A Bioanalytical Method Development and Validation for Simultaneous Determination of Velpatasvir and Sofosbuvir in Spiked Human Plasma

Abstract

INTRODUCTION Sofosbuvir ( Fig. 1) is a prodrug of 2'-deoxy-2'-fluoro-2'-C-methyl-uridine monophosphate that is phosphorylated intra cellularly to the active triphosphate form, used for the treatment of chronic hepatitis C. The nucleoside triphosphate is a nonobligate chain-terminating analogue of UTP that competes for incorporation at the HCV NS5B polymerase active site. Viral RNA synthesis is inhibited secondary to incorporation of the phosphorylated metabolite into nascent viral RNA by the HCV RNA-dependent RNA polymerase [1]. Velpatasvir (Fig. 1) A simple, precise, accurate and sensitive, isocratic RP-HPLC method has been developed and validated for the estimation of velpatasvir and sofosbuvir in plasma. Method was developed with, Intersil ODS C18 column (250 mm × 4.6 mm × 5 µ), mobile phase mixture of acetonitrile and water having pH of 6.5 in the ratio of (80:20 v/v) at flow rate 1 mL/min. at UV detector wavelength of 240 nm. Mobile phase pH was adjusted with 1 % o-phthalaldehyde (OPA). The retention times are 3.49 min for sofosbuvir and 6.15 min for velpatasvir, Quantitative linearity was obeyed in the concentration range of 0.1 to 0.8 µg/mL for sofosbuvir and 0.025 to 0.2 µg/mL for velpatasvir. The limit of detections are 0.006 µg/mL for sofosbuvir, 0.0017 µg/mL for velpatasvir and limit of quantifications are 0.0125 µg/mL for sofosbuvir, 0.003 µg/mL for velpatasvir for which indicates the sensitivity of the method. The average recovery of sofosbuvir is 100.75 and 99.25 % for velpatasvir. The high percentage recovery indicates that the proposed method is highly accurate. Bench top, Auto sampler, Freeze and Thaw stability test are performed.

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“…The literature survey revealed that there are few stability indicating RP-HPLC 5,6,7,8,9,10 and UV 11 spectrophotometric methods are available for the simultaneous estimation of Sofosbuvir and Velpatasvir in tablet dosage form. Few analytical methods for Sofosbuvir and other HCV drugs in biological fluids by UPLC-MS/MS have been reported 12,13,14,15,16 . However, a stability indicating UPLC method was not available.…”