to remove any further residues of carbon dioxide. The extract thus obtained was free from any solvent residues and in highest pure form.

Example 25

Preparation of extract from Cicer arietinum by Enzyme extraction

The shade dried material of seeds of Cicer arietinum was pulverized to powder and about 10 Kg each of powdered material placed in Stainless Steel container and 0.5% to 5% of cellulase and pectinase enzyme combination was added in 4 volumes of distilled water. The enzyme extraction was processed at the temperature of 55⁰C to 60⁰C under occasional stirring upto 4-6 hours, and then plant extracts were filtered and concentrated to dryness on rotatory evaporator or on steam bath at optimum temperature and under reduced pressure.

Example 26

Preparation of extract from Lawsonia inermis by percolation method: The shade dried material of leaves of Lawsonia inermis of pulverized to coarse powder and about 10 Kg each of powdered material placed in different percolators and extracted with n-hexane, acetone, ethyl alcohol, methanol, ethyl alcohol and water (1 : 1), methanol and water (1 : 1) and water at room temperature for 24 h to 48 h., then plant extracts were filtered and concentrated to dryness on rotatory evaporator or on steam bath at optimum temperature and under reduced pressure .

Example 27

Preparation of extract from Lawsonia inermis by hot-soxlation method: The coarse powdered material of Lawsonia inermis leaves was subjected to hot- soxlation by placing 10 Kg of material in each soxlator using solvents n-hexane, acetone, ethyl alcohol, methanol, ethyl alcohol and water (1 : 1), methanol and water (1 : 1) and water at refluxing temperature of each solvent and recycled the process until extraction is completed, then plant extracts were filtered and concentrated to dryness on rotatory evaporator or on steam bath at optimum temperature.

All extracts such as n-hexane extract (LI-I), acetone extract (LI-2), ethyl alcohol extract (LI-3), methanol extract (LI-4), ethyl alcohol and water (1 : 1) extract (LI-5), methanol and water ( 1 : 1 ) extract (LI-6) and water extract (LI-7) prepared from the leaves Lawsonia inermis by percolation method or hot-soxlation method were subjected to HPTLC (High Performance Thin Layer Chromatography) and HPLC (High performance - Liquid chromatography) in various mobile phases on precoated TLC plates (Merck) and ODS column for qualitative and quantitative estimation of marker compounds and active principles.

Example 28

Preparation of extract from Lawsonia inermis by Super Critical Fluid Extraction

The shade dried material of Lawsonia inermis leaves was pulverized to coarse powder and about 100 Kg of powdered material placed in a SCF extractor at the temperature of 40-50⁰C at high pressure of 300-350 bar using carbon dioxide as super critical fluid for extraction upto 4 to 6 hours and then the extract was collected in the collection vessel and evaporated at room temperature to remove any further residues of carbon dioxide, The extract thus obtained was free from any solvent residues and in highest pure form.

Example 29

Preparation of extract from Lawsonia inermis by Enzyme extraction The shade dried material of leaves of Lawsonia inermis was pulverized to powder and about 10 Kg each of powdered material placed in Stainless Steel container and 0.5% to 5% of cellulase and pectinase enzyme combination was added in 4 volumes of distilled water. The enzyme extraction was processed at the temperature of 55⁰C to 60⁰C under occasional stirring upto 4-6 hours, and then plant extracts were filtered and concentrated to dryness on rotatory evaporator or on steam bath at optimum temperature and under reduced pressure.

Example 30

Preparation of extract from Carica papaya by percolation method: The shade dried material of Carica papaya fruits was pulverized to coarse powder and about 10 Kg each of powdered material placed in different percolators and extracted with n-hexane, acetone, ethyl alcohol, methanol, ethyl alcohol and water (1 : 1), methanol and water ( 1 : 1 ) and water at room temperature for 24 h to 48 h., then plant extracts were filtered and concentrated to dryness on rotatory evaporator or on steam bath at optimum temperature and under reduced pressure.

Example 31

Preparation of extract from Carica papaya by hot-soxlation method: The coarse powdered material of Carica papaya fruits was subjected to hot-soxlation by placing 10 Kg of material in each soxlator using solvents n-hexane, acetone, ethyl alcohol, methanol, ethyl alcohol and water (1 : 1), methanol and water (1 : 1 ) and water at refluxing temperature of each solvent and recycled the process until extraction is completed, then plant extracts were filtered and concentrated to dryness on rotatory evaporator or on steam bath at optimum temperature.

All extracts such as n-hexane extract (CP-I), acetone extract (CP-2), ethyl alcohol extract (CP-3), methanol extract (CP-4), ethyl alcohol and water (1 : 1) extract (CP-5), methanol and water (1 :1) extract (CP-6) and water extract (CP-7) prepared from the Carica papaya fruits by percolation method or hot-soxlation method were subjected to HPTLC (High Performance Thin Layer Chromatography) and HPLC (High performance Liquid chromatography) in various mobile phases on precoated TLC plates (Merck) and ODS column for qualitative and quantitative estimation of marker compounds and active principles. Example 32

Preparation of extract from Carica papaya by Super Critical Fluid Extraction The shade dried material of Carica papaya fruits was pulverized to coarse powder and about 100 Kg of powdered material placed in a SCF extractor at the temperature of 40- 50⁰C at high pressure of 300-350 bar using carbon dioxide as super critical fluid for extraction upto 4 to 6 hours and then the extract was collected in the collection vessel and evaporated at room temperature to remove any further residues of carbon dioxide. The extract thus obtained was free from any solvent residues and in highest pure form.

Example 33

Preparation of extract from Carica papaya by Enzyme extraction The shade dried material of Carica papaya fruits was pulverized to powder and about 10 Kg each of powdered material placed in Stainless Steel container and 0.5% to 5% of cellulase and pectinase enzyme combination was added in 4 volumes of distilled water. The enzyme extraction was processed at the temperature of 55 C to 60 C under occasional stirring upto 4-6 hours, and then plant extracts were filtered and concentrated to dryness on rotatory evaporator or on steam bath at optimum temperature and under reduced pressure.

Example 34

Preparation of extract from Sapindus mukurrosi by percolation method: The shade dried material of Sapindus mukurrosi fruits was pulverized to coarse powder and about 10 Kg each of powdered rrlaterial placed in different percolators and extracted with n-hexane, acetone, ethyl alcohol, methanol, ethyl alcohol and water (1 : 1), methanol and water (1 : 1 ) and water at room temperature for 24 h to 48 h, then plant extracts were filtered and concentrated to dryness on rotatory evaporator or on steam bath at optimum temperature and under reduced pressure.

Example 35

Preparation of extract from Sapindus mukurrosi by hot-soxlation method: The coarse powdered material of Sapindus mukurrosi fruits was subjected to hot- soxlation by placing 10 Kg of material in each soxlator using solvents n-hexane, acetone, ethyl alcohol, methanol, ethyl alcohol and water (1 : 1), methanol and water (1 : 1) and water at refluxing temperature of each solvent and recycled the process until extraction is completed, then plant extracts were filtered and concentrated to dryness on rotatory evaporator or on steam bath at optimum temperature. All extracts such as n-hexane extract (SM-I), acetone extract (SM-2), ethyl alcohol extract (SM-3), methanol extract (SM-4), ethyl alcohol and water (1 : 1) extract (SM-5), methanol and water (1 : 1) extract (SM-6) and water extract (SM-7) prepared from the Sapindus mukurrosi fruits by percolation method or hot-soxlation method were subjected to HPTLC (High Performance Thin Layer Chromatography) and HPLC (High performance Liquid chromatography) in various mobile phases on precoated TLC plates (Merck) and ODS column for qualitative and quantitative estimation of marker compounds and active principles.

Example 36

Preparation of extract from Sapindus mukurrosi by Super Critical Fluid Extraction The shade dried material of Sapindus mukurrosi fruits was pulverized to coarse powder and about 100 Kg of powdered material placed in a SCF extractor at the temperature of 40-50⁰C at high pressure of 300-350 bar using carbon dioxide as super critical fluid for extraction upto 4 to 6 hours and then the extract was collected in the collection vessel and evaporated at room temperature to remove any further residues of carbon dioxide. The extract thus obtained was free from any solvent residues and in highest pure form.

Example 37

Preparation of extract from Sapindus mukurrosi by Enzyme extraction The shade dried material of Sapindus mukurrosi fruits was pulverized to powder and about 10 Kg each of powdered material placed in Stainless Steel container and 0.5% to 5% of cellulase and pectinase enzyme combination was added in 4 volumes of distilled water. The enzyme extraction was processed at the temperature of 55⁰C to 60⁰C under occasional stirring upto 4-6 hours, and then plant extracts were filtered and concentrated to dryness on rotatory evaporator or on steam bath at optimum temperature and under reduced pressure.

Example 38

Preparation of extract from Cucumis sativus by percolation method: The shade dried material of fruits of Cucumis sativus was pulverized to coarse powder and about 10 Kg each of powdered material placed in different percolators and extracted with n-hexane, acetone, ethyl alcohol, methanol, ethyl alcohol and water (1 : 1), methanol and water ( 1 : 1 ) and water at room temperature for 24 h to 48 h, then plant extracts were filtered and concentrated to dryness on rotatory evaporator or on steam bath at optimum temperature and under reduced pressure. Example 39

Preparation of extract from Cucumis sativus by hot-soxlation method: The coarse powdered material of Cucumis sativus fruits was subjected to hot- soxlation by placing 10 Kg of material in each soxlator using solvents n-hexane, acetone, ethyl alcohol, methanol, ethyl alcohol and water (1 : 1), methanol and water (1 : 1) and water at retluxing temperature of each solvent and recycled the process until extraction is completed, then plant extracts were filtered and concentrated to dryness on rotatory evaporator or on steam bath at optimum temperature.

All extracts such as n-hexane extract (CS-I ), acetone extract (CS-2), ethyl alcohol extract (CS-3), methanol extract (CS-4), ethyl alcohol and water (1 : 1) extract (CS-5), methanol and water ( 1 : 1 ) extract (CS-6) and water extract (CS-7) prepared from the fruits of Cucumis sativus by percolation method or hot-soxlation method were subjected to HPTLC (High Performance Thin Layer Chromatography) and HPLC (High performance Liquid chromatography) in various mobile phases on precoated TLC plates (Merck) and ODS column for qualitative and quantitative estimation of marker compounds and active principles.

Example 40

Preparation of extract from Cucumis sativus by Super Critical Fluid Extraction The shade dried material of Cucumis sativus fruits was pulverized to coarse powder and about 100 Kg of powdered material placed in a SCF extractor at the temperature of 40- 50⁰C at high pressure of 300-350 bar using carbon dioxide as super critical fluid for extraction upto 4 to 6 hours and then the extract was collected in the collection vessel and ev aporated at room temperature to remove any further residues of carbon dioxide. The extract thus obtained was free from any solvent residues and in highest pure form.

Example- 41

Preparation of extract from Cucumis sativus by Enzyme extraction The shade dried material of Cucumis sativus fruits was pulverized to powder and about 10 Kg each of powdered material placed in Stainless Steel container and 0.5% to 5% of cellulase and pectinase enzyme combination was added in 4 volumes of distilled water. The enzyme extraction was processed at the temperature of 55⁰C to 60⁰C under occasional stirring upto 4-6 hours, and then plant extracts were filtered and concentrated to dryness on rotatory evaporator or on steam bath at optimum temperature and under reduced pressure.

Example 42 Preparation of extract Glycine max from by percolation method: The shade dried material of Glycine max seeds was pulverized to coarse powder and about 10 Kg each of powdered material placed in different percolators and extracted with n- hexane, acetone, ethyl alcohol, methanol, ethyl alcohol and water (1 : 1), methanol and water ( 1 : 1 ) and water at room temperature for 24 h to 48 h, then plant extracts were filtered and concentrated to dryness on rotatory evaporator or on steam bath at optimum temperature and under reduced pressure.

Example 43

Preparation of extract from Glycine max by hot-soxlation method:

The coarse powdered material of Glycine max seeds was subjected to hot-soxlation by placing 10 Kg of material in each soxlator using solvents n-hexane, acetone, ethyl alcohol, methanol, ethyl alcohol and water (1 : 1), methanol and water (1 : 1) and water at re fluxing temperature of each solvent and recycled the process until extraction is completed, then plant extracts were filtered and concentrated to dryness on rotatory evaporator or on steam bath at optimum temperature.

All extracts such as n-hexane extract (GM-I ), acetone extract (GM-2), ethyl alcohol extract (GM-3), methanol extract (GM-4), ethyl alcohol and water (1 :1) extract (GM-5), methanol and water (1 : 1) extract (GM-6) and water extract (GM-7) prepared from the Glycine max seeds by percolation method or hot-soxlation method were subjected to HPTLC (High Performance Thin Layer Chromatography) and HPLC (High performance Liquid chromatography) in various mobile phases on precoated TLC plates (Merck) and ODS column for qualitative and quantitative estimation of marker compounds and active principles.

Example 44

Preparation of extract from Glycine max by Super Critical Fluid Extraction

The shade dried material of Glycine max seeds was pulverized to coarse powder and about 100 Kg of powdered material placed in a SCF extractor at the temperature of 40-50⁰C at high pressure of 300-350 bar using carbon dioxide as super critical fluid for extraction upto 4 to 6 hours and then the extract was collected in the collection vessel and evaporated at room temperature to remove any further residues of carbon dioxide. The extract thus obtained was free from any solvent residues and in highest pure form.

Example 45

Preparation of extract from Glycine max by Enzyme extraction

The shade dried material of Glycine max seeds was pulverized to powder and about 10 Kg each of powdered material placed in Stainless Steel container and 0.5% to 5% of cellulase and pectinase enzyme combination was added in 4 volumes of distilled water. The enzyme extraction was processed at the temperature of 55 C to 60 C under occasional stirring upto 4-6 hours, and then plant extracts were filtered and concentrated to dryness on rotatory evaporator or on steam bath at optimum temperature and under reduced pressure.

Example 46

Preparation of extract Rosmarinus officinalis from by percolation method: The shade dried material of Rosmarinus officinalis leaves was pulverized to coarse powder and about 10 Kg each of powdered material placed in different percolators and extracted with n-hexane, acetone, ethyl alcohol, methanol, ethyl alcohol and water (1 : 1), methanol and water (1 : 1 ) and water at room temperature for 24 h to 48 h, then plant extracts were filtered and concentrated to dryness on rotatory evaporator or on steam bath at optimum temperature and under reduced pressure.

Example 47

Preparation of extract from Rosmarinus officinalis by hot-soxlation method: The coarse powdered material of Rosmarinus officinalis leaves was subjected to hot- soxlation by placing 10 Kg of material in each soxlator using solvents n-hexane, acetone, ethyl alcohol, methanol, ethyl alcohol and water (1 : 1), methanol and water (1 : 1 ) and water at refluxing temperature of each solvent and recycled the process until extraction is completed, then plant extracts were filtered and concentrated to dryness on rotatory evaporator or on steam bath at optimum temperature.

All extracts such as n-hexane extract (RO-I), acetone extract (RO-2), ethyl alcohol extract (RO-3), methanol extract (RO-4), ethyl alcohol and water (1 : 1) extract (RO-5), methanol and water (1 : 1) extract (RO-6) and water extract (RO-7) prepared from the Rosmarinus officinalis leaves by percolation method or hot-soxlation method were subjected to HPTLC (High Performance Thin Layer Chromatography) and HPLC (High performance Liquid chromatography) in various mobile phases on precoated TLC plates (Merck) and ODS column for qualitative and quantitative < estimation of marker compounds and active principles.

Example 48

Preparation of extract from Rosmarinus officinalis by Super Critical Fluid Extraction

The shade dried material of Rosmarinus officinalis leaves was pulverized to coarse powder and about 100 Kg of powdered material placed in a SCF extractor at the temperature of 40-50⁰C at high pressure of 300-350 bar using carbon dioxide as super critical fluid for extraction upto 4 to 6 hours and then the extract was collected in the collection vessel and evaporated at room temperature to remove, any further residues of carbon dioxide. The extract thus obtained was free from any solvent residues and in highest pure form.

Example 49

Preparation of extract from Rosmarinus officinalis by Enzyme extraction The shade dried material of Rosmarinus officinalis leaves was pulverized to powder and about 10 Kg each of powdered material placed in Stainless Steel container and 0.5% to 5% of cellulase and pectinase enzyme combination was added in 4 volumes of distilled water. The enzyme extraction was processed at the temperature of 55⁰C to 60⁰C under occasional stirring upto 4-6 hours, and then plant extracts were filtered and concentrated to dryness on rotatory evaporator or on steam bath at optimum temperature and under reduced pressure.

Example 50

Preparation of extract from Herbal Blend by percolation method: The shade dried material of herbal blend of whole plants of Eclipta prostata and/or glycyrrhiza glabra roots and/or phyllanthus emplica fruits and/or of terminalia bellerica seeds/fruits and/or terminalia chebula fruits and /or Cicer arietinum seeds and/or Lawsonia inermis leaves in the ratio of 17: 1 1 :20: 10:20: 17:05 respectively mixed and pulverized to coarse powder and about 10 Kg each of herbal blend placed in different percolators and extracted with n-hexane, acetone, ethyl alcohol, methanol, ethyl alcohol and water (1 : 1). methanol and water (1 : 1) and water at room temperature for 24 h to 48 h., then plant extracts were filtered and concentrated to dryness on rotatory evaporator or on steam bath at optimum temperature and under reduced pressure .

Example 51

Preparation of extract from Herbal Blend by hot-soxlation method: The shade dried material of herbal blend of whole plants of Eclipta prostate and/or glycyrrhiza glabra roots and/or phyllanthus emblica fruits and/or of terminalia bellerica seeds/fruits and/or terminalia chebula fruits and /or Cicer arietinum seeds and/or Lawsonia inermis leaves in the ratio of 17: 1 1 :20: 10:20:;17:05 respectively was subjected to hot- soxlation by placing 10 Kg of herbal blend in each soxlator using solvents n-hexane, acetone, ethyl alcohol, methanol, ethyl alcohol and water (1 : 1), methanol and water (1 : 1 ) and water at refluxing temperature of each solvent and recycled the process until extraction is completed, then plant extracts were filtered and concentrated to dryness on rotatory evaporator or on steam bath at optimum temperature. All extracts such as n-hexane extract (HB-IM), acetone extract (HB- 1\2), ethyl alcohol extract (HB- 1\3), methanol extract (HB- 1\4), ethyl alcohol and water (1 : 1) extract (HB-1 \5), methanol and water (1 : 1) extract (HB-1\6) and water extract (HB-1V7) prepared from the herbal blend by percolation method or hot-soxlation method were subjected to HPTLC (High Performance Thin Layer Chromatography) and HPLC (High performance Liquid chromatography) in various mobile phases on precoated TLC plates (Merck) and ODS column for qualitative and quantitative estimation of marker compounds and active principles.

Example 52

Preparation of extract from herbal blend by Enzyme extraction The shade dried material of herbal blend of whole plants of Eclipta prostata and/or Glycyrrhiza glabra roots and/or Phyllanthus emblica fruits and/or of Terminalia bellerica seeds/fruits and/or Terminalia chebula fruits and /or Cicer arietinum seeds and/or Lawsonia inermis leaves in the ratio of 17: 1 1 :20: 10:20: 17:05 respectively was pulverized to powder and about 10 Kg each of powdered herbal blend was placed in Stainless Steel container and 0.5% to 5% of cellulase and pectinase enzyme combination was added in 4 volumes of distilled water. The enzyme extraction was processed at the temperature of 55⁰C to 60⁰C under occasional stirring upto 4-6 hours, and then plant extracts were filtered and concentrated to dryness on rotatory evaporator or on steam bath at optimum temperature and under reduced pressure.

Example 53

Preparation of extract from herbal blend by Super Critical Fluid Extraction The shade dried material of herbal blend of whole plants of Eclipta prostata and/or glycyrrhiza glabra roots and/or phyllanthus emblica fruits and/or of terminalia bellerica seeds/fruits and/or terminalia chebula fruits and /or Cicer arietinum seeds and/or Lawsonia inermis leaves in the ratio of 17: 1 1 :20: 10:20: 17:05 respectively was pulverized to coarse powder and about 100 Kg of powdered herbal blend was placed in a SCF extractor at the temperature of 40-50⁰C at high pressure of 300-350 bar using carbon dioxide as super critical fluid for extraction upto 4 to 6 hours and then the extract was collected in the collection vessel and evaporated at room temperature to remove any further residues of carbon dioxide. The extract thus obtained was free from any solvent residues and in highest pure form.

Example 54 Preparation of extract from Herbal Blend by percolation method: The shade dried material of herbal blend of fruits of Carica papaya and/or Sapindus mukurrosi fruits and/or Cucumis sativus fruits and/or Glycine max seeds and/or Glycyrrhiza glabra roots and/or Rosmarinus officinalis leaves in the ratio of 20:20: 15:20: 15: 10 respectively mixed and pulverized to coarse powder and about 10 Kg each of herbal blend placed in different percolators and extracted with n-hexane, acetone, ethyl alcohol, methanol, ethyl alcohol and water (1 : 1 ), methanol and water (1 : 1) and water at room temperature for 24 h to 48 h., then plant extracts were filtered and concentrated to dryness on rotatory evaporator or on steam bath at optimum temperature and under reduced pressure .

Example 55

Preparation of extract from Herbal Blend by hot-soxlation method:

The shade dried material of herbal blend of fruits of Carica papaya and/or Sapindus mukurrosi fruits and/or Cucumis sativus fruits and/or Glycine max seeds and/or Glycyrrhiza glabra roots and/or Rosmarinus officinalis leaves in the ratio of 20:20: 15:20: 15: 10 respectively was mixed, powdered and subjected to hot-soxlation by placing 10 Kg of herbal blend in each soxlator using solvents n-hexane, acetone, ethyl alcohol, methanol, ethyl alcohol and water (1 : 1 ), methanol and water (1 : 1 ) and water at refluxing temperature of each solvent and recycled the process until extraction is completed, then plant extracts were filtered and concentrated to dryness on rotatory evaporator or on steam bath at optimum temperature.

All extracts such as n-hexane extract (HB-2\1), acetone extract (HB-2\2), ethyl alcohol extract (HB-2\3), methanol extract (HB-2\4), ethyl alcohol and water (1 : 1) extract (HB-2\5), methanol and water (1 : 1) extract (HB-2\6) and water extract (HB-2Y7) prepared from the herbal blend by percolation method or hot-soxlation method were subjected to HPTLC (High Performance Thin Layer Chromatography) and HPLC (High performance Liquid chromatography) in various mobile phases on precoated TLC plates (Merck) and ODS column for qualitative and quantitative estimation of marker compounds and active principles.

Example 56

Preparation of extract from herbal blend by Enzyme extraction

The shade dried material of herbal blend of fruits of Carica papaya and/or Sapindus mukurrosi fruits and/or Cucumis sativus fruits and/or Glycine max seeds and/or Glycyrrhiza glabra roots and/or Rosmarinus officinalis leaves in the ratio of 20:20: 15:20: 15: 10 respectively mixed and pulverized to coarse powder and about 10 Kg each of powdered herbal blend was placed in Stainless Steel container and 0.5% to 5% of cellulase and pectinase enzyme combination was added in 4 volumes of distilled water. The enzyme extraction was processed at the temperature of 55⁰C to 60⁰C under occasional stirring upto 4- 6 hours, and then plant extracts were filtered and concentrated to dryness on rotatory evaporator or on steam bath at optimum temperature and under reduced pressure.

Example 57

Preparation of extract from herbal blend by Super Critical Fluid Extraction The shade dried material of herbal blend of fruits of Carica papaya and/or Sapindus mukurrosi fruits and/or Cucumis sativus fruits and/or Glycine max seeds and/or Glycynhiza glabra roots and/or Rosmarinus officinalis leaves in the ratio of 20:20: 15:20: 15: 10 respectively mixed and pulverized to coarse powder and about 100 Kg of powdered herbal blend was placed in a SCF extractor at the temperature of 40-50⁰C at high pressure of 300- 350 bar using carbon dioxide as super critical fluid for extraction upto 4 to 6 hours and then the extract was collected in the collection vessel and evaporated at room temperature to remove any further residues of carbon dioxide. The extract thus obtained was free from any solvent residues and in highest pure form.

Evaluation of efficacy and safety of Protein Shampoo-Gentle Daily Care (SLS/SLES Free) An open, phase III trial.

Synopsis:

This study was an open, non-comparative, non-randomized, phase III clinical trial, conducted as per the ethical guidelines of Declaration of Helsinki and was approved by Institutional Ethics Committee. Fifty healthy subjects of both sexes, from the age group of 20-45 years, who were willing to give informed consent, were enrolled in the study. All subjects were advised to wash hair with Protein Shampoo-Gentle Daily Care (SLS/SLES Free) twice a week for a period of 4weeks. All subjects were followed for a period of 4 weeks. The predefined primary efficacy endpoints were reduction in hair fall, hair breakage and split ends. Also cleansing effect on hair and scalp, hair conditioning effect, and softness of hair were evaluated. The predefined secondary safety endpoints for short- and long-term were assessed by incidence of adverse events and compliance to the therapy. All adverse events reported or observed were recorded with information about severity, date of onset, duration and action taken regarding the study drug. Statistical analysis was done according to intention-to-treat principles. A total 50 subjects were enrolled in the study. There was a significant reduction (p<0.05) in the mean score for hair fall, hair breakage and split ends at the end of 4 weeks. There were no clinically significant adverse reactions, either reported or observed, during the entire study period and overall compliance to the treatment was excellent. Therefore, it may be concluded that, "Protein Shampoo-Gentle Daily Care (SLS/SLES Free)" is effective and safe. It is beneficial in reducing hair fall, hair breakage and split ends.

Aim of the study:

This pilot study was planned to evaluate the clinical efficacy and safety (short- and long-term) of "Protein Shampoo-Gentle Daily Care (SLS/SLES Free)".

Study Design:

This pilot study was an open, non-comparative, non-randomized, phase III clinical trial, as per the ethical guidelines of Declaration of Helsinki, from 26/05/2009 to 13/07/2009.

Primary and secondary endpoints:

The predefined primary efficacy endpoints were reduction in hair fall, hair breakage and split ends. The predefined secondary safety endpoints for short- and long-term were assessed by incidence of adverse events and compliance to the therapy.

Inclusion criteria:

Fifty healthy subjects of both sexes, from the age group of 20-45 years, who were willing to give informed consent, were enrolled in the study.

Exclusion criteria:

Subjects with systemic illness, endocrine disorders, dermatological disorder, subjects with known history or present condition of allergic response to any cosmetic/Pharmaceutical products, toiletries or its components or ingredients in the test products and those who refused to give informed consent, were excluded from the study. Pregnant, and lactating women were also excluded from the study.

Study procedure:

Pre Study

Before being entering in the study all subjects were given detailed description about the investigational product, nature and duration of the study. Also subject's responsibilities after entering, the study was explained. Subjects were pre screened for the criteria indicated in the Subject Selection section. Only subjects Who meet the requirements of this section, have signed an informed consent form, subjects who were ready to follow instructions given by the investigator and have an updated medical history on file with the investigator were entered into the study. Study period: All enrolled subjects underwent a thorough clinical examination with special emphasis on examination of scalp and hair. Demographic data of all subjects entered in the study is given in table 1.

Table-!

All subjects were advised to wash hair with Protein Shampoo-Gentle Daily Care (SLS/SLES Free) twice a week for a period of 4 weeks. All subjects were followed for a period of 4 weeks. Subjects were told to restrict themselves to the "Protein Shampoo-Gentle Daily Care (SLS/SLES Free)" and not to use any other shampoo till the study is over.

Follow-up and assessment: All subjects were followed for a period of 4 weeks and at each follow-up visit, they were asked about the frequency of the application of Protein Shampoo-Gentle Daily Care (SLS/SLES Free). Assessment of hair condition was done objectively (by doctor) and also subjectively (by subject). Thorough examination was done after completion of week 2 and at the end of the study.

Assessment of Hairfall was done on the basis of 6-point scale ranging from 1-6. 1 : Nil

2: 1 -5 hair falls only once during combing. 3: 1 -5 hair falls each time during combing. 4: 6-10 hair falls each time during combing. 5: 10-30 hair falls each time during combing. 6: More than 30 hairs falls each time during combing.

Assessment of split ends was done on the basis of 4-point scale ranging from 1-4. I : Normal healthy hair. 2: less than 50% of hair with split ends. 3 : 50% of hair with split ends. 4: more than 50% of hair with split ends.

Assessment of hair breakages was done on the basis of 4-point scale ranging from 1 - 4.

1 : No hair breakages.

2: less than 10 hair breaks each time during combing. 3 : 10-30 hair breaks each time during combing. 4: more than 30 hair breaks each time during combing.

Assessment of cleansing of hair and scalp was done on the basis of 4-point scale ranging from 1 -4. 1 : Normal healthy scalp. 2: Minimal oily scalp and hair. 3: Moderately oily scalp and hair. 4: Very oily, itchy scalp and hair.

Assessment of hair conditioning and softness was done on the basis of 4-point scale ranging from 1-4. 1 : Normal healthy hair. 2: Minimal dry and dull hair. 3: Moderately dry and dull hair. 4: Very dry and dull hair.

Statistical analysis:

Statistical analysis was done according to intention-to-treat principles. The changes in various parameters from baseline values and the values after 2 weeks were evaluated by Repeated measures of ANOVA using Friedman Test followed by Dunnett's Multiple Test, p value of <0.05 was considered significant.

Results:

All subjects completed the study; no one was withdrawn from the study.

There was a significant reduction (p<0.05) in the mean score for hairfall, split ends and hair breakages. In subjective evaluation, majority of subjects experienced remarkable overall improvement in hair conditioning and softness. Also cleansing of hair and scalp was observed at the end of 4- week therapy (Table 2).

Table 2

Adverse Effects:

There were no clinically significant adverse reactions, either reported or observed, during the entire study period (table 3) and overall compliance to the treatment was excellent.

Table

Dermal Safety Evaluation of Protein Shampoo-Gentle Daily Care (SLS/SLESFree)

Days of application

Signs & Symptoms

Initial Week l Week 2

Conclusion:

Present study indicates good clinical efficacy of Protein Shampoo-Gentle Daily Care (SLS/SLES Free). It is beneficial in reducing hairfall, split ends and hair breakages.

While this invention has been described in detail with reference to certain preferred embodiments, it should be appreciated that the present invention is not limited to those precise embodiments. Rather, in view of the present disclosure, which describes the current best mode for practicing the invention, many modifications and variations would present themselves to those skilled in the art without departing from the scope and spirit of this invention.

Claims

We Claim:

1 . A herbal cleanser composition comprising extracts of herbs and surfactant selected from non-ionic surfactant and non-ethoxylated anionic surfactant and pharmaceutically acceptable excipients.

2. The composition according to claim 1 , wherein said extract of herbs is obtained from a blend of herbs selected from Eclipta prostate, Glycyrrhiza glabra, Phyllanthus emblica, Terminalia bellarcia, Terminalia chebula, Cicer arietinum, Lawsonia inermis, Carica papaya, Sapindus mukurrosi, Cucumis sativus, Glycine max, Rosmarinus officinalis and in various combination thereof.

3. The composition according to claim 2, wherein the blend is used in the range of O.OOOI % to 50%.

4. The composition according to claim 1 , wherein the blend of herbs is comprising Eclipta prostate, Glycyrrhiza glabra, Phyllanthus emblica, Terminalia bellarcia, Terminalia chebula, Cicer arietinum and Lawsonia inermis.

5. The composition according to claim 1 , wherein the blend of herbs is comprising Carica papaya, Sapindus mukurrosi, Cucumis sativus, Glycine max, Glycyrrhiza glabra and Rosmarinus officinalis

6. The composition according to claim 1, wherein the composition is formulated in various kinds of cleansers.

7. The composition according to claim 6, wherein the composition is formulated in a shampoo formulation.

8. The composition according to claim 1, wherein said composition is free of paraben components.

9. The composition according to claim 1, wherein said composition is devoid of using silicones.

10. The composition according to claim 1 , wherein said composition is prepared by the method comprising blending the herbs and extracting the resultant herbal blend employing super critical fluid extraction (SCFE), mixing the resultant extract of blend with non-ionic surfactant and non-ethoxylated anionic surfactant and pharmaceutically acceptable excipients.

1 1 . The composition according to claim 1 , wherein said composition is prepared by the method comprising blending the herbs and extracting the resultant herbal blend employing percolation or soxhalation using water, mixing the resultant extract of blend with non-ionic surfactant and non-ethoxylated anionic surfactant and pharmaceutically acceptable excipients.

12 The composition according to claim 1 , wherein said composition is prepared by the method comprising blending the herbs and extracting the resultant herbal blend employing enzymes, mixing the resultant extract of blend with non-ionic surfactant and non- ethoxylated anionic surfactant and pharmaceutically acceptable excipients.