Platinum(II) complexes of reduced amino acid ester Schiff bases: Synthesis, characterization, and antitumor activity

Article abstract of DOI:10.1007/s11164-012-0973-3

A series of platinum(II) complexes of reduced amino acid esters Schiff bases were synthesized as potential anticancer agents and characterized by ¹H NMR, EA, IR, and molar conductivity. These compounds were tested for their DNA interaction with salmon sperm DNA by ultraviolet spectrum and CD spectrum, and their in vitro anticancer activities have been validated against HL-60, KB, BGC-823, and Bel-7402 cell lines by MTT assay. The cytotoxicity of complexes 5d and 5f are better than cisplatin against Bel-7402 cell lines, and show a close cytotoxic effect against HL-60 cell line.

Full text of DOI:10.1007/s11164-012-0973-3

Res Chem Intermed
DOI 10.1007/s11164-012-0973-3
Platinum(II) complexes of reduced amino acid ester
Schiff bases: synthesis, characterization, and antitumor
activity
•
Li-Jun Li Cheng Wang Yu Qiao
•
•
•
Xiao-Yan Yang Xing-Xing Hua Jian-Long Du
•
Received: 12 September 2012 / Accepted: 8 December 2012
Ó Springer Science+Business Media Dordrecht 2012
Abstract A series of platinum(II) complexes of reduced amino acid esters Schiff
bases were synthesized as potential anticancer agents and characterized by 1H
NMR, EA, IR, and molar conductivity. These compounds were tested for their DNA
interaction with salmon sperm DNA by ultraviolet spectrum and CD spectrum, and
their in vitro anticancer activities have been validated against HL-60, KB, BGC-
823, and Bel-7402 cell lines by MTT assay. The cytotoxicity of complexes 5d and
5f are better than cisplatin against Bel-7402 cell lines, and show a close cytotoxic
effect against HL-60 cell line.
Keywords Platinum(II) complexes Á Reduced amino acid esters Schiff-bases Á
DNA interaction Á Antitumor activity
Introduction
Cancer is a biggest health hazard for humanity [1]. Despite the dramatic development
of antitumour drugs, the cancer death rate remains constant. Cisplatin is the important
currently used platinum-based anticancer drugs [2]. Currently cisplatin is being used
as an anticancer agent in several human cancers, particularly in ovarian, testicular,
bladder, head, and neck cancers [3, 4]. However, structural analogues to cisplatin have
not yet overcome cisplatin’s clinical limitations, particularly with regard to a dose-
limiting toxicity, such as nephrotoxicity, neurotoxicity, ototoxicity, nausea, etc., and
the lower activity (acquired resistance) in some of the most common cancers [5]. In
order to overcome severe side effects, numerous platinum-based complexes have
been synthesized, like carboplatin and oxaliplatin [6, 7]. Furthermore, there are some
L.-J. Li (&) Á C. Wang Á Y. Qiao Á X.-Y. Yang Á X.-X. Hua Á J.-L. Du (&)
Chemical Biology Key Laboratory of Hebei Province, Hebei University, Baoding 071002, China
e-mail: lljhbu@163.com
J.-L. Du
e-mail: dujl@hbu.edu.cn
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L.-J. Li et al.
other platinum anticancer drugs like platinum(II)-containing oxaliplatin, carboplatin,
picoplatin (JM473), nedaplatin (JM118), or redox-active platinum(IV)-based
satraplatin, tetraplatin, and ormaplatin [8].
Over the last few decades, transition metal complexes have been well studied
for their application as artificial nucleases, because of their diverse structural
features and the possibility to tune their redox potential through the choice of
ligands [10–12]. Transition metal complexes containing Schiff base ligands and
their reduced products are often used as artificial chemical nucleases, and some of
these complexes have proved to be efficient DNA cleavage reagents [13, 14].
Amino acids are the body’s essential nutrients. They are the basic unit of body
proteins and certain enzymes. Amino acids play an important physiological role in
the body. When amino acids are esterified and introduced to the drug molecules,
they can enhance the drug lipophilicity, ease drug toxicity, and improve drug
bioavailability. Most of the model studies of the metal complexes of Schiff base
ligands containing salicylaldehyde and amino acid esters have focused upon the
binding mode of these ligands [15–17]. Structural studies on the metal complexes
of reduced Schiff base ligands, derived from various amino acid esters and
salicylaldehyde, are well documented [18–20]. Compared to the Schiff bases, the
corresponding reduced Schiff bases are expected to be more stable and adaptable
to form conformationally flexible 5- or 6-membered rings upon complexation as
they are not constrained to be planar. Although there are numerous reports on
transition metal complexes of reduced Schiff bases derived from amino acids [21,
22], information on the corresponding derivatives of platinum(II) is still rare. We
have prepared a series of reduced Schiff bases derived from amino acid esters
coordinated to platinum(II), the interaction between salmon sperm DNA and these
complexes was investigated by ultraviolet spectrum and CD spectrum, and their
antitumor activities have also been tested on HL-60, Bel-7402, BGC-823, and KB
cell lines in vitro.
Results and discussion
Ligands 4a–4f were synthesized from salicylaldehyde and 5-bromsalicylaldehyde
with glycine ester. The platinum(II) complexes 5a–5f have been prepared by the
reaction of K2PtCl4 with reduced Schiff bases derived from glycine esters in a
mixture of CH3OH/H2O in the room temperature (See Scheme 1).
Comparison of the IR spectra of the free ligands with that of their Pt(II) complexes,
the (N–H) stretching frequencies were found in free ligands at about
3,303–3,352 cm-1, while in complexes they shifted to high wavenumbers at about
3,420–3,445 cm-1. On the other hand, the (C=O)carbonyl stretching frequency was
found at about 1,735–1,742 cm-1 in the free ligands, whereas it shifted to about
1,601–1,648 cm-1 in the complexes. In the spectra of metal complexes, the bands for
the phenolic group (C–O) shifted from about 1,256–1,266 cm-1 to 1,263–1,270 cm-1
,
compared to the free ligands. New bands appeared at about 611–620, 519–524, and
412–429 cm-1, and were assigned to tPt–OAr, tPt–N, and tPt–O=C, respectively. The IR
spectra of 4b and 5b are shown in Fig. 1. All of these indicated that nitrogen atoms of
123

Platinum(II) complexes of reduced amino acid ester Schiff-bases
O
N
R1
CHO
O
CH2Cl2
R2
OH
R2
OH
.
H2N
HCl
R1
Et3N
1
2
3
R1
O
N
H
NH R1
OH
O
Pt
NaBH4
K2PtCl4
R2
O
R2
Cl
CH3OH/H2O
4
5
a: R1=OMe R2=H; b: R1=OEt R2=H; c: R1=OMe R2=Br
d: R1=OEt R2=Br; e: R1=OMe R2=Cl; f: R1=OEt R2=Cl
Scheme 1 Synthetic pathway for the preparation of compounds
4b
5b
4000
3500
3000
2500
2000
1500
1000
500
0
Wavenumbers (cm-1)
Fig. 1 The IR spectrum of 4b and 5b
the imino groups and oxygen atoms of the carbonyl groups and phenolic groups are
coordinated to the metal ion (see Table 1).
1
Although the overall pattern of the H NMR spectra of the complexes 5a–5f
very closely resembled thoswe of the free ligands, the signals have been shifted to
lower fields. The mass spectra of the complexes 5a–5f have molecular peaks, and
the elemental analysis data of the complexes 5a–5f were in good agreement with
the calculated values. The conductivity data of all complexes were also measured.
The molar conductance values of platinum(II) complexes in deionized methanol
are in the range of 40–60 S cm2 mol-1 [23]. All complexes are therefore
electroneutral.
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L.-J. Li et al.
Table 1 Main IR absorptions of ligands and complexes (cm-1
)
t (N–H)
t (C=O-)
t (ph–O)
t (Pt–OAr)
t (Pt–N)
t (Pt–O=C)
4a
4b
4c
4d
4e
4f
3,352
3,332
3,309
3,303
3,309
3,303
3,420
3,445
3,445
3,426
3,440
3,423
1,735
1,739
1,742
1,737
1,741
1,736
1,631
1,645
1,645
1,643
1,648
1,601
1,256
1,256
1,266
1,266
1,265
1,265
1,270
1,265
1,268
1,265
1,264
1,263
5a
5b
5c
5d
5e
5f
412
429
420
429
420
429
520
519
520
519
524
519
619
611
620
615
615
611
Interaction between complexes and DNA
The interaction between DNA and the complexes was characterized in the
ultraviolet spectrum. Some DNA binding drugs produce direct DNA strand breaks.
Some possess planar multiring chromophores and are also able to bind noncova-
lently to DNA by intercalation between base pairs. Other drugs bind into the minor
groove of DNA [24]. In the wavelength range of 200–400 nm, DNA with different
concentrations of complexes (5c and 5d) was determined (see Figs. 2, 3). As can be
seen from Fig. 2, there was one positive peak at 258 nm (A = 1.4048) due to the
absorption of DNA. With the increased concentration of complexes 5c
(c5c:cDNA = 0.1, 0.3 and 0.5), the absorbance values were increased (A = 1.4930,
1.8788 and 2.3235), and the abosorption at 258 nm shifted to low wavelength
(k = 256, 255, and 251 nm). Also, in Fig. 3, there was one similar positive peak at
258 nm (A = 1.4548). When the concentration of complexes 5d (c5d:cDNA = 0.1,
0.3, and 0.5) was increased, the absorbance values were enhanced (A = 1.8461,
2.1749, and 2.2781); meanwhile, the abosorption at 258 nm shifed to low
wavelength (k = 257, 255, and 254 nm). This might be because the complex was
inserted into the DNA double helix structure and stacking with DNA base pairs
where p-electron accumulation occurred, thus the absorbance value increased and
maximum absorption wavelength was blue-shifted. The results confirmed that
hyperchromicity occurred after coordination and the binding mode of complexes
with DNA was right for intercalation [25–27].
CD (circular dichroism) is a very powerful technique to monitor the conforma-
tional state of the DNA double helix in solution. The CD spectrum of DNA
displayed a canonical B-form conformation with a large positive band at
270–280 nm and a negative band at 248 nm. Later, the CD spectra of 5d DNA
was recorded and displayed in Fig. 4, where the 275 nm band was due to base
stacking and 248 nm band to right-handed helicity. The decrease in the intensity of
123

Platinum(II) complexes of reduced amino acid ester Schiff-bases
5
c5c:cDNA = 0.5
c5c:cDNA = 0.3
c5c:cDNA = 0.1
DNA Blank
4
3
2
1
0
300
400
wavelength (nm)
Fig. 2 Ultraviolet spectrum of interaction between 5c and salmon sperm DNA
5
c5d:cDNA = 0.5
c5d:cDNA = 0.3
4
3
2
1
0
c5d:cDNA = 0.1
DNA Blank
300
400
wavelength (nm)
Fig. 3 Ultraviolet spectrum of interaction between 5d and salmon sperm DNA
the positive band and the increase in negative bands of 5d were exhibited, which
was typical of intercalation involving p-stacking and stabilization of the right-
handed B form of DNA. This indicates a intercalative binding producing a
consequent weakening of the base stacking interactions [28]. So the results
confirmed that, after coordination, the binding mode of complexes with DNA might
be intercalation.
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L.-J. Li et al.
12
10
8
6
4
2
0
-2
-4
Δε
-6
c5d:cDNA = 0.5
c5d:cDNA = 0.3
c5d:cDNA = 0.1
DNA Blank
-8
-10
-12
-14
-16
-18
-20
250
300
350
400
wavelength (nm)
Fig. 4 CD spectrum of interaction between 5d and salmon sperm DNA
Table 2 The cytotoxicity of complexes in vitro (IC50
)
Complex
IC50 (lM)
Bel-7402
HL-60
KB
BGC-823
5a
16.35
18.98
15.82
4.91
16.70
19.52
9.92
21.30
24.10
22.70
13.26
24.84
19.68
2.65
19.52
24.94
23.05
21.49
26.76
24.77
6.84
5b
5c
5d
3.01
5e
14.66
7.70
10.74
6.87
5f
Cisplatin
8.12
2.89
In vitro cytotoxic activity
The cytotoxic activities of 5a–5f were evaluated against four human cancer cell
lines consisting of HL-60, BGC-823, KB, and Bel-7402, and the results are listed in
Table 2. As shown in Table 2, almost all the complexes exerted cytotoxic effects
against the tested carcinoma cell lines with a lower IC50 value (\50 lM); moreover,
they have selectivity against the tested carcinoma cell lines. Complex 5c displayed
the best cytotoxicity among the six complexes against the tested carcinoma cell
lines. It can be seen that complexes 5d and 5f were more active against Bel-7402
cell lines than cisplatin, fairly active against HL-60, but less active than cisplatin
against the BGC-823 cell line. The complexes 5a, 5c, and 5e were fairly active
against cisplatin and against the Bel-7402 cell line. The complexes 5a–5c and 5e
demonstrated less active cytotoxicity than cisplatin against KB and the HL-60 cell
line, while all the complexes had less active cytotoxicity against the BGC-823 cell
line in compared with cisplatin.
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Platinum(II) complexes of reduced amino acid ester Schiff-bases
Conclusion
In this study, six platinum(II) complexes with reduced Schiff bases derived from
glycine ester as ligands were synthesized and characterized. All complexes were
tested for their DNA interaction ability with salmon sperm DNA using ultraviolet
spectrum and CD spectrum, and the results showed that after coordination the
binding mode of complexes with DNA might be intercalation. The in vitro
anticancer activities of the platinum complexes have been validated against HL-60,
KB, BGC-823, and Bel-7402 cell lines by MTT assay and the results show that the
complexes 5a–5f have selectivity against the tested carcinoma cell lines. The
complexes 5d and 5f have the best cytotoxicity among the six complexes, and
moreover their cytotoxicities are better than that of cisplatin against the Bel-7402
cell line, and display a fairly cytotoxic effect against the HL-60 cell line.
Platinum(II) complexes with reduced Schiff bases derived from glycine ester might
be a promising source of metal-based antitumor agents.
Experimental
All reagents and chemicals were purchased from commercial sources and used as
received. Salicylaldehyde and K2PtCl4 were of chemical grade, L-amino acids were
of analytical grade, MTT and salmon sperm DNA were from Sigma. Four different
human carcinoma cell lines: HL-60 (immature granulocyte leukemia), Bel-7402
(liver carcinoma), BGC-823 (gastrocarcinoma), and KB (nasopharyngeal carci-
noma) were obtained from the American Type Culture Collection.
Instrumentation and measurement
Elemental analysis were determined on a Exeter Analytical CE-440 elemental
analyzer. The IR spectra were recorded using KBr pellets and a Thermo Nicolet 380
1
spectrophotometer. The H NMR spectra were recorded in DMSO-d6 on a Bruker
AVIII 600 NMR spectrometer. The mass spectra were measured by Bruker apex-
ultra 7.0T. The interaction between DNA and complexes was measured on an
UV-3400 Toshniwal spectrophotometer. The CD spectrum was measured on an
MOS-450 (BioLogic).The conductivity values were determined on a DDSJ-308A
conductivity meter. Molar conductances at room temperature were measured in
1 9 10-3 M methanol using a DDS-12DW-type conductivity meter.
Synthesis of compounds
Ligands synthesis
5-Bromosalicylaldehyde was synthesized according to a published procedure [29].
Amino acid esters were synthesized according to a published procedure [30].
The synthesis of amino acid ester reduced Schiff bases were according to one-
pot. Amino acid esters (4.68 mmol) were added to the round-bottom flask
containing dichloromethane (20 mL), then triethylamine (4.75 mmol) and a certain
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L.-J. Li et al.
amount of anhydrous magnesium sulfate were added to the flask at room
temperature, stirring for 1 h. Then salicylaldehyde (4.80 mmol) was added to the
dichloromethane solution dropwise, stirring for another 36 h at room temperature.
Methanol solution (10 mL) containing sodium borohydride (7.02 mmol) was then
added to the above solution and stirred for 3 h, followed by rotary evaporation and
washing three times. The amino acid ester-reduced Schiff bases were purified by
column chromatography (ethyl acetate/petroleum ether) to give the white solid
product.
Complexes synthesis
Platinum complexes 5a–5f were synthesized by the following method: K2PtCl4
(0.0482 mmol) was added to a CH3OH/H2O (4 mL, v/v = 1/1) solution of reduced
Schiff bases 4a–4f (0.0461 mmol) at room temperature, and the mixture was
adjusted to pH 8–9 at the start, then stirred for 24 h, when the pH was down to about
7. The solution was heated in vacuo and recrystallized from CH3OH/CH2Cl2 in the
N2 protection.
Synthesis of methyl 2-(2-hydroxybenzylamino)acetate (4a)
White solid. (34.6 %): 1H NMR (600 MHz, DMSO-d6): d 7.198–7.224 (t, 1H, Ar–H),
7.003–7.015 (d, 1H, Ar–H), 6.872–6.886 (d, 1H, Ar–H), 6.800–6.825(t, 1H, Ar–H),
4.003 (s, 2H, –CH2–), 3.789 (s, 2H, –CH3), 3.482 (s, 2H, –CH2–); IR (KBr): 3,352
(N–H), 3,003, 1,735 (C=O), 1,587, 1,256 (ph–O), 1,130, 987, 858, 756 cm-1. Anal.
Calc. for C10H13NO3: C, 61.53; H, 6.71; N, 7.18. Found: C, 61.12; H, 6.95; N, 7.31.
Synthesis of ethyl 2-(2-hydroxybenzylamino)acetate (4b)
1
White solid. (42.5 %): H NMR (600 MHz, DMSO-d6): d 7.062–7.100 (m, 2H,
Ar–H), 6.720–6.748 (m, 2H, Ar–H), 4.087–4.122 (m, 2H, –CH2–), 3.348 (s, 2H,
–CH2–), 3.789 (s, 2H, –CH2–), 1.186–1.210 (t, 3H, –CH3); IR (KBr): 3,332 (N–H),
2,984, 1,739 (C=O), 1,589, 1,256 (ph–O), 1,130, 1,029, 855, 757 cm-1. Anal. Calc.
for C11H15NO3: C, 63.14; H, 7.23; N, 6.69. Found: C, 63.45; H, 6.89; N, 6.91.
Synthesis of methyl 2-(5-bromo-2-hydroxybenzylamino)acetate (4c)
1
White solid. (33.4 %): H NMR (600 MHz, DMSO-d6): d 7.313–7.317 (d, 1H,
Ar–H), 7.210–7.228 (dd, 1H, Ar–H), 6.703–6.717 (d, 1H, Ar–H), 3.722(s, 2H,
–CH2–), 3.635 (s, 3H, –CH3), 2.509 (s, 2H, –CH2–); IR (KBr): 3,309 (N–H), 2,921,
1,742 (C=O), 1,577, 1,266 (ph–O), 1,110, 990, 868, 763 cm-1. Anal. Calc. for
C10H12BrNO3: C, 43.82; H, 4.41; N, 5.11. Found: C, 42.58; H, 4.18; N, 5.38.
Synthesis of ethyl 2-(5-bromo-2-hydroxybenzylamino)acetate (4d)
1
White solid. (47.7 %): H NMR (600 MHz, DMSO-d6): d 7.309–7.314 (d, 1H,
Ar–H), 7.209–7.227 (dd, 1H, Ar–H), 6.703–6.717 (d, 1H, Ar–H), 4.079–4.114 (q,
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Platinum(II) complexes of reduced amino acid ester Schiff-bases
2H, –CH2–), 3.722 (s, 2H, –CH2–), 2.505–2.511 (t, 2H, –CH2–), 1.184–1.208 (s, 3H,
–CH3); IR (KBr): 3,303 (N–H), 2,971, 1,737 (C=O), 1,577, 1,266 (ph–O), 1,143,
992, 873, 763 cm-1. Anal. Calc. for C11H14BrNO3: C, 45.85; H, 4.90; N, 4.86.
Found: C, 45.58; H, 5.31; N, 4.60.
Synthesis of methyl 2-(5-chloro-2-hydroxybenzylamino)acetate (4e)
White solid. (51.8 %): 1H NMR(600 MHz, DMSO-d6): d 7.191–7.195 (d, 1H,
Ar–H), 7.088–7.106 (dd, 1H, Ar–H), 6.748–6.762 (d, 1H, Ar–H), 3.729 (s, 2H,
–CH2–), 3.638 (s, 3H, –CH3), 3.369 (s, 2H, –CH2–); IR (KBr): 3,309 (N–H), 2,924,
1,741 (C=O), 1,580, 1,265 (ph–O), 1,145, 992, 873, 762 cm-1. Anal. Calc. for
C10H12ClNO3: C, 52.30; H, 5.27; N, 6.10. Found: C, 52.66; H, 5.61; N, 5.82.
Synthesis of ethyl 2-(5-chloro-2-hydroxybenzylamino)acetate (4f)
1
White solid. (51.5 %): H NMR (600 MHz, DMSO-d6): d 7.190–7.193 (d, 1H,
Ar–H), 7.089–7.107 (dd, 1H, Ar–H), 6.745–6.759 (d, 1H, Ar–H), 4.079–4.115 (q,
2H, –CH2–), 3.721 (s, 2H, –CH2–), 3.366 (s, 2H, –CH2–), 1.184–1.207 (s, 3H,
–CH3); IR (KBr): 3,303 (N–H), 2,973, 1,763 (C=O), 1,581, 1,264 (ph–O), 1,143,
983, 871, 765 cm-1. Anal. Calc. for C11H14ClNO3: C, 54.22; H, 5.79; N, 5.75.
Found: C, 54.49; H, 5.41; N, 5.49.
Synthesis of Pt[methyl 2-(2-hydroxybenzylamino)acetate]Cl (5a)
1
Yellow Solid. (63.46 %): H NMR (600 MHz, DMSO-d6): d 7.088–7.169 (t, 1H,
Ar–H), 6.993–7.005 (d, 1H, Ar–H), 6.812–6.876 (dd, 2H, Ar–H), 4.021 (s, 2H,
–CH2–), 3.769 (s, 3H, –CH3), 3.463 (s, 2H, –CH2–); IR (KBr): 3,420 (N–H), 3,170,
1,631 (C=O), 1,404, 1,270 (ph–O), 1,119, 757, 709, 619 (Pt-OAr), 520 (Pt–N), 412
(Pt–O=C) cm-1. ESI–MS: 446.9964 [M ? Na]?. Anal. Calc. for C10H11ClNO3Pt:
C, 28.35; H, 2.62; N, 3.31; Found: C, 28.78; H, 2.26; N, 3.55.
Km = 45 S cm2 mol-1
.
Synthesis of Pt[ethyl 2-(2-hydroxybenzylamino)acetate]Cl (5b)
1
Yellow Solid. (65.78 %): H NMR (600 MHz, DMSO-d6): d 6.972–7.011 (m, 2H,
Ar–H), 6.702–6.726 (m, 2H, Ar–H), 4.108–4.151 (m, 2H, –CH2–), 3.368 (s, 2H,
–CH2–), 3.769 (s, 2H, –CH2–), 1.178–1.208 (t, 3H, –CH3); IR (KBr): 3,442 (N–H),
2,929, 1,643 (C=O), 1,470, 1,265 (ph–O), 1,123, 823, 670, 644 (Pt-OAr), 519
(Pt–N), 429 (Pt–O=C) cm-1. ESI–MS: 461.0121 [M ? Na]?. Anal. Calc. for
C11H13ClNO3Pt: C, 30.18; H, 2.99; N, 3.20; Found: C, 30.43; H, 2.67; N, 2.91.
Km = 48 S cm2 mol-1
.
Synthesis of Pt[methyl 2-(5-bromo-2-hydroxybenzylamino)acetate]Cl (5c)
1
Yellow Solid. (66.41 %): H NMR (600 MHz, DMSO-d6): d 7.289–7.296 (d, 1H,
Ar–H), 7.206–7.216 (dd, 1H, Ar–H), 6.710-6.726 (d, 1H, Ar–H), 3.730(s, 2H,
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L.-J. Li et al.
–CH2–), 3.641 (s, 3H, –CH3), 2.498 (s, 2H, –CH2–); IR (KBr): 3,445 (N–H), 2,921,
1,645 (C=O), 1,471, 1,268 (ph–O), 1,121, 822, 664, 615 (Pt–OAr), 520 (Pt–N), 420
(Pt–O=C) cm-1. ESI–MS: 524.9054 [M ? Na]?. Anal. Calc. for C10H10BrClNO3
Pt: C, 23.90; H, 2.01; N, 2.79. Found: C, 23.68; H, 2.43; N, 2.51. Km = 57 S cm2
mol-1
.
Synthesis of Pt[ethyl 2-(5-bromo-2-hydroxybenzylamino)acetate]Cl (5d)
1
Yellow Solid. (70.32 %): H NMR (600 M Hz, DMSO-d6): d 7.294–7.310 (d, 1H,
Ar–H), 7.198–7.216 (dd, 1H, Ar–H), 6.696–6.711 (d, 1H, Ar–H), 4.082–4.118 (m,
2H, –CH2–), 3.728 (s, 2H, –CH2–), 2.498–2.509 (t, 2H, –CH2–), 1.188–1.213 (s, 3H,
–CH3); IR (KBr): 3,426 (N–H), 2,927, 1,643 (C=O), 1,481, 1,268 (ph–O), 1,113,
881, 636, 615 (Pt–OAr), 519 (Pt–N), 429 (Pt–O=C) cm-1. ESI–MS: 538.9211
[M ? Na]?. Anal. Calc. for C11H12BrClNO3Pt: C, 25.57; H, 2.34; N, 2.71. Found:
C, 25.88; H, 2.61; N, 2.35. Km = 60 S cm2 mol-1
.
Synthesis of Pt[methyl 2-(5-chloro-2-hydroxybenzylamino)acetate]Cl (5e)
1
Yellow Solid. (68.44 %): H NMR (600 MHz, DMSO-d6): d 7.185–7.190 (d, 1H,
Ar–H), 7.080–7.098 (dd, 1H, Ar–H), 6.741–6.756 (d, 1H, Ar–H), 3.732 (s, 2H,
–CH2–), 3.644 (s, 3H, –CH3), 3.371 (s, 2H, –CH2–); IR (KBr): 3,440 (N–H), 2,926,
1,648 (C=O), 1,473, 1,264 (ph–O), 1,121, 824, 680, 618 (Pt–OAr), 524 (Pt–N), 420
(Pt-O=C) cm-1. ESI–MS: 480.9565 [M ? Na]?. Anal. Calc. for C10H10Cl2NO3Pt:
C, 26.21; H, 2.20; N, 3.06. Found: C, 26.62; H, 2.44; N, 2.76.
Km = 55 S cm2 mol-1
.
Synthesis of Pt[ethyl 2-(5-chloro-2-hydroxybenzylamino)acetate]Cl (5f)
1
Yellow Solid. (60.27 %): H NMR (600 MHz, DMSO-d6): d 7.182–7.186 (d, 1H,
Ar–H), 7.081–7.098 (dd, 1H, Ar–H), 6.738–6.750 (d, 1H, Ar–H), 4.083–4.119 (q,
2H, –CH2–), 3.728 (s, 2H, –CH2–), 3.374(s, 2H, –CH2–), 1.186–1.209 (s, 3H,
–CH3); IR (KBr): 3,423 (N–H), 2,912, 1,601 (C=O), 1,473, 1,263 (ph–O), 1,119,
825, 681, 611 (Pt–OAr), 519 (Pt–N), 429 (Pt–O=C) cm-1. ESI–MS: 494.9722
[M ? Na]?. Anal. Calc. for C11H12Cl2NO3Pt: C, 27.98; H, 2.56; N, 2.97. Found: C,
27.72; H, 2.85; N, 2.66. Km = 58 S cm2 mol-1
.
Determination of UV absorption spectra
An UV-3400 Toshniwal spectrophotometer emitting UV light was used mainly
between 200 and 400 nm. The salmon sperm DNA (M = 208.8 g/mol) was
dissolved in Tris–HCl (pH 7.5) buffer solution, and rested in 24 h at 4 °C. Then
complexes 5c and 5d were added in buffer solution in different concentrations
(ccomplex:cDNA = 0.1, 0.3 and 0.5), and rested for 24 h at 4 °C. The UV absorption
spectra was determined at room temperature (Dt = 1 s, n = 3).
123

Platinum(II) complexes of reduced amino acid ester Schiff-bases
Cell culture
Four different human carcinoma cell lines: HL-60, Bel-7402, BGC-823, and KB,
were cultured in RPMI-1640 medium supplemented with 10 % fetal bovine serum,
100 units/mL of penicillin and 100 lg/mL of streptomycin. Cells were maintained
at 37 °C in a humidified atmosphere of 5 % CO2 in air.
Solutions
The complexes were dissolved in DMSO at a concentration of 5 mM as stock
solution, and diluted in culture medium at concentrations of 0.1, 1.0, and 10 lM as
working solution. To avoid DMSO toxicity, the concentration of DMSO was less
than 0.1 % (v/v) in all experiments.
Cytotoxicity analysis
The cells harvested from the exponential phase were seeded equivalently into a
96-well plate, and then the complexes were added to the wells to achieve final
concentrations. Control wells were prepared by addition of culture medium. Wells
containing culture medium without cells were used as blanks. All experiments were
performed in quintuplicate. The MTT assay was performed as described by
Mosmann for HL-60 [31]. Upon completion of the incubation for 44 h, stock MTT
dye solution (20 mL, 5 mg/mL) was added to each well. After 4 h incubation,
2-propanol (100 mL) was added to solubilize the MTT formazan. The OD of each
well was measured on a microplate spectrophotometer at a wavelength of 570 nm.
The IC50 value was determined from a plot of % viability against dose of
compounds added.
Acknowledgments This work was supported by the Natural Science Foundation of Hebei Province
(No. B2012201109, No. B2010000223) and the Doctoral foundation of Hebei University (No. 2009-164).
References
1. M. Shaharyar, M.M. Abdullah, M.A. Bakht, J. Majeed, Eur. J. Med. Chem. 45, 114 (2010)
2. H. Khan, A. Badshah, G. Murtaz, M. Said, Z.R. Neuhausen, M. Todorova, Eur. J. Med. Chem. 46,
4071 (2011)
3. E. Wong, C.M. Giandomenico, Chem. Rev. 99, 2451 (1999)
4. E.R. Jamieson, S.J. Lippard, Chem. Rev. 99, 2467 (1999)
5. T.W. Hambley, Coord. Chem. Rev. 166, 181 (1997)
6. Y.P. Ho, K.K.W. To, X. Wang, G. Lin, X. Han, J. Med. Chem. 44, 2065 (2001)
7. Y.S. Kim, R. Song, D.H. Kim, M.J. Jun, Y.S. Sohn, Bioorg. Med. Chem. 11, 1753 (2003)
8. S.D. Adhikary, D. Bose, P. Mitra, K.D. Saha, V. Bertolasi, J. Dinda, New J. Chem. 36, 759 (2012)
9. Y.P. Ho, C.F. Steve, A. Yeung, K.K.W. To, Med. Res. Rev. 23, 633 (2003)
10. P.U. Maheswari, S. Roy, S. Barends, B. Kozlevcar, P. Gamez, J. Reedijk, J. Am. Chem. Soc. 128,
710 (2006)
11. J. Qian, W. Gu, H. Liu, F.X. Gao, L. Feng, S.P. Yan, D.Z. Liao, P. Cheng, Dalton Trans. 11, 1060
(2007)
12. J.L. Tian, L. Feng, W. Gu, G.J. Xu, S.P. Yan, D.Z. Liao, Z.H. Jiang, P. Cheng, Inorg. Biochem. 101,
96 (2007)
123

L.-J. Li et al.
13. J.H. Li, J.T. Wang, Z.W. Mao, L.N. Ji, J. Coord. Chem. 62, 446 (2009)
14. T.D. Tullius, J.A. Greenbaum, Curr. Opin. Chem. Biol. 9, 127 (2005)
15. K. Tatsumoto, A.E. Martell, R.J. Motekaitis, J. Am. Chem. Soc. 103, 6197 (1981)
16. K. Korhonen, R. Hamalainen, U. Turpeinen, Acta Crystallogr. Sect. C 40, 1175 (1984)
17. V. Kettmann, E. Fresova, Acta Crystallogr. Sect. C 49, 1932 (1993)
18. C.Y. Gao, X.F. Ma, J.L. Tian, D.D. Li, S.H. Yan, J. Coord. Chem. 63, 115 (2010)
19. B. Sreenivasulu, M. Vetrichelvan, F. Zhao, S. Gao, J.J. Vittal, Eur. J. Inorg. Chem. 4635 (2005)
20. R. Ganguly, B. Sreenivasulu, J.J. Vittal, Coord. Chem. Rev. 252, 1027 (2008)
21. X.F. Ma, D.D. Li, J.L. Tian, Y.Y. Kou, S.H. Yan, Trans. Metal. Chem. 34, 475 (2009)
22. C. Gao, X. Ma, J. Tian, D. Li, S. Yan, J. Coord. Chem. 63, 115 (2010)
23. W.J. Geary, Coord. Chem. Rev. 7, 81 (1971)
24. A. Skladanowski, P. Bozko, M. Sabisz, Chem. Rev. 109, 2951 (2009)
25. M. Wang, L.R. Hou, Z.N. Gao, T.L. Yang, Chin. J. Appl. Chem. 25, 1186 (2008)
26. L.N. Ji, Q.L. Zhang, J.G. Liu, Sci. China (Ser. B). 31, 193 (2001)
27. P.R. Cao, M.M. McHugh, T. Melendy, T. Beerman, Mol. Cancer Ther. 2, 651 (2003)
28. L. Xie, J. Cui, X. Qian, Y. Xu, J. Liu, R. Xu, Bioorg. Med. Chem. 19, 961 (2011)
29. C. Elzbieta, E. Zygmunt, K. Romuald, Dissertation. Pharm. 15, 369 (1964)
30. J. Li, Y. Sha, Molecules 13, 1111 (2008)
31. T. Mosmann, J. Immunol. Methods 65, 55 (1983)
123