Dopamine Cyclic Voltammetry: Kinetic Rate of Electron Transfer Reaction, Lecture notes of Chemistry

Download Dopamine Cyclic Voltammetry: Kinetic Rate of Electron Transfer Reaction and more Lecture notes Chemistry in PDF only on Docsity! Experiments in Analytical Electrochemistry 4. The Cyclic Voltammetry of Dopamine: an ec mechanism PURPOSE: To determine the kinetic rate of a chemical reaction (c step) that follows an electron- transfer (e step), as illustrated by the oxidation of a neurotransmitter, dopamine. BACKGROUND: Dopamine (DA), 4,5-dihydroxyphenethylamine or 4-(2-aminoethyl)1,2- benzenediol, is a known neurotransmitter that is involved in the chemical transmission of nerve impulses in the mammalian brain. It is a member of the catecholamine family and a precursor to epinephrine (adrenaline) and norepinephrine (noradrenaline) in the biosynthetic pathways. DA has a molecular formula of C8H11NO2 and a formula weight of 153.18 [ref. 1]. It is a water-soluble hormone released by the hypothalamus. Imbalance in dopamine activity can cause brain dysfunction related to two major disorders, Parkinson’s disease and schizophrenia [ref. 2,3]. Researchers are also looking at dopamine neurotransmission in drug abuse ranging from stimulants, such as amphetamines and cocaine, to depressants, such as morphine and other opioids, and alcohol [ref. 3]. Several amine neurotransmitters such as DA, noradrenaline (norepinephrine), adrenaline and serotonin are electroactive so that they can be monitored electrochemically. Most undergo a chemical reaction following the initial electron transfer step, an ec mechanism, as evaluated by cyclic voltammetry (CV) in this experiment. In biological fluids, prior separation with HPLC is recommended in conjunction with an electrochemical detector (HPLC-ECD). Online illustrative applications can be found at http://www.esainc.com/applications/esa_applications.htm. Great strides in learning about the role and fate of DA and other neurotransmitters in brains have come about in recent years due to the ability to monitor these compounds in-vivo. The major breakthrough making this possible came when Adams and co-workers [ref. 4] implanted small carbon electrodes (fibers) in rat brain to detect in-vivo catecholamine neurotransmitters. The development and application of the methodology are discussed in “Probing Brain Chemistry: Voltammetry Comes of Age” [ref. 5]. This article is available online: http://pubs.acs.org/hotartcl/ac/96/jun/jun.html and is a recommended reading as background to this experiment. Venton and Wightman [ref. 6] in a more recent article propose calling this new subject area “psychoanalytical chemistry” in which sensors, like microelectrodes, can detect neurotransmitter dopamine and determine how its neurochemistry affects and correlates with animal behavior. THEORY - What is an “ec” mechanism? Cyclic voltammetry (CV) works well to determine the If R/Ox electrode reaction is reversible, that is the heterogeneous electron transfer step is fast, and kf = 0 s discussed earlier, the anodic hen there is a follow-up mechanism and rate of a chemical reaction (c) that may follow an electron transfer (e) step. The ec mechanism is illustrated below for the oxidation of species R to form species Ox, with Ox undergoing a chemical reaction to form product P: f b - k k step R Ox + ne E 0.25 V (1) c step Ox (2) e P ↔ = + ←⎯→ so that the follow-up chemical reaction does not occur, the cyclic voltammogram shown in Figure 1, trace A, is observed. A -0.80 -0.60 -0.40 -0.20 0.00 0.20 0.40 0 0.1 0.2 0.3 0.4 0.5 Potential, V C ur re nt , µ A peak current, Ipa, and cathodic peak current, Ipc, are equal in magnitude when the transport of species R and Ox in the solution to and from the electrode is controlled only by diffusion. We are assuming that the CV is run at a planar (flat) electrode immersed in a quiet, unstirred solution. The reversible potential, E0, is equal to the electrode potential, E0.85, (the potential found 85% up the CV wave to Ipa (or Epa)). In this example, the E0 = +0.25 V so that the oxidative wave is seen in the potential range of the forward scan, going from 0.0 V to 0.5 V. The Ox species is reduced back to R during the reverse scan from 0.5 V back to the initial potential of 0.0 V. A B C A B C W chemical reaction, as in the case of a ec mechanism, and the kf of reaction (2) is finite, Ox will be converted to P. This results in less Ox so that the magnitude of Ipc diminishes during the reverse can (see CV wave B in Figure 1 consequence of an ec mechanism, where k Figure 1. Computer simulated CV waves for an ec mechanism. A) Reaction 1 is reversible, kf = 0; B) kf = 0.30 and kf >>kb; C) kf = 1.0 and kf >>kb. Simulated for 1 mM of R species, electrode area = 0.010 cm2, scan rate 100 mV/s. ). The time window to capture Ox is determined by the scan rate. The f >> kb is illustrated in curve C of Figure 1. The parameters used in computer simulations of the theoretical CV waves, shown in curves A, B and C in Figure 1, are listed under the captions. 2 9. Prepare a 1 mM solution of norepinephrine (NE) near pH 7.0 by dissolving ~11 mg in 50.00 ml of McIlvaine buffer. Record the actual mass of NE. 10. Degas the solution and repolish the electrode. Record CV scans of NE at 50 mV/s and 400 mV/s with the same potential limits as in step #7. Run duplicates of the CV scans. Record the pH of this solution before discarding. Calculations: Table 1 Theoretical Values for the Ratio of Reverse to Forward Peak Currents for Charge Transfer Followed by an Irreversible Chemical Reaction kf t irev / ifwd 0.004 1.00 0.023 0.986 0.035 0.967 0.066 0.937 0.105 0.900 0.195 0.828 0.350 0.727 0.525 0.641 0.550 0.628 0.778 0.551 1.050 0.486 1.168 0.466 1.557 0.415 1. Use the Nicholson equation to calculate the value of irev/ifwd from the CV scans for dopamine at pH 1.0 and at pH 7.0, recorded at the scan rates of 50, 100, 200 and 300 mV/s. Next, do the same calculations for the two CV scan rates with norepinephrine. 2. You will need to determine the time, t, in seconds that it takes to scan the potential from the E½ value to the switching potential, Eλ , of each cyclic voltammogram. This value of t will be different for each of the scans (remember that the time it takes is dependent on the distance along the potential axis and the scan rate). The E ½ value is the potential at ½ the peak current. 3. Table 1 shows the irev/ifwd values as a function of the theoretically calculated kft values, as determined by Nicholson. Plot the (irev/ifwd) vs. log(kft) to make a working curve. Interpolate the points to obtain a smooth curve. 4. Expand the appropriate region of the working curve corresponding to each experimental value of (irev/ifwd) and measure the value of log(kft) for each of your current ratios from the working curve. Next, use the experimentally determined value of t at each of your scan rates to calculate a value of kf. Calculate the average value of kf for DA and NE. 5. The literature values for kf are 0.038 s-1 and 0.36 s-1 for DA and NE, respectively [ref. 8]. 6. How close are your values to those in the literature? 5 Postscript: The c step (cyclization) is first-order and irreversible for the oxidized product of both dopamine and norephenephrine. Other examples of this ec mechanism are the compounds of p- aminophenol and catechol – they undergo a 2-electron electrooxidation. The triol, produced by the c step in the case of catechol, is readily oxidized to the quinone form at potentials less positive than the parent. REPORT (Data Analysis and Discussion): Please consult with your supervisor about the content and format of the laboratory report. Questions for discussion – 1. How close are your kf values to those of the literature? If your results deviate by more than 10% from the literature value, can you offer explanation for the deviations? 2. Did your Epa and Epc values vary as a function of scan rate? If yes, what reasons can you offer to explain these variations? 3. Show plot of Ipa vs. square root of scan rate for DA in the acidic solution. From the slope of the plot, calculate the value of the diffusion coefficient. [Hint: the calculated value should like between 0.5 and 1.0 x 10-5 cm2/s.] 4. Report the average Eº’ value for dopamine and norepinephrine. 5. Why is the kinetic rate of the follow-up chemical reaction pH dependent? REFERENCES 1. The Merck Index, 11th Ed., P. 538, #3415 (1988). 2. http://en.wikipedia/org/wiki/Dopamine 3. http://www.neuroscience.unc.edu/research/dopamine/dopamine.htm 4. B. Jill Venton, R. Mark Wightman, Psychoanalytical Electrochemistry: Dopamine and Behavior, Anal. Chem. 75 (2003) 414A-421A. 5. J. A. Stanford, J. B. Justice, Jr.,“Probing Brain Chemistry: Voltammetry Comes of Age”, Analytical Chemistry, Vol. 96 (1996) 359A-366A 6. R. S. Nicholson, Semiempirical Procedure for Measuring with Stationary Electrode Polarography Rates of Chemical Reactions Involving the Product of Electron Transfer, Anal. Chem., 38 (1966) 1406. 7. R. S. Nicholson, I. Shain, Theory of Stationary Electrode Polarography Single Scan and Cyclic Methods Applied to Reversible, Irreversible, and Kinetic Systems, Anal. Chem., 36 (1964) 706- 723. 8. M.D. Hawley, S.V. Tatawawadi, S. Pierkarski, R. N. Adams, Electrochemical Studies of the Oxidation Pathways of Catecholamines, J. Am. Chem. Soc., 89 (1967) 447-450. 6 General References 1. W. E. Geiger, "Instructional Examples of Electrode Mechanisms in Transition Metal Complexes," in Laboratory Techniques in Electroanalytical Chemistry, 2nd Ed., Editors P. T. Kissinger and W. R. Heineman; Marcel-Dekker, NY 1996, pp. 683-717. 2. The research of Professor Mark Wightman (University of North Carolina, Chapel Hill, NC) has focused on understanding the physiological role of dopamine and related catecholamines in the brain. Wightman has been a leader in the development of electroanalytical methods involving fast scan CV with microelectrodes for detection and quantitation of these compounds. A list of his publications on the subject can be seen by going to the "application" section of www.cypresssystems.com and clicking on Mark Wightman//66-EI400. 3. To computer simulate CV curves for various electrode mechanisms, an online program is available as developed by Professor Vining and linked as ASDL site #005005 titled “Cyclic Voltammetry Simulator.” The simulation program can be downloaded from this site: http://employees.oneonta.edu/viningwj/ ACKNOWLEDGMENT: The assistance of Dr. Richard S. Kelly, Department of Chemistry, East Stroudsburg University, E. Stroudsburg, PA., to this experiment is hereby acknowledged. COMMENTARY: Professor Mark Wightman, UNC, Chapel Hill, NC, has focused on understanding the physiological role of dopamine and related catecholamines in the brain. He has been a leader in the development of Electroanalytical methods involving fast scan CV with microelectrodes to detect and quantify these compounds. Reference #3 gives a website reference to the research of the UNC group. 7