Comprehensive Investigation of the Metabolism Profile of Norisoboldine in Rats Using UPLC-Q-TOF-MS/MS Method

Linderae Radix, a traditional Chinese medicine known for its anti-inflammatory, antirheumatic, and analgesic effects, contains norisoboldine (NOR) as its primary bioactive component. NOR exhibits significant anti-inflammatory effects in preclinical studies, particularly in rheumatoid arthritis and inflammatory bowel disease. Despite the various pharmacological activity of NOR, its metabolism and biotransformation patterns <i>in vivo</i> have yet to be elucidated. This study aimed to systematically elucidate the <i>in vivo</i> metabolic pathways of NOR, the excretion patterns of its metabolites, and the primary metabolic enzyme subtypes involved. Biosamples were collected from rats following oral administration of NOR (30 mg/kg). UPLC-Q-TOF-MS/MS was used to identify NOR metabolites in plasma, tissues, and excreta. Differences in the distribution of metabolites across excretion pathways were compared. Chemical inhibition assays and human recombinant enzyme experiments were conducted to determine the key cytochrome P450 (CYP450) enzyme subtypes involved. A total of 14 metabolites were identified, including 8 phase I metabolites and 6 phase II conjugates; the major metabolic reactions were hydrolysis, glucuronidation, sulfation, and dehydrogenation. The glycosylated conjugates (M13 - M15) predominantly underwent renal excretion via urine, whereas the hydrophobic metabolites (M2 - M9) were primarily eliminated through fecal routes due to limited aqueous solubility. The identification of CYP450 enzyme subtypes indicated that CYP3A1/2 and CYP2C11 were the key functional subtypes mediating NOR biotransformation. This study provides the first comprehensive characterization of NOR metabolism, demonstrating that CYP3A1/2 and CYP2C11 catalyze its conversion into 14 metabolites. These findings provide a theoretical basis for the clinical development of NOR and metabolic research on similar compounds.