In vitro and in vivo antioxidant potentials of Alchornea floribunda leaf extract, fractions and isolated bioactive compounds

Document Type: Original Research Article

Authors

1 Department of Pharmacology and Toxicology, Faculty of Pharmaceutical Sciences, Nnamdi Azikiwe University, Awka, Anambra State, Nigeria

2 Department of Pharmaceutical and medicinal Chemistry, Faculty of Pharmaceutical Sciences, Nnamdi Azikiwe University, Awka, Anambra State, Nigeria

3 Department of Pharmacology and Toxicology, Faculty of Pharmaceutical Sciences, University of Nigeria, Nsukka, Enugu State, Nigeria

Abstract


Objective: Alchornea floribunda leaves are widely used in ethnomedicine for the management of immuno-inflammatory disorders. We investigated the in vivo and in vitro antioxidant activity of the leaf extract, fractions and isolated compounds of A. floribunda.

Materials and
Methods: The ethyl acetate fraction of the methanol leaf extract was subjected to several chromatographic separations to isolate compounds 1-4. The structures of the isolated compounds were elucidated by a combination of 1D and 2D NMR and mass spectrometry. Oxidative stress was induced with carbon tetrachloride (CCl4). Further analysis on the isolated phenolic compounds were done using 2,2-diphenyl-1-picrylhydrazyl (DPPH), ferric reducing antioxidant power (FRAP) and hydrogen peroxide scavenging activity tests.

Results: The ethyl acetate fraction at 200 mg/kg produced significant (p<0.05) elevations of catalase enzyme activity and a significant (p<0.05) reduction in serum malondialdehyde. The chemical investigation of the ethyl acetate fraction led to the isolation of three flavans, (-) cathechin (1), (-) epicathechin (2), (+) epicathechin (3) and a flavanone, 2R, 3R dihydroquercitin (4). In hydrogen peroxide scavenging assay, (-) epicathechin exhibited an EC50 value of 8 μg/ml, similar to the standard ascorbic acid (EC50 = 8 μg/ml). (-) epicathechin showed scavenging of DPPH radical with EC50 value of 19 μg/ml while in the FRAP assay, it had EC50 value of 46 μg/ml which was lower than that of the standard, ascobic acid (EC50 = 66 μg/ml).

Conclusion: The medicinal uses of A. floribunda may be due to the antioxidant activities of its phenolic compounds.

Keywords

Main Subjects


Aebi H, 1984. Catalase In vitro. In: Methods in Enzymology: Oxygen Radicals in Biological Aebi H, 1984. Catalase in vitro. In Colowick SP, Kaplane NO eds. Methods in Enzymol, 105: 121-126.

Barrera G, 2012. Oxidative stress and lipid peroxidation products in cancer progression and therapy. ISRN Oncology, http://dx.doi.org/10.5402/2012/137289.

Burton GK and Jauniaux E, 2011. Oxidative stress. Best Pract Res Clin Obstet Gynaecol, 25: 287-299. 

Cartea ME, Francisco M, Soengas P, and Velasco P, 2011. Phenolic compounds in Brassica Vegetables. Molecules, 16: 251-280.

Di BD, Scapagnini G, Candore G, Castiglia L, Colonna-Romano G, Duro G, Nuzzo D, Lemolo F. Lio D, Pellicano M, Scafidi V, Caruso C, Vasto S, 2010. Immune-inflammatory responses and oxidative stress in Alzheimers disease: therapeutic implications. Curr Pharm Des, 16: 684-691.

Dragan A, Dusanka D, Drago B, and Nenad T, 2003. Structural-radical scavenging activity relationships of flavonoids. Croatica Chemica Acta, 76: 55-61.

Halliwell B and Gutteridge JMC, 2007. Free radicals in biology and medicine, 4th ed. Oxford university press, pp. 66-68. Oxford, UK.

Hamilton RT, Walsh ME, and Remmen HV, 2012. Mouse models of oxidative stress indicate a role for modulating healthy aging. J Clin Exp Pathol, S4:005. doi: 10.4172/2161-0681.S4-005.

Huang D, Ou B, Prior RL, 2005. The chemistry behind antioxidants capacity assays. J Agric Food Chem, 53: 1841-1856.

Iacopini P, Baldi M, Storchi P, Sebastiani L, 2008. Catechin, epicatechin, quercitin, rutin and reservarol in red grape: content, in vitro antioxidant activity and interactions. J Food Compos Anal, 21: 589-598.

Jones DP. 2008. Radical-free biology of oxidative stress. Am J Physiol Cell Physiol, 295: C849-68.

Kelly FT, 1998. Use of antioxidants in the prevention and treatment of disease. J Int Fed Clin Chem, 10: 21-23.

Khalili RMA, Shafekh SE, Norhayati AA, Fatahudin IM, Rahimah R, Norkamaliah H, Azimah AN, 2013. Total phenolic content and in vitro antioxidant activity of winged bean (Psophocarpus tetragonolobus). Pak J  Nutr, 12: 416-422.

Kim DO, Jeong SW, Lee CY, 2003. Antioxidant capacity of phenolic phytochemicals from various cultivars of plums. Food Chem, 81: 321-326.

Kolhir VK, Bykov VA, Baginskaja AI, Sokolov SY, Glazova NG, Leskova TE, Sakovich GS, Tjukavkina NA, Kolesnik YA, Rulenko IA, 2006. Antioxidant activity of a dihydroquercetin isolated from Larix gmelinii (Rupr.) wood. Phytother Res, 10: 478-482.

Kumar RS, Sivakumar T, Sunderam RS, Gupta M, Mazumdar UK, Gomathi P, Rajeshwar Y, Saravanan S, Kumar MS, Murugesh K and Kumar KA, 2005. Antioxidant and antimicrobial activities of Bauhinia racemosa L. stem bark. Braz J  Med Bio Res, 38: 1015-1024.

Kumaran A, Karunakaran JR, 2007. In vitro antioxidant activities of methanol extracts of five phyllanthus species from India. LWT-Food Sci  Technol, 40: 344-352.

Lobo V, Patil A, Phatak A, and Chandra N, 2010. Free radicals, antioxidants and functional foods: Impact on human health. Pharmacogn Rev, 4: 118-126.

Martin JE, Alizadeh BZ, Gonzalez-Gay MA, Balsa A, Pascual-Salcedo D, Fernandez-Gutierrez B, Raya E, Franke L, Vant SR, Coenen MJ, Van RP, Radstake TR, Koeleman BP, Martin J, 2010. Identification of the oxidative stress-related gene MSRA as a rheumatoid arthritis susceptibity locus by genome-wide pathway analysis. Arthritis Rheum, 62: 3183-3190.

Michalak A, 2006. Phenolic compounds and their antioxidant activity in plants growing under heavy metal. Pol J Environ Stud, 15: 523-530.

Moselhy SS, Ali HKH, 2009. Hepatoprotective effect of cinnamon extracts against carbon tetrachloride induced oxidative stress and liver injury in rats. Biol Res, 42: 93-98.

Nonaka G, Goto Y, Kinjo J, Nohara, T, Nishioka I, 1987. Tannins and Related Compounds. LII. Studies on the Constituents of the Leaves of           Thujopsis dolabrata SIEB. ZUCC. Chem  Pharm Bull, 35: 1105-1108.

Noundou XS, Krause RWM, Vuuren SFV, Ndinteh DT, Oliver DK, 2014. Antibacterial activity of the root stems and leaves of Alchornea floribunda. J Ethnopharmacol, 15: 1023-1027.

Nowak JZ, 2014. Oxidative stress, polyunsaturated fatty acids-derived oxidation products and bisretinoids as potential inducer of CNS diseases: focus on age-related macular degeneration. Pharmacol Rep, 65: 288-304.

OECD., 2011. Guidelines for the testing of chemicals. Acute Oral ToxicityAcute Toxic Class Method, Test no. 423, Organisation for Economic Co-operation and Development (OECD), Paris, France.

Odebiyi OO and Sofowora EA, 1978. Phytochemical screening of Nigerian medicinal plants II, Lloydia, 41: pp 234-235.

Ogbunugafor H, Igwo-Ezikpe M, Igwilo I, Ozumba N, Adenekan S, Ugochukwu C, Onyekwelu O and Ekechi A, 2012. Evaluation of antioxidant properties of Moringa Oleifera ethanol leaves extract and effect on serum lipid indices in rat. Maced J Med Sci, 5: 397-403.

Okawa M, Kinjo J, Nohara T, Ono M, 2001. DPPH (1,1-diphenyl-2-picrylhydrazyl) radical scavenging activity of flavonoids obtained from some medicinal plants. Bio Pharm Bull, 24: 1202-1205.

Okoye FBC, Osadeba PO, 2009. Studies on the mechanisms of anti-inflammatory activities of the extracts and fractions of Alchornea floribunda leaves. Asian J Trop Med, 2(3): 7-14.

Okoye FBC, Osadeba PO, 2010. A new anti-inflammatory flavonol glycoside from Alchornea floribunda leaves. Nat Prod Res, 24: 266-273.

Okoye FBC, Osadebe PO, Nworu CS, Okoye NN, Omeje EO, Esimone CO, 2011. Topical anti-inflammatory constituents of lipophilic leaf fractions of Alchornea floribunda and Alchornea cordifolia. Nat Prod Res, 25: 1941-1949.

Okoye FBC, Osadebe PO, Proksch P, Edrada-Ebe RAl, Nworu, CS, Esimone CO, 2010. Anti-inflammatory and membrane-stabilizing stigmastane steroids from Alchornea floribunda leaves. Planta Med, 76: 172-177.

Oluyemi KA, Okwuonu UC, Baxter DG, Oyesola TO, 2007. Toxic effects of methanolic extract of Aspilia africana leaf on the estrous cycle and uterine tissues of wister rats. Int J Morphol, 25: 609-614.

Pandey MK, Mittra P, Maheshwari PK, 2012. The lipid peroxidation product as a marker of oxidative stress in Epilapsy. J Clin Diagn Res, 6: 590-592.

Patel A, Patel NM, 2010. Determination of polyphenols and free radical scavenging activity of Tephrosia purpurea linn leaves (Leguminosae). Pharmacogn Res, 2: 152–158.

Ronald LP, Xianli W and Karen S, 2005. Standardized methods for the determination of antioxidant capacity and phenolics in foods and dietary supplements. J Agric Food Chem, 53: 4290-4302.

Rout SP, Choudary KA, Kar DM, Das L, Jain A, 2009. Plants in traditional medical system-future source of new drugs. Int J Pharm  Pharm Sci, 1: 1-23.

Sasaki YF, Kawaguchi S, Kamaya A, Ohshita M, Kabasawa K, Iwama K, Taniguchi K, Tsuda S, 2002. The comet assay with 8 mouse organs: results with 39 currently used food additives. Mutat Res/Genet Toxicol Environ Mutagen, 519: 103-109.

Soobrattee MA, Bahorun T, Aruoma OI, 2006. Chemopreventive actions of poly-phenolic compounds in cancer. Biofactors, 27: 19-35.

Sun M and Zigma S 1978. An improved spectrophotometer assay of superoxidant dismutase based on epinephrine antioxidation. Anal Biochem, 90: 81-89.

Thirunavukkarasu SV, Venketaraman S, Lokesh U, 2010. In vitro antioxidant and antibacterial activity of polyherbal Manasamitra vatakam drug. J Pharm Res, 3: 2042-3047.

 

Thomas C, Mackey MM, Diaz AA, Cox DP, 2009. Hydroxyl radical is produced via the fenton reaction in submitochondrial particles under oxidative stress: implications for disease associated with iron accumulation. Redox Rep, 14: 102-108.

Treas GE and Evans WC, 2002. Textbook of pharmacognosy, 15th ed Saunders Publishers, London. Pp 246-393.

Usoh IF, Akpan EJ, Etim EO, Farombi EO, 2005. Antioxidant actions of dried flower extracts of Hibiscus sabdariffa. Pakistan J Nutr, 4:135-141.

Veeru P, Kishor MP, Meenakshi M, 2009. Screening of medicinal plants for antioxidant activities. J Med Plant Res, 3:608-612.

Vijay P and Vimukta S, 2014. The role of natural antioxidants in oxidative stress induced diabetes mellitus. Res J Pharm Sci, 3: 1-6.

Vongtau HO, Abbah J, Chindo BA, Mosugu O, Salawu AO, Kwanashie HO, Gamaniel KS, 2005. Central inhibitory effects of the methanol extract of Neorautanenia mitis root in rats and mice. J Pharm Biol, 43: 113-120.

Yeh C and Yen G, 2006. Induction of hepatic antioxidant enzymes by phenolic acids in rats is accompanied by increased levels of multidrug resistance-associated protein 3 mRNA Expression.  J Nutr, 136: 11-15.

Zhang J, Li LX, Liu XH, Wang Y and Zhao DB, 2012. Flavonoids from Artemisia sphaerocephala and their free readical scavenging activity using DPPH. Chem Nat Compd, 48: 879-880.