From Light Harvesting to Intracellular Oxygen

Sensing: Chromophores and Their Novel Applications

in Photovoltaics and Molecular Cardiobiology

by

Rachael A. Carlisle

A dissertation submitted to The Johns Hopkins University in conformity

with the requirements for the degree of Doctor of Philosophy.

Baltimore, Maryland

October, 2007

UMI Number: 3288436

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Abstract

Chromophores, organic and inorganic molecules containing light absorbing moieties,

have been utilized throughout the scientific community. In the present work, derivatives

of the aromatic pyrene moiety and of the Ruthenium centered poylpyridyl complexes

have been used to sensitize nanocrystalline (anatase), mesoporous TiC^ and ZrC>2 thin

films. Electrical and energetic properties of these dyes were investigated in solutions as

well as on the thin film substrates. In a biological application, Ru(bpy)32+, tris (2,2'-

bipyridine) ruthenium(II) dichloride, a well characterized divalent ruthenium polypyridyl

dye, was used to measure the intracellular pC>2 of individual guinea pig ventricular

cardiomyocytes.

Chromophore-linker bipodal systems were used to anchor pyrene or ruthenium

polypyridyl complexes to the surface of T1O2 (anatase) and ZrC>2 nanoparticle thin films

in hopes of increasing the coupling and conjugation to the surface. This in turn would

improve electron injection into TiC^ and therefore produce more efficient dye- sensitized

solar cells. The properties of each dye varied with varying the spacer length and were

therefore extensively studied.

In addition to the bipodal rigid rod linkers, a pyrene "tripodal" system was adsorbed to

the metal oxide surfaces in a perpendicular orientation, preventing the possible pivoting

action of the rigid bipodal systems. Furthermore, this pyrene tripod derivative, with it's

large "footprint," helped to minimize sensitizer-sensitizer interactions when bound to the

surface.

ii

Chromophores have proven to be invaluable for efficient light harvesting energy for the

advancement of photovoltaics; however, many of these dyes, due to their tuneable

photophysical properties and energetics, have been useful in biologic and medicinal

research.

The balance of energy supply and demand is a crucial determinant of cardiovascular

health, but the mechanisms that regulate oxidative phosphorylation are still poorly

understood. While oxygen consumption (V02) can be measured in cell suspensions or

tissues, measurements of respiratory flux in single cells have proven challenging. Here,

we report a novel method for monitoring intracellular oxygen tension using a ruthenium

2+ 2+

polypyridyl complex, Ru(bpy)3 • Ru(bpy)3 was introduced into adult guinea pig

2+

cardiac cells via the whole cell patch clamp method, and Ru(bpy)3 photoluminescence,

mitochondrial redox potential, and sarcolemmal K,ATP currents were monitored

simultaneously as indices of the cellular metabolic status. Application of this method to

models of cardiovascular disease will provide novel insights into the role of

mitochondrial dysfunction under pathological conditions.

This work was performed under the advisement of Professor Gerald J. Meyer, PhD,

Department of Chemistry and Professor Brian O'Rourke, PhD, Institute of Molecular

Cardiobiology. This thesis was read by Prof. Brian O'Rourke, PhD, Prof. Gordon

Tomaselli, MD, Prof. John Toscano, PhD, and Prof. Tamara Hendrickson, PhD. The

tripodal/rigid-rod sensitizer work was performed in collaboration with Elena Galoppini

and co-workers at Rutgers University. Advisor: Dr. Brian O'Rourke

i i i

Acknowledgements

This dissertation is a written document of six years of research as a graduate student in

the Department of Chemistry. My success is not my accomplishment alone but is truly

an accomplishment of the many professors, doctors, collaborators, colleagues, friends,

and family that have supported me over the years.

Foremost, I would like to thank my thesis advisor, Dr. Brian O'Rourke for his patience,

support, long hours at the patch-clamp set-up and his humor. I never imagined I would

have been given the opportunity to combine two amazing fields, chemistry and

cardiobiology, to produce a primary thesis project. His immense knowledge in his field

has allowed for this project to become a success.

As a diligent young high school student, I was so intrigued by science; chemistry allowed

me to find answers to that never ending question "Why?" I always strived to attain more

knowledge of chemistry, and through extra out-of-class study with one person in

particular, I fell in love with the subject. I owe my fascination and success with

chemistry to Dr. Wilson, a great mentor, family friend, and dedicated Washington

Redskins fan.

Throughout my collegiate years at James Madison University, I realized how challenging

chemistry really was. I believe a great professor is not only one who can teach what

they know but also one who shows encouragement and applauds success. Dr. Frank

iv

Paloscay, Dr. J.J. Leary, and Dr. Brain Augustine are outstanding professors and will

always be remembered as having a hand in my success.

I would also like to thank the chemistry professors with whom I have had the privilege of

interacting over the last six years at Johns Hopkins. In particular, Dr. David Yarkony, for

whom I had the responsibility as a teacher's assistant, has been a great mentor. I

immensely appreciate his arranging for me to become part of the faculty in the Chemistry

Department at Villa Julie College and his support.

As a lecturer, I was able to work closely with Dr. Ellen Roskes, Department Chair. She

always showed compassion for her students and faculty which helped us succeed and

further develop as chemists.

I would like to thank my committee members, Dr. John Toscano, Dr. Tamara

Hendrickson, and Dr. Gordon Tomaselli for their guidance and input. Their suggestions

and scientific knowledge have helped to progress this research project.

I learned a great deal from members of my research group in the chemistry department;

Dr. Paul Hoertz and Dr. Bryan Bergeron have taught me a great deal regarding solar cell

chemistry and photovoltaics. Paul's dedication and passion for the science and as a

researcher was inspiring. I would also like to thank other members of the group for their

friendship and collaborations: Dr. Laura Bauer, Dr. Georg Hasslemann, Dr. Feng Liu, Dr.

Don Scaltrino, Dr. Nira Birnbaum, Dr. David Watson, Dr. Sherine O'bare, Dr. Arnold

v

Stux, Dr. Chris Clark, Dr. Tamae Ito, Dr. Jonathon Stromberg, Dr. Gerard Higgins,

Aaron Staniszewski, Hailong Xia, and Diane Wong-Verelle. Lastly, I would like to

thank Dr. Gerald J. Meyer for his advising and support.

I was welcomed into an outstanding group within the Institute of Molecular

Cardiobiology at the Johns Hopkins Medical Institute. Dr. Sonia Cortassa and Dr.

Miguel Aon are extremely knowledgeable and their input is greatly treasured. Dr.

Agnieszka Sidor, Dr. Anand Ganesan, Dr. Ting Liu, Dr. David Brown, Dr. Maria Celeste

Villa-Abrille, Dr. Lufang Xhou, Nicolas Sorarrain, Roger Ortines, Alice Ho, and Dr.

Brian Foster have each contributed to this dissertation through their valuable discussions.

Friends have always been a large part of my life and my success. I owe so much to those

who have kept me sane through the struggles of graduate school and who have celebrated

in my achievements along the way. A special thanks to Dr. Andras Marton, my favorite

Hungarian, lab-mate and dance partner. I would like to thank Dr. Sandra Sinishtaj and

Amanda Fond for all of the fun, laughter, tears, and gossip we have shared in graduate

school. I would especially like to thank my JMU girls: Stacey Barry, Nikki Hayden,

Melissa Moloney, Meghan Schablik, and Melissa Gilbert for always being there, even if

it is a phone call away; I have learned important life lessons from each of you and will

always cherish your friendship.

Lastly, I would like to thank those that have contributed the most to my success. My

parents have always put their children and our education as their highest priority. My

vi

parents have taught me to respect others, to aspire to be the best, have given me direction,

have offered me advice; but the most important thing of all I have learned is to love life.

I have always tried to emulate them, because they are the people I respect most in life.

Thank you for all of your support (not just financial). I also want to thank my sister

Catherine, who, despite her younger age and obsession with Walmart, is an amazing role

model and such an admirable person. Finally, I would like to thank my fiance, David, to

whom this dissertation is dedicated, for your love, patience, and understanding over the

years. I look forward to this next year and the many others to come.

vii

Table of Contents

Title i

Abstract ii

Acknowledgements iv

List of Tables xi

List of Schemes xii

List of Figures xiii

Chapter 1. Introduction to Organic and Inorganic Chromophores

1.1. Molecular Orbitals and Electronic Properties of Chromophores.

4

5

9

10

11

12

Chapter 2. Introduction to Light Harvesting Chromophores for Applications in

Photovoltaics

2.1. The Features of the Solar Spectrum. 14

2.2. Semiconductor Physics and Device Fabrication. 16

2.3. Dyes for Light Harvesting Energy. 17

2.4. Dye Sensitized Solar Cells (DSSCs). 19

2.5. References. 24

1.2.

1.3.

1.4.

1.5.

1.6.

1.7.

Chromophore Absorption.

Chromophore Emission.

Quantum Yields.

Lifetimes.

Conclusion.

References

viii

3.1.

3.2.

3.3.

3.4.

3.5.

3.6.

Introduction.

Experimental Section.

Results.

Discussion.

Conclusion.

References.

Chapter 3. Organic Rigid-Rod Linkers for Coupling Chromophores to Metal-

Oxide Surfaces

25

28

32

42

46

48

Chapter 4. Control of Intermolecular "Cross-Talk" between Chromophores on

Nanocrystalline Surfaces through the Isolation of Pyrene Rigid-Rod

Sensitizers

52

58

59

65

68

71

Chapter 5. Excited State Electron Transfer from Ru(II) Polypyridyl Complexes

Anchored to Nanocrystalline Ti02 through Rigid-Rod Linkers

75

80

85

104

112

114

4.1.

4.2.

4.3.

4.4.

4.5.

4.6.

Introduction.

Experimental Section.

Results.

Discussion.

Conclusions.

References.

5.1.

5.2.

5.3.

5.4.

5.5.

5.6.

Introduction.

Experimental Section.

Results.

Discussion.

Conclusions.

References.

IX

Chapter 6. Measurement of Oxygen Tension in Single Cardiomyocytes: 2+

Photoluminescence Quenching of Intracellular Ru(bpy)3 by Molecular Oxygen

120

125

130

150

154

155

APPENDIX 1: CURRICULUM VITAE 159

6.1.

6.2.

6.3.

6.4.

6.5.

6.6.

Introduction.

Experimental Section.

Results.

Discussion.

Conclusion.

References.

X

List of Tables

Table 3.1. Photophysical Properties of Pyrene Sensitizers. 36

Table 3.2. Electrochemical Properties of Pyrene Sensitizers. 40

Table 4.1. Photophysical Properties of Py-E-Ph-TRIPOD in solution and on 62

the surface.

Table 5.1. Surface Adduct Formation Constants and Limiting Surface 86

Coverages for Selected Ruthenium Sensitizers.

Table 5.2. Electrochemical Data for 1-3 and Reference Compounds in 92

Solution.

Table 5.3. Photophysical Properties of 1-5 and Other Ru Complexes in 94

CH3CN Solutions.

Table 5.4. Excited State Electron Yields, <t>inj, for Rigid Rods Bound to Ti02 103

Films Pretreated with Acid or Base.

Table 6.1. Photophysical Properties of Ru(bpy)32+ in Physiological 132

Tyrode's Solution at Room Temperature.

xi

List of Schemes

Scheme 6.1. The dynamic (collisional quenching) mechanism for Ru(bpy)3 126 by molecular oxygen. Once the Ru(bpy)32+ is excited by incident photons (h v), the excited state can either undergo A) a first-order PL decay (k~l) B) or quenching by colliding with triplet molecular oxygen. The latter leads to C) a bi-exponential decay (&'"') and a decreased lifetime (r ') .

xii

List of Figures

Figure 1.1. A generalized Jablonski-type diagram depicting the potential 3

energies of the HOMO and LUMO electrons. Absorbance of a

photon, fluorescence, phosphorescence, intersystem crossing

(&iSC), and vibrational cascade (kyc) are also illustrated.

Figure 1.2. Jablonski diagrams depicting different forms of radiative decays 8

of excited state electrons: a) High energy fluorescence from the

singlet excited state, S*, to the singlet ground state, S0. b) Lower

energy phosphorescence from the well-defined, lower-lying

triplet excited state, c) Lower energy photoluminescence from

the overlapping singlet-triplet excited state. Non-radiative decays

are also represented, km.

Figure 2.1. The solar spectral irradiance distribution (AM 1.5). This figure 15

represents the intensity of energies of the electromagnetic

spectrum that penetrate the atmosphere and reach the Earth's

surface.

Figure 2.2. Molecular structure of chlorophyll. This natural dye contains a 18

metal- centered porphyrin head which serves as the chromophore

moiety. The conjugated 7i-system is apparent in the structure.

The long hydrocarbon tail acts to anchor the chlorophyll in the

thylakoid membrane.

Figure 2.3. Regenerative Dye Sensitized Solar Cell (DSSC). After the 2 0

chromophore (sensitizer) absorbs a photon, an electron donated

from the excited state to the mesoporous semiconductor Ti02

nanoparticles. The injected electron then travels through the

xiii

mesoporous thin film to the fluorine-doped tin oxide (FTO) glass

substrate where it then proceeds through the electron circuit. The

oxidized form of the redox couple regenerates the ground state of

the sensitizer.

Figure 2.4. The Mechanistic Scheme of the Regenerative Dye Sensitized 21

Solar Cell (DSSC). The ground state sensitizer, S0, absorbs the

incident photon and promotes it to the excited state, S*. The

energy of the incident photon must be greater than or equal to

that of the energy gap between these two energy levels. The

photons are not absorbed by the FTO or the Ti02 because both

require energy of ultraviolet photons for absorption. The

electron can either undergo injection, kmj, or emission. The

injected electrons are then collected at the FTO and the external

circuit is utilized or recombination occurs, kKC, and regeneration

cannot be completed. Eg represents the band gap of the Ti02

semiconductor.

Figure 3.1. Schematic of the five bipodal pyrene-spacer-anchor systems 27

compared in this study. The phenylenethylynene spacer is

abbreviated (E-Ph).

Figure 3.2. Absorption spectra of (a) 1-3 in neat CH3CN and (b) 1-3 33

adsorbed to Ti02 in neat CH3CN. For both (a) and (b), solid

lines correspond to 1, dashed lines correspond to 2, and dashed-

dot lines correspond to 3.

Figure 3.3. Absorption spectra of 2-5 (a) in THF solution and (b) bound to 34

Ti02/glass.

xiv

Figure 3.4. a) Normalized emission spectra of Py-(E-Ph)2-Rod as a function 38

of surface coverage on mesoporous Zr02 nanoparticle thin films

immersed in acetonitrile at room temperature. The surface

coverage from left to right was: 0.068, 0.16, 0.22, 0.25, and 1.6 x

10" mol/cm . Superimposed on the data is the emission

spectrum (+) of a saturated ethylene glycol solution of Py-(E-

Ph)2-Rod. b) The green emission (left vial) represents the

excimer emission of Py-(E-Ph)2-Rod in an ethylene glycol

solution, while the Py-(E-Ph)2-Rod in neat acetonitrile,

representing the violet monomer emission. The samples were

excited with 350 nm light.

Figure 3.5. tsA spectrum observed after 417-nm laser light excitation (~3.1 41

mJ/cm2, 8 ns fwhm) of Py-(E-Ph)2-Rod/Ti02 immersed in

CH3CN. The data were recorded at 10 ns, 100 ns, 250 ns, and 1

u,s delays after the laser pulse. Overlaid is the normalized

absorption difference spectrum obtained 10 \xs after sensitizing

the triplet of Py-(E-Ph)2-Rod (4.0 x 10"5 M) in CH3CN using

[Ru(bpy)3](PF6)2 (4.0 x 10"5 M) and exciting with 532-nm laser

light (-11 mJ/cm2, 8-ns fwhm). Inset: Transient absorption

signals following 417-nm laser light excitation of Py-(E-Ph)2-

Rod/Ti02 immersed in 1.0 M LiC104/CH3CN monitored at 640

nm and displayed on a logarithmic time scale from 10"8 to 0.5 s.

Overlaid on the data is a kinetic fit (white line) to a sum of two

second-order rate constants.

Figure 3.6. The LUMO (a) and the HOMO (b) orbitals of the pyrene 4 4

chromophore in 1 - 3 derived from semi empirical calculations.

Note the pronounced derealization onto the rigid rod in 2 and 3.

xv

Figure 4.1. Schematic of the bipodal and tripodal linkers bound to metal 53

oxide (MO) nanoparticle surfaces. The linkers consist of a p-

phenyleneethynylene spacer of varying length, connecting the

chromophore to the anchoring group, a) Depicts the tripodal

linker with three COOH groups for attachment, b) Represents the

bipodal linker with two COOH groups for attachment.

Figure 4.2. The tripodal pyrene - spacer - anchoring system. The 54

phenylenethynylene spacer is abbreviated (E-Ph). The ball-and-

stick model illustrates the footprint size from a top-down view.

* footprint-size calculated by Galoppini and coworkers.

Figure 4.3. Three plausible techniques for eliminating excimer formation: 57

the addition of bulky substituents to the chromophore head

group, dilution of the sensitizer on the surface through the co-

adsorption of lengthy hydrocarbon chains, synthetically altering

the footprint size of the sensitizer through the addition of a three

point linker.

Figure 4.4. a) The extinction coefficient of the pyrene bipods 1-3 (discussed 60

in Chapter 3) and of the Pyrene-E-Ph-TRIPOD. The Tripod is

comparable in extinction coefficient and in the red-shift of the

absorbance band to that of the Py-E-Ph-ROD. b) The absorbance

spectra of pyrene bipods 1-3 and of the Pyrene-(E-Ph)-TRIPOD

on Ti02. The fundamental absorption band of the Ti02 is

obscured by the absorption of the sensitizers 2-3 and the tripod.

Figure 4.5. The structures absorbance and emission spectra of the Pyrene-E- 63

Ph-TRIPOD in CH3CN.

xvi

Figure 4.6. a) Emission dependence on surface coverage of the Py-(E-Ph)- 64

TRIPOD on Z1O2. As the surface coverage increases, the

monomer grows in and the excimer emission remains relatively

the same, b) A plot depicting the ratio of fluorescence intensity

of the monomer : excimer.

Figure 4.7. A schematic depicting a mesoporous nanoparticle Ti02 thin film 67

sensitized with Py-(E-Ph)-TRIPOD. The red arrows illustrate the

necking regions where the rc-stacking may occur by neighboring

sensitizer molecules, acting as the source of the expected

excimer.

Figure 4.8. Py-(E-Ph)-TRIPOD on Ti02 in the presence of neat CH3CN and 69

in the presence of 1.0M LiCLO^CHaCN. The cation affect

contributes to electron injection into the TiC>2 depicted by a

decrease in the emission intensity in the presence of LiC104.

Figure 5.1. Illustration of rigid-rod and tripodal linkers bound to metal oxide 77

(MO) nanoparticles' surface consisting of a bridge (b) of variable

length, anchoring groups (A), and a sensitizing dye (S). Similar

to that of the organic rigid-rod systems (Figure 4.1).

Figure 5.2. Structure of rigid-rod complexes and two reference complexes.

The rigid-rod linkers are made of (Ph-E)„ units and are 79

terminated with two methyl esters (COOMe) as the anchoring

groups (a). The number after ROD in the abbreviated name refers

to the number of Ph-E units in the linker. The counterion was

PF6_ in all cases. *These Ru derivatives were synthesized by

Elena Galoppini and co-workers at Rutgers University.

xvii

Figure 5.3. Surface adduct formation shown as plots of [Run]eq/r with 88

overlaid linear fits for the adsorption [Ru(bpy)2bpy-(EPh)2-

ROD]2+, [Ru(4,4'-Cl2-bpy)2bpy-(EPh)2-ROD]2+, [Ru(4,4'-Cl2.

bpy)2bpy]2+ to nanocrystalline Ti02.

Figure 5.4. a) Raman spectra of ester la (-) as a solid and la on Ti02 ( ). 89

The Ti02 film was cast on sapphire and was pretreated with acid,

b) IR spectra of the carboxylic acid prepared from ester la (-) on

a Ge and la on Ti02 ( ). The Ti02 film was cast on Ge and

was pretreated with acid.

Figure 5.5. Attenuated total reflectance IR spectra for 2b on Ti02 thin films

that were pretreated at pH = 1 and pH = 11.

90

Figure 5.6. Ground state absorbance spectra in acetonitrile a) Spectra of la, 96

lb, and lc. b) Spectra of 2a, 2b, and [Ru(4,4'-(Cl)2-bpy)2(bpy)]2+

5.

98 Figure 5.7. Transient absorbance spectra (kex = 417 nm) in CH3CN of (a) la.

The data were recorded at 10 ns (•), 0.5 jus (•), 1.0 us (A), 2.0 us

(•), and 10 us (•) delays after the laser pulse, (b) lb. The data

were recorded at 10 ns (•), 1.0 us (•), 2.5 us (A), 5 us (•), and 20

us (•) delays after the laser pulse, (c) lc. The data were recorded

at 0.1 us (•), 0.5 us (•), 1.0 us (A), 2.0 us (T), and 4.0 [is (•)

delays after the laser pulse.

Figure 5.8. Transient absorbance spectra (AeX = 417 nm) of Cl-substituted 99

complexes in acetonitrile at room temperature Ru(4,4'-(C1)2-

bpy)3(PF6)2 (•), 2a (•), and 2b (A). The data were recorded 10 ns

after the laser pulse.

xviii

Figure 5.9. Single wavelength kinetics monitored at the ground state-excited 101

state isosbestic point for la on pH = 1 pretreated Ti02 (black

line, top) and pH = 11 pretreated TiCh, 0.1 M LiC104 (red line,

bottom).

Figure 6.1. Chemical structure of Tris(2,2'-bipyridine) ruthenium(II) 124

dichloride, (Ru(bpy)32+ Cl2).

Figure 6.2. Normalized UV-Vis absorption (solid line) and steady state 131

emission (dashed line) spectra of Ru(bpy)32+ in physiological

Tyrode's solution. The spectra were collected at room

temperature.

Figure 6.3. Quenching of Ru(bpy)32+ is observed through a) the quenching of 135

Ru(bpy)32+ photoluminescence and through b) the shortening of

the Ru(bpy)32+ lifetime. Each trace was acquired in a potassium

glutamate pipette solution (previously described) purged with

either 100% nitrogen (black trace), in ambient air (red trace), or

purged with 100% oxygen (blue trace). The Insets of a and of b

depict the Stern-Volmer plots of the dynamic quenching,

represented by equation (2), as a function of photoluminescence

intensities and lifetimes, respectively. The Stern-Volmer

quenching constants (Ksv) were calculated from the slope of the

plots and using equation (3) to be 0.0025 +/. 0.0011 and 0.0027 +/. 0.0033.

Figure 6.4. After loading of the cardiomyocytes was complete (prior to time 137

zero of this plot), uncoupler (FCCP) was added and a

simultaneous rise green fluorescence (FAD oxidation) and the

red photoluminescence (dequenching of Ru(bpy)32+ ) occurred.

Shortly after, a rise in KATP current (black trace) was observed.

xix

These effects were reversed by exposure of the myocyte to the

inhibitor NaCN. INSET: A single cardiomyocyte loaded with 2+

O.lmM Ru(bpy)3 . The red circle in both images highlights the

location of the membrane-pipette interface. The image intensity 2+

represents the intracellular Ru(bpy)3 emission at 605nm a) in

the presence of a Tyrode's bath solution and b) in the presence

ofl.OmMFCCP.

Figure 6.5. a) The cardiomyocyte was exposed to several different 140

concentrations of the uncoupler, FCCP (in nM: 0, 10, 25, 50,

1000). b) An enlargement of the plot at lower FCCP

concentrations.

Figure 6.6. The normalized photoluminescence plot indicative of the loading 141

of Ru(bpy)32+ by the patch-clamp technique.

Figure 6.7. The effect of varying bath p02:. The cell was exposed to bath 143

solutions saturated with 100% 02 or 100% N2 with expected 2+

effects on Ru(bpy)3 PLI but no change in flavin fluorescence.

Figure 6.8. The flavoprotein fluorescence and Ru(bpy)3 145

photoluminescence were recorded in the presence of 4mM NaCN

purged with 100 %N2 or in ambient air. There is a larger change

in the intracellular concentration, depicted by the Ru(bpy)32+

photoluminescence, than in the flavoprotein oxidation. This

illustrates the effect of diffusion of oxygen from the extracellular

medium during inhibited respiration.

Figure 6.9. Calibration of the Ru(bpy)3 photoluminescence signal in the 146

presence of NaCN. a) The flavoprotein redox properties, the

Ru(bpy)32+ photoluminescence, and the external % O2 were

xx

recorded during a period of 4min exposure to NaCN purged with

100% N2, washout with Tyrode's solution, and then another

4min period of NaCN purged with 100% O2. The measurements

were completed at 37°C. b) Ru(bpy)32+ photoluminescence

values, taken during periods of NaCN (purged with either N2 or

O2), were plotted with their corresponding external O2

concentrations. A best fit line was used for calibration.

Figure 6.10. a) The cardiomyocyte (also used in the measurement in Figure

7.7) was stimulated at 2Hz from the start of this recoding to

induce twitching. The basal respiration, Ru(bpy)32+

photoluminescence signal, was measured to be 23.0 %C>2

(216uM). Oxidation of the flavoproteins was immediately

instigated upon stimulation. Before cell death, a spike in both

the Ru(bpy)32+ photoluminescence and the flavoprotein

fluorescence appeared with a measured intracellular %C«2 of

4.3% (40.4uM). b) The slope of the Ru(bpy)32+

photoluminescence curve that resulted from stimulation was fit

with a best fit line. The overall decrease in intracellular oxygen

content was measured to be 30.3uM/min.

Figure 6.11. Metabolic oscillations were induced in cardiomyocytes by a brief

period of high pC>2 (bath solution saturated with 100% 02).

100% N2 saturated solution was applied after -15 minutes but

oscillations were not suppressed.

xxi

Chapter 1. Introduction to Organic and Inorganic Chromophores

Archeological evidence has shown that the use of dyes has dated back to over 5000 years

ago. Natural dyes, derived from animals, plants and minerals, have been used to enhance

color through their affinity to a particular substrate.

The first synthetic dye was created from coal-tar in the mid 1800s by William Henry

Europe and finally, the vast expansion of the dye industry occurred in America. The

development of synthetic dyes had a significant impact on American industry and within

the scientific community, peaking in the 1970s.1

Chromophores, in particular, natural and synthetic, are the moieties of dyes that are

responsible for the absorption and the reflection of specific wavelengths in the visible

region of the electromagnetic spectrum. The energies absorbed, and therefore the color

of the dye, depend on the energy gap of specific molecular orbitals. The greater the

difference between the potential energy of the ground state and the excited state, the

shorter the wavelength absorbed. This relationship was first realized by Max Planck to

represent the quantum mechanical theory behind the distribution of energy of

electromagnetic radiation; his mathematical relationship (Equation 1.1) models the

radiation as harmonic oscillators of quantized energy.2

1

E = hv= hcfk (1.1)

Equation 1.1 states that the frequency, v, of the incident photon is directly proportional to

the ratio of the speed of light to the wavelegnth, c/X, and to the energy, E. Plank's

constant of proportionality is represented as h. Therefore, the greater the energy gap

between the quantized molecular orbitals, the greater the energy and frequency of the

photon absorbed, but the shorter its wavelength.

1.1. Molecular Orbitals and Electronic Properties of Chromophores.

Molecular orbitals and the corresponding potential energies of the electrons that reside

within these orbitals can be depicted by a Jablonski-like potential energy diagram, Figure

1.1.3 This generalized diagram illustrates the possible transitions between the electronic

states of a molecule, consisting of vertical and horizontal transitions.

The Born-Oppenheimer Approximation accounts for the vertical and horizontal electron

transitions which represent the distinction between electronic and nuclear motion,

respectively. This stepwise mathematical approximation, developed in 1927, first

establishes a wavefunction solution from the electronic Schrodinger equation with a

constant nuclear motion. 4 The possible vertical transitions in Figure 1.1 demonstrate the

time-independent nature of this solution. The lower potential energy well of the excited

state is slightly shifted along the reaction coordinate to represent nuclear solution to the

2

Figure 1.1. A generalized Jablonski-type diagram depicting the potential energies of the Highest Occupied Molecular Orbital (HOMO) and the Lowest Unoccupied Molecular Orbital (LUMO) electrons. Absorbance of a photon (—), fluorescence (—), phosphorescence (—), intersystem crossing rate (Arise), and vibrational cascade (kvc) are other electron transitions illustrated.

/t

fa

e m en © an

@

I ©

3

Schrodinger equation. This distinction is plausable due to the relative electron and

nuclear masses.

The potential energy wells are grouped horizontally by spin multiplicity and vertically by

energy. The higher energy wells illustrate the potential energy of the excited states while

the lower-lying energy wells represent the gound state electron configuration. The

horizontal lines in each of the wells symbolize the lowest lying vibronic energy level of

each state.

1.2. Chromophore Absorption.

Although the term "chromophore" encompasses a broad array of dyes, these dye

molecules must contain specific structural characteristics to absorb in the visible or near

ultraviolet regions of the electromagnetic spectrum. The light absorbing moiety of the

compound contains either a conjugated 7i-system or a ligand-coordinated transition metal.

Both allow for the absorption of an incident photon to provide the necessary energy for

an electron transition from the ground state to a higher energy excited state. The

promoted electron initially resides in the highest occuppied molecular orbital (HOMO).

The absorbance of a molecule can be defined as the fraction of incident light that

produces excited states:

Ax = ex • b • c = logioCV/) (1.2)

4

where s% is the molar extinction coefficient at a particular wavelength, b is the path

length, c is the sample concentration, and (/<//) is the fraction of the incident light that

produces excited states (I0 is the intensity of the incident light, while / is the intensity of

the absorbed light by the dye.) The absorbance at a single wavelength, A*., can be

measured from a simple ground state absorbance spectrum and is represented in Figure

1.1 as a violet-colored arrow. The intensity of the absorbance band is dependent on the

probability of the electrons occupying a particular vibronic level within the excited state.

Absorbance is mainly the highest energy electron transition achievable.

1.3. Chromophore Emission.

After absorption, the excited state electron maintains the energy donated by the absorbed

photon. This electron is short-lived and will eventually resume its stable position in the

original ground state, S0, releasing its stored energy; the released energy is equal to the

energy difference between the lowest vibronic level of the excited state and a particular

vibronic level of the ground state. Here, the intensity of the emission at a specific

wavelength depicts the probability of electrons populating a specific vibronic level of the

S0. The lowest energy singlet excited state is also commonly referred to as the lowest

unoccupied molecular orbital (LUMO). This energy is emitted as either a photon or as an

alternative form of non-emissive energy; therefore, chromophore emissions can be

categorized into two groups, radiative and non-radiative, respectively.

5

Radiative Pathways

Fluorescence is considered the fastest radiative electron transition and occurs from the

lowest vibronic level of the lowest lying singlet excited state, S*, to the singlet ground

state, S0 (signified by a blue arrow in Figure 1.1.) The wavelength of this radiative

emission depends on the vibronic level of the ground state to which the electron returns;

the energy of the emitted photon is therefore less than or equal in energy to that of

absorbance. There is no change in the spin multiplicity during fluorescence.

Phosphorescence, on the other hand, is a slower radiative process that emits photons of

lower energy than does fluorescence. This is due to a change in spin multiplicity within

the excited state and requires a non-radiative horizontal electron transition before

phosphorescence can occur. This fast transition from the singlet excited state, S*, to the

lower lying triplet excited state, T*, is referred to as intersystem crossing, klSQ. Once the

electron has reached the lowest vibronic energy level of the triplet state, it must once

again change spin multiplicity to overcome the spin forbidden transition back to the

singlet ground state. This spin forbidden transition accounts for the longer-lived triplet

excited state, and is represented by a red arrow in Figure 1.1.

Organic chromophores that possess a delocalized 7t-system are primarily considered as

fluorescent probes; the singlet and triplet excited states are both well defined; often the

triplet state is not involved in the decay pathway. These excited organic chromophores

are therefore short-lived, and high-energy radiation is both absorbed and emitted. In

6

contrast, inorganic chromophores, which possess a transition-metal center with

coordinated conjugated ligands, have "mixed" excited states; there is a large overlap of

the triplet and singlet excited states. The radiative emission can no longer be considered

solely fluorescent or solely phosphorescent but as some combination of the two.

Therefore a broader term is assigned to describe this mixed emission: photoluminescence,

Figure 1.2. Since the photophysical and electronic properties of inorganic molecules

differ from that of the organic fluorescent dyes, each type is found to have different novel

applications. The applications of both types of dyes were investigated and are reported

within this dissertation.

All emitted photons, whether undergoing fluorescence, phosphorescence, or

photoluminescence, must occupy the thermodynamically equilibrated excited state

(THEXI state), also known as the lowest lying vibronic level of the lowest energy excited

state, before transition to the ground state can occur. This phenomenon is known today

as Kasha's rule.5 Exceptions rarely occur but are possible in organic molecules whose

excited singlet states are separated by a large energy gap, allowing fluorescence to occur

from either state.

Non-radiative Pathways

Other pathways exist that release the energy stored in the excited state of the molecule

that do not produce a photon. These pathways are known to be non-radiative, and are

depicted by the curvilinear arrows in Figure 1.1 and in Figure 1.2. Inter system crossing

7

a)

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(kls<:) occurs when an electron changes spin multiplicity and potential energy well states

(S* ->T*), while internal conversion (not depicted) occurs without a spin flip between an

singlet excited state to another singlet excited state of lower energies (S* -^ S*).

Vibrational cascade can occur within a single potential energy well of either the excited

or ground state and releases energy through vibrations (infrared) rather than emitting

photons, and is represented in Figure 1.1 as kvc. Finally, energy can be released as non

specific non-radiative decay from the THEXI state to the ground state, k^, Figure 1.2.

The performance of the chromophore as an emitter is highly dependent on the energy of

the incident light, the absorbance efficiency (e), the electronic properties of the

chromophore, the quantum yields of the electron transitions, and the local environment.

1.4. Quantum Yields.

The efficiency for each electron transition can be defined as the fraction of occurrence of

the given quantum state relative to all other competing pathways; the pathways are all

expressed as rate constants associated with the competing transitions.

<|>i= ki/ktot (1.3)

where ()); is the quantum yield of transition i, k\ is the rate constant of transition i, and k^

is the sum of rate constants of all competitive transitions (Ar and k^.

9

Measurement of the quantum yield allows for direct comparison of emission efficiency of

chromophores.

1.5. Lifetimes.

Emission is a random depopulation of the excited state, and each excited state

chromophore of identical structure has the same probability of emitting a photon in a

given interval of time. Measuring the decay of this population produces an exponential

decay. The initial intensity of the decay, time = 0, is proportional to the quantity of

excited states formed. The lifetime is defined as the value which exists at 1/e of this

initial intensity. The lifetime, x, is also characterized by the inverse of the sum of the

radiative and non-radiative decays:

x = (kr + kmyx (1.4)

where ^ and k^ are the radiative and non-radiative decay constants, respectively. For a

single exponential, the lifetime also signifies the average time the chromophore remains

in the excited state. This may not be true for heterogeneous solutions; a multi-exponential

may exist.

As previously stated, excited state electrons of organic chromophores exhibit shorter

lifetimes on the orders 10"9s and faster, while inorganics typically exhibit longer lived

excited stated on the orders of 10"6s and slower.

10

1.6. Conclusion.

The structural, photophysical, and electrochemical properties of chromophores are

important characteristics to take into account when choosing a dye for a particular

research endeavor.

In chapters 3 and 4, several organic chromophores are investigated to reveal their

properties. These dyes were studied in hopes to find efficient light-harvesting sensitizers

for solar energy conversion.

In chapter 5, inorganic Ru-centered chromophores were also chosen to find the ideal

complexes for solar cells that would maximize the incident light to electrical energy

conversion efficiencieson TiC>2.

Finally, chapter 7 expores the development and application of a novel technique which

measures the intracellular oxygen tension in single guinea pig ventricular cardiomyocytes

using a Ru-centered dye as an oxygen sensor.

11

References.

Morris, Peter. "Does the Science Museum, London, have Perkin's Original Mauve

Dye? A Critical Reassessment of a Chemical Icon". History and Technology

2006,22(2), 119-130.

Atkins, P; de Paula, J. Physical Chemistry. 2001.

Turro, N.J.; Modern Molecular Photochemistry. The Benjamin/Cummings

Publishing Co., Inc. Menlo Park: California. 1978.

Levine, I.N. Quantum Chemistry; 5th Ed. Prentice Hall: New Jersey. 2000.

a) Lewis, G.N.; Kasha,M. Phosphorescence and the Triplet State. 1944, 66, 2100-

2116. b) Kasha, M. "Characterization of Electronic Transitions in Complex

Molecules." 1953. 14-19.

12

Chapter 2. Introduction of Light-Harvesting Chromophores for

Applications in Photovoltaics

The idea that man will one day be able to freely capture the sun's rays for harvesting

energy is a provocative concept. In today's society, especially, alternate forms of energy

have become the central focus within the political arena and the scientific community. It

is universally understood that the Earth's fossil fuels will inevitably be depleted and that

it is crucial to find an efficient replacement. It has been estimated that the fossil fuel

reserves will be exhausted over the next century.*

The first photovoltaic effects were observed over a century ago when E. Becquerel2

measured a voltage generated when a material was excited with light. Throughout the

early twentieth century, scientists endeavored to make advancements in this area;

however, it wasn't until the early 1970's when Gerisher, Memming, and Tributsch

initiated investigation into creating systems composed of molecular dyes proximate to a

semiconductor surface. Today, high-efficiency solar cells are desired not only to

decrease the cost of photovoltaic systems but to hopefully one day serve as a primary

means of achieving adequate energy in the future with no harmful emissions or

13

byproducts. The efficiency of such solar cells has reached 13-15% conversion due to

recent advancements by Michael Gratzel and his coworkers.3

The modern electrical solar device has been fabricated from semiconductor materials to

convert a fraction of the energy contained in sunlight to electrical energy. There is much

understanding required to design and construct an efficient solar cell. The features of the

solar spectrum, semiconductor physics, semiconductor device fabrication, and the dyes

for harvesting energy are several fundamental areas required for understanding and

improving the efficiency of regenerative solar energy devices.

2.1. The Features of the Solar Spectrum.

Before considering any type of synthetic energy conversion, the energy source must be

thoroughly explored; in this case, it is the sun. Outside the earth's atmosphere, 98% of

the solar energy radiation exists between 250 and 300 nm.4 The density of sunlight that

reaches the earth's surface depends on several factors: the atmospheric conditions such as

water vapor and ozone, the latitude at which the measurement is made, and the season.

For example, even though the difference is small, on a cloudless day when the sun is at

the zenith, the radiation is maximized. A standard for solar spectral irradiance

distribution is the air mass 1.5 (AMI.5) spectrum that has a broad maximum around 500

nm that tails into the IR region (Figure 2.1 .)5 Since the visible region yields the highest

intensity of energy reaching the Earth's surface, it is most advantageous to engineer solar

cell technology to utilize this region of the electromagnetic spectrum.

14

Figure 2.1. The solar spectral irradiance distribution (AM1.5).5 This figure represents the intensity of energies of the electromagnetic spectrum that penetrate the atmosphere and reach the Earth's surface.

15

2.2. Semiconductor Physics and Device Fabrication.

Semiconductors, unlike insulators and conductors, allow for an alteration in the

conductivity by simple manufacturing procedures and can carry current by either

negatively or positively charged carriers, i.e. electrons and holes. Recently, research and

development has expanded the area of semiconductor exploration to the creation of thin

films. There are several reasons for studying thin films; the cost of harvesting solar

energy is extremely expensive when dealing with single crystals and bulk materials.2 In

addition, there exist many deposition techniques for thin films as well as the numerous

elements available for synthesizing the semiconductors. In selecting a semiconductor,

the spectrum of the light source relative to the absorption of the material must be

considered. Out of the electromagnetic spectrum that comprises sunlight, only those

wavelengths with greater energy than the band gap, Eg, can be absorbed to produce

photocharges. Titanium dioxide, Ti02, is the relevant semiconductor examined herein.

Several forms of TiC>2 exist such as rutile and anatase; recently, electrodes have been

fabricated without using single-crystals. This accomplishment has triggered a steep

acceleration in the field. Anatase phase of titanium dioxide is a wide-band gap

semiconductor which has an indirect band gap with an energy of 3.2 eV.2 Because the use

of wide-band gap semiconductors in photovoltaic cells is hampered by the fact that they

utilize only a small portion of the solar spectrum, mainly the ultraviolet, sensitizing TiC>2

to visible light with dye molecules has become a primary research focus. Titanium

dioxide is undoubtedly one of the most important electrode materials in semiconductor

photoelectrochemistry. After pioneering by Fujishima and Honda in 1972,6 there has

16

been an increasing interest in employing this material as a light-harvesting electrode in

photochemical cells.

2.3. Dyes for Light Harvesting Energy.

An important part of the photovoltaic cell is the light-absorbing molecules called dyes or

sensitizers. In 1873, Vogel discovered organic dyes deposited on semiconductor halide

grains increased the sensitivity of these particles on the low energy side of the visible

spectrum.7 The synthetic organic and inorganic aromatic dyes investigated in this

dissertation resemble, in some respects, nature's own chlorophyll pigments utilized in the

photosynthetic process, Figure 2.2. Chlorophyll contains a chromophore head, consisting

of a magnesium metal centered porphyrin, and a hydrophobic hydrocarbon tail that

allows for the anchoring of the dye to the thylakoid membrane. The dyes investigated for

photovoltaics are similar in that they are anchored to the semiconductor TiC»2.

Furthermore, the synthetic dyes examined herein for light harvesting emulate

chlorophyll; in both the synthetic systems and the chlorophyll complex, specific energy

photons are absorbed, electrons are promoted to excited states of higher potential energy

and are donated to other components of the system with a lower reduction potential.

Chlorophyll donates its excited electrons to nearby intermediates of the electron transport

chain within the membrane for energy production.

17

Chromophore r N

Figure 2.2. Molecular structure of chlorophyll. This natural dye contains a metal-centered porphyrin head which serves as the chromophore moiety. The conjugated 7r-system is apparent in the structure. The long hydrocarbon tail acts to anchor the chlorophyll in the thylakoid membrane.

18

2.4. Dye-Sensitized Solar Cells.

Dye Sensitized Solar Cells (DSSC's), also referred to as Gratzel Cells, are forecasted to

be a competitive form of renewable energy due to their price to performance ratios.

The framework of a DSSC is represented in Figure 2.3 while a simplified, generally

accepted mechanism, incorporating the T1O2 and sensitizer energetics, of a regenerative

solar cell is shown in Figure 2.4. The basic components of the DSSC: electrodes

completing the external circuit, the photoanode containing the semiconductor/sensitizer

elements, the dark electrode, and the electrolyte for sensitizer ground state regeneration.

In Figure 2.3, external circuit is represented by the alligator clamps connecting the

fluorine-doped tin oxide (FTO) glass substrate (the photoanode) and the Pt electrode (the

dark electrode). The TiC"2 semiconductor nanoparticles are directly deposited onto the

FTO conductive glass substrate to create a 8-10 urn thick film. The redox couple is an

electrolyte solution, typically consisting of an iodide/tri-iodide (I'/V), that completes the

internal circuit and is represented by a reduced-donor (D) and an oxidized-acceptor (D+).

The mechanistic steps represented in Figure 2.4 is as follows:

1. Incident light (near UV or visible) is absorbed by the chromophore moiety of

the sensitizer (SD) and creates an excited state (S*). The competitive process,

the dashed arrow, represents the radiative and nonradiative emission of the

excited state, kr or k^.

19

o •

•

©7®

TiCh Semiconductor Nanoparticle

Ground State Chromophore

Excited State Chromophore

Redox CouDle

Figure 2.3. Regenerative Dye Sensitized Solar Cell (DSSC). After the chromophore (sensitizer) absorbs a photon, an electron donated from the excited state to the mesoporous semiconductor TiC«2 nanoparticles. The injected electron then travels through the mesoporous thin film to the fluorine-doped tin oxide (FTO) glass substrate where it then proceeds through the electronic circuit. The oxidized form of the redox couple regenerates the ground state of the sensitizer.

20

S

Ground State Potential Energy

Excited State Potential Energy

Potential Energy of the Conduction Band

Potential Energy of the Valance Band

Redox Couple

Figure 2.4. The Mechanistic Scheme of the Regenerative Dye Sensitized Solar Cell. The ground state sensitizer, S0, absorbs the incident photon and promotes it to the excited state, S*. The energy of the incident photon must be greater than or equal to that of the energy gap between these two energy levels. The photons are not absorbed by the FTO or the TiC>2 because both require energy of ultraviolet photons for absorption. The electron can then either undergo injection, k^, or emission ( ). The injected electrons then are collected at the FTO and the external circuit is utilized or recombination occurs, krec ( ), and regeneration cannot be completed. Eg ( ) represents the band gap of the TiC>2 semiconductor.

21

2. If the excited state is a stronger reductant than the Ti02 conduction band

(ECb), interfacial electron transfer is energetically favored. The electron is

injected (£inj) and travels through the Ti02 mesoporous thin film. The electron

in the conduction band can either be collected by the conductive glass at the

FTO/Ti02 interface (r|cou) or can undergo recombination to the ground state of

the sensitizer. Figure 2.3 also depicts the recombination of the injected

electron which decreases the overall efficiency of the DSSC. Dyes are often

designed in attempt to eliminate the krec.

3. These collected electrons travel through the external circuit until they reach

the counter Pt electrode.

4. Regeneration of the donor (D) is completed at the platinum electrode, and the

cell is fully restored.

The success of the Gratzel cell depends on the ability to absorb the photon, the quantum

yield for injecting the absorbed photon into the TiC>2 ((pinj), the collection efficiency of the

conductive glass substrate (r|coii), and the energetics of the redox couple. The overall

incident- photon-to-current efficiency (IPCE) can be defined as:

IPCE = a • ( inj • ricoii (3)

where a is the absorptance of the sensitizer. This directly measures the photocurrents

22

produced in the DSSC when irradiated with light and allows for comparison between

cells. Since the absorptance is dependent on wavelength of the incident light, the IPCE

also varies with the wavelength.

23

References.

Ciamician, G. Science 1912, 36, 385.

Gratzel, M. Nature 2001, 414, 338.

K. Nazeeruddin, M.K.; Bessho, T.; Le Cevey; Ito, S., C. Klein, F. De

Angelis, F.; Fantacci, S.; Comte, P.; Liska, P.; Imai, H.; Gratzel, M. J

Photochem Photobiol 2007, 185 (2-3), 331-337.

Fahrlenbruch, A.; Bube, R. Fundamentals of Solar Energy: Photovoltaic Solar

Energy Conversion. Academic Press: New York, 1983.

"WIRE." 24 July 1998. 1 April 2003.

http ://wire0. ises. org/wire/glossary. nsf/H/O?Open&000040F2

Fujishima, A; Honda, K. Nature 1972, 238, 37-38.

Vogel, H.W.; Photochemie and Beschriebung der Photographischen Chemikalien.

5th ed. Gustav Schmidt: Berlin. 1873.

24

Chapter 3. Organic Rigid-Rod Linkers for Coupling Chromophores to

Metal-Oxide Nanoparticles

3.1. Introduction.

Aromatic chromophores adsorbed to metal oxide surfaces can provide deep insights into

charge injection and recombination at the dye/metal-oxide interfaces. Previously

reported in literature are successful molecular-linkers such as siloxane1, carboxylic acid2,

acetyl acetonate3, amide4, or phosphonate5 functional groups physi- or chemi-sorbed to

metal oxide surfaces. At times, flexible methylene spacers have been synthesized

between the functional binding groups and the chromophore to avoid direct electron

interactions3'7. However, the flexibility of these molecules creates an unknown

orientation and unpredictable molecular interactions with the metal oxide surface.

Our collaborators at Rutger's University, Newark, Galoppini and coworkers, recently

developed a novel linker to replace the flexible spacers. The rigid spacers consist of p-

phenylenethynylene or w-phenylenethynylene bridges, (Ph-E)n, and are utilized to

examine photophysical properties and electron transfer processes at nanoparticle surfaces.

The linkers also allow for fully conjugated systems from the chromophore directly to the

25

surface. Furthermore, the length of the rod and, ideally, the distance of the chromophore

from the surface can be controlled; each phenylethyne unit is 5.4 A long and is rigid. The

results demonstrate that this is an important new approach for coupling molecular

components with nanostructured materials. '

The bipodal organic dyes presented are dimethyl 5-(l-pyrenylethynyl)isophthalate (2) \p-

Py-E-Ph-ROD], dimethyl 5-(4-(l-pyrenenylethynyl-phenylethynyl)isophthalate (3) [p-

Py-(E-Ph)2-ROD], dimethyl 5-(3-(l-pyrenenylethynyl-phenylethynyl)isophthalate (4) [m-

Py-(E-Ph)2-ROD], and dimethyl 5-(bis-3,5-(l-pyrenenylethynyl-

phenylethynyl)isophthalate (5) [Bis(7W-Py)-(E-Ph)2-ROD]. All dyes were synthesized

and provided by Elena Galoppini and co-workers, Figure 3.1,10 and were compared to the

commercially available pyrene carboxylic acid (1).

Overall, the purpose in using these rigid-rod organic molecules as dyes for sensitization is

clear. These organic sensitizers have well-defined spin states, vibronic structures, and

excimer emissions. Controlling the length of the rigid linker allows for specific

increases in molar extinction coefficients over the commercially available Py-CC^H.

These organic rigid-rod linkers were used to couple pyrene to the surface of Ti02

(anatase) and Zr02 nanoparticle thin films. The rigid-rods also shift to the red the long-

wavelength absorbance of the pyrene chromophore. The rigid-rod linkers afford high

surface coverages, -10" mol/cm , and high surface stabilities on the nanostructured metal

oxide films in acetonitrile. The appearance of a pyrene excimer-like emission on Zr02

26

Chromophore

^

> Linker

C 0 0 H HOOC" ^ "COOH HOOC" ^ "COOH

Py-C02H (1) Py-E-Ph-Rod (2) Py-(E-Ph)2-Rod (3)

J

HOOC ^-^ COOH

»i-Py-(E-Ph)j-ROD (4)

HOOC v COOH

Bis(m-Py)-(E-Ph)2-ROD (5)

Figure 3.1. Schematic of the five bipodal pyrene-spacer-anchor systems compared in this study. The phenylenethylynene spacer is abbreviated (E-Ph).

27

nanoparticles indicates that the bipodal rigid-rods do not spatially isolate the

chromophores effectively. The emission on HO2 is completely quenched, consistent with

quantitative electron injection into the semiconductor. Nanosecond transient absorption

measurements (AeXC = 417 nm, 8-10 ns fwhm, 3.1 mJ/cm2) indicate rapid excited-state

electron injection, k-mj > 108 s"1, and second-order recombination with an observed

average rate constant of &0bs = 2.5 ± 0.3 x 107 s"1, independent of which rigid-rod was

excited. Preliminary photoelectrochemical studies in regenerative solar cells with 0.5 M

LiI/0.05 M I2 indicate a quantitative conversion of absorbed photons into an electrical

current.

3.2 Experimental Section.

Metal Oxide Thin Film Preparation

Colloidal HO2 and ZrC>2 films were prepared by a previously described sol-gel technique

that produces nanocrystals of ~20nm in diameter and mesoporous films (after sintering at

450°C) of approximately 10-u,m thickness.3'11 For absorption and luminescence studies,

the particles were coated onto glass slides (VWR Scientific), and for

photoelectrochemical studies, they were coated onto fluorine-doped tin oxide conductive

glass (FTO; Libbey-Owens-Ford).

28

Binding.

The five pyrene compounds studied in detail are shown schematically below. Py-C02H,

(1), was purchased (Aldrich, 99%), and the para- rigid-rods, Py-E-Ph-Rod, (2), and Py-

(E-Ph)2-Rod, (3), were prepared as previously described.10 The meta- rigid-rigid rods, m-

Py-(E-Ph)2-ROD, (4) and Bis(/w-Py)-(E-Ph)2-ROD (5), were prepared as previously

described.12

The binding of Py-C02H as well as the pyrene-substituted rods to the Zr02 films was

accomplished by immersing the films in acetonitrile solutions of 1-5 overnight at room

temperature. The surface attachment of 1 as well as rigid-rods 2-5 to the Zr02 or Ti02

films resulted in the appearance of a pale yellow color. The surface coverage reached a

limiting value of ~8 x 10"8 mol/cm2 at high concentrations of the acetonitrile solutions of

1-3. Surface coverages of the carboxylic acid derivatives of 2-5 also approached ~8 x 10"

8 mol/cm2 at concentrations of -0.25mM THF solutions. The thin films were treated with

base by immersion in a pH=ll solution prior to binding of 2-5 (carboxylic acid

derivatives); this pretreatment was necessary for reaching high surface coverages.13

Adsorption Isotherms.

Surface binding of 2-5 was monitored spectroscopically by measuring the change in film

and solution absorbance after equilibrating the film overnight in solutions with known

concentrations of the sensitizers. In all cases, the surface coverages saturated at an

~0.25mM sensitizer concentration. The equilibrium binding for all sensitizers were

29

described by the Langmuir adsorption isotherm model3'14 from which the surface binding

constants (Xad) were abstracted using equation (3.1),

[ S J e 9 = _ J _ + [Sleg (3.1) r ^adl^o To

where [S]eq is the equilibrium concentration of the sensitizer (2-5), T0 is the saturated

surface coverage, and Tis the equilibrium surface coverage at a defined molar

concentration. Plots of [S]eq/ r versus [S]eq were fitted linearly to obtain the binding

constants Tad and surface coverages ro.

Spectroscopy.

Absorption spectra were acquired at ambient temperature in acetonitrile (Burdick and

Jackson, spectroscopic grade) for methyl ester derivatives or in THF for carboxylic

derivatives (Acros, spectroscopic grade) using a Hewlett-Packard 8453 diode array

spectrometer. Nanosecond transient absorbance spectra were acquired in a cuvette with

nitrogen-saturated acetonitrile or THF and 417-nm light excitation as previously

described.15

Steady-state fluorescence spectra were obtained with a Spex Fluorolog that had been

calibrated with a standard NIST tungsten-halogen lamp. Time-resolved fluorescence

decays were acquired with 316-nm excitation and a single-photon counting apparatus that

has been previously described.16 Fluorescence quantum yields for carboxylic acid

30

samples, <f>Fi, were calculated in THF with pyrene as a reference (<|>FI = 0.72) and using

equation (3.2):

<t>Fl = (VA.)(IAXlU/llr)2<|)r (3.2)

where Ar and As are the absorbances of the actinometer and the sample, respectively, Ir

and Is are the integrated PL intensities of the actinometer and the sample, respectively, ns

and nr are the indexes of refraction for the solvents used for the actinometer and for the

sample, respectively and (|)r is the quantum yield for pyrene in THF. The radiative (kr)

and non-radiative (Am) rate constants were calculated using equations (1.3) and (1.4).

Fourier transform infrared attenuated total reflectance (FTIR-ATR) spectra of sample

crystals and sensitized thin films were obtained using a Thermo Electron Corporation

Nicolet 6700 FT-IR and a Golden-Gate Apparatus.

Electrochemistry.

Cyclic voltammetry was performed with a BAS potentiostat under an atmosphere of

nitrogen at room temperature with a three-electrode configuration. A glassy carbon

electrode (2 nm diameter) or a sensitized Ti02 film was used as a working electrode

along with a platinum wire auxiliary electrode and a AgVAgCl reference electrode in

0.1M TBACIO4/CH3CN electrolyte solution. The potential of the Ag+/AgCl reference

electrode was referenced externally versus the ferrocene/ferroenium (Fc/Fc+) redox

couple. Compounds 2-5 show an irreversible first oxidation both in fluid and when

31

anchored to the TiCh when monitored at scan rates of lOOmV/s. Since the half-wave

potential (E1/2) could not be calculated because of the absence of the cathodic peak (EpC),

the anodic peak (Epa) potential is therefore quoted.

Photoelectrochemistry.

The photocurrent action spectra were obtained in a two-electrode sandwich cell

arrangement similar to figure 2.3. A 0.5M LiI/0.5M I2 acetonitrile solution was used as

the electrolyte. Quasi-monochromatic light from 400 to 600 nm was achieved with a 150

W Xe lamp coupled to a f/2 monochrometer, and incident irradiances were typically of

lmW/cm2.

Calculations.

Semi-empirical geometry optimization was performed at the AMI level using Spartan '02

by Wavefunction, Inc.

3.3 Results.

The absorbance spectra of 1 and rigid-rods 2 and 3 in neat acetonitrile and on TiCh,

Figure 3.2, show that the rigid-rods increase the extinction coefficient and shift to the red

the long-wavelength absorbance of the pyrene chromophore. The Uv-Vis absorbance

spectra of 2, 3, 4, and 5 in THF solutions and bound to TiCVglass are shown in Figure

3.3. A comparison between these spectra allows one to observe the affects of the

substitution position (para- or meta-) and of the number of PE units in the linker. The

32

a)

8.0x10

6.0x10

4.0x10'H CO

2.0x10

250 300 350 400 450

Wavelength, nm

b)

c

o

0.6

0.5

0.4

0.3

0.2 J

380 p-i-fc i. arT -

400 420 440 460

Wavelength, nm

480

Figure 3.2. Absorption spectra of (a) 1-3 in neat CH3CN and (b) 1-3 adsorbed to T1O2 in neat CH3CN. For both (a) and (b), solid lines correspond to 1 (—) , dashed lines correspond to 2 (——), and dashed-dot lines correspond to 3 (—) .

33

a)

HOOC^^XOOM

H O O C ^ C O O H

• 1 • 1 T~. 1 " • f * 1 ' "1

250 300 350 400 450 500

Wavelength, nm

b)

8 | 0.4 J

1

375 400 425 450 475 500 525 550 575 600 Wavelength, nm

Figure 3.3. Absorption spectra of 2-5 (a) in THF solution and (b) bound to TiCVglass. 2 ( - - - ) , 3 ( - ) , 4 ( - - - ) , 5 ( ).

34

band at higher energy in the spectra 3.2a and 3.3a was assigned to the n, it* transition of

the rigid-rods (E-Ph). The additional (E-Ph) group in 3 compared to 2 (both para rigid-

rods) increases the extinction coefficient and red shifts the absorbance band -17 nm of

the lower energy absorbance band assigned to the pyrene chromophore. Compound 4

(the meta rigid-rod), however, has the same spacer length as compound 3 but is blue

shifted and overlaps with the pyrene absorbance band of compound 2. In conclusion, an

increase in length of the spacer with a para substitution allows for the compound to

absorb further into the red and utilize more of the visible spectrum. Furthermore,

introducing a second pyrene moiety in the meta position increases the intensity of the

absorption, and therefore the molar extinction coefficient, by a factor of almost 2

(compound 4 vs. 5).

Absorption spectra on insulating colloidal ZrC>2 films were within experimental error the

same as those on TiCh (Figure 3.2b, 3.2b). Only a long-wavelength absorption tail into

the visible is observed when Py-CC^H is anchored to Ti02. The higher-energy vibronic

transitions, below 380 nm, are obscured by the fundamental absorption of the

semiconductor; however, the lower energy vibronic band is visible for compounds 2-5.

Light excitation of the pyrene compounds in fluid solution and anchored to Zr02 thin

films leads to room-temperature fluorescence. At low concentrations in THF, a single

emission band is observed. The fluorescence quantum yields were estimated using the

optically dilute technique with pyrene as a standard. Fluorescence decays were first

order, and the radiative and nonradiative rate constants were calculated using standard

equations9'17 (Table 3.1). Sonication of rigid-rods 2 and 3 in saturated ethylene glycol

35

Table 3.1: Photophysical Properties ofl-3a

sensitizer

Py-E-Ph-Rod (2)

Py-(E-Ph)2-Rod (3)

Py-C02H (1)

Pyrene

, ~B—

(e, RT'cm'1) 383

(5.3 x 104) 399

(8.2 x 104) 351

(3.2 x 104) 335g

(4.9 x ioY

A.f, nm

425

445

390

375,

395s

V

0.33

0.41

0.34

0.72*

x, nse

3.1

1.8

10.1

190"

Kr, S (xlO8)

1.1

2.3

0.34

3.8*

*nr) S (xlO8)

2.2

3.3

0.65

1.5"

aAll measurements were performed in ^-saturated CH3CN at room temperature. 6 Absorption maximum ±2 nm. Corrected singlet emission maximum, ±5 nm. d

Fluorescence quantum yield, ±5%. eExcited-state lifetime ±2%. f From ref 30 (in EtOH). gThis paper (in CH3CN). h From ref 21. The data provided is for pyrene in polar solvents.

36

solutions yielded suspensions of rigid-rods that display an additional emission band in the

green. Black-light excitation with visual inspection showed that the solution emission

was blue and that the green emission was from undissolved rigid-rod particles (Figure

3.4b). The introduction of E-Ph units yielded a sharp, 100-fold decrease in the pyrene

excited state lifetime and was accompanied by an unexpected increase in fluorescence

quantum yield to nearly unity. Radiative decay is the main deactivating pathway since

the quantum yields of most of the pyrene complexes are over 90%.

The fluorescence spectra of 1-5 bound to ZrC>2 thin films, an insulating-like substrate

with similar morphology as semiconductor TiC>2, were concentration-dependent. Shown

in Figure 3.4a are the fluorescence spectra of Py-(E-Ph)2-ROD/Zr02 as a function of

surface coverage. With increased surface coverage, a red shift and a broadening of the

fluorescence spectrum is observed with the appearance of a shoulder at -550 nm. Figure

3.4a illustrates dependence of compound 4 on the surface coverage; at low surface

coverage conditions (2.2x10"9 mol/cm2), the monomer and a weak excimer (excited state

dimer) emission were observed at -425 and ~525nm, respectively. As the surface

coverage increased, an excimer emission band appeared. No evidence for surface

decomposition or desorption was found by UV-vis spectroscopic measurements of the

supernatant acetonitrile or of the nanoparticle thin film.

Pyrene excimer emission of 2-5 on TiC>2 was not detected suggesting electron injection

from the excimer. Because the Zr02, as an insulator, is used as a control for prohibiting

electron injection, the overlayed spectrum suggests that the excimers are also formed

upon adsorption to the TiC>2 surface. The molecular parallel, cofacial 7i-stacking to form

37

a) Py-(E-Ph)

Wavelength, nm

b)

Figure 3.4. a) Normalized emission spectra of Py-(E-Ph)2-Rod as a function of surface coverage on mesoporous Zr02 nanoparticle thin films immersed in acetonitrile at room temperature. The surface coverage from left to right was: 0.068, 0.16, 0.22, 0.25, and 1.6 x 10"8 mol/cm2. Superimposed on the data is the emission spectrum (+) of a saturated ethylene glycol solution of Py-(E-Ph)2-Rod. b) The green emission (left vial) represents the excimer emission of Py-(E-Ph)2-Rod in an ethylene glycol solution, while the Py-(E-Ph)2-Rod in neat acetonitrile, representing the violet monomer emission. The samples were excited with 350 nm light.

38

the excimer only exist at short-ranges (~ <3.6A)18, and therefore suggest that the pyrene

molecules are closely packed on the semiconductor surface. Similar behavior was found

for all pyrene complexes. In conclusion, the excimer of the pyrene complexes efficiently

inject into HO2, acting as a sensitizer.

Cyclic voltammetry data of 1-5 reveal an irreversible oxidation (Table 3.2). The formal

reduction potentials E° (Py+/0) were estimated by extrapolating the measured anodic peak

potential as a function of scan rate. The excited-state reduction potentials E° (Py+/0*)

were estimated from these data and the free energy of the fluorescent excited state, AGes,

as previously described.9'19 The irreversible nature of this system can be explained by the

association of the resultant 7t-radical cation with a ground state molecule to form a

dimmer radical cation. Compounds 2-5 exhibit a more negative anodic peak potential

(Epa) than that found for either pyrene or Py-COOH, thus suggesting an increase in the

energy of the highest occupied molecular orbital (HOMO) of these compounds with

increasing conjugation. This trend is visible in the Epa potentials of compounds 1-3 that

display Epa potentials of 1.25, 1.21, and 1.16V respectively vs. Fc/Fc+.

Nanosecond transient absorption spectra of the pyrene-substituted rods in fluid solution

revealed a long-lived excited state assigned to the triplet state. The triplet yield was low

and could be increased by adding Ru(bpy)3(PF6)2 as a sensitizer.20 Pulsed light excitation

of 1-3 bound to Ti02 resulted in the appearance of a new transient absorption feature.

Figure 3.5 shows representative transient absorption spectra observed after pulsed 417-

nm light excitation of Py-(E-Ph)2-Rod/Ti02 at the indicated delay times and, for

comparison, the excited-state triplet difference spectra measured in acetonitrile solution.

39

Table 3.2: Electrochemical Properties of 1-3°

sensitizer

Py-E-Ph-Rod (2)

Py-(E-Ph)2-Rod (3)

Py-C02H (1)

Pyrene

E< w\ 1.18

1.15

1.29

1.16"

V E* '(Py+y°*),

-2.03

-1.97

-1.99

-2.17c

V AGes, eV

3.21

3.12

3.28

3.33c

"Formal reduction potentials reported versus SCE estimated from cyclic voltammetry data obtained by adding 1-3 dissolved in a minimal amount of CH2C12 to 0.1 M TBACIO4/CH3CN. b From ref 21, performed in CH3CN. c Calculated using data provided in ref 21 for pyrene in polar solvents.

40

-0.006 - i — i — i — i — i — i — i — i — i — i — i — i — i — i — i — i

350 400 450 500 550 600 650 700 750

Wavelength, nm

Figure 3.5. AA spectrum observed after 417-nm laser light excitation (-3.1 mJ/cm2, 8 ns fwhm) of Py-(E-Ph)2-Rod/Ti02 immersed in CH3CN. The data were recorded at 10 ns (•), 100 ns (red • ) , 250 ns (green A), and 1 (is (blue • ) delays after the laser pulse (solid lines with symbols). Overlaid is the normalized absorption difference spectrum obtained 10 (is after sensitizing the triplet of Py-(E-Ph)2-Rod (4.0 x 10"5 M) in CH3CN using [Ru(bpy)3](PF6)2 (4.0 * 10"5 M) and exciting with 532-nm laser light (-11 mJ/cm2, 8-ns fwhm) ( - • - • - • -). Inset: Transient absorption signals following 417-nm laser light excitation of Py-(E-Ph)2-Rod/Ti02

immersed in 1.0 M LiGCVCHaCN monitored at 640 nm and displayed on a logarithmic time scale from 10"8 to 0.5 s. Overlaid on the data is a kinetic fit (white line) to a sum of two second-order rate constants.

41

The inset shows single-wavelength kinetic traces. The formation of the transient state

could not be time resolved, and the recovery required milliseconds for completion. The

transient data was well described by a bi-second-order kinetic model.9 Average rate

constants of (2.5 ± 0.3) x 107 s"1 were measured for 2 and 3 at surface coverages of 7.9 x

10"9mol/cm2.

The photocurrent efficiency of the rods was quantified in a sandwich-cell arrangement

with a 0.5 M LiI/0.05 M I2 acetonitrile electrolyte. For 1-3, conditions were found where

the absorbed photons were converted to electrons with efficiencies within the

experimental error of unity. The photocurrents were stable for periods of hours with

negligible degradation under illumination conditions of ~1 mW/cm2.

3.4. Discussion.

The pyrene-terminated rigid-rod linkers bind to TiCh and ZrC>2 mesoporous thin films in

St 9

high surface coverages, -10" mol/cm . These values are comparable to those reported for

tripodal linkers with Ru(II) complexes and other inorganic complexes.9'21 The linkages

were robust, and decomposition of the surface-bound complexes was not observed on the

few hours time scale of the experiments. Strong electronic coupling between the pyrene

and the rigid-rod is clearly evident in the enhanced extinction coefficient and by a red

shift in the absorption spectrum as the number of phenylethyne groups is increased

(Figure 3.2a and 3.3a). The rigid-rods therefore increase the visible-light absorption by

pyrene, an important feature for solar energy conversion applications.

42

Semi-empirical calculations show the presence of a very pronounced derealization of the

LUMO orbital of the pyrene chromophore onto the rigid-rod in 2 and 3 (Figure 3.5). The

implications of this finding are two-fold. First, it suggests that a strong coupling, and

consequently fast electron transfer between the sensitizer and the semiconductor particle,

will exist even as the length of the rigid-rod increases. Second, it agrees with the

experimentally observed enhanced extinction coefficients and the spectral red shifts that

accompany the extension of the rod in 2 and 3. Indeed, in a similar agreement with

experiment, a 10 x 10 AMI-CI calculation predicts a nearly two-fold increase of the

oscillator strength in 3 in comparison with that of the unmodified 1.

Pyrene fluorescence ' ' has previously been used as a probe on titanium dioxide

surfaces24 and on sol-gel processed materials.25 Fluorescence on the HO2 nanocrystallites

studied here was highly quenched but was easily observed on insulating Zr02

nanoparticles. The emission spectra on ZrC>2 were concentration-dependent and displayed

a significant broadening and red shift with increased surface coverage (Figure 3.4).

Previous studies of pyrene physisorbed to Ti02 powders showed sharp fluorescence

spectra consistent with pyrene in a polar, protic-like environment.24 The broadening

observed for the rigid-rods was present at ~Vio of the maximum surface coverage,

suggesting some degree of pyrene aggregation on the nanoparticle surface. At surface

coverages near saturation, a shoulder was observed near the wavelength expected for the

pyrene excimer. This data indicates that the rigid-rod linkers inhibit pyrene-pyrene

interactions but do not fully eliminate them. Investigating whether the use of a linker

with a larger footprint, such as the tripod, eliminates the formation of exciplexes and

other aggregation phenomena is discussed in the following chapter.

43

a)

&

b)

P Figure 3.6. The LUMO (a) and the HOMO (b) orbitals of the pyrene chromophore in 1 - 3 derived from semi-empirical calculations. Note the pronounced derealization onto the rigid rod in 2 and 3.

44

Transient absorption and photoelectrochemical measurements clearly indicate that the

fluorescence quenching on HO2 is due to interfacial electron injection. The pyrene rigid-

rods are potent excited-state reductants (E° (Py+I0*) fa-1.9 V vs SCE), and interfacial

electron injection into the semiconductor is expected to be thermodynamically

favorable.21 Transient absorption spectroscopy reveals the presence of a single

photoproduct with an absorption spectrum clearly different than that expected for the

pyrene triplet, which is reasonably assigned to an interfacial charge-separated state,

Py+/Ti02(e"). The appearance of this product could not be time resolved for either of the

rigid-rods or Py-CC^H, consistent with rapid electron injection into the semiconductor,

kmi > 108 s"1.

Organic dyes strongly coupled to TiC>2 surfaces often display dye-to-particle charge-

transfer absorption bands.26'27 The overlap of pyrene and Ti02 absorption makes it

difficult to determine whether new electronic transitions of this type are present in the

rigid-rods. However, a broadened peak near 400 nm is observed in the photocurrent

action and absorption spectra of 2 and 3 anchored to TiC>2 that is approximately the same

energy as the lowest vibronic transition in the pyrene absorption spectrum. This spectral

feature, coupled with the long distance between the pyrene and the semiconductor

surface, is most consistent with injection from a pyrene excited state. Recombination of

the injected electron with the oxidized pyrene required milliseconds for completion, and

the kinetics were well described by a sum of two second-order rate constants.9 Average

rate constants for rigid-rods 2 and 3 at ~Vio of the maximum surface coverage were

within experimental error the same. This is not unexpected in view of literature

45

reports and the fact that semi empirical calculations show significant derealization

over the phenylethyne linker in the HOMO of rigid-rods 2 and 3.

The pyrene rigid-rods efficiently convert light into an electrical current in regenerative

solar cells with iodide as the electron donor.27 Energetically, a high photocurrent is

expected since the pyrene is a potent photoreductant and the pyrene radical cation is a

strong oxidant that is expected to oxidize iodide rapidly. In fact, when corrections were

made for transmitted light and for losses associated with the tin oxide substrate, the

absorbed photon-to-current efficiency was within experimental error of unity for both

rigid-rod linkers. High photocurrent efficiencies were observed at surface coverages

where excimer emission was evident on Zr02 surfaces, suggesting that either injection

occurs prior to excimer formation or that the excimer itself injects electrons into Ti02

efficiently.

3.5. Conclusions.

A study of rigid-rod molecules that can be anchored to metal oxide nanoparticles with

high surface coverages is reported. The conjugated phenyethynyl spacer of the rigid-rods

enhances the pyrene extinction coefficient and shifts the absorption spectrum toward the

red. The spectroscopic data indicates that the rigid-rods inhibit translational mobility on

the nanoparticle surface, but the observation of excimer-like emissions indicates that the

pyrene groups are not isolated. Light excitation of the rigid-rods on anatase Ti02

nanocrystallites reveals rapid and quantitative electron injection into the semiconductor.

46

This behavior may be useful for light detection and conversion at selected frequencies of

light. Furthermore, if this efficiency can be enhanced to longer wavelengths of light, then

rigid-rod chromophores would be expected to convert sunlight into electrical energy

efficiently.27 More fundamentally, pyrene rigid-rods, with their well-defined spin states,

vibronic structures, excimer emissions, and redox properties, provide unique insights into

interfacial electron transfer and excited states that are difficult to obtain from the more

commonly studied inorganic Ru(II) dyes.21'30

•Professor L. Brand and Dr. T. Dmitri at The Johns Hopkins University Biology Department allowed us to

use and technically assistanted with the single-photon counting apparatus.

47

3.6. References

1. (a) Ghosh, P.; Spiro, T. G. J. Am. Chem. Soc. 1980, 102, 5543-5549. (b)

Bookbinder, D. C; Wrighton, M. S.J. Electrochem. Soc. 1983, 130, 1080-1086.

(c) Miller, C. J.; Widrig, C. A.; Charych, D. H.; Majda, M. J. J. Phys. Chem.

1988, 92, 1928-1933. (d) Ford, W. E.; Rodgers, M. A. J. Phys. Chem. 1994, 98,

3822-3841.

2. Anderson, S.; Constable, E. C; Dare-Edwards, M. P.; Goodenough, J. B.;

Hamnett, A.; Seddon, K. R.; Wright, R. D. Nature (London) 1979, 280, 571-573.

3. Heimer, T. A; D'Arcangelis, S. T.; Farzad, F.; Stipkala, J. M.; Meyer, G. J. Inorg.

Chem. 1996, 35,5319-5324.

4. (a) Moses, P. R.; Murray, R. W. J. Electroanal. Chem. 1977, 77, 393-400. (b)

Shepard, V. R.; Armstron, N. R. J. Phys. Chem. 1979, 83, 1268-1276. (c) Fox, M.

A.; Nabs, F. J.; Voynick, T. A. J.Am. Chem. Soc. 1980,102, 4036-4042.

5. Zou, C; Wrighton, M. S. J. Am. Chem. Soc. 1990,112, 7578-7586.

6. (a) Pechy, P.; Rotzinger, F. P.; Nazeeruddin, M. K.; Kohle, O.; Zakeeruddin, S.

M.; Humphry-Baker, R.; Gratzel, M. Chem. Commun. 1995, 65-66. (b) Yan, S.

G; Hupp, J. T. J. Phys. Chem. 1996, 100, 6867-6870. (c) Saupe, G. B.; Mallouk,

T. E.; Kim, W.; Schmehl, R. H. J. Phys. Chem. B 1997, 101, 2508-2513. (d)

48

Gillaizeau-Gauthier, I.; Odobel, F.; Alebbi, M.; Argazzi, R.; Costa, E.; Bignozzi,

C. A.; Qu, P.; Meyer, G. Inorg. Chem. 2001, 40, 6073-6079.

7. Asbury, J. B.; Hao, E.; Wang, Y.; Lian, T. J. Phys. Chem. B 2000, 104, 11957-

11964.

8. Hoertz, P; Carlisle, R.; Meyer, G.; Wan, D; Piotrowiak, P.; Galoppini, E.

Nanoletters 2003, 3, 325.

9. a) Galoppini, E.; Guo, W.; Hoertz, P.; Qu, P.; Meyer, G. J. J. Am. Chem. Soc.

2002, 124, 7801-7811. b) Galoppini, E.; Guo, W.; Qu, P.; Meyer, G. J. J. Am.

Chem. Soc. 2001, 123, 4342-4343.

10. Wang, D.; Schlegel, J. M.; Galoppini, E. Tetrahedron 2002, 58, 6027-6032.

11. O'Regan, B.; Moser, J.; Anderson, M.; Gratzel, M. J. Phys. Chem. 1990, 94,

8720-8726

12. Tartula, O., Rochfors, J.Piotrowiak, P.; Galoppini, E.; Carlisle, R.; Meyer, G.J.; J

Phys Chem B 2006, 110, 15734-15741.

13. Wang, D.; Mendlesohn, R.; Galoppini, E.; Hoertz, P.; Carlisle, R.A.; Meyer, G.J.;

J Phys Chem B 2004, 108, 16642-16653. Kelly, C. A.; Thompson, D. W.; Farzad,

F.; Meyer, G. J. Langmuir 1999, 15, 731-737.

14. Langmuir, I.; J Am Chem Soc 1918, 40, 1362-1403.

49

15. Kelly, C.A.; Thompson, D.W.; Farzad, F.; Meyer, G.J.; Langmuir 1999, 15, 731-

737.

16. Dmitri, T.; Savtchenko, R.; Meadow, N.; Roseman, S.; Brand, L. J. Phys. Chem.

B 2002, 106, 3724-3734.

17. Lakowicz, J. R. Principles of Fluorescence Spectroscopy. 2nd ed.: Plenum Press:

New York, 1999.

18. Birks, J.B. Photophysics of Aromatic Molecules. John Wiley & Sons: London,

UK. 1970; pp301-307.

19. Rehm, D.; Weller, A. Isr. J. Chem. 1970, 8, 259. 17. de Carvalho, I. M. M ;

Gehlen, M. H. J. Photochem. Photobiol, A 1999, 122, 109.

20. Qu, P.; Meyer, G. J. Dye Sensitized Electrodes. In Electron Transfer in

Chemistry; Balzani, V., Ed.; Wiley-VCH: Weinheim, Germany, 2001; Vol. 4,

Chapter 2, pp 355-411.

21. Murov, S. L.; Carmichael, I.; Hug, G. L. Handbook of Photochemistry, 2nd ed.;

Marcel Dekker: New York, 1993.

22. (a) Forster, T. Angew. Chem., Int. Ed. Engl. 1969, 5, 333-343.

23. (a) Chandrasekaran, K.; Thomas, J. K. J. Am. Chem. Soc. 1983, 105, 6383-6389.

(b) Chandrasekaran, K.; Thomas, J. K. J. Colloid Interface Sci. 1985, 106, 532-

537.

50

24. Avnir, D. Ace. Chem. Res. 1995, 28, 328-334.

25. Moser, J.; Punchihewa, S.; Infelta, P.; Gratzel, M. Langmuir 1991, 7, 3012-3018.

26. Efficient sensitization by other organic dyes has been reported: (a) Enea, O.;

Moser, J.; Gratzel, M. J. Electroanal. Chem. 1989, 259, 59-65. (b) Ferrere, S.;

Gregg, B. New J. Chem. 2002, 26, 1155-1160.

27. Hasslemann, G. M.; Meyer, G. J. J. Phys. Chem. B 1999, 103, 7671-7675.

28. (a) Nelson, J. Phys. Rev. B 1999, 59, 1537'A. (b) Nelson, J.; Haque, S. A.; Klug, D.

R.; Durrant, J. R. Phys. Rev. B 2001, 63, 205321.

29. (a) Fiebig, T.; Kuhnle, W.; Staerk, H. Chem. Phys. Lett. 1998, 282, 7-15. (b)

Fiebig, T.; Stock, K.; Lochbrunner, S.; Riedle, E. Chem. Phys. Lett. 2001, 345,

81-88. (c) Pandurski, E.; Fiebig, T. Chem. Phys. Lett. 2002, 357, 272-278.

30. Jones, N. J. Am. Chem. Soc. 1945, 67, 2127-2150

51

Chapter 4. Control of Intermolecular "Cross-Talk" between

Chromophores on Nanocrystalline Surfaces through the Isolation of

Pyrene Rigid-Rod Sensitizers

In addition to the bipodal rigid rod linkers, a "tripodal" linker was developed at Rutgers

to allow for chromophores to adsorb to the metal oxide surfaces in a perpendicular

orientation, preventing the possible pivoting action of the rigid bipodal systems. Figure

4.1 illustrates the structural differences between the "bipods" and "tripods." This figure

depicts the sensitizer as having carboxylic acid functional groups for metal oxide

adsorption. This tripod-shaped linker carrying a pyrene chromophore on a phenylethynyl

arm and three COOH binding groups was synthesized to study how the size of the linker

footprint can influence the aggregation of organic dyes bound to metal oxide thin films.

The tripod has an adamantane core and a footprint of -52.4 A1 spanned by the three p-

carboxy-phenyl arms, see Figure 4.2.

4.1. Introduction.

A vast and diverse selection of photoactive molecular compounds referred to as

chromophores has been extensively investigated when bound to metal oxide surfaces.

Later in this dissertation, the use of anchored Ru(II) polypyridyl complexes to

52

Figure 4.1. Illustration of the bipodal and tripodal linkers bound to metal oxide (MO) nanoparticle surfaces. The linkers consist of a p-phenyleneethynylene spacer of varying length, connecting the chromophore to the anchoring group, a) Depicts the tripodal linker with three COOH groups for attachment, b) Represents the bipodal linker with two COOH groups for attachment.

53

Py-(E-Ph)-TRIPOD

Figure 4.2. The tripodal pyrene-spacer-anchoring system. The phenylenethynylene spacer is abbreviated (E-Ph). The ball-and-stick-model illustrates the footprint size from a "top-down" view.

*Footprint-size calculated bv GaloDDini and coworkers.

54

nanocrystalline TiC>2 through rigid-rod linkers is reported.1'2 In addition, I have

commented on the use of organic rigid-rod linkers for coupling pyrene chromophores to

metal oxide nanoparticles.3 These inflexible linkers included a varying number of

sequential phenylethyne units also termed "spacers," Figure 4.2. The pyrene tripod

studied herein, consisted of only a slightly insulating adamantine core and was found to

achieve maximum binding through three carboxylic acid functional groups (r~10"7mol

cm"2).

This novel rigid tripodal design ultimately allows for a perpendicular binding orientation

to the metal oxide surface, manipulation of the chromophore head-to-surface distance, an

increase and red shift in the molar extinction coefficient spectrum, and a larger

"footprint" binding site.

Organic dyes are interesting candidates for solar cell sensitizer applications due to their

tenability and absorption range. Pyrenes4, in addition to perylenes,5 anthracenes,6

porphyrines7 and other organic dyes with polyaromatic ring systems, have a tendency to

aggregate on TiC>2 nanoparticle surfaces. These excimers or exciplexes alter the

electronic levels and photophysical properties of the monomelic complex.

Pyrene's ability to form an excited state dimer was first reported by Forster and Kaspar8

and has been further investigated by Birks and co-workers.9 The excimer has become a

valuable means for studying molecular proximities and has facilitated our studies of

sensitizer-to-sensitizer distances. The pyrene excimer is an excited state complex that

55

has a well-defined and well-structured emission; therefore, studying intermolecular

"cross-talk" between pyrene sensitizers has become a primary focus. Previously, pyrene

rigid-rod excimer emission as a function of surface coverage has been examined; as the

concentration of pyrene rigid-rod increased, excimer emission increased.

Here, I report the results of the pyrene tripodal sensitizer, trimethyl (5-(7-(l-

pyrenylethynyl-phenyl)-l,3,5-triphenyl adamantine [Py-E-Ph-TRIPOD], Figure 4.2; used

in lieu of the rigid-rod linker primarily to explore intermolecular "cross-talk" by

attempting to prevent excimer formation. A larger footprint of 52.9 A2 allows the pyrene

tripods to sit at a fixed distance on the metal oxide surface to ensure the distance between

the pyrene chromophore heads are greater than 3.5A10 from each other; 3.5A is the

maximum distance at which two proximate pyrenes will form an excimer. This will

ultimately provide us with the insight required to measure the sensitizer-sensitizer

distances on nanocrystalline surfaces and the corresponding molecular interfacial

processes.

Methods for increasing sensitizer-sensitizer distances have been previously reported in

the literature; bulky substituents like t-butyl groups11 have been added to the

chromophore heads while methods of coadsorption of hydrocarbon chains12 to surfaces

have been employed, Figure 4.3. Tripodal spacers have been chosen as an alternate

means of preventing chromophore aggregation. This approach has advantages that the

other methods lack. The tripodal sensitizer surface coverage remains relatively high

compared

56

CH3 CH3

Bulky Substituents Co-Adsorption Tripodal Linker

Figure 4.3. Three plausible techniques for eliminating excimer formation: the addition of bulky substituents to the chromophore head group, dilution of the sensitizer on the surface through the co-adsorption of lengthy hydrocarbon chains, synthetically altering the footprint size of the sensitizer through the addition of a three point linker.

57

to the diluted surface of the chromophore/co-binder. Also, there is no need for further

alteration of chromophore with oversized substituents.

4.2. Experimental Section.

Metal Oxide Thin Film Preparation and Binding.

Nanocrystalline TiCh and Zr02 thin films were prepared on biological glass slides (VWR)

for absorption and photoluminescence experiments in a manner similar to that previously

reported.314 For photoelectrochemical studies, the films were deposited on fluorine

doped tin oxide 15Q conductive glass (Libbey-Owens-Ford). The colloidal sol-gel

technique produced transparent mesoporous thin films of approximately lO-um

thickness. Films that were acid pretreated were exposed to a pH = 1 aqueous sulfuric acid

solution while films that were basic pretreated films were submerged in a pH = 11

aqueous sodium hydroxide solution. After several minutes of exposure to pretreatment

solutions, the slides were rinsed thoroughly with neat acetonitrile and then immersed in

the pyrene sensitizer acetonitrile solution.

Adsorption Isotherms.

Surface binding of 2-5 was monitored spectroscopically by measuring the change in film

and solution absorbance after equilibrating the film overnight in solutions with known

concentrations of the sensitizers. In all cases, the surface coverages saturated at an

~0.25mM sensitizer concentration. The equilibrium binding for all sensitizers were

58

described by the Langmuir adsorption isotherm model from which the surface binding

constants (K^) were abstracted using equation (3.1) stated in chapter 3. Plots of [S]eq/ T

versus [S]eq were fitted linearly to obtain the binding constants K^ and surface coverages

r0.

Spectroscopy.

Absorption spectra were obtained at ambient temperature in neat acetonitrile (Burdick

and Jackson, spectroscopic grade) with a Varian Cary 50 UV-visible spectrophotometer.

Steady-state fluorescence spectra were acquired using a Spex Fluorolog that had been

calibrated with a standard NIST tungsten-halogen lamp.

Incident Photon-to-Current Efficiency (IPCE).

The photocurrent action spectra were obtained in a two-electrode sandwich cell

arrangement similar to figure 2.3. A 0.5M LiI/0.5M I2 acetonitrile solution was used as

the electrolyte. Quasi-monochromatic light from 400 to 600 nm was achieved with a 150

W Xe lamp coupled to a f/2 monochrometer, and incident irradiances were typically of

lmW/cm2.

4.3. Results.

The absorbance spectra of the pyrene tripod in neat acetonitrile, exhibits a significant

increase in the molar extinction coefficient compared to that of the commercially

available 9-pyrenecarboxylic acid (Aldrich), Figure 4.4a. In addition, there is a

59

a)

8.0x1(TH

^H 6.0x10

E

Py-C02H

Py-E-Ph-ROD Py-(E-Ph)2-ROD

-+-Py-E-Ph-TRIPOD

u

4.0x10

2.0x104

b)

8 c es s -

o XI

350 400

Wavelength, nm

-Py-C02H

Py-E-Ph-ROD Py-(E-Ph)2-R0D

-Py-E-Ph-TRIPOD

^ ' | ~TlTiiT^iin-rgii^ii|-iiiTBiiiin

380 400 420 440 460

Wavelength, nm

480

Figure 4.4. a) The extinction coefficient of the pyrene bipods 1-3 (discussed in Chapter 3) and of the Pyrene-(E-Ph)-TRIPOD. The Tripod is comparable in extinction coefficient and in the red-shift of the absorbance band to that of the Py-E-Ph-ROD. b) The absorbance spectra of pyrene bipods 1-3 and of the Pyrene-(E-Ph)-TRIPOD on TiC«2. The fundamental absorption band of the TiCh is obscured by the absorption of the sensitizers 2-3 and the tripod.

60

noticeable red shift of the low energy absorption band of the pyrene tripod. When bound

to metal oxide thin films, the high-energy absorption bands (7t->7i* transitions) observed

in solution are masked by the overwhelming fundamental absorption of the

semiconductor, Figure 4.4b. Absorbance spectra for the tripod on Zr02 are within

experimental error the same as those for the sensitizer on TiC>2. Table 4.1 shows the

photophysical properties of the Py-E-Ph-TRIPOD in solution and on the surface (ZrC>2

and TiCh).

It is known that at low concentrations of pyrene compounds in viscous solution2'3

and on ZrC>2,3 a single structured emission band is observed at room temperature due to

the fluorescence of the pyrene monomer. Figure 4.5 exhibits overlayed absorbance and

emission spectra of Py-E-Ph-TRIPOD in CH3CN. Furthermore, a second stuctureless

emission band appears at longer wavelengths (between 505 and 550nm) as the surface

coverage (T) is increased. This is caused by the aggregation of sensitizer molecules and

excimer formation. However, unlike the pyrene rigid-rods, when bound to Zr(>2, an

increase in the pyrene tripod surface coverage does not lead to either a large decrease in

monomer fluorescence or to a significant increase in the excimer emission, Figure 4.6.

There is only a negligible increase in the emission around 525nm due to the substantial

increase in monomer fluorescence intensity at 418nm. Furthermore, there was not a

decrease in the monomer band as seen for Py-(E-Ph)2-ROD, Figure 4.6a. To find the

exact increase in the monomer/excimer ratio, each band must be deconvoluted, Figure

4.6b. To fully understand the pyrene tripod emission on Zr02, the pyrene rigid-rods were

used for comparison. In contrast to the pyrene tripod, it was previously found that

61

Tab

le 4

.1.

Phot

ophy

sica

l Pro

pert

ies

of P

y-E

-Ph-

TR

IPO

D in

Sol

utio

n an

d on

the

Surf

ace.

SEN

SIT

IZE

R

Py-

(E-P

h)-T

RIP

OD

A. a

bs, n

m

(e, M

em)

384

(4.8

x 1

04)

A. P

L, n

m

mon

omer

415

A. P

L, n

m

exci

mer

N/A

T, n

s (k

m

kr, s

^ m

onom

er

(xlO

)

2.93

0.

098

0.33

knr,

s"1

(xlO

8)

3.1

On

Zr0

2

Py-

(E-P

h)-T

RIP

OD

A, a

b s,

nm

383

A, P

L, n

m

mon

omer

425

A, P

L, n

m

exci

mer

515

T, n

s T

, ns

mon

omer

ex

cim

er

2.72

11

.8

@49

5 nm

@

495

nm

^a

d

1.48

x

103

On

Ti0

2 A,

abs

» nm

A,

PL

, nm

m

onom

er

A- P

L, n

m

exci

mer

8 F

ootp

rint

D

ista

nce

to

surf

ace

Py-

(E-P

h)-T

RIP

OD

39

4 42

2 •5

2.4

A

12.4

A

(0 c 0)

a> o c a) o 0)

<u o

U.

1.6x106-

1.4x106-

1.2x106-

1.0x106-

8.0x105-

6.0x105-

4.0x105-

2.0x105-

0.0-

i

I ' - '' / I i'i ,' ' ;! / I

•. i i ' '• i i 1 1 ' . ; ' i ' ; i 1 1

; i f ! • i ' i

f' • i ' J ' i i !> • i ' ' ' /' i

i / \i i / l ' !

• / ! .' ;

/'

/ / 1 i 1

[ i |

" /

A \ \

\

i \ i \

— . 1 • 1 • / I s — • 1 . 1 • 300 350 400 450

Wavelength, nm

500

- 0.4

0.3

0.2

0)

o c n i_ o <n si <

0.1

0.0 550

Figure 4.5. The structures absorbance (—) and emission (—-) spectra of the Pyrene-E-Ph-TRIPOD in CH3CN.

63

a) 1.0x106

Inte

nsity

ce

nce

rest

o 3

8.0x10s-

6.0x10s-

4.0x10s-

-

2.0x10s-

0.0

Surface Coverage 2.45e-9 2.35e-8 2.31 e-7 3.69e-7 4.79e-7

450 500 550

Wavelength, nm

600 650

b)

E '5 x UJ "C Q) E o c o

1 6 -

1 4 -

12

10- |

8

6

4

2-T

0.00 1.50X10'7 3.00x10"7 4.50x10'

Surface Coverage 6.00x10'7

Figure 4.6. a) Emission dependence on surface coverage of the Py-(E-Ph)-TRIPOD on ZrC>2. As the surface coverage increases, the monomer grows in and the excimer emission remains relatively the same, b) A plot depicting the ratio of fluorescence intensity of the monomer : excimer.

64

excimer emission bands of Py-(EPh)n-ROD3 on Zr02 intensified as the sensitizer surface

concentration increased, Figure 3.4.

The Py-Tripod was found to have an increased surface coverage (T ~10"7 mol cm"2)

compared to that of the pyrene rigid-rods (T ~10'8 mol cm"2) previously described in

chapter 3. This difference may be due to the three-point attachment (three COOH rather

than two COOH binding groups) and increase in affinity of the Py-Tripod to the thin film

surfaces. IR data indicate that the tripodal complex binds through all three carboxylic

acid groups and is therefore normal to the surface (not shown).

Upon further studies of the pyrene tripod on Ti02 and upon comparison to that of the

pyrene rigid-rods on Ti02 thin films, it is realized that the excimer emission band

intensity for both models of the pyrene sensitizers have weakened, suggesting an

injection of electrons from the excited state. Both compounds on Ti02 exhibit neither

excimer emission nor a decrease in monomer emission intensity. No evidence for surface

desorption or decomposition was found by UV-vis spectroscopic measurements of the

nanocrystalline thin films.

4.4. Discussion.

The pyrene tripod was found to have an increased surface coverage (T-IO"7 mol cm"2) on

the metal oxide thin films compared to that of the pyrene rigid-rods. This may be due to

65

the increased number of carboxylic acid functional groups of the tripod linker. It is

assumed through IR data that all three carboxylic acid groups bind to the nanocrystalline

surface and therefore, the tripod sensitizer must be perpendicular to the surface; only a

single peak appears in the IR spectrum.

Fluorescence from pyrene in solution and as a probe on HO2 surfaces and on sol-gel

processed materials have been studied.3 However, a new pyrene tripodal sensitizer

design was used to control "cross-talk" between sensitizers. It is reported herein that

photoluminescence intensity of the pyrene excimer is decreased significantly for the

pyrene tripod on ZrC>2 because of its larger footprint. This novel tripodal design has

deterred the pyrene-pyrene interactions by increasing the distance between the pyrene

chromophore heads of adjacent sensitizers. Although the tripodal shape has been used

before in Ru sensitizers14, its purpose was not to unveil the sensitizer-sensitizer distances.

Here, it is known that for pyrene excimer formation to exist, the pyrene chromophores

must be at a maximum distance of 3.5A.10

Figure 4.6a illustrates the monomer and excimer photoluminescence dependence on

surface coverage. As T is increased, the monomer fluorescence increased considerably,

while the excimer emission displayed no appreciable change. Despite the static excimer

intensity, it was found that excimer formation could not be completely eliminated by the

larger footprint design. Figure 4.7 depicts the possible binding sites of the pyrene tripod.

At all surface coverages, sensitizers bind in the "necking" regions or pores of the

nanocrystalline films. It is plausible that adjacent pyrenes on these neighboring particles

66

Figure 4.7. A schematic depicting a mesoporous nanoparticle TiC>2 thin film sensitized with Py-(E-Ph)-TRIPOD. The red arrows illustrate the necking regions where the 7t-stacking may occur by neighboring sensitizer molecules, acting as the source of the expected excimer.

67

interact and form excited state dimers. Moreover, as T is increased, the large tripodal

footprint inhibits additional excimer formation in the non-necking regions of the thin

films, therefore increasing the overall monomer emission intensity.

It was observed that the excimer emission disappeared when the pyrene sensitizer was

bound to Ti02. Since ZrC>2 nanoparticles were used to make insulating thin films, there

was no electron injection or charge transfer from the pyrene to the film. HO2 thin films,

on the other hand, are used to study interfacial charge transfer because of its

semiconducting properties. The disappearance of the excimer photoluminescence band

on Ti02 must be due to the injection from the pyrene excimer. Figure 4.8 shows the

quenching of the photoluminescence on TiC>2 with the addition of LiGC>4. Li+ cations are

known to affect the quantum yield for injection; this process is completely reversible.13'15

4.5. Conclusion.

Tripodal linkers with large footprints may be useful to eliminate aggregation and

interactions between the chromophoric groups by increasing the distance between

sensitizer molecules.

To further characterize the theory of the excited state dimers forming in the necking

regions of the mesoporous nanoparticle thin films, similar measurements can be made on

planar insulating substrates. The excimer formation should be completely eliminated and

therefore only a single higher energy peak will appear in the visible emission spectrum

68

W/1.0M LiCIO. NeatCH3CN

111 1x105-|

450 500 550 600 650 700 750

Wavelength, nm

Figure 4.8. CH3CN (—

Py-(E-Ph)-Tripod on T1O2 in the presence of neat -) and in the presence of 1.0M LiCL04/CH3CN( ).

The cation affect contributes to electron injection into the Ti02 depicted by a decrease in the emission intensity in the presence of LiC104.

69

corresponding to the Py-Tripod monomer. However, with planar films, high surface

coverages usually cannot be achieved. Further experiments can be completed on planar

T1O2 to study electron injection properties.

Larger footprint pyrene tripods have already been synthesized at Rutgers by the

Galoppini group that will enable us to study charge injection efficiency as a function of

footprint area, of the length of the spacer, and of the amount of conjugation from the

chromophore to the surface.

70

References.

Galoppini, E.; Guo, W.; Qu, P.: Meyer, G.J. J. Am. Chem. Soc. 2001, 123,

4342-4343.

Galoppini, E.; Guo, W.; Hoertz, P.; Qu, P.: Meyer, G.J. J. Am. Chem. Soc. 2002,

124, 7801-7811.

a) Hoertz, P.; Carlisle, R.A.; Meyer, G.J.; Wang, D.; Piotrowiak, P.; Galoppini,

E. Nano lett. 2003, 3(3); 325-330. b) Taratula,0.; Rochford, J.; Piotrowiak, P.;

Galoppini, E.; Carlisle, R.A.; Meyer, G.J. Phys. Chem. B, 2006, 110 (32), 15734

-15741,

a) Khakhel, O.A. J. App. Spec. 2001, 62 (2), 280-286. b) Jones, G; Vullev,

V.I. J. Phys Chem A. 2001, 105, 6402-6406.

a) Mo, X.; Chen, H.Z.; Shi, M-M.; Wang, M. Chem. Phys. Lett. 2006, 417(4-6)

457-460.

Nishimura, T.; Nokashima, N.; Mataga, N; Chem.Phys. Lett. 1977, 46 (2), 334-

338.

71

a) Patemack, et. Al. Inorg. Chem. 1955, 33, 2062-2065. b) Koehrst, R.B.;

Hofstra, U.; Schaafsma, T.J.; Magnetic Resonance in Chemistry 2005, 26 (2),

167-172.

a) Kaspar; Forster Z Elektrochem 1955, 59976-59980. b) Forster, T. Agnew

Chem, Int. Ed. Engl. 1969, 8, 333-343.

Birks, J.B. Photophysics of Aromatic Molecules. John Wiley & Sons:

London, UK. 1970; pp301-307.

Lehrer, S. Subcellular Biochemistry, Volume 24. Proteins: Structure, Function,

and Engineering. B.B.Biswas and Siddhartha Roy, ed. Plenum Press: New

York, 1995.

Barfeindt, B.; Hannappel, T.; Storck, W.; Willig, W.F. JPhys Chem 1996, 100,

16463-16465.

a) Turro, N.J.; Lakshminarasimhan, P.H.; Jockusch, S.; O'Brien, S.P.;

Grancharov, S.G.; Redl, F. Nanoletters 2002, 2 (4), 325-328. b) Ipe, B.I.;

Thomas, K.G.; Barazzouk, S.; Hotchandani, S.; Kamat, P. J. Phys. Chem. B

2002,106, 18-21.

72

12. a) Heimer, T.A.; D'Arcangelis, S.T.; Farzad, F.; Stipkala, J.M.; Meyer, G.J.

Inorg Chem 1996, 35, 5319-5324, b) Langmuir, I. J Am Chem Soc 1918, 40,

1361-1403.

13. Wang, D.; Mendelsohn,R.; Galoppini, E.; Hoertz, P.G.; Carlisle, R.A.; Meyer,

G.J. Phys. Chem. B, 108 (43), 16642 -16653, 2004

14. Kelly, C.A.; Farzad, F.; Thompson, D.W.; Stipkala, J.M.; Meyer, G.J. Langmuir

1999, 15, 7047-7054.

73

Chapter 5. Excited State Electron Transfer from Ru(II) Polypyridyl

Complexes Anchored to Nanocrystalline Ti02 through Rigid-Rod

Linkers

Despite pyrene's unique photophysical properties and excimer emissions, solar cell

efficiencies will be increased with metal-centered ruthenium complexes due to their

abilities to absorb a wider range of the visible spectrum. Furthermore, ruthenium

polypyridyl complexes are easily synthesizable to achieve desired electronic and

thermodynamic properties. In addition, adding these highly conjugated spacers,

described in previous chapters herein, increase the molar extinction coefficient.

Rigid-rod linkers varying in length were used to bind Ru(II) polypyridyl complexes to the

surface of Ti02 (anatase) and of ZrOj nanoparticle thin films. The linkers were made of

/?-phenylenethynylene (Ph-E)„ bridges carrying two COOR anchoring groups at the end

and were capped with Ru(II) polypyridyl complexes as the sensitizing chromophores.

Two series of rigid-rod sensitizers were prepared: Ru complexes having bpy or 4,4'-(Cl)2-

bpy as the ancillary ligands. In the first series, the excited state was localized on the rigid-

rod linker, in the second series, the excited states were localized on the 4,4'-(Cl)2-bpy

74

ligands. The rigid-rod sensitizers with Ru(bpy)2 complexes did bind strongly (K^ ~ 105

M"1) with high surface coverages (~10"8 mol/cm2) on the nano structured metal oxide

films, Zr02 and anatase Ti02. The length of the fully conjugated rigid-rod linker

influences the photophysical properties of the sensitizer, and nanosecond transient

absorption measurements indicated long-lived metal-to-ligand charge-transfer (MLCT)

excited states (~2 |ns) with evidence for derealization onto the rigid-rod linker. The

interfacial electron transfer behavior on TiC>2 was found to be dependent on the Bransted

acidity or basicity of the surface. On base pretreated Ti02, the excited state electron

injection yields were low and could be increased by addition of LiC104 to an external

CH3CN solution. Under these conditions, a fraction of the injection process could be time

resolved on a 10 ns time scale. On acidic Ti02, ultrafast excited state electron injection

was observed for both series. Recombination was found to be second order with average

rate constants independent of which rigid-rod sensitizer was excited. For rigid-rod

sensitizers with Ru(4,4'-(Cl)2-bpy)2, there was evidence for a direct interaction between

the 4,4'-(Cl)2-bpy ligands and the Ti02 surface. Photophysical and interfacial electron

transfer properties of these Cl-substituted complexes were nearly independent of the

rigid-rod length.

5.1. Introduction.

Polypyridine complexes of Ru(II) are classical photosensitizing dyes or "sensitizers" for

nanocrystalline Ti02 (Gratzel) solar cells.1'2 The sensitizers are bound to anatase Ti02

nanoparticles through anchoring functional groups, such as carboxylic acid or

75

phosphonate groups, on the diimine ligand,la and the development of rigid molecular

linkers to the surface is attracting increasing attention.3 Rigid linkers varying in length

and structure that have the shape of tripods were developed in our laboratories to study

interfacial charge injection processes, Figure 5.1.4 Tripodal sensitizers, because of the

three-point attachment, stand perpendicularly to the surface and were useful in studies

that required a well-defined binding geometry and distance of the sensitizer (S) from the

semiconductor.5'6 There was evidence that the conjugated bridge (b) played a role in the

injection process,6 but the tetrahedral core (carbon or adamantane) was likely to act as an

insulating unit.

An alternative to tripodal linkers was to use rigid-rod linkers that provided a fully

conjugated bridge to the surface.7 Studies of rigid-rods substituted with pyrene sensitizers

and two COOH anchoring groups have been discussed in chapter 3.8 Light absorption by

pyrene resulted in quantitative, sub-nanosecond, electron injection into the T1O2

nanoparticles. Interestingly, the conjugated /?-phenylenethynylene, (Ph-E)„, bridge of the

rigid-rods considerably enhanced the pyrene extinction coefficient and red shifted the

absorption spectrum. Thus, the photon-to-electron conversion efficiency at single

wavelengths of light was a function of the bridge length and could be controlled at the

molecular level for optoelectronic applications. However, for regenerative solar cell

applications, Ru(II) polypyridine complexes are generally preferred to organic

chromophores because of their visible light harvesting efficiency, tunability, and

stability.13-2

76

rigid-rod linker -

Figure 5.1. Schematic illustration of rigid-rod and tripodal linkers bound to metal oxide (MO) nanoparticles' surface consisting of a bridge (b) of variable length, anchoring groups (A), and a sensitizing dye (S). Similar to that of the organic rigid-rod systems (Figure 4.1).

77

In this dissertation, two series of rigid-rod linkers varying in length and substituted with

Ru complexes are discussed. In the first series (la-c and 3), the ancillary ligands are bpy

for the Ru complexes; in the second series (2a-b) the ancillary ligands are 4,4'-(Cl)2-bpy,

Figure 5.2. The studies were performed on the compounds in solution as well as anchored

to nanocrystalline HO2 or Zr02 thin films. A prime motivation for these studies was to

ascertain whether tuning the sensitizer-nanoparticle electronic interactions by varying the

distance could be used to control the rate constants for charge injection and/or

recombination.9 More specifically, the goal is to find conditions where the injection

quantum yield, 4^, was still unity while the charge recombination rate was further

inhibited. Such conditions would not be expected to significantly improve efficiencies for

optimized sensitizers such as N3, c«-Ru(dcbpy)2(NCS)2. However, these conditions

would be useful for "black" sensitizers that have more negative Ru(III/II) reduction

potentials such that the rate constants for charge recombination and iodide oxidation are

competitive leading to less efficient collection of the injected electrons in the external

circuit.10 To test whether the fully conjugated rigid-rod linker made of (Ph-E)„ units

functions as a conduit for electron transport, specific compounds were designed where

the MLCT excited state was localized on ancillary 4,4'-(Cl)2-bpy ligands, rather than on

the rigid-rod. This design did indeed provide us with the opportunity to study "remote"

interfacial charge-transfer processes.

Since these studies were initiated on the tripodal4"6 and rigid-rod7'8 sensitizers, Kilsa et

al.11 have reported TiC>2 sensitization studies of rigid-rods of various lengths carrying

Ru(II) polypyridyl sensitizers and one COOH anchoring group. The lack of clear distance

78

Rutbpyfe*

.-RuCbpyfe2*

la: ROD^pyRu ' £ ^ SSSSSfa i * *QBH»~*^» **********

2*

A = COOMe

2+

2a: RODI-bpyRl^M'-CljtWk1* 2b: ROOa-bpyflmM'-Cljbpyfe* 5: bpyRiXM'-CJjbpyh**

Figure 5.2. Structure of rigid-rod complexes and two reference complexes. The rigid-rod linkers are made of (Ph-E)„ units and are terminated with two methyl esters (COOMe) as the anchoring groups (a). The number after ROD in the abbreviated name refers to the number of Ph-E units in the linker. The counterion was PF6- in all cases.

These Ru derivatives were synthesized by Elena Galoppini and co-workers at Rutgers University.

79

dependencies suggested that the rods were bound to HO2 with multiple orientations and

distances with respect to the surface.11 Although the rigid-rods reported herein are very

similar to those previously reported,11 they differ in several important respects. First, the

linkers described here have two anchoring groups instead of one, which may influence

the sensitizer-surface binding mode and orientation. Second, the bridge is made of p-

phenyleneethynylene spacers, (Ph-E)„, instead of />-phenylene units, (Ph)„. The (Ph-E)„

units, as opposed to (Ph)„, are able to assume a flat, fully conjugated conformation. The

degree to which the excited electron is dispersed into the bridge is therefore different, and

this could alter both the excited state and the interfacial electron transfer properties. In

fact, it was found that the photophysical properties of the Ru(II) rigid-rod sensitizers

described herein shared more commonalities with the oligomeric 4,4'-/?-

phenyleneethynylenes Ru(II) polypyridyl derivatives studied in considerable detail by

Schanze12 for applications in organic light emitting diodes and sensors.

5.2. Experimental Section.

Metal Oxide Thin Films

Transparent and mesoporous thin films of TiC>2 or ZrC>2 on glass slides were prepared in a

manner very similar to that previously described.13 ZrC>2, an insulator (band gap w5 eV),la

was used to study the excited state of bound molecules. The films were pretreated with

aqueous acid or with aqueous base prior to attachment of the sensitizers.14 Acid

pretreated films were prepared by immersing the films in pH = 1 H2SO4 (aq), while basic

80

films were pretreated using pH = 11 NaOH (aq). After this treatment, the films were

rinsed with CH3CN prior to immersion in an -0.1-1 mM CH3CN solution of the

sensitizer for 24 h. The sensitized films were thoroughly rinsed and then immersed in

pure solvent for several hours until desorption of weakly bound sensitizer molecules was

no longer detected (UV-vis of the solution).

Spectroscopy

UV- Vis A bsorbance

Ground state UV-vis absorbance measurements were performed on a Hewlett-Packard

8453 diode array spectrophotometer. The sensitized films were placed diagonally in a 1

cm square quartz cuvette with acetonitrile. An unsensitized film was used as the

reference. Transient absorption measurements were acquired with an apparatus that has

been previously described.13 A Xe lamp was used to probe, and 417 nm light from a D2-

filled Raman shifter pumped by a Nd:YAG Continuum Surelite II laser was used as the

excitation beam. Samples were nitrogen purged for at least 15 min or freeze-pump-

thawed prior to flash photolysis studies. Sensitized films were immersed in solutions of

acetonitrile, neat or with LiClCv Each kinetic trace was acquired averaging 60-150 laser

pulses. The quantum yields for excited state electron injection into TiC>2 were quantified

by comparative actinometry as previously described.19 A Ru(bpy)3(PF6)2 doped

poly(methyl methacrylate), PMMA, thin film, whose optical absorption and physical

dimensions were very similar to the sensitized TiC>2 films, was used as the actinometer.

The amplitudes of photoinduced absorption changes were converted to concentration

changes through Beer's law. A literature value for the difference of (-1.00 ± 0.09) x 104

81

M^cm"1 at 450 run between the extinction coefficients of the excited state and of the

ground state was used.20 Quantum yields for injection were measured most accurately by

probing at wavelengths that corresponded to isosbestic points between the ground and

excited states. The isosbestic points were determined on ZrC>2 films where only the

ground and MLCT excited states were observed. The extinction coefficients and

absorption spectra of the oxidized rigid-rods were determined by spectroelectrochemical

measurements.

Infrared Spectra.

IR spectra of the sensitizers were obtained by depositing two drops of a 1 mM CH3CN

solution of the sensitizer on a Ge or CaF2 window, allowing the solvent to evaporate. The

IR spectra of the Ti02-bound sensitizers were obtained in a transmission mode with films

cast on Ge or sapphire windows or by attenuated total reflectance (ATR) with a Golden

Gate Single Reflection Diamond ATR apparatus. In all cases, an unsensitized film acted

as the background. The spectra were collected with a Nexus 670 Thermo-Nicolet FTER or

a Mattson Research Series 1 FTIR spectrometer at 4 cm"1 resolution.

Raman Spectra.

Raman spectroscopy was carried out by using a HoloLab 5000/Raman Rxnl Confocal

Raman Spectrometer (Kaiser Optical Systems, Inc.) with an Invictus 785 nm laser diode.

A Teflon sample holder was used to measure the sensitizers (as powders), and the spectra

of the Ti02-bound sensitizers were obtained on films cast on a Sapphire window.

82

Photoluminescence.

Corrected photoluminescence (PL) spectra were obtained with a Spex Fluorolog that had

been calibrated with a standard tungsten-halogen lamp using procedures provided by the

manufacturer. Sensitized films were placed diagonally in a 1 cm square quartz cuvette,

immersed in acetonitrile, and purged with nitrogen for at least 15 min. The excitation

beam was directed 45 ° to the film surface, and the emitted light was monitored from the

front face of the surface-bound sample and from a right angle in the case of fluid

solutions. Photoluminescence quantum yield measurements, 4*PL, were performed using

the optically dilute technique with Ru(bpy)3(PF6)2 in acetonitrile as the actinometer, eq

5.1.21

<|)PL = ( A r / A s X I A X l l s / n r ) 2 ^ ( 5 . 1 )

AT and As were the absorbances of the actinometer and sample, respectively, Iv and Is were

the integrated photoluminescence of the actinometer and sample, respectively, nT and ns

were the refraction indexes for the solvents used for the actinometer and sample,

respectively, and 4>r was the quantum yield for Ru(bpy)3(PF6)2 in acetonitrile ($T =

0.062).22

Time-resolved photoluminescence decays were acquired in a similar optical arrangement

with excitation from a nitrogen-pumped dye laser that has been previously described.23

For solution studies, the traces were fit to a first order kinetic model. Values for radiative

83

and nonradiative constants, h and k^ respectively, were calculated from equations 5.2

and 5.3 with the measured quantum yields and lifetimes, x.

<J>PL = kj(b + km) (5.2)

<\>?L = kr*T (5.3)

Electrochemistry.

Cyclic voltammetry was performed in 0.1 M tetrabutylammonium perchlorate

(TBACIO4) CH3CN electrolyte in a standard three-electrode arrangement with a glassy

carbon or sensitized Ti02 working electrode, a Pt gauze counter electrode, and Ag/AgCl

or SCE as the reference electrode. A BAS model CV-50W potentiostat was used, and the

CV experiments were carried out at room temperature under argon.

Adsorption Isotherms.

Surface binding was monitored spectroscopically by measuring the change in film and

solution absorbance after soaking the film for 12 h in acetonitrile solutions with known

concentrations of the sensitizers. In all cases, the surface coverage saturated at high

sensitizer concentration. The equilibrium binding for all Ru-rod sensitizers were well

described by the Langmuir adsorption isotherm model24 from which surface binding

constants (Kad) were abstracted using equation 5.4,

84

mf}^= _J + [ R u i (5.4) r Kadr0 r0

where [Run]eq is the equilibrium sensitizer concentration, To is the saturation surface

coverage, and T is the equilibrium surface coverage at a defined molar concentration.

Plots of [Run]eq/ T versus [Run]eq were fitted linearly to obtain the binding constants KAd

and surface coverages T 0.

5.3. Results.

Binding to HO2 Films.

Adsorption isotherms for sensitizer binding to Ti02 in acetonitrile were measured at room

temperature. All equilibrium binding was well described by the Langmuir adsorption

isotherm model from which surface adduct formation constants and limiting surface

coverages were obtained.24 The surface coverages for la-c and 2a-b were about 10'8

mol/cm2 and were comparable to those obtained for tripodal sensitizers5 and for Ru

complexes directly bound through a deb (4,4'-(C02H)2-bpy) ligand.1^0 The binding

constants, approximately 2 x 105 M"1 for la-c, were consistently smaller than those

observed for the tripodal sensitizers (-10 x 105).5 The surface binding constants for

RODl-bpyRu(bpy)22+ (lb) and RODl-bpyRu(4,4'-(Cl)2-bpy)22+ (2b) on pH = 1

pretreated Ti02 films are reported in Table 5.1. The reference complex for Cl-substituted

rods 2a and 2b, Ru(4,4'-(Cl)2-bpy)2(bpy)2+ (5), had no obvious anchoring groups.

85

Table 5.1. Surface Adduct Formation Constants and Limiting Surface Coverages for Selected Ruthenium Sensitizers

sensitizer

Ru(bpy)2(deeb)"+

ROD2-bpyRu(bpy)2"+ (lb)

ROD2-bpyRu(4,4-Cl2bpy) 21+

Ru(4,4-(Cl)2-bpy)2(bpy)^(5)

(2b)

Kad, 10"5 M1

2.0±1.0

1.9± 1.0

3.7± 1.2 '

5.7± 1.3

r0 , mol/cm

5 ± 2 x 10-8

7.3 ± 0.2 x 10"*

3.9 ±0.3 x lO"8

4.8 ±0.4 x 1Q-X

86

However, the equilibrium binding constant for 5 was almost the same as that for Ru

complexes that have COOMe binding groups, such as (deeb)Ru(bpy)22+ and rigid-rods la

and lb , Figure 5.3. Since control experiments showed that [Ru(bpy)3]2+ was weakly

adsorbed, adsorption in the case of [Ru(4,4'-(Cl)2-bpy)2(bpy)]2+ must have been assisted

by the presence of the Cl-bpy ligands.

IR and Raman Spectroscopy.

The Raman spectra of la before and after binding differed only from contributions by the

characteristic anatase bands, Figure 5.4a. Both spectra exhibited bands typical of the

aromatic ring and of the C=C bond, but the carbonyl band was too broad and too weak to

give reliable information about the surface interaction(s). This was consistent with

previous Raman studies of Ru sensitizers on TiC>2.25 IR measurements were performed

for the rigid-rods before and after binding. The pH pretreatment was found to

significantly influence the binding process.14 All spectra of the nonbound esters displayed

intense bands at -1720 cm"1 (vC=0) for the carbonyl and at -1600 cm"1 (vC-^C(Ar)) for

the phenylene groups. The IR and Raman spectra of la before and after binding to acid-

pretreated pH = 1 TiC>2, shown in Figure 5.4, were representative of the IR and Raman

spectra obtained for the rigid-rods. Upon binding to TiC>2 films, the IR spectra exhibited a

small (7-12 cm"1) shift to lower wavenumbers of the carbonyl stretch, while the 1600

band due to the aromatic rings remained unchanged, Figure 5.4b.

Conversely, the IR spectra, of the rods bound to basic (pH = 11 pretreated) Ti02, shown in

Figure 5.5, displayed a broad carboxylate band at -1550-1650 cm"1, most consistent with

87

0-0 5.0x10s 1.0x10^ 1.5x10^ 2.0x10 4 2.5x10

[Run] ,M 1 Jeq

Figure 5.3. Surface adduct formation shown as plots of [Run]eq/r versus [Run]eq with overlaid linear fits for the adsorption [Ru(bpy)2bpy-(EPh)2-ROD]2+ (2, black squares with black line), [Ru(4,4'-Cl2-bpy)2bpy-(EPh)2-ROD]2+ (4, red circles with red line), and [Ru(4,4'-Cl2-bpy)2bpy] + (12, green triangles with green line) to nanocrystalline Ti02.

88

a)

12000-

3 8000-

1 c

4000-

0-

if'

it*

f

TiO, anatase r

1604

1 1 2217

1605 2218

~ l • T ' "'"1

b)

I

500 1000 1500 2000 2500

Raman Shift (cm'1)

1717 1606

1800 1750 1700 1650 1600 1550 1500

Wavenumbers (cm")

Figure 5.4. a) Raman spectra of ester la (-) as a solid and la on Ti02 ( ). The Ti02 film was cast on sapphire and was pretreated with acid, b) IR spectra of the carboxylic acid prepared from ester la (-) on a Ge and la on Ti02 ( ). The Ti02 film was cast on Ge and was pretreated with acid.

89

aoi2

aoi <H

% aooe c

s a: aooe

0.004

aooeH

aooo — i — T 1 1 1 1 1 | i | i | 1 1 1 1

1750 1700 1660 1600 1550 1500 1450 1400 1350

Wavenumber (cm"1)

Figure 5.5. Attenuated total reflectance IR spectra for 2b on T1O2 thin films that were pretreated at pH = 1 (—) and p H = l l ( - ) .

90

bidentate coordination modes. Interestingly, the rigid-rod sensitizers bound strongly to

Ti02 samples that were not pretreated with aqueous acid or basic solutions, and yielded

IR spectra with signatures for both the carbonyl and the carboxylate band indicative of a

mixture of binding modes.

The IR spectra of the rigid-rods obtained on Ti02 thin films, cast on germanium or

sapphire windows, were substrate independent. Germanium was preferred because it

combined resistance to aqueous acidic treatments with transparency in the IR region of

interest. The presence of a single carbonyl band after binding to an acidic TiC>2 surface

suggested that both groups were anchored in a monodentate ester-type bond..

Electrochemistry.

All rigid-rod sensitizers displayed quasireversible RuIIWI electrochemistry in CH3CN

electrolyte solution, Table 5.2. Complexes la, lb, and 3 showed Rum/n reduction

potentials at -1.3 V versus SCE that were, within experimental error, the same as those

reported previously for tripodal sensitizers with (Ph-E)„ bridges.5 These waves were

shifted to slightly more positive potentials compared to Ru(bpy)32+. A similar effect was

observed in 2a-b and in 5, as the presence of Cl-substituted ligands resulted in a

measurable positive shift in the RuIII/n potential to -1.40 V. Ligand-based reduction

potentials for the sensitizers in CH3CN solution are provided in Table 5.2.

The first reduction, E° (Ru2+/+), of la-c and 3 occurred at potentials more positive than

the corresponding reference complexes, that is, [Ru(bpy)3]2+ or [Ru(bpy)2(phen)]2+,

respectively. By comparison with the tripods5 and with other literature reports on Ru

91

Tab

le 5

.2. E

lect

roch

emic

al D

ata

for

1-3

and

Ref

eren

ce C

ompo

unds

in S

olut

ion"

Sens

itiz

er/

Com

plex

RO

Dl-

bpyR

u(bp

y)22+

(la

)

RO

D2-

bpyR

u(bp

y)22+

(lb

)

RO

D3-

bpyR

u(bp

y)22+

(lc

)

RO

Dl-

bpyR

u(4,

4-C

l 2bp

y)22+

(2a

)

RO

D2-

bpyR

u(4,

4-C

l 2bp

y)22+

(2b

)

Ru(

4,4-

(Cl)

2-bp

y)32+

(5)

RO

D2-

phen

Ru(

bpy)

22+ (

3)

Ru(

bpy)

32+b(4

)

[Ru(

bpy)

2(ph

en)]

2+b

E1

/2(R

u1M

1),

m

V

1300

1300

1287

1400

1390

1430

1300

1260

1230

£ 1/2

(Ruz+

/+),

m

V

-121

0

-123

0

-122

3

-116

0

-115

0

-108

0

-122

0

-134

0

-137

0

E1/

2(R

u+/0

),

mV

-148

0

-148

0

-142

9

-129

0

-128

0

-119

0

-148

0

-152

0

-153

0

mV

-820

-830

-850

-750

-760

-730

-930

-860

-910

are

repo

rted

vs

SCE

. All

mea

sure

men

ts w

ere

perf

orm

ed i

n 0.

1 M

TB

AC

IO4/

CH

3CN

. Fr

om G

alop

pini

and

co-

wor

kers

.5

complexes substituted with electron-withdrawing groups,12'26 this wave was assigned to

reduction of the bpy attached to the rigid-rod linker. The presence of 4,4'-(Cl)2-bpy

ligands in 2a-b and in 5 resulted first in reduction potentials that were -50 mV more

positive, consistent with the reduction of the Cl-substituted bpy. The excited state

reduction potentials, Ei/2(Rum/l1*), were calculated from the ground state potentials and

from the free energy stored in the thermally equilibrated MLCT excited state, AGes, using

equation 5.5:

£1/2(Runi/I1*) = £i/2(Ruin/I1) - AGeS (5.5)

AGes was estimated by drawing a tangent line to the high-energy side of the corrected

photoluminescence spectra. The excited state reduction potentials calculated for

unsubstituted bpy-based rods were nearly identical to those obtained for related bpy-

based tripods.5 Similarly, the potentials for phen-substituted ligand 3 were nearly

identical to those obtained for the phen-based tripods.5 Rigid-rods containing 4,4'-(Cl)2-

bpy were weaker excited state reductants by -70 mV than those with unsubstituted bpy.

Photophysical Studies.

Selected photophysical properties for 1-5 in argon-saturated acetonitrile at room

temperature are listed in Table 5.3 together with data for the reference complexes and

two tripodal sensitizers.

93

Tab

le 5

.3.

Phot

ophy

sica

l Pr

oper

ties

of

1-5

and

Oth

er R

u C

ompl

exes

in C

H3C

N S

olut

ions

0

Sen

sitiz

er/

Com

plex

RO

Dl-

bpyR

u(bp

y)2"

+ (

la)

RO

D2-

bpyR

u(bp

y)2z+

(lb

)

RO

D3-

bpyR

u(bp

y)22+

(lc

)

RO

Dl-

bpyR

u(4,

4-(C

l)2-

bpy)

2"+

RO

D2-

bpyR

u(4,

4-(C

l)2-

bpy)

2'!+

RO

D2-

phen

Ru(

bpy)

22+ (

3)

[Ru(

bpy)

3;r

Ru(

bpy)

2(ph

en)z+

[Ru(

bpy)

2(de

eb)]

2+(4

)

[Ru(

4,4-

Cl 2

-bpy

) 3]2+

[Ru(

4,4-

Cl 2

-bpy

) 2(b

py)]

^ (5

)

(2a)

(2b)

[Ru(

bpy)

2(bp

y-(E

-Ph)

-AdT

ripo

d )]

"*

[Ru(

bpy)

2(ph

en-(

E-P

h)A

dTri

pod

)fn

A*b

s> n

m

(e,

M *

cm

1)*

46

2 (1

.6 x

104)

465

(2.0

x 1

04)

465

(2.0

x 1

04)

465

(1.6

x 1

04)

466

(1.9

x 1

04)

450

(1.8

x 1

04)

450

450

475

(1.6

x 1

04)

466

461

(1.3

x 1

04 )

461

(1.9

x 1

04)

452

(1.6

x 1

04)

A.p

L,

nm

c

640

640

639

645

645

606

610

620

690

645

646

624

z, /

*sd

1.9

2.3

2.5

0.69

0.75

1.3

0.80

1.2

0.93

0.70

2.0

1.4

eVf

2.12

2.13

2.14

2.15

2.15

2.23

2.24

2.14

2.01

2.16

2.14

2.23

102

11

13

9.5

7.0

6.9

8.5

6.2

—

4.4

6.1 10

8.0

10

4,

n1

5.1

5.4

4.7 10

9.2

6.5

7.8

—

4.7

8.7

5.1

5.7

10

5 i"1

5.4

4.0

4.3 13

12

7.0 12

—

10

13

4.5

6.6

" A

ll m

easu

rem

ents

wer

e pe

rfor

med

at r

oom

tem

pera

ture

.* A

bsor

ptio

n m

axim

a, ±

2 nm

.c Cor

rect

ed p

hoto

lum

ines

cenc

e m

axim

a, ±

5 n

m/ E

xcite

d st

ate

lifet

ime

in

CH

3CN

, ±5%

. The

sol

utio

ns w

ere

deae

rate

d by

free

ze-p

ump-

thaw

or

by s

parg

ing

with

nitr

ogen

" H

alf-

wav

e po

tent

ials

(±2

0 m

V)

wer

e m

easu

red

at a

gla

ssy

carb

on w

orki

ng e

lect

rode

in

0.1

M T

BA

CIO

4/C

H3C

N s

olut

ion

usin

g A

g/A

gCl

as th

e re

fere

nce.

Dat

a ar

e re

porte

d vs

SC

E/ G

ibbs

free

ene

rgy

stor

ed i

n th

e th

erm

ally

equ

ilibr

ated

exc

ited

stat

e.8 D

ata

from

Gal

oppi

ni e

t al

.5b

The UV-visible absorption spectra for la-c, and for 2a, 2b, and 5, shown in Figure 5.6,

parts a and b, respectively, displayed a broad band in the visible region (400-500 nm) that

is typical of metal-to-ligand charge-transfer (MLCT) transitions.

These heteroleptic complexes show broad visible absorption bands and the individual Ru

-^ bpy, Ru ->4,4'-(Cl)2-bpy, or Ru -> rigid-rod linker charge transfer bands could not be

spectrally resolved. The narrow band at higher energy at -290 nm was assigned to the n,n

* transition of the ancillary ligands and the -350 nm band was assigned to the u,u*

transition of the rigid-rod (Ph-E)„ linker. As the number of (Ph-E) units increased, the

350 nm band increased in intensity and shifted to longer wavelengths of light. As

expected, this band is absent in the spectrum of 5. The spectral changes were most

significant when the number n of (Ph-E)„ units was increased from 1 to 2. As expected,

the %,TL* transition at -290 nm due to the ancillary bpy was insensitive to the number of

(Ph-E) units, Figure 5.6, parts a and b. Similar spectral features have been observed for

the tripodal complexes43,5 and oligomeric complexes of Ru.12'26 A comparison between

the absorption maxima for la-c and the reference Ru(bpy)32+ showed an -10 nm red shift

of the MLCT band in the presence of the linker, while the absorption maxima for 2a-b

and the reference complex Ru(4,4'-(Cl)2-bpy)32+ were identical.27 The absorption spectra

of la-c and 3 anchored to Ti02 and Zr02 were largely preserved, with only some minor

broadening. The UV spectrum of the reference complex 5b, however did show an

additional shoulder upon binding, possibly due to the Cl-Ti02 interactions.

All the rigid-rod sensitizers displayed room-temperature photoluminescence (PL) in

acetonitrile and when anchored to Ti02 or Zr02. The coincident PL maxima for 2a and

95

80000

60000

£ 40000 o

5 CO

20000

700

Wavelength (nm)

300 350 400 450 500 550 600

Wavelength (nm)

Figure 5.6. Ground state absorbance spectra in acetonitrile a) Spectra of la (-), lb (•••), and lc (- - -). b) Spectra of 2a (•••), 2b (- - -), and [Ru(4,4'-(Cl)2-bpy)2(bpy)]2+ 5 (-).

96

2b were centered at 645 nm and were identical to that of the [Ru(4,4'-(Cl)2-bpy)3] . The

PL decays were well described by a first-order kinetic model in fluid solution, and the

excited state lifetimes (T), along with the PL quantum yields, are listed in Table 5.3. As

the number of (Ph-E) increased, for instance from la to lc, the -[increased slightly from

1.9 us (n = 1) to 2.3 us (n = 2) and 2.5 us (n = 3). The excited state lifetime for the

nonsubstituted complex, Ru(bpy)32+ is 0.8 us. Time-resolved PL decays on TiC>2 and on

Z r d were nonexponential.19c

The transient absorption spectra of the rigid-rods in acetonitrile solutions were assigned

to the MLCT excited state. The kinetics were first order, and the rate constants agreed

well with the time-resolved PL data at all wavelengths monitored. A comparison of the

excited state absorption spectra of la-c uncovered distinct differences due the number of

(Ph-E) units; lb exhibits a stronger, red shifted excited state absorbance beyond 500 nm

compared to that of la, Figure 5.7. The spectra show a bleach of the MLCT absorption,

and for lb and lc, a bleach of the bridge u,u* transition(s).

The excited state absorption spectra for the Cl-substituted 2a and 2b did not display an

intense absorption band in the red spectral region and agreed well with the reference

complex that lacked the rigid-rod linker, [Ru(4,4'-(Cl)2-bpy)3]2+, Figure 5.8. The excited

state absorption spectra obtained on ZrC>2 thin films were within experimental error, the

same as that in CH3CN. Transient absorption spectra on rigid-rod sensitized Ti02

contained contributions from the MLCT excited state and from a Runi/Ti02(e") charge

separated state as described below.

97

a) M«OOC

M«OOC ^ r H

350 400 450 500 550 600 650 700

Wavelength (ran)

b)

c)

-0.04

UtoOOC

350 400 450 500 550 600 650 700

Wavelength (nm)

350 400 450 500 550 600 650 700

Wavelength (nm)

Figure 5.7. Transient absorbance spectra (kex = 417 nm) in CH3CN of a) la. The data were recorded at 10 ns (•), 0.5 u,s (•), 1.0 JLLS (A), 2.0 (is (•), and 10 pis ( • ) delays after the laser pulse, b) lb. The data were recorded at 10 ns (•), 1.0 us (•), 2.5 [is (A), 5 |is (•), and 20 (is ( • ) delays after the laser pulse, c) lc. The data were recorded at 0.1 us (•), 0.5 (is (•), 1.0 (is (A), 2.0 is (•), and 4.0 (is ( • ) delays after the laser pulse.

98

o

o x> <

x:

0.04

0,02

0.00

-0.02 H

I • i — • — i — • i • — r

300 350 400 450 500 550 600 650 700

Wavelength (ran)

Figure 5.8. Transient absorbance spectra (Kx = 417 nm) of Cl-substituted complexes in acetonitrile at room temperature Ru(4,4'-(C1)2-bpy)3(PF6)2 (•), 2a (•), and 2b (A). The data were recorded 10 ns after the laser pulse.

99

Inter facial Electron Transfer.

Nanosecond transient absorption was used to quantify interfacial electron transfer

dynamics and yields. These processes were influenced by the pH. This is expected, since

the conduction band edge shifts 59 mV/pH unit.28'la On acidic TiC>2, injection occurred

within our instrument response (probably on a femto to picosecond time scale6), and the

transient spectra were mainly from the Rum/Ti02(e") charge separated states, while on

base pretreated TiC>2 the MLCT excited state was predominately observed. To eliminate

possible contributions from the excited state, the measurements were made at

wavelengths corresponding to the ground-excited state isosbestic point. In this way, the

formation and loss of the interfacial injection and recombination processes could be

cleanly observed. On pH = 1 pretreated Ti02 samples, excited state electron injection into

Ti02 occurred faster than could be time resolved with a nanosecond laser for all samples.

On basic Ti02, the injection yield was very low and could be increased by the addition of

LiC104 to the acetonitrile solution in which the film was immersed. Figure 5.9 shows

absorption changes monitored at the ground-excited state isosbestic point for base

pretreated Ia/Ti02. The rise time observed corresponds to relatively slow electron

injection, k^ = 1.0 x 107 s"1. Typically, 10-30% of the injection process could be

observed over the first 100 ns for la-c/Ti02. Injection dynamics were not observed for

the rigid-rods anchored to acidic pretreated surfaces, for the reference [Ru(bpy)2(deeb)]2+,

or for both Cl-substituted rigid-rod compounds (2a and 2b). The injection is now being

studied by femtosecond laser spectroscopy.

100

0.000

o X> <

u 00

c

U

-0.005

-0.010- V ^ i W * ^ 0.0 5.0xl0"7 l.OxlO* 1.5x10*

Time (s)

Figure 5.9. Single wavelength kinetics monitored at the ground state-excited state isosbestic point for la on pH = 1 pretreated Ti02 (black line, top) and pH = 11 pretreated Ti02, 0.1 M LiC104 (red line, bottom).

101

Comparative actinometry was used to measure the quantum yields (^j) for excited state

electron transfer to the empty states of nanocrystalline Ti02. The injection yield were

found to decrease with irradiance, and extrapolation of the data to zero irradiance gave

the mj reported in Table 5.4.19 Injection quantum yields were measured for la-c on pH =

1 pretreated Ti02 and on pH = 11/Li+ pretreated TiC>2 with comparable surface coverages,

r(Run). In general, 4^ was above 0.60 for Ru-rigid-rod complexes that were bound to pH

= 1 pretreated films, while 4>i„j for pH = 11 pretreated films were lower and decreased

with rigid-rod length, Table 5.4.

The kinetics for charge recombination between the electron in TiC>2 and the oxidized

sensitizer were quantified on pH = 1 pretreated films at similar surface coverages (-2.3 x

10"8 mol/cm2). Transient data were obtained at the ground state-excited state isosbestic

point to avoid possible contributions from the excited states. The transient data was well

described by a bi-second-order kinetic model, equation 5.6:19c

AA = AAo - AAs + AAs (5.6)

1 + (kf/Ael) t(AAo - AAs) 1 + (kJAsl) t(AAs)

where AA was the absorbance change at time t, As was the molar extinction coefficient, /

was the optical path length, AAo was the initial amplitude (equal to the sum of the

contributions from the fast and slow components), kf was the recovery rate constant for

the fast component, AAS was the amplitude of the slow component, and ks was the

recovery rate constant of the slow component. The average rate constants for charge

102

Table 5.4. Excited State Electron Yields, ^ j , " for Rigid Rods Bound to Ti02 Films Pretreated with Acid or Base

Sensitizer

RODl-bpyRu(bpy)^+ (la)

ROD2-bpyRu(bpy)2"+ (lb)

ROD3-bpyRu(bpy)22+ (lc)

pH=l*

1.0

0.89

0.86

pH = lr*

0.77

0.20

<0.05c

" All injection yields were measured spectroscopically at room temperature, Kxc = 532.5 nm. b The Ti02 thin film pretreatment prior to binding of the rigid-rod sensitizers. pH = 1 pretreated samples were studied in neat acetonitrile; pH = 11 pretreated films were studied in 0.1 M LiC104 in acetonitrile. c Li+ addition caused some desorption, so this value was measured in neat acetonitrile.

103

recombination were found to be independent of the length of rigid-rod sensitizer used and

were within experimental error the same as that for the model complex Ru(bpy)2(dcb)2+.

The weights of the two components, -70% and -30% for the fast and slow components,

respectively, were also independent of the length of the rigid-rod sensitizer, with k0bs =

2.5 ±0.3 x io7s"1.

5.4. Discussion.

The photophysical and interfacial electron transfer properties of Ru(II) rigid-rod

sensitizers were quantified at room temperature in acetonitrile solution. The interfacial

pH was found to be important for both sensitizer binding and for photoinduced electron

transfer. Efficient electron injection was observed on acidic TiC>2, while long-lived

excited states were observed on basic TiC>2. Back electron transfer rate constants were

found to be independent of the length of the rigid-rod excited, consistent with a

previously proposed mechanism wherein transport of the injected electron to the oxidized

i n

sensitizer was rate limiting. These studies provide new details on surface binding,

molecular excited states, and interfacial electron injection. Below interpretations and

implications of these new findings are discussed.

Surface Binding.

Adsorption isotherms and IR studies of the rigid-rod sensitizers 1-3 have shown that all

bind strongly to nanocrystalline Ti02 films. In most studies, the methyl esters of the Ru

rigid-rods were reacted with the TiC>2 surface in acetonitrile at room temperature. The

104

observation of strong binding was interesting, considering that identical rigid-rod linkers

capped with an organic chromophore (pyrene) bound only when it was first converted

Q

into a carboxylate salt or a carboxylic acid. Tnpodal sensitizers behaved similarly:

tripods capped with organic chromophores such as pyrene or perylene did not bind

strongly under conditions where Ru tripods did.31'8 The fact that noncharged, aromatic

sensitizers required different binding conditions than the charged Ru complexes may

have been due to a variety of factors, such as differences in the solubility in organic

solvents and/or electrostatic interactions with the HO2 surface.

Infrared measurements showed that both ester groups reacted with HO2 to yield a single

binding mode whose chemical nature was dependent on the Brtfnsted acidity of the

surface. On acidic HO2 surfaces, a single asymmetric CO stretch was observed at higher

energy than the methyl ester, which we assigned to ester linkages. On basic Ti02, a broad

band centered at -1550-1575 cm"1 was observed at lower energy and was assigned to a

carboxylate binding mode. The fact that only a single binding mode was present on acidic

and on basic Ti02 indicated that both anchoring groups were in a similar environment.

Interestingly, on surfaces that were not pretreated with acid or base, spectroscopic

signatures of both binding modes were present. We should emphasize, however, that the

IR and surface binding studies do not demonstrate that the rigid-rods are perpendicular to

the surface or that they assumed a single orientation.

Ruthenium model compounds that contained the 4,4'-(Cl)2-bpy ligand(s) without obvious

anchoring groups were found to bind surprisingly well to Ti02. The rigid-rods that

contain 4,4'-(Cl)2-bpy ligands, that is, 2a and 2b, may bind with the Ru(4,4'-(Cl)2-bpy)2

105

"head-down" and the anchoring groups of the linker, all interacting with the HO2 surface.

This geometry clearly raises issues in the study of remote MLCT states, as discussed

later. In addition, the appearance of this unexpected kind of CLu-TiCVsurface binding

stresses the need to carefully evaluate the role of "nonbinding" groups introduced on

sensitizers to tune excited state and energetic properties.

3MLCT Excited State in Solution and Surface Bound.

The Ru rigid-rods have metal-to-ligand charge-transfer (MLCT) excited states.27 For the

parent compound Ru(bpy)32+, the preponderance of data supports a "localized" formalism

for the thermally equilibrated excited state in fluid solution, that is, Rum(bpy")(bpy)22+*

not Ruin(bpy"1/3)2+*.2'32 Furthermore, for compounds that contain different diimine

ligands, the excited state is expected to localize on the ligand that is most easily

reduced.27 It was therefore of interest to consider which ligand(s) the emissive excited

state was localized upon. A second and not completely unrelated issue concerns the

extent of charge transfer into the rigid-rod linker.

The (Ph-E)„ bridge in the rigid-rod linker and the 4,4'-(Cl)2-bpy are known to be electron

withdrawing and were therefore expected to stabilize the MLCT excited states.12'26 This

expectation was realized and it is clear that the rc* orbitals of the ligands decrease in

energy: bpy > rigid-rod bpy ~ rigid-rod phen > 4,4'-(Cl)2-bpy. Therefore, the emissive

excited state of Ru rigid-rod compounds that contain the 4,4'-(Cl)2-bpy ligand was

localized on this ligand, and this explains why their photophysical properties were nearly

independent of the Ph-E bridge. For the unsubstituted bpy based Ru rigid-rods, the

excited state was localized on the bpy (or phen) attached to the Ph-E bridge. The

106

spectroscopic data reveal the same localized excited states for the rigid-rods in fluid

acetonitrile solution and when anchored to ZrC>2 or Ti02 nanoparticles. Therefore, for the

Ru(bpy)2 rigid-rods, the thermally equilibrated excited state was localized on rigid-rod

linkers bound to the semiconductor surface.

It was of interest to compare Ru(bpy)2 rigid-rods where the excited state was localized on

a bpy with a variable length phenyl ethyne bridge, la-c. The visible absorption and

photoluminescence spectra of these rigid-rods were within experimental error the same.

Interestingly, the excited state lifetime increased for the two longer rigid-rods relative to

the shortest one. For a series of closely related Ru compounds, an increase in lifetime is

generally accompanied by a blue shift in the emission spectra in accordance with the

energy gap law.27 The increased lifetime observed here must have a different origin and

may reflect the phenylethyne substituents whose extended n-conjugation allows for

greater electron derealization. Excited state derealization disperses the electron into the

(Ph-E) bridge, thereby decreasing the average bond displacement energy, which

decreases vibrational overlap and hence the nonradiative rate constant.29

An alternative explanation for the long-lived "red" emissive excited state is that mixing

between MLCT and intraligand excited state(s) of the rigid-rod imparts more triplet

character and hence, a longer lived excited state. A mechanism where the MLCT excited

state transfers energy to a lower-lying triplet state of the rigid rod is ruled out by the

wavelength independent lifetimes measured by transient absorption spectroscopy and the

MLCT-like steady state PL spectrum. Nevertheless, excited state derealization and/or

107

mixing between the MLCT manifold and intraligand triplet states likely explain the long-

lived excited state behavior that does not follow the energy gap law.

Indirect evidence for derealization into the phenylethyne bridge is seen by the strong

excited state absorption in the >600 nm region. The first extension of the bridge (from la

to lb) resulted in significant changes in the intensity and spectrum, while further

elongation resulted in smaller differences. Previous researchers have also observed

intense near-IR absorption bands for phenylethyne substituted bpy ligands coordinated to

Ru(II) and have also attributed this to derealization of the excited state.12

Excited State Electron Injection into T1O2.

The quantum yield for excited state injection from the Ru rigid-rods to TiC>2 (4^), listed

in Table 5.4, was measured on acid (pH = 1) and base (pH =11) pretreated TiC>2 films on

the nanosecond time scale and therefore may not be the true injection yields as any sub-

nanosecond (geminate) recombination of the injected electron with the oxidized dye were

unaccounted for. Therefore, the ^ values reported here are best considered as lower

limits of the true injection yields. The 4^ on acid (pH = 1) pretreated Ti02 were near

unity, consistent with negligible geminate recombination. The 4^ values for la-c

decreased by about 15% as the number of (Ph-E) units in the bridge increased. This effect

may be due to the increasing distance of the complex from the surface.

At present, however, is not possible to determine whether this is a "distance dependent"

effect, since the orientation of the rods with respect to the surface is unknown. Even if

both COOR anchoring groups bind to the surface, the axis of the rod may not be

108

perpendicular to the surface and the molecules may bind at an angle (or angles) with

respect to the surface. In conclusion, the effective injection distance is unknown and it is

probably less than the Ru-center-to-0-bound-to-Ti02 distance calculated assuming that

the molecules are perpendicular (J(la) ~ 12 A, d(lb) ~ 19 A, d(lc) ~ 25 A).

On base (pH =11) pretreated Ti02, the injection yields were very low, 4^ < 0.05, Table

5.4. This is expected, as the conduction band edge shifts 59 mV/pH unit, resulting in poor

orbital overlap with the excited state sensitizer.28 It is known, however, that the addition

of "potential determining ions", such as Li+, can shift the conduction band edge and

promote electron injection under these conditions.33

A curious detail of the injection process is that the quantum yield decreased with

increased incident irradiance. This behavior was first reported by Kay and Gratzel and

was attributed to trap filling that shifts the quasi-Fermi level of TiC>2 toward the vacuum

level, making electron injection less favorable.34 If this explanation were correct, the

photoluminescence quantum yield should increase as 4^ decreases. However, here, and

in previous studies, we find that both quantum yields decrease with irradiance, indicating

that some other fast process is lowering the injection yield.190

The electron injection rate constants on acid pretreated TiCh films could not be time

resolved with the nanosecond laser used in this study (kmj »108 s"1). This result was

expected, since the rates of photoinduced interfacial electron transfer for Ru(II) sensitized

TiC>2 are known to occur on the femto to picosecond time scale.35 Recent ultrafast

measurements of tripodal sensitizers 15-24 A long bound to Ti02 revealed that the

injection occurs on a femto to picosecond time scale.6'35 It is noted that injection rate

109

constants Ainj >108 s'1 with (kj + k^) ~ 5 x 105 s"1 for the rods imply that the injection

quantum yields should be near unity, consistent with the actinometry measurements.

A fraction of the injection process for the rigid-rods with bpy as the ancillary ligand

could be time resolved on base pretreated Ti02 films immersed in 0.1 M LiC104

acetonitrile solution. As mentioned above, at basic pH the electron injection is less

favorable. Under these conditions, approximately 70-90% of the signal was instrument

response limited, while the remaining fraction could be quantified (k-^ ~ 1 x 107 s"1). This

is in agreement with the quantum yields measurements and strongly suggests that the

density of acceptor states for basic HO2 is significantly lower than for acidic TiCh.

The injection rate could not be time resolved, k^ > 108 s"1, for the rigid-rods containing

the 4,4'-(Cl)2-bpy ancillary ligand bound to acidic as well as basic TiC>2, despite the fact

that they are weaker excited state photoreductants than rigid-rods la-c. This finding

supports the notion that the 4,4'-(Cl)2-bpy ligands interact directly with the TiC>2 surface,

and that the molecules may bind "head down", thereby providing a new pathway for

electron injection.

In summary, a small fraction of the injection process could be observed on the

nanosecond time scale when the TiC>2 was base pretreated and when the excited state was

localized on a rigid-rod linker. All other experimental conditions led to instrument

response limited electron injection rate constants with nearly quantitative yields. The

faster and more efficient electron injection into acidic TiC>2 was attributed to a higher

density of unfilled states that overlap with the excited state sensitizer orbitals.

110

Femtosecond laser spectroscopy studies aimed to quantify the injection rates for la-c to

determine whether there are differences due to the linker's length are in progress.

Back Electron Transfer.

A specific goal of this study was to find conditions where the injection yield was unity

while the back electron transfer rate constant decreased with distance. The recombination

between the electrons in HO2 and the oxidized dye can lower the efficiencies of solar

cells. Hence, an attractive way to minimize this process would be to increase the distance

between the HO2 surface and the metal center, thereby decreasing the rate of

recombination. This effect was not realized. Back electron transfer rate constants were

found to be independent of the length of the rigid-rod excited. A similar observation was

made for the tripodal sensitizers, where the recombination rates between the electrons in

TiC>2 and the oxidized dye were found to be independent of the linker's length.5'6

Recombination of the injected electron with the oxidized dye required milliseconds for

completion, and the kinetics were well described by a sum of two second order rate

constants. Average rate constants for the rigid-rods la-c were within experimental error

the same. This is not unexpected in view of literature reports proposing a mechanism

wherein transport of the injected electron to the oxidized sensitizer is the rate limiting

step.30 The main process responsible for controlling the recombination rates is usually

assumed to be the electron transport by diffusion between trap sites on a semiconductor

nanoparticle surface (the Random Flight Model30). It is generally concluded that the

recombination reaction is governed by the energy redistribution of the electrons trapped

in the semiconductor, rather than by spatial diffusion. However, Clifford et al.36 have

111

recently observed slower recombination dynamics with single exponentials in dye cations

that are physically separated from the surface, an effect that is being studied by Tachiya

et al.37 The recent interest in the processes that regulate the recombination rates shows the

importance of having available model linkers such as the ones studied here. Based on

these data,36'37 It is anticipated that a distance dependence with TiC»2 materials that have a

lower density of trap states and at higher excitation irradiances will be observed.

5.5. Conclusions.

New rigid-rods Ru(II) polypyridyl complexes were synthesized by Galoppini and co­

workers and anchored to nanocrystalline T1O2 films for dye sensitization studies. In the

rods having unsubstituted bpy as ancillary ligands, the (Ph-E)„ bridge acts as a u-acceptor

group, resulting in derealization of the MLCT state on the linkers, as indicated by

measurable spectral shifts in the absorption, PL, and transient absorption spectra. When

the ancillary ligands contain CI groups, there is clear evidence that the MLCT is localized

on the 4,4'-Cl2-bpy. An unexpectedly strong binding between the 4,4'-Cl2-bpy and the

TiC>2 was observed, so that while the rigid-rods bind exclusively through the carboxylic

group, the rigid-rods with Cl-substituted ligands may also bind with the Ru complex

"head down". Ester-like or carboxylate binding modes to the TiC>2 surface were observed,

and conditions to obtain only one type of binding mode were found. The rigid-rods were

expected to exhibit diminished electronic coupling with the TiC>2 acceptor states relative

to complexes that are directly attached. This behavior was realized under conditions

112

where the density of Ti02 acceptor states was intentionally decreased by surface

pretreatments with basic aqueous solutions. Future studies will focus on ultrafast

spectroscopic measurements aiming at quantifying the electron injection process and on

the application of these new Ru rigid-rod sensitizers in regenerative solar cells.

113

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Hagfeldt, A.; Gratzel, M. Chem. Rev. 1995, 95, 49. (c) Kamat, P. V. Chem. Rev.

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Piotrowiak, P.; Galoppini, E.; Wei, Q.; Meyer, G. J.; Wiewior, P. J. Am. Chem.

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119

Chapter 6. Measurement of Oxygen Tension in Single

Cardiomyocytes: Photoluminescence Quenching of Intracellular

Ru(bpy)32+ by Molecular Oxygen

6.1. Introduction.

The balance of energy supply and demand is a crucial determinant of cardiovascular

health, but the mechanisms that regulate oxidative phosphorylation are still poorly

understood. While oxygen consumption (V02) can be measured in cell suspensions or

tissues, measurements of respiratory flux in single cells have proven challenging. Here,

we report a novel method for monitoring intracellular oxygen tension using the ruthenium

2+

polypyridyl complex, Ru(bpy)3 (tris(2,2'-bipyridine)ruthenium(II) dichloride), which

has a high photoluminescence quantum yield, a long lifetime, and is efficiently quenched 2+

by molecular oxygen. Ru(bpy)3 was introduced into adult guinea pig cardiac cells via 2+

the whole cell patch clamp method, and Ru(bpy)3 photoluminescence, mitochondrial

redox potential, and sarcolemmal K,ATP currents were monitored simultaneously as

indices of the cellular metabolic status. Significant dequenching of the Ru(bpy)3

emission was observed upon exposure of the myocytes to the mitochondrial uncoupler

FCCP, indicative of a rapid increase in V02 in parallel with the oxidation of

120

mitochondrial flavoproteins. These events were followed shortly thereafter by the

activation of K, ATP current (/KATP) and were reversed by inhibiting electron transfer to

oxygen with sodium cyanide. The Ru(bpy)32+ emission also responded reproducibly to

changes in perfusate p02, permitting quantitative calibration of the intracellular oxygen

concentration. Application of this method to models of cardiovascular disease will

provide novel insights into the role of mitochondrial dysfunction under pathological

conditions.

Most aspects of cellular function depend on a continuous supply of energy provided by

mitochondrial oxidative phosphorylation. The balance between oxygen supply and

demand is especially important in muscle, where the workload is continually varying and

spans a wide range. In addition to the basic need to supply ATP for ion transport and

contraction, mitochondrial respiration is also an important determinant of reactive oxygen

species (ROS) production, which may serve as either a molecular signaling pathway or a

toxic agent. Pathological models such as ischemia-reperfusion or hypoxia-reoxygenation

also bring about metabolic changes defined by the severity of oxygen depletion or

abundance.

In order to better understand how oxidative phosphorylation is regulated in the intact cell,

it would be very useful to have an index of metabolic flux by measuring intracellular O2

consumption while simultaneously measuring steady state changes in mitochondrial inner

membrane potential (A^m) and in redox state. Several attempts to measure O2 tension in

single cells have been reported previously, including measuring changes in extracellular

121

oxygen tension with a stable or oscillating electrode (fiber optic1 or polarigraphic2, by

phosphorescence quenching, or by examining the O2 - dependent quenching of

myoglobin fluorescence3. The latter method has an intrinsically low signal-to-noise ratio

and may be influenced by changes in the cellular environment (e.g., changes in pH or ion

concentrations). With extracellular methods, the pC>2 surrounding the cell is assumed to

be in equilibrium with the oxygen tension within the myocyte and may change depending

on the stirred and unstirred layers near the membrane and the geometry of the cell. These

methods are also subject to signal variation and, in some cases, consumption of O2 by the

electrode itself. An intracellular electrode technique, which pierces the cell to record

internal oxygen tension, has been previously reported.1 Thus, an easily recorded and/or a

less invasive approach are needed to expand upon and better understand the role of

molecular oxygen in cellular events.

In this regard, molecular sensors have proven to be an efficient means of measuring

intracellular concentration of ions, location and concentration of proteins, the intracellular

pH, mitochondrial membrane potentials, carbon dioxide and other gases, and

concentrations of reactive oxygen species and other radicals. Most available sensors

utilize the fluorescence of organic fluorophores for detection. However, for longer-lived

processes that require diffusion of the measured species, such as the measurement of

molecular oxygen, an inorganic sensor with a long-lived excited state is favored.

Inorganic dyes such as ruthenium polypyridyl complexes are widely used throughout the

scientific community in such areas as solar energy conversion4, photoluminescent

122

sensors5, chemiluminescence labels6, measuring DNA binding constants, and bioanalysis

of molecules8 due to their advantageous photophysical properties9'10. Fluctuations in

oxygen concentrations have also been probed by monitoring the steady state

photoluminescence of ruthenium polypyridyl complexes at varying temperatures11. For

quenching-based oxygen sensors, the most successful compounds have proven to be

those with high quantum yields and long lifetimes . Unlike organic probes, inorganic

dyes with transition metal centers also have red-shifted absorbance and emission spectra,

large Stokes shifts,13 broad absorbance and emission bands, and high photo and chemical

stability in both the ground and excited states14. Such intrinsic properties allow for more

efficient reactions to occur within the excited state lifetime of the ruthenium polypyridyl

dyes.

Here, we use Ru(bpy)32+, tris (2,2'-bipyridine) ruthenium(II) dichloride, a well

characterized divalent ruthenium polypyridyl dye, to measure the intracellular pC>2 of

guinea pig ventricular cardiomyocytes. Ru(bpy)32+ consists of a transition metal center

with three bipyridine chelating ligands, Figure 6.1. The most essential feature for

measuring pC>2 is the quenching efficiency of the Ru(bpy)32+ excited state by molecular

oxygen. The existence of a triplet excited state and its strong mixing with a low-lying

singlet excited state are responsible for the long, sub-microsecond lifetime.5 This long

lifetime allows for the oxygen to diffuse within the cardiomyocyte to the ruthenium

sensor, quenching its photoluminescence. If the lifetime were short-lived, the oxygen

may not reach a Ru(bpy)32+ excited state molecule during it's lifetime, and therefore,

quenching would not occur. Quenching is defined as a process which will reduce the

123

Figure 6.1. Chemical structure of Tris(2,2'-bipyridine) ruthenium(II) dichloride, (Ru(bpy)32+ CI2)

124

emission intensity of the sample either by the creation of a non-emissive dye-quencher

complex (static quenching) or by the depopulation of the excited state resulting from the

collision of the quencher during the excited state lifetime of the dye (dynamic

quenching).11 Dynamic quenching, which is commonly referred to as collisional

quenching, is the mechanism behind the Ru(bpy)32+ photoluminescence quenching by

molecular oxygen, Scheme 6.1.

These unique properties were exploited in the present study to record and image

intracellular O2 tension in single cardiomyocytes with Ru(bpy)32+ loaded into cells via the

whole cell patch clamp technique, and the sensitivity of the dye emission to external pC>2

changes and manipulation of mitochondrial respiratory rate were investigated.

6.2. Experimental.

Preparation of the Guinea Pig Ventricular Cardiomyocytes

Guinea pig ventricular cardiomyocytes were prepared as previously described.15 The

isolated cells were then stored at 37°C in Dulbecco's Modification of Eagle's Medium

(10-013 DMEM, Mediatech) containing 5% Fetal Bovine Serum (16140/071 FBS,

Invitrogen Corporation), 1% Penicillin-Streptomycin (30-001, 5000 ug/ml Mediatech),

and 1% HEPES Buffer Solution (15630, 1M, Gibco). All experiments were performed at

37°C after media exchange of DMEM with a modified Tyrode's solution containing

125

*o2 B

[Ru(bpy)3H 2 + i * 2+ . 3, -> [Ru(bpy)3

z • J02]

hv k hv;T=k'1 \ r, = kr1

Ru(bpy> 2+ Ru(bpy)32++ l02

Scheme 6.1. The dynamic (collisional quenching) mechanism for Ru(bpy)32+ by molecular oxygen. Once the Ru(bpy)32+ is excited by incident photons (hv), the excited state can either undergo A) a first-order PL decay (k~l) B) or quenching by colliding with triplet molecular oxygen. The latter leads to C) a bi-exponential decay (&'"1) and a decreased lifetime (Y).

126

(in mM): 140 NaCl, 5KC1, lMgCl2, 10 HEPES, 1 CaCl2, pH 7.5 (adjusted with NaOH),

supplemented with 10 mM glucose.

Patch-Clamp Technique

Physiological pipette solution containing Ru(bpy) s2+

The photoluminescent dye, Tris(2,2'-bipyridine)ruthenium (II) dichloride hexahydrate,

was made commercially available through Alfa Aesar and was used to reveal the changes

in oxygen tension within a single cardiomyocyte during changes in the metabolic state.

Due to the lack of membrane permeability of the plasma membrane to divalent Ru(II)

polypyridyl complexes, the technique of patch clamping was used to introduce the

Ru(bpy)32+ into the ventricular cardiomyocytes. The compound is water soluble and

dissolved readily in the physiological K-Glutamate pipette solution containing (in mM):

130 K-Glutamate, 19 KC1, 0.5MgCl2, 10 Na-HEPES, 1EGTA, pH 7.2 (adjusted with

KOH), supplemented with 50-100uM Ru(bpy)32+.

KATP Current Measurements

The whole-cell patch clamp technique was used to measure sarcolemmal KATP currents at

37°C. Borosilicate glass pipettes (outer diameter, 1.5mm; inner diameter, 1.0mm) were

pulled using Sutter Instruments Co. Model P-87 and polished to a 4-7 MO resistance with

a Narishige Scientific Instrument MF-83. Membrane currents were recorded in whole-

cell voltage clamp mode (Axopatch 200A amplifier, Digidata 1200B interface, Axon

Instruments). Electrophysiological signals were acquired, stored and analyzed using

127

custom-written software. The holding potential was -80mV, and the myocytes were

stimulated at 0.2 Hz with depolarizing pulses to +40mV for 100ms and then to 0 mV for

200ms before returning to the holding potential.

Optical Measurements

Mitochondrial Flavoprotein Fluorescence

The mitochondrial flavoprotein fluorescence, (arising predominantly from the oxidized

form of the flavins) was recorded at 525nm ±50 with excitation at the same wavelength

used to excite the Ru(bpy)32+ (480nm). The major component of mitochondrial

flavoprotein fluorescence arises from matrix dehydrogenases in equilibrium with the

NADH redox state; the signal was calibrated by completely oxidizing the mitochondrial

redox pool by applying the mitochondrial uncoupler carbonylcyanide-p-

trifluoromethoxyphenylhydrazone (FCCP; luM) and by inhibiting electron transfer to

oxygen, therefore completely reducing the flavoproteins with sodium cyanide. Both the

FCCP and the sodium cyanide solutions were diluted from concentrated stock solutions

into the physiological Tyrode's solution as described above.

Photoluminescence Recording of Myocytes

A two-channel photomultiplier tube assembly (PMT) (Photon Technology Inc. PTI-814)

was mounted on the side port of a Nikon Eclipse TE2000-E fluorescence microscope. A

dichroic mirror (550nm) was mounted in the emission cube to divide the emitted signal

into the short (530nm) and long (605nm) wavelength emission detectors. A Xe arc lamp

128

was used for excitation at 480nm using a dichroic mirror (490nm) mounted in the

microscope nosepiece.

Steady State Absorbance and Emission Measurements

Absorption spectra were acquired on a Cary Spectrometer using a 1cm x 1cm sealed

quartz cuvette to contain the sample. Steady state photoluminescence spectra of

Ru(bpy)32+ were recorded on a SPEX Fluorolog Spectrophotometer using a 450W Xe Arc

lamp as an excitation source. A Czeray-Turner monochrometer was used for emission

collection while the photoluminescence was measured with a Hamamatsu PMT.

Time Resolved Photoluminescence

The photoluminescence lifetime was obtained at room temperature using a Continuum

Surelite-III Nd:YAG pulsed laser to excite the Ru(bpy)32+. Third harmonic crystals were

used to produce 355nm laser excitation. A filter was positioned between the sample and

the emission monochrometer to reject scattered and frequency doubled photons. 610nm

light was monitored with a PMT mounted on a Czerny-Turner monochrometer and

collected with a Le-Croy digital Oscilloscope.

Extracellular p02 Measurements

An oxygen electrode (FOXY-R, Ocean Optics) was placed in the chamber containing the

perfusate solution. The sensor was adjacent to the cell, proximate to the coverslip

surface. The sensor was calibrated at 37°C in Tyrode's solution using the FOXY-CAL

129

in-house calibration system. Measurements were recorded using OOI sensor software

from Ocean Optics.

6.3. Results.

UV-Vis absorption and steady state emission spectra of the Ru(bpy)32+ in physiological

Tyrode's solution were normalized and are presented in Figure 6.2. Photophysical

properties for Ru(bpy)32+ measured in physiological Tyrode's solution at room

temperature are listed in Table 6.1.

Two absorption peaks appear in Figure 6.2 (bold line); the high-energy peak at 285nm

was assigned to the 7i- 7r* transition of the bipyridine ligands, while the broad low-

energy absorption band in the visible region (400-5 OOnm) depicts the metal-to-ligand

charge-transfer (MLCT) transition. The MLCT band, a red-shifted peak that is absent

from spectra of organic probes, allows for lower energy excitation of the sensor. The

extinction coefficient (s) of Ru(bpy)32+ in Tyrode's solution was found to be 1.4 x 104 M"

^m"1 for the ^^455 of the MCLT band. Monochromatic irradiation (455nm) of

Ru(bpy)32+ in physiological Tyrode's solution at room temperature resulted in the

observation of photoluminescence (PL) emission. The emission spectrum (dashed line)

exhibits a single broad peak red-shifted 150nm from the MLCT absorption peak. This

large Stoke's shift decreases the overlap of the absorbance and emission bands and

therefore eliminates the re-absorption at the wavelengths monitored.

130

0.00 - I ' 1 ' I ' 1 ' 1

300 400 500 600 700 800

Wavelength, nm

Figure 6.2. Normalized UV-Vis absorption (solid line) and steady state emission (dashed line) spectra of Ru(bpy)32+ in physiological Tyrode's solution. The spectra were collected at room temperature.

131

Tab

le 6

.1.

Phot

ophy

sica

l Pr

oper

ties

of

Ru(

bpy)

3 in

Phy

siol

ogic

al T

yrod

e's

Solu

tion

at R

oom

Tem

pera

ture

.

Sen

sor

Ru(

bpy)

32+

Fla

vin

Ade

nine

Din

ucle

otid

e

(FA

D)e

A-m

ax,a

bs>n

m

(e,

M-W

1)

285

/ 455

a

(1.4

x1

04)

-45

0

A-PL

, nm

615

-54

0

T, n

s*

605

N/A

x, n

sc

500

3.4 r

0.12

T, n

s"

107

N/A

o PL,

9

x10'

2

7.4

—

x1

0V

1

14.8

—

«nr>

x1

05s

1

18.5

—

" ^m

ax, ab

s of

the

7i->

7i*

/ML

CT

tra

nsit

ions

*'

c'd lif

etim

es w

ere

take

n in

sol

utio

n pu

rged

wit

h 10

0% n

itro

gen,

in

ambi

ent a

ir, a

nd p

urge

d w

ith

100%

oxy

gen,

res

pect

ivel

y.

e ref

eren

ce 1

1 fdo

uble

-exp

onen

tial

deca

y w

ith

lifet

imes

of

the

open

and

sta

cked

con

form

atio

ns,

resp

ecti

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" g i

ndex

of

ref

ract

ion

was

foun

d to

be

1.34

2 fo

r ph

ysio

logi

cal

salt

solu

tion

16

The photoluminescence quantum yield of Ru(bpy)3 varies depending on the solvent

used. The (|)PL was measured to be 0.073 in optically dilute physiological Tyrode's

solution, see Table 6.1. The quantum yield for photoluminescence is defined as the

fraction of molecules that emit a photon from the triplet excited state after direct

excitation by a source and is calculated using equation (6.1).12

<J)PL = (Ar/AsXIAXns/nr)2^ (6.1)

where Ar and As are the absorbances of the actinometer and the sample, respectively, Ir

and Is are the integrated photoluminescence intensities of the actinometer and the sample,

respectively, ns and nr are the indexes of refraction for the solvents used for the

actinometer and for the sample, respectively and §r is the quantum yield for Ru(bpy)32+ in

acetonitrile ((|)r = 0.062). n

The mechanism of quenching was first studied by Kautsy17 and is represented by the

Stern-Volmer equations (6.2) and (6.3).

IJI= rjt=\+ KSV[02] (6.2)

KSv = k2T0 (6.3)

133

where I0 and / are the emission intensities in the absence and in the presence of the

quencher, respectively, 70 and 7 are the lifetimes of the excited state in the absence and in

the presence of the quencher, respectively, Ksv is the Stern-Volmer quenching constant

and #2 is the bimolecular rate constant for the quenching of the excited state. In fluid

solution the Stern-Volmer plots of IJI and tj 7 versus quencher concentration are linear

with slope equal to Ksv. Therefore, with an increasing amount of oxygen present, the

photoluminescence intensity of the Ru(bpy)32+ in physiological Tyrode's solution is

quenched, Figure 6.3a. A total of 70.3% quenching of the Ru(bpy)32+

photoluminescence occurred once the oxygen-free solution was purged with 100% O2

directly. The inset represents a plot of the Stern-Volmer equation (6.2) as a function of

photoluminescence. A best-fit line, using equation (6.3), was drawn to estimate the

slope, KS\, to be 0.0025 +/- 0.0011. Due to the extended photophysical degradation

pathways within the excited state of metal-centered complexes, the lifetime becomes

much longer-lived than that of the organic probes, see Table 6.1. The time-resolved

photoluminescence decays were fit with a first-order kinetic model, and the k constant

was used to find the lifetime ( r = A:"1) of the excited state (Figure 6.3b.) The lifetime of

Ru(bpy)32+ was found to be 605ns, 500ns, and 170ns in physiological Tyrode's solution

purged with 100% nitrogen, in ambient air, and purged with 100% oxygen, respectively.

All measurements were acquired at room temperature. The inset depicts the Stern-

Volmer Plot as a function of lifetime as seen in equation (6.2). The Ksv was estimated

from the slope of the best-fit line, equation (6.3), and was found to be 0.0027+/- 0.0033.

Although 355nm excitation was used, variation of the wavelength of excitation will not

134

a)

550 600 650 700 750 800

Wavelength, nm

b) ~ 0.025-

c 0) £ 0.020-0 o S 0.015 u (A 0) | 0.010-

•g 0.005-

Q. 1 U U L . J J M A . J J

0.000

0.0 1.0x10 2.0x10*

Time, s

3.0x10

2+ Figure 6.3. Quenching of Ru(bpy)3 is observed through a) the 2+ quenching of Ru(bpy)3 photoluminescence and through b) the

2+ shortening of the Ru(bpy)3 lifetime. Each trace was acquired in a potassium glutamate pipette solution (previously described) purged with either 100% nitrogen (black trace), in ambient air (red trace), or purged with 100% oxygen (blue trace). The insets of a) and of b) depict the Stern-Volmer plots of the dynamic quenching, represented by equation (6.2), as a function of photoluminescence intensities and lifetimes, respectively. The Stern-Volmer quenching constants (Ksv) were calculated from the slope of the plots and using equation (6.3) to be 0.0025 V. 0.0011 and 0.0027 +/_ 0.0033.

135

change the lifetime kinetics nor will the concentration of the probe; the emission will

always originate from the lowest lying excited state.

The photophysical and redox properties of the endogenous mitochondrial flavoproteins

have been previously described.19 Several different mitochondrial matrix flavoproteins

exist that autofluoresce; however, the photophysics of the oxidized form (FAD, flavin

adenine dinucleotide) contribute most to the green fluorescence signal, depicted in Table

6.1.19'20 FAD is an organic cofactor for a number of mitochondrial enzymes, including

pyruvate dehydrogenase (PDH), a-hetogluterate dehydrogenase (a-KDH), Succinate

dehydrogenase(SDH), and the b-oxidation pathway of the electron transport flavin20

whose oxidation- reduction reaction occurs within the mitochondrial matrix. The short

lifetimes of FAD in aerated cells were found to be 3.4ns and 0.12ns for the stacked and

unstacked conformations, respectively11. The Xmax,abs was found to be ~450nm while the

?WX,PL was ~540nmu. This allowed for the simultaneous excitation of the FAD and the

loaded Ru(bpy)32+ with 480nm irradiation. The FAD green emission band resides within

the large Stoke's shift of the Ru(bpy)32+ complex; therefore, the green emission from the

FAD and the Ru(bpy)32+ photoluminescence were monitored simultaneously without

significant overlap of the emission bands.

Figure 6.4 illustrates the relationship between the green flavoprotein fluorescence, the red

Ru(bpy)32+ photoluminescence, and the sarcolemmal 7K,ATP- The myocyte was allowed to

load via whole cell patch clamp for -10-15 minutes until the Ru(bpy)32+

photoluminescence signal reached a steady state. After loading, the cell was exposed to

136

Time (min)

Figure 6.4. After loading of the cardiomyocyte was complete (prior to time zero of this plot), uncoupler (FCCP) was added, and a simultaneous rise in the green fluorescence (FAD oxidation) and the red photoluminescence (dequenching of Ru(bpy)32+) occurred. Shortly after, a rise in K,ATP current (black trace) was observed. These effects were reversed by exposure of the myocyte to the inhibitor NaCN. INSET: A single cardiomyocyte loaded with

2+

0.1 mM Ru(bpy)3 . The red circle in both images highlights the location of the membrane-pipette interface. The image intensity

2+

represents the intracellular Ru(bpy)3 emission at 605nm a) in the presence of a Tyrode's bath solution and b) in the presence of l.OmMFCCP.

137

the mitochondrial uncoupler, FCCP, Figure 6.4b. As a weak-acid protonophore, FCCP

can permeate the mitochondrial membrane due to the derealization of the electrons

throughout its 71-system.21'22 FCCP "short-circuits" the proton circuit and uncouples the

proton motive force from the generation of ATP at FiF0-ATPase. As a result, the rate of

respiration increases, and the oxidation of NADH and of the flavoproteins is enhanced

and an increase in the fluorescence of the latter occurs. In the process, oxygen is

consumed at Complex IV (cytochrome oxidase), therefore dequenching the Ru(bpy)32+

emission, resulting in an increase in photoluminescence intensity. Although respiration is

stimulated, the disruption of the protonmotive force causes a reversal of the FiFo-ATPase

and therefore, ATP consumption by the mitochondria.21 ATP hydrolysis eventually

exceeds the capacity of anaerobic metabolism (glycolysis) to compensate, accounting for

the delayed activation of the sarcolemmal K ATP channels ' ' as depicted in Figure 6.4a.

The addition of the electron transport chain inhibitor, NaCN, after washout of the FCCP,

completely reversed the effects of the uncoupler and causes maximal reduction of the

flavoprotein redox pool. ATP production and inhibition of KATP channels ensued. The

inset of Figure 6.4a illustrates the effects of 1.0 uM FCCP (b) on the emission of the

intracellular Ru(bpy)32+. There is an increase in photoluminescence as a result of the

dequenching of the ruthenium polypyridyl probe.

Ru(bpy)32+ loaded cells were then exposed to varying amount of the uncoupler, FCCP.

Cells are usually treated with 1.0 uM FCCP in the bath solution; this high concentration

allows for the cell to completely uncouple after a period of -1-2 minutes. Also, this

concentration can be applied to a cell for a period of only 5 minutes without causing

138

irreversible damage. However, the cardiomyocytes are found to be sensitive to much

lower concentration as seen in Figure 6.5. Here, FCCP solutions of increasing

concentration were titrated into the cell, but the period of exposure was lengthened to -10

minutes for each concentration to produce a response. As the concentration of FCCP

increased (in uM, 0.05, 0.1, 0.25, 0.50, 1.0) the emission intensity of the Ru(bpy)32+ as

well as the flavoproteins increased. This is indicative of uncoupling and of oxygen

consumption within the cell. The plot, Figure 6.5, illustrates a more rapid increase in

VO2 than in flavoprotein oxidation, when plotted as a function of Emiss/Emis0.

In order to achieve accurate quantitative measurements of the intracellular p02,

precautions were taken to ensure that the concentration of the loaded Ru(bpy)32+

remained unchanged. Loading via patch-clamp required -10-15 minutes, depending on

the pipette resistance, for the dye to reach an intracellular maximum concentration and

appear homogeneous throughout the cell. The emission intensity was recorded and

represented the steady-state photoluminescence intensity, Figure 6.6. Therefore, any

change in signal was directly correlated with a change in the intracellular oxygen

concentration and not due to a change in dye concentration. The variation in

intracellular p02 is a function of the rate of respiration and VO2 as well as a function of

the diffusion rate of molecular oxygen into the cell from the external solution and can be

represented by the following partial derivative,

d(pO?) = (srpOT) dt {8VO2;

dt + ext

5(E02}

8ext dt (6.4)

V02

139

a) 3.6

3.2 o

_l ^ 2.8-_ l Q. ,_ 2.4 O

£° 2.0

1.6-1

1.2

0.8 4 0

• Flavoprotein Fluorescence • Rutrisbpy Photoluminescence

200 400 600

[FCCP], nM

800 1000

b)

o

3.0-

2.5-

2.0-

1.5-

1.0- • 0

•

•

10

•

•

20 30 40

[FCCP], nM

•

•

50 60

Figure 6.5. a) The cardiomyocyte was exposed to several different concentrations of the uncoupler, FCCP (in nM: 0, 10, 25, 50, 1000). b) An enlargement of the plot at lower FCCP concentrations.

140

"8 N

1

!

0)

o c 0) o (A 0)

1.0

0.9

0.8

0.7

0.6

0.5

| 0.4

I

& & # *

i—'—r-0 2 8

— i — ' — i — • — i — • — i — • — i — • — i —

10 12 14 16 18 20

Time, min

Figure 6.6. The normalized photoluminescence plot indicative of the loading of Ru(bpy)3 technique.

2+ by the patch-clamp

141

where "ext" is the extracellular oxygen concentration. In order to assign corresponding

changes in intracellular oxygen concentration to each partial, the photoluminescence

intensity was monitored while holding one of the variables constant. It is known that the

penetration of oxygen into the cell occurs by simple diffusion through the cell membrane

Oft

until the intracellular and extracellular concentrations are in equilibrium. Therefore, an

external oxygen sensor was used to measure the bath pC>2, while respiration was inhibited

through the use of 4mM NaCN; the internal p02 during inhibition of respiration is a

direct measure of pC>2 changes in the bath. This allowed for the loaded dye to be

calibrated for the cell.

Figure 6.7 illustrates the effects of purging the bath solution with 100% O2 gas or 100%

N2 gas. An abrupt quenching of the Ru(bpy)32+ photoluminescence was observed upon

the addition of oxygen to the bath solution; in contrast, upon exposure of the cell to a

100% N2 purged Tyrode's solution, a dequenching of the signal occurred. The signal

changes were completely reversible. The flavoproteins were unaffected by changing the

external pC>2. Despite continuous purging with pure N2, the bath solutions were unable to

reach the extremely low pC>2 required to inhibit cytochrome oxidase and alter the basal

rate of respiration; a completely gas-tight chamber and purging lines would be required.

The KATP current was also unaffected by these bath changes.

142

3000 n Current Flavoprotein fluorescence (GREEN channel) Rutrisbpy emission (RED channel)

2500-

2000 4 - . . .

< 1500-

1000

500

0

,v-' /y

m

Oi

-..-4 0

180 €

- 125

- 100150

75

50

-I 25

120

90

c 0)

c

2 "3T o c 0)

o £ o 2

20 22 24 26

Time (min) 28

Figure 6.7. The effect of varying bath p02:. The cell was exposed to bath solutions saturated with 100% 0 2 or 100% N2 with expected effects

2+

on Ru(bpy)3 photoluminescence intensity but no change in flavin fluorescence.

143

In the next experiment, NaCN was also used to calibrate the Ru(bpy)3

photoluminescence signal. Figure 6.8 shows the effects of the NaCN purged with 100

%N2 and in ambient air; there is only a small difference in the flavoprotein fluorescence

when exposed to a higher pC>2 in external bath. On the other hand, intracellular oxygen,

represented by the Ru(bpy)32+ photoluminescence signal, increased significantly (a

decrease in the PLI).

A single ventricular cardiomyocyte was loaded with a 50 uM Ru(bpy)32+ K-glutamate

pipette solution. This solution was altered from its original composition to enhance Ca2+

cycling and contractions, providing an increase in the energy demand. EGTA was used

in low concentration (0.1 mM) to reduce Ca2+ buffering, while the Na+ concentration was

raised to 15mM. The cell was then exposed to 4mM NaCN for ~4 minutes and washed

out with a physiological Tyrode's solution. The pC>2 of the environment was measured

simultaneously with an external oxygen electrode. A calibration was made from the

recorded photoluminescence and the corresponding external pC^; Figure 6.9 illustrates

this calibration. The solubility coefficients for O2 in buffer solution and in myocytes, for

conversion of % O2 to uM O2, were found in literature to be 1.30xl0'6 M/mmHg and

1.74xl0"6 M/mmHg, respectively, at 37°C.27 The cell was also paced at 2Hz to measure

the respiration during stimulation. The resting intracellular oxygen concentration was

found to be 197uM (14.9% 0 2 or 113mmHg) and was reduced to 150uM (11.4% 0 2 or

86.7mmHg) once twitching occurred, see Figure 6.10. There was an overall decrease of

-30.3 uM/ min. Before death, the cell became oxidized and completely uncoupled, and

the intracellular oxygen concentration was reduced to 2.06uM (1.56% O2 or 11.8mmHg).

144

4.0VV-

2400

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1 1600 c o

w 1200-

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800

400

t

CN'/N2

•

•

CN" / N2

• 1 i

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Flavoprotein Fluorescence

RiXbpyJj2* Photduminescence

CN'/air

• 1 i ' i ' i '

1

i '

12 14 16 18 20 22

%a

2+ Figure 6.8. The flavoprotein fluorescence and Ru(bpy)3 photoluminescence were recorded in the presence of 4mM NaCN purged with 100 %N2 or in ambient air. There is a larger change in the intracellular concentration, depicted by the Ru(bpy)32+

photoluminescence, than in the flavoprotein oxidation. This illustrates the effect of diffusion of oxygen from the extracellular medium during inhibited respiration.

145

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Time (min)

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Error

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N

-0.93461 12.01264 13

I

15 20 25

%0„

I

30 i

35 40

Figure 6.9. Calibration of the Ru(bpy)3 photoluminescence signal in the presence of NaCN. a) The flavoprotein redox properties ( ), the Ru(bpy)32+

photoluminescence ( ), and the external % O2 ( ) were recorded during a period of 4min exposure to NaCN purged with 100% N2, washout with Tyrode's solution, and then another 4min period of NaCN purged with 100% O2. The measurements were completed at 37°C. b) Ru(bpy)32+ photoluminescence values, taken during periods of NaCN (purged with either N2 or O2), were plotted with their corresponding external O2 concentrations. A best-fit line was used for calibration.

146

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R SD N

Error

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3 0.6 0.8 1.0 1.2 1.4 1.6 1.8 2.0 2.2 2.4

Time, min

Figure 6.10. a) The cardiomyocyte (also used in the measurement in Figure 6.9) was stimulated at 2Hz from the start of this recoding to induce twitching. Under conditions of basal respiration, Ru(bpy)32+

photoluminescence signal ( ), was measured to be 14.9 %02 (197u.M). Oxidation of the flavoproteins ( ) was immediately instigated upon stimulation. Before cell death, a spike in both the Ru(bpy)32+

photoluminescence and the flavoprotein fluorescence appeared with a measured intracellular %02 of 1.56% (2.06uM). b) The slope of the Ru(bpy)32+ photoluminescence curve that resulted from stimulation was fit with a best fit line. The overall decrease in intracellular oxygen content was measured to be 30.3uM/min.

147

Throughout the duration of this experiment, the temperature of the open chamber was

held constant while the external electrode measured a relatively constant oxygen

concentration 208uM (21% O2 or 160mmHg). To ensure that metabolic oscillations did

not occur, 1.5mM reduced glutathione was added to the pipette solution to function as a

ROS scavenger.

Applications have proven successful using this novel method to study cellular respiratory

function during metabolic or oxidative stress. Guinea pig ventricular cardiomyocytes

were susceptible to intrinsic oscillations of energy metabolism and were associated with

cyclical activation of ATP- sensitive potassium currents, Figure 6.11. These oscillating

currents were not provoked here by an electrical stimulus or pacemaker activity;

metabolic oscillations developed after the cell was exposed for ~5 minutes to a solution

purged with 100% O2, constituting a form of oxidative stress. The green trace in Figure

6.11 represents the flavoprotein fluorescence; an increase in fluorescence intensity

coincides with a net oxidation of the flavins and an increase in respiration. The red trace,

exhibiting parallel changes in intensity, symbolizes the Ru(bpy)32+ photoluminescence,

which is indicative of the intracellular pC>2. There is a slight delay in the oscillatory KATP

current activation, which consistently shadows the signal of the flavin oxidation and the

Ru(bpy)32+ photoluminescence dequenching. This is characteristic of sarcolemmal KATP

currents; see Figure 6.4.15 The oscillating photoluminescence signal represents the pC>2

changes occurring directly due to respiration, while the slightly slower variation in the

baseline correlates to the exchanging of the bath solution to one purged with 100 % O2 (a

decrease at 5min) and then to one purged with 100% N2 (an increase at 16 min),

148

Figure 6.11. Metabolic oscillations were induced in cardiomyocytes by a brief period of high p02 (bath solution saturated with 100% 02). 100% N2 saturated solution was applied after -15 minutes, but oscillations were not suppressed.

149

respectively. These oscillations were best observed in freshly isolated and uncultured

ventricular cardiomyocytes.

6.4. Discussion.

The photophysical degradation pathways of excited state inorganic complexes differ

greatly from that of organic sensors; the lower energy triplet state of the metal complex

allows for these additional photophysical pathways in the excited state and therefore,

longer lived excited state lifetimes. The longer the lifetime of the oxygen probe, the

greater the probability of probe-quencher encounters during the excited state. This leads

to an overall more efficient and more sensitive oxygen probe. Using Ru(bpy)32+ to probe

the intracellular pC>2 has proven to be a successful, inexpensive tool that can be utilized in

future experiments to directly relate other known metabolic functions to intracellular

oxygen tension.

Many factors were taken into account when choosing a molecular oxygen probe.

Although Ru(bpy)32+ possessed the ideal photophysical and chemical characteristics and

energetics desired for our experiments, derivatives of this transition metal complex can

be synthesized. The synthetic addition of electron withdrawing ligands or additional %-

systems allows for fine-tuning of the electronic properties to achieve the desired spectral

characteristics, quantum yields, and lifetimes.28 Furthermore, the functionalization of

150

Ru(bpy)32+ with lipophilic ligands such as ethyl esters, may permit loading of the dye in

culture in lieu of utilizing the whole cell patch-clamp technique for dialysis.

However, the cell membrane impermeability to Ru(bpy)3 + can be viewed as an

advantage; it ensures that the dye does not accumulate in the mitochondrial matrix, in the

nucleus, or leak out of the cell. Accumulation of lipophilic probes in the matrix or in the

nuclear compartments can complicate interpretation of the results.22 Therefore, the

concentration of the Ru(bpy)32+, once steady state has been reached during dialysis,

remains unchanged, and any variations in signal are solely due to the changing

intracellular O2 levels. Once the experiment was complete, the original environmental

conditions were reestablished and the emission intensity of the dye within the cell was

again recorded; no change in intensity within instrumental error was observed. In

addition, this control illustrated that there was minimal photobleaching of the Ru(bpy)32+

throughout the duration of the experiments.

The loaded dye was found to be nontoxic to the isolated ventricular cardiomyocytes, but

the cells did exhibit some sensitivity to the photo-excited dye under prolonged periods or

irradiance. The cells were more sensitive to concentrations above 0.1 mM Ru(bpy)32+ in

the pipette solutions, depending on the intensity and duration of illumination. Neutral

density filters were used to decrease the intensity of the incident light, and a lower

frequency of light exposure was used to reduce free radical-mediated toxicity. It must be

noted that in solution, the excited state complex (exciplex) formed from the collision of

molecular oxygen ( O2) with the Ru(bpy)32+ excited state can result in a superoxide

151

product (202*")- Superoxide radicals, appear when the exciplex molecule, acting as an

electron donor, transfers an electron to a nearby 3C>2. Thus, photosensitivity of the cell is

believed to be due to the generation of the superoxide radical (O2"'). Although the yield

of O2"' is low in comparison to that of singlet oxygen ( O2) , a higher intensity excitation

leads to a greater number of excited states and therefore a proportional increase in

byproducts. Reduced glutathione, a thiol reagent, decreased the toxicity by acting as a

scavenger of free radical oxygen species byproducts (e.g. H2O2; not shown) through the

glutathione peroxidase.30 Endogenous superoxide dismutase (SOD) enzymes are present

in the cytopolasm and mitochondrial compartments to decrease the intracellular O2"

levels arising from respiration.30 Superoxide is a natural intrinsic byproduct of oxidative

phosphorylation; however, there is a maximum allowable threshold for O2"" concentration

that when crossed, can be toxic to cells. At lower power irradiation or concentrations of

Ru(bpy)32+ under ImM, O2"' production did not result in observable toxicity (increased

background leak current or hyper-contraction) Singlet oxygen, produced from O.lmM

Ru(bpy)32+, has proven to be a nontoxic, short-lived byproduct of this dynamic

quenching; it occurs in high yield, relative to that of the O2", also by electron transfer

from Ru(bpy)32+ to 302 .29 Any reduced Ru(bpy)32+ is regenerated by back-electron

transfer from these oxygen byproducts. In general, the most commonly produced

byproduct is triplet molecular oxygen that is released from the exciplex in its original

form.

This electron transfer from the ruthenium complex to the molecular oxygen is

thermodynamically and energetically favorable as a result of their compatible reduction

152

potentials. However, the high reduction potential of Ru(bpy)3 does not lead to direct

electron transfer to any complexes, electron shuttles, or flavoproteins of the electron

transport chain because the dye cannot penetrate the mitochondrial membrane where

these acceptor complexes reside. The CCD images of the Ru(bpy)32+ distribution also

displayed no significant accumulation in the mitochondria or nucleus.

Metabolic oscillations in fresh cells were observed after a brief period of exposure to

100% O2 purged physiological bath solution. These results demonstrated the sensitivity

of the ruthenium bipyridine sensor to rapid changes in internal oxygen concentration due

to respiration. Direct relationships can be made between the oxidation of the flavins, the

activation /KATP, and the intracellular oxygen concentration. In addition, relationships

between NADH redox, membrane potential, ROS concentrations, and /KATP during

metabolic oscillations have been previously reported. ' ' There is a phase shift

between the NADH oxidation and the activation of the sarcolemmal KATP current,15 and

an indirect relationship can be inferred from comparing the current results to the

literature. Therefore, as NADH is oxidized, there is a simultaneous oxidation of the

flavins and a decrease in intracellular pC>2 due to the uncoupling of the mitochondrial

respiration. A direct relationship between the NADH pool and the membrane potential

was investigated; the depolarization of the membrane coincided with the oxidation of

NADH. 32 In a computational model of the mitochondrial oscillator, Cortassa et al. found

that an increase in ROS preceded a depolarization of the mitochondrial membrane.

153

6.5. Conclusions.

The findings indicate that single myocyte oxygen measurements can be made to directly

observe changes in mitochondrial respiration, which will be a useful tool for investigating

fundamental mechanisms of energy supply-and-demand matching in the heart.

154

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Hitchman, M.C. Measurement of Dissolved Oxygen. Wiley, New York, 1978.

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Albani, J.; Alpert, B. Eur. J. Biochem. 1987, 162 (1), 175-178.

O'Regan, B.; Gratzel, M. Nature 1991, 353, 737-739.

Demas, J.N., Degraff, B.A. Anal Chem. 1991, 83 (17), 829-837.

Gerardi, R.D.; Barnett, N.W.; Lewis, S.W. Anal. Chim. Acta. 1999, 378, 1-43.

Liu, J.G.; Ye, B.H.; Li, H.; Zhen, Q.X.; Ji, L.N.; Fu, Y.H. J. Inorg. Biol. 1999,

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(a) Wu, X.J.; Choi, M.F.; Xiao, D. Analyst. 1999, 125, 157-162. (b) Choi, S.J.;

Choi, B.G.; Park, S.M. Anal. Chem. 2002, 74 (?), 1998-2002.

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9. Kalyanasundaram, K. Coord. Chem. Rev. 1982, 46, 159-244.

10. Balzani, V.; Bolette, F.; Grandolfi, M.T.; Maestru, M. Top. Curr. Chem. 1978, 75,

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11. Lakowicz, J.R. Principles of Fluorescence Spectroscopy; 2nd ed. Kluwer

Academic Publishers/ Plenum Publishers. New York: New York. 1999.

12. Demas, J.N.; Crosby, G.A. J. Phys. Chem. 1971, 75 (8), 991-1024.

13. Demas, J.N.; DeGraff, B.A. Anal. Chem. 1991, 63, 829A-837.

14. Fuller, Z.J.; Bare, W.D.; Kneas, K.A.; Xu, W.-Y.; Demas, J.N.; DeGraff, B.A.

Anal. Chem. 2003, 75, 2670-2677.

15. O'Rourke, B.; Ramza, B.M. ;Marban, E. Science. 1994, 265, 962-966.

16. Rolling, O.W. Transactions of the Kansas Academy of Science 1995, 98 (1-2),

72-75.

17. Kautsy, H. Trans. Faraday Soc. 1939, 35, 216-219.

18. Lewis, G.N.; Kasha, M. J. Amer. Chem. Soc. 1944, 66, 2100-2116.

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19. Chance, B.; Ernster, L.; Garland, P.B.; Lee, C.P.; Light, P.A.; Ohnishi, T.; Ragan,

C.I.; Wong, D. Proc. Natl. Acad. Sci. 1967, 57, 1498-505.

20. a) Kunz, W.S.; FEBS Lett. 1986, 195 (1-2), 92-96. b) Kunz, W.B.; Kunz, W.

Biochim. Biophys. Acta 1985, 841 (3), 237-246.

21. Nicholls, D.G.; Ferguson, S.J. Bioenergetics. Academic Press: London, UK.

2002.

22. (a) Benz, R.; McLaughlin, S., Biophys. J. 1983, 41, 381-398. (b) Noma, A.

Nature 1983, 305, 147-148.

23. Sasaki, N.; Sato, T.; Marban, E.; O'Rourke, B. Am. J. Heart Circ. Physiol. 2001,

280, H1882-H1888.

24. Romasko et al 1998

25. Nelson, D. L., Cox, M.M. Lehniger Principles of Biochemistry: 3 ed. Worth

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Hasenfuss, G. JMol Cell Cardiol 1998, 30, 1459-1470

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28. Dong, W.; Mendelsohn, R.; Galoppini, E.; Hoertz, P.G.; Carlisle, R.A.; Meyer,

G.J. J. Phys. Chem. B 2004, 108, 16641-16653.

29. Demas, J.N., Harris, E.W., McBride, R.P. J. Am. Chem. Soc. 1977, 99 (11),

3547-3551.

30. Chance, B., Williamson, J.R., Schoener, B. Biochem Z. 1965, 341, 357-377.

31. Cortassa, S, Aon, M.A, Winslow, R.L., O'Rourke, B. Biophys J. 2004, 87, 2060-

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2007, 557, 8-14.

158

CURRICULUM VITAE

EDUCATION

The Johns Hopkins University, Baltimore, MD

Ph.D. Candidate with emphasis on physical-inorganic chemistry applied to

cardiology, September 2007

Master of Arts Chemistry, April 2003

James Madison University, Harrisonburg, VA

Bachelor of Science in Chemistry, May 2001

Minor: Studio Art, Pre-medicine

HONORS

Member of the James Madison University Honors Program, 1997-2000

Recipient of the Madison Achievement Scholarship, 1997

159

EXPERIENCE

The Johns Hopkins University, Baltimore, MD

Research Assistant, April 2001 - Present

• Maintain and advance four major organic/ inorganic and cardiology research

projects

• Develop novel protocols to interface the two fields of chemistry and cardiology

• Perform Langendorff isolations of ventricular guinea pig cardiomyocytes

• Patch clamp individual ventricular cardiomyocytes to obtain electrophysiological

results

• Examine research results by means of chemical and physical analyses

• Create presentations and posters for local and national conference symposiums

• Present research results weekly to coworkers, postdoctoral fellows, professors,

and medical professionals

• Employ analytical and critical thinking skills to independently identify and solve

problems

• Utilize and cultivate written communication through journal publications

Teaching Assistant in Intermediate Chemistry, January 2003 - May 2003

• Administered personal help sessions to review and clarify lectures

• Organized and led review classes before exams

• Graded exams and finals

• Created curriculum for and taught several lectures to a 150-student class

160

Teaching Assistant in Physical Chemistry, August 2002 - December 2002

• Graded exams and homework problem sets

• Held individual help sessions

Teaching Assistant in Physical Chemistry Laboratory, August 2001 - May 2002

• Educated senior engineer and chemistry students on Fourier Transform Nuclear

Magnetic Resonance (FTNMR)

• Graded laboratory reports

Villa Julie College, Baltimore, MD

Instructor of Physical Chemistry (Thermodynamics and Quantum Chemistry), January

2005- Present

• Create a curriculum for both semesters of physical chemistry (Thermodynamics

and Quantum Chemistry)

• Educate senior level students on the states of matter, chemical equilibrium,

chemical kinetics, and electrochemistry

• Extend the students' knowledge of calculus by providing in depth mathematical

derivations of thermodynamic and quantum mechanical concepts

• Develop the students' understanding of classical and quantum mechanics,

quantum theory behind atoms and molecules, theory of spectroscopy and solid

state theory

• Design new laboratory curriculums in thermodynamics and quantum chemistry

161

Instructor of General Chemistry, Summer 2006

• Developed student knowled'ge of general chemistry theories and enhanced their

dexterity in problem solving

James Madison University, Harrisonburg, VA

Research Assistant in chemistry, 1999-2001

• Investigated the relationship between protein structure and function in

Escherichia coli protein, RecA

• Co-authored and illustrated a computer tutorial program for incoming inorganic

college students

Teaching Assistant in general chemistry laboratory, 1998-1999

• Promoted high-quality laboratory techniques

ACTIVITIES

Private Tutor, The Johns Hopkins University, Summer 2004- Present

• Prepare and home-tutor high school students for the chemistry Advanced

Placement exam

• Teach Intermediate Chemistry to several post-bachelor premedical students

162

Sigma Kappa Sorority, James Madison University

Sigma Kappa Sorority President, 2000

• Dedicated time to fostering sisterhood among its 160 members through

philanthropic, social and academic events

• Organized and ran weekly chapter meetings and rituals

• Acted as liaison between the chapter, the JMU sorority administration, and the

national counsel

Senior Ethics Committee and Standards Board Chair, 2001

• Promoted ethical conduct and behavior by confronting and reprimanding

members

Risk Manager, 1999

• Created new bylaws that helped reduce and eliminate negligent behavior

• Oversaw risk management committee to enforce existing policies

Best Buddies Organization

Vice President of Public Relations, 1997-2000

• Maintained a "best-buddy" relationship with mentally challenged and physically

disabled adults

• Established monthly events to involve the community with Best-Buddies

163

POSTERS

• Carlisle, R.A., O'Rourke, B. "Measurements of Oxygen Tension in Single

Cardiomyocytes." POSTER. Biophysical Society. Baltimore, MD. March

2007.

• Carlisle, R.A., O'Rourke, B. "Measurements of Oxygen Tension in Single

Cardiomyocytes: Photoluminescence Quenching of Intracellular Ru(bpy)32+ by

Molecular Oxygen." POSTER. Biophysical Society. Salt Lake City, UT.

February 2006.

• Carlisle, R.A. Meyer,G.J. "Organic Rigid- Rod Linkers for Coupling

Chromophores to Metal Oxide Nanoparticles." POSTER. National Science

Foundation. Washington, D.C. February 2004.

PRESENTATIONS

• Carlisle, R.A. "Measurements of Oxygen Tension in Single Cardiomyocytes:

Photoluminescence Quenching of Intracellular Ru(bpy)32+ by Molecular

Oxygen." PRESENTATION. University of Oslow. Oslow, Norway. February

2006.

164

• Carlisle, R.A. "Photophysical and Electrochemical Properties of Pyrene Rigid-

Rod Derivatives." PRESENTATION. The Johns Hopkins University:

Inorganic Nights; Baltimore, MD. September 2005.

PUBLICATIONS

• Taratula, O.; Rochford, J.; Piotrowiak, P.; Carlisle, R.A.; Meyer, G.J.;

Galoppinni, E. "Pyrene- Terminated Phenyenethynylene Rigid Linkers

Anchored to Metal Oxide Nanoparticles." J. Phys. Chem B 2006, 110, 15734-

15741.

• Hoertz, P.G.; Carlisle, R.A.; Meyer G.J.; Wang, D.; Mendelsohn, R.; Galoppini

E. "Excited State Electron Transfer from Ru(II) Polypyridyl Compounds

Anchored to Nanocrystalline Ti02 Through Rigid-Rod Linkers." J. Phys. Chem B

2004,108,16642-16653.

• G.J.; Wang, D.; Piotrowiak, P.; Galoppini, E.; Hoertz, P.G.; Carlisle, R.A.;

Meyer, "Organic Rigid-Rod Linkers for Coupling Chromophores to Metal Oxide

Nanoparticles." Nanoletters 2003,3, 325-330.

165

PROFESSIONAL ASSOCIATIONS

American Chemical Society, member

Materials Research Society, member

American Heart Association, member

Biophysical Society, member

166