A blue shift of the emission peak of my compound upon the addition of quencher was observed. I guess there is an electron transfer between it and quencher. But I don't know how to explain this blue shift. Anybody could give me some suggestions?
P.S.: There is no change in UV absorbance curve of the compound upon addition of quencher.
Could the quencher alter the polarity of the solvent leading to a blue shift, and not act to quench the emission of your compound?
The fluorescence spectrum may depend on the presence of quencher for a variety of reasons but in general this requires the presence of more than one emitting species. Even in the case of a single fluorophore in the presence of the excited-state solvent relaxation one may observe the blue shift of the fluorescence band upon addition of a dynamic quencher which shortens the excited-state lifetime and therefore gives more weight to the emission from only partially relaxed excited-states (which are higher in energy than totally solvent-relaxed states).
The maximum used quencher concentration is 1.0e-4 M. And this amount of quencher results in 14 nm blue shift of the emission maximum of my compound. Both blue shift and emission intensity quenching occur simultaneously. I prefer Aleksander's opinion that there is a process which can compete with the solvation dynamics. Does anybody know the rate of solvation dynamics of compound in DMF? Several 10 ps or several 100 ps?
You should consider the possibility that you have two populations emitting at different wavelengths, where one population is less available to the quencher e.g. multimers.
My hypothesis could perhaps be investigated by use of combinations of excitation and emission spectra
Excitation spectra recorded at different emission wavelengths, e.g. at your two peaks or broader apart, could look different and thereby confirming two populations. You could also look at emission spectra excited at different wavelengths. Perhaps in combination with varying the fluorophore concentration you should be able to confirm or rule out this hypothesis.
If the static quenching arises from ground-state complex formation, we could make a conclusion that the wavelength of maximum emission intensity of pure complex is shorter that of the fluorophore. In this case, you should record a UV-vis absorption spectrum of fluorophore-quencher mixture to confirm whether it is a real ground-state complex formation.
If the static quenching is interpreted as the result of the sphere-of-action model, I have no idea about how the blue shift can happen.
In case of the dynamic quenching, it is impossible that the biomolecular quenching constant can be larger the diffusion controlled value. If you can be sure that the quenching is caused by dynamic quenching, probably it means the relaxation processes , especially solvation relaxation, of your excited fluorophore is quite slow, which can be comparable with diffusion coefficient of the quencher in that solvent. I think this could only happen when the viscosity of your solvent is very high or the temperature of solution is very low.
Hi. I was getting crazy, it took me several days to answer this question, but I finally got it now.
Actually, this blue shift comes from the fact that the absorption of your quencher is A>0.1, inducing a photo-reabsorption process.
This leads to a coupling between the absorption of your quencher and the emission of your sample. If we approximate these spectra by gaussians, we can simulate such an interaction (y = \left |A \cdot e^{\frac{(x-m^{2})}{\sigma^{2}}} - B \cdot e^{\frac{(x-n^{2})}{\sigma^{2}}} \right |).
You will find as an attachement the simulated spectra.
Dear Yin Li,
For the generation of this document, Lucas Fortier used "Grapher" application, available on Mac OS. Gnuplot can also easily generate this type of file !
Hi Yin Li ,
A quencher typically plays a role to diminish the fluorescence. Quenchers typically does not shift the emission spectra. Depends on your situation, can you really use the word "quencher" to describe the compound which interfere with the emission of your fluorophore. This situation looks like the the addition of the compound (which you refer as the quencher) tends to generate some environmental changes which could hypsochromically shift the emission of your fluorophore. For example, some fluorophores can have tautomer forms which would have the same absorption profile.
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