water molecules may orient themselves to optimise their interactions with the
luminophore, there can be a significant shift in the maxima for light absorption
and light emission (Stokes shift). Indeed, a dye molecule is a good reporter on the
microenvironment [1]. In the gas phase where the ions are isolated, the Stokes shift
is expectedly smaller than in solution phase, which is also seen experimentally [2].
Measuring the spectrum of light emitted (i.e., dispersed fluorescence) from
isolated ions has one important benefit compared to measuring the action spectrum:
The experiment does not rely on the absorption of more than one photon, and
kinetic shifts do not need to be considered as ionic dissociation is not sampled. Of
particular relevance, spectroscopy of large protein ions can be done; action spectroscopy is limited to multiply charged anions where it is possible to sample
electron detachment (see Chap. 8 by Antoine and Dugourd). The difficulty, of
course, is to detect enough photons that are emitted in all directions from a low
number of ions (see instrumental setup description in Chap. 3 by Wyer) [3].
Experiments are normally done on mass-selected trapped ions, either in linear
quadrupole, Paul or Penning ion traps [4–7]. The collection angle is often low
due to poor accessibility to the trapped ions, and sensitive light detectors must be
used. Scattered light from the laser or ambient light reduces the signal-to-noise
ratio. Detection efficiencies of a few percent have been reported, which are enough
in some cases to produce beautiful fluorescence spectra, and even time-resolved
light emission has been followed on the nanosecond time scale [4, 8–10]. Fluorescence lifetimes seem to be longer for gaseous ions than for solvated ones [10].
Spectra have been recorded for several biomolecular ions, e.g., xanthene-based
rhodamine dyes [2, 3, 6, 10], fluorescein and derivatives [11, 12], and non-covalent
host–guest complexes [8].
It should be mentioned that light emission or direct internal conversion to the
electronic ground state are not the only pathways after photoexcitation. Indeed,
Kappes and co-workers [13] showed that continuous photoexcitation of rhodamine
6G cations led to the disappearance of fluorescence due to trapping of all the ions in
long-lived (seconds), dark triplet states. They found that the ground-state ions could
be recovered from quenching the triplet state in interactions with molecular oxygen
(triplet-state molecule), turning fluorescence back on when these were photoexcited.
Finally, information on the emitted photon can also be obtained indirectly:
Williams and co-workers [14] used mass spectrometry to “weigh” the photon that
is emitted after UV photoexcitation of hydrated protonated proflavine ions. Fewer
water molecules evaporate on average if an ion has emitted light compared to the
number that is lost following non-radiative population of the electronic ground state
(either directly by internal conversion or via a triplet state by intersystem crossing).
In other words, the energy of the emitted photon is not available for water loss.
Simply from the difference in the number of evaporated water molecules and the
known water binding energies, the energy of the emitted photon can be estimated!
The branching ratio provides the fluorescence quantum yield. Furthermore, as the
authors pointed out: “A key advantage of this indirect detection method is that all
dissociation products resulting from emission are observed, irrespective of the
direction of which the photon is emitted.” The collection efficiency is 100 %.
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S.B. Nielsen
luminophore, there can be a significant shift in the maxima for light absorption
and light emission (Stokes shift). Indeed, a dye molecule is a good reporter on the
microenvironment [1]. In the gas phase where the ions are isolated, the Stokes shift
is expectedly smaller than in solution phase, which is also seen experimentally [2].
Measuring the spectrum of light emitted (i.e., dispersed fluorescence) from
isolated ions has one important benefit compared to measuring the action spectrum:
The experiment does not rely on the absorption of more than one photon, and
kinetic shifts do not need to be considered as ionic dissociation is not sampled. Of
particular relevance, spectroscopy of large protein ions can be done; action spectroscopy is limited to multiply charged anions where it is possible to sample
electron detachment (see Chap. 8 by Antoine and Dugourd). The difficulty, of
course, is to detect enough photons that are emitted in all directions from a low
number of ions (see instrumental setup description in Chap. 3 by Wyer) [3].
Experiments are normally done on mass-selected trapped ions, either in linear
quadrupole, Paul or Penning ion traps [4–7]. The collection angle is often low
due to poor accessibility to the trapped ions, and sensitive light detectors must be
used. Scattered light from the laser or ambient light reduces the signal-to-noise
ratio. Detection efficiencies of a few percent have been reported, which are enough
in some cases to produce beautiful fluorescence spectra, and even time-resolved
light emission has been followed on the nanosecond time scale [4, 8–10]. Fluorescence lifetimes seem to be longer for gaseous ions than for solvated ones [10].
Spectra have been recorded for several biomolecular ions, e.g., xanthene-based
rhodamine dyes [2, 3, 6, 10], fluorescein and derivatives [11, 12], and non-covalent
host–guest complexes [8].
It should be mentioned that light emission or direct internal conversion to the
electronic ground state are not the only pathways after photoexcitation. Indeed,
Kappes and co-workers [13] showed that continuous photoexcitation of rhodamine
6G cations led to the disappearance of fluorescence due to trapping of all the ions in
long-lived (seconds), dark triplet states. They found that the ground-state ions could
be recovered from quenching the triplet state in interactions with molecular oxygen
(triplet-state molecule), turning fluorescence back on when these were photoexcited.
Finally, information on the emitted photon can also be obtained indirectly:
Williams and co-workers [14] used mass spectrometry to “weigh” the photon that
is emitted after UV photoexcitation of hydrated protonated proflavine ions. Fewer
water molecules evaporate on average if an ion has emitted light compared to the
number that is lost following non-radiative population of the electronic ground state
(either directly by internal conversion or via a triplet state by intersystem crossing).
In other words, the energy of the emitted photon is not available for water loss.
Simply from the difference in the number of evaporated water molecules and the
known water binding energies, the energy of the emitted photon can be estimated!
The branching ratio provides the fluorescence quantum yield. Furthermore, as the
authors pointed out: “A key advantage of this indirect detection method is that all
dissociation products resulting from emission are observed, irrespective of the
direction of which the photon is emitted.” The collection efficiency is 100 %.
106
S.B. Nielsen