Direct reductive alkylation of amino acids: Synthesis of bifunctional chelates for nuclear imaging

Article abstract of DOI:10.1055/s-2004-829120

A family of effective bifunctional chelators for technetium- and rhenium-based radiopharmaceuticals was conveniently synthesized in high yields through direct reductive N-alkylations of amino acids and their analogues with aldehydes, using NaBH(OAc)₃ as an efficient reagent. The mono-, di-, tetra- and even mixed alkylated amino acid derivatives were all prepared in one-pot synthesis.

Full text of DOI:10.1055/s-2004-829120

PAPER
1759
Direct Reductive Alkylation of Amino Acids: Synthesis of Bifunctional
Chelates for Nuclear Imaging
S
ynthesis of Bifunction
u
al Chelates
ra
a
Department of Chemistry, Syracuse University, Syracuse, NY 13244, USA
Fax +1(315)4434070; E-mail: jazubiet@syr.edu
b
Molecular Insight Pharmaceuticals, Inc., 160 Second St., Cambridge, MA 02142, USA
Received 30 February 2004; revised 30 March 2004
the side chain of amino acids. Such amino acid analogues
provide a tridentate donor set for chelation and an amino
acid functionality for attachment to biomolecules. We re-
cently developed a family of such single amino acid che-
Abstract: A family of effective bifunctional chelators for techne-
tium- and rhenium-based radiopharmaceuticals was conveniently
synthesized in high yields through direct reductive N-alkylations of
amino acids and their analogues with aldehydes, using
NaBH(OAc)3 as an efficient reagent. The mono-, di-, tetra- and lates (SAAC) that serve this function14,15 and can be
even mixed alkylated amino acid derivatives were all prepared in
readily incorporated into peptides via solid-phase synthe-
sis techniques.16
one-pot synthesis.
Key words: reductive alkylation, amino acids, bifunctional chela-
This approach affords significant flexibility in the choice
tors, radiopharmaceuticals, sodium triacetoxyborohydride
of donors for 99mTc coordination, combined with the con-
siderable advantages provided by application of routine
solid-phase synthetic techniques. Most significantly, the
Radiolabeled bioactive peptides are attractive vectors for
addition of a chelating moiety as a single amino acid sub-
targeting a variety of diseases through interaction with
stitution (or addition) to the peptide sequence should min-
specific cell surface receptors.1 Introduction of a molecu-
imize the perturbation of structure and function compared
lar entity that allows facile labeling with medically useful
to more elaborate chelating systems which are comprised
radionuclides such as Tc-99m for diagnosis and Re-186/
of three to four amino acids in addition to chemical tethers
188 for targeted therapy, without significant alteration of
or spacers. However, the exploitation of this approach to
the biopolymer’s structure and binding affinity, is desir-
labeling requires facile, high yield syntheses of the deriva-
able.2 Current approaches make use of polypeptides that
tized amino acids, preferably without isomerization.
can form chelate complexes with the radiometals. While
One synthetic approach involves reductive amination of
these peptidic chelates have been shown to provide a
aldehydes and ketones, a very useful reaction in modern
means of complexing both Tc and Re as the {M(V)-
organic preparations. In the case of amino acids, the re-
ductive alkylation with aldehydes is generally performed
after protection of the acid functionality as the ester,17 ren-
dering this a cumbersome three-step procedure. The re-
ports that detail the reductive alkylation of amino acids
with an open acid functionality (direct) are surprisingly
few. The preferred reagent for this purpose is
NaCNBH318, while NaBH419, H2/C-Pd(OH)2,20 and
NaHTe21 have occasionally been used. Although
NaCNBH3 is an excellent reagent, it is relatively toxic and
the isolated products are often contaminated with cyanide
residue. One limitation is that the reactions are restricted
to polar medium, which is sometimes sluggish and often
mitigated by poor yields. Furthermore, the scope of the
substrate is limited in heterogeneous hydrogenations as
many functional groups, such as nitro, alkene, alkyne,
OBn etc., are susceptible under the reaction condition.
The use of NaBH4 necessitates the preformed imine to
avoid the reduction of aldehyde, and also alkaline medi-
um, thereby limiting its practicality. Abdel-Magid et al.22
have recently demonstrated that NaBH(OAc)3 can effec-
tively be used as a mild reagent in the reductive amination
of aldehydes and ketones with shorter reaction time and
excellent yields. It is surprising, that with the exception of
a single example, there are no major reports of use of
oxo}+3 core,3–8 they suffer from limitations related to met-
al complex instability and to the uncertainty of metal com-
plex formation with the proposed metal binding sequence
versus the sequence responsible for receptor binding. Al-
ternatives to peptidic chelates, such as hydrazinonicotinic
acid (HYNIC), which is introduced commonly through
acylation of a lysine residue, provide more stable com-
plexation of Tc over peptidic chelates.9–11 However, the
coordination chemistry of the Tc-HYNIC complex is such
that metal binding sites are not completely saturated by
HYNIC, allowing both multimodal binding as well as
coligand lability and exchange which significantly influ-
ence the target and nontarget localization of the radiola-
beled peptide.12
Technetium tricarbonyl chemistry has recently been the
subject of considerable interest in radiopharmaceutical
development.13 Previous studies on the coordination
chemistry of the {M(CO)3}+1 core has established amine,
aromatic heterocyclic, and carboxylate donors as very ef-
fective chelating ligands.14 These observations led to the
design of tridentate chelators constructed as extensions of
SYNTHESIS 2004, No. 11, pp 1759–1766
x
x.
x
x
.
2
0
0
4
Advanced online publication: 01.07.2004
DOI: 10.1055/s-2004-829120; Art ID: M00904SS
© Georg Thieme Verlag Stuttgart · New York

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M. K. Levadala et al.
PAPER
NaBH(OAc)3 with free amino acids,23e even though there
are several reports in the literature for the corresponding
amino acid esters.23 We sought to exploit the above meth-
od to provide direct access to ligands useful for applica-
tions to nuclear medicine.
As part of our ongoing efforts to develop practical prepar-
ative methods for a broad-spectrum of bifunctional
ligands for nuclear imaging, we noted literature reports of
reductive alkylation of phenylalanine with pyridine-2-car-
boxaldehyde for which a 21% yield with NaCNBH3 had
been obtained for the monoalkyl derivative.18b We have
now investigated the dialkylation of the same system with
NaBH(OAc)3 in 1,2-dicholorethane (DCE) (Scheme 1).
The reaction was complete within two hours, and the
product 6 was isolated in 78% yield. Since DCE has been
used in similar reactions, we employed the same medium,
despite the insolubility of amino acids in DCE. It is note-
worthy that the reaction was homogeneous in DCE de-
spite the insolubility of amino acids while a suspension-
like mixture was evident in the polar solvent MeCN. The
rate of the reaction was significantly faster in chlorinated
solvents, DCE and CH2Cl2 compared to THF and MeCN.
The dipicolyl derivative of L-phenylalanine (compound 6)
was crystallized and structurally characterized by X-ray
crystallography. Figure 1 shows an ORTEP view of the
structure.24
Figure 1 An ORTEP view of the structure of 6, showing the atom-
labeling scheme and 50% probability ellipsoids.
kylation, the lysine dihydrochloride salt was treated with
4.5 equivalents of pyridine-2-carboxaldehyde and
NaBH(OAc)3 (5 equivalents) to afford the tetrapyridyl de-
rivative 10 in 79% yield, along with a small amount of the
reduced alcohol of the aldehyde. It is noteworthy that four
alkylation steps are performed in one-pot in the case of
such tetraalkyl amino acids, an observation consistent
with the direct use of the amino acid hydrochloride salt as
starting materials without any need for prior neutraliza-
tion or base addition to the reaction, at the expense of a
slight excess of aldehyde (0.5 equiv). In order to deter-
mine the generality of the protocol, we investigated the re-
ductive amination of a noncoordinating aryl aldehyde.
The required dialkylated product 11 was isolated in 74%
yield, in addition to the monoalkylation product in 12%
yield.
The preparative results of the reductive alkylation of var-
ious amino acids and their derivatives with different alde-
hydes are summarized in Tables 1 and 2. The amino acids
and heterocyclic aldehydes were chosen such that the
products could ultimately be used as chelating agents in
nuclear imaging with 99mTc. By appropriate stoichiomet-
ric manipulation of the amino acids and aldehydes, the
mono-, di-, and even tetraalkylated amino acid derivatives
could be obtained in good yields. In the case of tetraal-
Table 1 Preparative Di- and Tetraalkylation
Product
1
Amino Acid
Aldehyde
A
Producta
Time (h)
1
Yieldb (%)
86
Na-Fmoc-L-lysine
2
3
Na-Fmoc-L-lysine
Na-Fmoc-L-lysine
B
C
1
2
84
82
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PAPER
Synthesis of Bifunctional Chelates for Nuclear Imaging
1761
Table 1 Preparative Di- and Tetraalkylation (continued)
Product
Amino Acid
Aldehyde
E
Producta
Time (h)
2
Yieldb (%)
4
Na-Fmoc-L-lysine
86
5
6
Ne-Boc-D-lysine
A
A
1
2
83
78
Phenylalanine
7
8
Glycine
A
A
2
2
76
76
Valeric acid
9
Na-Fmoc-L-lysine
D
A
0.5
75
10
L-Lysine·2HCl
4
79c
11
Fmoc-D-lysine
F
2
74d
a New compounds gave satisfactory analytical data.
b Isolated yield.
c 12% of pyridin-2-methanol was also obtained.
d 10% of monoalkylated product was also isolated.
The simplified workup procedure involves the addition of Encouraged by the above results, we then investigated the
water to quench the reaction followed by extraction into monoalkylation of similar systems with stoichiometric
the organic phase. The methodology was also amenable to amounts of aldehyde and amine (compounds 12, 13). Un-
large scale synthesis, as we have successfully scaled up fortunately, the reaction resulted in the predominant for-
the reductive alkylation of amino acids (compound 1) to a mation of dialkylated product. However, it was
10 g scale.
subsequently found that the dialkylation could be signifi-
cantly suppressed by forming the imine25 in situ, i.e., re-
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M. K. Levadala et al.
PAPER
Table 2 Representative Monoalkylation and Mixed Dialkylation Products of This Study
Product
Amino Acid
Valeric acid
RCHO
A
R1CHO
-
Producta
Time (h)
0.5
Yieldb (%)
12
71
13
14
Na-Boc-D-lysine
Na-Boc-D-lysine
A
A
-
0.5
2
73
B
75
73
70
15
16
Na-Boc-D-lysine
A
A
C
D
2
2
Na-Fmoc-D-lysine
a New compounds gave satisfactory analytical data.
b Isolated yield, the corresponding dialkylated product was isolated in 10–15% yields in all cases.
fluxing the mixture of amino acid and aldehyde in DCE minutes, followed by addition of NaBH(OAc)3 and the
for 10 minutes under inert atmosphere (Scheme 2), fol- second carboxaldehyde at ambient temperature in one pot
lowed by treatment with NaBH(OAc)3 at ambient temper- to obtain the dialkyl derivatives in good yields (Table 2).
ature. This enabled us to isolate the monoalkylated
products 12 and 13 in good yields with the formation of
Representative rhenium complexes were prepared in
quantitative yields by reacting (NEt4)2[Re(CO)3Br3]26
dialkylated products reduced to 10–15%. This small
with the ligands in methanol under reflux conditions.14
amount of dialkylation may be explained in light of the
The overall complex charges of the compounds of the
observation for amino esters by Abdel-Magid et al.23c that
general class [Re(CO)3L]n (n = –1, 0, +1) were deter-
the monoalkylated product itself would add to the imine to
mined by the identity of the donor groups. Thus, the
ligands 1–8, 14 and 15 provided cationic complexes,
while ligands 16 and 9, after deprotection of tert-butyl
form a dialkyliminium ion, which could then be reduced
by NaBH(OAc)3 to afford the dialkylated product. We
found that other aprotic solvents CH2Cl2, THF and MeCN
group followed by reacting with (NEt4)2[Re(CO)3Br3],
along with protic solvents such as methanol were equally
yielded neutral and anionic metal complexes, respective-
effective for the formation of imine.
ly. The 99mTc analogues were prepared by the reaction of
We next focused on mixed dialkylation of amino acids for [Tc(CO)3(H2O)3]+ with the ligands.27 The complex with
which there were no direct and single-step methods avail- ligand 1 was challenged in 1 mM cysteine and 1 mM his-
able in the literature. For this endeavor, the amino acid tidine in PBS buffer (pH 7.4) for 24 hours at 37 °C to de-
was refluxed with the appropriate aldehyde in DCE for 10 termine its stability against ligand exchange and
Scheme 1 Synthesis of the dialkyl derivatives of amino acids
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Synthesis of Bifunctional Chelates for Nuclear Imaging
1763
Scheme 2 General procedure for the synthesis of mono- and mixed-alkylated (R ºR1) derivatives of amino acids.
using standard pulse programs with gradients. HSQC spectra were
collected with 4 scans per increment, or 64 scans per increment (de-
pending on whether the natural abundance peaks were desired), into
2048 × 512 points with no sample spinning and no zero-filling.
Electrospray mass spectrometry (ES-MS) was performed on a
Fisons Platform quadrupole instrument where samples were dis-
solved in 1:1:1 mixture of MeOH–THF–MeCN. For compounds an-
alyzed in the positive ion mode, 3% AcOH was added. For
compounds run in negative mode, one drop of 0.10 M NaCl solution
was added. High-resolution masses are within 5 ppm of theortical
values. The crystal structure of ligand 6 was studied on a Bruker dif-
fractometer equipped with the SMART CCD system using graph-
ite-monochromated Mo-Ka radiation (l = 0.71073Å). The data
collection was carried out at 90(2) K. The data was corrected for
Lorentz polarization effects and absorption corrections were made
using SADABS. All calculations were performed using SHELXTL.
The structure was solved by direct methods and all of the non-hy-
drogen atoms were located from the initial solution. After locating
all the non-hydrogen atoms in the structure, the model was refined
against F2 initially using isotropic and later anisotropic thermal dis-
placement parameters until the final value of D/smax was less than
0.001. At this point the hydrogen atoms were located from the elec-
tron density difference map and a final cycle of refinements was
performed, until the final value of D/smax was again less than 0.001.
No anomalies were encountered in the refinement of the structure.
decomposition. The sample was analyzed after 1, 4 and 24
hours. Minimal decomposition (< 2%) was observed even
after 24 hours of incubation. The stable ‘closed’ shell oc-
tahedral coordination of {M(CO)3}+1, i.e., low spin d6
Re(I)/Tc(I), renders substitution reactions via either disso-
ciative or associative mechanisms unlikely. Furthermore,
preliminary biodistribution experiments with the repre-
sentative neutral, anionic and cationic complexes are even
more encouraging as no significant localization of the
complex in any organ of rat was observed.28 These results
suggested that this important family of bifunctional
ligands can be functionalized with different biologically
active molecules without significantly altering the biodis-
tribution properties of the drug molecule. Our recent
report16b demonstrated that the ligand could be conve-
niently incorporated into a small peptide sequences via
solid phase synthesis while retaining its expected biodis-
tribution properties. These latter results demonstrate the
versatility and numerous advantage of using the single
amino acid chelate system to prepare peptide-targeted
Tc(I) and Re (I) radiopharmaceuticals.
In conclusion, a mild method has been developed for the
direct reductive alkylation of amino acids and also for the
rapid assembly of useful bifunctional chelators for appli-
cation to nuclear medicine. The methodology offers nu-
merous advantages: preparative convenience, relatively
nontoxic reagents, base free workup, and mixed dialkyla-
tion in one pot. Our further efforts are directed towards
rapid generation of a diverse set of bifuntional chelators to
allow coordination for other metal radionuclides.
Dialkylation Reactions; 6-[Bis(pyridin-2-yl)methyl]amino-2-
(9H-fluoren-9-ylmethoxycarbonyl)amino-1-hexanoic Acid (1);
Typical Procedure
To a mixture of Fmoc-L-lysine (10.0 g, 27.1 mmol) and
NaBH(OAc)3 (14.4 g, 67.95 mmol) in DCE (150 mL), was added 2-
pyridinecarboxaldehyde (6.4 g, 57.0 mmol) in DCE (15 mL) at 0 °C
under argon. The suspension was stirred at r.t. for 1 h. The reaction
mixture was decomposed with H2O (100 mL) and diluted with
CHCl3 (100.0 mL). The separated organic layer was washed with
H2O and brine, dried (Na2SO4) and concentrated under reduced
pressure. The residue was purified through a pad of silica gel using
MeOH–CHCl3 (1:6) as eluent to provide the dipyridylmethyl deriv-
ative of Fmoc-L-lysine (12.85 g, 86%).
1H NMR (300 MHz, CDCl3): d = 10.85 (br s, 1 H), 8.50 (d, J = 5.10
Hz, 2 H), 7.70 (d, J = 7.24 Hz, 2 H), 7.55 (m, 4 H), 7.46 (m, 2 H),
7.32 (t, J = 7.72, 2 H), 7.22 (t, J = 7.52, 2 H), 7.09 (m, J = 6.20 Hz,
2 H), 6.0 (br s, 1 H), 4.29 (m, 3 H), 4.17 (t, J = 6.20 Hz, 1 H), 3.86
(br s, 4 H), 2.57 (t, 2 H), 1.90–1.20 (m, 6 H).
All reagents were weighed in air and the reactions were conducted
under an atmosphere of air, unless otherwise indicated. Anhydrous
DCE, pyridine-2-carbaldehyde, thiazole-2-carbaldehyde, 1-methyl-
1H-imidazole-2-carbaldehyde, quinoline-2-carbaldehyde, benzal-
dehyde, NaBH(OAc)3 were all obtained from Aldrich. All other re-
agents were purchased from commercial sources and used as
received. Flash column chromatography was done with silica gel 60
(240–400 mesh). Analytical TLC was performed using Merck
glass-backed 0.2 mm silica gel 60 F-254 plates and visualized by ul-
traviolet light, I2, 5% phosphomolybdic acid or 1% ninhydrin in
13C NMR (300 MHz, CDCl3): d = 175.67, 157.13, 155.93, 148.03,
143.75, 143.56, 140.87, 137.04, 127.36, 126.76, 124.92, 123.44,
122.35, 119.63, 66.40, 58.66, 54.34, 53.68, 46.85, 32.21, 25.70,
22.71.
1
EtOH. H and 13C NMR spectra were recorded on a Bruker DPX
300 spectrometer, and all peak positions are relative to TMS. HSQC
spectra were acquired on a Bruker Avance 500 MHz spectrometer,
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M. K. Levadala et al.
PAPER
HRMS: m/z calcd for C33H35N4O4 (M + H+): 551.2652; found:
551.2672.
13C NMR (300 MHz, CDCl3): d = 174.94, 159.28, 148.09, 139.40,
137.45, 129.68, 128.51, 126.46, 123.08, 122.52, 66.45, 56.36,
35.67.
6-[Bis(thiazol-2-yl)methyl]amino-2-(9H-fluoren-9-ylmethoxy-
carbonyl)amino-1-hexanoic Acid (2)
Yield: 84%.
1H NMR (300 MHz, CDCl3): d = 11.45 (br s, 1H, CO2H), 8.20-7.60
(m, 4H), 7.59 (t, J = 7.3 Hz, 2H), 7.35 (t, J = 7.2 Hz, 2H), 7.30-7.10
(m, 4H), 5.86 (d, J = 8.1 Hz, 1H), 4.62-4.32 (m, 3H), 4.19 (t,
J = 6.90 Hz, 1H), 4.17 (s, 4H), 2.60 (m, 2H), 2.00-1.30 (m, 6H,
CH2).
13C NMR (300 MHz, CDCl3): d = 175.04, 171.42, 156.14, 143.96,
143.76, 141.88, 141.23, 127.70, 127.07, 125.18, 119.96,119.82,
66.92, 55.33, 53.82, 47.15, 32.38, 26.73, 22.77.
HRMS: m/z calcd for C29H30N4O4S2Na (M+Na+): 585.1600; found:
585.1653.
HRMS: m/z calcd for C21H22N3O2 (M + H+): 348.1707; found:
348.1721 and calcd for C21H21N3O2 + Na (M + Na+): 370.1525;
found: 370.1501.
2-[Bis(pyridin-2-yl)methyl]amino-1-ethanoic Acid (7)
Yield: 76%.
1H NMR (300 MHz, MeOH-d4): d = 8.29 (d, J = 5.1 Hz, 2 H), 7.60
(t, J = 9.0 Hz, 2 H), 7.30 (d, J = 7.8 Hz, 2 H), 7.12 (t, J = 6.2 Hz, 2
H), 4.10 (s, 4 H), 3.39 (m, 2 H).
13C NMR (300 MHz, MeOH-d4): d = 173.05, 156.10, 149.76,
139.31, 125.15, 124.77, 59.77, 57.77.
5-[Bis(pyridin-2-yl)methyl]amino-1-pentanoic Acid (8)
Yield: 83%.
6-[Bis(1-methyl-1H-imidazol-2-yl)methyl]amino-2-(9H-fluo-
ren-9- ylmethoxycarbonyl)amino-1-hexanoic Acid (3)
Yield: 82%.
1H NMR (300 MHz, CDCl3): d = 7.73 (d, J = 7.20 Hz, 2 H), 7.61 (t,
J = 7.35 Hz, 2 H), 7.36 (t, J = 7.35 Hz, 2 H), 7.27 (t, J = 6.90 Hz, 2
H), 6.95 (s, 2 H), 6.74 (s, 2 H), 5.96 (br s, 1 H), 4.40–4.15 (m, 3 H),
4.14 (t, J = 6.90 Hz, 1 H), 3.73 (q, J = 27.6, 14.1 Hz, 4 H), 3.46 (s,
6 H), 2.54 (m, 2 H), 2.00–1.20 (m, 6 H, CH2).
1H NMR (300 MHz, CDCl3): d = 8.51 (d, J = 4.8 Hz, 2 H), 7.64 (t,
J = 7.5 Hz, 2 H), 7.51 (d, J = 7.8 Hz, 2 H), 7.13 (t, J = 5.1 Hz, 2 H),
3.78 (s, 4 H), 2.54 (t, J = 6.30 Hz, 2 H), 2.26 (t, J = 6.60 Hz, 2 H),
1.59 (m, 4 H).
13C NMR (300 MHz, CDCl3): d = 177.00, 159.45, 148.49, 137.12,
123.41, 122.34, 59.87, 54.12, 34.38, 26.49, 22.91.
6-[Bis(tert-butoxycarbonylmethylamino)]-2-(9H-fluoren-9-yl-
methoxycarbonylamino)hexanoic Acid (9)
Yield: 75%.
1H NMR (300 MHz, MeOH-d4): d = 7.72 (d, J = 7.2 Hz, 2 H), 7.57
(d, J = 7.5 Hz, 2 H), 7.35 (t, J = 7.2 Hz, 2 H), 7.26 (t, J = 7.5 Hz, 2
H), 5.62 (d, J = 8.1 Hz, 1 H), 4.40–4.34 (m, 3 H), 4.18 (t, J = 6.9 Hz,
1 H), 3.39 (s, 4 H), 2.68 (t, J = 7.5 Hz, 2 H), 1.96–1.25 (m, 24 H).
13C NMR (300 MHz, MeOH-d4): d = 174.57, 170.45, 156.08,
144.05, 143.91, 141.29, 127.70, 127.11, 125.25, 119.97, 81.12,
66.92, 55.62, 53.91, 53.75, 47.22, 32.34, 28.19, 27.38, 22.82.
13C NMR (300 MHz, CDCl3): d = 156.16, 145.05, 144.33, 144.13,
141.41, 127.80, 127.23, 125.47, 121.83, 120.07, 66.84, 54.77,
49.38, 47.40, 33.02, 32.31, 25.72, 22.61.
ESI-MS: m/z = 579.2 (M+ + Na), 557.3 (M+ + 1).
6-[Bis(quinolin-2-yl)]methyl]amino-2-(9H-fluoren-9-ylmeth-
oxycarbonyl)amino-1-hexanoic Acid (4)
Yield: 86%.
1H NMR (300 MHz, CDCl3): d = 10.23 (br s, 1 H), 8.07 (d, J = 8.4
Hz, 2 H), 8.01 (d, J = 8.7 Hz, 2 H), 7.75–7.15 (m, 16 H), 6.05 (br s,
1 H), 4.40–4.10 (m, 8 H), 2.81 (m, 2 H), 2.04–138 (m, 6 H, CH2).
13C NMR (300 MHz, CDCl3): d = 175.85, 158.17, 156.19, 146.80,
144.06, 143.91, 141.23, 137.24, 131.59, 129.93,128.30, 127.64,
127.07, 126.63, 125.25, 121.30, 119.52, 66.81, 59.88, 54.67, 47.19,
32.50, 26.05, 22.88.
HRMS: m/z calcd for C33H44N2O8 + Na (M + Na+): 619.2989;
found: 619.2960.
2,6-[Bis(pyridin-2-yl)methyl]diamino-1-hexanoic Acid (10)
Yield: 79%.
1H NMR (300 MHz, CDCl3): d = 8.39 (m, 4 H), 7.72–7.40 (m, 6 H),
7.33 (d, J = 7.8 Hz, 2 H), 7.03 (m, 4 H), 4.0 (q, 4 H), 3.69 (s, 4 H),
3.30 (t, J = 7.2 Hz, 1 H), 2.43 (t, J = 7.2 Hz, 2 H), 1.94–1.16 (m, 6
H).
HRMS: m/z calcd for C41H38N4O4 (M + H+): 651.2965; found:
651.2936.
2-[Bis(pyridin-2-yl)methyl]amino-6-(tert-butoxycarbonyl)ami-
no-1-hexanoic Acid (5)
Yield: 83%.
13C NMR (300 MHz, CDCl3): d = 175.45, 159.87, 159.64, 148.78,
148.24, 137.13, 136.61, 123.00, 122.28, 122.01, 64.21, 60.32,
56.54, 54.38, 29.51, 27.09, 24.66.
HRMS: m/z calcd for C30H35N6O2 (M + H+): 511.2816; found:
511.2828.
1H NMR (300 MHz, CDCl3): d = 8.50 (d, J = 4.2 Hz, 2 H), 7.59 (t,
J = 7.5 Hz, 2 H), 7.30 (d, J = 7.8 Hz, 2 H), 7.15 (t, J = 6.9 Hz, 2 H),
4.80 (br s, 1 H), 4.04 (q, J = 19.2, 14.9 Hz, 4 H), 3.39 (t, J = 7.2 Hz,
1 H), 3.07 (m, 2 H), 2.00–1.40 (m, 15 H).
13C NMR (300 MHz, CDCl3): d = 175.62, 159.27, 156.11, 148.30,
137.31, 123.15, 122.45, 79.00, 64.62, 56.29, 40.30, 29.84, 28.84,
28.50, 24.15.
6-Dibenzylamino-2-(9H-fluoren-9-ylmethoxycarbonyl)amino-
1-hexanoic Acid (11)
Yield: 74%.
1H NMR (300 MHz, CDCl3): d = 9.18 (br s, 1 H), 7.90 (d, J = 7.50
Hz, 2 H), 7.67 (t, J = 7.80 Hz, 2 H), 7.45–7.2 (m, 14 H), 6.24 (d, 1
H), 4.34 (m, 3 H), 4.21 (t, J = 6.20 Hz, 1 H), 4.06 (br s, 4 H), 2.74
(m, 2 H), 2.06–1.20 (m, 6 H).
13C NMR (300 MHz, CDCl3): d = 177.14, 156.31, 144.20, 143.99,
141.27, 131.92, 130.52, 129.94, 129.94, 128.62, 128.21, 127.69,
127.40, 127.11, 125.33, 119.96, 66.80, 56.13, 55.09, 51.47, 47.25,
32.67, 24.12, 22.77.
2-[Bis(pyridin-2-yl)methyl]amino-3-phenyl-1-propionic Acid
(6)
Yield: 78%.
1H NMR (300 MHz, CDCl3): d = 8.50 (d, J = 4.20 Hz, 2 H), 7.56 (t,
J = 7.5 Hz, 2 H), 7.32–7.08 (m, 9 H), 4.10 (dd, J = 15.6 Hz, 4 H),
3.81 (dd, J = 5.6 Hz, 1 H), 3.39 (dd, J = 5.1 Hz, 1 H), 3.08 (dd,
J = 9.9 Hz, 1 H).
ESI-MS: m/z = 449.2 (M+ + Na).
Synthesis 2004, No. 11, 1759–1766 © Thieme Stuttgart · New York

PAPER
Synthesis of Bifunctional Chelates for Nuclear Imaging
1765
Monoalkylation Reactions; 2-[tert-Butoxycarbonyl]amino-6-
[(pyridin-2-yl)methyl]amino-1-hexanoic Acid (13); Typical
Procedure
HRMS: m/z calcd for C22H33N5O4 (M + H+): 432.2605; found:
432.2583.
A solution of Boc-D-lysine (2.0 g, 8.12 mmol) and 2-pyridinecar-
boxaldehyde (0.87 g, 8.12 mmol) in DCE (20 mL) was refluxed for
10 min under argon. The reaction mixture was cooled to 0 °C, and
treated with NaBH(OAc)3 (2.06 g, 9.74 mmol). The suspension was
stirred at r.t. until the completion of reaction. The monopyridyl-
methyl derivative of Boc-D-lysine was obtained (2.73 g, 73%) along
with 10% of bis(pyridyl) derivative as side product following the
same purification procedure as mentioned earlier.
1H NMR (300 MHz, MeOH-d4): d = 8.50 (d, J = 4.2 Hz, 1 H), 7.80
(t, J = 7.8 Hz, 1 H), 7.45 (d, J = 7.8 Hz, 1 H), 7.29 (t, J = 5.1 Hz, 1
H), 3.94 (m, 1 H), 3.88 (s, 2 H), 2.61 (t, J = 7.3 Hz), 1.85–1.3 (m,
15 H).
6-[(tert-Butoxycarbonyl)methyl,(pyridin-2-yl)methyl]amino-2-
[9H-fluoren-9-ylmethoxycarbonyl]amino-1-hexanoic Acid (16)
Yield: 70%.
1H NMR (300 MHz, CDCl3): d = 13.43 (br s, 1 H), 8.57 (d, J = 7.5
Hz, 1 H), 7.74–7.59 (m, 6 H), 7.36 (t, J = 7.5 Hz, 2 H), 7.295 (t,
J = 7.5 Hz, 2 H), 7.20 (t, J = 6.0 Hz, 1 H), 5.88 (d, J = 7.8 Hz, 1 H)
4.35 (m, 3 H), 4.20 (t, J = 6.2 Hz, 1 H), 4.02 (s, 2 H), 3.48 (s, 2 H),
2.72 (br t, 2 H), 2.05–1.49 (m, 15 H).
13C NMR (300 MHz, CDCl3): d = 175.64, 170.15, 157.82, 156.14,
147.60, 144.10, 143.92, 141.28, 138.12, 127.69, 127.09, 125.26,
124.24, 122.90, 120.32, 81.41, 66.86, 58.37, 55.11, 54.33, 53.87,
47.23, 32.50, 27.92, 26.62, 22.77.
HRMS: m/z calcd for C33H40N3O6 (M + H+): 574.2911; found:
574.2935 and calcd for C33H39N3O6 + Na (M + Na+): 596.2731;
found: 596.2739.
13C NMR (300 MHz, MeOH-d4): d = 179.91, 160.09, 157.77,
149.98, 138.79, 124.23, 123.89, 80.02, 57.28, 55.20, 50.10, 34.36,
30.40, 28.93, 24.54.
5-[(Pyridin-2-yl)methyl]amino-1-pentanoic Acid (12)
Yield: 71%.
Acknowledgment
1H NMR (300 MHz, CDCl3): d = 8.40 (d, J = 4.2 Hz, 1 H), 7.54 (t,
J = 7.8 Hz, 1 H), 7.15 (d, J = 8.1 Hz, 1 H), 7.04 (t, J = 4.8 Hz, 1 H),
4.59 (s, 2 H), 3.21 (t, J = 5.7 Hz, 1 H), 2.32 (t, J = 5.4 Hz, 1 H), 1.69
(m, 4 H).
This work was supported by a grant from the National Institutes of
Health, National Institutes of Allergy and Infection Diseases, STTR
1R41 A1044080-01.
13C NMR (300 MHz, CDCl3): d = 170.14, 157.04, 148.82, 137.01,
122.33, 122.06, 52.03, 48.14, 32.18, 23.05, 21.18.
References
(1) Fischman, A. J.; Babich, J. W.; Strauss, H. W. J. Nucl. Med.
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Mixed-Alkylation Reactions; 2-[tert-Butoxycarbonyl]amino-6-
[(pyridin-2-yl)methyl,(thiazol-2-yl)methyl]amino-1-hexanoic
Acid (14); Typical Reaction
(2) (a) Yeh, S. M.; Sherman, D. G.; Meares, C. F. Anal.
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(4) Rao, T. N.; Adhikesavalu, D.; Camerman, A.; Fritzberg, A.
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Stevenson, D. A.; Kushner, S. A.; McElgin, W. T.; Mozley,
P. D.; Kung, H. F. J. Med. Chem. 1997, 40, 9; and references
cited therein.
A solution of Boc-D-lysine (2.0 g, 8.12 mmol) and 2-pyridinecar-
boxaldehyde (0.87 g, 8.12 mmol) in DCE (10 mL) was refluxed for
10 min under argon. The reaction mixture was cooled to 0 °C, and
treated sequentially with NaBH(OAc)3 (4.30 g, 20.3 mmol) and 2-
thiazolecarboxyaldehyde(0.91 g, 8.12 mmol). The reaction mixture
was stirred at r.t. until the completion of reaction and purified as
mentioned above to obtain the mixed derivative of Boc-D-lysine
(3.39 g, 75%) along with 12% of the bis-alkyl derivative as a side
product.
1H NMR (300 MHz, CDCl3): d = 8.55 (d, J = 5.1 Hz, 1 H), 7.75 (t,
J = 7.5 Hz, 1 H), 7.68 (d, J = 3.3 Hz, 1 H), 7.64 (d, J = 7.8 Hz, 1 H),
7.25 (d, J = 3.3 Hz, 1 H), 7.22 (t, J = 6.3 Hz, 1 H), 5.3 (d, J = 8.7
Hz, 1 H), 4.28 (m, 1 H), 3.99 (s, 2 H), 3.87 (s, 2 H) 2.56 (t, J = 7.2
Hz, 2 H), 1.92–1.3 (m, 15 H).
13C NMR (300 MHz, CDCl3): d = 175.46, 171.67, 158.54, 155.68,
147.97, 142.23, 137.90, 123.59, 122.81, 119.63, 79.57, 59.29,
55.36, 54.20, 53.63, 32.81, 28.51, 26.73, 22.98.
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L. S.; Webb, E. G.; Marmon, M. E.; Vanderheyden, J. L.;
Srinivasan, A. Bioconjugate Chem. 1997, 8, 407. (b)Wong,
E.; Fauconnier, T.; Bennett, S.; Valliant, J.; Nguyen, T.; Lau,
F.; Lu, L. F. L.; Pollak, A.; Bell, R. A.; Thornback, J. R.
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(8) Numerous examples of peptide-based bifunctional chelators
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Representative examples include: (a) MAG3 types
(MAG3 = mercaptoacetylglycylglycylglycine): Liu, S.;
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Srinivasan, A.; Rao, T. N.; Hobson, L. J.; Reno, J. M.;
HRMS: m/z calcd for C21H31N4O4S (M + H+): 435.2061; found:
435.2025 and calcd for C21H30N4O4S + Na (M + Na+): 457.1880;
found: 457.1859.
2-[tert-Butoxycarbonyl]amino-6-[(1-methyl-1H-imidazol-2-
yl)methyl,(pyridin-2-yl)methyl]amino-1-hexanoic Acid (15)
Yield: 73%.
1H NMR (300 MHz, CDCl3): d = 10.19 (br s, 1 H), 8.52 (d, J = 4.2
Hz, 1 H), 7.62 (t, J = 7.5 Hz, 1 H), 7.31 (d, J = 7.8 Hz, 1 H), 7.14 (t,
J = 6.2 Hz, 1 H), 6.94 (s, 1 H), 6.73 (s, 1 H), 5.42 (br s, 1 H), 4.17
(m, 1 H), 3.38–3.64 (m, 4 H), 3.55 (s, 3 H), 2.48 (m, 2 H), 1.95–1.20
(m, 15 H).
13C NMR (300 MHz, CDCl3): d = 176.41, 158.70, 155.67, 148.90,
145.07, 136.95, 124.84, 123.81, 122.42, 121.76, 79.13, 60.19,
54.44, 54.24, 49.68, 33.33, 32.78, 28.56, 25.81, 22.84.
Synthesis 2004, No. 11, 1759–1766 © Thieme Stuttgart · New York

1766
M. K. Levadala et al.
PAPER
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(24) Table 1, Entry 6 (C22 H25 N3 O3, formula weight 379.45):
Orthorhombic, space group P212121 with a = 7.3392(4) Å,
b = 15.0539(7) Å, c = 17.9286(9) Å, V = 1980.82(17) Å3,
Z = 4, R1 = 0.0556, wR2 = 0.1079 (F2, all data). Full details
on the crystal structure of 6 are available from the authors.
(25) The imine to aldehyde ratio was in the range of 1: 0.10–0.20,
as determined by 1H NMR spectrum.
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Gramlich, V.; Schubiger, P. A. J. Chem. Soc., Dalton Trans.
1994, 2815.
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1999, 121, 6076.
(28) Maresca, K. P.; Levadala, M. K.; Banerjee, S. R.; Babich, J.
W.; Zubieta, J., unpublished results.
Synthesis 2004, No. 11, 1759–1766 © Thieme Stuttgart · New York