Proton transfer and supramolecular complex formation between Nile Blue and tetraundecylcalix[4]resorcinarene - A fluorescence spectroscopic study

Article abstract of DOI:10.1039/b202637f

The interactions between a calixarene host comprising dissociable protons, and an organic dye guest existing in a neutral base and in a protonated form, have been studied on model systems obtained by dissolving Nile Blue base (1) and tetraundecylcalix[4]r

Full text of DOI:10.1039/b202637f

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Proton transfer and supramolecular complex formation between
Nile Blue and tetraundecylcalix[4]resorcinarene—a fluorescence
spectroscopic study
2
Miklós Kubinyi,*a,b János Brátán,a András Grofcsik,a,b László Biczók,b Benedek Poór,b
István Bitter,c Alajos Grün,c Botond Bogátid and Klára Tóthd
a Department of Physical Chemistry, Budapest University of Technology and Economics,
1521 Budapest, Hungary. E-mail: kubinyi@mail.bme.hu; Fax: (ϩ36)-1-463-3767;
Tel: (ϩ36)-1-463-2137
b Chemical Research Center, Hungarian Academy of Sciences, 1525 Budapest, P. O. B. 17,
Hungary
c Department of Organic Chemical Technology, Budapest University of Technology and
Economics, 1521 Budapest, Hungary
d Institute of General and Analytical Chemistry, Budapest University of Technology and
Economics, 1521 Budapest, Hungary
Received (in Cambridge, UK) 15th March 2002, Accepted 31st July 2002
First published as an Advance Article on the web 28th August 2002
The interactions between a calixarene host comprising dissociable protons, and an organic dye guest existing in a
neutral base and in a protonated form, have been studied on model systems obtained by dissolving Nile Blue base
(1) and tetraundecylcalix[4]resorcinarene (2) in two apolar solvents, dichloromethane and toluene. Steady state
absorption and fluorescence spectroscopy and fluorescence lifetime measurements were applied as experimental
methods. It has been established that the equilibrium composition of these systems is controlled predominantly by
three reactions: (1) formation of the weakly bound complex 1ؒ2; (2) simultaneous formation of the tightly bound
ionic species with zero net charge 1؉ؒ2؊ (1؉ = protonated form of 1, 2؊ = monophenolate form of 2); (3) protonation
of the latter product in a subsequent step yielding 1؉ؒ2. The equilibrium constants for these reactions have been
determined from the absorption spectra by an iterative procedure. The high value of the equilibrium constant for
reaction (2) in toluene (9.8 × 106) is in accordance with the strongly solvatophobic nature of cation 1؉ in this solvent.
The fluorescence lifetime of neutral base 1 increases whereas that of its protonated form, 1؉, markedly decreases
upon complexation. These effects have been interpreted in terms of intra- and intermolecular decay channels.
calix[6]arene, calix[8]arene and calix[4]resorcinarene) deriv-
Introduction
atives were 2-anilinonaphthalene,5 8-anilinonaphthalene-1-
sulfonate anion,6,8 pyrene,7–9 pyrene-modified N-alkylpyr-
idinium cations,10,11 2-(2Ј-hydroxyphenyl)-4-methyloxazole12
and the cationic dyes Auramine O,13 Brilliant Cresyl Blue14 and
Acridine Red.15
The strategic goal of calixarene chemistry is the synthesis of
novel supramolecular receptors capable of selective recognition
of charged and neutral species.1 The calixarene core in these
hosts can play two different roles. In the majority of these
receptors the macrocyclic skeleton is merely a lipophilic matrix
of appropriate conformation holding the properly designed
coordination sphere where the ionic guest can be bound
through non-covalent interactions.2 (Some exceptions are
known where cation–π interactions between the guest—a soft
metal ion3 or an organic ammonium ion4—and the calixarene
core take part in the binding process.) The situation is quite
different with organic guests, which can penetrate into the calix-
arene cavity, forming a great variety of inclusion complexes.
Of these, the calixarene complexes with ‘fluorescent probes’—
aromatic hydrocarbons and fluorescent dyes—have received
special attention, the properties of which can be very efficiently
studied in dilute solutions by combining UV–visible absorption
and fluorescence spectroscopic methods.
The first papers in this field were published by Shinkai and
co-workers,5–8 who demonstrated that such experiments provide
invaluable information on the nature of the host–guest inter-
action and on the ring-size selectivity of the host in these sys-
tems. Following these studies there has been a growing interest
in this research area, as an important part of the investigation
of various types of nanocavities. The fluorescent guests studied
in supramolecular structures with calixarene (calix[4]arene,
A great number of dyes, including some of the fluorescent
dyes mentioned above, have a basic and at least one (singly)
protonated form. As has been pointed out by Shinkai et al.,16
the interaction between these indicator dyes and calixarene
based supramolecular receptors in protic solvents can be
described by a complicated reaction scheme, in which the pro-
ton transfer from the solute to the free and complexed forms of
the basic guest is taken into account. The scheme was applied to
the analyses of systems formed by p-sulfonated calixarene hosts
and indicator dye guests, the latter in neutral and in mono-
cationic form (Phenol Blue, Anthrol Blue)16 or in monocationic
and in dicationic form (4-[4-(dimethylamino)styryl]-1-methyl-
pyridinium iodide),17 in aqueous solutions.
Besides the proton transfer between such dye guests and
solvent molecules, an additional type of proton transfer may
take place between the guest and the host, if the host comprises
dissociable protons. In the present work we studied the role of
the latter process in a model system. We have chosen Nile Blue
as a dye guest existing in a basic and in a protonated form (1
and 1؉ in Scheme 1, cation 1؉ is often referred to as “Nile Blue
A”) and tetraundecylcalix[4]resorcinarene (2 in Scheme 2, its
monophenolate form will be denoted by 2؊) as a host with
1784
J. Chem. Soc., Perkin Trans. 2, 2002, 1784–1789
This journal is © The Royal Society of Chemistry 2002
DOI: 10.1039/b202637f

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Table 1 Absorption and fluorescence spectral data of Nile Blue A (1؉) perchlorate and of Nile Blue base (1)–tetraundecylcalix[4]arene (2) mixtures
Solute composition/M
Absorption spectrum
Fluorescence spectrum
λmax/nm (relative fluorescence intensity)
Solvent
[1؉]
[1]0
[2]0
λmax/nm (ε/MϪ1 cmϪ1
)
Dichloromethane
λex = 505 nm
λex = 585 nm
2 × 10Ϫ6
647 (97 400)
505 (45 200)
649 (96 500)
660 (1160)
2 × 10Ϫ6
2 × 10Ϫ6
579 (886)
595 (36)
λex = 490 nm
4 × 10Ϫ5
4 × 10Ϫ5
666 (103)
λex = 585 nm
645 (43)
Toluene
Satd. soln.a
627 (—)
490 (36 800)
647 (55 900)
2 × 10Ϫ6
2 × 10Ϫ6
544 (271)
567 (9)
658 (20)
a Estimated concn. ∼ 1.2 × 10Ϫ7 M.
solvents (water, alcohols), we obtained equilibrium mixtures of
1 and its protonated form, 1؉, before adding any 2 to the solu-
tions. In these solvents, as well as in aprotic, polar solvents (e.g.
acetonitrile), upon the addition of 2 no change in the spectrum
was observed. In aliphatic, aromatic and chlorinated hydro-
carbons the mixing resulted in substantial changes in the
spectrum. Of the latter systems, toluene and dichloromethane
solutions were studied in detail. The absorption and fluor-
escence spectra of pure 1 and of pure 1؉-perchlorate were
taken, then the spectra of mixtures with the same initial con-
centration of 1 (2 × 10Ϫ6 M) and various initial concentrations
of 2 (from 5 × 10Ϫ7 to 4 × 10Ϫ5 M) were recorded in the two
solvents (1 M = 1 mol dmϪ3).
Scheme 1
The data of the absorption spectra and fluorescence spectra
of the solutions of pure 1, of pure 1؉-perchlorate, and of the
mixtures with the highest 2 excess are summarized in Table 1.
The variations of absorption and fluorescence spectra with the
amount of added 2 are displayed in Figs. 1 and 2, respectively.
Scheme 2
dissociable protons. Nile Blue is a versatile oxazine dye with
many applications in analytical chemistry, photoscience,
materials science and molecular biology. The proton addition
of neutral base 1 and the reverse process in various types of
solvents were the subject of a thorough spectroscopic study by
Douhal.18 The characteristics of the S0
S1 transition and of
the following deactivation of cation 1؉ in polar solvents are
known in detail from our former time-resolved spectroscopic
measurements19–21 and quantum chemical calculations.22
Calixresorcinarene 2 forms stable hydrogen-bonded complexes
with strongly polar guests like sugars and these complexes are
soluble in apolar solvents.23
In this work, in addition to steady state UV–visible absorp-
tion and fluorescence spectroscopy, fluorescence lifetime
measurement has also been applied as an experimental method.
The recently published results of the first fluorescence decay
experiments, carried out on an organic dye guest–calixarene
host system, prove that these measurements provide important
information on the nature of host–guest interactions.12
Fig.
1
Absorption spectra of Nile Blue base (1)–tetraundecyl-
calix[4]resorcinarene (2) mixtures (a) in dichloromethane and (b) in
toluene. Initial concentrations: [1]0 = 2 × 10Ϫ6 M in each solution,
[2]0 = (A) 0, (B) 1 × 10Ϫ6, (C) 2 × 10Ϫ6, (D) 3 × 10Ϫ6, (E) 4 × 10Ϫ6
,
(F) 6 × 10Ϫ6, (G) 8 × 10Ϫ6, (H) 1.2 × 10Ϫ5, (I) 1.6 × 10Ϫ5, (J) 4 × 10Ϫ5 M.
These spectra indicate that Nile Blue exists in these systems in
at least five different forms: (i) unreacted base 1, (ii) a supra-
molecular complex consisting of a neutral host and a neutral
guest, 1ؒ2, (iii) a supramolecular complex of ionic nature,
1؉ؒ2؊, (iv) complexed cation 1؉ؒ2 and (v) uncomplexed cation
1؉. The 1 : 1 stoichiometry of 1؉ؒ2؊ has been directly proved
with the help of the ‘continuous variations’ method. The com-
positions of the other two complexes could not be determined
Results and discussion
Steady state spectra
Solutions of 1 and 2 prepared with various solvents were mixed
and the interactions between the two solutes were monitored
by comparing the UV–visible absorption spectra of the pure
components to those of the mixtures. On dissolving 1 in protic
J. Chem. Soc., Perkin Trans. 2, 2002, 1784–1789
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The presence of 1؉ؒ2؊, a supramolecular structure of ionic
nature, is indicated by the appearance of two overlapping bands
with maxima around 585 and 645 nm in the absorption spectra
in Fig. 1. The intensities show that in both solvents the concen-
tration of this species rapidly increases with the amount of the
added host up to ∼ [2]0 = 4 × 10Ϫ6 M, the addition of further 2 to
the systems leads to the conversion of this product into 1؉ؒ2.
The range of the visible absorption of 1؉ؒ2؊ clearly shows the
cationic character of its Nile Blue moiety, and through this
the ionic nature of the whole supramolecular structure. 1؉-
perchlorate dissolved in polar or amphiphilic solvents has a
single absorption band in the visible range (λmax = 635 nm in
water, 628 nm in ethanol, 627 nm in butan-1-ol, 624 nm in
chloroform).24 In contrast, in 1-chloronaphthalene the absorp-
tion band (λmax = 646 nm) has a shoulder on the lower wave-
length side,25 the relative intensity of which does not change
with the concentration,18 thus, it does not seem to be related to
aggregation. Instead, the splitting may be a consequence of the
imperfect solvation of the oppositely charged ions. Nile Blue
may be present in the form of tight ion pairs in this rather
apolar solvent, in other words, the electrostatic interaction
between cation 1؉ and the perchlorate anion is expected to be
stronger in 1-chloronaphthalene than in more polar solvents.
The electrostatic forces may be even stronger between the 1؉
and 2؊ moieties of the ionic complex in the systems studied,
where the solvent molecules are unable to weaken the electro-
static part of the host–guest interactions, which may be the
reason for the splitting observed in the spectra in Fig. 1.
As can be seen in Fig. 2d, the fluorescence of the supra-
molecular systems in toluene is very weak when they are irradi-
ated at 585 nm, where the free and the two complexed forms
of cation 1؉ absorb strongly. This confirms that the cations are
present predominantly in the complexed forms, as 1؉ؒ2 and
1؉ؒ2؊. Probably, neither of the complexes exhibits detectable
emission in toluene, the weak fluorescence of the solutions with
high excess of 2 may arise from a small amount of free cations.
On the other hand, dichloromethane solutions show fluor-
escence, when irradiated at the above wavelength. The
fluorescence intensity increases with the amount of 2 in the
systems, but even in cases of high excess of 2, where the absorp-
tion spectra indicate that almost the whole amount of base 1 is
converted to protonated species, it remains weaker by one order
of magnitude than that of pure 1؉-salts. This suggests that
there is only a small amount of strongly fluorescing free 1؉
cations in these systems, and—in contrast to the toluene
solutions—the complexation of the cations in the forms of
1؉ؒ2؊ and 1؉ؒ2 results in a dramatic reduction of fluorescence,
but not in complete quenching.
Fig. 2 Fluorescence spectra of 1–2 mixtures (a) in dichloromethane,
excitation wavelength (λex) = 505 nm; (b) in dichloromethane, λex = 585
nm; (c) in toluene, λex = 490 nm; (d) in toluene, λex = 585 nm. Initial
concentrations as in Fig. 1.
with simple standard methods, because their formation can be
observed only in the presence of the host in high excess. The
presence of each complex in the solutions, and their stoichio-
metries, however, have been established by calculating the equi-
librium constants of the occurring reactions from the absorp-
tion spectra, by the steady-state fluorescence spectra and by the
fluorescence lifetime measurements, the details of which will be
given later in this paper.
The position of the absorption band belonging to the Nile
Blue base is not sensitive to the addition of 2 to the solutions.
The formation of 1ؒ2 is indicated primarily by the fluorescence
measurements. The fluorescence band of the base loses inten-
sity and shifts gradually to higher wavelengths with increasing 2
content. In the case of CH2Cl2 solutions the maximum of this
band, λmax, is located at 579 2 nm in the spectra of all systems
with initial calixresorcinarene concentrations [2]0 = 0–8 × 10Ϫ6
M, then it is shifted gradually to 595 nm observed at [2]0 =
4 × 10Ϫ5 M, the highest applied excess of the host. In toluene
the corresponding band, appearing at 544 nm in the absence of
2, shows a clear shift to higher wavelengths even at low 2
contents, e.g. it appears at 560 nm in the spectrum of the solu-
tion with [2]0 = 8 × 10Ϫ6 M. As can be seen in Fig. 2c, the shape
of the band also changes in the same interval (the shoulder
gradually disappears). This is followed by a further moderate
shift to 567 nm, as [2]0 is increased to 4 × 10Ϫ5 M. These results
suggest that the adduct 1ؒ2 is more stable in a strongly apolar
medium, like toluene.
A substantial reduction of the fluorescence intensity has been
reported for the majority of supramolecular systems studied so
far, with cationic dye guests and calixarene hosts having
dissociable protons,10,11,14,15 but there are also examples where
such systems exhibit fluorescence enhancement.12,13 (All these
systems were studied in aqueous solutions.)
Reliable absorption and fluorescence spectral data for the
pure protonated dye, 1؉, (in the form of its perchlorate salt)
could be obtained only in dichloromethane. The solubility of
this salt is very low in toluene and the absorption spectra indi-
cate that a part of the dissolved cations is instantly deproton-
ated.
The applied absorption and fluorescence spectroscopic
methods do not provide much information about the inclusion
type nature of the complexes formed by 1 and 2. The inclusion
of dye molecules by calixarene receptors can be studied effi-
1
ciently by measuring the shifts in the H NMR spectra of the
The absorption bands arising from free 1؉ cation, and from
its complexed forms, 1؉ؒ2 and 1؉ؒ2؊, strongly overlap, thus,
they cannot be easily separated in the spectra.
dye moieties upon the addition of calixarenes, as was shown
in two papers by Shinkai et al.16,17 The solubility of their
hosts, guests and complexes in D2O was high enough for such
measurements. We made an effort to carry out similar experi-
ments by measuring the 1H NMR spectra of 1, of 1؉ (perchlor-
ate salt), of 2 and of 1–2 mixtures in two solvents—CDCl3 and
CD2Cl2—in which the UV–VIS spectra indicate complex form-
ation. These experiments clearly indicated the proton transfer
from 2 to 1, but did not give reliable information concerning the
inclusion type structure of the complex. In CDCl3 the solubility
of 1؉-perchlorate is too low for NMR measurements, in CD2Cl2
The absorption feature with a single maximum around 650
nm, which becomes predominant at high 2 concentrations, has
been assigned to 1؉ؒ2. As can be seen in Table 1, the λmax and
εmax values for complex 1؉ؒ2 and for uncomplexed cation 1؉ in
dichloromethane are very close. The corresponding data for the
toluene solutions cannot be directly compared, because of
the aforementioned poor solubility of 1؉-perchlorate in this
solvent.
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J. Chem. Soc., Perkin Trans. 2, 2002, 1784–1789

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the solubility of 2 is too low to prepare mixtures which contain
at least the ‘ionic’ complex 1؉ؒ2؊ in higher concentration.
Reactions and equilibrium constants
In the majority of studies on supramolecular systems the form-
ation of a single supramolecular product is observed, when the
stoichiometry of the complex can be justified, and the associ-
ation constant can be determined with the help of a simple
Benesi–Hildebrand plot of the absorbance values or of the
fluorescence intensities. Since the spectra revealed the existence
of three different supramolecular complexes in our systems, this
route could not be followed. Instead, various schemes with par-
allel and consecutive reactions have been set up and tested by
checking if the values obtained for the equilibrium constants
for the individual reactions are independent of the com-
position. The equilibrium concentrations of the reactants and
of the products have been determined from the absorption
spectra. The spectral data of twelve solutions in each solvent
have been taken into account, with the same total Nile Blue
concentration of 2 × 10Ϫ6 M: solution of pure base 1, ten mix-
tures with various amounts of added calixresorcinarene from
[2]0 = 5 × 10Ϫ7 to 8 × 10Ϫ6 M, and a mixture with [2]0 = 4 × 10Ϫ5
M. The spectrum of the latter system has been taken as the
spectrum of 1؉ؒ2. The concentration of the free 1؉ cations was
taken as zero and the absorption of the free host (in the visible
range) was neglected. By application of Beer’s law for such a
system, the spectrum—the absorbance of a sample in a cell with
light-pass λ—can be expressed as a sum of the contributions
from the components, which are present in their equilibrium
concentrations:
Fig. 3 Resolution of the absorption spectra of a 1–2 mixture in (a)
dichloromethane and (b) toluene. Initial concentrations: [1]0 = 2 × 10Ϫ6
M, [2]0 = 4 × 10Ϫ6 M.
(4)
(5)
(6)
A(λ) = {ε1(λ)ؒ[1] ϩ ε1ؒ2 (λ)ؒ[1ؒ2] ϩ
؉
؊
ε1 (λ)ؒ[1 ؒ2 ] ϩ ε1 ؒ2 (λ)ؒ[1؉ؒ2]}ؒl
؉
؉
ؒ2؊
The visible absorption spectrum of 1ؒ2 was taken to be
identical with that of 1 for the ten systems involved in the
analysis, which seems to be a reasonable assumption at least for
dichloromethane solutions, where the effect of the added 2 is
insignificant even on the fluorescence behavior. The spectrum
of 1؉ؒ2؊, a component available only in equilibrium mixtures,
was obtained at the end of an iterative subtraction of the bands
representing 1 (together with 1ؒ2) and 1؉ؒ2. The result of this
procedure is illustrated in Fig. 3, showing the contributions of
the components to the spectrum of two selected systems: one in
dichloromethane and one in toluene.
Denoting the fractions of 1 converted to 1ؒ2 and to 1؉ؒ2؊ by
α and β, respectively, and the fraction of the latter product
converted to 1؉ؒ2 by γ, the equilibrium concentrations of the
products and the reactants are
[1ؒ2] = [1]0ؒα
(7)
(8)
[1؉ؒ2؊] = [1]0ؒβؒ(1 Ϫ γ)
[1؉ؒ2] = [2Ϫ] = [1]0ؒβؒγ
[1] = [1]0ؒ(1 Ϫ α Ϫ β)
The analysis has led to the conclusion that our systems are
governed by three reactions. The weakly bound and the ionic
complexes are formed in simultaneous processes
(9)
(10)
(11)
1 ϩ 2
1ؒ2
(1)
and
[2] = [2]0 Ϫ [1]0ؒ(α ϩ β ϩ βؒγ)
1 ϩ 2
1؉ؒ2؊
(2)
As a first estimation, the value of α (and of K1) was taken as
zero, and the values for K2 and K3 were calculated from the
concentrations available from the resolved spectra. In the case
of dichloromethane solutions, the values obtained for K2 and
K3 were independent of the composition, which seems to con-
firm that K1 is of low value in this solvent. In contrast, in tolu-
ene the figures obtained for K2 and K3 showed a slight, but
systematic variation with the amount of the added 2. The equi-
librium constants for the latter systems were computed in an
iteration process, in which K2 and K3 were calculated using
values for α for the ten systems corresponding to various
assumed values for K1, as long as K2 and K3 became independ-
ent of the composition of the mixtures.
The ionic complex is then protonated in a subsequent step:
1؉ؒ2؊ ϩ 2 1؉ؒ2 ϩ 2؊
(3)
The experimental spectra do not provide direct information
on reaction (1). As can be seen in Fig. 1, at lower 2 concen-
trations the formation of the ionic complex—reaction (2)—
predominates in dichloromethane and this is the only observ-
able reaction in toluene. As the concentration of 2 is increased,
the ionic complex is gradually protonated and in systems with
large 2 excess 1 is converted almost completely into 1؉ؒ2.
Thermodynamically the systems can be characterized by the
apparent equilibrium constants K1, K2 and K3 belonging to
reactions (1), (2) and (3), respectively
The calculated equilibrium constants are presented in Table
2. As can be seen, the association constants, particularly that
for the formation of 1؉ؒ2؊ in toluene, are significantly higher
J. Chem. Soc., Perkin Trans. 2, 2002, 1784–1789
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Table 2 Equilbrium constants for reactions in Nile Blue base–
tetraundecylcalix[4]resorcinarene mixtures
Values for τF for 1 have not been found in the literature. The
observation that the addition of 2 does not influence the rate of
fluorescence decay in dichloromethane—τF has a constant
Solvent
K1/105 MϪ1
K2/105 MϪ1
K3
value of 1260
20 ps—is in accord with the result of the
steady-state experiments: reaction (1) does not play an import-
ant role in this solvent. In contrast, the lifetimes measured at
545 nm in toluene provide further evidence for the formation of
1ؒ2. The value for τF is 390 ps for pure 1 and 550 ps for complex
1ؒ2, the latter measured at the highest applied excess of 2. The
decay curves of the solutions with lower 2 concentrations have
seemingly also a single-exponential character with lifetimes
between the two above values, in other words, the lifetimes
of the two co-existing species cannot be obtained separately in
the fitting procedure. (Model calculations by Gardini et al.27
demonstrated that the correct values for two relatively close
fluorescence lifetimes may not be obtained from the analysis
of the decay curves arising from the two coexisting species,
instead a single exponential function may give a satisfactory
fit.)
Dichloromethane
Toluene
Chlorobenzene
—
2.0 0.5
—
7.5 1.3
0.012 0.003
0.023 0.005
0.34 0.06
98
8
2.8 0.5
than the values (with 102–104 order of magnitude) typical of
organic guest–calixarene systems.6,8,11,13–15,17,26 Equilibrium con-
stants in the range 105–107 were obtained for the complexation
of pyrene by water soluble calixarenes in aqueous solutions.6,8 It
seems plausible that cation 1؉, solvatophobic in apolar solvents
(like pyrene is solvatophobic in water), forms a very stable
complex with anion 2؊. The solubility of this complex in apolar
media is due partly to the zero net charge, partly to the isolation
of the oppositely charged functional groups by the aromatic
rings and the long aliphatic chains.
In order to study the behavior of the system in a saturated
and in a chlorinated aromatic hydrocarbon, analogous experi-
ments have been carried out in n-heptane and in chlorobenzene.
In the former solvent, in which the solubility of 2 is poor, the
simultaneous formation of the “neutral” (1ؒ2) and the “ionic”
(1؉ؒ2؊) complexes could be observed in slow heterogeneous
reactions by measuring the absorption and fluorescence spectra
repeatedly. In chlorobenzene the system behaved in a similar
manner as in dichloromethane. The equilibrium constants for
reactions (2) and (3) have been evaluated, their values are also
presented in Table 2.
Fluorescence lifetimes for cation 1؉ in various sets of sol-
vents were reported in Refs. 19,20,28–30. Its lifetime decreases
with increasing solvent polarity, from ∼3100 ps measured in
chloroform to ∼400 ps measured in water. Our result for the
dichloromethane solution practically coincides with the τF
value of 2900 ps reported in Refs. 29,30. Lifetimes for 1؉-salts
dissolved in an apolar solvent such as toluene have not been
found in the literature, the value in Table 3 fits the trend
observed for decreasing solvent polarity.
The fluorescence of Nile Blue in the presence of 2 in toluene
was very weak when the 632 nm laser was applied for excitation,
which did not allow reliable measurement of the fluorescence
lifetimes of the cationic species in this solvent. In accordance
with the steady-state spectra in Fig. 2, much stronger fluor-
escence was observed when the dichloromethane solutions were
excited with this laser, which allowed recording the above men-
tioned decay curves with double exponential character. As can
be seen in Table 3, the value of τF belonging to the dominant
component decreases with increasing concentrations of the
calixresorcinarene up to [2]0 = 8 × 10Ϫ6 M, and it remains
unchanged at higher 2 concentrations. Its lowest value of
∼2100 ps can be assigned to 1؉ؒ2. The longer decays observed
at lower 2 contents may be associated with the presence of free
Fluorescence lifetimes
In order to separate the fluorescence emissions of the pure and/
or complexed forms of base 1 from those of pure and/or com-
plexed forms of cation 1؉, the exciting pulses were produced by
two semiconductor lasers, operating at 404 nm (in the absorp-
tion range of the former species) and at 632 nm (in the absorp-
tion region of the latter species); the emission monochromator
was set to wavelengths 545 and 660 nm, close to the fluor-
escence maxima of neutral and cationic forms of Nile Blue,
respectively. Most of the decay curves show a single exponential
character. The exceptions are those obtained by exciting the
cationic species in dichloromethane, which have been found
biexponential, with two strongly different amplitudes. The
(dominant) fluorescence lifetimes (τF) obtained with the two
different excitation and observation wavelengths are presented
in Table 3.
1
؉ cations in low concentrations. The relative amplitude of the
minor component increases with the concentration of 2 up to
[2]0 = 4 × 10Ϫ6 M, and shows an opposite trend above this
value, suggesting that it originates from the ionic complex
1؉ؒ2؊. The amplitude of this component is too small for
Table 3 Fluorescence lifetimes of Nile Blue base (1) and of Nile Blue A cation (1؉) in pure solvents and in Nile Blue base (1)–tetraundecyl-
calix[4]arene (2) mixtures
Solute composition/M
Fluorescence spectrum/psa
1
1؉
λex = 404 nm
λob = 545 nm
λex = 632 nm
λob = 660 nm
Solvent
[1؉]
[1]0
[2]0
Dichloromethane
2 × 10Ϫ6
2910
2 × 10Ϫ6
2 × 10Ϫ6
2 × 10Ϫ6
2 × 10Ϫ6
2 × 10Ϫ6
2 × 10Ϫ6
1260
1260
1240
1270
1280
1270
2 × 10Ϫ6
2690
2430
2110
2110
2090
3210
4 × 10Ϫ6
8 × 10Ϫ6
1.6 × 10Ϫ5
3 × 10Ϫ5
Toluene
∼ 6 × 10Ϫ8
2 × 10Ϫ6
2 × 10Ϫ6
2 × 10Ϫ6
2 × 10Ϫ6
2 × 10Ϫ6
2 × 10Ϫ6
390
380
390
430
470
550
2 × 10Ϫ6
4 × 10Ϫ6
8 × 10Ϫ6
1.6 × 10Ϫ5
3 × 10Ϫ5
a Estimated errors within 5%.
1788
J. Chem. Soc., Perkin Trans. 2, 2002, 1784–1789

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accurate determination of the respective lifetime. Its estimated
value is ∼ 800 ps.
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As pointed out in our former study, the fluorescence decay of
cation 1؉ has an intramolecular and an intermolecular
channel.21 The predominant intramolecular process is probably
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amino group. The intermolecular channel is of significance in
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Conclusion
The combined application of steady-state absorption and fluor-
escence spectroscopy and of fluorescence lifetime measure-
ment, to identify the supramolecular products and to determine
the equilibrium constants for the reactions occurring in this
model system may be helpful in the analysis of many related
systems, where proton transfer between the host and the guest
may take place. In addition, knowledge of the properties of the
Nile Blue–calixresorcinarene complexes in apolar solvents may
be interesting in relation to the applications of this dye in more
complex apolar environments. This is the case when Nile
Blue salts (and in particular, the salts of its lipophilised deriv-
atives) are employed as indicator dyes in optical sensors with
hydrophobic polymer matrices;31 or when they are applied in
photodynamic therapy studies, as special photosensitizers
which localize in the lysosome of tumor cells.32
Experimental
Materials
Nile Blue A perchlorate was purchased from Fluka and used as
received. Its basic form, 1, was prepared by precipitating it from
the aqueous solution of 1؉-perchlorate with NaOH, extracting
with CH2Cl2 and evaporating the solvent. Calixresorcinarene 2
was prepared through the cyclooligomerisation of resorcinol
and dodecanal as described in the literature.33
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Instrumentation
All spectroscopic measurements were carried out at 25 ЊC. The
UV–VIS absorption spectra were recorded using a GBC Cintra
10E spectrometer. The fluorescence spectra were taken by using
a Perkin-Elmer LB50 spectrofluorimeter.
Fluorescence lifetimes were measured with the time-
correlated single-photon counting technique. The 404 and 632
nm exciting pulses were provided by two Picoquant diode
lasers, with pulse duration of 60 and 90 ps, respectively. The
emission was collected at a 90Њ angle through an Oriel 0.25 m
monochromator, located in front of a Hamamatsu H5783
photosensor module. The output signal was connected to a
Picoquant Timeharp 100 computer board module (channel
width 36 ps). Data were analyzed by a non-linear least-squares
deconvolution method.
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Photobiol., 2000, 71, 747; (b) G. Singh, M. Espiritu, X. Y. Shen,
J. G. Hanlon and A. J. Rainbow, Photochem. Photobiol., 2001, 73,
651.
33 L. M. Tunstad, J. A. Tucker, E. Dalcanale, J. Weiser, J. A. Bryant,
J. C. Sherman, R. C. Helgeson, C. B. Knobler and D. J. Cram,
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Acknowledgements
The authors are grateful to the Hungarian Science Research
Foundation (grant numbers T 25561 and T 34990) for financial
support.
J. Chem. Soc., Perkin Trans. 2, 2002, 1784–1789
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