D. Zhong and A. H. Lewin
CH3OH (5 mL) in another volumetric flask (5 mL). The HPLC peak equilibrated with hexane (6mL). Crude [3H]-9 (200mCi) in
areas of aliquots (5, 10, 15, 20, and 25 mL) of the solution were toluene was mixed with hexane (12 mL) and the mixture was
measured at 235 nm and a calibration curve was generated passed through the cartridge. The cartridge was eluted with
(Figure 1) for use in the determination of the specific activity of hexane (2 mL), hexane/EtOAc (1/1, 2 Â 2 mL), EtOAc (2 Â 2 mL).
[3H]-1.
Most of the activity was found in the hexane/EtOAc (1/1,
2 Â 2 mL) and EtOAc (2Â 2 mL) eluents (171 mCi/91% radio-
chemical purity in the combined fraction).
21-Acetoxy-6a,9,-difluoro-11b-hydroxy-16a,17-[(1-methy-
lethylidene)bis(oxy)]pregna-1,4-diene-3,20-one (21-O-acetyl
fluocinolone acetonide, (2)
11b,21-Diacetoxy-6a,9,-difluoro-16a,17-[(1-methylethylide-
ne)bis(oxy)]pregna-1,4-diene-3,20-one (1b,21-O-,O-diacetyl
fluocinolone acetonide, 8)
To a mixture of fluocinolone acetonide (1) (203 mg, 0.44 mmol)
in CH2Cl2 (5 mL, anhydrous) was added acetic anhydride
(0.050 mL, 54 mg, 0.53 mmol, 1.2 eq.) followed by a solution A solution of 1 (201 mg, 0.44 mmol) in pyridine (1.3 mL, 1.277 g,
(1 M) of trimethylsilyl trifluoromethanesulfonate (0.040 mL, 16.1 mmol, anhydrous) and acetic anhydride (0.67 mL, 0.724 g,
442 mg, 2.0 mmol) in CH2Cl2. The mixture was stirred at RT for 7.09 mmol) was heated under N2 for 2 h. The solution was
35 min at which time more acetic anhydride (50 mL, 54 mg, cooled to RT, stirred at RT for 2 h, then mixed with H2O (01C,
0.53 mmol, 1.2 eq.) was added. Stirring at RT was continued for 20 mL). The mixture was centrifuged at 3000 rpm for 2 min and
35 min more, then the excess acetic anhydride was destroyed by the supernatant was removed. The white solids were washed
addition of CH3OH (0.5 mL). The mixture was mixed with H2O with H2O (01C, 2 Â 20 mL), freeze-dried overnight, leaving a
(10 mL) and CH2Cl2 (20 mL). The organic phase was separated, white solid (208 mg) containing 77% of 21-acetate 2 and 23% of
washed with H2O (10 mL), then dried over Na2SO4. The solvent 11,21-diacetate 8.
was evaporated, leaving a white solid (130 mg, 0.26 mmol, 59%),
The white solid (206 mg) was mixed with 4-dimethyamino-
m.p. 42501C. 1H NMR (300 MHz, CDCl3) d (ppm): 0.94 (s, 3H, pyridine (DMAP, 35 mg, 0.29 mmol), pyridine (1.3 mL, 1.277 g,
H-18), 1.22 (s, 3H, H-23), 1.44 (s, 3H, H-22), 1.53 (s, 3H, H-19), 16.1 mmol, anhydrous), and acetic anhydride (0.67 mL, 724 mg,
1.58–1.87 (m, 4H, H-7a, H-12b, H-15), 2.19 (s, 3H, H-24), 2.08–2.59 7.09 mmol). The light yellow solution was stirred at RT for 18 h,
(m, 4H, H-7b, H-8, H-12a, H-14), 4.42 (d, J = 8.2 Hz, 1H, H-11), mixed with H2O (01C, 30 mL). The mixture was centrifuged at
4.9 (bs, 2H, H-21), 5.00 (d, J = 4.6 Hz, 1H, H-16), 5.50 3000 rpm for 2 min and the supernatant was removed. The off-
(qd, J = 48.7 Hz, 1.7 Hz, 1H, H-6), 6.38 (dd, J = 10.1 Hz, 1.7 Hz, white solids were washed with H2O (01C, 30 mL), citric acid (01C,
1H, H-2), 6.44 (m, 1H, H-4), 7.13 (dd, J = 10.1 Hz, 1.0 Hz, 1H, H-1). 0.1 M, 2 Â 30 mL), H2O (01C, 2 Â 30 mL), then freeze-dried over-
13C NMR (CDCl3, 75.5 MHz) d (ppm): 16.7, 20.8, 23.4, 23.5, 26.1, night, leaving a white solid (8) (207 mg, 0.38 mmol, yield: 86%,
1
26.9, 32.1, 32.2, 32.3, 32.5, 33.9, 34.2, 34.4, 37.5, 43.1, 45.9, 48.2, purity: 97%), m.p. 42501C H NMR (300 MHz, CDCl3) d (ppm):
48.5, 68.0, 71.8, 72.4, 82.2, 85.5, 87.9, 97.7, 97.8, 112.1, 121.5, 0.84 (s, 3H, H-18), 1.22 (s, 3H, H-23), 1.37(s, 1H, H-22), 1.44 (s, 3H,
121.7, 130.6, 150.7, 161.4, 161.6, 171.0, 185.8, 203.9. MS m/z H-19), 1.57–1.88 (m, 4H, H-7a, H-12b, H-15), 2.13 (s, 3H, H-24),
calcd for C29H32F2O7: 494.21; found (APCI) 495 (M1H). Anal 2.17 (s, 3H, H-25), 2.19-2.54 (m, 4H, H-7b, H-8, H-12a, H-14), 4.73
calcd for C29H32F2O7: C 63.15; H 6.52; F 7.68; found: C 62.89; H (d, J = 17.8 Hz, 1H, H-21), 4.96 (d, 1H, J = 17.9 Hz , H-21), 5.01 (d,
6.53; F 7.47.
J = 4.6 Hz, 1H, H-16), 5.31 (qd, J = 48.6 Hz, 1.8 Hz, 1H, H-6),
5.46–5.50 (m, 1H, H-11), 6.38 (dd, J = 10.1 Hz, 1.8 Hz, 1H, H-2),
6.45 (m, 1H, H-4), 6.78 (dd, J = 10.1 Hz, 1.4 Hz, 1H, H-1), in good
agreement with the literature,9 13C NMR (CDCl3, 75.5 MHz)
d (ppm): 16.3, 20.7, 21.8, 23.3, 23.4, 26.0, 26.8, 32.6, 32.8, 32.9,
33.0, 33.5, 33.8, 34.0, 34.2, 42.7, 42.8, 45.4, 67.8, 71.3, 71.3, 82.0,
85.2, 87.6, 97.4, 112.2, 121.8, 122.0, 131.2, 148.9, 169.0, 170.5,
185.1, 203.8, MS (APCI) m/z = 537 (M1H1).
[1,2-3H]-21-acetoxy-6a,9,-difluoro-11b-hydroxy-16a,17-[(1-
methylethylidene)bis(oxy)]pregn-4-ene-3,20-one
(21-O-
acetyl 1,2-dihydrofluocinolone acetonide, [3H]-9)
The light yellow solution of 2 (3.8 mg, 0.0077 mmol), Wilkinson’s
catalyst (chlorotris(triphenylphosphine)-rhodium (I)) (3.8mg,
0.0041mmol) in toluene (filtered through an aluminum
oxide pad packed in a disposable glass pipette, 0.8 mL) and
THF (filtered through an aluminum oxide pad packed in a
disposable glass pipette, 0.4 mL) in a round bottomed flask (2 mL)
equipped with a stirrer bar was de-gassed three times on a
[1,2-3H]-11b,21-diacetoxy-6a,9,-difluoro-16a,17-[(1-methy-
lethylidene)bis(oxy)]pregn-4-ene-3,20-one (1b,21-O-,O-dia-
cetyl dihydrofluocinolone acetonide,[3H]-11)
tritiation system by freezing/evacuating/thawing/the content in Crude [3H]-9 recovered from a failed DDQ oxidation (255 mCi,
the flask. The mixture was stirred under tritium gas (658mmHg) 5.6 mg DDQ, and 0.27 mL benzene) was purified using the
for 48h. The black reaction mixture was filtered through a procedure described in the preparation of [3H]-9 affording
Celite pad packed in a disposable glass pipette. The reaction flask 90.3 mCi (89% radiochemically pure). This material was com-
was washed with CH3OH (4Â 1 mL) and the washings were bined with separately purified [3H]-9. The total (261 mCi) was
filtered through the same Celite pad. The combined filtrate combined with DMAP (4-dimethylaminopyridine) (2.5 mg,
was concentrated to dryness using a rotary evaporator. The 0.02 mmol), pyridine (0.13 mL, 128 mg, 1.62 mmol), and acetic
residue was dissolved in CH3OH (2mL) and the solution was anhydride (0.067 mL, 72 mg, 0.71 mmol) in a 2-dram vial and
mixed with H2O (9 mL). The slightly cloudy solution was passed stirred under N2 at RT for 15 h. The reaction mixture was mixed
through a Supelco Discovery DSC-18 SPE cartridge (500mg/3mL, with hexane (4 mL)/EtOAc (1 mL), and the solution was washed
activated with 3 mL CH3OH and equilibrated with 3 mL H2O with citric acid (01C, 0.1 M, 3 Â 2 mL), H2O (2 mL), dried over
before use). The cartridge was washed with H2O (4mL) and Na2SO4, leaving [3H]-11 (238 mCi/97% radiochemical purity).
eluted with CH3OH (2mL), leaving 500.2 mCi crude [3H]-9 in the The solvent was evaporated and the residue was dried by
CH3OH solution. A portion of this material was purified as follows. dissolving the residue in CH3CN (1 mL) and evaporating the
A Supelco LC-Alumina N SPE cartridge (500mg/3mL) was solvent twice.
Copyright r 2009 John Wiley & Sons, Ltd.
J. Label Compd. Radiopharm 2009, 52 103–109