Two-step regio- and stereoselective syntheses of [19F]- and [18F]-2-deoxy-2-(R)-fluoro-β-D-allose

Article abstract of DOI:10.1016/j.carres.2005.12.002

Replacement of specific hydroxyl groups by fluorine in carbohydrates is an ongoing challenge from chemical, biological, and pharmaceutical points of view. A rapid and efficient two-step, regio- and stereoselective synthesis of 2-deoxy-2-(R)-fluoro-β-d-allose (2-(R)-fluoro-2-deoxy-β-d-allose; 2-FDβA), a fluorinated analogue of the rare sugar, d-allose, is described. TAG (3,4,6-tri-O-acetyl-1,5-anhydro-2-deoxy-d-arabino-hex-1-enitol or 3,4,6-tri-O-acetyl-d-glucal), was fluorinated in anhydrous HF with dilute F ₂ in a Ne/He mixture or with CH₃COOF at -60°C. The fluorinated intermediate was hydrolyzed in 1 N HCl and the hydrolysis product was purified by liquid chromatography and characterized by 1D ¹H, ¹³C, and ¹⁹F NMR spectroscopy as well as 2D NMR spectroscopy and mass spectrometry. In addition, ¹⁸F-labeled 2-deoxy-2-(R)-fluoro-β-d-allose (2-[¹⁸F]FDβA) was synthesized for the first time, with an overall decay-corrected radiochemical yield of 33 ± 3% with respect to [¹⁸F]F₂, the highest radiochemical yield achieved to date for electrophilic fluorination of TAG. The rapid and high radiochemical yield synthesis of 2-[¹⁸F] FDβA has potential as a probe for the bioactivity of D-allose.

Full text of DOI:10.1016/j.carres.2005.12.002

Carbohydrate
RESEARCH
Carbohydrate Research 341 (2006) 457–466
Two-step regio- and stereoselective syntheses of
[19F]- and [18F]-2-deoxy-2-(R)-fluoro-b-D-allose
b
Rezwan Ashique,a,b Raman V. Chirakal,a,b, Donald W. Hughes and
*
Gary J. Schrobilgenb,*
aDepartment of Nuclear Medicine, Hamilton Health Sciences, 1200 Main Street West, Hamilton, Ontario, Canada L8N 3Z5
bDepartment of Chemistry, McMaster University, Hamilton, Ontario, Canada L8S 4M1
Received 13 September 2005; received in revised form 2 December 2005; accepted 12 December 2005
Available online 17 January 2006
Abstract—Replacement of specific hydroxyl groups by fluorine in carbohydrates is an ongoing challenge from chemical, biological,
and pharmaceutical points of view. A rapid and efficient two-step, regio- and stereoselective synthesis of 2-deoxy-2-(R)-fluoro-b-D-
allose (2-(R)-fluoro-2-deoxy-b-D-allose; 2-FDbA), a fluorinated analogue of the rare sugar, D-allose, is described. TAG (3,4,6-tri-O-
acetyl-1,5-anhydro-2-deoxy-D-arabino-hex-1-enitol or 3,4,6-tri-O-acetyl-D-glucal), was fluorinated in anhydrous HF with dilute F2
in a Ne/He mixture or with CH3COOF at ꢀ60 ꢀC. The fluorinated intermediate was hydrolyzed in 1 N HCl and the hydrolysis prod-
uct was purified by liquid chromatography and characterized by 1D 1H, 13C, and 19F NMR spectroscopy as well as 2D NMR spec-
troscopy and mass spectrometry. In addition, 18F-labeled 2-deoxy-2-(R)-fluoro-b-D-allose (2-[18F]FDbA) was synthesized for the
first time, with an overall decay-corrected radiochemical yield of 33 3% with respect to [18F]F2, the highest radiochemical yield
achieved to date for electrophilic fluorination of TAG. The rapid and high radiochemical yield synthesis of 2-[18F]FDbA has poten-
tial as a probe for the bioactivity of D-allose.
ꢁ 2005 Elsevier Ltd. All rights reserved.
Keywords: 2-Deoxy-2-(R)-fluoro-D-allose; PET tracer; Regio- and stereoselective fluorination; Elemental fluorine; Fluorine-18
1. Introduction
to advantage for the diagnosis and management of can-
cer by use of the 18F-labeled analogue of 2-deoxy-D-glu-
cose, 2-[18F]fluoro-2-deoxy-D-glucose (2-[18F]FDG), as
a glucose tracer in conjunction with positron emission
tomography (PET).
Substitution of a hydroxyl group by fluorine usually
results in molecules that are metabolized without C–F
bond attack. Deoxyfluorinated sugars therefore allow
studies of carbohydrate transport and metabolism and
are key to the isolation of specific biochemical reaction
sequences from general metabolic pathways, especially
in the functional imaging sciences.1 Furthermore,
18F-labeled (t1/2 = 110 min) sugars play an important
role in medical applications, including the diagnoses
of pathophysiological processes.2 For example, the ele-
vated glucose metabolism in tumor cells has been used
There have been several recent publications related to
the biological functions of D-allose. For example,
D-allose provides very effective protection against neu-
trophil-related postischemic injury of liver tissue3 and
inhibits segmented neutrophil production and lowers
platelet count without detrimental side effects.4 Such
characteristics make D-allose a candidate for the treat-
ment of diseases such as chronic myelogenous leukemia.
By analogy with glucose and its fluorinated derivative,
2-FDG, it appeared desirable to synthesize 2-deoxy-2-
fluoro-D-allose, which, when labeled with 18F, could
serve as a probe of the bioactivity of D-allose.
*
Corresponding authors. Tel.: +1 905 521 2100x76893; fax: +1 905
546 1125 (R.V.C.); tel.: +1 905 525 9140x23306; fax: +1 905 522
2509 (G.J.S.); e-mail addresses: chriraklr@mcmaster.ca; schrobil@
mcmaster.ca
Johansson and Lindberg5 have reported the only syn-
thesis of 2-deoxy-2-fluoro-D-allose prior to the present
0008-6215/$ - see front matter ꢁ 2005 Elsevier Ltd. All rights reserved.
doi:10.1016/j.carres.2005.12.002

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R. Ashique et al. / Carbohydrate Research 341 (2006) 457–466
work. Their lengthy, multi-step synthesis was carried out
by the reaction of an anhydrous chloroform (ethanol-
free) solution of the sugar epoxide, methyl 4,6-di-O-acet-
yl-2,3-anhydro-a-D-allopyranoside, with BF3 in HF at
ꢀ70 ꢀC. Hydrolysis and deprotection of the reaction
intermediate resulted in 2-deoxy-2-fluoro-D-altrose,
68% of which was epimerized to 2-deoxy-2-fluoro-D-
allose in 33% aqueous trimethylamine at 60 ꢀC over a
period of 5 h. The final reaction product was a mixture
of the a- and b-anomers. Although the authors referred
ization occurred at C3, with the configuration at C3
changing from S in the starting material to R in the
product.10
Drawing on the synthetic approach of Lundt and
Pedersen10 and the established synthetic procedure used
to introduce fluorine at C2 in hexapyranoses,6,7,12–14
a rapid and efficient two-step synthesis of 2-deoxy-2-
(R)-fluoro-b-D-allose (2-(R)-fluoro-2-deoxy-b-D-allose;
2-FDbA), resulting from electrophilic fluorination of
TAG, is reported in this paper. A detailed 1H, 13C,
and 19F NMR study has provided the first unambiguous
structural characterization of 2-FDbA. In addition, the
synthesis of 18F-labeled 2-FDbA (2-[18F]FDbA), using
[18F]F2 as the electrophilic fluorinating agent, is reported
for the first time.
1
to unpublished H and 19F NMR data to support their
structural characterization, no NMR parameters were
provided.
One of the most prevalent synthetic procedures used
to introduce fluorine at C2 in carbohydrate molecules
is electrophilic addition of fluorine to glycals. Such
compounds often show high regio- and stereoselectivity
in addition reactions.2 Adamson et al.6,7 first reported
the syntheses of 2-deoxy-2-fluoro sugars by addition
of CF3OF to 3,4,6-tri-O-acetyl-D-glucal (TAG) dis-
solved in CFCl3 at ꢀ80 ꢀC followed by acid catalyzed
hydrolysis of the reaction intermediates. The synthesis
produced 2-fluoro-2-deoxy-D-glucose (2-FDG) and
2-fluoro-2-deoxy-D-mannose (2-FDM) in good yields.
Since then, TAG has been one of the most widely used
precursors for the syntheses of 2-deoxy-2-fluoro-
sugars.
The introduction of a fluorine atom at C2 by addition
to glycals is effected by so-called ‘electrophilic fluorinat-
ing reagents, Rdꢀ–Fd+’. Because C2 of a glycal is more
electronegative than C1 as a result of resonance contri-
butions –O–C(1)@C(2) M –O+@C(1)–Cꢀ(2), the fluo-
rine of R–F always adds to C2. Reagents in this
category include CF3OF, F2 diluted with an inert gas,
CH3COOF, and XeF2.8 Because almost all electrophilic
fluorinating agents are the products of direct fluorina-
tion by F2, it is desirable to use F2 directly as the electro-
philic fluorinating agent, thereby eliminating synthetic
steps.
2. Results and discussion
2.1. Synthesis of 2-FDbA
The synthesis of 2-FDbA was accomplished by electro-
philic fluorination of commercially available TAG with
F2 or CH3COOF in aHF at ꢀ60 ꢀC, followed by acid
catalyzed hydrolysis and purification using liquid chro-
matography (Scheme 1). The compound was character-
ized by conventional 1D 1H, 13C, and 19F NMR
spectroscopy, selective 1D 1H and 2D 1H–1H and
1H–13C correlation spectroscopy experiments, and by
mass spectrometry.
2.2. Structural characterization of 2-FDbA by 1D and 2D
NMR spectroscopy
Among the methods for determining the compositions
and stereochemistries of sugar mixtures, NMR spectro-
scopy has emerged as the most reliable.15 The structures
of monosaccharides and fluorinated monosaccharides
can be readily analyzed, in most instances, by conven-
Lundt and Pedersen9–11 extensively studied the reac-
tions of TAG and other 3,4,6-tri-O-substituted glycals
with and in anhydrous hydrogen fluoride (aHF) solvent
and in mixed solvent media containing HF. The reaction
of TAG with aHF resulted in a 1,2-unsaturated 3,4-
dioxolenium ion, which was stable in aHF at ꢀ70 ꢀC
for several hours. The dioxolenium ion was hydrolyzed
in the presence of a small amount of water and epimer-
tional 1D NMR techniques and by reference to the H
1
and 13C NMR parameters of related compounds.16
Moreover, the conformations of the deoxyfluorinated
1
sugars can be reliably determined by H NMR spectro-
scopy.17 Whereas resonances of non-anomeric protons
of different conformers usually overlap, those of anomeric
protons are often well separated from each other.15
1
In cases where H spectral data are difficult to obtain,
OAc
6
OH
5
O
O
F
OH
F2 or AcOF
aHF
1N HCl
130 oC
17 min
4
1
several minor products
+
OAc
3
OAc
-60 o
C
OH
2
OH
TAG
Scheme 1. Synthesis of 2-(R)-fluoro-2-deoxy-b-D-allose (2-FDbA).
2-FDβA

R. Ashique et al. / Carbohydrate Research 341 (2006) 457–466
459
19F NMR spectroscopy can be used to probe the config-
uration and/or conformation of fluorocarbohydrates.18
The 1D 1H NMR spectrum (Fig. 1) of the products of
Scheme 1 was too complex to permit full and unambig-
uous analysis and extraction of NMR parameters. How-
ling between H1 and F, and established that the fluo-
rine atom was bonded to C2. Moreover, the larger
coupling (8.2 Hz) showed that the monosaccharide pos-
sessed a trans-diaxial arrangement of hydrogen atoms at
C1 and C2. As a result, the absolute configuration at C1
was assigned as R, i.e., the fluorocarbohydrate had the
b-form. Furthermore, comparison of the magnitudes
of 3JH1,H2 and 3JH1,F measured in the present study with
those of the b-anomers of 2-FDG (3JH1,H2 = 7.8 Hz;
3JH1,F = 2.5 Hz)19,20 and 2-fluoro-2-deoxy-D-galactose
1
ever, the H resonances corresponding to the anomeric
proton could be readily assigned. The full NMR struc-
tural characterization of 2-FDbA was accomplished by
1
carrying out selective 1D H and 2D correlation spec-
troscopy experiments. The 1H NMR parameters for
2-FDbA are summarized in Table 1.
3
(3JH1,H2 = 7.3 Hz; JH1,F = 3.0 Hz)21–23 indicated that
The 1H NMR spectrum showed only one type of ano-
meric proton resonance. Thus, it could be immediately
concluded that the major carbohydrate product synthe-
sized in Scheme 1 existed in either the a- or b-form. The
doublet of doublets (dd), centered at 5.21 ppm, was as-
signed to the anomeric proton, H1. The smaller coupling
(1.4 Hz) was characteristic of an axial–equatorial coup-
2-fluoro-2-deoxy-D-allose predominantly existed as the
b-form in equilibrated aqueous solutions of the prod-
ucts. This is consistent with the observation that the
anomeric equilibrium for the D-ribo- and D-allo-pyra-
nose series favors the b-D-pyranose form.24–26 This is
also in agreement with the reported compositions of
D-allose and D-gulose and their amino sugar derivatives
in equilibrium mixtures in which the b-pyranose forms
of the sugars dominate (Table 2).
1
A H–1H gradient COSY spectrum (Fig. 2) showed a
strong correlation between the H1 resonance and a dou-
blet of doublets of doublets centered at 4.38 ppm. Selec-
tive 1D TOCSY experiments and coupling constants
were used to assign the multiplets to H2. The magni-
2
3
tudes of the JH2,F,
3JH2,H1, and JH2,H3 couplings
unequivocally established the absolute configuration at
1
C2 as R. The H–1H COSY and selective 1D TOCSY
experiments were also used to assign the H3 resonance
to the doublet of pseudo-triplets at 4.51 ppm. The larger
(8.9 Hz) coupling is characteristic of equatorial–equato-
rial coupling between H3 and fluorine bonded to C2.
Each pseudo-triplet resulted from the overlap of two
sets of doublets having an average J-value of 3.0 Hz
and resulted from the three-bond equatorial–axial coup-
Figure 1. The 1H NMR spectrum (D2O solvent at 25 ꢀC) of the
products resulting from the electrophilic fluorination of TAG in aHF.
The spectrum was processed using Gaussian multiplication and
presaturation of the HDO solvent resonance at 4.77 ppm. Dots (•)
3
3
3
lings, JH2,H3 and JH3,H4. The magnitudes of JH3,F
,
3
3JH3,H4, and JH3,H2 confirmed the R-configuration at
C3.
and asterisks ( ) denote H4 and H60 resonances, respectively. The
*
In addition to correlation between H2 and H3, the
1H–1H COSY spectrum showed correlation between
H3 and the proton(s) responsible for the complex multi-
plet centered at 3.78 ppm. Although it was certain that
this multiplet included the H4 resonance, it was appar-
ent from peak integration that the multiplet corre-
sponded to two hydrogen atoms. A selective 1D
COSY experiment showed that the multiplet’s complex-
ity and integrated intensity resulted from overlap of the
H4 and H60 resonances. The magnitudes of the trans-
diaxial coupling between H4 and H5 (10.0 Hz) and the
three-bond axial–equatorial coupling between H4 and
H3 (3.0 Hz) confirmed that the absolute configuration
at C4 was R. A long-range coupling (1.4 Hz) between
H4 and the fluorine bonded to C2 was also observed.
The H5 and the diastereotopic H6 and H60 resonances
dagger ( ) denotes an unassigned ‘triplet’ (d(1H) 4.39 ppm; 1.6 Hz
splitting).
Table 1. 1H NMR parameters for 2-FDbA
1H
Multiplicitya
Coupling constant (Hz)
Chemical
shift (ppm)
H1
H2
H3
H4
H5
H6
H60
5.21
4.38
4.51
3.78
3.90
3.97
3.78
dd
3JH1,F = 1.4
3JH1,H2 = 8.2
3JH2,H3 = 3.1
3JH3,H4 = 3.0
ddd
dddb
ddd
ddd
dd
2JH2,F = 47.0
3JH3,F = 8.9
4JH4,F = 1.6
3JH4,H5 = 10.0
3JH5,H6 = 2.3
JH5;H6 ¼ 5:8
3
0
2
0
JH6;H6 ¼ 12:3
dd
a The abbreviations denote doublet of doublets (dd) and doublet of
doublets of doublets (ddd).
1
were assigned by the use of a H–1H COSY experiment
b This multiplet appears as
a
doublet of ‘triplets’ because
3
3JH3,H4 ꢁ JH2,H3
.
as well as by the magnitude of the coupling between H4

460
R. Ashique et al. / Carbohydrate Research 341 (2006) 457–466
Table 2. The compositions (%) of D-allose and D-gulose and their amino sugars in D2O
Compound
Temp (ꢀC)
Pyranose (%)
Furanose (%)
References
a
b
a
b
D-Allose
D-Gulose
2-Acetamido-2-deoxy-D-allose
2-Acetamido-2-deoxy-D-gulose
31
22
23–25
23–25
14
16
14
17
77.5
3.5
—
9
5
3
5
6
27
28
26
26
81
72
74
3
Figure 3. The 13C NMR spectrum (D2O solvent at 25 ꢀC) of the
products resulting from the electrophilic fluorination of TAG in aHF.
Figure 2. The 1H–1H gradient COSY spectrum (D2O solvent at 25 ꢀC)
of the products resulting from the electrophilic fluorination of TAG in
aHF.
Table 3. 13C NMR parameters for 2-FDbA
13C Chemical shifta
(ppm)
Multiplicityb
Coupling constant (Hz)
2JC1,F = 24.5
1JC2,F = 185.5
2JC3,F = 15.9
3JC4,F = 5.5
and H5. The doublet of doublets centered at 3.90 ppm
was assigned to H5. As noted above, the 10.0 Hz cou-
C1
C2
C3
C4
C5
C6
91.93
90.67
70.09
67.05
74.47
61.75
(96.4)
(74.2)
(74.3)
(69.5)
(76.5)
(64.1)
d
d
d
d
s
3
pling arose from JH5,H4. The two smaller couplings,
3
3
0
5.8 and 2.3 Hz, corresponded to JH5,H6 and JH5;H6
,
respectively, which arose from coupling of H5 to the dia-
stereotopic H6 (3.97 ppm) and H60 (3.78 ppm) protons.
Assignments of the 13C resonances (Fig. 3 and Table
3) were aided by the 13C–19F coupling constants and
by a 13C–1H heteronuclear single quantum correlation
(HSQC) experiment. The 13C NMR spectrum provided
further evidence that Scheme 1 produced 2-FDbA as
the major carbohydrate product. The 13C resonance of
the anomeric carbon, C1, appeared as a doublet (d) at
the highest frequency, 91.93 ppm, with 2JC1,F = 24.5 Hz.
The chemical shift of C2, bearing the fluorine ligand and
being adjacent to the anomeric carbon atom, occurred
at 90.67 ppm and gave rise to a one-bond 13C–19F cou-
pling (185.5 Hz) that was consistent with fluorine
bonded to an sp3 hybridized carbon. Carbon-3 reso-
nance appeared at 70.09 ppm and the magnitude of
s
a Chemical shifts in parentheses are those of b-D-allose.29
b The abbreviations denote doublet (d) and singlet (s).
those of C3 and C4, because C5 is bonded to the ring
oxygen. The C6 resonance (61.75 ppm) appeared at the
lowest frequency.
The 1D 19F NMR spectrum of the products (Fig. 4)
showed an intense doublet of doublets of pseudo-triplets
centered at ꢀ199.99 ppm, corresponding to 2-FDbA,
along with a weak, unassigned doublet of doublets cen-
tered at ꢀ199.44 ppm. The largest coupling (47.0 Hz)
was assigned to the two-bond coupling between fluorine
and H2. The three-bond coupling between fluorine and
H3 was 8.2 Hz. Finally, the pseudo-triplet resulted from
overlap of two doublets (average J-value, ca. 1.4 Hz)
2
the JC3,F coupling was 15.9 Hz. The three-bond cou-
3
4
pling between C4, which appeared at 67.05 ppm, and
the fluorine environment was 5.5 Hz. The chemical shift
of C5 (74.47 ppm) appeared at higher frequency than
corresponding to JF,H1 and JF,H4. The 1D 19F NMR
spectrum confirmed that 2-FDbA was the major prod-
uct of the electrophilic fluorination of TAG in aHF. It

R. Ashique et al. / Carbohydrate Research 341 (2006) 457–466
461
confirmed that 2-FDbA was the major product. The
spectrum showed the deprotonated molecular ion,
[MꢀH]ꢀ, corresponding to 2-FDbA at m/z 181. How-
ever, the base peak in the spectrum appeared at m/z
217, which corresponds to the chloride adduct of neutral
2-FDbA.
2.4. Proposed mechanistic route to 2-FDbA
Fluorination of glycals by F2 and AcOF in polar sol-
vents and aqueous solutions usually results in difluoro
and 2-deoxy-2-fluoro intermediates, respectively, in
which the electrophilic fluorinating agent is added across
the double bond to give a syn-arrangement.12–14 The
absence of an anti-isomer is attributed to the fact that,
unlike other halogens, fluorine cannot form a halonium
ion. Therefore, the initial fluorocarbonium ion interme-
diate resulting from electrophilic attack of C2 by fluo-
rine is not bridged.35 The carbonium ion rapidly
combines with the counter ion, Fꢀ or AcOꢀ. Both the
fluorine ligand and acetoxy group on C1 are susceptible
to hydrolysis in dilute acid media.
Figure 4. The 19F NMR spectrum (D2O solvent at 30 ꢀC) of the
products resulting from the electrophilic fluorination of TAG in aHF.
The spectrum was processed using Gaussian multiplication. Asterisks
( ) denotes an unassigned resonance (‘doublet of doublets’) arising
*
2
from a minor component (d(19F), 199.44 ppm; JF,H, 44.35 Hz and
3JF,H, 7.67 Hz) characteristic of a fluorocarbohydrate.
is significant that neither 2-FDG nor 2-FDM was pro-
duced in this reaction, whereas electrophilic fluorina-
tions of TAG with F2 and AcOF in any other solvent
medium result in both 2-FDG and 2-FDM13,30–34 (also
see synthesis of 2-[18F]FDbA).
Lundt and Pedersen10 proposed that the reaction of
TAG with aHF (Scheme 2) proceeded by protonation
of the oxygen atom bonded to C3 and subsequent cleav-
age of the O-acetyl group was likely assisted by attack
from the acyloxy group at C4. These workers showed,
from a freshly prepared aHF solution of TAG that the
double bond of the corresponding 1,2-unsaturated 3,4-
dioxolenium ion was initially intact at ꢀ70 ꢀC. After a
period of 24 h, the 1H NMR spectrum showed
that aHF had added across the double bond of the
2.3. Mass spectrometric results
Negative ion electrospray ionization mass spectrometry
of the reaction products resulting from Scheme 1 also
OAc
OAc
OAc
O
O
O
aHF
aHF
-70 oC
<1 h
-
+
F
OAc
OAc
O
O
+
O
TAG
O
-70 oC
HF
>24 h
H2O HF
OAc
OAc
O
O
F
F
OAc
O
+
O
H2O
OH
O
F
OH
OAc
Scheme 2. Reaction of TAG with aHF.10

462
R. Ashique et al. / Carbohydrate Research 341 (2006) 457–466
OAc
O
OAc
F
OAc
OAc
O
O
O
_
O
O
H
F
H
+
+
δ
_
+
O
O
F
−
OAc
OAc
O
O
_
δ
+
O
TAG
F
+
+F
− F
R
H
−
δ
δ
O
O
+
δ
δ
OH
OAc
OH
OAc
O
F
O
F
O
F
OH
1N HCl
17 min
130 oC
-HF
95:5 ACN:H2O
+
R
O
OH
OH
_
+
_
R
O
OH
OAc
F
2-FDβA
+
− F
δ
δ
H
R = F or AcO
Scheme 3. A proposed partial mechanism for the synthesis of 2-FDbA by electrophilic fluorination of TAG in aHF.
dioxolenium ion in a Markovnikov fashion. The pres-
ence of a small amount of water hydrolyzed the dioxo-
lenium ion and resulted in epimerization at C3.10
In the present study, fluorinations were routinely car-
ried out at ꢀ60 ꢀC within 30 min following addition of
aHF to TAG, thus avoiding significant HF addition to
the double bond of the 1,2-unsaturated dioxolenium
ion prior to fluorination. A proposed partial mechanism
for the synthesis of 2-FDbA in the present work is
provided in Scheme 3, which incorporates the existing
synthetic route to 2-deoxy-2-fluoro sugars and the
proposed synthetic route10 that gives rise to the 1,2-
unsaturated 3,4-dioxolenium ion and its epimerization
at C3.
The exclusive existence of the b-anomer of 2-deoxy-2-
(R)-fluoro-D-allose, as indicated in the 1H and 13C NMR
spectra, is consistent with the data presented in Table 2.
The high stereoselectivity of TAG toward fluorination in
aHF to give 2-deoxy-2-(R)-fluoro-D-allose, however, has
yet to be accounted for.
2.6. Synthesis of 18F-labeled 2-FDbA
To determine an overall product yield with respect to F2,
the electrophilic fluorination of TAG in aHF was car-
ried out using [18F]F2 as the electrophilic fluorinator.
The overall decay-corrected radiochemical yield (RCY)
of the resulting 2-[18F]FDbA was 33 3% with respect
to [18F]F2, which is the highest RCY achieved to date
for electrophilic fluorination of TAG using [18F]F2 in
any solvent system. The radiochemical purity of
2-[18F]FDbA was 96.3 3 and 85 7%, as determined
by radio-HPLC and radio-TLC, respectively. Further-
more, 19F NMR studies showed that neither
2-[18F]FDG nor 2-[18F]FDM were produced. The
former is widely used for the study of local glucose
metabolism in humans and the latter is a signi-
ficant contaminant when 2-[18F]FDG is produced by
electrophilic fluorination methods. The absence of
2-[18F]FDG and 2-[18F]FDM in 2-[18F]FDbA is a signi-
ficant factor when used for PET imaging purposes,
because the presence of either contaminant would
contribute to background noise, reducing the signal-to-
noise ratio in the resulting image, and would unnecessar-
ily increase the radiation dose to the patient.
2.5. Synthesis of 2-FDbA in weakly acidic media
The reactivities of F2 toward TAG in weakly acidic
media, namely trifluoroacetic acid (TFA) and formic
acid (FA), were also studied. These reactions, however,
provided significantly lower stereoselectivities for 2-
FDbA than those carried out in aHF. In both TFA
and FA, electrophilic fluorination of TAG produced 2-
FDG, 2-FDM and 2-FDbA in 1.3:1.0:1.6 ratios, based
on their integrated 19F NMR spectra. This demonstrates
that TAG does not completely convert to the 1,2-unsat-
urated 3,4-dioxolenium ion in weakly acidic media and
that a significant amount of the precursor is intact at
the time of fluorination.
3. Conclusion
A reliable and rapid two-step highly regio- and stereose-
lective synthesis of 2-FDbA by electrophilic fluorination
of TAG in aHF has been achieved. The high regioselec-
tivity results from exclusive fluorination at C2. Stereo-
selectivity resulted in an R configuration at C2 of the
product, whereas all other known electrophilic fluorina-
tions of TAG result in products having both R and S

R. Ashique et al. / Carbohydrate Research 341 (2006) 457–466
463
configurations at C2. Such high regio- and stereoselec-
tivities are often hard to achieve from electrophilic fluo-
rinations. The total synthesis time was approximately
45 min, which is more rapid and efficient than the exist-
ing synthesis of 2-deoxy-2-fluoro-D-allose reported by
Johansson and Lindberg.5 In addition, 2-[18F]FDbA
has been synthesized for the first time with an overall
decay-corrected RCY of 33 3% with respect to [18F]F2.
Work is underway to determine the potential applica-
tion of 2-[18F]FDbA in PET as a tracer for D-allose
metabolism.
enriched [18O]O2 and irradiated for 30 min using a
30 lA proton beam (production shoot). After irradiation,
the [18O]O2 was recovered from the target by condensing
it at ꢀ196 ꢀC onto molecular sieves (Varian VacSorb)
contained in a 316 stainless steel Whiteyꢂ cylinder
(75 mL). The target was then evacuated to remove resid-
ual 18O, flushed with neon (ca. 7 atm), and re-evacuated.
The target was then filled with 1% F2 (30–50 lmol) in
neon, pressurized to 20 atm with neon, and irradiated
for 15 min in a 15 lA proton beam (recovery shoot).
The [18F]F2 was sporadically released from the target
into a continuous stream of helium (10 mL/min) until
the target pressure dropped to 2 atm. Helium was used
as the sweep gas to transfer [18F]F2 from the target into
the hot cell and to help dissipate the heat evolved during
the fluorination reaction.
4. Experimental
Caution: It is recommended that proper first-aid treat-
ment procedures36–38 be known and available to all lab-
oratory personnel prior to repeating portions of this
study that deal with the use of aHF and F2. Skin contact
with even small amounts of aHF or F2 may result in
painful burns if immediate and proper treatment is not
given. Any incident involving direct contact with liquid
aHF, HF vapor, F2 gas, and aqueous solutions of HF
must be aggressively treated and brought to the atten-
tion of qualified medical personnel for appropriate fol-
low-up treatment.
4.4. Electrophilic fluorinations of TAG in aHF using F2
and [18F]F2
TAG (20 mg; 73.5 lmol) was loaded into a 5/16 in.
o.d. · 5/32 in. i.d. FEP (perfluoroethylene/perfluoro-
propylene co-polymer) reaction vessel and attached to
a Kel-F (trifluorochloropolyethylene polymer) Y-piece
with 1/4 in. o.d. · 1/8 in. i.d. ends. One arm of the Y-
piece was connected through a Kel-F valve to a vacuum
line used to dispense aHF. A 1/16 in. o.d. · 1/32 in. i.d.
length of Teflon tubing was fed through a 1/4 in.
o.d. · 1/16 in. Teflonꢂ Swagelokꢂ reducing union and
connected to the remaining arm of the Y-piece. The Tef-
lonꢂ tube was fed into and to the bottom of the FEP
reaction vessel and sealed at the 1/16 in. Teflonꢂ union
connection. Anhydrous HF was condensed into the
reaction vessel at ꢀ196 ꢀC. The reaction vessel and con-
tents were equilibrated at ꢀ60 ꢀC and the reaction vessel
was disconnected from the vacuum line. The Teflonꢂ
tube was connected to a stainless steel vacuum manifold
(28.5 mL) used to dispense F2. The initial manifold pres-
sure of 8 atm of 1% F2 (ca. 95 lmol) in neon was
adjusted to 27 atm with helium. The remaining arm of
the Y-piece was connected to a separate 1/16 in. o.d.
Teflonꢂ tube, which was led into a 1 N NaOH solution
that served to trap the HF and unreacted F2 gases as
they emerged from the reaction vessel. Approximately
20 atm of the resulting F2/Ne/He mixture (ca. 70 lmol
of F2) was passed through the substrate solution.
4.1. Standard techniques
The use of radioactive isotopes was carried out in a safe
and effective manner in compliance with all require-
ments of the Canadian Nuclear Safety Commission
(CNSC) and Radioisotope Protection Committee
(RPC).
4.2. Materials
Enriched [18O]O2 (18O, 99 at. %, Isotec), neon (99.999%,
Air Products), 1% F2 in neon (Canadian Liquid Air),
helium (99.9999%, Matheson), anhydrous hydrogen
fluoride (Air Products, 99.9%), trifluoroacetic acid
(Caledon, 99%), formic acid (Sigma–Aldrich; 98–
100%), 3,4,6-tri-O-acetyl-D-glucal (Aldrich, 98%), anhy-
drous ethyl ether (Aldrich, 99.0%), and HPLC grade
acetonitrile (Caledon, 99.8%) were used without further
purification and/or drying. Sterile, deionized water was
used in all aqueous procedures.
When electrophilic fluorinations were carried out
using [18F]F2, the reactor configuration remained un-
changed except that the Teflonꢂ tube was connected to
the [18F]F2 target line. A helium sweep gas line was con-
nected to the [18F]F2 target line (between the target and
the reaction vessel) and helium was slowly passed
through the line. Aliquots of [18F]F2 gas were released
from the target by periodically opening and closing the
target valve. Fluorine-18 labeled F2 gas and the helium
sweep gas were passed through the reaction vessel
4.3. Production of [18F]F2
Fluorine-18 labeled F2 was produced by the 18O(p,n)18F
nuclear reaction using a Siemens RDS 112 proton cyclo-
tron operating at 11 MeV by the use of ‘double shoot’
method39 in the Nuclear Medicine Department, Hamil-
ton Health Sciences as previously described.40 An alumi-
num target (11 mL) was pressurized to 15–17 atm with

464
R. Ashique et al. / Carbohydrate Research 341 (2006) 457–466
containing the substrate solution. The effluent gas from
the reaction vessel was trapped in the NaOH solution
before it was vented into the hot cell. The amount of
[18F]F2 that had reacted was determined by counting
the amount of radioactivity present in the reaction
mixture.
4.7. Hydrolysis of the reaction intermediate resulting from
electrophilic fluorination of TAG
The reaction intermediate was recovered from the reac-
tion vessel by dissolution in a 5% water/CH3CN, fol-
lowed by two rinsings with the same solvent mixture.
The combined rinsings were then passed through a silica
Sep-Pakꢂ (Waters Corporation, Milford, Massachu-
setts, USA) and collected in a 10 mL Reacti-Vialꢂ
equipped with a magnetic stirring bar. The solvent was
evaporated to dryness under a slow stream of nitrogen
gas in a Reacti-Thermꢂ (Pierce) set at 130 ꢀC. A
2.0 mL aliquot of 1 N HCl was added to the Reacti-
Vialꢂ and the residue was hydrolyzed at 130 ꢀC for
17 min.
Once the desired amount of F2 or [18F]F2 gas had
passed through the substrate solution, the reaction ves-
sel was disconnected from the F2 line and the NaOH
trap and the HF solvent was removed under dynamic
vacuum and trapped in a FEP U-tube cooled to
ꢀ196 ꢀC.
4.5. Gas–solid phase generation of CH3COOF and
[18F]CH3COOF from KOAc(HOAc)1.5
The complex, KOAc(HOAc)1.5, was prepared as previ-
ously described.41 A 2 in. long 1/4 in. o.d. · 1/8 in. i.d.
FEP column, fitted with a 1/4 in. o.d. · 1/16 in. Teflonꢂ
Swagelokꢂ reducing union on one end and a 1/4 in. to
1/16 in. stainless steel Swagelokꢂ reducing union on
the other end, was packed to a depth of 1 in. with
KOAc(HOAc)1.5. The Teflonꢂ Swagelokꢂ union was
connected to a 1/16 in. o.d. stainless steel F2 line and
8 atm of 1% F2 in neon was diluted to 27 atm with
4.8. Separation and purification of the hydrolysis product
by liquid chromatography (LC)
A 1 cm o.d. · 17 cm long column (Bio-Rad) was packed
with ion retardation resin (Bio-Rad AGꢂ 11 A8, 50–100
mesh) and the column was washed with water. An alu-
mina Sep-Pakꢂ (Waters Corporation, Milford, Massa-
chusetts, USA) was conditioned with 10 mL of water
and attached to the end of the ion exchange column.
A C18 Sep-Pakꢂ (Waters Corporation, Milford, Massa-
chusetts, USA) was conditioned with 4 mL of absolute
ethanol followed by 10 mL of water and attached to
the end of the alumina Sep-Pakꢂ. The end of the C18
Sep-Pakꢂ was attached to a 30 mL multi-dose vial
through a 0.2 lM filter.
The hydrolysis product was added to the top of the
LC column and eluted with 10 mL of water into the
30 mL multi-dose vial. Ionic species and unhydrolyzed
organic compounds remained on the column while
ꢂ10 mL of the hydrolyzed product solution was col-
lected in the multi-dose vial.
helium inside
a stainless steel vacuum manifold
(28.5 mL). Approximately 20 atm of the resulting F2/
Ne/He mixture was passed through the column contain-
ing the KOAc(HOAc)1.5 complex, which resulted in the
formation of CH3COOF gas.
In the case of [18F]CH3COOF, the KOAc(HOAc)1.5
column was connected, through the Teflonꢂ Swagelokꢂ
union, to a 1/16 in. stainless steel tube leading from the
[18F]F2 target cell. After irradiation, the target gas was
slowly metered through the KOAc(HOAc)1.5 complex,
forming [18F]CH3COOF gas.
4.6. Electrophilic fluorinations of TAG in aHF using
CH3COOF and [18F]CH3COOF
4.9. NMR spectroscopy
The reaction conditions were identical to those
described above. The KOAc(HOAc)1.5 column used to
generate CH3COOF and [18F]CH3COOF gas was con-
nected to a 1/16 in. o.d. · 1/32 in. i.d. Teflonꢂ tube,
which was fed through one of the arms of a Kel-F Y-
piece into a 5/16 in. o.d. · 5/32 in. i.d. FEP reaction ves-
sel containing TAG (20 mg; 73.5 lmol) in aHF (ca.
1 mL) and connected as described above. The fluorina-
tion was carried out using the procedure described
above. Once the desired amount of CH3COOF or
[18F]CH3COOF had passed through the substrate solu-
tion, the reaction vessel was disconnected from
the CH3COOF or [18F]CH3COOF line and the NaOH
trap. Anhydrous HF solvent was removed under vac-
uum by pumping through an FEP U-tube cooled to
ꢀ196 ꢀC.
NMR samples were prepared by drying an aqueous
solution of the final products on a rotary evaporator
and redissolving the resulting thin film in D2O. Samples
were allowed to equilibrate for in excess of 36 h before
recording their NMR spectra. Samples were locked to
deuterium in D2O and spectra were acquired without
spinning the samples. Spectra were externally referenced
with respect to neat TMS (1H and 13C, 25 ꢀC) and neat
CFCl3 (19F, 30 ꢀC).
Proton and 13C NMR spectra were recorded at
600.130 and 150.903 MHz, respectively, on a Bruker
Avance 600 MHz NMR spectrometer using a 5-mm tri-
ple broad band inverse probe. The sample temperature
was maintained at 25 ꢀC by means of a BVT 3000 digital
temperature controller.

R. Ashique et al. / Carbohydrate Research 341 (2006) 457–466
465
Proton NMR spectra were obtained in 48 scans in
64 K data points over a 5.00 kHz spectral width corre-
sponding to an acquisition time of 6.55 s and a digital
resolution of 0.08 Hz/point. The residual HDO solvent
peak was suppressed by presaturation during a 1.5 s
relaxation delay between acquisitions. The data were
zero-filled to 128 K before Fourier transformation.
The free induction decays (FIDs) were processed using
Gaussian multiplication with a Gaussian broadening
of 0.1 and ꢀ2.00 Hz line broadening.
using the pulsed field gradient version of the heteronu-
clear single quantum coherence (HSQC) pulse sequence.
The FIDs in the F2 (1H) dimension were recorded over a
5.00 kHz spectral width in 2 K data points. The 256
FIDs in the F1 (13C) dimension were obtained over a
27.10 kHz spectral width. Each FID was acquired in
32 scans. The fixed delays during the pulse sequence
were a 1.0 s relaxation delay and a delay for polarization
1
transfer of 0.001786 s. The 90ꢀ H and 13C pulses were
8.0 and 14.0 ls, respectively. The data were processed
using a sine-bell squared window function shifted by
p/2 in both dimensions and a linear prediction to 256
data points in the F1 dimension followed by zero-filling
to 1 K.
Proton correlation spectroscopy (COSY) spectra were
recorded in the absolute value mode using the pulse
sequence 90ꢀ-t1-45ꢀ-ACQ and included pulse field gradi-
ents for coherence selection. The data were acquired in
16 scans for each of the 256 FIDs that contained 2 K
data points in the F2 dimension over a 5.00 kHz spectral
width. The 1H 90ꢀ pulse width was 8.0 ls. A 1.0 s relax-
ation delay was used between acquisitions. Zero-filling
in the F1 dimension produced a 1 K · 1 K data matrix
with a digital resolution of 4.88 Hz/point in both dimen-
sions. During the 2D Fourier transformation, a sine-bell
squared window function was applied to both dimen-
sions. The transformed data were then symmetrized.
Selective 1D total correlation spectroscopy (TOCSY)
1H spectra with pulsed field gradients were recorded
over an 8.09 kHz spectral width in 64 K data points cor-
responding to a 4.05 s acquisition time. Selective excita-
tion was provided by Gaussian-shaped pulses with a
180ꢀ pulse width corresponding to 83.4 ms. This pulse
was followed by the standard TOCSY MLEV-17 spin-
lock pulse sequence. The 90ꢀ spin-lock pulse width was
63.0 ls. A 1.0 s relaxation delay was used. The spin-lock
period was 100 ms, which was followed by a z-filter that
contained 10 variable delay times ranging from 4 to
18 ms. The transmitter offset was adjusted to the fre-
Fluorine-19 NMR spectra were recorded at
470.592 MHz on a Bruker Avance DRX-500 NMR
spectrometer using a 5-mm broad band inverse probe
with the 1H coil tuned to the 19F frequency. The spectra
were obtained over a 6.89 kHz spectral width in 64 K
data points corresponding to an acquisition time of
4.76 s and a data point resolution of 0.11 Hz/point.
Adequate signal-to-noise was achieved in 60 scans.
The 19F pulse width was 2.5 ls (30ꢀ pulse angle) and a
relaxation delay of 1.0 s was used. The FIDs were pro-
cessed using Gaussian multiplication with a Gaussian
broadening of 0.2 and ꢀ1.50 Hz line broadening. The
data were zero-filled to 128 K before Fourier transfor-
mation. The sample temperature was maintained at
30 ꢀC by means of a Bruker BVT 3000 digital variable
temperature unit.
4.10. Mass spectrometry
The mass spectrometric analyses were carried out on a
Waters Micromass Quattro Ultima triple quadrupole
mass spectrometer equipped with an electrospray ioniza-
tion interface. Samples were prepared in water, which
was also used as the blank. The blank and analytes were
injected by use of a pneumatically assisted Rheodyne
7010 injector equipped with a 10 lL injection loop at a
flow rate of 10 lL/min.
1
quency of the H resonance being selectively excited.
The spectra were acquired in 200 scans and the FIDs
were processed using exponential multiplication with a
0.20 Hz line broadening and were zero-filled to 128 K
of memory before Fourier transformation.
1
A selective 1D COSY H spectrum was obtained in
320 scans over an 8.09 kHz spectral width in 64 K data
points corresponding to a 4.05 s acquisition time. The
data were processed in the same manner as for the selec-
Acknowledgment
1
tive 1D TOCSY H spectra discussed above.
The 13C NMR spectrum was acquired over a
36.231 kHz spectral width in 53,100 scans in 32 K data
points with an acquisition time of 0.45 s and a data
point resolution of 1.11 Hz/point. The 13C pulse width
was 4 ls (30ꢀ pulse angle) and a relaxation delay of
0.5 s was employed. The FIDs were zero-filled to 64 K
prior to Fourier transformation and were processed
using exponential multiplication with a line broadening
of 4.0 Hz and linear back-prediction.
We thank Dr. Helene P. A. Mercier for her assistance in
´ `
preparing and critiquing this manuscript.
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