Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy 303 (2023) 123205

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Spectrochimica Acta Part A:

Molecular and Biomolecular Spectroscopy
journal homepage: www.journals.elsevier.com/spectrochimica-acta-part-a-
molecular-and-biomolecular-spectroscopy

Solvent-directed multiple correspondence fluorescent probe for highly

selective and sensitive detection of Cu2+ and Mg2+
Xianfa Zhang a, Shuaibing Yu a, Xuliang Pang a, Xiaochen Ren a, Bo Zhang a, Jinming Kong b,
Lianzhi Li a, *
a
School of Chemistry and Chemical Engineering, Liaocheng University, Liaocheng 252059, PR China
b
School of Environmental and Biological Engineering, Nanjing University of Science and Technology, Nanjing 210094, PR China

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 solvent-directed Schiff base fluores­

cent probe was synthesized.
• The probe displayed excellent selectivity
and sensitivity for Cu2+ in PBS buffer.
• The probe displayed excellent selectivity
and sensitivity for Mg2+ in acetonitrile.
• The interactions of probe with Cu2+ and
Mg2+ were characterized by NMR, IR
and XPS.

A R T I C L E I N F O A B S T R A C T

Keywords: A solvent-directed, new Schiff base multiple correspondence fluorescent probe, (E)-2-(2-hydroxybenzylidene)
Schiff base hydrazine-1-carboxamid (L), was synthesized for selective sensing of Cu2+ and Mg2+ ions. L showed excellent
Fluorescence probe selectivity and high sensitivity toward Cu2+ in “turn off” mode with a detection limit of 40.5 nM in 10 mM, pH =
Solvent-directed
7.0 PBS buffer. Contrary to that, when acetonitrile was used as the solvent, L exhibited highly selective and
Copper(II) ion
sensitive fluorescence sensing ability for Mg2+ in “turn on” mode with a detection limit of 9.5 nM. L can co­
Magnesium(II) ion
ordinate to Cu2+ and Mg2+ in a 1:1 molar ratio, respectively, evidenced by Job’s plot analysis. Their binding
modes were investigated by NMR, IR and XPS spectroscopies. Moreover, the satisfied results were obtained when
L was used to detect Cu2+ and Mg2+ in real water samples.

1. Introduction physiological processes in organisms [2]. Lack of Cu2+ in humans may

cause anemia, coronary heart diseases and bone abnormalities [3], but
Copper is an essential and the third abundant metal element in high concentration of Cu2+ in human will lead to serious diseases such as
human body [1]. It plays a crucial role in many fundamental neurodegenerative diseases, liver, kidney damage or even death [4].

* Corresponding author at: School of Chemistry and Chemical Engineering, Liaocheng University, Liaocheng 252059, PR China.
E-mail address: lilianzhi1963@163.com (L. Li).

https://doi.org/10.1016/j.saa.2023.123205
Received 7 May 2023; Received in revised form 21 July 2023; Accepted 24 July 2023
Available online 25 July 2023
1386-1425/© 2023 Elsevier B.V. All rights reserved.
X. Zhang et al.
In order to maintain Cu2+ content in the human body at a reasonable
level, the copper ion intake should be limited. The World Health Orga­
nization (WHO) and United States Environmental Protection Agency
(USEPA) recommended the maximum acceptable levels of Cu2+ in
drinking water are 2.0 and 1.3 mg/L, respectively [5,6]. Because the
wastewater containing Cu2+ from industrial and agricultural cause
water pollution and has an adverse effect on human health, it is neces­
sary to develop an effective detection method to detect the Cu2+ in
environment.
Similarly, Mg2+ is one of the most important elements [7], and plays
a crucial role in various biological processes and environmental appli­
cations. Mg2+ plays significant physiological roles in a large number of
cellular processes, such as membrane protein activities [8], biochemical
process of cell proliferation [9] and most enzyme activities [10].
Abnormal content of magnesium in human body associates with several
diseases, for example, Mg2+ deficiency may led to a series of chronic
diseases such as hypertension, diabetes, coronary heart disease and
osteoporosis [11,12].
Due to their importance of Cu2+ and Mg2+ in biochemical and
physiological processes, several analytical methods such as atomic ab­
sorption spectrum (AAS) [13], ion-selective electrodes (ISES) [14],
inductive coupled mass atomic emission spectrometry (ICP-AES) [15]
and inductive coupled plasma mass spectroscopy (ICP-MS) [16] have
been developed for their determination. However, these methods need
sophisticated and expensive equipment and operational procedures are
also hard to handle, and require experienced operators with high data
analysis skills. Therefore, colorimetric and fluorescent chemosensor
methods have been considered for metal ion sensing because of its high
selectivity, sensitivity, cheap and easy to operate [17–21].
In recent years, fluorescent probes with selectivity for multi-metal
ions that have cost advantage and real-time applications have received
a great deal of attention and become an attractive research area [22].
Several multi-metal ion probes have been reported such as for the de­
tections of Hg2+ and Cu2+ [23,24], Mg2+ and Zn2+ [7,9,25]. However,
to design more effective multi-responsive probes for different metal ions
still remains a challenge.
Therefore, it is an urgent task to design a fluorescence probe to detect
Cu2+ and Mg2+. Schiff bases and their corresponding complexes have
been extensively studied on the aspects of pharmaceutical chemistry,
catalytic synthesis, luminescent materials and analytical chemistry [26].
In this paper, we designed and synthesized a new Schiff base for multiple
correspondence fluorescent probe of metal ions, (E)-2-(2-hydrox­
ybenzylidene) hydrazine-1-carboxamid (L), derived from the conden­
sation of semicarbazide hydrochloride and salicylaldehyde. It has been
found that the fluorescence property of L showed distinct solvent effects.
Interestingly, due to the solvatochromic property of L, we can make it as
a multi-responsive fluorescent probe by changing the solvent. As in
acetonitrile, L itself showed weak fluorescence and showed a selective
fluorescence “turn on” response when Mg2+ was added to the solution.
When the solvent is 10 mM, pH = 7.0 PBS:ethanol (99:1, v/v), L itself
showed strong fluorescence signal and its fluorescence can be quenched
in the presence of Cu2+. Therefore, L was a solvent-directed multiple
correspondence fluorescent probe for highly selective and sensitive
detection of Cu2+ and Mg2+.
2. Experimental
2.1. Materials and instruments
Semicarbazide hydrochloride was purchased from Tianjin Chemical
Reagent First Factory (Tianjin, China). Salicylaldehyde was obtained
from Aladdin Reagent (Shanghai) Co., Ltd (Shanghai, China). Deuter­
ated DMSO for NMR measurements was purchased from Cambridge
Isotope Laboratories (CIL). N,N-Dimethylformamide (DMF) and
dimethyl sulfoxide (DMSO) were purchased from Tianjin Fengchuan
chemical Reagent Technologies Co., Ltd. (Tianjin, China). Metal nitrate
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Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy 303 (2023) 123205
and chloride, tetrahydrofuran, acetonitrile and other chemical reagents
were analytical grade unless noted. The solvents and reagents were used
as received.
The elemental analysis was conducted using an Elementar Vario EL
Cube apparatus (Germany). 1H NMR measurements were measured
with a Bruker AVANCE NEO 500 MHz spectrometer (Swiss) with
DMSO‑d6 used as solvent and tetramethylsilane (TMS) as an internal
standard. Fourier transform infrared (FTIR) spectra were obtained using
a Nicolet 5700 spectrometer (Thermo Electron Corp., USA). Fluores­
cence spectra were measured on a Hitachi F-7000 fluorescence spec­
trofluorometer (Hitachi Inc., Japan). UV–visible spectra were acquired
on a UH-4150 UV–visible absorption spectrophotometer (Hitachi Inc.,
Japan). API 4500 QTRAP Mass Spectrometer (Applied Biosystems/MDS
SCIEX, USA). X-ray photoelectron spectroscopy (XPS) measurements
were performed using ESCALAB Xi + X-ray photoelectron spectroscopy
(Thermo Fisher Scientific Inc, USA).
2.2. Synthesis and characterization of Schiff base (L)
A mixed solution of semicarbazide hydrochloride (0.111 g, 1.0
mmol) and salicylaldehyde (0.122 g, 1.0 mmol) in ethanol (10 mL) was
refluxed for 4 h. After that, a white precipitate was formed and then
cooled to room temperature. The precipitate was separated out and
washed with ethanol for three times. The precipitate was recrystallized
from ethanol, 0.192 g crystals was obtained with the yield of 82.3 %. 1H
NMR (500 MHz, DMSO‑d6) δ 10.17 (s, 1H), 9.96 (s, 1H), 8.14 (s, 1H),
7.75 (d, J = 7.6 Hz, 1H), 7.17 (m, 1H), 6.85 (d, J = 8.1 Hz, 1H), 6.81 (t, J
= 7.5 Hz, 1H), 6.38 (s, 2H). Elemental analysis calculated for C8H9N3O2:
C, 53.63; H, 5.06; N, 23.45. Found: C, 52.96; H, 4.977; N, 23.55. The
synthesis routine of Schiff base is shown in Scheme S1.
2.3. Stock solution
The nitrates salts or chlorinated salts of Ag+, Al3+, Ba2+, Ca2+, Cd2+,
Co2+, Cr3+, Cu2+, Fe3+, Hg2+, K+, Li+, Mg2+, Mn2+, Na+, Ni2+, Pb2+,
Sr2+, Zn2+ were dissolved in double distilled water to prepare a 10 mM
stock solution. Stock solution of L (1.0 mM) for detecting Mg2+ was
prepared in acetonitrile. Stock solution of L (1.0 mM) for detecting Cu2+
was first dissolved in ethanol then dilution with 10 mM, pH = 7.0 PBS
buffer (PBS:ethanol = 99:1, v/v).
2.4. Spectral measurements
Fluorescence and UV–Vis absorption spectra of compound L were
measured by fluorescence and UV–Vis instruments at room temperature.
Added 10 μM L stock solution to the cuvette followed by the addition of
the requisite volume of cation solution using a microliter syringe. Upon
addition of aliquot of each metal ion, the solution was well-mixed and
then the spectrum was measured. For measuring the fluorescence
spectra, both excitation and emission slit widths were set at 5 nm. The
emission spectra were acquired with the excitation wavelength at 320
nm for Cu2+ detection, and at 370 nm for Mg2+ detection. UV–Vis
spectra were conducted in 3 mL cuvette in the wavelength range of
200–500 nm.
3. Results and discussion
3.1. Fluorescence emission property of L in different solvents
Fluorescence emission property of L was investigated in different
solvents. As shown in Fig. 1a, it can be found that the fluorescence
emission spectrum of L has different intensity in different solvents. L
showed maximum emission in 10 mM, pH = 7.0 PBS:ethanol (99:1, v/v),
with a maximum emission peak at 480 nm. In other solvents, it has very
weak fluorescence emission intensity.
X. Zhang et al. Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy 303 (2023) 123205

Fig. 1. (a) Fluorescence emission spectra of L (10 μM) in different solvents. (b) Fluorescence spectra of L (10 μM) in the presence of metal ions (1 eq) in 10 mM, pH
= 7.0 PBS:ethanol (99:1, v/v). (c) Selective fluorescence “turn on” response of L (10 μM) towards Mg2+ in acetonitrile.

3.2. Selectivity of L towards Cu2+
In order to study the fluorescence behavior of L towards various
metal ions in 10 mM, pH = 7.0 PBS:ethanol (99:1, v/v) system, we
studied the fluorescence properties of L (excited at 320 nm) in the
presence of various metal ions (Ag+, Al3+, Ba2+, Ca2+, Cd2+, Co2+, Cr3+,
Cu2+, Fe3+, Hg2+, K+, Li+, Mg2+, Mn2+, Na+, Ni2+, Pb2+, Sr2+, Zn2+). As
shown in Fig. 1b, upon the addition of 1 eq. of various metal ions, only
when added the Cu2+ into the solution, can it induced a strong fluo­
rescence quenching, while other metal ions showed very weak fluores­
cence quenching. This may be attributed to the formation of L-Cu2+
complex between L and Cu2+, due to the paramagnetic nature of Cu2+
which quenched excited state of phenylmethyleneamine fluorophore by
intramolecular charge transfer (ICT) [27].
3.3. Selectivity of L towards Mg2
Since Schiff base L showed significant solvatochromic property and
exhibited weak emission intensity in solvents, solvents including
acetonitrile, methanol, EtOH, DMF, DMSO and THF were tested if L
could be used as a fluorescence “turn on” probe for metal ions in these
solvents. The metal ion selectivity of L was tested in these solvents with
metal ions (Ag+, Al3+, Ba2+, Ca2+, Cd2+, Co2+, Cr3+, Cu2+, Fe3+, Hg2+,
K+, Li+, Mg2+, Mn2+, Na+, Ni2+, Pb2+, Sr2+, Zn2+), respectively. As
shown in Fig. 1c, Mg2+ could significantly enhance the fluorescence
intensity of L in acetonitrile, but other metal ions had no significant
effects on fluorescence intensity of L under the same conditions. Result
indicated that L exhibit a specific fluorescence response toward Mg2+ in
acetonitrile. L itself showed very weak fluorescence emission at 442 nm
due to the C = N isomerization, which has long been known as the main
decay process [28,29]. The fluorescence intensity of L at 442 nm
increased almost 68 fold upon the addition of 1 equiv. Mg2+. The sig­
nificant fluorescent enhancement may be attributed to the formation of
complex L-Mg2+, which inhibited the isomerization of C = N bond,
increased the rigidity of the molecule, and produced the chelated
enhanced fluorescence (CHEF) effects [28,30–32]. This result suggested
that L could be used as a fluorescence “turn on” probe for Mg2+ in
acetonitrile solution.
3.4. Turn off fluorescence response of L toward Cu2+
3.4.1. Uv–vis absorption and fluorescence spectral studies
When the 10 mM, pH = 7.0 PBS:ethanol (99:1, v/v) solution of L was
titrated with Cu2+, UV–Vis and fluorescence spectral changes were
observed. Initially, UV–Vis absorption spectrum of L (10 μM) showed an
absorption peak at 275 nm and a shoulder peak at about 310 nm
(Fig. 2a), possibly due to the π-π* transition and the intermolecular
charge transfer (ICT) transition, respectively [27]. Upon gradual addi­
tion of Cu2+ (0–9.9 μM), the original absorption bands of L decreased
and two new absorption peaks appeared at about 243 nm and 355 nm,
with two clear isobestic points observed at 260 nm and 329 nm, infer­
ring that the reaction between L and Cu2+ has occurred and the for­
mation of new complex. From the fluorescence titration experiment, it
was found that the addition of Cu2+ gradually decreased the fluores­
cence emission intensity of L, and it was almost completely quenched
while the concentration of Cu2+ reached 9.9 μM (Fig. 2b). The Fluo­
rescence quenching phenomenon may be due to the metal to ligand
charge transfer (MLCT) reaction between L and paramagnetic Cu2+,

Fig. 2. (a) UV–Vis absorption spectral response of L (10 μM) upon gradual addition of Cu2+ in 10 mM, pH = 7.0 PBS:ethanol (99:1, v/v). [Cu2+] (1–7) = 0, 1.66,
3.32, 4.98, 6.62, 8.26 and 9.9 μM, respectively. (b) Fluorescence spectral response of L (10 μM) upon gradual addition of Cu2+ in 10 mM, pH = 7.0 PBS:ethanol (99:1,
v/v), [Cu2+] (1–11) = 0, 0.999, 1.996, 2.99, 3.98, 4.98, 5.96, 6.95, 7.94, 8.92 and 9.9 μM, respectively. Inset: image of L in the absence and presence of 1 eq Cu2+
when observed under 365 nm UV light. (c)The curve of F0-Fx with Cu2+concentration (0.999–7.94 μM). F0 and Fx represent the fluorescence intensity of L in the
absence and presence of Cu2+.

3
X. Zhang et al.
resulting in chelation enhanced quenching (CHEQ) [27].
The fluorescence titration data showed a good linear relationship
between the decrease of fluorescence intensity of L and Cu2+ concen­
tration in the range of 0.999–7.94 μM, with a correlation coefficient of
R2 = 0.997 (Fig. 2c). The detection limit, calculated according to the
equation LOD = 3σ/S method, was 40.5 nM, which was much lower than
United States Environmental Protection Agency (USEPA) maximum
allowable level of Cu2+ (20 μM) in drinking water [33]. As shown in
Table S1, this prober was better than most probers reported by previous
references.
3.4.2. Effect of pH
The fluorescence intensity of L in the absence and presence of Cu2+ in
aqueous solutions at different pH are shown in Fig. S1. The pH was
adjusted with HCl and NaOH aqueous solution. The fluorescence in­
tensity of L decreased significantly when it was in the acidic solution
(pH < 5). When the pH > 10, the fluorescence quenching of L by Cu2+
was weak, which may be attributed to the partial hydrolysis of Cu2+ ion.
In contrast, the fluorescence quenching of the L in the range of pH
4.0–9.0 was strong, and the quenching efficiency reached a maximum at
pH 7.0. Therefore, pH 7.0 is the most suitable system for detecting Cu2+.
3.4.3. Binding capability
To determine the binding stoichiometry between L and Cu2+ com­
plex, the fluorescence spectra of L at different [Cu2+]/([Cu2+] + [L])
concentration ratios were determined (Fig. S2). As shown in the Job’s
plot (Fig. 3a), the fluorescence intensity of L at 480 nm shows a turning
point at a [Cu2+]/([Cu2+] + [L]) ratio of 0.5. This means that the
binding stoichiometry between L and Cu2+ was 1:1. The binding con­
stant (K) can be obtained from the fluorescence titration spectra by the
Benesi–Hildebrand equation (F0 − Fmax)/(F0 − Fx) = 1 + 1/K[M]n [34].
Where F0 is the intensity of L, Fmax is the fluorescence intensity of L in the
presence of Cu2+ at complete interaction state, Fx is the intensity of L at
an intermediate Cu2+ concentration. K is the binding constant, [M] is the
concentration of Cu2+, n is the number of Cu2+ ion bound to each L (here
n = 1). As shown in Fig. 3b, the K value was calculated to be 8.58
(±0.025) × 104 M− 1 (R2 = 0.994).
3.4.4. Competitive study
High selectivity is an important standard to design the success of a
probe. To further explore the selectivity of the L and the possible
interference that other metal ions and anions may cause when detecting
Cu2+, competition experiments were carried out. The fluorescence
Fig. 3. (a) Job’s plot of L–Cu2+ complex in 10 mM, pH = 7.0 PBS:ethanol (99:1, v/v). The total concentration of L and Cu2+ was 10 μM. λex = 320 nm. (b) Benesi-
Hildebrand plot to estimate the binding constant of L–Cu2+complex.
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Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy 303 (2023) 123205
spectra of L were measured in the presence of 1 eq. of different metal
ions and anions, and further addition of 1 eq. Cu2+. As shown in Fig. S3,
in the presence of other metal ions and different anions, there is almost
no interference in the detection of Cu2+. The result suggested that L has
an excellent specificity for the Cu2+.
3.4.5. Application in real water samples analysis
Due to the high selectivity of L for the detection of Cu2+, the
experiment for detecting Cu2+ in real water samples was carried out.
The water from the lake of our school was collected and filtered. Direct
addition of lake water failed to influence the fluorescence of L, therefore,
different concentrations of Cu2+ were spiked in lake water samples and
the recovery percentage was analyzed. As shown in Table S2, the Cu2+
concentration determined was close to the spiked samples. This result
means that the L can be applied to detect Cu2+ in real water samples.
3.4.6. Binding mode study
In order to investigate the binding mode of L with Cu2+, FTIR, XPS
and NMR spectra were carried out. As shown in Fig. S4, in the FTIR
spectrum of L, the peaks at 3463.8 and 3151.4 cm− 1 may be attributed to
NH and NH2 group of L. After addition 1 eq. of Cu2+, the FTIR spectrum
of L was changed, the peak at 3151.4 cm− 1 was disappeared, and the
peak at 3463.8 cm− 1 was shifted to 3407.5 cm− 1. The peaks at 1677.2
and 1599.2 cm− 1 attributed to C = O and C = N group of L were shifted
to 1654.4 and 1603.1 cm− 1, respectively. These changes indicated that
the nitrogen atoms of NH2 and C = N participated in the binding to Cu2+.
In addition, the FTIR spectrum of [L–Cu2+] also contained an intense
peak of NO−3 group at 1384.46 cm− 1 [28,35].
To better confirm the binding mode, X-ray photoelectron spectros­
copy was carried out (Fig. 4). As shown in Fig. 4a, the photoemission
peaks of Cu 2p, O 1 s, N 1 s and C 1 s can be found in the sample, which is
attributed to the presence of L-Cu2+ complex [36]. As can be seen from
Cu 2p spectrum in Fig. 4b, peaks at 934.9 eV and 955 eV are ascribed to
Cu 2p3/2 and Cu 2p1/2, with a shake-up satellite peak at 940–945 eV,
indicating that Cu is in + 2 oxidation state [37]. N 1 s spectrum of L can
be further deconvoluted into three peaks as can be seen from Fig. 4c. The
peaks at ~400.2 eV, ~399.5 eV and ~400.8 eV can be corresponded to
–NH2, –NH- and –HC = N-, respectively [38,39]. N 1 s spectrum of L +
Cu2+ complex (Fig. 4d) can be further deconvoluted into four peaks at
~399.6 eV, ~399.3 eV, ~399.9 eV and ~406.2 eV, which can be cor­
responded to –NH2, –NH-, –HC = N- and NO–3 respectively. The binding
energy of –NH2 and –HC = N- were obviously decreased, which sug­
gested that Cu2+ bound to the nitrogen atom of –NH2 and –HC = N-.
X. Zhang et al.
Moreover, the decreased binding energy was due to the strong electro­
negativity of Cu2+ and the probable electron transfer between Cu2+ and
L [40]. Similarly, as can be seen from Fig. 4e, the O1s peak of L at 531.1
eV can be identified as C-O. The O1s spectrum of L + Cu2+ complex
(Fig. 4f) can be deconvoluted two peaks at 531.9 and 532.6 eV, corre­
sponding to the Cu–O bonding and the O − C bonding, respectively [41].
This suggested that the oxygen atom of phenolic-OH bound to Cu2+.
1
H NMR titration experiment was carried out in DMSO‑d6 solution,
as shown in Fig. S5. The peak at 9.958 ppm of OH (Ha) proton became
Fig. 4. (a) XPS spectra of L and complex L-Cu2+. (b) XPS spectra of Cu 2p. (c) N 1 s XPS spectra of L. (d) N 1 s XPS spectra of L + Cu2+. (e) O 1 s XPS spectra of L. (f) O
1 s XPS spectra of L + Cu2+.
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Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy 303 (2023) 123205
weak and almost disappeared with increasing concentration of Cu2+,
indicating that –OH of L can coordinate to Cu2+ by loss of H atom. The
peaks at 6.383 ppm, 8.141 ppm and 10.169 ppm are attributed to the
amide NH2 protons, –C = N– proton and –N–H– proton, respectively.
With the addition of Cu2+, the peak for –NH2 (Hd) protons at 6.383 ppm
shifted to 6.350 ppm and the intensity decreased. The –C = N– (Hb)
proton signal at 8.141 ppm shifted to 8.159 ppm, at the same time the
proton signal of the–N–H– (Hc) at 10.169 ppm shifted to 10.190 ppm,
which was owing to the reduction of local electron density by the
X. Zhang et al. Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy 303 (2023) 123205

Fig. 5. (a) UV–Vis absorption spectral response of L (10 μM) upon gradual addition of Mg2+ in acetonitrile. (1–4) = 0, 3.32, 6.62 and 9.9 μM. (b) Fluorescence
titration spectra of L (10 μM) upon gradual addition of Mg2+ in acetonitrile. (1–11) = 0, 0.999, 1.996, 2.99, 3.98, 4.98, 5.96, 6.95, 7.94, 8.92 and 9.9 μM. Inset: image
of L in the absence and presence of 1 eq Mg2+ observed under 365 nm UV light. (c) The linear relationship of the fluorescence intensity at 442 nm with Mg2+
concentration.

chelation of Cu2+ with the nitrogen atom of –C = N– and the oxygen 3.5.2. Binding capability
atom of phenolic-OH. Moreover, the broadening of 1H NMR spectral As seen in the Job’s plot (Fig. 6a), the turning point value of the ratio
signals may arise due to the usual paramagnetic effect of copper ion was 0.5. This means that the binding stoichiometry between L and Mg2+
[41]. These changes in 1H NMR spectra, FTIR spectra and XPS analysis was 1:1. The binding constant (K) of the L-Mg2+ complex can be ob­
suggested that Cu2+ may be coordinated by the oxygen atom of the tained from the fluorescence titration spectra by the Benesi–Hildebrand
phenolic –OH, the nitrogen atoms of –C = N– and the –NH2. equation (Fmax − F0)/(Fx − F0) = 1 + 1/K[M]n [34] as shown in Fig. 6b.
Where Fmax is the fluorescence intensity of L in the presence of Mg2+ at
complete interaction state, F0 is the intensity of L, Fx is the fluorescence
3.5. Turn on fluorescence response of L to Mg2+
intensity of L at an intermediate Mg2+ concentration. K is the binding
constant, [M] is the concentration of Mg2+, n is the number of Mg2+
3.5.1. Fluorescence and UV–Vis absorption spectral studies
bound to each L (here n = 1). The value of K was calculated to be 1.35
During the titration of L with Mg2+ in acetonitrile, UV–Vis absorp­
(±0.02) × 105 M− 1 (R2 = 0.998).
tion spectral changes were observed. As shown in Fig. 5a, L showed an
absorption peak at 315 nm in acetonitrile, this may be attributed to the
3.5.3. Competitive studies
transfer of the intramolecular charge (ICT) [28]. Upon gradual addition
The competitive experiments were conducted by measuring the
of Mg2+ to the L in acetonitrile solution, the original absorption peaks of
fluorescence intensity at 442 nm of L upon the addition of 1 eq. of
L decreased and a new broad absorption band was observed at 360 nm.
different metal ions, and then further addition of 1 eq. Mg2+. The result
A distinct isobestic point observed at 331 nm indicated that the reaction
is shown in Fig. S6, indicating that the various cations tested had little
between L and Mg2+ was clean and only one new product was produced
interference in the detection of Mg2+ except for Zn2+.
[42]. Based on the fluorescence titration spectra as shown in Fig. 5b, a
linear relationship between the fluorescence intensity at 442 nm and
3.5.4. Application in real water samples analysis
Mg2+ concentration was obtained in the range of 0.999–9.99 μM (R2 =
Due to the high selectivity of L for the detection of Mg2+, the
0.996) as is shown in Fig. 5c. The limit of detection (LOD) of L toward
application of L to real samples was studied to detect Mg2+. The tap
Mg2+ was calculated to be 9.5 nM according to the equation LOD = 3σ/
water samples in our laboratory were collected and filtered. The tap
S. The comparison of LOD of L with other fluorescence probers for Mg2+
water samples were diluted with acetonitrile(1:9 v/v)，then added tap
is shown in Table S3, indicating that L was better than those probers
water samples to the L, which had no influence on the fluorescence of L.
which reported by previous references.

Fig. 6. (a) Job’s plot of the L-Mg2+ complex in acetonitrile. The total concentration of L and Mg2+ was 10 μM. λex = 370 nm. (b) Benesi-Hildebrand plot to estimate
the binding constant of L-Mg2+complex.

6
X. Zhang et al.
Fig. 7. (a) XPS spectra of L and complex L-Mg2+. (b) XPS spectra of Mg 1 s. (c) N 1 s XPS spectra of L. (d) N 1 s XPS spectra of L + Mg2+. (e) O 1 s XPS spectra of L. (f)
O 1 s XPS spectra of L + Mg2+.
Therefore, different concentrations of Mg2+ were spiked in tap water
samples and the recovery percentage was analyzed. As shown in
Table S4, the Mg2+ concentration determined by L was close to that of
spiked. This result means that the L can be applied to detect Mg2+ in real
water samples.
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Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy 303 (2023) 123205
3.5.5. Binding mode study
The binding mode of Mg2+ with L was investigated by FTIR, XPS and
NMR spectral experiments. As shown in Fig. S7, the FTIR spectrum of L
contains peaks for the NH and NH2 groups at 3463.8 and 3151.4 cm− 1,
respectively. When L was coordinated to Mg2+ ion, the peak at 3151.4
cm− 1 of NH2 was disappeared, the peaks at 3463.8 of NH was shifted to
3491.9 cm− 1. The peaks at 1677.2 and 1599.2 cm− 1 of C = O and C = N
X. Zhang et al.
were shifted to 1692.39 and 1622.19 cm− 1, respectively. These findings
showed that the nitrogen atom of NH2 and C = N participated in the
binding to Mg2+. In addition, the FTIR spectrum of [L–Mg2+] also con­
tains an intense peak for the NO−3 group at 1384.5 cm− 1 [28,35],
inferring that NO−3 can coordinate to Mg2+.
The XPS spectra were shown in Fig. 7a, the photoemission peaks of
Mg 1 s, O 1 s, N 1 s, and C 1 s can be found in the samples, which is
attributed to the presence of L-Mg2+ complex [36]. As can be seen from
Fig. 7b, a peak at 1303.9 eV can be ascribed to Mg 1 s. N 1 s spectra of L
can be further deconvoluted into three peaks as can be seen from Fig. 7c,
the peaks at ~400.2 eV, ~399.5 eV and ~400.8 eV can be corresponded
to –NH2, –NH- and –HC = N-, respectively [38,39]. N 1 s spectra of L +
Mg2+ complex (Fig. 7d) can be further deconvoluted into four peaks at
~399.7 eV, ~399.3 eV, ~400 eV and ~406.8 eV, which can be attrib­
uted to –NH2, –NH-, –HC = N- and NO–3, respectively. The binding en­
ergies of –NH2 and –HC = N- were obviously decreased, it suggested that
Mg2+ bound to nitrogen atoms of –NH2 and –HC = N-. Similarly, as can
be seen from Fig. 7(e), the O1s peak of L at 531.1 eV can be identified as
C-O. The O1s of L + Mg2+ complex (Fig. 7f) can be deconvoluted two
peaks at 531.5, and 532.6 eV, which can be identified as the Mg-O
bonding and O − C bonding, respectively. Results showed that the ox­
ygen atom of phenolic-OH bound to Mg2+.
1 Supplementary data to this article can be found online at https://doi.
H NMR titration spectra carried out in DMSO‑d6 solution were
depicted in Fig. S8. The peak at 10.169 ppm was attributed to the –N–H–
proton. The peak at 6.383 ppm, 8.141 ppm and 9.958 ppm were
attributed to the amide NH2, –C = N– and OH protons, respectively.
With the addition of Mg2+ to L, the proton signal of the phenolic OH
(Ha) at 9.958 ppm became broader and the intensity decreased, and it
shifted to 10.002 ppm. The peak for NH2 (Hd) proton at 6.383 ppm
became broader and the intensity decreased. Meanwhile, the peak
shifted to 6.386 ppm. The –C = N– (Hb) proton signal at 8.141 ppm
shifted to 8.149 ppm, at the same time the proton signal of the–N–H–
(Hc) at 10.169 ppm shifted to 10.177 ppm, these were owing to the
reduction of local electron density, which was induced by the chelation
of Mg2+ with the nitrogen atom of –C = N– and the phenolic-OH. The
whole changes in 1H NMR spectra, FTIR spectra and XPS analysis indi­
cated that Mg2+ may be coordinated with the oxygen atom of the
phenolic –OH, the nitrogen atoms of –C = N– and the NH2.
4. Conclusions
In conclusion, we synthesized a Schiff base multiple correspondence
fluorescent probe (L) formed by the condensation of semicarbazide hy­
drochloride and salicylaldehyde. L has different fluorescence properties
in different solvents, which can be used in the highly sensitive and se­
lective detection of Cu2+ and Mg2+, respectively, by regulating solvent
system. L exhibited high selectivity and sensitivity toward Cu2+ in 10
mM, pH = 7.0 PBS: ethanol (99:1, v/v) solution in a fluorescence turn off
mode with the detection limit of 40.5 nM. While in acetonitrile, L
showed a high selectivity and sensitivity for Mg2+ detection in a fluo­
rescence turn on mode with the detection limit as low as 9.5 nM.
Furthermore, this fluorescent probe exhibited good practicality for lake
water and tap water samples without performing tedious sample pre­
treatments. Furthermore, the binding ratios and binding constants of
Cu2+ and Mg2+ with L were obtained by data analysis, and the possible
binding modes were elucidated by spectroscopic studies. We hope that
this probe can provide a strategy for designing more excellent multiple
correspondence fluorescent probe for metal ions in the future, and has
practical application in the environmental detection.
CRediT authorship contribution statement
Xianfa Zhang: Conceptualization, Investigation, Methodology, Data
curation, Writing – original draft. Shuaibing Yu: Investigation. Xuliang
Pang: Investigation. Xiaochen Ren: Investigation. Bo Zhang: Investi­
gation, Formal analysis. Jinming Kong: Funding acquisition, Writing –
8
Spectrochimica Acta Part A: Molecular and Biomolecular Spectroscopy 303 (2023) 123205
review & editing. Lianzhi Li: Conceptualization, Supervision, Funding
acquisition, Writing – review & editing.
Declaration of Competing Interest
The authors declare that they have no known competing financial
interests or personal relationships that could have appeared to influence
the work reported in this paper.
Data availability
Data will be made available on request.
Acknowledgements
This work was supported by the Scientific Research Foundation of
Liaocheng University, China (318011919), and the National Natural
Science Foundation of China (21974068).
Appendix A. Supplementary data
org/10.1016/j.saa.2023.123205.
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