ABSTRACT DEVELOPMENT OF A TEACHING COULOMETRY INSTRUMENT FOR THE DIRECT DETERMINATION OF SULFUR COMPOUNDS AND OF ZINC INDIRECTLY by Jeralyne Beatriz Padilla Mercado The development and characterization of a teaching coulometry instrument for on-line data acquisition is described. A constant current source connected to nonisolated Pt electrodes in a 150-mL beaker served as the cell where the iodine titrant is electrochemically generated and allowed to react with the analyte reducing agent. A photodiode monitored the darkening of the solution due to the starch-iodine complex permitting this titration curve to be stored by a multifunctional chemical analysis system used for teaching for subsequent graphical analysis. Characterization of the instrument for direct analyte determination is performed with ascorbic acid, thiols, thiosulfate, and bisulfite. The number of electrons per mole of thiol for iodine titration of glutathione and N-acetylcysteine varied as a function of pH, indicating different reaction pathways. Ascorbic acid and the thiols are determined in dietary supplements with a recovery of 90- 100%. The indirect determination of zinc after its complexation with cysteine was performed in alkaline media. The titration endpoint times of cysteine with zinc are proportionally longer as compared to cysteine itself. The determination of ascorbic acid and zinc in a supplement could be titrated sequentially without changing the sample. Recovery of zinc ranged from 96-112% with a RSD range of 6-11%. DEVELOPMENT OF A TEACHING COULOMETRY INSTRUMENT FOR THE DIRECT DETERMINATION OF SULFUR COMPOUNDS AND OF ZINC INDIRECTLY A Thesis Submitted to the Faculty of Miami University in partial fulfillment of the requirements for the degree of Master of Science by Jeralyne Beatriz Padilla Mercado Miami University Oxford, Ohio 2017 Advisor: Neil D. Danielson Reader: Jiangjiang Zhu Reader: Dominik Konkolewicz Reader: Richard Bretz ©2017 Jeralyne Beatriz Padilla Mercado This Thesis titled DEVELOPMENT OF A TEACHING COULOMETRY INSTRUMENT FOR THE DIRECT DETERMINATION OF SULFUR COMPOUNDS AND OF ZINC INDIRECTLY by Jeralyne Beatriz Padilla Mercado has been approved for publication by The College of Arts and Science and Department of Chemistry and Biochemistry ____________________________________________________ Neil D. Danielson ______________________________________________________ Jiangjiang Zhu _______________________________________________________ Dominik Konkolewicz _______________________________________________________ Richard Bretz Table of Contents Chapter 1. Introduction P. 1 Chapter 2. Iodine coulometry with on-line photocell detection for a multifunctional chemical analysis (MCA) system P. 21 Chapter 3. Indirect determination of zinc by thiol complexation and iodine coulometric titration with photodiode detection P. 59 Chapter 4. Conclusions and future directions P. 75 Appendix. Oxygen meter finger probe studies P. 77 iii List of Tables Chapter 2 Table 2.1. Analytes determine in commercial products P. 27 Table 2.2. Average number of electrons per mole of analyte (% RSD). P. 28 Table 2.3. Standards and commercial products determined With the home-built coulometer. P. 30 Chapter 3 Table S1. Complete label listings of ingredients in the commercial samples. P. 74 Appendix Table A.1. Reproducibility studies with electric toothbrush. P. 81 Table A. 2. Attempts to make calibration curves of permanent marker paper chromatography separations. P. 82 Table A.3. Filter paper with permanent marker spots trials. P. 82 Table A.4. Trials of IR active compounds. P. 83 Table A.5. Teflon tape, nitrile glove, and salicylic acid sample in oximeter. P. 84 Table A.6. Blank trials with transfer pipette and Teflon tape. P. 85 iv List of Figures Chapter 1 Figure 1.1. A constant-current coulometer. P. 4 Figure 1.2. Coulometer with isolated electrodes and salt bridge versus coulometer with nonisolated electrodes. P. 4 Figure 1.3. Helical amylose units encasing 3I2 units. P. 7 Figure 1.4. Ascorbic acid structure. P. 15 Figure 1.5. Glutathione structure. P. 15 Figure 1.6. N-acetylcysteine structure. P. 15 Figure 1.7. Thiosulfate structure. P. 15 Figure 1.8. Bisulfite structure. P. 15 Figure 1.9. Cysteine structure. P. 17 2- Figure 1.10. Zncys2 complex at alkaline pH. P. 17 Chapter 2 Figure 2.1. Picture of constant-current coulometry instrument. From left to right: current source, electrodes in coulometric cell on top of magnetic photodiode and magnetic stirrer, and current-to-voltage converter circuit. MeasureNet station in the back. P. 24 Figure 2.2. Typical titration plot. P. 24 Figure S1. Proper alignment of electrodes, stir bar, and photocell. P. 33 Figure S2. Circuit diagram of the current-to-voltage converter. P. 34 Figure S3. Close-up picture of the breadboard used to construct the current-to-voltage converter circuit. P. 34 Figure S4. Titration plot of ascorbic acid titration using the 1 cm and 2 cm platinum electrodes as the anode and cathode, respectively. P. 35 v Figure S5. Titration plot of ascorbic acid titration with 2 cm and 1 cm platinum electrodes as anode and cathode. P. 35 Figure S6. Ascorbic acid titration plot using the 1 cm Pt cylinder as the anode and the medium counter electrode (Platinum Inlay Cat. No. 476060 from Corning). P. 36 Figure S7. N-acetylcysteine titration plot using a dialysis membrane to cover the 1 cm cylinder cathode. P. 36 Chapter 3 Figure 3.1. Zinc-cysteine complex structure as described in the literature. P. 61 Figure 3.2. (a) The raw plot (voltage vs. time), (b) first derivative plot (first derivative versus average time), and (c) the normalized voltage plot (normalized voltage vs. time). P. 64 Figure 3.3. Titration of cysteine and cysteine with zinc standard showed the least variability in endpoint measurements when using the buffer at its pKa, 9.2. P. 66 Figure 3.4. Effect of stir rate on titration endpoints. P. 67 Figure 3.5. Interference studies on cysteine titration endpoints. P. 69 Figure S1. Calibration curve of 1.24 x 10-1 mM cysteine with 0 - 1.74 x 10-5 mM zinc nitrate hexahydrate. The first four and last four points were taken 5 days apart using the same cysteine stock solution. P. 72 Figure S2. Cysteine and cysteine with zinc endpoints titrated in two different temperature baths. P. 72 Figure S3. Five-point calibration curve of titration of 1.3 x 10-1 mM cysteine with zinc standard ranging from 1.2 x 10-2 to 3.6 x 10-2 mM zinc. P. 73 vi Figure S4. Calibration curve of constant cysteine concentration with increasing zinc concentrations for ascorbic acid and zinc combined titrations. P. 73 Appendix Figure A.1. Heme group in hemoglobin. P. 78 Figure A.2. Picture of oximeter used in these studies. P. 79 Figure A.3. Green marker dot on electric toothbrush. P. 81 vii Dedication I want to dedicate this thesis to my parents, Ada and Rafael. I have worked tirelessly to reach my goals and make them proud. Thanks to their support and continuous words of encouragement I can say I am a master in chemistry. viii Acknowledgements Words cannot adequately express how grateful I am to have had Dr. Neil D. Danielson as my thesis advisor. It has been an honor for me to work with him during these past two years. I greatly appreciate his time and effort in helping me with my research and writing. He always goes above and beyond and makes the best out of every mentoring opportunity. From my time in the Danielson Research Group I will take with me his love of teaching, creativity, and resourcefulness. I also want to thank my committee members for taking the time to read my thesis and for providing helpful feedback on it. Dr. Jiangjiang Zhu for agreeing to be my graduate committee chair, Dr. Dominik Konkolewicz for his input during a crucial time of my zinc studies, and Dr. Richard Bretz for his insights. I am indebted to Dr. Zuleika Medina Torres, Prof. Edgardo Ortíz Nieves, and Dr. Stacey López Rivera for encouraging me to pursue graduate studies. I hope I am making you proud. Lastly, I want to thank the Danielson Research Group, the great friends I have met at Miami: Caitlyn; Andrew; Bryce, and my husband, Jean Pall, for their patience and support throughout this journey. ix Chapter 1. Introduction The coulometry research described in this thesis is two-fold in nature. Specific aim 1 is the development and characterization of a teaching coulometry instrument compatible with a multi-chemical analysis (MCA) station network. The versatility of iodine coulometry for sulfur compounds as well as ascorbic acid is demonstrated. Specific aim 2 describes an analytical chemistry research project involving the indirect determination of zinc using cysteine and iodine coulometry. These aims are explained in more detail at the end of this chapter. What immediately follows is a summary of the fundamentals of coulometry, the variety of applications, and how coulometry has been presented in the teaching literature. I. Fundamentals of coulometry Coulometry is an electrochemical method used to quantitate inorganic and organic analytes by measuring the current and time needed to change their oxidation state. Coulometry has many advantages when compared to other analytical procedures. First, it is exceptionally good when small amounts of sample are measured. Second, unstable titration reagents that would not be used otherwise can be generated in situ, and third its quick analysis time makes automation possible. Constant-current coulometry requires straightforward data analysis because the measurement charge (Q) is directly proportional to the product of current (I) and time (t) as shown in the following equation: Q = It (1.1) Furthermore, Faraday’s law can be used to relate the measurement charge to the amount of analyte in solution: molesanalyte = Q/nF (1.2) where n equals the moles of electrons participating in the redox reaction and F is Faraday’s constant, 96485.31 C/mol.1 Faraday’s constant represents the charge corresponding to one mole of electrons.