Y. Li et al. / Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy 234 (2020) 118277
7
Table 1
Comparison of analytical performance of the three sensors.
Sensor
Carrier
Carrier size (μm)
Detection limit (μM)
Detection range (μM)
Response time (min)
Number of recycles
I
Chloracetyl
Chloromethyl
Chloromethyl
200
40
40
0.439
0.065
0.072
1–9
0.5–4
0.5–4
4
4
3
≥3
≥3
≥3
PS-RB-2
PS-R6G-2
PS-R6G-2 were significantly different from PS-Cl, with longer diameter
and rougher surface. The obvious changes in surface morphology indi-
cated that the immobilization was successful.
obtained two electrons to form negative ion. In the LUMO of neutral RB-
2, electrons were mainly delocalized on the pyrene ring. When electron
transitions took place, some electrons entered into the negative HOMO,
and then delocalized on the rhodamine ring. Compound R6G-2 had
quite similar electron distributions. The vertical electrophilic potential
of neutral RB-2 and R6G-2 was 311.0 kj/mol and 410.3 kj/mol, respec-
tively. However, their adiabatic electrophilic potentials (65.5 kj/mol
for RB-2 and 148.2 kj/mol for R6G-2) decreased, which meant it was
easier to get electrons when the structure changed. In comparison, the
narrower energy gap made RB-2 with a higher reactivity, which led to
better fluorescent property.
In addition, EDTA titration was carried out to explore the regenera-
tion ability. EDTA, which had stronger chelation with Hg(II), made fluo-
rescence disappear along with free sensor regain. As shown in Fig. 8, PS-
RB-2 and PS-R6G-2 could be reused for more than three times in aceto-
nitrile. However, a decrease in fluorescence intensity was presented
with the repeat time increase, which might due to the loss of some rho-
damine derivatives on the surface of polystyrene microspheres. In addi-
tion, the surface color constantly changed in the process of reuse,
indicating the rhodamines immobilized on the microspheres presented
an “off-on” state.
3.2. Fluorescence properties
To explore the sensing properties, the fluorescence titration of PS-
RB-2 and PS-R6G-2 with Hg(II) was conducted. The recognition was
finished in 4 min with negligible fluorescence changes at pH values of
6.0–9.0. Upon incremental addition of Hg(II), enhanced fluorescence
emission was observed at 574 nm (Fig. 4). For PS-R6G-2, Hg(II)
expressed a linear concentration range from 0.5 to 4 μM with a correla-
tion coefficient of 0.996. The detection limit was determined to
0.072 μM based on 3s/k where s was standard deviation and k was the
slope of calibration plot [39,40]. However, sensor PS-RB-2 displayed a
much higher fluorescence response and a lower detection limit of
0.065 μM in comparison, which was also better than sensor I (Scheme
1b). Moreover, the microspheres surface showed a color change from
yellow to rosy red after the addition of Hg(II), which illustrated that sen-
sors could perform fluorescence “off-on” response and visual recogni-
tion of Hg(II).
As shown in Fig. 5, weak fluorescence enhancement was observed
with the addition of Ca(II), Na(I), K(I), Ni(II), Pb(II), Mg(II), Zn(II), Cu
(II), Mn(II), Ag(I), Cd(II), Fe(II), Cr(III) and Ba(II) ions (black bars). How-
ever, when Hg(II) ions were added, significant variation was observed
(red bars). Compared to PS-R6G-2, PS-RB-2 had better selectivity and
fluorescence response to Hg(II) along with obvious color change, from
yellow to rosy red, whereas other ions remained yellow. Its anti-
interference ability to Cu(II) was also better than sensor I (Scheme
1b). From the selectivity of other metal ions, PS-RB-2 had the similar
performance with sensor I.
To explore the influences of different polystyrene microspheres, the
analytical performances of PS-RB-2 and PS-R6G-2 were compared with
sensor I (Scheme 1b). Sensor I was employed chloroacetylated polysty-
rene microspheres (200 μm) as carriers and rhodamine RH as recogni-
tion probe. As shown in Table 1, PS-RB-2 and PS-R6G-2 with smaller
size had lower detection limit, whereas other properties such as re-
sponse time, selectivity and recyclability were no degradation. The de-
crease in size would increase the specific surface area and the loading
amount. Moreover, the stability of chloroacetylated microspheres was
lower than chloromethyl microspheres due to the greater viability of
chloracetyl group. During the synthesis of chloroacetylated micro-
spheres, other side reactions also occurred, which reduced the amount
of Cl and then reduced the loading rate. In addition, PS-RB-2 and PS-
R6G-2 could be used at r.t. with no special conditions and the operation
was relatively simple. Compared with some Hg(II) chemical sensors
[13,14], although they have low detection limit and excellent selectivity,
bad reusing property restricted their large scale application. The modi-
fication of small-molecule fluorescent probe on solid phase carriers
such as polystyrene microspheres can solve this problem. However,
the introduction of larger size carriers may reduce their detection per-
formance, such as detection limit. Thus, chloromethyl microspheres
with smaller size were selected as carriers, and solid-phase sensors
3.3. Detection mechanism
Detection mechanism of PS-RB-2 and PS-R6G-2 with Hg(II) were
shown in Fig. 6. At first rhodamine existed in a closed lactam spirocycle
with nonfluorescence. With the addition of Hg(II), PS-RB-2 and PS-
R6G-2 could chelate with Hg(II) via N and O atoms, which caused the
cycle open along with apparent color change and strong fluorescence
emission at 574 nm. To further explain the fluorescent difference from
theoretic level, compounds RB-2 and R6G-2 were selected as templates,
and the DFT calculations of RB-2 and R6G-2 with Hg(II) were carried out
(Fig. 7).
The frontier-orbital energies are closely related to molecular activity
in general [41]. EHOMO is a rough measure of electron-donating ability,
whereas ELUMO acts in reverse [42,43]. The comparison of DFT results
for RB-2 and R6G-2 with Hg(II) were shown in Fig. 7, where the positive
phase was symbolized with red and the negative phase green. Both of
them have delocalized π systems. It is easier for the vertical transition
of delocalized π electrons from HOMO to LUMO. It could be seen that
the HOMO-LUMO energy gap of neutral RB-2 (0.131 a.u.) was smaller
than R6G-2 (0.149 a.u.). The narrow gap implies a higher chemical reac-
tivity because it is energetically favorable to add electrons to a low-lying
LUMO or extract electrons from a high-lying HOMO so to form an acti-
vated complex in any potential reaction [44]. It indicated that the elec-
tronic transfer in RB-2 was easier and its chelation with Hg(II) might
possess a relatively higher reactivity, which correlated well with the
fluorescent results. When the chelation took place, the neutral molecule
Table 2
Recovery results for spiked Hg(II) in real samples using the developed method.
Sample
Original
(μM)
Added
(μM)
Detected
(μM)
Recovery
(%)
Relative
error
(%)
0
0
0
1.0
2.0
3.0
1.0
2.0
3.0
1.0
2.0
3.0
0.926
2.116
2.811
1.061
1.969
3.136
0.913
2.082
3.175
92.60
105.80
93.70
106.10
98.45
104.52
91.30
104.10
105.84
7.40
5.80
6.30
6.10
1.55
4.52
8.70
4.10
5.84
Water
0.0021
0.0021
0.0021
0
0
0
Lake water
Crucian
carp