Faculty of Biosciences Division of Plant Biology Department of Biological and Environmental Sciences University of Helsinki Faculty of Pharmacy Division of Pharmaceutical Biology University of Helsinki and Institute for Preventive Nutrition, Medicine and Cancer Folkhälsan Research Center Helsinki Traditional medicinal uses and biological activities of some plant extracts of African Combretum Loefl., Terminalia L. and Pteleopsis Engl. species (Combretaceae) Pia Fyhrquist Academic dissertation To be presented with the permission of the Faculty of Biosciences of the University of Helsinki, for public criticism in Auditorium XV (4072) at University Main Building, Unioninkatu 34, on November 16th, 2007, at 12 noon Helsinki 2007 1 Supervisors: Prof. Carl-Adam Hæggström, Ph.D. Botanical Museum of the Finnish Museum of Natural History University of Helsinki Finland Prof. Raimo Hiltunen, Ph.D. Faculty of Pharmacy Division of Pharmaceutical Biology University of Helsinki Finland Prof. Pia Vuorela, Ph.D. Department of Biochemistry and Pharmacy Åbo Akademi University Finland Reviewers: Prof. Riitta Julkunen-Tiitto, Ph.D. Natural Product Research Laboratory Department of Biology University of Joensuu Finland Prof. Sinikka Piippo, Ph.D. Botanical Museum of the Finnish Museum of Natural History University of Helsinki Finland Opponent: Prof. Jacobus N. Eloff, Ph.D. Phytomedicine programme Department of Paraclinical Sciences Faculty of Veterinary Science University of Pretoria South Africa © Pia Fyhrquist 2007 ISBN 978-952-10-4056-6 (printed version) ISBN 978-952-10-4057-3 (pdf; http://ethesis.helsinki.fi) Yliopistopaino, Helsinki 2007 2 TABLE OF CONTENTS ACKNOWLEDGEMENTS .............................................................................................................6 LIST OF ORIGINAL PUBLICATIONS .........................................................................................8 LIST OF ABBREVIATIONS ..........................................................................................................9 BOTANICAL TERMS ..................................................................................................................11 ABSTRACT...................................................................................................................................12 1. INTRODUCTION......................................................................................................................15 2. REVIEW OF THE LITERATURE............................................................................................18 2.1. Traditional Medicine in Africa .......................................................................................................................... 18 2.1.1. Documentation of African traditional medicine............................................................................................ 18 2.1.2. The importance of traditional medicine in different African countries ......................................................... 19 2.1.3. The practice of traditional medicine in the African community.................................................................... 20 2.1.4. Information on the medicinal uses of plants in Africa is in danger of disappearing ..................................... 21 2.2. Description, brief systematics and geographical extension of the Combretaceae family.............................. 22 2.2.1. The genus Terminalia L. ............................................................................................................................... 23 2.2.2. The genus Combretum Loefl......................................................................................................................... 28 2.2.3. The genus Pteleopsis Loefl. .......................................................................................................................... 31 2.3. Conventional antibiotics and the problem of microbial resistance................................................................. 33 2.3.1. Conventional antibiotics and their targets in bacterial cells .......................................................................... 33 2.3.2. Antifungal agents and their targets in fungal cells ........................................................................................ 34 2.4. Plant derived antimicrobials .............................................................................................................................. 35 2.4.1. Defense chemicals produced by plants.......................................................................................................... 35 2.4.1.1. Phenolic compounds.............................................................................................................................. 36 2.4.1.2. Terpenes ................................................................................................................................................ 39 2.4.2. Antimicrobial effects of plant extracts .......................................................................................................... 39 2.5. Antimicrobial effects and traditional medicinal uses of some species of Terminalia, Combretum and Pteleopsis reported in the literature ......................................................................................................................... 40 2.5.1. Terminalia species......................................................................................................................................... 40 2.5.1.1. African species of Terminalia ............................................................................................................... 41 2.5.1.2. Asian species of Terminalia .................................................................................................................. 50 2.5.2. Combretum species ....................................................................................................................................... 53 2.5.3. Pteleopsis species.......................................................................................................................................... 69 2.6. Anticancer drugs................................................................................................................................................. 70 2.6.1. Cytotoxic effects of species of Combretum reported in the literature ........................................................... 72 2.6.2. Cytotoxic and antimutagenic effects of some species of Terminalia according to the literature ..................78 3 3. AIMS OF THE STUDY.............................................................................................................83 4. EXPERIMENTAL .....................................................................................................................85 4.1. Selection of study area, plant material and bioactivity assays ........................................................................ 85 4.2. Ethnobotanical and -pharmacological survey in Tanzania ............................................................................. 87 4.2.1. Description of the study area......................................................................................................................... 87 4.2.1.1. Brief description of Miombo and Combretum woodlands..................................................................... 88 4.2.2. Methods for the fieldwork............................................................................................................................. 89 4.2.2.1. Ethnobotanical fieldwork (I).................................................................................................................. 89 4.2.2.2. Botanical fieldwork (I) .......................................................................................................................... 93 4.3. Extraction and fractionation of the plant material for antimicrobial and cytotoxicity assays................... 100 4.3.1. Crude extracts (I, II, III) .............................................................................................................................. 100 4.3.2. Fractionation with medium pressure liquid chromatography (MPLC) and thin layer chromatography (TLC) (II) ......................................................................................................................................................................... 100 4.4. Biological assays ................................................................................................................................................ 101 4.4.1. Bacterial and fungal strains (I, II)................................................................................................................ 101 4.4.2. Assay for antibacterial and antifungal activity (I, II) .................................................................................. 101 4.4.2.1. Estimating minimum inhibitory concentrations, MIC (I, II) ............................................................... 102 4.4.3. Cell lines (III).............................................................................................................................................. 102 4.4.4. Assays for measuring cytotoxic activities of the extracts (III) .................................................................... 103 4.4.4.1. Alamar Blue method...........................................................................................................................