Selective Determination of Entecavir in the Presence of its Oxidative Degradate by Spectrophotometric and Chromatographic Methods.

Background: Entecavir (ENT) is an antiretroviral agent prescribed for the treatment of the hepatitis B virus(HBV) and human immunodeficiency virus(HIV).

Objective: Development and validation of three simple, sensitive, selective, and precise methods for determination of ENT in the presence of its oxidative degradation product (ENT deg.).

Method: The first method was based on second derivative (D2) spectrophotometry through measuring the peak amplitude of D2 spectra at 293.6 nm. The second one is mean centering of the ratio spectra (MCR), which enabled measurement of the peak amplitude at 280.0 nm. The third method was HPLC, where ENT was separated from ENT deg. using a Zobrax C18 column and methanol:water (30:70, v/v) with pH 3 as a mobile phase. The three developed methods were validated according to the International Conference on Harmonization guidelines.

Results: Linearity range of ENT was 5.00-50.00 μg/mL for both D2 and MCR. However, higher sensitivity was achieved using HPLC (1.00-50.00 μg/mL). Accuracy of ENT were 100.60 ± 0.547%, 101.55 ± 1.2071%, and 100.61 ± 1.207% for D2, MCR, and HPLC methods, respectively, and precision was within 1.280.

Conclusions: The developed methods were successfully applied for the determination of ENT in Tecavir® tablets without interference from ENT deg. They showed no significant difference in comparison with the official method and they can be applied in the quality analysis of ENT with high selectivity, accuracy, and precision.

Highlights: ENT was quantified using two spectrophotometric (D2 and MCR) methods and an HPLC method in presence of ENT deg. The proposed methods were applied to analysis of ENT tablets with high selectivity, sensitivity, and accuracy.