Dilruba Yeasmin
Department of Botany, University of Rajshahi, Rajshahi 6205, Bangladesh
Rawnak Jahan Swarna
Department of Botany, University of Rajshahi, Rajshahi 6205, Bangladesh
Mst. Samima Nasrin
Department of Botany, University of Rajshahi, Rajshahi 6205, Bangladesh
,Sarwar Parvez
Department of Botany, University of Rajshahi, Rajshahi 6205, Bangladesh
Mohammad Firoz Alam
Department of Botany, University of Rajshahi, Rajshahi 6205, Bangladesh
Chrysanthemum, Phytochemical analysis, Antioxidant, DPPH, Free radicals.
Department of botany, Rajshahi University Rajshahi-6205, Bangladesh,
Crop-Soil-Water Management
Plant material- White and yellow colour flower of chrysanthemum morifolium were collected from local farmer of Jessore, Bangladesh and pink colour flower of chrysanthemum morifolium was collected from local farmer of Rajshahi, Bangladesh. The plant material was collected in December- January, 2014. Dr. A. H. M. Mahbubur Rahman, Associate Professor, Department of botany, Rajshahi University Rajshahi-6205, Bangladesh, confirmed the taxonomic identification of the plant. Chemicals and Reagents-2, 2-Diphenyl-1-picrylhydrazyl (DPPH), ascorbic acid, methanol, ethanol, Dragendroff’s, Hager’s, Mayer’s, Wagner’s and Tannic acid reagents. Spot phytochemical analysis Preparation of sample- Initially well washed flowers were dried in hot air oven. The dried materials were coarsely powdered as a fine powder. Spot phytochemical screening of three colours (pink, yellow and white) flower of C. morifolium were carried out by using the following protocols as described below for the presence of alkaloids, flavonoids, glycosides, saponins, tannins, terpenoids and phenols. Determination of Alkaloids- 5 g fine powder was mixed up to moistered with 10 ml 2% HCl and heated in water at 60 °C for one hour. After cooling the sample was filtered through Whatmann No. 1 filter paper. Two drops of filtrates were put on a microscopic groove slide with one drop of the alkaloid detecting reagent. The relative abundance of precipitate, if any, formed in the plant sample with the reagents was considered as the presence of alkaloid (Aplin and Cannon 1971). Determination of Flavonoids- About 10 g plant sample was extracted with 20 ml ethanol (1:2) in aspirator bottle by soaking about 72 hours. After that few drops of conc. HCl were added to the alcoholic extract resulting red color, indicates the presence of flavonoids (Farnsworth 1985). Determination of Glycosides- A small amount of sample solution was mixed with 2ml of glacial acetic acid containing 1-2 drops of 2% solution of FeCl3. The mixture was then poured into another test tube containing 2ml of concentrated H2SO4. A brown ring at the interphase indicated the presence of cardiac glycosides (Trease and Evans 1989). Determination of Saponins- 20 ml water is added to 150 mg fine powder and shaken vigorously; layer of foam formation indicates the presence of saponins (Siddiqui and Ali 1997). Determination of Tannins- 2g fine powder was extracted with 10 ml distilled water (1:5), and was boiled for about 20 to 25 minutes. After cooling the extract was filtered. The filtrate was taken on 3 microscopic slides, two drops on each. Then to the first slide one drop 10% NaCl, to the second 1% gelatin and to the third 1% gelatin + 10% NaCl were added. The appearance of a white precipitate on the second and third was taken as positive test for tannins (Wall et al.1954). Determination of Terpenoids- 2ml of sample was mixed in 5ml of chloroform and concentrated H2SO4 2ml was carefully added to form layer. A reddish brown coloration of the interface was formed show positive result for the presence of terpenoids (Harborne 1973). ?Determination of Phenols- 5ml of sample was mixed with 2ml of 2% solution of FeCl3. A blue-green or black coloration indicated the presence of phenols. Antioxidant activity- Antioxidant activity of three colours (pink, yellow and white) flower of C. morifolium were estimated for their free radical scavenging activity by using DPPH (2, 2-Diphenyl-1-Picrylhydrazyl) as described by Hsu et al. (2007) with some modification. Experimental procedure of Antioxidant activity test- First, various concentrations like 20, 40, 60, 80, 100 mg/ml of sample in methanol were prepared. 2 ml of methanol solution of plant sample or standard at different concentrations was taken in test tube. 3 ml of 0.1 mm methanol solution of DPPH was added into the test tube. The test tube was incubating at room temperature for 30 minutes in dark place to complete the reaction. Then the absorbance of the solution was measured at 517 nm using a spectrophotometer against control. Ascorbic acid was used as standards (positive control). Then the percentage (%) inhibition activity was calculated according to Pavlov et al. (2002). % I= {(AO– A1)/AO}*100; Where, AO is the absorbance of the control, and A1 is the absorbance of the sample or standard. Sample was analyzed in two replications and data presented as mean (±) standard deviation (SD). Then % of inhibition was plotted against blank concentration and from the graph IC50 was calculated. IC50 value, the concentration of sample required for 50% scavenging of DPPH free radical are completed (Mandal et al., 2009). Statistical analysis- Statistical analysis (ANOVA) and Least Significant Difference (LSD) test was used to speculate further if there was a significant difference within varieties, various concentrations. P values <0.05 were considered as significant.
International Journal of Biosciences | IJB |, Vol. 9, No. 2, p. 69-77, 2016
Journal