"New Chromatographic Analytical Conditions for the Separation and Determination of a Standard Mixture of Phytosterols"

Abstract

This research aims to develop a reverse-phase high-performance liquid chromatography (RP-HPLC) method and validate its suitability for the separation and identification of a mixture of phytosterols, which are key components of food and herbal products known for their beneficial effects in managing hyperlipidemia. Chromatographic analysis was performed using a Thermo Hypersil™ RP C8 HPLC column (250 × 4.6 mm I.D.) and 5 µm particle size. A mixture of (Acetonitrile:Water) in a ratio of (80:20)% was used as the mobile phase under isocratic elution conditions, with a flow rate of 1.8 mL/min at a temperature of 45°C and 20 μL as an injection volume. The chromatograms were recorded at 195 nm using Diode Array Detector (DAD). Validation of the method was evaluated based on linearity, specificity, limit of detection (LOD), and limit of quantification (LOQ), according to the guidelines of the International conference on Harmonization (ICH). This reliable method ensures high accuracy and a rapid sample preparation process for the precise determination of sterol content in food products, herbal formulations, and vegetable oil, facilitating quality, safety, and efficacy monitoring, all within an analysis time of no more than 23 minutes.