Studies on Respiratory Enzymes in Rice Kernel

Glutathione reductase as an acidic flavoprotein, has been isolated from the acidic protein fraction of rice embryos and purified by procedures involving ammonium sulfate fractionation, gel filration on Sephadex G–75 and G–100, ion exchange chromatography on CM-and DEAE-Sephadex and finally hydroxylapatite column chromatography. The preparation was homogeneous when examined by ultracentrifugation and almost pure on polyacrylamide gel electrophoresis. The flavoprotein exhibited an absorption spectrum characteristic of glutathione reductase having absorption maxima at 275, 370, 379, and 463 mμ with a clear double peak between 370 and 380 mμ and shoulders at around 430 and 490 mμ. The absorption ratio of A₂₇₅/A₄₆₃ and A₄₆₃/A₃₇₉ were 8.15 and 1.06, respectively. The purified enzyme was highly specific for NADPH and oxidized glutathione. The preparation had the average catalytic activity of 150 μmoles of NADPH oxidized per min per mg of protein.