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Antimicrobial, Cytotoxic and Antioxidant Activites of Arisaema Utile

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Antimicrobial, Cytotoxic and Antioxidant Activites of Arisaema Utile Sofi Mubashir et al. / Journal of Pharmacy Research 2012,5(3),1368-1370 Research Article Available online through ISSN: 0974-6943 http://jprsolutions.info Antimicrobial, Cytotoxic and Antioxidant activites of Arisaema utile Sofi Mubashir*, Wajaht A.Shah*µ * Department of Chemistry, University of Kashmir, Hazratbal Srinagar-190006, J&K-India. Received on:10-12-2011; Revised on: 15-01-2012; Accepted on:12-02-2012 ABSTRACT Most of the species from genus Araceae have a history of use in folk medicine for the treatment of various infectious diseases. Therefore the present study was carried out in order to evaluate the antimicrobial, cytotoxic and antioxidant potential of Arisaema utile. The methanolic extracts of the aforesaid plant was evaluated for its antimicrobial activities using Agar well diffusion method, where 100mg/ml of the extract showed between 14-24mm inhibitory zones on the test organisms. The methanolic extract of the same plant was screened for its possible cytotoxic activities through Sulpharhodamine-B (SRB) assay against a panel of human cancer cell lines including THP-1 (leukemia), A-549(lung), HCT-15(Colon), Cervix (Hela) and Prostrate (PC-3), which revealed increase in growth inhibition during 48 hour incubation. The aforementioned plant extract also showed prominent DPPH radical scavenging activity. Key words: Arisaema utile; Anti-Microbial; Cytotoxic; antioxidant. INTRODUCTION The World Health Organization estimates that 80% of the world’s inhabit- America some two species are known at present. The majority of Arisaema ants rely mainly on traditional medicines for their health care [1]. Many species grow in humus-rich, well-aerated soils in mountain meadows and medicinal plants have proved to successfully aid in various ailments leading slopes as well as open spots in lowland to high altitude woods, few species to mass screening for their therapeutic components. Today, the search for grow in pure loamy soils in sunny rock crevices. The objective of the natural compounds rich in antioxidant, anticancer and antimicrobial proper- present study was to evaluate antimicrobial, cytotoxic and antioxidant ac- ties is escalating due to their medicinal importance in controlling many tivities of Arisaema utile belonging to Araceae family related chronic disorders such as cancer and cardiovascular diseases. Anti- oxidants aid in the prevention by scavenging the excess free radicals in the 2. MATERIALS AND METHODS body. Cancer is currently a leading cause of death and growing evidence relates its occurrence to the oxidative damage to DNA, proteins and lipid in 2.1 Plant Material the body [2]. It has been estimated that approximately two-thirds of anti- The Arisaema utile plant material was collected from district pulwama cancer drugs approved worldwide up to 1994 were derived from plant of Jammu & Kashmir, India. The localities were the plant material was sources [3]. The rapid emergence of multiple drug resistant strains of patho- collected are usually situated between 2400-4600m. Voucher specimen of gens to current antimicrobial agents has generated an urgent intensive search Arisaema utile bearing specimen no 27911, was deposited at KASH her- for new antibiotics from medicinal plants. Many medicinal plants have barium in centre of plant taxonomy, University of Kashmir, Srinagar, J & K, been screened extensively for their antimicrobial potential worldwide [4- India. 6]. Antimicrobial compounds of plant origin may occur in stems,roots,leaves,bark,flowers and fruits of plants. Plant derived phytoal- 2.2 Extraction and Isolation exin [7], isothiocynates [8], allicins, anthocyanins [9] essential oils [10], Shade dried and powdered plant material was subjected to soxhlet tannins, and terpenoids [11-13] have demonstrated antimicrobial activites. extraction with organic solvents in increasing order of their Further, plant phenolic compounds have been found to possess potent polarity(Petroleum ether, chloroform and Methanol).The extracts thus antioxidant [14,15, 16-19], antimicrobial and anticancer activities [20,21]. obtained were concentrated by vacuum evaporation using rotary evapora- These compounds are bacteriocidal or bacteriostatic influencing lag time, tor. growth rate and maximum growth of microorganisms. A significant oppor- tunity exists to identify new, natural plant derived antimicrobial agents for 2.3 Antimicrobial Assay treatment of diseases or as food or cosmetic preservatives. The effect of plant extracts on microorganisms have been studied by a very large number of researchers in different parts of the world. The aim of the The genus Arisaema commonly known as “Cobra Lilies” or “Jack in the present study was to evaluate the antimicrobial potential of Arisaema utile Pulpit” is made up of more than 250 herbaceous species, which are distrib- against different bacterial strains. uted throughout temperate to tropical areas on all continents with the ex- ception of Australia, Europe and South America. Their main distributional 2.3.1 Microbial Strains and Culture Media range is located from the western Himalayas, southeast to China and Japan, Gram positive and gram negative bacterial strains were obtained from Mi- whereas in North America only two to three species exist and in Central crobial Type Culture Collection (MTCC), Institute of Microbial Technol- ogy (IMTECH) Chandigarh, India. The bacterial strains used were Pseudomonas aeruginosa MTCC 1688, Proteus vulgaris MTCC 426, *Corresponding author. Bacillus subtilis MTCC 441, Staphylococcus epidermidis MTCC 435, Sta- phylococcus aureus MTCC 96. Bacterial strains were grown on nutrient Dr. Syed Wajaht Amin Shah agar plates at 370C and maintained on nutrient agar slants at 250C. Department of Chemistry, University of Kashmir, Hazratbal 2.3.2 Agar Well Diffusion Method Srinagar-190006, J&K-India. The antibacterial susceptibility tests were carried out using the agar well diffusion assay. The bacterial cultures were developed for 24 hours and Journal of Pharmacy Research Vol.5 Issue 3.March 2012 1368-1370 Sofi Mubashir et al. / Journal of Pharmacy Research 2012,5(3),1368-1370 were later transferred into boiling tubes containing 20ml of liquid nutrient methanolic extract of Arisaema utile may help to discover new chemical agar. The contents of the tubes were transferred to petri plates. After 10 classes of antibiotic substances that could serve as selective agents for minutes of solidification of the agar, petri plates were punched in the form infectious disease, chemotherapy and control. Results of these studies of wells. Later these agar wells were filled with 20µl of the plant extract indicate that further searches and characterizations of Arisaema utile for (100mg/ml) dissolved in methanol. The incubation was carried out for 24 antimicrobial compounds are warranted. In addition, researches on syner- hours at 37 0C. After the incubation period, the antimicrobial activity was gistic combinations of extracts with broad spectrum or a high degree of evaluated by measuring the width of the zones of inhibition. Kenamycin inhibition against a particular micro organism would seem worthwhile. As (10µg per disc) was used as positive control and pure methanol was used as the search for new antimicrobial agents intensifies, these plant extracts may negative control. provide attractive alternate sources of molecules for consideration. 2.4 Cytotoxic Studies Table -1 Antimicrobial activity of Arisaema utile. 2.4.1 Human Cell Lines and Culture Zones of inhibition (in mm) The optimum density of seeded cell suspension were introduced to each Materials Methanolic Standard* Control** well of 96-well plates (Iwaki) and exposed to particular concentration of /microorganisms extract the plant extract. In the cultured RPMI-1640 medium, supplemented with P. aeruginosa 14 35 - known cytotoxic agent Paclitaxel and Mitomycin-C (Sigma-Aldrich, Fluka, P.vulgaris 23 35 - UK) as positive controls. The cells were incubated with sample for 48 S.aureus 19 35 - hours incubation. Fixed the cells in ice cold TCA for 1 hour at 40oC. The B.Subtilis 21 35 - S. epidermidis 24 35 - plates were washed with distilled water and allowed to dry in the air. Sulpharhodamine-B (SRB) solution (0.4%) was added to each well of dry *Standarol : Kanamycin; **Control : Methanol; (-) : No inhibition 96-well plates and allowed staining at room temperature for 30-minutes.The 3.2 Cytotoxic Activity unbound SRB solution was removed by washing the plates quickly with In order to understand the effects of Arisaema utile on human cancer cell 1% (v/v) acetic acid. The bound SRB dye was solubilised by adding 100µl lines, experiments were carried using cultured THP-1(leukemia), A-549(lung), of 10mM unbuffered Tris base (PH=10.5) to each well and shaken for 5- HCT-15(Colon), Cervix(Hela) and PC-3 (Prostrate) cell lines by SRB as- minutes on shaker platform. The plates were read in a 96-well plate reader say. Cell lines were exposed to concentration of 100µg for 48 hours, which at 540nm. reduced the viability of these cell lines. As shown in Table 2, the extract was 2.4.2 Cytotoxic Assay active mainly against Leukemia (THP-1) and lung (A-549) cancer lines. Sulpharhodamine-B assay was performed against five human cancer cell Table-2 In-vitro Cytotoxic activity of Arisaema utile. lines namely THP-1(Leukemia), A-549 (Lung), HCT-15 (Colon), PC-3 (Prostrate) and Hela (Cervix), which revealed increase in growth inhibition Tissue type Leukemia Lung Colon Cervix Prostrate during 48 hour incubation at the concentration of 100µg of the sample. Cell line type THP-1 A546 HCT15 Hela PC-3 DMSO control was set up separately to cancel out the cell death occurred Material Conc.(µg) by DMSO, which was used as a solvent for dissolving samples homoge- Arisaema utile 100 83 14 15 30 1 Paclitaxel 1x10-6 13 61 17 6 7 neously. The results depicted that the inhibition of different human cancer -6 cell lines of varying tissue origin with 100µg imparted cellular cytotoxic Mitomycin-C 1x10 23 43 21 4 67 effects on all the cell lines that were tested. 2.3.4 Antioxidant activity.
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