2.3. Extraction of lipidsRices of e

2.3. Extraction of lipids

Rices of each cultivar (1 kg) were milled by a domestic miller (Zojirushi Ltd. BR-CA25 Osaka, Japan): well-milled rice (WMR) brans and half-milled rice (HMR) brans, respectively, before extraction. The total lipids from the brans (20 g) were extracted for 10 min at 0 °C with 300 ml chloroform/methanol (2:1, v/v), following the Folch procedure (Folch, Lee, & Sloane-Stanley, 1957). These solvents contained 0.01% butylated hydroxytoluene (BHT) to inhibit the oxidative degradation of lipids during analysis. Extraction was repeated thrice and then, 20 ml aqueous KCl (0.75%) were added to the combined extracts. After phase separation, the chloroform layer was withdrawn, dried over anhydrous Na2SO4, filtered, and the filtrate was concentrated under vacuum in a rotary evaporator at 35 °C. Solventless lipids were weighted to determine the lipid content of the rice brans and then transferred to a 25-ml brown-glass volumetric flask with chloroform/methanol (2:1, v/v).