Reversible photoswitching of dye-doped core-shell nanoparticles

Article abstract of DOI:10.1039/c1cc14800a

We present a simple and versatile mechanism for the reversible photoswitching of dye-doped core-shell nanoparticles. Photochromic dithienylethenes are incorporated into the outer shell, close enough to the dyes entrapped in the core to efficiently quench them by energy transfer when photoconverted with UV light. The emission can be switched back on by irradiation with λ > 450 nm.

Full text of DOI:10.1039/c1cc14800a

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Reversible photoswitching of dye-doped core–shell nanoparticlesw
Damiano Genovese,a Marco Montalti,a Luca Prodi,*a Enrico Rampazzo,a Nelsi Zaccheroni,a
b
b
b
b
¨
Oliver Tosic, Kai Altenhoner, Florian May and Jochen Mattay
Received 3rd August 2011, Accepted 2nd September 2011
DOI: 10.1039/c1cc14800a
We present a simple and versatile mechanism for the reversible
photoswitching of dye-doped core–shell nanoparticles. Photochromic
dithienylethenes are incorporated into the outer shell, close enough to
the dyes entrapped in the core to efficiently quench them by energy
transfer when photoconverted with UV light. The emission can be
switched back on by irradiation with k 4 450 nm.
ca. 7 nm, even though the effective hosting region is probably
thinner, since hydrophobic molecules are expected to stay far from
the external water environment. Such guest molecules are thus
hosted within a very small region, close to each other and to the
core-embedded dyes so that energy transfer processes can occur.
We thought then to exploit this innovative architecture to
realize a very versatile nanometric photoswitch of simple
preparation. We have inserted into the outer shell of the
nanoparticles a suitable photochromic compound, whose
ability as an energy acceptor for the dyes buried in the silica
core depends on its (open or closed) form.
The possibility of switching fluorescence signals on and off is
the basis for most advanced imaging techniques based on
fluorescence. Most super-resolution microscopy techniques
(STORM, PALM, SOFI)1–4 are based on a common idea:
using information stored in several different images to create
one upsized image. Algorithms thus extract details from every
image of a sequence to reconstruct super-resolved images. If the
object under investigation is identical in all frames, collecting
more frames and/or with a longer acquisition time can only
result in sharper images, but resolution will not be pushed
beyond the diffraction limit. Such a limit can be overtaken
only if an ensemble of emitters, though not moving within a
certain number of frames, changes its global appearance by
partially turning on and off intermittently, showing all its
sub-diffraction features within the frame series. In this framework,
switchable fluorescent probes are now a major issue, since their
brightness and the signal difference between the on and off states
determine the performance of super-resolution techniques.
Among the different families of photochromic compounds,
dithienylethenes are widely studied because of their favourable
properties such as high fatigue resistance and thermal
stability.9 The photocyclization reaction leading to the close-
ring isomer is usually triggered by UV-light with wavelengths
below 350 nm, while the ring opening reaction is carried out
with visible light (4450 nm). In our experiments, we used the
dithienylethene derivative PS,10,11 in its open (PS-o) and
closed (PS-c) forms (Fig. 1). The photoconversion from
PS-o to PS-c leads to several changes in the UV bands and,
more importantly, to a new broad absorption band in the
visible region, centred at 630 nm, displaying an interesting
molar extinction coefficient (e = 11 800 MÀ1 cmÀ1, Fig. 2).
The two forms of PS are well soluble in acetonitrile and other
organic solvents, but they are insoluble in water.
We have recently reported on the synthesis and characterization
of a new family of silica core/polyethylene glycol (PEG) shell
nanoparticles,5–7 featuring extremely interesting properties: high
brightness, emission wavelength spanning all visible and
near-infrared spectral regions, water solubility, low toxicity,
and functional groups at the external shell for selective targeting
of biomolecules.6 Furthermore, we have also shown that the
polymeric shell is able to reversibly host apolar molecules.8
Hosted dyes experience a significant spatial confinement
because of the small structure of the outer shell. DLS, TEMw
and AFM5 measurements assign to this shell a thickness of
We prepared NPs covalently doped with R (a Rhodamine B
derivative, Fig. 1) 0.25% mol/mol vs. TEOS (R@NPs), using
direct micelles of Pluronic F127 in water as nanometric
templates.8 These nanoparticles are stable for months and
no dye leakage is observed. R@NPs present absorption and
emission spectra analogous to the ones of R in solution and
a fluorescence quantum yield of 0.35.8 R was chosen as the
doping agent because of the large match between its emission
and the absorption band of PS only in its close-ring form, as
shown in Fig. 3. If PS is added to a suspension of R@NPs
10À7 M under physiological conditions (PBS 1 Â buffer) from
a concentrated acetonitrile solution, it is rapidly included into
the NPs’ shell, as demonstrated by ultrafiltration experiments.8
In fact, if the suspension of NPs and PS is centrifuged through
a 100 kD cut-off membrane, PS is found only in the retentate
together with the NPs, and not in the filtrated solution
(see Fig. S4 in the ESIw). Moreover, PS still undergoes
photoconversion under irradiation with UV and visible light
(Fig. 3) in the presence of NPs, although with a slightly
a Dipartimento di Chimica ‘‘G. Ciamician’’,
`
Universita degli Studi di Bologna, via Selmi 2, 40126 Bologna, Italy.
E-mail: luca.prodi@unibo.it; Fax: +39 051 2099456;
Tel: +39 051 2099481
b Organic Chemistry I, Department of Chemistry, Bielefeld University,
Universitatsstr. 25, 33615 Bielefeld, Germany
¨
w Electronic supplementary information (ESI) available: Details on
the synthesis of PS and of the nanoparticles, ultrafiltration, dynamic
light scattering, TEM, photophysical measurements, and fluorescence
microscopy. See DOI: 10.1039/c1cc14800a
c
This journal is The Royal Society of Chemistry 2011
Chem. Commun., 2011, 47, 10975–10977 10975

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Fig. 3 Absorption and emission spectra of a suspension containing
R@NPs and PS-o (dark lines, solid and dashed respectively), or
R@NPs and PS-c, i.e. after UV irradiation (gray lines, solid and
dashed respectively).
PS into its close-ring form. The emission spectrum does not change
in shape, but its intensity is drastically reduced by roughly 95%
(Fig. 3, gray dashed line), proving that an efficient energy transfer
process has been triggered on. The inverse process takes place if the
same solution is irradiated with visible light (l 4 450 nm). The
absorption band centred at 630 nm decreases, and R@NPs
emission is fully recovered to its initial intensity. This process could
be repeated many times (420) with only minor changes in the
initial and final states (Fig. 4). These cycles were performed in a low
volume cuvette (50 mL), which ensured that the incident excitation
beam irradiated the whole sample. It has to be noted here that the
observed emission intensity decrease cannot be attributed to inner
filter effects if not for a negligible amount (o5%),12 because of the
low absorbance of PS-c under our experimental conditions
(Fig. 3). The microscopy experiments (see below), in which the
optical path is very short, further support this conclusion.
Fig. 1 Molecular structures of the rhodamine B derivative (R) and the
dithienylethene in the two forms (PS-o and PS-c) obtained by VIS and
UV irradiation, respectively. The cartoon represents the photoswitchable
energy transfer process, and consequent emission quenching, within the
proposed core–shell nanostructure.
It is important to stress that, although the absorption coefficient
of PS is not very high, the efficiency of the energy transfer process is
close to unity, indicating that a very high electronic communication
can be established among the dyes buried in the silica core and
the species present in the outer PEG shell, allowing the design
of systems suitable for many research needs.
Fig. 2 Absorption spectra of PS-o (dark line) and PS-c (gray line) in
acetonitrile.
The adduct of R@NPs and PS was also tested using a
wide-field fluorescence microscope. We used a continuous
reduced efficiency and photostability compared to the ideal
behaviour observed in acetonitrile solutions (Fig. 2).
In a first experiment we wanted to prove that irradiation with
UV light (320 nm) could lead to the photoconversion of PS in
its close-ring form and consequently to the quenching of the
emission of R, while visible light irradiation (l 4 450 nm) could
reset the system back to the initial fluorescent configuration. We
thus irradiated and monitored through absorption and emission
spectroscopy a volume of 2.5 mL of a water suspension of
1.6 Â 10À7 M R@NPs upon addition of 10 ml of 6.4 Â 10À4 M
acetonitrile solution of PS added in the dark, corresponding to
an average of ca. 13 PS per NP. The resulting absorption
spectrum, before irradiation, is the sum of the spectra of
R@NPs and PS-o. The emission spectrum is identical in shape
and magnitude to that of R@NPs before addition of PS-o,
confirming that FRET does not occur at this stage.
Fig. 4 Real time R@NPs fluorescence (lemission = 590 nm) monitored
during several photoconversion cycles. Data collected on a quartz
cuvette for low volume samples (optical path length 0.3 cm, cuvette
volume 50 mL). Irradiation at 320 nm (black dots and lines) and at
580 nm with a Xenon lamp (red dots and lines).
After 1 min irradiation at 320 nm, the appearance of an
absorption band at 630 nm indicates the photoconversion of
c
10976 Chem. Commun., 2011, 47, 10975–10977
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In this context, a further improvement of the system is
possible, in particular enhancing the absorption coefficient
(to obtain a more favourable Forster radius), the efficiency
¨
and robustness of the photoswitch in a water-based environ-
ment, a research that is in progress in our laboratories.
Worth noticing is the nature of the nanoparticles, which are
water soluble, non-toxic and membrane-permeable, and thus
they respond to the most stringent criteria for in vivo and in vitro
investigations. Moreover, the photoswitching mechanism that
we propose is highly versatile, and many different fluorescent
and photochromic dyes can be advantageously used, allowing
fine tuning of the spectral and photophysical properties of the
nanostructure. This strategy can thus be able to produce
structures having the features required to meet different
research needs.
We gratefully acknowledge financial support by the Italian
Ministry of University and Research (MIUR, grant PRIN
2009Z9ASCA), by the program NanoSci-E+ (financed
project "INOFEO"), by the German Ministry of Research
and Education (BMBF, grant 13N9234) and by Bielefeld
University.
Fig. 5 Wide-field image of R@NPs and PS deposited on a glass
coverslip before (a) and after (b) UV irradiation. (c) Normalized
fluorescence intensity of all particles in the field of view upon turning
on and off the UV irradiation, under constant 488 nm laser excitation.
For single particles, see Fig. S5 (ESIw).
Notes and references
laser excitation at 488 nm (filtered to obtain a 10 mW power)
and a cooled CCD camera to image the nanoparticles
dispersed on a glass coverslip from a drop of water suspension
containing R@NPs of 5 Â 10À9 M and PS of 1 Â 10À4 M. The
irradiation from the top of the glass slide with a UV lamp
resulted in a strong emission quenching, as clearly visible in
Fig. 5b. Once the UV irradiation was turned off, the
continuously running 488 nm excitation reset the system back
to the initial fluorescent configuration by photoconverting
PS-c in its uncoloured open-ring form PS-o. Fig. 5 displays
five cycles of these quenching steps followed by fluorescence
recovery.
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In conclusion, we proved by means of fluorescence spectroscopy
and wide-field fluorescence microscopy that the luminescence of
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in the core. This makes the emission quenching strongly
dependent on the state of PS, thus switchable with light.
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c
This journal is The Royal Society of Chemistry 2011
Chem. Commun., 2011, 47, 10975–10977 10977