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Photophysical behaviors of single fluorophores localized on zinc oxide nanostructures.

Fu Y, Zhang J, Lakowicz JR - Int J Mol Sci (2012)

Bottom Line: In this report we studied photophysical behaviors of single fluorophores in proximity to zinc oxide nanostructures by single-molecule fluorescence spectroscopy and time-correlated single-photon counting (TCSPC).Single fluorophores on ZnO surfaces showed enhanced fluorescence brightness to various extents compared with those on glass; the single-molecule time trajectories also illustrated pronounced fluctuations of emission intensities, with time periods distributed from milliseconds to seconds.The fluorescence fluctuation dynamics were found to be inhomogeneous from molecule to molecule and from time to time, showing significant static and dynamic disorders in the interfacial electron transfer reaction processes.

View Article: PubMed Central - PubMed

Affiliation: Center for Fluorescence Spectroscopy, Department of Biochemistry and Molecular Biology, School of Medicine, University of Maryland School of Medicine, 725 Lombard Street, Baltimore, MD 21201, USA; E-Mails: jianzhang@umaryland.edu (J.Z.); jlakowicz@umaryland.edu (J.R.L.).

ABSTRACT
Single-molecule fluorescence spectroscopy has now been widely used to investigate complex dynamic processes which would normally be obscured in an ensemble-averaged measurement. In this report we studied photophysical behaviors of single fluorophores in proximity to zinc oxide nanostructures by single-molecule fluorescence spectroscopy and time-correlated single-photon counting (TCSPC). Single fluorophores on ZnO surfaces showed enhanced fluorescence brightness to various extents compared with those on glass; the single-molecule time trajectories also illustrated pronounced fluctuations of emission intensities, with time periods distributed from milliseconds to seconds. We attribute fluorescence fluctuations to the interfacial electron transfer (ET) events. The fluorescence fluctuation dynamics were found to be inhomogeneous from molecule to molecule and from time to time, showing significant static and dynamic disorders in the interfacial electron transfer reaction processes.

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Schematic diagram of a model of photoinduced processes in a dye-deposited ZnO system. The ZnO energy levels are shown at the left, the relevant energy levels for the dye molecule are shown on the right. The optical transition between the ground electronic state (S0) and the excited electronic state (S1) is illustrated by the arrow labeled as hν.
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f3-ijms-13-12100: Schematic diagram of a model of photoinduced processes in a dye-deposited ZnO system. The ZnO energy levels are shown at the left, the relevant energy levels for the dye molecule are shown on the right. The optical transition between the ground electronic state (S0) and the excited electronic state (S1) is illustrated by the arrow labeled as hν.

Mentions: Single molecules were studied using a stage-scanning confocal microscope equipped with time-correlated single photon counting (TCSPC) module. Figure 2a,b show typical 10 μm × 10 μm images for single Cy5 molecules on a coverslip and a ZnO nanoparticle covered surface, respectively. The individual bright spots in the images are attributed to the single-molecule emission as evidenced by several criteria [37–39]. (1) The emission spot is about the same size order as the diffraction-limited size of the laser focus, which is about 300 nm in this confocal setup; (2) The signal brightness is comparable as expected for single-molecule emission considering the properties of the molecule, the excitation intensity, and the efficiency of optical filters; (3) The density of the spots increases proportional to the concentration of dye in the solution. Figure 2 shows characteristic fluorescence intensity time traces of single Cy5 molecules on bare glass (Figure 2c) and on ZnO substrate (Figure 2d–g), respectively. Typical single-molecule trajectories show constant fluorescence emission before photobleaching and the intensity drops to the background abruptly in a single step. Many traces of single fluorophores on ZnO exhibit pronounced fluctuations between bright and dark states over the course of their trajectories accompanied by a significant increase in emission rate as shown in Figure 2d–g. In contrast to the relatively continuous emission on glass surface in Figure 2c, increase in the number of “on/off” events was observed occasionally in the trajectories (Figure 2d–g) with “off” time ranging from sub-seconds to seconds. The fluorescence blinking observed from molecules on bare glass is contributed to temporary trapping of the excited states in long-lived triplet states of the dye molecules [27]. We believe that the single-molecule fluorescence intensity fluctuation is not due to triplet state blinking during the experiment. Under ambient conditions, the time scale of the triplet blinking is approximately shorter than 1 ms [27] which is clearly distinguished from the fluorescence intensity fluctuation time of subseconds to seconds as depicted in Figure 2d–g, the triplet blinking is thus not responsible for the observed long “off” times. Additionally, fluorescence counts depicted are integrated in 5 ms binning time, the fast fluorescence intensity fluctuations or blinking of the dye molecules (on sub-millisecond levels) are averaged out herein, as we observed a relatively steady fluorescence intensity level as illustrated in Figure 2c. It has been reported that excited organic fluorophores on a semiconductor coated surface show rapid reversible electron transfer and exhibit similar dramatic fluctuation behaviors [5–7,24,29]. The occurrences of the longer “off” time are clearly related to the proximity of ZnO surface and we assume that the stochastic fluctuation behaviors such as those in Figure 2d–g are the results of electron transfer (ET) processes to the ZnO nanostructures: electron injection from the fluorophore into the conduction band of the semiconductor leads to the loss of fluorescence, and charge recombination leads to the return of fluorescence [5,24]. A similar model of the photophysical processes may be represented by Figure 3. The forward electron transfer (FET) kinetics in various dye-semiconductor systems have been studied with rates in the femtoseconds to several hundred picoseconds range, the injected electron is then localized on the ZnO semiconductor surfaces followed by a backward electron transfer (BET) process. Electron injection from the fluorophore into the conduction band of the ZnO leads to the loss of fluorescence, and charge recombination leads to the return of fluorescence. The long “off” periods in the fluorescence trajectory is herein the result of electron transfer process quenching the fluorescence, and the “on” state is due to natural fluorescence emission cycles.


Photophysical behaviors of single fluorophores localized on zinc oxide nanostructures.

Fu Y, Zhang J, Lakowicz JR - Int J Mol Sci (2012)

Schematic diagram of a model of photoinduced processes in a dye-deposited ZnO system. The ZnO energy levels are shown at the left, the relevant energy levels for the dye molecule are shown on the right. The optical transition between the ground electronic state (S0) and the excited electronic state (S1) is illustrated by the arrow labeled as hν.
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC3472795&req=5

f3-ijms-13-12100: Schematic diagram of a model of photoinduced processes in a dye-deposited ZnO system. The ZnO energy levels are shown at the left, the relevant energy levels for the dye molecule are shown on the right. The optical transition between the ground electronic state (S0) and the excited electronic state (S1) is illustrated by the arrow labeled as hν.
Mentions: Single molecules were studied using a stage-scanning confocal microscope equipped with time-correlated single photon counting (TCSPC) module. Figure 2a,b show typical 10 μm × 10 μm images for single Cy5 molecules on a coverslip and a ZnO nanoparticle covered surface, respectively. The individual bright spots in the images are attributed to the single-molecule emission as evidenced by several criteria [37–39]. (1) The emission spot is about the same size order as the diffraction-limited size of the laser focus, which is about 300 nm in this confocal setup; (2) The signal brightness is comparable as expected for single-molecule emission considering the properties of the molecule, the excitation intensity, and the efficiency of optical filters; (3) The density of the spots increases proportional to the concentration of dye in the solution. Figure 2 shows characteristic fluorescence intensity time traces of single Cy5 molecules on bare glass (Figure 2c) and on ZnO substrate (Figure 2d–g), respectively. Typical single-molecule trajectories show constant fluorescence emission before photobleaching and the intensity drops to the background abruptly in a single step. Many traces of single fluorophores on ZnO exhibit pronounced fluctuations between bright and dark states over the course of their trajectories accompanied by a significant increase in emission rate as shown in Figure 2d–g. In contrast to the relatively continuous emission on glass surface in Figure 2c, increase in the number of “on/off” events was observed occasionally in the trajectories (Figure 2d–g) with “off” time ranging from sub-seconds to seconds. The fluorescence blinking observed from molecules on bare glass is contributed to temporary trapping of the excited states in long-lived triplet states of the dye molecules [27]. We believe that the single-molecule fluorescence intensity fluctuation is not due to triplet state blinking during the experiment. Under ambient conditions, the time scale of the triplet blinking is approximately shorter than 1 ms [27] which is clearly distinguished from the fluorescence intensity fluctuation time of subseconds to seconds as depicted in Figure 2d–g, the triplet blinking is thus not responsible for the observed long “off” times. Additionally, fluorescence counts depicted are integrated in 5 ms binning time, the fast fluorescence intensity fluctuations or blinking of the dye molecules (on sub-millisecond levels) are averaged out herein, as we observed a relatively steady fluorescence intensity level as illustrated in Figure 2c. It has been reported that excited organic fluorophores on a semiconductor coated surface show rapid reversible electron transfer and exhibit similar dramatic fluctuation behaviors [5–7,24,29]. The occurrences of the longer “off” time are clearly related to the proximity of ZnO surface and we assume that the stochastic fluctuation behaviors such as those in Figure 2d–g are the results of electron transfer (ET) processes to the ZnO nanostructures: electron injection from the fluorophore into the conduction band of the semiconductor leads to the loss of fluorescence, and charge recombination leads to the return of fluorescence [5,24]. A similar model of the photophysical processes may be represented by Figure 3. The forward electron transfer (FET) kinetics in various dye-semiconductor systems have been studied with rates in the femtoseconds to several hundred picoseconds range, the injected electron is then localized on the ZnO semiconductor surfaces followed by a backward electron transfer (BET) process. Electron injection from the fluorophore into the conduction band of the ZnO leads to the loss of fluorescence, and charge recombination leads to the return of fluorescence. The long “off” periods in the fluorescence trajectory is herein the result of electron transfer process quenching the fluorescence, and the “on” state is due to natural fluorescence emission cycles.

Bottom Line: In this report we studied photophysical behaviors of single fluorophores in proximity to zinc oxide nanostructures by single-molecule fluorescence spectroscopy and time-correlated single-photon counting (TCSPC).Single fluorophores on ZnO surfaces showed enhanced fluorescence brightness to various extents compared with those on glass; the single-molecule time trajectories also illustrated pronounced fluctuations of emission intensities, with time periods distributed from milliseconds to seconds.The fluorescence fluctuation dynamics were found to be inhomogeneous from molecule to molecule and from time to time, showing significant static and dynamic disorders in the interfacial electron transfer reaction processes.

View Article: PubMed Central - PubMed

Affiliation: Center for Fluorescence Spectroscopy, Department of Biochemistry and Molecular Biology, School of Medicine, University of Maryland School of Medicine, 725 Lombard Street, Baltimore, MD 21201, USA; E-Mails: jianzhang@umaryland.edu (J.Z.); jlakowicz@umaryland.edu (J.R.L.).

ABSTRACT
Single-molecule fluorescence spectroscopy has now been widely used to investigate complex dynamic processes which would normally be obscured in an ensemble-averaged measurement. In this report we studied photophysical behaviors of single fluorophores in proximity to zinc oxide nanostructures by single-molecule fluorescence spectroscopy and time-correlated single-photon counting (TCSPC). Single fluorophores on ZnO surfaces showed enhanced fluorescence brightness to various extents compared with those on glass; the single-molecule time trajectories also illustrated pronounced fluctuations of emission intensities, with time periods distributed from milliseconds to seconds. We attribute fluorescence fluctuations to the interfacial electron transfer (ET) events. The fluorescence fluctuation dynamics were found to be inhomogeneous from molecule to molecule and from time to time, showing significant static and dynamic disorders in the interfacial electron transfer reaction processes.

Show MeSH
Related in: MedlinePlus