3
Table 1. Optical properties of compounds 1-7 in various solvents at 3-10 µM concentrations.
H-derivatives
OH-derivatives
5
NR2-derivatives
7a
214b (original)
neutral anionic
485
404
520
485
6
114a
314c
(original)
4
7b
(original)
neutral
anionic
561(56.5)
572(40)
22500
neutral
424(14)
564(27)
4000
anionic
Absa
Emb
368
427(-c)
490(55) 511(49.5)
476(-c)
670(4)
511(- c)
674(2)
466(15)
554(77)
11500
Water
EtOH
430(~10)
532(65)
638(5)
3000
564(5)
2500
c
c
c
Brightness
Absa
-
-
-
-
-
-
416
483
-
365
436(-c)
512
532
-
565(52.5) 430(14.5) 529(58) 496(46.5)
498(18)
650(19)
3500
510(26)
653(13)
3500
479(20)
543(65)
13000
Emb
426(~20)
510(30)
573(33)
17500
550(61)
9000
653(9)
5000
555(35)
16500
c
Brightness
Absa
-
-
d
360
425(19)
497(86)
16500
533
540
-
563(49.5) 422(17.5)
-
485(45)
552(48)
22000
487(21)
641(29)
6000
498(30)
646(20)
6000
402
469(73)
-
458(23)
532(80)
18500
d
CH3CN
EtOAc
Emb
420(55)
571(27)
13500
524(77)
13500
-
d
Brightness
Absa
-
362
-
d
428(19.5)
484(95)
18500
533
543
-
567(48.5) 429(17.5)
-
484(46.5)
538(67)
31000
486(22)
607(40)
9000
498(30)
617(31)
9500
410
465
-
467(23.5)
525(80)
19000
d
Emb
421(63)
-
574(19)
9000
511(88)
15500
-
d
Brightness
Absa
-
d
365
435(20)
483(98)
19500
529
541
-
567(50.5)
576(7)
3500
433(18)
504(94)
17000
-
489(52.5) 492(22.5) 508(31.5)
414
464
-
474(21)
523(86)
18000
d
Dioxane
Emb
422(78)
-
-
532(78)
41000
c
595(45)
10000
602(34)
10500
d
d
Brightness
-
peak maximum in nm (extinction coefficient in (mM cm)-1), peak maximum in nm (fluorescence quantum yield in %)), non-soluble enough, anionic
a
b
form cannot be obtained.
Solvatochromism was especially noticeable for 7 amine
Conclusion
derivatives. These compounds were also characterized by a
significant variation in FQY depending on the properties of the
In this paper, we proposed a new group of derivatives of a
solvent. The analysis of the behavior of these compounds using
conformationally locked GFP chromophore, containing
a
the Kamlet-Taft model (see ESI part 3) showed that their
solvatochromic properties are similar to the properties of other
previously obtained conformationally locked amino BDIs. Thus,
both compounds 7 demonstrate the dipole moment and acidity
increase upon excitation.
naphthalene residue. The novel dyes are characterized by a
noticeable red-shift of emission and absorption spectra in contrast
with the corresponding derivatives that contain only one benzene
ring. Also, all the obtained compounds are characterized by a
significant Stokes shift and most of them have no pH-dependent
transformation under physiological conditions. Thus, the
presented dyes are excellent candidates for application as novel
fluorescent tags, which can potentially be used for multicolor
labeling together with other dyes with small Stokes shift.
Compounds 7 are noteworthy as they demonstrate not only a
large Stokes shift but also a significant variation of FQY, which
can be used in actively developing fluorogenic labeling
technologies.
Previously, we suggested that this FQY variation can be
explained by the formation of an additional non-fluorescent
excited state characterized by a significant degree of charge
separation and planar geometry (PICT state). Formation of this
state (as well as FQY quenching) cannot be prevented by
conformational fixation of the benzylidene moiety, as it is not
accompanied by a loss of planarity. But for simple amino-BDI,
these changes could be partially blocked by the introduction of
rigid substituents at the nitrogen atom, for instance, by the
introduction of a pyrrolidine residue.17 However, the introduction
of this group did not significantly affect the variation of FQY in
the case of naphthalene derivatives – the characteristics of 7a and
7b derivatives were similar. The introduction of the pyrrolidine
fragment allowed shifting the spectral maxima to the long-wave
region but the spread of FQY over the solvents remained
practically unchanged. Such a variation can make it difficult to
use these compounds as common fluorescent dyes but enable
their use as fluorogenic dyes – compounds that have weak
fluorescence in the free-state but strongly fluoresce when binding
to a specific target.11
Acknowledgments
The authors gratefully acknowledge financial support from the
grant MK-1216.2019.3 of the President of the Russian
Federation.
References and notes
1. (a) Suzuki T, Matsuzaki T, Hagiwara H, Aoki T, Takata K. Acta
Histochem Cytochem. 2007; 40: 131-137; (b) Sahoo H. RSC Adv.
2012; 2: 7017-7029.
2. (a) Goncalves MST. Chem. Rev. 2009; 109: 190-212; (b) Christie
RM. Rev. Prog. Col. 1993; 23: 1-18.
3. (a) Yang Y, Lowry M, Xu X, Escobedo JO, Sibrian-Vazquez M,
Wong L, Schowalter CM, Jensen TJ, Fronczek FR, Warner IM,
Strongin RM. Proc. Natl. Acad. Sci. U. S. A. 2008; 105: 8829-834;
(b) Azuma E, Nakamura N, Kuramochi K, Sasamori T, Tokitoh N,
Sagami I, Tsubaki K. J. Org. Chem. 2012; 77: 3492-3500; (c)
Sibrian-Vazquez M, Escobedo JO, Lowry M, Fronczek FR, Strongin
RM. J. Am. Chem. Soc. 2012; 134: 10502-10508.
4. Chevalier A, Renard PY, Romieu P. Chemistry (Easton). 2014; 20:
8330-8337.
5. Ni Y, Zeng W, Huang KW, Wu J. Chem. Commun. 2013; 49: 1217-
1219.
6. Araneda JF, Piers WE, Heyne B, Parvez M, McDonald R. Angew.
Chem. Int. Ed. Engl. 2011; 50: 12214-12217.
The study of the spectra in water showed that acidification of
solutions of compounds 7 or basification of solutions of
compounds 4 and 5 is followed by a noticeable shift of the
maxima of absorption and emission. These shifts are explained
by the respective protonation of the amino group and
deprotonation of the phenol moiety. Deprotonation of phenols 4
and
5 led to a noticeable bathochromic shift and was
characterized by pKa 6.0 and 8.3, respectively. Protonation of 7a
and 7b led to a hypsochromic shift and occurred at a pH of ~3-4
(see ESI part 4, Table S4).