A preparative, spectroscopic anti equilibrium study of some phenyl-2- thiazoline fluorophores for aluminium(III) detection

Article abstract of DOI:10.1039/b001905o

As part of a study of fluorophores for selective Al³⁺ detection and of potential use in the study of intracellular Al³⁺, the preparative, UV- visible and fluorescence spectroscopic, and Al³⁺ coordination characteristics of five ligands containing the conjugated phenolic substituted thiazoline chromophore of the natural siderophore, pyochelin, 2- [2-(2-hydroxyphenyl)-4,5-dihydro-1,3-thiazol-4-yl]-3-methyl-1,3-thiazolane-4- carboxylic acid) 1, are reported. The five ligands are 2-(2-hydroxyphenyl)- 4,5-dihydro-1,3-thiazole-4-carboxylic acid, 2, 2-(4,5-dihydro-1,3-thiazol-2- yl)phenol, 3, 4-bromo-2-(4,5-dihydro-1,3-thiazol-2-yl)phenol, 4, 2-(4,5- dihydro-1,3-thiazol-2-yl)-5-nitrophenol, 5, and 2-(4,5-dihydro-1,3-thiazol-2- yl)-4-nitrophenol, 6. All form stable binary Al³⁺ complexes in 75% methanol and 25% water in the case of 2, and 73.2% methanol, 24.4% water and 2.4% N,N- dimethylformamide in the case of 3-6, and exhibit substantial UV-visible absorbance changes on coordination. However, while 2, 3 and 4 fluoresce strongly when coordinated to Al³⁺, 5 and 6 do not, probably because their fluorescence is quenched through a twisted intramolecular charge transfer process. All five ligands show selectivity in complexing for Al³⁺ over other metal ions and, in the case of 2, 3 and 4, represent an initial stage in the development of Al³⁺ specific fluorophores for biological use.

Full text of DOI:10.1039/b001905o

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A preparative, spectroscopic and equilibrium study of some
phenyl-2-thiazoline Ñuorophores for aluminium(III) detection
Stephanie G. Lambert, Jo-Anne M. Taylor, Kate L. Wegener, Susan L. Woodhouse,
Stephen F. Lincoln* and A. David Ward
Department of Chemistry, University of Adelaide, Adelaide, SA 5005, Australia.
E-mail: stephen.lincoln=adelaide.edu.au
Received (in Montpellier, France) 8th March 2000, Accepted 12th April 2000
Published on the Web 6th June 2000
As part of a study of Ñuorophores for selective Al3` detection and of potential use in the study of intracellular
Al3`, the preparative, UVÈvisible and Ñuorescence spectroscopic, and Al3` coordination characteristics of Ðve
ligands containing the conjugated phenolic substituted thiazoline chromophore of the natural siderophore,
pyochelin, 2-[2-(2-hydroxyphenyl)-4,5-dihydro-1,3-thiazol-4-yl]-3-methyl-1,3-thiazolane-4-carboxylic acid) 1, are
reported. The Ðve ligands are 2-(2-hydroxyphenyl)-4,5-dihydro-1,3-thiazole-4-carboxylic acid, 2, 2-(4,5-dihydro-1,3-
thiazol-2-yl)phenol, 3, 4-bromo-2-(4,5-dihydro-1,3-thiazol-2-yl)phenol, 4, 2-(4,5-dihydro-1,3-thiazol-2-yl)-5-
nitrophenol, 5, and 2-(4,5-dihydro-1,3-thiazol-2-yl)-4-nitrophenol, 6. All form stable binary Al3` complexes in 75%
methanol and 25% water in the case of 2, and 73.2% methanol, 24.4% water and 2.4% N,N-dimethylformamide in
the case of 3È6, and exhibit substantial UVÈvisible absorbance changes on coordination. However, while 2, 3 and 4
Ñuoresce strongly when coordinated to Al3`, 5 and 6 do not, probably because their Ñuorescence is quenched
through a twisted intramolecular charge transfer process. All Ðve ligands show selectivity in complexing for Al3`
over other metal ions and, in the case of 2, 3 and 4, represent an initial stage in the development of Al3` speciÐc
Ñuorophores for biological use.
Aluminium is the third most abundant element in the earthÏs
crust, but until recently its biological availability has been low
because it is predominantly bound as Al3` in aluminosilicates
the detection of Al3` it is necessary that the excited state of
the pyochelin chromophore is sufficiently stabilised through
Al3` coordination to generate signiÐcant Ñuorescence. The
ligands studied are 2-(2-hydroxyphenyl)-4,5-dihydro-1,3-
thiazole-4-carboxylic acid, 2, 2-(4,5-dihydro-1,3-thiazol-2-yl)
phenol, 3,4-bromo-2-(4,5-dihydro-1,3-thiazol-2-yl)phenol, 4,
2-(4,5dihydro-1,3-thiazol-2-yl)-5-nitrophenol, 5, and 2-(4,5-
dihydro-1,3-thiazol-2-yl)-4-nitrophenol, 6. This appears to be
one of the Ðrst systematic studies of complexation by Al3`
speciÐc Ñuorophores.
and [Al(OH) ], which have low aqueous solubilities.1h4
3
However, the advent of acid rain in industrial regions has
lowered soil pH and released Al3` into ground water with a
deleterious e†ect on forest and aquatic systems. The use of
aluminium utensils and containers for food and beverage
handling,5,6 and Al3` containing pharmaceuticals2 have also
increased Al3` bioavailability. Although at the physiological
pH of 7.4 the low solubility of Al(OH) limits uncomplexed
3
[Al3`] to O10~11 mol dm~3 in plasma, its concentration as
citrate, transferrin and adenosine and guanosine triphosphate
complexes may be substantially higher.7,8 Consequently, there
has been increasing interest in the transport of Al3` at the
cellular level,9,10 and a need for the development of a sensitive
non-destructive detection method for intracellular Al3`. Fluo-
rescent emission from Ca2` and Zn2` speciÐc Ñuorophores
has proved to be a particularly sensitive method for determin-
ing the in situ variations of the intracellular levels of these
ions.11h15 This study represents an initial phase in the devel-
opment of Al3` speciÐc Ñuorophores for intracellular use.
Pyochelin, 2-[2-(2-hydroxyphenyl)-4,5-dihydro-1,3-thiazol-
4-yl]-3-methyl-1,3-thiazolane-4-carboxylic acid, 1, isolated
from Psedomonas aeruginosa, is a siderophore that forms a red
complex with Fe3`.16 Because of its propensity to coordinate
Fe3`, pyochelin is likely to coordinate another hard Lewis
acid, Al3`, and thereby provide a starting point for the devel-
opment of Al3` speciÐc Ñuorophores. The pyochelin chromo-
phore is the conjugated phenolic substituted thiazoline
moiety, which exhibits absorbance maxima at 218 (10 500),
248 (10 500) and 310 nm (4200 dm3 mol~1 cm~1) where the
Ðgures in parentheses are molar absorbances.16 Accordingly,
this study is predominantly concerned with the coordination
by Al3` of ligands containing the pyochelin chromophore and
the e†ect of such coordination on the UVÈvisible absorbance
and Ñuorescence of these ligands. To act as a Ñuorophore for
Results and discussion
Ligand synthesis and characterisation
Reported synthetic methods were used to obtain 2 and 3 in
good yield.17,18 The general method for the preparation of the
DOI: 10.1039/b001905o
New J. Chem., 2000, 24, 541È546
541
This journal is ( The Royal Society of Chemistry and the Centre National de la Recherche ScientiÐque 2000

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thiazolines 4È6 is shown in Scheme 1. Conversion of a
benzaldehyde derivative to the corresponding benzonitrile
derivative by reÑuxing with hydroxylamine hydrochloride in
formic acid was followed by formation of the thiazoline ring
through reaction of the benzonitrile derivative with 2-
aminoethanethiol hydrochloride in the presence of tri-
ethylamine. To obtain sufficient solubility of the ligands for
the studies discussed below it was necessary to use a 75%
methanolÈ25% aqueous (v : v) solvent mixture to make up
solutions. Because of their slow dissolution rates, stock solu-
tions of 3È6 were prepared in N,N-dimethylformamide (DMF)
so that their Ðnal solutions were 73.2% methanol, 24.4%
water and 2.4% DMF by volume. Under the conditions of
this study 2 exhibits two pK s and 3È6 one pK each as listed
in Table 1. Ligands 2È6 show strong absorbance in the UVÈ
visible region but none shows signiÐcant Ñuorescence in the
free state.
It is usually necessary for biological probe molecules to
possess signiÐcant lipophilic character so that they can tra-
verse the cell membrane. As a consequence, such molecules
may possess low aqueous solubilities and are consequently
studied in aqueous solvent mixtures to increase their concen-
to maintain a constant [H`]. However, 3È6 do not complex
Al3` at this high [H`] and accordingly their coordination
was studied in the absence of added acid. These observations
are consistent with Al3` competing with H` for coordination
sites in the dianion of 2, 2@, which probably acts as a tridentate
ligand while the monoanions of 3È6, 3@È6@, probably act as
bidentate ligands as shown in Schemes 2 and 3, respectively.
Ligand 2. The variation of the UV-visible spectrum of 2
with [Al3`]
is shown in Fig. 1, from which it is seen that
total
three isosbestic points (259, 279 and 328 nm) exist consistent
with the chromophore experiencing two dominant environ-
ments. Fitting of an algorithm for 1 : 1 complexation to the
absorbance data in the ranges 290È315 nm and 335È375 nm
gave a formation constant K \ 2390 ^ 20 dm3 mol~1 for
[Al(2@)]` and a derived spectrum of coordinated 2@ with
maxima at 270 (14 900) and 350 nm (6600 dm3 mol~1 cm~1),
where the molar absorbances appear in parentheses. Under
the same conditions, the spectrum of free 2 is characterised by
maxima at 258 (9800), 275 (7700) and 311 (5600) and a shoul-
der at 350 nm (2600 dm3 mol~1 cm~1). The Ðtting of algo-
rithms incorporating the additional formation of either 1 : 2
or 2 : 1 complexes to these data showed no evidence for their
formation in either this system or in the other systems con-
sidered below.
a
a
trations to those required for pK determinations and UVÈ
a
visible spectroscopic studies as is the case in this work. A
similar situation prevailed for our Zn2` speciÐc Ñuoro-
phore,
2-methyl-8-(toluene-p-sulfonamido)-6-quinolyloxy-
The isosbestic point at 328 nm was chosen as the excitation
wavelength for the Ñuorescence study of the coordination of 2
by Al3`. The concentration of 2 was held constant at
3.80 ] 10~7 mol dm~3 to optimise the Ñuorescence measure-
ments over the range 400È500 nm in the presence of Al3`. The
acetic acid,15 which has been successfully used in the study of
intracellular Zn2`.13,14
UV–visible and Ñuorescence equilibrium studies
increase in Ñuorescence of 2 with [Al3`]
is shown in Fig. 2,
from which it is seen that the broad Ñuorescence band shows
total
Hydrated Al3` is strongly acidic and precipitates as
[Al(OH) ] at pH P 5. This precluded the use of pH-titrimetric
a systematic increase in intensity with increase in [Al3`]
.
3
total
methods use to determine the pK s of 2È6 for the determi-
When an algorithm for 1 : 1 complexation is Ðtted to these
a
nation of Al3` coordination of these ligands. However, it was
both convenient and necessary to study the coordination of
ligands 2È6 by UVÈvisible and Ñuorimetic methods to assess
their suitability as Al3` speciÐc Ñuorophores. Most available
bu†ers either complex Al3`, or cause it to precipitate as a salt,
or both, under the conditions of this study. Thus, the coordi-
nation of 2 was studied in 10~3 mol dm~3 HClO solutions
4
Scheme 2
Scheme 1
Scheme 3
Table 1 Ligand 2È6 pK s and stability constants (K) of the 1 : 1 complexes of their conjugate bases 2ºÈ6º with Al3` at 298.2 K
a
K/dm3 mol~1b
(UVÈvisible)
K/dm3 mol~1b
Liganda
2/2@
pK
(Ñuorescence)
a
4.62 ^ 0.01c
11.98 ^ 0.01c
(2.39 ^ 0.02) ] 103d
(1.83 ^ 0.01) ] 103d
3/3@
4/4@
5/5@
6/6@
[12.0
(5.57 ^ 0.12) ] 103
(7.08 ^ 0.11) ] 103
(1.20 ^ 0.04) ] 104
(4.48 ^ 0.20) ] 104
(3.19 ^ 0.04) ] 103
(3.89 ^ 0.03) ] 103
11.12 ^ 0.01
9.81 ^ 0.01
8.38 ^ 0.01
a For 2 all solutions were 75% methanol and 25% water, while for 3È6 solutions were 73.2% methanol, 24.4% water and 2.4% DMF by volume.
In all solutions I \ 0.10 mol dm~3 (NaClO ). b The K values determined by UVÈvisible spectrophotometry are \2 greater than those deter-
4
mined by Ñuorimetry. This may be because the concentrations of 2, 3 and 4 were 315, 10 and 10 times greater in the spectrophotometric studies
than in the Ñuorimetric studies, respectively; while the [Al3`]
ranges were similar in both types of study, the equilibrium shown in Scheme 2
total
and the analogous equilibria for 3 and 4 were slightly more to the left in the Ñuorimetric studies. c pK s for the carboxylic acid and phenol
a
groups, respectively. d Also 0.001 mol dm~3 in HClO .
4
542
New J. Chem., 2000, 24, 541È546

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Fig. 3 Variation of the UVÈvisible absorption spectrum of
1.20 ] 10~4 mol dm~3 4 with added Al(ClO ) in 73.2% methanol,
Fig. 1 Variation of the UVÈvisible absorption spectrum of
4 3
24.4% water and 2.4% DMF by volume solution, 0.10 mol dm~3 in
1.20 ] 10~4 mol dm~3 2 with added Al(ClO ) in 75% methanol and
4 3
NaClO at 298.2 K. The absorbance increases at 270 and 360 and
25% water by volume solution, 0.10 mol dm~3 in NaClO and 0.001
4
4
decreases at 320 nm as [Al(ClO ) ] increases in the sequence: 0,
mol dm~3 in HClO at 298.2 K. The absorbance increases at 270 and
4 3
4
2.0 ] 10~5, 6.0 ] 10~5, 1.0 ] 10~4, 1.5 ] 10~4, 2.0 ] 10~4,
350 and decreases at 300 nm as [Al(ClO ) ] increases in the sequence:
4 3
3.0 ] 10~4, 4.0 ] 10~4, 5.0 ] 10~4, 6.0 ] 10~4, 7.0 ] 10~4,
8.0 ] 10~4, 9.0 ] 10~4, 1.00 ] 10~3, 1.20 ] 10~3 and 1.40 ] 10~3
mol dm~3.
0, 2.0 ] 10~5, 4.0 ] 10~5, 6.0 ] 10~5, 8.0 ] 10~5, 1.0 ] 10~4,
1.6 ] 10~4, 2.0 ] 10~4, 3.0 ] 10~4, 4.0 ] 10~4, 4.6 ] 10~4,
5.0 ] 10~4, 6.0 ] 10~4, 7.2 ] 10~4, 7.6 ] 10~4, 8.0 ] 10~4,
9.2 ] 10~4, 9.6 ] 10~4, 1.00 ] 10~3 and 1.20 ] 103 mol dm~3.
Ñuorescence data a formation constant K \ 1830 ^ 10 dm3
mol~1 is obtained. The Ñuorescence maxima for [Al(2@)]`
occurs at 429 nm. The increased Ñuorescence of coordinated 2@
probably reÑects a combination of the increased rigidity intro-
duced into its structure through coordination with the conse-
quent loss of Ñuorescence quenching of vibrational and
rotational modes, and Ñuorophore deprotonation and coordi-
nation, which increases aromatic character and the probabil-
experiences two dominant environments in the complexation
equilibrium and K \ 7080 ^ 110 dm3 mol~1 for [Al(4@)]2`
was obtained from absorbance data collected at 1 nm inter-
vals over the range 340È380 nm. The spectrum of 3 shows a
shoulder at 255 (11 000) and a maximum at 310 nm (4460 dm3
mol~1 cm~1). In [Al(3@)]2` the spectrum of 3@ shows a shoul-
der at 270 (7300) and broad maxima at 320 (3570) and 345 nm
(2970 dm3 mol~1 cm~1). Isosbestic points appear at 290 and
322 nm in the spectra of 1.20 ] 10~4 mol dm~3 solutions of 3
ity of nÈp electronic transitions.
A similar explanation
probably applies for the increased Ñuorescence of coordinated
3@ and 4@ discussed below.
containing increasing [Al3`]
over the range 0È1.20 ] 10~3
total
mol dm~3, consistent with the chromophore experiencing two
dominant environments in the complexation equilibrium, and
K \ 5570 ^ 120 dm3 mol~1 was determined for [Al(3@)]2`
from absorbance data collected at 1 nm intervals over the
range 330È370 nm. Because the solutions of 3 and 4 studied
Ligands 3 and 4. The UV-visible spectra of both 3 and 4
vary systematically with [Al3`]
as exempliÐed by 4 in Fig.
total
3. The spectrum of free 4 shows a shoulder at 254 (8300) and a
maximum at 323 nm (3920 dm3 mol~1 cm~1). The spectrum
of [Al(4@)]2` shows a shoulder at 270 (5490), a broad shoulder
at 280È300 (2350) and a broad maximum at 335È360 nm
(3200 dm3 mol~1 cm~1) when coordinated by Al3`. The
isosbestic points at 290 and 335 nm show the chromophore
were not bu†ered a small decrease in pH occurs as [Al3`]
total
increases. This was not incorporated into either of the above
stability constant determinations or those discussed below.
The isosbestic point at 335 nm was chosen as the excitation
wavelength for the Ñuorescence study of the coordination of 4
by Al3`. Free 4 has a very low Ñuorescence but coordination
by Al3` induces substantial Ñuorescence in 4@ as seen in Fig. 4.
The broad Ñuorescence band shows a systematic increase in
intensity with increase in [Al3`] , and these systematic
total
changes in Ñuorescence collected at 1 nm intervals over the
range 410È450 nm yielded K \ 3890 ^ 30 dm3 mol~1 for
[Al(4@)]2`. The Ñuorescence maximum for [Al(4@)]2` occurs at
430 nm. The isosbestic point at 322 nm was chosen as the
excitation wavelength for the Ñuorescence study of the coordi-
nation of 3 by Al3`. Free 3 also has a very low Ñuorescence
but coordination of 3@ by Al3` induces substantial Ñuores-
cence. The Ñuorescence band of [Al(3@)]2` is broad with a
maximum at 420 nm. The systematic increase in Ñuorescence
intensity observed for 1.20 ] 10~5 mol dm~3 solutions of 3 in
which [Al3`]
increased over the range 0È1.20 ] 10~3 mol
total
dm~3, gave K \ 3190 ^ 40 dm3 mol~1 from Ñuorescence
data collected at 1 nm intervals over the range 400È440 nm.
Fig. 2 Variation of the Ñuorescence of 3.80 ] 10~7 mol dm~3 2 with
added Al(ClO ) in 75% methanol and 25% water by volume solu-
4 3
tion, 0.10 mol dm~3 in NaClO and 0.001 mol dm~3 in HClO at
4
4
298.2 K. The excitation wavelength was 328 nm. The Ñuorescence
increases as [Al(ClO ) ] increases in the sequence: 0, 2.0 ] 10~5,
Ligands 5 and 6. The UVÈvisible spectra of both 5 and 6
4 3
4.0 ] 10~5, 6.0 ] 10~5, 8.0 ] 10~5, 1.0 ] 10~4, 1.6 ] 10~4,
exhibit signiÐcant changes with [Al3`]
as exempliÐed by 5
2.0 ] 10~4, 3.0 ] 10~4, 4.0 ] 10~4, 4.6 ] 10~4, 5.0 ] 10~4,
6.0 ] 10~4, 7.2 ] 10~4, 7.6 ] 10~4, 8.0 ] 10~4 9.2 ] 10~4,
9.6 ] 10~4, 1.00 ] 10~3 and 1.20 ] 10~3 mol dm~3.
total
in Fig. 5. The free ligand shows maxima at 280 (11 100) and
350 (4250), and 5@ exhibits maxima at 280 (12 900) and 378 nm
New J. Chem., 2000, 24, 541È546
543

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attributable to the formation of twisted intramolecular charge
transfer (TICT) states by these ligands.19h21 The nitro-
aromatic units of 5 and 6 undergo intramolecular charge
transfer in the excited state with the aromatic ring and nitro
group acting as the electron donor and acceptor, respectively.
In the ground state and the initial excited state the aromatic
and nitro groups are coplanar and achieve maximum conjuga-
tion. However, a TICT excited state may form from the initial
excited state where the planes of the aromatic ring and the
nitro group are twisted at 90¡ to each other, conjugation
between the two entities is lost and complete transfer of an
electronic charge between them results. Sometimes emission
may be observed from both the initial and TICT excited
states. This is observed for neither 5 nor 6 in the free state and
for neither 5@ nor 6@ in the coordinated state, consistent with
the occurrence of non-emissive decay to the ground state
through energy dissipation into vibrational modes.
Fig. 4 Variation of the Ñuorescence of 1.20 ] 10~5 mol dm~3 4 with
added Al(ClO ) in 73.2% methanol, 24.4% water and 2.4% DMF by
4 3
volume solution, 0.10 mol dm~3 in NaClO at 298.2 K. The excita-
4
tion wavelength was 335 nm. The Ñuorescence increases as
Coordination of ligands 2–6 by other metal ions
[Al(ClO ) ] increases in the sequence: 0, 4.0 ] 10~5, 8.0 ] 10~5,
4 3
1.2 ] 10~4, 1.6 ] 10~4, 2.0 ] 10~4, 2.4 ] 10~4, 2.8 ] 10~4,
Under the same acidic conditions as for the study of Al3`
coordination of 2 described above, the spectrum of 2 showed
very small increases in absorbance over the range 250È370 nm
in the presence of 1.2 ] 10~3 mol dm~3 Zn2`, and moderate
absorbance increases in the presence of 1.2 ] 10~3 mol dm~3
Cd2`. However, these changes were very small compared to
those induced by Al3`, consistent with a weaker coordination
3.2 ] 10~4, 3.6 ] 10~4, 4.0 ] 10~4, 4.4 ] 10~4, 4.8 ] 10~4,
5.2 ] 10~4 and 5.6 ] 10~4 mol dm~3.
(3940 dm3 mol~1 cm~1) when it is coordinated to Al3`. The
isosbestic points at 317 and 364 nm show the chromophore
experiences two dominant environments in the complexation
equilibrium and K \ 11 960 ^ 430 dm3 mol~1 was derived for
[Al(5@)]2` from absorbance data collected at 1 nm intervals
over the range 370È410 nm. In the free state 6 shows a
maximum at 318 (9400), which changes to 337 (13 300) and a
shoulder appears at 375È400 nm (3500 dm3 mol~1 cm~1)
when 6@ is coordinated by Al3`. The isosbestic points at 265,
316 and 380 nm show the chromophore experiences two dom-
inant environments in the complexation equilibrium and
K \ 44 800 ^ 2000 dm3 mol~1 for [Al(6@)]2` derived from
absorbance data collected at 1 nm intervals over the range
320È360 nm for 1.20 ] 10~4 mol dm~3 solutions of 6 in
of these two ions. In the absence of added HClO , but other-
4
wise under the same conditions, Zn2` induced slightly larger
changes in the spectrum of 2, and the spectral changes
induced by Cd2` also increased, but still Al3` induced much
greater changes in the spectrum of 2. (The pH of the latter
solutions were 4.3, 5.4 and 5.0 for Al3`, Zn2` and Cd2`,
respectively, consistent with decreasing protolysis of coordi-
nated water as the surface charge density of the metal ion
decreases.) Under neither set of conditions did the spectrum of
2 show signiÐcant change in the presence of the biologically
important K`, Mg2` and Ca2` ions. Overall, these data are
qualitatively consistent with 2 in its dianionic form, 2@,
showing signiÐcant selectivity for complexing Al3`.
which [Al3`]
dm~3.
increased over the range 0È1.20 ] 10~3 mol
total
Under the same conditions as for the Al3` studies, the
spectra of 3È6 showed very small increases in absorbance over
the range 250È370 nm in the presence of 1.2 ] 10~3 mol
dm~3 Zn2`, and slightly larger absorbance increases in the
presence of 1.2 ] 10~3 mol dm~3 Cd2` and Pb2`. Compared
with the changes induced by Al3`, these changes were very
small, consistent with a stronger coordination of Al3`.
Neither K`, Mg2` nor Ca2` induced signiÐcant spectral
changes. These data are qualitatively consistent with 3È6
showing signiÐcant selectivity for complexing Al3`.
Neither 5 nor 6 are Ñuorescent in the free state and the
Al3` complexes of 5@ and 6@ show no Ñuorescence. This is
For a ligand to be useful as a Ñuorophore in the detection
of a given metal ion it has to possess substantial selective
complexing character for that metal ion. Given the close
relationship of 2È6 to pyochelin, 1, it is probable that these
ligands will strongly complex Fe3`. However, because the dÈd
electronic transitions of this d5 ion quench ligand Ñuorescence
the coordination of 2È6 by Fe3` was not studied. For the
same reason other transition metal ions with d1Èd9 electronic
conÐgurations were not examined. However, the data report-
ed here indicates that biologically ubiquitous Zn2` could
interfere with the detection of intracellular Al3` and accord-
ingly we propose to modify our Ñuorophores to enhance their
selectivity for Al3` for such use.
Fig. 5 Variation of the UVÈvisible absorption spectrum of
1.21 ] 10~4 mol dm~3 5 with added Al(ClO ) in 73.2% methanol,
4 3
24.4% water and 2.4% DMF by volume solution, 0.10 mol dm~3 in
NaClO at 298.2 K. The absorbance increase at 270 and 360 and
4
Experimental
decreases at 320 nm as [Al(ClO ) ] increases in the sequence: 0,
4 3
3.0 ] 10~5, 6.0 ] 10~5, 9.0 ] 10~5, 1.2 ] 10~4, 1.6 ] 10~4,
Potentiometric titrations were carried out using an Orion
Ross SureÑow 81-72 BN combination electrode, an Orion SA
720 potentiometer and a Metrohm E665 Dosimat autoburette
2.0 ] 10~4, 3.0 ] 10~4, 4.0 ] 10~4, 5.0 ] 10~4, 6.0 ] 10~4,
7.0 ] 10~4, 8.0 ] 10~4, 9.0 ] 10~4, 1.00 ] 10~3 and 1.20 ] 10~3 mol
dm~3.
544
New J. Chem., 2000, 24, 541È546

View Article Online
interfaced to an XT/4-8086 computer. The reference com-
partment of the combination electrode was Ðlled with 0.10
3.40, t, J 8.4 Hz, SCH . 13C NMR (CDCl ) d: 171.55, C2N;
2
3
158.32, C1; 135.55, 132.84, C5, C3; 119.03, C6; 117.90, 110.19,
C2, C4; 63.36, CH N; 32.06, CH S.
mol dm~3 NaClO in 75% methanol and 25% water (v : v)
4
2
2
solvent and allowed to attain equilibrium over 2 days before
use. The pK s of 2 and 3È6 were determined from triplicate
2-(4,5-Dihydro-1,3-thiazol-2-yl)-5-nitrophenol, 5. A solution
of 2-hydroxy-4-nitrobenzonitrile (2.26 g, 0.013 mol), 2-
aminoethanethiol hydrochloride (1.90 g, 0.016 mol) and tri-
ethylamine (2.71 g, 0.026 mol) in ethanol (50 cm3) was
degassed and placed under a nitrogen atmosphere. The solu-
tion was reÑuxed for 50 min and cooled to room temperature,
forming crystals of 5, which were collected by vacuum Ðl-
tration and washed with ethanol (1.95 g, 63%). MP 150È
151 ¡C. (Found: C, 48.22; H, 3.37; N, 12.52; S, 14.45%.
C H N O S requires C, 48.21; H, 3.60; N, 12.49; S, 14.30%)
a
titrations of 10 cm3 of 1.00 ] 10~3 mol dm~3 ligand in 75%
methanol and 25% water by volume and 73.2% methanol,
24.4% water and 2.4% DMF by volume solutions, respec-
tively, thermostatted at 298.2 K in a titration vessel against
0.10 mol dm~3 NaOH in 75% methanol and 25% water by
volume solution delivered from a microburette. A Ðne stream
of nitrogen, which had been bubbled through successive traps
containing 0.10 mol dm~3 sodium hydroxide and 75% meth-
anol and 25% water by volume solvent, respectively, was
bubbled continuously through the magnetically stirred titra-
9
8 2 3
IR: 2724 (OH), 1614 (C2C), 1573 (C2C), 1340 cm~1 (CO). MS
tion solution to remove carbon dioxide. Ligand pK s were
m/z: 224 (M`). 1H NMR (CDCl ) d: 13.13, s, OH; 7.83, d, J
a
3
derived from the titration data using the programme
2.2 Hz, H6; 7.71, dd, J 2.2, 8.6 Hz, H4; 7.55, d, J 8.6 Hz, H3;
SUPERQUAD22 on a Digital Venturis 575 computer.
4.59, t, J 8.6 Hz, NCH ; 3.56, t, J 8.4 Hz, SCH . 13C NMR
2
2
UVÈvisible spectra were measured from 250 to 450 nm at 1
nm intervals at a scan rate of 1 nm s~1 and a spectral band-
width of 1.0 nm with a Varian Cary 2200 spectrophotometer
interfaced to an IBM-compatible 486DX personal computer.
Fluorescence spectra were measured from 350 to 600 nm at
0.5 nm intervals at a scan rate of 4 nm s~1 with a Perkin
Elmer LS50B Ñuorimeter interfaced to a Digital 75 MHz
Pentium computer. The entrance and exit slit widths were
adjusted in the range 0.4 to 0.6 nm for each ligand according
to the intensity of emitted light. Solutions were thermostatted
at 298.2 K in 1 ] 1 cm quartz cells for both sets of measure-
ments. Stability constants were determined by Ðtting algo-
rithms for 1 : 1, 1 : 1 and 1 : 2 and 1 : 1 and 2 : 1
complexations (Al3` : ligand) to the absorbance and Ñuores-
cence data using an inhouse programme, which also generated
the coordinated ligand spectra. The 1 : 1 algorithm yielded the
better data Ðts for each system.
(CDCl ) d: 171.86, C2N; 159.87, C1; 150.24, C4; 131.48, C3;
3
121.01, C2; 113.26, 112.52, C6, C4; 63.46, NCH ; 32.11,
2
SCH .
2
2-(4,5-Dihydro-1,3-thiazol-2-yl)-4-nitrophenol, 6. A solution
of 2-hydroxy-5-nitrobenzonitrile (987 mg, 6.02 mmol), 2-
aminoethanethiol hydrochloride (845 mg, 7.44 mmol) and tri-
ethylamine (1.2 g, 11.77 mmol) in ethanol (25 cm3) was
degassed and placed under a nitrogen atmosphere. The solu-
tion was reÑuxed for 2 h after which a precipitate began to
form. The reaction mixture was cooled to room temperature
and yellow needle crystals of 6 were collected by vacuum Ðl-
tration and washed with ethanol (820 mg, 61%). MP 163È
166 ¡C. (Found: C, 48.06; H, 3.36; N, 12.52; S, 14.48%.
C H N O S requires C, 48.21; H, 3.60; N, 12.49; S, 14.30%)
9
8 2 3
IR: 3000 (OH), 1594 (C2C), 1334 cm~1 (CO). MS m/z: 224
(M`). 1H NMR (CDCl ) d: 13.80, s, OH; 8.33, d, J 2.8 Hz,
H3; 8.24, dd, J 3.0, 9.0 Hz, H5; 7.07, d, J 9.4 Hz, H6; 4.53, t, J
Uncorrected melting points were determined using a KoÑer
hot-stage apparatus attached to a Reichert microscope. Infra-
red (IR) spectra were recorded in nujol mulls on either a
Hitachi 270-30 grating spectrophotometer or an ATI Mattson
Genesis FT spectrophotometer. Thin layer chromatography
(TLC) was carried out on Kieselgel 60 F254 (Merck) on alu-
minium backed plates. NMR spectra were recorded on a
Varian Gemini 200 spectrophotometer operating at 199.8
MHz (1H) and 50.4 MHz (13C). Mass spectra were recorded
using a VG ZAB 2HF spectrometer operating at 70 eV.
3
8.2 Hz, NCH ; 3.48, t, J 8.2 Hz, SCH . 13C NMR (CDCl ) d:
2
2
3
172.20, CN; 164.76, C1; 139.64, C4; 128.19, 126.98, C5, C3;
117.98, C6; 115.78, C2; 63.17, NCH ; 32.24, SCH .
2
2
Acknowledgements
Funding by the Australian Research Council is gratefully
acknowledged.
Syntheses
N,N-Dimethylformamide (Aldrich) was distilled under
reduced pressure and stored under dry nitrogen. Water was
puriÐed using the Milli-Q reagent system. Bulk methanol was
puriÐed by distillation prior to use. The reagents used in
ligand preparations were either of good commercial quality or
were prepared by standard methods. Ligands 2 and 3 were
prepared by literature methods and were characterised by IR
spectroscopy, NMR spectroscopy and molecular mass.17,18
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545

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