A Preliminary Study on the Effect of Traditional Ayurvedic Purifying Methods of Semicarpus...

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Journal of Herbal Medicine and Toxicology 4 (2) 237-247 (2010)

ISSN : 0973-4643 Original Article

A PRELIMINARY STUDY ON THE EFFECT OF TRADITIONALAYURVEDIC PURIFYING METHODS OF SEMECARPUS

ANACARDIUM LINN. NUTS – A PHYSICO-CHEMICAL ANDPOWDER MICROSCOPIC STUDY

Venkateshwarlu G, Saraswathi P, Shantha T R, Shiddamallayya,

Kishore K R, Sridhar B N

Drug Standardization research Unit, National Ayurveda Dietetics Research Institute, (Central Council for

Research in Ayurveda & Siddha, Dept. of AYUSH, Ministry of Health & F. W, Govt. of India, New Delhi),

Ashoka Pillar, Jayanagar Bangalore-11

email: [email protected]

Abstract : Bhallataka - Semicarpus anacardium Linn seeds form one of the important

ingredients used in many medicinal formulations in Ayurveda for therapeutic and

Rasayana (rejuvenative) effects. They are reported to be potent antioxidants

corroborating their usage as rasayana (rejuvenator) in Ayurveda. Nonetheless, if used

without processing known as shodhana as per Ayurveda they have toxic effects. The

procedure of shodhana has been mentioned in ancient classics as a procedure that

removes external and internal doshas (impurities) and makes the material more potent,

effective, safe, assailable and homogeneous with out any adverse effects. But there

have been no studies to look into the changes in physiochemical and phytochemical

properties of the drug before and after shodhana. Hence in order to observe and

interpret the changes that might occur during shodhana and present it in the

contemporary light this study was planned incorporating Physicochemical analysis,

TLC and powder microscopy studies.

Keywords: Bhallataka, Semicarpus anacardium, Marking nut, Shodhana, Physico

chemical and Phytochemical studies, Powder microscopy

INTRODUCTION

Ayurveda is an ancient medical science for the

restoration of health and prevention of diseases

(patwardhan et al., 2009; Sharma et al., 2007).

Semecarpus anacardium (SA) nut finds in use in

variety of Ayurvedic medicinal preparations and is

effective against many diseases especially those that

are not amenable to allopathic medicines. It is called

as bhallataka in Ayurveda and commonly known as

marking nut in India. Chemical and phytochemical

analyses reveal the presence of biflavonoids, phenolic

compounds, bhilawanols, minerals, vitamins and amino

acids. A variety of nut extract preparations from this

source are effective in various diseases, viz. arthritis,

infections, tumours etc., (Premalatha., 2000;

Ramprasath et al., 2006), SA has been shown to

possess property of inducing apoptosis (Thatte et a.l,

2000). It has potent antioxidant properties (Verma et

al, 2009). Clinically compound preparations with SA

have been found to be safe including in pregnant

women (Murty, 1974). However, studies also have

reported cytotoxic effect of SA (Smith et al., 1995)

and toxic at dose dependent levels in animal studies

(Kesava Rao et al., 1979). Spermicidal effect of SA

in rats also has been reported (Sharma et.,al

2003).Toxic Phenols in SA causes dermatitis and

rashes (Goldsmith and Norman, 1943). A comparative

study of various purifying methods including the

traditional one using brick powder has been reported

Journal of Herbal Medicine & Toxicology

238

and it has been found that the oily compound of the

SA nut has pro oxidant properties (Tripathi et al.,

2008). But there has been no conclusive studies

looking into physicochemical and powder microscopy

analysis of SA nuts before and after traditional

Ayurvedic purification method. Processing of

medicinal ingredients used in Ayurveda to detoxify

them is known as shodana (Mohapatra et al., 2007).

shodhana not only reduces the toxic effects but also

enhances the therapeutic effect of the drugs at times

imparts additional qualities* (CS,su.1). Hence this

study was planned.

Brief description of the drug in Ayurveda

Pharmacodynamics and pharmacokinetics in

Ayurveda is explained in terms of attributes of an

ingredient used as medicine or food the description in

brief are as follows; Rasa [tastes viz., Madhura

(sweet), Amla (sour) Lavana (saline), Katu (pungent)

and Kashaya (astringent)]. Guna [properties (effect

it has on the body after ingestion and assimilation

including the nature of its interaction with digestive

juices), viz., laghu (light for digestion), guru (heavy

for digestion). ruksha (dryness) and so on]. Veerya

[potencies (release or conservation of energy during

digestion and metabolism) viz., ushna veerya (releases

energy during digestion and metabolism) and sheeta

veerya (conserves energy during digestion and

metabolism)]. Vipaka [post digestive effect on

metabolism viz., madhura (sweet), amla (sour), katu

(pungent)]. Based on these the probable action of a

drug or food can be predicted and understood in terms

of action on a dosha (vata pitta or kapha) known as

Doshaghnata. Karma is systemic action

vyadhiharatva or prabhava is specific action on a

particular disease. The details of bhallataka are given

in table1.

3. Known facts about bhallataka

The pericarp of the fruit contains a bitter and powerful

astringent principle. The black corrosive juice of the

pericarp has tarry oil consisting of 90% of an oxy-

acid named Anacardiac acid and 10% of a higher

non-volatile alcohol called Cardol (Naidu et al., 1925).

The crude extracts were found to be very toxic and

after purification, the toxicity was found to increase

as evident from LD50 values (Patwardhan et al.,

1986).

Ayurvedic method of administration after purification

has shown reduction in toxicity with maintained

efficacy (Patwardhan B.K.,et al 1988).

In order to verify the claims about the beneficial effect

of Shodhana, a preliminary study on the effect of

shodhana on the Bhallataka seeds have been

undertaken and also studied its physicochemical &

phytochemical changes of the drug before and after

shodhana with its comparative studies and detoxified

findings

4. Aim of the study

To identify change in phytochemical (TLC) and

botanical parameters (microscopy) in bhallataka seeds

with traditional purificatory procedures called

shodhana

MATERIALS AND METHODS

Materials used

Ballataka seeds -250gm.

Ishtika Churna (Brick powder)- Q.S. for gharshana

Jala (Water)- Q.S. for kshalana

Gunny bag for pottali,

5.2. Purification method

The Bhallataka seeds (250gms) were purchased

from the local market and the thalamus part of the

Bhallataka seeds was removed and made in to four

pieces of each seed. The cut pieces of Bhallataka

seeds were mixed with sufficient quantity of ishtika

churna (coarse brick powder) and made in to a pottali

by using a gunny bag. The mixtures was kept for 7

days then after these seeds were rubbed thoroughly.

The purified seeds were then washed in hot water

and dried in sun light and stored in moisture free

container (Ref. Rasmritam-Parishishtam-8,prishta-

147). The final quantity of sodhita bhallataka seeds

was 200gms.

5.3. Physico-Chemical Analysis

Both samples of seeds i.e. 1). Unpurified. 2). Purified

in Ishtika curna (coarse brick powder) were powdered

and used for the analysis. Physico-chemical &

preliminary phytochemical analysis of two samples

were carried out employing standard procedures and

239

Venkateshwarlu et al.

using GPR grade reagents [(WHO, 1996) British

Pharmacopoea, Indian Pharmacopoea ].

5.4. Thin layer chromatographic studies (TLC)

Comparative TLC was done using two solvent

systems a) 93:7- Toluene: Ethyl acetate solvent

Table 1 : Ayurvedic description of Bhallataka

S. No. Ayurvedic characteristic Properties

1 Rasa (taste) Katu (pungent)

2 Guna (quality) Laghu (light)

3 Veerya (potency) Ushna (hot)

4 Vipaka (post digestive effect) Madhura (sweet)

5 Doshagnata (effect on doshas) Vata kapha hara (pacifying)

6 Karma (action) Medhya (intellect promoting) agnivardhaka (digestive & metabolic

corrective)

7 Vyadhiharatva (indications) Kushta (skin disorders), gulma (space occupying lesions in the

abdomen), krimi(parasitic infestations) arshas (hemorrhoids)

8 Bhallataka Yoga (preparations

with SA)

Bhallataka rasayana, Bhallataka modaka, Amritha Bhallataka leha,

Sanjeevani vati, Bhallataka ghritha and Bhallataka avaleha

Table 2 : Showing values of physicochemical parameters before and after shodhana

S. No Parameters (%) Fresh seed (Before shodhana) Seed (After shodhana)

1 Weight 250 gms 200 gms

2 Ash content 1.88 13.98

3 Water soluble ash 0.69 6.68

4 Acid insoluble ash 0.50 3.76

5 Solubility in ethanol 10.50 9.34

6 Solubility in water 4.35 7.59

7 pH 6.2 6.9

8 Loss on drying at 105ºC 1.98 2.15

Table 3 : Rf values of the SA seeds before and after shodhana (for polyphenols)

S.No. Sample Extractive

s

Adsorben

t

Solvent

system

Viewing

reagent

Rf Values

1 Semicarpus

anacardium Seeds

(Before purification)

Methanol Silica gel

60 F254

Toluene: Ethyl

acetate (93:7)

Iodine vapour 0.10, 0.24,

0.47, 0.71,

0.76, 0.85

2 Semicarpus

anacardium Seeds

(After purification)

Methanol Silica gel

60 F254

Toluene: Ethyl

acetate (93:7)

Iodine vapour 0.18, 0.27,

0.30, 0.36,

0.43, 0.57,

0.71, 0.91


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Table 4 : Rf values of the SA seeds before and after shodhana (for oils and fats)

S.No. Sample Extractives Adsorbent Solvent

system

Viewing

reagent

Rf Values

1 Semicarpus

anacardium Seeds

(Before

purification)

Methanol Silica gel

60 F254

Benzene:

Ethyl acetate

(6:1)

Iodine

vapour

0.10, 0.18,

0.23, 0.32,

0.38, 0.58,

0.82, 0.90

2 Semicarpus

anacardium Seeds

(After purification)

Methanol Silica gel

60 F254

Benzene:

Ethyl acetate

(6:1)

Iodine

vapour

0.10, 0.18,

0.23, 0.32,

0.38, 0.58,

0.90

Table 5 : Organoleptic characteristics of bhallataka seeds

S. No. Parameter Characteristic (before shodhana) Characteristic (after shodhana)

1 color Brownish black dark brown

2 touch Greasy coarse Dry coarse

3 odour odourless odourless

4 consistency Very hard Moderately hard

Table 6 : Showing description of microscopy images

S. no Figure no. Description

1 1-4 Different fragments of tissues

2 5-6 Elongated small brown colored epicarp cells of epidermis.

3 7 Elongated stone cells with heavily lignified cell walls

4 8 Parenchyma cells and xylem vessels with helical thickenings

5 9 Parenchyma cells with reddish tannin contents

6 10 Thin walled rounded parenchyma cells.(Mesocarp cells)

7 11 Elongated ,thin walled parenchyma cells

8 12 Xylem vessels with helical thickenings

9 13 Xylem vessels with helical to reticulate thickenings

10 14 Rounded to elongated stone cells with heavily lignified cell walls.

11 15 Parenchyma cells with oilgloubles

12 16 Brown colored thick walled parenchyma cells

13 17 Parenchyma cells with reddish tannin content

14 18 Thin walled parenchyma cells

15 19 Fragments of endocarp cells, which are elongated, covered with thin cuticle and with

reddish content of tannin.

16 20 Reticulate to spiral xylem vessel.

17 21 Abundant small oilgloubles

18 22 Simple, oval to rounded starch grains

19 23 Compressed, elongated stone cells with lignified thickenings, lumen narrow

241

system for characterization of polyphenols and (b)

6:1- Benzene: Ethyl acetate solvent system for

characterization of oils and fats (Igon & Stahl, 1969).

The iodine vapour and long wave length (365 nm)

ultra violet images were evaluated.

5.4. Powder macro and microscopy

Sodhita bhallataka seeds were then powdered and

organoleptic characteristics were noted. Powder was

then treated with chloral hydrate solution and water,

observed under the microscope for diagnostic and other

features and photomicrographs were taken before and

after shodhana.

RESULTS

6.1. Physicochemical analysis of bhallataka seeds

before and after shodhana

The values of physico-chemical parameters before

and after shodhana are given in Table 2

6.2. Thin Layer Chromatography studies before and

after shodhana in 93:7 -Toluene: Ethyl acetate solvent

medium

The iodine vapour view and U-V view of T L

Chromatograms are shown in images 1, 1a, 2, 2a and

Rf values in Table 3.

6.3. Thin Layer Chromatography studies before and

after shodhana in 6:1 Benzene: Ethyl acetate solvent

medium

The iodine vapour view and U-V view of T L

Chromatograms are shown in images 3, 3a and Rf

values in Table 4.

6.4. Powder Macroscopy of Bhallataka seed powder

The macroscopy of bhallataka powder is shown in

image 4 and characteristics in table 5.

6.5. Powder microscopy

The diagnostic features of bhallataka seed powder

(Prasad et al, 1968) were identified in the sample and

are as follows.



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243



244

245

Presence of fragments of small, thin walled epidermal

cells.

Presence of fragments of elongated parenchyma cells

filled with reddish content of tannin of endocarp cells.

Presence of oilgloubles in parenchymatous cells.

Presence of compressed stone cells, lumen narrow,

with lignified walls .

Presence of fragments of xylem vessels with pitted ,

reticulate and helical thickenings. .

Presence of thinwalled parenchymatous cells with

reddish content of tannin.

The images and detailed descriptions of powder

microscopy of bhallataka seeds are shown in figures

1-23 and detailed descriptions are depicted in table 6

DISCUSSION

The objective of shodhana in Ayurveda is to make

the medicinal ingredient fit for a formulation and for

human consumption in anticipation of a desired disease

modifying effect. This concept hence is indicative of

quality control awareness of ancient scientists.

Nevertheless, the conventional interpretation of a

reductionist approach in detoxifying process, of late

can lead to misunderstanding of the intentions of

processes like shodhana with which they were

originally designed. This is because process like

shodhana might bring key changes that are beyond

just presence or absence of some chemical

constituent.

7.1. Physicochemical analyses

Physicochemical analyses showed the following; Loss

of 50 gms after shodhana might be attributed to the

washing and transfer of contents during physical

cleansing. Increase in water soluble ash and

corresponding increase in pH value turning it towards

neutral suggests that increase in alkaline salts

associated with shodhana making it less corrosive.

Increased water insoluble ash might be attributed to

increased amount of silica when the seeds were mixed

with brick powder. Insignificant change in ethanol

solubility might suggest that the process has not

brought in any change in the ethanol soluble fraction

of the polyphenols that have been attributed with

antioxidant activity. Whereas increased water

solubility was seen. This might be because of

increased fraction of water soluble tannins that most

of the times possess free radical scavenging activity.

Change in values of loss on drying was also

insignificant. The above results hence suggests that

shodhana procedure brings about increasing the

alkalinity of a more acidic form, apart from maintaining

the phenolic constituents of the seeds that may be

useful in bringing about free radical scavenging activity

usually noted in bhallataka seeds.

7.2. Thin Layer Chromatography

In the present study Methanol extracts were subjected

to TLC in Toluene Ethyl acetate (93:7) solvent system

which is specific among others, for poly-phenols. The

compounds corresponding to certain Rf values present

before shodhana (0.10, 0.24, 0.47, 0.76, 0.85) were

absent after shodhana. Similarly compounds

corresponding to certain Rf values (0.18, 0.27, 0.30,

0.36, 0.43, 0.57, 0.91) were present only after

shodhana. This suggests that shodhana must have

brought extensive rearrangement in the phenolic

constituents of bhallataka that might contribute to the

multidimensional attributed effects.

Further, TLC of Methanol extracts of the seeds before

and after shodhana in Benzene: Ethyl acetate (6:1)

solvent system specific to oils and fats revealed that

almost all the compounds corresponding to certain

Rf values (0.10, 0.18, 0.23, 0.32, 0.38, 0.58, 0.82, 0.90)

present before shodhana were present after shodhana

except that corresponding to 0.82. Besides, the

intensity of the chromatogram was reduced after

shodhana which means that shodhana has brought in

some change in the oily fraction of the constituents

of bhallataka by removing a certain compound that

makes it non toxic. Hence suggesting shodhana does

not necessarily mean complete removal of the tarry

oily portion, rather change in the chemical architecture

of this portion to make it non toxic and safe for human

consumption.

7.3. Powder Microscopy

Powder Microscopical studies after shodana revealed

that, all the microscopical characters are found to be

present in powder except calcium oxalate crystals

are not present in powder form. The powder study of

the fruit after purification has been described

highlighting its important characters, which will be



246

useful in identification of the fruit in powder form after

purification. The powder study revealed that, when it

is compared with that of the microscopical studies of

the fruit (S.Prasad etal 1968) it appears to be same in

having all the characters in powder form, even after

purification. It can be concluded that, all the characters

appears to be similar before and after purification of

the drug except in the absence of calcium oxalate

crystals in purified powder. Calcium oxalate crystals

are common biomineralization product in higher plants

(Jáuregui et al., 2003). They have been used as

microscopic markers in authentication of certain herbs

like ginseng (Yap et al., 2009). It also helps in

differentiating as well as characterization of herbal

products like in Fourier Transform Infra Red

Spectroscopy (FTIR) (Liang et al., 2009).

crystallochemical characterization of calcium oxalate

crystals can prove to be useful in identification of

different morphologies shown by these crystals grown

inside specialized cells in plants (Jáuregui et al., 2003).

Hence presence and absence of Calcium oxalate

crystals can be promising in its utility as a authentic

markers for bhallataka before and after shodhana.

Further, after absorption cannot be metabolized and

is excreted by the kidney into urine, where it binds to

calcium forming an insoluble salt that may precipitate

to form kidney stones (Massey et al., 2001). Hence

removal of calcium oxalate crystals after shodhana

is strong evidence of purification that makes bhallataka

safe for human consumption.

7.4. Summary and conclusion

Bhallataka seeds form one of the important ingredients

used in many medicinal formulations in Ayurveda.

They are reported to be potent antioxidants

corroborating their usage as rasayana (rejuvenator)

in Ayurveda. Nonetheless, if used without processing

known as shodhana as per Ayurveda they have toxic

effects. The procedure of shodhana has been

mentioned in ancient classics as a procedure that

removes external and internal doshas (impurities) and

makes the material more potent, effective, safe,

assailable and homogeneous with out any adverse

effects. But there have been no studies to look into

the changes in physicohemical and phytochemical

properties of the drug before and after shodhana.

Hence in order to observe and interpret the changes

that might occur during shodhana and present it in

the contemporary light this study was planned.

Physicochemical analysis, TLC and powder

microscopy studies on the drug before and after

shodhana revealed that the ancient detoxification

procedure brings about changes in pH, alters

phytochemical composition and removes calcium

oxalate crystals respectively making the drug fit for

human consumption.

ACKNOWLEDGEMENT

Authors are thankful to Director General, C.C.R.A.S,

New Delhi for providing facilities and encouragement.

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