Characterization of Photoinduced Isomerization and Intersystem Crossing of the Cyanine Dye Cy3

Abstract

Several important photophysical properties of the cyanine dye Cy3 have been determined by laser flash photolysis. The triplet-state absorption and photoisomerization of Cy3 are distinguished by using the heavy-atom effects and oxygen-induced triplet --> triplet energy transfer. Furthermore, the triplet-state extinction coefficient and quantum yield of Cy3 are also measured via triplet-triplet energy-transfer method and comparative actinometry, respectively. It is found that the triplet --> triplet (T1-->Tn) absorptions of trans-Cy3 largely overlap the ground-state absorption of cis-Cy3. Unlike what occurred in Cy5, we have not observed the triplet-state T1-->Tn absorption of cis-Cy3 and the phosphorescence from triplet state of cis-Cy3 following a singlet excitation (S0-S1) of trans-Cy3, indicating the absence of a lowest cis-triplet state as an isomerization intermediate upon excitation in Cy3. The detailed spectra of Cy3 reported in this paper could help us interpret the complicated photophysics of cyanine dyes.

Single molecule fluorescence fluctuations of the cyanine dyes linked covalently to DNA

“…In order to quantitatively analyze the observed on-time and off-time in the fluorescence intensity time-traces, we set a suitable threshold between the on-and off-levels according to the count rate distribution in Figure 1 In our previous studies [1,7,8,21] , we intensively studied the photophysical properties of triplet state and isomerization of free-Cy3 and free-Cy5. Cyanine dyes are believed to adopt an all-trans configuration in its ground state [1,8,22,23] . For the light-induced isomerization of cyanine dyes Cy5 and Cy3 studied in this work, we can use the following model shown in Figure 2(a) to interpret the observed photophysics of cyanine dyes.…”

“…The results indicate that the single molecule fluorescence fluctuation and the resulting second autocorrelation analysis used in this work are powerful methods for determining the triplet state and isomerization dynamics, which could be the simple techniques and complementary to other spectroscopic techniques, such as fluorescence decay measurement and laser flash photolysis to study the photophysical processes [1,7,8] . …”

“…Much information is available about the photophysical and photochemical properties of cyanine dyes in the past years [1,[7][8][9][10] . When cyanine dyes are used in the study of protein-DNA, protein-protein interactions [11,12] , and DNA structure [3][4][5][6]13] , etc., the single molecule photophysical properties could be changed compared with free cyanine dyes after the cyanine dyes were covalently attached to those biomolecules [14,15] .…”

“…The occurrence of on-off blinking has attracted a lot of interests because the origin of off state may concern many causes, such as triplet state and isomerization. During single molecule measurements, isomerization (or torsional motion) is also a relaxation pathway in competition with intersystem crossing, where the isomerization will yield a compound with different absorption and emission properties, which is another reason for off state during single molecule measurements [1,7,8,19] . Therefore, the investigation on on-off blinking characteristics of single cyanine dyes will give information on the populations of the triplet state and dynamics of isomerization, which could be an alternative technique to the time-resolved spectroscopy such as laser photolysis and fluorescence decay measurements.…”

Photoinduced charge transfer in porphyrin-C60 oligomer

“…Phosphorescence measurement was carried on a setup developed in our laboratory [17]. Phosphorescence spectra were measured with a high-quality emission spectrophotometer (Jobin Yvon TRIAX 320) with highly sensitive LN-CCD as the detector (Jobin Yvon, CCD-3000V) combined with a high-speed rotated sector wheel for chopping the laser and emission beam in turn.…”