HIGH-PERFORMANCE LIQUID-CHROMATOGRAPHIC ANALYSIS OF 2'-FLUORO-2',3'-DIDEOXYADENOSINE AND 2'-FLUORO-2',3'-DIDEOXYINOSINE IN DOG PLASMA AND THE URINE

A high performance liquid chromatographic assay was developed and vali dated for a simultaneous determination of 2'-fluoro-2',3'-dideoxyadeno sine (FddA) and its metabolite, 2'-fluoro-2',3'-dideoxyinosine (FddI) in dog plasma and urine. In vitro, FddA and FddI exhibit activity agai nst human immunodeficiency virus (HIV). A solid phase extraction was a pplied to extract FddA, FddI, and the internal standard (IS; 3',5'-anh ydrothymidine) from the biomatrices. The processed samples were chroma tographed using a C8 column coupled with a mobile phase consisting of monobasic phosphate, dibasic phosphate, ethylene glycol monomethyl eth er, and water. Detection was performed at 257 nm. The nominal retentio n times were 9, 14, and 26 min for FddI, IS, and FddA, respectively. T he lower limits of quantitation were 0.1 and 2.0 mu g/mL in plasma and urine, respectively, for both analytes. The: accuracy of the assay de viated less than or equal to 10% from the nominal concentrations, and the precision was less than or equal to 44% coefficient of variation, In either matrix, both analytes were stable for at least three freeze- thaw cycles and in the injection media for at least 54 h. The extracti on recoveries of the analytes were greater than 80%. The application o f this assay was demonstrated in a preliminary pharmacokinetic study o f FddA and FddI in dogs, Two male dogs per dose level received a 100, 250, or 500 mg/kg oral dose of FddA once daily for 14 clays. The early appearance of FddI in plasma (0.25 h; the first sampling time) and gr eater plasma levels of FddI than FddA (>50-fold of C-max), suggested t hat the conversion of FddA to FddI was rapid and extensive. Renal excr etion appeared to be the major route of elimination of FddI.