That control by the analysis of the difficulty mechanism that a secondary living body by cystine oxidizes and the food factor

1. 氧化胱氨酸形成的反应条件检查：金属-抗坏血酸或金属-过氧化氢体系（芬顿反应）检查各种蛋白质中氧化胱氨酸的形成。由于胰岛素本身的不稳定性，不能用于该反应体系，而血清蛋白在37℃作用24小时后仍能通过氨基酸分析检测出氧化的胱氨酸。氧化半胱氨酸形成的氧化因子考察：氧化蛋白中由Fenton反应（铜离子+过氧化氢）产生的羟基自由基是形成氧化半胱氨酸最有效的氧化剂。然而，除了某种疾病（威尔逊病）外，人体很难存在高浓度的铜离子。因此，在非金属催化条件下，对氧化因子进行了重新研究。结果表明，脂质过氧化（LOOH）对胱氨酸和其他含硫氨基酸的氧化效率最高。氧化半胱氨酸和氧化蛋氨酸（=蛋氨酸亚砜）的选择性比过氧化氢高5倍。氧化胱氨酸的次生生物氧化损伤：发现含有氧化胱氨酸的氧化修饰蛋白与硫醇（谷胱甘肽等）发生反应，产生活性氧（ROS）。这种结合通过产生超氧化物（O2）使天然蛋白质和DNA断裂。山葵异硫氰酸酯对氧化还原的控制：筛选与谷胱甘肽反应产生ROS的食物因子。选择葱属蔬菜和十字花科蔬菜。其中山葵异硫氰酸酯（6-甲基亚砜基己基异硫氰酸酯）在RL34细胞中产生了大量的ROS。

1. Examination of the reaction condition of the oxidized cystine formation : Metal-ascorbic acid or metal-hydrogen peroxide systems (Fenton reaction) were examined for the formation of oxidized cystine in various proteins. Insulin couldn't used for this reaction system because of its unstable nature, but serum proteins were able to detect the oxidized cystine with the amino acid analysis even though in 37℃ for 24 hours.2. Examination of oxidative factors for the oxidized cystine formation : Hydroxyl radical generated by Fenton reaction (copper ion + hydrogen peroxide) was the most effective oxidizing agent for the formation of oxidized cystine in oxidative protein. However, high concentration of copper ion was hard to exist in human body except a certain illness (Wilson disease). So the oxidative factors were re-examined under the non-metal catalyzed conditions. As a result, lipid peroxide (LOOH) caused the oxidation of cystine and the other sulfur-containing amino acids with the best efficiency. Oxidized cystine and oxidized methionine (= methionine sulfoxide) were formed with LOOH about 5 times higher selectivity than hydrogen peroxide.3. Secondary biological oxidative damages by oxidized cystine : Oxidatively modified protein, which contains oxidized cystine, was found to react with thiols (glutathione, etc.) to generate reactive oxygen species (ROS). And this combination made the fragmentation of native protein and DNA by generating superoxide (O2).4. The redox control by wasabi isothiocyanate : The food factor, which can generate ROS from the reaction with glutathione, was screened. The Allium vegetables and the crusifer vegetables were selected. In particular, wasabi isothiocyanate (6- methylsulfinylhexyl isothiocyanate) made great amount of ROS in RL34 cells.