Talanta 197 (2019) 199–203

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Talanta
journal homepage: www.elsevier.com/locate/talanta

A novel hydrophilic polymer-based anion exchanger grafted by quaternized T

polyethyleneimine for ion chromatography
Zhanqiang Yanga, Zongying Lia, Feifang Zhanga, Bingcheng Yanga, , Shengmao Zhangb
⁎

a
Shanghai Key Laboratory of New Drug Design, School of Pharmacy, East China University of Science and Technology, Shanghai 200237, China
b
Engineering Research Center for Nanomaterials, Henan University, Kaifeng 475004, China

ARTICLE INFO ABSTRACT

Keywords: A novel hydrophilic polymer-based anion exchanger has been described for ion chromatography (IC). It is
Ion chromatography prepared by grafting polyethyleneimine (PEI) onto the poly(glycidylmethacrylate-divinylbenzene) (GMA-DVB)
Anion exchanger substrate, followed by hydrolytic treatment to convert the residual epoxide groups onto the surface of GMA-DVB
Polyethylenimine substrates to hydroxyl groups, finally quaternized PEI by a molecule containing epoxide group. Hydrophobic
Poly (glycidylmethacrylate-divinylbenzene)
surface of GMA-DVB substrate can be converted to hydrophilic, aiming to reduce unwanted non-ionic interaction
Hydrophilic polymer
from the substrate. After optimizing the reaction conditions, the obtained anion exchanger showed good se-
paration towards standard inorganic anions under the suppressed mode and complete elution of seven common
inorganic anions could be achieved in less than 16 min by using potassium bicarbonate eluent. It also demon-
strated fast dynamic kinetic and enabled the separation of 7 anions within 3 min at the isocratic mode. Column
efficiency of the anion exchanger was up to 34,000 /m.

1. Introduction including styrene/divinylbenzene) (PS/DVB), ethylvinylbenzene/divi-

Ion chromatography (IC) has been a well-established technique for
the determination of ionic analytes, especially to inorganic anions. The
development of novel anion exchanger is always an active subject in the
field of IC, which bring many applications for IC towards various
samples [1]. Generally, an anion exchanger comprises three parts, a
supporting substrate, spacer arm and the functional groups, in which
the functional groups are fixed onto the surface of the supporting
substrate by a spacer arm. Although the functional groups are created
as ion exchange sites, the substrate composition often has effect on the
final separation by influencing the charge density of the anion ex-
changer or causing unwanted non-ionic interaction with target anions.
The latter is especially true for some polarizable anions onto the anion
exchanger with hydrophobic surface, in which the polarizable anions
have much higher affinity onto the anion exchanger owing to reverse-
phase interaction, then leading to long retention time, peak tailing and
poor peak shape [2–4]. The use of hydrophilic substrate for supporting
anion exchanger can solve such problems to a degree. Thus the devel-
opment of hydrophilic anion exchanger has aroused much interest in
recent years [5–8].
Presently, organic polymer substrates have been predominantly
used in IC system owing to their wider pH tolerance range against ex-
treme pH conditions used in IC. Four types of polymeric substrate
nylbenzene copolymers, polyvinyl alcohol and polymethacrylate resins
are commonly used in IC [9,10]. Recently, the copolymer of glyci-
dylmethacrylate (GMA) and divinylbenzene (GMA-DVB) being chro-
matographic substrate has been paid some attention since such copo-
lymer can be readily modified to introduce desirable functional groups
via epoxide groups in its structure [2,11–14].
Another manner to shield the substrate surface from unwanted non-
ionic interactions with polarizable anions was to use hyperbranched
hydrophilic polymer or branched polyamines [5,6,15]. Poly-
ethylenimine (PEI), a hydrophilic polyamine, contains rich primary,
secondary and tertiary amine groups [16,17]. Thus PEI itself can serve
as anion exchange sites [16] or can be further modified to obtain am-
monium quaternary groups, e.g, use 1,4-butanediol diglycidyl ether
(BDDE) to modify PEI to quaternized amino groups [12,17]. Shchukina
et al. reported an anion exchanger by grafting BDDE quaternized
branched PEI onto the surface of animated PS-DVB substrate, in which
the PS-DVB substrate was firstly modified by acylation and then re-
ductive amination to obtain amine groups, and a BDDE was used as a
spacer arm or linker to bond PEI onto PS-DVB substrate [17]. The ob-
tained anion exchanger demonstrated effective separation for inorganic
anions. Zhang et al. reported a hyperbranched condensation polymer-
based anion exchanger applied for of protein analysis by exposing in an
alternating fashion to graft PEI onto GMA-DVB and then BDDE in a

⁎
Corresponding author.
E-mail address: bcyang@ecust.edu.cn (B. Yang).

https://doi.org/10.1016/j.talanta.2019.01.024
Received 16 November 2018; Received in revised form 2 January 2019; Accepted 5 January 2019
Available online 08 January 2019
0039-9140/ © 2019 Elsevier B.V. All rights reserved.
Z. Yang et al.
repetitive manner [12]. The protein recovery could be obviously in-
creased with the cycle of the reaction between PEI and BDDE.
Herein we have described the preparation of an anion exchanger by
grafting branched PEI onto the GMA-DVB substrate. To improve hy-
drophility of the exchanger, a hydrolytic treatment has been for the first
time used to convert residual epoxide groups associated with GMA-DVB
substrate to hydroxyl groups, and the PEIs were further quaternization
via BEED or a molecule containing an epoxide group. The obtained
anion exchanger has been used for the separation of inorganic anions.
2. Experimental
2.1. Reagents and materials
Glycidyl methacrylate (GMA, 99.5%), divinylbenzene (DVB, 80%),
azobisisobutyronitrile (AIBN,) 1,4-butanedioldiglycidyl ether (BDDE,
95%) and branched polyethyleneimine (PEI) (average MW~600,
~1800 and ~10,000) were purchased from Aladdin (Shanghai, China).
Polyvinylpyrrolidone (PVP, K-30) was purchased from Juyuan
(Shanghai, China). The other reagents were analytical grade, which
purchased from Lingfeng Corp. (Shanghai, China). Unless other stated,
the solutions were prepared by ultrapure water with specific resistivity
of 18.2 MΩ cm.
2.2. Instrumentation
The chromatographic experiments were carried out by IC-2010
system (TOSOH Corp, Japan) operated under the suppressed mode and
a L-3000 HPLC system (RIGOL, Suzhou, China) equipped with ultra-
violet absorbance detector (UVD) for performing non-suppression
mode. An electrodialytic membrane suppressor was from Minghao
Chromatographic Corp. (Suzhou, China). Unless otherwise stated, the
separations were performed at 30 °C and the flow rate of eluent was
1 ml/min.
The SEM was performed on JSM-6360LV Scanning Electron
Microscope–Energy Dispersive X-ray Spectroscope (HITACHI, Japan).
Elemental analysis was carried out on a Varil EL III elemental analyzer
(Elementar, Germany). Fourier transform infrared spectroscopy (FT-IR)
was performed on a Nicolet 6700. The porous properties of the anion
exchange stationary phase was characterized by Micromeritics
TristarⅡ3020.
2.3. Preparation of anion exchanger
The whole preparation procedures of the anion exchanger mainly
consist of four steps, the synthesis of substrate, PEI grafting, hydrolytic
treatment and quaternization, as illustrated in Fig. 1.
Firstly, the synthesis of substrates was much similar to that pre-
viously described [13] with slight modification. Briefly, polystyrene
seed microspheres were synthesized via dispersion polymerization
method as follows. 3.0 g PVP and 100 ml water/ethanol solution (5/95,
v/v) were added to a 250 ml three-neck round bottom flask, and then
0.7 g AIBN and 20 ml styrene were added dropwise under nitrogen
protection. The mixture was then heated in water bath at 70 °C for 24 h.
After filtering and washing with water and ethanol subsequently, the
polystyrene seeds were obtained. 2 g polystyrene seeds and 30 ml 0.2%
SDS (m/v) aqueous solution were added to a 1000 ml three-neck round
bottom flask. With the addition of 7.7 ml dibutyl phthalate (DBP) and
30 ml 0.2% SDS solution, the mixture was stirred at 30 °C for 24 h. To
realize the second swelling, an emulsion solution was prepared by
mixing 2.0 g benzyl peroxide (BPO), 28 ml GMA, 31.2 ml DVB and
66.8 ml toluene with 600 ml 2% polyvinyl alcohol (PVA) aqueous so-
lution containing 2.5 g SDS. After emulsifying with an ultrasonic
treatment, the emulsion solution was added to the above flask. After
swelling at 30 °C for 24 h, the temperature of water bath was increased
to 70 °C under nitrogen atmosphere for 24 h. After filtering and washing
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Talanta 197 (2019) 199–203
with excess hot water and ethanol, the crude substrate microspheres
were obtained, and followed by drying at 60 °C for 12 h to obtain the
final substrates as named by GMA-DVB.
Secondly, 3.0 g dry GMA-DVB and 1.2 g PEI were added to a 100 ml
three-neck round bottom flask containing 50 ml ethanol. By ultrasonic
treatment of 5 min, the mixture was allowed to react at 70 °C for 12 h.
The PEI modified GMA-DVB microspheres were obtained by filtering
and washing with ethanol and followed by drying at 60 °C for 12 h, as
named by GR.
Thirdly, 2.5 g dry GR microspheres were dispersed into 100 ml di-
lute H2SO4 in a flask and then allowed to react at 65 °C for 6 h. After
filtering and washing with water and ethanol successively and then
drying at 60 °C for 12 h, the obtained hydrolytic GR were named as H-
GR.
Finally, 2.5 g dry H-GR were dispersed into 46 ml ethanol and so-
nicated until the microspheres were well suspended. After adding a
certain volume of BDDE (or a molecule containing epoxide group), the
mixture was reacted at 65 °C for 4 h. The final product was collected by
filtration, washing with ethanol and drying in the oven at 60 °C for 12 h,
as named by H-GR-BDDE.
The alternative route was that GR was directly reacted with BDDE as
mentioned above except that no hydrolytic treatment process was in-
volved.
The anion exchangers were in slurry packed into a stainless-steel
column with pure water as slurry and propulsion solvent under the
packing pressure of 5000 psi. When the packing procedure was over,
the obtained anion exchanger was washed with pure water at 0.2 ml/
min for at least 6 h prior to use.
3. Results and discussion
3.1. Characterization of anion exchangers
Scanning electron micrographs of the GMA-DVB microspheres and
anion exchangers were shown in Fig. 2. Good monodispersity was ob-
served for both microspheres and their average diameter was ∼9.3 µm
(GMA-DVB) and 9.6 µm (H-GR-BDDE). The surface area and porosity
was characterized via N2 adsorption-desorption method, as provided in
support information (SI)-Table 1. Relative to GMA-DVB, the micro-
spheres after modification of PEI (anion exchanger) showed lower
surface areas and pore volumes, which may result from the bonding PEI
in the inner pores of the substrates.
Elemental analysis showed an obvious increase of nitrogen content
(0.075–2.1%) after grafting with quaternized PEI as compared with
initial GMA-DVB, as provided in SI-Table 2. This indicated successful
introduction of PEI onto the GMA-DVB particles. The effective anion
exchange capacity was measured to be ∼ 130 μmol/column by the
breakthrough method as described previously [18].
3.2. Preparation of anion exchangers
3.2.1. The choice of the functional reagent
Since PEI contains rich primary, secondary and tertiary amino
groups, PEI-functionalized substrates can in principle behave as an
anion exchanger [16]. In our initial exploration, the PEI-functionalized
GMA-DVB substrates (GR) were directly used as anion exchanger. Un-
fortunately, the common inorganic anions were poorly retained and
baseline separation couldn’t be achieved even using a very dilute eluent
(e.g. 0.5 mM NaHCO3). This may result from much less capacity owing
to less protonated amino groups in such alkaline eluent. One solution to
address this problem is to convert secondary and tertiary amino groups
to quaternary ammonium, in which the effective capacity can be kept
well in a wide pH range. Herein three reagents containing an epoxide
group including epoxypropane (PO), glycidol (GL) and BDDE were
chosen to be quaternized reagent. The obtained three anion exchangers
demonstrated increased retention relative to GR, as shown in SI-Fig. 1.
Z. Yang et al. Talanta 197 (2019) 199–203

Fig. 1. The synthesis route of H-GP-R anion exchanger.

Fig. 2. SEM of GMA-DVB microspheres (a) and H-GP-BDDE anion exchanger (b).

For the model anions, GR-BDDE behaved best in terms of peak shape
and separation resolution, probably resulting from its higher hydro-
philicity. Thus such reagent was selected in the further study.

3.2.2. Hydrolytic treatment of GR

Although GR-BDDE demonstrated effective separation towards
model anions, the separation efficiency was still poor, especially to
bromide and nitrate. As mentioned above, some unwanted non-ion
exchange interactions between the analytes and the substrate may exist.
This is more pronounced for some polarizable anions such as nitrate or
iodide. Such non-ionic interaction did exist for bromide and nitrate, as
indicated by the decrease of retention and improved peak shape by
adding small amount of methanol in the eluent (the eluent concentra-
tion was kept constant) (data not shown). Thus it is necessary to in-
crease the hydrophilicity of GR. Considering large steric hindrance of
PEI molecule, some possible residual unreacted epoxy groups of GR
may exist and hydrolytic treatment of GR was carried out to convert
epoxide groups into hydroxyl groups, aiming to increase the surface
hydrophilicity and in turn to reduce the unwanted interaction from the
substrate. Here, the hydrolytic way was investigated by converting the
epoxy groups into hydroxyl group via reacting with dilute sulfuric acid.
After hydrolytic treatment, the chromatographic performance of the
Fig. 3. Chromatograms achieved by both anion exchangers. Conditions: eluent:
obtained anion exchanger (H-GR) was evaluated by separating five
5 mM KHCO3; 4.6 mm i.d.× 150 mm length; inject volume: 20 μL; analyte
model anions using a bicarbonate eluent, as shown in Fig. 3. Clearly, concentration (μM), IO3- (50), BrO3- (200), NO2- (50), Br- (100), NO3- (50); flow
relative to GR, H-GR showed obvious improvement in terms of peak rate: 1.0 ml/min; detection wavelength: 210 nm; column temperature: 35 °C.
shape and shorter retention towards strongly-retained anions. This

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should result from the improvement of surface hydrophilicity of anion

exchanger. Such was also confirmed by the longer retention of iodate, a
highly hydrophilic and a much less retained anion onto common anion
exchangers. More important, the separation resolution was not affected
by such hydrolytic treatment. Thus such hydrolytic treatment was used
to prepare the final anion exchanger.

3.2.3. The effect of the molecular weight of the PEI attachment

PEI is a polymeric branched molecule, and its polymerization de-
gree determines the final M.W. The PEIs with different M.W mean
different the amounts of amino groups being active sites. Here three
PEIs with different M.W (M.W 600, M.W 1800 and M.W 10,000) were
selected to prepare the anion exchangers (H-GR-BDDE). The chroma-
tographic properties of the obtained anion exchangers were char-
acterized, as illustrated in SI-Fig. 2. Baseline separation of the five
model anions could be achieved onto three anion exchangers. A general
trend was that the retention time of model anions decreased with the
increase of the M.W of PEI. A possible explanation was as follows. With
the increase of the M.W of PEI, the steric effect increases gradually,
leading to less PEI that can be grafted onto the substrate surface. By
comparison, the anion exchanger functionalized with M.W of 1800
demonstrated better peak shape and reasonable retention time. Thus Fig. 4. Separation of seven conventional anions on H-GP-BDDE anion ex-
such PEI was chosen for the further study. changer. Conditions: eluent, 10 mM KHCO3; column: 4.6 mm i.d.× 250 mm
length; analytes concentration (μM), F- (50), Cl- (50), NO2- (50), Br- (50), NO3-
3.2.4. Effect of the reaction temperature on GMA-DVB and PEI attachment (50), SO42- (50), HPO42- (100); other conditions same as Fig. 2.
The effect of the reaction temperature on GMA-DVB and PEI at-
tachment was explored in the range of 60 −75 °C. The obtained four
anion exchangers under different reaction temperature were evaluated
by separating five model anions, as shown in SI-Fig. 3. Baseline se-
paration of five model anions could be achieved on all of the obtained
anion exchangers prepared. Generally, the retention temperature had
less effect on the separation in the tested temperature range. In term of
separation resolution of bromide and nitrate (which is often regarded as
a pair of anions uneasy to separate), the reaction temperature of 70 °C
was regarded as the optimal value.

3.2.5. Effect of the reaction time of GMA-DVB and PEI attachment

The reaction time of GMA-DVB and PEI was explored on the anion
exchanger separation performance by varying the reaction time from
6 h to 24 h, as illustrated in SI-Fig. 4. Baseline separation of the five
model anions can be achieved on all anion exchangers. When the re-
action time increased from 6 h to 12 h, the retention time and the se-
paration resolution of bromide/nitrate had a slight increase, e.g. the
separation resolution of bromide and nitrate increased from 1.58 to 1.9.
Meanwhile, the theoretical plate count of nitrite increased from
28,000 /m to 34,000 /m. When further increasing the reaction time up
to 18 h or 24 h, the retention time of the five model anions were es-
sentially unchanged. So 12 h was chosen to be the optimal reaction Fig. 5. Fast separation of seven anions onto H-GR-BDDE anion exchanger.
time. Conditions: eluent, 10 mM KHCO3; column: 4.6 mm i.d.× 100 mm length; other
conditions same as Fig. 4. Note: the background level was not the real value and
3.3. Chromatographic performance of anion exchanger it was deliberately elevated by adding a given value, aiming to show clearly.

The anion exchanger (H-GR-BDDE) obtained under the optimal

conditions was evaluated by the separation of seven standard inorganic
anions under the mode of suppressed conductance detection, as pro-
vided in Fig. 4. For the eluent of 10 mM KHCO3, a good separation of
seven standard inorganic anions could be achieved in less than 16 min
and the elution orders were same with common commercially anion
exchangers. The column showed relative high separation efficiency and
good peak shape. Take nitrate as an example, the theoretical plate
count and the asymmetry factor were 34,000 /m and 1.31, respectively.
Ion exchange was found to dominate the retention mechanism, as in-
dicated by linear decrease of the retention with increasing eluent con-
centration (data not shown).
Fast analysis is obviously desirable since it can significantly reduce
the analysis time. Here the use of H-GR-BDDE column with short length
(100 mm) for fast analysis has been explored. As provided in Fig. 5,
seven standard inorganic anions onto the column could be eluted out at
the typical flow rate of 1 ml/min in less than 6 min. When increasing
the flow rate up to 2 ml/min, the complete elution could be achieved in
less than 3 min and the system backpressure (9.8 MPa) was still ac-
ceptable. Under such conditions, baseline separation could be still
realized, indicating fast dynamic kinetic performance of such anion
exchanger. Such property will be useful for the fast analysis.
H-GR-BDDE column also demonstrated good mechanical stability,
as proved by the highly linear plot between the applied rate and the
produced backpressure drop (up to 3000 psi) (data not shown). This can
contribute to high crosslinking degree of GMA-DVB substrate. The re-
tention of the anions was studied over 3500 column volume of

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continuous operation with 7 mM KHCO3 as eluent. The retention times
of all anions were less than 0.34% (RSD, n = 15). The intra-day and
inter-day RSD of the retention time of the model anions are less than
0.14% and 0.18%, respectively. These indicated excellent stability of
the anion exchanger (SI-Fig. 5). The column has been used off and on
for over than 6 months and no observable change was found, indicating
good long-term stability. Clearly, relative to hydroxide eluent, bi-
carbonate is a rather mild eluent, which will be beneficial to long-term
stability of anion exchanger.
3.4. Separation of polarizable anions by H-GR-BDDE column
In consideration of high hydrophilic property of H-GR-BDDE
column, it was used to separate polarizable anions. Two polarizable
anions including iodide, and thiocyanate with other common anions
were chosen to be model and the chromatogram was illustrated in SI-
Fig. 6. By using 13 mM KHCO3 as the eluent, the separation could be
completed in 16 min. The asymmetric factors were 1.06 and 1.21 for
the peaks of I- and SCN-, respectively and their corresponding theore-
tical plate numbers were 23,160 /m and 20,336 /m. Relative to iodide,
SCN- still had strong retention, which may be caused by the interactions
with some unsheltered area on the substrate surface. Such has been
confirmed by adding some amount of methanol in the eluent, in which
the obvious decrease of retention time of polarizable anions. More work
is needed for further increase the hydrophility of the substrate.
Nevertheless, these results indicated that the H-GR-EGDGE have a
promising application for detecting polarizable anions.
3.5. Effect of the column temperature on separation
The column operated in elevated temperature would enhance the
kinetic and transport property of the analytes due to the decrease of the
viscosity of eluent and the increase of the analyte diffusivity. Here, the
effect of temperature on the performance of H-GR-BDDE column was
explored, as shown in SI-Fig. 7. In the tested range of 30–45 °C, good
linear relationship between logarithm of the retention factor (lnk′) and
reciprocal absolute temperature existed for all the tested analytes. The
retention of the model anions slightly decreased with the increase of
column temperature except dihydrogen phosphate and fluoride. Lower
operation temperature offered better separation of dihydrogen phos-
phate and sulfonate.
4. Conclusion
A new anion exchanger based on GMA-DVB substrate grafted by a
quaternized PEI has been described for IC. By hydrolytic treatment, the
hydrophobic GMA-DVB substrate can be manipulated into hydrophilic
owing to the introduction of hydroxyl groups. This can reduce some
unwanted non-ionic interactions from the substrate to a degree. Under
the suppression mode, the obtained anion exchanger showed good se-
paration towards seven common inorganic anions in 16 min, which
makes it promising for the routine analysis of water. Another desirable
feature of such anion exchanger is its fast dynamic kinetic performance,
which is believed to have applications for fast analysis.
Acknowledgements
This work was sponsored by the National Key Research and
Development Program of China (2018YFC1603500) and National
Natural Science Foundation of China (No. 21477037). BCY acknowl-
edges financial support from Fundamental Research Funds for the
Central Universities (22221818014).
Appendix A. Supplementary material
Supplementary data associated with this article can be found in the
online version at doi:10.1016/j.talanta.2019.01.024.
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