A New HPLC Method for the Simultaneous Analysis of Plasma Retinol, Tocopherols, and Carotenoids and its Application in Cancer Epidemiology
An isocratic high-performance liquid chromatography (HPLC) method for the simultaneous determination of various fat-soluble vitamins and carotenoids has been developed. The method utilizes a Radial-Pak C18, 8 mm i.d. × 10 cm, 5-µm column and an elution solvent composed of methanol, acetonitrile, and chloroform. Retinol, α-tocopherol, α-carotene, β-carotene, lycopene, zeaxanthin, and two other unidentified carotenoids can be clearly separated and quantified in one HPLC run using α-tocopheryl acetate or tocol as the internal standard. The eluted peaks are quantified by either a photodiode-array detector at preprogrammed wavelengths at the absorption maxima of the compounds or by a dual-wavelength detector at 280 and 436 nm. The total run time is 16 min. With an automatic injector and a programmable detector the system allows unattended operation. The within-run and day-to-day coefficients of variation range from 1 to 8%. In addition, the system can monitor the absorption spectra of the eluent during the HPLC run; this allows the spectral identification of various compounds separated in the same run. The application of this new method in cancer epidemiology was demonstrated in a joint U.S.-China cooperative study on nutrition and esophageal cancer.
KeywordsHPLC Chloroform Lipase Carotenoid Palmitate
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