One-step equilibrium direct-solvent extraction was conducted by the method of Butsat and Siriamornpun (2010) with some modifications. Each sample (1 g) was extracted with acetone at a ratio of 1:10 w/v, vortexed at maximum speed for 1 min then centrifuged at 2500 rpm for 20 min, after which the solvent was removed. The residual was further extracted twice, and the supernatants were combined before evaporating them to dryness
under nitrogen gas. The determinations were made in triplicate. The contents of c-oryzanol and tocopherols were determined using HPLC. The crude extracts were dissolved in the mobile phase and filtered through a 0.45 lm pore size syringe-driven filter. The RP-HPLC system (Shimadzu) consisted of an auto sampler and column oven equipped with Inertsil ODS (4.6 mm 250 mm, 5 lm) with mobile phase of acetonitrile/methanol (25:75, v/v), flow rate 1.5 mL/min and photodiode-array detector at 292 nm for the analysis of tocopherols and at 325 nm for the analysis of c-oryzanol. Calibration curves were constructed with the external standards.