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Antimicrobial potentiality ofPhyllanthus amarus against drugresistant pathogensAvijit Mazumder a , Arun Mahato a & Rupa Mazumder aa Department of Pharmaceutical Sciences , Birla Institute ofTechnology , Mesra, Ranchi – 835 215, IndiaPublished online: 18 Aug 2006.

To cite this article: Avijit Mazumder , Arun Mahato & Rupa Mazumder (2006) Antimicrobialpotentiality of Phyllanthus amarus against drug resistant pathogens, Natural Product Research:Formerly Natural Product Letters, 20:04, 323-326, DOI: 10.1080/14786410600650404

To link to this article: http://dx.doi.org/10.1080/14786410600650404

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Natural Product Research, Vol. 20, No. 4, April 2006, 323–326

Antimicrobial potentiality of Phyllanthus amarusagainst drug resistant pathogens

AVIJIT MAZUMDER*, ARUN MAHATO and RUPA MAZUMDER

Department of Pharmaceutical Sciences, Birla Institute of Technology,Mesra, Ranchi – 835 215, India

(Received 29 September 2005; in final form 3 March 2006)

The antimicrobial potentiality of the methanolic extract of Phyllanthus amarus (Family:Euphorbiaceae) was studied against some drug resistant pathogenic bacterial strains by discdiffusion and agar dilution method. The extract showed significant concentration-dependentantibacterial activity particularly against gram-negative microbes. The study illustratedthe claim of the usefulness of the plant in dysenteric and diarrheal infections and alsosuggested its use in fever. The antibacterial action was mainly due to the isolated phyllanthin.

Keywords: Phyllanthus amarus; Euphorbiaceae; Antimicrobial

1. Introduction

Phyllanthus amarus is an erect annual herb cultivated throughout India. It has beenfound that the local tribes of Chotonagpur region of India use the leaves of this plantas an antitussive and an astringent. The plant has been used by the local tribesof the Chotanagpur region in India for treatment of jaundice, sores, fever, dysentery,and diarrheal infections [1]. The plant was reported to have hepatoprotective activity[2]. Sane et al. [3] in 1997 reported the efficacy of the plant in inhibiting the hepatitis Bvirus. The leaves were found to possess antipyretic activity [4]. The present investigationwas undertaken to evaluate the antibacterial efficacy of P. amarus on some drugresistant microbes and thereby justify the folklore claim of the plant in the traditionalsystem of Indian medicine.

2. Materials and methods

2.1. Plant material and extraction

Phyllanthus amarus (Euphorbiaceae) were collected in April 2004 from the Ranchidistrict of Jharkhand, India. The plant was then identified by Scientist-in-charge,

*Corresponding author. Tel.: þ91-651-2275339. Fax: þ91-651-2275401. Email: avijitmazum@yahoo.com

Natural Product Research

ISSN 1478-6419 print/ISSN 1029-2349 online � 2006 Taylor & Francis

http://www.tandf.co.uk/journals

DOI: 10.1080/14786410600650404

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Central National Herbarium, Botanical Survey of India, Shibpur, Howrah (CNH/I-I(89)/2004-Tech II/1323) and a voucher specimen is also retained by our laboratoryfor future reference. The whole plant was then air-dried after washing and pulverizedin a mechanical grinder. The dried powder was extracted with methanol in a Soxhletapparatus (yield 2.4% w/w with respect to dried powdered extract).

2.2. Test microorganisms

The overnight nutrient broth culture of Shigella dysenteriae 1, S. dysenteriae 2,S. boydii 8, Staphylococcus aureus ML267, S. aureus NCTC 7447, Streptococcuspneumoniae 7465, Escherichia coli ROW 7/12, E. coli CD/99/1, Bacillus subtilisCD/99/1, Salmonella typhimurium ATCC 6539, Vibrio cholerae 8531, Pseudomonasaeruginosa 7241, and Klebsiella pneumoniae RM 8/98 were used in the present study.All these strains were found to show a high degree of resistance when tested against thestandard antibacterial including ciprofloxacin.

2.3. Determination of antimicrobial activity

One loopful (loop diameter: 3mm) of an overnight grown nutrient broth culture of thetest organisms was added by checker board technique to the marked quadrant ofthe sterile 100mm petridishes containing various concentrations of the extractin nutrient agar (5, 10, 25, 50, 100, 200, 400, 800, and 1000 mgmL�1) (table 1). Thespot inoculated plates were incubated at 37�C for 24 h to determine MIC of the extractagainst the microorganisms [5].

Antibacterial activity was then also tested by the disc diffusion method. The sterilenutrient agar cooled to 48–50�C seeded with the test organisms was poured inpresterilized petridishes. When the agar solidified, the total surface of the agar wasdivided into six quadrants and six filter paper discs (diameter: 6mm) containingdifferent concentrations of the extract, standard drug (sparfloxacin), and one control

Table 1. Determination of MIC of extract against various microorganisms.

Inhibitory effect at concentrations of extract in mgmL�1

Name of organisms 0* 5 10 25 50 100 200 400 800 1000

Bacillus subtilis CD/99/1 þ þ þ þ þ þ þ þ � �

Staphylococcus aureus ML267 þ þ þ þ � � � � � �

Staphylococcus aureus NCTC 7447 þ þ þ þ þ � � � � �

Shigella dysenteriae 1 þ þ þ � � � � � � �

Shigella dysenteriae 2 þ þ þ � � � � � � �

Shigella boydii 8 þ þ þ � � � � � � �

Escherichia coli ROW 7/12 þ þ þ � � � � � � �

Escherichia coli CD/99/1 þ þ þ þ � � � � � �

Vibrio cholerae 8531 þ þ þ þ þ � � � � �

Pseudomonas aeruginosa 7241 þ þ þ þ þ þ þ � � �

Salmonella typhimurium ATCC 6539 þ þ þ þ þ þ þ � � �

Klebsiella pneumoniae RM8/98 þ þ þ þ þ þ þ � � �

Streptococcus pneumoniae 7465 þ þ þ þ þ þ þ � � �

(0*)Control without any extract; þ)Growth; �)No growth; �) Inhibited growth).

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(10% dimethyl sulfoxide) were placed in the middle of each zone. All the plates wereincubated at 37�C for 48 h. The zone of inhibition was calculated by measuring thediameter of zone of no bacterial growth around filter paper disc. For each set of data,an average of three independent determinations was recorded [6,7].

2.4. Phytochemical studies

The extract was tested by preliminary phytochemical screening for the detection of thephytoconstituents [8,9].

3. Results and discussion

The preliminary phytochemical studies revealed the presence of alkaloids, tannins,and flavanoids in the methanolic extract. Shigella spp. were found to be inhibited at theleast concentration (25mgmL�1) and found to show the maximum diameter of zoneof inhibition at the largest tested concentration of 1000 mgmL�1 comparable withsparfloxacin. E. coli was the next highly sensitive culture as it is inhibited at a slightlyhigher concentration of the extract (50 mgmL�1) and showed a zone of inhibitionslightly smaller than that produced by the strains of Shigella spp. V. cholerae was thenext susceptible strain as the extract had a MIC of 100 mgmL�1 with a greater zone ofinhibition as compared to that produced by the next susceptible strains of S. aureusthough having the same MIC as the former. Similar sort of results were obtained withall the remaining microbes. Thus it was seen that methanol extract posses significantantimicrobial activity against all the tested strains; maximum inhibitory effect wasnoted against Shigella spp., E. coli, V. cholerae, and S. aureus. The extract was foundto be moderately active against S. typhimurium, P. aeruginosa, B. subtilis, and otherpneumonia causing strains of Klebsiella and Streptococcus spp. The study thus justifiedthe use of the plant in dysenteric and diarrheal infections by the tribals. The activity

Table 2. Antibacterial activity of methanol extract of P. amarus.

Diameter of zone of inhibition (mm)a

Concentration of extract (mgmL�1)Sparfloxacin10 mgmL�1Bacteria 200 400 800 1000

Bacillus subtilis CD/99/1 6.0 6.5 7.5 8.0 18.5Staphylococcus aureus ML267 7.0 8.0 9.0 10.5 24.0Staphylococcus aureus NCTC 7447 7.5 9.0 10.0 11.0 24.5Shigella dysenteriae 1 10.5 11.5 13.5 17.0 22.5Shigella dysenteriae 2 10.0 11.5 13.5 16.5 23.0Shigella boydii 8 11.0 13.0 14.5 18.0 23.5Escherichia coli ROW 7/12 10.0 12.0 14.0 16.0 28.0Escherichia coli CD/99/1 10.0 12.0 14.0 16.0 27.5Vibrio cholerae 8531 8.5 11.0 12.5 15.0 29.0Pseudomonas aeruginosa 7241 7.0 7.5 8.0 8.5 24.5Salmonella typhimurium ATCC 6539 7.0 8.0 8.5 9.0 24.0Klebsiella pneumoniae RM8/98 6.0 6.5 7.5 8.5 20.0Streptococcus pneumoniae 7465 6.0 7.0 8.0 9.0 20.5

a Each value is an average of three values. Solvent used is dimethyl sulfoxide.

Antimicrobial potentiality of P. amarus 325

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of the extract against S. aureus supported the use of the plant against fever (table 2).However the exact mechanism of action is yet to be determined.

References

[1] A.V. Sala. Indian Medicinal Plants: A Compendium of 500 Species, Vol. IV, pp. 252–253, OrientLongman, New Delhi, India (1996).

[2] D.W. Unander, B.S. Blumberg. Econ Bot., 45, 225 (1997).[3] R.T. Sane, J.L. Chawla, V.V. Kuber. Indian Drugs, 34, 11 (1997).[4] R. Mehrotra, S. Rewat, D.P. Kulshestra, G.K. Patnaik, B.N. Dhawan. Indian J. Med. Res., 93A, 71

(1991).[5] Rupa Mazumder, S.G. Dastidar, S.P. Basu, Avijit Mazumder, S.K. Singh. Phytotherapy Research, 18(10),

824 (2004).[6] Avijit Mazumder, B.P. Saha, S.P. Basu, Rupa Mazumder. Indian J. Nat. Prod., 19(3), 20 (2003).[7] M.J. Pelczar, E.C.S. Chan, N.R. Kreig. Microbiology Concepts and Application, Intl. Edn, pp. 578–587,

McGraw Hill, New York (1993).[8] J.B. Harborne. Phytochemical Methods, pp. 280–288, Chapman and Hall, New York (1976).[9] G.E. Trease, W.C. Evans. Pharmacognosy, 12th Edn, p. 539, ELBS Publication, London, UK (1985).

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