Relaxometric, Structural, and Dynamic NMR Studies of DOTA-like Ln(III) Complexes (Ln = La, Gd, Ho, Yb) Containing a p-Nitrophenyl Substituent

Article abstract of DOI:10.1021/ic950981u

We report here the synthesis of the ligand 1-((p-nitrophenyl)carboxymethyl)-4,7,10-tris(carboxymethyl)-1,4,7,10- tetraazacyclododecane (3) and its La(III), Gd(III), Ho(III), and Yb(III) complexes. The introduction of the p-nitrophenyl substituent on the methylenic carbon of one acetate group does not alter the overall chelating ability of 3 with respect to the parent DOTA ligand. The [Gd(3)]^- complex displays a slightly higher relaxivity than that of [Gd(DOTA)]^-, mainly as a consequence of a longer molecular reorientational correlation time (τ_R), due to the increased molecular dimension of the complex, and of limited changes to the other relaxation parameters. A further increase of relaxivity has been observed upon formation of an inclusion compound with β-cyclodextrin. The solution structure and dynamics were thoroughly investigated by high-resolution NMR spectroscopy. Of the four possible enantiomeric pairs that could be present in the solutions of monosubstituted derivatives of [Ln(DOTA)]^- complexes, the proton spectra of Ho and Yb derivatives are consistent with the occurrence of only two isomeric species whose structures have been elucidated through analysis of dipolar shifts and 2D-EXSY data. In both species the bulky aromatic group has replaced the acetate proton pointing outward from the coordination cage. Unlike in the DOTA case, in [Ln(3)]^- complexes the isomerization process involves the inversion of the ethylenic groups of the macrocycle rather than the motion of the acetate arms. This behavior is rationalized in terms of steric crowding at the substituent site: minimization of the steric interactions between the aromatic group and the macrocyclic ring protons results in both structural and dynamic selectivity. Interestingly, in the case of the diamagnetic La(III) complex the variable temperature behavior of the ¹³C NMR spectra is consistent with an exchange process involving one major species and at least one minor isomer of very low concentration, whose relative population increases with temperature. This causes a persistent exchange broadening of the resonances over a wide range of temperatures.

Full text of DOI:10.1021/ic950981u

2726
Inorg. Chem. 1996, 35, 2726-2736
Relaxometric, Structural, and Dynamic NMR Studies of DOTA-like Ln(III) Complexes
(Ln ) La, Gd, Ho, Yb) Containing a p-Nitrophenyl Substituent
Silvio Aime,*,† Mauro Botta,† Giuseppe Ermondi,† Enzo Terreno,† Pier Lucio Anelli,‡
Franco Fedeli,‡ and Fulvio Uggeri‡
Dipartimento di Chimica Inorganica, Chimica Fisica e Chimica dei Materiali, Universita` di Torino,
Via P. Giuria 7, I-10125 Torino, Italy, and Bracco S.p.A., Via E. Folli 50, I-20134 Milano, Italy
ReceiVed July 28, 1995X
We report here the synthesis of the ligand 1-((p-nitrophenyl)carboxymethyl)-4,7,10-tris(carboxymethyl)-1,4,7,-
10-tetraazacyclododecane (3) and its La(III), Gd(III), Ho(III), and Yb(III) complexes. The introduction of the
p-nitrophenyl substituent on the methylenic carbon of one acetate group does not alter the overall chelating ability
of 3 with respect to the parent DOTA ligand. The [Gd(3)]- complex displays a slightly higher relaxivity than
that of [Gd(DOTA)]-, mainly as a consequence of a longer molecular reorientational correlation time (τR), due
to the increased molecular dimension of the complex, and of limited changes to the other relaxation parameters.
A further increase of relaxivity has been observed upon formation of an inclusion compound with â-cyclodextrin.
The solution structure and dynamics were thoroughly investigated by high-resolution NMR spectroscopy. Of the
four possible enantiomeric pairs that could be present in the solutions of monosubstituted derivatives of
[Ln(DOTA)]- complexes, the proton spectra of Ho and Yb derivatives are consistent with the occurrence of only
two isomeric species whose structures have been elucidated through analysis of dipolar shifts and 2D-EXSY
data. In both species the bulky aromatic group has replaced the acetate proton pointing outward from the
coordination cage. Unlike in the DOTA case, in [Ln(3)]- complexes the isomerization process involves the
inversion of the ethylenic groups of the macrocycle rather than the motion of the acetate arms. This behavior is
rationalized in terms of steric crowding at the substituent site: minimization of the steric interactions between the
aromatic group and the macrocyclic ring protons results in both structural and dynamic selectivity. Interestingly,
in the case of the diamagnetic La(III) complex the variable temperature behavior of the 13C NMR spectra is
consistent with an exchange process involving one major species and at least one minor isomer of very low
concentration, whose relative population increases with temperature. This causes a persistent exchange broadening
of the resonances over a wide range of temperatures.
Introduction
linear6 and cyclic7 polyamino carboxylate Gd(III) complexes.
To limit such effects an alternative approach may be pursued
by introducing the substituent on the methylenic carbon of the
acetate group.
Very high thermodynamic and kinetic stability1,2 coupled with
a favorable, long electronic relaxation time3,4 (τS) makes
[Gd(DOTA)]- a superb prototype of a contrast agent (CA) for
magnetic resonance imaging (MRI). Currently the search for
new CAs which display increased selectivity toward organs and
tissues is based, as far as possible, on maintaining these peculiar
properties. Thus, one may envisage a possible route to a new
generation of CAs that involves introducing suitable function-
alities on the surface of a [Gd(DOTA)]- structure. Previous
work dealing with the transformation of an acetate into a
functionalized amide group showed that such a change is
accompanied by a drastic shortening of the electronic relaxation
time of the Gd(III) ion.5 This undesirable feature is due to
changes in the coordination cage when the oxygen of a
carboxylate functionality is substituted by a carboxamido donor
group as shown in other cases involving amide derivatives of
Another point of interest in the study of these DOTA-like
Ln(III) complexes is associated with the assessment of their
solution structures and dynamics. The solid state X-ray
structures determined for DOTA complexes of Eu,8 Gd,9 Y,9,10
Ho,11 and Lu11 have shown that the lanthanide chelates are
isostructural with the octadentate macrocyclic ligand arranged
about the metal ion to give a square antiprismatic coordination
geometry. The twist angle between the two square planes is
about 39°. We have recently shown12 that in solution the
complexes are present in two isomeric forms differing essentially
in the layout of the acetate arms but displaying an identical
staggered conformation of the macrocyclic ring. The coordina-
tion polyhedron of the second isomer can be described as an
“inverted” square antiprism where the twist angle between the
* Corresponding author. Telephone: (+39)-11-6707520. FAX: (+39)-
11-6707524. E-mail: aime@silver.ch.unito.it.
† Universita` di Torino.
(6) Aime, S.; Botta, M.; Fasano, M.; Paoletti, S.; Anelli, P. L.; Uggeri,
F.; Virtuani, M. Inorg. Chem. 1994, 33, 4707.
(7) Aime, S.; Anelli, P. L.; Botta, M.; Fedeli, F.; Grandi, M.; Paoli, P.;
Uggeri, F. Inorg. Chem. 1992, 31, 2422.
(8) Spirlet, M. R.; Rebizant, J.; Desreux, J. F.; Loncin, F. Inorg. Chem.
1984, 23, 359.
‡ Bracco S.p.A.
X Abstract published in AdVance ACS Abstracts, April 1, 1996.
(1) Wang, X.; Jin, T.; Comblin, V.; Lopez-Mut, A.; Merciny, E.; Desreux,
J. F. Inorg. Chem. 1992, 31, 1095.
(2) Toth, E.; Bru¨cher, E.; Lazar, I.; Toth, I. Inorg. Chem. 1994, 33, 4070.
(3) Koenig, S. H.; Brown, R. D., III. Prog. NMR Spectrosc. 1990, 22,
487.
(4) Aime, S.; Botta, M.; Ermondi, G.; Fedeli, F.; Uggeri, F. Inorg. Chem.
1992, 31, 1100.
(9) Chang, C. A.; Francesconi, L. C.; Malley, M. F.; Kumar, K.;
Gougoutas, J. Z.; Tweedle, M. F.; Lee, D. W.; Wilson, L. J. Inorg.
Chem. 1993, 32, 3501.
(10) Parker, D.; Pulukkody, K.; Smith, F. C.; Batsanov, A.; Howard, J. A.
K. J. Chem. Soc., Dalton Trans. 1994, 689.
(5) Sherry, A. D.; Brown, R. D., III; Geraldes, C. F. G. C.; Koenig, S.
H.; Kuan, K.-T.; Spiller, M. Inorg. Chem. 1989, 28, 620.
(11) Aime, S.; Botta, M.; Bombieri, G. Inorg. Chim. Acta, in press.
(12) Aime, S.; Botta, M.; Ermondi, G. Inorg. Chem. 1992, 31, 4291.
S0020-1669(95)00981-5 CCC: $12.00 © 1996 American Chemical Society

DOTA-like Ln(III) Complexes
Inorganic Chemistry, Vol. 35, No. 10, 1996 2727
kg) to give a precipitate which was crystallized several times from H2O.
After drying in Vacuo (KOH, 2 kPa, 50 °C) 2 (285 g; 60%) was obtained
as an orange solid: mp 236 °C (dec); HPLC (method A) retention time,
4.8 min (purity, 98.4%);13C NMR (D2O + NaOD) 45.7, 46.4, 48.1,
50.5, 71.6, 126.6, 133.7, 147.4, 149.7, 160.3.
Chart 1
1-((p-Nitrophenyl)carboxymethyl)-4,7,10-tris(carboxymethyl)-
1,4,7,10-tetraazacyclododecane (3). A suspension of 2 (200.6 g, 0.34
mol) and bromoacetic acid (183.4 g, 1.32 mol) in H2O (500 mL), cooled
to 15-20 °C with an ice-water bath, was adjusted to pH 10 by slow
addition (80 min) of 5 N NaOH (650 mL). The reaction mixture was
maintained at pH 10 (by addition of 5 N NaOH by means of a pH-stat
apparatus) and room temperature for 4 h. Acidification (pH 2.5) of
the reaction mixture with 47% HBr gave a yellow precipitate which
was filtered and washed with H2O (600 mL) and MeOH (2 × 1 L).
The solid was dissolved in 1 N NaOH (550 mL) and the solution
dripped into H2O (500 mL) containing 47% HBr (66 mL). The pH
was adjusted to 2.5 with NaOH and the precipitate then filtered and
washed with H2O (2 × 1.5 L). After drying, the solid (crude 3, 156 g,
86% by HPLC, containing NaBr, 8.4%) was dissolved in H2O (12 L)
and loaded onto a column of Amberlite IR 120 (12 L, H+ form). This
was eluted first with H2O and then with 2.5 N NH4OH. The alkaline
solution was evaporated to dryness and the residue dissolved in H2O
and evaporated. This procedure was repeated three times. The residue
was dissolved in H2O (6 L) and the solution acidified to pH 3.2 by
addition of Amberlite IR 120 resin (200 mL, H+ form). After filtration,
the solution was loaded onto a column of Amberlite XAD 2 (400 mL)
which was eluted with H2O. The eluted solution was concentrated and
dried (P2O5, 2 kPa, 40 °C) to yield 3 (50.3 g; 28%) as an orange solid:
mp 131 °C (dec); HPLC (method B) retention time, 6.5 min (purity,
99%); 13C NMR (D2O + DCl) 44.9, 47.1, 48.9, 50.6, 52.4, 53.0, 53.8,
54.6, 55.4, 56.2, 64.4, 125.6, 132.8, 140.4, 149.1, 169.2, 169.6, 174.1,
175.9.
Complexation Procedure. Ligand 3 (2.1 g, 4 mmol) was suspended
in H2O (20 mL) and solubilized by addition of 2 N NaOH (4 mL). A
solution of lanthanide trichloride (4 mmol) in H2O (10 mL) was added
and the pH slowly adjusted to 6.5 and maintained at this value by
addition of 2 N NaOH using a pH-stat apparatus. After 10 h the
solution was filtered through a 0.45 µm filter and desalted by
electrodialysis. The pH of the solution was adjusted to 6.5 with 1 N
NaOH. The solvent was then evaporated and the residue dried (P2O5,
50 °C, 2 kPa).
Na[La(3)]. Yield 69%. White solid, mp >250 °C; HPLC (method
B) retention time, 31.0 min (purity, 97.8%).
Na[Gd(3)]. Yield 66%. White solid, mp >250 °C; HPLC (method
B) retention time, 24.0 min (purity, 98.4%).
two square planes is about 23°. The substitution of one of the
two nonequivalent acetate protons for a Y-group can be expected
to affect the number of isomeric species in solution and their
interconversion processes.
To investigate all these different aspects we synthesized the
p-nitrophenyl derivative of the DOTA ligand (3; Chart 1), a
lanthanide chelator recently described by Cheng et al.,13 and
its complexes with diamagnetic (La) and paramagnetic (Gd, Ho,
and Yb) ions. Furthermore the NO2 substituent on the para
carbon of the phenyl group represents a potential site for further
modification of the paramagnetic probe. This represents a
means of improving the specificity towards desired target tissues,
a requirement of increasing importance in diagnostic and
therapeutic applications of metal chelates.
Experimental Section
Electrodialyses were carried out with an instrument, model ED 0.004
from Hydro Air Research (Zerbo di Opera, Milan, Italy), fitted with
seven cation- and five anion-exchange STX membranes (total exchange
surface: 0.004 m2) and operated at 12 V. HPLC analyses were
performed on a Merck-Hitachi Lichrograph modular liquid chromato-
graph using the following methods: (A) E. Merck Lichrospher 100
RP-8 column, 5 µm, 250 × 4 mm, thermostated at 40 °C; eluent, 30%
MeCN 70% 0.017 M aqueous H3PO4; flow rate, 1.0 mL/min; UV, 280
nm; (B) E. Merck Lichrospher 100 RP-8 column, 5 µm, 250 × 4 mm,
thermostated at 40 °C; eluent, 5% MeCN 95% 0.017 M aqueous H3-
PO4; flow rate, 1.0 mL/min; UV, 280 nm. 1H NMR and 13C NMR
spectra were recorded on Bruker AC 200, JEOL EX-90, and JEOL
1
EX-400 spectrometers. The H NMR spectrum of 2 is not reported
because the broad and overlapping resonances are of no use for the
structural assignment. Elemental analyses within commonly accepted
limits (H, (0.2%; C, N, Ln, Na, (0.4%) were obtained for all new
compounds.
Organic and inorganic reagents were purchased from E. Merck,
Darmstadt, Germany, and used without further purification. LaCl3,
GdCl3‚6H2O, HoCl3‚6H2O, and YbCl3‚6H2O were obtained from
Aldrich (Milan, Italy).
Na[Ho(3)]. Yield 66%. White solid, mp >250 °C; purity >95%
1
(by H NMR).
Na[Yb(3)]. Yield 66%. White solid, mp >250 °C; HPLC (method
B) retention time, 19.6 min (purity, 98.7%).
NMR Measurements. The variable temperature longitudinal solvent
proton relaxation rates were obtained for 1-2 mM solutions of [Gd(3)]-
on a Stelar Spinmaster spectrometer (Stelar, Mede (PV), Italy) operating
at 20 MHz, by means of the standard inversion-recovery technique
(16 experiments, 4 scans). A typical 90° pulse width was 3.5 µs and
the reproducibility of T1 data was (0.5%. The variable field relaxation
rates were acquired with a field cycling Koenig-Brown relaxometer
(University of Florence, Italy) which operates over a continuum of
magnetic field strengths from 2.5 × 10-4 to 1.4 T (corresponding to
0.01-50 MHz proton Larmor frequencies). Details of the instrument
and of the data acquisition procedure are given elsewhere.15 Samples
for high-resolution measurements were prepared in D2O (99.95%;
Merck, Germany) at 50-80 mM concentrations. A drop of tert-butyl
alcohol was added to the solution in the NMR tube as an internal
chemical shift reference (δH ) 0.0 ppm; δC ) 31.3 ppm).
r-Bromo-4-nitrobenzeneacetic acid (1) was prepared from 4-ni-
trobenzeneacetic acid following the general methodology for R-bro-
mination reported by Harpp et al.14 Caution must be used in handling
the title compound as it is highly vescicating!
r-(p-Nitrophenyl)-1,4,7,10-tetraazacyclododecane-1-acetic Acid
Trishydrobromide (2). A solution of 1 (233.5 g, 0.8 mol) in CH2Cl2
(1.6 L) was dripped over 2 h into a stirred solution of 1,4,7,10-
tetraazacyclododecane (275.2 g, 1.6 mol) in H2O (1.6 L) maintained
at 15-20 °C in an ice-water bath. After being stirred for 2 h at 20 °C
the organic phase was separated. The aqueous phase was washed with
CH2Cl2 (2 × 100 mL) and acidified with 47% HBr (320 mL) to give
an amorphous precipitate. This was decanted and the aqueous phase
washed with CH2Cl2 (3 × 150 mL) and concentrated (to 1 kg).
Addition of EtOH (2.5 L) led to the precipitation of a white solid
(1,4,7,10-tetraazacyclododecane trihydrobromide, 197 g). The solution
was concentrated to eliminate EtOH and the residue diluted with H2O
(to 1 kg). Acidification with 47% HBr (750 mL) gave a precipitate
which was filtered and dissolved in H2O (2.5 L). A small amount of
insoluble solid was filtered off. The solution was concentrated (to 1
Results and Discussion
Synthesis of Ligand 3 and [Ln(3)]- Complexes. Ligand 3
was synthesized according to Scheme 1. The procedure for
(13) Cheng, R.; et al. Dow Chemical Co. WO 89/12631; 28 Dec 1989.
(14) Harpp, D. N.; Bao, L. Q.; Black, C. J.; Gleason, J. G.; Smith, R. A.
J. Org. Chem. 1975, 40, 3420.
(15) Koenig, S. H.; Brown, R. D., III. NMR Spectroscopy of Cells and
Organisms; Gupta, R. K., Ed.; CRC Press: Boca Raton, 1987; Vol.
II.

2728 Inorganic Chemistry, Vol. 35, No. 10, 1996
Aime et al.
Scheme 1a
a
(i) H2O-CH2Cl2, room temperature. (ii) BrCH2COONa, H2O (pH 10), room temperature (iii) LnCl3, H2O (pH 6.95).
R-bromination of carboxylic acids, as reported by Harpp et al.,14
was used for the preparation of R-bromo-4-nitrobenzeneacetic
acid (1). Monoalkylation of 1,4,7,10-tetraazacyclododecane
with 1 was achieved in fairly good yield using a 100% excess
of the macrocycle in a two-phase (CH2Cl2-H2O) reaction
medium. Carboxymethylation of 2 with bromoacetic acid in
water at pH 10 gave ligand 3.
Sodium salts of the LnIII(3) complexes were prepared in H2O
at pH 6.5 (by adding aqueous NaOH) from 3 and the appropriate
LnIIICl3. The excess NaCl was eliminated by electrodialysis.
Ligand 3 has previously been obtained at Dow by a procedure
involving the stepwise alkylation of 1,4,7,10-tetraazacyclodode-
cane with R-bromoesters.13 More recently the same group has
reported that the crucial monoalkylation of 1,4,7,10-tetraaza-
cyclododecane can be achieved in very good yield if sufficiently
hindered R-bromoesters are used as electrophiles in a 1:1
stoichiometric ratio in CHCl3.16 In the same paper the crystal
structure of a ligand closely related to 2 is also reported.
NMR Characterization. In D2O, at 25 °C and pD ) 7.5,
1
the 400 MHz H NMR spectrum of 3 consists of three well-
defined resonances at 8.3 ppm (d, JH-H ) 8.6 Hz, 2), 7.8 ppm
(d, JH-H ) 8.6 Hz, 2), and 5.1 ppm (broadened singlet, 1),
corresponding respectively to the meta, ortho, and methylenic
protons of the p-nitrophenyl acetate group. In addition there
are a high number of ill-defined resonances between 4.2 and
2.6 ppm which are attributable to the 22 protons of the 3 acetate
and 4 ethylenediamine groups. As the temperature is increased,
Figure 1. Variable temperature 13C NMR spectra of ligand 3 in D2O
(pD ) 7.5).
an overall reduction and sharpening of the resonances in this
latter region suggest that a dynamic rearrangement is taking
place on the NMR time-scale. Elucidation of this process is
ppm are detected. The two higher field resonances can easily
be assigned to carboxylate groups in which the negative charge
is partially removed through the formation of hydrogen bonds
with positively charged nitrogen atoms of the macrocyclic ring.
In fact, although we have not carried out protonation studies
on this ligand, we may reasonably assume that at pH values
close to neutrality two of the nitrogen atoms are in the protonated
form, as found in other related tetraazamacrocyclic ligands.17
better tackled on the basis of the variable temperature (VT) 13
C
NMR spectra (Figure 1), in particular by looking at the region
of the carboxylate absorption. At 275 K (and up to room
temperature) four resonances at 178.6, 177.6, 170.9, and 168.7
(16) Kruper, W. J., Jr.; Rudolf, P. R.; Langhoff, C. A. J. Org. Chem. 1993,
58, 3869.

DOTA-like Ln(III) Complexes
Inorganic Chemistry, Vol. 35, No. 10, 1996 2729
Thus, the intramolecular interaction of two carboxylate anions
with two NH+ centers results in an overall stereochemical
rigidity which in turn leads to a low- symmetry solution
structure, as manifested by the inequivalence of both the proton
and carbon nuclei in the low-temperature NMR spectra.
As the temperature is increased the two carboxylate reso-
nances at 170.9 and 168.7 ppm broaden, collapse, and then
merge into a single resonance at 170.2 ppm. From the chemical
shift difference of the two peaks in the low-temperature limiting
spectrum and the coalescence temperature we may estimate18
the free energy of activation for this dynamic process to be
∆Gq318 ) 61.5 ( 1 kJ mol-1. Analogous behavior is observed
in the methylenic region in which the high-temperature 13C
NMR limiting spectrum is consistent with the presence of a
mirror plane in the molecule. This may be accounted for in
terms of a fast proton exchange over the four nitrogen basic
sites of the macrocycle accompanied by a free rotation of the
side arms.
Relaxometric Studies of the [Gd(3)]- Complex. A 1 mM
aqueous solution of [Gd(3)]- at 25 °C, pH 7.3, and a proton
Larmor frequency of 20 MHz shows a relaxivity of 5.4 mM-1
s-1, a value clearly higher than that of the parent compound
[Gd(DOTA)]- (4.7 mM-1 s-1). The relaxivity at this frequency,
which corresponds to a magnetic field strength within the range
of values routinely employed in MRI applications, is normally
taken as a direct indication of the ability of a paramagnetic
complex to enhance the water proton relaxation rates. There-
fore, these data indicate that the chemical modification of one
side arm of the DOTA ligand not only does not compromise
the relaxation efficiency of the corresponding Gd(III) complex
but, on the contrary, it promotes roughly a 15% increase in
relaxivity. This result is not surprising if compared with what
is known about the magnetic relaxation properties of Gd(III)
complexes with both linear and macrocyclic polyamino poly-
carboxylic ligands.3 The relaxivity enhancement of the solvent
arises from random fluctuations in time of the magnetic dipolar
coupling interaction between the unpaired electrons of the Gd-
(III) ion and the water proton nuclei.19 This electron-nucleus
coupling can be conveniently described as taking place through
two distinct mechanisms: (i) inner sphere, due to the water
molecules present, in number q, in the coordination sites of the
metal ion, and (ii) outer sphere, which involves all the
nonbonded water molecules surrounding the complex at a given
time (normally, second sphere solvent molecules are not
explicitly considered). Modulation of the dipolar interaction,
which is responsible for the nuclear spin solvent relaxation,
arises from (i) chemical exchange (τM is the corresponding
correlation time), reorientation of the paramagnetic complex
(τR), and electron spin relaxation (τS) and from (ii) translational
diffusion (τD) and electron spin relaxation. Moreover, both
contributions have a pronounced magnetic field dependence.3
The data accumulated over the last 10 years allow some general
guidelines to be drawn. At the imaging fields normally adopted
and at 25 °C the outer sphere water molecules are responsible
for a relaxivity contribution of ∼2.0 mM-1 s-1, which may
account for up to 50% of the total relaxivity for complexes
characterized by a q value of 1, as is normally the case for
compounds such as MRI CAs. The outer sphere component is
thus expected to be quite similar for complexes of similar
Figure 2. Temperature dependence of the relaxivity for [Gd(3)]- in
H2O at 20 MHz and pH ) 7.2.
molecular size, as evidenced from its direct dependence from
τD ) a2/D, where a is the minimum distance between the
paramagnetic center and the diffusing solvent molecules with
values in the range 3.6-5.0 Å3,4,20 and D is the sum of the
diffusion constants for the solvent and the metal complex (D
= (2.0-3.0) × 10-5 cm2 s-2 at 25 °C).3,4,6,20 Since in the high-
field region τS plays a negligible role the observed differences
in relaxivity can hardly be ascribed to this contribution. The
inner sphere component has a value of ∼2.5-3.5 mM-1 s-1
,
for small Gd(III) chelates with q ) 1 at 20 MHz and 25 °C,
depending on the value of the ratio τR/r6, where r is the distance
between the metal ion and the protons of the coordinated water
molecules, within the range 3.0-3.2 Å. In principle, this
relaxation mechanism could be influenced by the rate of the
chemical exchange of water from the coordination site to the
bulk. If the exchange rate is slow with respect to the relaxation
rate (1/T1M) of the bound water protons, then the relaxation
enhancement is partially quenched, as recently observed for a
Gd(III) complex with a diamide derivative of DTPA.6,21
However, this effect is generally not observed at frequencies
of 20 MHz or above, near room temperature. Often the
condition of fast exchange is rather arbitrarily assumed, even if
it could easily be checked by looking at the temperature
dependence of the relaxivity. In fact the paramagnetic contribu-
tion of the solvent longitudinal relaxation rate is given by the
following equation:
Nq
1
R1p
)
(1)
55.6 T1M + τM
where N is the molar concentration of the metal complex.
Because of the opposite temperature dependences of T1M and
τM two limiting cases can be envisaged: (i) fast exchange (T1M
. τM); R1p increases by decreasing the temperature and (ii) slow
exchange (T1M , τM); R1p decreases by decreasing the tem-
perature. The temperature dependence of the relaxivity of
[Gd(3)]- has been investigated between 1 and 70 °C at 20 MHz
and pH ) 6.8 and clearly shows (Figure 2) a monoexponential
decrease of R1p with T, as expected for the fast exchange
condition. In conclusion, the measured relaxivity value of 5.4
mM-1 s-1 is likely to be a consequence of a longer τR value
due to the increased molecular dimension of the complex with
consequent decreased tumbling rate, thus implicitly indicating
the presence of a single coordinated water molecule. Further
(17) (a) Desreux, J. F.; Merciny, E.; Loncin, M. F. Inorg. Chem. 1981, 20,
987. (b) Ascenso, J. R.; Delgado, R.; Frau`sto da Silva, J. J. R. J. Chem.
Soc., Perkin Trans. 2 1985, 781.
(18) Friebolin, H. Basic One- and Two-Dimensional NMR Spectroscopy,
2nd ed.; VCH: Weinheim, Germany, 1993; pp 293-296.
(19) Banci, L.; Bertini, I.; Luchinat, C. Nuclear and Electron Relaxation;
VCH: Weinheim, Germany, 1991.
(20) (a) Aime, S.; Batsanov, A. S.; Botta, M.; Howard, J. A. K.; Parker,
D.; Senanayake, K.; Williams, G. Inorg. Chem. 1994, 33, 4696. (b)
Hernandez, G.; Brittain, H. G.; Tweedle, M. F.; Bryant, R. G. Inorg.
Chem. 1990, 29, 985.
(21) Gonza`les, G.; Powell, D. H.; Tissie`res, V.; Merbach, A. E. J. Phys.
Chem. 1994, 98, 53.

2730 Inorganic Chemistry, Vol. 35, No. 10, 1996
Aime et al.
Table 1. Best Fitting Parameters for the NMRD Profiles of Figure
3a
temperature (°C)
parameter
S0 (ps)
10
25
39
τ
271.0(6)
15.0(2)
113.0(3)
282.0(20)
1.56(0.1)
420.0(10)
7.4(1)
81.0(2)
100.0(20)
2.40(0.2)
497.0(12)
4.9(1)
58.0(2)
81.0(30)
3.15(0.2)
τV (ps)
τR (ps)
τ
M (ns)
D (cm2 s-1) × 105
a Numbers in parentheses represent standard deviations in mean
parameter estimates on 1000 simulated relaxivity data sets obtained
by introducing repeatedly into the experimental data set random errors
of 1% and estimating best parameters.
Figure 3. 1/T1 NMRD profiles of a 1 mM aqueous solution of
[Gd(3)]-. The solid curves through the data points were calculated with
the parameters of Table 1. The lower curves represent the outer sphere
contribution to proton relaxivity.
methylenic resonances in the case of the diamagnetic La(III)
complex, using a well-established procedure.25,26 By introducing
these values into the paramagnetic equations and by using τM,
τS0, τV, and r as adjustable parameters, the profiles at 25 °C
could be fitted satisfactorily. For the water exchange lifetime
support to the hypothesis of q ) 1 comes from the comparison
with published data. Tweedle et al.22 have plotted the relaxivity
values at 40 °C vs the hydration number q, as determined by
luminescence studies, for a series of amino carboxylato Gd(III)
complexes of similar size. The plot gave a straight line with
an intercept of 2.0 ( 0.3 mM-1 s-1 (corresponding to the outer
sphere component) and a slope of 1.7 ( 0.1 mM-1 s-1 per
coordinated water molecule (corresponding to the inner sphere
contribution). We found for [Gd(3)]-, at the same temperature,
a relaxivity of 4.1 mM-1 s-1, a value which clearly indicates
the presence of one water molecule in the inner coordination
sphere of the complex. Therefore, the introduction of a phenyl
group on the methylenic carbon of an acetate arm of the DOTA
ligand does not seem to have a negative effect on its potential
coordinating ability.
These qualitative observations can be validated by a more
thorough analysis of the magnetic field dependence, i.e., by
measuring the NMRD (nuclear magnetic relaxation dispersion)
profiles.3,19 This technique allows the full exploitation of the
large information content of paramagnetic relaxation. We have
measured the NMRD profiles for [Gd(3)]- at 10, 25, and 39
°C (Figure 3) on the field-cycling relaxometer developed by
Koenig and Brown, over a wide range of magnetic fields
covering the proton Larmor frequencies from 0.01 to 50 MHz.
The experimental data can be fitted by the equations describing
the inner (Solomon-Bloembergen-Morgan equations19) and
outer sphere (Freed equation)23 relaxation mechanisms to give
the following set of structural and dynamic relaxation param-
eters: q, r, a, D, τR, τM, τS0, τV. The latter two parameters can
be entered into the Bloembergen and Morgan equation that
describes the field dependence of τS and represent its zero field
value and the correlation time for the collision of the complex
with the solvent molecules, respectively. The number of
parameters is too large to give a reliable set of best-fitting values,
and thus some of them have to be estimated or derived from
other data. According to the discussion above the hydration
number q was taken as equal to 1; a and D were given values
of 3.6 Å and 2.4 × 10-5 cm2 s-1 (at 25 °C), respectively, which
were obtained from the analysis of NMRD profiles for the “outer
sphere” complex [Gd(TETA)]-;24 a value of 80 ( 2 ps for the
reorientational correlation time τR was calculated at 25 °C by
measuring the 13C relaxation rates and NOE factors of some
τ
M an initial value of 0.2 µs was tentatively assigned, since this
was recently calculated by the 17O-NMR technique for DOTA
and DTPA complexes.27 The fitting results have a rather limited
depencence from the value of this parameter, despite a better
agreement with the experimental data being obtained for a τM
of 0.1 µs. Finally, when τR was also used as a variable
parameter the best value obtained was 81 ps. The profiles at
10 and 39 °C were fitted by using the value of 3.02 Å calculated
for r at 25 °C and by using τR as an adjustable parameter. The
best-fitting parameters are reported in Table 1, and the corre-
sponding calculated profiles are shown as continuous lines
through the data points in Figure 3. The parameters show the
expected temperature dependence: τR,τM, and τV increase with
decreasing temperature, while D and τS0 show the opposite
behavior. The mean residence lifetime of the coordinated water
molecule is much better determined from the data taken at 10
°C since the condition of fast exchange, at least in the low
magnetic field region, no longer holds. This results in partial
quenching of the relaxivity at low fields, as revealed by a
decrease in the ratio of the relaxivities at 0.01 and 20 MHz
which passes from 2.4 at 25 °C to 2.1 at 10 °C. In each of the
three profiles it is clear that, at any frequency, the inner sphere
relaxivity is largely controlled by the reorientational correlation
time of the complex whose contribution to τC ranges from 71%
(0.01 MHz, 10 °C) to 97% (50 MHz, 39 °C). This is well-
known for [Gd(DOTA)]- where it has been attributed to an
unusually long electronic relaxation time. However, for [Gd(3)]-
it is not obvious since the chemical modification of a carboxylic
group into a carboxamide has previously been shown to be
accompanied by a drastic reduction of τS0 and therefore of the
low-field relaxivity.5,7 Here the introduction of a phenyl
substituent, without changing the type of donor groups of the
ligand, has only a limited effect on τS0, and as a consequence
the relaxivity of [Gd(3)]- is higher than the relaxivity of
[Gd(DOTA)]- whatever the magnetic field strength.
As discussed above, one of the main aims in the preparation
of [Gd(3)]- was the assessment of the ability of a functionalized
paramagnetic carrier to interact with a number of macromo-
lecular substrates, in both covalent and noncovalent modes. A
pronounced relaxivity (more exactly the inner sphere compo-
nent) enhancement is expected as a result of the remarkable
lengthening of the reorientational correlation time for the bound
form. In this regard, the noncovalent binding is particularly
attractive for it ensures both the integrity of the complex and
(22) Zhang, X.; Chang, C. A.; Brittain, H. G.; Garrison, J. M.; Telser, J.;
Tweedle, M. F. Inorg. Chem. 1992, 31, 5597.
(23) Freed, J. J. Chem. Phys. 1978, 68, 4034.
(24) Aime, S.; Botta, M. Unpublished data.
(25) Aime, S.; Botta, M.; Ermondi, G. J. Magn. Reson. 1991, 92, 572.
(26) Kim, W. D.; Kiefer, G. E.; Maton, F.; McMillan, K.; Muller, R. N.;
Sherry, A. D. Inorg. Chem. 1995, 34, 2233.
(27) Micskei, K.; Helm, L.; Bru¨cher, E.; Merbach, A. E. Inorg. Chem. 1993,
32, 3844.

DOTA-like Ln(III) Complexes
Inorganic Chemistry, Vol. 35, No. 10, 1996 2731
Figure 4. Plot of the proton relaxation enhancement for [Gd(3)]-, at
20 MHz and 25 °C, as a function of â-cyclodextrin concentration. The
solid curve was calculated with the binding parameters (KD and ꢀb)
given in the text.
its stability. We tested the ability of [Gd(3)]- to interact with
the hydrophobic cavity of â-cyclodextrin (â-CD) which was
previously found to act as a good model system by forming
inclusion complexes of relatively high stability with suitably
functionalized Gd(III) complexes.28 The measurement of water
proton relaxation times, at a fixed frequency, provides an easy
and accurate tool for determining the strength of the interaction
and the extent of the relative relaxation enhancement (ꢀb) of
the inclusion complex. A titration of a 0.5 mM aqueous solution
of the [Gd(3)]- complex with â-CD at pH 7.4 and 25 °C (Figure
4) allowed us to determine a value of 1.7 × 10-2 M for the
dissociation constant KD for the equilibrium
Figure 5. Schematic representation of the two enantiomeric pairs for
the [Ln(DOTA)]- complexes (Y ) H) and of the four isomers for the
[Ln(3)]- complexes (Y ) C6H5NO2) in the case of an R configuration
of the chiral carbon atom. Labels ac, a, and e identify acetate, axial,
and equatorial protons respectively.
temperatures of the two distinct resonances for the ethylenic
carbons.12,30,31 On the other hand, the exchange between the
two isomeric forms of the complexes may take place either by
rotation of the acetate arms while maintaining the same
conformation of the macrocylic ring (A T B; C T D) or by
inversion of the ethylenic groups without changing the orienta-
tion of the acetate arms (A T C; B T D).12,31,32
{[Gd(3)]-/â-CD} f [Gd(3)] + â-CD
This KD value is almost 1 order of magnitude higher than
corresponding values obtained for related linear and cyclic
polyamino carboxylate Gd(III) complexes containing one or
more (benzyloxy)methyl substituents.28 As expected from the
similar size of the two types of inclusion compounds, their
relaxivity enhancements are very similar (ꢀb = 2). The larger
KD value of [Gd(3)]- is presumably a consequence of a poorer
fit within the â-CD cavity compared with complexes containing
the CH2-O-CH2-C6H5 substituent. In fact, the presence of
a nitro group in the para position is expected to strengthen the
binding and favor the inclusion complex formation, as found
in the case of monosubstituted benzenes.29 Thus the steric
hindrance due to the direct attachment of the aromatic group to
the substituted carbon appears to play a major role in controlling
the strength of the interaction. Tentatively, we may imagine
that the steric crowding does not allow an efficient inclusion
of the aromatic ring inside the cyclodextrin cavity and thus the
fully exploitation of the hydrophobic interaction.
The substitution of an acetate proton with a group Y (in our
case Y ) p-nitrophenyl) introduces a chiral center in the ligand
and a site of asymmetry in the complexes which doubles the
number of possible isomeric species in solution. Unlike in the
case of [Ln(DOTA)]- complexes, A and D (and B and C) are
no longer enantiomers but diasteromers and are therefore
characterized by distinct resonances in their NMR spectra. If
the macrocyclic ligand is a racemic mixture, we need to indicate
the configuration (R or S) of the chiral carbon atom of the ligand
along with the four arrangements (A-D) of the ligand itself
about the metal ion in order to identify completely all the
possible stereoisomers. Thus, in the solutions of Ln(III)
complexes with monosubstituted derivatives of DOTA we might
expect up to eight stereoisomers of the type XW (X ) A-D; W
) R, S). It is worth noting that the mirror image of the isomer
A still corresponds to the structural model D, that the enantio-
meric form of B corresponds to the structural model C and vice
versa, and therefore that the eight stereoisomers constist of four
enantiomeric pairs (Figure 6) able to mutually interconvert.
Finally, the lack of axial symmetry removes the signal
degeneracy found in the NMR spectra of the [Ln(DOTA)]-
complexes and therefore a large number of resonances are
expected for each isomer in the proton (27) and carbon (22)
spectra.
Solution Structure and Dynamics of [Ln(3)]- Complexes
(Ln ) La, Ho, and Yb). A. General Considerations. The
basic structural and dynamic features characterizing the aqueous
solutions of [Ln(DOTA)]- complexes are schematically sum-
marized and graphically represented in Figure 5, where A and
D represent the square antiprismatic enantiomers and B and C
the “inverted” square antiprismatic enantiomeric geometries. The
chirality arises from the two opposite elicities in which the
achiral ligand can wrap around the metal ion. Interconversion
within each enantiomeric pair occurs through inversion of the
ethylenic groups and concerted or stepwise rotation of the acetate
arms: this process is easily and clearly observed in the VT 13C-
NMR spectra since it results in the equilibration at high
[La(3)]-. The 139La NMR spectrum, recorded at 70 °C,
shows a single, broad resonance centered at 320 ( 5 ppm. This
value is very close to that reported for [La(DOTA)]- and fully
consistent with the empirical rules which ascribe a downfield
shift, with respect to the resonance for the aquoion, of 50 and
30 ppm for each coordinated nitrogen and oxygen atom,
respectively.33 This clearly indicates that the substitution does
(28) Aime, S.; Botta, M.; Panero, M.; Grandi, M.; Uggeri, F. Magn. Reson.
Chem. 1991, 29, 923.
(29) Bender, M. L.; Komiyama, M. Cyclodextrin Chemistry; Springer-
Verlag: Berlin, 1978; Chapter 3.
(30) Desreux, J. F. Inorg. Chem. 1980, 19, 1319.
(31) Hoeft, S.; Roth, K. Chem. Ber. 1993, 126, 869.
(32) Vincent, J.; Desreux, J. F. Inorg. Chem. 1994, 33, 4048.
(33) Geraldes, C. F. G. C.; Sherry, A. D. J. Magn. Reson. 1986, 66, 274.

2732 Inorganic Chemistry, Vol. 35, No. 10, 1996
Aime et al.
the chemical shift difference between the high-field carboxylate
peaks increases by 0.3 ppm on going from 0 to 90 °C.
1
The set of H NMR spectra essentially reproduce the same
basic features and do not add further information. This unusual
dynamic behavior can be accounted for by an exchange process
involving one major species and one (or more) minor isomer
in very low concentration at low temperatures but whose
population increases with temperature (endothermic isomeriza-
tion process). Support for this hypothesis comes from the
analysis of the paramagnetic complexes [Ln(DOTA)]- (Ln )
Eu-Yb) that show in their proton NMR spectra two distinct
sets of resonances corresponding to two isomeric species. In
this case the integration of corresponding pairs of resonances
yields a direct estimation of the equilibrium constant for the
isomerization process and from its temperature dependence it
is evident that the major f minor isomer transformation
represents an endothermic process.34 Finally, it may be noted
that the three ethylenic resonances at lower field show a very
limited temperature dependence, both in terms of line broadening
and chemical shift variation. This can be interpreted as the result
of an exchange process between peaks with a small frequency
separation and thus characterized by a lower coalescence
temperature.
Figure 6. Symbolic representation of the eight possible stereoisomers
for [Ln(3)]- complexes. The labels A-D refer to the coordination
geometries of Figure 5. Labels R and S indicate the configuration of
the chiral carbon atom of the ligand. The lines connect pairs of
enantiomers.
[Ho(3)]-. The remarkable stereochemical rigidity normally
encountered with lanthanide(III) complexes of 12-membered
tetraazamacrocyclic rings often allows good high-resolution
proton NMR spectra to be obtained in the case of highly
paramagnetic ions such as Tb, Dy, and Ho. The advantage is
that the extremely large chemical shift range (up to 1000 ppm!)
characteristic of the spectra of these complexes permits the
detection of well-separated peaks for all the nonequivalent
protons of the ligand and, in fortunate cases, separate sets of
resonances for the different isomeric species present in solution.
The 1H NMR spectrum of [Ho(3)]- in D2O is shown in Figure
8. The spectrum was recorded for a 0.08 M solution at 25 °C
and at 90 MHz since, to minimize severe magnetic field-induced
line broadening,35 it is preferable to operate at low magnetic
field strengths. Twenty-two main resonances and a series of
other low-intensity peaks cover a range of about 470 ppm. As
mentioned above, the lack of a C4 symmetry axis removes the
signal degeneracy found in the spectra of DOTA derivatives
and therefore each peak of [Ho(DOTA)]- is split into a set of
four resonances. Thus, in the absence of any fluxional process,
we expect to observe 27 distinct signals. On this basis the partial
assignment of the spectrum in Figure 8 is possible. Apart from
casual overlapping of some resonances (two equatorial protons
at -66.0 ppm, one axial and one acetic proton at 62.4 ppm,
and two acetic protons at 178.5 ppm), only two peaks for the
ortho and meta protons are present at 43.9 and 17.3 ppm,
respectively, to indicate the occurrence of a freely rotating
aromatic group. We may note, by analogy with DOTA
derivatives,35 that these resonances belong to a species corre-
sponding to a square antiprismatic coordination geometry, i.e.,
to structural models A or D of Figure 5.
Figure 7. Variable temperature 13C NMR spectra (carbonyl and
aliphatic regions only) of [La(3)]- in D2O (pD ) 7.5) at 9.4 T.
not alter the potential octadenticity of the macrocyclic ligand,
in full agreement with the relaxometric data. Interestingly, the
bandwidth at half-height for [La(3)]- (∆ν1/2 ) 4950 Hz) is larger
than that for [La(DOTA)]- (∆ν1/2 ) 3120 Hz),33 and this
directly reflects differences in the transverse relaxation rates
which are largely dominated by the quadrupolar mechanism.
On this basis such a difference can be easily accounted for by
the slightly different values of the corresponding reorientational
correlation times and the higher degree of asymmetry for
[La(3)]-. On the other hand, a broader resonance could also
arise from the presence in solution of isomeric species origi-
nating NMR signals with small differences in their chemical
shift values.
However, the low-temperature limiting 13C NMR spectrum
(Figure 7a) shows 12 narrow, distinct peaks in the aliphatic
region, corresponding to the 12 different aliphatic carbon atoms
of the ligand, thus indicating the presence of just one of the
four possible structural isomers. Comparison with the analogous
spectrum of the DOTA derivative and a heterocorrelated 2D-
COSY experiment allows a rather straightforward assignment
of the resonances: carboxylate at 180.7(1), 180.6(2), and 178.9-
(1) ppm, aromatic at 148.6(1), 139.7(1), 134.2(2), and 124.3
(2) ppm, acetate at 70.0 (CH), 61.2, 61.0, and 60.9 ppm, and
ethylenic at 54.7, 54.5, 54.1, 50.7, 50.0, 49.7, 48.6, and 46.2
ppm. Upon increasing the temperature an overall broadening
of all the resonances, particularly marked for those correspond-
ing to the ethylenic ring carbons, is observed over the range
10-40 °C, followed by their sharpening up at higher temper-
atures (Figure 7). No change in the number of 13C resonances
is detected as the process takes place, whereas a differential
temperature-dependent behavior is shown by their chemical shift
In addition to the main resonances another set of low-intensity
signals covering a range of about 300 ppm (from -170 to 125
ppm) is clearly detectable and most likely attributable to the
presence of a minor isomeric species. Again, comparison with
the spectra of DOTA complexes enables the assignment of some
of the resonances to an isomer characterized by an “inverted”
square antiprismatic structure of type B or C. Particularly
diagnostic in this regard are the four peaks in the -170 to -125
(34) Aime, S.; et al. Manuscript in preparation.
(35) Aime, S.; Barbero, L.; Botta, M.; Ermondi, G. J. Chem. Soc., Dalton
Trans. 1992, 225.
t
values (relative to the signal of BuOH to which a δ ) 31.3
ppm was assigned throughout each experiment). For example

DOTA-like Ln(III) Complexes
Inorganic Chemistry, Vol. 35, No. 10, 1996 2733
1
Figure 8. 90 MHz H NMR spectrum for a 0.08 M solution of [Ho(3)]- in D2O at pD ) 7.2 and 25 °C. The labels w and r indicate HDO and
ButOH (internal reference) peaks, respectively. The resonances of the major isomer have been labeled according to the scheme of Figure 5.
ppm range corresponding to four axial protons of the macro-
cyclic ring. The two isomers have at 25 °C a relative population
of 88:12, as determined by integration of corresponding sets of
axial peaks at high field, and represent one of the four possible
pairs A/B, A/C, D/B and D/C. Further insight into the solution
structures of these species can be gained by considering the
acetate proton resonances in detail. In [Ln(DOTA)]- complexes
two distinct resonances, ac1 and ac2, are detected for the acetate
protons for each isomer in the NMR limiting spectrum at low
temperature. The two signals differ in their chemical shift
values as a consequence of the different location of the protons
with respect to the symmetry axis of the complex. They also
present rather different bandwidths which reflect differences in
the metal-proton distances.12,35 For the main isomer (enan-
tiomeric pair A/D) of [Ho(DOTA)]- the “broad” resonance
(associated with the proton at a shorter distance) falls at 160.7
ppm and the “narrow” peak of ac1 at 52.4 ppm. With reference
to the schematic models of Figure 5 the “broad” resonance
corresponds to the acetate proton close to the macrocyclic ring
protons. By increasing the temperature the two signals ac1 and
ac2 mutually exchange as a result of the enantiomerization
processes of the type A T D and B T C. In the monosubsti-
tuted derivatives of DOTA the acetate protons give seven distinct
resonances in the NMR spectra, divided in two subsets of four
and three peaks. The substituent may replace either an ac1
proton to give enantiomeric pair AR/DS (CR/BS) or an ac2 proton
to give the other enantiomeric pair DR/AS (BR/CS). In the first
case we would expect to observe a group of four “broad”
resonances in the spectrum at low field and a group of three
“narrow” resonances at higher field, whereas the opposite
situation would occur in the second case. In the spectrum of
[Ho(3)]- we clearly observe four “broad” signals in the range
160-185 ppm and three “narrow” peaks in the range 50-65
ppm. This indicates that for this complex the main isomer in
aqueous solution corresponds to the enantiomeric pair AR/DS.
Unfortunately, the acetate signals for the minor isomer are buried
under more intense peaks and no inference can be made.
A final comment should be made concerning the close
correspondence observed between the proton chemical shift
values for the resonances of [Ho(DOTA)]- and the average
values for the related sets of four signals for [Ho(3)]-. Since
the paramagnetic shift is given by the sum of both contact
(through bond) and dipolar (through space) contributions,36 the
close analogy between the 1H NMR spectra of the two
complexes means that (i) a very similar unpaired electron
delocalization exists in both compounds. This is to be expected
because the donor atoms involved in the coordination of the
metal ion are the same. (ii) The proton nuclei have quite similar
geometrical coordinates. (iii) the principal magnetic susceptibil-
ity axis for [Ho(3)]- deviates only slightly from the axis
perpendicular to the plane of the four coordinated oxygens.
These latter two points can be further confirmed through
quantitative analysis of the dipolar shifts which is better
accomplished for the Yb derivative for which the contact shift
has the smallest theoretical contribution.36,37
1
[Yb(3)]-. The H NMR spectrum of this complex (Figure
9) clearly reveals the presence of two isomers in solution at a
ratio of 3:1, which roughly corresponds to the isomeric ratio
found for [Yb(DOTA)]-12,31 and confirms the increase of the
population of the minor isomer along the lanthanide series. The
(36) Sherry, A. D.; Geraldes, C. F. G. C. In Lanthanide Probes in Life,
Chemical and Earth Sciences: Theory and Practice; Bu¨nzli, J.-C. G.,
Choppin, G. R., Eds.; Elsevier: Amsterdam, 1989.
(37) Golding, R. M.; Halton, M. P. Aust. J. Chem. 1972, 25, 2577.

2734 Inorganic Chemistry, Vol. 35, No. 10, 1996
Aime et al.
Table 2. Calculated and Experimental Protona NMR Shifts for the
AR/DS Enantiomeric Pair of [Yb(3)]- at 5 °C and Best Fitting
Parameters
proton
δνcalc
δνsper
proton
δνcalc
δνsper
H1
H2
H3
H4
H5
H6
H7
H8
H9
H10
H11
H12
160.5
155.2
151.0
146.4
30.5
29.7
30.1
25.1
24.4
22.0
19.1
18.1
157.3
154.3
157.3
142.2
29.4
29.0
25.3
23.0
19.7
19.4
17.8
15.7
H13
H14
H15
H16
H17
H18
H19
H20
H21
H22
H23
-43.0
-45.0
-45.5
-46.0
-52.5
-54.8
-62.5
-94.6
-95.5
-101.0
-104.0
-48.1
-44.9
-45.2
-50.5
-58.1
-58.1
-62.6
-98.9
-100.2
-101.2
-111.9
D1
5254
D2
-250
R
0.045
a Labeled according to Figure 10.
Figure 9. 90 MHz 1H NMR spectrum for a 0.08 M solution of [Yb(3)]-
in D2O at pD ) 7.2 and 25 °C. The labels w and r indicate HDO and
ButOH (internal reference) peaks, respectively. Assignment of the
resonances for the major isomer is given in Table 2.
profile for the Gd complex was used. The data were taken for
the Ho complex since its higher value of the effective magnetic
moment results in a larger Curie spin contribution to the
relaxation rate and therefore in more accurate results. Once
the distances for the different protons have been calculated, from
the dipolar contribution to the relaxation rate (first term of eq
3), the value of the electronic relaxation time τS can be
obtained.35 The very similar τS value, 0.12 ( 0.2 ps, calculated
from the data for the different peaks provides an internal check
to validate the accuracy of the procedure. This value is also
rather similar to those calculated for [Ho(DOTA)]- (0.15 ps)35
and [Ho(H2O)9]3+ (0.27 ps)38 to confirm that this parameter is
rather insensitive to the chemical environment of the metal ion.
The calculated distance values were used as input adjustable
parameters in the best-fitting procedure.
On the basis of the close similarity between the chemical
shifts of [Yb(3)]- (when for each type of proton the average of
the four resonances is considered) and those of [Yb(DOTA)]-
which is indicative of a close similarity of the corresponding
structures and which further reinforces the considerations made
before for the Ho derivative, our analysis of the dipolar shift
was restricted to the two square antiprismatic structures, AR and
DR, as starting models. As the method is based on comparing
experimental shifts with those calculated through eq 2, by using
geometrical factors extracted from the model structures, we used
a procedure that is able to modify systematically the internal
coordinates of the model to get the best agreement between
calculated and experimental shifts. Different constraints were
used (suitable range of variability for distances, dihedral, and
bond angles) with the aim of maintaining a real physical
meaning for the model. As suggested previously, to test the
quality of the obtained model for any orientation of the magnetic
axes we used an agreement factor R, defined as
1H resonances of [Yb(3)]- are sharper than those of the
corresponding Ho complex, and despite the smaller chemical
shift range an even higher number of resonances are observed.
The partial assignment of the resonances follows from a
comparison with the spectrum of [Yb(DOTA)]-. The spectrum
reproduces the same basic features found for the Ho complex:
four “broad” acetate peaks at low field and three “narrow”
acetate signals at high field. However, in this case a set of
four “broad” small intensity resonances is clearly detected for
the acetate protons of the minor isomer to indicate for this
species a solution structure of the type CR/BS. To confirm these
qualitative hypotheses on the solution structures of Ho and Yb
1
complexes with ligand 3 the H NMR spectrum was quantita-
tively and carefully analyzed as previously reported for
[Yb(DOTA)]-.12 The analysis was restricted to the resonances
of the principal isomer since only in this case was it possible
to clearly distinguish each of the 25 different peaks. Unlike
the axially symmetric DOTA complex, the analysis of the
dipolar-induced shift on the proton resonances by the paramag-
netic metal ion in [Yb(3)]- has to be based on the use of the
complete form of the dipolar equation:36
3 cos2 θ - 1
sin2 θ cos 2φ
δνi ) D1
+ D2
(2)
r3
r3
where D1 and D2 represent the magnetic susceptibility factors
and r, θ, and φ are the polar coordinates of the ligand nucleus
i with respect to the magnetic susceptibility axes of the complex.
The r distances between the paramagnetic center and a given
proton atom were independently evaluated by exploiting the
Curie contribution to the relaxation rates of the proton reso-
nances for the related [Ho(3)]- complex, according to a well-
established procedure.35 In this case we measured the longi-
tudinal relaxation rate 1/T1 at three magnetic field strengths (2.1,
6.3, and 9.0 T) at 25 °C. If the value of the molecular
reorientational correlation time τR is known, then, from the field
dependence of the relaxation rate, the distances r can be
evaluated according to the equation
(δν calc - δνiexp 2
)
∑
i
i
R )
(4)
(δν exp 2
)
∑
i
i
where δcalc is the chemical shift value calculated with eq 2 for
a given proton for each isomeric structural model for a given
orientation of the magnetic axes.
As expected, a very good agreement factor, R ) 0.045, was
obtained upon fitting the more intense set of resonances to the
structural model AR of Figure 5. The data analysis was
performed on the spectrum recorded at 5 °C to ensure, as much
as possible, the occurrence of static structures. The comparison
2
2
4γ2µeff
3r6
γ2µeff4H0
6τR
1
T1
)
τS +
(3)
2 6
2
2
[
]
5(3kT) r 1 + ωH τR
(38) Bertini, I.; Capozzi, F.; Luchinat, C.; Nicastro, G.; Xia, Z. J. Phys.
Chem. 1993, 97, 6351.
The τR value of 81 ps derived from the analysis of the NMRD

DOTA-like Ln(III) Complexes
Inorganic Chemistry, Vol. 35, No. 10, 1996 2735
Figure 10. Structures of the two isomers present in the aqueous
solutions of [Yb(3)]-: (a) major isomer (enantiomer AR, calculated by
analysis of the dipolar shifts) with proton labeling scheme and (b) minor
isomer (enantiomer CR).
Scheme 2
between observed and calculated shifts is reported in Table 2
and the structure (for the enantiomer AR) derived from the best-
fitting procedure is shown in Figure 10a. An analogous
procedure for the minor isomer does not result in a good fit
because of the incomplete spectral assignment arising from the
fact that several peaks are masked by the more intense
resonances of the principal isomer. However, further insights
into the structure of this species are provided by a simple 2D-
EXSY experiment which clarifies the details of the exchange
pathway between the two isomers. According to the scheme
of Figure 5 the enantiomer AR can interconvert either to BR
through the motion of the acetate arms or to CR through the
inversion of the conformation of the ethylenediamine groups.
If we focus our attention on the macrocyclic ring protons we
see that the two exchange pathways interconvert different proton
pairs according to Scheme 2.
1
The H 2D-EXSY spectrum of [Yb(3)]- reported in Figure
11 clearly shows all the proton connectivities which correspond
nicely to case A T C of Scheme 2.
In conclusion the two isomers found in the aqueous solution
of the La, Ho, and Yb complexes of ligand 3 correspond to
structural models A and C (Figure 10). In both structures the
substituent has replaced the same type of acetate proton, the
one located at a longer distance from the metal ion and pointing
outward from the coordination cage. This minimizes the steric
interaction of the bulky p-nitrophenyl group with the macro-
cyclic ring protons and in this regard free rotation of the aromatic
substituent in these species has been observed. Besides
structural selectivity the steric factors also play a major role in
controlling the dynamics of the complexes. Unlike the DOTA
case the interconversion process between the two isomers
involves only a rearrangement of the macrocycle: the steric
encumbrance of the substituent does not allow the acetate arm
to slide over the surface of the coordination cage.
Figure 11. Expanded regions (top, low field; bottom, high field) of
the 400 MHz 2D-EXSY spectrum for a 0.08 M solution of [Yb(3)]-
in D2O at pD ) 7.2 and 25 °C with a mixing time tm ) 5 ms and
presaturation of the HDO peak. Data were collected with 1024 points
in F2 (128 scans; 112 000 Hz) and 512 points in F1, using a delay
between pulses of 0.5 s. Data were processed using a sine square-bell
function in F1 and F2. The asterisks indicate the resonances of the minor
isomer. For clarity reasons the four axial peaks of the minor isomer in
the range -25 to -43 ppm have not been labeled.
Conclusions
The relaxometric, structural, and dynamic properties of
aqueous solutions of lanthanide(III) complexes of the p-

2736 Inorganic Chemistry, Vol. 35, No. 10, 1996
Aime et al.
nitrophenyl derivative of DOTA have been investigated by low-
and high-resolution NMR techniques. The relaxivity of the Gd
complex is higher than that of [Gd(DOTA)]- over a wide range
of magnetic field strengths as a result of a slight increase of the
molecular dimension and a negligible decrease of the electronic
relaxation time of the metal ion. The relaxivity is enhanced
slightly after formation of an inclusion compound with â-cy-
clodextrin, although steric crowding at the substituent site has
a negative influence on the strength of the interaction as
indicated by the high value of the calculated dissociation
constant.
Interesting effects due to chemical modification of the parent
DOTA ligand have been observed on the solution structures
and dynamics of the complexes. While four enantiomeric pairs
are expected in the solution of monosubstituted DOTA deriva-
tives, only two are actually observed. These differ in the relative
orientation of the two square planes formed by the donor atoms,
and their structures can be described as square antiprismatic
and “inverted” square antiprismatic, respectively. On the other
hand, in both isomers the substituent is located in a strictly
analogous position, pointing outward from the coordination cage,
a position that reduces steric repulsion with the protons of the
macrocyclic ring. Accordingly, the two isomers mutually
interconvert through a concerted rotation of the four ethylene-
diamine rings, as evidenced by a 2D-EXSY experiment for the
Yb(III) complex. The acetate groups are not involved in this
dynamic process. Such steric control of the structural and
dynamic features of the [Ln(3)]- complexes could likely be
removed by introducing a spacing aliphatic chain between the
bulky aromatic group and the substituted acetate group. In this
case we should be able to observe all four isomeric species and,
more importantly, the increased motional freedom and reduced
steric constraint of the pendant substituent should favor the
creation of noncovalent interactions with suitable substrates and
thus the optimization of the relaxivity of the Gd(III)-chelate
for different biomedical applications.
IC950981U