Introductory Lab Manual
For Electrogenerated Chemiluminescence
Created By: Amy L. Hruska and Paul L. Walsh
Introduction to ECL: Background Information
Electrogenerated Chemiluminescence, also called electrochemiluminescence and abbreviated as ECL, is the generation of light emitting species from electrochemical oxidation and reduction at an electrodes surface.1 A voltage or voltage pattern applied to the electrode causes electron transfer at the electrode’s surface to form intermediates. These intermediates react to produce an excited state molecule near the electrode. As the excited molecule returns to the ground state, energy is released in the form of light. This can be seen in the following general mechanism for light emission by ECL:1
(1)A + e- A-(reduction potential)
(2)D D+ + e-(oxidation potential)
(3)A- + D+ A* + D (or D* + A)
(4)A* (or D*) A (or D) + hv.
In this scheme, A is an electron acceptor, which is reduced at the electrode’s surface, and D is an electron donor, which is oxidized at the electrode’s surface. A square wave potential pattern is applied to the electrode with voltages chosen to reduce A and oxidize D. Note that the electron donor and acceptor may be a different molecule or the same molecule.
ECL emission has been shown to occur by three main types of reaction schemes, which fall into two larger groups: annihilation ECL and co-reactant ECL.1 The more specific relations are called the S- Route and T-Route systems (annihilation), and oxidative-reductive or reductive-oxidative systems (co-reactant). In the S-Route, or energy sufficient system, the change in enthalpy generated in creating the excited species that exhibits emission is greater than the change in energy necessary to create the singlet species from the ground state.1,2 Unlike the other ECL mechanisms, once the oxidized and reduced species of the luminophores react by the S-Route, the emitting species is created and has sufficient energy to emit a photon. A common example of a luminophore which undergoes ECL by this mechanism is tris(2,2’-bipyridyl) ruthenium (II) dichloride hexahydrate (Ru(bpy)32+) system. The reaction scheme for the generation of ECL emission for this system is
(5)Ru(bpy)32+ - e- Ru(bpy)33+E= +1.2V vs SCE
(6)Ru(bpy)32+ + e- Ru(bpy)3+ E= -1.4V vs SCE
(7)Ru(bpy)33+ + Ru(bpy)3+ Ru(bpy)32+ * + Ru(bpy)3
(8)Ru(bpy)32+* Ru(bpy)32+ + hv (~2.1eV, 610nm).
Another example of a luminophore that undergoes ECL by the S-Route mechanism is 9,10-dephenylanthracene (DPA).1,3 The reaction scheme that DPA follows is
(9)DPA + e- DPA·- E = -1.6 V vs. Ag
(10)DPA DPA·+ + e- E = +1.2 V vs. Ag
(11)DPA·+ + DPA·- DPA* + DPA
(12)DPA* DPA + hv.
Above, the DPA·- and DPA·+ represent the reduced and oxidized free radical species, respectively, and DPA* represents the excited species undergoing ECL emission. The oxidized and reduced species are created at the surface of the electrode by applying a square wave that cycles between the two potentials necessary to make DPA·- and DPA·+.3,4 The cycling of potentials to create both a reduced and an oxidized species is true of all S-Route systems. These two radicals then go on to react and create the excited emitting species that fall to the ground state as emitted light.
While the S-Route is an “energy sufficient” system, some systems are known as “energy deficient.” An energy deficient system proceeds by a slightly different mechanism known as the T-Route or triplet-triplet annihilation pathway. The T-Route mechanism gets its name because there are triplet states which are formed during the oxidation and reduction step that must undergo an annihilation reaction to create the singlet excited species.1 The singlet state varies from the triplet state because the electrons have paired spins in the singlet state (one spin up, the other spin down), while they are the same spin in the triplet state. In the T-Route mechanism, Equation 4 is replaced with the following emissive pathway:
(13)3A* + 3A* (or 3D* + 3D*) 1A* + 1A* (or 1D* + 1D*) + hv
(14)1A* (or 1D*) A (or D) + hv.
Equation 3 from the scheme above only provides enough energy to excite the luminophore into the first excited triplet state. This excited state may undergo non-radiative decay back to the ground state, or it may react via Equation 13 to produce an excited singlet state of the luminophore. This excited single state will then return to the ground state with the emission of light. . DPA reacts with N,N,N’,N’-tetramethyl-p-phenylenediamine (TMPD) via the T-Route and ECL with tris-(2,2’-bipyridine)-ruthenium(II) demonstrates another example.
(15)DPA + e- DPA·- E = -1.89 V vs. SCE
(16)TMPD TMPD·+ + e- E = +0.24 V vs. SCE
(17)DPA·- + TMPD·+3DPA* + TMPD
(18)3DPA* + 3DPA* 1DPA* + DPA
(19)1DPA* DPA + hv.
As shown above, the oxidized and reduced species react to form a triplet excited species in the T-Route system that reacts with another triplet to form an excited species in the singlet state; this excited singlet molecule has sufficient energy to undergo emission down to its ground state. Similar to the S-Route system, a square potential waveform is applied to the surface of the electrode in order to create both the oxidized and reduced species.1 Also, it is important to note that many reactions proceed by both the T-Route and S-Route mechanisms to varying degrees.
ECL may also be achieved at a single potential by the oxidative-reductive system. In this system, a co-reactant forms a strong reducing agent upon oxidation at the electrode. This reducing agent then reacts with the luminophore to form an excited state. This system is the most complicated because, instead of having a singlet reactant, the primary emitting species reacts with a co-reactant to form the emitting species.1 An example of this system is Ru(bpy) 32+ and tripropylamine (TPrA). The mechanism for it is
(20)Ru(bpy) 32+ Ru(bpy) 33+ + e-
(21)TPrA [TPrA]+ + e-
(22)[TPrA]+ + e- TPrA + H+
(23)Ru(bpy) 33+ + TPrA Ru(bpy) 32+* + Products
(24)Ru(bpy) 32+* Ru(bpy) 32+ + hv.
Both the primary emitting species (Ru(bpy)32+) and the co-reactant (TPrA) are oxidized at the surface of the electrode while applying a single potential, hence the reaction scheme name of oxidative-reductive.1 If the primary emitting species and co-reactant are reduced at the surface of the electrode, the system is called a reductive-oxidative system. The oxidized TPrA then undergoes de-protonation and becomes a highly reactive free radical. The oxidized Ru(bpy)32+ reacts with the tripropylamine radical to form the excited species of Ru(bpy)32+* which decays to its ground state by emitting a photon. The potential at which the co-reactant and primary emitting species oxidize must be very close to one another so that when a single potential is applied to the surface of the electrode both oxidations can occur simultaneously. Since Ru(bpy) 32+ is one of the very few ECL emitting compounds which have an oxidation potential within the solvent window for water, when combined with TPrA as a co-reactant, chemiluminescence is made possible at a single potential applied in aqueous solutions (the potentials are between water’s oxidation, +1.2V, and reduction, -1.2V which is why water can be used as a solvent).5
References
1.Richter, Mark M. Electrochemiluminescence. Chemical Reviews, 2003, A – AH.
2.Faulkner, L. R.; Glass, R. S. Electrogenerated Chemiluminescence from the Tris(2,2’-
bipyridine)ruthenium(II) System. An Example of S-Route Behavior. J. Phys. Chem.
1981, 85, 1160-1165.
3.Bard, A. J.; Maloy, J. T. Electrogenerated Chemiluminescence. VI. Studies of the
Efficiency and Mechanisms of 9,10-Diphenylanthracene, Rubene, and Pyrene Systems at
a Rotating-Ring-Disk Electrode. J. Am. Chem. Soc.1971, 93, 5968-5981.
4.Wightman, R. M.; Collinson, M. M. High-Frequency Generation of
Electrochemiluminescence at Microelectrodes. Anal. Chem.1993, 65, 2576-2582.
5.Harvey, David. Modern Analytical Chemistry. McGraw Hill, Boston, 2000, Chapter 11.
Introduction to ECL Experiment #1: Electronics Review
Purpose:
This experiment seeks to review the concepts of electronics covered in University Physics as well as to introduce new researchers to some more advanced electronics concepts and materials not covered in prerequisite courses.
Materials:
Breadboard
Protoboard
Wiring kit
3 – Resistors
1 – 10 kΩ Variable Resistor
Multimeter
1 - 9V batteries with connecting ports
Background:
The use of electronics in the ECL lab is of the utmost importance. Not only is it necessary for the generation of ECL emission, but it is also important for truly understanding how to take the measurements of the ECL emission. Electronics has become an integral part of analytical chemistry over the past 20 years with the advent of more specialized and sensitive instrumentation. It is therefore necessary for all researchers to have an excellent background in the subject beyond what is covered in the introductory physics classes. Before beginning this experiment, make sure to review the basic concepts covered in those introductory physics classes such as University Physics. Topics include: electric fields, electric potential differences, basic circuits, Ohm’s Law, Kirchoff’s Current Law, Kirchoff’s Voltage Law, AC versus DC power sources, resistors in series and parallel, oscilloscopes and function generators.
Procedure:
1. Study the multimeter provided. Make sure that you understand what each setting measures, and how to attach the leads to make the correct measurements. If there is something you are unsure of, make sure to ask a fellow researcher or your instructor. Connect the leads to make a resistance measurement and set the multimeter to do the same. Take three different resistors and look up their resistance by the color code. Record the order of the colors and the corresponding resistance. Now measure the resistance of each of the resistors with the multimeter and record your results.
2. Next, pick up the breadboard and study it. If there is an instruction manual for it, read through it. On the breadboard, there are three knobs to one side. One is black and the other two are red. The black knob is for connecting to electrical ground. Connecting to ground is much like completing a circuit. It gives the electric current a place to flow, and if two ends of a circuit are both attached to ground, it is the same as making a loop circuit like those introduced in University Physics. The two red ports can be used if a common potential is to be applied throughout the circuit. Along the two long sides of the breadboard, there are 4 sets of holes stretching the length of the board. These are called power buses because each of the rows are connected the length of the board. This allows for a common ground or potential to be applied to any part of the board you want. In order to test to see how the various ports are connected, place wires in various places and see where no resistance is measured. If the resistance between 2 ports is about zero ohms, then they are connected. Make a sketch of the breadboard and label which parts are connected in a way that makes sense to you in the space provided.
3. First, measure the voltage of the battery. Then make a series connection between the three resistors and one 9 V battery. Make sure that you have at least two 10 kΩ resistors before connecting the batteries to the resistors. Next, measure the voltage drop across each of the resistors and measure the current between each of the resistors as well as between the two end resistors and the battery. Also, measure the voltage drop across all three resistors, and the current before each of the resistors and after all the resistors. Record all measurements taken in the space provided. Use the circuit diagram below to help you in creating the circuit.
4. Next, create the same circuit, but instead of creating a complete circuit, use ground as your area of low potential to allow the current to flow there. Measure the voltage drop across each of the resistors and measure the current between each of the resistors as well as between the two end resistors and the battery. Also, measure the voltage drop across all three resistors, and the current before each of the resistors and after all the resistors. Record all measurements taken in the space provided. Use the circuit diagram below to help you in creating the circuit.
5. Then connect the three resistors in parallel with one another. Measure the voltage drop across each of the resistors and measure the current between each of the resistors as well as between the two end resistors and the battery. Also, measure the voltage drop across all three resistors, and the current before each of the resistors and after all the resistors. Record all measurements taken in the space provided. Use the circuit diagram below to help you in creating the circuit.
6. Then create the same circuit using ground as the area of low potential. Measure the voltage drop across each of the resistors and measure the current between each of the resistors as well as between the two end resistors and the battery. Also, measure the voltage drop across all three resistors, and the current before each of the resistors and after all the resistors. Record all measurements taken in the space provided. Use the circuit diagram below to help you in creating the circuit.
7. Once all the measurements of the series and parallel circuits have been completed, create the series circuit from Step 4, except replace one of the resistors with a variable resistor. Make sure you connect the correct lead to ground. If you are unsure, ask a fellow researcher or the instructor. Measure the resistance of the variable resistor when the knob is turned completely counter-clockwise as well as completely clockwise. Next, measure the voltage drop across the resistor when the knob is turned completely counter-clockwise. Also measure the voltage drop across the resistor when the knob is turned ¼, ½, ¾ of its full rotation as well as the entire rotation clockwise. Also measure the resistance of the variable resistor. Record all measurements taken in the space provided.
8. Take the Proto-Board given and study it. If there is a user’s manual for it, read through it. Along the top of the board, there is a ground connection much like on the Breadboard. The other three red terminals are labeled +5V, +15V, and -15V. These are ports to which you can make connections to supply these voltages to the circuit. There is also a power switch and an AC power cord on the Proto-Board which are necessary to supply the voltages to the terminals. There are power busses and terminals just as on the Breadboard. Make connections and test the resistance between various ports to test which are connected and which are not. Make a schematic sketch of the Proto-Board in the space provided.
9. Now connect the +5V port to one of the terminals on the Proto-Board and connect the 10 kΩ variable resistor in series with a 5 kΩ resistor. Measure the potential at the end of the circuit versus ground for when the variable resistor is turned completely counter-clockwise, as well as ¼, ½, ¾ of the way clockwise, and completely clockwise. Also be sure to measure the resistance of the variable resistor. Record the data in the space provided.
Data/ Observations:
1. Give the color code of each of the resistors provided, its expected resistance, and the actual resistance measured by the multimeter.
2. Draw the Breadboard, and schematically show how the terminals are connected to one another, including the power buses.
3. What is the true voltage of the 9 V battery?
4. Report the values for the voltage drops, and current for the two series circuits.
5.Report the values for the voltage drops and current for the two parallel circuits.
6. Record the voltage drop across the variable resistor in the series circuit when the knob is turned completely counterclockwise, and about 1/4, 1/2, 3/4, and all of the way clockwise.
7.Draw a schematic representation of the Proto Board and show how the various ports are interconnected.
8. Record the voltage drop across the entire circuit made in Step 9 when the variable resistor is completely clockwise, as well as 1/4, 1/2, 3/4, and entirely clockwise. Also, record the resistance of the variable resistor.
Calculations/ Questions:
1. Using the measured values for the 9V battery and the resistors, calculate the theoretical values for the current (I) and the voltage drops across each of the resistors for the series loop circuit. Calculate percent errors for each of the values as well and report them below. What are the possible sources of error? Do Ohm’s Law and Kirchoff’s Voltage Law remain true?
2.Using the measured values for the 9V batter and the resistors, calculate the theoretical values for the current (I) and the voltage drops across each of the resistors for the parallel loop circuit. Calculate percent errors for each of the values as well and report them below. What are the possible sources of error? Do Ohm’s Law and Kirchoff’s Current Law remain true?
3.Are there any differences between the values obtained with the loop circuits versus the circuits which use ground?
4. What might the circuit created in Step 9 of the procedure be used for?
Introduction to ECL Experiment #2: Cyclic Voltammetry
Purpose:
The purpose of this experiment is to give the student an understanding of the theory and method of cyclic voltammetry. This will be accomplished by observing the effect of scan rate, concentration, and supporting electrolyte on the CV scan.
Materials:
BAS CV-1B Voltammograph
Houston Instruments Omniographic 100 X-Y Recorder and pen
Multimeter
Lined graph paper
12 in ruler
4 - 25 mL volumetric flask
1 – 100 mL volumetric flask
1 – 200 mL volumetric flask
5 - 50 mL beaker
2 mm Pt working electrode
Pt auxiliary electrode
Ag/AgCl reference electrode
Chemicals:
Analytical Reagent Grade KNO3
Reagent Grade K3Fe(CN)6
Background:
Before beginning this laboratory, be sure to read the section on cyclic voltammetry in your Analytical textbook. Only the highlights will be covered in this laboratory. Cyclic voltammetry (CV) is an analytical technique used to detect electrochemically active species in solution. CV is a little different from the techniques that are learned in General Chemistry and Analytical Chemistry in that it is performed on single solutions as opposed to two solution systems. When two solutions, which have the correct reduction potentials, are attached by a circuit and salt bride, the REDOX reactions occur creating a current through the wire, and therefore a potential difference is also created between the two solutions. In a single solution sample, a potential is applied to the various electrodes in the solution to make electrochemically reversible reactions occur.