A highly selective probe for fluorescence turn-on detection of Fe3+ ion based on a novel spiropyran derivative

Article abstract of DOI:10.1016/j.molstruc.2019.127481

A new spiropyran bearing multiple phenolic hydroxyl groups is synthesized. It acts as a chemosensor for selective detection of Fe³⁺ ions in both organic and aqueous solution. The optimal pH range for Fe³⁺ fluorescent detection by the sensor is pH 4.5–10.5. Upon addition of Fe³⁺ to the sensor solution the fluorescence emission intensity is enhanced by around 50 fold. Other common competitive metal ions including Li⁺, Na⁺, K⁺, Ag⁺, Cu²⁺, Fe²⁺, Zn²⁺, Co²⁺, Ni²⁺, Mn²⁺, Sr²⁺, Hg²⁺, Ca²⁺, Mg²⁺, Al³⁺ and Cr³⁺ do not substantially interfere with the selective detection of Fe³⁺ by the sensor. Changes in the fluorescence emission intensity with the equivalent ratio of Fe³⁺/sensor exhibits an approximate linear relationship, from which the concentration of Fe³⁺ ion may be estimated. The detection limit of Fe³⁺ ion by the sensor is measured to be 1.93 × 10–7 M. A plausible sensing mechanism for selective Fe³⁺ ion detection by the sensor is proposed based on the Job's plot measurement.

Full text of DOI:10.1016/j.molstruc.2019.127481

Journal Pre-proof
3+
A highly selective probe for fluorescence turn-on detection of Fe ion based on a
novel spiropyran derivative
Ruiqing Zhang, Luping Hu, Zhenxiang Xu, Yanxi Song, Hongqi Li, Xin Zhang,
Xucheng Gao, Mengxuan Wang, Chunying Xian
PII:
S0022-2860(19)31590-X
DOI:
https://doi.org/10.1016/j.molstruc.2019.127481
MOLSTR 127481
Reference:
To appear in: Journal of Molecular Structure
Received Date: 22 October 2019
Revised Date: 21 November 2019
Accepted Date: 25 November 2019
Please cite this article as: R. Zhang, L. Hu, Z. Xu, Y. Song, H. Li, X. Zhang, X. Gao, M. Wang, C. Xian,
3+
A highly selective probe for fluorescence turn-on detection of Fe ion based on a novel spiropyran
derivative, Journal of Molecular Structure (2019), doi: https://doi.org/10.1016/j.molstruc.2019.127481.
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3+
A highly selective probe for fluorescence turn-on detection of Fe ion
based on a novel spiropyran derivative
a
a
b
c
a,
a
Ruiqing Zhang , Luping Hu , Zhenxiang Xu , Yanxi Song , Hongqi Li *, Xin Zhang , Xucheng
a
a
a
Gao , Mengxuan Wang , Chunying Xian
a
Key Laboratory of Science and Technology of Eco-Textiles, Ministry of Education, College of
Chemistry, Chemical Engineering & Biotechnology, Donghua University, 2999 North Renmin
Road, Shanghai 201620, P. R. China.
b
Penglai Xinguang Pigment Chemical Co., Ltd, Penglai 265601, P. R. China
c
School of Environmental Science and Engineering, Donghua University, 2999 North Renmin
Road, Shanghai 201620, P. R. China.
ABSTRACT
A new spiropyran bearing multiple phenolic hydroxyl groups is synthesized. It acts as
3
+
a chemosensor for selective detection of Fe ions in both organic and aqueous
3
+
solution. The optimal pH range for Fe fluorescent detection by the sensor is pH
3
+
4
.5‒10.5. Upon addition of Fe to the sensor solution the fluorescence emission
intensity is enhanced by around 50 fold. Other common competitive metal ions
+
+
+
+
2+
2+
2+
2+
2+
2+
2+
2+
2+
2+
including Li , Na , K , Ag , Cu , Fe , Zn , Co , Ni , Mn , Sr , Hg , Ca , Mg ,
3
+
3+
3+
Al and Cr do not substantially interfere with the selective detection of Fe by the
sensor. Changes in the fluorescence emission intensity with the equivalent ratio of
3
+
Fe /sensor exhibits an approximate linear relationship, from which the concentration
3
+
3+
of Fe ion may be estimated. The detection limit of Fe ion by the sensor is
–
7
3+
measured to be 1.93 × 10 M. A plausible sensing mechanism for selective Fe ion
detection by the sensor is proposed based on the Job’s plot measurement.
3
+
Keywords: Fluorescence; probe; spiropyran; Fe
1
. Introduction
As a kind of transitional metal, iron is the metal element of the second highest
crust content and is also the essential trace element in human body. It mainly exists in
the form of hemoglobin and has great effect in oxygen supply of human body. On the
3
+
other hand, excessive intake of Fe may result in a disturbance of physiological
function and several diseases [1‒4]. Therefore convenient and effective identification
3
+
and detection of Fe ions in the organism and in the environment is of high
significance. In this respect selective and sensitive fluorescent probes for rapid
3
+
detection of Fe ions aroused much interest from scientists and a number of
3
+
fluorescent Fe probes have been developed by utilizing various fluorogenic
platforms such as coumarin [5‒9], rhodamine [10‒12], 1,8-naphthalimide [13‒16],

pillar[5]arene [17,18], quantum dots [19‒21], and metal-organic frameworks (MOFs)
3
+
als [22‒24]. Chemosensors for fluorescent or colorimetric detection of Fe ions based
on photochromic spiropyran scaffold have also been reported [25‒27]. However, the
3
+
selectivity of these sensors toward Fe ions is substantially weakened by other
3+
3+
trivalent ions Al and Cr . In view that spiropyran is an extraordinarily versatile
photochrome [28] and spiropyran-based fluorescent and/or colorimetric probes [29]
have been developed for highly selective and sensitive detection of various analytes
+
2+
2+
2+
2+
2+
including pH [30‒32], Li [33], Mg [34], Ca [35], Co [36], Cu [37], Zn [38],
2
+
2+
3+
3+
3+
‒
‒
‒
Hg [39], Pb [40], Al [41], Cr [42], La [43], CN [44], F [45], Cl [46],
pyrophosphate [47], CO [48], H O [49], H S [50], H S/SO [51], hydrazine [52],
2
2
2
2
2
2
cysteine (Cys)/homocysteine (Hcy) [53], γ-glutamyl-cysteinyl-glycine (GSH) [54],
β-amyloid peptide oligomers [55], lysozyme [56], DNA G-quadruplexes [57], aniline
pollutants [58], acidic food spoilage [59], ultraviolet (UV) light intensity [60], and
mechanical stress [61], we envisage to design and synthesize a fluorescent
3
+
chemosensor which can detect Fe ions selectively and sensitively even in the
3
+
3+
presence of other trivalent ions Al and Cr based on spiropyran, following our
recent work concerning study on a spiropyran-appended polysiloxane which can be
3
+
+
2+
used for probing pH changes and Fe ions and sequential detection of Ag and Hg
ions [62]. Herein we report the synthesis, characterization and recognition behavior of
3
+
the new spiropyran-based fluorogenic Fe sensor.
2. Experimental section
2
.1. Materials and instrumentation
All materials and solvents employed in this study were of analytical grade. All
chemical reagents were purchased from commercial sources and used as received
unless other statements. 1,2,3,3-Tetramethylindolindolenium iodide was synthesized
according to literature method [63]. Melting point was determined on a WRS-2A
1
capillary melting apparatus and the quoted temperatures were uncorrected. H NMR
1
3
and C NMR spectra were recorded on a Bruker AM 400 spectrometer. DMSO-d or
6
CDCl was used as solvent and chemical shifts recorded were internally referenced to
3
Me Si (0 ppm). IR spectra were obtained on a Thermo Electron Corporation Nicolet
4
3
80 FT-IR spectrophotometer from Thermo Fisher Scienific Company. Mass spectra
were obtained on an Agilent 6000 LC-MS instrument (ESI, pos mode, 70 to 1000 amu).
Fluorescence spectra were recorded on a FS5 spectrofluorometer from Edinburgh

Instruments Co., Ltd. UV-Vis spectra were recorded on a UV-1800 spectrophotometer
from Shimadzu Instruments (Suzhou) Co., Ltd..
2
.2. Synthesis of sensor SP
1
,2,3,3-Tetramethylindolindolenium iodide (6.35 g, 21 mmol) was dissolved in
anhydrous methanol (100 mL) and heated to reflux. Then triethylamine (1 mL) was
added and the solution was stirred for 15 min. 5-Nitrosalicylaldehyde (3.34 g, 20
mmol) was added and the mixture was refluxed for 2 h. After reaction the mixture
was allowed to cool to room temperature and methanol was removed by evaporation.
The formed orange solid was recrystallized with dichloromethane/ethanol. Yellow
solid (5.88 g) of the target product 1,3,3-trimethyl-6´-nitrospiro[chromene-2,2´-
indoline] was obtained in 91% yield after filtration and drying. Mp 178–180 °C
1
(
literature [64] value: 178–179 °C). H NMR (400 MHz, CDCl ) δ 8.05–8.03 (m, 2H),
3
7
.25–7.23 (m, 1H), 7.12 (dd, J = 7.3, 1.3 Hz, 1H), 6.99–6.88 (m, 2H), 6.80 (d, J = 8.6
Hz, 1H), 6.59 (d, J = 7.7 Hz, 1H), 5.89 (d, J = 10.3 Hz, 1H), 2.77 (s, 3H), 1.32 (s, 3H),
.22 (s, 3H).
,3,3-Trimethyl-6´-nitrospiro[chromene-2,2´-indoline] (6.45 g, 20 mmol) and
SnCl (18.96 g, 0.10 mol) were added to a flask under N atmosphere. Anhydrous
1
1
2
2
ethanol (100 mL) was added and the reaction mixture was heated to reflux for 2 h.
After reaction the mixture was cooled to room temperature and filtered. The filtrate
was concentrated and 5% aqueous NaOH was added to the residue. The organic phase
was extracted with dichloromethane for three times, washed with water and dried.
After filtration the solvent was removed by evaporation and the residue was purified
by column chromatography with petroleum ether/ethyl acetate (3:1, v/v) as eluent to
give the reduction product 1,3,3-trimethyl-6´-aminospiro[chromene-2,2´-indoline]
1
(
3.18 g) as brown solid in 54% yield. Mp 139–140 °C. H NMR (400 MHz, CDCl ) δ
3
7.16–7.14 (m, 1H), 7.05 (d, J = 8.3 Hz, 1H), 6.81–6.79 (m, 1H), 6.74 (d, J = 8.8 Hz,
1H), 6.55–6.44 (m, 3H), 6.41 (d, J = 7.9 Hz, 1H), 5.67 (d, J = 10.9 Hz, 1H), 2.85 (s,
3H), 1.45 (s, 3H), 1.30 (s, 3H).
1
,3,3-Trimethyl-6´-aminospiro[chromene-2,2´-indoline] (0.73 g, 2.5 mmol) and
2
,5-dihydroxybenzaldehyde (0.33 g, 2.4 mmol) were dissolved in dichloromethane
(20 mL). To the solution was added 5 drops of acetic acid and the mixture was stirred
at room temperature for 1 h. After reaction the mixture was washed with water and
the organic layer was separated then dried. The filtration was concentrated and the
residue was purified by column chromatography with petroleum ether/ethyl acetate

(5:1, v/v) as eluent to afford the target sensor SP (0.30 g) as red-brown solid in 30%
yield. Mp 146–148 °C. IR (KBr pellet): ν 3318, 1644, 1609, 1481, 1450, 1270, 953
–
1 1
cm . H NMR (400 MHz, CDCl ) δ 8.43 (s, 1H), 7.19 (s, 1H), 7.14–7.12 (m, 2H),
3
7
1
1
1
1
.03–7.00 (m, 3H), 6.97 (d, J = 2.5 Hz, 1H), 6.85–6.76 (m, 7H), 6.69 (d, J = 8.6 Hz,
H), 6.47 (d, J = 7.7 Hz, 1H), 5.69 (d, J = 10.2 Hz, 1H), 2.68 (s, 3H), 1.25 (s, 3H),
1
3
.11 (s, 3H). C NMR (101 MHz, CDCl ) δ 158.49, 154.06, 152.88, 147.06, 146.78,
3
39.68, 135.58, 127.98, 126.62, 121.17, 120.49, 119.49, 118.55, 118.27, 118.22,
18.18, 116.85, 116.09, 114.79, 105.83, 103.63, 50.85, 27.91, 24.85, 19.12. MS (ESI):
+
m/z 413 (M + 1).
2
.3. General UV-vis and fluorescence spectra measurements
All spectral analyses were accomplished at room temperature in 0.1 M Tris-HCl
buffer solution (DMF/H O = 9:1, v/v, pH = 7.10). The concentration of sensor SP and
2
–
5
metal ions was 4 × 10 M for UV-vis and fluorescence measurement. Sensor SP was
dissolved in DMF and solutions of metal ions were prepared with nitrate or chloride
salts in water. Each sample was added to a 10 mL volumetric flask by using a 20‒200
ꢀ
L liquid transfer gun and the solution was well mixed before the spectra were
measured. Fluorescence titration experiments were performed in 2 mL cuvette by
successive addition of prepared solutions at an excitation wavelength of 378 nm with
both excitation and emission slits of 5.0 nm. UV-vis absorption spectra were
measured in the range of 290‒700 nm.
3
. Results and discussions
3
.1. Synthesis and structural characterization of sensor SP
The novel spiropyran-based sensor SP was synthesized by a three-step protocol
from 1,2,3,3-tetramethylindolindolenium iodide and 5-nitrosalicylaldehyde, as
showed in Scheme 1. Condensation between 1,2,3,3-tetramethylindolindolenium
iodide and 5-nitrosalicylaldehyde in the presence of triethylamine produced the target
product 1,3,3-trimethyl-6´-nitrospiro[chromene-2,2´-indoline] in 91% yield. The
1
nitrospiropyran was characterized by both melting point measurement and H NMR
spectrum. The data were in good accordance with those reported in literature [64].
Reduction of the nitrospiropyran to the corresponding aminospiropyran by SnCl2
afforded a 54% yield. The structure of the obtained aminospiropyran compound was
1
characterized by melting point measurement and H NMR spectrum and the spectral

data were in good accordance with those reported in literature [65]. Imination reaction
between 2,5-dihydroxybenzaldehyde and the aminospiropyran catalyzed by acetic
acid in dichloromethane afforded sensor SP in 30% yield. The chemical structure of
1
13
the sensor SP was fully characterized by H NMR, C NMR, IR and mass spectrum.
All of the data in the spectra were in good accordance with the structure.
Scheme 1
3
+
3
.2. Effect of medium on fluorescent detection of Fe ions by sensor SP
3
+
‒5
Fluorescence spectra of sensor SP and Fe solution (4 × 10 M) in different solvents
including EtOH, MeOH, MeCN, DMF and DMSO at an excitation wavelength of 378
nm were measured and showed in Fig. 1. It was found that the solution exhibited the
maximum fluorescence emission at around 450 nm and the fluorescence intensities in
DMF and DMSO were much higher that those in other solvents EtOH, MeOH and
MeCN. Based on the results DMF was chosen as the solvent in the following
determination, considering that DMSO has a higher viscosity and strongly irritating
odor though DMSO shows a little higher fluorescence intensity that that of DMF.
Fig. 1
It is important for a practical metal ion probe to be usable in an aqueous medium.
3
+
‒5
Therefore the fluorescence spectra of solutions containing Fe ion (4 × 10 M) and
‒
5
sensor SP (4 × 10 M
) in DMF/H O of different ratios (DMF/H O = 9:1, 8:2, 7:3, 6:4,
2 2
5
:5, 4:6, 3:7, 2:8, 1:9, v/v) were determined at an excitation wavelength of 378 nm and
were showed in Fig. 2. It could be observed from Fig. 2 that the solution exhibited the
highest fluorescence intensity in 9:1 DMF/H O. With the increase in the ratio of water,
2
the fluorescence emission intensity decreased sharply. The solution in 7:3 DMF/H O
2
was fairly fluorescent and further increase in the water content led to very weak
fluorescence of the solution. Thus the optimal sensing medium was 9:1 DMF/H O,
2
which was used in the following determination.
Fig. 2
The fluorescence spectra of sensor SP before and after addition of common metal
+
+
+
+
2+
2+
2+
2+
2+
2+
2+
2+
2+
2+
ions (Li , Na , K , Ag , Cu , Fe , Zn , Co , Ni , Mn , Sr , Hg , Ca , Mg ,
3
+
3+
3+
Al , Cr , Fe ) in DMF/H O (9:1, v/v) were measured and showed in Fig. 3. The
2
very weak fluorescence emission of the probe SP appeared at 450 nm when it was

excited at a wavelength of 378 nm. The fluorescence intensity of the probe SP was
3
+
3+
enhanced by addition of Fe and Al while addition of other metal ions failed to
3
+
3+
cause substantial change in the fluorescence emission. Addition of Fe and Al
induced an enhancement in the fluorescence intensity of the probe SP by around 50
fold and 8 fold, respectively. Therefore the probe SP can be used for selective
3
+
detection of Fe in an aqueous medium.
Fig. 3
.3. Anti-interference ability of sensor SP for the fluorescent detection of Fe ions
3
+
3
In order to investigate the interference of competitive metal ions on the sensing
3
+
selectivity of the probe SP toward Fe ion, changes in fluorescence intensity at 450 nm
3
+
of sensor SP upon addition of Fe ion (1 equivalent) and/or other competitive metal
ions (1 equivalent) were recorded and showed in Fig. 4. It was visible that to the
3
+
2+
3+
fluorescence turn-on probe SP for Fe ion, addition of 1 equivalent of Cu or Al ion
reduced the fluorescence intensity by about 20%. Other competitive ions did not cause
3
+
obvious interference with the selective detection of Fe ion.
Considering that competitive metal ions may exist in a large amount and mix with
3
+
the analyte such as Fe ion, changes in fluorescence intensity at 450 nm of sensor SP
3
+
upon addition of Fe ion (1 equivalent) and other competitive metal ions (5 equivalent)
3
+
2+
2+
were measured and showed in Fig. 5. In this case the excessive Al , Hg and Fe
3
+
ions interfered with the selective detection of Fe by sensor SP, bringing about
decrease in the fluorescence intensity by 40‒43%. While the presence of excessive
2
+
3+
Cu ion caused serious interference with the selective detection of Fe by sensor SP,
reducing the fluorescence intensity by nearly 84%. Other competitive ions caused
3
+
relatively small interference to the selective detection of Fe ion.
Fig. 4
Fig. 5
The real samples especially waste water samples usually contain multiple metal
ions. Therefore the anti-interference ability to multiply mixed metal ions of a sensor
for a specific metal ion reflects the ‘real’ anti-interference ability of the sensor.
3
+
Changes in fluorescence intensity at 450 nm of sensor SP upon addition of Fe (1
equivalent) and 5 kinds of other competitive metal ions together (each 1 equivalent) in

DMF/H O (9:1, v/v) were determined and showed in Fig. 6. Groups containing 5 kinds
2
of competitive metal ions for anti-interference test were picked up at random on the
premise to ensure each group contains a monovalent and a trivalent metal ion. The
+
2+
2+
2+
3+
following groups of metal ions were chosen for the test: Li , Hg , Zn , Ni and Al
+
2+
2+
2+
3+
+
2+
2+
2+
3+
in a group; Na , Cu , Ca , Mg and Cr in a group; K , Fe , Mn , Sr and Al in a
+
2+
2+
2+
3+
+
2+
2+
2+
3+
group; Ag , Zn , Co , Fe and Cr in a group; and Ag , Cu , Sr , Mn and Al in
a group. It can be seen that each group containing 5 kinds of competitive metal ions
exhibits only a little decrease in the fluorescence intensity compared to the sensor
3
+
SP/Fe system, indicating that the multiply mixed competitive metal ions, containing
2
+
2+
2+
3+
2+
3+
2+
3+
Cu or Fe alone, or Hg /Al , Fe /Al , and Cu /Al together, had negligible
3
+
interference to the selective detection of Fe ion by sensor SP.
2
+
The paramagnetic property of Cu would bring about a fluorescence quenching
2
+
2+
effect and Fe /Hg ions also undergo electron or energy transfer to result in a certain
degree of fluorescence quenching. Generally the fluorescence quenching phenomenon
is positively correlated with the concentration of the quenching ions [66‒69]. This is
2
+
2+
2+
why Cu , Hg and Fe ions cause significant decrease in the fluorescence intensity
when these ions are present alone in 5-fold equivalent while the mixed metal ions (each
having a much lower concentration) only bring about a little interference to the
fluorescence intensity.
3
+
3+
The enhancement in fluorescence intensity of the probe SP by Fe and Al
3
+
reaches about 50 fold and 8 fold, respectively. When Al exists in large quantities, the
3
+
response of the sensor SP to Fe is weakened heavily and results in the decrease of the
overall fluorescence intensity. However, the total fluorescence intensity is only slightly
3
+
3+
reduced if Al and Fe ions are present in equal amount. Overall, the sensor SP
3
+
exhibited good anti-interference ability for specific detection of Fe ion.
Fig. 6
3
+
3
.4. Effect of UV irradiation and pH on sensor SP for the detection of Fe ions
‒
5
Fluorescence spectra of sensor SP (4 × 10 M in DMF/H O, 9:1, v/v) before and
2
after UV light irradiation at an excitation wavelength of 378 nm were recorded as
showed in Fig. 7. Solution of probe SP was orange-yellow. After the solution was
irradiated with UV light for 2 h the color did not change obviously. However, the
fluorescence emission intensity at around 450 nm was enhanced greatly after the
solution of probe SP was irradiated with UV light for 2 h. It indicated that the weakly
fluorescent ring-closed spirobenzopyran isomer gradually transformed into the strongly
fluorescent ring-opened merocyanine isomer upon irradiation with UV light, in
consistence with the previously reported results observed on photochromic spiropyrans

3
+
[
33‒37]. Addition of 1 equivalent of Fe to probe SP led to obvious enhancement in
the fluorescence emission intensity. Irradiation of the solution with UV light for 2 h did
not cause much increase in the fluorescence emission intensity, as showed in Fig. 8. It
3
+
might indicate that the probe SP was readily ring-opened by addition of Fe ion,
resulting in intense enhancement of the fluorescence emission. The sensor SP
3
+
underwent a ring-opening process in the presence of Fe ion then the ring-opened
3
+
merocyanine isomer complexed with Fe ion [34‒38].
Fig. 7
Fig. 8
3
+
Effect of pH on the fluorescent sensing behavior of probe SP toward Fe ion was
investigated by measurement of the fluorescence spectra of sensor SP containing 1
3
+
equivalent of Fe in DMF/H O (7:3, v/v) at different pH values (pH = 1.5, 2.5, 3.5,
2
4
.5, 5.5, 6.5, 7.5, 8.5, 9.5, 10.5, 11.5, 12.5, 13.5) and an excitation wavelength of 378
nm, as showed in Fig. 9. It was clearly visible that the solution exhibited high
fluorescence emission intensity in the pH range of pH 4.5‒10.5. The fluorescence
intensity decreased by 50% more or less when the pH value was below 3.5. Strong
alkaline (pH > 11.5) media resulted in fluorescence quenching of the solution. The
3
+
results reveal that the probe SP can be used to recognize Fe in a wide pH range except
3
+
strong alkaline conditions, superior to many fluorescent probes for Fe ion [9‒13]. The
3
+
optimal pH range for Fe fluorescent detection by probe SP is pH 4.5‒10.5.
Fig. 9
3
+
3
.5. Detection limit of sensor SP for sensing Fe ions
3
+
Fluorescence spectra of sensor SP containing different concentrations of Fe ion
4 µM, 6 µM, 8 µM, 10 µM, 12 µM, 14 µM, 16 µM, 18 µM, 20 µM) in DMF/H O (9:1,
(
2
v/v) at an excitation wavelength of 378 nm were measured and the changes in the
3
+
fluorescence emission intensity of the solution with Fe ion concentration were
showed in Fig. 10. It was undoubted that the fluorescence intensity of the solution
3
+
increased gradually with increase in Fe ion concentration. A line could be drawn
3
+
based on the intensity of the fluorescence emission band and the concentration of Fe
3
+
ion within Fe concentration range from 4 µM to 8 µM and from 8 µM to 20 µM, as
showed in Fig. 10. The first linear fitting equation can be depicted as Y = 47002X +

1
01413 with R = 0.99488 (4 µM‒8 µM), from which the detection limit can be
–
7
calculated to be 1.93 × 10 M. The second linear fitting equation can be depicted as Y
14055X + 379360 with R = 0.99352 (10 µM‒20 µM). Thus sensor SP may be
=
3
+
utilized for quantitative determination of Fe ion within a concentration range of
4
‒20 µM.
Fig. 10
.6. Study on Fe ion sensing mechanism by sensor SP
3
+
3
3
+
Investigation on possible mode of complexation between probe SP and Fe ion
was carried out by determination of the Job's plot. The total concentration of probe SP
3
+
and Fe ion was kept at 0.2 mM and changes in the fluorescence intensity of the
3
+
solution containing sensor SP and Fe ion with the molar ratio of sensor SP were
measured as showed in Fig. 11. It can be seen from the Job’s plot that the maximum
fluorescence emission intensity appears at 0.25. Thus it can be inferred that probe SP
3
+
and Fe ion form a complex in 1:3 ratio. The plausible mode of the complexation
3
+
between sensor SP and Fe ion is showed in Fig. 12, which reveals the isomeric
structure of spiropyran SP (closed, weakly fluorescent spiropyran isomer) and the
3
+
3+
Fe -induced ring-opened Fe complex (opened, strongly fluorescent merocyanine
3
+
isomer). In other works concerning fluorescent chemosensors for Fe ion, the
3
+
reported stoichiometry ratio between the sensor and Fe ion is 1:1 generally [12–15].
3
+
In our case the formation of 1:3 SP–Fe complexation may be attribute to the specific
SP structure bearing multiple phenolic hydroxyl groups which readily complex with
3
+
Fe ions.
Fig. 11
Fig. 12
4
. Conclusions
A new spiropyran-based fluorescent probe has been developed and investigated.
The probe can transform from weakly fluorescent closed loop to strongly fluorescent
3
+
opened loop upon UV irradiation or addition of Fe ion. The fluorescence emission of
3
+
the probe solution is intensely enhanced by adding Fe ion in DMF/H O (9:1, v/v). The
2
fluorescence enhancement is not obviously interfered by addition of one or more other

3
+
common competitive metal ions. The probe can be utilized for sensing Fe ion in a
3
+
wide pH range except strong alkaline conditions and the optimal pH range for Fe
fluorescent detection is pH 4.5‒10.5. Investigation on the changes in the fluorescence
3
+
emission intensity of the solution containing the sensor and Fe ion with the
3
+
equivalent ratio of Fe /sensor reveals an approximate linear relationship. Thus the
3
+
sensor may be useful for quantitative determination of Fe ion within a concentration
3
+
–7
range of 4‒20 µM. The detection limit of Fe ion by the sensor is 1.93 × 10 M.
3
+
Job’s plot indicates complexation between the sensor and Fe ion in a stoichiometry
ratio of 1:3. It may be attribute to the characteristic sensor structure bearing multiple
3
+
phenolic hydroxyl groups which readily complex with Fe ions. These findings well
demonstrate the versatility of spiropyrans as selective fluorescent probes for various
3
+
metal ions including Fe ion.
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OH
Et N/MeOH
reflux
3
CHO
+
91%
N O
NO2
N
I
NO2
SnCl /HCl
EtOH, reflux
2
2,5-dihydroxybenzaldehyde
AcOH/CH Cl
2
2
54%
N O
HO
NH2
30%
N O
N
OH
SP
Scheme 1. Synthetic route of chemosensor SP

3
+
‒5
Fig. 1. Fluorescence spectra of solutions containing Fe ion (4 × 10 M) and sensor
‒5
SP (4 × 10 M) in different solvents (EtOH, MeOH, MeCN, DMF, DMSO) at an
excitation wavelength of 378 nm.

Fig. 2. Fluorescence spectra of solutions containing Fe3+ ion (4 × 10 M) and sensor
‒5
‒5
SP (4 × 10 M) in DMF/H
2
O of different ratios (DMF/H O = 9:1, 8:2, 7:3, 6:4, 5:5,
2
4
:6, 3:7, 2:8, 1:9, v/v) at an excitation wavelength of 378 nm.

‒
5
Fig. 3. Fluorescence emission spectra of sensor SP (4 × 10 M) before and after
‒5
+
+
+
+
2+
2+
addition of solution (4 × 10 M) of different metal ions (Li , Na , K , Ag , Cu , Fe ,
2
+
2+
2+
2+
2+
2+
2+
2+
3+
3+
3+
Zn , Co , Ni , Mn , Sr , Hg , Ca , Mg , Al , Cr , Fe ) in DMF/H
at an excitation wavelength of 378 nm.
2
O (9:1, v/v)

Fig. 4. Changes in fluorescence intensity at 450 nm of sensor SP upon addition of
Fe3+ (1 equivalent) and/or other competitive metal ions (1 equivalent). Black bars
3
+
denote responses of individual metal ions while red bars show responses of Fe ion in
the presence of other coexisting metal ions (in 9:1 DMF/H
wavelength of 378 nm).
2
O, at an excitation

Fig. 5. Changes in fluorescence intensity at 450 nm of sensor SP upon addition of
3
+
Fe (1 equivalent) and other competitive metal ions (5 equivalent) in DMF/H
v/v) at an excitation wavelength of 378 nm.
2
O (9:1,

Fig. 6. Changes in fluorescence intensity at 450 nm of sensor SP upon addition of
Fe3+ (1 equivalent) and 5 kinds of other competitive metal ions together (each 1
3
+
equivalent) in DMF/H O (9:1, v/v) at an excitation wavelength of 378 nm: bar 1. Fe ;
2
3
+
+
2+
2+
2+
3+
3+
+
2+
2+
2+
3+
bar 2. Fe , Li , Hg , Zn , Ni , Al ; bar 3. Fe , Na , Cu , Ca , Mg , Cr ; bar 4.
3
+
+
2+
2+
2+
3+
3+
+
2+
2+
2+
3+
3+
Fe , K , Fe , Mn , Sr , Al ; bar 5. Fe , Ag , Zn , Co , Fe , Cr ; bar 6. Fe ,
+
2+
2+
2+
3+
Ag , Cu , Sr , Mn , Al .

‒5
Fig. 7. Fluorescence spectra of sensor SP (4 × 10 M in DMF/H
2
O, 9:1, v/v) before
and after UV light irradiation for 2 h at an excitation wavelength of 378 nm.

‒
5
3+
Fig. 8. Fluorescence spectra of sensor SP (4 × 10 M) containing 1 equivalent of Fe
in DMF/H O (9:1, v/v) before and after UV light irradiation for 2 h at an excitation
wavelength of 378 nm.
2

‒
5
3+
Fig. 9. Fluorescence spectra of sensor SP (4 × 10 M) containing 1 equivalent of Fe
in DMF/H
0.5, 11.5, 12.5, 13.5) with an excitation wavelength of 378 nm.
2
O (7:3, v/v) at different pH values (pH = 1.5, 2.5, 3.5, 4.5, 5.5, 6.5, 7.5, 8.5, 9.5,
1

‒
5
Fig. 10. Changes in fluorescence intensity of sensor SP (4 × 10 M) containing
3+
different concentrations of Fe (4 µM, 6 µM, 8 µM, 10 µM, 12 µM, 14 µM, 16 µM,
8 µM, 20 µM) in DMF/H O (9:1, v/v) at an excitation wavelength of 378 nm.
1
2

3
+
Fig. 11. Job’s plot obtained from fluorescence intensity of sensor SP and Fe ion
3
+
solution ([SP] + [Fe ] = 200 μM, in 9:1 DMF/H O, v/v) at an excitation wavelength
2
of 378 nm.

O
Fe3+
N
HO
Fe3+
O
N O
N
N
O
Fe3+
OH
Fe3+
SP
weakly fluorescent
strongly fluorescent
3
+
Fig. 12. Plausible mode of complexation between sensor SP and Fe ion.