Amphiphilic meso(sulfonate ester fluoroaryl)porphyrins: Refining the substituents of porphyrin derivatives for phototherapy and diagnostics

Article abstract of DOI:10.1016/j.tet.2012.08.007

A set of amphiphilic fluorinated porphyrins appended with sulfonate ester groups were synthesized and fully characterized. The reaction proceeds efficiently, with high yields, with an improved methodology. Their potential use as imaging and phototherapeutic agents was assessed measuring relevant photophysical properties. It is shown that these porphyrins have good photostability, long triplet lifetimes (between 47 μs and 102 μs), high singlet oxygen quantum yields (0.74≤FD≤1.00), low fluorescence quantum yields (<0.04) and sharp 19F NMR peaks. The data on the new meso(sulfonate ester fluoroaryl)porphyrins illustrate the potential of perfluorinated sulfonate esters to improve physical properties relevant for cancer imaging and photodynamic therapy.

Full text of DOI:10.1016/j.tet.2012.08.007

Tetrahedron 68 (2012) 8767e8772
Contents lists available at SciVerse ScienceDirect
Tetrahedron
journal homepage: www.elsevier.com/locate/tet
Amphiphilic meso(sulfonate ester fluoroaryl)porphyrins: refining the substituents
of porphyrin derivatives for phototherapy and diagnostics
a
b
b,
a
a,c
*
~
ꢀ
Ana V.C. Simoes , Agnieszka Adamowicz , Janusz M. Da˛browski , Mario J.F. Calvete , Artur R. Abreu
,
b
a
a,
*
_
Grazyna Stochel , Luis G. Arnaut , Mariette M. Pereira
a Chemistry Department, University of Coimbra, Rua Larga 3004-535, Coimbra, Portugal
b
ꢀ
Faculty of Chemistry, Jagiellonian University, Ingardena 3, 30-060 Krakow, Poland
c Edifício Bluepharma, Rua da Bayer, S. Martinho do Bispo, 3045-016 Coimbra, Portugal
a r t i c l e i n f o
a b s t r a c t
Article history:
Received 31 May 2012
Received in revised form 2 August 2012
Accepted 2 August 2012
Available online 16 August 2012
A set of amphiphilic fluorinated porphyrins appended with sulfonate ester groups were synthesized and
fully characterized. The reaction proceeds efficiently, with high yields, with an improved methodology.
Their potential use as imaging and phototherapeutic agents was assessed measuring relevant photo-
physical properties. It is shown that these porphyrins have good photostability, long triplet lifetimes
(between 47
m
s and 102
m
s), high singlet oxygen quantum yields (0.74ꢀFDꢀ1.00), low fluorescence
quantum yields (<0.04) and sharp 19F NMR peaks. The data on the new meso(sulfonate ester fluoroaryl)
porphyrins illustrate the potential of perfluorinated sulfonate esters to improve physical properties
relevant for cancer imaging and photodynamic therapy.
Keywords:
Porphyrins
Anticancer agents
Imaging agents
Singlet oxygen
Photodynamic therapy
Photosensitizers
Ó 2012 Elsevier Ltd. All rights reserved.
1. Introduction
Molecules that could combine properties adequate for both
therapy and diagnostics, when associated with clinically relevant
Porphyrins are useful in many medical and industrial applica-
tions, such as phototherapy, photodiagnosis, photocatalysis, solar
energy conversion, and optoelectronics.1e3 The properties of the
triplet state of these molecules are particularly important because
it acts as the immediate precursor of reactive oxygen species (ROS).
Another key factor in the above mentioned application is strong
absorption of light in the visible region of the spectrum.3,4
Photodynamic therapy (PDT) is a selective cancer phototherapy
under active investigation by researchers both in fundamental
science and in the clinic.5 Cancer PDT involves administration of
a photosensitizer, which preferentially localizes in the tumors,
followed by irradiation with visible or near-infrared light. The
photosensitizer absorbs light and in the presence of molecular
oxygen transfers energy to oxygen producing ROS, especially sin-
glet oxygen, which are responsible for selective tumor destruction,
tumor-associated vascular damage, and activation of anti-tumor
immune responses.6,7
tools, are particularly appealing.1 Porphyrin-based photosensitizers
were found to accumulate readily in tumors and have been ex-
tensively studied in this area because of their high vascular per-
meability, as well as of their affinity for proliferating endothelium,
and the lack of lymphatic drainage in tumors.8 There is also evi-
dence that the amphiphilic character of the photosensitizer is an
important factor.9,10 Several approaches have been proposed in
order to modify the lipophilicity of the photosensitizer.11 Besides
the chemical and photophysical properties of such designed sen-
sitizers, it is interesting to see if they can be imaged in neoplastic
tissue. Fluorinated derivatives are particularly attractive in this
respect, as the 19F nucleus is an interesting probe for nuclear
magnetic resonance (NMR) imaging. Examples of 19F NMR imaging
of photosensitizer or other drugs in vivo have already been re-
ported and were recently reviewed.12 Fluorine atoms have been
introduced into biologically active compounds to modulate their
physicochemical and pharmacokinetic properties and improve
their potency with respect to biomedical aplications.13,14 Moreover,
fluorescent sensitizers may also be used as diagnostic agents in
photodetection of cancer.1 These properties suggest that the de-
velopment of fluorinated compounds for use as PDT sensitizers and
* Corresponding authors. Tel.: þ351 239854474; fax: þ351 239827703; e-mail
addresses: jdabrows@chemia.uj.edu.pl (J.M. Da˛browski), mmpereira@qui.uc.pt
(M.M. Pereira).
0040-4020/$ e see front matter Ó 2012 Elsevier Ltd. All rights reserved.
http://dx.doi.org/10.1016/j.tet.2012.08.007

~
A.V.C. Simoes et al. / Tetrahedron 68 (2012) 8767e8772
8768
19F magnetic resonance imaging probes can be relevant contribu-
tion to this field.
We recently reported the synthesis of meso-halogenated
sulfonated or sulfonamide porphyrins15e17 and their dihydro18 and
tetrahydro19e21 analogues. We have shown that they have a low
toxicity in the dark, high phototoxicity and a tendency to accu-
mulate in a range of cancer cells. The most promising candidates for
new PDT-agents are the sulfonamide substituted bacteriochlorins,
which produce high yields of singlet oxygen, superoxide ions and
hydroxyl radicals,22e24 extending the average tumor growth delay
by 44 days, with respect to a control group in DBA mice bearing S91
melanoma tumors.25 We have also shown that the fluorinated
porphyrins belonging to this series of compounds have relatively
long singlet lifetimes and sizeable fluorescence quantum yields.
In this paper we offer additional strategies to incorporate fluo-
rine atoms in related porphyrins. We describe a simple and efficient
synthetic method to tune the amphiphilicity of sulfonate ester
porphyrins by selecting the type and number of fluorine atoms as
well as the length of the alkyl sulfonate ester chains. We also de-
scribe the photophysical properties of the new sulfonate ester
fluorinated porphyrins as well as their 19F NMR spectra in order to
evaluate their potential as PDT sensitizers and/or 19F NMR probes.
2. Results and discussion
2.1. Synthesis
Scheme 1. Reagents and conditions: (i) 1, HSO3Cl, 60 ꢁC, 1.5 h, 97%; 2, HSO3Cl, 100 ꢁC,
2 h, 91%; (ii) alcohol, NaOH (aq), THF, rt, 2 h, yields greater than 75%.
The 5,10,15,20-tetrakis(2-fluorophenyl)porphyrin 1 and 5,10,15,
20-tetrakis(2,6-difluorophenyl)porphyrin 2 were prepared by one-
pot condensation of pyrrole with the corresponding fluorinated
benzaldehyde using acetic acid/nitrobenzene as solvent and oxi-
dant, accordingly to our previously described methodology.16,26,27
Using this synthetic approach, crystals with high purity from
fluorinated porphyrins 1 and 2 were obtained by direct crystalli-
zation from the reaction medium, with 20% and 10%, respectively,
in good agreement with literature.16,26,27 In order to synthesize the
desired fluorinated amphiphilic porphyrins, containing sulfonate
ester appendices, the derivatization of 1 and 2 into the corre-
sponding chlorosulfonated porphyrins was achieved by mixing the
porphyrins with an excess of chlorosulfonic acid.
Table 1
Yields of meso(sulfonate ester fluoroaryl)porphyrins (5aee; 6aee)
Alcohol
FP(SO2Cl)4
5
Yield%
F2P(SO2Cl)4
6
Yield%
Derivatives
Derivatives
HOC3H7
HOC3H4F3
HOC4H9
HOC4H3F6
OHC5H6F5
a-FPC3H7
b-FPC3H4F3
c-FPC4H9
d-FPC4H3F6
e-FPC5H6F5
94
96
89
84
95
a-F2PC3H7
b-F2PC3H4F3
c-F2PC4H9
d-F2PC4H3F6
e-F2PC5H6F5
78
85
82
81
82
So, in a typical reaction, 1 was mixed with chlorosulfonic acid at
room temperature and the reaction was maintained at 60 ꢁC, for
1.5 h. After cooling to room temperature, dichloromethane was
added and the acid was removed via continuous extraction with
water. After evaporation of the solvent the 5,10,15,20-tetrakis(2-
fluoro-5-chlorosulfophenyl)porphyrin 3 was obtained with 97%
yield. The 5,10,15,20-tetrakis(2,6-difluoro-3-chlorosulfophenyl)
porphyrin 4 was obtained in 91% yield in a similar procedure, by
reaction of porphyrin 2, chlorosulfonic acid at 100 ꢁC for 2 h.
The chlorosulfonic acid group is susceptible to react with alco-
hols, in basic conditions.28,29 In a typical experiment, the chlor-
osulfonyl porphyrin derivative was dissolved in dry THF and slowly
added to a solution of the desired alcohol (40 equiv) in the presence
of a concentrated aqueous solution of NaOH (8 equiv) in a small
amount of THF. The reaction was kept at room temperature for 1 h
until full consumption of the starting material was observed.
The corresponding sulfonate esters (5aee; 6aee, see Scheme 1)
were obtained, after work-up, in good yields (78e96%), in-
dependently of the alcohol used (see Table 1). This synthetic
strategy allowed us not only to prepare a family of amphiphilic
porphyrins, containing several sulfonate ester appendices, but also
the inclusion of one or more fluorine atoms in the porphyrin
structure.
in all the compounds. However, it should be emphasized that the
compounds 5b and 6b possess an extra narrow intense peak at
d
z63.8 and 5d and 6d at d z73.0, due to the presence of a CF3
fragment in the porphyrin sulfonate ester appendices, which allows
them to be potential probes for 19F MRI. These fluorinated amphi-
philic compounds may find use as 19F MRI probes for cancer de-
tection because the fluorine atom has 100% NMR active isotope
with intrinsic high sensitivity in biological systems,12 and it is
well established that porphyrins preferentially accumulate in
tumors.8
2.2. Spectroscopic characterization of sulfonate ester
substituted fluorinated porphyrins
The ground state absorption spectrum of the representative
porphyrin 6d-F2PC4H3F6, recorded at room temperature in ethanol
is presented in Fig. 1. The spectrum shows the characteristic bands
from free base porphyrins with the D2h symmetry, described by
a four orbitals model.30 The intense Soret band is observed around
410 nm and four bands between 500 and 650 nm are assigned to
the (0,0) and (1,0) vibronic progressions of the Qx and Qy bands,
Table 2. The longest-wavelength absorption bands at 641e649 nm
are crucial for PDT process, because only red light has sufficient
tissue penetration ability. Using measured absorbance for various
As expected, all 19F NMR aromatic signals appeared as multi-
plets (d z90.7e99.8) not only due to the coupling with adjacent
protons, but also due to the presence of a mixture of atropisomers

~
A.V.C. Simoes et al. / Tetrahedron 68 (2012) 8767e8772
8769
2.3. Photophysical properties of sulfonate ester substituted
fluorinated porphyrins
The kinetics of the transient absorption changes (
DA) of the
porphyrins in solution was determined by laser flash photolysis.
The spectra were constructed from fitting data from the sets of the
decay curves, collected with 10 nm intervals. The transient decay
times in the presence of oxygen are independent from the moni-
toring wavelength and are typical of triplet states quantitatively
transferring energy to molecular oxygen. Triplet lifetimes are pre-
sented in Table 3. In argon-saturated ethanol solution, the decays
remain monoexponential (inset in Fig. 2) and lifetimes increase up
to 100 ms. Transient absorption spectrum of 5d-FPC4H3F6 in ethanol
is shown in Fig. 2. The strong negative absorbance change is due to
the bleaching of the ground-state absorption and the clearly de-
fined pseudoisosbestic points reflect the fact that all molecules
return to this ground state, which also means that photo-
degradation is negligible. The rate constant of energy transfer from
the triplet state of the photosensitizer to molecular oxygen (kq) can
be calculated from the triplet lifetimes in the absence (nitrogen
saturated solution), s0T, and in the presence of oxygen (air saturated
Fig. 1. Electronic absorption and fluorescence spectra of F2PC4H3F6 (6d) in ethanol, at
room temperature. The fluorescence was obtained with excitation at 411 nm.
Table 2
solution), sT, using the relationship:
Absorption and fluorescence properties of fluorinated porphyrins in ethanol
Absorption
Fluorescence
Table 3
Triplet state lifetimes in the presence and absence of oxygen determined in ethanol,
with respective oxygen quenching rate constants at room temperature and singlet
oxygen quantum yields for selected porphyrins
a
l
(nm), ε (mMꢂ1 cmꢂ1
)
l
(nm)
FF
B
Qy
Qy
Qx
Qx
(0e0)
(1e0)
656;
709
648;
712
646;
712
647;
709
(0e1)
505
13.7
508
18.2
508
12.2
505
19.4
513
7
(0e0)
535
3.27
538.5
3.79
539
2.89
536
3.96
549
(0e1)
583
4.53
584.5
5.97
585
3.98
583
6.73
581
2.5
(0e0)
648.5
1.6
641.5
1.02
641.5
0.84
649
0
Porphyrin
sT
[
m
s]
sT,air [ns]
kq (ꢄ109)[Mꢂ1 sꢂ1
]
FD
6b-F2PC3H4F3
5d-FPC4H3F6
5a-FPC3H7
410
208
412.5
1000
412
248
410
286
413
160
0.039
0.032
0.031
0.030
0.008
6b-F2PC3H4F3
5d-FPC4H3F6
5a-FPC3H7
92ꢅ41
102ꢅ36
47ꢅ6
385ꢅ141
361ꢅ130
337ꢅ21
343ꢅ15
1.23ꢅ0.45
1.31ꢅ0.47
1.40ꢅ0.09
1.38ꢅ0.06
0.74ꢅ0.03
0.81ꢅ0.02
0.74ꢅ0.02
1.00ꢅ0.04
6d-F2PC4H3F6
80ꢅ34
6d-F2PC4H3F6
ClPOHb
2.18
633
0.5
638;
698
1.2
a
The error associated with FF is 0.001.
From Ref. 20 determined in PBS, room temperature.
b
concentrations of this series of porphyrins, the molar absorption
coefficients were determined from Beer’s law. Their absorption
band maxima and corresponding molar absorption coefficients are
presented in Table 2. Aggregation does not seem to occur in the mM
concentration range, because as the concentrations were increased,
the longest wavelength absorption band was not displaced to
longer wavelengths, and its bandwidth at half height did not in-
crease. Among the studied porphyrins, 6d-F2PC4H3F6 has the most
promising spectroscopic properties. The Qx band of this porphyrin
derivative is four times more intense than that of published
sulfonated and chlorinated porphyrin (ClPOH)15 and is further
displaced to the red (633/649 nm). Considering that ClPOH was
shown to be efficient photosensitizer in vitro against different
cancer cells,15 we expect that the sulfonate ester substituents
studied in this work may increase the phototoxicity of the
sensitizers.
Fig. 2. Time-resolved transient absorption spectra of 5d-FPC4H3F6 in ethanol, mea-
sured by laser flash photolysis, 20 ꢁC, lexc¼355 nm. Inset: decay curve of the triplet
state signal of the FPC4H3F6 (5d) in the absence of oxygen.
The fluorescence spectrum of 6d-F2PC4H3F6 shown in Fig. 1 is
representative of the fluorescence of these porphyrins. The fluo-
rescence excitation spectra of all the compounds correspond well
with their absorption spectra and confirm the purity and non-
aggregation of the samples. Earlier studies with other haloge-
nated porphyrins established that the presence of bulky ortho-
chloro groups reduce the tendency of porphyrins to aggregation.14
The fluorescence quantum yields of these porphyrins were also
determined according to published procedures22 and calculated
relatively to that of TPP (FF¼0.11). All fluorescence data are given in
Table 2.
ꢀ
ꢁ.
kq
¼
1=sT ꢂ 1=s0T ½O2ꢃ
and the oxygen concentration in ethanol, [O2]¼2.1ꢄ10ꢂ3 M.31 In
fact, sT 0>> T and the actual value of s0T is irrelevant to calculate the
s
value of kq reported in Table 3.
Spin statistics require that kq should be 1/9 of the diffusion rate
constant, kdiff. The value of kdiff was estimated from the fluores-
cence intensity of 5,10,15,20-tetrakis(4-sulfophenyl)porphyrin
(PSO3H) in the absence and in the presence of oxygen, its lifetime in
the absence of oxygen and the oxygen concentration in ethanol.

~
A.V.C. Simoes et al. / Tetrahedron 68 (2012) 8767e8772
8770
Porphyrins triplets are quenched with a rate constants kq value in
atoms in sulfonate ester substituents of tetraphenylporphyrin de-
rivatives improves photophysical and photochemical properties
relevant for PDT, with respect to the properties of other haloge-
nated porphyrins that already showed relevant efficacy.15,22 Pho-
tosensitizers with such substituents are worth further
investigations for PDT and NMR imaging.
the range 1.2e1.4ꢄ109 Mꢂ1 sꢂ1
,
which is similar to
kq¼1.4ꢄ109 Mꢂ1 sꢂ1 determined for TPP.22 Taking into account that
kdiff¼5.4ꢄ109 Mꢂ1 sꢂ1 in ethanol at 20 ꢁC, our data are in good
agreement with literature values for similar systems.32 The values
of kq reported in Table 3 indeed approach be 1/9 kdiff, as in related
compounds.15,20
The internal heavy-atom effect assists halogenated porphyrins,
to generate triplet states with nearly unit quantum yields.25 The
low fluorescence quantum yields obtained for all the fluorinated
porphyrins reported in this work suggests that intersystems
crossing to the triplet state are also a major singlet deactivation
pathway for these molecules. The triplet state of the photosensi-
tizer is the precursor of reactive oxygen species, namely singlet
oxygen. Following the excitation of the porphyrins at 355 nm, we
3. Conclusions
Amphiphilic meso(sulfonate ester fluoroaryl)porphyrins can be
readily synthesized from economical starting materials. Their ab-
sorption bands at w649 nm show values of ε of porphyrins with
perfluorinated sulfonate esters representing a four fold improve-
ment over those of photodynamically active sulfonated porphy-
rins.15 Moreover, the additional fluorine atoms contribute to
increase singlet oxygen quantum yields via the internal heavy-atom
effect in the intersystem crossing from the singlet to the triplet
state of the photosensitizers. The 19F nucleus is also potentially
useful as a probe for in vivo imaging with NMR spectroscopy, par-
ticularly in compounds 5b and 6b that possess a CF3 group in the
sulfonate ester appendices. Among the compounds studied in this
work, 6d-F2PC4H3F6 has the most relevant photophysical properties
and the highest singlet oxygen quantum yield for PDT, which sug-
gests that perfluorinated sulfonate esters may help improve the
PDT efficacy of tetraphenylporphyrin derivatives.
detected a species emitting at 1270 nm with a lifetime of 13e18
These values are in good agreement with lifetime of singlet oxygen
phosphorescence in the ethanol, which is 15
s,32 confirming the
ms.
m
origin of the emission measured. Also, the shape of emission
spectra taken between 1250 and 1300 nm (Fig. 3) corresponds to
that of singlet oxygen. Singlet oxygen quantum yields (FD) were
obtained by comparing the intensity of singlet oxygen emission at
1270 nm in an air-equilibrated sample containing a given sensitizer
against the intensity obtained from an optically matched sample
containing a reference sensitizer. We used phenalenone as a refer-
ence and used its literature value for the singlet oxygen quantum
yield in ethanol, 0.95.33 Table 3 presents the singlet oxygen quan-
tum yields (FD) measured in ethanol according to the procedure
recently recommended by us, that makes use of the linear de-
pendence between singlet oxygen emission intensity and the en-
ergy of the exciting laser pulse at low laser energies.22 The reason
why the lines presented in Fig. 3 are not crossing through zero is
probably related with scatter light heating the powermeter, which
is not coming from the laser bean. Any scattered or ambient light
can change the 0 of the powermeter. The error associated with FD is
4% or less.
4. Experimental
4.1. Instrumentation
1H- and 19F NMR spectra were obtained using a Bruker Avance III
400 MHz spectrometer and 376.5 MHz, respectively. The chemical
shifts are given in parts per million (ppm) relative to tetrame-
thylsilane at
d
0.00 ppm for proton spectra and relative to tri-
fluoroacetic acid at
d
0.00 ppm for 19F spectra. Mass spectra ESI-FIA-
TOF were acquired using a Micromass Q-TOF2 spectrometer con-
taining a Z-spray source, an electrospray probe and an injection
syringe. Flash column chromatography was performed with silica
gel grade 60, 70e230 mesh. Purity of the compounds was estab-
lished by HPLC and is greater than 95% in all cases. The HPLC
analysis were obtained in a Agilent 1100 series, equipped with
a UVevisible and DAD detector Agilent 1100 series, using a Agilent
Technologies Zorbax ODS, 5
nitrile/water as eluent, depending on the compound.
m
m, 4.6ꢄ250 mm and gradient aceto-
4.2. Spectroscopic and photophysical measurements
Absorption and fluorescence spectra were measured by stan-
dard techniques using Shimadzu UV-2100 and SPEX Fluoromax
3.22 spectrophotometers, respectively. The reference employed for
fluorescence quantum yield measurements was TPP (FF¼0.11).
Time-resolved singlet oxygen phosphorescence measurements
were made with a modification of an Applied Photophysics LKS.60
flash photolysis spectrometer, and using the third harmonic of
a Nd:YAG laser (Spectra-Physics Quanta Ray GCR 130, 5e6 ns
FWHM) for excitation and HP Infinium (500 MHz, 1 GSa/s) or Tek-
tronix DPO 7254 (2.5 GHz, 40 GSa/s) oscilloscopes. Singlet oxygen
emission was detected using a Hamamatsu R5509-42 photo-
multiplier, cooled to 193 K in a liquid nitrogen chamber (Products
for Research, model PC176TSCE005). Interposition of a Melles Griot
cold mirror (03MCS005), that reflects more than 99% of the incident
light in the 400e700 nm range, and of a Scotch RG665 filter,
eliminated from the infrared signal all harmonic contributions
of the sensitizer emission in the 400e900 nm range. A 600
line diffraction grating was mounted in place of a standard one to
improve spectral resolution and sensitivity in the NIR. This
Fig. 3. 1270 nm emission intensity as a function of the laser energy. Inset: singlet
molecular oxygen emission intensity as a function of wavelength for 5a-FPC3H7.
It is especially interesting that the porphyrin with the highest
number of fluorine atoms, 6d-F2PC4H3F6, is capable of generating
singlet molecular oxygen with unit quantum yield. In fact, in-
creasing the number of fluorine atoms increases FD but does not
change kq appreciably. This suggests that the increase in FD is
probably related with an increase in triplet quantum yield and,
therefore, to a heavy-atom effect. The introduction of fluorine

~
A.V.C. Simoes et al. / Tetrahedron 68 (2012) 8767e8772
8771
equipment allows for spectral identification of the singlet oxygen
phosphorescence and measurement of singlet oxygen lifetime in
the nanosecond and microsecond ranges. Fitting to the experi-
mental data the decays of the singlet molecular oxygen emissions
(I0D) measured in ethanol for the reference (phenalenone,
FD¼0.95ꢅ0.02) and for porphyrins, at a given laser intensity, we
obtain a relation between emission intensities that is identical to
the relation between the singlet molecular oxygen quantum yields.
The actual singlet oxygen quantum yields were obtained from the
ꢂ2.84 (s, 2H, eNH); dF, ppm (376.5 MHz, CDCl3), ꢂ100.09 to
ꢂ100.20 (m, 4F); HRMS (ESI-FIA-TOF) [MþH]þ found: 1175.2281,
calculated for [C56H51F4N4O12S4] 1175.2244.
4.5.2. 5,10,15,20-Tetrakis[2-fluoro-5-(3-trifluoropropyloxy)-sulfonyl-
phenyl] porphyrin 5b-FPC3H4F3. Yield¼96%; mp >250 ꢁC; dH, ppm
(400 MHz, CDCl3) 8.80 (s, 8H, b-H), 8.76e8.72 (m, 4H, PheH), 8.44
(t, 4H, J¼4 Hz, PheH), 7.79e7.75 (m, 4H, PheH), 4.53e4.50 (m, 8H,
eOeCH2e), 2.69e2.63 (m, 8H, eCH2CF3), ꢂ2.85 (s, 2H, eNH); dF
,
linear dependence between ID0 and the energy of the laser pulse Eh
,
n
ppm (376.5 MHz, CDCl3) ꢂ63.73 (s, 12F, eCF3), ꢂ98.94 to ꢂ99.05
(m, 4F, PheF); HRMS (ESI-FIA-TOF) [MþH]þ 1391.1186, calculated
for [C56H39F16N4O12S4] 1391.1114.
for the range of laser energies where a good linear relationship was
found. Good linearity was observed up to 12 mJ/pulse with fluori-
nated porphyrins.
4.5.3. 5,10,15,20-Tetrakis[2-fluoro-5(-3-butyloxy)-sulfonylphenyl]
porphyrin 5c-FPC4H9. Yield¼89%; mp >250 ꢁC; dH, ppm (400 MHz,
4.3. General porphyrin synthesis
CDCl3) 8.80 (s, 8H, b-H); 8.74e8.70 (m, 4H, PheH), 8.44e8.41 (m,
The synthesis of porphyrins 1 and 2 was carried out using the
nitrobenzene method16,26,27 prepared by the condensation of the
desired fluorinated arylaldehyde (43 mmol) with pyrrole (3 mL,
43 mmol) in a mixture of acetic acid (140 mL, 2.45 mol) and ni-
trobenzene (70 mL, 0.68 mol) at 120 ꢁC. The solution was cooled to
room temperature and 50 mL of methanol was added to promote
precipitation. The crystals of the porphyrins were filtered off,
washed with methanol and dried. By this method, crystals of por-
phyrins were obtained without further purification procedures.
Porphyrins 1 and 2 were obtained with yields of 20% and 10%, re-
spectively. Their identity was confirmed by NMR, FAB and micro-
analysis, with good agreement with literature data.16,26,27
4H, PheH), 7.74 (t, 4H, J¼8 Hz, PheH), 3.76e3.73 (m, 8H,
eOeCH2e), 1.80e1.76 (m, 8H, eCH2e), 1.49e1.43 (m, 8H, eCH2e),
0.96e0.92 (m, 12H, eCH3), ꢂ2.84 (s, 2H, eNH); dF, ppm (376.5 MHz,
CDCl3) ꢂ99.88 to ꢂ100.21 (m, 4F) HRMS (ESI-FIA-TOF) [M]þ
1230.2750, calculated for [C60H58F4N4O12S4] 1230.2870.
4.5.4. 5,10,15,20-Tetrakis[2-fluoro-5-(2,2,3,4,4,4-hexafluorobutyloxy)-
sulfonylphenyl] porphyrin 5d-FPC4H3F6. Yield¼84%; mp >250 ꢁC; dH
,
ppm (400 MHz, CDCl3), 8.78e8.74 (m, 12H,
b
-HþPheH), 8.47e8.45
(m, 4H, PheH), 7.80 (t, 4H, J¼8 Hz, PheH), 5.10e4.96 (m, 4H, CHeF),
4.70e4.55 (m, 8H, OeCH2), ꢂ2.85 (s, 2H, NH); dF, ppm (376.5 MHz,
CDCl3) ꢂ73.89 to ꢂ73.99 (m, 12F, CF3), ꢂ97.77 to ꢂ97.87 (m, 4F, Ph-
F), ꢂ113.02 to ꢂ119.64 (m, 8F, CF2), ꢂ211.10 to ꢂ211.20 (m, 4F,
CFeH); HRMS (ESI-FIA-TOF) [MþH]þ 1663.0679, calculated for
[C60H35F28N4O12S4] 1663.0609.
4.4. General procedure for porphyrin chlorosulfonation
Chlorosulfonation of the fluorinated porphyrins was carried out
according to a method previously developed.19 The required por-
phyrin (200 mg) and chlorosulfonic acid (10 mL, 150 mmol) were
stirred at 60 ꢁC for 1 and 100 ꢁC for 2 during 1.5 h and 1 h, re-
spectively. After this period, dichloromethane (200 mL) was added
to the solution. A continuous water extraction was carried out, with
stirring, until neutralization. The dichloromethane solution was
then washed with sodium hydrogen carbonate and dried over an-
hydrous Na2SO4. The purification by column chromatography in
silica gel using dichloromethane as eluent, and subsequent solvent
evaporation yielded the desired chlorosulfonated porphyrins as
purple crystals, in agreement with the literature.19
4.5.5. 5,10,15,20-Tetrakis[2-fluoro-5-(4,4,5,5,5-pentafluoropentyloxy)-
sulfonylphenyl] porphyrin 5e-FPC5H6F5. Yield¼95%; mp >250 ꢁC; dH
,
ppm (400 MHz, CDCl3) 8.79 (s, 8H, b-H), 8.75e8.69 (m, 4H, PheH),
8.46e8.42 (m, 4H, PheH), 7.77 (t, 4H, J¼16 Hz, PheH), 4.39 (t, 8H,
J¼8 Hz, eOeCH2e), 2.30e2.20 (m, 8H, eCH2eCF2), 2.16e2.11 (m,
8H, eCH2e), ꢂ2.84 (s, 2H, NH); dF, ppm (376.5 MHz, CDCl3) ꢂ84.31
(s, 12F, CF3), ꢂ99.27 to ꢂ99.47 (m, 4F, Ph-F), ꢂ116.99 (s, 8F, CF2);
HRMS (ESI-FIA-TOF) [MþH]þ 1647.1533, calculated for
[C64H47F24N4O12S4] 1647.1612.
4.5.6. 5,10,15,20-Tetrakis[2,6-difluoro-3-(3-propyloxy)-sulfonyl-
phenyl] porphyrin 6a-F2PC3H7. Yield¼78%; mp >250 ꢁC; dH, ppm
4.5. General procedure for the synthesis of sulfonate ester
fluorinated porphyrins
(400 MHz, CDCl3) 8.85 (s, 8H,
b
-H), 8.52e8.46 (dd, 4H, J¼8z, 16 Hz,
PheH), 4.38e4.34 (m, 8H, eOeCH2e), 1.85e1.76 (m, 8H, eCH2),
0.97 (t, J¼8 Hz, 12H, eCH3), ꢂ2.83 (s, 2H, NH); dF, ppm (376.5 MHz,
CDCl3) ꢂ94.87 to ꢂ95.68 (m, 4F), ꢂ96.88 to ꢂ99.80 (m, 4F); HRMS
(ESI-FIA-TOF) [MþH]þ 1247.1957, calculated for [C56H47F8N4O12S4]
1247.1940.
In an ice bath cooled flask, (ca. 5 ꢁC) the corresponding alcohol
was placed (40 equiv) and a concentrated solution of NaOH
(8 equiv), dissolved in a minimum amount of dry THF, was added
with stirring, for half hour. Then, the chlorosulfonated porphyrin
(0.1 mmol), dissolved in dry THF (4 mL) was added to the flask. The
mixture was allowed to stir at room temperature and TLC con-
trolled. After the reaction finished (ca. 1 h), CH2Cl2 (50 mL) was
added and the solution was washed with distilled water
(3ꢄ100 mL) and then dried over anhydrous Na2SO4. The solvent
was concentrated by rotary evaporation then purified by column
chromatography in silica gel using dichloromethane/ethyl acetate
as eluent, and subsequent solvent evaporation yielded the re-
spective sulfonate ester fluorinated porphyrins.
4.5.7. 5,10,15,20-Tetrakis[2,6-difluoro-3(-3-trifluoropropyloxy)-sulfo-
nylphenyl] porphyrin 6b-F2PC3H4F3. Yield¼85%; mp >250 ꢁC; dH
,
ppm (400 MHz, CDCl3) 8.84 (s, 8H,
b
-H), 8.49 (t, 4H, J¼8 Hz, PheH),
7.61 (t, 4H, J¼8 Hz, PheH), 4.60e4.57 (m, 8H, OeCH2e), 2.67e2.57
(m, 8H, eCH2CF3), ꢂ2,84 (s, 2H, eNHe); dF, ppm (376.5 MHz, CDCl3)
ꢂ63.85 to ꢂ63.96 (m, 12F, CF3), e93.69 to ꢂ94.22 (m, 8F, PheF),
ꢂ98.78 to ꢂ98.94 (m, 4F, PheF); HRMS (ESI-FIA-TOF) [MþH]þ
found: 1463.0828, calculated for [C56H35F20 N4O12S4] 1463.0737.
4.5.8. 5,10,15,20-Tetrakis[2,6-difluoro-3(-3-butyloxy)-sulfonylphenyl]
porphyrin 6c-F2PC4H9. Yield¼82%; mp >250 ꢁC; dH, ppm (400 MHz,
4.5.1. 5,10,15,20-Tetrakis[2-fluoro-5-(3-propyloxy)-sulfonylphenyl]
porphyrin 5a-FPC3H7. Yield¼94%; mp >250 ꢁC; dH, ppm (400 MHz,
CDCl3) 8.89 (s, 8H, b-H); 8.59e8.54 (m, 4H, PheH), 7.67e7.64 (m,
CDCl3) 8.81 (s, 8H,
b
-H), 8.76e8.70 (m, 4H, PheH), 8.45e8.41 (m,
4H, PheH), 3.77e3.74 (m, 8H, eOeCH2e), 2.18e2.16 (m, 8H,
eCH2e), 1.87e1.84 (m, 8H, eCH2e), 1.27e1.25 (m, 12H, eCH3),
ꢂ2,82 (s, 2H, eNH); dF, ppm (376.5 MHz, CDCl3) ꢂ90.74 to ꢂ91.14
4H, PheH), 7.74 (t, 4H, J¼8 Hz, PheH), 4.29 (td, 8H, J¼4, 8 Hz,
eOeCH2e), 1.88e1.79 (m, 8H, eCH2), 1.02 (t, 12H, J¼8 Hz, eCH3),

~
A.V.C. Simoes et al. / Tetrahedron 68 (2012) 8767e8772
8772
(m, 4F), ꢂ97.15 to ꢂ97.48 (m, 4F); HRMS (ESI-FIA-TOF) [MþH]þ
5. Agostinis, P.; Berg, K.; Cengel, K. A.; Foster, T. H.; Girotti, A. W.; Golinick, S. O.;
Hahn, S. M.; Hamblin, M. R.; Juzeniene, A.; Kessel, D.; Korbelik, M.; Moan, J.; Mroz,
P.; Nowis, D.; Piette, J.; Wilson, B. C.; Golab, J. CA Cancer J. Clin. 2011, 61, 250.
6. Dougherty, T. J.; Gomer, C. J.; Henderson, B. W.; Jori, G.; Kessel, D.; Korbelik, M.;
Moan, J.; Peng, Q. J. Natl. Cancer Inst. 1998, 90, 889.
7. Castano, A. P.; Mroz, P.; Hamblin, M. R. Nat. Rev. Cancer 2006, 6, 535.
8. Dolmans, D.; Fukumura, E. D.; Jain, R. K. Nat. Rev. Cancer 2003, 3, 380.
9. Adams, K. R.; Berembaum, M. C.; Bonnett, R.; Nizhnik, A. N.; Salgado, A. J. Chem.
Soc., Perkin Trans. 1 1992, 1465.
found: 1303.2483, calculated for [C60H55F8N4O12S4] 1303.2493.
4.5.9. 5,10,15,20-Tetrakis[2,6-difluoro-3-(2,2,3,4,4,4-hexafluorobutyloxy)-
sulfonylphenyl] porphyrin 6d-F2PC5H3F6. Yield¼81%; mp >250 ꢁC; dH
,
ppm (400 MHz, CDCl3) 8.81 (s, 8H, b-H), 8.52e8.49 (m, 4H, PheH),
7.67e7.50 (m, 4H, PheH), 5.01e4.87 (m, 4H, CHeF), 4.72e4.63 (m, 8H,
CH2), ꢂ2.80 (s, 2H, NH); dF, ppm (376. 5 MHz, CDCl3) ꢂ72.91 to ꢂ74.13
(m, 12F, CF3), ꢂ92.47 to ꢂ93.43 (m, 4F, PheF), ꢂ98.73 to ꢂ99.88 (m, 4F,
Ph-F), ꢂ113.19 to ꢂ120.32 (m, 8F, CF2), ꢂ211.19 (s, 4F, CFeH);
HRMS (ESI-FIA-TOF) [M]þ found: 1734.9030, calculated for
[C60H30F32N4O12S4] 1734.0232.
~
ꢀ
10. Pereira, M. M.; Monteiro, C. J. P.; Simoes, A. V. C. S.; Pinto, M. A.; Arnaut, L. G.; Sa,
~
G. F. F.; Silva, E. F. F.; Rocha, L. B.; Simoes, S.; Formosinho, S. J. J. Porphyrins
Phathlocyanines 2009, 13, 567.
11. Zheng, G.; Potter, W. R.; Camacho, S. H.; Missert, J. R.; Wang, G.; Bellnier, D. A.;
Henderson, B. W.; Rodgers, M. A. J.; Dougherty, T. J.; Pandey, R. K. J. Med. Chem.
2001, 44, 1540.
12. Goslinski, T.; Piskorz, J. J. Photochem. Photobiol., C 2011, 12, 304.
13. Lepre, C. A.; Moore, J. M.; Peng, J. W. Chem. Rev. 2004, 104, 3641.
14. Dalvit, C.; Fagerness, P. E.; Hadden, D. T. A.; Sarver, R. W.; Stockman, B. J. J. Am.
Chem. Soc. 2003, 125, 7696.
4.5.10. 5,10,15,20-Tetrakis[2,6-difluoro-3-(4,4,5,5,5-penta-
fluoropentyloxy)-sulfonylphenyl] porphyrin 6e-F2PC5H6F5. Yield¼82%;
15. Dabrowski, J. M.; Pereira, M. M.; Arnaut, L. G.; Monteiro, C. J. P.; Peixoto, A. F.;
mp >250 ꢁC; dH, ppm (400 MHz, CDCl3) 8.79 (t, 8H, J¼12 Hz,
b-H),
Karocki, A.; Urbanska, K.; Stochel, G. Photochem. Photobiol. 2007, 83, 897.
~
ꢀ
16. Monteiro, C. J. P.; Pereira, M. M.; Pinto, S. M. A.; Simoes, A. V. C.; Sa, G. F. F.;
8.50e8.43 (m, 4H, PheH), 7.57e754 (m, 4H, PheH), 4.39 (s, 4H,
OeCH2e), 2.14e1.93 (m, 16H, eCH2eCF2þeCH2e), ꢂ2.90 (s, 2H,
NH); dF, ppm (376,5 MHz, CDCl3) ꢂ84.33 to ꢂ84.46 (m, 12F, eCF3),
ꢂ91.08 to ꢂ91.30 (m, 4F, PheF), ꢂ98.99 to ꢂ99.39 (m, 4F, Ph-F),
ꢂ117.00 to ꢂ117.19 (m, 8F, CF2); HRMS (ESI-FIA-TOF) [M]þ found:
1718.2811, calculated for [C64H42F28N4O12S4] 1718.1235.
~
Arnaut, L. G.; Formosinho, S. J.; Simoes, S.; Wyatt, M. F. Tetrahedron 2008, 64,
132.
17. Dabrowski, J. M.; Krzykawska, M.; Arnaut, L. G.; Pereira, M. M.; Monteiro, C. J. P.;
Simoes, S.; Urbanska, K.; Stochel, G. ChemMedChem 2011, 6, 1715.
18. Dabrowski, J. M.; Arnaut, L. G.; Pereira, M. M.; Monteiro, C. J. P.; Urbanska, K.;
~
Simoes, S.; Stochel, G. ChemMedChem 2010, 5, 1770.
19. Pereira, M. M.; Monteiro, C. J. P.; Simoes, A. V. C.; Pinto, S. M. A.; Abreu, A. R.; Sa,
G. F. F.; Silva, E. F. F.; Rocha, L. B.; Dabrowski, J. M.; Formosinho, S. J.; Simoes, S.;
Arnaut, L. G. Tetrahedron 2010, 66, 9545.
Acknowledgements
20. Pereira, M. M.; Monteiro, C. J. P.; Peixoto, A. F. In Target in Heterocyclic Systems;
Attanasi, O. A., Spinelli, D., Eds.; Italian Society of Chemistry: 2008; p 258.
~
21. Pereira, M. M.; Abreu, A. R.; Goncalve, N. P. F.; Calvete, M. J. F.; Simoes, A. V. C.;
A.V.C.S. thanks FCT/QREN/FEDER Portugal for PhD grant: SFRH/
BD/65699/2009 and also PEST/C/QUI/UI 0313/2011; PTDC/QUI-QUI/
112913/2009 for funding. NMR data was obtained at the Nuclear
Magnetic Resonance Laboratory of the Coimbra Chemistry Centre
(www.nmrccc.uc.pt), Universidade de Coimbra, supported in part
by grant REEQ/481/QUI/2006 from FCT, POCI-2010 and FEDER,
Portugal. This work was supported in part by the International PhD-
studies program at the Faculty of Chemistry Jagiellonian University
within the Foundation for Polish Science MPD Program co-financed
by the EU European Regional Development Fund. J.M.D. thanks the
Ministry of Science and Higher Education for Iuventus Plus grant nr
IP2011009471.
ꢀ
Monteiro, C. J. P.; Arnaut, L. G.; Eusebio, M. E. S.; Canotilho, J. Green Chem. 2012, ,
http://dx.doi.org/10.1039/C2GC35126A
22. Silva, E. F. F.; Serpa, C.; Dabrowski, J. M.; Monteiro, C. J. P.; Arnaut, L. G.;
Formosinho, S. J.; Stochel, G.; Urbanska, K.; Simoes, S.; Pereira, M. M. Chem.dEur.
J. 2010, 16, 9273.
23. Dabrowski, J. M.; Urbanska, K.; Arnaut, L. G.; Pereira, M. M.; Abreu, A. R.;
~
Simoes, S.; Stochel, G. ChemMedChem 2011, 6, 465.
24. Dabrowski, J. M.; LArnaut, G.; Pereira, M. M.; Urbanska, K.; Simoes, S.; Stochel,
G.; Cortes, L. Free Radical Biol. Med. 2012, 52, 1188.
25. Dabrowski, J. M.; Arnaut, L. G.; Pereira, M. M.; Urbanska, K.; Stochel, G. Med-
ChemComm 2012, 3, 502.
26. Gonsalves, A. M. d. A. R.; Varejao, J. M. T. B.; Pereira, M. M. J. Heterocycl. Chem.
~
1991, 28, 635.
27. Johnstone, R. A. W.; Nunes, M. L. P. G.; Pereira, M. M.; Rocha Gonsalves, A. M. d.
A.; Serra, A. C. Heterocycles 1996, 43, 1423.
28. Miller, S. C. J. Org. Chem. 2010, 75, 4632.
29. Meshram, G. A.; Patil, V. D. Tetrahedron Lett. 2009, 50, 1117.
30. Gouterman, M. J. Mol. Spectrosc. 1961, 6, 138.
References and notes
31. Gonc¸ alves, P. J.; Aggarwal, L. P. F.; Marquezin, C. A.; Ito, A. S.; De Boni, L.; Barbosa
Neto, N. M.; Rodrigues, J. J., Jr.; Zílio, S. C.; Borissevitch, I. E. J. Photochem. Pho-
tobiol., A 2006, 181, 378.
1. Vicente, M. G. H. Curr. Med. Chem.: Anti-Cancer Agents 2001, 1, 175.
2. Ethirajan, M.; Chen, Y.; Joshi, P.; Pandey, R. K. Chem. Soc. Rev. 2010, 40, 340.
3. Szaci1owski, K.; Macyk, W.; Drzewiecka-Matuszek, A.; Brindell, M.; Stochel, G.
Chem. Rev. 2005, 105, 2647.
32. Bonnett, R.; McGarvey, D. J.; Harriman, A.; Land, E. J.; Truscott, T. G.; Winfield, U.
J. Photochem. Photobiol. 1988, 48, 271.
33. Murov, S. L.; Carmichael, I.; Hug, G. L. Handbook of Photochemistry, 2nd ed.;
Marcel Dekker: New York, NY, 1993.
4. Pineiro, M.; Carvalho, A. L.; Pereira, M. M.; Gonsalves, A. M. d’ A. R.; Arnaut, L.
G.; Formosinho, S. J. Chem.dEur. J. 1998, 4, 2299.