Synthesis, crystal structure, and SOD-like activity of two copper(II) complexes with hydroxymethyl pendants

Article abstract of DOI:10.1080/00958972.2014.940335

Dinuclear and mononuclear copper(II) complexes (1, [Cu₂(HL)₂(L)₂](ClO₄)₂; 2, [Cu(HL)₂(phen)] (ClO₄)₂, where HL = (N-methyl-2-methylol)imidazole, phen = 1,10-phenanthroline) have been synthesized and characterized by single crystal X-ray diffraction. The SOD-like activity (IC₅₀) of 1 and 2 were measured to be 0.10 ± 0.01 and 0.19 ± 0.01 μM by NBT assay at pH 7.8. The higher SOD activity of 1 could be contributed to the coordination configuration and the labile hydroxymethyl pendants. Electrochemistry and the frontier molecular orbital energies of the complexes were also studied. To mimic the active site of Cu, Zn-SOD, new copper(II) complexes (1, [Cu₂(HL)₂(L)₂](ClO₄)₂; 2, [Cu(HL)₂(phen)](ClO₄)₂; where HL = (N-methyl-2-methylol)imidazole, phen = 1,10-phenanthroline) have been synthesized and characterized by elemental analysis, IR, and single-crystal X-ray diffraction. Copper(II) in 1 is four-coordinate by a N₂O₂ plane with two copper(II) ions bridged with two oxygens from the deprotonated hydroxylmethyl pendants. Each Cu²⁺ in 2 is coordinated by four nitrogens from two HL and one co-ligand of 1,10-phenanthroline. Electrochemistry of the complexes was studied by cyclic voltammetry. The atomic net charges distribution and frontier molecular orbital energies were obtained by Gaussian 98 program with DFT method at B3LYP/lanl2dz level. The SOD-like activity (IC₅₀) of 1 and 2 were measured as 0.10 ± 0.01 and 0.19 ± 0.01 μM by NBT assay at pH 7.8. The higher SOD activity of 1 could be attributed to the coordination configuration and the labile hydroxymethyl pendants.

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Synthesis, crystal structure, and
SOD-like activity of two copper(II)
complexes with hydroxymethyl
pendants
ab
a
a
a
Ying-Hua Zhou , Da-Liang Sun , Jun Tao , Li-Qing Chen , Ya-Fan
a
a
ac
Huang , Ying-Kun Li & Yong Cheng
a
College of Chemistry and Materials Science, Anhui Normal
University, Wuhu, PR China
b
Center for Nano Science and Technology, Anhui Normal
University, Wuhu, PR China
Click for updates
c
The Key Laboratory of Functional Molecular Solids, Ministry of
Education, Anhui Normal University, Wuhu, PR China
Accepted author version posted online: 30 Jun 2014.Published
online: 22 Jul 2014.
To cite this article: Ying-Hua Zhou, Da-Liang Sun, Jun Tao, Li-Qing Chen, Ya-Fan Huang, Ying-
Kun Li & Yong Cheng (2014) Synthesis, crystal structure, and SOD-like activity of two copper(II)
complexes with hydroxymethyl pendants, Journal of Coordination Chemistry, 67:14, 2393-2404,
DOI: 10.1080/00958972.2014.940335
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Journal of Coordination Chemistry, 2014
Vol. 67, No. 14, 2393–2404, http://dx.doi.org/10.1080/00958972.2014.940335
Synthesis, crystal structure, and SOD-like activity of two
copper(II) complexes with hydroxymethyl pendants
YING-HUA ZHOU*†‡, DA-LIANG SUN†, JUN TAO†, LI-QING CHEN†,
YA-FAN HUANG†, YING-KUN LI† and YONG CHENG†§
†College of Chemistry and Materials Science, Anhui Normal University, Wuhu, PR China
‡Center for Nano Science and Technology, Anhui Normal University, Wuhu, PR China
§The Key Laboratory of Functional Molecular Solids, Ministry of Education, Anhui Normal
University, Wuhu, PR China
(
Received 8 March 2014; accepted 6 June 2014)
Dinuclear and mononuclear copper(II) complexes (1, [Cu
ClO , where HL = (N-methyl-2-methylol)imidazole, phen = 1,10-phenanthroline) have been syn-
thesized and characterized by single crystal X-ray diffraction. The SOD-like activity (IC50) of 1 and
were measured to be 0.10 ± 0.01 and 0.19 ± 0.01 μM by NBT assay at pH 7.8. The higher SOD
2 2 2 4 2 2
(HL) (L) ](ClO ) ; 2, [Cu(HL) (phen)]
(
4 2
)
2
activity of 1 could be contributed to the coordination configuration and the labile hydroxymethyl
pendants. Electrochemistry and the frontier molecular orbital energies of the complexes were also
studied.
To mimic the active site of Cu,Zn-SOD, new copper(II) complexes (1, [Cu
Cu(HL) (phen)](ClO ; where HL = (N-methyl-2-methylol)imidazole, phen = 1,10-phenanthroline)
have been synthesized and characterized by elemental analysis, IR, and single-crystal X-ray diffrac-
tion. Copper(II) in 1 is four-coordinate by a N plane with two copper(II) ions bridged with two
2 2 2 4 2
(HL) (L) ](ClO ) ; 2,
[
2
4 2
)
2 2
O
2+
oxygens from the deprotonated hydroxylmethyl pendants. Each Cu in 2 is coordinated by four
nitrogens from two HL and one co-ligand of 1,10-phenanthroline. Electrochemistry of the com-
plexes was studied by cyclic voltammetry. The atomic net charges distribution and frontier molecu-
lar orbital energies were obtained by Gaussian 98 program with DFT method at B3LYP/lanl2dz
level. The SOD-like activity (IC50) of 1 and 2 were measured as 0.10 ± 0.01 and 0.19 ± 0.01 μM by
NBT assay at pH 7.8. The higher SOD activity of 1 could be attributed to the coordination
configuration and the labile hydroxymethyl pendants.
Keywords: Copper complex; Crystal structure; Superoxide dismutase; Imidazole; Mimic
*
Corresponding author. Email: yhzhou@mail.ahnu.edu.cn
2014 Taylor & Francis
©

2
394
. Introduction
Y.-H. Zhou et al.
1
ÁÀ
Superoxide anion (O ) is a natural byproduct of the normal metabolism of oxygen and has
2
the important functions of cell signaling and homeostasis. However, during environmental
ÁÀ
stress, the level of O2 can increase dramatically, inducing host molecular and cellular dam-
ages to lipids and proteins [1–3]. Copper, zinc superoxide dismutase (Cu,Zn-SOD) is an
ubiquitous enzyme with an essential role in antioxidant defense through catalyzing the dis-
ÁÀ
mutation of O2 to hydrogen peroxide and dioxygen under physiological conditions [4, 5].
Due to limitations such as solution instability, immunogenicity, bell-shaped dose response
curves, short half-lives, costs of production and proteolytic digestion, the direct utilization
of such enzyme as pharmaceutical agents still has many problems [6]. Therefore, research
of Cu,Zn-SOD mimic has been carried out towards obtaining stable, nontoxic, and inexpen-
sive low molecular weight biomimetic molecules, which are able to catalyze superoxide dis-
mutation and provide important therapeutic applications [7]. To mimic Cu,Zn-SOD, many
copper(II) complexes with polyamine and porphyrin derivative ligands were reported. How-
ever, the SOD-like activities of mimics were relatively low and the water solubility of mim-
ics should be improved [8–11]. Imidazole compounds widely exist in metalloenzymes of
mammals due to coordination diversity and favorable hydrophilicity [12–14]. The flexible
hydroxymethyl arm can be expected to adapt to the different requirements of configuration
and proper basicity for copper ions. Furthermore, oxygen of the hydroxymethyl arm may
mimic the role of water molecule bound to the copper site of Cu ,Zn -SOD [15]. Metal
2
2
hydroxides and alkoxides play important roles on activation of metal proximate alcoholic
OH groups in carbonic anhydrase and carboxypeptidase [16, 17]. Therefore, we herein syn-
thesize two di/mono-copper(II) complexes based on the imidazole derivative containing
hydroxymethyl pendant groups (1, [Cu (HL) (L) ](ClO ) ; 2, [Cu(HL) (phen)](ClO ) ;
2
2
2
4 2
2
4 2
where HL = (N-methyl-2-methylol)imidazole, phen = 1,10-phenanthroline), which have been
characterized by X-ray crystallography. The atomic net charge distribution and frontier
molecular orbital energies were obtained by Gaussian 98 with the DFT method at B3LYP/
lanl2dz level. Moreover, the electrochemistry and SOD activity of 1 and 2 were
determined.
2
. Experimental
Caution: Although no problems were encountered in this work, transition-metal perchlorate
salts are potentially explosive and should thus be prepared in small quantities and handled
with care.
2.1. Materials and instrumentation
Organic reagents were reagent grade and solvents used in this research were purified by
standard procedures. Water used in all physical measurement experiments was double dis-
tilled. An improved method of solvothermal synthesis has been applied for the preparation
of (N-methyl-2-methylol)imidazole from N-methyl-1H-imidazole with a higher yield, and
1
was confirmed by elemental analyses and H NMR spectra [18, 19]. Then, the mono- or di-
nuclear complexes were synthesized by reaction of ligand and copper(II) perchlorate. Ele-
mental analyses for C, H, and N were obtained on a Vario EL III. Infrared spectroscopy on

Copper(II) complexes with hydroxymethyl pendants
2395
KBr pellets was performed on a Shimadzu IR Prestige-21 infrared spectrophotometer from
−
1
4
000–400 cm . Electronic spectra were measured on a Shimadzu UV-2450 spectrophotom-
eter. The redox potentials of the complexes were determined by CH Instrument 400c using
the conventional three-electrode system of glass carbon electrode as the working electrode,
platinum wire as the counter electrode, and Ag/AgCl electrode as the reference electrode.
All electrospray ionization mass spectrometry measurements were carried out using a Finni-
gan LCQ DECA XP PLUS ion trap spectrometer operating in the positive ion mode and
equipped with an orthogonal ESI source (Thermo Electron Corporation, USA).
2
.2. Synthesis of (N-methyl-1H-imidazol-2-yl)methanol (HL)
To a solution of N-methyl-1H-imidazole (15.00 g, 0.18 M) in 20 mL anhydrous acetonitrile
in a 100 mL polytetrafluoroethylene flask was added a solution of paraformaldehyde (30.00
g, 1.00 M) in 20 mL anhydrous acetonitrile with vigorous stirring under nitrogen. After
placing in the sealed high pressure reactor and heating to 393 K for 16 h, the reaction was
cooled to room temperature and evaporated under reduced pressure to dryness. The residue
was poured into acetone (100 mL) to give a precipitate. The crude product was recrystal-
lized from a solution of ethyl acetate and anhydrous methanol (v/v, 2 : 1). Colorless cubic
crystals suitable for X-ray analysis were obtained in 92% yield. Anal. Calcd for C H N O:
5
8 2
1
C, 53.56; N, 24.98; H, 7.19%. Found: C, 53.36; N, 24.88; H, 7.31%. H NMR (300 MHz,
CDCl ): δ 6.86 (2H; CH-4,5), 4.63 (2H; CH ), 3.73 (3H, CH ); C NMR (500 MHz,
1
3
3
H
2
3
D O): δ 32.74 (–CH ), 55.08 (CH OH), 123.28 (CH-5), 126.32 (CH-4), 146.92 (CH-2).
2
C
3
2
2
.3. Synthesis of [Cu (HL) (L) ](ClO ) (1)
2 2 2 4 2
To 10 mL methanol solution of HL (0.224 g, 2.0 mM) was slowly added a solution of Cu
ClO ) ·6H O (0.370 g, 1.0 mM) in 30 mL ethanol with stirring. The pH of the solution
(
4
2
2
was adjusted to about 8 with triethylamine. After reluxing for 8 h, the mixture was cooled
to room temperature and filtered. The filtrate was kept at room temperature. Three days
later, blue cubic crystals suitable for X-ray analysis were obtained with 62% yield. Anal.
Calcd for C H Cl N O Cu : C, 31.10; N, 14.51; H, 3.91%. Found: C, 30.92; N, 14.16;
2
0
30
2
8
12
2
−
1
H, 4.23%. IR (KBr disk, cm ): 3439 (s, br), 3138 (m), 2375 (s), 2343 (s), 2301 (s), 1634
s), 1597 (s), 1508 (m), 1342 (m), 1122 (s), 1073 (s), 752 (m), 667 (s), 626 (s), 416 (s).
(
2
.4. Synthesis of [Cu(HL) (phen)](ClO ) (2)
2 4 2
To 10 mL methanol solution of HL (0.224 g, 2.0 mM) was slowly added a solution of Cu
(
ClO ) ·6H O (0.370 g, 1.0 mM) in 30 mL ethanol with stirring. After stirring for 30 min,
4 2 2
1
,10-phenanthroline (0.180 g, 1.0 mM) was added. Then, after reluxing for 8 h, the mixture
was cooled to room temperature and filtered. The filtrate was kept at room temperature.
Three days later, blue cubic crystals suitable for X-ray analysis were obtained with 68%
yield. Anal. Calcd for C H Cl N O Cu: C, 39.62; N, 12.60; H, 3.63%. Found: C, 39.46;
2
2
24
2 6 10
−
1
N, 12.51; H, 3.91%. IR (KBr disk, cm ): 3407 (s, br), 3131 (m), 2367 (s), 1621 (s), 1524
s), 1424 (s), 1342 (s), 1277 (m), 1222 (m), 1107 (s), 1068 (s), 926 (m), 847 (s), 766 (s),
20 (s), 672 (s), 620 (s).
(
7

2
396
.5. X-ray crystallography
Y.-H. Zhou et al.
2
Single crystals of 1 and 2 were used for X-ray diffraction analyses by mounting on the tip
of a glass fiber in air using a Bruker Smart Apex CCD diffractometer at 291 K with graph-
ite-monochromated Mo Kα radiation (λ = 0.71073 Å). Diffraction intensities for the com-
plexes were collected using the ω-scan technique. The structures were solved by direct
methods using SHELXS-97 [20] and subsequent Fourier difference techniques, and refined
2
anisotropically by full-matrix least-squares on F using SHELXL-97 [21]. All nonhydrogen
atoms were refined anisotropically and all hydrogens were located in the Fourier difference
maps. Further crystallographic data and experimental details for structural analyses of 1 and
2
are summarized in table 1. Selected bond distances and angles are listed in table 2.
2
.6. DFT calculation
The atomic coordinate positions of 1 and 2 were based on the data of the crystal structures,
À
in which the solvent molecules and ClO4 were ignored. DFT calculations with a hybrid
Functional B3LYP method at the LanL2DZ basis set were performed with Gaussian 98 soft-
ware package [22]. In 1, the charge and multiplicity is respectively set as +2, 3 with
restricted-open spin due to two unpaired electrons in two copper(II), while +2, 2 in 2
because of one unpaired electron in Cu(II). All calculations were performed on a Pentium
IV computer using the high convergence criteria. For 1, 64 atoms, 392 basis functions,
1
024 primitive gaussians, 138 alpha electrons, and 136 beta electrons are involved in the
Table 1. Crystal data, data collection, and structure refinement parameters for HL, 1, and 2.
Compound
HL
1
2
Empirical formula
Formula weight
Temperature (K)
λ/(Å)
Crystal system
Space group
a (Å)
b (Å)
c (Å)
α (°)
β (°)
C
5
H
8
N
2
O
C
20
H
30Cl
2
Cu
2
N
8
O
12
C
22 2 6 11
H24Cl CuN O
112.13
293
0.71073
Orthorhombic
Pbca
12.3358(12)
7.0839(7)
13.1696(13)
90.00
90.00
90.00
1150.8(2), 8
1.294
0.093
772.50
293
0.71073
Monoclinic
P2 /n
7.3523(4)
10.7269(6)
18.9324(11)
90.00
91.2180(10)
90.00
1492.81(15), 1
1.719
1.676
788
682.91
293
0.71073
Triclinic
P − 1
8.0008(12)
12.0485(18)
15.977(2)
83.140(2)
76.398(2)
74.060(2)
1436.9(4), 2
1.578
1
γ (°)
Volume (Å ), Z
3
−
3
Calculated density (Mg m
Absorption coefficient (mm
F(0 0 0)
)
1
−
)
1.013
698
480
θ Range for data collection (°)
Limiting indices
3.09–27.44
−12 ≤ h ≤ 15
2.15–27.67
−9 ≤ h ≤ 9
−13 ≤ k ≤ 13
−24 ≤ l ≤ 23
3463/0/205
12,761
1.76–25.35
−9 ≤ h ≤ 9
−12 ≤ k ≤ 14
−19 ≤ l ≤ 19
5211/0/392
10,405
−9 ≤ k ≤ 9
−17 ≤ l ≤ 16
Data/restraints/parameters
Total reflections
1310/0/76
9019
Unique reflections (Rint
)
0.0268
0.0259
0.0203
Completeness
Goodness-of-fit on F
99.8%
1.019
99.3%
1.044
98.7%
1.055
2
R
R
1
/wR
/wR
2
[I > 2σ(I)]
0.0395/0.1102
0.0431/0.1144
0.239/−0.143
0.0306/0.0815
0.0389/0.0865
0.420/−0.364
0.0565/0.1652
0.0651/0.1727
0.625/−0.586
1
2
[all data]
−
3
Δρmax, min (e Å
)

Copper(II) complexes with hydroxymethyl pendants
2397
Table 2. Selected bond lengths (Å) and angles (°) for 1 and 2.
1
Cu(1)–O(1)
Cu(1)–O(1A)
O(1)–Cu(1)–O(1A)
O(1)–Cu(1)–N2
O(1A)–Cu(1)–N2
1.9350(15)
1.9378(15)
78.11(7)
176.76(7)
99.73(7)
Cu(1)–N(2)
Cu(1)–N(3)
O(1)–Cu(1)–N3
O(1A)–Cu(1)–N3
N2–Cu(1)–N3
1.9523(17)
1.9632(19)
82.61(7)
150.69(8)
100.29(8)
2
Cu(1)–N(1)
1.977(4)
Cu(1)–N(6)
2.028(4)
Cu(1)–N(3)
1.981(4)
Cu(1)–N(5)
2.031(4)
N(1)–Cu(1)–N(3)
N(1)–Cu(1)–N(6)
N(3)–Cu(1)–N(6)
90.95(15)
172.07(14)
93.86(15)
N(1)–Cu(1)–N(5)
N(3)–Cu(1)–N(5)
N(6)–Cu(1)–N(5)
94.55(15)
169.92(15)
81.67(15)
Note: Symmetry transformations used to generate equivalent atoms for 1: A, −x + 1, −y + 2, −z.
calculation. For 2, 55 atoms, 340 basis functions, 898 primitive gaussians, 116 alpha elec-
trons, and 116 beta electrons are involved in the calculation.
2
.7. Cyclic voltammetry
A CHI400c computerace instrument was employed to obtain cyclic voltammograms in
dimethylformamide solutions of the complex (1 mM) at room temperature (298 K) under
nitrogen using 0.1 M tetrabutylammonium perchlorate (TBAP) as supporting electrolyte. A
glass carbon working electrode, a platinum auxiliary electrode, and a Ag/AgCl reference
electrode were used to obtain cyclic voltammograms. Measured potentials were calibrated
+
through an internal ferrocene (Fc /Fc) standard.
2
.8. SOD-like activity
ÁÀ
Superoxide anion (O ) was generated in an enzymatic (xanthine/xanthine oxidase) system
2
ÁÀ
in the presence or absence of test complex, and O2 production was determined by monitor-
ing the reduction of NBT to monoformazan dye at 560 nm at 298 ± 0.1 K [23–26]. An
appropriate amount of xanthine oxidase was added to a mixture of 500 μM NBT, 500 μM
xanthine, and 0–1.0 μM complex dissolved in 50 mM phosphate buffer (pH 7.8) to cause a
variation of absorbance (ΔA560/Δtmin) of 0.025 ± 0.005. The percentage inhibition (% I) of
+
NBT formation was calculated from equation: (%) I = (A − A )/A × 100, in which A and
0
s
0
0
+
A are the maximum absorbance values due to NBT at 560 nm in the absence and in the
s
presence of the complex. By plotting the % I as a function of complex concentration, the
IC50 values were calculated.
3
3
. Results and discussion
.1. UV–vis, IR spectroscopy of 1 and 2
The electronic absorption spectra of 1 and 2 in freshly prepared methanol solution were
obtained from 200 to 1100 nm at room temperature. As shown in figure S1 (see online
supplemental material at http://dx.doi.org/10.1080/00958972.2014.940335), the electronic
spectra of free HL displayed intense absorption at 221 nm due to an n→π* transition, which
was shifted to 216 and 232 nm for 1 and 2, respectively. The broad band at 303 nm in 1

2
398
Y.-H. Zhou et al.
was assigned to the ligand-to-metal charge transfer (LMCT) transition. The band at 295 nm
in 2 was assigned to LMCT transition. The low energy bands at 500–1000 nm corresponded
to d–d transitions. The broad band at 811 nm increased and shifted to 725 and 642 nm for 1
and 2, respectively, indicating that copper(II) binds with the ligand. As shown in figure S2,
−
1
−1
IR spectra show absorptions at 3440 cm for 1 and 3410 cm for 2, which can be attrib-
−
1
−1
uted to the stretching vibrations of O–H. Absorptions at 3148 cm for 1 and 3125 cm for
originate from stretching vibrations of imidazole–H. The peak at 1634 cm was assigned
to the C=C stretching frequency. The bands around 1508 and 1342 cm were attributed to
v(C=N) and v(C–N), respectively. Most important, the strong bands at 1122 and 626 cm
for 1 (2 at 1096 and 615 cm ) can be ascribed to vibration of perchlorate in the copper(II)
complexes [27–29].
−
1
2
−1
−
1
−
1
3
.2. Crystal structure analysis
Ligands 1 and 2 have been structurally characterized by X-ray crystallography (figure 1).
The single-crystal X-ray analysis reveals that [Cu (HL) (L) ](ClO ) (1) crystallizes in the
2
2
2
4 2
monoclinic space group P2 /n and exhibits a dinuclear structure, which contains two Cu(II)
1
−
centers, two L anions, two [HL], and two non-coordinated perchlorates. Selected bond
lengths and angles are listed in table 2. Each copper ion is at the center of the N O donor
2
2
set derived from three ligands in a square-planar geometry with Cu1 deviation 0.2320(2) Å
out from the plane. The two bridged oxygens and two copper(II) ions are practically copla-
nar with deviations from the least-squares plane lower than 0.002(1) Å. The dihedral angle
between the N O plane and Cu O plane is 23.2(1)°. The bond length of Cu1–O1(1.938
2
2
2 2
(2) Å) is typical for Cu–O complexes, while those of Cu–N (Cu(1)–N(2), 1.952(2) Å; Cu
(1)–N(3), 1.963(2) Å) are also in the normal range [30–32]. The distance of the Cu(1)Á Á ÁO
(2) (2.430(2) Å) is obviously longer than that of Cu(1)–O(1), due to O(2) from the alcohol
hydroxyl whereas O1 is from the deprotonated alcohol hydroxyl. Therefore, Cu(1) is four-
coordinate by O(1), O(1A), N(2), and N(3). The conformation of the dinuclear copper(II)
ions and four ligands is centrosymmetric, resulting in uniform coordination for each copper
(
II). The CuÁ Á ÁCu distance in each binuclear complex is 3.007 Å, close to that commonly
observed in dinuclear copper(II) complex based on the Robson-type macrocycle ligand
33]. [Cu(HL) (phen)](ClO ) (2) crystallizes in the monoclinic space group P − 1. Its
[
2
4 2
2
+
asymmetric unit contains one Cu , two HL, one 1,10-phenanthroline, and two uncoordi-
2
+
nated perchlorates. As shown in figure 1(3), each Cu is coordinated by two nitrogens
from 1,10-phenanthroline and two nitrogens from two HL. The coordination environment
of Cu(II) can be regarded as a slightly distorted planar geometry.
3
.3. DFT calculation
By analysis of Mulliken population, some information of atom net charges of 1 and 2 were
obtained (table 3). The net charge of Cu(II) is less than the original value (+2). In 1, the net
negative charge of O(1), O(1A), N(2), and N(3) are higher than that of O(2), N(1), and N
(4), suggesting that copper ion was preferentially coordinated by two oxygens from depro-
tonated hydroxyl and two nitrogens from two imidazole groups. For 2, the net negative
charges of O(1), O(2), N(2), and N(4) are obviously lower than that of other N atoms, indi-
cating that the above two hydroxyl oxygens from two ligands did not take part in complex-
ation. The frontier molecular orbital energies of 1 and 2 are listed in table 4, where the

Copper(II) complexes with hydroxymethyl pendants
2399
Figure 1. (1) ORTEP view of (N-methyl-2-methylol)imidazole (HL). Thermal ellipsoids are drawn at 30% proba-
bility level. (2) ORTEP view of [Cu (HL) (L) ](ClO . Thermal ellipsoids are drawn at 30% probability level. The
A” labeled atoms are at equivalent positions (−x + 1, −y + 2, −z). (3) ORTEP view of [Cu(HL) (phen)](ClO
Thermal ellipsoids are drawn at 30% probability level.
2
2
2
4 2
)
“
2
4 2
) .
occupied molecular orbital energies are all negative, indicating the complex is stable. Com-
pared with that of 2, the energy difference (ΔE = ELUMO − EHOMO) of 1 is lower, indicating
ÁÀ
that copper ion of 1 has a stronger potential of accepting lone electron pair of O2 as an
active center of the antioxidative reagent [34].
3
.4. Cyclic voltammetry
The electrochemical properties of 1 and 2 have been studied by cyclic voltammetry (CV) in
degassed DMF solution under nitrogen using 0.1 M TBAP as supporting electrolyte. A
glass carbon working electrode, a platinum auxiliary electrode, and a Ag/AgCl reference

2
400
Y.-H. Zhou et al.
Table 3. Mulliken atomic charges of some atoms of 1 and
2
at the B3LYP/LanL2DZ level.
1
2
Atom
Charge
Atom
Charge
Cu(1)
N(1)
N(2)
N(3)
N(4)
O(1)
O(2)
1.3057
−0.3944
−0.6026
−0.5900
−0.3800
−0.9391
−0.2154
Cu(1)
N(1)
N(2)
N(3)
N(4)
N(5)
N(6)
O(1)
O(2)
1.3475
−0.5652
−0.3544
−0.5792
−0.3331
−0.5250
−0.5053
−0.2485
−0.2460
Table 4. Frontier molecular orbital energies (eV)
of 1 and 2 at the B3LYP/LanL2DZ level.
Energies (eV)
1
2
HOMO
LUMO
ΔE
−6.3342
−4.7723
1.5619
−7.1915
−4.3922
2.7993
+
electrode were used to obtain cyclic voltammograms. Under these conditions, the Fc /Fc
standard couple occurred at 544 mV, identical to that reported previously [35]. The data for
1
and 2 are collected in table 5 and the cyclic voltammograms of the complexes are shown
in figure 2. Complex 1 showed the quasi-reversible redox waves with E = 0.28 V versus
1
/2
+
Fc /Fc, ΔE = 0.12 V, and i /i = 1.06, which was typical of reversible Cu(II)/Cu(I) process
p
pa pc
for the dinuclear copper(II) complex. The CV of 2 exhibits one reversible one-electron
+
redox couple at E1/2 = 0.13 V versus Fc /Fc, ΔE = 0.09 V. It should be pointed out that only
p
one pair of redox peaks was detected in the dinuclear 1, indicating that the environments of
the two copper ions are very similar [34, 36]. It is in agreement with the centrosymmetric
conformation of the dicopper complex from crystal structure analysis. The potentials of 1
can be assigned to one-electron process for each Cu(II)/Cu(I) couple and two-electron pro-
cess for the dinuclear copper(II) complex. The difference in Cu(II)/Cu(I) redox potential
between 1 and 2 demonstrates that the coordination environments around the copper(II)
ions of the two complexes are different. An adequate Cu(II)/Cu(I) redox potential for effec-
tive catalysis of superoxide radical must be required between −0.36 V versus Ag/AgCl for
À
À
O /O and 0.69 V versus Ag/AgCl for O /H O [15]. Therefore, 1 and 2 can be applied to
2
2
2
2 2
mimic superoxide dismutase, because the redox potentials of the two complexes in DMF
were located in the allowed ranges of an SOD mimic.
+
−3
Table 5. Redox potential data vs. Fc /Fc for 10 M solution of 1 and
−
1
2
in DMF containing 0.1 M TBPA and scan rate 100 mV s .
Complex
E
pa/V
E
pc/V
E
1/2/V
ΔE/V
1
2
0.34
0.17
0.22
0.08
0.28
0.13
0.12
0.09

Copper(II) complexes with hydroxymethyl pendants
2401
−
1
Figure 2. Cyclic voltammograms of 1 and 2 in DMF at 298 K and 100 mV s scan rate.
3
.5. Electronspray ionization mass spectrometry
To prove the state of 1 and 2 in the phosphate buffer solutions at pH 7.8, mass spectra
ESI, positive-detection mode) were performed. Sample solutions were injected into the ion
(
−
1
source with the addition of methanol at a flow rate of 5 mL min . As shown in figure S3,
for 1, the prominent peak at 286.67 m/z was assigned to the [Cu (HL) (L) ] double-charge
2
+
2
2
2
positive species, due to loss of the two non-coordinated perchlorate anions. The peaks at
+
+
1
[
13.33, 135.42, 323.25, 398.00, and 461.17 m/z may be related to [HL + H] , [HL + Na] ,
2
2 2 2 2 2 4 2 2 2 2
+
2+
2+
Cu (HL) (L) + 4H O] , [Cu (HL) (L) ] , and [Cu (HL) (L) + 4 MeOH] , respectively.
As for 2, the peaks at 113.25 and 181.50 m/z are attributed to HL and phen, respectively.
+
The prominent peak for 2 at 354.50 m/z corresponds to [Cu(L)(phen)] , due to the hydroxyl
deprotonation of HL at pH 7.8 and further providing the coordination of oxygen, resulting
2
+
in the loss of one HL from [Cu(HL) (phen)] . The peak at 423.25 m/z can be ascribed to
2
+
the solvent adduct [Cu(L)(phen)+2H O + 2MeOH] .
2
3
.6. SOD-like activity
The SOD-like (SOD = superoxide dismutase) activities of 1 and 2 were investigated by
NBT assay [23–26]. The imidazole derivative with a hydroxymethyl arm provides a hydro-
philic environment similar to that in the active site of the native enzyme. The concentrations
of the complex required to attain 50% inhibition of the reduction (defined as IC ) were
5
0
determined, and IC50 value of bovine erythrocyte SOD is 0.042 ± 0.01 μM, almost identical
to 0.04 μM value reported [37]. From the control experiments of copper(II) perchlorate, the
IC50 values of free copper(II) is 4.01 μM, which is very close to that reported previously
ÁÀ
[
38]. The ligand did not exhibit O2 scavenger effects under the same conditions (IC50
>
1
000 μM). As shown in figure 3, IC50 values of 1 and 2 at pH 7.8, obtained from the plot
of inhibition percentage versus copper(II) ion concentration, are 0.10 and 0.19 μM, respec-
tively. Despite the lower activity than the native enzyme, 1 and 2 are among the relatively
active model compounds of SOD enzyme [39, 40]. The difference in IC50 values for the
two complexes should be ascribed to different structures between 1 and 2. For 2, the CuN4
coordination is different from that of the natural enzyme. Meanwhile, phenanthroline as the
rigid co-ligand in 2 is unfavorable for accommodating the geometrical transformation from

2
402
Y.-H. Zhou et al.
+
Figure 3. Percentage of inhibition of superoxide radicals by NBT absorption at 560 nm in the presence of bovine
erythrocyte SOD (BeSOD) (●), 1 (■), and 2 (▲) at pH 7.8 phosphate buffer.
Cu(II) to Cu(I) during the catalytic process. The higher activity of 1 may be attributed to
the two labile hydroxymethyl groups in the coordination, which are proposed to be easily
À
ÁÀ
substituted by O2 in the catalytic process [15], which is similar to O displacing water
2
ÁÀ
bound to the copper site in the mechanism of O2 dismutation by native SOD [40–42].
4
. Conclusion
We report the syntheses, structures, and properties of two new copper(II) complexes based
on imidazole with a hydroxymethyl pendant, [Cu (HL) (L) ](ClO ) (1) and [Cu
2
2
2
4 2
(
HL) (phen)](ClO ) (2). The quasi-reversible Cu(II)/Cu(I) redox waves of 1 and 2 were
2 4 2
determined by cyclic voltammetry. The atomic net charges distribution and frontier molecu-
lar orbital energies were obtained by calculation. Complex 1 exhibits high SOD-like activ-
ity, which can be attributed to the CuN O coordination configuration and the labile
2
2
hydroxymethyl pendants.
Supplementary material
Crystallographic data for HL, 1 and 2 have been deposited with the Cambridge Crystallo-
graphic Data Center as supplementary publication CCDC reference numbers 959840,
9
59839 and 981394, respectively. These data can be obtained free of charge via http://
www.ccdc.cam.ac.uk/conts/retrieving.html, or from the Cambridge Crystallographic Data
Center, 12 Union Road, Cambridge CB2 1EZ, UK; Fax: (+44) 1223 336 033; or Email:
deposit@ccdc.cam.ac.uk.

Copper(II) complexes with hydroxymethyl pendants
2403
Funding
This work was supported by the National Natural Science Foundation of China [grant number 20901002].
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