2
J. A. Christensen, J. T. Fletcher / Tetrahedron Letters xxx (2014) xxx–xxx
NaN3
K2CO3
CuSO4
Na ascorbate
TMS
N
N
N
Br
N
+
I
N
t-BuOH/H2O
I
r.t., 24h
1 (92%)
N
N
N
N
N
N
Pd(PPh3)4
CuI
N
N
N
2
N
N
N
+
I
THF/TEA
50oC, 24h
2 (93%)
N
N
N
N
N
N
Pd(PPh3)4
CuI
N
N
N
N
N
+
N
2
I
THF/TEA
50oC, 24h
3 (50%)
Scheme 1. Preparation of sensors 2 and 3.
These sensors were originally designed as next generation ana-
Figures 1 and 2. As Ni(II) concentrations increased, a large hypso-
chromic shift and an emission intensity increase were observed
for each compound. Ratiometric comparisons of emission intensi-
ties for 2 (365/450 nm) and 3 (375/435 nm) were used to define
‘turn-on’ sensor responses. Ratiometric analysis summaries for all
analyte:sensor combinations assayed using 2 and 3 are shown in
Figures 3 and 4.
This method of data analysis shows clearly that the only ana-
lytes varying significantly from control responses are Ni(II), Cu(II)
and V(III). It is noteworthy that the positive ratiometric responses
generated for both Cu(II) and V(III) result from simple emission
logs of ethynylarene chemosensors operating via a conformational
restriction mechanism,5d,g,14 such that in the absence of analyte the
arene subunit would be capable of free rotation about its alkyne
bonds resulting in a baseline fluorescence emission signal. Upon
binding of analyte a rigidification of the central ethynylarene unit
would be enforced that results in an increase in fluorescence inten-
sity and a possible change in emission wavelength depending on
the conformation of the ethynylarene bonds in the analyte bound
state.13 Due to the distance between the chelating units in the sen-
sor, such rigidification would likely not be driven by cooperative
interaction between chelating units of the same molecule, but
rather by cooperative intermolecular binding of multiple sensor
molecules via analyte coordination.12b
quenching, which is
a significantly different spectroscopic
response than the blue-shifted signal intensification observed for
Ni(II).15 Based on these observations, it can be claimed that both
2 and 3 serve as selective fluorescence chemosensors for Ni(II)
under aqueous conditions.
Two different assays were performed to determine the revers-
ibility of Ni(II) binding with 2 and 3. Titration of the standard
assays for both 2 and 3 with Ni(II) using the competitive chelator
EDTA degraded the observed ratiometric signals back to baseline
levels.15 In addition, simple dilution of samples showing turn-on
responses from 10-fold to 1000-fold resulted in a return to baseline
levels.15 Each of these results indicates that the interactions of 2
and 3 with Ni(II) generating turn-on signals are reversible. It is
noteworthy that the turn-on signal for 2 persisted at significantly
Because the goal of this study was to identify sensors capable of
sensing aqueous solutions of metal cation analytes, it was impor-
tant to evaluate 2 and 3 in an aqueous environment. While each
compound is insoluble in water itself, both 2 and 3 can be pre-
dissolved in DMSO and diluted 1:1 or 1:2 into water without pre-
cipitating. Significant differences in the UV–visible absorbance and
fluorescence emission spectra for 2 and 3 are observed in compar-
ing DMSO and DMSO/water mixtures (Supplementary Material).
Under aqueous conditions the fluorescence emission for each
was significantly red shifted and quite broad relative to the DMSO
samples, while the absorbance spectra for each also varied signifi-
cantly. It is proposed that the differences in emission for 2 and 3
under varying solvent conditions are due to aggregate formation
promoted by the decreased solubility of these compounds in an
aqueous environment. Due to the variation in fluorescence emis-
sion promoted by environmental changes, these compounds were
considered attractive candidates for sensor applications.
High-throughput fluorescence screening1c,5g,14 was used to
examine the spectroscopic responses of these ethynylarenes in the
presence of various cations in DMSO/water solvent systems. DMSO
stock solutions of 2 and 3 were mixed with varying concentrations
of aqueous metal chloride salt solutions in 96 well plates. The
resulting solutions of 50 lM sensor mixed with 50–250 lM metal
cations in 1:1 DMSO/water and 1:2 DMSO/water for 2 and 3, respec-
tively were analyzed for fluorescence changes induced by the
increasing metal concentrations. In these assays, 2 was excited at
kmax = 290 nm and monitored from 320 nm to 550 nm and 3 was
excited at kmax = 320 nm and monitored from 350 to 600 nm.
Among the 22 metal cations evaluated in this study, both 2 and
3 displayed unique responses to the Ni(II) analyte, as illustrated in
Figure 1. Emission of 2 in response to increasing amounts of Ni(II) analyte in 1:1
DMSO/H2O solution.