Lab 6. Procedure.

Parts of this lab are adapted from Schwenz and Moore¹ and analogous experiment at Dartmouth College².
You will collect data with a lab partner (who should be noted in your write-up). However, all the data analysis and write-up should be done without collaboration.

In this lab you will study quenching of Ru(bpy)₃²⁺phosphorescence by time resolved and steady state measurements of its luminescence. You will compare the two techniques and identify different mechanisms of quenching. Following quenchers will be used: O₂, Fe(H₂O)₆³⁺, Fe(H₂O)₆²⁺ , Fe(CN)₆^3- , and Fe(CN)₆^4- . You and your partner will be given much freedom in the design of the experiment. Please make sure to plot a strategy among yourselves before coming to the laboratory. This simple act of communication and preparation will save valuable time in the laboratory.

Before the experiment

• Record room temperature
• Section A. You should have a few glass pipetts, a 60 mL plastic syringe with a narrow teflon tubing approximately 6 cm long and the two gas tanks - with oxygen and nitrogen, both equipped with a regulator and a purging device as in the picture on the right. The purpose of this purging device is to have either O₂ or N₂ at atmospheric pressure by means of a slow flow of gas through it. Pumping known volume of each gas will provide a measured mixture of the gases.

• Section B. You should have following stock solutions available:
  • 1.8 x 10^-4 M of Ru(bpy)₃²⁺ in water
  • 0.5 M HCl solution
  • 14.8 mM Fe(H₂O)₆³⁺ (made from FeCl₃•6H₂O) in 0.5 M HCl (in order to prevent the formation of ferric hydroxide and other dimeric ferric species)
  • 21.8 mM Fe(H₂O)₆²⁺ (made from FeCl₂•6H₂O) in 0.5 M HCl
  • 11.5 mM Fe(CN)₆^3- (made from K₃Fe(CN)₆) in water
  • 8.8 mM Fe(CN)₆^4- (made from K₄Fe(CN)₆) in water
• record a fluorescence profile of *Ru(bpy)₃²⁺ and absorption spectra for Fe(H₂O)₆³⁺ , Fe(H₂O)₆²⁺ , Fe(CN)₆^3- and Fe(CN)₆^4- at low concentrations (so that absorbance is less than 1 in a 1 cm cuvette, you can use a plastic cuvette for this)
• By mixing the above stock solutions in appropriate proportions, prepare following stock solutions (in 25 mL volumetric flasks) for your measurements (and label them):
  • 9 x 10^-5 M of Ru(bpy)₃²⁺ in water
  • 9 x 10^-5 M of Ru(bpy)₃²⁺ in 0.25 M HCl
  • 9 x 10^-5 M of Ru(bpy)₃²⁺ + 1 mM Fe(H₂O)₆³⁺ in 0.25 M HCl
  • 9 x 10^-5 M of Ru(bpy)₃²⁺ + 10 mM Fe(H₂O)₆²⁺ in 0.25 M HCl
  • 9 x 10^-5 M of Ru(bpy)₃²⁺ + 1 mM Fe(CN)₆^3- in water
  • 9 x 10^-5 M of Ru(bpy)₃²⁺ + 1 mM Fe(CN)₆^4- in water
• For your Stern-Volmer plot, you will need to vary the concentration of a quencher. Using your prelab calculations, make 4 or 5 solutions (~ 5 mL each) with each quencher at different concentrations suitable for the Stern-Volmer plot. Since you measuring the lifetimes, concentration of of Ru(bpy)₃²⁺complex does not have to be constant.
•  

During the experiment

The experimental arrangement of equipment is shown in the Figure below.

The N₂-laser (VSL-337 from LSI) delivers 4 ns pulses at 337 nm of about 100 mJ each. This wavelength is shorter than the maximum of Ru(bpy)₃²⁺ absorption but we can always adjust its concentration to gain absorbance to the desired value around 1. The triggering photodiode (ET 2000 from EG&G) initiates recording the measurements by the detecting photodiode (DET2 from Thor Labs). The signal is collected on a digital oscilloscope (TDS-210 form Tektronix) which is interfaced to a computer. The data from the oscilloscope will be transferred to a computer via serial port using Wavestar program, also from Tektronix. You can review the help file for this program before starting the lab either on the lab computer or by downloading this file wvstar.hlp (warning, its size is 510 k). You will need to use probably 250 ns/division timescale and either 5 mV or 10 mV sensitivity scale on the oscilloscope (correct setup should be stored as a setup 5: Push Save/Recall|Setup scroll to 5|Recall). For running an experiment you need to:

- turn on the Thor photodiode
- turn on the scope (choose setup 5 as described above)
- put your sample into the cuvette holder (you should use a quartz cuvette)
- turn on the laser
- push the RUN button
- when acquisition is complete (takes less than 30 sec), trasfer the kinetic to the computer

Section A. Quenching by O₂

• fill a quartz cuvette approximately 1/2 full with your stock solution of 9 x 10^-5 M of *Ru(bpy)₃²⁺ in water and record its luminescence kinetics using above procedure; store your data; the *Ru(bpy)₃²⁺ lifetime in this solution should be shortened (quenched) in accordance to the natural abundance of O₂ in atmosphere
• purge the solution with N₂ gas by slow bubbling via a glass pipett for 5 min and record the luminescence kinetics; make sure that the solution has reached the room temperature; the Ru(bpy)₃²⁺ lifetime in this solution should correspond to the unquenched lifetime
• now is a tricky part: you have to make three or four measurements at intermediate oxygen concentrations; my suggestion is to use 60 cc syringe
  • fill it with a mixture of the two gases by sucking each gas through the outlet of the corresponding purging device
  • purge the mixture slowly through the solution and put the cap on the cuvette as soon as you finish
  • with the cap on, shake the solution gently for 1 min
  • repeat the procedure with the same gas ratio again at least five times
  • after waiting a few minutes for the solution to warm up, make the lifetime measurements as before
• the highest oxygen concentration will be achieved by purging the solution with pure O₂ gas directly from the tank by slow bubbling via a glass pipett for 5 min; again, make sure that the solution has reached the room temperature; record the luminescence kinetics and measure the lifetime

Section B. Quenching by ions

• record luminescence kinetics of *Ru(bpy)₃²⁺ in water and 0.25 M HCl solutions as before, without deoxygenation; measure the lifetimes - those will be the lifetimes for the Stern-Volmer evaluation of quenching in corresponding ionic solutions
• you already, presumably, made 4 or 5 solutions (~ 5 mL each) with each quencher at different concentrations suitable for the Stern-Volmer plot; record luminescence kinetics for each of them, measure the lifetimes

After you finish

• clean up after yourself
• show you notebook and the setup to the TA or the instructor before you leave

Tentatively, plan to run O₂ quenching on a first day and quenching by ions on the last two days. On a third day, if you have time, you might progress to your own project. It is encouraged that you discuss the project with the instructor, so that appropriate adjustments can be made.

References

1. Physical Chemistry: Developing a Dynamic Curriculum, Ed. R. W. Schwenz and R. J. Moore, 1993, American Chemical Society, Washington, DC.
2. Chem 10 Lab manual, Department of Chemistry, Dartmouth College

Last updated 01/09/06.