ANTIFUNGAL ACTIVITY OF Curcuma neilgherrensis Wt. A WILD MEDICINAL PLANT.

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1 Page3903 Indo American Journal of Pharmaceutical Research, 2013 ISSN NO: Journal home page: INDO AMERICAN JOURNAL OF PHARMACEUTICAL RESEARCH ANTIFUNGAL ACTIVITY OF Curcuma neilgherrensis Wt. A WILD MEDICINAL PLANT. D. Chaithra*, N.Yasodamma, C. Alekhya Department of Botany, Sri Venkateswara University, Tirupati, Andhra Pradesh, India ARTICLE INFO Article history Received 30/04/2013 Available online 31/05/2013 Keywords Curcuminoids, Cosmetics, Hepatoprotective, Antiarthritis, Skin Diseases, Aspergillus Niger, Candida Albicans. ABSTRACT Indian Curcuma is well known to cure skin infections and its traditional uses as facial cosmetic. Curcuminoids and volatile oils of Curcuma species are known for their therapeutic activities as antiasthamatic, antitumour, stomachic, hepatoprotective, antiarthritis, antiseptic and to cure menstrual disorders. C.neilgherrensis is a wild Curcuma of Eastern Ghats also possesses essential oils, phenols, flavonoids, tannins, saponins and alkaloids. To prove its therapeutic activities against antiasthamatic, antitumour, stomachic, hepatoprotective, antiarthritis, antiseptic, antifungal activity of C.neilgherrensis leaves and rhizome extracts was carried out. Both leaf and rhizome methanol and alcohol extracts are proved most effective on Candida albicans than Aspergillus niger. But antifungal activity of all extracts showed most effective than the reference drug Nystatin and also the MIC values are proved with lowest concentrations. Hence the bioactive constituents may be isolated and further the drug may be formulated to prove its therapeutic activities in support of its herbal uses. Corresponding author D. Chaithra Department of Botany, Sri Venkateswara University, Tirupati, Andhra Pradesh, India Please cite this article in press as D. Chaithra et.al. ANTIFUNGAL ACTIVITY OF Curcuma neilgherrensis Wt. A WILD MEDICINAL PLANT. Indo American Journal of Pharm Research.2013:3(05). Copy right 2013 This is an Open Access article distributed under the terms of the Indo American journal of Pharmaceutical Research, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

2 Page3904 INTRODUCTION: The genus Curcuma, comprises nearly 80 species, have been used in traditional systems of medicine (Ayurveda, Siddha, Unani) since a long time. Among them the least studied is C.neilgherrensis which is known to possess tremendous therapeutic potency has been attributed due to the presence of various secondary metabolites such as essential oils, phenols, flavonoids, tannins, saponins and alkaloids which possess anti-inflammatory, Cholagogue, hepatoprotective, blood purifier, antioxidant, antimicrobial, skin diseases, anti-asthmatic, antitumour, stomachic, carminative and regenerator of liver tissue. It is also used for chronic hepatitis, antiarthritis, antiseptic and menstrual disorders. [1-8]. C. neilgherrensis produce starchy rhizomes which are used as remedy for infections, antimicrobial, inflammations, gastric and skin disorders by the local herbalists but have not been evaluated pharmacognostically. The present study aims to investigate antifungal activity of C. neilgherrensis leaf and rhizome extracts. MATERIALS AND METHODS: Collection and identification of Plant material: Plant material C. neilgherrensis (Zingiberaceae) was collected from Tirumala and Talakona along the Seshachalam Hill Ranges during the months of April September, 2011, authenticated by Prof. N.Yasodamma and a voucher specimens DC 921, DC 922 were prepared and preserved in the herbarium Department of Botany, S.V.University, Tirupati as per the standard method [9]. Rhizomes and leaves were collected, thoroughly washed, cut in to pieces and further dried under shade at 28 ± 2 º C for about 10 days. The dried parts were ground well in to a fine powder in a mixer grinder and sieved to particle size of mm. The powders were stored in a polythene bags at room temperatures. Extracts preparation: Shade dried leaf and rhizome powders were subjected to soxhlet extractions with Methanol and Ethanol. Simultaneously cold water and hot water extracts also prepared. The above obtained semisolid extracts were preserved in airtight bottles at 4 ºC in the refrigerator until further use. Antifungal activity of leaf and rhizome extracts: Test organisms: Fungal cultures of Candida albicans (ATTC-10231) causes kidney, liver damages, convulsions, hemorrhages of lungs and brain and Aspergillus niger (ATCC-16404) causes infections of mucous membrane of the mouth (Thrush), vagina and alimentary tract, skin diseases physiological disorders and obesity. These organisms were procured from the department of Microbiology, S.V. University and were further maintained on nutrient agar slants at 4 0 C until further use. Preparation of Inoculum: Active cultures were prepared by transferring a loop full of cells from the stock cultures to test tubes of Potato Dextrose agar medium and were incubated without agitation for 24hrs at 37 C. Well diffusion method: Antifungal activity of plant extracts was determined using agar well diffusion method with slight modifications of Perez [10]. Potato dextrose agar was inoculated by spreading the selected fungal inoculums on the media. Wells (9 mm diameter) were punched in the agar and filled with plant crude extracts of 10mg/well; control wells containing pure solvents (negative control) or standard antibiotic (positive control) Nystatin (10mg/well). The plates were incubated at 25+2ºC for 48 hours for fungal growth. The antifungal activity was assessed by measuring the diameter of the zone of inhibition for the respective drug. The data of crude drugs activity is given the mean of quadruplicates along with the standard error. MIC of leaf and rhizome extracts compare with Nystatin 10mg: Minimum Inhibitory Concentration is defined as the lowest concentration where no visible turbidity is observed in the test tube (fungal concentration).the Vollekova method modified by Usman was employed [11, 12]. In this method the broth dilution technique was used where plant extracts were prepared to the highest concentration of 10mg/ml (stock concentration). By adding sterile distilled water and serially diluted (two fold dilution) using the potato dextrose broth and later inoculated with 0.2ml standardized suspension of the test organisms. After 18hrs of incubation at 37 C., the test tubes were observed for turbidity.the lowest concentration of the tube that did not show any visible growth can be considered as the MIC. RESULTS: Antifungal Activity of leaf and rhizome extracts: [Table-I, Graph-I, Plate-I] Both leaf and rhizome extracts showed most effective nearly double the activity than the reference drug Nystatin with 10.2 to 12.1mm on both the tested fungal strains respectively. A. niger mm and C.albicans mm of zone of inhibition. And it is also observed C.albicans is most susceptible than A.niger with methanol and alcohol extracts followed by hot water and cold water extracts.

3 Page3905 Zone of Inhibition in mm Table: I: Antifungal Activity of Leaf and Rhizome extracts (10 mg/well) Organism Leaf Rhizome NYS C.W H. W Al ME C.W H. W AL ME A.niger 19.2±0 21.3± 28.1± 27.2± 23.6± 25.5± 28.9± 29.5± 10.2± C.albicans 20.3±0 24.0± 27.7± ± 26.5± ± Values are mean Inhibition zone (mm) ± S.D of quadruplicate. C. W =Cold Water; H.W =Hot Water; AL = Alcohol; ME =Methanol; NYS: Nystatin ± 0.36 Graph: I: Antifungal Activity of Leaf and Rhizome extracts Antifungal Activity ± ± C.W H. W Al Me C.W H. W Al Me A.niger C.albicans Leaf Rhizome NYS Parts/ Extracts C. W =Cold Water; H.W =Hot Water; AL = Alcohol; Me =Methanol; NYS: Nystatin. Plate: I: Antifungal Activity Aspergillus niger LEAF Candida albicans

4 Page3906 Aspergillus niger RHIZOME Candida albicans 1, 2, 3, 4: Quadruplicates for 10mg/well. C. W =Cold Water; H.W =Hot Water; AL = Alcohol; ME =Methanol. CONTROL Aspergillus niger Candida albicans NYS: Nystatin. MIC of leaf and rhizome extracts compare with Nystatin 10mg: [Table: II, Graph: II] Minimum Inhibitory Concentrations on C.albicans ranges from to 2.5mg and on A.niger ranges from to 2.5mg. The lowest concentrations observed with methanol and alcohol extracts of both leaf and rhizome extracts at 0.156mg on C.albicans. Table: II: MIC values of Leaf and Rhizome extracts compare with Nystatin 10mg: Organism Leaf Rhizome C.W H. W Al Me C.W H. W Al Me A.niger C.albicans

5 Page3907 Conc. in mg Graph: II: MIC of leaf and rhizome extracts compare with Nystatin 10mg MIC- Fungal Activity C.W H. W Al Me C.W H. W Al Me A.niger C.albicans Leaf Rhizome NYS Parts/ Extracts C. W =Cold Water; H.W =Hot Water; AL = Alcohol; Me =Methanol; NYS: Nystatin. DISCUSSION: Antibacterial activity of Xanthorrhizol isolated from methanol extracts of C. xanthorrhiza rhizome against oral micro organisms resulted highest activity on four Streptococcus species which causes dental carries and also against Actinomyces viscosus and Porphyromonas gingivalis responsible for peridontitis. Whereas C.albicans and Lacto bacillus were somewhat resistant to Xanthorrhizol ranging MBC 4 to 8 µg/ml and MIC 2-4 µg to that of Chlorhexidine 2-4 µg/ml MBC and 1-4 µg MIC [13]. C. zedoaria and C. malabarica rhizome hexane and acetone extracts against six bacterial and two fungal strains exhibited effective activity than other extracts and C.malabarica is most efficient than C.zedoaria with MIC values ranging from 0.01 to 0.15 mg and 0.01 to mg with C.malabarica extracts, S.aureus is most susceptible. [14] C.longa volatile oils and Curcuminoids hydro methanol extracts were tested against B. subtilis, K. pneumonia, E.coli, Enterobacter aerogenes, Pseudomonas aeruginosa, S. aureus and Proteus mirabilis bacterial strains and on two fungal strains. A.niger and C.albicans effective activity was observed with Curcuminoid extracts than volatile oils on both bacteria and fungi, than the standard drugs Kanamycin and Flucanazole respectively [15]. Aqueous and Hydro alcoholic extracts of C.longa on S.aureus and C.albicans is very effective. [16]. Essential oils isolated from C. xanthorriza as Xanthorhizol, Camphere, Curcumin, α- pinane, β- pinene, Myrecene, Linalool antimicrobial activity against bacterial and fungal resulted most effective on E.coli, B.amyloliquefaciens, K.pneumoniae, P. aeruginosa followed by others. On fungal strains relatively less activity is observed. [17]. CONCLUSION: Antifungal activity on both the strains with leaf and rhizome aqueous, methanol and alcohol extracts proved most effective to that of the control drug Nystatin and the MIC values ranges very low in relation to that of the herbal uses. Antifungal activity with isolated compounds of C.longa and C. xanthorrhiza, C.malabarica is also equal to that of C.neilgherrensis crude extracts inhibition on fungal strains is most promising and very recommendable for future drug designing studies. ACKNOWLEDGEMENTS The authors are grateful to the University Grants Commission (UGC) New Delhi for Financial assistance. We are also indebted to the Department of Botany, S.V.U College of Sciences, Sri Venkateswara University, Tirupati, Andhra Pradesh, India for providing the space and facilities to complete the above Research work. The authors are grateful in this regard. REFERENCES: 1. Rangachari, D. Flora of Chittoor District, Ph.D. Thesis, S.V.University, Tirupati, India, Pullaiah. T, Flora of Andhra Pradesh. Scientific publishers, New Pali Roads, Jodhpur, India. 5A, Jadav S. N., Conservation assessment and management planning (CAMP) for medicinal plants of Andhra Pradesh. Medicinal Plant Conservation Center (MPCC) and FRLHT, Hyderabad, 2001: Yesodaram. K and K.A. Sujana. Wild Edible Plants Traditionally used by the Tribes in the Parambikulam Wildlife Sanctuary, Kerala, India. Natural Product Radiance, 2007, 6(1); Arinathan. V, Mohan.VR, John De Britto and Murugan. C. Wild edibles used by Palliyars of the Western Ghats, Tamilnadu, Indian Journal of Traditional Knowledge, 2007, 6 (1) : Gantait A, Barman T and Mukherjee P. Validated method for Determination of Curcumin in Turmeric Powder. Indian Journal of Traditional Knowledge, 2011, 10 (2):

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