EVALUATION OF HYDROGEN PEROXIDE SCAVENGING ACTIVITY OF METANOLIC EXTRACTS OF MORINGA (M. STENOPETALA) LEAVES

Marye Mulugeta Kassaw 1* , Tamiru Gashaw Gete 1 and Amha Admasie 2 . 1. College of Natural and Computational Sciences, Chemistry of Department Wolaita Sodo University, Ethiopia. 2. College of Health Sciences, School of Public Health Wolaita Sodo University, Ethiopia. ...................................................................................................................... Manuscript Info Abstract ......................... ........................................................................ Manuscript History

An antioxidant is a chemical that prevents the oxidation of other chemicals. They protect the key cell components by neutralizing the damaging effects of free radicals, which are natural by-products of cell metabolism (Badarinath et al., 2010 andMilan et al., 2010). In the organism, hydrogen peroxide is generated by activated phagocytes used to kill several bacterial and fungal strains, additionally under physiological conditions by peroxisomes and oxidative enzymes. An increase in the rate of production or a decrease in the rate of removal disrupts this balance and increases the levels of reactive oxygen species and free radicals. This indicates harmful effects of free radicals and other oxidants (M. Rama et al. 2011 andSerhat et al., 2012). Free radicals are responsible for causing a large number of diseases including cancer, cardiovascular disease, neural disorders, alzheimer's disease, mild cognitive impairment, parkinson's disease, alcohol induced liver disease, ulcerative colitis, aging and atherosclerosis (Nur et al., 2013, Nooman et al., 2008 and Aurelia, M. P., and Gheorghe, P. N., 2011). The plants have a natural antioxidants scavenge harmful free radicals from our body (Ranju et al., 2009). In 2002 World Health Organization report, medicinal plants would be the best source to obtain a variety of drugs. In developed countries about 80% of plants are used in traditional medicine (Adamu et al., 2004).

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Hydrogen peroxide is a weak oxidizing agent and can inactivate a few enzymes directly, usually by oxidation of essential thiol (-SH) groups. It can cross cell membranes rapidly, and inside the cell, H 2 O 2 probably reacts with Fe 2+ , and possibly Cu 2+ ions to form hydroxyl radical which may be the origin of many of its toxic effects. Thus, the removing of H 2 O 2 is very important for antioxidant defense in cell or food systems (Mahesh et al., 2010, Ranju et al., 2011and Serhat et al., 2012. Moringa plant family contains several phytochemicals and some of which are high interest because of their medicinal value (Mahesh et al., 2010). Moringaceae stenopetala is one of a species moringaceae, which have been found in Ethiopia. M. stenopetala is an important food and medicinal (heart failure, high blood pressure, and arrhythmia) plants that has been used for most southwestern Ethiopians, they called Aleco and/or Sheferaw. It is cultivated as a crop plant (Daljit et al., 2013). To quantify the antioxidant properties are important to human life. Therefore, Moringaceae stenopetala plants have been investigated for better understanding their medicinal properties.
Hence, the aim of this study was undertaken to evaluate the methanol leaves extract of moringa stenopetala for its antioxidant activity by using hydrogen peroxide scavenging method.

Sampling and Extraction:-
The fresh mature leaves of moringa stenopetala were collected from in wolaita Sodo, southern Ethiopia, March 2014. The plant was authenticated by senior expert Botanists, Wolaita Sodo University, Ethiopia. The moringa stenopetala leaves was dried under shade and crushed to coarse powder and the powdered moringa stenopetala leaves was taken for soxhlet extraction using methanol as solvent. The extraction was done 20 g of dried powdered material was extracted with 150 mL of methanol for three hours. The obtained extracts were filtered over Whatman No. 1 paper and the filtrate was collected and the fraction after filtration was dried under reduced pressure to get the crude dried fraction. The yield of dried fraction of methanol extract of methanolic leaves extract of moringa stenopetala was collected for further analysis (Vibha et  A solution of hydrogen peroxide (UNI-CHEM, 43 mM) is prepared in phosphate buffer (SAMIR TECH CHEM, 0.1 M, pH 7.4). The absorbance's of hydrogen peroxide is determined by 230 nm using a Model 752 UV-Vis spectrophotometer. Extracts (20, 40 and 60 µg/mL) in distilled water is added to hydrogen peroxide and absorbances at 230 nm were determined five times in 10 min interval against a blank solution containing phosphate buffer without hydrogen peroxide. For each concentration, a separate blank sample was used for background subtraction. The values of percentage scavenging capacity were calculated for various concentrations of the extract. Tests were conceded out in triplicate.

Statistical analysis:-
The results of these investigations are means and standard deviation of three measurements. The different concentrations of extracts have significantly different efficiencies and times variation is significantly greater than the variation due to the random error of measurement were tested by two-way ANOVA. A p value of 0.01 was taken to be significant.

Results and Discussion
The results of this study, Scavenging capacity of hydrogen peroxide in moringa stenopetala different concentration of H 2 O 2 containing extract was also evaluated in a series of the 10 min period of the assay and BHT as reference compound was comparable shown below table 1 and figure 1. 20 µg/mL methanol extracted moringa stenopetala exhibited 67.341-84.574% scavenging activity on hydrogen peroxide. On the other hand, using the same moringa stenopetala, 40 µg/mL and 60 µg/mL exhibited 50.50-64.336% and 30.412-47.083% hydrogen peroxide scavenging activity respectively. Scavenging activity of hydrogen peroxide in moringa stenopetala and BHT as reference compound was not remarkably different and shown to be 90.245 % to 42.5 %.  Results show that the scavenging activity values on hydrogen peroxide 20 μg/mL of the extracts of moringa stenopetala leaves increased than that of 40 μg/mL and 60 μg/mL. Whereas decreased scavenging activity was exhibited at concentration of 60 µg/mL. The maximum scavenging activity was exhibited for all concentration at 10 minutes. Therefore moringa stenopetala is biologically advantageous for cells to control the amount of hydrogen peroxide that is allowed to accumulate.