Investigation of novel substituted zinc and aluminium phthalocyanines for photodynamic therapy of epithelial breast cancer

Article abstract of DOI:10.1016/j.dyepig.2019.107592

A series of phthalonitrile ligands were synthesized by nucleophilic substitution reaction using the hydroxyl or sulfanyl group precursors and the nitro moiety of the nitrophthalonitrile to yield corresponding oxy or sulfanyl bridged ligands. These ligands were subsequently subjected to cyclocondensation reaction with diamagnetic metal ions like zinc and aluminium to afford symmetrically substituted zinc and aluminium phthalocyanine (Pc)complexes and polymers. The ligands and Pc complexes were characterized by ¹H nuclear magnetic resonance, fourier transform infrared, ultraviolet visible and mass spectrometric techniques. Additionally, thermal gravimetric, and elemental analyzer were used for characterization of the Pc complexes. The photophysical and photochemical behaviour of the Pc complexes were investigated in dimethyl sulfoxide. Additionally, the complexes were tested against epithelial breast cancer cells for photodynamic therapy (PDT)effect. The substituted ZnPc complexes afforded higher singlet oxygen quantum yields as compared to the AlPc analogue. All the complexes showed innocuous invitro dark cytotoxicity and moderate PDT effect.

Full text of DOI:10.1016/j.dyepig.2019.107592

Accepted Manuscript
Investigation of novel substituted zinc and aluminium phthalocyanines for
photodynamic therapy of epithelial breast cancer
Imadadulla Mohammed, David O. Oluwole, Manjunatha Nemakal, Lokesh Koodlur
Sannegowda, Tebello Nyokong
PII:
S0143-7208(19)30545-5
DOI:
https://doi.org/10.1016/j.dyepig.2019.107592
Article Number: 107592
Reference:
DYPI 107592
To appear in: Dyes and Pigments
Received Date: 11 March 2019
Revised Date: 30 April 2019
Accepted Date: 28 May 2019
Please cite this article as: Mohammed I, Oluwole DO, Nemakal M, Sannegowda LK, Nyokong T,
Investigation of novel substituted zinc and aluminium phthalocyanines for photodynamic therapy of
epithelial breast cancer, Dyes and Pigments (2019), doi: https://doi.org/10.1016/j.dyepig.2019.107592.
This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to
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ACCEPTED MANUSCRIPT
Synthesis of novel substituted zinc and aluminium phthalocyanines for
photodynamic therapy of epithelial breast cancer
a
b
a
Imadadulla Mohammed , David O. Oluwole , Manjunatha Nemakal , Lokesh Koodlur
a
b
Sannegowda* , Tebello Nyokong* .
a
Department of Chemistry, Vijayanagara Sri Krishnadevaraya University, Vinayakanagara,
Ballari-583105, Karnataka, INDIA
b
Center for Nanotechnology Innovation, Department of Chemistry, Rhodes University,
Grahamstown 6140, South Africa
Table of Content Graphic

ACCEPTED MANUSCRIPT
1
Investigation of novel substituted zinc and aluminium phthalocyanines for
photodynamic therapy of epithelial breast cancer
a
b
a
Imadadulla Mohammed , David O. Oluwole , Manjunatha Nemakal , Lokesh Koodlur
a
b
Sannegowda* , Tebello Nyokong* .
a
Department of Chemistry, Vijayanagara Sri Krishnadevaraya University, Vinayakanagara,
Ballari-583105, Karnataka, INDIA
b
Center for Nanotechnology Innovation, Department of Chemistry, Rhodes University,
Grahamstown 6140, South Africa

ACCEPTED MANUSCRIPT
2
Abstract
A series of phthalonitrile ligands were synthesized by nucleophilic substitution reaction using
the hydroxyl or sulfanyl group precursors and the nitro moiety of the nitrophthalonitrile to
yield corresponding oxy or sulfanyl bridged ligands. These ligands were subsequently
subjected to cyclocondensation reaction with diamagnetic metal ions like zinc and aluminium
to afford symmetrical substituted zinc and aluminium phthalocyanine (Pc) complexes and
1
polymers. The ligands and Pc complexes were characterized by H-nuclear magnetic
resonance, fourier transform infrared, ultraviolet visible and mass spectrometric techniques.
Additionally, thermal gravimetric, and elemental analyzer were used for characterization of
the Pc complexes. The photophysical and photochemical behaviour of the Pc complexes were
investigated in dimethyl sulfoxide. Additionally, the complexes were tested against epithelial
breast cancer cells for photodynamic therapy (PDT) effect. The substituted ZnPc complexes
afforded higher singlet oxygen quantum yields as compared to the AlPc analogue. All the
complexes showed innocuous invitro dark cytotoxicity and moderate PDT effect.
Key words: Zn phthalocyanine, Al phthalocyanine, polymer, amide bond, singlet oxygen
quantum yield, photodynamic therapy.
*
Corresponding authors: kslokesh@vskub.ac.in; T.nyokong@ru.ac.za

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3
1
. Introduction
Metallophthalocyanines (MPcs) are conjugated planar aromatic macrocycles having 18π–
electron ring system with four isoindole units linked via nitrogen atoms [1, 2]. MPcs offer
structural flexibility according to the utility and application by tailoring the Pc ring with different
functionality leading to improved solubility. Different MPcs can be synthesized by altering the
central metal cations at the core structure [3-6]. Phthalocyanine moiety has sufficient space to
accommodate metal cations in its central cavity.
Nowadays, MPcs and their derivatives have been frequently used in photodynamic therapy
(PDT) [7-10]. The singlet oxygen species are considered as the major cytotoxic species
responsible for photodynamic therapy (PDT) effect against tumorigenic cells. Typically, the
photosensitizer (PS) gets excited to its triplet state by light of appropriate wavelength (630-800
nm) and the excited PS interacts with its environment to produce highly reactive intermediates
(singlet oxygen, superoxide anion, and hydroxyl radical) that are cytotoxic leading to tumor
regression either directly by cell inactivation and/or indirectly by the destruction of the tumor
vascular microcirculation [9-11]. Most of the PS used in PDT [12-14] are tetrapyrrolic in nature;
chlorin, porphyrin, phthalocyanine and bacteriochlorin. Among these, MPcs are known for their
minimal dark cytotoxicity and efficacious PDT activity [15]. Especially Pc derivatives with
diamagnetic metal ions like zinc and aluminium have been explored for PDT because of their
ability to generate high singlet oxygen quantum yield which are essential for PS application in
PDT [16,-18]. Hence, there are lot of reports and efforts to enhance the efficiency of the singlet
oxygen production by tuning and modifying the functional groups at the periphery of the Pc
macrocycle [19]. Herein, we report for the first time the synthesis of MPc complexes with
different peripheral functionalities, namely zinc tetra 3-[(4-nitrophenyl)amino]-1-phenylpropan-
1
-one-3-(phenoxy) phthalocyanine [Pc 1], tetra 3-amino-propan-1-one-1,3-bis(phenoxy) zinc
phthalocyanine polymer [Pc 2], zinc tetra 4-(4-(p-tolyimino)methyl)phenoxy phthalocyanine

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4
[
Pc-3], tetra (E)-(naphthalen-1-yl)({4-[(E)-2-(naphthalen-1-yl)diazen-1-yl]phenyl})diazene-
bis(oxy) zinc phthalocyanine polymer [Pc 4], tetra 3H-spiro[2-benzofuran-3,9'-xanthene]-3-one-
',6'-bis(oxy) zinc phthalocyanine polymer [Pc 5], zinc tetra 2-methylpropanoic acid-2-oxy
3
phthalocyanine [Pc 6] and aluminium-chloro tetra pyridin-1-ium-1-olate-2-sulfanyl
phthalocyanine [Pc 7]. The efficiency of the MPcs with different functionalities were
systematically investigated for their singlet oxygen production and PDT effect.
2
. Materials and Methods:
4
-Nitrophthalonitrile, ammonium molybdate, urea, ZnCl
hydroxybenzaldehyde, 4-hydroxyacetophenone, citric acid, 2-naphthol, 2-mercaptopyridine-N-
oxide, α-hydroxyisobutyric acid, 4-nitroaniline, -toludine, N,N-dimethylformamide, n-
2 3
, AlCl , P-phenylenediamine, p-
P
pentanol, methanol, ethanol, spectroscopy grade dimethyl sulfoxide (DMSO), 1,3–
diphenylisobenzofuran (DPBF) and unsubstituted zinc phthalocyanine were acquired from
®
Sigma Aldrich . Cultures of human breast adenocarcinoma cells (MCF–7) were obtained from
®
Cellonex . Dulbecco's modified Eagle's medium (DMEM) with phenol red, phenol red free
DMEM, trypan blue, Dulbecco's phosphate–buffered saline (DPBS) and trypsin were purchased
®
from Sigma Aldrich . 100 µg/mL–Penicillin–100 unit/mL–streptomycin–amphotericin B
®
mixture, heat-inactivated fetal bovine serum (FBS) were acquired from Biowest . Cell
®
®
2
proliferation neutral red reagent WST–1 (Roche ), 96 well cell culture plates (Nest ), 25 cm
2
®
and 75 cm vented flasks were obtained from Porvair .
2
.1 Equipment:
Vario EL III CHNS elemental analyzer was used for the determination of elemental
compositions (C, H, N, S) of the synthesized complexes. Absorption spectra were recorded on a
Perkin Elmer Lambda 950 spectrophotometer for the Pc complexes. FTIR spectra were recorded
−
1
as KBr pellet in 4000 to 500 cm using Perkin Elmer Spectrum-Two spectrophotometer.
Thermogravimetric analysis was carried out on a STA6000 machine in the temperature range 30
°
1
to 700 C in oxygen atmosphere. The H NMR spectra were recorded on a Bruker AMX-400
using either deuterated methanol (MeOD-d4) or DMSO-d6 or CDCl solvent and mass spectra
3

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5
were recorded on Bruker ESQUIRE 3000 Mass Spectrometer using MS (ESI) technique for the
ligands and phthalocyanine molecules.
Illuminations for singlet oxygen quantum yields were performed using a general electric quartz
®
lamp (300 W). A 600 nm glass cut off filter (Schott ) and water filter were employed to filter off
ultra-violet and infrared radiation, respectively. An interference filter, 670 nm with a bandwidth
of 40 nm, was placed in the light path between the sample holder and 600 nm glass cut off filter.
®
Light intensities were measured with a POWER MAX 5100 (Molelectron detector
1
5
-2 -1
incorporated) power meter and were found to be 4.3 × 10 photons cm s . For the singlet
oxygen generation studies, samples and standards were mixed with the singlet oxygen quencher (
̴
-5
-3
3
x 10 mol dm DPBF was used for all mixtures, to avoid chain reactions) in DMSO at a
mixing ratio of 1:1 and the depletion of the quencher in the presence of samples or standard was
spectroscopically monitored at ~417 nm using a predetermined time intervals. The rate of the
photodegradation of the quencher in the presence of samples or standard was used in the
determination of the singlet oxygen quantum yields. Fluorescence excitation and emission
®
spectra were measured on a Varian Eclipse spectrofluorometer using a 360–1100 nm filter,
absorbance at the excitation wavelength was adjusted to ~0.05. Fluorescence lifetimes were
measured using a time correlated single photon counting setup (TCSPC) (FluoTime 300,
®
®
Picoquant GmbH) with a diode laser (LDH–P–670, Picoquant GmbH, 20 MHz repetition rate,
4 ps pulse width) [20].
4
®
Illumination source for PDT studies was obtained from Modulight Medical Laser system (ML)
710–680 channel Turnkey laser system coupled with a 2 × 3 W channel at 680 nm, cylindrical
7
output channels, aiming beam, integrated calibration module, foot/hand switch pedal,
subminiature version A connectors and safety interlocks. Illumination kit for in vitro PDT studies
has capacity to hold 127.76 × 85.48 mm 96 well cell culture plate [21].
2
2
®
The MCF–7 cells were cultured in 25 cm and 75 cm vented flasks (Porvair ) in a humidified
®
atmosphere incubator with ∼5% CO
2
and physiological temperature of 37 °C (HealForce ). The
®
cells were viewed under phase contrast using a Zeiss AxioVert. A1 Fluorescence LED (FL–
LED) inverted microscope and the cell viability was measured using cell proliferation neutral red
®
®
reagent WST–1 (Roche ) with a Synergy 2 multi–mode microplate reader (BioTek ).
2
.2. Synthesis of substituted phthalonitrile ligands:
2
.2.1. Synthesis of 3-(4-hydroxy-phenyl)-3-(4-nitro-phenylamino)-1-phenyl-propan-1-one (i):

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The mixture of 4-nitroaniline (3 g, 21 mM), acetophenone (2.5 g, 21 mM), p-
hydroxybenzaldehyde (2.6 g, 21 mM) and citric acid (0.8 g, 20 mM) were charged into a flask
containing 10 mL ethanol [22, 23]. The reaction mixture was stirred at room temperature for 24
h. The reaction was monitored by TLC. After the reaction completion, the crude product was
cooled to room temperature and poured into beaker containing ice water, the precipitate obtained
was filtered and washed with excess of deionized water until it was free from acid. The crude
product was purified by recrystallization from aq. ethanol. The resultant product was dried over
°
P O . Yield. 85%. Melting point. 122 C.
2
5
-1
FTIR (cm ): 1550 (C=C stretching), 1700 (-C=O stretching), 2890 (C-H stretching) and 3200-
3
550 (-OH stretching).
1
H-NMR(300 MHz, MeOD ): δ 8.04-7.63 (m, 4 Ar-H), δ 7.58-7.30 (m, 4 Ar-H), δ 7.14 (d, J =
8
.8 Hz, 1H), 6.75 (d, J = 8.8 Hz, 1H), δ 6.68-6.60 (m, 1H) δ 6.41 (m, 2H) 5.01 (dd, J= 5, 8.4 Hz,
H), 3.54 (dd, J= 8.4, 17.0 Hz, 1H), δ 3.38-3.25 (m, 1H), δ 2.26-2.60 (m, 1H) .
1
Anal. For: C H N O : Calc. C, 69.60; H, 5.01; N, 7.73; O, 17.66. Found: C, 70.00; H, 4.80; N,
2
1
18
2
4
7
.40.
MS (ESI): calc: 362.3 for C H N O ; m/z: 380 [M +H+NH ] , 317 [M+H-NO ] .
+
+
2
1
18
2
4
3
2
2
.2.2. Synthesis
of
4-{4-[1-(nitro-phenylamino)-3-oxo-3-phenyl-propyl]-phenoxy}-
phthalonitrile (ii):
Compound i (5.0 g, 0.0139 M) and 4-nitrophthalonitrile (2.4 g, 0.0139 M) were dissolved in 20
ml of DMF and K CO (1.93 g, 0.039 M) was added to the mixture in three portions within 30
2
3
min. with continuous stirring at room temperature. The reaction mixture was maintained at room
temperature and stirred for 72 h under inert atmosphere. Completion of the reaction was
confirmed by TLC and the product was poured into ice water with constant stirring to yield
brownish precipitate. The precipitate was washed with excess of deionized water and dried. The
crude product was recrystallized from ethanol and dried over P O . Yield: 80%. Melting point.
2
5
°
1
04 C.
-1
FTIR (cm ): 1560 (C=C stretching), 1692 (C=O stretching), 2915(C-H stretching), 2224 (-CN
stretching), and 3365 (-NH stretching).

ACCEPTED MANUSCRIPT
7
1
3
HNMR: (400 MHz, CDCl ): δ 1.24 (s, 2H), 3.48 (d, J = 8.0 Hz, 1H), 5.03 (s, 1H), 6.48 (d, J=8.0
Hz, 1H), 6.62 (d, J= 7.5 Hz, 1H), 6.79 (d, J=8.0 Hz, H), 7.23 (d, J= 7.5 Hz, 2H), 7.41 (d, J= 1.2
Hz, 2H), 7.55 (d, J= 8.0 Hz, 2H), 7.74 (d, J=8.0 Hz, 2H), 8.00 (dd, J= 1.6, 8.4 Hz, 1H), 8.58 (d,
J= 8.0Hz, 2H), 8.65 ppm (d, J= 8.0 Hz, 1H).
Anal. For C H N O : Calc. C, 71.30; H, 4.13; N, 11.47; O, 13.10. Found: C, 70.90; H, 4.54; N,
2
9
20
4
4
1
1.57.
.2.3. Synthesis of 3-amino-1,3-bis(4hydroxyphenyl]propane-1-one (iii):
2
The mixture of 4-hydroxyacetophenone (3.0 g, 22 mM), vaniline (3.35 g, 22mM), ammonia
0.27 ml, 22 mM) and citric acid (0.8 g, 20mM) were transferred into a flask containing 10 ml of
(
ethanol and the reaction mixture was stirred at room temperature for 24 h. Upon reaction
completion, the mixture was poured into the beaker containing ice water and the obtained
precipitate was filtered and washed with excess of deionised water until it is free from acid. The
product was recrystallized from aq. ethanol. The resultant product was dried over P O . Yield.
2
5
°
8
5%. Melting point. 72 C.
-1
FTIR (cm ): 1580 (C=C stretching), 1680 (C=O stretching), 2900 (C-H stretching) and 3210-
3
500 (-OH stretching).
1
H-NMR (400 MHz, DMSO-d6): δ 2.10 (d, J=7.3 Hz, 2H), 3.50 (s, 1H), 4.80 (d, J=7.3 Hz, 2H),
.08 (br s, 2H), 6.40-7.00 (m, 4H), 7.02 (d, J=7.0 Hz, 2H), 8.15 (d, J=7.0 Hz, 2H).
Anal. For: C15 : C, 70.02; H, 5.88; N, 5.44; O, 18.66. Found: C, 69.82; H, 5.68; N, 5.84.
5
H15NO
3
2
.2.4. Synthesis
of
4-(4-{1-amino-3-[4-(3,4-dicyanophenoxy)phenyl]-3-
oxopropyl}phenoxy)benzene-1,2-dicarbonitrile (iv):
Compound iv was synthesised using the procedure employed for compound ii but compound iii
was used instead of i. The precipitate was washed with excess of deionized water and the crude
°
product was recrystallized from ethanol and dried over P O . Yield: 80%. Melting point. 117 C
2
5
-1
FTIR (cm ): 1553(C=C stretching), 1650 (C=O stretching), 2930 (C-H stretching), 2232 (-CN
stretching), and 3330 (-NH stretching).
2
1
H-NMR (400 MHz, DMSO-d6): δ 1.24 (d, J=7.3 Hz, 1H), 2.59 (d, J=7.3 Hz, 2H), 3.83 (s, 2H),
δ 7.29-7.35 (m, 3H), 7.36-7.44 (m, 2H), 7.46-7.53 (m, 1H), 7.65-7.71 (m, 2H), 7.80 (d, J = 2.4

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8
Hz, 1H), 7.94 (d, J = 2.4 Hz, 1H), 8.07 (d, J = 8.8 Hz, 1H), 8.10-8.14 (m, 4H), 8.17 (d, J = 8.0
Hz, 2H).
Anal. For C32
3.32.
H
21
N
5
O
4
: Calc. C, 71.24; H, 3.92; N, 12.98; O, 11.86. Found: C, 70.86; H, 4.15; N,
1
+
MS (ESI): calc: 540.44 for C H N O ; m/z: 541 [M + H] .
3
2
21
5
4
2
.2.5. Synthesis of 4-(p-tolyimino) methyl) phenol (v):
A mixture of p-hydroxy benzaldehyde (2.0 g, 0.016 M) and p- toluidine (1.75 g, 0.016 M) were
dissolved in 10 mL of methanol and 0.2 mL of triethylamine was added to the mixture. The
reaction mixture was refluxed with constant stirring for 4 h to form light yellow colour product.
The crude product was recrystallized to form a pure (E)-4-(p-tolyimino) methyl) phenol. Yield:
°
8
0%. Melting point. 149 C.
-1
FTIR (cm ): 1545 (C=C stretching), 1600 (C=N stretching), 2890 (C-H stretching) and 3010-
3
250 (-OH).
1
H-NMR (400 MHz, DMSO-d6): δ 2.10 (s, 3H), 4.90 (br s, 1H), 7.05 (d, J= 7.2 Hz, 2H), 7.21-
7
.23 (m, 4H), 7.48 (d, J= 7.2 Hz, 2H), 8.64 (s, 1H).
Anal. for: C H NO: Calc. C, 79.59; H, 6.20; N, 6.63; O, 7.57. Found: C, 79.35; H, 6.32; N,
1
4
13
6
.75.
.2.6. Synthesis of 4-(4-(p-tolyimino)methyl)phenoxy phthalonitrile (vi):
2
Compound vi was synthesized using the same protocol applied for compound ii but compound v
was used instead of i. After the completion of the reaction, the product was extracted with
ethylacetate and washed with water. The organic layer was separated and dried over anhydrous
°
Na SO . Yield: 70%. Melting point. 132 C.
2
4
-1
FTIR (cm ): 1610 and 1475 (C=C stretching), 3030 (C-H stretching) and 2230 (-CN stretching).
1
H-NMR (400 MHz, DMSO-d6): δ 2.30 (s, 3H), 7.44 (dd, J= 1.6, 8.4 Hz, 1H), 7.91 (d, J= 8.0
Hz, 2H), 7.94 (d, J= 1.2 Hz, 1H), 8.16 (d, J= 7.0 Hz, 1H), 8.50 (s, 1H).
Anal. For C H N O: Calc. C, 78.3; H, 4.48; N, 12.46; O, 4.74. Found: C, 78.65; H, 3.06; N,
2
2
15
3
1
2.80.

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9
2
.2.7. Synthesis of 1-[(E)-2-{4-[(E)-2-(2-hydroxynaphthalen-1-yl)diazen-1-yl]phenyl}diazen-1-
yl]naphthalen-2-ol (vii):
-[(E)-2-{4-[(E)-2-(2-hydroxynaphthalen-1-yl)diazen-1-yl]phenyl}diazen-1-yl]naphthalen-2-ol
vii) was synthesized by diazotization of p-phenylenediamine (PPDA) and then coupling with 2-
1
(
naphthol. The procedure is as follows: sodium nitrite (3.34 g in 25 mL cold water) was slowly
°
added to PPDA (2.5 g in 10 mL dil. HCl) and the reaction was maintained at 0-5 C. After the
complete addition of sodium nitrite, the solution was kept for 20 min with constant stirring and
maintained at same temperature to yield diazonium salt solution [24]. Then the diazonium salt
solution was mixed with cold solution of 2-naphthol (7.2 g in 30 mL of 10% aq. NaOH solution)
to yield dark brown product (vii) which was filtered to obtain a solid product and it was washed
with excess of cold double distilled water to remove the unreacted 2-naphthol. Yield: 90%.
°
Melting point: 118 C
-1
FT-IR (cm ): 1600 (C=C aromatic stretching), 2980 (C-H stretching), 3200-3500 (-OH
stretching),
1
H NMR: (400 MHz, DMSO-d ): δ 7.09 (q, J = 2.4 Hz, 2H), 7.11-7.23 (m, 1H), 7.26 (t, J = 6.0
6
Hz, 1H), 7.76 (d, J = 3.2 Hz, 1H), 7.80 (d, J = 8.8 Hz, 2H), 9.75 (br.s, 1H).
Anal. for C H N O : Calc. C, 74.63; H, 4.34; N, 13.39; O, 7.65. Found: C, 74.20; H, 4.20; N,
2
6
18
4
2
1
3.57.
MS (ESI): Calc; 418.44 g/mol for C H N O , m/z: 420 (M+1), 278.2 [fragment ion (M+2-
2
6
18
4
2
(molecular weight of 2-naphthol))].
2
.2.8. Synthesis of ligand 4-({1-[(E)-2-{4-[(E)-2-[2-(3,4-dicyanophenoxy)naphthalen-1-
yl]diazen-1-yl]phenyl}diazen-1-yl]naphthalen-2-yl}oxy)benzene-1,2-dicarbonitrile (viii):
Compound viii was prepared by using the same procedure employed for the synthesis of
compound ii but compound vii was used instead of compound i. The precipitate was washed with

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10
2 5
excess water and recrystallized from aq. ethanol and dried over P O . Yield. 80%. Melting point:
°
2
20 C.
-1
FT-IR (cm ): 1587(C=C stretching), 2928 (C-H stretching) and 2224 (-CN stretching)
1
6
H-NMR (400 MHz, DMSO-d ): δ 7.36-7.39 (m, 1H), 7.48 (q, J = 2.4 Hz, 1H), 7.52-7.57 (m,
2
2
H), 7.72 (d, J = 2.4 Hz, 1H), 7.90 (t, J = 2.8 Hz, 2H), 7.93-7.98 (m, 1H), 8.12 (d, J = 8.8 Hz,
H).
Anal. for C H N O : Calc. C, 75.22; H, 3.31; N, 16.31; O, 4.77. Found: C, 75.42; H, 3.18; N,
4
2
22
8
2
1
6.65.
.2.9. Preparation of 4-[3'-(3,4-dicyanophenoxy)-3H-spiro[2-benzofuran-3,9'-xanthene]-3-
2
oneoxy]benzene-1,2-dicarbonitrile (ix):
Compound ix was prepared by using the same procedure described for compound ii but
fluorescein was used instead of compound i. The precipitate was washed with excess of water
2 5
and the crude product was recrystallized from aq. ethanol and dried over P O . Yield: 80%.
°
Melting Point: 122 C
-1
FTIR (cm ): 1610 (C=C), 3010 (C-H), 3300-3500 (-OH) and 2226 (-CN).
1
HNMR (400 MHz, DMSO-d6): δ 7.24 (d, J = 12.0 Hz, 4H), 7.45 (q, J = 4.0 Hz, 6H), 7.87-7.89
(m, 1H), 7.91-7.99 (m, 6H), 8.11 (d, J = 12.0 Hz, 2H).
Anal. for C H N O : Calc. C, 76.28; H, 3.11; N, 9.62; O, 10.9. Found: C, 78.92; H, 4.12; N,
3
7
18
4
4
1
0.50.
+
MS (ESI): Calc. 583 m/z for C H N O : m/z: 585 (M+2H) .
3
7
18
4
4
2
.2.10. Synthesis of 2-(3,4-dicyanophenoxy)-2-methylpropanoic acid (x):
Compound x was prepared as explained for compound ii by using α-hydroxyisobutyric acid
instead of compound i. The obtained crude product was purified by 230x400 silica gel
3
chromatography using 9:1 CHCl /MeOH as eluent. Yield: 70%.
°
Melting point: 129 C.
-1
FTIR (cm ): 1750 (-C=O stretching), 1600 (C=C stretching), 2910 (C-H stretching) and 2218 (-
CN stretching).
1
H-NMR (400 MHz, DMSO-d6): δ 1.23 (s, 6H), 7.27 (dd, J=1.8, 9.2 Hz, 1H), 7.41 (d, J= 8.0 Hz,
1
H), 7.49 (d, J= 1.0 Hz, 1H), 11.2 (br s, 1H).

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11
10 2 3
Anal. for C12H N O : Calc. C, 62.30; H, 4.38; N, 12.17; O, 20.85. Found: C, 62.47; H, 4.69; N,
1
2.6.
.2.11. Synthesis of 2-[(3,4-dicyanophenyl)sulfanyl]pyridin-1-ium-1-olate (xi):
2
Compound xi was prepared as explained for compound ii but 2-mercaptopyridine-N-oxide was
employed instead of compound i. The reaction product was extracted with ethylacetate and
washed with water. The organic layer was separated and dried over P O . Yield: 70%. Melting
2
5
°
point. 176 C.
-1
FTIR (cm ): 1590 (C=C), 2950 (C-H), 2231 (-CN).
1
H-NMR (400 MHz, DMSO-d6): δ 6.88 (d, J= 10.0 Hz, 1H), 7.20 (dd, J= 1.4, 8.6 Hz, 1H), 7.44
(d, J= 1.2 Hz, 1H), 7.80 (d, J=7.0 Hz, 1H), 8.10 (dd, J= 1.2, 9.8 Hz, 1H), 8.40 (d, J=7.0 Hz, 1H).
Anal. for C H N OS: Calc. C, 61.4; H, 3.17; N, 16.5; O, 6.29; S, 12.6. Found: C, 61.72; H, 34.4;
1
3
8
3
N, 16.55; S, 12.43.
2
.3. Synthesis of substituted phthalocyanines:
2
.3.1. Synthesis of Zinc tetra 3-[(4-nitrophenyl)amino]-1-phenylpropan-1-one-3-phenoxy
phthalocyanine [Pc 1]:
2
Compound ii (1.5 g, 0.003 M) and ZnCl (0.104 g (0.0008 M) were transferred into a flask
containing 10 ml of 1-pentanol and 0.1mL of DBU [25]. The reaction mixture was refluxed at
°
~
140 C with constant stirring for 24 h. Upon reaction completion, the mixture was allowed to
attain the room temperature and methanol was added to the crude product to yield dark green
product. The precipitate was then filtered and successively washed with methanol and acetone.
2 5
The product is then dried over P O . Yield: 75%.
-1
FTIR (cm ): 760, 872, 980, 1090, 1111 (phthalocyanine skeletal vibrations) and 3360 (-NH
stretching).
UV-Visible, nm (log ε):356 (5.24), 700(5.12).
Anal. For C H N O Zn: Calc. C, 68.99; H, 3.99; N, 11.10; O, 12.68; Zn,3.24. Found: C,
1
16 80 16 16
7
0.18; H, 3.85; N, 11.40; Zn, 3.04.

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12
2
.3.2. Synthesis of tetra 3-amino-propan-1-one-1,3-bis(phenoxy) zinc phthalocyanine polymer
[Pc 2]:
Tetra 3-amino-propan-1-one-1,3-bis(phenoxy) zinc phthalocyanine polymer [Pc 2] was prepared
using the procedure for Pc-1 but using compound iv was employed instead of ii. Compounds
with two phthalonitrile units will produce the symmetrical metallophthalocyanine polymers as in
the literature [26-28]. Yield: 75%.
-1
FTIR (cm ): 765, 880, 990, 1085, 1115 (phthalocyanine skeletal vibrations) and 2220 (weak, -
CN stretching for peripheral Pcs)
UV-Visible, nm (log ε): 266 (5.23), 678 (5.10)
Anal. For: C H N O Zn: Calc. C, 69.10; H, 4.29; N, 10.51; O, 12.01; Zn, 4.09. Found: C,
9
2
68 12 12
6
9.40; H, 4.39; N, 10.31; Zn, 3.89.
2
.3.3. Synthesis of Zinc tetra 4-(4-(p-tolyimino)methyl)phenoxy phthalocyanine [Pc-3]:
Pc-3 was obtained using the same synthetic route employed for Pc–1 but compound vi was used
instead of ii. Yield: 75%.
-1
FTIR (cm ): 748, 878, 969, 1071 and 1109 (phthalocyanine skeletal vibrations), 1610 (C=N
stretching) 2920 (-CH stretching).
UV-Visible, nm (log ε): 355(5.16), 681(4.95).
60 12 4
Anal. For C88H N O Zn: Calc. C, 74.78; H, 4.27; N, 11.8; O, 4.52; Zn, 4.62. Found: C, 76.80;
H, 5.34; N, 12.4; Zn, 4.8.
60 12 4
MS (ESI): Calc; 1414.88 g/mol for C88H N O Zn, m/z: 1415.93 (M-1).
2
1
.3.4. Preparation of substituted tetra (E)-(naphthalen-1-yl)({4-[(E)-2-(naphthalen-1-yl)diazen-
-yl]phenyl})diazene-bis(oxy) zinc phthalocyanine polymer [Pc 4]:
Tetra (E)-(naphthalen-1-yl)({4-[(E)-2-(naphthalen-1-yl)diazen-1-yl]phenyl})diazene-
bis(oxy) zinc phthalocyanine polymer [Pc 4] was synthesised as described Pc-1 but compound
viii was used instead of ii. Yield: 70%.

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13
-1
FT-IR (cm ): 759, 893, 964, 1104, 1133 (phthalocyanine skeletal vibrations) and 2215 (-CN
stretching)
UV-Visible, nm (log ε): 356(5.19), 683(4.77).
Anal. For: C H N O Zn: Calc. C, 72.80; H, 3.59; N, 14.98; O, 5.70; Zn, 2.92. Found: C,
1
36 80 24
8
7
3.15; H, 3.74; N, 14.80; Zn, 2.72.
2
.3.5. Preparation of tetra 3H-spiro[2-benzofuran-3,9'-xanthene]-3-one-3',6'-bis(oxy) zinc
phthalocyanine polymer [Pc 5]:
Pc-5 was synthesized as described for Pc-1 but compound ix was used instead of ii. Yield: 70%.
-1
FTIR (cm ): 770, 898, 960, 1110, 1131 (phthalocyanine skeletal vibrations) and 2226 (-CN
stretching).
UV-Visible, nm (log ε): 288 (4.44), 692 (4.95).
Anal. For: C H N O Zn: Calc. C, 73.67; H, 3.41; N, 5.92; O, 13.54; Zn, 3.46. Found: C,
1
16 64
8
16
7
3.55; H, 3.53; N, 5.80; Zn, 3.58.
2
.3.6. Synthesis of Zinc tetra 2-methylpropanoic acid-2-oxy phthalocyanine [Pc 6]:
Pc-6 was synthesized as described for Pc-1 but compound x was used instead of compound ii.
Yield: 65%.
-1
FTIR (cm ): 745, 885, 958, 1094, 1129 (phthalocyanine skeletal vibrations) and 1765(C=O
-1
stretching of -COOH) and 3110-3450 cm (-OH stretching of -COOH).
UV-Visible, nm (log ε): 290(5.00), 693(4.86).
Anal. for C H N O Zn: Calc. C, 58.40; H, 4.0; N, 11.35; O, 19.4; Zn, 6.62. Found: C, 58.71;
4
8
40
8
12
H, 3.70; N, 12.56; Zn, 7.05.
MS (ESI): Calc; 986.37 g/mol for C H N O Zn, m/z: 985.5(M+1).
4
8
40
8
12
2
.3.7. Synthesis of Aluminium chloro tetra pyridin-1-ium-1-olate-2-sulfanyl phthalocyanine [Pc
7
]:
Pc-7 was synthesized as described for Pc-1 but compound xi and AlCl were used instead of
3
2
compound ii and ZnCl . Yield: 65%.

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14
-1
FTIR (cm ): 755, 892, 970, 1005, 1125 (phthalocyanine skeletal vibrations)
UV-Visible, nm (log ε): 331 (5.04), 687(5.06).
Anal. for C52
32 12 4 4
H N O S : Calc. C, 57.80; H, 2.96; N, 20.75; S, 11.88; Cl, 3.28; Al, 2.50. Found:
C, 58.22; H, 2.50; N, 20.8; S, 12.21
3
3
. Results and discussions
.1. Synthesis and characterization of the Pc complexes
The synthesis of the ligands and substituted zinc and aluminium phthalocyanine complexes
and polymers were carried out as shown in Table 1 and Scheme 1. The first step involves the
preparation of hydroxyl or sulfanyl precursors for all the Pcs. The second step involves the base
catalyzed nucleophilic aromatic nitro displacement of 4-nitrophthalonitrile with hydroxyl or
sulfanyl group containing precursors to yield phthalonitrile/bisphthalonitrile ligands [29,30]. The
phthalonitrile ligands undergo cyclotetramerization to form the symmetrical substituted
phthalocyanine complexes, whereas bisphthalonitrile forms polymeric phthalocyanines. In the
first step of polymerization, four molecules of corresponding bisphthalonitrile ligand reacts to
give substituted octacyanophthalocyanine. Then the peripheral cyano groups of phthalocyanine
units interact at high temperature to yield fused ring structure joined at the sides to form ladder
polymeric Pcs. This type of conjugated ladder polymer has advantages over single stranded
conjugated polymer in terms of chemical resistance, thermal stability, mechanical strength and
greater molecular and structural order [31].
The reactions were monitored by TLC and the synthesized ligands were purified and
characterized with, FT-IR, NMR, mass spectroscopy and melting point (NMR and mass spectral
data are provided in supplementary information). The phthalocyanine complexes and polymers
were successfully purified to afford blue to green product with solubility in DMSO, DMF and
concentrated H SO . The polymeric Pc’s are less soluble compared to the monomeric Pc’s.
2
4
3
.1.1. FTIR Characterization:
FT-IR spectra for all the synthesised ligands and phthalocyanine complexes were
-1
recorded in the region 400 to 4000 cm . The FTIR spectra of compounds i to xi are shown in Fig
-1
1
, all the compounds showed vibrational band at 1540-1620 cm which corresponnd to C=C

ACCEPTED MANUSCRIPT
15
-1
stretch and 2870-3020 cm corresponding to C-H stretch. Compounds i, iii, v and vii showed
-1
broad peaks at 3100-3450 cm which is assigned to -OH.
-1
In addition, compound i-iv showed vibration band at 1650-1700 cm corresponding to
-1
C=O, i and ii showed peak at 3340-3350 cm which corresponds to –NH (secondary amine) and
-1
compounds iii and iv showed bands at 3300-3330 cm corresponding to -NH . A peak was
2
-1
observed for compound v at 1600 cm which can be assigned to C=N. The IR spectra for
-1
compound vi exhibited vibrational bands at 1610 and 1475 cm corresponding to C-C ring
-1
skeletal stretch vibrations and the 720 cm band to -CH out of plane bending vibrations.
-1
Compound ix showed peak at 1750 cm accountable to the C=O peak and compound x at 1750
-1
-1
cm for C=O of the carboxylic acid with broad peak at 3110-3450 cm for –OH of the
-1
carboxylic acid. Compound xi showed peak at 1270 cm may be due to the NO of pyridine-N-
oxide [32].
-1
Ligands ii, iv, vi, viii, ix, x and xi showed sharp peak at 2200-2290 cm corresponding to
–
CN group. The appearance of -CN group and disappearance of –OH or –SH peak confirms the
formation of the conjugated phthalonitriles . In addition to TLC, IR was useful in monitoring the
completion of the formation of cyano ligands from their counter hydroxy ligands by monitoring
-1
the appearance of cyano group peak at ~2250 cm with the complete disappearance of the
-1
-1
hydroxy or sulfanyl group at ~3300 cm or 2550 cm . All the phthalocyanine complexes
-1
showed peaks at 745-760, 875-895, 965-990, 1004-1110 and 1111-1135 cm which can be
assigned to the phthalocyanine skeletal vibrations [33]. Additionally, the IR spectra of Pcs
showed disappearance of –CN peak in Pc-1, Pc-3, Pc-6 and Pc-7 indicating the consumption of
cyano groups to form phthalocyanine macrocycle. Whereas in polymeric Pc’s (Pc-2, Pc-4 and
-1
Pc-5), though the intensity of the cyano peak at 2200-2290 cm was decreased but did not
disappear completely due to the terminal –CN peak of polymeric Pc’s. The peaks of the polymer
were slightly lesser in intensity and broader in nature as compared to the monomeric Pcs.
3
.1.2. UV-Vis spectra:
The UV Visible spectra for all the Pcs were recorded by dissolving 1mg dry
phthalocyanine in 5 mL DMSO. The UV-Vis spectra were recorded in the range of 250 to 900
-1
nm at a scan rate of 240 nm min .

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16
Figure ES-12 shows the absorption spectra of the Pc’s. All the synthesized Pc’s showed
absorption peaks at 250-280, 335-360, 414-450, 620-650 (shoulder peak) and 685-705 nm, Table
1
. The peak observed in the UV region at 335-360 nm correspond to B-band and the peak
observed at 620-650 nm can be accounted for the oligomer and dimeric forms of the
phthalocyanine macrocycle and the intense Q-band was observed at 685-705 nm [34]. The
absorption spectra of Pc-4, Pc-5, Pc-6 and Pc-7 showed split Q-band. The splitting of the Q-
band in Pc-6 and Pc-7 suggests aggregation of the macrocycle whereas the splitting in case of
Pc-4 and Pc-5 may be due to the polymeric nature of the macrocycle. The deep intense
blue/green colour of the phthalocyanines is due to the peak at 670-700 nm which arises because
of the π-π* transitions of the conjugated macrocycle [35].
3
.1.3. TGA studies:
TGA studies provide information about the thermal stability and decomposition behaviour of the
synthesized phthalocyanine compounds. The thermogram for the thermal degradation of the
phthalocyanines is shown in Fig. 2.
°
It was observed from the thermograms that the monomeric Pcs are stable up to 300 C
°
and then starts to degrade in air atmosphere in the region of 350–490 C which leads to the rapid
loss of weight corresponding to the degradation of core structure of phthalocyanine molecule.
°
Whereas, the polymeric Pcs were found to be stable up to 500 C and then the polymeric
°
molecule starts to degrade in air atmosphere in the region of 520–650 C. Pc-4 shows higher
thermal stability than other two polymeric Pcs due to the extended conjugation and
°
delocalization of π-electrons. The stable TGA curve after 520 C corresponds to the respective
o
metal oxide formed after complete degradation of the Pcs. The weight remaining after 490 C in
the TG curve is theoretically equivalent to metal oxide of the sample taken for TGA analysis.
3
.1.4. NMR and Mass spectra:
NMR and mass spectroscopy gives the information about the structural and molecular mass of
1
the compounds. In particular H-NMR gives the information about the chemical environment of
the protons present in the compound. The number of non-equivalent protons are equal to the
1
number of peaks in the NMR spectra. H-NMR spectra was recorded for all the synthesized
compounds (i-xi) in different deuterated solvents, such as for compound (i) in deuterated
4
methanol (methanol-d ), compound (ii) in CDCl3 and compounds (iii-xi) in deuterated DMSO

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17
(DMSO-d6). The synthesized compounds are having different types of protons like aromatic
protons, aliphatic protons, -OH, and amine groups. The aromatic protons show peaks around 6-8
δ, aliphatic protons (-CH at 0.9-1.5 δ, -CH at 1.2-1.8 δ and –CH at 1.5-2.1 δ) at 1-2.2 δ, -OH at
3
2
3
-10 δ and primary and secondary amines at 5-6.5 δ. All the precursor compounds exhibited
broad singlet peak for –OH and –SH group proton, whereas this peak disappeared in their
1
corresponding ligands confirming the formation of respective ligands. In addition, the H-NMR
peaks obtained for different ligands are well matching with the theoretical data for the proposed
structures.
Mass spectra gives the information about the molecular weight and the fragmentation of
+
-
the compounds. The synthesized ligands showed (M or M ) molecular ion peaks in mass spectra
which are well coinciding with the theoretical molecular weight of the compounds. Some ligands
showed fragment ion peaks which are responsible for the detachment of one part of the molecule
-1
from the ligand. The mass spectra of Pc-3 displayed peak at 1415.9 corresponding to M
+
1
molecular ion, Pc-6 showed peak at 985.5 corresponding to M molecular ion. The NMR and
mass spectra of the precursors, ligands and complexes suggests the successful formation of pure
desired compounds.
3
.2. Photophysicochemical Parameters
The fluorescence (Φ ) and singlet oxygen (Φ
∆
) quantum yields of the Pc complexes were
determined using comparative methods reported in the literatures [36-38]. Unsubstituted ZnPc
F
dissolved in DMSO was used as a standard (Φ =0.20 [37], and Φ
∆
= 0.67 [38] in DMSO using
F
DPBF as a quencher).
3
.2.1. Fluorescence (Φ ) quantum yields and lifetimes (τ )
F F
Figure 3 (using Pc-2 and Pc-6 as examples) shows the normalised absorption, emission and
excitation spectra and figure ES 11 (for all the Pc complexes) shows the ground state electronic
absorption, emission and excitation spectra of Pc complexes in DMSO. The absorption and
excitation spectra were similar and mirror images of the emission spectra for Pc–2. This applied
to all non-aggregated complexes. Pc–6 and other aggregated complexes showed a slightly
different behaviour with absorption spectra different from the excitation spectra due to the
presence of aggregates in the absorption spectra. Aggregates do not fluoresce. Aggregation in

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18
Pc’s is often judged by the broadening or split in the Q band with the high energy band being due
to the so called “H” aggregates [39].
The fluorescence quantum yields and lifetimes of the Pc complexes were measured in DMSO
with Pc–5 accounting for the highest Φ value (0.16) while Pc–4 and Pc–6 accounted for the
F
least Φ value (0.02), Table 2. Low Φ values are due to aggregation. A typical fluorescence
F
F
decay curve is shown in Figure 4 for Pc complexes (using Pc–5 as an example). A single
fluorescence lifetime was observed for all the Pc complexes. Additionally, the fluorescence
quantum yields of the Pc complexes afforded corresponding fluorescence lifetimes which is
expected since there is a direct correlation between the former and latter except for Pc–6 which
showed a decrease in its fluorescence lifetime despite the relatively high fluorescence quantum
yield, it afforded.
3
.2.2. Singlet Oxygen Quantum Yield
The singlet oxygen quantum yields of the Pc complexes were determined using photochemical
method involving DPBF as singlet oxygen quencher which was spectroscopically monitored
from 0 s to 120 s at ~417 nm. Concentrations of Pc complexes for the determination of singlet
-6
-6
oxygen quantum yield are as follows, Pc–1 (8.12 × 10 M), Pc–2 (7.95 × 10 M), Pc–3 (1.15 ×
-5
-5
-5
-5
-5
1
0 M), Pc–4 (2.02 × 10 M), Pc–5 (1.11 × 10 M), Pc–6 (1.35 × 10 M) and Pc–7 (1.06 × 10
M). The characteristic change in absorbance profile of DPBF in the presence of PS is shown in
Figure 5 (using Pc–2 as an example). A gradual decrease in the absorbance of DPBF was
observed suggesting the production of singlet oxygen. There was no change in the intensity of
the Q band for all the Pc complexes on exposure to illumination inferring the photostability of
the Pc complexes. Pc–1, Pc–6 and Pc–7 showed a relatively low singlet oxygen quantum yields
which might be attributed to plausible aggregation as earlier shown in their UV–vis spectra
(Table 2).
3
.3 Cell studies
Cell culture and treatment
The in vitro dark cytotoxicity and photodynamic therapy activities of the photosensitizers (PS)
were performed as described in the literature [25, 40]. Details of the protocol are provided in the
supporting information.

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19
The cell viability represents the fraction of active cells after PS treatment and incubation for 24 h
often expressed as a normalized percentage. Prior to PDT studies, the in vitro dark cytotoxicity
of the PS were carried out in the absence of illumination to ascertain their applicability for PDT
since dark cytotoxicity of PS is undesirable in PDT due to plausible lack of selectivity and
specificity towards tumorigenic cells as opposed to healthy cells.
3
.3.1. In vitro dark cytotoxicity
Figure 6A and 6B shows the histograms for the in vitro dark cytotoxicity and PDT activities of
PS against MCF–7 cells (using Pc–5 as an example for the gradient PS concentrations). All the
PS accounted for ≥81% viable cells within the concentration range of 5–80 µg/mL inferring
minimal in vitro dark cytotoxicity which is a good indication for PS meant for PDT application,
Figure 6A and 6B. The overall confluence of the cells in the presence of PS were relatively
unaltered as compared to the control cells, Figure not shown.
The Synergy 2 multi–mode microplate reader was used to assess the cellular uptake of the PS
[41]. From Figure 7 (using Pc-3 as an example), it can be observed that the cellular uptake of the
PS was concentration dependent due to the increase in the cellular uptake as the PS concentration
increases. There was steady cellular uptake of the PS as the concentration increases. The efficient
steady uptake of the PS by the cell is advantageous for PDT due to the availability of the
unchanged PS in the cells which will inevitably elicit pharmacological response on exposure to
light.
3
.3.2. In vitro photodynamic therapy evaluation
The in vitro PDT evaluation of the PS was carried out at the same concentrations and
experimental conditions as the in vitro dark cytotoxicity studies except for irradiation of the
-2
incubated cells having the PS at 680 nm and laser beam dosimetry of 85 J.cm , respectively.
Significant decrease in the cell viability of the cells having PS exposed to light was observed
in comparison to the ones without irradiation. PDT activity increased moderately with PS
concentration increases, Fig. 6A. Of importance is the high PDT activity observed for the Pcs 5–
7
with less than 50% viable cells as compared Pcs 1–4 which showed greater than 50% viable
cells across all the tested concentrations (5–80 µg/mL) using 80 µg/mL as an example for all the
complexes, Figure 6C. Pcs 6 and 7 showed the highest PDT effect with 57% cell death while Pc

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20
3
accounted for 33% cell death. It is important to note that the latter showed the highest singlet
oxygen quantum yield (Table 2) as compared to the former or other Pcs considered in this study
but afforded the least PDT activity, Figure 6C. It has been reported that the PDT activity of PS
is dependent on factors such as cell type, cellular uptake and localization [42]. However, the PS
with high singlet oxygen quantum yields could have suffered from low cellular uptake and
localization as compared to the others with low singlet oxygen quantum yields which showed
high PDT since they differ greatly on their ring substituents. The overall confluence of the
cellular micrographs in the presence of PS were significantly altered as compared to the control
cells evidenced by the photo–micrographs, Figure 8 (using Pcs 1 and 2 as examples). From the
photo–micrographs, it can be deduced that the confluence of the cells having the PS were low as
compared to the control cells which is a good indication for the PDT activity of the PS.
4
. Conclusion:
The present work demonstrates the successful synthesis of novel Zn and Al phthalocyanine
derivatives in pure state. All the Pcs showed significant PDT activity compared to the in vitro
dark cytotoxicity behavior, but Pc–1 to Pc–4 showed ≥50% viable cells within the tested
concentration range of 5–80 µg/mL. The remarkable PDT activity was shown by Pc–5 to Pc–7
with Pc–5 showing less than 50% viable cells at 80 µg/mL, while Pc–6 and Pc–7 achieved the
same efficacy at 40 µg/mL and 80 µg/mL. Though, Pc–6 and Pc–7 exhibited low singlet oxygen
quantum yields in DMSO plausibly due to aggregation, but showed higher PDT activity
∆
compared to the other PS (Pc–1 to Pc–5) with larger Φ values. This phenomenon could be
ascribed to better interaction of the PS (Pc–6 and Pc–7) in slightly amphiphilic media compared
to the lipophilic ones. On visualization of the bright field fluorescence micrographs of the
placebo cells and treated cells (cells having PS), significant alteration in the overall confluence
of the treated cells was observed compared to the control cells. The observed spaces in the
treated cells could be attributed to the cytocidal activity of the PS against the MCF–7 cells on
exposure to light.
Acknowledgements:
This work was supported by the Department of Science and Technology (DST)
Innovation and National Research Foundation (NRF), South Africa through DST/NRF South
African Research Chairs Initiative for Professor of Medicinal Chemistry and Nanotechnology

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(UID 62620), DST-SERB, grant No. SERB/ F/9388/ 2016-17, CSIR EMR-II grant, VGST K-
FIST support (GRD No.555), DST-FIST grant and Directorate of Minorities, Karnataka
Government (GOKDOM) is also acknowledged.

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Table I: Synthetic scheme for different ligands
Pcs
Pc-1
Synthesis of Ligand
R
NO
2
O
HO
NH
2
O
NH
CN
CN
NO2
M=Zn
X=O;
Z=H
+
2
O N
+
+
HO
O
HN
NO2
O
i
NO
2
O
NH
O
CN
CN
ii
O
O
NH2
HO
Pc-2
Polymer
M=Zn
X=O;
CN
CN
+
NH
3
+O2N
O
NH2
+
HO
OH
HO
O
iii
O
NH2
O
O
Z=O
CN
CN
CN
CN
iv
CN
CN
Pc-3
M=Zn
X=O;
Z=H
OH
O
CHO
OH
CN
CN
+
+ O
2
N
N
N
NH2
N
v
vi