Raman and resonance Raman spectroscopies are employed to explore the degree of solvent-induced symmetry breaking int he ground and excited electronic states of the nominally symmetric linear molecules CS₂ and I₃-. A signature of broken symmetry is intensity in the antisymmetric stretching fundamental. CS₂ breaks symmetry only slightly in all solvents examined, and the effects on the nuclear dynamics of the S₃ excited state appear to be minimal. In I₃-, asymmetry is pronounced in alcoholic solvents but undetectable in acetonitrile. MCSCF/RISM calculations support the experimental result and indicate that solvent stabilization of separated charges renders a wide range of asymmetric ground-state structures energetically accessible. Recent femtosecond time-domain studies of the photodissociated process indicate a significant influence of asymmetry of the reactant on the dynamics of product formation.

In this paper, we describe a new approach to simulating many-body molecular dynamics on coupled electronic surfaces. The method is based on a semiclassical limit of the quantum Louisville equation, which yields equations of motion for classical-like distribution functions describing both nuclear probability densities on the coupled surfaces and the coherences between the electronic states. The Hamiltonian dynamics underlying the evolution of these distributions is augmented by nonclassical source and sink terms, which allow the flow of probability between the coupled surfaces and the corresponding formation and decay of electronic coherences. We show that this approach reproduces the familiar Landau-Zehnder transition probability in the limit of weak electronic coupling. In addition, we describe a trajectory-based implementation in the context of a conventional molecular dynamics simulation.

The electron transfer dynamics of 9-anthracene carboxylic acid bound to TiO₂ nanoparticles in ethanol has been examined by a combination of transient absorption and time- resolved anistropy measurements. The results from these experiments show that the forward electron transfer reaction is very fast, <EQ 360 fs for anatase TiO₂ and <EQ 1 ps for rutile TiO₂. The back electron transfer reaction occurs on a slower time scale: 33 +/- 1 ps for the anatase particles and 54 +/- 1 ps for the rutile particles. The rate of the back electron transfer reaction for anatase TiO₂ is also very sensitive to the addition of small amounts of water to the ethanolic solutions. For example, adding 1 percent water by volume decreases the average back electron transfer time from 33 ps to 20 ps. The water also produces a red-shift in the absorption spectra of the TiO₂ particles. These observations show that the back electron transfer reaction is in the Marcus inverted region.

Ultrafast IR vibrational echo experiments, which are used to examine liquids, glasses, and proteins are described. Like the NMR spin echo and other NMR pulse sequences, the vibrational echo can extract dynamical and spectroscopic information that cannot be obtained from a vibrational absorption spectrum. The vibrational echo measures the homogeneous vibrational linewidth even if the absorption line is massively inhomogeneously broadened. When combined with pump-probe experiments, the homogeneous pure dephasing is obtained. Conducting these experiments as a function of temperature provides information in dynamics and intermolecular interactions. The nature of the method and the experimental procedures are outlined. Experimental results are presented for the metal carbonyl solute, Rh(CO)₂acac, in liquid and glassy dibutyl phthalate. The dynamics of the CO ligand bound at the active site of the protein myoglobin is also examined and compared with that in several myoglobin mutants. The results provide insights into protein dynamics and how protein structural fluctuations are communicated to a ligand bound at the active site.

A novel optimized heterodyne method which recovers the complete electric field of any four-wave mixing signal at its point of origin is demonstrated. A tracer pulse is sent along the signal path and characterized at the sample by frequency-resolved optical gating. Spectral interferometry is used to determine the phase difference between the tracer and a reference pulse, the absorptive change in tracer phase in the sample, and the reference-signal phase difference. Together, these measurements allow calculation of the signal phase at the sample. The phase of three pulse scattering signals from solutions of the dye IR144 in methanol determines the absolute signal emission time within 0.5 fs.

Membrane-bound receptor proteins regulate the transmission of signals at the junctions between cells of the mammalian nervous system. Upon binding a specific neurotransmitter these receptor proteins form transient channels through which inorganic ions flow, leading to a change in the transmembrane voltage of a cell. A newly developed laser- pulse photolysis technique, with a microsecond(s) time resolution, allows one to determine the rate constants for both the formation and closing of the transmembrane channel, the dissociation constant for the ligand-binding site that controls channel opening, and the concentration of the receptor in the cell membrane, and gives information about the rate of transient inactivation of the receptor. The technique allows one to determine the binding constants of inhibitors to the closed- and open-channel forms independently. THe use of the chemical kinetic method is illustrated.

The dynamic response of a protein to photoinitiation of folding can reveal new information about the early stages of secondary and tertiary structure formation, information beyond that obtainable from x-ray crystallography, NMR, and other steady state methods. The combined efforts of time- resolved studies using different experimental probes have already observed several events in the early stages of protein and peptide folding. However, in terms of the 'big folding picture', our understanding of protein folding is limited. Information on the folding mechanisms for a range of peptides and proteins is important to address the commonality of folding intermediates and pathways, but it is also useful to thoroughly understand the process of folding in at least one protein. To obtain such an understanding of one protein it is useful to employ a number of spectroscopic approaches We present here the results of time-resolved absorption, CD and MCD studies of the folding reactions of reduced cytochrome c after ligand photolysis of the CO adduct. We also discuss our results and the results of other studies on cytochrome c to demonstrate the advantage of using a number of different approaches for understanding protein folding.

The technique of femtosecond coherence spectroscopy is applied to the heme proteins myoglobin (Mb) and hemoglobin (Hb). Studies of field driven coherences lead to power spectra that are in good agreement with resonance Raman scattering experiments. Studies of the NO bound derivatives of Mb and Hb reveal rapid photolysis and non-radiative relaxation to the ground electronic state. The ensuing nuclear response is oscillatory and displays strong coupling of the NO photolysis reaction to the iron-histidine vibration and to the heme doming mode, which we locate at approximately 40 cm^-1. The doming mode was previously assigned to a mode at approximately 80 cm^-1, which we now believe is actually the first overtone of the doming motion. Other modes at approximately 120 cm^-1 and 160 cm^-1 are also exposed by the new data, suggesting that a progression of doming harmonics is excited to the strong coupling of this mode to the photolysis reaction.

Linear and nonlinear spectroscopy of liquid water is studied with molecular dynamics computer simulation techniques. The electronic structure variation of each solvent molecule with its local environment is effected via a truncated adiabatic basis-set description. By the inclusion of both linear and nonlinear electronic response, this accounts for the instantaneous adjustment of the water dipole moment and polarizability to the fluctuating local electric field. It also allows for the electronic relaxation effects associated with excitations through a mixing of different excited electronic configurations. By employing the TAB/10D potential model developed recently in our group, the electronic absorption, far-IR, depolarized Raman scattering and optical Kerr effect spectroscopy of water are examined under ambient conditions.

The photolysis, geminate recombination and vibrational relaxation of the low affinity ferrous myoglobin nicotinate complex have been studied by femtosecond transient absorption spectroscopy. This is an interesting system due to the peculiar interaction between ligand and protein fluctuations. This ligand is bulky and affects the naturally occurring protein fluctuations in a way similar to a doorstop precluding a door from closing totally. The whole Q band absorption transient spectrum of the photoproduct has been monitored starting from 100 fs to 100 ps. The time evolution of the spectrum has clearly shown two distinct phases, a vibrational cooling process occurring within 4 ps after the photolyzing pulse and a geminate rebinding process with a time constant of 28.8 +/- 0.1 ps. The transient spectra show different cooling rates for the different excited normal modes. The geminate rebinding process appears to be complete within 100 ps and hence appears to be the fastest geminate recombination process reported to date for a hemoprotein. This is the first report on the photolysis of a ferrous heme adduct with a nitrogenous base, previously considered as photoinert.

The electronic characteristics and rapid dynamics associated with bacteriochlorophyll dimers in photosynthetic systems are investigated using novel ultrafast anisotropy techniques. The excitonic structure of isolated subunits (B820) from the core (LH-1) light harvesting complex of Rs. Rubrum and the reassociated complex (B873) is revealed in coherent anisotropy responses following impulsive excitation. For B820, the oscillatory anisotropy responses indicate excitonic splitting frequencies ranging from 370 cm^-1 to 490 cm^-1 indicating significant inhomogeneity in the excitonic spectrum. The complete set of wavelength dependent anisotropy result is analyzed to reveal the sources of inhomogeneity for B820; correlated distributions of dimer energetic parameters are necessary to reproduce the results. The coherent response of reassociated B873 complexes exhibit multiple frequencies, revealing the extended excitonic structure of the aggregates. In other experiments, the electronic properties of the photosynthetic reaction center and the rapid electronic dynamics prior to charge separation are investigated by both 'two color' anisotropy. A variety of excitation and detection conditions provide the first room temperature characterization of the excitonic structure of the special pair, and a detailed description of rapid energy transfer to and within the special pair. The results presented here stress the importance of delocalized excitation in both the light harvesting antenna complexes and the photosynthetic reaction center.

The general theoretical and experimental principles of optical pump-THz probe spectroscopy of chemical reactions in liquids is presented. Background on specific difficulties encountered in the experimental observation is reviewed. Chiefly, signal-to-noise ratios currently limit the quality of information that can be extracted from optical pump-THz prove data on chemical reactions. This issue is shown to be connected to the assumption of linear responses. The problem of interpretation of frequency dependent absorption coefficient data of neat liquids and solutions is addressed, and reasonably successful methods for analysis of the data are presented. Transient absorption coefficient spectra of GaAs illustrates the type of information this spectroscopic method can extract form condensed phase samples.

We present a method to determine system-bath correlation functions from third order optical coherence measurements. The importance of these correlation functions for understanding solvation dynamics is explained. A physical argument is made to explain why one coherence measurement, the photon echo peak shift, should strongly reflect system- bath dynamics. Finally, this method is applied to the system of bacteriochlorophyll in tetrahydrofuran solution.

The design and performance of a cavity-dumped Ti:sapphire oscillator and short pulse-pumped amplifier system is presented. Dumped pulses as short as 12 fs nJ in energy, with 70-80 percent dumping efficiency are made possible by a stretched dumper-arm configuration. The oscillator yields stable, reliable, and reproducible pulses. Pulse duration and phase structure are analyzed via wavelength-resolved nonresonant scattering. Single and multi-pass amplification schemes and performance results are described. The optical continuum generated using single-pass amplified pulses is characterized by frequency-resolved cross-correlations.

In situ surface plasmon resonance (SPR) spectroscopy methods are used to investigate the process by which thin films form at solid/solutions interfaces. Representative kinetics data are presented for the adsorption of two different types of molecular adsorbates onto gold substrates. The first system involves adsorption of alginic acid, an acidic polysaccharide. The second system involves the adsorption of single-stranded DNA oligomers. Quantitative two-color SPR kinetics data are of sufficiently high quality that distinguishing between various kinetics models is now possible. Commonly used Langmuir adsorption models cannot adequately describe some of the adsorption kinetics data. However, these data are very well described by a model developed to include diffusion at the interface as well as adsorption/description. This more general model is found to work very well for a variety of chemically distinct adsorbate/substrate systems. Analysis of high quality in- situ SPR data can identify distinct differences in kinetics and many be useful for distinguishing between different types of adsorbate/surface interactions. This capability may prove very useful for gaining insight into mechanistic differences in the process by which molecular films at solid/liquid interfaces.

The new in situ optical technique of electrochemically modulated surface plasmon resonance is described and applied to the measurement of the electrostatic fields inside noncentrosymmetric zirconium phosphate films. In situ EM-SPR measurements on noncentrosymmetric ZP films yield a value for the change in electric field strength of 4 X 10³ V/cm for a change in electrode potential of +/- 25 mV. This electric field strength value indicates that there is substantial ion penetration into the film in the electrochemical environment. Both the phase and magnitude of the surface optical response in the EM-SPR measurements are used to distinguish the molecular and metal electrode contributions to the overall optical signal. These two EM- SPR contributions are identified and separated in a quantitative fashion through a series of theoretical Fresnel calculations.

Both human lung surfactant protein SP-B and its amino terminus alter the phase behavior of palmitic acid (PA) monolayers by inhibiting the formation of condensed phases and creating a new fluid PA-protein phase. This phase increase the compressibility of the monolayers by forming a network that separates condensed phase domains at coexistence and persists to high surface pressures. The network changes the monolayer collapse nucleation from a heterogenous to a more homogenous process through isolating individual condensed phase domains. This results in higher surface pressures at collapse, and monolayers easier to respread on expansion, factors essential to the in vivo function of lung surfactant. The network is stabilized by low line tension between the coexisting phases are confirmed by the formation of extended linear domains or 'stripe' phases. Similar stripes are found in monolayers of fluorescein-labeled amino terminus, suggesting that the reduction n line tension is due to the protein. Comparison of isotherm data and observed morphologies of monolayers containing amino terminus with those containing amino terminus with those containing the full length SP-B protein shows that the peptide retains most of the native activity of the protein, which may lead to cheaper and more effective synthetic replacement formulations.

Work from our laboratory on vibrational sum frequency spectroscopic investigations of molecular ordering at the carbon tetrachloride-water interface is reviewed. Simple charged surfactants adsorbed at the liquid-liquid interface are seen to induce alignment of interfacial water molecules to a degree which is dependent on the induced surface potential. Saturation of water molecule alignment occurs at a surfactant surface concentration corresponding to a calculated surface potential of approximately 160 mV. In complementary studies, the relative degree of hydrocarbon chain ordering within monolayers of symmetric phosphatidylcholines of different chain lengths is inferred by the relative signal contributions of the methyl and methylene symmetric stretch modes. The degree of hydrocarbon chain disorder observed depends strongly on the method of monolayer preparation. By one method, a decrease in hydrocarbon chain order is seen with increasing chain length. Another method of monolayer formation yielded very well ordered hydrocarbon chains for the longest chain phosphatidylcholine studied, and showed much greater disorder in shorter chain species which was comparable to the other preparation method. These studies are a foundation for further work with this technique geared towards understanding molecular-level structural features in membrane-like assemblies and surface biochemical interactions of relevance to biomedical research.

Molecular dynamics computer simulations are used to study the electronic absorption line shapes of adsorbed chromophores at several liquid interfaces. Specifically considered are the liquid/vapor interface of water and the interface between water and a number of different organic liquids which are characterized by different dielectric constants, structure and polarizability. The chromophore used in the calculations is modeled after the common dye molecule DEPNA. Both non-polarizable and polarizable liquid and solute models are considered. The calculations demonstrate the effect of solvent polarity on the spectra, in agreement with recent experiments. These calculations demonstrate the effect of solvent polarity on the spectra, in agreement with recent experiments. These calculations also highlight the important effect due to the microscopic structure of the interface. A comparison of the results with predictions of continuum models is presented. Although these models can qualitatively account for the effect of interface polarity on the spectra, they must be extended to include structural aspects of the interface for better quantitative agreement.

The imaging characteristics of cantilevered NSOM probes operating in a tapping-mode feedback arrangement are discussed and compared to conventional tips employing the shear-force feedback method. Images form a wide range of samples are presented to demonstrate the surface tracking capabilities over both high and low topology samples, in addition to the low fluorescence detection limits possible utilizing the new tips. The results show that the cantilevered tip operating in a tapping-mode arrangement offers enhanced force imaging of the sample topology without compromising the low detection limits or high spatial resolution of the NSOM fluorescence images. The examples discussed here indicate that the new design will be particularly useful for applications involving biological samples that frequently exhibit complex surface topologies.

The development of fluorescence spectroscopy and imaging on the single molecule level has provided a new method to investigate various phenomena in condensed phase at liquid helium and room temperature unobscured by ensemble averaging. Single molecule spectroscopy allows studying the photophysical behavior of chromophores as well as using those chromophores as a probe for their local environment. A crucial problem that has to be overcome for single molecule studies in the liquid phase is Brownian motion: only partially or completely immobilized molecules allow extended observation. Here we report on the use of water-based polyacrylamide gels as a promising medium for single molecule investigations at room temperature with wide-field total internal reflection microscopy. The gel framework dramatically reduced Brownian motion of small fluorescent dye molecules. Observation of the diffusion of these molecules served as a probe for the inner structure of the gels. Furthermore these water-based gels form a useful medium for single molecule studies of biological systems in vitro.

While optical microscopy is an everyday tool in the biological sciences, significant advances could be made if optical resolution were increased. We have evaluated the utility of near-field optical microscopy, with its demonstrated sub-diffraction optical resolution, for fluorescence studies of living cells. We show that the incorporation of a fluorescent feedback method, photon- density feedback, permits near-field studies of living cells. With this approach it is possible to make local high- resolution point measurements from living cells without impacting the physiology of the system under study. By integrating near-field illumination fibers, with confocal detection, traditional cameras and PMT-based detection methods, we have developed a near-field microscope for biological studies which also permits traditional lens-based optical studies of biological system With this system we are able to make immediate comparisons between confocal and near-field data from the same biological system.

A new approach to near-field optical microscopy is presented. The method relies on the highly enhanced fields at sharp metal tips under proper laser illumination. These fields are laterally confined to the tip size and can be used to locally excite the sample surface. Detection of nonlinear responses ensure sufficient background discrimination. The strong field gradients close to the tip give also rise to a trapping force towards the tip. Therefore, the proposed scheme is also promising for optical trapping and alignment of dielectric particles in aqueous environments at the nanometer scale. The paper presents the result of self-consistent 3D field calculations. Starting with a discussion of commonly used aperture probes, the field distributions for the novel scheme are presented.

The STM-electroluminescence technique is shown to be a valuable tool for characterizing optoelectronic properties and understanding structure-function relationships in heterogenous or disordered material on nanometer length scales. The intensity of photon emission induced by tunneling electrons from rough Au films is found to depend on the surface feature size. This size-dependent photon emission yield is shown to agree with the theoretically predicted trend based on the inelastic electron tunneling mechanism. Correlated STM 'topography' and electroluminescence measurement of polypyridine (PPy) showed electroluminescence almost exclusively result from low conductivity regions of the film .This anomalous correlation between STM topography and photon emission maps of PPy films is interpreted as the consequence of the spatial variation of the carrier mobility. The results have important implications for understanding the underlying physics of electroluminescence of polymer films as well as for development of optoelectronic devices based on polymeric materials.

We summarize recent progress aimed at observing biochemical and biological dynamics using confocal microscopy with 3D spatial resolution down to a few hundred nm and temporal resolution to 15 fs. We also review recent control of population dynamics using tailored ultrafast pulses, i.e. quantum control. Progress is described for i) feedback control, ii) multiphoton control, and iii) molecular (pi) pulse. Finally, using ultrafast light pulses, we combine confocal and quantum control techniques to produce a new way to measure the microscopy chemical environment, int his case pH, potentially with a spatial resolution of a few hundred nanometers.

The dynamics induced in matter by a short optical pump pulse can be measured by time-resolved x-ray diffraction without resorting to additional and often unknown information as required in optical pump and probe experiments. Several theoretical aspects of such measurements are considered here: elastic versus inelastic scattering, quantum interference among electronic states, physical implications of temporal- and spatial-averaging, and the coherence of x- ray beams. Based on these considerations, it is possible to use inelastic scattering for studying curve crossing in molecular systems and electronic coherence in electronic materials, in addition to probing nuclear dynamics on an excited potential energy surface. With certain modifications, the time-dependent analysis presented here can be extended to other experimental methods including electron diffraction and x-ray absorption.

Nanosecond x-ray pulses have been used for time resolved x- ray diffraction to measure the transient structure of Pt and GaAs crystals caused by laser pulse heating. A direct imaging x-ray CCD system with high spatial resolution allows the detection for the lattice deformations of the order of 10^-4 A, induced by a laser pulse energy of a few millijoules.

The mixed-order semiclassical molecular dynamics method is used for the calculation of quantum time correlation functions in extended systems. The method allows the consistent treatment of a selected number of degrees of freedom to second-order in the stationary phase approximation through the Herman and Kluk propagator, while the rest of the system is treated to zeroth-order, using frozen Gaussians. The formulation is applied to calculate the absorption spectrum, of the B $IMP X transition of Cl₂ isolated in solid Ar, a spectrum that shows zero-phonon lines and phonon sidebands with relative intensities that depend on the excited state vibrational level. The explicit simulation of quantum time correlation functions of the system consisting of 321 degrees of freedom, reproduces the spectrum and allows its interpretation in terms of the underlying molecular motions.IN order to extend the semiclassical methods to longer timescale a new extension of Herman-Kluk propagator is developed, which combines classical propagation of trajectories for length where the initial value propagator remains accurate, followed by Monte-Carlo regeneration of the ensemble of trajectories and continuation of propagation. This new method is tested for the calculation of long time dynamics in a 1D Morse oscillator.

Femtosecond laser pulses focused inside liquid helium initiate an excitation sequence that leads to formation of molecular Rydberg states He₂ detected by fluorescence spectroscopy. Unlike in the case of longer laser pulses, the excitations may be created in a controllable way, at light intensities below dielectric breakdown. The initial step is ionization of He atoms, as demonstrate by charge separation in external electric field. A sequence of the subsequent processes is proposed, which accounts for rapid production of He₂, in less than 10 ns, observed by nanosecond time- resolved laser induced fluorescence following the excitation pulse. the lowest triplet state excimers He₂(³a), probed in the latter experiment, are long-lived and survive in concentrations of the order of 10¹¹-10¹² cm^-3 on a millisecond time scale. Femtosecond time- resolved spectroscopy was performed on He₂* molecules in liquid He, using the pump-probe sequence He₂*(³a) + 790 nm yields He₂*(³c), He₂*(³c) + 790 nm yields He₂*(³f). The observed decay of the transient signals with characteristic time 3.5 ps is thought to be due to solvent motion corresponding to the relaxation of the liquid helium 'bubble' around the intermediate He₂*(³c) state.

The advances in room temperature single molecule detection (SMD) and single molecule spectroscopy (SMS) by laser induced fluorescence provide new tools for testing semi- classical and quantum mechanical theories of light-matter interaction and molecular interactions. They also provide the tools for the study of individual synthetic and biological macromolecules under physiological conditions. Two properties of a single fluorescent probe attached to a macromolecule can be exploited to provide local structural information. The first, its the very high sensitivity of the fluorophore to its immediate local environment, including the sensitivity to the presence of other fluorophores and quenchers near-by. The second is its unique absorption and emission transition dipoles, which can be interrogated by polarized light. Recently, we have developed 'molecular rulers' which use SMD/SMS to measure nanometer distances and distance changes of macromolecules. We also demonstrated the ability to measure orientations and orientation changes of macromolecules with very high accuracy. The spectroscopic signature of the fluorescent light coming from two different color labels which are attached to two sites on the macromolecule is a sensitive measure for dynamic conformations. Distance changes between two sites on the macromolecule are measured via single-pair fluorescence resonance energy transfer and orientation changes are measured by detecting changes in the dipole orientation of a rigidly attached probe or changes in the degree of rotational diffusion of a tethered probe via single molecule fluorescence polarization anisotropy.

Near-field optics has been utilized for a variety of applications. Using near-field optical probe and microscopy, we have devised a method to introduce a near-field probe into live cultured vascular smooth muscle cell and NG108-15 neuroblastoma cells. We have effectively monitored cellular responses, with excellent spatial and fast temporal resolutions, to drug stimulation. Near-field optical probes enable the visualization of functional response in living cells. We have also nanofabricated the first single molecule optical probe. A single dye molecule, carbocyanine dye C18, is immobilized on a near-field optical probe by physical immobilization. We are able to control the preparation process by selecting the dye molecule concentrations and the interaction times of the probe with the DiI solution. The single DiI molecule probe's optical and spectroscopic properties have been characterized. Photobleaching of a single DiI molecule probe occurs as a discrete and total extinction of its fluorescence. We have also developed ultrasensitive detection schemes using near-field optical probes. Biomolecule immobilization has been carried out on optical fiber probes. Ultrasensitive biochemical sensors for glutamate and lactate have been prepared and characterized.