Determination of egg yolk xanthophylls by isocratic high-performance liquid chromatography

An isocratic HPLC method was developed for the determination of eight xanthophylls (lutein, capsanthin, zeaxanthin, canthaxanthin, beta-apo-8'-carotenal, ethyl-8'-apo-beta-carotene-8'-oate, citranaxanthin and beta-cryptoxanthin; registered as additives in poultry feeding) in egg yolks. Optimum separation of all-Eisomers of these xanthophylls was achieved in less than 18 min on a ProntoSIL C-30 column at 27 degrees C using acetone-methanol-0.5 M triethylammonium acetate buffer pH 7 14:5:1 (v/v) as the mobile phase with a flow rate of 1 mL/min using spectrophotometric detection at 450 nm. Other mobile phases were also found suitable, including acetone-water 93:7 (v/v) and acetone-methanol 1:4 (v/v) and the influences of column temperature on the separation and addition of triethylammonium acetate buffer pH 7 to the mobile phase on enhancement of the peak areas were evaluated. Preparation of test solution from yolks included a short vortexing of 0.5 g of yolk in 10 mL of acetone, followed by 15 min magnetic stirring under nitrogen and centrifugation. The method was validated for 5 analytes. The calibration range was between 0.04 and 2 mu g/mL and the mean recovery of the analytes (95%) and the intra-day precision of the method (RSD less than 5%) on three levels in triplicates were obtained. Analyses of eggs from four husbandry classes showed the presence of up to four xanthophylls (lutein, zeaxanthin, canthaxanthin and ethyl-8'-apo-beta-carotene-8'-oate) and traces of beta-cryptoxanthin. (C) 2013 Elsevier B.V. All rights reserved.