RP-LC Method for the Determination of Seven Antiviral Drugs and Bioanalytical Application for Simultaneous Determination of Lamivudine and Penciclovir in Human Plasma

Abstract

An RP-LC method was developed and validated for separation and determination of seven antiviral drugs in their drug substances and drug products, using a BDS Hypersil C8 column at ambient temperature. The mobile phase was composed of 0.01 M KH2PO4 (pH 4):acetonitrile in gradient mode and the quantification was achieved at 245 nm for acyclovir, ganciclovir, penciclovir and valacyclovir hydrochloride, and at 220 nm for lamivudine, oseltamivir phosphate and ribavirin. Run time was 8 min, except for the oseltamivir-containing mixture where the run time was extended to 20 min. The validation of the method was assessed according to ICH guidelines. Linearity, accuracy and precision were satisfactory over the concentration ranges 3–100 µg mL−1 for acyclovir, ganciclovir and penciclovir; 5–100, 4–100, 7–120 and 5–120 µg mL−1 for valaciclovir HCl, lamivudine, oseltamivir phosphate and ribavirin, respectively. The developed method is suitable for the quality control and routine analysis of the cited drugs separately or in combinations. Moreover, the proposed method can be used for counterfeit drug detection. Also, simultaneous determination of lamivudine and penciclovir in spiked human plasma has been developed, using valaciclovir hydrochloride as an internal standard. Protein precipitation using perchloric acid solution was used for the extraction of the drugs from plasma. With modified chromatographic conditions, quantification was achieved at 254 nm. Validation of the method was performed according to European Medicines Agency guidelines with linearity over the range 0.056–4.0 and 0.14–10.0 μg mL−1 for lamivudine and penciclovir, respectively.