z Extraction and Determination of γ-oryzanol and α-tocopherol: The experimentation method modified from [24] and [26] involved 1 g of soaking powder of rice formulas accepted by consumers (before and after cooking) in acetone at a ratio of 1:10 (w/v). The mixture was then spun with a centrifuge machine at maximum speed for 1 min and at 2,500 rpm for 20 min. Supernatants will be obtained after the solvent has been separated. Supernatants were then dehydrated under nitrogen.
Measurement of γ-oryzanol and α-tocopherol levels by the HPLC method was performed by dissolving rough extracts in the mobile phase and filtering by using a syringe-driven filter with a diameter of 0.45 μm. Analysis was performed using a RP-HPLC system machine (Shimadzu). 20 μl of the sample were injected through the security guard-column. Column size was 4.60 × 250 mm, 4 μl. Column temperature was controlled at 45°C. The mobile phase solvent, acetonitrile/methanol (25:75, v/v), was used at a flow rate of 1.5 ml/min. The wavelength was detected at 292 nm for measuring α-tocopherol levels and 325 nm for measuring γ-oryzanol levels.
2.2.3. Determination of Antioxidant Activity
2.2.3.1 In Vitro
2.2.3.1.1. Free-Radical Scavenging Activity on DPPH•
The DPPH method tested ability to scavenge DPPH• radicals. The experimentation method modified from [24] and [27] mixed 100 μl in a volume of rough extract from rice with 0.1 mM of concentrated DPPH• solution at a volume of 1.9 ml. The obtained mixture will be fermented in a dark space at room temperature