Simultaneous determination of a p38 MAP kinase inhibitor and its amide hydrolyzed metabolite in Cynomolgus monkey plasma by LC–MS/MS, and its application to a toxicokinetic study

A LC–MS/MS method was developed for the determination of a p38 MAP kinase inhibitor (Compound I) and its amide hydrolyzed metabolite (M7) in Cynomolgus monkey plasma over the concentration range of 1.00–1000 ng/mL. Stable isotope labeled compounds (d 3-Compound I and d 3-M7) were used as internal st...

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Bibliographic Details
Published in:Journal of pharmaceutical and biomedical analysis 2011-07, Vol.55 (5), p.1104-1110
Main Authors: Akrami, Anna, Hsieh, Faye, Almon, Valerie, Bruenner, Bernd A., James, Christopher, Wong, Philip
Format: Article
Language:English
Subjects:
acetonitrile
Acetonitriles - chemistry
Administration, Oral
Amides - chemistry
Analysis
Analytical, structural and metabolic biochemistry
Animals
Area Under Curve
atmospheric pressure
Biological and medical sciences
Calibration
Chemistry Techniques, Analytical
Chromatography, Liquid - methods
Cynomolgus
Enzyme Inhibitors - analysis
Enzyme Inhibitors - blood
Female
Food and Drug Administration
Formates - chemistry
Fundamental and applied biological sciences. Psychology
General pharmacology
hydrolysis
Inflammation
ionization
Ions
LC–MS/MS
Macaca fascicularis
Male
Medical sciences
metabolites
mitogen-activated protein kinase
monitoring
oral administration
p38 MAP kinase inhibitor
p38 Mitogen-Activated Protein Kinases - antagonists & inhibitors
Pharmacology. Drug treatments
Quality Control
Reproducibility of Results
reversed-phase liquid chromatography
stable isotopes
Tandem Mass Spectrometry - methods
Water - chemistry
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Summary:A LC–MS/MS method was developed for the determination of a p38 MAP kinase inhibitor (Compound I) and its amide hydrolyzed metabolite (M7) in Cynomolgus monkey plasma over the concentration range of 1.00–1000 ng/mL. Stable isotope labeled compounds (d 3-Compound I and d 3-M7) were used as internal standards (IS). Samples were prepared using protein precipitation in the 96-well format with a 30 μL plasma sample volume. Chromatographic separation was performed with reversed-phase liquid chromatography on a Varian Monochrom C 18 (100 mm × 2.00 mm, 5 μm) analytical column. The mobile phases were 5 mM ammonium formate in acetonitrile/water (95/5, v/v) pH 7.0 and 5 mM ammonium formate in acetonitrile/water (5/95, v/v) pH 7.0. Gradient elution, at a flow rate of 550 μL/min, was used to separate Compound I and M7. Positive atmospheric pressure chemical ionization was utilized with detection by multiple reaction monitoring (MRM). Total run time was about 3.2 min. This method was validated following the current Food and Drug Administration (FDA) guidance for bioanalytical method validation. The intra- and inter-day precision (% CV) and accuracy (% bias) at all concentrations tested were below 15% for both analytes. The mean recoveries for Compound I, M7, d 3-Compound I, and d 3-M7 were 106%, 107%, 108% and 105%, respectively. The method was successfully applied to support a GLP toxicokinetic study in Cynomolgus monkeys after oral administration of Compound I. A total of 48 samples (∼12.5% of the total number of samples) were selected for incurred sample reanalysis (ISR). The % difference between the reassay concentrations and the original concentrations were all less than 20% of their mean values and met the acceptance criteria for ISR.
ISSN:0731-7085
1873-264X
DOI:10.1016/j.jpba.2011.03.022