Determination of carotenoid pigments, retinol, and alpha-tocopherol in feeds, tissues, and blood serum by normal phase high performance liquid chromatography.
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abstract
A normal phase HPLC method employing a hexane:ethyl acetate mobile phase solvent gradient for the simultaneous separation and quantification of carotenoid pigments and vitamins A and E has been developed. The gradient consists of a 10-min linear change from 1 to 2 mL/min and 95:5 to 70:30 hexane:ethyl acetate mobile phase, which is followed by 7 min of isocratic mobile phase of 70:30 hexane:ethyl acetate at 2 mL/min. The stationary phase consisted of a 4-mu silica column. A photodiode array detector generated simultaneous chromatogram data at three wavelengths (294, 325, and 445 nm) with broad spectrum data at peaks. The method produced good separation of trans-lutein and zeaxanthin.