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Abstract
A sensitive and selective liquid chromatography-mass spectrometry (LC-MS) method for determination of tolbutamide and its metabolite hydroxytolbutamide in rat plasma was developed and validated. The analytes and internal standard omeprazole were extracted from plasma by liquid-liquid extraction using ethyl acetate, and chromatographically separated on a Zorbax SB-C18 column (2.1 mm × 50 mm, 3.5 µm) using acetonitrile-0.1% formic acid as the mobile phase with gradient elution. Electrospray ionization (ESI) source was applied and operated in positive ion mode, and selected ion monitoring (SIM) mode used to quantify tolbutamide and its metabolite hydroxytolbutamide. The assay was linear over the range 10–20000 ng/mL for tolbutamide and 5–800 ng/mL for hydroxytolbutamide, with a lower limit of quantification (LLOQ) of 10 ng/mL for tolbutamide and 5 ng/mL for hydroxytolbutamide. Intra- and inter-day precision were less than 12% and the accuracy were in the range 88.8–109.7%. This developed method was successfully used for determination of tolbutamide and metabolite hydroxytolbutamide in rat plasma for pharmacokinetic study.
ACKNOWLEDGMENT
This work was supported by Shanghai Key Laboratory Program of Forensic Medicine (Institute of Forensic Science, Ministry of Justice, China), No. KF1101; Wenzhou Municipal Science and Technology Agency-funded projects of Zhejiang Province, China, No. Y20090143; and National Youth Natural Science Foundation of China, No. 81102297.