In-vitro Antibacterial and Antioxidant Activities of Sandalwood (Santalum Album)

Abstract

Santalum album commonly known as Sandalwood is used traditionally for health and
wellness. It is an evergreen and hemi-parasitic tree and has a long history in Indian religious
rituals and traditional Chinese medicine. Due to its wide application in cosmetics and
therapeutics, we have done this study to explore the possibility of using aqueous extract
of S. album as antibacterial and antioxidant agent. The S. album extract was prepared
in distilled water. The activity of aqueous extract was evaluated against eight bacterial
pathogens including two strains of Escherichia coli, one each of Klebsiella pneumoniae,
Staphylococcus aureus, Bacillus subtilis, Pseudomonas aeruginosa, Aeromonas species
and Klebsiella oxytoca. The anti-oxidant activity was analyzed by two most common radical
scavenging assays of FRAP (ferric reducing antioxidant power) and DPPH (1,1- diphenyl-
2-picrylhydrazyl). Results showed that S. album had strongest inhibitory activity against S.
aureus (MTCC 902) i.e. 87% whereas; it showed no inhibition against E.coli (ATCC 25922)
and B. subtilis (MTCC736). The S. album extract showed DPPH radical scavenging activity
in a concentration–dependent manner with maximum scavenging of 64% in presence of
500μl of aqueous extract. The FRAP assay also proved antioxidant potential of S. album
with the highest value of 0.628mM at 200μl of aqueous extract.

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Citation: Shamsi TN, Parveen R, Afreen S, Azam M, Fatma T, Haque QMR and Fatima S. In-vitro Antibacterial
and Antioxidant Activities of Sandalwood (Santalum Album). Austin J Biotechnol Bioeng. 2014;1(2): 3.
Austin J Biotechnol Bioeng - Volume 1 Issue 2 - 2014
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Fatima et al. © All rights are reserved
Austin Journal of Biotechnology &
Bioengineering
Open Access
Full Text Article
Abstract
Santalum album commonly known as Sandalwood is used traditionally for health and
wellness. It is an evergreen and hemi-parasitic tree and has a long history in Indian religious
rituals and traditional Chinese medicine. Due to its wide application in cosmetics and
therapeutics, we have done this study to explore the possibility of using aqueous extract
of S. album as antibacterial and antioxidant agent. The S. album extract was prepared
in distilled water. The activity of aqueous extract was evaluated against eight bacterial
pathogens including two strains of Escherichia coli, one each of Klebsiella pneumoniae,
Staphylococcus aureus, Bacillus subtilis, Pseudomonas aeruginosa, Aeromonas species
and Klebsiella oxytoca. The anti-oxidant activity was analyzed by two most common radical
scavenging assays of FRAP (ferric reducing antioxidant power) and DPPH (1,1- diphenyl-
2-picrylhydrazyl). Results showed that S. album had strongest inhibitory activity against S.
aureus (MTCC 902) i.e. 87% whereas; it showed no inhibition against E.coli (ATCC 25922)
and B. subtilis (MTCC736). The S. album extract showed DPPH radical scavenging activity
in a concentration–dependent manner with maximum scavenging of 64% in presence of
500μl of aqueous extract. The FRAP assay also proved antioxidant potential of S. album
with the highest value of 0.628mM at 200μl of aqueous extract.
Keywords: Antibacterial activity; Antioxidant activity; Santalum album; Aqueous extract
Material and Methodology
Plant material
Sandalwood purchased from Local Ayurvedic Clinic.
Chemicals and reagents
All solvents and chemicals (analytical grade) used for antioxidant
and antibacterial assay were purchased from Merck and Himedia.
DPPH and TPTZ were purchased from Sigma-Aldrich.
Test microorganism
e following eight clinical isolates of bacteria were used for
the study: Two species of Escherichia coli, one each of Klebsiella
pneumoniae, Staphylococcus aureus, Bacillus subtilis, Pseudomonas
aeruginosa, Aeromonas species and Klebsiella oxytoca. All these
cultures were maintained on nutrient agar plates at 4OC.
Methodology
Preparation of aqueous extract
e Sandalwood was ground nely and then strained through
muslin cloth. 1 gram of sample was soaked for 2 hours in 20 ml of
distilled water (50 mg/ml). e sample was then centrifuged and
the supernatant was picked which served as aqueous extract for the
further studies.
Antibacterial Assay
Antimicrobial activity of the aqueous extract was tested against
three gram-positive bacteria (Bacillus subtilis, Aeromonas species.
and Staphylococcus aureus) and ve gram-negative bacteria (two of
Escherichia coli and one each of Klebsiella pneumoniae, Klebsiella
oxytoca and Pseudomonas aeruginosa). Overnight cultures were
prepared in Luria broth (LB) media by inoculation with a single
Abbreviations
DPPH: 1,1-diphenyl-2-picrylhydrazyl; FRAP: Ferric Reducing
Antioxidant Power; mg: Milligram; ml: Milliliter; CFU: Colony
Forming Units; OD: Optical Density; TPTZ: 2,4,6-Tripyridyl-s-
Triazine
Introduction
In present world of medical and pharmaceutical advancement,
microbes have evolved with resistance against the drugs by changing
their metabolism and genetic structure [1,2]. ese drug resistant
microorganisms are more pathogenic with high mortality rate and
have become a threat against human race. To overcome microbial
drug resistance, scientists are looking forward for the development of
alternative and novel drugs [3]. Natural sources such as plants, algae
and animals provide an array of natural medicinal compounds for
the treatment of various infectious diseases [4-6]. Studies by various
researchers have proved that plants are one of the major sources for
drug discovery and development [7,8].
Free radicals are inevitably produced in biological systems
and also encountered exogenously, and are known to cause
various degenerative disorders like mutagenesis, carcinogenesis,
cardiovascular disturbances and ageing [9].
Sandalwood, Santalum album, has been used since ancient times
for religious purposes in incense, in fragrances, and as medicine.
Various types of sandalwood trees grow in dierent countries of the
world [10].
e present research has been conducted to study the medicinal
properties like antimicrobial and antioxidant potential of aqueous
extract of Santalum album so that they can be a hope in the eld of
phytodrugs.
Research Article
In-vitro Antibacterial and Antioxidant Activities of
Sandalwood (Santalum Album)
Shamsi TN1¶, Parveen R1¶, Afreen S2, Azam M2,
Fatma T2, Haque QMR2 and Fatima S1*
1Department of Biotechnology, Jamia Millia Islamia,
India
2Department of Biosciences, Jamia Millia Islamia, India
¶These authors contributed equally to this work
*Corresponding author: Sadaf Fatima, Department of
Biotechnology, Jamia Millia Islamia, New Delhi-110025,
India
Received: July 10, 2014; Accepted: August 10, 2014;
Published: August 11, 2014
Austin
Publishing Group
A

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colony from agar plates and incubated at 37OC for 12 hrs. Overnight
cultures were diluted with fresh LB media to approximately 104
colonies forming units (CFU) and incubated at 37OC for 12-14 hrs
in the presence of S. album compared to the growth of the control
culture where only media and bacterial inoculum was taken. e
experiment was repeated twice for the conrmation. e percentage
inhibition was calculated by using the formula:
Percentage Inhibition (%) = [(dc - dt)/dc] x 100,
where dc and dt represent OD600 of control and treated sample
strains respectively.
Antioxidant Activity
DPPH Assay
e antioxidant activity of S. album and the standard was checked
on the basis of the free radical scavenging eect of the stable 1,
1-diphenyl-2-picrylhydrazyl (DPPH) by the method of Braca et al.
with minor modications [11]. A range of diluted working solutions
of the S. album were prepared in distilled water. Ascorbic acid (1
mg/ml) in distilled water was used as standard. 0.1mM DPPH was
prepared in ethanol and 500μl of this solution was mixed with 500μl
of working sample solutions and standard solution separately. ese
solution mixtures were kept in dark for 30 min and optical density
was measured at 517 nm using Spectrophotometer. 0.1mM DPPH
solution was used as control. e range of diluted aqueous extracts
was taken as blank. e optical density were recorded and DPPH
scavenging was calculated using the formula given below:
DPPH scavenging Activity (%) = [(dc - dt)/dc] x 100,
where dc and dt represent OD517 of control and test sample
respectively.
FRAP Assay
Antioxidant activity assay was also done following the ferric-
reducing antioxidant power (FRAP) method described by Benzie &
Strain method with minor modications [12]. FRAP reagents was
freshly prepared by mixing 10 ml acetate buer (300mM, pH 3.6),
1 ml 2,4,6-tris (2-pyridyl)-S-triazine (TPTZ) solution (10mM TPTZ
in 40mM/L HCl) and 1 ml FeCl3 (20mM) water solution. A range of
diluted working solutions of the S. album were prepared in distilled
water. Each sample (200 μl) was added in 1.5 ml of freshly prepared
FRAP reagent and mixed and aer 5 min, absorbance was measured
at 593 nm, using FRAP working solution as blank. Ascorbic acid
was used as standard. e results were expressed in mM Fe2+/ml of
aqueous extract. Higher absorbance indicates higher reducing power.
Results
Antibacterial Assay
Antimicrobial assay of the aqueous extract was examined against
various bacterial strains by accessing the percentage inhibition in
presence of S. album compared to the control where only media and
cultures were added. e results suggested that S. album exhibits
bactericidal property in-vitro i.e. the growth of microorganisms was
inhibited in its presence as shown in Table 1.
It was found that S. album had strongest inhibitory activity against
S. aureus (MTCC 902) i.e. 87% whereas, it showed no inhibition
against E.coli (ATCC 25922) and B. subtilis (MTCC736) as shown in
Figure 1.
Antioxidant Activity
DPPH assay
DPPH radical scavenging assay is the most widely used method
for screening antioxidant activity, since it can accommodate many
samples in a short period and detect active ingredients at low
concentration. e decrease in the absorbance of the DPPH radical
caused by antioxidant was due to the scavenging of the radical by
hydrogen donation. It is visually noticeable as the colour changes
from purple to yellow. S. album showed DPPH radical scavenging
activity in a concentration–dependent manner as shown in the gure
(Figure 2).
FRAP Assay
e ferric reducing antioxidant power of S. album is presented
in gure (Figure 3). e results showed that FRAP value of S. album
increase in the concentration-dependent manner. e highest
absorbance of FRAP was observed in S. album at 200 μl and the lowest
was that in at 20μl i.e 0.628 and 0.078 as compared to standard i.e.
0.210 and 0.965 respectively. ese concentrations were eective to
react with ferric tripyridyltriazine (FeIII-TPTZ) complex and produce
a blue colored ferrous tripyridyltriazine (FeII-TPTZ). From the
observations, it is clear that S. album showed fair antioxidant activity
comparable to ascorbic acid.
S.No. Bacterial strain Percentage Inhibition
E.coli (ATCC 25922) 0
K. pneumoniae (ATCC 700603) 69.2
S. aureus (MTCC 902) 87
B. subtilis (MTCC 736) 0
E.coli (MTCC 443) 78.1
P. aeruginosa (MTCC 2453) 65.5
Aeromonas spp. (A10 MDR) 76.8
K. oxytoca (A13 MDR) 77.4
Table 1: Percentage Growth inhibition of various bacterial strains in presence
of S. album.
0
10
20
30
40
50
60
70
80
90
100
E.coli (ATCC
25922)
K.
pneumoniae
(ATCC
700603)
S. aureus
(MTCC 902)
B. subtilis
(MTCC 736)
E.coli (MTCC
443)
P.
aeruginosa
(MTCC 2453)
Aeromonas
spp. (A10
MDR)
K. oxytoca
(A13 MDR)
% inhibition
Microorganisms
Figure 1: Graphical representation of % inhibition of Antibacterial potential
of S. album.

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Discussion
Antioxidant and antimicrobial properties possessed by various
plant extracts have recently been of great interest in both research
and food industry, due to their possible use as natural additives which
emerged from a growing tendency to replace synthetic antioxidants
0
10
20
30
40
50
60
70
80
90
100
0 100 200 300 400 500 600
DPPH radical scavenging
Amount(μl)
S. album
Ascorbic Acid
Figure 2: DPPH scavenging assay of S. album in comparison with Ascorbic
acid.
0
0.2
0.4
0.6
0.8
1
1.2
0 50 100 150 200 250
FRAP value (mM)
Amount (μl)
Ascorbic acid
S. album
Figure 3: FRAP values of S. album in comparison with Ascorbic acid.
with natural ones. Owing to the antioxidant and antibacterial activities
exhibited by the plant extracts, we have done this study to nd out
their possible role in food and pharmaceutical industries. Also,
results show that aqueous extract of S. album possesses the potent
antioxidant & antimicrobial substances which may be responsible for
its anti-tumor, anti-carcinogenic activity and remedy for hepatitis
B viral infection mechanism as well as justify the basis of using this
plant’s extract as traditional remedies.
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Citation: Shamsi TN, Parveen R, Afreen S, Azam M, Fatma T, Haque QMR and Fatima S. In-vitro Antibacterial
and Antioxidant Activities of Sandalwood (Santalum Album). Austin J Biotechnol Bioeng. 2014;1(2): 3.
Austin J Biotechnol Bioeng - Volume 1 Issue 2 - 2014
Submit your Manuscript | www.austinpublishinggroup.com
Fatima et al. © All rights are reserved