International Journal of Ayurvedic Medicine

Many herbo-mineral remedies individually or in combination have been recommended in various medical treatises for the cure of different diseases. Gandhaka Rasayana is a compound preparation, being used by Ayurvedic physicians in clinical practice for thousands of years, in the treatment of skin diseases of bacterial and fungal origin. So the drug is selected to study its anti-fungal effect on Candida albicans. Candidiasis is the most common mycotic disorder of the skin affecting the immune-compromised patients and those on the anti-biotic therapy. Hence the anti-candidal activity was examined in the study. For the study Broth Dilution method was adopted to study the Minimum Inhibitory Concentration (MIC) and Minimum Contact Time (MCT) of the Gandhaka Rasayana, following NCCLS guidelines. Results: Gandhaka rasayana showed antifungal effect against Candida albicans with Minimum inhibitory concentration (MIC) of 5.00mg/ml of Brain Heart Infusion (BHI) Broth and Minimum Contact Time (MCT) above one hour

T, Prasanna Kumar

GS, Vijay Kumar

Devi, Yumnam Dhanesori

ISSN: 0976-5921 
International Journal of Ayurvedic Medicine, 2010, 1(2), 93-99 
93 
IN-VITRO ANTIFUNGAL ACTIVITY OF GANDHAKA RASAYANA 
Prasanna Kumar T
*
, Vijay Kumar GS
1
, Yumnam Dhanesori Devi
2
 
 
1
Professor and HOD, Department of Microbiology, JSS Medical College, Mysore, 
2
P.G. Scholar, Department of P.G. Studies in Rasashastra, JSS Ayurveda Medical College, Mysore, 
dr.dhanesori@gmail.com 
 
Abstract 
 
Many herbo-mineral remedies individually or in combination have been recommended in various 
medical treatises for the cure of different diseases. Gandhaka Rasayana is a compound 
preparation, being used by Ayurvedic physicians in clinical practice for thousands of years, in 
the treatment of skin diseases of bacterial and fungal origin. So the drug is selected to study its 
anti-fungal effect on Candida albicans. Candidiasis is the most common mycotic disorder of the 
skin affecting the immune-compromised patients and those on the anti-biotic therapy. Hence the 
anti-candidal activity was examined in the study. For the study Broth Dilution method was 
adopted to study the Minimum Inhibitory Concentration (MIC) and Minimum Contact Time 
(MCT) of the Gandhaka Rasayana, following NCCLS guidelines. Results: Gandhaka rasayana 
showed antifungal effect against Candida albicans with Minimum inhibitory concentration 
(MIC) of 5.00mg/ml of Brain Heart Infusion (BHI) Broth and Minimum Contact Time (MCT) 
above one hour.  
 
Key words: Candidiasis, Minimum inhibitory concentration, Minimum contact time, Gandhaka 
rasayana,  
 
Introduction: 
Fungal diseases of the skin can be divided 
into superficial mycoses and deep mycoses. 
Superficial mycoses are infections restricted 
to invasion of horny structures like the 
stratum corneum while the deep mycoses 
infect deeper layers. Deep mycoses are 
much more common in immuno-
compromised patients including these with 
AIDS, transplanted patients on 
corticosteroid or immuno-suppressive 
therapy. Candidiasis is one of the most 
common fungal infections affecting both the 
superficial as well as deeper layers of the 
skin and mucous membranes. Though 
candidiasis can be caused by various species 
of Candida (like C. albicans, C.stellatoidea 
tropicalis, C. krusei, C. viswanathii etc), 
Candida albicans is a causative species in 
about 80-90% of cases. Candida albicans is 
a normal inhabitant of gastro-intestinal tract, 
oral and vaginal cavities. Various 
manisfestation of candidiasis include 
mucocutaneous lesions (like oral thrush, 
vulvovaginitis, conjunctivitis etc), various 
skin and nail infections (commonly found in 
areas macerated by perspiration like axillae, 
groin, perineum, diapered area of infants, 
etc). Deeply invasive candidiasis is often 
preceded by increased superficial 
colonization of mouth, vagina, GIT, skin 
etc., mostly following broad spectrum 
antibiotic therapy. Factors which decrease 
the immune-competence of the person (e.g.: 
diabetes mellitus, HIV infections, 
corticosteroid therapy etc.) favour its 
growth. Candida can pass from colonized 
surface into deep tissue when integrity of 
skin or mucosa is violated. Local factors like 
* Corresponding Author:  
Assistant professor, Department of P.G. Studies in 
Rasashastra, JSS Ayurveda Medical College, 
Mysore,  
E-Mail: drprasannakumart@gmail.com,  
Tel: +91-9916482581 
Prasanna Kumar T et al., In-Vitro Anti-Fungal Activity of Gandhaka Rasayana 
94 
third degree burns, trauma and wounds, 
surgery, intravenous drug abuse may prove 
favourable for deep invasion.  
To find an affective Ayurvedic remedy for 
the treatment of the candidiasis Gandhaka 
rasayana is selected for the study. 
Gandhaka Rasayana is one of the important 
formulations used in the treatment all types 
of kusthas (skin diseases), nadivrana 
(sinuses) and kostha gata roga (Gastro-
intestinal disorders). Thus based on its use 
on certain skin diseases, since ancient times, 
the present study “In-Vitro Anti-Fungal 
Activity of Gandhaka Rasayana” was 
undertaken to prove its efficacy as an Anti-
fungal agent especially on Candida 
albicans. 
 
Aims and objectives: 
 To carry out Purification of the raw 
drug – Gandhaka (sulphur) and. 
 To prepare Gandhaka Rasayana 
 To find the Anti-fungal action of the 
prepared Gandhaka Rasayana and  
 To determine the Minimum 
inhibitory concentration (MIC) & 
Minimum Fungal dosage (MFD) of 
prepared sample against Candida 
albicans. 
Materials and methods: 
Raw materials were collected from genuine 
sources and subjected initially for the 
purification process. 
 
Gandhaka Shodhana (Purification of 
Sulphur ):(1) (Rasa ratna samuchaya 3/21-
23) 
Materials: Gandhaka – 500 gm (Figure 1), 
Godugdha – 2 lts, Mud pot, Sthali yantra, 
Cloth, lace or wire. 
Method: Bhudhara method (Figure 2) 
Procedure: A mud pot was taken and filled 
with 2 lts of Godugdha (Cow milk) and a 
cloth is tied to the mouth of the pot with a 
lace or wire. On the cloth 500 gm of 
Gandhaka was placed and is covered with 
Sthali yantra (Lid). Then the pot is placed in 
a pit made in the ground such that the pot 
along with the lid exactly fits into it. Then it 
is covered with sufficient number of 
Vanyopalas (Cow dung cakes). The cow 
dung cakes are then burnt till they become 
ash. The Gandhaka melts and drops into the 
milk leaving the impurities in the cloth. 
After the ashes get self cooled, the mud pot 
is taken from the pit and the Gandhaka is 
collected from the milk and washed in hot 
water.  Thus the pure Gandhaka (Figure 3), 
devoid of impurities is obtained. This is used 
for the preparation of the compound 
Gandhaka Rasayana. 
 
Observations: 
Gandhaka Before 
shodhana  
After 
shodhana(b) 
Weight 500g 496g 
Appearance  Yellow, 
crystalline 
Small 
globular and 
lustrous 
 
Preparation of Gandhaka Rasayana:(5)
 
Materials:  
 Suddha gandhaka (purified sulphur),  
 Godugdha (Cow Milk),  
 Chaturjata Kwatha (Decoction 
prepared by the combination of Tvak 
(Cinnamonum zeylanicum Lume.), 
Patraka (Cinnamum tamala Nees.), 
Ela (Elettaria cardamomum Maton.) 
and Nagakesara (Mesua ferrea)),  
 Guduci swarasa (Juice of the leaves 
of Tinospora Cordifolia),  
 Triphala kwatha (Decoction 
prepared by the combination of the 
fruits of Hareetaki (Terminalia 
chebula), Vibhitaki (Terminalia 
belerica) and Amalaki (Phyllanthus 
embelica)),  
 Sunthi kwatha (Decoction of dried 
ginger (Zingiber officinalis),   
ISSN: 0976-5921 
International Journal of Ayurvedic Medicine, 2010, 1(2), 93-99 
95 
 Bhringaraja swarasa (Juice of the 
whole plant of Eclipta alba) and  
 Ardraka swarasa (Juice of the wet 
rhizomes of Zingiber officinalis) 
 Kharala yantra 
Method: Bhavana (Trituration) 
Procedure: Suddha gandhaka (purified 
sulphur) was subjected to bhavana 
(trituration) first with Godugdha  (Cow 
milk) followed by the trituration with 
Chaturjata Kwatha, Guduci swarasa, 
Triphala kwatha, Sunthi kwatha, 
Bhringaraja swarasa and Ardraka swarasa, 
for 8 times each. Then the Gandhaka is 
collected and mixed with equal quantity of 
Sugar and preserved in an airtight bottle. 
This compound is called as Gandhaka 
Rasayana. 
 
Anti Fungal Study: 
After the drug is prepared, the antifungal 
study was conducted in MICROBIOLOGY 
DEPARTMENT OF JSS MEDICAL 
COLLEGE, MYSORE. The study is as 
follows. 
Procedure to know the Minimum 
Inhibitory Concentration (MIC): 
Preparatory procedures:  
A) To make Gandhaka Rasayana suitable 
for the study, it was made into fine 
powder and stock solution was prepared 
by adding 10 ml of BHI broth (Figure 5) 
to 100 mg of Gandhaka rasayana. The 
stock solution was then diluted to 
prepare three different samples of 
different concentration of the drug and 
were named as follows.  
Sample I – Containing Gandahaka 
Rasayana in a concentration of 10 
mg/ml of the BHI Broth.    
Sample II – Containing Gandahaka 
Rasayana in a concentration of 5 mg/ml 
of the BHI Broth.    
Sample III – Containing Gandahaka 
Rasayana in a concentration of 2.5 
mg/ml of the BHI Broth.    
B) Control groups: Two solutions named as 
positive control group and negative 
control group were taken as follows:  
Positive control group: In which fungal 
suspension was maintained to check the 
viability of organisms. 
Negative control group: Was only BHI broth 
for validation of the test was used.  
C) A suspension was prepared with standard 
strain of Candida albicans matching to 
0.5 Mc Farland turbidity standards.  
 
Method:  
To study the antifungal activity, In-
vitro study by Broth Dilution Method was 
adopted. In this five test tubes were taken 
and filled with 1 ml of the samples of 
Sample I, sample II, Sample III, Positive 
control group and Negative control group 
respectively. To the test tubes named as 
Sample I, Sample II and Sample III, 1 loop 
full of Candida albicans matching to 0.5 Mc 
Farland turbidity standards was added. Then 
all the five test tubes (sample group and 
control group) were incubated at 37
0
C for 24 
hrs.  
After 24 hrs of incubation, the test 
tubes were taken out on wet mount 
separately for each sample and were 
examined microscopically for viability of 
the Candida. Then one loop full of material 
was taken out from each test tube aseptically 
and inoculated on five different Sabouraud’s 
dextrose agar (SDA) plates separately for 
the sample groups and the control groups 
respectively. All the plates were then 
incubated at 37
0
C for 24 hours to promote 
the fungal growth. 
On the third day the plates were 
examined for presence or absence of fungal 
growth (colony formation). 
  
Procedure adopted to know the Minimum 
Contact Time (MCT):  
5.0mg of the prepared sample 
(Gandhaka Rasayana) was added to 5ml of 
Prasanna Kumar T et al., In-Vitro Anti-Fungal Activity of Gandhaka Rasayana 
96 
BHI broth containing Candida albicans. 
Subcultures were prepared from the solution 
at different times i.e., immediate, after half-
an-hour, one hour, one and half hour, two 
hours, two and half hours, three hours and 
four hours, on the SDA (Sabouraud’s 
dextrose agar) and were incubated at 37
o 
C 
for 24 hours for the viability of the Candida. 
After 24 hours, the SDA media was 
examined for the growth of the fungi. 
 
Observations and results:  
Minimum Inhibitory Concentration 
(MIC): (Table 1) 
After 24 hours of incubation of  SDA 
plates, positive control group and the 
Sample III were showing the growth of the 
Candida (Figure 6), indicating that the 
Candida were viable through out the 
experiment and the Concentration of 
Gandhaka Rasayana at 2.5 mg/ml of BHI 
Broth was not sufficient to inhibit the 
growth of the Candida. Thus it can be said 
that the Minimum Inhibitory Concentration 
(MIC) of Gandhaka Rasayana for Candida is 
5 mg/ml of BHI Broth. 
 
Table 1: Showing the Effect of different 
concentrations of Gandhaka rasayana on 
the Candida cultures 
SDA  plates Candida albicans 
Positive control group Growth present 
Negative control 
group 
Growth absent 
Sample I Growth absent 
Sample II Growth absent 
Sample III Growth present  
 
Minimum Contact Time (MCT): (Table 
2) 
It was observed after 24 hours of 
incubation that the growth of Candida was 
seen till half an hour of the adding of the 
Gandhaka Rasayana and was absent later. 
This indicates that the minimum contact 
time (MCT) of Gandhaka Rasayana to act 
anti-fungally should be above one hour. 
 
Table 2: Showing the Effect of Gandhaka 
rasayana on the Candida cultures 
inoculated at different times 
Time of subculture after 
inoculation 
Growth 
Immediately after adding 
sample 
Present 
Half an hour Present 
One hour Absent 
One & Half hour Absent 
Two hours Absent 
Two & Half hour Absent 
Three hours Absent 
Four hours Absent 
 
 
Discussion and conclusion:  
Ayurvedic System of medicine has its 
long history of therapeutic potential. Many 
drugs used by the Ayurvedic physicians 
have good therapeutic efficacy, but they lack 
scientific validity regarding the standard 
dosage patterns and the duration of the 
treatment to be adopted. Hence an 
experimental proof in this will help the 
mankind to use the medicine much 
efficaciously in the therapeutic practice.  
In the present study, Gandhaka 
Rasayana, a well known Ayurvedic 
compound was taken for the in-vitro anti-
fungal study especially on the Candida 
albicans. Gandhaka Rasayana has a wide 
range of therapeutic actions as mentioned in 
the Ayurvedic classics. It is used in diseases 
like Kushta, Kandu etc., which show a clear 
involvement of bacterial and fungal activity. 
Thus the drug is selected for the study to 
know its anti-fungal activity. 
Candidiasis is the most common 
mycotic disorder affecting the immune-
compromised patients like Diabetes, AIDS, 
ISSN: 0976-5921 
International Journal of Ayurvedic Medicine, 2010, 1(2), 93-99 
97 
post-transplat cases etc. So for this study, 
Candida albicans was selected from the 
fungal organisms as it is the most common 
cause of the candidiasis.  
Standard sample of the Gandhaka 
Rasayana is prepared and is used. Broth 
dilution method was adopted to know the 
Minimum inhibitory concentration (MIC) 
and the Minimum Contact Time (MCT).  
From the above study it was observed 
that the Minimum inhibitory Concentration 
of the of Gandhaka Rasayana on the 
Candida cultures is above 5 mg/ml of the 
BHI Broth and the minimum Inhibitory 
Concentration is above one hour. 
Thus it can be concluded that 
Gandhaka Rasayana has the anti-fungal 
activity on Candida albicans organisms 
when studied in-vitro. 
Further research has to be carried out 
to find out the effect of Gandhaka Rasayana 
on Candida albicans and to know its effects 
in-vivo and complications of using the drug.  
 
REFERENCES: 
1. Satpute D. Ashok, Rasa Ratna 
Samucchaya of Vagbhatta Acharya, 
First Edition, Delhi; Chaukamba 
Sanskrit Pratishthan; 2003. p. 316-PP3. 
2. Acharya Panashara Radhakrisna, 
Sharangdhara Samhita of Sharangdhara, 
Fourth Edition, Allahabad;  Shri 
Bhaidyanath Ayurved Bhavan Ltd.; 
1994. p.612 
3. Indradeva Tripathi, Rajanighantu of 
Pandit Narahari, Fourth Edition, 
Varanasi, Chowkhamba Krisnadas 
Academy; 2006. p.708 
4. Chunekar K.C, Pandey G.S, Sri 
Bhavaprakasa Nighantu of 
Bhavamishra, First Edition, Varanasi; 
Chaukhambha Bharati Academy; 2006. 
P.984 
5. Gulrajsharmamishra, Ayurveda 
prakasha of Madhav upadhyaya, 
Varanasi;2
nd
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Vishwabharati;1999 p.504 
6. Baveja,C.P, Text book of Microbiology, 
Second Edition, New Delhi; Arya 
Publication; 2005. p.626 
7. Anonymous, Pharmacopoeial Standards 
for Ayurvedic Formulations, Revised 
Edition, New Delhi; CCRAS; 1987.  
8. Ananthanarayanan R, C. K. Textbook of 
Microbiology, 4
th
 Edition, Madras; 
Jayaram Paniker, Orient Longman 
Limited; 1990.  
9. Ghosh B. N, Pharmacology, Materia 
Medica and Therapeutics, 18
th
 Edition, 
Hilton & Co;  
10. Nadakrani K.M, Nadakarni A.K, Indian 
Materia Medica Volume II, 2
nd
 Edition, 
Bombay; Popular Prakashana; 1992. 
P.47 
11. Satoskar R.S. and Bhandarkar S.D., 
Pharmacology and 
Pharmacotherapeutics, 12
th
 Edition, 
Bombay; Popular Prakashan; 1991.  
12. Behl P.N, Practice of Dermatology, 
New Delhi; CDS Publishers; 1988.  
13. Harrison; Principles of Internal 
Medicine, 17
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 Edition, USA; McGraw-
Hill              Publication; 2009.p.2607 
14. Clayton Thomas-Taber's Cyclopedic 
Medical Dictionary, Edi. XVIIIth, 1998,  
15. Jaypee  Brothers Medical Publishers, 
New Delhi. 
16. Norman Evers G. D, Elsdon. C., The 
analysis of drugs and chemicals, 
Company Ltd; 1929.  
17. Davidson, Davidson’Principles and 
Practice of Medicine, 18
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 Edition,  
Great Britain; ELBS; 1999.p.1381 
 
 
Prasanna Kumar T et al., In-Vitro Anti-Fungal Activity of Gandhaka Rasayana 
98 
 
 
  
Figure 1: Impure Gandhaka (Sulphur) Figure 2: Bhudara puta  
 
 
Figure 3: Purified Gandhaka Figure 4: Gandhaka Rasayana 
 
 
 
Figure 5: SAMPLE WITH BHI BROTH 
Figure 6: CANDIDA ALBICANS 
 
ISSN: 0976-5921 
International Journal of Ayurvedic Medicine, 2010, 1(2), 93-99 
99 
 
Figure 7: Subculture of Candida on SDA at different intervals of time 
 
A.  Immediately after adding sample B.  After half an hour 
C.  After One hour D.  After One & Half hour 
E.  After Two hours F.  After Two & Half hour 
G.  After Three hours H.  After Four hours 
 
 
 
 
***** 
 


English

IN-VITRO ANTIFUNGAL ACTIVITY OF GANDHAKA RASAYANA

Picrorrhiza kurroa Royle ex Benth is commonly known as Kutki belonging to family Scrophulariaceae. It is useful as a laxative, liver-stimulant, improving lactation, appetite stimulant, fabrifuge and as beneficial in bronchial asthma. This plant is native to the Himalayan region and is part of Ayurvedic medicine as a treatment for various diseases. The leaf, bark and the underground parts of the plant, mainly rhizomes are widely used in the traditional Indian (Ayurvedic) systems of medicine since ancient times. Rhizomes of Picrorrhiza kurroa Royle ex Benth plant material was subjected to microscopic characterization and physico-chemical studies. TLC has also been studied to fix the quality standards of this drug. The experiment has revealed a set of diagnostic characters essential for its standardization. Photomicrography and TLC profile were employed to fix standards. In this article attempt has been made to standardize the Picrorrhiza kurroa Royle ex Benth obtained from reliable source for its physico-chemical parameters like loss on drying, total ash value, acid insoluble ash, water soluble extract, alcohol soluble extract, pH etc.&nbsp

K, Meena A

Krishna, C M

M, Rao M

K N, Sunil Kumar

Preet, Komal

M, Padhi M

Babu, Ramesh

International Journal of Ayurvedic Medicine, 2010, 1(1), 41-47 
 
 
1* Corresponding Author, National Institute of Ayurvedic Pharmaceutical Research, Patiala-147001, Punjab.  
Tele Fax. No. +91-175-2212393; +91-9465159789 (M), E-mail: ajaysheera@yahoo.co.in.  
1. National Institute of Ayurvedic Pharmaceutical Research, Patiala-147001, Punjab, 2. Central Council for 
Research in Ayurveda and Siddha, Janakpuri, Delhi, 3. National Institute of Indian Medical Heritage, 
Hyderabad, India, 4. Captain Srinivasa Murti Research Institute for Ayurveda and Siddha Drug Development, 
Chennai. 
41 
Evaluation of Phamacognostic and Physicochemical  
parameters of Picrorrhiza kurroa Royle ex Benth 
 
A K Meena
1
*, M M Rao
1
, C M Krishna
3
, Komalpreet
1
,  
K N Sunil Kumar
4
, M M Padhi
2
, Ramesh Babu
2 
 
 
Abstract 
Picrorrhiza kurroa Royle ex Benth is commonly known as Kutki belonging to family 
Scrophulariaceae. It is useful as a laxative, liver-stimulant, improving lactation, appetite 
stimulant, fabrifuge and as beneficial in bronchial asthma. This plant is native to the 
Himalayan region and is part of Ayurvedic medicine as a treatment for various diseases. The 
leaf, bark and the underground parts of the plant, mainly rhizomes are widely used in the 
traditional Indian (Ayurvedic) systems of medicine since ancient times. Rhizomes of 
Picrorrhiza kurroa Royle ex Benth plant material was subjected to microscopic characterization 
and physico-chemical studies. TLC has also been studied to fix the quality standards of this drug. The 
experiment has revealed a set of diagnostic characters essential for its standardization. 
Photomicrography and TLC profile were employed to fix standards. In this article attempt has 
been made to standardize the Picrorrhiza kurroa Royle ex Benth obtained from reliable 
source for its physico-chemical parameters like loss on drying, total ash value, acid insoluble 
ash, water soluble extract, alcohol soluble extract, pH etc. 
 
Keywords: Picrorrhiza kurroa Royle ex Benth, Rhizomes, Ayurvedic 
 
Introduction 
Picrorrhiza kurroa Royle.ex Benth 
is commonly known as Kutki belonging to 
family Scrophulariaceae. It is useful as a 
laxative, liver-stimulant, sialogogue, 
appetite stimulant, fabrifuge and as 
beneficial in bronchial asthma. The plant 
and its formulations are widely used in 
therapy of epidemic jaundice. Clinical 
studies have been carried out with the root 
powder of the plant in patients with viral 
hepatitis with significant improvement in 
symptoms like anorexia, nausea and 
vomiting. It is bitter, carminative, 
digestant and has a cooling effect and is 
used as a cardiotonic, antipyretic and 
anthelmintic. It is also used in diabetes, 
jaundice, liver, spleen disorders and skin 
diseases. The Kutki plant is native to the 
Himalayan region. It is used in Ayurvedic 
system of medicine for treating primarily 
gastrointestinal problems such as 
indigestion and constipation. Usually the 
root or rhizome of the plant is used 
medicinally. Picrorrhiza kurroa Royle ex 
Benth is a small perennial herb growing in 
the hilly areas of the North-Western 
Himalayan region in India and Nepal. 
Although it shows anti-oxidant, anti-
inflammatory and immunomodulatory 
activities, it is most valued for its 
hepatoprotective effect. The bitter 
rhizomes of Picrorrhiza kurroa Royle ex 
Benth have been used for thousands of 
years in India to treat people with 
indigestion (1) and constipation due to 
insufficient digestive secretion (2). 
Picrorrhiza kurroa Royle ex Benth is 
considered as a trophorestorative herb for 
the liver as well as a potent 
immunostimulant (3). Its constituent, 
picroliv is also reported to possess 
choleretic effect (4), and prevent hepatic 
AK Meena et al., Evaluation of Phamacognostic and Physicochemical parameters of Picrorrhiza kurroa Royle ex Benth 
42 
injury caused by ethanol (5) and 
chemicals.  
Picrorrhiza kurroa Royle ex Benth 
is a small, hairy perennial herb with small 
white or pale bluish flowers in cylindrical 
spikes and woody rhizomes. The dried 
rhizomes of the plant constitute the drug. 
Picrorrhiza kurroa Royle ex Benth is 
highly beneficial in treating liver cirrhosis 
among adults. Its root is given in 
powdered form. It stimulates the liver to 
produce more bile. Picrorrhiza kurroa 
Royle ex Benth is also used with good 
results in the treatment of ascites, a disease 
characterized by the accumulation of fluid 
in the peritoneal cavity of the abdomen.  
 Picrorrhiza kurroa Royle ex Benth 
is one of the major drugs useful in 
jaundice. It is beneficial in the treatment of 
dyspepsia. It strengthens the stomach and 
promotes its action, while improving 
appetite and stimulating the secretion of 
gastric juices. It cures periodic attacks like 
hysteria, epilepsy and convulsions and 
promotes the secretion of bile. It has 
antibiotic properties and is a mild 
purgative. Its main chemical constituents 
are glycosides picroside I, II and III, 
picrorhizin, kutkoside, kurrin, kuthinol, 
kutkiol, kutkisterol, kutkoside, androsin, 
apocynin, drosin and cucurbitacin. 
Apocynin is a catechol that has been 
shown to inhibit neutrophil oxidative burst 
in addition to being a powerful anti-
inflammatory agent while the 
curcubitacins have been shown to be 
highly cytotoxic and possess antitumor 
effects. 
Here, we attempted to study the 
physico-chemical standardization 
parameters as well as analyze the crude 
drug for its authenticity by microscopic 
characterization.  
 
MATERIALS AND METHODS 
Plant material 
The sample of Picrorrhiza kurroa 
Royle ex Benth was collected from local 
market of Patiala, Punjab. Specimen was 
identified and authenticated at the National 
Institute of Ayurvedic Pharmaceutical 
Research (NIAPR), Patiala (Punjab). 
Samples were studied for physiochemical 
evaluation. 
 
Methods for Physicochemical 
Parameter 
Moisture content 
4 g of the sample was taken and 
heated in an oven at 105°C for 5 hour in a 
previously weighed 100 ml beaker. It was 
cooled in desiccators and weighed. The 
procedure was repeated till constant 
weight is obtained. The percentage of loss 
in weight of the sample was calculated.  
Deterioration time of the plant 
material depends upon the amount of 
water present in plant material. If the water 
content is high, the plant can be easily 
deteriorated due to fungal attack. The loss 
on drying at 105°C of Picrorrhiza kurroa 
Royle ex Benth rhizome was found to be 
7.83 %. 
 
Determination of Total ash value 
2 g of the sample was taken 
accurately in a previously ignited and 
tarred Silica dish. The material was spread 
evenly and ignited in a muffle furnace by 
gradually increasing the temperature to 
600
o
C until it is white, indicating the 
absence of carbon. The crucible was 
cooled in desiccators and allowed to stand 
for 30 minutes and weighed.  
Total ash value of plant material 
indicated the amount of minerals and 
earthy materials attached to the plant 
material. Analytical results showed total 
ash value of Picrorrhiza kurroa Royle ex 
Benth rhizome was 5.92 %. 
 
Determination of Acid insoluble ash 
value 
To the dish containing the total ash, 
25 ml of 20 % Hydrochloric acid was 
added covered with a watch glass and 
boiled gently for 5 minutes. The watch 
was rinsed with a hot water and added to 
the crucible. The residue was washed with 
the hot water till the washings were neutral 
International Journal of Ayurvedic Medicine, 2010, 1(1), 41-46 
43 
to the litmus. The insoluble material was 
collected and again placed in a same 
crucible and again ignited for 6 hr. to 
constant weight. The residue was cooled a 
desiccators for 30 minutes and weighed.  
Percentage of acid insoluble as was 
calculated. The amount of acid-insoluble 
siliceous matter present in the Picrorrhiza 
kurroa Royle ex Benth rhizome was 
2.14%. 
 
Determination of Water soluble 
extractive value 
4 g of the sample was taken in a 
glass stoppered flask.100 ml of distilled 
water was added. The flasks were shaken 
occasionally for 6 hours and then allowed 
to stand for 18 hours. The extract was 
filtered and 25 ml of the filtrate was 
pipette out in a pre-weighed 100 ml beaker 
and evaporated to dryness on a water 
bath.It was kept in a hot air oven for 5 hr 
at l05°C, cooled in a desiccators for 30 
minutes and weighed. The procedure was 
repeated till constant weight. 
The water-soluble extractive value 
indicated the presence of sugar, acids and 
inorganic compounds. The water soluble 
extractive value in the Picrorrhiza kurroa 
Royle ex Benth rhizome sample was found 
to be 33.10%. 
 
Determination of Alcohol soluble 
extractive Value 
Same procedure as for the water 
soluble extractive value was followed. 
Instead of water, rectified spirit was taken 
as a solvent. 
The alcohol soluble extractive 
values indicated the presence of polar 
constituents like phenols, alkaloids, 
steroids, glycosides, flavonoids and 
secondary metabolites present in the plant 
sample. The alcohol soluble extractive 
value was found to be 25.40% in the 
Picrorrhiza kurroa Royle ex Benth 
rhizome. 
 
 
 
Determination of pH Value 
10% aqueous solution of sample 
was prepared and used for determining the 
pH value by pH meter. The pH value of 
Picrorrhiza kurroa Royle ex Benth 
rhizome was found to be 4.84 
 
RESULT AND DISCUSSION 
Microscopy 
Rhizome: 
Transverse section of the rhizomes 
showed outermost thin brown colored 
cork, lacunose black colored bark, a circle 
of vasculature formed by 5 to 7 ovate 
vascular bundles. 
 TS of the rhizome (Figure 1) 
showed 8 to 12 layers of rectangular cork 
cells without any intercellular spaces. 
Cells of the cortex were found thick and 
loosely arranged in definite radial rows. It 
showed ovate vascular bundles in between 
the cortex and pith. In the centre there was 
parenchymatous pith with pitted walls. 
Simple starch grains were found in the 
parenchyma of the cortex and the pith. 
Detailed TS of the root was similar 
to that of rhizome in histology but it 
showed a central small circular vascular 
bundle formed by xylem in the centre 
surrounded by phloem.  
 
Powder: 
Microscopy of the powder (Figure 
2) showed plenty of thick-walled 
parenchymatous cells with starch grains. 
Cortex cells were found circular to oval. 
Cork cells in transversely cut mode and in 
surface view were found to contain 
fragments of pitted vessels and tracheids 
and groups of cells with orange colored 
content. 
 
Physico-chemical parameters: 
Different physicochemical 
parameters like Total ash, acid insoluble 
ash, water soluble extractive value, ethanol 
soluble extractive value, loss on drying at 
105ºC and pH etc. were studied. (Table 1)  
 
AK Meena et al., Evaluation of Phamacognostic and Physicochemical parameters of Picrorrhiza kurroa Royle ex Benth 
44 
 
 
Figure 1. TS of rhizome of 
Picrorrhiza kurroa Royle ex Benth 
Figure 2. Powder microscopy of Picrorrhiza kurroa 
Royle ex Benth 
(a. starch grains; b. cork in surface view; c. 
fragments of pitted vessels; d. transversely cut 
cork; e. cortical parenchyma; f. pigment cells.) 
 
Table 1. Physicochemical parameters of Picrorrhiza kurroa Royle ex Benth 
S. No. Name of Parameters Results 
1.  pH (10% aqueous solution (v/w) 4.84 
2.  Loss on drying at 105oC (% w/w) 8.96 
3.  Total Ash (% w/w) 5.92 
4.  Acid-insoluble ash  (% w/w) 2.14 
5.  Water-soluble extractive  (% w/w) 33.10 
6.  Alcohol-soluble extractive (% w/w) 25.40 
 
Thin Layer Chromatography (Figure 3) 
The thin layer chromatography of 
the Kutki was performed and observed 
under 366 and 254 nm. The plates were 
also exposed to the iodine vapours and to 
the vanillin-sulphuric acid solution for 
Derivatisation (Fig: 3). Under 366 nm the 
plate showed no spots indicating that the 
Kutki contains no components absorbing at 
this wavelength. At 254 nm, the samples 
showed 2-3 spots towards the baseline 
indicating the highly polar nature of the 
compounds. After exposing to the iodine 
vapours, the plate showed few spots in 
between the base line and solvent front 
indicating the presence of compounds 
which can be fixed by the iodine. After 
dipping the plate in a vanillin-sulphuric 
acid solution, the plate showed different 
colored compounds indicating the 
presence of compounds with strong 
chromophoric groups and multiple bonds.  
The Rf of all the spots was 
calculated and mentioned in the table 2. 
 
Table 2. TLC profile of Picrorrhiza kurroa Royle ex Benth 
S.No 254 nm After Derivatisation 
with Iodine  in visible 
light 
After Derivatisation with 
Vanillin sulphuric acid in 
visible light 
Colour Rf. Colour Rf. Colour Rf. 
1.  Purple 0.11 Yellow 0.09 Purple 0.11 
International Journal of Ayurvedic Medicine, 2010, 1(1), 41-46 
45 
2.  Purple 0.16 - - Purple 0.16 
3.  Purple 0.36 Yellow 0.36 Pink 0.36 
4.  - - - - Pink 0.41 
5.  - - - - Violet 0.51 
6.  - - - - Light Pink 0.55 
7.  Purple 0.72   Violet 0.72 
8.  - - Yellow 0.82 Pink 0.82 
 
Figure 3: TLC profile of Picrorrhiza kurroa Royle ex Benth rhizome  
(Toluene: Ethyl acetate 7.5:2.5) 
λ 254 nm 
 
After Derivatisation with 
Iodine  in visible light 
After Derivatisation with 
Vanillin Sulphuric acid in 
visible light 
   
 
            
Conclusion 
Morphology as well as various 
phamacognostic aspects of the rhizome 
sample was studied and described along 
with physio-chemical parameters and 
TLC. These parameters will be useful in 
authentification and identifying the 
adulterants and quality control of raw 
drugs. Whole plant of Picrorrhiza kurroa 
Royle ex Benth exhibited a set of 
diagnostic characters, which will help to 
identify the drug in dried condition. It has 
been concluded from this study that 
estimation of these parameters is highly 
essential for raw drugs or plant parts used 
for the preparation of compound 
formulation. The periodic assessment is 
essential for quality assurance and safer 
use of herbal drugs. 
 
Acknowledgement 
Authors acknowledge the 
secretarial assistance rendered by Ms. 
Rekha in the preparation of this paper.  
 
 
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AK Meena et al., Evaluation of Phamacognostic and Physicochemical parameters of Picrorrhiza kurroa Royle ex Benth 
46 
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