Research Article COMPARATIVE ASSESSMENT OF PHYTOCHEMICALS AND ANTIOXIDANT POTENTIAL OF METHANOLIC AND AQUEOUS EXTRACTS OF CLERODENDRUM COLEBROOKIANUM WALP. LEAF FROM NORTH-EAST INDIA ABHISHEK DAS a , DIPANKAR CHAUDHURI b , NIKHIL BABAN GHATE b , ANUPAM CHATTERJEE a , NRIPENDRANATH MANDAL b* a Department of Biotechnology & Bioinformatics, North-Eastern Hill University, Shillong 793022, Meghalaya, b Division of Molecular Medicine, Bose Institute, Kolkata 700054, West Bengal, India. Email: mandaln@rediffmail.com Received: 24 July 2013, Revised and Accepted: 24 Aug 2013 ABSTRACT Objective: The present study was undertaken, to evaluate the in vitro antioxidant capacity as well as phytochemical quantification of Clerodendrum colebrookianum. Methods: A 70% methanolic extract (CCLM) and aqueous extract (CCLA) of C. colebrookianum Walp. leaves to evaluate different in vitro tests in diversified fields including the total antioxidant activity and the extent of their abilities to scavenge for various reactive oxygen species (ROS), reactive nitrogen species (RNS) and the analysis of various phytochemicals present in both the extracts followed by an attempt to clarify the active compounds through High performance liquid chromatography (HPLC) conferring antioxidant capacity. Results: Results confirmed a better efficacy of CCLM in projecting total antioxidant activity and scavenging of DPPH, hydroxyl, superoxide, nitric oxide and HOCl radicals. CCLM also displayed impressive amounts of phytochemicals like phenolic & flavonoid compounds, carbohydrates, tannins and alkaloids. HPLC chromatogram revealed the presence of compounds that represent the retention peaks of the compared standard phytocompounds-tannic acid, quercetin, catechin, reserpine, ascorbic acid and gallic acid. Conclusion: These results confirm the possible role of the 70% methanolic extract as a promising free radical scavenger and as a potent antioxidant source. Keywords: Free radicals, HPLC, Phytochemical analysis. INTRODUCTION The use of traditional medicine is widespread and plants provide a large source of natural antioxidants that might serve as leads for the development of novel drugs[1]. Normally formed as resultant by- products of a plethora of metabolic processes, the reactive oxygen species (ROS) and reactive nitrogen species (RNS), apart from playing a beneficial role in physiological processes, are more pronounced for their detrimental effects on living systems. The overproduction of ROS, resulting either from mitochondrial electron transport chain or excessive stimulation of NAD(P)H leads to oxidative stress thereby causing damage to cell membranes, lipids, proteins and nucleic acids[2]. These deleterious changes are implicated in the pathogenesis of various human diseases such as myocardial and cerebral ischemia, atherosclerosis, diabetes, rheumatoid arthritis, inflammation, cancer-initiation and ageing process. To deal with oxidative stress, antioxidant enzymes including superoxide dismutases (SOD), catalase (CAT), glutathione peroxidase (GSH-Px) etc., along with other intrinsic repair systems try to establish the redox equilibrium, but again fail occasionally, to entirely prevent the damage. This has encouraged the development of chemical antioxidants in the past few years and many have been clinically accepted, but their wide application has been restricted due to negative side effects and escalating costs[3]. Polyphenols, flavonoids, tannins and other bioactive substances of herbal origin display various biological roles, including antioxidant potential, free radical scavenging ability, anti-inflammatory, anti-diabetic, anticarcinogenic activities[4], and work as potential iron chelators[5]. Thus, many eyes drifted towards evaluating the natural antioxidants of plant origin for their respective efficacies which, on the other hand, showed lesser or no side effects. The East Indian Glory Bower Clerodendrum colebrookianum Walp., one of the most well-known among ~400 species of Clerodendrum, is an impressive inclusion in the folklore medicinal practices of North- East India for the treatment of various ailments and disorders since ages. Apart from antihypertensive property[6], its use has been reported for dealing with infant anti- colics pain, diabetes[7], helminthic infections[8], stomach disorder and headache[9] and for the treatment of cough, dysentery and some skin diseases[10]. Among the Khasi and Jaintia tribes in Meghalaya where it is commonly known as Sla Jarem, consuming the water after boiling the leaves is a traditional practice for the treatment of high blood pressure, malaria and liver troubles and in case of rheumatic pains, application of the warmed leaf - paste on the affected area is a common traditional practice. Experimental evidence on high-fat diet fed Wister rats show a declining level of total cholesterol, triglycerides and LDL, upon treatment with extracts of C. colebrookianum[6]. A dose dependent hypoglycemic effect is also observed with the aqueous extract of C. colebrookianum[11]. The present study aims towards the antioxidant and free radical scavenging activities of aqueous and 70% methanolic extracts of C. colebrookianum leaves followed by the analysis of the availability and abundance of different phytochemicals. MATERIALS AND METHODS Chemicals 2,2′-azinobis-(3-ethylbenzothiazoline-6-sulfonic acid) (ABTS) was procured from Roche diagnostics, Mannheim, Germany, and 6- hydroxy-2,5,7,8-tetramethylchroman-2-carboxylic acid (Trolox) was obtained from Fluka, Buchs, Switzerland. Potassium persulfate (K2S2O8), 2-deoxy-2-ribose, ethylene diammine tetraacetic acid (EDTA), ascorbic acid, trichloroacetic acid (TCA), mannitol, nitro blue tetrazolium (NBT), reduced nicotinamide adenine dinucleotide (NADH), phenazine metho-sulfate (PMS), sodium nitroprusside (SNP), 1,10-phenanthroline, sulphanilamide, naphthyl ethylenediamine dihydrochloride (NED), L-histidine, lipoic acid, sodium pyruvate, quercetin and ferrozine were obtained from Sisco Research Laboratories Pvt. Ltd, Mumbai, India. HPLC grade acetonitrile, ammonium acetate, hydrogen peroxide, potassium hexacyanoferate, Folin-ciocalteu reagent, sodium carbonate, mercuric chloride, potassium iodide, anthrone, vanillin, thiourea, 2,4-dinitro-phenylhydrazine, sodium hypochlorite, aluminium chloride, xylenol orange, butylated hydroxytoluene (BHT) and N,N- dimethyl-4-nitrosoaniline were taken from Merck, Mumbai, India. 1,1-diphenyl-2-picrylhydrazyl (DPPH), gallic acid, (+) catechin and International Journal of Pharmacy and Pharmaceutical Sciences ISSN- 0975-1491 Vol 5, Issue 4, 2013 A A c c a a d d e e m mi i c c S Sc c i i e e n n c c e e s s