Chemistry Reference
In-Depth Information
[95]. The attraction of DIBO is that its synthesis is extremely simple, it is nontoxic
and its hydroxyl can be employed to attach a variety of probes. Furthermore, it was
found that the rate of cycloaddition of DIBO can be enhanced by modifying the eight-
membered ring. For example, oxidation of the hydroxyl moiety to ketone resulted
in the compound that exhibits a fivefold increase in the rate of cycloaddition [96].
Keto-DIBO as well as its oxime analog possessed additional interesting properties
and it was found that the cyclooctynes were fluorescent whereas this was not the
case for the corresponding triazole products. This type of fluoro-switches can, for
example, be exploited for monitoring reactions in real time.
Several cyclooctynes have been described that exhibit even faster rates of cycload-
dition by introducing sp
2
-hybridized ring-atoms. For example, the groups of van Delft
[97] and Popik [98] independently reported a more reactive DIBO derivative that has
a nitrogen atom within the cyclooctyne ring. The cycloaddition reaction between aza-
dibenzocyclooctyne (DIBAC or ADIBO) and benzyl azide was found to be five times
faster compare to DIBO k
DIBAC
0.057 M
−
1
.s
−
1
.DIBAC
was also successfully employed for protein modification and surface functionaliza-
tion. Further enhancement of the reaction rate k
0.3 M
−
1
.s
−
1
;k
DIBO
≈
≈
0.96 M
−
1
.s
−
1
was achieved by
using BARAC [99], which is a dibenzo-fused lactam. Due to its high reactivity, very
low concentration (250 nM) of BARAC-Fluor (BARAC conjugated to fluorescein)
was required to image living cells metabolically labeled with azido-sialic acid. In an
attempt to render BARAC fluorogenic, it was modified by a coumarin moiety [100].
Although increase in fluorescence was observed upon reaction with an azide, the
low fluorescence quantum yield
≈
0.04 of the triazole product, combined with
a relatively high-energy wavelength of excitation (
F
≈
≈
300 nm) makes coumBARAC
unsuitable for biological imaging.
Recent DFT calculations by Goddard [101] have suggested that although the aryl
rings of DIBO promote the rate of cycloaddition by increasing the ring strain, the effect
is somewhat tempered by unfavorable steric interactions between the approaching
azide and the ortho-hydrogen atom of the aryl rings. Therefore, it was anticipated
that removal of an aryl ring of DIBO could lead to compounds with enhanced rates
of cycloaddition. Indeed, Bertozzi and coworkers demonstrated that DIFBO [102],
which is a hybrid of DIBO and DIFO, was found to be highly reactive and underwent
spontaneous homotrimerization in solution. For analytical purposes, DIFBO was
ingeniously stabilized as an inclusion complex with
-cyclodextrin, enabling long-
term storage and used as a reagent for rapid cycloaddition with azides (k
0.22
M
−
1
.s
−
1
). Unfortunately, to date no biological applications have been reported, most
probably due to its high instability. Furthermore, a monobenzylated cyclooctyne
(COMBO) [103] has been designed that exhibits similar reaction kinetics (k
≈
0.23
M
−
1
.s
−
1
) as DIFBO, but is chemically stable at room temperature over a 2-day
period, but quickly degrades at 37
◦
C (30% degradation after 9 hours). To underscore
its stability within complex biological environment, a COMBO-fluorescein conjugate
was successfully used for imaging azido-labeled sialic acids present on the surface
of living cells.
Although increasing the number of sp
2
-like centers within the cyclooctyne ring
has proved to be a successful strategy to enhance the rate of cycloaddition, it renders
≈
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