Isolation of Lamotrigine 2-N-glucuronide from Guinea-pig Urine

Lamotrigine (L T) is a novel anticonvulsant. Its major metabolite in human is 2-N-glucuronide (L T -2NG). In order to investigate the metabolic characteristics of L T in our laboratory, a reference standard of L T -2NG was required. The purpose of this experiment was to isolate pure L T -2NG from the urine of L T -treated guinea pigs. The pooled urine of guinea pigs fed with L T was eluted with methanol through XAD-2 column. L T-2NG in the eluent was purified by semipreparative HPLC equipped with a C8 column and a UV detector set at 267 nm. The mobile phase for HPLC was 0.0lM ammonium acetate (pH 6.6) containing12% of methanol. The isolated L T-2NG was confirmed by mass, 1H NMR and 13C NMR spectroscopic analysis. The molecular ion 432.1, a downfield anomeric proton at 5.39 ppm, and an upfield shift (-6.9 ppm) of the triazine ring C-3 indicate attachment of the glucuronide to the N-2 of LT. These spectra were identical with the reported spectra of L T -2NG isolated from human urine.