title: A rapid HPLC column switching method for sample preparation and determination of β-carotene in food supplements.
abstract: A simple and automated HPLC column-switching method with rapid sample pretreatment has been developed for quantitative determination of β-carotene in food supplements. Commercially samples of food supplements were dissolved in chloroform with help of saponification with 1M solution of sodium hydroxide in ultrasound bath. A 20-min sample dissolution/extraction step was necessary before chromatography analysis to transfer β-carotene from solid state of food supplements preparations (capsules,tablets) to chloroform solution. Sample volume - 3μL of chloroform phase was directly injected into the HPLC system. Next on-line sample clean-up was achieved on the pretreatment precolumn Chromolith Guard Cartridge RP-18e (Merck), 10×4.6mm, with a washing mobile phase (methanol:water, 92:8, (v/v)) at a flow rate of 1.5mL/min. Valve switch to analytical column was set at 2.5min in a back-flush mode. After column switching to the analytical column Ascentis Express C-18, 30×4.6mm, particle size 2.7μm (Sigma Aldrich), the separation and determination of β-carotene in food supplements was performed using a mobile phase consisting of 100% methanol, column temperature at 60°C and flow rate 1.5mL/min. The detector was set at 450nm. Under the optimum chromatographic conditions standard calibration curve was measured with good linearity - correlation coefficient for β-carotene (r(2)=0.999014; n=6) between the peak areas and concentration of β-carotene 20-200μg/mL. Accuracy of the method defined as a mean recovery was in the range 96.66-102.40%. The intraday method precision was satisfactory at three concentration levels 20, 125 and 200μg/mL and relative standard deviations were in the range 0.90-1.02%. The chromatography method has shown high sample throughput during column-switching pretreatment process and analysis in one step in short time (6min) of the whole chromatographic analysis.
authors.affiliation: Department of Analytical Chemistry, Faculty of Pharmacy, Charles University, Heyrovského 1203, Hradec Králové 500 05, Czech Republic.
authors.forename: Ivana
authors.initials: I
authors.lastname: Brabcová
authors.forename: Markéta
authors.initials: M
authors.lastname: Hlaváčková
authors.forename: Dalibor
authors.initials: D
authors.lastname: Satínský
authors.forename: Petr
authors.initials: P
authors.lastname: Solich
mesh_terms: D002851:Chromatography, High Pressure Liquid / Q000295:instrumentation / Q000379:methods*; D019587:Dietary Supplements / Q000032:analysis*; D012680:Sensitivity and Specificity; D052616:Solid Phase Extraction; D019207:beta Carotene / Q000032:analysis* / Q000302:isolation & purification
chemical_list: D019207:beta Carotene