Formation of very stable and selective Cu(II) complexes with a non-macrocyclic ligand: Can basicity rival pre-organization?

Article abstract of DOI:10.1039/c0dt00453g

The synthesis of ligand L based on a 2,6-bis[(N,N-bis(methylene phosphonic acid)aminomethyl] pyridine scaffold is described. Potentiometry combined with UV-Vis absorption spectrophotometric titrations were used to determine the protonation constants of the ligand and the stability constants of its corresponding Cu(II), Ni(II), Zn(II) and Ga(III) cations (0.1 M NaClO ₄, 25.0 °C). The physico-chemical approach revealed very large stability constants for Cu(II) complexation (logK_CuL = 22.71(7)) reflected in a very high pCu^II value of ～ 15.5 (pH = 7.4, [L]_tot = 10^-5 M, [Cu]_tot = 10^-6 M), close to those measured for the strong methylphosphonate functionalized cyclen chelators. Based on a literature survey, a correlation is proposed between the pK values of branched polyamine ligands and their stability constants for Cu(II) complexation, allowing for an estimation of the latter on the basis of the protonation constants of L. Ligand L was also shown to be very selective towards Cu(II) compared to the other cations studied (ΔlogK > 4). UV-Vis spectroscopy and kinetic measurements indicated that the formation of the cupric complexes with L is very fast, which, in combination with all other properties, makes it an excellent non-cyclic target for Cu(II) radiopharmaceutical within the frame of ⁶⁴Cu positron emission tomography imaging and radiotherapy. The Royal Society of Chemistry 2010.

Full text of DOI:10.1039/c0dt00453g

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Formation of very stable and selective Cu(II) complexes with a
non-macrocyclic ligand: can basicity rival pre-organization?†
Sabah Abada,a Alexandre Lecointre,a Mourad Elhabiri*b and Lo¨ıc J. Charbonnie`re*a
Received 10th May 2010, Accepted 5th July 2010
DOI: 10.1039/c0dt00453g
The synthesis of ligand L based on a 2,6-bis[(N,N-bis(methylene phosphonic acid)aminomethyl]
pyridine scaffold is described. Potentiometry combined with UV-Vis absorption spectrophotometric
titrations were used to determine the protonation constants of the ligand and the stability constants of
its corresponding Cu(II), Ni(II), Zn(II) and Ga(III) cations (0.1 M NaClO4, 25.0 ◦C). The
physico–chemical approach revealed very large stability constants for Cu(II) complexation (logKCuL
=
22.71(7)) reflected in a very high pCuII value of ~ 15.5 (pH = 7.4, [L]tot = 10-5 M, [Cu]tot = 10-6 M), close
to those measured for the strong methylphosphonate functionalized cyclen chelators. Based on a
literature survey, a correlation is proposed between the pK values of branched polyamine ligands and
their stability constants for Cu(II) complexation, allowing for an estimation of the latter on the basis of
the protonation constants of L. Ligand L was also shown to be very selective towards Cu(II) compared
to the other cations studied (DlogK > 4). UV-Vis spectroscopy and kinetic measurements indicated that
the formation of the cupric complexes with L is very fast, which, in combination with all other
properties, makes it an excellent non-cyclic target for Cu(II) radiopharmaceutical within the frame of
64Cu positron emission tomography imaging and radiotherapy.
Up until now, most of the BFCs for Cu(II) coordination11
Introduction
were concerned with cyclen and cyclam based structures and
their branched or cross-bridged analogues (R π H in Chart 1).11
Although, they were undoubtedly the best candidates regarding
their in vitro stability, they sometime present drawbacks such as
tedious synthetic protocols (especially for the introduction of the
labeling functions for grafting on biomaterials), slow kinetics for
Cu(II) complexation often due to a “proton sponge” effect,12 or
weak in vivo stabilities.13
Copper cation plays a major role in many biological systems
and misregulation of its homeostasis can result in severe illness.1
For instance, there is significant evidence to suggest that Cu(II),
among others (e.g. Zn(II) and Fe(III)), is directly involved in the
pathogenesis of Alzheimer’s disease.2 Indeed, b-Amyloid peptides
(Ab) display high affinities for Cu(II) (KD ~ 1 mM),3 which
mediates its precipitation as insoluble and toxic aggregates.4 These
metal ions being highly concentrated in the neocortex of AD
patients,5 the Cu(II) binding by Ab might cause the production
of reactive oxygen species (ROS), which are also associated to
neurodegeneration.5,6 It was shown that Zn(II)- or Cu(II)-induced
Ab precipitation is reversed by treating the aggregates with
exogenous ligands,7 justifying the search for biocompatible Cu(II)
chelators as potential therapeutic agents.8
Concomitantly, the increasing number of available research cy-
clotrons worldwide, together with the advances in the production
of 64Cu from 64Ni,9 recently led to a revival of the interest in 64Cu
and 67Cu chelation and in its application to positron emission
tomography (PET) imaging and radiotherapy.10 For this purpose,
various bifunctional chelators (BFCs)11 or copper(II) complexing
reagents12 have been designed to firmly and selectively hold 64Cu
and to deliver it to the biological targets.
Chart 1 Chemical structures of cyclen, cyclam and LH8
In our quest to fulfil the criteria for 64Cu complexation: i.e. (i)
strong Cu(II) complexation; (ii) selectivity towards Zn(II), Ni(II)
and endogenous cations; (iii) fast kinetic of complexation at room
temperature and (iv) stability towards reduction to Cu(I), we
developed the synthesis of ligand L, a 2,6-diaminomethylpyridine
functionalized with four methanephosphonic acid functions.
aLaboratoire d’Inge´nierie Mole´culaire Applique´e a` l’Analyse, IPHC, UMR
7178 CNRS-UdS, ECPM, 25 rue Becquerel, 67087, Strasbourg Cedex 2,
France. E-mail: l.charbonn@unistra.fr
Results and discussion
bLaboratoire de Physico-Chimie Bioinorganique, Institut de Chimie, UMR
7177 CNRS-UdS, ECPM 25, rue Becquerel, 67087, Strasbourg, Cedex 2,
France. E-mail: elhabiri@chimie.u-strasbg.fr
Synthesis of the ligand
† Electronic supplementary information (ESI) available: 1H and 13C-NMR
spectra of L; list of ligands used for Fig. 4 and corresponding references
of the literature. See DOI: 10.1039/c0dt00453g
The synthetic protocol is summarized in Scheme 1. Starting
from the commercially available 2,6-dihydroxymethyl pyridine,
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Scheme 1 Synthesis of ligand LH8 (i) PBr3, DMF, 84%. (ii) HMTA, CHCl3, 80 ◦C, 3 h; conc. HCl, MeOH, reflux, 17 h, quant.14 (iii) EtOH, NaOH,
reflux, 1 h; HPO(OEt)2, 37% formaldehyde in H2O, 0 ◦C; 100 ◦C, 60%. (iv) TMSBr, CH2Cl2; MeOH, 83%.
2,6-dibromomethylpyridine was obtained by a bromination with
PBr3. Following a literature procedure, a Delepine-type reaction
afforded the 2,6-diaminomethylpyridine 1,14 which was further
converted to the tetramethylphosphonic ethyl ester 2 by reaction
with formaldehyde and diethylphosphite. The phosphonic acid
functions were recovered by hydrolysis of the phosphonic esters
with TMSBr followed by methanolysis to afford LH8 as its
hydrated hydrobromic salt.
and logKLH2 values are more than two orders of magnitude
higher than the corresponding ones measured for the analogue
L¢, which can be explained by the stronger repulsions of the
2-
negative charges of the –PO3 moieties with respect to the –
CO2- units, which prevails over the electron withdrawing effect.18,19
Markedly different hydrogen bonding between the protonated
ammonium units and the negatively charged phosphonates or
carboxylate moieties can also largely account for the very dif-
ferent acido–basic properties.20 Intricate hydrogen bond networks
involving the pyridine chromophore cannot be, indeed, excluded
since a spectrophotometric titration vs. pH of the free ligand L
indicated significant variations of the pyridine-centered transitions
(hypochromic shift), particularly when the two tertiary amines and
the two first methanephosphonates are protonated.
Physico–chemical studies. Using a combination of pH-metric
and UV-Vis absorption spectrophotometric vs. pH titrations, it
was possible to determine the protonation constants (KLHx) of L8-
(i.e. the fully deprotonated ligand) and the corresponding stability
constants (KMLHx) for the formation of the metal complexes with
Cu(II), Zn(II), Ni(II) and Ga(III) (water; I = 0.1 M NaClO4;
T = 25.0(2) ◦C). All the thermodynamic data are gathered in
Table 1, together with some relevant data obtained for ligand
L¢, an analogue of L containing carboxylic acids instead of the
phosphonic functions in L, and DO2P, one of the most efficient
polyaminomethanephosphonic ligand for Cu(II) complexation, to
the best of our knowledge.15
The four following protonation constants, which were accu-
rately determined, were attributed to the first protonation of the
four phosphonate functions. The second successive protonation
constant for each –PO3H- functions were assumed to be lower
than 101 M-1 in agreement with numerous other systems.21 Lastly,
the logKH value related to the pyridine unit is also estimated to
be lower than 2.19 Therefore, under our experimental conditions
Within the pH range studied (2.5 < pH < 12), ligand L
displayed two basic protonation constants above pH 10 (Table 1),
which were unambiguously assigned, by analogy with L¢, to the
protonation of the two tertiary amine functions. The logKLH
2-
(starting pH ~ 2.5), ligand L exists as a LH6 protonated species
(Fig. 1).
The Cu(II) complexes with L were then characterized and
quantified by pH-metric and spectrophotometric titrations vs.
pH. Only protonated monocupric monochelates CuLHy were
identified with y = 0 to 4 (logKCuLH = 8.24(8), logKCuLH2 = 7.15(9),
logKCuLH3 = 5.7(1), logKCuLH4 = 4.0(2)). Interestingly, the stability
constant of the Cu(II) complex with L is rather high and represents
one of the most stable cupric species reported until now for non-
macrocyclic branched compounds (to be compared for example
with ethylene-diamine-tetramethylene-phosphonic acid EDTPA
Table 1 Successive protonation constants (logKLHx), formal stability
b
constants (logKML) and pCuII values for ligands L,a L¢ and DO2Pc
b
La
L¢
DO2Pc
logKLH
logKLH2
logKLH3
logKLH4
logKLH5
logKLH6
logKCuL
logKNiL
logKZnL
logKGaL
pCuIId
11.21(2)
10.29(2)
8.04(4)
6.49(6)
5.53(8)
8.95
7.85
3.38
2.48
12.80(8)
10.92(2)
8.47(2)
6.39(2)
with logKCuL = 23.2, pCuII = 12.7 at pH 7.4 and [EDTPA]tot = 10-5
M
and [Cu(II)]tot = 10-6 M).22 This very high stability was further
4.19(9)
confirmed by determining the conditional stability constant K*
CuL*
22.71(7)
16.50(3)
17.84(4)
16.31(9)
14.4d/15.5e
15.69(2)
28.7
under acidic conditions (logK*
= 4.25(4) and 5.82(9) at pH 1.0
CuL*
15.84(2)
21.2(3)
17.8d
and 2.0, respectively, L* stands for the protonated form of ligand
L at either pH 2.0 or pH 1.0 and K* designates the conditional
stability constant). Fig. 1 displays the speciation diagrams of the
protonated cupric species with L for a ratio [Cu(II)]tot/[L]tot = 1 as
a function of pH.
If we now compare the pCuII values measured at pH 7.4
(Table 1), we clearly observe that the substitution of the terminal
carboxylate functions in L¢ by phosphonates in L induces a
sizeable stabilization of the Cu(II) complex by almost two orders of
magnitude, which can be related to the increase of the global amino
basicity in L (logbLH2 = 21.50) with respect to L¢ (logbL¢H2 = 16.80).
As no protonation constants for the cupric complexes with DO2P
are available in the reported literature, the pCuII values reported in
12.7d/12.7e
a Solvent: water; I = 0.1 M (NaClO4); T = 25.0(2) ◦C. Errors = 3s with s
= standard deviation. KLHx = [LHx]/[[LHx-1][H] and KML = [ML]/[L][M].
Charges have been omitted for the sake of clarity. logKCu(OH) = -6.29 and
logKCu(OH)2 = -13.1; logKZn(OH) = -7.89 and logKZn(OH)2 = -14.92; logKNi(OH)
= -8.1 and logKNi(OH)2 = -16.87 (16a); logKGa(OH) = -2.65 (16b). b L¢ =
2,6-Bis[bis((carboxymethyl)amino)methyl]pyridine (17); solvent: water;
T = 25 ◦C. c DO2P = 1,4,7,10-Tetraazacyclododecane-1,7-◦bis(methane
phosphonic acid) (15); solvent: water; I = 0.1 M (KCl); T = 25 C. d pCuII =
-log[Cu(II)]free for [Cu(II)]tot = 10-6 M, [L]tot = 10-5 M, pH = 7.4 and
calculated without considering the protonation constants of the Cu(II)
complex. e pCuII calculated with the protonation constants of the Cu(II)
complex.
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Table 2 Spectroscopic parameters of the protonated Cu(II) complexes
formed with L.a
Absorption bands lmax (e)/nm(M-1 cm-1)
Species
Pyridine p → p*
N → Cu CT
Cu(II) d–d
CuL6-
266 (5520)
266 (5670)
264 (5490)
359 (3350)
329 (2140)
315 (2395)
703 (236)
702 (153)
653 (153)
CuLH5-
4-
CuLH2
a Solvent: water; I = 0.1 M (NaClO4); T = 25.0(2) ◦C.
Table 3 Formal stability constants (logKML) for ligand L, and related
polyaminomethanephosphonate analogues
L
logKCuL logKNiL
logKZnL
logKCoL
a,25
R = –CH3
14.32
17.2
9.59 (4.73)
11.7 (5.5)
10.44 (3.88) 9.27 (5.05)
2-
R = –CH2PO3
14.6 (2.6)
14.0 (3.2)
(NTPA)b,26
R = –C2H5
c,27
13.26
12.41
8.14 (5.12)
7.58 (4.83)
9.33 (3.93)
9.21 (3.2)
7.95 (5.31)
7.75 (4.66)
c,27
R =
9.73
3.94 (5.79)
9.12 (0.61)
3.31 (6.42)
d,28
EDTPAb,29
Ld
23.21
22.71
16.38 (6.83)
16.50 (6.21)
18.76 (4.45) 17.11 (6.1)
17.84 (4.87) nd
Solvent: H2O; T = 25 ◦C.a I = 0.1 M (KCl). b I = 0.1 M KNO3. c I = 0.2 M
(KCl). d I = 0.1 M (NaClO4). The values in parentheses correspond to the
difference logKCuL - logKML
.
Fig. 1 Speciation diagrams of the species present in solution as a function
of pH for L ([L]tot = 2 ¥ 10-3 M, top) and for a stoichiometric mixture of
L and Cu(II) ([L]tot = [Cu(II)]tot = 2 ¥ 10-3 M, bottom). Solvent: water; I =
0.1 M (NaClO4), T = 25.0(2) ◦C.
the worst case. These differences are in good agreement with
those calculated for comparable polyaminomethanephosphanate
derivatives (Table 3).
Table 1 were calculated without considering the protonated Cu(II)
species for the sake of comparison. Taking into account the overall
thermodynamic constants, we can calculate higher pCuII values of
15.5 and 12.7 for ligand L and L¢, respectively. The comparison
of the pCuII data measured for the tetraazacyclododecane-based
ligand DO2P and for the non-macrocyclic compound L shows a
stabilization of about three orders of magnitude (pCuII = 17.8 and
14.4 for DO2P and L, respectively). Interestingly, this improved
stability can also be related to the higher global branched amine
basicity of DO2P (logbDO2PH4 = 23.72) by comparison with L
(logbLH2 = 21.5) and emphasizes that, despite the lack of preor-
ganization of L with respect to DO2P, the Cu(II) complexes with
L are very stable species in solution. The formal stability constants
of the metal complexes with M(II) = Zn(II), Ni(II) and Ga(III) have
been determined by potentiometric methods and are gathered in
Table 1. The corresponding stability sequence satisfactorily follows
the Irving–Williams23 order with the Cu(II) complexes being the
more stable due to Jahn–Teller distortions.24 Importantly and as
anticipated,23 the complexation of Cu(II) by L is very selective
when compared to those of Zn(II), Ni(II) or Ga(III), with almost
five orders of magnitude of difference in logKML (Table 1) in
Fig. 2 displays the variation of the pM values (M = Cu(II), Zn(II),
Ni(II) and Ga(III)) vs. pH, which clearly illustrates the expected
Fig. 2 Evolution of the pM values (pM = -log[M]free with M = Cu(II),
Zn(II), Ni(II) and Ga(III)) as a function of pH. Solvent: water; I = 0.1 M
(NaClO4); T = 25.0(2) ◦C; [L]tot = 10-5 M and [M]tot = 10-6 M.
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selectivity of ligand L toward Cu(II) with respect to the other
cations studied over a wide range of acidity.
to the weakening of the PO32-–Cu(II) bonds upon protonation in
favor of a strengthening of the N–Cu(II) bonds. Lastly, a broad
and weak absorption band (e ~ 150–240 M-1 cm-1) was evidenced
in the visible region (lmax 650–700 nm) and was attributed to
d–d transitions on Cu(II), indicative of an octahedral or square
pyramidal geometry,33 while d–d transitions are observed above
800 nm for triangular based bipyramidal complexes.34 The hyp-
sochromic shift of ~ 50 nm upon protonation of CuLH5- species
UV-Vis absorption spectroscopy
Fig. 3 displays the evolution of the UV-Vis absorption spectra
of the CuL complex as a function of pH, together with the
corresponding electronic spectra of the Cu(II) species formed along
the titration, which were calculated by statistical methods.30 Table 2
summarizes the main spectroscopic parameters associated with
these Cu(II) species. Whatever the pH, the absorption spectra
are dominated by a structured band at ca. 265 nm (e265 ~ 5.5 ¥
103 M-1 cm-1) related to the p → p* transitions of the pyridine
moiety.31 In addition, the electronic spectra are characterized by
an absorption band of weaker intensity (e ~ 2.0–3.4 ¥ 103 M-1 cm-1),
with a maximum of absorption ranging from 315 nm to 359 nm,
depending on the protonation state of the Cu(II) complex.
These absorption bands induced by Cu(II) complexation most
likely correspond to Namino → Cu(II) charge transfer (LMCT)
transitions.27a,32
4-
to afford CuLH2 most likely indicates a drastic change of the
Cu2+ coordination geometry.33 The involvement of the N pyridine
atom in the metal coordination sphere cannot be excluded and can
be related to the extra-stabilization of the Cu(II) complexes with
L (pCuII = 15.5, Table 1) when compared with those formed with
EDTPA (pCuII = 12.7). Solid state structures of comparable linear
or macrocyclic analogues points out the potential participation of
the aromatic ring nitrogen atom to Cu(II) coordination.19,35 Our
spectroscopic data correlate with those obtained with bis- and tris-
(methanephosphonate) derivatives of a 14-membered tetraaza-
macrocycle containing pyridine.19 Thorough physico–chemical
studies with appropriate models are currently under progress to
fully elucidate the structural and geometrical properties of these
Cu(II) complexes with L.
Thanks to these spectroscopic probes and preliminary kinetic
experiments, it was noticed that the copper complexation is instan-
taneous, an important criterion for the use of such compounds
within the frame of 64Cu and 67Cu complexation for PET imaging
or radiotherapeutic applications.
The 1 : 1 Cu : L composition was also confirmed by isolation
of the Cu(II) complex and its characterization by electrospray
mass spectrometry in the negative mode. The spectrum displays a
major peak at m/z = 572.955, which unambiguously corresponds
to the [CuLH5]- species, as evidenced by its corresponding
isotopic distribution. Adducts containing sodium cations were
also detected (Fig. 4).
Fig. 3 Spectral changes vs. pH recorded for an equimolar solution of
Cu(II) and L (water; I = 0.1 M (NaClO4); T = 25.0(2) ◦C; [L]tot = 1.41 ¥
10-4 M; [Cu(II)]tot = 1.40 ¥ 10-4 M, top) and electronic absorption spectra
of the protonated cupric complexes with L (bottom). Solvent: water; I =
0.1 M (NaClO4); T = 25.0(2) ◦C; [L]tot = 1.41 ¥ 10-4 M; [Cu(II)]tot = 1.40 ¥
10-4 M. Inset: evolution of the absorbances at 350 and 310 nm.
Fig. 4 ES/MS spectra of the [CuL]Na6 complex in water. Inset: expansion
of the region of the molecular peak (a) and its simulated pattern (b).
Discussion
The stepwise hypsochromic shifts of 30 nm and 14 nm of the
LMCT band upon protonation (Fig. 3, bottom) can be ascribed
In order to better understand the origin of such a strong
Cu(II) chelation, we focused our attention towards the hypothesis
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proposed by Lukes and co-workers, who postulated that, rather
than the chemical structure and topography of the ligand, the
two first pK values of the amine backbone are the dominating
parameters for the complexation of Cu(II).18 Fig. 4 summarizes
a compilation of some 49 compounds available in the literature,
corresponding to linear or macrocyclic branched polyaza ligands
for which the logKCuL and the pK are well documented. A full
description of the ligands used and the corresponding references
are provided in the ESI.†
As evidenced in Fig. 5, the hypothesis proposed by Lukes and
co-workers18 can be confirmed and a linear correlation of logKCuL
with the sum of the two first pKs can be evidenced. On the basis of a
simple linear regression, we can obtain a first rough approximation
of the stability constants for branched polyaza ligands with Cu(II)
using the two first pK values of the ligands and the following
equation :
2,6-dibromomethylpyridine, the latter being obtained according
to the procedure described below.
Synthesis of 2,6-dibromomethylpyridine
2,6-Dihydroxymethylpyridine (3.70 g, 26.6 mmol) was dissolved
in DMF (15 mL). The flask was immersed in an ice bath and
vigorously stirred and PBr3 (16.6 g; 5.75 mL; 61.2 mmol) was
slowly added dropwise. The reaction mixture rapidly turned black.
The temperature was raised to r.t. and the solution was stirred
for 4 h. Water (20 mL) was slowly added and the aqueous layer
was extracted with Et2O. The organic extracts were dried over
Na2SO4, filtered and evaporated to dryness. The solid residue was
purified by column chromatography (SiO2, CH2Cl2) to afford 1
as a white solid in 84% yield (5.90 g). All analyses correspond to
those reported in the literature.37
logKCuL = 1.20 ¥ (pK1 + pK2) - 3.63 (R2 = 0.916)
(1)
Synthesis of compound 2
To the hydrochloride salt of 2,6-diaminomethylpyridine (810 mg,
3.3 mmol)14 and ethanol (20 mL), was added NaOH (395 mg,
9.9 mmol). The mixture was refluxed for 1 h. The solvent was
removed under reduced pressure and the resulting residue was
taken up with HPO(OEt)2 (1.82 g, 13.2 mmol). The mixture was
stirred for 10 min in an ice bath and 1.6 mL of formaldehyde (37%)
was slowly added, while keeping the temperature below 10 ◦C. The
reaction mixture was stirred at r.t. for 30 min and at 100 ◦C for 14 h.
The cooled solution was concentrated under reduced pressure.
The crude product was purified by column chromatography over
silica gel (CH2Cl2–MeOH gradient from 100 : 00 to 88 : 12) to give
compound 2 (1.42 g, 60%) as a colourless oil. 1H NMR (CDCl3,
200 MHz): d 7.62 (t, J = 7.7 Hz, 1 H), 7.43 (d, J = 7.5 Hz, 2 H),
4.08 (q, J = 7.3 Hz, 16 H), 4.05 (s, 4 H), 3.17 (d, J = 10.0 Hz, 8
Fig. 5 Representation of the logKCuL values as a function of the sum of
the two first pK values of the amines of linear (stars) and cyclic (squares)
branched polyaza ligands (only one pK was used for monoamines). The
black dot represents the value determined for L by potentiometry.
1
H), 1.27 (t, J = 7.1 Hz, 24 H). 13C { H} NMR (CDCl3, 50 MHz):
d 157.8, 136.9, 122.1, 62.4 (t, J = 8.0 Hz), 61.9 (d, J = 7.0 Hz),
50.2 (dd, J = 158.7 Hz, J = 8.6 Hz), 16.4 (d, J = 6.5 Hz). 31P NMR
(CDCl3, 400 MHz): d 24.4. IR (cm-1, ATR): n 2979 (w), 2930 (w),
2909 (w), 1680 (s), 1442 (w), 1390 (w), 1230 (s, nP=O), 1160 (m),
1013 (s), 957 (s). ESI+/MS (MeOH–H2O): m/z = 738.3 ([M + H]+,
100%). Anal. Calcd. for C27H55N3P4O12, 2H2O: C, 41.91; H, 7.68;
N, 5.43. Found: C, 41.88; H, 7.30; N, 5.80.
Using eqn (1), the logKCuL value estimated for L is equal to 22.2,
in reasonably good agreement with the value of 22.7 determined
in this study by potentiometric method.
Synthesis of LH8
Experimental
Compound
2 (650 mg, 0.88 mmol) was dissolved in
Synthesis of the ligand
Materials and methods
dichloromethane (10 mL) and TMSBr (4.7 mL, 35.2 mmol) was
added. The solution was stirred at r.t. for 24 h. A second addition of
TMSBr (4.7 mL, 35.2 mmol) was made and the mixture was stirred
at r.t. for 24 h. The solvent was removed under reduced pressure,
the resulting residue was taken up with MeOH (10 mL) and stirred
for 2 h at r.t. The evaporation, dissolution procedure was repeated
with MeOH (10 mL) and the solution was stirred for 24 h at
r.t. The solvent was removed under vacuum and the residue was
dissolved in a minimum of MeOH. Addition of Et2O resulted in the
formation of a precipitate, which was collected by centrifugation
and dried under vacuum to afford ligand H8L1 with 81% yield
(450 mg). 1H NMR (D2O, 300 MHz): d 7.96 (t, J = 7.8 Hz, 1 H),
7.50 (d, J = 7.8 Hz, 2 H), 4.95 (s, 4 H), 3.64 (d, J = 12.4 Hz, 8 H).
Column chromatography and flash column chromatography were
performed on silica (0.063–0.200 mm, Merck) or silica gel (40–
63 mm, Merck) or on standardized aluminium oxide (Merck,
Activity II–III). DMF was distilled under reduced pressure. Other
solvents were used as purchased. 1H and 13C NMR spectra
were recorded on Bruker AC 200, Avance 300 and Avance 400
spectrometers operating at 200, 300 or 400 MHz, respectively for
1H. 31P NMR (161.9 MHz) spectra were recorded on an Avance 400
apparatus. Chemical shifts are given in ppm, relative to residual
protiated solvent.36 IR spectra were recorded on a Nicolet 380 FT-
IR spectrometer (Thermo Scientific) as solid samples. Compound
1
13C { H} NMR (D2O, 75 MHz): d 149.4, 139.9, 123.9, 59.3, 52.3
1
14 was obtained according to literature procedures, starting from
(d, J = 137.8 Hz). 31P NMR (D2O, 400 MHz): d 7.98. IR (cm-1,
Dalton Trans., 2010, 39, 9055–9062 | 9059
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ATR): n 2940 to 2580 (w, br, nOH), 1618 (m), 1426 (m), 1160 (s,
methyl orange and with phenolphthalein (Prolabo, purum) as the
indicators. The cell was thermostated at 25.0(2) ◦C by the flow of
a Lauda E200 thermostat. A stream of Argon, pre-saturated with
water vapor, was passed over the surface of the solution. The Glee
program40 was applied for the glass electrode calibration (standard
electrode potential E0/mV and slope of the electrode/mV pH-1)
and to check carbonate levels of the NaOH solutions used (<
5%). The potentiometric data of L and its metal complexes (about
300 points collected over the pH range 2.5–11.5) were refined
with the Hyperquad 200041 program which uses non-linear least-
squares methods.42 Potentiometric data points were weighted by
a formula allowing greater pH errors in the region of an end-
point than elsewhere. The weighting factor Wi is defined as the
n
P–O), 919 (s, nP=O). Anal. Calcd. for C11H23N3O12P4.HBr·2H2O: C,
20.96; H, 4.48; N, 6.66. Found: C, 21.06; H, 4.48; N, 6.11.
Synthesis and characterization of [CuL]Na6·7H2O
To a solution of ligand L (47.7 mg, 0.09 mmol) in H2O (2 mL)
was added CuCl2·2H2O (15.8 mg, 0.09 mmol) dissolved in H2O
(2 mL), resulting in a deep blue solution. The mixture was stirred
for 1 h at r.t. The pH was raised with a diluted aqueous NaOH
solution and the solution was concentrated to ca. 0.5 mL. Addition
of acetone resulted in the formation of a greenish precipitate,
which was collected by centrifugation and dried under reduced
pressure to yield [CuL]Na6·7H2O (52 mg, 91%). Anal. Calcd. for
C11H15CuN3Na6O12P4·7H2O: C, 15.86; H, 3.51; N, 5.05. Found: C,
15.88; H, 3.48; N, 4.94. ESI-/MS (MeOH–H2O): m/z = 572.955
([CuLH5]-, 100%); 595.7 ([CuLH4Na]-, 15%). IR (cm-1, ATR): n
3185 (m, br, nOH), 1644 (m), 1604 (m) 1442 (m), 1117 (s, nP–O), 1037
(s, nP–O), 963 (s, nP=O).
2
reciprocal of the estimated variance of measurements: Wi = 1/si =
1/[sE + (dE/dV)2sV2] where sE (0.1 mV) and sV (0.005 mL)
are the estimated variance of the potential and volume readings,
respectively. The constants were refined by minimizing the error-
square sum, U, of the potentials:
2
2
2
N
2
U =
Wi (Eobs,i − Ecal,i )
∑
Physico–chemical studies
i
Starting materials and solvents
At least three titrations were treated either as single sets or as
separated entities, for each system, without significant variation
in the values of the determined constants. The quality of fit was
judged by the values of the sample standard deviation, S, and
Copper(II) perchlorate (Cu(ClO4)2·6H2O, Fluka, purum p.a.),
zinc(II) perchlorate (Zn(ClO4)2·6H2O, Ventron, Alfa Produkte,
98.9%), nickel(II) perchlorate (Ni(ClO4)2·6H2O, Fluka, purum
p.a.) and gallium(III) nitrate (Ga(NO3)3.xH2O, Alfa Aesar,
2
the goodness of fit, c (Pearson’s test). At sE = 0.1 mV (0.023
R
Puratronicꢀ) are commercial products, which were used without
s
pH) and sV = 0.005 mL, the values of S in different sets of
2
further purification. Distilled water was further purified by passing
it through a mixed bed of ion-exchanger (Bioblock Scientific
R3-83002, M3-83006) and activated carbon (Bioblock Scientific
ORC-83005) and was de-oxygenated by CO2- and O2-free argon
(Sigma Oxiclear cartridge) before use. All the stock solutions were
prepared by weighing solid products using an AG 245 Mettler
Toledo analytical balance (precision 0.01 mg). The ionic strength
was maintained at 0.1 M with sodium perchlorate (NaClO4·H2O,
Merck, ◦p.a.), and all the measurements were carried out at
25.0(2) C. The metal contents of the solutions were ascertained
according to classical colorimetric titrations.38
titrations were between 0.8 and 1.2, and c was below 22. The
scatter of residuals vs. pH was reasonably random, without any
significant systematic trends, thus indicating a good fit of the
experimental data. The successive protonation constants were
calculated from the cumulative constants determined with the
program. The uncertainties in the logK values correspond to
the added standard deviations in the cumulative constants. The
distribution curves of the protonated species of L and its metal
complexes as a function of pH were calculated using the Hyss2009
program.43
CAUTION! Perchlorate salts combined with organic ligands
are potentially explosive and should be handled in small quantities
and with the adequate precautions.39
Spectrophotometric titrations vs. pH
Spectrophotometric titration as a function of pH of the free
ligand L was first performed. Stock solution of L (1.51 ¥
10-4 M) was prepared by quantitative dissolution of a solid
sample in deionised water and the ionic strength was adjusted
to 0.1 M with NaClO4 (Fluka, puriss). An aliquot of 40 mL of the
solution was introduced into a jacketed cell (Metrohm) maintained
at 25.0(2) ◦C (Lauda E200 thermostat). The free hydrogen ion
concentration was measured with a combined glass electrode
(Metrohm 6.0234.500, Long Life) and an automatic titrator
system 794 Basic Titrino (Metrohm). The Ag/AgCl reference
glass electrode was filled with NaCl (0.1 M, Fluka, p.a.) and
was calibrated as a hydrogen concentration probe as described
above. The initial pH was adjusted to ~ 2 with HClO4 (Fluka, ~
70%), and the titration of the free ligand L (2.5 < pH < 11.2)
was then carried out by addition of known volumes of NaOH
solutions (BdH, AnalaR) with an Eppendorf microburette. Special
care was taken to ensure that complete equilibration was attained.
Absorption spectra vs. pH were recorded using a Varian Cary
Potentiometric titrations
The potentiometric titrations of ligand L (2.2–2.5 ¥ 10-3 M) and
its metal complexes ([M]tot/[L]tot ~ 1) were performed using an
automatic titrator system 794 Basic Titrino (Metrohm) with a
combined glass electrode (Metrohm 6.0234.500, Long Life) filled
with 0.1 M NaCl in water and connected to a microcomputer
(Tiamo light 1.2 program for the acquisition of the potentiometric
data). The combined glass electrode was calibrated as a hydrogen
concentration probe by titrating known amounts of perchloric
acid (~ 1.3 ¥ 10-2 M from HClO4, Fluka, ~ 70%) with CO2-
free sodium hydroxide solution (~ 10-1 M from NaOH, BdH,
AnalaR).40 The HClO4 and NaOH solutions were freshly prepared
just before use and titrated with sodium tetraborate decahydrate
(B4Na2O7·10H2O, Fluka, puriss, p.a.) and potassium hydrogen
phthalate (C8H5KO3, Fluka, puriss, p.a.), respectively, using
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50 spectrophotometer fitted with Hellma optical fibers (Hellma,
041.002-UV) and an immersion probe made of quartz suprazil
(Hellma, 661.500-QX). Spectrophotometric titration of the cupric
complexes with L was thereafter carried out. About one equivalent
of Cu(II) perchlorate ([Cu(II)]tot = 1.40 ¥ 10-4 M) was added to
40 mL of L (1.41 ¥ 10-4 M) in a jacketed cell (Metrohm) maintained
Ministry of Higher Education and Scientific Research is also
gratefully acknowledged for the financial support of S.A.
Notes and references
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Conclusions
In conclusion, we have developed a straightforward synthesis of
a new non-macrocyclic ligand which forms very stable complexes
with Cu(II). The kinetics of Cu(II) complexation is fast and the
Cu(II) coordination is very selective for Cu(II) with respect to
Ni(II) and Zn(II), its neighbours in the d series. These coordination
properties thus represent a very promising approach for copper(II)
complexation with high potential within the frame of 64Cu
chelation for PET imaging and radiotherapy. Current efforts are
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Acknowledgements
This work was supported by the Centre National de la Recherche
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9062 | Dalton Trans., 2010, 39, 9055–9062
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The Royal Society of Chemistry 2010
©