New Cyanoarylporphyrazines with High Sensitivity of Photophysical Parameters to Viscosity as Promising Agents for Photodynamic Therapy

Article abstract of DOI:10.1134/S1070363220020140

Abstract: Photophysical properties and photodynamic activity for a series of cyanoarylporphyrazine pigments have been analyzed depending on the nature of the substituents and their position in the aromatic ring. Replacement of the fluorine atom in the par

Full text of DOI:10.1134/S1070363220020140

ISSN 1070-3632, Russian Journal of General Chemistry, 2020, Vol. 90, No. 2, pp. 249–256. © Pleiades Publishing, Ltd., 2020.
Russian Text © The Author(s), 2020, published in Zhurnal Obshchei Khimii, 2020, Vol. 90, No. 2, pp. 274–283.
New Cyanoarylporphyrazines with High Sensitivity
of Photophysical Parameters to Viscosity as Promising Agents
for Photodynamic Therapy
S. A. Lermontovaa, T. S. Lyubovaa, E. Yu. Ladilinaa, V. I. Plekhanovb, I. V. Balalaevac,
V. P. Boyarskiid, and L. G. Klapshinaa,b,*
a Razuvaev Institute of Organometallic Chemistry of the Russian Academy of Sciences, Nizhny Novgorod, 603950 Russia
b Institute of Applied Physics of the Russian Academy of Sciences, Nizhny Novgorod, 603950 Russia
c Lobachevsky State University of Nizhny Novgorod, Nizhny Novgorod, 603950 Russia
d St. Petersburg State University, St. Petersburg, 198504 Russia
*e-mail: klarisa@iomc.ras.ru
Received June 24, 2019; revised June 24, 2019; accepted June 27, 2019
Abstract—Photophysical properties and photodynamic activity for a series of cyanoarylporphyrazine pigments
have been analyzed depending on the nature of the substituents and their position in the aromatic ring. Replace-
ment of the fluorine atom in the para-position with the methoxy group leads to significant increase in the ratio
of the dark and photoinduced cytotoxicities, i.e. potential therapeutic index of porphyrazine as photosensitizer in
photodynamic cancer therapy.
Keywords: cyanoarylporphyrazines, photosensitizers, photoinduced and dark cytotoxicity, fluorescent
molecular rotors, optical probes of intracellular viscosity
DOI: 10.1134/S1070363220020140
Photodynamic cancer therapy is a popular method of
oncological diseases treatment exploiting the properties
of a photosensibilizer which is selectively accumulated
in the tumor and can be activated by visible or infrared
radiation with the appropriate wavelength [1–6]. Such
activation leads to conversion of the excited singlet
state of the photosensibilizer in the triplet one. Certain
pigments in the singlet state are prone to generate
active forms of oxygen: radicals and/or singlet oxygen
exhibiting short lifetime yet extremely high cytotoxicity
[7, 8]. Hence, three non-toxic components of the therapy
(photosensibilizer, oxygen, and light) can efficiently
kill cancer cells in the tumor and/or disrupt its blood-
vascular system. Most of the photosensibilizers known
to date belong to a class of fluorescent tetrapyrrole
pigments based on porphyrines and phthalocyanines
derivatives [9, 10]. The prospects of the application of
porphyrazines as the agents of photodynamic cancer
therapy have been recently demonstrated as well [11–
13]. Photophysical and cytotoxic properties of a novel
class of tetrapyrroles, cyanoarylporphyrazine pigments
exhibiting unique potential towards diverse biomedical
applications (photodynamic therapy, bimodal fluore-
scent/magnetic resonance diagnostics, and probing
of intracellular viscosity), have been recently studied
at Razuvaev Institute of Organometallic Chemistry,
Russian Academy of Sciences [14–20].
Similar to other tetrapyrrole photosensibilizers, these
compounds can act as both therapeutic and diagnostic
agents, since they are selectively accumulated in a tumor
which allows their imaging via fluorescence. The use of
a photosensibilizer in dark or irradiation in the absence
of a photosensibilizer does not bring the cytotoxic effect.
It should be noted that the use of oxygen active forms
in cancer photodynamic therapy is advantageous over
other treatments in oncology, since the tumor cells are
selectively affected, and, hence, normal cells are not
damaged in the absence of irradiation and at low inher-
ent (dark) cytotoxicity of the photosensibilizer. This fact
underscores the importance of the ratio between dark and
photoinduced cytotoxicity, an important indicator of the
photosensibilizer safety.
Herein we present the study of photophysical and cy-
totoxic properties of several novel cyanoarylporphyrazine
pigments as well as those synthesized by us earlier [14, 17,
249

250
LERMONTOVA et al.
Scheme 1.
R
CN
N
N
NC
R
R
NH
N
N
N
N
HN
CN
NC
R
ꢀ
1 7
R = Ph (1), 4-FC H (2), 4-MeOC H (3), 2-MeC H (4),
6
4
6
4
6 4
2-MeOC H (5), 3-MeO-4-EtOC H (6), C F (7).
6
4
6
3
6 5
Scheme 2.
R
H
CN
CN
R
CN
R
CN
CN
R
H
NCS
CH2(CN)2
KCN, HCl
O
NC
CN
NC
ꢀ
14 16
ꢀ
8 10
ꢀ
11 13
R
CN
R
N
CN
NC
R
R
NC
R
NH
N
C
O
O
CN
Yb
N
N
R
N
N
NC
R
TFA
N
HN
N
N
N
N
N
Yb
N
CN
N
N
R
CN
NC
NC
R
, ,
3 5 6
ꢀ
17 19
R = 4-MeOC H (3, 8, 11, 14, 17), 2-MeOC H (5, 9, 12, 15, 18), 4-MeOC H (6, 10, 13, 16, 19).
6
4
6
4
6 4
20] and demonstrate strong effect of nature and position
of the substituents in the phenyl ring on the magnitude
and ratio of the photoinduced and dark cytotoxicity.
in the phenyl ring did not significantly affect the position
of the Q-band. Hence, the introduced substituents did not
significantly influence the π-conjugation system of the
macrocycle, except for the pentafluorophenyl fragment
in compound 7 which led to considerable red shift of its
Q-band, the underlying reason remaining unclear.
Scheme 1 shows the structure of novel and earlier
described [14, 18, 20] cyanoarylporphyrazines 1–7 con-
sidered in this study. Novel porphyrazines 3, 5, and 6
were synthesized according to Scheme 2.
Acharacteristic feature of the considered series of the
porphyrazines was strong sensitivity of the photophysical
parameters to local viscosity. That was due to the possibil-
ity of the radiationless relaxation of the excited state of the
cyanoarylporphyrazine molecule (dark state) consisting in
the excitation energy dissipation via intramolecular rota-
tion of the mobile fragments of the dye upon excitation
of the porphyrazine with light and charge transfer from
the π-donor aryl groups to the electron-accepting CN-
Figure 1 displays the absorption and emission spectra
of the new porphyrazines 3, 5, and 6. Their absorption
spectra were typical of tetrapyrrole macrocycles.
The photophysical properties of the considered por-
phyrazines which are crucial for the estimation of a pho-
tosensibilizer potential for cancer photodynamic therapy
application are collected in Table 1. It is interesting to
notice that in most cases the presence of the substituents
RUSSIAN JOURNAL OF GENERAL CHEMISTRY Vol. 90 No. 2 2020

NEW CYANOARYLPORPHYRAZINES WITH HIGH SENSITIVITY
251
Fig. 1. Absorption (a) and emission (b) (λex = 580 nm) spectra of porphyrazines 3, 5, and 6 in water.
groups [18, 21–23]. The increase in the medium viscosity
(3)
hindered the intramolecular rotation and, hence, increased
the fluorescence quantum yield and its decay time. The
relation of those photophysical parameters for the “fluo-
rescent molecular rotors” to viscosity is expressed by
well-known Förster–Hoffman equations (1).
The new cyanoarylporphyrazines (3, 5, and 6) as well
as the earlier obtained compounds 1, 2, 4, and 7 [14, 20]
turned out to be typical fluorescent molecular rotors, as
confirmed by good agreement with the Förster–Hoffman
equations (Fig. 2).
log Φ = z + αlgη,
(1)
The dependences of the rate constants of radiation
and radiationless transitions on the medium viscosity
(Fig. 3) confirmed that indeed the new porphyrazine
pigments were fluorescent molecular rotors. The plotted
data showed that the rate constant of the radiationless
transition was sharply decreased with the increase in
the viscosity, whereas the rate constant of the radiation
transition was almost viscosity-independent.
with Φ being the quantum yield, z and α being constant
parameters, and η being the viscosity.
A similar equation is known for fluorescence lifetime
[24]:
log τ = log (z/kr) + αlog η,
(2)
with τ being the lifetime of the excited state, η being the
viscosity, z and α being constant parameters, and kr being
rate constant of the radiationless transition.
The above-described photophysical features of the
cyanoarylporphyrazine photosensibilizers opens the
possibility of their use as local viscosity probes (includ-
ing the intracellular viscosity). Viscosity is an important
parameter determining the functional status and viability
Rate constants of the radiation and radiationless tran-
sitions for the porphyrazines were calculated from the
values of the fluorescence parameters (quantum yield Φf
and lifetime τav) using equation (3) [23].
Table 1. Absorption (Q-band) and red fluorescence maximums, quantum yield of fluorescence, and molar absorptivity (log ε)
of porphyrazines 1–7 in watera
Comp. no.
Q-band, nm
579
log ε
4.23
4.42
4.63
4.53
4.62
4.68
4.52
λem, nm
650
Φ, %b
0.3
αc
References
[10]
1
0.5
0.6
0.5
0.3
0.5
0.6
0.4
2
579
650
0.3
[4, 7, 10]
3
576
645
0.7
4
585
650
0.4
[4, 10]
[10]
5
586
655
0.6
6
7
576
650
0.5
606
695
0.1
a c = 5 μmol/L.
b Quantum yield was calculated relative to the Cresol Violet, λex = 580 nm.
c Value of parameter α in the Förster–Hoffman equations.
RUSSIAN JOURNAL OF GENERAL CHEMISTRY Vol. 90 No. 2 2020

252
LERMONTOVA et al.
toxicity of the porphyrazine. Therefore, compound 3 is
potentially more efficient photosensibilizer in the cancer
photodynamic therapy, since the decrease in the inherent
toxicity reduced the impact of the porphyrazine on normal
tissues in the absence of irradiation, and the potential
therapeutic index IC50(dark)/IC50(light) for compound 3
was significantly increased. In general, the data in Table
2 suggested that the meta- and ortho-substituents did not
alter the photodynamic activity. It is interesting to notice
that the therapeutic index was the lowest in the case of
the unsubstituted phenyl group.
In summary, we revealed the details of fine tuning of
the photophysical and photodynamic properties of the
cyanoarylporphyrazines for the enhancement of their
potential as photosensibilizers in cancer photodynamic
therapy and optical probing of the intracellular viscosity.
Fig. 2. Quantum yield and lifetime of porphyrazines 3, 5,
and 6 fluorescence as functions of medium viscosity (model
ethanol–glycerol mixtures).
of a cell. It is the intracellular viscosity which determines
the diffusion of various solutes and, therefore, the rate
of bimolecular intracellular reactions related to nutrition,
energy metabolism, and diverse signaling processes af-
fecting the cell viability.
EXPERIMENTAL
IR spectra of the suspensions in vaseline oil were ob-
tained using an FSM 1201 spectrometer. UV/vis absorp-
tion spectra were recorded using a PerkinElmer Lambda
25 spectrometer. 1Н NMR spectra were recorded using a
BrukerAvance DPX-200 instrument (200 MHz) at 25°С.
Stationary fluorescence was studied using a PerkinElmer
LS 55 spectrometer (300–800 nm). Mass-chromatograms
were obtained using a PolarisQ/TraceGCUltra chro-
mato–mass spectrometer. Dynamic viscosity of binary
ethanol(methanol)–glycerol mixtures was measured using
an SVM 3000 Stabinger viscometer (Anton Paar) with
accuracy of ±0.35%. The porphyrazines were dissolved
via mechanical mixing and ultrasonication during at least
15 min.
To evaluate the novel cyanoarylporphyrazines as
potential photosensibilizers, we measured their dark and
photoinduced cytotoxicity towards the epidermoid human
carcinoma cells, line А431. These parameters are typi-
cally determined via the MTT test [25], i.e. measurement
of a compound concentration decreasing the cells growth
(or inducing their death) by 50% (IC50). That value was
measured both in dark and under light; the obtained IC50
values are collected in Table 2. The listed data demonstrat-
ed noticeable photodynamic activity of the studied series
of compounds. The highest photoinduced cytotoxicity
was revealed for the cyanoarylporphyrazines containing
fluorine atoms in the peripheral aryl groups. The substi-
tution of the para-fluorine atom (2) with the OMe group
(3) marginally reduced the photoinduced cytotoxicity, yet
significantly (almost twofold) decreased the dark cyto-
The syntheses were performed using anhydrous
solvents. 2-Methoxybenzaldehyde, 4-methoxybenzalde-
hyde, 3-methoxy-4-ethoxybenzaldehyde, malonodinitrile,
and N-chlorosuccinimide were purchased from Sigma
ꢀꢀꢀꢁɚꢂꢀꢀꢀꢀꢀꢀꢀꢀꢀꢀꢀꢀꢀꢀꢀꢀꢀꢀꢀꢀꢀꢀꢀꢀꢀꢀꢀꢀꢀꢀꢀꢀꢀꢀꢀꢀꢀꢀꢀꢀꢀꢀꢀꢀꢀꢀꢀꢁEꢂꢀꢀꢀꢀꢀꢀꢀꢀꢀꢀꢀꢀꢀꢀꢀꢀꢀꢀꢀꢀꢀꢀꢀꢀꢀꢀꢀꢀꢀꢀꢀꢀꢀꢀꢀꢀꢀꢀꢀꢀꢀꢀꢀꢀꢀꢀꢁFꢂ
Viscosity, cP
Viscosity, cP
Viscosity, cP
Fig. 3. Rate constants of radiation (1) and radiationless (2) transitions of porphyrazines 3 (a), 5 (b), and 6 (c) as functions of the
ethanol–glycerol mixtures viscosity at 25°С.
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NEW CYANOARYLPORPHYRAZINES WITH HIGH SENSITIVITY
253
Table 2. The IC50 values under irradiation (λ = 615–635 nm) and dark cytotoxicity of cyanoarylporphyrazines with different
aryl groups at the macrocycle periphery
Comp. no.
IC50(light), mol/L
3.7×10–6
8.0×107
IC50(dark), mol/L
1.1×10–5
6.9×10–6
2.1×105
IC50(dark)/ IC50(light)
1
2
3
4
5
6
7
3.0
8.6
1.3×106
16.1
4.7
3×106
1.4×105
2.4×106
1.1×105
4.6
2.4×106
1.2×105
5.0
9×107
3.6×106
4 .0
Aldrich. Bis(indenyl)ytterbium was synthesized as de-
scribed elsewhere [26].
Synthesis of 2-aryl-1,1,2-tricyanoethanes 11–13.
The corresponding 2-aryl-1,1-dicyanoethylene (10 mmol)
was dissolved in 150 mL of EtOH and 1.30 g (20 mmol)
of KCN dissolved in 80 mL of water was added; the
reaction mixture was then diluted with 240 mL of water.
The obtained mixture was stirred during 45 min at room
temperature. 6 mL of 37% HCl was added, and the reac-
tion mixture was cooled on an ice bath. The precipitate
was filtered off, washed with water, and dried at room
temperature under reduced pressure.
Porphyrazines 1, 2, 4, and 7 were synthesized as
described earlier [14, 17, 20]. The aryltricyanoethylenes
were obtained via the three-stage synthesis from the cor-
responding substituted benzaldehydes [27].
Synthesis of 2-aryl-1,1-dicyanoethylenes 8–10.
0.99 g (15 mmol) of malonodinitrile and 2 drops of pi-
peridine were added to a solution of 15 mmol of the sub-
stituted benzaldehyde in 100 mL of EtOH. The reaction
mixture was stirred during 24 h at room temperature. The
precipitate was filtered off, washed with water (4×80 mL),
and dried at room temperature under reduced pressure.
2-(4-Methoxyphenyl)-1,1,2-tricyanoethane (11).
Yield 76%, white substance. IR spectrum, ν, cm–1: 2250
1
(C≡N). H NMR spectrum (CDCl3), δ, ppm (J, Hz ):
7.42 d (2Н, 3J = 8.7), 7.01 d (2Н, 3J = 8.7), 4.42 d [1Н,
CH(СN)–CH(CN)2, J = 5.9], 4.23 d [1Н, CH(СN)–
CH(CN)2, 3J = 5.9], 3.85 s (3Н, CH3О). Mass spectrum,
m/z (Irel, %): 211 (5).
2-(4-Methoxyphenyl)-1,1-dicyanoethylene (8).
Yield 75%, yellow solid. IR spectrum, ν, cm–1: 2223
(C≡N). 1H NMR spectrum (CDCl3), δ, ppm (J, Hz): 3.91 s
(3Н, CH3О), 7.01 d (2Н, СНАr, J = 8.9), 7.90 d (2Н,
СНАr, 3J = 8.9), 7.65 s [1Н, CH=C(CN)2]. Mass spectrum,
m/z (Irel, %): 184 (100).
3
3
2-(2-Methoxyphenyl)-1,1,2-tricyanoethane (12).
Yield 83%, white substance. IR spectrum, ν, cm–1: 2257
(C≡N). 1H NMR spectrum (CDCl3), δ, ppm (J, Hz): 3.94 s
(3Н, CH3О), 4.45 d [1Н, CH(СN)–CH(CN)2, 3J = 5.8],
2-(2-Methoxyphenyl)-1,1-dicyanoethylene (9).
Yield 78%, light yellow solid. IR spectrum, ν, cm–1: 2224
(C≡N). 1H NMR spectrum (CDCl3), δ, ppm (J, Hz): 3.93
3
4.82 d [1Н, CH(СN)–CH(CN)2, J = 5.8], 7.00 d (1Н,
3
3
s (3H, CH3О), 6.99 d (1H, J = 8.5 Hz), 7.08 t (1H, J
= 7.7 Hz), 7.59 t (1H, 3J = 7.2 Hz), 8.18 d (1H, 3J = 9.1
Hz), 8.31 s [1H, CH=C(CN)2]. Mass spectrum, m/z (Irel,
%): 184 (100).
СНАr, 3J = 8.3), 7.50 t (1Н, СНАr, 3J = 7.9), 7.58 d (1Н,
СНАr, 3J = 6.3). Mass spectrum, m/z (Irel, %): 211 (8).
2-(3-Methoxy-4-ethoxyphenyl)-1,1,2-tricyanoeth-
ane (13). Yield 88%, white substance. IR spectrum, ν,
cm–1: 2255 (C≡N). 1H NMR spectrum (CDCl3), δ, ppm (J,
Hz): 1.49 t (3Н, CH3CH2О, 3J = 7.0), 3.92 s (3Н, CH3О),
4.21 q (2Н, CH3CH2О, 3J = 7.0), 4.21 d [1Н, CH(СN)–
CH(CN)2, 3J 5.8], 4.39 d [1Н, CH(СN)–CH(CN)2, 3J =
5.8], 6.91–7.05 m (3Н, СНАr). Mass spectrum, m/z (Irel,
%): 255 (10).
2-(3-Methoxy-4-ethoxyphenyl)-1,1-dicyanoethyl-
ene (10). Yield 65%, yellow solid. IR spectrum, ν, cm–1:
2220 (C≡N). 1H NMR spectrum (CDCl3), δ, ppm (J, Hz):
1.52 t (3Н, CH3CH2О, 3J = 7.0), 3.93 s (3Н, CH3О), 4.21 q
(2Н, CH3CH2О, 3J = 7.0), 6.94 d (1Н, СНАr, 3J = 8.4),
7.39 d (1Н, СНАr, 3J = 7.4), 7.63 s (1Н, СНАr), 7.68 s [1Н,
CH=C(CN)2]. Mass spectrum, m/z (Irel, %): 228 (100).
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254
LERMONTOVA et al.
Synthesis of 1-aryl-1,1,2-tricyanoethylenes 14–16.
3406 (N–H), 2840 (O–CН3), 2201 (С≡N), 1603 (C=N),
1512, 1497 (CAr–CAr); 1255, 1217, 1043, 1028 (CAr–O–
CAlk). UV spectrum (Н2О), λmax, nm: 365 (Soret band),
576 (Q-band). 1H NMR spectrum (CDCl3), δ, ppm (J, Hz):
3.87 s (12Н, CH3О), 7.02 d (8Н, СНАr, 3J = 8.8), 7.44 d
(8Н, СНАr, 3J = 8.8), 7.56 s (2Н, NH). Mass spectrum, m/z
(Irel, %): 838 [M]+, 857 [M + Н2О]+, 808 [M – СН3О]+.
1.20 g (9.0 mmol) of N-chlorosuccinimide was added to
the corresponding 2-aryl-1,1,2-tricyanoethane (6.4 mmol)
dissolved in 100 mL of Et2O. The reaction mixture was
stirred at cooling (ice bath). After 1 h, 150 mL of water
was added, the organic layer was separated and washed
with water (3×150 mL). The solvent was removed under
reduced pressure. The product was sublimated under
reduced pressure.
Tetra(2-methoxyphenyl)tetracyanoporphyrazine
(5). Yield 28%, black solid. IR spectrum (KBr), ν, cm–1:
3435 (N–H), 3066 (CAr–Н), 2838 (O–CН3), 2205 (С≡N),
1598 (C=N); 1504, 1493 (CAr–Car); 1253, 1216, 1043,
1023 (CAr–O–CAlk). UV spectrum (Н2О), λmax, nm:
312, 380 (Soret band), 586 (Q-band). 1H NMR spectrum
(CDCl3), δ, ppm (J, Hz): 3.77 s (12Н, CH3О), 7.05 t (4H,
СНАr, 3J = 7.5), 7.15 d (8H, СНАr, 3J = 8.2), 7.52 t (4H,
2-(4-Methoxyphenyl)-1,1,2-tricyanoethylene (14).
Yield 69%, yellow solid. IR spectrum, ν, cm–1: 2226
(C≡N). 1H NMR spectrum (CDCl3), δ, ppm (J, Hz): 3.96 s
3
(3Н, CH3О), 7.08 d (2Н, СНАr, J = 9.1), 8.10 d (2Н,
СНАr, 3J = 9.1). Mass spectrum, m/z (Irel, %): 209 (100).
2-(2-Methoxyphenyl)-1,1,2-tricyanoethylene (15).
Yield 72%, yellow solid. IR spectrum, ν, cm–1: 2240
(C≡N). 1H NMR spectrum (CDCl3), δ, ppm (J, Hz): 3.99 s
(3Н, CH3О), 7.07 t (1Н, 3J = 8.3), 7.15 d (1Н, СНАr, 3J =
7.6), 7.55 d (1Н, СНАr, 3J = 6.6), 7.64 t (1Н, СНАr, 3J =
7.1). Mass spectrum, m/z (Irel, %): 209 (100).
3
СНАr, J = 8.8), 8.47 s (2Н, NH). Mass spectrum, m/z
(Irel, %): 838 [M]+, 824 [M – СН3]+, 808 [M – СН3О]+.
Tetra(3-methoxy-4-ethoxyphenyl)tetracyanopor-
phyrazine (6). Yield 34%, black solid. IR spectrum
(KBr), ν, cm–1: 3403 (N–H), 2853 (O–CAlk), 2201 (С≡N),
1597 (C=N); 1515, 1500 (CAr–CAr); 1257, 1217, 1030
(CAr–O–CAlk). UV spectrum (Н2О), λmax, nm: 383 (Soret
band), 576 (Q-band). 1H NMR spectrum (CDCl3), δ, ppm
(J, Hz): 1.42 t (12Н, CH3CH2О, 3J = 6.9), 3.76 s (12Н,
2-(3-Methoxy-4-ethoxyphenyl)-1,1,2-tricyano-
ethylene (16). Yield 95%, orange solid. IR spectrum, ν,
cm–1: 2220 (C≡N). 1H NMR spectrum (CDCl3), δ, ppm (J,
Hz): 1.54 t (3Н, CH3CH2О, 3J = 7.0), 3.95 s (3Н, CH3О),
4.25 q (2Н, CH3CH2О, 3J = 7.0), 7.00 d (1Н, СНАr, 3J =
8.3), 7.70–7.75 m (2Н, СНАr). Mass spectrum, m/z (Irel,
%): 253 (50).
3
CH3О), 4.13 q (8Н, CH3CH2О, J = 7.0), 6.97–7.09 m
(12Н, СНАr), 7.50 s (2Н, NH). Mass spectrum, m/z (Irel,
%): 1014 [M]+.
Measurement of photoinduced IC50(light) and
dark IC50(dark) cytotoxicity. Viability of the cells was
evaluated using the MTT test [25]. The optical density
was measured using a Synergy MX microplate spectro-
photometer (BioTek, USA) at 570 nm. The cells viability
was estimated from the ratio of the absorbance of the
formazan solution in the sample to that in the reference
specimen (without irradiation). In the case of the dark
cytotoxicity the irradiation was not performed.
Synthesis of ytterbium complexes 17–19. Asolution
of 2.3 mmol of the corresponding 2-aryl-1,1,2-tricyano-
ethylene 14–16 in 5 mL of degassed THF was added in
small portions under inert atmosphere to as solution of
0.25 g (0.46 mmol) of bisindenyl ytterbium(II) complex
in 5 mL of THF.After keeping for 1 day, the solution was
filtered in vacuum. To remove the unreacted 2-aryl-1,1,2-
tricyanoethylene and its ytterbium complex, the solution
as washed with degassed toluene until discoloration. The
reaction product was dried under reduced pressure.
Measurement of average fluorescence decay time
for compounds 3, 5, and 6. The porphyrazines fluores-
cence lifetime was measured over the picosecond time
scale using the time-correlated single photon counting
(TCSPC) technique. The measuring system included the
TCSPC components from Becker & Hickl, GmbH. The
data collection window was set to 0–50 ns with 1024
channels. The excitation was performed using an SC-450
picosecond fiber laser (Fianium Ltd, Великобритания)
with an adjustable (580–595 nm) output filter (Chroma
Tech., USA). The fluorescence signal was detected over
640–700 nm. Exponential fluorescence decay was due to
Synthesis of porphyrazines 3, 5, 6. A solution of
0.14 mmol of the corresponding ytterbium complex
17–19 in 2 mL of trifluoroacetic acid was stirred at room
temperature during 30 min. 30 mL of water was them
added, and the formed precipitate was centrifuged off and
thoroughly washed with water to neutral reaction of the
medium. The reaction product was purified by column
chromatography (silica gel 60, 40–60 μm, eluent: THF).
Tetra(4-methoxyphenyl)tetracyanoporphyrazine
(3). Yield 30%, black solid. IR spectrum (KBr), ν, cm–1:
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NEW CYANOARYLPORPHYRAZINES WITH HIGH SENSITIVITY
255
6. Niedre, M., Patterson, M.S., and Wilson, B.C., Photo-
the fundamental features of the relaxation kinetics of the
chem.
fluorophore excited state. The decay curves were analyzed
to reveal the best multiexponent model using SPCImage
software (Becker & Hickl, GmbH), followed by the data
processing in Excel. The decay time (τav) was calculated
as the average lifetime of the fluorescent components
weighted by the integral intensity.
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Here ai and τi are the amplitudes and lifetimes of the
components obtained via fitting of the experimental decay
curves using the chosen multiexponent model. The mea-
surements were performed using standard 96-well plates.
Temperature of the solutions of the cyanoarylporphyr-
azines in the ethanol–glycerol mixtures was maintained
constant using a ThermoStat plus laboratory thermostat
(Eppendorf, GmbH).
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FUNDING
This study was financially supported by the President of
Russia (grant MK-1469.2019.3, synthesis and study of prop-
erties of porphyrazine 1) and performed using the equipment
of Analytical Center, Institute of Organometallic Chemistry,
Russian Academy of Sciences in the scope of the state task
(no. АААА-А19-119011690055-0).
https://doi.org/10.1073/pnas.0912811107
CONFLICT OF INTEREST
No conflict of interest was declared by the authors.
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