52g/55Mn-Labelled CDTA-based trimeric complexes as novel bimodal PET/MR probes with high relaxivity

Article abstract of DOI:10.1039/c8dt04996c

trans-1,2-Diaminocyclohexane-N,N,N′,N′-tetraacetic acid (CDTA) labelled with a mixture of paramagnetic ⁵⁵Mn(ii) and β⁺-emitting ^52gMn(ii) offers access to bimodal Positron Emission Tomography/Magnetic Resonance (PET/MR) tr

Full text of DOI:10.1039/c8dt04996c

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ISSN 1477-9226
PAPER
Joseph T. Hupp, Omar K. Farha et al.
Efficient extraction of sulfate from water using
framework
a Zr-metal–organic
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DOI: 10.1039/C8DT04996C
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52g/55Mn-Labelled CDTA-based trimeric complexes as novel bimodal
PET/MR probes with high relaxivity
Received 00th January 20xx,
Accepted 00th January 20xx
Marie R. Brandt,a Christian Vanasschen,a Johannes Ermert,*a Heinz H. Coenena and Bernd
Neumaiera,b
DOI: 10.1039/x0xx00000x
www.rsc.org/
Trans-1,2-diaminocyclohexane-N,N,N‘,N‘-tetraacetic acid (CDTA)
labelled with a mixture of paramagnetic 55Mn(II) and β+-emitting
52gMn(II) offers access to bimodal Positron Emission Tomography
/Magnetic Resonance (PET/MR) tracers. To enhance the number
of NMR-active nuclei and simultaneously improve the
longitudinal relaxivity r1, a complex composed of three CDTA
units was designed. Accordingly, a functionalised tris-CDTA-
1,3,5-tris-triazolobenzene was prepared and labelled with c.a.
and n.c.a. 52gMn. Relaxivity measurements of the 55Mn-complex
showed an enhancement of r1 of 144% in comparison to the Mn-
CDTA monomer. Moreover, the trimer was equipped with an
additional linker functionality suitable for conjugation with
biomolecules, enabling interaction with specific molecular
targets.
Fig. 1), this has been clinically applied as Teslascan® for the
detection of liver lesions in MR imaging studies. However,
Teslascan® was withdrawn from the market due to its insufficient
in vivo stability4. The free ion Mn2+ is neurotoxic, why chelating
agents that form highly inert and stable complexes have to be
applied in order to avoid manganese release from the complex and
to obtain high tissue contrast. Recent work revealed that trans-
CDTA-complexes, which were labelled isotopically with mixtures of
[
52g/55Mn]Mn(II), showed high serum stability and T1 contrast
enhancement.5 Furthermore, manganese complexes with trans-
CDTA derivatives as ligands have recently been the subject of
searching for MR imaging probes free of gadolinium.6a-c
The aim of this work was the development of multimeric trans-
CDTA-based manganese (II) complexes with enhanced MR
sensitivity and relaxivity.
In recent years, bimodal Positron Emission Tomography/ Magnetic
Resonance (PET/MR) imaging contrast agents and their application
have sparked great research interest with regard to the synergistic
combination of both imaging methods1a-e. MR imaging offers a
very high resolution, however, its sensitivity is relatively low. In
contrast, PET provides a very high sensitivity whereas the
resolution is deficient. The sensitivity and resolution mismatch
between the modalities can be counterbalanced by applying
mixtures of paramagnetic MR contrast agents in millimolar
amounts spiked with the corresponding isotopically labelled PET-
compound. Both compounds will exhibit the same in vivo
biodistribution pattern, since both are chemically identical.
3-
OH
O
HO
O
O
P
P
O
HO
OH
N
N
Mn
O
N
N
O
O
O
Figure 1: Mn-DPDP (Teslascan®), a manganese based T1 contrast agent.
A major disadvantage of manganese complexes, besides their
limited in vivo stability, is their lower average longitudinal
relaxivity (r1), owing to their lower number of unpaired electrons
in comparison to the clinically established gadolinium.
The positron emitting nuclide 52gMn (t1/2 = 5.5d) with its low
+
maximum positron energy (Eβ max = 576 keV) is well suited for PET-
applications and has already been used in several in vivo studies2a-
For compensation of this drawback of Mn compounds as MR
contrast agents, an extensive search for stable Mn(II) complexes
with appropriate relaxivity had to be carried outc. To enhance the
relaxivity of a metal complex, the use of rigid ligand scaffolds and/
or multiple metal ions in one molecule is advantageous, as
d. Furthermore, paramagnetic Mn2+ represents an attractive
alternative to Gd3+, generally used in MR contrast agents3.
Coordinated as the dipyridyl phosphate chelate 55Mn-DPDP “(see
a. Institute of Neuroscience and Medicine, INM-5: Nuclear Chemistry,
Forschungszentrum Jülich, Jülich, Germany
cFor explanation of the extensive physical justification of the relaxation
behaviour of metal complexes it is referred to appropriate literature.7-10
b. Institute of Radiochemistry and Experimental Molecular Imaging, University Clinic
Cologne, Cologne, Germany
*E-mail: j.ermert@fz-juelich.de
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Mastarone et al. and Zhu et al. reported on a trimeric Gd-
complex11 and a hexameric Mn-complex12, respectively.
position 1 will only marginally, if at all, influence theVcioewmAprtlieclxeaOtniolinne,
DOI: 10.1039/C8DT04996C
an isomer separation was not attempted and is not indicated in
the schemes given here.
Here, bimodal contrast agents with three trans-CDTA chelating
units labelled with the positron emitting radionuclide 52gMn and
paramagnetic 55Mn are presented. Based on our previous study
and results therein13, the functional group for the coupling
reaction was inserted at position 4 of the CDTA backbone, in order
to minimize ring strain and steric effects on the coordination sites.
CuI
Pd(PPh3)2Cl2
OEtVal
TMS
OEtVal
TMS
OH
OVal
EtVal-Br
Br
Br
K2CO3 Br
Br
KF
EtOH,
RT, 18 h
97%
TMS
Et3N,
75°C, 18 h
79%
DMF,
50 °C, 24 h
98%
Br
Br
TMS
5b
O
EtVal =
O
However, upon derivatization of the cyclohexane ring a similar
effect must be taken into account as observed before, marginally
impairing the thermodynamic complex stability, while increasing
the dissociation half-life of the substituted complex. The rigidity of
the trimeric complex with three paramagnetic Mn ions results in
an enhanced longitudinal relaxivity r1, while on the other hand the
aminocarboxylate chelator scaffold provides sufficient complex
stability. In favour of enhancing the rigidity of the molecule by
limiting the intermolecular rotation, the connection between the
CDTA backbone and the aromatic centre of the trimer was kept as
short as possible.
Scheme 2: Synthetic route to triethynylbenzene 5b with ethyl valerate
functionalization, according to Mastarone et al.11
Click coupling between
4 and the commercially available
triethynyl-benzene 5a or the functionalized triethynylbenzene 5b
was performed in a THF/water mixture with copper(II) sulfate and
sodium ascorbate as reducing agent (see Scheme 3). The
progression of the reaction was monitored by mass spectrometry.
Although 1,3-dipolar cycloadditions are known as rapid reactions
converting alkynes and azides into triazoles in the presence of only
catalytic amounts of copper, here equimolar amounts of copper
and ascorbate had to be employed. Additionally, quantitative
conversion to the cycloaddition product was not observed before
7 days. The long reaction time can be attributed to steric hindrance
and/or interaction between the Cu(II) ions and the chelating units,
although these were protected. Experiments with different copper
sources (CuI and Cu(MeCN)4·PF6) or other reducing agents /
additives, like 2,6-lutidine, L-histidine and tris(benzyltriazolyl-
methyl)amine (TBTA) did not result in higher yields or shorter
reaction times. In addition, the application of other solvents
(dichloromethane, tetrahydrofuran) was ineffective. A higher
reaction temperature even turned out to be counterproductive
due to decomposition of the starting material.
The trans-CDTA azide 4 was synthesized in five steps from racemic
cyclohexene-1-methanol with an overall yield of 12% as previously
reported (see Scheme 1).13
1. TBDPSCl, imidazole
N3
N3
NH2
NH2
83%
H2 , Pd/C
MeOH
93%
2. Mn(OAc)3, NaN3
CH3CN/TFA, -20°C
50%
OH
OTBDPS
OTBDPS
1
2
CO2tBu
CO2tBu
CO2tBu
CO2tBu
CO2tBu
CO2tBu
1.
BrCH2CO2tBu
K2CO3, DMF, 50°C
63%
N
N
N
N
1.
MsCl, Et3N
2.
TBAF, THF
81%
2.
NaN3
65%
CO2tBu
CO2tBu
COOtBu
OH
N3
N
N
COOtBu
COOtBu
3
4
N3
Scheme 1: Synthetic route to obtain the azide functionalized CDTA unit 4,
according to Vanasschen et al.13
4
COOtBu
N
N
R
N
N
R
N
CuSO4, NaOAsc
THF, H2O
N
N
The triethynyl unit 5b was produced according to Mastarone et al.
via Sonogashira coupling of ethyl valerate functionalized
tribromophenol with trimethylsilyl acetylene followed by
deprotection (see Scheme 2).11
50 °C, 7 d
6a-b
6a Y = 29%
6b Y = 25%
N
N
5a-b)
a) R = H
COOtBu
b) R = O(CH2)4CO2CH2CH3
It is important to consider that the introduction of the azide groups
in the second reaction step is trans-selective. Since racemic
cyclohex-3-en-1-ylmethanol was used as starting material, a
mixture of four trans-3,4-diazide isomers was formed. NMR-
analysis clearly showed an equatorial conformation of the azide
groups in solution, yielding diastereomeric pairs with 90% of the
side chain in axial and 10 % in equatorial position. The latter
products could be removed completely by repeated column
chromatography. This finding is in accordance with Habala et al.14
and Galanski et al.15, and proof is also further discussed in the
Supporting Info of Ref. 13.). As the absolute configuration of
N
N
COOtBu
COOtBu
=
COOtBu
Scheme 3: Click chemistry procedure to synthesize the trimeric CDTA
structures 6a and 6b.
Regarding the subsequent reaction steps and the high tendency of
Cu(II) ions to form complexes with tBu-protected CDTA ligands,
Cu(II) had to be completely removed before the deprotection of
the ester groups and the radiolabelling step. Therefore, a thorough
2 | Dalton Trans., 2018, 00, 1-3
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purification method of the trimeric ligand was necessary. To this
end, the crude reaction mixture was incubated with DTPA for 24 h
and the residue was purified by column chromatography on
Chelex®100 resin. After that, column purification on silica was
carried out to remove an excess azide and other side products.
difficult due to their size, polarity and multiple charges. After
DOI: 10.1039/C8DT04996C
different stationary phases, buffers and eluents were examined,
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HPLC purification was carried out on a reversed phase column,
using a mixture of PBS buffer in acetonitrile as eluent. This method
allowed separating the radioactive complexes sufficiently. The
chromatograms showed only one radioactive component, which
was identified as the intact radio complex by co-elution with the
isotopic (non-radioactive) 55Mn-compounds.
The trimeric structures 6a and 6b were obtained in moderate
yields of 29% and 31%, respectively. Quantitative deprotection of
the tBu-protected carboxylate groups succeeded by treatment
with trifluoroacetic acid. Using this method, two different trimeric
structures with trans-CDTA-units were obtained. Additionally, a
trans-CDTA trimer with an ethyl valerate moiety (TTB-CDTA, 7b)
was received, bearing a potential binding site for the conjugation
of targeting molecules.
O
O
O
6+
O
N
Mn*
N
O
O
O
O
N
N
COOH
COOH
N
COOH
N
N
O
R
O
O
O
COOH
O
O
N
N
N
O
N
O
N
N
N
N
N
N
Mn*
Mn*
N
N
O
O
N
O
O
HOOC
HOOC
N
N
O
COOH
COOH
N
N
HOOC
O
N
O
N
52g/55
52g/55
N
7a
Mn][Mn( )]R = H
[
[
N
O
N
N
7b
Mn][Mn( )]R = O-(ethyl)valerate
N
7a
TTB-(CDTA)3
COOH
COOH
HOOC
COOH
COOH
N
HOOC
HOOC
Figure 4: Molecular structure of isotopically labelled
52g/55Mn][Mn(7b)]
[
52g/55Mn][Mn(7a)]) and
[
N
N
N
O
O
O
To determine the longitudinal relaxivity r1 of the trimeric Mn-CDTA
complexes, T1 of cold [Mn3(7a)] was measured at four different
concentrations at 25 °C and 37 °C. The longitudinal relaxation time
T1 correlates with the concentration [CA] according to eq. 1
HOOC
HOOC
N
N
N
HOOC
N
N
N
N
N
N
7b
EtVal-TTB-(CDTA)3
HOOC
COOH
N
HOOC
1
1
1
1
HOOC
COOH
=
푟1[퐶퐴]
―
= 푟1[퐶퐴]
(1)
or
푇1
푇1,0
푇1
푇1,0
Figure 2: Structures of free acids 7a and 7b after deprotection.
1
Linear fitting of the corrected relaxation rates (푇1
) against
―
Labelling of the trimeric chelator systems 7a and 7b (see Fig. 2)
with 55Mn, spiked with 52gMn, was performed in 1 M NaOAc buffer
at room temperature according to the protocol reported recently5.
After a reaction time of 30 min, no free non-chelated 52gMn2+ ions
were detected by radio-TLC in all cases (RCY > 99%, see Fig. 3; for
additional information concerning the radio-TLC procedure, see
Vanasschen et al.5).
푇1,0
1
the concentration of contrast agent [CA] gave r1 as slope (see Fig.
1
5), where 푇 is the longitudinal relaxation rate of pure water.
1,0
Figure 3: Radio-TLC images of [52gMn][Mn(7a)] (left) and [52gMn][Mn(7b)]
Figure 5: Linear regression of Mn-concentration against corrected
longitudinal relaxation rate.
(centre) in comparison to [52gMn]Mn2+ (right).
Due to the rigidity of the trimeric complex relaxivity values of
8.8 mM-1s-1 per paramagnetic ion at 25 °C and 6.6 mM-1s-1 at 37 °C,
respectively, were measured. This value corresponds to an
enhancement of 144% in comparison to the [Mn(CDTA)]2-
monomer (see Table 1). The relaxivity values per molecule
The radiochemical yield (RCY) was calculated as percentage of
activity in the product fraction related to the activity in an aliquot
of the reaction mixture (in Fig. 3 marked as ‘Control sample’) as
determined by radio-TLC or radio-HPLC, respectively. HPLC
purification of the radiolabelled complexes 7a,b (see Fig. 4) was
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(26.4 mmol-1s-1 at 25 °C and 19.8 mmol-1s-1 at 37 °C) exceeded the
relaxivity of the Mn-CDTA monomer by more than 630%. These
results are in good accordance with relaxivity values determined
for a hexameric [Mn(EDTA)]2- complex reported by Zhu et al.12.
Notably, the stability of the Mn-CDTA-complex is much higher than
the stability of corresponding EDTA chelates. Therefore, Mn-CDTA
complexes are significantly better suited for in vivo applications.
were energy and efficiency calibrated with certified radiation point
DOI: 10.1039/C8DT04996C
sources (Co-60, Ba-133, Eu-152, Ra-226) from the Physikalisch-
Technische Bundesanstalt (Braunschweig, Germany).
Radio-TLC measurements were carried out on 60F254 silica plates
(Merck, Darmstadt, Germany), using 25% aq. NH3/MeOH/H2O
2:1:1 as mobile phase; radio-TLC were recorded using a Packard
Instant Imager (Packard Instrument Company, Meriden, CT, USA).
The radioactivity detection limit (LOD) was determined by serial
dilution: LOD ≤ 3 Bq for an exposure time of 30 min.
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Table 1: Longitudinal relaxivities of relevant contrast agents
20 MHz
Contrast agent
25 °C
4.7
7.4
37 °C
3.8
6.8
8.2
Reference
Radio-HPLC was performed on a Knauer Azura P 4.1S pump, a
Knauer Azura UVD 2.1S UV/VIS detector (Knauer Wissenschaftliche
Geräte GmbH, Berlin, Germany) and an EG&G Ortec ACE NaI(Tl)
radioactivity detector with photomultiplier (EG & G Ortec, Oak
Ridge, TN, USA). The radioactivity detection limit was determined
by serial dilution: LOD ≤ 0.3 kBq. As mobile phase PBS/CH3CN 1:1
(PBS = phosphate buffered saline, pH 7.4) and as RP stationary
phase Synergi 4µ polar RP 80 Å, 250 x 4.6 mm (Phenomenex Inc.,
Aschaffenburg, Germany) were used for co-elution experiments of
radioactive and non-radioactive complexes with a flow rate of 0.7
mL/min. The UV detection wavelength for all measurements was
230 nm.
Dotarem
Mn(H2O)6
16
17
12
2+
Mn(EDTA)-hexamer
Mn(CDTA)
[Mn(7a)]
3.6
8.8
18
6.6
this work
Conclusions
In continuation of our previous studies5,13 a novel synthetic
strategy is presented here towards bimodal PET/MR contrast
agents with high T1 relaxivity.
NMR spectra were measured on
a Bruker Avance 600
Two trimeric compounds were synthesized, bearing three CDTA
chelating units. One compound contains an additional
functionalization for the conjugation of specific biomolecules to
address specific molecular targets.
spectrometer with 600.15 MHz (1H-NMR) and 150.9 MHz (13C-
NMR) (Bruker BioSpin GmbH, Rheinstetten, Germany) or a Varian
Inova 400 spectrometer (Agilent Technologies, Waldbronn,
Germany) with 400.1 MHz (1H-NMR) and 100.62 MHz (13C-NMR),
respectively.
Relaxivity measurements on [Mn3(TTB-CDTA)] ([Mn3(7a)] showed
a considerably enhanced T1 relaxivity compared to the Mn-CDTA
monomer. Thus, this compound should enable higher sensitivity
due to the higher number of NMR-active Mn ions and enhanced T1
relaxivity. Furthermore, radiolabelling of the trimeric compound
The longitudinal relaxation times of water protons (T1) were
measured at 20 MHz on a Bruker NMS 120 Minispec apparatus,
using the inversion recovery method (180° – τ – 90°). The
temperature of the samples was kept at 25.0 ± 0.2 °C or 37 ± 0.2 °C
and controlled with a circulating water bath.
High-resolution mass spectrometry was performed using a hybrid
linear ion trap FT-ICR mass spectrometer LTQ-FT Ultra (Thermo
Fisher Scientific, Bremen, Germany) equipped with
delivered
[
[
52gMn][Mn(TTB-CDTA)
([52gMn][Mn(7a)])
in
and
high
52gMn][Mn(val-TTB-CDTA)
([52gMn][Mn(7b)])
radiochemical yields providing the basis for further
characterization and PET/MR evaluation.
a 7 T
superconducting magnet. The electrospray ionisation (ESI) source
was operated in the positive or negative mode, respectively.
Low-resolution mass spectrometry was measured using a Finnigan
Automass Multi spectrometer (Thermoquest, Biberach, Germany).
55Mn concentrations in solution were determined by inductively
coupled plasma mass spectrometry (ICP-MS) using an Agilent 7500
instrument with quadrupole mass analyser and collision cell
(Agilent Technologies, Frankfurt am Main, Germany). The
instrument was operated with helium as collision gas to minimize
spectral interferences by cluster ions. Quantification was
performed by external calibration using Rh as the internal
standard.
Experimental
Chemicals
PBS buffer was purchased from Life Technologies (Thermo Fisher
Scientific, Schwerte, Germany), Chelex®100 resin from Bio-Rad
(München, Germany) and organic solvents for HPLC from VWR
(Langenfeld, Germany). All other chemicals were obtained from
Sigma Aldrich GmbH (Schnelldorf, Germany) and used without
further purification.
Production of n.c.a. [52gMn]MnCl2
For centrifugation, a small 5415 C Centrifuge from Eppendorf
(Hamburg, Germany) or a Heraeus Biofuge Stratos centrifuge
(Thermo Fisher Scientific Inc., Schwerte, Germany) was used.
52gMn was produced as reported earlier19a,b by proton irradiation
of a natCr target followed by ion exchange chromatography.
Instruments
Organic syntheses
Gamma spectroscopy was performed with ORTEC gamma-ray
spectrometers (AMETEK GmbH, Meerbusch, Germany), which
Tetra-tert-butyl-4-(azidomethyl)-trans-1,2-diaminocyclohexan-
N,N,N’,N’-tetraacetate (4) was synthesized according to
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Vanasschen et al.13 5-(2,4,6-triethinylphenoxy)-pentanoic acid
ethyl ester (5) was produced according to Mastarone et al.11
trifluoroacetic acid (TFA) at room temperature. After 18 h, 10 mL
DOI: 10.1039/C8DT04996C
Et2O was added to the reaction vessel. The resulting suspension
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was transferred to a falcon tube and centrifuged. After removal of
the supernatant, the yellowish solids were washed with 2 x 5 mL
Et2O and dried on the air. Compounds 7a and 7b were obtained as
TFA salts in quantitative yields (7a: 15 μmol, 22 mg; 7b: 15 μmol,
25 mg).
Due to their poor solubility, the free ligands were transferred into
the corresponding zinc complexes for NMR spectroscopy. For this,
2.2 µmol (≈3 mg of 7a/7b) were suspended in 1 mL water and the
pH adjusted to 7, using small amounts of 1 M NaOH. To that 105
µL of a 0.11 M ZnCl2 (11 µmol, 5 eq.) solution was added and the
pH readjusted to 6. After 18 h of stirring, the solution was passed
through a Millex® Millipore filter and the solvent was removed.
The crystalline residue was analysed without further purification.
7a (NMR as zinc complex).
1H-NMR (400 MHz, D2O) δ: 8.21 (s, 3H, Htriazin); 7.82 (s, 2H, Harom);
4.34 (s, 6H,HCHN), 3.58 – 2.86 (m, 24H,12 x CH2), 2.65 – 2.30 (m, 9H,
Haliph), 1.90 – 1.30 (m, 12 H, Haliph).
13C-NMR (151 MHz, D2O) δ: 180.87, 177.99, 177.83, 177.67,
176.73, 171.05, 166.83, 161.52, 160.44, 153.06, 148.72, 146.59,
143.16, 131.02, 129.93, 122.57, 122.22, 63.29, 59.09, 52.19, 51.63,
50.91, 33.64, 25.40, 24.36, 19.06. HR-MS (FT-ICR, free ligand,
negative mode): 1352.50243 ([M-H]-·, calculated 1352.50366 for
C57H74N15O24-); 675.74818 [M-2H]2-(calculated 675.74819 for
C57H73N15O242-).
Click chemistry procedure
To obtain TTB-(CDTAtBu)3 6a and EtVal-TTB-(CDTAtBu)3 6b, 0.17
mmol (115 mg, 3.3 eq.) of the azide 4 was dissolved in 4.5 mL THF.
0.05 mmol (1 eq.) of the triethynyl unit (5a or 5b) in 0.5 mL THF
was added to the reaction vessel, followed by 700 µL (0.25 mmol.
5 eq.) of a 0.34 mM copper(II)sulfate solution and 127 mg
(0.64 mmol, 12 eq.) sodium ascorbate in 800 µL water. The
reaction vial was closed with a septum and the mixture stirred at
50 °C. The progress of the reaction was monitored by mass
spectrometry. After completion at ≈ 7 d, the reaction mixture was
poured into a mixture of 10 mL 0.4 M DTPA solution and 20 mL
EtOAC and stirred vigorously for 24 h. Upon phase separation, the
DTPA extraction was repeated for another 24 h. Then, phase
separation, the solvent was removed and the organic residue was
suspended in 10 mL of MeOH and rinsed through a short column
loaded with Chelex®100 resin to remove the remaining copper
traces. Following the evaporation of MeOH, the crude product was
purified via column chromatography on silica (100% EtOAc, Rf =
0.1). The products were obtained as yellow-brownish gels.
6a Yield: 29.0 mg, 15 µmol, 29%.
1H-NMR (600 MHz, CDCl3) δ: 8.05-8.04 (m, 3H, Htriazol); 7.72-7.71
(m, 3H, Harom); 3.51-3.37 (m, 32H, Haliph); 3.11 (s, 2H, Haliph); 3.02 (s,
2H, Haliph); 2.94 (s, 2H, Haliph); 2.35-1.60 (m, 21H, Haliph); 1.44 (s,
120H, Haliph).
7b (NMR as zinc complex).
1H-NMR (400 MHz, D2O) δ: 8.42 (s, 1H, Htriazin); 8.31 (s, 2H, Htriazin);
8.12 (s, 2H, Harom); 4.08 (s, 4H,Haliph), 3.80 – 3.70 (m, 10H, Haliph);
3.36 (m, 18H, Haliph); 2.90 (s, 6H, Haliph); 2.73 (s, 4H, Haliph); 2.58 (2,
3H,Haliph); 2.70 (m, 5H, Haliph); , 2.19 (m, 3H, Haliph), 1.95 – 1.70 (m,
18 H, Haliph); 1.51 – 1.39 (m, 18H, Haliph); 1.18 (m, 3H, Haliph).
13C-NMR No resilient 13C-NMR could be measured due to poor
solubility in any solvent.
13C-NMR (151 MHz, CDCl3) δ: 172.5; 170.5; 158.2; 157.0; 134.6;
133.8; 128.6; 126.7; 124.5; 79.6; 72.69; 65.8; 53.1; 46.0; 41.0; 40.4;
39.8; 37.7; 34.4;, 33.0; 30.9; 29.3; 28.7; 28.0; 27.9; 27.1; 25.6; 23.5;
22.7; 21.9; 21.7; 21.6; 19.41; 18.4; 13.4; 13.3.
ESI-MS (m/z): 1014.78 ([M+2H]2+; calculated 1014.64002 for
C105H173N15O242+); 676.89 ([M+3H]3+, calculated 676.76244 for
C105H174N15O243+).
MS (MALDI-TOF, free ligand): 1493.8 ([M+H]+; calculated
1493.55745 for C70H94N9O27+).
6b Yield: 34.0 mg, 17 µmol, 33%.
1H-NMR (600 MHz, CDCl3) δ: 8.49 (s, 2H, Htriazol), 8.14 (s, 2H, Harom);
7.89 (s, 1H, Htriazol); 4.40-4.20 (m, 2H, Haliph), 4.11-4.06(m, 4H,
Haliph), 3.56 (s, 4H, Haliph); 3.50-3.33 (m, 20H, Haliph); 3.10 (s, 1H,
Haliph); 3.01 (s, 1H, Haliph); 2.92 (s, 3H, Haliph); 2.82 (s. 1H, Haliph);
2.75(s. 1H, Haliph); 2.42-2.10 (m, 9H, Haliph); 2.04-1.55 (m, 22H,
Haliph); 1.43-1.40 (m, 116H, Haliph); 1.22 (t, 3J = 7.5 Hz, 3H, HCH3).
13C-NMR (151 MHz, CDCl3) δ: 171.54; 171.53; 159.5; 158.2; 126.2;
125.8; 123.3; 80.9; 68.7; 68.3; 60.5; 54.3; 47.2; 43.3; 42.1; 41.6;
41.0; 37.8; 35.6; 34.0; 33.8; 29.9; 29.5; 28.3, 22.9; 21.7; 14.7; 14.4;
14.3; 12.11.
Syntheses of [Mn3(7a)] and [Mn3(7b)]
2.2 µmol (≈3 mg of 7a/7b), respectively, were suspended in 1 mL
water and the pH adjusted to 7 using small amounts of 1 M NaOH.
To that 420 µL of a 15 mM MnCl2·4H2O (6.3 µmol; 0.95eq per
chelator unit) solution was added and the pH readjusted to 6. After
18 h of stirring, the solution was passed through a Millex®
Millipore filter and the solvent was removed. The residues were
taken up in 5 mL of water and filtered through short Chelex®100
columns in order to remove any traces of free manganese. The
high NaCl content in the product did not allow to determine the
yield of [Mn3(7a,b)] and Zn-complexes of 7a,b gravimetrically.
TLC (25% NH3(aq)/MeOH/H2O 2:1:1) : Rf [Mnx(7a)]y- : 0.95; HPLC Rt
[Mnx(7a)]y-: 3.13 min.20
ESI-MS m/z: 1087.12 ([M+2H]2+ calculated: 1087.67935 for
C112H185N15O272+); 725.39 ([M+3H]3+ calculated: 724.78866 for
C112H185N15O273+).
TLC (25% NH3(aq)/MeOH/H2O 2:1:1) : Rf [Mnx(7b)]y-: 0.95; HPLC Rt
[Mnx(7b)]y-: 3.15 min.20
Deprotection procedure
To obtain TTB-(CDTA)3 7a and EtVal-TTB-(CDTA)3 7b: 15 µmol
(29 mg 6a, 34 mg 6b) of the protected ligands were stirred in 2 mL
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Dalton Trans., 2018, 00, 1-3 | 5
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COMMUNICATION
Journal Name
Radiochemistry
Ramsay, Y.-F. Yen, D. E. Sosnovik, P. Caravan, Radiology 2018, 286,
865-872; c) K. Pota, Z. Garda, F. K. KáDlmOáI:n1,0.J1.03LV9.i/eCBwa8ADrrrtTiica0led4Oa9,9nl6iDnCe.
Esteban-Gomez, C. Platas-Iglesias, I. Tóth, E. Brücher, and G.
Tircso, New J. Chem. 2018, 42, 8001-8011.
Radiosyntheses of n.c.a.
[
52gMn][Mn(7a)] and n.c.a.
[
52gMn][Mn(7b)]
5 mg (3.3 µmol) of the free ligands 7a or 7b were dissolved in
500 µL 1 M NaOAc buffer (pH = 6), ≈20 MBq [52gMn]MnCl2 added
as aqueous solution (20 µL) and the mixture was stirred until
complete consumption of Mn. The reaction was monitored by
RadioTLC.
TLC (25% NH3(aq)/MeOH/H2O 2:1:1) : Rf [52gMn][Mnx(7a)]y-: 0.95.
Rt[52gMn]2+ = 0.
HPLC Rt [52gMn][Mnx(7a)]y-: 3.13 min.
RCY: > 99% (no detection of free [52gMn]Mn2+).
TLC (25% NH3(aq)/MeOH/H2O 2:1:1) : Rf [52gMn][Mnx(7a)]y-: 0.95;
Rt[52gMn]2+ = 0.
HPLC Rt [52gMn][Mnx(7a)]y-: 3.15 min.
RCY: > 99% (no detection of free [52gMn]Mn2+).
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7746–7760.
Radiosyntheses of c.a. [52gMn][Mn(7a)] and c.a.[52gMn][Mn(7b)]
5 mg (≈3.3 µmol) of the free ligands 7a or 7b were dissolved in 500
µL 1 M NaOAc buffer (pH = 6), 1 eq. (3.3 µmol) MnCl2 added as
aqueous solution and the pH readjusted to 6 by addition of small
amounts 0.1 M NaOH. Then, ≈20 MBq [52gMn]MnCl2 were added
as aqueous solution (20 µL), and the mixture was stirred until
complete consumption of Mn. The reaction was monitored by
RadioTLC (analytical data see above).
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Acknowledgement
The authors thank Stefan Spellerberg for nuclide production and
Kai Giesen for support with organic syntheses.
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6 | Dalton Trans., 2018, 00, 1-3
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