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 the separation of stigmasterol, campesterol and β-sitosterol in Prunus spinosa L. (blackthorn) fruit extracts by HPLC system. Samples were prepared by Soxhlet extraction method and separated on a C18 column using acetonitrile-methanol mobile phase and photodiode array detector (PDA). The optimized method resulted in a linear calibration curve ranging from 1.70–130 μg mL–1 for all three phytosterols. Analyses of external phytosterol standards showed good linearity (R2 of 0.998 to 0.999); LOD and LOQ were determined to be 0.32–9.30 μg mL–1 and 0.98–28.1 μg 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.