Fluorescent sensing of pyrophosphate and bis-carboxylates with charge neutral PET chemosensors

Article abstract of DOI:10.1021/ol026004l

(Matrix presented) The fluorescent photoinduced electron transfer (PET) chemosensors 2 and 3 were designed for the recognition of anions possessing two binding sides such as dicarboxylates and pyrophosphate; the anion recognition in DMSO takes place throu

Full text of DOI:10.1021/ol026004l

ORGANIC
LETTERS
2
002
Vol. 4, No. 15
449-2452
Fluorescent Sensing of Pyrophosphate
and Bis-carboxylates with Charge
2
†
Neutral PET Chemosensors
,‡
‡,§
‡
Thorfinnur Gunnlaugsson,* Anthony P. Davis, John E. O’Brien, and
‡
Mark Glynn
Department of Chemistry, Trinity College Dublin, Dublin 2, Ireland, and School of
Chemistry, UniVersity of Bristol, Cantock’s Close, Bristol BS8 1TS, UK
gunnlaut@tcd.ie
Received April 11, 2002
ABSTRACT
The fluorescent photoinduced electron transfer (PET) chemosensors 2 and 3 were designed for the recognition of anions possessing two
binding sides such as dicarboxylates and pyrophosphate; the anion recognition in DMSO takes place through the two charge neutral thiourea
receptor sites with concomitant PET quenching of the anthracene moiety. The anion binding of acetate, phosphate, and pyrophosphate to 2
1
and 3 was also evaluated by using H NMR in DMSO-d
6
.
3
Over the past few years, fluorescent and luminescent
chemosensors for the detection of cations have been suc-
cessfully developed.1,2 Conversely, the development of
optically based anion sensors has been less successful. Given
the important role of anions in biology, clinical diagnostics,
and environmental monitoring, the need for easily synthe-
sized fluorescent anion chemosensors is of great importance.
4,5
A variety of anion receptors have been reported, including
many capable of luminescent sensing. However, rather few
have the simplicity and accessibility which is ideally required
for practical devices.
We have been interested in the development of lumines-
cent chemosensors for the detection of cations, anions, and
neutral molecules. We are currently particularly interested
in developing luminescent anion chemosensors where the
anion recognition takes place at charge neutral recognition
6
†
Affectionately dedicated to Sigr u´ nar Ingibjargar G ´ı slad o´ ttur on the
7
occasion of her 70th birthday.
‡
Department of Chemistry, Trinity College Dublin.
School of Chemistry, University of Bristol.
§
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(
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1
1
0.1021/ol026004l CCC: $22.00 © 2002 American Chemical Society
Published on Web 06/25/2002

sites with concomitant changes in the photophysical proper-
ties of a lumophore by modulation of a Photoinduced
Electron Transfer (PET) mechanism. We have chosen to
thiourea moieties that can form hydrogen bonding complexes
with bis-anions. To the best of our knowledge, these
chemosensors are the first examples of charge neutral
fluorescent PET sensors that show ideal PET behavior for
bis-anions.
8
demonstrate such anion recognition of biologically relevant
-
-
2 4
anions such as H PO and AcO in DMSO by employing
simple thiourea and urea recognition sites, connected to an
anthracene fluorescent moiety by a covalent spacer. Here
the anion recognition takes place through hydrogen bonding
Sensors 2 and 3 (Scheme 1) can be described as being
designed as “receptor-spacer-fluorophore-spacer-recep-
9
between the thiourea hydrogens and the anion. Such
chemosensors should in principle show ideal PET behavior
upon anion recognition, i.e., only the quantum yield and the
lifetime of the excited-state emission should be modulated
Scheme 1. The Synthesis of PET Anion Chemosensors 2 and
3
10
upon anion recognition. In this letter we demonstrate such
PET fluorescent sensing of anions flanked with two binding
1
1
sites. Such fluorescence sensing is both exceptional and
of great physiological relevance since many dicarboxylates
are components of various metabolic processes, and pyro-
phosphate is the product of ATP hydrolysis under cellular
1
2
conditions. However, it has up to now been difficult to
4
,5
achieve without the use of structurally complicated hosts.
With this in mind we developed 2 and 3, which have two
(6) Liao, J. H.; Chen, C. T.; Fang, J. M. Org. Lett. 2002, 4, 561. Fabbrizzi,
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1
3
tor” conjugates where the anion recognition takes place at
the two-thiourea moieties. They are easily synthesized, and
simple modification to the thiourea moiety (by incorporating
aromatic or aliphatic electron withdrawing groups) can be
used to “tune” the anion sensitivity and selectivity, as the
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4
Chem. Soc., Perkin Trans. 2 2002, 141. Gunnlaugsson, T.; Mac Donaill,
D. A.; Parker, D. J. Am. Chem. Soc. 2001, 123, 12866. Gunnlaugsson T.;
Davis, A. P.; Glynn, M. Chem. Commun. 2001, 2556. Gunnlaugsson, T.
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acidity of the thiourea hydrogens is modulated. 2 and 3 were
synthesized in good yield (Scheme 1) from 9,10-diamino-
methylanthracene (1). The synthesis of this starting material
has been described previously in the literature, using Gabriel
synthesis. However, due to the insolubility of the bis-
phthalimide intermediate the yield of 1 was found to be
extremely poor. With this in mind we synthesized 1 using
an alternative method that involved the initial synthesis of
,10-bis-bromomethylanthracene in one step in 75% yield.
Accordingly, 1 was synthesized from this bis-bromide
intermediate with hexamethylenetetramine in anhydrous
1
4
(
8) Rurack, K. Spectrochim. Acta, Part A 2001, 57, 2161. Amendola,
V.; Fabbrizzi, L.; Mangano, C.; Pallavicini, P. Acc. Chem. Res. 2001, 34,
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4
1
(
15
9
A. D. J. Org. Chem. 2001, 66, 7313. B u¨ hlmann, P.; Nishizawa, S.; Xiao
K. P.; Umezawa, Y. Tetrahedron 1997, 53, 1647. Kelly R. R.; Kim, M. H.
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S.; Hamilton, A. D. J. Am. Chem. Soc. 1993, 115, 369. Dixon, R. P.; Geib,
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F.; Goswami, S.; Chang, S.-K.; Geib, S. J.; Hamilton, A. D. J. Am. Chem.
Soc. 1990, 112, 7393.
16
3
CHCl under inert atmosphere. This method gave 1 in 85%
yield as a crude product that could be used without further
purification. The two sensors 2 and 3 were subsequently
(
10) Nishizawa, S.; Kaneda, H.; Uchida T.; Teramae, N. J. Chem. Soc.,
Perkin Trans. 2 1998, 2325.
11) Mei M.; Wu, S. New J. Chem. 2001, 25, 471. Watanabe, S.; Higashi,
(
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Mu n˜ oz, M. C. Tetrahedron 1997, 53, 2629.
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994, 116, 9397.
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(
2450
Org. Lett., Vol. 4, No. 15, 2002

made by reacting 1 with phenyl and 4-(trifluoromethyl)-
phenylisothiocyanate, respectively, in dry CH Cl under
2
2
argon at room temperature. The resulting light-yellow
precipitate was collected by filtration, washed several times
with cold CH
Cl or CHCl . All products were analyzed with conventional
methods. The H NMR of 2 and 3 in DMSO-d
the high symmetry of the sensors, with only two sets of
aromatic signals and a single resonance for the -CH
2 2 2
Cl , and recrystallized from either hot CH -
2
3
1
7
1
6
indicated
2
-
spacer. For 2, the thiourea protons appeared as two reso-
nances at 9.68 and 8.38 ppm, respectively.
The sensitivity and the selectivity of these sensors toward
a series of mono- and bis-anions was evaluated by observing
the changes in their fluorescence emission spectra in DMSO
1
+
and in the H NMR upon anion titration (with (C
4 9 4
H ) N
(
TBA) salts) in DMSO-d . The fluorescence emission spectra
6
of 2 consisted of three bands at 409, 430, and 455 nm when
excited at 378 nm. Upon addition of monodentate anions
-
-
such as AcO and H
PO
2 4
the emission was ca. 70-95%
“
switched off” or quenched due to the formation of the
anion-receptor complex. No other spectral changes were
observed in the emission spectra, i.e., there was no evidence
of either exciplex or excimer emissions.10 Concurrently the
changes in the absorption spectra (peaks at 358, 378, and
Figure 1. The changes in the fluorescence emission spectra of 2
upon addition of pyrophosphate.
400 nm) of the anthracene moiety were only minor. Similar
results were observed for 3. This is a typical PET behavior
since the receptors are separated from the fluorophore by
observed in the emission spectra (Figure 1 for the titration
of 2 with pyrophosphate). For the two organic anions, the
fluorescence emission of 2 was “switched off” by ca. 70%
and 86% for glutarate and malonate, respectively ([anion]
the two -CH
2
- spacers; the only interaction between the
two moieties is via electron transfer. Upon addition of
-
-
spherical anions such as Cl and Br no significant quench-
ing was observed, ruling out quenching by the heavy atom
)
40mM in all cases). Similar reduction was seen in the
fluorescence emission of 3. The changes in the quantum
yields of fluorescence (Φ ) of 2 and 3 upon anion sensing
-
effect. However, F quenched the emission effectively
F
(
∼98%) due to its small size and high charge density. The
-
were measured in comparison with that of 9,10-dimethyl-
anthracene (9,10-DMA). For 2, these were measured to be
addition of AcO to a solution of 2 with a 40 mM
background concentration of Cl quenched the emission to
the same degree as seen previously for AcO , indicating that
the two receptors were selectively binding AcO over Cl .
Plotting the changes of the fluorescence intensity at 430 nm
as a function of pA (-log[anion]) gave, in all cases,
-
-
-
-
-
sigmoidal profiles (see Figure 2 for AcO ). However, for
both 2 and 3, these profiles changed over ca. 3-4 pA units.
This can be regarded as an indication of a possible 2:1
binding. This was further confirmed by observing the changes
1
5
in H NMR of the thiourea protons upon titration. With
-
-
either H
thiourea resonances were gradually shifted downfield by
2.5 ppm, confirming the formation of anion-receptor
complexes. Analysis of the changes in the “inner proton”
2
PO
4
or AcO (0 f 5-6 equiv of TBA salts), the
>
(
8.38 ppm) vs concentration showed 1:2 binding for both of
-
these anions as seen for H
2
PO
4
in Figure 3.
When 2 and 3 were titrated with TBA salts of the
biologically important bis-anions such as glutarate, malonate,
and pyrophosphate (with tris(tetrabutylammonium) hydrogen
pyrophosphate), the emission spectra were also quenched.
As before, no other significant spectral changes were
Figure 2. Fluorescence titration curve for 2 when measured at
(
17) Calculated for 2 (C32H24N4S2F6): C, 59.80; H, 3.76; N, 8.72.
430 nm (OD ) 0.1) in DMSO: malonate (b), pyrophosphate ([),
Found: C, 59.82; H, 3.76; N, 8.70. Calculated for 3 (C30H26N4S2): C, 71.12;
H, 5.17; N, 11.06. Found: C, 71.10; H, 5.17; N, 11.10. H NMR (400 MH,
1
glutarate (2), and acetate (9) .
DMSO-d6)for 2 and 3 are available as Supporting Information.
Org. Lett., Vol. 4, No. 15, 2002
2451

the quenching was even more efficient, being ca. 95% for 2
Figure 2) (Φ ) 0.001) and 90% (Φ ) 0.017) for 3. No
(
F
F
other spectral changes were seen in the emission spectra.
Plotting the emission changes at 430 nm vs pA gave
sigmoidal curves for all the bis-anions (Figure 3). Impor-
tantly, the emission is “switched off” over two pA units, for
pyrophosphate and malonate, indicating 1:1 binding and
simple equilibrium. For the larger glutarate anion, the
emission was “switched off” over ca. 3 pA units. From these
changes (Figure 2) the binding constant logâ of 3.74((0.05),
2.34((0.05), and 3.40((0.05) was determined for glutarate,
malonate, and pyrophosphate, respectively, for 2. For 3, these
values were found to be 3.07 ((0.05), 3.15((0.05), and 2.02-
((0.05) for pyrophosphate, glutarate, and malonate, respec-
tively. To investigate these binding interactions in greater
1
detail, we carried out a H NMR titration on 2 using
pyrophosphate. From these changes, Figure 3, a logâ of 3.81-
((0.05) was determined, with a clear 1:1 binding, which is
in good agreement with that seen above. For such recognition
to take place, the anion would have to bridge the anthracene
moiety. Although this did not affect the absorption spectra
of 2 to any great extent (see Supporting Information), then
upon closer examination an isosbestic point was observed
Figure 3. 1H NMR titration of 2 with phosphate (red) and
pyrophosphate (blue) in DMSO-d
6
.
.012 and 0.011 for glutarate and malonate, respectively.18
0
We believe that this relatively high degree of fluorescence
quenching is due to the increase in the reduction potential
of the thiourea receptor moieties after anion recognition. This
affects the rate of the electron transfer from the HOMO of
the receptor to the excited state of the fluorophore, i.e., ∆GET
becomes more negative upon anion recognition and hence
more thermodynamically favorable. This causes the emission
to be “switched off”. We were unable to demonstrate this
by measuring the changes in the redox potential for the
receptor since the thiourea was irreversibly oxidized. How-
1
at ca. 406 nm. However, the H NMR spectra of 2 showed
somewhat greater changes for the anthracene resonances
1
upon titration with pyrophosphate than that seen in the H
-
NMR titration of H
2
PO
4
. We are currently investigating
these anion-binding features in greater detail, and carrying
out modifications on these chemosensors to further enhance
the bis-ion selectivity and sensitivity.
ever, Φ
receptor sites, gave a Φ
substantially larger than that of 2, Φ
.11, in DMSO. This implies that PET is active prior to the
F
measurements of 9,10-DMA, which lacks the two
) 0.87 in DMSO, which is
) 0.047, and 3, Φ )
Acknowledgment. We thank Enterprise Ireland, Kinerton
Ltd., and Trinity College Dublin for financial support and
Dr. Hazel M. Moncrieff for helpful discussions.
F
F
F
0
anion recognition, but becomes even more so after anion
recognition. The addition of 10 mM of AcO or pyrophos-
phate did not affect the Φ of 9,10-DMA. For pyrophosphate
F
Supporting Information Available: Synthesis of 1, 2,
and 3 and absorption spectrum of 2 upon titration with
pyrophosphate. This material is available free of charge via
the Internet at http://pubs.acs.org.
-
(18) Barnes, R. L.; Briks, J. B. Proc. R. Soc. London, Ser. A 1966, 291,
5
70.
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