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International Journal of Pharmagenesis

IJP 1(1), January-June 2010, pp. 113-117

Original Research Article

Biological Control of Phytopathogenic Bacteria Pantoea Agglomerans


Using Medicinal Plant Extracts

Varaprasad Bobbarala, 2Varahalarao Vadlapudi, 2K. Chandrasekhar Naidu and


1*

3
Somasekhar Penumajji
1
For U Biosciences, A/4A, Park lane Residency, East point colony, Visakhapatnam-17, A. P, India
2
Department of Botany, Andhra University, Visakhapatnam-3.
3
Vivimed labs Limited, 2nd, 4th Floor, Veeranag towers, Habsiguda, Hyderabad, A.P.

Abstract: We have examined antimicrobial activities of forty-nine medicinal plants extracts that have been
popularly used as folk medicines. Hexane, chloroform and methanolic extracts were investigated in vitro for
antibacterial activity against Pantoea agglomerans using agar well diffusion technique. The plant extracts inhibited
the growth of all the test organisms. P. pterophorus appears to be more effective as an antibacterial agent than all
the medicinal plant extracts tested. This study has to some extent validated the medicinal potential of medicinal
plants. Hence this plant can be further subjected to isolation of the therapeutic antimicrobials and further
pharmacological evaluation.
Keywords: Medicinal plants, Folk medicines, Antimicrobials, P. pterophorus, Pantoea agglomerans

Introduction used for folk medicinal purposes, are numerous


One of the greatest achievements of medical and diverse. Biologically active compounds
science has been the control and management of present in the medicinal plants have always been
infectious diseases. Indigenous plants, widely of great Interest to scientists working in this field.
used for folk medicinal purposes, are numerous Antimicrobial properties of medicinal plants are
and diverse. For a long period of time, plants have being increasingly reported from different parts
been a valuable source of natural products for of the world [1-3]. Although hundreds of plant
maintaining human health, especially n the last species have been tested for antimicrobial
decade, with more intensive studies for natural properties, the vast majority of have not been
therapies. Thus plants should be investigated to adequately evaluated [4].
better understand their properties, safety and In recent years this interest to evaluate plants
efficiency. The use of plant extracts and phyto possessing antibacterial activity for various
chemicals, both with known antimicrobial diseases is growing [5]. Thus, this research work
properties, can be of great significance in seeks to establish the microbial activities of some
therapeutic treatments. Many plants have been plants and to justify the ethno botanical uses of
used because of their antimicrobial traits, which some of these plants. Considering the vast
are due to compounds synthesized in the potentiality of plants as sources for antimicrobial
secondary metabolism of the plant. The potential drugs with reference to antibacterial and
of higher plants as source for new drugs is still antifungal agents, a systematic investigation was
largely unexplored. Indigenous plants, widely undertaken to screen the local flora for P.
agglomerans activity from forty nine medicinal
Corresponding Author: Varaprasad Bobbarala
E-mail: varaprasadphd@rediffmail.com plants. A medicinal use of plants is the oldest form
114 International Journal of Pharmagenesis, 1(1) 2010

of healthcare known to mankind. This situation, ml of nutrient agar was dispensed into sterile
the undesirable side effect of certain antibiotics, universal bottles these were then inoculated with
and the emergence of previously uncommon 0.2 ml of cultures mixed gently and poured into
infections, has forced scientists to look for new sterile petri dishes. After setting a number 3-cup
antimicrobial substances from various sources, borer (6mm) diameter was properly sterilized by
such as medicinal plants [6, 7]. A large number flaming and used to make three to five uniform
of plants in different location around the world cups/wells in each petri dish. A drop of molten
have been extracted and semi-purified to nutrient agar was used to seal the base of each
investigate individually their antimicrobial cup. The cups/wells were filled with 50µ of the
activity [8]. extract concentration of 500mg/ml, 250mg/ml
and 100mg/ml and allow diffusing for 45
Materials and Methods minutes. The solvents used for reconstituting the
extracts were similarly analyzed. The plates were
The medicinal plants belong to forty nine
incubated at 37°c for 24 hours for bacteria. The
different plant species were collected selectively
above procedure is allowed for fungal assays but
from Andhra University surroundings and
except the media potato dextrose agar instead of
nearest villages, Viskhapatnam, Andhra Pradesh,
nutrient agar and incubates at 25°c for 48 hours.
India. The authenticated samples of plant parts
The zones of inhibition were measured with
were shade dried, cut into small pieces and
antibiotic zone scale in mm and the experiment
powdered in a mixer grinder the residues (crude
was carried out in duplicates. The extracts and
extracts) obtained were finally dried under
the phytochemicals that showed antimicrobial
vacuum.
activity were later tested to determine the
The extraction method employed here is a Minimal Inhibitory Concentration (MIC) for each
known amount of coarsely powdered plant bacterial and fungal sample.
materials of different plant species were
successively extracted with organic solvents like Minimum Inhibitory Concentration (MIC) Assays
hexane, chloroform and methanol basing on order
Based on the preliminary screening chloroform
of their polarity using soxhlet apparatus. The
and methanolic extracts were found to have
different extracts obtained were subsequently
potent antimicrobial activity and Minimum
concentrated under reduced pressure to get their
Inhibitory Concentrations (MIC) of the extracts
corresponding residues. The extracts were
was determined according to Elizabeth, A final
screened for antimicrobial activity using the
concentration of 0.5% (v/v) Tween-20 (Sigma)
method described under the section
was used to enhance crude extract solubility. A
Pantoes agglomerans was procured from series of two fold dilution of each extract, ranging
Microbial Type Culture Collection MTCC, from 0.2 to 100 mg/ml, was prepared. After
IMTECH, Chandigarh. Active culture was sterilization, the medium was inoculated with 3µl
generated by inoculating a loop full of culture in aliquots of culture containing approximately 105
separate 100mL nutrient broth and incubating on CFU/ml of each organism of 24 hours slant
a shaker at 37 o C overnight. The cells were culture in aseptic condition and transferred into
harvested by centrifuging at 4000 rpm for 5 min, sterile 6 inch diameter petri dishes and allowed
washed with normal saline, spun at 4000 rpm for to set at room temperature for about 10 minutes
5 min again and diluted in normal saline to obtain and then kept in a refrigerator for 30 minutes.
5 x 105cfu/mL.
After the media solidified a number 3-cup
borer (6mm) diameter was properly sterilized by
Determination of Antimicrobial Activity flaming and used to make three to five uniform
The crude extracts of the different plant parts of cups/wells in each petri dish. A drop of molten
different species were subjected to antimicrobial nutrient agar was used to seal the base of each
assay using the agar well diffusion method of cup. Different plant crude extracts ranging from
Murray et al., [9] modified by Olurinola [10] 20 0.2 to 100 mg/ml were added to the cups/wells
Biological Control of Phytopathogenic Bacteria Pantoea Agglomerans using Medicinal Plant Extracts 115

of each petri dish and the control plates without Results and Discussion
plant extract. Inhibition of organism growth in (Graph 1a&b) In the present study different
the plates containing test crude extracts was concentrations of methanol extracts of inhibited
judged by comparison with growth in blank activity but their effectiveness varied. The zones
control plates. The MICs were determined as the of inhibition ranged from 7 to 24 mm.
lowest concentration of extracts inhibiting visible
growth of each organism on the agar plate. Graph 2 Minimum inhibitory concentration
Similarly the MICs of methanolic extracts were (MIC) (Volume per well: 50ìl, Borer size used:
determined against all other microorganisms. The 6mm, ‘x’ axis = Medicinal plants, ‘y’ axis =0-300
results were given in = Extract concentration in mg/ml)

0-25: Zone of Inhibition in mm; A: 100mg/ml; B: 300mg/ml; C: 500mg/ml; Includes Diameter of Well (6 mm)

0-25: Zone of Inhibition in mm; A: 100mg/ml; B: 300mg/ml; C: 500mg/ml; Includes Diameter of Well (6 mm)
116 International Journal of Pharmagenesis, 1(1) 2010

Successful prediction of botanical compounds antimicrobial compounds. The results of present


from plant material is largely dependent on the investigation clearly indicate that the antibacterial
type of solvent used in the extraction procedure. activity vary with the species of the plants and
Among the forty nine selected plants extracts plant material used. Thus, the study ascertains the
screened against P. agglomerans. Medicinal plants value of plants used as biocide which could be of
of A. vasica, B. diffusa, C. roseus, E. crassipes, and T. considerable interest to the development of new
cordifolia plants showed lowest value (7 mm) drugs. It can be concluded that the methanolic
whereas A. farnaciana, P. pterophorus and G. arborea extract of P. pterophorus showed high antibacterial
have significant activity. There is no antimicrobial activity against phytopathogenic bacterium P.
activity were found with A. ilcifolius, C. sativum, agglomerans. The antibacterial activity is in the
H. bengalenses, P. rubrum and S. grandiflora. function of the extract concentration. It is
Among the significant activity produced plants therefore recommended that the nature and the
P. pterophorus showed highest activity of 20mm number of the active antimicrobial principles
inhibition zone at even at low concentration and involved in P. pterophorus plant extract to be
MIC at 5µg/well (Graph.2). The antimicrobial studied in detail.
properties of medicinal plants have been
explained by the chemical association of active References
substances. The hexane extract appears to have [1] Saxena K. Antimicrobial Screening of Selected
less antibacterial and antifungal activity than the Medicinal Plants from India. Journal of
chloroform and methanolic extracts from the Ethnopharmacology 1997; 58(2): 75-83.
above results it can be concluded that methanol [2] Nimri L. F., Meqdam M. M., Alkofahi A. Antibacterial
extract of P. pterophorus have greater potential as Activity of Jordanian Medicinal Plants. Pharmacologycal
Biology 1999; 37(3): 196-201.
antimicrobial agent against P. agglomerans.
Comparisons with pertinent data from literature [3] Saxena V. K., Sharma R. N. Antimicrobial Activity of
Essential oil of Lankana Aculeata. Fitoterapia 1999; 70(1):
indicate that, according to the methodology 59-60.
adopted in studies on antimicrobial activity, the [4] Balandrin M. F., Klocke J. A., Wurtele E. S., Bollinger
most diverse results can be obtained. W. H. Natural Plant Chemicals: Sources of
Industrial and Medicinal Materials Science 1985; 228:
Conclusion 1154-1160.
[5] Clark A. M., Hufford C. D. American Chemical Society
Plant based products have been effectively (ACS Symposium Series 534), Washington, D.C; 1993.
proven for their utilization as source for pp. 228-241.
Biological Control of Phytopathogenic Bacteria Pantoea Agglomerans using Medicinal Plant Extracts 117

[6] Marchese A., Shito G. C. Resistance Patterns of Lower [8] Draughon F. A. Use of Botanicals as Bio Preservatives
Respiratory Tract Pathogens in Europe. Int J. in Foods. Food Technol. 2004; 58: 20-28.
Antimicrobial Agents 2001; 16: 25-29. [9] Murray P. R., Baron E. J., Pfaller M. A., Tenover F. C.,
[7] Poole K. Overcoming Antimicrobial Resistance by Yolken H. R. Manual of Clinical Microbiology, 6 th
Targeting Resistance Mechanisms. J. Pharmacy and Edition. ASM Press, Washington. DC 1995. 15-18.
Pharmacol, 2001; 53: 283-284. [10] Olurinola P. F. A Laboratory Manual of Pharmaceutical
Microbiology. Idu, Abuja, Nigeria 1996; 69-105.

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