Time-dependent Spectral Shift Research Articles

Determination of time-resolved fluorescence emission spectra and anisotropies of a fluorophore-protein complex using frequency-domain phase-modulation fluorometry.

We report the first time-resolved fluorescence emission spectra and time-resolved fluorescence anisotropies obtained using frequency-domain fluorescence spectroscopy. We examined the fluorophore p-2-toluidinyl-6-naphthalenesulfonic acid (TNS) in viscous solvents and bound to the heme site of apomyoglobin using multifrequency phase fluorometers. Fluorescence phase shift and modulation data were obtained at modulation frequencies ranging from 1 to 200 MHz. For time-resolved emission spectra, the impulse response for the decay of intensity at each emission wavelength was obtained from the frequency response of the sample at the same emission wavelength. The decays have negative pre-exponential factors, consistent with a time-dependent spectral shift to longer wavelengths. These multiexponential decays were used to construct the time-resolved emission spectra, which were found to be in good agreement with earlier spectra obtained from time-domain measurements. Additionally, time-resolved anisotropies were obtained from the frequency-dependent phase angle differences between the parallel and perpendicularly polarized components of the emission. The rotational correlation times of TNS bound to apomyoglobin are consistent with those expected for this probe rigidly bound to the protein. TNS in propylene glycol also displayed a single exponential decay of anisotropy. These results, in conjunction with the previous successful resolution of multiexponential decays of fluorescence intensity (Lakowicz, J. R., Gratton, E., Laczko, G., Cherek, H., and Limkeman, M. (1984) Biophys. J., in press; Gratton, E., Lakowicz, J. R., Maliwal, B. P., Cherek, H., Laczko, G., and Limkeman, M. (1984) Biophys. J., in press) demonstrate that frequency-domain measurements provide information which is, at a minimum, equivalent to that obtainable from time-domain measurements.

Synthesis and Characterization of a Fluorescence Probe of the Phase Transition and Dynamic Properties of Membranes.

We describe the synthesis and characterization of a new fluorescence probe whose emission spectra, anisotropies, and wavelength-dependent decay times are highly sensitive to the phase state of phospholipid vesicles. This probe is 6-palmitoyl-2-[[2-(trimethylammonio)ethyl]methylamino] naphthalene chloride (Patman). The emission maximum of Patman shifts from 425 to 470 nm at the bilayer transition temperatures. The spectral properties of Patman reveal nanosecond time-dependent spectral shifts, which are the result of membrane relaxation around the excited state of Patman. The apparent fluorescence lifetimes of Patman are strongly dependent upon the emission wavelength, and the fluorescence phase and modulation data prove that the spectral shifts are due to an excited-state process, and not ground-state heterogeneity. As expected, the fluorescence anisotropies reflect the phase transitions of the bilayers. In addition, the anisotropies are dependent upon the emission wavelength because the duration of the excited state varies across the emission spectrum. The different apparent lifetimes across the emission spectrum allow the relaxed and unrelaxed emission spectra to be resolved by phase-sensitive detection of fluorescence. Also, the emission spectra of Patman show marked shifts to longer wavelengths as the excitation wavelength is increased. These red-edge excitation shifts are sensitive to the temperature and phase state of the bilayers.

Time-resolved Emission Spectra of 2-Acetylanthracene/Methanol/Hexane System. Evidence for the Photoassociation of a 1:2 Hydrogen Bonded Complex

Abstract The deconvolved time-resolved emission spectra of 2-acetylanthracene in hexane containing 0.1 mol dm−3 of methanol revealed a considerable time-dependent spectral shift associated with the photoassociation of the hydrogen-bonded complex which includes two methanol molecules.

Analysis of excited-state processes by phase-modulation fluorescence spectroscopy.

Fluorescence phase shift and demodulation methods were used in the analysis of excited-state reactions and to investigate solvent relaxation around fluorophores in viscous solvents. The chosen samples illustrate the results expected for fluorophores bound to biological macromolecules. These moderately simple samples served to test the theoretical predictions described in the preceding paper (J.R. Lakowicz and A.B. Balter, Biophys. Chem. 16 (1982) 99.) and to illustrate the characteristic features of phase-modulation data expected from samples which display time-dependent spectral shifts. The excited-state protonation of acridine and exciplex formation between anthracene and diethylaniline provided examples of one-step reactions in which the lifetimes of the initially excited and the reacted species were independent of emission wavelength. Using these samples we demonstrated the following: (1) Wavelength-dependent phase shift and demodulation values can be used to prove the occurrence of an excited-state process. Proof is obtained by observation of phase angles (ø) larger than 90 degrees or by measurement of ratios of m/cos ø greater than 1, where m is the modulation of the emission relative to that of the excitation. (2) For a two-state process the individual emission spectra of each state can be calculated from the wavelength-dependent phase and modulation data. (3) The phase difference or demodulation factor between the initially excited and the reacted states reveals directly the fluorescence lifetime of the product of the reaction. (4) Phase-sensitive detection of fluorescence can be used to prove that the lifetimes of both the initially excited and the reacted states are independent of emission wavelength. (5) If steady-state spectra of the individual species are known, then phase-sensitive emission spectra can be used to measure the lifetimes of the individual components irrespective of the extent of spectral overlap. (6) Spectral regions of constant lifetime can be identified by the ratios of phase-sensitive emission spectra. In addition, we examined 6-propionyl-2-dimethylaminonaphthalene (PRODAN) and N-acetyl-L-tryptophanamide (NATA) in viscous solvents where the solvent relaxation times were comparable to the fluorescence lifetimes. Using PRODAN in n-butanol we used m/cos ø measurements, relative to the blue edge of the emission, to demonstrate that solvent relaxation requires more than a single step. For NATA in propylene glycol we used phase-sensitive detection of fluorescence to directly record the emission spectra of the initially excited and the solvent relaxed states. These measurements can be easily extended to fluorophores which are bound to proteins and membranes and are likely to be useful in studies of the dynamic properties of biopolymers.

Polar Fluorescent State of 1- and 2-Acylanthracenes. II. The Perturbation of Protic Solvents

Abstract The steady-state and time-resolved fluorescence spectra were measured for protic solutions of 1- and 2-acetyl- or 1- and 2-benzoylanthracenes at various temperatures. It has been demonstrated that the perturbation of protic solvents on the fluorescence spectra is caused by a combination of the orientational dipolar solvent-solute relaxation and the hydrogen bond interaction. Based on the time-dependent spectral shift and the temperature dependence of fluorescence polarization observed for 2-acylanthracene, the relaxation process of the excited solute has been supposed to involve the level inversion of dual fluorescent states, which respond differently to the solvation of the system.

Laser photoionization study of time dependent spectral shifts of localized electrons in ethanol glass. Temperature effects

ADVERTISEMENT RETURN TO ISSUEPREVArticleNEXTLaser photoionization study of time dependent spectral shifts of localized electrons in ethanol glass. Temperature effectsMasaaki Ogasawara and Larry KevanCite this: J. Phys. Chem. 1978, 82, 4, 378–382Publication Date (Print):February 1, 1978Publication History Published online1 May 2002Published inissue 1 February 1978https://doi.org/10.1021/j100493a002RIGHTS & PERMISSIONSArticle Views16Altmetric-Citations8LEARN ABOUT THESE METRICSArticle Views are the COUNTER-compliant sum of full text article downloads since November 2008 (both PDF and HTML) across all institutions and individuals. These metrics are regularly updated to reflect usage leading up to the last few days.Citations are the number of other articles citing this article, calculated by Crossref and updated daily. Find more information about Crossref citation counts.The Altmetric Attention Score is a quantitative measure of the attention that a research article has received online. Clicking on the donut icon will load a page at altmetric.com with additional details about the score and the social media presence for the given article. Find more information on the Altmetric Attention Score and how the score is calculated. Share Add toView InAdd Full Text with ReferenceAdd Description ExportRISCitationCitation and abstractCitation and referencesMore Options Share onFacebookTwitterWechatLinked InReddit PDF (659 KB) Get e-Alerts

Kinetics of electron trapping reactions in amorphous solids; a non-Gaussian diffusion model

Decay patterns of trapped electrons in aqueous glasses have been analyzed over the time span of more than eight decades in terms of a time-dependent "rate constant" of the form ${t}^{\ensuremath{\alpha}\ensuremath{-}1}$, $0<\ensuremath{\alpha}<1$, which can be derived from the long-tail hopping-time distribution of Scher and Montroll. The curve fitting of experiments in our model is quite satisfactory with $\ensuremath{\alpha}\ensuremath{\sim}0.1$ and with another adjustable parameter which depends on the trapping species and varies within one order of magnitude from one system to another. The time-dependent spectral shift for the absorption spectrum of trapped electrons can also be fitted by $\ensuremath{\alpha}\ensuremath{\sim}0.1$, providing fairly direct evidence that non-Gaussian diffusion of electrons from shallow to deep traps underlies the ${t}^{\ensuremath{\alpha}\ensuremath{-}1}$ dependence.

Shift of emission band upon the excitation at the long wavelength absorption edge. III. Temperature dependence of the shift and correlation with the time dependent spectral shift

The edge-excitation red shift previously observed for quinine and related compounds has been shown to exist also for 8-anilinonaphthalene-1-sulfonic acid. The excitation energy dependent spectral shift observed here and the time dependent spectral shift observed by Chakrabarti and Ware have been interpreted satisfactorily in terms of one and the same mechanism. The previously suggested mechanism of the red shift in which the solute–solvent interaction plays an essential role is thus supported. The solvent reorientation relaxation time has been estimated from two different sets of experimental data, one concerned with the excitation energy dependent shift and the other with the time dependent shift. The estimated relaxation times are shown to be entirely different from the dielectric relaxation times of the solvent.

Pulse radiolysis studies of time dependent spectral shifts of the solvated electron in ethanol and deuterated ethanol glasses at 76 K

The initial (?200 nsec) spectra of the trapped electron are similar in C2H5OH, C2H5OD, C2D5OH, and C2D5OD glasses at 76 K with λmax ∼1300 nm. The kinetics of the spectral shifts to the stable values of λmax near 540 nm are similar for C2H5OH and C2H5OD and resemble the kinetics of a series of first order processes or of tunneling reactions. Structure seems to be present in the partially relaxed spectra and may result from specific geoemetrical orientations of first solvation shell molecules.

Nanosecond time-resolved emission spectroscopy of a fluorescence probe bound to L-α-EGG lecithin vesicles

The monophoton counting technique was used to measure nanosecond time-resolved fluorescence emission spectra of 2-p-toluidino-naphthalene-6-sulfonate (2,6 p-TNS) adsorbed to lipid bilayer vesicles. A time-dependent red shift in the emission maximum was observed and the rate of this red shift was shown to be temperature dependent. Analysis of fluorescence decay curves obtained at different emission wavelengths indicates that the time-dependent spectral shifts are due to an excited-state reaction such as solvent relaxation or an excited-state interaction between the chromophore and a polar residue of the phospholipid.

Nanosecond Time-Resolved Emission Spectroscopy of 1-Anilino-8-Naphthalene Sulfonate

Nanosecond time-resolved emission spectroscopy and fluorescence lifetime measurements are reported for 1-anilino-8-naphthalene sulfonate. Time-dependent spectral shifts are observed that are interpreted in terms of dipole reorientation of the solvent around the electronically excited molecule. Both the radiative and nonradiative transitions probabilities are observed to be strongly solvent dependent.

Nanosecond Time-Resolved Emission Spectroscopy: Spectral Shifts due to Solvent-Excited Solute Relaxation

Nanosecond time-resolved emission spectral techniques have been applied to the problem of the origin and nature of the well-known temperature-dependent spectral shifts characteristic of the aminophthalimides in alcohol solvents. It is demonstrated that the temperature-dependent spectral shifts are in fact due to time-dependent spectral shifts. At least two relaxation times characterize this phenomenon. One relaxation time is observed to be subnanosecond in character and may be associated with the exciplex that presumably is present in the system. The other relaxation time is presumably associated with the nonspecific dipolar reorientation although it has distinctly different characteristics from the solvent dielectric relaxation time. Wavelength-dependent fluorescence decay that can be explained by the time dependence of the emission spectrum is also observed.