Molecular recognition directed porphyrin chemosensor for selective detection of nicotine and cotinine

Article abstract of DOI:10.1039/b006055k

The first example of a metalloporphyrin based fluorescent chemosensor for selective detection of dinitrogen alkaloids such as nicotine and cotinine in solution by using a 'two-point' binding strategy and a modified fluorescence analysis procedure is reported.

Full text of DOI:10.1039/b006055k

Molecular recognition directed porphyrin chemosensor for selective detection
of nicotine and cotinine
Gollapalli R. Deviprasad and Francis D’Souza*
Department of Chemistry, Wichita State University, Wichita, KS 67260-0051, USA.
E-mail: dsouza@wsuhub.uc.twsu.edu
Received (in Corvallis, OR, USA) 25th July 2000, Accepted 14th August 2000
First published as an Advance Article on the web 18th September 2000
The first example of a metalloporphyrin based fluorescent
chemosensor for selective detection of dinitrogen alkaloids
such as nicotine and cotinine in solution by using a ‘two-
point’ binding strategy and a modified fluorescence analysis
procedure is reported.
Nicotine, a dinitrogen alkaloid is a major drug of abuse, acting
as a potent agonist of the nicotinic acetylcholine receptor
(nAChR). Recently, nicotinic agonists and antagonists have
been considered as promising therapeutic agents for a variety of
conditions including the treatment of pain, cognitive and
attention deficits, Parkinson’s disease, Tourette’s syndrome,
and anxiety.1 Nicotine is also the most abundant and potent
pharmacological agent in tobacco and tobacco smoke, hence, it
is of considerable interest to medicine and society.2 Several
methods using HPLC, GC-MS or capillary electrophoresis are
available for quantification of nicotine in tobacco, tobacco
smoke, pharmaceutical agents, atmospheric air (an issue related
to passive smoking), and in urine samples (involves cotinine, a
metabolic product of nicotine).3 Though these methods offer
good detection sensitivity, they often suffer from low selectiv-
ity. Hence, a selective method for sensitive detection of nicotine
and cotinine is highly desirable.
a deshielding effect (up to 0.5 ppm) upon binding to 1. Control
experiments performed using meso-tetraphenylporphyrinato-
zinc, (TPP)Zn, (a compound without the side pendant arm) or
compound 5, bearing a methyl ester group at the substituted
phenyl ring, reveal similar large deshielding of pyridyl protons
of 1 indicating that the pyridyl group of 1 binds to the central
zinc.9
The UV-visible absorption spectral studies reveal red shifted
Soret and visible bands upon addition of 1 to a solution of the
investigated receptor porphyrins confirming that the pyridyl
entity of 1 binds to the central zinc.10 The formation constant K
The development of sensors/biosensors for selective detec-
tion of analytes and improved transduction methods for higher
detection sensitivity is considered to be a challenge in modern
chemistry.4 Among the different techniques utilized for devel-
oping chemical sensors, molecular fluorescence is an important
one because of its high sensitivity of detection down to a single
molecule, recognition and/or self-assembly directed selectivity,
on–off switchability, sub-nanometer spatial resolution with sub-
micron visualization, and sub-millisecond temporal resolution.5
Among the different fluorophores that could be utilized to
develop fluorescent sensors/chemosensors, porphyrins are at-
tractive candidates because of their relatively high fluorescence
quantum yields and the many different established synthetic
procedures of functionalization.6 In porphyrin based chemo-
sensors, sensing is often achieved either by having suitable
receptors at the ring periphery, by metal axial ligation, or a
combination of both.6 In the present study, we have successfully
designed a porphyrin based fluorescence chemosensor for
selective detection of nicotine and cotinine in solution. Our
approach utilizes: (i) a ‘two-point’ binding mechanism which
involves axial ligation of the nicotine pyridine entity through
the zinc ion of the porphyrin cavity, and, hydrogen bonding of
the pyrrolidine ring nitrogen to a carboxylic acid or amide group
located on one of the phenyl rings of a tetraphenylporphyr-
inatozinc macrocycle and (ii) a modified porphyrin fluores-
cence data analysis procedure.
The binding of the dinitrogen alkaloid, 1 or 2, to the newly
1
synthesized, receptor porphyrin,† 3 or 4 was studied by H
NMR and UV-visible absorption spectral methods. Fig. 1
1
depicts H NMR spectra of 1, 3, and, a mixture of 1 and 3 in
CDCl3. Upon binding to 3, the o-pyridyl protons of 1 experience
a shielding up to 5 ppm while the m- and p-pyridyl protons and
the pyrrolidine ring protons of 1 experience less shielding (1–2
ppm). The protons of the substituted phenyl ring of 3 experience
Fig. 1 1H NMR spectrum of (a) nicotine, 1 (31 mM), (b) 1 (31 mM) +
porphyrin, 3 (27 mM), and (c), 3 (12 mM) in CDCl3 at 298 K.
DOI: 10.1039/b006055k
Chem. Commun., 2000, 1915–1916
This journal is © The Royal Society of Chemistry 2000
1915

Table 1 Formation constant, K, and the thermodynamic parameters for zinc
porphyrin–alkaloid complexes in toluene
The inset in Fig. 2 shows such plots obtained for nicotine and
cotinine binding. A linear relationship is obtained for the
emission peak intensity ratio against the alkaloid concentration.
This procedure is found to work well in non-coordinating
solvents such as toluene, o-dichlorobenzene or acetonitrile.
These plots offer the much-needed selectivity with respect to
the presence of other axially coordinating nitrogenous bases. As
shown in the inset of Fig. 2, the intensity ratio of the emission
bands for a strongly coordinating ligand such as pyridine does
not change significantly in the employed concentration range.
These results along with the higher binding constants suggest
that the present ‘two-point’ binding and fluorescence analysis
procedure offers the much-needed selectivity for dinitrogen
alkaloid detection. Further studies to expand this novel
approach of employing a ‘two-point’ binding and modified
fluorescence analysis procedure for developing porphyrin
chemosensors for selective detection of compounds of bio-
logical and societal importance are in progress.
DG, KJ DH, KJ DS, J K21
Porphyrin Alkaloid Ka, M21 a
mol21 a mol21
mol21
3
1
2
1
2
1
2
1
2
455.6 3 103
42.3 3 103
63.1 3 103
17.5 3 103
6.4 3 103
7.0 3 103
6.9 3 103
7.0 3 103
232.28 265.75 2112.31
226.39 247.21
227.38 249.12
224.21 240.25
221.69 226.13
221.92 226.44
221.91 226.34
221.93 226.33
269.87
272.96
253.83
214.89
215.17
214.87
214.77
4
5
(TPP)Zn
a At 298 K.
calculated from the Scatchard method11 of UV-visible absorp-
tion titration curves, is listed in Table 1. The binding constants
for (TPP)Zn binding to alkaloids under these solution condi-
tions is also given for comparison. The K values for 1 binding
to 3 is found to be nearly two orders of magnitude higher than
that observed for binding of either 1 to 5 or 1 to (TPP)Zn. The
binding of alkaloids, 1 or 2 to porphyrins 3 or 4 are stronger as
revealed by the K values and this effect could be attributed to the
‘two-point’ mode of binding. As expected, the K values for
binding of 1 or 2 to 5 are comparable to that observed for
alkaloid binding to (TPP)Zn indicating the absence of any
hydrogen bonding between the methyl ester group of 5 with the
pyrrolidine ring nitrogen of either 1 or 2. The calculated
thermodynamic parameters from the Van’t Hoff plots of lnK vs.
T21 for 5 or (TPP)Zn binding to 1 or 2 also draw similar
conclusions. Interestingly, both DH and DS decrease upon
binding to 3 or 4 as compared to that observed for binding to 5
or (TPP)Zn. These results indicate that the enthalpy change is a
main factor responsible for the observed higher stability of the
‘two-point’ bound porphyrin–alkaloid systems.
The authors are thankful to the donors of the Petroleum
Research Fund, administered by the American Chemical
Society, and Wichita State University for financial help.
Notes and references
† Free-base forms of porphyrins 3–5 were synthesized by reacting
stoichiometric amounts of pyrrole, benzaldehyde and the appropriate ortho
substituted benzaldehyde in propionic acid followed by column chromatog-
raphy purification on either basic alumina or silica gel. The ortho
substituted benzaldehydes, (2-formylphenoxy)acetic acid and (2-formyl-
phenoxy)acetamide were synthesized according to the literature procedure
given in ref. 7a and 7b, respectively. Zinc insertion was carried out
according to the standard procedure (ref. 8). The molecular integrity of all
the synthesized free-base and zinc porphyrins was established from FAB
1
1
mass, elemental analysis and H NMR studies (see Fig. 1 and text for H
NMR results).
‡ The details of the fluorescence quenching mechanism will be published
elsewhere.
Fig. 2 shows the fluorescence emission spectra of porphyrin
3 in the presence of various amounts of 1 in toluene. The zinc
porphyrin emission bands located at 605 and 650 nm decrease
in intensity with the appearance of an isosbestic point at 670 nm
indicating the presence of only one equilibrium in solution. It is
observed that the decrease in intensity of the 650 nm band is
much more than the 605 nm band.‡ Similar spectral features are
observed for porphyrin 3 or 4 binding with either compound 1
or 2.
1 (a) A. W. Banon, M. W. Decker, M. W. Holladay, P. Curzon, D.
Donnelly-Robers, P. S. Puttfarcken, R. S. Bitner, A. Diaz, A. H.
Dickenson, R. D. Porsolt, M. Williams and S. P. Arneric, Science, 1998,
279, 77; (b) M. W. Holladay, M. J. Dart and J. K. Lynch, J. Med. Chem.,
1997, 40, 4169; (c) R. M. Elgen, J. C. Hunter and A. Dray, Trends
Pharmacol. Sci., 1999, 20, 337; (d) M. B. Brennan, Chem. & Eng. News,
2000, 78, 23.
2 (a) N. L. Benowitz, Annu. Rev. Med., 1986, 37, 21; (b) P. E. McBride,
Med. Clin. North Am., 1992, 76, 333; (c) I. P. Stolerman and M. J. Jarvis,
Psychopharmacology, 1995, 117, 2; (d) K. A. Perkins, N. Benowitz, J.
Henningfield, P. Newhouse, O. Pomerleau and G. Swan, Addiction,
1996, 91, 129.
In order to quantitate these results, we monitored the intensity
ratio of these two bands as a function of alkaloid concentration.
3 (a) H. W. A. Teeuwen, F. J. W. Aalders and J. M. V. Rossum, Mol. Biol.
Rep., 1989, 13, 165 and references cited therein; (b) B. Siegmund, E.
Leitner and W. Pfannhauser, J. Agric. Food Chem., 1999, 47, 3117 and
references cited therein.
4 See for example, (a) Handbook of Biosensors and Electronic Noses,
Medicine, Food, and the Environment, ed. E. Kress-Rogers, CRC Press,
1997; (b) Biosensor and Chemical Sensor Technology, eds. K. R.
Rogers, A. Mulchandani, W. Zhou, ACS Symposium Series 613,
American Chemical Society, Washington, DC, 1995; (c) Biosensor
Technology, Fundamentals and Applications, eds. R. P. Buck, W. E.
Hatfield, M. Umana, E. F. Bowden, Marcel Dekker, Inc., 1990; (d) B.
Eggins, Biosensors, An Introduction, Wiley, New York, 1996.
5 (a) Fluorescent Chemosensors for Ion and Molecule Recognition, ed.
A. W. Czarnik, ACS Symposium Series 538, American Chemical
Society, Washington, DC, 1993; (b) A. P. de Silva, H. Q. N. Gunaratne,
T. Gunnlaugsson, A. J. M. Huxley, C. P. McCoy, J. T. Rademacher and
T. E. Rice, Chem. Rev., 1997, 97, 1515; (c) J. R. Lakowicz, Principles
of Fluorescence Spectroscopy, 2nd Edn. Kluwer/Plenum, New York,
1999.
6 The Porphyrin Handbook, ed. K. M. Kadish, K. M. Smith and R.
Guilard, vol 6, Academic Press, New York, 2000.
7 (a) Org. Synth., 1966, 46, 28; (b) Org. Synth. Coll. Vol. 4, 1963, 486.
8 G. D. Dorough, J. R. Miller and F. M. Huennekens, J. Am. Chem. Soc.,
1951, 73, 4315.
Fig. 2 Fluorescence emission spectrum of 3 (3.8 mM) in the presence of
various amounts of 1 in toluene (lex = 420 nm). The inset figure shows the
relationship between the intensity ratio of the emission bands, I604/I650 of 3
in the presence of (a) nicotine, (b) cotinine and, (c) pyridine substrates.
9 F. D’Souza, Y.-Y. Hsieh and G. R. Deviprasad, Inorg. Chem., 1996, 35,
5747.
10 M. Nappa and J. S. Valentine, J. Am. Chem. Soc., 1978, 100, 5075.
11 G. Scatchard, Ann. N. Y. Acad. Sci., 1949, 51, 661.
1916
Chem. Commun., 2000, 1915–1916