Fluorescence methods are being used increasingly in biochemical, medical, and chemical research. This is because of the inherent sensitivity of this technique. and the favorable time scale of the phenomenon of fluorescence. 8 Fluorescence emission occurs about 10- sec (10 nsec) after light absorp tion. During this period of time a wide range of molecular processes can occur, and these can effect the spectral characteristics of the fluorescent compound. This combination of sensitivity and a favorable time scale allows fluorescence methods to be generally useful for studies of proteins and membranes and their interactions with other macromolecules. This book describes the fundamental aspects of fluorescence. and the biochemical applications of this methodology. Each chapter starts with the -theoreticalbasis of each phenomenon of fluorescence, followed by examples which illustrate the use of the phenomenon in the study of biochemical problems. The book contains numerous figures. It is felt that such graphical presentations contribute to pleasurable reading and increased understand ing. Separate chapters are devoted to fluorescence polarization, lifetimes, quenching, energy transfer, solvent effects, and excited state reactions. To enhance the usefulness of this work as a textbook, problems are included which illustrate the concepts described in each chapter. Furthermore, a separate chapter is devoted to the instrumentation used in fluorescence spectroscopy. This chapter will be especially valuable for those perform ing or contemplating fluorescence measurements. Such measurements are easily compromised by failure to consider a number of simple principles."

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Principles of fluorescence spectroscopy

For more than 70 years, researchers in several areas of science have been intrigued by the physical and chemical properties of the lowest excited states of molecular oxygen. With two singlet states lying close above its triplet ground state, the O2 molecule possesses a very unique configuration, which gives rise to a very rich and easily accessible chemistry, and also to a number of important photophysical interactions. In particular, photosensitized reactions of the first excited state, O2(∆g), play a key role in many natural photochemical and photobiological processes, such as photodegradation and aging processes including even photocarcinogenesis. Reactions of O2(∆g) are associated with significant applications in several fields, including organic synthesis, bleaching processes, and, most importantly, the photodynamic therapy of cancer, which has now obtained regulatory approval in most countries for the treatment of several types of tumors. The development of both applications and novel observation techniques has strongly accelerated during the past few years. Significant recent advances include, for example, the development of novel luminescent singlet oxygen probes,1-4 the time-resolved detection of O2(∆g) in a transmission microscope,5 the first time-resolved measurements of singlet oxygen luminescence in vivo,6 and the observation of oxygen quenching of triplet-excited single molecules.7 Experimental and theoretical studies on the mechanisms of photosensitized formation of excited O2 states and of their deactivation have been performed for almost 40 years. While most early liquid-phase studies were exclusively concerned with O2(∆g), recent technological advances also made possible time-resolved investigations of the second excited state, O2(Σg), which can be formed in competition with O2(∆g) in many cases. A significant number of * Corresponding author. Tel.: ++49 69 79829448. Fax: ++49 69 79829445. E-mail: R.Schmidt@chemie.uni-frankfurt.de. 1685 Chem. Rev. 2003, 103, 1685−1757

Physical mechanisms of generation and deactivation of singlet oxygen.

CHAPTER 3 – Photosensitized Oxidation and Singlet Oxygen: Consequences in Biological Systems

https://science.sciencemag.org/content/sci/162/3857/963.full.pdf

Mechanisms of Photosensitized Oxidation

Photosensitized oxygenations and the role of singlet oxygen

The quenching of triplet‐state molecules by molecular oxygen is examined theoretically. Two different quenching processes are considered: namely, (i) quenching by transfer of electronic excitation energy from the triplet state of the donor molecule to oxygen, resulting in a ground‐state donor molecule and an electronically excited singlet‐state oxygen molecule; and (ii) quenching by enhancement of intersystem crossing in the donor molecule, resulting in a vibrationally excited donor molecule in its ground electronic state and a ground‐state molecule. The relative importance of these two quenching processes is determined almost entirely by Franck—Condon factors, rather than by electronic factors. From our analysis we conclude that quenching by transfer of electronic excitation energy from the triplet‐state molecule to oxygen is usually 100–1000 times faster than quenching by enhanced intersystem crossing.Exchange interactions appear to be less important in promoting the radiationless transitions than do in...

Role of Singlet Excited States of Molecular Oxygen in the Quenching of Organic Triplet States

In this paper we have examined the fluorescence emission spectra, intensity, and decay characteristics and the excitation spectra of pure pyrene crystals and of perylene‐doped pyrene crystals at temperatures down to 1.7°K. From these observations we conclude that energy transfer in pyrene crystals occurs by four different mechanisms with the following characteristics: (i) Reabsorption of emitted photons: important at high guest concentrations or with thick samples. (ii) Monomer exciton migration: Operates at all temperatures with a transfer rate constant on the order of ∼1012 sec−1. (iii) Excimer exciton migration: important only at temperatures above about 130°K, thermally activated with an apparent activation energy of 112 cm−1 and a transfer rate constant of 4×106 sec−1 at room temperature. (iv) Forster transfer from excimers to perylene guest molecules: important only with perylene concentrations greater than about 10−3 mole/mole, or at slightly lower concentrations at reduced temperatures. In additio...

Investigation of Energy‐Transfer Mechanisms in Pyrene Crystals

A theoretical investigation of the excited electronic states of the equilateral triangular H3+ ion is reported. Ab initio calculations were performed for the lowest energy states of symmetries 3A1′, 1A2″, 3A2″, 1E′, and 3E′, using the single‐center expansion configuration‐interaction method. For each state we have obtained the potential‐energy curve in D3h symmetry. The calculated state energies are expected to be in error by at most 0.01 hartree (6 kcal/mole). Two of the excited states 1E′ and 3E′ were found to be purely repulsive with respect to the totally symmetric nuclear coordinate, while three excited states 1A2″, 3A2″, and 3A1′ showed minima with respect to the totally symmetric nuclear coordinate. However, arguments based on group‐theoretical correlation between our calculated D3h state energies and the energies of possible dissociation products suggest that none of the excited states considered achieves an absolute energy minimum in D3h symmetry. For the allowed electric dipole transitions 1A2″ ...

Single‐Center Calculations on the Electronically Excited States of Equilateral H3+ Ion

Single‐center wavefunctions, employing up to 100 configuration‐interaction terms, are used to compute electric, magnetic, and spectral properties of the 1A1′ electronic ground state of equilateral triangular H3+. The best values for several computed properties are: electronic energy, E=−1.3405 hartree; equilibrium internuclear distance, R=1.65 bohr; electric quadrupole moment, Θzz= −1.2540 B; and Larmor contribution to the mean diamagnetic susceptibility, χ¯L=−3.2693×10−6 cgs·emu. Expectation values for the squared electronic coordinate operators 〈 x12〉, 〈 y12〉, 〈 z12〉, and 〈 r12〉 are also given. The derivative of the electric quadrupole moment with respect to symmetric motion of the nuclei was calculated to be ∂ Θzz/∂ R=−2.5531 B/A, while the derivative of the electric dipole moment (center‐of‐mass origin) with respect to molecular bending was calculated to be ∂ μx/∂ φ=−0.0193 D/deg. These latter results permit prediction of the strengths of the quadrupole‐allowed A1′ and dipole‐allowed E′ vibrational fu...

Electric, Magnetic, and Spectral Properties of H3+ Ground State Calculated from Single‐Center Wavefunctions

The ``anomalous'' emission properties of perylene—pyrene mixed crystals have been re‐examined. The results of this study demonstrate that the unusual character of the mixed‐crystal emission spectra is primarily due to reabsorption of ``normal'' monomeric perylene emission, rather than to a very large excited‐state perylene—pyrene interaction as previously suggested. Studies of the emission and absorption properties of perylene in liquid solutions containing pyrene indicate, however, that there is a significant excited‐state perylene—pyrene interaction. This excited‐state interaction shifts the perylene emission over 150 A to the red, reduces the intensity, and produces a loss of some of the vibronic structure of the perylene emission. Because the lowest excited singlet states of perylene and pyrene are far from degenerate in energy, the excited‐state perylene—pyrene interaction cannot be due to a resonance interaction. Preliminary experimental results suggest that the excited‐state interaction is of charge‐transfer character.

Kenji Kawaoka

Papers

Role of Singlet Excited States of Molecular Oxygen in the Quenching of Organic Triplet States

Investigation of Energy‐Transfer Mechanisms in Pyrene Crystals

Single‐Center Calculations on the Electronically Excited States of Equilateral H3+ Ion

Electric, Magnetic, and Spectral Properties of H3+ Ground State Calculated from Single‐Center Wavefunctions

Excited‐State Perylene—Pyrene Interactions: Relation to the Interpretation of Excimer Emission