A Comparative study for quality assessment of various location samples of Withania somnifera Dunal.

 

Ajay Kumar Meena2, Brijendra Singh1*, Vijay Simha,             V. Gautam1 and M. M. Rao2

1School of Pharmaceutical Sciences, Shobhit University, Meerut, UP, (India)

2National Institute of Ayurvedic Pharmaceutical Research, Patiala – 147001 (Punjab)

 

ABSTRACT:

Ashwagandha is a reputed drug mentioned in the ancient books of Ayurveda and Siddha for the treatment of stress, hypertension, rejuvenative, sound sleep, brain functions and reducing the high cholesterol levels. It is a stout shrub that reaches a height of 170 cm (5.6 ft). For the standardization of this drug Physico-chemical and Phyto-chemical parameters were carried out such as moisture content, ash values, pH values, extractability in water and alcohol were carried out. Thin Layer Chromatography studies were also carried out to ascertain the quality of this drug. The present study summarizes the difference in two places of Withania somnifera Dunal. This study also helps to identify the right place for cultivation of medicinal plants and quality assessment.

 

KEYWORDS: Withania somnifera Dunal, Ayurveda, Physicochemical, Standardization, Quality

 

INTRODUCTION:

Withania somnifera Linn. also known as Ashwagandha, Indian ginseng, Winter cherry, Ajagandha, Kanaje Hindi, Amukkuram in Malayalam and Samm, is a plant in Solanaceae or nightshade family and is commonly used in Indian traditional health care systems. The plant is said to have a potential property of pacifying ‘Vata’ in herbal drugs compared therapeutic value of its roots with Panax ginseng 1-3. The genus Withania is an important member of the family Solanaceae. Twenty-three species of the genus Withania have been reported of these, only two Withania somnifera (Linn.) Dunal and Withania coagulans (Linn.) Dunal have been reported from India.

 

The dried roots of the shrub, Withania somnifera, commonly known as ‘Ashwagandha’, are important ingredients in Ayurvedic medicine. Roots of Withania contain several alkaloids of medicinal value. These include 13 Dragendroff-positive compounds, withasomine and visamine 4. It is found throughout the drier parts of India, it is widely grown in the provinces of Madhya Pradesh, Uttar Pradesh, plains of Punjab and northwestern parts of India like Gujarat and Rajasthan 5-7. Wild Withania somnifera genotypes may possess genes important for the development of new varieties of this important species 8.

 

The stems are around 3 to 4 feet in height. One plant survives for up to 4 to 5 years. The leaves are bowl shaped, small and without thorns. It remains green for 12 months near big trees and ponds9. The roots are rough, white from within, strong, transparent, thick and one to one and half feet long. The seeds are sown in rainy season and harvested in the spring season.10,11

It is an ingredient in many formulations prescribed for a variety of musculoskeletal conditions (e.g., arthritis, rheumatism), and as a general tonic to increase energy, improve overall health and longevity, and prevent disease in athletes, the elderly, and during pregnancy12-15. Also used in Anxiety and Depression16-18, Chronic Stress18-20.

 

MATERIALS AND METHODS:

Plant material was collected in Sep. 2009 from Gwalior M.P. (Sample- 1) and Patiala, Punjab (Sample- 2) and specimen was identified and authenticated at the Pharmacognocy Laboratory, National Institute of Ayurvedic Pharmaceutical Research (NIAPR), Patiala (Punjab).

 

RESULT AND DISCUSSION:

Analysis of Various Parameters:

Different physicochemical parameters like Total ash, acid insoluble ash, water soluble ash, ethanol soluble extractive value, water soluble extractive value, loss on drying, pH, TLC and Phyto-chemical (Chemical Test) of both sample Withania somnifera Dunal was determined by different methods Table 1.

 

Physicochemical parameters:

Total ash:

Weigh 2g of the sample accurately in a previously ignited and tarred Silica dish. Spread the material evenly and ignite in a muffle furnace by gradually increasing the temperature to 600oC until it is white, indicating the absence of carbon. Cool the dish in desiccators and weigh. If carbon free ash cannot be obtained in this manner, cool the dish and moisten the residue with about 2 ml of water or a saturated solution of Ammonium nitrate. Dry on a water-bath, and then ignite in the muffle furnace to constant weight. Cool the dish in desiccators for 30 minutes, and then weigh. Calculate the percentage of total ash of air-dried material.

 

Acid insoluble ash:

Find out the total ash of the sample as described under determination of Ash. To the dish Containing the total ash, add 25ml of 1:5 Hydrochloric acid, cover with a watch glass and boil gently for 5 minutes. Rinse the watch glass with 5 ml of hot water and add the washings to the dish. Collect the insoluble matter on an ash less filter paper (Whatman No. 41) and wash with hot water until the residue is free from acid. Transfer the filter paper containing the insoluble matter to the original dish, dry and ignite to constant weight. Cool the dish in desiccators for 30 minutes, and then weigh. Calculate the percentage of Acid insoluble - ash of the air-dried material.

 

Loss on Drying:

Weigh 4g of the sample in a previously weighed 100 ml beaker and heat in an oven at 105°C for 5 hours. Cool in desiccators and weigh. Repeat the procedure till constant weight is obtained. Calculate the percentage of loss in weight of the sample.

Water-soluble extractive:

Weigh accurately 4 g of the sample in a glass stoppered flask. Add 100 ml of distilled water, with shake occasionally for 6 hours and then allow standing for 18 hours. Filter rapidly taking care not to lose any solvent and pipette out 25ml of the filtrate in a pre-weighed 100 ml beaker and evaporate to dryness on a water bath. Keep it in an air oven at l05°C for 6 hours, cool in desiccators for 30 minutes and weigh. Repeat the experiment twice, and take the average value.

 

Alcohol-soluble extractive:

Weigh accurately 4 g of the sample in a glass stoppered flask. Add 100 ml of distilled alcohol shake occasionally for 6 hours and then allow standing for 18 hours. Filter rapidly taking care not to lose any solvent and pipette out 25ml of the filtrate in a pre-weighed 100 ml beaker and evaporate to dryness on a water bath. Keep it in an air oven at 105°C for 6 hours, cool in desiccators for 30 minutes and weigh. Calculate the percentage of Alcohol extractable matter of the sample. Repeat the experiment twice, and take the average value.

 

Table 1.   Analysis of Various Parameters

S. No.

Parameters

Sample 1

Sample 2

1

Description

Yellowish

Yellowish

2

Loss on drying at 105 0C

7.35 %

7.67 %

3

Total ash

4.61 %

4.78 %

4

Acid-insoluble ash

0.83 %

0.26 %

5

Water-soluble ash

0.635 %

1.24  %

6

Water-soluble extractive

26.15 %

24.82 %

7

Alcohol-soluble extractive

12.12 %

11.5%

 

Preliminary Phytochemical Studies:

Preliminary phytochemical results showed the presence or absence of certain phytochemicals in the roots of Withania somnifera Dunal .the result have been presented in Table 2.

 

Table 2 - Preliminary phytochemical test for different solvent extract of root of Withania somnifera Dunal

S. No.

Test

Sample 1

Sample 2

1.         

Alkaloid

+ ve

+ ve

2.         

coumarine

-ve

-ve

3.         

Flavone

+ ve

+ ve

4.         

Steroid

+ ve

+ ve

5.         

Tannin

+ ve

+ ve

6.         

Glycoside/Sugar

+ ve

+ ve

7.         

Terpenoid

+ ve

+ ve

8.         

Saponin

+ ve

+ ve

+ ve Present, - ve Not present.

 

Thin Layer Chromatography Methodology:

4g of the sample was soaked in 40 ml of rectified spirit (90%) with occasional shaking for 18 hrs, boiled for 10 minutes and filtered. The filtrate was evaporated and extracted with Chloroform. The soluble portion was filtered, concentrated and made up to 10 ml in standard flask. It was applied on (E. Merck) Aluminium plate pre-coated with Silica gel 60 F254 of 0.2 mm thickness using Linomat IV applicator. The plate was developed in


 

Table 3: Thin layer chromatographic

S. No

254 nm

366nm

After derivatization in visible light

Sample 1

Sample2

 

Sample 1

Sample 2

 

Sample 1

Sample 2

Colour

Rf.

Rf

Colour

Rf

Rf

colour

Rf

Rf

1

Darkgrey

0.30

0.31

-

-

-

Pale blue

0.09

0.08

2

Darkgrey

0.42

0.43

Pale blue

0.42

0.44

Blue

0.29

0.30

3

Pale grey

0.47

0.49

-

-

-

Grey

0.43

0.45

4

Pale grey

0.59

0.61

-

-

-

Grey

0.52

0.54

5

Pale grey

0.66

0.67

Pale blue

0.69

0.71

-

-

-

6

Pale grey

0.74

0.76

-

-

-

-

-

-

7

Dark grey

0.85

0.86

Blue

0.83

0.84

Grey

0.77

0.77

8

Dark grey

0.96

0.97

-

-

-

-

-

-

9

-

-

-

-

-

-

Dark grey

0.97

0.98

 

 

Toluene: Ethyl acetate: Acetic acid (6: 4: 0.5 v/v). After air drying the plate was visualized in UV 254 and 366 nm. The plate was then dipped in Vanillin -Sulphuric acid and heated in air oven at 105°C till the spots appeared are given in  Table 3 .

 

CONCLUSION:

The present study revealed different physicochemical parameters which can be used for authentication of the crude drug samples. Physicochemical as well as various aspects of the samples were studied. It has been concluded from this study, there are difference between parameters of both the samples shows the effect of environmental conditions. This study also helps to identify the right place for cultivation of medicinal plants and quality assessment.

 

ACKNOWLEDGEMENT:

The authors are grateful to college authorities and Director of CCRAS for constant support throughout this work.

 

REFERENCES:

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Received on 14.12.2011

Modified on 26.12.2011

Accepted on 01.01.2012        

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Research Journal of Pharmacognosy  and Phytochemistry. 4(1): Jan. - Feb. 2012, 14-17