CHARACTERIZATION OF MAJOR CAROTENOIDS IN WATER CONVOLVULUS (IPOMOEA-AQUATICA) BY OPEN-COLUMN, THIN-LAYER AND HIGH-PERFORMANCE LIQUID-CHROMATOGRAPHY

The major carotenoids present in water convolvulus (Ipomoea aquatica) were characterized by open-column chromatography, thin-layer chromatography (TLC), and high-performance liquid chromatography (HPLC). A 1:1 mixture of activated magnesium oxide and diatomaceous earth was used as the major adsorbent to separate carotenes, monohydroxy, dihydroxy and polyoxy pigments by open-column chromatography. Carotenes and cryptoxanthin were eluted with hexane-acetone at 96:4 and 90:10, respectively. Lutein, violaxanthin and neoxanthin were eluted with hexane-acetone-methanol at 85:15:0.2, 85:15:0.2 and 85:15:1.5, respectively. The elution sequence of lutein and violaxanthin was dependent on the amount of methanol present. A lutein band containing lutein and lutein epoxide was further separated by TLC. An HPLC isocratic solvent system of acetonitrile-methanol-ethyl acetate (75:15:10) was found to be appropriate for determining the reproducibility of retention time with respect to separated bands obtained by open-column chromatography. Each band was identified by comparing the absorption spectra and retention time with reference standards. The major carotenoids present in water convolvulus were beta-carotene, lutein, lutein epoxide, violaxanthin and neoxanthin. The amount of each major carotenoid was also determined.