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World Journal of Pharmaceutical Research

Boopathi et al. SJIF Impact


World Journal of Pharmaceutical Factor 7.523
Research
Volume 6, Issue 3, 617-627. Research Article ISSN 22777105

EVALUATION OF ANTIMICROBIAL ACTIVITY OF TECOMA STANS


AND MUNTINGIA CALABURA

T. Boopathi1*, K. Gopalasatheeskumar2, S. Parthiban1, G. Sangeetha3, M. Thanga


Kokila3 and T. Manimaran4

1
SSM College of Pharmacy, Jambai, Erode-638312, Tamilnadu, India.
2
Jaya College of Paramedical Sciences, College of Pharmacy, Thiruninravur, Chennai
602024 Tamilnadu, India.
3
Madurai Medical College, Madurai-625020, Tamilnadu, India.
4
Jkk Nataraja College of Pharmacy, Salem-638183. Tamilnadu, India.

ABSTRACT
Article Received on
25 Dec. 2016,
In the present study, the antimicrobial activity of leaves of Tecoma
Revised on 14 Jan. 2017, stans and Muntingia calabura were performed by using disc diffusion
Accepted on 04 Feb. 2017
method. The antimicrobial activity of the extracts were studied in
DOI: 10.20959/wjpr20173-7820
different concentrations against Eight pathogenic bacterial strains,
three Gram-positive (Staphylococcus aureus, Streptomyces filvissimus,
*Corresponding Author Basillus subtilis) and five Gram-negative (Escherichia
T. Boopathi
coli, Pseudomonas aeruginosa, Klebsiella pneumonia, proteus
SSM College of Pharmacy,
mirabilis, shigella flexneri) and two fungal strains (Aspergillus niger &
Jambai, Erode-638312,
Tamilnadu, India. Candida albicans). These strains have been selected for the basis of its
application purpose of further formulation study. The ethanolic extract
of Tecoma stans was found to be active on most of the clinically isolated microorganism and
fungi, as compare with standard drugs. The activities of these extracts are found to be quiet
comparable with the standard antibiotics screened under similar conditions. So these extracts
can be used as an external antiseptic in prevention and treatment of bacterial infections. The
present study justified the claimed uses of leaves in the traditional system of medicine to treat
various infectious disease caused by the microbes.

KEYWORDS: Tecoma stans, Muntingia calabura, Disc diffusion method, Antimicrobial,


Ethanolic extract.

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INTRODUCTION
A majority of the world's population in developing countries still relies on herbal medicines
to meet its health needs. Herbal medicines are often used to provide first-line and basic health
service, both to people living in remote areas where it is the only available health service, and
to people living in poor areas where it offers the only affordable remedy. Even in areas where
modern medicine is available, the interest on herbal medicines and their utilization have been
increasing rapidly in recent years.

Microbial infections pose a health problem throughout the world, and plants are a possible
source of antimicrobial agents. Medicinal plants contain active principles which can be used
as an alternative to cheap and effective herbal drugs against common bacterial infections.[1]
Even though pharmacological industries have produced a number of new antibiotics in the
last three decades, resistance to these drugs by microorganisms has increased. In general,
bacteria have the genetic ability to transmit and acquire resistance to drugs, which are utilized
as therapeutic agents.

A wide range of antibiotics (e.g. erythromycin, tetracyclines, trimethoprim, sulfonamides,


gentamicin, etc) are being used at present for treating wound infections. Bacterial resistance
to antibiotics is a major therapeutic problem and the rate at which new antibiotics are being
produced is slowing.[2] The increasing failure of chemotherapeutics and antibiotic resistance
exhibited by pathogenic microbial infectious agents, has led to the screening of several
medicinal plants for their potential antibacterial activity. There are several reports in the
literature regarding the antibacterial activity of plant extracts.[3]

According to one of the hypotheses discussed in the literature, molecules of antimicrobial


peptides interact with the bacterial membrane giving rise to the formation of a
transmembrane cluster (probably, an ion channel). This causes decrease in the membrane
potential value and subsequent cytolysis.[4] The present study was conducted to compare the
antimicrobial activity from two medicinal plants Tecoma stans and Muntingia calabura. The
therapeutic utility and efficacy of these plants in various diseases like diabetes, diuretic,
cancer, lungs cancer, stomach pain, gastric ulcer is well documented among ethnic
populations. Here we are using different stains to find out the antimicrobial activity such as
S.fulvissimus, K.Pneumonia, S.flexneri, E.coli, B.subtitis, S.pyrogenes, P.auruginosa,
P.mirabis, A.niger, C.albicans.[5,6]

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Boopathi et al. World Journal of Pharmaceutical Research

The present work was aimed to study antimicrobial activity of Tecoma stans and Muntingia
calabura. To achieve the objectives the plan is executed to extraction of the plant and
Evaluation of Antimicrobial activity.

METERIALS AND METHODS


Meterials
The leaves of Tecoma stans and Muntingia calabura were collected from the surrounding
areas of Erode district, Tamilnadu, India. The samples were identified and authenticated
(Reg.No: PARC/2015/3073) at Institute of Herbal Botany, PLANT ANATOMY
RESEARCH CENTRE, Chennai.

Methods
Extraction procedure
The leaves were dried under shade and then made in to a coarse powder with a mechanical
grinder. Powdered dried leaves were passed through sieve No. 40 and stored in an airtight
container for further use. Both petroleum ether and ethanol extraction were carried out by
using Soxhlet extractor.

Preparation of petroleum ether extract


The dried powder material of leaves (100gm) were first extracted with petroleum ether (60-
80o) in a Soxhlet apparatus and after complete extraction (48 hrs), the solvent was removed
by distillation under reduced pressure and resulting semisolid mass was vacuum dried using
rotary flash evaporator.

Preparation of ethanolic extract


After the extraction with petroleum ether the same plant material were dried and again
extracted with ethanol (99.9% v/v) in Soxhlet apparatus and after complete extraction (48 hr)
the solvent was removed by distillation under reduced pressure and resulting semisolid mass
was vacuum dried using rotary flash evaporator.

All the extracts were subjected to various chemical tests to detect the presence of different
phytoconstituents.

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Anti-microbial activity
Test Microorganisms
Gram-positive organisms (Staphylococcus aureus, streptomyces fulvissimus, Bacillus
subtilis),Gram-Negative organisms (Klebsiella pneumoniae, Pseudomonas aeruginosa,
Escherichia coli, Shigella flexneri, proteus mirabilis) and a fungal yeast Candida albicans
were chosen based on their clinical and pharmacological importance. They were maintained
on Mueller-Hinton Agar medium. Twenty-four hour old pure cultures were prepared for use
each time.

Growth media
The bacterial and fungal stock cultures were incubated for 24 hours at 37C on nutrient agar
and potato dextrose agar (PDA) medium respectively, following refrigeration storage at 4C.
The bacterial strains were grown in Mueller-Hinton agar (MHA) plates at 37C (the bacteria
were grown in the nutrient broth at 37C and maintained on nutrient agar slants at 4C),
whereas the fungal strains were grown in Sabouraud dextrose agar and PDA media,
respectively, at 28C. The stock cultures were maintained at 4C. These mediums were
prepared according to the manufacturers instruction, autoclaved and dispensed at 30 ml per
plate in 12 x 12cm Petri dishes. Set plates were incubated overnight to ensure sterility before
use.[7,8]

Experimental Procedure
The agar diffusion method was used to evaluate the antimicrobial activity. Bacteria were
cultured overnight at 370C in Mueller Hinton Broth and fungi at 280C for 48hrs in Potato
Dextrose Broth used as inoculums. Final inoculums, using 100-1 of suspension containing
108 CFV/ml of bacteria and 104 spore/ml of fungi spread on Mueller Hinton Agar (MHA) and
Potato Dextrose agar (PDA) medium, respectively. The disc (6mm in diameter) was
impregnated with 10l of 100mg/ml (1mg/disc) extracts placed on seeded agar.
Oxytetracyclines (10g/disc) was used as positive control for bacteria and Ketoconzole
(10g/disc) for fungi. The test plates were incubated at 370C for 24hrs for bacteria and at
280C for 48hrs for fungi depending on the incubation time required for a visible growth.At
the end of the incubation period the media were observed for Zone of inhibition.[9,10]

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RESULTS AND DISCUSSION


Qualitative Phytochemical Analysis
In the phytochemical studies, petrolium ether, ethanolic extracts of leaves of Tecoma stans
and Muntingia calabura showed the presence various phytoconstituents.

Preliminary phytochemical screening of Tecoma stans revealed the presence of bioactive


compounds such as glycosides, alkaloids, tannins, flavonoids and saponins in ethanolic
extract and fixed oil,gums, mucilage in petroleum ether extract. On the other hand the
petroleum ether extract of Mutingia calabura contained fixed oil, steroids and tannins. While
the ethanolic extract showed the presence of carbohydrates, glycosides, tannins, flavonoids,
steroids, saponins and terpenoids. Different phytochemicals have been found to possess a
wide range of activities, which may help in protection against chronic diseases.The results are
given in Table-1.

Table 1: Data showing the preliminary phytochemical screening of the leaves extracts
of Tecoma stans and Muntingia calabura.
Tecoma stans Muntingia calabura
Phytoconstituents Petroleum Ethanol Petroleum Ethanol
Ether (99.9% v/v) Ether (99.9% v/v)
Carbohydrate Absent Absent Absent Present
Glycosides Absent Present Absent Present
Alkaloids Absent Present Absent Absent
Phytosterol and
Absent Absent Present Present
steroids
Protein& Amino Acid Absent Absent Absent Absent
Tannins Absent Present Present Present
Flavonoids Absent Present Absent Present
Saponins Absent Present Absent Present
Terpenoids Absent Absent Absent Present
Fixed oil and fats Present Absent Present Absent
Gums and mucilage Present Absent Present Absent

ANTIMICROBIAL ACTIVITY
Antimicrobial properties of medicinal plants are being increasingly reported from different
parts of the world. The World Health Organization estimates that plant extract or their active
constituents are used as folk medicine in traditional therapies of 80% of the world's
population. In the present work, the extracts obtained from Tecoma stans and Muntingia
calabura show strong activity against most of the tested bacterial and fungal strains. The
results were compared with standard antibiotic drugs.

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Table 2: Antibacterial study of ethanolic extract of Tecoma stans


1000g/ml 500g/ml 250g/ml 125g/ml 62.5g/ml Oxytetracycline
Microorganisms
(mm) (mm) (mm) (mm) (mm) (1mg/ml) (mm)
Escherichia coli
14 10 09 06 0 15
(MTCC 1687)
Klebsiella pneumonia
16 08 04 03 0 18
(MTCC 7162)
Proteus mirabilis
12 10 09 07 02 13
(MTCC 9242)
Shigella flexneri
14 08 05 0 0 15
(MTCC 1457)
Stephylococcus
aureus 17 13 08 02 0 20
(MTCC 96)
Streptomyces
Fulvissimus 15 08 06 03 0 16
(MTCC 7336)
Bacillus subtilis
21 14 04 0 0 22
(MTCC 736)
Pseudomonas
Aeruginosa 10 04 0 0 0 11
(MTCC 2488)

Table 3: Antifungal study of ethanolic extract of Tecoma stans.


1000g/ml 500g/ml 250g/ml 125g/ml 62.5g/ml Ketaconazole(1mg/ml)
Microorganisms
(mm) (mm) (mm) (mm) (mm) (mm)
Candida albicans
15 08 07 05 0 17
(MTCC 282)
Aspergillus niger
18 09 07 06 04 20
(MTCC 183)

Table 4: Antibacterial study of ethanolic extract of Muntingia calabura


1000g/ml 500g/ml 250g/ml 125g/ml 62.5g/ml Oxytetracycline
Microorganisms
(mm) (mm) (mm) (mm) (mm) (1mg/ml) (mm)
Escherichia coli
13 10 05 0 0 14
(MTCC 1687)
Klebsiella pneumonia
14 11 03 0 0 16
(MTCC 7162)
Proteus mirabilis
15 14 12 07 03 18
(MTCC 9242)
Shigella flexneri
12 11 0 0 0 13
(MTCC 1457)
Stephylococcus aureus
17 13 08 05 0 18
(MTCC 96)
Streptomyces
Fulvissimus 13 11 08 07 0 15
(MTCC 7336)
Bacillus subtilis 18 16 10 08 05 21

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(MTCC 736)
Pseudomonas
Aeruginosa 08 05 0 0 0 15
(MTCC 2488)

Table 5: Antifungal study of ethanolic extract of Muntingia calabura.


Ketaconazole
1000g/ml 500g/ml 250g/ml 125g/ml 62.5g/ml
Microorganisms (1mg/ml)
(mm) (mm) (mm) (mm) (mm)
(mm)
Candida albicans
0 0 0 0 0 16
(MTCC 282)
Aspergillus niger
16 10 06 03 0 18
(MTCC 183)

Ethanolic extract of Tecoma Ethanolic extract of Muntingia


Micro-organism
stans calabura

Escherichia coli

Klebsiella
pneumonia

Proteus mirabilis

Shigella flexneri

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Stephylococcus
aureus

Streptomyces
fulvissimus

Basillus subtilis

Pseudomonas
aeruginosa

Fig. 1-32: Antibacterial activity of ethanolic extracts of Tecoma stans and Muntingia
calabura.

Microorganism Ethanolic extract of Tecoma stans Ethanolic extract of Muntingia calabura


Aspergilus
niger

Candida
albicans

Fig. 33-40: Antifungal activity of ethanolic extracts of Tecoma stans and Muntingia
calabura.

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Boopathi et al. World Journal of Pharmaceutical Research

In the present study, the antimicrobial activity of leaves of Tecoma stans and Muntingia
calabura were performed by using disc diffusion method. The antimicrobial activity of the
extracts were studied in different concentrations (62.5, 125, 250, 500, and 1000 g/ml)
against Eight pathogenic bacterial strains, three Gram-positive (Staphylococcus
aureus, Streptomyces filvissimus, Basillus subtilis,) and five Gram-negative (Escherichia
coli, Pseudomonas aeruginosa, Klebsiella pneumonia, proteus mirabilis, shigella flexneri) and
two fungal strains (Aspergillus niger & Candida albicans). These strains have been selected
for the basis of its application purpose of further formulation study.

As compared with standard drugs, the results revealed that in the extracts for bacterial
activity against staphylococcus aureus and Basillus subtilis were more sensitive as compared
with Pseudomonas aeruginosa , and for fungal activity, Aspergillus niger shows good result
as compare with Candida albicans .The result of this study showed that ethanolicextracts of
Tecoma stans have varied antibacterial activities against the tested organisms The growth
inhibition zone measured ranged from 10-21mm for all the sensitive bacteria, and ranged
from 15- 18mm for fungal strains.

Muntingia calaburaleaf extract also showed appreciable activity on all test bacteria
(0818mm inhibition zone) except Candida albicans fungal strain. It did not inhibit Candida
albicans fungal strain.

The ehanolic extract of Tecoma stans showed greater antimicrobial activities than the
Muntingia calabura extract. The leaf ethanolic extract of Tecoma stans showed pronounced
antibacterial and antifungal activities against all the microorganisms tested (10-21mm and
15-18mm inhibition zone).[11,12]

The antimicrobial effect of ethanol extracts against these organisms may be due to the ability
of the ethanol to extract some of the active properties of these plants glycosides, alkaloids,
tannins, flavonoids, saponins and other secondary metabolites which are reported to be
antimicrobial.[13,14] The results show that the extract of Tecoma stans was found to be more
effective against all the microbes tested compared to Muntingia calabura extract.

CONCLUSION
In the present study, the herbal extracts of Tecoma stans and Muntingia calabura leaves
examined for antimicrobial activity. The ethanolic extract of Tecoma stans was found to be

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Boopathi et al. World Journal of Pharmaceutical Research

active on most of the clinically isolated microorganism and fungi, as compare with standard
drugs. The activities of these extracts are found to be quiet comparable with the standard
antibiotics screened under similar conditions. So these extracts can be used as an external
antiseptic in prevention and treatment of bacterial infections. The present study justified the
claimed uses of leaves in the traditional system of medicine to treat various infectious disease
caused by the microbes. However, further studies are needed to better evaluate the potential
effectiveness of the crude extracts as the antimicrobial agents. The present results will form
the basis for selection of plant species for further investigation in the potential discovery of
new natural bioactive compounds. Further studies which aimed at the isolation and structure
elucidation of antibacterial active constituents from the plant have been initiated.

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