Carotenoids in the extract were analyzed by high performance-liquid chromatography (HPLC) equipped with STR-ODS-II column (5 lm; 4.6 mm 250 mm)(Nacalai Tesque, Inc., Japan) and UV–visible detector (UV-970, Jasco, Japan) as reported by Machmudah et al.(2008a). A mixture of 2-propanol:acetonitrile:methanol:water in 52:39:5:4 v/v ratio was used as a mobile phase t a flow rate of 1.2 mL/min. The sample dissolved in the solvent mixture, consisting of 1:2 chloroform:acetone, was injected in 20 ll units. The column temperature was controlled at 30 C by a column heater (Sugai U- 620, Japan) and detection was measured at 450 nm. Detector signals were recorded using BORWIN Chromatography software. Peaks of lycopene and b-carotene were identified by comparing the retention time in the extract with standard compounds. The content of lycopene and b-carotene in the extract were estimated by comparing the peak areas with their respective standards. Calibration curves of the standards were prepared by plotting standard concentrations (0.25–4 mg/mL) versus their peak areas detected at 5.7 and 8.9 min for lycopene and b-carotene, respectively. All samples were analyzed in duplicate. The recovery of lycopene and b-carotene in the extract at each extraction condition is cited as mean ± standard deviation.