Spectrum Correction in Study of Solvation Dynamics by Fluorescence Non-collinear Optical Parametric Ampli cation Spectroscopy

Femtosecond time-resolved fluorescence non-collinear optical parametric amplification spectroscopy can extract the curve of spectral gain from its parametric superfluorescence. This unique spectrum correction method enables fluorescence non-collinear optical parametric amplification spectroscopy acquiring the genuine transient fluorescence spectrum of the studied system. In this work we employ fluorescence non-collinear optical parametric amplification spectroscopy technique to study the solvation dynamics of DCM dye in ethanol solution, and confirm that genuine solvation correlation function and shift of peak frequency can be derived from transient fluorescence spectra after the spectral gain correction. It demonstrates that fluorescence non-collinear optical parametric amplification spectroscopy can benefit the research fields, which focuses on both fluorescence intensity dynamics and fluorescence spectral shape evolution.     

Long-range corrected time-dependent density functional study on fluorescence of 4,4'-dimethylaminobenzonitrile

Dual fluorescence of 4,4(')-dimethylaminobenzonitrile (DMABN) was theoretically investigated on the basis of long-range corrected time-dependent density functional theory. Excited-state geometry optimization states and single-point energy calculations with and without solvent effect were carried out. It has been explained that DMABN emits dual fluorescence only in polar solvents through locally excited (LE) and charge transfer (CT) states. It was, however, concluded from this study that although the main spectrum of dual fluorescence in acetonitrile solvent is clearly due to twisted intramolecular CT fluorescence, small secondary fluorescence in acetonitrile may also emanate from CT fluorescence during the DMABN twisting process. This conclusion is supported by an experimental interpretation on polarization spectroscopy. It was also found that the optimized DMABN geometries have certain wagging angles for the CT state and no wagging angle for the LE state. This may support an early experimental hypothesis that the dual fluorescence of DMABN is induced by the wagging mode due to vibronic coupling between LE and CT states. Consequently, the authors propose a fluorescence mechanism of DMABN in gas phase and in acetonitrile solvent: the main absorption proceeds to the CT state in both situations. In gas phase, single fluorescence is chiefly emitted from the LE state through the internal conversion from CT to LE states. Dual fluorescence in acetonitrile solvent may only be emitted from the CT state.     

Solvent effect in an “isolated” solvated molecule in a free jet: Intramolecular charge-transfer state of solvated 9,9'-bianthryl

The fluorescence spectra of bare and solvated 9,9'-bianthryl were recorded in a free Jet. In the case of bare bianthryl, only the fluorescence from the locally excited (LE) state was observed, even on excitation to a high vibronic state. In contrast, the excitation of the 0-0 band of 9,9'-bianthryl solvated by acetone gave fluorescence from the charge-transfer (CT) state. The ratio of the intensity of the CT fluorescence to that of the LE fluorescence increases with increasing degree of solvation. Solvation by cyclohexane did not give CT fluorescence.     

Light-induced redistribution and photobleaching of protoporphyrin in erythrocytes from patients with erythropoietic protoporphyria: an explanation of the rapid fading of fluorocytes

Erythrocytes from patients with erythropoietic protoporphyria (EPP) contain large amounts of protoporphyrin. By fluorescence microscopy it has been found that the erythrocytes show red fluorescence which fades very rapidly. During irradiation of erythrocytes from patients with EPP, a decrease in the fluorescence intensity and a red shift in the fluorescence emission maximum is observed. Since the human eye has a diminishing sensitivity in the red part of the spectrum, the red shift will augment the fluorescence decrease observed in the microscope. The decrease in fluorescence intensity is greater than what would be expected by photobleaching of protoporphyrin alone. We suggest that the rapid fading of fluorocytes observed in a fluorescence microscope can be explained both by photoinduced detachment of protoporphyrin from hemoglobin followed by a redistribution to the erythrocyte membrane and by protoporphyrin photobleaching.     

Fluorescence resonance energy transfer by quencher adsorption into hydrogels containing fluorophores

We synthesized anionic hydrogels containing fluorophores and investigated the adsorption of a cationic quencher having an amino group into hydrogels by fluorescence resonance energy transfer (FRET). FRET from the fluorophore to the quencher in hydrogels was examined by fluorescence intensity and fluorescence decay using a fluorescence spectrophotometer and femtosecond laser spectroscopy. The fluorescence intensity of the fluorophore‐containing hydrogels decreased rapidly with increasing amounts of adsorbed cationic quencher. The fluorescence emission of the fluorophore in the quencher‐adsorbed hydrogels containing fluorophores decayed more rapidly than that of the original hydrogels. The aforementioned result indicates that the fluorescence of the fluorophore‐containing hydrogels is quenched due to FRET from the fluorophore to the quencher as the cationic quenchers can approach the fluorophores in hydrogels by electrostatic interactions. © 2006 Wiley Periodicals, Inc. J Polym Sci Part B: Polym Phys 44: 3245–3252, 2006     

Distance-dependent metal-enhanced fluorescence from Langmuir-Blodgett monolayers of alkyl-NBD derivatives on silver island films

We described the effect of fluorophore distance from the silver island films (SIFs) on the metal-enhanced fluorescence (MEF) from two newly developed long-chain nitrobenzoxadiazole derivatives (NBD-C16 and NBD-C18). The well-established Langmuir-Blodgett technique is used to deposit the fluorophores at defined distances from the SIFs surface, and an inert amphiphilic stearic acid is used to control the distance. NBD probes deposited directly on the SIFs surface show the highest metal-enhanced fluorescence of approximately 32-fold, and both of the probes that were studied show a consistent decrease in metal-enhanced fluorescence when increasing the distance from the fluorophore to the SIFs surface. The lowest fluorescence enhancement of approximately 4-fold is observed for the probes located 90 nm from the SIFs surface. Additionally, we also have noticed the shortest fluorescence lifetimes for the NBD probes deposited directly onto the SIFs surface, and the lifetimes are consistently increased when increasing the distances between the fluorophore and SIFs surfaces. These contrasting spectral changes, enhanced fluorescence, and decreased fluorescence lifetimes are in accordance with an increase in the rate of radiative decay for fluorophores near the silver particles. The present study provides significant information on the effect of fluorophore distance on the metal-enhanced fluorescence phenomenon.     

Excitation emission and time-resolved fluorescence spectroscopy of selected varnishes used in historical musical instruments

The analysis of various varnishes from different origins, which are commonly found on historical musical instruments was carried out for the first time with both fluorescence excitation emission spectroscopy and laser-induced time-resolved fluorescence spectroscopy. Samples studied include varnishes prepared using shellac, and selected diterpenoid and triterpenoid resins from plants, and mixtures of these materials. Fluorescence excitation emission spectra have been collected from films of naturally aged varnishes. In parallel, time-resolved fluorescence spectroscopy of varnishes provides means for discriminating between short- (less than 2.0 ns) and long-lived (greater than 7.5 ns) fluorescence emissions in each of these complex materials. Results suggest that complementary use of the two non destructive techniques allows a better understanding of the main fluorophores responsible for the emission in shellac, and further provides means for distinguishing the main classes of other varnishes based on differences in fluorescence lifetime behaviour. Spectrofluorimetric data and time resolved spectra presented here may form the basis for the interpretation of results from future in situ fluorescence examination and time resolved fluorescence imaging of varnished musical instruments.     

External electric field effects on fluorescence of pyrene butyric acid in a polymer film: concentration dependence and temperature dependence

Fluorescence spectra and electrofluorescence spectra (plots of the electric field-induced change in fluorescence intensity as a function of wavelength) have been measured at different temperatures for pyrene butyric acid (PBA) in a PMMA film at different concentrations. At a low concentration of 0.5 mol % where fluorescence emitted from the locally excited state of PBA (LE fluorescence) is dominant, LE fluorescence spectra show only the Stark shift in the presence of an electric field (F), which results from the difference in molecular polarizability between the ground and emitting states. At a high concentration of 10 mol % where the so-called sandwich-type excimer fluorescence (EX(1)) is dominant, both EX(1) and LE fluorescence are quenched by F. Another fluorescence assigned to a partially overlapped excimer (EX(2)) also exists at room temperature, and this emission is enhanced by F. As the temperature decreases, three fluorescence emissions whose electric field effects are different from each other become clear besides EX(1) and LE fluorescence, indicating that at least five fluorescence components exist at high concentrations at low temperatures. At a medium concentration of 5 mol % where EX(1) is comparable in intensity to the LE fluorescence, the intensity of EX(1) is not affected by F at any temperature, but LE fluorescence and EX(2) are markedly influenced by F at room temperature, and four fluorescence emissions are confirmed at low temperatures.     

Autofluorescence spectroscopy of normal and malignant human breast cell lines

The fluorescence of tryptophan, reduced nicotinamide adenine dinucleotide (phosphate) (NAD(P)H) and flavin adenine dinucleotide (FAD) were characterized in normal human breast cells as well as in malignant human breast cells of similar and dissimilar genetic origins. Fluorescence measurements of each cell line were made over a wide range of cell concentrations, and the fluorescence per cell was determined from the slope in the linear range of the fluorescence intensity vs cell concentration plot. All of the malignant cells showed a statistically significant decrease in the tryptophan fluorescence per cell relative to that of the normal cells. No statistically significant differences were observed in the NAD(P)H or FAD fluorescence per cell between the normal and any of the malignant cell types. NAD(P)H fluorescence was also imaged from monolayers of the normal and malignant cells (of similar genetic origin) using two-photon fluorescence microscopy. A statistically significant decrease in the NAD(P)H fluorescence with malignancy was observed, suggesting that fluorescence imaging of single cells or the cell monolayer preparation may provide more contrast than volume-averaged fluorescence measurements of cells in suspension. In conclusion, the differences in normal and malignant human breast tissue fluorescence spectra may be attributed in part to differences in the intrinsic cellular fluorescence of normal and malignant breast epithelial cells.     

Modulated Fluorescence in LB Films Based on DADQs—A Potential Sensing Surface?

Novel fluorescent Langmuir-Blodgett (LB) films have been constructed from three different amphiphilic dicynaoquinodimethanes (DADQs). The DADQs varied in functional group structure, which had an impact on the LB film structure and the fluorescence properties. As the fluorescence of DADQs competes with non-radiative decay (conformational change), the packing and/or free volume in the LB film will influence the average fluorescence lifetime and integrated intensity. The pristine (blank) LB films were then exposed to a selection of non-fluorescent target analytes (some with environmental relevance) and the fluorescence was measured and analyzed relative to the pristine LB film. Exposure of the LB films to selected target analytes results in a modulation of the fluorescence, both with respect to average fluorescence lifetime and integrated intensity. The modulation of the fluorescence is different for different DADQ LB films and can be attributed to restricted non-radiative decays or charge transfer reactions between target analyte and DADQ LB film. The response from the DADQ LB films shows that these systems can be developed into sensing surfaces based on fluorescence measurements.     

Photophysics of alpha,omega-diphenyloctatetraene in the vapor phase

Fluorescence and fluorescence excitation spectra of diphenyloctatetraene vapor have been measured at different temperatures from 98 to 136 degrees C and at different buffer gas pressures from 0 to 300 Torr. The fluorescence quantum yields were determined as functions of the excitation energy and buffer gas pressure. It is shown that diphenyloctatetraene vapor exhibits weak fluorescence from the S2 (1(1)Bu) state in addition to the fluorescence from the S1 (2(1)Ag) state. The quantum yield of the S1 fluorescence is shown to decrease with decreasing pressure and with increasing excitation energy. The electronic relaxation processes of diphenyloctatetraene vapor are discussed based on the pressure and excitation-energy dependence of the fluorescence quantum yield.     

Solvent Effects on Spectral Property and Dipole Moment of the Lowest Excited State of Coumarin 343 Dye

Steady-state absorption and fluorescence spectra, and time-resolved fluorescence spectra of coumarin 343 (C343) were measured in different solvents. The effect of the solvent on the spectral properties and dipole moment of the lowest excited state of C343 were investigated. It was found that the absorption and fluorescence spectra red-shifted slightly and strongly with increasing solvent polarity, respectively, because the charge distribution of the excited state leaded to the increasing difference between the absorption and fluorescence spectra with increasing solvent polarity. The dipole moment of the lowest excited state of C343was determined from solvatochromic measurements and the quantum chemical calculation, and the results obtained from these two methods were fully consistent. Investigations of the time-resolved fluorescence of C343 in different solvents indicated that the fluorescence lifetimes increased nearly linearly with increasing solvent polarity from 3.09 ns in toluene to 4.45 ns in water. This can be ascribed to the intermolecular hydrogen bonding interactions between C343 and hydrogen donating solvents.     

Relationship between collagen autofluorescence of the human cervix and menopausal status

The goal of this study was to evaluate the effect of different menopausal states (pre- and post-) on the endogenous fluorescence of normal cervical tissues. In particular, the average fluorescence as well as the interpatient and intrasample variability in the average fluorescence of the epithelium and stroma were evaluated as a function of pre- and postmenopausal states. High-resolution fluorescence images at excitation-emission wavelengths of 440, 520 nm and 365, 465 nm were obtained from epithelia and stroma of freeze-trapped cervical tissue blocks maintained at -196 degrees C. The fluorescence images were recorded using a low temperature optical scanner. Fluorescence images from a normal sample population (n = 27) were quantitatively analyzed, and the average epithelial and stromal fluorescence intensities were obtained. Data grouped according to menopausal status (pre- vs post-) showed statistically significant differences (P < 0.002) in stromal fluorescence. In particular, the cervical stroma of postmenopausal women showed (1) significantly greater average fluorescence and (2) greater interpatient and intrasample variability in the fluorescence, relative to that of premenopausal women. These results provide evidence for changes in collagen cross-linking with menopause.     

Nanostructure origins of C60 fluorescence in pyridine

The nanostructure origins of three distinct and strong fluorescence bands of C60 in room-temperature pyridine are reported for the first time. Fluorescence study and in-situ TEM observation on C60-pyridine solutions with different setting times exhibit that the blue fluorescence peaks centered at 440 nm originate from C60 nanoparticles; the yellow-green fluorescence band located at 575 nm derives from C60 lace-like cluster; and the salmon pink fluorescence band around 700 nm arises from C60 microbulk in solution. The conclusions are supported further by investigation on the C60-toluene system as well.     

Self‐Assembly and Aggregate‐Induced Enhanced Emission of Amphiphilic Fluorescence Dyes in Water and in the Solid State

1‐Cyano‐1,2‐bis(biphenyl)ethene (CNBE) derivatives with a hexa(ethylene glycol) group as an amphiphilic side chain were synthesized and the self‐assembling character and fluorescence behavior were investigated. The amphiphilic derivatives showed aggregate‐induced enhanced emission (AIEE) in water and in the solid state. The fluorescence quantum yield increased as the rigidity of the aggregates increased (i.e., in ethyl acetate<in water<in the solid state). As determined from measurements of fluorescence spectra, fluorescence quantum yields, and fluorescence lifetimes, a key factor for the enhanced emission is suppression of the nonradiative decay process arising from restricted molecular motion. Additionally, the difference in the emission rate constant is not negligible and can be used to interpret the difference in fluorescence quantum yield in water and in the solid state.     

Rapid amplitude-modulation of a diarylethene photoswitch: en route to contrast-enhanced fluorescence imaging

A water soluble diarylethene (DAE) derivative that displays exceptionally intense fluorescence from the colorless open form has been synthesized and characterized using UV/vis spectroscopy and fluorescence microscopy. We show that the bright emission from the open form can be rapidly switched using amplitude modulated red light, that is, by light at wavelengths longer than those absorbed by the fluorescent species. This is highly appealing in any context where undesired background fluorescence disturbs the measurement, e.g., the autofluorescence commonly observed in fluorescence microscopy. We show that this scheme is conveniently applicable using lock-in detection, and that robust amplitude modulation of the probe fluorescence is indeed possible also in cell studies using fluorescence microscopy.

A water soluble diarylethene derivative displaying exceptionally bright fluorescence in the open isomeric form has been used for emission amplitude-modulation. We apply this scheme in fluorescence microscopy, aiming to suppress undesired background.     

一种pH荧光探针的合成及光谱研究

设计合成了一种pH荧光分子探针2,5-双(4-羟基-苯亚甲基)环戊酮,并对其光谱性能进行了研究.pH滴定实验表明:探针的紫外吸收和荧光光谱对溶液的pH值有很强的依赖性.当体系溶液由酸性变为碱性时,探针紫外吸收光谱发生明显的红移,并伴有溶液颜色的显著变化.荧光光谱强度在酸性条件下随pH的变化不大,而在碱性条件下荧光强度则...     

Synthesis and fluorescence study of 7-azaindole in DNA oligonucleotides replacing a purine base

The fluorescence spectroscopy of 7-azaindole (7aIn) incorporated in DNA oligonucleotides is investigated. Incorporation of 7aIn into DNA oligonucleotides is accomplished through standard solid-phase phosphoramidite chemistry. Fluorescence emission of the 7aIn chromophore shifts slightly to the red (from 386 nm to 388 nm) upon glycosylation at the N-1 position, but its relative fluorescence quantum yield increases 23 times, from 0.023 to 0.53. Upon incorporation into DNA, the fluorescence emission of 7aIn is greatly quenched with fluorescence quantum yields of 0.020 and 0.016 in single and double strand DNA, respectively. The fluorescence emission for 7aIn in DNA oligonucleotides shifts to the blue with an emission maximum at 379 nm. Both the strong fluorescence quenching and the blue shift of the emission spectrum signify that 7aIn is stacked with neighboring DNA bases in both single and double strand DNA. As the duplex DNA melts due to temperature increase, the fluorescence of the 7aIn chromophore increases, indicating the transition from the less fluorescent duplex DNA to the more fluorescent single strand DNA. Since this fluorescent 7aIn is a structural analog of purine, its fluorescence property may be utilized as a probe for studying nucleic acid structure and dynamics.     

Triphenylmethane dyes as fluorescent probes for G-quadruplex recognition

Triphenylmethane (TPM) dyes normally render rather weak fluorescence due to easy vibrational deexcitation. However, when they stack onto the two external G-quartets of a G-quadruplex (especially intramolecular G-quadruplex), such vibrations will be restricted, resulting in greatly enhanced fluorescence intensities. Thus, TPM dyes may be developed as sensitive G-quadruplex fluorescent probes. Here, fluorescence spectra and energy transfer spectra of five TPM dyes in the presence of G-quadruplexes, single- or double-stranded DNAs were compared. The results show that the fluorescence spectra of four TPM dyes can be used to discriminate intramolecular G-quadruplexes from intermolecular G-quadruplexes, single- and double-stranded DNAs. The energy transfer fluorescence spectra and energy transfer fluorescence titration can be used to distinguish G-quadruplexes (including intramolecular and intermolecular G-quadruplexes) from single- and double-stranded DNAs. Positive charges and substituent size in TPM dyes may be two important factors in influencing the binding stability of the dyes and G-quadruplexes.     

Fluorescence decay of α-chymotrypsin studied by the picosecond-resolved single photon-counting technique

The fluorescence decay of the multi-tryptophan-containing enzyme α-chymotrypsin in Tris buffer (pH 7.8) at room temperature was studied using a frequency-doubled, synchronously-pumped picosecond rhodamine-6G laser excitation source with time-correlated single photon-counting detection. The fluorescence decay parameters were computed with a non-linear least-squares iterative reconvolution program. The goodness-of-fit was tested with well-known graphical methods such as residuals plots and the autocorrealtion function. Numerical tests (reduced chi-square, ordinary runs test and the Durbin—Watson statistic) were included to improve the reliability of the residuals analysis. Normal distribution of the weighted residuals was checked with the normal probability plot, and with computation of the mean and standard deviation of the weighted residuals. α-Chymotrypsin exhibited triple-exponential fluorescence decay kinetics with decay times of 615 ± 76 ps, 1.7 ± 0.2 ns, and 4.3 ± 0.3 ns. The fractional fluorescence contributions depended on the emission wavelength. The fluorescence spectra of the components contributing to the total fluorescence were calculated from the steady-state fluorescence spectrum and fluorescence decays at different emission wavelengths, and from convoluted time-resolved emission spectra and a fluorescence decay measurement.