The antimicrobial activity of ethanol extract obtained from Rauvolfia tetraphylla was tested against bacterial species of Escherichia coli, Streptococcus lactis, Enterobacter aerogenes, Alcaligenes faecalis, Pseudomonas aeruginosa, Proteus vulgaris, and fungal species of Fusarium oxysporum, Alternaria helianthii, Curvularia lunata, Aspergillus niger, and Penicillium spp.  Better antimicrobial activity was observed when the extracts showed maximum activity against E. coli, Enterobacter aerogenes, and Alcaligenes faecalis. Among different fungi tested, A. niger and Penicillium spp were found to be more sensitive to crude extract when compared to others

Babu, S. Saravana

Harisaranraj, R.

Suresh, K.

English

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Ethnobotanical Leaflets 12: 586-90. 2008.  Studies on In Vitro Antimicrobial Activity of Ethanol Extract of Rauvolfiatetraphylla Suresh K*, S. Saravana Babu and Harisaranraj R. Department of Plant Biology and Plant BiotechnologyChikkaiah Naicker College, Erode. (T.N.) INDIA* sureshbioteck@yahoo.com Issued 11 August 2008  ABSTRACT The antimicrobial activity of ethanol extract obtained from Rauvolfia tetraphylla was tested against bacterialspecies of Escherichia coli ATCC 69314, Streptococcus lactis NCIM 50038, Enterobacter aerogenes NCIM 2340,Alcaligenes faecalis ATCC 15246, Pseudomonas aeruginosa NCIM 2200, Proteus vulgaris ATCC 6380 and fungalspecies of Fusarium oxysporum NCIM 1008, Alternaria helianthii ATCC 201540, Curvularia lunata ATCC 34477,Aspergillus niger NCIM 1207 and Penicillium spp NCIM 741. Better antimicrobial activity was observed when theextracts showed maximum activity against E. coli, Enterobacter aerogenes, Alcaligenes faecalis. Among differentfungi tested A. niger and Penicillium spp were found to be more sensitive to crude extract when compared to others. Key Words: Antibacterial, Antifungal, Ethanol extract, Rauvolfia tetraphylla. Introduction: Medicinal plants as a group comprise approximately 8000 species and account for around 50% of all thehigher flowering plant species of India. Millions of rural households use medicinal plants in a self-help mode. Overone and a half million practitioners of the Indian System of Medicine in the oral and Codified streams use medicinalplants in preventive, promotive and curative applications. There are estimated to be over 7800 manufacturing unitsin India. In recent years, the growing demand for herbal product has led to a quantum jump in volume of plantmaterials traded within and across the countries. In recent years, secondary plant metabolities (Phytochemicals),previously with unknown pharmacological activities, have been extensively investigated as a source of medicinalagents (Krishnaraju A V, et al, 2005). Thus it is anticipated that phytochemicals with adequate antibacterial efficacywill be used for the treatment of the bacterial infections (Balandrin M F, et al 1985). Approximately 20% of the plants found in the world have been submitted to pharmacological or biologicaltests (Suffredini et al., 2004). The systemic screening of antimicrobial plant extracts represents a continuous effort tofind new compounds with the potential to act against multi-resistant pathogenic bacteria and fungi. A special featureof higher angiospermic plants is their capacity to produce a large number of organic chemicals of high structuraldiversity. The so-called secondary metabolites (Evans et al., 1986), which are divided into different categories basedon their mechanism of function like chemotherapeutic, bacteriostatic, bactericidal and antimicrobial (Purohit andMathur, 1999). The accumulation of phytochemicals in the plant cell cultures had been studied for more than thirtyyears and the generated knowledge had helped in realization of using cell cultures for production of desiredphytochemicals (Castello et al., 2002). The antimicrobial activity of Solanaceae and Apocynaceae members were well documented in the literature.These include Cestrum diurnum (Bhattacharjee et al., 2005), Capsicum annum (Cichewicz and Thorpe, 1996),Withania spp. (Ramzi et al., 2005), Picralima nitida (Nkere and Lroegbu, 2005), Nerium oleander (Hussain andGorsi, 2004), Alstonia macrophylla, Alstonia cholaris, Voacanga foetida, Wrightia spp. (Hadi and Bremner, 2001),Rauvolfia serptentina (Siddique et al., 2004). The leaf extract of the herbaceous plant, Rauvolfia tetraphylla (Apocynaceae), is used for treatment ofcholera, eye disease and fever. It is also used as antihypertensive, as well as in intestinal disorders, diarrhea anddysentery (Anonymous, 1969). The principle aim of the present work was to study the antimicrobial activity ofRauvolfia tetraphylla, a photochemical study based on the in-vitro screening of ethanol crude extract was done. Plant material The fresh matured leaves of the R. tetraphylla were collected randomly during the month of January-February, from the Kolli Hills, Namakkal (District), Tamilnadu, India. The plant species were identified by referringthe standard morphological characteristic features (keys) according to the Flora of Madras Presidency. Preparation of Extracts The dried Leaves were powdered and soaked in the ethanol for about 10-15 days then this cold extract issubjected to distillation at low temperature under reduced pressure in rotary flash evaporator and concentrated onwater bath to get the crude extract. Likewise, the powdered leaf which is subjected to soxhlation is exhaustivelyextracted with ethanol for 48 hours. The solvent was distilled off at lower temperature under reduced pressure inrotary flash evaporator and concentrated on water bath to get the crude extract. Disc diffusion method The bioassay for bacterial strains was employed by disc diffusion method (Ergene et,al 2006). Filter paperdiscs (Whatman No. 1) of 5 mm diameter were loaded with crude extracts. Discs were completely dried andsterilized. 100 µl of cultures were spread on sterilized nutrient agar media; impregnated discs were placed on it andincubated for 24 hrs at 37ºC. Streptomycin discs (10 µg/disc) were used as a standard drug. The diameter of zone ofinhibition in mm was recorded after incubation. The experiment was performed in triplicates and average diameter ofzone of inhibition was obtained. Screening of antifungal activity The antifungal activity was determined by the poison food technique. The test fungus is allowed to grow onpoisoned plate with ethanol extract. It was observed that reduction in colony diameter and extent of sporulation. Theeffect of sample on the fungal growth was determined by measuring the diameter of the colony obtained on poisonedplate. RESULTS AND DISCUSSION The disc diffusion method for antibacterial activity showed significant reduction in bacterial growth in termsof zone of inhibition around the disc. Among bacterial forms tested, E. coli, Enterobacter aerogenes and Alcaligenesfaecalis were found to be more sensitive to crude extract. Other bacterial forms were inhibited by the extract. Thezone of inhibition increased on increasing the concentration of extract in disc. This showed the concentrationdependent activity (Table 1). The results of poison food technique revealed antifungal nature of the constituentspresent in the crude leaf extract of Rauvolfia tetraphylla. Among different fungi tested A. niger and Penicillium sppwere found to be more sensitive to crude extract when compared to others. A considerable reduction in thesporulation was also recorded. The antibacterial activity crude extract is shown in Table 1. The extracts showed maximum activity against E.coli, Enterobacter aerogenes and Alcaligenes faecalis. These data revealed that extracts of R. tetraphylla exhibitedsignificant antibacterial activity. In testing, inhibition zone increased with increase in drug concentrations and thusexhibiting concentration dependent activity.The plants are the vital source of innumerable number of antimicrobialcompounds. Several phytoconstituents like flavanoids (Tsuchiya et al., 1996), phenolics and polyphenols (Mason andWasserman, 1987), tannins (Ya et al., 1988), terpenoids (Scortichini and Pia Rossi, 1991), sesquiterpenes (Goren,1996) etc., are effective antimicrobial substances against a wide range of microorganisms. Table 1.  Antibacterial activity of Rauvolfia tetraphylla. Sl. No. Organism Plant extract used Average Inhibition zone in mm 1 Escherichia coli  Ethanolextractfromseeds ofRauvolfiatetraphylla 26.8 13.3 10.62 Streptococcuslactis24.3 12.6 10.93 Enterobacteraerogenes29.6 14.9 11.54 Alcaligenesfaecalis21.2 13.3 10.45 Pseudomonasaeruginosa20.6 11.4 9.66 Pseudomonasaeruginosa25.6 22.5 18.2 Table 2.  Antifungal activity of Rauvolfia tetraphylla. Sl.No. Test organism Average colonydiameter in mm incontrol platesAverage colonydiameter in mmin plates with  extract 1 Fusarium oxysporum 33 262 Alternaria helianthii 36 313 Curvularia lunata 39 284 Aspergillus niger 24 125 Penicillium spp 29 14  CONCLUSION The antimicrobial activity of R. tetraphylla may be attributed to the various phytochemical constituentspresent in the crude extract. The purified components may have even more potency with respect to inhibition ofmicrobes. The work carried was a basic approach to find out the antimicrobial activity in R. tetraphylla. Furtherworks on the types of phytoconstituents and purification of individual groups of bioactive components can reveal theexact potential of the plant to inhibit several pathogenic microbes. ACKNOWLEDGMENTAuthors are gratefully thankful to Prof. K. Sekar, Lecturer (SG), A.A Government Arts College, Namakkal,Tamilnadu, India for constant encouragement and timely help during the present work. We are also grateful to theDirectors, BIIOGENEIC – Institute of Biotechnology for providing laboratory facilities.  REFERENCES 1. Anonymous (1969). The Wealth of India. Publications and information directorate, Vol.8. CSIR, New Delhi. 2. Balandrin M F , Kjocke A J, Wurtele et al, Natural Plant chemicals sources of Industrial and mechanicalmaterials, Science 228-1154-160. 1985. 3. Bhattacharjee I, Ghosh A, Chandra G (2005). Antimicrobial activity of essential oil of Cestrum diurnum (L.)(Solanales: Solanaceae). Afr. J. Biotechol. 4: 371-374. 4. Castello M, Phatak A, Chandra N, Sharon M (2002). Antimicrobial activity of crude extracts from plant partsand  corresponding calli of FBixa orellana L. Indian J. Exp. Biol. 40: 1378-1381. 5. Cichewicz RH, Thorpe PA, (1996). The antimicrobial properties Chilli peppers (Capsicum species) and theirused in Mayan medicine. J. Ethnopharmacol. 52: 61-70. 6. Ergene, A., Guler, P., Tan, S., Mirici, S., Hamzaoglu, E. and Duran, AAntimicrobial and antifungal activity ofHeracleum sphondylium subsp.artivinense. Afr.J. Biotechnology, Vol 5(11), 1087-1089. 2006. 7. Evans JS, Pattison E, Morris P, (1986). Antimicrobial agents from plant cell culture, in: secondary metabolitesin plant cell culture. Morris P, Scraggs A, Stanfford A, Fowler M. Cambridge University, London. p.12. 8. Goren, N., Woerdenbag, H. and Bozok-Johansson, C. Cytotoxic and antibacterial activities of sesquiterpenelactones isolated from Tanacetum praeteritum subsp. praeteritum. Planta Medica. 62: 419-422. 1996. 9. Hadi H, Bremner B, (2001). Initial studies on alkaloids form Lombok medicinal plants. Molecules 6: 117-129. 10. Hussain MA, Gorsi MS (2004). Antimicrobial activity of Nerium oleander Linn. Asi. J. Plant Sci. 3: 177-180. 11. Krisharaju, A, V., Rao T V N, and Sundararaju. 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Siddique NA, Bari MA, Naderuzzaman ATM, Khatun N, Rahman MH, Sultana RS, Matin MN, Shahnewaz S,Rahman MM (2004). Collection of indigenous knowledge and identification of endangered medicinal plantsby questionnaire survey in Barind tract of Bangladesh. J. Biol. Sci. 4(1): 72-80. 18. Suffredini JB, Sader HS, Goncalves AG, Reis AO, Gales AC, Varella AD, Younes RN (2004). Screening ofantimicrobial extracts from plants native to the Brazilian Amazon rainforest and Atlantic forest. Brazil. J.Med. Biol. Res. 37: 379-384. 19. Tsuchiya, H., Sato, M., Miyazaki, T., Fujiwara, S., Tanigaki, S., Ohyama, M., Tanaka, T. and Iinuma, M.Comparative study on the antibacterial activity of phytochemical flavanones against methicillinresistantStaphylococcus aureus. Journal of Ethnopharmacology. 50: 27-34. 1996. 20. Ya, C., Gaffney, S.H., Lilley, T.H. and Haslam, E. Carbohydrate-polyphenol complexation. p. 553. In:Hemingway, R.W. and Karchesy, J.J. (ed.), Chemistry and significance of condensed tannins. Plenum Press;New York. 1988.  

Studies on \u3cem\u3eIn Vitro\u3c/em\u3e Antimicrobial Activity of Ethanol Extract of \u3cem\u3eRauvolfia tetraphylla\u3c/em\u3e

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Studies on \u3cem\u3eIn Vitro\u3c/em\u3e Antimicrobial Activity of Ethanol Extract of \u3cem\u3eRauvolfia tetraphylla\u3c/em\u3e

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