RP-HPLC Method for Estimating Norethindrone in Plasma and Tissues Following Administration of a Controlled Release Nanoparticulate Liquid Formulation

Abstract

The purpose of this investigation was to develop and validate a simple reversed phase-high-performance liquid chromatography method coupled with UV detector estimating norethindrone in plasma and tissues. A Thermo Scientific C18 column (250×4.6 mm ID, 5 μm pore size) and a mobile phase consisting of deionized water:acetonitrile (60:40, v/v) were used. The flow was isocratic at a rate of 1.3 ml/min and the wavelength of detection was 245 nm. Estradiol was used as internal standard. Validation of linearity, accuracy and precision, limits of detection and quantification, specificity and recovery was carried out according to the International Conference of Harmonization guidelines. The method was used in estimating the bioavailability of a controlled release nanoparticulate liquid formulation, administered to dogs. It was also used estimating adhesion potential of the same formulation to the GIT of mice. The method was simple and only liquid-liquid extraction was used in case of plasma samples. However, sample preparation was more complex in the case of tissue samples. It was linear in the range studied, accurate and precise. It was specific and the excipients used in preparing the formulation did not interfere with the method. The recovery was high and consistent. The bioavailability of the drug was enhanced after the administration of the nanoparticulate liquid formulation as compared to unformulated drug. The formulation adhered to the stomach and intestines for 48 h. After 48 h the concentration reached undetectable levels. In conclusion, a simple, linear, accurate and precise reversed phase-high-performance liquid chromatography method coupled with UV detection was developed, validated, and used successfully for the determination of norethindrone in plasma and adhesion to GIT tissues. The cost of the analysis was expected to be low sine the extraction procedure was simple and no radiolabeled internal standard was useds.