Global Journal of Pharmacology 7 (3): 288-293, 2013

ISSN 1992-0075
© IDOSI Publications, 2013
DOI: 10.5829/idosi.gjp.2013.7.3.75167

Screening for Antimicrobial Activity of the Stem Bark of Bauhinia purpurea Linn
1
Yogita Sardessai, 2Roocha R. Desai,
2
Arun B. Joshi and 2Maya P. Bhobe

1
Department of Microbiology, Goa College of Pharmacy, Panaji-Goa, 400601, India
2
Department of Pharmacognosy, Goa College of Pharmacy, Panaji-Goa, 400601, India

Abstract: The aim of the present study was, to carry out the phytochemical and antimicrobial screening of the
n-hexane extract of the stem bark of Bauhinia purpurea. The preliminary phytochemical studies revealed the
presence of fatty acids, triterpenoids, steroids, alkaloids and phytol esters. The antimicrobial screening of the
n-hexane extract of the stem bark of Bauhinia purpurea by Well Diffusion and Tube Dilution Method was
carried out against 6 bacterial strains; Bacillus subtilis, Staphylococcus aureus, Escherichia coli,
Pseudomonas aeruginosa, Salmonella typhimurium, Klebsiella species and 2 fungal strains; Aspergillus niger
and Claviceps purpurea. The present evaluation revealed that the n-hexane extract exhibited activity towards
all the tested microbial strains showing a broad spectrum of activity against gram positive as well as gram
negative microorganisms.

Key words: Bauhinia purpurea Leguminosae Tube Dilution Well Diffusion

INTRODUCTION resistant strains. Despite a great step towards

development of new antibiotic drug therapies there has
Natural products obtained from the plant resources been a decline in advancement of new drugs.
have been the major supplements to combat many serious The rapid emergence of multiple drug resistance
diseases in the developing countries [1]. With the strains of pathogens to current antimicrobial agents has
advancement of modern medicinal systems, herbal generated an urgent intensive for new antibiotics from
medicines have always maintained their popularity in medicinal plants. Many medicinal plants have been
treatment of diseases due to their easy availability and screened extensively for their antimicrobial potential
safe effectiveness. The capacity of herbs to produce worldwide. Free radicals which have one or more unpaired
innumerable bioactive secondary metabolites has electrons (superoxide, hydroxyl, peroxyl) are produced in
increased their usage in healing systems. normal or pathological cell metabolism and the
Natural products, as a basis of new drugs, have a compounds that can scavenge free radicals have great
great promise and it is gratifying to note that the World potential in ameliorating the diseases and pathological
Health Organization is showing abiding interest in plant cells [3,4].
derived medicines, described in the folk medicine of The genus Bauhinia L is often called as ‘Orchid
different countries. According to the WHO (2001), 80% of Tree’ of ornamental value [5]. Bauhinia purpurea (Linn.)
the world population uses natural remedies and traditional is a medium sized deciduous flowerings tree, bark ashy to
medicines [2]. dark brown belonging to the family Leguminosae and
During the last few decades, the number of upcoming subfamily Caesalpinioidae [6, 7]. It is an erect tree with a
infectious diseases is increasing in the developing round, symmetrical, moderate dense crown of 25 feet
countries [2]. Plant based products may be a new boon to growing to a height of 20-35 feet tall [8]. The plant
the development of antibiotics effective against antibiotic commonly known as Purple Orchid Tree or Butterfly Tree

Corresponding Author: Roocha Ramesh Desai, (Pharmacognosy), Department of Pharmacognosy and Phytochemistry,
Goa College of Pharmacy, Panaji-Goa, 403001, India, Tel: +09823888599 (M),
Fax: +2226883.
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is a native from the foot of the West Himalayas, Khasia

Mountains and also found distributed in other parts of
India and China [9].
The plant is well known for its traditional uses to
treat roundworm infections, conjunctivitis, anthrax,
ulcerations, dysentery, blood-poisoning, leprosy, lung
and skin diseases. The use of different parts of the plant
like roots as carminative and flowers as laxative are folk
claimed. The bark is commonly used for treatment of
diarrhoea and other gastrointestinal complaints [10,11]. Leaf and Stem Bark of Bauhinia purpurea linn.

Salmonella typhimurium ATCC 23564, Kleibsiella

MATERIAL AND METHODS species and 3 fungal strains like Aspergillus niger ATCC
10864, Claviceps purpurea NCIM No. 1046 and Candida
Authentication and Collection of the Plant Material: albicans.
The stem bark of B. purpurea was collected from Altinho,
Panaji - Goa during November, 2011. It was identified and Determination of Antimicrobial Activity: The n-hexane
authenticated by Prof. G. I. Hukkeri, Dept. of Botany, extract of stem bark of B. purpurea was subjected to
Dhempe College of Arts and Science, Miramar - Goa, antibacterial as well as antifungal screening by Well
India. Diffusion Method (Cup Plate Method) [13-15] and Broth
Dilution method (Tube Dilution Method). Mueller Hinton
Preparation of n-hexane Extract: The stem bark of Agar/Broth and Sabouraud’s Dextrose Agar/Broth were
B. purpurea was collected from Altinho, Panaji - Goa used as the seed medium for the antibacterial and
during the month of November, 2011. It was dried under antifungal screening respectively. The Minimum
shade. The dried stem bark was powdered (500g) and Inhibitory Concentration was performed by two-fold
exhaustively extracted by maceration with n-hexane for dilution of the test extract in the respective medium under
three days. After three days, the n-hexane layer was sterile conditions [14, 16]. The stock inoculums of the
decanted off. The process was repeated thrice. The bacteria and fungi were prepared in sterile normal saline
solvent from the total extract was distilled off and the (0.9%) previously autoclaved at 1200C with 15lbs bar
concentrate was evaporated to a syrupy consistency and pressure. The fungal and bacterial cultures were
then evaporated to dryness (5g). maintained on potato dextrose agar and nutrient agar
slants respectively and refrigerated at 50C. The inoculum
Preliminary Phytochemical Analysis: Qualitative was verified by streaking on specific medium for colony
analysis was carried out of the crude n-hexane extract identification and purification. Appropriate controls were
which revealed the presence of fatty acids, triterpenoids, maintained.
steroids, alkaloids and phytol esters. The plates were observed visually and the diameter
of zones was measured using an mm scale. The activity
Isolation of Phytoconstituents from N-hexane Soluble was indicated by the presence of clear zones around the
Fraction: Chromatographic elution’s led to the well size. The bioassay was repeated thrice and the mean
isolation of 9 plant constituents namely Myristic was recorded to check the effectiveness of the procedure.
Acid, Octadecanoic Acid, 9, 12- Octadecadienoic The MIC was determined by turbidometric method by
Acid, isopropyl-24-methyl-pentacosanoate, Stigmasterol, measuring the Optical Density using Elico colorimeter
-Amyrin, -Sitosterol, Lupeol and ethyl 9, 12- (Filter No. 60). The MIC results were further reinforced by
Hexadecadienoate [12]. determining viable counts using pour plate method.

Microbial Strains: The microbial strains used for the RESULTS

antimicrobial screening techniques were acquired from
National Chemical Laboratories, Pune. The bacterial The phytochemical screening led to the isolation of
strains used for the study are Bacillus subtilis ATCC many bioactive phytoconstituents. The n-hexane extract
6633, Staphylococcus aureus ATCC 6538, Escherichia was found to be significantly effective towards bacterial
coli ATCC 35218, Pseudomonas aeruginosa ATCC 19429, as well as fungal strains showing a broad spectrum of

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Table 1: Effect of n-hexane extract of Bauhinia purpurea against various bacterial strains by Well Diffusion Method.
Diameter of Zone of Inhibition in mm
----------------------------------------------------------------------------------------------------------------------------------------------------
Bacterial Strains
-----------------------------------------------------------------------------------------------------------------------------------------------------
Sample SA BS PA ST EC KS
Extract 17 16 39 21 18 12
Standard (Streptomycin) 40 39 50 28 15 34
Concentration of the stock solution was 25mg/ml.
SA (Staphylococcus aureus ATCC 6538), BS (Bacillus subtilis ATCC 6633), PA (Pseudomonas aeruginosa ATCC 19429), ST (Salmonella typhimurium
ATCC 23564), EC (Escherichia coli ATCC 35218), KP (Klebsiella species) and 2 fungal strains; AN (Aspergillus niger ATCC 10864), CP (Claviceps
purpurea NCIM No. 1046).

Table 2: Effect of concentration of n-hexane extract on Bacterial cultures by Tube Dilution Method
Concentration (µg/ml) SA BS PA ST EC KS
+ve 0.76 0.56 0.63 0.71 0.70 0.53
31.25 0.56 0.42 0.66 0.74 0.71 0.59
62.5 0.55 0.38 0.53 0.54 0.65 0.47
125 0.52 0.26 0.44 0.34 0.62 0.41
250 0.23 0.22 0.39 0.30 0.51 0.30
500 0.22 0.19 0.35 0.25 0.41 0.25
1000 - - - - - -
Concentration of the stock solution was 2mg/ml.
Extract control O.D.-0.10 (turbidity of the extract)
-Indicates no growth

Table 3: Effect of concentration of n-hexane extract on fungal cultures by Broth Dilution Method
AN CP
Sample -------------------------------------------------------------------------- -----------------------------------------------------------------------------
Concentration (µg/ml) 1000 500 250 125 62.5 31.25 1000 500 250 125 62.5 31.25
Extract - - - + + + - - - - - +
Concentration of the stock solution was 2mg/ml.
+ positive indicating presence of growth -negative indicating absence of growth

activity. The extract shows significant antibacterial diffusion and MIC methods (Tables 1 & 2). The large zone
activity towards Bacillus subtilis, Staphylococcus of inhibition obtained with P. aeruginosa is particularly
aureus, Escherichia coli, Pseudomonas aeruginosa, significant as the incidence of drug resistance in this
Salmonella typhimurium, Klebsiella species and culture is well known and many of its strains are
antifungal activity towards the fungal strains; implicated in nasoconial infections. Off late there are many
Aspergillus niger and Claviceps purpurea as cited below bioactive results regenerated mainly focussing on
(Tables 1-3). antimicrobial activity of aqueous and methanolic
extracts of B. purpurea [2, 20- 22]. This paper reports
DISCUSSION the antimicrobial activity of n-hexane extract of the stem
bark of B. purpurea Linn and further accelerates our
Bauhinia traditionally is well known to have study on bioactive compounds.
multiple uses in medicine. Since it is used in treatment of It was seen from the present study that there was
several infectious diseases such as diarrhoea, fever, negligible growth seen at 500µg/ml for most bacterial
dysentery, skin diseases, cancer etc [17]; it was decided cultures and no growth at 1000µg/ml. (Table 2). This
to analyse the extract against various bacterial and fungal further indicates that the MIC value lies between 500µg/ml
strains. S. aureus, P. aeruginoasa and E.coli, well known to 1000µg/ml. From the above results it is evident that as
to be a causative agent for boils, skin infections, the concentration of the n-hexane extract increases the
abscesses, dysentery and diarrhoea [18, 19] appears to be viable count decreases indicating the bioactivity of the
strongly inhibited by n-hexane extract by both well plant extract (Fig 1).

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Fig. 5:
Fig. 1:

Fig. 6:
Fig. 2:
In our study the chromatographic elution led to
the isolation of Myristic acid, Octadecanoic acid
(Stearic acid), 9,12-Octadecadienoic acid, Stigmasterol,
-Amyrin, -Sitosterol and Lupeol. It is well documented
that the above constituents possess antimicrobial activity
and hence this confirms and reinforces our study on
antimicrobial activity of n-hexane extract of the stem bark
of B. purpurea [23-28].

CONCLUSION

The present study establishes that the n-hexane

Fig. 3: extract of stem bark of B. purpurea shows antibacterial as
well as antifungal activity. The phytochemical
investigation led to the isolation of steroids, triterpenoids,
alkaloids, fatty acids and phytol esters. The above
antimicrobial activity may be attributed to the presence of
these bioactive constituents in the plant.

ACKNOWLEDGEMENT

The authors would like to convey great appreciation

and gratitude towards National Chemical Laboratories
(NCL), Pune for their cooperation, fast service and
Fig. 4: support for providing microbial authentic cultures. Sincere

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heartfelt thanks to Dr. Gopalkrishna Rao, Principal, Goa 12. Desai, R.R., A.B. Joshi and M.P. Bhobe, 2013.
College of Pharmacy, Panaji- Goa, for extending a helping Phytochemical investigation of the hexane extract of
hand and encouraging our research programme. The stem bark of Bauhinia purpurea linn. Der Pharma
authors are also grateful to Prof. G. I. Hukkeri, Dept. of Chemica, 5(3): 116-121.
Botany, Dhempe College of Arts and Science, Miramar - 13. Sethil kumar, S. and M. Kamraj, 2011. Antimicrobial
Goa, India for his excellent contribution in the activity of Cucumis anguria L. By agar well diffusion
authentication of the plant material. method. Botany Research International, 4(2): 41-42.
14. Farrukh, R., M.A. Zargar, A. Akhtar, S.A. Tasduq and
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