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PHYTOCHEMICAL SCREENING and ANTIMICROBIAL ACTIVITY of FLOWER EXTRACT of Euphorbia Milii

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PHYTOCHEMICAL SCREENING and ANTIMICROBIAL ACTIVITY of FLOWER EXTRACT of Euphorbia Milii BMR Phytomedicine www.advancejournals.org Open Access Scientific Publisher Research Article PHYTOCHEMICAL SCREENING AND ANTIMICROBIAL ACTIVITY OF FLOWER EXTRACT OF Euphorbia Milii Megha Vyas1, Binita Desai1 1Department of microbiology, Shree Ramakrishna Institute of computer education And Applied Sciences, Atwalines, Surat-395 001, India Correspondence should be addressed to Megha Vyas Received February 27, 2018; Accepted March 31, 2018; Published April 07, 2018; Copyright: © 2018 Megha Vyas et al. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. Cite This Article: Vyas, M., Desai, B.(2018). Phytochemical screening and antimicrobial activity of flower extract of Euphorbia milii. BMR Phytomedicine, 4(1). 1-6 ABSTRACT An ornamental plant Euphorbia milii (Euphorbiaceae) is plays a role in folk medicine. The Chinese use it as a cure for cancer, and some Brazilians believe that it can cure warts. Phytochemical studies showed the presence of cardiac glycosides, steroids/phytosterols, anthocyanin, terpenoids, flavonoids and tannins.The Antimicrobial activity studies of hexane, ethyl acetate,chloform,petroleum ether,and water extracts of flowers of Euphorbia milii were performed on gram positive organisms(Bacillus subtilis, Bacillus megaterium,Staphylococcus aureus, Entrococci ) and gram negative organisms (Escherichia coli and Proteus vulgaris, Pusodomonas aeruginosa) by using cup plate method. The hexane extracts in the concentration of 5μg/ml, have shown considerable inhibition zone on as compared to other extracts.Bacillus subtlis and Bacillus megaterium have not show inhibition zone from solvent extract which was compared with the inhibition zone produced by standard amikacin sulphate (1μg/ml.). KEYWORDS:.Euphorbia milii, Anti-microbial, Phytochemical. INTRODUCATION when dried and processed as powder, inhibit the growth of Aspergillus. [1 nfectious diseases caused by bacteria ,fungi, viruses and I Table 1: Taxonomical Classification. parasites are still risk to public health, due to evolvement of microbial resistance to synthetic drug.[4] About 80% people Kingdom Plantae use tradition medicines, which compounds obtained from Division Magnoliophyta medical plant. Plant contains some organic compounds Class Magnoliopsida which show some physiological effect on human body. Phytochemical are secondary metabolites with obscure Order Malpighiales function .Phytochemical derived from any part of the plant Family euphorbiaceae Genus Euphorbia The family euphorbiaceae consists of 2000 species (Davis et Species E.milii al.,1988).The genus euphorbia is the largest genus of Origin Madagascar medical plant . E.milii is commonly known as “crown of Flowering Yes 1 thrown”.[1] The part of plants that grow above ground that Fragrant Slightly used for make medicine. Fungi of the genus Aspergillus Growing case No extra care requires produce a toxic substance called aflatoxin, which Temperature 30-40⁰c contaminates crops (e.g., corn and peanuts) and causes Vernacular names Crown of thrown, Christ human diseases. Aflatoxin has even been implicated as a plant, Christ thorn contributing factor in liver cancer. Euphorbia milii flowers, PM 18|Volume 4|Issue 1|2018 BMR Phytomedicine Milin, an extract of Euphorbia milii latex, is a glycosylated Pseudomonas aeruginosa(MTCC NO:2297))bacteria were serine protease (an enzyme that breaks down protein and has used[9], which were obtained from microbiology laboratory a sugar attached to it). Because it is more stable than most of Ramakrishna institute. Tests for susceptibility were proteases, it will be useful to food processers and makers of determined using modified agar well diffusion method to detergents who have been using proteases in their test the antibacterial activity of the different solvent operations. [1,3] fractions. The nutrient agar was used as medium. All the cultures were taken in triplicates at incubation temperature Phytochemical studies of E. milii revealed the Presence of of 37°C for 24 to 72 hours. In a sterile Petri-dish, test Flavanoids, Terpenoids, Tannins.Flavanoids are yellow organism broth culture (0.1mL) was placed with sterile pigments,which occure in plant kingdom either in free state nutrient agar medium (20mL). Holes were bored in to the or as a glycosides or associated with tannins.These are medium and each fraction was added (0.2mL). Amikacisn known as anthoxanthins.[2]. sulphate was the standard antimicrobial agent at a concentration of 2 mg /ml. it was incubated at 37°C for 24h. MATERIALS AND METHODS The diameter of the zone of inhibition of microbial growth in the plate was measured in millimeter (mm).[2,3,9] Collection of plat material and identification Phytochemical screening Flower of Euphorbia milii was collected from Jahngirpara, Surat. Identification of plant material was done by The chemical tests were performed on the water, hexane, Department of Botany, P. T. Science College, chloform, ethyl acetate, petroleum ether extracts of E. milii Athwalines,Surat followed by the standard procedures for identification of the chemical constituents.[1,5,10] Plant material Test for alkaloids The collected flowers were washed with running water 2-3 times and with distilled water. The flowers were air dried Each fraction (0.2g) was warmed with 2% H₂SO₄ (2.0ml) under shade for 10-15 days. The flower were crushed to fine for two minutes. The reaction mixture was filtered and few powder with help of electric grinder and stored for further drops of Dragendrof’s reagent were added to filtrate. Orange analysis.[1,3] red precipitation showed the presence of alkaloids moiety. Aqueous extraction Test for tannins 20 grams of dried plant material was extracted in 100 ml water for 2 hour on rotary shaker. After 2 hour it was Each extract in small quantity was mixed with water and filtered through whattman filter paper-1 and centrifuged at heated on water bath and filtered. To the filtrate, few drops 5000 rpm for 15 minute. The supernatant was collected and of FeCl₃ and a dark green solution was obtained which used for further phytochemical analysis.[4] indicate the presence of tannins. Ethyl acetate and petroleum ether extraction Test for anthraquinone 20 grams of dried plant material was extracted in 100 ml of Each extract (0.5g) was boiled with 10% HCl for few Ethyl acetate and Petroleum ether. The flasks were covered minutes. The reaction mixtures was then filtered and with aluminum foil and kept on rotatory shaker at 191 rpm allowed to cool. Equal volume of CHCl₃ was added to each for 24 hour at room temperature. After 24 hour the solution filtrate along with few drops of 10% NH₃ and heated. Rose- was filtered through whattman filter paper-1. The filtrates pink color formation was obtained which indicate the were collected and were concentrated up to dryness by presence of anthraquinones. keeping it in hot air oven at 35⁰C. The stock solution (0.5 mg/ml) of each extract was prepared in Dimethyl sulfoxide Test for glycosides (DMSO).[1,4] Each extract (0.6g) was hydrolyzed with HCl and Methanol,n-hexen,chloroform fraction neutralized with NaOH solution and few drops of Fehling’s solution A and B were added. Red precipitate were formed 10 grams of dried plant material was soaked in 25 ml which indicate the presence of glycosides. methanol for 5 days and then subjected to repeated extraction with 25 3 until exhaustion of plant material. The Test for reducing sugars obtained were then evaporate at below 55⁰c.crude methenolic extracts were suspended in water and Extract was treated with Benedict’s reagent and boil for 5 successively partitioned with n-hexen and chloroform.[5,8] minutes in water bath .Formation of an orange red precipitate indicated the presence of reducing sugars. Antimicrobial activity 2 Test for saponins Three strains of Gram-positive (Staphylococcus aureus(MTCC NO:737), Bacillus subtilis(MTCC NO:441), Each extract (0.2g) was shaken with distilled water (5.0mL) Bacillus megaterium(MTCC NO:2949), Entrococci(MTCC and boiled. Frothing (appearance of creamy miss of small NO:439)) and three of Gram negative (Escherichia bubbles) was observed showed the presence of saponins. coli(MTCC NO:119), Proteus vulgaris(MTCC NO:426), PM 18|Volume 4|Issue 1|2018 BMR Phytomedicine Test for flavonoids Test for terpenoids Each extract (0.2g) was dissolved in diluted NaOH and few Each extract (0.2g) was mixed with CHCl₃ (2.0mL) and drops of HCl were added. A yellow solution turned into carefully concentrated H₂SO₄(3.0mL) was added to form a colorless which indicate the presence of flavonoids. layer. Test for. phlobatanins Test for phenol Each extract (0.5g) was dissolved in distilled water and Treat the extract with 3ml of 10% lead acetate solution. A filtered. The filtrate was then boiled with 2% HCl solution. bulky white precipite indicate the presence of phenolic Red precipitates were obtained which showed the presence compounds. of phlobatanins. RESULT Test for steroids Phytochemical analysis Acetic anhydride (2.0mL) was added to mixture of extract (0.5g) and H₂SO₄ (2.0mL) of each extract. The change of The primary Phytochemical screening of different extraction color from violet to blue or green in some samples showed was done and ascertain bioactive compound are presence. the presence of steroids. The result show in Table-2. Table 2: Phytochemical screening of n-hexane, chloroform, ethyl acetate, petroleum ether and water
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