Synthesis and photoactivity of a Pt(II) complex based on an o-nitrobenzyl-derived ligand

Article abstract of DOI:10.1016/j.ica.2012.06.047

Development of photoactivatable platinum complexes was an attractive approach to improve the anticancer behavior of Pt-based agents. In this work, a new photolabile Pt(II) complex, NMPt, was prepared using a photosensitive compound, 2-hydroxy-2-(2-nitrophenyl)acetic acid (HNPAC). X-ray crystallographic data disclosed that Pt(II) center of NMPt adopts a square-planar geometry similar to that of nedaplatin. In NMPt molecule, the Pt(II) center is coordinated by two oxygen atoms of the hydroxyl and carboxyl groups in HNPAC ligand and two ammine ligands. The quick conversion of NMPt (100 μM) in PBS buffer upon irradiation at 365 nm was observed, displaying a distinct absorption bathochromic shift from 265 to 300 nm. The ¹H NMR determination also confirmed the UV irradiation-induced conversion of NMPt complex. The CD spectroscopic determination revealed that NMPt is able to distort the conformation of CT-DNA only upon UV-irradiation. In addition, agarose gel analysis of plasmid pBR 322 DNA demonstrated that NMPt induces plasmid DNA nicking upon UV-irradiation with a distinct dose- and time-dependence. Preliminary results showed that the cytotoxicity of NMPt at lower concentration (5-20 μM) against human breast adenocarcinoma (MCF-7) cells is comparable to that of nedaplatin upon UV irradiation, while its cytotoxicity is very low without UV irradiation. Therefore, UV irradiation-induced DNA distortion and damage might be correlated to the photo-induced cytotoxicity enhancement for NMPt. This study provides an interesting approach for the exploration of photoactivatable Pt(II) complex as anticancer agent, although the photo-induced anticancer activity of NMPt remains to be improved.

Full text of DOI:10.1016/j.ica.2012.06.047

Inorganica Chimica Acta 393 (2012) 198–203
Contents lists available at SciVerse ScienceDirect
Inorganica Chimica Acta
journal homepage: www.elsevier.com/locate/ica
Synthesis and photoactivity of a Pt(II) complex based
on an o-nitrobenzyl-derived ligand
⇑
⇑
Di Liu, Jialiu Ma, Wen Zhou, Weijiang He , Zijian Guo
State Key Laboratory of Coordination Chemistry, School of Chemistry and Chemical Engineering, Nanjing University, Nanjing 210093, China
a r t i c l e i n f o
a b s t r a c t
Article history:
Available online 20 July 2012
Development of photoactivatable platinum complexes was an attractive approach to improve the anti-
cancer behavior of Pt-based agents. In this work, a new photolabile Pt(II) complex, NMPt, was prepared
using a photosensitive compound, 2-hydroxy-2-(2-nitrophenyl)acetic acid (HNPAC). X-ray crystallo-
graphic data disclosed that Pt(II) center of NMPt adopts a square-planar geometry similar to that of
nedaplatin. In NMPt molecule, the Pt(II) center is coordinated by two oxygen atoms of the hydroxyl
and carboxyl groups in HNPAC ligand and two ammine ligands. The quick conversion of NMPt
Metals in Medicine Special Issue
Keywords:
Platinum complex
Photoactivation
Cytotoxicity
Tumor
(100 lM) in PBS buffer upon irradiation at 365 nm was observed, displaying a distinct absorption bath-
ochromic shift from 265 to 300 nm. The 1H NMR determination also confirmed the UV irradiation-
induced conversion of NMPt complex. The CD spectroscopic determination revealed that NMPt is able
to distort the conformation of CT-DNA only upon UV-irradiation. In addition, agarose gel analysis of plas-
mid pBR 322 DNA demonstrated that NMPt induces plasmid DNA nicking upon UV-irradiation with a dis-
tinct dose- and time-dependence. Preliminary results showed that the cytotoxicity of NMPt at lower
DNA
concentration (5–20 lM) against human breast adenocarcinoma (MCF-7) cells is comparable to that of
nedaplatin upon UV irradiation, while its cytotoxicity is very low without UV irradiation. Therefore,
UV irradiation-induced DNA distortion and damage might be correlated to the photo-induced cytotoxic-
ity enhancement for NMPt. This study provides an interesting approach for the exploration of photoac-
tivatable Pt(II) complex as anticancer agent, although the photo-induced anticancer activity of NMPt
remains to be improved.
Ó 2012 Elsevier B.V. All rights reserved.
1. Introduction
complexes are the most studied prodrugs due to their more inert
kinetics than Pt(II) complexes, and their intracellular activation
Platinum-based anticancer drugs, including cisplatin, carbo-
platin, nedaplatin and oxaliplatin have been widely used in clinical
treatment of different cancers. However, their clinical application
is limited due to their low specificity for tumor tissue, distinct
resistance and high toxicity [1]. Various strategies to improve the
anticancer behavior of platinum complexes have been adopted.
Pt(II) complexes of novel structures such as monofuctional com-
pounds, multinuclear compounds, have been developed to improve
their activity and anticancer spectrum [2]. In addition, hybrid anti-
cancer compounds has also been investigated via integrating Pt(II)
complex with other anticancer agents such as photodynamic ther-
apy agent [3]. To decrease the side effects of Pt-based anticancer
agents, approaches for targeting delivery systems have been ex-
plored [4], except for common strategies of integrating targeting
groups for tumor with Pt complexes [5]. On the other hand, activat-
ing platinum-based prodrugs at specific sites is an attractive alter-
native to reduce the side effects of these compounds. Pt(IV)
via reduction to Pt(II) species by biological reducing agents should
be helpful to overcome certain disadvantages of Pt(II)-based clini-
cal drugs [6]. The tumor-targeting property of Pt(IV) complexes has
been demonstrated by Saldler and co-workers, i.e. active Pt(II) spe-
cies were released by photoactivation of non-toxic diazo Pt(IV)
complexes of octahedral geometry [7]. This approach displays a
site-specificity for photoactivation, which inspires scientists to de-
velop novel Pt-based complexes for the practical tumor-targeting
treatment, and photosensitive Pt(II) complex has also been investi-
gated [8].
Herein, a new photosensitive Pt(II) complex, NMPt, was con-
structed to investigate its anticancer activity especially upon
photo-irradiation (Scheme 1). In this complex, the ligand, 2-hydro-
xy-2-(2-nitrophenyl)acetic acid (HNPAC) is a derivative of photo-
sensitive o-nitrobenzyl alcohol. Both of its hydroxyl group and
carboxylic anion coordinate to the same Pt(II) center with a chelat-
ing effect similar to that of nedaplatin. As the normal o-nitrobenzyl
alcohol, the photo-induced internal redox process will reduce its
nitro group to nitroso group, and the hydroxyl group is oxidized
to carbonyl group in the mean time [9]. In fact, this photosensitiv-
ity has been frequently utilized to form photo-caged molecules in
⇑
Corresponding authors. Tel.: +86 25 83597066; fax: +86 25 83314502.
E-mail addresses: heweij69@nju.edu.cn (W. He), zguo@nju.edu.cn (Z. Guo).
0020-1693/$ - see front matter Ó 2012 Elsevier B.V. All rights reserved.
http://dx.doi.org/10.1016/j.ica.2012.06.047

D. Liu et al. / Inorganica Chimica Acta 393 (2012) 198–203
199
Scheme 1. Photo-induced conversion of o-nitrobenzyl alcohol and the proposed photo-induced conversion of NMPt complex.
biological studies [10]. Therefore, the Pt(II) chelating effect of li-
gand should be decreased distinctly, which may result in a more
hydrolytical Pt(II) species, altering its anticancer behavior upon
photo-irradiation.
the monoclinic space group of P21/c (Fig. 2), and there are only
one complex molecule and one water molecule in the unit cell.
The Pt center was coordinated in a square-planar geometry by
two amine nitrogen atoms and two oxygen atoms from hydroxyl
and carboxyl groups of HNPAC ligand. With the similar structure
to that of nedaplatin, the related coordination bonds around Pt(II)
center are almost the same to those found in nedaplatin. For exam-
ple, the length of Pt1–N1 bond is 2.05 Å, which is almost identical
to 2.04 Å found in nedaplatin [13]. Detailed structure parameters
have been shown in Table 1. Each water molecule form two hydro-
gen bonds (Ow–H. . .O) with two carboxylic O atoms from two dif-
ferent NMPt molecules, in addition, this water molecule still form
the third hydrogen bond (Ow. . .H–N) with NH3 from the third
NMPt molecule.
2. Results and discussion
2.1. Synthesis and characterization of NMPt
Ligand HNPAC was prepared from o-nitrophenylaldehyde via a
two step procedure reported by Rossi and Kao [11]. Then, complex
NMPt was prepared as a pale yellow powder using a modified pro-
cedure for nedaplatin preparation reported by Wang and co-work-
ers [12]. In the MS spectrum of NMPt, three signals of m/z 446.92,
870.67 and 1295.17 were observed. These signals can be assigned
to [M+Na]+, [2M+Na]+ and [3M+Na]+, respectively by comparing
the determined isotopic distribution patterns with the calculated
ones given by ISOPRO 3.0 (M = NMPt, Fig. 1). This result suggests
this complex was stable in the condition of MS determination. Sin-
gle crystals suitable for X-ray crystallographic determination was
obtained by recrystallizing NMPt from methanol in fridge at 4 °C.
Structural resolution disclosed that this complex crystallizes in
2.2. Photolysis of NMPt
Although the photosensitivity of o-nitrobenzyl derivatives has
been reported [9,10], the photosensitivity of NMPt formed by
HNPAC should be determined. Since NMPt has fine aqueous solu-
bility, and the UV–Vis spectrum of NMPt was investigated in PBS
buffer (100 lM) of pH 7.4 upon UV irradiation (365 nm) in a
photoreactor. NMPt displays a stable absorption spectrum without
UV irradiation, showing a minor band centered at 262 nm with an
extinction coefficient of 4217.9 Mꢀ1cmꢀ1. Upon UV irradiation, the
UV–Vis spectra of NMPt were recorded every 20 s, and a new
absorption band centered at 300 nm was observed upon UV irradi-
ation. This band increased gradually with the irradiation time
accompanied by a gradual decrease of former band at 262 nm
(Fig. 3), implying there is a photo-induced conversion of NMPt
complex. The UV–Vis spectra became stable in 3 min, displaying
the photo-induced conversion has been finished, and the final
extinction coefficient at 300 nm is 5203.4 Mꢀ1cmꢀ1. The clear iso-
bestic points at 255 and 276 nm also confirmed the conversion
process, and the solution of NMPt turned from colorless to pale
brown. Since the absorbance at 300 nm is the characteristic band
of o-nitrosophenyl group, a typical change of o-nitrobenzyl alcohol
derivatives to o-nitrosophenyl group upon UV irradiation should
Fig. 1. (A) Observed isotopic distribution patterns (IDPs) of main signals found in
the ESI-MS spectrum of complex NMPt. (B) Calculated IDPs of [NMPt + Na]+,
[2NMPt + Na]+, and [3NMPt + Na]+.
Fig. 2. Molecular structure of NMPt. Water molecule and hydrogen atoms were
omitted for clarity.

200
D. Liu et al. / Inorganica Chimica Acta 393 (2012) 198–203
Table 1
Selected bond lengths (Å) and angles (°) for NMPt.
Pt(1)–N(1)
Pt(1)–N(2)
Pt(1)–O(1)
Pt(1)–O(2)
2.045(7)
2.030(8)
2.005(6)
2.054(7)
O(1)–Pt(1)–N(1)
92.9(3)
85.1(3)
91.7(3)
90.5(3)
176.7(2)
O(1)–Pt(1)–O(2)
N(2)–Pt(1)–O(2)
N(2)–Pt(1)–N(1)
N(1)–Pt(1)–O(2)
be occurred. The bathochromic shift of absorbance from 262 to
300 nm for NMPt complex suggested
a
Norrish type II
photoreaction.
The photosensitivity of complex NMPt (5 mM) has also been
investigated using 1H NMR spectroscopy in D2O. Similar to the
UV–Vis determination, stable 1H NMR spectrum was observed
and new peaks were observed in the down field only upon UV irra-
diation. As shown in Fig. 4, the three new minor signals labeled
with asterisk can be assigned as signals for protons b0, c0 and d0.
It is the result of down-field shift of signals for protons b, c and d
due to the photo-induced conversion of NMPt to complex 1. It is
clear that the conversion of NMPt to complex 1 will decrease the
density of delocalized electrons of phenyl ring due to the formation
carbonyl group at the para position of nitroso group, which caused
the down-field shift for the signals of the three protons on aromatic
ring. According to the rule to estimate chemical shift of protons on
aromatic ring, this Norrish type II conversion of HNPAC would in-
duce a neglectable down-field shift for proton a, which is consis-
tent with our observed spectra. The 1H NMR data implied that
only 10% of NMPt has been converted into complex 1 in D2O after
10 min of UV irradiation. Longer photo-irradiation resulted in a
distinct brown precipitate.
Fig. 4. 1H NMR spectra of NMPt in D2O determined before and after 10 min of
irradiation. The signals labeled with asterisks are the new signals observed after
irradiation in the down field.
helicity and base stacking can be altered by the irradiated complex.
These results suggested that the photo-irradiation is able to con-
vert NMPt of poor DNA binding ability into species which can
interact with DNA effectively.
2.4. Agarose gel electrophoresis of plasmid pBR322 DNA in the
presence of NMPt
Agarose gel assay of supercoiled DNA (plasmid pBR322 DNA)
mixing with NMPt has also been carried out to explore the
photo-irradiation effect of NMPt on the interaction between plas-
2.3. CD spectra of CT-DNA in the presence of NMPt
mid DNA and NMPt. Therefore, DNA (200 ng/
with NMPt (0–100 M) at 37 °C with or without UV irradiation
for 3 h. As can be seen from Fig. 6, incubating plasmid DNA with
NMPt (100 M) in dark for 3 h resulted in supercoiled DNA (form
lL) was incubated
l
It was well recognized that the formation of covalent cross links
between platinum complex and DNA is closely related to the anti-
cancer activity of platinum-based drugs. Therefore, the DNA bind-
ing behavior of NMPt was investigated with or without UV
irradiation to explore whether photo-irradiation would alter its
DNA binding behavior. As shown in Fig. 5, the CD spectra of CT-
DNA (0.1 mM) displayed that 40 h of incubation with NMPt at
37 °C in dark did not induce any obvious change at all tested
[NMPt]/[DNA] ratio (0–2.0), implying poor DNA binding ability of
NMPt. However, if the mixture of CT-DNA and NMPt was UV-irra-
diated for 3 min before incubation, both the negative (245 nm) and
positive (275 nm) bands of CD spectrum of CT-DNA decreased in
intensity after 40 h of incubation in dark. Higher complex concen-
tration resulted in a larger decrement. It seems that both the DNA
l
I) with very minor nicked DNA (Form II, lane 1). Photo-irradiation
of DNA with no NMPt complex for 3 h displayed a similar gel pat-
tern with only nicked DNA being slightly enhanced (Lane 2). The
gel patterns shown in lanes 1 and 2 suggested that either NMPt
or UV irradiation alone could not disturb supercoiled DNA. Distinct
enhancement of nicked DNA was observed only upon UV irradia-
tion in the presence of NMPt (Lanes 3–6). The accompanied de-
crease of form I DNA was also observed in the mean time. Higher
NMPt concentration resulted in larger amount of nicked DNA
and smaller amount of form I DNA. Besides the distinct dose-
dependent nicking efficiency of supercoiled DNA promoted by
NMPt, the cleavage to form nicked DNA can also be promoted by
1.0
0.5
0.4
0.8
0.3
0.6
0.2
0
60
120 180 240
UV exposure / s
0.4
0.2
0.0
(a)
200 250 300 350 400 450 500 550
Wavelength (nm)
Fig. 3. (a) UV–Vis spectra of 100 lM NMPt in PBS buffer (100 lM, pH 7.4) determined upon UV irradiation (365 nm, 4 min) in a photoreactor. The spectra were recorded with
an interval of 20 s. Inset is the temporal profile of absorbance at 300 nm upon UV-irradiation. (b) Photograph of non-irradiated and UV-irradiated NMPt (100 lM) solutions in
PBS buffer.

D. Liu et al. / Inorganica Chimica Acta 393 (2012) 198–203
201
10
10
8
[NMPt]/[DNA]
[NMPt]/[DNA]
3-min irradiation
8
6
0
0
0.2
0.5
1.0
2.0
0.2
0.5
1.0
2.0
6
4
4
2
2
0
0
-2
-4
-6
-2
-4
-6
(a)
320
(b)
320
220
240
260
280
300
220
240
260
280
300
Wavelength (nm)
Wavelength (nm)
Fig. 5. CD spectra of CT-DNA (0.1 mM) treated with complex NMPt at different [NMPt]/[DNA] ratios (0–2.0). (a) Spectra determined after 40 h of incubation with NMPt at
37 °C in dark; (b) spectra determined after 40 h of incubation with NMPt which has been UV-irradiated at 365 nm for 3 min.
1.0
NMPt in dark
NMPt with irradiation
Nedaplatin in dark
Nedaplatin with irradiation
0.8
0.6
0.4
0.2
0.0
5
10
20
50
100
200
NMPt concentration
(
µ
M
)
Fig. 7. Cytotoxicity of NMPt and nedaplatin against MCF-7 cells in dark or upon UV
irradiation at 365 nm.
cell lines. For nedaplatin, no distinct photo-induced enhancement
of inhibition rate was observed at all tested concentrations. The
distinct photo-induced enhancement of inhibition rate might be
correlated to the photo-induced conversion of NMPt, which en-
hances the DNA binding ability and alters DNA conformation. In
addition, the photo-induced DNA nuclease activity may also con-
tribute to the photo-induced enhancement of anticancer activity
at lower concentration. It should be noted that NMPt displayed a
very lower cytotoxicity to MCF-7 cells at lower complex concentra-
Fig. 6. (a) Agarose gel electrophoresis patterns of plasmid pBR322 DNA (200 ng/
incubated with complex NMPt of different concentrations at 37 °C. Concentration of
NMPt for Lanes 2–6: 0, 10 M, 20 M, 50 M, 100 M, and the UV-irradiation time
is 3 h. Lane 1: control of DNA incubated with 100 M NMPt in dark. (b) The related
histogram of percentages of Form I and Form II DNA induced by photo-irradiation in
the presence of NMPt complex.
lL)
l
l
l
l
l
tion (610 lM) in dark, when compared with nedaplatin. However,
photo-irradiation enhanced its cytotoxicity to be comparable to
that of nedaplatin at same concentration without UV irradiation.
increasing the photo-irradiation time at the same NMPt concentra-
tion. All these results indicated that photo-irradiation is able to
induce the cleavage of DNA in the presence of NMPt.
3. Conclusion
2.5. Cytotoxicity of NMPt against human breast adenocarcinoma cells
(MCF-7 cells)
A new photo-sensitive complex, NMPt, was prepared with a
photo-sensitive o-nitrobenzyl derivative as ligand. This complex
possesses a similar structure to that of nedaplatin, and displays a
distinct photo-induced conversion upon UV irradiation. Enhanced
DNA binding and cleavage activities were also observed upon UV
irradiation. Although the inhibition rate of NMPt to MCF-7 cells
is low in dark at lower concentration, photo-irradiation promotes
its cytotoxicity to a level comparable to that of nedaplatin in dark
at the same concentration. Further biological investigations are
warranted to reveal the molecular mechanism of NMPt, which will
provides helpful insights into the rational design of photoactivat-
able platinum-based antitumor complexes.
On the basis of the above mentioned experimental results, the
cytotoxicity of NMPt against human breast adenocarcinoma cells
(MCF-7) was determined respectively with or without UV irradia-
tion. Nedaplatin was taken as a positive control for comparison due
to their similar coordination structure. For NMPt, the distinct
photo-induced enhancement of inhabitation rate was observed
when the incubation concentration is lower (Fig. 7). For example,
NMPt showed almost no inhibition to MCF-7 cells at 5
ꢁ13% inhibition rate was observed upon photo-irradiation. The
M of NMPt, while it was en-
lM, while
inhibition rate in dark is ꢁ3% at 10
l
hanced to ꢁ18% upon photo-irradiation. The photo-irradiation in-
duced enhancement factor for NMPt is above 6-fold at these two
cases. Interestingly, when the NMPt concentration was increased
4. Experimental
from 20 to 200
for inhibition rate was reduced quickly, and the photo-induced
enhancement factor at 100
M is ꢁ1.1-fold. This demands more
detailed biological investigation and further experiments on other
l
M, the irradiation-induced enhancement factor
4.1. Material and methods
l
All chemicals and solvents were of reagent grade and used as
received. 1H NMR spectra were recorded on a Bruker DRX-500

202
D. Liu et al. / Inorganica Chimica Acta 393 (2012) 198–203
spectrometer. Electrospray mass spectra were determined using
an LCQ electron spray mass spectrometer (ESI-MS, Finnigan). The
isotopic distribution patterns for the complex were simulated by
ISOPRO 3.0 program. UV–Vis spectra were recorded with a UV-2550
photospectrometer (Shimadzu). X-ray crystallographic data were
recorded on a Siemens Bruker SMART CCD diffractometer using
irradiation in photoreactor with a filter of 365 nm. The DNA without
NMPt was irradiated and incubated as control. In addition, DNA
incubated NMPt (100
was also measured as a control. After being mixed with 2
l
M) incubated at 37 °C without irradiation
L loading
l
buffer, all the quenched samples were loaded respectively onto a 1%
agarose gel. Electrophoresis was carried out in TAE buffer (40 mM
Tris–acetate/1 mM EDTA) at 60 V for 2–3 h. Gels were stained with
ethidium bromide and visualized using UVP gel doc system, and
the gel patterns were analyzed with Quantity One 4.6.2.
graphite monochromatized Mo K
a radiation (k = 0.71037 Å).
The crystal structure was resolved by direct methods using the
program SHELXL-97 [12]. Photolysis experiments were all performed
in photoreactor offered by Xujiang company, Nanjing.
4.6. Cytotoxicity of NMPt against MCF-7 cells
4.2. Preparation of NMPt
Cytotoxicity of NMPt against MCF-7 cell lines were tested by
MTT assay. MCF-7 cells were seeded in a 96-well plate and incu-
bated in Dulbecco’s modified Eagle medium (DEME) overnight.
NMPt in PBS buffer solution (pH 7.4) were added to the culture
medium to a desired concentration. The irradiation was carried
out by UV irradiation at 365 nm for 3 min. Then the cells were
Ligand HNPAC was synthesized according to the reported pro-
cedure [11]. NMPt were synthesized in dark using a modified pro-
cedure for nedaplatin [12]. Therefore, cisplatin (600 mg) were
suspended in 25 mL water and 624 mg Ag2SO4 were added with
stirring. After being stirred at room temperature for 24 h, the pre-
cipitates were removed by filtration. The obtained filtrate was
mixed with a methanolic solution (20 mL) containing 375 mg
HNPAC. Then, an aqueous solution prepared by dissolving
599 mg Ba(OH)2ꢂ8H2O in 20 mL water was added dropwise into
the mixture in 6 h with stirring. After stirring in dark at room tem-
perature for 48 h, the precipitate was removed by filtration and
50 mL methanol was added. Then, the solvents were removed in
vacuo and the residue was washed by methanol and acetone in se-
quence. NMPt (310 mg) was obtained as pale yellow powders with
a yield of 38%. The single crystal for crystal structure resolution
was obtained by slow evaporation of its methanolic solution. 1H
NMR (500 MHz, DMSO-d6, d, ppm): 8.14 (d, J = 7.6 Hz, 1H), 7.71
(d, J = 8.0 Hz, 1H), 7.62 (t, J = 7.3 Hz, 1H), 7.41 (t, J = 7.7 Hz, 1H),
5.55 (s, 1H). ESI-MS (m/z, positive mode): found 446.9, calcd.
447.0 for [M+Na]+; found 870.7, calcd. 871.1 [2M+Na]+; found
1294.7, calcd. 1295.1 for [3M+Na]+.
incubated for 48 h and 20
wells, respectively. The medium were removed after 4 h of incuba-
tion. Then, 200 L DMSO were added to each well and the absor-
lL MTT (5 mg/mL) were added to the
l
bance was measured at 570 nm using plate reader (Varioskan
Flash, Thermo Scientific). Each assay was performed independently
three times. The inhibition rate was assessed using the following
equation:
%Inhibition rate ¼ ðODcontrol ꢀ ODdrugÞ=ðODcontrol ꢀ ODblanckÞ ꢃ 100
Acknowledgments
We thank the National Basic Research Program of China (No.
2011CB935800) and National Natural Science Foundation of China
(No. 10979019, 21131003, 90713001 and 21021062) for financial
support.
4.3. UV–Vis and 1H NMR spectra of NMPt upon UV irradiation
Appendix A. Supplementary material
One hundred micromolar of NMPt in PBS buffer (100 lM, pH
7.4) were added into a cylinderic quartz cuvette and irradiated in
a photoreactor equipped with a filter of 365 nm, and UV–Vis spec-
tra were recorded every 20 s until no change was observed.
In addition, 1H NMR spectra of NMPt in D2O (5 mM) were deter-
mined also before and after UV irradiation for 10 min.
Supplementary data associated with this article can be found, in
the online version, at http://dx.doi.org/10.1016/j.ica.2012.06.047.
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