ISOLATION AND IDENTIFICATION OF A MAJOR METABOLITE OF SFZ-47 IN THE RABBIT URINE

AIMTo isolate and identify a glucuronide metabolite of SFZ-47 ［3H-1,2-dihydro-2-(4-methyl-phenylamino)methyl-1-pyrrolizinone］, which is difficult to synthesize because it undergoes hydrolysis and intramolecular acyl migration at physiological pH, in rabbit urine. METHODSTwo rabbits were ig 200 mg doses of SFZ-47. Urine was collected for 24 h, adjusted to pH 4.0 with acetic acid and lyophilized. The residues were reconstituted in 25 mL methanol and centrifuged at 5 000 r·min^-1 for 10 min. The supernatant was filtered (0.45 μm) and then isolated with semi-preparative reversed phase HPLC. The eluent collected from individual peaks was evaporated by rotary evaporation and freeze-drying. Compounds were then identified with electrospray ion trap mass spectrometry and ¹HNMR spectroscopy. RESULTSThe ¹HNMR and ESI-MSⁿ results indicate that the metabolite is the 1-O-acyl β-D-glucuronide conjugate of 4-(3H-1,2-dihydro-1-pyrrolizinone-2-methylamino) benzoic acid. CONCLUSIONThis method was shown to be rapid and simple and gave excellent resolution from endogenous constituents in urine, and it is suitable for preparation of the glucuronide metabolites of SFZ-47 and its analogues.