Anthony W. Parker

Toward control of electron transfer in donor-acceptor molecules by bond-specific infrared excitation

Electron transfer (ET) from donor to acceptor is often mediated by nuclear-electronic (vibronic) interactions in molecular bridges. Using an ultrafast electronic-vibrational-vibrational pulse-sequence, we demonstrate how the outcome of light-induced ET can be radically altered by mode-specific infrared (IR) excitation of vibrations that are coupled to the ET pathway. Picosecond narrow-band IR excitation of high-frequency bridge vibrations in an electronically excited covalent trans-acetylide platinum(II) donor-bridge-acceptor system in solution alters both the dynamics and the yields of competing ET pathways, completely switching a charge separation pathway off. These results offer a step toward quantum control of chemical reactivity by IR excitation. Vibrational excitation can modulate electron transfer probabilities in real time. Big impact from a well-placed shake Since the advent of ultrashort laser pulses, chemists have sought to steer reaction trajectories in real time by setting particular molecular vibrations in motion. Using this approach, Delor et al. have demonstrated a markedly clear-cut influence on electron transfer probabilities along the axis of a platinum complex. The complex comprised donor and acceptor fragments—which respectively give and take electrons upon ultraviolet excitation—bridged together by triply bonded carbon chains linked to the metal center. By selectively stimulating the carbon triple-bond stretch vibration with an infrared pulse, the authors could induce substantial changes in the observed electron transfer pathways between the fragments. Science, this issue p. 1492

Vibrationally quantum-state-specific reaction dynamics of H atom abstraction by CN radical in solution

Molecular vibrations in a solution-phase reaction are detected at a level of detail rivaling that of gas-phase studies. Solvent collisions can often mask initial disposition of energy to the products of solution-phase chemical reactions. Here, we show with transient infrared absorption spectra obtained with picosecond time resolution that the nascent HCN products of reaction of CN radicals with cyclohexane in chlorinated organic solvents exhibit preferential excitation of one quantum of the C-H stretching mode and up to two quanta of the bending mode. On time scales of approximately 100 to 300 picoseconds, the HCN products undergo relaxation to the vibrational ground state by coupling to the solvent bath. Comparison with reactions of CN radicals with alkanes in the gas phase, known to produce HCN with greater C-H stretch and bending mode excitation (up to two and approximately six quanta, respectively), indicates partial damping of the nascent product vibrational motion by the solvent. The transient infrared spectra therefore probe solvent-induced modifications to the reaction free energy surface and chemical dynamics.

Picosecond Transient Infrared Study of the Ultrafast Deactivation Processes of Electronically Excited B-DNA and Z-DNA Forms of [poly(dG-dC)]2†

Unravelling the ultrafast processes within DNA is a challenge that continues to exploit the boundaries of both spectroscopic and computational capabilities. However, such studies are essential for the understanding of the nature and dynamics of the UV-activated processes that precede DNA damage and lead to mutagenesis and ultimately cause a number of diseases. Considerable effort has focused on the photophysics and photochemistry of the individual base components which have very short electronically excited singlet state lifetimes (< 1 ps). In contrast, UV excitation of the polynucleotide systems produces additional species that have much longer lifetimes, 4] which is currently a main topic of scientific interest and debate. Questions remain as to whether the electronic excited states of these polymeric forms of DNA are localized on a single base or delocalized over a number of bases. Furthermore, the structural features of polymeric DNA raise a number of additional questions, such as their influence on the excited-state properties and relaxation dynamics of base-stacking interactions, hydrogen bonding, hydration, and conformation. The advantages of picosecond time-resolved infrared spectroscopy (ps-TRIR) are that it can yield structural details about transient species and also that it allows the ground-state depletion to be directly monitored. DNA is only weakly emissive and excited-state decay occurs predominantly through nonradiative channels, thus making ps-TRIR an ideal tool for DNA investigations. ps-TRIR also provides information about such dark processes, which are difficult to observe by the traditional transient absorption and not detectable by transient fluorescence techniques. For these reasons, ps-TRIR has recently been used to study the photodynamics and photoreactions of mononucleotides and polynucleotide DNA. By using ps-TRIR, we have identified a number of processes following direct excitation at 267 nm. For example, the cooling of the vibrationally hot ground state (2–4 ps) of mononucleotides was observed. Direct spectroscopic evidence of a longer-lived state (34 ps) in 5’-dCMP was assigned, partially on the basis of the IR band position, to the nNp* dark state. [5] The participation of this latter state in the relaxation dynamics was predicted by computational work and inferred from transient absorption work by Hare et al. We have also studied G-rich polynucleotides that are known to participate in tetrad stacking formation. In addition, the photoionization of the B form of [poly(dG-dC)]2 by direct excitation at 200 nm has been investigated, and resulted in a structural fingerprint to identify DNA damage. The relaxation processes and electron transfer from DNA to an intercalated metal complex excited at 400 nm has also been reported. Herein, we focus on the ultrafast dynamics of doublestranded poly(dG-dC). This is interesting for a number of reasons. Firstly, the steady-state absorption of the G and C bases in the polymeric form indicates the presence of electronic interactions between the bases, which in turn alter the excited state dynamics from that of the parent bases. It was recently shown that the excited states of the bases in [poly(dG-dC)]2 decay at rates faster than those observed for the individual nucleotides, with a particular role ascribed to charge-transfer states, a process that is predicted to be followed by rapid proton transfer between the G and C units. Secondly, if, as predicted, proton transfer acts to modulate the decay of locally excited (LE) states, then the characteristic IR signatures of the deprotonated guanine and protonated cytosine products should be detectable by psTRIR. Thirdly, [poly(dG-dC)]2 can adopt the unusual lefthanded Z-DNA structure. Thus a comparison of how the base stacking arrangements in the structurally distinct righthanded Band left-handed Z-DNA forms influences the photophysical processes of the G–C base pairs is possible. Finally, and in particular, the recent characterization of a nNp* dark state for 5’-dCMP, with a strong IR identifiable transient band at 1574 cm 1 poses questions as to the possible role of this state in C-containing polynucleotide chemistry. The transient IR spectra for B-form double-stranded [poly(dG-dC)]2 show strong bleaching and weaker transient features (Figure 1a). The ground-state IR spectra of the individual bases can be considered to have three regions: the carbonyl stretching region (1640–1700 cm ), where both C and G bases absorb, the G ring region (1550–1600 cm ) and [*] G. W. Doorley, D. A. McGovern, Prof. J. M. Kelly, Dr. S. J. Quinn School of Chemistry, Centre for Synthesis and Chemical Biology Trinity College Dublin, Dublin 2 (Ireland) E-mail: quinnsu@tcd.ie

Time-Resolved Resonance Raman Spectroscopy: Exploring Reactive Intermediates

The study of reaction mechanisms involves systematic investigations of the correlation between structure, reactivity, and time. The challenge is to be able to observe the chemical changes undergone by reactants as they change into products via one or several intermediates such as electronic excited states (singlet and triplet), radicals, radical ions, carbocations, carbanions, carbenes, nitrenes, nitrinium ions, etc. The vast array of intermediates and timescales means there is no single “do-it-all” technique. The simultaneous advances in contemporary time-resolved Raman spectroscopic techniques and computational methods have done much towards visualizing molecular fingerprint snapshots of the reactive intermediates in the microsecond to femtosecond time domain. Raman spectroscopy and its sensitive counterpart resonance Raman spectroscopy have been well proven as means for determining molecular structure, chemical bonding, reactivity, and dynamics of short-lived intermediates in solution phase and are advantageous in comparison to commonly used time-resolved absorption and emission spectroscopy. Today time-resolved Raman spectroscopy is a mature technique; its development owes much to the advent of pulsed tunable lasers, highly efficient spectrometers, and high speed, highly sensitive multichannel detectors able to collect a complete spectrum. This review article will provide a brief chronological development of the experimental setup and demonstrate how experimentalists have conquered numerous challenges to obtain background-free (removing fluorescence), intense, and highly spectrally resolved Raman spectra in the nanosecond to microsecond (ns–μs) and picosecond (ps) time domains and, perhaps surprisingly, laid the foundations for new techniques such as spatially offset Raman spectroscopy.

Optically trapped probes with nanometer-scale tips for femto-Newton force measurement

We describe the development of a novel force probe, controlled by multiple optical traps, with a nanometer-scale tip that protrudes outside the direct laser radiation field. We have measured forces to an accuracy of 240?fN, which enables future experiments that probe photo-sensitive components (such as biological cells) and non-transparent objects. The probes were produced using two methods, electron beam lithography and two-photon polymerization, with the latter providing approximately twice as much trapping stiffness.

Induction of persistent double strand breaks following multiphoton irradiation of cycling and G1-arrested mammalian cells-replication-induced double strand breaks.

DNA double strand breaks (DSBs) are amongst the most deleterious lesions induced within the cell following exposure to ionizing radiation. Mammalian cells repair these breaks predominantly via the nonhomologous end joining pathway which is active throughout the cell cycle and is error prone. The alternative pathway for repair of DSBs is homologous recombination (HR) which is error free and active during S‐ and G2/M‐phases of the cell cycle. We have utilized near‐infrared laser radiation to induce DNA damage in individual mammalian cells through multiphoton excitation processes to investigate the dynamics of single cell DNA damage processing. We have used immunofluorescent imaging of γ‐H2AX (a marker for DSBs) in mammalian cells and investigated the colocalization of this protein with ATM, p53 binding protein 1 and RAD51, an integral protein of the HR DNA repair pathway. We have observed persistent DSBs at later times postlaser irradiation which are indicative of DSBs arising at replication, presumably from UV photoproducts or clustered damage containing single strand breaks. Cell cycle studies have shown that in G1 cells, a significant fraction of multiphoton laser‐induced prompt DSBs persists for >4u2003h in addition to those induced at replication.

The role of CN and CO ligands in the vibrational relaxation dynamics of model compounds of the [FeFe]-hydrogenase enzyme

The vibrational dynamics of (μ-propanedithiolate)Fe(2)(CO)(4)(CN)(2)(2-), a model compound of the active site of the [FeFe]-hydrogenase enzyme, have been examined via ultrafast 2D-IR spectroscopy. The results indicate that the vibrational coupling between the stretching modes of the CO and CN ligands is small and restricted to certain modes but the slow growth of off-diagonal peaks is assigned to population transfer processes occurring between these modes on timescales of 30-40 ps. Analysis of the dynamics in concert with anharmonic density functional theory simulations shows that the presence of CN ligands alters the vibrational relaxation dynamics of the CO modes in comparison to all-carbonyl model systems and suggests that the presence of these ligands in the enzyme may be an important feature in terms of directing the vibrational relaxation mechanism.

A 100 kHz Time-Resolved Multiple-Probe Femtosecond to Second Infrared Absorption Spectrometer

We present a dual-amplifier laser system for time-resolved multiple-probe infrared (IR) spectroscopy based on the ytterbium potassium gadolinium tungstate (Yb:KGW) laser medium. Comparisons are made between the ytterbium-based technology and titanium sapphire laser systems for time-resolved IR spectroscopy measurements. The 100u2009kHz probing system provides new capability in time-resolved multiple-probe experiments, as more information is obtained from samples in a single experiment through multiple-probing. This method uses the high repetition-rate probe pulses to repeatedly measure spectra at 10u2009µs intervals following excitation allowing extended timescales to be measured routinely along with ultrafast data. Results are presented showing the measurement of molecular dynamics over >10 orders of magnitude in timescale, out to 20u2009ms, with an experimental time response of <200u2009fs. The power of multiple-probing is explored through principal component analysis of repeating probe measurements as a novel method for removing noise and measurement artifacts.

Dynamic position and force measurement for multiple optically trapped particles using a high-speed active pixel sensor

A high frame rate active pixel sensor designed to track the position of up to six optically trapped objects simultaneously within the field of view of a microscope is described. The sensor comprises 520 x 520 pixels from which a flexible arrangement of six independent regions of interest is accessed at a rate of up to 20 kHz, providing the capability to measure motion in multiple micron scale objects to nanometer accuracy. The combined control of both the sensor and optical traps is performed using unique, dedicated electronics (a field programmable gate array). The ability of the sensor to measure the dynamic position and the forces between six optically trapped spheres, down to femtonewton level, is demonstrated paving the way for application in the physical and life sciences.

Two-photon excitation with pico-second fluorescence lifetime imaging to detect nuclear association of flavanols

Two-photon excitation enabled for the first time the observation and measurement of excited state fluorescence lifetimes from three flavanols in solution, which were ~1.0 ns for catechin and epicatechin, but <45 ps for epigallocatechin gallate (EGCG). The shorter lifetime for EGCG is in line with a lower fluorescence quantum yield of 0.003 compared to catechin (0.015) and epicatechin (0.018). In vivo experiments with onion cells demonstrated that tryptophan and quercetin, which tend to be major contributors of background fluorescence in plant cells, have sufficiently low cross sections for two-photon excitation at 630 nm and therefore do not interfere with detection of externally added or endogenous flavanols in Allium cepa or Taxus baccata cells. Applying two-photon excitation to flavanols enabled 3-D fluorescence lifetime imaging microscopy and showed that added EGCG penetrated the whole nucleus of onion cells. Interestingly, EGCG and catechin showed different lifetime behaviour when bound to the nucleus: EGCG lifetime increased from <45 to 200 ps, whilst catechin lifetime decreased from 1.0 ns to 500 ps. Semi-quantitative measurements revealed that the relative ratios of EGCG concentrations in nucleoli associated vesicles: nucleus: cytoplasm were ca. 100:10:1. Solution experiments with catechin, epicatechin and histone proteins provided preliminary evidence, via the appearance of a second lifetime (τ(2)=1.9-3.1 ns), that both flavanols may be interacting with histone proteins. We conclude that there is significant nuclear absorption of flavanols. This advanced imaging using two-photon excitation and biophysical techniques described here will prove valuable for probing the intracellular trafficking and functions of flavanols, such as EGCG, which is the major flavanol of green tea.