HPLC determination of acyclovir in human serum and its application in bioavailability studies

Abstract

A sensitive, accurate and rapid reverse phase HPLC method was described to quantitate levels of acyclovir in human plasma. The drug, internal standard (metronidazole) and phosphate buffer (0.05 M) were added to serum samples and vortexed for 30 sec. A mixture of isopropyl alcohol:dichloromethane (60:40) was then added and vortexed for 3 min. Samples were centrifuged and the supernatant layer was separated, evaporated to dryness under nitrogen gas stream, reconstituted in mobile phase and, an aliquot of 50 μl was analyzed on a μ-bondapack C₁₈ (250 × 3.9 mm) column, with 3% acetonitrile in deionised water and 0.5% orthophosphoric acid, (pH 2.5) at 254 nm. The standard curve covering 100-1500 ng/ml concentration range, was linear, relative errors were within 0.79 to 17.4% and the CV% ranged from 3.81 to 18.2. The limits of quantitation and detection of the method were 100 ng/ml and 25 ng/ml, respectively. The method was suitable for bioavailability and pharmacokinetic studies of acyclovir in humans and applied in a randomized, two-way cross over bioequivalence study of two different acyclovir preparations with twelve subjects and with a one-week washout period.