Published:Journal of Chromatographic Science,
ISSN 0021-9665Volume
40, Number 4, April, pp. 214-218
Assay of TNP-470 and Its Two Major Metabolites in Human Plasma
by High-Performance Liquid Chromatography–Mass Spectrometry
Carl T. Whalen[1], Glenn
D. Hanson[1], Kathy J. Putzer[1], Michael D. Mayer[2], and Darcy J. Mulford[2]
[1]Covance Laboratories Inc., 3301 Kinsman Blvd., Madison, WI 53704 and [2]TAP
Pharmaceutical Products, Inc., 675 N. Field Drive, Lake Forest, IL 60045
In this study, a sensitive, specific assay for the determination
of TNP-470 and its two major metabolites M-IV (also know as AGM-1883) and M-II
in human plasma is reported. The assay involves liquid–liquid extraction
of acidified plasma followed by reversed-phase high-performance liquid chromatography–atmospheric
pressure chemical ionization–tandem mass spectrometry. A liquid–liquid
extraction using an organic solvent mixture (methyl-tert-butyl-ether–hexane,
1:1, v/v) is used in place of solid-phase extraction because it provides consistent
recoveries for all analytes, including the internal standard. Retention times
for the analytes and internal standard are less than 7 min. Within- and between-day
precision is £ 6.5% and £ 13.3% relative standard deviation, respectively,
for the three analytes. The lower limits of quantitation are 0.25, 0.5, and
1.0 ng/mL for TNP-470, M-IV, and M-II, respectively.
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