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The Determination of the Rate Constant and Reaction Mechanism for Ru(bpy)32+ Luminescence Quenching
Author: Megan Kraus Lab Partners: Megan Moyer, Marley Pillion, Stefan Kozak Submitted: March 22nd, 2012 (CHEM 457, Section 4) Abstract In this experiment, the basic principles of luminescence spectroscopy and quenching were understood through the calculation of the rate constant of luminescence quenching of ruthenium (II) trisbipyridine, also known as Ru(bpy)32+ by oxygen. Three Ru(bpy)32+ samples of varying oxygen concentration (air-saturated, nitrogen saturated, and oxygen-saturated) were used in this experiment. The laser photolysis of the nitrogen-saturated sample was used to find the fluorescence rate plus the radiationless rate constant, (kf+knr), which was calculated to be
. The quenching rate constant was determined by plotting the concentrations of oxygen with the ratio of the integration areas from emission data of each Ru(bpy)3 sample and
2+

applying the Stern-Volmer equation to find the value of the quenching rate constant, kq, divided by the (kf+knr). The (kf+knr) was previously determined yielding a kq value of The collision rate constant, kd, of water was also calculated and found to be .

Introduction The principles of luminescence spectroscopy are mainly due to light induced chemical changes in photochemical reactions. The chemical change is due to the excitation of a molecules electrons to a higher energy level from the molecules ground state. The ground state of a molecule exists due to all the electrons residing in their lowest energy level and most stable orbital. When the electrons are excited, they return to the more stable ground state in various ways including luminescence, where light is emitted. Ruthenium (II) trisbipyridine (Ru(bpy)32+), was the molecule used in this experiment. The Ru(bpy)32+s electrons absorbed UV light and became excited to a high energy state. The excited electrons then lost their energy and returned to ground state via radiation or radiationless mechanisms.1 Two radiation mechanisms are fluorescence and phosphorescence. These mechanisms occur due to the quantum level interactions of the electrons in the molecule. Electrons in the ground state are opposite spin paired (Pauli Exclusion Principle) and when one becomes excited it can stay opposite and return to ground state (fluorescence) or flip and become parallel then go through a transition excited state before returning to ground state (phosphorescence) shown in the equations below for Ru(bpy)32+.1 Fluorescence Phosphorescence Where the h emitted is not the same h that was used to excite the electrons and is usually the a longer wavelength.2

Radiationless or non-radiative mechanisms are where an electron in a molecule returns to ground state without emitting a photon. One way in which this is done is through quenching. The mechanism of quenching deactivates the excited state by reacting it with a quencher molecule.3 In this experiment, Ru(bpy)32+ is quenched by oxygen. Radiation and radiationless mechanisms have rate constants, k. The quantum yield, , is the ratio of these rate constants where the sum of all the rates. In this experiment, the rate constants were determined by first finding the rate constants of radiative decay, kf+knr, where the excited state decays in the absence of a quencher shown in Eq. 1.
[ ]

equals the rate of formation of the product divided by

)[

] (1)

The solution to Eq. 1 is shown in Eq. 2 below: [ ] [ ] ( ) (2)

Using Eq. 2, a plot of the natural log of fluorescence intensity versus time will yield a graph with a slope of (kf + knr). After the rate constant, (kf + knr) is found, the quenching rate constant can then be determined. The quenching rate is proportional to the concentration of the oxygen present in the sample, shown in Eq. 3 below.
[ ]

][

(3)

The quenching rate constant can be determined using quantum yields of Ru(bpy)3+2 in the absence of a quencher as well as in the presence of a quencher. The quantum yield of Ru(bpy)3+2 in the absence of a quencher is shown below in Eq. 4 and in the presence of a quencher shown in Eq. 5 below.
[ [ ] [ ] ]

(4)

[ [ ] [ ]

] [ ][ ] [ ]

(5)

Using these quantum yields, the Stern-Volmer equation can be used to find a relationship between the kq and (kf+knr) by finding the ratio of Eq. 4 and Eq. 5 shown in Eq. 6. [ ]

(6)

Eq. 6 will be used to calculate the quenching rate by utilizing fluorescence spectroscopy data and concentrations of oxygen in the samples. The collision rate constant will also be found for Ru(bpy)3+2 by using Eq. 7. (7) where R is the universal gas constant, T is the temperature and is the viscosity. With these basic concepts and equations above, the basic principles of luminescence and quenching will be investigated as well as the rate constants determined.

Experimental This procedure was taken from chapter 10 of the CHEM 457 Experimental Physical Chemistry, Spring 2012 Lab Packet.1 An absorption spectrum was taken for a sample of air saturated Ru(bpy)32+ using a Cary 4000 Varian UV-Vis spectrophotometer to find the excitation wavelength of the molecule. Before the Ru(bpy)32+ was analyzed, an air blank was analyzed first. The scan for the cuvette filled with the air saturated Ru(bpy)32+ sample was taken between the wavelengths 250 and 650 nm to yield an excitation wavelength of 450 nm. This excitation wavelength was used for the emission spectra on the Horiba Scientific Fluorolog Spectrofluorimeter. The emission analysis was conducted on air saturated, nitrogen saturated and oxygen saturated Ru(bpy)32+ samples within the range of 500-850 nm. A graph of emission versus wavelength was found for each sample and used to find integrate the area under the curve for data analysis. The nitrogen saturated Ru(bpy)32+ sample was made by bubbling nitrogen through the air saturated cuvette sample for 5 minutes to rid the sample of oxygen. The oxygen saturated sample was made the same way except oxygen was bubbled through instead of nitrogen. Additional analysis was performed on the nitrogen sample with a photolysis apparatus (SRS NL-100). The sample was struck with UV light at a wavelength of 337.1 nm and the fluorescence intensity of Ru(bpy)32+ was collected and sent to a Teletronix TDS 2022B TwoChannel Digital Storage Oscilloscope which converted the data to a graph of intensity versus time.

Results The analysis from the air saturated Ru(bpy)32+ sample from the UV-Vis shows the excitation wavelength at 450 nm. This wavelength was then used for the emission spectra. The graph of the absorbance versus wavelength is shown in Fig. 1.

Absorbance vs. Wavelength

0.6 0.5 Absorbance 0.4 0.3 0.2 0.1 0 350 400 450 500 550 600 650 700 Wavelength (nm)

Fig. 1. The Plot of Absorbance vs. Wavelength to Find an Excitation Wavelength of 450 nm After the excitation wavelength was found, the emission spectrum of the Ru(bpy)3+2 samples (air saturated, nitrogen saturated and oxygen saturated) were taken. The areas under the curve of the emission vs wavelength for each sample of varying concentrations of oxygen were then found using the integration tool. In order to apply the Stern-Volmer equation, a ratio of the quantum yields is needed, therefore the integration areas were made into ratios with respect to a standard area, Areao. Because the nitrogen saturated sample doesnt have any oxygen present, its integration area was the Areao. This area is representative of the total amount of radiative emission because in the absence of oxygen, there are no excited molecules relaxing due to quenching. The concentration of oxygen was also needed for the Stern-Volmer Equation and

calculated using Henrys Law. Fig. 2 illustrates a table that shows the area, area ratio and concentration of O2.
Sample Air Saturated Nitrogen Saturated Oxygen Saturated [O2] (M) 2.68E-04 0 0.00129 Area ratio 1.5297 1 3.4678 Area 6.01E+07 9.19E+07 2.65E+07

Fig. 2. Table of Integration Areas from Emission Curves, Ratio of areas used in SternVolmer Equation and Concentration of Oxygen Using the Stern-Volmer equation, a plot of (Areao/Area) versus the concentration of oxygen yielded a straight line with a slope equal to = shown in Fig. 3.

Stern-Volmer Plot
4 3.5 3 areao/area 2.5 2 1.5 1 0.5 0 0.00E+00 2.00E-04 4.00E-04 6.00E-04 8.00E-04 1.00E-03 1.20E-03 1.40E-03 Concentration of Oxygen (M) y = 1908.4x + 1.0081 R = 0.9999

Fig. 3. Stern-Volmer Plot of (Areao/Area) versus the concentration of oxygen In order to find the quenching rate from the value obtained from the Stern-Volmer Plot, the combined rate of radiative and nonradiative decay was needed to be found. Ru(bpy)3+2,

without the presence of a quencher (oxygen), decayed through radiative and nonradiative mechanisms. The nitrogen saturated sample was used to find these combined rate constants because there was no oxygen present. The plot of the natural log of intensity versus time with a slope equal to -(kf + knr) is shown in Fig. 4.

ln (Intensity) vs. Time

0 -0.5 0 -1 -1.5 -2 -2.5 -3 -3.5 -4 -4.5 -5 0.0000005 0.000001 0.0000015 0.000002

Ln (Intensity)

y = -1.56E+06x - 1.49E+00 R = 9.95E-01

Time (s)

Fig. 4. Plot of the Natural Log of Intensity Versus Time in the Absence of a Quencher (Nitrogen Sample) With the (kf + knr) = was then calculated to be using Eq. 7. now known, the quenching rate constant, kq,
. The rate of collision was also calculated to be

Discussion Although I could not find any literature values for the rate constants, I feel that the results obtained from this experiment seemed reasonable in relation to one another. The results show that the quenching rate is much larger than the combined rate constants of radiative and

nonradiative decay. This comparison confirms that oxygen is a very effective quencher of Ru(bpy)3+2 and therefore when oxygen is present in a Ru(bpy)3+2 solution, it decays the excited state molecules much faster. This also explains why the area below the curve for oxygen saturated Ru(bpy)3+2 decreased greatly compared to the other samples. The significance of the collision rate constant and quenching rate constant is further explained in report question 1 in the appendix of this report. Some sources of error that could have occurred during this lab was during the nitrogen purge. The experiment advised that nitrogen was to bubble through the sample for 5 minutes. This is assuming that five minutes is an adequate amount of time to completely rid the sample of oxygen. One way the lab could have been improved is with the nitrogen and oxygen purging. The method used to purge the samples left much room for oxygen contamination from the surrounding air. A way to improve this would be to use a septum to hold the sample and puncture it with a vent needle. With this method, there is less exposure to the air because you are not required to remove a cap to remove the vent needle. Overall I felt that the lab was successful in confirming the chemistry and concepts behind luminescence spectroscopy. Conclusion The Ru(bpy)3+2 solutions saturated with air, oxygen and nitrogen with varying oxygen concentrations of 0.00027 M, 0.00129 M, and 0.0 M respectively were used to determine the rate constants. With the nitrogen saturated sample, which was void of oxygen, the quencher, the sum

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of the radiative and non radiative rate constants was found to be

. The

fluorescence spectrometry was used to calculate the quantum yield for each of the samples by utilizing the integration area under the curves of the emission scan data. The nitrogen purged sample was used for the standard reference area in the Stern-Volmer plot which was a graph of (Areao/Area) versus oxygen

concentration. The slope of the linear fit was From this, the quenching rate was determined to be collision rate constant was also determined and calculated to be

which is also equal to

. The

. From this

collision rate constant, it was found that the average collision time was shorter than the average quenching time of oxygen from the excited state of Ru(bpy)3+2 confirming that there was successful quenching. Acknowledgements The author would like to acknowledge my lab partners, Marley Pillion, Stefan Kozak and Megan Moyer, for their combined help in performing the procedure, Dr. Milosavljevic and the TAs, Ms. Shivangi Nangia and Mr. Stuart Friesen for their help in understanding the concepts behind the experiment as well as the Pennsylvania State University for allowing us to use the laboratory equipment.

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References 1. Milosavljevic, B.H. Lab Packet for CHEM 457: Experimental Physical Chemistry, Chapter 10. University Press: University Park, 2012. 2. Holler, F.J., Skoog, D.A., Crouch, S.R. Principles Of Instrumental Analysis. 2006 3. OReilly, J.E. Fluorescence Experiments With Quinine. Journal of Chemical Education. 52 (9). 1975 4. Birks, J.B. Excimers. Reports on Progress in Physics. Vol 38. 1975 5. Atkins, P., Paula, J., Friedman, R. Quanta, Matter and Change: A Molecular Approach to Physical Change. W. H. Freeman and Company, New York. 2009 Appendix Report Questions:
1.

The calculated collision rate, kd, using Eq. 7, was found because of its connection to the quenching rate constant. The reciprocal of kq can also be known as the average time it takes for an excited molecule of Ru(bpy)3+2 to be quenched by oxygen. With this being said, the reciprocal of kq is equal to
The reciprocal of kd is equal to which is the amount of

time it takes for two reactants in the solution of water to collide with each other. From the two reciprocals, it is easy to see that the time of the collision between

Ru(bpy)3+2 and O2 is shorter than the time it takes for quenching collision to occur.
2. One major source of error in the calculation of the quenching rate constant

was the atmospheric pressure reading used in Henrys law to find the concentration of oxygen. The atmospheric pressure was found using the

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barometer in the organics lab that is an entire floor below where the samples were being analyzed.
3. There are three common mechanisms for biomolecular quenching of an

excited state molecule. The Forster resonance energy transfer uses a mechanism of dipole-dipole coupling of the excited state molecule and the quenching molecule. The Exciplex mechanism is where the eximer is quenched when the excited state and quencher form a dimer molecule.4 The third mechanism is the Dexter electron transfer mechanism where the overlap of the excited state molecule and donor orbital creates a shorter distance for quenching to occur.5

Sample Calculations with Uncertainty: A. [O2] (Henrys Law)

Atmospheric Pressure = 745.5 mmHg= Percent O2 in the air = 20.95 % Partial Pressure of O2 in atmosphere = (0.2095) ( Henrys constant for O2: Air Sample: Pa) = Pa Pa

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Oxygen Sample: [ ] [ ]

B. Calculation of Quenching Rate Constant, kq with uncertainty

C. Uncertainty: ( ( ) )

D. Kd Calculation

14 Regression Analysis: A. Stern-Volmer Plot: Regression Statistics Multiple R 0.999974 R Square 0.999949 Adjusted R Square 0.999897 Standard Error 0.013159 Observation s 3 ANOVA df Regression Residual Total 1 1 2 Coefficien ts Intercept X Variable 1 1.008065 1908.41 SS 3.375444 0.000173 3.375617 Standard Error 0.010398 13.66911 MS 3.37544 4 0.00017 3 F 19492.3 1 Significan ce F 0.00456

t Stat 96.9491 1 139.614 8

P-value 0.00656 6 0.00456

Lower 95% 0.875948 1734.728

Upper 95% 1.14018 3 2082.09 3

Lower 95.0% 0.87594 8 1734.72 8

Upper 95.0% 1.14018 3 2082.09 3

B. Natural Log of Intensity vs. time: Regression Statistics Multiple R 0.997668 R Square 0.995342 Adjusted R Square 0.995339 Standard Error 0.054806 Observation s 1781 ANOVA df SS MS F Significan ce F

15 1141.78 1 0.00300 4 380131. 4

Regression Residual Total

1 1779 1780 Coefficien ts

1141.781 5.343491 1147.125 Standard Error

t Stat 564.063 616.548

P-value

Lower 95%

Upper 95% 1.4827 6 155239 6

Lower 95.0%

Upper 95.0%

Intercept

-1.48793

0.002638

-1.4931

X Variable 1

-1557350

2525.919

-1562304

-1.4931 1.48276 156230 155239 4 6