DNA photocleavage by a cationic BODIPY dye through both singlet oxygen and hydroxyl radical: New insight into the photodynamic mechanism of BODIPYs

Article abstract of DOI:10.1002/cphc.201200224

Two new NIR-absorbing BODIPY dyes, each bearing two pyridinium groups, are synthesized and their DNA-binding affinities and DNA photocleavage abilities examined in depth. While one BODIPY dye photocleaves DNA mainly through singlet oxygen, the other photo

Full text of DOI:10.1002/cphc.201200224

DOI: 10.1002/cphc.201200224
DNA Photocleavage by a Cationic BODIPY Dye through
Both Singlet Oxygen and Hydroxyl Radical: New Insight
into the Photodynamic Mechanism of BODIPYs
[
a, b]
[a]
[a]
[a]
[a]
Jianguang Wang,
Yuanjun Hou, Wanhua Lei, Qianxiong Zhou, Chao Li,
[a]
[a]
Baowen Zhang, and Xuesong Wang*
Two new NIR-absorbing BODIPY dyes, each bearing two pyridi-
nium groups, are synthesized and their DNA-binding affinities
and DNA photocleavage abilities examined in depth. While
one BODIPY dye photocleaves DNA mainly through singlet
oxygen, the other photocleaves DNA through both singlet
oxygen and hydroxyl radical. To the best of our knowledge,
this is the first example of a hydroxyl radical being involved in
the photodynamic behavior of BODIPY-type dyes. EPR experi-
ments confirm the ability of these and several related BODIPYs
to generate superoxide anion radical and hydroxyl radical. This
finding may shed light on the mechanism of BODIPY-based
photodynamic therapy (PDT) and open a new avenue for de-
velopment of more efficient BODIPY-type PDT agents.
1
. Introduction
Photodynamic therapy (PDT) is a minimally invasive treatment
for a variety of cancers by interaction of three elements: light,
a photosensitizer, and oxygen. When exposed to proper wave-
lengths of light, the photosensitizer can be activated and un-
dergo electron/energy transfer through its triplet excited state
to generate cytotoxic reactive oxygen species (ROS), mainly
aza-BODIPYs further by attaching four iodine atoms on the dye
[10]
skeleton. In 2005, Nagano et al. disclosed that an iodinated
1
BODIPY is also a good O photosensitizer, but the examined
2
[11]
dye suffers from short absorption wavelength. Akkaya et al.
extended the absorption bands of brominated or iodinated
BODIPYs into the phototherapeutic window by incorporating
conjugation structures at 3- and 5-positions of the BODIPY
1
[1]
singlet oxygen ( O ). Although several porphyrin photosensi-
2
[12]
tizers, for example, Photofrin, are approved for clinical treat-
ment of certain types of cancers, they are far from ideal. One
of the main drawbacks of these photosensitizers is the relative-
ly weak absorbance in the phototherapeutic window of 600–
core. Recently, Ng and co-workers also reported a series of
unsymmetrical distyryl iodinated BODIPYs that show high in
[13]
vitro PDT activities. Additionally, You and co-workers found
that fusion of brominated thiophene rings with the BODIPY
1
[14]
900 nm. For Photofrin, the drawbacks also include the compli-
core may improve O2 quantum yield remarkably.
While
1
cated components from which it is composed and long-term
much interest focused on the O2 efficiency, absorption
[
2]
skin photosensitivity after PDT. Such limitations encourage
great efforts to search for new and more effective photosensi-
tizers, including porphyrin-type photosensitizers such as chlor-
window, and water solubility of the BODIPY family, another im-
1
portant factor, that is, the bioavailability of O , should also be
2
1
paid proper attention. O is a highly reactive species but has
2
[
3]
in and bacteriochlorin, and non-porphyrin-type photosensitiz-
ers such as cyanine dyes, squaraines, and boradiazaindacenes
a short apparent lifetime (ca. 2.0 ms), a low apparent diffusion
ꢀ6
2
coefficient (4ꢀ10 cm s), and thus a very limited sphere of ac-
tivity (about 155 nm in radius) in biological systems, and this
implies the importance of the binding ability of PDT agents
[
2e,4]
(abbreviated hereafter as BODIPY).
The BODIPYs are versatile organic dyes which have found
[
5]
wide applications in biological labeling and imaging, lumines-
[
5]
[5,6]
[7]
cent devices, chemical sensors,
solar cells, and photoca-
[
a] Dr. J. G. Wang, Prof. Y. J. Hou, Dr. W. H. Lei, Dr. Q. X. Zhou, Dr. C. Li,
Prof. B. W. Zhang, Prof. X. S. Wang
Key Laboratory of Photochemical Conversion
and Optoelectronic Materials
[
8]
talytic hydrogen generation, due to their excellent thermal
and photochemical stability, high fluorescence quantum yield,
intense absorption profile, good solubility, and chemical ro-
bustness. The intense absorption of BODIPYs in the near-infra-
red (NIR) region makes them suitable for PDT application, but
Technical Institute of Physics and Chemistry
Chinese Academy of Sciences
Beijing 100190 (China)
the low intersystem crossing (ISC) efficiency severely hampers
Fax: (+86)10-6487-9375
E-mail: g203@mail.ipc.ac.cn
1
the ability to generate O and thus the photodynamic activity
2
xswang@mail.ipc.ac.cn
of BODIPYs. In 2002, O’Shea and co-workers successfully dealt
[
b] Dr. J. G. Wang
with this problem by introduction of bromine atoms into
Graduate University of Chinese Academy of Sciences
Beijing 100049 (China)
1
a series of aza-BODIPYs to give the resulting dyes high O
2
[
9]
quantum yields and potent photodynamic activity. Recently,
Supporting information for this article is available on the WWW under
http://dx.doi.org/10.1002/cphc.201200224.
1
Ramaiah and co-workers improved the O quantum yields of
2
ChemPhysChem 2012, 13, 1 – 10
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X. S. Wang et al.
[
15]
toward their biotarget. DNA is one of the main targets of
a new avenue for the development of novel and more efficient
BODIPY-based PDT agents
[
16]
many anticancer drugs, including PDT agents. As a result,
DNA photocleavers, particularly activated by visible and NIR
light, are highly pursued due to their promising potential in
PDT. Owing to the negatively charged character of DNA, a vari- 2. Results and Discussion
ety of positively charged dye molecules were scrutinized as
2
.1. Synthesis, Photophysical, and Photochemical Properties
DNA photocleavers to take advantage of the attractive electro-
static interaction, which may enhance the binding affinity of
the dye molecules toward DNA and therefore improve the bio-
The BODIPY 1 was synthesized by following reported proce-
[6a]
dures. Bromination or iodination of 1 led to formation of
BODIPY 2 or 3, which, after further condensation with 4-pyridi-
necarboxaldehyde, transformed into BODIPY 4 or 5, respective-
ly. By reaction with iodomethane, target BODIPYs 6 and 7 were
finally obtained, as shown in Scheme 1. Both 6 and 7 were
[
17]
availability of the ROS. Though several BODIPYs or aza-BODI-
PYs bearing either quaternary ammonium groups or protonat-
able pyridyl groups were investigated as potential PDT
[
9d,12d,13b,18]
agents,
there are no reports on the interactions of
1
cationic BODIPYs and DNA up to now, which is believed to be
helpful for better understanding of their PDT mechanism.
In this work, two BODIPY dyes 6 and 7 bearing pyridinium
groups (Scheme 1) were synthesized and their binding features
and photocleavage activities toward DNA were examined in
depth. Unexpectedly, while 7 photocleaves DNA exclusively
structurally identified by H NMR and HRMS. For comparison,
1–5 were also isolated and structurally characterized by
1
H NMR and EI-MS or MALDI-TOF MS.
Figure 1a shows the visible absorption spectra of 1–7 in ace-
tonitrile. Compared to 1, bromination or iodination at 2- and
6-positions of 1 leads to about 25 nm (for 2) and 31 nm (for 3)
redshift in absorption maxima (Table 1). Further conjugation
with pyridyl-substituted ethylene at 3- and 5-positions gives
rise to another 100 nm of bathochromic shift. Finally, methyla-
tion at pyridyl N atoms makes the visible absorption bands of
6 and 7 totally move into the phototherapeutic window, with
absorption maxima centered at 648 and 649 nm, respectively.
Moreover, the molar extinction
1
1
through O , 6 photocleaves DNA through both O and hy-
2
2
droxyl radical (COH). Further experiments indicate that 1, 2, and
can also generate COH with different efficiencies. To the best
3
of our knowledge, this is the first example that BODIPYs are
able to generate COH on visible and NIR irradiation. Since COH
is the most active among all ROS, these results may open
coefficients of 6 and 7 at their
absorption maxima are 4.5ꢀ
4
ꢀ1 ꢀ1
1
0 cm
m
and
5.6ꢀ
4
ꢀ1 ꢀ1
1
0 cm m , respectively, much
larger than that of the Q bands
of general porphyrin-based pho-
tosensitizers.
Similar to the absorption spec-
tra, the fluorescence emission
spectra
a marked redshift from 1, over
–5, to 6 and 7, as shown in Fig-
also
experience
2
ure 1b and Table 1. As expected,
the fluorescence quantum yields
of 2 and 3 decrease significantly
with respect to 1, mainly due to
the intramolecular heavy-atom
effect of bromine or iodine
atoms. For 6 and 7, the fluores-
cence quantum yields decrease
further than for 2 and 3, which
may be attributed to intramolec-
ular charge transfer (ICT) rather
than energy-gap law, since 2
and 4 or 3 and 5 have similar
fluorescence quantum yields
We also measured the oil/
water partition coefficients of 6
and 7. Due to the presence of
two pyridinium groups, both 6
and 7 display amphiphilic char-
Scheme 1. Synthesis of the examined BODIPY dyes.
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DNA Photocleavage by a Cationic BODIPY Dye
Figure 2. Time-dependent photobleaching of AMDA sensitized by 6, 7, or
MB in PBS. The absorbance changes of AMDA at 400 nm were monitored as
a function of irradiation (l>600 nm) time. A control experiment was carried
out in the absence of photosensitizer.
trast, no absorbance decrease was observed in the absence of
1
6
and 7, that is, O stems from the excited 6 or 7. With MB as
2
1
reference, the O quantum yields of 6 and 7 are estimated to
2
be 0.10 and 0.22, respectively, in good agreement with the
stronger heavy-atom effect of iodine compared to bromine.
1
The ability of 6 and 7 to generate O was also confirmed by
2
EPR experiments. As shown in Figure 3, irradiation of an air-sa-
turated CH CN solution of 6 or 7 and TEMP with a 532 nm
3
laser led to a three-line EPR spectrum with equal intensity and
N
hyperfine coupling constants of a =16.0 G, which is assigna-
1
[23]
Figure 1. Visible absorption (a) and normalized fluorescence emission (b)
spectra of 1–7 (1.0 mm) in acetonitrile.
ble to TEMPO (the adduct of TEMP and O ). The TEMPO
2
signal intensity of 7 is larger than that of 6, in line with the
AMDA bleaching experiments. Additionally, we also
compared the TEMPO signal intensities of 1–5 as
Table 1. Photophysical properties of BODIPYs 1–7.
shown in Figure S1 (Supporting Information). The
1
O generation efficiencies follow the order 1<2<
[
a]
ꢀ1 ꢀ1 [a]
[a]
[d]
[f]
[e]
2
Compound
l
abs(max) [nm]
e (cm
m
)
l
em(max) [nm]
F
fl
F
D
P
c
3
and 4<5, in accordance with the heavy-atom
[
[
b]
b]
[
1
2
3
4
5
6
7
497
522
528
620
620
648
649
77000
68000
68000
57000
64000
45000
56000
514
544
556
638
643
680
686
0.71
0.18
0.032
0.19
0.04
0.095
0.014
n.d.
n.d.
n.d.
n.d.
n.d.
n.d.
n.d.
n.d.
n.d.
n.d.
effect.
b]
[
c]
2.2. DNA Binding Properties
[
c]
[
[
c]
c]
0.10 0.29
0.22 0.34
Before examining the DNA photocleavage activities
of 6 and 7, their binding toward CT DNA was first
studied by absorption titration, circular dichroism
spectroscopy, viscosity experiments, and EB dis-
placement assay.
[a] Absorption maxima, molar extinction coefficients at absorption maxima, and fluores-
cence maxima measured in acetonitrile. [b] Fluorescence quantum yields determined at
[
19]
258C by using fluorescein in 0.1m aqueous NaOH solution as standard (Ffl =0.85 ).
[
c] Fluorescence quantum yields determined at 258C by using MB in ethanol as stan-
[
20]
dard (Ffl =0.04 ). [d] Singlet oxygen quantum yields determined by the AMDA
Absorption titration is one of the simplest and
most useful methods to investigate the binding in-
teractions of small molecules, particularly dyes,
toward DNA. Figure 4 shows the absorption spec-
tral changes of 7 on titration of CT DNA. At the be-
[
33]
D
bleaching method, taking MB in PBS buffer as reference (F =0.52 ). [e] n-Octanol/
water partition coefficients. [f] n.d.: not determined.
acter, with n-octanol/water partition coefficients of 0.29 and
.34, respectively. Amphiphilicity is expected to be an advant-
age for PDT agents by simplifying formulation and favoring cel-
ginning of titration (0ꢁRꢁ1.5, R=[CT DNA]/[7]), significant
hypochromism (42% at R=1.5 vs. R=0) and a bathochromic
shift around 670–750 nm were observed. Further addition of
CT DNA (1.5ꢁRꢁ6), however, led to a hyperchromic effect
(43% at R=3 vs. R=1.5) and a hypsochromic shift around
600–700 nm. At R>6, the absorption band of 7 shows hypo-
chromism and bathochromic shift again. Titration of 6 by CT
DNA gave similar results (Figure S2, Supporting Information).
Though the spectral changes are somewhat complex, particu-
0
[
21]
lular internalization.
1
The O generation efficiencies of 6 and 7 in aqueous solu-
2
[22]
tions were evaluated by the AMDA bleaching method. On ir-
radiation with light above 600 nm, the absorbance of AMDA at
4
00 nm decreased gradually in the presence of 6 or 7 as
1
a result of its reaction with generated O (Figure 2). In con-
2
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X. S. Wang et al.
creases the p–p* transition energy of the ligand and results in
a bathochromic shift. On the other hand, the coupling p* orbi-
tal is partially filled by electrons, and thus decreases the transi-
[24]
tion probabilities and leads to hypochromism.
Circular dichroism measurements provide more information
on the binding interactions of 6 and 7 toward CT DNA. As
shown in Figure 5 and Figure S3 (Supporting Information), no
CD signals can be detected within the range of 550–750 nm
Figure 3. EPR signals obtained on irradiation of air-saturated CH
tions of TEMP (50 mm) and 6 or 7 (1 mm) with a 532 nm laser.
3
CN solu-
Figure 5. ICD spectra of 6 and 7 (10 mm) in the presence of CT DNA (50 mm).
for PBS solutions of 6 and 7. However, the presence of CT DNA
results in a negative CD signal within the absorption bands of
6
and 7, which indicates a ligand–DNA interaction. Intercala-
tors usually exhibit small induced CD (ICD) signals,
ꢀ
1
ꢀ1
<
10m cm at the maximum of the ICD signal, although the
magnitude of the signal does depend on the oscillator
[25]
strength of the ligand transitions. In our case, De values are
ꢀ
1
ꢀ1
ꢀ1
ꢀ1
ꢀ
5.96m cm for 6 and ꢀ8.25m cm for 7, in good agree-
ment with the intercalation binding mode.
More convincing evidence for the intercalation of 6 and 7
with CT DNA comes from viscosity experiments. Intercalators
such as EB generally cause a remarkable specific viscosity en-
hancement of DNA, as the result of the elongation of the DNA
helix caused by insertion of planar aromatic chromophores be-
[26]
tween base pairs. As shown in Figure 6, both 6 and 7 can in-
Figure 4. Absorption spectral changes of 7 (5 mm) on titration of CT DNA (0–
.1 mm) in PBS buffer (pH 7.4). a) 0ꢁRꢁ1.5, b) 1.5ꢁRꢁ6 and c) 6ꢁRꢁ20
R=[CT DNA]/[7]).
0
(
larly at 1.5ꢁRꢁ6, the overall trends are hypochromism and
bathochromic shift. Such spectral responses suggest an inter-
calation binding mode between CT DNA and 6 or 7. Intercala-
tion leads to a strong coupling between the p* orbital of the
ligand and the p orbitals of the DNA base pairs, and thus de-
Figure 6. Relative specific viscosities of CT DNA at 30.08C in 5 mm PBS
(
pH 7.2) as a function of the concentration ratio of photosensitizer and CT
DNA ([CT DNA]=165 mm, [6 or 7]=0–80 mm).
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DNA Photocleavage by a Cationic BODIPY Dye
crease the specific viscosity of CT DNA more efficiently than
EB, and this confirms their intercalation binding toward DNA.
An EB displacement assay was carried out to determine the
apparent binding constants Kapp of 6 and 7 to CT DNA. On ad-
dition of 6 or 7 (Figure 7) to the EB/CT DNA solution, the EB
fluorescence was quenched effectively due to its displacement
generation more efficiently than 7 allows one to recognize the
key role of DNA affinity, which can improve the bioavailability
1
1
of O and compensate the low O quantum yield.
2
2
To examine the photodynamic mechanisms behind the DNA
photocleavage, a series of ROS scavengers, including NaN for
3
1
[28]
ꢀ· [29]
O , SOD and catalase for superoxide anion radical (O ),
2
2
[30]
and KI for COH, were utilized to modulate the photocleavage
activities of 6 and 7. As shown in Figure 9, only NaN inhibits
3
Figure 9. Comparison of agarose gel electrophoresis patterns of photo-
cleaved supercoiled pBR322 DNA (0.1 mm in base pairs) by 7 (30 mm) in Tris/
CH
: DNA+7+SOD (500 UmL ); lane 3: DNA+7+KI (0.1 mm); lane 4:
DNA+7+NaN (0.1 mm); lane 5: DNA+7; lane 6: DNA alone; 50 min irradi-
3
COOH/EDTA buffer (pH 7.4). Lane 1: DNA+7+catalase (0.05 mm); lane
ꢀ
1
2
3
ation with light of l>600 nm. SC and NC denote supercoiled circular and
nicked circular form, respectively. NC% represents the percentage of the NC
form.
Figure 7. Fluorescence quenching of the EB (5 mm)–CT DNA (10 mm) solution
in PBS by 7 (0–2.5 mm). The arrow indicates increasing concentration of 7.
7
ꢀ1
from DNA by 6 or 7, from which K of 5.7ꢀ10 m for 6 and
the photocleavage activity of 7 markedly, and this indicates
app
7
ꢀ1
1
9
.8ꢀ10 m for 7 can be obtained. We also measured the
binding constant of MB, a well known intercalator, under the
a O mechanism or Type II mechanism for 7. In contrast, both
2
[
27]
NaN and KI can remarkably restrict the photocleavage activity
3
same conditions. The higher affinities of 6 and 7 toward CT
of 6 (Figure 10), which suggests that 6 exerts photodynamic
activity through both Type I and Type II mechanisms. To the
best of our knowledge, this is the first example that COH is gen-
7
ꢀ1 [35]
DNA compared EB (K =1.0ꢀ10 m ) and MB (Kapp =2.7ꢀ
app
7
ꢀ1
1
0 m ) can at least partly ascribed to their two positive charg-
es.
erated on NIR irradiation of a BODIPY-type photosensitizer and
participates in the damage of DNA.
2.3. DNA Photocleavage
The DNA photocleavage activities of 6 and 7 were character-
ized by using pBR322 plasmid DNA as target. On NIR irradia-
tion (ꢂ600 nm), both 6 and 7 can lead to DNA cleavage from
supercoiled circular form to nicked circular form (Figure 8).
Control experiments show that irradiation and the presence of
Figure 10. Comparison of agarose gel electrophoresis patterns of photo-
cleaved supercoiled pBR322 DNA (0.1 mm in base pairs) by 6 (30 mm) in Tris/
CH
3
COOH/EDTA buffer (pH 7.4). Lane 1: DNA+6+catalase (0.05 mm); lane
6
or 7 are necessary for DNA cleavage. The higher photocleav-
ꢀ1
2
: DNA+6+SOD (500 UmL ); lane 3: DNA+6+KI (0.1 mm); lane 4:
age activity of 7 compared to 6 can be attributed to its higher
3
DNA+6+NaN (0.1 mm); lane 5: DNA+6; lane 6: DNA alone; 50 min irradi-
1
ation with the light of l>600 nm. SC and NC denote supercoiled circular
and nicked circular form, respectively. NC% represents the percentage of the
NC form.
DNA affinity and larger O quantum yield. We also examined
2
the DNA photocleavage activity of MB under the same condi-
tions (Figure S4, Supporting Information). The cleavage activi-
1
ties follow the order 7>MB>6. The fact that MB sensitizes O
2
ꢀ
·
2
.4. O2 and COH Generation
Spin-trapping EPR is a powerful technique to detect formation
of O2Cꢀ and COH. We used DMPO as spin-trapping agent for
ꢀ
both O C and COH. As shown in Figure 11b, 532 nm laser irradi-
2
ation of an air-saturated acetonitrile solution of 6 or 7 and
DMPO led to an EPR signal. Control experiments indicate that
6 or 7, DMPO, irradiation, and oxygen are all necessary for ap-
pearance of this signal. We tentatively ascribe it to the signal
Figure 8. Comparison of agarose gel electrophoresis patterns of photo-
cleaved supercoiled pBR322 DNA (0.1 mm in base pairs) by 6 and 7 (30 mm)
in Tris/CH COOH/EDTA buffer (pH 7.4). Lane 1: DNA+7 (50 min irradiation);
3
lane 2: DNA+7 (25 min irradiation); lane 3: DNA+7 (in dark); lane 4:
DNA+6 (50 min irradiation); lane 5: DNA+6 (25 min irradiation); lane 6:
DNA+6 (in dark); lane 7: DNA alone (50 min irradiation); lane 8: DNA alone
ꢀ [31]
of DMPO–O C , though the signal intensity is weak and the
2
signal/noise ratio is low, probably due to the low absorptivity
(in dark). SC and NC denote supercoiled circular and nicked circular form, re-
spectively.
of 6 or 7 at the irradiation wavelength. If water is allowed into
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X. S. Wang et al.
Figure 12. Transient absorption spectra of 7 following 640 nm laser pulse
excitation (recorded at time delays of 0–500 ns). Inset: transient decay at
[
37]
4
20 nm, from which a triplet excited state lifetime of 475 ns was obtained
by mono-exponential fitting.
allow full interaction of the triplet excited states of 6 and 7
1
with O , supported by their O -generating abilities. As a result,
2
2
ꢀ
t is not likely the decisive factor in the generation of O C and
T
2
COH. The intersystem crossing efficiency is not the critical factor
ꢀ
either, since 1 generates O2C and COH most efficiently among
1
–3, just like the cases of 6 and 7.
ꢀ
The generation of O C and COH must involve electron trans-
2
fer; therefore, the redox potentials of 1–3 and 6, 7 were mea-
sured (Supporting Information Figure S7 and Table 2). The trip-
Figure 11. EPR signals obtained on irradiation of air-saturated solutions of
DMPO (50 mm) and 6 or 7 (1 mm) with a 532 nm laser in acetonitrile con-
taining 17% water (a) and in dry acetonitrile (b).
Table 2. Redox potentials, triplet excited state energies, and oxidation
potentials of 1–3 and 6, 7.
the solution, irradiation causes a four-line EPR signal with in-
tensity ratio of about 1:2:2:1 and hyperfine coupling constants
[b]
BODIPY
E
T
[eV]
E [V]
E [V] (vs. SCE)
N
H
[c]
[d]
of a =a =14.9 G in the case of 6 (Figure 11a). Furthermore,
(vs. SCE)
Eox
Ered
[28]
the presence of mannitol, a well-known scavenger of COH,
1
2
3
6
7
2.30
2.15
2.12
1.71
1.71
ꢀ1.36
ꢀ1.27
ꢀ1.26
ꢀ1.22
ꢀ1.19
0.94
0.88
0.86
0.49
0.52
ꢀ1.15
ꢀ0.91
ꢀ0.93
ꢀ0.28
ꢀ0.29
can inhibit signal generation (Figure S6a, Supporting Informa-
tion).These results suggest the signal can be attributed to
[
31]
DMPO–COH, which in turn supports the assignment of EPR
ꢀ
signal occurring in Figure 11a, because O C can transform into
2
COH in aqueous solutions. Clearly, 6 sensitizes O2Cꢀ and COH
more efficiently than 7, in line with the DNA photocleavage ex-
[a] Triplet excited state energy. [b] Triplet excited state oxidation poten-
tial. [c] Oxidation peak potential. [d] Reduction peak potential.
ꢀ
periments (Figures 9 and 10). We also examined O C genera-
2
ꢀ
tion sensitized by 1–3. The O C generation abilities follow the
2
order 1>2>3 (Figure S5, Supporting Information). In aqueous
let excited state energies (estimated from the phosphores-
cence maxima measured at 77 K) and triplet excited state oxi-
dation potential (calculated by the subtracting the triplet excit-
ed state energy from the ground-state oxidation potential) are
also included in Table 2. Clearly, electron transfer from the trip-
ꢀ
solution, the signal transformation from DMPO–O C to DMPO–
2
COH was observed again (Figure S6, Supporting Information).
We found that O has a negligible effect on the fluorescence
2
quantum yields of the examined BODIPYs. For example, the
ꢀ
fluorescence quantum yields of 1 in Ar- and air-saturated ace-
let excited states of 1–3 and 6, 7 to O to form O C is a thermo-
2
2
ꢀ
ꢀ
tonitrile are 0.73 and 0.71, respectively. Thus, O C and COH gen-
dynamically allowed process (the redox potential of O /O C
2
2
2
[32]
eration should stem from the triplet excited state of 1–7,
rather than the singlet excited state.
couple is ꢀ0.50 V vs. NHE or ꢀ0.74 V vs. SCE). The more neg-
ative triplet excited state oxidation potentials of 1 (about
100 mV compared to 2 and 3) and 6 (about 30 mV compared
To explore the role of the triplet excited state, the transient
absorption spectra of 6 and 7 were measured. Taking 7 as an
example, laser irradiation at 640 nm led to transient absorption
spectra, as shown in Figure 12, from which a triplet excited
state life time t of 475 ns was obtained. Similarly, t of 6 was
ꢀ
to 7) may partly account for their higher O C and COH efficien-
2
cy due to the higher driving forces for electron transfer from
the excited state to O . Further studies on other BODIPYs are
2
underway and will undoubtedly help to better understand the
T
T
ꢀ
measured to be 1229 ns. Both lifetimes are long enough to
underlying mechanism for their sensitization of O C and COH.
2
&6
&
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DNA Photocleavage by a Cationic BODIPY Dye
The redox potentials were measured on an EG&G Model 283 Po-
tentiostat/Galvanostat in a three-electrode cell with a platinum-
wire working electrode, a platinum-plate counterelectrode, and
a SCE reference electrode. Cyclic voltammetry was conducted at
ꢀ1
a scan rate of 100 mVs in Ar-saturated, anhydrous acetonitrile so-
lution containing 0.1m tetra-n-butylammonium hexafluorophos-
phate as supporting electrolyte.
Scheme 2. Proposed DNA photodamage mechanism for BODIPY 6.
The EPR spectra were taken at room temperature on a Bruker ESP-
3
00E spectrometer at 9.8 GHz, X-band with 100 kHz field modula-
tion. Samples were injected quantitatively into quartz capillaries
and illuminated in the cavity of the EPR spectrometer with an
Nd:YAG laser at 532 nm (5–6 ns pulse width, 10 Hz repetition fre-
quency, 30 mJ/pulse energy).
On the basis of the above discussion, the photodynamic
mechanism of 6 can be depicted as in Scheme 2. Though O2C
ꢀ
plays a role in the generation of COH, a restricting effect of
Methods: All experiments involving CT DNA were performed in
phosphate buffered saline (PBS, pH 7.4), unless otherwise noted.
CT DNA solutions were prepared by dispersing the desired amount
of DNA in buffer solution by stirring overnight at a temperature
below 48C. The concentration of CT DNA was expressed as the
concentration of nucleotides and was calculated by using the
SOD and catalase on DNA photocleavage was not observed
ꢀ
(
Figure 10). This suggests that their interaction with O C can
2
ꢀ
not compete with disproportionation of O C to COH under our
2
experimental conditions. Our work, for first time, demonstrates
1
that BODIPY dyes can serve as photosensitizers of not only O ,
2
ꢀ1
ꢀ1
but also O2Cꢀ and COH, which may open a new structure-opti-
molar absorptivity of 6600m cm at 260 nm.
1
1
mization strategy for more efficient BODIPY-based PDT agents.
Measurement of O quantum yield: The O quantum yields of the
2
2
examined BODIPYs were determined by the AMDA bleaching
1
method, taking MB as reference whose O quantum yield in aque-
2
[33]
ous solution was reported to be 0.52. The photooxidation of
AMDA sensitized by the examined BODIPYs was carried out by
using an Oriel 91192 solar simulator as light source and a 600 nm
long-pass optical filter to remove the short-wavelength light. Typi-
cally, 3 mL of BODIPY sample (ca. 5 mm, all samples were adjusted
to the same optical density at 650 nm) was mixed with 100 mL of
1 mm AMDA, and then subjected to photobleaching in a standard
3
. Conclusions
New NIR-absorbing cationic BODIPY dyes 6 and 7 were synthe-
sized, and their binding affinities and photocleavage activities
toward DNA characterized in detail. It is noteworthy that 6
photocleaves DNA through both O and COH, while 7 photo-
cleaves DNA mainly through O . The ability of a BODIPY dyes
1
2
1
2
1
cm path length quartz cuvette. The photoreactions were fol-
1
to generate O and COH was demonstrated for the first time in
2
lowed spectrophotometrically by detecting the absorbance de-
crease of AMDA at 400 nm as a function of irradiation time. All
samples were air-saturated and tested at room temperature.
this work, which is helpful for better understanding the
BODIPY-based PDT mechanism and for further development of
more efficient BODIPY-type PDT agents.
Gel electrophoretic DNA photocleavage: DNA photocleavage abili-
ties of the examined BODIPYs were evaluated by using supercoiled
pBR322 plasmid DNA as target. A mixture of 5 mL of supercoiled
pBR322 DNA (1 mm in base pairs) in PBS (pH 7.4), 5 mL BODIPY
Experimental Section
(
300 mm in CH CN), and 40 mL PBS (pH 7.4) was irradiated under an
3
Materials: 2,4-Dimethylpyrrole, 4-pyridinecarboxaldehyde, 2,3-di-
chloro-5,6-dicyano-1,4-benzoquinone (DDQ), boron trifluoride di-
ethyl etherate (BF ·Et O, 98%), azobisisobutyronitrile (AIBN), tetra-
n-butylammonium hexafluorophosphate, benzaldehyde, bromosuc-
cinimide (NBS), iodomethane, and piperidine were purchased from
Alfa Aesar. Catalase, ethidium bromide (EB), superoxide dismutase
Oriel 91192 Solar Simulator with a glass filter to cut off the light
below 600 nm. After irradiation, 20 mL of gel loading buffer was
added. The sample was then subjected to agarose gel (1%) elec-
trophoresis (Tris/acetic acid/EDTA buffer, pH 8.0) at 80 V for about
3
2
ꢀ
1
1
.5 h. The gel was stained with 1 mgL EB for 1 h, and then ana-
lyzed with a Gel Doc XR system (Bio-Rad).
(
(
(
SOD), sodium azide (NaN ), mannitol, 2,2,6,6-tetramethylpiperidine
TEMP), 5,5-dimethyl-1-pyrroline-N-oxide (DMPO), calf thymus DNA
CT DNA), gel loading buffer, and Tris base were products of
3
n-Octanol/water partition coefficients: n-Octanol/water partition
coefficients P were measured at room temperature by a reported
c
[34]
Sigma-Aldrich. The supercoiled pBR322 plasmid DNA was obtained
from TaKaRa Biotechnology Company. 9,10-Anthracenediyl-bis(me-
thylene)dimalonic acid (AMDA, ꢂ90%) was provided by Fluka.
Methylene blue (MB) was a product of Beijing Chemical Works.
method. In brief, solutions of BODIPYs (1 mm) in equal volumes
of PBS (pH 7.4, 1 mL) and n-octanol (1 mL) were mixed and sonicat-
ed for 30 min. After separation by centrifugation, the amounts of
BODIPY in each phase were determined by measuring the absorp-
tion spectra after dilution with acetonitrile, and the results were
the average of three independent measurements.
Spectroscopic measurements: UV/Vis absorption spectra were re-
corded on a Shimadzu UV-2450 spectrophotometer. Fluorescence
emission spectra were run on a Hitachi F-4500 fluorescence spec-
Viscosity measurements: Viscosity was measured with an Ubbe-
lohde viscometer immersed in a constant-temperature bath at
1
trophotometer. H NMR spectra were obtained on a Bruker DMX-
1
/3
4
00 spectrophotometer. High-resolution mass spectra (HRMS), EI
308C and a stopwatch. The data were presented as (h/h0) versus
[BODIPY]/[DNA], where h is the specific viscosity of DNA in the
mass spectra (EI-MS), and MALDI TOF mass spectra (MALDI TOF-
MS) were determined on a Bruker Daltonics Inc. APEX II FT-ICR
mass spectrometer, GCT Premier (Waters) mass spectrometer, and
Microflex (Bruker) mass spectrometer, respectively. Circular dichro-
ism (CD) spectra were recorded on a JASCO-J810 spectrometer.
presence of photosensitizer and h that of DNA alone in 5 mm PBS
0
(pH 7.2). Viscosities were calculated from the observed flow time of
DNA-containing solutions (t) corrected for the buffer alone (t ): h=
0
(tꢀt )/t .
0
0
ChemPhysChem 0000, 00, 1 – 10
ꢁ 2012 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
www.chemphyschem.org
&
7
&
ÞÞ
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X. S. Wang et al.
EB displacement assay: Aliquots (15 mL each time) of the BODIPY
give 20 mg of 4 or 110 mg of 5 as a dark green solid. Yield: 10%
1
or MB solution (1.0 mm in CH CN) were added to a 2 mL solution
for 4 and 46% for 5. BODIPY 4: H NMR (400 MHz, CDCl ): d=1.46
3
3
of EB (5 mm) and CT DNA (10 mm in base pairs) in PBS, and the mix-
tures allowed to stand for 15 min before fluorescence measure-
ments. The fluorescence from EB was measured with 490 nm exci-
tation. The apparent binding affinity Kapp was calculated from Equa-
tion (1), where [drug] is the concentration of the BODIPY or MB at
(s, 6H), 7.30–7.32 (m, 2H), 7.57–7.60 (m, 7H), 7.90 (d, J=16.7 Hz,
2H), 8.06 (d, J=16.7 Hz, 2H), 8.69 (d, J=6.0 Hz, 4H);. MALDI-TOF
1
MS: 658.37. BODIPY 5: H NMR (400 MHz, CDCl ): d=1.48 (s, 6H),
3
7.30–7.32 (m, 2H), 7.50–7.52 (m, 4H), 7.57–7.59 (m, 3H), 7.83 (d,
J=16.7 Hz, 2H), 8.06 (d, J=16.7 Hz, 2H), 8.68 (d, J=6.0 Hz, 4H);
MALDI-TOF MS: 754.44.
5
0% reduction of EB fluorescence and K the binding constant of
EB
7
ꢀ1 [35]
EB toward CT DNA (1ꢀ10 m ):
[36]
BODIPYs 6 and 7 were synthesized as follows: A mixture of 4
10 mg, 0.015 mmol) or 5 (50 mg, 0.066 mmol), 10 mL of iodome-
(
KEB½EBꢃ ¼ Kapp½drugꢃ
ð1Þ
thane, and 15 mL of DMF was stirred at 508C for 10 h. The reaction
mixture was then poured into 200 mL of diethyl ether, and the pre-
cipitate collected by filtration and recrystallized from DMF to give
Transient absorption and triplet excited state lifetime: Nanosecond
transient absorption measurements were performed on a LP-920
laser flash photolysis setup (Edinburgh). Excitation at 640 nm with
a power of 2.0 mJ/pulse from a computer-controlled Nd:YAG laser/
OPO system from Opotek (Vibrant 355 II) operating at 10 Hz was
directed to the sample with an optical absorbance of 0.4 at the ex-
citation wavelength. The laser and analyzing light beam passed
perpendicularly through a 1 cm quartz cell. The signals were de-
tected by a Tektronix TDS 3012B oscilloscope and R928P photo-
multiplier, and finally analyzed by Edinburgh analytical software
1
1 mg of 6 or 65 mg of 7 as a green solid. Yield: 77% for 6 and
1
9
6
1
5% for 7. BODIPY 6: H NMR (400 MHz, [D ]DMSO): d=1.44 (s,
6
H), 4.31 (s, 6H), 7.52–7.54 (m, 2H), 7.66–7.67 (m, 3H), 7.91 (d, J=
6.7 Hz, 2H), 8.13 (d, J=16.5 Hz, 2H), 8.33(d, J=6.5 Hz, 4H), 8.94 (d,
J=6.5 Hz, 2H); MALDI-TOF MS: 687.41; ESI-HRMS: m/z calcd for
2
4
+
1
C H BBr F N : 688.0809; found: 688.0815. BODIPY 7: H NMR
33
29
2
2
(
400 MHz, [D ]DMSO): d=1.47 (s, 6H), 4.31 (s, 6H), 7.50–7.53 (m,
6
2
2
H), 7.65–7.68 (m, 3H), 7.89 (d, J=16.7 Hz, 2H), 8.07 (d, J=16.5 Hz,
H), 8.33 (d, J=6.5 Hz, 4H), 8.93 (d, J=6.5 Hz, 2H); MALDI-TOF
(
LP920). All samples used in flash photolysis experiments were dea-
2
+
MS: 784.00; ESI-HRMS: m/z calcd for C H BI F N : 784.0532;
33
29
2
2
4
erated for 30 min with argon before measurements.
found: 784.0543.
Synthesis: BODIPY 1 was synthesized by using the general synthet-
[6a]
ic method for BODIPY dyes. Pyrrole (230 mL, 2.25 mmol, 214 mg)
and benzaldehyde (106 mg, 1.0 mmol) were dissolved in 200 mL of
absolute dichloromethane under nitrogen atmosphere. Two drops
of trifluoroacetic acid were added and solution was stirred at room
temperature until benzaldehyde was completely consumed, moni-
tored by TLC. Then 272 mg of DDQ (1.20 mmol) was added. After
stirring for another 15 min, 5 mL of triethylamine and 5 mL of
BF ·Et O were added. The reaction mixture was stirred at room
Acknowledgements
This work was financially supported by NNSFC (21172228,
21101163, 20873170) and CAS (KGCX2YW-389).
3
2
Keywords: DNA cleavage · dyes/pigments · photodynamic
therapy · photosensitizers · reactive oxygen species
temperature for 2 h and washed with deionized water three times.
After removal of organic solvent, the obtained solid was purified
by column chromatography on silica gel (CH Cl /hexane 1/4 as
2
2
1
eluent) to give 95 mg of 1 as a nacarat solid. Yield: 29%. H NMR
400 MHz, CDCl ): d=1.37 (s, 6H), 2.56 (s, 6H), 5.98 (s, 2H), 7.26–
[
1] a) T. J. Dougherty, C. J. Gomer, B. W. Henderson, G. Jori, D. Kessel, M.
Korbelik, J. Moan, Q. Peng, J. Natl. Cancer Inst. 1998, 90, 889–905;
b) I. J. Macdonald, T. J. Dougherty, J. Porphyrins Phthalocyanines 2001, 5,
105–129; c) D. E. J. G. J. Dolmans, D. Fukumura, R. K. Jain, Nat. Rev.
Cancer. 2003, 3, 380–387; d) C. A. Robertson, D. H. Evans, H. Abra-
hamse, J. Photochem. Photobiol. B 2009, 96, 1–8; e) J. P. Celli, B. Q.
Spring, I. Rizvi, C. L. Evans, K. S. Samkoe, S. Verma, B. W. Pogue, T. Hasan,
Chem. Rev. 2010, 110, 2795–2838.
(
3
7
.30 (m, 2H), 7.46–7.52 (m, 3H); EI-MS: m/z 324.16.
[12a–b]
BODIPY 2 was prepared as follows:
1 (220 mg, 0.68 mmol),
azoisobutyronitrile (AIBN, 223 mg, 1.36 mmol), and N-bromosucci-
nimide (NBS, 242 mg, 1.36 mmol) were heated to reflux in 15 mL
of CCl for 30 min. After removal of the solvent, the obtained solid
4
was subjected to column chromatography on silica gel with
[
2] a) R. Bonnett, Chem. Soc. Rev. 1995, 24, 19–33; b) S. K. Pushpan, S. Ven-
katraman, V. G. Anand, J. Sankar, D. Parmeswaran, S. Ganesan, T. K.
Chandrashekar, Curr. Med. Chem. 2002, 2, 187–207; c) M. Wainwright,
Anti-Cancer Agents Med. Chem. 2008, 8, 280–291; d) J. F. Lovell, T. W. B.
Liu, J. Chen, G. Zheng, Chem. Rev. 2010, 110, 2839–2857; e) M. R. Detty,
S. L. Gibson, S. J. Wagner, J. Med. Chem. 2004, 47, 3897–3915.
CH Cl /hexane (1/5) as eluent to give 265 mg of 2 as a nacarat
2
2
1
solid. Yield: 81%. H NMR (400 MHz, CDCl ): d=1.36 (s, 6H), 2.61
3
(
s, 6H), 7.24–7.26 (m, 2H), 7.47–7.59 (m, 3H); EI-MS: m/z 481.96.
[12c–d,13]
BODIPY 3 was prepared as follows:
Iodic acid (232 mg,
1
.32 mmol) dissolved in a 15 mL of water was added dropwise
over 20 min to a solution of 1 (195 mg, 0.60 mmol) and iodine
335 mg, 1.32 mmol) in 250 mL of ethanol. The mixture was heated
[3] a) P. K. Frederiksen, S. P. McIlroy, C. B. Nielsen, L. Nikolajsen, E. Skovsen,
M. Jørgensen, K. V. Mikkelsen, P. R. Ogilby, J. Am. Chem. Soc. 2005, 127,
255–269; b) T. Lu, P. Shao, I. Mathew, A. Sand, W. F. Sun, J. Am. Chem.
(
Soc. 2008, 130, 15782–15783; c) M. Obata, S. Hirohara, R. Tanaka, I. Ki-
noshita, K. Ohkubo, S. Fukuzumi, M. Tanihara, S. Yano, J. Med. Chem.
to reflux for 1 h. After removal of solvent, the crude product was
purified by silica gel column chromatography with CH Cl /hexane
2
2
2
009, 52, 2747–2753; d) Y. Y. Huang, P. Mroz, T. Zhiyentayev, S. K.
(
1/5) as eluent to afford 310 mg of 3 as bright red needles. Yield:
Sharma, T. Balasubramanian, C. Ruziꢂ, M. Krayer, D. Fan, K. E. Borbas, E.
Yang, H. L. Kee, C. Kirmaier, J. R. Diers, D. F. Bocian, D. Holten, J. S. Lind-
sey, M. R. Hamblin, J. Med. Chem. 2010, 53, 4018–4027.
1
8
7
9%. H NMR (400 MHz, CDCl ): d=1.38 (s,6H), 2.65 (s, 6H), 7.21–
3
.25 (m, 2H), 7.49–7.56 (m, 3H); EI-MS: m/z 575.95.
[
12,13]
[4] a) M. Wainwright, Chem. Soc. Rev. 1996, 25, 351–359; b) D. Ramaiaht, A.
Joy, N. Chandrasekhar, N. V. Eldho, S. Das, M. V. George, Photochem. Pho-
tobiol. 1997, 65, 783–790; c) V. Rapozzi, L. Beverina, P. Salice, G. A.
Pagani, M. Camerin, L. E. Xodo, J. Med. Chem. 2010, 53, 2188–2196.
BODIPYs 4 and 5 were synthesized as follows:
148 mg of 2 or
1
77 mg of
3 (0.308 mmol), 4-pyridinecarboxaldehyde (82 mg,
.77 mmol, 72 mL), 220 mL of glacial acetic acid, and 270 mL of pi-
peridine in 15 mL of benzene were heated to reflux for 16 h. After
removal of solvent, the crude product was purified by thin-layer
chromatography on silica gel with CHCl /CH OH (1/5) as eluent to
0
[
5] a) R. Ziessel, G. Ulrich, A. Harriman, New. J. Chem. 2007, 31, 496–501;
b) A. Loudet, K. Burgess, Chem. Rev. 2007, 107, 4891–4932; c) G. Ulrich,
R. Ziessel, A. Harriman, Angew. Chem. 2008, 120, 1202–1219; Angew.
3
3
&8
&
www.chemphyschem.org
ꢁ 2012 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
ChemPhysChem 0000, 00, 1 – 10
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DNA Photocleavage by a Cationic BODIPY Dye
Chem. Int. Ed. 2008, 47, 1184–1201; d) A. C. Benniston, G. Copley, H.
Lemmetyinen, N. V. Tkachenko, ChemPhysChem 2010, 11, 1685–1692.
[18] D. O. Frimannsson, M. Grossi, J. Murtagh, F. Paradisi, D. F. O’Shea, J. Med.
Chem. 2010, 53, 7337–7343.
[
6] a) W. W. Qin, M. Baruah, A. Stefan, M. Van der Auweraer, N. Boens, Chem-
PhysChem 2005, 6, 2343–2351; b) Y. Yamada, Y. Tomiyama, A. Morita, M.
Ikekita, S. Aokia, ChemBioChem 2008, 9, 853–856; c) J. G. Wang, Y. J.
Hou, C. Li, B. W. Zhang, X. S. Wang, Sens. Actuators B 2011, 157, 586–
[19] a) J. N. Demasa, G. A. Crosby, J. Phys. Chem. 1971, 75, 991–1024; b) D. P.
Kennedy, C. M. Kormos, S. C. Burdette, J. Am. Chem. Soc. 2009, 131,
8578–8536.
[20] J. Olmsted, J. Phys. Chem. 1979, 83, 2581–2584.
5
2
93; d) D. P. Wang, Y. Shiraish, T. Hirai, Chem. Commun. 2011, 47, 2673–
675; e) S. C. Dodani, S. C. Leary, P. A. Cobine, D. R. Winge, C. J. Chang, J.
[21] M. K e˛ pczy n´ ski, R. P. Pandian, K. M. Smith, B. Ehrenberg, Photochem. Pho-
tobiol. 2002, 76, 127–134.
Am. Chem. Soc. 2011, 133, 8606–8616; f) N. Boens, W. Qin, M. Baruah,
W. M. D. Borggraeve, A. Filarowski, N. Smisdom, M. Ameloot, L. Crovet-
to, E. M. Talavera, J. M. Alvarez-Pez, Chem. Eur. J. 2011, 17, 10924–
[22] N. A. Kuznetsova, N. S. Gretsova, O. A. Yuzhakova, V. M. Negrimovskii,
O. L. Kaliya, E. A. Luk’yanets, Russ. J. Gen. Chem. 2001, 71, 36–41.
[23] a) Y. Lion, M. Delmelle, A. Vorst, Nature 1976, 263, 442–443; b) J. Moan,
E. Wold, Nature 1979, 279, 450–451.
10934.
[
7] a) D. Kumaresan, R. P. Thummel, T. Bura, G. Ulrich, R. Ziessel, Chem. Eur.
J. 2009, 15, 6335–6339; b) T. Rousseau, A. Cravino, E. Ripaud, P. Leriche,
S. Rihn, A. D. Nicola, R. Ziessel, J. Roncali, Chem. Commun. 2010, 46,
[24] a) A. M. Pyle, J. P. Rehmann, R. Meshoyrer, C. V. Kumar, N. J. Turro, J. K.
Barton, J. Am. Chem. Soc. 1989, 111, 3051–3058; <lit b>D. S. Raja,
N. S. P. Bhuvanesh, K. Natarajan, Inorg. Chem. 2011, 50, 12852–12866.
[25] a) R. Lyng, T. Hꢃrd, B. Norden, Biopolymers 1987, 26, 1327–1345; b) S.
Allenmark, Chirality 2003, 15, 409–422; c) N. C. Garbett, P. A. Ragazzon,
J. B. Chaires, Nat. Protoc. 2007, 2, 3166–3172.
5082–5084.
[
8] R. P. Sabatini, T. M. McCormick, T. Lazarides, K. C. Wilson, R. Eisenberg,
D. W. McCamant, J. Phys. Chem. Lett. 2011, 2, 223–227.
[
9] a) J. Killoran, L. Allen, J. F. Gallagher, W. M. Gallagher, D. F. O’Shea, Chem.
Commun. 2002, 17, 1862–1863; b) A. Gorman, J. Killoran, C. O’Shea, T.
Kenna, W. M. Gallagher, D. F. O’Shea, J. Am. Chem. Soc. 2004, 126,
[26] a) S. Mahadevan, M. Palaniandavar, Inorg. Chem. 1998, 37, 693–700;
b) S. Mahadevan, M. Palaniandavar, Inorg. Chem. 1998, 37, 3927–3934;
c) R. W. Y. Sun, C. K. L. Li, D. L. Ma, J. J. Yan, C. N. Lok, C. H. Leung, N.
Zhu, C. M. Che, Chem. Eur. J. 2010, 16, 3097–3113.
10619–10631; c) W. M. Gallagher, L. T. Allen, C. O’Shea, T. Kenna, M. Hall,
A. Gorman, J. Killoran, D. F. O’Shea, Br. J. Cancer 2005, 92, 1702–1710;
d) A. Byrne, W. M. Gallagher, D. F. O’Shea, J. Am. Chem. Soc. 2005, 127,
[27] R. Rohs, H. Sklenar, R. Lavery, B. Roder, J. Am. Chem. Soc. 2000, 122,
2860–2866.
1
6360–16311; e) A. T. Byrne, A. E. O’Connor, M. Hall, J. Murtagh, K.
[28] S. Mettath, B. R. Munson, R. K. Pandey, Bioconjugate Chem. 1999, 10,
94–102.
[29] Y. Sun, Y. J. Hou, Q. X. Zhou, J. R. Chen, B. W. Zhang, X. S. Wang, J. Inorg.
Biochem. 2011, 105, 978–984.
O’Neill, K. M. Curran, K. Mongrain, J. A. Rousseau, R. Lecomte, S. McGee,
J. J. Callanan, D. F. O’Shea, W. M. Gallagher, Br. J. Cancer 2009, 101,
1565–1573.
[
[
10] N. Adarsh, R. R. Avirah, D. Ramaiah, Org. Lett. 2010, 12, 5720–5723.
11] T. Yogo, Y. Urano, Y. Ishitsuka, F. Maniwa, T. Nagano, J. Am. Chem. Soc.
[30] S. Hirayama, R. Ueda, K. Sugata, Energy. Convers. Mgmt. 1995, 36, 685–
688..
2
005, 127, 12162–12163.
[31] a) J. R. Harbour, M. L. Hair, J. Phys. Chem. 1978, 82, 1397–1399; b) S. J.
Xu, X. X. Zhang, S. Chen, M. H. Zhang, T. Shen, Photochem. Photobiol.
Sci. 2003, 2, 871–876.
[32] a) D. T. Sawyer, Acc. Chem. Res. 1981, 14, 393–400; b) V. V. Pavlishchuk,
A. W. Addison, Inorg. Chim. Acta 2000, 298, 97–102.
[33] a) J. M. Fernandez, M. D. Bilgin, L. I. Grossweiner, J. Photochem. Photo-
biol. B 1997, 37, 131–140; b) R. W. Redmond, J. N. Gamlin, Photochem.
Photobiol. 1999, 70, 391–475.
[
12] a) Z. Dost, S. Atilgan, E. U. Akkaya, Tetrahedron 2006, 62, 8484–8488;
b) S. Atilgan, Z. Ekmekci, A. L. Dogan, D. Guc, E. U. Akkaya, Chem.
Commun. 2006, 42, 4398–4400; c) S. Erbas, A. Gorgulu, M. Kocakusako-
gullari, E. U. Akkaya, Chem. Commun. 2009, 33, 4956–4958; d) S. Ozlem,
E. U. Akkaya, J. Am. Chem. Soc. 2009, 131, 48–49.
[
13] a) H. He, P. C. Lo, S. L. Yeung, W. P. Fong, D. K. P. Ng, J. Med. Chem. 2011,
5
4, 3097–3102; b) H. He, P. C. Lo, S. L. Yeung, W. P. Fong, D. K. P. Ng,
Chem. Commun. 2011, 47, 4748–4750.
14] S. G. Awuah, J. Polreis, V. Biradar, Y. You, Org. Lett. 2011, 13, 3884–3887.
[34] S. J. Wagner, A. Skripchenkol, D. Robinenel, J. W. Foley, L. Cincotta, Phot-
ochem. Photobiol. 1998, 67, 343–349.
[
[
[
15] P. R. Ogilby, Chem. Soc. Rev. 2010, 39, 3181–3209.
16] a) J. W. Hofman, F. Zeeland, S. Turker, H. Talsma, S. A. G. Lambrechts,
D. V. Sakharov, W. E. Hennink, C. F. Nostrum, J. Med. Chem. 2007, 50,
[35] M. Lee, A. L. Rhodes, M. D. Wyatt, S. Forrow, J. A. Hartley, Biochemistry
1993, 32, 4237–4245.
[36] a) P. Hambright, E. B. FleischerL, Inorg. Chem. 1970, 9, 1757–1761; b) P.
Hambright, T. Gore, M. Burton, Inorg. Chem. 1976, 15, 2314–2315.
[37] a) S. Mula, K. Elliott, A. Harriman, R. Ziessel, J. Phys. Chem. A 2010, 114,
10515–10522; b) J. Y. Liu, M. E. El-Khouly, S. Fukuzumi, D. K. P. Ng,
Chem. Asian J. 2011, 6, 174–179.
1485–1494; b) K. Kawai, Y. Osakada, M. Fujitsuka, T. Majima, J. Phys.
Chem. B 2007, 111, 2322–2326; c) A. Ikeda, Y. Doi, M. Hashizume, J. Ki-
kuchi, T. Konishi, J. Am. Chem. Soc. 2007, 129, 4140–4141.
ˇ
[
17] a) G. Viola, F. DallAcqua, N. Gabellini, S. Moro, D. Vedaldi, H. Ihmels,
ChemBioChem 2002, 3, 550–558; b) P. Wang, L. G. Ren, H. P. He, F. Liang,
X. Zhou, Z. Tan, ChemBioChem 2006, 7, 1155–1159; c) Q. X. Zhou, W. H.
Lei, Y. Sun, J. R. Chen, C. Li, Y. J. Hou, X. S. Wang, B. W. Zhang, Inorg.
Chem. 2010, 49, 4729–4731; d) Y. Sun, Y. J. Hou, Q. X. Zhou, W. H. Lei,
J. R. Chen, X. S. Wang, B. W. Zhang, Inorg. Chem. 2010, 49, 10108–
Received: March 15, 2012
10116.
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ChemPhysChem 0000, 00, 1 – 10
ꢁ 2012 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
www.chemphyschem.org
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ARTICLES
J. G. Wang, Y. J. Hou, W. H. Lei,
New photocleavage mechanism: New
cationic BODIPY dye 6 is found, for the
Q. X. Zhou, C. Li, B. W. Zhang, X. S. Wang*
first time, to be able to generate singlet
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oxygen ( O ), superoxide anion radical
2
Cꢀ) and hydroxyl radical (COH) simulta-
(O
2
DNA Photocleavage by a Cationic
BODIPY Dye through Both Singlet
Oxygen and Hydroxyl Radical: New
Insight into the Photodynamic
Mechanism of BODIPYs
neously on irradiation (lꢂ600 nm).
1
Both O
and
COH participate in photo-
2
cleavage of DNA by this BODIPY dye.
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www.chemphyschem.org
ꢁ 2012 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
ChemPhysChem 0000, 00, 1 – 10
ÝÝ
These are not the final page numbers!