Cost-effective green chromatographic method for the simultaneous determination of four commonly used direct-acting antiviral drugs in plasma and various pharmaceutical formulations

Abstract

This article presents the simultaneous determination of multiple recently approved direct-acting antiviral (DAA) regimens using a sensitive, robust method. Reversed-phase high-performance liquid chromatographic method (RP-HPLC) has been developed to represent a green alternative of other reported methods for the simultaneous determination of the most common four DAAs used for chronic hepatitis C treatment, including sofosbuvir (SF), ledipasvir (LD), daclatasvir (DC), velpatasvir (VP) and tenofovir (TO) as internal standard (IS) in bulk, pharmaceutical dosage form and plasma samples. This method was performed with an environment-friendly mobile phase with the least amount of energy consumption and waste generation. Chromatographic conditions have been optimized in order to obtain well-resolved symmetrical peaks within a reasonable elution time. Separation was achieved using Waters XBridge BEH C18 Column (4.6 × 150 mm) using isocratic elution mode at a flow rate 0.8 mL.min−1 with the mobile phase 0.1% (v/v) triethylamine acidified water with ortho-phosphoric acid (OPA) pH 2.53 and acetonitrile in the ratio (70:30 v/v). Sample extraction was done using diethyl ether for the analyte and IS extraction from the plasma samples. The suggested method was validated within the concentration range (0.4–20 μg.mL−1) for all the studied drugs according to the ICH guidelines. Calibration plots showed a linear relationship for SF, LD, DC and VP with correlation coefficients ≥ 0.9995 and acceptable recoveries. The method has been applied for the analysis of the studied drugs in plasma samples and pharmaceutical dosage forms without any matrix interference and with % RSD less than 2%, indicating the method to be precise. The suggested method was also assessed using the analytical Eco-scale tool and Green Analytical Procedure Index (GAPI) by comparing the method with other reported methods in terms of sample preparation, consuming hazardous reagents, instrument energy consumption and waste generation. Eco-scale and GAPI results proved the greenness of our proposed method. Moreover, the proposed method has the advantage of analyzing several pharmacologically related drugs in the same run simultaneously, which allows the analysis of alternative therapeutic formulations under the same running conditions. Thus, much effort and time is saved for developing a specific method for every pharmaceutical mixture. This promotes the proposed method to be used for both quality control analysis and any bioavailability study for the simultaneous determination of any of the four antiviral drugs in the pharmaceutical formulation or plasma samples without any harmful effect on the environment or human health.