Isolation and characterization of a novel quaternary ammonium-linked glucuronide of lamotrigine.
Lamotrigine (LTG) is a novel triazine anticonvulsant currently undergoing clinical trials. LTG N-glucuronide, the major human metabolite of LTG, was isolated from human urine by means of XAD-2 column chromatography and semi-preparative HPLC. The structure of the suspected lamotrigine 2-N-glucuronide was proven by mass spectroscopy and NMR spectroscopy, along with chemical and enzymatic hydrolysis studies. High resolution fast atom bombardment mass spectrometry and Electrospray tandem mass spectrometry of the glucuronide gave an M+ ion at 432.0 amu and a fragment ion at 256.0 (M - 176)+ amu. The proton NMR of the glucuronide indicated the presence of a glucuronic acid moiety. A downfield anomeric proton (5.35-5.60 ppm) implied direct attachment to the aromatic triazine ring. Carbon-13 NMR of the glucuronide revealed an upfield shift (delta = -7.0 ppm) of the C-3 carbon of the triazine ring compared to LTG, indicating attachment of the glucuronide to the N-2 position. Chemical degradation or rearrangement of the glucuronide occurs at neutral pH to produce an unknown product (RP-1), while at basic pH a different unknown product (RP-2) is formed. The glucuronide is unusually stable at acidic pH. Treatment of the glucuronide with beta-glucuronidase resulted in hydrolysis to LTG, and enzymatic hydrolysis was inhibited by saccharo-1,4-lactone.
Sinz
M W
MW
Department of Medicinal Chemistry, College of Pharmacy, University of Minnesota, Minneapolis 55455.
Remmel
R P
RP
D000641:Ammonia / Q000737:chemistry; D000927:Anticonvulsants / Q000652:urine*; D002851:Chromatography, High Pressure Liquid; D002855:Chromatography, Thin Layer; D005965:Glucuronates / Q000652:urine; D005966:Glucuronidase / Q000378:metabolism; D006801:Humans; D006868:Hydrolysis; D000077213:Lamotrigine; D009682:Magnetic Resonance Spectroscopy; D013058:Mass Spectrometry; D014227:Triazines / Q000652:urine*
D000927:Anticonvulsants; D005965:Glucuronates; D014227:Triazines; D000641:Ammonia; D005966:Glucuronidase; D000077213:Lamotrigine