i ii EVALUATION OF SOME CHEMICAL CONSTITUENTS OF SELECTED ENERGY DRINKS BY NURUDEEN SULAIMAN ABBAS DEPARTMENT OF CHEMISTRY FACULTY OF SCIENCE AHMADU BELLO UNIVERSITY, ZARIA NIGERIA AUGUST, 2014 iii EVALUATION OF SOME CHEMICAL CONSTITUENTS OF SELECTED ENERGY DRINKS By Nurudeen Sulaiman ABBAS, B.Sc. CHEMISTRY (ILORIN) 2010 M.Sc./Scie/5339/2011-2012 A THESIS SUBMITTED TO THE SCHOOL OF POST GRADUATE STUDIES, AHMADU BELLO UNIVERSITY ZARIA, IN PARTIAL FULFILLMENT OF THE REQUIREMENTS FOR THE AWARD OF MASTER OF SCIENCE DEGREE IN ANALYTICAL CHEMISTRY DEPARTMENT OF CHEMISTRY, FACULTY OF SCIENCE AHMADU BELLO UNIVERSITY, ZARIA NIGERIA AUGUST, 2014 iv Declaration I hereby declared that the work in this thesis entitled ―Evaluation of some chemical constituents of selected energy drinks‖ has been carried out by me in the Department of Chemistry. The information derived from the literature has been duly acknowledged in the text and a list of references provided. No part of this thesis was previously presented for another degree or diploma at this or any other institution. ____________________________ _________________________ Abbas Nurudeen Sulaiman Date v Certification This thesis entitled EVALUATION OF SOME CHEMICAL CONSTITUENTS OF SELECTED ENERGY DRINKS by Nurudeen Sulaiman ABBAS meets the regulations governing the award of the degree of Master of Science in Analytical Chemistry of Ahmadu Bello University, and is approved for its contribution to knowledge and literary presentation. ______________________ _____________________ Prof. C.E Gimba Date Chairman, Supervisory committee ________________________ _______________________ Dr. S.E Abechi Date Member, Supervisory committee _______________________ _______________________ Prof. V.O Ajibola Date Head of Department ________________________ ________________________ Prof. A.A Joshua Date Dean, School of Postgraduate Studies vi Dedication This thesis is dedicated to Almighty Allah and to my family who has supported me throughout the course of this work. vii Acknowledgement All praises and thanks are due to Allah ―the Exalted‖ Who in His Infinite mercies, granted me protection and ability to do this work. I would like to express my deepest gratitude to my supervisors Prof. C.E Gimba and Dr. S.E Abechi, whose constructive criticisms and encouragement made it possible for the successful completion of this project. Their patience and invaluable contribution guided me throughout the research period for which I am extremely grateful. My gratitude goes to Head of Department Prof. V. O. Ajibola, and other members of staff especially Mr. Celestine Ochigbo, Mallam M. Bashir, Mallam Ja‘far Adam, Mallam Ahmad Saleh and Mallam Ashir Adamu for their contribution and support. My profound and utmost gratitude goes to my family, especially my parents and my fiancée (Zainab Oshun) for their continuous love, prayers and supports. I sincerely appreciate the support, kindness and encouragement of my aunty, Mrs. Al- Hassan and family. I am also thankful to my colleagues and fellow students in the Department especially my sister, Oshomah Ummulkhair, Francis Peter, Abdullahi Amina and others too numerous to be mentioned. Thank you and God bless you all. viii Abstract This research work examined and compares the physicochemical properties and some chemical constituents of selected energy drinks. Fourteen (14) brands of energy drinks samples consisting eleven (11) liquid and three (3) powdered forms were randomly purchased. All samples were analyzed for their physicochemical properties (pH, turbidity, conductivity and total dissolved solids), trace and heavy metals, aspartame, sugar and caffeine contents. Results showed that the physicochemical properties (i.e. pH, turbidity, conductivity and total dissolved solids) ranged from 4.47 ± 0.012 - 5.96 ± 0.012, 8 ± 0.577 – 592 ± 1.155 NTU, 2.21 ± 0.006 – 1975 ± 1.732 µs/cm, and 243 ± 0.577 – 1064 ± 0.577 mg/L respectively. Energy drinks analyzed all fell within the FDA recommended range for the physicochemical properties analyzed. Iron, calcium, zinc and potassium were found in all the energy drinks and their concentration ranged from 1.961 ± 0.0003 - 0.294 ± 0.0005 mg/L, 2.763 ± 0.0009 - 19.310 ± 0.0015 mg/L, 0.045 ± 0.0001 - 13.887 ± 0.0037 mg/L, and 2.0 to 2500 mg/L respectively. The copper, lead and manganese concentration of energy drinks ranged from 0.002 ± 0.0002 - 0.102 ± 0.0003 mg/L, 0.028 ± 0.0006 - 0.209 ± 0.0009 mg/L and 0.003 ± 0.0001 - 0.024 ± 0.0002 mg/L respectively. The concentration of copper and manganese were below the MCL of 1.0 mg/L and 0.05 mg/L respectively while lead had a concentration above the MCL of 0.01 mg/L. Cadmium was not detected in all energy drinks except for sample EJ which had a concentration of 0.102 ± 0.0003 mg/L and exceeded the MCL of 0.005 mg/L. The caffeine, aspartame and sugar concentrations ranged from 1.11 mg/L – 2487.13 mg/L, 6.51 mg/L – 1491.19 mg/L, and 16.98 – 1686.73 mg/L respectively. Caffeine and aspartame concentrations in all the energy drink samples were below the FDA set standard of 400 mg/L and 3000 mg/L respectively except for sample AL which had a concentration above the set standard for ix caffeine. Though the analyzed parameters were mostly below the set standards, especially caffeine, aspartame and sugar, it is important that the pattern of consumption of these drinks must be monitored to minimize ingestion of excess doses of harmful substances to prevent the reported adverse effects. x Table of Contents Cover Page i Title page ii Declaration iii Certification iv Dedication v Acknowledgement vi Abstract vii Table of Contents ix List of Figures xv List of Tables xvi List of Abbreviations xvii CHAPTER ONE 1.0 INTRODUCTION 1 1.1 Energy Drinks 2 1.2 Contents of Energy Drinks 3 1.2.1 Caffeine 3 1.2.2 Taurine 4 1.2.3 Guarana 4 1.2.4 Glucuronolactone 5 1.2.5 Ginseng 6 1.2.6 Vitamins 6 1.2.7 Sugar 8 xi 1.3 Justification 8 1.4 Aim and Objectives 9 CHAPTER TWO 2.0 LITERATURE REVIEW 10 2.1 Review Work on Energy Drinks 10 2.2 Caffeine 11 2.2.1 Metabolism of caffeine 12 2.2.2 Caffeine and health 14 2.2.3 Research works on caffeine 15 2.3 Aspartame 16 2.3.1 Chemistry of aspartame 17 2.3.2 Biochemical data 18 2.4 Heavy Metal 21 2.4.1 Lead 22 2.4.2 Cadmium 22 2.4.3 Copper 23 2.4.4 Zinc 24 2.4.5 Manganese 24 2.4.6 Iron 25 2.4.7 Calcium 25 xii 2.4.8 Potassium 26 2.5 Carbohydrates 27 2.5.1 Sugar 27 2.5.2 Added sugars 28 2.5.3 Sugar-sweetened beverage 28 2.5.4 Sugar-sweetened beverages and health risks 29 2.6 Research Work on Beverages 30 CHAPTER THREE 3.0 MATERIALS AND METHODS 34 3.1 Materials 34 3.1.1 Chemicals and reagents 34 3.1.2 Preparation of stock solution 34 3.1.2.1 Preparation of standard solution for AAS 34 3.1.2.2 Preparation of standard solution for HPLC 35 3.1.2.3 Preparation of standard solution for UV 35 3.1.3 Apparatus and equipments 36 3.1.4 Sample collection 37 3.1.5 Sample preparation 37 3.1.5.1 Sample preparation for AAS 37 3.1.5.2 Sample preparation for HPLC 37 xiii 3.1.5.3 Sample preparation for UV 38 3.2 Analysis of Physicochemical Properties 38 3.2.1 Determination of pH 38 3.2.2 Determination of conductivity 38 3.2.3 Determination of turbidity 39 3.2.4 Total dissolved solid 39 3.3 Elemental Analysis 40 3.4 Determination of Carbohydrates (Sugar) 40 3.5 Determination of Caffeine and Aspartame 40 3.5.1 Preparation of pH 4.0 and pH 7.0 buffer solution 40 3.5.2 Buffer preparation 41 3.5.3 Mobile phase preparation 41 3.6 Statistical Analysis 41 CHAPTER FOUR 4.0 RESULTS 42 4.1 Physicochemical Parameters of Samples 42 4.2 Metal Concentrations 42 4.2.1 Concentration of heavy metals 42 4.2.2 Concentrations of essential metals 42 xiv 4.3 Concentrations of Caffeine, Aspartame and Sugar 42 CHAPTER FIVE 5.0 DISCUSSION 49 5.1 Physicochemical Parameters of the Samples 49 5.1.1 pH 49 5.1.2 Turbidity 50 5.1.3 Total dissolved solids 51 5.1.4 Conductivity 52 5.2 Metal Concentrations 52 5.2.1 Heavy metals 52 5.2.1.1 Cadmium 53 5.2.1.2 Lead 54 5.2.1.3 Copper 54 5.2.1.4 Manganese 55 5.2.1.5 Zinc 56 5.2.2 Essential Metals 57 5.2.2.1 Iron 58 5.2.2.2 Calcium 58 5.2.2.3 Potassium 59 xv 5.3 Caffeine, Aspartame and Sugar Concentrations in Energy Drinks 60 5.3.1 Caffeine 60 5.3.2 Aspartame 63 5.3.3 Sugar 65 CHAPTER SIX 6.0 Conclusion 68 6.1 Recommendations 70 Reference 72 Appendices 88 xvi List of Figures Figure 2.1: Structure of caffeine and adenosine 12 Figure 2.2: Metabolism of caffeine 13 Figure 2.3: Metabolism of aspartame 20 Figure 4.1: A comparison of the calculated concentration of caffeine in energy drinks with the maximum US FDA standard 47 Figure 4.2: A comparison of the calculated concentration of aspartame in energy drinks with the minimum and maximum US FDA standard 48 xvii List of Tables Table 4.1: pH, turbidity, total dissolved solids and conductivity of energy drinks 43 Table 4.2: Cadmium, copper, zinc, lead and manganese concentrations of energy drink 44 Table 4.3: Iron, calcium and potassium concentrations of energy drinks 45 Table 4.4: Concentration of sugar, caffeine and aspartame 46 xviii List of Abbreviations AAS Atomic Absorption Spectroscopy ATP Adenosine Triphosphate CYP1A2 Cytochrome P450 1A2 CYP2A6 Cytochrome P450 2A6 DKP 5-benzyl-3, 6-dioxo-2-piperazine acetic acid DNA Deoxyribonucleic Acid EU European Union FAO Food and Agriculture Organization FDA Food and Drinks Administration GC–MS Gas Chromatography–Mass Spectrometry HDL High Density Lipoprotein Cholesterol HPLC High Performance Liquid Chromatographic NAT2 N-acetyl transferase 2 NAG N-acetyl-beta-glucosamynidase NNS National Nutrition Survey RDA Recommended Daily Allowance RNA Ribonucleic Acid SCF Scientific Committee for Food SSB Sugar Sweetened Beverages USA United States of America UV Ultraviolet Visible Spectroscopy WHO World Health Organization XO xix CHAPTER ONE 1.0 INTRODUCTION Energy drinks refer to beverages that contain large doses of caffeine and other legal stimulants such as taurine, carbohydrates, glucuronolactone, inositol, niacin, panthenol, and β-complex vitamins which are considered as source of energy (Attila and Çakir, 2009).