Validation of an Isocratic HPLC Method for Simultaneous Estimation of Major Phytosterols in Prunus spinosa L. extracts

Abstract

This study aimed to develop a rapid method for separation of stigmasterol, campesterol and β-sitosterol in Prunus spinosa L. (sloe) fruit extracts by High Performance Liquid Chromatography system. Samples were prepared by Soxhlet extraction method and separated on a high strength silica C18 column using acetonitrile-methanol mobile phase and Photodiode Array Detector. The optimized method resulted in a linear calibration curve ranging from 1.7 ng mL^-1 to 130 ng mL^-1 for all three phytosterols. Analyses of internal and external phytosterol standards showed good linearity (R² of 0.998 to 0.999); LOD and LOQ were determined to be 2.33×10^-7–2.18×10^-4 and 7.07×10^-7–6.60×10^-4 mg mL^-1, respectively. Repeatability and reproducibility precision analyses showed acceptable values of RSD %. β-sitosterol was the predominant phytosterol (51.53–81.03 % of total) among all samples. Method validation parameters indicated that this analytical method can be applied for accurate and precise determination of campesterol, stigmasterol and β-sitosterol, in selected extracts.