Water-soluble phosphonate-substituted BODIPY derivatives with tunable emission channels

Article abstract of DOI:10.1021/ol200969r

Water-soluble BODIPY dyes have been readily obtained by introduction of phosphonate fragments either on the boron for the green and yellow emitting dyes or on the side chain for the red emitting dyes. Hydrolysis of the phosphonate is realized at the end o

Full text of DOI:10.1021/ol200969r

ORGANIC
LETTERS
2011
Vol. 13, No. 12
3072–3075
Water-Soluble Phosphonate-Substituted
BODIPY Derivatives with Tunable
Emission Channels
Thomas Bura and Raymond Ziessel*
ꢀ
Laboratoire de Chimie Organique et Spectroscopies Avancees (LCOSA), UMR 7515
au CNRS, Ecole de Chimie, Polymeres, Materiaux de Strasbourg (ECPM),
ꢁ
ꢀ
25 rue Becquerel, 67087 Strasbourg, Cedex 02, France
ziessel@unistra.fr
Received April 13, 2011
ABSTRACT
Water-soluble BODIPY dyes have been readily obtained by introduction of phosphonate fragments either on the boron for the green and yellow
emitting dyes or on the side chain for the red emitting dyes. Hydrolysis of the phosphonate is realized at the end of the reaction sequence and
allows isolation of the targets by precipitation. All these novel dyes are soluble and fluorescent in water with quantum yields in the 23À59% range
and emission wavelength spanning from 667 to 509 nm.
Over the past decade there has been a renewed interest in
the design of highly luminescent dyes for use as probes in
biological systems.1 The major drawbacks with neutral
organic dyes are their low solubility in polar solvents and
their tendency to decrease solvent contact by forming
nonemissive aggregates.2 Furthermore, most of these
dyes display fluorescence features sensitive to environ-
mental factors such as solvent polarity or association
with hydrophobic biomolecules or with organized mat-
ter such as micelles, vesicles and thin films.3 Numerous
fluorescent molecules (e.g., fluorescein, rhodamine,
acridine, anthracene, phenanthrene, pyrene, quinoline,
benzofuran, dansyl, naphthalimide, squaraines, cyanines
and indacene) display a rich variety of optical pro-
perties but challenges remain concerning: (i) water
solubility, (ii) facility of synthesis and purification pro-
cedures, (iii) control of nonradiative decay pathways
and (iv) conversion of often fragile organic frameworks
to more robust species.
To engender water solubility and other requirements
imposed for use in bioassays, sophisticated functionaliza-
tion of basic dye skeletons is usually necessary. Currently,
propelled by scientific reviews highlighting their exceptional
chemical stability and promising spectroscopic features,
there is a major interest in the engineering of difluoro-
bora-diaza-s-indacene (BODIPY) dyes.4 Advantages of
the basic BODIPY core are its absorptive properties in the
visible region and the absence of deactivation complications
due to triplet state population. However, the synthetic
methodologies for construction of water-soluble dyes are
limited, and the provision of larger quantities may represent
a bottleneck for further studies and development in biome-
dical analysis. Various strategies leading to water-soluble
BODIPY dyes have been envisaged by introduction of oligo-
(ethyleneglycol) chains,5 R-galactosylceramide,6 nucleotides,7
(1) Haugland, R. P.; Spence, M. T. Z.; Johnson, I. D.; Basey, A. The
Handbook: A Guide to Fluorescent Probes and Labeling Technologies,
10th ed.; Molecular Probes: Eugene, OR, 2005.
(2) (a) Lakowicz, J. R. Principles of Fluorescence Spectroscopy, 3rd
ed.; Springer: Heidelberg, 2006. (b) Kasha, M.; Rawls, H. R.; El-Bayoumi,
M. A. Pure Appl. Chem. 1965, 11, 371.
(4) (a) Ziessel, R.; Ulrich, G.; Harriman, A. New J. Chem. 2007, 31,
496. (b) Loudet, A.; Burgess, K. Chem. Rev. 2007, 107, 4891. (c) Ulrich,
G.; Ziessel, R.; Harriman, A. Angew. Chem., Int. Ed. 2008, 47, 1184.
(5) (a) Atilgan, S.; Ozdemir, T.; Akkaya, E. U. Org. Lett. 2008, 10,
4065. (b) Zhu, S.; Zhang, J.; Vegesna, G.; luo, F.-T.; Green, S. A.; Liu, H.
Org. Lett. 2011, 13, 438.
(3) Bergstroem, F.; Mikhalyov, I.; Haeggloef, P.; Wortmann, R.; Ny,
T.; Johansson, L. B. A. J. Am. Chem. Soc. 2002, 124, 196.
r
10.1021/ol200969r
Published on Web 05/20/2011
2011 American Chemical Society

sulfonated peptides,8 N,N-bis(2-hydroxyethyl)amines,9
carboxylates,10 sulfonates,11 or betaine12 units. In most
cases, relatively good solubility in water was reached but
the absence of aggregates and strong fluorescence were only
observed with the less conjugated dyes emitting in the 500À
540 nm range. Despite great synthetic effort, little success
was achieved for the more extended dyes emitting in the
650À700 nm window and synthetic methodologies remain
elusive and purification procedures extremely tedious.7,13
In this light, it may be noted that most key natural
biomolecules (ATP, DNA, etc.) are constructed with
phosphate residues, which ensure good solubility and
stability in water under daylight conditions. Herein, we
report a convenient strategy based on the grafting of
alkylphosphonate residues onto the boron center or the
styryl side arms of BODIPY dyes to achieve water solubi-
lity and fluorescence properties without the formation of
aggregates even with the dyes emitting in the red part of the
electromagnetic spectrum.
the nucleophile and TEA as base to quench the nascent
acid.18
Hydrolysis of both the carboxylic ester and the phos-
phonate to the corresponding monophosphonic acid was
brought about in one step using NaOH. The water-soluble
dyes 7a and 7b were isolated by crystallization from the
reaction mixture without the need to purify them by
reverse-phase chromatography. Notice that the presence
of the carboxylate group certainly facilitate water solubi-
lity and prevent dye aggregation through charge repulsion.
Surprisingly, the distyryl derivatives 6c could not be
properly hydrolyzed under the given conditions and pro-
vided an intractable mixture of dyes.
Scheme 1. Synthesis of BODIPY Phospohnates
Our first goal was the efficient preparation of a variety of
dyes bearing an ethyleneglycol chain with a TIPS-protected
alcohol at one end and tethered to the boron center via an
ethynyl bond (3aÀc in Scheme 1). The pivotal derivative 214
was prepared in 83% yield from the TIPS-monoprotected
diethyleneglycolate15 and propargyl bromide. The Grignard
derivatives of 2 readily react with the fluoro derivatives
1aÀc in excellent yields applying a protocol previously
developed by us.16 In the second step, 2 M HCl can be used
to deprotect the alcohol. The corresponding alkoxides were
prepared with NaH or t-BuOK and allowed to react with
diethoxy(phosphonate) trifluoromethylsulfonate17 provid-
ing a mixture of mono- and disubstituted compounds 5aÀc.
Despite the use of an excess of the triflate, complete
substitution could not be attained. The disubstituted
derivatives 5aÀc were isolated in only 35% yields but
were efficiently transformed to the corresponding
ethylcarboxylic esters 6aÀc via a carboalkoxylation
promoted by [Pd(PPh3)2Cl2] under a flow of CO at
atmospheric pressure and in the presence of EtOH as
To circumvent this problem and to further extend the
methodology to blue dyes, an alternative suggested by
earlier work of Lindsey on chlorins19 is to link the phos-
phonate to the side arm and to graft a simple ethyleneglycol
chain onto the boron center. In practice, it was found that
the method outlined in Schemes 3 and 4 must be adapted to
the nature of the aryl residues carrying the alkylphospho-
nate fragments. We were fortunate first to be able to prepare
blue dyes 8 and 9 with a TIPS-protected phenol or naphthol
aldehydes using established procedures (Scheme 2).20,21
From these building blocks, it was possible to substitute
the fluoro ligands by alkyne derivatives, leading to dyes
(6) Vo-Hoang, Y.; Micouin, L.; Ronet, C.; Gachelin, G.; Bonin, M.
ChemBioChem. 2003, 4, 27.
(7) Giessler, K.; Griesser, H.; Gohringer, D.; Sabirov, T.; Richert, C.
Eur. J. Org. Chem. 2010, 3611.
(8) Niu, S. L.; Ulrich, G.; Ziessel, R.; Kiss, A.; Renard, P.-Y.;
Romieu, A. Org. Lett. 2009, 11, 2049.
(9) Jiao, L.; Li, J.; Zhang, S.; Wei, C.; Hao, E.; Vicente, M. G. H. New
J. Chem. 2009, 33, 1888.
€
(10) (a) Li, L.; Han, J.; Nguyen, B.; Burgess J. Org. Chem. 2008, 73,
1963. (b) Dodani, S. C.; He, Q.; Chang, C. J. J. Am. Chem. Soc. 2009,
€
131, 18020. (c) Dilek, O.; Bane, S. L. Bioorg. Med. Chem. Lett. 2009, 19,
6911. (d) Brellier, M.; Duportail, G.; Baati, R. Tetrahedron Lett. 2010,
51, 1269.
(11) (a) Boyer, J. H.; Haag, A. M.; Sathyamoorthi, G.; Soong, M. L.;
Thangaraj, K.; Pavlopoulos, T. G. Heteroat. Chem. 1993, 4, 39. (b) Li, L.;
Han, J.; Nguyen, B.; Burgess, K. J. Org. Chem. 2008, 73, 1963.
(12) Niu, S. L.; Ulrich, G.; Retailleau, P.; Harrowfield, J.; Ziessel, R.
Tetrahedron Lett. 2009, 50, 3840.
(13) Niu, S. L.; Massif, C.; Ulrich, G.; Ziessel, R.; Renard, P.-Y.;
Romieu, A. Org. Biomol. Chem. 2011, 9, 66.
(14) Ulrich, G.; Ziessel, R.; Niu, S. L.; Haeffele, A.; Bura, T. WO
2010076516 A1 20100708.
(18) El Ghayoury, A.; Ziessel, R. J. Org. Chem. 2000, 65, 7757.
(19) Borbas, K. E.; Chandrashaker, V.; Muthiah, C.; Kee, H. L.;
Holten, D.; Lindsey, J. S. J. Org. Chem. 2008, 73, 3145.
(20) (a) Dost, Z.; Atilgan, S.; Akkaya, E. U. Tetrahedron 2006, 62,
8484. (b) Atilgan, S.; Ekmekci, Z.; Dogan, A. L.; Guc, D.; Akkaya, E. U.
Chem. Commun. 2006, 4398. (c) Ziessel, R.; Bura, T.; Olivier, J.-H.
Synlett 2010, 2304.
(15) Moody, C. J.; Miller, D. J. Tetrahedron 1998, 54, 2257.
(16) Goze, C.; Ulrich, G.; Ziessel, R. Org. Lett. 2006, 8, 4445.
(17) Xu, Y.; Flavin, M. T.; Xu, Z.-Q. J. Org. Chem. 1996, 61, 7697.
(21) Bozdemir, O. A.; Sizmen, F.; Buyukcakir, O.; Guliyev, R.;
Cakmak, Y.; Akkaya, E. U. Org. Lett. 2010, 12, 1400.
Org. Lett., Vol. 13, No. 12, 2011
3073

Scheme 2. Synthesis of TIPS Protected Phenol- and
Naphtol-Based Distyryl BODIPY Dyes
Scheme 4. Synthesis of Naphtol-Based BODIPY Phosphonate
Alkylation of the naphthol hydroxyl in 17 was accom-
plished with diethoxy(phosphonate)trifluoromethylsulf-
onate. Simultaneously, the carboxylic acid was esterified, as
10 and 15 in excellent yields. At this stage, the reaction
sequence for the phenol and naphthol dyes must diverge due
to solubility and reactivity reasons. For the phenol-based
protocol depicted in Scheme 3, after deprotection of the
TIPS in basic conditions and nucleophilic substitution using
diethoxy(phosphonate) trifluoromethylsulfonate, the de-
sired phosphonated derivative 12 was easily prepared.
Conversion of the phenyliodo residue to the correspond-
ing ethylester 13 is feasible using the carboalkoxylation
reaction referred to above. Finally, concomitant hydrolysis
of the carboxy- and phosphonate esters gave rise to the target
derivative 14 which precipitated during the course of the
reaction and appeared to be perfectly soluble and highly
fluorescent in water (vide infra). Complete hydrolysis of the
phosphonate to the phosphonic acid moieties was not
achievable at this stage even when using a high concentration
of base in different solvents or using TMS-Br as an activator.
1
clearly seen from the H and 31P NMR spectra, the latter
displaying two phosphorus signals at 18.9 and 19.1 ppm
(in CDCl3) for 18 whereas a single signal at 20.1 ppm
(in CD3OD) was observed for dye 13. As would be expected
in light of previous observations, hydrolysis under basic con-
ditions leads to the monophosphonic and carboxylic acids in
dye 19. This dye precipitated during the course of the reaction
and could be isolated pure by centrifugation (see Figure 1b).
It appeared to be highly soluble and fluorescent in water.
The proton NMR of the prototypic dyes 7b and 19 are
shown in Figure 1. In both cases, integration of the ethyl
signal at 3.96 and 1.24 ppm versus the AB quartet of the
phenyl-carboxylateisdiagnostic forthemonohydrolysis of
Scheme 3. Synthesis of Phenol-Based BODIPY Phosphonates
For the naphthol derivative 15, an alternative route had
to be devised involving first the formation of the carbox-
yester 16 and then the simultaneous deprotection of the
TIPS and saponification of the ester, leading to dye 17 in
excellent yields (Scheme 4).
Figure 1. 1H NMR of dyes (a) 7b and (b) 19 in d4-MeOH.
Org. Lett., Vol. 13, No. 12, 2011
3074

the phosphonate. For 7b, the presence of two type of ethyl
groups confirms this statement. For 19, the doublet at 4.23
ppm with a JPH coupling constant of 10.2 Hz is easly
identifiable and the doublet at 8.35 with a JHH = 16.4 Hz
confirms the trans conformation of the vinyl functions.
Furthermore, the presence of a singlet at 6.86 ppm assigned
to the β-pyrrolic protons of the dipyrromethene core con-
firms the symmetry of the molecule. The well-defined NMR
spectra and expected integrales confirm the absence of
aggregates and impurities as well as the molecular structures.
The photophysical properties of the new dyes were
measured in alcohols and water and the data are collected
in Table 1. The absorption spectra of the final compounds
7aÀb, 14 and 19 exhibit a major absorption peak at the
expected wavelength with extinction coefficients in the range
60 000 to 66 000 MÀ1cmÀ1 for the classical BODIPY and
100 000 to 120 000 MÀ1cmÀ1 for the more extended dyes.
This transition corresponds to the S0fS1 transition.22
Figure 3. Absorption, emission and excitation spectra measured
in water: green and violet lines, dye 7a; blue and red lines, dye 14.
For the excitation spectra (dashed lines), the emission is plotted
at 560 nm for 7a and at 700 nm for 14.
Note that the weaker absorption in the UV window
corresponds to an S0fS2 transition and to an internal
charge transfer band (ICTB) for dyes 14 and 19 (Figures 2
and 3).21 By irradiation in the lower energy absorption
bands, strong fluorescence is observed and the shape of the
emission bands shows excellent mirror symmetry with the
lowest-energy absorption band, suggesting that the chro-
mophore has a similar geometry in the ground and first
excited states. Furthermore, the high quantum yields, the
weak Stokes shift around 400 cmÀ1 and the nanosecond
excited state lifetime regime are in keeping with a singlet
excited state. The radiative deactivation rates are higher
than the nonradiative deactivation rates for dyes 7aÀb and
conversely for the red emitting dyes 14 and 19. Note also
that these dyes entail a significant red-shift of the emission
extending to 830 nm in case of dye 19. It is also interesting
that the spectroscopic properties of dyes 7aÀb, 14 and 19
are very similar in EtOH, MeOH and pure water, exclud-
ing the formation of aggregates.8 As shown in Figures 2
and 3, the excitation spectra (in dashed lines) match the
absorption spectra fairly well over the entire absorption
window (250À700 nm). Finally, it appears that the aqu-
eous solutions of the dyes are stable for weeks without
significant degradation or flocculation of the dyes.
Table 1. Selected Optical Data Measured in Polar Solution
b
b
λabs
dye (nm)
ε
λF
τF
kr
knr
M
À1 cmÀ1 (nm) (ns) ΦF 108 sÀ1 108 sÀ1 solvent
a
7a 500
7a 500
7b 518
7b 516
14 638
14 638
19 649
19 651
66 000 509 4.8 0.59 12.2
64 500 509 5.0 0.61 12.1
8.5 EtOH
7.8 H2O
60 000 531 7.5 0.73
60 500 529 6.4 0.68 10.7
106 000 654 5.3 0.45 8.5
102 000 655 3.5 0.42 12.0
9.7
3.6 EtOH
5.0 H2O
10.4 MeOH
16.6 H2O
15.0 MeOH
29.9 H2O
120 500 664 4.3 0.35
118 500 667 2.8 0.23
8.1
5.3
a Using Rhodamine 6G as reference, Φ = 0.78 in water,23 λex = 480
nm for 7a,b and using tetramethoxydiisoindomethene-difluoroborate (φf
= 0.51 in methanol)24 λex = 570 nm for 14 and λex = 580 nm for 19.
b Calculated using the following equations: kr = Φf /τf ; knr = (1-Φf)/τf,
assuming that the emitting state is produced with unit quantum efficiency.
In short, a convergent method has been successfully
developed for the synthesis of water-soluble BODIPY
dyes emitting light in the 490 to 750 nm range. The
proposed method uses alkylphosphonate as a tool
to carry out the synthesis allowing purification of each
synthetic intermediate by conventional methods. At the
end of the procedure, hydrolysis led to the free carbox-
ylate as well as the ethylphosphonate as substituents.
The desired dyes were obtained purely by precipita-
tion without the need for column chromatography.
Figure 2. Absorption, emission and excitation spectra measured
in water: green and violet lines, dye 7b; blue and red lines, dye 19.
For the excitation spectra (dashed lines), the emission is plotted
at 560 nm for 7b and at 730 nm for 19.
Supporting Information Available. Synthetic proce-
dures and analytical data reported herein. This material
is available free of charge via the Internet at http://pubs.
acs.org.
(22) Karolin, J.; Johansson, L. B.-A.; Strandberg, L.; Ny, T. J. Am.
Chem. Soc. 1994, 116, 7801.
(23) Olmsted, J. J. Phys. Chem. 1979, 83, 2581–2584.
(24) Ulrich, G.; Goeb, S.; De Nicola, A.; Retailleau, P.; Ziessel, R.
Synlett 2007, 1517.
Org. Lett., Vol. 13, No. 12, 2011
3075