Development and validation of HPLC/UV-procedure for efavirenz quantitative determination

Abstract

Efavirenz is a synthetic compound from the group of non-nucleoside reverse transcriptase inhibitors used for treatment of HIV infection. There are cases of acute poisoning due to administration of efavirenz, including cases of suicide attempts. The aim is to develop HPLC/UVprocedure of efavirenz quantification using the system of HPLC-analyzer MiLiChrome® A-02. HPLC microcolumn of Ø2×75 mm dimension and reversed phase ProntoSIL 120-5-C18 AQ, 5 μm were used as an analytical column. Eluent A (0.2 M LiClO4 – 0.005 M HClO4) and Eluent B (acetonitrile) were used as the mobile phase components. The mobile phase was run in gradient elution mode, namely from 5% to 100% Eluent B for 40 min, then 100% Eluent B for 3 min. All analysis was carried out at 40°С and flow rate of 100 μl/min. Detection was performed at 247 nm. The specificity of the used chromatographic conditions has been confirmed in relation to other antiretroviral medicine. Retention time for efavirenz is 11.95 min. Ethanol has been proposed for preparation of efavirenz solutions. To prove the possibility of the proposed procedure application in further analysis its step-by-step validation has been carried out in the variants of the method of calibration curve, method of standard and method of additions. Such validation parameters as in process stability, linearity/calibration model, accuracy and precision (repeatability) have been estimated by model solutions. New procedure of efavirenz quantitative determination by the method of HPLC/UV has been developed; its acceptability for application has been shown