ELECTRONIC AND VIBRATIONAL DYNAMICS OF HEME MODEL COMPOUNDS-AN ULTRAFAST SPECTROSCOPIC STUDY By JAGANNADHA REDDY CHALLA Submitted in partial fulfillment of the requirements For the degree of Doctor of Philosophy Dissertation Advisor: Dr. M. Cather Simpson Department of Chemistry CASE WESTERN RESERVE UNIVERSITY AUGUST, 2007 CASE WESTERN RESERVE UNIVERSITY SCHOOL OF GRADUATE STUDIES We hereby approve the dissertation of ______________________________________________________ candidate for the Ph.D. degree *. (signed)_______________________________________________ (chair of the committee) ________________________________________________ ________________________________________________ ________________________________________________ ________________________________________________ ________________________________________________ (date) _______________________ *We also certify that written approval has been obtained for any proprietary material contained therein. Dedicated to My loving uncle Subba Reddy TABLE OF CONTENTS TABLE OF CONTENTS i LIST OF TABLES v LIST OF FIGURES viii LIST OF ABBREVIATIONS xxiii ACKNOWLEDGEMENTS xxv ABSTRACT xxvi CHAPTER 1. INTRODUCTION 1 1.1 Heme proteins and biological significance 1 1.2 Heme model compounds 4 1.3 Electronic dynamics 10 1.4 Vibrational dynamics 15 1.5 Objectives 19 1.6 References 21 CHAPTER 2. EXPERIMENTAL METHODS AND INSTRUMENTATION 30 2.1 Introduction 30 2.2 Transient absorption spectroscopy 32 2.3 Time resolved resonance Raman spectroscopy (TR3S) 33 2.4 Generation and amplification of ultrashort pulses 36 i 2.5 Pulse duration measuerement of ultrashort pulses 38 2.6 CLARK MXR INC. CPA-1000 fs laser system 39 2.7 Single color pump-probe setup 42 2.8 Quantronix Integra fs laser system 46 Wavelength tunability 48 Fs visible and IR OPA 49 Ps visible OPA 53 CHAPTER 3. THE ELECTRONIC AND VIBRATIONAL DYNAMICS OF II FEOEP-2MeIm IN CH2Cl2 67 3.1 Introduction 67 3.2 Materials and methods 70 3.3 Results 73 3.4 Discussion 87 3. 5 Conclusions 96 3.6 References 97 CHAPTER 4. THE ELECTRONIC AND VIBRATIONAL DYNAMICS OF II FeOEP-(Im)2 in CH2Cl2 104 4.1 Introduction 104 4.2 Materials and methods 107 4.3 Results 110 ii 4.4 Discussion 120 4.5 Conclusions 125 4.6 References 126 CHAPTER 5. THE ELECTRONIC DYNAMICS of FeIIOEP-2MeIm and II FeOEP-(Im)2 in CH2Cl2 UPON Q-BAND EXCITATION 133 5.1 Introduction 133 5.2 Materials and methods 136 5.3 Results and discussion 138 5.4 Conclusions 146 5.5 References 147 CHAPTER 6. EFFECT OF METHYL GROUPS ON THE VIBRATIONAL DYNAMICS OF PARA-NITROANILINE 149 6.1 Introduction 149 6.2 Materials and methods 157 6.3 Results 159 NNDMPNA 163 2-MPNA 166 2, 6-DMPNA 170 iii 6.4 Discussion 173 6.5 Conclusions 178 6.6 References 179 APPENDIX 187 BIBLIOGRAPHY 194 iv LIST OF TABLES Table 2-1 Summary of the fs UV-visible OPA output wavelengths and their energies. I: idler, S: signal, SHI: second harmonic of idler, SHS: second harmonic of signal, FHI: fourth harmonic of idler, FHS: fourth harmonic of signal, SHSFI: second harmonic of SFI, SHSFS: second harmonic of SFS, 2B: Type I BBO crystal, 3B: Type II BBO crystal, 5B: Type I BBO crystal, 6B: Type I BBO crystal, H: horizontal, V: vertical. --------------------------------------- 52 Table 2-2 Summary of the fs IR OPA output wavelengths and their energies. I: idler, S: signal, NDF: nonlinear difference frequency, 8B: Type I AgGaS2 crystal, 9: Type I GaSe crystal, H: horizontal, V: vertical. ---------------------------------------------------------------------- 53 Table 2-3 Summary of the ps UV-vis OPA TWNB output energies. S: signal, SHS: second harmonic of signal, H: horizontal, V: vertical. -------- 54 Table 3-1 Vibrational Dynamics in Photoexcited FeOEP-2MeIm. aDetermined from simulations of the kinetic traces in Figure 3-3 using the fit decay life time, a pump/probe relative flux of 0.4 and convolution with a 600 fs Gaussian function. bFrom fitting only the decay portions of the kinetic traces in Figure 3-3. cFrom fitting only the bleach recovery portions of the kinetic traces in Figure 3- 5. ----------------------------------------------------------------------------- 79 Table 3-2 Kinetic Analysis for TA experimental Data and Simulations. ar2 > v 0.99 for all fits except the last three experimental ones, which were 0.987, 0.979, 0.966. ------------------------------------------------- 82 Table 3-3 Parameters used in TA simulations. aSee text for details of the b equations used to simulate the TA data. The ν (0) values for S0* -1 are red-shifted 300 cm relative to the values for the thermal S0 ground state spectrum. The ω(0) values for S0* are 1.2 x the values for S0. --------------------------------------------------------------- 85 Table 4-1 Exponential Fit parameters to Experimental TA Measurements of II FE OEP-(Im)2 in CH2Cl2 using Soret band pump white light continuum probe. R^2 values in order from top: 0.99, 0.99, 0.99, 0.96, 0.97, 0.98, 0.99, 0.99, 0.98, 0.98, 0.95, 0.95, 0.94, 0.98, 0.96, 0.97, 0.93, and 0.94. ------------------------------------------------------- 114 Table 5-1 Exponential Fits to Experimental TA Measurements of FeIIOEP- 2MeIm in CH2Cl2 following excitation at 541 nm. ------------------- 141 Table 5-2 Exponential Fits to Experimental TA Measurements of FeIIOEP- (Im)2 in CH2Cl2 following excitation at 541 nm. ---------------------- 145 Table 6-1 Selected experimental and calculated vibrational frequencies (in cm-1). aExperimental data in DMSO obtained with 403 nm (PNA), 403.5 nm (NNDMPNA), 410.5 nm (2-MPNA), 410.35 nm (2, 6- DMPNA). bScaling factor 0.98; cAssignment and Mode descriptions according to Varsanyi45 and Kozich et al15, 41. --------- 162 Table 6-2 Effect of methyl group substitution on the vibrational energy relaxation lifetimes of PNAa. aLifetimes obtained by plotting the vi transient integrated or average peak intensity as a function of time delay. bLifetimes of PNA obtained from Gunaratne et al. ChemPhysChem 2005, 6, 1157-63. -------------------------------------- 175 Table 6-3 Effect of methyl group substitution on the vibrational energy relaxation lifetimes of PNAa.aLifetimes obtained using change in vibrational frequency as a function of the time delay. bLifetimes of PNA obtained from Gunaratne et al. ChemPhysChem 2005, 6, 1157-63. --------------------------------------------------------------------- 175 vii LIST OF FIGURES Figure 1-1 The chemical structure of Fe protoporphyrin IX also known as protoheme. ------------------------------------------------------------------ 1 Figure 1-2 The chemical structure of the heme prosthetic group present in deoxy form of the Mb. ---------------------------------------------------- 3 Figure 1-3 Edge on view of the heme group present in Hb and Mb. The porphyrin macrocycle is domed in five-coordinate form and changes to planar upon O2 binding with simultaneous change from high to low spin state. ---------------------------------------------- 3 Figure 1-4 The chemical structure of 2-methyl imidazole complex of iron octaethyl porphyrin [FeIIOEP-2MeIm]. It is a five-coordinate high- spin heme. ------------------------------------------------------------------ 5 Figure 1-5 The chemical structure of bis-imidazole complex of iron II octaethylporphyrin [Fe OEP-(Im)2] It is a six-coordinate, low- spin heme. ------------------------------------------------------------------ 6 II Figure 1-6 The UV-vis absorption spectrum of Fe OEP-2MeIm in CH2Cl2. The Q-band is an order of magnitude weaker than B band. --------- 7 II Figure 1-7 The UV-vis absorption spectrum of [Fe OEP-(Im)2] in CH2Cl2. The six-coordinate, low-spin ferrous compound has two band on the Q band region corresponding to Q00 and Q01. --------------------- 8 II Figure 1-8 The Stokes resonance Raman spectrum of Fe OEP-2MeIm in CH2Cl2 obtained with 413 nm excitation. ------------------------------ 9 viii II Figure 1-9 The Stokes resonance Raman spectrum of Fe OEP-(Im)2 in CH2Cl2 obtained with 410 nm excitation. ------------------------------ 9 Figure 1-10 Parallel and sequential intermediate excited electronic state decay models of Petrich et al. and Franzen et al. representing the electronic relaxation dynamics of both ligated and unligated heme proteins as well that of an iron PPIX. ----------------------------------- 13 Figure 1-11 Vibrationally hot ground electronic state models of Kholodenko et al and Ye et al. representing the electronic and ground state vibrational relaxation of heme proteins. -------------------------------- 14 Figure 1-12 General experimental scheme representing the monitoring of intramolecular vibrational energy redistribution (IVR) and intermolecular vibrational energy relaxation (VER) using Raman scattering following IR or UV-vis excitation. ------------------------- 16 Figure 2-1 General Pump-Probe scheme in time-resolved spectroscopy. ------- 30 Figure 2-2 Energy level diagrams for Stokes and anti-Stokes Raman scattering. ------------------------------------------------------------------- 34 Figure 2-3 Principle of chirped pulse amplification. The femtosecond seed pulse is stretched in time prior to amplification and compressed back to its near original duration subsequent to amplification. ------ 37 Figure 2-4 The schematic layout of the various components of Clark-MXR Inc. CPA-1000 fs laser system. ------------------------------------------ 39 Figure 2-5 The second harmonic spectrum of the CPA-1000 output after frequency doubling in a 30 mm long KDP crystal. The nonlinear ix Gaussian and Lorentzian peak fits together with the corresponding bandwidths are shown. The spectral