Asian Pacific Journal of Tropical Medicine (2011)654-657 654 Contents lists available at ScienceDirect Asian Pacific Journal of Tropical Medicine journal homepage:www.elsevier.com/locate/apjtm Document heading doi: Antifungal activity of Ocimum sanctum Linn. (Lamiaceae) on clinically isolated dermatophytic fungi Balakumar S1, Rajan S2, Thirunalasundari T3, Jeeva S4* 1Department of Chemistry and Biosciences, SASTRA University, Srinivasa Ramanujan Centre, Kumbakonam- 612001, Tamil Nadu, India 2Department of Microbiology, M.R. Government Arts College, Mannargudi, Thriuvarur District, Tamil Nadu, India 3Department of Biotechnology, Bharathidasan University, Tiruchirappalli-620 024, Tamil Nadu, India 4Centre for Biodiversity and Biotechnology, Department of Botany, Nesamony Memorial Christian College, Marthandam-629 165, Tamil Nadu, India ARTICLE INFO ABSTRACT Article history: Objective: Ocimum sanctum To assess antifungal activity of leaves against dermatophytic fungi. Received 21 February 2011 Methods: Ocimum sanctum Antifungal activity of leaves was measured by 38 A NCCLS method. Received in revised form 20 April 2011 (MIC) (MFC) Accepted 15 July 2011 Minimum inhibitory concentrationOcimum sanctum and minimum fungicidal concentrationResults: ofOcimum various extracts and fractions of leaves were also determined. Available online 20 August 2011 sanctum leaves possessed antifungal activity against clinically isolated dermatophytes at the concentration of 200 g/mL. MIC and MFC were high with water fraction (200 g/mL) against Keywords: 毺 Conclusions: Ocimum sanctum 毺 dermatophytic fungi used. has antifungal activity, and the leaf Antifungal activity extracts may be a useful source for dermatophytic infections. EpidermophytonDermatophytosis floccosum MicrosporumMinimum fungicidal concentration sp OcimumMinimum sanctum inhibitory concentration Trichophyton sp 1. Introduction (WHO) estimates that about three quarters of the world population currently use herbs and other forms of traditional system of medicines for treating their diseases. Fungal infections are one of the major health problems Herbal formulations are popular among rural and urban in tropical countries, and tinea is one of the most common community in India[9-12]. Part of the reason is their fungal infection. Climate of the region, living conditions popularity, acceptability and the belief that all natural like nutrition, hygiene and socio-economic status, products are safe[13-17]. Recent studies proved that the genetic susceptibilities and other such conditions play plants used in traditional medicine contain a wide range an important role in the prevalence of dermatophytic of bioactive compoundsOcimum thatsanctum can be used to treat infectious infection in a particular region. Dermatophytes invade diseases[18-22]. is a medicinal shrub and in to the keratinophilic region of the body and cause is commonly called as Tulsi. Traditionally the leaves of dermatophytosis. The drugs used against dermatophytosis this plant were used as a medicine for asthma and other [9-12] exhibit several side effects and have limited efficacy. respiratory problems .Ocimum No report sanctum is available on the Hence there is a distinct need for the discovery of new, antifungal activity of the . Biologically safer and effective antifungal drugs[1-4]. Herbal medicines active compounds and extracts isolated from such plant are popular as remedies for diseases by vast majority of species were used in herbal medicine and they have been population in the world[5-8]. The world health organization the centre of interest as on date. This work was carried out to investigate the antifungal Ocimum sanctum *Corresponding author: Dr. S. Jeeva, Assistant Professor, Centre for Biodiversity activity of the leaves of against the and Biotechnology, Department of Botany, Nesamony Memorial Christian College, clinical isolates of dermatophytic fungi from the patients Marthandam , Tamil Nadu, India -629 165. Tel: +91 9952202112 attending the dermatology section of Bharath Heavy E-mail: [email protected] Electrical Limited (BHEL) Hospital, Trichirappalli, India Balakumar S et al./Asian Pacific Journal of Tropical Medicine (2011)654-657 655 and Annal Gandhi Memorial Government Hospital, growth inhibition needed at least (291.66依28.86) 毺g/mL Tiruchirappalli, India. of extracts and fractions. Among extracts alcoholic extract showedTrichophyton best activity. mentagrophytes MIC required to inhibit the growth of 2. Materials and methods was (166Trichophyton.66依14.43) rubrum毺g/mL and (383.33依28.86) 毺g/mL againstMicrosporum gypseum, (366.66依38.11) 毺g/mL againstMicrosporum nanum , The plant material used in this study was collected from (383.66依14.43) 毺g/mL against Epidermophyton floccosum, and Tiruchirappalli, Tamil Nadu, India. It was identified and (366.66依57.73) 毺g/mL against authenticated by the Botanist of Department of Plant (Table 1). Among fractions methanol fraction exhibited best Biotechnology, Bharathidasan University, Tiruchirappalli. activity againstTrichophyton all dermatophytic mentagrophytes fungus: (125.00依25.00) Fresh leaves were collected and shade dried. The dried 毺g/mL against Trichophyton rubrum , (350.00依66.14) leaves were ground to powder and stored in an Ociair mtightum 毺g/mL againstMicrosporum gypseum , (375.00依25.00) 毺g/ sanctumcontainer until further use. Known quantity of mL against Epidermophyton floccosum, and (383.33依14.43) 毺g/ leaf powder was subjected for cold extraction with mL against ). water and 100% ethyl alcohol and the aqueous extracts were The results of preliminary quantitative phytochemicalOcimum collected. The extracts were dried in vacuum desicator and screeningsanctum of aqueous and alcoholic extracts of were stored in a sterile Ocimum container sanctum for further use. revealed the presence of multiple polar and non- Known quantity of coarse powder was polar chemical constituents (Table 2). Steroids, terpenoids, also successively extracted with various solvents like flavonoids, phenolic compounds, lignin, proteins, hexane, benzene, chloroform, ethyl acetate, methanol carbohydrates were present in both extracts. Hexane fraction and water. Different fractions collected were filtered and showed the availability of steroid, terpenoids, benzene evaporated in vacuum. Coding was given to various extracts extract also indicated the availability of steroid. Alkaloid and fractions and was stored till use. The dried extracts was also available in benzene fraction along with steroid. and fractions were weighed and dissolvedin-vitro in 5% dimethyl Chloroform and ethylacetate fractions yielded alkaloid, sulfoxide (DMSO) and was used for testing of its terpenoids and flavonoids. Methanol and water fractions activity against dermatophytes. showed the presence of terpenoids, flavonoids, phenolic Five differentTrichophyton clinical mentagrophytes isolates of dermatophyticTrichophyton compounds. Though charbohydrates, reducing sugars, rubrumfungi likeMicrosporum canis Microsporum, gypseum proteins and amimoacids were available in all the extracts Epidermophyton, floccosum , and and fractions, they were not effectively involved in any of were taken for this study. the inhibitory activities of fungus as they are the primary The selected isolates were grown on sabouraud dextrose metabolites.Ocimum sanctum agar (SDA). Twenty one day old culture of dermatophytic showedOcimum the presence sanctum of proteins in all fungi was scraped with a sterile scalpel and macerated the fractions and extracts. showed seven with sterile distilled water. The suspension was adjusted to eleven peaks in various fractions in HPTLC. The Rf was spectrophotometrically to an absorbance of 0.600 at 450 nm. between 0.03 and 0.90. Tannins andOcimum steroids sanctum were the two By this way the fungal inoculum was prepared. For further major secondary metabolites seen in . study known quantity of this inoculum was used. Susceptibility testing was performed by the reference 4. Discussion broth micro dilution method[23,24]. Minimum fungicidal concentration (MFC) and minimum inhibitory concentration℃ (MIC) were determined after 21 days of incubation at 35 The incidence of dermatophytosis has been high since by 38 A NCCLS method. Medicinal plants are the good source 1980s. It is still a serious problem in today. This is because of macromolecules andOcimum secondary sanctum metabolites. To know the of the antifungal drug resistance of the pathogen and the side phytoconstituents of , the extracts were effect exhibited by the drugs used to cure fungal diseases. subjected to the analysis of macromolecules and secondary Hence, there is a great demand for an efficient and safer metabolites by using thin layer chromatography and high treatment. Medicinal herbs could be an alternate source performance thin layer chromatography methods. for the treatment of dermatophytosis. Herbal medicines are generally safer and free from side effects. Another important reason for this revival is that the effectiveness of many 3. Result [1] traditional medicines is now an accepted fact . Ocimum sanctum This study showed that the leaf extracts of MIC value is used to evaluate antimicrobial nature of plant has fungicidal activity. Few previous studies have extracts and minimum quantity of antimicrobial compound comprehensively investigated the activity of medicinal plant required to kill or arrest multiplication of all microorganisms leaves, bark extracts and oil against dermatophytes and [19] presentOcimum in the sanctum medium or body fluid.