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Dennis Final Thesis.Pdf KWAME NKRUMAH UNIVERSITY OF SCIENCE AND TECHNOLOGY, KUMASI, GHANA Evaluation of the anti-proliferative effect, antioxidant and phytochemical constituents of Ficus pumila Linn. By Dennis Torkornoo, BSc A Thesis submitted to the Department of Biochemistry and Biotechnology, Kwame Nkrumah University of Science and Technology in partial fulfillment of the requirements for the Degree of MASTER OF PHILOSOPHY (BIOCHEMISTRY) COLLEGE OF SCIENCE OCTOBER, 2017 DECLARATION I hereby declare that this submission is my own work towards the MPhil and that, to the best of my knowledge, it contains no material previously published by another person nor material which has been accepted for the award of any other degree of the University, except where due acknowledgement has been made in the text. ……………………………..…… …………………………..…… Dennis Torkornoo Date (PG4575715) Certified by: ……………………………..…… …………………………..…… Dr. Christopher Larbie Date (Supervisor) Certified by: ……………………………..…… …………………………..…… Dr. Peter Twumasi Date (Head of Department) ii ABSTRACT Plants, such as Ficus pumila, have been indispensable in treating and managing diverse forms of diseases including cancer. About 80% of the people living in developing countries depend on medicinal plants for their healthcare needs based on the knowledge of traditional use of medicinal plants. This study was aimed at evaluating the heavy metal content of the raw powder and extract of the plant, the phytochemical constituents, antioxidant effect by the use of DPPH assay, the total phenolic content using Folin Ciocalteu assay and the cytotoxic effect using the MTT Assay of Ficus pumila ethanolic extract, methanolic and hydro fractions on liver cancer cells (HepG2), Leukemic cell (Jurkat) and normal liver cells (Chang). FTIR analysis was also done to determine the functional groups of the active components of the F. pumila fractions. The structure of the compound of the most active fraction was determined using Gas Chromatography-Mass Spectrometry (GC-MS). Alkaloids, terpenoids, flavonoids, cardiac glycoside, saponins and tannins were present in the ethanolic extract of F. pumila. The heavy metal analysis revealed the presence of Iron in both the raw powder (1.97 ± 0.11 mg/l) and extract (0.92 ± 0.02 mg/L). Zinc was also detected in both the raw powder (1.19 ± 0.00 mg/l) and extract (0.6595 ± 0.02 mg/l). These levels were below the permissible level set by FAO/WHO. The results from the FTIR revealed the presence of alkynes, alkyl halides, aromatics and aliphatic amines common to all fractions and compounds such as Phenol, 2,4 –bis (1,1-Dimethylethyl) and Dodecane, 2,6, 10-trimethyl were detected in the samples by GC-MS. The DPPH assay also showed that all the fractions scavenged DPPH free radical in a dose dependent manner as compared to the positive control (Ascorbic acid). The hydro fraction had the strongest DPPH scavenging activity (EC50 =0.09 ± 0.00 mg/ml). Phenolic compounds were present in all the fractions with the hydro fraction having the highest level (16950 ± 331.95 mg GAE/100g). The MTT assay revealed that all the samples were cytotoxic against the cancer cell lines but only methanolic fraction was selective towards the Jurkat cell lines (Selectivity Index = 2.822) with IC50 value of 248.1 µg/ml. The most active fraction in terms of the antioxidant assay was the hydro fraction and with respect to the cytotoxicity assay was the methanolic fraction. This increase the prospect that this plant contains compound(s) which could serve as leads for novel anticancer drugs. iii ACKNOWLEDGEMENT I would like to give thanks to the Almighty God for the gift of life and for seeing me through this entire work. I would also like to express my sincerest gratitude to my supervisor Dr. Christopher Larbie for his continuous support in my postgraduate study and research, for his patience, motivation, enthusiasm and immense knowledge. His guidance helped me in all the time of research and writing of this thesis. I could not have imagined having a better supervisor, advisor and mentor for my postgraduate study. I would also like to thank Dr. Regina Appiah-Opong, Dr. Bright Adu, Mr. Caleb Sinclear and Ms. Eunice Dotse of Noguchi Memorial Institute for Medical Research, and Mr. Nat Yaw, Technician, KNUST for their assistance during the laboratory sessions. God bless you all. I would also like to thank all my friends, especially Jasmine and Sefakor, and mates who helped me throughout the work. I would also like to thank my family; my mother Mrs. Amponsah Gloria for supporting me spiritually and financially throughout my life. The financial support of the KNUST Research Fund (KReF) is greatly appreciated. God bless you. iv TABLE OF CONTENT DECLARATION ............................................................................................................ ii ABSTRACT .................................................................................................................. iii ACKNOWLEDGEMENT ............................................................................................ iv TABLE OF CONTENT ................................................................................................. v LIST OF FIGURES ................................................................................................... viii LIST OF TABLES ........................................................................................................ ix CHAPTER ONE ............................................................................................................. 1 1.0 INTRODUCTION .................................................................................................... 1 1.1 PROBLEM STATEMENT .................................................................................... 2 1.2 STUDY OBJECTIVE ............................................................................................ 3 1.2.1 Specific Objectives .............................................................................................. 4 1.3 JUSTIFICATION .................................................................................................. 4 CHAPTER TWO ............................................................................................................ 5 2.0 LITERATURE REVIEW ........................................................................................ 5 2.1 CANCER ............................................................................................................... 5 2.1.1 Oncogene Theory ................................................................................................ 5 2.2 GLOBAL CANCER TRENDS: A GROWING DISPARITY ................................. 6 2.2.1 Disparities in incidence ....................................................................................... 6 2.2.2 Disparity in mortality .......................................................................................... 7 2.3 CAUSES OF CANCER IN DEVELOPING COUNTRIES .................................... 8 2.3.1 Infectious diseases ............................................................................................... 8 2.3.2 Environmental and dietary factors ..................................................................... 10 2.3.3 Effects of tobacco .............................................................................................. 11 2.4 IMMUNODIAGNOSIS OF CANCER ................................................................. 14 2.5 IMMUNOLOGIC DETECTION OF TUMOUR ANTIGENS .............................. 15 2.5.1. Detection of myeloma proteins produced by plasma cell tumours ..................... 15 2.5.2. Detection of Αlpha-Fetoprotein (AFP) .............................................................. 15 2.5.3. Carcinoembryonic Antigen ............................................................................... 16 2.5.4. Detection of prostate-specific antigen ............................................................... 16 2.5.5. Cancer Antigen – 125 ....................................................................................... 17 v 2.5.6. Other markers .................................................................................................. 17 2.6 PREVENTION OF CANCER .............................................................................. 17 2.6.1 Tumour Immunoprophylaxis ............................................................................. 19 2.7 TREATMENT REGIMES OF CANCER AND EFFECTS ................................... 20 2.8 FREE RADICALS AND ANTIOXIDANTS ........................................................ 21 2.8.1 Free radicals ...................................................................................................... 21 2.8.2 Antioxidants ...................................................................................................... 22 2.9 PHYTOCHEMICALS ......................................................................................... 23 2.9.1 How phytochemicals work ................................................................................ 23 2.10 MEDICINAL PLANTS AS ANTICANCER AGENTS ...................................... 24 2.10.1 Mechanistic action of anticancer agents ........................................................... 25 2.10.1 Anticancer activities of some plants................................................................. 26 2.11. FICUS PUMILA LINN. .................................................................................... 28 2.11.1 Classification .................................................................................................
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