Investigations into the bovine serum albumin binding and fluorescence properties of Tb (III) complex of a novel 8-hydroxyquinoline ligand

Article abstract of DOI:10.1016/j.saa.2014.07.089

A novel ligand, 2-methyl-6-(8-quinolinyl)-dicarboxylate pyridine (L), and its corresponding Tb (III) complex, Na₄Tb(L)₂Cl ₄·3H₂O, were successfully prepared and characterized. The luminescence spectra showed that the ligand L was an efficient sensitizer for Tb (III) luminescence. The interaction of the complex with bovine serum albumin (BSA) was investigated through fluorescence spectroscopy under physiological conditions. The Stern-Volmer analysis indicated that the fluorescence quenching was resulted from static mechanism. The binding sites (n) approximated 1.0 and this meant that interaction of Na ₄Tb(L)₂Cl₄·3H₂O with BSA had single binding site. The results showed van der Waals interactions and hydrogen bonds played major roles in the binding reaction. Furthermore, circular dichroism (CD) spectra indicated that the conformation of BSA was changed.

Full text of DOI:10.1016/j.saa.2014.07.089

Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy 135 (2015) 953–958
Contents lists available at ScienceDirect
Spectrochimica Acta Part A: Molecular and
Biomolecular Spectroscopy
journal homepage: www.elsevier.com/locate/saa
Investigations into the bovine serum albumin binding and fluorescence
properties of Tb (III) complex of a novel 8-hydroxyquinoline ligand
Mingming Zhao a,b, Ruiren Tang a, , Shuai Xu a
⇑
a
School of Chemistry and Chemical Engineering, Central South University, Changsha 410083, PR China
Hunan Police Academy, Changsha 410138, PR China
b
h i g h l i g h t s
g r a p h i c a l a b s t r a c t
ꢀ
A novel 8-HQ ligand and its Tb (III)
complex were synthesized and
characterized.
ꢀ
ꢀ
The fluorescence properties of Tb (III)
complex was studied.
The binding characteristics of
Na
4
Tb(L)
2
Cl
4
2
ꢁ3H O with BSA was
investigated.
ꢀ
Investigating the structural changes
of BSA by CD spectra.
a r t i c l e i n f o
a b s t r a c t
Article history:
Received 19 April 2014
Received in revised form 12 July 2014
Accepted 29 July 2014
Available online 9 August 2014
A novel ligand, 2-methyl-6-(8-quinolinyl)-dicarboxylate pyridine (L), and its corresponding Tb (III) com-
plex, Na
4
Tb(L)
2
Cl
4
ꢁ3H
2
O, were successfully prepared and characterized. The luminescence spectra
showed that the ligand L was an efficient sensitizer for Tb (III) luminescence. The interaction of the com-
plex with bovine serum albumin (BSA) was investigated through fluorescence spectroscopy under phys-
iological conditions. The Stern–Volmer analysis indicated that the fluorescence quenching was resulted
from static mechanism. The binding sites (n) approximated 1.0 and this meant that interaction of
Keywords:
Na
4
Tb(L)
2
Cl
4
ꢁ3H
2
O with BSA had single binding site. The results showed van der Waals interactions
8
-Hydroxyquinoline
and hydrogen bonds played major roles in the binding reaction. Furthermore, circular dichroism (CD)
spectra indicated that the conformation of BSA was changed.
Tb (III) complex
Fluorescence property
BSA binding study
Ó 2014 Elsevier B.V. All rights reserved.
Circular dichroism spectroscopy
Introduction
earth ions (RE(III)) are weak since intra-configuration 4f–4f transi-
tions in trivalent lanthanide ions are forbidden (Laporte rule). In
Since a brightly photoluminescent lanthanide complex was first
reported by Weissman in 1942 [1], the properties of lanthanide
organic complexes had been extensively studied. Within recently
years, lanthanide complexes have enormous applications in many
areas, such as chemical, medical, and sensor application [2,3]. The
absorption and emission spectra intensity of the trivalent rare
order to overcome this drawback and obtain outstanding lumines-
cent properties, an organic ligand with excellent absorption coeffi-
cient and high efficient ligand-to-RE(III) ions energy transfers,
which acts as an ‘‘antenna’’ [4], is very necessary. Based on differ-
ent ligands and the central RE(III), many novel ligands and com-
plexes have been designed and synthesized [5]. Some of that
have excellent lanthanide binding capacity and antenna effects.
8
-Hydroxyquinoline (8-HQ) is one of the most important chelates
⇑
Corresponding author. Tel.: +86 731 88836961; fax: +86 731 88879616.
E-mail address: trr@mail.csu.edu.cn (R. Tang).
for metal ions and has significant applications in a variety of
http://dx.doi.org/10.1016/j.saa.2014.07.089
386-1425/Ó 2014 Elsevier B.V. All rights reserved.
1

9
54
M. Zhao et al. / Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy 135 (2015) 953–958
investigations involving metal complexes [6]. It can be matched
the first main group of metal ions, transition metal ions, rare earth
metal ions [7–9]. 8-HQ has a great conjugate structure so that it
can be part of good antenna ligand [10]. It has been used to build
lots of highly sensitive fluorescent chemosensors and photoelectric
materials [11–13]. On the other hand, 2,6-dipicolinic acid (DPA) is
an excellent lanthanide binding substance whose N and O atoms
have strong coordination ability in coordination reaction [14,15].
Meanwhile, it possesses biological activity in the body of the ani-
mal and plant [16–18]. Its derivatives and complexes are studied
extensively and used in all sorts of fields [19]. Taking into account
the above advantages of 8-hydroxyquinoline and 2,6-dipicolinic
acid, we designed and synthesized a new ligand combining 8-HQ
ligand and DPA moieties, expecting it has excellent lanthanide
binding capacity and antenna effects.
spectra were measured with a Bruker-400 MHz nuclear magnetic
resonance spectrometer with CDCl
reference.
3
as solvent and TMS as internal
Luminescence measurements were made on a Hitich F-4500
spectrophotometer, the widths of both the excitation and emission
slit were set to 5 nm with the photomultiplier tube voltage at
700 V. Thermal gravimetric analysis (TGA) were performed in the
nitrogen atmosphere using a Netzsch TG 209 thermal gravimetric
ꢃ1
analyzer at a heating rate of 10 °C min from 25 to 750 °C. CD
measurements were carried out on a J-810 spectropolarimeter
(Jasco, Tokyo, Japan) in a cell of path length 1.0 mm at room
temperature.
Preparation of ligand (L)
As one of the most abundant carrier proteins, serum albumin
plays an important role in the transport and disposition of endog-
enous and exogenous ligands present in blood [20,21]. Recently,
the interactions of serum albumin with lanthanide complexes have
arose much interest owing to their applications in a great deal of
medical and chiroptical systems [22]. So, it is very meaningful to
Synthesis of 6-(methoxycarbonyl)-pyridine-2-carboxylic acid (2)
A solution of (1) (5.85 g, 30.0 mmol) in methanol (150 mL) was
cooled to 0 °C. After KOH pellets (1.76 g, 31.0 mmol) were added,
the reaction mixture was stirred at 0 °C for 2 h and then at room
temperature for 24 h. The solvent was removed under reduced
2
pressure, and the residue was suspended in H O (100 mL) and
explore the action mode between Na
serum albumin (BSA), which can provide a theoretical basis for
their potential medicinal value.
4
Tb(L)
2
Cl
4
ꢁ3H
2
O and bovine
extracted with ethyl acetate (3 ꢂ 30 mL). The aqueous layers were
acidified to pH 3 with 1 M diluted HCl solution and extracted with
chloroform (5 ꢂ 30 mL). The collected organic layers were dried
Here, in order to provide a clue for drug design and pharmaceu-
tical research in this work, the novel ligand of 2-methyl-6-(8-quin-
olinyl)-dicarboxylate pyridine (L) and its Tb (III) complex were
designed and synthesized (Scheme 1). The fluorescence property
of the complex was researched in detail. Furthermore, the interac-
tions of the complex with BSA were investigated through fluores-
cence quenching, binding sites, binding mode, CD spectra under
physiological conditions, etc.
2 4
over anhydrous Na SO . The chloroform was removed in vacuum
to provide the desired product as a white solid (2.80 g, yield:
1
5
3
2%). m.p. 144–146 °C; H NMR (400 MHz, CDCl ): d 8.45 (m, 2H,
Py-3, 5), 8.18 (t, J = 6.50 Hz, 1H, Py-4), 3.93 ppm (s, 3H, CH
3
); IR
ꢃ1
(
KBr), v/cm : 3073, 2964, 2852, 1725, 1581, 1325.
Synthesis of 2-methyl-6-(8-quinolinyl)-dicarboxylate pyridine (L)
-Hydroxyquinoline (8-HQ) (1.6 g, 11 mmol) and (2) (2.0 g,
1 mmol) were dissolved in 60 mL of CH Cl (CH Cl , dried by
) at room temperature. To this stirring solution was slowly
added DMAP (0.27 g, 3 mmol) and 20 mL of a solution of 2.53 g
Cl under dried condition at room
8
1
2
2
2
2
Experimental
2 5
P O
Materials
(13 mmol) of EDCꢁHCl in CH
2
2
temperature. After 48 h of stirring, organic phase was washed by
water (3 ꢂ 30 ml) to clear out EDCꢁHCl and DMAP. A white precip-
itate was obtained by evaporated under pressure. The grey solid
was recrystallized over AcOEt and petroleum ether. The white
8
-Hydroxyquinoline (8-HQ) was purchased from Beijing (China)
Medicine Co. Ltd.; BSA, obtained from Sigma Chemical Co. Ltd., was
dissolved in 0.1 M Tris–HCl buffer solution (pH = 7.40, 50 mM
ꢃ5
NaCl). BSA stock solution (1.0 ꢂ 10 mol/L) was kept in the fridge
product was recrystallized over acetone and H
compound (L) (1.90 g, yield:56%). m.p. 157–159 °C; H NMR
(400 MHz, CDCl ): d 8.88(d, J = 3.60 Hz, 1H, 8-HQ-2), 8.62 (m, 2H,
Py-3, 5), 8.25 (t, J = 6.50 Hz, 1H, Py-4), 8.11–7.46 (m, 5H, 8-HQ-
2
O to give the title
1
at 0–4 °C. And other chemicals were of A.R. grade without further
purification. Doubly distilled and deionized water were used in
the whole experiments.
3
1
3
Melting points were determined on a XR-4 apparatus (ther-
mometer uncorrected); Elemental analysis was carried out by a
Perkin Elmer 2400 elemental analyzer; Infrared spectra were
recorded on a Nicolet NEXUS 670 FT-IR spectrophotometer using
3,4,5,2,7), 4.06 ppm (s, 3H, CH
3 3
); C NMR(400 MHz, CDCl ): d
52.9 (CH ); 113.4, 118.9, 123.1, 127.7, 128.8, 128.9, 131.1, 137.3,
3
138.8, 141.2, 149.0, 149.0, 151.7, 154.2 (all Ph-C and Py-C), 158.7
ꢃ1
(COO-8-HQ), 167.0 ppm (COOCH
3
); IR (KBr), v/cm : 3064, 2955,
KBr discs in the 400–4000 cm region; 1H NMR and C NMR
ꢃ1
13
1737, 1575, 1232, 1113.
Scheme 1. The synthetic route of ligand L.

M. Zhao et al. / Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy 135 (2015) 953–958
955
Synthesis of Tb (III) complex
The complex Na Tb(L) Cl
steps. A solution of L (1 mmol) in ethanol (10 ml) was added drop-
wise to the solution of TbCl (0.5 mmol) in ethanol (10 ml) under
4
2
4
2
ꢁ3H O was prepared in the following
3
stirring at 60 °C for 24 h. Then the pH value of the mixture was
adjusted to 6 by adding an aqueous solution of sodium hydroxide
(
1 mol/L). The yellow precipitate was collected by filtration,
washed three times with ethanol and chloroform mixture (1:1,
v:v), dried in vacuum at 60 °C one day.
Results and discussion
Composition and properties of the complex
Elemental analytical
The elemental analytical data for the complex is presented in
Table 1, which is in good agreement with the values calculated
from the molecular formula, Na
4
Tb(L)
2
Cl
4
ꢁ3H
2
O. The complex is
Fig. 1. IR Spectrum of ligand L and Tb (III) complex. A: L, B: Na
4 2
Tb(L) Cl
4
ꢁ3H
2
O.
yellowish powder, stable under atmospheric condition and soluble
in DMF, DMSO and methanol, a little soluble in H
insoluble in benzene, diethyl ether and toluene.
2
O and acetone,
involved with the coordination, and the chemical structure of com-
plex may be as shown in Scheme 2.
FT-IR spectra
Fluorescence properties of complex
The FT-IR spectra and characteristic absorption bands of the
free ligand L and its Tb (III) complex are shown in Fig. 1 and Table 2.
The excitation and emission spectra of the Tb (III) complex is
ꢃ4
ꢃ1
examined in C
2
H
5
OH solution (c = 2.0 ꢂ 10 mol/L) at ambient
The FT-IR spectra of the free ligand L shows bands 1738 cm
,
ꢃ1
ꢃ1
ꢃ1
temperature (Fig. 2). The luminescence data for the Tb (III) com-
1
579 cm , 1500 cm , 1231 cm , which can be assigned to
(C@O), Py(C@N), 8-HQ(C@N) and (CAN) of the ligand, respec-
(C@O) absorption bands in the ligand L are coincided
so that only one (C@O) absorption bands in the FT-IR spectra. In
plex are listed in Table 4. The maximum excitation wavelength
m
m
m
m
*
of the complex Na
sition centered at the ligand L. The complex displays the character-
4
Tb(L)
2
Cl
4
ꢁ3H
2
O is 277 nm, due to the
p
–
p
tran-
tively. Two
m
m
istic line emission of 4f–4f transition of Tb (III) ion, consisting of
comparison with the ligand, the FT-IR spectra of the Tb (III) com-
plex are changed greatly, the band
ligand disappears and the new band appears at 1625 cm . The
absorption band assign to the coordinated
around 489 cm . These evidences clearly show that two carbonyl
groups participate in coordination reaction. The bands
5
7
5
7
ꢃ1
four main lines at 490 nm ( D
4
5
? F
6
), 545 nm ( D
4
? F
5
),
m
(C@O) at 1738 cm in free
5
7
7
ꢃ1
585 nm ( D
4
? F
4
) and 621 nm ( D
4
? F
3
), and the emission
band at 545 nm is obviously stronger than the other emission
bands [23]. Still, each emission band is very narrow and the width
of half band is about several nanometers, indicating that the com-
plex has high color purity and the ligand is a comparative good
organic chelator to sensitize fluorescence of Tb (III) ion [24].
m(TbAO) is found at
ꢃ1
m
Py (C@N),
ꢃ1
m
8-HQ(C@N) and
m
(CAN) peak are slightly shifted to 1568 cm
,
ꢃ1
ꢃ1
1
486 cm , 1322 cm for the complex Na
4
Tb(L)
2
Cl
4
ꢁ3H
2
O. These
indicate that pyridine and 8-hydroxyquinoline nitrogen atoms par-
Interactions with BSA
Fluorescence spectrum
ticipate in coordination reaction. The band v(OAH) vibration at
ꢃ1
3
414 cm , is relatively intense and can be assigned to solvated
water.
ꢃ5
A 2.5 mL solution containing 1.0 ꢂ 10 mol/L BSA was titrated
ꢃ3
by successive addition of 1.0 ꢂ 10 mol/L stock solution of
Thermal gravimetric analysis
Na
4
Tb(L)
2
Cl
4
ꢁ3H
2
O and the concentration of it varied from 0 to
The thermal gravimetric analysis of the complex is given in
Fig. S1. (Fig. S1–S4 are shown in support information). There are
two main successive mass loss stages in the TGA curves. The first
stage of the decomposition appears within 30–170 °C correspond-
ing to the loss of the solvated water (Weight loss: 5.25%, Calcd:
ꢃ4
1
.4 ꢂ 10 mol/L. Titrations were done manually by using micro-
injector. An excitation wavelength (Ex) of 280 nm was chosen in
the experiment. The excitation and emission slit widths (5 nm
each) and scan rate (240 nm/min) were constantly maintained
for all experiments. The fluorescence spectra were recorded at
4
.92%). And the second loss stage ranging from 250 to 450 °C is
2
98 K, 308 K and 318 K in the range of 300–440 nm, respectively.
attributed to the loss of L (Weight loss: 57.68%, Calcd: 56.20%),
which is confirmed by comparing the observed and the calculated
mass of the complex. The detailed data are listed in Table 3. Fur-
thermore, the TGA proves that it has the relatively high thermal
stability.
The temperatures of the samples were kept by recycled water
throughout the experiment.
Analysis of fluorescence quenching of BSA by the complex
The effect of Na
4
Tb(L)
2
Cl
4
ꢁ3H
2
O on the fluorescence emission
The results of EA, TGA and FT-IR spectroscopy indicate that the
ligand L have two carbonyl groups and two nitrogen atom what
spectra of BSA (PH = 7.40) at 298 K is shown in Fig. 3, and the cor-
responding calculated results are given in Table 5. As shown in
Fig. 3, the natural fluorescence of BSA at around 340 nm is gradu-
ally quenched by the increasing concentration of the complex,
Table 1
Result of elemental analysis data for Tb (III) complex.
while the complex Na
in this range. Results above indicate that strong interaction exist
between Na Tb(L) Cl O and BSA, and the microenvironment
4
Tb(L)
2
Cl
4
ꢁ3H
2
O has no intrinsic fluorescence
Complex
C, H, N Found (calc.)
C (%)
4
2
4
ꢁ3H
2
H (%)
N (%)
around the chromophore of BSA is also changed [25]. In order to
shed light on the fluorescence quenching mechanism, the
Na
4 2
Tb(L) Cl
4
ꢁ3H
2
O
37.36 (37.22)
2.80 (2.73)
5.07 (5.11)

9
56
M. Zhao et al. / Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy 135 (2015) 953–958
Table 2
Comparison between IR spectra of ligand L and complex.
Complex
m
(OH)
m
(C@O)
1738
3414 1625
mPy(C@N) m8-HQ(C@N) m(CAN) m(ReAO)
L
1579
1568
1500
1486
1231
1322
Na
4
Tb(L)
2
Cl
4
ꢁ3H
2
O
489
Table 3
TGA data of Na
4
Tb(L)
2
Cl
4
ꢁ3H
2
O.
Complex
Stage Temperature
Mass loss (%)
found (calc.)
Probable lost
molecules
range (°C)
Na
4
Tb(L) Cl
2 4
ꢁ3H
2
O
I
II
30–170
250–450
5.25 (4.92)
57.68 (56.20)
2
3H O
2L
Fig. 3. The quenching effect of Na
4
Tb(L)
2
Cl
4
ꢁ3H
2
O on fluorescence intensity of BSA
ꢃ5
ꢃ5
at 298 K; kex = 280 nm; c(BSA) = 1.0 ꢂ 10 mol/L; c(L)/(10 mol/L). (a–j): 0, 1, 2, 3,
4, 6, 8, 10, 12 and 14, respectively.
Table 5
Fluorescence quenching constants of complex-BSA at three different temperatures.
Complex
T
K
sv
k
q
aR2
4
ꢃ1
12
ꢃ1 ꢃ1
(
K)
(ꢂ10 L mol
)
(ꢂ10 L mol
S )
Na
4
Tb(L)
2
4
Cl ꢁ3H
2
O
298 1.05
1.05
0.88
0.72
0.99607
0.99806
0.99646
3
3
08 0.88
18 0.72
a
R is the correlation coefficient of Stern–Volmer equation.
fluorescence quenching data are analyzed by the Stern–Volmer
equation (Eq. (1)) [26]. The graphs plotted for F /F against [Q] from
Eq. (1) at different temperatures are shown in Fig. S2.
Scheme 2. The chemical structure of Na
4
Tb(L)
Cl
2 4
ꢁ3H
2
O.
0
F
F
0
¼
1 þ K
sv
½Qꢄ ¼ 1 þ
s
0
k
q
½Qꢄ
ð1Þ
where F and F are the fluorescence intensities in the absence and
0
presence of quencher, respectively, sv is the Sterne–Volmer
K
quenching constant, [Q] is the concentration of quencher, is the
s
0
ꢃ8
average fluorescence lifetime of biomolecule at about 10 s [27].
is the biomolecule quenching rate constant and the value of
the maximum scattering collision quenching constant is
k
q
1
0
ꢃ1 ꢃ1
2
.0 ꢂ 10 L mol
S
[28]. The results show the value of k
Tb(L) Cl O, which is
ꢁ3H
. The slope of the Sterne–Volmer
q
is the
rate constants of BSA quench by Na
4
2
4
2
1
0
ꢃ1 ꢃ1
greater than 2.0 ꢂ 10 L mol
S
4
3
3
quenching constants (Ksv
)
are 1.05 ꢂ 10 , 8.8 ꢂ 10 , 7.2 ꢂ 10
ꢃ1
(
L mol ) at 298 K, 308 K and 318 K, which inversely correlated
with the temperature. These evidences indicate that the main
quenching mechanism of BSA by the Na
a static quenching procedure [29].
4
Tb(L)
2
Cl
4
2
ꢁ3H O should be
Binding constant and binding sites
Fig. 2. Excitation (left) and emission (right) spectra of Na
4
Tb(L)
Cl
2 4
ꢁ3H
2
O
The relationship between fluorescence intensity and the con-
centration of quenchers can be described by the following Eq. (2)
(
c = 2.0 ꢂ 10ꢃ4 mol/L, C
2 5
H OH).
a
for static quenching, the apparent K is the binding constant and
n is the number of binding sites per BSA can be expressed [30]:
Table 4
Fluorescence data for Tb (III) complex.
F
0
ꢃ F
log
¼ log K
a
þ n log½Qꢄ
ð2Þ
F
Complex
Na Tb(L)
k
ex (nm)
k
em (nm)
F
Assignment
Fig. S3 shows the double-logarithm algorithm curve, the results of
a
K and n are listed in Table 6. It is noticed that the binding constant
5
5
5
5
7
4
Cl
2 4
ꢁ3H
2
O
277
490
3242
4670
634
D
D
D
D
4
4
4
4
– F
6
5
4
3
7
4
5
5
6
45
85
21
– F
values are equal to 10 and decrease with the increase of tempera-
ture due to reduction of the stability of Na Tb(L) Cl O-BSA sys-
ꢁ3H
tem. On the other hand, the correlation coefficient R is about 0.995,
7
– F
7
4
2
4
2
147
– F

M. Zhao et al. / Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy 135 (2015) 953–958
957
Table 6
The binding constant (K
a
), binding sites (n) and relative thermodynamic parameters for the Tb (III) complex with BSA at three different temperatures.
ꢃ1
bR2
ꢃ1
ꢃ1 ꢃ1
ꢃ1
cR2
Complex
T (K)
K
a
(L mol
)
n
D
H (kJ mol
)
D
S (J mol
K
)
D
G (kJ mol
)
4
Na
4 2
Tb(L) Cl
4
ꢁ3H
2
O
298
3.5 ꢂ 10
2.5 ꢂ 10
1.9 ꢂ 10
1.18
1.12
1.08
0.99861
0.99945
0.99088
ꢃ20.65
ꢃ20.56
ꢃ20.47
4
4
3
08
18
ꢃ23.26
ꢃ8.76
0.99521
3
b
c
R2 is the correlation coefficient of double-logarithm equation.
2
R
is the correlation coefficient of the value of DH, DS.
which proves that the interaction between the complex and BSA is
in accordance with the site-binding model underlined in the above
equation. The above results clearly indicates that Na
4
Tb(L)
2
Cl
4
ꢁ3H
2
O
quenches the fluorescence emission of BSA with higher binding
affinity and experiences a static quenching process. The binding
capacity (n) varies in the range from 1.08 to 1.18, therefore that
the number of binding sites approximates 1.0 and the interaction
of Na
4
Tb(L)
2
Cl
4
2
ꢁ3H O with BSA has single binding site.
Thermodynamic parameters and binding mode
In general, the working forces between the proteins and ligands
mainly include four types: hydrophobic force, electrostatic interac-
tions, van der Waals interactions and hydrogen bonds. To illustrate
the interaction of L with BSA, the thermodynamic parameters (
S) are calculated from the Van’t Hoff equation at three different
temperatures [30].
DH,
D
D
RT
H
DS
R
ln K
a
¼ ꢃ
þ
ð3Þ
is the effective binding constant at the corresponding
Fig. 4. CD spectra of the BSA–Na
4
Tb(L)
2
Cl
4
ꢁ3H
2
O system at pH 7.40, 298 K. (A)
ꢃ6
ꢃ6
ꢃ6
where K
a
5.0 ꢂ 10 mol/L BSA; (B) 5.0 ꢂ 10 mol/L BSA and 5.0 ꢂ 10 mol/L
Na
4
Tb(L)
2
Cl
4
ꢁ3H
2
O.
temperature and R is the gas constant. The calculation is carried
out at three different temperatures, which are 298 K, 308 K, and
3
18 K. The enthalpy change (
DH) is calculated from the slope of
a
%helix ¼ ½ðꢃMRE208 ꢃ 4000Þ=33000 ꢃ 4000ꢄ ꢂ 100
where MRE208 is the MRE value observed at 208 nm, 4000 is the MRE
ð6Þ
the van’t Hoff relationship. The free energy change (
DG) is then esti-
mated from the following relationship:
of the b-form and random coil conformation cross at 208 nm, and
D
G ¼
D
H ꢃ T
D
S
ð4Þ
DH
33,000 is the MRE value of a pure
Fig. 4 shows the CD spectra of BSA and BSA–Tb (III) complex,
which characteristic features of the typical ( + b) helix structure
with negative bands at 208 and 223 nm. The binding of Na Tb(L)2-
Cl O to BSA cause a decrease in band intensity without any
ꢁ3H
significant shift of the peaks, indicating a decrease of the -helical
content in protein [32]. Using the Eq. (5) and (6), the calculated
results exhibit a reduction of -helix structures from 60.53% to
7.17% at Na Tb(L) Cl O /BSA molar ratio of 1:1. From the
ꢁ3H
above results, it is apparent that the effect of Na Tb(L) Cl
ꢁ3H
on BSA causes a conformational change of the protein, with the loss
of -helical stability.
a-helix at 208 nm [34].
Fig. S4, by plotting the data in Table 6, shows the values of
and S obtained for the binding site from the slopes and ordinates
at the origin of the fitted lines. When H < 0 or S > 0, the
H ꢅ 0,
main acting force is electrostatic force; when H < 0, S < 0, the
main acting force is van der Waals interactions and hydrogen
bonds; when H > 0, S > 0, the main force is hydrophobic force
31]. The negative enthalpy ( H) and entropy ( S) values of the
interaction between Na Tb(L) Cl O and BSA indicate that van
ꢁ3H
a
D
4
D
D
D
D
D
4
2
a
D
D
a
[
D
D
5
4
2
4
2
4
2
4
2
4
2
4
2
O
der Waals interactions and hydrogen bonds play major roles in
the binding reaction. Furthermore, the negative free energy change
D
a
G and DH suggests the binding process is exothermic and
spontaneous.
Conclusions
Circular dichroism spectroscopy
As CD spectra is one of the sensitive methods for monitoring the
conformational changes in the protein [32]. The induced ellipticity
is obtained by the ellipticity of the Na
subtracting the ellipticity of Na Tb(L)
length and expressed in degrees. To quantify the results, the mean
residue ellipticity (MRE) in deg cm /dmol , and the
tents of free and bound BSA are calculated using the following
respective equations [33]:
In summary, a novel ligand, 2-methyl-6-(8-quinolinyl)-dicar-
boxylate pyridine (L), and its corresponding Tb (III) complex were
1
13
synthesized and characterized by EA, H NMR, C NMR, FT-IR and
TGA. The study of their luminescence properties showed the Tb
4
Tb(L)
2
Cl
4
ꢁ3H
2
O–BSA mixture
4
2
Cl
4
ꢁ3H
2
O at the same wave-
(
III) complex could be sensitized efficiently by the ligand. The
interactions of Na Tb(L) Cl O with BSA were investigated
4
2
4
ꢁ3H
2
2
ꢃ1
a-helix con-
through the fluorescence spectroscopy under physiological condi-
tions. The probable quenching mechanism of the fluorescence of
BSA with Tb (III) complex was static quenching and the single
binding site. Van der Waals interactions and hydrogen bonds
played major roles in the binding reaction. The CD spectra indi-
MRE ¼ h=C
p
nl10
ð5Þ
where h is the observed dichroism in millidegree, C
concentration of the BSA, n is the number of amino acid residues
in protein (583) and l is the optical length of the cell (0.1 cm).The
p
is the molar
cated that the interaction of Na
4
Tb(L)
2
Cl
4
ꢁ3H
2
O with BSA caused
conformational change of BSA. These results give important insight
into interactions of the rare earth complexes and BSA, which shows
great reference value for research and application in drug design
and pharmacodynamic fields.
a-helical content of BSA is calculated from the MRE value at
2
08 nm using the equation:

9
58
M. Zhao et al. / Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy 135 (2015) 953–958
[
[
14] L.Q. Shen, Z.F. Yang, R.R. Tang, Spectrochim. Acta A 98 (2012) 170–177.
15] Y.M. Luo, J. Li, L.X. Xiao, R.R. Tang, X.C. Tang, Spectrochim. Acta A 72 (2009)
03–708.
Acknowledgement
7
Financial support from the National Natural Science Foundation
of China (No. 21071152, China) is gratefully acknowledged.
[16] X.C. Su, H.B. Liang, S. Simonsen, J. Am. Chem. Soc. 130 (2008) 10486–10487.
[17] U. Karhunen, J. Rosenberg, U. Lamminmaki, T. Soukka, Anal. Chem. 83 (2011)
9011–9016.
[
18] Q.F. Li, Y. Yang, A. Maleckis, G. Otting, X.C. Su, Chem. Commun. 48 (2012)
2704–2706.
Appendix A. Supplementary material
[
[
[
19] S.S. Krishnmurthy, S. Soundarajan, Z. Anorg, Allg. Chem. 348 (1996) 309–312.
20] D.C. Carter, J.X. Ho, Adv. Protein Chem. 45 (1994) 153–203.
21] L.A. MacManus-Spencer, M.L. Tse, P.C. Hebert, H.N. Bischel, R.G. Luthy, Anal.
Chem. 82 (2010) 974–981.
22] G. Zolese, G. Falcioni, E. Bertoli, R. Wozniak, Z. Wypych, E. Gratton, A.
Ambrosini, Proteins 40 (2000) 39–48.
Supplementary data associated with this article can be found, in
the online version, at http://dx.doi.org/10.1016/j.saa.2014.07.089.
[
[
References
23] T.Y. Zhu, K. Ikarashi, T. Ishigaki, K. Uematsu, K. Toda, H. Okawa, M. Sato, Inorg.
Chim. Acta 362 (2009) 3407–3414.
[
[
1] S.I. Weissman, J. Chem. Phys. 10 (1942) 214–217.
2] V. Tsaryuk, V. Zolin, K. Zhuravlev, V. Kudryashova, J. Legendziewicz, R. Szostak,
J. Alloys Compd. 451 (2008) 153–157.
[24] Z.F. Zhang, R.R. Tang, J. Mol. Struct. 1010 (2012) 116–122.
[25] T.H. Wang, Z.M. Zhao, L. Zhang, L. Ji, J. Mol. Struct. 937 (2009) 65–69.
[26] Y.C. Liu, Z.Y. Yang, J. Organomet. Chem. 694 (2009) 3091–3101.
[27] N. Zhou, Y.Z. Liang, P. Wang, J. Photochem. Photobiol. A 185 (2007) 271–
276.
[28] W.R. Ware, J. Phys. Chem. 66 (1962) 455–458.
[29] X.J. Guo, X.W. Han, J. Tong, C. Guo, W.F. Yang, J.F. Zhu, B. Fu, J. Mol. Struct. 966
(2010) 129–135.
[
3] R.X. Ma, Z.M. Chen, Z.H. Gao, R.F. Wang, J.J. Zhang, Synth. Met. 159 (2009)
1
272–1276.
[
[
4] E. Niyama, H.F. Brito, M. Cremona, Spectrochim. Acta A 61 (2005) 2643–2649.
5] C.M.G. dos Santos, A.J. Harte, S.J. Quinn, T. Gunnlaugsson, Coordin. Chem. Rev.
2
52 (2008) 2512–2527.
[
[
[
6] M. Iwamuro, Y. Wada, T. Kitamura, T.J. Adachi, Chem. Lett. 28 (1999) 539–540.
7] C. Schmitz, H.W. Schmidt, M.J. Thelakkat, Chem. Mater. 12 (2000) 3012–3019.
8] E. Garribba, G. Micera, D. Sanna, E. Lodyga-Chruscinska, Inorg. Chim. Acta 348
[30] Y.J. Hu, O.Y. Yu, A.M. Bai, R.M. Zhao, Y. Liu, Biol. Trace Elem. Res. 136 (2010) 8–
10.
[31] Y.T. Sun, H.T. Zhang, Y. Sun, Y.P. Zhang, H. Liu, J.H. Cheng, S.Y. Bi, H.Q. Zhang, J.
Lumin. 130 (2010) 270–279.
[32] H. Gao, L.D. Lei, J.Q. Liu, Q. Kong, X.G. Chen, Z.D. Hu, J. Photochem. Photobiol. A
167 (2004) 213–221.
(
2003) 97–106.
[
9] R.I.R. Blyth, J. Thompson, Y. Zou, R. Fink, E. Umbach, G. Gigli, R. Cingolani,
Synth. Met. 139 (2003) 207–213.
[
[
[
[
10] H.B. Xu, J. Li, L.X. Shi, Z.N. Chen, Dalton Trans. 40 (2011) 5549–5556.
11] M.D. Shults, B. Imperiali, J. Am. Chem. Soc. 125 (2003) 14248–14249.
12] S.Y. Moon, N.R. Cha, Y.H. Kim, S.K. Chang, J. Org. Chem. 69 (2004) 181–183.
13] H. Zhang, L.F. Han, K.A. Zachariasse, Y.B. Jiang, Org. Lett. 19 (2005) 4217–4220.
[33] Y. Li, W. He, J. Liu, F. Sheng, Z. Hu, X. Chen, Biochim. Biophys. Acta 1722 (2005)
15–21.
[34] Z.X. Lu, T. Cui, Q.L. Shi, First ed., Science Press, Beijing, 1987.