Eosin phosphorescence

The intensity of the delayed fluorescence emission from eosin decreases as the temperature is lowered and this indicates that an energy barrier is involved. Since the delayed fluorescence is spectrally identical to normal fluorescence, emission must occur from the lowest vibrational level of Si. However, the fact that the lifetime is characteristic of phosphorescence implies that the excitation originates from T,. The explanation of this requires a small Si-Ti energy gap, where T, is initially populated by intersystem crossing from Si. Ti to Si intersystem crossing then occurs by thermal activation. 

Photoinduced electron transfer from eosin and ethyl eosin to Fe(CN)g in AOT/heptane-RMs was studied and the Hfe time of the redox products in reverse micellar system was found to increase by about 300-fold compared to conventional photosystem [335]. The authors have presented a kinetic model for overall photochemical process. Kang et al. [336] reported photoinduced electron transfer from (alkoxyphenyl) triphenylporphyrines to water pool in RMs. Sarkar et al. [337] demonstrated the intramolecular excited state proton transfer and dual luminescence behavior of 3-hydroxyflavone in RMs. In combination with chemiluminescence, RMs were employed to determine gold in aqueous solutions of industrial samples containing silver alloy [338, 339]. Xie et al. [340] studied the a-naphthyl acetic acid sensitized room temperature phosphorescence of biacetyl in AOT-RMs. The intensity of phosphorescence was observed to be about 13 times higher than that seen in aqueous SDS micelles. 

Phosphorescence measurements of eosin in ethanol place the 0-0 transition of 3Si 3S0 at 7150 A.98 Recent determinations of phosphorescence spectra of xanthene dyes in EPA at 77°K show these transitions to occur in the region of 7000 A 94,103 (Table I). The triplet energies of these dyes, 3Bi, are thus about 40 kcal/mole and the energy separations between the excited singlet and triplet states, AE = Elai — 3si, are about 12-13 kcal/mole. 

In the delayed emission spectrum of eosin in glycerol or ethanol two bands are present, the relative intensities of which are strongly temperature-dependent (see Fig. 12). The visible band at 1.8 has a contour identical with that of the fluorescence band. It no doubt corresponds to the visible phosphorescence observed by Boudin.26 To interpret the results it was assumed that this band of delayed fluorescence was produced by thermal activation of the eosin triplet to the upper singlet level followed by radiative transition from there to the ground state. The far red band was assumed to correspond to the direct transition from the triplet level to the ground state and was therefore called phosphorescence. To determine the relationship between the intensities of the two bands we write the equations for the formation and consumption of triplet molecules as follows ... 

Triplet-Singlet Phosphorescence eosin, proflavine hydrochloride, phenanthrene. 

Reticulum ATPase [105,106], Owing to the long-lived nature of the triplet state, Eosin derivatives are suitable to study protein dynamics in the microsecond-millisecond range. Rotational correlation times are obtained by monitoring the time-dependent anisotropy of the probe s phosphorescence [107-112] and/or the recovery of the ground state absorption [113— 118] or fluorescence [119-122], The decay of the anisotropy allows determination of the mobility of the protein chain that cover the binding site and the rotational diffusion of the protein, the latter being a function of the size and shape of the protein, the viscosity of the medium, and the temperature. 

Fluorescein and related quinonelike dyes such as eosin have found extensive use as low-energy sensitizers, especially in photooxidation studies. The efficiency of intersystem crossing in fluorescein varies markedly with pH. For the neutral molecule, the quantum yield of fluorescence is only 0.31, while it is 0.91 for the dianion.449 In strong acid solutions, however, protonated fluorescein is strongly phosphorescent. In this state (dissolved in boric acid) fluorescein was the first compound for which T-T absorption in an excited triplet state was observed.7... 

Figure 1.9. (A) E-type Fluorescence and phosphorescence emission spectra, ex = 473 nm, of Eosin in a cuvette at different temperatures. Insert Eosin immobilized in PVA sandwiched between two silvered and unsilvered slides at 25C EF - Enhancement Factor. RT - Room Temperature. (B) experimental sample geometry. (C) Real-color photographs of Eosin emission from glass and SIFs, before and after 2 mins heating, ex = 473 mn. SIFs - Silver Island Films. The real- color photographs were taken through an emission filter (488 mn razor edge).

B. Phosphorescence lifetimes are typicaUy near l-IO ms. Assume that the nahiral fifetime for phosphores cence emission of these compounds is 10 ms and that the nonra< ative dec rates of the two compomds are the same for the triplet state as for the singlet state. Estim e the phosphorescence quantum yields of eosin and Er B at room tempoature. 

Fig. 8 The simultaneous generation of singlet oxygen phosphorescence and erbimn(in) luminescence by an eosin-modified DTPA complex in buffered D20 (pD 8.3), dtanonstrating the ctnnpa-rable (very weak) luminescence intensities of erbium(III) luminescence and singlet oxygtai emission. Upon deoxygenation, the erbium(ni) luminescence yield increases, which shows the competition between oxygen and cu-biumflll) in the energy transfer liom eosin...

In Section 2.5 we described the use of time-resolved fluorescence anisotropy for monitoring protein motion on the nanosecond timescale. For motion on much longer timescales, time-resolved phosphorescence anisotropy can be used instead. The latter technique has been employed, for example, to examine the rotational motion of membrane-boimd proteins labelled with the triplet probe eosin (57, 58). Prior to making the measurements, the protein is labelled with eosine-maleimide as described in Protocol 2. 

Eosine labelling of proteins for time-resolved phosphorescence measurements... 

E-type delayed fluorescence. As defined by the lUPAC Gold book [2], this is the process in which the first excited singlet state becomes populated by a thermally activated radiationless transition from the first excited triplet state. Since in this case the populations of the singlet and triplet states are in thermal equilibrium, the lifetimes of delayed fluorescence and the concomitant phosphorescence are equal. This process takes its name from eosin and is typically observed with dyes, where the Si-Ti gap is small. 

Eosin and its derivatives show weak room temperature phosphorescence and are particularly useful as phosphorescent probes for measuring the rotational properties of proteins and other biomolecules in solution and in membranes. Also useful for FRET studies and fiuorescence recovery after photobleaching (FRAP) measurements of diffusion. 

DLA Phosphorescence of Eosin-labeled Fatty Acids in Phospholipid Bilayers... 

We have investigated further the rotation of DMPC lipids by incorporating two eosin fatty acid probes (dodecanoyl- and hexadecanoyl-amidoeosin) and measuring the time-dependent phosphorescent anisotropies (37). The eosin moieties of these reporter molecules are located close to the membrane surface. Figure 7 shows typical experimental results at two temperatures. A number of features serve to illustrate the type of information provided by such studies. The phosphorescence emission at both temperatures displayed a time-dependent anisotropy which could be fit to an equation of the form... 

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