A cobalt(ii) complex with unique paraSHIFT responses to anions

Article abstract of DOI:10.1039/c7cc00619e

ParaSHIFT agents have shown promise in detecting chemical targets in biological systems by magnetic resonance, but few studies have used transition metal complexes for this purpose. Here we report our investigations into CoMe₆trenCl (tren = tris(2-aminoethyl)amine) as a paraSHIFT agent. The paramagnetic region of the ¹H NMR spectrum shows characteristic spectral profiles in the presence of fluoride, acetate, lactate and citrate in aqueous solution. These distinctive NMR shifts of each anion are maintained even in mixtures of anions.

Full text of DOI:10.1039/c7cc00619e

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A cobalt(II) complex with unique paraSHIFT
responses to anions†
Cite this:DOI: 10.1039/c7cc00619e
a
a
b
a
E. S. O’Neill, J. L. Kolanowski, P. D. Bonnitcha and E. J. New*
Received 23rd January 2017,
Accepted 6th March 2017
DOI: 10.1039/c7cc00619e
rsc.li/chemcomm
ParaSHIFT agents have shown promise in detecting chemical targets of transition metal complexes, which can transfer the relaxation to
in biological systems by magnetic resonance, but few studies have the bulk water by a mechanism termed paramagnetic chemical
9
used transition metal complexes for this purpose. Here we report our exchange saturation transfer (paraCEST). This has been utilised in
1
0,11
12
investigations into CoMe
6
trenCl (tren = tris(2-aminoethyl)amine) as a a variety of transition metal complexes, including Fe(II),
Ni(II),
1
13
paraSHIFT agent. The paramagnetic region of the H NMR spectrum and cobalt, with the main challenge being the intrinsically
shows characteristic spectral profiles in the presence of fluoride, low ‘‘per-molecule’’ sensitivity of these agents.
acetate, lactate and citrate in aqueous solution. These distinctive An alternative approach is to use magnetic resonance spectro-
1
4–16
NMR shifts of each anion are maintained even in mixtures of anions. scopy (MRS), which observes resonances other than the bulk
water signal. In order to avoid the background from the protons
The focus of medical research has shifted from primarily questions of water and fat, the observed shift must be distinct from the
1
17
of structure to those of chemistry, and there is therefore a need for endogenous diamagnetic region of H NMR. This can be
imaging techniques that report on the chemical environments of achieved by probing a nucleus that is not typically present
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9
tissues and cells. Magnetic resonance imaging (MRI) is one of in biological systems, such as F. Responsive agents of this
the most powerful imaging techniques, offering particularly high type have been prepared by incorporating this nuclide in para-
1
8–21
spatial resolution, and recent research efforts have therefore magnetic systems.
Clinical application of these agents
focussed on MRI protocols that can provide molecular information may be limited by sensitivity challenges, and the less routine
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9
17
as well as structural. This has precipitated the development of availability of detection coils suitable for F. Alternatively, to
responsive MRI contrast agents, which change the relaxation of achieve improved sensitivity and minimised background signal,
1,2
the bulk water protons upon chemical stimuli. Such systems are and compatibility with existing MRI instrumentation, a strategy
1
limited, however, by the concentration dependence of the response, involving paramagnetically-shifted H signals (paraSHIFT) can
due to the high backgrounds observed arising from both the be employed. By this method, pH and/or temperature-responsive
2
2,23
intrinsic properties of tissue and in particular the residual relaxa- complexes of a number of lanthanoids, including ytterbium,
tion effect of the contrast agent even in its inactivated form.
3,4
24
25
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praseodymium, thulium, and dysprosium have been developed
We and others have addressed this challenge by utilising and their utility in biological systems has been confirmed, with
2
6
transition metals that can be converted from diamagnetic to the detection limit as low as tens of mM.
paramagnetic forms upon external stimulus. For example, in While lanthanoid complexes predominate literature on shift
Fe(II) systems the switch from the low spin diamagnetic to high spin agents, it has long been known that first row transition metal
1
27,28
paramagnetic form was achieved upon ligand-centred chemical ions can be applied to paramagnetic H NMR spectroscopy.
5–7
reduction or enzymatic activity, while we have shown that the However, transition metal complexes for paraSHIFT imaging
biologically-tuned reduction of Co(III) to Co(II) gives similar off-to-on applications have been reported only recently, with Fe(II) and
effects. Another solution has been to use the same diamagnetic Co(II) systems utilised as temperature sensors. In line with
to paramagnetic switch to alter the readily-exchangeable protons our interest in cobalt systems, we sought to identify a scaffold
that could sense a chemical target. We report here our observa-
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tions of the differential response of paramagnetically-shifted
protons on a cobalt complex upon binding to different anions.
a
School of Chemistry, The University of Sydney, NSW 2006, Australia.
E-mail: elizabeth.new@sydney.edu.au
b
In our previous studies of cobalt-tris-2-pyridylmethylamine
Sydney Medical School, Royal North Shore Hospital, St. Leonards, NSW 2065,
Australia
systems, we observed a large paramagnetic shift of almost
8
†
Electronic supplementary information (ESI) available. See DOI: 10.1039/c7cc00619e 100 ppm for some ligand protons. In order to maximise this
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Fig. 2 Paramagnetic H NMR of CoMe
6
TrenCl (1 mM) upon addition of
various anions (lactate, citrate, acetate and fluoride) that induce unique
chemical shifts of the methyl protons of CoMe
pH 6.5, 9.4 T).
6 2 2
TrenCl (H O : D O 90 : 10,
1
Fig. 1 Paramagnetic H NMR region of CoMe
inset) in D O at 9.4 T.
6
TrenCl (10 mM, structure
2
signal, we sought a ligand bearing a large number of equivalent
protons. Cobalt(II) complexes of tris(2-aminoethyl)amine (tren) have
30–32
previously been widely studied.
methyl analogue of this system (CoMe
We identified the hexa-N-
TrenCl) (Fig. 1 inset) as an
6
appropriate model complex, as it has up to 18 equivalent protons.
This complex has previously been studied for its photophysical and
binding properties, and has demonstrated potential utility as an
3
3
atom-transfer radical-polymerisation (ATRP) radical mediator
3
4
and single molecule magnet. In addition, the observed effects
of different anions on the ligand field of the Co(II) centre in the
solid state indicated that this complex may be responsive to
3
0
anions in solution.
We prepared CoMe
6
5 1
TrenCl following a reported procedure
H NMR spectroscopy of the complex
3
(Scheme S1, ESI†).
revealed the presence of significantly-shifted broad proton signals
at 180–130 ppm (Fig. 1). In order to identify the origin of these
main signals, we prepared the (d -Me) Tren ligand and complex.
3 6
We observed the clear loss of the signals in the 180–130 ppm
region compared to the non-deuterated analogue prepared by
the same synthetic method (Fig. S1, ESI†), confirming that
these signals arise from the methyl groups.
Given the previously-reported interactions of the complex with
30
anions, we sought to investigate the effects of various anions on
the paramagnetic region. Interestingly, we observed a distinctive
spectral profile within the 180–130 ppm region for each added
anion (Fig. 2), indicating that the paramagnetic environment of
the methyl groups is diagnostic of the anion bound.
Fig. 3 Investigations of the interaction of NaF with CoMe
6 2 2
Tren (H O : D O
9
0: 10, pH 6.5, 9.4 T). (a and b) Sodium fluoride binding affinity determina-
1
tion by paramagnetic H NMR (1 mM CoMe
6
Tren), (c) UV-Visible absorption
Tren (5 mM),
Of the anions tested, fluoride showed the sharpest and most
ratio between peak intensities at 637 nm and 605 nm of CoMe
6
intense signal, which increased upon the gradual addition (d) water longitudinal relaxation rate by inversion recovery sequences in the
1
9
presence of CoMe
6
Tren (1 mM), and (e) F NMR shift of NaF with varying
of the anion (Fig. 3a). By monitoring peak intensity (Fig. 3b),
À1
[NaF] and fixed CoMe Tren (1 mM), relative to NaF signal at À120.05 ppm
6
we were able to establish a weak 1 : 1 binding (29 Æ 7 M ). We
6
without CoMe Tren.
could also monitor the ratio of the peaks at 140 and 165 ppm,
which demonstrates a linear dependence on fluoride concentration
up to 40 mM (Fig. S2, ESI†). Importantly, this correlation is An alternative method of studying binding events to a para-
independent of the cobalt concentration, enabling the quantifica- magnetic centre, which has been extensively utilised to inves-
À
tion of F in unknown samples. This binding affinity was consis- tigate metalloprotein–ligand interactions, is to monitor the T
1
3
6
tent with that obtained from absorption spectroscopy, in which relaxation of the bulk water signal. In our case, the relaxivity
fluoride addition led to increases in the peaks at 455 and 635 nm, decreased upon the addition of F , consistent with a lower
À
and a concomitant decrease at 605 nm (Fig. 3c and Fig. S3, ESI†). accessibility of water molecules to the paramagnetic metal
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Table 1 Association binding constants of fluoride to CoMe TrenCl cal-
culated by different methods
À1
Method
K
a
(M
)
T
UV
1
31 Æ 6
30 Æ 4
29 Æ 7
34 Æ 5
pNMR intensity
19
F NMR
centre (Fig. 3d). We were also able to monitor the binding event
1
9
by observing the chemical shift of the fluoride anion in
F
NMR (Fig. 3e and Fig. S4, ESI†). The presence of CoMe
6
TrenCl
1
9
À
caused a downfield shift in the F NMR spectrum relative to F
alone. This peak then shifted upfield with subsequent addition of
excess fluoride. The binding affinities calculated for all of these
methods were in agreement, within error (Table 1). In terms of
1
6
Fig. 5 Paramagnetic region of H NMR spectrum of CoMe Tren (1 mM,
À
sensitivity, it is possible to detect 1 mM F using a 1 mM solution H O : D O 90 : 10, pH 6.5, 9.4 T) in the presence of a 1 : 1 : 1 mixture of
2
2
lactate, acetate and fluoride (50 mM; purple). Spectra of the complex in the
presence of single anions (50 mM) are also shown (blue – lactate, red –
acetate and green – fluoride).
of cobalt complex, with a total experiment time of approximately
1
minute (signal : noise ratio 44; Fig. S5a, ESI†).
We then investigated the binding properties of the complex
towards lactate, acetate and citrate (Fig. 4 and Fig. S6, ESI†).
Acetate showed similarly weak binding to fluoride, with lactate
an order of magnitude weaker. By contrast, the binding affinity
of citrate is markedly tighter, and its spectral form includes two
signals at shifts consistent with those of both lactate and
acetate-bound forms (Fig. 2). Taken together, this might indi-
cate a cooperative mode of binding for citrate.
best signal : noise ratio (49), and requires the shortest experi-
mental acquisition time (approximately 1 minute; Fig. S5d, ESI†).
Competition studies with fluoride and lactate show that
lactate can be displaced upon fluoride addition (Fig. S7, ESI†),
confirming the reversibility of binding. Interestingly, even in
mixtures of anions, the unique signal for each anion is preserved,
rather than an average signal (Fig. 5). These results demonstrate
that this system is uniquely poised to simultaneously detect
several different anions in mixtures.
As for fluoride, the sensitivity of the complex is sufficient to
detect, within 1–20 min, 1 mM of these anions (signal : noise ratio
3
7
38
4
4), which correspond to the reported levels of lactate, acetate
39
and citrate in biological fluids (Fig. S5b–d, ESI†). Significantly,
In summary, we have demonstrated that the model para-
magnetic CoMe TrenCl complex can be used to report on the
6
citrate, which shows the tightest binding of the three offers the
presence of a variety of anions in aqueous solutions by observing the
distinct chemical shift of the N-Me protons. Significantly, this system
contributes an alternative method for fluoride detection, currently
40
predominated by irreversible reaction-based probes. Furthermore,
the unique spectral fingerprint for each individual anion opens
up the possibility for ‘‘multicolour’’ magnetic resonance. Future
investigation into the detailed mechanism of binding for each
anion will enable the rational design of novel scaffolds with
varied selectivities and binding affinities towards applications in
environmental and biological imaging.
This work was supported by an Australian Postgraduate Award
(ESO), a Heart Research Australia Research Support Grant (2014-06),
the Australian Research Council (DP150100649, DE120102687), and
the Westpac Bicentennial Foundation (EJN). The authors acknow-
ledge the facilities and the scientific and technical assistance of
Dr Ian Luck at the NMR Facility in the School of Chemistry at the
University of Sydney. We are grateful to Professor David Parker,
Durham University for advice and helpful discussions.
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Fig. 4 Paramagnetic H-NMR spectra and the results of corresponding
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longitudinal relaxation studies of CoMe trenCl (1 mM) with the addition of
9704–9721.
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