Development of LC-MS Method for Determination of Ursolic Acid: Application to the Analysis of Ursolic Acid in Staphylea Holocarpa Hemsl

Ursolic acid is a hydroxy pentacyclic triterpene, which has a chemoprotective activity in human. A reliable and reproducible liquid chromatography-mass spectrometric assay (LC-MS) was developed for the determination of ursolic acid in laboratory-made mixtures and in leaves and twigs extracts of Staphylea holocarpa Hemsl. The methanolic solution of the extracted ursolic acid was chromatographically analyzed using Shim Pack CLC-CN, C18 (150 x 6 mm, 5 mu) column and a mobile consisting of methanol-1% acetic acid solution (4:1) at a flow rate of 1.0 ml min(-1). The mass spectrometer (LCQ-Finnigan) was programmed in the positive single ion monitoring (SIM) to permit detection and quantitation of ursolic acid in MS-SIM mode at m/z 439.2, 411.2 and 390.9. Linear correlation (r > 0.99) of the peak area and the concentration of ursolic acid over the concentration range 0.25-10 microg ml(-1) was obtained. The relative standard deviation (%R.S.D.) and percentage deviation from the nominal concentrations (%DEV) were found to be 3.03-3.59% and -4.5 to +6.2%, respectively. Analysis of laboratory-made mixtures containing known concentrations of ursolic acid, as quality control samples, gave a mean recovery percentage of 97.8%. Application of the proposed method for the analysis of leaves and twigs extracts of S. holocarpa Hemsl. gave mean percentage contents of ursolic acid of 0.95 and 0.25%, respectively.