Determination of retinol and carotenoids in selected Malaysian food products using high-performance liquid chromatography (HPLC)

The presented method is focused on simultaneous determination of retinol and two carotenoids (lutein, β-carotene) in selected Malaysian food products. A Dionex Ultimate 3000 Series High-Performance Liquid Chromatography system equipped with two Dionex Acclaim Polar Advantage II C18 columns was utilized in this study. Simultaneous elution of retinol, lutein and β-carotene was achieved within 45 min with gradient profile of methanol and ultra-pure water (95:5–100:0–95:5) programmed at 0.8 mL/min flow rate and 30 °C. Retinol was acquired at 325 nm while lutein and β-carotene were acquired at 440 nm. Limits of detection and quantification of retinol and lutein were at 0.10, 0.34 µg/mL, and 0.08, 0.26 µg/mL; respectively while higher levels were observed for β-carotene at 1.06 and 3.54 µg/mL. Linearity test indicated R2 of 0.996–0.999, along with high accuracy (71.92–116.29%) and excellent repeatability for all analytes. Extraction efficiency was determined using standard addition into representative food product with recovery obtained in the range of 61.86–116.90% for all three analytes. The developed in-house HPLC method incorporates an enclosed solid-phase extraction system that is believed to further improve sample purification and analytes detection. On the whole, the developed on-line SPE-HPLC technique and efficient sample pre-treatment is deemed a reliable approach for simultaneous determination of retinol, lutein and β-carotene, as analyzed among the selected Malaysian food products. Additionally, the described procedure could also benefit routine analysis of these analytes in other food matrices or for determination of other analytes with similar chemistry and faster elution.