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Antimicrobial Agents and Chemotherapy, November 1999, p. 2716-2719, Vol. 43, No. 11
Department of Clinical Pharmacology, School
of Pharmacy, Tokyo University of Pharmacy & Life Science, Tokyo
192-03, Japan,1 and Drug Research Unit,
Department of Clinical Pharmacy, School of Pharmacy, University of
California, San Francisco, California
94143-06222
Received 8 June 1998/Returned for modification 5 September
1998/Accepted 30 August 1999
Ribavirin is an antiviral agent used in the treatment of chronic
hepatitis C virus infection. One of the limitations associated with the
use of ribavirin is a reversible anemia caused by its accumulation in
erythrocytes. Therefore, it is of interest to determine ribavirin
levels in erythrocytes, as well as in plasma, as these measurements may
be predictive of hematotoxicity. In the present study, we describe a
high-performance liquid chromatographic (HPLC) assay for ribavirin in
whole blood to estimate concentrations of free ribavirin and
phosphorylated anabolites in erythrocytes. Since ribavirin exists
primarily as phosphorylated anabolites (mono-, di-, and triphosphates)
in erythrocytes, whole-blood extracts were initially dephosphorylated
with acid phosphatase. The enzyme-treated samples were subjected to
phenyl boronic acid column extraction for cleanup. The
purified fraction was analyzed by reversed-phase HPLC, which was
optimized for determination of ribavirin levels in whole blood. The
recoveries of ribavirin from whole blood ranged from 63.1 to 90.7% at
concentrations ranging from 1.67 to 40.0 µM. Intra- and interassay
variations estimated at these concentrations were 3.2 to 10.4 and 4.7 to 11.7%, respectively. This method was used to quantitate ribavirin
in samples both treated and untreated with acid phosphatase to estimate
the extent of intracellular phosphorylation in erythrocytes. The method
was also used to evaluate the effects of dipyridamole, a nucleoside
transporter inhibitor, on ribavirin disposition in erythrocytes in
in vitro experiments.
0066-4804/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.
High-Performance Liquid Chromatographic
Determination of Ribavirin in Whole Blood To Assess Disposition
in Erythrocytes
*
Corresponding author. Mailing address: Drug Research
Unit, Building 100, San Francisco General Hospital, 1000-1 Potrero
Ave., San Francisco, CA 94110. Phone: (415) 476-1148. Fax: (415)
476-0307. E-mail: aweeka{at}itsa.ucsf.edu.
Antimicrobial Agents and Chemotherapy, November 1999, p. 2716-2719, Vol. 43, No. 11
0066-4804/99/$04.00+0
Copyright © 1999, American Society for Microbiology. All rights reserved.
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