Western Michigan University ScholarWorks at WMU Master's Theses Graduate College 6-1996 An Investigation on Nonenzymatic Autoxidation of Phenolic Compounds in Natural Waters Lei Jiang Follow this and additional works at: https://scholarworks.wmich.edu/masters_theses Part of the Chemistry Commons Recommended Citation Jiang, Lei, "An Investigation on Nonenzymatic Autoxidation of Phenolic Compounds in Natural Waters" (1996). Master's Theses. 4357. https://scholarworks.wmich.edu/masters_theses/4357 This Masters Thesis-Open Access is brought to you for free and open access by the Graduate College at ScholarWorks at WMU. It has been accepted for inclusion in Master's Theses by an authorized administrator of ScholarWorks at WMU. For more information, please contact [email protected]. AN INVESTIGATION ON NONENZYMATIC AlITOXIDATIONOF PHENOLIC COMPOUNDS IN NATURAL WATERS by Lei Jiang A Thesis Submitted to the Facultyof The GraduateCollege in partial fulfillment of the requirements forthe Degree of Master of Arts Department of Chemistry WesternMichigan University Kalamazoo,Michigan June 1996 / ACKNOWLEDGMENfS First, I am very grateful to the members of my committee, Dr. Schreiber, Dr. Howell and Dr. Berndt. Without their constant guidance and support, it would have beenimpossible for me to finish the thesis. Second, I would like to express my sincere gratitude to the Department of Chemistry for continuous support throughout my study. I appreciate all the encouragement from our faculty members, staff and graduate students. I enjoyed my study and research in such a family-like environment. To my husband, my father and my sister, I extend my wholehearted appreciation. They are my source of inspiration and my continuous drive to finish in excellence. Finally, I dedicated this thesis to my beloved mother. Lei Jiang 11 AN INVESTIGATION ON NONENZYMATICAUTOXIDATION OF PHENOLIC COMPOUNDS IN NATURAL WATERS Lei Jiang, M.A. WesternMichigan University, 1996 The nonenzymatic autoxidation of the phenolic compoundschlorogenic acid in aqueous solution was studied. The study demonstrated that the oxidation rate of chlorogenic acid was strongly dependent on pH. The higher the pH, the faster the rate of oxidation. The involvement of phenolate ions and radicals in oxidation reactions are two possible reasons for the pH dependency. The presence of a trace amounts of reducible transition metal ions, such as copper(II), dramatically increases the oxidation rate of chlorogenic acid. Light also increases the oxidation rate of chlorogenic acid. However, the photochemical reaction was a much slower process. The TLC and HPLC results show that the oxidation of chlorogenic acid produces a mixture. The percentage of the various components in the mixture was affected by oxidation time. The NMR results indicate that under strong alkaline conditions, some chlorogenic acids were initially hydrolyzed into caffeic acid and quinic acid. The study also showed that the oxidation of caffeic acid was pH dependent. The oxidation rate of caff eic acid was much faster than that of chlorogenic acid under acidic and neutral conditions. Both NMR and UV /VIS results demonstratedthat the conjugated double bond on the side chain of chlorogenic acid was involved in the oxidation reaction. TABLE OF CONTENTS ACKNOWLEDGMENTS................................................................. 11 LIST OFT ABLES......................................................................... V LIST OF FIG URES .................................. , ................................ .'... vi CHAPTER I. INTRODUCTION................. :.............................................. 1 Motivation. .. .. .. .. .. .. .. .. .. .. .. .. .. .. .. .. .. .. .. .. .. 1 Phenolic Compounds........................................................ 1 Humic Substances ......-.................... ................................. 2 TransitionMetals in Natural Waters....................................... 6 Pul1)0se of Study .. .. .. .. .. .. .. .. .. .. .. .. .. .. .. .. .. .. 8 II. LITERATURE REVIEW......................................................... 10 Chlorogenic Acid ............................................................ 10 Enzymatic Oxidation of Phenolic Compounds........................... 12 Nonenzymatic Oxidation of Phenolic Compounds. .. .. .. .. .. .. 13 III. EXPERIMENTALMErHODS................................................. 18 Materials. .. .. .. .. .. .. .. .. .. .. .. .. .. .. .. .. .. .. .. .. 18 Propertiesof Materials ...................................................... 18 Purity Checking.............................................................. 19 Sample Preparation.......................................................... 20 Aqueous Sample . .. .. .. .. .. .. .. .. .. .. .. .. .. .. .. 20 Recovery of Oxidation Products. .. .. .. .. .. .. .. .. .. 22 Instrumental Technique ..................................................... 22 Differential Scanning Calorimetry (DSC)........................... 22 lll Table of Contents-continued CHAPTER FreezeDry .. .. .. .. .. .. .. .. .. .. ... .. .. .. .. .. 23 Ultraviolet -Visible Spectrophotometry(UV NIS)................. 23 Thin-layer Chromatography(TLC) .................................. 23 High-performance Liquid Chromatography (HPLC).............. 24 Nuclear Magnetic Resonance .. .. .. .. .. .. .. .. .. .. .. 25 IV. RESULTS AND DISCUSSIONS .............................................. 26 Factors Related to Nonenzymatic Oxidation.............................. 26 Effect of pH............................................................. 26 Effect of Transition Metal Copper(II)................................ 36 Effect of Light.......................................................... 40 Separation of Reaction Mixtures... .. .. .. .. .. .. .. .. .. 42 TLC...................................................................... 42 HPLC.................................................................... 45 Identification of Reaction Mixtures . .. .. .. .. .. .. .. .. 47 V. CONCLUSIONS.................................................................. 51 APPENDICES A. Proton NMR Spectraof Caffeic Acid, Quinic Acid, Chlorogenic Acid and Its Oxidation Products....................................................... 53 BIBLIOGRAPHY.......................................................................... 61 IV UST OFTABLES 1. Solubility of Chlorogenic Acid in Different Solvents........................... 19 2. TLC Results fora Reaction Mixture of Chlorogenic Acid...................... 42 3. TLC Results forDifferent Reaction Mixtures.................................... 43 4 The Effect of Elution Solvents on Separationof a Reaction Mixture.......... 44 5. HPLCResults of the Separationof a ReactionMixture of Chlorogenic Acid ................................................................................... 46 6. Chemical Shifts ( o, ppm) of 1H NMR for Standards and Reaction Mixtures .. .. .. .. .. .. .. .. .. .. .. .. .. .. .. .. .. .. .. 48 7. Relative Peak Area at DifferentChemical Shifts in lH NMR Spectra........ 49 V LIST OF FIGURES 1. Scheme for the Fractionationof Humic Substances............................. 3 2. Structure of Humic Acid According to Fuchs.................................... 4 3. The Polyphenol Model.............................................................. 5 4. Proposed Mechamism forthe Reaction of CopperComplexes With Catechol............................................................................... 7 5. The Structure of 3-Caffeoylquinic Acid (IUPAC: 5-Caffeoylquinic Acid)... 10 6. Enzymatic Reaction Catalyzed by Chlorogenate CaffeoylTransfe rase........ 14 7. Structure of Dimer Isomers From the Oxidation of CaffeicAcid .............. 15 8. Proposed Mechanism of Dimer Formation . .. .. .. .. 16 9. Structures of Caffeic Acid and Quinic Acid . .. 18 10. Differential Scanning Calorimetry Thermogramof Chlorogenic Acid .. 21 11. UV/VIS Spectrumof ChlorogenicAcid (0.1 mM) at pH= 3.................. 27 12. UV/VIS Spectra of Chlorogenic Acid (0.1 mM) at pH= 10.5................. 27 13. UV/VIS Spectraof Chlorogenic Acid (0.1 mM) at pH= 12................... 30 14. UV/VIS Spectraof Chlorogenic Acid (0.05 mM) at pH= 7 .......... .. .. 30 15. Effect of pH on Oxidation of Chlorogenic Acid (0.05 mM).................... 31 16. Effect of pH on Oxidation of Caffeic Acid (0.05 mM).......................... 33 17. Effect of pH on Oxidation of CaffeicAcid (0.05 mM) in the Presence of Quinic Acid.... .. .. .. .. .. .. .. .. .. .. .. 34 18. Comparison of the Effect of pH on Different Systems.......................... 35 19. Catalytic Effect of Copper(II) on Oxidationof Chlorogenic Acid (0.05 mM)............................................................................ 38 20. UV/VIS Spectra of Chlorogenic Acid (0.1 mM) at pH= 7 in the Presence of Copper(Il).............................................................. 38 VI List of Figures-continued 21. Oxidation of Chlorogenic Acid (0.05 mM) at DifferentCopper(II) Concentrations .. .. .. .. .. .. .. .. 39 22. ChlorogenicAcid Oxidation With a Trace of Cu(II) (0.3 ppm)................ 40 23. Effectof Light on Oxidation of Chlorogenic Acid (0.05 mM) Without Copper(II).......... .. .. .. .. .. .. .. .. ..... .. .. .. .. .. .. .. 41 24. Effect of Light on Oxidation of Chlorogenic Acid (0.05 mM) With Copper(II).. .. .. .. .. .. .. .. .... .. .. .. 41 25. Peak Assignments for Chlorogenic Acid According to SADTLER............ 47 vu CHAPTER.I INTRODUCTION Motivation There are a variety of organic