Synthesis, structures and urease inhibition studies of copper(II) and nickel(II) complexes with bidentate N,O-donor Schiff base ligands

Article abstract of DOI:10.1016/j.jinorgbio.2011.12.006

Five mononuclear copper(II) and nickel(II) complexes of Schiff base ligands derived from 4-hydroxyphenethylamine and 2-phenylethylamine were synthesized and determined by single crystal X-ray analysis. The crystal structures of these complexes presented t

Full text of DOI:10.1016/j.jinorgbio.2011.12.006

Journal of Inorganic Biochemistry 108 (2012) 22–29
Contents lists available at SciVerse ScienceDirect
Journal of Inorganic Biochemistry
journal homepage: www.elsevier.com/locate/jinorgbio
Synthesis, structures and urease inhibition studies of copper(II) and nickel(II)
complexes with bidentate N,O-donor Schiff base ligands
a
a
Xiongwei Dong a, Yuguang Li a,b, , Zuowen Li , Yongming Cui , Hailiang Zhu
⁎
b,⁎⁎
a
School of Chemistry and Chemical Engineering, Wuhan Textile University, Wuhan 430073, PR China
State Key Laboratory of Pharmaceutical Biotechnology, Nanjing University, Nanjing 210093, PR China
b
a r t i c l e i n f o
a b s t r a c t
Article history:
Five mononuclear copper(II) and nickel(II) complexes of Schiff base ligands derived from 4-
hydroxyphenethylamine and 2-phenylethylamine were synthesized and determined by single crystal X-ray anal-
ysis. The crystal structures of these complexes presented the square planar coordination geometry at the metal
center. The inhibitory activity of all the obtained complexes was tested in vitro against jack bean urease. It was
found that Schiff base copper(II) complexes, namely [Cu(C15H13BrNO2)2]·2(C6H7N) (1), [Cu(C15H12Br2-
NO2)2]·2(DMF) (2), Cu(C19H16NO2)2 (3) and Cu(C19H16NO)2 (5), showed strong inhibitory activity against
jack bean urease (IC50 =1.45–3.59 μM), while Schiff base nickel(II) complex, [Ni(C19H16NO2)2]·2(DMF) (4),
exhibited weak inhibitory activity (IC50 >50 μM). Their structure–activity relationships were further discussed.
© 2011 Elsevier Inc. All rights reserved.
Received 8 September 2011
Received in revised form 25 November 2011
Accepted 21 December 2011
Available online 29 December 2011
Keywords:
Copper(II) complexes
Schiff base ligands
X-ray structure
Urease inhibitors
Molecular docking
1. Introduction
inhibitors has so far relied upon extended screen tests because of
the low efficiency and negative side effects of the presently available
Urease (urea amidohydrolase; E.C.3.5.1.5) is a nickel-containing
enzyme that catalyzes the rapid hydrolysis of urea to form ammonia
and carbamate in a variety of algae, bacteria, fungi and plants [1–5].
It participates in environmental nitrogen transformations to supply
these organisms with a nitrogen source for growth [3]. On the other
hand, the reaction catalyzed by the dinuclear nickel active site of ure-
ase causes an accumulation of ammonia and an abrupt pH increase,
which has negative side effects in agriculture and health. For exam-
ple, urease serves as a virulence factor in pathogens that are respon-
sible for the development of kidney stones, pyelonephritis, peptic
ulcers, and other disease states [3,4]. In another context, urease can
severely decrease the efficiency of urea fertilizers to cause the release
of large amounts of ammonia and further induce plant damage by
ammonia toxicity and soil pH increase [5]. Therefore, the capability
to control the rate of the enzymatic urea hydrolysis using urease in-
hibitors is an important goal to pursue.
inhibitors against plant, bacterial and fungal ureases. In earlier stud-
ies, we have investigated the inhibition of jack bean urease by a series
of Schiff base metal complexes and their urease inhibitory activity
was found [20–22]. An interesting observation is that their reported
biological activities are significantly influenced by metal center and
the positions of the substituent groups such as halogen atoms in the
aromatic ring. The full mechanism of the action is unclear yet. As a
continuation of our work on Schiff base complexes as the urease in-
hibitor, the urease inhibitory activities of Schiff base copper(II) com-
plexes 1, 2, 3 and 5, and nickel(II) complex 4 were investigated and
reported in this paper. Here, Schiff base ligands HL1, HL2 and HL3
were obtained from the condensation of 4-hydroxyphenethylamine
with 5-bromosalicyladehyde, 3,5-dibromosalicyladehyde and 2-
hydroxy-1-naphthaldehyde, respectively, while ligand HL4 was
prepared from the reaction of 2-phenylethylamine with 2-hydroxy-
1-naphthaldehyde (Scheme 1). A preliminary docking study was
carried out using the DOCK program to gain an understanding of
urease inhibitory activity of complexes 1–5 and their structure–
activity relationships were also described within this paper.
Recently, urease inhibition studies have attracted increasing at-
tention [6–9] and the numerous urease inhibitors have also been
reported [10–19]. Among the known inhibitors of urease, hydroxamic
acids, phosphoramides and thiols are the best recognized urease in-
hibitors [10–13]. However, the discovery of new and more efficient
2. Experimental section
2.1. Materials and physical measurements
⁎
Correspondence to: Y. Li, School of Chemistry and Chemical Engineering, Wuhan Tex-
Urease (from jack beans, type III, activity 22 units/mg solid), HEPES
(Ultra) buffer and urea (Molecular Biology Reagent) were from Sigma.
All other chemicals and solvents were purchased from Aldrich and
tile University, Wuhan 430073, PR China. Tel./fax: +86 27 8761 1776.
⁎⁎ Corresponding author. Tel./fax: +86 27 8761 1776.
E-mail address: liyg2010@wtu.edu.cn (Y. Li).
0162-0134/$ – see front matter © 2011 Elsevier Inc. All rights reserved.
doi:10.1016/j.jinorgbio.2011.12.006

X. Dong et al. / Journal of Inorganic Biochemistry 108 (2012) 22–29
23
4-methylpyridine (5 mL) or Ni(NO3)2·6H2O (0.5 mmol) in methanol
(5 mL). The resulting solution was stirred for 30 min at room temperature
and then filtered. The filtrate was kept in air for about 7 days, forming
block crystals. The crystals were isolated, washed three times with dis-
tilled water and dried in a vacuum desiccator containing anhydrous CaCl2.
h
ꢀ
ꢁ i
Cu L1
2ð4−methylpyridineÞ
ð1Þ
⋅
2
Brown black solid, yield: 235 mg (53%). IR (KBr, cm−1): 3447,
2901, 2599, 1618, 1516, 1460,1386, 1322, 1259, 1174, 1069, 1013,
823, 701, 648, 512, 484, 431. UV–visible [DMSO–H2O (1:1 v/v), λ/
nm (ε/M−1 cm−1)]: 370 (9,740), 294 (11,850), 258 (28,480). Anal.
Calcd for C42H40Br2CuN4O4: C, 56.80; H, 4.54; N, 6.31. Found: C,
56.96; H, 4.71; N, 6.10%.
h
ꢀ
ꢁ i
Cu L2
2ðDMFÞ
⋅
ð2Þ
2
Brown black solid, yield: 282 mg (56%). IR (KBr, cm−1): 3422, 2929,
1664, 1620, 1513, 1446, 1384, 1321, 1243, 1157, 1102, 858, 826, 707,
669, 523, 489, 432. UV–visible [DMSO–H2O (1:1 v/v), λ/nm (ε/
M−1 cm−1)]: 376 (8,550), 268 (20,730), 256 (25,270). Anal. Calcd for C-
36H38Br4CuN4O6: C, 42.99; H, 3.81; N, 5.57. Found: C, 42.76; H, 3.92; N,
5.31%.
h
ꢀ
ꢁ i
Cu L3
ð3Þ
2
Brown black solid, yield: 200 mg (62%). IR (KBr, cm−1): 3370, 3062,
Scheme 1. The Schiff base ligands HL1, HL2, HL3 and HL4 used for the syntheses of com-
plexes 1–5.
2914, 2856, 1610, 1546, 1511, 1439, 1415, 1364, 1343, 1305, 1249,
1190, 1139, 1097, 1033, 965, 824, 748, 647, 603, 563, 524, 492, 463,
419. UV–visible [DMSO–H2O (1:1 v/v), λ/nm (ε/M−1 cm−1)]: 414
(8,650), 396 (13,290), 380 (12,080), 310 (23,180), 262 (29,370). Anal.
Calcd for C38H32CuN2O4: C, 70.85; H, 5.01; N, 4.35. Found: C, 70.71; H,
5.07; N, 4.28%.
used as received. Distilled water was used for all procedures. 1H NMR
spectra were recorded on a Bruker DRX-500 spectrometer. Chemical
shifts were reported in ppm from tetramethylsilane as an internal stan-
dard (1H, 0.00 ppm). Mass spectra were obtained on a Micromass GC–
TOF for EI–MS (70 eV). Elemental analyses were performed using an
elementar vario EL III elemental analyzer. Infrared spectra were
obtained on a Nexus 870 FT-IR spectrophotometer using KBr pellets in
the 4000–400 cm−1 range. UV–visible spectra were measured on a Shi-
madzu UV-160 A spectrophotometer using DMSO–H2O (1:1 v/v) sol-
vents in the 800–200 nm range. The enzyme inhibitory activity was
measured on a Bio-Tek Synergy™ HT microplate reader.
h
ꢀ
ꢁ i
Ni L3
2ðDMFÞ
⋅
ð4Þ
2
Brownish yellow solid, yield: 188 mg (48%). IR (KBr, cm−1): 3167,
3062, 2926, 2809, 1662, 1611, 1542, 1511, 1442, 1412, 1368, 1344,
1237, 1195, 1142, 1102, 1058, 1032, 970, 934, 826, 749, 664, 570, 534,
477, 421. UV–visible [DMSO–H2O (1:1 v/v), λ/nm (ε/M−1 cm−1)]: 416
(7,280), 400 (7,410), 308 (8,920), 272 (16,850), 263 (17,730), 252
(19,710). Anal. Calcd for C44H46NiN4O6: C, 67.27; H, 5.90; N, 7.13. Found:
C, 67.01; H, 5.97; N, 7.01%.
2.2. General procedure for the synthesis of Schiff base ligands HL1–4
(exemplified by HL3)
h
ꢀ
ꢁ i
Cu L4
ð5Þ
4-Hydroxyphenethylamine (0.274 g, 2.0 mmol) was added to the
solution of 2-hydroxy-1-naphthaldehyde (0.344 g, 2.0 mmol) in
methanol (20 mL). The mixture was refluxed in methanol at 65 °C
within 1 h. Subsequently, the solution was filtered and kept in air
for about 7 days, to afford yellow block-shaped crystals of HL3.
Yield: 460 mg (79%), m.p. 197 °C dec. 1H NMR (500 MHz, d6-DMSO)
δ: 13.99 (s, 1H), 9.20 (s, 1H), 8.99 (d, J=10.5 Hz, 1H), 7.97–6.69 (m,
6H), 7.08 (d, J=8.0 Hz, 2H), 6.67 (d, J=8.0 Hz, 2H), 3.82 (t, 2H),
2.87 (t, 2H). EI–MS (70 eV): m/z 291 (M+, 5%). IR (KBr, cm−1):
3424, 2908, 2593, 1633, 1546, 1510, 1447, 1365, 1241, 1190, 1090,
855, 827, 748, 507, 487, 481, 435, 409. UV–visible [DMSO–H2O
(1:1 v/v), λ/nm (ε/M−1 cm−1)]: 416 (12,360), 400 (11,760), 308
(10,130), 268 (12,510), 262 (17,380), 252 (20,070). Anal. Calcd for
2
Brown black solid, yield: 205 mg (67%). IR (KBr, cm−1): 3444,
3025, 2907, 1612, 1540, 1500, 1462, 1410, 1365, 1202, 1091, 1027,
958, 825, 743, 696, 522, 499, 415. UV–visible [DMSO–H2O (1:1 v/v),
λ/nm (ε/M−1 cm−1)]: 420 (6,330), 400 (6,060), 312 (6,780), 248
(11,660). Anal. Calcd for C38H32CuN2O2: C, 74.55; H, 5.27; N, 4.58.
Found: C, 74.43; H, 5.36; N, 4.33%.
2.4. Crystal structure determinations
X-ray crystallographic data [23,24] were collected on a Bruker
SMART Apex II CCD diffractometer using graphite-monochromated
Mo Kα (λ=0.71073 Å) radiation. The collected data were reduced
using the SAINT program, and empirical absorption corrections
were performed using the SADABS program. The structures were
solved by direct methods and refined against F2 by full-matrix least-
squares methods using the SHELXTL version 5.1. All of the non-
hydrogen atoms were refined anisotropically. All other hydrogen
atoms were placed in geometrically ideal positions and constrained
C19H17NO2: C, 78.33; H, 5.88; N, 4.81. Found: C, 78.11; H, 5.96; N, 4.60.
2.3. General method for the preparation of complexes 1–5
The Schiff base ligands HL1–4 (1 mmol) were dissolved in the solvent
mixture (10 mL) of methanol and N,N-dimethyl formamide (DMF)
(1:1), respectively, which was added to Cu(NO3)2·3H2O (0.5 mmol) in

24
X. Dong et al. / Journal of Inorganic Biochemistry 108 (2012) 22–29
Table 2
to ride on their parent atoms. The crystallographic data for the Schiff
base ligands (HL2, HL3) and Schiff base metal complexes (1, 2, 3, 4
and 5) are summarized in Tables 1 and 2.
Crystal data for HL2, HL3 and complexes 1–5.
Compound
HL2
HL3
Empirical formula
C15H13Br2NO2
399.08
C19H17NO2
291.34
Orthorhombic
Pbca
12.5136(17)
11.4059(16)
20.092(3)
90
90
90
291(2)
2867.7(7)
8
2.5. Measurement of jack bean urease inhibitory activity
Molecular weight
Crystal system
Space group
a (Å)
b (Å)
c (Å)
α (°)
β (°)
γ (°)
Orthorhombic
P212121
8.1072(16)
8.8634(17)
21.038(4)
90
90
90
298(2)
1511.7(5)
4
1.754
784
5.362
2958/0/182
1.003
The measurement of urease activity was carried out according to
the literature reported by Tanaka [25]. Generally, the assay mixture,
containing 25 μL of jack bean urease (12 kU/L) and 25 μL of the tested
complexes of different concentrations (dissolved in DMSO/H2O mix-
ture (1:1 v/v)), was preincubated for 1 h at 37 °C in a 96-well assay
plate. After preincubation, 200 μL of 100 mM HEPES (N-[2-hydroxy-
ethyl] piperazine-N′-[2-ethanesulfonic acid]) buffer [26] pH=6.8
containing 500 mM urea and 0.002% phenol red were added and in-
cubated at 37 °C. The reaction was measured by micro plate reader
(570 nm), which was required to produce enough ammonium car-
bonate to raise the pH of a HEPES buffer from 6.8 to 7.7, the end-
point being determined by the color of phenol red indicator [27].
T (K)
V (Å3)
Z
ρcalc. (g·cm−3
F(000)
)
1.350
1232
0.088
2527/0/203
1.001
μ(Mo-Kα) (mm−1
)
Data/restraint/parameters
Goodness-of-fit on F2
Final R1, wR2 [I>2σ(I)]
0.0364/0.0716
0.0634/0.0790
2.6. Docking simulations
Molecular docking of the inhibitor with the three-dimensional
structure of jack bean urease (entry 3LA4 in the Protein Data Bank)
was carried out using the DOCK 4.2 program suite [28–31]. The graphi-
cal user interface AutoDockTools (ADT 1.4.5) was performed to setup
every inhibitor–enzyme interaction, where all hydrogen atoms were
added, Gasteiger charges were calculated and nonpolar hydrogen
atoms were merged to carbon atoms. The Ni initial parameters are set
as r=1.170 Å, q=+2.0, and van der Waals well depth of 0.100 kcal/
mol [32]. As performed by the graphical user interface AutoDockTools,
the catalytic center and the peripheral anionic site of the target protein
were scanned to evaluate the modeled binding mode of the inhibitor–
urease complex. The flexible docking of the ligand structures was
done by the Lamarckian genetic algorithm (LGA), searching for favor-
able bonding conformations of the ligands at the sites of the target pro-
tein. The docking procedure of Schiff base copper(II) complexes 1, 2, 3
and 5 with the enzyme active site of jack bean urease was performed
as described previously by our group [33,34].
the bidentate Schiff base ligands. The Schiff base ligands HL1, HL2 and
HL3 were prepared by the reaction of 4-hydroxyphenethylamine with
the corresponding 5-bromosalicylaldehyde, 3,5-dibromosalicyl-alde-
hyde and 2-hydroxy-1-naphthaldehyde in nearly 70–80% yield in
methanol, while the ligand HL4 was obtained by the reaction of 2-
phenylethylamine with 2-hydroxy-1-naphthaldehyde. These Schiff
base ligands are the stable solids and can be stored without precautions.
In general, Schiff base ligands HL1–4 may dissolve in the polar solvent
such as methanol and N,N-dimethyl formamide (DMF). Treatment of
the Schiff base ligands HL1–4 with the respective metal salt,
Cu(NO3)2·3H2O and Ni(NO3)2·6H2O, in a 2:1 M ratio at ambient tem-
perature led to the copper(II) complexes 1, 2, 3 and 5, and nickel(II)
complex 4. Crystals of complexes 1–5 suitable for X-ray diffraction
could be isolated from 4-methylpyridine/methanol or DMF/methanol
after slow evaporation of the solvent over a period of a week. The spec-
troscopic data is in good agreement with the chemical formula pro-
posed for Schiff base complexes 1–5.
3. Results and discussion
3.2. Spectroscopic studies (Supplementary data)
3.1. Synthesis
On the basis of the structure found and a comparison with spectra of
related complexes [35–37], the IR spectra of complexes 1–5 have been
tentatively assigned. The broadness of the ν(O\H) band between
3447 and 3167 cm−1 may be attributed to the phenolic hydroxyl
Condensation of salicylaldehydes or salicylaldehyde derivatives
with primary amines leads to the formation of an important class of
Table 1
Crystal data for HL2, HL3 and complexes 1–5.
Complex
1
2
3
4
5
Empirical formula
C42H40Br2CuN4O4
888.14
Monoclinic
C2/c
38.069(3)
5.7043(5)
19.7619(16)
90.00
114.206(2)
90.00
298(2)
3914.1(6)
4
1.507
1804
C36H38Br4CuN4O6
1005.88
Triclinic
C38H32CuN2O4
644.20
Triclinic
P−1
9.069(2)
10.030(2)
10.157(2)
67.743(4)
63.737(4)
69.342(5)
298(2)
747.5(3)
1
1.431
335
0.777
C44H46N4NiO6
785.56
Monoclinic
P21/c
18.276(2)
10.2304(12)
11.0151(13)
90.00
106.473(2)
106.473(2)
291(2)
1975.0(4)
2
1.321
828
0.545
C38H32CuN2O2
612.20
Triclinic
Molecular weight
Crystal system
Space group
a (Å)
b (Å)
c (Å)
α (°)
β (°)
γ (°)
P−1
P−1
6.2178(6)
9.2270(11)
18.1787(18)
78.042(2)
84.892(2)
70.359(2)
291(2)
5.1785(3)
15.8614(10)
18.1411(11)
90.2320(10)
96.6460(10)
93.4460(10)
298(2)
T (K)
V (Å3)
960.73(18)
1
1.739
499
4.777
1477.30(15)
2
1.376
638
0.777
Z
ρcalc. (g·cm−3
F(000)
)
μ(Mo-Kα) (mm−1
)
2.646
Data/restraint/parameters
Goodness-of-fit on F2
Final R1, wR2 [I>2σ(I)]
3825/0/243
1.009
0.0450, 0.1046
3762/26/235
1.003
0.0329, 0.0894
2898/0/206
0.943
0.0660, 0.0793
3874/0/253
1.002
0.0485, 0.1116
5718/0/391
1.003
0.0364, 0.0836

X. Dong et al. / Journal of Inorganic Biochemistry 108 (2012) 22–29
25
Fig. 1. One-dimensional zigzag chain structure of HL2. (symmetry codes: (i) 1−x, −1/2+y, 1/2−z; (ii) 1−x, 1/2+y, 1/2−z).
group of complexes 1–5. The IR spectra of complexes 1–5 exhibit strong
absorption between 1620 and 1610 cm−1, assignable to the v(C_N)
absorption. It can be observed that the v(C_N) vibration band suffers
a negative shift in complexes 2 and 3 with respect to the corresponding
free ligand HL2 and HL3 at 1643 and 1633 cm−1, thus indicating that the
imine nitrogen atom is involved in coordination to the metal ion. The
appearances of the strong absorption at 1664 and 1662 cm−1 in 2 and
4 were attributed to the v(C_O) absorption of the lattice DMF mole-
cule. The broad strong absorption between 1546 and 1500 cm−1 in all
complexes could be reasonably attributed to the presence of the
v(C_C) stretching vibration of the aromatic ring backbone. The UV–
vis spectra for the Schiff base complexes 1–5 were obtained in assay
conditions (DMSO/H2O, 1:1 v/v). The electronic spectra of the Schiff
base copper(II) complexes 1 and 2 are very similar and show the mod-
erately intense UV band near 370 nm originating from a phenolate to
metal ion charge-transfer [38,39]. The absorption maxima of complex
2 are blue-shifted compared to 1, probably because of the acceptor ef-
fect of the 3-bromine substituent. In the electronic spectra of complexes
3, 4 and 5, the intense higher-energy bands at around 260 nm are attrib-
uted to an intraligand charge transfer (π→π*). The formation of com-
plexes 3 and 5 in DMSO/H2O mixture is supported by the strong and
broad charge-transfer band at 310 and 312 nm, and also by the evolu-
tion of a shoulder in the close UV range between 380 and 420 nm, orig-
inated from a charge transfer band. The shoulder centered at 388 nm in
the UV–vis spectrum of 3 is blue-shifted relative to its free ligand HL3.
Similar spectra are observed for Schiff base nickel(II) complex 4 and
its free ligand HL3, whereas 4 exhibits somewhat weaker absorption
bands appeared at 308 nm, 400 nm, 416 nm than the corresponding
HL3 ligand. In the electronic spectra of all complexes 1–5, bands
associated with d→d transitions were not detected, perhaps due to
the intensity of the charge transfer and intraligand transitions [40].
3.3. Crystal structure description
The study shows that the Schiff base ligands HL1, HL2, HL3 and HL4
are much alike (Scheme 1). These bidentate N,O-donor ligands are
able to coordinate as a monoanionic species through their deproto-
nated phenolic hydroxyl groups. Analogous N,O-bidentate chelating
Schiff base species has been obtained from the condensation of salicy-
laldehyde with 4-hydroxyphenethylamine in our group [41]. Unfor-
tunately, the Schiff base ligands HL1 and HL4 could not be
crystallized in a form suitable for X-ray single crystal diffraction stud-
ies and thus only the crystal structures of HL2 and HL3 have been de-
termined. The solid-state structures of the Schiff base ligands HL2 and
HL3 are shown in Figs. 1 and 2. In the ligands HL2 and HL3 derived
from 3,5-dibromosalicylaldehyde and 2-hydroxy-1-naphthaldehyde,
the C_N bond distances are 1.286(6) and 1.307(3)Å, respectively. In-
terestingly, the Schiff base ligands HL2 and HL3 were linked into a zig-
zag chain by the intramolecular O\H⋯N hydrogen bonds and the
intermolecular O\H⋯O hydrogen bonds between the corresponding
adjacent molecules [O1⋯N1 2.553(4)Å, O2⋯O1# 2.720(4)Å for HL2;
O1⋯N1 2.592(3), O2⋯O1# 2.662(3)Å for HL3].
The Schiff base metal complexes 1–5 with general formula M(L)2,
where L=L1, L2, L3 and L4, were obtained from the combination of
two equivalents of the Schiff base ligands HL1–4 with a metal ion
(M=Cu2+, Ni2+). The molecular structures of complexes 1–5 were
determined by single crystal X-ray analysis as shown in Figs. 3–7.
All five complexes afford the square planar coordination geometry,
Fig. 2. One-dimensional zigzag chain structure of HL3. (symmetry codes: (i) 3/2−x, −y, −1/2+z; (ii) 3/2−x, −y, 1/2+z).

26
X. Dong et al. / Journal of Inorganic Biochemistry 108 (2012) 22–29
Fig. 3. Ball-and-stick representation of the molecular structure of 1 (symmetry codes: (i) 1/2−x, 5/2−y, −z; (ii) 1/2−x, 5/2+y, 1/2−z; (iii) x, −y, −1/2+z), atoms are shown as
sphere of arbitrary diameter.
Fig. 4. Ball-and-stick representation of the molecular structure of 2 (symmetry codes: (i) 1−x, −y, 1−z; (ii) 2−x, −y, −z; (iii) −1+x, y, 1+z), atoms are shown as sphere of
arbitrary diameter.
where the metal ion is four-coordinated by two imine N atoms and
two phenolic O atoms from two Schiff base ligands (L) in a trans po-
sition. Selected bond distances and bond angles at the metal center
for complexes 1–5 are listed in Table 3. Single crystal X-ray diffraction
reveals that Schiff base copper(II) complexes 1 and 2 crystallize in the
monoclinic C2/c (No. 15) space group and triclinic P-1 (No. 2) space
group, respectively. As depicted in Figs. 3 and 4, complexes 1 and 2
are the mononuclear copper(II) species with formula [Cu(L1)2]·2(4-
methylpyridine) and [Cu(L2)2]·2DMF, to afford the square planar
trans-[CuN2O2] coordination geometry, where the Cu(II) atom lies
on a center-of-inversion. The average Cu\O and Cu\N bond dis-
tances of 1.882(2) and 2.004(2)Å in complex 1 are slightly shorter
than that of 1.906(2) and 2.010(2)Å observed in 2, respectively,
probably due to the electron withdrawing effect of the bromine sub-
stituent. This suggests that the position of the bromine atom in the ar-
omatic ring has significant effect on the coordination abilities of the
ligands [39]. The Cu(L1)2 moiety in the crystal structure of 1 is linked
with two co-crystallized 4-methylpyridine solvent molecules through
the intermolecular O2\H2⋯N2 hydrogen bonds [O2⋯N2 2.713(5)Å]
(Fig. 3), while the Cu(L2)2 moiety in 2 is connected with two DMF
molecules through the intermolecular O2\H2⋯O3 hydrogen bonds
[O2⋯O3 2.588(8)Å] (Fig. 4).
3, the crystal structure of 4 consists of a discrete mononuclear Ni(L3)2
unit and two lattice DMF molecules. The coordination geometry of the
nickel(II) atom in the Ni(L3)2 unit closely resembles that observed for
complex 3. Analogous copper(II) and nickel(II) species of Schiff base
ligands derived from 4-hydroxyphenethylamine have been reported
by our group [20]. The bond distances of Ni1\O1 and Ni1\N1 are
1.825 (2) and 1.911(2)Å, which are comparable with the correspond-
ing values reported for analogous square planar Ni(II) species
[20,36,42]. The Ni(L3)2 moiety in 4 is further connected with two
DMF molecules through the intermolecular O2\H2⋯O3 hydrogen
The X-ray crystal structure reveals that complexes 3 and 4 are the
mononuclear copper(II) and nickel(II) species of formula Cu(L3)2 and
[Ni(L3)2]·2DMF, respectively (Figs. 5 and 6). These two complexes
have the same bidentate backbone of the chelate Schiff base ligand
HL3 and the same metal coordination spheres. In contrast to complex
Fig. 5. Ball-and-stick representation of the molecular structure of 3 (symmetry code:
(i) 1−x, 1−y, 1−z), atoms are shown as sphere of arbitrary diameter.

X. Dong et al. / Journal of Inorganic Biochemistry 108 (2012) 22–29
27
Fig. 6. Ball-and-stick representation of the molecular structure of 4 (symmetry codes: (i) 1−x, 1−y, −z; (ii) −x, 1−y, −z; (iii) 1+x, y, z), atoms are shown as sphere of arbitrary
diameter.
bonds [O2⋯O3 2.663(3)Å] (Fig. 6). In contrast to complexes 1–3, the
solid-state structure of the Schiff base copper(II) complex 5 derived
from 2-phenylethylamine contains two crystallographically indepen-
dent molecules. The molecular structure of one of the two rather sim-
ilar complex units, Cu(L4)2, is represented in Fig. 7. The copper(II)
atom in each mononuclear Cu(L4)2 unit of 5 also lies on a crystallo-
graphic inversion center (symmetry codes: 1−x, 1−y, 1−z for
Cu1; 1−x, 2−y, −z for Cu2). The average bond distances of
Cu1\O1 and Cu1\N1 are 1.879(2) and 1.998(2)Å, respectively,
while the average bond distances of Cu2\O2 and Cu2\N2 are
1.881(2) and 2.006(2)Å.
derived from 4-hydroxyphenethylamine is much more potent than
complex 5 derived from 2-phenylethylamine. This defines the mini-
mal substitution patterns in the aromatic ring for obtaining the po-
tent activity. Among the four Schiff base copper(II) complexes 1, 2,
3 and 5 tested, the most potent activity was observed in complex 3
only. These observations are in agreement with the previously
reported Schiff base copper(II) complexes derived from 4-
hydroxyphenethylamine [20]. When compared with complex 3, the
Schiff base nickel(II) complex 4 exhibits weaker urease inhibitory ac-
tivity (IC50 =52 μM) under the same condition. The results indicate
that inhibitory activities of Schiff base metal complexes as the urease
inhibitor depend on not only the organic ligands but also the central
ions.
3.4. Inhibitory activity against jack bean urease
The Schiff base ligands HL1–4 and the corresponding copper(II) 1,
2, 3 and 5 and nickel(II) complex 4 were screened for inhibitory activ-
ity against jack bean urease (see Table 4). It was found that all the
synthesized ligands HL1, HL2, HL3 and HL4 exhibited no ability to in-
hibit the jack bean urease. Compared with the standard inhibitor
acetohydroxamic acid (AHA, IC50 =63.00 μM), the Schiff base cop-
per(II) complexes 1, 2, 3 and 5 displayed potent inhibitory activity
against jack bean urease. Generally, heavy metal ions are believed to
inhibit the urease by binding to the sulfhydryl groups of cysteines,
and possibly nitrogen-(histidine) and oxygen-(aspartic and glutamic
acids) in the urease active site [43–46]. Thus it can be seen that coor-
dination to copper(II) ion resulted in the improved inhibitory activity
[47–49]. It should be noted that in terms of the inhibitory strength to-
wards jack bean urease the Schiff base copper(II) complexes studied
form the order: 3>1>2>5. Here, complex 1 is more potent than
complex 2 as a result of the difference of the electron-withdrawing
bromine substituent on the aromatic ring. Interestingly, complex 3
3.5. Molecular docking study
The binding models of Schiff base copper(II) complexes 1, 2, 3 and
5 with jack bean urease were simulated using the Dock program to
validate their structure–activity relationships (Fig. 8) [28–31]. The re-
sults revealed that the complex molecules were well filled in the ac-
tive pocket of jack bean urease. Additional interactions have been
established in a variety of conformations because of the flexibilities
of the chelating phenolic oxygen atom and the amino acid residues
Table 3
Selected bond lengths (Å) and angles (°) for complexes 1–5.
1 (i=1/2−x, 5/2−y, −z)
Cu(1)\O(1)
1.882(2)
1.882(2)
2.004(2)
2.004(2)
O(1)\Cu(1)\N(1)
O(1)i\Cu(1)\N(1)
O(1)\Cu(1)\O(1)i
91.57(10)
88.43(10)
180.0
Cu(1)\O(1)i
Cu(1)\N(1)
Cu(1)\N(1)i
2 (i=1−x, -y, 1−z)
Cu(1)\O(1)
1.906(2)
1.906(2)
2.010(2)
2.010(2)
O(1)\Cu(1)\N(1)
O(1)i\Cu(1)\N(1)
O(1)\Cu(1)\O(1)i
91.70(9)
88.29(9)
179.999(1)
Cu(1)\O(1)i
Cu(1)\N(1)
Cu(1)\N(1)i
3 (i=1−x, 1−y, 1−z)
Cu(1)\O(1)
1.900(3)
1.900(3)
1.987(3)
1.987(3)
O(1)\Cu(1)\N(1)
O(1)a\Cu(1)\N(1)
O(1)\Cu(1)\O(1)a
90.27(12)
89.73(12)
180.000(1)
Cu(1)\O(1)a
Cu(1)\N(1)
Cu(1)\N(1)a
4 (i=1−x, 1−y, −z)
Ni(1)\O(1)
Ni(1)\O(1)i
Ni(1)\N(1)
Ni(1)\N(1)i
1.8255(16)
1.8255(16)
1.911(2)
O(1)\Ni(1)\N(1)
O(1)i\Ni(1)\N(1)
O(1)\Ni(1)\O(1)i
87.82(8)
92.18(8)
180.000(2)
1.911(2)
5 (i=1−x, 1−y, 1−z; ii=1−x, 2−y, −z)
Cu(1)\O(1)
Cu(1)\O(1)i
Cu(1)\N(1)
Cu(1)\N(1)i
Cu(2)\O(2)
Cu(2)\O(2)ii
Cu(2)\N(2)
1.8793(14)
1.8793(14)
1.9981(16)
1.9981(16)
1.8814(14)
1.8814(14)
2.0055(16)
O(1)\Cu(1)\N(1)
O(1)i\Cu(1)\N(1)
O(1)\Cu(1)\O(1)i
Cu(2)\N(2)ii
O(2)\Cu(2)\N(2)
O(2)ii\Cu(2)\N(2)
O(2)\Cu(2)\O(2)ii
90.84(6)
89.16(6)
179.999(1)
2.0055(16)
90.53(6)
89.47(6)
180.0
Fig. 7. Ball-and-stick representation of the molecular structure of 5 (symmetry code:
(i) 1−x, 1−y, 1−z), atoms are shown as sphere of arbitrary diameter.

28
X. Dong et al. / Journal of Inorganic Biochemistry 108 (2012) 22–29
Table 4
and other intermolecular interactions with the amino acid residues such
as His407, His409, His492, His593 and Asp633. In addition, the distances
between two nickel(II) atoms in the active site of urease and the respec-
tive copper(II) atom of complexes 1 and 2 were calculated (9.030(2) and
9.002(2)Å for complex 1; 8.971(2) and 8.703(2)Å for complex 2).
In the best docking conformation (Fig. 8c), as expected, the aro-
matic ring of complex 3 was stacked against the imidazole and car-
bonyl moieties of His492, Asp494, His519, His593 and Ala636,
respectively. The phenolic OH group of complex 3 derived from 4-
hydroxyphenethylamine forms one hydrogen bond with the carbox-
ylate group of Asp494. The hydrogen-bonding distance and angle of
Asp494 O⋯H\Ocomplex-3 are 2.422(2)Å and 156.9(2)°, respectively.
In contrast, no hydrogen bond was found between complex 5 derived
from 2-phenylethylamine and the amino acid residues of the urease
active site (Fig. 8d). This probably causes the activity difference of
complexes 3 and 5 as urease inhibitors. In addition, as a result of
the strong intermolecular hydrogen bond formed between the chelat-
ing Schiff base ligand and the active site of the enzyme, the binding
energy (−3.64 kcal/mol) and the lowest intermolecular energy
(−5.84 kcal/mol) in the modeled structure of 3-urease complex are
slightly lower than the corresponding values (−3.36 kcal/mol and
−5.70 kcal/mol) observed in the 5-urease complex. The results fur-
ther demonstrate exceptional difference of urease inhibitory activity
of these copper(II) complexes.
Inhibition of jack bean urease by Schiff base ligands and com-
plexes 1–5.
Tested materials
IC50 (μM)
HL1–4
1
2
3
4
5
>100
2.80
3.22
1.45
52.00
3.59
63.00
Acetohydroxamic acid
of jack bean urease. The optimized cluster (20 occurrences)
was ranked by energy level in the best conformation of the inhibi-
tor–urease modeled structures, where the binding energy of the
amino acid residues with the corresponding copper(II) complexes 1,
2, 3 and 5 shows −7.60 kcal/mol, −6.62 kcal/mol, −3.64 kcal/mol,
−3.36 kcal/mol, and the lowest intermolecular energy presents
−9.79 kcal/mol, −8.81 kcal/mol, −5.84 kcal/mol, −5.70 kcal/mol,
respectively. Besides, some hydrophobic interactions also exist in
the corresponding inhibitor–urease complex.
The binding modes of complexes 1 and 2 in the enzyme active site
were modeled as depicted in Fig. 8a and b. It should be noted that the phe-
nolic O atom of complexes 1 and 2 forms one hydrogen bond with the
side chain N\H of Arg439, respectively. The hydrogen-bonding distance
and angle of Arg439 N\H⋯Ocomplex-1 are 2.652(2)Å and 140.8(2)°,
while that of Arg439 N\H⋯Ocomplex-2 are 2.907(2)Å and 143.0(2)°. This
is due to the presence of a different bromine substituent in the aromatic
ring of 1 and 2 derived from 4-hydroxyphenethylamine. In the inhibi-
tor–urease complex structures, both 1 and 2 form some hydrophobic in-
teractions with the amino acid residues such as Ala440 and Met637,
4. Conclusion
This paper describes the synthesis, crystal structures and urease
inhibitory activities of five new Schiff base copper(II) and nickel(II)
complexes with the bidentate N,O-donor Schiff base ligands. It was
found that this class of complexes exhibited strong inhibitory activity
Fig. 8. Modeled structures of complexes of designed inhibitors 1 (a), 2 (b), 3 (c) and 5 (d) with jack bean urease. Hydrogen bonds are presented as green dotted lines.

X. Dong et al. / Journal of Inorganic Biochemistry 108 (2012) 22–29
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This work was financially supported by the National Natural
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