1358
these results support the scheme shown in Figure 1b as a major
¹
reaction mechanism. Furthermore, probe 1 reacted with OCl
efficiently and quickly (Figure S214), thus seeming suitable as
¹
an OCl-sensing probe. Unfortunately, however, the solubility
of probe 1 in water was too low (0.73 mM at maximum in
phosphate buffer pH 7.4) for its application in further biological
studies.
To improve solubility, we prepared probe 2, in which the
p-amino group of 1 was monoalkylated by a triethylene glycol
unit. During the derivatization study of 1, we found that
N-monoalkylation did not affect the reactivity between the
¹
p-aminophenyl alkyl ether and OCl, whereas N-bisalkylation
suppressed the reactivity completely (data not shown). There-
¹
fore, probe 2, which is soluble in water up to 15 mM, was OCl
¹
reactive. The reactivity of probe 2 with OCl, which was
analyzed using HPLC, was very high and resulted in 80 and
100% consumption of 2 after incubation with only 1 and 2
¹
equivalents of OCl, respectively. On the other hand, the probe
¹
2 was almost intact after incubation without OCl, suggesting
the stability of the probe under physiological conditions and
¹
good reactivity to OCl.
¹
With a water-soluble and OCl-reactive probe in hand, we
Figure 2. (a) 19F NMR spectra of probe 2 and trifluoroethanol (100
¯M). (b) 1H and 19F chemical-shift-selective imagings (11.7 T) of probe 2
and trifluoroethanol (10 mM each). Samples were dissolved in phosphate
buffer (pH 7.4, 100 mM) containing 150 mM NaCl and 0.1% DMF.
moved on to the visualization of OCl using 19F MRI, which
¹
was the aim of the present work. 19F NMR analyses of the
authentic samples, probe 2 and the predicted product trifluoro-
ethanol, gave 19F signals at ¹75.1 and ¹77.7 ppm, respectively
(Figure 2a). The observed 19F chemical-shift difference between
the probe and the product was 2.6 ppm, which was sufficient to
visualize each compound separately using 19F chemical-shift-
selective MRI. Figure 2b depicts phantom images (11.7 T) of
¹75.1 ppm (probe 2) but produced no 19F signal at ¹77.7 ppm
(trifluoroethanol), suggesting that NO reacts with probe 2 but
does not undergo o-dealkylation via an ipso-substitution path-
way.
1
¹
these compounds. H MRI visualized both samples because of
Because of the OCl-specific production of trifluoroethanol,
¹
the presence of H2O (left panel in Figure 2b). In contrast, probe
2 and the product (trifluoroethanol) were visualized separately
using 19F chemical-shift-selective imaging (second-right and
right panels in Figure 2b, obtained based on probe 2- and
trifluoroethanol-selective 19F pulse frequencies, respectively).
Then, we applied probe 2 to the detection and imaging of
as described above, probe 2 yielded targeted imaging of OCl
(Figure 3b). 19F chemical-shift-selective imaging (trifluoroeth-
anol 19F selective) gave a clear signal for probe 2 only in the
¹
presence of OCl (bottom panel in Figure 3b). The specificity of
the reaction was high; the presence of other ROS or RNS
produced no such signals. These results show clearly that the
designed compound 2 functions as a 19F MRI probe for the
¹
¹
OCl in 19F MR modality. After addition of OCl, the 19F signal
¹
of probe 2 (¹75.1 ppm) shifted to a new peak at ¹77.7 ppm,
highly specific detection and imaging of OCl.
¹
¹
corresponding to trifluoroethanol, in a clear OCl-dose-depend-
In conclusion, we designed an OCl-sensing 19F MRI
ent manner (Figure S314). The comparison of 19F NMR peak
integrals allowed us to calculate that probe 2 (100 ¯M) produced
trifluoroethanol with a reaction yield of 56% via a reaction with
probe. The advantages of this probe were at least threefold.
The first advantage was reactivity. The probe, which contained a
p-aminophenyl alkyl ether scaffold, reacted expeditiously with
¹
¹
2 equivalents of OCl. Because of the efficient probe-to-product
OCl to produce trifluoroethanol with a high conversion yield.
¹
conversion, this OCl-dose-dependent consumption of probe 2
The second advantage was specificity. The probe produced
¹
and generation of trifluoroethanol, i.e., the presence or absence
trifluoroethanol only after reaction with OCl. The third
¹
of OCl, was well visualized using 19F chemical-shift-selective
advantage was its applicability to 19F MRI. Because of the
imaging (Figure S414). These data indicate the utility of
sufficient 19F chemical-shift change concomitant with probe-to-
¹
¹
compound 2 as an OCl-imaging probe in 19F MR modality.
product conversion, OCl was detected and visualized using 19
F
In addition to the high reactivity and good conversion yield
obtained, probe 2 also exhibited high specificity for OCl.
chemical-shift-selective imaging. To the best of our knowledge,
this is the first MRI probe that yields OCl imaging with a high
¹
¹
HPLC analyses revealed that probe 2 remained intact after
reaction with most of the biologically important ROS (H2O2,
specificity among a variety of ROS and RNS. Our next
challenge is to adapt the probe to in vivo applications, e.g.,
evaluation, optimization, and improvement of the biodistribution
or biokinetics of the probe. Along these lines, and in addition to
the 19F MRI application described here, 19F MR spectroscopy
using a much larger voxel, which could provide the possibility
for probe/product ratiometric analysis, may also be a promising
target of this probe. Further work is now underway in our
laboratory.
•¹
•
ROO•, O2 , and OH) and reactive nitrogen species (RNS;
¹
¹
ONOO ), with the exception of OCl and NO (Figure S514).
However, and interestingly, the analysis of the reaction of probe
2 with ROS or RNS in 19F NMR modality showed that probe
2 produced a 19F NMR peak at ¹77.7 ppm, corresponding to
¹
trifluoroethanol, only after reaction with OCl (Figure 3a).
Reaction with NO decreased the intensity of the 19F signal at
Chem. Lett. 2011, 40, 1357-1359
© 2011 The Chemical Society of Japan
Doura, Tomohiro
