Magnesium lithospermate B is excreted rapidly into rat bile mostly as methylated metabolites, which are potent antioxidants.

To elucidate the in vivo pharmacological activities of magnesium lithospermate B (MLB), an active constituent of Radix Salviae Miltiorrhizae, in the rat, its metabolic fate both in vivo and in vitro was investigated. High-performance liquid chromatography revealed that four major metabolites with lower polarity were excreted into bile after intravenous and oral administration of MLB. The metabolites present in combined samples of bile from rats after intravenous injection were isolated and purified by column chromatography and identified as four meta-O-methylated products, namely 3-monomethyl- (M1), 3,3"'-dimethyl- (M2), 3,3"-dimethyl-, and 3,3",3"'-trimethyl-lithospermic acid B according to their spectroscopic characteristics ((1)H, (13)C NMR, (1)H-(1)H correlation spectroscopy, (1)H-detected multiple quantum coherence, and heteronuclear multiple bond coherence combined with positive ion fast atom bombardment-mass spectroscopy). After administration of MLB at an intravenous dose of 4 mg/kg or an oral dose of 100 mg/kg, the total biliary recovery of the four metabolites after 30 h reached 95.5 +/- 2.4% (with approximately 90% recovered within 2 h) or 5.5 +/- 0.7%, respectively. The metabolic pathway was proposed to involve sequential formation of the four methylated metabolites. Incubation of MLB, M1, M2, or M4 in rat hepatic cytosol in the presence of S-adenosyl-l-methionine demonstrated the formation of all four metabolites, which indicated that the enzyme responsible for the biotransformation is catechol O-methyltransferase. MLB and its main metabolites M1 and M2 showed potent 1,1-diphenyl-2-picrylhydrazyl radical-scavenging activities, the activity of M1 being stronger than those of caffeic acid (the monomer form of MLB) and alpha-tocopherol (a representative antioxidant) but weaker than that of MLB. The rapid and high biliary excretion levels of these metabolites suggested that they could undergo enterohepatic circulation in rats and that they might thereby be largely responsible for the pharmacological effects of MLB.