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Full text of DOI:10.1002/jlcr.706

JOURNAL OF LABELLED COMPOUNDS AND RADIOPHARMACEUTICALS
J Label Compd Radiopharm 2003; 46: 661–668.
Published online in Wiley InterScience (www.interscience.wiley.com). DOI: 10.1002/jlcr.706
Research Article
Automated commercial synthesis system for
1
8
preparation of O-(2-[ F]fluoroethyl)-L-tyrosine
by direct nucleophilic displacement on a resin
column
1
,
2
1
Ganghua Tang *, Xiaolan Tang , Mingfang Wang , Lei Luo ,
1
1
Manquan Gan and Zuhan Huang
1
1
Nan Fang PET Centre, Nan Fang Hospital, The First Military Medical
University, Guangzhou 510515, China
2
Department of Applied Chemistry, South China Agricultural University,
Guangzhou 510642 , China
Summary
A slightly modified automated commercial synthesis system for preparation of
8
1
O-(2-[ F]fluoroethyl)-l-tyrosine (FET), an amino acid tracer for tumor
imaging with positron emission tomography, is described. Direct nucleophilic
1
fluorination of [ F]fluoride with 1,2-di(4-methylphenylsulfonyloxy)ethane
8
on a quaternary 4-(4-methylpiperidinyl)-pyridinium functionalized polystyrene
1
anion exchange resin gave 1-[ F]-2-(4-methylphenylsulfonyloxy)ethane,
8
1
8
then [ F]fluoroalkylation of l-tyrosine yielded FET. The overall
radiochemical yield with no decay correction was about 8–10%, the whole
synthesis time was about 52 min, and the radiochemical purity was above
9
5%. Copyright # 2003 John Wiley & Sons, Ltd.
1
8
Key Words: O-(2-[ F]fluoroethyl)-l-tyrosine; quaternary 4-aminopyridinium
resin; automated synthesis
*Correspondence to: G. Tang, Nang Fang PET Centre, Nan Fang Hospital, The First Military
Medical University, Guangzhou 510515, China. E-mail: tghtgh@china.com
Contract/grant sponsor: Presidents’ Foundation of Nan Fang Hospital.
Copyright # 2003 John Wiley & Sons, Ltd.
Received 26 August 2002
Revised 14 February 2003
Accepted 15 February 2003

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G. TANG ET AL.
Introduction
1
-[ F]Fluoro-2-deoxy-D-glucose (FDG) as a tracer of glucose meta-
8
2
bolism has been widely used to the diagnosis and treatment evaluation
1
for oncology with positron emission tomography (PET). However,
clinical FDG PET studies demonstrated several limitations such as
substantial uptake of FDG in tumor, human and experimental
inflammatory lesions. Uptake of FDG in acute inflammatory reactions
2
gives rise to false positive results in a considerable percentage of cases.
1
8
O-(2-[ F]fluoroethyl)-l-tyrosine (FET), a recently developed amino
acid tracer, has shown high uptake in experimental tumors and in
3–5
patients with brain tumors. In addition, almost no uptakes of FET in
activated inflammatory cells with experimental acute abscess model and
2
,6
in inflammation within lymph nodes are found. The routine synthesis
of FET with high radiochemical yield has been reported, however, the
3
separation and purification of FET used HPLC, adding difficulty to
performing a fully automated synthesis of FET.
We report here that the PETtrace FDG MicroLab can be applied to
the fully automated synthesis of FET.
Results and discussion
Chemistry
The synthetic procedure designing for the preparation of O-(2-
19
1
9
[
F]fluoroethyl)-l-tyrosine ( FET) is based on the selective O-
7
alkylation of tyrosine. Dimethylsulfoxide (DMSO) was particularly
effective in promoting base-catalyzed condensation and a lot of alkyl
ethers of l-tyrosine were successfully synthesized from O-alkylation of
7
tyrosine in this solvent without protecting the amino group. Under the
conditions of l-tyrosine dissolved in a mixture of DMSO and aqueous
sodium hydroxide and treated with the appropriate alkyl halide at
moderate temperature, no significant N-alkylation was detected. The
Copyright # 2003 John Wiley & Sons, Ltd.
J Label Compd Radiopharm 2003; 46: 661–668

AUTOMATED COMMERCIAL SYNTHESIS SYSTEM
663
ether ester, major by-product resulting from dialkylation, could be
avoided by carrying out the reaction with a slight excess of the amino
acid or eliminated by saponification with dilute base before isolation of
1
9
the product. For this purpose the high yield of FET was obtained
from the reaction of l-tyrosine dissolved in the solution of 2 equivalent
of 10% sodium hydroxide and about 3–10 volume of DMSO at
ae
19
1
7
0–90 i . The structure of FET is confirmed by MS and H NMR.
Radiochemistry
Synthesis of FET via a two-step procedure as shown in Figure 1 was
similar to that of FDG. First, direct nucleophilic substitution of
1
8
[
F]fluoride with 1,2-di(4-methylphenylsulfonyloxy)ethane (1) on a
quaternary 4-(4-methylpiperidinyl)- pyridinium functionalized polystyr-
18
ene anion exchange resin gave 1-[ F]-2-(4-methylphenylsulfonylox-
18
y)ethane (2). Second, [ F]fluoroethylation of l-tyrosine with (2) and
saponification with dilute sodium hydroxide solution yielded FET (3).
8
Hence, commercial PETtrace FDG MicroLab with only minor
modifications could be used for the fully automated preparation of
FET. The typical chromatograms of FET were consistent with those of
19
the standard FET by HPLC analysis (FET retention time R =
t
4
.3 min) and TLC analysis (FET R =0.22). After decay of final product
f
(
FET injection), product characterization was also confirmed by mass
+
spectrum, MS (EI) m/z: 226 (M ), furthermore, only little l-tyrosine
and no DMSO were detected by HPLC and GC. FET (3) was prepared
in the overall radiochemical yield of 8–10% with no decay correction
and in the radiochemical purity of more than 95% within 52 min
18
synthesis time. No O-(2-[ F]fluoroethyl)-d-tyrosine was found by
enantiomeric analysis.
Figure 1. Synthesis of FET by direct nucleophilic exchange on an anion exchange
column
Copyright # 2003 John Wiley & Sons, Ltd.
J Label Compd Radiopharm 2003; 46: 661–668

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G. TANG ET AL.
The growing importance of FET as a positron-emitting radio-
pharmaceutical for amino-acid metabolic study makes it necessary to
synthesize this compound using a simple method. This work indicates
that the fully automated synthesis of FET on commercial PETtrace
FDG MicroLab can be prepared routinely for clinical use though
radiochemical yield is low. Also, the fully automated synthesis by direct
nucleophilic exchange on a quaternary 4-aminopyridinium resin has the
following advantages: Firstly, the Sep-Pak cartridges are used to purify
and separate the radiolabeled intermediate product and the last product
3
instead of HPLC system, which results in simplification of the
operation steps and shortening of the whole synthesis time. Secondly,
due to the small overall size of the resin column and heating block, the
1
8
assembly fits easily into a shielded hot cell. The trapping of [ F]fluoride
ion, nucleophilic radiolabeling reagents transport and all radioactive
solution transfers done by means of Teflon tubing, helium pressure,
remote syringes, or vaccum are performed in shielded state, with no
operator handling necessary except to remove the final product vial.
These factors keep the radiation exposure to personnel at a minimum.
Thirdly, the amount of training required and effort involved in
8
,9
operating this system also are minimal.
1
Fourthly, nucleophilic
8
[
F]fluorination procedure on a resin column eliminates some problems
1
which have arisen in other reactions with resolubilized [ F]fluoride ion
8
+
3,10
using counterions such as Kryptofix 222/K
1
and tetraalkylammo-
18
as phase transfer reagents. Loss of [ F]fluoride on
1,12
nium salts
reaction vessel walls need not be considered. Furthermore, the use of a
resin column eliminates the need to remove phase transfer reagents with
9
toxic chemicals such as Kryptofix 222 and tetraalkylammonium salts.
Experimental
Isotope availability
18
No-carrier-added aqueous [ F]fluoride ion was produced on a
PETtrace cyclotron (GE Co.) by irradiation of a 1.5 ml water target
1
8
using a 16.5 MeV proton beam on 95% enriched [ O]water by the
1
8
18
O(p, n) F nuclear reaction and was transferred to the synthesis
1
module. Typical production: 400–500 mCi (14.8–13.5 GBq) of [ F]F
8
À
8
at the end of bombardment for a 25 mA, 30 min irradiation.
Copyright # 2003 John Wiley & Sons, Ltd.
J Label Compd Radiopharm 2003; 46: 661–668

AUTOMATED COMMERCIAL SYNTHESIS SYSTEM
665
Experimental apparatus
The fully automated synthesis of FET was performed in a lead-shielded
8
hot cell using a modified PETtrace FDG MicroLab (GE Co.). The
PETtrace FDG MicroLab is a computer controlled automated radio-
1
8
À
chemistry system which will produce FDG from [ F]F . All chemical
transformation and FDG production take part within a disposable
process Cassette. One Cassette is required for each batch production
run of FDG. The Cassette is able to be used for the synthesis of FET.
1
9
19
Synthesis of O-(2-[ F]fluoroethyl)-l-tyrosine ( FET)
1
9
19
O-(2-[ F]fluoroethyl)-l-tyrosine ( FET) was prepared according to the
7
method described by Solar et al. A solution of 0.646 g (3.5 mmol) l-
tyrosine in 2.800 g (7.0 mmol) 10% aqueous sodium hydroxide was
added to 14 ml of dimethyl sulfoxide and heated in a oil bath to 808C.
To this was added, with stirring, 0.450 g (3.5 mmol) 1-fluoro-2-
bromoethylane. Heating and stirring were continued for 2 h and the
reaction mixture was then poured into 18 g of crushed ice. The pH was
adjusted to about 7.5 and the resulting precipitate was filtered off,
washed with water and dried. The crude product was recrystallized from
1
0% acetic acid to give 0.270 g (1.2 mmol, 34%) of cold standard. H
6
NMR (CDCl ) 6.7–7.3 (m, 4 H), 4.7 (t, 2 H), 4.3 (t, 2 H), 3.4 (m, 1 H),
3
+
.8 (m, 2 H) MS (EI) m/z: 226 (M )
2
18
Automated production of O-(2-[ F]fluoroethyl)-l-tyrosine (FET)
The fully automated radiochemical synthesis of FET was modified from
the PETtrace FDG MicroLab by a two-step reaction sequence that
18
consisted of F-fluorination of 1,2-ditosyloxyethane and subsequent
8
1
F-fluoroethylation of unprotected l-tyrosine as follows. The gener-
1
8
ated F-fluoride target water was transferred to the target water
reservoir on the FDG Cassette, then transferred to one of the syringes
and passed through the solid-phase extraction column where the
[
18F]FÀ was trapped. The solid-phase consists of a 2%-crosslinked
polystyrene matrix (Merrifields resin, approximately 1.5 meq of active
group/g) where the trapping agent is 4-(4-methylpiperidinyl)pyridinium
cation. The resin is placed in the column, which was included in the
FDG Cassette. The used target water was recovered via a separate
18
À
collecting vial. The resin-bound [ F]F was dried by passing
Copyright # 2003 John Wiley & Sons, Ltd.
J Label Compd Radiopharm 2003; 46: 661–668

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G. TANG ET AL.
acetonitrile as the mobile phase instead of water at about 1008C and the
target water receiver in the Cassette was rinsed. 1,2-Ditosyloxyethane
(10 mg, 0.027 mmol) dissolved in dry acetonitrile (2 ml) was passed
through the heated solid-phase extraction column and was reacted with
À
18
18
the trapped [ F]F on the column. 1-[ F]-2-tosyloxyethane was
18
formed and was transferred to the [ F]fluoroethylation vessel. Then,
the acetonitrile was evaporated. The mixture of l-tyrosine (9.1 mg,
5
0 mol), 100 g/l sodium hydroxide solution (4 mg/40 l, 100 mol) and
DMSO (0.5 ml) was added and the reaction mixture was heated at
1
8
1
008C for 20 to form FET. [ F]Fluoroethylation was halted by
stopping the heating and adding 0.15 mol/l phosphate-buffered saline
PBS, pH 7.40) (2 ml). The final product was transferred to the product
(
syringe and further through a silica Sep-Pak cartridge. FET was eluted
from the serial silica Sep-Pak cartridge, C-18 Sep-Pak cartridge and
Al O Sep-Pak cartridge with PBS (pH 0.74). Eluate was passed
2
3
through 0.22 m sterile filtration to the product vial in a separate
radiation shielded container.
18
Analysis of O-(2- F-fluoroethyl)-l-tyrosine (FET)
The radiochemical purity was analysed by HPLC on an analytical
reverse phase column (RP-18, 150 Â 4.6 mm) eluted with ethanol/water/
acetic acid (10/87.5/2.5, v/v/v) and 2.5 g/l ammonium acetate (pH 3.0) at
a flow of 1 ml/min. The effluent was monitored for UV absorbance and
radioactivity (FET retention time Rf: 4.3 min). Thin-layer chromato-
graphy was performed with acetonitrile/water (95/5, v/v) as the solvent
system (FET R =0.22). The chemical purity was determined by HPLC,
f
TLC and gas chromatography (GC). Enantiomeric analysis was carried
out with HPLC using a reversed-phase C18 column and a chiral mobile
1
3
phase.
Conclusion
1
8
A fully automated preparation of O-(2-[ F]fluoroethyl)-l-tyrosine
FET) is performed in a lead-shielded hot cell using a modified
(
1
8
commercial PETtrace FDG MicroLab. Collection of [ F]fluoride ion
from target water, drying and nucleophilic fluorination reaction of
1
8
[
F]fluoride with 1,2-di(4-methylphenylsulfonyloxy)ethane are all
rapidly completed on a quaternary 4-(4-methylpiperidinyl)pyridinium
Copyright # 2003 John Wiley & Sons, Ltd.
J Label Compd Radiopharm 2003; 46: 661–668

AUTOMATED COMMERCIAL SYNTHESIS SYSTEM
667
1
functionalized polystyrene anion exchange resin column to give 2-[ F]-
8
1
8
1
-(4-methylphenylsulfonyloxy)ethane. Then, [ F]fluoroalkylation of l-
18
tyrosine with 1-[ F]-2-(4-methylphenylsulfonyloxy) ethane forms FET.
The synthesis of FET is performed in the whole synthesis time of about
5
2 min with overall radiochemical yield about 8–10% (no-decay
correction) and radiochemical purity above 95%.
Acknowledgements
This work was supported by the President’s Foundation of Nan Fang
Hospital, First Military Medical University.
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