WO2012160569A1 - "process for extraction of ashwagandha (withania somnifera) roots" - Google Patents

"process for extraction of ashwagandha (withania somnifera) roots" Download PDF

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Publication number
WO2012160569A1
WO2012160569A1 PCT/IN2011/000894 IN2011000894W WO2012160569A1 WO 2012160569 A1 WO2012160569 A1 WO 2012160569A1 IN 2011000894 W IN2011000894 W IN 2011000894W WO 2012160569 A1 WO2012160569 A1 WO 2012160569A1
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Prior art keywords
withanoside
roots
extraction
followed
ashwagandha
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PCT/IN2011/000894
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French (fr)
Inventor
Jayesh Panalal CHOKSI
Pranav Jayesh CHOKSI
Mafatlal Tribhovandas DAVE
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Gufic Biosciences Limited
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Priority to IN1556MU2011 priority Critical
Priority to IN1556/MUM/2011 priority
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Publication of WO2012160569A1 publication Critical patent/WO2012160569A1/en

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    • AHUMAN NECESSITIES
    • A61MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61KPREPARATIONS FOR MEDICAL, DENTAL, OR TOILET PURPOSES
    • A61K36/00Medicinal preparations of undetermined constitution containing material from algae, lichens, fungi or plants, or derivatives thereof, e.g. traditional herbal medicines
    • A61K36/18Magnoliophyta (angiosperms)
    • A61K36/185Magnoliopsida (dicotyledons)
    • A61K36/81Solanaceae (Potato family), e.g. tobacco, nightshade, tomato, belladonna, capsicum or jimsonweed

Abstract

Discloses a process for extraction of Withanoside IV and Withanoside V from Ashwagandha (Withania somniferd) roots and a pharmaceutical composition containing the Withanosides IV and V along with pharmaceutically acceptable excipients, for the treatment of relieving anxiety and stress.

Description

PROCESS FOR EXTRACTION OF ASHWAGANDHA
(WITHANIA SOMNIFERA) ROOTS"
TECHNICAL FIELD OF THE INVENTION;
The present invention relates to a process for extraction of active constituents from Ashwagandha (Withania somnifera) roots. The present invention further relates to a pharmaceutical composition comprising active constituents from Ashwagandha (Withania somnifera) roots along with pharmaceutically acceptable excipient, useful for the treatment of relieving anxiety and stress.
BACKGROUND OF THE INVENTION:
Withania somnifera, also known as Ashwagandha, Indian ginseng, Winter cherry, is a plant in the Solanaceae or nightshade family. It grows as a short shrub (35-75 cm) with a central stem from which branch extend radially in a star pattern (stellate) and covered with a dense matte of wooly hairs (tomentose). The flowers are small and green, while the ripe fruit is orange-red and has milk-coagulating properties. The plant also has long brown tuberous roots that are used for medicinal purposes. It is cultivated in many of the drier regions of India such as Manasa, Neemuch, and Jawad tehsils of the Mandsaur District of Madhya Pradesh, Punjab, Sind and Rajasthan.
In Ayurveda, the roots of W. somnifera are used to prepare many herbal medicines. It is claimed to possess aphrodisiac, sedative, rejuvenative and life prolonging properties. It is traditionally used to treat the symptoms and conditions associated with chronic fatigue, dehydration, bone weakness, muscle weakness and tension, loose teeth, thirst, impotency, premature ageing, emaciation, debility, constipation, senility, rheumatism, nervous exhaustion, memory loss, neurodegenerative disorders, spermatorrhoea.
US Patent Publication No. US20100285064 discloses a process for preparing a Withania somnifera fraction rich in withanolides and a vaccine comprising a " Withania somnifera fraction" as an adjuvant. US Patent No. 6153198 discloses a high purity Withania somnifera extract composition, process for obtaining extract composition and pharmaceutical and nutritional use products thereof.
US Patent No. 7108870 discloses an improved process of analytical and quantitative isolation of withaferin-A from Withania somnifera.
WIPO Publication No. WO2010013254 (corresponding of Indian Patent Application No. 1775/MUM/2008) relates to a Withania somnifera plant extract, composition comprising the extract and process for preparation of the extract.
Indian Patent Application No. 1283/MUM/2009 describes a method of extraction from Withania somnifera (WS) and one or more fractions containing the pharmacologically active ingredients extracted from WS using the method of extraction.
Roots of Ashwagandha contains steroidal Lactones, Withanosides I, II, III, Glycowithanolides A, D, E, F, G, H, I, J, K, L, M, WS-1 P and S, alkaloids like withanone, withaferin A, withasomine, somniferine, withanine, cuscohygrine, anhygrine, tropine, pseudotrophine, anaferine, choline, tropanol, pseudotropanol, isopelletriene, withasomnine, starch etc.
The taste of roots used to be bitter and sweet astringent. Roots used to be 1/3 to 1/2 m long, thin to thick whitish brown from outside, creamish from inside. Withania somnifera is used in many diseases because its constituents have good effect on immune system, nervous system, reproductive system, circulatory system, respiratory system, digestive system, excretory/ urinary system etc.
The quantity/amount of desired/active Withanosides will be more when compared to the same amount of crude drug (root) powder. Moreover, crude drug powder loses its effect in 6 to 12 months, whereas extract remain stable for 5 to 7 years, even if stored at room temperature. Hence, the present inventors have come up with a process wherein the enriched extract of Ashwagandha (Withania somnifera) roots containing Withanosides IV and Withanosides V can be achieved. The invention further discloses a pharmaceutical composition comprising Withanosides IV and Withanosides V along with pharmaceutically acceptable excipients, useful for relieving anxiety and stress.
OBJECT OF THE INVENTION:
Thus, the object of the present invention is to provide a process for extraction of active constituents i.e. Withanosides IV and Withanosides V from Ashwagandha (Withania somnifera) roots and a pharmaceutical composition comprising said Withanosides, along with pharmaceutically acceptable excipients, useful for relieving anxiety and stress.
SUMMARY OF THE INVENTION:
The present invention discloses a process for extraction of Withanosides JV and Withanosides V from Ashwagandha (Withania somnifera) roots, comprising the following six major steps:-
1. Sorting and sieving of roots, to remove dust & contaminants
2. Pulverization of roots
3. Cold extraction and filtration
4. Hot extraction and filtration
5. Concentration of extract
6. Drying of extract
The present invention further discloses the pharmaceutical composition comprising Withanosides IV and Withanosides V, along with pharmaceutically acceptable excipients, useful for relieving anxiety and stress.
BRIEF DESCRIPTION OF FIGURE:
FIG 1 : Flow chart for Extraction of Ashwagandha. DETAILED DESCRIPTION OF THE INVENTION:
The invention will now be described in detail in connection with certain preferred and optional embodiments, so that various aspects thereof may be more fully understood and appreciated.
The plant material used in the present invention i.e. Ashwagandha {Withania somnifera) roots are collected from northern, western and central parts of India.
In an embodiment, the present invention provides a process for extraction of active constituents i.e. Withanoside IV and Withanoside V from Ashwagandha {Withania somnifera) roots. The extract is standardized to contain Withanoside IV and Withanoside
V within the range of not less than 0.8 % and not less than 0.4 % respectively.
In a preferred embodiment, a process for extraction of Withanoside IV and Withanoside
V from Ashwagandha {Withania somnifera) roots comprises following steps^
a) Sorting and sieving of roots to remove dust and contaminants;
b) pulverizing of roots to obtain pulverized powder;
c) cold extraction of pulverized powder of step (b) with polar solvent followed by centrifuging to obtain the residue;
d) subjecting to hot extraction of residue obtained in step (c) by adding it to polar solvent followed by refluxing and centrifuging;
e) collecting the filtrates of step (c) and step (d) after centrifugation, followed by distillation to obtain an extract;
f) adding and mixing methyl paraben IP, Propyl paraben IP, dicalcium phosphate anhydrous IP and Colloidal silicone Dioxide IP to the extract of step (e) to obtain pasty mass;
g) transferring the pasty mass of step (f) to evaporator, followed by stirring and heating to obtain granular powder of 30 to 60 mesh.
Accordingly, in another preferred embodiment, the present invention provides a process for extraction of active constituents i.e. Withanoside IV and Withanoside V from Ashwagandha {Withania somnifera) roots, comprises the following steps:- Step 1: Sorting and sieving of roots to remove dust and contaminants
Ashwagandha roots are cut into small pieces and spread on SS sieve of size 10 mesh to 30 mesh. The contaminants, infected roots or adhered dust etc. from the roots are removed by sieving or simply by hand picking. After sorting and sieving, roots are filled in clean bag for Pulverization.
Step 2: Pulverization of roots
Suitable commercial pulverization machine with 30 to 60 mesh sieve is used for pulverization and plastic bag is used for collecting the pulverized material.
Step 3: Cold extraction and filtration
Mixing and stirring fixed quantity of pulverized Ashwagandha powder of Step 2 with sufficient quantity of polar solvent in first vessel. Heating the mixture from RT to below boiling point and maintain there for a period of 8 to 15 hours with continuous stirring. Cool the mixture at room temperature followed by centrifugation (I) to obtained residue. The filtrate of centrifuge (I) is taken in a second vessel for concentration. The residue is charged to first vessel for hot extraction.
Step 4: Hot extraction and filtration
The residue obtained from Step 3 is added to the first vessel containing enough of polar solvent (as used before) (30% recovered polar solvent and 70% fresh polar solvent, wherein the recovered polar solvent is from Ashwagandha of previous batch), followed by heating and refluxing at from 60°C to 100°C over a period of 1 to 4 hours. The mixture is cooled to R.T and discharged the mass in SS tank followed by centrifugation (11). After centrifuge, the filtrate is added to the second vessel of Step 3.
Step 5: Concentration of extract
The filtrates of Step 3 and Step 4 are collecting in second vessel and heating slowly and further raising the temperature of mixture to distill out polar solvent, followed by cooling to R.T. Discharging pasty mass in previously weighed SS storage tank. The test sample is sent for quality control and to the extract obtained is added methyl paraben IP, Propyl paraben IP and mixed well followed by addition of Dicalcium phosphate anhydrous IP and Colloidal silicon dioxide IP in the quantity based on analysis of test sample and total quantity of the extract, and further mixing well to get the pasty mass.
Step 6: Drying of extract
Transferring the pasty mass of Step 5 to evaporator, followed by stirring and heating from RT to 100°C, till it is converted into granular powder. Cool to R.T. Passing the granular powder from pulvarizer or multimill, to get the final dry powder of 30 to 60 mesh.
The polar solvent used in the abovementioned process is selected from lower alcohols such as methanol, ethanol, acetone, isopropyl alcohol etc., ketones such as acetone, methyl ethyl ketone, diethyl ketone, ethyl butyl ketone etc., ethers such as dimethyl ether, diethyl ether, tetrahydrofuran (THF) etc.
The polar solvent used in the abovementioned process is preferably methanol.
The desired percentages of Withanoside IV and Withanoside V extracted from Ashwagandha (Withania somniferd) roots are NLT 0.8 % and NLT 0.4 % respectively.
In another embodiment of the present invention, the active constituents Withanoside IV and Withanoside V, along with pharmaceutically acceptable excipients are formulated into pharmaceutical composition and oral dosage form.
Accordingly, the pharmaceutically acceptable excipients are selected from the group comprising diluents, binders, wetting agents, disintegrants and lubricants.
Suitable excipients such as Lactose, Starch, Sodium starch glycolate, AC.Disol, Talcum, Magnesium stearate etc. can be used.
In another embodiment of the present invention, the active constituents Withanoside IV and Withanoside V are mixed with other suitable diluents and disintegrants; followed by granulation with appropriate binding agent, dried, lubricated and compressed into tablet or capsule. Accordingly, pharmaceutical composition of the present invention is in the form of tablet, powder, syrup or capsule.
In another embodiment of the present invention, the pharmaceutical compositions comprising Withanoside IV and Withanoside V are used for relieving anxiety and stress.
In another embodiment of the present invention, the amounts of active constituents such as Withanoside IV and Withanoside V are analyzed by using HPLC method. The results are shown in Table 1.
In another embodiment of the present invention, the presence of residual polar solvent, in final dried extract is analyzed by using GC method. The specifications of residual solvents are shown in Table 2.
In another embodiment of the present invention, trial batches using various polar solvents were analyzed for the presence of Withanoside IV and Withanoside V. The results are shown in Table 3.
Table 1 : Results of HPLC Analysis:
Figure imgf000009_0001
Table 3:
Figure imgf000010_0001
EXAMPLES:-
The following example, which includes preferred embodiments, will serve to illustrate the practice of this invention, it being understood that the particulars shown are by way of example and for purpose of illustrative discussion of preferred embodiments of the invention.
Example 1: Extraction of Ashwagandha roots
Ashwagandha roots were cut into small pieces and spread on 20 mesh SS sieve to remove dust and contaminants. Pulverize the said roots through pulverization machine with 50 mesh sieve to obtain pulverized powder. Mixed and stirred the said pulverized powder with methanol and heated to about 40°C for 12 hours with continuous stirring. Cool the mixture at R.T and centrifuged (1) it, to obtain the residue. Then subjecting to hot extraction of the residue by adding methanol, followed by heating and refluxing at 95°C for 2 hrs. The mixture is cooled to R.T and discharged the mass followed by centrifugation (II). Collected both the filtrates and heated to distill out methanol, followed by cooling to R.T. to obtain an extract. Added and mixed methyl paraben IP, Propyl paraben IP, dicalcium phosphate anhydrous IP and Colloidal silicone Dioxide IP to the extract and mixed well to obtain pasty mass. Transferred the said pasty mass to evaporator, followed by stirring and heating to 90 - 95°C, till it is converted into granular powder. Passed the granular powder from pulvarizer or multimill, to get the final dry powder of 50 to 60 mesh. Specification of Ashwagandha extract:
Figure imgf000011_0001
Results of 3 trials based on HPLC analysis are summarized as follows:
Figure imgf000011_0002
Solvent by (Limit: NLT 3000ppm) (Limit: NLT 3000ppm) (Limit: NLT 3000ppm) GC
7. Withanoside 1.50% 1.31% 1.15%
(a) IV (Limit: NLT 0.8%) (Limit: NLT 0.8%) (Limit: NLT 0.8%)
7. Withanoside 0.92% 0.82% 0.91%
(b) V (Limit: NLT 0.4) (Limit: NLT 0.4) (Limit: NLT 0.4)
8. Arsenic Less than 7ppm Less than 7ppm Less than 7ppm
(Limit: Less than 7ppm) (Limit: Less than 7ppm) (Limit: Less than 7ppm
9. Heavy metal ί Less than 50ppm Less than 50ppm Less than 50ppm
(Limit: Less than 50 ppm) (Limit: Less than 50 ppm) (Limit: Less than 50 ppm)
10. Microbial Complies as per Report No. Complies as per Report Complies as per Report Limit Test MGB/BED/430 No. MGB/BED/431 No. MGB/BED/432
(Limit: (Limit: (Limit:
Total Aerobic count: Total Aerobic count: Total Aerobic count:
<1000cfu/gm <1000cfu/gm <1000cfu/gm Yeast and mound: Yeast and mound: Yeast and mound: <100cfu/gm <100cfu/gm <100cfu/gm Coliforms: Should be Coliforms: Should be Coliforms: Should be negative negative negative
Salmonella/Shigolla/E.coli Salmonella/Shigolla/E.coli Salmonella/Shigolla/E.coli Should be negative Should be negative Should be negative
Example 2: Extraction of Ashwagandha roots
Ashwagandha roots were cut into small pieces and spread on 20 mesh SS sieve to remove
dust and contaminants. Pulverize the said roots through pulverization machine with 50
mesh sieve to obtain pulverized powder. Mixed and stirred the said pulverized powder
with ethanol and heated to about 40°C for 12 hours with continuous stirring. Cool the
mixture at R.T and centrifuged (I) it, to obtain the residue. Then subjecting to hot
extraction of the residue by adding ethanol, followed by heating and refluxing at 95°C for
2 hrs. The mixture is cooled to R.T and discharged the mass followed by centrifugation
(II). Collected both the filtrates and heated to distill out ethanol, followed by cooling to
R.T. to obtain an extract. Added and mixed methyl paraben IP, Propyl paraben IP,
dicalcium phosphate anhydrous IP and Colloidal silicone Dioxide IP to the extract and
mixed well to obtain pasty mass. Transferred the said pasty mass to evaporator, followed
by stirring and heating to 90 - 95°C, till it is converted into granular powder. Passed the
granular powder from pulvarizer or multimill, to get the final dry powder of 50 to 60
mesh.

Claims

We claim,
1. A process for extraction of Withanoside IV and Withanoside V from Ashwagandha (Withania sotnnifera) roots comprises following steps:- a) Sorting and sieving of roots to remove dust and contaminants;
b) pulverizing of roots to obtain pulverized powder;
c) cold extraction of pulverized powder of step (b) with polar solvent followed by centrifuging to obtain the residue;
d) subjecting to hot extraction of residue obtained in step (c) by adding it to polar solvent followed by refluxing and centrifuging;
e) collecting the filtrates of step (c) and step (d) after centrifugation, followed by distillation to obtain an extract;
f) adding and mixing methyl paraben IP, Propyl paraben IP, dicalcium phosphate anhydrous IP and Colloidal silicone Dioxide IP to the extract of step (e) to obtain pasty mass;
g) transferring the pasty mass of step (f) to evaporator, followed by stirring and heating to obtain granular powder of 30 to 60 mesh.
2. The process for extraction of Withanoside IV and Withanoside V as claimed in claim 1 , wherein said polar solvent is selected from lower alcohols such as methanol, ethanol, acetone, isopropyl alcohol etc., ketones such as acetone, methyl ethyl ketone, diethyl ketone, ethyl butyl ketone etc., ethers such as dimethyl ether, diethyl ether, tetrahydrofuran (THF) etc.
3. The process for extraction of Withanoside IV and Withanoside V as claimed in claim 1, wherein said polar solvent is preferably methanol.
4. The pharmaceutical composition comprising Withanoside IV and Withanoside V obtained by process according to claim 1, along with pharmaceutically acceptable excipients.
5. The pharmaceutical composition according to claim 4, wherein said composition is formulated into oral dosage form.
6. The pharmaceutical composition according to claim 5, wherein said oral composition is in the form of tablet, powder, syrup or capsule.
PCT/IN2011/000894 2011-05-23 2011-12-26 "process for extraction of ashwagandha (withania somnifera) roots" WO2012160569A1 (en)

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Cited By (2)

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WO2017068600A1 (en) 2015-10-22 2017-04-27 Benny Antony A process to enhance the bioactivity of ashwagandha extracts
US9987323B2 (en) 2015-10-22 2018-06-05 Benny Antony Process to enhance the bioactivity of Ashwagandha extracts

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Cited By (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2017068600A1 (en) 2015-10-22 2017-04-27 Benny Antony A process to enhance the bioactivity of ashwagandha extracts
US9987323B2 (en) 2015-10-22 2018-06-05 Benny Antony Process to enhance the bioactivity of Ashwagandha extracts
US10166266B2 (en) 2015-10-22 2019-01-01 Benny Antony Process to enhance the bioactivity of ashwagandha extracts
US10251927B2 (en) 2015-10-22 2019-04-09 Benny Antony Process to enhance the bioactivity of Ashwagandha extracts
US20210113646A1 (en) * 2015-10-22 2021-04-22 Benny Antony Process to enhance the bioactivity of ashwagandha extracts

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