Synthesis and Radiolabelling of Ipratropium and Tiotropium for Use as PET Ligands
57
tropenol 7 was taken onto the next step without further purification. The
1H NMR and IR spectra were consistent with the data reported in the
literature.[20]
57.8, 56.1, 42.5, 31.0 (one signal overlapping). m/z (ESI+) 378 (M + H,
100%). (HRMS (ESI+) calc. 378.0834. C18H20NO4S2 (M + H) found
378.0824.)
2-Hydroxy-2,2-di(thiophen-2ꢀ-yl)acetic Acid Methyl Ester 6[22]
(1R,2R,4S,5S,7s)-7-[2ꢀ-Hydroxy-2ꢀ,2ꢀ-di(thiophen-2ꢀꢀ-yl)
acetoxy]-9,9-dimethyl-9-aza-3-oxatricyclo[3.3.1.02,4
nonan-9-ium 2 Trifluoroacetate (Tiotropium 2
Trifluoroacetate)
]
Ester 6 was synthesized according to the literature with only minor
modifications.[21,22] Purification of the crude reaction mixture by flash
chromatography (EtOAc/hexane 1/9) gave methyl ester 6[22] (75%) as
white crystalline plates that darkened on standing. Rf (EtOAc/hexane
1/9) 0.21. ν (thin film)/cm−1 3477 (OH), 3105, 2960, 2924, 2849 (C–H),
1722 (C=O ester) cm−1. δH (200 MHz, CDCl3) 7.29 (2H, dd, J 5.1 &
A solution of epoxide 13 (27 mg, 0.0716 mmol, 1.0 equiv.) in
anhydrous DMF (0.9 mL) was treated with methyl iodide (44.6 µL,
0.716 mmol, 10 equiv.) dropwise at room temperature. The reaction
mixture was heated at 57◦C for 4.5 h. After cooling to room tem-
perature, milliQ water (5 mL) was added. The aqueous phase was
washed with ethyl acetate and then lyophilized to dryness. Purification
by reverse-phase HPLC (Alltech Altima C18 10 µm, 22 × 300 mm2,
ꢀ
ꢀ
1.2, C5 -H), 7.16 (2H, dd, J 3.6 & 1.3, C3 -H), 6.98 (2H, dd, J 5.1 &
ꢀ
3.7, C4 -H), 4.68 (1H, s, OH), 3.90 (3H, s, OCH3). δC (75 MHz, CDCl3)
172.8, 145.6, 126.8, 126.0, 125.9, 76.5, 54.1. m/z (EI+) 237 (M-OH,
27%), 195 (M-C2H3O2, 48), 111 (C5H3OS, 100). A second fraction
afforded di(thiophen-2-yl)methanone 11,[23] for which the 1H NMR and
IR spectra were consistent with the data reported in the literature.[23]
acetonitrile/water 40/60 with 0.05% trifluoroacetic acid, 7 mL min−1
)
afforded tiotropium 2 trifluoroacetate (27.2 mg, 97%) as a white solid.
Rt (Spherisorb S10 SCX 10 µm, 10 × 250 mm2, acetonitrile/0.1 M
NaH2PO4 40/60, 5 mL min−1) 9.10 min. ν (KBr plate)/cm−1 3427
(OH), 1742 (C=O ester), 1686 (COO−). δH (200 MHz, MeOD) 7.43
(1R,3s,5R)-2ꢀ-Hydroxy-2ꢀ,2ꢀ-di(thiophen-2ꢀꢀ-yl)acetic Acid
8-Methyl-8-azabicyclo[3.2.1]oct-6-en-3-yl Ester 12[24]
ꢀꢀ
ꢀꢀ
(2H, dd, J 5.1 & 1.2, C5 -H), 7.17 (2H, dd, J 3.6 & 1.2, C3 -H), 7.02
The procedure was adapted from that of Nilles and Schuetz.[22]
To a round-bottomed flask charged with sodium hydride (67.1 mg,
2.80 mmol, 2.4 equiv.) was added ester 6 (293.7 mg, 1.16 mmol,
1.0 equiv.) followed by a pre-dried (CaSO4) solution of tropenol 7
(193 mg, 1.39 mmol, 1.2 equiv.) in n-heptane (13.9 mL). The reaction
mixture was heated at reflux under a nitrogen atmosphere for 2.5 h.After
cooling, the mixture was quenched carefully with water (10 mL) and
dichloromethane (10 mL). The organic phase (top layer) was separated
and the aqueous phase extracted with dichloromethane (2 × 15 mL).The
combined organic layers were washed with brine, dried (Na2SO4), fil-
tered, and concentrated to dryness to give a tan coloured solid. Purifica-
tion by flash chromatography (MeOH/CH2Cl2/NH3 10/90/1) afforded
ester 12[24] (241 mg, 58%) as a white solid. Rf (MeOH/CH2Cl2/NH3
10/90/1) 0.30. ν (thin film)/cm−1 3472 (OH), 3101, 3069, 2955, 2934,
2870 (C–H), 1728 (C=O ester). δH (200 MHz, CDCl3) 7.29 (2H, dd,
ꢀꢀ
(2H, dd, J 5.1 & 3.7, C4 -H), 5.22 (1H, t, J 6.0, C7-H), 4.04 (2H, m,
C1-H, C5-H), 3.40 (2H, s, C2-H, C4-H), 3.34 (3H, s, N-CH3), 3.10 (3H,
s, N-CH3), 2.79 (2H, ddd, J 18.0 & 6.0 & 4.2, C6-Ha, C8-Ha), 2.02 (2H,
d, J 17.6, C6-Hb, C8-Hb). δC (75 MHz, MeOD) 171.5, 147.4, 127.8,
127.5, 127.2, 78.1, 66.8, 65.1, 57.2, 55.0, 48.2, 29.8. m/z (ESI+) 392
(M, 100%). (HRMS (ESI+) calc. 392.0990. C19H22NO4S2 (M) found
392.0996.) The HPLC retention time for tiotropium 2 trifluoroacetate
correlated with that of a commercially derived sample of tiotropium
bromide.
Radiochemistry
General Procedures
A small septum-sealed vial containing a solution of the labelling
precursor (1.5 mg) dissolved in anhydrous DMF (200 µL) was cooled
to −40◦C and [11C]methyl iodide was transferred into the vial by a
stream of helium carrier gas for 2 min. The solution was heated at 88◦C
in a water bath for 5 min and then diluted with 200 µL of HPLC sol-
vent. The mixture was injected into a HPLC and the effluent from the
column was monitored with both UV and radioactivity detectors. The
peak corresponding to the carbon-11 labelled product was collected in
a rotary evaporator, the solvent was removed under reduced pressure,
and then the residue was diluted with water. The radiochemical purity
and specific radioactivity was determined by HPLC (Waters Spherisorb
S10 SCX, 5 µm, 4.6 × 250 mm2, acetonitrile/0.1 M NaH2PO4 50/50,
2 mL min−1).The area of the absorbance peak corresponding to the radi-
olabelled product of known radioactivity was measured at 210 nm (for
1) or 238 nm (for 2) by an automated integrating recorder and compared
to a standard curve relating mass to UV absorbance.
ꢀꢀ
ꢀꢀ
J 5.1 & 1.2, C5 -H), 7.16 (2H, dd, J 3.7 & 1.2, C3 -H), 7.00 (2H, dd,
ꢀꢀ
J 5.0 & 3.6, C4 -H), 5.63 (2H, s, C6-H, C7-H), 5.11 (1H, t, J 6.0, C3-H),
3.33 (2H, m, C1-H, C5-H), 2.25 (3H, m, C2-Ha, C4-Ha, OH), 2.23 (3H,
s, N-CH3), 1.69 (2H, d, J 14.6, C2-Hb, C4-Hb). δC (50 MHz, CDCl3)
171.3, 145.7, 131.5, 126.7, 126.0, 125.7, 71.0, 65.2, 41.3, 33.1 (one
signal overlapping). m/z (ESI+) 362 (M + H, 100%).
(1R,2R,4S,5S,7s)-2ꢀ-Hydroxy-2ꢀ,2ꢀ-di(thiophen-2ꢀꢀ-yl)acetic
Acid 9-Methyl-9-aza-3-oxatricyclo[3.3.1.02,4]non-7-yl
Ester 13[24]
The procedure was adapted from that of Rapp and Sobbota.[24]
To a mixture of ester 12 (142 mg, 0.393 mmol, 1.0 equiv.) and vana-
dium(v) oxide (7.2 mg, 0.0393 mmol, 0.1 equiv.) was added DMF
(2.3 mL) followed by 30% hydrogen peroxide (107 µL, 0.944 mmol,
2.4 equiv.) dropwise. The reaction mixture was stirred at 57◦C for 4.5 h.
After cooling to 40◦C, the reaction was quenched with sodium bisul-
fite (a mixture of NaHSO3 and Na2S2O5) and then acidified with
2 M hydrochloric acid. After stirring at 40◦C for 10 min, the mixture
was cooled to room temperature and diluted with 5% sodium hydro-
gencarbonate solution. The product was extracted into ethyl acetate
(4 × 10 mL) and the combined organic layers washed with brine, dried
(Na2SO4), filtered, and concentrated under vacuum. Residual DMF
was removed under high vacuum. Flash chromatographic purification
(MeOH/CH2Cl2/NH3 6/94/1) afforded epoxide 13[24] (119 mg, 80%)
as a white solid. Rf (MeOH/CH2Cl2/NH3 10/90/1) 0.56. Rt (Spherisorb
S10 SCX 10 µm, 10 × 250 mm2, acetonitrile/0.1 M NaH2PO4 40/60,
5 mL min−1) 7.20 min. ν (thin film)/cm−1 3481 (OH), 3103, 3039, 2959,
2945 (C–H), 1732 (C=O ester). δH (200 MHz, CDCl3) 7.31 (2H, dd, J
Synthesis of [11C]Ipratropium 1 Iodide [11C]1
The labelling was conducted according to the general procedure
to afford [11C]ipratropium 1 iodide [11C]1 (>98% chemical and
radiochemical purity). Rt (Spherisorb S10 SCX 10 µm, 10 × 250 mm2,
acetonitrile/0.1 M NaH2PO4 60/40, 6 mL min−1) 6.67 min. The radio-
chemical yield was 61 3 MBq (n = 3) at end-of-synthesis (0.3%,
calculated from [11C]CO2, non-decay corrected) and the specific
radioactivity at end-of-synthesis including formulation and quality
control (35 min) was measured at 11 GBq µmol−1
.
Synthesis of [11C]Tiotropium 2 Iodide [11C]2
The labelling was conducted according to the general procedure to
afford [11C]tiotropium 2 iodide [11C]2 (>98% chemical and radio-
chemical purity). Rt (Spherisorb S10 SCX 10 µm, 10 × 250 mm2,
acetonitrile/0.1 M NaH2PO4 40/60, 6 mL min−1) 6.68 min. The radio-
chemical yield calculated was 129 8 MBq (n = 3) at end-of-synthesis
(0.5%, calculated from [11C]CO2, non-decay corrected) and the specific
ꢀꢀ
ꢀꢀ
5.1 & 1.2, C5 -H), 7.12 (2H, dd, J 3.6 & 1.2, C3 -H), 6.99 (2H, dd, J
ꢀꢀ
5.1 & 3.6, C4 -H), 5.13 (1H, t, J 5.4, C7-H), 4.81 (1H, br s, OH), 3.09
(2H, m, C1-H, C5-H), 3.00 (2H, s, C2-H, C4-H), 2.47 (3H, s, N-CH3),
2.16 (2H, dt, J 15.7 & 5.2, C6-Ha, C8-Ha), 1.60 (2H, d, J 15.4, C6-Hb,
C8-Hb). δC (50 MHz, CDCl3) 171.4, 145.2, 126.8, 126.2, 126.1, 70.5,