Rapid quantitation of (−)-2′-deoxy-3′-thiacytidine in human serum by high-performance liquid chromatography with ultraviolet detection |
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Authors: | Xiao-Jian Zhou Jean-Pierre Sommadossi |
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Affiliation: | Department of Pharmacology and Toxicology, Division of Clinical Pharmacology, The Liver Center, Center for AIDS Research, University of Alabama at Birmingham, 1670 University Boulevard, VH 019G, Birmingham, AL 35294-0019, USA |
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Abstract: | A rapid, sensitive and specific high-performance liquid chromatographic (HPLC) assay was developed and validated for the measurement of (−)-2′-deoxy-3′-thiacytidine (3TC) in human serum. The method included precipitation of serum proteins by trichloroacetic acid (20%, w/v) treatment followed by centrifugation. The resulting supernatant was directly injected and 3TC was isocratically chromatographed on a reversed-phase C18 column using a mixture of phosphate buffer and methanol (88.3:11.7, v/v) and monitored at 280 nm. The limit of quantitation was 20 ng/ml using 100 μl of serum. The standard curve was linear within the range of 20–10 000 ng/ml. Replicate analysis of three quality control samples (40–1500 ng/ml) led to satisfactory intra- and itner-assay precision (coefficient of variation from 3.0 to 12.9%) and accuracy (deviation from −6.3 to 9.7%). Moreover, sample treatment processes including human immunodeficiency virus (HIV) heat-inactivation, exposure at room temperature and freezing-thawing cycles did not influence the stability of the analyte. This assay was successfully applied to the determination of 3TC serum levels in HIV-infected patients. In addition, preliminary results indicated that this procedure may also be extended to the measurement of 3TC in human plasma and urine. |
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Keywords: | (− )-2′ -Deoxy-3′ -thiacytidine |
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