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nitrilotriacetic acid residue, 152 nucleotides, 153 and sugars. 154 Most of them
show similar optical properties in aqueous solutions as their water-
insoluble counterparts in organic solvents. Despite the fact that most of
the reported water-soluble BDPs are the ones absorbing at rather short
wavelength ( < 650 nm, with a few examples of water-soluble BDPs or
aza-BDPs absorbing in red and near-IR), 134,141 the water-solubilizing
groups reported so far, in principle, can be used for long-wavelength
absorbing derivatives. Monovalent bioconjugatable BDP and aza-BDP
derivatives have been also prepared in straightforward manner. 155
4.2. Energy-transfer dyads for increasing the Stokes shift
of BDPs
The inherent optical property of BDPs that potentially may hamper their ap-
plication in vivo is their small Stokes shift (20-30 nm). This can be a potential
problem, given the relatively narrow absorption bands in BDPs and aza-
BDPs. Hence, to achieve efficient excitation, BDPs need to be excited very
close to their absorption maxima. While modifications of BDP structures
reported so far do not allow substantial increase of the Stokes shift, a potential
solution can be the assembling of two different BDP derivatives in energy-
transfer dyads. In an energy-transfer dyad, two chromophores are covalently
connected by a nonconjugated bridge so that each chromophore retains the
optical properties that it had as a monomer. 156-158 The excitation of the
chromophore absorbing at the shorter wavelength (donor) causes energy
transfer to the chromophore with the longer wavelength of absorption
(acceptor) and consequently emission of the acceptor. If the quantum
efficiency of energy transfer is high and there are no other competitive
processes (such as electron transfer), an energy-transfer dyad behaves as a
single chromophore with the excitation wavelength of the donor and the
emission wavelength of the acceptor (see Fig. 3.4 ).
The critical aspect in constructionof energy-transfer dyads is the linker con-
necting the donor and acceptor, which determines the mechanism of energy
transfer and thereby the choiceof the donor and the acceptor. In dyads inwhich
the donor and the acceptor are connected by a fully nonconjugated linker (such
as an alkyl or a peptide chain), the dominant mechanism of energy transfer is
through-space F¨rster resonance energy transfer (commonly referred to as a
FRET). Efficient FRET requires a large spectral overlap, that is, overlap be-
tween the emission band of the donor and the absorption band of the acceptor,
and thus limits the choice of both pairs of chromophores that fulfill this require-
ment. On the other hand, a conjugated linker that provides, to some extent,
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