Development and validation of an HPLC method for determination of oleanolic acid content and partition of oleanolic acid in submicron emulsions.

The aim of this study was to develop and validate a simple HPLC method for the quantitative determination of the oleanolic acid (OA) content and partition coefficient of OA in a submicron emulsion-based formulation (SE-OA). A Diamonsil C18 (150 mm x 4.6 mm, 5 microm) column was eluted with a mobile phase consisting of methanol/water (95:5, v/v). The analyses were performed at 35 +/- 1 degrees C with a flow rate of 1.0 mL/min and variable wavelength detector (VWD) at 210 nm. The calibration curve was linear over a concentration range of 2-100 microg/mL with a correlation coefficient of 0.999. The LOD and LOQ were 0.1 and 1 microg/mL, respectively. The individual spike recovery of OA ranged from 99.88 to 100.28%. The percent relative standard deviations (% R.S.D.) of intra-day and inter-day analyses were less than 3.1%. The validation results confirmed that the method is specific, linear, accurate, precise, robust and sensitive for its intended use. The present method was successfully applied to the determination of the OA content and partition coefficient of OA in SE-OA during the early stage of formulation development.