The scope of this review is the nature and dynamics of the singlet excited electronic states created in nucleic acids and their constituents by UV light. Interest in the UV photochemistry of nucleic acids has long been the motivation for photophysical studies of the excited states, because these states are at the beginning of the complex chain of events that culminates in photodamage. UV-induced damage to DNA has profound biological consequences, including photocarcinogenesis, a growing human health problem.1-3 Sunlight, which is essential for life on earth, contains significant amounts of harmful UV (λ < 400 nm) radiation. These solar UV photons constitute one of the most ubiquitous and potent environmental carcinogens. This extraterrestrial threat is impressive for its long history; photodamage is as old as life itself. The genomic information encoded by these biopolymers has been under photochemical attack for billions of years. It is not surprising then that the excited states of the nucleic acid bases (see Chart 1), the most important UV chromophores of nucleic acids, are highly stable to photochemical decay, perhaps as a result of selection pressure during a long period of molecular evolution. This photostability is due to remarkably rapid decay pathways for electronic energy, which are only now coming into focus through femtosecond laser spectroscopy. The recently completed map of the human genome and the ever-expanding crystallographic database of nucleic acid structures are two examples that illustrate the richly detailed information currently available about the static properties of nucleic acids. In contrast, much less is known about the dynamics of these macromolecules. This is particularly true of the dynamics of the excited states that play a critical role in DNA photodamage. Efforts to study nucleic acids by time-resolved spectroscopy have been stymied by the apparent lack of suitable fluorophores. In contrast, dynamical spectroscopy of proteins has flourished thanks to intrinsically fluorescent amino acids such as tryptophan, tyrosine, and phenylalanine.4 The primary UVabsorbing constituents of nucleic acids, the nucleic acid bases, have vanishingly small fluorescence quantum yields under physiological conditions of temperature and pH.5 In fact, the bases were frequently described as “nonfluorescent” in the early literature. * To whom correspondence should be addressed. E-mail: kohler@ chemistry.ohio-state.edu. Phone: (614) 688-3944. Fax: (614) 2921685. 1977 Chem. Rev. 2004, 104, 1977−2019

Ultrafast Excited-State Dynamics in Nucleic Acids

The combined results of ab initio electronic-structure calculations and spectroscopic investigations of jet-cooled molecules and clusters provide strong evidence of a surprisingly simple and general mechanistic picture of the nonradiative decay of biomolecules such as nucleic bases and aromatic amino acids. The key role in this picture is played by excited singlet states of πσ* character, which have repulsive potential-energy functions with respect to the stretching of OH or NH bonds. The 1πσ* potential-energy functions intersect not only the bound potential-energy functions of the 1ππ* excited states, but also that of the electronic ground state. Via predissociation of the 1ππ* states and a conical intersection with the ground state, the 1πσ* states trigger an ultrafast internal-conversion process, which is essential for the photostability of biomolecules.
In protic solvents, the 1πσ* states promote a hydrogen-transfer process from the chromophore to the solvent. Calculations for chromophore–water clusters have shown that a spontaneous charge-separation process takes place in the solvent shell, yielding a microsolvated hydronium cation and a microsolvated electron. These results suggest that the basic mechanisms of the complex photochemistry of biomolecules in liquid water can be revealed by experimental and theoretical investigations of relatively small chromophore–water clusters.

Excited-state hydrogen detachment and hydrogen transfer driven by repulsive 1πσ* states: A new paradigm for nonradiative decay in aromatic biomolecules

Radiationless deactivation pathways of excited gas phase nucleobases were investigated using mass-selected femtosecond resolved pump-probe resonant ionization. By comparison between nucleobases and methylated species, in which tautomerism cannot occur, we can access intrinsic mechanisms at a time resolution never reported so far (80 fs). At this time resolution, and using appropriate substitution, real nuclear motion corresponding to active vibrational modes along deactivation coordinates can actually be probed. We provide evidence for the existence of a two-step decay mechanism, following a 267 nm excitation of the nucleobases. The time resolution achieved together with a careful zero time-delay calibration between lasers allow us to show that the first step does correspond to intrinsic dynamics rather than to a laser cross correlation. For adenine and 9-methyladenine a first decay component of about 100 fs has been measured. This first step is radically increased to 200 fs when the amino group hydrogen atoms of adenine are substituted by methyl groups. Our results could be rationalized according to the effect of the highly localized nature of the excitation combined to the presence of efficient deactivation pathway along both pyrimidine ring and amino group out-of-plane vibrational modes. These nuclear motions play a key role in the vibronic coupling between the initially excited pipi(*) and the dark npi(*) states. This seems to be the common mechanism that opens up the earlier phase of the internal conversion pathway which then, in consideration of the rather fast relaxation times observed, would probably proceed via conical intersection between the npi(*) relay state and high vibrational levels of the ground state.

Excited states dynamics of DNA and RNA bases: characterization of a stepwise deactivation pathway in the gas phase.

Gas-phase spectroscopy lends itself ideally to the study of isolated molecules and provides important data for comparison with theory. In recent years, we have seen enormous progress in the study of biomolecular building blocks in the gas phase. The motivation for such work is threefold: (a) It is important to distinguish between intrinsic molecular properties and properties that result from the biological environment. (b) Gas-phase spectroscopy of clusters provides insights into fundamental interactions and into microsolvation. (c) Gas-phase data support quantum-chemical calculations. This review focuses on the current status of (poly)amino acids and DNA bases. Recent results help elucidate structure and hydrogen-bonded interactions, as well as showcase a successful interplay between theory and experiment.

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Gas-phase spectroscopy of biomolecular building blocks.

The photophysics and photochemistry of DNA is of great importance due to the potential damage of the genetic code by UV light. Quantum mechanical studies have played a key role in interpretating the results of modern time-resolved pump-probe spectroscopy, and in elucidating the main photoactivated reactive paths. This review provides a concise, complete picture of the computational studies carried out, approximately, in the past decade. We start with an overview of the photophysics of the nucleobases in the gas phase and in solution. We discuss the proposed mechanisms for ultrafast decay to the ground state, that involve conical intersections, consider the role of triplet states, and analyze how the solvent modulates the photophysics. Then we move to larger systems, from dinucleotides to single- and double-stranded oligonucleotides. We focus on the possible role of charge transfer and delocalized or excitonic states in the photophysics of these systems and discuss the main photochemical paths. We finish with an outlook on the current challenges in the field and future directions of research.

Quantum Mechanical Studies on the Photophysics and the Photochemistry of Nucleic Acids and Nucleobases

Electronic spectra of the jet-cooled DNA base adenine were obtained by the resonant two-photon ionization (R2PI) and the laser induced fluorescence (LIF) techniques. The 0–0 band to the lowest electronically excited state was found to be located at 35 503 cm−1. Well-resolved vibronic structures were observed up to 1100 cm−1 above the 0–0 level, followed by a slow rise of broad structureless absorption. The lowest electronic state was proposed to be of nπ* character, which lies ∼600 cm−1 below the onset of the ππ* state. The broad absorption was attributed to the extensive vibronic mixing between the nπ* state and the high-lying ππ* state.

Resonant two-photon ionization and laser induced fluorescence spectroscopy of jet-cooled adenine

L-phenylalanine (Phe), one of the aromatic amino acids, and its hydrated clusters were generated in supersonic expansion and investigated by resonant two-photon ionization. Excitation spectra of Phe and Phe–(H2O)1 were obtained near their S0–S1 origins. We found that, by comparing the experimental results with the density functional theory and ab initio calculations, the water in Phe–(H2O)1 tends to form a cyclic hydrogen bond at the carboxyl group while inducing little change in the corresponding monomer structure. No sign of water making bridged hydrogen bonds with both polar groups was found. In order to form the cyclic hydrogen bond, hydration takes place only with the conformers whose carboxyl hydrogen is free, i.e., not occupied in the intramolecular hydrogen bonding with the amino nitrogen in the monomer.

Resonant two-photon ionization study of jet-cooled amino acid: L-phenylalanine and its monohydrated complex

Conformation-dependent ionization energies of L-phenylalanine.

Conformation-dependent ionization of l-phenylalanine: structures and energetics of cationic conformers

Toxic chemicals inside building materials have long-term harmful effects on human bodies. To prevent secondary damage caused by the evaporation of latent chemicals, it is necessary to detect the chemicals inside building materials at an early stage. Deep Raman spectroscopy is a potential candidate for on-site detection because it can provide molecular information about subsurface components. However, it is very difficult to spectrally distinguish the Raman signal of the internal chemicals from the background signal of the surrounding materials and to acquire the geometric information of chemicals. In this study, we developed hyperspectral wide-depth spatially offset Raman spectroscopy coupled with a data processing algorithm to identify toxic chemicals, such as chemical warfare agent (CWA) simulants in building materials. Furthermore, the spatial distribution of the chemicals and the thickness of the building material were also measured from one-dimensional (1D) spectral variation.

Jiha Sung

Papers

Resonant two-photon ionization and laser induced fluorescence spectroscopy of jet-cooled adenine

Resonant two-photon ionization study of jet-cooled amino acid: L-phenylalanine and its monohydrated complex

Conformation-dependent ionization energies of L-phenylalanine.

Conformation-dependent ionization of l-phenylalanine: structures and energetics of cationic conformers

Hyperspectral depth-profiling with deep Raman spectroscopy for detecting chemicals in building materials