Directly oxidized chemiluminescence of 2-substituted-4,5-di (2-furyl)-1H -imidazole by acidic potassium permanganate and its analytical application for determination of albumin

Article abstract of DOI:10.1007/s10895-011-0849-8

In the paper, 2,4,5-tri(2-furyl)-1H-imidazole (TFI) and 2-phenyl-4,5-di(2-furyl)-1H-imidazole (PDFI), were chosen to investigate chemiluminescence (CL) properties of 2-substituted-4,5-di(2-furyl)-1H- imidazoles. The directly oxidized CL of analytes by potassium permanganate (KMnO₄) was in detail studied. The KMnO₄ could directly oxidize TFI/PDFI to produce strong CL emission in acidic solution. The effects of experimental conditions were investigated. Under the optimal conditions, the effect of albumin on the TFI/PDFI-KMnO₄ system was investigated. It was found that the addition of albumin into the system could induce enhancement of CL signal, and the enhanced CL intensity is linearly related to the logarithm of concentration of albumin. Based on this study, a novel CL method has been developed for the determination of albumin with high sensitivity and good selectivity. The method was applied to the determination of albumin in human serum samples, and the results were in agreement with those obtained by the bromcresol green (BCG) method. The relative errors for the analytical results were from -5.8% to 4.2%. These new phenomena would further enable people to exploit more CL analytical application of the heterocyclic imidazole derivatives. Springer Science+Business Media, LLC 2009.

Full text of DOI:10.1007/s10895-011-0849-8

J Fluoresc (2011) 21:1607–1615
DOI 10.1007/s10895-011-0849-8
ORIGINAL PAPER
Directly Oxidized Chemiluminescence of 2-Substituted-4,5-di
(2-Furyl)-1H -Imidazole by Acidic Potassium Permanganate
and its Analytical Application for Determination of Albumin
Jing Kang & Yumin Zhang & Zhongxiu Huang & Lu Han &
Jieli Tang & Shuaijun Wang & Yihua Zhang
Received: 22 November 2010 /Accepted: 12 January 2011 /Published online: 28 January 2011
#
Springer Science+Business Media, LLC 2011
Abstract In the paper, 2,4,5-tri(2-furyl)-1H-imidazole
TFI) and 2-phenyl-4,5-di(2-furyl)-1H-imidazole (PDFI),
Introduction
(
were chosen to investigate chemiluminescence (CL) prop-
erties of 2-substituted-4,5-di(2-furyl)-1H-imidazoles. The
directly oxidized CL of analytes by potassium permanganate
Over the years, heterocyclic imidazole derivatives have
attracted considerable attention because of their unique
optical properties [1–3]. These compounds play a very
important role in chemistry as mediators for synthetic
reactions, primarily as a means for preparing functionalized
materials [4]. Imidazole nucleus forms the main structure of
some well-known components of human organisms, i.e. the
(
KMnO ) was in detail studied. The KMnO could directly
4 4
oxidize TFI/PDFI to produce strong CL emission in acidic
solution. The effects of experimental conditions were
investigated. Under the optimal conditions, the effect of
albumin on the TFI/PDFI-KMnO system was investigated.
amino acid histidine, vitamin B , a component of DNA
4
12
It was found that the addition of albumin into the system
could induce enhancement of CL signal, and the enhanced
CL intensity is linearly related to the logarithm of concen-
tration of albumin. Based on this study, a novel CL method
has been developed for the determination of albumin with
high sensitivity and good selectivity. The method was
applied to the determination of albumin in human serum
samples, and the results were in agreement with those
obtained by the bromcresol green (BCG) method. The
relative errors for the analytical results were from −5.8% to
base structure, purines, histamine and biotin. It is also
present in structures of many natural or synthetic drug
molecules, i.e. azomycin, cimetidine and metronidazole [5].
Phenylimidazoles have been studied because of their
important laser properties [6, 7]. Further substitution by
phenyl groups results in other significant optical properties.
An important imidazole derivative is lophine (2,4,5-
triphenylimidazole). Recently, a number of lophine deriva-
tives were synthesized based on lophine skeleton substituted
at the ortho-, meta- and para-substituted in the 2-phenyl ring
and para-substituted in the 4- and 5-aryl rings according to
slightly modified procedure of the Debus method [8–13]. A
variety of lophine analogues having 2-pyridyl or 2-furyl
group at both 4- and 5-positions of heterocyclic imidazole
derivatives have been reported [12, 13]. Among the
derivatives, compounds carrying a 2-furyl group showed
strong photoluminescence (PL) intensities, while those
having a 2-pyridyl group gave very weak intensities [4].
Hitherto, many heterocyclic imidazole derivatives have been
synthesized and studied with regard to their ultraviolet (UV),
PL and chemiluminescence (CL) properties [11, 12].
4
.2%. These new phenomena would further enable people to
exploit more CL analytical application of the heterocyclic
imidazole derivatives.
.
Keywords Chemiluminescence 2,4,5-tri(2-furyl)-1H-
.
.
imidazole 2-phenyl-4,5-di(2-furyl)-1H-imidazole
.
Potassium permanganate Albumin
:
:
:
:
:
J. Kang Y. Zhang L. Han J. Tang S. Wang Y. Zhang (*)
College of Chemistry, Jilin University,
Changchun 130012, People’s Republic of China
e-mail: yihuazh47@yahoo.com.cn
The CL can be defined as emission of light (ultraviolet,
visible or infrared) from a molecule or atom in an
electronically excited state produced by a chemical reaction
at ordinary temperature without any associated generation
Z. Huang
Qianwei Hospital, Jilin University,
Changchun 130012, People’s Republic of China

1
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J Fluoresc (2011) 21:1607–1615
biological oxidants like flavin adenine dinucleotide (FAD),
nicotinamide adenine dinucleotide phosphate (NADP), etc.
O
O
O
O
N
NH
N
NH
KMnO , however, in various homogeneous and heteroge-
4
neous media, as well as on solid supports and in solvent
free conditions, provides excellent results when used in a
O
large number of oxidation processes. The KMnO as a
4
versatile oxidizing agent has been used to generate CL in
many different medium (such as H SO , HCl, H PO ,
(a) TFI
(b) PDFI
2
4
3
4
Fig. 1 Molecular structures of TFI and PDFI
NaHCO and NaOH) [22–24]. In 1920, Grinberg was the
3
first to report the use of acidified KMnO as a reagent for
4
of heat. The CL has attracted a great deal of attention as an
interesting and useful detection method in analytical
chemistry [14–17]. The CL detection has a number of
advantages: (i) high detection sensitivity, (ii) wide linear range
of signal response, (iii) inexpensive reagent and apparatus,
CL, as he investigated the oxidation of pyrogallol, also in
the presence of hydrogen peroxide. Between 1936 and
1939, Audubert reported the oxidation of glucose by
KMnO to generate CL but did not specify whether the
4
reaction was conducted under acidic or alkaline conditions.
Over 40 years later, Mizuno et al. observed, using photon
counting, what they described as extra-weak CL from the
(
iv) easy and rapid measurement. Lophine is a well-known
potential CL compound synthesized by Radziszewski (1877).
It has been used for analysis of some metal ions [18–21] and
chlorinated compounds [20]. The study of CL of heterocy-
clic imidazole derivatives was limited to system of H O as
well-known oxidation of oxalic acid by KMnO
[22]. The
4
acidic solutions of KMnO have been extensively studied
4
due to generating CL during the oxidation of both organic
compounds and inorganic species [25–27]. Because of its
importance as an analytical reagent, significant attention
has also been given to elucidate the mechanism of the
reaction with organic or inorganic substrates [28, 29].
In this paper, two 2-substituted-4,5-di(2-furyl)-1H-
imidazoles, 2,4,5-tri(2-furyl)-1H-imidazole (TFI) and 2-
phenyl-4,5-di(2-furyl)-1H-imidazole (PDFI), were synthe-
sized according to the reported methods [30, 31] and their
2
2
oxidant for the CL reaction [4]. In this paper, the direct
oxidation of 2-substituted-4,5-di(2-furyl)-1H-imidazoles by
acidic potassium permanganate (KMnO ) was applied to
4
producing CL signals.
KMnO , an important oxidant in many organic and
4
inorganic redox reactions, involves the Mn(VII) entity,
which is renowned for its versatility. The KMnO oxidation
4
process is eco-friendly and has gained importance in green
chemistry. Chemical oxidation is full of illustrations of the
use of many uni-atomic metallic ions as oxidants, e.g., Ag
molecular structures are shown in Fig. 1. The KMnO
4
could directly oxidize TFI and PDFI to produce strong CL
emission in acidic conditions. The effects of experimental
(
I), Fe(III), Ce(IV), Cr(VI), Mn(VII), etc., and some large
Fig. 2 Synthesis of TFI and
PDFI
O
O
O
CHO
O
O
O
CuSO 5H O
O
4
2
VB1
OH
O
NaOH
N
O
O
O
N
NH
O
O
CH COONH4
3
O
CH COOH
3
O
CHO
O
O
O
O
O
CH COONH4
3
N
N H
O
+
CH COOH
3

J Fluoresc (2011) 21:1607–1615
1609
Fig. 3 Schematic diagram of
the steady-injection CL system.
(
a)
W
Black box
R
A: KMnO
solution; P
4
solution; B: HCl
, P : peristaltic
air
W
1
2
pump; SL: sample loop; V:
eight-way valve; R: chemifold;
S: sample cell; PMT: photomul-
tiplier tube; PC: computer; W:
waste. a Loading position.
b Injection position
BPCL
A
SL1
P1
SL2
S
W
W
V
PMT
B
air
air
PC
P2
(
b)
Black box
R
W
BPCL
A
B
P1
SL1
SL2
W
W
S
W
V
PMT
air
PC
P2
conditions were investigated. Under the optimal conditions,
the experiments on the effect of albumin on the TFI/PDFI-
acetate, acetic acid, ethyl acetate, methanol, ammonium
sulfate solution, CuSO ·5H O, MgSO , NaOH, KMnO ,
4
2
4
4
KMnO CL system was carried out and the result indicated
H SO , HCl, HNO and H PO were purchased from
2 4 3 3 4
4
that albumin could effectively enhance the CL signal of this
reaction. The enhanced CL intensity is linearly related to the
logarithm of concentration of albumin in the certain
concentration range. The method was applied to the
determination of albumin in human serum samples and the
obtained results were satisfactory.
Beijing Chemical Plant in China. Furfural, petroleum ether
and benzaldehyde were purchased from Tianjin Guangfu
Fine Chemical Research Institute in China. Bovine serum
albumin (BSA) and human serum albumin (HSA) were
purchased from Sigma.
Synthesis of TFI and PDFI
Experimental
The synthesis of TFI and PDFI is shown in Fig. 2. First,
furil was synthesized from furfural by benzoic condensation
and oxidation. TFI was synthesized according to the
literature [30]. A mixture of 1 g furil (5.26 mmol) and
4.05 g ammonium acetate (52.6 mmol) in 20 mL acetic acid
was heated and refluxed. After completion of the reaction,
Reagents and Chemicals
All the reagents were of analytical reagent grade and all
solutions were prepared with ultra-pure water. Ammonium
Fig. 4 Absorption spectra (A)
and PL spectra (B) of TFI and
(A)
(B) 180
1
0
0
0
0
.00
.75
.50
.25
.00
PDFI. Concentration: TFI (curve
a
a
−
6
150
20
a), 2×10 mol/L; PDFI (curve
−6
b), 2×10 mol/L
1
b
b
90
b
a
60
30
0
-
0.25
2
50 300 350 400 450 500 550 600
Wavelength (nm)
250 300 350 400 450 500 550 600 650
Wavelength (nm)

1
610
J Fluoresc (2011) 21:1607–1615
4
000
500
000
500
000
500
000
600
a
3
3
2
2
1
1
a
5
4
00
00
b
b
300
2
1
00
00
0
Injection times
5
00
0
0
10
20
30
40
50
60
0.0
0.2
0.4
0.6
0.8
1.0
1.2
Time (S)
Concentration of HCl (mol/L)
Fig. 5 CL kinetic curves of TFI and PDFI. Concentration: a TFI,
Fig. 7 Influence of HCl concentration on the CL intensity. Concen-
0
.1 mmol/L; H
2
SO
4
, 0.5 mol/L; KMnO
4
, 1 mmol/L. b PDFI,
tration: a TFI, 0.1 mmol/L; KMnO , 1 mmol/L. b PDFI, 0.1 mmol/L;
4
0
.1 mmol/L; H SO
2
4
, 0.5 mol/L; KMnO
4
, 1 mmol/L
KMnO , 1 mmol/L. The error bars denote the standard deviation of
4
the values with the three same determinations
the mixture was cooled to room temperature, diluted with
00 mL of water, and then neutralized with a 20% NaOH
1
reaction mixture was heated and refluxed. After furil was
completely reacted, the reaction mixture was cooled to
room temperature, poured into 100 mL of water, and then
neutralized with an aqueous solution of 20% NaOH to pH 9.
The mixture was extracted with ethyl acetate. The organic
layer was separated, washed with water, and dried over
aqueous solution to pH 9. The mixture was extracted with
ethyl acetate, and the solvent was removed. The ethyl
acetate was evaporated by rotary evaporation. The crude
product was further purified by column chromatography
using a mixture of petroleum ether and ethyl acetate (3:1) as
eluents. Then TFI was recrystallized from methanol. Yellow
single crystals were obtained by slow evaporation of the
solvent at ambient temperature.
PDFI was synthesized according to the literature [31]. A
mixture of 0.82 g furil (4.32 mmol) and 4.05 g ammonium
acetate (52.60 mmol) in 20 mL acetic acid was stirred to
form homogeneous solution at room temperature. Then,
benzaldehyde (4.75 mmol) was added to the mixture. The
MgSO . The ethyl acetate was evaporated in vacuo, and the
4
residue was purified by recrystallization after column
chromatography on Chemapol silica gel (eluent, petroleum
ether/EtOAc=3/1) to obtain PDFI.
Characterization of TFI and PDFI
The TFI and PDFI were characterized by melting point, IR,
MS and NMR. The results obtained by elemental analysis
600
500
400
300
200
100
0
TFI
PDFI
600
a
5
4
3
2
00
00
00
00
b
0
1
2
3
4
5
HCl
HNO3
H PO4
H SO4
Concentration of KMnO (mmol/L)
4
3
2
Fig. 6 The effect of acids on CL intensity. Concentration: TFI,
.1 mmol/L; PDFI, 0.1 mmol/L; acids, 0.5 mol/L; KMnO , 1 mmol/L.
The error bars denote the standard deviation of the values with the
three same determinations
Fig. 8 Influence of KMnO
4
concentration on the CL intensity.
0
4
Concentration: a TFI, 0.1 mmol/L; HCl, 0.75 mol/L. b PDFI,
0.1 mmol/L; HCl, 0.75 mol/L. The error bars denote the standard
deviation of the values with the three same determinations

J Fluoresc (2011) 21:1607–1615
1611
1
200
000
Apparatus
a
1
The CL analysis was conducted on a laboratory-built steady
injection CL system. The schematic diagram of the system
is shown in Fig. 3. The steady injection Analysis Processor
FIA-3100 (Beijing Wantuo Instruments Co. Ltd.) consists
of two peristaltic pumps, a sixteen-hole eight-way valve
and a digital-system to control the time and pump pressure.
PTFE tube (0.8 mm i.d.) was used as connection material in
the steady system. The CL emission was detected by an
ultra-weak luminescence analyzer (type BPCL manufac-
tured at the Institute of Biophysics, Chinese Academy of
Sciences, Beijing, China). The acquisition and treatment of
data were performed with BPCL software running under
Windows XP.
800
600
400
200
0
b
1
000
8
6
4
00
00
00
a
b
200
0
0.00 0.02 0.04 0.06 0.08 0.10
Concentration of TFI/PDFI (mmol/L)
0
.0
0.1
0.2
0.3
0.4
0.5
Concentration of TFI/PDFI (mmol/L)
Fig. 9 Influence of TFI/PDFI concentration on the CL intensity. The
inset displays the linear relationship between ΔI and the concentration
PL spectra were recorded on a RF-5301 spectrofluorimeter
Shimadzu, Japan). The absorption spectra were recorded on
an Australian GBC Cintra 10e UV–vis Spectrometer within
the wavelength range from 200 to 800 nm.
(
of TFI/PDFI. a TFI-KMnO
Concentration: a HCl, 0.75 mol/L; KMnO
.75 mol/L; KMnO , 0.5 mmol/L. The error bars denote the standard
4
system and b PDFI-KMnO
4
system.
4
, 0.75 mmol/L. b HCl,
0
4
deviation of the values with the three same determinations
Procedure
were in conformity with the theoretical results. The results
are described as follows.
Experimental results were obtained using the following
operation parameters: pump rate, 60 rpm/min; sample loop
volume, 300 μL; sampling time, 12 s; sample injection
time, 20 s; the PMT negative voltage, −900 V; the integral
time of the CL signal, 60 s. 200 μL of TFI/PDFI solution
(or solution containing TFI/PDFI and albumin) was first
added into the sample cell (colorless glass tube 1 cm i.d.),
2
,4,5-Tri(2-Furyl)-1H-Imidazole (TFI)
−1
M.p. 196–197°C. IR (KBr) ν (cm ): 3417, 3114, 2926,
1
1
627, 1538, 1477, 1430, 1380, 1201, 1016, 887, 748. H
NMR (500 MHz, CDCl ), δ (ppm): 10.48 (s, 1H), 7.42
3
(
1
s, 2H), 7.36 (s, 1H), 6.92 (d, J=3.3 Hz, 3H), 6.46 (dd, J=3.0,
.7 Hz, 2H), 6.41 (dd, J=3.1, 1.6 Hz, 1H). MS (m/z): (M +
and then KMnO and HCl solution were synchronously
injected into the sample cell using the steady injection
system.
4
+
H) 267.3 (Calcd 266.25). Calcd for C H N O : C, 67.65;
1
5 10 2 3
H, 3.79; N, 10.53. The elemental analysis gave the molecular
formula C H N O (Found: C, 67.53; H, 3.71; N, 10.45).
In sequence 1 (Fig. 3a), pumps P1 and P2 were
activated, and valve V was in the loading position. The
pump P was used to deliver KMnO solution into the
1
5 10 2 3
1
4
2
-Phenyl-4,5-di(2-Furyl)-1H-Imidazole (PDFI)
sample loop (SL ) and the pump P was used to introduce
1 1 2
HCl solution into the sample loop (SL ). In sequence 2
(Fig. 3b), pumps P and P were activated, and valve V
1 2
was in the injection position. The pumps P and P were
1 2
2
2
−
1
M.p.197–198°C. IR (KBr) ν (cm ): 3118, 3054, 1599,
1
1
551, 1481, 1404, 1233, 1090, 886, 732, 590. H NMR
(
300 MHz, CDCl ), δ (ppm): 7.92 (dd, J=8.2, 1.5 Hz, 2H),
used to deliver the air current. The KMnO solution and
3
4
7
3
2
1
.57–7.33 (m, 5H), 6.99 (d, J=3.3 Hz, 2H), 6.53 (dd, J=
.4, 1.8 Hz, 2H). MS (m/z): (M + H) 277.9 (Calcd
HCl solution were simultaneously pumped at the same rate
separately into chemifold R where they were mixed. The
mixed solution was carried into sample cell S and mixed
with TFI/PDFI (or solution containing TFI/PDFI and
albumin) in the sample cell S. CL signal was measured
+
76.29). Calcd for C H N O : C, 73.80; H, 4.38; N,
1
7 12 2 2
0.14. The elemental analysis gave the molecular formula
C H N O (Found: C, 73.55; H, 4.34; N, 9.95).
1
7 12 2 2
Table 1 Linear calibration equation for TFI/PDFI
System
Calibration curve (C, mmol/L)
Linear range (mol/L)
R
Detection limit (mol/L)
RSD (%)
TFI-KMnO
4
ΔI=8077C + 1.513
ΔI=4976C–1.295
1.0×10−8–1.0×10−4
5.0×10−8–7.5×10−5
0.9935
0.9973
3.6×10−9
3.4×10−8
2.0
3.0
PDFI-KMnO
4
The RSD was estimated at 5.0×10− mol/L for TFI/PDFI (n=11)
6

1
612
J Fluoresc (2011) 21:1607–1615
Fig. 10 Calibration curves for
the determination of HSA and
(
A)1200
1000
(B)1200
1000
800
BSA. A TFI-KMnO
and B PDFI-KMnO
4
system
system
4
HSA
8
6
4
2
00
00
00
00
0
600
BSA
HSA
BSA
400
200
0
0
.0 0.5 1.0 1.5 2.0 2.5 3.0
0.5 1.0 1.5 2.0 2.5 3.0 3.5 4.0
Logarithm of concentration of
albumin (nmol/L)
Logarithm of concentration of
albumin (nmol/L)
and recorded. After determination, the mixed solution in
the sample cell S was emptied. The sample cell S was
washed and dried. All experiments were performed in
triplicate.
Results and Discussion
Absorption Spectra, PL Spectra and CL Kinetic Curves
of TFI and PDFI
The concentration of analyte was quantified by measur-
ing the change of CL intensity. ΔI = I –I , where I and I
are CL signals in the absence and presence of analyte,
Figure 4 shows the absorption spectra (Fig. 4A) and PL
spectra (Fig. 4B) of TFI and PDFI. It can be seen from
Fig. 4A that the first excitonic absorption peaks of the TFI
and PDFI appear at 316 and 314 nm, respectively. The PL
spectra show that the excitation peaks of TFI and PDFI are
at around 319 and 317 nm, respectively, corresponding with
the emission peaks of 422 and 435 nm. The CL reaction
s
0
0
s
respectively.
Preparation of Sample
Human serum samples obtained from the Qianwei
Hospital of Jilin University were analysed. 300 μL of
sample and an equal volume of normal saline (NS) were
mixed, and added into a 2 mL centrifuge tube. 600 μL of
saturated ammonium sulfate solution was added drop-
wise into the centrifuge tube when the solution was
stirred continuously. The solution was at rest for 1 h, and
then centrifugalized for 30 mins at 4000 rpm/min. The
upper solution was added into a flask and the lower
sediment was dissolved in 600 μL of NS. 300 μL of
saturated ammonium sulfate solution was added drop-
wise into resulted solution from the sediment when the
solution was stirred continuously. The solution was
centrifugalized for 30 mins at 4000 rpm/min. Then the
upper solution obtained was also added into the flask. To
purify the solution, the free ions were removed via
dialysis for 24 h in water (4°C)
between TFI/PDFI and KMnO was further investigated in
4
acid medium. The KMnO can, in the available concentra-
4
tion range, directly oxidize TFI and PDFI to generate strong
CL emission. The dynamic CL intensity-time profiles of the
TFI-KMnO4 and PDFI-KMnO4 systems are shown in
Fig. 5.
Effect of Acids
The effects of kinds of acids, including HCl, HNO , H PO
4
3
3
and H SO , on the TFI/PDFI-KMnO CL system were
2
4
4
examined. The results are shown in Fig. 6. It can be seen that
the highest emission was observed in HCl medium. Hence,
the HCl solution was chosen as the reaction medium.
The effect of the concentration of HCl on the CL
intensity of TFI/PDFI-KMnO system was examined in the
4
Table 2 Standard curves for albumins
System
Albumin
Calibration curve (C, nmol/L)
r
Linear range (mol/L)
Detection limit (mol/L)
RSD (%)
TFI-KMnO
4
BSA
HSA
BSA
HSA
ΔI=259.4lgC + 44.94
ΔI=365.8lgC–39.89
ΔI=316.5lgC–170.9
ΔI=165.7lgC + 169.8
0.9952
0.9952
0.9990
0.9945
1.0×10−9–1.0×10−6
5.0×10−9–5.0×10−7
1.0×10−8–5.0×10−6
5.0×10−9–1.0×10−6
3.5×10−10
1.2×10−9
4.5×10−9
1.7×10−9
2.5
1.6
3.1
1.8
PDFI-KMnO
4
The RSD was estimated under 1.0×10−7 mol/L for albumin (n=11)

J Fluoresc (2011) 21:1607–1615
1613
Table 3 Tolerance of foreign
substances
Substance
Concentration
TFI-KMnO
change of ΔI (%)
4
system
Concentration
(μmol/L)
PDFI-KMnO
change of ΔI (%)
4
system
(
μmol/L)
Na , Cl−
+
+
500
500
500
500
500
500
500
500
25
−0.3
2.7
500
500
500
500
500
500
250
100
25
−3.1
−0.8
−4.4
0.4
K , Cl−
Ca , Cl−
2
+
−1.8
0.6
Mg , Cl−
2
+
Ba , Cl−
2
+
−2.1
4.9
−0.5
−2.5
3.0
3
+
2−
Al , SO
4
Zn , Cl−
2
+
−3.2
0.9
2
+
2−
Cu , SO
4
1.3
2
+
2−
Mn , SO
4
−1.4
4.5
−4.5
−1.2
1.4
Fe , Cl−
2
+
50
10
Carbamide
Glucose
500
500
500
25
2.7
500
500
500
10
0.6
−4.3
−4.6
4.6
L-Tryptophan
L-Tyrosine
−3.7
4.9
DL-β-Phenylalanine
L-Asparagine
500
250
3.2
250
500
−3.8
3.9
Concentration of HSA is
−0.4
−
7
1
.0×10 mol/L
range of 0.01–1.2 mol/L, and the results are shown in
Fig. 7. The CL intensity increases when the HCl concen-
tration increases from 0.01 to 0.75 mol/L, and the intensity
starts to decrease when the concentration is higher than
and 0.5 mmol/L KMnO were chosen for further research,
respectively.
4
Effect of the Concentration of TFI/PDFI
0
.75 mol/L. Therefore, the optimum HCl concentration was
chosen to be 0.75 mol/L for TFI/PDFI-KMnO system.
The effect of TFI/PDFI concentration on the CL intensity of
the studied system was tested. From Fig. 9, it can be seen
that the concentration of the TFI/PDFI has great influence
on the CL intensity. With the increase of concentration of
TFI/PDFI, the CL intensity increases when the concentra-
tion of TFI/PDFI is lower than 0.15 mmol/L, and the
intensity decreases when the concentration of TFI/PDFI is
higher than 0.15 mmol/L. Therefore, the optimum concen-
tration of TFI/PDFI was chosen to be 0.15 mmol/L.
There is a good linear relationship between the enhanced
CL intensity and the concentration of TFI/PDFI in the range
of low concentration (Fig. 9 inset). The regression
equations of calibration curves, detection limits and relative
standard deviations (RSDs) are summarized in Table 1. The
results demonstrate that the proposed CL system may be
used to detect TFI/PDFI.
4
Effect of the Concentration of KMnO4
The effect of the KMnO concentration on the CL intensity
4
was studied, and the plot of CL intensity versus KMnO4
concentration is shown in Fig. 8. From Fig. 8, it can be seen
that the CL intensity increases with the increase of the
concentration of KMnO . The CL intensity reaches a
4
maximum when the concentrations of KMnO4 are
0
.75 mmol/L and 0.5 mmol/L for TFI-KMnO and PDFI-
4
KMnO systems, respectively. When the concentrations of
4
KMnO are higher than 0.75 mmol/L and 0.5 mmol/L for
4
TFI-KMnO and PDFI-KMnO systems, respectively, the
4
4
CL intensity of the TFI/PDFI-KMnO system decreases
4
with the increase of KMnO concentration. So 0.75 mmol/L
4
Table 4 Determination of HSA
in human serum samples
Sample
Bomocresol
green (g/L)
TFI-KMnO
(g/L) (n=5)
4
system
Relative
error (%)
PDFI-KMnO
(g/L) (n=5)
4
system
Relative
error (%)
1
2
3
4
5
6
47
50
46
48
50
52
46±1.5
−2.1
2.0
45±0.6
50±1.4
45±1.8
46±2.1
51±1.8
50±1.5
−4.3
0.0
51±2.1
45±1.1
50±1.0
52±1.9
49±1.9
−2.2
4.2
−2.2
−4.2
2.0
4.0
−5.8
−3.8

1
614
J Fluoresc (2011) 21:1607–1615
Determination of Albumin
and the obtained results were satisfactory. By comparison
with some existing methods [32–34], the proposed method
has advantages of high selectivity and sensitivity, instrumen-
tal simplicity, low cost and wide linear response range for
determination of albumin. This may intrigue researchers into
gaining a new interest in investigating the CL property of
heterocyclic imidazole derivatives.
The influences of albumins BSA and HSA on the CL
intensity of the TFI/PDFI-KMnO system were examined.
It was found that the addition of albumin into the system
could induce enhancement of CL signal. Based on the
enhancing of CL for TFI/PDFI-KMnO system by albumin,
4
4
a sensitive CL enhancing method has been developed for
the determination of albumin. The calibration graphs were
constructed under the optimal conditions, and the results are
shown in Fig. 10. It can be seen from Fig. 10 that the CL
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