N1'-(p-[18F]fluorobenzyl)naltrindole (p-[18F]BNTI) as a potential PET imaging agent for DOP receptors

Article abstract of DOI:10.1002/jlcr.1095

The N1'-(p-fluorobenzyl)naltrindole 5 has been synthesized by reaction of 3-O-benzyl NTI 3 with p-fluorobenzylbromide under phase transfer catalysis. The subsequent 3-O-benzyldeprotection of 4 in HBr/CH₃COOH gave the target compound 5 in three

Full text of DOI:10.1002/jlcr.1095

JOURNAL OF LABELLED COMPOUNDS AND RADIOPHARMACEUTICALS
J Label Compd Radiopharm 2006; 49: 857–866.
Published online in Wiley InterScience (www.interscience.wiley.com). DOI: 10.1002/jlcr.1095
Research Article
N1’-(p-[18F]Fluorobenzyl)naltrindole (p-[18F]BNTI) as
a potential PET imaging agent for DOP receptors
Eyup Akgun1,2,3,*,y, Munawwar Sajjad1,2,z and Philip S. Portoghese3
¨
1 PET Imaging Service, VAMC, Minneapolis, MN 55417, USA
2 School of Medicine, Department of Radiology, University of Minnesota, Minneapolis,
MN 55455, USA
3 College of Pharmacy, Department of Medicinal Chemistry, University of Minnesota,
Minneapolis, MN 55455, USA
Summary
The N1’-(p-fluorobenzyl)naltrindole 5 has been synthesized by reaction of 3-O-benzyl
NTI 3 with p-fluorobenzylbromide under phase transfer catalysis. The subsequent 3-
O-benzyldeprotection of 4 in HBr/CH3COOH gave the target compound 5 in three
steps from naltrindole 2. p-FBNTI 5 is a novel delta opioid receptor antagonist
(Ki=0.00312 nM) and antagonizes the delta opioid (DOP) agonist, DPDPE, with a
Ke=1.55 nM in the mouse vas deferens preparation. Using the same synthetic strategy
the synthesis of p-[18F]BNTI 10 was undertaken. The final yield was 4% and the
specific activity varied in a range of 250–400 mCi/mmol. Copyright # 2006 John
Wiley & Sons, Ltd.
Received 27 April 2006; Revised 18 May 2006; Accepted 19 May 2006
Key Words: FBNTI; delta opioid receptors; fluorine-18; positron emission tomography
Introduction
Opioid receptors (ORs) belong to the superfamily of G protein-coupled
receptors (GPCRs). The existence of at least four types of opioid receptors
(MOP, KOP, DOP and NOP) is well established.1,2
*Correspondence to: Eyup Akgun, College of Pharmacy, Department of Medicinal Chemistry, University of
¨
Minnesota, Minneapolis, MN 55455, USA. E-mail: akgun001@umn.edu
yCurrent address: College of Pharmacy, Department of Medicinal Chemistry, University of Minnesota,
Minneapolis, MN 55455, USA.
zCurrent address: Department of Nuclear Medicine, The State University of New York at Buffalo, NY
14214, USA.
Contract/grant sponsor: DA01533
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E. AKGUN ET AL.
858
Pharmacological and radioligand binding studies have led to the proposal
for opioid receptors subtypes. However, the cloning of ORs has confirmed
only four main types that are coupled to inhibitory G proteins.3 The DOP
receptors are one of the four major opioid receptors that are present in the
central nervous system and in the periphery. In the central nervous system
DOP receptors are found in the neocortex, striatum, olfactory areas,
substantai nigra, amygdala and the nucleus accumbens.4 Activation of these
receptors results in many physiological and behavioral effects ranging, from
modulation of antinociception, mood, sensory system, motor integration and
cognative functions. In addition, delta opioid receptors are also present in
several immunocompetent cells, which suggest they may play an important
role in regulating the immune system.5
Pharmacological studies of DOP receptors have suggested the existence of at
least two subtypes (delta1 and delta2)6,7 which has not been substantiated by
cloning.8,9 These pharmacological results can be explained on the basis of two
possibilities. The first is that DOP receptors form heterodimers10–13 with other
receptors and these heterodimers have different phenotypical behaviors.14 The
second possibility is that the putative subtypes of delta opioid receptors
represent different affinity states of the same receptors due to dimerization and
or higher-order oligomerization.15
To address these issues, a full characterization of delta opioid receptors is
necessary. The in vivo distribution profiles can provide information about the
DOP receptors in general and their changes in pathological conditions in
particular. One useful tool for this is positron emission tomography (PET)
which is a non-invasive technique.16 PET utilizes specifically designed radio-
tracers with affinity and selectivity for target receptors. Ligand development is
the crucial part of PET studies. Therefore, we were interested in preparation a
selective DOP receptor ligand to aid the mapping delta opioid receptors in vivo.
As N1’-benzylnaltrindole (BNTI) 1 is known to be a delta2 selective DOP
receptors antagonist,17 we chose to synthesize p-fluorobenzylnaltrindole (p-
FBNTI) 5 based on its close structural similarity to BNTI 1, because p-FBNTI 5
was expected to have similar binding profile for DOP receptors.
We have prepared p-FBNTI18,19 5 and determined that it bound specifically
to delta opioid receptors. p-FBNTI 5 antagonized DPDPE with a
Ke=1.55 nM in the mouse vas deferens prep. The synthesis of corresponding
radiolabeled p-[18F]BNTI 10 was undertaken because the 18F (t1/2=110 min)
positron emitter should be good candidate for PET study of DOP receptors in
the central nervous system. The following report summarizes our results.
Results and discussion
The synthesis of p-fluorobenzylnaltrindole 5 started with protection of
naltrindole20 2 as 3-O-benzylnaltrindole 3 prior to fluorobenzylation. The
Copyright # 2006 John Wiley & Sons, Ltd.
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FBNTI AS A POTENTIAL PET IMAGING AGENT
859
N
N
N
OH
OH
O
OH
a
N
H
N
N
H
O
O
OH
OH
OR
3
2
BNTI 1
N
N
OH
OH
b
c
N
N
O
O
OH
OR
F
F
4
5
R=C6H5CH2
a) Acetone, K2CO3, benzylbromide ; b) CH2Cl2, tetrabutylammonium hydrogen sulfate,
NaOH (50%), p-fluorobenzyl bromide; c) i.CH3COOH/HBr(1:1), 90 °C, ii. NaHCO3
Scheme 1. Synthesis of N1’-(p-fluorobenzyl)naltrindole p-FBNTI 5
synthetic route involved the transfer of p-fluorobenzyl bromide under phase
transfer catalysis to the indole nitrogen of NTI 2. The reaction was conducted
in the presence of a catalytic amount of tetrabutylammonium hydrogen sulfate
(TBAHS) and NaOH (50%) in methylene chloride,21 affording [3-O-benzyl,
N1’-(fluorobenzyl)]NTI 4 in a yield of 95% which was characterized clearly by
1H and 13C-NMR analyzes. Elemental analysis of compound and mass spectra
confirmed unequivocally its constitution. Using HBr in acetic acid the 3-O-
benzyl protective group was removed selectively. This deprotection method
was very effective and the reaction was complete in 10 min, affording
p-fluorobenzyl NTI 5 in over 90% yield. The structure of the product was
assigned with the aid of NMR spectroscopic data. High-resolution mass and
elemental analysis also confirmed clearly the identity of the compound
(Scheme 1).
In its biological evaluation, the in vitro characteristics of p-FBNTI 5 showed
high affinity for delta opioid receptors (Ki=0.00312 nM). Weak binding was
observed for the m and k compounds [3H]DAMGO (Ki=87.4 ꢀ 33.8 nM) and
[3H]U69593 (Ki>1000 nMl ).18,19 The binding assay (Inhibition of 3H-
Diprenorphine in HEK 293 cells) showed the following selectivity ratios of
m=d ¼ 13814 and k=d ¼ 41987 (see Figure 1).
The key compound p-[18F]fluorobenzaldehyde 7 was prepared in a yield of
58.8 ꢀ 11.6% by reaction of 18Fꢁ with p-trifluoromethylammonium-benzal-
dehyde triflate 6.22–25 This starting material was prepared according to a
known procedure.22 Reduction of 7 with LiBH4 and subsequent treatment
Copyright # 2006 John Wiley & Sons, Ltd.
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860
100
80
60
40
20
0
MOP
KOP
DOP
10-13 10-12 10-11 10-10 10-9 10-8 10-7 10-6
Concentration (M)
Figure 1. Binding of 5 to opioid receptor-transfected HEK 293 cells
with HBr (48%) gave p-[18F]fluoro benzyl bromide 8 in a yield of 68.5%.
Compound 8 was transferred via N2-flow in pentane into a flask containing
phase transfer catalyst tetrabutylammonium hydrogen sulfate (TBAHS),
KOH and (3-O-benzyl NTI) 3 in methylene chloride/toluene (1:1). This
reaction gave the intermediate 9, which on treatment with HBr/CH3COOH
provided the final product 10. The yield was 4%, and the specific activity
varied between 250 and 400 mCi/mmol decay corrected to EOB. The average
time for the radiochemical synthesis was 3.5 h from the end of bombardment.
Because of its very high DOP selectivity, p-[18F]BNTI 10 complements existing
PET imaging agents such as N1’-([11C]methyl)naltrindole20 and N1’-
([18F]fluoroethyl)naltrindole.26 Application of p-[18F]BNTI 10 in vivo studies
will help to image DOP receptors in human brain and their changes as a
consequence of diseases. Imaging DOP receptors and their distribution in vivo
using positron emission tomography will be very helpful to understand
interaction of DOP receptors with other GPCRs in general, and with MOP
and KOP receptors in particular.27 In this regard, a bivalent opioid ligand
labeled with positron emitter should visualize the opioid heterodimers and
their changes in vivo with neuronal cross-reactivity.
Experimental
Solvents and chemicals were purified prior to use. Melting points were
determined with Mel-Temp Laboratory Devices and uncorrected. IR spectra
were recorded with a Perkin-Elmer 1600 FTIR. 1H NMR (300.007 MHz) and
13C NMR (75.462 MHz) were obtained with a Varian Gemini-300 instrument.
Chemical shifts are reported in ppm (d) relative to internal Me4Si in CDCl3.
High-resolution fast bombardment mass spectroscopy (HRFABMS) was done
at the University of Minnesota Mass Spectroscopy Facility. Elemental analysis
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FBNTI AS A POTENTIAL PET IMAGING AGENT
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was determined by M-H-W Laboratory in Phoenix, Arizona. Analytical TLC
was done on Baker-flex, silica gel IB2-F plates. The analytical HPLC
equipment consisted of Waters 600E pumps, Waters 490E UV absorbance
detector (254 nm), a NaI (TI) crystal (2’’) Beckman 170 Radioisotope detector
and Hewlett–Packard 3396A integrators. Preparative HPLC consisted of
spectral series UV 100 in series (254 nm) with NaI detector for monitoring
radioactivity. Analytical (Waters Bondapak C-18, 3.9 ꢂ 300 mm) and semi
preparative (Alltech, Adsorbospere C-18, HS 7U, 10 ꢂ 300 mm) reverse-phase
HPLC columns were used. A Hewlett–Packard 3396A integrator recorded the
HPLC chromatograms. Radioactivity was measured with a Capintec radio-
isotope dose calibrator. NTI 1 was prepared from naltrexone as described in
the literature.28,29
(Cyclopropylmethyl)-6,7-dehydro-4,5-epoxy-3-benzyloxy-14-hydroxy-6,7-2’,3’-
indolomorphinan (3-O-benzyl NTI) 3
A solution of NTI 2 (1.1 g, 2.45 mmol) in dry acetone (25 ml) containing
K2CO3 (10 equivalent) and benzyl bromide (461 mg, 2.7 mmol, 1.1 eq.) was
refluxed for 5 h, and then the mixture was cooled to room temperature and
filtered. The mother liquor was portioned between water and methylene
chloride where the compound was extracted with methylene chloride
(3 ꢂ 50 ml) and dried over anhydrous Na2SO4. The solvent was evaporated
and the compound was purified by gravity column chromatography [SiO2,
CH2Cl2/MeOH (20:1)] to give the final product as a white solid.
Yield: 1.27 g (85%); mp>1158C decomposed.; IR (Film, CHCl3) u 3500–
3100 (w), 3050 (w), 2950 (s), 2850 (m), 1505 (s), 1438 (m) cmꢁ1 1H-
;
NMR(CDCl3) d 8.1 (s, 1 H, NH), 7.42 (d, J=7.74 Hz, 1 H, Ar), 7.7–7.4 (m,
6 H, Ar), 7.3 (t, J=7.0 Hz, 1 H, Ar), 7.05 (t, J=7.86 Hz, 1 H, Ar), 6.6 (AB,
J
AB=8.25 Hz, 2 H, Ar), 5.68 (s, 1 H, H5), 5.05 (AB, JAB=11.9 Hz, 2 H,
benzylic protons), 3.49–2.2 (m, 9 H, aliphatic protons), 1.77 (dd, J=3.99 Hz,
J=2.0 Hz, 2 H, aliphatic protons), 1.3 (s, 1 H, OH), 0.9 (m, 1 H), 0.6
(d, 8.19 Hz, cyclopropyl protons), 0.2 (d, J=4.68 Hz, 2 H, cyclopropyl
protons).; 13C-NMR (CDCl3) d 144.1, 141.2, 136.9, 136.7, 131.2, 129.3,
128.1, 127.9, 127.7, 126.6, 126.1, 121.8, 118.8, 118.3, 115.3, 111.3, 110, 84.5,
72.1, 70.2, 61.6, 52.6, 47.2, 43.4, 31.1, 31, 22.8, 22.1, 9.3, 3.9, 3.6;
(FABHRMS): 505.2491, C33H33N2O3 requires (M+H)+=505.2491. Analy-
tically calculated for C33H33N2O3 C, 78.54: H, 6.39: N, 5.55. Found C, 78.65:
H, 6.53: N, 5.29.
(Cyclopropylmethyl)-6,7-dehydro-4,5-epoxy-3-benzyloxy-14-hydroxy-6,7,2’,3’-
[1’-(p-fluorobenzyl)]-indolomorphinan [3-O-benzyl, N1’-(p-fluorobenzyl)]NTI 4
A solution of 3-O-benzyl NTI 3 (148 mg, 0.29 mmol) in methylene chloride
(15 ml) containing tetrabutylammonium hydrogen sulfate (20 mg) was mixed
Copyright # 2006 John Wiley & Sons, Ltd.
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with NaOH (50%, 15 ml). To this reaction mixture was added p-fluorobenzyl
bromide (60.8 mg, 1.1 eq.) dissolved in methylene chloride (5 ml) and this
mixture was stirred for 2 h at room temperature. The reaction mixture was
diluted with ice-water and the organic layer was removed. The aqueous layer
was extracted with methylene chloride (2 ꢂ 25 ml) and the combined extracts
were dried over anhydrous sodium sulfate. After filtration and evaporation of
the solvent, the residual compound was purified by flash chromatography
[SiO2, CH2Cl2/MeOH (20:1)]. A white powder was obtained which was
crystallized from CH2Cl2/pentane.
Yield: 132 mg (74.5%); mp>948C decomposed.; IR (Film, CHCl3) u 3450–
3400(br, OH), 3060 (w), 2900 (s), 2850 (m), 1625 (m), 1587 (m), 1500 (s) cmꢁ1
;
1H-NMR(CDCl3) d 7.48 (d, J=7.41 Hz, 1 H, Ar), 7.4–7.0 (m, 11 H, Ar), 6.48
(t, J=8.5 Hz, 1 H, Ar), 6.5 (AB, JAB=8.25 Hz, 2 H, Ar), 5.72 (s, 1 H, H5), 5.6
(AB, JAB=16.5 Hz, 2 H, benzylic protons), 4.98 (AB, JAB=4.95 Hz, 2 H,
benzylic protons), 3.49–2.2 (m, 9 H, aliphatic protons), 1.85 (dd, J=3.99 Hz,
J=2.0 Hz, 2 H, aliphatic protons), 1.3 (s, 1 H, OH), 0.95 (m, 1 H), 0.6
(d, J=7.6 Hz, cyclopropyl protons), 0.2 (d, J=4.17 Hz, 2 H, cyclopropyl
protons); 13C-NMR (CDCl3) d 142, 138, 137.9, 132(JCꢁ1,F=236 Hz), 129.3,
127.3 (3JCꢁ3,F=9 Hz), 126.8, 126.5, 125.7, 125.4, 121.6, 118.2, 118.1, 117.7,
115.5, 114.5, 114.5(d, 2JCꢁ2,F=22.5 Hz), 110.1, 109, 83.4, 71.6, 70.6, 61.3, 58.6,
47.2, 45.9, 42.7, 30.8, 28.2, 22.3, 8.6, 3.2, 3.0; (FABHRMS): 613.2880,
C40H37N2O3F requires (M+H)+=613.2854. Analytically calculated for
C40H37N2O3F C, 78.40: H, 6.08: N, 4.57. Found C, 78.32: H, 6.19: N, 4.56.
N1’-(p-fluoromethyl)-17-(cyclopropylmethyl)-6,7-didehydro-4,5-epoxy-3,14-di-
hydroxyindolo[2’,3’,6,7]morphinan (FBNTI).HBr.1/2H2O 5
A solution of 4 (735 mg, 1.2 mmol) was dissolved in acetic acid/ HBr (5 ml,
1:1). The entire mixture was heated to 908C within 2 h. Subsequently, this
mixture was poured into saturated NaHCO3 solution. The resulting gum-like
material was washed with water until the filtrate was neutral. The solid
material then was dissolved in ethyl acetate (5 ml) and allowed to precipitate as
a white powder. The powder was washed with cold ethyl acetate and ether.
The HPLC analysis {MeOH/(NH4)2PO4 [85:15], flow rate 1 ml/min,
tR=9.1 min} showed the product analytically pure.
Yield: 535 mg (85.3%); mp>1808C decomposed.; IR (Film, CHCl3) u 3500–
3050(br, OH), 2950 (m), 2900 (m), 2850 (m), 1600 (m), 1550 (m), 1500 (s),
1
1495 (s) cmꢁ1; H-NMR(CDCl3) d 9.3 (s(br), 1H, OH), 7.49 (d, J=7.62 Hz,
1H, Ar), 7.15–6.9 (m, 7H, Ar), 6.6 (AB, JAB=8.25 Hz, 2H, Ar), 5.70 (s, 1H,
H5), 5.4 (AB, JAB=16.7 Hz, 2H, benzylic protons), 4.98, 3.8–2.2 (m, 9H,
aliphatic protons), 1.77 (dd, J=3.99 Hz, J=2.0 Hz, 2H, aliphatic protons), 1.3
(s, 1H, OH), 0.9 (m, 1H), 0.58 (d, J=7.6 Hz, cyclopropyl protons), 0.2 (d,
J=5.1 Hz, 2H, cyclopropyl protons); 13C-NMR (CDCl3) d 142.5, 138.7, 137,
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FBNTI AS A POTENTIAL PET IMAGING AGENT
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132(JCꢁ1,F=236 Hz), 129.9, 127.6 (3JCꢁ3,F=8.2 Hz), 126.6, 125.4, 122.8,
2
119.4, 119.2, 119.1, 116.7, 115.4(d, JCꢁ2,F=22.5 Hz), 111.4, 109.7, 85, 72.6,
62.3, 59.5, 46.5, 43.8, 45.9, 31.5, 29.1, 23.2, 9.5, 4.2, 3.9, 1.82; (FABHRMS):
523.2384, C33H31N2O3F requires (M+H)+=523.2389. Analytically calcu-
lated for C33H32N2O3BrF.1/2H2O C, 64.71: H, 5.43: N, 4.61. Found C, 64.71:
H, 5.52: N, 4.35.
Receptor binding assays
The modified receptor binding assays30 were applied as follows: Competition
binding experiments were performed using HEK 293 cells genetically modified
to produce wild-type MOP, KOP, or DOP receptors. Ten concentrations of
the tested compounds (50 mL) were added to test tubes containing 0.1 nM 3H-
diprenorphine ( ꢃ 0.5–1.0 ꢂ KD) (50 mL) and whole cells (75 mm2 plate, 80–
90% confluent) suspended in 12.5 ml HEPES buffer (25 mM, pH=7.4)
(400 mL). Final volume was 500 mL. Non-specific binding was measured using
10 mM naloxone. Assays were incubated at room temperature for 90 min and
then filtered using a Brandel M-48 tissue harvester through Whatman GF/C
filter paper that was pre-soaked in 0.25% poly(ethyleniminie). Filters were
washed three times with ice-cold HEPES buffer (see above), and the
radioactivity counted using a Beckmann LS 6500 liquid scintillation counter.
All measurements were performed in triplicate. IC50 values were calculated
using PRISM software utilizing non-linear regression of the data normalized
to fit a sigmoidal dose–response curve with a variable slope (100% defined at
concentration=0 (total binding) and 0% defined at the value of non-specific
binding). Ki values were determined from the Cheng-Prusoff 31 equation
assuming a single-site-binding model. Values reported are mean IC50 and
Ki ꢀ standard error of the mean (SEM) of three or more independent
experiments.
Radiolabeling procedure
In preparation for the synthesis of N1’-p-([18F]fluorobenzyl naltrindole 10, the
synthesis of p-[18F]fluorobenzyl bromide 8 was undertaken as depicted in
Scheme 2. Potassium [18F]fluoride was obtained by bombarding [18O]H2O
(Isotec, 1.2 mL, 98% isotopic abundance) with 15 MeV protons and
transferred under helium pressure to hot cell through a polyethylene tubing
(1/16’’ i.d.) in a Reacti-vial containing ꢄ5 mg potassium carbonate (99.995%
pure). This solution was concentrated to 0.1 ml. KrytofixR [2.2.2] (ꢄ5–7 mg) in
acetonitrile (1 ml) was added and the mixture transferred under helium
pressure to a vessel purged with argon gas. The potassium [18F]fluoride was
dried via azeotropic removal of water with acetonitrile (3 ꢂ 0.5 ml) at 1208C.
The precursor, p-trifluoromethylammonium-benzaldehyde triflate 6, dissolved
in 0.3 ml DMSO was added to the reaction vessel containing the [18F]fluoride,
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CHO
CHO
CH Br
2
a
b,c
d
18
18
N(CH ) TfO
F
3 4
F
6
7
8
N
N
OH
OH
e
N
N
O
O
OH
OR
18
18
F
F
9
10
R=C H CH
6
5
2
18
a) F-/K222/K CO ; b) THF, LiBH ; c)HBr(48%); d) CH Cl , tetrabutylammonium hydrogen sulfate,
2
3
4
2
2
°
NaOH (50%), 3-O-benzylnaltrindole 3 ; e) i.CH COOH/HBr(1:1), 90 C, ii. NaHCO
3
3
Scheme 2. Synthesis of N1’-(p-[18F]fluorobenzyl)naltrindole p-[18F]BNTI 10
and the mixture was heated at 858C for 5 min. After cooling for 2 min at room
temperature, the mixture was quenched with 5 ml water and passed through a
C-18 Sep-Pak (Waters, Milford, MA) which was sequentially washed with 5 ml
0.01 M HCl and water. The radioactive p-[18F]fluorobenzaldehyde 7 was
eluted with pentane and passed through an anhydrous Na2SO4 column into a
flask containing 0.3 ml LiBH4 (2 M in THF). This mixture was stirred for
3 min at room temperature. Finally, the solvent was evaporated at 908C. The
residual material was cooled with methanol/ice and treated with HBr(48%,
2 ml) and then heated at 908C for 10 min. The resulting mixture was cooled
with ice-water for 3 min, diluted with water, and then passed through a C-18
Sep-Pak (Waters, Milford, MA) which was rinsed with a solution of NaHCO3
(3 ml, 1 M), and the radioactive p-[18F] fluorobenzyl bromide 8 then was eluted
with pentane (3 ml), and the eluent passed through a anhydrous Na2SO4/
K2CO3 column into a flask containing TBAHS (5 mg), KOH (100 ml, 30 M) in
methylene chloride/toluene (1:1, 4 ml). To this mixture (3-O-benzyl NTI) 3 was
added and the entire mixture was heated at 408C until the pentane was
evaporated. Then this mixture was heated at 908C for 15 min. On cooling,
water was added to the reaction mixture, and the product was extracted with
methylene chloride and transferred to a round bottom flask. Most of
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FBNTI AS A POTENTIAL PET IMAGING AGENT
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methylene chloride was evaporated, and then a mixture of acetic acid and HBr
(48%) [1:1, 2 ml] was added to the flask and heated at 908C for 15 min. This
mixture then was quenched with saturated sodium bicarbonate solution and
the product was extracted with ether.
The final purification of p-[18F]BNTI 10 was achieved via a semi preparative
reverse-phase HPLC column using acetonitrile/(NH4)2HPO4 (4 mM) in a
mixture of 40:60 (flow rate 5 ml/min) for 15 min then the solvent mixture
changed to 60:40. The fraction containing p-[18F]BNTI 10 (tR=32 min) was
collected and evaporated to dryness. The residual was dissolved in saline
containing 10% ethanol and transferred to a sterile, pyrogen-free bottle. The
retention time of the p-[18F]BNTI 10 was 9.2 min using methanol/dibasic
ammonium phosphate [85:15] with a flow rate 1 ml/min. Radiochemical purity
and specific activity were determined by analytical HPLC. Specific activity was
calculated by relating the area of the UV absorbance peak of carrier p-FBNTI
5 in an aliquot of p-[18F]BNTI 10 of known radioactivity to the peak area of a
standard sample of p-FBNTI. p-[18F]BNTI 10 of >99% radiochemical purity
was obtained at the end of synthesis with the specific activity varied in a range
of 250–400 mCi/mmol, and radiochemical yield of 4% based upon initial [18F]-
fluoride activity.
Acknowledgements
We thank Mike Powers for the binding experiments. This was supported by
research grant DA01533. We also thank Mr Rashid Syed for assistance with
cyclotron operation.
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