High-performance liquid chromatographic analysis of 2'-fluoro-2',3'-dideoxyadenosine and 2'-fluoro-2',3'-dideoxyinosine in dog plasma and urine.

A high-performance liquid chromatographic assay was developed and validated for a simulataneous determination of 2'-fluoro-2' 3'-dideoxyadenosine (FddA) and its metabolite, 2'-fluoro-2',3'-dideoxyinosine (Fddl) in dog plasma and urine. In vitro, FddA and Fddl exhibit activity against human immunodeficiency virus (HIV). A solid phase extraction was applied to extract FddA, Fddl, and the internal standard (IS; 3',5'-anhydrothymidine) from the biomatrices. The processed samples were chromatographed using a C8 column coupled with a mobile phase consisting of monobasic phosphate, dibasic phosphate, ethylene glycol monomethyl ether, and water. Detection was performed at 257 nm. The nominal retention times were 9, 14, and 26 min for Fddl, IS, and FddA, respectively. The lower limits of quantitation were 0.1 and 2.0 micrograms/mL in plasma and urine, respectively, for both analytes. The accuracy of the assay deviated < or = 10% from the nominal concentrations, and the precision was < or = 14% 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 extraction recoveries of the analytes were greater than 80%. The application of this assay was demonstrated in a preliminary pharmacokinetic study of FddA and Fddl in dogs. Two male dogs per dose level received a 100, 250, or 500 mg/kg oral dose of FddA once daily for 14 days. The early appearance of Fddl in plasma (0.25 h; the first sampling time) and greater plasma levels of Fddl than FddA (> 50-fold of Cmax), suggested that the conversion of FddA to Fddl was rapid and extensive. Renal excretion appeared to be the major route of elimination of Fddl.

Pharmacokinetics of 2',3'-dideoxyadenosine in dogs.

The pharmacokinetics of 2',3'-dideoxyadenosine (ddAdo) and 2'-3'-dideoxyinosine (ddIno) were determined after intravenous bolus administration and long-term intravenous infusion of ddAdo in dogs. ddAdo was rapidly deaminated to ddIno and ddAdo plasma concentrations were only a fraction of ddIno concentrations. The total body clearance of ddAdo exceeded the literature value for the cardiac output of the dog, indicating an extremely rapid metabolism, and the existence of extrahepatic metabolism. Urinary excretion of unchanged ddAdo was a minor route of elimination (approximately 1%). The pharmacokinetics of ddIno was determined assuming complete conversions of ddAdo to ddIno. ddIno elimination was dose-dependent with total body clearance ranging from 4 to 55 ml/min/kg in individual animals. The plasma half-life was approximately 30 min after most routes of administration, but increased to approximately 60 min in two animals receiving a large intravenous dose of 500 mg/kg. ddIno penetrated into the cerebrospinal fluid to a limited extent, reaching concentrations of 3-11% of those in plasma. Urinary excretion of unchanged ddIno accounted for approximately 20% of the administered dose of ddAdo, while uric acid and hypoxanthine were minor urinary metabolites. Concentrations exceeding the in vitro minimal viral inhibitory concentration (2.4 micrograms/mL) could be safely maintained in plasma for a 10-day period. Infusions which gave cerebrospinal fluid concentrations of 12 to 17 micrograms/mL resulted in dose limiting myelosuppression and intestinal toxicity, after less than 10 days of infusion. Orally administered ddAdo was absorbed as ddIno, with bioavailabilities ranging from 28 to 93% in experiments where no emesis occurred. These studies indicate the rapid in vivo conversion of ddAdo to ddIno, and support the selection of ddIno over ddAdo for further drug development.