Highly effective discrimination of catecholamine derivatives via FRET-on/off processes induced by the intermolecular assembly with two fluorescence sensors

Article abstract of DOI:10.1039/c3cc45077e

We proudly presented the novel sensor system based on intermolecular assembled complexes of two fluorescence sensors and a suitable catecholamine acting as a guest linker by the reaction between boronic acid and aldehyde moieties of the sensors with the diol and amine units of catecholamines, respectively. The FRET-on/off induced by the assembly could discriminate structural similarity of catecholamines.

Full text of DOI:10.1039/c3cc45077e

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Highly effective discrimination of catecholamine
derivatives via FRET-on/off processes induced by the
intermolecular assembly with two fluorescence
sensors†
Cite this: Chem. Commun., 2013,
4
9, 9287
Received 8th July 2013,
Accepted 10th August 2013
DOI: 10.1039/c3cc45077e
a
b
a
Anusak Chaicham, Somboon Sahasithiwat, Thawatchai Tuntulani and
a
Boosayarat Tomapatanaget*
www.rsc.org/chemcomm
We proudly presented the novel sensor system based on intermolecular to develop the fluorescence sensor for the highly selective detection
assembled complexes of two fluorescence sensors and a suitable and discrimination among DA, EPI and NE under the significant
catecholamine acting as a guest linker by the reaction between boronic emission changes via the FRET process induced by the inter-
acid and aldehyde moieties of the sensors with the diol and amine molecular assembled complex. This conceptual system is expected
units of catecholamines, respectively. The FRET-on/off induced by the to offer the promising advantages not only reducing the complicated
assembly could discriminate structural similarity of catecholamines.
design and synthesis of the fluorescence compound containing both
donor and acceptor fluorophores but also reducing the waste from
Catecholamines including epinephrine (EPI), norepinephrine synthesis and saving cost. Herein, we demonstrated a FRET-on
NE) and dopamine (DA) as the neurotransmitters, which have process induced by an intermolecular assembly of coumarin alde-
structural similarity, consisting of catechol and amino groups hyde (CA) and pyrene boronic acid (PBA) with catecholamine as a
(
1
,2
involved in a variety of central nervous system functions.
Generally, the amount of DA in body fluids is an essential
guest linker, as shown in Fig 1.
There are a number of fluorescence sensors for catecholamine
3
indicator of patients suffering from Parkinson’s disease in the detection previously reported providing the fluorescence quenching
clinical diagnosis. However, a critically elevated amount of EPI in the upon binding with catecholamine. In the challenge searching for
human blood certainly leads to the increase in blood pressure and the suitable donor fluorophore for catecholamine detection, we
4
heart failure. Furthermore, the patients suffering from tumors of found that pyrene boronic acid (PBA), upon binding to catechol
adrenal glands are found to have large amounts of EPI and NE in based catecholamine, exhibited the fluorescence enhancement.
5
their blood and urine. Specific detection of each catecholamine Undoubtedly, pyrene boronic acid is an appropriate candidate
species is a challenging task for sensing purpose. Regarding the of the donor fluorophore for the FRET process. Moreover, sensor
optical detection, there are only a few reports concerning the PBA is a commercially available chemical substance. Sensor CA,
6
chemosensor of catecholamine, particularly, bearing boronic acid an acceptor fluorescence sensor, could form an iminium ion
and aldehyde for covalently binding with catechol and amine groups
of catecholamine species to form a boronate ester and an iminium
7
ion, respectively. However, most fluorescent chemosensors reported
previously displayed the chelation-enhanced fluorescent quenching
8
(
CHEQ) effect upon adding catecholamines. Concerning the
discrimination of each catecholamine, many reports regarding
fluorescent chemosensors showed the non-specific binding
9
among EPI, DA and NE. None of the fluorescence chemosensors
for catecholamine sensing employed the Fluorescence Resonance
Energy Transfer (FRET) process as a tool in detection. This alerts us
a
Department of Chemistry, Faculty of Science, Chulalongkorn University, Bangkok,
Thailand. E-mail: tboosayarat@gmail.com; Fax: +66-2-254-1309;
Tel: +66-2-218-7642
b
National Metal and Materials Technology Center, National Science and
Technology Development Agency, Pathumthani 12120, Thailand
†
Electronic supplementary information (ESI) available: Fluorescence spectra,
Fig. 1 Conceptual design of an intermolecular assembled complex for catechol-
mass spectroscopy and binding study method. See DOI: 10.1039/c3cc45077e
amine sensing by a FRET-on process.
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with primary amines. On the other hand, sensor PBA could react
Extensive studies on discrimination of catecholamine derivatives
with the catechol group producing the boronate ester whose give rise to the most fascinating sensing approach by the induced-fit-
emission band overlaps with the absorption band of sensor CA. binding of primary amine based catecholamines and aldehyde
12
Prior to verification of the covalent binding affinity of both based acceptor fluorescence sensor producing an iminium ion
fluorescence sensors and guests in this aspect, the pK values of sensor and a consequent FRET-on process. As anticipated, the suitable
a
PBA should be firstly examined by fluorescence spectroscopy. The pH fluorescence sensor containing aldehyde based coumarin (CA) was
a
profile (Fig. S1, ESI†) indicated that the pK values of sensor PBA are selected to be the acceptor part properly coupled with the donor
approximately 6.5. These small pK values implied that the electron fluorescence sensor PBA resulting in the FRET-on process (Fig S8 in
a
rich pyrene ring would stabilize the electron deficient boronic acid to the ESI†). The FRET efficiency for the couple of sensors PBA and CA
3
preferentially rearrange in the sp -hybidized orbital around boron could be optimized by varying the concentration of sensor CA in the
1
0,11
atoms.
Therefore, the usually used pH of 7.4 in normal sensing solution mixture of sensor PBA and DA as summarized in Table S1
3
purposes could benefit the reaction between the sp -hybridized boronic and Fig S9 in the ESI.† The highest FRET efficiency was obtained at
acid based sensor PBA and diol based catecholamine.
The sensor PBA (1 mM) exhibited a strong emission band in the intensity of the sensor. Furthermore, time-resolved FRET (trFRET)
region of 375–425 nm corresponding to monomeric pyrene species. spectra (Fig. S10 in the ESI†) showed the decrease in fluorescence
Actually, the catecholamine contains the air-sensitive catechol group intensity of the PBA excimer band at 451 nm and a concomitant
which is easily oxidized by oxygen. To suppress the oxidization of increase in fluorescence intensity of the CA band at 487 nm, and
À5
3 Â 10 M of CA as evidenced from the largest fluorescence
2 2 3
catechol based catecholamine, Na S O , a reducing agent, was added thus strongly supported that the FRET mechanism occurred in
into the solution system for all manipulation. Upon the addition of our detection system.
DA, NE and EPI, a significant red shift assigned to the excimer band
centered at 451 nm was observed (Fig. 2).
The addition of sensor CA to sensor PBA led to a decrease in
the emission band of monomeric pyrene and an increase in the
We hypothesized that the reaction of catechol based catechol- emission band at 487 nm of sensor CA. Upon adding Lys, Tym,
amine and pyrene boronic acid induced the excimer fluorescence or Glu, the fluorescence responses observed were similar to
spectrum. As a single fluorescence sensor, PBA is a promisingly those of the mixture solution of sensors PBA and CA (in Fig 3).
selective fluorescence probe for catecholamines over other biogenic
After adding DA or NE to solution of sensors PBA and CA, a large
amines. To investigate the binding affinity between sensor PBA and quenching in the monomeric band of sensor PBA and a large
catecholamines, the stoichiometry of sensor PBA and catechol- enhancement of the emission band at 487 nm of sensor CA were
amines including DA, NE and EPI was evaluated by Job’s continuous obviously observed (Fig. 3). Upon the addition of EPI, the excimer
variation method showing the binding mode at a 1 : 1 ratio of sensor band at 451 nm was still observed and also the disappearance of the
PBA and guests as illustrated in Fig. S2–S4 (in the ESI†). The logK emission band at 487 nm for sensor CA supported the non-FRET
values of sensor PBA and DA, NE and EPI examined by fluorometric process. It is indicative of non-binding of EPI and sensor CA. As a
À1
titration are 4.23, 4.10 and 4.35 M , respectively. The log K values of result of the different fluorescence spectra, this system could identify
the binding between sensor PBA and various catecholamines show a EPI selectively from other catecholamine derivatives. Successful
small difference. Considering the fluorescence titration of sensor discrimination of EPI from DA and NE depends strongly on the
PBA and catecholamines, the emission band of sensor PBA and DA reaction between their amine units on the side chain to the aldehyde
assigned to excimer fluorescence at 451 nm was gradually developed moiety of sensor CA to form iminium species. Only primary amines
(Fig. S5 in the ESI†). Similarly, the feature of fluorescence spectra in of DA and NE can react with the aldehyde moiety of sensor CA, while
the case of NE and EPI is identical to those in the case of DA possibly the secondary amine of EPI cannot. Importantly, DA and NE suitably
caused by the same reaction of catechol group and boronic acid act as guest linkers of sensors PBA and CA inducing the FRET-on
based sensors (Fig. S6 and S7 in the ESI†).
process. To confirm our hypothesis of the self-assembled reaction of
Fig. 2 Fluorescence spectra of sensor PBA (1 mM) in buffer solution (100 mM
Fig. 3 Fluorescence spectra of the mixed sensors of PBA (1 mM) and CA (30 mM)
Na
2
S
2
O
3
, 50 mM HEPES, 20 mM NaCl, pH 7.4) with various biogenic amines and
2 2 3
in buffer solution (100 mM Na S O , 50 mM HEPES, 20 mM NaCl, pH 7.4) with
inset: the relative fluorescence intensity of sensor PBA (lex = 340 nm) in the
various biogenic amines and inset: relative fluorescence intensity of the mixed
presence of various biogenic amines.
sensors of PBA and CA (lex = 340 nm) in the presence of various biogenic amines.
9
288 Chem. Commun., 2013, 49, 9287--9289
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expected to further give a high sensing performance for DA in a
strip test. All sensing properties of this adaptive self-assembled
complex with NE are analogous to those with DA. Furthermore,
this proposed self-assembled complex method was applied to
determine the amount of DA in two real urine samples, and the
results are displayed in Table S2 in the ESI.† Average %
Fig. 4 The proposed structure of the PBA–DA–CA complex inducing the FRET recoveries of the spike samples of DA to the solution of sensors
process.
PBA and CA were 106. Our method thus gave good analytical
characteristics of the sensor in dopamine detection.
In summary, we have successfully developed a new sensing
approach based on the adaptive assembled complexes of two
fluorescence sensors inducing the FRET process to discriminate
effectively among catecholamines. The case of a single sensing
element of sensor PBA offered the promising selectivity of catechol-
amines over other biogenic amines by the especially covalent binding
between boronic acid and catechol groups of catecholamine. For two
sensing elements, the additional sensor CA could separate EPI from
other catecholamines. Based on a basic reaction of sensor PBA and
the catechol group as well as sensor CA and primary amines of
Fig. 5 Photographs of (a) the solution of sensor PBA, sensor CA and combi-
catecholamine, this is the first report to explore the intermolecular
nation of sensors PBA and CA in the absence and in the presence of dopamine
and (b) the fluorescence images of sensor PBA; P, sensor CA; C, the mixture of assembled complexes by properly coupled fluorescence chemosensors
sensors PBA and CA in the absence of DA; PC and in the presence of DA; PCD.
to detect catecholamine via the FRET-on process. On the basis of
these findings, the possibility of the intermolecular assembled sensors
based on non-complicated sensory molecules by using a specific guest
both sensors and guest, the ESI-Ms spectrum in Fig S11 in the ESI† as a linker highlighted a powerful tool for sensing purposes in real-
showed the intense peak at m/z of 609.67 which clearly supported the time analysis in the field of medical diagnosis.
proposed structure of the PBA–DA–CA complex as presented in Fig. 4.
This work was supported by the Thailand Research Fund (TRF),
The detection limit of the combination probes of sensors CA the Commission on Higher Education (CHE) (RSA5680015 and
and PBA toward DA and EPI examined by monitoring the RTA5380003) and the Higher Education Research Promotion
emission band at 487 and 451 nm in HEPES pH 7.4 solution and National Research University Project of Thailand, Office of
provided 1.41 and 0.69 mM, respectively, in the linear relation- the Higher Education Commission (AM1006A-56), and the
ship in the concentration range from 16.7 to 47.4 mM for DA Ratchadaphiseksomphot Endowment Fund of Chulalongkorn Uni-
and 2.0–56.0 mM for EPI (Fig. S12 and S13 in the ESI†).
versity (RES560530126-AM). A.C. is a PhD student under the Royal
Since the PBA–CA system can discriminate EPI from DA and Golden Jubilee Program (PHD/0235/2552) of the TRF and CHE.
NE by the non-FRET approach, our PBA–CA system would offer
an excellent way for the detection of DA in the FRET-on process. Notes and references
Moreover, the clinical diagnosis usually monitors the amount
1
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2
3
4
Applications of the sensor system PBA–CA as naked-eye sensors
in aqueous solution and solid support are demonstrated in Fig 5.
Upon exposure to the 356 nm UV–visible light, the solution of the
sensor system in the presence of DA exhibited the color brightness
changing from sky blue to green luminescence corresponding to
the emitting light from sensor CA. In comparison, the solution of
sensor PBA in the presence of DA displayed the blue brightness
corresponding to the excimer band of sensor PBA. In the case of
sensor CA upon binding with DA, the luminescence remained
unchanged even though sensor CA is able to form an iminium ion
with primary amine based DA.
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On the solid support, the filter papers were painted with
PBA–CA ink in the word ‘‘P’’, ‘‘C’’, ‘‘PC’’ and ‘‘PCD’’ by using a
1
paintbrush. Then, all prepared filter papers were immersed in 11 J. P. Lorand and J. O. Edwards, J. Org. Chem., 1959, 24, 769.
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the solution of DA. Interestingly, the green luminescence of the
filter paper labeling PCD containing sensors PBA–CA and DA
was obviously observed. In this regard, the PBA–CA system was
(
3
This journal is c The Royal Society of Chemistry 2013
Chem. Commun., 2013, 49, 9287--9289 9289