Efficient regioselective ring opening of activated aziridine-2-carboxylates with [18F]fluoride

Article abstract of DOI:10.1002/open.201402081

Aziridines can undergo a range of ring-opening reactions with nucleophiles. The regio- and stereochemistry of the products depend on the substituents on the aziridine. Aziridine ring-opening reactions have rarely been used in radiosynthesis. Herein we rep

Full text of DOI:10.1002/open.201402081

DOI: 10.1002/open.201402081
Efficient Regioselective Ring Opening of Activated
Aziridine-2-Carboxylates with [18F]Fluoride
Christina Schjoeth-Eskesen,[a, b] Paul Robert Hansen,[c] Andreas Kjaer,[a, b] and Nic Gillings*[a]
Aziridines can undergo a range of ring-opening reactions with
nucleophiles. The regio- and stereochemistry of the products
depend on the substituents on the aziridine. Aziridine ring-
opening reactions have rarely been used in radiosynthesis.
Herein we report the ring opening of activated aziridine-2-car-
boxylates with [18F]fluoride. The aziridine was activated for nu-
cleophilic attack by substitution of various groups on the aziri-
dine nitrogen atom. Fluorine-18 radiolabelling was followed by
ester hydrolysis and removal of the activation group. Totally re-
gioselective ring opening and subsequent deprotection was
achieved with tert-butyloxycarbonyl- and carboxybenzyl-acti-
vated aziridines to give a-[18F]fluoro-b-alanine in good radio-
chemical yield.
Introduction
Nucleophilic ring opening of aziridines is an efficient way of
synthesising a variety of organic compounds.[1,2] The strain in
the three-membered ring of aziridine allows it to undergo
a range of ring-opening reactions with nucleophiles.[3] The
presence of specific substituents on the aziridine can control
the regio- and stereochemistry of the ring-opening reaction.[4]
For non-activated aziridines a strong nucleophile or acid cata-
lyst is required to open the ring. The reactivity of the aziridine
ring toward nucleophiles is increased by having an activation
group on the aziridine ring nitrogen atom to stabilise the neg-
ative charge and to make the nitrogen a better leaving
group.[5,6] The nucleophile can attack either of the two carbon
atoms in the aziridine ring. For 2-substituted aziridines ring-
opening can lead to two regioisomeric amine products. For
cases in which an acyl substituent is used, attack is favoured
on the more hindered aziridine ring carbon atom with azide,
halide, or cyanide as the nucleophile, but is favoured on the
less hindered carbon with alcohols as the nucleophile.[7]
Nucleophilic attack of aziridines by fluoride ions has been
studied much less than other halides.[8] The synthesis of b-fluo-
roamines has been explored by ring opening of aziridines. Flu-
orination reagents such as HF, HF/pyridine, and diethylamino-
sulfur trifluoride (DAST) have been used.[9,10,11] Ring opening
was recently carried out with the use of KF·2H2O/Bu4NHSO4,
KF·2H2O, or KHF2 in an ionic liquid and triethylamine trihydro-
fluoride.[12,13,14] However, these methods are not convenient
[15]
1
when using the radionuclide fluorine-18 (t = =109.7 min).
2
Fluorine-18 is one of the most frequently used radionuclides
for positron emission tomography (PET), which is an important
technique for imaging and studying biochemical processes in
vivo. PET uses organic compounds labelled with short-lived
positron-emitting radionuclides and relies on rapid synthetic
pathways that can incorporate radionuclides into these com-
pounds.[16–18] Only a few publications can be found on the use
of aziridines for radiosynthesis.[8,15,19–24]
The aim of this study was to investigate ring-opening reac-
tions of aziridine-2-carboxylates with [18F]fluoride in aziridines
with various nitrogen-activating groups. Regioselectivity of
ring opening was studied, and reaction conditions and depro-
tection protocols were optimised. Using aziridine-2-carboxy-
lates, fluorine-18 ring opening followed by de-esterification
and removal of the activating group will yield two possible iso-
mers: a-[18F]fluoro-b-alanine or b-[18F]fluoroalanine (Scheme 1).
[a] C. Schjoeth-Eskesen, Prof. Dr. A. Kjaer, Dr. N. Gillings
Department of Clinical Physiology, Nuclear Medicine & PET
Copenhagen University Hospital Rigshospitalet
Blegdamsvej 9, 2100 Copenhagen Ø (Denmark)
E-mail: nic@pet.rh.dk
[b] C. Schjoeth-Eskesen, Prof. Dr. A. Kjaer
Cluster for Molecular Imaging, Faculty of Health Sciences
University of Copenhagen
Results and Discussion
Blegdamsvej 3B, 2100 Copenhagen Ø (Denmark)
[c] Dr. P. R. Hansen
The ring-opening reaction with fluorine-18 was investigated by
using methylaziridine-2-carboxylates activated with five differ-
ent groups. The initial approach to synthesise these com-
pounds was from commercially available triphenylmethyl
(trityl)-protected methylaziridine-2-carboxylate. Removal of the
trityl group proved to be problematic, and it was only possible
to synthesise the p-nitrobenzenesulfonyl (nosyl)- and toluene-
sulfonyl (tosyl)-activated aziridine-2-carboxylates with this ap-
Department of Drug Design & Pharmacology
University of Copenhagen
Universitetsparken 2, 2100 Copenhagen Ø (Denmark)
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This is an open access article under the terms of the Creative Commons
Attribution-NonCommercial-NoDerivs License, which permits use and
distribution in any medium, provided the original work is properly cited,
the use is non-commercial and no modifications or adaptations are
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dissolved in either N,N-dimethyl-
formamide (DMF) or dimethyl
sulfoxide (DMSO) and reacted
with each of the five aziridine-2-
carboxylates. The ring-opening
reaction was investigated at vari-
ous temperatures using conven-
tional or microwave heating.
Scheme 1. Aziridine ring opening with [18F]fluoride followed by removal of activating group G and methyl ester
hydrolysis to afford a-[18F]fluoro-b-alanine (1) or b-[18F]fluoroalanine (2).
proach. The remaining aziridines with tert-butyloxycarbonyl
(Boc), carboxybenzyl (Cbz), and fluorenylmethyloxycarbonyl
(Fmoc) activating groups were synthesised directly from meth-
ylaziridine-2-carboxylate (Scheme 2). Because previous studies
demonstrated the lack of reactivity of non-activated aziridines
toward nucleophilic ring opening in the absence of a Lewis
acid,[24] for example, the non-activated aziridine-2-carboxylate
was not tested.
Initial radiolabelling experiments were performed using the
Boc-activated aziridine, and the results are summarised in
Table 1. Labelling with [18F]fluoride using K222/K2CO3 as base
gave lower yields than with TEAHCO3. Microwave heating led
to increased yields with decreased reaction times. Longer reac-
tion times by microwave heating did not lead to higher yields
owing to decomposition of the aziridine precursor during pro-
longed exposure to high temperature and base.
Seo et al. demonstrated that
the milder base forms of Chro-
mafix columns could be benefi-
cial for radiolabelling yields by
preserving the integrity of the
radiolabelling precursor.[25] As
can be seen in Table 1, the phos-
phate form gave much better re-
sults than the bicarbonate form
(entries 10 and 11). Due to the
simplicity of this approach, this
may be advantageous, although
Scheme 2. Synthesis of activated methylaziridine-2-carboxylates. Reagents and conditions: a) TFA, CHCl3, MeOH,
08C, 2.5 h; b) p-TsCl, EtOAc, H2O, RT, 48 h; c) NsCl, EtOAc, H2O, RT, 18 h; d) Boc2O, Et3N, MeCN, RT, 18 h; e) CbzCl,
Et3N, MeCN, RT, 18 h; f) Fmoc-OSu, Et3N, THF, H2O, RT, 2 h.
Labelling conditions for aziridine ring-opening with fluo-
rine-18
Table 1. [18F]Fluoride labelling of Boc-activated methylaziridine-2-carbox-
ylate.
The first step in aziridine ring opening was to find the optimal
labelling conditions. [18F]Fluoride was prepared by proton irra-
diation of enriched oxygen-18 water and thus initially available
as an aqueous solution. To provide reactive [18F]fluoride, it was
firstly trapped on a small purification cartridge to remove (and
recover) most of the water, and then eluted from the cartridge
using a phase-transfer catalyst. Two different approaches were
investigated for this procedure. After trapping of aqueous
[18F]fluoride on a quaternary methylammonium (QMA) column
(anion exchange, silica based), it was eluted with either a Kryp-
tofix solution containing potassium carbonate (K222/K2CO3) or
a tetraethylammonium bicarbonate solution (TEAHCO3), and
subsequently dried by azeotropic distillation with acetonitrile.
Recently, a faster and easier method for preparation of anhy-
drous [18F]fluoride using Chromafix polymer cartridges (anion
exchange) was published.[25] In this second approach
[18F]fluoride can be quantitatively trapped on the resin and,
following rinsing with methanol to remove water, eluted with
an anhydrous solution of tetrabutylammonium mesylate
(TBAOMs) in methanol and dried without the need for numer-
ous azeotropic drying cycles. The Chromafix columns are avail-
able in the bicarbonate (PS-HCO3) form, but can be easily con-
verted into less basic forms by flushing with a potassium phos-
phate solution (PS-K2PO4) prior to [18F]fluoride trapping, for
example. For all methods, after drying the [18F]fluoride, it was
Entry
Base[a]
Solvent
T [8C][b]
t [min]
Yield [%][c]
1
2
3
4
5
6
7
8
K222/K2CO3
K222/K2CO3
K222/K2CO3
TEAHCO3
TEAHCO3
TEAHCO3
TEAHCO3
TEAHCO3
TEAHCO3
DMSO
DMSO
DMSO
DMSO
DMSO
DMSO
DMF
DMSO
DMSO
DMSO
DMSO
100
130
130 MW
100
130
130 MW
130 MW
150
150 MW
130
20
20
10
20
20
10
10
10
10
20
20
5
17
15
15
25
45
36
31
43
3
[d]
9
10
11
À
TBAOMs/HCO3
TBAOMs/PO4
3À
130
40
[a] Base concentrations: K2CO3 8 mm, TEAHCO3 19 mm, TBAOMs 8 mm.
[b] MW=microwave. [c] Ring-opening yields are based on percentage ra-
diochemical conversion determined by radio-HPLC. [d] Concentration of
K2CO3 solution: 19 mm.
ring-opening yields were not quite as high as with the QMA/
TEAHCO3 approach. Overall, the best yields for labelling of the
Boc-activated aziridine were obtained with TEAHCO3 as the
base and microwave heating at 1308C for 10 min.
The solvent was changed to DMF to see if this could im-
prove the yield. The reaction with TEAHCO3 and DMF was
tested at various temperatures and also by microwave heating.
All reactions in DMF (Boc, Cbz, tosyl) gave consistently lower
yields than obtained for similar reactions in DMSO (for exam-
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ChemistryOpen 2015, 4, 65 – 71 66

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ple, see entry 7 in Table 1). DMSO was thus chosen as solvent
for all subsequent labelling reactions.
Table 3. [18F]Fluoride labelling of tosyl-activated methylaziridine-2-car-
boxylate.
The ring-opening reaction of the Cbz-activated aziridine
with [18F]fluoride failed completely with K222/K2CO3. Ring
opening was observed if the base was changed to TEAHCO3,
and microwave heating further increased the yields. Chromafix
columns were tested with the microwave conditions, but the
results were not promising. The best labelling conditions for
the Cbz-activated aziridine were obtained using TEAHCO3 with
microwave heating at 1308C for 10 min (Table 2).
Entry
Base[a]
Solvent
T [8C][b]
t [min]
Yield [%][c]
1
2
3
4
5
7
8
K222/K2CO3
K222/K2CO3
TEAHCO3
TEAHCO3
TEAHCO3
DMSO
DMSO
DMSO
DMSO
DMSO
DMSO
DMSO
100
130
90
90 MW
100 MW
90
20
20
20
20
10
20
10
10
4
26
34
72
3
À
TBAOMs/HCO3
TBAOMs/PO4
3À
100 MW
1
[a] Base concentrations: K2CO3 8 mm, TEAHCO3 19 mm, TBAOMs 8 mm.
[b] MW=microwave. [c] Ring-opening yields are based on percentage ra-
diochemical conversion determined by radio-HPLC.
Table 2. [18F]Fluoride labelling of Cbz-activated methylaziridine-2-carbox-
ylate.
Entry
Base[a]
Solvent
T [8C][b]
t [min]
Yield [%][c]
Concentration of activated aziridine for labelling
1
2
3
4
5
6
7
8
K222/K2CO3
K222/K2CO3
TEAHCO3
TEAHCO3
TEAHCO3
TEAHCO3
TBAOMs/HCO3
TBAOMs/PO4
DMSO
DMSO
DMSO
DMSO
DMSO
DMSO
DMSO
DMSO
100
130
100
130
130 MW
150 MW
130 MW
130 MW
20
20
20
20
10
10
10
10
0
0
Different concentrations of the activated aziridine precursor for
labelling were tested to determine the effect on ring-opening
yield. For these experiments the Boc-activated aziridine-2-car-
boxylate was used, and labelling with [18F]fluoride was per-
formed with TEAHCO3 and microwave conditions at 1308C for
10 min. The concentration of TEAHCO3 and the volume were
kept constant. The correlation between aziridine precursor
concentration and ring-opening yield determined by radio-
HPLC is shown in Figure 1. At precursor concentrations of 10
and 20 mgmLÀ1 the radiochemical yield was ~60%. Increasing
precursor concentrations only marginally increased the radio-
chemical yield, and thus a concentration of 10-20 mgmLÀ1 was
considered to be adequate.
23
29
48
26
4
À
3À
8
[a] Base concentrations: K2CO3 8 mm, TEAHCO3 19 mm, TBAOMs 8 mm.
[b] MW=microwave. [c] Ring-opening yields are based on percentage ra-
diochemical conversion determined by radio-HPLC.
Initial radiolabelling attempts using the Fmoc-activated aziri-
dine failed. The labelling reactions were tested at 100 and
1308C with K222/K2CO3 and TEAHCO3. HPLC analysis showed
decomposition of the aziridine precursor under these basic re-
action conditions, presumably due to cleavage of the Fmoc
group with the resultant decreased reactivity of the aziridine.
The milder base TBAOMs in combination with the Chromafix
columns (both PS-HCO3 and PS-K2PO4) was tested, but again
did not lead to any labelling. Fmoc was thus abandoned as an
aziridine activating group.
[18F]Fluoride labelling experiments with the nosyl-activated
aziridine were first performed at 100 and 1308C with K222/
K2CO3 and TEAHCO3. No labelling was observed after 20 min
for both conditions. The UV trace on the HPLC indicated de-
composition of the aziridine precursor. The stability of the
nosyl-activated aziridine was thus investigated by heating in
DMSO and showed that the optimal temperature was between
50 and 608C. Radiolabelling was not possible at such low tem-
peratures, and the nosyl group was thus abandoned as a suita-
ble aziridine activating group.
Figure 1. Correlation between Boc-activated methylaziridine-2-carboxylate
concentration and radiochemical yield in ring opening with [18F]fluoride (re-
action conditions: TEAHCO3, DMSO, 1308C MW, 10 min).
The best conditions for [18F]fluoride labelling of the tosyl-ac-
tivated aziridine involved the use of TEAHCO3 and microwave
heating at 1008C (Table 3). These conditions led to a high radi-
ochemical yield of 72%. The tosyl-activated aziridine was not
stable at temperatures above 1008C, and higher temperatures
were detrimental to the radiochemical yield. As with the Boc-
and Cbz-activated aziridines, labelling with K222/K2CO3 and the
Chromafix method gave low yields.
Base concentration
Various concentrations of TEAHCO3 were tested for labelling
the Boc-activated aziridine-2-carboxylate using standard condi-
tions. The volume and precursor concentration were kept con-
stant. The correlation between base concentration in the reac-
tion and ring-opening yield is illustrated in Figure 2. The high-
est radiochemical yield was achieved at a base concentration
of 7.5 mm; higher base concentrations appeared to be slightly
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product. Removal of the tosyl activating group was attempted
by using hydrochloric acid and sulfuric acid.[26] Treatment with
H2SO4 converted the two products into one radioactive prod-
uct. The product was analysed by a second HPLC method to
determine the regioselectivity, but it did not correspond to any
of the two reference compounds. Due to the small masses in-
volved in working with high specific radioactive fluorine-18, it
was not possible to determine the identity of this product. Fur-
ther work using the tosyl-activated aziridine was not pursued.
The Boc group on the 18F-ring-opened products could easily
be removed by heating in aqueous HCl, and these conditions
also hydrolysed the ester in the same step. The Cbz group was
removed by adaptation of a published method using palladi-
um acetate, triethylsilane, and triethylamine.[27] The small
amount of triethylamine in the deprotection step was found to
be sufficient to hydrolyse the methyl ester as well. This depro-
tection method was found to be far superior to hydrogenoly-
sis, using hydrogen gas together with palladium over charcoal,
and allowed Cbz cleavage and de-esterification in one step
within 10 min, compared with 40 min for the reported hydro-
genolysis approach.[8,15]
Figure 2. Correlation between TEAHCO3 concentration and radiochemical
yields in the ring opening of Boc-activated methylaziridine-2-carboxylate
with [18F]fluoride (reaction conditions: 5 mg precursor, DMSO, 1308C MW,
10 min).
detrimental. The radiochemical yield was slightly higher than
observed previously with an equivalent base concentration
(Table 1, entry 6). This is explained by the use of a freshly pre-
pared TEAHCO3 solution.
The ring-opening product for labelling of the Boc- and Cbz-
activated aziridines after ester hydrolysis and activation group
removal was determined to be a-[18F]fluoro-b-alanine
(Figure 3). [18F]Fluoride exclusively attacked the aziridine at the
most substituted a-carbon, and none of the other regioisomer
was detected. Interestingly, reports on ring opening of non-ac-
tivated isopropylaziridine-2-carboxylate with HF/pyridine
showed the complete opposite regioselectivity, with attack at
the unsubstituted b-carbon atom.[28,29] Ring opening of a 2-car-
boxylic ester substituted aziridinium salt by reaction with tetra-
n-butylammonium fluoride demonstrated exclusive attack at
the a-carbon atom, in agreement with our results, whilst the
2-methyl ether analogue gave only 25% attack at the a-
carbon atom.[30] In contrast, a recent report using triethylamine
Regioselectivity
Nucleophilic ring opening of aziridines can lead to two prod-
ucts due to reaction at both of the aziridine carbon atoms. For
aziridine-2-carboxylates treated with [18F]fluoride, both a-
[18F]fluoro-b-alanine and b-[18F]fluoroalanine may be produced
following ester hydrolysis and removal of N-activating groups.
We succeeded in developing an HPLC analysis whereby these
two compounds could be well separated (retention times of
3.3 and 5.8 min, Rs=6), thus allowing for identification of the
[18F]-labelled products by co-injection of authentic reference
compounds.
Ring opening with [18F]fluoride was only successful for aziri-
dine-2-carboxylates with Boc, Cbz, or tosyl as the activating
group. The yields of the ring-opening reaction were similar for
Boc and Cbz and highest for the more activating tosyl group.
With all three aziridines, two initial radiolabelled products were
observed on radio-HPLC analysis. The first eluting peak was
identified as the N-activated ring-opening product after hydrol-
ysis of the ester function. Surprisingly, the base in the labelling
reaction and the high temperatures were able to hydrolyse the
methyl ester. The later eluting peak was the intact N-activated
ring-opening product. Unsurprisingly, the extent of ester hy-
drolysis during the labelling reaction was found to depend on
the base concentration. No hydrolysis was observed at
TEAHCO3 concentrations below 0.2 mm, and only 4% ring-
opening product was seen at this base concentration. To avoid
ester hydrolysis, a more stable ester could be used. Previously
we had prepared the N-Boc-activated aziridine with an isopro-
pyl ester instead of the methyl, and reaction with [18F]fluoride
afforded intact ring-opening product (unpublished data). The
yield of intact N-activated ring-opening product determined
by HPLC was confirmed by TLC. Under the TLC conditions
used, there was inadequate separation of the hydrolysed prod-
uct from [18F]fluoride to enable precise quantification of this
Figure 3. Analytical HPLC trace of radiolabelling product after deprotection
spiked with reference a-fluoro-b-alanine. The radioactivity trace (top) is shift-
ed to compensate for the time delay between the UV and radiodetectors.
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trihydrofluoride for ring opening of an N-deactivated ethylaziri-
dine-2-carboxylate showed only 34% of the a-fluoro product,
with the remainder due to attack at the b-carbon.[14] Van Oos-
ten et al. reported a fluorine-18 attack on 2-methylaziridine
with Cbz and benzoyl (Bz) as activating group on the nitrogen.
Two regioisomers were formed for both aziridine ring opening
reactions, but with opposite regioselectivity. The Cbz 2-methyl-
aziridine was more reactive at the less hindered 28 carbon,
whilst the Bz 2-methylaziridine showed opposite regioselectivi-
ty.[8,15] The authors also reported very different regioselectivity
for both aziridines on fluorine-19 ring opening with HF. In this
case, only attack at the methyl-substituted 38 carbon was ob-
served. Roehn et al. tested aziridine ring opening with
[18F]fluoride on a model compound, a 2-carboxylic acid benzyl
amide substituted aziridine. They tested Bz and three different
sulfonyl activating groups. The Bz aziridine was unsuccessful,
but the ring-opening yields were high with sulfonyl activating
groups. The best activating group was used on three biomole-
cules: two peptides and a thymidine derivative. The ring open-
ing with [18F]fluoride for the peptide-based aziridines gave low
yields, but the yield for the thymidine was 87%. The only prod-
uct shown for the ring opening was with [18F]fluoride attack at
the less hindered carbon, but regioselectivity of ring opening
was not discussed in the report. Furthermore, no attempts
were made to remove the aziridine activating group subse-
quent to radiolabelling.[21] Recently an ApoSense compound,
[18F]2-(5-(dimethylamino)naphthalene-1-sulfonamido)-2-(fluoro-
methyl)butanoic acid ([18F]NST732), was successfully prepared
by nucleophilic ring opening of a sulfonyl-activated aziridine
precursor.[22] In this case, there were two regioisomeric prod-
ucts formed at an approximate 60:40 ratio, with attack on the
most substituted carbon atom slightly favoured. There are two
earlier reports on fluorine-18 ring opening of aziridines, but in
both cases radiochemical yields were low.[19,20]
tivity depends more on the nitrogen-activating groups as pre-
viously noted.[34] The relatively small ionic radius of fluoride is
assumed to minimise the influence of steric effects on regiose-
lectivity. Thus, we can rationalise the observed regioselectivity
of ring opening of activated aziridine-2-carboxylates with
[18F]fluoride and conclude that this is directed mainly by elec-
tronic effects due to the carboxylic ester substituent.
Conclusions
[18F]Fluoride ring-opening reactions of five N-activated methyl-
aziridine-2-carboxylates were studied. Fmoc activation was not
suitable due to its instability under basic conditions. The nosyl
group was only stable at temperatures up to 608C, and no
ring opening occurred at this temperature. The tosyl-activated
aziridine-2-carboxylate was somewhat unstable at high tem-
peratures; nonetheless, good fluorine-18 incorporation was ob-
served with microwave heating at 1008C. Unfortunately, fol-
lowing deprotection no identifiable radioactive products were
obtained. Both Boc- and Cbz-activated aziridine-2-carboxylates
underwent successful regioselective ring opening with
[18F]fluoride in high radiochemical yield to produce a-
[18F]fluoro-b-alanine after ester hydrolysis and removal of the
activating group. Nucleophilic attack by [18F]fluoride occurred
exclusively at the most substituted carbon atom.
a-Fluoro-b-alanine has long been known as a major catabo-
lite of the chemotherapeutic drug 5-fluorouracil (5-FU), and is
known to exhibit neurotoxicity in patients treated with 5-FU if
its formation is not suppressed.[35] The radiosynthesis of a-
[18F]fluoro-b-alanine via a bromine-to-fluorine exchange reac-
tion has been reported in two conference abstracts,[36,37] but
no biological studies were reported. This radiopharmaceutical
may be useful for non-invasively studying the fate of 5-FU in
humans. Furthermore, fluorine-18 ring opening of aziridines
may be extended to the preparation of [18F]-labelled peptides.
Whilst there are numerous examples of ring opening of aziri-
dines with fluoride, these predominantly use acidic conditions
(e.g., HF/pyridine) or a Lewis acid (e.g., BF3), often with non-ac-
tivated aziridines, in which the reaction appears to be facilitat-
ed by initial protonation of the nitrogen to form an aziridinium
ion.[7] Nucleophilic ring opening of activated aziridines is gen-
erally considered to occur via an SN2 mechanism,[31] and there
are numerous reports that support this.[32] Ring opening of
a chiral aziridine-2-carboxylate with [11C]cyanide was previously
shown to lead to racemic products,[24] and preliminary results
of ring opening of the same aziridine with [18F]fluoride ap-
peared to show the same racemisation (unpublished data).
This lack of retention of stereochemistry can be rationalised by
a base-catalysed racemisation of the initially formed ring-
opened product under high temperature and basic reaction
conditions.[24,33] For this reason, the current work was not fo-
cussed on stereoselectivity. The observed exclusive nucleophil-
ic attack at the most substituted carbon atom in our experi-
ments can be rationalised by the strong electron-withdrawing
nature of the carboxylate group. For previously reported reac-
tions of [18F]fluoride with 2-methylaziridines,[8,15] there is no
electron-withdrawing substituent on the aziridine ring; there-
fore a mixture of regioisomers is produced, and the regioselec-
Experimental Section
General
Methylaziridine-2-carboxylate was purchased from FluoroChem
and H-b-fluoro-d,l-Ala-OH from Bachem. Other chemicals and sol-
vents were purchased from Sigma–Aldrich. Silica gel 60 (40–63 mm)
was purchased from Merck. QMA cartridges were purchased from
Merck Millipore, and Chromafix PS-HCO3 cartridges were purchased
from ABX (Radeberg, Germany). H and 13C NMR spectra were ac-
1
quired with a 400 MHz Bruker Avance II instrument equipped with
TopSpin software. HRMS was performed using an Agilent 1200
HPLC system coupled via a T-splitter with a Bruker microTOF-Q II
mass spectrometer with an electrospray ionisation interface. Mass
spectra were acquired in positive mode. [18F]Fluoride was pro-
duced by the 18O(p,n)18F reaction with proton irradiation of 18O-en-
riched water (Taiyo Nippon Sanso Corp.) using an EclipseTM HP Cy-
clotron (Siemens Healthcare). An automated Scansys module was
used for drying the [18F]fluoride and microwave reactions. HPLC
analysis was performed on a Gilson HPLC with a Dionex UV lamp
(UVD170U) and a Scansys radiodetector. Aziridine radiolabelling
products were analysed on a Phenomenex Kinetex column (2.6 mm
C18, 50ꢁ4.6 mm) at a flow rate of 1.5 mLminÀ1. The mobile phase
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was 0.1% TFA in MeCN/H2O 15:85 for compounds 3 and 5, 20:80
for compounds 4 and 6, and 40:60 for compound 7 (HPLC meth-
od A). The reference compounds H-b-fluoro-d,l-Ala-OH and a-
fluoro-b-alanine were analysed on a Phenomenex Luna column
(5 mm C18 100 ꢂ, 150ꢁ4.6 mm) at a flow rate of 1 mLminÀ1, and
mobile phase consisting of 5 mm sodium decanesulfonate and
25 mm potassium dihydrogen phosphate, pH 2.6 (HPLC method B).
MgSO4 and concentrated. The compound was purified by flash
chromatography in 10 to 20% EtOAc in heptane to give a colour-
less oil (321 mg, 81%). Rf =0.32 (heptane/EtOAc 3:1); HPLC tR:
1
8.7 min (method A); H NMR (400 MHz, CDCl3): d=3.79 (s, 3H), 3.05
(dd, 1H, J=3.2, 5.4 Hz), 2.54 (dd, 1H, J=1.4, 3.2 Hz), 2.43 (dd, 1H,
J=1.4, 5.4 Hz), 1.47 ppm (s, 9H); 13C NMR (100 MHz, CDCl3): d=
168.8, 159.5, 82.0, 52.5, 34.8, 31.2, 27.8 ppm; HRMS (ESI): m/z calcd
for C9H15NO4 +Na+: 224.0899 [M+Na+]; found 224.0893.
1-Benzyl 2-methylaziridine-1,2-dicarboxylate (6): Benzyl chloro-
formate (778 mg, 4.56 mmol) was added to a solution of methyla-
ziridine-2-carboxylate (419 mg, 4.14 mmol) in MeCN (10 mL) and
Et3N (0.6 mL). The solution was stirred at room temperature over-
night and concentrated. The residue was dissolved in CH2Cl2,
washed with NaHCO3, dried over MgSO4 and concentrated. The
compound was purified by flash chromatography in 10 to 20%
EtOAc in heptane to give a colourless oil (639 mg, 66%). Rf =0.19
(heptane/EtOAc 3:1); HPLC tR: 12.7 min (method A); 1H NMR
(400 MHz, CDCl3): d=7.32–7.42 (m, 5H), 5.17 (s, 2H), 3.74 (s, 3H),
3.13 (dd, 1H, J=3.2, 5.4 Hz), 2.63 (dd, 1H, J=1.3, 3.2 Hz), 2.51 ppm
(dd, 1H, J=1.3, 5.4 Hz); 13C NMR (100 MHz, CDCl3): d=168.6, 160.7,
141.1, 128.4, 68.6, 52.6, 34.8, 31.3 ppm; HRMS (ESI): m/z calcd for
C12H13NO4 +H+: 236.0923 [M+H+]; found 236.0912.
Synthesis
Methyl 1-tosylaziridine-2-carboxylate (3)[38]: Trifluoroacetic acid
(TFA; 1.8 mL, 23.5 mmol) was added dropwise to a solution of
methyl (S)-(À)-1-tritylaziridine-2-carboxylate (501 mg, 1.46 mmol) in
CHCl3 and MeOH (2.6 mL, 50:50) at 08C. The solution was stirred at
08C for 2.5 h. The solvents were removed, and the remaining solid
was dried with Et2O (5ꢁ4 mL). The residue was dissolved in Et2O
(30 mL) and H2O (30 mL). The ether layer was extracted with H2O
(20 mL). The combined aqueous fractions were neutralised with
solid NaHCO3 and diluted with EtOAc (30 mL). The mixture was
cooled to 08C and p-toluenesulfonyl chloride (278 mg, 1.46 mmol)
was added. The solution was stirred at room temperature for 48 h.
The organic and aqueous phases were separated, and the aqueous
phase was extracted with EtOAc (40 mL). The organic phases were
combined and concentrated. The compound was purified by flash
chromatography in 25% EtOAc in heptane to give a colourless oil
(253 mg, 68%). Rf =0.61 (heptane/EtOAc 1:1); HPLC tR: 7.4 min
1-((9H-Fluoren-9-yl)methyl) 2-methylaziridine-1,2-dicarboxylate
(7): 9-Fluorenylmethyl N-succinimidylcarbonate (1.51 g, 4.94 mmol)
was added to a solution of methylaziridine-2-carboxylate (454 mg,
4.49 mmol) in THF (6 mL), H2O (2 mL), and Et3N. The solution was
stirred at room temperature for 2 h at pH 8–9. The mixture was
concentrated, and the residue was diluted with 10% citric acid and
extracted with EtOAc. The combined organic layers were washed
with brine, dried over MgSO4, and concentrated. The compound
was purified by flash chromatography in 5 to 20% EtOAc in hep-
tane to give a colourless oil (978 mg, 67%). Rf =0.27 (heptane/
1
(method A); H NMR (400 MHz, CDCl3): d=7.86 (d, 2H, J=8.1 Hz),
7.37 (d, 2H, J=8.1 Hz), 3.75 (s, 3H), 3.36 (dd, 1H, J=4.1, 7.1 Hz),
2.78 (d, 1H, J=7.1 Hz), 2.58 (d, 1H, J=4.1 Hz), 2.47 ppm (s, 3H);
13C NMR (100 MHz, CDCl3): d=167.2, 145.3, 134.0, 129.9, 128.2,
52.9, 35.7, 31.9, 21.7 ppm; HRMS (ESI): m/z calcd for C11H13NO4S+
H+: 256.0644 [M+H+]; found 256.0633.
1
EtOAc 3:1); HPLC tR: 7.3 min (method A); H NMR (400 MHz, CDCl3):
Methyl 1-((4-nitrophenyl)sulfonyl)aziridine-2-carboxylate (4): TFA
(1.8 mL, 23.5 mmol) was added dropwise to a solution of methyl
(S)-(À)-1-tritylaziridine-2-carboxylate (502 mg, 1.46 mmol) in CHCl3
and MeOH (2.6 mL, 50:50) at 08C. The solution was stirred at 08C
for 2.5 h. The solvents were removed, and the remaining solid was
dried with Et2O (5ꢁ4 mL). The residue was dissolved in Et2O
(30 mL) and H2O (30 mL). The ether layer was extracted with H2O
(20 mL). The combined aqueous fractions were neutralised with
solid NaHCO3 and diluted with EtOAc (30 mL). The mixture was
cooled to 08C, and p-nitrobenzenesulfonyl chloride (324 mg,
1.46 mmol) was added. The solution was stirred at room tempera-
ture overnight. The organic and aqueous phases were separated,
and the aqueous phase was extracted with EtOAc (40 mL). The or-
ganic phases were combined and concentrated. The compound
was purified by flash chromatography in heptane/EtOAc (2:1) to
give a yellow solid (332 mg, 79%). Rf =0.32 (heptane/EtOAc 2:1);
d=7.79 (d, 2H, J=7.5 Hz), 7.63 (d, 2H, J=7.5 Hz), 7.43 (t, 2H, J=
7.5 Hz), 7.35 (t, 2H, J=7.5 Hz), 4.51 (dd, 1H, J=7.2, 10.5 Hz), 4.39
(dd, 1H, J=7.2, 10.5 Hz), 4.27 (t, 1H, J=7.2 Hz), 3.77 (s, 3H), 3.13
(dd, 1H, J=3.2, 5.4 Hz), 2.64 (dd, 1H, J=1.2, 3.2 Hz), 2.49 ppm (dd,
1H, J=1.2, 5.4 Hz); 13C NMR (100 MHz, CDCl3): d=168.7, 160.8,
143.5, 141.3, 127.8, 127.1, 125.1, 120.0, 68.7, 52.8, 46.9, 34.8,
31.2 ppm; HRMS (ESI): m/z calcd for C19H17NO4 +H+: 324.1236 [M+
H+]; found 324.1227.
Radiochemistry
[18F] Fluorination with K222/K2CO3 and TEAHCO3: Aqueous
[18F]fluoride was trapped on a Waters QMA column and eluted
with either
a K222/K2CO3 solution (1 mL; 35 mm K222 and
36 mmK2CO3, 6% H2O in MeOH) or a TEAHCO3 solution (1 mL;
75 mm, H2O/MeOH 1:4). The solvent was removed by helium flow
at 1108C for 10 min, and excess H2O was removed by azeotropic
evaporation with MeCN (2ꢁ1 mL) at 1108C. The dried [18F]fluoride
was dissolved in either DMF or DMSO (2 mL).
1
HPLC tR: 8.4 min (method A); H NMR (400 MHz, CDCl3): d=8.43 (d,
2H, J=8.7 Hz), 8.21 (d, 2H, J=8.7 Hz), 3.78 (s, 3H), 3.48 (dd, 1H,
J=4.2, 7.1 Hz), 2.91 (d, 1H, J=7.1 Hz), 2.68 ppm (d, 1H, J=4.2 Hz);
13C NMR (100 MHz, CDCl3): d=166.6, 151.0, 143.0, 129.5, 124.5,
53.1, 36.2, 32.5 ppm; HRMS (ESI): m/z calcd for C10H10N2O6S+H+:
287.0338 [M+H+]; found 287.0330.
[18F] Fluorination with Chromafix and HCO3
:
Aqueous
À
[18F]fluoride was trapped on a Chromafix (PS-HCO3) column. The
column was washed with MeOH (2 mL) and the radioactivity
eluted with TBAOMs in MeOH (0.05m, 0.6 mL). The solvent was re-
moved at 1008C for 10 min with helium flow. The dried
[18F]fluoride was dissolved in DMSO (2 mL).
1-(tert-Butyl) 2-methylaziridine-1,2-dicarboxylate (5): Di-tert-butyl
dicarbonate (475 mg, 2.18 mmol) was added to a solution of meth-
ylaziridine-2-carboxylate (200 mg, 1.98 mmol) in MeCN (5 mL) and
Et3N (0.28 mL). The solution was stirred at room temperature over-
night. The mixture was concentrated, and the residue was dis-
solved in EtOAc (40 mL) and H2O (20 mL). The layers were separat-
ed, and the aqueous layer was extracted with EtOAc (3ꢁ40 mL).
The combined organic layers were washed with brine, dried over
[18F] Fluorination with Chromafix and K2PO4À: A Chromafix (PS-
HCO3) column was washed with 0.2m K3PO4 (5 mL) and dried with
air. Aqueous [18F]fluoride was trapped on the column, and the
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ChemistryOpen 2015, 4, 65 – 71 70

www.chemistryopen.org
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column was washed with MeOH (2 mL) and the radioactivity
eluted with TBAOMs in MeOH (0.05m, 0.6 mL). The solvent was re-
moved at 1008C for 10 min with helium flow. The dried
[18F]fluoride was dissolved in DMSO (2 mL).
General procedure for aziridine labelling with [18F]fluoride:
[18F]Fluoride in DMSO (0.1–0.2 mL, 10–550 MBq) was added to the
N-activated aziridine solution (5–8 mg) in DMSO. The solution was
heated by either conventional heat or microwave heating. Micro-
wave reactions were performed at 300 W in a closed vial. The yield
of the reaction was determined by HPLC method A.
[18F]Fluoride labelling with various precursor concentrations:
[18F]Fluoride in TEAHCO3 and DMSO (0.1 mL, 170–460 MBq) was
added to compound 5 (0.3–29.4 mg) in DMSO. The reaction was
heated at 1308C by microwave for 10 min. The yield of the reac-
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[18F]Fluoride labelling with various base concentrations:
[18F]Fluoride in TEAHCO3 and DMSO (2.5–500 mL, 40–1100 MBq)
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[18F]fluoride labelling of compound 5 were separated from the free
[18F]fluoride by collecting the peaks from HPLC. The Boc group and
methyl ester were removed by addition of 1m HCl and heating at
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purity of >99%. The product was analysed by HPLC method B and
identified by co-injection of reference a-fluoro-b-alanine. Due to
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detection of the UV detector.
From [18F]-labelled compound 6: The two products formed in the
[18F]fluoride labelling of compound 6 were separated from free
[18F]fluoride by collecting the peaks from HPLC. The Cbz group and
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>99%. The product was analysed by HPLC method B.
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This work was generously supported by the Danish Research
Council, the Novo Nordisk Foundation, the Lundbeck Foundation,
the Svend Andersen Foundation, Innovationsfonden, Director Mi-
chael Hermann Nielsen Memorial Fund, the Else and Mogens
Wedell-Wedellsborg Foundation, and the John and Birthe Meyer
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Keywords: aziridines
·
fluorine-18
·
radiochemistry
·
regioselectivity · ring opening
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ꢀ 2015 The Authors. Published by Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
ChemistryOpen 2015, 4, 65 – 71 71