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Development and validation of a sensitive LC‐MS/MS method with electrospray ionization for quantitation of doxofylline in human serum: application to a clinical pharmacokinetic study
Author(s) -
Sreenivas Nimmagadda,
Narasu M. Lakshmi,
Shankar B. Prabha,
Mullangi Ramesh
Publication year - 2008
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.984
Subject(s) - chemistry , chromatography , electrospray ionization , formic acid , protein precipitation , selected reaction monitoring , detection limit , pharmacokinetics , analyte , acetonitrile , electrospray , mass spectrometry , tandem mass spectrometry , pharmacology , medicine
A highly sensitive and specific LC‐MS/MS method has been developed and validated for the estimation of doxofylline (DFL) with 300 µL human serum using imipramine as the internal standard (IS). The API‐3000 LC‐MS/MS was operated under multiple reaction‐monitoring mode using the electrospray ionization technique. The assay procedure involved direct precipitation of DFL and IS from human serum with acetonitrile. The resolution of peaks was achieved with formic acid (pH 2.5):acetonitrile (10:90, v/v) on an Amazon C 18 column. The total chromatographic run time was 3.0 min and the elution of DFL and IS occurred at approximately 1.46 and 2.15 min, respectively. The MS/MS ion transitions monitored were 267.5 → 181.1 for DFL and 281.1 → 86.2 for IS. The method was proved to be accurate and precise at linearity range of 1.00–5000 ng/mL with a correlation coefficient ( r ) of ≥0.999. The method was rugged with 1.00 ng/mL as lower limit of quantitation. The intra‐ and inter‐day precision and accuracy values were found to be within the assay variability limits as per the FDA guidelines. The developed assay method was applied to a pharmacokinetic study in human volunteers following oral administration of DFL tablet. Copyright © 2008 John Wiley & Sons, Ltd.