Photoreversible fluorescence modulation of a rhodamine dye by supramolecular complexation with photosensitive cyclodextrin

Article abstract of DOI:10.1002/anie.200701396

(Chemical Equation Presented) A complex light switch: Photoreversible fluorescence modulation was achieved by supramolecular complexation of a guest rhodamine molecule (RhB) with spiropyran-modified cyclodextrin (see picture). Upon UV irradiation, the chromophore within the cyclodextrin cavity can transfer its energy to the spiropyran moiety, while upon visible-light irradiation, no energy transfer occurs. Thus, light can be used to turn "on" and "off" the fluorescence emission of a dye inside the cyclodextrin cavity.

Full text of DOI:10.1002/anie.200701396

Angewandte
Chemie
DOI: 10.1002/anie.200701396
Fluorescence Modulation
Photoreversible Fluorescence Modulation of a Rhodamine Dye by
Supramolecular Complexation with Photosensitive Cyclodextrin**
Shuizhu Wu,* Yulan Luo, Fang Zeng, Jian Chen, Yanan Chen, and Zhen Tong
Materials with properties that can be optically modulated are
of high interest in many scientific areas, including biotech-
nology and optics.[1] Photosensitive—in particular photochro-
mic—compounds can be used to achieve this purpose, as they
can be converted reversibly by light between two states with
different spectroscopic properties.[2–5] This attribute can then
be used to modulate or switch various functions at the
molecular or supramolecular level by using light as a trigger.
Spiropyran, a promising photochromic compound, undergoes
reversible structural transformations in response to external
inputs, such as light.[4,6,7] This special property of the
spiropyran molecules was used to construct complex, opti-
cally controlled integrated logic gates and molecular switches
composed of dyad or triad molecules with spiropyran as one
of their units.[8–10] In this dyad or triad method, the modulation
of various photonic or optoelectronic processes was usually
realized by fluorescence quenching through energy transfer.
In this approach to a light-controlled molecular switch or
logic gate, several issues still need to be addressed: Firstly, the
specific fluorophore must be covalently linked to the spiro-
pyran molecule (or linked to it through a spacer) to form the
dyad or the triad; hence, in this approach, only one dyad or
triad can be used for a specific application. Secondly, self-
quenching is still observed as a result of the self-aggregation
of the fluorophore moieties. Thirdly, the dyads or triads are
usually hydrophobic, which limits their applications in hydro-
philic environments; in addition, they lack the necessary
biocompatibility for applications in the fields of biotech-
nology.
realized. This strategy is easily achievable, and the guest
rhodamine molecules can be readily replaced by other
chromophore compounds. In addition, the self-quenching of
RhB can be decreased by supramolecular complexation. The
strategy could provide a facile method for reversible fluores-
cence modulation, controlled by light, for a wide range of
chromophores, which is essential in applications such as
reversible bioimaging. At the same time, the chromophore
complexes will have the necessary biocompatibility and
hydrophilicity for applications in the field of biotechnology
because of the presence of the oligosaccharide cyclodextrin.
Photosensitive b-cyclodextrin was synthesized and based
on mass spectrometry (MS), high-performance liquid chro-
matography (HPLC), NMR spectroscopy, and elemental
analysis (see the Supporting Information), it was estimated
that two spiropyran moieties were present per b-cyclodextrin
molecule; the compound was thus designated as CDSP-2. The
structure and reversible fluorescence modulation of the
supramolecular complex formed by CDSP-2 and RhB is
shown in Scheme 1.
The spiropyran molecules can adopt one of two stable
states, namely, the open-ring state, which is known as the
protonated merocyanine (McH) form (upon UV light irradi-
ation), and the closed-ring state, which is known as the spiro
(SP) form (upon visible-light irradiation).[19,20] The as-pre-
It is well known that cyclodextrin (CD) can form
supramolecular inclusion complexes with small-molecule
chromophore compounds that fit into its 5–8 Š cavity;[11–13]
in this way, the fluorescence intensity, biocompatibility, and
photostability of the guest chromophore compounds can be
enhanced.[14,15] Herein, we synthesized photosensitive cyclo-
dextrin (CDSP) by covalently attaching spiropyran moieties
onto b-CD. Rhodamine B[16–18] (RhB) was used as a model
fluorophore to form a supramolecular complex with the
CDSP, and photoreversible fluorescence modulation was thus
[*] Dr. S. Wu, Y. Luo, Dr. F. Zeng, J. Chen, Y. Chen, Dr. Z. Tong
Department of Polymer Science & Engineering
South China University of Technology
Guangzhou 510640 (China)
Fax: (+86)20-8711-4649
E-mail: shzhwu@scut.edu.cn
[**] This work was supported by NSFC (Project No. 50573023) and
NCET.
Scheme 1. Supramolecular complex (composed of photosensitive b-
cyclodextrin and RhB) and photoreversible fluorescence modulation.
The position of RhB—included in the b-cyclodextrin cavity—was
estimated according to 2D-NMR measurements.
Supporting information for this article is available on the WWW
under http://www.angewandte.org or from the author.
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pared CDSP-2 solution in the dark (that is, the SP form of the
photosensitive cyclodextrin) exhibited an absorption band at
about 340 nm. After UV irradiation (at 365 nm), a new
absorption band at 550 nm appeared as a result of the
formation of the McH form (see the Supporting Information).
The formation of a complex between RhB and the CDSP-
2 was verified by 2D NMR experiments (see the Supporting
Information). Rotating-frame Overhauser-effect spectrosco-
py (ROESY) of an equimolar mixture of photosensitive b-CD
and T-shaped guest RhB molecules displays clear nuclear
Overhauser effect (NOE) correlations between the H-3 and
H-5 protons of b-CD and the methyl protons of the
diethylamino groups in RhB as well as NOE correlations
between the same protons of b-CD and the aromatic protons
of the diethylaminophenyl group in RhB. Since both the H-3
and H-5 protons are located inside the b-CD cavity, and the
H-5 protons are found near the primary (narrow) side of this
cavity while the H-3 protons are located near its secondary
(wide) side, these NOE correlations indicate that the RhB
molecule is accommodated inside the b-CD cavity.
The complexation between RhB and the photosensitive b-
CD was further demonstrated by means of absorption and
fluorescence spectra of RhB in the absence and presence of
CDSP-2 (see the Supporting Information). Complexation of
RhB by CDSP-2 causes a sharpening and a slight bath-
ochromic shift of the absorption band. The latter effect is
characteristic for the immersion of the guest in a less polar
environment. The shift in fluorescence is not pronounced,
which suggests that the relaxation of the dye through geo-
metrical and solvent effects is smaller, as expected for
inclusion in a less polar and more confined environment.[14]
However, at the same concentration of RhB, the fluorescence
intensity of the complex is stronger than that of a pure RhB
solution, which is probably because the supramolecular
complexation between RhB and CDSP-2 reduces the self-
quenching of RhB resulting from self-aggregation. A widely
used method[21] was employed to determine the complexation
stoichiometry, and a complexation ratio of 1:1 for RhB and
CDSP-2 was found (see the Supporting Information).
The fluorescence modulation for the stoichiometric RhB–
CDSP-2 complex upon exposure to UV or visible light is
illustrated in Figure 1. After irradiating the complex solution
with UV light (at 365 nm, 12 W) for five minutes, it was
immediately moved into the fluorescence spectrometer to
perform the emission measurements (at an excitation wave-
length of 546 nm); in this case, the characteristic fluorescence
emission of RhB (at 589 nm) was significantly quenched.
However, after irradiation with visible light (at 460 nm, 15 W)
for ten minutes, the intensity of this fluorescence signal was
recovered (see Figure 1a). During the fluorescence measure-
ments, 546-nm light was used to excite the RhB molecules;
this light could also convert the McH forms of spiropyran into
SP forms. However, the rate of the ring-closure reaction from
McH to SP (on the ten-minute scale)[22] is much lower than
that of the energy-transfer process (on the nanosecond scale).
In a one-minute fluorescence measurement carried out under
weak irradiation (at 546 nm), only a small portion of McH
spiropyran could be converted into SP spiropyran. The
remaining McH form of spiropyran quenched the RhB
Figure 1. a) Fluorescence-emission spectra of the RhB–CDSP-2 solu-
tion (in ethanol/DMSO 4:1, v/v; concentration: 3”10À5 m, slit: 2.5 nm,
lex =546 nm) . b) Photographs of the RhB–CDSP-2 complex solution
after: I) ten minutes of visible-light irradiation and II) five minutes of
UV irradiation.
emission, while the SP form caused a background fluores-
cence (Figure 1a, sample II).
In Figure 1b, the appearance of the RhB–CDSP-2 solu-
tion after visible-light irradiation is clearly different from that
after UV irradiation. Bright fluorescence can be seen in the
former case under ambient light (which includes the excita-
tion wavelength of RhB).
The reversible nature of the fluorescence modulation in
the RhB–CDSP complexes upon exposure to alternating
cycles of UV and visible light is illustrated in Figure 2. In an
ethanol/dimethyl sulfoxide (DMSO) solution, five minutes of
UV irradiation and ten minutes of visible-light irradiation can
reversibly turn “on” and “off” the fluorescence of RhB.
From the above results, it is clear that the closed-ring form
of the spiropyran moieties caused no change in the fluores-
cence of RhB, while the open-ring (McH) form did. The
energy of the first-excited-singlet state of RhB was estimated
to be 2.18 eV from the average frequencies of the absorption
and emission bands. Similar calculations, based on absorption
and emission maxima,[23] gave first-excited-singlet-state ener-
gies of 3.65 eV (lmax = 340 nm) for the SP-form spiropyran
moieties of CDSP-2, and of 2.10 eV (lmax = 552 nm, lem
=
630 nm) for the McH-form moieties. These results suggest
that energy transfer from RhB to the SP form of CDSP-2 is
impossible, while transfer to the McH form of the compound
may occur. Therefore, no fluorescence quenching of RhB
could be observed before ring opening of the spiropyran
moieties. However, upon UV illumination, these moieties
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Chemie
Experimental Section
b-CD, N,N’-dicyclohexylcarbodiimide (DCC), 4-(dimethylamino)pyr-
idine (DMAP), RhB, tetra-n-butylammonium hexafluorophosphate
(TBAHFP), and 3-iodopropanoic acid were purchased from Sigma–
Aldrich. 2,3,3-trimethylindolenine was received from Acros Organics.
N,N-dimethylformamide (DMF), DMSO, ethyl methyl ketone, eth-
anol, and methanol were analytically pure solvents (and were distilled
before use).
During the synthesis process, all the reaction vessels were
wrapped with aluminum foil to ensure that the reaction proceeded
in the dark. The carboxy-containing spiropyran l-(b-carboxyethyl)-
3’,3’-dimethyl-6-nitrospiro(indoline-2’,2
(SpCOOH) was synthesized according to the literature method.[26]
2,3,3-trimethylindolenine (0.06 mol), 3-iodopropanoic acid
[2H-1]
benzopyran)
Figure 2. Fluorescence-emission intensity of the RhB–CDSP-2 solution
(in ethanol/DMSO 4:1, v/v; concentration: 3”10À5 m, slit: 2.5 nm,
lex =546 nm) recorded at 589 nm immediately after several UV and
visible-lightirradiation cycles.
(0.06 mol), and ethyl methyl ketone (5 mL) were heated under
nitrogen at 1008C for three hours. The resulting solid was dissolved in
water, and the solution was washed with chloroform. Evaporation of
water gave l-(b-carboxyethyl)-2,3,3- trimethylindolenine iodide (75%
yield). This iodide (0.04 mol), 5-nitrosalicylaldehyde (0.04 mol), and
piperidine (0.04 mol) were dissolved in ethyl methyl ketone, and the
solution was heated under reflux for three hours. After allowing the
solution to stand over night, the product precipitated as a yellow
crystalline powder, which was collected by filtration and washed with
methanol to give the product SpCOOH (78% yield).
CDSP-2: SpCOOH (8 mmol) and b-cyclodextrin (3 mmol) were
added to anhydrous DMF (30 mL) in the presence of DCC
(0.021 mol) and DMAP (0.0021 mol) and stirred at 258C for 24 h.
The mixture was then filtered, and the filtrate was precipitated in cold
methanol. Afterwards, the product was dissolved in DMF and re-
precipitated in methanol (several times). Then, the product was
washed with large amounts of deionized water, and the material was
further purified by means of silica-gel chromatography [using ethyl
acetate/petroleum ether (2:8, v/v) and then acetonitrile/water (10:1,
v/v) as eluents] (28% yield).
adopted the open-ring (McH) form, thus making energy
transfer from the RhB molecules to the spiropyran centers
more efficient; this resulted in an clear fluorescence quench-
ing of rhodamine.
There are several requirements on the energy-transfer
process:[24] In this case, there is an excited-state energy-level
match between RhB and the open-ring spiropyran moieties,
and a spectral overlap between the fluorescence emission of
RhB (lem = 589 nm, from 570 to 650 nm) and the absorption
of the open-ring spiropyran moieties (lmax = 552 nm, from 500
to 610 nm). In addition, the distance between RhB (which is
located inside the CDSP-2 cavity) and the open-ring spiro-
pyran moieties is estimated to be 1.1–1.4 nm based on the
bond lengths, the position of the RhB molecule included in
CDSP-2, and the height of the CD molecule; this distance is
within the Förster radius.[24] All these conditions can meet the
requirements for a fluorescence resonance energy transfer
(FRET) process.[24] To investigate whether the fluorescence
quenching is a result of electron transfer, the electrochemical
potentials of CDSP-2 were measured by means of cyclic
voltammetry. No differences were observed in the oxidation
or reduction waves when the SP-form spiropyran moieties
were photoisomerized into the McH form (see the Supporting
Information), which suggests that the reduction and oxidation
potentials were not affected by the photoinduced structural
changes. These results are consistent with previous reports on
similar spiropyran derivatives.[23,25] Therefore, fluorescence
quenching of RhB through electron quenching is unlikely; we
believe that it could probably be the result of a FRET process
instead.
1H NMR, 13C NMR, and 2D-NMR spectra were recorded on a
Bruker Avance 400 MHz NMR spectrometer. UV/Vis spectra were
recorded on a Hitachi U-3010 UV/Vis spectrophotometer. Fluores-
cence spectra were recorded on a Hitachi F-4500 fluorescence
spectrophotometer. The cyclic-voltammetric experiments were car-
ried out with a CHI 660 setup (CH Instruments Inc.).
For the fluorescence and UV/Vis absorption measurements, RhB
was dissolved in ethanol/DMSO (4:1, v/v) and mixed with an
equimolecular amount of photosensitive cyclodextrin in ethanol/
DMSO (4:1, v/v). The mixture was stirred for 48 h in the dark. The
solvent was then evaporated, and the mixture was washed with
deionized water and dried under vacuum at 658C for 100 h. The
resulting powder was dissolved in deuterated DMSO for the 2D-
NMR measurements.
Received: March 31, 2007
Revised: May 25, 2007
Published online: August 7, 2007
Keywords: chromophores · cyclodextrins · energy transfer ·
.
fluorescence · supramolecular chemistry
In summary, we can use light to reversibly switch “on” and
“off” the fluorescence emission of a rhodamine dye by
including the dye molecules in the cavity of a spiropyran-
modified b-cyclodextrin. Energy transfer could be the reason
for fluorescence quenching by the McH form of CDSP-2. This
study could open up possibilities for achieving facile fluores-
cence modulation—controlled by light—for various chromo-
phore compounds.
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