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Research Article
Antimicrobial Activities of Leaf Extracts of
Guava (Psidium guajava L.) on Two Gram-Negative
and Gram-Positive Bacteria
Copyright 2013 Bipul Biswas et al. This is an open access article distributed under the Creative Commons Attribution License,
which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.
Aim. To determine the antimicrobial potential of guava (Psidium guajava) leaf extracts against two gram-negative bacteria
(Escherichia coli and Salmonella enteritidis) and two gram-positive bacteria (Staphylococcus aureus and Bacillus cereus) which are
some of foodborne and spoilage bacteria. The guava leaves were extracted in four different solvents of increasing polarities (hexane,
methanol, ethanol, and water). The efficacy of these extracts was tested against those bacteria through a well-diffusion method
employing 50 L leaf-extract solution per well. According to the findings of the antibacterial assay, the methanol and ethanol
extracts of the guava leaves showed inhibitory activity against gram-positive bacteria, whereas the gram-negative bacteria were
resistant to all the solvent extracts. The methanol extract had an antibacterial activity with mean zones of inhibition of 8.27 and
12.3 mm, and the ethanol extract had a mean zone of inhibition of 6.11 and 11.0 mm against B. cereus and S. aureus, respectively.
On the basis of the present finding, guava leaf-extract might be a good candidate in the search for a natural antimicrobial agent.
This study provides scientific understanding to further determine the antimicrobial values and investigate other pharmacological
properties.
1. Introduction that help to treat and manage various diseases. The many
parts of the plant have been used in traditional medicine
Recently there has been a lot of attention focused on pro- to manage conditions like malaria, gastroenteritis, vomiting,
ducing medicines and products that are natural. Several diarrhea, dysentery, wounds, ulcers, toothache, coughs, sore
fruits and fruit extracts, as well as arrowroot tea extract [1] throat, inflamed gums, and a number of other conditions
and caffeine [2], have been found to exhibit antimicrobial [35]. This plant has also been used for the controlling
activity against E. coli O157:H7. This suggests that plants of life-changing conditions such as diabetes, hypertension,
which manifest relatively high levels of antimicrobial action and obesity [3, 610]. In this study, we aim to evaluate
may be sources of compounds that can be used to inhibit the total extracts of P. guajava leaves, growing at Fort
the growth of foodborne pathogens. Bacterial cells could be Valley State University, using various aqueous and organic
killed by the rupture of cell walls and membranes and by the solvents to establish if it is effective against killing or
irregular disruption of the intracellular matrix when treated inhibiting the growth of foodborne bacterium Staphylo-
with plant extracts [1]. coccus aureus, Escherichia coli, Salmonella enteritidis, and
The guava (Psidium guajava) is a phytotherapic plant used Bacillus cereus which can cause foodborne illness and
in folk medicine that is believed to have active components spoilage.
2 International Journal of Microbiology
(a)
(b)
(c)
(d)
(e)
Figure 2: (a) Existence of saponin tests; L to R: n-hexane, methanol, ethanol, and distilled water extracts. (b) Existence of phenols and tannins
tests; L to R: n-hexane, methanol, ethanol, and distilled water extracts. (c) Existence of terpenoids tests; L to R: n-hexane, methanol, ethanol,
and distilled water extracts. (d) Existence of flavonoids tests; L to R: n-hexane, methanol, ethanol, and distilled water extracts. (e) Existence
of glycosides tests; L to R: n-hexane, methanol, ethanol, and distilled water extracts.
of guava leaf and was found that a compound called aglycone wrapped in aluminum foil to avoid evaporation and exposure
quercetin is responsible for spasmolytic activities, which is to light for 3 days at room temperature. The flasks were placed
formed when flavonoids of guava leaves are hydrolyzed by the on a platform shaker at 70 rpm. After 3 days of soaking in
gastrointestinal fluids. solvent, the mixtures were transferred to 50 mL tubes and
centrifuged for 10 min at 4,000 rpm at 25 C. The supernatant
2. Materials and Methods was collected and stored at 4 C until use.
2.1. Preparation of Plant Extract. The leaf samples were 2.3. Phytochemical Analysis. Chemical tests for the screening
collected from the guava trees growing at the Specialty Plant and identification of bioactive chemical constituents in the
House at Fort Valley State University. Random leaf samples guava were carried out with the extracts using the standard
were collected into plastic zip lock bags with appropriate procedure as described [3234]. For each test, 1 mL of each
labeling and stored in an ice cooler until being transported solvent extract was used for analysis, in exception for the
to the laboratory for extraction. saponin test in which 3 mL solvent extract was used.
2.6. Test for Terpenoids (Salkowskis Test). Extract was mixed Table 1: Phytochemical constituents of Psidium guajava extracts.
with 2 mL of chloroform. Then 2 mL of concentrated sulfuric
acid was added carefully and shaken gently. A reddish brown Phenols
Extracts and Saponins Terpenoids Flavonoids Glycosides
coloration of the interphase was formed to show positive
tannins
results for the presence of terpenoids (Figure 2(c)).
n-Hexane
Methanol + + + +
2.7. Test for Flavonoids (Shinoda Test). Extract was mixed
with magnesium ribbon fragments, and concentrated Ethanol + + + +
hydrochloric acid was added drop wise. Orange, red, pink, Distilled
+ + + + +
or purple coloration indicates the presence of flavonoids water
(Figure 2(d)). +: presence of constituent (positive); : absence of constituent (negative).
Table 2: Antibacterial activity of Psidium guajava leaves of the properties of guava. The results also indicate that the plant
screened solvents extracts. extracts have no antibacterial effect on the Gram-negative
Zone of inhibition (mm) bacteria, showing that they do not contain active ingredients
Plant extracts against the organisms. The observed inhibition of Gram-
B. cereus S. aureus E. coli S. entertidis
positive bacteria, Bacillus cereus and Staphylococcus aureus,
n-Hexane
suggests that guava possesses compounds containing antibac-
Ethanol 6.11 0.60 11.0 0.52 terial properties that can effectively suppress the growth
Methanol 8.27 0.44 12.3 0.78 when extracted using methanol or ethanol as the solvent.
Water Comparisons with related data from the literature indicate
Inhibition zones are the mean including borer (5 mm) diameter standard that according to the different methodologies of studies
deviation. on antibacterial activity, the most diverse outcomes can be
: no inhibitory activity. obtained. This study provides scientific insight to further
determine the antimicrobial principles and investigate other
pharmacological properties of guava. On the basis of the
3.2. Antibacterial Activity. The results of the study indi- present finding, P. guajava leaves possess the capabilities
cated that only two of the crude solvent extracts prepared of being a good candidate in the search for a natural
from the leaves of Psidium guajava, methanol and ethanol, antimicrobial agent against infections and/or diseases caused
showed inhibitory activity against bacteria (Table 2). Only by B. cereus and S. aureus.
Gram-positive bacteria, Bacillus cereus and Staphylococcus
aureus, were susceptible to the two extracts, while neither
of the Gram-negative bacterium showed any inhibition. At Acknowledgments
10 mg/50 L, the methanol extract had a slightly higher This work was apprehended by the financial support provided
antibacterial activity with mean zones of inhibition 8.27 and to one of the authors, Kimberly Rogers, (MS student) through
12.3 mm than ethanol extract with mean zone of inhibition the Advancing Graduate Education in the STEM Disciplines
6.11 and 11.0 mm against B. cereus and S. aureus, respec- for the Underserved African American and Low Income
tively. The resistance of the Gram-negative bacteria could be American Population grant (FVSU Project no. 20000-
attributed to its cell wall structure. Gram-negative bacteria 1017000-11100-61000-331079, Dr. Anand K. Yadav, PD/PI)
have an effective permeability barrier, comprised of a thin funded by the College of Agriculture, Family Sciences and
lipopolysaccharide exterior membrane, which could restrict Technology, Fort Valley State University by the United States
the penetration of the extruding the plant extract. It has Department of Education.
been reported earlier that Gram-negative bacteria are usually
more resistant to the plant-origin antimicrobials and even
show no effect, compared to Gram-positive bacteria [42 References
44]. Gram positive bacteria have a mesh-like peptidoglycan
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