Published:Journal of Chromatographic Science,
ISSN 0021-9665Volume
41, Number 5, May/June 2003, pp 255-260
A Sensitive and Specific Method for the Determination of Total
Ribavirin in Human Red Blood Cells by Liquid Chromatography–Tandem Mass
Spectrometry
Li-Tain Yeh, Mai Nguyen, and Chin-Chung Lin
Drug Development Department, Ribapharm Inc., 3300 Hyland Avenue, Costa Mesa,
CA 92626
A sensitive and specific method using high-performance liquid
chromatography (LC)–tandem mass spectrometry (MS) for the analysis of
total ribavirin in human red blood cells (RBC) is developed and validated.
The method involves the addition of an internal standard and perchloric acid,
the conversion of ribavirin phosphorylated metabolites to ribavirin, purification
with a solid-phase exchange cartridge, and LC–MS–MS analysis. The
MS–MS is selected to monitor m/z 245–113 for ribavirin and m/z
250–113 for [13C]ribavirin using positive electrospray ionization. The
calibration curve is linear over a concentration of 100–10,000 ng/mL
with a limit of quantitation of 100 ng/mL. Mean interassay accuracy for quality
control (QC) at 100, 1000, and 10,000 ng/mL are 101.8%, 99.4%, and 98.8%, respectively.
Mean interassay precision (%CV) for QC at 100, 1000, and 10,000 ng/mL are 5.0%,
5.0%, and 2.5%, respectively. Extractibility of total ribavirin from RBC is
confirmed with RBC obtained from a [14C]ribavirin-dosed monkey. The method
is used to determine the free and total ribavirin concentration in human RBC
obtained from hepatitis C patients treated with ribavirin.
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