A cationic porphyrin-based self-assembled film for mercury ion detection

Article abstract of DOI:10.1016/j.tetlet.2008.01.138

A cationic 5,15-(p-(9,9-bis(6-trimethylammoniumhexyl)fluorenylethynyl)phenyl)porphyrin tetrabromide was synthesized and the self-assembled films were used for Hg²⁺ detection in aqueous media. The detection response is based on fluorescence quenching of the porphyrin molecule upon coordination with Hg²⁺. The detection shows high selectivity for Hg²⁺ over Cu²⁺, Zn²⁺, Pb²⁺, Cd²⁺, Mn²⁺, Ni²⁺, Co²⁺ and Ca²⁺. A linear response toward Hg²⁺ in a concentration range of 1 × 10^-10-1 × 10^-6 M was observed for the film with a detection limit of 0.1 nM. The cationic porphyrin film shows higher stability and significant improvement in detection sensitivity, as compared to other porphyrin-based sensors. The amphiphilic cationic nature of the porphyrin synthesized also allows for the direct deposition of a porphyrin layer on a bare glass surface through self-assembly.

Full text of DOI:10.1016/j.tetlet.2008.01.138

Available online at www.sciencedirect.com
Tetrahedron Letters 49 (2008) 2311–2315
A cationic porphyrin-based self-assembled film
for mercury ion detection
Zhen Fang, Bin Liu *
Department of Chemical and Biomolecular Engineering, 4 Engineering Drive 4, National University of Singapore, Singapore 117576, Singapore
Received 23 November 2007; revised 14 January 2008; accepted 31 January 2008
Available online 7 February 2008
Abstract
A cationic 5,15-(p-(9,9-bis(6-trimethylammoniumhexyl)fluorenylethynyl)phenyl)porphyrin tetrabromide was synthesized and the
self-assembled films were used for Hg2+ detection in aqueous media. The detection response is based on fluorescence quenching of
the porphyrin molecule upon coordination with Hg2+. The detection shows high selectivity for Hg2+ over Cu2+, Zn2+, Pb2+, Cd2+
,
Mn2+, Ni2+, Co2+ and Ca2+. A linear response toward Hg2+ in a concentration range of 1 ꢀ 10ꢁ10–1 ꢀ 10ꢁ6 M was observed for the
film with a detection limit of 0.1 nM. The cationic porphyrin film shows higher stability and significant improvement in detection
sensitivity, as compared to other porphyrin-based sensors. The amphiphilic cationic nature of the porphyrin synthesized also allows
for the direct deposition of a porphyrin layer on a bare glass surface through self-assembly.
Ó 2008 Elsevier Ltd. All rights reserved.
Keywords: Cationic porphyrin; Fluorene; Fluorescence quenching; Mercury; Self-assembly
There has been growing interest in using porphyrins as
active materials for analytical applications owing to their
large absorption coefficient and well tunable fluorescence
emission.1 In addition, porphyrins show high sensitivity
to metal ions, and display photometric changes in the
region of the Soret band (ꢂ400 nm, S0?S2) as well as the
minor Q bands (500–700 nm, S0?S1) upon metal ion
chelation.2 Large Stoke’s shifts (>200 nm) are observed
for most porphyrins because of the fast vibration relaxa-
tion from S2?S1, while the maximum emission is
located in the Q-bands and can be attributed to S1?S0
transitions. Prominent fluorescence quenching occurs once
metalloporphyrins are formed.3 This property has been
widely used for sensing heavy metal ions in aqueous
solutions.4–8
ilized 5,10,15,20-tetraphenylporphyrin (H2tpp) embedded
in a polyvinyl chloride (PVC) membrane was operated
through ion exchange between hydrogen and mercury(II)
in aqueous solutions, which showed a detection limit
(DL) of 4.0 ꢀ 10ꢁ8 M.4 By incorporating alkylated b-
cyclodextrin (b-CD) in the H2tpp solution, significant fluo-
rescence enhancement (up to 45-fold) was observed, which
further improved the detection limit to 2.0 ꢀ 10ꢁ9 M.5
However, both the sol–gel membrane and b-CD enhanced
neutral molecule based sensors suffered from the low stabil-
ity of porphyrin in aqueous media and could only work
within a certain pH range. To improve the stability and
function of porphyrin-based sensors, amphiphilic porphy-
rins, such as 5,10,15,20-tetra(p-sulfonatophenyl)porphyrin
(TPPS), were developed. A sol–gel membrane of TPPS
could afford a detection limit of 7 ꢀ 10ꢁ9 M with improved
stability as compared to neutral porphyrin-based mercury
sensors.6 The TPPS-based sol–gel films, however, are not
suitable for long-term use since the leaching of porphyrin
increased with time. To solve the problem, a N-trimethoxy-
silylpropyl-N0,N00,N000-trimethylammonium chloride (TMAC)
Both neutral and charged porphyrins have been used for
the detection of mercury ions in aqueous media. A lyoph-
*
Corresponding author. Tel.: +65 65168049; fax: +65 67791936.
E-mail address: cheliub@nus.edu.sg (B. Liu).
0040-4039/$ - see front matter Ó 2008 Elsevier Ltd. All rights reserved.
doi:10.1016/j.tetlet.2008.01.138

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Z. Fang, B. Liu / Tetrahedron Letters 49 (2008) 2311–2315
functionalized substrate was developed to allow self-assem-
bly of TPPS films through electrostatic interactions, which
allowed the colorimetric detection of Hg2+ in a concentra-
tion range of 2.5 ꢀ 10ꢁ8–5 ꢀ 10ꢁ7 M.7 This detection strat-
egy suffered from the interference of many other metal
ions, such as Cu2+, Pb2+ and Fe3+. Recently self-assembled
films of 5-(p-trimethylammonium trismethyleneoxy-
phenyl)-10,15,20-triphenylporphyrin chloride on glass were
also reported, which have shown good mechanical stability
with a detection limit of around 1 lM for mercury ions in
aqueous solutions.8
The fluorescence quantum yield of most porphyrins is
less than 20%, which is due to the high yield of non-irradi-
ation decay and intersystem crossing from excited singlet
state (S1) to excited triplet state (T1).9 To improve the
detection sensitivity of porphyrin-based optical sensors, it
is of importance to develop porphyrin derivatives with high
quantum yields. In addition, the degeneration of probe
molecules from the solid substrate is one of the most seri-
ous problems for the long-term operation of porphyrin
based solid state sensors.4,6 It became our aim to develop
high quantum yield porphyrin derivatives that could
anchor the substrate with high stability. In this Letter, we
report the synthesis and characterization of a new cationic
porphyrin, 5,15-(p-(9,9-bis(6-trimethylammoniumhexyl)-
fluorenylethynyl)phenyl) porphyrin tetrabromide (1) and
its application to mercury ion detection as a self-assembled
film on glass.
Dipyrromethane (2) and 2-bromo-9,9-bis(6-bromo-
hexyl) fluorene (3) were synthesized according to previous
reports.10,11 Coupling of 3 and trimethylsilylacetylene
under Sonogashira conditions afforded 2-(9,9-bis(6-bromo-
hexyl)fluoren-7-yl)ethynyltrimethylsilane (4). After the
removal of the trimethylsilyl group in a KOH/THF/
methanol solution, 9,9-bis(6-bromohexyl)-2-ethynyl-9H-
fluorene (5) was coupled with p-bromobenzylaldehyde to
give p-(2-(9,9-bis(6-bromohexyl)-9H-fluoren-7-yl)ethynyl)-
benzaldehyde (6) in a yield of 51%. Porphyrin 7 was pre-
pared from 6 and 2 by a modified MacDonald condensa-
tion.12 Continuous extraction with methanol was carried
out to remove the trace chloranil. The residue was then
recrystallized from chloroform and hexane to afford 7 as
a purple solid. MALDI-TOF gave a clear mass spectrum
of the neutral porphyrin 7, which was consistent with the
expected structure (see Fig. S1 in the Supplementary data).
The 1H NMR spectrum of 7 shows signals at d 10.36, 9.44,
and 9.12 ppm, representing the protons of porphyrin. The
signal of the two delocalized hydrogens was found at
ꢁ3.10 ppm (Scheme 1).
The cationic porphyrin 1 was synthesized through the
amination of the neutral porphyrin 7 by the addition of tri-
methylamine to a solution of 7 in THF. Compound 1 was
i
NH
HN
N
H
37%
2
BrC6H12
C6H12Br
BrC6H12
C6H12Br
Br
ii
70%
iii
Br
Si
56%
3
4
BrC6H12
C6H12Br
BrC6H12
C6H12Br
v
CHO
iv
51%
96%
6
5
70%
NH
N
N
BrC6H12
C6H12Br
vi
BrC6H12
C6H12Br
HN
65%
7
NH
N
N
Br(H3C)3NC6H12
vii
C6H12N(CH3)3Br
Br(H3C)3NC6H12
C6H12N(CH3)3Br
HN
65%
1
Scheme 1. Synthesis of 5,15-(p-(9,9-bis(6-trimethylammoniumhexyl)fluorylethynyl)phenyl) porphyrin tetrabromide (1). Reagents and conditions: (i)
paraformaldehyde, acetic acid, MeOH, 22 h, argon; (ii) KOH, (C4H9)4NBr, 1,6-dibromohexane, 75 °C; (iii) (Ph3P)2PdCl2, CuI, triethylamine,
trimethylsilylacetylene, 8 h, nitrogen, 70 °C; (iv) KOH, THF, MeOH, H2O, 1 h; (v) p-bromobenzylaldehyde, (Ph3P)2PdCl2, CuI, diisopropylamine,
overnight, nitrogen, 70 °C; (vi) dipyrromethane 2, CH2Cl2, trifluoroacetic acid, 15 h, nitrogen; chloranil, 50 °C, 1 h; (vii) trimethylamine, THF, ꢁ78 °C to
room temperature, 12 h; MeOH, 20 h.

Z. Fang, B. Liu / Tetrahedron Letters 49 (2008) 2311–2315
2313
obtained as a purple solid in 65% yield. As compared to the
1H NMR spectrum of 7, the signal due to –CH2Br in the
fluorene chain at d 3.33 ppm disappeared; meanwhile a
new resonance at d 3.07 ppm corresponding to –N(CH3)3
groups appeared for the cationic porphyrin. The chemical
shifts for the porphyrin, phenyl, and fluorene protons did
not change obviously upon amination. However, the signal
at d ꢁ3.10 ppm disappeared for 1, which is possibly a result
of rapid proton exchange between porphyrin and deuter-
ated methanol used for the NMR measurement. The 13C
NMR spectrum of 1 clearly showed seven carbons for the
side chain and one corresponding to the C-9 of fluorene.
Due to the limited solubility of 1 in methanol and DMSO
(<5 mg/mL), only a few carbons with respect to phenyl and
the porphyrin macrocycle were observed in the 13C NMR
spectrum.
Figure 1 shows the absorption and emission spectra of
cationic porphyrin 1 in methanol and in self-assembled film
state (procedure for film preparation is outlined in Supple-
mentary data). The cationic porphyrin exhibits strong
absorption at 408 nm, with four minor Q-bands at 502,
537, 577 and 631 nm, respectively, in methanol. The Soret
band at 408 nm could be attributed to the p–p* transition
from the ground state S0 to the second excited singlet state
S2. The four Q-bands are attributed to p–p* transitions
from the ground state S0 to the different vibration levels
of the first excited singlet state S1.13 Due to rapid vibronic
relaxation from S2 to S1, only emission at 635 nm and
699 nm was observed and there were no emission peaks
near the Soret band. The absorption and emission maxima
of the self-assembled films are red-shifted by ꢂ10 nm com-
pared to that for 1 in methanol, which is due to the inten-
sive intermolecular interaction in the film state.14 The
quantum yields (U) for 1 in methanol and 7 in THF were
measured using quinine sulfate in 0.1 N H2SO4 as the stan-
dard (U = 54%). The neutral porphyrin 7 in THF has a U
value of 15%, which is slightly higher than H2tpp (ꢂ13% in
toluene).15 Cationic 1 has a quantum yield of ꢂ10% in
methanol, which is similar to TPPS (12% in water).16 The
effect of aggregation was investigated by adding water to
a methanol solution of 1. The fluorescence was quenched
upon the addition of water, and the emission maximum
was slightly red-shifted (Fig. S3 in the Supplementary
data). This observation indicates that although the sp3 car-
bon bridge in propeller-like fluorene would impede the reg-
ular packing of molecule 1, the strong p–p interaction of
porphyrin moieties predominates for aggregation, which
leads to fluorescence quenching and emission red shift.
Figure 2 shows the effect of Hg2+ on the fluorescence
quenching of the cationic porphyrin film self-assembled
on a glass slide. With the addition of Hg2+, both emission
bands at 643 nm and 705 nm decreased gradually. The
fluorescence quenching suggests the interaction between
porphyrin and Hg2+. The relationship between the fluores-
cence intensity and the mercury concentration is shown in
Figure 3. The logarithm of the relative fluorescence value
(F ꢁ F0)/(F1 ꢁ F) is given as a linear correlation of the log-
arithm of Hg2+ concentration in the range from 10ꢁ10 to
10ꢁ6 M, which can be explained using Eq. 8 in the Supple-
mentary data. In Figure 3, F0 and F1 are defined as the
fluorescence intensities in the absence and presence of mer-
cury ions, respectively. The linear quenching indicates a 1:1
(10ꢁ10–10ꢁ6 M) complex formation as in Eq. 1 (Supple-
mentary data), which is similar to the previous reports.3–5
The well fitted curve could serve as a calibration standard
to quantify Hg2+ concentration in solution. The detection
limit is estimated to be approximately 10ꢁ10 M, where the
fluorescence quenching is equal to three times the standard
deviation of blank films. As compared to previously
reported porphyrin-based mercury sensors,1–7 the
improved detection sensitivity for 1 indicates that at low
[Hg2+], the effective binding between metal ions and the
porphyrin core is improved. This is due to electron delocali-
zation along the conjugation between the fluorenyl-
ethynylphenyl moieties and the central porphyrin ring,
where the relatively electron rich fluorenylethynylphenyl
groups donate electrons to increase the electron density
of the porphyrin macrocycle.17
1000
1
UV in MeOH
UV in Film
FL in MeOH
FL in Film
1.0
0.8
0.6
0.4
0.2
0.0
800
600
10
400
200
0
650
700
Wavelength/nm
750
350 400 450 500 550 600 650 700 750
Wavelength/nm
Fig. 2. Fluorescence spectra of the cationic porphyrin film after immer-
sion in various Hg2+ solutions (from 1 to 10, [Hg2+] = 0, 0.1, 1, 10, 60,
100 nM, 1.0, 10.0, 30.0, 100 lM).
Fig. 1. UV and fluorescence (FL) spectra of 1 (1 ꢀ 10ꢁ6 M) in MeOH and
in film state.

2314
Z. Fang, B. Liu / Tetrahedron Letters 49 (2008) 2311–2315
soaked and dried for 10 cycles (see Fig. S2 in the Supple-
1.0
0.5
mentary data). Furthermore the freshly prepared porp-
hyrin films are stable when stored under dry and dark
conditions at ambient temperature. The loss of fluorescence
was less than 1.5% after the films had been stored for three
weeks. Both short-term and long-term measurements
showed that these self-assembled films are stable, which
make them suitable for the detection and determination
of Hg2+ in aqueous solution.
In conclusion, we have synthesized a cationic porphyrin
and developed a simple fluorometric method for the detect-
ing and quantifying mercury ions in aqueous solution. The
detection is based on the fluorescence quenching of self-
assembled films on glass slides. The deposited films are
highly stable in operation and storage, and are very sensi-
tive and selective for mercury detection. A detection limit
of ꢂ10ꢁ10 M was achieved, which is among the lowest
for porphyrin-based Hg2+ detection. Under optimized con-
ditions (e.g., temperature, pH, film thickness, etc.), further
improvement is expected to yield more efficient Hg2+
sensors. As compared to the previously reported methods
which utilize commercial porphyrins or their derivatives
with slight modifications, the strategy we have developed
for the synthesis of cationic porphyrins sets an example
for future development of fine-tuned porphyrin-based
materials for different sensor applications.
y=0.3983x+2.5638
R2=0.9932
0.0
-0.5
-1.0
-1.5
-10
-9
-8
-7
-6
-5
log(Hg2+
)
Fig. 3. log [(F ꢁ F0)/(F1 ꢁ F)] versus log (Hg2+).
By adding different amounts of potentially interfering
ions (Cu2+, Zn2+, Pb2+, Cd2+, Mn2+, Ni2+, Co2+ and
Ca2+) to the sample solution containing 0.3 lM Hg2+
(ꢂ50% quenching as compared to blank), the selectivity
of the porphyrin film was investigated and the results are
shown in Table 1. The contaminants do not show signifi-
cant interference in Hg2+ determination even at 1000-fold
excess as compared to that of mercury. This outcome
shows the high selectivity of the self-assembled porphyrin
films toward Hg2+
.
To evaluate the stability of the porphyrin films, blank
films were repeatedly immersed into pure water and dried
under nitrogen before fluorescence measurement. Due to
the negatively charged nature of the glass surface, static
interactions bind the cationic porphyrin to the slide
surface. As compared to 5-(p-trimethylammonium tris-
methyleneoxyphenyl)-10,15,20-triphenylpoyphyrin chloride,
where only one positive charge was attached to the porp-
hyrin ring, the increase in positive charge (four for 1)
greatly reduces the leaching of porphyrin into solution.
For the blank films of 1, there was no obvious change in
film fluorescence intensity even after the films had been
Acknowledgment
The authors are thankful to the National University of
Singapore (NUS ARF R-279-000-197-112/133 and R-
279-000-234-123) for financial support.
Supplementary data
Synthesis and characterization of porphyrins, operation
principles, preparation of metal ions solutions, procedures
to self-assemble the porphyrin films and the fluorescence of
blank films. Supplementary data associated with this article
can be found, in the online version, at doi:10.1016/j.tetlet.
2008.01.138.
Table 1
Effect of different ions on Hg2+ detectiona
Entry
Concentration (M) Relative fluorescence
change value,
b
References and notes
(F2 ꢁ F1)/F1 (%)
Ca2+ (chloride)
Cd2+ (sulfate)
Cu2+(chloride)
Co2+(chloride)
Mn2+(sulfate)
3 ꢀ 10ꢁ4
3 ꢀ 10ꢁ4
3 ꢀ 10ꢁ5
3 ꢀ 10ꢁ4
3 ꢀ 10ꢁ4
3 ꢀ 10ꢁ4
3 ꢀ 10ꢁ4
3 ꢀ 10ꢁ4
3 ꢀ 10ꢁ4 c
ꢁ1.3
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