3-[2-(8-Quinolinyl)benzoxazol-5-yl]alanine derivative - A specific fluorophore for transition and rare-earth metal ion detection

Article abstract of DOI:10.1016/j.tet.2004.09.084

A novel aromatic amino acid, 3-[2-(8-quinolinyl)benzoxazol-5-yl]alanine derivative was synthesized as a potential Zn(II) and rare-earth metal, Eu(III) and Tb(III), ion chemosensor. The fluorophore was obtained using lead tetraacetate in DMSO to oxidize the Schiff base obtained from N-Boc-3-amino-tyrosine methyl ester and quinoline-8-carboxaldehyde. Preliminary photophysical properties of this ligand show that it possesses the properties necessary to be an effective chemosensor for Zn²⁺, Tb³⁺ and Eu³⁺ ions. Graphical Abstract

Full text of DOI:10.1016/j.tet.2004.09.084

Tetrahedron 60 (2004) 11889–11894
3-[2-(8-Quinolinyl)benzoxazol-5-yl]alanine derivative—a specific
fluorophore for transition and rare-earth metal ion detection
*
´ ´
Katarzyna Guzow, Magda Milewska, Dominik Wroblewski, Artur Giełdon and Wiesław Wiczk
´
´
Faculty of Chemistry, University of Gdansk, Sobieskiego 18, 80-952 Gdansk, Poland
Received 1 July 2004; revised 1 September 2004; accepted 23 September 2004
Available online 20 October 2004
Abstract—A novel aromatic amino acid, 3-[2-(8-quinolinyl)benzoxazol-5-yl]alanine derivative was synthesized as a potential Zn(II) and
rare-earth metal, Eu(III) and Tb(III), ion chemosensor. The fluorophore was obtained using lead tetraacetate in DMSO to oxidize the Schiff
base obtained from N-Boc-3-amino-tyrosine methyl ester and quinoline-8-carboxaldehyde. Preliminary photophysical properties of this
ligand show that it possesses the properties necessary to be an effective chemosensor for Zn2C, Tb3C and Eu3C ions.
q 2004 Elsevier Ltd. All rights reserved.
1. Introduction
2. Results and discussion
Considerable attention has been recently focused on the
development of compounds capable to respond selectively,
via changes in UV–vis absorption and fluorescence spectra,
to the presence of specific ions.1–8 Among numerous
analytical methods of cation detection, those based on
fluorescence sensors offer several distinct advantages in
terms of sensitivity, selectivity, response time and local
observation.1,5–8 The oldest class of fluorescence sensor
molecules are fluorescence ligands which are mostly
heteroaromatic ring systems often substituted by potentially
metal ion chelating functional groups which acts as both the
recognition and signaling site.4 Well known group of ligands
for heavy and transition metal ions consists of 8-amino or
8-hydroxyquinoline and its derivatives.9,10 Its selectivity/
sensitivity can be modified by appending to diaza-18-
crown-6.11,12 Depending on the 8-quinoline derivative
used, this chemosensor is selective to Zn2C or Cd2C
ions.9–12 In spite of the presence of numerous fluorescence
sensors,4–8 there is still a need for new ones—more selective
and sensitive. In this paper we describe a new chemosensor
containing an 8-quinoline moiety appended to position 2 of
3-(benzoxazol-5-yl)-alanine (Bx(8-Q)) (Fig. 1). This com-
pound has been synthesized according to the procedure
developed previously in our laboratory13,14 and it is a
representative of a group of new fluorescent aromatic amino
acids the spectral and photophysical properties of which
depend on the substituent in position 2.15
A new fluorophore, N-Boc-3-[2-(8-quinolinyl)benzoxazol-
5-yl]alanine methyl ester (Bx(8-Q)) (5), was synthesized
adapting the procedure published previously (Fig. 1).13,14
N-Boc-3-nitrotyrosine methyl ester (1) was reduced to
N-Boc-3-aminotyrosine methyl ester (2) by means of
catalytic hydrogenation in methanol (H2/Pd/C). The inter-
mediate Schiff base (4) was prepared in absolute ethanol
from 2 and quinoline-8-carboxaldehyde (3), obtained from
8-methylquinoline using selenium dioxide. Cyclization of
the Schiff base to the benzoxazole structure was performed
using lead tetraacetate in DMSO giving the product (5) in
10% yield.
The interaction of Bx(8-Q) with the following ions: LiC,
NaC, KC, Mg2C, CaC2, Ba2C, Sr2C, AgC, Co2C, Cd2C
,
Cu2C, Zn2C, Tb3C, Eu3C was studied by means of
absorption and steady-state fluorescence spectroscopy.
The studied ions can be divided into three groups. The
first contains alkaline and alkaline earth metal ions which do
not cause changes in the absorption spectrum of the ligand.
The exceptions are lithium and magnesium ions the
interaction of which with the ligand causes a small increase
of absorbance of the long-wave part of the absorption
spectrum, but these changes do not allow calculation of an
equilibrium constant. All ions from this group caused a
relatively small increase of the fluorescence quantum yield
without changing the shape or position of the emission band
of the ligand (Table 1).
Keywords: Chemosensor; Benzoxazolyl-alanine; Fluorophore; 8-
Quinoline.
Transition metal ions, belonging to the second group, cause
substantial changes of absorption and fluorescence spectra.
* Corresponding author. Tel.: C48 58 34 50 353; fax: C48 58 34 50 472;
e-mail: ww@chem.univ.gda.pl
0040–4020/$ - see front matter q 2004 Elsevier Ltd. All rights reserved.
doi:10.1016/j.tet.2004.09.084

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K. Guzow et al. / Tetrahedron 60 (2004) 11889–11894
Figure 1. Scheme of synthesis and structure of ligand (Bx(8-Q)).
A spectrophotometric and spectrofluorimetric titrations of
the ligand using Zn2C ion are shown in Figure 2.
which is additionally confirmed by the absorbance–
absorbance (A–A) diagram (Fig. 3).
An increase of metal ion concentration, up to a ligand/metal
ion ratio equal to one, causes an increase of absorbance of
the long-wave absorption band (at about 350, 275 and
240 nm) and a decrease at about 300 nm, moreover, sharp
isosbestic points can be detected. A further increase of the
ion concentration causes a blue shift of the absorption
spectrum with simultaneous broadening of isosbestic points
and the formation of a new peak at the long-wavelength
band of the spectrum. Such behaviour indicates on the
formation of a new type of the complex with different
stoichiometry to the previous one present in the solution,
The A–A diagram composes of two linear segments
intersecting at ligand:metal ratio about 1:1, indicating that
the system involves two titration steps.16 However, the
contribution of the second complex to the absorption spectra
is insufficient for the calculation of stability constants of
each complex formed, thus the overall stability constant
(log bZ11.7) is presented in Table 1. A relatively high
standard error of the calculated overall binding constant is a
result of the fact that the formation of a higher order
complex, the presence of which is visible in the A–A
diagram, has not been considered in calculations. Similar
Table 1. Absorption and fluorescence data for ligand and its metal ion complexes in acetonitrile
Absorption
Fluorescence
3
350 (MK1 cmK1
)
% of ligand 3350 (MK1 cmK1
)
log b
Q.Y.
% of ligand Q.Y.
t (ns)
L
Li
Na
K
Mg
Ca
Ba
Sr
LCd
L2Co
L2Ni
L2Cu
L2Zn
LEu
LTb
Ag
2550
4430
2550
2550
6250
2550
2550
2550
15,690
14,920
15,400
22,040
16,570
10,590
10,270
2550
—
74
0
0
145
0
0.024
0.093
0.032
0.04
0.223
0.026
0.037
0.046
1
0.004
0.003
0.005
0.955
0.12
—
387
133
167
930
108
154
192
4167
17
12
21
3980
500
458
480
nd
nd
0
0
485
485
505
764
550
315
303
0
5.14G0.15
10.96G0.50
11.47G0.45
8.12G1.98
11.70G3.12
7.57G0.78
4.33G0.26
—
4.98
5.68
4.38
4.39
0.11
0.116
nd, not determined.

K. Guzow et al. / Tetrahedron 60 (2004) 11889–11894
11891
at 435 nm) versus a metal/ligand molar ratio are presented.
The absorbance and fluorescence intensity curves versus
metal/ligand ratio differ substantially, especially for the
Cd2C ion. Lower values of normalized fluorescence
intensity compared to the normalized absorbance (less
pronounced for Zn2C ion) clearly indicate lower stability of
the complex in the excited state compared to the ground
state. In both cases, up to the M:L ratio of about 0.25, an
invariant fluorescence intensity can be observed indicating
dissociation of the excited complex. Because of that, the
stability constants of the complexes in the excited state
cannot be determined. Additionally, an increase of Zn2C ion
concentration above the 1:2 M:L ratio causes the quenching
of complex fluorescence because of the formation of a new
1:1 ligand:Zn2C complex, confirmed by the intensity–
intensity (I–I) diagram (Fig. 5).
Figure 2. Absorption and fluorescence titrations of the ligand with Zn2C
ion in acetonitrile.
behaviour was also observed for the complexes of the ligand
with Co2C, Cu2C and Ni2C ions, whereas for Cd2C, the
A–A diagram reveals the presence of only one type of
complex (ML) with the logarithm of stability constant equal
to about 5 (Table 1) (data not shown). The calculated
stoichiometry reveals that all ions mentioned in this group
form ML2 complexes with relatively high overall stability
constants (log b about 11, except Cu2C for which log b is
about 8, Table 1). As in the case of Zn2C, it was impossible
to calculate the individual stability constant for the other
ML complexes.
Not only are stoichiometry and ground state stability
constant different for the Zn2C and Cd2C complexes, but
also the fluorescence lifetimes are different. For the Cd2C
complex (for M:L ratio about 16) the fluorescence lifetime
is 4.98 ns (lemZ430 nm, c2RZ1.18) whereas for the Zn2C
complex the fluorescence lifetime is 5.68 ns (lemZ430 nm,
c2RZ1.10).
The third class of ions consists of the luminescent rare-earth
metal ions, Eu3C and Tb3C. Spectrophotometric and
spectrofluorimetric titrations are presented in Figure 6. As
for the transition metal ions, the addition of Eu3C or Tb3C
to the ligand solution in acetonitrile causes a formation of a
new absorption band with a maximum at about 370 nm and
simultaneously an increase of the absorbance at about
250 nm and decrease in the range of 300–350 nm.
A common property of the transition metal ions studied is
the almost invariable position of the fluorescence spectrum
of the ligand. For Zn2C and Cd2C ions, the fluorescence
band of the ML2 complex was shifted bathochromically by
about 15 nm compared to the emission spectrum of the pure
ligand (Fig. 2). The property which differentiates the studied
ions is the fluorescence quantum yield of a given complex
measured at high metal-to-ligand ratio (M:LO20). The
divalent paramagnetic ions (Cu2C, Ni2C, Co2C) quench
almost totally the ligand fluorescence, whereas diamagnetic
ions (Zn2C and Cd2C) enhance the fluorescence intensity,
resulting in a fluorescence quantum yield equal to 1.0 for
Cd2C ion and 0.95 for Zn2C (close to 1.0 at M:LZ1:1)
(Table 1). Such behaviour is typical for this type of ligand
and ions.4 In Figure 4 the changes of normalized absorbance
(measured at 350 nm) and fluorescence intensity (measured
The A–A diagrams for Eu3C and Tb3C ions are straight
lines, indicating on the formation of only one type of the
complex. The calculation reveals that the stoichiometry of
these complexes are ML with binding constants about two
times higher for Eu3C than Tb3C ion (Table 1).
In contrast to the transition metal ions, increasing the Eu3C
or Tb3C ion concentration causes a decrease of the ligand
Figure 4. Changes in the normalized absorbance and fluorescence quantum
yield upon addition of Zn(ClO4)2 or Cd(ClO4)2 to the solution of Bx(8-Q) in
acetonitrile.
Figure 3. Absorbance–absorbance (A–A) diagram for titration of Bx(8-Q)
with Zn2C ion.

11892
K. Guzow et al. / Tetrahedron 60 (2004) 11889–11894
Figure 7. Changes in the normalized absorbance and fluorescence quantum
yield upon addition of Eu(ClO4)3 or TbCl3 to the solution of Bx(8-Q) in
acetonitrile.
Figure 5. Intensity–intensity (I–I) diagram for titration of Bx(8-Q) with
Zn2C ion.
emission band and simultaneously an increase of a new
emission band centered at about 530 nm. In Figure 7 the
dependencies of the changes of the normalized absorption
measured at 365 nm and emission intensities of the ligand
(at 425 nm) and complex intensity (at 525 nm) versus the
molar ratio of metal to ligand are depicted.
A different behaviour, compared to above mentioned ions,
was observed during a titration with AgC ion. With the
increase of AgC ion concentration there was no change in
the absorption spectrum of the ligand, as in the case of large
alkaline and alkaline earth metal ions (NaC, KC, Ca2C
,
Ba2C, Sr2C), whereas in the excited state this ion interacted
with the ligand similarly to the Eu3C and Tb3C, causing an
increase of the long-wave fluorescence band with the
maximum at 530 nm up to the M:L ratio higher than 20.
As in the case of the Zn2C and Cd2C complexes, the curves
displaying the complex fluorescence increase are situated
lower than the corresponding absorption curves, indicating a
lower constant binding in the excited state compared to the
ground state. On the initial part of these curves a flat region
is clearly seen confirming the differences in stability
constants in the ground and excited state. The difference
in the ground state stability constant as well as the different
shape of the curves illustrating the absorption or emission
changes with the increase of metal ion concentration
between the Eu3C and Tb3C ions, having very similar ion
radii and electron configurations, is probably due to the
influence of the anion on the complex formation because
europium perchlorate and terbium chloride were used for
the titration. However, the fluorescence quantum yield
(0.12) and the fluorescence lifetime (4.38 ns), (Eu:L
ratioZ5, Tb:L ratioZ16) of both complexes are very
similar (Table 1).
The different interactions of ions studied with the ligand
cannot be explained taking into account ionic radii only.
The change of the absorption spectrum of the ligand caused
by small ions, LiC and Mg2C, and the lack of such changes
in the case of alkaline or alkaline earth metal ions as well as
AgC having ionic radii greater than 1 A and simultaneous
˚
changes in absorption spectrum caused by transition metal
ions or luminescent rare-earth metal ions having ionic radii
similar to Ca2C or NaC indicates that not only ionic radius
but also the polarizability and/or second ionization potential
should be considered.17 While the quenching of ligand
fluorescence is predominantly connected with the nature of
metal ion, the ligand fluorescence enhancement can result
from changes of geometry in the excited state or be induced
by the ion. The fact that all ions studied alter the
fluorescence properties of the ligand suggests chelation-
induced changes of the relatively close lying ligand centered
np* and pp* states. The lower binding affinity in the excited
state is probably caused by the lower, compared to the
ground state, charge density on the benzoxazolyl nitrogen
atom and the increase of the dihedral angle between the
benzoxazolyl and quinolinyl moiety (for about 208) as
results from the theoretical calculations using a semi-
empirical molecular-orbital PM3 method18 with semi-
conductor-like screening model for solvation (COSMO)19
implementated in the MOPAC 2002 package.20 The
increase of the ligand flexibility in the excited state, caused
by a decrease of interaction between two aromatic moieties,
probably facilitates adjusting the ligand chelation cavity to
an ion and simultaneously suppresses the radiationless
deactivation via torsional motions. The above discussion
does not fully explain all observed phenomena and
Figure 6. Absorption and fluorescence titrations of the ligand with Tb3C
ion in acetonitrile.

K. Guzow et al. / Tetrahedron 60 (2004) 11889–11894
11893
additional work are currently ongoing in our laboratory and
will be published in due course.
excitation source was a NanoLed 03 (UV led 370 nm) from
IBH. The half-width of the response function of apparatus
measured using a Ludox solution as a scatter was about
1.0 ns. The excitation wavelength was isolated using an
interference filter (lmaZ372 nm, spectral band-width about
7 nm) whereas emission wavelengths were isolated using a
monochromator (about 12 nm spectral band-width).
Fluorescence decay data were fitted by the iterative
convolution to the sum of exponents
3. Conclusions
The formation of highly fluorescent complexes of Bx(8-Q)
ligand with the transition metal ions Cd2C and Zn2C as well
as with rare-earth metal ion and substantial changes in the
absorption spectra and fluorescence lifetime cause that this
ligand can be used as a chemosensor for these ions.
Moreover, much higher constant binding determined for
Zn2C ion over other ions studied indicate that Bx(8-Q) is
preferred as a chemosensor for this particular ion. However,
the described ligand possesses a remarkable advantage over
other chemosensors for Zn2C ion presented in the literature.
It has an amino acid moiety allowing it to be incorporated
into a peptide chain and/or used for the synthesis of a
fluorescent cell-permeable compound for the study of
intracellular zinc chemistry.
X
IðtÞ Z
aiexpðKt=tiÞ;
(1)
i
where ai is the pre-exponential factor obtained from the
fluorescence intensity decay analysis and ti is the decay
time of the ith component. The adequacy of the exponential
decay fitting was judged by visual inspection of the plots of
weighted residuals and by the statistical parameter c2R and
shape of the autocorrelation function of the weighted
residuals and serial variance ratio (SVR).
4.2. Synthesis
4. Experimental
4.1. General
4.2.1. Quinoline-8-carboxaldehyde (3). 8-Methylquino-
line (10 g, 70 mmol) was heated to 100 8C. Selenium
dioxide (7.91 g, 71 mmol) was added in small portions
with stirring during 2 h. Then the reaction mixture was
cooled to the room temperature and extracted with boiling
methanol. After solvent evaporation, the residue was
crystallized from MeOH/diethyl ether. The obtained yellow
solid was purified by means of column chromatography
using as an eluent AcOEt/petroleum ether (1:5).
Recrystallization from AcOEt/petroleum ether gave the
title compound (0.80 g, 5.08 mmol, 7%) as a white solid
(RfZ0.49, AcOEt/petroleum ether (1:3); tRZ12.8 min).
8-Methylquinoline, selenium dioxide and lead tetraacetate
were purchased from Lancaster whereas 3-nitro-L-tyrosine
was purchased from Fluka. The following compounds were
prepared according to literature procedures: 3-nitro-L-
tyrosine methyl ester23 and N-Boc-3-nitro-L-tyrosine methyl
ester.24 Quinoline-8-carboxaldehyde was synthesized
according to a modified literature procedure.25 N-Boc-3-
[2-(8-quinolinyl)benzoxazol-5-yl]alanine methyl ester was
synthesized adapting the procedure published previously.13,14
TLC was carried out on Merck silica gel plates (Kieselgel
60 F254). The spots were revealed using a UV lamp (254 nm,
366 nm). All solvent ratios are in volume parts. The
purification was carried out by means of column chroma-
tography (Merck, Silica gel 60, 0.040–0.063 mm). The
purity of the obtained compounds was checked by means of
analytical RP-HPLC (Kromasil column, C-8, 5 mm, 250 mm
long, IDZ4.5 mm) with detection at 223 nm. The mobile
phase was a gradient running from 0 to 100% of B (AZ
water with addition of 0.01% trifluoroacetic acid, BZ80%
of an aqueous solution of acetonitrile with addition of 0.08%
trifluoroacetic acid) over 60 min. Melting points (mp) were
determined in capillary tubes using Gallenkamp Griffin
Mp 96–97 8C; (Found: C, 75.98; N, 8.82; H, 4.42. C10H7NO
requires C, 76.42; N, 8.91; H, 4.49%); nmax (KBr) 3347.4
(w), 2868.5 (m), 2711.4 (w), 1681.5 (s), 1572.6 (s), 1498.4
(s), 1403.2 (s), 1320.2 (s), 1248.0 (s), 1164.9 (m), 1131.1
(m), 1035.9 (m), 873.1 (s), 828.8 (s), 787.1 (s), 766.4 (s),
642.5 (s) cmK1; dH (400 MHz, CDCl3) 11.5 (1H, s, CHO),
0
9.06 (1H, dd, JZ1.6, 4.0 Hz, C2 H), 8.34 (1H, dd, JZ1.6,
0
0
7.4 Hz, C7 H), 8.25 (1H, dd,0 JZ1.2, 8.2 Hz, C4 H), 8.11
(1H, dd, JZ1.6, 8.2 Hz, C5 H), 7.68 (1H, t, JZ7.8 Hz,
0
0
C6 H), 7.52 (1H, dd, JZ4.0, 8.4 Hz, C3 H); dC (100 MHz,
0
0
CDCl3) 192.58 (CHO), 151.505 (C2 ), 147.84 (C9 ), 140.00
0
0
0
0
(C7 ), 136.53 (C4 ), 134.43 (C5 ), 131.95 (C8 ), 129.55 (C10 ),
0
0
126.46 (C6 ), 122.03 (C3 ); m/z (FAB) 158 (80, MHC), 131
(100%).
1
MPA-350.MB2.5 apparatus and are uncorrected. H NMR
and 13C NMR spectra were recorded on a Varian, Mercury-
400 BB spectrometer (400 MHz) in CDCl3. Infrared spectra
were recorded on a Bruker IFS-66 instrument. Mass spectra
were recorded on a MASSLAB TRIO-3 instrument (FAB)
or Bruker Biflex III (MALDI-TOF). Elemental analysis was
achieved on a Carlo Erba CNSO Eager 200 instrument.
Absorption spectra were measured on a Perkin–Elmer
Lambda 40P spectrophotometer. Fluorescence spectra
were measured on a Perkin–Elmer LS-50B spectrofluori-
meter. MeCN used in our studies was HPLC grade.
Quantum yields (QY) were calculated using as a reference
quinine sulphate in 0.5 M H2SO4 (QYZ0.53G0.0221). The
fluorescence lifetimes were measured with a time-correlated
single-photon counting apparatus Edinburgh CD-900. The
4.2.2. N-(tert-Butyloxycarbonyl)-3-[2-(8-quinolinyl)ben-
zoxazol-5-yl]-L-alanine methyl ester (5). A mixture of N-
Boc-3-nitro-L-tyrosine methyl ester (1.38 g, 4.08 mmol) and
5% palladium on active carbon in MeOH (30 mL) was
stirred under a hydrogen atmosphere at room temperature
for 90 min (TLC monitoring (CH2Cl2/MeOH/AcOH
100:10:1), RfZ0.72). The catalyst was filtered off and the
solvent evaporated in vacuo to give the brownish, oily
product which was dissolved in absolute EtOH (5 mL) and
mixed with the solution of quinoline-8-carboxaldehyde
(0.59 g, 3.76 mmol) in absolute EtOH (7 mL). The
mixture was stirred at rt overnight (TLC monitoring

11894
K. Guzow et al. / Tetrahedron 60 (2004) 11889–11894
(AcOEt/petroleum ether 2:5), RfZ0.67), giving the Schiff
base as a yellow solid (1.6 g, 3.56 mmol, 87%). The
obtained Schiff base was filtered off and dissolved in
DMSO (10 mL) and lead tetraacetate (2.78 g, 6.27 mmol)
was added. The mixture was stirred at rt for about an hour
(TLC monitoring (AcOEt/petroleum ether 1:1), RfZ0.32)
and then dissolved in ethyl acetate (AcOEt) and washed in
turns with a saturated aqueous solution of NaCl (!1), a 5%
solution of NaHCO3 (!2), a saturated aqueous solution of
NaCl (!3) and dried over anhydrous MgSO4. The solvent
was evaporated in vacuo and the product was isolated by
means of column chromatography using as an eluent
AcOEt/petroleum ether (1:1). The crude product was
recrystallized from AcOEt/petroleum ether, giving the
title compound (0.17 g, 0.39 mmol, 10%) as a white solid
(tRZ36.2 min).
References and notes
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Mp 106–107 8C; (Found: C, 67.40; N, 9.32; H, 5.71.
C25H25N3O5 requires C, 67.10; N, 9.39; H, 5.63%); nmax
(KBr) 3358.5 (m), 2969.0 (w), 1751.6 (s), 1736.4 (s), 1686.0
(s), 1525.5 (s), 1439.4 (m), 1351.5 (m), 1297.9 (m), 1259.0
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(s), 1210.9 (m), 1166.4 (s), 1056.4 (w), 791.6 (m) cmK1; dH
0
(400 MHz, CDCl3) 9.18 (1H, dd, JZ2.0, 4.2 Hz, C2 H),
0
8.54 (1H, dd, JZ1.2, 7.4 Hz, C5 H), 8.27 (1H, dd, JZ2.0,
10. Kimber, M. C.; Mahadevan, B. M.; Lincoln, S. F.;
Ward, A. D.; Tiekink, E. R. T. J. Org. Chem. 2000, 65,
8204–8209.
0
0
8.4 Hz, C4 H), 8.03 (1H, dd, JZ1.2, 8.2 Hz, C7 H), 7.71
0
(1H, d, JZ7.2 Hz, C6 H), 7.68 (1H, s, C4H), 7.60 (1H, d, JZ
0
8.0 Hz, C7H), 7.53 (1H, dd, JZ4.4, 8.2 Hz, C3 H), 7.16 (1H,
dd, JZ1.6, 8.2 Hz, C6H), 5.03 (1H, d, JZ8.0 Hz, NH), 4.64
(1H, dd, JZ5.6, 13.6 Hz, CaH), 3.74 (3H, s, OCH3), 3.23–
11. Prodi, L.; Montalti, M.; Zaccheroni, N.; Bradshaw, J. S.; Izatt,
R. M.; Savage, P. B. Tetrahedron Lett. 2001, 42, 2941–2944.
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Zaccheroni, N. Tetrahedron 2001, 57, 7623–7628.
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Acta 2001, 84, 1086–1092.
3.28 (2H, m, CbH2), 1.40 (9H, s, (CH3)3); dC (100 MHz,
0
CDCl3) 172.36 (CO)0, 157.00 (C2), 152.08 (C2 ), 152.00
(HNCO), 146.09 (C9 ), 146.00 (C8), 136.85 (C9), 132.91
0
0
0
0
(C8 ), 132.57 (C5), 131.97 (C4 ), 128.99 (C10 ), 126.71 (C5 ),
0
0
126.48 (C6), 126.20 (C6 ), 121.94 (C3 ), 121.15 (C4), 110.90
(C7), 80.12 (C(CH3)3), 54.84 (Ca), 52.54 (OCH3), 38.47
(Cb), 28.53 ((CH3)3); m/z (MALDI-TOF) 448 (MHC).
14. Guzow, K.; Szabelski, M.; Malicka, J.; Karolczak, J.; Wiczk,
W. Tetrahedron 2002, 58, 2201–2209.
15. Guzow, K.; Mazurkiewicz, K.; Szabelski, M.; Ganzynkowicz,
R.; Karolczak, J.; Wiczk, W. Chem. Phys. 2003, 295, 119–130.
16. Polster, J.; Lachmann, H. Spectrometric Titrations. Analysis of
Chemical Equilibria; VCH: New York, 1989.
4.3. UV–vis and fluorescence titration experiments
A solution of ligand (concentration about 5!10K5 M for
absorption and 5!10K6 M for emission) in acetonitrile was
treated with increasing amounts of a solution of perchlorate
salts of the cation of interest, except for terbium ions
where the chloride salt was used, (concentration about
5!10K4 M) containing a ligand at the same concentration
as in the cuvette at room temperature. After each addition of
aliquots using Hamilton microsyringe with micrometer
screw, the UV–vis or fluorescence spectrum was recorded.
When the titration was complete the spectra were
implemented into the STOICHIO software for binding
constant calculations.22
17. Irving, H.; Wiliams, R. J. P. Nature (London) 1948, 162,
746–747.
18. Steward, J. J. P. J. Comput. Chem. 1986, 10, 221–264.
19. Klamt, A.; Schu¨rmann, G. I. J. Chem. Soc., Perkin Trans. 2
1993, 799–805.
20. Steward, J. J. P. MOPAC 2002, Fujitsu Limited, 2001.
21. Lakowicz, J. R. Principles of Fluorescence Spectroscopy,
2nd ed.; Kluwer Academic/Plenum: New York, 1999.
22. Kostrowicki, J.; Liwo, A. Comput. Chem. 1987, 11,
195–210.
23. Wunsch, E. In Mhller, E., Ed.; Synthese von Peptiden;
Houben-Weyl, Methoden der organischen Chemie; Georg
Thieme Verlag: Stuttgart, 1974; Vol. 15, p 317.
24. Stewart, J. M.; Young, J. D. Solid Phase Peptide Synthesis;
Chemical Company: Rockford, IL, 1984; p 63.
¨
Acknowledgements
25. Teague, C. E. Jr.; Roe, A. J. Am. Chem. Soc. 1951, 73,
688–689.
This work was supported by the State Committee for
Scientific Research (KBN Poland) under grant
1005/T09/2003/24.