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Antioxidant and Analgesic Activities of Ethanol Leaf Extract of Brownea Coccinea

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Antioxidant and Analgesic Activities of Ethanol Leaf Extract of Brownea Coccinea Advancement in Medicinal Plant Research Vol. 3(2), pp. 69-74, June 2015 ISSN: 2354-2152 Full Length Research Paper Antioxidant and analgesic activities of ethanol leaf extract of Brownea coccinea Abeer Sarwar1, Tamanna Binte Huq1, Tasnia Malik1 and Biplab Kumar Das2* 1Department of Pharmaceutical Sciences, North South University, Bashundhara, Baridhara, Dhaka 1229, Bangladesh. 2Department of Pharmaceutical Chemistry, University of Dhaka, Dhaka 1000, Bangladesh. Accepted 5 May, 2015 ABSTRACT The current study aimed to investigate the analgesic and antioxidant properties of ethanol extract of Brownea coccinea leaves (EEBL). The initial phytochemical study revealed the presence of tannins, flavonoids, saponins, gums, terpenoids, steroids and reducing sugars. The antioxidant property was checked by using DPPH free radical scavenging method which was conducted at doses of 25, 50, 100, 200, 400 and 800 µg/ml. Antioxidant activity was also tested by reducing power capacity (Fe3+ to Fe2+) at concentrations of 25, 50, 100 and 200 µg/ml. All the results illustrated the presence of antioxidant property of B. coccinea. The analgesic activity was tested in mice at doses of 250 and 500 mg/kg by Hot Plate Method and Acetic Acid Induced Writhing Method. The analgesic analysis was conducted on Swiss Albino mice and the results clearly exhibited significant (p < 0.001 to 0.05) activity. Keywords: Brownea coccinea, Fabaceae, phytochemical, antioxidant, analgesic, EEBL. *Corresponding author: E mail- [email protected], Tel- +8801720051994. INTRODUCTION Plants are used as natural remedies by mankind from the Alzheimer’s, cancer and diabetes (Bae et al., 2009; very beginning of civilization (Seidl, 2002) and a large Parejo et al., 2002). Plants contain a large variety of free number of modern drugs that are used today had been radical scavenging molecules such as phenolic obtained from natural sources on the basis of their use in compounds (e.g. phenolic acids, flavonoids, quinones, folk culture. An estimated 80% of the population in and tannins), nitrogen compounds (e.g. alkaloids, developing countries depends on medicinal plants to amines), vitamins and other endogenous metabolites rich meet their primary healthcare needs (Hostettmann and in antioxidant activity (Cai et al., 2003; Zheng and Wang, Marston, 2002). 2001). One of the major contributing factors for developing Brownea coccinea which is commonly known as degenerative diseases like atherosclerosis, ischemic Scarlet Flame Bean, Mountain Rose and Copper Hoop heart disease, ageing, diabetes mellitus, cancer etc, is belongs to the subfamily Caesalpinioideae of the family oxidative stress (Ames et al., 1993). Oxidative process Fabaceae and is a native plant of Tropical South America results in the formation of free radicals such as (Klitgaard, 1991; Hanelt, 2001). The plant is used in .- . superoxide anion radical (O2 ), hydroxyl radical (OH ) and Guyana for treating gynecological disorders like non-free-radical species like H2O2 and singled oxygen dysmenorrhea and menorrhagia (De Filipps et al., 2004; (Huda Faujan et al., 2009). Antioxidants have the ability Roth and Lindorf, 2013). In Bangladesh, the plant is of curbing free radical chain reaction. Plants are one of known as Supti and is distributed in regions of Chittagong the best sources for natural antioxidants and and Sylhet. The Chakma tribe of Bangladesh uses the consumption of plants rich in antioxidants have been root and leaves of this plant for treating gynecological shown to reduce the risk of degenerative diseases like problems (Roy et al., 2008). Based upon the folkloric use Adv Med Plant Res 70 of this plant the present study had been designed to Antioxidant activity of plant extracts investigate the antioxidant and analgesic activities of DPPH free radical scavenging assay ethanol extract of B. coccinea leaves. The antioxidant activity of EEBL was assessed according to the method established by Brand-William et al. (1995) with slight MATERIALS AND METHODS modifications. The extracts (25, 50, 100, 200, 400 and 800 μg/ml) were prepared in 90% ethanol. The positive control was ascorbic Plant materials acid and was prepared at concentration of 25, 50, 100, 200, 400 and 800 μg/ml. DPPH solution was prepared in ethanol and 5 ml of The leaves of B. coccinea were collected from Bangladesh National the solution was mixed with the same volume of extract and the Herbarium, Dhaka, Bangladesh during the month of May 2013 and positive control ascorbic acid solution. After incubating the solutions were identified by the experts of Bangladesh National Herbarium for 30 min, the absorbance was read at 517 nm using a UV (BNH), Dhaka. (Accession number: 38374). Spectrophotometer. The percentage of radical scavenging by the sample was determined by comparing it with that of the ascorbic acid control group. Preparation of crude extracts The leaves of the selected plant were washed with water, Reducing power capacity separated from undesirable materials. They were then dried at room temperature (24 to 26°C) for two days. The fully dried leaves The reducing power capacity of EEBL was carried out according to were then grinded to coarse powder and were stored in zipper bag the method determined by Oyaizu (1986). 1.0 ml of the extract in refrigerator at +4°C for two weeks. After sieving, 400 g of solution of concentrations 25, 50, 100 and 200 μg/ml was mixed powered material obtained was taken in a clean, flat-bottomed with 2.5 ml of phosphate buffer (0.2 M, pH 6.6) and 2.5 ml of 1% glass container and soaked in 600 ml of 95% ethanol. The potassium ferricyanide. The mixture was then incubated at 50°C for container with its contents was sealed and kept for a period of 7 20 min. After incubation, 2.5 ml of trichloroacetic acid was added to days accompanying occasional shaking and stirring. The whole the mixture followed by centrifugation at 3000 rpm for 10 min. 2.5 mixture then underwent a coarse filtration by a piece of clean, white ml of the supernatant was mixed with 2.5 ml of distilled water and cotton material. Then it was filtered through Whatman filter paper. 0.5 ml of ferric chloride. The absorbance of the resulting solution The filtrate (Ethanol extract) obtained was evaporated by Rotary was measured at 700 nm against a blank by using UV evaporator (Bibby RE-200, Sterilin Ltd., UK) at 5 to 6 rpm and at Spectrophotometer. An increase in the absorbance of the reaction 68°C temperature. It rendered a gummy concentrate of dark bottle mixture indicates increasing reducing power. green color. The gummy concentrate was designated as crude ethanol extract of B. coccinea leaves (EEBL). Then the crude Analgesic activity of plant extracts extract was dried in freeze drier and preserved at +4°C for two weeks. Hot plate test method The hot-plate test employed for measurement of analgesic activity Phytochemical test was done in accordance to the method previously described by Lanhers et al. (1992) and which was modified by Mahomed and The phytochemical screening of EEBL was carried out to identify Ojewole (2004). Mice were divided into four groups- control, the presence of tannins, flavonoids, saponins, gums, steroids, positive control and test groups consisting of five mice in each alkaloids, reducing sugars and terpenoids by known methods group. The mice of each group were placed in the beaker (on the (Harborne, 1998; Sazada et al., 2009). The screening of the extract hot plate) in order to obtain its response to electrical heat induced was performed using the following reagents and chemicals: pain stimulus. Licking of the paws or jumping out of the beaker was Alkaloids with Wagner reagent, flavonoids with the use of taken as an indicator of the animal’s response to heat-induced pain concentrated HCl, tannins with 10% Ferric Chloride, and saponins stimulus. The time for each mouse to lick its paws or jump out of the with ability to produce suds. Gum was tested using Molish reagents beaker was taken as reaction time. Before treatment, the reaction and concentrated sulfuric acid, steroids with sulfuric acid, reducing time was taken once. Each of the test mice were thereafter treated sugar with the use of -napthol and hydrochloric acid and with distilled water, Diclofenac (10 mg/kg of body weight) and EEBL terpenoids with chloroform and concentrated sulfuric acid. These at the doses of 250 mg/kg and 500 mg/kg of body weight orally. phytochemicals are assumed to be responsible for biological The temperature of the hot plate was maintained at 54 ± 1°C. The activities of plants. Therefore before going for studying biological test sample, control and standard drug were given 30 min prior to activities, phytochemical screening is required in order to ascertain the beginning of the experiment. The mice were observed before the presence of phytochemicals from which we can make an initial and at 60, 120, 180 and 240 min after administration. The reaction assessment regarding activities. time was recorded when the mice licked their fore or hind paws or jumped prior to and 0, 60, 120, 180 and 240 min after oral administration of the samples. Percent analgesic score was Test animal calculated as: Swiss-albino mice aged 4 to 5 weeks with average weight of 22 to (PAS) = Tb-Ta/Tb × 100 30 g were used for the analgesic activity study. The mice were collected from the animal house of Department of Pharmaceutical Where, Tb = Reaction time (in second) before drug administration; Sciences of North South University and were housed in cages Ta = Reaction time (in seconds) after drug administration. under standard environmental conditions of room temperature 24 ± 1°C and 55 to 65% relative humidity. The animals were provided Acetic acid induced writhing test method with standard laboratory food and distilled water ad libitum and maintained at natural day night cycle.
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