Design, synthesis and photophysical properties of 8-hydroxyquinoline-functionalized tripodal molecular switch as a highly selective sequential pH sensor in aqueous solution

Article abstract of DOI:10.1039/c4ra16345a

The development and photophysical properties of a biomimetic analogue of microbial siderophore from quinolobactin have been described. The putative analogue, 5,5′-(2(((8-hydroxyquinolin-5-yl)methylamino)methyl)2-methylpropane-1,3-diyl)-bis(azanediyl)-bis(methylene)diquinolin-8-ol (TAME5OX), was synthesized with three bidentate 8-hydroxyquinoline (8HQ) groups, connected to a 1,1,1-tris(aminomethyl)ethane framework, and was selected for its potential applications in chemical and biological fields. A combination of absorption and emission spectrophotometry, potentiometry, electrospray mass spectrometry, NMR, IR and theoretical investigation was used to fully characterize TAME5OX. The intense fluorescence from TAME5OX is quenched intermittently under acidic and basic conditions due to the photoinduced intramolecular electron transfer from excited N-pyridyl to hydroxyl moiety of 8HQ units. This renders the ligand an OFF-ON-OFF type of pH-dependent fluorescent sensor. DFT was employed for optimization and evaluation of vibrational modes, excitation and emission properties of the protonated, neutral, deprotonated states of the analogue. Anomalous enhancement observed in the fluorescence spectra of the neutral form of the sensor can be attributed to subtle structural differences from its cationic and anionic forms. Plausible explanations for low fluorescence of the acidic as well as basic form are provided. This journal is

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Design, synthesis and photophysical properties of 8ꢀhydroxyquinolineꢀfunctionalized
tripodal molecular switch as a highly selective sequential pH sensor in aqueous solution
1
2
1
Rifat Akbar , Minati Baral and B K Kanungo *
1
Department of Chemistry, Sant Longowal Institute of Engineering & Technology, Longowal, Punjab−148106,
India
2
Department of Chemistry, National Institute of Technology Kurukshetra, Haryana−136119, India
b.kanungo@gmail.com
Abstract
The development and photophysical properties of biomimitic analogue of microbial
siderophore from quinolobactin have been described. The impudent analogue, 5,5'ꢀ(2(((8ꢀ
hydroxyquinolinꢀ5ꢀyl)methylamino)methyl)2ꢀmethylpropaneꢀ1,3ꢀdiyl)bis(azanediyl)ꢀbisꢀ
(methylene)diquinolinꢀ8ꢀol, (TAME5OX), has been synthesized by three bidentate
8
−hydroxyquinoline (8HQ) groups, connected to 1,1,1−tris(aminomethyle) ethane
framework, was selected for its potential applications in chemical and biological fields. A
combination of absorption and emission spectrophotometry, potentiometry, electrospray mass
spectrometry, nmr, ir and theoretical investigation were used to fully characterize
TAME5OX. The intense fluorescence from TAME5OX get quenched intermittently under
acidic and basic conditions due to the photoinduced intramolecular electron transfer (PET)
from excited N−pyridyl to hydroxyl moiety of 8HQ units. This renders the ligand as
OFF‒ON‒OFF type of pHꢀdependent fluorescent sensor. DFT was employed for
optimization and evaluation of vibrational modes, excitation and emission properties of the
protonated, neutral, deprotonated states of the analogue. Anomalous enhancement observed
in the fluorescence spectra of the neutral form of sensor can be attributed to subtle structural
differences from its cationic and anionic forms. Plausible explanations for low fluorescence
of the acidic as well as basic form, were provided.
1

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Keywords: biomimitic analogue; quinolobactin; microbial siderophore; fluorescence; DFT;
OFF‒ON‒OFF sensor.
*Correspondence: B. K. Kanungo, Department of Chemistry, Sant Longowal Institute of
Engineering & Technology, Longowalꢀ148106, Punjab, INDIA.
Email: b.kanungo@gmail.com; Ph. No. : +91ꢀ1672ꢀ253166 (O), +91ꢀ1672ꢀ253167 (R); FAX:
+
91ꢀ1672ꢀ280057.
1. Introduction
Out of seven potential quinolinols, only 8ꢀhydroxyquinoline (8ꢀHQ) can readily form a fiveꢀ
1
membered chelate complex with metal ions. An important property that makes 8ꢀHQ even
2
more interesting as a chelator is the significant change in its fluorescence behaviour.
Therefore, 8ꢀHQ has been used extensively to construct highly sensitive fluorescent
chemosensors for sensing and imaging of metal ions of important biological and/or
3
ꢀ6
3+
environmental significance, and its chelates, in particular those with Al , are major
7
components for organic lightꢀemitting diodes (OLEDs). Due to their high thermal stability,
8
ꢀ9
excellent electronꢀtransport properties, and unique luminescent properties,
many
derivatives of 8ꢀhydroxyquinoline (for example, substituted at 2, 3,or 7 position in tri or
tetrapodal orchestration, and/or coordinated with different metal ions) (Figure 1) have also
attracted considerable interest. The importance of metalloquinolates in analytical chemistry is
10
also well known. In addition, a large number of 8ꢀhydroxyquinoline derivatives have been
synthesized and shown to be bioactive. For example, it was reported that some 8ꢀ
1
1
hydroxyquinoline derivatives possess antitumor and antimicrobial activities.
It is known that the excitedꢀstate intramolecular proton transfer (ESIPT), from 8ꢀOH
to the quinolinoꢀN atom, makes 8ꢀHQ weakly fluorescent and metal binding to 8ꢀHQ blocks
2
,5
the ESIPT channel, thereby restoring the fluorescence. 8ꢀHQ could hence be employed to
build fluorescent chemosensors with increased fluorescence signal upon metal binding via
2

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Figure 1. Structures of Oxinobactin, Sulfoxinobactin, Tsox, TsoxMe, COX2000 and Oꢀ
TRENSOX.
ESIPT suppression. To enhance metal binding selectivity of 8ꢀHQ, introduction of additional
moieties (central unit and connecting groups) at the Cꢀ2, Cꢀ3 and/or Cꢀ7 positions, adjacent to
3
ꢀ6
the original binding sites in 8ꢀHQ (Figure 1), has been extensively examined. In the case
where 8ꢀHQ was substituted with methyleneamine at the Cꢀ2 or Cꢀ7 position, a photoinduced
4
electron transfer (PET) process leading to fluorescence quenching was identified, and metal
binding resulting in PET suppression was also established, which mechanism is currently
intensively employed in constructing fluorescent chemosensors for transition metal ions
1
2
operating under fluorescence enhancement mode. Moreover, due to unique pyridyl −N and
−
OH structural features of 8−HQ unit, these end groups under acidic and basic conditions are
able to act as a weak acid and a weak base, respectively, associated with the equilibrium
3

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+
∼−OH form and deprotonated quinolinate N∼
O−
between the protonated quinolinium NH
13
form, this moiety appears to be able to act as the pH indicator for both acidic and basic
systems. However, despite the large number of investigations on the fluorescent sensors with
1
4
the 8ꢀHQ moiety as receptor for metal ions, there still exists very less exploration on the
tripodal fluorescent sensors with 8ꢀHQ moiety as pH indicator to the best of our knowledge.
For the intention of preparing versatile fluorescent sensors for the acidic and/or basic
systems, novel fluorescent compounds concurrently possessing three efficient 8ꢀHQ receptor
units should be developed. It is noted, however, that these investigations hitherto reported are
1
5ꢀ18
limited mainly to 8ꢀHQ derivatives with substituents on the Cꢀ2, Cꢀ3 or Cꢀ7 position.
Derivation at Cꢀ5 position of the 8ꢀHQ has not been explored in detail. A possible reason
could be that due to the para position of pyridyl N and OH groups in this arrangement, not
much room remains for encapsulation of metal ions in the central cavity of ligand which
facilitate coordination with more strain. However, further exploration revealed that
substitution at the Cꢀ2, Cꢀ3 and/or Cꢀ7 positions in the 8ꢀHQ moiety mainly tunes the
coordinating property of the pyridyl N group, while substitution at the Cꢀ5 positions induces
13
change in the binding ability of the OH group. A preliminary molecular modeling study
with molecular mechanics also attests the fact. As a consequence, designing and preparing
novel fluorescent sensor in which the central unit is attached at the Cꢀ5 position are of
interest. In the present paper, the novel 8ꢀhydroxyquinoline based tripodal chelator, namely
5
,5'ꢀ(2ꢀ(((8ꢀhydroxyquinolinꢀ5ꢀyl)methylamino)methyl)ꢀ2ꢀmethylpropaneꢀ1,3ꢀ
diyl)bis(azanediyl bis(methylene)diquinolinꢀ8ꢀol, (TAME5OX), (Scheme 1) was
reported, in which design relies on a short tris(aminomethyl)ethane (TAME) backbone as an
anchor symmetrically fitted with three individual 8−hydroxyquinoline (8HQ) moieties at the
C−5 position. The spacer bears a secondary amine function, and its length has been chosen to
achieve a tight orientation of binding units. In continuation to our work on acyclic tripodal
4

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1
9ꢀ20
chelates with 8ꢀhydroxyquinoline units,
in this present communication, the synthesis,
characterization and the detailed photophysical investigation of the new ligand is described to
explore the pH dependent fluorescent behaviour of chelator.
2. Experimental
2.1 Materials and Methods
All manipulations were carried out under an atmosphere of nitrogen using standard cannula
technique. All reagents were purchased from Sigma−Aldrich at the highest commercial
quality, Alfa Aesar or Acros Organics, and used without further purification unless otherwise
stated. Solvents were distilled freshly, refluxed over appropriate drying agents following
standard procedures and kept under argon atmosphere. Melting points were determined on
melting point apparatus and are uncorrected. Reactions were monitored by thin layer
chromatography (TLC), performed on silica gel G plates using hexane and ethyleacetate
(7:3), chloroform and methanol (4:1) as eluents; developed TLC plates were visualized by
iodine vapours. Elemental analysis (CHN) was performed on the elemental analyzer Euro
Vector S.P.A−81328 (Euro EA). Infra−red spectra were recorded using an Agilent
Technologies Cary 630 FTIR spectrometer equipped with a universal attenuated total
1
reflection (ATR) sampler and samples were directly used for obtaining IR spectrum. H and
1
3
C NMR spectra were recorded in DMSO–d using a Joel 300 (MHz) ECX spectrometer.
6
Chemical shifts are reported in δ values related to an internal reference of tetramethylsilane
TMS). Mass analysis was performed in a TOF MS ES+ instrument. For solubility reason the
(
ligand was converted to its hydrochloride salt and potentiometric and spectrophotometric
studies were carried out in water purified by a Millipore Milli−Q water purification system.
Ionic strength was adjusted with 0.1M KCl. All the stock solutions were prepared by
weighing appropriate amounts by using GR−202 electronic balance (precision 0.01 mg) in
Millipore grade deionized water. The exact concentration of KOH (0.1 M) was determined
5

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potentiometrically using 0.1 M solutions of succinic acid, potassium hydrogen phthalate
KHP) and oxalic acid as primary standards and then the exact strength of HCl (0.1N) was
(
determined by same method using standardized KOH. All the solutions were prepared with
Millipore grade deionized water immediately before use, which was deoxygenated and
flushed continuously with Ar (U grade) to exclude CO and O . All measurements were
2
2
carried out at 25.0 ± 0.2°C maintained with the help of Julabo F−25 thermostat.
2.2 Synthesis of TAME5OX
The synthesis of ligand was carried out by chloromethylation of the oxine 2 followed by
condensation with tripodal 1,1,1 tris (aminomethyl) ethane) 1c as depicted in Scheme 1. The
synthesis of 5ꢀchloromethylꢀ8ꢀhydroxyquinoline hydrochloride 2a was carried out with
21
higher yields by the reported procedure. The synthesis of 1,1,1 Tris (aminomethyl)ethane
22
(
TAME) 1c was carried out under multistep reactions by the reported procedure. The
detailes of procedure for synthesis of 1c and 2a are presented in supplementary material.
2.2.1
5,5'ꢀ(2ꢀ(((8ꢀhydroxyquinolinꢀ5ꢀyl)methylamino)methyl)ꢀ2ꢀmethylpropaneꢀ1,3ꢀ
diyl)bis(azanediyl)bis(methylene)diquinolinꢀ8ꢀol (TAME5OX) (3).
To a suspension of 2a (138.28 mg, 0.6mmol) was added 1,1,1 tris (aminomethyl) ethane 1c
2 3
(23.4 mg, 0.2 mmol) in an acetone−water mixture and K CO (82.92 mg, 0.6 mmol) was used
as an acid acceptor. The mixture was refluxed with stirring on magnetic stirrer for 3 hrs. The
suspension gets dissolved upon reflux and the colour changed from yellow to brown with the
progress of reaction. The resulting mixture which contained brownish green precipitate after
cooling at room temperature, was poured into ice cold (50 mL) water and then filtered. The
solid product was collected, washed several times with diethylether and dried in vaccuo to
6

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i
ii
N3
N3
HO
OH
OH
TsO
OTs
N3
OTs
1
1a
1b
iii
Cl
iv
H N
2
NH2
N
N
.
HCl
NH2
OH
OH
2
2a
1c
v
N
N
H
H
NH
HO
OH
N
N
HO
N
3
°
Scheme 1: Reagents and conditions: (i) NaOH:H
2
O, TsCl, THF, 0 C, 6 hrs; (ii) NaN
3
,
°
DMSO, 137 C, 12 hrs, under N
2
; (iii) LiAlH
4
, dry THF, reflux, 18 hrs, under N
, acetone:water, reflux, 3 hrs.
2
; (iv) HCHO,
3
7% HCl, 50°C, 7 hrs, dry HCl gas;(v) K
2
CO
3
afford TAME5OX 3 as a off white colour powder (0.085mg, 0.17mmol, 77%). m.p. 160°C;
1
HNMR (DMSO d6, 25°C, TMS): δ = 1.18(s, 3H, CH
3
2 2
); 2.05(s, 6H, CH ); 2.4 (s, 6H, CH );
3
3
4
.61 (s, 3H, NH); 8.42(d, J H,H = 6.8 Hz, 3H, Ar–H); 8.84(d, J H,H = 7.8 Hz, 3H, Ar–H);
3
3
6
.91–6.94(t, J H,H = 7.6 Hz, 3H, Ar–H); 7.49ꢀ7.55(dd, J H,H = 8.0 Hz, 6H, Ar–H); 9.74(s, 1H,
1
3
OH) ppm. CNMR (DMSO d ): δ = 37.01 (CH ), 57.45 (CH ), 62.66 (CH ), 111.44, 117.88,
6
3
2
2
ꢀ
1
1
3
7
5
21.93, 127.57; 135.97, 138.34, 148.87, 152.51 (C=N), 153.58 (C–O). IR (ATR) ν(cm ) =
36 6 3
470, 1594, 1454, 1396, 1239, 1172, 1080. Elemental analysis Calcd (%) for C35H N O :
+
1.41 C; 6.16 H; 14.28 N; found: 71.66 C; 6.21 H; 14.35.N. MS (ES ), m/Z (I,%)
+
98.2[M+3] (20).
7

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2.3 Solution Thermodynamics: Potentiometric, Spectrophotometric and Fluorescence
measurements
The pH meter, absorption and luminescence spectrophotometer used, experimental details,
calibration of the electrode, titration procedures and quantum yield determination were used
1
9
ꢀ4
as described before. Hydrochloride salt of ligand (5ml, 1×10 M) was titrated with
standardized 0.095 M KOH for potentiometric titrations for the determination of protonation
and formation constant, respectively. Spectrophotometric titrations of a 50mL solution of
ꢀ5
TAME5OX (5×10 M) in aqueous medium and acidified with approximately 5mL (0.1M)
HCl, ionic strength maintained at 0.1 M with KCl were monitored as a function of pH over
the range 1.6−10.8 by titrating with freshly prepared standardized (0.1M) NaOH solution.
After each addition of NaOH and adjustment of pH, it was ensured that complete equilibrium
was attained (ca. 5 minutes) before measurement of the pH and UV−visible absorption
spectrum. The method adopted for the fluorescence titrations are similar to the
spectrophotometric titrations. Fluorescence spectra were registered with excitation at 385 nm
and all excitation and emission slit widths at 5.0 nm unless otherwise indicated.
2.4 Theoretical Calculations
All computations were carried out on a Pentium IV 6.0 GHz machine with a Linux operating
system. Initial geometry optimization and conformational analysis of the TAME5OX and its
20
protonated and deprotonated species were performed using the approach already described.
The PM6 optimized molecular structures were then used for DFT level calculations. All the
calculations including geometry optimization, harmonic vibrational frequency analysis, NBO
analysis and ground state electronic transitions were performed by employing the B3LYP
2
(
Becke three parameter Lee−Yang−Parr) exchange−correlation functional with the
all−electron 6–31G* basis set was used. All possible protonated and deprotonated states of
+
6
+5
+4
+3
+2
ligand (TAME5OX) described experimentally [(LH
9
) , (LH
8
) , (LH
7
) , (LH
6
) , (LH
5
) ,
8

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+
1
ꢀ1
ꢀ2
ꢀ3
(LH
4
) , (LH
3
), (LH
2
) , (LH) and (L) ] were also optimized at DFT level. To each
optimized molecule, the Hessian was calculated for the resultant stationary points and all
were characterized as true minima (i.e., no imaginary frequencies). The cartesian (x, y, z)
coordinates of minimized structures are reported as supplementary information (Table S1ꢀ
S10). Dispersion corrected DFT calculations were carried out for all the molecules using
23
B3LYPꢀD3 functional
with zeroꢀdamping. The theoretically obtained dispersion and
dispersion corrected energies for these structures are reported as supplementary information
1
13
(
Table S11). H and C NMR chemical shifts were calculated from the optimized molecule,
by the Guage−Including Atomic Orbital (GIAO). All relative chemical shifts (δ) are given
with respect to the absolute shielding values (σ) obtained at the same computational level (δ
=
(σ −σ). The calculated vibrational wavenumbers and chemical shifts were compared with
ref
the experimental data of TAME5OX. To obtain the electronic transition energies (which
include some account of electron correlation) in ground state for various acid−base species of
TAME5OX, Time−Dependent Density−Functional Theory (TD−DFT) with same functional
and basis sets as for geometry optimizations, were used.
The excited states were computed at the TD−DFT level, the cam−B3LYP density
16
functional with the all electron Pople double−ζ basis set including one diffusion functions
on heavier atoms 6−31G*. The low−lying excited−state structure ( 1) was optimized using
S
the cam−B3LYP. This functional makes use of the Coulomb attenuation method and has been
specifically parameterized with electron excitation processes in mind. To obtain properties of
the first excited state (S1) such as structure, free energy, absorption and emission energies,
three excited states were always included in the calculations. Emission wavelengths were
evaluated on the TD−DFT (cam−B3LYP) optimized structures of the excited state. All of the
2
3
computational results have been performed using the Gaussian 09 being used for spectra
convolution and molecular orbital plots.
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3. Results and Discussion
3.1 Design, Synthesis and Characterization
For an effecient metal chelator for trivalent metal ions with high complexing ability requires
a topology which has a pivotal basic skeleton with suitable spacer length with attachment
position of the binding unit; it is essential that the three electron donating arms should
arrange themself around the tripodal ligand to work cooperatively for uptake of the cation.
Keeping the above fact in view, we tried to design a suitable molecular edifice (TAME5OX)
containing tris (aminomethyl) ethane as a central unit, anchored on a quaternary carbon, and
extended by three diverging arms bearing the oxine coordination sites, connected at the C−5
15ꢀ18
position of the 8−hydroxyquinoline than commonly used Cꢀ2, Cꢀ3, or Cꢀ7 positions
. The
whole synthesis of the TAME5OX was performed through a multistep pathway by following
the general plan as depicted in Scheme 1, leading to a more reliable procedure.
Commercially available (Sigma−Aldrich) 1,1,1−tris(hydroxymethyl)ethane, 1, the
synthetic precursor bearing a methyl group at the bridgehead carbon, was converted to its
tritosylate 1a by a nucleophilic substitution reaction with tosyl chloride in THF. Intermediate
tritosylate 1a was isolated in high yield as white crystalline solid which was further reacted
with sodium azide in DMSO to afford the corresponding triazide 1b. The crude azide was
obtained as slightly yellow oil and was used immediately after work−up to avoid explosion
risks (see caution in section 4). Subsequent reduction of the triazide with LiAlH gave the
required triamine (1,1,1 tris amino methyethane) 1c in good yield. Since the triamine is a
malodorous air sensitive liquid, it was converted to its corresponding salt as stable white solid
[24]
by passing dry HCl gas into it. Triamine has been prepared by reported procedure
slight changes.
with
The derivatization of oxine starts from 8ꢀhydroxyquinoline 2, a very cheap precursor,
which is easily transformed into a 5−chloromethyle derivative 2a by chloromethylation of
1
0

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21
8
−hydroxyquinoline at 5−position (Scheme 1). The chloromethylation step is best done with
2
5
formaldehyde (40%) and HCl (37%). It has been reported that the synthesis of the free base
a could be carried out by the reaction of its hydrochloride salt with sodium hydrogen
2
26
carbonate(NaHCO ) ; however, 2a is unstable and readily undergo hydrolysis to form
3
25
hydroxymethyl quinolinol. Hence, the hydrochloride salt of 2a was directly used for
coupling. The synthesis of the final product, novel tripodal ligand based on C symmetrical
carbon anchor (TAME5OX 3), involve direct condensation of hydrochloride salts of 1c and
a. The reaction was performed by a conventional method, refluxing in acetone water
mixture (because of insolubility of hydrochloride salt of 1c and 2a in organic solvents) in
presence of K CO for 3 hrs, which led formation of the final product, TAME5OX, 3. The
3
2
2
3
compound was isolated as an air−stable brownish green colour solid, recrystalised from
chloroform as pure compound with 96% yield and was characterized by CHN analysis, FTIR,
1
13
H, COZY and C NMR and mass spectrometry.
3.1.1 Infrared spectra
Figure 2 shows comparison of experimental and theoretical IR spectra of the TAME5OX.
ꢀ1
The absorption peak at 3399 cm of TAME5OX is ascribed to the −OH stretching of
8
−hydroxyquinoline unit. The band observed at lower frequency than the normal position of
ꢀ1
free oxine, 3730−3520 cm , is attributed to presence of intramolecular hydrogen bonding in
1
the ligand, which can be also seen in H NMR of TAME5OX. The appearance of bands at
ꢀ1
2
925 and 1469 cm are due to N−H stretching and bending vibrations, respectively of
CH −NH−CH − spacer, and no peak attributable to unreacted amine or chloromethylene
group was present. The absorption peaks around 1599, 1556 and 1389 cm are due to the
−
2
2
ꢀ1
25
aromatic carbon−carbon stretching vibrations due to 8−hydroxyquinoline nucleus. The
ꢀ1
weak bands around 2112 and 1786 cm may be due to asymmetric and symmetric stretching
1
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ꢀ1
ꢀ1
vibrations of methylene groups (−CH
2
−), and near 1300 cm and 1320 cm are due to
asymmetric and symmetric stretching vibrations of the −CH
2
−NH−CH
2
−bridge.
N
H
N
H
HO
NH
OH
N
N
HO
N
Figure 2: Infra−red (IR) spectrum of TAME5OX (a) experimental (KBr) (b) theoretical
(calculated by B3LYP/6−31G*).
In order to confirm the assignments of IR frequencies, theoretical calculations were
carried out by DFT method at B3LYP/6ꢀ31G* level. The calculated IR frequencies of the
TAME5OX are shown in Figure 2. For comparison, the theoretical and experimental values
are also given in table 1. The calculated vibrational frequencies are obtained at higher values
1
2

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than experimental ones, but their trend are corroborate with the experimental data indicating
that theoretical results can be implemented for the interpretation of experimental results. The
differences in calculated and experimental spectra may be mainly due to, somewhat, to the
fact that the experimental spectra were measured in the solid phase and the theoretical spectra
were obtained in the gas phase. However, a linear correlation obtained between the
theoretical and experimental data (Figure 3) gave an acceptable correlation.
Figure 3: Correlation between the experimental and theoretical IR frequencies of
TAME5OX
3.1.2 NMR Spectra
1
The H NMR of TAME5OX presents set of signals given in section 2.3 with the assignments
proposed on the basis of the numbering scheme are given in ‘a’ of Figure 4. On proton NMR
2 3
spectra of the ligand, signals were observed characteristics to OH, CH, CH and CH protons.
Signals due to CH of 8−hydroxyquinoline were observed in the aromatic region of the
spectrum in the range of 8.87−6.9 ppm (inset of figure 4) which showed two doublets at 8.4
and 8.8 ppm, one double doublet at 7.5 and one triplet around 6.9 ppm. Singlets at 1.1, 2.0
and 2.4 ppm are assigned to the protons of one methyl (−CH
3
2
) and two methylene (−CH )
1
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groups: one attached to carbon of centre unit, and the other to 8−hydroxyquinoline unit,
respectively. TAME5OX showed a signal at 4.61 ppm due to NH proton and board peak at
(a)
1
.16
7
.06
3.81
2.47
2.47
3.81
7.06
6.80
6.80
N
H
.44
N
H
28 .. 04 4
2
.0 2.47
2.0
NH
8
9
.83
HO
OH 9.83
7.06
N
7. 8. 60 0
3.8 17 .60
N
6
8
.86
8.86
8.44
9
.83 HO
N
7.60
8.86
(b)
1
Figure 4: HNMR spectrum of TAME5OX: (a) experimental (DMSO, 300 MHz), inset
showing splitting of protons of aromatic part of ligand, and (b) calculated (applying
DFT/B3LYP/6−31G* method).
9.74 ppm for hydroxyl proton of quinoline ring; this weak and broad band of hydroxyl
protons observed were most probably resulted from intramolecular H−bonding of OH proton
27
with N atom of quinoline unit.
1
4

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13
The C NMR spectrum of TAME5OX, with the assignments proposed on the basis of
the numbering scheme, is given in Figure 5. The ligand showed bands at 34, 56 and 61 ppm
due to the CH and CH −NH−CH carbons, respectively. Signals at 148, 139, 135, 127, 126,
3
2
2
122, 120 and 110 ppm attributed to the CH groups belong to aromatic oxine unit, whereas the
peak at 153 ppm corresponds to carbon attached to −OH present in the oxine unit.
1
3
Figure 5: C NMR spectrum of TAME5OX (DMSO, 300 MHz), insets showing chemical
shifts due to aromatic carbons (left), and aliphatic carbons (right).
1
13
The theoretically calculated H and C NMR chemical shifts of TAME5OX obtained
through DFT/B3LYP method have been compared with the experimental data in Figue 4, b
and Table 1. The methyl protons showed singlet peak at 1.2 ppm. The methylene protons of
2
spacer showed signals in the spectrum at 2.05 ppm due −CH group attached to central unit
carbon and at 2.47 ppm due −CH group attached to 8−hydroxyquinoline unit. Five peaks at
2
1
5

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6
.8, 7.2, 7.6, 8.4 and 8.9 were calculated for the aromatic −CH protons of
−hydroxyquinoline unit in contrast to experimental findings where only four signals were
8
obtained for these protons. The double doublet appeared at 6.94 ppm in the experimental
1
HNMR spectrum of TAME5OX due to 2 −CH protons were calculated theoretically as two
doublets separately at 6.8 and 7.2 ppm as shown in Figure 7b. In addition the experimentally
observed peak for hydroxyl proton at 9.74 ppm, was calculated theoretically at 9.78 ppm
which was found merged with aromatic protons of oxine group in both cases due to
intramolecular hydrogen bonding. The calculated chemical shifts gave qualitative trends
although the values are not in quantitative agreement with the experiment measurements,
1
13
whereas the linearity in curves between the calculated H and C NMR values with their
respective experimental values (Figure 6, a−b) gave a perfect correlation.
1
13
Figure 6: Correlation between the experimental and theoretical (a) H NMR, and (b)
C
NMR, data of TAME5OX.
3.1.3 Mass Spectra
+
High−resolution electrospray ionization time of flight mass spectrometry TOF MS(ES ) was
used to identify the product by evaluation of the molecular ion peak of TAME5OX (Figure
1
6

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7). The spectrum of TAME5OX showed two peaks at m/Z 598.2(20%) and 599.3(10%)
+
+
corresponding to the neutral [M] and protonated [M+H] form of ligand, respectively.
Figure 7: MS(ES+) mass spectrum of TAME5OX.
1
Table 1: Calculated (B3LYP/6−31G*) and Experimental (DMSO, 300 MHz) NMR ( H and
1
3
ꢀ1
C) Chemical Shifts (ppm), and IR Frequencies (KBr, cm ) for TAME5OX.
NMR
IR
Proton B3LYP/631G*
Experimental Carbon
B3LYP/631G* Experimental B3LYP/631G* Experimental
H1
1.18
2.05
2.5
1.2
C1
C2
C3
C4
C5
36.78
41.23
55.99
61.72
111.1
37.01
42.65
57.45
62.66
111.44
3619
2998
1649
1556
1383
3470
2932
1594
1454
1396
H
2
2.0
H3
2.4
H
H
4
5
4.73
7.45
3.61
7.41
H
6
6.94
7.5
6.85
7.55
8.43
8.85
C6
C7
C8
C9
116.65
122.04
128.31
135.82
117.88
121.93
127.57
135.97
1152
1087
1022
1239
1172
1080
H7
H
8
8.42
8.84
H9
H
9
9.98
9.74
C10
C11
C12
C13
137.86
149.14
154.21
157.39
138.34
148.87
152.51
153.58
3.2 Ligand Properties: Thermodynamics and Spectrophotometry
3.2.1 Ligand Deprotonation Constants
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7

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In order to evaluate the competition between the protons and metal ions for the coordination
sites, it was necessary to determine first the acid−base properties of the ligand. The
deprotonation constants of TAME5OX were determined separately by potentiometry,
UV−visible and luminescence spectrophotometry methods. Owing to insolubility of
hexadentate ligand in water, it was dissolved in appropriate amount of standard 0.1M HCl to
make it water soluble by converting into its hydrochloride salt. The neutral TAME5OX is
addressed as LH
potentially nona−protic acid and is denoted as (H
sites: three phenolate oxygen atoms, three pyridine nitrogens and three secondary amine
nitrogen atoms of pendant arms. The extracted p values (protonation equilibria) obtained
3
, whereas the fully protonated form of TAME5OX is considered as a
6
+
9
L) . It possesses nine potential protonation
K
a
from titrations in the pH range 1.6−12.5, are defined by the following equations:
ꢁꢂꢃꢄꢅꢆꢇꢀ
ꢈꢄꢅꢇꢉ
⇋ ꢀꢁꢊꢄꢅꢆꢇꢀꢋꢄꢅꢇꢉ + ꢌꢁꢉ
ꢐꢑH ꢒ ꢔꢕꢏꢀꢁꢂꢃꢄꢅꢆꢇꢀꢈꢄꢅꢇꢉꢐꢔꢔꢔꢔꢀꢌ = 1 − 9ꢇ
ꢀ
(1)
(2)
ꢍꢎꢅ = ꢏꢀꢁꢊꢄꢅꢆꢇꢀ
ꢋꢄꢅꢇꢉ
ꢉ ꢓ
6
+
The potentiometric titration of the protonated form (H L) of the ligand was carried out in
9
º
the p[H] range of 1.2−10.5 in 0.1 M KCl with NaOH at 25 C. Due to its poor solubility in
ꢀ3
ꢀ4
water (as precipitation below pH 7.0 at concentration 1×10 M−1×10 M), the final
ꢀ
5
concentration was maintained to 1×10 M and data were analyzed by the HYPERQUAD
28
program. Analysis of potentiometric curve (Figure 8) allowed the determination of nine
deprotonation constants (the number in parentheses corresponds to the standard deviation in
the last significant digit): log K
1
= 9.89(4), log K
2
= 8.92(5), log K
3
= 8.42(4), log K
4
=
6
.71(3), log K = 6.38(2), log K = 5.88(1), log K
5
6
7
= 4.60(2), log K
8
= 4.12(1) and log K
9
=
1
8

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ꢀ5
Figure 8: Potentiometric titration curve of TAME5OX (1×10 M). Solvent H O, I = 0.1M
2
°
(
KCl), T= 25(2) C, and ‘a’ is the moles of base added per mole of TAME5OX present.
Symbols and solid lines represent the experimental and calculated data, respectively.
3.64(6). UV−visible spectrophotometric titrations were also carried out showing three buffer
regions in the pH range of 1.97−10.81: one, in the pH range of 1.97−6.69 (‘a’ of Figure 9;
corresponding spectra exhibit three isosbestic points at 217, 250 and 272 nm), second region
in the pH range 7.53−9.35 (Figure 11b, spectra exhibit three more isosbestic points at 277,
295 and 315 nm) and the third buffer region in the pH range 9.62−10.81 (‘b’ of Figure 9,
spectra overlayed exhibit λmax at 325nm). Analysis of spectrophotometric data performed by
29
the program Hypspec gave a model with same number of absorbing species (as observed by
potentiometric titration) involving three proton exchange in each pH range and also nine
deprotonation constants, which are in good agreement with that determined by potentiometric
method.
In consideration to further support the number and nature of species obtained by both
potentiometric and spectrophotometric methods and the corresponding deprotonation
ꢀ5
constants discussed above, luminescence titrations were also carried out for 1×10 M solution
of TAME5OX from 1.97−10.81 pH range. The typical example of experimental data for the
luminescence titration of the ligand is depicted in Figure 10. The best refinement of the
1
9

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luminescence−pH data by the Hypspec program corresponds to the presence of nine species,
in concurrence with the potentiometric and spectrophometric data; the values of Log Ks are
returned in table 2 (the number in parentheses corresponds to the standard deviation in last
significant digit).
−
5
Figure 9: UV−vis absorption spectra of TAME5OX (1.0×10 M) as a function of pH: (a) pH
º
=
1.97−6.69; (b) pH = 7.53−10.81. Solvent: H
2
O, I = 0.1M (KCl), T = 25.0(2) C.
Figure 10: Dependence of the fluorescence emission spectra of aqueous solution of
ꢀ5
TAME5OX (1.0 × 10 M) on the change of the pH value from acidic to basic range
°
(
1.97−10.95 pH), T = 25.0(3) C (excitation and emission slit widths of 5.0 nm) with an
excitation wavelength of 385 nm.
2
0

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[a]
Table 2. Protonation constants
(logK) of TAME5OX, (p = potentiometry, s =
spectrophotometry and f = fluorometrically) and ligands containing 8
subunits.
−hydroxyquinoline
Ligand
log K
1
log K
2
log K
3
average
log K1ꢀ3
log K
4
log K
5
log K
6
average
log K4ꢀ6
log K
7
log K
8
log K
9
average
log K7ꢀ9
[
b]
p
s
p
s
p
s
p
p
s
f
p
s
f
p
s
f
p
s
f
p
s
f
p
s
f
TAME5OX
9.89(4)
8.92(5)
8.42(4)
9.07
9.06
9.00
6.71(8)
6.79(7)
6.63(3)
6.38(2)
6.35(1)
6.30(5)
5.88(4)
5.82(3)
5.80(1)
6.32
6.32
6.24
4.60(4)
4.65(5)
4.63(3)
4.12(2)
4.08(1)
4.16(1)
3.64(4)
3.67(3)
3.62(6)
4.12
4.13
4.13
s
9
9
.82(5)f
8.97(4)f
8.39(3)f
f
.79(3)
8.87(6)
8.36(7)
[
c]
Oxinobactin
8.51(3)
7.84(1)
7.78(2)
7.03(1)
6.90(4)
6.18(2)
7.73
7.02
4.07(7)
2.75(9)
~2
2.28(2)
1.42(3)
3.02
2.07
4.07(7)
2.75(9)
~2
2.28(2)
1.42(3)
3.02
2.07
[
c]
Sulfoxinobactin
2.78
2.78
[b,d]
Tsox
TsoxMe
O−TRENSOX
9.46(3)
9.19(7)
8.62
8.44(4)
8.20(9)
8.18
6.4(1)
6.65(11)
7.44
8.1
8.01
8.08
4.4(1)
3.2(1)
ꢀ
2.55
1.8(1)
ꢀ
1.83
3.13
ꢀ
2.46
4.4(1)
3.2(1)
ꢀ
2.55
1.8(1)
ꢀ
1.83
3.13
ꢀ
2.46
[b,d]
3.94(14)
3.01
3.94(14)
3.01
[
b,e]
a Numbers in parentheses represent the standard deviation in the last significant digit. b In water (I = 0.1M KCl). c In methanol/water
(
80/20 w/w, I = 0.1M NaClO4). c Reference [30]. d Reference [32]. e Reference [31].
All constants are reported in Table 2 together with some similar ligands containing
30
8
−hydroxyquinoline or its sulphonate derivative (oxinobactin, sulfoxinobactin),
1
9
31
(
TMOM5OX) and (O−TRENSOX) based on the backbone of tris(2−aminoethyl)amine);
the structures of these ligands are also given in Figure 2 for comparison. The three highest
log K values found between 8.42 and 9.89 and three lowest values between 3.64 and 4.60 are
assigned to the deprotonation of the hydroxyl and pyridinium nitrogen moieties of the
three quinolinate groups, respectively, whereas the log K between 5.88 and 6.71 were
attributed to the secondary amines for these ligands. Inspite of the differences in solvents or
other experimental conditions, the log K values of TAME5OX and these reported ligands can
be compared: the average value for the log K of the oxygen sites, 9.07 for TAME5OX, is
much higher than the corresponding value (7.73 for oxinobactin, 7.02 for sulfoxinobactin,
8.08 for O−TRENSOX and 8.65 for Tsox). The same trend was observed for protonation of
the nitrogen sites: the average value is 4.45 for TAME5OX and (3.02 for oxinobactin, 2.07
for sulfoxinobactin, 2.46 for O−TRENSOX and 3.01 for Tsox). This differences seems to be
due to the absence of an electron withdrawing effects of sulphonate group in case of
30
oxinobactin and sulfoxinobactin and carbonyl group of amide linkage at ortho position to
31,32
hydroxyl moieties.
The higher values of TAME5OX could be due to electron donating
2
1

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effect of CH
2
−NH−CH
2
linkage attached at 5−position (para to hydroxyl group) of
8
−hydroxyquinoline. Moreover, the deprotonation constant (log K) ranges for the nitrogen
and oxygen atoms differ from the statistical factor of (log 4 = 0.6), which points to
cooperativity taking place between the three arms of the tripodal ligand, probably via intra−
or intermolecular hydrogen bonds, similar to the situation for oxinobactin and Tsox, but in
contrast to O−Trensox.
Figure 11: Species distribution curves of TAME5OX containing species, computed from the
ꢀ5
protonation constants given in Table 2. Calculated for [TAME5OX] = 1×10 M, Solvent :
tot
°
H
2
O, I = 0.1M (KCl), T = 25.0(2) C.
The corresponding distribution curves of TAME5OX obtained from the log K values are
presented in Figure 11 which indicate that in acidic solution TAME5OX initially exists 100%
in the fully protonated form as LH below pH 2.9. As the pH is increased, deprotonation
starts, the ligand loses two protons subsequently from the N−pyridyl of oxine unit with
formation of LH and LH which exist between pH 0.8−4.0 and 1.2−4.8 with maximum
9
8
7
concentration of 28% and 81% at pH 2.8 and ~3.4 respectively. Further release of protons
lead formation of LH , LH and LH successively in the pH range of 2.9−6.5 with maximum
6
5
4
2
2

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concentration 42%, 49% and 63% at pH 4.1, 4.8 and 5.2, respectively. Further deprotonation
leads to the formation of LH , the neutral form of TAME5OX, in pH range of 4.9−7.4 with
maximum concentration 57% at pH 6.3. Other species LH and LH were formed after pH 5.2
3
2
and 7.5, with maximum concentration of 82% and 61%, respectively. No isosbestic points in
the electronic spectra (‘b‘ Figure 10) were observed above the pH 9.62 and the spectra
overlapped with each other.The fully deprotonated ligand (L) was predominant above pH
9.0 with 100% concentration.
Figure 12: Correlation between the experimental log K and calculated ∆Gº of TAME5OX.
In order to validate the assignment of experimental protonation constants,
aqueous−phase free energy for the entire representative species of TAME5OX were
calculated by density functional theory by B3LYP/6ꢀ31G* quantum mechanical approach.
For an acid species AH, the pK
a
defined as the negative logarithm of the dissociation constant
ꢀ
+
of the reaction AH = A + H , is given by the thermodynamics relation pK AH/2.303
a
= ∆Gaq,
RT. The change in free energy in aqueous−phase is obtained according to the following
equation:
∆
Gꢖꢗ,ꢘꢙ = ꢚGꢖꢗ,ꢘ
ꢛ
+ꢔGꢖꢗ,ꢙ ꢝ − Gꢖꢗ,ꢘꢙꢔꢔꢔꢔꢔꢔꢔꢔꢔꢔꢔꢔꢔꢔꢔꢔꢔꢔꢔꢔꢔꢔꢔꢔꢔꢔꢔꢔꢔꢔꢔꢔꢔꢔꢔꢔꢔꢔꢔꢔꢔꢔꢔꢔꢔꢀ3ꢇ
ꢜ
2
3

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For which the proton free energy at 298 K and 1 atm is:
Gꢖꢗ,ꢙ
ꢜ
= 2.5RT−TꢀS° =1.48−7.76
=
− 6.28 kcal=mol
The greater acidity belongs to the group in which its hydrogen release is easier and there must
be a decrease in ꢀG values in the deprotonation process. A clear decreaseꢔ in ꢀG was
observed in case of TAME5OX (Figure 12) as the deprotonation take place from protonated
6
+
N− of 8−hydroxyquinoline from fully protonated free ligand, LH
9
, followed by protonated
amines of pendant arms and hydroxyl groups of binding units. The validity of this
theoretically calculated ꢀ
Gaq for the different species of TAME5OX was compared with the
2
calculated experimental pK
a
, which resulted an acceptable correlation with R = 0.98956.
3.2.2 Absorption spectra of ligand (TAME5OX) and its protonated species
−1
−1
The ligand TAME5OX shows two peaks 255 and 310 nm (ε = 110, 000 M cm and 60, 100
−1
−1
M cm , respectively) in the UVꢀvisible region assigned to π→π* and n→π* transitions.
Since, a variety of species are formed with change on pH, the large spectral changes (Figure
9
, a−b) observed in the spectrophotometric titration of the TAME5OX as a function of pH is
indicative of a change in the protonation and deprotonation behaviour of the ligand. At pH
.9, the electronic spectrum of the TAME5OX in its protonated form is symbolized by a
≤
2
−1
−1
strong absorption band at higher energies 263 nm, (ε = 110, 000 M cm ) assigned for
−
1
−1
π→π* transitions and a broad band at 380 nm, (ε = 5, 000 M cm ) assigned for n→π*
transition, respectively. Upon rising pH above 2.9, the deprotonation of the pyridinium
−1
−1
nitrogen results in hypochromic shifts (263 nm, ε = 72,000 M cm and 380 nm, ε = 2, 300
−1
−1
M cm ) with the simultaneous ascent in absorbance towards lower wavelength, 230 nm (ε
−
1
−1
=
32, 000 M cm ) which results in the formation of three isosbestic points at 217, 250 and
2
4

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272 nm as shown in Figure 9 . The isosbestic points assert the formation of three protonated
species of the ligand in the pH range of 1.97–6.69, consequently which can be well
comprehended from species distribution curves (Figure 11).
Above pH 6.7, the deprotonation of protonated secondary amines of TAME5OX, is
−1
−1
characterized by a main absorption band centered at 308 nm, (ε = 60, 100 M cm ) with a
−
1
−1
shoulder at about 279 nm, (ε =38, 000 M cm ) attributed to n→π* and π→π* transitions,
respectively, and the broad band around 380 nm get diminished in this pH range (‘b’ of
Figure 9). The deprotonation results in the bathochromic and hypochromic shifts with the rise
of pH from acidic to neutral medium. The formation of three more isosbestic points at 276
nm, 295 nm and 315 nm, confirms the formation of LH
6 5 4
, LH and LH species, respectively.
Similarly, under basic conditions the electronic spectra of ligand exhibits two bands, one at
−
1
−1
higher energies was red shifted (about 17 nm) to 325 nm, (ε = 28, 000 M cm ) and low
energy band was also hyperchromically red shifted (about 14 nm) to 290 nm, (ε = 40, 000
−1
−1
M cm ) (Figure 9), and evidenced the deprotonation of hydroxyl function from neutral form
LH ) of TAME5OX to form LH , LH and L upon variation of pH upto 10.81. Bands at λ >
00 nm have attributed to n→π* state with charge−transfer character while the band around
(
3
2
3
2
31
85 nm concerns the quinoline ring. It should be noticed that the band due to protonation of
N disappears in this pH range. No spectral change was observed on further pH variation
py
upto 11.5.
To corroborate the above argument for the deprotonation behavior of ligand, the
electronic spectra of different species of TAME5OX obtained were calculated by
time−dependent density functional theory (TD−DFT) at the B3LYP/6−31G* level. The
calculated spectra for the neutral (LH ), protonated and deprotonated species (LH , LH , LH ,
3
9
8
7
LH
6 5 4 2
, LH , LH and LH , LH) were obtained by stepwise protonation and deprotonation of
N−, NH− and OH− groups, respectively (Figure 13). The experimental spectra of TAME5OX
2
5

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gave major variations for the π−π* transition; the peak of protonated (LH
9
, LH
8
and LH
7
) at
262 nm (‘a’ of Figure 13) shifted bathochromically with formation of LH
6
, LH
5
and LH
4
,
show transitions at 274, 276 and 278 nm, respectively. Similar variations were also observed
in the calculated electronic spectra (‘b’ of Figure 13) and the corresponding calculated values
Figure 13: pH−dependent electronic spectra (absorption) of the nine species as a function of
molar absorptivity and wavelength (a) predicted from Hypspec using experimental data and
(b) calculated through TD−DFT/B3LYP method by employing 6−31G* basis set, for the
protonation and deprotonation of ground state geometrical optimized neutral species (LH
TAME5OX.
3
) of
are reported in table 3. The calculated π−π* and n−π* transitions for the LH
6 5 4
, LH and LH
obtained at 245 nm and 280 nm, shifted to 260 nm and 300 nm for the species LH , LH and
3
2
LH respectively. The simulated electronic absorption spectra for corresponding predicted
species are in good concurrence in the contour, number of absorption bands and relative
intensities with the species except small red−shifts in LH , LH and LH forms, which are due
3
2
to the impossibility of considering all the symmetrical orbitals in TD−DFT calculation. Also,
strong absorption peaks were obtained from theoritical calculations at 264 nm for LH and at
species, which
9
the same position with comparatively lower intensities for LH
corroborates well with the experimental results.
8 7
and LH
2
6

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Table 3: Computed Optical Properties: Absorption−S0 and emission−S1; wavelengths,
oscillator strengths (fcalc), electronic transitions and energies calculated at the
TD−DFT/6−31G* level of theory, using DFT/B3LYP and DFT/cam−B3LYP for ground state
and excited state geometry, respectively, for different species of the TAME5OX.
S0
S1
Species
λnm (abs)
f
calc
Nature
Transition
Excitation energy)
λnm(em)
f
calc
Nature
Transition
(Emission energy)
(
6
5
+
+
(
(
H
9
L)
270.54
272.34
0.3281
HOMOꢀ2→LUMO+1
110→114 (1.186 eV)
425.64
425.89
0.1091
0.1581
HOMOꢀ1→LUMO+1
HOMO→LUMO+1
111→114 (0.162 eV)
H
H
8
7
L)
L)
0.1794
0.1563
HOMOꢀ1→LUMO
HOMOꢀ2→LUMO
111→113 (0.883 eV)
110→113 (0.726 eV)
112→114 (0.293 eV)
4
3
+
+
(
(
274.89
220.84
519.95
522.83
0.1674
0.1748
HOMOꢀ2→LUMO
HOMOꢀ1→LUMO
110→112 (0.385 eV)
111→112 (0.571eV)
H L)
0.1068
0.0921
HOMOꢀ1→LUMO
HOMO→LUMO
111→113 (0.590eV)
112→113 (0.513 eV)
6
3
10.23
2
+
(
(
(
(
(
(
H
H
H
H
5
4
3
2
L)
L)
L)
L)
223.60
19.37
0.0821
0.0731
HOMO→LUMO+1
HOMOꢀ1→LUMO
112→ 114 (0494 eV)
111→113 (0.489 eV)
526.64
528.93
530.37
474.67
470.23
429.34
0.1921
0.2107
0.2193
0.2116
0.1875
0.1845
HOMOꢀ1→LUMO
HOMO→LUMO+1
HOMOꢀ1→LUMO+1
HOMO→LUMO
111→ 112 (0.836 eV)
112→114 (1.243 eV)
111→114 (1.391 eV)
112→113 (1.287 eV)
111→113 (0.723eV)
110→113 (0.596 eV)
3
+
225.30
37.42
0.0675
0.0598
HOMOꢀ1→LUMO+2
HOMOꢀ1→LUMO
111→115 (0.314 eV)
111→113 (0.379 eV)
3
229.91
50.42
0.0401
0.0463
HOMOꢀ2→LUMO+1
HOMO→LUMO
110→114 (0.253 eV)
112→113 (0.288 eV)
3
ꢀ
234.98
72.87
0.0374
0.0412
HOMOꢀ1→LUMO+1
HOMO→LUMO
111→114 (0.192 eV)
112→113 (0.219 eV)
3
2
ꢀ
HL)
244.26
76.05
0.0283
0.0403
HOMOꢀ1→LUMO
HOMO→LUMO
111→113 (0.175 eV)
112→113 (0.209 eV)
HOMOꢀ1→LUMO
HOMOꢀ2→LUMO
3
3ꢀ
L)
245.16
79.08
0.0138
0.0294
HOMO→LUMO+1
HOMO→LUMO
112→114 (0.159 eV)
112→113 (0.184 eV)
3
3.2.3 Fluorescence Spectra of ligand (TAME5OX)
In general, the binding moiety of 8−hydroxyquinoline serves as a fluorophore because of the
significant enhancement of fluorescence that is known to occur when the excited state
intramolecular proton transfer (ESIPT) between the hydroxyl group and quinoline nitrogen is
33
suppressed upon binding of a metal cation. However, some HQ derivatives are poorly
34
luminescent due to an intramolecular photoinduced proton transfer (PPT) process between
the hydroxyl group (which is relatively strong photoacid) and the nearby quinoline nitrogen
(which is, in turn, a strong photo base). In addition, in protic media, intermolecular PPT
process involving solvent molecules can also occur, further decreasing the fluorescence
quantum yield of this kind of fluorophore. The 8−hydroxyquinoline ligands are indeed
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DOI: 10.1039/C4RA16345A
characterized by strong intramolecular hydrogen bonding that may be photoinduced to
35,36
undergo proton transfer in the excited state.
In the present tripodal ligand containing three units of 8−hydroxyquinoline, upon
excitation into the ligand transitions, a maximum of emission at about 425 nm was observed
(Figure 10). This fluorescence emission originates from an excited–state intramolecular
proton transfer (ESIPT), and enhanced fluorescence with absolute quantum yield ≈0.091)
may be attributed to presence of three units of 8−hydroxyquinoline in TAME5OX.
3.2.3.1 pH−Dependent Fluorescence Spectra
From survey of pHꢀdependent photophysical properties of 8ꢀhydroxyquinoline compounds, it
is seen that, the fluorescence intensity of these compounds changes in a monotonous way,
depending only on the change of pH, which gets increased or decreased along with the
change in the pH value in a consistent way due to the only one photoinduced electron transfer
mechanism between pyridyl −N and −OH group of quinoline unit, resulting in the OFF−ON
3
7
or ON−OFF type of fluorescent pH sensors. Moreover, there are reports on the pH sensors
based on the opposite photoinduced electron transfer processes between a receptor and
38
signaling unit. To explore the pH−dependent optical properties of TAME5OX with unique
pyridyl−N and −OH structural characteristics, the pH vs fluorescence titration experiment
was carried out in aqueous medium. An appropriate volume of 0.1M HCl and 0.1M NaOH
were used for protonation of pyridyl −N group and deprotonation of the −OH group,
respectively. As shown in Figure 9, the π−π* and n−π* transitions in the electronic
absorption spectrum of TAME5OX exhibit the exemplary behaviour (red and hypochromic
shifts) along with changing the pH of the system from acidic to basic (as discussed in
protonation of TAME5OX). In good comparison, the changes in the blue−green emission of
the ligand maxima located at 425 nm which was excited at 385 nm, were monitored as a
function of pH ranging from 1.97−10.81 (Figure 10) at room temperature. The emission
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RSC Advances
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DOI: 10.1039/C4RA16345A
intensity at 425 nm (with meagre quantum yield) as well as maximum wavelength remains
almost unchanged, no substantial change in behaviour of the fluorophore was observed upon
measuring emission under acidic conditions within the pH range of 1.97−2.88. Most
interestingly, with an increase in the pH from 3.04 to 6.69, resulted the evolution of emission
peak at 520 nm upto ∼5 fold (could be attributed to the competition between the
intramolecular photon induced proton transfer), while no change in blue−green emission at
425 nm. Further rise of pH resulted the intense fluorescence of TAME5OX gradually get blue
shifted (about 50 nm) towards 470nm, along with quenching of fluorescence signal about 1, 2
and 3−fold at 7.53, 7.94 and 8.27 pH, respectively. After addition of more base (0.1M NaOH)
(pH > 8.5), the deprotonation effect again evidenced in growth of fluorescence signal upto
3
−fold which promptly got blue shifted to original blue−green emission wavelength (420
nm). Surprisingly, with an increase in the pH from 8.9 to 10.9, the intense fluorescence of the
same compound under basic conditions also gets gradually quenched in a similar manner as
observed under acidic conditions, (cf. Figure 10). It is worthnoting that more significant
changes under physiological pH and less significant changes under acidic and basic medium
confirm that protonation of N atom lowers the LUMO energy and decreases the vertical
S1→S0 transition energy, and the deprotonation of the hydroxyl function also destabilize the
HOMO, thus closing the gap giving rise to smaller transition, while in neutral form the
HOMO−LUMO gap increases which corresponds significant red shifts along with intense
fluorescence. Thus, among all the species formed during the pH range of 1.8−10.9, the
neutral species, LH , was found to be more fluorescent that predominates under physiological
3
pH, while the more protonated and deprotonated species were found to be less fluorescent.
These results divulge the typical characteristics of a pH−fluorescent probe through the
photoinduced proton transfer (PPT) enhancement process and photoinduced electron transfer
34,38
(PET) quenching processes between pyridyl−N and −OH group for this tripodal chelator.
2
9