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Development and validation of a high‐sensitivity liquid chromatography/tandem mass spectrometry (LC/MS/MS) method with chemical derivatization for the determination of ethinyl estradiol in human plasma
Author(s) -
Shou Wilson Z.,
Jiang Xiangyu,
Naidong Weng
Publication year - 2004
Publication title -
biomedical chromatography
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.4
H-Index - 65
eISSN - 1099-0801
pISSN - 0269-3879
DOI - 10.1002/bmc.329
Subject(s) - chemistry , chromatography , derivatization , liquid chromatography–mass spectrometry , tandem mass spectrometry , mass spectrometry , human plasma
An ultra‐sensitive liquid chromatography/tandem mass spectrometry (LC/MS/MS) method for the analysis of oral contraceptive ethinyl estradiol (EE) was developed and validated over the curve range of 2.5–500 pg/mL using 1 mL of human plasma sample. Ethinyl estradiol and the internal standard, ethinyl estradiol tetra‐deuterated (EE‐d4), were extracted from the plasma matrix with methyl t ‐butyl ether, derivatized with dansyl chloride and then back‐extracted into hexane. The hexane phase was evaporated to dryness, reconstituted and injected onto the LC/MS/MS system. The chromatographic separation was achieved on a Luna C 18 column (50 × 2 mm, 5 µm) with an isocratic mobile phase of 20:80 (v/v) water:acetonitrile with 1% formic acid. The ofine derivatization procedure introduced the easily ionizable tertiary amine function group to EE. This greatly improved analyte sensitivity in electrospray ionization and enabled us to achieve the desired lower limit of quantitation at 2.5 pg/mL. This high sensitivity method can be used for therapeutic drug monitoring or supporting bio‐equivalence and drug–drug interaction studies in human subjects. Copyright © 2004 John Wiley & Sons, Ltd.