Fluorescence upconversion

Figure B2.1.4 Fluorescence upconversion spectrometer based on the use of off-axis elliptical reflectors for the collection and focusing of fluorescence. Symbols used el, c2, off-axis elliptical reflectors s, sample x, nonlinear crystal. (After Jimenez and Fleming [21].)...

The instrument response fiinction (IRF) for the fluorescence upconversion experiment, then, caimot be shorter than the intensity cross-correlation fiinction, which can be obtained usmg an mstniment like that shown in figure B2.1.4... 

Figure B2.1.5 Fluorescence upconversion traces obtained at two observation wavelengdis (fiill circles, 570 mn open circles, 650 mn) at room temperature with an oxazine dye, phenoxazone, in methanol solvent. Figure courtesy of Professor S Rosenthal (Vanderbilt University).

The main cost of this enlianced time resolution compared to fluorescence upconversion, however, is the aforementioned problem of time ordering of the photons that arrive from the pump and probe pulses. Wlien the probe pulse either precedes or trails the arrival of the pump pulse by a time interval that is significantly longer than the pulse duration, the action of the probe and pump pulses on the populations resident in the various resonant states is nnambiguous. When the pump and probe pulses temporally overlap in tlie sample, however, all possible time orderings of field-molecule interactions contribute to the response and complicate the interpretation. Double-sided Feymuan diagrams, which provide a pictorial view of the density matrix s time evolution under the action of the laser pulses, can be used to detenuine the various contributions to the sample response [125]. 

Underwood, D. E., Kippeny, T. and Rosenthal, S. J. (2001) Ultrafast carrier dynamics in CdSe nanocrystals determined by femtosecond fluorescence upconversion spectroscopy. /. Phys. Chem. B, 105,436-443. 

Gustavsson T, Sharonov A, Markovitsi D (2002) Thymine, thymidine and thymidine 5 -monophosphate studied by femtosecond fluorescence upconversion spectroscopy. Chem Phys Lett 351 195... 

Gustavsson T, Sarkar N, Lazzarotto E, Markovitsi D, Improta R (2006) Singlet excited-state dynamics of uracil and thymine derivatives a femtosecond fluorescence upconversion study in acetonitrile. Chem Phys Lett 429 551-557... 

Wang H, Zhang H, Abou-Zied OK et al (2003) Femtosecond fluorescence upconversion studies of excited-state proton-transfer dynamics in 2-(20-hydroxyphenyl)benzoxazole (HBO) in liquid solution and DNA. Chem Phys Lett 367 599-608... 

Fujino, T., and Tahara, T. 2004. Characterization and performance of femtosecond fluorescence upconversion microscope. Appl. Phys. B 79 145. 

The fluorescence quenching of Pe and derivatives has been investigated by fluorescence upconversion. Excitation was performed with the frequency-doubled output of a Ti Sapphire amplifier. The instrument response time was around 240 fs with 0.4 mm thick samples. The data were analysed by iterative reconvolution of the instrument response function with trial functions. For most samples, measurements were carried out at three different wavelengths (438, 475, and 490 nm). Global fits were done with all the available data. 

Fig. 2. Time-profiles of the fluorescence at 475 nm upon excitation at 400 nm of Pe, PeCN, PeOH and PeMe in DMA monitored by fluorescence upconversion and best triexponential fits.

Fluorescence upconversion and time correlated single photon counting... 

Precise measurements of the excited state lifetimes of the DNA constituents were not available till very recently, mainly due to the limited time resolution of conventional spectroscopic techniques. Studying the DNA nucleosides by transient absorption spectroscopy, Kohler and co-workers observed a very short-lived induced absorption in the visible which they assigned to the first excited state [5,6]. The lifetimes observed were all well below 1 picosecond. The first femtosecond fluorescence studies of DNA constituents were performed using the fluorescence upconversion technique. Peon and Zewail [7] reported that the excited state lifetimes of DNA/RNA nucleosides and nucleotides all fall in the subpicosecond time, thus corroborating the results obtained by transient absorption. 

At the same time we have, in a series of papers, pursued the study of DNA constituents using the fluorescence upconversion technique [8-11]. 

Femtosecond fluorescence upconversion studies of chiral self-assembled supramolecules in solution... 

Sample preparation was given elsewhere [2]. Femtosecond fluorescence upconversion and picosecond time-correlated single-photon-counting set-ups were employed for the measurement of the fluorescence transients. The system response (FWHM) of the femtosecond fluorescence up-conversion and time-correlated single-photon-counting setups are 280 fs and 16 ps, respectively [3] The measured transients were fitted to multiexponential functions convoluted with the system response function. After deconvolution the time resolution was 100 fs. In the upconversion experiments, excitation was at 350 nm, the transients were measured from 420 nm upto 680 nm. Experiments were performed under magic angle conditions (to remove the fluorescence intensity effects of rotational motions of the probed molecules), as well as under polarization conditions in order to obtain the time evolution of the fluorescence anisotropy. 

Figure 2a shows a few fluorescence upconversion transients as measured for 1 dissolved in n-heptane, under magic angle conditions. The transients show multi-exponential decay behavior... 

First results from our fluorescence upconversion experiments are shown in Fig. 2, which displays the solvation-functions of C343 in bulk water and adsorbed on ZrC>2 nanoparticles. The response in bulk water confirms the previously reported results of bimodal dynamics [8] and a corresponding behaviour can be found for the dye bound to Z1O2, indicating that similar processes are involved. The results from biexponential fits to the solvation function S(t) of C343 in pure water and at the ZrCh-water interface are listed in Table 1. In both cases we find a fast decay time of about 100 fs and a slower decay of about 750 fs. We can see that the individual decay times stay similar and only the relative contributions change, resulting in an overall somewhat faster solvation for adsorbed dyes. 

Figure 7. Schematic of the fluorescence upconversion apparatus. The notation is defined in the text. From Ref. 6 with permission, from J. I mag. Sci. 33, 53 (1989). Copyright Society for Imaging Science and Technology.

For alcohol solvents, measurements were made with time-correlated single photon counting. The remaining measurements were made with the fluorescence upconversion system. The transients in alcohol solvents were fitted with a single exponential kinetic function. The kinetics in acetone is also well described by a single exponential, but in benzonitrile, dimethyl-sulfoxide, and propylene carbonate the kinetics were modeled with a biexponential decay. 

Fig. 11 Illustration of the excited state relaxation derived from experimental results obtained for poly(dA).poly(dT) by steady-state absorption and fluorescence spectroscopy, fluorescence upconversion and based on the modeling of the Franck-Condon excited states of (dA)io(dT)io. In red (full line) experimental absorption spectrum yellow circles arranged at thirty steps represent the eigenstates, each circle being associated with a different helix conformation and chromophore vibrations.

In this section, I will discuss some of the more recent developments in continuum solvation dynamics in polar solvents. Some of these deal with incorporation of realistic models for chromophores [8,43 16] used in fluorescence-upconversion experiments, others with improvements in modeling of the solution dielectric properties [47,48], including incorporation solvent dielectric response over a wide frequency range [43,44, 46,48] into theories of SD. 

Rising components of 10 ps, attributable to intramolecular H-atom transfer are clearly observed in the fluorescence upconversion traces of both hypericin and hypocrellin A (Figs. 1.3 and 1.4, respectively). For simplicity, in the rest of our discussion, we refer to this shorter-lived component as the 110-ps component. The amplitude of the rising component is emission wavelength dependent and occurs on the blue edge of the emission spectra. The clear and complementary observation in fluorescence of the 10-ps component in both hypericin and hypocrellin A is a crucial link in providing a unified model of the hypericin and... 

Figure 1.3. (Top) Fluorescence upconversion transient for hypericin in ethanol at /,cm = 576 nm. The fit curve is described hy the following equation (with background subtracted) F(t) — —0.21 exp (—f/6.5ps) + 1.00 exp (—r/oo). (Bottom) At /.em = 653 nm, however, there is no rising component in the fluorescence trace. Similar behavior is observed for hypocrellin A (Fig. 1.4). The excitation wavelength was the second harmonic of our unamplified Ti sapphire oscillator, 414 nm. The panel below the kinetic trace displays the residuals between the fit and the data.

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