Water-soluble cationic gallium(III) and indium(III) phthalocyanines for photodynamic therapy

Article abstract of DOI:10.1016/j.jinorgbio.2009.12.011

The new tetra-non-peripheral and peripheral 2-mercaptopyridine substituted gallium(III) and indium(III) phthalocyanine complexes (np-GaPc, p-GaPc, np-InPc and p-InPc) and their quaternized derivatives (Qnp-GaPc, Qp-GaPc, Qnp-InPc and Qp-InPc) have been sy

Full text of DOI:10.1016/j.jinorgbio.2009.12.011

Journal of Inorganic Biochemistry 104 (2010) 297–309
Contents lists available at ScienceDirect
Journal of Inorganic Biochemistry
journal homepage: www.elsevier.com/locate/jinorgbio
Water-soluble cationic gallium(III) and indium(III) phthalocyanines
for photodynamic therapy
a
a,b,*
Mahmut Durmu ßs , Vefa Ahsen
a
Gebze Institute of Technology, Department of Chemistry, PO Box 141, Gebze 41400, Kocaeli, Turkey
TUBITAK-Marmara Research Centre, Materials Institute, PO Box 21, Gebze 41470, Kocaeli, Turkey
b
a r t i c l e i n f o
a b s t r a c t
Article history:
Received 31 May 2009
Received in revised form 8 December 2009
Accepted 16 December 2009
Available online 18 January 2010
The new tetra-non-peripheral and peripheral 2-mercaptopyridine substituted gallium(III) and indium(III)
phthalocyanine complexes (np-GaPc, p-GaPc, np-InPc and p-InPc) and their quaternized derivatives
(Qnp-GaPc, Qp-GaPc, Qnp-InPc and Qp-InPc) have been synthesized and characterized. The quaternized
complexes show excellent solubility in water, which makes them potential photosensitizer for use in
photodynamic therapy (PDT) of cancer. Photophysical and photochemical properties of these phthalocy-
anines were investigated. General trends are described for quantum yields of photodegradation, fluores-
cence and fluorescence lifetimes as well as singlet oxygen quantum yields of these compounds. In this
study, the effects of the position of the substituents, the nature of the metal ion and quaternization of
the substituents on the photophysical and photochemical parameters of the gallium(III) and indium(III)
phthalocyanines are also reported. This study also presented the ionic gallium(III) and indium(III) phthal-
ocyanines strongly bind to bovine serum albumin (BSA).
Keywords:
Phthalocyanine
Water soluble
Gallium
Indium
Photodynamic therapy
Photosensitizer
Ó 2009 Elsevier Inc. All rights reserved.
1
. Introduction
these dyes. Closed shell and diamagnetic ions, such as Zn2+, Ga3+
and Si , give Pc complexes with both high triplet yields and long
lifetimes [11–14].
4
+
Phthalocyanines (Pc) and metallophthalocyanines (MPc) have
been studied in great deal for many years, mostly in terms of their
uses as dyes and catalysts [1–3]. Recently, they have also found
applications in many fields ranging from industrial [4], technolog-
ical [5,6] to medical [7,8]. The Pc macrocycle can engage most me-
tal ions in its cavity; hence scores of different MPc complexes have
been synthesized. The nature of the metal ion coordinated within
the Pc cavity plays a vital role in the properties, reactions and of
course, applications of the resulting MPc. MPc derivatives in which
the central metal is diamagnetic and non-transitional, are photoac-
tive, and are often employed in photosensitization and energy con-
version [9,10]. Worth emphasizing, is the Pcs’ application as
photosensitizers in the photodynamic therapy (PDT) of tumours.
MPc complexes have been proved as highly promising photosensi-
tizers for PDT due to their intense absorption in the red region of
the visible light. High triplet state quantum yields and long triplet
lifetimes are required for efficient sensitization of them. The
photophysical properties of the Pc dyes are strongly influenced
by the presence and nature of the central metal ion. Complex struc-
ture of Pc with transition metals gives short triplet lifetimes to
The photophysics and photochemistry of gallium(III) and indiu-
m(III) Pc complexes are well documented [15–18]. Such studies re-
veal that these complexes show great promise for photocatalytic
and photosensitizing applications [19,20]. However, studies on
water-soluble gallium(III) and indium(III) Pc complexes are scarce
in the literature [21,22].
In PDT administration, the drug is injected into the patient’s
blood stream, and since the blood itself is a hydrophilic system,
water solubility becomes crucial for a potential photosensitizer
in PDT. The advantages of MPcs bearing cationic substituents over
those with neutral and anionic substituents are numerous [23],
and include the following: (i) they are able to improve water solu-
bility and prevent aggregation [24]; aggregation seriously compro-
mises the PDT value of the photosensitizer, (ii) they are more
efficient as PDT agents [25] and also improve cell uptake [26],
(iii) they are selectively localized in the cell mitochondria, which
when impaired induces apoptosis [27].
Thiol-derivatized MPc complexes show rich spectroscopic and
photochemical properties. For example, they are known to absorb
at longer wavelengths (>700 nm) [28,29] than other MPc com-
plexes. Therefore these complexes have a very useful feature for
applications in optoelectronics, near-IR devices and PDT.
The advantages that go with cation-substituted MPcs are too
striking to overlook, hence our interest in these complexes. In this
*
Corresponding author. Address: Gebze Institute of Technology, Department of
Chemistry, PO Box 141, Gebze 41400, Kocaeli, Turkey. Tel.: +90 2626053106; fax:
+90 2626053101.
E-mail address: ahsen@gyte.edu.tr (V. Ahsen).
0
162-0134/$ - see front matter Ó 2009 Elsevier Inc. All rights reserved.
doi:10.1016/j.jinorgbio.2009.12.011

2
98
M. Durmu ßs , V. Ahsen / Journal of Inorganic Biochemistry 104 (2010) 297–309
work, the synthesis, photophysical and photochemical studies on
new non-peripheral and peripheral tetra-substituted non-ionic
and quaternized ionic gallium(III) and indium(III) Pc complexes
furan (THF), acetone, ethanol and dimethylformamide (DMF)
were dried as described in Perrin and Armarego [32] before use.
Zinc phthalocyanine, anhydrous gallium(III) chloride, anhydrous
(Schemes 1 and 2) are presented. The aim of our ongoing research
2 3
indium(III) chloride, K CO , dimethylsulphate (DMS) were pur-
is to synthesis water-soluble gallium(III) and indium(III) Pc com-
plexes to be used as potential PDT agents, since gallium(III) and in-
dium(III) Pc complexes show good photophysical and
photochemical properties which are very useful for PDT studies.
Herein, we report the synthesis, characterization and spectroscopic
behaviour as well as photophysical (fluorescence quantum yields
and lifetimes) and photochemical (singlet oxygen and photodegra-
dation quantum yields) properties of gallium(III) and indium(III) Pc
complexes tetra-substituted at the non-peripheral (np-GaPc and
np-InPc) and peripheral (p-GaPc and p-InPc) positions with 2-
mercaptopyridine group (Scheme 1) and their quaternized (Qnp-
GaPc and Qnp-InPc for non-peripheral, Qp-GaPc and Qp-InPc for
peripheral position) complexes (Scheme 2).
Bovine serum albumin (BSA) and human serum albumin (HSA)
are major plasma proteins, which contribute significantly to phys-
iological functions and display effective drug delivery roles [30,31],
hence the investigation of binding of drugs with albumin is of
interest. A spectroscopic investigation of the binding of the
water-soluble gallium(III) and indium(III) Pc complexes (Qnp-
GaPc, Qp-GaPc, Qnp-InPc and Qp-InPc) to BSA is also presented
in this work.
chased from Aldrich. 2-Mercaptopyridine, 1,8-diazabicyclo[5.4.0]
undec-7-ene (DBU), 9,10-antracenediyl-bis(methylene)dimalonoic
acid (ADMA) and 1,3-diphenylisobenzofuran (DPBF) were pur-
chased from Fluka. Preparative thin layer chromatography was
performed on silica gel 60 P F254. 3-Nitrophthalonitrile (1) [33],
4-nitrophthalonitrile (2) [34], 3-(2-mercaptopyridine) phthalonit-
rile (3) and 4-(2-mercaptopyridine) phthalonitrile (4) [35] were
synthesized and purified according to literature procedures.
2.2. Equipment
Absorption spectra in the UV–visible region were recorded with
a Shimadzu 2001 UV spectrophotometer. Fluorescence excitation
and emission spectra were recorded on a Varian Eclipse spectroflu-
orometer using 1 cm pathlength cuvettes at room temperature. IR
spectra (KBr pellets) were recorded on a Bio-Rad FTS 175C FTIR
spectrometer. The mass spectra were acquired on a Bruker Dalton-
ics (Bremen, Germany) MicrOTOF mass spectrometer equipped
with electronspray ionization (ESI) source. The instrument was
operated in positive ion mode using a m/z range of 50–3000. The
capillary voltage of the ion source was set at 6000 V and the capil-
lary exit at 190 V. The nebulizer gas flow was 1 bar and drying gas
flow 8 mL/min. The drying temperature was set at 200 °C for quat-
2
. Experimental
ernized complexes (Qnp-GaPc, Qp-GaPc, Qnp-InPc and Qp-InPc).
1
2.1. Materials
6
H NMR spectra were recorded in DMSO-d solutions on a Varian
5
00 MHz spectrometer. Elemental analyses were obtained with a
Quinoline, DMSO, methanol, n-hexane, ethylacetate, CCl
4
, dieth-
Thermo Finnigan Flash 1112 Instrument. Although the elemental
analysis data for one or two atoms for several compounds are
3
ylether, chloroform (CHCl ), dichloromethane (DCM), tetrahydro-
RS
SR
N
CN
CN
N
N
N
N
CN
CN
RSH, DMF
K CO , RT
MCl , quinoline
3
M
N
N
2
3
180 C, 7 hrs
O
NO2
N
SR
SR
1
3
SR
M= Ga(III)
In(III)
np-GaPc
np-InPc
SR
N
N
O N
CN
CN
CN
N
N
N
2
RSH, DMF RS
K CO , RT
MCl , quinoline
3
RS
M
N
N
N
SR
CN
2
3
180 C, 7 hrs
O
2
4
SR
R
=
M= Ga(III)
In(III)
p-GaPc
p-InPc
N
Scheme 1. Synthesis of tetra-2-mercaptopyridine substituted gallium(III) and indium(III) phthalocyanine complexes.

M. Durmu ßs , V. Ahsen / Journal of Inorganic Biochemistry 104 (2010) 297–309
299
4
+
RS
R'S
SR
N
N
N
SR'
N
N
N
N
N
Dimethylsulphate, DMF
N
N
M
N
N
M
N
2SO42-
N
O
1
20 C, 12h
N
SR
N
SR'
SR
SR'
M= Ga(III)
In(III)
np-GaPc
np-InPc
M= Ga(III)
In(III)
Qnp-GaPc
Qnp-InPc
SR
SR'
4
+
N
N
N
N
N
N
N
N
Dimethylsulphate, DMF
SR' 2SO42-
RS
M
N
N
N
SR
R'S
N
N
M
N
N
O
1
20 C, 12h
N
SR
SR'
M= Ga(III)
In(III)
p-GaPc
p-InPc
M= Ga(III)
In(III)
Qp-GaPc
Qp-InPc
R
=
=
N
R'
+ N
CH3
Scheme 2. Synthesis of tetra-2-mercaptopyridine substituted quaternized gallium(III) and indium(III) phthalocyanine complexes.
somewhat (>0.5%) unsatisfactory, the ESI-MS results and other
spectroscopic analysis ( H NMR, IR) data reasonably support the
employed as the standard. The absorbance of the solutions at the
excitation wavelength ranged between 0.04 and 0.05.
1
chemical formulas.
Natural radiative life times (
chemCAD program which uses the Strickler–Berg equation [39].
The fluorescence lifetimes ( ) were evaluated using Eq. (2).
0
s ) were determined using Photo-
Photo-irradiations were done using a General Electric quartz
line lamp (300 W). A 600 nm glass cut off filter (Schott) and a water
filter were used to filter off ultraviolet and infrared radiations
respectively. An interference filter (Intor, 670 or 700 nm with a
band width of 40 nm) was additionally placed in the light path be-
fore the sample. Light intensities were measured with a POWER
MAX5100 (Molelectron detector incorporated) power meter.
s
F
s
s
F
U
F
¼
ð2Þ
0
2
2
.4. Photochemical parameters
.4.1. Singlet oxygen quantum yields
Singlet oxygen quantum yield (UD) determinations were car-
2
2
.3. Photophysical parameters
ried out using the experimental set-up described in the literature
[40,41]. Typically, a 3 mL portion of the respective unsubstituted,
peripheral and non-peripheral tetra-substituted gallium(III) and
indium(III) Pc solutions (absorbance ꢁ1.0 at the irradiation wave-
length) containing the singlet oxygen quencher was irradiated in
the Q band region with the photo-irradiation set-up described in
.3.1. Fluorescence quantum yields and lifetimes
Fluorescence quantum yields (
comparative method (Eq. (1)) [36,37],
U
F
) were determined by the
2
F ꢀ AStd
ꢀ
g
g
U
F
¼
U
F
ðStdÞ F
ð1Þ
2
Std
Std ꢀ A ꢀ
D
references [40,41]. U values were determined in air using the rel-
ative method with ZnPc (in DMSO) or ZnPcSmix (in aqueous media)
as references. DPBF and ADMA were used as chemical quenchers
for singlet oxygen in DMSO and aqueous media, respectively. Eq.
(3) was employed for the calculations:
where F and FStd are the areas under the fluorescence emission
curves of the samples (Schemes 1 and 2) and the standard, respec-
tively. A and AStd are the respective absorbances of the samples and
standard at the excitation wavelengths, respectively.
the refractive indices of solvents used for the sample and standard,
2
2
Std
g
and
g
are
Std
R ꢀ I
Std
D
abs
UD
¼
U
ð3Þ
Std
R
ꢀ Iabs
respectively. Unsubstituted ZnPc (in DMSO) ( = 0.18) [38] was
U
F

3
00
M. Durmu ßs , V. Ahsen / Journal of Inorganic Biochemistry 104 (2010) 297–309
Std
BSA
where
U
is the singlet oxygen quantum yields for the standard
Qp-GaPc, Qnp-InPc and Qp-InPc) respectively; KSV , the Stern–Vol-
mer quenching constant; k , the bimolecular quenching constant;
and
D
Std
Std
ZnPc (U
¼ 0:67 in DMSO) [42] and ZnPcSmix
(U
¼ 0:45 in aque-
q
D
D
ous media) [41], R and RStd are the DPBF (or ADMA) photobleaching
s
F(BSA), the fluorescence lifetime of BSA.
s
F(BSA) is known to be
obtained from the plots
versus [Pc], the value of k may be determined (Eq. (7)).
q
BSA
SV
rates in the presence of the respective samples (Schemes 1 and 2)
and standards, respectively. Iabs and I are the rates of light absorp-
10 ns [47–49], thus from the values of K
Std
abs
BSA
BSA
of F0 =F
tion by the samples and standards, respectively. To avoid chain
reactions induced by DPBF (or ADMA) in the presence of singlet
oxygen [43], the concentration of quenchers (DPBF or ADMA) was
2.5. Synthesis
ꢃ5
lowered to ꢁ3 ꢂ 10 M. Solutions of sensitizer (absorbance = 1 at
2.5.1. 1,(4)-Tetrakis-[(2-mercaptopyridine)phthalocyaninato]
chlorogallium(III) (np-GaPc, Scheme 1)
the irradiation wavelength) containing DPBF (or ADMA) were pre-
pared in the dark and irradiated in the Q band region using the
set-up described above. DPBF degradation at 417 nm and ADMA
degradation at 380 nm were monitored. The light intensity
A
mixture of anhydrous gallium(III) chloride (0.74 g,
4.22 mmol), 3-(2-mercaptopyridine) phthalonitrile (3) (1.00 g,
4.22 mmol), DBU (0.1 mL, 0.7 mmol) and quinoline (5 mL, doubly
distilled over CaH ) was stirred at 180 °C for 7 h under nitrogen
2
atmosphere. After cooling, the solution was dropped in the n-hex-
ane. The green solid product was precipitated and collected by fil-
tration and washed with n-hexane. The crude product was
dissolved in chloroform. After concentrating, the dark green waxy
product was precipitated with hot n-hexane. Furthermore the
crude green product was purified by column chromatography over
1
5
ꢃ1
ꢃ2
6
.65 ꢂ 10 photons s cm was used for
U
D
determinations.
2.4.2. Photodegradation quantum yields
Photodegradation quantum yield (
ried out using the experimental set-up described in literature
40,41]. Photodegradation quantum yields were determined using
d
U ) determinations were car-
[
Eq. (4):
Al
2 3
O using ethyl acetate/methanol (5:1) solvent system as an elu-
ðC
0
ꢃ C
t
Þ ꢀ V ꢀ N
A
U
d
¼
ð4Þ
ent. Green solid product was obtained. MP > 240 °C, Yield: 0.36 g
I
abs ꢀ S ꢀ t
(
32%). UV/visible (UV/Vis) (DMSO): kmax nm (log
e
) 346 (4.53),
ꢃ1
where C
0
and C
t
are the samples concentrations before and after
642 (4.22), 716 (4.88). IR [(KBr)
(C@N), 1570 (C@C), 1449, 1418, 1317, 1107, 908, 762, 744.
m
max/cm ]: 3045 (Ar-CH), 1595
1
irradiation respectively, V is the reaction volume, N the Avogadro’s
A
H
constant, S the irradiated cell area and t the irradiation time, Iabs is
NMR (DMSO-d , m = multiplet): d, ppm 9.62–9.57 (m, 4H, Pyri-
6
the overlap integral of the radiation source light intensity and the
dyl-H), 8.66–8.59 (m, 4H, Pyridyl-H), 8.34–8.09 (m, 8H, Pyridyl-H
and Pc-H), 7.86–7.78 (m, 8H, Pyridyl-H and Pc-H), 7.39–7.33 (m,
16
absorption of the samples. A light intensity of 2.23 ꢂ 10 pho-
ꢃ1
ꢃ2
tons s cm was employed for
U
d
determinations.
4H, Pc-H). Calc. for C52
28 12 4
H N S GaCl: C 59.24, H 2.68, N 15.94%;
Found: C 58.91, H 2.44 N 16.09%. MS (ESI-MS) m/z: Calc. 1054.1;
+
+
2
.4.3. Binding of quaternized gallium(III) and indium(III) Pc complexes
Found: 1055.0 [M+1] , 1019.2 [MꢃCl] .
to BSA
The binding of the quaternized gallium(III) and indium(III) Pc
complexes (Qnp-GaPc, Qp-GaPc, Qnp-InPc and Qp-InPc) to BSA
were studied by spectrofluorometry at room temperature. An
aqueous solution of BSA (fixed concentration) was titrated with
varying concentrations of the respective quaternized gallium(III)
and indium(III) Pc solutions. BSA was excited at 280 nm and fluo-
rescence recorded between 290 nm and 500 nm. The steady dimi-
nution in BSA fluorescence with increase in quaternized
gallium(III) and indium(III) Pc concentrations was noted and used
in the determination of the binding constants and the number of
binding sites on BSA, according to Eq. (5) [44–46].
2.5.2. 1,(4)-Tetrakis-[(2-mercaptopyridine)phthalocyaninato]
chloroindium(III) (np-InPc, Scheme 1)
Synthesis and purification was as outlined for np-GaPc except
anhydrous indium(III) chloride was employed instead of anhy-
drous gallium(III) chloride. The amounts of the reagents employed
were: 3 (1.00 g, 4.22 mmol), DBU (0.1 mL, 0.7 mmol), anhydrous
indium(III) chloride (0.93 g , 4.22 mmol) in quinoline (5 mL). The
crude green product was purified by column chromatography over
2 2
silica gel using ethyl acetate and then CH Cl /methanol (10:1) sol-
vent system as an eluent. MP > 240 °C, Yield: 0.45 g (39%). UV/Vis
(DMSO): kmax nm (log
e
) 345 (4.62), 642 (4.31), 714 (4.99). IR
ꢃ1
ꢀ
ꢁ
[
(KBr)
m
max/cm ]: 3049 (Ar-CH), 1627 (C@N), 1575 (C@C), 1449,
ðF ꢃ FÞ
0
1
log
¼ log K
b
þ n log½Pcꢄ
ð5Þ
6
1417, 1327, 1102, 898, 757, 742. H NMR (DMSO-d ): d, ppm
ðF ꢃ F Þ
1
9
.10–8.92 (m, 4H, Pyridyl-H), 8.69–8.51 (m, 4H, Pyridyl-H), 8.11–
7.93 (m, 8H, Pyridyl-H and Pc-H), 7.79–7.58 (m, 8H, Pyridyl-H
and Pc-H), 7.30–7.24 (m, 4H, Pc-H). Calc. for C52 InCl: C
where F
and presence of quaternized Pc complexes (Qnp-GaPc, Qp-GaPc,
Qnp-InPc and Qp-InPc) respectively; F , the fluorescence intensity
of BSA saturated with quaternized Pc complexes; K , the binding
constant; n, the number of binding sites on a BSA molecule; and
0
and F are the fluorescence intensities of BSA in the absence
28 12 4
H N S
56.81, H 2.57, N 15.29%; Found: C 56.41, H 2.19 N 15.83%. MS
1
+
(ESI-MS) m/z: Calc. 1098.2; Found: 1099.0 [M+1] .
b
[
Pc] the concentration of quaternized Pc complexes. Plots of
2.5.3. 2,(3)-Tetrakis-[(2-mercaptopyridine)phthalocyaninato]
chlorogallium(III) (p-GaPc, Scheme 1)
h
i
ðF
0
ꢃFÞ
log
against log[Pc] would provide the values of n (from the
ðFꢃF
1
Þ
Synthesis and purification was as outlined for np-GaPc except
slope) and K
b
(from the intercept).
4
-(2-mercaptopyridine) phthalonitrile (4) was employed instead
The changes in BSA fluorescence intensity were related to quat-
ernized Pc concentrations by the Stern–Volmer relationship (Eq.
(
of 3-(2-mercaptopyridine) phthalonitrile (3). The amounts of the
reagents employed were: 4 (1.00 g, 4.22 mmol), DBU (0.1 mL,
6)):
0
.7 mmol), anhydrous gallium(III) chloride (0.74 g, 4.22 mmol) in
BSA
0
BSA
quinoline (5 mL). The crude green product was purified by column
chromatography over silica gel using ethyl acetate and then
CH Cl /methanol (10:1) solvent system as an eluent. MP > 240 °C,
2 2
F
F
BSA
¼
1 þ KSV ½Pcꢄ
ð6Þ
ð7Þ
BSA
and KSV is given by Eq. (7):
Yield: 0.45 g (41%). UV/Vis (DMSO): kmax nm (log
e
) 362 (4.63),
ꢃ1
BSA
SV
622 (4.39), 695 (5.16). IR [(KBr)
m
max/cm ]: 3049 (Ar-CH), 1645
K
¼ k
q
s
FðBSAÞ
1
(
C@N), 1573 (C@C), 1448, 1417, 1309, 1118, 919, 764, 747. H
where F0 and FBSA are the fluorescence intensities of BSA in the
absence and presence of quaternized Pc complexes (Qnp-GaPc,
BSA
6
NMR (DMSO-d , br = broad): d, ppm 9.73–9.58 (m, 8H, Pyridyl-H),
8.57–8.49 (m, 8H, Pyridyl-H and Pc-H), 7.80 (br, 4H, Pyridyl-H),

M. Durmu ßs , V. Ahsen / Journal of Inorganic Biochemistry 104 (2010) 297–309
301
Table 1
8H, Pyridyl-H), 8.84–8.81 (m, 4H, Pc-H), 8.57–8.51 (m, 4H, Pyri-
dyl-H), 8.03 (br, 4H, Pyridyl-H), 7.75 (br, 4H, Pc-H), 7.52 (br, 4H,
Absorption, excitation and emission spectral data for unsubstituted, non-peripheral
and peripheral tetra-substituted gallium(III) and indium(III) phthalocyanine com-
plexes in DMSO and water.
Pc-H). Calc. for C52
28 12 4
H N S InCl: C 56.81, H 2.57, N 15.29%; Found:
C 56.35, H 2.20 N 15.90%. MS (ESI-MS) m/z: Calc. 1098.2; Found:
Compound Solvent Q band (log
e)
Excitation Emission Stokes shift
Ex, (nm) Em, (nm)
+
1
099.0 [M+1] .
k
max
,
k
k
Stokes
D ,
(nm)
(nm)
GaPca
np-GaPc
p-GaPc
DMSO
DMSO
DMSO
680 5.15
716 4.88
695 5.16
701 5.02
680
717
694
699
691
737
708
708
702
702
697
700
732
712
711
–
11
21
13
7
16
13
12
14
18
17
8
2.5.5. 1,(4)-Tetrakis-[(N-methyl-2-
mercaptopyridine)phthalocyaninato] chlorogallium(III) sulphate
(Qnp-GaPc, Scheme 2)
Qnp-GaPc DMSO
Compound np-GaPc (190 mg, 0.177 mmol) was heated to
120 °C in freshly distilled DMF (5 mL) and excess DMS (0.168 mL,
1.77 mmol) was added dropwise. The mixture was stirred at
2
H O
686, 653 4.52, 4.53 689
Qp-GaPc
DMSO
689 4.94
685 4.83
686 4.46
714 4.99
695 5.17
689
686
689
727
697
696
–
2
H O
InPca
np-InPc
p-InPc
DMSO
DMSO
DMSO
1
20 °C for 12 h. After this time, the mixture was cooled to room
temperature and the product was precipitated with hot acetone
and collected by filtration. The green solid product was washed
successively with hot ethyl acetate, CHCl , n-hexane, CCl and
3 4
diethylether. The resulting hygroscopic product was dried over
Qnp-InPc DMSO
703 4.95
2
H O
685, 652 4.59, 4.57
692 4.80
686, 649 4.40, 4.46 700
–
19
15
Qp-InPc
DMSO
700
711
701
2
H O
phosphorous pentoxide. MP > 240 °C, Yield: 0.16 g (86%). UV/Vis
a
Data from [55].
(DMSO): kmax nm (log
e
) 337 (4.60), 631 (4.30), 701 (5.02). IR
ꢃ1
[
(KBr)
mmax/cm ]: 3046 (Ar-CH), 2953 (CH), 1614 (C@N), 1564
1
(
C@C), 1491, 1245 (S@O), 1105 (S@O), H NMR (DMSO-d
6
): d,
).
O): C 48.16, H 3.61, N 12.03%;
Found: C 48.83, H 3.17, N 12.77%. MS (ESI-MS) m/z: Calc. for
7
C
2
.49–7.45 (m, 4H, Pc-H), 7.30 (br, 4H, Pc-H). Calc. for
GaCl: C 59.24, H 2.68, N 15.94%; Found: C 58.67, H
.79 N 16.22%. MS (ESI-MS) m/z: Calc. 1054.1; Found: 1055.0
ppm 8.58–8.10 (m, 28H, Pc-H and Pyridyl-H), 4.91 (m, 12H, CH
Calc. for C56 GaCl (+5H
3
52
28 12 4
H N S
H
50
N
12
O
S
13
6
2
+
+
+
[
M+1] , 1019.2 [MꢃCl] .
56 40 12 8 6 4
C H N O S GaCl, 1306.5; Found 1079.3 [M-Cl-2SO ] .
2.5.4. 2,(3)-Tetrakis-[(2-mercaptopyridine)phthalocyaninato]
2.5.6. 1,(4)-Tetrakis-[(N-methyl-2-
chloroindium(III) (p-InPc, Scheme 1)
mercaptopyridine)phthalocyaninato] chloroindium(III) sulphate
(Qnp-InPc, Scheme 2)
Synthesis and purification was as outlined for np-GaPc except
4
-(2-mercaptopyridine) phthalonitrile (4) was employed instead
Synthesis and purification was as outlined for Qnp-GaPc except
compound np-InPc was employed instead of compound np-GaPc.
The amounts of the reagents employed were: np-InPc (0.190 g,
0.177 mmol), excess DMS (0.168 mL, 1.77 mmol) in DMF (5 mL).
of 3-(2-mercaptopyridine) phthalonitrile (3) and anhydrous indiu-
m(III) chloride was employed instead of anhydrous gallium(III)
chloride. The amounts of the reagents employed were: 4 (1.00 g,
4
.22 mmol), DBU (0.1 mL, 0.7 mmol), anhydrous indium(III) chlo-
MP > 240 °C, Yield: 0.15 g (77%). UV/Vis (DMSO): kmax nm (log
e
)
ꢃ1
ride (0.93 g, 4.22 mmol) in quinoline (5 mL). The crude green prod-
uct was purified by column chromatography over silica gel using
ethyl acetate and then CH
an eluent. MP > 240 °C, Yield: 0.53 g (46%). UV/Vis (DMSO): kmax
nm (log
323 (4.60), 632 (4.29), 703 (4.95). IR [(KBr)
m
max/cm ]: 3055 (Ar-
CH), 2948 (CH), 1614 (C@N), 1563 (C@C), 1486, 1238 (S@O),
1
2
Cl
2
/methanol (10:1) solvent system as
1159 (S@O), H NMR (DMSO-d
H and Pyridyl-H), 4.86 (m, 12H, CH
(+4H O): C 47.24, H 3.40, N 11.81%; Found: C 47.67, H 3.09, N
12.56%. MS (ESI-MS) m/z: Calc. for C56 InCl, 1351.6;
6
): d, ppm 9.32–7.86 (m, 28H, Pc-
3 48 12 12 6
). Calc. for C56H N O S InCl
e) 372 (4.72), 623 (4.44), 695 (5.17). IR [(KBr)
m
max
/
2
ꢃ1
cm ]: 3054 (Ar-CH), 1601 (C@N), 1573 (C@C), 1448, 1417, 1338,
40 12 8 6
H N O S
118, 911, 760, 742. 1H NMR (DMSO-d
+
1
6
): d, ppm 9.37–9.08 (m,
4
Found 1123.3 [M-Cl-2SO ] .
2
1
0
.5
p-InPc
p-GaPc
2
.5
1
np-InPc
np-GaPc
p-GaPc
np-InPc
p-InPc
np-GaPc
p-InPc
np-InPc
p-GaPc
.5
np-GaPc
0
300
400
500
600
700
800
Wavelength (nm)
Fig. 1. Absorption spectra of tetra-substituted gallium(III) and indium(III) phthalocyanines (np-GaPc, p-GaPc, np-InPc and p-InPc) in DMSO. Concentration = 1 ꢂ 10ꢃ5 M.

3
02
M. Durmu ßs , V. Ahsen / Journal of Inorganic Biochemistry 104 (2010) 297–309
Qnp-GaPc-in DMSO
1
1
1
.6
.4
.2
1
(
a)
Qp-GaPc-in DMSO
Qnp-GaPc-in DMSO
Qnp-GaPc-in water
Qp-GaPc-in DMSO
Qp-GaPc-in water
Qp-GaPc-in water
Qnp-GaPc-in DMSO
Qp-GaPc-in water
Qnp-GaPc-in water
Qp-GaPc-in DMSO
0.8
0.6
0.4
0.2
0
Qnp-GaPc-in water
300
400
500
600
700
800
Wavelength (nm)
1
.4
.2
1
(b)
Qnp-InPc-in DMSO
Qnp-InPc-in water
Qp-InPc-in DMSO
Qp-InPc-in water
1
Qnp-InPc-in DMSO
Qp-InPc-in DMSO
0.8
0.6
0.4
0.2
0
Qnp-InPc-in water
Qp-InPc-in water
Qp-InPc-in DMSO
Qp-InPc-in water
Qnp-InPc-in DMSO
Qnp-InPc-in water
300
400
500
600
700
800
Wavelength (nm)
Fig. 2. Absorption spectra of: (a) tetra-substituted quaternized gallium(III) phthalocyanines (Qnp-GaPc and Qp-GaPc) and (b) tetra-substituted quaternized indium(III)
ꢃ5
phthalocyanines (Qnp-InPc and Qp-InPc) in DMSO and water. Concentration = 1 ꢂ 10 M.
2.5.7. 2,(3)-Tetrakis-[(N-methyl-2-
The amounts of the reagents employed were: p-InPc (0.190 g,
mercaptopyridine)phthalocyaninato] chlorogallium(III) sulphate (Qp-
GaPc, Scheme 2)
Synthesis and purification was as outlined for Qnp-GaPc except
compound p-GaPc was employed instead of compound np-GaPc.
The amounts of the reagents employed were: p-GaPc (0.190 g,
0.177 mmol), excess DMS (0.168 mL, 1.77 mmol) in DMF (5 mL).
MP > 240 °C, Yield: 0.14 g (72%). UV/Vis (DMSO): kmax nm (log
e
)
ꢃ1
361 (4.46), 625 (4.15), 692 (4.80). IR [(KBr)
m
max/cm ]: 3055 (Ar-
CH), 2953 (CH), 1616 (C@N), 1565 (C@C), 1489, 1226 (S@O),
1
1111 (S@O), H NMR (DMSO-d
H and Pyridyl-H), 4.57 (m, 12H, CH
(+H O): C 49.11, H 3.09, N 12.27%; Found: C 49.17, H 3.79, N
12.70%. MS (ESI-MS) m/z: Calc. for C56 InCl, 1351.6;
6
): d, ppm 9.47–7.46 (m, 28H, Pc-
0
.177 mmol), excess DMS (0.168 mL, 1.77 mmol) in DMF (5 mL).
3 42 12 9 6
). Calc. for C56H N O S InCl
MP > 240 °C, Yield: 0.15 g (81%). UV/Vis (DMSO): kmax nm (log
e
)
2
ꢃ1
3
60 (4.49), 619 (4.19), 689 (4.94). IR [(KBr)
mmax/cm ]: 3052 (Ar-
40 12 8 6
H N O S
+
CH), 2949 (CH), 1616 (C@N), 1565 (C@C), 1493, 1223 (S@O),
4
Found 1123.3 [M-Cl-2SO ] .
1
1
094 (S@O), H NMR (DMSO-d
6
): d, ppm 9.99–7.35 (m, 28H, Pc-
). Calc. for C56 GaCl
O): C 48.16, H 3.61, N 12.03%; Found: C 48.73, H 3.31, N
2.95%. MS (ESI-MS) m/z: Calc. for C56 GaCl, 1306.5;
H and Pyridyl-H), 4.53 (m, 12H, CH
3
50 12 13 6
H N O S
(
+5H
2
3. Results and discussion
1
40 12 8 6
H N O S
+
Found 1079.3 [M-Cl-2SO
4
] .
3.1. Ground state electronic absorption and fluorescence spectra
2
.5.8. 2,(3)-Tetrakis-[(N-methyl-2-
The electronic spectra of the studied gallium(III) and indium(III)
Pc complexes showed characteristic absorption in the Q band re-
gion at around 685–716 nm in DMSO, Table 1. The B-bands were
observed at around 350 nm (Figs. 1 and 2). The spectra showed
monomeric behaviour evidenced by a single (narrow) Q band,
mercaptopyridine)phthalocyaninato] chloroindium(III) sulphate (Qp-
InPc, Scheme 2)
Synthesis and purification was as outlined for Qnp-GaPc except
compound p-InPc was employed instead of compound np-GaPc.

M. Durmu ßs , V. Ahsen / Journal of Inorganic Biochemistry 104 (2010) 297–309
303
typical of metallated phthalocyanine complexes in DMSO [50]. In
DMSO, the Q bands were observed at: 716 (np-GaPc), 695 (p-
GaPc), 701 (Qnp-GaPc), 689 (Qp-GaPc), 714 (np-InPc), 695 (p-
InPc), 703 (Qnp-InPc), and 692 (Qp-InPc), Table 1. The red-shifts
were observed for gallium(III) and indium(III) Pc complexes fol-
lowing substitution. In DMSO, the quaternization caused 15 nm
and 11 nm blue-shifts for non-peripheral gallium(III) and indiu-
m(III) Pc complexes, respectively. The quaternization also caused
The Q bands of the non-peripheral substituted complexes are
red-shifted when compared to the corresponding peripheral
substituted complexes in DMSO (Figs. 1 and 2). The red-shifts are
21 nm between np-GaPc and p-GaPc, 12 nm between Qnp-GaPc
and Qp-GaPc, 19 nm between np-InPc and p-InPc, and 11 nm be-
tween Qnp-InPc and Qp-InPc. The observed red spectral shifts are
typical of Pcs with substituents at the non-peripheral positions and
have been explained in the literature [51,52]. The B-bands are
6
nm and 3 nm blue-shifts for peripheral gallium(III) and indiu-
1 2
broad due to the superimposition of the B and B bands in the
m(III) Pc complexes, respectively, which is smaller than non-
peripheral blue-shifts. The lone pair electrons of the nitrogen atom
of the substituents in non-quaternized complexes (np-GaPc, p-
GaPc, np-InPc and p-InPc) are delocalized into the ring and ulti-
mately leading to a red-shifting. However when these lone pair
electrons are engaged as in quaternized complexes (Qnp-GaPc,
Qp-GaPc, Qnp-InPc and Qp-InPc) and its mesomeric contribution
to the ring electron density is lost.
320 to 370 nm region. In water, the absorption spectra of quatern-
ized complexes (Qnp-GaPc, Qnp-InPc and Qp-InPc) showed cofa-
cial aggregation, as evidenced by the presence of two non-
vibrational peaks in the Q band region, Fig. 2a and b, Table 1. The
lower energy (red-shifted) bands at 686 for Qnp-GaPc, 685 for
Qnp-InPc and 686 nm for Qp-InPc are due to the monomeric spe-
cies, while the higher energy (blue-shifted) bands at 653 for Qnp-
GaPc, 652 for Qnp-InPc and 649 nm for Qp-InPc is due to the
0.6
0.5
0.4
0.3
0.2
0.1
0
Qp-InPc in water + Triton X
Qp-InPc in water + Triton X
Qp-InPc in water
Qp-InPc in water
Qp-InPc-in water
Qp-InPc-in water+Triton X
300
400
500
600
700
800
Wavelength (nm)
Fig. 3. Absorption spectral changes for complex Qp-InPc observed on addition of Triton X-100 (0.1 mL) in water. [Qp-InPc] = 1 ꢂ 10ꢃ5 M.
1.8
1.6
1.4
1.2
1
A
B
C
D
E
F
0.8
0.6
0.4
0.2
0
3
00
400
500
600
700
800
Wavelength (nm)
Fig. 4. Aggregation behaviour of Qp-GaPc in DMSO at different concentrations: 18 ꢂ 10 (A), 16 ꢂ 10ꢃ6 (B), 14 ꢂ 10ꢃ6 (C), 12 ꢂ 10ꢃ6 (D), 10 ꢂ 10ꢃ6 (E), 8 ꢂ 10ꢃ6 (F) M.
ꢃ6
(
Inset: Plot of absorbance versus concentration).

3
04
M. Durmu ßs , V. Ahsen / Journal of Inorganic Biochemistry 104 (2010) 297–309
aggregated species. Surprisingly, the Qp-GaPc did not show aggre-
gation in water (Fig. 2a). The Q band of Qp-InPc in DMSO is dis-
torted (Fig. 2b). We suggest that Qp-InPc complex is more
aggregated than Qnp-InPc in DMSO. Aggregation is generally
decreasing the intensity of the Pc molecules.
at different concentrations in DMSO (Fig. 4, for complex Qp-GaPc
as an example). The studied gallium(III) and indium(III) Pc com-
plexes did not show aggregation in DMSO.
Fig. 5 shows fluorescence emission, absorption and excitation
spectra of complexes p-GaPc and np-InPc in DMSO and Qnp-GaPc
in aqueous solution as examples of the studied gallium(III) and in-
dium(III) Pc complexes. Fluorescence emission peaks were listed in
Table 1. The observed Stokes shifts were within the region ob-
served for gallium(III) and indium(III) Pc complexes. All gallium(III)
Pc complexes (np-GaPc, p-GaPc, Qnp-GaPc and Qp-GaPc) showed
similar fluorescence behaviour in DMSO (Fig. 5a for complex p-
GaPc as an example). The excitation spectra were similar to
absorption spectra and both were mirror images of the fluorescent
spectra for all gallium(III) Pc complexes in DMSO. The proximity of
the wavelength of each component of the Q band absorption to the
Q band maxima of the excitation spectra for all gallium(III) Pc com-
plexes suggest that the nuclear configurations of the ground and
excited states are similar and not affected by excitation. For
studied indium(III) Pc complexes (np-InPc, p-InPc, Qnp-InPc
Addition of Triton X-100 (0.1 mL) to an aqueous solution of
quaternized indium(III) Pc complexes (Qnp-InPc and Qp-InPc)
ꢃ5
(
Concentration = 1.0 ꢂ 10 ) brought about considerable increase
in intensity of the low energy side of the Q band (Fig. 3), suggesting
that the molecules are aggregated and that addition of Triton X-
1
00 breaks up the aggregates. But spectrum of non-peripheral
substituted quaternized gallium(III) Pc complex (Qnp-GaPc) did
not show any change with the addition of Triton X-100 to an aque-
ous solution of this complex. This suggests that Triton X-100 mol-
ecules could be penetrate among the indium(III) Pc complexes due
to the larger indium metal.
In this study, the aggregation behaviour of the gallium(III) and
indium(III) Pc complexes (np-GaPc, np-InPc, p-GaPc, p-InPc,
Qnp-GaPc, Qnp-InPc, Qp-GaPc and Qp-InPc) were investigated
1
000
1.6
(a)
Emission
8
6
4
2
00
00
00
00
0
Absorption
1.2
Excitation
0
0
0
.8
.4
500
550
600
650
700
750
800
850
Wavelength (nm)
1
1
1
1
60
1.2
1
(b)
40
20
00
Excitation
Absorption
0.8
80
60
40
20
0
0.6
Emission
0
0
0
.4
.2
500
550
600
650
700
750
800
850
Wavelength (nm)
7
6
5
4
3
2
1
00
0.5
0.4
Excitation
(c)
00
00
00
00
00
00
0
Absorption
0.3
0.2
0.1
0
Emission
750
5
00
550
600
650
700
800
850
Wavelength (nm)
Fig. 5. Absorption, excitation and emission spectra of: (a) p-GaPc in DMSO, (b) np-InPc in DMSO and (c) Qnp-GaPc in water. Excitation wavelengths: 660 nm for p-GaPc and
np-InPc and 625 nm for Qnp-GaPc.

M. Durmu ßs , V. Ahsen / Journal of Inorganic Biochemistry 104 (2010) 297–309
305
and Qp-InPc), the shapes of excitation spectra were different from
the absorption spectra in that the Q band of the excitation showed
splitting unlike the narrow Q band of the absorption in DMSO
(in both of DMSO and water) were higher than indium(III) Pc com-
plexes because of the indium is very heavy metal, Table 2. The
peripheral substituted complexes show marginally higher
F
U val-
(
Fig. 5b for complex np-InPc as an example). This suggests that
ues in both of DMSO and water, compared to non-peripheral
substituted derivatives, suggesting not as much of quenching of
the excited singlet state by peripheral substitution compared to
the non-peripheral substitution. The non-ionic phthalocyanine
there are changes in the molecule following excitation most likely
due to loss of symmetry, or change in aggregation status following
excitation. Thus the Q band maxima of the excitation and absorp-
tion spectra were different (Table 1) due to the differences in the
ground and excited state species. The difference in the behaviour
of indium(III) Pc complexes on excitation could be due to the larger
indium metal being more displaced from the core of the Pc ring,
and the displacement being more pronounced on excitation hence
a loss of symmetry. In aqueous media for the water-soluble quat-
ernized gallium(III) and indium(III) Pc complexes (Qnp-GaPc, Qp-
GaPc and Qp-InPc), the excitation spectra were different from
absorption spectra in that one Q band is observed instead of the
split Q band observed in absorption (Fig. 5c for complex Qnp-GaPc
as an example). This observation is typical of aggregated species.
Aggregated MPc complexes are not known [53] to fluoresce since
aggregation lowers the photoactivity of molecules through dissipa-
tion of energy by aggregates. The quaternized non-peripheral
substituted indium phthalocyanine complex (Qnp-InPc) showed
very low fluorescence in aqueous media.
complexes (np-GaPc, np-InPc, p-GaPc, p-InPc) show larger
F
U val-
ues compared to the corresponding quaternarized ionic Pc com-
plexes (Qnp-GaPc, Qnp-InPc, Qp-GaPc and Qp-InPc) in DMSO.
Comparing DMSO and water, the quaternarized ionic Pc complexes
(Qnp-GaPc, Qnp-InPc, Qp-GaPc and Qp-InPc) show larger
F
U val-
ues in DMSO than in water. This is attributed to the higher viscos-
ity of DMSO than that of water. As expected on grounds of the
higher viscosity of DMSO compared to that of water,
F
U values of
the quaternarized ionic complexes (Qnp-GaPc, Qnp-InPc, Qp-GaPc
and Qp-InPc) are higher in DMSO than in water. The relationship
between
F
U and the viscosity (g) of the solvent has been derived,
and is given by the Forster–Hoffmann [56]:
log ¼ C þ x log
U
F
g
ð8Þ
where C is a temperature-dependent constant and x, a fluorophore
dependent constant.
Fluorescence lifetime (
stays in its excited state before fluorescing, and its value is directly
related to that of ; i.e. the longer the lifetime, the higher the
F
s ) refers to the average time a molecule
3.2. Fluorescence lifetimes and quantum yields
U
F
quantum yield of fluorescence. Any factor that shortens the fluo-
rescence lifetime of a fluorophore indirectly reduces the value of
U . Such factors include internal conversion and intersystem cross-
F
ing. As a result, the nature and the environment of a fluorophore
determine its fluorescence lifetime.
The fluorescence quantum yields (
U
F
) for non-ionic gallium(III)
and indium(III) Pc complexes (np-GaPc, np-InPc, p-GaPc and p-
InPc) in DMSO and for quaternized ionic complexes (Qnp-GaPc,
Qnp-InPc, Qp-GaPc and Qp-InPc) in both DMSO and water are gi-
U values of all studied gallium(III) and indiu-
F
m(III) Pc complexes are typical of gallium(III) and indium(III) Pc
complexes [54,55]. The quaternized ionic Pc complexes showed
ven in Table 2. The
s
F
values (Table 2) were calculated using the Strickler–Berg
equation. The values of the substituted gallium(III) and indiu-
s
F
m(III) Pc complexes are lower compared to unsubstituted gal-
lium(III) and indium(III) Pc complexes in DMSO, suggesting more
aggregation in water. The
the mixture of the monomer-aggregated Pc species in the water.
The values of the substituted gallium(III) Pc complexes are low-
F
U values of these complexes are for
quenching by substitution.
F
s values are lower for non-peripheral
U
F
complexes (np-GaPc, np-InPc, Qnp-GaPc and Qnp-InPc) when
compared to peripheral complexes (p-GaPc, p-InPc, Qp-GaPc and
Qp-InPc), Table 2, suggesting more quenching by peripheral sub-
stitution compared to non-peripheral substitution. However the
er compared to unsubstituted gallium(III) Pc complex in DMSO,
which implies that the presence of the 2-mercaptopyridine substit-
uents certainly results in fluorescence quenching. For indium(III)
Pc complexes, the
F
U value of the peripheral substituted indiu-
s
F
values are typical for gallium(III) and indium(III) Pc complexes
m(III) Pc complex is higher than unsubstituted indium(III) Pc com-
plex while the non-peripheral substituted indium(III) Pc complex
is lower than unsubstituted indium(III) Pc complex which suggest-
ing more quenching by peripheral substitution compared to non-
[
53,54]. The values of gallium(III) Pc complexes (in both of
s
F
DMSO and water) were higher than indium(III) Pc complexes be-
cause of the indium is very heavy metal, Table 2.
The natural radiative lifetime (
fluorescence (k ) values are also given in Table 2. But there was
no clear trend found for and k values among the studied gal-
lium(III) and indium(III) Pc complexes.
0
s ) and the rate constants for
peripheral substitution. The
F
U values of gallium(III) Pc complexes
F
s
0
F
Table 2
Photophysical and photochemical parameters of unsubstituted, non-peripheral and
peripheral tetra-substituted gallium(III) and indium(III) phthalocyanine complexes in
DMSO and water.
3.3. Singlet oxygen quantum yields
Compound Solvent
U
F
s
F
(ns)
s
0
ak
(s
F
/108
)
U
10
d
/
U
D
ꢃ1
ꢃ4
(
ns)
Energy transfer between the triplet state of photosensitizers
GaPcb
np-GaPc
p-GaPc
DMSO
DMSO
DMSO
DMSO
0.30
0.20
0.22
0.11
0.021
0.12
0.074
0.018
0.017
0.029
0.0028
–
3.71
3.41
3.50
0.78
0.38
1.57
1.41
0.90
0.17
0.26
0.024
11.96 0.83
14.22 0.70
9.71 1.03
9.72 1.03
16.99 0.58
13.10 0.76
19.06 0.52
50.20 0.19
9.94 1.00
9.18 1.09
8.84 1.13
0.09
0.27
14.44
41.91
0.41
38.61
0.10
0.34
0.36
21.23
0.68
3.73
22.42
1.29
0.41
0.67
0.76
0.52
0.29
0.75
0.26
0.67
0.75
0.85
0.57
0.80
0.79
0.40
and ground state molecular oxygen leads to the production of sin-
glet oxygen. There is a necessity of high efficiency of transfer of en-
ergy between excited triplet state of MPc and ground state of
oxygen to generate large amounts of singlet oxygen, essential for
PDT.
The singlet oxygen quantum yields (U ), give an indication of
D
the potential of the complexes as photosensitizers in applications
where singlet oxygen is required, (e.g. for Type II mechanism).
Qnp-GaPc
H
2
O
Qp-GaPc
DMSO
H
2
O
InPcb
np-InPc
p-InPc
DMSO
DMSO
DMSO
DMSO
The
D
U values were determined using a chemical method (DPBF
Qnp-InPc
H
2
O
–
0.36
–
–
in DMSO and ADMA in water). The disappearance of DPBF or ADMA
was monitored using UV–vis spectrophotometer (Fig. 6a using
DPBF for complex p-GaPc in DMSO and Fig. 6b using ADMA for
complex Qnp-InPc in water). Many factors are responsible for
the magnitude of the determined quantum yield of singlet oxygen
Qp-InPc
DMSO
0.021
0.0008 <0.1
17.27 0.58
38.00 0.26
H
2
O
a
k
F
is the rate constant for fluorescence. Values calculated using k
F F F
= U /s .
b
Data from [55].

3
06
M. Durmu ßs , V. Ahsen / Journal of Inorganic Biochemistry 104 (2010) 297–309
0
0
0
0
0
0
0
.7
.6
.5
.4
.3
.2
.1
0
0.8
0.6
0.4
0.2
0
(a)
y = -0.0092x + 0.6446
2
R
= 0.9867
0
5
10
15
Time (s)
20
25
30
0
5
1
s (top)
s
0 s
15 s
0 s
5 s
0 s (bottom)
2
2
3
3
00
400
500
600
700
800
Wavelength (nm)
1
1
.4
.2
1
1
.4
(
b)
0
.8
y= -0.001x + 1.2553
R2 = 0.9824
1
.2
1
0.6
0
0
.4
.2
0
0
100
200
Time (s)
300
400
500
0.8
0.6
0.4
0.2
0
0
6
1
1
2
3
s (top)
0 s
20 s
80 s
40 s
00 s
360 s
20 s (bottom)
4
300
400
500
600
700
800
Wavelength (nm)
Fig. 6. A typical spectrum for the determination of singlet oxygen quantum yield of: (a) p-GaPc in DMSO using DPBF as a singlet oxygen quencher and (b) Qnp-InPc in water
ꢃ5
using ADMA as a singlet oxygen quencher. Concentration = 1 ꢂ 10 M. (Inset: plots of DPBF or ADMA absorbance versus time.)
1
1
.4
.2
1
1.2
1
0
.8
.6
.4
.2
y = -0.0058x + 1.1836
R2 = 0.966
0
0
2
s (top)
0 s
0
0
0
0
0
0
.8
.6
.4
.2
0
40 s
0 s
80 s (bottom)
0
6
0
10
20
30
40
Time (s)
50
60
70
80
300
400
500
600
700
800
Wavelength (nm)
Fig. 7. The photodegradation of p-InPc in DMSO showing the disappearance of the Q band and appearance of the reduction band at 20 s intervals. (Inset: plot of Q band
absorbance versus time.)
including; triplet excited state energy, ability of substituents and
solvents to quench the singlet oxygen, the triplet excited state life-
time and the efficiency of the energy transfer between the triplet
excited state and the ground state of oxygen. There was no de-
crease in the Q band of formation of new bands during
minations (Fig. 6).
D
U deter-
Table 2 shows that the values of
D
U are higher for substituted
gallium(III) and indium(III) Pc complexes (np-GaPc, np-InPc,

M. Durmu ßs , V. Ahsen / Journal of Inorganic Biochemistry 104 (2010) 297–309
307
A
[
Pc]=0
B
C
D
E
[
Pc]=saturated
F
G
290
340
390
440
490
Wavelength (nm)
Fig. 8. Fluorescence emission spectral changes of BSA (C = 3.00 ꢂ 10 M) on addition of varying concentrations of Qnp-InPc in water. [Qnp-InPc]: A = 0, B = 1.66 ꢂ 10ꢃ6
ꢃ5
,
ꢃ6
ꢃ6
ꢃ6
ꢃ6
C = 3.33 ꢂ 10 , D = 5.00 ꢂ 10 , E = 6.66 ꢂ 10 , F = 8.33 ꢂ 10 M, G = saturated with Qnp-InPc .
p-GaPc, p-InPc, Qnp-GaPc, Qp-GaPc and Qp-InPc) when com-
pared to respective unsubstituted gallium(III) and indium(III) Pc
except for complex Qnp-InPc which is lower than unsubstituted
InPc due to a little aggregation in DMSO. The peripheral substi-
tuted complexes (p-GaPc, Qp-GaPc, p-InPc, and Qp-InPc) are
absorption in this region has longer singlet oxygen lifetimes than
water, resulting in large values in DMSO. However the value
of for Qnp-InPc is higher in water (0.80) than in DMSO (0.57).
Quaternarized ionic gallium(III) and indium(III) Pc complexes
(Qnp-GaPc, Qp-GaPc, Qnp-InPc, and Qp-InPc) showed lower
U
D
U
D
U
D
higher
np-GaPc, Qnp-GaPc, np-InPc, and Qnp-InPc) in DMSO. On the
contrary in water, the non-peripheral substituted complexes (np-
GaPc, Qnp-GaPc, np-InPc, and Qnp-InPc) are higher values
when compared to the peripheral complexes (p-GaPc, Qp-GaPc,
p-InPc, and Qp-InPc).
U
D
values when compared to the non-peripheral complexes
values when compared to corresponding non-ionic complexes
(np-GaPc, p-GaPc, np-InPc, and p-InPc) in DMSO.
(
The
D
U values of indium(III) Pc complexes (in both of DMSO
U
D
and water) are higher than gallium(III) Pc complexes because of
the indium is very heavy metal. The populations of the molecules
on the triplet state are higher for indium(III) Pc complexes than
gallium(III) Pc complexes.
Table 2 shows that lower
solutions compared to in DMSO. The low
U
D
values are observed in aqueous
in water compared
U
D
to other solvents such as deuterated water and DMSO was ex-
plained [57] by the fact that singlet oxygen absorbs at 1270 nm,
and water, which absorbs around this wavelength has a great effect
on singlet oxygen lifetime, while DMSO which exhibits little
3.4. Photodegradation studies
Degradation of the molecules under irradiation can be used to
study their stability and this is especially important for those
Qnp-InPc
Qnp-GaPc
3
2.6
2.2
1.8
1.4
1
Qp-GaPc
Qp-InPc
0.00
1.00
2.00
3.00
4.00
5.00
6.00
7.00
8.00
9.00
[
Pc] (µM)
Fig. 9. Stern–Volmer plots of tetra substituted gallium(III) and indium(III) phthalocyanines quenching of BSA in water. [BSA] = 3.00 ꢂ 10 M in water. [Pc] = 0, 1.66 ꢂ 10ꢃ6
ꢃ5
,
ꢃ6
ꢃ6
ꢃ6
ꢃ6
3
.33 ꢂ 10 , 5.00 ꢂ 10 , 6.66 ꢂ 10 , 8.33 ꢂ 10 M.

3
08
M. Durmu ßs , V. Ahsen / Journal of Inorganic Biochemistry 104 (2010) 297–309
ꢃ4
molecules intended for use in photocatalysis. The collapse of the
absorption spectra without any distortion of the shape confirms
clean photodegradation not associated with phototransformation
into different forms of MPc absorbing in the visible region.
d
and 10 . The U values, found in this study, are similar gallium(III)
and indium(III) Pc complexes having different substituents on the
phthalocyanine ring in literature [20–22,54,55]. Stable zinc phtha-
ꢃ6
locyanine complexes show
U
d
values as low as 10 and for unsta-
ꢃ3
Fig. 7 shows that for gallium(III) and indium(III) Pc complexes,
there was a change in spectra following photodegradation. The
spectral changes involved a decrease in the Q band and an increase
in the absorption near 590 nm, suggesting that this band is due to
reduction products of the complexes. These spectral changes were
only observed in DMSO. The first ring reduction in MPc complexes
is characterized by a decrease in the Q band and the formation of
weak bands between 500 and 600 nm [58]. When the degradated
solutions of the gallium(III) and indium(III) Pc complexes were oxi-
dized with bromine as oxidation agent the intensity of the Q band
was increased and the reduction band at around 600 nm was dis-
appeared. Thus, we propose that during photodegradation, the gal-
lium(III) and indium(III) Pc complexes were partly transformed to
ble molecules, values of the order of 10 have been reported [53].
It seems studied gallium(III) and indium(III) Pc complexes show
also similar
d
U values and stability with zinc Pc complexes.
3.5. Interaction of quaternized ionic gallium(III) and indium(III)
phthalocyanine complexes with BSA
Fig. 8 shows the fluorescence emission spectra of BSA in the
presence of different concentrations of Qnp-InPc in water as an
example. The quaternized gallium(III) and indium(III) Pc com-
plexes are mixtures of aggregated and unaggregated species. The
total concentrations of the complexes are mixture of the monomer
and aggregated species. We calculated the percentage aggregation
ꢃ3
an anion (Pc ) species. This type of transformation has been ob-
served before during the photodegradation of gallium(III) and in-
dium(III) Pc complexes [54]. The suggested mechanism for the
(
% Agg) of these complexes using the equation described in the lit-
erature [59,60]. The aggregation percentages are 6% for Qnp-GaPc,
0% for Qnp-InPc, 50% for Qp-GaPc and 42% for Qp-InPc. The non-
1
ꢃ3
formation of Pc in the presence of H donors is shown by Eqs.
peripheral substituted Pc complexes having lower % Agg values
which may be due to reduced aggregation tendencies when the
substitution is at the non-peripheral position of the Pc skeleton
(
9)–(11):
3
! MPcꢅ
MPc þ h
m
ð9Þ
ð10Þ
ð11Þ
[
61]. The BSA fluorescence at 348 nm is mainly attributable to
3
MPc þ S ꢃ H ! MPc þ S ꢃ HÅþ
ꢅ
Åꢃ
tryptophan residues in the macromolecule. BSA and the respective
quaternized ionic Pc complexes exhibit reciprocated fluorescence
quenching on one another; hence it was possible to determine
Stern–Volmer quenching constants (KSV). The slope of the plots
shown at Fig. 9 gave KSV values and listed in Table 3, suggest that
BSA fluorescence quenching is more effective for quaternized non-
peripheral substituted Pc complexes (Qnp-GaPc and Qnp-InPc)
than quaternized peripheral substituted Pc complexes (Qp-GaPc
and Qp-InPc) in water. Using the approximate fluorescence life-
time of BSA (10 ns) [48,49], the bimolecular quenching constant
Åꢃ
Åꢃ
MPc þ O
S ¼ solvent
2
! MPc þ O
2
All the studied gallium(III) and indium(III) Pc complexes showed
about the same stability with
ꢃ3
d
U of the order of between 10
Table 3
Binding and fluorescence quenching data for interaction of BSA with quaternized
gallium(III) and indium(III) phthalocyanine complexes in water.
q
(k ) was determined (Eq. (7)). These values are of the order of
Compound
K
BSA=10
5
ðMꢃ1
Þ
k
q
/1013 (M
ꢃ1 ꢃ1
s
)
K
b
/10 (M
ꢃ6
ꢃ1
)
n
13
ꢃ1 ꢃ1
10
ꢃ1 ꢃ1
SV
10
M
s
, which exceed the proposed value of 10
M
s
Qnp-GaPc
Qp-GaPc
Qnp-InPc
Qp-InPc
2.28
1.79
2.62
1.28
2.28
1.79
2.62
1.28
4.78
5.49
4.22
7.41
1.26
1.33
1.24
1.23
for diffusion-controlled (dynamic) quenching (according to the
Einstein–Smoluchowski approximation) at room temperature
[
62]. This also, is an indication that the mechanism of BSA quench-
ing by quaternized Pc complexes (Qnp-GaPc, Qp-GaPc, Qnp-InPc
0.4
Qnp-InPc
Qnp-GaPc
Qp-GaPc
0
.15
0.1
Qp-InPc
-
-0.35
-0.6
-0.85
-5.9
-5.8
-5.7
-5.6
-5.5
-5.4
-5.3
-5.2
-5.1
-5
log[Pc]
ꢃ5
Fig. 10. Determination of tetra substituted gallium(III) and indium(III) phthalocyanine-BSA binding constant (and number of binding sites on BSA). [BSA] = 3.00 ꢂ 10 M and
ꢃ6
ꢃ6
ꢃ6
ꢃ6
ꢃ6
[
Pc] = 0, 1.66 ꢂ 10 , 3.33 ꢂ 10 , 5.00 ꢂ 10 , 6.66 ꢂ 10 , 8.33 ꢂ 10 M in water.

M. Durmu ßs , V. Ahsen / Journal of Inorganic Biochemistry 104 (2010) 297–309
309
and Qp-InPc) is not diffusion-controlled (i.e., not dynamic quench-
ing, but static quenching). The k values are larger for quaternized
non-peripheral substituted Pc complexes (Qnp-GaPc and Qnp-
InPc) than quaternized peripheral substituted Pc complexes (Qp-
GaPc and Qp-InPc) in water. The binding constants (K ) and num-
b
ber of binding sites (n) on BSA were obtained using Eq. (5) and the
results are shown in Table 3. The slope of the plots shown at Fig. 10
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[
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than the quaternized non-peripheral substituted Pc complexes
[
[
(Qnp-GaPc and Qnp-InPc), probably due to steric effect at the
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sity of tryptophan with quaternized Pc concentration indicates that
these complexes readily bind to BSA, which implies that the Pc
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photophysical and photochemical properties of new non-periphe-
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and quaternized ionic gallium(III) and indium(III) Pc complexes.
The effect of quaternization on these properties is also presented.
Solvent effect (DMSO or water) on the photophysical and photo-
chemical properties of the quaternized derivative is investigated.
Although, the photophysical and photochemical properties rele-
vant for photosensitization gave more attractive values in DMSO,
the values in water are still good enough for PDT applications. This
work will certainly enrich the hitherto scanty literature on the
potentials of cationic phthalocyanines as photosensitizers in PDT.
All studied gallium(III) Pc complexes (np-GaPc, p-GaPc, Qnp-GaPc
and Qp-GaPc) showed similar fluorescence behaviour with MPcs.
The difference in the behaviours of indium(III) Pc complexes on
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U
F
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complexes are very low due to presence of a heavier indium atom
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[
[
[
plexes have good
peripheral indium(III) Pc complex (Qnp-InPc) has high
U
D
values. In particular, quaternarized non-
value
0.80) in water (Photosens , which has been in use for PDT has
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D
of 0.42). Thus, these complexes show potential as Type II
4
photosensitizers for photodynamic therapy of cancer. Photoreduc-
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during photodegradation in DMSO. This study reveals that the
water-soluble complexes bind strongly to serum albumin; hence
they can easily be transported in the blood.
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Acknowledgement
This work was supported by Gebze Institute of Technology (Pro-
ject number: BAP-2007-A-01).
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