Purpose: To evaluate the sensitivity requirement for LC-MS/MS as an analytical tool to support human microdosing study with sub-pharmacological dose, investigate proportionality of pharmacokinetics from the microdose to therapeutic human equivalent doses in rats and characterize circulating metabolites in rats administered with the microdose.
Materials and methods: Five drugs of antipyrine, metoprolol, carbamazepine, digoxin and atenolol were administered orally to male Sprague-Dawley rats at 0.167, 1.67, 16.7, 167 and 1,670 microg/kg doses. Plasma samples were extracted using either solid phase extraction or liquid-liquid extraction, and analyzed using LC-MS/MS.
Results: Using 100 microl of plasma sample, the lower limit of quantitation for antipyrine (10 pg/ml), carbamazepine (1 pg/ml), metoprolol (5 pg/ml), atenolol (20 pg/ml), and digoxin (5 pg/ml) were achieved using an API 5000. Proportional pharmacokinetics were observed from 0.167 microg/kg to 1,670 microg/kg for antipyrine and carbamazepine and from 1.67 to 1,670 microg/kg for atenolol and digoxin, while metoprolol exhibited a non-proportional pharmacokinetics relationship. Several metabolites of carbamazepine were characterized in plasma from rats dosed at 1.67 mug/kg using LC-MS/MS.
Conclusions: This study has shown the promise of sensitive LC-MS/MS method to support microdose pharmacokinetics and drug metabolism studies in human.