G. Srinivas et al.
spectroscopic methods. 1H-NMR (400 MHz, DMSO-d6): d: 3.3 (s, 3H), 4.3 autoradiography. The radioactivity in the ethyl acetate extract was
(d, 2H), 4.6 (m, 1H), 5.0 (d, 1H), 6.4 (s, 1H), 7.6 (d, 2H), 7.8 (d, 2H) 8.6 (s, 1.5 mCi (55.5 MBq), radiochemical yield was 53.57%, and the overall
1H); Mass: m/z,381 (M+ Na)+; IR: (KBr) Vmax (cmÀ1) 1275.90 (C–O), yield of the product was 30% based on [14C]-methyl iodide. The final
1368.39 (SO), 1537.31 (CC), 1683.00 (CO), 3029.62 (C–H), 3320.86 (N–H), product on radiochromatographic assay was found to be more than
3453.81 (O–H); and its [a]2D0 À18.02 (c = 1.0, DMF) at 25 ꢀC. Melting 96% radiochemically pure. Its weight was 10.75 mg and specific activity
point of the inactive florfenicol prepared was found to be 154 ꢀC ~50 mCi (1.85 GBq/mmol).
(uncorrected). They were in good agreement with the reported values
for the authentic sample of non-labeled florfenicol.
Results and discussion
Synthesis of (4S,5R)-4-(fluoromethyl)-5-(4-[14C]-methylsulfonyl)-
In the literature, several approaches for the synthesis of florfenicol
phenyl)- oxazolidin-2-one (compound III)
have been reported.3,4,6–16 But none of the reported procedures
Sodium sulfite (429 mg, 3.4 mmol) and sodium bicarbonate (286 mg,
3.4 mmol) were taken in a single necked 25-mL capacity round bottomed
flask having a magnetic bar. Water (5 mL) was then added to it and the
mixture was heated to 70 ꢀC for 2 h, with stirring. It was then allowed
to cool. Then, compound II (400 mg, 1.3 mmol) was added to the reaction
flask and the mixture was then heated at 70 ꢀC for 3–4 h. The solution
was stirred overnight at room temperature. Dry THF (5 mL) was then
added. A side-arm connection was provided to the flask through which
a narrow tube was introduced that almost touched the bottom of the
flask. Through another side-arm, a cold finger type condenser was
could be adopted as such because our requirement was for the
[S-methyl-14C]-florfenicol.
Introduction of the radioisotopic label at the latest possible
stage of the synthesis has generally been recognized as ideal on
consideration of yield, cost, risk, and so on. As a rule of thumb,
the production of radiolabeled compounds of high radiochemical
purity and specific activity, in enhanced or moderate yields,
demands novel and/or facile synthetic schemes and also clean
workup procedures. In view of the aforementioned, for obtaining
provided in which a dry ice-acetone bath was kept for condensing any a high-specific activity product, the radiochemical synthesis was
un-reacted [14C]-methyl iodide getting released from the reaction flask.
[
carried out on a submicromolar scale without adding the carrier
14C]-Methyl iodide [5 mCi, 185 MBq., specific activity 57 mCi (2.11 GBq/
and employing the newly designed and developed method after
standardizing the procedure using inactive materials.
Compound I required for use as the precursor for the [S-
methyl-14C]-florfenicol synthesis was prepared in-house starting
from cinnamyl alcohol employing multiple steps of a reported
procedure17 with certain modifications. This work will be reported
elsewhere.
mmol)] contained in a break seal tube was connected to the system.
The reaction mixture was cooled to À45 ꢀC using acetone-solid CO2 bath.
The system was continuously flushed with nitrogen gas. The break seal
was then broken with a magnetic hammer and [14C]-methyl iodide was
allowed to bubble through the reaction mixture along with nitrogen.
After completion of the transfer of [14C]-methyl iodide to the reaction
flask, the stirring of the mixture was continued for a further period of
6 h while raising the temperature of the reaction mixture to 40 ꢀC. The
Chlorosulfonation of compound
I
gave the sulfonyl
product formation was checked and confirmed by means of TLC chloride compound II after purification by silica gel column
(silica gel). The crude [14C]-labeled product formed was isolated by
chromatography in 36.5%. This was followed by sodium salt
extracting with dichloromethane (5 mL Â 3). The product, on further
formation in situ by treatment with aqueous sodium sulfite and
sodium bicarbonate and introduction of [14C]-methyl group by
chromatographic analysis, was found to match with the spot of the
prepared sample of non-radioactive product. The radioactivity of the
product obtained was 3.5 mCi (129.5 MBq). The radiochemical yield
was 70%.
Synthesis of (1R, 2S)-2-amino-3-fluoro-1-(4-[14C]-methylsulfonyl)-
phenyl) propan-1-ol (compound IV)
In a cylindrical glass tube, compound III (3.5 mCi, 129.5 MBq) and dioxane
(10 mL) were taken. After cooling the solution at 0 ꢀC 4.5 N, sulphuric acid
(5 mL) was added slowly with stirring. The glass tube was then flame
sealed and covered with wire gauze. The reaction mixture was heated
at 110–120 ꢀC for 9–10 h in an oil bath. The [14C] product was isolated
by extraction with dichloromethane (5 mL Â 4). The product was assayed
for radioactivity. It was found to contain 2.8 mCi (103.6 MBq) activity and
the radiochemical yield was 80%.
Synthesis of [S-methyl-14C]-florfenicol (compound V)
In a single necked round bottomed flask, compound IV (2.8 mCi,
103.6 MBq) and dry methanol (3 mL) were taken. Methyl-2,2-
dichloroacetate (0.69 g, 4 mmol) and triethylamine (0.1 mL) were added
to it with stirring. The reaction mixture was subsequently heated at
45 ꢀC with stirring for 7–8 hrs. It was then allowed to reach at room
temperature. The solution was evaporated and the final crude product
was purified by preparative thin layer silica gel chromatography using
the solvent system—30% ethyl acetate in hexane. After drying the
TLC, it was kept for autoradiography. From the TLC and autoradiograph,
the product band region corresponding to the florfenicol was identified
and the portion of silica gel containing [S-methyl-14C]-florfenicol was
scraped-off. It was extracted with ethyl acetate (5 mL Â 5). The final
purified product was spotted, chromatographed on TLC and kept for
Scheme 1. Synthesis of [S-methyl-14C]-florfenicol
J. Label Compd. Radiopharm 2013, 56 382–384
Copyright © 2013 John Wiley & Sons, Ltd.
Srinivas
