G Model
CCLET 2989 1–5
2
Y.-Y. Huang et al. / Chinese Chemical Letters xxx (2014) xxx–xxx
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X-ray crystal data were collected on the Bruker Smart APEX II CCD
diffractometer. Elemental analyses were measured on a Vario EL III
analyzer. IR spectra were acquired on the Bruker Tensor 27
spectrometer in KBr disks. Mass spectra were performed on the
Bruker micrOTOF-Q II ESI-Q-TOF LC/MS/MS Spectrometer. NMR
spectra were recorded on the Varian Inova-400 MHz spectrometer
(at 400 MHz for 1H and 100 MHz for 13C). The chemicals and
reagents were obtained from Sigma-Aldrich Co., LLC. Analytical
thin layer chromatography was performed using Merck 60 GF254
silica gel coated glass plates. Silica gel (0.200–0.500 mm, 60A, J&K
Scientific Ltd.) was used for column chromatography.
Synthesis of the probes: Fluorescein hydrazide was synthesized
according to Ref. [12]. Fluorescein hydrazine (3.46 g, 0.01 mol) was
then dissolved in 50 mL ethanol. The cinnamyl aldehydes
(0.01 mol) were added in 30 min. The reaction mixture was
refluxed for 12 h. After cooling to the room temperature, the
solvent was evaporated and then washed with anhydrous ether to
obtain a white or light yellow powder.
J = 7.9 Hz), 6.10 (s, 1H), 6.01 (s, 1H). MS (ESI) m/z 333.0757 [M+H]+,
Calcd. for C20H12O5 332.0685.
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General procedure: The stock solutions of probes were prepared
at 500
mmol/L concentration. The solutions of ions, including LiCl,
NaCl, KCl, BaCl2, MgCl2, CaCl2, CdCl2, MnCl2, CoCl2, ZnCl2, NiCl2,
CuCl2, HgCl2, PbCl2, CrCl3, FeCl3, AlCl3, SnCl4, NaNO3, Na2CO3,
NaHCO3, Na2SO4, Na3PO4, Na2HPO4, NaH2PO4, NaOAc, H2O2, and
NaOCl, were also prepared at 500
mmol/L in MeOH/H2O (4:6, v/v).
The fluorescence intensity was recorded at 533 nm with an
excitation wavelength at 490 nm. The absorption data were
recorded at 517 nm.
Crystal growth and conditions: White single crystals of the
probes were obtained at room temperature from a mixed solution
of CH2Cl2–CH3CN by slow evaporation and then mounted on the
goniometer of a single crystal diffractometer. The crystal data have
˚
been collected at 296 K by using Mo Ka radiation (l = 0.710713 A)
and the w/v scan mode and Analyzed for Lorentz and polarization
effect (SADABS). The structure was solved using the direct method
and refined by full-matrix least-squares fitting on F2 by SHELX-97.
Fluorescent imaging: Human Osteosarcoma MG-63 cells were
cultured in Dulbecco’s Modified Eagle Medium (DMEM) supple-
mented with 10% FBS, at 37 8C in a humidified atmosphere of 5%
CO2 and 95% air. The cells were then cultured for 2 h until they
plated on glass-bottomed dishes. The growth medium was then
removed and the cells were washed with DMEM without FBS and
Probe 1: White powder. mp: 289–290 8C. Anal. Calcd. for
C29H20N2O4: H: 4.38, C: 75.64, N: 6.08. Found: H: 4.36, C: 75.69, N:
6.09. 1H NMR (400 MHz, DMSO-d6, TMS):
d 9.92 (s, 2H), 8.93 (d, 1H,
J = 9.1 Hz), 7.88 (d, 1H, J = 7.2 Hz), 7.58 (dd, 4H, J = 14.4, 7.3 Hz),
7.31 (dd, 3H, J = 15.0, 7.5 Hz), 7.04 (d, 1H, J = 7.0 Hz), 6.95 (d, 1H,
J = 16.0 Hz), 6.76 (dd, 1H, J = 15.9, 9.2 Hz), 6.64 (s, 2H), 6.49 (dd, 4H,
J = 21.3, 8.6 Hz). 13C NMR (100 MHz, DMSO-d6, TMS):
d 163.9,
158.5, 153.0, 151.9, 151.2, 140.4, 135.6, 133.9, 129.0, 128.7, 128.4,
128.0, 127.3, 126.2, 123.6, 123.2, 112.4, 110.0, 102.6, 65.2. MS (ESI)
m/z 461.1489 [M+H]+, Calcd. for C29H20N2O4 460.1423.
incubated with 20
three times with PBS and imaged. Then the cells were supple-
mented with 200 mol/L NaOCl in the growth medium for 4 h at
mmol/L of the probes for 4 h at 37 8C, washed
m
Probe 2: White powder. mp: 277–278 8C. Anal. Calcd. for
C30H22N2O4: H: 4.67, C: 75.94, N: 5.90. Found: H: 4.66, C: 75.99, N:
37 8C and imaged [13].
5.89. 1H NMR (400 MHz, DMSO-d6, TMS):
d
9.90 (s, 2H), 8.85 (s, 1H),
3. Results and discussion
139
7.90 (d, 1H, J = 7.3 Hz), 7.69–7.55 (m, 2H), 7.37 (d, 4H, J = 6.5 Hz),
7.28 (s, 1H), 7.13 (d, 1H, J = 7.3 Hz), 6.67 (d, 3H, J = 18.6 Hz), 6.46 (s,
The probes were prepared by the synthetic route outlined in
Scheme 1. The facile formylation reaction of fluorescein and
hydrazine hydrate gave fluorescein hydrazide, which was further
treated with cinnamyl aldehydes in EtOH solution at reflux
temperature to yield the target probes. The probes were
characterized by X-ray single crystal analysis in combination with
elemental analysis, IR, NMR, and MS spectra. The X-ray crystal
structural investigation clearly showed the special Schiff base
functional group (Fig. 1). The structural differences between the
probes are a result of the different substitution groups, which were
further investigated to have obvious effects on the fluorescent
properties.
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4H), 1.80 (s, 3H). 13C NMR (100 MHz, DMSO-d6, TMS):
d 163.3,
158.6, 154.5, 152.4, 150.2, 137.8, 136.1, 134.8, 133.8, 129.4, 129.1,
128.4, 128.1, 127.8, 123.9, 123.2, 112.3, 110.4, 102.5, 65.4, 11.9. MS
(ESI) m/z 475.1652 [M+H]+, Calcd. for C30H22N2O4 474.1582.
Probe 3: Yellow powder. mp: 284–285 8C. Anal. Calcd. for
C29H19BrN2O4: H: 3.55, C: 64.58, N: 5.19. Found: H: 3.54, C: 64.59,
N: 5.21. 1H NMR (400 MHz, DMSO-d6, TMS):
d 9.91 (s, 2H), 9.08 (s,
1H), 7.92 (d, 1H, J = 7.2 Hz), 7.77 (d, 2H, J = 7.1 Hz), 7.67–7.56 (m,
3H), 7.40 (d, 3H, J = 7.6 Hz), 7.15 (d, 1H, J = 7.3 Hz), 6.64 (s, 2H), 6.47
(s, 4H). 13C NMR (100 MHz, DMSO-d6, TMS):
d 163.6, 158.6, 152.4,
150.3, 148.9, 139.0, 134.5, 134.2, 129.8, 129.3, 129.0, 128.4, 128.0,
124.0, 123.4, 120.6, 112.3, 110.1, 102.5, 65.8. MS (ESI) m/z
537.0463 [MÀH]À, Calcd. C29H19BrN2O4 538.0528.
We commenced our investigation by first verification of the
sensing media. The combination of MeOH/H2O (4:6, v/v) was
proved to be highly efficient (Figs. S1–S3 in Supporting informa-
tion). The selective properties of the probes were then examined
under simulated physiological conditions (PBS buffer, pH 7.4)
based on the investigation of pH responses using the solutions with
pH values from 1.7 to 13.0 (Figs. S4–S6 in Supporting information).
The probes exhibited similar high selectivity toward OClÀ (Fig. 2,
Red bars, Fig.S7–S9 in Supporting information) among the various
Synthesis of the product of probe 1 upon addition of OClÀ: Probe
1 (0.46 g, 1 mmol) was dissolved in 20 mL ethanol. The 1.0 mL of
30% NaOCl solution was then added. The reaction mixture was
refluxed for 1 h. After cooling to the room temperature, the
mixture was filtrated and solvent evaporated under reduced
pressure and the resulting powder was purified by column
chromatography on silica gel (eluent: ethyl acetate only) yielding
an orange powder. 1H NMR (400 MHz, DMSO-d6, TMS):
1H, J = 8.2 Hz), 7.54 (s, 3H), 7.30 (d, 4H, J = 7.4 Hz), 6.81 (d, 2H,
d
7.80 (d,
testing ions including Li+, Na+, K+, Mg2+, Ba2+, Ca2+, Mn2+, Ni2+, Hg2+
,
,
Co2+, Zn2+, Pb2+, Cu2+, Cd2+, Fe3+, Al3+, Cr3+, Sn4+, SO42À, ClÀ, CO3
2À
Scheme 1. Synthesis of the probes.
Please cite this article in press as: Y.-Y. Huang, et al., High efficient probes with Schiff base functional receptors for hypochlorite sensing