Structures and esterolytic reactivity of novel binuclear copper(ii) complexes with reduced l-serine Schiff bases as mimic carboxylesterases

Article abstract of DOI:10.1039/d0dt01823f

Three novel binuclear copper(ii) complexes with reduced l-serine Schiff bases were synthesized and their structures were analyzed with single-crystal X-ray diffraction and DFT calculations. The crystal data revealed that all of these binuclear complexes a

Full text of DOI:10.1039/d0dt01823f

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DOI: 10.1039/D0DT01823F
ARTICLE
Structures and Esterolytic Reactivity of Novel Binuclear Copper(II)
Complexes with Reduced L-Serine Schiff Bases as Mimic
Carboxylesterases
Received 00th January 20xx,
Accepted 00th January 20xx
Qin Zhang,a Jun Shu,a Yajun Zhang,b Zhigang Xu,b Jian Yue,a Xiaoqiang Liu, **a Bin Xu,a Zhongzhu Chen
b and Weidong Jiang *a
DOI: 10.1039/x0xx00000x
Three novel binuclear copper (II) complexes with reduced L-serine Schiff bases were synthesized, and their structures were
analyzed with single crystal X-ray diffraction and DFT calculation. Crystal data reveal all of these binuclear complexes are
chiral. Both of 5-halogenated (bromo- and chloro-) binuclear complexes exhibit right-hand helix structural character.
Interestingly, 5-methyl-containing analogue constructs a two-dimensional pore structure. In this paper, esterolysis reactivity
of the as-prepared complexes show that in the hydrolysis of p-nitrophenyl acetate (PNPA) these three complexes provide
26, 18, 40-fold rate acceleration as compared to spontaneous hydrolysis of PNPA at pH 7.0, respectively. Under selected
conditions, in buffered aqueous solution excess three orders of magnitude rate enhancement were observed for the
catalytic hydrolysis of another carboxylic ester, p-nitrophenyl picolinate (PNPP). Furthermore, these complexes efficiently
promote the PNPP hydrolysis in a micellar solution of cetyltrimethylammonium bromide (CTAB), giving rise to excess four
orders of magnitude rate enhancement which is approximate 2.0~3.2 times higher than that in buffer.
towards different substrates including esters,3 amides 4 and plasmid
DNA.5 In general, chemists are eagerly anticipant of the successful
construction of the active centers in which includes three essential
domains (e.g., the binding site, catalytic site and hydrophobic cave) 6
that should be regarded to be typical structural characters of
enzymes.
Introduction
Carboxylesterases (CESs) as B-type esterolases play vital role in some
physiological activity,1 thereby maintaining the normal physiological
function of the body. In addition, natural esterases display high
activity and selectivity due to multiple molecular activation and
specific substrate binding.2 Owing to intrinsic environmental
sensitivity of natural CESs, it was highly limited that the expansion of
the application of CESs from the physiological activities to other
fields. To overcome the above defects, most of scientists devoted to
develop various artificial esterases and evaluated their activities
As for natural metallohydrolases, central metal ions in active
domain play vital role in catalytic hydrolysis reactions.7 That is, at
least four key roles should be considered for central metal 8: (1)
activating substrate; (2) activating nucleophilic reagent; (3) activating
leaving group; (4) stabilizing negative tetrahedron transition state
(TS). Along this line of consideration, a great deal of metal complexes
as enzyme models were designed to simulate those features of
natural CESs.9 As we know, Schiff bases metal complexes with
features of convenient synthesis and structural modification were
widely applied in the fields of biological applications,10 functional
materials11 and artificial metalloenzyme.12 Among these applications
mentioned above, Schiff bases metal complexes were widely
a. School of Chemistry and Environmental Engineering, Sichuan University of Science
& Engineering, Sichuan Zigong 643000, P. R. China
E-mail: jwdxb@suse.edu.cn; xqliu@suse.edu.cn
College of Pharmacy, National & Local Joint Engineering Research Center of
b
Targeted and Innovative Therapeutics, Chongqing Key Laboratory of Kinase
Modulators as Innovative Medicine, Chongqing University of Arts and Sciences,
Chongqing 402160, China.
Electronic Supplementary Information (ESI) available: crystal structures of the two
reduced L-serine Schiff bases and three copper(II) complexes, molecule structures
and topological structures of binuclear copper(II) complexes, representations of the
charge density difference, HOMO and LUMO orbitals for two halogenated
complexes, thermal analysis and electronic spectra of three binuclear copper(II)
complexes used in this work, representative UV-vis absorbance scan for the
hydrolysis of PNPA and PNPP by Cu2L12, ESI-MS spectra for the Cu2L22-promoted
hydrolysis of PNPP or PNPA, Crystal parameters of binuclear copper(II) complexes
with reduced Schiff bases, selected bond lengthen(Å) and angle (deg) of binuclear
copper(II) complexes, Calculated Addison Tau factors (τ) and geometry for the
central copper(II) of binuclear copper(II) complexes, DFT-calculated Mulliken
charges of the two central copper(II) ions of each complexes used here, relative
activities of PNPP hydrolysis between various catalytic systems. Crystallographic
Information Files (CIF) were also provided.
13
selected as mimic hydrolases to hydrolytically cleave the varied
esters, proteins and DNA. And, further elucidations involve in the
effects of various structural modulation strategies (steric hindrance,
electronic effect, rigid-flexible adjustment, etc) on hydrolytic
reactivity.
Owing to good biocompatibility of amino acid Schiff bases and
corresponding metal complexes, and thereby studying their design
and functions has become a hot pot. At present, some amino acid
Schiff base complexes have been evaluated their activities for the
14
hydrolysis of esters besides their biological activity.15 Generally,
See DOI: 10.1039/x0xx00000x
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appropriate rigid-flexibility of the active site is required for native
enzymes to which the substrate could be bound as described in
‘Induce-Fit Hypothesis’ by D. E. Koshland 16 and the developed theory
by C. L. Tsou.17 Though those mimic metallohydrolases with amino
acid Schiff bases exhibited good activities for esterolysis, their rigid
structures probably limit the binding of substrate on the central
metal during the substrate hydrolysis. Taking account into this
reason, herein we decided to synthesize three reduced L-Serine
Schiff bases and their copper (II) complexes with more flexible frame
(Scheme 1). Furthermore, catalytic activities of the as-prepared
complexes as potential artificial metallohydrolases were evaluated
for the hydrolysis of p-nitrophenyl acetate (PNPA). In order to
investigate different substrate selectivity, a similar substrate (p-
nitrophenyl picolinate, PNPP) was used as reference for evaluating
the different selectivity of these complexes under comparable
conditions. With regard to native enzymes, additionally, catalytically
View Article Online
DOI: 10.1039/D0DT01823F
Cu2L1
2
Cu2L2
2
active site and substrate binding domain generally exist inside
18
hydrophobic pocket of enzymes.
This hydrophobic domain is
essential to high activity and selectivity of natural enzymes. To
simulate this feature of enzymes, many researchers have
constructed hydrophobic microenvironment of some artificial
19
esterolase models mainly including metallosurfactants or mixed
20
systems of metal complexes and various micelles. Hence, in the
present work we are intrigued to study a selected micellar effect of
cationic cetyltrimethylammonium bromide (CTAB) on the PNPP
hydrolysis promoted by the as-synthesized binuclear copper(II)
complexes with reduce L-Serine Schiff bases.
Cu2L3
2
Figure 1. Asymmetrical units of the as-synthesized binuclear copper(II) complexes
with reduced L-serine Schiff bases.
‘ONO’ tridentate ligand. Different geometries for these complexes
are obtained. The crystal of Cu2L12 (CCDC No. 2004683) containing 5-
methyl group is orthorhombic with unit-cell parameters: a=7.527,
b=14.125, and c=23.965 Å; space group P212121/c; Z=4. Four complex
Scheme 1. Synthesis route of binuclear copper (II) complexes with reduced L-Serine
molecules are distributed in each crystal cell of Cu2L1 , and the other
2
Schiff Bases.
two complexes display monoclinic crystal system with P21/c space
group with each crystal cell including two complex molecules (left
Results and Discussion
2.1 Structures Analysis of Binuclear Copper Complexes with
Reduced L-Serine Schiff Bases
column of Figure 2). As for Cu2L1 , an asymmetric coordination
sphere was assigned to their two copper centers. Penta-coordinated
2
structure of the Cu(2) cation with slightly distorted rectangular-
21
pyramidal configuration with Addison Tau factor
(τ=0.26) was
In the present work, three binuclear copper(II) complexes were
successfully synthesized with a coordination reaction of the as-
synthesized reduced L-serine Schiff bases and cupric acetate
monohydrate. Scheme 1 shows the synthesis route of desired
binuclear copper(II) complexes. Single crystals of these complexes
were successfully collected and further be structural analysed. Figure
1 depicts the asymmetrical units of three binuclear copper(II)
complexes on the basis of original crystal data shown in Figure S-1.
Corresponding molecule structures and 2D network structures were
given in Figure S-2. Collected crystal data and selected bond
parameters of three complexes were shown in Tables S-1 and S-2,
respectively.
appeared as companying with the binding of one water molecule.
The slightly distorted octahedral configuration feature (τ=0.08) was
assigned to the hexacoordinate Cu(1) cation of Cu2L1 . Interestingly,
2
the 2D staggered network structure (the right graph of Figure S-2A)
of forming a smaller cavity (marked with broken orange-yellow
circle) was constructed via the Cu(1) coordination site filled by
intermolecular interactions with alkoxo oxygen (O7#1) and one
carboxylic oxygen (O8#) of L-serine. In terms of two halogenated
complexes (Cu2L2 : CCDC No. 2004684, Cu2L3 : CCDC No. 2004685),
2
2
one can observe the almost same characteristics for the molecular
structure and crystal packing shown in Figures S-2B and 2C. Namely,
the one Cu(II) ion of them has a slightly distorted tetrahedral
configuration and the other one is square planar, which is confirmed
by τ factors shown in Table S-3. Intriguingly, 1D periodic right-hand
Molecule structures in Figure S-2 reveal that each binuclear
copper(II) complex used is phenoxo-bridged dicopper(II) one with
2 | J. Name., 2012, 00, 1-3
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DOI: 10.1039/D0DT01823F
-4.522eV
-3.961eV
Figure 3. Representations of the charge density difference (left column), HOMO
(central column) and LUMO (right column) orbitals of Cu2L12. Those DFT-calculated
results for Cu2L22 and Cu2L32 are shown in Figure S-3 for clarity.
In addition, the thermal analysis reveals that all of three complexes
possess high thermostability and have no definite melt point (Figure
S-4A-C). Thermogravimetric (TG) curves illustrate that these
complexes undergo gradually thermal decomposition and lose some
small molecules or structural fragments with respective 46.65%,
51.62% and 60.14% of residual mass in the operated temperature
range. As shown in differential thermal gravity (DTG) curves, three
complexes sharply decompose in the range of 225 °C to 260 °C with
Figure 2. Crystal cells (left column) and recorded PXRD (right column) of three
binuclear copper(II) complexes.
respective peak temperature (e.g., 231.89°C for Cu2L1 , 240.36°C for
2
helix structure and 2D supramolecular network texture were Cu2L2 , 246.42°C for Cu2L3 ). Mass losing of 5.54% shall be assigned
2
2
originated from π–π* stacking with approximate 4.0 Å spacing to the departure of two crystal waters of Cu2L22 companying with an
between two benzene rings respectively located in neighboring 1D apparent endothermic peak around 100°C shown in DSC analysis.
helix structures. The Cu-Cu distance of each binuclear Cu(II) complex Also, DSC curve of Cu2L3 shows the losing of two crystal waters
2
is about 3.0Å that is match with the favourable metal-metal distance around 75°C. Combining DSC with TG of Cu2L1 , it was found that
2
of most of natural enzymes. 22 As shown in the right column of Figure there is 3.18% of mass lose but no endothermic signal implying the
2, recorded PXRD patterns were given for three binuclear copper(II) exist of a small amount of adsorbed water in the Cu2L1 sample.
2
complexes. A serial of sharp peaks in a 2θ range of 10° to 50° suggest Overall, thermal analysis shows 5-bromo-substituted Cu2L3
2
a high degree of crystallization for each binuclear copper(II) complex possesses the best thermal stability.
used here.
The UV-vis electronic spectra of these three complexes were
In this work, all the density functional theory (DFT) calculations recorded in a wavelength region of 190 to 600nm (Figure S-4D). It
were carried out by DMol3 code. The generalized gradient was found that each complex has a strong and sharp absorption in
approximation (GGA) with Perdew-Burke- Ernzerhof (PBE) functional the near-UV wavelength range (around 210nm). The electronic
was used to describe the exchange and correlation functionals. In spectra show a single absorption band at 209 nm for Cu2L1 and
2
order to accurately describe noncovalent forces, Grimme is applied Cu2L2 , and 211 nm for bromo-containing Cu2L3 . The absorptive
2
2
to dispersion correction for DFT. For more detailed parameters positions around 210 nm of these bands are typical of π-π* transition
setting of simulation, see the previous work of Liu et al. 23 Both the of multiply substituted benzene ring. Red shift of 2nm on λmax and
charge density difference and the molecular orbital interactions are strengthened absorption were observed in characteristic absorption
respectively illustrated in Figure 3 (for Cu2L1 ) and Figure S-3 (for of Cu2L3 . This is probably contributed to a n-π conjunction
2
2
Cu2L2 and Cu2L3 ), blue area indicates electron accumulation, inspiration and hyperchromicity due to the introducing of bromine
2
2
whereas the yellow region shows the electron depleted. Judging group as auxochrome group. According to Lambert-Beer law,
from the charge density difference, copper atoms in binuclear furthermore, related molar absorption coefficients (ɛ) are
copper(II) complexes are electron-deficient, and corresponding determined and listed in Table 1. The ɛ values show an increased
Mulliken charges (Table S-4) are predicted to be 0.569 ~ 0.573 e for trend along with the introducing of the 5-substituted groups as
Cu2L1 , 0.576 ~ 0.586 e for Cu2L22 and 0.580 ~ 0.581 e for Cu2L3 . To following order: CH3 < Cl < Br. The biggest absorbance with a value of
2
2
obtain a more detailed qualitative description of the binuclear 2.401 was observed for bromo-containing Cu2L32 compared to Cu2L1
copper (II) complexes with reduced L-serine Schiff bases, the HOMO and Cu2L22 at the same sample concentration.
2
and LUMO orbitals of them have been calculated. As can be observed
in Figure 3, the HOMOs are mainly ligand π orbitals for the
centrosymmetric square-plannar of binuclear copper (II) complexes,
whereas the d orbitals of copper (II) ion have small contribution to
orbitals of these complexes. The copper (II) metal centers use dxy
type orbitals, not dx2-y2 orbitals, for a σ∗ interaction with the p orbitals
of N and O atoms.
Table 1 Electronic spectra parameters of three copper(II) complexes at 25°C
Electronic
Catalyst
λmax (nm)
Abs
ɛ (L・ mol-1・cm-1)
Transition
Cu2L1
209
209
1.748
1.976
26220
29640
π→π*
π→π*
2
Cu2L2
2
Cu2L3
211
2.401
36015
π→π*
2
Conditions: 25°C, pH 7.0, Ccomplex =6.67×10-6 mol L-1.
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2.2 Hydrolysis Reactivity and Proposed Mechanisms of Binuclear cause significant difference in activity in spite Voiefw AirntidclievOidnulinael
Copper(II) Complexes Towards Carboxylic Esters
exceptions (Figures 4A’-C’). This result is poDssOibI:l1y0a.1s0c3r9ib/De0dDtTo01w8e2a3Fk
electrostatic and steric effects of these substitutes that influencing
the activity of these binuclear copper(II) complexes. DFT-calculated
Mulliken charges in Table S-4 denote small difference between these
three complexes, which also provides evidence for a neglectable
impact of Lewis acidity on reactivities of these complexes.
In the current work, we mainly focus on the activity of binuclear
copper(II) complexes with reduced L-serine Schiff base in the
hydrolysis of p-nitrophenyl acetate (PNPA) as classic carboxylic ester
model. Besides, another analogue (p-nitrophenyl picolinate, PNPP)
was selected as candidate for comparing the possible selectivity of
catalyst to the two substrates. Figure S-5A shows the spectral change
To evaluate the substrate selectivity of the prepared copper
of the PNPA hydrolysis by Cu2L1 as example in buffered aqueous complexes, herein we compared hydrolytic behaviour of PNPA with
2
solution. The absorption strength at 272nm of substrate PNPA that of PNPP as a contrast substrate (Figure 5) under selected
gradually decreases along with the extension of reaction time. conditions (pH 7.0, Csubstrate=2.0×10-4 mol L-1, Ccomplex= 1×10-5 mol L-1).
Moreover, strengthened absorbance at 400nm indicates the The obtained observations shown in Figure 5A-C reveal that Cu2L12 is
increase in concentration of detectable product, p-nitrophenolate relatively much reactive over the other two halogenated complexes,
anion. The changed trend of UV spectra reveals that the current Cu2L2 and Cu2L3 . In non-micellar solution, moreover, all of
2
2
substrate PNPA is able to be catalytically hydrolysed by Cu2L1 . complexes displayed good reactivity by three orders of magnitude
2
Similar trend of spectral change (Figure S-5B) was observed for the acceleration in comparison with the background rate (k0=7.8×10-6 s-1
Cu2L12 -induced hydrolysis of PNPP as a reference substrate.
25) of PNPP spontaneous hydrolysis in buffered aqueous solution.
Respectively, 6949-, 3862- and 3880-fold rate enhancement was
obtained for the PNPP hydrolysis respectively promoted by Cu2L1 ,
2
Cu2L22 and Cu2L3 .
2
Cu2L12+CTAB
0.200mM
0.200mM
Cu2L22+CTAB
100
50
0
100
50
0
=53.76*10-3s-1
A
B
△
△
×
Cu2L1
△
=65.38*10-3s-1
2
Cu2L2
2
=54.11*10-3s-1
△
=30.55*10-3s-1
CTAB
CTAB
6.99
7.00
pH
7.01
6.99
7.00
pH
7.01
0.200mM
Cu2L32+CTAB
1000
800
600
400
200
0
100
50
0
D
C
=62.82*10-3s-1
△
1
Cu
L
2
Cu2L3
2
2
L
2
2
Cu
2
=30.18*10-3s-1
△
3
L
2
Cu
2
CTAB
6.99
7.00
pH
7.01
PNPP/PNPA
Figure 5. Hydrolysis rates (A-C) of PNPP in different catalytic systems. Figure 5D
shows the relative activity of PNPP against PNPA. Conditions: pH 7.00, 25.0 °C,
[complex]=1×10-5 mol L-1, [PNPA]=2.0×10-4 mol L-1, [PNPP]=2.0×10-4 mol L-1,
[CTAB]=1.0×10-3 mol L-1.
Figure 4. Dependence of the hydrolysis rate constants of PNPA upon pH in the
presence of as-prepared copper(II) complexes (left, A-B), and relative activity
graphs (right, A’-C’). Conditions: 25 °C, I=0.1M KCl, Ccomplex=1×10-5 mol L-1.
With respect to substrate specificity of natural enzymes, it is
mainly contributed to special binding of substrate molecules to
binding sites of enzymes surface.26 According to the hydrolysis
process of esters induced by metallohydrolases, the formation of
In the hydrolytic kinetic determination, the substrate
concentration should be more than 20 times of the catalyst
concentration. As shown in Figures 4A-C, three copper(II) complexes
display gradually increase tendency of kob against pH for the PNPA
hydrolysis. Under a comparable condition (pH 7.0, 25 °C, I=0.1 KCl),
reactive substrate-enzyme binary complex benefits the hydrolysis
27
reaction.
In the current work, each complex exhibited
distinguishable reactivity towards the hydrolysis of PNPA and PNPP
(Figure 5D). The hydrolysis of PNPP is 1047-, 860- and 379-fold faster
than that of PNPA. The intrinsic structure of each substrate should
be the main cause for the different respond to the selectivity of each
catalyst used. With regard to PNPA and PNPP, completely different
mechanisms shown in Figure 6 lead to remarkable difference in
hydrolysis rate. PNPA hydrolysis undergoes an intermolecular
nucleophilic attack of metal-bound OH- on the carbonyl of free PNPA
methyl-substituted Cu2L1 achieves approximately 26-fold rate
2
24
enhancement in contrast with spontaneous hydrolysis rate
of
PNPA. Meanwhile, we observed eighteen and forty-fold rate
acceleration for 5-chloro- or 5-bromo-substituted complexes (Cu2L2 ,
2
Cu2L3 ). As a whole, in the operated ranges of pH and substrate
2
concentration three 5-substituted groups (-CH3, -Cl, -Br) does not
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molecule during which no binary catalyst-PNPA complex forms
(please refer to ESI-MS spectrum shown in Figure S-6). Conversely,
an intramolecular nucleophilic attack was involved in the hydrolysis
of PNPP even though no direct ESI-MS evidence (Figure S-6) for the
formation of catalyst-PNPP intermediate was temporarily obtained
that is probably attributed to highly rapid hydrolysis process of PNPP
in the presence of the as-synthesized complexes. As described in
previous reports,28 the intramolecular mechanism for PNPP
hydrolysis includes the formation of active PNPP-complex
intermediate via a coordination of central Cu(II) ion with pyridine N
atom inside PNPP, which possesses a few advantages concerning
lower energy barrier and closer substrate-to-nucleophile space
against the intermolecular one. Thereupon, PNPP hydrolysis
promoted by each binuclear copper(II) complex achieves much
obvious rate enhancement compared to PNPA hydrolysis. To some
extent this feature provides evidence for the substrate specificity of
natural hydrolases,29 suggesting the artificial enzymes are required
favourable substrate structures.
View Article Online
DOI: 10.1039/D0DT01823F
Scheme 2. Distinguishable distribution (left) of substrate and catalyst in two media,
e.g., buffered aqueous solution and CTAB micellar solution. Right illustration shows
the micellar catalysis hydrolysis of PNPP by the as-prepared complexes.
with buffered aqueous medium. The CTAB-promoted hydrolysis of
PNPP in the absence of binuclear copper(II) complex only gives a
small kob value of 9.0×10-5 s-1 showing undesired rate enhancement
(about 11.5-fold) as compared to PNPP spontaneous hydrolysis.
Table S-5 lists relative activities of PNPP hydrolysis between various
catalytic systems each other, indicating that these three copper(II)
complexes indeed provide dominative contribution to the rate
enhancement of PNPP hydrolysis in contrast with sole CTAB micelles.
32
Obvious acceleration probably results from three major factors
(see Scheme 2) deriving from the intrinsic structural property of
CTAB micelles. First, reactive molecules including hydrophobic PNPP
and complexes are facilely solubilized in the micelle phase of CTAB,
leading to a higher local concentration of reactants 33 in CTAB micelle
phase relative to the lower one in aqueous phase. Second, positive
Intermolecular One
head groups of CTAB micelles provide additional stability of negative
34
tetrahedron transition state.
Owing to higher local OH-
concentration in the positive head groups region of CTAB micelles,
finally, the Cu(II)-bound water undergoes an easier deprotonation to
produce nucleophilic hydroxide species compared to bulk phase.
This experimental phenomenon is in accordance with those previous
reports concerning the increased local basicity due to electrostatic
attraction of positive head groups of cationic micelles.33, 35
Intramolecular One
Conclusions
In our work, three novel binuclear copper(II) complexes with
reduced L-serine Schiff bases possess interesting crystal forms, and
exhibit better reactivity towards the hydrolysis of p-nitrophenyl
acetate (PNPA). Moreover, contrast test was performed for
evaluating activities of these complexes in the hydrolysis of another
analogue, PNPP (p-nitrophenyl picolinate). The observations shown
that PNPP hydrolysis is much faster than the hydrolysis of PNPA,
which is attributed to more beneficial intramolecular nucleophilic
mechanism of PNPP hydrolysis under comparable conditions. The
introducing of CTAB micelles exhibited positive effect on the PNPP
hydrolysis in the presence of the as-prepared complexes, announcing
2~3 folds rate enhancement as compared to that in buffered
aqueous solution.
Figure 6. Proposed mechanism of PNPA (top) and PNPP (bottom) hydrolysis
promoted by Cu2L22 as an example.
The hydrophobic domain of natural enzyme plays a vital role in
catalysis process via hydrophobic and electrostatic interactions.30
‘Hydrophobic pocket’ having low dielectric constants facilitates the
insertion of substrate molecules and the facial generation of active
nucleophile (e.g., OH-). 31 Therefore, in the current work the micellar
effect of traditional surfactant CTAB was investigated for evaluating
possible positive function of CTAB micelles in the PNPP hydrolysis. As
shown in Figure 5A-C, excess four orders of magnitude rate
enhancement achieved for each catalyst (13842-fold for Cu2L1 ,
2
12054-fold for Cu2L2 , 11934-fold for Cu2L3 ) in the CTAB micellar
2
2
Experimental
Chemicals and Instruments
solution in comparison with the auto-hydrolysis of PNPP at pH 7.0,
25°C. Also, for the complex-promoted PNPP hydrolysis the
introducing of CTAB micelles led to 2~3 times acceleration in contrast
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All of reagents were analytical grade and used without further d6): δ7.45 (1H), 7.31 (1H), 6.80 (1H), 3.98-3.96 (2H), 3.76 (1H), 3.65
View Article Online
DOI: 10.1039/D0DT01823F
purification. Potassium chloride as buffered reagent was purchased (1H), 3.25 (1H).
from Sigma-Aldrich Co. Tris-H+ [tris(hydroxymethyl) aminomethane]
Synthesis and single crystals of binuclear copper(II) complexes with
reduced L-serine Schiff bases. Cu2L1 : 0.5mmol of HL1 was dissolved
was purchased from Chengdu Kelong Chemical Co. Surfactant CTAB
and three kinds of 5-substituted salicylaldehydes were products of
2
with super-pure water under boiling condition, and then an aqueous
Beijing J&K Scientific Ltd. The reduced L-serine Schiff bases as desired
solution of 0.6 mmol Cu(OAc)2·H2O was dropwise added into the
ligands and their respective binuclear copper(II) complex were
solution of HL1. Dark green transparent solution was obtained after
synthesized by the method described below. Purified water with high
refluxing for 1h. The dark green filtrate was obtained and cooled to
resistance of 18 MΩ, which is from a Water Purification System (Nex
room temperature. Dark green solids were collected for acting as
Power 1000, Human Corporation, South Korea), was used
catalyst. To determine absolute structure of this complex, dark green
throughout for kinetic determinations. PNPA as aim substrate was
single crystal of Cu2L12 suitable for X-ray diffraction was successfully
provided by our lab, and PNPP as contrast substrate was prepared
obtained by natural diffusion method. Yield of 78.95%. m. p. >450.0
using previous route. 25 Stock solutions of PNPA and PNPP (Cstock=0.02
°C (partial decomposition with about 46.65% residual mass). m/z:
M) were freshly prepared with anhydrous acetonitrile. Hydrolysis
calc. for C22H26Cu2N2O9 589.53; found 590.54 (M+H)+, 602. 53
rates of PNPA or PNPP was determined by monitoring the changing
(M+Na)+. Anal. calcd. For C22H26Cu2N2O9: C 44.82, H 4.45, N 4.75;
concentration of p-nitrophenolate anion as detectable product at
found C 44.81, H 4.46, N 4.74%. Similar preparation was adopted for
400nm on a GBC Cintra 1010 UV-vis spectrometer (Australia GBC
Scientific Equipment Pty Ltd).
green Cu2L22 and Cu2L3 , and their single crystals were also collected
2
for X-ray diffraction determination. Cu2L2 : yield 82.66%. m. p.>450.0
2
The 1H NMR spectrum was recorded at 600 MHz on a Bruker °C (partial decomposition with about approximate 51.62% residual
Avance-600 NMR spectrometer (Bruke Corporation, Germany). Mass mass). m/z: calc. for C20H24Cl2Cu2N2O10 650.41; found 651.41 (M+H)+,
spectra of the three complexes were recorded on AGILENT 1100 LC- 673.42 (M+Na)+. Anal. calcd. For C20H24Cl2Cu2N2O10: C 36.93, H 3.72,
MS (Agilent, USA). Elemental analysis was carried out on CHNOS N 4.31; found C 36.95, H 3.73, N 4.29%. Cu2L3 : yield 88.12%. m. p.
2
Elemental Analyzer Vario EL Cube (Germany Elementar Co.). Melting >450.0 °C (partial decomposition with about 60.14% residual mass).
point determination was conducted on X-5-type Digital Precision m/z: calc. for C20H20Br2Cu2N2O10 735.28; found 736.29 (M+H)+,
Melting Point Apparatus (Beijing Fukai Instrument Co., Ltd., China). 778.28 (M+Na)+. Anal. calcd. For C20H20Br2Cu2N2O10: C 32.67, H 2.74,
Single Crystal X-ray Diffraction was recorded with Bruker Smart- N 3.81; found C 32.66, H 2.76, N 3.80%.
Apex-II X-Ray Single-Crystal Diffractometer (Bruke Corporation,
Germany). Differential scanning calorimetry (DSC) was determined
with DSC200F3 (Netzch, Germany). Thermal analysis (TG-DTG) was
Hydrolysis Kinetics
investigated with Netzsch STA409 PC thermal analyzer (Netzch, Kinetics of p-nitrophenyl acetate (PNPA) hydrolysis promoted by the
Germany). ESI-MS was carried out using Shimadzu LCMS-IT-TOF as-resulting binuclear copper(II) complexes were carried out in a pH
(Shimadzu Co., Kyoto, Japan).
range of 7.00-8.50 under room temperature. The glass vials
containing solution of the buffer, metal complex was thermally
equilibrated in a thermostatic compartment. Hydrolysis of PNPA was
performed by injecting the PNPA stock solution (an initial
concentration of 2.0×10−2 mol L−1) into a 3mL buffer system
containing the complex of desired concentration. PNPA
concentration was added at least 20 folds excess over the catalyst,
resulting in pseudo-first-order kinetics. Pseudo first-order rate
constants (kob) were determined by monitoring the appearance of p-
nitrophenolate anions (NP) as one product at 400nm. For the aim of
comparing substrate selectivity, in addition, this work carried out a
control experiment about the catalytic hydrolysis of another
substrate (PNPP) promoted by the as-prepared complexes at
comparable conditions. Micellar effect of CTAB on the complex-
induced PNPP hydrolysis was evaluated under CTAB concentration
over its critical micelle concentration (CMC=9.2×10-4 mol L-1). Control
run for the PNPP hydrolysis only by CTAB micelles was also
performed. All of data reported here are averages of two or three
runs with uncertainty of less than 5%.
General synthesis of reduced L-serine Schiff bases (HL). L-serine
(7mmol) and sodium hydroxide (7mmol) dissolved in 20mL of
methanol with stirring, gaining colourless and transparent solution.
Then, methanol solution of 5-substituted salicylaldehyde (7mmol)
was dropwise added into methanol solution of L-serine. The reaction
was further maintained for 3 hours under room temperature. The
temperature of yellow reaction residue was controlled at 0-4°C
following by the adding of sodium borohydride (NaBH4, 7mmol) as
reductant. The obtained colourless and transparent solution was
adjusted to approximate pH 5.0-6.0 with glacial acetic acid, gradually
producing amounts of white precipitates. White precipitates were
washed with anhydrous diethyl ether and deionized water for three
times during filtration under reduced pressure. Crude precipitates
were recrystallized with a mixed solvent of anhydrous methanol and
water. Collected white crystals were dried under vacuum at 70°C,
generating desired reduced L-serine Schiff bases as aim products. HL1
(containing 5-methyl one): white crystal. Yield 80.6%. m. p. 193-
1
195°C. H NMR (600MHz, DMSO): δ7.05 (1H), 6.99 (1H), 6.75 (1H),
4.02 (2H), 3.79 (1H), 3.65 (1H), 3.24 (1H), 2.19 (3H). HL2 (containing
5-chloro one): white crystal. Yield 89.2%. m. p. 202.4-204.0°C. 1H
NMR (600 MHz, DMSO-d6): δ7.32 (1H), 7.19 (1H), 6.84 (1H), 3.97
(2H), 3.75 (1H), 3.64 (1H), 3.25 (1H). HL3 (containing 5-bromo one):
white crystal. Yield 78.5%. m.p.195-197°C. 1H NMR (600 MHz, DMSO-
Conflicts of Interest
There are no conflicts to declare.
6 | J. Name., 2012, 00, 1-3
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ARTICLE
Termine, A. Golemme, M. Ghedini, N. Bellec, E. Pini, I. Rimoldi and N.
Godbert, ACS Appl. Mater. Interf., 2015, 7, 4019-4028.
Acknowledgements
View Article Online
DOI: 10.1039/D0DT01823F
This work was supported by National Natural Science Foundation of
China (No. 21771129), Postgraduate Research Project of Chongqing
University of Arts and Sciences (No. M2019 ME09). Authors highly
appreciate Dr. Qianli Li for analyzing crystal structures. The Sichuan
University of Science & Engineering High Performance Computing
Center is acknowledged for providing computational.
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DOI: 10.1039/D0DT01823F
Table of Contents
Structures and Esterolytic Reactivity of Novel Binuclear Copper(II)
Complexes with Reduced L-Serine Schiff Bases as Mimic
Carboxylesterases
Qin Zhang,a Jun Shu,a Yajun Zhang,b Zhigang Xu,b Jian Yue,a Xiaoqiang Liua**, Bin Xu,a Zhongzhu Chenb
and Weidong Jiang a*
Three novel binuclear copper(II) complexes with reduced L-serine Schiff bases exhibited significant
activity difference for the hydrolysis of PNPA and PNPP, suggesting favourable intramolecular
nucleophilic pathway for PNPP. The introducing of CTAB micelles arouses 2.0 ~ 3.2 folds rate
enhancement for PNPP hydrolysis against buffered aqueous solution.
1000
800
600
1
Cu L
2
2
2
400
200
0
Cu L
2
2
3
Cu L
2
2
PNPP/PNPA