PROPAGATION AND QUALITY ASSESSMENT FOR THE INTRODUCTION OF GREYIA RADLKOFERI INTO COMMERCIALIZATION by NOLUYOLO NOGEMANE Submitted in accordance with the requirements for the degree of Doctor of Philosophy in the subject Agriculture at the UNIVERSITY OF SOUTH AFRICA SUPERVISOR: Dr G. Prinsloo CO-SUPERVISOR: Prof N. Lall 15 June 2017 DECLARATION Name: _____________________________________________________________ Student number: _____________________________________________________________ Degree: _____________________________________________________________ PROPAGATION AND QUALITY ASSESSMENT FOR THE INTRODUCTION OF GREYIA RADLKOFERI INTO COMMERCIALIZATION I declare that the above dissertation/thesis is my own work and that all the sources that I have used or quoted have been indicated and acknowledged by means of complete references. ________________________ _____________________ SIGNATURE DATE i PREFACE A section of the research study presented in this thesis has been accepted for publication in a peer-reviewed journal and parts of this research study were presented at conferences. Scientific publication in peer-reviewed journal: Nogemane N. and Prinsloo G., 2017. Seed Germination and Vegetative production of Greyia radlkoferi, an Indigenous Tree of Cosmeceutical Importance. Acta Horticulturae: In press Scientific publications in preparation: Nogemane N., Lall N., Prinsloo G., 2017. Metabolomics profile and anti-tyrosinase activity of Greyia radlkoferi leaf extracts in response to seasonal changes. Nogemane N., Lall N., Prinsloo G., 2017.Chemical composition and biological activities of Greyia radlkoferi leaf extracts processed by different drying methods using 1H-NMR based metabolomic approach as a quality control tool. International conferences: Nogemane N. and Prinsloo G., 2016. 1H-NMR based metabolomics approach to understanding the influence of seasons on the metabolomics profile of Greyia radlkoferi. 12th Annual Conference of the Metabolomics Society, Dublin, Ireland. Poster presentation. Nogemane N. and Prinsloo G., 2016. Effects of temperature on seed germination of Greyia radlkoferi, an indigenous tree of cosmeceutical importance. 7th International Symposium for Seed Transplant and Stand Establishment of Horticultural Crops (SEST, seed Indaba Africa). University of Pretoria, Gauteng, South Africa. Oral presentation. Local conferences: Nogemane N. and Prinsloo G., 2016. 1H-NMR-based metabolomics approach to understanding the drying effects on the secondary metabolites of Greyia radlkoferi. South African Association of Botanists (SAAB). University of Free State, Bloemfontein, South Africa. Oral presentation. ii Nogemane N. and Prinsloo G., 2016. 1H-NMR based metabolomics approach to understanding the influence of seasons on the metabolomics profile of Greyia radlkoferi. Conference for Southern African Biochemistry and Informatics for Natural Products (SABINA) 2016. Protea Kruger Gate, Mpumalanga, South Africa. Poster presentation. iii ACKNOWLEDGEMENTS My heartfelt gratitude goes to my mother, Kholeka Nogemane, for all her love and support throughout the duration of my studies. I would like to acknowledge my son Lingomso Nogemane, who has brought me so much joy and happiness. Many thanks also goes to my late father, GV Mabusela for being there for me until his departure- ‘I know you would have been proud’. To my supervisors, Dr Prinsloo and Prof. Lall, I am very grateful for your supervision and guidance during the course of my studies. I have become a better researcher because of your constant positive criticism. My appreciation also goes to the members of the Medicinal Plant Research Group under the leadership of Dr Prinsloo in the Department of Agriculture and Animal Health of the University of South Africa. Your comments and suggestions were most valuable to see me through this study. A special ‘thanks’ goes to Cynthia Marokane for assistance with laboratory techniques and some data collection. Your help is appreciated. Lastly, I would like to acknowledge the University of South Africa for the AQIP grant, which did not only fund the study but offered me time off work in order to undertake my PhD study on full-time basis. The Department of Science and Technology is also acknowledged for partly funding this study. Without funding, this work would not have been possible. iv TABLE OF CONTENTS LIST OF FIGURES…………………………………………………………………………………………………………………………….…….ix LIST OF TABLES…………………………………………………………………………………………………………………………………….xi APPENDICES…………………………………………………………………………………………………………………………………………xii ABSTRACT……………………………………………………………………………………………………………………………………………xiii CHAPTERS CHAPTER 1: GENERAL INTRODUCTION ................................................................................................1 1.1 BACKGROUND ..................................................................................................................................... 1 1.2 AIM AND OBJECTIVES.......................................................................................................................... 5 1.3 STRUCTURE OF THE THESIS ................................................................................................................. 6 1.4 REFERENCES ........................................................................................................................................ 8 CHAPTER 2: LITERATURE REVIEW ..................................................................................................... 11 2.1 GREYIA RADLKOFERI ......................................................................................................................... 11 2.1.1 Botanical description ................................................................................................................. 11 2.1.2 Distribution and Habitat ............................................................................................................ 12 2.1.3 Ethnobotanical and Phytochemical studies ............................................................................... 13 2.2 INTRODUCING MEDICINAL PLANTS INTO CULTIVATION .................................................................. 14 2.2.1 Seed germination ....................................................................................................................... 15 2.2.2 Vegetative production ............................................................................................................... 16 2.3 QUALITY DETERMINANTS OF RAW MEDICINAL PLANTS AND THEIR PRODUCTS ............................. 17 2.4 CULTIVATION OF MEDICINAL PLANTS: A POSSIBLE SOLUTION TO QUALITY PRODUCTS ................. 19 2.5 BRIEF HISTORY INTO CULTIVATION OF SOUTH AFRICAN INDIGENOUS PLANTS FOR COMMERCIALIZATION ............................................................................................................................ 21 2.6 PLANT METABOLOMICS AS A QUALITY CONTROL TOOL FOR MEDICINAL PLANTS .......................... 22 2.6.1 The metabolomics approach ..................................................................................................... 23 2.6.2 NMR as a metabolomic tool....................................................................................................... 25 v 2.6.3 Use of metabolomics in traditional medicine ............................................................................ 25 2.7 REFERENCES ...................................................................................................................................... 32 CHAPTER 3 ....................................................................................................................................... 44 DEVELOPMENT OF VEGETATIVE AND SEED PROPAGATION PROTOCOLS FOR THE EFFICIENT CULTIVATION OF GREYIA RADLKOFERI .............................................................................................. 44 ABSTRACT ................................................................................................................................................ 44 3.1 INTRODUCTION ................................................................................................................................. 45 3.2 MATERIALS AND METHODS .............................................................................................................. 48 3.2.1 Propagation by seeds ................................................................................................................. 48 3.2.2 Propagation by stem cuttings .................................................................................................... 49 3.3 RESULTS............................................................................................................................................. 50 3.3.1 Seed germination response to temperature ............................................................................. 50 3.3.2 Propagation by stem cuttings .................................................................................................... 51 3.4 DISCUSSION ....................................................................................................................................... 55 3.5 CONCLUSIONS ................................................................................................................................... 57 3.6 REFERENCES ...................................................................................................................................... 59 CHAPTER 4 ......................................................................................................................................