A CuII2 Paramagnetic Chemical Exchange Saturation Transfer Contrast Agent Enabled by Magnetic Exchange Coupling

Article abstract of DOI:10.1021/jacs.6b03060

The ability of magnetic exchange coupling to enable observation of paramagnetic chemical exchange saturation transfer (PARACEST) in transition metal ions with long electronic relaxation times (_s) is demonstrated. Metalation of the dinucleating, tetra(carboxamide) ligand HL with Cu²⁺ in the presence of pyrophosphate (P₂O₇)^4- affords the complex [LCu^II₂(P₂O₇)]^-. Solution-phase variable-temperature magnetic susceptibility data reveal weak ferromagnetic superexchange coupling between the two S = 1/2 Cu^II centers, with a coupling constant of J = +2.69(5) cm^-1, to give an S = 1 ground state. This coupling results in a sharpened NMR line width relative to a GaCu analogue, indicative of a shortening of _s. Presaturation of the amide protons in the Cu₂ complex at 37 °C leads to a 14% intensity decrease in the bulk water ¹H NMR signal through the CEST effect. Conversely, no CEST effect is observed in the GaCu complex. These results provide the first example of a Cu-based PARACEST magnetic resonance contrast agent and demonstrate the potential to expand the metal ion toolbox for PARACEST agents through introduction of magnetic exchange coupling.

Full text of DOI:10.1021/jacs.6b03060

Subscriber access provided by UNIV OF NEBRASKA - LINCOLN
Communication
II2
A Cu PARACEST Contrast Agent Enabled by Magnetic Exchange Coupling
Kang Du, and T. David Harris
J. Am. Chem. Soc., Just Accepted Manuscript • DOI: 10.1021/jacs.6b03060 • Publication Date (Web): 08 Jun 2016
Downloaded from http://pubs.acs.org on June 12, 2016
Just Accepted
“Just Accepted” manuscripts have been peer-reviewed and accepted for publication. They are posted
online prior to technical editing, formatting for publication and author proofing. The American Chemical
Society provides “Just Accepted” as a free service to the research community to expedite the
dissemination of scientific material as soon as possible after acceptance. “Just Accepted” manuscripts
appear in full in PDF format accompanied by an HTML abstract. “Just Accepted” manuscripts have been
fully peer reviewed, but should not be considered the official version of record. They are accessible to all
readers and citable by the Digital Object Identifier (DOI®). “Just Accepted” is an optional service offered
to authors. Therefore, the “Just Accepted” Web site may not include all articles that will be published
in the journal. After a manuscript is technically edited and formatted, it will be removed from the “Just
Accepted” Web site and published as an ASAP article. Note that technical editing may introduce minor
changes to the manuscript text and/or graphics which could affect content, and all legal disclaimers
and ethical guidelines that apply to the journal pertain. ACS cannot be held responsible for errors
or consequences arising from the use of information contained in these “Just Accepted” manuscripts.
Journal of the American Chemical Society is published by the American Chemical
Society. 1155 Sixteenth Street N.W., Washington, DC 20036
Published by American Chemical Society. Copyright © American Chemical Society.
However, no copyright claim is made to original U.S. Government works, or works
produced by employees of any Commonwealth realm Crown government in the course
of their duties.

Page 1 of 9
Journal of the American Chemical Society
1
2
3
4
5
6
7
8
9
A CuII PARACEST Contrast Agent Enabled by Magnetic Exchange
Coupling
2
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
Kang Du and T. David Harris*
Department of Chemistry, Northwestern University, Evanston IL 60208-3113
Supporting Information Placeholder
Nevertheless, these agents rely on through-space pseudocontact
ABSTRACT: The ability of magnetic exchange coupling to
enable observation of paramagnetic chemical exchange saturation
transfer (PARACEST) in transition metal ions with long electron-
shifts of protons, rather than through-bond contact shifts, due to
the contracted 4f orbitals,1 and therefore necessitate placement of
exchangeable protons within close proximity to the metal ion.
ic relaxation times (τs) is demonstrated. Metalation of the dinucle-
Moreover, the ionic nature of lanthanides, in conjunction with
their redox- and spin-state inertness, limits the design of respon-
sive agents.1e,3b, 6 Compared to lanthanide complexes, transition
metal PARACEST agents exhibit smaller hyperfine shifts, up to
135 ppm vs water,4f but offer more tunability through coordina-
tion and redox chemistry. Indeed, recent work has demonstrated
the ability of transition metal ions with short values of τs to en-
gender PARACEST agents that are stable under physiological
conditions.4cde Nevertheless, metal ions with large values of elec-
ating, tetra(carboxamide) ligand HL with Cu2+ in the presence of
4
−
pyrophosphate (P2O7) affords the complex [LCuII2(P2O7)]−.
Solution-phase variable-temperature magnetic susceptibility data
reveal weak ferromagnetic superexchange coupling between the
1
two S = /2 CuII centers, with a coupling constant of J = +2.69(5)
1
cm− , to give an S = 1 ground state. This coupling results in a
sharpened NMR linewidth relative to a GaCu analogue, indicative
of a shortening of τs. Presaturation of the amide protons in the Cu2
complex at 37 °C leads to a 14% intensity decrease in the bulk
5
tronic spin and higher oxidative stability, such as S = /2 MnII or
1
water H NMR signal through the CEST effect. Conversely, no
FeIII, or metal ions that exhibit even more kinetically inert substi-
tution, such as CrIII, are precluded from consideration for
PARACEST, as they exhibit long values of τs = 10−8 to 10−9 s and
therefore induce severe NMR line broadening.7 Taken together,
these observations underscore the need for a general strategy to
design transition metal-based PARACEST agents with short elec-
tronic relaxation times.
CEST effect is observed in the GaCu complex. These results pro-
vide the first example of a Cu-based PARACEST magnetic reso-
nance contrast agent, and demonstrate the potential to expand the
metal ion toolbox for PARACEST agents through introduction of
magnetic exchange coupling.
PARACEST magnetic resonance imaging (MRI) contrast
agents represent an emerging class of molecules that offer several
key advantages over conventional Gd-based proton relaxation
agents, including the ability to facilitate on/off contrast switching
and an inherent response to environmental parameters such as pH
and temperature.1 The mechanism through which PARACEST
agents generate contrast exploits labile ligand protons within close
proximity to the paramagnetic center. Here, irradiation at the res-
onant frequency of the exchangeable protons, along with concom-
itant chemical exchange of these protons with bulk water protons,
induces a suppression of bulk water signal and thus an image
darkening.2 Importantly, the paramagnetic metal imposes a large
hyperfine shift on the resonant frequency of the exchangeable
protons, so as to minimize interference from resonances associat-
ed with surrounding tissue and to permit rapid proton exchange.
Despite the potential utility of PARACEST agents, a fundamental
limitation toward their implementation arises due to proton nucle-
ar spin relaxation enhancement induced by paramagnetic metal
centers, which leads to spectral broadening and thus low sensitivi-
ty and resolution. In order to reduce proton relaxation effects, the
electronic relaxation time (τs) of the paramagnetic metal must be
minimized so that it is not resonant with the proton nuclear spin.1
To date, reported PARACEST agents have taken the form of
non-Gd lanthanide1, 3 and first-row transition metal (FeII, CoII,
NiII) complexes.4 These ions display short values of τs, typically in
the range of 10−11 to 10−12 s, owing to low-lying excited states that
arise from spin-orbit coupling and/or zero-field splitting. Lantha-
nide PARACEST agents are air-stable and tend to exhibit large
hyperfine shifts, as high as 720 ppm vs water,5 stemming from
their large magnetic moments and strong magnetic anisotropy.
A paramagnetic metal ion displays a long τs when the elec-
tronic ground state is energetically well-isolated from excited
states, with no significant zero-field splitting or spin-orbit cou-
pling. A representative example is CuII, where the d9, S = 1/2 elec-
tronic configuration and Jahn-Teller distortion result in a long τs
of 10−8 to 10−9 s.7 Nevertheless, several CuII complexes with
2
weak magnetic superexchange coupling between metal centers
have been shown to exhibit much shorter values, as small as τs
~10−11 s.8 As the separation of ground and excited state is corre-
lated to the magnitude of exchange constant J, the presence of
weak coupling leads to a low-lying excited state that can facilitate
fast electronic relaxation and therefore a short τs. Moreover, the S
= 1 ground or excited state, resulting from ferro- or antiferromag-
netic coupling, respectively, can possess non-negligible zero-field
splitting that further decreases τs and can contribute to increasing
the hyperfine shift.7 Despite the potential of magnetic exchange
coupling to serve as a tool to shorten τs, no multinuclear transition
metal complexes have been reported as PARACEST agents. Here-
in, we demonstrate the utility of magnetic exchange coupling to
enable realization of a CuII2 PARACEST agent.
Dinuclear CuII complexes supported by a phenoxo-centered
2
tetrapyridyl ligand have previously been shown to exhibit solu-
bility and stability in aqueous solution and weak exchange cou-
pling of |J| ≤ 25 cm−1.8,9 Building from these results, we targeted
the analogous tetra(carboxamide) ligand HL, with the intent that
the carboxamide groups could provide CEST-active protons (see
Figure 1). This ligand was synthesized through an SN2 reaction
between 2,2′-iminodiacetamide and 2,6-bis(bromomethyl)-4-
methylphenol (see Experimental Section and Scheme S1 in the
ACS Paragon Plus Environment

Journal of the American Chemical Society
Page 2 of 9
1
2
3
4
5
6
Figure 1. Synthesis of ligand HL.
7
8
9
Supporting Information). The ancillary ligand, pyrophosphate,
(P2O7)4−, was selected owing to its tendency to coordinate metal
ions with higher thermodynamic stability than do water and coor-
dinating anions in aqueous solution.10 Reaction of two equivalents
of Cu(NO3)2·3H2O with one equivalent each of HL and K4(P2O7)
in water gave a brown solution. Subsequent slow evaporation of
this solution afforded brown plate-shaped crystals of
H[LCu2(P2O7)]·0.5KNO3·3H2O (1). To provide a related uncou-
pled species for comparison to 1, a GaCu analogue was synthe-
sized. Here, HL was treated with one equivalent of
Ga(NO3)3·H2O, followed by one equivalent each of
Cu(NO3)2·3H2O and Na(H3L′)·3H2O (H4L′ = alendronic acid), to
give LGaCuL′·4NaNO3·7.1EtOH (2; see Figure 2 right). Tetraan-
ionic alendronate was employed in place of (P2O7)4− in order to
impart water solubility to the neutral complex.
Figure 2. Left: Crystal structure of [LCu2(P2O7)]−, as observed in
1. Cyan, pink, red, blue, and gray spheres represent Cu, P, O, N,
and C atoms, respectively; H atoms are omitted for clarity. Right:
Molecular structure of LGaCuL′, as observed in 2.
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
ics. Here, peak maxima corresponding to the LMCT and d-d tran-
sition shifted from 398 to 432 nm ( = 735 M−1 cm−1) and 726 to
779 nm ( = 104 M−1 cm−1), respectively, with each conversion
proceeding through an isosbestic point (see Figure S3). These
spectral changes suggest that aquo or hydroxo ligands are dis-
placed by (P2O7)4− ligands. In addition, the d-d transition at 779
nm lies between that of [Cu(H2O)6]2+ (810 nm,  = 12 M−1 cm−1)
and [Cu(EDTA)]2− (735 nm,  = 85 M−1 cm−1), confirming the
preservation of the elongated CuII octahedral geometry in solu-
tion.13 Fitting the absorbance data at 246 nm from titrations at
lower concentration (40.2 μM) gave a dissociation constant of Kd
= 9(2) μM (see Figures S3-4), confirming the high affinity of
(P2O7)4−. The diffuse-reflectance spectrum collected for a solid
sample of 1 features peaks with maxima at 436 and 788 nm (see
Figure S5), and an electrospray mass spectrum exhibits three ma-
jor patterns corresponding to adducts of the anionic complex of 1
(see Figure S6). Taken together, these data lead us to conclude
that the structure of [LCu2(P2O7)]− determined from X-ray diffrac-
tion is preserved in aqueous HEPES solution. Finally, the spec-
trum of 2 exhibits similar features to that of 1 (see Figures S7-8),
with the LMCT and d-d transitions at 418 nm ( = 297 M−1) and
784 nm ( = 75 M−1 cm−1), respectively, only slightly shifted.
These spectral similarities suggest that the Cu centers in 1 and 2
feature similar coordination environments in aqueous solution.
Variable-temperature dc magnetic susceptibility data were
collected to determine the sign and magnitude of magnetic cou-
pling in the Cu2 complex. To approximate the magnetic interac-
tions in aqueous solution, magnetic data were collected on a solu-
tion of 1 in a 1:1 H2O/glycerol mixture.14 The corresponding plot
of MT vs T is shown in Figure 3. Due to the error associated with
mass determination of such a small amount of solute, the number
of moles of Cu2 complex was determined by a fixed g = 2.0629
obtained from X-band EPR, along with the assumption that the
high-temperature values of MT correspond to two non-interacting
Compound 1 crystalizes in the space group P−1, with an
asymmetric unit that contains two nearly identical Cu2 complexes
(see Figure 2 left and Table S1). The two CuII centers of each
molecule reside in distorted octahedral coordination environ-
ments, with significantly elongated Jahn-Teller axes along the
Oamide-Cu-Ophenoxy and Ophosphate-Cu-Namine vectors. As a result, all
four pendant amides are structurally inequivalent in the solid
state. The pyrophosphate unit acts as a tetradentate, dinucleating
ligand that coordinates the two Cu centers in a 2-4 configura-
tion. This binding mode presumably engenders a chelate effect of
(P2O7)4− that results in preferential coordination of the Cu2 com-
plex over water and other anions in solution. The mean Cu···Cu
distance of 3.658(1) Å is comparable to those reported for alkoxo-
or phenoxo-bridged CuII complexes that feature acetate bridging
2
ligands,7 and the mean Cu-O-Cu angle of 123.8(2)° is similar to
those reported for Cu2 complexes with weak antiferromagnetic or
ferromagnetic exchange coupling.11
In anticipation of CEST experiments, we sought to confirm the
integrity and structure of [LCu2(P2O7)]− in buffered aqueous solu-
tion. Toward this end, the kinetic and thermodynamic profiles of
complexation for Cu2+ ions by L− and (P2O7)4− in aqueous solu-
tion were monitored by UV/Visible spectroscopy. First, small
aliquots of a 50 mM aqueous solution of Cu(NO3)2·3H2O were
titrated into a pH 7 buffer solution of 5 mM 4-(2-hydroxyethyl)-1-
piperazineethanesulfonic acid (HEPES) containing 0.317 mM HL
(see Figure S1). Prior to Cu addition, the spectrum of HL was
characterized by one major peak at 285 nm, which we tentatively
assign as a - transition. Upon Cu addition, two new features at
398 nm ( = 580 M−1 cm−1) and 726 nm ( = 129 M−1 cm−1)
emerged, which we assign as ligand-metal charge transfer
(LMCT) and d-d transitions, respectively, based on literature
precedent.8bc Spectral changes ceased within 3 minutes following
each addition, suggesting relatively rapid Cu complexation by L−.
Moreover, the peak intensity at 398 nm is linearly dependent on
the concentration of Cu2+ and reaches a maximum value upon
addition of two equivalents of Cu2+ (see Figure S2), suggesting
the formation of a 1:2 complex of L− and Cu2+ with a dissociation
constant well below millimolar. The resulting species is likely the
diaquo adduct [LCu2(H2O)4]3+ or the µ-hydroxo adduct [LCu2(µ-
OH)]2+, as have been observed in related tetrapyridyl-supported
CuII2 complexes.12
1
S = /2 centers (see Experimental Section and Figure S10). With
decreasing temperature, MT remains relatively constant to ca.
100 K, whereupon it undergoes a gradual increase to a maximum
value of MT = 0.929 cm3K/mol at 6 K. Below 6 K, MT decreas-
es sharply to a minimum value of 0.853 cm3K/mol at 2 K. The
increase in MT below 100 K stems from ferromagnetic coupling
between the CuII ions, likely dominated by superexchange through
the phenoxo bridge, to give an S = 1 ground state.
The data were fit in the range 10-300 K to the Van Vleck equa-
tion considering the spin Hamiltonian ꢀ = −2J(ꢁCu1·ꢁCu2), where
ꢁCu1 and ꢁCu2 are the spin operators.15 The fit to the data gave an
exchange constant of J = +2.69(5) cm−1, indicating a weak ferro-
magnetic Cu···Cu interaction (see Figure 3). This interaction re-
sults in an energetic separation of the S = 1 ground state and S = 0
excited state of only 5.38 cm−1 (see Figure 3, inset), which sug-
Titration of the Cu2 solution with a buffered pH 7 aqueous
solution of K4(P2O7) revealed similarly rapid complexation kinet-
ACS Paragon Plus Environment

Page 3 of 9
Journal of the American Chemical Society
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
Figure 3. Variable-temperature dc magnetic susceptibility data
for 1 in 1:1 H2O/glycerol, collected under an applied field of 1 T.
The red line corresponds to a fit of the data. Inset: Spin ladder
and graphical representation of the energetic separation between
S = 1 ground and S = 0 excited state.
Figure 4. CEST spectra of 10 mM of 1 (top) and 30 mM of 2
(bottom) solutions in 50 mM of HEPES buffer at pH 7 and 37 °C,
with a 2 s presaturation pulse of 21 μT.
gests that 1 may feature a considerably shortened τs relative to a
monocopper complex with CuII in similar coordination.
ration at the water proton resonance. For 1, in stark contrast to 2, a
peak was observed at 29 ppm corresponding to a 14% decrease in
intensity of the water signal due to CEST. Although significant,
the CEST effect is likely limited due to incomplete presaturation
of the broad resonance of the amide protons. The presence of a
slight shoulder on the water resonance in 2 likely corresponds to
CEST arising from carboxamide protons coordinated to the dia-
magnetic GaIII center. Since the T1 of water protons can signifi-
cantly influence the CEST effect,1b these values were measured
for 20 mM solutions of 1 and 2, in addition to pure water, as T1 =
352(4) ms, 152(1) ms, and 4.36(4) s, respectively. The slightly
lower value for 2 relative to 1 is consistent with more efficient
proton relaxation stemming from a longer s. For 1, the CEST
peak intensity at 25 °C decreases to 7.8%, suggesting that the
exchange rate of the carboxamide protons is in the slow exchange
regime with respect to the saturation offset (see Figure S12). The
exchange rate at 37 °C was estimated at kex = 420(20) s−1 by em-
ploying the omega plot method (see Figure S13),16 and this value
is comparable to those reported for mononuclear Fe-based
The relaxation of proton nuclear spins in 1 and 2 was probed
1
by H NMR to obtain a qualitative comparison of τs. Here, elec-
tronic spins with long values of τs induce a shortening of spin-
lattice relaxation time (T1) in proximal nuclear spins, which leads
to a broadening of the NMR spectra. Such an analysis has been
previously employed to obtain a qualitative measure for τs in re-
lated Cu2 complexes.7,8b Importantly, the presence of similar local
coordination at CuII in 1 and 2 suggests that the hyperfine shifts of
analogous protons on the two complexes should be similar. In the
case of 1, the 1H NMR spectrum collected for a 10 mM H2O solu-
tion at 37 °C features three paramagnetically-shifted resonances at
12, 14, and 29 ppm vs H2O. Inversion recovery experiments of the
three peaks gave values of T1 = 8(1), 13(1) and 22(4) ms, respec-
tively (see Experimental Section and Figure S13), suggesting that
the T1 shortening of the peaks by the CuII centers is comparable
for all three resonances. In addition, the peak at 29 ppm is absent
in the spectrum of 1 in D2O, indicating that this peak corresponds
to the exchangeable amide protons. The presence of only one
amide resonance suggests fast interchange among the four pen-
dant arms in solution that leads to an average of four inequivalent
amides. As such, the broadness of the amide resonance, compared
to the other two paramagnetic peaks, is likely due to the combina-
tion of fast molecular dynamics and proton exchange, rather than
relaxation induced by CuII. In contrast, an aqueous sample con-
taining 30 mM of 2 in either D2O or H2O gave a spectrum that is
absent of resolvable resonances in the paramagnetic region (see
Figure S11), presumably due to line broadening induced by CuII.
This comparison provides strong qualitative evidence that the Cu2
complex in 1 features a shorter τs than does the GaCu complex in
2, likely resulting from the presence of weak magnetic exchange
coupling in 1.8b
PARACEST agents with pendant carboxamide groups.4d Interest-
1
ingly, despite containing S =
/ metal centers with negligible
2
single-ion magnetic anisotropy, 1 gives rise to a CEST peak shift
and intensity that is comparable to some reported mononuclear
PARACEST agents comprised of the high-spin, high-anisotropy
FeII ion.4d In sum, these experiments demonstrate the ability of 1
to provide MR contrast through the CEST effect.
Preliminary experiments were carried out to probe the stability
of 1 to reduction, pH variation, and ion substitution. The cyclic
voltammogram of 1 shows a reduction wave at ca. −500 mV vs.
NHE (see Figure S14), suggesting that 1 would not undergo re-
duction under physiological conditions.17 In addition, the UV-Vis
spectra of 1 confirm the integrity of [LCu2(P2O7)]− in the pH
range 6-8 or in the presence of excess Na2CO3 and NaH2PO4 (see
Figure S15-16). However, in the presence of excess Zn2+ ions, the
LMCT is red shifted by 20 nm and the d-d transition disappears
(see Figure S17), suggesting the presence of ZnII substitution.
The foregoing results demonstrate that the limited scope of
metal ions suitable for PARACEST can be expanded through
introduction of magnetic exchange coupling in multinuclear com-
plexes. Efforts are underway to extend this initial proof-of-
concept investigation to metal ions with larger values of spin and
higher stability and to apply this strategy to the design of respon-
sive PARACEST agents.
The dramatic sharpening of the NMR linewidths for the Cu2
complex relative to the GaCu analogue indicates the possibility to
1
observe the CEST effect. We therefore collected H NMR spectra
on solutions of 1 and 2 in 50 mM HEPES buffer at pH 7 and 37
°C on an 11.75 T NMR spectrometer (see Figure 4). The resulting
CEST spectra, also known as Z spectra, were constructed by plot-
1
ting the intensity of the water H NMR signal (MZ/M0, where M0
and MZ correspond to the bulk water signal before and after pre-
saturation at a given frequency, respectively) against the presatu-
ration frequency (i.e. frequency offset) relative to the bulk water
frequency, set to 0 ppm. For 1 and 2, complete disappearance of
the water signal was observed at 0 ppm, arising from direct satu-
ACS Paragon Plus Environment

Journal of the American Chemical Society
Page 4 of 9
1
2
3
4
ASSOCIATED CONTENT
Supporting Information
39, 51. (g) Ratnakar, S. J.; Viswanathan, S.; Kovacs, Z.; Jindal, A.
K.; Green, K. N.; Sherry, A. D. J. Am. Chem. Soc. 2012, 134, 5798.
(h) Hingorani, D. V.; Randtke, E. A.; Pagel, M. D. J. Am. Chem.
Soc. 2013, 135, 6396. (i) Ekanger, L. A.; Ali, M. M.; Allen, M. J.
Chem. Commun. 2014, 50, 14835.
Crystallographic data for 1, additional experimental details, spec-
troscopic data, mass spectrometric data, and CIF for 1. This mate-
rial is available free of charge via the Internet at
http://pubs.acs.org.
5
6
7
8
(7) (a) Bertini, I.; Luchinat, C. NMR of Paramagnetic Molecules in
Biological Systems; The Benjamin/Cummings Publishing Company,
Inc.: Menlo Park, 1986. (b) Bertini, I.; Luchinat, C. Coord. Chem.
Rev. 1996, 150, 1. (c) Bertini, I.; Luchinat, C.; Parigi, G. Solution
NMR of Paramagnetic Molecules: Applications to Metallobiomole-
cules and Models; Elsevier Science B. V.: Amsterdam, 2001.
(8) (a) Nishida, Y.; Shimo, H.; Maehara, H.; Kida, S. J. Chem. Soc.
Dalton Trans. 1985, 1945. (b) Murthy, N. N.; Karlin, K. D.; Bertini,
I.; Luchinat, C. J. Am. Chem. Soc. 1997, 119, 2156. (c) Siluvai, G.
S.; Murthy, N. N. Polyhedron 2009, 28, 2149.
(9) These values correspond to the Hamiltonian ꢀ = −2J(ꢁCu1·ꢁCu2).
(10) Lee, D. H.; Im, J. H.; Son, S. U.; Chung, Y. K.; Hong, J.-I. J. Am.
Chem. Soc. 2003, 125, 7752.
(11) Osꢀrio, R. E. H. M. B.; Peralta, R. A.; Bortoluzzi, A. J.; de Almeida,
V. R.; Szpoganicz, B.; Fischer, F. L.; Terenzi, H.; Mangrich, A. S.;
Mantovani, K. M.; Ferreira, D. E. C.; Rocha, W. R.; Haase, W.;
Tomkowicz, Z.; dos Anjos, A.; Neves, A. Inorg. Chem. 2012, 51,
1569.
9
AUTHOR INFORMATION
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
Corresponding Author
dharris@northwestern.edu
Notes
The authors declare no competing financial interests.
ACKNOWLEDGMENT
This research was funded by the Chemistry of Life Processes
Institute, Northwestern University, and the Air Force Research
Laboratory (AFRL) under agreement number FA8650-15-5518.
We thank Drs. L. Liu, I.-R. Jeon and J. M. Zadrozny for helpful
discussions, Dr. Y. Zhang, Prof. J. S. Anderson, and L. M. Lilley
for experimental assistance, and Prof. T. J. Meade for use of his
HPLC system.
(12) Torelli, S.; Belle, C.; Gautier-Luneau, I.; Pierre, J. L.; Saint-Aman,
E.; Latour, J. M.; Le Pape, L.; Luneau, D. Inorg. Chem. 2000, 39,
3526.
(13) Baker, A. T. J. Chem. Educ. 1998, 75, 98.
(14) Note that the peak maxima in the UV/Visible spectra of 1 in H2O
and 1:1 H2O/glycerol are identical (see Figure S9).
(15) (a) Magnetic susceptibility data were simulated using the program
MagProp: Tregenna-Piggott, P. L. W.; MagProp (part of the NIST
DAVE
software
suite),
version
2.0,
2008,
http://www.ncnr.nist.gov/dave (b) Azuah, R. T.; Kneller, L. R.; Qiu,
Y. M.; Tregenna-Piggott, P. L. W.; Brown, C. M.; Copley, J. R. D.;
Dimeoo, R. M. J. Res. Natl. Inst. Stand. Technol. 2009, 114, 341.
(16) Dixon, W. T.; Ren, J.; Lubag, A. J. M.; Ratnakar, J.; Vinogradov, E.;
Hancu, I.; Lenkinski, R. E.; Sherry, A. D. Magn. Reson. Med. 2010,
63, 625.
REFERENCES
(1) (a) Zhang, S.; Winter, P.; Wu, K.; Sherry, A. D. J. Am. Chem. Soc.
2001, 123, 1517. (b) Zhang, S.; Merritt, M.; Woessner, D. E.; Len-
kinski, R. E.; Sherry, A. D. Acc. Chem. Res. 2003, 36, 783. (c) Zhou,
J.; van Zijl, P. C. M. Prog. Nucl. Magn. Reson. Spectrosc. 2006, 48,
109. (d) Ali, M. M.; Liu, G.; Shah, T.; Flask, C. A.; Pagel, M. D.
Acc. Chem. Res. 2009, 42, 915. (e) Viswanathan, S.; Kovacs, Z.;
Green, K. N.; Ratnakar, S. J.; Sherry, A. D. Chem. Rev. 2010, 110,
2960. (f) Terreno, E.; Castelli, D. D.; Viale, A.; Aime, S. Chem. Rev.
2010, 110, 3019.
(17) Tsitovich, P. B.; Bums, P. J.; McKay, A. M.; Morrow, J. R. J. Inorg.
Biochem. 2014, 133, 143.
(2) Ward, K. M.; Aletras, A. H.; Balaban, R. S. J. Magn. Reson. 2000,
143, 79.
(3) Selected references: (a) Zhang, S.; Michaudet, L.; Burgess, S.; Sher-
ry, A. D. Angew. Chem. Int. Ed. 2002, 41, 1919. (b) Aime, S; Cas-
telli, D. D.; Terreno, E. Angew. Chem. Int. Ed. 2002, 41, 4334. (c)
Terreno, E.; Castelli, D. D.; Cravotto, G.; Milone, L.; Aime, S. In-
vest. Radiol. 2004, 39, 235. (d) Woods, M.; Woessner, D. E.; Zhao,
P.; Pasha, A.; Yang, M.-Y.; Huang, C.-H.; Vasalitiy, O.; Morrow, J.
R.; Sherry, A. D. J. Am. Chem. Soc. 2006, 128, 10155.
(4) Selected references: (a) Dorazio, S. J.; Tsitovich, P. B.; Siter, K. E.;
Spernyak, J. A.; Morrow, J. R. J. Am. Chem. Soc. 2011, 133, 14154.
(b) Tsitovich, P. B.; Morrow, J. R. Inorg. Chim. Acta 2012, 393, 3.
(c) Olatunde, A. O.; Dorazio, S. J.; Spernyak, J. A.; Morrow, J. R. J.
Am. Chem. Soc. 2012, 134, 18503. (d) Dorazio, S. J.; Morrow, J. R.
Inorg. Chem. 2012, 51, 7448. (e) Dorazio, S. J.; Olatunde, A. O.;
Spernyak, J. A.; Morrow, J. R. Chem. Commun. 2013, 49, 10025. (f)
Tsitovich, P. B.; Spernyak, J. A.; Morrow, J. R. Angew. Chem. Int.
Ed. 2013, 52, 13997. (g) Jeon, I.-R.; Park, J. G.; Haney, C. R.; Har-
ris, T. D. Chem. Sci. 2014, 5, 2461.
(5) Woods, M.; Woessner, D. E.; Sherry, A. D. Chem. Soc. Rev. 2006,
35, 500.
(6) Selected references: (a) Zhang, S.; Trokowski, R.; Sherry, A. D. J.
Am. Chem. Soc. 2003, 125, 15288. (b) Zhang, S.; Malloy, C. R.;
Sherry, A. D. J. Am. Chem. Soc. 2005, 127, 17572. (c) Trokowski,
R.; Ren, J.; Kálmán, F. K.; Sherry, A. D. Angew. Chem. Int. Ed.
2005, 44, 6920. (d) Que, E. L.; Chang, C. J. J. Am. Chem. Soc. 2006,
128, 15942. (e) Huang, C.-H.; Morrow, J. R. J. Am. Chem. Soc.
2009, 131, 4206. (f) Que, E. L.; Chang, C. J. Chem. Soc. Rev. 2010,
ACS Paragon Plus Environment

Page 5 of 9
Journal of the American Chemical Society
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
236x67mm (300 x 300 DPI)
ACS Paragon Plus Environment

Journal of the American Chemical Society
Page 6 of 9
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
413x178mm (150 x 150 DPI)
ACS Paragon Plus Environment

Page 7 of 9
Journal of the American Chemical Society
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
204x164mm (150 x 150 DPI)
ACS Paragon Plus Environment

Journal of the American Chemical Society
Page 8 of 9
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
220x227mm (150 x 150 DPI)
ACS Paragon Plus Environment

Page 9 of 9
Journal of the American Chemical Society
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
39
40
41
42
43
44
45
46
47
48
49
50
51
52
53
54
55
56
57
58
59
60
44x23mm (300 x 300 DPI)
ACS Paragon Plus Environment