The longest characteristic time of rotational anisotropy decays (rotational cor-relation time)τrot for compoundsB(105), B⊥B (107) and BkB(116) was de-termined at different temperatures (figure 57). For B andBkB the anisotropy decays were fitted by an monoexponential fit (depolarization only due to rota-tion), whereas forB⊥Ba biexponential fit was used.
T = 30 °C B(105, black symbols) in DEHP at different temperatures. Black lines repre-sent the best monoexponential (B,BkB) and biexponential (B⊥B) fits. This figure was published in an article with the title "Bichromophoric Compounds with Orthogonally and Parallelly Arranged Chromophores Separated by Rigid Spacers".[160] Copyright Wiley-VCH Verlag GmbH & Co. KGaA. Re-produced with permission.
In order to calculate the effective hydrodynamic volumeVhfor each compound, the rotational correlation timeτrotwas plotted against η/kBT. Whereη is the viscosity of the solvent (DEHP)[161] at the corresponding temperatureTandkB is the Boltzmann constant (figure 58).
Effective hydrodynamic volumes 14.3
0 2 4 6 8 10 12
0 5 10 15
τrotns/
η/(kBT) / (1018 s m-3)
B B||B B T
B
Figure 58:Rotational (longest) characteristic time of anisotropy decaysτrotofB(105), B⊥B(107) andBkB(116) in DEHP as a function of solvent viscosity. The lines represent the best linear fits. This figure was published in an article with the title "Bichromophoric Compounds with Orthogonally and Parallelly Arranged Chromophores Separated by Rigid Spacers".[160]Copyright Wiley-VCH Verlag GmbH & Co. KGaA. Reproduced with permission.
According to the relation[162]
τrot = ηVh
kBT , (34)
the slope equalsVh. The resulting effective hydrodynamic volumesVhand radii of spheresrspherewith these volumes are given in table 13.
Table 13:Calculated hydrodynamic volumesVhand corresponding sphere radiirsphere.
compound Vh/nm3 rsphere/nm
B(105) 6.86 1.18
B⊥B(107) 12.0 1.42
BkB(116) 13.6 1.48
Figure 59 shows ground state energy minimized DFT structures of compounds B,B⊥BandBkBdrawn inside spheres which correspond to the hydrodynamic
Chapter 14 TIME-RESOLVED FLUORESCENCE ANISOTROPY EXPERIMENTS . . .
correlation times extracted from time-resolved anisotropy measurements are in good agreement.
Figure 59:Orthographic projections of DFT structures of B (105), B⊥B (107) and BkB (116) drawn inside spheres corresponding to the effective hydrody-namic volumes. This figure was published in an article with the title "Bichro-mophoric Compounds with Orthogonally and Parallelly Arranged Chro-mophores Separated by Rigid Spacers".[160] Copyright Wiley-VCH Verlag GmbH & Co. KGaA. Reproduced with permission.
15 Evidence for intramolecular EET by antibunching experiments
We also addressed intramolecular EET of bichromophores BkB (116) and B⊥B(107) by fluorescence antibunching experiments in solution, allowing to determine the number of emitters per bichromophore (see chapter 3.3).[111,115,163]
These experiments and the data processing were kindly carried out by Dr. Haisen Ta using a custom-made setup and custom-made software (see section 18 for de-tails).
With very short excitation pulses (∼70 psτf=1.0 ns), every chromophore can be excited only once and emit not more than one photon (antibunching). Mul-tiple photons detected within a short period of time (<25 ns), can result from either multiple fluorescent molecules diffusing through the detection volume or multiple emitters in the same molecule. The former contribution can be quan-tified by cross correlation of the two independent detection channels at large lag timet≈1 s (figure 60). Interestingly, the cross correlation has the same am-plitude as at t≈0 (i.e. the occurrence of multiple photon events is the same), indicating that none of the bichromophore compounds emits two photons at the same time. By fitting the measured autocorrelation function, we found that the numbers of emitters for compoundsB⊥BandBkBandBare all very sim-ilar (1.18±0.12, 1.00±0.05 and 1.22±0.08, respectively).[117] This means that these compounds behave as single quantum systems. Highly efficient EET with nearly quantitative quantum yield enabling singlet-singlet annihilation[117,164]is suggested as a most probable explanation (see section 3.3.5).
Chapter 15 EVIDENCE FOR INTRAMOLECULAR EET BY ANTIBUNCHING . . .
Correlation / (counts2 ns-2)Correlation / (counts2 ns-2)
Figure 60:Measured cross correlation functions of antibunching experiments.
Left: Magnification at zero lag time t≈0 (blue curve) and at large lag time t+∆t where ∆t was chosen to be 1 s (red curve); Right: Cross correlation function normalized by its value at t=∞; Dots correspond to the measured values of the cross correlation and solid lines correspond to fitting of the cross-correlation function to the experimental data (model functions 25 and 26). This figure was published in an article with the title
"Bichromophoric Compounds with Orthogonally and Parallelly Arranged Chromophores Separated by Rigid Spacers".[160] Copyright Wiley-VCH Verlag GmbH & Co. KGaA. Reproduced with permission.
Part III
Conclusion and Outlook
In this thesis a new approach for the design of bichromophoric systems with well-defined geometry was developed. As shown in figure 61, application of adamantane and 2-(2-adamantylidene)adamantane as rigid bridges between two chromophores led to assemblies with perpendicular and parallel oriented chromophores, respectively. Linking the chromophores by spiro junctions to both sides of the bridge provides rigid bichromophoric systems with fixed rel-ative orientation and precisely defined distance between the chromophores.
Figure 61:Adamantane and 2-(2-adamantylidene)adamantane as rigid linkers between two chromophores provide an access to assemblies with perpendicular and parallel arranged chromophores.
The fluorophores cannot be varied freely and need to meet several requirements:
First, they need to be capable of being attached to the adamantane scaffold. As shown in scheme 29, this was achieved by metal-halogen exchange of biaromatic halogenides24and25(chromophore precursors). The resulting alcohol was cy-clized under acidic conditions to give spiro compounds29or31.
X
Scheme 29:Attaching dye precursors24or25to adamantane derivatives23aand23b; Y=Br, I; R1-4=Ar, electron withdrawing/donating group.
For spectroscopic studies, a decent brightness (product of extinction coefficient and fluorescence quantum yield) is required. Furthermore, a pair of two fluo-rophores with spectrally separated absorption/emission bands is necessary for
their emission. A series of new fluorophores were developed in the course of im-proving their properties. Finally, a pair of "blue" and "red" fluorophores was ob-tained; these were used for the design of model compounds and bichromophores shown in figure 62.
Figure 62:Bichromophoric compounds with rigid linkers arranged in orthogonal (B⊥B, B⊥R) and parallel geometry (BkB,BkR), as well as model com-poundsBandR.
To characterize the photophysical properties of the fluorophores, model com-pounds B and R were synthesized (figure 62). Then, bichromophores with two identical (blue/blue) and different fluorophores (blue/red) were synthesized.
BichromophoresB⊥BandB⊥Rwith perpendicular arranged chromophores as well asBkBandBkRwith the parallel arrangement were prepared.
The novel bichromophoric systems are perfectly suited for the study of in-tramolecular electronic energy transfer (EET) between chromophores at well-defined geometry. The high rigidity and well-well-defined geometry of the systems could be advantageous relative to most systems reported up to now, which are less rigid and prone to rotation and bending of single bonds distorting the
molec-The common approach for explaining through-space EET is the Förster theory, where the EET is described as resonance energy transfer between two point-dipoles. According to Förster’s theory, the resonance energy transfer between perpendicularly oriented chromophores should not occur. However, an impor-tant requirement for this approximation is a sufficiently large distance between the fluorophores. This distance has to be much larger than the dimensions of the fluorophores. Therefore it becomes clear, that Förster’s theory may not cor-rectly describe the EET in the bichromophores presented here. The distances between the chromophores are about 0.8 and 1.3 nm for the perpendicular and parallel geometry, respectively. This separation equals roughly the dimension of the chromophores. Therefore it is no surprise to observe deviations from the FRET theory.
All studied bichromophoric molecules showed efficient EET, independently of the relative chromophore orientation. Steady-state absorption and emission spectroscopy of B⊥R and BkR revealed EET with near quantitative transfer efficiency due to the absence of the donor fluorescence. In bichromophoreB⊥B very fast EET was proven by time-resolved fluorescence anisotropy measure-ments. The results showed very fast anisotropy decay to a non-zero anisotropy value. This was explained by a fast back and forth energy transfer between two fluorophores. To address the energy transfer between the chromophores inBkB, fluorescence antibunching experiments were carried out. The results confirmed, that the bichromophores behave as single emitters, i.e. both parts of the same bichromophore never emit photons at the same time. We assume, that singlet-singlet annihilation (SSA) is accountable for this behavior. SSA provides evidence for fast intramolecular EET between two simultaneously excited chromophores.
Having in mind, that Förtster’s dipole-dipole approximation may not be valid at these short distances, it was examined whether FRET may explain the efficient EET observed for the bichromophores introduced here. The quantum yield, the fluorescence lifetime and the spectral overlap were taken from the model com-poundsBandR. To determine the orientation factorκ2, the relative orientation of the emission and absorption transition dipole moments and the distance be-tween the chromophores were determined using density functional theory (DFT) and time-dependend density functional theory (TD-DFT). The results are sum-marized in table 14.
Table 14:Calculated distance between chromophores |~RDA|, orientation factor κ2, Förster radius R0 and resulting FRET efficiency EFRET for bichromophores based on photophysical properties of model compounds B and R and DFT/TD-DFT calculations.
Compound κ2 |~RDA|/nm R0/nm EFRET
B⊥B(107) 6.0·10-4 0.84 0.85 0.51
BkB(116) 1.0 1.33 2.93 0.99
B⊥R(114) 1.0·10-4 0.84 0.77 0.38
BkR(118) 1.0 1.33 3.61 1.00
Geometries and orientation of transition dipole moments were calculated using DFT and TD-DFT methods, see chapter 19 for details.
Since all DFT studies were performed in gas-phase (neglecting any solvent-effects) the results should be interpreted with caution. Nonetheless, the results indicate that even at these short interchromophoric distances the FRET approx-imation gives a FRET efficiency for compoundsB⊥BandB⊥Rmuch smaller 1.
The calculation results do not completely explain the efficient EET in the bichro-mophores.
Additionally, the Förster point-dipole approximation may be not valid at these short distances due to multipole-multipole Coulombic interactions. At the same time, the classical Dexter mechanism seems to be unlikely due to the vanish-ingly small orbital overlap of the twoπ-systems, which are separated by a bulky saturated hydrocarbon spacer. However, the rigidity of the σ-framework may enhance a through-bond (superexchange) interactions. Finally, despite the rigid-ity of the adamantane linkers, deviations from the equilibrium geometry caused by vibrations in the excited state cannot be completely disregarded. To iden-tify the EET mechanism further experiments with bichromophoric compounds possessing longer rigid linkers are required.
Part IV
Experimental Part
16 General remarks
16.1 Solvents and reagents
The used solvents had pro analysi grade. Anhydrous solvents were dried over 3 Å or 4 Å molecular sieves. Commercially available substances were used with-out further purification. 2,6-adamantanedione was purchased from Ambinter (a trademark of Greenpharma S.A.S), Orléans, France.