Highly sensitive salicylic fluorophore for visual detection of picomole amounts of Cu2+

Article abstract of DOI:10.1039/c3ra44239j

Four new fluorophores containing multiple fluorogenic branches of phenylene-ethynylene and salicylic acid Cu²⁺ binding sites are synthesized and evaluated as Cu²⁺ sensors in aqueous media. The fluorophore, F4, having a triphenylamine core and three fluorogenic branches, possesses superior sensitivity, selectivity and emission properties for fluorescence quenching detection of Cu²⁺. This fluorophore exhibits the highest Cu²⁺ quenching sensitivity in aqueous solution; K _sv = 5.79 × 10⁶ M^-1. The relatively high fluorescence emission of this fluorophore provides excellent visual contrast in a solid state paper-based sensor that enables naked eye detection of Cu ²⁺ at the picomole level. The Royal Society of Chemistry.

Full text of DOI:10.1039/c3ra44239j

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PAPER
Highly sensitive salicylic fluorophore for visual
detection of picomole amounts of Cu2+†
Cite this: RSC Adv., 2013, 3, 25215
Nakorn Niamnont,a Nattaporn Kimpitak,b Gamolwan Tumcharern,c
d
d
*
Paitoon Rashatasakhon and Mongkol Sukwattanasinitt
Four new fluorophores containing multiple fluorogenic branches of phenylene-ethynylene and salicylic
acid Cu2+ binding sites are synthesized and evaluated as Cu2+ sensors in aqueous media. The
fluorophore, F4, having a triphenylamine core and three fluorogenic branches, possesses superior
sensitivity, selectivity and emission properties for fluorescence quenching detection of Cu2+. This
Received 7th August 2013
Accepted 16th September 2013
fluorophore exhibits the highest Cu2+ quenching sensitivity in aqueous solution; Ksv ¼ 5.79 ꢀ 106 Mꢁ1
.
DOI: 10.1039/c3ra44239j
The relatively high fluorescence emission of this fluorophore provides excellent visual contrast in a solid
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state paper-based sensor that enables naked eye detection of Cu2+ at the picomole level.
polymers (CPs) have been selected for detection of Cu2+for their
high uorescence quenching sensitivity caused by the ampli-
Introduction
Cu2+ is an important ion which plays a critical role as a catalytic
cofactor for a variety of metalloenzymes, including superoxide
dismutase, cytochrome c oxidase, tyrosinase and nuclease.1
Several research works to establish the detection of Cu2+ ions in
living cells at the micromolar level have been reported.2–6
However, under overexposure conditions, Cu2+can accumulate
within tissues and organs and exhibit toxicity causing oxidative
stress and disorders associated with neurodegenerative
diseases e.g. Alzheimer's, Wilson's and Parkinson's diseases.7–10
A Cu2+ level above 0.6 mmol in urine collected over 24 hours is
strongly indicative of Wilson's disease and other abnormal
conditions.11–13 The World Health Organization (WHO) recom-
mends that the concentration of Cu2+ in drinking water should
not exceed 2 mg Lꢁ1 (31 mM).14 Thus, there is the need for
sensing methods which can detect low doses of Cu2+ in bio-
logically relevant aqueous samples. Among various modes of
sensing, the observation and measurement of uorescence
signal change is one of the most sensitive, rapid, simple and
non-destructive.15–18 In certain systems, a naked eye detection of
Cu2+ is possible even at nanomole levels.19–26 Conjugated
cation effect originating from their efficient intramolecular
energy and electron transfer-processes.27–31
In comparison with a polymer, a small uorescent molecule
has a better dened structure but lacks signal amplication as it
usually contains only one uorogenic unit. To attain the
amplication effect, a well-dened structure and sufficient
water solubility, we have recently focused on the molecular
design of sensing compounds based on star-shaped and
dendritic structures with multiple uorogenic units and ionis-
able peripheral groups.32–35 We have found that the 1,3,5-tri-
phenylbenzene uorophore F1 containing salicylic end groups
was an excellent Cu2+ uorescence quenching sensor with a
quenching efficiency of 1.6 ꢀ 106 Mꢁ1
.
34 Despite high selectivity
and sensitivity, F1 possesses a low quantum efficiency (0.007)
which required the addition of a non-ionic surfactant to
enhance its initial uorescence signal. For naked eye detection
of uorescence quenching, a uorophore with a higher
quantum efficiency is more desirable. We noted that the poor
quantum yield of F1 was partially caused by self-quenching,
originating from its hydrophobic planar structure which tends
to form some sort of H-aggregate in water and the solid state. In
this work, we therefore decided to synthesize four new
analogues (F2–F5) of F1 (Fig. 1) with the aim that these uo-
rophores would have a higher quantum efficiency suitable for
applications for naked eye detection.
aOrganic Synthesis & Natural Products Research unit, Department of Chemistry,
Faculty of Science, King Mongkut's University of Technology Thonburi, Bangkok
10140, Thailand. Fax: +66 24708843
bProgram of Petrochemistry and Polymer Science, Faculty of Science, Chulalongkorn
University, Bangkok 10330, Thailand
cNational Nanotechnology Center, National Science and Technology Development
Agency, Pathumthani 12120, Thailand
dOrganic Synthesis Unit, Department of Chemistry, Faculty of Science and Nanotec-CU
Center of Excellence on Food and Agriculture, Chulalongkorn University, Bangkok
10330, Thailand. E-mail: smongkol@chula.ac.th; Tel: +66 2218 7620
Experimental section
Apparatus, reagents and chemicals
Methyl 4-iodosalicylate, 1,8-diazabicyclo[5.4.0]undec-7-ene
(DBU), copper(I) iodide, iodine monochloride and sodium
chloride were reagent grade and purchased from Sigma-Aldrich.
† Electronic supplementary information (ESI) available: 1H and 13C NMR data and
photophysical properties of uorophores. See DOI: 10.1039/c3ra44239j
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Fluorescence spectra were acquired on a Varian Carry Eclipse
spectrouorometer (Varian, USA) from solution samples in a
1 ꢀ 1 cm2 quartz cuvette cell. The uorophore solutions were
prepared in 10 mM phosphate buffer saline (pH 7.4). The stock
solutions of metal ions were prepared in Milli-Q water.
Synthesis and characterization
Compound 1 in Scheme S1†. A mixture of methyl 4-ethynyl-
2-hydroxybenzoate (0.50 g, 2.84 mmol), PdCl2(PPh3)2 (0.11 g,
0.16 mmol), CuI (0.03 g, 0.16 mmol) in toluene (15 mL), was
added to DBU (1.0 mL) and the mixture was stirred at RT for
24 h. Aer the combined ltrate was evaporated, the residue
was eluted through a silica gel column by gradient solvents
from pure hexane to methylene chloride–hexane (3/1 v/v) as an
eluent. 1 was obtained as a light-yellow solid (0.44 g, 88% yield),
1H NMR (CDCl3, 400 MHz): d (ppm) 10.81 (s, 2H), 7.86 (d, J ¼ 8.0
Hz, 2H), 7.83 (s, 2H), 7.28 (s, 2H), 7.18 (d, J ¼ 12.0 Hz, 2H), 3.98
(s, 6H). 13C NMR (CDCl3, 100 MHz): d (ppm) 170.2, 161.4, 130.1,
129.9, 122.8, 122.7, 120.8, 112.7, 91.3, 52.6; MALDI-TOF m/z
calcd for C20H14O6, 350.079; found, 350.367.
Fig. 1 Structures of molecules F1–F5.
Compound 2 in Scheme S1†. A mixture of 4,7-dibromobenzo-
[c]-1,2,5-thiadiazle (2BrDZ) (133.5 mg, 0.45 mmol), PdCl2(PPh3)2
(28 mg, 0.04 mmol), CuI (8.0 mg, 0.04 mmol) and methyl 4-
ethynyl-2-hydroxybenzoate (179.6 mg, 1.02 mmol) in toluene
(15ꢂmL) was added to DBU (0.5 mL) and the mixture was stirred at
70 C for 12 h. Aer the combined ltrate was evaporated, the
residue was eluted through a silica gel column by gradient
solvents from pure hexane to methylene chloride–hexane (1/1 v/v)
as an eluent to afford 2 which was obtained as a yellow green solid
(139.5 mg, 64% yield). 1H NMR (CDCl3, 400 MHz): d (ppm) 10.81
(s, 2H), 7.86 (d, J ¼ 8.0 Hz, 2H), 7.83 (s, 2H), 7.28 (s, 2H), 7.18 (d,
J ¼ 12.0 Hz, 2H), 3.98 (s, 6H); 13C NMR (CDCl3, 100 MHz): d (ppm)
170.2, 161.4, 154.4, 133.0, 130.1, 130.0, 122.8, 121.0, 117.3, 113.0,
96.6, 88.0, 52.6.
Table 1 Photophysical properties of F2–F5 in PBS (10 mM, pH 7.4)
Absorption
Emission
Compound
labs (nm)
log 3
lem (nm)
Fa (%)
F1b
F2
F3
F4
F5
316
395
322
373
379
4.28
3.61
4.44
4.84
4.80
465
532
434
506
480
0.7
0.2
0.3
3.2
0.4
a
Quinine sulfate in 0.1 M H2SO4 (F ¼ 54%) was used as the standard.
b
The data are taken from ref. 34.
Compound 3 in Scheme S1†. A mixture of 4,40,400-triiodo-
triphenylamine (T3I) (2.00 g, 3.21 mmol), PdCl2(PPh3)2 (0.11 g,
0.16 mmol), CuI (0.03 g, 0.16 mmol), methyl 4-ethynyl-2-
Bis(triphenyl-phosphine)palladium(II) dichloride (PdCl2(PPh3)2), hydroxybenzoate (1.24 g, 7.06 mmol) in toluene (30 mL) was
potassium hydroxide, potassium carbonate, potassium chlo- added to DBU (2.0 mL) and the mixture was stirred at RT for
ride, potassium hydrogen carbonate and dipotassium hydrogen 24 h. Aer the combined ltrate was evaporated and the residue
phosphate were purchased from Fluka (Switzerland). Trime- was eluted through a silica gel column by gradient solvents
thylsilylacetylene was purchased from GFS Chemical. Metal from pure hexane to methylene chloride–hexane (3/1 v/v) as an
ions (Ag+, Al3+, Ba2+, Ca2+, Cd2+, Co2+, Cu2+, Fe2+, Fe3+, Hg2+, K+, eluent. The fractions were combined and the solvents were
Li+, Mg2+, Mn2+, Na+, Ni2+, Pb2+, Sr2+ and Zn2+) for uorescent removed to afford the desired product as a yellow-orange solid
measurements were purchased from Sigma-Aldrich and Fluka (0.32 g, 13% yield). 1H NMR (CDCl3, 400 MHz): d (ppm) 10.78 (s,
(Switzerland). All column chromatography was operated using 3H), 7.80 (d, J ¼ 8.0 Hz, 3H), 7.45 (d, J ¼ 8.0 Hz, 6H), 7.08–7.12
Merck silica gel 60 (70-230 mesh). Thin layer chromatography (m, 9H), 7.00–7.03 (m, 3H), 3.96 (s, 9H). 13C NMR (CDCl3, 100
(TLC) was performed on silica gel plates (Merck F245). Solvents MHz): d (ppm) 170.3, 161.4, 147.2, 133.3, 130.8, 130.0, 124.2,
used for extraction and chromatography were commercial grade 122.5, 120.3, 117.6, 112.1, 92.6, 88.7, 52.5. MALDI-TOF m/z calcd
and distilled before use. Milli-Q water was used in all experi- for C48H33NO9, 767.216; found, 766.807.
ments unless specied otherwise.
Compound 4 in Scheme S1†. A mixture of T3I (2.00 g, 3.21
1
Dendritic uorophore was characterized with H-NMR and mmol), PdCl2(PPh3)2 (0.11 g, 0.16 mmol), CuI (0.03 g, 0.16
13C-NMR spectra from sample solutions in CD3OD by a Varian mmol), methyl 4-ethynyl-2-hydroxybenzoate (1.24 g, 7.06 mmol)
Mercury NMR spectrometer (Varian, USA) at 400 MHz and in toluene (30 mL) was added to DBU (2.0 mL) and the mixture
Bruker 400 MHz, respectively. Absorption spectra were was stirred at RT for 24 h. Aer the combined ltrate was
measured by a Varian Carry 50 UV-Vis spectrophotometer. evaporated and the residue was eluted through a silica gel
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(0.06 g, 0.08 mmol), CuI (0.02 g, 0.08 mmol) and trimethyl-
silylacetylene (0.33 g, 3.3 mmol) in toluene (10 mL) was added to
DBU (1 mL) and the mixture was stirred at room temperature for
3 h. The reaction mixture was then ltered and the solid was
washed with toluene (3 ꢀ 15 mL). The ltrate was evaporated
and the residue was eluted through a silica gel column by
gradient solvents starting from pure hexane to dichloro-
methane–hexane (1/4 v/v) as an eluent to afford 5 as a yellow
1
solid (0.81 g, 42% yield). H NMR (CDCl3, 400 MHz): d (ppm)
10.78 (s, 2H), 7.80 (d, J ¼ 8.0 Hz, 2H), 7.41 (d, J ¼ 8.0, 8.0 Hz,
6H), 7.11 (s, 2H), 7.03 (dd, 8H), 3.96 (s, 6H), 0.25 (s, 9H); 13C
NMR (CDCl3, 100 MHz): d (ppm) 170.2, 161.3, 147.1, 133.1,
130.7, 129.8, 123.9, 122.3, 120.2, 117.2, 111.9, 104.8, 94.2, 92.5,
88.5, 52.4, 0.0.
Compound 6 in Scheme S1†. A mixture of 5 (1.00 g, 1.45
mmol) and K2CO3 (0.059 g, 0.15 mmol) in dichloromethane
(15 mL) and methanol (15 mL) was stirred at room temperature
for 24 h. The organic layer was separated and the aqueous phase
was extracted with dichloromethane (2 ꢀ 50 mL) and was then
dried over anhydrous MgSO4. The solvent was evaporated and
the residue was eluted through a silica gel column by gradient
solvents starting from pure hexane to dichloromethane–hexane
(1/4 v/v) as an eluent to afford H2SA0 (6) as a brown solid (0.74 g,
Fig. 2 Emission spectra of (a) F2, (b) F3, (c) F4 and (d) F5 (1 mM) in the absence
and presence of Cu2+ (10 mM) in PBS pH 7.4.
1
83% yield). H NMR (CDCl3, 400 MHz): d (ppm) 10.79 (s, 2H),
7.80 (d, J ¼ 8.0 Hz, 2H), 7.59 (d, J ¼ 8.0 Hz, 1.5H), 7.43 (d, J ¼ 8.0
Hz, 4.5H), 7.00–7.12 (m, 8.5H), 6.89 (d, J ¼ 8.0 Hz, 1.5H), 3.96 (s,
6H), 3.08 (s, 1H); 13C NMR (CDCl3, 100 MHz): d (ppm) 170.3,
161.4, 147.2, 133.6, 133.3, 130.0, 124.4, 124.1, 122.5, 120.3,
117.5, 112.1, 92.6, 88.7, 83.5, 52.5.
Compound 7 in Scheme S1†. A mixture of 6 (0.50 g, 0.41
mmol), PdCl2(PPh3)2 (0.02 g, 0.02 mmol), CuI (0.004 g, 0.02
mmol) in toluene (15 mL) was added DBU (1.0 mL) and the
mixture was stirred at RT for 24 h. Aer the combined ltrate was
evaporated, the residue was eluted through a silica gel column by
gradient solvents from pure hexane to methylene chloride–
hexane (3/1 v/v) as an eluent. 7 was obtained as a dark yellow solid
(0.33 g, 65% yield). 1H NMR (CDCl3, 400 MHz): d (ppm) 10.79 (s,
4H), 7.80 (d, J ¼ 8.0 Hz, 4H), 7.45 (d, J ¼ 8.0 Hz, 12H), 7.01–7.12
(m, 20H), 3.96 (s, 12H). 13C NMR (CDCl3, 100 MHz): d (ppm)
170.3, 161.4, 147.0, 133.3, 130.8, 130.0, 124.4, 122.5, 120.4, 117.8,
116.6, 112.1, 92.5, 88.8, 81.9, 74.3, 52.5. MALDI-TOF m/z calcd for
Fig. 3 Fluorescence quenching of (a) F4 and (b) F5 (5 mM) by various metal ions
(5 mM) in PBS (10 mM, pH 7.4). The corresponding fluorescence spectra are shown
in the insets.
C
80H52N2O12, 1232.352; found, 1232.357.
F2. A mixture of 2 (0.50 g, 1.03 mmol) in THF (15 mL) and
methanol (15 mL) was added to saturated KOH aqueous solu-
ꢂ
tion (0.5 mL) and the mixture was heated to 70 C. Aer 24 h,
column by gradient solvents from pure hexane to methylene the solution was evaporated and the residue was dissolved in
chloride–hexane (2/1 v/v) as an eluent. The fractions were water (20 mL). Approximately 50 g of ice was then added to the
combined and the solvents were removed to afford the desired aqueous solution. The mixture was acidied and stored in the
product as a dark yellow solid (0.76 g, 33% yield), 1H NMR refrigerator for 1 h. The product was ltered to afford F2 as a
(CDCl3, 400 MHz): d (ppm) 10.78 (s, 2H), 7.80 (d, J ¼ 8.0 Hz, 2H), green solid (0.34 g, 72% yield). 1H NMR (DMSO-d6, 400 MHz): d
7.59 (d, J ¼ 8.0 Hz, 2H), 7.43 (d, J ¼ 8.0 Hz, 4H), 7.11 (d, J ¼ 8.0 (ppm) 8.00 (s, 2H), 7.85 (d, J ¼ 8.0 Hz, 2H), 7.16–7.18 (m, 4H);
Hz, 2H), 7.00–7.06 (m, 6H), 6.89 (d, J ¼ 8.0 Hz, 2H), 3.96 (s, 6H). 13C NMR (CDCl3, 100 MHz): d (ppm) 171.1, 160.7, 153.5, 133.3,
13C NMR (CDCl3, 100 MHz): d (ppm) 170.2, 161.3, 147.1, 146.4, 130.9, 128.1, 122.3, 119.6, 116.0, 114.1, 95.7, 87.8.
138.6, 133.1, 130.7, 129.8, 127.0, 123.6, 122.3, 120.2, 117.2,
111.9, 92.5, 88.5, 87.5, 52.4.
F3. A mixture of dimethyl 1 (0.50 g, 1.43 mmol) in THF
(15 mL) and methanol (15 mL) was added to saturated KOH
Compound 5 in Scheme S1†. A mixture of methyl 2-hydroxy- aqueous solution (0.5 mL) and the mixture was heated to 70 ꢂC.
5-iodobenzoate (I2SA0) (4) (2.00 g, 2.78 mmol), PdCl2(PPh3)2 Aer 24 h, the solution was evaporated and the residue was
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F4. A mixture of 3 (0.50 g, 0.41 mmol) in THF (15 mL) and
methanol (15 mL) was added to saturated KOH aqueous solu-
ꢂ
tion (0.5 mL) and the mixture was heated to 70 C. Aer 24 h,
the solution was evaporated and the residue was dissolved in
water (20 mL). Approximately 50 g of ice was then added and the
aqueous solution was acidied and stored in the refrigerator for
1 h. The product was ltered to afford F4 as a yellow solid (0.43
g, 92% yield). Mp: >200 ꢂC to decompose. 1H NMR (CD3OD, 400
MHz): d (ppm) 7.72 (d, J ¼ 8.0 Hz, 3H), 7.35 (d, J ¼ 8.0 Hz, 6H),
6.84–6.96 (m, 12H). 13C NMR (CD3OD, 100 MHz): d (ppm)
173.02, 162.82, 148.38, 134.23, 131.62, 131.55, 125.28, 123.14,
120.58, 118.87, 113.77, 92.94, 89.45. MALDI-TOF m/z calcd for
C
45H27NO9, 725.169; found, 725.210.
F5. A mixture of 7 (0.50 g, 0.41 mmol) in THF (15 mL) and
methanol (15 mL) was added to saturated KOH aqueous solu-
ꢂ
tion (0.5 mL) and the mixture was heated to 70 C. Aer 24 h,
the solution was evaporated and the residue was dissolved in
water (20 mL). Approximately 50 g of ice was then added to the
aqueous solution, acidied and kept in a refrigerator for 1 h.
The product was ltered to afford F5 as a yellow solid (0.39 g,
82% yield). Mp: >200 ꢂC to decompose. 1H NMR (CD3OD,
400 MHz): d (ppm) 7.91 (d, 4H), 7.57 (m, 12H), 7.14 (m, 20H);
13C NMR (CD3OD, 100 MHz): d (ppm) 173.0, 162.7, 148.0, 143.5,
135.2, 134.4, 131.6, 125.4, 123.4, 120.7, 119.0, 113.5, 93.3, 89.9;
MS-ESꢁ m/z calcd for C76H44N2O12, 1176.2894, 293.07 [M]4ꢁ
Fig. 4 Bar chart displays fluorescence intensity ratio ((I0 ꢁ I)/(I0 ꢁ IM)) of (a) F4
and (b) F5 in the presence of Cu2+ (5 mM) and another interfering ion (50 mM).
found: 292.83 [M]4ꢁ
.
Paper-based sensor
F4 solution (1.0 mM) in ethanol was pipetted at 1.0 mL onto the
surface of marked lter paper and allowed to air dry to generate
uorescent spots under an ordinary 20 W black light lamp.
Filtered water samples, i.e. drinking, mineral, Milli-Q and river
(Chaophraya), were pipetted on top of the uorescent spots.
Aer air drying, the images of the lter paper under the black
light were photographically recorded.
Fig. 5 Stern–Volmer plots for fluorescence quenching of F4 and F5 (1 mM) by
Cu2+ in PBS (10 mM, pH 7.4).
Results and discussion
The synthesis of uorophores F2–F5 was accomplished by
Sonogashira coupling36,37 and acetylenic homocoupling38,39 fol-
lowed by base-catalyzed hydrolysis of the salicylate esters
(Scheme S1†). The photophysical properties of F2–F5 (spectra
provided in Fig. S1†) in phosphate buffer saline (PBS), pH 7.4
are compiled in Table 1 along with those of F1. The uorophores
showed the maximum absorption wavelength (lab) in the range
of 316–395 nm. The order of lab (F2 > F5 > F4 > F3 > F1) denotes
dissolved in water (20 mL). Approximately 50 g of ice was then
added to the aqueous solution. The mixture was acidied and
stored in the refrigerator for 1 h. The product was ltered to
1
afford 2SAꢁ as a light-yellow solid (0.42 g, 91% yield). H NMR
(CD3OD, 400 MHz): d (ppm) 7.82 (d, J ¼ 8.0 Hz, 2H), 7.00–7.03
(m, 8H), 7.64; 13C NMR (CDCl3, 100 MHz): d (ppm) 131.9, 131.8,
130.4, 123.7, 123.2, 120.9, 91.5.
Table 2 Comparison of Cu2+ detection limit of F4 with other optical sensing materials
Detection
Sensing materials
Solvent
Signal detected
limit
Ref.
Iridium(III) complex (ZIr2)
Imidazole-CP microspheres
AuNPs
F4
PIPES buffer, pH 7.0
Acetate–acetic buffer, pH 6.5
Tris–HCl, pH 7.4
PBS buffer, pH 7.4
Paper-based
Turn-on emission
Turn-off emission
Absorption
Turn-off emission
35 ppb
1 ppb
0.6 ppb
0.5 ppb
10 pmol
15
55
56
This work
Turn-off emission (visual observation)
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sensitivity, selectivity and its initial uorescence quantum
yield. In PBS pH 7.4, F4 and F5 gave the detection limits (at 3ꢀ
noise) of 0.5 ppb and 6.9 ppb, respectively, well below the
WHO limit of 2 mg Lꢁ1 (2 ppm) recommended for drinking
water.14
Paper-based solid state sensors are very economical, envi-
ronmentally friendly and convenient to use.49–53 It is important
to emphasize here that, compared with our previous uo-
rophore F1, F4 not only possesses a 5 times higher quantum
yield but also has a Ksv value 3 times higher.34 This new uo-
rophore is thus tested for feasibility as a Cu2+ sensor detectable
by the naked eye. A series of 1.0 mL F4 solutions in ethanol was
dropped onto a piece of lter paper and allowed to air dry to
Fig. 6 Photographic images of F4 (0.1 nmol) paper-based detection of Cu2+
under a 20 W black light.
the HOMO–LUMO gap associated with the delocalization of the generate rows of bright blue uorescent spots containing 1.0
p-conjugated system. The observed molar extinction coeffi- nmol of F4. In the sensing test, 1.0 mL of each water sample,
cients of all uorophores were well above 1 ꢀ 104 except for that i.e. drinking, mineral, Milli-Q and river water containing
of F2 which was probably underestimated due to its poor solu- various concentrations of Cu2+ (0–500 mM) was dropped on top
bility in water at this physiological pH .
of the uorescent dots. Under an ordinary black light, a dark
All uorophores showed maximum emission wavelengths quenching area within the blue emission spots of F4 was
(lem) in the range of 434–532 nm with the uorescence readily observed for the water samples containing at least 40
quantum yield of 0.2–3.2%. Among the four new uorophores, pmol of Cu2+, representing the naked eye detection limit of
only F4 showed higher quantum yield than F1 probably due to this method.
the decrease of the self-quenching process, originating from its
propeller-like conformation which precludes the intermolecular detection in aqueous solution, the Cu2+ detection limit of F4 is
H-type p–p interaction.40–44 one of the lowest as shown in Table 2.15,19,54 To the best of our
In comparison with other sensing materials used for Cu2+
To evaluate the Cu2+ sensing ability of these new salicylic knowledge, the paper-based sensor constructed from micro-
uorophores, the uorescence signals of F2–F5 solution in PBS liter-drops presented in this work is also one of the most
pH 7.4 were recorded in the absence and presence of Cu2+ sensitive and convenient techniques for the naked eye detection
(10 mM). As illustrated in Fig. 2, only the uorescence signals of of Cu2+ (Fig. 6).
F4 and F5 are quenched by Cu2+, while those of F2 and F3 do not
show any response. The results indicated that not only the
salicylic acid binding sites but also the p-conjugated systems
Conclusion
are important for Cu2+ sensing ability. Since the uorescence Our comparison study on uorophores F1–F5 containing
quenching mechanism of Cu2+ is mostly accepted as a photo- phenylene-ethynylene uorescent units with salicylic acid
induced electron transfer (PET) process,45–48 the quenching termini identied F4 as the most sensitive and selective sensor
ability should thus depend on the energy of the LUMO level of for Cu2+ detection in aqueous media. Not only its multiple
the electron donating uorophore matching that of the Cu2+ salicylic binding units but also its optimal LUMO level are
complex, the electron accepting unit. In other words, the LUMO important for efficient uorescence quenching by Cu2+. F4 also
level of F2 is probably too low while that of F3 is too high to possesses high absorptivity and emission efficiency for Cu2+
allow the transfer of an excited electron from the uorophores which provides excellent visual contrast for naked eye detection
to the Cu2+–salicylate complex.
of Cu2+ at the picomole level on a solid state paper-based
The uorescence quenching selectivity of F4 and F5 by metal sensor.
ions (5 mM) such as Ag+, Al3+, Ba2+, Ca2+, Cd2+, Co2+, Fe2+, Fe3+,
Hg2+, K+, Li+, Mg2+, Mn2+, Na+, Ni2+, Pb2+, Sr2+ and Zn2+ was
Acknowledgements
evaluated. We found that the uorescence signal of F4 was most
effectively quenched by Cu2+ (over 20 times) and somewhat The authors gratefully thank the Thailand Nanotechnology
quenched by Fe2+ (ꢃ2 times) (Fig. 3a). On the other hand, F5 Center (NANOTEC, NSTDA), the IIAC Chulalongkorn University
showed lower quenching sensitivity and selectivity for Cu2+ over Centenary Academic Development Project, the National
Fe2+ (Fig. 3b). The interference test with the interfering metal Research University Project of Thailand, Office of the Higher
ions at 10 times concentration showed negligible interference Education Commission (AM1006A), the Thai Government
of all ions tested on F4 (Fig. 4a) but discernible interference of Stimulus Package 2 (TKK2555, SP2), the Project for Establish-
Fe2+ tested on F5 (Fig. 4b).
The Stern–Volmer plots of the uorescence quenching Products and Agriculture supported by the Thai government
ment of Comprehensive Center for Innovative Food, Health
against the Cu2+ concentration gave linear lines with the slopes stimulus package
2
(TKK2555, SP2), the Ratch-
corresponding to the quenching efficiency (Ksv) of 5.79 ꢀ 106 adaphiseksomphot Endowment Fund (90th Anniversary CU
Mꢁ1 and 1.31 ꢀ 106 Mꢁ1 for F4 and F5, respectively (Fig. 5). Fund), the Thailand Research Fund (TRF-MRG5680031), and
Clearly, F4 is a better Cu2+ sensor than F5 in terms of KMUTT Research Fund for nancial support.
This journal is ª The Royal Society of Chemistry 2013
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25220 | RSC Adv., 2013, 3, 25215–25220
This journal is ª The Royal Society of Chemistry 2013